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What is the function of a viral peplomer? | The coronavirus peplomer protein S is responsible for attachment and fusion during viral entry as well as for the induction of cell to cell fusion.
Since tissue affinities are a function of the viral peplomer-mediated attachment of virus to cells and are often directly related to pathogenicity, | Viral proteins of two strains of infectious bronchitis virus (IBV), which have
different tissue trophism and serology, were separated on the basis of their
isoelectric points (pI). The viruses have four structural proteins; the protein
of greatest serological importance is found at the peplomer tip. The viral
structural proteins separated by isoelectric focusing were identified by
comparison to SDS-PAGE separations. Three protein bands were identical in pI and
one protein band showed a difference in pI between strains. When the renatured
viral proteins were Western blotted and reacted with strain-specific antiserum,
antigen-antibody complexing was seen only at points corresponding to the
strain-specific variant bands. For IBV strain Mass-41, antigen-antibody
complexing occurred at a pI of 6.8, and, for IBV strain Ark-99, at 7.2. No cross
reaction of antisera was observed for either strain. Since tissue affinities are
a function of the viral peplomer-mediated attachment of virus to cells and are
often directly related to pathogenicity, it appears that altered pathogenicity
of strains of IBV may be detected by alteration of pI of the proteins.
Classification by pI of proteins of at least the smaller viruses allows
untypeable, highly pathogenic or persistent strains of these viruses to be
characterized on the basis of variant proteins. Numerous studies have demonstrated that the spike glycoprotein of coronaviruses
bears major determits of pathogenesis. To elucidate the antigenic structure
of the protein, a panel of monoclonal antibodies was studied by competitive
ELISA, and their reactivities were assayed against fragments of the murine
coronavirus murine hepatitis virus strain A59 S gene expressed in prokaryotic
vectors. An immunodomit linear domain was localized within the predicted
stalk, S2, of the peplomer. It is recognized by several neutralizing antibodies.
Other domains were also identified near the proteolytic cleavage site, in the
predicted globular head, S1, and in another part of the stalk. Furthermore,
competition results suggest that the immunodomit functional domain forms part
of a complex three-dimensional structure. Surprisingly, some antibodies which
have no antiviral biological activities were shown to bind the immunodomit
neutralization domain. The coronavirus peplomer protein S is responsible for attachment and fusion
during viral entry as well as for the induction of cell to cell fusion. While
several regions within S have been shown to influence the ability to induce
fusion, the region of the protein actually responsible for fusion, the fusion
peptide, has not yet been identified. We identified two hydrophobic peptides
(peptides #1 and #2) within MHV-A59 S2 as possible fusion domains. This was
based on hydrophobicity, conservation among coronavirus S proteins and the
prediction of a sided helix conformation. Using site directed mutagenesis and an
in vitro cell to cell fusion assay we showed that substitution of hydrophobic
amino acids with charged amino acids, within the predicted hydrophobic face of
either of these two peptides eliminated fusion. Within peptide #1 substitution
of the same hydrophobic amino acids with other hydrophobic amino acids or
substitution of polar amino acids with charged or polar amino acids had little
effect on fusion. Thus peptides #1 and #2 remain likely candidates for the MHV
fusion peptide. A third previously identified peptide within S2 (Chambers et
al., 1990) is unlikely as a fusion peptide as it is not well conserved among
coronaviruses and substitution within the hydrophobic face with charged amino
acids does not effect fusion. |
Have toll-like receptor 2 activators been found in food? | Yes, toll-like receptor 2 activators (TLR2) have been found in food. | BACKGROUND: Toll-like receptor 2 (TLR2) is a widely expressed pattern
recognition receptor critical for innate immunity. TLR2 is also a key regulator
of mucosal immunity implicated in the development of allergic disease. TLR2
activators are found in many common foods, but the role of TLR2 in oral
tolerance and allergic sensitization to foods is not well understood.
OBJECTIVE: The purpose of this study was to evaluate the impacts of TLR2
expression and TLR2 activation on oral tolerance to food antigens in a murine
model.
METHODS: Mice were fed ovalbumin (OVA) or peanut butter with or without the
addition of low doses of TLR2 activators Pam3 CSK4 or FSL-1. Oral tolerance was
assessed by analysing antibody responses after a systemic antigen challenge.
OVA-specific Tregs were assessed in the Peyer's patches, mesenteric lymph nodes,
and spleen in wild-type and TLR2(-/-) mice. Low-dose Pam3 CSK4 was also tested
as an oral adjuvant.
RESULTS: Oral tolerance was successfully induced in both wild-type and TLR2(-/-)
recipient mice, with an associated regulatory T-cell response. Oral TLR2
activation, with low-dose Pam3 CSK4 or FSL-1, during oral antigen exposure was
found to alter oral tolerance and was associated with the development of
substantial IgE and IgA responses to foods upon systemic challenge. Low-dose
oral Pam3 CSK4 treatment also selectively enhanced antigen-specific IgA
responses to oral antigen exposure.
CONCLUSIONS AND CLINICAL RELEVANCE: TLR2 is not necessary for oral tolerance
induction, but oral TLR2 activation modulates humoral IgE and IgA responses
during tolerance development. Low-dose Pam3 CSK4 is also an effective oral
adjuvant that selectively enhances IgA production. These observations are
pertinent to the optimization of oral allergen immunotherapy and oral vaccine
development. |
Describe the Open Targets platform | The Open Targets platform is a data integration and visualization platform that provides evidence about the association of known and potential drug targets with diseases. The platform is designed to support identification and prioritization of biological targets for follow-up. Each drug target is linked to a disease using integrated genome-wide data from a broad range of data sources. The platform provides either a target-centric workflow to identify diseases that may be associated with a specific target, or a disease-centric workflow to identify targets that may be associated with a specific disease. Users can easily transition between these target- and disease-centric workflows. The Open Targets Validation Platform is accessible at https://www.targetvalidation.org. | BACKGROUND: We present the Europe PMC literature component of Open Targets - a
target validation platform that integrates various evidence to aid drug target
identification and validation. The component identifies target-disease
associations in documents and ranks the documents based on their confidence from
the Europe PMC literature database, by using rules utilising expert-provided
heuristic information. The confidence score of a given document represents how
valuable the document is in the scope of target validation for a given
target-disease association by taking into account the credibility of the
association based on the properties of the text. The component serves the
platform regularly with the up-to-date data since December, 2015.
RESULTS: Currently, there are a total number of 1168365 distinct target-disease
associations text mined from >26 million PubMed abstracts and >1.2 million Open
Access full text articles. Our comparative analyses on the current available
evidence data in the platform revealed that 850179 of these associations are
exclusively identified by literature mining.
CONCLUSIONS: This component helps the platform's users by providing the most
relevant literature hits for a given target and disease. The text mining
evidence along with the other types of evidence can be explored visually through
https://www.targetvalidation.org and all the evidence data is available for
download in json format from https://www.targetvalidation.org/downloads/data . The recently developed Open Targets platform consolidates a wide range of
comprehensive evidence associating known and potential drug targets with human
diseases. We have harnessed the integrated data from this platform for novel
drug repositioning opportunities. Our computational workflow systematically
mines data from various evidence categories and presents potential repositioning
opportunities for drugs that are marketed or being investigated in ongoing human
clinical trials, based on evidence strength on target-disease pairing. We
classified these novel target-disease opportunities in several ways: (i) number
of independent counts of evidence; (ii) broad therapy area of origin; and (iii)
repositioning within or across therapy areas. Finally, we elaborate on one
example that was identified by this approach. BACKGROUND: The Open Targets Platform integrates different data sources in order
to facilitate identification of potential therapeutic drug targets to treat
human diseases. It currently provides evidence for nearly 2.6 million potential
target-disease pairs. G-protein coupled receptors are a drug target class of
high interest because of the number of successful drugs being developed against
them over many years. Here we describe a systematic approach utilizing the Open
Targets Platform data to uncover and prioritize potential new disease
indications for the G-protein coupled receptors and their ligands.
RESULTS: Utilizing the data available in the Open Targets platform, potential
G-protein coupled receptor and endogenous ligand disease association pairs were
systematically identified. Intriguing examples such as GPR35 for inflammatory
bowel disease and CXCR4 for viral infection are used as illustrations of how a
systematic approach can aid in the prioritization of interesting drug discovery
hypotheses. Combining evidences for G-protein coupled receptors and their
corresponding endogenous peptidergic ligands increases confidence and provides
supportive evidence for potential new target-disease hypotheses. Comparing such
hypotheses to the global pharma drug discovery pipeline to validate the approach
showed that more than 93% of G-protein coupled receptor-disease pairs with a
high overall Open Targets score involved receptors with an existing drug
discovery program.
CONCLUSIONS: The Open Targets gene-disease score can be used to prioritize
potential G-protein coupled receptors-indication hypotheses. In addition,
availability of multiple different evidence types markedly increases confidence
as does combining evidence from known receptor-ligand pairs. Comparing the
top-ranked hypotheses to the current global pharma pipeline serves validation of
our approach and identifies and prioritizes new therapeutic opportunities. The Open Targets Platform integrates evidence from genetics, genomics,
transcriptomics, drugs, animal models and scientific literature to score and
rank target-disease associations for drug target identification. The
associations are displayed in an intuitive user interface
(https://www.targetvalidation.org), and are available through a REST-API
(https://api.opentargets.io/v3/platform/docs/swagger-ui) and a bulk download
(https://www.targetvalidation.org/downloads/data). In addition to target-disease
associations, we also aggregate and display data at the target and disease
levels to aid target prioritisation. Since our first publication two years ago,
we have made eight releases, added new data sources for target-disease
associations, started including causal genetic variants from non genome-wide
targeted arrays, added new target and disease annotations, launched new
visualisations and improved existing ones and released a new web tool for batch
search of up to 200 targets. We have a new URL for the Open Targets Platform
REST-API, new REST endpoints and also removed the need for authorisation for API
fair use. Here, we present the latest developments of the Open Targets Platform,
expanding the evidence and target-disease associations with new and improved
data sources, refining data quality, enhancing website usability, and increasing
our user base with our training workshops, user support, social media and
bioinformatics forum engagement. |
Does radiotherapy for Hodgkin disease increases risk for lung cancer? | Yes, radiotherapy for Hodgkin disease is associated with increased risk for lung cancer. | The risk of second cancers (SCs) was assessed in 744 patients with Hodgkin's
disease (HD) admitted to The Netherlands Cancer Institute from 1966 to 1983.
Sixty-nine SCs were observed one month or more after start of first treatment.
These included 14 cases of lung cancer, nine cases of non-Hodgkin's lymphoma
(NHL), 16 cases of leukemia, and six cases of the myelodysplastic syndrome
(MDS). The median interval between the diagnosis of HD and that of second lung
cancer, NHL, and leukemia was 8.1, 13.3, and 5.7 years, respectively. The
overall relative risks (RR) (observed/expected [O/E] ratios) of developing lung
cancer, NHL, and leukemia were 4.9 (95% confidence limit [CL], 2.7 to 8.2), 31.0
(95% CL, 14.2 to 58.9) and 45.7 (95% CL, 26.1 to 74.2), respectively. At 15
years the cumulative risk of developing an SC amounted to 20.6% +/- 2.9%. The
15-year estimates of lung cancer, NHL, and leukemia were 6.2% +/- 1.9%, 5.9% +/-
2.1% and 6.3% +/- 1.7%, respectively. Increased lung cancer risk following HD
has not frequently been clearly demonstrated before; that we were able to
demonstrate such risk may be due to the completeness of follow-up over long
periods that could be achieved in this study. Excess lung cancer risk was only
noted in treatment regimens with radiotherapy (RT); also, all lung cancers arose
in irradiation fields. Excess risk of leukemia was only found in treatment
regimens involving chemotherapy (CT). For NHL, combined modality treatment was
shown to be the most important risk factor. Risk of lung cancer and NHL
increased with time since diagnosis. A time-dependent covariate analysis (Cox
model) performed on leukemia and MDS showed an increasing risk with intensity of
CT, age (greater than 40 years), and a splenectomy. The risk of lung and breast cancer is significantly increased after therapy for
Hodgkin's disease (HD), but there are few data that describe the molecular
profiles of these tumors. We investigated the genetic abnormalities in second
primary lung (n = 19) and breast cancers (n = 19) that follow therapy for HD
("post-HD cancers") and compared these with changes observed in corresponding
tumor types (57 lung and 20 breast cancers) arising in the general population
("sporadic cancers"). DNA obtained from archival tissues was examined using
PCR-based analyses for loss of heterozygosity and microsatellite alterations
(MAs) at several chromosomal regions, TP53 and K-ras gene mutations, and
frameshift mutations at minisatellite sequences at the coding regions of several
genes (TGF-betaRII, IGFIIR, BAX, hMSH6, and hMSH3). The occurrence of loss of
heterozygosity at all chromosomal regions taken together and frequencies at most
individual areas were similar for the post-HD and sporadic cancers for both lung
and breast sites. The overall frequency of MAs in the post-HD tumors was
substantially greater (lung, 2.4-fold, P = 0.004; breast, 4.2-fold, P = 0.16)
than that in the respective sporadic cancers. No differences in the pattern of
TP53 and K-ras mutations were detected between post-HD and sporadic cancers. No
mutations were detected at the minisatellite sequences examined. MAs, which
reflect widespread genomic instability, occur at greatly increased frequency in
post-HD lung and breast cancers. Although the mechanisms underlying the
development of increased MAs are unknown, they have been associated with
immunosuppression and radiation exposure. Future research should address the
role that MAs, as well as other influences, may play in the development of
neoplasias that occur after therapy for HD. BACKGROUND: Lung cancer is a frequent cause of death in patients cured of
Hodgkin's disease, but the contributions of chemotherapy, radiotherapy, and
smoking are not well described. We quantified the risk of treatment-associated
lung cancer, taking into account tobacco use.
METHODS: Within a population-based cohort of 19 046 Hodgkin's disease patients
(diagnosed from 1965 through 1994), a case-control study of lung cancer was
conducted. The cumulative amount of cytotoxic drugs, the radiation dose to the
specific location in the lung where cancer developed, and tobacco use were
compared for 222 patients who developed lung cancer and for 444 matched control
patients. All statistical tests were two-sided.
RESULTS: Treatment with alkylating agents without radiotherapy was associated
with increased lung cancer risk (relative risk [RR] = 4.2; 95% confidence
interval [CI] = 2.1 to 8.8), as was radiation dose of 5 Gy or more without
alkylating agents (RR = 5.9; 95% CI = 2.7 to 13.5). Risk increased with both
increasing number of cycles of alkylating agents and increasing radiation dose
(P for trend <.001). Among patients treated with mechlorethamine, vincristine,
procarbazine, and prednisone (MOPP), risk increased with cumulative amounts of
mechlorethamine and procarbazine (P<.001) when evaluated separately.
Statistically significantly elevated risks of lung cancer were apparent within
1-4 years after treatment with alkylating agents, whereas excess risk after
radiotherapy began 5 years after treatment and persisted for more than 20 years.
Risk after treatment with alkylating agents and radiotherapy together was as
expected if individual excess risks were summed. Tobacco use increased lung
cancer risk more than 20-fold; risks from smoking appeared to multiply risks
from treatment.
CONCLUSIONS: Past treatments with alkylating agents and radiation therapy for
Hodgkin's disease were associated with an increased risk of lung cancer in a
dose-dependent and additive fashion. The precise risk estimates, however, should
be interpreted cautiously, given the possible residual and enhancing effects of
tobacco. PURPOSE: Hodgkin lymphoma (HL) survivors face an increased risk of
treatment-related lung cancer. Screening with low-dose computed tomography
(LDCT) may allow detection of early stage, resectable cancers. We developed a
Markov decision-analytic and cost-effectiveness model to estimate the merits of
annual LDCT screening among HL survivors.
METHODS AND MATERIALS: Population databases and HL-specific literature informed
key model parameters, including lung cancer rates and stage distribution,
cause-specific survival estimates, and utilities. Relative risks accounted for
radiation therapy (RT) technique, smoking status (>10 pack-years or current
smokers vs not), age at HL diagnosis, time from HL treatment, and excess
radiation from LDCTs. LDCT assumptions, including expected stage-shift,
false-positive rates, and likely additional workup were derived from the
National Lung Screening Trial and preliminary results from an internal phase 2
protocol that performed annual LDCTs in 53 HL survivors. We assumed a 3%
discount rate and a willingness-to-pay (WTP) threshold of $50,000 per
quality-adjusted life year (QALY).
RESULTS: Annual LDCT screening was cost effective for all smokers. A male smoker
treated with mantle RT at age 25 achieved maximum QALYs by initiating screening
12 years post-HL, with a life expectancy benefit of 2.1 months and an
incremental cost of $34,841/QALY. Among nonsmokers, annual screening produced a
QALY benefit in some cases, but the incremental cost was not below the WTP
threshold for any patient subsets. As age at HL diagnosis increased, earlier
initiation of screening improved outcomes. Sensitivity analyses revealed that
the model was most sensitive to the lung cancer incidence and mortality rates
and expected stage-shift from screening.
CONCLUSIONS: HL survivors are an important high-risk population that may benefit
from screening, especially those treated in the past with large radiation fields
including mantle or involved-field RT. Screening may be cost effective for all
smokers but possibly not for nonsmokers despite a small life expectancy benefit. Lung cancer (LC) represents the most common solid tumor in survivors of
Hodgkin's disease (HD), and the assessment of the mutational status of oncogenic
driver mutations in LC is now standard. We compiled clinical and mutation data
(EGFR, KRAS, and ALK) from the medical records of patients with LC and a remote
history of HD. 13 cases of LC following HD were seen, including seven with
mutational data. Two had EGFR mutations, none had KRAS mutations or ALK
translocations. Our conclusions are limited by the small sample size, however
this report reinforces the need to identify driver mutations in lung cancers. PURPOSE: Hodgkin lymphoma (HL) survivors have an increased risk of
cardiovascular disease (CD), lung cancer, and breast cancer. We investigated the
risk for the development of CD and secondary lung, breast, and thyroid cancer
after radiation therapy (RT) delivered with deep inspiration breath-hold (DIBH)
compared with free-breathing (FB) using 3-dimensional conformal RT (3DCRT) and
intensity modulated RT (IMRT). The aim of this study was to determine which
treatment modality best reduced the combined risk of life-threatening late
effects in patients with mediastinal HL.
METHODS AND MATERIALS: Twenty-two patients with early-stage mediastinal HL were
eligible for the study. Treatment plans were calculated with both 3DCRT and
IMRT on both DIBH and FB planning computed tomographic scans. We reported the
estimated dose to the heart, lung, female breasts, and thyroid and calculated
the estimated life years lost attributable to CD and to lung, breast, and
thyroid cancer.
RESULTS: DIBH lowered the estimated dose to heart and lung regardless of
delivery technique (P<.001). There was no significant difference between IMRT-FB
and 3DCRT-DIBH in mean heart dose, heart V20Gy, and lung V20Gy. The mean breast
dose was increased with IMRT regardless of breathing technique. Life years lost
was lowest with DIBH and highest with FB.
CONCLUSIONS: In this cohort, 3DCRT-DIBH resulted in lower estimated doses and
lower lifetime excess risks than did IMRT-FB. Combining IMRT and DIBH could be
beneficial for a subgroup of patients. PURPOSE: To evaluate the risk factors associated with lung cancer (LC) and other
second neoplasms (SN) in Hodgkin lymphoma (HL) survivors.
METHODS: We retrospectively analyzed the clinical characteristics and outcomes
of 604 patients treated in our institution between 1968 and 2012.
RESULTS: 90 out of 604 patients developed SN: 27 LC and 63 other SN. The median
time elapsed until LC and other SN was 16.5 and 11.8 years, respectively (p =
0.003). In the LC group, 85.5 % of patients were male and 84.6 % smokers (HR 7,
95 % CI 2.4-20.7, p < 0.001). Radiotherapy (RT) doses applied were higher in the
SN group with an increased risk of LC (HR: 4.0 95 % CI 1.1-11.6, p = 0.010) and
other SN (HR: 3.3 95 % CI 1.6-6.7 p = 0.001) with doses higher than 42 Gy. No
association was found between alkylating agents and development of SN. In LC,
the most frequent histology was adenocarcinoma with an elapsed time after HL of
13.2 years in early stages and 21.3 in advanced (p = 0.02). Median OS after a
diagnosis of LC was 12.6 months ranging from 5.9 (in cases presenting due to
symptoms) to 49.1 (incidentally diagnosed cases) (p = 0.005).
CONCLUSIONS: RT treatment, especially with doses higher than 42 Gy, and smoking
increase the risk of SN after HL. In this series, LC patients with early stages
had a shorter elapsed time from HL diagnosis and longer OS, therefore the role
of LC screening in HL survivors should be prospectively evaluated and smoking
cessation counseling ought to be a key aspect during follow-up. |
Does radiotherapy for prostate cancer increase bladder cancer risk? | Yes, radiotherapy for prostate cancer is associated with increased bladder cancer risk. | PURPOSE: Pre-prostate specific antigen era series demonstrated an increased risk
of bladder cancer and rectal cancer in men who received radiotherapy for
prostate cancer. We estimated the risk of secondary bladder cancer and rectal
cancer after prostate radiotherapy using a contemporary population based cohort.
MATERIALS AND METHODS: We identified 243,082 men in the Surveillance,
Epidemiology and End Results database who underwent radical prostatectomy or
radiotherapy for prostate cancer between 1988 and 2003. We estimated the
incidence rate, standardized incidence ratio and age adjusted incidence rate
ratio of subsequent bladder cancer and rectal cancer associated with radical
prostatectomy, external beam radiotherapy, brachytherapy, and a combination of
external beam radiotherapy and brachytherapy.
RESULTS: The relative risk of bladder cancer developing after external beam
radiotherapy, brachytherapy and external beam radiotherapy-brachytherapy
compared to radical prostatectomy was 1.88, 1.52 and 1.85, respectively.
Compared to the general United States population the standardized incidence
ratio for bladder cancer developing after radical prostatectomy, external beam
radiotherapy, brachytherapy and external beam radiotherapy-brachytherapy was
0.99, 1.42, 1.10 and 1.39, respectively. The relative risk of rectal cancer
developing after external beam radiotherapy, brachytherapy and external beam
radiotherapy-brachytherapy compared to radical prostatectomy was 1.26, 1.08 and
1.21, respectively. The standardized incidence ratio for rectal cancer
developing after radical prostatectomy, external beam radiotherapy,
brachytherapy and external beam radiotherapy-brachytherapy was 0.91, 0.99, 0.68
and 0.86, respectively.
CONCLUSIONS: Men who receive radiotherapy for localized prostate cancer have an
increased risk of bladder cancer compared to patients undergoing radical
prostatectomy and compared to the general population. The risk of rectal cancer
is increased in patients who receive external beam radiotherapy compared to
radical prostatectomy. Patients should be counseled appropriately regarding
these risks. PURPOSE OF REVIEW: Prostate cancer is the most common cancer diagnosed in men
and remains the second most lethal maligcy. Most patients undergoing
treatment elect for radical prostatectomy or radiation. As the number of
patients treated has increased and survival improved, delayed complications of
these modalities has assumed increased importance. Recent studies report an
increased risk of certain cancers after radiation for prostate cancer. This
review aims to summarize recent data.
RECENT FINDINGS: Recent studies have confirmed the association of prostate
radiation with secondary cancers. The most common secondary maligcy is
bladder carcinoma. We have treated 44 patients with bladder cancer who had
radiation therapy for prostate cancer. At diagnosis, 60% had tumor, which
invaded the bladder muscle (T2 or greater disease). The mean latency from
radiation to diagnosis of bladder cancer was 5.5 years.
SUMMARY: Radiation therapy for prostate cancer is associated with an increased
risk of bladder cancer. In our series, patients presented at higher stage than
expected from population-based studies of bladder cancer. Patients and their
physicians should be aware of such risks when choosing therapy for prostate
cancer. Hematuria following radiation therapy for prostate cancer should be
investigated rather than being attributed to radiation-induced cystitis. Radiotherapy for prostate cancer is associated with an increased incidence of
secondary bladder cancer (BC). We investigated the incidence,
clinicopathological characteristics, and prognosis of BC after radiotherapy,
surgical therapy, and primary androgen-deprivation therapy (ADT) for prostate
cancer. This study included 1,334 Japanese patients with prostate cancer treated
with radiotherapy (n=631), surgical therapy (n=437), and primary ADT (n=266).
During the median follow-up period of 51.2, 44.8, and 45.5 months, secondary BC
occurred in 14 (2.2%), 5 (1.1%), and 0 (0%) of patients with prostate cancer
treated with radiotherapy, surgical therapy, and primary ADT, respectively. The
10-year BC-free survival rate was 91.3% in the radiotherapy group, 97.4% in the
surgical therapy group, and 100% in the primary ADT group. The rates of
intravesical recurrence, progression to muscle-invasive BC, and BC-specific
death might be higher in secondary BC after radiotherapy compared with after
surgical therapy. There was a significant difference in the incidence of
secondary BC among different therapeutic modalities for prostate cancer in
Japanese men, indicating significantly lower comorbidity rates of secondary BC
after primary ADT for prostate cancer compared with radiotherapy. OBJECTIVE: To determine the association between exposure to radiotherapy for the
treatment of prostate cancer and subsequent second maligcies (second primary
cancers).
DESIGN: Systematic review and meta-analysis of observational studies.
DATA SOURCES: Medline and Embase up to 6 April 2015 with no restrictions on year
or language.
STUDY SELECTION: Comparative studies assessing the risk of second maligcies
in patients exposed or unexposed to radiotherapy in the course of treatment for
prostate cancer were selected by two reviewers independently with any
disagreement resolved by consensus.
DATA EXTRACTION AND SYNTHESIS: Two reviewers independently extracted study
characteristics and outcomes. Risk of bias was assessed with the
Newcastle-Ottawa scale. Outcomes were synthesized with random effects models and
Mantel-Haenszel weighting. Unadjusted odds ratios and multivariable adjusted
hazard ratios, when available, were pooled.
MAIN OUTCOME MEASURES: Second cancers of the bladder, colorectal tract, rectum,
lung, and hematologic system.
RESULTS: Of 3056 references retrieved, 21 studies were selected for analysis.
Most included studies were large multi-institutional reports but had moderate
risk of bias. The most common type of radiotherapy was external beam; 13 studies
used patients treated with surgery as controls and eight used patients who did
not undergo radiotherapy as controls. The length of follow-up among studies
varied. There was increased risk of cancers of the bladder (four studies;
adjusted hazard ratio 1.67, 95% confidence interval 1.55 to 1.80), colorectum
(three studies; 1.79, 1.34 to 2.38), and rectum (three studies; 1.79, 1.34 to
2.38), but not cancers of the hematologic system (one study; 1.64, 0.90 to 2.99)
or lung (two studies; 1.45, 0.70 to 3.01), after radiotherapy compared with the
risk in those unexposed to radiotherapy. The odds of a second cancer varied
depending on type of radiotherapy: treatment with external beam radiotherapy was
consistently associated with increased odds while brachytherapy was not. Among
the patients who underwent radiotherapy, from individual studies, the highest
absolute rates reported for bladder, colorectal, and rectal cancers were 3.8%,
4.2%, and 1.2%, respectively, while the lowest reported rates were 0.1%, 0.3%,
and 0.3%.
CONCLUSION: Radiotherapy for prostate cancer was associated with higher risks of
developing second maligcies of the bladder, colon, and rectum compared with
patients unexposed to radiotherapy, but the reported absolute rates were low.
Further studies with longer follow-up are required to confirm these findings. OBJECTIVE: Although it is well known that radiotherapy for prostate cancer
increases comorbid rate of secondary bladder cancer, the effect of aging and
smoking with radiotherapy on incidence rate of secondary bladder cancer remains
unknown. Then, this study investigated the combinational effect of external beam
radiotherapy for prostate cancer and aging or smoking on comorbid rate of
secondary bladder cancer.
METHODS: This study included 754 Japanese patients with prostate cancer treated
with radiotherapy (n = 319) and radical prostatectomy (n = 435) from 2000
through 2013. The relationship between therapeutic modality for prostate cancer
as well as age or smoking status and comorbid rate of secondary bladder cancer
was examined.
RESULTS: During the median follow-up period of 4.3 and 3.1 years, secondary
bladder cancer occurred in 11 (3.4%) and 5 (1.1%) of patients with prostate
cancer treated with external beam radiotherapy and radical prostatectomy,
respectively. The 5-year bladder cancer-free survival rate was 97.3% in the
external beam radiotherapy group and 99.4% in the radical prostatectomy group.
Age (hazard ratio = 1.15, P = 0.027) and ever smoking (hazard ratio = 5.65, P
= 0.011) were significant predictive factors of secondary bladder cancer
incidence in the external beam radiotherapy cohort, but not in the radical
prostatectomy cohort. Inversely, among men with ever smoking, but not among
older men, external beam radiotherapy (hazard ratio = 9.64, P = 0.0052) was a
significant risk factor of secondary bladder cancer.
CONCLUSIONS: Taken together, these findings suggest that smoking history might
be one of criteria to choose radical prostatectomy than external beam
radiotherapy for prostate cancer, and that age would not be a criterion for
therapeutic selection in terms of secondary bladder cancer. Radiation therapy represents an alternative treatment to radical prostatectomy
in the management of clinically localized prostate cancer. Radiation-induced
second neoplasms are defined by a latency period of at least 5 years, location
within the field of radiation therapy, and a histology which differs from the
primary tumor. Based on the data in the literature, there is a consistently
increased risk of bladder cancer (HR: 1.67, 95% CI 1.55-1.80), rectal cancer
(HR: 1.79, 95% CI 1.34-2.38), and colorectal cancer (HR: 1.79, 95% CI 1.34-23.8)
following percutaneous radiation therapy. Following brachytherapy only an
increased for the development of bladder cancer (HR: 2.14, 95% CI 1.03-3.94) has
been observed. The incidence of second neoplasms increases significantly and
continuously with the posttreatment time interval. Although bladder cancers
following RT of the prostate are usually more locally advanced and of high
grade, no negative impact in terms of overall survival and cancer-specific
survival has been observed. Symptoms or findings of microhematuria need to be
examined thoroughly after radiation therapy to identify bladder cancer quite
early. PURPOSE: To analyse the rate of secondary maligcies observed in a series of
675 prostate cancer patients who underwent a permanent implant brachytherapy
between 1999 and 2003, and to compare the incidence with the expected rate in a
matched general French population.
MATERIAL AND METHODS: The cohort included low-risk patients and a selection of
"favourable-intermediate" risk patients. All patients were homogeneously treated
using an intraoperative dynamic planning prostate brachytherapy technique, with
loose 125-iodine seeds and a prescription dose of 145Gy. The mean follow-up was
132 months.
RESULTS: The 10-year overall survival for the entire cohort was 92% (95%
confidence interval [CI]: 90-94). The 10-year relapse-free survival rate was 82%
(95% CI: 79-85). Overall, 61 second cancers were registered. When comparing with
a matched general French population, the standard incidence ratio (SIR) for
bladder cancer was 1.02 (95% CI: 0.46-1.93). For colorectal cancer, the SIR was
0.45 (95% CI: 0.19-0.89). For lung cancer, the SIR was 0.38 (95% CI: 0.17-0.76).
The SIR for all cancers was 0.61 (95% CI: 0.47-0.79). When excluding secondary
colorectal and lung cancers (both with low SIRs in this series), the SIR for all
cancers was 1.06 (95% CI: 0.77-1.29).
CONCLUSION: With a mean follow-up of more than 11 years, this series does not
detect any excess risk of second cancers associated with permanent implant
prostate brachytherapy. However, due to power limitation, a small increase in
the risk of secondary maligcies cannot be totally ruled out. BACKGROUND: Long-term survival can be achieved in patients affected by localized
prostate cancer (PCa) treated with either radical prostatectomy (RP) or external
beam radiotherapy (EBRT). However, development of a second primary tumor is
still poorly investigated.
OBJECTIVE: To investigate the impact of RP and EBRT on subsequent risk of
developing bladder (BCa) and/or rectal cancer (RCa) among PCa survivors.
DESIGN, SETTING, AND PARTICIPANTS: A total of 84397 patients diagnosed with
localized PCa, treated with RP or EBRT between 1988 and 2009, and older than 65
yr of age were identified in the Surveillance, Epidemiology, and End Results
Medicare insurance program-linked database. Our primary objective was to
investigate the effect of EBRT and RP on the second primary BCa and RCa
incidence.
OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Multivariable competing-risk
regression analyses were performed to assess the risk of developing a second
primary BCa or RCa.
RESULTS AND LIMITATIONS: Of the 84397 individuals included in the study, 33252
(39%) were treated with RP and 51145 (61%) with EBRT. Median follow-up was 69
months, and follow-up periods for patients who did not develop BCa, RCa, or
pelvic cancer were 68, 69, and 68 mo, respectively. A total of 1660 individuals
developed pelvic tumors (1236 BCa and 432 RCa). The 5- and 10-yr cumulative BCa
incidence rates were 0.75% (95% confidence interval [CI]: 0.64-0.85%) and 1.63%
(95% CI: 1.45-1.80%) versus 1.26% (95% CI: 1.15-1.37%) and 2.34% (95% CI:
2.16-2.53%) for patients treated with RP versus EBRT, respectively. The 5- and
10-yr cumulative RCa incidence rates were 0.32% (95% CI: 0.25-0.39%) and 0.73%
(95% CI: 0.61-0.85%) versus 0.36% (95% CI: 0.30-0.41%) and 0.69% (95% CI:
0.60-0.79%) for patients treated with RP versus EBRT, respectively. On
multivariable competing risk regression analyses, treatment with EBRT was
independently associated with the risk of developing a second primary BCa
(hazard ratio: 1.35, CI: 1.18-1.55; p<0.001), but not RCa (p=0.4). Limitations
include lack of information regarding the dose of radiotherapy and the
retrospective nature with the implicit risk of selection bias.
CONCLUSIONS: Patients treated with EBRT are at increased risk of developing a
second primary BCa compared with those treated with RP. However, no differences
were found considering RCa incidence in patients treated with RP or EBRT within
the first 5 yr after primary therapy. These results need to be validated in a
well-designed randomized prospective trial.
PATIENT SUMMARY: We retrospectively analyzed the risk of developing a second
primary bladder or rectal cancer during follow-up for patients treated with
radical prostatectomy or external beam radiotherapy for a localized prostate
cancer. We found that those treated with external beam radiotherapy are at an
increased risk of developing a second primary bladder cancer tumor. |
What is ESN364? | systemic administration of an NK3R antagonist (ESN364) prolongs the LH interpulse interval in ovarectomized ewes and significantly lowers plasma LH and FSH concentrations in castrated nonhuman primates (Macaca fascicularis). Moreover, daily oral dosing of ESN364 throughout the menstrual cycle in M fascicularis lowered plasma estradiol levels in a dose-dependent manner, although nadir levels of estradiol were maintained well above menopausal levels. Nevertheless, estradiol levels during the follicular phase were sufficiently inhibited at all doses to preclude the triggering of ovulation as evidenced by the absence of the LH surge and failure of a subsequent luteal phase rise in plasma progesterone concentrations, consistent with the absence of normal cycle changes in the uterus. Apart from the point at surge, FSH levels were not altered over the course of the menstrual cycle. These effects of ESN364 were reversible upon cessation of drug treatment.
ESN364 was well-tolerated and rapidly bioavailable with linear pharmacokinetics and no drug accumulation with repeated, daily oral administration. Drug treatment dose-dependently decreased basal LH, but not FSH, and consequently decreased estradiol and progesterone (in women) as well as testosterone (in men). The hormonal changes in women corresponded to delayed ovulation, decreased endometrial thickening, impeded follicular maturation, and prolongation of the menstrual cycle. Drug effects were rapidly reversible. Oral administration of the NK3R antagonist, ESN364, suppressed the hypothalamic-pituitary-gonadal axis in healthy volunteers by selective modulation of gonadotropin secretion, leading to a restrained decrease in ovarian hormone levels in women. These results suggest that ESN364 may offer therapeutic benefit in the treatment of women's health disorders with a mitigated risk of menopausal-like adverse events. | Women's health disorders such as uterine fibroids and endometriosis are
currently treated by GnRH modulators that effectively suppress the
hypothalamic-pituitary-gonadal axis. The neurokinin-3 receptor (NK3R) is an
alternative target with an important role in the modulation of this axis. In
this report, we demonstrate that systemic administration of an NK3R antagonist
(ESN364) prolongs the LH interpulse interval in ovarectomized ewes and
significantly lowers plasma LH and FSH concentrations in castrated nonhuman
primates (Macaca fascicularis). Moreover, daily oral dosing of ESN364 throughout
the menstrual cycle in M fascicularis lowered plasma estradiol levels in a
dose-dependent manner, although nadir levels of estradiol were maintained well
above menopausal levels. Nevertheless, estradiol levels during the follicular
phase were sufficiently inhibited at all doses to preclude the triggering of
ovulation as evidenced by the absence of the LH surge and failure of a
subsequent luteal phase rise in plasma progesterone concentrations, consistent
with the absence of normal cycle changes in the uterus. Apart from the point at
surge, FSH levels were not altered over the course of the menstrual cycle. These
effects of ESN364 were reversible upon cessation of drug treatment. Together
these data support the proposed role of neurokinin B-NK3R signaling in the
control of pulsatile GnRH secretion. Furthermore, in contrast to GnRH
antagonists, NK3R antagonists induce a partial suppression of estradiol and
thereby offer a viable therapeutic approach to the treatment of ovarian sex
hormone disorders with a mitigated risk of menopausal-like adverse events in
response to long-term drug exposure. CONTEXT: Women's health disorders are commonly treated by agents that suppress
the hypothalamic-pituitary-gonadal axis. NK3 receptor antagonism modulates this
axis with distinct pharmacology compared to existing therapies.
OBJECTIVE: The study aim was to evaluate safety, pharmacokinetics, and
pharmacodynamics on gonadotropins and sex hormones after single- and
multiple-dose administration of an NK3R antagonist to healthy men and women.
DESIGN AND SETTING: This was a first-in-human, double-blind, placebo-controlled,
combined single and multiple ascending dose trial.
PARTICIPANTS: Forty-one men and 24 regularly cycling women participated in the
study.
INTERVENTION(S): In part 1 of the study, men received single oral doses of 3-180
mg or placebo. In part 2, men received placebo or 20, 60, or 180 mg each day for
10 days. In part 3, women received placebo or 20, 60, or 180 mg each day for 21
days, where dosing was initiated on day 3 ± 2 after menses.
MAIN OUTCOME MEASURE(S): Safety, tolerability, pharmacokinetics, and
pharmacodynamics on circulating levels of LH, FSH, testosterone, estradiol, and
progesterone, in addition to physiological biomarkers of endometrial thickening,
follicle growth, and the duration of the menstrual cycle were evaluated.
RESULTS: ESN364 was well-tolerated and rapidly bioavailable with linear
pharmacokinetics and no drug accumulation with repeated, daily oral
administration. Drug treatment dose-dependently decreased basal LH, but not FSH,
and consequently decreased estradiol and progesterone (in women) as well as
testosterone (in men). The hormonal changes in women corresponded to delayed
ovulation, decreased endometrial thickening, impeded follicular maturation, and
prolongation of the menstrual cycle. Drug effects were rapidly reversible.
CONCLUSIONS: Oral administration of the NK3R antagonist, ESN364, suppressed the
hypothalamic-pituitary-gonadal axis in healthy volunteers by selective
modulation of gonadotropin secretion, leading to a restrained decrease in
ovarian hormone levels in women. These results suggest that ESN364 may offer
therapeutic benefit in the treatment of women's health disorders with a
mitigated risk of menopausal-like adverse events. |
Do genes with monoallelic expression contribute proportionally to genetic diversity in humans? | No, genes with monoallelic expression contribute disproportionately to genetic diversity in humans. | An unexpectedly large number of human autosomal genes are subject to monoallelic
expression (MAE). Our analysis of 4,227 such genes uncovers surprisingly high
genetic variation across human populations. This increased diversity is unlikely
to reflect relaxed purifying selection. Remarkably, MAE genes exhibit an
elevated recombination rate and an increased density of hypermutable sequence
contexts. However, these factors do not fully account for the increased
diversity. We find that the elevated nucleotide diversity of MAE genes is also
associated with greater allelic age: variants in these genes tend to be older
and are enriched in polymorphisms shared by Neanderthals and chimpanzees. Both
synonymous and nonsynonymous alleles of MAE genes have elevated average
population frequencies. We also observed strong enrichment of the MAE signature
among genes reported to evolve under balancing selection. We propose that an
important biological function of widespread MAE might be the generation of
cell-to-cell heterogeneity; the increased genetic variation contributes to this
heterogeneity. |
Has MLE4901 been tested in phase III clinical trials? | No, MLE4901 has been tested in phase 2, randomised, double-blind, placebo-controlled trial. | BACKGROUND: Hot flushes affect 70% of menopausal women and often severely impact
physical, psychosocial, sexual, and overall wellbeing. Hormone replacement
therapy is effective but is not without risk. Neurokinin B signalling is
increased in menopausal women, and has been implicated as an important mediator
of hot flushes.
METHODS: This phase 2, randomised, double-blind, placebo-controlled,
single-centre, crossover trial assessed the effectiveness of an oral neurokinin
3 receptor antagonist (MLE4901) on menopausal hot flushes. Eligible participants
were healthy women aged 40-62 years, having seven or more hot flushes in every
24 h of which some were reported as being severe or bothersome, who had not had
a menstrual period for at least 12 months, and who had not been taking any
medication shown to improve menopausal flushes in the preceding 8 weeks.
Participants received 4 weeks of MLE4901 (40 mg, orally, twice daily) and
placebo (orally, twice daily) in random order separated by a 2 week washout
period. Randomisation was completed by a central computer, and participants were
allocated to treatment number in numerical order. The primary outcome was the
total number of hot flushes during the final week of both treatment periods.
Analyses were by intention to treat and per protocol using generalised linear
mixed models and standard crossover analysis. All analyses were prespecified in
the study protocol. The trial is registered at ClinicalTrials.gov, number
NCT02668185.
FINDINGS: 68 women were screened between Feb 3 and Oct 10, 2016, of which 37
were randomly assigned and included in an intention-to-treat analysis. 28
participants completed the trial and were included in a per-protocol analysis.
MLE4901 significantly reduced the total weekly number of hot flushes by 45
percentage points (95% CI 22-67) compared with the placebo (intention-to-treat
adjusted means: placebo 49·01 [95% CI 40·81-58·56] vs MLE4901 19·35
[15·99-23·42]; adjusted estimate of difference 29·66 [17·39-42·87], p<0·0001).
Treatment was well tolerated. Three participants developed a transaminase rise
(alanine aminotransferase 4·5-5·9 times the upper limit of normal) with a normal
bilirubin 28 days after starting MLE4901, which normalised within 90 days.
INTERPRETATION: Treatment with a neurokinin 3 receptor antagonist (MLE4901)
could be practice changing as it safely and effectively relieves hot flush
symptoms without the need for oestrogen exposure. Larger scale studies of longer
duration are now indicated.
FUNDING: UK Medical Research Council and National Institute for Health Research. |
Does ESN364 activate the hypothalamic-pituitary-gonadal axis? | No, the NK3R antagonist, ESN364, suppressed the hypothalamic-pituitary-gonadal axis in healthy volunteers by selective modulation of gonadotropin secretion | CONTEXT: Women's health disorders are commonly treated by agents that suppress
the hypothalamic-pituitary-gonadal axis. NK3 receptor antagonism modulates this
axis with distinct pharmacology compared to existing therapies.
OBJECTIVE: The study aim was to evaluate safety, pharmacokinetics, and
pharmacodynamics on gonadotropins and sex hormones after single- and
multiple-dose administration of an NK3R antagonist to healthy men and women.
DESIGN AND SETTING: This was a first-in-human, double-blind, placebo-controlled,
combined single and multiple ascending dose trial.
PARTICIPANTS: Forty-one men and 24 regularly cycling women participated in the
study.
INTERVENTION(S): In part 1 of the study, men received single oral doses of 3-180
mg or placebo. In part 2, men received placebo or 20, 60, or 180 mg each day for
10 days. In part 3, women received placebo or 20, 60, or 180 mg each day for 21
days, where dosing was initiated on day 3 ± 2 after menses.
MAIN OUTCOME MEASURE(S): Safety, tolerability, pharmacokinetics, and
pharmacodynamics on circulating levels of LH, FSH, testosterone, estradiol, and
progesterone, in addition to physiological biomarkers of endometrial thickening,
follicle growth, and the duration of the menstrual cycle were evaluated.
RESULTS: ESN364 was well-tolerated and rapidly bioavailable with linear
pharmacokinetics and no drug accumulation with repeated, daily oral
administration. Drug treatment dose-dependently decreased basal LH, but not FSH,
and consequently decreased estradiol and progesterone (in women) as well as
testosterone (in men). The hormonal changes in women corresponded to delayed
ovulation, decreased endometrial thickening, impeded follicular maturation, and
prolongation of the menstrual cycle. Drug effects were rapidly reversible.
CONCLUSIONS: Oral administration of the NK3R antagonist, ESN364, suppressed the
hypothalamic-pituitary-gonadal axis in healthy volunteers by selective
modulation of gonadotropin secretion, leading to a restrained decrease in
ovarian hormone levels in women. These results suggest that ESN364 may offer
therapeutic benefit in the treatment of women's health disorders with a
mitigated risk of menopausal-like adverse events. |
Which type of variants can be called by the VarDict algorithm? | VarDict is a novel and versatile variant caller for both DNA- and RNA-sequencing data. It simultaneously calls SNV, MNV, InDels, complex and structural variants, expanding the detected genetic driver landscape of tumors. It performs local realignments on the fly for more accurate allele frequency estimation. | Accurate variant calling in next generation sequencing (NGS) is critical to
understand cancer genomes better. Here we present VarDict, a novel and versatile
variant caller for both DNA- and RNA-sequencing data. VarDict simultaneously
calls SNV, MNV, InDels, complex and structural variants, expanding the detected
genetic driver landscape of tumors. It performs local realignments on the fly
for more accurate allele frequency estimation. VarDict performance scales
linearly to sequencing depth, enabling ultra-deep sequencing used to explore
tumor evolution or detect tumor DNA circulating in blood. In addition, VarDict
performs amplicon aware variant calling for polymerase chain reaction
(PCR)-based targeted sequencing often used in diagnostic settings, and is able
to detect PCR artifacts. Finally, VarDict also detects differences in somatic
and loss of heterozygosity variants between paired samples. VarDict reprocessing
of The Cancer Genome Atlas (TCGA) Lung Adenocarcinoma dataset called known
driver mutations in KRAS, EGFR, BRAF, PIK3CA and MET in 16% more patients than
previously published variant calls. We believe VarDict will greatly facilitate
application of NGS in clinical cancer research. |
Has ORMD-0801 been tested in patients? | Yes, ORMD-0801 has been tested in patients. | The unpredictable behavior of uncontrolled type 1 diabetes often involves
frequent swings in blood glucose levels that impact maintece of a daily
routine. An intensified insulin regimen is often unsuccessful, while other
therapeutic options, such as amylin analog injections, use of continuous glucose
sensors, and islet or pancreas transplantation are of limited clinical use. In
efforts to provide patients with a more compliable treatment method, Oramed
Pharmaceuticals tested the capacity of its oral insulin capsule (ORMD-0801, 8 mg
insulin) in addressing this resistant clinical state. Eight Type I diabetes
patients with uncontrolled diabetes (HbA1c: 7.5-10%) were monitored throughout
the 15-day study period by means of a blind continuous glucose monitoring
device. Baseline patient blood glucose behavior was monitored and recorded over
a five-day pretreatment screening period. During the ensuing ten-day treatment
phase, patients were asked to conduct themselves as usual and to self-administer
an oral insulin capsule three times daily, just prior to meal intake. CGM data
sufficient for pharmacodynamics analyses were obtained from 6 of the 8 subjects.
Treatment with ORMD-0801 was associated with a significant 24.4% reduction in
the frequencies of glucose readings >200 mg/dL (60.1 ± 7.9% pretreatment vs.
45.4 ± 4.9% during ORMD-0801 treatment; p = 0.023) and a significant mean 16.6%
decrease in glucose area under the curve (AUC) (66055 ± 5547 mg/dL/24 hours vs.
55060 ± 3068 mg/dL/24 hours, p = 0.023), with a greater decrease during the
early evening hours. In conclusion, ORMD-0801 oral insulin capsules in
conjunction with subcutaneous insulin injections, well tolerated and effectively
reduced glycemia throughout the day.
TRIAL REGISTRATION: Clinicaltrials.gov NCT00867594. |
Was vivotif licensed in Europe and the US at the same time? | No, vivotif was licensed in Europe in 1983 and in the US in 1989. | Cases of diarrhoeal disease number from 1.7 to 5 billion per year worldwide. One
of the main causes of diarrhoeal disease is typhoid fever, which is a
potentially life-threatening multi-systemic illness. According to the most
recent estimates, a total of 26.9 million typhoid fever episodes occurred in
2010. The geographical distribution of the disease differs widely; in developed
countries, the incidence rate per 100,000 per year varies from < 0.1 to 0.3, and
the disease mainly affects people who travel to endemic areas located in low-
and middle-income countries. Low- and middle-income countries are mainly
affected owing to the lack of clean water and proper sanitation. In the fight
against this plague, prevention is fundamental, and vaccination against typhoid
is an effective measure. Vivotif® is an oral live attenuated vaccine which
contains a mutated strain of Salmonella (Ty21a) and reproduces the natural
infection. The vaccine was first licensed in Europe in 1983 and in the US in
1989, and over the years it has proved efficacious and safe. It is indicated for
adults and children from 5 years of age upwards. Specifically, in the most
developed countries, vaccination is suggested for highrisk population groups and
particularly for international travellers to destinations where the risk of
contracting typhoid fever is high. It must also be borne in mind that
international travel is increasing. Indeed, international tourist arrivals
totalled 1,184 million in 2015 and, on the basis of current trends,
international travel is expected to grow by 3-4% in 2017. Vivotif® appears to be
a powerful means of disease prevention, the importance of which is highlighted
by the spread of antibiotic-resistant strains of Salmonella typhy (S. typhi). |
Are stem cell transplants used to treat acute kidney injury? | Yes, stem cell transplantation is becoming the treatment of choice for complicated acute kidney injury. | OBJECTIVE: This is a retrospective study for risk assessment of acute kidney
injury after allogeneic hematopoietic stem cell transplantation (allo HSCT)
based on the Acute Kidney Injury Network (AKIN) criteria.
METHODS: Two hundred and eighty-nine consecutive patients who received allo HSCT
were studied retrospectively to identify the risk factors for AKI according to
the AKIN criteria. The incidence of AKI based on AKIN staging and overall
survival (OS) was evaluated using Cox proportional hazard regression models
treating each AKIN stage as a time-dependent covariate.
PATIENTS: We identified a total of 180 patients who developed AKI within 100
days after allo HSCT; AKI was classified as stage 1 in 88 patients (30.5%),
stage 2 in 46 patients (15.9%) and stage 3 in 46 patients (15.9%).
RESULTS: Patients who developed stage 3 AKI had a significantly worse survival
compared to those who developed no AKI or lower stage AKI (HR: 7.6, 95%CI:
4.8-12.1; p<0.001). Multivariate analysis for risks for developing AKI revealed
an episode of sepsis or sinusoidal obstruction syndrome (SOS) and the use of
liposomal amphotericin as a major cause of the severe stage of AKI.
CONCLUSION: On the basis of our analysis, sepsis, hemorrhagic cystitis, and
acute GVHD were associated with severe AKI, and SOS was associated any stage of
AKI. Animal studies have shown that mesenchymal stromal cell (MSC) infusions improve
acute kidney injury (AKI) outcomes when administered early after
ischemic/reperfusion injury or within 24 hours after cisplatin administration.
These findings have spurred several human clinical trials to prevent AKI.
However, no specific therapy effectively treats clinically obvious AKI or
rescues renal function once advanced injury is established. We investigated if
noninvasive image-guided pulsed focused ultrasound (pFUS) could alter the kidney
microenvironment to enhance homing of subsequently infused MSC. To examine the
efficacy of pFUS-enhanced cell homing in disease, we targeted pFUS to kidneys to
enhance MSC homing after cisplatin-induced AKI. We found that pFUS enhanced MSC
homing at 1 day post-cisplatin, prior to renal functional deficits, and that
enhanced homing improved outcomes of renal function, tubular cell death, and
regeneration at 5 days post-cisplatin compared to MSC alone. We then
investigated whether pFUS+MSC therapy could rescue established AKI. MSC alone at
3 days post-cisplatin, after renal functional deficits were obvious,
significantly improved 7-day survival of animals. Survival was further improved
by pFUS and MSC. pFUS prior to MSC injections increased IL-10 production by MSC
that homed to kidneys and generated an anti-inflammatory immune cell profile in
treated kidneys. This study shows pFUS is a neoadjuvant approach to improve MSC
homing to diseased organs. pFUS with MSC better prevents AKI than MSC alone and
allows rescue therapy in established AKI, which currently has no meaningful
therapeutic options. |
What is the interaction between WAPL and PDS5 proteins? | We propose that Wapl and Pds5 directly modulate conformational changes of cohesin to make it competent for dissociation from chromatin during prophase. | The cohesin complex establishes sister chromatid cohesion during S phase. In
metazoan cells, most if not all cohesin dissociates from chromatin during
mitotic prophase, leading to the formation of metaphase chromosomes with two
cytologically discernible chromatids. This process, known as sister chromatid
resolution, is believed to be a prerequisite for synchronous separation of
sister chromatids in subsequent anaphase. To dissect this process at a
mechanistic level, we set up an in vitro system. Sister chromatid resolution is
severely impaired upon depletion of Wapl from Xenopus egg extracts. Exogenously
added human Wapl can rescue these defects and, remarkably, it can do so in a
very short time window of early mitosis. A similar set of observations is made
for Pds5, a factor implicated previously in the stabilization of interphase
cohesion. Characteristic amino acid motifs (the FGF motifs) in Wapl coordinate
its physical and functional interactions with Pds5 and cohesin subunits. We
propose that Wapl and Pds5 directly modulate conformational changes of cohesin
to make it competent for dissociation from chromatin during prophase. Evidence
is also presented that Sgo1 plays a hitherto underappreciated role in
stabilizing cohesin along chromosome arms, which is antagonized by the mitotic
kinases polo-like kinsase (Plk1) and aurora B. Cohesin is a ring-shaped protein complex that plays a crucial role in sister
chromatid cohesion and gene expression. The dynamic association of cohesin with
chromatin is essential for these functions. However, the exact nature of cohesin
dynamics, particularly cohesin translocation, remains unclear. We evaluated the
dynamics of individual cohesin molecules on DNA and found that the cohesin core
complex possesses an intrinsic ability to traverse DNA in an adenosine
triphosphatase (ATPase)-dependent manner. Translocation ability is suppressed in
the presence of Wapl-Pds5 and Sororin; this suppression is alleviated by the
acetylation of cohesin and the action of mitotic kinases. In Xenopus laevis egg
extracts, cohesin is translocated on unreplicated DNA in an ATPase- and Smc3
acetylation-dependent manner. Cohesin movement changes from bidirectional to
unidirectional when cohesin faces DNA replication; otherwise, it is incorporated
into replicating DNA without being translocated or is dissociated from
replicating DNA This study provides insight into the nature of individual
cohesin dynamics and the mechanisms by which cohesin achieves cohesion in
different chromatin contexts. The ring-shaped ATPase machine, cohesin, regulates sister chromatid cohesion,
transcription, and DNA repair by topologically entrapping DNA. Here, we propose
a rigid scaffold model to explain how the cohesin regulators Pds5 and Wapl
release cohesin from chromosomes. Recent studies have established the Smc3-Scc1
interface as the DNA exit gate of cohesin, revealed a requirement for ATP
hydrolysis in ring opening, suggested regulation of the cohesin ATPase activity
by DNA and Smc3 acetylation, and provided insights into how Pds5 and Wapl open
this exit gate. We hypothesize that Pds5, Wapl, and SA1/2 form a rigid scaffold
that docks on Scc1 and anchors the N-terminal domain of Scc1 (Scc1N) to the Smc1
ATPase head. Relative movements between the Smc1-3 ATPase heads driven by ATP
and Wapl disrupt the Smc3-Scc1 interface. Pds5 binds the dissociated Scc1N and
prolongs this open state of cohesin, releasing DNA. We review the evidence
supporting this model and suggest experiments that can further test its key
principles. The cohesin complex topologically encircles chromosomes and mediates sister
chromatid cohesion to ensure accurate chromosome segregation upon cell division.
Cohesin also participates in DNA repair and gene transcription. The
Nipped-B-Mau2 protein complex loads cohesin onto chromosomes and the Pds5-Wapl
complex removes cohesin. Pds5 is also essential for sister chromatid cohesion,
indicating that it has functions beyond cohesin removal. The Brca2 DNA repair
protein interacts with Pds5, but the roles of this complex beyond DNA repair are
unknown. Here we show that Brca2 opposes Pds5 function in sister chromatid
cohesion by assaying precocious sister chromatid separation in metaphase spreads
of cultured cells depleted for these proteins. By genome-wide chromatin
immunoprecipitation we find that Pds5 facilitates SA cohesin subunit association
with DNA replication origins and that Brca2 inhibits SA binding, mirroring their
effects on sister chromatid cohesion. Cohesin binding is maximal at replication
origins and extends outward to occupy active genes and regulatory sequences.
Pds5 and Wapl, but not Brca2, limit the distance that cohesin extends from
origins, thereby determining which active genes, enhancers and silencers bind
cohesin. Using RNA-seq we find that Brca2, Pds5 and Wapl influence the
expression of most genes sensitive to Nipped-B and cohesin, largely in the same
direction. These findings demonstrate that Brca2 regulates sister chromatid
cohesion and gene expression in addition to its canonical role in DNA repair and
expand the known functions of accessory proteins in cohesin's diverse functions. |
Is deletion at 6q24.2-26 associated with shorter survival for ovarian cancer patients? | No, the 6q24.2-26 deletion is an independent marker of favorable outcome in high-grade serous ovarian carcinoma (HGSOC) patients with potential clinical value as it can be analyzed by FISH on tumor sections and guide the selection of patients towards more conservative therapeutic strategies in order to reduce side-effects and improve quality of life. | Author information:
(1)Human Genetics Group, Spanish National Cancer Research Center (CNIO), C/
Melchor Fernández Almagro 3, 28029, Madrid, Spain.
(2)Structural Computational Biology Group, Spanish National Cancer Research
Center (CNIO), C/ Melchor Fernández Almagro 3 28029, Madrid, Spain.
(3)Cancer Epidemiology Centre, Cancer Council Victoria, 615 St Kilda Road,
Melbourne 3004, Australia; Center for Epidemiology and Biostatistics, Melbourne
School of Population and Global Health, The University of Melbourne, Level 3,
207 Bouverie Street Carlton, Melbourne 3010, Victoria, Australia.
(4)Molecular Cytogenetics Group, Spanish National Cancer Research Center (CNIO),
C/ Melchor Fernández Almagro 3, 28029 Madrid, Spain.
(5)Departments of Genetics, Reproduction, and Fetal Medicine, IBIS, University
Hospital Virgen del Rocio/CSIC/University of Seville, Avda. Manuel Siurot, s/n.,
41013 Sevilla, Spain; Biomedical Network Research Centre on Rare Diseases
(CIBERER), Spain.
(6)Pathology Department, Fundación Jiménez Díaz, Avda. Reyes Católicos, 2, 28040
Madrid, Spain.
(7)Oncology Department, Hospital General de Albacete, Calle Hermanos Falco, 37,
02006 Albacete, Spain.
(8)Oncology Department, Fundación Hospital Alcorcón, Calle Valdelaguna, 1, 28922
Alcorcón, Spain.
(9)Medical Oncology Service, Oncologic Center Clara Campal, Calle Oña, 10, 28050
Madrid, Spain.
(10)Breast Cancer Clinical Research Unit, Spanish National Cancer Research
Center (CNIO), C/ Melchor Fernández Almagro 3, 28029 Madrid, Spain.
(11)Medical Oncology Service, Hospital Sant Pau, Carrer de Sant Quintí, 89,
08026 Barcelona, Spain.
(12)Familial Cancer Unit and Medical Oncology Department, Hospital 12 de
Octubre, Avda de Córdoba, s/n, 28041 Madrid, Spain.
(13)Medical Oncology Service, Instituto de Investigación Sanitaria Gregorio
Marañón, Universidad Complutense, Calle Doctor Esquerdo, 46, 28007 Madrid,
Spain.
(14)Obstetrics and Gynecology Department, Institut Universitari Dexeus, Carrer
de Sabino Arana, 5, 08028 Barcelona, Spain.
(15)Laboratory of Genetics, Hospital Donostia, Calle Doctor Begiristain, 117,
20080 San Sebastián, Spain.
(16)Department of Internal Medicine, Hospital Severo Ochoa, Avd. de Orellana,
s/n., 28911 Madrid, Spain.
(17)Human Genetics Group, Spanish National Cancer Research Center (CNIO), C/
Melchor Fernández Almagro 3, 28029, Madrid, Spain; Biomedical Network Research
Centre on Rare Diseases (CIBERER), Spain.
(18)Familial Cancer Clinical Unit, Spanish National Cancer Research Center
(CNIO), C/ Melchor Fernández Almagro 3, 28029 Madrid, Spain; Biomedical Network
Research Centre on Rare Diseases (CIBERER), Spain.
(19)Molecular Cytogenetics Group, Spanish National Cancer Research Center
(CNIO), C/ Melchor Fernández Almagro 3, 28029 Madrid, Spain; Biomedical Network
Research Centre on Rare Diseases (CIBERER), Spain.
(20)Medical Oncology Department, University Hospital Virgen del Rocio, Avda.
Manuel Siurot s/n., 41013 Sevilla, Spain.
(21)Pathology Department, Hospital Universitario Ramón y Cajal, Ctra. de
Colmenar Viejo, km. 9,100, 28034 Madrid, Spain.
(22)Human Genetics Group, Spanish National Cancer Research Center (CNIO), C/
Melchor Fernández Almagro 3, 28029, Madrid, Spain; Biomedical Network Research
Centre on Rare Diseases (CIBERER), Spain. Electronic address: [email protected]. |
What is the function of a protein kinase? | Protein kinases are enzymes that add a phosphate (PO4) group to a protein, and can modulate its function. | Protein kinases are known primarily for their ability to phosphorylate protein
substrates, which constitutes an essential biological process. Recently,
compelling evidence has accumulated that the functions of many protein kinases
extend beyond phosphorylation and include an impressive spectrum of
non-catalytic roles, such as scaffolding, allosteric regulation, or even
protein-DNA interactions. How the conserved kinase fold shared by all metazoan
protein kinases can accomplish these diverse tasks in a specific and regulated
manner is poorly understood. In this review, we analyze the molecular mechanisms
supporting phosphorylation-independent signaling by kinases and attempt to
identify common and unique structural characteristics that enable kinases to
perform non-catalytic functions. We also discuss how post-translational
modifications, protein-protein interactions, and small molecules modulate these
non-canonical kinase functions. Finally, we highlight current efforts in the
targeted design of small-molecule modulators of non-catalytic kinase functions,
a new pharmacological challenge for which structural considerations are more
important than ever. Diacylglycerol kinase (DGK) phosphorylates diacylglycerol (DG) to produce
phosphatidic acid (PA). Mammalian DGK comprises ten isozymes (α-κ) and regulates
a wide variety of physiological and pathological events, such as cancer, type II
diabetes, neuronal disorders and immune responses. DG and PA consist of various
molecular species that have different acyl chains at the sn-1 and sn-2
positions, and consequently, mammalian cells contain at least 50 structurally
distinct DG/PA species. Because DGK is one of the components of
phosphatidylinositol (PI) turnover, the generally accepted dogma is that all DGK
isozymes utilize 18:0/20:4-DG derived from PI turnover. We recently established
a specific liquid chromatography-mass spectrometry method to analyze which PA
species were generated by DGK isozymes in a cell stimulation-dependent manner.
Interestingly, we determined that DGKδ, which is closely related to the
pathogenesis of type II diabetes, preferentially utilized 14:0/16:0-,
14:0/16:1-, 16:0/16:0-, 16:0/16:1-, 16:0/18:0- and 16:0/18:1-DG species
(X:Y = the total number of carbon atoms: the total number of double bonds)
supplied from the phosphatidylcholine-specific phospholipase C pathway, but not
18:0/20:4-DG, in high glucose-stimulated C2C12 myoblasts. Moreover, DGKα mainly
consumed 14:0/16:0-, 16:0/18:1-, 18:0/18:1- and 18:1/18:1-DG species during cell
proliferation in AKI melanoma cells. Furthermore, we found that 16:0/16:0-PA was
specifically produced by DGKζ in Neuro-2a cells during retinoic acid- and serum
starvation-induced neuronal differentiation. These results indicate that DGK
isozymes utilize a variety of DG molecular species derived from PI
turnover-independent pathways as substrates in different stimuli and cells. DGK
isozymes phosphorylate various DG species to generate various PA species. It was
revealed that the modes of activation of conventional and novel protein kinase
isoforms by DG molecular species varied considerably. However, PA
species-selective binding proteins have not been found to date. Therefore, we
next attempted to identify PA species-selective binding proteins from the mouse
brain and identified α-synuclein, which has causal links to Parkinson's disease.
Intriguingly, we determined that among phospholipids, including several PA
species (16:0/16:0-PA, 16:0/18:1-PA, 18:1/18:1-PA, 18:0/18:0-PA and
18:0/20:4-PA); 18:1/18:1-PA was the most strongly bound PA to α-synuclein.
Moreover, 18:1/18:1-PA strongly enhanced secondary structural changes from the
random coil form to the α-helix form and generated a multimeric and proteinase
K-resistant α-synuclein protein. In contrast with the dogma described above, our
recent studies strongly suggest that PI turnover-derived DG species and also
various DG species derived from PI turnover-independent pathways are utilized by
DGK isozymes. DG species supplied from distinct pathways may be utilized by DGK
isozymes based on different stimuli present in different types of cells, and
individual PA molecular species would have specific targets and exert their own
physiological functions. Several studies have revealed that cyclin-dependent kinases (CDK) can mediate
phosphorylation of steroid receptors at multiple sites, including serine 81 of
the androgen receptor (AR). Phosphorylation of S81 is required for AR nuclear
translocation, an association with chromatin and also regulates endogenous
AR-regulated transcription in response to hormones. Up to date,
S81-phosphorylation has been studied using different CDK inhibitors.
Nevertheless, most inhibitors are non-selective or have unknown selectivity. We
investigated the selectivity of commercially available CDK inhibitors and
identified compounds that will be suitable for further studies to identify the
CDKs responsible for S81-AR phosphorylation. We confirmed the positive impact of
CDK1 and CDK9 on phosphorylation of S81-AR and its transcriptional activity.
Although CDK1-mediated phosphorylation was previously shown to occur during
mitosis, our experiments did not confirm this finding. By using chemical and
genetic inhibition techniques, we identified that CDK2 contributes to S81-AR
phosphorylation and transactivation while CDK4 was not shown to be involved in
this process. Src family kinases (SFKs) are a family of protein tyrosine kinases containing
nine members: Src, Lyn, Fgr, Hck, Lck, Fyn, Blk, Yes, and Ylk. Although SFK
activation is a major immediate signaling event in LPS/Toll-like receptor 4
(TLR4) signaling, its precise role has remained elusive due to various
contradictory results obtained from a certain SFK member-deficient mice or
cells. The observed inconsistencies may be due to the compensation or redundancy
by other SFKs upon a SFK deficiency. The chemical rescuing approach was
suggested to induce temporal and precise SFK activation in living cells, thereby
limiting the chance of cellular adaption to a SFK-deficient condition. Using the
rescuing approach, we demonstrate that restoring SFK activity not only induces
tyrosine phosphorylation of TLR4, but also inhibits LPS-induced NFκB and JNK1/2
activation and consequently suppresses LPS-induced cytokine production. TLR4
normally recruits TIR domain-containing adaptors in response to LPS, however,
temporally restored SFK activation disrupts the LPS-induced association of MyD88
and Mal/Tirap with TLR4. Additionally, using kinase-dead SFK-Lyn (Y397/508F) and
constitutively active SFK-Lyn (Y508F), we found that the kinase-dead SFK
inhibits TLR4 tyrosine phosphorylation with reduced binding affinity to TLR4,
while the kinase-active SFK strongly binds to TLR4 and promotes TLR4 tyrosine
phosphorylation, suggesting that SFK kinase activity is required for TLR4
tyrosine phosphorylation and TLR4-SFK interaction. Together, our results
demonstrate that SFK activation induces TLR4 tyrosine phosphorylation,
consequently dissociating MyD88 and Mal/Tirap from TLR4 and inhibiting
LPS-induced inflammatory responses, suggesting a negative feedback loop
regulated by SFK-induced tyrosine phosphorylation in TLR4. Precise regulation of kinetochore-microtubule attachments is essential for
successful chromosome segregation. Central to this regulation is Aurora B
kinase, which phosphorylates kinetochore substrates to promote microtubule
turnover. A critical target of Aurora B is the N-terminal "tail" domain of Hec1,
which is a component of the NDC80 complex, a force-transducing link between
kinetochores and microtubules. Although Aurora B is regarded as the "master
regulator" of kinetochore-microtubule attachment, other mitotic kinases likely
contribute to Hec1 phosphorylation. In this study, we demonstrate that Aurora A
kinase regulates kinetochore-microtubule dynamics of metaphase chromosomes, and
we identify Hec1 S69, a previously uncharacterized phosphorylation target site
in the Hec1 tail, as a critical Aurora A substrate for this regulation.
Additionally, we demonstrate that Aurora A kinase associates with inner
centromere protein (INCENP) during mitosis and that INCENP is competent to drive
accumulation of the kinase to the centromere region of mitotic chromosomes.
These findings reveal that both Aurora A and B contribute to
kinetochore-microtubule attachment dynamics, and they uncover an unexpected role
for Aurora A in late mitosis. |
Which was the first cholera vaccine approved in the US? | Vaxchora is the first vaccine approved by the Food and Drug Administration for the prophylaxis of cholera infection. | Vaxchora is the first vaccine approved by the Food and Drug Administration for
the prophylaxis of cholera infection. Cholera, a potentially life-threatening
bacterial infection that occurs in the intestines and causes severe diarrhea and
dehydration, has a low incidence in the U.S., but a high incidence in Africa,
Southeast Asia, and other locations around the world. These areas draw travelers
from the U.S., so cholera can present in patients who return from visits to
these regions. Previous means of prophylaxis included the use of doxycycline for
the prevention of traveler's diarrhea, but doxycycline is not specific for
cholera. With the approval of Vaxchora, a live attenuated, single-dose, oral
suspension vaccine, travelers can now visit these areas with less chance of
contracting the bacterium Vibrio cholerae, which causes cholera infections. |
List 3 PD-L1 inhibitors on the market as of 2018. | Atezolizumab (Tecentriq), Avelumab (Bavencio), and Durvalumab (Imfinzi) are PD-L1 inhibitors | Author information:
(1)Département de biopathologie et département de recherche translationnelle et
d'innovations, centre Léon-Bérard UNICANCER, 28, rue Laennec, 69008 Lyon,
France; Inserm U1209/CNRS 5309, Grenoble-Alpes université, Institute for
Advanced Biosciences, 38700 La Tronche, France. Electronic address:
[email protected].
(2)Département de biologie et pathologie médicales, Gustave-Roussy, 114, rue
Edouard-Vaillant, 94805 Villejuif, France; Inserm U981, Gustave-Roussy, 94805
Villejuif, France.
(3)Institut du thorax Curie-Montsouris, institut Curie, 75005 Paris, France;
Université Claude-Bernard Lyon 1, université de Lyon, 69622 Villeurbanne,
France.
(4)Inserm 1052, CNRS 5286, centre de recherche en cancérologie de Lyon, institut
de cancérologie des Hospices Civiles de Lyon (IC-HCL), service de pneumologie,
hôpital Louis-Pradel, 69008 Lyon, France.
(5)Département de pathologie, hôpital Cochin, université Paris Descartes,
Assistance publique-hôpitaux de Paris, 74014 Paris, France.
(6)Inserm UMR1152, département de pathologie, hôpital Bichat, université Paris
Diderot, 75018 Paris, France.
(7)Département de pathologie, IUCT Oncopôle, CHU de Toulouse, 31059 Toulouse,
France.
(8)Département de pathologie, hôpital européen Georges-Pompidou, Assistance
publique-hôpitaux de Paris, 75015 Paris, France; Inserm UMR-S970, Paris centre
de recherche cardiovasculaire, Georges-Pompidou European Hospital, 75015 Paris,
France.
(9)Département de Pathologie, hôpital Nord, Assistance publique-hôpitaux de
Marseille, Aix-Marseille Université, CRCM, 13015 Marseille, France.
(10)Service d'anatomie pathologique, hôpital Tenon AP-HP, 75020 Paris, France;
UPMC université Paris 06, GRC n(o) 04, Theranoscan, 75252 Paris, France.
(11)Département de pathologie, CHRU de Nancy, 54035 Nancy, France; Inserm 1256,
université de Lorraine, 54505 Vandœuvre-lès-Nancy, France.
(12)Centre National Référent MESOPATH, Base Clinicobiologique nationale
MESOBANK, Registre multicentrique MESONAT centre Leon-Bérard, département de
biopathologie, 69008 Lyon, France.
(13)Inserm U1087, institut du Thorax, service d'anatomie et cytologique
pathologiques, hôpital Hotel-Dieu, CHU de Nantes, 44093 Nantes, France.
(14)UMR Inserm 1240 IMoST, Centre Jean-Perrin, département de pathologie,
université Clermont-Auvergne, 63011 Clermont-Ferrand, France. |
Are there negative enhancers? | Yes, negative enhancers are also called gene silencers. | We have previously identified a silencer (negative enhancer) in glutathione
transferase P (GST-P) gene which is strongly and specifically induced during
hepatocarcinogenesis of the rat. At least three trans-acting factors bind to
multiple cis-elements located in this silencer. One of these factors, SF-B
(Silencer Factor B) specifically binds to GPS1 (GST-P Silencer 1) and has been
cloned by a Southwestern protocol. Analysis of DNA and deduced amino acid
sequence reveals that SF-B clone is most likely identical to an IL-6 inducible
trans-activator LAP/IL6-DBP. Binding efficiency of SF-B to GPS1 is
indistinguishable from that to IL6-responsive element found in C-reactive
protein gene. The possibility that SF-B/LAP/IL6-DBP functions as a dual positive
and negative regulator is discussed. Calcium-dependent protease (CANP, Calpain) is an intracellular protease involved
in essential cellular functions mediated by calcium. To understand the mechanism
regulating the expression of CANP at the transcriptional level, we isolated a
human gene for the large subunit of mCANP (CANP mL) and analyzed its 5'-region.
The transcription initiation sites were mapped to multiple positions (-142 to
-103, A of initiation ATG as +1). The upstream region lacks typical promoter
elements such as TATA and CAAT boxes and is characterized by its high GC content
(-300 to -20, 70% GC content). Functional analyses of the 5'-region by a
transient expression assay on HeLa cells revealed that the region (-202 to -80)
has a promoter activity. The upstream half of the promoter region (-202 to -130)
acts as an upstream promoter element in an orientation-independent manner.
Upstream of the promoter region are tandemly reiterated multiple regulatory
regions (-2.5k to -690, -690 to -460, -460 to -260, and -260 to -202), each of
which negatively regulates the CANP mL gene promoter as well as heterologous
promoters in an orientation-independent manner. The presence of a cellular
factor(s) mediating the action of these positive (promoter) and negative
regulatory elements was suggested by an in vivo competition assay. The negative
regulation of transcription mediated by these reiterated cis-acting elements and
trans-acting factor(s) may play an essential role in the expression of the CANP
mL gene. The E1a gene of adenovirus encodes two proteins, 289 and 243 amino acids long,
which have positive (transactivator) and negative (enhancer repressor) RNA
polymerase II transcriptional regulatory properties and cell transformation
activities including cooperation with an activated ras gene. The E1a
transforming functions more closely correlate with the repressor property than
with transactivation in that both E1a proteins express the repressor and
transformation functions while only the 289-amino-acid protein is an efficient
transactivator. To understand whether the transcriptional regulatory activities
of E1a are related to its ras cooperation activity, we generated a series of
mutant E1a expression vectors by linker insertion mutagenesis of the
289-amino-acid protein. Here we describe a new class of mutants which although
defective for enhancer repression still can cooperate with the ras oncogene in
cell transformation. The mutants are also defective in transcription
transactivation. Our data suggest that enhancer repression and transformation
via ras cooperation are separate E1a functions and that cooperation with ras
does not rely on either of the RNA polymerase II transcription regulatory
functions of E1a. We also show that mutations which inactivate enhancer
repression are not confirmed to a single critical domain necessary for
repression. We therefore propose that the integrity of the overall configuration
of the E1a proteins is important for the repression activity. Muscle development involves the coordinated regulation of transcription of
muscle-type-specific genes and their encoded proteins during myogenesis. We show
here that transcriptional regulation of the Drosophila tropomyosin I (TmI) gene
during myogenesis is under the control of at least two muscle enhancer regions
located within the first intron of the gene. Together these enhancer regions
contain multiple muscle-type-specific positive and negative cis-acting elements
which together contribute toward full expression of the gene. One of these
enhancers is contained within a 355-bp fragment that is sufficient to direct
high levels of temporally regulated expression from a heterologous promoter in
all muscles of transgenic flies. Dissection of this enhancer region into smaller
fragments has allowed us to identify a 91-bp enhancer fragment sufficient for
directing expression in all somatic and visceral muscles of the larva and adult
but not in the indirect flight muscles and tergal depressor of the trochanter or
jump muscles of the adult. We also show that this somatic/visceral muscle
element(s) can be repressed through an adjacent negative control region,
suggesting that the regulation of expression in these muscles is under dual
control during both phases of myogenesis. We propose a model in which
transcriptional regulation of the Drosophila TmI gene is controlled by the
cooperative interaction of multiple positive and negative cis-acting regulatory
elements that control the temporal and muscle-type pattern of expression. The
distribution of enhancer elements and their control of TmI gene expression are
similar to those regulating transcription of the muscle promoter of the TmII
gene and provide a framework for the coordinate expression of the two genes. Substantial evidence supports the hypothesis that enhancers are critical
regulators of cell-type determination, orchestrating both positive and negative
transcriptional programs; however, the basic mechanisms by which enhancers
orchestrate interactions with cognate promoters during activation and repression
events remain incompletely understood. Here we report the required actions of
LIM domain-binding protein 1 (LDB1)/cofactor of LIM homeodomain protein
2/nuclear LIM interactor, interacting with the enhancer-binding protein
achaete-scute complex homolog 1, to mediate looping to target gene promoters and
target gene regulation in corticotrope cells. LDB1-mediated enhancer:promoter
looping appears to be required for both activation and repression of these
target genes. Although LDB1-dependent activated genes are regulated at the level
of transcriptional initiation, the LDB1-dependent repressed transcription units
appear to be regulated primarily at the level of promoter pausing, with LDB1
regulating recruitment of metastasis-associated 1 family, member 2, a component
of the nucleosome remodeling deacetylase complex, on these negative enhancers,
required for the repressive enhancer function. These results indicate that
LDB1-dependent looping events can deliver repressive cargo to cognate promoters
to mediate promoter pausing events in a pituitary cell type. |
What is the function of WAPL protein on cohesin? | Human Wapl is a cohesin-binding protein that promotes sister-chromatid resolution in mitotic prophase We show that the human ortholog of Wapl is a cohesin-binding protein that facilitates cohesin's timely release from chromosome arms during prophase. | BACKGROUND: The linkage between duplicated chromosomes (sister chromatids) is
established during S phase by the action of cohesin, a multisubunit complex
conserved from yeast to humans. Most cohesin dissociates from chromosome arms
when the cell enters mitotic prophase, leading to the formation of metaphase
chromosomes with two cytologically discernible chromatids. This process is known
as sister-chromatid resolution. Although two mitotic kinases have been
implicated in this process, it remains unknown exactly how the cohesin-mediated
linkage is destabilized at a mechanistic level.
