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Molecular Biotechnology Jan 2021The repressor element 1 (RE1) silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF) modulates the expression of genes with...
The repressor element 1 (RE1) silencing transcription factor/neuron-restrictive silencing factor (REST/NRSF) modulates the expression of genes with RE1/neuron-restrictive silencing element (RE1/NRSE) sites by recruiting the switch independent 3 (SIN3) factor and the REST corepressor (COREST) to its N and C-terminal repressor domain, respectively. Both, SIN3 and COREST assemble into protein complexes that are composed of multiple subunits including a druggable histone deacetylase (HDAC) enzyme. The SIN3 core complex comprises the eponymous proteins SIN3A or SIN3B, the catalytically active proteins HDAC1 or HDAC2, the histone chaperone retinoblastoma-associated protein 46/retinoblastoma-binding protein 7 (RBAP46/RBBP7) or RBAP48/RBBP4, the SIN3-associated protein 30 (SAP30), and the suppressor of defective silencing 3 (SDS3). Here, we overcome a bottleneck limiting the molecular characterization of the REST/NRSF-SIN3 transcriptional corepressor complex. To this end, SIN3 genes were amplified from the complementary DNA of neural stem/progenitor cells, and expressed in a baculovirus/insect cell expression system. We show that the isolates bind to DNA harboring RE1/NRSE sites and demonstrate that the histone deacetylase activity is blocked by small-molecule inhibitors. Thus, our isolates open up for future biomedical research on this critical transcriptional repressor complex and are envisioned as tool for drug testing.
Topics: Animals; Baculoviridae; Benzamides; Co-Repressor Proteins; Depsipeptides; Gene Library; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Nerve Tissue Proteins; Neural Stem Cells; Pyrimidines; Recombinant Proteins; Regulatory Elements, Transcriptional; Repressor Proteins; Sf9 Cells; Sin3 Histone Deacetylase and Corepressor Complex
PubMed: 33130996
DOI: 10.1007/s12033-020-00283-7 -
Bioorganic & Medicinal Chemistry Letters Nov 2020Several novel indirubin-based N-hydroxybenzamides, N-hydropropenamides and N-hydroxyheptanamides (4a-h, 7a-h, 10a-h) were designed using a fragment-based approach with...
Design, synthesis and evaluation of novel indirubin-based N-hydroxybenzamides, N-hydroxypropenamides and N-hydroxyheptanamides as histone deacetylase inhibitors and antitumor agents.
Several novel indirubin-based N-hydroxybenzamides, N-hydropropenamides and N-hydroxyheptanamides (4a-h, 7a-h, 10a-h) were designed using a fragment-based approach with structural features extracted from several previously reported HDAC inhibitors, such as SAHA (vorinostat), MGCD0103 (mocetinostat), nexturastat A and PXD-101 (belinostat). The biological results reveal that our compounds showed excellent cytotoxicity toward three common human cancer cell lines (SW620, PC-3 and NCI-H23) with IC values ranging from 0.09 to 0.007 µM. The cytotoxicity of the compounds was equipotent or even up to 10-times more potent than adriamycin and up to 205-times more potent than SAHA. Among the series of N-hydroxypropenamides, compounds 10a-d were the most potent HDAC inhibitors as well as cytotoxicity toward the cell lines tested. In addition, the strong inhibitory activites toward HDAC of our compounds were observed with IC values of below-micromolar range. Especially, compound 4a inhibited HDAC6 with an IC value of 29-fold lower than that against HDAC2 isoform. Representative compounds 4a and 7a were found to significantly arrest SW620 cells at G0/G1 phase. Compounds 7a and 10a were found to strongly induce apoptosis in SW620 cells. Docking studies revealed some important features affecting the selectivity against HDAC6 isoform. The results clearly demonstrate the potential of the indirubin-hydroxamic acid hybrids and these compounds should be very promising for further development.
Topics: Amides; Antineoplastic Agents; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Drug Design; Drug Screening Assays, Antitumor; Histone Deacetylase 2; Histone Deacetylase 6; Histone Deacetylase Inhibitors; Humans; Indoles; Molecular Structure; Structure-Activity Relationship
PubMed: 32916298
DOI: 10.1016/j.bmcl.2020.127537 -
European Review For Medical and... Apr 2020To investigate the effect and mechanism of mocetinostat on diminishing epidural fibrosis. Dysregulated wound repair usually occurs after injury or surgery and is...
