Activating transcription factor 5 (ATF5) is a cellular prosurvival transcription factor within the basic leucine zipper (bZip) family that is involved in cellular differentiation and promotes ...cellular adaptation to stress. Recent studies have characterized the oncogenic role of ATF5 in the development of several different types of cancer, notably glioblastoma. Preclinical assessment of a systemically deliverable dominant-negative ATF5 (dnATF5) biologic has found that targeting ATF5 results in tumor regression and tumor growth inhibition of glioblastoma xenografts in mouse models. In this review, we comprehensively and critically detail the current scientific literature on ATF5 in the context of cellular differentiation, survival, and response to stressors in normal tissues. Furthermore, we will discuss how the prosurvival role of ATF5 aides in cancer development, followed by current advances in targeting ATF5 using dominant-negative biologics, and perspectives on future research.
Summary
Background
Eczema control has been identified as an important outcome by key stakeholders in eczema research (including patients, carers, healthcare professionals and researchers) but no ...validated instruments for the domain have been identified.
Objectives
To develop a measurement instrument to capture a patient's perspective of eczema control that is suitable for use in eczema clinical trials.
Methods
Best practice for the development of a patient‐reported outcome was followed. A mixed‐methods approach was used to develop and refine a conceptual framework, generate, refine and select items and to test the distribution and construct validity of the final scale. The mixed‐methods approach involved expert panel meetings (including patient representatives, healthcare professionals and methodologists), and data collection using a focus group, cognitive interviews and an online survey with people with eczema and caregivers. Multivariable linear regression was used in the item selection process.
Results
Fourteen expert panel members co‐produced the instrument, with input from people with eczema and caregivers via a focus group (n = 6), cognitive interviews (n = 13) and an online survey (n = 330). The resulting instrument, Recap of atopic eczema (RECAP), is a seven‐item questionnaire that captures eczema control via self or caregiver report. The development process aimed to ensure good content validity and feasibility. Initial testing suggested no floor or ceiling effects and good construct validity. Hypothesized correlation with the Patient‐Oriented Eczema Measure was confirmed r(258) = 0·83, P < 0·001.
Conclusions
RECAP has the potential to improve reporting of eczema control in research and clinical practice. Further exploration of measurement properties is required.
Linked Comment: Pattinson and Bundy. Br J Dermatol 2020; 183:418–419.
What's already known about this topic?
Eczema control has been identified as an important outcome by key stakeholders in eczema research (including patients, carers, healthcare professionals and researchers).
Qualitative studies suggest eczema control is a multifaceted and individual experience and no instrument has been identified that captures eczema control in this way.
What does this study add?
We have developed Recap of atopic eczema (RECAP), a seven‐item questionnaire to capture the experience of eczema control in all ages and eczema severities; there are two versions: a self‐reported version for adults and older children with eczema, and a caregiver‐reported version for younger children with eczema.
Designed with input from people with eczema, caregivers and healthcare professionals to ensure good content validity.
Initial testing of score distributions and construct validity suggests good measurement properties.
What are the clinical implications of the work?
The RECAP instrument is appropriate and feasible for measuring eczema control in clinical trials and may also be useful in routine practice.
Linked Comment: Pattinson and Bundy. Br J Dermatol 2020; 183:418–419.
Plain language summary available online
Introduction
Mutations in
isocitrate dehydrogenase 1/2
(IDH
mut
) identify a subset of gliomas that exhibit epigenetic dysregulation via aberrant DNA methylation. These tumors are ultimately fatal ...and lack effective therapeutic strategies. Considering the epigenetic dysregulation of IDH
mut
gliomas, we hypothesized that epigenetic-targeting drugs may yield therapeutic benefits in gliomas bearing IDH
mut
. One set of targets includes the bromodomain and extraterminal (BET) family of transcriptional coactivators.
Methods
We used TCGA data from glioma patients to determine whether BET proteins affect patient survival differently based on IDH status. Follow-up experiments using a set of IDH wildtype/mutant glioma cultures, as well as an IDH wildtype glioblastoma cell line expressing exogenous
R132H IDH1
, focused on cell health assays to investigate whether IDH
mut
was associated with increased sensitivity to the BET inhibitor JQ1. Immunoblots were used to evaluate the molecular response to JQ1 in these cultures.