RESULTS: The wings apart-like (Wapl) protein was originally identified as a gene
product that potentially regulates heterochromatin organization in Drosophila
melanogaster. We show that the human ortholog of Wapl is a cohesin-binding
protein that facilitates cohesin's timely release from chromosome arms during
prophase. Depletion of Wapl from HeLa cells causes transient accumulation of
prometaphase-like cells with chromosomes that display poorly resolved sister
chromatids with a high level of cohesin. Reduction of cohesin relieves the
Wapl-depletion phenotype, and depletion of Wapl rescues premature sister
separation observed in Sgo1-depleted or Esco2-depleted cells. Conversely,
overexpression of Wapl causes premature separation of sister chromatids. Wapl
physically associates with cohesin in HeLa-cell nuclear extracts. Remarkably, in
vitro reconstitution experiments demonstrate that Wapl forms a stoichiometric,
ternary complex with two regulatory subunits of cohesin, implicating its
noncatalytic function in inactivating cohesin's ability to interact with
chromatin.
CONCLUSIONS: Wapl is a new regulator of sister chromatid resolution and promotes
release of cohesin from chromosomes by directly interacting with its regulatory
subunits. Cohesin establishes sister-chromatid cohesion from S phase until mitosis or
meiosis. To allow chromosome segregation, cohesion has to be dissolved. In
vertebrate cells, this process is mediated in part by the protease separase,
which destroys a small amount of cohesin, but most cohesin is removed from
chromosomes without proteolysis. How this is achieved is poorly understood.
Here, we show that the interaction between cohesin and chromatin is controlled
by Wapl, a protein implicated in heterochromatin formation and tumorigenesis.
Wapl is associated with cohesin throughout the cell cycle, and its depletion
blocks cohesin dissociation from chromosomes during the early stages of mitosis
and prevents the resolution of sister chromatids until anaphase, which occurs
after a delay. Wapl depletion also increases the residence time of cohesin on
chromatin in interphase. Our data indicate that Wapl is required to unlock
cohesin from a particular state in which it is stably bound to chromatin. The classical X shape of mitotic human chromosomes is the consequence of two
distinct waves of cohesin removal. First, during prophase and prometaphase, the
bulk of cohesin is driven from chromosome arms by the cohesin antagonist WAPL.
This arm-specific cohesin removal is referred to as the prophase pathway [1-4].
The subsequent cleavage of the remaining centromeric cohesin by Separase is
known to be the trigger for anaphase onset [5-7]. Remarkably the biological
purpose of the prophase pathway is unknown. We find that this pathway is
essential for two key mitotic processes. First, it is important to focus Aurora
B at centromeres to allow efficient correction of erroneous
microtubule-kinetochore attachments. In addition, it is required to facilitate
the timely decatenation of sister chromatids. As a consequence, WAPL-depleted
cells undergo anaphase with segregation errors, including both lagging
chromosomes and catees, resulting in micronuclei and DNA damage. Stable WAPL
depletion arrests cells in a p53-dependent manner but causes p53-deficient cells
to become highly aneuploid. Our data show that the WAPL-dependent prophase
pathway is essential for proper chromosome segregation and is crucial to
maintain genomic integrity. During the cell cycle, duplicated sister chromatids become physically connected
during S phase through a process called sister-chromatid cohesion. Cohesion is
terminated during the metaphase-to-anaphase transition to trigger
sister-chromatid segregation. The establishment and dissolution of cohesion are
highly regulated by the cohesin complex and its multitude of regulators. In
particular, the cohesin regulator Wapl promotes the release of cohesin from
chromosomes during both interphase and mitosis. Here, we describe in vitro
protein binding assays between Wapl and a cohesin subcomplex, and cellular
assays in human cells that probe the functions of Wapl in cohesin release. The spatial organization of chromosomes influences many nuclear processes
including gene expression. The cohesin complex shapes the 3D genome by looping
together CTCF sites along chromosomes. We show here that chromatin loop size can
be increased and that the duration with which cohesin embraces DNA determines
the degree to which loops are enlarged. Cohesin's DNA release factor WAPL
restricts this loop extension and also prevents looping between incorrectly
oriented CTCF sites. We reveal that the SCC2/SCC4 complex promotes the extension
of chromatin loops and the formation of topologically associated domains (TADs).
Our data support the model that cohesin structures chromosomes through the
processive enlargement of loops and that TADs reflect polyclonal collections of
loops in the making. Finally, we find that whereas cohesin promotes chromosomal
looping, it rather limits nuclear compartmentalization. We conclude that the
balanced activity of SCC2/SCC4 and WAPL enables cohesin to correctly structure
chromosomes. The cohesin complex topologically encircles chromosomes and mediates sister
chromatid cohesion to ensure accurate chromosome segregation upon cell division.
Cohesin also participates in DNA repair and gene transcription. The
Nipped-B-Mau2 protein complex loads cohesin onto chromosomes and the Pds5-Wapl
complex removes cohesin. Pds5 is also essential for sister chromatid cohesion,
indicating that it has functions beyond cohesin removal. The Brca2 DNA repair
protein interacts with Pds5, but the roles of this complex beyond DNA repair are
unknown. Here we show that Brca2 opposes Pds5 function in sister chromatid
cohesion by assaying precocious sister chromatid separation in metaphase spreads
of cultured cells depleted for these proteins. By genome-wide chromatin
immunoprecipitation we find that Pds5 facilitates SA cohesin subunit association
with DNA replication origins and that Brca2 inhibits SA binding, mirroring their
effects on sister chromatid cohesion. Cohesin binding is maximal at replication
origins and extends outward to occupy active genes and regulatory sequences.
Pds5 and Wapl, but not Brca2, limit the distance that cohesin extends from
origins, thereby determining which active genes, enhancers and silencers bind
cohesin. Using RNA-seq we find that Brca2, Pds5 and Wapl influence the
expression of most genes sensitive to Nipped-B and cohesin, largely in the same
direction. These findings demonstrate that Brca2 regulates sister chromatid
cohesion and gene expression in addition to its canonical role in DNA repair and
expand the known functions of accessory proteins in cohesin's diverse functions. |
What is romiplostim targeting? | Romiplostim (ROM) is a thrombopoietin (TPO)-receptor-agonist approved for treatment of adults with chronic immune thrombocytopenia (ITP). | |
Please list 3 drugs that have EGFR as their primary target. | There are a number of drugs that target EGFR. The best known targeted therapies include: Epidermal growth factor receptor (EGFR) monoclonal antibodies (cetuximab, panitumumab, zalutumumab and nimotuzumab), EGFR tyrosine kinase inhibitors (gefitinib, erlotinib, lapatinib, afatinib and dacomitinib) | Lung cancer is the foremost cause of cancer-related deaths world-wide. Both, the
major forms of lung cancer, Non-small cell lung cancer (NSCLC) and Small cell
lung cancers (SCLC), have responded effectively to chemo-, radiation and
adjuvant-therapies. Tumor removal through surgery also appeared as a good
therapeutic strategy. However, these therapies demonstrated unfavourable
side-effects, and hence novel drugs targeting lung cancer emerged essential.
Activation of epidermal growth factor receptor (EGFR)-tyrosine kinases is a key
reason for lung cancer progression. Two important strategies that have
attenuated lung cancers were through treatments with EGFR-tyrosine
kinase-inhibitors, erlotinib and gefitinib, or EGFR-neutralizing antibodies,
cetuximab and bevacizumab. A major advantage with erlotinib and gefitinib was
their role in second and third-line treatments following chemotherapies. Phase
II/III clinical trials showed that combinatorial treatment of tyrosine kinase
(TK)-inhibitors with chemotherapeutics, such as docetaxel and pemetrexed, caused
significant improvements in progression-free survival and overall survival.Phase
I and II clinical studies also revealed that combination of tyrosine
kinase-inhibitors with the EGFR-targeted antibodies was an effective approach
for treating lung cancer. However, patients having T790M-mutations within EGFR
gene were resistant to erlotinib and gefitinib. Alternatively, another
second-generation EGFR-tyrosine kinase-inhibitor, afatinib, that could
circumvent the problem of drug resistance has been developed as lung cancer
therapy. The current review focuses on the role of EGFR in lung cancer
progression and apprises about the EGFR-targeted therapies. The review also
informs on the adverse side-effects of these therapies and enlightens the need
for safer therapeutic regimens to eradicate this dreaded disease. Epidermal growth factor receptor (EGFR) is a known target in cancer therapy and
targeting the receptor has proven to be extremely successful in treating cancers
that are dependent on EGFR signaling. To that effect, targeted therapies to EGFR
such as Cetuximab, Panitumumab-monoclonal antibodies and Gefitinib,
Erlotinib-tyrosine kinase inhibitors have had success in therapeutic scenarios.
However, the development of resistance to these drugs makes it necessary to
combine anti- EGFR therapies with other inhibitors, so that resistance can be
overcome by the targeting of alternate signaling pathways. On the other hand,
components of the inflammatory pathway, within and around a tumor, provide a
conducive environment for tumor growth by supplying numerous cytokines and
chemokines that foster carcinogenesis. Interleukin 6 (IL-6) is one such cytokine
that is found to be associated with inflammation-driven cancers and which also
plays a crucial role in acquired resistance to anti-EGFR drugs. The EGFR and
IL-6 signaling pathways crosstalk in multiple ways, through various mediators
and downstream signaling pathways driving resistance and hence co-targeting them
has potential for future cancer treatments. Here we provide an overview on the
crosstalk between the EGFR and IL-6 pathways, and discuss how co-targeting these
two pathways could be a promising combination therapy of the future. |
What is the role of STAG1/STAG2 proteins in differentiation? | STAG1/STAG2 proteins are tumour suppressor proteins that suppress cell proliferation and are essential for differentiation. | A mutant version of p53 (p53-121F), in which phenylalanine replaces the 121st
serine residue, can induce apoptosis more effectively than wild-type p53
(wt-p53). In view of this observation, we considered that one or more
apoptosis-related p53-target genes might be preferentially induced by p53-121F.
We carried out cDNA microarray analysis to identify such genes, using mRNAs
isolated from LS174T colon-cancer cells infected by adenovirus vectors
containing either p53-121F (Ad-p53-121F) or wt-p53 (Ad-p53). The STAG1 gene was
one of the transcripts showing higher expression levels in cells infected with
Ad-p53-121F as opposed to Ad-wtp53. The encoded product appears to contain a
transmembrane domain, and binding motifs for SH3 and WW. In two other cancer
cell lines, the expression of STAG1 mRNA was induced in response to various
genotoxic stresses in a p53-dependent manner; moreover, enforced expression of
STAG1 led to apoptosis in several additional cancer cell lines. Suppression of
endogenous STAG1 using the RNA-interference method reduced the apoptotic
response, whether induced by Ad-p53-121F or Ad-p53. These results suggest that
STAG1, a novel transcriptional target for p53, mediates p53-dependent apoptosis,
and might be a good candidate for next-generation gene therapy. Sonic hedgehog (Shh) is necessary for sustaining the proliferation of neural
stem cells (NSCs), yet little is known about its mechanisms. Whereas Gli1, Gli2,
and Gli3, the primary mediators of Shh signaling, were all expressed in
hippocampal neural progenitors, Shh treatment of NSCs induced only Gli1
expression. Acute depletion of Gli1 in postnatal NSCs by short-hairpin RNA
decreased proliferation, whereas germline deletion of Gli1 did not affect NSC
proliferation, suggesting a difference in mechanisms of Gli1 compensation that
may be developmentally dependent. To determine whether Gli1 was sufficient to
enhance NSC proliferation, we overexpressed this mitogen and were surprised to
find that Gli1 resulted in decreased proliferation, accumulation of NSCs in the
G2/M phase of cell cycle, and apoptosis. In contrast, Gli1-expressing
lineage-restricted neural precursors demonstrated a 4.5-fold proliferation
enhancement. Expression analyses of Gli1-expressing NSCs identified significant
induction of Gadd45a and decreased cyclin A2 and Stag1 mRNA, genes involved in
the G2-M transition and apoptosis. Furthermore, Gadd45a overexpression was
sufficient to partially recapitulate the Gli1-induced G2/M accumulation and cell
death of NSCs. In contrast to normal stem cells, tumor-derived stem cells had
markedly higher basal Gli1 expression and did not undergo apoptosis with further
elevation of Gli1. Our data suggest that Gli1-induced apoptosis may serve as a
protective mechanism against premature mitosis and may give insight into
mechanisms by which nonmaligt stem cells restrain hyperproliferation in the
context of potentially transforming mitogenic signals. Tumor-derived stem cells
apparently lack these mechanisms, which may contribute to their unrestrained
proliferation and maligt potential. Replicated sister chromatids are held together until mitosis by cohesin, a
conserved multisubunit complex comprised of Smc1, Smc3, Scc1, and Scc3, which in
vertebrate cells exists as two closely related homologues (SA1 and SA2). Here,
we show that cohesin(SA1) and cohesin(SA2) are differentially required for
telomere and centromere cohesion, respectively. Cells deficient in SA1 are
unable to establish or maintain cohesion between sister telomeres after DNA
replication in S phase. The same phenotype is observed upon depletion of the
telomeric protein TIN2. In contrast, in SA2-depleted cells telomere cohesion is
normal, but centromere cohesion is prematurely lost. We demonstrate that loss of
telomere cohesion has dramatic consequences on chromosome morphology and
function. In the absence of sister telomere cohesion, cells are unable to repair
chromatid breaks and suffer sister telomere loss. Our studies elucidate the
functional distinction between the Scc3 homologues in human cells and further
reveal an essential role for sister telomere cohesion in genomic integrity. Author information:
(1)School of Bioscience and Engineering, South China University of Technology,
Guangzhou, Guangdong 510006, P.R. China Physics Department, North Carolina State
University, Raleigh, North Carolina, NC 27695, USA.
(2)Physics Department, North Carolina State University, Raleigh, North Carolina,
NC 27695, USA.
(3)Department of BioSciences, Rice University, Houston, TX 77005, USA Institute
of Microbiology and College of Life Sciences, Zhejiang University, Hangzhou,
Zhejiang 310058, P.R. China.
(4)Department of BioSciences, Rice University, Houston, TX 77005, USA.
(5)Department of Neurology, The First Affiliated Hospital of Zhengzhou
University, Zhengzhou, He 450014, P.R. China.
(6)Biomanufacturing Training and Education Center, North Carolina State
University, Raleigh, North Carolina, NC 27695, USA.
(7)Division of Biophysics, Universität Osnabrück, Barbarstrasse 11, 49076
Osnabrück, Germany.
(8)School of Biosciences, University of Kent, Canterbury, Kent CT2 7NJ, UK.
(9)Department of Environmental and Occupational Health, University of
Pittsburgh, PA 15213, USA.
(10)Kimmel Center for Biology and Medicine at the Skirball Institute, Department
of Pathology, New York University School of Medicine, New York, NY 10016, USA.
(11)Physics Department, North Carolina State University, Raleigh, North
Carolina, NC 27695, USA [email protected]. |
What can we measure with the TSA-Seq method? | Mapping 3D genome organization relative to nuclear compartments using TSA-Seq as a cytological ruler. TSA-Seq, a new mapping method capable of providing a "cytological ruler" for estimating mean chromosomal distances from nuclear speckles genome-wide and for predicting several Mbp chromosome trajectories between nuclear compartments without sophisticated computational modeling. | While nuclear compartmentalization is an essential feature of three-dimensional
genome organization, no genomic method exists for measuring chromosome distances
to defined nuclear structures. In this study, we describe TSA-Seq, a new mapping
method capable of providing a "cytological ruler" for estimating mean
chromosomal distances from nuclear speckles genome-wide and for predicting
several Mbp chromosome trajectories between nuclear compartments without
sophisticated computational modeling. Ensemble-averaged results in K562 cells
reveal a clear nuclear lamina to speckle axis correlated with a striking spatial
gradient in genome activity. This gradient represents a convolution of multiple
spatially separated nuclear domains including two types of transcription "hot
zones." Transcription hot zones protruding furthest into the nuclear interior
and positioning deterministically very close to nuclear speckles have higher
numbers of total genes, the most highly expressed genes, housekeeping genes,
genes with low transcriptional pausing, and super-enhancers. Our results
demonstrate the capability of TSA-Seq for genome-wide mapping of nuclear
structure and suggest a new model for spatial organization of transcription and
gene expression. |
Which method is behind HipMCL? | HipMCL is a high-performance parallel implementation of the Markov clustering algorithm for large-scale networks. Despite its popularity, MCL's scalability to cluster large datasets still remains a bottleneck due to high running times and memory demands. | Biological networks capture structural or functional properties of relevant
entities such as molecules, proteins or genes. Characteristic examples are gene
expression networks or protein-protein interaction networks, which hold
information about functional affinities or structural similarities. Such
networks have been expanding in size due to increasing scale and abundance of
biological data. While various clustering algorithms have been proposed to find
highly connected regions, Markov Clustering (MCL) has been one of the most
successful approaches to cluster sequence similarity or expression networks.
Despite its popularity, MCL's scalability to cluster large datasets still
remains a bottleneck due to high running times and memory demands. Here, we
present High-performance MCL (HipMCL), a parallel implementation of the original
MCL algorithm that can run on distributed-memory computers. We show that HipMCL
can efficiently utilize 2000 compute nodes and cluster a network of ∼70 million
nodes with ∼68 billion edges in ∼2.4 h. By exploiting distributed-memory
environments, HipMCL clusters large-scale networks several orders of magnitude
faster than MCL and enables clustering of even bigger networks. HipMCL is based
on MPI and OpenMP and is freely available under a modified BSD license. |
What is the function of CR elements in B-cells? | After addition of culture supernatant from BCG-activated macrophages CR- B cells cooperate with both unprimed and primed T helper cells. | B cells that carry the complement receptor (CR+) were separated from B cells
that lack the complement receptor (CR-) by velocity sedimentation or by passage
through C-coated Sephadex columns. The kinetics of responses to bacterial
lipopolysaccharide (LPS) in both B cell subpopulations were determined in three
assay procedures: 1) incorporation of radioactive thymidine into DNA; 2)
incorporation of radioactive leucine into immunoglobulin; 3) enumeration of
cells forming polyclonal antibody to the 2,4,6-trinitrophenyl hapten. Although
both subpopulations of B cells responded to LPS, they differed in the time
course. CR- B cells responded with a delay of approximately 24 hr as compared
with the response of CR+ B cells. The implications to the ontogenetic status of
CR+ and CR- B subpopulations are discussed. A Sephadex G-10 column coated with antigen-antibody complexes and complement
retains complement receptor-bearing (CR+) mouse spleen cells. The effluent is
rich in thymus-derived cells (T cells), and contains bone marrow-derived cells
(B cells) which carry surface immunoglobulin (Ig), Ir-associated antigen (Ia),
and Fc receptors, but no complement receptors (CR-). Although both
unfractionated and CR- B cell populations are capable of producing antibody to
red cell antigens, they differ in their requirements for the initiation of the
response. Unfractionated B cells cooperate with primed as well as unprimed
helper T cells; macrophages are required for this cooperation but can be
replaced by 2-mercaptoethanol. CR- B cells cooperate with primed but not with
unprimed T cells provided macrophages are added to cultures. After addition of
culture supernatant from BCG-activated macrophages CR- B cells cooperate with
both unprimed and primed T helper cells. |
What is drug target for olaparib? | Olaparib(Lynparza) is a PARP inhibitor, inhibiting poly ADP ribose polymerase (PARP), an enzyme involved in DNA repair. | The poor prognosis for patients with esophagogastric cancers (EGC) has resulted
in an increased focus on the use of targeted agents in this disease. Targets
include epidermal growth factor receptor (EGFR), vascular endothelial growth
factor (VEGF), Her2, mammalian target of rapamycin (mTOR), MET, poly
(ADP-ribose) polymerase (PARP) and claudin 18.2 (CLDN18.2). Trastuzumab, an
anti-Her2 antibody, was approved by the U.S. FDA in 2010 as first-line therapy
in combination with chemotherapy for Her2-positive disease. Since then,
strategies targeting Her2 that have been successful in Her2-positive breast
cancer, have failed in EGC. The one remaining study, the phase III Jacob study
with pertuzumab, has yet to be presented. The anti-VEGF receptor 2 antibody,
ramucirumab has been investigated as second-line therapy in 2 phase III trials,
which resulted in improved survival, with subsequent FDA approval of ramucirumab
in the second-line setting. Therapies targeting EGFR have been evaluated in a
number of phase III studies, all of which have been negative. Phase III
investigation of an mTOR inhibitor did not improve survival, although biomarker
studies are awaited which may identify subgroups of patients that may benefit
from its use. The results of the trials targeting MET in EGC have been
disappointing, raising doubts about the usefulness of further testing agents
that inhibit the MET pathway. PARP inhibition with olaparib, warrants further
investigation, possibly in combination with other targeted therapies or immune
checkpoint inhibition and in a biomarker-selected population. The identification
of CLDN18.2 and its targeting with claudiximab is very promising and will be
further investigated in a phase III study. BACKGROUND: Poly (ADP-ribose) polymerase inhibitors (PARPi) prevent
single-stranded DNA repair. Olaparib is a PARPi approved for the treatment of
BRCA mutant ovarian and breast carcinoma. Emerging clinical data suggest a
benefit of combining olaparib with immunotherapy in prostate cancer patients
both with and without somatic BRCA mutations.
METHODS: We examined if olaparib, when combined with IgG1 antibody-dependent
cellular cytotoxicity (ADCC)-mediating monoclonal antibodies (mAbs) cetuximab
(anti-EGFR), or avelumab (anti-PD-L1), would increase tumor cell sensitivity to
killing by natural killer (NK) cells independently of BRCA status or mAb target
upregulation. BRCA mutant and BRCA wildtype (WT) prostate carcinoma cell lines
were pretreated with olaparib and then exposed to NK cells in the presence or
absence of cetuximab or avelumab.
RESULTS: NK-mediated killing was significantly increased in both cell lines and
was further increased using the ADCC-mediating mAbs. Pre-exposure of NK cells to
recombit IL-15/IL-15Rα further increased the lysis of olaparib treated tumor
cells. In addition, olaparib treated tumor cells were killed to a significantly
greater degree by engineered high-affinity NK cells (haNK). We show here for the
first time that (a) olaparib significantly increased tumor cell sensitivity to
NK killing and ADCC in both BRCA WT and BRCA mutant prostate carcinoma cells,
independent of PD-L1 or EGFR modulation; (b) mechanistically, treatment with
olaparib upregulated death receptor TRAIL-R2; and (c) olaparib significantly
enhanced NK killing of additional tumor types, including breast, non-small cell
lung carcinoma, and chordoma.
CONCLUSIONS: These studies support the combined use of NK- and ADCC-mediating
agents with correctly timed PARP inhibition. |
What virus is the Gardisil vaccine used for? | Gardisil is a quadrivalent HPV vaccine would have been useful in the prevention of infections with human papillomavirus. | |
Which cells mature in the human thymus? | Thymus progenitor cells mature in the human thymus through differentiation into cardiomyocytes and fibroblasts. | The finding of peanut agglutinin (PNA) binding capacity, supposed to be a marker
of immature lymphocytes, within the T-cell population of the human thymus (58%)
and tonsil (10%) prompted the comparison of maturation stages of PNA binding
(PNA+) and nonbinding (PNA-) T cells in both organs. The proliferative response
after mitogenic stimulation of purified PNA+ fractions was significantly less
than that of purified PNA- fractions. The results of mitogen dose-response
experiments, of variation in time of culture harvest, and of addition of
irradiated allogeneic peripheral blood non-T cells indicated the intrinsic
mitogen unresponsiveness of cells in the PNA+ fractions. The mitogen response of
tonsil fractions was higher than that of thymocyte fractions. Cells with an
immature immunologic phenotype were enriched in the thymocyte PNA+ fraction, and
almost absent in the tonsil fractions. Both tonsil fractions contained cells
with the immunologic phenotype of mature T cells, and showed a purine
interconversion enzyme makeup comparable to mature T lymphocytes. It is
concluded that the tonsil PNA+ T cell is a functionally immature lymphocyte
which is in a further maturation stage than PNA+ or PNA- thymocytes. The
presence of PNA+ T cells outside the thymus is of relevance for the clinical
evaluation of PNA binding assays and suggests the occurrence of T-cell
maturation within the tonsil environment. CD4+CD8+ double-positive (DP) T cells represent a minor subpopulation of T
lymphocytes found in the periphery of adult rats. In this study, we show that
peripheral DP T cells appear among the first T cells that colonize the
peripheral lymphoid organs during fetal life, and represent approximately 40% of
peripheral T cells during the perinatal period. Later their proportion decreases
to reach the low values seen in adulthood. Most DP T cells are small size
lymphocytes that do not exhibit an activated phenotype, and their proliferative
rate is similar to that of the other peripheral T cell subpopulations. Only
30-40% of DP T cells expresses CD8beta chain, the remaining cells expressing
CD8alphaalpha homodimers. However, both DP T cell subsets have an intrathymic
origin since they appear in the recent thymic emigrant population after
injection of FITC intrathymically. Functionally, although DP T cells are
resistant to undergo apoptosis in response to glucocorticoids, they show poor
proliferative responses upon CD3/TCR stimulation due to their inability to
produce IL-2. A fraction of DP T cells are not actively synthesizing the CD8
coreceptor, and they gradually differentiate to the CD4 cell lineage in
reaggregation cultures. Transfer of DP T lymphocytes into thymectomized SCID
mice demonstrates that these cells undergo post-thymic maturation in the
peripheral lymphoid organs and that their CD4 cell progeny is fully
immunocompetent, as judged by its ability to survive and expand in peripheral
lymphoid organs, to proliferate in response to CD3 ligation, and to produce IL-2
upon stimulation. There is evidence for both physiological functions of the natural
neurotransmitter, acetylcholine, and pharmacological actions of the plant
alkaloid, nicotine, on the development and function of the immune system. The
effects of continuous exposure to nicotine over a 12-day course of fetal thymus
organ culture (FTOC) were studied, and thymocytes were analyzed by flow
cytometry. In the presence of very low concentrations of nicotine many more
immature T cells (defined by low or negative TCR expression) and fewer mature T
cells (intermediate or high expression of TCR) were produced. In addition, the
numbers of cells expressing CD69 and, to a lesser extent, CD95 (Fas) were
increased. These effects took place when fetal thymus lobes from younger (13-14
days gestation) pups were used for FTOC. If FTOC were set up using tissue from
older (15-16 days gestation pups), nicotine had little effect, suggesting that
it may act only on immature T cell precursors. Consistent with an increase in
immature cells, the expression of recombinase-activating genes was found to be
elevated. Nicotine effects were partially blocked by the simultaneous addition
of the nicotinic antagonist d-tubocurarine. Furthermore, d-tubocurarine alone
blocked the development of both immature and mature murine thymocytes,
suggesting the presence of an endogenous ligand that may engage nicotinic
acetylcholine receptors on developing thymocytes and influence the course of
normal thymic ontogeny. Transgenic mice expressing the E7 protein of HPV16 from the keratin 14 promoter
demonstrate increasing thymic hypertrophy with age. This hypertrophy is
associated with increased absolute numbers of all thymocyte types, and with
increased cortical and medullary cellularity. In the thymic medulla, increased
compartmentalization of the major thymic stromal cell types and expansion of
thymic epithelial cell population is observed. Neither an increased rate of
immature thymocyte division nor a decreased rate of immature thymocyte death was
able to account for the observed hypertrophy. Thymocytes with reduced levels of
expression of CD4 and/or CD8 were more abundant in transgenic (tg) mice and
became increasingly more so with age. These thymic SP and DP populations with
reduced levels of CD4 and/or CD8 markers had a lower rate of apoptosis in the tg
than in the non-tg mice. The rate of export of mature thymocytes to peripheral
lymphoid organs was less in tg animals relative to the pool of available mature
cells, particularly for the increasingly abundant CD4lo population. We therefore
suggest that mature thymocytes that would normally die in the thymus gradually
accumulated in E7 transgenic animals, perhaps as a consequence of exposure to a
hypertrophied E7-expressing thymic epithelium or to factors secreted by this
expanded thymic stromal cell population. The K14E7 transgenic mouse thus
provides a unique model to study effects of the thymic epithelial cell
compartment on thymus development and involution. BACKGROUND: Generation of functional (CD4+)(CD8-)CD25+ regulatory T cells (Treg)
in the murine thymus depends on FoxP3. Removal of the thymus from neonatal mice
has been shown to result in a multiple organ autoimmune disease phenotype that
can be prevented by introducing the FoxP3+ Treg population to the animal. It has
therefore, been proposed that functional FoxP3+ Treg cells are not made in the
neonatal thymus; however, it remains unclear when and where functional
(FoxP3+)(CD4+)(CD8-)CD25+ thymocytes are generated in postnatal thymus.
RESULTS: We report that neither FoxP3 mRNA nor protein is expressed in
(CD4+)(CD8-)CD25+, or (CD4+)(CD8-)CD25- thymocytes until 3-4 days post birth,
despite the presence of mature (CD4+)(CD8-)CD25+/- thymocytes in the thymus by
1-2 days after birth. (FoxP3-)(CD4+)(CD8-)CD25+ thymocytes from day 2 newborn
mice show no Treg activity. Interestingly, we are able to detect low numbers of
FoxP3+ thymocytes dispersed throughout the medullary region of the thymus as
early as 3-4 days post birth. Expression of FoxP3 is induced in embryonic day 17
fetal thymus organ culture (FTOC) after 4-6 days of in vitro culture. Treatment
of FTOCs with thymic stromal derived lymphopoietin (TSLP) enhanced expression of
FoxP3, and blocking the TSLP receptor reduces FoxP3 expression in FTOC.
Furthermore, TSLP stimulates FoxP3 expression in purified (CD4+)CD8- thymocytes,
but not in (CD4+)CD8+, (CD4-)CD8+ and (CD4-)CD8- thymocytes.
CONCLUSION: Expression of FoxP3 or Treg maturation is ontogenically distinct and
kinetically delayed from the generation of (CD4+)(CD8-)CD25+ or
(CD4+)(CD8-)CD25- thymocytes in the postnatal thymus. TSLP produced from
medullary thymic epithelia cells (mTEC) contributes to the expression of FoxP3
and the maturation of natural regulatory T cells. Overall, these results suggest
that the development of Treg cells requires paracrine signaling during late
stages of thymocyte maturation that is distinct from signaling during positive
or negative selection. Regain of immunocompetence after myeloablation and bone marrow cell (BMC)
reconstitution essentially depends on T progenitor homing into the thymus and
intrathymic T cell maturation. CD44 facilitates progenitor homing and settlement
in the bone marrow and is known as a T progenitor marker. In search for
improving regain of immunocompetence after BMC reconstitution, we explored
whether the CD44 standard (CD44 s) and/or variant isoforms CD44v6 and CD44v7
contribute to thymus repopulation and thymocyte maturation. Antibody-blocking
studies and cells/mice with a targeted deletion of CD44v6/7 or CD44v7 revealed
that CD44s, but not CD44v6 and CD44v7, has a major impact on progenitor cell
homing into the thymus. Instead, CD44v6 strengthens apoptosis resistance and
expansion of early thymocytes. CD44v6-induced apoptosis resistance, most strong
in double-negative (DN) thymocytes, is accompanied by Akt activation.
CD44v6-induced proliferation of DN cells proceeds via activation of the MAPK
pathway. At later stages of T cell maturation, CD44 acts as an accessory
molecule, initiating and supporting TCR/CD3 complex-mediated signal transduction
in double-positive and single-positive thymocytes. Thus, CD44 plays a major role
in thymus homing. In addition, CD44v6 is important for survival and expansion of
early thymocytes. These findings suggest that strengthening CD44v6 expression on
lymphoid progenitors could well contribute to accelerated regain of
immunocompetence. In the thymus, a T-cell repertoire able to confer protection against infectious
and noninfectious agents in a peptide-dependent, self-MHC-restricted manner is
selected. Direct detection of Ag-specific thymocytes, and analysis of the impact
of the expression of the MHC-restricting allele on their frequency or function
has never been studied in humans because of the extremely low precursor
frequency. Here, we used a tetramer-based enrichment protocol to analyze the ex
vivo frequency and activation-phenotype of human thymocytes specific for self,
viral and tumor-antigens presented by HLA-A*0201 (A2) in individuals expressing
or not this allele. Ag-specific thymocytes were quantified within both CD4CD8
double or single-positive compartments in every donor. Our data indicate that
the maturation efficiency of Ag-specific thymocytes is poorly affected by HLA-A2
expression, in terms of frequencies. Nevertheless, A2-restricted T-cell lines
from A2(+) donors reacted to A2(+) cell lines in a highly peptide-specific
fashion, whereas their alloreactive counterparts showed off-target activity.
This first ex vivo analysis of human antigen-specific thymocytes at different
stages of human T-cell development should open new perspectives in the
understanding of the human thymic selection process. Positive selection occurs in the thymic cortex, but critical maturation events
occur later in the medulla. Here we defined the precise stage at which T cells
acquired competence to proliferate and emigrate. Transcriptome analysis of late
gene changes suggested roles for the transcription factor NF-κB and interferon
signaling. Mice lacking the inhibitor of NF-κB (IκB) kinase (IKK) kinase TAK1
underwent normal positive selection but exhibited a specific block in functional
maturation. NF-κB signaling provided protection from death mediated by the
cytokine TNF and was required for proliferation and emigration. The interferon
signature was independent of NF-κB; however, thymocytes deficient in the
interferon-α (IFN-α) receptor IFN-αR showed reduced expression of the
transcription factor STAT1 and phenotypic abnormality but were able to
proliferate. Thus, both NF-κB and tonic interferon signals are involved in the
final maturation of thymocytes into naive T cells. The mRNA for dopamine receptors of type D1, D3, D5, but not type D2, was
detected in the thymus of rats starting from day 16 of embryonic development
(E16). Dopamine at concentrations of 10-8-10‒6 M inhibited fetus thymocyte
response to mitogen, confirming the functionality of the receptors and the
possibility of a direct effect of dopamine on the developing thymus.
Pharmacological inhibition of catecholamine synthesis in the crucial period of
thymus development leads to long-term changes in the T-system immunity due to
increased production of natural regulatory T-lymphocytes. The presence and
functional activity of dopamine receptors in the fetal thymus indicates its
ability to influence the development of the immune system of rats during
ontogeny. |
What are the eRNA-producing centers (EPCs)? | Active enhancers in mammals produce enhancer RNAs (eRNAs) that are bidirectionally transcribed, unspliced, and unstable. Enhancer regions are also enriched with long noncoding RNA (lncRNA) transcripts, which are typically spliced and substantially more stable. DNase hypersensitive sites with evidence of bidirectional transcription are called eRNA-producing centers (EPCs). EPCs found very close to transcription start sites of lncRNAs exhibit attributes of both enhancers and promoters, including distinctive DNA motifs and a characteristic chromatin landscape. These EPCs are associated with higher enhancer activity, driven at least in part by the presence of conserved, directional splicing signals that promote lncRNA production, pointing at a causal role of lncRNA processing in enhancer activity. | Active enhancers in mammals produce enhancer RNAs (eRNAs) that are
bidirectionally transcribed, unspliced, and unstable. Enhancer regions are also
enriched with long noncoding RNA (lncRNA) transcripts, which are typically
spliced and substantially more stable. In order to explore the relationship
between these two classes of RNAs, we analyzed DNase hypersensitive sites with
evidence of bidirectional transcription, which we termed eRNA-producing centers
(EPCs). EPCs found very close to transcription start sites of lncRNAs exhibit
attributes of both enhancers and promoters, including distinctive DNA motifs and
a characteristic chromatin landscape. These EPCs are associated with higher
enhancer activity, driven at least in part by the presence of conserved,
directional splicing signals that promote lncRNA production, pointing at a
causal role of lncRNA processing in enhancer activity. Together, our results
suggest that the conserved ability of some enhancers to produce lncRNAs augments
their activity in a manner likely mediated through lncRNA maturation. |
Describe Brain Radiation Information Data Exchange (BRIDE) approach | BRIDE (Brain Radiation Information Data Exchange) is a data integration platform that acts as a knowledge broker for LDIR researchers to facilitate molecular research on the systems biology of LDIR response in mammals. Its flexible design can capture a range of experimental information for genomics, epigenomics, transcriptomics, and proteomics. | Author information:
(1)Department of Genetics, Development & Molecular Biology, School of Biology,
Aristotle University of Thessalonica, 54124, Thessalonica, Greece.
(2)Institute of Radiation Biology, Helmholtz Zentrum München, German Research
Center for Environmental Health GmbH, 85764, Neuherberg, Germany.
(3)Present address: Department of Biochemistry and Molecular Biology, University
of Southern Denmark, Campusvej 55, 5230, Odense M, Denmark.
(4)Radiobiology Unit, Belgian Nuclear Research Centre (SCK•CEN), B-2400, Mol,
Belgium.
(5)School of Medicine, IISPV, "Rovira i Virgili" University, Sant Llorens 21,
43201, Reus, Spain.
(6)Laboratory of Radiation Biology & Biomedicine, Agenzia Nazionale per le Nuove
Tecnologie, l'Energia e lo Sviluppo Economico Sostenibile (ENEA) Centro Ricerche
Casaccia, 00123, Rome, Italy.
(7)National Research Center for Radiation Medicine of the National Academy of
Medical Sciences of Ukraine, Melnykov str. 53, Kyiv, 04050, Ukraine.
(8)Erasmus Medical Center, 3015GE, Rotterdam, The Netherlands.
(9)Institute of Radiation Biology, Helmholtz Zentrum München, German Research
Center for Environmental Health GmbH, 85764, Neuherberg, Germany.
[email protected].
(10)Radiobiology Unit, Belgian Nuclear Research Centre (SCK•CEN), B-2400, Mol,
Belgium. [email protected].
(11)Department of Genetics, Development & Molecular Biology, School of Biology,
Aristotle University of Thessalonica, 54124, Thessalonica, Greece.
[email protected].
(12)Biological Process & Computation Laboratory (BCPL), Chemical Process &
Energy Resources Institute (CPERI), Centre for Research & Technology Hellas
(CERTH), Thessalonica, 57001, Greece. [email protected]. |
When was vivotif first licenced in Europe? | The vaccine vivotif was first licensed in Europe in 1983. | Cases of diarrhoeal disease number from 1.7 to 5 billion per year worldwide. One
of the main causes of diarrhoeal disease is typhoid fever, which is a
potentially life-threatening multi-systemic illness. According to the most
recent estimates, a total of 26.9 million typhoid fever episodes occurred in
2010. The geographical distribution of the disease differs widely; in developed
countries, the incidence rate per 100,000 per year varies from < 0.1 to 0.3, and
the disease mainly affects people who travel to endemic areas located in low-
and middle-income countries. Low- and middle-income countries are mainly
affected owing to the lack of clean water and proper sanitation. In the fight
against this plague, prevention is fundamental, and vaccination against typhoid
is an effective measure. Vivotif® is an oral live attenuated vaccine which
contains a mutated strain of Salmonella (Ty21a) and reproduces the natural
infection. The vaccine was first licensed in Europe in 1983 and in the US in
1989, and over the years it has proved efficacious and safe. It is indicated for
adults and children from 5 years of age upwards. Specifically, in the most
developed countries, vaccination is suggested for highrisk population groups and
particularly for international travellers to destinations where the risk of
contracting typhoid fever is high. It must also be borne in mind that
international travel is increasing. Indeed, international tourist arrivals
totalled 1,184 million in 2015 and, on the basis of current trends,
international travel is expected to grow by 3-4% in 2017. Vivotif® appears to be
a powerful means of disease prevention, the importance of which is highlighted
by the spread of antibiotic-resistant strains of Salmonella typhy (S. typhi). |
What is herceptin? | Herceptin is a second generation tyrosine kinase inhibitor, that serves as an effective and approved oral therapy for patients with HER2-positive breast cancer. | BACKGROUND & OBJECTIVE: Herceptin is a humanized monoclonal antibody for
treating the patients with metastatic breast cancers overexpressing human
epidermal growth factor receptor (HER-2). Herceptin combined with doxorubicin
markedly increased the incidence of cardiac dysfunction in clinical trials. The
current study was designed to investigate the cardiotoxicity aggravated by
Herceptin combined with doxorubicin in rats.
METHODS: Thirty-six female rats were randomized into four groups (n=9): control
group, Herceptin group, doxorubicin group, Herceptin combined with doxorubicin
group. The value of malon dialdehyde (MDA) was measured by thiobarbituric acid
(TBA) method and the activity of glutathione peroxidase (GSH-Px) was measured by
5,5'-dithiobis-2-nitrobenzoic acid (DTNB) method. The damage and apoptosis of
myocardium were observed by electron microscopy and tdt-mediated dUTP nick end
labeling (TUNEL). The ratio of myocardial apoptosis cells was quantified using
flow cytometry.
RESULTS: The value of MDA in control group and Herceptin group were
(0.70+/-0.03) and (0.73+/-0.05) nmol x(mg x protein)(-1), respectively; the
activity of GSH-PX in control group and Herceptin group were (81.31+/-0.13) and
(78.43+/-0.15) U x(mg x protein)(-1),respectively;the two groups were similar.
The value of MDA in doxorubicin group and Herceptin combined with doxorubicin
group were (0.88+/-0.10) and (0.94+/-0.13) nmol x(mg x protein)(-1),
respectively; the activity of GSH-PX in doxorubicin group and Herceptin combined
with doxorubicin group were(67.88+/-0.24) and(63.37+/-0.28) U x(mg x
protein)(-1),respectively. The most prominent ultrastructural damages were
mitochondrial swelling with fragmentation of cristae and vacuoles formation in
doxorubicin group and Herceptin combined with doxorubicin group. Herceptin
markedly increased the ratio of apoptosis of myocardial cells from (5.35+/-0.27)
% in doxorubicin group to (8.27+/-0.38) % in Herceptin combined with doxorubicin
group as measured by flow cytometry (P< 0.05).
CONCLUSION: Herceptin alone had little toxic effects on rat cardiomocytes, but
Herceptin combined with doxorubicin can aggravate rat cardiotoxicity. Herceptin
can also increase the number of the apoptosis cell in rat myocardium. Her2 (erbB2/neu) is overexpressed in 25-30% of human breast cancers. Herceptin
is a recombit humanized Her2 antibody used to treat breast cancer patients
with Her2 overexpression. Over a 5-month selection process, we isolated clones
of BT474 (BT) human breast carcinoma cells (BT/Her(R)) that were resistant to
Herceptin in vitro. In BT/Her(R) subclones, cell-surface, phosphorylated and
total cellular Her2 protein remained high in the continuous presence of
Herceptin. Likewise, the levels of cell-surface, phosphorylated, and total
cellular Her3 and EGFR were either unchanged or only slightly elevated in
BT/Her(R) subclones relative to BT cells. One BT/Her(R) subclone had
substantially upregulated cell-surface EGFR, but this did not correlate with a
higher relative resistance to Herceptin. In looking at the downstream PI-3K/Akt
signaling pathway, phosphorylated and total Akt levels and Akt kinase activities
were all sustained in BT/Her(R) subclones in the presence of Herceptin, but
significantly downregulated in BT cells exposed to Herceptin. Whereas BT cells
lost sensitivity to the PI-3K inhibitor LY294002 in the presence of Herceptin,
BT/Her(R) subclones were equally sensitive to this agent in the presence and
absence of Herceptin. This suggests that BT/Her(R) subclones acquired a
Herceptin-resistant mechanism of PI-3K signaling. BT/Her(R) subclones were also
sensitive to the EGFR kinase inhibitor AG1478 in the presence of Herceptin, to
the same extent as BT cells. The BT/Her(R) subclones provide new insights into
mechanisms of Herceptin resistance and suggest new treatment strategies in
combination with other inhibitors targeted to signal transduction pathways. BACKGROUND: Herceptin (trastuzumab) is a humanized monoclonal antibody that is
approved for the treatment of metastatic breast cancer patients whose tumors
overexpress Her2 (erbB2/neu). Up to 70% of Her2-positive breast cancers
demonstrate a response to Herceptin-based therapies, but resistance almost
inevitably arises within a year of the initial response. To help understand the
mechanism of Herceptin resistance, we isolated clonal variants of Her2-positive
BT474 human breast cancer cells (BT/Her(R)) that are highly resistant to
Herceptin. These cell lines exhibit sustained PI3K/Akt signaling as an essential
component of Herceptin-resistant proliferation. Several genes in the protein
kinase A (PKA) signaling network have altered expression in BT/Her(R) cells,
including PPP1R1B, which encodes a 32 kDa protein known as Darpp-32 and its
amino-terminal truncated variant, t-Darpp. The purpose of the current work was
to determine the role of Darpp-32 and t-Darpp in Herceptin resistance.
METHODOLOGY AND RESULTS: We determined expression of Darpp-32 and t-Darpp in
BT/Her(R) cells selected for resistance to Herceptin. Subsequently, cDNAs
encoding the two isoforms of Darpp-32 were transfected, separately and together,
into Her2-positive SK-Br-3 breast cancer cells. Transfected cells were tested
for resistance to Herceptin and Herceptin-mediated dephosphorylation of Akt. DNA
binding activity by the cAMP response element binding protein (CREB) was also
measured. We found that BT/Her(R) cells overexpressed t-Darpp but not Darpp-32.
Moreover, t-Darpp overexpression in SK-Br-3 cells was sufficient for conferring
resistance to Herceptin and Herceptin-mediated dephosphorylation of Akt.
Darpp-32 co-expression reversed t-Darpp's effects on Herceptin resistance and
Akt phosphorylation. t-Darpp overexpression led to increased CREB binding
activity, which was also reversible by Darpp-32.
CONCLUSIONS: t-Darpp and Darpp-32 appear to have antagonistic effects on
Herceptin resistance. We present a unified model by which these effects might be
mediated via the PKA regulatory network. BACKGROUND: Trastuzumab (Herceptin(®)) is a humanized monoclonal antibody
targeting the human epidermal growth factor receptor 2 (HER2) and is used in the
treatment of HER2-overexpressing breast and gastric cancer. FTMB is being
developed as a biosimilar of trastuzumab.
OBJECTIVE: In this combined dose-escalation and bioequivalence study of parallel
design, the pharmacokinetic profile of FTMB was compared with Herceptin(®).
METHODS: Healthy male volunteers received single doses of 0.5, 1.5, 3.0 or 6.0
mg/kg FTMB, or placebo, in consecutive dose-escalation cohorts to assess the
safety profile. Thereafter, the 6 mg/kg cohort was expanded to establish
bioequivalence between FTMB (Test) and Herceptin(®) (Reference) based on an
acceptance interval of 80.0-125.0 %. In total, 118 subjects were enrolled in the
study.
RESULT: The mean area under the concentration-time curve from time zero to
infinity (AUC∞) was 1,609 µg·day/mL (Test) and 1,330 µg·day/mL (Reference). The
log-transformed geometric mean Test/Reference (T/R) ratio for AUC∞ was 89.6 %
(90 % confidence interval [CI] 85.1-94.4), demonstrating bioequivalence. For the
secondary endpoint, the maximum concentration observed (Cmax), the geometric
mean T/R ratio was 89.4 % (90 % CI 83.4-95.9). Non-linear, target-mediated
pharmacokinetics were also observed. Adverse events other than the documented
side effects of Herceptin(®) (fever, influenza-like illness, and fatigue) did
not occur. No signs of cardiotoxicity were observed.
CONCLUSIONS: This bioequivalence study with a trastuzumab biosimilar in healthy
male volunteers demonstrated bioequivalence of FTMB with Herceptin(®). FTMB was
well tolerated in doses up to 6 mg/kg. Non-linear target elimination was also
observed in the pharmacokinetic profile of trastuzumab. Optimal adoptive cell therapy (ACT) should contribute to effective cancer
treatment. The unique ability of natural killer (NK) cells to kill cancer cells
independent of major histocompatibility requirement makes them suitable as ACT
tools. Herceptin, an antihuman epidermal growth factor receptor-2 (anti-HER2)
monoclonal antibody, is used to treat HER2+ breast cancer. However, it has
limited effectiveness and possible severe cardiotoxicity. Given that Herceptin
may increase the cytotoxicity of lymphocytes, we explored the possible
augmentation of NK cell cytotoxicity against HER2+ breast cancer cells by
Herceptin. We demonstrated that Herceptin could interact with CD16 on NK cells
to expand the cytotoxic NK (specifically, CD56dim) cell population.
Additionally, Herceptin increased NK cell migration and cytotoxicity against
HER2+ breast cancer cells. In a pilot study, Herceptin-treated NK cells shrunk
lung nodular metastasis in a woman with HER2+ breast cancer who could not
tolerate the cardiotoxic side effects of Herceptin. Our findings support the
therapeutic potential of Herceptin-treated NK cells in patients with HER2+ and
Herceptin-intolerant breast cancer. Trastuzumab (Herceptin®), a monoclonal antibody against the ErbB2 (HER2)
receptor, has significantly improved clinical outcomes for HER2+ breast cancer
patients. However, the drug also has known cardiotoxic side effects through
mechanisms that are not fully understood. Here we utilized human induced
pluripotent stem cell-derived cardiomyocytes (iPS-CMs) to model
trastuzumab-related cardiotoxicity in vitro. We demonstrate that cardiotoxic
effects of ErbB2 inhibition by trastuzumab can be recapitulated only when the
cardioprotective effects of ErbB2/4 signaling is observed. We observed no
cardioprotective effects of ErbB2/4 signaling without cellular stress
(doxorubicin exposure in this study). In addition to neuregulin-1 (NRG-1), we
show that heparin-binding epidermal growth factor-like growth factor (HB-EGF)
also provides cardioprotective effects for iPS-CMs. Finally, we demonstrate a
simple, high-throughput co-culture platform utilizing iPS-CMs and endothelial
cells that is capable of detecting trastuzumab-related cardiotoxicity. We
conclude that iPS-CMs can recapitulate trastuzumab-related cardiotoxicity, and
may be used to elucidate additional modes of toxicity of trastuzumab and related
compounds. HER2-positive breast cancer correlates with more aggressive tumor growth, a
poorer prognosis and reduced overall survival. Currently, trastuzumab
(Herceptin), which is an anti-HER2 antibody, is one of the key drugs. There is
evidence indicating that conjugation of trastuzumab with chemotherapy drugs,
such as doxorubicin (DOX), for multiple targets could be more effective.
However, incomplete penetration into tumors has been noted for those conjugates.
Compared to an antibody, peptides may represent an attractive alternative. For
HER2, a similar potency has been observed for a 12-amino-acid anti-HER2 peptide
mimetic YCDGFYACYMDV-NH2 (AHNP, disulfide-bridged) and full-length trastuzumab.
Thus, a peptide, GPLGLAGDDYCDGFYACYMDV-NH2, which consists of AHNP and an MMP-2
cleavable linker GPLGLAGDD, was first designed, followed by conjugation with DOX
via a glycine residue at the N-terminus to form a novel DOX-peptide conjugate
MAHNP-DOX. Using HER2-positive human breast cancer cells BT474 and SKBR3 as in
vitro model systems and nude mice with BT474 xenografts as an in vivo model,
this conjugate was comprehensively characterized, and its efficacy was evaluated
and compared with that of free DOX. As a result, MAHNP-DOX demonstrated a much
lower in vitro IC50, and its in vivo extent of inhibition in mice was more
evident. During this process, enzymatic cleavage of MAHNP-DOX is critical for
its activation and cellular uptake. In addition, a synergistic response was
observed after the combination of DOX and AHNP. This effect was probably due to
the involvement of AHNP in the PI3K-AKT signaling pathway, which can be largely
activated by DOX and leads to anti-apoptotic signals. Breast cancer (BCa) is the most common cancer affecting women worldwide.
Overexpression of human epidermal growth factor receptor 2 (HER2) occurs in
~20-25% of invasive ductal breast carcinomas and is associated with the more
aggressive phenotype. Herceptin, a humanized antibody against HER2, is a
standard therapy in HER2-overexpressing cases. Approximately one-third of
patients relapse despite treatment. Therefore numerous studies have investigated
the molecular mechanisms associated with Herceptin resistance. An interaction
between HER2 signalling and steroid hormone receptor signalling pathways has
been previously investigated, but the effect of this relationship on Herceptin
resistance has never been studied. The present study analysed an impact of the
steroid hormone, progesterone (PG), on Herceptin-dependent cell growth
inhibition. Results indicated that Herceptin-inhibited proliferation of breast
cancer cell lines overexpressing HER2 (BT474 and MCF/HER2) in 3D culture is
abolished by PG. Furthermore, results demonstrated that PG led to the activation
of HER2/HER3-mediated signalling. Moreover, PG treatment induced a shift of
Herceptin-dependent cell cycle arrest in G1 phase towards S and G2 phases with
concomitant upregulation of cyclin-dependent kinase 2 (CDK2) and downregulation
of CDK inhibitor p27Kip1. These results demonstrate for the first time PG
involvement in the failure of Herceptin treatment in vitro. The present
observations suggest that cross-talk between PG- and HRG/HER2-initiated
signalling pathways may lead to the acquisition of resistance to Herceptin in
patients with BCa. BACKGROUND: Trastuzumab (Herceptin® [H]) is the standard of care for
HER2-positive locally advanced/metastatic breast cancer and
gastric/gastroesophageal junction (GEJ) cancer. However, there is a paucity of
data available on long-term H treatment of patients. The Rollover Protocol (ROP)
Study was conducted to report safety data for patients with HER2-positive
locally advanced/metastatic breast and gastric/GEJ cancer who have received
long-term H therapy (≥ 5 years and ≥ 3 years for breast and gastric/GEJ cancer,
respectively).
METHODS: The ROP Study was a single-arm, multicenter, international continuation
trial of H in patients who had previously completed a global Roche-sponsored
trial with H therapy, had stable disease, and were receiving H at the end of the
lead-in trial. Patients with chronic heart failure during the lead-in trial
could be included following a risk-benefit analysis. The primary objectives were
to provide H therapy to patients with HER2-overexpressing locally
advanced/metastatic breast or gastric/GEJ cancer at the end of the lead-in
study, and to follow the long-term outcomes and long-term overall safety in
these patients.
RESULTS: Twenty-five of 69 patients enrolled in the ROP Study received long-term
H therapy (19 breast cancer and 6 gastric/GEJ cancer). The median duration of H
treatment for patients with breast cancer was 8 years 7 months, and 5 years
2 months for patients with gastric/GEJ cancer. The cardiac status of the
patients remained stable over time, with no serious cardiac adverse events or
marked changes in left ventricular ejection fraction (LVEF). The median overall
worst LVEF measurement was 57.0%, and no patients experienced an LVEF of < 45%
(range 47-63%). There were no serious adverse events related to study treatment.
CONCLUSIONS: These results suggest that H has an acceptable safety profile and
was well tolerated in patients who received long-term H therapy (≥ 5 years and ≥
3 years for breast and gastric/GEJ cancer, respectively). Further investigation
and reporting of long-term H therapy would be valuable.
TRIAL REGISTRATION: This study was retrospectively registered on March 24, 2016
with Clinicaltrials.gov, number NCT02721641 . BACKGROUND: 25% of breast cancer patients suffer from aggressive HER2-positive
tumours that are characterised by overexpression of the HER2 protein or by its
increased tyrosine kinase activity. Herceptin is a major drug used to treat HER2
positive breast cancer. Understanding the molecular events that occur when
breast cancer cells are exposed to Herceptin is therefore of significant
importance. Dual specificity phosphatases (DUSPs) are central regulators of cell
signalling that function downstream of HER2, but their role in the cellular
response to Herceptin is mostly unknown. This study aims to model the initial
effects of Herceptin exposure on DUSPs in HER2-positive breast cancer cells
using Boolean modelling.
RESULTS: We experimentally measured expression time courses of 21 different
DUSPs between 0 and 24 h following Herceptin treatment of human MDA-MB-453
HER2-positive breast cancer cells. We clustered these time courses into patterns
of similar dynamics over time. In parallel, we built a series of Boolean models
representing the known regulatory mechanisms of DUSPs and then demonstrated that
the dynamics predicted by the models is in agreement with the experimental data.
Furthermore, we used the models to predict regulatory mechanisms of DUSPs, where
these mechanisms were partially known.
CONCLUSIONS: Boolean modelling is a powerful technique to investigate and
understand signalling pathways. We obtained an understanding of different
regulatory pathways in breast cancer and new insights on how these signalling
pathways are activated. This method can be generalized to other drugs and longer
time courses to better understand how resistance to drugs develops in cancer
cells over time. |
Has saracatinib been tested in clinical trials? | Yes, saracatinib has been tested in multiple clinical trials. | INTRODUCTION: Despite significant progress, a disease-modifying therapy for
Alzheimer's disease (AD) has not yet been developed. Recent findings implicate
soluble oligomeric amyloid beta as the most relevant protein conformation in AD
pathogenesis. We recently described a signaling cascade whereby oligomeric
amyloid beta binds to cellular prion protein on the neuronal cell surface,
activating intracellular Fyn kinase to mediate synaptotoxicity. Fyn kinase has
been implicated in AD pathophysiology both in in vitro models and in human
subjects, and is a promising new therapeutic target for AD. Herein, we present a
Phase Ib trial of the repurposed investigational drug AZD0530, a Src family
kinase inhibitor specific for Fyn and Src kinase, for the treatment of patients
with mild-to-moderate AD.
METHODS: The study was a 4-week Phase Ib multiple ascending dose, randomized,
double-blind, placebo-controlled trial of AZD0530 in AD patients with
Mini-Mental State Examination (MMSE) scores ranging from 16 to 26. A total of 24
subjects were recruited in three sequential groups, with each randomized to
receive oral AZD0530 at doses of 50 mg, 100 mg, 125 mg, or placebo daily for
4 weeks. The drug:placebo ratio was 3:1. Primary endpoints were safety,
tolerability, and cerebrospinal fluid (CSF) penetration of AZD0530. Secondary
endpoints included changes in clinical efficacy measures (Alzheimer's Disease
Assessment Scale - cognitive subscale, MMSE, Alzheimer's Disease Cooperative
Study - Activities of Daily Living Inventory, Neuropsychiatric Inventory, and
Clinical Dementia Rating Scale - Sum of Boxes) and regional cerebral glucose
metabolism measured by fluorodeoxyglucose positron emission tomography.
RESULTS: AZD0530 was generally safe and well tolerated across doses. One subject
receiving 125 mg of AZD0530 was discontinued from the study due to the
development of congestive heart failure and atypical pneumonia, which were
considered possibly related to the study drug. Plasma/CSF ratio of AZD0530 was
0.4. The 100 mg and 125 mg doses achieved CSF drug levels corresponding to brain
levels that rescued memory deficits in transgenic mouse models. One-month
treatment with AZD0530 had no significant effect on clinical efficacy measures
or regional cerebral glucose metabolism.
CONCLUSIONS: AZD0530 is reasonably safe and well tolerated in patients with
mild-to-moderate AD, achieving substantial central nervous system penetration
with oral dosing at 100-125 mg. Targeting Fyn kinase may be a promising
therapeutic approach in AD, and a larger Phase IIa clinical trial of AZD0530 for
the treatment of patients with AD has recently launched.
TRIAL REGISTRATION: ClinicalTrials.gov: NCT01864655. Registered 12 June 2014. OBJECTIVES: Thymic maligcies are rare, and options are limited for metastatic
disease. Src plays a role in normal thymic epithelial maturation, and its
inhibition with the oral compound saracatinib was postulated to be effective in
controlling thymic maligcy.
MATERIALS AND METHODS: Patients with unresectable thymic maligcy were treated
with saracatinib 175mg by mouth daily in 28 days cycles with radiographic
evaluation at cycle 2 day 1 for safety, then cycle 3 day 1 and every 8 weeks
thereafter. Response was evaluated by RECIST 1.0. A two-stage optimal design was
used, powered to detect a true response rate of 20%.
RESULTS: 21 patients were enrolled at two institutions, 12 of them with thymoma,
9 with thymic carcinoma. Thymoma patients received a median of 4.5 cycles and
thymic carcinoma patients a median of 1 cycle. There were no responses, so
accrual was halted after the first stage per protocol. 9 patients had stable
disease beyond the first assessment. Median time to progression was 5.7 months
for thymoma patients and 3.6 months for thymic carcinoma patients. Saracatinib
was well tolerated.
CONCLUSION: Src inhibition by saracatinib did not produce any radiographic
responses, though some patients did experience stable disease. Though negative,
this study shows the feasibility of completing a trial in this rare disease, and
of accruing reasonably significant numbers of thymic carcinoma patients. More
clinical trials are required for this population (NCT00718809). BACKGROUND: Fyn is a kinase that is upregulated in a subset of metastatic
castration-resistant prostate cancer. Saracatinib potently inhibits Fyn
activation. We have noted a relationship between Fyn expression and directional
motility, a cellular process related to metastasis. As such we hypothesized that
treatment with saracatinib would increase the time required to develop new
metastatic lesions.
METHODS: Patients with metastatic castration-resistant prostate cancer that had
progressed after docetaxel were eligible for enrollment. This study was executed
as a randomized discontinuation trial. During a lead-in phase of two 28-Day
cycles, all patients received saracatinib. Afterward, patients with
radiographically stable disease were randomized to either saracatinib or
placebo. Patients continued treatment until evidence of new metastasis.
RESULTS: Thirty-one patients were treated. Only 26% of patients had stable
disease after 8 weeks and thus proceeded to randomization. This required early
termination of the study for futility. The 70% of patients who progressed after
the lead-in phase exhibited expansion of existing lesions or decompensation due
to clinical progression without new metastatic lesions. Fatigue was reported in
more than 25% of patients (all grades) with only two patients experiencing grade
3 toxicity. Other grade 3 adverse events included dehydration, thrombocytopenia,
and weakness.
CONCLUSIONS: This study was unable to determine if saracatinib had potential as
metastasis inhibitor. Metastasis inhibition by saracatinib may still be viable
in an earlier time in the disease history. |
What animal is thought to be the host for the Coronavirus causing MERS? | The animal thought to be the host for the Coronavirus causing MERS is camels. | Since the emergence of Middle East respiratory syndrome coronavirus (MERS-CoV)
in 2012, there have been a number of clusters of human-to-human transmission.
These cases of human-to-human transmission involve close contact and have
occurred primarily in healthcare settings, and they are suspected to result from
repeated zoonotic introductions. In this study, we sequenced whole MERS-CoV
genomes directly from respiratory samples collected from 23 confirmed MERS cases
in the United Arab Emirates (UAE). These samples included cases from three
nosocomial and three household clusters. The sequences were analysed for changes
and relatedness with regard to the collected epidemiological data and other
available MERS-CoV genomic data. Sequence analysis supports the epidemiological
data within the clusters, and further, suggests that these clusters emerged
independently. To understand how and when these clusters emerged, respiratory
samples were taken from dromedary camels, a known host of MERS-CoV, in the same
geographic regions as the human clusters. Middle East respiratory syndrome
coronavirus genomes from six virus-positive animals were sequenced, and these
genomes were nearly identical to those found in human patients from
corresponding regions. These data demonstrate a genetic link for each of these
clusters to a camel and support the hypothesis that human MERS-CoV diversity
results from multiple zoonotic introductions. Coronavirus (CoV) infections are commonly associated with respiratory and
enteric disease in humans and animals. In 2012, a new human disease called
Middle East respiratory syndrome (MERS) emerged in the Middle East. MERS was
caused by a virus that was originally called human coronavirus-Erasmus Medical
Center/2012 but was later renamed as Middle East respiratory syndrome
coronavirus (MERS-CoV). MERS-CoV causes high fever, cough, acute respiratory
tract infection, and multiorgan dysfunction that may eventually lead to the
death of the infected individuals. The exact origin of MERS-CoV remains unknown,
but the transmission pattern and evidence from virological studies suggest that
dromedary camels are the major reservoir host, from which human infections may
sporadically occur through the zoonotic transmission. Human to human
transmission also occurs in healthcare facilities and communities. Recent
studies on Middle Eastern respiratory continue to highlight the need for further
understanding the virus-host interactions that govern disease severity and
infection outcome. In this review, we have highlighted the major mechanisms of
immune evasion strategies of MERS-CoV. We have demonstrated that M, 4a, 4b
proteins and Plppro of MERS-CoV inhibit the type I interferon (IFN) and nuclear
factor-κB signaling pathways and therefore facilitate innate immune evasion. In
addition, nonstructural protein 4a (NSP4a), NSP4b, and NSP15 inhibit
double-stranded RNA sensors. Therefore, the mentioned proteins limit early
induction of IFN and cause rapid apoptosis of macrophages. MERS-CoV strongly
inhibits the activation of T cells with downregulation of antigen presentation.
In addition, uncontrolled secretion of interferon ɣ-induced protein 10 and
monocyte chemoattractant protein-1 can suppress proliferation of human myeloid
progenitor cells. |
Which molecules are inhibited by Gilteritinib? | Gilteritinib is a novel, dual FLT3/AXL inhibitor with promising early phase trial data for acute myeloid leukemia. | Advances in the understanding of the molecular basis for acute myeloid leukemia
(AML) have generated new potential targets for treatment. Fms-like tyrosine
kinase 3 (FLT3) is one of the most frequently mutated genes in AML and mutations
in this gene are associated with poor overall survival. AXL plays a role in the
activation of FLT3 and has been implicated in the pathogenesis of AML. The
studies reported here evaluated the ability of a novel FLT3/AXL inhibitor,
gilteritinib, to block mutated FLT3 in cellular and animal models of AML.
Initial kinase studies showed that gilteritinib, a type I tyrosine kinase
inhibitor, was highly selective for both FLT3 and AXL while having weak activity
against c-KIT. Gilteritinib demonstrated potent inhibitory activity against the
internal tandem duplication (FLT3-ITD) and FLT3-D835Y point mutations in
cellular assays using MV4-11 and MOLM-13 cells as well as Ba/F3 cells expressing
mutated FLT3. Gilteritinib also inhibited FLT3-F691 mutations, although to a
lesser degree, in these assays. Furthermore, gilteritinib decreased the
phosphorylation levels of FLT3 and its downstream targets in both cellular and
animal models. In vivo, gilteritinib was distributed at high levels in
xenografted tumors after oral administration. The decreased FLT3 activity and
high intratumor distribution of gilteritinib translated to tumor regression and
improved survival in xenograft and intra-bone marrow transplantation models of
FLT3-driven AML. No overt toxicity was seen in mouse models treated with
gilteritinib. These results indicate that gilteritinib may be an important
next-generation FLT3 inhibitor for use in the treatment of FLT3
mutation-positive AML. BACKGROUND: Internal tandem duplication mutations in FLT3 are common in acute
myeloid leukaemia and are associated with rapid relapse and short overall
survival. The clinical benefit of FLT3 inhibitors in patients with acute myeloid
leukaemia has been limited by rapid generation of resistance mutations,
particularly in codon Asp835 (D835). We aimed to assess the highly selective
oral FLT3 inhibitor gilteritinib in patients with relapsed or refractory acute
myeloid leukaemia.
METHODS: In this phase 1-2 trial, we enrolled patients aged 18 years or older
with acute myeloid leukaemia who either were refractory to induction therapy or
had relapsed after achieving remission with previous treatment. Patients were
enrolled into one of seven dose-escalation or dose-expansion cohorts assigned to
receive once-daily doses of oral gilteritinib (20 mg, 40 mg, 80 mg, 120 mg, 200
mg, 300 mg, or 450 mg). Cohort expansion was based on safety and tolerability,
FLT3 inhibition in correlative assays, and antileukaemic activity. Although the
presence of an FLT3 mutation was not an inclusion criterion, we required ten or
more patients with locally confirmed FLT3 mutations (FLT3mut+) to be enrolled in
expansion cohorts at each dose level. On the basis of emerging findings, we
further expanded the 120 mg and 200 mg dose cohorts to include FLT3mut+ patients
only. The primary endpoints were the safety, tolerability, and pharmacokinetics
of gilteritinib. Safety and tolerability were assessed in the safety analysis
set (all patients who received at least one dose of gilteritinib). Responses
were assessed in the full analysis set (all patients who received at least one
dose of study drug and who had at least one datapoint post-treatment).
Pharmacokinetics were assessed in a subset of the safety analysis set for which
sufficient data for concentrations of gilteritinib in plasma were available to
enable derivation of one or more pharmacokinetic variables. This study is
registered with ClinicalTrials.gov, number NCT02014558, and is ongoing.
FINDINGS: Between Oct 15, 2013, and Aug 27, 2015, 252 adults with relapsed or
refractory acute myeloid leukaemia received oral gilteritinib once daily in one
of seven dose-escalation (n=23) or dose-expansion (n=229) cohorts. Gilteritinib
was well tolerated; the maximum tolerated dose was established as 300 mg/day
when two of three patients enrolled in the 450 mg dose-escalation cohort had two
dose-limiting toxicities (grade 3 diarrhoea and grade 3 elevated aspartate
aminotransferase). The most common grade 3-4 adverse events irrespective of
relation to treatment were febrile neutropenia (97 [39%] of 252), anaemia (61
[24%]), thrombocytopenia (33 [13%]), sepsis (28 [11%]), and pneumonia (27
[11%]). Commonly reported treatment-related adverse events were diarrhoea (92
[37%] of 252]), anaemia (86 [34%]), fatigue (83 [33%]), elevated aspartate
aminotransferase (65 [26%]), and increased alanine aminotransferase (47 [19%]).
Serious adverse events occurring in 5% or more of patients were febrile
neutropenia (98 [39%] of 252; five related to treatment), progressive disease
(43 [17%]), sepsis (36 [14%]; two related to treatment), pneumonia (27 [11%]),
acute renal failure (25 [10%]; five related to treatment), pyrexia (21 [8%];
three related to treatment), bacteraemia (14 [6%]; one related to treatment),
and respiratory failure (14 [6%]). 95 people died in the safety analysis set, of
which seven deaths were judged possibly or probably related to treatment
(pulmonary embolism [200 mg/day], respiratory failure [120 mg/day], haemoptysis
[80 mg/day], intracranial haemorrhage [20 mg/day], ventricular fibrillation [120
mg/day], septic shock [80 mg/day], and neutropenia [120 mg/day]). An
exposure-related increase in inhibition of FLT3 phosphorylation was noted with
increasing concentrations in plasma of gilteritinib. In-vivo inhibition of FLT3
phosphorylation occurred at all dose levels. At least 90% of FLT3
phosphorylation inhibition was seen by day 8 in most patients receiving a daily
dose of 80 mg or higher. 100 (40%) of 249 patients in the full analysis set
achieved a response, with 19 (8%) achieving complete remission, ten (4%)
complete remission with incomplete platelet recovery, 46 (18%) complete
remission with incomplete haematological recovery, and 25 (10%) partial
remission INTERPRETATION: Gilteritinib had a favourable safety profile and
showed consistent FLT3 inhibition in patients with relapsed or refractory acute
myeloid leukaemia. These findings confirm that FLT3 is a high-value target for
treatment of relapsed or refractory acute myeloid leukaemia; based on activity
data, gilteritinib at 120 mg/day is being tested in phase 3 trials.
FUNDING: Astellas Pharma, National Cancer Institute (Leukemia Specialized
Program of Research Excellence grant), Associazione Italiana Ricerca sul Cancro. Acute myeloid leukemia (AML) is a heterogeneous disease with cure rates of only
30-40% in patients <60 years old. Cytogenetic and molecular markers have
improved our understanding of the different prognostic entities in AML. FLT3
mutations are present in 30-40% of AML cases, conferring a poor prognosis with
reduced survival. AXL activates FLT3, impacting adversely on outcome. Both FLT3
and AXL constitute promising molecular targets. ASP2215 (gilteritinib) is a
novel, dual FLT3/AXL inhibitor with promising early phase trial data
(NCT02014558). A Phase III randomized multicenter clinical trial, comparing
ASP2215 to salvage chemotherapy in relapsed/refractory AML with FLT3-mutations
is now open to recruitment (NCT02421939). Trial design and objectives are
discussed here. FLT3-ITD is the most frequent tyrosine kinase mutation in acute myeloid leukemia
(AML) associated with poor prognosis. We previously reported that activation of
STAT5 confers resistance to PI3K/Akt inhibitors on the FLT3-ITD-positive AML
cell line MV4-11 and 32D cells driven by FLT3-ITD (32D/ITD) but not by FLT3
mutated in the tyrosine kinase domain (32D/TKD). Here, we report the involvement
of Pim kinases expressed through STAT5 activation in acquisition of this
resistance. The specific pan-Pim kinase inhibitor AZD1208 as well as PIM447 in
combination with the PI3K inhibitor GDC-0941 or the Akt inhibitor MK-2206
cooperatively downregulated the mTORC1/4EBP1 pathway, formation of the
eIF4E/eIF4G complex, and Mcl-1 expression leading to activation of Bak and Bax
to induce caspase-dependent apoptosis synergistically in these cells. These
cooperative effects were enhanced or inhibited by knock down of mTOR or
expression of its activated mutant, respectively. Overexpression of Mcl-1
conferred the resistance on 32D/ITD cells to combined inhibition of the PI3K/Akt
pathway and Pim kinases, while the Mcl-1-specific BH3 mimetic A-1210477
conquered the resistance of MV4-11 cells to GDC-0941. Furthermore,
overexpression of Pim-1 in 32D/TKD enhanced the mTORC1/Mcl-1 pathway and
partially protected it from the PI3K/Akt inhibitors or the FLT3 inhibitor
gilteritinib to confer the resistance to PI3K/Akt inhibitors. Finally, AZD1208
and GDC-0941 cooperatively inhibited the mTORC1/Mcl-1 pathway and reduced viable
cell numbers of primary AML cells from some FLT3-ITD positive cases. Thus, Pim
kinases may protect the mTORC1/4EBP1/Mcl-1 pathway to confer the resistance to
the PI3K/Akt inhibitors on FLT3-ITD cells and represent promising therapeutic
targets. Internal tandem duplications in fms-like tyrosine kinase 3 (FLT3-ITDs) are
common in acute myeloid leukemia (AML) and confer a poor prognosis. A sensitive
and specific assay for the detection of minimal residual disease (MRD) in
FLT3-ITD mutated AML could guide therapy decisions. Existing assays for MRD in
FLT3-ITD AML have not been particularly useful because of limited sensitivity.
We developed a sensitive and specific MRD assay for FLT3-ITD mutations using
next-generation sequencing. The initial validation of this assay was performed
by spiking fixed amounts of mutant DNA into wild-type DNA to establish a
sensitivity of detection equivalent to ≥1 FLT3-ITD-containing cell in 10 000,
with a minimum input of 100 000 cell equivalents of DNA. We subsequently
validated the assay in bone marrow samples from patients with FLT3-ITD AML in
remission. Finally, we analyzed bone marrow samples from 80 patients with
FLT3-ITD relapsed/refractory AML participating in a trial of a novel FLT3
inhibitor, gilteritinib, and demonstrated a relationship between the mutation
burden, as detected by the assay, and overall survival. This novel MRD assay is
specific and 2 orders of magnitude more sensitive than currently available
polymerase chain reaction- or next-generation sequencing-based FLT3-ITD assays.
The assay is being prospectively validated in ongoing randomized clinical
trials. Controversy exists whether internal tandem duplication of FMS-like tyrosine
kinase 3 (FLT3-internal tandem duplication [ITD]) allelic ratio (AR) and/or
length of the ITD should be taken into account for risk stratification of
pediatric acute myeloid leukemia (AML) and whether it should be measured on RNA
or DNA. Moreover, the ITD status may be of relevance for selecting patients
eligible for FLT3 inhibitors. Here, we included 172 pediatric AML patients, of
whom 36 (21%) harbored FLT3-ITD as determined on both RNA and DNA. Although
there was a good correlation between both parameters ARspearman = 0.62 (95%
confidence interval, 0.22-0.87) and ITDlengthspearman = 0.98 (95% confidence
interval, 0.90-1.00), only AR ≥ 0.5 and length ≥48 base pairs (bps) based on RNA
measurements were significantly associated with overall survival (AR: Plogrank =
.008; ITDlength: Plogrank = .011). In large ITDs (>156 bp on DNA) a remarkable
90-bp difference exists between DNA and RNA, including intron 14, which is
spliced out in RNA. Ex vivo exposure (n = 30) to FLT3 inhibitors, in particular
to the FLT3-specific inhibitor gilteritinib, showed that colony-forming capacity
was significantly more reduced in FLT3-ITD-AR ≥ 0.5 compared with ITD-AR-low and
ITD- patient samples (P < .001). RNA-based FLT3-ITD measurements are recommended
for risk stratification, and the relevance of AR regarding eligibility for
FLT3-targeted therapy warrants further study. Gilteritinib, a novel, highly specific, potent fms-like tyrosine kinase 3/AXL
inhibitor, demonstrated antileukemic activity in patients with
relapsed/refractory (R/R) acute myeloid leukemia (AML). In this open-label phase
1 study (NCT02181660), Japanese patients (aged ≥18 years) with R/R AML received
once-daily gilteritinib, escalating from 20 to 300 mg/d. Primary endpoints were
safety/tolerability, including the maximum tolerated dose (MTD) and the
recommended dose (RD); secondary endpoints were antileukemic activity and
pharmacokinetics (PK). Twenty-four Japanese patients with R/R AML received
once-daily oral gilteritinib in 1 of 6 dose-escalation cohorts (20, 40, 80, 120,
200, and 300 mg/d). Gilteritinib was well tolerated. The MTD was 200 mg/d;
dose-limiting toxicities were grade 3 tumor lysis syndrome (120 mg/d; n = 1);
and grade 3 elevated blood lactate dehydrogenase, amylase, blood creatine
phosphokinase levels, and syncope (all n = 2; 300 mg/d). The RD was 120 mg/d.
The most common drug-related grade ≥3 adverse events were thrombocytopenia
(n = 4 [16.7%]) and increased blood creatine phosphokinase (n = 3 [12.5%]).
Gilteritinib had a dose-proportional PK profile. Among patients with mutated
fms-like tyrosine kinase 3, the overall response rate (ORR) was 80% (n = 4 of 5;
complete remission [CR] with incomplete platelet recovery, 1 [20%]; CR with
incomplete hematologic recovery, 2 [40%]; partial remission (PR), 1 [20%]).
Among patients with wild-type fms-like tyrosine kinase 3, ORR was 36.4%; (n = 4
of 11; CR, 1 [9.1%]; CR with incomplete platelet recovery, 2 [18.2%]; PR, 1
[9.1%]). In conclusion, gilteritinib was well tolerated and demonstrated
antileukemic activity in a Japanese R/R AML population. The ROS1 tyrosine kinase inhibitor (TKI) crizotinib has shown dramatic effects
in patients with non-small cell lung cancer (NSCLC) harboring ROS1 fusion genes.
However, patients inevitably develop resistance to this agent. Therefore, a new
treatment strategy is required for lung tumors with ROS1 fusion genes. In the
present study, lung cancer cell lines, HCC78 harboring SLC34A2-ROS1 and ABC-20
harboring CD74-ROS1, were used as cell line-based resistance models.
Crizotinib-resistant HCC78R cells were established from HCC78. We
comprehensively screened the resistant cells using a phosphor-receptor tyrosine
kinase array and RNA sequence analysis by next-generation sequencing. HCC78R
cells showed upregulation of HB-EGF and activation of epidermal growth factor
receptor (EGFR) phosphorylation and the EGFR signaling pathway. Recombit
HB-EGF or EGF rendered HCC78 cells or ABC-20 cells resistant to crizotinib. RNA
sequence analysis by next-generation sequencing revealed the upregulation of AXL
in HCC78R cells. HCC78R cells showed marked sensitivity to EGFR-TKI or anti-EGFR
antibody treatment in vitro. Combinations of an AXL inhibitor, cabozantinib or
gilteritinib, and an EGFR-TKI were more effective against HCC78R cells than
monotherapy with an EGFR-TKI or AXL inhibitor. The combination of cabozantinib
and gefitinib effectively inhibited the growth of HCC78R tumors in an in vivo
xenograft model of NOG mice. The results of this study indicated that
HB-EGF/EGFR and AXL play roles in crizotinib resistance in lung cancers
harboring ROS1 fusions. The combination of cabozantinib and EGFR-TKI may
represent a useful alternative treatment strategy for patients with advanced
NSCLC harboring ROS1 fusion genes. Internal tandem duplication (ITD) in Fms-like tyrosine kinase 3 (FLT3) is
frequently observed in acute myeloid leukemia (AML). Quizartinib, gilteritinib,
and midostaurin are inhibitors against FLT3-ITD that have good efficacy for
FLT3-ITD-positive AML patients. Long-term administration leads to drug
resistance through acquired tyrosine kinase domain (TKD) mutations in FLT3-ITD,
such as N676K, F691L, D835V, and Y842C. Here, our screen to detect inhibitors
capable of overcoming resistance to FLT3 inhibitors identified heat shock
protein (HSP) 90 inhibitors as potential candidates. Although Ba/F3 cells
expressing FLT3-ITD with TKD mutations (Ba/F3-ITD+N676K, Ba/F3-ITD+F691L,
Ba/F3-ITD+D835V, and Ba/F3-ITD+Y842C) showed various resistance patterns to FLT3
inhibitors compared with Ba/F3-ITD cells that express FLT3-ITD lacking TKD
mutations, they were more sensitive to HSP90 inhibitors than Ba/F3 cells.
Notably, the Ba/F3-ITD+D835V cells were the most sensitive to HSP90 inhibitors.
Treatment with HSP90 inhibitors downregulated FLT3 and its downstream signaling
and induced G1 arrest followed by apoptosis in Ba/F3-ITD+N676K, Ba/F3-ITD+F691L,
Ba/F3-ITD+Y842C, and especially Ba/F3-ITD+D835V cells at lower concentrations
compared with Ba/F3-ITD cells. The downregulation of FLT3-ITD+D835V was caused
by rapid proteolysis in autophagy. Similar results were also observed in the
quizartinib-resistant MV4-11 cells, QR1 and QR2, which were established by
culturing cells in the presence of quizartinib and harbored FLT3-ITD+D835H and
FLT3-ITD+D835V, respectively, in a single allele. Interestingly, the efficacies
of HSP90 inhibitors in QR cells are reversely correlated with that of quizartib,
but not to gilteritinib and midostaurin. Collectively, HSP90 inhibitors are good
candidates to overcome drug resistance in AML with various FLT3-ITD TKD
mutations. Gilteritinib (Xospata®) is an orally available small molecule receptor tyrosine
kinase inhibitor developed by Astellas Pharma in collaboration with Kotobuki
Pharmaceutical for the treatment of acute myeloid leukaemia (AML) harbouring
FMS-like tyrosine kinase 3 (FLT3) mutations. Gilteritinib inhibits FLT3 (STK1 or
FLK2), AXL (UFO or JTK11) and anaplastic lymphoma kinase (ALK or CD246).
Gilteritinib inhibits FLT3 signalling in cells expressing FLT3 internal tandem
duplication (ITD), tyrosine kinase domain mutation FLT3-D835Y and the double
mutant FLT3-ITD-D835Y, thereby inducing apoptosis. Gilteritinib also binds to
and inhibits the wild-type and mutated forms of ALK, resulting in reduced tumour
cell proliferation in cancer cell types that overexpress the mutation.
Gilteritinib is approved in Japan for the treatment of relapsed or refractory
AML with FLT3 mutation. Recently, it was also approved in the USA for the
treatment of adult patients who have relapsed or refractory AML with a FLT3
mutation, as detected by an FDA-approved test. Clinical development of
gilteritinib is underway in several countries worldwide. Development for
non-small cell lung cancer and solid tumours has been discontinued. Activating internal tandem duplication (ITD) and tyrosine kinase domain (TKD)
point mutations in Fms-like tyrosine kinase 3 (FLT3) occur in approximately 30%
of patients with acute myeloid leukemia (AML), and confer a poor prognosis with
standard cytarabine/anthracycline or azacitidine-based chemotherapy regimens.
Gilteritinib is a highly-specific, potent FLT3/AXL inhibitor with demonstrated
activity against FLT3-ITD and FLT3-TKD mutations. Compared with salvage
chemotherapy, treatment with once-daily oral gilteritinib demonstrated a
clinical benefit in patients with FLT3-mutated relapsed/refractory AML, which
led to its recent approval in Japan and the United States. We investigated the
effects of gilteritinib combined with cytarabine plus daunorubicin/idarubicin,
or combined with azacitidine in human FLT3-ITD-positive (FLT3-ITD +) AML cell
lines and xenografted mouse models. Gilteritinib induced G1 arrest and apoptosis
in a dose-dependent manner. The addition of cytarabine, daunorubicin,
idarubicin, or azacitidine potentiated apoptosis. Gilteritinib alone or combined
with cytarabine, daunorubicin, idarubicin, or azacitidine, inhibited
anti-apoptotic protein expression in MV4-11 cells. In xenografted mice,
administration of cytarabine, idarubicin, or azacitidine in combination with
gilteritinib had little impact on plasma or intratumor PK profiles of
gilteritinib, cytarabine, idarubicin, or azacitidine. Gilteritinib combined with
chemotherapy reduced tumor volume to a greater extent than either gilteritinib
or chemotherapy alone. Of note, the addition of cytarabine plus
daunorubicin/idarubicin led to tumor regression in mice, with complete
regression observed in six out of eight mice in both triple combination groups.
These findings support the investigation of gilteritinib combined with
chemotherapy in patients with FLT3-ITD + AML, including those who are ineligible
for intensive chemotherapy. Gilteritinib is a potent and selective FLT3 kinase inhibitor with single-agent
clinical efficacy in relapsed/refractory FLT3-mutated acute myeloid leukemia
(AML). In this context, however, gilteritinib is not curative, and response
duration is limited by the development of secondary resistance. To evaluate
resistance mechanisms, we analyzed baseline and progression samples from
patients treated on clinical trials of gilteritinib. Targeted next-generation
sequencing at the time of AML progression on gilteritinib identified
treatment-emergent mutations that activate RAS/MAPK pathway signaling, most
commonly in NRAS or KRAS. Less frequently, secondary FLT3-F691L gatekeeper
mutations or BCR-ABL1 fusions were identified at progression. Single-cell
targeted DNA sequencing revealed diverse patterns of clonal selection and
evolution in response to FLT3 inhibition, including the emergence of RAS
mutations in FLT3-mutated subclones, the expansion of alternative wild-type FLT3
subclones, or both patterns simultaneously. These data illustrate dynamic and
complex changes in clonal architecture underlying response and resistance to
mutation-selective tyrosine kinase inhibitor therapy in AML. SIGNIFICANCE:
Comprehensive serial genotyping of AML specimens from patients treated with the
selective FLT3 inhibitor gilteritinib demonstrates that complex, heterogeneous
patterns of clonal selection and evolution mediate clinical resistance to
tyrosine kinase inhibition in FLT3-mutated AML. Our data support the development
of combinatorial targeted therapeutic approaches for advanced AML.See related
commentary by Wei and Roberts, p. 998.This article is highlighted in the In This
Issue feature, p. 983. FMS-like tyrosine kinase 3- internal tandem duplication (FLT3-ITD) remains as
one of the most frequently mutated genes in acute myeloid leukemia (AML),
especially in those with normal cytogenetics. The FLT3-ITD and FLT3-TKD
(tyrosine kinase domain) mutations are biomarkers for high risk AML and are
associated with drug resistance and high risk of relapse. Multiple FLT3
inhibitors are in clinical development, including lestaurtinib, tandutinib,
quizartinib, midostaurin, gilteritinib, and crenolanib. Midostaurin and
gilteritinib have been approved by FDA for Flt3 mutated AML. Gilteritinib
(ASP2215, Xospata) is a small molecule dual inhibitor of FLT3/AXL. The ADMIRAL
study showed that longer overall survival and higher response rate are
associated with gilteritinib in comparison with salvage chemotherapy for relapse
/refractory (R/R) AML. These data from the ADMIRAL study may lead to the therapy
paradigm shift and establish gilteritinib as the new standard therapy for R/R
FLT3-mutated AML. Currently, multiple clinical trials are ongoing to evaluate
the combination of gilteritinib with other agents and regimens. This study
summarized clinical trials of gilteritinib for AML. BACKGROUND: Patients with relapsed or refractory acute myeloid leukemia (AML)
with mutations in the FMS-like tyrosine kinase 3 gene (FLT3) infrequently have a
response to salvage chemotherapy. Gilteritinib is an oral, potent, selective
FLT3 inhibitor with single-agent activity in relapsed or refractory FLT3-mutated
AML.
METHODS: In a phase 3 trial, we randomly assigned adults with relapsed or
refractory FLT3-mutated AML in a 2:1 ratio to receive either gilteritinib (at a
dose of 120 mg per day) or salvage chemotherapy. The two primary end points were
overall survival and the percentage of patients who had complete remission with
full or partial hematologic recovery. Secondary end points included event-free
survival (freedom from treatment failure [i.e., relapse or lack of remission] or
death) and the percentage of patients who had complete remission.
RESULTS: Of 371 eligible patients, 247 were randomly assigned to the
gilteritinib group and 124 to the salvage chemotherapy group. The median overall
survival in the gilteritinib group was significantly longer than that in the
chemotherapy group (9.3 months vs. 5.6 months; hazard ratio for death, 0.64; 95%
confidence interval [CI], 0.49 to 0.83; P<0.001). The median event-free survival
was 2.8 months in the gilteritinib group and 0.7 months in the chemotherapy
group (hazard ratio for treatment failure or death, 0.79; 95% CI, 0.58 to 1.09).
The percentage of patients who had complete remission with full or partial
hematologic recovery was 34.0% in the gilteritinib group and 15.3% in the
chemotherapy group (risk difference, 18.6 percentage points; 95% CI, 9.8 to
27.4); the percentages with complete remission were 21.1% and 10.5%,
respectively (risk difference, 10.6 percentage points; 95% CI, 2.8 to 18.4). In
an analysis that was adjusted for therapy duration, adverse events of grade 3 or
higher and serious adverse events occurred less frequently in the gilteritinib
group than in the chemotherapy group; the most common adverse events of grade 3
or higher in the gilteritinib group were febrile neutropenia (45.9%), anemia
(40.7%), and thrombocytopenia (22.8%).
CONCLUSIONS: Gilteritinib resulted in significantly longer survival and higher
percentages of patients with remission than salvage chemotherapy among patients
with relapsed or refractory FLT3-mutated AML. (Funded by Astellas Pharma;
ADMIRAL ClinicalTrials.gov number, NCT02421939.). |
Is αCGRP a member of the CGRP family? | Yes, aCGRP, a 37-residue-long peptide hormone, is a novel amyloidogenic member of the CGRP family. | Author information:
(1)Section of Cell Biology and Biophysics, Department of Biology, School of
Sciences, National and Kapodistrian University of Athens, Panepistimiopolis,
Athens 157 01, Greece. Electronic address: [email protected].
(2)Section of Cell Biology and Biophysics, Department of Biology, School of
Sciences, National and Kapodistrian University of Athens, Panepistimiopolis,
Athens 157 01, Greece. Electronic address: [email protected].
(3)Section of Cell Biology and Biophysics, Department of Biology, School of
Sciences, National and Kapodistrian University of Athens, Panepistimiopolis,
Athens 157 01, Greece. Electronic address: [email protected].
(4)Section of Cell Biology and Biophysics, Department of Biology, School of
Sciences, National and Kapodistrian University of Athens, Panepistimiopolis,
Athens 157 01, Greece. Electronic address: [email protected].
(5)Department of Pharmacy, University of Patras, Patras 26504, Greece.
Electronic address: [email protected].
(6)Section of Cell Biology and Biophysics, Department of Biology, School of
Sciences, National and Kapodistrian University of Athens, Panepistimiopolis,
Athens 157 01, Greece. Electronic address: [email protected].
(7)Section of Cell Biology and Biophysics, Department of Biology, School of
Sciences, National and Kapodistrian University of Athens, Panepistimiopolis,
Athens 157 01, Greece. Electronic address: [email protected]. |
Which is the most common monogenic cause of common variable immunodeficiency (CVID) in Europeans? | Loss-of-function nuclear factor kB subunit 1 (NFKB1) variants are the most common monogenic cause of common variable immunodeficiency in Europeans. | Author information:
(1)Immunology Division, Hospital Universitari Vall d'Hebron (HUVH), Vall
d'Hebron Research Institute (VHIR), Department of Cell Biology, Physiology and
Immunology, Autonomous University of Barcelona (UAB), Barcelona, Catalonia,
Spain; Jeffrey Model Foundation Excellence Center, Barcelona, Catalonia, Spain.
(2)Area of Clinical and Molecular Genetics, Hospital Universitari Vall d'Hebron
(HUVH), Barcelona, Catalonia, Spain.
(3)Pathology Department, Hospital Universitari Vall d'Hebron (HUVH), Barcelona,
Catalonia, Spain.
(4)Pneumology Department, Hospital Universitari Vall d'Hebron (HUVH), Barcelona,
Catalonia, Spain.
(5)Exocrine Pancreas Research Unit, Department of Digestive Diseases, Hospital
Universitari Vall d'Hebron (HUVH), Autonomous University of Barcelona (UAB),
CiberEHD, Barcelona, Catalonia, Spain.
(6)Jeffrey Model Foundation Excellence Center, Barcelona, Catalonia, Spain;
Pediatric Infectious Diseases and Immunodeficiencies Unit (UPIIP), Hospital
Universitari Vall d'Hebron (HUVH), Vall d'Hebron Research Institute (VHIR),
Universitat Autònoma de Barcelona (UAB), Barcelona, Catalonia, Spain.
(7)Immunology Division, Hospital Universitari Vall d'Hebron (HUVH), Vall
d'Hebron Research Institute (VHIR), Department of Cell Biology, Physiology and
Immunology, Autonomous University of Barcelona (UAB), Barcelona, Catalonia,
Spain; Jeffrey Model Foundation Excellence Center, Barcelona, Catalonia, Spain;
Area of Clinical and Molecular Genetics, Hospital Universitari Vall d'Hebron
(HUVH), Barcelona, Catalonia, Spain. Electronic address: [email protected]. |
List 3 human diseases caused by viruses in the family Paramyxoviridae. | Measles, mumps and encephalitis are diseases caused by viruses in the family Paramyxoviridae. | Many of the common respiratory illnesses of infancy and childhood are caused by
viruses of the Paramyxoviridae family, in particular measles virus, respiratory
syncytial (RS) virus and parainfluenzavirus type 3 (PI3). Effective measles
vaccine was developed by classical methods, but these same methods have failed
to provide vaccines to control RS and PI3 virus infections. The WHO Programme
for Vaccine Development was initiated in 1983 to encourage the application of
the new biotechnologies to continuing problems, such as the acute virus-induced
respiratory diseases of childhood. At a meeting of research workers held in July
1986 under the auspices of this programme, renewed optimism was expressed
concerning the prospects for immunoprophylaxis of RS virus-induced disease.
Animal models are now available for evaluation of the immunogenic potential of
candidate vaccines. Vaccinia/RS recombit viruses have been produced which
have allowed the immunogenic properties of individual RS virus proteins to be
defined. Complete protection without the exacerbation of disease, which earlier
had accompanied the use of formalin-inactivated vaccines, has been achieved in
animals immunized with vaccinia virus recombits expressing the F protein;
partial protection was obtained using G protein gene vectors. PI3 appears to be
an inherently stable virus and evidence from animal experiments suggests that
bovine PI3 might be suitable for use as a live vaccine in man. BACKGROUND: Previous controversy was generated over the hypothesis that a
paramyxovirus such as measles or vaccination against such viruses might be
causally associated with inflammatory bowel disease (IBD). We aimed to determine
if Crohn's disease (CD) or ulcerative colitis (UC) subjects are more likely to
be seropositive for measles, mumps, or rubella than controls.
METHODS: Using our population-based University of Manitoba IBD Research Registry
we recruited CD (n = 235) and UC (n = 137) subjects ages 18-50 years for a study
involving detailed questionnaires and venipuncture. We accessed the
population-based databases of Manitoba Health (single provincial health insurer)
to get age-, gender-, and geography-matched non-IBD controls (n = 310). We used
a standard enzyme-linked immunosorbent assay (ELISA) to measure serum
antibodies.
RESULTS: Seropositivity for measles and mumps was similar in controls (98.1%,
78.4%, respectively) as in CD (96.2%, 72.3% respectively) and in UC (95.5%,
74.6%, respectively). However, controls were significantly more likely to be
seropositive for rubella (98.1%) than were CD cases (91.0%, P < 0.0002) or UC
cases (93.3%, P = 0.01). Males accounted for the significantly lower rates of
seropositivity to rubella with CD. While we determined that significantly more
controls than CD were vaccinated, we cannot be sure if the increased rate of
rubella seropositivity in controls is secondary to wildtype or
vaccine-associated infection.
CONCLUSIONS: These data suggest there is no association of having acquired
measles, mumps, or rubella (by natural infection or through vaccination) and CD
or UC. If anything, these data may suggest some protective effect of having
acquired rubella infection or vaccine against acquiring CD. Measles are a systemic infectious disease caused by a single stranded
ribonucleic acid virus (measles virus) from the paramyxovirus family. Typically,
the disease is characterized by a two-phase course. After an average incubation
period of 8 to 11 days, initial symptoms such as fever, cough, coryza and
conjunctivitis appear. Two thirds of the patients shows a white-marked ethema
on the buccal mucosa (Koplik's spots). After disappearance of these symptoms, a
second increase of temperature and the typical measles exanthema, a brownish-red
maculopapular rash, appear. Infection with measles virus induces transient
immunodeficiency that favours the formation of several complications. Some of
them, e. g. encephalitic diseases, are severe and associated with a high
mortality. Measles are world-wide distributed and belong to the ten most
frequent infectious diseases in some less developed countries. The disease is
associated with a high mortality in some African and South-East Asian countries,
in particular in children aged less than 12 months. Of particular note, measles
are the most important cause of blindness in children in population with
borderline vitamin A status. In Germany, the number of reported measles cases
has been declined dramatically since the introduction of a vaccine more than
four decades ago. However, regional outbreaks or small epidemics still occur.
Because there is no specific antiviral treatment, therapy of measles is
symptomatic and depends on the manifestation of the disease. The most important
prevention strategy is immunization with a life-attenuated vaccine that can be
applied as monovaccination or in combination with mumps and rubella virus (MMR
vaccination) or mumps, rubella and varicella virus (MMRV vaccination). BACKGROUND: Measles virus (MV) is a member of the Paramyxoviridae family and an
important human pathogen causing strong immunosuppression in affected
individuals and a considerable number of deaths worldwide. Currently, measles is
a re-emerging disease in developed countries. MV is usually quantified in
infectious units as determined by limiting dilution and counting of plaque
forming unit either directly (PFU method) or indirectly from random distribution
in microwells (TCID50 method). Both methods are time-consuming (up to several
days), cumbersome and, in the case of the PFU assay, possibly operator
dependent.
METHODS/FINDINGS: A rapid, optimized, accurate, and reliable technique for
titration of measles virus was developed based on the detection of virus
infected cells by flow cytometry, single round of infection and titer
calculation according to the Poisson's law. The kinetics follow up of the number
of infected cells after infection with serial dilutions of a virus allowed
estimation of the duration of the replication cycle, and consequently, the
optimal infection time. The assay was set up to quantify measles virus,
vesicular stomatitis virus (VSV), and human immunodeficiency virus type 1
(HIV-1) using antibody labeling of viral glycoprotein, virus encoded fluorescent
reporter protein and an inducible fluorescent-reporter cell line, respectively.
CONCLUSION: Overall, performing the assay takes only 24-30 hours for MV strains,
12 hours for VSV, and 52 hours for HIV-1. The step-by-step procedure we have set
up can be, in principle, applicable to accurately quantify any virus including
lentiviral vectors, provided that a virus encoded gene product can be detected
by flow cytometry. Negative-strand (NS) RNA viruses comprise many pathogens that cause serious
diseases in humans and animals. Despite their clinical importance, little is
known about the host factors required for their infection. Using vesicular
stomatitis virus (VSV), a prototypic NS RNA virus in the family Rhabdoviridae,
we conducted a human genome-wide siRNA screen and identified 72 host genes
required for viral infection. Many of these identified genes were also required
for infection by two other NS RNA viruses, the lymphocytic choriomeningitis
virus of the Arenaviridae family and human parainfluenza virus type 3 of the
Paramyxoviridae family. Genes affecting different stages of VSV infection, such
as entry/uncoating, gene expression, and assembly/release, were identified.
Depletion of the proteins of the coatomer complex I or its upstream effectors
ARF1 or GBF1 led to detection of reduced levels of VSV RNA. Coatomer complex I
was also required for infection of lymphocytic choriomeningitis virus and human
parainfluenza virus type 3. These results highlight the evolutionarily conserved
requirements for gene expression of diverse families of NS RNA viruses and
demonstrate the involvement of host cell secretory pathway in the process. The Paramyxoviridae family includes many viruses that are pathogenic in humans,
including parainfluenza viruses, measles virus, respiratory syncytial virus, and
the emerging zoonotic Henipaviruses. No effective treatments are currently
available for these viruses, and there is a need for efficient antiviral
therapies. Paramyxoviruses enter the target cell by binding to a cell surface
receptor and then fusing the viral envelope with the target cell membrane,
allowing the release of the viral genome into the cytoplasm. Blockage of these
crucial steps prevents infection and disease. Binding and fusion are driven by
two virus-encoded glycoproteins, the receptor-binding protein and the fusion
protein, that together form the viral "fusion machinery." The development of
efficient antiviral drugs requires a deeper understanding of the mechanism of
action of the Paramyxoviridae fusion machinery, which is still controversial.
Here, we review recent structural and functional data on these proteins and the
current understanding of the mechanism of the paramyxovirus cell entry process. Respiratory syncytial virus (RSV) belongs to the family Paramyxoviridae and is
the single most important cause of serious lower respiratory tract infections in
young children, yet no highly effective treatment or vaccine is available. To
clarify the potential for an anti-G mAb, 131-2G which has both anti-viral and
anti-inflammatory effects, to effectively treat RSV disease, we determined the
kinetics of its effect compared to the effect of the anti-F mAb, 143-6C on
disease in mice. Treatment administered three days after RSV rA2-line19F (r19F)
infection showed 131-2G decreased breathing effort, pulmonary mucin levels,
weight loss, and pulmonary inflammation earlier and more effectively than
treatment with mAb 143-6C. Both mAbs stopped lung virus replication at day 5
post-infection. These data show that, in mice, anti-G protein mAb is superior to
treating disease during RSV infection than an anti-F protein mAb similar to
Palivizumab. This combination of anti-viral and anti-inflammatory activity makes
131-2G a promising candidate for treating for active human RSV infection. Paget's disease of bone (PDB) is a common skeletal disorder characterised by
focal abnormalities of increased and disorganised bone turnover. Genetic factors
play a central role in the pathogenesis of PDB but environmental factors also
contribute. Measles virus (MV), respiratory syncytial virus (RSV) and canine
distemper virus (CDV) have all been implicated as potential disease triggers but
the data are conflicting. Since chronic paramyxovirus infection with measles is
known to be accompanied by increased production of antiviral antibodies, we have
analysed circulating concentrations of antibodies to MV, CDV, and RSV as well as
mumps, rubella and varicella zoster virus (VZV) in 463 patients with PDB and 220
aged and gender-matched controls. We also studied the relation between viral
antibody concentrations and various markers of disease severity and extent in
460 PDB patients. A high proportion of cases and controls tested positive for
antiviral antibodies but there was no significant difference in circulating
antibody concentrations between PDB cases and controls for MV, CDV, RSV, rubella
or VZV. However, mumps virus antibody levels were significantly higher in the
PDB cases (mean ± SD = 3.1 ± 0.84 vs. 2.62 ± 0.86. p < 0.001). There was no
association between disease severity and circulating antibody concentrations to
any of the viruses. In conclusion, we found no evidence to suggest that PDB is
associated with abnormalities of immune response to measles or other
paramyxoviruses, although there was evidence of a greater antibody response to
mumps. The results do not support that hypothesis that PDB is associated with a
persistent infection with measles or other paramyxoviruses. BACKGROUND: Mumps is a Paramyxoviridae virus. This disease was rampant prior to
introduction of the measles, mumps, and rubella vaccine, resulting in decreased
incidence. This disease has demonstrated several outbreaks.
OBJECTIVE: This review provides a focused evaluation of mumps, an update on
outbreaks, management recommendations, and ways to decrease transmission.
DISCUSSION: Clusters of mumps outbreaks continue to occur. The virus is a
paramyxovirus, a single-stranded RNA virus. The vaccine can provide lifelong
immunity if administered properly, though prior to 1967 and introduction of the
vaccine, the virus was common. In the past decade, there have been several
notable outbreaks. Humans are the only known hosts, with disease spread through
exposure to droplets and saliva. Factors affecting transmission include age,
compromised immunity, time of year, travel, and vaccination status. Upper
respiratory symptoms, fever, and headache are common, with unilateral or
bilateral parotitis, and the virus may spread to other systems. Diagnosis is
clinical, though polymerase chain reaction and immunoglobulin testing are
available. This review provides several recommendations for vaccine in
pregcy, patients living in close quarters, health care personnel, and those
immunocompromised. Treatment is generally supportive, with emphasis on proper
isolation to prevent widespread outbreaks. Although reporting regulations and
procedures vary by state, mumps is reportable in most states.
CONCLUSIONS: Mumps is an easily spread virus. Although vaccination is the most
effective way to prevent transmission, early recognition of the disease is
crucial. As an emergency physician, it is important to recognize the clinical
presentation, recommended testing, treatment, and isolation procedures. Respiratory viruses of human origin infect wild apes across Africa, sometimes
lethally. Here we report simultaneous outbreaks of two distinct human
respiratory viruses, human metapneumovirus (MPV; Pneumoviridae: Metapneumovirus)
and human respirovirus 3 (HRV3; Paramyxoviridae; Respirovirus, formerly known as
parainfluenza virus 3), in two chimpanzee (Pan troglodytes schweinfurthii)
communities in the same forest in Uganda in December 2016 and January 2017. The
viruses were absent before the outbreaks, but each was present in ill
chimpanzees from one community during the outbreak period. Clinical signs and
gross pathologic changes in affected chimpanzees closely mirrored symptoms and
pathology commonly observed in humans for each virus. Epidemiologic modelling
showed that MPV and HRV3 were similarly transmissible (R0 of 1.27 and 1.48,
respectively), but MPV caused 12.2% mortality mainly in infants and older
chimpanzees, whereas HRV3 caused no direct mortality. These results are
consistent with the higher virulence of MPV than HRV3 in humans, although both
MPV and HRV3 cause a significant global disease burden. Both viruses clustered
phylogenetically within groups of known human variants, with MPV closely related
to a lethal 2009 variant from mountain gorillas (Gorilla beringei beringei),
suggesting two independent and simultaneous reverse zoonotic origins, either
directly from humans or via intermediary hosts. These findings expand our
knowledge of human origin viruses threatening wild chimpanzees and suggest that
such viruses might be differentiated by their comparative epidemiological
dynamics and pathogenicity in wild apes. Our results also caution against
assuming common causation in coincident outbreaks. |
Which disease is caused by de novo VPS4A mutations? | Mutations in the VPS4A gene, which encodes the alpha-subunit of the lysosomal sorting enzyme, beta-N-acetylhexosaminidase 4, are the cause of multisystem disease type 4 or Ferroportin disease. | Author information:
(1)Cambridge Institute for Medical Research, University of Cambridge, Cambridge
CB2 0XY, UK; Department of Medical Genetics, University of Cambridge, Cambridge
CB2 0QQ, UK.
(2)Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità,
Rome 00161, Italy.
(3)Department of Haematology, NHS Blood and Transplant Centre, University of
Cambridge, Cambridge CB2 0XY, UK; NIHR BioResource, Cambridge University
Hospitals NHS Foundation Trust, Cambridge Biomedical Campus, Cambridge CB2 0QQ,
UK.
(4)Department of Medical Genetics, University of Cambridge, Cambridge CB2 0QQ,
UK; European Molecular Biology Laboratory - European Bioinformatics Institute
(EMBL-EBI), Wellcome Genome Campus, Hinxton, Cambridgeshire CB10 1SA, UK.
(5)Microscopy Area, Core Facilities, Istituto Superiore di Sanità, Rome 00161,
Italy.
(6)Department of Medical Genetics, University of Cambridge, Cambridge CB2 0QQ,
UK.
(7)Cambridge Institute for Medical Research, University of Cambridge, Cambridge
CB2 0XY, UK; Department of Pathology, University of Cambridge, Cambridge CB2
1QP, UK.
(8)Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità,
Rome 00161, Italy; Genetics and Rare Diseases Research Division, Ospedale
Pediatrico Bambino Gesù, IRCCS, Rome 00146, Italy.
(9)Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino
Gesù, IRCCS, Rome 00146, Italy.
(10)Genomics England, London, UK.
(11)Fondazione Policlinico Universitario A. Gemelli-IRCCS, Rome 00168, Italy.
(12)Fondazione Policlinico Universitario A. Gemelli-IRCCS, Rome 00168, Italy;
Università Cattolica del Sacro Cuore, Rome 00168, Italy.
(13)Unit of Mechanisms, Biomarkers and Models, Department of Environment and
Health, Istituto Superiore di Sanità, Rome 00161, Italy.
(14)Department of Clinical Genetics, Great Ormond Street Hospital, London WC1N
3JH, UK.
(15)Department of Paediatric Neurology, Cambridge University Hospitals NHS
Foundation Trust, Cambridge CB2 0QQ, UK.
(16)Department of Clinical Genetics, Liverpool Women's Hospital, Liverpool L8
7SS, UK.
(17)Department of Medical Genetics, Guys' and St Thomas' NHS Foundation Trust,
London SE1 9RT, UK.
(18)Clinical Genetics, Birmingham Women's and Children's NHS Foundation Trust,
Birmingham B15 2TG, UK.
(19)Phoenix Children's Hospital, Phoenix, AZ 76109, USA.
(20)Cook Children's Medical Centre, Fort Worth, TX 76104, USA.
(21)Colchester Hospital, East Suffolk and North Essex NHS Foundation Trust,
Essex CO4 5JL, UK.
(22)Ophthalmology Department, Cambridge University Hospitals NHS Foundation
Trust, Cambridge CB2 0QQ, UK.
(23)Department of Neuroscience and Brain Technologies, Istituto Italiano di
Tecnologia, Genova 16163, Italy; Area of Neuroscience, SISSA, Trieste 34136,
Italy.
(24)Genetics and Rare Diseases Research Division, Ospedale Pediatrico Bambino
Gesù, IRCCS, Rome 00146, Italy. Electronic address: [email protected].
(25)Cambridge Institute for Medical Research, University of Cambridge, Cambridge
CB2 0XY, UK; Department of Medical Genetics, University of Cambridge, Cambridge
CB2 0QQ, UK. Electronic address: [email protected]. |
What is the target of a drug pidilizumab? | Pidilizumab is a a humanised monoclonal antibody that targets programmed death-1 pathway. | Author information:
(1)Philippe Armand and Edie Weller, Dana-Farber Cancer Institute; David E.
Avigan, Beth Israel Deaconess Medical Center; Yi-Bin Chen, Massachusetts General
Hospital, Boston, MA; Arnon Nagler, Chaim Sheba Medical Center, Tel-Hashomer;
Reuven Or, Hadassah Medical Center, Jerusalem; Rinat Rotem-Yehudar, CureTech,
Yavne, Israel; Steven M. Devine, The Ohio State University Comprehensive Cancer
Center, Ohio State University, Columbus; Hillard M. Lazarus, Case Western
Reserve University and University Hospitals Case Medical Center, Cleveland, OH;
Mark S. Kaminski, University of Michigan, Ann Arbor, MI; H. Kent Holland,
Northside Hospital; Edmund K. Waller, Winship Cancer Institute, Emory
University, Atlanta, GA; Jane N. Winter and Leo I. Gordon, Robert H. Lurie
Comprehensive Cancer Center, Northwestern University Feinberg School of
Medicine; Stephanie A. Gregory, Rush Medical Center, Chicago, IL; James R.
Mason, Scripps Clinic; Edward D. Ball, Moores University of California at San
Diego Cancer Center, University of California, San Diego; John M. Timmerman and
David Andorsky, University of California, Los Angeles, Los Angeles, CA; Joseph
W. Fay, Baylor Research Institute, Baylor University Medical Center, Dallas;
Chitra M. Hosing, The University of Texas MD Anderson Cancer Center, Houston,
TX; David A. Rizzieri, Duke Cancer Center, Durham, NC; Joseph P. Uberti,
Karmanos Cancer Institute, Detroit, MI; Markus Y. Mapara, University of
Pittsburgh School of Medicine and Cancer Institute, Pittsburgh, PA. BACKGROUND: Endogenous or iatrogenic antitumour immune responses can improve the
course of follicular lymphoma, but might be diminished by immune checkpoints in
the tumour microenvironment. These checkpoints might include effects of
programmed cell death 1 (PD1), a co-inhibitory receptor that impairs T-cell
function and is highly expressed on intratumoral T cells. We did this phase 2
trial to investigate the activity of pidilizumab, a humanised anti-PD1
monoclonal antibody, with rituximab in patients with relapsed follicular
lymphoma.
METHODS: We did this open-label, non-randomised trial at the University of Texas
MD Anderson Cancer Center (Houston, TX, USA). Adult (≥18 years) patients with
rituximab-sensitive follicular lymphoma relapsing after one to four previous
therapies were eligible. Pidilizumab was administered at 3 mg/kg intravenously
every 4 weeks for four infusions, plus eight optional infusions every 4 weeks
for patients with stable disease or better. Starting 17 days after the first
infusion of pidilizumab, rituximab was given at 375 mg/m(2) intravenously weekly
for 4 weeks. The primary endpoint was the proportion of patients who achieved an
objective response (complete response plus partial response according to Revised
Response Criteria for Maligt Lymphoma). Analysis was by intention to treat.
This trial is registered with ClinicalTrials.gov, number NCT00904722.
FINDINGS: We enrolled 32 patients between Jan 13, 2010, and Jan 20, 2012. Median
follow-up was 15.4 months (IQR 10.1-21.0). The combination of pidilizumab and
rituximab was well tolerated, with no autoimmune or treatment-related adverse
events of grade 3 or 4. The most common adverse events of grade 1 were anaemia
(14 patients) and fatigue (13 patients), and the most common adverse event of
grade 2 was respiratory infection (five patients). Of the 29 patients evaluable
for activity, 19 (66%) achieved an objective response: complete responses were
noted in 15 (52%) patients and partial responses in four (14%).
INTERPRETATION: The combination of pidilizumab plus rituximab is well tolerated
and active in patients with relapsed follicular lymphoma. Our results suggest
that immune checkpoint blockade is worthy of further study in follicular
lymphoma.
FUNDING: National Institutes of Health, Leukemia and Lymphoma Society, Cure
Tech, and University of Texas MD Anderson Cancer Center. Significant enthusiasm currently exists for new immunotherapeutic strategies:
blocking the interaction between programmed death-1 receptor on T-cells and
programmed death-ligand 1 on tumor cells to boost immune system stimulation to
fight cancer. Immunomodulation with the antiprogrammed death-1/programmed
death-ligand 1 monoclonal antibodies has shown to mediate tumor shrinkage and
extend overall survival from several pivotal phase I/II studies in melanoma,
renal cell carcinoma, and non-small cell lung cancer. This has prompted multiple
large ongoing phase III trials with the expectation for fast-track FDA approvals
to satisfy unmet medical needs. Compounds targeting the programmed death-1
pathway that are in clinical trials fall into two major categories, namely
antiprogrammed death-1 antibodies: Nivolumab, MK-3475, and pidilizumab; and
antiprogrammed death-ligand 1 antibodies: MPDL3280A, BMS-936559, MEDI4736, and
MSB0010718C. We reviewed the clinical efficacy and safety of each compound based
upon major registered clinical trials and published clinical data. Overall,
response rate of more than 20% is consistently seen across all these trials,
with maximal response of approximately 50% achieved by certain single
antiprogrammed death-1 agents or when used in combination with cytotoxic
T-lymphocyte antigen-4 blockade. The responses seen are early, durable, and have
continued after treatment discontinuation. Immune-related adverse events are the
most common side effects seen in these clinical trials. Overall, the skin and
gastrointestinal tract are the most common organ systems affected by these
compounds while hepatic, endocrine, and neurologic events are less frequent.
These side effects are low grade, manageable, and typically resolve within a
relatively short time frame with a predictable resolution pattern given proper
management. We therefore propose detailed guidelines for management of major
immune-related adverse events that are anticipated with antiprogrammed
death-1/programmed death-ligand 1 therapies based on general experience with
other monoclonal antibodies and the established management algorithms for
immune-related adverse events for cytotoxic T-lymphocyte antigen-4 blockade with
ipilimumab. We anticipate that the antiprogrammed death-1 strategy will become a
viable and crucial clinical strategy for cancer therapy. INTRODUCTION: The programmed death-1 (PD-1) immune checkpoint pathway is an
emerging target in the treatment of hematologic maligcies. Pidilizumab is an
mAb that binds to PD-1 and is a safe and well-tolerated therapy. Recent data
have shown clinical activity utilizing this strategy in diffuse large B-cell
lymphoma (DLBCL).
AREAS COVERED: The role of PD-1 expression in hematologic maligcies is
explored. Recent clinical trials including the results of a Phase I trial in
hematologic maligcies and a Phase II trial of pidilizumab following
autologous hematopoietic stem-cell transplant (AHSCT) are reviewed.
EXPERT OPINION: We review data that suggest that PD-1 is a promising target in
the treatment and management of DLBCL. Changes in immune subsets following
administration of pidilizumab are felt to represent on-target responses. The
improvement in progression-free survival (PFS) following AHSCT supports a
response to therapy. Importantly, the improvement in PFS for patients with
positive FDG-PET/CT following AHSCT indicating residual disease further supports
direct activity of pidilizumab in DLBCL. Immune-regulatory mechanisms are used by cancer to hide from the immune system.
Advances and in-depth understanding of the biology of melanoma and its
interaction with the immune system have led to the development of some of
antagonistic antibodies to the programmed death 1 pathway (PD-1) and one of its
ligands, programmed death ligand 1 (PD-L1), which are demonstrating high
clinical benefit rates and tolerability. Blocking the immune-regulatory
checkpoints that limit T-cell responses to melanoma upon PD-1/PD-L1 modulation
has provided clinically validated targets for cancer immunotherapy. Combinations
with other anti-melanoma agents may result in additional benefits. Nivolumab,
pembrolizumab (formerly known as MK-3475 and lambrolizumab), and pidilizumab are
anti-PD-1 antibodies in clinical development for melanoma, non-small cell lung
cancer, renal cell carcinoma, head and neck cancers, lymphoma, and several other
cancers. Long-term survivors already have been reported with these therapies. In
this review, we discuss the current state of anti-PD-1 agents, the evidence in
the literature to support the combination of anti-PD-1 antibodies with other
anti-cancer agents and discuss the future directions for rational design of
clinical trials that keep on increasing the number of long-term survivors. Advances in understanding of the immune microenvironment have highlighted the
role of immunosuppressive T cell, myeloid, dendritic and monocytic
sub-populations in inhibition of the anti-tumor immune response. The role of B
cells in modulating the immune response to solid tumors as well as lymphoid
maligcies is less well understood. Murine models of autoimmune disease have
defined B regulatory cell (Breg) subsets with immune suppressive activity,
including B cell subsets that express IL-10, and transforming growth factor-β,
which can facilitate T regulatory cell recruitment and expansion. Multiple
murine tumor models point to the existence of similar immune suppressive B cell
sub-populations that can migrate into tumor deposits and acquire an immune
suppressive phenotype, which then leads to attenuation of the local anti-tumor
immune response. Other murine models of viral or chemically induced skin
carcinogenesis have identified a pivotal role for B cells in promoting
inflammation and carcinogenesis. While many human solid tumors demonstrate
significant B cell infiltration and/or tertiary lymphoid structure formation,
the functional properties of tumor-infiltrating B cells and their effects on
immunity are poorly understood. Recent successes in early Phase I/II trials
using anti-checkpoint inhibitor antibodies such as nivolumab or pidilizumab
directed against PD-1 in the setting of Hodgkin's and non-Hodgkin's lymphomas
validate the therapeutic utility of reversing B cell-mediated immune
suppression. Further studies to define Breg subsets, and mechanisms of
suppression, may provide new avenues for modulation of the immune response and
meaningful therapeutic intervention in both lymphoid and solid tumors. PURPOSE OF REVIEW: The development of 'immune checkpoint inhibitors' or drugs
targeting the programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1) axis
has been a stunning success of cancer immunotherapy. This review provides a
timely overview of the biology and function of the PD-1 pathway and discusses
the rationale for therapeutic inhibition of this pathway in lymphoma.
RECENT FINDINGS: Recent studies have evaluated the prevalence and prognostic
implications of PD-1, PD-L1/2 expression in various lymphoma subtypes. We
present an overview of the clinical trials evaluating pidilizumab, nivolumab,
and pembrolizumab in patients with lymphoid maligcies, and highlight some of
the more promising agents in this class, currently in development. Finally, we
discuss biomarkers that may predict response to therapy in patients with
lymphoma across these clinical trials.
SUMMARY: A plethora of clinical trials are in progress testing immune checkpoint
inhibitors in many subtypes of lymphoma, which will define their role both as a
monotherapy and in combination with other biologic agents. BACKGROUND: This meta-analysis has been conducted to determine the risk of
elevated transaminases associated with immune checkpoint inhibitors use in
patients with cancer.
METHODS: Studies eligible for our analysis included randomized Phase II and III
trials of patients with cancer on ipilimumab, nivolumab, pembrolizumab,
tremelimumab and pidilizumab, which describe events of elevated transaminases
[alanine aminotransferase (ALT) and aspartate aminotransferase (AST)].
RESULTS: Initial database search revealed 210 relevant citations. After
excluding noneligible studies, 10 trials were considered eligible for the
quantitative synthesis. The RR of all-grade elevated ALT and AST was 2.36 (95%
CI 1.20-4.66; p = 0.01) and 1.53 (95% CI 0.73-3.22; p = 0.26), respectively,
whereas for high-grade elevated ALT and AST, it was 11.27 (95% CI 5.38-23.63; p
< 0.0001) and 4.9 (95% CI 2.97-8.09; p < 0.0001), respectively.
CONCLUSIONS: Our study has shown that the use of immune checkpoint inhibitors
has a causal relationship to an increased risk of high-grade elevated ALT and
AST. Clinicians using these agents should be attentive of this risk. AIM: We performed a meta-analysis of the risk of selected gastrointestinal
toxicities associated with immune checkpoint inhibitors.
PATIENTS & METHODS: Eligible studies included randomized trials of patients with
solid tumors on ipilimumab, nivolumab, pembrolizumab, tremelimumab, pidilizumab
and atezolizumab, describing events of diarrhea, vomiting or colitis.
RESULTS: After exclusion of ineligible studies, a total of ten clinical trials
were considered eligible for the meta-analysis. The relative risk of all-grade
diarrhea, vomiting and colitis was 1.64 (95% CI: 1.19-2.26; p = 0.002), 0.72
(95% CI: 0.49-1.07; p = 0.1), 10.35 (95% CI: 5.78-18.53; p < 0.00001),
respectively.
CONCLUSION: Our meta-analysis has demonstrated that immune checkpoint inhibitors
are associated with a significantly increased risk of all grade and high-grade
colitis. BACKGROUND: The standard treatment of advanced melanoma has been changing in
recent years. Palliative chemotherapy is being replaced by more efficient
targeted therapies and modern immunotherapies based on antibodies against
checkpoints of the immune response (so called checkpoint inhibitors). Todays
standard ipilimumab (ant-iCTLA 4 antibody) could significantly prolong overall
survival and achieved longterm disease control in about 20% of patients. There
are other perspective immune modulating agents, such as anti-PD 1 antibodies
(nivolumab, pembrolizumab, pidilizumab) and anti-PD L1 antibodies. Unique
mechanism of action is accompanied by new types of immunerelated adverse events.
AIM: The aim of the article is to summarize current knowledge about the toxicity
of these antibodies and propose solutions in routine clinical practice. Diffuse intrinsic pontine glioma (DIPG) is an incurable disease with a median
overall survival of 10 months. Immune modulating antibodies have recently
emerged as a highly promising treatment modality in multiple cancer types. We
present results from the first study to evaluate the immune modulating antibody
MDV9300 (pidilizumab) in pediatric patients with DIPG. All patients aged 3 years
and older, diagnosed with DIPG between February 2014 and June 2015 in Israel,
were offered to participate in the study. Enrolled patients were started on
biweekly 6 mg/kg MDV9300 after radiation completion. Treatment was continued
until disease progression on imaging. Patients were followed biweekly for the
occurrence of neurological deficit toxicities and side effects. Secondary
endpoints were event free survival and overall survival. Of 13 children
diagnosed with DIPG during the study period, nine were enrolled in the study.
The patients underwent radiotherapy and none had chemotherapy. A total of 83
cycles of MDV9300 (range 2-16) were applied. The main side effects were
neutropenia (CTCAE grade 1-3), mild to moderate fatigue, and acute elevation of
blood pressure. Four patients died within 1 year of the diagnosis, another three
died within 2 years and two children are still alive nearly 30 months from
diagnosis, with stable disease. The median event free survival is 9.3 months
(range 6.8-24) and the median overall survival is 15.6 months (range 6.9-28).
Preliminary results demonstrate that MDV9300 treatment is safe and may be
effective in the treatment of children with DIPG. |
List the proteins defining the triple negative breast cancer. | The so called "Triple Negative Breast Cancer" (TNBC) represents approximately 15-20% of breast cancers. This acronym simply means that the tumour does not express oestrogen receptor (ER) and progesterone receptor (PR) and does not exhibit amplification of the human epidermal growth factor receptor 2 (HER2) gene. | BACKGROUND: Triple negative breast cancers (TNBCs) are a diverse and
heterogeneous group of tumors that by definition lack estrogen and progesterone
receptors and amplification of the HER-2 gene. The majority of the tumors
classified as TNBCs are highly maligt, patients are usually young and only a
subgroup of patients responds to conventional chemotherapy with a favorable
prognosis. Various studies have been reported in western literature on TNBCs,
all highlighting the poor prognosis of this subtype. However, extensive data
from India is lacking.
AIM: The aim of this study was to analyze the epidemiological and clinical
profile of TNBCs at our institute.
MATERIALS AND METHODS: This was the retrospective study carried out in Tertiary
Cancer Care Center in South India. Case files of all breast cancer patients were
reviewed from the hospital database registered in 1 year and TNBC patients were
selected for the study. Patient's characteristic, treatment, and histological
features were analyzed.
RESULTS: A total of 322 patients were registered during the period of 1 year and
26% (84/322) of total patients were TNBC. Median age of presentation was 44.5
years. About 94% (79/84) of patients had first full-term delivery before the age
of 30 years. The most common presenting symptom was left sided breast lump.
Locally advanced and early breast cancer (EBC) was 51% (43/84) and 42% (36/84),
respectively. Metastatic breast cancer was seen in five patients. The highest
numbers of patients were node negative disease (36.9%) [31/84], followed by N1
30.95% (26/84). Most of the patients had high-grade tumor. 94% (34/36) of cases
of EBC had undergone upfront modified radical mastectomy. Invasive ductal
carcinoma was the predomit histology except one who had medullary carcinoma.
Twenty-four patients received neoadjuvant chemotherapy (NACT). There was no
pathological complete remission, but all patients responded to NACT. Metastatic
disease was seen in five patients. All patients had bone metastasis.
CONCLUSIONS: TNBCs are highly aggressive subtype, with high grade with limited
treatment options and very poor prognosis. Incidence is more in our country than
the western literature. Even in our country also the incidence is varies in
different region. TNBCs are significantly associated with young aged patients.
There was a lack of association between tumor size and lymph node positivity. The so called "Triple Negative Breast Cancer" (TNBC) represents approximately
15-20% of breast cancers. This acronym simply means that the tumour does not
express oestrogen receptor (ER) and progesterone receptor (PR) and does not
exhibit amplification of the human epidermal growth factor receptor 2 (HER2)
gene. Despite this unambiguous definition, TNBCs are an heterogeneous group of
tumours with just one common clinical feature: a distinctly aggressive nature
with higher rates of relapse and shorter overall survival in the metastatic
setting compared with other subtypes of breast cancer. Because of the absence of
well-defined molecular targets, cytotoxic chemotherapy is currently the only
treatment option for TNBC. In the last decades, the use of more aggressive
chemotherapy has produced a clear improvement of the prognosis in women with
TNBC, but this approach results in an unacceptable deterioration in the quality
of life, also if some support therapies try to relieve patients from distress.
In addition, there is the general belief that it is impossible to further
improve the prognosis of TNBC patients with chemotherapy alone. In view of that,
there is a feverish search for new "clever drugs" able both to rescue
chemo-resistant, and to reduce the burden of chemotherapy in chemo-responsive
TNBC patients. A major obstacle to identifying actionable targets in TNBC is the
vast disease heterogeneity both inter-tumour and intra-tumour and years of study
have failed to demonstrate a single unifying alteration that is targetable in
TNBC. TNBC is considered the subtype that best benefits from the neoadjuvant
model, since the strong correlation between pathological Complete Response and
long-term Disease-Free-Survival in these patients. In this review, we discuss
the recent discoveries that have furthered our understanding of TNBC, with a
focus on the subtyping of TNBC. We also explore the implications of these
discoveries for future treatments and highlight the need for a completely
different type of clinical trials. Triple negative breast cancer (TNBC) is a type of breast cancer (BC) that does
not express the oestrogen and the progesterone receptors and the human epidermal
growth factor receptor type 2 (HER2). Since there are no positive markers to
reliably classify TNBC, these tumours are not yet treated with targeted
therapies. Perhaps for this reason they are the most aggressive form of breast
carcinomas. However, the clinical observation that these patients do not carry a
uniformly dismal prognosis, coupled with data coming from pathology and
epidemiology, suggests that this negative definition is not capturing a single
clinical entity, but several. We critically evaluate this evidence in this
paper, reviewing clinical and epidemiological data and new studies that aim to
subclassify TNBC. Moreover, evidence on the role of tumour infiltrating
lymphocytes (TILs) on TNBC progression, response to chemotherapy and patient
outcome have been published. The heterogeneity, observed even at TILs level,
highlights the idea that TNBC is much more than a single disease with a unique
treatment. The exploration of the immune environment present at the tumour site
could indeed help in answering the question 'How many diseases is TNBC' and will
help to define prognosis and eventually develop new therapies, by stimulating
the immune effector cells or by inhibiting immunological repressor molecules. In
this review, we focus on the prospect of the patient's diverse immune signatures
within the tumour as potential biomarkers and how they could be modulated to
fight the disease. |
Han Wistar and Sprague Dawley are breeds of what laboratory animal? | Han-Wistar and Sprague-Dawley rats | In the present study the pineal gland was examined in 2 outbred stocks and 6
inbred strains of rats some of which were pigmented to varying degrees, to see
whether inbreeding affects the variability and whether differences exist between
albino and pigmented rats. The animals were kept under 12 h light: 12 h darkness
(12 L:12 D) and killed 7 h after the onset of light and darkness, respectively.
The parameters examined were pineal protein content, serotonin and melatonin
levels and hydroxyindole-O-methyltransferase (HIOMT) activity. All the
parameters examined revealed interstrain differences, independently of whether
the data were expressed per pineal or per mg protein. The variation coefficients
for the various parameters were relatively high. They were mostly smaller when
the data were expressed per pineal rather than per mg protein. No striking
differences existed between the variation coefficients in inbred and outbred
rats. When pineal size and the melatonin-related parameters expressed per pineal
were used to assess the melatonin-synthesizing capacity of the pineal glands, it
was found that the outbred Wistar and Sprague-Dawley rats and the inbred
LEWIS-derived (LEW/Han) rats, all of which were albinos, had the most active
pineals. Intermediate activity was noted in the hooded E3/Han and BDE/Han and
the albino BDII/Han rats. The smallest and least active pineals were found in
the totally pigmented BN/Han and DA/Han rats. The results taken together show
that different stocks and strains exhibit significant differences in pineal size
and melatonin-forming capacity. Albino rats appear to have larger and more
active pineals than pigmented rats. Many of the studies conducted to examine the developmental and reproductive
toxicity potential of candidate pharmaceuticals use the Sprague-Dawley rat as
the animal model. This is due in part to the large database for this outbred rat
available for comparison of litter data, and the low incidence of fetal
malformations and variations. The following study was conducted to generate
information on potential embryo-fetal developmental defects and litter data in
another outbred stock of rat, the Wistar Hannover. One hundred fifty pregt
female Wistar Hannover rats (Tac:Glx:WIfBR) were dosed orally once per day with
distilled water from Gestation Days (GD) 6 through 17 covering the time from
implantation to closure of the hard palate (GD0 = day of insemination).
Caesarean sections were performed on Day 20 of gestation. All fetuses were
examined for external, visceral and skeletal malformations and variations.
Macroscopic and histomorphologic examinations were also completed for the F0
females at termination. The percent pregt (88%) and litter size (average
10.6) were found to be lower than that commonly reported for the Sprague-Dawley
rat (Crl:CD (SD)BR; 95.4% and 14.6, respectively). Pre-implantation loss
(14.1%), post-implantation loss (7.4%) and percent resorptions (7.2%) occurred
at a higher incidence than typically seen in the Sprague-Dawley rat (5.9, 5.6
and 5.1%, respectively). The average fetal body weights for both the female and
male rats were lower than those typically seen in the Sprague-Dawley rat.
External, visceral and skeletal examination of the F1 fetuses revealed numerous
malformations and variations which also occurred at higher incidences than those
reported for the Sprague-Dawley rat. Routine macroscopic and histomorphologic
examination showed there were no changes that would be interpreted to have
impaired mating performance, fertility or gestation. Thus, this study provides
information on the reproductive effects and the background incidence of
embryo-fetal development defects that could be used for comparison to those
identified when using this outbred rat for developmental and reproductive
toxicity studies, as well as for comparison to the more commonly used rat stock,
the Sprague-Dawley rat. For the parameters evaluated, the Wistar Hannover rat
had greater variability and an increased incidence of spontaneous malformations
as compared to the Crl:CD (SD)BR Sprague-Dawley rat. These findings should be
considered if this stock of rat is selected in the conduct of developmental and
reproductive toxicity studies. NMR spectroscopic and statistical methods have been applied to investigate the
biochemical variations within and between two phenotypically normal rat strains.
The 600 MHz (1)H NMR spectra of urine were acquired as part of a series of drug
toxicity studies from 450 control rat urine samples from each of two strains of
laboratory rat (Han Wistar and Sprague Dawley). The spectra were data-reduced to
256 intensity descriptors over a range of delta 0.2-10.0. The spectral variation
was analysed both within and between strains in terms of the mean, standard
deviation, skewness and kurtosis of each descriptor. It is demonstrated that
spectral intensities corresponding to a number of endogenous metabolites do not
show Gaussian distributions and there is evidence for bimodality for some
metabolites. Additionally, despite the visual similarity of the NMR spectra from
the two strains of rat, the descriptor distributions and the statistics derived
from them revealed differences in the metabolite profiles, which clearly
distinguished the two populations. This work is of value in the determination of
biochemical normality and variability, and thus can be used to investigate, and
place confidence limits on the biochemical deviations, which arise as a
consequence of genetic modification or pathophysiological events. Time-related changes in the incidences of spontaneous neoplasms in skin (fibroma
and keratoacanthoma), thyroid (C-cell and follicular cell adenomas/carcinomas),
uterus (stromal polyp), testes (Leydig cell tumor) and hemolymphoreticular
system (mesenteric lymph node hemangioma and maligt granular lymphocytic
leukemia) were assessed statistically in Wistar, Sprague-Dawley and F344 rats
employed by the BASF, Germany and major European contract research organizations
over the last 20 years. Negative trends (5 out of 80 cases) were observed for
skin fibromas in F344 males, for follicular cell adenomas in Han Wistar females
and in Sprague-Dawley males and females, and for follicular cell carcinomas in
Sprague-Dawley males. Positive trends (8 out of 80 cases) were observed for skin
keratoacanthomas in Han Wistar males, for C-cell adenomas in BASF Wistar males
and females, for stromal polyps in Han Wistar and Sprague-Dawley females, and
for mesenteric lymph node hemangiomas in Han Wistar and Sprague-Dawley males and
in BASF Wistar females. In 67 out of 80 cases there were no statistically
significant trends. Tumor drift was not common but occurred far more often in
outbred rat strains (Wistar and Sprague-Dawley) than in the inbred rat strain
(F344). This observation suggests that tumor predisposition is genetically
determined, that tumor drift is primarily caused by genetic drift and that
non-genotoxic carcinogens operate by facilitating the expression of tumor
predisposition in target cells. A previous report of this work (Ringeissen et al. 2003) described the use of
nuclear magnetic resoce (NMR) spectroscopy coupled with multivariate
statistical data analysis (MVDA) to identify novel biomarkers of peroxisome
proliferation (PP) in Wistar Han rats. Two potential biomarkers of peroxisome
proliferation in the rat were described, N-methylnicotinamide (NMN) and
N-methyl-4-pyridone-3-carboxamide (4PY). The inference from these results was
that the tryptophan-nicotinamide adenine dinucleotide (NAD(+)) pathway was
altered in correlation with peroxisome proliferation, a hypothesis subsequently
confirmed by TaqMan analysis of the relevant genes encoding two key enzymes in
the pathway, aminocarboxymuconate-semialdehyde decarboxylase (EC 4.1.1.45) and
quinolinate phosphoribosyltransferase (EC 2.4.2.19). The objective of the
present study was to investigate these data further and identify other
metabolites in the NMR spectrum correlating equally with PP. MVDA Partial Least
Squares (PLS) models were constructed that provided a better prediction of PP in
Wistar Han rats than levels of 4PY and NMN alone. The resulting Wistar Han rat
predictive models were then used to predict PP in a test group of Sprague Dawley
rats following administration of fenofibrate. The models predicted the presence
or absence of PP (above on arbitrary threshold of >2-fold mean control) in all
Sprague Dawley rats in the test group. The aims of the present study were to determine cytochrome P450 enzyme activity
in six strains of experimental rodents (n = 5/sex/species): ICR, C57BL/6 and
DBA/2 mice; Sprague Dawley and Wistar rats; and Dunkin Hartley guinea pigs.
After animals were treated with the typical inducers β-naphthoflavone (BNF),
dexamethasone (DEX) and phenobarbital (PB), the levels of O-dealkylation of
ethoxyresorufin (EROD), methoxyresorufin (MROD), pentoxyresorufin (PROD) and
benzyloxyresorufin (BROD) activity were determined using responsive catalytic
reactions to study CYP1A1, CYP1A2 and CYP2B, respectively. A maximal induction
of EROD and MROD was found in BNF-treated animals from all strains (2.4- to
15.1-fold) except DBA/2 (0.9- to 1.8-fold). C57BL/6 mice had the strongest
BNF-induced EROD (15.1-fold) and MROD (8.3-fold) activities. No differences in
BNF-induced EROD and MROD activities were observed between males and females.
However, the EROD activity of Wistar rats and the MROD activity of Sprague
Dawley rats were higher in males than females. DEX induced PROD activity only in
mice (1.3- to 7.1-fold), but not in rats and guinea pigs (0.2- to 1.1-fold).
However, induction of BROD activity was found in DEX-treated mice and rats (1.5
to 12.5-fold), but not in guinea pigs (0.3 to 0.4-fold). PB caused a significant
elevation of PROD (1.7- to 10.4-fold) and BROD (31- to 13.2-fold) activities in
all the animals. PB-induced BROD activity was higher in females than males in
Sprague Dawley rats. These observations strongly suggest that the choice of
experimental animal strain, species and inducer is of critical importance for
studies of drug metabolism and interaction. BACKGROUND: A cross-laboratory analytic evaluation of a commercially available
human inhibin B ELISA for measuring inhibin B in rat serum and plasma has been
undertaken.
METHODS: Dilution linearity, spiked recovery, intra- and inter-assay precision,
functional sensitivity, matrix effects, and frozen stability were assessed
across five laboratories. Reference ranges were generated for male Sprague
Dawley and Han Wistar rats.
RESULTS: Acceptable performance was defined as an overall assay coefficient of
variation ≤ 20% with an intraday LLOQ ≤ 20 pg/ml. Intra- and inter-assay
precision and functional sensitivity (≤6.4 pg/ml) generally met these criteria,
but with occasional evidence of greater variability, particularly at lower
concentrations. Dilution linearity was acceptable with occasional low recovery.
Acceptable recovery of kit calibrators from rat serum confirmed the absence of
matrix effects. Matched serum and plasma samples gave comparable results. The
signal increased on freezing, remained constant for ≥3 freeze-thaw cycles and
was generally stable for at least 8 weeks. Mean inhibin B ranged from 33.5 to
140.6 pg/ml in adult rats across laboratories, with some evidence for a decline
from 6 to 9 weeks of age. Power calculations using preliminary reference range
data indicated 10 animals/group would generally detect a 40% decrease in inhibin
B at AstraZeneca, but laboratories with lower control values would require
larger groups.
CONCLUSIONS: The assay meets the analytical performance criteria; however,
precision at the low end of the standard curve, biological variability, and low
control values observed in some laboratories indicate that the utility of the
assay may be limited in some laboratories. A substantial quantity of data on Sprague-Dawley (SD) and Hannover Wistar rats
strains have been published concerning their source, diet, and housing
conditions, as well as the incidences of nonneoplastic lesions and neoplasms
observed in different laboratories. Differences between the commonly used rat
strains provided by different breeders (i.e., CD (SD) vs. Harlan Sprague-Dawley
strain or Crl: WI(Han) vs. Wistar Hannover (Han)-derived strain, continued
breeding by RCC Ltd., Switzerland, thereafter continued breeding by Harlan) may
include, but are not limited to, body weight, incidence, and onset of major
nonneoplastic lesions and neoplasms, and these can impact the development of a
nonclinical safety program. Fisher 344 (F344) and SD rat strains generally have
the highest tumor incidences, exceeding that in Wistar rats. Certain tumors are
more commonly observed in one strain, and for some, the difference in incidence
may be so significant that the tumor may even be considered characteristic for a
specific strain (e.g., thymoma in Wistar and amphophilic renal adenoma in SD). In 2006 the National Toxicology Program (NTP) of the FDA shifted to the
preferred use of Wistar-Han rats from the more commonly used Sprague-Dawley (SD)
strain - and industry followed. While European laboratories preferred the
Wistar-Han line, there was a paucity of relevant historical control data in many
US research institutions for the new "industry standard" rat strain. In 2010 the
NTP reversed its decision and shifted back to SD rats because of reproductive
issues with the Wistar strain. For post hoc comparative analyses, we report
minimal practical differences in Functional Observational Battery (FOB) data
from a large sample of male and female Wistar-Han and SD rats. In summarizing
data from the preclinical safety evaluations of the CNS effects of new drugs
using the FOB, it is crucial to understand the value of not only how the
functional expression of drug effects in the rat are predictive of the human
response, but also how and why they differ. What we can predict from the
behavioral and physiological response of the designated test system to drug
administration is the foundation of "generalizability" to the human's response.
Here, we conclude that the use of either SD or WH rat strains in standard CNS
safety studies provide equivalent supportive data for CNS safety assessment
required for IND approval under the harmonized guidelines. Knowledge of inter-strain and inter-gender differences in drug metabolism
studies is important for animal selection in pharmacokinetic and toxicological
studies. The effects of rat strain and gender in in vitro metabolism were
investigated in Sprague Dawley (SD) and Wister Han (WH) rats based on the
hepatocyte metabolic profiles of 14 small molecule drugs. Similarities were
found between the hepatocyte metabolic clearances of SD and WH strains,
suggesting that only one strain can be confidently used for the evaluation of
hepatic clearance. Neither strain of rat was preferable over the other to cover
human metabolites. Higher similarities in metabolic pathways were found between
the same gender than the same strain. Differences in metabolite identities,
metabolite formation rates and potential biotransformation pathways were
observed between SD and WH rat strains. Eleven metabolites from six drugs were
"disproportionally" formed between SD and WH rats. The use of a specific rat
strain model and gender for ADME and toxicity testing should, therefore, be
carefully considered as metabolic profiles may differ, even though metabolic
clearance was similar between SD and WH rats. |
Which is the role of the IFIT1 gene in Systemic Lupus Erythematosus (SLE)? | Systemic Lupus Erythematosus (SLE) is caused by a protein called interferon-induced protein with tetratricopeptide repeats 1 (IFIT1). IFIT1 is the first gene described as a candidate gene for SLE, and may function activating Rho proteins through interaction with Rho/Rac guanine nucleotide exchange factor (RHG). | OBJECTIVE: To identify disease-related genes and immune-regulatory pathways in
the pathogenesis of systemic lupus erythematosus (SLE) by using gene expression
profiling and protein-protein interaction analysis.
METHODS: Peripheral white blood cell gene expression profiles of 10 SLE patients
were determined by oligonucleotide microarray analysis. Clustering of the gene
expression profile was compared with the clinical immune phenotype. SLE-induced
genes that were over- or under-expressed were determined and independently
validated using a real-time polymerase chain reaction (PCR) method. To study
their potential function and the possible pathways involved, a candidate gene
was cloned and a GST (glutathione S-transferase) fusion protein was expressed in
Escherichia coli. The fusion protein was further purified using the glutathione
Sepharose 4B system, and was treated as bait to capture prey from SLE peripheral
white blood cell lysate. MALDI-TOF (matrix-assisted laser
desorption/ionization-time-of-flight) mass spectrometry was then performed to
determine the prey protein.
RESULTS: Similarity was found between the gene expression profile and the immune
phenotype clusters of the SLE patients. More than 20 disease-associated genes
were identified, some of which have not been related to SLE previously. Of these
genes, a cluster of interferon-induced genes were highly correlated. IFIT1
(interferon-induced with tetratricopeptide repeats 1) was one of these genes,
and overexpression of its mRNA was confirmed independently by real-time PCR in a
larger population (40 SLE patients and 29 normal controls). An IFIT1 protein-
protein interaction study showed that IFIT1 may interact with Rho/Rac guanine
nucleotide exchange factor.
CONCLUSION: The gene expression profile seems to be the molecular basis of the
diverse immune phenotype of SLE. On the basis of the SLE-related genes found in
this study, we suggest that the interferon-related immune pathway is important
in the pathogenesis of SLE. IFIT1 is the first gene described as a candidate
gene for SLE, and may function by activating Rho proteins through interaction
with Rho/Rac guanine nucleotide exchange factor. IFIT1 and the
interferon-related pathway may provide potential targets for novel interventions
in the treatment of SLE. OBJECTIVE: To investigate the protein-to-protein interaction of
interferon-induced protein with tetratricopeptide repeats 1 (IFIT1), a newly
discovered systemic lupus erythematosus (SLE) related up-regulated gene, and its
possible function.
METHODS: Peripheral blood of 40 SLE patients was obtained to extract total RNA
and synthesized cDNA. Real-time PCR was used to determine the IFIT1 expression
at transcript level. Peripheral blood of another 10 SLE patients was extracted
to obtain specimens of white blood cell lysate. Molecular cloning and a modified
gluthion S-transferase (GST)-pull down assay were used to capture the protein
interacting with IFIT1 in the specimens of white blood cell lysate. MALDI-TOF
mass spectrometry (MS) was preformed to identify the captured protein that could
interact with IFIT1. Twenty-nine sex and age-matched healthy persons were used
as controls.
RESULTS: By real-time PCR showed that the IFIT1Delta Ct value (x +/- s) was
2.344 +/- 1.200 in the SLE patients and was 3.734 +/- 1.274 in the controls (P <
0.001), showing a significant up-regulation in SLE patients. IFIT1 was cloned
and GST-IFIT1 fusion protein was expressed in Escherichia coli. GST-IFIT1 fusion
protein was further purified using Glutathione Sepharose 4B column, and was
treated as bait to capture prey from peripheral white blood cell lysate of SLE
patients. MALDI-TOF MS detected protein interaction between Rho/Rac guanine
nucleotide exchange factor and IFIT1.
CONCLUSION: IFIT1 may interact with Rho/Rac guanine nucleotide exchange factor,
and regulate the activation of Rho/Rac proteins, thus being involved in the
pathogenesis of SLE. Renal damage is the major cause of SLE associated mortality, and IFIT1expression
was elevated in SLE cases in accordance of previous studies. Therefore, we
conducted an animal study to identify the role of IFIT1 expression in renal
pathological changes.18 female MRL/lpr mice and same number of female BALB/c
mice were enrolled in present study. Quantitative analysis of urine protein,
Complement C3 and C4, and anti-ds DNA antibody were conducted. HE and PAS
staining and TEM analysis were employed to observe the pathological changes in
renal tissue. Significant elevation on urine protein and anti-dsDNA and
reduction on Complement C3 and C4 were observed in MRL/lpr mice when comparing
the controls in same age. Staining and TEM analysis observed several
pathological changes in glomerulus among MRL/lpr mice, including cellular
enlargement, basement membrane thickening, and increased cellularcasts. The
linear regression analysis found the optical density of IFIT1 was inversely
associated with F-actin, Nephrin, and Podocin, but not Synatopodin. In summary,
IFIT1 expression is associated with podocytes damage, and capable of suppressing
some proteins essential to glomerular filtration. |
Has tocilizumab been assessed against Covid-19? | Preliminary clinical results have indicated that tocilizumab, can improve the outcomes of patients with severe or critical COVID-19 while maintaining a good safety profile. | |
Variants in which genes cause nonsyndromic retinal degeneration? | Variants in DYNC2H1, IFT81, USH2A and ABHD12 can cause nonsyndromic retinal degeneration. | OBJECTIVE: To identify the genetic causes underlying autosomal recessive
retinitis pigmentosa (arRP) and to describe the associated phenotype.
DESIGN: Case series.
PARTICIPANTS: Three hundred forty-seven unrelated families affected by arRP and
33 unrelated families affected by retinitis pigmentosa (RP) plus noncongenital
and progressive hearing loss, ataxia, or both, respectively.
METHODS: A whole exome sequencing (WES) analysis was performed in 2 families
segregating arRP. A mutational screening was performed in 378 additional
unrelated families for the exon-intron boundaries of the ABHD12 gene. To
establish a genotype-phenotype correlation, individuals who were homozygous or
compound heterozygotes of mutations in ABHD12 underwent exhaustive clinical
examinations by ophthalmologists, neurologists, and otologists.
MAIN OUTCOME MEASURES: DNA sequence variants, best-corrected visual acuity,
visual field assessments, electroretinogram responses, magnetic resoce
imaging, and audiography.
RESULTS: After a WES analysis, we identified 4 new mutations (p.Arg107Glufs*8,
p.Trp159*, p.Arg186Pro, and p.Thr202Ile) in ABHD12 in 2 families (RP-1292 and
W08-1833) previously diagnosed with nonsyndromic arRP, which cosegregated with
the disease among the family members. Another homozygous mutation (p.His372Gln)
was detected in 1 affected individual (RP-1487) from a cohort of 378 unrelated
arRP and syndromic RP patients. After exhaustive clinical examinations by
neurologists and otologists, the 4 affected members of the RP-1292 had no
polyneuropathy or ataxia, and the sensorineural hearing loss and cataract were
attributed to age or the normal course of the RP, whereas the affected members
of the families W08-1833 and RP-1487 showed clearly symptoms associated with
polyneuropathy, hearing loss, cerebellar ataxia, RP, and early-onset cataract
(PHARC) syndrome.
CONCLUSIONS: Null mutations in the ABHD12 gene lead to PHARC syndrome, a
neurodegenerative disease including polyneuropathy, hearing loss, cerebellar
ataxia, RP, and early-onset cataract. Our study allowed us to report 5 new
mutations in ABHD12. This is the first time missense mutations have been
described for this gene. Furthermore, these findings are expanding the spectrum
of phenotypes associated with ABHD12 mutations ranging from PHARC syndrome to a
nonsyndromic form of retinal degeneration. Defects in USH2A cause both isolated retinal disease and Usher syndrome (ie,
retinal disease and deafness). To gain insights into isolated/nonsyndromic USH2A
retinopathy, we screened USH2A in 186 probands with recessive retinal disease
and no hearing complaint in childhood (discovery cohort) and in 84 probands with
recessive retinal disease (replication cohort). Detailed phenotyping, including
retinal imaging and audiological assessment, was performed in individuals with
two likely disease-causing USH2A variants. Further genetic testing, including
screening for a deep-intronic disease-causing variant and large
deletions/duplications, was performed in those with one likely disease-causing
change. Overall, 23 of 186 probands (discovery cohort) were found to harbour two
likely disease-causing variants in USH2A. Some of these variants were
predomitly associated with nonsyndromic retinal degeneration ('retinal
disease-specific'); these included the common c.2276 G>T, p.(Cys759Phe) mutation
and five additional variants: c.2802 T>G, p.(Cys934Trp); c.10073 G>A,
p.(Cys3358Tyr); c.11156 G>A, p.(Arg3719His); c.12295-3 T>A; and c.12575 G>A,
p.(Arg4192His). An allelic hierarchy was observed in the discovery cohort and
confirmed in the replication cohort. In nonsyndromic USH2A disease, retinopathy
was consistent with retinitis pigmentosa and the audiological phenotype was
variable. USH2A retinopathy is a common cause of nonsyndromic recessive retinal
degeneration and has a different mutational spectrum to that observed in Usher
syndrome. The following model is proposed: the presence of at least one 'retinal
disease-specific' USH2A allele in a patient with USH2A-related disease results
in the preservation of normal hearing. Careful genotype-phenotype studies such
as this will become increasingly important, especially now that high-throughput
sequencing is widely used in the clinical setting. PURPOSE: IFT81, a core component of the IFT-B complex, involved in the
bidirectional transport of ciliary proteins, has been recently implicated in
syndromic ciliopathies. However, none of the IFT-B core complex proteins have
been associated with nonsyndromic retinal dystrophies. Given the importance of
ciliary transport in photoreceptor function and structural maintece, we
sought to investigate the impact of IFT (intraflagellar transport) mutations in
nonsyndromic retinopathies.
METHODS: Whole exome sequencing was performed on 50 cone-rod dystrophy (CRD)
patients that were previously screened for mutations in known retinal disease
genes. The impact of candidate mutation was studied using in vitro cell system
and in vivo zebrafish assay to determine the pathogenicity of the variant.
RESULTS: Compound heterozygous mutations in IFT81, including one nonsense
(c.1213C>T, p.R405*) and one missense variant (c.1841T>C, p.L614P), were
identified in a nonsyndromic CRD proband. Extensive functional analyses of the
missense variant in cell culture and zebrafish strongly suggests its pathogenic
nature. Loss of IFT81 impairs ciliogenesis and, interestingly, the missense
variant displayed significantly reduced rescue of ciliogenesis in the IFT81
knockdown in vitro system. Consistently, dramatic reduction of rescue efficiency
of the ift81 mutant zebrafish embryo by mRNA with the missense variant was
observed, further supporting its pathogenicity.
CONCLUSIONS: Consistent with the function of the IFT-B complex in the
maintece of photoreceptor cilium, we report a case of mutations in a core
IFT-B protein, IFT81. This represents the first report of mutations in IFT81 as
a candidate gene for nonsyndromic retinal dystrophy, hence expanding the
phenotype spectrum of IFT-B components. |
Describe the mechanism of action of Omecamtiv Mecarbil. | Omecamtiv mecarbil is a novel, selective cardiac myosin activator that has been shown to improve cardiac function and to decrease ventricular volumes, heart rate, and N-terminal pro-B-type natriuretic peptide in patients with heart failure. | Decreased cardiac contractility is a central feature of systolic heart failure.