Results
We identified that high BRD4 expression associated with decreased survival only in IDH
mut
glioma patients. Cell viability analysis showed that IDH
mut
sensitized glioma cells to delayed cytotoxicity (10 days) in response to JQ1. Early effects of JQ1 (3 days) were primarily antiproliferative, with IDH
mut
glioma exhibiting a modest increase in sensitivity. Finally, exogenous
R132H IDH1
expression in a resistant IDH wildtype cell line recapitulated the JQ1-mediated delayed cytotoxicity seen in our endogenous IDH
mut
glioma cells.
Conclusion
Overall, these data suggest that BRD4 enhances malignancy primarily in gliomas bearing IDH
mut
and is associated with greater sensitivity to BET inhibition. The finding that BET inhibition primarily exhibits delayed cytotoxicity may be overlooked in conventional short endpoint dose–response assays. Follow-up mechanistic and animal studies will help address the translational potential of these findings.
Seizures are a frequent complication of adult-type diffuse gliomas, and are often difficult to control with medications. Gliomas with mutations in isocitrate dehydrogenase 1 or 2 (IDHmut) are more ...likely than IDH-wild type (IDHwt) gliomas to cause seizures as part of their initial clinical presentation. However, whether IDHmut is also associated with seizures during the remaining disease course, and whether IDHmut inhibitors can reduce seizure risk, are unclear. Clinical multivariable analyses showed that preoperative seizures, glioma location, extent of resection, and glioma molecular subtype (including IDHmut status) all contributed to postoperative seizure risk in adult-type diffuse glioma patients, and that postoperative seizures were often associated with tumor recurrence. Experimentally, the metabolic product of IDHmut, d-2-hydroxyglutarate, rapidly synchronized neuronal spike firing in a seizure-like manner, but only when non-neoplastic glial cells were present. In vitro and in vivo models recapitulated IDHmut glioma-associated seizures, and IDHmut inhibitors currently being evaluated in glioma clinical trials inhibited seizures in those models, independent of their effects on glioma growth. These data show that postoperative seizure risk in adult-type diffuse gliomas varies in large part by molecular subtype, and that IDHmut inhibitors could play a key role in mitigating such risk in IDHmut glioma patients.
Introduction
A large subset of diffusely infiltrative gliomas contains a gain-of-function mutation in isocitrate dehydrogenase 1 or 2 (IDH1/2
mut
) which produces 2-hydroxglutarate, an inhibitor of ...α-ketoglutarate-dependent DNA demethylases, thereby inducing widespread DNA and histone methylation. Because histone deacetylase (HDAC) enzymes are localized to methylated chromatin via methyl-binding domain proteins, IDH1/2
mut
gliomas may be more dependent on HDAC activity, and therefore may be more sensitive to HDAC inhibitors.
Methods
Six cultured patient-derived glioma cell lines, IDH1
wt
(n = 3) and IDH1
mut
(n = 3), were treated with an FDA-approved HDAC inhibitor, panobinostat. Cellular cytotoxicity and proliferation assays were conducted by flow cytometry. Histone modifications and cell signaling pathways were assessed using immunoblot and/or ELISA.
Results
IDH1
mut
gliomas exhibited marked upregulation of genes associated with the HDAC activity. Glioma cell cultures bearing IDH1
mut
were significantly more sensitive to the cytotoxic and antiproliferative effects of panobinostat, compared to IDH1
wt
glioma cells. Panobinostat caused a greater increase in acetylation of the histone residues H3K14, H3K18, and H3K27 in IDH1
mut
glioma cells. Another HDAC inhibitor, valproic acid, was also more effective against IDH1
mut
glioma cells.
Conclusion
These data suggest that IDH1
mut
gliomas may be preferentially sensitive to HDAC inhibitors. Further, IDH1
mut
glioma cultures showed enhanced accumulation of acetylated histone residues in response to panobinostat treatment, suggesting a direct epigenetic mechanism for this sensitivity. This provides a rationale for further exploration of HDAC inhibitors against IDH1
mut
gliomas.
Background
Atopic eczema (also known as eczema) is a chronic, inflammatory skin condition that often afflicts patients’ health and well‐being. The Harmonising Outcome Measures for Eczema (HOME) ...initiative recommends that ‘long‐term control of eczema’ is measured in all clinical trials 3 months or longer in duration. However, little has been published on what eczema control means to those living with or treating atopic eczema.
Objectives
To (i) develop understanding of what eczema control means to patients, carers and clinicians and (ii) explore the feasibility and acceptability of different ways of measuring eczema control in the long term.
Methods
Online focus groups explored patients/carers experiences in the UK, the United States, the Netherlands, France, Sweden and Japan, and an international online survey gathered views of clinicians. The framework method was used to analyse the focus groups, and thematic analysis was used to analyse survey data. All findings were integrated into a theoretical framework to create overarching themes that cut across these diverse groups.