Existing drugs increase cardiac contractility indirectly through signaling
cascades but are limited by their mechanism-related adverse effects. To avoid
these limitations, we previously developed omecamtiv mecarbil, a small-molecule,
direct activator of cardiac myosin. Here, we show that it binds to the myosin
catalytic domain and operates by an allosteric mechanism to increase the
transition rate of myosin into the strongly actin-bound force-generating state.
Paradoxically, it inhibits adenosine 5'-triphosphate turnover in the absence of
actin, which suggests that it stabilizes an actin-bound conformation of myosin.
In animal models, omecamtiv mecarbil increases cardiac function by increasing
the duration of ejection without changing the rates of contraction. Cardiac
myosin activation may provide a new therapeutic approach for systolic heart
failure. BACKGROUND: Omecamtiv mecarbil (OM) is a novel inotropic agent that prolongs
systolic ejection time and increases ejection fraction through myosin ATPase
activation. We hypothesized that a potentially favorable energetic effect of
unloading the left ventricle, and thus reduction of wall stress, could be
counteracted by the prolonged contraction time and ATP-consumption.
METHODS AND RESULTS: Postischemic left ventricular dysfunction was created by
repetitive left coronary occlusions in 7 pigs (7 healthy pigs also included). In
both groups, systolic ejection time and ejection fraction increased after OM
(0.75 mg/kg loading for 10 minutes, followed by 0.5 mg/kg/min continuous
infusion). Cardiac efficiency was assessed by relating myocardial oxygen
consumption to the cardiac work indices, stroke work, and pressure-volume area.
To circumvent potential neurohumoral reflexes, cardiac efficiency was
additionally assessed in ex vivo mouse hearts and isolated myocardial
mitochondria. OM impaired cardiac efficiency; there was a 31% and 23% increase
in unloaded myocardial oxygen consumption in healthy and postischemic pigs,
respectively. Also, the oxygen cost of the contractile function was increased by
63% and 46% in healthy and postischemic pigs, respectively. The increased
unloaded myocardial oxygen consumption was confirmed in OM-treated mouse hearts
and explained by an increased basal metabolic rate. Adding the myosin ATPase
inhibitor, 2,3-butanedione monoxide abolished all surplus myocardial oxygen
consumption in the OM-treated hearts.
CONCLUSIONS: Omecamtiv mecarbil, in a clinically relevant model, led to a
significant myocardial oxygen wastage related to both the contractile and
noncontractile function. This was mediated by that OM induces a continuous
activation in resting myosin ATPase. BACKGROUND AND PURPOSE: Omecamtiv mecarbil (OM) is a novel cardiac myosin
activator drug for inotropic support in systolic heart failure. Here we have
assessed the concentration-dependent mechanical effects of OM in permeabilized
cardiomyocyte-sized preparations and single skeletal muscle fibres of
Wistar-Kyoto rats under isometric conditions.
EXPERIMENTAL APPROACHES: Ca2+ -dependent active force production (Factive ), its
Ca2+ sensitivity (pCa50 ), the kinetic characteristics of Ca2+ -regulated
activation and relaxation, and Ca2+ -independent passive force (Fpassive ) were
monitored in Triton X-100-skinned preparations with and without OM (3nM-10 μM).
KEY RESULTS: In permeabilized cardiomyocytes, OM increased the Ca2+ sensitivity
of force production (ΔpCa50 : 0.11 or 0.34 at 0.1 or 1 μM respectively). The
concentration-response relationship of the Ca2+ sensitization was bell-shaped,
with maximal effects at 0.3-1 μM OM (EC50 : 0.08 ± 0.01 μM). The kinetics of
force development and relaxation slowed progressively with increasing OM
concentration. Moreover, OM increased Fpassive in the cardiomyocytes with an
apparent EC50 value of 0.26 ± 0.11 μM. OM-evoked effects in the diaphragm muscle
fibres with intrinsically slow kinetics were largely similar to those in
cardiomyocytes, while they were less apparent in muscle fibres with fast
kinetics.
CONCLUSIONS AND IMPLICATIONS: OM acted as a Ca2+ -sensitizing agent with a
downstream mechanism of action in both cardiomyocytes and diaphragm muscle
fibres. The mechanism of action of OM is connected to slowed
activation-relaxation kinetics and at higher OM concentrations increased
Fpassive production. Heart failure became a leading cause of mortality in the past few decades with a
progressively increasing prevalence. Its current therapy is restricted largely
to the suppression of the sympathetic activity and the renin-angiotensin system
in combination with diuretics. This restrictive strategy is due to the potential
long-term adverse effects of inotropic agents despite their effective influence
on cardiac function when employed for short durations. Positive inotropes
include inhibitors of the Na+/K+ pump, β-receptor agonists, and
phosphodiesterase inhibitors. Theoretically, Ca2+ sensitizers may also increase
cardiac contractility without resulting in Ca2+ overload; nevertheless, their
mechanism of action is frequently complicated by other pleiotropic effects.
Recently, a new positive inotropic agent, the myosin activator omecamtiv
mecarbil, has been developed. Omecamtiv mecarbil binds directly to β-myosin
heavy chain and enhances cardiac contractility by increasing the number of the
active force-generating cross-bridges, presumably without major off-target
effects. This review focuses on recent in vivo and in vitro results obtained
with omecamtiv mecarbil, and discusses its mechanism of action at a molecular
level. Based on clinical data, omecamtiv mecarbil is a promising new tool in the
treatment of systolic heart failure. The small molecule drug omecamtiv mecarbil (OM) specifically targets cardiac
muscle myosin and is known to enhance cardiac muscle performance, yet its impact
on human cardiac myosin motor function is unclear. We expressed and purified
human β-cardiac myosin subfragment 1 (M2β-S1) containing a C-terminal Avi tag.
We demonstrate that the maximum actin-activated ATPase activity of M2β-S1 is
slowed more than 4-fold in the presence of OM, whereas the actin concentration
required for half-maximal ATPase was reduced dramatically (30-fold). We find OM
does not change the overall actin affinity. Transient kinetic experiments
suggest that there are two kinetic pathways in the presence of OM. The domit
pathway results in a slow transition between actomyosin·ADP states and increases
the time myosin is strongly bound to actin. However, OM also traps a population
of myosin heads in a weak actin affinity state with slow product release. We
demonstrate that OM can reduce the actin sliding velocity more than 100-fold in
the in vitro motility assay. The ionic strength dependence of in vitro motility
suggests the inhibition may be at least partially due to drag forces from weakly
attached myosin heads. OM causes an increase in duty ratio examined in the
motility assay. Experiments with permeabilized human myocardium demonstrate that
OM increases calcium sensitivity and slows force development (ktr) in a
concentration-dependent manner, whereas the maximally activated force is
unchanged. We propose that OM increases the myosin duty ratio, which results in
enhanced calcium sensitivity but slower force development in human myocardium. Omecamtiv mecarbil is a selective, small-molecule activator of cardiac myosin
that is being developed as a potential treatment for heart failure with reduced
ejection fraction. Here we determine the crystal structure of cardiac myosin in
the pre-powerstroke state, the most relevant state suggested by kinetic studies,
both with (2.45 Å) and without (3.10 Å) omecamtiv mecarbil bound. Omecamtiv
mecarbil does not change the motor mechanism nor does it influence myosin
structure. Instead, omecamtiv mecarbil binds to an allosteric site that
stabilizes the lever arm in a primed position resulting in accumulation of
cardiac myosin in the primed state prior to onset of cardiac contraction, thus
increasing the number of heads that can bind to the actin filament and undergo a
powerstroke once the cardiac cycle starts. The mechanism of action of omecamtiv
mecarbil also provides insights into uncovering how force is generated by
molecular motors.Omecamtiv mecarbil (OM) is a cardiac myosin activator that is
currently in clinical trials for heart failure treatment. Here, the authors give
insights into its mode of action and present the crystal structure of OM bound
to bovine cardiac myosin, which shows that OM stabilizes the pre-powerstroke
state of myosin. BACKGROUND: Clinical treatment of heart failure is still suffering from limited
efficacy and unfavorable side effects. The recently developed group of agents,
the myosin motor activators, act directly on cardiac myosin resulting in an
increased force generation and prolongation of contraction. The lead molecule,
omecamtiv mecarbil is now in human 3 stage. In addition to the promising
clinical data published so far, there are new in vitro results indicating that
the effect of omecamtiv mecarbil on contractility is rate-dependent.
Furthermore, omecamtiv mecarbil was shown to activate cardiac ryanodine
receptors, an effect that may carry proarrhythmic risk.
METHODS: These new results, together with the controversial effects of the drug
on cardiac oxygen consumption, are critically discussed in this review in light
of the current literature on omecamtiv mecarbil.
RESULTS: In therapeutically relevant concentrations the beneficial inotropic
effect of the agent is not likely affected by these new results - in accordance
with the good clinical data. At supratherapeutic concentrations, however,
activation of cardiac ryanodine receptors may increase arrhythmia propensity,
and the stronger effect on diastolic than systolic cell shortening, observed at
higher pacing frequencies, may decrease or offset the inotropic effect of
omecamtiv mecarbil.
CONCLUSION: Further studies with definitely supratherapeutical concentrations of
omecamtiv mecarbil should be designed to map the actual risk of these
potentially harmful side-effects. Heart failure continues to be a major global health problem with a pronounced
impact on morbidity and mortality and very limited drug treatment options
especially with regard to inotropic therapy. Omecamtiv mecarbil is a
first-in-class cardiac myosin activator, which increases the proportion of
myosin heads that are tightly bound to actin and creates a force-producing state
that is not associated with cytosolic calcium accumulation. Phase I and phase II
studies have shown that it is safe and well tolerated. It produces
dose-dependent increases in systolic ejection time (SET), stroke volume (SV),
left ventricular ejection fraction (LVEF), and fractional shortening. In the
ATOMIC-AHF trial, intravenous (IV) omecamtiv mecarbil did not improve dyspnoea
overall but may have improved it in a high-dose group of acute heart failure
patients. It did, however, increase SET, decrease left ventricular end-systolic
diameter, and was well tolerated. The COSMIC-HF trial showed that a
pharmacokinetic-based dose-titration strategy of oral omecamtiv mecarbil
improved cardiac function and reduced ventricular diameters compared to placebo
and had a similar safety profile. It also significantly reduced plasma
N-terminal-pro B-type natriuretic peptide compared with placebo. The GALACTIC-HF
trial is now underway and will compare omecamtiv mecarbil with placebo when
added to current heart failure standard treatment in patients with chronic heart
failure and reduced LVEF. It is expected to be completed in January 2021. The
ongoing range of preclinical and clinical research on omecamtiv mecarbil will
further elucidate its full range of pharmacological effects and its clinical
usefulness in heart failure. Clinical treatment of heart failure is still not fully solved. A novel class of
agents, the myosin motor activators, acts directly on cardiac myosin resulting
in an increased force generation and prolongation of contraction. Omecamtiv
mecarbil, the lead molecule of this group, is now in human phase 3 displaying
promising clinical performance. However, omecamtiv mecarbil is not selective to
myosin, because it readily binds to and activates cardiac ryanodine receptors
(RyR-2), an effect that may cause complications in case of overdose. In this
study, in silico analysis was performed to investigate the docking of omecamtiv
mecarbil and other structural analogues to cardiac myosin heavy chain and RyR-2
to select the structure that has a higher selectivity to myosin over RyR-2. In
silico docking studies revealed that omecamtiv mecarbil has comparable affinity
to myosin and RyR-2: the respective Kd values are 0.60 and 0.87 μmol/L. Another
compound, CK-1032100, has much lower affinity to RyR-2 than omecamtiv mecarbil,
while it still has a moderate affinity to myosin. It was concluded that further
research starting from the chemical structure of CK-1032100 may result a better
myosin activator burdened probably less by the RyR-2 binding side effect. It
also is possible, however, that the selectivity of omecamtiv mecarbil to myosin
over RyR-2 cannot be substantially improved, because similar moieties seem to be
responsible for the high affinity to both myosin and RyR-2. Heart failure is a life-threatening condition that occurs when the heart muscle
becomes weakened and cannot adequately circulate blood and nutrients around the
body. Omecamtiv mecarbil (OM) is a compound that has been developed to treat
systolic heart failure via targeting the cardiac myosin heavy chain to increase
myocardial contractility. Biophysical and biochemical studies have found that OM
increases calcium (Ca2+) sensitivity of contraction by prolonging the myosin
working stroke and increasing the actin-myosin cross-bridge duty ratio. Most
in vitro studies probing the effects of OM on cross-bridge kinetics and muscle
force production have been conducted at subphysiological temperature, even
though temperature plays a critical role in enzyme activity and cross-bridge
function. Herein, we used skinned, ventricular papillary muscle strips from rats
to investigate the effects of [OM] on Ca2+-activated force production,
cross-bridge kinetics, and myocardial viscoelasticity at physiological
temperature (37°C). We find that OM only increases myocardial contractility at
submaximal Ca2+ activation levels and not maximal Ca2+ activation levels. As
[OM] increased, the kinetic rate constants for cross-bridge recruitment and
detachment slowed for both submaximal and maximal Ca2+-activated conditions.
These findings support a mechanism by which OM increases cardiac contractility
at physiological temperature via increasing cross-bridge contributions to
thin-filament activation as cross-bridge kinetics slow and the duration of
cross-bridge attachment increases. Thus, force only increases at submaximal Ca2+
activation due to cooperative recruitment of neighboring cross-bridges, because
thin-filament activation is not already saturated. In contrast, OM does not
increase myocardial force production for maximal Ca2+-activated conditions at
physiological temperature because cooperative activation of thin filaments may
already be saturated. A novel myosin activator, omecamtiv mecarbil (OM), is a cardiac inotropic agent
with a unique new mechanism of action, which is thought to arise from an
increase in the transition rate of myosin into the actin-bound force-generating
state without increasing calcium (Ca2+) transient. There remains, however,
considerable controversy about the effects of OM on cardiac contractility and
energy expenditure. In the present study, we investigated the effects of OM on
left ventricular (LV) mechanical work and energetics, i.e., mechanoenergetics in
rat normal hearts (CTL) and failing hearts induced by chronic administration of
isoproterenol (1.2 mg/kg/day) for 4 weeks (ISO-HF). We analyzed the LV
end-systolic pressure-volume relation (ESPVR) and the linear relation between
the myocardial oxygen consumption per beat (VO2) and systolic pressure-volume
area (PVA; a total mechanical energy per beat) in isovolumically contracting rat
hearts at 240- or 300-bpm pacing in the absence or presence of OM. OM did not
change the ESPVR in CTL and ISO-HF. OM, however, significantly decreased the
slope of VO2-PVA relationship in both CTL and ISO-HF, and significantly
increased the mean VO2 intercept without changes in basal metabolism in ISO-HF.
These results suggested that OM improved the oxygen cost of PVA (contractile
efficiency) with the unchanged LV contractility in both CTL and ISO-HF but
increased VO2 for Ca2+ handling in excitation-contraction (E-C) coupling in
ISO-HF. We concluded that OM improves contractile efficiency in normal and
failing hearts but increases O2 consumption of Ca2+ handling in failing hearts
in isovolumically contracting rat model. A central factor in the pathogenesis of heart failure (HF) with reduced ejection
fraction is the initial decrease in systolic function. Prior attempts at
increasing cardiac contractility with oral drugs have uniformly resulted in
signals of increased mortality at pharmacologically effective doses. Omecamtiv
mecarbil is a novel, selective cardiac myosin activator that has been shown to
improve cardiac function and to decrease ventricular volumes, heart rate, and
N-terminal pro-B-type natriuretic peptide in patients with chronic HF. The
GALACTIC-HF (Global Approach to Lowering Adverse Cardiac outcomes Through
Improving Contractility in Heart Failure) trial tests the hypotheses that
omecamtiv mecarbil can safely improve symptoms, prevent clinical HF events, and
delay CV death in patients with chronic HF. The GALACTIC-HF trial is an
international, multicenter, randomized, double-blind, placebo-controlled,
event-driven cardiovascular outcomes trial. More than 8,000 patients with
chronic symptomatic (New York Heart Association functional class II to IV) HF,
left ventricular ejection fraction ≤35%, elevated natriuretic peptides, and
either current hospitalization for HF or history of hospitalization or emergency
department visit for HF within a year of screening will be randomized to either
oral placebo or omecamtiv mecarbil employing a pharmacokinetic-guided dose
titration strategy using doses of 25, 37.5, or 50 mg twice daily. The primary
efficacy outcome is the time to cardiovascular death or first HF event. The
study has 90% power to assess a final hazard ratio of approximately 0.80 in
cardiovascular death, the first secondary outcome. The GALACTIC-HF trial is the
first trial examining whether selectively increasing cardiac contractility in
patients with HF with reduced ejection fraction will result in improved clinical
outcomes. (Registrational Study With Omecamtiv Mecarbil/AMG 423 to Treat Chronic
Heart Failure With Reduced Ejection Fraction [GALACTIC-HF]; NCT02929329). Omecamtiv mecarbil (OM), an activator of cardiac myosin, strongly affects
contractile characteristics of the ventricles and, to a much lesser extent, the
characteristics of atrial contraction. We compared the molecular mechanism of
action of OM on the interaction of atrial and ventricular myosin with actin
using an optical trap and an in vitro motility assay. In concentrations up to
0.5 μM, OM did not affect the step size of a myosin molecule but reduced it at a
higher OM level. OM substantially prolonged the interaction of both isoforms of
myosin with actin. However, the interaction characteristics of ventricular
myosin with actin were more sensitive to OM than those of atrial myosin. Our
results, obtained at the level of isolated proteins, can explain why the impact
of OM in therapeutic concentrations on the contractile function of the atrium is
less significant as compared to those of the ventricle. Omecamtiv mecarbil (OM) is a selective cardiac myosin activator (myotrope),
currently in Phase 3 clinical investigation as a novel treatment for heart
failure with reduced ejection fraction. OM increases cardiac contractility by
enhancing interaction between myosin and actin in a calcium-independent fashion.
This study aims to characterize the mechanism of action by evaluating its
simultaneous effect on myocyte contractility and calcium-transients (CTs) in
healthy canine ventricular myocytes. Left ventricular myocytes were isolated
from canines and loaded with Fura-2 AM. With an IonOptix system, contractility
parameters including amplitude and duration of sarcomere shortening, contraction
and relaxation velocity, and resting sarcomere length were measured. CT
parameters including amplitude at systole and diastole, velocity at systole and
diastole, and duration at 50% from peak were simultaneously measured. OM was
tested at 0.03, 0.1, 0.3, 1, and 3 µmol\L concentrations to simulate therapeutic
human plasma exposure levels. OM and isoproterenol (ISO) demonstrated
differential effects on CTs and myocyte contractility. OM increased
contractility mainly by prolonging duration of contraction while ISO increased
contractility mainly by augmenting the amplitude of contraction. ISO increased
the amplitude and velocity of CT, shortened duration of CT concurrent with
increasing myocyte contraction, while OM did not change the amplitude, velocity,
and duration of CT up to 1 µmol\L. Decreases in relaxation velocity and
increases in duration were present only at 3 µmol\L. In this translational
myocyte model study, therapeutically relevant concentrations of OM increased
contractility but did not alter intracellular CTs, a mechanism of action
distinct from traditional calcitropes. Left ventricular systolic dysfunction is the hallmark pathology in heart failure
with reduced ejection fraction. Increasing left ventricular contractility with
beta-adrenergic receptor agonists, phosphodiesterase-3 inhibitors, or
levosimendan has failed to improve clinical outcomes and, in some situations,
increased the risk of sudden cardiac death. Beta-adrenergic receptor agonists
and phosphodiesterase-3 inhibitors retain an important role in advanced heart
failure. Thus, there remains an unmet need for safe and effective therapies to
improve left ventricular systolic function. Two novel cardiac myotropes,
omecamtiv mecarbil and danicamtiv, target cardiac myosin to increase left
ventricular systolic performance. Neither omecamtiv mecarbil nor danicamtiv
affects cardiomyocyte calcium handling, the proposed mechanism underlying the
life-threatening arrhythmias associated with cardiac calcitropes and calcium
sensitizers. Phase 2 clinical trials have demonstrated that these cardiac myosin
activators prolong left ventricular systolic ejection time and promote left
ventricular and atrial reverse remodeling. At higher plasma concentrations,
these agents may be associated with myocardial ischemia and impaired diastolic
function. An ongoing phase 3 clinical trial will estimate the clinical efficacy
and safety of omecamtiv mecarbil. An additional study of these agents, which
have minimal hemodynamic and renal effects, is warranted in patients with
advanced heart failure refractory to guideline-directed neurohormonal blockers. BACKGROUND: Chronic heart failure with reduced ejection fraction impairs
health-related quality of life (HRQL). Omecamtiv mecarbil (OM)-a novel activator
of cardiac myosin-improves left ventricular systolic function and remodeling and
reduces natriuretic peptides. We sought to evaluate the effect of OM on symptoms
and HRQL in patients with chronic heart failure with reduced ejection fraction
and elevated natriuretic peptides enrolled in the COSMIC-HF trial (Chronic Oral
Study of Myosin Activation to Increase Contractility in Heart Failure).
METHODS: Patients (n=448) were randomized 1:1:1 to placebo, 25 mg of OM BID, or
to pharmacokinetically guided dose titration (OM-PK) for 20 weeks. The Kansas
City Cardiomyopathy Questionnaire was administered to assess HRQL at baseline,
16 weeks, and 20 weeks. The primary scores of interest were the Total Symptom
Score, Physical Limitation Scale, and Clinical Summary Score.
RESULTS: Mean change in score from baseline to 20 weeks for the Total Symptom
Score was 5.0 (95% CI, 1.8-8.1) for placebo, 6.6 (95% CI, 3.4-9.8) for OM 25 mg
(P=0.32 versus placebo), and 9.9 (95% CI, 6.7-13.0) for OM-PK (P=0.03 versus
placebo); for the Physical Limitation Scale, it was 3.1 for placebo (95% CI,
-0.3 to 6.6), 6.0 (95% CI, 3.1-8.9) for OM 25 mg (P=0.12), and 4.3 (95% CI,
0.7-7.9) for OM-PK (P=0.42); for the Clinical Summary Score, it was 4.1 (95% CI,
1.4-6.9) for placebo, 6.3 (95% CI, 3.6-9.0) for OM 25 mg (P=0.19), and 7.0 (95%
CI, 4.1-10.0) for OM-PK (P=0.14). Differences between OM and placebo were
greater in patients who were more symptomatic at baseline.
CONCLUSIONS: HRQL as measured by the Total Symptom Score improved in patients
with heart failure with reduced ejection fraction assigned to the OM-PK group
relative to placebo. Ongoing trials are prospectively testing whether OM
improves symptoms and HRQL in heart failure with reduced ejection fraction.
Registration: URL: https://www.clinicaltrials.gov; Unique identifier:
NCT01786512. Collaborators: Teerlink JR, Díaz R, Santa Fe R, Felker GM, McMurray JJV, Metra
M, Solomon SD, Malik FI, Kurtz CE, Abbasi S, Knusel B, Hucko T, Groarke J,
Honarpour N, Legg JC, Sharpsten L, Varin C, Konstam MA, Butler J, Dargie H,
Massaro J, Greenberg BH, Januzzi JL, Lesko LJ, Upshaw JN, Lopes R, Jones WS,
Alexander KP, Al-Khatib SM, Harrison RW, Jordan JD, Kong DF, Mathews R, McGarrah
RW, Metha RH, Melloni C, Povsic TJ, Shah S, Claggett B, Adams K, Ad I, Arias
Mendoza A, Biering-Sørensen T, Böhm M, Bonderman D, Cleland J, Corbalan Herreros
R, Crespo Leiro M, Dahlstrom U, Diaz R, Echeverria LE, Fang J, Filippatos G,
Fonseca C, Goncalvesova E, Goudev A, Howlett J, Jiang L, Lanfear D, Li J, Lund
M, Mareev V, Momomura SI, O'Meara E, Parkhomenko A, Ponikowski P, Ramires F,
Serpytis P, Sliwa K, Spinar J, Suter T, Tomcsanyi J, Vandekerckhove H, Vinereanu
D, Davila C, Voors A, Yilmaz MB, Zannad F, Besada DA, Majul CR, Litvak Bruno MR,
Sassone S, Avaca HA, Rasmussen M, Aiub JR, Hominal MA, Perna E, Garcia Duran RO,
Schiavi L, Lobo Marquez LL, Gomez Vilamajo OA, Mackinnon I, Leon de la Fuente
RA, Montana OR, Novaretto L, Ahuad Guerrero RA, Garcia Brasca D, Prado A,
Garrido MA, Luquez H, Martinez DF, Nicolosi L, Parody ML, Zaidman C, Colombo
Berra F, Ibañez J, Zapata G, Amuchastegui M, Caccavo A, Colque R, Diez M, Poy C,
Salomone OA, Vogel D, Bordonava AP, Ferdez A, French J, Atherton J, Hamilton
A, Begg A, Abhayaratna W, Judkins C, Macdonald P, De Pasquale C, McKenzie S,
Amerena J, Szto G, Kearney L, Zimmet H, Sverdlov A, Beltrame J, Korczyk D,
Sindone A, Moertl D, Huber K, Huelsmann M, Jakl-Kotauschek G, Ablasser K,
Fruhwald F, Ebner C, Siostrzonek P, Drexel H, Poelzl G, Dujardin K, Dupont M,
Buysschaert I, Lancellotti P, Pierard L, Droogne W, Chouchane I, Silveira F,
Rassi S, Reis G, Pimentel Filho P, Simoes MV, Braga JC, Giorgeto FE, Duda N,
Ferraz A, Pederneiras Jaeger C, Rech RL, Saraiva JF, Tognon A, Cardoso J, Greco
O, Sanali Paiva M, Paolino B, Coelho Filho O, Nigro Maia L, da Silva R, Canesin
M, Ferreira Rossi PR, Zytynski Moura LA, Ribas Fortes JA, Cerci RJ, Ferdes
Manenti ER, Leaes PE, Beck da Silva Neto L, Barroso de Souza WK, Bacal F, Chaves
R, Vidotti MH, Brito F, Melo de Barros E Silva PG, Soares Piegas L, Todorov G,
Tzekova M, Mincheva V, Manolova A, Vasilev I, Tisheva-Gospodinova S, Petrov I,
Postadzhiyan A, Velikov C, Dimov B, Constance C, Phaneuf DC, Mielniczuk L,
Pandey AS, Senaratne M, Zieroth S, Savard D, Stewart R, Huynh T, Al-Hesayen A,
Giannetti N, Moe G, Bourgeois R, Ezekowitz J, Hartleib M, Sussex B, Babapulle M,
Chehayeb R, Gaudet D, McKelvie R, Nguyen V, Roth S, Gupta M, Pesant Y, Rupka D,
Bhargava R, Costa-Vitali A, Proulx G, Vega M, Potthoff S, Schnettler Cid MC,
Villablanca Sepulveda AM, Lanas Zanetti FT, Corbalan R, Saavedra Gajardo VA,
Conejeros Kindel C, Pincetti Jofre CP, Cobos Segarra JL, Rodriguez Venegas ME,
Yanez Hidalgo M, Vejar Jalaf MG, Li W, Zhang J, Fu X, Zhang X, Li D, Wang Z, Qu
Y, Zheng Z, Tang H, Yang P, Zhang Y, Zheng Y, Mi Y, Huang H, Bu P, Chen G, Chen
J, Han Y, Li Z, Ma S, Yang X, Yuan Z, Dong Y, Li Z, Mahemuti A, Niu W, Yang Z,
Zhang Y, Sun Y, Wu W, Liu F, Yan J, Li Y, Wang Y, Zhang S, Zhou C, Cui H, Li J,
Li T, Han Q, Wei Y, Echeverria LE, Manzur Jattin F, Accini Mendoza JL, Castaño
Osorio W, Sanchez Vallejo G, Luengas CA, Coronel Arroyo JA, Moncada Corredor MA,
Saldarriaga Giraldo CI, Botero Lopez R, Molina de Salazar DI, Urina Triana M,
Atehortua Lopez LH, Olaya Rojas P, Velez Pelaez S, Lopez Pareja M, Cadena
Bonfanti A, Polasek R, Monhart Z, Sochor K, Motovska Z, Belohlavek J, Busak L,
Krupicka J, Tyl P, Jerabek O, Podpera I, Skrobakova J, Peterka K, Spacek R, Cech
V, Kellnerova I, Nechvatal L, Pozdisek Z, Houra M, Kryza R, Machova V, Brabec T,
Cepelak M, Stepek D, Zeman K, Klimsa Z, Koleckar P, Schee A, Spinarova L, Coufal
Z, Jeppesen J, Vraa S, Wiggers H, Nyvad O, Nielsen T, Gadsboll N, Kaiser-Nielsen
P, Eifer Moller J, Videbaek L, Galinier M, Lefebvre JM, Tartiere JM, De Geeter
G, Roubille F, Ricci JE, Salvat M, Gueffet JP, Decoulx E, Berdague P, Jondeau G,
Mewton N, Ovize M, De Groote P, Donal E, Isnard R, Sabatier R, Trochu JN, Damy
T, Georges JL, Rosamel Y, Picard F, Aboyans V, Laperche T, Mitrovic V,
Taggeselle J, Störk S, Ebelt H, Genth-Zotz S, Rassaf T, Duengen HD, Mittag M,
Menck N, von Haehling S, Zeymer U, Boehm M, Frankenstein L, Killat H, Bourhaial
H, Beug D, Horacek T, Pfister R, Sandri M, Westenfeld R, Kadel C, Karvounis H,
Patsilinakos S, Mantas I, Karavidas A, Giamouzis G, Tsioufis K, Naka K, Tziakas
D, Parissis J, Styliadis I, Barbetseas I, Manolis A, Kochiadakis G, Parthenakis
F, Herczeg B, Nagy L, Nyolczas N, Toth K, Merkely B, Laszlo Z, Mark L, Szakal I,
Papp A, Bezzegh K, Peterfai E, Lakatos F, Hajko E, Papp A, Forster T, Lupkovics
G, Mohacsi A, Salamon C, Aradi D, Andreka P, Szasz G, Zilahi Z, Kazinczy R,
Margonato A, Agostoni P, Fucili A, Piovaccari G, Senni M, Ambrosio G, Carluccio
E, Bilato C, Frigerio M, Indolfi C, Sinagra G, Brunetti ND, Di Biase M, Perna G,
Pini D, Volterrani M, De Ferrari GM, Leonardi S, Raineri C, Achilli F, Bonaduce
D, Mortara A, Musumeci G, Francisco Polo Friz HE, Rossini R, Tocchetti CG,
Vincenzi A, Cavallini C, Floresta AM, Zaca V, Giudici V, Quinto Villani G,
Higashino Y, Oishi S, Wada A, Yasaka Y, Fukuzawa S, Kawata H, Onoue K, Saito Y,
Koike A, Koizumi T, Masuda S, Meno H, Mitsuo K, Nakayama K, Taguchi S, Takahashi
N, Takenaka T, Tanabe J, Tsuboi H, Watanabe N, Yoshida T, Amano T, Ishikawa M,
Kida K, Kubota T, Nakamura K, Sakamoto T, Sato N, Shimomura M, Uehara H,
Yamamoto F, Yuge M, Akatsuka Y, Doi M, Domae H, Ebato M, Fujii K, Fujiwara W,
Gohara S, Hata Y, Hirayama A, Izawa H, Kagiyama S, Kanda J, Kitaoka H, Kiyokawa
H, Makino K, Matsumoto T, Michishita I, Miura S, Miyazaki T, Nakahama M,
Nakamura A, Nakazato Y, Ogawa T, Okumura T, Okumura Y, Sakai T, Saku K, Sato Y,
Shimizu W, Sugino H, Suzuki M, Takagi A, Takaishi H, Tanaka T, Terasaki T,
Tsujimoto M, Ueda Y, Ujino K, Usui M, Yamamoto M, Yoshikawa M, Ajioka M, Ando K,
Asakura M, Asano H, Fujii S, Hara H, Inomata T, Isshiki T, Kadokami T, Kai H,
Kasai T, Kawamitsu K, Kawasaki T, Koga T, Komiyama N, Maejima Y, Manita M,
Miyamoto N, Nagase K, Node K, Numaguchi K, Sakata Y, Serikawa T, Sugimura K,
Takama N, Tatebe S, Ueno H, Urabe Y, Hidaka T, Hiroi S, Iseki H, Ito H, Kajinami
K, Kawakami H, Kihara Y, Momiyama Y, Mori M, Morita Y, Okishige K, Sakagami S,
Takeishi Y, Terasawa A, Utsu N, Badariene J, Celutkiene J, Sakalyte G, Slapikas
R, Jarasuniene D, Garcia Castillo A, De Los Rios Ibarra MO, Ramos Lopez GA,
Bayram Llamas EA, Llamas Esperon GA, Salcido Vazquez E, Garcia Gonzalez R,
Arenas Leon JL, Gonzalez SL, Arias Mendoza MA, Contreras Rodriguez A, Mendez
Machado GF, Alpizar Salazar M, Bazzoni Ruiz AE, De Leon Flores AM, Pagola
Carrasco JA, Reyes Araiza R, Römer T, Remmen J, Van Eck J, Elvan A, Kedhi E,
Smilde T, van de Wal R, Lok D, Schaap J, van der Sluis A, Linssen G, Magro M,
Willems F, Schellings D, van der Zwaan C, van Hal J, Beelen D, Boswijk D,
Hermans W, van Huysduynen-Monraats P, Van Kesteren H, Scott R, Hart H, Szczasny
M, Blicharski T, Kafara M, Stankiewicz A, Skonieczny G, Zabowka M, Kania G,
Borkowski T, Kopaczewski J, Pawlowicz L, Spyra J, Wlodarczyk A, Sciborski R,
Balsam P, Drozdz J, Sobkowicz B, Konieczynska M, Lelonek M, Bednarkiewicz Z,
Trebacz J, Jankowski P, Sidor M, Berkowski P, Chmielak Z, Lenartowska L, Nessler
J, Straburzynska-Migaj E, Kalarus Z, Kowalski R, Kalecinska-Krystkiewicz E, Gola
Z, Pijanowski Z, Wozakowska-Kaplon B, Cymerman K, Rynkiewicz A, Miekus P,
Monteiro P, Morais Sarmento P, Almeida F, Duarte T, Oliveira L, Soares Goncalves
S, Santos L, Brito D, Stanciulescu G, Spiridon MR, Militaru C, Podoleanu CG,
Zdrenghea D, Popescu MI, Macarie C, Giuca A, Mitu F, Voicu OC, Dorobantu M,
Lighezan D, Stamate S, Bykov A, Kobalava Z, Zrazhevskiy K, Semenova I,
Vishnevsky A, Shutemova E, Tereschenko S, Shvarts Y, Barbarash O, Lukyanov Y,
Voevoda M, Dovgolis S, Dronov D, Goloshchekin B, Sitnikova M, Ezhov M, Tarasov
N, Kotelnikov M, Kostenko V, Solovev O, Boytsov S, Goncharov I, Myasnikov R,
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List the types of defensins expressed in humans. | Defensins are antimicrobial peptides that participate in the innate immunity of hosts. Humans constitutively and/or inducibly express α- and β-defensins, which are known for their antiviral and antibacterial activities. | Defensins are antimicrobial peptides that participate in the innate immunity of
hosts. Humans constitutively and/or inducibly express α- and β-defensins, which
are known for their antiviral and antibacterial activities. This review
describes the application of human defensins. We discuss the extant experimental
results, limited though they are, to consider the potential applicability of
human defensins as antiviral agents. Given their antiviral effects, we propose
that basic research be conducted on human defensins that focuses on RNA viruses,
such as human immunodeficiency virus (HIV), influenza A virus (IAV), respiratory
syncytial virus (RSV), and dengue virus (DENV), which are considered serious
human pathogens but have posed huge challenges for vaccine development for
different reasons. Concerning the prophylactic and therapeutic applications of
defensins, we then discuss the applicability of human defensins as antivirals
that has been demonstrated in reports using animal models. Finally, we discuss
the potential adjuvant-like activity of human defensins and propose an
exploration of the 'defensin vaccine' concept to prime the body with a
controlled supply of human defensins. In sum, we suggest a conceptual framework
to achieve the practical application of human defensins to combat viral
infections. |
What do HA and NA stand for with respect to the flue virus, e.g. H1N1? | VaxArray assays for influenza hemagglutinin (HA) and neuraminidase (NA) have been developed to address this need. | The hemagglutinin (HA) and neuraminidase (NA) external glycoprotein antigens of
H1N1 and H3N2 subtypes of epidemiologically important influenza A viruses
prevalent during recent decades were subjected to intensive antigenic analysis
by four different methods. Prior to serological analysis with polyclonal rabbit
antisera, HA and NA antigens of four viruses of each subtype were segregated by
genetic reassortment to forestall nonspecific steric hindrance during
antigen-antibody combination. This analysis has demonstrated that with respect
to antigenic phenotype, HA and NA proteins have evolved at different rates. With
H1N1 viruses, an arrest of significant evolution of the NA discordant with the
continuing antigenic drift of HA was found in the 1980-1983 period. It is
probable that the different and independent rates of evolution of HA and NA
reflect the greater selective pressure of HA antibodies, which forces the more
rapid emergence of HA escape mutants. The slower antigenic change found for NA
further supports the potential for NA-specific infection-permissive immunization
as a useful stratagem against influenza. The ability of a single dose of plasmid DNA encoding neuraminidase (NA) or
hemagglutinin (HA) from influenza virus A/PR/8/34 (PR8) (H1N1) to protect
against homologous virus infection was examined in BALB/c mice. In the present
study, mice were immunized once with 30 microg of NA or HA DNA by
electroporation. Four weeks or 28 weeks after immunization, mice were challenged
with a lethal dose of homologous virus and the ability of NA or HA DNA to
protect the mice from influenza was evaluated. We found that a single
inoculation of NA DNA could provide protection against influenza virus challenge
as well as long-term protection against viral infection. Whereas, the mice
immunized with a single dose of HA DNA could not be protected. In addition,
neonatal mice immunized with a single dose of 30 microg of NA DNA could be
provided with significant protection against viral infection. Influenza is an infectious disease caused by RNA viruses of the family
Orthomyxoviridae. The new influenza H1N1 viral stain has emerged by the genetic
combination of genes from human, pig, and bird's H1N1 virus. The influenza virus
is roughly spherical and is enveloped by a lipid membrane. There are two
glycoproteins in this lipid membrane; namely, hemagglutinin (HA) which helps in
attachment of the viral strain on the host cell surface and neuraminidase (NA)
that is responsible for initiation of viral infection. We have developed
homology models of both Hemagglutinin and Neuraminidase receptors from H1N1
strains in eastern India. The docking studies of B-Sialic acid and O-Sialic acid
in the optimized and energy-minimized homology models show important H-bonding
interactions with ALA142, ASP230, GLN231, GLU232, and THR141. This information
can be used for structure-based and pharmacophore-based new drug design. We have
also calculated ADME properties (Human Oral Absorption (HOA) and % HOA) for
Oseltamivir which have been subject of debate for long. Two surface glycoproteins of influenza virus, haemagglutinin (HA) and
neuraminidase (NA), play opposite roles in terms of their interaction with host
sialic acid receptors. HA attaches to sialic acid on host cell surface receptors
to initiate virus infection while NA removes these sialic acids to facilitate
release of progeny virions. This functional opposition requires a balance. To
explore what might happen when NA of an influenza virus was replaced by one from
another isolate or subtype, in this study, we generated three recombit
influenza A viruses in the background of A/PR/8/34 (PR8) (H1N1) and with NA
genes obtained respectively from the 2009 pandemic H1N1 virus, a highly
pathogenic avian H5N1 virus, and a lowly pathogenic avian H9N2 virus. These
recombit viruses, rPR8-H1N1NA, rPR8-H5N1NA, and rPR8-H9N2NA, were shown to
have similar growth kinetics in cells and pathogenicity in mice. However, much
more rPR8-H5N1NA and PR8-wt virions were released from chicken erythrocytes than
virions of rPR8-H1N1NA and rPR8-H9N2NA after 1 h. In addition, in MDCK cells,
rPR8-H5N1NA and rPR8-H9N2NA infected a higher percentage of cells, and induced
cell-cell fusion faster and more extensively than PR8-wt and rPR8-H1N1NA did in
the early phase of infection. In conclusion, NA replacement in this study did
not affect virus replication kinetics but had different effects on infection
initiation, virus release and fusion of infected cells. These phenomena might be
partially due to NA proteins' different specificity to α2-3/2-6-sialylated
carbohydrate chains, but the exact mechanism remains to be explored. In the vast majority of influenza A viruses characterized to date, hemagglutinin
(HA) is the receptor-binding and fusion protein, whereas neuraminidase (NA) is a
receptor-cleaving protein that facilitates viral release but is expendable for
entry. However, the NAs of some recent human H3N2 isolates have acquired
receptor-binding activity via the mutation D151G, although these isolates also
appear to retain the ability to bind receptors via HA. We report here the
laboratory generation of a mutation (G147R) that enables an N1 NA to completely
co-opt the receptor-binding function normally performed by HA. Viruses with this
mutant NA grow to high titers even in the presence of extensive mutations to
conserved residues in HA's receptor-binding pocket. When the receptor-binding NA
is paired with this binding-deficient HA, viral infectivity and red blood cell
agglutination are blocked by NA inhibitors. Furthermore, virus-like particles
expressing only the receptor-binding NA agglutinate red blood cells in an
NA-dependent manner. Although the G147R NA receptor-binding mutant virus that we
characterize is a laboratory creation, this same mutation is found in several
natural clusters of H1N1 and H5N1 viruses. Our results demonstrate that, at
least in tissue culture, influenza virus receptor-binding activity can be
entirely shifted from HA to NA. In nearly all characterized influenza viruses, hemagglutinin (HA) is the
receptor-binding protein while neuraminidase (NA) is a receptor-cleaving protein
that aids in viral release. However, in recent years, several groups have
described point mutations that confer receptor-binding activity on NA, albeit in
laboratory rather than natural settings. One of these mutations, D151G, appears
to arise in the NA of recent human H3N2 viruses upon passage in tissue culture.
We inadvertently isolated the second of these mutations, G147R, in the NA of the
lab-adapted A/WSN/33 (H1N1) strain while we were passaging a heavily engineered
virus in the lab. G147R also occurs at low frequencies in the reported sequences
of viruses from three different lineages: human 2009 pandemic H1N1 (pdmH1N1),
human seasonal H1N1, and chicken H5N1. Here we reconstructed a representative
G147R NA from each of these lineages and found that all of the proteins have
acquired the ability to bind an unknown cellular receptor while retaining
substantial sialidase activity. We then reconstructed a virus with the HA and NA
of a reported G147R pdmH1N1 variant and found no attenuation of viral
replication in cell culture or change in pathogenesis in mice. Furthermore, the
G147R virus had modestly enhanced resistance to neutralization by the Fab of an
antibody against the receptor-binding pocket of HA, although it remained
completely sensitive to the full-length IgG. Overall, our results suggest that
circulating N1 viruses occasionally may acquire the G147R NA receptor-binding
mutation without impairment of replicative capacity.
IMPORTANCE: Influenza viruses have two main proteins on their surface: one
(hemagglutinin) binds incoming viruses to cells, while the other (neuraminidase)
helps release newly formed viruses from these same cells. Here we characterize
unusual mutant neuraminidases that have acquired the ability to bind to cells.
We show that the mutation that allows neuraminidase to bind cells has no
apparent adverse effect on viral replication but does make the virus modestly
more resistant to a fragment of an antibody that blocks the normal
hemagglutinin-mediated mode of viral attachment. Our results suggest that
viruses with receptor-binding neuraminidases may occur at low levels in
circulating influenza virus lineages. Influenza type A viruses are classified into subtypes based on their two surface
proteins, hemagglutinin (HA) and neuraminidase (NA). The HA protein facilitates
the viral binding and entering a host cell and the NA protein helps the release
of viral progeny from the infected cell. The complementary roles of HA and NA
entail their collaboration, which has important implications for viral
replication and fitness. The HA protein from early strains of pandemic 2009 H1N1
of swine origin preferentially binds to human type receptors with a weak binding
to avian type receptors. This virus caused several human deaths in December 2013
in Texas, USA, which motivated us to investigate the changes of genetic features
that might contribute to the surged virulence of the virus. Our time series
analysis on the strains of this virus collected from 2009 to 2013 implied that
the HA binding preference of this virus in USA, Europe, and Asia has been the
characteristic of swine H1N1 virus since 2009. However, its characteristic of
seasonal human H1N1 and its binding avidity for avian type receptors both were
on steady rise and had a clear increase in 2013 with American strains having the
sharpest surge. The first change could enhance the viral transmission and
replication in humans and the second could increase its ability to cause
infection deep in lungs, which might account for the recent human deaths in
Texas. In light of HA and NA coadaptation and evolutionary interactions, we also
explored the NA activity of this virus to reveal the functional balance between
HA and NA during the course of virus evolution. Finally we identified amino acid
substitutions in HA and NA of the virus that were critical for the observed
evolution. Genetic variation of influenza neuraminidase (NA), unlike for hemagglutinin
(HA), has not been fully characterized. Therefore, we determined the relation
between mutations in the NA and HA genome segments of 205 influenza A/H3N2
viruses isolated from patients in Japan during the five seasons from 2010 to
2015. The amino acid (AA) sequences of the NA and HA proteins in these isolates
were then determined. In the 2011-2012 season, there was the emergence of
isolates with NA and HA sequences containing AA93G (NA93G) and AA278K (HA278K),
respectively (24/48 isolates, 50.0%). This was in contrast to NA93D-HA278N being
detected exclusively in the previous 2010-2011 season (24/24 isolates, 100.0%).
The isolates with the NA93G-HA278K substitutions became predomit in the
following 2012-2013 season (95.8%, 46/48 isolates). The NA and HA phylogenetic
trees of the 2011-2012 and 2012-2013 seasons were segregated by clades with
NA93D-HA278N or NA93G-HA278K. In the subsequent 2013-2014 and 2014-2015 seasons,
the strong relationship between NA93D-HA278N and NA93G-HA278K observed in the
previous seasons, was no longer present and NA93G-HA278N (33/52 isolates, 63.5%
in the 2014-2015 season) became predomit. In addition, the clades within the
NA and HA trees could no longer be segregated based on NA AA93 and HA AA278.
These findings suggest that the co-mutation of NA and HA AA sequences is present
and may contribute to the formation of an epidemic lineage. The fifth wave of A(H7N9) virus infection in China from 2016 to 2017 caused
great concern due to the large number of individuals infected, the isolation of
drug-resistant viruses, and the emergence of highly pathogenic strains.
Antibodies against neuraminidase (NA) provide added benefit to
hemagglutinin-specific immunity and may be important contributors to the
effectiveness of A(H7N9) vaccines. We generated a panel of mouse monoclonal
antibodies (MAbs) to identify antigenic domains on NA of the novel A(H7N9) virus
and compared their functional properties. The loop formed in the region of
residue 250 (250 loop) and the domain formed by the loops containing residues
370, 400, and 430 were identified as major antigenic regions. MAbs 1E8, 2F6,
10F4, and 11B2, which recognize these two antigenic domains, were characterized
in depth. These four MAbs differ in their abilities to inhibit cleavage of small
and large substrates (methyl-umbelliferyl-acetyl neuraminic acid [MU-NANA] and
fetuin, respectively) in NA inhibition assays. 1E8 and 11B2 did not inhibit NA
cleavage of either MU-NANA or fetuin, and 2F6 inhibited cleavage of fetuin
alone, whereas 10F4 inhibited cleavage of both substrates. All four MAbs reduced
the in vitro spread of viruses carrying either the wild-type N9 or N9 with
antiviral-resistant mutations but to different degrees. These MAbs have
different in vivo levels of effectiveness: 10F4 was the most effective in
protecting mice against challenge with A(H7N9) virus, 2F6 was less effective,
and 11B2 failed to protect BALB/c mice at the doses tested. Our study confirms
that NA-specific antibodies can protect against A(H7N9) infection and suggests
that in vitro properties can be used to rank antibodies with therapeutic
potential.IMPORTANCE The novel A(H7N9) viruses that emerged in China in 2013
continue to infect humans, with a high fatality rate. The most recent outbreak
resulted in a larger number of human cases than previous epidemic waves. Due to
the absence of a licensed vaccine and the emergence of drug-resistant viruses,
there is a need to develop alternative approaches to prevent or treat A(H7N9)
infection. We have made a panel of mouse monoclonal antibodies (MAbs) specific
for neuraminidase (NA) of A(H7N9) viruses; some of these MAbs are effective in
inhibiting viruses that are resistant to antivirals used to treat A(H7N9)
patients. Binding avidity, inhibition of NA activity, and plaque formation
correlated with the effectiveness of these MAbs to protect mice against lethal
A(H7N9) virus challenge. This study identifies in vitro measures that can be
used to predict the in vivo efficacy of NA-specific antibodies, providing a way
to select MAbs for further therapeutic development. Author information:
(1)Department of Medicine, Section of Rheumatology, the Knapp Center for Lupus
and Immunology, University of Chicago, Chicago, IL 60637, USA.
(2)Department of Microbiology, Icahn School of Medicine at Mount Sinai, New
York, NY 10029, USA.
(3)The Committee on Immunology, University of Chicago, Chicago, IL 60637, USA.
(4)Department of Chemical Engineering, University of Texas at Austin, Austin, TX
78731, USA.
(5)Division of Viral Products, Center for Biologics Evaluation and Research,
Food and Drug Administration, Silver Spring, MD 20993, USA.
(6)Center for Vaccine Biology & Immunology, Department of Microbiology &
Immunology, University of Rochester Medical Center, Rochester, NY 14642, USA.
(7)Division of Infectious Disease, Department of Medicine, University of
Rochester Medical Center, Rochester, NY 14642, USA.
(8)Emory Vaccine Center, Department of Pediatrics, Division of Infectious
Disease, Emory University School of Medicine, Atlanta, GA 30322, USA.
(9)Department of Microbiology, Icahn School of Medicine at Mount Sinai, New
York, NY 10029, USA. Electronic address: [email protected].
(10)Department of Medicine, Section of Rheumatology, the Knapp Center for Lupus
and Immunology, University of Chicago, Chicago, IL 60637, USA. Electronic
address: [email protected]. Practical methods to measure the potency of influenza vaccines are needed as
alternatives for the standard single radial immunodiffusion (SRID) assay.
VaxArray assays for influenza hemagglutinin (HA) and neuraminidase (NA) have
been developed to address this need. In this report, we evaluate the use of
these assays to assess the potency of HA and NA of an A/H3N2 subunit vaccine by
determining the correlation between the amounts measured by VaxArray and the
immunogenicity in mice. The antibody response after one and two doses of five
formulations of the vaccine ranging from 5 µg/mL to 80 µg/mL of HA, was measured
by hemagglutination inhibition (HAI) and neuraminidase inhibition (NAI) assays.
For hemagglutinin, vaccine potency determined by VaxArray was equivalent to
potency measured SRID and these amounts were predictive of immunogenicity, with
excellent correlation between potency measured by VaxArray and the HAI geometric
mean titers (GMT). Likewise, the amount of NA measured by VaxArray was
predictive of the NAI GMT. The VaxArray NA assay reported non-detectable levels
of intact NA for a sample that had been heat degraded at 56 °C for 20 h,
demonstrating that the assay measures the native, active form of NA. Similarly,
the HA potency measured by VaxArray in this heat-treated sample was very low
when a monoclonal antibody was used to detect the amount of antigen bound.
Importantly, the force degraded sample induced low HAI titers and the NAI titers
were not measurable, supporting the conclusion that the VaxArray HA and NA
assays measure the immunogenic forms of these A/H3N2 antigens. This study
indicates that VaxArray assays can be used to assess the potency of HA and NA
components in influenza vaccines as a proxy for immunogenicity. The effectiveness of influenza vaccines against circulating A(H1N1)pdm09 viruses
was modest for several seasons despite the absence of antigenic drift of
hemagglutinin (HA), the primary vaccine component. Since antibodies against HA
and neuraminidase (NA) contribute independently to protection against disease,
antigenic changes in NA may allow A(H1N1)pdm09 viruses to escape from
vaccine-induced immunity. In this study, analysis of the specificities of human
NA-specific monoclonal antibodies identified antigenic sites that have changed
over time. The impact of these differences on in vitro inhibition of enzyme
activity was not evident for polyclonal antisera until viruses emerged in 2013
without a predicted glycosylation site at amino acid 386 in NA. Phylogenetic and
antigenic cartography demonstrated significant antigenic changes that in most
cases aligned with genetic differences. Typical of NA drift, the antigenic
difference is observed in one direction, with antibodies against conserved
antigenic domains in A/California/7/2009 (CA/09) continuing to inhibit NA of
recent A(H1N1)pdm09 viruses reasonably well. However, ferret CA/09-specific
antiserum that inhibited the NA of A/Michigan/45/2015 (MI/15) very well in
vitro, protected mice against lethal MI/15 infection poorly. These data show
that antiserum against the homologous antigen is most effective and suggest the
antigenic properties of NA should not be overlooked when selecting viruses for
vaccine production.IMPORTANCE The effectiveness of seasonal influenza vaccines
against circulating A(H1N1)pdm09 viruses has been modest in recent years,
despite the absence of antigenic drift of HA, the primary vaccine component.
Human monoclonal antibodies identified antigenic sites in NA that changed early
after the new pandemic virus emerged. The reactivity of ferret antisera
demonstrated antigenic drift of A(H1N1)pdm09 NA from 2013 onward. Passive
transfer of serum raised against A/California/7/2009 was less effective than
ferret serum against the homologous virus in protecting mice against a virus
with the NA of more recent virus, A/Michigan/45/2015. Given the long-standing
observation that NA-inhibiting antibodies are associated with resistance against
disease in humans, these data demonstrate the importance of evaluating NA drift
and suggest that vaccine effectiveness might be improved by selecting viruses
for vaccine production that have NAs antigenically similar to those of
circulating influenza viruses. The H1N1 influenza pandemic vaccine has been developed from the
A/California/07/09 (Cal) virus and the well-known high-yield A/Puerto Rico/8/34
(PR8) virus by classical reassortment and reverse genetics (RG) in eggs.
Previous studies have suggested that Cal-derived chimeric hemagglutinin (HA) and
neuraminidase (NA) improve virus yields. However, the cell-based vaccine of the
H1N1 pandemic virus has been less investigated. RG viruses that contained
Cal-derived chimeric HA and NA could be rescued in Madin-Darby canine kidney
cells that expressed α2,6-sialyltransferase (MDCK-SIAT1). The viral growth
kinetics and chimeric HA and NA properties were analyzed. We attempted to
generate various RG viruses that contained Cal-derived chimeric HA and NA, but
half of them could not be rescued in MDCK-SIAT1 cells. When both the 3'- and
5'-terminal regions of Cal HA viral RNA were replaced with the corresponding
regions of PR8 HA, the RG viruses were rescued. Our results were largely
consistent with those of previous studies, in which the N- and C-terminal
chimeric HA slightly improved virus yield. Importantly, the chimeric HA,
compared to Cal HA, showed cell fusion ability at a broader pH range, likely due
to amino acid substitutions in the transmembrane region of HA. The rescued RG
virus with high virus yield harbored the chimeric HA capable of cell fusion at a
broader range of pH. Hemagglutinin (HA) and neuraminidase (NA) are major glycoproteins expressed on
the surface of influenza virus. They have complementary and antagonistic
functions that facilitate in the life cycle of the virus. The functional
equilibrium generated between HA and NA can impact the evolution and adaptation
of influenza virus strains within the human reservoir. This functional
equilibrium is referred to as the "HA-NA balance". An imbalanced HA-NA can
restrict the multiplication and transmission capacity of influenza viruses.
Moreover, this equilibrium is likely a limiting factor against species crossover
for the virus. In light of such considerations, the HA-NA balance should be
precisely studied to gain a better understanding of the emergence of pandemic
and seasonal influenza virus strains. This review describes the concept of the
HA-NA balance, the methods used to study it, plus a discussion of the HA-NA
balance in the evolution of the pandemic influenza A H1N1 strains that plagued
the world in 1918 and 2009. |
Is the TFR1 gene dispensable for erythropoiesis? | Yes. The TFR1 gene is a key part of the mechanism by which the body delivers iron to the red blood cells. It is not dispensable for erythropoiesis. | Soluble transferrin receptor-1 (sTfR1) concentrations are increased in the
plasma under two conditions that are associated with increased iron absorption,
i.e. iron deficiency and increased erythropoiesis. To determine the possible
role of sTfR1 as a signaling mechanism for iron absorption, a hydrodynamic gene
transfer technique was established to express transfected plasmid constructs of
human sTfR1 (hsTfR1) and murine sTfR1 (msTfR1) from the livers of C57BL/6 mice.
Iron absorption, serum iron levels and hepcidin expression were then measured.
The hydrodynamic gene transfer technique proved to be an effective approach to
achieving sustained expression of sTfR1 in mice. Although expression of high
levels of sTfR1 significantly increased serum iron levels, repeated experiments
showed that neither hsTfR1 nor msTfR1 had any effect on iron absorption or
hepcidin mRNA expression levels. Thus, despite its attractiveness as a potential
modifier of iron absorption, sTfR1 levels do not exert a regulatory effect on
iron absorption. Sorting of endocytic ligands and receptors is critical for diverse cellular
processes. The physiological significance of endosomal sorting proteins in
vertebrates, however, remains largely unknown. Here we report that sorting nexin
3 (Snx3) facilitates the recycling of transferrin receptor (Tfrc) and thus is
required for the proper delivery of iron to erythroid progenitors. Snx3 is
highly expressed in vertebrate hematopoietic tissues. Silencing of Snx3 results
in anemia and hemoglobin defects in vertebrates due to impaired transferrin
(Tf)-mediated iron uptake and its accumulation in early endosomes. This impaired
iron assimilation can be complemented with non-Tf iron chelates. We show that
Snx3 and Vps35, a component of the retromer, interact with Tfrc to sort it to
the recycling endosomes. Our findings uncover a role of Snx3 in regulating Tfrc
recycling, iron homeostasis, and erythropoiesis. Thus, the identification of
Snx3 provides a genetic tool for exploring erythropoiesis and disorders of iron
metabolism. PURPOSE OF REVIEW: The type 1 transferrin receptor (TfR1) is well known as a key
player in erythroid differentiation through its role in iron uptake. Recently,
it has been demonstrated that TfR1 could also have signaling functions in
erythroid cells. Moreover, the second transferrin receptor, TfR2, whose
signaling functions in hepatic cells are well established, was recently shown to
be a partner of the erythropoietin receptor (EpoR) and thereby likely to play a
role in erythroid differentiation.
RECENT FINDINGS: This review reports recent findings regarding the specificities
of the regulation of TfR1 expression and iron uptake in erythroblasts. The newly
discovered noncanonical actions of TfR1 and TfR2 in erythroid cells are also
discussed.
SUMMARY: Erythrocytes contain more than 60% of the iron of the body and each
day, differentiating erythroid cells uptake around 20 mg of iron for heme
synthesis. Accordingly, TfR1 is one of the most abundant membrane proteins of
the erythroblasts and it is not surprising that specific regulations regarding
both its expression and its mechanism of action operate in erythroblasts. The
signaling functions of both TfR1 and TfR2 in erythroid cells were unexpected and
these recent findings open a new field of research regarding the last steps of
erythroid differentiation and their regulation. To identify novel regulators of erythropoiesis, we performed independent forward
genetic screens using the chemical mutagen ENU in mice. Among progeny displaying
microcytic red-cell phenotypes, 7 independent mouse strains harboring mutations
within the transferrin receptor gene Tfrc were identified. Six of the mutants,
including the previously described red blood cell 6 (RBC6) strain, displayed
reduced erythroblast CD71 expression and midgestation lethality of homozygotes
(E12.5-E14.5), and 1 novel strain, RBC21, displayed a variable phenotype with
sustained CD71 expression and late homozygous lethality (E18.5). Standard iron
studies were normal in the RBC21 mutant, but intracellular ferritin was
significantly reduced. The microcytic phenotype seen in the RBC21 strain was the
result of impaired binding of transferrin to the receptor. Neither RBC6 nor
RBC21 responded to iron replacement therapy. These studies describe how point
mutations of the transferrin receptor can cause a microcytic anemia that does
not respond to iron therapy and would not be detected by routine iron studies,
such as serum ferritin. BACKGROUND: Fetal and neonatal brain iron content is compromised at the time of
anemia, suggesting that screening for iron deficiency by measuring hemoglobin is
inadequate to protect the brain. Reticulocyte hemoglobin (Ret-He) reflects
iron-deficient (ID) erythropoiesis prior to anemia.
METHODS: At postnatal day (P), 10 and 20 iron-sufficient rat pups were fostered
to ID dams to produce a postnatal ID (PNID) group, which was compared to 20
iron-sufficient (IS) pups fostered by IS dams. Pups were assessed from P13 to
P15 for hemoglobin, hematocrit, reticulocyte count, and Ret-He. Hippocampal iron
status was assessed by transferrin receptor-1 (Tfrc-1) and divalent metal
transporter-1 (Slc11a2) mRNA expression.
RESULTS: At P13, brain iron status was similar between groups; only Ret-He was
lower in the PNID group. At P14, the PNID group had lower Ret-He, hematocrit,
mean corpuscular volume (MCV), and reticulocyte percentage (RET%). Tfrc-1
expression was increased, consistent with brain iron deficiency. Both Ret-He and
MCV correlated with brain iron status at P14 and P15.
CONCLUSIONS: Ret-He was the only red cell marker affected prior to the onset of
brain ID. The clinical practice of using anemia as the preferred biomarker for
diagnosis of iron deficiency may need reconsidering. Transferrin receptor 1 (Tfr1) mediates the endocytosis of diferric transferrin
in order to transport iron, and Tfr1 has been suggested to play an important
role in hematopoiesis. To study the role of Tfr1 in hematopoiesis, we generated
hematopoietic stem cell (HSC) specific Tfr1 knockout mice. We found that Tfr1
conditional knockout mice reached full term but died within one week of birth.
Further analyses revealed that Tfr1-deficient HSC had impaired development of
all hematopoietic progenitors except thrombocytes and B lymphocytes. In
addition, Tfr1-deficient cells had cellular iron deficiency, which blocked the
proliferation and differentiation of hematopoietic precursor cells, attenuated
the commitment of hematopoietic lineages, and reduced the regeneration potential
of HSC. Notably, hemin rescued the colony-forming capacity of Tfr1-deficient
HSC, whereas expressing a mutant Tfr1 that lacks the protein's iron-transporting
capacity failed to rescue hematopoiesis. These findings provide direct evidence
that Tfr1 is essential for hematopoiesis through binding diferric transferrin to
supply iron to cells. |
Which are the predominant rotavirus genotypes around the world? | The predominant RV genotypes circulating all over the world are G1P[8], G2P[4], G3P[8], G4P[8], and G9P[8], while G12[P6] and G12[P8] are emerging genotypes. | Comprehensive reviews of pre licensure rotavirus strain prevalence data
indicated the global importance of six rotavirus genotypes, G1P[8], G2P[4],
G3P[8], G4P[8], G9P[8] and G12P[8]. Since 2006, two vaccines, the monovalent
Rotarix (RV1) and the pentavalent RotaTeq (RV5) have been available in over 100
countries worldwide. Of these, 60 countries have already introduced either RV1
or RV5 in their national immunization programs. Post licensure vaccine
effectiveness is closely monitored worldwide. This review aimed at describing
the global changes in rotavirus strain prevalence over time. The genotype
distribution of the nearly 47,000 strains that were characterized during
2007-2012 showed similar picture to that seen in the preceding period. An
intriguing finding was the transient predomice of heterotypic strains, mainly
in countries using RV1. Unusual and novel antigen combinations continue to
emerge, including some causing local outbreaks, even in vaccinated populations.
In addition, vaccine strains have been found in both vaccinated infants and
their contacts and there is evidence for genetic interaction between vaccine and
wild-type strains. In conclusion, the post-vaccine introduction strain
prevalence data do not show any consistent pattern indicative of selection
pressure resulting from vaccine use, although the increased detection rate of
heterotypic G2P[4] strains in some countries following RV1 vaccination is
unusual and this issue requires further monitoring. Since its discovery 40 years ago, rotavirus (RV) is considered to be a major
cause of infant and childhood morbidity and mortality particularly in developing
countries. Nearly every child in the world under 5 years of age is at the risk
of RV infection. It is estimated that 90% of RV-associated mortalities occur in
developing countries of Africa and Asia. Two live oral vaccines, RotaTeq (RV5,
Merck) and Rotarix (RV1, GlaxoSmithKline) have been successfully deployed to
scale down the disease burden in Europe and America, but they are less effective
in Africa and Asia. In April 2009, the World Health Organization recommended the
inclusion of RV vaccination in national immunization programs of all countries
with great emphasis in developing countries. To date, 86 countries have included
RV vaccines into their national immunization programs including 41 Global
Alliance for Vaccines and Immunization eligible countries. The predomit RV
genotypes circulating all over the world are G1P[8], G2P[4], G3P[8], G4P[8], and
G9P[8], while G12[P6] and G12[P8] are emerging genotypes. On account of the
segmented genome, RV shows an enormous genetic diversity that leads to the
evolution of new genotypes that can influence the efficacy of current vaccines.
The current need is for a global RV surveillance program to monitor the
prevalence and antigenic variability of new genotypes to formulate future
vaccine development planning. In this review, we will summarize the previous and
recent insights into RV structure, classification, and epidemiology and current
status of RV vaccination around the globe and will also cover the status of RV
research and vaccine policy in Pakistan. |
Which TREX mRNA export complex subunits have been implicated in neurodevelopmental disorders? | Multiple TREX mRNA export complex subunits, e.g. THOC1, THOC2, THOC5, THOC6, THOC7, have been implicated in neurodevelopmental disorders (NDDs), neurodegeneration and cancer. | Author information:
(1)Adelaide Medical School and the Robinson Research Institute, The University
of Adelaide, Adelaide, SA, Australia.
(2)Genetics of Learning Disability Service, Hunter Genetics, Waratah, NSW,
Australia.
(3)School of Women's and Children's Health, University of New South Wales,
Randwick, NSW, Australia.
(4)Faculty of Biology, Medicine and Health, Division of Evolution and Genomic
Sciences, School of Biological Sciences, University of Manchester, Manchester,
United Kingdom.
(5)Manchester Centre for Genomic Medicine, St. Mary's Hospital, Manchester
University NHS Foundation Trust, Health Innovation Manchester, Manchester,
United Kingdom.
(6)Department of Neuroscience, Erasmus MC University Medical Center, Rotterdam,
Netherlands.
(7)ENCORE Expertise Centre for Neurodevelopmental Disorders, Erasmus MC
University Medical Center, Rotterdam, Netherlands.
(8)Genetic Medicine, Department of Pediatrics, University of California, San
Francisco, San Francisco, CA, United States.
(9)North West Thames Regional Genetics Service, Northwick Park Hospital, Harrow,
United Kingdom.
(10)Nottingham Clinical Genetics Service, Nottingham University Hospitals NHS
Trust, and the 100,000 Genomes Project and the Genomics England Research
Consortium, Nottingham, United Kingdom.
(11)Division of Pediatric Neurology, Medical University of South Carolina,
Charleston, SC, United States.
(12)Department of Pediatrics, McMaster University Medical Centre, Hamilton, ON,
Canada.
(13)Department of Diagnostic Genomics, PathWest, Nedlands, WA, Australia.
(14)Division of Pathology and Laboratory Medicine, Medical School, University of
Western Australia, Crawley, WA, Australia.
(15)Faculty of Health and Medical Sciences, University of Western Australia
Medical School, Perth, WA, Australia.
(16)Genetic Services of Western Australia, Undiagnosed Diseases Program,
Department of Health, Government of Western Australia, Perth, WA, Australia.
(17)Linear Clinical Research, Perth, WA, Australia.
(18)Child Health Evaluative Sciences, Research Institute, The Hospital for Sick
Children, and Institute of Health Policy Management and Evaluation, University
of Toronto, Toronto, ON, Canada.
(19)Genome Diagnostics, Department of Paediatric Laboratory Medicine, The
Hospital for Sick Children, and Laboratory Medicine and Pathobiology, University
of Toronto, Toronto, ON, Canada.
(20)Department of Paediatrics, Division of Clinical and Metabolic Genetics, The
Hospital for Sick Children, Toronto, ON, Canada.
(21)Department of Clinical Genetics, Erasmus MC University Medical Center,
Rotterdam, Netherlands.
(22)Royal Devon and Exeter NHS Foundation Trust, Exeter, United Kingdom.
(23)Neuroradiology, Royal North Shore Hospital, Sydney, NSW, Australia.
(24)Childhood Disability Prevention, South Australian Health and Medical
Research Institute, Adelaide, SA, Australia. |
What is the use of the Liverpool Elbow Score? | The Liverpool Elbow Score (LES) is a newly developed, validated elbow-specific score. It has been widely used to assess the outcomes of total elbow replacement in various conditions. | BACKGROUND: Chronic lateral elbow epicondylitis is a tendinosis with
angiofibrolastic degeneration of the wrist extensors' origin. Healing of this
lesion is reported with the use of autologous blood as well as with
platelet-rich plasma (PRP).
PURPOSE: A comparative study of these 2 treatments was conducted in an effort to
investigate the possible advantages of PRP.
STUDY DESIGN: Randomized controlled trial; Level of evidence, 1.
METHODS: Twenty-eight patients were divided equally into 2 groups, after blocked
randomization. Group A was treated with a single injection of 3 mL of autologous
blood and group B with 3 mL of PRP under ultrasound guidance. A standardized
program of eccentric muscle strengthening was followed by all patients in both
groups. Evaluation using a pain visual analog scale (VAS) and Liverpool elbow
score was performed at 6 weeks, 3 months, and 6 months.
RESULTS: The VAS score improvement was larger in group B at every follow-up
interval but the difference was statistically significant only at 6 weeks, when
mean improvement was 3.8 points (95% confidence interval [CI], 3.1-4.5) in group
B (61.47% improvement) and 2.5 points (95% CI, 1.9-3.1) in group A (41.6%
improvement) (P < .05). No statistically significant difference was noted
between groups regarding Liverpool elbow score.
CONCLUSION: Regarding pain reduction, PRP treatment seems to be an effective
treatment for chronic lateral elbow epicondylitis and superior to autologous
blood in the short term. Defining details of indications, best PRP
concentration, number and time of injections, as well as rehabilitation protocol
might increase the method's effectiveness. Additionally, the possibility of cost
reduction of the method might justify the use of PRP over autologous whole blood
for chronic or refractory tennis elbow. PURPOSE OF THE STUDY: The aim of the study was to compare two surgical methods
of treating diaphyseal fractures of the humerus.
MATERIAL AND METHODS: A prospective randomised study on the surgical treatment
of humeral diaphyseal fractures was carried out between September 2009 and
January 2013. The patients indicated for surgical treatment fell into two groups
according to the technique used as follows: minimally invasive plate
osteosynthesis (MIPO) with a locking compression plate (LCP; Synthes,
Switzerland); depending on the fracture type and course of fracture line, either
a straight narrow or a Philos or a metaphyseal LCP was used (group 1); and
intramedullary nailing (EHN, Synthes, Switzerland) (group 2). A total of 49
patients were entered into the study. The men-to-women ratio was about 1:1 and
the average age was 52 years (18 to 83). Of them, 45 patients with 46 humeral
fractures were followed up and evaluated. The injury was caused by a fall from
standing or while walking (n=21), traffic accident (n=16), sports activity (n=3)
or work-related activity (n=5). A single injury was treated in 72% and multiple
trauma in 28% of the patients. Using the AO classification, fractures were
diagnosed as types A, B and C in 25, 10 and 11 patients, respectively. The
patients were randomised into the groups using a computer programme allowing for
the maintece of group homogeneity. Each patient received information on the
method to be used in a sealed envelope.
RESULTS: The average injury-surgery interval was 6 days (range, 0 to 26). The
average operative time was 128 minutes (80 to 200). The average intra-operative
exposure to X-ray was 8 minutes (3 to 20). The average hospital stay was 20 days
(4 to 100) The average rehabilitation time till functional recovery was 17 weeks
(3 to 37), the time of bone union by radiographic assessment was 20 weeks (12 to
40). The functional outcome was assessed using the Constant-Murley (CM) and
Liverpool Elbow Score (LES) systems. The relative CM/LES score, as compared with
the healthy extremity, was 95/100. Excellent and good results were achieved in
89% and complications were recorded in 26% of the patients. The most frequent
complication was impingement syndrome or prolonged rehabilitation both in the
patients with proximal humerus fractures managed with the Philos locking plate
and in those treated by intramedullary nailing, although this was not
necessarily due to a technical error, i.e., osteosynthetic material protrusion.
Iatrogenic radial nerve injury was found only in one patient treated by
intramedullary nailing and was associated with traction during reduction and
nail insertion. Pseudoarthrosis was recorded in one patient of each group and
there were no infectious complications.
DISCUSSION: Intramedullary nailing has not yet shown such good outcomes in
humeral as in tibial fractures. The results of metaanalysis have indicated that
humeral fractures treated by plate osteosynthesis have fewer complications and
require repeat surgery less frequently. However, as shown by most recent
studies, this difference is getting smaller. The MIPO technique was adopted at
our department several years ago and the aim was to compare our results with
those of other centres. The studies so far published have show that MIPO and
conventional open plate osteosynthesis give comparable outcomes. CONCLUSIONS The
statistical evaluation using the unpaired t-test did not show any significant
differences in either the functional results or the number of complications
between the two methods. Both achieved about 90% of excellent and good results
and had 26% of complications. The only significant difference was found in the
length of operative time (136 min in MIPO versus 117 min in EHN). With use of
the Chi-Square test, a significant correlation between AO fracture type and
treatment outcome in the MIPO group was found, i.e., no poor result was recorded
for type A fractures, as assessed by the CM/LES score. No such correlation
between the fracture type and the functional outcome was seen in the EHN group. BACKGROUND: Total elbow arthroplasty (TEA) is increasingly used for the
treatment of advanced elbow conditions to reduce pain and improve function.
However, TEA is still associated with a higher complication rate than total hip
and knee arthroplasty despite advances in the design and surgical techniques.
This prospective clinical study reports the outcome of the Discovery Elbow
System (Biomet, Warsaw IN, USA), which has been in clinical use in the United
Kingdom since 2003.
METHODS: The study included a total of 100 Discovery Elbows (April 2003 to
January 2010) with a minimum 2-year follow-up, including 75 primary and 25
revisions (60% women and 40% men; mean age, 62 years). Outcome was assessed by
means of the Liverpool Elbow Score, pain experience, patient satisfaction, range
of motion, and radiographic imaging.
RESULTS: The mean follow-up period was 48.5 months (range, 24-108 months). The
Liverpool Elbow Score improved from 3.79 to 6.36 (P < .001). The percentage of
pain-free patients was substantially increased from 7% preoperatively to 64% at
the final follow-up. The patient satisfaction rate was over 90%. The
flexion-extension arc and pronation-supination arc increased from 72° to 93° and
from 86° to 111°, respectively (P < .001). Major postoperative complications
included deep infection (2%), progressive aseptic loosening requiring revision
(primary, 5%; revision 12%), persistent ulnar neuropathy (3%), and
periprosthetic fracture (primary, 6.8%; revision, 8%).
CONCLUSION: The Discovery Elbow System resulted in improved function, reduced
pain, and high patient satisfaction. Long-term results are required to assess
the survivorship of this system. BACKGROUND: The available literature on the use of a cementless total elbow
arthroplasty (TEA) design and its results are limited. This clinical study
reports the outcome of the cementless Discovery elbow system.
METHODS: Patients were operated on by a single surgeon between 2007 and 2014.
Nineteen patients (20 elbows) were available for review, 2 women (1 bilateral
TEA) and 17 men. The age of the patients ranged from 27 to 75 years (mean, 48
years). The mean follow-up was 61.8 months (range, 12-156 months). Patients were
assessed for range of motion, pain, and satisfaction level. Outcome scores
included the Mayo Elbow Performance Score, the Liverpool Elbow Score, and the
12-Item Short Form Health Survey (version 1). Radiographs were reviewed to
evaluate for loosening.
RESULTS: The mean Mayo Elbow Performance Score was 77.25, and the mean Liverpool
Elbow Score was 6.76. The mean flexion range was 123°, and the mean extension
lag was 35°. The mean pronation was 59°, and the mean supination was 58°. On
radiologic evaluation, there were no signs of loosening; however, in 2 cases,
nonprogressive radiolucent lines were observed. No signs of infection were
detected at final follow-up, and no elbows were revised. More than 90% of
patients were satisfied with the overall outcome.
CONCLUSION: The cementless TEA seems to be a reliable option for treatment of
varying elbow diseases. Long-term results are needed to assess the survivorship
of this design. BACKGROUND: The use of self-assessment questionnaires in addition to clinical
evaluations is gradually increasing. Liverpool Elbow Scale (LES) is an
elbow-specific outcome score that provides a comprehensive assessment of by both
the clinicians and patients. However, it has not been adapted and validated to
Turkish language.
OBJECTIVE: To conduct the translation, cross-cultural adaptation and validation
of Liverpool Elbow Score-patient answered outcome (LES-PAQ) into Turkish for
patients with elbow fracture.
DESIGN: Study of diagnostic accuracy/assessment scale.
METHODS: This study was carried out in three consecutive phases: translation,
cross-cultural adaptation and validation. In the third phase, we used the Quick
Disabilities of the Arm, Shoulder and Hand (Quick-DASH), Mayo Elbow Performance
Score (MEPS) and 12-Item Short Form Survey (SF-12) physical health score.
RESULTS: Sixty-one patients were included for the analysis. Neither a ceiling
nor a floor effect was observed. Cronbach's α coefficient was 0.89. Intraclass
correlation coefficient was 0.94 (95% CI 0.89 to 0.96; p < 0.001). SEM was 0.28
and MDC95 was 0.79. The LES-PAQ showed a high negative correlation with the
Quick-DASH (r = -0.72, p < 0.001) and high positive correlation with MEPS
(r = 0.77, p < 0.001), and with SF-12 physical health subscale (r = 0.73,
p < 0.001).
CONCLUSIONS: The Turkish version of the LES-PAQ is a reliable and valid tool for
the assessment of the patients with elbow fracture. |
Is Bcl-2-like protein 1 an pro apoptotic protein? | No,
it is an anti-apoptotic protein. | Mutations in the myocilin gene (MYOC) are causative for 10% of cases with
juvenile open-angle glaucoma and 3-4% of those with primary open-angle glaucoma.
Myocilin is a secreted protein with relatively ill-defined matricellular
properties. Despite its high expression in the eye, myocilin-deficient mice have
originally been reported to have no obvious ocular phenotype. Here we revisited
the ocular phenotype of myocilin-deficient mice and detected a higher number of
neurons in their inner (INL) and outer (ONL) nuclear layers, as well as a higher
number of retinal ganglion cells (RGC) and their axons. The increase in retinal
neurons appears to be caused by a decrease in programmed developmental cell
death, as apoptosis of retinal neurons between postnatal days 4 and 10 was found
to be attenuated when compared to that of wildtype littermates. In contrast,
when Myoc(-/-) mice were crossed with βB1-crystallin-MYOC mice with ectopic
overexpression of myocilin in the eye, no differences in developmental
apoptosis, RGC number and INL thickness were observed when compared to wildtype
littermates. The amounts of the anti-apoptotic Bcl-2-like protein 1 (BCL2L1,
Bcl-xL) and its mRNA were increased in retinae of Myoc(-/-) mice, while lower
amounts of BCL2L1 and its mRNA were detected in mixed
Myoc(-/-)/βB1-crystallin-MYOC mice. The structural differences between Myoc(-/-)
mice and wildtype littermates did not result in functional differences as
measured by electroretinography. Noteworthy though mixed
Myoc(-/-)/βB1-crystallin-MYOC mice with ocular overexpression of myocilin had
significant cone function deficits. Myocilin appears to modulate apoptotic death
of retinal neurons likely by interacting with the intrinsic apoptotic pathway. The aim of the present study was to investigate licochalcone-E (Lico-E)-induced
apoptosis and the associated apoptotic signaling pathway in FaDu cells, a human
pharyngeal squamous carcinoma cell line. Treatment with Lico-E exhibited
significant cytotoxicity on FaDu cells in a concentration-dependent manner. The
IC50 value of Lico-E in FaDu cells was ~50 µM. Treatment with Lico-E increased
the number of dead FaDu cells. Furthermore, chromatin condensation, which is
associated with apoptotic cell death, was observed in FaDu cells treated with
Lico-E for 24 h. By contrast, Lico-E did not produce cytotoxicity or increase
the number of dead cells when applied to human normal oral keratinocytes
(hNOKs). Furthermore, chromatin condensation was not observed in hNOKs treated
with Lico-E. Treatment with Lico-E increased the expression of Fas ligand and
the cleaved form of caspase-8 in FaDu cells. Furthermore, treatment with Lico-E
increased the expression of pro-apoptotic factors, including apoptosis regulator
BAX, Bcl-2-associated agonist of cell death, apoptotic protease-activating
factor 1, caspase-9 and tumor suppressor p53, while decreasing the expression of
anti-apoptotic factors, including apoptosis regulator Bcl-2 and Bcl-2-like
protein 1 in FaDu cells. The expression of cleaved caspases-3 and poly
(ADP-ribose) polymerase was significantly upregulated following treatment with
Lico-E in FaDu cells, while Lico-E-induced apoptotic FaDu cell death was
partially suppressed by treatment with Z-VAD-FMK, a pan caspase inhibitor.
Therefore, Lico-E-induced oral cancer (OC) cell-specific apoptosis is mediated
by the death receptor-dependent extrinsic and mitochondrial-dependent intrinsic
apoptotic signaling pathways. In conclusion, these data suggested that Lico-E
exhibits potential chemopreventive effects and warrants further developed as a
chemotherapeutic agent against OC. |
Glucoraphanin from broccoli can help reduce obesity , yes or no? | Yes, there is evidence that glucoraphanin from broccoli can help reduce obesity. | Obesity is a low-grade sustained inflammatory state that causes oxidative stress
in different metabolic tissues, which leads to insulin resistance and
nonalcoholic fatty liver disease (NAFLD). Particularly, obesity-induced
metabolic endotoxemia plays an important role in the pathogenesis of insulin
resistance and inflammation. Nuclear factor erythroid 2-related factor 2 (Nrf2)
is a key regulator of antioxidant signaling that serves as a primary cellular
defense against the cytotoxic effects of oxidative stress. Pharmacological
stimulation of Nrf2 mitigates obesity and insulin resistance in mice; however,
Nrf2 activators are not clinically available due to biosafety concerns. A recent
study demonstrated that glucoraphanin, a precursor of the Nrf2 activator
sulforaphane, ameliorates obesity by enhancing energy expenditure and browning
of white adipose tissue, and attenuates obesity-related inflammation and insulin
resistance by polarizing M2 macrophages and reducing metabolic endotoxemia.
Thus, this review focuses on the efficiency and safety of glucoraphanin in
alleviating obesity, insulin resistance, and NAFLD. Abbreviations: ALT, Alanine
aminotransferase; AMPK, AMP-activated protein kinase; ATMs, Adipose tissue
macrophages; BAT, Brown adipose tissue; CDDO-Im,
2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid-imidazolide; CDDO-Me, CDDO-methyl
ester; DIO, High-fat-diet-induced obese; FFA, Free fatty acid; FGF, Fibroblast
growth factor; GTP, Glutamyl transpeptidase; HFD, High-fat diet; IKKβ, Inhibitor
of κB-kinase β; IL, Interleukin; JNK, C-Jun N-terminal kinase; KD, Knockdown;
Keap1, Kelch-like ECH-associated protein 1; KO, Knockout; LPS,
Lipopolysaccharide; NADPH, Nicotinamide adenine dinucleotide phosphate; NAFLD,
Non-alcoholic fatty liver disease; NF-κB, Nuclear factor-κB; Nrf2, Nuclear
factor E2-related factor 2; ROS, Reactive oxygen species; T2D, Type 2 diabetes;
TLR, Toll-like receptor; TNF, tumor necrosis factor; UCP, Uncoupling protein;
WAT, White adipose tissue. |
What is the function of the zelda transcription factor in D. melanogaster? | The zinc-finger TF zelda (zld) is essential for the maternal-to-zygotic transition (MZT) in Drosophila melanogaster, where it directly binds over thousand cis-regulatory modules to regulate chromatin accessibility. | In all animals, the initial events of embryogenesis are controlled by maternal
gene products that are deposited into the developing oocyte. At some point after
fertilization, control of embryogenesis is transferred to the zygotic genome in
a process called the maternal-to-zygotic transition. During this time, many
maternal RNAs are degraded and transcription of zygotic RNAs ensues. There is a
long-standing question as to which factors regulate these events. The recent
findings that microRNAs and Smaug mediate maternal transcript degradation have
shed new light on this aspect of the problem. However, the transcription
factor(s) that activate the zygotic genome remain elusive. The discovery that
many of the early transcribed genes in Drosophila share a cis-regulatory
heptamer motif, CAGGTAG and related sequences, collectively referred to as
TAGteam sites raised the possibility that a dedicated transcription factor could
interact with these sites to activate transcription. Here we report that the
zinc-finger protein Zelda (Zld; Zinc-finger early Drosophila activator) binds
specifically to these sites and is capable of activating transcription in
transient transfection assays. Mutant embryos lacking zld are defective in
cellular blastoderm formation, and fail to activate many genes essential for
cellularization, sex determination and pattern formation. Global expression
profiling confirmed that Zld has an important role in the activation of the
early zygotic genome and suggests that Zld may also regulate maternal RNA
degradation during the maternal-to-zygotic transition. BACKGROUND: In embryos the maternal-to-zygotic transition (MTZ) integrates
post-transcriptional regulation of maternal transcripts with transcriptional
activation of the zygotic genome. Although the molecular mechanisms underlying
this event are being clarified in Drosophila melanogaster, little is know about
the embryogenic processes in other insect species. The recent publication of
expressed sequence tags (ESTs) from embryos of the global pest species Ceratitis
capitata (medfly) has enabled the investigation of embryogenesis in this species
and has allowed a comparison of the embryogenic processes in these two related
dipteran species, C. capitata and D. melanogaster, that shared a common ancestor
80-100 mya.
RESULTS: Using a novel PCR-based sexing method, which takes advantage of a
putative LTR retrotransposon MITE insertion on the medfly Y chromosome, the
transcriptomes of individual early male and female embryos were analysed using
RT-PCR. This study is focused on two crucial aspects of the onset of embryonic
development: sex determination and cellular blastoderm formation. Together with
the three known medfly genes (Cctransformer, Cctransformer2 and Ccdoublesex),
the expression patterns of other medfly genes that are similar to the D.
melanogaster sex-determination genes (sisterlessA, groucho, deadpan, Sex-lethal,
female lethal d, sans fille and intersex) and four cellular blastoderm formation
genes (Rho1, spaghetti squash, slow-as-molasses and serendipity-alpha) were
analyzed, allowing us to sketch a preliminary outline of the embryonic process
in the medfly. Furthermore, a putative homologue of the Zelda gene has been
considered, which in D. melanogaster encodes a DNA-binding factor responsible
for the maternal-to-zygotic transition.
CONCLUSIONS: Our novel sexing method facilitates the study of i) when the MTZ
transition occurs in males and females of C. capitata, ii) when and how the
maternal information of "female-development" is reprogrammed in the embryos and
iii) similarities and differences in the regulation of gene expression in C.
capitata and D. melanogaster. We suggest a new model for the onset of the sex
determination cascade in the medfly: the maternally inherited Cctra transcripts
in the female embryos are insufficient to produce enough active protein to
inhibit the male mode of Cctra splicing. The slow rate of development and the
inefficiency of the splicing mechanism in the pre-cellular blastoderm
facilitates the male-determining factor (M) activity, which probably acts by
inhibiting CcTRA protein activity. Maternally contributed mRNAs and proteins control the initial stages of
development following fertilization. During this time, most of the zygotic
genome remains transcriptionally silent. The initiation of widespread zygotic
transcription is coordinated with the degradation of maternally provided mRNAs
at the maternal-to-zygotic transition (MZT). While most of the genome is
silenced prior to the MZT, a small subset of zygotic genes essential for the
future development of the organism is transcribed. Previous work in our
laboratory and others identified the TAGteam element, a set of related
heptameric DNA-sequences in the promoters of many early-expressed Drosophila
genes required to drive their unusually early transcription. To understand how
this unique subset of genes is regulated, we identified a TAGteam-binding factor
Grainyhead (Grh). We demonstrated that Grh and the previously characterized
transcriptional activator Zelda (Zld) bind to different TAGteam sequences with
varying affinities, and that Grh competes with Zld for TAGteam occupancy.
Moreover, overexpression of Grh in the early embryo causes defects in cell
division, phenocopying Zld depletion. Our findings indicate that during early
embryonic development the precise timing of gene expression is regulated by both
the sequence of the TAGteam elements in the promoter and the relative levels of
the transcription factors Grh and Zld. The earliest stages of development in most metazoans are driven by maternally
deposited proteins and mRNAs, with widespread transcriptional activation of the
zygotic genome occurring hours after fertilization, at a period known as the
maternal-to-zygotic transition (MZT). In Drosophila, the MZT is preceded by the
transcription of a small number of genes that initiate sex determination,
patterning, and other early developmental processes; and the zinc-finger protein
Zelda (ZLD) plays a key role in their transcriptional activation. To better
understand the mechanisms of ZLD activation and the range of its targets, we
used chromatin immunoprecipitation coupled with high-throughput sequencing
(ChIP-Seq) to map regions bound by ZLD before (mitotic cycle 8), during (mitotic
cycle 13), and after (late mitotic cycle 14) the MZT. Although only a handful of
genes are transcribed prior to mitotic cycle 10, we identified thousands of
regions bound by ZLD in cycle 8 embryos, most of which remain bound through
mitotic cycle 14. As expected, early ZLD-bound regions include the promoters and
enhancers of genes transcribed at this early stage. However, we also observed
ZLD bound at cycle 8 to the promoters of roughly a thousand genes whose first
transcription does not occur until the MZT and to virtually all of the thousands
of known and presumed enhancers bound at cycle 14 by transcription factors that
regulate patterned gene activation during the MZT. The association between early
ZLD binding and MZT activity is so strong that ZLD binding alone can be used to
identify active promoters and regulatory sequences with high specificity and
selectivity. This strong early association of ZLD with regions not active until
the MZT suggests that ZLD is not only required for the earliest wave of
transcription but also plays a major role in activating the genome at the MZT. In past years, much attention has focused on the gene networks that regulate
early developmental processes, but less attention has been paid to how multiple
networks and processes are temporally coordinated. Recently the discovery of the
transcriptional activator Zelda (Zld), which binds to CAGGTAG and related
sequences present in the enhancers of many early-activated genes in Drosophila,
hinted at a mechanism for how batteries of genes could be simultaneously
activated. Here we use genome-wide binding and expression assays to identify Zld
target genes in the early embryo with the goal of unraveling the gene circuitry
regulated by Zld. We found that Zld binds to genes involved in early
developmental processes such as cellularization, sex determination,
neurogenesis, and pattern formation. In the absence of Zld, many target genes
failed to be activated, while others, particularly the patterning genes,
exhibited delayed transcriptional activation, some of which also showed weak
and/or sporadic expression. These effects disrupted the normal sequence of
patterning-gene interactions and resulted in highly altered spatial expression
patterns, demonstrating the significance of a timing mechanism in early
development. In addition, we observed prevalent overlap between Zld-bound
regions and genomic "hotspot" regions, which are bound by many developmental
transcription factors, especially the patterning factors. This, along with the
finding that the most over-represented motif in hotspots, CAGGTA, is the Zld
binding site, implicates Zld in promoting hotspot formation. We propose that Zld
promotes timely and robust transcriptional activation of early-gene networks so
that developmental events are coordinated and cell fates are established
properly in the cellular blastoderm embryo. The transcription factor Zelda plays a pivotal role in promoting the maternal to
zygotic transition during embryogenesis in Drosophila melanogaster. However,
little is known about its role later in development. Here we are showing that
Zelda is essential for proper wing development through gain and loss of function
experiments. Zelda's transcript variants RB, RC and RD are present in imaginal
wing discs of third instar larvae and the production of 2 protein isoforms of
∼180 and ∼70kD was detected in the same tissue. In ChIP experiments using larval
wing discs, Zelda was found to bind to a region of the optomotor-blind gene,
suggesting an interaction with a Dpp target that promotes wing growth and
patterning. Zygotic genome activation (ZGA) is a major genome programming event whereby the
cells of the embryo begin to adopt specified fates. Experiments in Drosophila
and zebrafish have revealed that ZGA depends on transcription factors that
provide large-scale control of gene expression by direct and specific binding to
gene regulatory sequences. Zelda (Zld) plays such a role in the Drosophila
embryo, where it has been shown to control the action of patterning signals;
however, the mechanisms underlying this effect remain largely unclear. A recent
model proposed that Zld binding sites act as quantitative regulators of the
spatiotemporal expression of genes activated by Dorsal (Dl), the morphogen that
patterns the dorsoventral axis. Here we tested this model experimentally, using
enhancers of brinker (brk) and short gastrulation (sog), both of which are
directly activated by Dl, but at different concentration thresholds. In
agreement with the model, we show that there is a clear positive correlation
between the number of Zld binding sites and the spatial domain of enhancer
activity. Likewise, the timing of expression could be advanced or delayed. We
present evidence that Zld facilitates binding of Dl to regulatory DNA, and that
this is associated with increased chromatin accessibility. Importantly, the
change in chromatin accessibility is strongly correlated with the change in Zld
binding, but not Dl. We propose that the ability of genome activators to
facilitate readout of transcriptional input is key to widespread transcriptional
induction during ZGA. In nearly all metazoans, the earliest stages of development are controlled by
maternally deposited mRNAs and proteins. The zygotic genome becomes
transcriptionally active hours after fertilization. Transcriptional activation
during this maternal-to-zygotic transition (MZT) is tightly coordinated with the
degradation of maternally provided mRNAs. In Drosophila melanogaster, the
transcription factor Zelda plays an essential role in widespread activation of
the zygotic genome. While Zelda expression is required both maternally and
zygotically, the mechanisms by which it functions to remodel the embryonic
genome and prepare the embryo for development remain unclear. Using
Cas9-mediated genome editing to generate targeted mutations in the endogenous
zelda locus, we determined the functional relevance of protein domains conserved
amongst Zelda orthologs. We showed that neither a conserved N-terminal zinc
finger nor an acidic patch were required for activity. Similarly, a previously
identified splice isoform of zelda is dispensable for viability. By contrast, we
identified a highly conserved zinc-finger domain that is essential for the
maternal, but not zygotic functions of Zelda. Animals homozygous for mutations
in this domain survived to adulthood, but embryos inheriting these
loss-of-function alleles from their mothers died late in embryogenesis. These
mutations did not interfere with the capacity of Zelda to activate transcription
in cell culture. Unexpectedly, these mutations generated a hyperactive form of
the protein and enhanced Zelda-dependent gene expression. These data have
defined a protein domain critical for controlling Zelda activity during the MZT,
but dispensable for its roles later in development, for the first time
separating the maternal and zygotic requirements for Zelda. This demonstrates
that highly regulated levels of Zelda activity are required for establishing the
developmental program during the MZT. We propose that tightly regulated gene
expression is essential to navigate the MZT and that failure to precisely
execute this developmental program leads to embryonic lethality. Connecting the developmental patterning of tissues to the mechanistic control of
RNA polymerase II remains a long-term goal of developmental biology. Many key
elements have been identified in the establishment of spatial-temporal control
of transcription in the early Drosophila embryo, a model system for
transcriptional regulation. The dorsal-ventral axis of the Drosophila embryo is
determined by the graded distribution of Dorsal (Dl), a homolog of the nuclear
factor κB (NF-κB) family of transcriptional activators found in humans [1, 2]. A
second maternally deposited factor, Zelda (Zld), is uniformly distributed in the
embryo and is thought to act as a pioneer factor, increasing enhancer
accessibility for transcription factors, such as Dl [3-9]. Here, we utilized the
MS2 live imaging system to evaluate the expression of the Dl target gene short
gastrulation (sog) to better understand how a pioneer factor affects the kinetic
parameters of transcription. Our experiments indicate that Zld modifies
probability of activation, the timing of this activation, and the rate at which
transcription occurs. Our results further show that this effective rate increase
is due to an increased accumulation of Dl at the site of transcription,
suggesting that transcription factor "hubs" induced by Zld [10] functionally
regulate transcription. In the endopterygote Drosophila melanogaster, Zelda is an activator of the
zygotic genome during the maternal-to-zygotic transition (MZT). Zelda binds
cis-regulatory elements (TAGteam heptamers), making chromatin accessible for
gene transcription. Zelda has been studied in other endopterygotes:
Apis mellifera and Tribolium castaneum, and the paraneopteran Rhodnius prolixus.
We studied Zelda in the cockroach Blattella germanica, a hemimetabolan, short
germ-band, and polyneopteran species. B. germanica Zelda has the complete set of
functional domains, which is typical of species displaying ancestral features
concerning embryogenesis. Interestingly, we found D. melanogaster TAGteam
heptamers in the B. germanica genome. The canonical one, CAGGTAG, is present at
a similar proportion in the genome of these two species and in the genome of
other insects, suggesting that the genome admits as many CAGGTAG motifs as its
length allows. Zelda-depleted embryos of B. germanica show defects involving
blastoderm formation and abdomen development, and genes contributing to these
processes are down-regulated. We conclude that in B. germanica, Zelda strictly
activates the zygotic genome, within the MZT, a role conserved in more derived
endopterygote insects. In B. germanica, zelda is expressed during MZT, whereas
in D. melanogaster and T. castaneum it is expressed beyond this transition. In
these species and A. mellifera, Zelda has functions even in postembryonic
development. The expansion of zelda expression beyond the MZT in endopterygotes
might be related with the evolutionary innovation of holometabolan
metamorphosis. DATABASES: The RNA-seq datasets of B. germanica, D. melanogaster,
and T. castaneum are accessible at the GEO databases GSE99785, GSE18068,
GSE63770, and GSE84253. In addition, the RNA-seq library from T. castaneum adult
females is available at SRA: SRX021963. The B. germanica reference genome is
available as BioProject PRJNA203136. |
What does tsDMARD stand for? | tsDMARDs are targeted synthetic disease-modifying antirheumatic drugs. | The treatment of rheumatoid arthritis (RA) has been transformed with the
introduction of biologic disease modifying anti-rheumatic drugs (bDMARD) and
more recently, targeted synthetic DMARD (tsDMARD) therapies in the form of
janus-kinase inhibitors. Nevertheless, response to these agents varies such that
a trial and error approach is adopted; leading to poor patient quality of life,
and long-term outcomes. There is thus an urgent need to identify effective
biomarkers to guide treatment selection. A wealth of research has been invested
in this field but with minimal progress. Increasingly recognized is the
importance of evaluating synovial tissue, the primary site of RA, as opposed to
peripheral blood-based investigation. In this mini-review, we summarize the
literature supporting synovial tissue heterogeneity, the conceptual basis for
stratified therapy. This includes recognition of distinct synovial
pathobiological subtypes and associated molecular pathways. We also review
synovial tissue studies that have been conducted to evaluate the effect of
individual bDMARD and tsDMARD on the cellular and molecular characteristics,
with a view to identifying tissue predictors of response. Initial observations
are being brought into the clinical trial landscape with stratified biopsy
trials to validate toward implementation. Furthermore, development of tissue
based omics technology holds still more promise in advancing our understanding
of disease processes and guiding future drug selection. OBJECTIVES: Abatacept, a biologic DMARD, was associated with respiratory adverse
events in a small subgroup of RA patients with chronic obstructive pulmonary
disease (COPD) in a trial. Whether this potential risk is specific to abatacept
or extends to all biologics and targeted synthetic DMARDs (tsDMARDs) is unclear.
We assessed the risk of adverse respiratory events associated with biologic and
tsDMARDs compared with conventional synthetic DMARDs (csDMARDs) among RA
patients with concomitant COPD in a large, real-world cohort.
METHODS: We used a prevalent new-user design to study RA patients with COPD in
the US-based MarketScan databases. New users of biologic DMARDs and/or tsDMARDs
were matched on time-conditional propensity scores to new users of csDMARDs.
Adverse respiratory events were estimated using Cox models comparing current use
of biologic/tsDMARDs with csDMARDs.
RESULTS: The cohort included 7424 patients initiating biologic/tsDMARDs and 7424
matched patients initiating csDMARDs. The adjusted hazard ratio of hospitalized
COPD exacerbation comparing biologic/tsDMARD vs csDMARD was 0.76 (95% CI: 0.55,
1.06), while it was 1.02 (95% CI: 0.82, 1.27) for bronchitis, 1.21 (95% CI:
0.92, 1.58) for hospitalized pneumonia or influenza and 0.99 (95% CI: 0.87,
1.12) for outpatient pneumonia or influenza. The hazard ratio of the combined
end point of COPD exacerbation, bronchitis and hospitalized pneumonia or
influenza was 1.04 (95% CI: 0.89, 1.21).
CONCLUSION: In this large, real-world comparative safety study, biologic and
tsDMARDs, including abatacept, were not associated with an increased risk of
adverse respiratory events when compared with csDMARDs in patients with RA and
COPD. OBJECTIVE: To provide real-world evidence about the reasons why Australian
rheumatologists cease biologic (b) and targeted synthetic (ts) disease-modifying
antirheumatic drugs (DMARD) when treating patients with rheumatoid arthritis
(RA), and to assess (1) the primary failure rate for first-line treatment, and
(2) the persistence on second-line treatments in patients who stopped first-line
tumor necrosis factor inhibitors (TNFi).
METHODS: This is a multicenter retrospective, noninterventional study of
patients with RA enrolled in the Australian Optimising Patient outcome in
Australian RheumatoLogy (OPAL) dataset with a start date of b/tsDMARD between
August 1, 2010, and June 30, 2017. Primary failure was defined as stopping
treatment within 6 months of treatment initiation.
RESULTS: Data from 7740 patients were analyzed; 6914 patients received
first-line b/tsDMARD. First-line treatment was stopped in 3383 (49%) patients;
1263 (37%) were classified as primary failures. The most common reason was "lack
of efficacy" (947/2656, 36%). Of the patients who stopped first-line TNFi, 43%
(1111/2560) received second-line TNFi, which resulted in the shortest median
time to stopping second-line treatment (11 months, 95% CI 9-12) compared with
non-TNFi. The longest second-line median treatment duration after first-line
TNFi was for patients receiving rituximab (39 months, 95% CI 27-74).
CONCLUSION: A large proportion of patients who stopped first-line TNFi therapy
received another TNFi despite evidence for longer treatment persistence on
second-line b/tsDMARD with a different mode of action. Lack of efficacy was
recorded as the most common reason for making a switch in first-line treatment
of patients with RA. In recent years tremendous progress has been made in the therapeutic management
of rheumatoid arthritis. Rheumatologists now have a large armamentarium of
highly efficient drugs with different mechanisms of action at their disposal.
These new drugs consist of biologicals (biological disease-modifying
antirheumatic drugs, bDMARDs) as well as targeted synthetic DMARDs (tsDMARD). A
common feature of these new drugs for treatment of rheumatoid arthritis is that
the molecular target of the drug is known, which is not the case for
conventional DMARDs. With the help of the new drugs, the therapeutic goal of
inducing remission in patients with rheumatoid arthritis has become reality for
many patients. Nevertheless, there is still a significant proportion of patients
who do not adequately respond to all available drugs, leaving room for still
further improvement. This review gives a short overview on the currently
available and effective substances for the treatment of rheumatoid arthritis. The progress in the understanding of the pathophysiology of rheumatic diseases
provided a rational basis for the development of biologic disease‑modifying
antirheumatic drugs (bDMARDs) and targeted synthetic DMARDs (tsDMARDs), which
have completely revolutionized the treatment of inflammatory conditions. These
agents differ in terms of their effectiveness for controlling specific rheumatic
diseases depending on the pivotal cytokine driving the inflammatory process.
Cytokine blockers were the first to be developed and rapidly expanded. They
include agents that act against tumor necrosis factor α (TNF‑α) (etanercept,
infliximab, adalimumab, golimumab, and certolizumab pegol) and interleukin (IL)
6 (tocilizumab and sarilumab), IL‑1 (anakinra, canakinumab, and rilonacept),
IL‑17 (secukinumab and ixekizumab), and IL-12/23 (ustekinumab) receptors.
Lymphocyte‑targeting agents include rituximab and belimumab, which act against B
cells by different mechanisms, and abatacept, which is a T cell costimulation
modulator. tsDMARDs, also known as small‑molecule inhibitors, are oral drugs
based on a novel strategy to treat inflammatory diseases. Janus kinase (JAK)
inhibitors (tofacitinib, baricitinib, and upadacitinib) and phosphodiesterase 4
inhibitors (apremilast) form this group. The major concern with the use of
bDMARDs and tsDMARDs is a higher risk of infections. Performance of blood tests
as well as screening for tuberculosis and hepatitis viral infection are
mandatory prior to biologic therapy initiation. Adherence to an immunization
program is also recommended. Whenever possible, the choice of bDMARDs and
tsDMARDs should be guided by the patient's comorbidities. There have been
limited data on the use of these drugs during pregcy, but anti‑TNF‑α therapy,
rituximab, and anakinra seem to be safe. Biologic agents are expensive, but
biosimilars have emerged as a cost‑effective option with a potential to treat a
greater number of patients. |
Which conditions are manifested by TRIM8 mutations? | TRIM8 mutations are associated with epilepsy, epileptic encephalopathy, developmental delay and intellectual disability. | Mutations in the TRIM8 gene have been described in patients with severe
developmental delay, intellectual disability and epilepsy. Only six patients
have been described to date. All the previous mutations were truncating variants
clustered in the C-terminus of the protein. A previous patient with
TRIM8-related epileptic encephalopathy was reported to have nephrotic syndrome.
Here we describe the clinical, radiological and histological features of an
8-year-old male patient with a TRIM8 mutation who, in contrast to previous
patients, had only mild intellectual disability and well-controlled epilepsy.
The patient was found to have proteinuria at 2 years of age. Renal biopsy
findings were suggestive of focal segmental glomerulosclerosis. His kidney
function declined and peritoneal dialysis was started at 5 years of age. He
underwent renal transplant at 7 years of age. Trio-based whole genome sequencing
identified a novel de novo heterozygous frameshift mutation in TRIM8
(NM_030912.2) c.1198_1220del, p.(Tyr400ArgfsTer2). This patient is further
evidence that TRIM8 mutations cause a syndrome with both neurological and renal
features. Our findings suggest the spectrum of TRIM8-related disease may be
wider than previously thought with the possibility of milder neurodevelopmental
problems and/or a more severe, progressive renal phenotype. We highlight the
need for proteinuria screening in patients with TRIM8 mutations. |
What is LY-CoV555? | LY-CoV555 is an anti-spike neutralizing antibody targeting the SARS-CoV-2 that has been tested for patients with Covid-19. | BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes
coronavirus disease 2019 (Covid-19), which is most frequently mild yet can be
severe and life-threatening. Virus-neutralizing monoclonal antibodies are
predicted to reduce viral load, ameliorate symptoms, and prevent
hospitalization.
METHODS: In this ongoing phase 2 trial involving outpatients with recently
diagnosed mild or moderate Covid-19, we randomly assigned 452 patients to
receive a single intravenous infusion of neutralizing antibody LY-CoV555 in one
of three doses (700 mg, 2800 mg, or 7000 mg) or placebo and evaluated the
quantitative virologic end points and clinical outcomes. The primary outcome was
the change from baseline in the viral load at day 11. The results of a
preplanned interim analysis as of September 5, 2020, are reported here.
RESULTS: At the time of the interim analysis, the observed mean decrease from
baseline in the log viral load for the entire population was -3.81, for an
elimination of more than 99.97% of viral RNA. For patients who received the
2800-mg dose of LY-CoV555, the difference from placebo in the decrease from
baseline was -0.53 (95% confidence interval [CI], -0.98 to -0.08; P = 0.02), for
a viral load that was lower by a factor of 3.4. Smaller differences from placebo
in the change from baseline were observed among the patients who received the
700-mg dose (-0.20; 95% CI, -0.66 to 0.25; P = 0.38) or the 7000-mg dose (0.09;
95% CI, -0.37 to 0.55; P = 0.70). On days 2 to 6, the patients who received
LY-CoV555 had a slightly lower severity of symptoms than those who received
placebo. The percentage of patients who had a Covid-19-related hospitalization
or visit to an emergency department was 1.6% in the LY-CoV555 group and 6.3% in
the placebo group.
CONCLUSIONS: In this interim analysis of a phase 2 trial, one of three doses of
neutralizing antibody LY-CoV555 appeared to accelerate the natural decline in
viral load over time, whereas the other doses had not by day 11. (Funded by Eli
Lilly; BLAZE-1 ClinicalTrials.gov number, NCT04427501.). Facing the COVID-19 global healthcare crisis, scientists worldwide are
collaborating to develop prophylactic and therapeutic interventions against the
disease. Antibody therapeutics hold enormous promise for the treatment of
COVID-19. In March 2020, the Chinese Antibody Society, in collaboration with The
Antibody Society, initiated the "COVID-19 Antibody Therapeutics Tracker"
("Tracker") (https://chineseantibody.org/covid-19-track/) program to track the
antibody-based COVID-19 interventions in preclinical and clinical development
globally. The data are collected from the public domain and verified by
volunteers on an ongoing basis. Here, we present exploratory data analyses and
visualization to demonstrate the latest trends of COVID-19 antibody development,
based on data for over 150 research and development programs and molecules
included in the "Tracker" as of 8 August 2020. We categorized the data mainly by
their targets, formats, development status, developers and country of origin.
Although details are limited in some cases, all of the anti-SARS-CoV-2 antibody
candidates appear to target the viral spike protein (S protein), and most are
full-length monoclonal antibodies. Most of the current COVID-19 antibody
therapeutic candidates in clinical trials are repurposed drugs aimed at targets
other than virus-specific proteins, while most of these virus-specific
therapeutic antibodies are in discovery or preclinical studies. As of 8 August
2020, eight antibody candidates targeting the SARS-CoV-2 S protein have entered
clinical studies, including LY-CoV555, REGN-COV2, JS016, TY027, CT-P59,
BRII-196, BRII-198 and SCTA01. Ongoing clinical trials of SARS-CoV-2
neutralizing antibodies will help define the utility of these antibodies as a
new class of therapeutics for treating COVID-19 and future coronavirus
infections. |
List blood marker for Non-Hodgkin lymphoma. | Soluble interleukin-2 receptor-α, CXC chemokine ligand 13, soluble CD30, and soluble tumor necrosis factor receptor-2 were individually positively associated, and B-cell activating factor of the tumor necrosis factor family inversely associated, with all non-Hodgkin lymphoma and one or more subtypes.
GALECTIN-3 AS A PROGNOSTIC BIOMARKER IN PATIENTS WITH NON-HODGKIN LYMPHOMA. | The aim of the study - to evaluate the prognostic value of galectin-3 for
cumulative survival in patients with controlled non-Hodgkin lymphoma. Eighty two
out subjects with full or partial remission of non-Hodgkin lymphoma were
enrolled in the study. Observation period was up to 12 months. Blood samples for
biomarkers measurements were collected. ELISA method for measurements of
circulating level of Gal-3 and NT-pro-brain natriuretic peptide (NT-proBNP) was
used. Hemodynamic evaluation was performed by transthoracic echocardiography.
Fifty five cumulative clinical events occurred in 21 patients (25.6%) within the
follow-up, with their distribution being as follows: 5 cardiovascular deaths, 24
cardiac arrhythmias, 8 cardiac ischemic events, 3 strokes, 9 chronic heart
failures and 6 hospital admissions for cardiovascular reasons. Circulating
levels of Gal-3 in subjects without and with cardiovascular events were 5.37
ng/ml (95% confidence interval [CI]=2.90-7.85 ng/ml) and 13.97 ng/ml (95%
CI=7.82-20.11 ng/ml) (P=0.03) respectively. The results of regression analysis
showed directly related circulating Gal-3 with E/Em (r=0.45, P=0.045), T2DM
(r=0.38, P=0.01). Multivariate logistic regression revealed independent
predictive value of circulating Gal-3 for 12 months cumulative cardiovascular
events (odds ratio [OR]=1.11; 95% CI=1.05-1.25; P=0.005). In fact, Gal-3,
NT-pro-BNP, GFR, and LVEF remained statistically significant predictors for
cumulative cardiovascular events, whereas T2DM, hypertension, obesity did not.
Increased circulating Gal-3 associates with increased 12 months cumulative
cardiovascular events among patients with documented non-Hodgkin lymphoma. Inflammation and B-cell hyperactivation have been associated with non-Hodgkin
lymphoma development. This prospective analysis aimed to further elucidate
pre-diagnosis plasma immune marker profiles associated with non-Hodgkin lymphoma
risk. We identified 598 incident lymphoma cases and 601 matched controls in
Nurses' Health Study and Health Professionals Follow-up Study participants with
archived pre-diagnosis plasma samples and measured 13 immune marker levels with
multiplexed immunoassays. Using multivariable logistic regression we calculated
Odds Ratios (OR) and 95% Confidence Intervals (CI) per standard deviation unit
increase in biomarker concentration for risk of non-Hodgkin lymphoma and major
histological subtype, stratifying additional models by years (<5, 5 to <10, ≥10)
after blood draw. Soluble interleukin-2 receptor-α, CXC chemokine ligand 13,
soluble CD30, and soluble tumor necrosis factor receptor-2 were individually
positively associated, and B-cell activating factor of the tumor necrosis factor
family inversely associated, with all non-Hodgkin lymphoma and one or more
subtypes. The biomarker combinations associated independently with lymphoma
varied somewhat by subtype and years after blood draw. Of note, the unexpected
inverse association between B-cell activating factor and chronic lymphocytic
leukemia/small lymphocytic lymphoma risk (OR: 95%CI: 0.51, 0.43-0.62) persisted
more than ten years after blood draw (OR: 0.70; 95%CI: 0.52-0.93). In
conclusion, immune activation precedes non-Hodgkin lymphoma diagnosis by several
years. Decreased B-cell activating factor levels may denote nascent chronic
lymphocytic leukemia many years pre-diagnosis. |
Are bacteria in the genus Clostridium facultative anaerobes? | Clostridia belong to those bacteria which are considered as obligate anaerobe, e.g. oxygen is harmful or lethal to these bacteria. | Clostridia belong to those bacteria which are considered as obligate anaerobe,
e.g. oxygen is harmful or lethal to these bacteria. Nevertheless, it is known
that they can survive limited exposure to air, and often eliminate oxygen or
reactive derivatives via NAD(P)H-dependent reduction. This system does
apparently contribute to survival after oxidative stress, but is insufficient to
establish long-term tolerance of aerobic conditions. Here we show that
manipulation of the regulatory mechanism of this defence mechanism can trigger
aerotolerance in the obligate anaerobe Clostridium acetobutylicum. Deletion of a
peroxide repressor (PerR)-homologous protein resulted in prolonged
aerotolerance, limited growth under aerobic conditions and rapid consumption of
oxygen from an aerobic environment. The mutant strain also revealed higher
resistance to H2O2 and activities of NADH-dependent scavenging of H2O2 and
organic peroxides in cell-free extracts increased by at least one order of
magnitude. Several genes encoding the putative enzymes were upregulated and
identified as members of the clostridial PerR regulon, including the heat shock
protein Hsp21, a reverse rubrerythrin which was massively produced and became
the most abundant protein in the absence of PerR. This multifunctional protein
is proposed to play the crucial role in the oxidative stress defence. Cancer has become the second ranking cause of death in the industrialized world.
Conventional anti-cancer therapies such as surgery, radiotherapy, and
chemotherapy are effective in the treatment of solid tumors only to some extent.
Furthermore, they are often associated with severe side effects. Use of bacteria
as alternative cancer therapeutics has sporadically been followed over more than
a century. The potential to target and colonize solid tumors could be shown for
many different bacteria in the meantime. Such bacteria are either obligate
anaerobic bacteria like Clostridium or Bifidobacterium or facultative anaerobic
like Escherichia coli or Salmonella. Here we describe bacterial strains that
were successfully applied mostly in animals bearing model tumors, although first
clinical trials have been reported as well. Our review mainly concentrates on
Salmonella enterica serovar Typhimurium (S. Typhimurium) since these bacteria
were studied most intensively thus far. Importantly, S. Typhimurium were shown
not only to colonize large established tumors but also exhibit the property to
invade and affect metastases. We report on a potential mechanism by which such
bacteria can invade solid tumors. Furthermore, we describe several successful
attempts in which the bacteria have been used as carriers for recombit
therapeutic molecules that render bacteria more powerful in eradication of the
established tumor. Such attempts should be considered starting points on the way
to an effective and safe tumor therapy with the help of bacteria. Author information:
(1)Genomic and Applied Microbiology & Göttingen Genomics Laboratory,
Georg-August-University Göttingen, Göttingen, Germany.
(2)The Clostridia Research Group, BBSRC/EPSRC Synthetic Biology Research Centre,
School of Life Sciences, Centre for Biomolecular Sciences, University of
Nottingham, Nottingham, United Kingdom.
(3)Unilever, Research and Development, Bedford, United Kingdom.
(4)The Clostridia Research Group, BBSRC/EPSRC Synthetic Biology Research Centre,
School of Life Sciences, Centre for Biomolecular Sciences, University of
Nottingham, Nottingham, United Kingdom [email protected]. Clostridium difficile is a spore-forming obligate anaerobe that is a leading
cause of healthcare-associated infections. C. difficile infections begin when
its metabolically dormant spores germinate in the gut of susceptible
individuals. Binding of bile salt germits to the Csp family pseudoprotease
CspC triggers a proteolytic signaling cascade consisting of the Csp family
protease CspB and the cortex hydrolase SleC. Conserved across many of the
Clostridia, Csp proteases are subtilisin-like serine proteases that activate
pro-SleC by cleaving off its inhibitory pro-peptide. Active SleC degrades the
protective cortex layer, allowing spores to resume metabolism and growth. This
signaling pathway, however, is differentially regulated in C. difficile, since
CspC functions both as a germit receptor and regulator of CspB activity. CspB
is also produced as a fusion to a catalytically inactive CspA domain that
subsequently undergoes interdomain processing during spore formation. In this
study, we investigated the role of the CspA pseudoprotease domain in regulating
C. difficile spore germination. Mutational analyses revealed that the CspA
domain controls CspC germit receptor levels in mature spores and is required
for optimal spore germination, particularly when CspA is fused to the CspB
protease. During spore formation, the YabG protease separates these domains,
although YabG itself is dispensable for germination. Bioinformatic analyses of
Csp family members suggest that the CspC-regulated signaling pathway
characterized in C. difficile is conserved in related Peptostreptococcaceae
family members but not in the Clostridiaceae or Lachnospiraceae. Our results
indicate that pseudoproteases play critical roles in regulating C. difficile
spore germination and highlight that diverse mechanisms control spore
germination in the Clostridia. Clostridium is a large genus of obligate anaerobes belonging to the Firmicutes
phylum of bacteria, most of which have a Gram-positive cell wall structure. The
genus includes significant human and animal pathogens, causative of potentially
deadly diseases such as tetanus and botulism. Despite their relevance and many
studies suggesting that they are not a monophyletic group, the taxonomy of the
group has largely been neglected. Currently, species belonging to the genus are
placed in the unnatural order defined as Clostridiales, which includes the class
Clostridia. Here, we used genomic data from 779 strains to study the taxonomy
and evolution of the group. This analysis allowed us to 1) confirm that the
group is composed of more than one genus, 2) detect major differences between
pathogens classified as a single species within the group of authentic
Clostridium spp. (sensu stricto), 3) identify inconsistencies between taxonomy
and toxin evolution that reflect on the pervasive misclassification of strains,
and 4) identify differential traits within central metabolism of members of what
has been defined earlier and confirmed by us as cluster I. Our analysis shows
that the current taxonomic classification of Clostridium species hinders the
prediction of functions and traits, suggests a new classification for this
fascinating class of bacteria, and highlights the importance of phylogenomics
for taxonomic studies. Antimicrobial resistance continues to rise on a global scale, affecting the
environment, humans, animals and food systems. Use of natural antimicrobials has
been favoured over synthetic molecules in food preservation owing to concerns
over the adverse health effects of synthetic chemicals. The continuing need for
novel natural antimicrobial compounds has spurred research to investigate
natural sources, such as bacteria, for antimicrobials. The
antimicrobial-producing potential of bacteria has been investigated in numerous
studies. However, the discovery of antimicrobials has been biased towards
aerobes and facultative anaerobes, and strict anaerobes such as Clostridium spp.
have been largely neglected. In recent years, genomic studies have indicated the
genetic potential of strict anaerobes to produce putative bioactive molecules
and this has encouraged the exploration of Clostridium spp. for their
antimicrobial production. So far, only a limited number of antimicrobial
compounds have been isolated, identified and characterised from the genus
Clostridium. This review discusses our current knowledge and understanding of
clostridial antimicrobial compounds as well as recent genome mining studies of
Clostridium spp. focused at identification of putative gene clusters encoding
bacterial secondary metabolite groups and peptides reported to possess
antimicrobial properties. Furthermore, opportunities and challenges in the
identification of antimicrobials from Clostridium spp. using genomic-guided
approaches are discussed. The limited studies conducted so far have identified
the genus Clostridium as a viable source of antimicrobial compounds for future
investigations. |
Do nematodes contain architectural proteins like CTCF? | insulator protein CTCF has been secondarily lost in derived nematodes like C. elegans. The most highly enriched motif (LM1) corresponds to the X-box motif known from yeast and nematode. | Conserved noncoding elements (CNEs) constitute the majority of sequences under
purifying selection in the human genome, yet their function remains largely
unknown. Experimental evidence suggests that many of these elements play
regulatory roles, but little is known about regulatory motifs contained within
them. Here we describe a systematic approach to discover and characterize
regulatory motifs within mammalian CNEs by searching for long motifs (12-22 nt)
with significant enrichment in CNEs and studying their biochemical and genomic
properties. Our analysis identifies 233 long motifs (LMs), matching a total of
approximately 60,000 conserved instances across the human genome. These motifs
include 16 previously known regulatory elements, such as the histone 3'-UTR
motif and the neuron-restrictive silencer element, as well as striking examples
of novel functional elements. The most highly enriched motif (LM1) corresponds
to the X-box motif known from yeast and nematode. We show that it is bound by
the RFX1 protein and identify thousands of conserved motif instances, suggesting
a broad role for the RFX family in gene regulation. A second group of motifs
(LM2*) does not match any previously known motif. We demonstrate by biochemical
and computational methods that it defines a binding site for the CTCF protein,
which is involved in insulator function to limit the spread of gene activation.
We identify nearly 15,000 conserved sites that likely serve as insulators, and
we show that nearby genes separated by predicted CTCF sites show markedly
reduced correlation in gene expression. These sites may thus partition the human
genome into domains of expression. BACKGROUND: The zinc finger (ZF) protein CTCF (CCCTC-binding factor) is highly
conserved in Drosophila and vertebrates where it has been shown to mediate
chromatin insulation at a genomewide level. A mode of genetic regulation that
involves insulators and insulator binding proteins to establish independent
transcriptional units is currently not known in nematodes including
Caenorhabditis elegans. We therefore searched in nematodes for orthologs of
proteins that are involved in chromatin insulation.
RESULTS: While orthologs for other insulator proteins were absent in all 35
analysed nematode species, we find orthologs of CTCF in a subset of nematodes.
As an example for these we cloned the Trichinella spiralis CTCF-like gene and
revealed a genomic structure very similar to the Drosophila counterpart. To
investigate the pattern of CTCF occurrence in nematodes, we performed
phylogenetic analysis with the ZF protein sets of completely sequenced
nematodes. We show that three ZF proteins from three basal nematodes cluster
together with known CTCF proteins whereas no zinc finger protein of C. elegans
and other derived nematodes does so.
CONCLUSION: Our findings show that CTCF and possibly chromatin insulation are
present in basal nematodes. We suggest that the insulator protein CTCF has been
secondarily lost in derived nematodes like C. elegans. We propose a switch in
the regulation of gene expression during nematode evolution, from the common
vertebrate and insect type involving distantly acting regulatory elements and
chromatin insulation to a so far poorly characterised mode present in more
derived nematodes. Here, all or some of these components are missing. Instead
operons, polycistronic transcriptional units common in derived nematodes,
seemingly adopted their function. The CCCTC-binding factor (CTCF) is multi-functional, ubiquitously expressed, and
highly conserved from Drosophila to human. It has important roles in
transcriptional insulation and the formation of a high-dimensional chromatin
structure. CTCF has a paralog called "Brother of Regulator of Imprinted Sites"
(BORIS) or "CTCF-like" (CTCFL). It binds DNA at sites similar to those of CTCF.
However, the expression profiles of the two proteins are quite different. We
investigated the evolutionary trajectories of the two proteins after the
duplication event using a phylogenomic and interactomic approach. We find that
CTCF has 52 direct interaction partners while CTCFL only has 19. Almost all
interactors already existed before the emergence of CTCF and CTCFL. The unique
secondary loss of CTCF from several nematodes is paralleled by a loss of two of
its interactors, the polycomb repressive complex subunit SuZ12 and the
multifunctional transcription factor TYY1. In contrast to earlier studies
reporting the absence of BORIS from birds, we present evidence for a multigene
synteny block containing CTCFL that is conserved in mammals, reptiles, and
several species of birds, indicating that not the entire lineage of birds
experienced a loss of CTCFL. Within this synteny block, BORIS and its genomic
neighbors seem to be partitioned into two nested chromatin loops. The high
expression of SPO11, RAE1, RBM38, and PMEPA1 in male tissues suggests a possible
link between CTCFL, meiotic recombination, and fertility-associated phenotypes.
Using the 65,700 exomes and the 1000 genomes data, we observed a higher number
of intergenic, non-synonymous, and loss-of-function mutations in CTCFL than in
CTCF, suggesting a reduced strength of purifying selection, perhaps due to less
functional constraint. |
What is the primary indication of tocilizumab? | Tocilizumab is considered first-line treatment for rheumatoid arthritis. | INTRODUCTION: Approximately 30% of patients with rheumatoid arthritis (RA)
respond inadequately to conventional-synthetic disease-modifying anti-rheumatic
drugs (csDMARDs). However, widespread use of biologic DMARDs (bDMARDs) and
targeted-synthetic (tsDMARDs) is limited by cost. We formulated updated
recommendations for eligibility criteria for government-assisted funding of
bDMARDs/tsDMARDs for RA patients in Singapore.
MATERIALS AND METHODS: Published guidelines regarding use of bDMARD and tsDMARDs
were reviewed. We excluded those without a systematic literature review, formal
consensus process or evidence grading. Separately, unpublished national
reimbursement guidelines were included.
RESULTS: Eleven recommendations regarding choice of disease activity measure,
initiation, order of selection and continuation of bDMARD/tsDMARDs were
formulated. A bDMARD/tsDMARD is indicated if a patient has: (a) at least
moderately active RA with a Disease Activity Score in 28 joints/erythrocyte
sedimentation rate (DAS28-ESR) score of ≥3.2; (b) failed ≥2 csDMARD strategies,
1 of which must be a combination; (c) received an adequate dose regimen of
≥3 months for each strategy. For the first-line bDMARD/tsDMARD, either tumor
necrosis factor inhibitors (TNFi), non-TNFi (abatacept, tocilizumab, rituximab),
or tsDMARDs, may be considered. If a first-line TNFi fails, options include
another TNFi, non-TNFi biologic or tsDMARDs. If a first-line non-TNFi biologic
or tsDMARD fails, options include TNFi or another non-TNF biologic or tsDMARD.
For continued bDMARD/tsDMARD subsidization, a patient must have a documented
DAS28-ESR every 3 months and at least a moderate European League Against
Rheumatism response by 6 months.
CONCLUSION: These recommendations are useful for guiding funding decisions,
making bDMARD/tsDMARDs usage accessible and equitable in RA patients who fail
csDMARDs. |
Describe efforts on Sarcoma from the 100,000 Genomes Project | The largest whole genome sequencing (WGS) endeavour involving cancer and rare diseases was initiated in the UK in 2015 and ran for 5 years. Despite its rarity, sarcoma ranked third overall among the number of patients' samples sent for sequencing. A specialist sarcoma centre recruited close to 1000 patients to the project. WGS data was generated from 597 patients, but samples from the remaining approximately 400 patients were not sequenced. This was largely accounted for by unsuitability due to extensive necrosis, secondary to neoadjuvant radiotherapy or chemotherapy, or being placed in formalin. The number of informative genomes produced was reduced further by a PCR amplification step. Overall, diagnoses were modified for 3% of patients following review of the WGS findings. Continued refinement of the variant-calling bioinformatic pipelines is required as not all alterations were identified when validated against histology and standard of care diagnostic tests. Further research is necessary to evaluate the impact of germline mutations in patients with sarcoma, and sarcomas with evidence of hypermutation. Despite 50% of the WGS exhibiting domain 1 alterations, the number of patients with sarcoma who were eligible for clinical trials remains small, highlighting the need to revaluate clinical trial design. | The largest whole genome sequencing (WGS) endeavour involving cancer and rare
diseases was initiated in the UK in 2015 and ran for 5 years. Despite its
rarity, sarcoma ranked third overall among the number of patients' samples sent
for sequencing. Herein, we recount the lessons learned by a specialist sarcoma
centre that recruited close to 1000 patients to the project, so that we and
others may learn from our experience. WGS data was generated from 597 patients,
but samples from the remaining approximately 400 patients were not sequenced.
This was largely accounted for by unsuitability due to extensive necrosis,
secondary to neoadjuvant radiotherapy or chemotherapy, or being placed in
formalin. The number of informative genomes produced was reduced further by a
PCR amplification step. We showed that this loss of genomic data could be
mitigated by sequencing whole genomes from needle core biopsies. Storage of
resection specimens at 4 °C for up to 96 h overcame the challenge of freezing
tissue out of hours including weekends. Removing access to formalin increased
compliance to these storage arrangements. With over 70 different sarcoma
subtypes described, WGS was a useful tool for refining diagnoses and identifying
novel alterations. Genomes from 350 of the cohort of 597 patients were analysed
in this study. Overall, diagnoses were modified for 3% of patients following
review of the WGS findings. Continued refinement of the variant-calling
bioinformatic pipelines is required as not all alterations were identified when
validated against histology and standard of care diagnostic tests. Further
research is necessary to evaluate the impact of germline mutations in patients
with sarcoma, and sarcomas with evidence of hypermutation. Despite 50% of the
WGS exhibiting domain 1 alterations, the number of patients with sarcoma who
were eligible for clinical trials remains small, highlighting the need to
revaluate clinical trial design. |
What is the goal of the RadRAT calculator? | Radiation risk assessment tool (RadRAT) can be used to estimate the lifetime risk of radiation-related cancer with uncertainty intervals following a user-specified exposure history. The calculator can be used to estimate lifetime cancer risk from both uniform and non-uniform doses that are acute or chronic. It is most appropriate for low-LET radiation doses < 1 Gy, and for individuals with life-expectancy and cancer rates similar to the general population in the US. | Risk projection methods allow for timely assessment of the potential magnitude
of radiation-related cancer risks following low-dose radiation exposures. The
estimation of such risks directly through observational studies would generally
require infeasibly large studies and long-term follow-up to achieve reasonable
statistical power. We developed an online radiation risk assessment tool
(RadRAT) which can be used to estimate the lifetime risk of radiation-related
cancer with uncertainty intervals following a user-specified exposure history
(https://irep.nci.nih.gov/radrat). The uncertainty intervals constitute a key
component of the program because of the various assumptions that are involved in
such calculations. The risk models used in RadRAT are broadly based on those
developed by the BEIR VII committee for estimating lifetime risk following
low-dose radiation exposure of the US population for eleven site-specific
cancers. We developed new risk models for seven additional cancer sites, oral,
oesophagus, gallbladder, pancreas, rectum, kidney and brain/central nervous
system (CNS) cancers, using data from Japanese atomic bomb survivors. The
lifetime risk estimates are slightly higher for RadRAT than for BEIR VII across
all exposure ages mostly because the weighting of the excess relative risk and
excess absolute risk models was conducted on an arithmetic rather than a
logarithmic scale. The calculator can be used to estimate lifetime cancer risk
from both uniform and non-uniform doses that are acute or chronic. It is most
appropriate for low-LET radiation doses < 1 Gy, and for individuals with
life-expectancy and cancer rates similar to the general population in the US. BACKGROUND: To project risks of developing cancer and the number of cases
potentially induced by past, current, and future computed tomography (CT) scans
performed in the United Kingdom in individuals aged <20 years.
METHODS: Organ doses were estimated from surveys of individual scan parameters
and CT protocols used in the United Kingdom. Frequencies of scans were estimated
from the NHS Diagnostic Imaging Dataset. Excess lifetime risks (ELRs) of
radiation-related cancer were calculated as cumulative lifetime risks,
accounting for survival probabilities, using the RadRAT risk assessment tool.
RESULTS: In 2000-2008, ELRs ranged from 0.3 to 1 per 1000 head scans and 1 to 5
per 1000 non-head scans. ELRs per scan were reduced by 50-70% in 2000-2008
compared with 1990-1995, subsequent to dose reduction over time. The 130 750
scans performed in 2015 in the United Kingdom were projected to induce 64 (90%
uncertainty interval (UI): 38-113) future cancers. Current practices would lead
to about 300 (90% UI: 230-680) future cancers induced by scans performed in
2016-2020.
CONCLUSIONS: Absolute excess risks from single exposures would be low compared
with background risks, but even small increases in annual CT rates over the next
years would substantially increase the number of potential subsequent cancers. |
List the deadliest viruses in the world. | The filoviruses, Ebola virus (EBOV) and Marburg virus (MARV), are among the deadliest viruses that cause disease in humans, with reported case fatality rates of up to 90% in some outbreaks.
WHO ranks HIV as one of the deadliest diseases.
Influenza virus | The Ebola and Marburg viruses are some of the deadliest viruses in the world. In
this study a series of G-rich DNA sequences derived from these types of viruses
which possess the potential to form G-quadruplex structures are analyzed. A set
of DNA oligonucleotides derived from original viral isolates was used as a
representative modeling sequence with which to demonstrate the influence of
thiazole orange on circular dichroism (CD) spectral profiles. The results show
the unique profile of the induced CD (ICD) signal in the visible region caused
by interactions between the ligand and G-quadruplexes. This ligand was found to
stabilize the G-quadruplex structure and can also induce topological changes and
facilitate G-quadruplex multimerization. Thus, the ICD signatures can be used to
determine whether specific unknown sequences can form G-quadruplex motifs. The
viral sequences were analyzed using standard spectral and electrophoretic
methods. In addition, the ability to target G-quadruplexes located in
filoviruses offers researchers attractive therapeutic targets which would be of
particular use in the development of novel antiviral therapies. This article is
part of a Special Issue entitled "G-quadruplex" Guest Editor: Dr. Concetta
Giancola and Dr. Daniela Montesarchio. The healthcare system faces various challenges in human immunodeficiency virus
(HIV) therapy due to resistance to Anti-Retroviral Therapy (ART) as a
consequence of the evolutionary process. Despite the success of antiretroviral
drugs like Zidovudine, Zalcitabine, Raltegravir WHO ranks HIV as one of the
deadliest diseases with a mortality of one million lives in 2016. Thus, there
emerges an urgency of developing a novel anti-retroviral agent that combat
resistant HIV strains. The clinical development of ART from a single drug
regimen to current triple drug combination is very slow. The progression in the
structural biology of the viral envelope prompted the discovery of novel
targets, which can be demonstrated a proficient approach for drug design of
anti-retroviral agents. The current review enlightens the recent updates in the
structural biology of the viral envelope and focuses on CCR5 as a validated
target as well as ways to overcome CCR5 resistance. The article also throws
light on the SAR studies and most prevalent mutations in the receptor for
designing CCR5 antagonists that can combat HIV-1 infection. To conclude, the
paper lists diversified scaffolds that are in pipeline by various pharmaceutical
companies that could provide an aid for developing novel CCR5 antagonists. |
What monoclonal antibody drugs are used to treat late stage melanoma? | Nivolumab, ipilimumab, vemurafenib, and dabrafenib are used to treat late stage melanoma | PURPOSE: L612, a human IgM monoclonal antibody produced by an EBV-transformed
human B-cell line, binds to ganglioside GM3 and kills GM3-positive human
melanoma cells in the presence of complement. It has been shown to be effective
in some patients with late-stage melanoma. L612 consists of hexameric IgM (about
20%), pentameric IgM (about 74%), and other minor IgM molecules. Because
hexameric IgM activates complement more effectively than pentameric IgM, we
developed and evaluated a hexamer-domit recombit IgM for clinical
applications.
EXPERIMENTAL DESIGN: Chinese hamster ovary (CHO) cells were transfected with
heavy- and light-chain genes of L612, with or without the joining-chain gene.
Antitumor effects of the recombit IgM secreted from CHO cells were evaluated
in vitro and in vivo.
RESULTS: Recombit IgM secreted from CHO cells without the joining chain
(designated CA19) was approximately 80% hexameric, whereas recombit IgM from
CHO cells transfected with heavy-, light-, and joining-chain genes (designated
CJ45) was about 90% pentameric. Both CA19 and CJ45 recombit IgMs caused
complement-dependent cytotoxicity against human and mouse melanoma cell lines,
but the amount of CA19 required for 50% specific cytotoxicity was 5 to 10 times
smaller. I.v. injection of CA19 compared with CJ45 or native L612 elicited more
profound antitumor activity in nude rats bearing a GM3-positive mouse melanoma
xenograft.
CONCLUSIONS: A hexamer-domit human IgM against GM3 may provide a more potent
treatment option for patients with GM3-positive melanoma. Use of monoclonal anti-CTLA4 antibodies represents a new promising strategy to
block the activation of immunosuppressive CTLA-4 and thus induce tumour
regression. This review is mainly focused on the report of existing data on the
clinical use of Ipilimumab (formerly MDX-010) in the treatment of metastatic
melanoma. Several phase I and II trials have been conducted to evaluate safety
and efficacy of this form of immunotherapy either alone or in combination with
vaccines or chemotherapy in patients with stage III or stage IV melanoma.
Results from these studies are presented, patented and discussed. The mechanism
of ipilimumab action may take time to induce an anti-tumour immune response and
thus it is recommended that ipilimumab therapy should be carried out for at
least 12 weeks, even in the presence of early progressive disease. Objective
response of around 15% has been reported in patients treated with ipilimumab.
However, ipilimumab-mediated objective response and stable disease tend to be
durable. The therapy with ipilimumab is associated with different side effects
classified as immune-related adverse events (IRAEs). The most common IRAEs are
enterocolitis and dermatitis. Majority of IRAEs disappear with the
discontinuation of ipilimumab anti-CTLA-4 therapy. Several phase II/III trials
are ongoing to evaluate ipilimumab alone or in combination with other
therapeutic modalities. Results from these trials are awaited. PURPOSE: The primary objective of this phase I/II study was to determine the
safety and pharmacokinetic profile of either transfectoma- or a
hybridoma-derived ipilimumab. Secondary objectives included determination of a
maximum-tolerated dose and assessment of clinical activity.
PATIENTS AND METHODS: Eighty-eight patients with unresectable stage III or IV
melanoma with at least one measurable lesion were treated. Mean age was 59
years, with 65% male and 35% female patients, and 79% of patients had received
prior systemic therapy. Single doses of ipilimumab up to 20 mg/kg (group A,
single dose), multiple doses up to 5 mg/kg (group A, multiple dose), and
multiple doses up to 10 mg/kg (group B) were administered.
RESULTS: Single dosing up to 20 mg/kg of transfectoma antibody was well
tolerated, as were multiple doses up to 10 mg/kg without a maximum-tolerated
dose. In group B, dose-limiting toxicity was seen in six of 23 melanoma
patients. Grade 3 or 4 immune-related adverse events (irAEs) were observed in
14% of patients (12 of 88 patients), and grade 1 or 2 irAEs were seen in an
additional 58%. The half-life of ipilimumab was 359 hours. In group B, there was
one partial response (23+ months), one complete response (21+ months), and seven
patients with stable disease (SD), for a disease control rate of 39%. Two
patients in group B with SD had slow, steady decline in tumor burden that was
ongoing at 1 year of observation.
CONCLUSION: Ipilimumab has activity in patients with metastatic melanoma. Late
responses were observed in patients with prolonged SD. Melanoma is a maligcy that is highly curable in the early stages but has
devastating consequences in later stages due to lack of response to traditional
treatments. Improved understanding of the basic science of tumorigenesis has
helped lead to novel targeted therapies which are producing beneficial results
in patients with melanoma. Enhancement of the immune system by blockade of the
cytotoxic T-lymphocyte associated antigen-4 by the monoclonal antibody
ipilimumab is now approved by the United States Food and Drug Administration
(FDA) for use in patients with unresectable melanoma. The approval of this drug
was based on the first ever data in melanoma showing an improvement in overall
survival. New advances in targeting components of the mitogen-activated protein
kinase pathway are showing impressive responses in clinical trials in most
patients harboring activating mutations in BRAF. Thus, this is a new era in the
management of melanoma and we review the recent progress made in treating
patients with advanced disease. Melanoma is the deadliest form of skin cancer. Ipilimumab, a novel
immunotherapy, is the first treatment shown to improve survival in patients with
metastatic melanoma in large randomized controlled studies. The most concerning
side effects reported in clinical studies of ipilimumab fall into the category
of immune-related adverse events, which include enterocolitis, dermatitis,
thyroiditis, hepatitis, hypophysitis, uveitis, and others. During the course of
routine clinical care at Mount Sinai Medical Center, frequent hepatotoxicity was
noted when ipilimumab was administered at a dose of 3 mg/kg according to Food
and Drug Administration (FDA) guidelines. To better characterize these adverse
events, we conducted a retrospective review of the first 11 patients with
metastatic melanoma treated with ipilimumab at the Mount Sinai Medical Center
after FDA approval. Aspartate aminotransferase (AST) and alanine
aminotransferase (ALT) elevation, as defined by the National Cancer Institute's
Common Terminology Criteria for Adverse Events, each occurred in six of 11 cases
(≥grade 1), a notably higher frequency than could be expected on the basis of
the FDA licensing study where elevations were reported in 0.8 and 1.5% of
patients for AST and ALT, respectively. Grade 3 elevations in AST occurred in
three of 11 patients as compared with 0% in the licensing trial. All cases of
transaminitis resolved when ipilimumab was temporarily withheld without
administration of immunosuppressive medication. During routine clinical care of
late-stage melanoma patients with ipilimumab, physicians should monitor patients
closely for hepatotoxicity and be aware that toxicity rates may differ across
populations during ipilimumab therapy. IMPORTANCE: This case series highlights the risk of uveitis in patients treated
with vemurafenib for unresectable or metastatic cutaneous melanoma.
OBJECTIVE: To assess the occurrence and severity of uveitis as an adverse effect
of vemurafenib therapy.
DESIGN, SETTING, AND PATIENTS: In this observational small case series, data
were collected successively from May 1, 2012, through February 31, 2013, from
patients with clinical signs of ocular inflammation treated with vemurafenib at
the Department of Ophthalmology, Cochin-Hôtel-Dieu Hospital.
MAIN OUTCOMES AND MEASURES: Patients' demographics, vemurafenib dosages, and the
intervals between the onset of treatment and the first ocular symptoms were
recorded. The characteristics of ocular inflammatory manifestations were
analyzed. The effect of the discontinuation of vemurafenib therapy on ocular
manifestations was assessed, as well as the effect of rechallenging when
vemurafenib was reintroduced.
RESULTS: Seven patients (mean [SD] age, 74.7 [4.0] years) had uveitis. The
vemurafenib dose was 960 mg twice per day in 6 patients and a half dose in 1
patient. The mean (SD) time until the appearance of ocular signs was 5.6 (2.3)
months (range, 19 days to 7 months), and inflammation ranged from mild or
low-grade anterior uveitis to severe explosive panuveitis complicated by retinal
detachment. Signs of ocular inflammation were always bilateral. Optical
coherence tomography revealed a macular edema in only 1 of the 7 patients.
Clinical improvement occurred when vemurafenib therapy was stopped in 5 of 7
patients. The rechallenge at treatment reintroduction was positive in 2 of 7
patients.
CONCLUSIONS AND RELEVANCE: This small case series highlights that uveitis can be
a noteworthy adverse effect of vemurafenib therapy in patients with metastatic
cutaneous melanoma. However, these cases of uveitis were usually restricted to
the anterior segment and manageable with local corticosteroid therapy, which
justified the continuation of vemurafenib therapy because the benefits regarding
the patients' survival were greater than the risk to their vision. Until recently, a treatment for advanced melanoma with a tolerable side effect
profile has remained elusive. Therefore, despite its relatively rare occurrence,
melanoma continues to cause the majority of skin cancer deaths, with less than
15% of those affected with late-stage disease surviving 10 years or more.
Historically, the mainstay of treatment has been single-agent chemotherapy or
immunotherapy with the alkylating agent dacarbazine and interleukin-2 or
interferon. Cytotoxic chemotherapy with dacarbazine demonstrated poor response
rates and little or no survival benefit, whereas IL-2 and interferon, although
showing durable responses, are associated with poor side effect profiles (1, 2).
However, with the elucidation of the molecular biology and oncogenic pathways
involved in melanoma, agents targeted against mutations in the mitogen-activated
protein kinase (MAPK) pathway including BRAF and MEK inhibitors have
demonstrated prolonged survival and more manageable side effect profiles
relative to traditional chemotherapy. Simultaneously, an evolved understanding
of the immunologic basis for the development and regression of melanoma lead to
the discovery of immune checkpoint inhibitors, which confer a similar survival
benefit. Foremost among these was ipilimumab, a monoclonal antibody against the
negative regulatory checkpoint molecule cytotoxic T-lymphocyte protein 4
(CTLA-4), which was the first drug in the management of metastatic melanoma to
confer a survival benefit. However, treatment is complicated by a high rate of
grade 3 and 4 immune-related adverse events and limited response. Nivolumab, a
fully human monoclonal antibody against programmed cell death protein 1 (PD-1),
has shown a survival benefit in an open-label phase II trial, and was the first
PD-1 inhibitor to be approved worldwide. With a favorable side effect profile
and ongoing trials in combination with extant therapies, nivolumab shows
substantial potential to further augment the options for an effective treatment
in maligt melanoma. The anti-programmed cell death-1 (PD-1) monoclonal antibodies pembrolizumab and
nivolumab have been contingently approved for the treatment of patients with
advanced melanoma based on their durable response, high response rate, and
favorable safety profile. Mature survival data from randomized phase III trials
are eagerly awaited to confirm their position as the standard-of-care frontline
or second-line therapy in advanced melanoma management algorithm. The
immune-related adverse events associated with these novel agents are somewhat
different than those of ipilimumab, considering the manifestation of pneumonitis
and acute renal failure. Active research is ongoing to identify biomarkers
predictive of clinical benefit to the anti-PD1 monoclonal antibodies, to expand
their utility in other disease settings, and to explore their safety and
efficacy in combination with other therapeutic agents. Uswered questions
concerning optimal dosing schedule, treatment duration, and therapy sequencing
will also need to be addressed in future investigations. The 37kDa/67kDa laminin receptor (LRP/LR) is a non-integrin laminin receptor
which is overexpressed in tumorigenic cells and supports progression of cancer
via promoting metastasis, angiogenesis and telomerase activity and impediment of
apoptosis. The present study investigates the role of LRP/LR on the metastatic
potential of early (A375) and late (A375SM) stage maligt melanoma cells. Flow
cytometry revealed that both early and late stage maligt melanoma cells
display high levels of LRP/LR on their cell surface. Flow cytometry and western
blot analysis showed that late stage maligt melanoma cells display
significantly higher total and cell surface LRP/LR levels in comparison to early
stage maligt melanoma cells and the poorly invasive breast cancer (MCF-7)
control cell line. Targeting LRP/LR using the LRP/LR specific antibody IgG1-iS18
resulted in a significant reduction of the adhesive potential to laminin-1 and
the invasive potential through the 'ECM-simulating' Matrigel™ of both early and
late stage maligt melanoma cells. Furthermore, Pearson's correlation
coefficient confirmed that increased LRP levels correlate with the increased
invasive and adhesive potential in early and late stage melanoma cells. Thus,
blocking LRP/LR using the IgG1-iS18 antibody may therefore be a promising
therapeutic strategy for early and late stage maligt melanoma treatment. BACKGROUND: Checkpoint blockade with ipilimumab provides long-term survival to a
significant proportion of patients with metastatic melanoma. New approaches to
increase survival and to predict which patients will benefit from treatment are
needed. This phase II trial combined ipilimumab with carboplatin/paclitaxel (CP)
to assess its safety, efficacy, and to search for peripheral and tumor-based
predictive biomarkers.
METHODS: Thirty patients with untreated unresectable/metastatic melanoma were
treated with ipilimumab and CP. Adverse events (AEs) were monitored and response
to treatment was evaluated. Tumor tissue and peripheral blood were collected at
specified time points to characterize tumor immune markers by
immunohistochemistry and systemic immune activity by multiplex assays and flow
cytometry.
RESULTS: Eighty three percent of patients received all 5 cycles of CP and 93%
completed ipilimumab induction. Serious AEs occurred in 13% of patients, and no
treatment-related deaths were observed. Best Overall Response Rate (BORR) and
Disease Control Rate (DCR) were 27 and 57%, respectively. Median overall
survival was 16.2 months. Response to treatment was positively correlated with a
higher tumor CD3+ infiltrate (immune score) at baseline. NRAS and BRAF mutations
were less frequent in patients who experienced clinical benefit. Assessment of
peripheral blood revealed that non-responders had elevated baseline levels of
CXCL8 and CCL4, and a higher proportion of circulating late differentiated B
cells. Pre-existing high levels of chemokines (CCL3, CCL4 and CXCL8) and
advanced B cell differentiation were strongly associated with worse patient
overall survival. Elevated proportions of circulating CD8+/PD-1+ T cells during
treatment were associated with worse survival.
CONCLUSIONS: The combination of ipilimumab and CP was well tolerated and
revealed novel characteristics associated with patients likely to benefit from
treatment. A pre-existing systemic inflammatory state characterized by elevation
of selected chemokines and advanced B cell differentiation, was strongly
associated with poor patient outcomes, revealing potential predictive
circulating biomarkers.
TRIAL REGISTRATION: Clinicaltrials.gov , NCT01676649 , registered on August 29,
2012. |
Which transcription factor controls Drosophila's Hes genes? | The Notch/Hes axis represses a cohort of transcription factor genes . In Drosophila, activation of the Notch receptor induces transcriptional repressors encoded by the hairy/Enhancer of split (HES) genes, which shut off achaete-scute transcription . The molecular details of how Hes and Hey proteins control transcription are still poorly understood . | Hes2 encodes a mammalian basic helix-loop-helix transcriptional repressor
homologous to the products of Drosophila hairy and Enhancer of split. Here, we
isolated and characterized the mouse Hes2 gene. This gene consists of four
exons, and all the introns are located within the protein-coding region at
positions homologous to those of other Hes genes. On the inter-specific
backcross analyses, mouse Hes2 is mapped to the distal region of Chromosome 4
near the Hes3 and Hes5 loci, which are different from the Hes1 locus on
Chromosome 16. Upstream of the transcription initiation site, there are GC-rich
regions, but a typical TATA box is not present. Transient transfection analyses
demonstrated that, while Hes1 and Hes5 promoter activities are significantly
upregulated by the active form of Notch, a key regulator of cellular
differentiation, Hes2 and Hes3 promoter activities are not. These results
suggest that Hes genes are functionally classified into two groups: those that
are regulated by Notch and those that are not. HES transcriptional repressors are important components of the Notch pathway
that regulates neurogenesis from Drosophila to vertebrates. These proteins are
normally induced by Notch activity and inhibit neural commitment by antagonizing
the activity of proneural genes. We describe here four chick hes genes that are
expressed during neurogenesis: three hes5-like genes (hes5-1, hes5-2 and hes5-3)
and one hes6-like (hes6-2). We show that hes6-2 represses transcription of the
hes5 genes, thus functioning as a negative regulator of Notch signaling.
Conversely, hes6-2 may be repressed by hes5 activity. In cells committing to
differentiation, we find that hes6-2 is up-regulated by proneural genes and
contributes to the proneural program of neuronal commitment by preventing Notch
activity in these cells. In neural progenitors, Notch signaling produces an
initial burst of hes5 activity, which represses hes6-2. However, as hes5
transcription declines due to negative auto-regulation, hes6-2 may become active
and inhibit the remaining hes5 activity to end Notch signaling. These cells can
then enter a new cycle of fate decisions and will be kept as progenitors if a
new pulse of Notch activity occurs. Maintece of progenitors during vertebrate
neurogenesis therefore requires that these cells go through successive cycles of
Notch activity. We propose that the hes5/hes6 circuitry of negative
cross-regulations is a conserved feature of the Notch pathway that underlies
these cycles in neural progenitors. Hes and Hey genes are the mammalian counterparts of the Hairy and
Enhancer-of-split type of genes in Drosophila and they represent the primary
targets of the Delta-Notch signaling pathway. Hairy-related factors control
multiple steps of embryonic development and misregulation is associated with
various defects. Hes and Hey genes (also called Hesr, Chf, Hrt, Herp or
gridlock) encode transcriptional regulators of the basic helix-loop-helix class
that mainly act as repressors. The molecular details of how Hes and Hey proteins
control transcription are still poorly understood, however. Proposed modes of
action include direct binding to N- or E-box DNA sequences of target promoters
as well as indirect binding through other sequence-specific transcription
factors or sequestration of transcriptional activators. Repression may rely on
recruitment of corepressors and induction of histone modifications, or even
interference with the general transcriptional machinery. All of these models
require extensive protein-protein interactions. Here we review data published on
protein-protein and protein-DNA interactions of Hairy-related factors and
discuss their implications for transcriptional regulation. In addition, we
summarize recent progress on the identification of potential target genes and
the analysis of mouse models. BACKGROUND: Establishment and maintece of a functional central nervous system
(CNS) requires a highly orchestrated process of neural progenitor cell
proliferation, cell cycle exit, and differentiation. An evolutionary conserved
program consisting of Notch signalling mediated by basic Helix-Loop-Helix (bHLH)
transcription factor activity is necessary for both the maintece of neural
progenitor cell character and the progression of neurogenesis; however,
additional players in mammalian CNS neural specification remain largely unknown.
In Drosophila we recently characterized Hamlet, a transcription factor that
mediates Notch signalling and neural cell fate.
METHODOLOGY/PRINCIPAL FINDINGS: Hamlet is a member of the Prdm (PRDI-BF1 and RIZ
homology domain containing) proto-oncogene transcription factor family, and in
this study we report that multiple genes in the Prdm family (Prdm6, 8, 12, 13
and 16) are expressed in the developing mouse CNS in a spatially and temporally
restricted manner. In developing spinal cord Prdm8, 12 and 13 are expressed in
precise neuronal progenitor zones suggesting that they may specify discrete
neuronal subtypes. In developing telencephalon Prdm12 and 16 are expressed in
the ventricular zone in a lateral to medial graded manner, and Prdm8 is
expressed in a complementary domain in postmitotic neurons. In postnatal brain
Prdm8 additionally shows restricted expression in cortical layers 2/3 and 4, the
hippocampus, and the amygdala. To further elucidate roles of Prdm8 and 16 in the
developing telencephalon we analyzed the relationship between these factors and
the bHLH Hes (Hairy and enhancer of split homolog) effectors of Notch
signalling. In Hes null telencephalon neural differentiation is enhanced, Prdm8
expression is upregulated, and Prdm16 expression is downregulated; conversely in
utero electroporation of Hes1 into the developing telencephalon upregulates
Prdm16 expression.
CONCLUSIONS/SIGNIFICANCE: Our data demonstrate that Prdm genes are regulated by
the Notch-Hes pathway and represent strong candidates to control neural class
specification and the sequential progression of mammalian CNS neurogenesis. The Notch pathway plays an important role in ovary development in invertebrates
like Drosophila. However its role for the mammalian ovary is unclear. Mammalian
Hes genes encode transcriptional factors that mediate many of the activities of
the Notch pathway. Here, we have studied the function of Hes1 during embryonic
development of the mouse ovary. We find that Hes1 protein is present in somatic
cells and oocyte cytoplasm and decreases between E15.5 and P0. Conventional Hes1
knock-out (KO), Hes1 conditional KO in the ovarian somatic, and chemical
inhibition of Notch signaling decrease the total number, size and maturation of
oocytes and increase the number of pregranulosa cells at P0. These defects
correlate with abnormal proliferation and enhanced apoptosis. Expression of the
proapoptotic gene Inhbb is increased, while the levels of the antiapoptotic and
oocyte maturation marker Kit are decreased in the Hes1 KO ovaries. Conversely,
overactivation of the Notch pathway in ovarian somatic cells increases the
number of mature oocytes and decreases the number of pregranulosa cells.