Results
Eight focus groups with patients (16 years+) and eight groups with carers of children took place (N = 97). Sixty‐two people took part in the survey. Eczema control was described as a multifaceted construct involving changes in disease activity, the treatment and management of the condition and psychological, social and physical functioning. Patient/carer measurement allows personal accounts and frequent measurement, whilst clinician measurement was deemed less subjective. The burden on patients/carers and issues for analysing and interpreting data should be considered.
Conclusions
This study formed the basis of judging the content validity and feasibility of measurement instruments/methods to assess control of eczema in clinical trials. This online approach to an international qualitative study is an example of how core outcome set developers with limited resources can engage with multiple stakeholder groups on an international basis to inform consensus meeting discussions.
Robust and reproducible protocols to efficiently reprogram adult canine cells to induced pluripotent stem cells are still elusive. Somatic cell reprogramming requires global chromatin remodeling that ...is finely orchestrated spatially and temporally. Histone acetylation and deacetylation are key regulators of chromatin condensation, mediated by histone acetyltransferases and histone deacetylases (HDACs), respectively. HDAC inhibitors have been used to increase histone acetylation, chromatin accessibility, and somatic cell reprogramming in human and mice cells. We hypothesized that inhibition of HDACs in canine fibroblasts would increase their reprogramming efficiency by altering the epigenomic landscape and enabling greater chromatin accessibility. We report that a combined treatment of panobinostat (LBH589) and vitamin C effectively inhibits HDAC function and increases histone acetylation in canine embryonic fibroblasts
in vitro
, with no significant cytotoxic effects. We further determined the effect of this treatment on global chromatin accessibility
via
Assay for Transposase-Accessible Chromatin using sequencing. Finally, the treatment did not induce any significant increase in cellular reprogramming efficiency. Although our data demonstrate that the unique epigenetic landscape of canine cells does not make them amenable to cellular reprogramming through the proposed treatment, it provides a rationale for a targeted, canine-specific, reprogramming approach by enhancing the expression of transcription factors such as CEBP.
Despite being widely regarded as generalist predators, amphibians exhibit a diversity of tooth shapes and dentition patterns, which may indicate the influence of dietary specialization on the ...evolution of tooth morphology. Very few studies have analysed the relationship between tooth morphology and diet (i.e., prey items) in amphibians, and those existing studies are highly speculative. We investigated the evolution of salamander teeth and the relationship between tooth morphology and diet in a phylogenetically independent fashion. We used a phylogeny of 23 species of salamander representing three families (Ambystomatidae, Plethodontidae, and Salamandridae) to, first, analyse the divergence of tooth morphology and its relationship to phylogeny and, second, to analyse the relationship between tooth morphology and diet diversity. We used electron scanning microscopy and a statistical comparative approach using Spatial Evolutionary and Ecological Analysis (SEEVA) and phylogenetic generalized least‐squares regression in R. Our results indicated significant divergence in tooth morphology at major phylogenetic splits. Moreover, there was a significant, phylogenetically independent relationship between tooth morphology and diet diversity. The relationship between diet and tooth morphology indicates not only a reflection of phylogenetic history, but also a degree of dietary specialization, indicating that evolution in tooth morphology has had an adaptive aspect in relation to salamander diet.