Fertility is also reduced by either Hes1 deletion or Notch pathway
overactivation. In conclusion, our data suggest that the Notch-Hes1 pathway
regulates ovarian somatic cell development, which is necessary for oocyte
survival and maturation. Hes genes, encoding basic helix-loop-helix (HLH) transcriptional repressors, are
mammalian homologues of Drosophila hairy and Enhancer of split genes, both of
which are required for normal neurogenesis in Drosophila. There are seven
members in the human Hes family, Hes1-7, which are expressed in many tissues and
play various roles mainly in development. All Hes proteins have three conserved
domains: basic HLH (bHLH), Orange, and WRPW domains. The basic region binds to
target DNA sequences, while the HLH region forms homo- and heterodimers with
other bHLH proteins, the Orange domain is responsible for the selection of
partners during heterodimer formation, and the WRPW domain recruits
corepressors. Hes1, Hes5, and Hes7 are known as downstream effectors of
canonical Notch signaling, which regulates cell differentiation via cell-cell
interaction. Hes factors regulate many events in development by repressing the
expression of target genes, many of which encode transcriptional activators that
promote cell differentiation. For example, Hes1, Hes3, and Hes5 are highly
expressed by neural stem cells, and inactivation of these genes results in
insufficient maintece of stem cell proliferation and prematurely promotes
neuronal differentiation. Recently, it was shown that the expression dynamics of
Hes1 plays crucial roles in proper developmental timings and fate-determination
steps of embryonic stem cells and neural progenitor cells. Here, we discuss some
key features of Hes factors in development and diseases. Neural stem cells divide during embryogenesis and juvenile life to generate the
entire complement of neurons and glia in the nervous system of vertebrates and
invertebrates. Studies of the mechanisms controlling the fine balance between
neural stem cells and more differentiated progenitors have shown that, in every
asymmetric cell division, progenitors send a Delta-Notch signal to their sibling
stem cells. Here, we show that excessive activation of Notch or overexpression
of its direct targets of the Hes family causes stem-cell hyperplasias in the
Drosophila larval central nervous system, which can progress to maligt
tumours after allografting to adult hosts. We combined transcriptomic data from
these hyperplasias with chromatin occupancy data for Dpn, a Hes transcription
factor, to identify genes regulated by Hes factors in this process. We show that
the Notch/Hes axis represses a cohort of transcription factor genes. These are
excluded from the stem cells and promote early differentiation steps, most
likely by preventing the reversion of immature progenitors to a stem-cell fate.
We describe the impact of two of these 'anti-stemness' factors, Zfh1 and Gcm, on
Notch/Hes-triggered tumorigenesis. |
Is tocilizumab a tumor necrosis factor inhibitor? | No, tocilizumab, is a non-TNFi DMARD. | INTRODUCTION: Approximately 30% of patients with rheumatoid arthritis (RA)
respond inadequately to conventional-synthetic disease-modifying anti-rheumatic
drugs (csDMARDs). However, widespread use of biologic DMARDs (bDMARDs) and
targeted-synthetic (tsDMARDs) is limited by cost. We formulated updated
recommendations for eligibility criteria for government-assisted funding of
bDMARDs/tsDMARDs for RA patients in Singapore.
MATERIALS AND METHODS: Published guidelines regarding use of bDMARD and tsDMARDs
were reviewed. We excluded those without a systematic literature review, formal
consensus process or evidence grading. Separately, unpublished national
reimbursement guidelines were included.
RESULTS: Eleven recommendations regarding choice of disease activity measure,
initiation, order of selection and continuation of bDMARD/tsDMARDs were
formulated. A bDMARD/tsDMARD is indicated if a patient has: (a) at least
moderately active RA with a Disease Activity Score in 28 joints/erythrocyte
sedimentation rate (DAS28-ESR) score of ≥3.2; (b) failed ≥2 csDMARD strategies,
1 of which must be a combination; (c) received an adequate dose regimen of
≥3 months for each strategy. For the first-line bDMARD/tsDMARD, either tumor
necrosis factor inhibitors (TNFi), non-TNFi (abatacept, tocilizumab, rituximab),
or tsDMARDs, may be considered. If a first-line TNFi fails, options include
another TNFi, non-TNFi biologic or tsDMARDs. If a first-line non-TNFi biologic
or tsDMARD fails, options include TNFi or another non-TNF biologic or tsDMARD.
For continued bDMARD/tsDMARD subsidization, a patient must have a documented
DAS28-ESR every 3 months and at least a moderate European League Against
Rheumatism response by 6 months.
CONCLUSION: These recommendations are useful for guiding funding decisions,
making bDMARD/tsDMARDs usage accessible and equitable in RA patients who fail
csDMARDs. |
Describe manifestations of KIF1A-related disorders in children | KIF1A-related disorders (KRD) were first described in 2011 and the phenotypic spectrum has subsequently expanded to encompass a range of central and peripheral nervous system involvement. A study identified twelve individuals from 10 families. Eight different mutations were present, including four novel mutations. Two patients displayed a very severe phenotype including congenital contractures, severe spasticity and/or dystonia, dysautonomia, severe sensorimotor polyneuropathy and optic atrophy, significant white matter changes on brain MRI, respiratory insufficiency, and complete lack of neurodevelopmental progress. The remaining 10 patients represented a spectrum of severity with common features including a movement disorder with spasticity and/or dystonia, subtle features of dysautonomia, sensory axonal neuropathy, varying degrees of optic atrophy and of learning and/or behavioural difficulties, and subtle or absent-but sometimes progressive-changes in white matter on MRI. Epilepsy was common among the more severely affected children. | |
Describe the mechanism of action of pitolisant. | Pitolisant is an antagonist/inverse agonist of the human histamine H3 receptor. It is used for treatment of narcolepsy. | OBJECTIVE: Narcolepsy is a rare disabling sleep disorder characterized by
excessive daytime sleepiness and cataplexy (sudden loss of muscle tone). Drugs
such as pitolisant, which block histamine H3 autoreceptors, constitute a newly
identified class of stimulants because they increase brain histamine and enhance
wakefulness in animal and human adult narcolepsy.
METHODS: We report our experience with the off-label use of pitolisant in 4
teenagers with narcolepsy/cataplexy with severe daytime sleepiness, refractory
to available treatments (modafinil, methylphenidate, mazindol, sodium oxybate,
and D-amphetamine).
RESULTS: All teenagers developed their disease during childhood (11.3 ± 2.4
years; 50% boys) and were 17.3 ± 0.8 years old at the time of pitolisant
therapy. Pitolisant treatment was increased from 10 to 30 mg (n = 1) and 40 mg
(n = 3). The adapted Epworth Sleepiness Score decreased from 14.3 ± 1.1 to 9.5 ±
2.9 (P = 0.03) with pitolisant alone to 7 ± 3.4 when combined with mazindol (n =
1), methylphenidate (n = 1), or sodium oxybate plus modafinil (n = 1). Mean
sleep onset latency increased from 31 ± 14 minutes to 36 ± 8 minutes (P = 0.21)
on the maintece of wakefulness test. The severity and frequency of cataplexy
were slightly improved. Adverse effects were minor (insomnia, headache, hot
flushes, leg pain, and hallucinations) and transitory, except for insomnia,
which persisted in 2 teenagers. The benefit was maintained after a mean of 13
months.
CONCLUSIONS: Pitolisant could constitute an acceptable alternative for the
treatment of refractory sleepiness in teenagers with narcolepsy. OBJECTIVE AND DESIGN: Pitolisant (BF2.649) is a selective inverse agonist for
the histamine H(3) receptor and was developed for the treatment of excessive
daytime sleepiness in Parkinson disease, narcolepsy, and schizophrenia. Since
H(3)-ligands can decrease inflammatory pain, we tested Pitolisant in
inflammatory and neuropathic pain models. MATERIALS AND TREATMENTS: Behavioral
effects of pitolisant and the structural different H(3) receptor inverse
agonists ciproxifan and ST-889 were tested in zymosan-induced inflammation and
the spared nerve injury model for neuropathic pain.
METHODS: Responses to mechanical and thermal stimuli were determined. Calcium
imaging was performed with primary neuronal cultures of dorsal root ganglions.
RESULTS: Clinically relevant doses of pitolisant (10 mg/kg) had no relevant
effect on mechanical or thermal pain thresholds in all animal models. Higher
doses (50 mg/kg) dramatically increased thermal but not mechanical pain
thresholds. Neither ciproxifan nor ST-889 altered thermal pain thresholds. In
peripheral sensory neurons high concentrations of pitolisant (30-500 μM), but
not ciproxifan, partially inhibited calcium increases induced by capsaicin, a
selective activator of transient receptor potential vanilloid receptor 1
(TRPV1). High doses of pitolisant induced a strong hypothermia.
CONCLUSION: The data show a dramatic effect of high dosages of pitolisant on the
thermosensory system, which appears to be H(3) receptor-independent. Pitolisant (Wakix™) is an inverse agonist of the histamine H3 receptor that is
being developed by Bioproject. Oral pitolisant is approved in the EU for the
treatment of narcolepsy with or without cataplexy in adults. Pitolisant has
received a Temporary Authorization of Use in France for this indication in case
of treatment failure, intolerance or contraindication to currently available
treatment. Pitolisant has orphan drug designation in the EU and the USA. In the
pivotal HARMONY I trial, pitolisant significantly decreased excessive daytime
sleepiness versus placebo in adults with narcolepsy with or without cataplexy
(primary endpoint). Pitolisant also significantly decreased cataplexy rate
versus placebo in these patients. This article summarizes the milestones in the
development of pitolisant leading to this first approval for narcolepsy. Previous studies have demonstrated that the histamine H3 receptor inverse
agonist thioperamide potentiates the stimulant and rewarding effects of cocaine.
However, these potentiating effects of thioperamide do not necessarily result
from H3 receptor blockade since thioperamide is an imidazole-based compound
capable of enhancing plasma cocaine concentrations by blocking cytochrome P450
activity. In contrast, Pitolisant is a non-imidazole H3 receptor inverse agonist
that has already been tested in clinical trials but it remains to be determined
whether this compound also potentiates the behavioral effects of cocaine. The
present study tested the effects of Pitolisant on locomotion, on cocaine-induced
hyperactivity and on the development of conditioned place preference induced by
cocaine (2 and 8mg/kg, i.p.) in male C57BL/6J mice. Pitolisant was injected
30min before each cocaine-pairing session. Locomotion recorded on the first
cocaine-pairing session was used to test the effects of Pitolisant on the
locomotor effects of cocaine. Our results show that doses of Pitolisant higher
than 10mg/kg depressed locomotion. When injected alone at doses that did not
affect locomotion, Pitolisant (2.5-10mg/kg, i.p.) had no rewarding properties in
the place conditioning technique. Additionally, Pitolisant did not significantly
alter cocaine-induced hyperactivity and cocaine-induced conditioned place
preference. Taken together, our study indicates that Pitolisant has no addictive
properties alone. Moreover, this compound does not significantly affect the
stimulant and rewarding effects of cocaine. These results add further evidence
to support the hypothesis that a pharmacokinetic interaction is involved in the
ability of thioperamide to potentiate cocaine's psychomotor effects. The Gi/o protein-coupled histamine H3 receptor is distributed throughout the
central nervous system including areas like cerebral cortex, hippocampus and
striatum with the density being highest in the posterior hypothalamus, i.e. the
area in which the histaminergic cell bodies are located. In contrast to the
other histamine receptor subtypes (H1, H2 and H4), the H3 receptor is located
presynaptically and shows a constitutive activity. In detail, H3 receptors are
involved in the inhibition of histamine release (presynaptic autoreceptor),
impulse flow along the histaminergic neurones (somadendritic autoreceptor) and
histamine synthesis. Moreover, they occur as inhibitory presynaptic
heteroreceptors on serotoninergic, noradrenergic, dopaminergic, glutamatergic,
GABAergic and perhaps cholinergic neurones. This review shows for four functions
of the brain that the H3 receptor represents a brake against the wake-promoting,
anticonvulsant and anorectic effect of histamine (via postsynaptic H1 receptors)
and its procognitive activity (via postsynaptic H1 and H2 receptors). Indeed, H1
agonists and H3 inverse agonists elicit essentially the same effects, at least
in rodents; these effects are opposite in direction to those elicited by
brain-penetrating H1 receptor antagonists in humans. Although the benefit for H3
inverse agonists for the symptomatic treatment of dementias is inconclusive,
several members of this group have shown a marked potential for the treatment of
disorders associated with excessive daytime sleepiness. In March 2016, the
European Commission granted a marketing authorisation for pitolisant (WakixR)
(as the first representative of the H3 inverse agonists) for the treatment of
narcolepsy. BACKGROUND: Histaminergic neurons are crucial to maintain wakefulness, but their
role in cataplexy is unknown. We assessed the safety and efficacy of pitolisant,
a histamine H3 receptor inverse agonist, for treatment of cataplexy in patients
with narcolepsy.
METHODS: For this randomised, double-blind, placebo-controlled trial we
recruited patients with narcolepsy from 16 sleep centres in nine countries
(Bulgaria, Czech Republic, Hungary, Macedonia, Poland, Russia, Serbia, Turkey,
and Ukraine). Patients were eligible if they were aged 18 years or older,
diagnosed with narcolepsy with cataplexy according to version two of the
International Classification of Sleep Disorders criteria, experienced at least
three cataplexies per week, and had excessive daytime sleepiness (defined as an
Epworth Sleepiness Scale score ≥12). We used a computer-generated sequence via
an interactive web response system to randomly assign patients to receive either
pitolisant or placebo once per day (1:1 ratio). Randomisation was done in blocks
of four. Participants and investigators were masked to treatment allocation.
Treatment lasted for 7 weeks: 3 weeks of flexible dosing decided by
investigators according to efficacy and tolerance (5 mg, 10 mg, or 20 mg oral
pitolisant), followed by 4 weeks of stable dosing (5 mg, 10 mg, 20 mg, or 40
mg). The primary endpoint was the change in the average number of cataplexy
attacks per week as recorded in patient diaries (weekly cataplexy rate [WCR])
between the 2 weeks of baseline and the 4 weeks of stable dosing period.
Analysis was by intention to treat. This trial is registered with
ClinicalTrials.gov, number NCT01800045.
FINDINGS: The trial was done between April 19, 2013, and Jan 28, 2015. We
screened 117 patients, 106 of whom were randomly assigned to treatment (54 to
pitolisant and 52 to placebo) and, after dropout, 54 patients from the
pitolisant group and 51 from the placebo group were included in the
intention-to-treat analysis. The WCR during the stable dosing period compared
with baseline was decreased by 75% (WCRfinal=2·27; WCRbaseline=9·15;
WCRfinal/baseline=0·25) in patients who received pitolisant and 38%
(WCRfinal=4·52; WCRbaseline=7·31; WCRfinal/baseline=0·62) in patients who
received placebo (rate ratio 0·512; 95% CI 0·43-0·60, p<0·0001).
Treatment-related adverse events were significantly more common in the
pitolisant group than in the placebo group (15 [28%] of 54 vs 6 [12%] of 51;
p=0·048). There were no serious adverse events, but one case of severe nausea in
the pitolisant group. The most frequent adverse events in the pitolisant group
(headache, irritability, anxiety, and nausea) were mild or moderate except one
case of severe nausea. No withdrawal syndrome was detected following pitolisant
treatment; one case was detected in the placebo group.
INTERPRETATION: Pitolisant was well tolerated and efficacious in reducing
cataplexy. If confirmed in long-term studies, pitolisant might constitute a
useful first-line therapy for cataplexy in patients with narcolepsy, for whom
there are currently few therapeutic options.
FUNDING: Bioprojet, France. On 31 March 2016, the European Commission issued a decision for a marketing
authorisation valid throughout the European Union (EU) for pitolisant (Wakix)
for the treatment of narcolepsy with or without cataplexy in adults. Pitolisant
is an antagonist/inverse agonist of the human histamine H3 receptor. The dose
should be selected using an up-titration scheme depending on individual patient
response and tolerance and should not exceed 36 mg/day. The main evidence of
efficacy of pitolisant was based on two Phase III clinical trials. The
improvement on excessive daytime sleepiness was shown against placebo in the
Harmony I study (-3.33 points; 95% confidence interval (CI) [-5.83; -0.83];
p = 0.024) and in Harmony CTP (-3.41 points; 95% CI [-4.95; -1.87]; p < 0.0001).
The daily cataplexy rate in Harmony I improved against placebo with a rate ratio
(rR) of 0.38 whilst in the Harmony CTP the ratio of improvement on weekly
cataplexy rate against placebo was 0.512. The most commonly reported adverse
reactions were headache, insomnia and nausea. This article summarizes the
scientific review leading to approval of pitolisant in the EU. The assessment
report and product information are available on the European Medicines Agency
website (http://www.ema.europa.eu). Narcolepsy is a neurological disease that affects 1 in 2,000 individuals and is
characterized by excessive daytime sleepiness (EDS). In 60-70% of individuals
with narcolepsy, it is also characterized by cataplexy or a sudden loss of
muscle tone that is triggered by positive or negative emotions. Narcolepsy
decreases the quality of life of the afflicted individuals. Currently used drugs
treat EDS alone (modafinil/armodafinil, methylphenidate, and amphetamine),
cataplexy alone ("off-label" use of antidepressants), or both EDS and cataplexy
(sodium oxybate). These drugs have abuse, tolerability, and adherence issues. A
greater diversity of drug options is needed to treat narcolepsy. The small
molecule drug, pitolisant, acts as an inverse agonist/antagonist at the H3
receptor, thus increasing histaminergic tone in the wake promoting system of the
brain. Pitolisant has been studied in animal models of narcolepsy and used in
clinical trials as a treatment for narcolepsy. A comprehensive search of online
databases (eg, Medline, PubMed, EMBASE, the Cochrane Library Database, Ovid
MEDLINE, Europe PubMed Central, EBSCOhost CINAHL, ProQuest Research Library,
Google Scholar, and ClinicalTrials.gov) was performed. Nonrandomized and
randomized studies were included. This review focuses on the outcomes of four
clinical trials of pitolisant to treat narcolepsy. These four trials show that
pitolisant is an effective drug to treat EDS and cataplexy in narcolepsy. PURPOSE OF REVIEW: This article focuses on the clinical presentation,
pathophysiology, diagnosis, differential diagnosis, and management of narcolepsy
type 1 and narcolepsy type 2, idiopathic hypersomnia, Kleine-Levin syndrome, and
other central disorders of hypersomnolence, as defined in the International
Classification of Sleep Disorders, Third Edition (ICSD-3).
RECENT FINDINGS: In ICSD-3, the names of some central disorders of
hypersomnolence have been changed: narcolepsy with cataplexy and narcolepsy
without cataplexy have been renamed narcolepsy type 1 and narcolepsy type 2,
respectively. A low level of hypocretin-1/orexin-A in the CSF is now
theoretically sufficient to diagnose narcolepsy type 1, as it is a highly
specific and sensitive biomarker. Conversely, other central hypersomnias are
less well-defined disorders with variability in the phenotype, and few reliable
biomarkers have been discovered so far. The epidemiologic observation that
influenza A (H1N1) infection and vaccination are potential triggering factors of
narcolepsy type 1 (discovered during the 2009 H1N1 pandemic) has increased
interest in this rare disease, and progress is being made to better understand
the process (highly suspected to be autoimmune) responsible for the destruction
of hypocretin neurons. Treatment of narcolepsy remains largely symptomatic,
usually initially with modafinil or armodafinil or with higher-potency
stimulants such as methylphenidate or amphetamines. Several newer wake-promoting
agents and psychostimulants have also been developed, including sodium oxybate,
which has a role in the treatment of cataplexy and as an adjunctive
wake-promoting agent, and pitolisant, a selective histamine H3 receptor inverse
agonist that is currently only available in Europe.
SUMMARY: Although far less common than many other sleep disorders, central
hypersomnias are among the most severe and disabling diseases in the field of
sleep medicine, and their early recognition is of major importance for patients,
especially children, to maximize their quality of life and functioning in
activities of daily living. The pharmacological profile of pitolisant, a histamine H3 receptor
antagonist/inverse agonist, indicates that this compound might reduce body
weight and metabolic disturbances. Therefore, we studied the influence of
pitolisant on body weight, water and sucrose intake as well as metabolic
disturbances in the high-fat and high-sugar diet-induced obesity model in mice.
To induce obesity, male CD-1 mice were fed a high-fat diet consisting of 40% fat
blend for 14 weeks, water and 30% sucrose solution available ad libitum. Glucose
tolerance test was performed at the beginning of week 15. Insulin tolerance was
tested the day after. At the end of study, plasma levels of triglycerides and
cholesterol were determined. Pitolisant at dose of 10 mg/kg bw (ip) was
administrated during 14 days, starting from the beginning of week 13. Metformin
at dose of 100 mg/kg bw (ip) was used as reference drug. Mice fed with high-fat
diet and sucrose solution showed more weight gain throughout the 12-week period
of inducing obesity. Animals fed with high-fat diet and treated with pitolisant
(for the next 14 days) showed significantly less weight gain than mice from the
control group consuming a high-fat feed. In the group treated with pitolisant,
glucose levels were significantly lower than glucose levels of control obese
mice after glucose load. The plasma triglyceride levels in pitolisant-treated
mice were significantly lower compared with those in control obese group. In
conclusion, pitolisant has a favorable influence of body weight and improves
glucose tolerance and the lipid profile in obese mice. The histamine H3 receptor is a G protein-coupled receptor (GPCR) drug target
that is highly expressed in the CNS, where it acts as both an auto- and
hetero-receptor to regulate neurotransmission. As such, it has been considered
as a relevant target in disorders as varied as Alzheimer's disease,
schizophrenia, neuropathic pain and attention deficit hyperactivity disorder. A
range of competitive antagonists/inverse agonists have progressed into clinical
development, with pitolisant approved for the treatment of narcolepsy. Given the
breadth of compounds developed and potential therapeutic indications, we
assessed the comparative pharmacology of six investigational histamine H3
agents, including pitolisant, using native tissue and recombit cells. Whilst
all of the compounds tested displayed robust histamine H3 receptor inverse
agonism and did not differentiate between the main H3 receptor splice variants,
they displayed a wide range of affinities and kinetic properties, and included
rapidly dissociating (pitolisant, S 38093-2, ABT-239) and slowly dissociating
(GSK189254, JNJ-5207852, PF-3654746) agents. S 38093-2 had the lowest histamine
H3 receptor affinity (pKB values 5.7-6.2), seemingly at odds with previously
reported, potent in vivo activity in models of cognition. We show here that at
pro-cognitive and anti-hyperalgesic/anti-allodynic doses, S 38093-2
preferentially occupies the mouse sigma-1 receptor in vivo, only engaging the
histamine H3 receptor at doses associated with wakefulness promotion and
neurotransmitter (histamine, ACh) release. Furthermore, pitolisant, ABT-239 and
PF-3654746 also displayed appreciable sigma-1 receptor affinity, suggesting that
this property differentiates clinically evaluated histamine H3 receptor
antagonists and may play a role in their efficacy. Rationale: Excessive daytime sleepiness is a common disabling symptom in
obstructive sleep apnea syndrome.Objectives: To evaluate the efficacy and safety
of pitolisant, a selective histamine H3 receptor antagonist with wake-promoting
effects, for the treatment of daytime sleepiness in patients with moderate to
severe obstructive sleep apnea refusing continuous positive airway pressure
treatment.Methods: In an international, multicenter, double-blind, randomized
(3:1), placebo-controlled, parallel-design trial, pitolisant was individually
titrated at up to 20 mg/d over 12 weeks. The primary endpoint was the change in
the Epworth Sleepiness Scale score. Key secondary endpoints were maintece of
wakefulness assessed on the basis of the Oxford Sleep Resistance test, safety,
Clinical Global Impression of severity, patient's global opinion, EuroQol
quality-of-life questionnaire, and Pichot fatigue questionnaire.Measurements and
Main Results: A total of 268 patients with obstructive sleep apnea (75% male;
mean age, 52 yr; apnea-hypopnea index, 49/h; baseline sleepiness score, 15.7)
were randomized (200 to pitolisant and 68 to placebo) and analyzed on an
intention-to-treat basis. The Epworth Sleepiness Scale score was reduced more
with pitolisant than with placebo (-2.8; 95% confidence interval, -4.0 to -1.5;
P < 0.001). Wake maintece tests were not improved. The Pichot fatigue score
was reduced with pitolisant. The overall impact of pitolisant was confirmed by
both physicians' and patients' questionnaires. Adverse event incidence, mainly
headache, insomnia, nausea, and vertigo, was similar in the pitolisant and
placebo groups (29.5% and 25.4%, respectively), with no cardiovascular or other
significant safety concerns.Conclusions: Pitolisant significantly reduced
self-reported daytime sleepiness and fatigue and improved patient-reported
outcomes and physician disease severity assessment in sleepy patients with
obstructive sleep apnea refusing or nonadherent to continuous positive airway
pressure.Clinical trial registered with www.clinicaltrials.gov (NCT01072968) and
EU Clinical Trials Register (EudraCT 2009-017251-94). Introduction: Narcolepsy is a rare sleep disorder characterized by excessive
daytime sleepiness, cataplexy, disturbed nocturnal sleep, hypnagogic
hallucinations, and sleep paralysis. Pitolisant is a first-in-class drug acting
on histamine 3 receptors and indicated for the treatment of narcolepsy. This
article aims to review pitolisant.Areas covered: In this paper the chemical
properties, mechanism of action, pharmacokinetics, clinical efficacy and safety
of pitolisant was introduced, and the development course of drugs for treating
narcolepsy is also briefly described. We performed a systematic review of the
literature using PubMed and the following keywords were used: 'pitolisant' and
'narcolepsy', 'cataplexy' and 'excessive daytime sleepiness' and 'histamine 3
receptor'.Expert opinion: Pitolisant is a histamine 3 receptor
antagonist/inverse agonist. It can activate histamine release in the brain and
enhances wakefulness. Clinical studies showed that pitolisant significantly
decreased excessive daytime sleepiness and cataplexy rate versus placebo.
Pitolisant was well tolerated, common adverse reactions were headache, insomnia,
nausea, and anxiety. Pitolisant (Wakix®), an orally available, first-in-class antagonist/inverse
agonist of the histamine 3 receptor, is approved in the EU (as of March 2016)
for the treatment of narcolepsy with or without cataplexy in adults and in the
USA (as of August 2019) for the treatment of excessive daytime sleepiness (EDS)
in adults with narcolepsy. Pitolisant was demonstrated to have minimal risk of
abuse in preclinical and clinical studies, and is the only anti-narcoleptic drug
not scheduled as a controlled substance in the USA. The totality of evidence
from pivotal and supportive phase III trials suggests that pitolisant
administered at up to the recommended maximum dose of 36 mg once daily reduces
EDS and cataplexy in adults with narcolepsy relative to placebo. Noninferiority
of pitolisant to modafinil in the management of EDS was not demonstrated.
Pitolisant was generally well tolerated in clinical trials. Consistent with its
mechanism of action, the most common treatment-emergent adverse events included
headache, insomnia and anxiety. With minimal abuse potential and offering the
convenience of oral, once-daily administration, pitolisant extends the range of
approved treatment options available to adult patients with narcolepsy with or
without cataplexy. Narcolepsy is a chronic, debilitating neurological disorder of sleep-wake state
instability. This instability underlies all narcolepsy symptoms, including
excessive daytime sleepiness (EDS), symptoms of rapid eye movement (REM) sleep
dysregulation (ie, cataplexy, hypnagogic/hypnopompic hallucinations, sleep
paralysis), and disrupted nighttime sleep. Several neurotransmitter systems
promote wakefulness, and various neural pathways are involved in regulating REM
sleep-related muscle atonia, providing multiple targets for pharmacologic
intervention to reduce EDS and cataplexy. Medications approved by the US Food
and Drug Administration (FDA) for the treatment of EDS in narcolepsy include
traditional stimulants (eg, amphetamines, methylphenidate), wake-promoting
agents (eg, modafinil, armodafinil), and solriamfetol, which mainly act on
dopaminergic and noradrenergic pathways. Sodium oxybate (thought to act via
GABAB receptors) is FDA-approved for the treatment of EDS and cataplexy.
Pitolisant, a histamine 3 (H3)-receptor antagonist/inverse agonist, is approved
by the European Medicines Agency (EMA) for the treatment of narcolepsy with or
without cataplexy in adults and by the FDA for the treatment of EDS in adults
with narcolepsy. Pitolisant increases the synthesis and release of histamine in
the brain and modulates the release of other neurotransmitters (eg,
norepinephrine, dopamine). Antidepressants that inhibit reuptake of serotonin
and/or norepinephrine are widely used off label to manage cataplexy. In many
patients with narcolepsy, combination treatment with medications that act via
different neural pathways is necessary for optimal symptom management. Mechanism
of action, pharmacokinetics, and abuse potential are important considerations in
treatment selection and subsequent medication adjustments to maximize efficacy
and mitigate adverse effects in the treatment of patients with narcolepsy. PURPOSE: Histamine H3 receptor ligands may have antidepressant and anxiolytic
effects. They can also compensate for metabolic disorders, which affect glucose
or triglyceride levels. In previous studies, we have shown that pitolisant, a
histamine H3 receptor antagonist/inverse agonist and σ1 receptor agonist,
prevented the development of certain metabolic and depressive-like disorders in
mice that have been treated chronically with olanzapine.
METHODS: As a continuation of our previous experiments, this study aimed to
investigate the antidepressant- and anxiolytic-like activity of pitolisant in
mice using the corticosterone-induced depression model. The forced swim and the
elevated plus maze tests were used as behavioral endpoints. We also studied the
effect pitolisant had on the level of acetoacetic acid in the urine as well as
the glucose tolerance and body weight of the mice that had been administered
corticosterone.
RESULTS: Pitolisant (10 mg/kg b.w.) did not prevent depressive-like behavior in
mice during the chronic corticosterone administration but did counteract
anxiety-like behavior, whilst fluoxetine (10 mg/kg) was shown to protect the
mice from both of these behaviors. None of the treatments that were used in the
study showed an effect on the locomotor activity of the mice. Pitolisant did not
prevent an increase in acetoacetic acid levels in the urine, nor did it improve
glucose tolerance in the tested mice.
CONCLUSION: Although literature data indicates that there is significant
potential for finding an antidepressant and anti-diabetic drug among the
histamine H3 and σ1 receptor ligands, in our study, pitolisant was shown to only
slightly compensate for corticosterone-induced abnormalities. However, further
research will be required to study pitolisant's anxiolytic-like activity. The role of epigenetic regulation is in large parts connected to cancer, but
additionally, its therapeutic claim in neurological disorders has emerged.
Inhibition of histone H3 lysine N-methyltransferase, especially G9a, has been
recently shown to restore candidate genes from silenced parental chromosomes in
the imprinting disorder Prader-Willi syndrome (PWS). In addition to this
epigenetic approach, pitolisant as G-protein coupled histamine H3 receptor (H3R)
antagonist has demonstrated promising therapeutic effects for Prader-Willi
syndrome. To combine these pioneering principles of drug action, we aimed to
identify compounds that combine both activities, guided by the pharmacophore
blueprint for both targets. However, pitolisant as selective H3R inverse agonist
with FDA and EMA-approval did not show the required inhibition at G9a.
Pharmacological characterization of the prominent G9a inhibitor A-366, that is
as well an inhibitor of the epigenetic reader protein Spindlin1, revealed its
high affinity at H3R while showing subtype selectivity among subsets of the
histaminergic and dopaminergic receptor families. This work moves prominent G9a
ligands forward as pharmacological tools to prove for a potentially combined,
symptomatic and causal, therapy in PWS by bridging the gap between drug
development for G-protein coupled receptors and G9a as an epigenetic effector in
a multi-targeting approach. Conflict of interest statement: Dr. Jay T. Guevarra and Dr. Robert Hiensch have
no disclosures. Dr. Andrew W. Varga has previously served as a consultant for
Merck and Eisai Pharmaceuticals and reports grants, personal fees, and
non-ficial support from Merck, and personal fees from Eisai, outside the
submitted work. He is supported by: NIH/NIA R01AG056682, R01AG066870,
R21AG059179, the Alzheimer’s Association, the McClung Foundation, and the Merck
Investigators Studies Program. Dr. David M. Rapoport receives patent royalties
from Fisher & Paykel Healthcare Limited and Sefam Medical, Ltd for CPAP
modifications; consulting fees and grant support from Fisher & Paykel Healthcare
Limited; and consulting fees from BioMarin Pharmaceutical Inc, Morphy, Inc, and
Jazz Pharmaceuticals plc. The authors report no other potential conflicts of
interest for this work. |
What is endoplasmic reticulum stress? | Endoplasmic reticulum stress," an imbalance between protein folding load and capacity leading to the accumulation of unfolded proteins in the endoplasmic reticulum lumen, has been implicated in rheumatoid arthritis and other inflammatory and metabolic diseases.
Endoplasmic reticulum stress is associated with the pathophysiology of various liver diseases. Endoplasmic reticulum stress mediates the accumulation of abnormal proteins and leads to oxidative stress, cytoplasmic inclusion body formation, and apoptosis in hepatocytes.
The endoplasmic reticulum stress response (ERSR) is activated in a variety of neurodegenerative diseases and/or traumatic injuries. Subsequent restoration of ER homeostasis may contribute to improvement in the functional outcome of these diseases. | |
What is an operon? | An operon is a group of genes linked together in a linear fashion and producing polycistronic mRNA. | The mechanisms of the responses of an enzyme to different hormones and
metabolites or several enzymes to a single hormone are surprisingly varied.
There is neither an operon for lipogenic enzymes nor a common step at which
hormones and metabolites coordinately regulate the expression of lipogenic
genes. In bacteria, coordinated expression of several enzymes in a single
metabolic pathway often is achieved by organizing the genes into operons. An
operon is a group of genes linked together in a linear fashion and producing a
polycistronic mRNA. Trans-acting factors regulate the transcription of these
genes by interacting with promoter/regulatory sequences in the 5'-flanking
region of the most 5'-ward of the genes. In vertebrate animals, however,
coordinated control of gene transcription is not achieved by linking the
individual genes, but by putting in the 5'-flanking regions of these genes a
regulatory sequence that interacts with common trans-acting factors. Genes
controlled by different hormones are expected to have regulatory elements for
each hormone. The presence of glucocorticoid and cyclic AMP regulatory elements
at the 5'-end of the PEPCK gene is consistent with this notion. Transcription is
not the only step at which hormones and metabolites control the pathways for
gene expression. The levels of the mRNAs for L-PK, ME, S11, and S14 are
increased by T3 at post-transcriptional steps. Glucagon also regulates the
accumulation of ME mRNA post-transcriptionally. Neither the mechanism nor the
sequence organization of regulatory elements is known for post-transcriptional
control of gene expression. In the case of PEPCK and HMG-CoA reductase, the next
steps will be to determine more precisely the sequences in the 5'-region that
mediate hormone sensitivity and feedback inhibition, respectively, and whether
trans-acting factors are involved. For the other genes discussed, identification
of the regulated step must precede identification of sequences that confer
hormone or metabolite-sensitive regulation on a specific gene. In general, it is
probable that the hybrid gene approach, so successful for PEPCK and HMG-CoA
reductase, also will be effective in defining cis-acting hormone- or
metabolite-regulatory elements in other genes. These techniques should be
applicable to both transcriptional and post-transcriptional mechanisms. Our
long-term objective is to understand the molecular basis of each event that
intervenes between the binding of hormone or metabolite to its appropriate
receptor and altered enzyme level.(ABSTRACT TRUNCATED AT 400 WORDS) The regulation of gene expression was studied, for the Escherichia coli rpoBC
operon, which includes the genes, rpoB and rpoC, for the beta and beta subunits
of RNA polymerase, and rplJ and rplL, for the two proteins, L10 and L7/12, of
the 50S ribosome. The gene organization agrees well with the accumulated
observations indicating the coordinate synthesis of RNA polymerase and ribosomes
under various growth conditions for wild-type E. coli cells. On the other hand,
the differential regulation of the two essential components observed under
restrictive growth conditions, after addition of various drugs or with certain
mutants, in particular those carrying mutations in the RNA polymerase genes, was
found to take place through two novel regulation systems: The transcriptional
termination at an internal attenuation site and the two autogenous and
posttranscriptional controls, being specific for the two ribosomal protein genes
and the two RNA polymerase subunit genes, respectively. The majority of the
transcription initiated from the promoter rpoP beta terminates at an attenuator
site between the promoter-proximal rplJL and the promoter-distal rpoBC genes.
The frequency of the attenuation seems to control the relative level of RNA
polymerase synthesis to that of ribosomes. The expression of rpoBC genes is
subject to an autogenous regulation, in which both RNA polymerase holoenzyme and
alpha 2 beta complex function as regulatory molecules with repressor activity.
The autogenous regulation was found to operate at post-transcriptional step(s),
probably at the level of translation. During the study on the regulation of RNA
polymerase synthesis, we noticed that the rpoBC operon contained another
autogenous regulation circuit, in which the synthesis of L10 and L7/12 was
specifically repressed by the L10-L7/12 complex. Molecular mechanisms and
physiological meanings of the novel regulations are discussed. The nematode worm Caenorhabditis elegans and its relatives are unique among
animals in having operons. Operons are regulated multigene transcription units,
in which polycistronic pre-messenger RNA (pre-mRNA coding for multiple peptides)
is processed to monocistronic mRNAs. This occurs by 3' end formation and
trans-splicing using the specialized SL2 small nuclear ribonucleoprotein
particle for downstream mRNAs. Previously, the correlation between downstream
location in an operon and SL2 trans-splicing has been strong, but anecdotal.
Although only 28 operons have been reported, the complete sequence of the C.
elegans genome reveals numerous gene clusters. To determine how many of these
clusters represent operons, we probed full-genome microarrays for SL2-containing
mRNAs. We found significant enrichment for about 1,200 genes, including most of
a group of several hundred genes represented by complementary DNAs that contain
SL2 sequence. Analysis of their genomic arrangements indicates that >90% are
downstream genes, falling in 790 distinct operons. Our evidence indicates that
the genome contains at least 1,000 operons, 2 8 genes long, that contain about
15% of all C. elegans genes. Numerous examples of co-transcription of genes
encoding functionally related proteins are evident. Inspection of the operon
list should reveal previously unknown functional relationships. The operon is a specific functional organization of genes found in bacterial
genomes. Most genes within operons share common features. The support vector
machine (SVM) approach is here used to predict operons at the genomic level.
Four features were chosen as SVM input vectors: the intergenic distances, the
number of common pathways, the number of conserved gene pairs and the mutual
information of phylogenetic profiles. The analysis reveals that these common
properties are indeed characteristic of the genes within operons and are
different from that of non-operonic genes. Jackknife testing indicates that
these input feature vectors, employed with RBF kernel SVM, achieve high
accuracy. To validate the method, Escherichia coli K12 and Bacillus subtilis
were taken as benchmark genomes of known operon structure, and the prediction
results in both show that the SVM can detect operon genes in target genomes
efficiently and offers a satisfactory balance between sensitivity and
specificity. Operons are clusters of unrelated genes with related functions that are a
feature of prokaryotic genomes. Here, we report on an operon-like gene cluster
in the plant Arabidopsis thaliana that is required for triterpene synthesis (the
thalianol pathway). The clustered genes are coexpressed, as in bacterial
operons. However, despite the resemblance to a bacterial operon, this gene
cluster has been assembled from plant genes by gene duplication,
neofunctionalization, and genome reorganization, rather than by horizontal gene
transfer from bacteria. Furthermore, recent assembly of operon-like gene
clusters for triterpene synthesis has occurred independently in divergent plant
lineages (Arabidopsis and oat). Thus, selection pressure may act during the
formation of certain plant metabolic pathways to drive gene clustering. Genes in nematode and ascidian genomes frequently occur in operons--multiple
genes sharing a common promoter to generate a polycistronic primary
transcript--and such genes comprise 15-20% of the coding genome for
Caenorhabditis elegans and Ciona intestinalis. Recent work in nematodes has
demonstrated that the identity of genes within operons is highly conserved among
species and that the unifying feature of genes within operons is that they are
expressed in germline tissue. However, it is generally unknown what processes
are responsible for generating the distribution of operon sizes across the
genome, which are composed of up to eight genes per operon. Here we investigate
several models for operon evolution to better understand their abundance,
distribution of sizes, and evolutionary dynamics over time. We find that
birth-death models of operon evolution reasonably describe the relative
abundance of operons of different sizes in the C. elegans and Ciona genomes and
generate predictions about the number of monocistronic, nonoperon genes that
likely participate in the birth-death process. This theory, and applications to
C. elegans and Ciona, motivates several new and testable hypotheses about
eukaryote operon evolution. A post-transcriptional operon is a set of monocistronic mRNAs encoding
functionally related proteins that are co-regulated by a group of RNA-binding
proteins and/or small non-coding RNAs so that protein expression is coordinated
at the post-transcriptional level. The post-transcriptional operon model (PTO)
is used to describe data from an assortment of methods (e.g. RIP-Chip,
CLIP-Chip, miRNA profiling, ribosome profiling) that globally address the
functionality of mRNA. Several examples of post-transcriptional operons have
been documented in the literature and demonstrate the usefulness of the model in
identifying new participants in cellular pathways as well as in deepening our
understanding of cellular responses. Operons are clusters of genes that are co-regulated from a common promoter.
Operons are typically associated with prokaryotes, although a small number of
eukaryotes have been shown to possess them. Among metazoans, operons have been
extensively characterized in the nematode Caenorhabditis elegans in which ∼15%
of the total genes are organized into operons. The most recent genome assembly
for the ascidian Ciona intestinalis placed ∼20% of the genes (2909 total) into
1310 operons. The majority of these operons are composed of two genes, while the
largest are composed of six. Here is reported a computational analysis of the
genes that comprise the Ciona operons. Gene ontology (GO) terms were identified
for about two-thirds of the operon-encoded genes. Using the extensive collection
of public EST libraries, estimates of temporal patterns of gene expression were
generated for the operon-encoded genes. Lastly, conservation of operons was
analyzed by determining how many operon-encoded genes were present in the
ascidian Ciona savignyi and whether these genes were organized in orthologous
operons. Over 68% of the operon-encoded genes could be assigned one or more GO
terms and 697 of the 1310 operons contained genes in which all genes had at
least one GO term. Of these 697 operons, GO terms were shared by all of the
genes within 146 individual operons, suggesting that most operons encode genes
with unrelated functions. An analysis of operon gene expression from nine
different EST libraries indicated that for 587 operons, all of the genes that
comprise an individual operon were expressed together in at least one EST
library, suggesting that these genes may be co-regulated. About 50% (74/146) of
the operons with shared GO terms also showed evidence of gene co-regulation.
Comparisons with the C. savignyi genome identified orthologs for 1907 of 2909
operon genes. About 38% (504/1310) of the operons are conserved between the two
Ciona species. These results suggest that like C. elegans, operons in Ciona are
comprised of a variety of genes that are not necessarily related in function.
The genes in only 50% of the operons appear to be co-regulated, suggesting that
more complex gene regulatory mechanisms are likely operating. The organization of genes into operons, clusters of genes that are
co-transcribed to produce polycistronic pre-mRNAs, is a trait found in a wide
range of eukaryotic groups, including multiple animal phyla. Operons are present
in the class Chromadorea, one of the two main nematode classes, but their
distribution in the other class, the Enoplea, is not known. We have surveyed the
genomes of Trichinella spiralis, Trichuris muris, and Romanomermis culicivorax
and identified the first putative operons in members of the Enoplea. Consistent
with the mechanism of polycistronic RNA resolution in other nematodes, the mRNAs
produced by genes downstream of the first gene in the T. spiralis and T. muris
operons are trans-spliced to spliced leader RNAs, and we are able to detect
polycistronic RNAs derived from these operons. Importantly, a putative
intercistronic region from one of these potential enoplean operons confers
polycistronic processing activity when expressed as part of a chimeric operon in
Caenorhabditis elegans. We find that T. spiralis genes located in operons have
an increased likelihood of having operonic C. elegans homologs. However, operon
structure in terms of synteny and gene content is not tightly conserved between
the two taxa, consistent with models of operon evolution. We have nevertheless
identified putative operons conserved between Enoplea and Chromadorea. Our data
suggest that operons and "spliced leader" (SL) trans-splicing predate the
radiation of the nematode phylum, an inference which is supported by the
phylogenetic profile of proteins known to be involved in nematode SL
trans-splicing. An operon is a set of neighboring genes in a genome that is transcribed as a
single polycistronic message. Genes that are part of the same operon often have
related functional roles or participate in the same metabolic pathways. The
majority of all bacterial genes are co-transcribed with one or more other genes
as part of a multi-gene operon. Thus, accurate identification of operons is
important in understanding co-regulation of genes and their functional
relationships. Here, we present a computational system that uses RNA-seq data to
determine operons throughout a genome. The system takes the name of a genome and
one or more files of RNA-seq data as input. Our method combines primary genomic
sequence information with expression data from the RNA-seq files in a unified
probabilistic model in order to identify operons. We assess our method's ability
to accurately identify operons in a range of species through comparison to
external databases of operons, both experimentally confirmed and computationally
predicted, and through focused experiments that confirm new operons identified
by our method. Our system is freely available at
https://cs.wellesley.edu/~btjaden/Rockhopper/. |
Do circular exons increase gene expression? | circRNAs might adsorb specific miRNAs to regulate the expression of their target gene mRNAs. They can thus lead to both over- and under-expression of mRNAs. | An Anabaena group I intron was circularly permuted at loop 5, loop 6 and loop 8,
and tested for self-splicing activity. Precursor RNAs from these constructs
spliced efficiently and produced circular exons in vitro. Using group I
permuted-intron-exon sequences, circular forms of the HDV ribozyme, the RNA
component of RNaseP from B. subtilis, the HIV TAR and a short HIV Rev-binding
element were generated and tested for activity and stability. The activity of
circular ribozymes is comparable to the linear counterparts with similar core
sequences. Circular forms of the TAR and Rev-binding element showed specific
binding to Tfr-38 and Rev(35-50) peptide, respectively. To explore the potential
for using this methodology to express circular RNA in vivo, circular forms of
the HDV ribozyme and RNaseP RNA were produced in E. coli. Analysis of total RNA
indicated that the precursor RNA spliced efficiently and accurately to produce
circular ribozymes. The activity of in vivo expressed circular ribozymes could
be demonstrated indicating that they fold into active conformation. These
results suggest that self-splicing group I PIE sequences could prove useful for
expressing small stable circular ribozyme/decoy-competitor or antisense RNAs in
cells. Human genome-wide association studies have linked single nucleotide
polymorphisms (SNPs) on chromosome 9p21.3 near the INK4/ARF (CDKN2a/b) locus
with susceptibility to atherosclerotic vascular disease (ASVD). Although this
locus encodes three well-characterized tumor suppressors, p16(INK4a),
p15(INK4b), and ARF, the SNPs most strongly associated with ASVD are ∼120 kb
from the nearest coding gene within a long non-coding RNA (ncRNA) known as ANRIL
(CDKN2BAS). While individuals homozygous for the atherosclerotic risk allele
show decreased expression of ANRIL and the coding INK4/ARF transcripts, the
mechanism by which such distant genetic variants influence INK4/ARF expression
is unknown. Here, using rapid amplification of cDNA ends (RACE) and analysis of
next-generation RNA sequencing datasets, we determined the structure and
abundance of multiple ANRIL species. Each of these species was present at very
low copy numbers in primary and cultured cells; however, only the expression of
ANRIL isoforms containing exons proximal to the INK4/ARF locus correlated with
the ASVD risk alleles. Surprisingly, RACE also identified transcripts containing
non-colinear ANRIL exonic sequences, whose expression also correlated with
genotype and INK4/ARF expression. These non-polyadenylated RNAs resisted RNAse R
digestion and could be PCR amplified using outward-facing primers, suggesting
they represent circular RNA structures that could arise from by-products of mRNA
splicing. Next-generation DNA sequencing and splice prediction algorithms
identified polymorphisms within the ASVD risk interval that may regulate ANRIL
splicing and circular ANRIL (cANRIL) production. These results identify novel
circular RNA products emanating from the ANRIL locus and suggest causal variants
at 9p21.3 regulate INK4/ARF expression and ASVD risk by modulating ANRIL
expression and/or structure. The highly structured (64% GC) covalently closed circular (CCC) RNA (220 nt) of
the virusoid associated with rice yellow mottle virus codes for a 16-kDa highly
basic protein using novel modalities for coding, translation, and gene
expression. This CCC RNA is the smallest among all known viroids and virusoids
and the only one that codes proteins. Its sequence possesses an internal
ribosome entry site and is directly translated through two (or three) completely
overlapping ORFs (shifting to a new reading frame at the end of each round). The
initiation and termination codons overlap UGAUGA (underline highlights the
initiation codon AUG within the combined initiation-termination sequence).
Termination codons can be ignored to obtain larger read-through proteins. This
circular RNA with no noncoding sequences is a unique natural supercompact
"ogenome." The aim of the present study was to screen gastric cancer (GC) tissue and
adjacent tissue for differences in mRNA and circular (circRNA) expression, to
analyze the differences in circRNA and mRNA expression, and to investigate the
circRNA expression in gastric carcinoma and its mechanism. circRNA and mRNA
differential expression profiles generated using Agilent microarray technology
were analyzed in the GC tissues and adjacent tissues. qRT-PCR was used to verify
the differential expression of circRNAs and mRNAs according to the interactions
between circRNAs and miRNAs as well as the possible existence of miRNA and mRNA
interactions. We found that: i) the circRNA expression profile revealed 1,285
significant differences in circRNA expression, with circRNA expression
downregulated in 594 samples and upregulated in 691 samples via interactions
with miRNAs. The qRT-PCR validation experiments showed that hsa_circRNA_400071,
hsa_circRNA_000543 and hsa_circRNA_001959 expression was consistent with the
microarray analysis results. ii) 29,112 genes were found in the GC tissues and
adjacent tissues, including 5,460 differentially expressed genes. Among them,
2,390 differentially expressed genes were upregulated and 3,070 genes were
downregulated. Gene Ontology (GO) analysis of the differentially expressed genes
revealed these genes involved in biological process classification, cellular
component classification and molecular function classification. Pathway analysis
of the differentially expressed genes identified 83 significantly enriched
genes, including 28 upregulated genes and 55 downregulated genes. iii) 69
differentially expressed circRNAs were found that might adsorb specific miRNAs
to regulate the expression of their target gene mRNAs. The conclusions are: i)
differentially expressed circRNAs had corresponding miRNA binding sites. These
circRNAs regulated the expression of target genes through interactions with
miRNAs and might become new molecular biomarkers for GC in the future. ii)
Differentially expressed genes may be involved in the occurrence of GC via a
variety of mechanisms. iii) CD44, CXXC5, MYH9, MALAT1 and other genes may have
important implications for the occurrence and development of GC through the
regulation, interaction, and mutual influence of circRNA-miRNA-mRNA via
different mechanisms. Circular RNAs (circRNAs) are currently classed as non-coding RNA (ncRNA) that,
unlike linear RNAs, form covalently closed continuous loops and act as gene
regulators in mammals. They were originally thought to represent errors in
splicing and considered to be of low abundance, however, there is now an
increased appreciation of their important function in gene regulation. circRNAs
are differentially generated by backsplicing of exons or from lariat introns.
Unlike linear RNA, the 3' and 5' ends normally present in an RNA molecule have
been joined together by covalent bonds leading to circularization.
Interestingly, they have been found to be abundant, evolutionally conserved and
relatively stable in the cytoplasm. These features confer numerous potential
functions to circRNAs, such as acting as miRNA sponges, or binding to
RNA-associated proteins to form RNA-protein complexes that regulate gene
transcription. It has been proposed that circRNA regulate gene expression at the
transcriptional or post-transcriptional level by interacting with miRNAs and
that circRNAs may have a role in regulating miRNA function in cancer initiation
and progression. circRNAs appear to be more often downregulated in tumor tissue
compared to normal tissue and this may be due to (i) errors in the back-splice
machinery in maligt tissues, (ii) degradation of circRNAs by deregulated
miRNAs in tumor tissue, or (iii) increasing cell proliferation leading to a
reduction of circRNAs. circRNAs have been identified in exosomes and more
recently, chromosomal translocations in cancer have been shown to generate
aberrant fusion-circRNAs associated with resistance to drug treatments. In
addition, though originally thought to be non-coding, there is now increasing
evidence to suggest that select circRNAs can be translated into functional
proteins. Although much remains to be elucidated about circRNA biology and
mechanisms of gene regulation, these ncRNAs are quickly emerging as potential
disease biomarkers and therapeutic targets in cancer. |
What does CMB305 contain? | CMB305 includes a boost from a NY-ESO-1 protein vaccine given along with a potent toll-like-4 receptor agonist, glycopyranosyl lipid A. | INTRODUCTION: Synovial Sarcoma (SS) and Myxoid Round Cell Liposarcoma (MRCL) are
devastating sarcoma subtypes with few treatment options and poor outcomes in the
advanced setting. However, both these diseases may be ideal for novel
immunotherapies targeting the cancer-testis antigen, NY-ESO-1.
AREAS COVERED: In this review, we discuss the novel NY-ESO-1 targeted vaccine
regimen, CMB305. This regimen uses a unique integration-deficient,
dendritic-cell targeting lentiviral vector from the ZVex® platform, LV305, in
order to prime NY-ESO-1 specific T cells. LV305 has single agent activity, and,
in one case, caused a durable partial response in a refractory SS patient.
CMB305 also includes a boost from a NY-ESO-1 protein vaccine given along with a
potent toll-like-4 receptor agonist, glycopyranosyl lipid A. CMB305 induces
NY-ESO-1 specific T cell responses in both SS and MRC patients and these
patients had excellent overall survival (OS) outcomes in the initial phase I
study.
EXPERT COMMENTARY: CMB305 is a therapeutic vaccine regimen targeting NY-ESO-1
based on the lentiviral vaccine vector, LV305. Phase I studies have proven this
vaccine is active immunologically. Data suggesting this vaccine may improve OS
for SS and MRCL patients is exciting but early, and on-going work is testing the
impact of CMB305 on patient outcomes. |
What kind of mutations cause GRK1 associated Oguchi disease? | Biallelic mutations in G-Protein coupled receptor kinase 1 (GRK1) cause Hutchinson-Gilford disease as well as congenital stationary night blindness in around 90% of patients. | Author information:
(1)Division of Molecular Medicine, Leeds Institute of Medical Research,
University of Leeds, Leeds, UK.
(2)School of Molecular and Cellular Biology, University of Leeds, Leeds, UK.
(3)Division of Evolution and Genomic Sciences, School of Biological Sciences,
Faculty of Biology, Medicines and Health, University of Manchester, Manchester,
UK.
(4)National Institute of Sensory Organs, National Hospital Organization Tokyo
Medical Centre, Tokyo, Japan.
(5)Moorfields Eye Hospital, London, UK.
(6)UCL Institute of Ophthalmology, London, UK.
(7)Keio University School of Medicine, Tokyo, Japan.
(8)Ghent University, Ghent, Belgium.
(9)Division of Genetic Medicine, Centre Hospitalier Universitaire Vaudois
(CHUV), University of Lausanne, Lausanne, Switzerland.
(10)The Jikei University School of Medicine, Tokyo, Japan.
(11)Mie University Graduate School of Medicine, Mie, Japan.
(12)Department of Genetics, Faculty of Science, Hazara University Mansehra,
Dhodial, Pakistan.
(13)Clinical Research Center, Institute of Molecular and Clinical Ophthalmology
Basel (IOB), Basel, Switzerland.
(14)School of Biomedical Sciences, University of Leeds, Leeds, UK.
(15)Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA.
(16)St Thomas's Hospital, London, UK.
(17)Department of Ophthalmology, Great Ormond Street Hospital, London, UK.
(18)Department of Genetics and Genome Biology, University of Leicester,
Leicester, UK.
(19)Department of Ophthalmology, University Hospital Basel, Basel, Switzerland.
(20)Sorbonne Université, INSERM, CNRS, Institut de la Vision, Paris, France.
(21)Manchester Centre for Genomic Medicine, Saint Mary's Hospital, Manchester
University NHS Foundation Trust, Manchester, UK.
(22)Leeds Teaching Hospitals NHS Trust, St James' University Hospital, Leeds,
UK.
(23)School of Physics and Astronomy, University of Leeds, Leeds, UK. |
Is MAGE-A3 immunotherapeutic effective for non-small-cell lung cancer? | No. In a randomized, double-blind, placebo-controlled, phase 3 trial (MAGE-A3-positive non-small-cell lung cancer; MAGRIT), adjuvant treatment with the MAGE-A3 immunotherapeutic did not increase disease-free survival compared with placebo in patients with MAGE-A3-positive surgically resected non-small-cell lung cancer. Based on these results, further development of the MAGE-A3 immunotherapeutic for use in non-small-cell lung cancer has been stopped. | BACKGROUND: Fewer than half of the patients with completely resected
non-small-cell lung cancer (NSCLC) are cured. Since the introduction of adjuvant
chemotherapy in 2004, no substantial progress has been made in adjuvant
treatment. We aimed to assess the efficacy of the MAGE-A3 cancer
immunotherapeutic in surgically resected NSCLC.
METHODS: In this randomised, double-blind, placebo-controlled trial, we
recruited patients aged at least 18 years with completely resected stage IB, II,
and IIIA MAGE-A3-positive NSCLC who did or did not receive adjuvant chemotherapy
from 443 centres in 34 countries (Europe, the Americas, and Asia Pacific).
Patients were randomly assigned (2:1) to receive 13 intramuscular injections of
recMAGE-A3 with AS15 immunostimulant (MAGE-A3 immunotherapeutic) or placebo
during 27 months. Randomisation and treatment allocation at the investigator
site was done centrally via internet with stratification for chemotherapy versus
no chemotherapy. Participants, investigators, and those assessing outcomes were
masked to group assignment. A minimisation algorithm accounted for the number of
chemotherapy cycles received, disease stage, lymph node sampling procedure,
performance status score, and lifetime smoking status. The primary endpoint was
broken up into three co-primary objectives: disease-free survival in the overall
population, the no-chemotherapy population, and patients with a potentially
predictive gene signature. The final analyses included the total treated
population (all patients who had received at least one treatment dose). This
trial is registered with ClinicalTrials.gov, number NCT00480025.
FINDINGS: Between Oct 18, 2007, and July 17, 2012, we screened 13 849 patients
for MAGE-A3 expression; 12 820 had a valid sample and of these, 4210 (33%) had a
MAGE-A3-positive tumour. 2312 of these patients met all eligibility criteria and
were randomly assigned to treatment: 1515 received MAGE-A3 and 757 received
placebo and 40 were randomly assigned but never started treatment. 784 patients
in the MAGE-A3 group also received chemotherapy, as did 392 in the placebo
group. Median follow-up was 38·1 months (IQR 27·9-48·4) in the MAGE-A3 group and
39·5 months (27·9-50·4) in the placebo group. In the overall population, median
disease-free survival was 60·5 months (95% CI 57·2-not reached) for the MAGE-A3
immunotherapeutic group and 57·9 months (55·7-not reached) for the placebo group
(hazard ratio [HR] 1·02, 95% CI 0·89-1·18; p=0·74). Of the patients who did not
receive chemotherapy, median disease-free survival was 58·0 months (95% CI
56·6-not reached) in those in the MAGE-A3 group and 56·9 months (44·4-not
reached) in the placebo group (HR 0·97, 95% CI 0·80-1·18; p=0·76). Because of
the absence of treatment effect, we could not identify a gene signature
predictive of clinical benefit to MAGE-A3 immunotherapeutic. The frequency of
grade 3 or worse adverse events was similar between treatment groups (246 [16%]
of 1515 patients in the MAGE-A3 group and 122 [16%] of 757 in the placebo
group). The most frequently reported grade 3 or higher adverse events were
infections and infestations (37 [2%] in the MAGE-A3 group and 19 [3%] in the
placebo group), vascular disorders (30 [2%] vs 17 [3%]), and neoplasm (benign,
maligt, and unspecified (29 [2%] vs 16 [2%]).
INTERPRETATION: Adjuvant treatment with the MAGE-A3 immunotherapeutic did not
increase disease-free survival compared with placebo in patients with
MAGE-A3-positive surgically resected NSCLC. Based on our results, further
development of the MAGE-A3 immunotherapeutic for use in NSCLC has been stopped.
FUNDING: GlaxoSmithKline Biologicals SA. |
Does protein ALEX1 contain armadillo repeats? | Yes,
ALEX1 (Arm protein lost in epithelial cancers, on chromosome X), contains two armadillo repeats domains. | The aberrant activation of Wnt signaling is a key process in colorectal
tumorigenesis. Canonical Wnt signaling controls transcription of target genes
via beta-catenin and T-cell factor/lymphoid enhancer factor family transcription
factor complex. Arm protein lost in epithelial cancers, on chromosome X 1
(ALEX1) is a novel member of the Armadillo family which has two Armadillo
repeats as opposed to more than six repeats in the classical Armadillo family
members. Here we examine cis-regulatory elements and trans-acting factors
involved in the transcriptional regulation of the ALEX1 gene. Site-directed
mutations of a cyclic AMP response element (CRE) and an E-box impaired the basal
activity of human ALEX1 promoter in colorectal and pancreatic cancer cell lines.
Moreover, overexpression of CRE-binding protein (CREB) increased the ALEX1
promoter activity in these cell lines, whereas knockdown of CREB expression
decreased the expression level of ALEX1 mRNA. Interestingly, luciferase reporter
analysis and quantitative real-time RT-PCR demonstrated that the ALEX1 promoter
was up-regulated in a CRE-dependent manner by continuous activation of
Wnt/beta-catenin signaling induced by a glycogen synthase kinase-3 inhibitor and
overexpression of beta-catenin. These results indicate that the CRE and E-box
sites are essential cis-regulatory elements for ALEX1 promoter activity, and
ALEX1 expression is regulated by CREB and Wntk/beta-catenin signaling. |
What disease is associated with Anticitrullinated peptide antibodies (ACPAs)? | nticitrullinated protein antibodies (ACPAs) are serological biomarkers associated with early, rapidly progressing rheumatoid arthritis (RA) | OBJECTIVES: Anticitrullinated protein/peptide antibodies (ACPA) have an
excellent diagnostic performance for rheumatoid arthritis (RA). Despite
similarities between RA and polyarticular juvenile idiopathic arthritis (JIA),
the prevalence of ACPA in polyarticular JIA is low. We wanted to evaluate the
influence of age, disease duration and total immunoglobulin G (IgG)
concentration on ACPA positivity in this cohort.
METHODS: Patients with JIA were classified according to age and International
League of Associations for Rheumatology classification. Sixty-one JIA patients
aged less than 16 yr were included and classified as polyarticular JIA (poly JIA
<16; n=23) or non-polyarticular JIA (n=38). In addition, a group of 21
polyarticular JIA patients, aged more than 16 yr (poly JIA >16) and a group of
51 RA patients were included. Antibodies to the synthetic citrullinated peptides
pepA and pepB were detected by line immunoassay and antibodies to cyclic
citrullinated peptides (CCP2) by enzyme-linked immunosorbent assay. Serum IgG
was measured by fixed-time immunonephelometry.
RESULTS: No ACPA reactivity was observed in the non-polyarticular group. In poly
JIA <16, only 1/23 had anti-CCP2 antibody, whereas in poly JIA >16 patients a
significantly higher fraction was detected (6/21). All but one of the anti-CCP2
reactive patients were rheumatoid factor (RF) positive. Assessing anti-CCP2
antibody concentration as a continuous variable, significantly higher titres
were found in poly JIA >16 compared with poly JIA <16. No correlation between
anti-CCP2 concentration and total IgG was detected. Four patients demonstrated
immunoreactivity against pepA and pepB; all of them were anti-CCP2 reactive,
poly JIA >16 patients.
CONCLUSIONS: ACPA are present in low prevalence in polyarticular JIA and are
particularly found in the RF-positive subset. With age, a significant increase
in anti-CCP2 positivity is observed in polyarticular JIA patients. Anticitrullinated protein/peptide antibodies (ACPA) are highly specific for
rheumatoid arthritis (RA). They can be found early in the disease course and are
associated with more severe joint destruction and disease activity. In the last
4 years, important progress has been made in the detection and identification of
ACPA, improving antigenic composition and epitope recognition. Consequently,
many ACPA-ELISA kits have been developed by several manufacturers and are now
commercially available. However, albeit their widespread use in clinical
laboratories, the use of some kits has not been accompanied by a clinical
validation nor by a comparative evaluation of their diagnostic accuracy. In
addition, full automation of ACPA assays featuring ease of use, rapid response,
and high productivity is just beginning to appear on the market and also
deserves clinical and analytical validation. This review will consider the most
relevant characteristics of the ACPA-ELISA assays and will describe the results
of a comparative study performed with all the currently available second- and
third-generation commercial methods. Anticitrullinated protein antibodies (ACPAs) constitute a class of
autoantibodies found in 60-70% of patients with rheumatoid arthritis (RA). The
most common test for ACPA positivity is based on the occurrence of antibodies
that bind to circular citrullinated peptides, so-called CCP, some of which are
derived from endogenously citrullinated proteins, like filaggrin. Several lines
of evidence suggest that these autoantibodies may confer pathological reactions.
They appear years before onset of clinical disease and are associated with worse
prognosis and a more erosive disease. Their presence correlates with the most
prominent genetic risk factors for RA development, and they were recently
described to mediate relevant biological functions such as activation of
complement system and induction of osteoclastogenesis. The development of new
drugs that specifically target these autoantibodies is an appealing and novel
approach. Herein, we briefly review the autoimmune condition of RA,
characterized by the presence of ACPA, and we describe how the neutralization of
autoantibodies might become a novel pharmacological principle. OBJECTIVE: Anticitrullinated protein/peptide antibodies (ACPA) are implicated in
rheumatoid arthritis (RA) pathogenesis and linked to the shared epitope (SE).
Citrulline modification is very similar to a different modified amino acid,
homocitrulline. We investigated antihomocitrullinated protein/ peptide antibody
(AHCPA) specificity for RA, whether ACPA were also able to bind
homocitrullinated targets, and whether the SE could accommodate
homocitrullinated peptide.
METHODS: Homocitrullinated fibrinogen was used to screen sera from patients with
RA, psoriatic arthritis, and systemic lupus erythematosus, and healthy subjects
for AHCPA using ELISA. Homocitrullination sites on fibrinogen were identified by
mass spectrometry. ACPA were affinity-purified using a synthetic citrullinated
peptide and tested for binding to homocitrullinated protein/peptide. Inhibition
of antihomocitrullinated fibrinogen antibody binding was examined.
Homocitrullinated peptide interaction with the SE was studied using computer
modeling.
RESULTS: IgG antihomocitrullinated fibrinogen antibodies were found specifically
in 49% of patients with RA. Enrichment of ACPA by affinity purification from 5
patients with RA also enriched AHCPA. Serum AHCPA was inhibited by citrullinated
fibrinogen and more significantly by homocitrullinated fibrinogen. Computer
modeling indicated that the SE could accommodate a homocitrullinated peptide
without steric hindrance. Mass spectrometry identified that 89/103 lysines of
fibrinogen could be homocitrullinated, and 5 peptides that could be both
citrullinated and homocitrullinated and are predicted to bind the SE.
CONCLUSION: Antihomocitrullinated fibrinogen antibodies are specific to RA. The
presence of AHCPA coincides with ACPA, and AHCPA copurifies with ACPA in
affinity purification and is inhibited by citrullinated and homocitrullinated
antigens. Thus AHCPA and ACPA are frequently cross-reactive and
homocitrullinated proteins/peptides may bind the SE. Anti-citrullinated protein/peptide antibodies (ACPAs) are detected in rheumatoid
arthritis (RA) sera and because of their strict association with the disease are
considered marker antibodies, probably endowed with pathogenic potential.
Antibody affinity is one of the parameters affecting pathogenicity. Three
diagnostic citrullinated peptides-viral citrullinated peptide 1 (VCP1) and VCP2
derived from Epstein-Barr virus (EBV)-encoded proteins and histone citrullinated
peptide 1 (HCP1) derived from histone H4-were synthesized as tetrameric multiple
antigen peptides and immobilized on sensor chips CM5 type in a Biacore T100
instrument. Specific binding of purified antibodies from RA patients to the
three peptides was analyzed by surface plasmon resoce using two
arginine-containing sequences as controls. Employing a 1:1 binding model for
affinity constant calculation, ACPAs interacted with VCP1 and VCP2 with lower
apparent affinity (10(-6) M>KD>10(-7) M) and interacted with HCP1 with higher
apparent affinity (KD=10(-8) M). The results indicate that the binding to
citrullinated peptides is characterized by wide differences in affinity, with
slower association and faster dissociation rates in the case of antibodies to
viral citrullinated peptides as compared with antibodies specific for the
histone peptide. This biosensor analysis shows the high cross-reactivity of
purified ACPAs that bind other citrullinated peptides besides the one used for
purification. Anticitrullinated peptide/protein antibodies (ACPA), which are highly specific
for rheumatoid arthritis (RA), may be found in some patients with other systemic
autoimmune diseases. The clinical significance of ACPA in patients with
antisynthetase syndrome (ASS), a systemic disease characterized by the
association of myositis, interstitial lung disease, polyarthralgia, and/or
polyarthritis, has not yet been evaluated with regard to phenotype, prognosis,
and response to treatment. ACPA-positive ASS patients were first identified
among a French multicenter registry of patients with ASS. Additionally, all
French rheumatology and internal medicine practitioners registered on the Club
Rhumatismes et Inflammation web site were asked to report their observations of
ASS patients with ACPA. The 17 collected patients were retrospectively studied
using a standardized questionnaire and compared with 34 unselected ACPA-negative
ASS patients in a case-control study. All ACPA-positive ASS patients suffered
from arthritis versus 41% in the control group (P < 0.0001). The number of
swollen joints was significantly higher (7.0 ± 5.0 vs 2.9 ± 3.9, P < 0.005),
with a distribution resembling that of RA. Radiographic damages were also more
frequent in ACPA-positive ASS patients (87% vs 11%, P < 0.0001). Aside from a
significantly higher transfer factor for carbon monoxide in ACPA-ASS patients,
lung, muscle, and skin involvements had similar incidences, patterns, and
severity in both groups. Although Nonbiologic treatments were similarly used in
both groups, ACPA-positive patients received biologics more frequently (59% vs
12%, P < 0.0008), mostly due to refractory arthritis (n = 9). Eight patients
received anti-Cluster of differentiation 20 (CD20) monoclonal antibodies (mAbs)
with good efficacy and tolerance, whereas 2 of the 5 patients treated with
antitumor necrosis factor drugs had worsened myositis and/or interstitial lung
disease. After a >7-year mean follow-up, extra-articular outcomes and survival
were not different. ACPA-positive ASS patients showed an overlapping RA-ASS
syndrome, were at high risk of refractory erosive arthritis, and might
experience ASS flare when treated with antitumor necrosis factor drugs. In
contrast, other biologics such as anti-CD20 mAb were effective in this context,
without worsening systemic involvements. INTRODUCTION: Patients with clinically suspect arthralgia (CSA) have, according
to their rheumatologists, an increased risk of rheumatoid arthritis (RA), but
their actual outcome is unexplored. This longitudinal study investigated (1)
progression from CSA to clinically detectable arthritis and (2) associations of
clinical factors, serological factors (among which are anticitrullinated peptide
antibodies (ACPAs)) and MRI-detected subclinical inflammation with arthritis
development.
METHODS: 150 patients with CSA were followed for ≥6 months. At baseline,
clinical and serological data were collected and unilateral 1.5 T-MRI of
metacarpophalangeal (MCP), wrist and metatarsophalangeal (MTP) joints was made.
MRI scoring was done according to the RA MRI scoring system. Subclinical MRI
inflammation was defined based on MRI results of 193 symptom-free persons.
RESULTS: During follow-up (median=75 weeks, IQR=41-106 weeks), 30 patients
developed clinical arthritis; 87% did so <20 weeks after inclusion. In
multivariable analyses, age, localisation of initial symptoms in small and large
joints (compared with small joints only), C-reactive protein level,
ACPA-positivity and subclinical MRI inflammation significantly associated with
arthritis development; ACPA and MRI inflammation were most strongly associated
(HR (95% CI) respectively, 6.43 (2.57 to 16.05) and 5.07 (1.77 to 14.50)). After
1-year follow-up, 31% of the patients with MRI inflammation and 71% of the
ACPA-positive patients with MRI inflammation had progressed to arthritis.
Forty-three per cent of the patients that developed arthritis within 1 year were
ACPA-negative; 78% of them had subclinical MRI inflammation at baseline. When
MRI inflammation was absent arthritis development was infrequent (6% in all
patients with CSA and 3% in ACPA-negative patients with CSA).
CONCLUSIONS: Subclinical MRI inflammation precedes clinical arthritis with a few
months. Subclinical MRI inflammation is, independent of other factors such as
ACPA, associated with arthritis development. Rheumatoid arthritis (RA) is an autoimmune connective tissue disease, associated
with the presence of anti-citrullinated protein antibodies (ACPA). These
antibodies have been found in approximately 70% of patients suffering from RA
and they are currently used for diagnosis of RA. Although they exhibit an
absolute need for citrulline for antibody reactivity, no precise cognate antigen
for these antibodies has been determined. In this study, we analyzed the
reactivity of ACPA to various citrullinated peptides by modified enzyme-linked
immunosorbent assays, in order to determine the dependency of specific amino
acids for antibody reactivity. A non-human protein (ovalbumin) and antigens
directly related to RA were used as templates for synthesis of non-modified and
citrullinated peptides, becoming potential target epitopes. Mainly peptides
containing a Cit-Gly motif were recognized by ACPAs, while no particular amino
acids N-terminal of citrulline were found to be essential for antibody
reactivity. Moreover, ACPA reactivity was not restricted to antigens known to be
associated with ACPA-positive RA alone, but also to proteins without relation to
RA, primarily illustrating that any protein in theory can be turned into an RA
autoantigen, by introducing Cit-Gly motifs. Knowledge about the interaction
between ACPAs and their citrullinated targets is important for understanding
autoimmune ACPA responses in RA, which are known to contribute to the
pathophysiology. OBJECTIVE: Citrullinated proteins have been found within atherosclerotic plaque.
However, studies evaluating the association between anti-citrullinated protein
antibodies (ACPAs) and imaging measures of atherosclerosis in patients with
rheumatoid arthritis (RA) have been limited to seroreactive citrullinated
fibrinogen or citrullinated vimentin and have rendered contradictory results.
Therefore, our objective was to evaluate this association using an extended
panel of ACPAs in a larger sample of RA patients without clinical cardiovascular
disease (CVD).
METHODS: ACPAs were identified using a custom Bio-Plex bead assay in 270
patients from 2 independent RA cohorts without clinical CVD, with the first one
consisting of 195 patients and the other of 75 patients. Coronary artery calcium
(CAC) was assessed by computed tomography as a measure of coronary artery
disease.
RESULTS: High levels of anti-citrullinated histone H2B antibodies were strongly
associated with higher CAC scores, compared with lower antibody levels
(P = 0.001); this remained significant after adjustment for traditional CV and
RA-specific risk factors (P = 0.03). No association between levels of ACPAs and
CAC progression at 3 years was seen (P = 0.09); however, the number of
progressors was small (n = 92).
CONCLUSION: Higher levels of ACPAs targeting Cit-histone H2B were associated
with higher CAC scores when compared to lower antibody levels, suggesting a
potential role for histone citrullination seroreactivity in atherosclerosis. BACKGROUND: Anticitrullinated protein antibodies (ACPAs) are serological
biomarkers associated with early, rapidly progressing rheumatoid arthritis (RA),
including more severe disease and joint damage. ACPA testing has become a
routine tool for RA diagnosis and prognosis. Furthermore, treatment efficacy has
been shown to vary by ACPA-positive status. However, it is not clear if the
economic burden of patients with RA varies by ACPA status.
OBJECTIVE: To determine if the economic burden of RA varies by patient ACPA
status.
METHODS: IMS PharMetrics Plus health insurance claims and electronic medical
record (EMR) data from 2010-2015 were used to identify patients with incident
RA. Patients were aged ≥ 18 years, had ≥ 1 inpatient or ≥ 2 outpatient claims
reporting an RA diagnosis code (ICD-9-CM code 714.0), and had an anticyclic
citrullinated peptide (anti-CCP; a surrogate of ACPA) antibody test within 6
months of diagnosis. Incident patients were defined as those who had no claims
with an RA diagnosis code in the 6 months before the first observed RA
diagnosis. The primary outcome of interest was RA-related medical expenditures,
defined as the sum of payer- and patient-paid amounts for all claims with an RA
diagnosis code. Secondary outcomes included health care utilization metrics such
as treatment with a disease-modifying antirheumatic drug (DMARD) and physician
visits. Generalized linear regression models were used for each outcome,
controlling for ACPA-positive status (defined as anti-CCP ≥ 20 AU/mL), age, sex,
and Charlson Comorbidity Index score as explanatory variables.
RESULTS: Of 647,171 patients diagnosed with RA, 89,296 were incident cases, and
47% (n = 42,285) had an anti-CCP test. After restricting this sample to patients
with a linked EMR and reported anti-CCP test result, 859 remained, with 24.7% (n
= 212) being ACPA-positive. Compared with ACPA-negative patients, adjusted
results showed that ACPA-positive patients were more likely to use either
conventional (71.2% vs. 49.6%; P < 0.001) or biologic (20.3% vs. 11.8%; P <
0.001) DMARDs during the first year after diagnosis and had more physician
visits (5.58 vs. 3.91 times per year; P < 0.001). Annual RA-associated total
expenditures were $7,941 for ACPA-positive and $5,243 for ACPA-negative patients
(Δ = $2,698; P = 0.002). RA-associated medical expenditures were $4,380 for
ACPA-positive and $3,427 for ACPA-negative patients (Δ = $954; P = 0.168),
whereas DMARD expenditures were $3,560 and $1,817, respectively (Δ = $1,743; P =
0.001).
CONCLUSIONS: RA-related economic burden is higher for patients who are
ACPA-positive compared with those who are ACPA-negative. Providers may wish to
inform patients diagnosed with ACPA-positive RA about the likely future disease
and economic burden in hopes that both stakeholders can be more proactive in
addressing them.
DISCLOSURES: Funding for this research was contributed by Bristol-Myers Squibb.
Patel and Price are employees and stockholders of Bristol-Myers Squibb. Shafrin
and Tebeka are employees of Precision Health Economics, a health care consulting
firm that received funding from Bristol-Myers Squibb to conduct this study.
Michaud has received a grant from Pfizer and is employed by the National Data
Bank for Rheumatic Diseases, which has received funds from Amgen, Bristol-Myers
Squibb, Eli Lilly, Janssen, Pfizer, and Regeneron. Study concept and design were
contributed by Shafrin, Price, Patel, and Michaud. Shafrin, Price, and Patel
collected the data, and all authors contributed equally to data analysis. The
manuscript was written by Shafrin and Tebeka and revised by Shafrin, Price,
Patel, and Michaud. BACKGROUND: Anti-citrullinated peptides antibodies (ACPA) are specific for
rheumatoid arthritis and have been implicated in disease pathogenesis. ACPA
examination is a new component of ACR/EULAR 2010 classification criteria for
rheumatoid arthritis. ACPA positivity predicts a more erosive disease course
with severe joint damage and extra-articular manifestations.
OBJECTIVES: To evaluate the benefits of ACPA examination in patients with early
undifferentiated arthritis and patients with rheumatoid arthritis.
METHODS: We examined patients with arthritis and tested them for ACPA
positivity. In every individual patient we evaluated if ACPA examination was
necessary to establish the diagnosis of rheumatoid arthritis, or to change
treatment, or if the diagnosis could have been established without ACPA
examination (ACR/EULAR 2010 classification criteria was met without ACPA
scoring).
RESULTS AND CONCLUSIONS: The study was placed in Slovak Republic. We examined
833 patients with arthritis. There were 43 patients, or 62% of a subgroup of 69
who were ACPA positive whose ACPA examination was not needed-ACR/EULAR criteria
was met without ACPA scoring. This number represents 5.1% of the total number
examined. There were 15 patients, or 22% of the subgroup and 1.8% of the total
whose diagnosis was revised to rheumatoid arthritis due to ACPA
positivity-ACR/EULAR criteria were met solely with ACPA scoring. There were 11
patients (16% and 1.3%) whose medication was changed due to ACPA positivity.
ACPA examination is useful in 3.1% of all examined patients. When we correlate
data on ACPA positive patients, 38% of the patients profit from ACPA
examinations. Considering the relatively low price of ACPA testing, this
examination should not be excluded. |
What is the function of emergency granulopoiesis? | ARIH2 encodes TRIAD1, an E3 ubiquitin ligase required for termination of emergency granulopoiesis and leukemia suppressor function in AML . The Interferon Consensus Sequence Binding Protein (Icsbp/Irf8) Is Required for Termination of Emergency GranulopOiesis . Emergency granulopsis is the enhanced production of neutrophils by hematopoietic stem and progenitor cells (HSPCs) upon infection . | Infections and inflammation trigger neutrophilias that are supported by a
hematopoietic program of accelerated granulopoiesis known as emergency
granulopoiesis. The intrinsic factors that drive reactive neutrophilias and
emergency granulopoiesis have been inferred but not demonstrated. Here, we show
that alum cannot elicit reactive neutrophilias in IL-1R type I (IL-1RI)(-/-)
mice, whereas other inflammatory responses, including eosinophilia and Ab
production, remain intact. Analysis of this specific impairment revealed an
uticipated role for IL-1RI in supporting increased proliferation by
granulocyte/macrophage progenitors and, surprisingly, multipotent progenitors
and hematopoietic stem cells (HSC). Indeed, HSC and multipotent progenitor
proliferative responses were most suppressed in IL-1RI(-/-) mice, suggesting a
critical role for their proliferation in inflammatory granulopoiesis. Whereas
IL-1 drives increased HSC proliferation directly in vitro, IL-1RI expression by
radiation-resistant host cells was both necessary and sufficient for
alum-induced HSC, multipotent progenitor, and granulocyte/macrophage progenitor
proliferation and reactive neutrophilias in radiation chimeric mice. Thus, IL-1
plays a necessary, but indirect, role in the support of alum-induced
neutrophilias by expanding both pluripotent and myeloid progenitor compartments
to accelerate granulopoiesis. Granulocyte colony-stimulating factor (G-CSF) mediates "emergency"
granulopoiesis during infection, a process that is mimicked by clinical G-CSF
use, yet we understand little about the intracellular signaling cascades that
control demand-driven neutrophil production. Using a murine model with
conditional deletion of signal transducer and activator of transcription 3
(STAT3) in bone marrow, we investigated the cellular and molecular mechanisms of
STAT3 function in the emergency granulopoiesis response to G-CSF administration
or infection with Listeria monocytogenes, a pathogen that is restrained by G-CSF
signaling in vivo. Our results show that STAT3 deficiency renders hematopoietic
progenitor cells and myeloid precursors refractory to the growth-promoting
functions of G-CSF or L monocytogenes infection. STAT3 is necessary for
accelerating granulocyte cell-cycle progression and maturation in response to
G-CSF. STAT3 directly controls G-CSF-dependent expression of
CCAAT-enhancer-binding protein β (C/EBPβ), a crucial factor in the emergency
granulopoiesis response. Moreover, STAT3 and C/EBPβ coregulate c-Myc through
interactions with the c-myc promoter that control the duration of C/EBPα
occupancy during demand-driven granulopoiesis. These results place STAT3 as an
essential mediator of emergency granulopoiesis by its regulation of
transcription factors that direct G-CSF-responsive myeloid progenitor expansion. The mechanisms by which innate immune signals regulate alloimmune responses
remain poorly understood. In the present study, we show by intravital 2-photon
microscopy direct interactions between graft-infiltrating neutrophils and donor
CD11c(+) dendritic cells (DCs) within orthotopic lung allografts immediately
after reperfusion. Neutrophils isolated from the airways of lung transplantation
recipients stimulate donor DCs in a contact-dependent fashion to augment their
production of IL-12 and expand alloantigen-specific IFN-γ(+) T cells. DC IL-12
expression is largely regulated by degranulation and induced by TNF-α associated
with the neutrophil plasma membrane. Extended cold ischemic graft storage
enhances G-CSF-mediated granulopoiesis and neutrophil graft infiltration,
resulting in exacerbation of ischemia-reperfusion injury after lung
transplantation. Ischemia reperfusion injury prevents immunosuppression-mediated
acceptance of mouse lung allografts unless G-CSF-mediated granulopoiesis is
inhibited. Our findings identify granulopoiesis-mediated augmentation of
alloimmunity as a novel link between innate and adaptive immune responses after
organ transplantation. Granulopoiesis is tightly regulated to meet host demands during both
"steady-state" and "emergency" situations, such as infections. The transcription
factor CCAAT/enhancer binding protein β (C/EBPβ) plays critical roles in
emergency granulopoiesis, but the precise developmental stages in which C/EBPβ
is required are unknown. In this study, a novel flow cytometric method was
developed that successfully dissected mouse bone marrow cells undergoing
granulopoiesis into five distinct subpopulations (#1-5) according to their
levels of c-Kit and Ly-6G expression. After the induction of candidemia, rapid
mobilization of mature granulocytes and an increase in early granulocyte
precursors accompanied by cell cycle acceleration was followed by a gradual
increase in granulocytes originating from the immature populations. Upon
infection, C/EBPβ was upregulated at the protein level in all the granulopoietic
subpopulations. The rapid increase in immature subpopulations #1 and #2 observed
in C/EBPβ knockout mice at 1 d postinfection was attenuated. Candidemia-induced
cell cycle acceleration and proliferation of hematopoietic stem/progenitors were
also impaired. Taken together, these data suggest that C/EBPβ is involved in the
efficient amplification of early granulocyte precursors during
candidemia-induced emergency granulopoiesis. Emergency granulopoiesis is a component of the innate immune response that is
induced in response to infectious or inflammatory challenge. It is characterized
by the rapid expansion and differentiation of granulocyte/monocyte progenitor
(GMP) populations, which is due in part to a shortened S-phase of the cell
cycle. We found that IRF8 (also known as ICSBP), an interferon regulatory
transcription factor that activates phagocyte effector genes during the innate
immune response, activates the gene encoding Fanconi C (Fancc) in murine myeloid
progenitor cells. Moreover, IRF8-induced Fancc transcription was augmented by
treatment with IL-1β, an essential cytokine for emergency granulopoiesis. The
Fanconi pathway participates in repair of stalled or collapsed replication forks
during DNA replication, leading us to hypothesize that the Fanconi pathway
contributes to genomic stability during emergency granulopoiesis. In support of
this hypothesis, Fancc(-/-) mice developed anemia and neutropenia during
repeated, failed episodes of emergency granulopoiesis. Failed emergency
granulopoiesis in Fancc(-/-) mice was associated with excess apoptosis of HSCs
and progenitor cells in the bone marrow and impaired HSC function. These studies
have implications for understanding the pathogenesis of bone marrow failure in
Fanconi anemia and suggest possible therapeutic approaches. Systemic bacterial infection induces a hematopoietic response program termed
"emergency granulopoiesis" that is characterized by increased de novo bone
marrow (BM) neutrophil production. How loss of local immune control and
bacterial dissemination is sensed and subsequently translated into the switch
from steady-state to emergency granulopoiesis is, however, unknown. Using
tissue-specific myeloid differentiation primary response gene 88
(Myd88)-deficient mice and in vivo lipopolysaccharide (LPS) administration to
model severe bacterial infection, we here show that endothelial cells (ECs) but
not hematopoietic cells, hepatocytes, pericytes, or BM stromal cells, are
essential cells for this process. Indeed, ECs from multiple tissues including BM
express high levels of Tlr4 and Myd88 and are the primary source of granulocyte
colony-stimulating factor (G-CSF), the key granulopoietic cytokine, after LPS
challenge or infection with Escherichia coli. EC-intrinsic MYD88 signaling and
subsequent G-CSF production by ECs is required for myeloid progenitor lineage
skewing toward granulocyte-macrophage progenitors, increased colony-forming unit
granulocyte activity in BM, and accelerated BM neutrophil generation after LPS
stimulation. Thus, ECs catalyze the detection of systemic infection into
demand-adapted granulopoiesis. Expression of the E3 ubiquitin ligase Triad1 is greater in mature granulocytes
than in myeloid progenitor cells. HoxA10 actives transcription of the gene
encoding Triad1 (ARIH2) during myeloid differentiation, but the contribution of
increased Triad1 expression to granulocyte production or function is unknown.
Mice with bone marrow-specific disruption of the ARIH2 gene exhibit constitutive
inflammation with tissue infiltration by granulocytes and B cells. In contrast,
disruption of the HOXA10 gene in mice neither constitutively activates the
innate immune response nor significantly alters steady-state granulopoiesis.
This study explores the impact of HoxA10-induced Triad1 expression on emergency
(stress) granulopoiesis. We found that mice with HOXA10 gene disruption
exhibited an overwhelming and fatal emergency granulopoiesis response that was
characterized by tissue infiltration with granulocytes, but reversed by
re-expression of Triad1 in the bone marrow. We determined that HoxA9 repressed
ARIH2 transcription in myeloid progenitor cells, antagonizing the effect of
HoxA10 on Triad1 expression. Also, we found that differentiation-stage-specific
ARIH2 transcription was regulated by the tyrosine phosphorylation states of
HoxA9 and HoxA10. Our studies demonstrate a previously undescribed role for
HoxA10 in terminating emergency granulopoiesis, suggesting an important
contribution by Hox proteins to the innate immune response. Emergency granulopoiesis refers to the increased production of neutrophils in
bone marrow and their release into circulation induced by severe infection.
Several studies point to a critical role for G-CSF as the main mediator of
emergency granulopoiesis. However, the consequences of G-CSF stimulation on the
transcriptome of neutrophils and their precursors have not yet been investigated
in humans. In this work, we examine the changes in mRNA expression induced by
administration of G-CSF in vivo, as a model of emergency granulopoiesis in
humans. Blood samples were collected from healthy individuals after 5 d of G-CSF
administration. Neutrophil precursors were sorted into discrete stages of
maturation by flow cytometry, and RNA was subjected to microarray analysis. mRNA
levels were compared with previously published expression levels in
corresponding populations of neutrophil precursors isolated from bone marrow of
untreated, healthy individuals. One thousand one hundred and ten mRNAs were
differentially expressed >2-fold throughout terminal granulopoiesis. Major
changes were seen in pathways involved in apoptosis, cytokine signaling, and TLR
pathways. In addition, G-CSF treatment reduced the levels of four of five
measured granule proteins in mature neutrophils, including the proantibacterial
protein hCAP-18, which was completely deficient in neutrophils from
G-CSF-treated donors. These results indicate that multiple biological processes
are altered to satisfy the increased demand for neutrophils during G-CSF-induced
emergency granulopoiesis in humans. Interferon consensus sequence-binding protein (Icsbp) is required for
terminating emergency granulopoiesis, an episodic event responsible for
granulocyte production in response to infections and a key component of the
innate immune response. Icsbp inhibits the expression of Stat3 and C/ebpβ,
transcription factors essential for initiating and sustaining granulopoiesis,
and activates transcription of Fanconi C (FANCC), a DNA repair protein. In prior
studies, we noted accelerated bone marrow failure in Fancc-/- mice undergoing
multiple episodes of emergency granulopoiesis, associated with apoptosis of bone
marrow cells with unrepaired DNA damage. Additionally, we found increased
expression of Fanconi C and F proteins during emergency granulopoiesis. These
findings suggest that Icsbp protects the bone marrow from DNA damage by
increasing activity of the Fanconi DNA repair pathway, but the mechanisms for
FANCC activation during initiation of emergency granulopoiesis are unclear. In
this study, we observed that Stat3 and C/ebpβ activate FANCC transcription and
contribute to DNA repair. Our findings indicate that FancC expression is
increased during Stat3- and C/ebpβ-induced initiation of emergency
granulopoiesis by these transcription factors and is maintained through
termination by Icsbp. Our work reveals that Stat3- and C/ebpβ-mediated FancC
expression is a critical component for initiating and sustaining key innate
immune responses. Emergency granulopoiesis is a hematopoietic program of stem cell-driven
neutrophil production used to counteract immune cell exhaustion following
infection. Shigella flexneri is a Gram-negative enteroinvasive pathogen
controlled by neutrophils. In this study, we use a Shigella-zebrafish (Danio
rerio) infection model to investigate emergency granulopoiesis in vivo We show
that stem cell-driven neutrophil production occurs in response to Shigella
infection and requires macrophage-independent signaling by granulocyte
colony-stimulating factor (Gcsf). To test whether emergency granulopoiesis can
function beyond homoeostasis to enhance innate immunity, we developed a
reinfection assay using zebrafish larvae that have not yet developed an adaptive
immune system. Strikingly, larvae primed with a sublethal dose of Shigella are
protected against a secondary lethal dose of Shigella in a type III secretion
system (T3SS)-dependent manner. Collectively, these results highlight a new role
for emergency granulopoiesis in boosting host defense and demonstrate that
zebrafish larvae can be a valuable in vivo model to investigate innate immune
memory.IMPORTANCEShigella is an important human pathogen of the gut. Emergency
granulopoiesis is the enhanced production of neutrophils by hematopoietic stem
and progenitor cells (HSPCs) upon infection and is widely considered a
homoeostatic mechanism for replacing exhausted leukocytes. In this study, we
developed a Shigella-zebrafish infection model to investigate stem cell-driven
emergency granulopoiesis. We discovered that zebrafish initiate granulopoiesis
in response to Shigella infection, via macrophage-independent signaling of
granulocyte colony-stimulating factor (Gcsf). Strikingly, larvae primed with a
sublethal dose of Shigella are protected against a secondary lethal dose of
Shigella in a type III secretion system (T3SS)-dependent manner. Taken together,
we show that zebrafish infection can be used to capture Shigella-mediated stem
cell-driven granulopoiesis and provide a new model system to study stem cell
biology in vivo Our results also highlight the potential of manipulating stem
cell-driven granulopoiesis to boost innate immunity and combat infectious
disease. Under stress conditions such as infection, inflammation, and hematopoietic
recovery following chemotherapy or transplantation, the hematopoietic system is
required to meet the increasing demands, especially from myeloid cells.
Therefore, an understanding of the molecular mechanism underlying stress
hematopoiesis is clinically imperative. We previously showed that C/EBPβ, which
is a transcription factor required for emergency granulopoiesis, plays a pivotal
role at the level of hematopoietic stem/progenitor cells under stress
conditions. Upon exposure to stress, the C/EBPβ protein is upregulated in the
hematopoietic stem cells. A close examination of C/EBPβ knockout mice revealed
that C/EBPβ regulates the proliferation and differentiation of hematopoietic
stem cells at the cost of the self-renewing activity. Further elucidation of the
functions and regulation of C/EBPβ in hematopoietic stem cells will facilitate
an understanding of stress hematopoiesis. Emergency granulopoiesis is a very important strategy to supply efficient
neutrophil number in response to infection. However, molecular mechanism
involved in this process remains unclear. Here, we found that administration of
WKYMVm, an immune modulating peptide, to septic mice strongly increased
neutrophil number through augmented emergency granulopoiesis. WKYMVm-induced
emergency granulopoiesis was blocked not only by a formyl peptide receptor 2
(FPR2) antagonist (WRW4), but also by FPR2 deficiency. As progenitors of
neutrophils, Lin-c-kit＋Sca-1- cells expressed FPR2. WKYMVm-induced emergency
granulopoiesis was also blocked by a phospholipase C inhibitor (U-73122). These
results suggest that WKYMVm can stimulate emergency granulopoiesis via FPR2 and
phospholipase C enzymatic activity. [BMB Reports 2018; 51(8): 418-423]. |
What was the predominant rotavirus genotype in the pre-vaccine era, in Australia? | G1P[8] was the dominant genotype in Australia in the prevaccine era (1995-2006). | BACKGROUND: Introduction of rotavirus vaccines into national immunization
programs (NIPs) could result in strain selection due to vaccine-induced
selective pressure. This study describes the distribution and diversity of
rotavirus genotypes before and after rotavirus vaccine introduction into the
Australian NIP. State-based vaccine selection facilitated a unique comparison of
diversity in RotaTeq and Rotarix vaccine states.
METHODS: From 1995 to 2015, the Australian Rotavirus Surveillance Program
conducted genotypic analysis on 13051 rotavirus-positive samples from children
<5 years of age, hospitalized with acute gastroenteritis. Rotavirus G and P
genotypes were determined using serological and heminested multiplex
reverse-transcription polymerase chain reaction assays.
RESULTS: G1P[8] was the domit genotype nationally in the prevaccine era
(1995-2006). Following vaccine introduction (2007-2015), greater genotype
diversity was observed with fluctuating genotype domice. Genotype
distribution varied based on the vaccine implemented, with G12P[8] domit in
states using RotaTeq, and equine-like G3P[8] and G2P[4] domit in states and
territories using Rotarix.
CONCLUSIONS: The increased diversity and differences in genotype domice
observed in states using RotaTeq (G12P[8]), and in states and territories using
Rotarix (equine-like G3P[8] and G2P[4]), suggest that these vaccines exert
different immunological pressures that influence the diversity of rotavirus
strains circulating in Australia. IntroductionRotavirus vaccination with the live-attenuated monovalent (a G1P[8]
human rotavirus strain) two-dose Rotarix vaccine was introduced in England in
July 2013. Since then, there have been significant reductions in rotavirus
gastroenteritis incidence.AimWe assessed the vaccine's impact on rotavirus
genotype distribution and diversity 3 years post-vaccine
introduction.MethodsEpidemiological and microbiological data on genotyped
rotavirus-positive samples between September 2006 and August 2016 were supplied
by EuroRotaNet and Public Health England. Multinomial multivariable logistic
regression adjusting for year, season and age was used to quantify changes in
genotype prevalence in the vaccine period. Genotype diversity was measured using
the Shannon's index (H') and Simpson's index of diversity (D).ResultsWe analysed
genotypes from 8,044 faecal samples. In the pre-vaccine era, G1P[8] was most
prevalent, ranging from 39% (411/1,057) to 74% (527/709) per year. In the
vaccine era, G1P[8] prevalence declined each season (35%, 231/654; 12%,
154/1,257; 5%, 34/726) and genotype diversity increased significantly in 6-59
months old children (H' p < 0.001: D p < 0.001). In multinomial analysis, G2P[4]
(adjusted multinomial odds ratio (aMOR): 9.51; 95% confidence interval (CI):
7.02-12.90), G3P[8] (aMOR: 2.83; 95% CI: 2.17-3.81), G12P[8] (aMOR: 2.46; 95%
CI: 1.62-3.73) and G4P[8] (aMOR: 1.42; 95% CI: 1.02-1.96) significantly
increased relative to G1P[8].ConclusionsIn the context of reduced rotavirus
disease incidence, genotype diversity has increased, with a relative change in
the domit genotype from G1P[8] to G2P[4] after vaccine introduction. These
changes will need continued surveillance as the number and age of vaccinated
birth cohorts increase in the future. BACKGROUND: Since 2006, the New Vaccine Surveillance Network has conducted
active, population-based surveillance for acute gastroenteritis (AGE)
hospitalizations and emergency department (ED) visits in 3 United States
counties. Trends in the epidemiology and disease burden of rotavirus
hospitalizations and ED visits were examined from 2006 to 2016.
METHODS: Children < 3 years of age hospitalized or visiting the ED with AGE were
enrolled from January 2006 through June 2016. Bulk stool specimens were
collected and tested for rotavirus. Rotavirus-associated hospitalization and ED
visit rates were calculated annually with 2006-2007 defined as the prevaccine
period and 2008-2016 as the postvaccine period. Rotavirus genotype trends were
compared over time.
RESULTS: Over 11 seasons, 6954 children with AGE were enrolled and submitted a
stool specimen (2187 hospitalized and 4767 in the ED). Comparing pre- and
postvaccine periods, the proportion of children with rotavirus dramatically
declined for hospitalization (49% vs 10%) and ED visits (49% vs 8%). In the
postvaccine era, a biennial pattern of rotavirus rates was observed, with a
trend toward an older median age. G1P[8] (63%) was the predomit genotype in
the prevaccine period with a significantly lower proportion (7%) in the
postvaccine period (P < .001). G2P[4] remained stable (8% to 14%) in both
periods, whereas G3P[8] and G12P[8] increased in proportion from pre- to
postvaccine periods (1% to 25% and 17% to 40%), respectively.
CONCLUSIONS: The epidemiology and disease burden of rotavirus has been altered
by rotavirus vaccination with a biennial disease pattern, sustained low rates of
rotavirus in children < 3 years of age, and a shift in the residual genotypes
from G1P[8] to other genotypes. |
Missense mutations in which genes cause X-linked developmental and epileptic encephalopathy? | GRIA3 missense mutation is cause of an x-linked developmental and epileptic encephalopathy. Missense variants in the N-terminal domain of the A isoform of FHF2/FGF13 also cause an X-linked developmental and epileptic encephalopathy. | PURPOSE: GRIA3, encoding subunit 3 of glutamate ionotropic AMPA receptor, is
associated with X-linked intellectual disability (ID), dysmorphic features, and
non-syndromic epilepsy. We aimed to characterize electro-clinical features of
patients with GRIA3 variants.
METHODS: We report a patient carrying a hemizygous missense variant c.2359 G > A
(p.Glu787Lys) inGRIA3 gene. Following a literature search, we also reviewed
clinical, electrophysiological, radiological, and genetic features of 19
patients with GRIA3 mutations.
RESULTS: This 26-month-old boy had developmental delay, early onset refractory
myoclonic epilepsy, and non-convulsive refractory status epilepticus. In
published reports, epilepsy was in 6 of 19 patients carrying different
genotypes, though epilepsy and electroencephalogram features were not completely
defined. Out of the 6 patients, one presented with generalized tonic-clonic
seizures, two with myoclonic and clonic events (one also presented with
epileptic spasms), and one with atypical absences and myoclonic jerks.
Information on type of epilepsy was unavailable for 3 cases. Epilepsy onset was
early in life and there was potential tendency for myoclonic/clonic events. The
epilepsy was difficult to treat and prognosis is poor. Severity of ID ranged
from mild to severe and was variably associated with bipolar affective disorder
and autistic spectrum disorders. Other neurological features included hypotonia,
asthenic body habitus with poor muscle bulk, and hyporeflexia.
CONCLUSION: Our report expands knowledge on the electro-clinical and molecular
spectrum of GRIA3 variants. Larger investigations will better define the
prevalence of epilepsy, the epileptic phenotype, and syndromic features
underlying GRIA3 variants. Author information:
(1)Institute of Medical Genetics, University Hospital of Wales, Cardiff CF14
4XW, UK; Division of Cancer and Genetics, School of Medicine, Cardiff
University, Cardiff CF14 4XN, UK. Electronic address: [email protected].
(2)Department of Biological Sciences, Hunter College of City University, 695
Park Avenue, New York, NY 10065, USA; Program in Biology, Graduate Center of
City University, 365 Fifth Avenue, New York, NY 10016, USA.
(3)Neurology and Molecular Neuroscience Research, Institute of Life Science,
Swansea University Medical School, Swansea University, Swansea SA2 8PP, UK.
(4)Neurology and Molecular Neuroscience Research, Institute of Life Science,
Swansea University Medical School, Swansea University, Swansea SA2 8PP, UK;
Neurology department, Morriston Hospital, Swansea Bay University Hospital Health
Board, Swansea SA6 6NL, UK.
(5)Paediatric Neurology, University Hospital of Wales, Heath Park, Cardiff CF14
4XW, UK.
(6)Division of Cancer and Genetics, School of Medicine, Cardiff University,
Cardiff CF14 4XN, UK.
(7)Institute of Medical Genetics, University Hospital of Wales, Cardiff CF14
4XW, UK.
(8)Manchester Centre for Genomic Medicine, Manchester University NHS Foundation
Trust and Institute of Human Development, University of Manchester, Manchester
M13 9WL, UK.
(9)Department of Paediatric Neurology, Royal Manchester Children's Hospital,
Oxford Road, Manchester M13 9WL, UK.
(10)Department of Neurology, Salford Royal Hospital NHS Foundation Trust, Stott
Lane, Salford M6 8HD, UK.
(11)Department of Radiology, Alder Hey Children's NHS Foundation Trust, Eaton
Road, Liverpool L12 2AP, UK.
(12)West Midlands Regional Genetics Service, Clinical Genetics Unit, Birmingham
Women's Hospital, Birmingham B15 2TG, UK.
(13)Department of Paediatrics, McMaster University, 1200 Main St. W., Hamilton,
ON L8N 3Z5, Canada.
(14)Department of Pediatrics, Peking University First Hospital, Xicheng
District, Beijing 100034, China.
(15)Cipher Gene Ltd, Beijing, China.
(16)Genomics England, London EC1M 6BQ, UK.
(17)Neurology and Molecular Neuroscience Research, Institute of Life Science,
Swansea University Medical School, Swansea University, Swansea SA2 8PP, UK;
Faculty of Medicine and Health, Camperdown, University of Sydney, NSW 2006,
Australia.
(18)Neurology and Molecular Neuroscience Research, Institute of Life Science,
Swansea University Medical School, Swansea University, Swansea SA2 8PP, UK; Kids
Neuroscience Centre, Kids Research, Children Hospital at Westmead, Sydney, NSW
2145, Australia; Brain and Mind Centre, Faculty of Medicine and Health,
University of Sydney, NSW 2050, Australia. |
Does radiotherapy for cervical cancer increases risk of colon cancer? | Yes, there is epidemiological evidence to suggest that radiotherapy for cervical cancer increases risk for colon cancer. | Incidence of second primary cancers was evaluated in 7,127 women with invasive
cancer of the cervix uteri, diagnosed between 1935 and 1978, and followed up to
38 years (average, 8.9 yr) in Connecticut. Among 5,997 women treated with
radiation, 449 developed second primary cancers compared with 313 expected
(relative risk = 1.4) on the basis of rates from the Connecticut Tumor Registry.
Excess incidence was noticeable 15 years or more after radiotherapy and
attributed mostly to cancers of sites in or near the radiation field, especially
the bladder, kidneys, rectum, corpus uteri, and ovaries. No excess was found for
these sites among the 1,130 nonirradiated women. The ratio of observed to
expected cancers for these sites did not vary appreciably by age at irradiation.
The data suggested that high-dose pelvic irradiation was associated with
increase in cancers of the bladder, kidneys, rectum, ovaries, corpus uteri, and
non-Hodgkin's lymphoma but, apparently, not leukemia, Hodgkin's disease, breast
cancer, or colon cancer. BACKGROUND: Given the extended survival of patients diagnosed with cervical
cancer, the large number of these women treated with radiotherapy, and the
presence in this population of established cancer risk factors such as human
papillomavirus (HPV) infection and cigarette smoking, it is important to clarify
long-term trends in second cancer risk.
METHODS: Using data from 104,760 one-year survivors of cervical cancer reported
to 13 population-based cancer registries in Denmark, Finland, Norway, Sweden,
and the United States, we calculated standardized incidence ratios (SIRs) for
second cancers overall and cancers at particular sites among women with cervical
cancer, including cervical cancer patients who were treated or not treated with
radiation, over more than 40 years of follow-up. Cox regression models were used
to assess the time-varying association of radiotherapy with risk of second
cancers and to assess the interaction of radiation treatment with age at
diagnosis. All statistical tests were two-sided.
RESULTS: Among 104,760 one-year survivors of cervical cancer, the risk of all
second cancers taken together was increased to a statistically significant
extent (n = 12,496; SIR = 1.30; 95% confidence interval [CI] = 1.28 to 1.33).
Compared with the general population, in both radiotherapy (N = 52,613) and
no-radiotherapy groups (N = 27,382), risks for HPV-related cancers (of the
pharynx, genital sites, and rectum/anus) and smoking-related cancers (of the
pharynx, trachea/bronchus/lung, pancreas, and urinary bladder) were elevated to
a statistically significant extent. Cervical cancer patients treated with
radiotherapy, but not those who did not receive radiotherapy, were at increased
risk for all second cancers and cancers at heavily irradiated sites (colon,
rectum/anus, urinary bladder, ovary, and genital sites) beyond 40 years of
follow-up compared with women in the general population. The association of
radiotherapy with second cancer risk was modified by age at cervical cancer
diagnosis for rectum/anus, genital sites, and urinary bladder, with higher
hazard ratios for second cancer at younger ages of cervical cancer. After
adjustment for competing mortality, the 40-year cumulative risk of any second
cancer was higher among women diagnosed with cervical cancer before age 50
(22.2%; 95% CI = 21.5% to 22.8%) than among women diagnosed after age 50 (16.4%;
95% CI = 16.1% to 16.9%).
CONCLUSION: Cervical cancer patients treated with radiotherapy are at increased
risk of second cancers at sites in close proximity to the cervix beyond 40 years
of follow-up. Because advances in therapy have increased long-term survival for women with
cervical cancer, it is important to study the risk of secondary primary
maligcies in high-dose organ areas. From the 1973-2009 National Cancer
Institute Surveillance, Epidemiology, and End Results (SEER) program, we studied
the risk of developing cancer of the colon and rectum in 64,507 cervical cancer
patients over 35 years after initial radiation treatment. We also assessed
change in risk over time. Kaplan-Meier estimator for survival curve and Cox
proportional hazards models was used. More than half (52.6%) of the cervical
cancer patients received radiation treatment. In the analyses adjusted for
race/ethnicity, age, marital status, surgery status, stage and grade, the risk
of colon cancer between those both with and without XRT diverged beginning at
approximately 8 years. After 8 years, the hazard ratio for developing colon
cancer was 2.00 (95% CI 1.43-2.80) for women with radiation versus those without
radiation treatment. The risk of rectal cancer diverged after 15 years of
follow-up (HR 4.04, 95% CI 2.08-7.86). After 35 years of follow-up, the absolute
risk of developing colon cancer was 6.5% for those who received radiation versus
2.5% for those without, and 3.7 versus 0.8% for rectum. The risk of colon and
rectum cancer over 20 years of follow-up after radiation remained the same
across three eras (1973-1980, 1981-1990, and 1991-2000). Radiation-induced
second cancers of the colon and rectum may occur 8 years after radiation
treatment for cervical cancer. |
List human antibody isotypes. | IgA
IgE
IgG
IgM
IgD | Human immunoglobulin D (IgD) occurs most abundantly as a membrane-bound antibody
on the surface of mature B cells (mIgD). IgD possesses the longest hinge
sequence of all the human antibody isotypes, with 64 residues connecting the Fab
and Fc fragments. A novel rapid purification scheme of secreted IgD from the
serum of an IgD myeloma patient using thiophilic (T-gel) and lectin affinity
chromatography gave a stable, homogeneous IgD preparation. Synchrotron X-ray
scattering and analytical ultracentrifugation of IgD identified the solution
arrangement of its Fab and Fc fragments, and thereby its hinge structure. The
Guinier X-ray radius of gyration R(G) of 6.9(+/-0.1)nm showed that IgD is more
extended in solution than the immunoglobulin subclass IgA1 (R(G) of 6.1-6.2nm).
Its distance distribution function P(r) showed a single peak at 4.7nm and a
maximum dimension of 23nm. Velocity experiments gave a sedimentation coefficient
of 6.3S, which is similar to that for IgA1 at 6.2S. The complete IgD structure
was modelled using molecular dynamics to generate IgD hinge structures, to which
homology models for the Fab and Fc fragments were connected. Good scattering
curve fits were obtained with 18 semi-extended best fit IgD models that were
filtered from 8500 trial models. These best-fit models showed that the IgD hinge
does not correspond to an extended polypeptide structure. The averaged solution
structure arrangement of the Fab and Fc fragments in IgD is principally T-shaped
and flexible, with contribution from Y-shaped and inverted Y-shaped structures.
Although the linear sequence of the IgD hinge is much longer, comparison with
previous scattering modelling of IgA1 and IgA2(m)1 suggests that the hinge of
IgA1 and IgD are more similar than might have been expected, Both possess
flexible T-shaped solution structures, probably reflecting the presence of
restraining O-linked sugars. B cells have recently entered the stage as an important accessory player in type
1 diabetes (T1D) etiopathogenesis. Experimental studies suggest regulatory
functions of vitamin D on B cells. However, only a few human studies, with
considerable methodological limitations, have been conducted within this field.
Our objective was to investigate whether higher 25-hydroxyvitamin D (25(OH)D)
concentrations were inversely associated with β-cell autoantigens glutamic acid
decarboxylase (isoform 65) (GADA) and insulinoma-associated antigen-2A (IA-2A).
Further, we also wanted to examine the relationship between 25(OH)D and total
antibody concentrations. We randomly selected 500 patients with newly diagnosed
T1D and 500 siblings for 25(OH)D, antibody and genetic analysis from the
population-based Danish Registry of Childhood and Adolescent Diabetes. The
relative change (RC) in the mean concentration of GADA, IA-2A and antibody
isotypes by a 10 nmol/l increase in 25(OH)D concentration was modelled by a
robust log-normal regression model. We found no association between 25(OH)D and
GADA [adjusted RC per 10 nmol/l increase: 1.00; 95% confidence interval (CI):
0.98-1.02] and IA-2A [adjusted RC per 10 nmol/l increase: 0.92; CI: 0.76-1.12].
Further, 25(OH)D was not associated with the total concentration of antibody
isotypes [immunoglobulin (Ig)A, IgE, IgG and IgM]. All null findings were
unaltered after adjustment for genetic variation in the vitamin D pathway.
Physiological concentrations of 25(OH)D are unlikely to have a clinically
important effect on antibody concentrations in a paediatric population of newly
diagnosed patients with T1D and their healthy siblings. Autoantibodies to breast and other cancers are commonly present in cancer
patients. A method to rapidly produce these anti-cancer autoantibodies in the
lab would be valuable for understanding immune events and to generate candidate
reagents for therapy and diagnostics. The purpose of this report is to evaluate
sentinel nodes (SNs) of breast cancer patients as a source of anti-cancer
antibodies. Radiotracer lymphatic mapping in 29 patients with breast cancer
confirmed the identity of the SNs which provided source cells for this study.
Flow cytometry demonstrated ~28% of the MNCs were B cells and ~44% of the B
cells were class switched memory B cells. EBV-induced proliferation of B cells
yielded tumor binding antibodies from 3 wells per 1000 but cultures were too
unstable for detailed evaluations. Hybridomas generated by electrofusion
produced IgG (48%), IgM (34%) and IgA (18%) antibody isotypes which were
screened for binding to a panel of breast cancer cells of the major molecular
subtypes. Tumor lysate binding was observed in 28% of the hybridoma clones and
10% of these bound whole tumor cells with unique binding patterns. More detailed
evaluation of selected clones showed binding to the patient's own tumor. SNs are
removed from more than 100,000 breast cancer patients in the US per year.
Samples from these lymph nodes represent a substantial opportunity to generate
anticancer antibodies. Author information:
(1)Graduate School of Public Health and Center for Vaccine Research, University
of Pittsburgh, Biomedical Science Tower 3, room 9052, 3501 5th Avenue,
Pittsburgh, PA 15261, USA. Electronic address: [email protected].
(2)Sanofi Pasteur, One Discovery Drive, Swiftwater, PA, 18370, USA.
(3)Aggeu Magalhaes Institute, Oswaldo Cruz Foundation (FIOCRUZ), Av. Prof.
Moraes Rego, s/n - Cidade Universitária - Campus da UFPE, CEP: 50.740-465,
Recife, Pernambuco, Brazil.
(4)Aggeu Magalhaes Institute, Oswaldo Cruz Foundation (FIOCRUZ), Av. Prof.
Moraes Rego, s/n - Cidade Universitária - Campus da UFPE, CEP: 50.740-465,
Recife, Pernambuco, Brazil; School of Medical Science, University of Pernambuco,
Recife, Brazil.
(5)DST-NRF Centre of Excellence in Epidemiological Modelling and Analysis
(SACEMA), Stellenbosch, Western Cape, South Africa.
(6)Graduate School of Public Health and Center for Vaccine Research, University
of Pittsburgh, Biomedical Science Tower 3, room 9052, 3501 5th Avenue,
Pittsburgh, PA 15261, USA.
(7)Graduate School of Public Health and Center for Vaccine Research, University
of Pittsburgh, Biomedical Science Tower 3, room 9052, 3501 5th Avenue,
Pittsburgh, PA 15261, USA; Aggeu Magalhaes Institute, Oswaldo Cruz Foundation
(FIOCRUZ), Av. Prof. Moraes Rego, s/n - Cidade Universitária - Campus da UFPE,
CEP: 50.740-465, Recife, Pernambuco, Brazil. Electronic address:
[email protected]. |
What is CRAO in the context of the eye? | central retinal artery occlusion (CRAO) is an ophthalmological emergency, the retinal analog of a stroke. | Central retinal-artery obstruction (CRAO) is a devastating complication after
retrobulbar anesthesia, a procedure which was previously recommended routinely
to immobilize the eye and reduce discomfort during laser surgery. Recent
developments in treatment technique, involving scatter laser applications given
over several sessions with smaller spot sizes and shorter durations, have
virtually eliminated the need for retrobulbar injections and the risks of
retrobulbar hemorrhage, which can cause increased intraocular pressure and
culminate in closure of the arterial circulation. We present the case of a
patient with proliferative sickle retinopathy who sustained such a complication
after direct photocoagulation treatment of sea-fans elsewhere and offer
alternative treatment techniques which we have used for the past ten years that
eliminate this hazard. Central retinal artery occlusion (CRAO) is considered to be an acute stroke of
the eye that results in profound visual loss. Spontaneous recovery rates are
poor. Most CRAOs are caused by thromboembolism in the central retinal artery.
Current standard therapies for CRAO that aim to restore perfusion to the retina
and optic nerve head have not been shown to alter the natural course of the
disease. Thrombolytic therapy for acute management of CRAO has shown promise in
nonrandomized studies with regard to improving visual outcomes. A randomized
controlled trial will be required to confirm the efficacy of thrombolytic
therapy before it can be recommended for use in CRAO in daily clinical practice. Central retinal artery occlusion (CRAO) is an ophthalmological emergency
situation. Known risk factors are arterial hypertension, cardial arrhythmia,
arteriosclerosis, hypercholesterolemia and diabetes. Elderly patients should be
examined for an arteritic genesis. Young patients (< 45 years) without typical
risk factors may suffer from thrombophilia. There is no uniform recommendation
on how to treat non-arteritic CRAO. Many different interventions have been
suggested in the literature, i. e., massaging the eye, systemic or local
reduction of intraocular pressure, anticoagulation, either systemically
administered venous thrombolysis or supraselective intra-arterial thrombolysis.
In this review we present the causes of CRAO and diagnostic means to detect
causes; we also critically discuss previously described therapeutic options. It
is our aim to provide a guide through the necessary interdisciplinary
diagnostics in co-operation with internal medicine and neurology and to
recommend a multimodal therapy in patients with non-arteritic CRAO. Central retinal artery occlusion (CRAO) causes ischemic stroke of the eye. We
report a case of CRAO that was successfully treated with intravenous recombit
tissue plasminogen activator (rt-PA) and review the current literature. A
64-year-old right-handed man presented to the emergency department with acute
left eye amaurosis. An ophthalmologic assessment revealed a left afferent
pupillary defect, minimal visual acuity, macular edema with a cherry red spot,
and multiple emboli in the inferotemporal arcade of the left eye. A neurologic
examination was otherwise nonfocal; neuroimaging was normal. Acute CRAO was
diagnosed, and rt-PA was administered intravenously 185 minutes after symptom
onset. A repeat examination 4.5 hours after treatment found improved vision,
reduced macular edema, and no emboli. An ophthalmologic evaluation 10 days later
found a visual acuity of 20/200 in the left eye and bilateral arterial sclerosis
without evidence of retinal emboli or macular edema. This case illustrates that
intravenous rt-PA may be an effective therapeutic option for CRAO in select
patients. Given the current literature and the recommended established safety
window for thrombolytics in acute ischemic cerebral stroke, it is reasonable to
administer intravenous treatment for CRAO within 4.5 hours after symptom onset.
Nevertheless, it is critical that a prospective clinical trial confirm the
efficacy, safety, and time window for treatment. β(2)-Microglobulin (β(2) M) has been reported to be elevated in patients with a
variety of neoplasms and inflammatory disorders, and is believed to be a
sensitive although nonspecific marker for lymphocyte activation and/or
proliferation. In order to investigate the role of inflammation in the
pathogenesis of various types of cataract, the authors measured β(2)M
concentrations in the aqueous humor and serum of patients with senile cataracts
(82 eyes), cataracts secondary to uveitis (16 eyes) and cataracts associated
with atopic dermatitis (eight eyes). In addition, measurements were made in six
patients with rhegmatogenous retinal detachment (RRD) and three patients with
central retinal artery occlusion (CRAO) for comparison. The average aqueous
β(2)M was increased in eyes with uveitic cataracts (678 μg/1) and RRD (533
μg/1), when compared to eyes with senile cataracts (265 μg/1), atopic cataracts
(309 μg/1) and CRAO (122 μg/1). However, comparison of β(2)M to albumin
aqueous-to-serum ratios (protein coefficient analysis) revealed that the aqueous
β(2)M elevation was specific in only uveitic cataracts, with the elevation in
RRD being most likely due to breakdown of the bloodocular barrier. Higher
aqueous β(2)M concentrations were also found in cataracts with a posterior
subcapsular cataract component, although this was related to a higher percentage
of uveitic cataracts in this group. There was no statistically significant
difference found in association with a past medical history of diabetes
mellitus, hypertension or heart disease. These results are discussed in the
context of the pathogenesis of cataract and the role of β(2)M in inflammatory
processes of the eye. Central retinal artery occlusion (CRAO) is an ocular emergency and is the ocular
analogue of cerebral stroke. It results in profound, usually monocular vision
loss, and is associated with significant functional morbidity. The risk factors
for CRAO are the same atherosclerotic risk factors as for stroke and heart
disease. As such, individuals with CRAO may be at risk of ischemic end organ
damage such as a cerebral stroke. Therefore, the management of CRAO is not only
to restore vision, but at the same time to manage risk factors that may lead to
other vascular conditions. There are a number of therapies that has been used in
the treatment of CRAO in the past. These include carbogen inhalation,
acetazolamide infusion, ocular massage and paracentesis, as well as various
vasodilators such as intravenous glyceryl trinitrate. None of these "standard
agents" have been shown to alter the natural history of disease definitively.
There has been recent interest shown in the use of thrombolytic therapy,
delivered either intravenously or intra-arterially by direct catheterisation of
the ophthalmic artery. Whilst a number of observational series have shown that
the recovery of vision can be quite dramatic, two recent randomised controlled
trials have not demonstrated efficacy. On the contrary, intra-arterial delivery
of thrombolytic may result in an increased risk of intracranial and systemic
haemorrhage, while the intravenous use of tissue plasminogen activator (tPA) was
not shown to be efficacious within 24 h of symptom onset. Nevertheless, both of
these studies have shown one thing in common, and that is for treatment to be
effective in CRAO, it must be deployed within a short time window, probably
within 6 h of symptom onset. Therefore, while CRAO is a disease that does not
have a treatment, nevertheless it needs to follow the same principles of
treatment as any other vascular end organ ischaemic disease. That is, to attempt
to reperfuse ischemic tissue as quickly as possible and to institute secondary
prevention early. Central retinal artery occlusion (CRAO) is an ophthalmic emergency and the
ocular analogue of cerebral stroke. Best evidence reflects that over
three-quarters of patients suffer profound acute visual loss with a visual
acuity of 20/400 or worse. This results in a reduced functional capacity and
quality of life. There is also an increased risk of subsequent cerebral stroke
and ischaemic heart disease. There are no current guideline-endorsed therapies,
although the use of tissue plasminogen activator (tPA) has been investigated in
two randomized controlled trials. This review will describe the pathophysiology,
epidemiology, and clinical features of CRAO, and discuss current and future
treatments, including the use of tPA in further clinical trials. IMPORTANCE: Sickle cell disease (SCD) is characterized by vaso-occlusive crisis.
In the eye, central retinal artery occlusion (CRAO) is a rare complication in
SCD, with only 1 previous report of bilateral, concurrent CRAO. We report a case
of bilateral, concurrent CRAO in a patient with SCD, possibly precipitated by
the use of phosphodiesterase 5 inhibitors.
OBSERVATIONS: A 37-year-old African American woman with a known medical history
significant for SCD and pulmonary arterial hypertension who was receiving
treatment with tadalafil, a phosphodiesterase 5 inhibitor, developed bilateral,
concurrent CRAO that persisted after exchange transfusion.
CONCLUSIONS AND RELEVANCE: Bilateral CRAO secondary to SCD is extremely rare,
with only 1 previous case report in the literature. The use of phosphodiesterase
5 inhibitors is an additional risk factor and may have contributed to the
development of concurrent CRAO in this patient. Systemic lupus erythematosus (SLE) is a chronic, multisystem, autoimmune disease
that can affect any part of the human body including the eyes. Common blinding
ocular manifestations include central retinal artery occlusion (CRAO), central
retinal vein occlusion (CRVO), severe vaso-occlusive retinopathy, and optic
nerve involvement. Antiphospholipid syndrome (APS) in lupus is usually
associated with large vessel occlusions and needs prompt treatment with
anticoagulant. We are reporting two cases of APS in SLE patients that presented
with CRVO (case 1) and vaso-occlusive lupus retinopathy (case 2). Both cases
were positive for antiphospholipid antibody (APA) and were treated with
immunosuppression, anticoagulant, and laser treatment. Thus, screening for APA
is vital in SLE patients with lupus retinopathy, as prompt treatment with
anticoagulants is important to prevent further vascular thrombosis, which
worsens the visual prognosis. BACKGROUND: Central retinal artery occlusion (CRAO) is an ocular emergency and
most of the cases present with painless sudden persistent loss of vision in the
range of counting fingers to perception of light. The presentation of CRAO is
associated with a variety of medical conditions. We report a rare case of CRAO
associated with persistent truncus arteriosus (PTA) and single atrium in a
female patient.
CASE PRESENTATION: A 23-year-old woman was admitted due to sudden painless
visual loss in the left eye. On examination visual acuity of light-perception
was noted in the left eye with a left relative afferent pupillary defect.
Fundoscopic examination revealed retinal ischemic whitening, constriction of the
arteriole and venule with segmentation and typical "cherry-red spot" suggesting
CRAO. The patient was treated with ocular massage and anterior chamber
paracentesis. She was commenced on 150 mg of aspirin and also received
hyperbaric oxygen therapy. An echocardiogram revealed PTA and single atrium. A
diagnosis of CRAO associated with PTA and single atrium was made.
CONCLUSION: The ophthalmologist should enquire about congenital and acquired
cardiac abnormalities in patients with CRAO and consider such abnormalities to
be possible sources of emboli. Central retinal artery occlusion (CRAO) is a devastating ocular emergency
characterized by acute painless visual loss in the ipsilateral eye. We describe
the case of acute non-arteritic CRAO associated fusiform internal
carotid-ophthalmic artery aneurysm with intraluminal thrombus. Despite the
rarity of this condition, we suggest that carotid-ophthalmic artery aneurysm
should be included in the differential diagnosis of CRAO. Presentation of a combination of branch retinal artery occlusion (BRAO)/central
retinal artery occlusion (CRAO) and central retinal vein occlusion (CRVO) in
systemic lupus erythematosus (SLE) is extremely rare. Herein, we have presented
the case of a 29-year-old female with SLE, who simultaneously developed
bilateral CRVO and BRAO/CRAO in the absence of antiphospholipid syndrome (APS)
as a catastrophic form of clinical flare. A combinatorial diagnosis of CRVO and
BRAO/CRAO should be considered during clinical flare-up in a patient with SLE
who presents with rapidly progressive visual loss. BACKGROUND: Central retinal artery occlusion (CRAO) is an ophthalmological
emergency, the retinal analog of a stroke. To date there is no consensus or
national guidelines on how this disorder should be managed. As academic
neurologists and ophthalmologists treat CRAO frequently, we set out to
understand how these clinicians approach patients with CRAO with a national
survey.
METHODS: We identified university-associated teaching hospitals offering
vascular neurology, neuro-ophthalmology and/or retina fellowships in the US and
asked the directors of the programs to respond to questions in an open response
format to profile the acute management of CRAO at their institution.
RESULTS: We found remarkable heterogeneity in the approach to acute treatment of
patients with CRAO among the 45 institutions that responded to the survey. Only
20% had a formal policy, guideline or white paper to standardize the approach to
treatment. The primary treating physician was an ophthalmologist, neurologist,
or neuro-ophthalmologist 44, 27, and 4% of the time, respectively; 24% were
co-managed acutely by neurology and ophthalmology. Intravenous fibrinolysis was
offered to selected patients in 53% of institutions, and was the preferred
initial treatment in 36%. When the acute treatment team involved a vascular
neurologist, fibrinolysis was more likely to be considered a first-line
treatment (p < 0.05). Anterior chamber paracentesis, ocular massage and
hyperbaric oxygen therapy were offered 42, 66 and 7% of the time, respectively,
while 9% of institutions offered no treatment. Anterior chamber paracentesis was
more likely to be offered at programs where neurologists were not involved in
treating CRAOs (p < 0.001). At 35% of institutions, patients with acute CRAO
were not routinely referred to a general emergency room for initial evaluation
and treatment. Carotid imaging was routinely obtained by 89% of programs,
magnetic resoce imaging of the brain by 69%, echocardiogram by 62%,
laboratory screening for an inflammatory state by 27% and retinal angiography by
30%. The thoroughness of vascular risk factors' screening was greater in
programs that routinely referred acute CRAO cases to the emergency department.
CONCLUSIONS: This survey shows that there is significant variability in
treatment practices for acute CRAO in the US. Because of the high
cerebrovascular and cardiovascular risk reported in this population of patients,
it is notable that the approach to risk factor screening is also highly variable
and many programs do not routinely refer patients to an emergency department for
urgent evaluation. Finally, there appears to be equipoise among treatment teams
regarding the efficacy of systemic fibrinolysis, as 53% of programs report a
willingness to treat at least some patients with this modality. Central retinal artery occlusion (CRAO) is a medical emergency that, if not
treated, may result in irreversible loss of vision. It continues to be an
important cause for acute painless loss of vision. Amaurosis fugax or "transient
CRAO" has long been considered an equivalent of transient cerebral ischemic
event. Animal models, in addition to data from retrospective and randomized
clinical studies, provide valuable insights into the time interval for
irreversible retinal ischemia. Subset analyses from 2 large studies of patients
with CRAO show benefit from treatment with thrombolysis within 6 hours from
symptoms onset. Significant workflow improvements after the intra-arterial
therapy trials for acute ischemic stroke have occurred world over in last 5
years. Patients with CRAO are uniquely suited to receive maximum benefits from
the changes in workflow for treatment of patient's acute ischemic stroke. Just
as in clinical triage of acute ischemic stroke, correct and timely diagnosis of
patients with CRAO may help in preventing visual loss. The approach to acute
ocular ischemia should mimic that used for acute brain ischemia. Comprehensive
stroke centers would be ideal triage centers for these patients in view of
availability of multidisciplinary participation from vascular neurology,
neuroendovascular surgery, and ophthalmology. Time is Retina! Central retinal artery occlusion (CRAO) is a neuro-ophthalmological emergency.
There is a finite time window for acute interventions such as revascularization
(e.g. intravenous thrombolysis-IVT) and retinal oxygenation (e.g. hyperbaric
oxygen therapy-HBOT) therapies. Case 1: A 35-year-old female presented with CRAO
in the right eye (OD) confirmed by fluorescein angiography (FA) and optical
coherence tomography (OCT). She underwent 4 sessions of HBOT (100% O2 at 2.4
atmosphere absolute for 90 min). Afterwards, visual defect on the nasal field
was kept but visual acuity (VA) improved from counting fingers to 1.0. Case 2: A
65-year-old male presented with CRAO in his left eye (OS) with 1.5 h of
evolution. Orbital sonography and OCT confirmed the presence of an embolus and
he underwent IVT with rTPA (0.9 mg/kg). VA improved from light perception to
0.1. Case 3: A 21-year-old male presented acute visual loss in his OD with 2.5 h
of evolution. OCT and retinography identified CRAO. He was submitted to IVT
(rTPA-0.9 mg/kg) followed by 12 sessions of HBOT. VA improved from hand motion
to 1.0. Our case series depicts the approaches and possible outcomes in acute
management of an infrequent, but highly morbid, cerebroretinovascular disorder.
Future clinical trials are warranted to tackle current difficulties in CRAO
treatment. Vision loss is a known rare complication of prone positioning during surgery.
Vision loss following prone surgery is most commonly attributed to direct
pressure on the eye but can also be caused by central retinal artery occlusion
(CRAO) in the absence of pressure on the eye. Central retinal artery occlusion
has not been previously described following prone transcranial surgery for
craniosynostosis. We present two cases of monocular CRAO following prone
calvarial expansion. A multidisciplinary root cause analysis suggested that
raised intracranial pressure and intraoperative tranexamic acid may have been
risk factors for the development of CRAO in these cases as no conventional risk
factors for CRAO following prone surgery were present. Because of this, we
retrospectively reviewed all prone transcranial procedures performed at the
Oxford Craniofacial Unit for the presence of raised intracranial pressure and
intraoperative tranexamic acid use. A total of 662 prone procedures have been
performed between 1994 and March, 2019. Tranexamic acid has been used routinely
in all transcranial procedures since 2012 and in the last 311 consecutive prone
cases. Fifty-one (7.7%) prone procedures were performed for raised intracranial
pressure, and tranexamic acid was used in the 33 most recent of these. Since the
implementation of standard intraoperative administration of tranexamic acid
there have been 2 cases of CRAO following prone surgery. The overall incidence
of CRAO was 0.3% but was 6% in the context of raised intracranial pressure and
tranexamic acid use. Prone positioning raised intracranial pressure and
tranexamic acid use together may represent a potent combination of risk factors
for CRAO. |
Is yeast fbp1 affected by glucose starvation stress? | The chromatin configuration is altered into an accessible state within 290 bp downstream from the initiation site of metabolic-stress-induced lncRNAs (mlonRNAs) in the promoter of the fission yeast fbp1 gene, whose transcription is massively induced upon glucose starvation . We investigated the mechanisms by which chromatin is reconstituted . | The specific induction of genes in response to distinct environmental stress is
vital for all eukaryotes. To study the mechanisms that result in selective gene
responses, we examined the role of the fission yeast Tup1 family repressors in
chromatin regulation. We found that chromatin structure around a cAMP-responsive
element (CRE)-like sequence in ade6-M26 that is bound by Atf1.Pcr1
transcriptional activation was altered in response to osmotic stress but not to
heat and oxidative stresses. Such chromatin structure alteration occurred later
than the Atf1 phosphorylation but correlated well with stress-induced
transcriptional activation at ade6-M26. This chromatin structure alteration
required components for the stress-activated protein kinase (SAPK) cascade and
both subunits of the M26-binding CREB/ATF-type protein Atf1.Pcr1. Cation stress
and glucose starvation selectively caused chromatin structure alteration around
CRE-like sequences in cta3(+) and fbp1(+) promoters, respectively, in
correlation with transcriptional activation. However, the tup11Delta tup12Delta
double deletion mutants lost the selectivity of stress responses of chromatin
structure and transcriptional regulation of cta3(+) and fbp1(+). These data
indicate that the Tup1-like repressors regulate the chromatin structure to
ensure the specificity of gene activation in response to particular stresses.
Such a role for these proteins may serve as a paradigm for the regulation of
stress response in higher eukaryotes. BACKGROUND: Glucose is a signaling molecule which regulates multiple events in
eukaryotic organisms and the most preferred carbon source in the fission yeast
Schizosaccharomyces pombe. The ability of this yeast to grow in the absence of
glucose becomes strongly limited due to lack of enzymes of the glyoxylate cycle
that support diauxic growth. The stress-activated protein kinase (SAPK) pathway
and its effectors, Sty1 MAPK and transcription factor Atf1, play a critical role
in the adaptation of fission yeast to grow on alternative non-fermentable carbon
sources by inducing the expression of fbp1+ gene, coding for the gluconeogenic
enzyme fructose-1,6-bisphosphatase. The cell integrity Pmk1 pathway is another
MAPK cascade that regulates various processes in fission yeast, including cell
wall construction, cytokinesis, and ionic homeostasis. Pmk1 pathway also becomes
strongly activated in response to glucose deprivation but its role during
glucose exhaustion and ensuing adaptation to respiratory metabolism is currently
unknown.
RESULTS: We found that Pmk1 activation in the absence of glucose takes place
only after complete depletion of this carbon source and that such activation is
not related to an endogenous oxidative stress. Notably, Pmk1 MAPK activation
relies on de novo protein synthesis, is independent on known upstream activators
of the pathway like Rho2 GTPase, and involves PKC ortholog Pck2. Also, the
Glucose/cAMP pathway is required operative for full activation of the Pmk1
signaling cascade. Mutants lacking Pmk1 displayed a partial growth defect in
respiratory media which was not observed in the presence of glucose. This
phenotype was accompanied by a decreased and delayed expression of transcription
factor Atf1 and target genes fbp1+ and pyp2+. Intriguingly, the kinetics of Sty1
activation in Pmk1-less cells was clearly altered during growth adaptation to
non-fermentable carbon sources.
CONCLUSIONS: Unknown upstream elements mediate Pck2-dependent signal
transduction of glucose withdrawal to the cell integrity MAPK pathway. This
signaling cascade reinforces the adaptive response of fission yeast to such
nutritional stress by enhancing the activity of the SAPK pathway. It has been postulated that a myriad of long noncoding RNAs (lncRNAs) contribute
to gene regulation. In fission yeast, glucose starvation triggers lncRNA
transcription across promoter regions of stress-responsive genes including fbp1
(fructose-1,6-bisphosphatase1). At the fbp1 promoter, this transcription
promotes chromatin remodeling and fbp1 mRNA expression. Here, we demonstrate
that such upstream noncoding transcription facilitates promoter association of
the stress-responsive transcriptional activator Atf1 at the sites of
transcription, leading to activation of the downstream stress genes. Genome-wide
analyses revealed that ∼50 Atf1-binding sites show marked decrease in Atf1
occupancy when cells are treated with a transcription inhibitor. Most of these
transcription-enhanced Atf1-binding sites are associated with stress-dependent
induction of the adjacent mRNAs or lncRNAs, as observed in fbp1 These
Atf1-binding sites exhibit low Atf1 occupancy and high histone density in
glucose-rich conditions, and undergo dramatic changes in chromatin status after
glucose depletion: enhanced Atf1 binding, histone eviction, and histone H3
acetylation. We also found that upstream transcripts bind to the Groucho-Tup1
type transcriptional corepressors Tup11 and Tup12, and locally antagonize their
repressive functions on Atf1 binding. These results reveal a new mechanism in
which upstream noncoding transcription locally magnifies the specific activation
of stress-inducible genes via counteraction of corepressors. Antisense RNA has emerged as a crucial regulator of opposite-strand
protein-coding genes in the long noncoding RNA (lncRNA) category, but little is
known about their dynamics and decay process in the context of a stress
response. Antisense transcripts from the fission yeast fbp1 locus (fbp1-as) are
expressed in glucose-rich conditions and anticorrelated with transcription of
metabolic stress-induced lncRNA (mlonRNA) and mRNA on the sense strand during
glucose starvation. Here, we investigate the localization and decay of antisense
RNAs at fbp1 and other loci, and propose a model to explain the rapid switch
between antisense and sense mlonRNA/mRNA transcription triggered by glucose
starvation. We show that fbp1-as shares many features with mRNAs, such as a
5'-cap and poly(A)-tail, and that its decay partially depends upon Rrp6, a
cofactor of the nuclear exosome complex involved in 3'-5' degradation of RNA.
Fluorescence in situ hybridization and polysome fractionation show that the
majority of remaining fbp1-as localizes to the cytoplasm and binds to
polyribosomes in glucose-rich conditions. Furthermore, fbp1-as and antisense RNA
at other stress-responsive loci are promptly degraded via the cotranslational
nonsense-mediated decay (NMD) pathway. These results suggest NMD may potentiate
the swift disappearance of antisense RNAs in response to cellular stress. The arrangement of nucleosomes in chromatin plays a role in transcriptional
regulation by restricting the accessibility of transcription factors and RNA
polymerase II to cis-acting elements and promoters. For gene activation, the
chromatin structure is altered to an open configuration. The mechanism for this
process has been extensively analyzed. However, the mechanism by which
repressive chromatin is reconstituted to terminate transcription has not been
fully elucidated. Here, we investigated the mechanisms by which chromatin is
reconstituted in the fission yeast Schizosaccharomyces pombefbp1 gene, which is
robustly induced upon glucose starvation but tightly repressed under
glucose-rich conditions. We found that the chromatin structure in the region
upstream from fbp1 is closed by a two-step process. When cells are returned to
glucose-rich medium following glucose starvation, changes in the nucleosome
pattern alter the chromatin configuration at the transcription factor binding
site to an inaccessible state, after which the nucleosome density upstream from
fbp1 gradually increases via histone loading. Interestingly, this histone
loading was observed in the absence of the Tup family corepressors Tup11 and
Tup12. Analysis of strains carrying either gene disruptions or mutations
affecting nine fission yeast histone chaperone genes demonstrated that the
histone chaperone Asf1 induces nucleosome loading during glucose repression.
These data establish a previously unappreciated chromatin reconstitution
mechanism in fbp1 repression. |
What does csDMARD stand for? | csDMARDS are conventional synthetic disease-modifying antirheumatic drugs. | OBJECTIVE: To determine whether intensive combinations of conventional synthetic
disease-modifying antirheumatic drugs (csDMARDS) achieve similar clinical
benefits more cheaply than high-cost biologics such as tumor necrosis factor
inhibitors (TNFi) in patients with active rheumatoid arthritis (RA) whose
illness has failed to respond to methotrexate and another DMARD.
METHODS: We used within-trial cost-effectiveness and cost-utility analyses from
health and social care and 2 societal perspectives. Participants were recruited
into an open-label, 12-month, pragmatic, randomized, multicenter, 2-arm,
noninferiority trial in 24 rheumatology clinics in England and Wales. Costs were
linked with the Health Assessment Questionnaire (HAQ; primary outcome) and
quality-adjusted life years derived from 2 measures (Short-Form 36 health survey
and EuroQol 5-domain 3-level instrument).
RESULTS: In total, 205 participants were recruited, 104 in the csDMARD arm and
101 in the TNFi arm. Participants in the csDMARD arm with poor response at 6
months were offered TNFi; 46 participants (44%) switched. Relevant cost and
outcome data were available for 93% of participants at 6-month follow-up and for
91-92% of participants at 12-month follow-up. The csDMARD arm had significantly
lower total costs from all perspectives (6-month health and social care adjusted
mean difference -£3,615 [95% confidence interval (95% CI) -4,104, -3,182];
12-month health and social care adjusted mean difference -£1,930 [95% CI -2,599,
-1,301]). The HAQ score showed benefit to the csDMARD arm at 12 months (-0.16
[95% CI -0.32, -0.01]); other outcomes/follow-ups showed no differences.
CONCLUSION: Starting treatment with csDMARDs, rather than TNFi, achieves similar
outcomes at significantly lower costs. Patients with active RA and who meet the
National Institute for Health and Care Excellence criteria for expensive
biologics can be treated with combinations of intensive csDMARDs in a
cost-effective manner. |
What is caused by SCUBE2 loss-of-function? | Scube2 (-/-) caused defective endochondral bone formation and impaired Ihh-mediated chondrocyte differentiation and proliferation as well as osteoblast differentiation of -/- bone-marrow mesenchymal stromal-cell cultures. | Signal peptide-CUB-EGF domain-containing protein 2 (SCUBE2) belongs to a
secreted and membrane-tethered multidomain SCUBE protein family composed of
three members found in vertebrates and mammals. Recent reports suggested that
zebrafish scube2 could facilitate sonic hedgehog (Shh) signaling for proper
development of slow muscle. However, whether SCUBE2 can regulate the signaling
activity of two other hedgehog ligands (Ihh and Dhh), and the developmental
relevance of the SCUBE2-induced hedgehog signaling in mammals remain poorly
understood. In this study, we first showed that as compared with SCUBE1 or
SCUBE3, SCUBE2 is the most potent modulator of IHH signaling in vitro. In
addition, gain and loss-of-function studies demonstrated that SCUBE2 exerted an
osteogenic function by enhancing Ihh-stimulated osteoblast differentiation in
the mouse mesenchymal progenitor cells. Consistent with these in vitro studies
and the prominent roles of Ihh in coordinating skeletogenesis, genetic ablation
of Scube2 (-/-) caused defective endochondral bone formation and impaired
Ihh-mediated chondrocyte differentiation and proliferation as well as osteoblast
differentiation of -/- bone-marrow mesenchymal stromal-cell cultures. Our data
demonstrate that Scube2 plays a key regulatory role in Ihh-dependent
endochondral bone formation. |
Is Semagacestat effective for Alzheimer's Disease? | No. In a placebo controlled clinical trial, semagacestat did not improve cognitive status, and patients receiving the higher dose had significant worsening of functional ability. The trial was terminated due to unexpected aggravation of cognitive deficits and side effects. | The recent failure of semagacestat in two large Phase III studies questions the
value of γ-secretase inhibitors in treating Alzheimer's disease. Understanding
the reasons of this setback may be important for the future research on
effective treatments for this devastating disease. Several second-generation active β-amyloid (Aβ) vaccines and passive Aβ
immunotherapies are under clinical investigation with the aim of boosting Aβ
clearance from the brain of the Alzheimer's disease (AD) patients. However, the
preliminary cognitive efficacy of bapineuzumab, a humanized anti-Aβ monoclonal
antibody, appears uncertain. Moreover, the occurrence of vasogenic edema and,
more rarely, brain microhemorrhages, especially in apolipoprotein E ϵ4 carriers,
have led to abandoning of the highest dose of the drug. Solanezumab, another
humanized anti-Aβ monoclonal antibody, was shown to neutralize soluble Aβ
oligomers, which is believed to be the more neurotoxic Aβ species. Phase II
studies showed a good safety profile of solanezumab while studies on
cerebrospinal and plasma biomarkers documented good signals of pharmacodynamic
activity. However, the preliminary equivocal cognitive results obtained with
bapineuzumab as well as the detrimental cognitive effects observed with
semagacestat, a potent γ-secretase inhibitor, raise the possibility that
targeting Aβ may not be clinically efficacious in AD. The results of four
ongoing large Phase III trials on bapineuzumab and two Phase III trials on
solanezumab will tell us if passive anti-Aβ immunization is able to alter the
course of this devastating disease, and if Aβ is still a viable target for
anti-AD drugs. Drugs currently used for the treatment of Alzheimer's disease (AD) produce
limited clinical benefits, and there is no disease-modifying therapy yet
available. Compounds that inhibit or modulate γ-secretase, the pivotal enzyme
that generates β-amyloid (Aβ), are potential therapeutics for AD. This article
briefly reviews the profile of γ-secretase inhibitors and modulators that have
reached the clinic. Studies in both transgenic and non-transgenic animal models
of AD have indicated that γ-secretase inhibitors, administered by the oral
route, are able to lower brain Aβ concentrations. However, scanty data are
available on the effects of these compounds on brain Aβ deposition after
prolonged administration. γ-Secretase inhibitors may cause abnormalities in the
gastrointestinal tract, thymus, spleen, skin, and decrease in lymphocytes and
alterations in hair color in experimental animals and in man, effects believed
to be associated with the inhibition of the cleavage of Notch, a transmembrane
receptor involved in regulating cell-fate decisions. Unfortunately, two large
Phase III clinical trials of semagacestat in mild-to-moderate AD patients were
prematurely interrupted because of the observation of a detrimental cognitive
and functional effect of the drug. These detrimental effects were mainly
ascribed to the inhibition of the processing of an unknown substrate of
γ-secretase. It has been also hypothesized that the detrimental cognitive
effects observed after semagacestat administration are due to the accumulation
of the neurotoxic precursor of Aβ (the carboxy-terminal fragment of amyloid
precursor protein, APP, or CTFβ) resulting from the block of the γ-secretase
cleavage activity on APP. Some non-steroidal anti-inflammatory drugs and other
small organic molecules have been found to modulate γ-secretase shifting its
cleavage activity from longer to shorter Aβ species without affecting Notch
cleavage. However, two large Phase III studies in mild AD patients with
tarenflurbil, a putative γ-secretase modulator, were also completely negative.
The failure of tarenflurbil was ascribed to low potency and brain penetration.
New more selective γ-secretase inhibitors and more potent, more brain penetrant
γ-secretase modulators are being developed with the hope of overcoming the
previous setbacks. Further understanding of the reasons of the failures of these
γ-secretase-based drugs in AD may be important for the future research on
effective treatments for this devastating disease. Neurological and psychiatric disorders are frequently associated with disruption
of various cognitive functions, but development of effective drug treatments for
these conditions has proven challenging. One of the main obstacles is the poor
predictive validity of our preclinical animal models. In the present study the
effects of the γ-secretase inhibitor semagacestat was evaluated in preclinical
in vivo electrophysiological models. Recently disclosed Phase III findings on
semagacestat indicated that Alzheimer's disease (AD) patients on this drug
showed significantly worsened cognitive function compared to those treated with
placebo. Since previous studies have shown that drugs impairing cognitive
function (including scopolamine, NMDA (N-methyl-D-aspartate) receptor
antagonists, and nociceptin receptor agonists) disrupt or decrease power of
elicited theta oscillation in the hippocampus, we tested the effects of acute
and sub-chronic administration of semagacestat in this assay. Field potentials
were recorded across the hippocampal formation with NeuroNexus multi-site
silicon probes in urethane anesthetized male C57BL/6 mice; hippocampal CA1 theta
oscillation was elicited by electrical stimulation of the brainstem nucleus
pontis oralis. Sub-chronic administration of semagacestat twice daily over 12
days at a dose known to reduce beta-amyloid peptide (Aβ) level [100 mg/kg, p.o.
(per oral)] diminished power of elicited hippocampal theta oscillation. Acute,
subcutaneous administration of semagacestat (100 mg/kg) produced a similar
effect on hippocampal activity. We propose that the disruptive effect of
semagacestat on hippocampal function could be one of the contributing mechanisms
to its worsening of cognition in patients with AD. As it has been expected, both
acute and sub-chronic administrations of semagacestat significantly decreased
Aβ40 and Aβ42 levels but the current findings do not reveal the mode of action
of semagacestat in disrupting hippocampal oscillation. BACKGROUND: Alzheimer's disease is characterized by the presence of cortical
amyloid-beta (Aβ) protein plaques, which result from the sequential action of
β-secretase and γ-secretase on amyloid precursor protein. Semagacestat is a
small-molecule γ-secretase inhibitor that was developed as a potential treatment
for Alzheimer's disease.
METHODS: We conducted a double-blind, placebo-controlled trial in which 1537
patients with probable Alzheimer's disease underwent randomization to receive
100 mg of semagacestat, 140 mg of semagacestat, or placebo daily. Changes in
cognition from baseline to week 76 were assessed with the use of the cognitive
subscale of the Alzheimer's Disease Assessment Scale for cognition (ADAS-cog),
on which scores range from 0 to 70 and higher scores indicate greater cognitive
impairment, and changes in functioning were assessed with the Alzheimer's
Disease Cooperative Study-Activities of Daily Living (ADCS-ADL) scale, on which
scores range from 0 to 78 and higher scores indicate better functioning. A
mixed-model repeated-measures analysis was used.
RESULTS: The trial was terminated before completion on the basis of a
recommendation by the data and safety monitoring board. At termination, there
were 189 patients in the group receiving placebo, 153 patients in the group
receiving 100 mg of semagacestat, and 121 patients in the group receiving 140 mg
of semagacestat. The ADAS-cog scores worsened in all three groups (mean change,
6.4 points in the placebo group, 7.5 points in the group receiving 100 mg of the
study drug, and 7.8 points in the group receiving 140 mg; P=0.15 and P=0.07,
respectively, for the comparison with placebo). The ADCS-ADL scores also
worsened in all groups (mean change at week 76, -9.0 points in the placebo
group, -10.5 points in the 100-mg group, and -12.6 points in the 140-mg group;
P=0.14 and P<0.001, respectively, for the comparison with placebo). Patients
treated with semagacestat lost more weight and had more skin cancers and
infections, treatment discontinuations due to adverse events, and serious
adverse events (P<0.001 for all comparisons with placebo). Laboratory
abnormalities included reduced levels of lymphocytes, T cells, immunoglobulins,
albumin, total protein, and uric acid and elevated levels of eosinophils,
monocytes, and cholesterol; the urine pH was also elevated.
CONCLUSIONS: As compared with placebo, semagacestat did not improve cognitive
status, and patients receiving the higher dose had significant worsening of
functional ability. Semagacestat was associated with more adverse events,
including skin cancers and infections. (Funded by Eli Lilly; ClinicalTrials.gov
number, NCT00594568.) OBJECTIVE: Semagacestat, a γ-secretase inhibitor, demonstrated an unfavorable
risk-benefit profile in a Phase 3 study of patients with Alzheimer's disease
(IDENTITY trials), and clinical development was halted. To assist in future
development of γ-secretase inhibitors, we report detailed safety findings from
the IDENTITY study, with emphasis on those that might be mechanistically linked
to γ-secretase inhibition.
RESEARCH DESIGN AND METHODS: The IDENTITY trial was a double-blind,
placebo-controlled trial of semagacestat (100 mg and 140 mg), in which 1537
patients age 55 years and older with probable Alzheimer's disease were
randomized. Treatment-emergent adverse events (TEAEs) are reported by body
system along with pertinent laboratory, vital sign, and ECG findings.
RESULTS: Semagacestat treatment was associated with increased reporting of
suspected Notch-related adverse events (gastrointestinal, infection, and skin
cancer related). Other relevant safety findings associated with semagacestat
treatment included cognitive and functional worsening, skin-related TEAEs, renal
and hepatic changes, increased QT interval, and weight loss. With few
exceptions, differences between semagacestat and placebo treatment groups were
no longer significant after cessation of treatment with active drug.
CONCLUSIONS: Many of these safety findings can be attributed to γ-secretase
inhibition, and may be valuable to researchers developing γ-secretase
inhibitors. BACKGROUND: In a recent report, 76 weeks' treatment with a gamma-secretase
inhibitor (semagacestat) was associated with poorer cognitive outcomes in
Alzheimer's disease (AD).
OBJECTIVE: We sought to examine the effect of semagacestat treatment on
neuropsychiatric symptoms (NPS).
METHODS: 1,537 participants with mild to moderate AD were randomized to 76
weeks' treatment with placebo versus two doses of semagacestat. NPS were
assessed with the Neuropsychiatric Inventory (NPI-Total and subdomains).
Cognition was assessed with the Alzheimer's Disease Assessment Scale-Cognitive
(first 11 items, ADAS11). Mixed-Model Repeated Measures was used to compare the
effects of treatment assignment on change in NPI-total and subdomains over time.
Survival analysis was used to assess the treatment effect on time to first
worsening of NPS (NPI-Total ≥10 or NPI subdomain ≥4) for subjects with no or
minor NPS at baseline.
RESULTS: Participants on high dose semagecestat (140 mg) had greater increase in
NPI-Total and greater risk of incident first worsening in NPI-Total and in
subdomains of aberrant motor behavior, appetite, depression/dysphoria, and
sleep. ADAS11 increased more in participants whose NPI-Total increased.
CONCLUSION: In participants with mild to moderate AD, high dose semagacestat
treatment was associated with greater severity and faster worsening of NPS in a
pattern resembling an agitated depression. Increased NPS was associated with
cognitive decline regardless of treatment assignment. These findings suggest
that greater NPS may be the result of gamma-secretase treatment and emphasize
the importance of monitoring NPS as potential adverse events in trials of novel
treatments for AD. Author information:
(1)Neuropsychiatry, Department of Integrated Medicine, Division of Internal
Medicine, Osaka University Graduate School of Medicine, Suita, Osaka 565-0871,
Japan.
(2)Department of Neuropathology, Faculty of Life and Medical Sciences, Doshisha
University, Kizugawa, Kyoto 619-0225, Japan.
(3)Neuropsychiatry, Department of Integrated Medicine, Division of Internal
Medicine, Osaka University Graduate School of Medicine, Suita, Osaka 565-0871,
Japan; Pain and Neurology, Shionogi & Co. Ltd., Osaka, Osaka 561-0825, Japan.
(4)Department of Molecular Virology, Research Institute for Microbial Diseases,
Osaka University, Suita, Osaka 565-0871, Japan.
(5)Departments of Neurology and Molecular Genetics, Brain Research Institute,
Niigata University, Niigata, Niigata 951-8520, Japan.
(6)Pain and Neurology, Shionogi & Co. Ltd., Osaka, Osaka 561-0825, Japan.
(7)Department of Psychiatry, Osaka University Health Care Center, Toyonaka,
Osaka 560-0043, Japan.