Abstract
Glioblastoma (GBM) is an incurable form of brain cancer with a median survival of ~15 months. Identification of a CpG Island Methylator Phenotype (CIMP) subtype of GBM (G-CIMP) represents a ...significant clinical discovery as these patients have an enhanced survival, with a median survival of 3 years. G-CIMP is characterized by a mutation in isocitrate dehydrogenase 1 or 2 (IDH1/2) which results in production of the oncometabolite 2-hydroxglutarate, an inhibitor of α-ketoglutarate-dependent DNA demethylases. This mutation occurs early in gliomagenesis and further results in aberrant DNA methylation and widespread transcriptional repression. Histone deacetylase (HDAC) enzymes are localized to methylated chromatin via methyl-binding domain (MBD) proteins, so we hypothesized that IDH mutant GBM exhibits enhanced reliance on HDAC activity, which is functionally significant to their cell proliferation and survival. Using a panel of 6 patient-derived cell lines grown in defined growth media, we show that G-CIMP is significantly more sensitive in vitro to the clinically approved pan histone deacetylase (HDAC) inhibitor panobinostat, with IDH mutant GBM cells exhibiting ~4 and ~10 fold lower IC/EC50 values in flow cytometric cell viability and apoptosis assays compared to IDH wild type (WT) GBM cells, respectively. For IDH mutant GBM, the average IC50 value was 13.5 nM for cell viability assays and the average EC50 value was 4.8 nM for apoptosis assays. Induction of cleaved caspase 3, a marker of apoptosis, was observed only in IDH mutant GBM cells when exposed to 10 nM panobinostat over 5 days. Analysis of proliferation via BrdU incorporation assays show that treatment with 10 nM panobinastat for 48 hours results in a 90% reduction in proliferation of IDH mutant GBM compared to only 25% reduction in IDH WT GBM cells. Molecular analysis via western blot shows that various acetylated chromatin marks, i.e. H3K14/18/27Ac, are preferentially upregulated in our IDH mutant GBM cell lines when exposed to 10 nM panobinostat for 48 hours. This was also supported by ELISA data showing that IDH mutant GBM cells have ~2 fold increase in histone H3 acetylation compared to IDH WT cells in response to 10 nM and 50 nM panobinostat. These data led us to perform a HDAC activity assay with nuclear extracts from our IDH WT and mutant GBM cells, but this assay showed no significant difference in basal HDAC activity. Overall, our results support a hypothesis where IDH mutant GBMs are more sensitive to panobinostat via preferential uptake of the drug. These data ultimately suggest that G-CIMP tumor cells display a profound sensitivity to the cytotoxic and antiproliferative effects mediated by exposure to the HDAC inhibitor panobinostat. These studies provide a strong foundation for future preclinical work evaluating the use of HDAC inhibitors as a personalized cancer therapy to treat G-CIMP.
Citation Format: Thomas K. Sears, Kevin D. Woolard. The HDAC inhibitor panobinostat elicits preferential cytotoxic and antiproliferative effects in IDH mutant glioblastoma abstract. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1917.
Abstract
Patients with glioblastoma frequently develop progressive disease following treatment. This is believed to occur because of an inherent or acquired resistance to chemo- and radiotherapy ...within residual tumor burden. However, how discrete tumor subpopulations coordinate tumor recurrence and progression is still poorly understood. Here, we describe the isolation of glioma stem cells (GSCs) from a patient prior to treatment (0203), and then from multiple sites in the brain following tumor recurrence. These sites include the recurrent tumor at the primary site (T3/T4), as well as disseminated tumor cells in the contralateral hemisphere (T6/T7). Whole exome sequencing identifies loss of all unique genomic signatures within the original tumor cells (0203) following treatment, suggesting eradication of the original tumor bulk. Moreover, genomic signatures of the recurrent tumor (T3/T4) clusters alongside disseminated subpopulations of tumor cells isolated from the contralateral hemisphere (T6/T7), suggesting that tumor recurrence is driven by repopulation of the original tumor location. Copy number variants (CNVs) between the original tumor (0203), the recurrent tumor (T3/T4), and distant tumor sites (T6/T7), identify a more primordial genome in distant tumor sites (T6/T7), which lack canonical CNVs found in the original tumor and in other subpopulations (deletion of PTEN or amplification of EGFR). Thus, tumor dissemination likely occurs early in disease development. We are able to model the patient's disease progression by using co-culture techniques and orthotopic xenograft studies. Our data indicates that 0203 cells actively suppress migration of disseminated tumor sub-populations (T6/T7) in vitro and in vivo, which is replicated by using cell-free conditioned media. In stark contrast, the induction of apoptosis in 0203 cells (or treating with conditioned media from apoptotic 0203 cells) rapidly recruits migration of disseminated cells (T6/T7) in a co-culture system. Co-culture of disseminated GSCs (T6/T7) with viable 0203 cells also results in suppression of pathways implicated in glioma migration, such as AKT, CXCR4, TrkA, and CLCN3; suppression that is reversed upon induction of apoptosis in 0203 cells. These data indicate that while widespread tumor dissemination occurs early in glioblastoma, a dominant subpopulation may actively suppress proliferation of separate, disseminated subpopulations. Moreover, abrogating this suppression following therapy triggers migration and proliferation of specific, distant tumor sub-populations, resulting in tumor recurrence. This suggests that glioblastoma recurrence may not be as simple as selection of radioresistant cells, rather it may result from the active recruitment of disseminated subpopulations, which migrate to the original tumor microenvironment.
Citation Format: Kevin D. Woolard, Patrick Huang, Thomas K. Sears, Matthew Settles. Functional heterogeneity in glioblastoma: cell-cell communication regulates temporospatial events in tumor recurrence abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 202.
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