(8)Neuropsychiatry, Department of Integrated Medicine, Division of Internal
Medicine, Osaka University Graduate School of Medicine, Suita, Osaka 565-0871,
Japan. Electronic address: [email protected]. |
List enzymes that removes histone modifications. | Histone deacetylases
Lysine Specific Demethylases | Lysine residues across the proteome are modified by posttranslational
modifications (PTMs) that significantly enhance the structural and functional
diversity of proteins. For lysine, the most abundant PTM is ɛ-N-acetyllysine
(Kac), which plays numerous roles in regulation of important cellular functions,
such as gene expression (epigenetic effects) and metabolism. A family of
enzymes, namely histone deacetylases (HDACs), removes these PTMs. A subset of
these enzymes, the sirtuins (SIRTs), represent class III HDAC and, unlike the
rest of the family, these hydrolases are NAD+-dependent. Although initially
described as deacetylases, alternative deacylase functions for sirtuins have
been reported, which expands the potential cellular roles of this class of
enzymes. Currently, sirtuins are investigated as therapeutic targets for the
treatment of diseases that span from cancers to neurodegenerative disorders. In
the present book chapter, we review and discuss the current literature on novel
ɛ-N-acyllysine PTMs, targeted by sirtuins, as well as mechanism-based sirtuin
inhibitors inspired by their substrates. Protein methylation has an important role in the regulation of chromatin, gene
expression and regulation. The protein methyl transferases are genetically
altered in various human cancers. The enzymes that remove histone methylation
have led to increased awareness of protein interactions as potential drug
targets. Specifically, Lysine Specific Demethylases (LSD) removes methylated
histone H3 lysine 4 (H3K4) and H3 lysine 9 (H3K9) through
formaldehyde-generating oxidation. It has been reported that LSD1 and its
downstream targets are involved in tumor-cell growth and metastasis. Functional
studies of LSD1 indicate that it regulates activation and inhibition of gene
transcription in the nucleus. Here we made a discussion about the summary of
histone lysine demethylase and their functions in various human cancers. |
Roughly how many base pairs are in the human mitochondrial genome or mtDNA? | The mitochondrial genome, mtDNA, is 16569 base pairs. | It is now clear that molecular defects in human mitochondrial DNA play a
significant role in human disease. Mitochondrial DNA mutations range from single
base changes in the 16.5 kilobase-pair genome up to large deletions and
rearrangements. Independently of the actual cause of a given mutation, it is
possible to predict at least some of the consequences of changes in
mitochondrial DNA sequence. This paper reviews our overall understanding of the
mode and mechanism of mitochondrial DNA replication and transcription and how
this relates to mitochondrial gene expression. This provides a background to
anticipate the nature and extent of mitochondrial DNA sequence changes that
might be of physiological consequence. The 16569 base pairs of the mitochondrial DNA encode with a specific genetic
code 13 proteins involved in the respiratory chain complex formation. Nuclear
gene products also contribute to the formation of these complexes. In the first
point, the organization and expression of the mtDNA are described with the main
characteristics of the enzymatic complexes as well as nuclear gene expression.
New information concerned with mitochondrial DNA deletions and mutations are
described particularly with respect to Kearns-Sayre Syndrome. Denaturing high pressure liquid chromatography (dHPLC) is an efficient method
for discovery of unknown mutations by heteroduplex analysis of PCR fragments.
For comprehensive mutation scanning of the whole 16.569 bp human mitochondrial
genome, we developed a set of 67 primer pairs defining overlapping PCR fragments
that are well suited for heteroduplex analysis. The aim of our optimization
efforts was to ensure that point mutations are detectable at every nucleotide
position of each amplicon. Some GC-rich regions of mitochondrial DNA (mtDNA)
were found to have unfavourable melting profiles in all possible amplicons,
therefore requiring GC-clamps at the end of one or both oligonucleotide PCR
primers. Following detection of a heteroduplex pattern by dHPLC, our primers can
also be employed for DNA sequencing to identify the underlying mutation. In case
of heteroplasmic mutations with a low proportion of mutant mtDNA, a fragment
collector is useful to recover the heteroduplex peak, which contains mutant and
wildtype DNA molecules in a 1:1 ratio. BACKGROUND: Somatic mutations of mitochondrial DNA (mtDNA) are increasingly
being recognized in many human cancers, but automated sequencing of 16.5 kb of
DNA poses an onerous task. We have recently described an oligonucleotide
microarray (MitoChip) for rapid and accurate sequencing of the entire
mitochondrial genome (Zhou et al., J Mol Diagnostics, 8: 9_14, 2006), greatly
facilitating the analysis of mtDNA mutations in cancer. In this report, we
perform a comprehensive cataloging of somatic mutations in the mitochondrial
genome of human pancreatic cancers using our novel array-based approach.
MATERIALS AND METHODS: MitoChip analysis was performed on DNA isolated from 15
histologically confirmed resection specimens of pancreatic ductal
adenocarcinomas. In all cases, matched nonneoplastic pancreatic tissue was
obtained as germline control for mtDNA sequence. DNA was extracted from
snap-frozen cryostat-embedded specimens and hybridized to the sequencing
microarray after appropriate polymerase chain reaction amplification and
labeling steps. The vast majority of somatic mutational analyses of mtDNA in
human cancers utilize lymphocyte DNA as germline control, without excluding the
potential for organ-specific polymorphisms. Therefore, we also examined a series
of 15 paired samples of DNA obtained from nonneoplastic pancreata and
corresponding EBV-immortalized lymphoblastoid cell lines to determine whether
lymphocyte DNA provides an accurate surrogate for the mtDNA sequence of
pancreatic tissue.
RESULTS: We sequenced 497,070 base pairs of mtDNA in the 15 matched samples of
pancreatic cancer and nonneoplastic pancreatic tissue, and 467,269 base pairs
(94.0%) were assigned by the automated genotyping software. All 15 pancreatic
cancers demonstrated at least one somatic mtDNA mutation compared to the control
germline DNA with a range of 1-14 alterations. Of the 71 somatic mutations
observed in our series, 18 were nonsynonymous coding region alterations (i.e.,
resulting in an amino acid change), 22 were synonymous coding region
alterations, and 31 involved noncoding mtDNA segments (including ribosomal and
transfer RNAs). Overall, somatic mutations in the coding region most commonly
involved the ND4, COI, and CYTB genes; of note, an A-G transition at nucleotide
position 841 in the 12sRNA was observed in three independent samples. In the
paired analysis of nonneoplastic pancreata and lymphoblastoid cell line DNA, 14
nucleotide discrepancies were observed out of 226,876 nucleotide sequences (a
concordance rate of 99.99%), with 9 samples demonstrating a perfect match across
all bases assigned.
CONCLUSIONS: Our findings confirm that somatic mtDNA mutations are common in
pancreatic cancers, and therefore, have the potential to be a clinically useful
biomarker for early detection. Further, our studies confirm that lymphocyte DNA
is an excellent, albeit not perfect, surrogate for nonneoplastic pancreatic
tissues in terms of being utilized as a germline control. Finally, our report
confirms the utility of a high-throughput array-based platform for mtDNA
mutational analyses of human cancers. The human mitochondrial genome consists of a multicopy, circular dsDNA molecule
of 16,569 base pairs. It encodes for 13 proteins, two ribosomal genes, and 22
tRNAs that are essential in the generation of cellular ATP by oxidative
phosphorylation in eukaryotic cells. Germline mutations in mitochondrial DNA
(mtDNA) are an important cause of maternally inherited diseases, while somatic
mtDNA mutations may play important roles in aging and cancer. mtDNA
polymorphisms are also widely used in population and forensic genetics.
Therefore, methods that allow the rapid, inexpensive and accurate sequencing of
mtDNA are of great interest. One such method is the Affymetrix GeneChip Human
Mitochondrial Resequencing Array 2.0 (MitoChip v.2.0) (Santa Clara, CA). A
direct comparison of 93 worldwide mitochondrial genomes sequenced by both the
MitoChip and dideoxy terminator sequencing revealed an average call rate of
99.48% and an accuracy of > or =99.98% for the MitoChip. The good performance
was achieved by using in-house software for the automated analysis of additional
probes on the array that cover the most common haplotypes in the hypervariable
regions (HVR). Failure to call a base was associated mostly with the presence of
either a run of > or =4 C bases or a sequence variant within 12 bases up- or
downstream of that base. A major drawback of the MitoChip is its inability to
detect insertions/deletions and its low sensitivity and specificity in the
detection of heteroplasmy. However, the vast majority of haplogroup defining
polymorphism in the mtDNA phylogeny could be called unambiguously and more
rapidly than with conventional sequencing. Human mitochondrial DNA, the 25th chromosome, is a 16 569 base pair long
circular molecule, that encoders a variety of genes for the translational
machinery of the mitochondrion, as well as 13 structural proteins, that are all
subunits of the respiratory chain (RC). A variety of alterations of
mitochondrial DNA (mtDNA) are now functionally and genetically linked to human
disease, including encephalomyopathies, Leber's hereditary optic neuropathy,
diabetes mellitus, some neurogenerative diseases and even ageing. The
mitochondrial encephalomyopathies are a clinically, biochemically and
genetically heterogeneous group of disorders. Alterations of mtDNA include point
mutations, encephalomyopathies, as well as in the other human diseases, are
reviewd. In humans, mitochondrial DNA (mtDNA) is a 16,569-bp double-stranded circular
molecule, encoding 37 genes, and is exclusively transmitted from the mother.
According to the recent findings from many studies of mitochondrial diseases
caused by nuclear gene mutations, the accumulation of somatic mtDNA mutations in
tissues has been expected to contribute toward age-associated mitochondrial
dysfunction and a life span. Human mitochondria harbor an essential, high copy number, 16,569 base pair,
circular DNA genome that encodes 13 gene products required for electron
transport and oxidative phosphorylation. Mutation of this genome can compromise
cellular respiration, ultimately resulting in a variety of progressive metabolic
diseases collectively known as 'mitochondrial diseases'. Mutagenesis of mtDNA
and the persistence of mtDNA mutations in cells and tissues is a complex topic,
involving the interplay of DNA replication, DNA damage and repair, purifying
selection, organelle dynamics, mitophagy, and aging. We briefly review these
general elements that affect maintece of mtDNA, and we focus on nuclear genes
encoding the mtDNA replication machinery that can perturb the genetic integrity
of the mitochondrial genome. The small (16,569 base pair) human mitochondrial genome plays a significant role
in cell metabolism and homeostasis. Mitochondrial DNA (mtDNA) contributes to the
generation of complexes which are essential to oxidative phosphorylation
(OXPHOS). As such, mtDNA is directly integrated into mitochondrial biogenesis
and signaling and regulates mitochondrial metabolism in concert with
nuclear-encoded mitochondrial factors. Mitochondria are a highly dynamic,
pleiomorphic network that undergoes fission and fusion events. Within this
network, mtDNAs are packaged into structures called nucleoids which are actively
distributed in discrete foci within the network. This sensitive organelle is
frequently disrupted by insults such as oxidants and inflammatory cytokines, and
undergoes genomic damage with double- and single-strand breaks that impair its
function. Collectively, mtDNA is emerging as a highly sensitive indicator of
cellular stress, which is directly integrated into the mitochondrial network as
a contributor of a wide range of critical signaling pathways. There are approximately 1500 proteins that are needed for mitochondrial
structure and function, most of which are encoded in the nuclear genome (Calvo
et al., 2006). Each mitochondrion has its own genome (mtDNA), which in humans
encodes 13 polypeptides, 22 tRNAs and 2 rRNAs required for oxidative
phosphorylation. The mitochondrial genome of humans and most vertebrates is
approximately 16.5kbp, double-stranded, circular, with few non-coding bases.
Thus, maintaining mtDNA stability, that is, the ability of the cell to maintain
adequate levels of mtDNA template for oxidative phosphorylation is essential and
can be impacted by the level of mtDNA mutation currently within the cell or
mitochondrion, but also from errors made during normal mtDNA replication,
defects in mitochondrial quality control mechanisms, and exacerbated by
exposures to exogenous and/or endogenous genotoxic agents. In this review, we
expand on the origins and consequences of mtDNA instability, the current state
of research regarding the mechanisms by which mtDNA instability can be overcome
by cellular and chemical interventions, and the future of research and
treatments for mtDNA instability. Mitochondria are essential cytoplasmic organelles that generate energy (ATP) by
oxidative phosphorylation and mediate key cellular processes such as apoptosis.
They are maternally inherited and in humans contain a 16,569-base-pair circular
genome (mtDNA) encoding 37 genes required for oxidative phosphorylation.
Mutations in mtDNA cause a range of pathologies, commonly affecting
energy-demanding tissues such as muscle and brain. Because mitochondrial
diseases are incurable, attention has focused on limiting the inheritance of
pathogenic mtDNA by mitochondrial replacement therapy (MRT). MRT aims to avoid
pathogenic mtDNA transmission between generations by maternal spindle transfer,
pronuclear transfer or polar body transfer: all involve the transfer of nuclear
DNA from an egg or zygote containing defective mitochondria to a corresponding
egg or zygote with normal mitochondria. Here we review recent developments in
animal and human models of MRT and the underlying biology. These have led to
potential clinical applications; we identify challenges to their technical
refinement. |
Do nematodes contain a CTCF gene? | Our findings show that CTCF and possibly chromatin insulation are present in basal nematodes. We suggest that the insulator protein CTCF has been secondarily lost in derived nematodes like C. elegans. | Conserved noncoding elements (CNEs) constitute the majority of sequences under
purifying selection in the human genome, yet their function remains largely
unknown. Experimental evidence suggests that many of these elements play
regulatory roles, but little is known about regulatory motifs contained within
them. Here we describe a systematic approach to discover and characterize
regulatory motifs within mammalian CNEs by searching for long motifs (12-22 nt)
with significant enrichment in CNEs and studying their biochemical and genomic
properties. Our analysis identifies 233 long motifs (LMs), matching a total of
approximately 60,000 conserved instances across the human genome. These motifs
include 16 previously known regulatory elements, such as the histone 3'-UTR
motif and the neuron-restrictive silencer element, as well as striking examples
of novel functional elements. The most highly enriched motif (LM1) corresponds
to the X-box motif known from yeast and nematode. We show that it is bound by
the RFX1 protein and identify thousands of conserved motif instances, suggesting
a broad role for the RFX family in gene regulation. A second group of motifs
(LM2*) does not match any previously known motif. We demonstrate by biochemical
and computational methods that it defines a binding site for the CTCF protein,
which is involved in insulator function to limit the spread of gene activation.
We identify nearly 15,000 conserved sites that likely serve as insulators, and
we show that nearby genes separated by predicted CTCF sites show markedly
reduced correlation in gene expression. These sites may thus partition the human
genome into domains of expression. BACKGROUND: The zinc finger (ZF) protein CTCF (CCCTC-binding factor) is highly
conserved in Drosophila and vertebrates where it has been shown to mediate
chromatin insulation at a genomewide level. A mode of genetic regulation that
involves insulators and insulator binding proteins to establish independent
transcriptional units is currently not known in nematodes including
Caenorhabditis elegans. We therefore searched in nematodes for orthologs of
proteins that are involved in chromatin insulation.
RESULTS: While orthologs for other insulator proteins were absent in all 35
analysed nematode species, we find orthologs of CTCF in a subset of nematodes.
As an example for these we cloned the Trichinella spiralis CTCF-like gene and
revealed a genomic structure very similar to the Drosophila counterpart. To
investigate the pattern of CTCF occurrence in nematodes, we performed
phylogenetic analysis with the ZF protein sets of completely sequenced
nematodes. We show that three ZF proteins from three basal nematodes cluster
together with known CTCF proteins whereas no zinc finger protein of C. elegans
and other derived nematodes does so.
CONCLUSION: Our findings show that CTCF and possibly chromatin insulation are
present in basal nematodes. We suggest that the insulator protein CTCF has been
secondarily lost in derived nematodes like C. elegans. We propose a switch in
the regulation of gene expression during nematode evolution, from the common
vertebrate and insect type involving distantly acting regulatory elements and
chromatin insulation to a so far poorly characterised mode present in more
derived nematodes. Here, all or some of these components are missing. Instead
operons, polycistronic transcriptional units common in derived nematodes,
seemingly adopted their function. The CCCTC-binding factor (CTCF) is multi-functional, ubiquitously expressed, and
highly conserved from Drosophila to human. It has important roles in
transcriptional insulation and the formation of a high-dimensional chromatin
structure. CTCF has a paralog called "Brother of Regulator of Imprinted Sites"
(BORIS) or "CTCF-like" (CTCFL). It binds DNA at sites similar to those of CTCF.
However, the expression profiles of the two proteins are quite different. We
investigated the evolutionary trajectories of the two proteins after the
duplication event using a phylogenomic and interactomic approach. We find that
CTCF has 52 direct interaction partners while CTCFL only has 19. Almost all
interactors already existed before the emergence of CTCF and CTCFL. The unique
secondary loss of CTCF from several nematodes is paralleled by a loss of two of
its interactors, the polycomb repressive complex subunit SuZ12 and the
multifunctional transcription factor TYY1. In contrast to earlier studies
reporting the absence of BORIS from birds, we present evidence for a multigene
synteny block containing CTCFL that is conserved in mammals, reptiles, and
several species of birds, indicating that not the entire lineage of birds
experienced a loss of CTCFL. Within this synteny block, BORIS and its genomic
neighbors seem to be partitioned into two nested chromatin loops. The high
expression of SPO11, RAE1, RBM38, and PMEPA1 in male tissues suggests a possible
link between CTCFL, meiotic recombination, and fertility-associated phenotypes.
Using the 65,700 exomes and the 1000 genomes data, we observed a higher number
of intergenic, non-synonymous, and loss-of-function mutations in CTCFL than in
CTCF, suggesting a reduced strength of purifying selection, perhaps due to less
functional constraint. |
Is tocilizumab a csDMARD? | No, tocilizumab is a biological DMARD (bDMARD). | |
What is caused by heterozygous lamin B1 and lamin B2 variants? | Heterozygous lamin B1 and lamin B2 variants cause primary microcephaly and define a novel laminopathy. | Lamin B1 plays an important role in the nuclear envelope stability, the
regulation of gene expression, and neural development. Duplication of LMNB1, or
missense mutations increasing LMNB1 expression, are associated with
autosomal-domit leukodystrophy. On the basis of its role in neurogenesis, it
has been postulated that LMNB1 variants could cause microcephaly. Here, we
confirm this hypothesis with the identification of de novo mutations in LMNB1 in
seven individuals with pronounced primary microcephaly (ranging from -3.6 to -12
SD) associated with relative short stature and variable degree of intellectual
disability and neurological features as the core symptoms. Simplified gyral
pattern of the cortex and abnormal corpus callosum were noted on MRI of three
individuals, and these individuals also presented with a more severe phenotype.
Functional analysis of the three missense mutations showed impaired formation of
the LMNB1 nuclear lamina. The two variants located within the head group of
LMNB1 result in a decrease in the nuclear localization of the protein and an
increase in misshapen nuclei. We further demonstrate that another mutation,
located in the coil region, leads to increased frequency of condensed nuclei and
lower steady-state levels of lamin B1 in proband lymphoblasts. Our findings
collectively indicate that de novo mutations in LMNB1 result in a domit and
damaging effect on nuclear envelope formation that correlates with microcephaly
in humans. This adds LMNB1 to the growing list of genes implicated in severe
autosomal-domit microcephaly and broadens the phenotypic spectrum of the
laminopathies. Author information:
(1)MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine,
University of Edinburgh, Edinburgh, UK.
(2)Yorkshire Regional Genetics Service, Leeds Teaching Hospitals NHS Trust,
Department of Clinical Genetics, Chapel Allerton Hospital, Leeds, UK.
(3)West Midlands Regional Genetics Service, Birmingham Women's NHS Foundation
Trust, Birmingham Women's Hospital, Edgbaston, Birmingham, UK.
(4)Northern Ireland Regional Genetics Service, Belfast City Hospital, Belfast,
UK.
(5)Liverpool Centre for Genomic Medicine, Liverpool Women's Hospital, Liverpool,
UK.
(6)Faculty of Medicine, University of Southampton, Southampton, UK.
(7)Wessex Clinical Genetics Service, University Hospital Southampton, University
Hospital Southampton NHS Foundation Trust, Southampton, UK.
(8)Wessex Clinical Genetics Service, Princess Anne Hospital, University Hospital
Southampton NHS Foundation Trust, Southampton, UK.
(9)Human Development and Health, Faculty of Medicine, University of Southampton,
Southampton, UK.
(10)Department of Radiology, Royal Hospital for Sick Children, Edinburgh, UK.
(11)Department of Clinical Genetics, Aberdeen Royal Infirmary, Scotland, UK.
(12)Clinical Genetics Service, Nottingham University Hospitals NHS Trust, City
Hospital Campus, Nottingham, UK.
(13)MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine,
University of Edinburgh, Edinburgh, UK. [email protected]. |
What is the risk for secondary cancer after proton beam therapy? | Proton beam therapy is associated with lower risk of secondary cancer when compared to other radiation therapy approaches. It allows excellent dose localization by administration of a high dose to the tumor while minimizing damage to surrounding normal tissues. Therefore it is more commonly used in children. | The risk of induction of a second primary cancer after a therapeutic irradiation
with conventional photon beams is well recognized and documented. However, in
general, it is totally overwhelmed by the benefit of the treatment. The same is
true to a large extent for the combinations of radiation and drug therapy. After
fast neutron therapy, the risk of induction of a second cancer is greater than
after photon therapy. Neutron RBE increases with decreasing dose and there is a
wide evidence that neutron RBE is greater for cancer induction (and for other
late effects relevant in radiation protection) than for cell killing. Animal
data on RBE for tumor induction are reviewed, as well as other biological
effects such as life shortening, maligt cell transformation in vitro,
chromosome aberrations, genetic effects. These effects can be related, directly
or indirectly, to cancer induction to the extent that they express a "genomic"
lesion. Almost no reliable human epidemiological data are available so far. For
fission neutrons a RBE for cancer induction of about 20 relative to photons
seems to be a reasonable assumption. For fast neutrons, due to the difference in
energy spectrum, a RBE of 10 can be assumed. After proton beam therapy (low-LET
radiation), the risk of secondary cancer induction, relative to photons, can be
divided by a factor of 3, due to the reduction of integral dose (as an average).
The RBE of heavy-ions for cancer induction can be assumed to be similar to
fission neutrons, i.e. about 20 relative to photons. However, after heavy-ion
beam therapy, the risk should be divided by 3, as after proton therapy due to
the excellent physical selectivity of the irradiation. Therefore a risk 5 to 10
times higher than photons could be assumed. This range is probably a pessimistic
estimate for carbon ions since most of the normal tissues, at the level of the
initial plateau, are irradiated with low-LET radiation. Probabilities for secondary cancer incidence have been estimated for a patient
with Hodgkin's disease for whom treatment has been planned with different
radiation modalities using photons and protons. The ICRP calculation scheme has
been used to calculate cancer incidence from dose distributions. For this
purpose, target volumes as well as critical structures have been outlined in the
CT set of a patient with Hodgkin's disease. Dose distributions have been
calculated using conventional as well as intensity-modulated treatment
techniques using photon and proton radiation. The cancer incidence has been
derived from the mean doses for each organ. The results of this work are: (a)
Intensity-modulated treatment of Hodgkin's disease using nine photon fields (15
MV) results in nearly the same cancer incidence as treating with two opposed
photon fields (6 MV). (b) Intensity-modulated treatment using nine proton fields
(maximum energy 177.25 MeV) results in nearly the same cancer incidence as
treating with one proton field (160 MeV). (c) Irradiation with protons using the
spot scanning technique decreases the avoidable cancer incidence compared to
photon treatment by a factor of about two. This result is independent of the
number of beams used. Our work suggests that there are radiotherapy indications
in which intensity-modulated treatments will result in little or no reduction of
cancer incidence compared to conventional treatments. However, proton treatment
can result in a lower cancer incidence than photon treatment. The purpose of this work was to compare the risk of developing a second cancer
after craniospinal irradiation using photon versus proton radiotherapy by means
of simulation studies designed to account for the effects of neutron exposures.
Craniospinal irradiation of a male phantom was calculated for
passively-scattered and scanned-beam proton treatment units. Organ doses were
estimated from treatment plans; for the proton treatments, the amount of stray
radiation was calculated separately using the Monte Carlo method. The organ
doses were converted to risk of cancer incidence using a standard formalism
developed for radiation protection purposes. The total lifetime risk of second
cancer due exclusively to stray radiation was 1.5% for the passively scattered
treatment versus 0.8% for the scanned proton beam treatment. Taking into account
the therapeutic and stray radiation fields, the risk of second cancer from
intensity-modulated radiation therapy and conventional radiotherapy photon
treatments were 7 and 12 times higher than the risk associated with scanned-beam
proton therapy, respectively, and 6 and 11 times higher than with passively
scattered proton therapy, respectively. Simulations revealed that both passively
scattered and scanned-beam proton therapies confer significantly lower risks of
second cancers than 6 MV conventional and intensity-modulated photon therapies. PURPOSE: To assess the risk of a secondary maligt neoplasm (SMN) from proton
therapy relative to intensity-modulated radiation therapy (IMRT) using X-rays,
taking into account contributions from both primary and secondary sources of
radiation, for prostate cancer.
METHODS AND MATERIALS: A proton therapy plan and a 6-MV IMRT plan were
constructed for 3 patients with early-stage adenocarcinoma of the prostate.
Doses from the primary fields delivered to organs at risk of developing an SMN
were determined from treatment plans. Secondary doses from the proton therapy
and IMRT were determined from Monte Carlo simulations and available measured
data, respectively. The risk of an SMN was estimated from primary and secondary
doses on an organ-by-organ basis by use of risk models from the Committee on the
Biological Effects of Ionizing Radiation.
RESULTS: Proton therapy reduced the risk of an SMN by 26% to 39% compared with
IMRT. The risk of an SMN for both modalities was greatest in the in-field
organs. However, the risks from the in-field organs were considerably lower with
the proton therapy plan than with the IMRT plan. This reduction was attributed
to the substantial sparing of the rectum and bladder from exposure to the
therapeutic beam by the proton therapy plan.
CONCLUSIONS: When considering exposure to primary and secondary radiation,
proton therapy can reduce the risk of an SMN in prostate patients compared with
contemporary IMRT. Proton beam therapy (PT) offers improved sparing of normal tissue, thus
potentially reducing the risk for treatment related late sequelae and induction
of secondary cancer. In addition, it can be an instrument for intensification of
local therapy ("dose escalation") for disease currently not sufficiently
controlled. Up to now, more than 70 000 patients have been treated with PT
worldwide. In particular, tumors of the ocular fundus, the base of skull and
prostate were targeted. In recent years an increasing number of children got
treated, predomitly when suffering from sarcomas or brain tumors. In Europe,
treatment was applied so far mainly in Switzerland (PSI) or France (Orsay).
However, availability of particle therapy is about to increase considerably
within the next years. The German Working Group of paediatric radiation oncology
APRO=Arbeitsgemeinschaft für Pädiatrische Radioonkologie) is currently trying to
embed PT in the multidisciplinary concepts of the GPOH, and PT experts are
appointed for each relevant study. In addition, prospective documentation of the
applied PT is to be performed according to the RISK protocol. Still, it is
mandatory to reach out for the integration of PT into our cooperative network
and the multidisciplinary trials. Best practice solutions have to be established
in order to provide high quality and transparency of any applied particle
therapy. INTRODUCTION: The concern of secondary cancer induction and normal tissue
complications have motivated a more frequent use of protons in radiotherapy (RT)
of children. However, proton RT is likely to be less robust to anatomical
changes occurring during therapy. In this study we present a recent clinical
case to illustrate this issue.
MATERIAL AND METHODS: A five-year-old boy with a highly proliferating maligt
intracranial nerve sheath tumour underwent a partial resection prior to RT and
developed a post-surgery oedema close to the surgical cavity. RT was delivered
with volumetric modulated arc therapy (VMAT) to a total tumour dose of 61.2 Gy.
The most critical organs at risk (ORs) were the right optical nerve, brainstem
and chiasm. Proton plans were constructed for the purpose of this study. In
order to simulate a worst-case scenario, the extent of the oedema observed in
the last part of the treatment was used to modify the oedema on the planning
computed tomography (CT). Both the photon and proton plans were then
re-calculated, as follows: Scenario A: Treatment planning based on the planning
CT with oedema and dose calculated as if it was delivered without oedema.
Scenario B: Treatment planning on the modified planning CT without oedema, but
re-calculated with oedema. These two scenarios were compared to the situation
where the oedema was present at treatment planning and unchanged during RT.
RESULTS: Total dose to critical ORs remained unchanged for the photon plans,
with changes within 0.3 Gy for the normal tissues and nearly identical target
coverage. For protons, scenario A led to increased maximal doses in all critical
ORs, 5.1 Gy in the brainstem, 6.1 Gy in the chiasm and 6.4 Gy in the right
optical nerve. For scenario B the proton plans resulted in a loss in target
coverage.
CONCLUSION: This case study shows that RT with protons were far less robust to
anatomical changes than when treated with photons, emphasising the increased
need for adaptive approaches in RT with protons. BACKGROUND: To compare proton beam therapy (PBT) and intensity-modulated
radiation therapy (IMRT) with conformal radiation therapy (CRT) in terms of
their organ doses and ability to cause secondary cancer in normal organs.
METHODS: Five patients (median age, 4 years; range, 2-11 years) who underwent
PBT for retroperitoneal neuroblastoma were selected for treatment planning
simulation. Four patients had stage 4 tumors and one had stage 2A tumor,
according to the International Neuroblastoma Staging System. Two patients
received 36 Gy, two received 21.6 Gy, and one received 41.4 Gy of radiation. The
volume structures of these patients were used for simulations of CRT and IMRT
treatment. Dose-volume analyses of liver, stomach, colon, small intestine,
pancreas, and bone were performed for the simulations. Secondary cancer risks in
these organs were calculated using the organ equivalent dose (OED) model, which
took into account the rates of cell killing, repopulation, and the neutron dose
from the treatment machine.
RESULTS: In all evaluated organs, the mean dose in PBT was 20-80% of that in
CRT. IMRT also showed lower mean doses than CRT for two organs (20% and 65%),
but higher mean doses for the other four organs (110-120%). The risk of
secondary cancer in PBT was 24-83% of that in CRT for five organs, but 121% of
that in CRT for pancreas. The risk of secondary cancer in IMRT was equal to or
higher than CRT for four organs (range 100-124%).
CONCLUSION: Low radiation doses in normal organs are more frequently observed in
PBT than in IMRT. Assessments of secondary cancer risk showed that PBT reduces
the risk of secondary cancer in most organs, whereas IMRT is associated with a
higher risk than CRT. We report a woman with maligt meningioma diagnosed 9 years after the
treatment of a choroidal melanoma with proton beam therapy. The risk of
secondary cancers is a well-known adverse late effect of radiation therapy,
especially with the use of advanced techniques such as intensity-modulated
radiation therapy. However, this risk may be less with the use of proton beam
therapy. A 79-year-old woman presented with symptoms of enophthalmos, ptosis and
paralysis of the left medial rectus muscle. She had previously been successfully
treated for a choroidal melanoma of the left eye with proton beam therapy (total
dose: 60 cobalt gray equivalents) following local resection. MRI showed a lesion
in the left cavernous sinus with extension into the orbit and a subsequent
biopsy revealed a papillary meningioma. The cavernous tumor was treated with
photon radiotherapy (total dose: 54Gy) which achieved an initial partial
response. However, 8 months later the tumor extensively metastasized to the
skull and the spine and the patient died 1 year after the treatment. The
incidence of secondary maligcies after proton beam therapy is low but not
negligible, therefore, it must be taken into account when planning a treatment
as secondary tumors may present with a highly aggressive behaviour. Radiation-induced secondary maligcies are a significant, yet uncommon cause
of morbidity and mortality among cancer survivors. Secondary maligcy risk is
dependent upon multiple factors including patient age, the biological and
genetic predisposition of the individual, the volume and location of tissue
irradiated, and the dose of radiation received. Proton therapy (PRT) is an
advanced particle therapy with unique dosimetric properties resulting in reduced
entrance dose and minimal to no exit dose when compared with standard photon
radiation therapy. Multiple dosimetric studies in varying cancer subtypes have
demonstrated that PRT enables the delivery of adequate target volume coverage
with reduced integral dose delivered to surrounding tissues, and modeling
studies taking into account dosimetry and radiation cell biology have estimated
a significantly reduced risk of radiation-induced secondary maligcy with PRT.
Clinical data are emerging supporting the lower incidence of secondary
maligcies after PRT compared with historical photon data, though longer
follow-up in proton treated cohorts is awaited. This article reviews the current
dosimetric and clinical literature evaluating the incidence of and risk factors
associated with radiation-induced secondary maligcy following PRT. Recent progress in the treatment for pediatric maligcies using a combination
of surgery, chemotherapy, and radiotherapy has improved survival. However, late
toxicities of radiotherapy are a concern in long-term survivors. A recent study
suggested reduced secondary cancer and other late toxicities after proton beam
therapy (PBT) due to dosimetric advantages. In this study, we evaluated the
safety and efficacy of PBT for pediatric patients treated in Japan. A
retrospective observational study in pediatric patients who received PBT was
performed. All patients aged <20 years old who underwent PBT from January 1983
to August 2014 at four sites in Japan were enrolled in the study. There were 343
patients in the study. The median follow-up periods were 22.6 months
(0.4-374.3 months) for all patients and 30.6 months (0.6-374.3 months) for
survivors. The estimated 1-, 3-, 5-, and 10-year survival rates were 82.7% (95%
CI: 78.5-87.0%), 67.4% (61.7-73.2%), 61.4% (54.8-67.9%), and 58.7% (51.5-65.9%),
respectively. Fifty-two events of toxicity ≥ grade 2 occurred in 43 patients.
Grade 4 toxicities of myelitis, visual loss (two cases), cerebral vascular
disease, and tissue necrosis occurred in five patients. This study provides
preliminary results for PBT in pediatric patients in Japan. More experience and
follow-up with this technique are required to establish the efficacy of PBT in
this patient population. The number of patients treated by proton beam therapy in Japan since 2000 has
increased; in 2016, 11 proton facilities were available to treat patients.
Notably, proton beam therapy is very useful for pediatric cancer; since the
pediatric radiation dose to normal tissues should be reduced as much as possible
because of the effect of radiation on growth, intellectual development,
endocrine organ function and secondary cancer development. Hepatocellular
carcinoma is common in Asia, and most of the studies of proton beam therapy for
liver cancer have been reported by Japanese investigators. Proton beam therapy
is also a standard treatment for nasal and paranasal lesions and lesions at the
base of the skull, because the radiation dose to critical organs such as the
eyes, optic nerves and central nervous system can be reduced with proton beam
therapy. For prostate cancer, comparative studies that address adverse effects,
safety, patient quality of life and socioeconomic issues should be performed to
determine the appropriate use of proton beam therapy for prostate cancer.
Regarding new proton beam therapy applications, experience with proton beam
therapy combined with chemotherapy is limited, although favorable outcomes have
been recently reported for locally advanced lung cancer, esophageal cancer and
pancreatic cancer. Therefore, 'chemoproton' therapy appears to be a very
attractive field for further clinical investigations. In conclusion, there are
cost issues and considerations regarding national insurance for the use of
proton beam therapy in Japan. Further studies and discussions are needed to
address the use of proton beam therapy for several types of cancers, and for
maintaining the quality of life of patients while retaining a high cure rate. To investigate the amount that radiation-induced secondary cancer would be
reduced by using proton beam therapy (PBT) in place of intensity-modulated X-ray
therapy (IMXT) in pediatric patients, we analyzed lifetime attributable risk
(LAR) as an in silico surrogate marker of the secondary cancer after these
treatments. From 242 pediatric patients with cancers who were treated with PBT,
26 patients were selected by random sampling after stratification into four
categories: (i) brain, head and neck, (ii) thoracic, (iii) abdominal, and (iv)
whole craniospinal (WCNS) irradiation. IMXT was replanned using the same
computed tomography and region of interest. Using the dose-volume histograms
(DVHs) of PBT and IMXT, the LARs of Schneider et al. were calculated for the
same patient. All the published dose-response models were tested for the organs
at risk. Calculation of the LARs of PBT and IMXT based on the DVHs was feasible
for all patients. The means ± standard deviations of the cumulative LAR
difference between PBT and IMXT for the four categories were (i) 1.02 ± 0.52%
(n = 7, P = 0.0021), (ii) 23.3 ± 17.2% (n = 8, P = 0.0065), (iii) 16.6 ± 19.9%
(n = 8, P = 0.0497) and (iv) 50.0 ± 21.1% (n = 3, P = 0.0274), respectively (one
tailed t-test). The numbers needed to treat (NNT) were (i) 98.0, (ii) 4.3, (iii)
6.0 and (iv) 2.0 for WCNS, respectively. In pediatric patients who had undergone
PBT, the LAR of PBT was significantly lower than the LAR of IMXT estimated by in
silico modeling. Although a validation study is required, it is suggested that
the LAR would be useful as an in silico surrogate marker of secondary cancer
induced by different radiotherapy techniques. Proton beam therapy (PBT) is a potential new alternative to treatment with
photon radiotherapy that may reduce the risk of late toxicity and secondary
cancer, especially for pediatric tumors. The goal of this study was to evaluate
the long-term benefits of PBT in cancer survivors. A retrospective observational
study of pediatric patients who received PBT was performed at four institutions
in Japan. Of 343 patients, 62 were followed up for 5 or more years. These
patients included 40 males and 22 females, and had a median age of 10 years
(range: 0-19 years) at the time of treatment. The irradiation dose ranged from
10.8 to 81.2 GyE (median: 50.4 GyE). The median follow-up period was 8.1 years
(5.0-31.2 years). The 5-, 10- and 20-year rates for grade 2 or higher late
toxicities were 18%, 35% and 45%, respectively, and those for grade 3 or higher
late toxicities were 6%, 17% and 17% respectively. Univariate analysis showed
that the irradiated site (head and neck, brain) was significantly associated
with late toxicities. No maligt secondary tumors occurred within the
irradiated field. The 10- and 20-year cumulative rates for all secondary tumors,
maligt secondary tumors, and maligt nonhematologic secondary tumors were
8% and 16%, 5% and 13%, and 3% and 11%, respectively. Our data indicate that PBT
has the potential to reduce the risk of late mortality and secondary maligcy.
Longer follow-up is needed to confirm the benefits of PBT for pediatric tumors. INTRODUCTION: The potential of proton therapy to improve the sparing of the
healthy tissue has been demonstrated in several studies. However, even small
doses delivered to the organs at risk (OAR) may induce long-term detriments
after radiotherapy. In this study, we investigated the possibility to reduce the
risk of radiation-induced secondary cancers with intensity modulated proton
therapy (IMPT), when used for radiosurgery of liver metastases.
MATERIAL AND METHODS: Ten patients, previously treated for liver metastases with
photon-beam based stereotactic body radiation therapy (SBRT) were
retrospectively planned for radiosurgery with IMPT. A treatment plan comparison
was then performed in terms of calculated risk of radiation-induced secondary
cancer. The risks were estimated using two distinct models (Dasu et al., 2005;
Schneider et al., 2005, 2009). The plans were compared pairwise with a two-sided
Wilcoxon signed-rank test with a significance level of 0.05.
RESULTS: Reduced risks for induction of fatal and other types of cancers were
estimated for the IMPT plans (p<0.05) with the Dasu et al.
MODEL: Using the Schneider et al. model, lower risks for carcinoma-induction
with IMPT were estimated for the skin, lungs, healthy part of the liver,
esophagus and the remaining part of the body (p<0.05). The risk of observing
sarcomas in the bone was also reduced with IMPT (p<0.05).
CONCLUSION: The findings of this study indicate that the risks of
radiation-induced secondary cancers after radiosurgery of liver metastases may
be reduced, if IMPT is used instead of photon-beam based SBRT. Cancer is a major cause of childhood death, with central nervous system (CNS)
neoplasms being the second most common pediatric maligcy, following
hematological cancer. Treatment of pediatric CNS maligcies requires
multimodal treatment using a combination of surgery, chemotherapy, and
radiotherapy, and advances in these treatments have given favorable results and
longer survival. However, treatment-related toxicities have also occurred,
particularly for radiotherapy, after which secondary cancer, reduced function of
irradiated organs, and retarded growth are significant problems. Proton beam
therapy (PBT) is a particle radiotherapy with excellent dose localization that
permits treatment of liver and lung cancer by administration of a high dose to
the tumor while minimizing damage to surrounding normal tissues. Thus, PBT has
the potential advantages for pediatric cancer. In this context, we review the
current knowledge on PBT for treatment of pediatric CNS maligcies. In the past few years, proton therapy has taken the centre stage in treating
various tumour types. The primary contribution of this study is to investigate
the tumour control probability (TCP), relapse time and the corresponding
secondary cancer risks induced by proton beam radiation therapy. We incorporate
tumour relapse kinetics into the TCP framework and calculate the associated
second cancer risks. To calculate proton therapy-induced secondary cancer
induction, we used the well-known biologically motivated mathematical model,
initiation-inactivation-proliferation formalism. We used the available in vitro
data for the linear energy transfer (LET) dependence of cell killing and
mutation induction parameters. We evaluated the TCP and radiation-induced second
cancer risks for protons in the clinical range of LETs, i.e. approximately 8
$\mathrm{keV/\mu m}$ for the tumour volume and 1-3 $\mathrm{keV/\mu m}$ for the
organs at risk. This study may serve as a framework for further work in this
field and elucidates proton-induced TCP and the associated secondary cancer
risks, not previously reported in the literature. Although studies with a
greater number of cell lines would reduce uncertainties within the model
parameters, we argue that the theoretical framework presented within is a
sufficient rationale to assess proton radiation TCP, relapse and carcinogenic
effects in various treatment plans. We show that compared with photon therapy,
proton therapy markedly reduces the risk of secondary maligcies and for
equivalent dosing regimens achieves better tumour control as well as a reduced
primary recurrence outcome, especially within a hypo-fractionated regimen. |
List the essential aminoacids. | Leucine
Isoleucine
Valine
Tryptophan
Cysteine
Methionine
Lysine
Phenylalanine | The effects on growth, body composition, and metabolism of a protein-restricted
diet supplemented with essential aminoacids, the calcium-ketoacids of valine,
leucine, isoleucine, and phenylalanine, and the calcium-hydroxyacid of
methionine, were investigated in seven growth-retarded children with chronic
renal failure. During 0.4 to 1.0 years of treatment there were significant
increases in growth velocity and upper arm circumference SD scores, body cell
mass (intracellular water calculated as tritium space minus corrected sodium
bromide space) and serum transferrin. Blood urea and urea:creatinine ratio fell
in all children. Renal function assessed from plasma creatinine and 51Cr EDTA
clearance did not change significantly. During treatment there was an increase
in plasma calcium, high levels necessitating brief interruption of therapy in
two children, and a decrease in plasma phosphate. Levels of serum parathormone
fell in all children, and were correlated inversely with plasma calcium and
positively with plasma phosphate. Abnormalities compared to control children in
blood aminoacid and branched-chain ketoacid levels were unaffected by treatment.
These results suggest that a protein-restricted diet supplemented with essential
aminoacids and calcium-keto and hydroxyacids may be useful to improve linear
growth and nutritional status in children with chronic renal failure, and that a
reduction in hyperparathyroidism may be partly responsible for some of the
beneficial effects observed. BACKGROUND: Plant food lacks vitamin B12, vitamin D and higher n-3
polyunsaturated fatty acids. Essential aminoacids methionine and lysine can be
found in significantly lower amounts. On the contrary, the culinary and
technologically non-processed plant food and whole-grain products contain
essential nutrients in a highly condensed form. The aim of the study was to
compare nutritional status of adults on alternative or on traditional diet and
sequels of the diet to body metabolism.
METHODS AND RESULTS: The group on alternative diet consisted of 89
lacto-ovo-vegetarians (age 38.7 +/- 0.6 years, average duration of vegetarianism
7.8 years). Control group on traditional diet (omnivores, n = 84) was formed as
an average sample. Nutritional regime was determined using dietetic
questionnaire on the food intake regularity. Vegetarians consume optimal amount
of fat (along with recommendations of OVD) with predomice of vegetal lipids.
They have low intake of cholesterol (62.8 mg), recommended ratio of saturated
fatty acids (SFA), monounsaturated (MUFA), and polyunsaturated (PUFA)
6.5:10.6:8.9 energetic percent. Their ratio of linolic:alpha-linolenic acid
(10.4:1) also corresponds with recommendations. In traditional diet, the content
of lipids and energy usually exceeds the norm of OVD (by 33% or 19%
respectively), cholesterol intake is much higher (512.2 mg, 200 mg is
recommended as a maximum). Higher is the amount taken of SFA (11.2 energetic %,
recommended 7%), and not sufficient is the intake of alpha-linolenic acid (68%
of OVD). People on alternative diet have low plasma levels of risk lipid
parameters and significantly higher levels of antisclerotic substances. As a
result of significantly higher intake of fruits and vegetables, plant oil,
sprouts, seeds, and whole-grain food the plasma levels of antioxidative vitamins
are in vegetarians higher then threshold. It reduces the risk of the
free-radical disease. On the contrary, vegetarians have deficits in methionine
intake, and 15% of them have hypoproteinemia is (0% in omnivores). Low plasma
levels of iron and calcium, occurrence of hyposideremia (16% versus 2%) and
hypocalcemia (21% versus 8%) corresponds with intake of vegetal absorption
inhibitors (fytolic acid, oxalic acid, roughage). Frequently a mild form of
hyperhomocysteinemia is found (28% versus 5%), resulting vitamin B12 deficit.
CONCLUSIONS: Vegetarian diet is optimal for prevention of free-radical diseases,
especially those of the cardiovascular system. It may bring a risk from the
point of view of low iron and calcium absorption, low intake of methionine and
occurrence of mild forms of hyperhomocysteinemia. In traditional diet, total
lipid content should be lowered, amount of vegetable oil with alpha-linolenic
acid should be elevated as well as fruit and vegetable consummation. Whole grain
food and oily seeds should be included into the daily food. Alpha-Lactalbumin is the main whey protein in human milk rising 2,44 g/L in
mature milk. It has a key function in the synthesis of lactose from glucose and
galactose in the mammary gland although this compound has also other beneficial
effects on the infant health due to the high proportion of essential aminoacids
(tryptophan and cysteine). It seems also to increase iron absorption in the
digestive track, and in in vitro experiments, linked to oleic acid (HAMLET
complex), has shown anticarcinogenic effects against cellular tumor such as
human papilloma. In addition, this complex has been reported to exhibit
antimicrobial properties against Streptococcus pneumoniae, Haemophilus
influenzae, enteropathogenic strains of Escherichia coli and Salmonella
thypimurium. However, the in vivo synthesis of HAMLET complex during milk
digestion has not been proved yet. Infant formula have been improved
considerably during the last decades not only adapting nutrient concentrations
to infants requirements but also by the addition of new bioactive ingredients
such as alpha-lactalbumin, to have the same functional effect as in breast fed
babies. Author information:
(1)Renal, Vascular and Diabetes Research Laboratory, Instituto de Investigación
Sanitaria Fundación Jiménez Díaz, Universidad Autónoma de Madrid, Spain and
Spanish Biomedical Research Centre in Diabetes and Associated Metabolic
Disorders (CIBERDEM), Spain. Electronic address: [email protected].
(2)Renal, Vascular and Diabetes Research Laboratory, Instituto de Investigación
Sanitaria Fundación Jiménez Díaz, Universidad Autónoma de Madrid, Spain and
Spanish Biomedical Research Centre in Diabetes and Associated Metabolic
Disorders (CIBERDEM), Spain. Electronic address: [email protected].
(3)Renal, Vascular and Diabetes Research Laboratory, Instituto de Investigación
Sanitaria Fundación Jiménez Díaz, Universidad Autónoma de Madrid, Spain and
Spanish Biomedical Research Centre in Diabetes and Associated Metabolic
Disorders (CIBERDEM), Spain. Electronic address: [email protected].
(4)Division of Hematology, Fundación Jiménez Díaz, Madrid, Spain. Electronic
address: [email protected].
(5)Division of Endocrinology, Fundación Jiménez Díaz, Madrid, Spain. Electronic
address: [email protected].
(6)Department of Pharmacology, Faculty of Medicine, Universidad Autónoma de
Madrid, Spain. Electronic address: [email protected].
(7)Renal, Vascular and Diabetes Research Laboratory, Instituto de Investigación
Sanitaria Fundación Jiménez Díaz, Universidad Autónoma de Madrid, Spain and
Spanish Biomedical Research Centre in Diabetes and Associated Metabolic
Disorders (CIBERDEM), Spain. Electronic address: [email protected].
(8)Renal, Vascular and Diabetes Research Laboratory, Instituto de Investigación
Sanitaria Fundación Jiménez Díaz, Universidad Autónoma de Madrid, Spain and
Spanish Biomedical Research Centre in Diabetes and Associated Metabolic
Disorders (CIBERDEM), Spain. Electronic address: [email protected]. |
What does Retapamulin treat? | Retapamulin is a small molecule covalently binding and inhibiting the bacterium Staphylococcus aureus (MRSA). | Retapamulin is a semisynthetic pleuromutilin derivative being developed as a
topical antibiotic for treating bacterial infections of the skin. It is potent
in vitro against susceptible and multidrug-resistant organisms commonly
associated with bacterial skin infections. We report detailed mode of action
studies demonstrating that retapamulin binds to the bacterial ribosome with high
affinity, inhibits ribosomal peptidyl transferase activity, and partially
inhibits the binding of the initiator tRNA substrate to the ribosomal P-site.
Taken together, these data distinguish the mode of action of retapamulin from
that of other classes of antibiotics. This unique mode of action may explain the
lack of clinically relevant, target-specific cross-resistance of retapamulin
with antibacterials in current use. INTRODUCTION: Retapamulin is a novel, topical antibacterial of the pleuromutilin
class in development for the treatment of secondarily infected traumatic lesions
of the skin.
METHODS: The efficacy, safety, and tolerability of topical retapamulin ointment,
1% for 5 days twice daily was evaluated in 2 identical, randomized,
double-blind, double-dummy, multicenter studies vs oral cephalexin, 500 mg twice
daily for 10 days, in 1904 patients with secondarily infected traumatic lesions.
RESULTS: Clinical success rates were 89.5% in protocol-adherent patients
receiving retapamulin compared with 91.9% for cephalexin (treatment difference,
-2.5% [95% confidence interval, -5.4% to 0.5%]). In patients with Staphylococcus
aureus or Streptococcus pyogenes at baseline, clinical success was 89.2%
(365/409) for retapamulin and 92.6% (63/68) for cephalexin. Safety and
tolerability were similar between treatments. Noncompliance (defined as using or
taking <80% of doses) was recorded in 8.0% (51/636) of patients taking
cephalexin compared with 0.39% (5/1268) of patients receiving retapamulin.
CONCLUSIONS: Retapamulin offers a novel, effective, and convenient topical
treatment for secondarily infected traumatic lesions. OBJECTIVES: Retapamulin is the first agent of the pleuromutilin class formulated
as a topical antibacterial for treating skin infections. The aim of this study
was to determine the antimicrobial activity of retapamulin by determining the
minimal inhibitory concentration (MIC) values of this new drug and comparators
against a wide range of anaerobic bacteria of human origin.
METHODS: The in vitro activity of retapamulin and six comparators (amoxicillin,
amoxicillin/clavulanic acid, ceftriaxone, imipenem, clindamycin and
metronidazole) was evaluated against 232 anaerobic clinical isolates. MICs were
determined by the CLSI reference agar dilution method (M11-A6).
RESULTS: Ceftriaxone, clindamycin and amoxicillin/clavulanic acid resistance
rates were 54%, 42% and 9.6%, respectively, within the Bacteroides fragilis
group. Despite high resistance rates to various antibiotics, retapamulin
inhibited 37/52 (71%) strains of the B. fragilis group and 85/87 (98%) of the
other Gram-negative bacilli at a concentration of 2 mg/L or less. All the
investigated strains of Clostridium perfringens were inhibited by 1 mg/L
retapamulin. Three strains of C. difficile and one strain of C. clostridioforme
demonstrated decreased susceptibility to retapamulin. Based on inhibitory
concentrations, retapamulin was more active than clindamycin, metronidazole and
ceftriaxone against Propionibacterium acnes and anaerobic Gram-positive cocci,
as all isolates were inhibited by <or=2 mg/L.
CONCLUSIONS: At <or=2 mg/L, retapamulin inhibited 90% of all 232 anaerobes
tested, whereas overall resistance rates for the comparators were as follows:
co-amoxiclav, 2%; metronidazole, 12%; clindamycin, 15% and ceftriaxone, 20%. The
broad anaerobic spectrum demonstrated by retapamulin in vitro is attractive.
Pending further clinical investigation, retapamulin may offer an alternative
treatment for anaerobic skin infections in this era of increasing resistance. Retapamulin is a semisynthetic pleuromutilin compound with in vitroactivity
against Gram-positive bacteria, no cross-resistance to other classes of
antimicrobial agents in current use and a low potential for development of
resistance. A 1% ointment formulation has been developed for clinical use, and a
placebo-controlled trial of impetigo in 210 patients produced significantly
higher rates of clinical and microbiological success compared with placebo -
85.6 versus 52.1% and 91.2 versus 50.9%, respectively. Additional comparative
studies in over 1900 patients showed noninferiority to topical fusidic acid and
oral cephalexin and a low frequency of adverse events. In 2007, retapamulin was
approved in the USA for topical treatment of impetigo caused by Streptococcus
pyogenes and methicillin-susceptible Staphylococcus aureus, and in the EU for
topical treatment of impetigo and infected wounds caused by S. pyogenes and S.
aureus, with approvals including adults and children over 9 months of age. Retapamulin is the first agent in the new pleuromutilin class of antibacterials
to become commercially available for clinical use in humans. Retapamulin acts as
a potent inhibitor of bacterial protein synthesis and has a unique mode of
antibiotic action. To date, retapamulin has not demonstrated any clinically
relevant, target-specific crossresistance with other antibiotic classes, and has
shown a low potential for resistance selection in vitro. In preclinical studies,
retapamulin demonstrated pronounced in vitro activity against staphylococcal,
streptococcal and anaerobic Gram-positive clinical isolates associated with skin
and skin structure infections. Clinical pharmacology studies showed low systemic
exposure with topical use of retapamulin, and a favorable tolerability profile.
In clinical efficacy trials involving pediatric and adult patients who received
retapamulin twice daily for five days, retapamulin was highly effective in the
treatment of impetigo, secondarily infected traumatic lesions and secondarily
infected dermatitis. Further, the clinical efficacy and safety profile of
retapamulin was comparable to that of commonly used oral and topical
antibiotics. Retapamulin was also clinically effective against isolates
resistant to existing therapies. As a 1% ointment, retapamulin has been approved
in the United States for the treatment of impetigo and in Europe for the
shortterm treatment of impetigo and infected small lacerations, abrasions and
sutured wounds. Topical retapamulin (Altabax, Altargo) is the first pleuromutilin antibacterial
approved for the treatment of uncomplicated superficial skin infections caused
by Staphylococcus aureus (excluding meticillin-resistant S. aureus [MRSA]) and
Streptococcus pyogenes in patients aged > or = 9 months. In the EU, retapamulin
is indicated for use in patients with impetigo or with infected small
lacerations, abrasions or sutured wounds (without abscesses); in the US, it is
indicated for use in patients with impetigo. Retapamulin has a novel site of
action on bacterial ribosomes. In clinical trials in patients with impetigo,
topical retapamulin 1% ointment twice daily for 5 days (the approved regimen)
was superior to placebo; treatment with retapamulin was noninferior to that with
topical fusidic acid. In patients with secondarily infected traumatic lesions,
treatment with retapamulin was noninferior to that with oral cefalexin, although
the efficacy of retapamulin was reduced in patients with MRSA infections or
superficial abscesses. Retapamulin was well tolerated in both paediatric and
adult patients, and the majority of adverse events were of mild to moderate
severity. Thus, the introduction of topical retapamulin 1% ointment extends the
treatment options available in the management of impetigo and uncomplicated
secondarily infected traumatic lesions. Retapamulin is a new topical pleuromutilin antibiotic for the treatment of skin
and skin-structure infections, including impetigo. In vitro studies indicate
that retapamulin has a unique mode of action that minimizes the potential for
target-specific cross-resistance with other antibacterials and a limited
potential for resistance development. Its spectrum of activity includes the most
likely causative pathogens Staphylococcus aureus and Streptococcus pyogenes. In
the Global Surveillance Program, retapamulin was highly active in vitro,
including against strains of S. aureus resistant to methicillin, mupirocin or
fusidic acid. In clinical studies, retapamulin was noninferior to fusidic acid
and oral cefalexin, achieving per-pathogen success rates of 86-99%. Topical
retapamulin has a good safety profile and is associated with high patient
compliance. In atopic dermatitis (AD), the stratum corneum of patients appears to have
alterations that predispose them to colonization and invasion by various
bacteria, most notably Staphylococcus aureus (S. aureus). This bacterial
co-existence is accepted to be an important factor in AD disease activity.
Exactly when to initiate antimicrobial treatment is controversial, but such
intervention, when warranted, has repeatedly been demonstrated to improve the
course of AD. However, the increase in antibiotic resistance presents a
therapeutic challenge in the management of AD patients, which highlights the
need for novel mechanism topical antibacterial agents. Retapamulin is a
relatively new pleuromutilin antibiotic designed for topical use. In vitro
studies have demonstrated its low potential for the development of antibacterial
resistance and high degree of potency against Gram-positive bacteria found in
skin infections, including many S. aureus strains that are resistant to
methicillin, fusidic acid, and mupirocin. Clinical studies exploring the
treatment of secondarily infected dermatitis reveal that the efficacy of topical
retapamulin is comparable to a 10-day course of oral cephalexin or to topical
fusidic acid. Retapamulin appears to be a much needed antimicrobial option for
treating the AD population due to their common carriage of bacterial pathogens
and frequency of infectious complications. Staphylococcus aureus skin infections represent a significant public health
threat because of the emergence of antibiotic-resistant strains such as
methicillin-resistant S. aureus (MRSA). As greater understanding of protective
immune responses and more effective antimicrobial therapies are needed, a S.
aureus skin wound infection model was developed in which full-thickness scalpel
cuts on the backs of mice were infected with a bioluminescent S. aureus
(methicillin sensitive) or USA300 community-acquired MRSA strain and in vivo
imaging was used to noninvasively monitor the bacterial burden. In addition, the
infection-induced inflammatory response was quantified using in vivo
fluorescence imaging of LysEGFP mice. Using this model, we found that both IL-1α
and IL-1β contributed to host defense during a wound infection, whereas IL-1β
was more critical during an intradermal S. aureus infection. Furthermore,
treatment of a USA300 MRSA skin infection with retapamulin ointment resulted in
up to 85-fold reduction in bacterial burden and a 53% decrease in
infection-induced inflammation. In contrast, mupirocin ointment had minimal
clinical activity against this USA300 strain, resulting in only a 2-fold
reduction in bacterial burden. Taken together, this S. aureus wound infection
model provides a valuable preclinical screening method to investigate cutaneous
immune responses and the efficacy of topical antimicrobial therapies. Impetigo is a common childhood skin infection. There are reports of increasing
drug resistance to the currently used topical antibiotics including fusidic acid
and mupirocin. Retapamulin is a newer topical agent of pleuromutilin class
approved by the Food and Drug Administration for treatment of impetigo in
children and has been recently made available in the Indian market. It has been
demonstrated to have low potential for the development of antibacterial
resistance and a high degree of potency against poly drug resistant
Gram-positive bacteria found in skin infections including Staphylococcus aureus
strains. The drug is safe owing to low systemic absorption and has only minimal
side-effect of local irritation at the site of application. INTRODUCTION: Retapamulin, a topical pleuromutilin that selectively inhibits
bacterial protein synthesis, is approved for treatment of impetigo and
secondarily infected traumatic lesions in adults and in children older than 9
months of age.
OBJECTIVE: A 5-year study was conducted to monitor prescription use in children
younger than 9 months of age.
METHODS: Annual prescription events were monitored in the UK Clinical Practice
Research Datalink (CPRD) and the Clinformatics™ DataMart Multiplan (IMPACT), a
product of OptumInsight Life Sciences, Inc. (Eden Prairie, MN, USA), from the
USA.
RESULTS: In the CPRD, of 148 prescriptions, three (2 %) were identified in
children aged less than 9 months between years 2008 and 2011. In IMPACT, of
59,210 claims for retapamulin in children, 1,951 (3.3 %) were categorized as
definitive, or uncertain for, less than 9 months of age between 2007 and 2011.
CONCLUSION: Retapamulin prescription events in children aged less than 9 months
were relatively low compared with other recent estimations of off-label
pediatric medicines. Our report provides a framework for future investigations
and discussions that may facilitate off-label reporting schemes and promote
pediatric drug safety. Impetigo is the most common bacterial skin infection in children two to five
years of age. There are two principal types: nonbullous (70% of cases) and
bullous (30% of cases). Nonbullous impetigo, or impetigo contagiosa, is caused
by Staphylococcus aureus or Streptococcus pyogenes, and is characterized by
honey-colored crusts on the face and extremities. Impetigo primarily affects the
skin or secondarily infects insect bites, eczema, or herpetic lesions. Bullous
impetigo, which is caused exclusively by S. aureus, results in large, flaccid
bullae and is more likely to affect intertriginous areas. Both types usually
resolve within two to three weeks without scarring, and complications are rare,
with the most serious being poststreptococcal glomerulonephritis. Treatment
includes topical antibiotics such as mupirocin, retapamulin, and fusidic acid.
Oral antibiotic therapy can be used for impetigo with large bullae or when
topical therapy is impractical. Amoxicillin/clavulanate, dicloxacillin,
cephalexin, clindamycin, doxycycline, minocycline,
trimethoprim/sulfamethoxazole, and macrolides are options, but penicillin is
not. Natural therapies such as tea tree oil; olive, garlic, and coconut oils;
and Manuka honey have been anecdotally successful, but lack sufficient evidence
to recommend or dismiss them as treatment options. Treatments under development
include minocycline foam and Ozenoxacin, a topical quinolone. Topical
disinfectants are inferior to antibiotics and should not be used. Empiric
treatment considerations have changed with the increasing prevalence of
antibiotic-resistant bacteria, with methicillin-resistant S. aureus,
macrolide-resistant streptococcus, and mupirocin-resistant streptococcus all
documented. Fusidic acid, mupirocin, and retapamulin cover
methicillin-susceptible S. aureus and streptococcal infections. Clindamycin
proves helpful in suspected methicillin-resistant S. aureus infections.
Trimethoprim/sulfamethoxazole covers methicillin-resistant S. aureus infection,
but is inadequate for streptococcal infection. BACKGROUND: Cutaneous bacterial infections are common in children and adults and
frequently are caused by Staphylococcus aureus (S. aureus). Treatment failures
with topical agents are not uncommon and have been shown to be secondary to
bacterial resistance.
OBJECTIVE: To determine clinical and bacteriological efficacy of retapamulin
ointment 1% in treatment of patients with cutaneous bacterial infections caused
by methicillin-resistant S. aureus (MRSA) and other bacteria.
METHODS: Prospective, nonrandomized, uncontrolled, open label, single center
trial conducted between April 2008 and November 2012 that evaluated efficacy of
retapamulin ointment 1% in the treatment of impetigo, folliculitis, and other
minor soft tissue infections in children and adults. Fifty patients, who
presented to a dermatology outpatient clinic and were clinically diagnosed with
impetigo, folliculitis, or minor soft tissue infection suitable for treatment
with a topical antibiotic, were screened. Thirty-eight patients were enrolled
and received treatment: topical retapamulin ointment 1% twice daily for 5 days.
Seven patients were MRSA positive and qualified for the primary efficacy
population. One patient withdrew due to an adverse event. Clinical and
microbiological exams were performed at baseline and follow-up 5 to 7 days later
to assess clinical, microbiological, and therapeutic responses. Primary outcome
was clinical response at follow-up in primary efficacy population with MRSA
isolated as the baseline pathogen. Secondary outcomes included clinical,
microbiologic, and therapeutic responses in patients who were culture positive
for any species of bacteria.
RESULTS: Clinical response at follow-up in the primary efficacy population
(MRSA-positive patients) was not sufficiently powered to demonstrate
significance; however, outcomes were excellent, with 7 of 7 patients
demonstrating clinical success (5 of 7) or clinical improvement (2 of 7) at
follow-up. Barring lack of significance due to small total sample size for
patients who were culture positive for any species of bacteria (n = 35), overall
success rates were favorable for clinical, microbiologic, and therapeutic
responses with values of 66%, 97%, and 69%, respectively. Adverse events (AEs)
were mild or moderate in severity. No serious AEs were reported.
CONCLUSION: Safety profile appears favorable given the low number of AEs. Study
design limits conclusions that can be drawn. Nevertheless, this study supports
use of topical retapamulin 1% ointment in treatment of cutaneous bacterial
infections, particularly those caused by S. aureus, including MRSA. Invasive infections due to Staphylococcus aureus, including
methicillin-resistant S. aureus are prevalent and life-threatening. Combinations
of antibiotic therapy have been employed in many clinical settings for improving
therapeutic efficacy, reducing side effects of drugs, and development of
antibiotic resistance. Pleuromutilins have a potential to be developed as a new
class of antibiotics for systemic use in humans. In the current study, we
investigated the relationship between pleuromutilins, including valnemulin,
tiamulin, and retapamulin, and 13 other antibiotics representing different
mechanisms of action, against methicillin-susceptible and -resistant S. aureus
both in vitro and in an experimental Galleria mellonella model. In vitro
synergistic effects were observed in combination of all three study
pleuromutilins with tetracycline (TET) by standard checkerboard and/or time-kill
assays. In addition, the combination of pleuromutilins with ciprofloxacin or
enrofloxacin showed antagonistic effects, while the rest combinations presented
indifferent effects. Importantly, all study pleuromutilins in combination with
TET significantly enhanced survival rates as compared to the single drug
treatment in the G. mellonella model caused by S. aureus strains. Taken
together, these results demonstrated synergy effects between pleuromutilins and
TET against S. aureus both in vitro and in vivo. |
Which histone mark distinguishes active from inactive enhancers? | Histone H3K27ac separates active from poised enhancers and predicts developmental state . In contrast, elements of the second class 'poised enhancers' are linked to genes inactive in hESCs . They are involved in orchestrating early steps in embryogenesis, such as gastrulation, mesoderm formation and neurulation . | Developmental programs are controlled by transcription factors and chromatin
regulators, which maintain specific gene expression programs through epigenetic
modification of the genome. These regulatory events at enhancers contribute to
the specific gene expression programs that determine cell state and the
potential for differentiation into new cell types. Although enhancer elements
are known to be associated with certain histone modifications and transcription
factors, the relationship of these modifications to gene expression and
developmental state has not been clearly defined. Here we interrogate the
epigenetic landscape of enhancer elements in embryonic stem cells and several
adult tissues in the mouse. We find that histone H3K27ac distinguishes active
enhancers from inactive/poised enhancer elements containing H3K4me1 alone. This
indicates that the amount of actively used enhancers is lower than previously
anticipated. Furthermore, poised enhancer networks provide clues to unrealized
developmental programs. Finally, we show that enhancers are reset during nuclear
reprogramming. Cell-fate transitions involve the integration of genomic information encoded by
regulatory elements, such as enhancers, with the cellular environment. However,
identification of genomic sequences that control human embryonic development
represents a formidable challenge. Here we show that in human embryonic stem
cells (hESCs), unique chromatin signatures identify two distinct classes of
genomic elements, both of which are marked by the presence of chromatin
regulators p300 and BRG1, monomethylation of histone H3 at lysine 4 (H3K4me1),
and low nucleosomal density. In addition, elements of the first class are
distinguished by the acetylation of histone H3 at lysine 27 (H3K27ac), overlap
with previously characterized hESC enhancers, and are located proximally to
genes expressed in hESCs and the epiblast. In contrast, elements of the second
class, which we term 'poised enhancers', are distinguished by the absence of
H3K27ac, enrichment of histone H3 lysine 27 trimethylation (H3K27me3), and are
linked to genes inactive in hESCs and instead are involved in orchestrating
early steps in embryogenesis, such as gastrulation, mesoderm formation and
neurulation. Consistent with the poised identity, during differentiation of
hESCs to neuroepithelium, a neuroectoderm-specific subset of poised enhancers
acquires a chromatin signature associated with active enhancers. When assayed in
zebrafish embryos, poised enhancers are able to direct cell-type and
stage-specific expression characteristic of their proximal developmental gene,
even in the absence of sequence conservation in the fish genome. Our data
demonstrate that early developmental enhancers are epigenetically pre-marked in
hESCs and indicate an unappreciated role of H3K27me3 at distal regulatory
elements. Moreover, the wealth of new regulatory sequences identified here
provides an invaluable resource for studies and isolation of transient, rare
cell populations representing early stages of human embryogenesis. Enhancers play a pivotal role in regulating the transcription of distal genes.
Although certain chromatin features, such as the histone acetyltransferase P300
and the histone modification H3K4me1, indicate the presence of enhancers, only a
fraction of enhancers are functionally active. Individual chromatin marks, such
as H3K27ac and H3K27me3, have been identified to distinguish active from
inactive enhancers. However, the systematic identification of the most
informative single modification, or combination thereof, is still lacking.
Furthermore, the discovery of enhancer RNAs (eRNAs) provides an alternative
approach to directly predicting enhancer activity. However, it remains
challenging to link chromatin modifications to eRNA transcription. Herein, we
develop a logistic regression model to unravel the relationship between
chromatin modifications and eRNA synthesis. We perform a systematic assessment
of 24 chromatin modifications in fetal lung fibroblast and demonstrate that a
combination of four modifications is sufficient to accurately predict eRNA
transcription. Furthermore, we compare the ability of eRNAs and H3K27ac to
discriminate enhancer activity. We demonstrate that eRNA is more indicative of
enhancer activity. Finally, we apply our fibroblast trained model to six other
cell-types and successfully predict eRNA synthesis. Thus, we demonstrate the
learned relationships are general and independent of cell-type. We provided a
powerful tool to identify active enhancers and reveal the relationship between
chromatin modifications, eRNA production and enhancer activity. BACKGROUND: Transcriptional regulation in multi-cellular organisms is a complex
process involving multiple modular regulatory elements for each gene. Building
whole-genome models of transcriptional networks requires mapping all relevant
enhancers and then linking them to target genes. Previous methods of enhancer
identification based either on sequence information or on epigenetic marks have
different limitations stemming from incompleteness of each of these datasets
taken separately.
RESULTS: In this work we present a new approach for discovery of regulatory
elements based on the combination of sequence motifs and epigenetic marks
measured with ChIP-Seq. Our method uses supervised learning approaches to train
a model describing the dependence of enhancer activity on sequence features and
histone marks. Our results indicate that using combination of features provides
superior results to previous approaches based on either one of the datasets.
While histone modifications remain the domit feature for accurate
predictions, the models based on sequence motifs have advantages in their
general applicability to different tissues. Additionally, we assess the
relevance of different sequence motifs in prediction accuracy showing that even
tissue-specific enhancer activity depends on multiple motifs.
CONCLUSIONS: Based on our results, we conclude that it is worthwhile to include
sequence motif data into computational approaches to active enhancer prediction
and also that classifiers trained on a specific set of enhancers can generalize
with significant accuracy beyond the training set. BACKGROUND: Regulated gene expression controls organismal development, and
variation in regulatory patterns has been implicated in complex traits. Thus
accurate prediction of enhancers is important for further understanding of these
processes. Genome-wide measurement of epigenetic features, such as histone
modifications and occupancy by transcription factors, is improving enhancer
predictions, but the contribution of these features to prediction accuracy is
not known. Given the importance of the hematopoietic transcription factor TAL1
for erythroid gene activation, we predicted candidate enhancers based on genomic
occupancy by TAL1 and measured their activity. Contributions of multiple
features to enhancer prediction were evaluated based on the results of these and
other studies.
RESULTS: TAL1-bound DNA segments were active enhancers at a high rate both in
transient transfections of cultured cells (39 of 79, or 56%) and transgenic mice
(43 of 66, or 65%). The level of binding signal for TAL1 or GATA1 did not help
distinguish TAL1-bound DNA segments as active versus inactive enhancers, nor did
the density of regulation-related histone modifications. A meta-analysis of
results from this and other studies (273 tested predicted enhancers) showed that
the presence of TAL1, GATA1, EP300, SMAD1, H3K4 methylation, H3K27ac, and CAGE
tags at DNase hypersensitive sites gave the most accurate predictors of enhancer
activity, with a success rate over 80% and a median threefold increase in
activity. Chromatin accessibility assays and the histone modifications H3K4me1
and H3K27ac were sensitive for finding enhancers, but they have high false
positive rates unless transcription factor occupancy is also included.
CONCLUSIONS: Occupancy by key transcription factors such as TAL1, GATA1, SMAD1,
and EP300, along with evidence of transcription, improves the accuracy of
enhancer predictions based on epigenetic features. The fundamental repeating unit of eukaryotic chromatin is the nucleosome.
Besides being involved in packaging DNA, nucleosome organization plays an
important role in transcriptional regulation and cellular identity. Currently,
there is much debate about the major determits of the nucleosome architecture
of a genome and its significance with little being known about its role in stem
cells. To address these questions, we performed ultra-deep sequencing of
nucleosomal DNA in two human embryonic stem cell lines and integrated our data
with numerous epigenomic maps. Our analyses have revealed that the genome is a
determit of nucleosome organization with transcriptionally inactive regions
characterized by a "ground state" of nucleosome profiles driven by underlying
DNA sequences. DNA sequence preferences are associated with heterogeneous
chromatin organization around transcription start sites. Transcription, histone
modifications, and DNA methylation alter this "ground state" by having distinct
effects on both nucleosome positioning and occupancy. As the transcriptional
rate increases, nucleosomes become better positioned. Exons transcribed and
included in the final spliced mRNA have distinct nucleosome profiles in
comparison to exons not included at exon-exon junctions. Genes marked by the
active modification H3K4m3 are characterized by lower nucleosome occupancy
before the transcription start site compared to genes marked by the inactive
modification H3K27m3, while bivalent domains, genes associated with both marks,
lie exactly in the middle. Combinatorial patterns of epigenetic marks (chromatin
states) are associated with unique nucleosome profiles. Nucleosome organization
varies around transcription factor binding in enhancers versus promoters. DNA
methylation is associated with increasing nucleosome occupancy and different
types of methylations have distinct location preferences within the nucleosome
core particle. Finally, computational analysis of nucleosome organization alone
is sufficient to elucidate much of the circuitry of pluripotency. Our results,
suggest that nucleosome organization is associated with numerous genomic and
epigenomic processes and can be used to elucidate cellular identity. The normal adult human mammary gland is a continuous bilayered epithelial
system. Bipotent and myoepithelial progenitors are prominent and unique
components of the outer (basal) layer. The inner (luminal) layer includes both
luminal-restricted progenitors and a phenotypically separable fraction that
lacks progenitor activity. We now report an epigenomic comparison of these three
subsets with one another, with their associated stromal cells, and with three
immortalized, non-tumorigenic human mammary cell lines. Each genome-wide
analysis contains profiles for six histone marks, methylated DNA, and RNA
transcripts. Analysis of these datasets shows that each cell type has unique
features, primarily within genomic regulatory regions, and that the cell lines
group together. Analyses of the promoter and enhancer profiles place the luminal
progenitors in between the basal cells and the non-progenitor luminal subset.
Integrative analysis reveals networks of subset-specific transcription factors. Enhancer activation is a critical step for gene activation. Here we report an
epigenetic crosstalk at enhancers between the UTX (H3K27 demethylase)-MLL4 (H3K4
methyltransferase) complex and the histone acetyltransferase p300. We
demonstrate that UTX, in a demethylase activity-independent manner, facilitates
conversion of inactive enhancers in embryonic stem cells to an active
(H3K4me1+/H3K27ac+) state by recruiting and coupling the enzymatic functions of
MLL4 and p300. Loss of UTX leads to attenuated enhancer activity, characterized
by reduced levels of H3K4me1 and H3K27ac as well as impaired transcription. The
UTX-MLL4 complex enhances p300-dependent H3K27 acetylation through UTX-dependent
stimulation of p300 recruitment, while MLL4-mediated H3K4 monomethylation,
reciprocally, requires p300 function. Importantly, MLL4-generated H3K4me1
further enhances p300-dependent transcription. This work reveals a previously
unrecognized cooperativity among enhancer-associated chromatin modulators,
including a unique function for UTX, in establishing an "active enhancer
landscape" and defines a detailed mechanism for the joint deposition of H3K4me1
and H3K27ac. |
What does DMARD stand for? | DMARD stands for disease-modifying antirheumatic drug. | OBJECTIVE: To determine whether intensive combinations of conventional synthetic
disease-modifying antirheumatic drugs (csDMARDS) achieve similar clinical
benefits more cheaply than high-cost biologics such as tumor necrosis factor
inhibitors (TNFi) in patients with active rheumatoid arthritis (RA) whose
illness has failed to respond to methotrexate and another DMARD.
METHODS: We used within-trial cost-effectiveness and cost-utility analyses from
health and social care and 2 societal perspectives. Participants were recruited
into an open-label, 12-month, pragmatic, randomized, multicenter, 2-arm,
noninferiority trial in 24 rheumatology clinics in England and Wales. Costs were
linked with the Health Assessment Questionnaire (HAQ; primary outcome) and
quality-adjusted life years derived from 2 measures (Short-Form 36 health survey
and EuroQol 5-domain 3-level instrument).
RESULTS: In total, 205 participants were recruited, 104 in the csDMARD arm and
101 in the TNFi arm. Participants in the csDMARD arm with poor response at 6
months were offered TNFi; 46 participants (44%) switched. Relevant cost and
outcome data were available for 93% of participants at 6-month follow-up and for
91-92% of participants at 12-month follow-up. The csDMARD arm had significantly
lower total costs from all perspectives (6-month health and social care adjusted
mean difference -£3,615 [95% confidence interval (95% CI) -4,104, -3,182];
12-month health and social care adjusted mean difference -£1,930 [95% CI -2,599,
-1,301]). The HAQ score showed benefit to the csDMARD arm at 12 months (-0.16
[95% CI -0.32, -0.01]); other outcomes/follow-ups showed no differences.
CONCLUSION: Starting treatment with csDMARDs, rather than TNFi, achieves similar
outcomes at significantly lower costs. Patients with active RA and who meet the
National Institute for Health and Care Excellence criteria for expensive
biologics can be treated with combinations of intensive csDMARDs in a
cost-effective manner. |
Can SMAD6 variants cause craniosynostosis? | Yes, SMAD6 variants can cause craniosynostosis. |