OBJECTIVE
To investigate the effect and mechanism of mocetinostat on diminishing epidural fibrosis. Dysregulated wound repair usually occurs after injury or surgery and is featured by excessive scar tissue contributed by fibrosis. Increasing researches demonstrated that histone acetylation, an epigenetic alteration, plays a crucial role in fibrosis. However, the mechanism of the complicated process remains unclear. In the current study, the effect of histone deacetylase (HDAC) inhibitor mocetinostat in a rat model of epidural fibrosis was detected, and it was discovered that mocetinostat suppressed myofibroblast activation and increased apoptosis by reducing Akt/GSK3b signaling.
PATIENTS AND METHODS
First, the levels of histone acetylation in the patients' epidural fibroblasts were analyzed. Then, mRNAs and proteins obtained from human fibroblasts following TGF-β activation and mocetinostat treatment in vitro were used to examine the influence of mocetinostat on the activation and survival of fibroblasts, so as to explore the related mechanism of mocetinostat. The laminectomy model was established in rats to observe the therapeutic effect of mocetinostat on epidural scar tissues.
RESULTS
In this research, it was found that the increase of HDAC1 in human dura scar was accompanied by the aggravation of fibrosis. In addition, cell assay demonstrated that mocetinostat inhibited fibroblast activation and accelerated apoptosis by inhibiting Akt/GSK3b pathway. In the rat model, mocetinostat weakened scar hyperplasia and collagen deposition and effectively inhibited the process of epidural fibrosis.
CONCLUSIONS
The above results indicate that mocetinostat inhibits HDAC1 expression and decreases the conduction of the AKT/GSK3b pathway in fibroblasts, leading to myofibroblast activation and apoptosis elevation. Hence, mocetinostat ameliorates epidural fibrosis.
Topics: Animals; Apoptosis; Benzamides; Cells, Cultured; Epidural Space; Fibrosis; Humans; Laminectomy; Myofibroblasts; Pyrimidines; Rats; Rats, Sprague-Dawley
PubMed: 32373984
DOI: 10.26355/eurrev_202004_21029 -
British Journal of Pharmacology Jan 2021Epigenetic mechanisms, including DNA methylation and histone post-translational modifications (PTMs), have been known to regulate chromatin structure and... (Review)
Review
Epigenetic mechanisms, including DNA methylation and histone post-translational modifications (PTMs), have been known to regulate chromatin structure and lineage-specific gene expression during cardiovascular development and disease. However, alterations in the landscape of histone PTMs and their contribution to the pathogenesis of incurable cardiovascular diseases such as pulmonary hypertension (PH) and associated right heart failure (RHF) remain largely unexplored. This review focusses on the studies in PH and RHF that investigated the gene families that write (histone acetyltransferases), read (bromodomain-containing proteins) or erase (histone deacetylases [HDACs] and sirtuins [SIRT]) acetyl moieties from the ε-amino group of lysine residues of histones and non-histone proteins. Analysis of cells and tissues isolated from the in vivo preclinical models of PH and human pulmonary arterial hypertension not only confirmed significant alterations in the expression levels of multiple HDACs, SIRT1, SIRT3 and BRD4 proteins but also demonstrated their strong association to proliferative, inflammatory and fibrotic phenotypes linked to the pathological vascular remodelling process. Due to the reversible nature of post-translational protein acetylation, the therapeutic efficacy of numerous small-molecule inhibitors (vorinostat, valproic acid, sodium butyrate, mocetinostat, entinostat, tubastatin A, apabetalone, JQ1 and resveratrol) have been evaluated in different preclinical models of cardiovascular disease, which revealed the promising therapeutic benefits of targeting histone acetylation pathways in the attenuation of cardiac hypertrophy, fibrosis, left heart dysfunction, PH and RHF. This review also emphasizes the need for deeper molecular insights into the contribution of epigenetic changes to PH pathogenesis and therapeutic evaluation of isoform-specific modulation in ex vivo and in vivo models of PH and RHF. LINKED ARTICLES: This article is part of a themed issue on Risk factors, comorbidities, and comedications in cardioprotection. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.1/issuetoc.
Topics: Acetylation; Cell Cycle Proteins; Histones; Humans; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Nuclear Proteins; Protein Processing, Post-Translational; Transcription Factors
PubMed: 31749139
DOI: 10.1111/bph.14932 -
Sarcoma 2019[This corrects the article DOI: 10.1155/2018/2068517.].
Corrigendum to "SARC018_SPORE02: Phase II Study of Mocetinostat Administered with Gemcitabine for Patients with Metastatic Leiomyosarcoma with Progression or Relapse following Prior Treatment with Gemcitabine-Containing Therapy".
[This corrects the article DOI: 10.1155/2018/2068517.].
PubMed: 31534435
DOI: 10.1155/2019/7608743 -
Cancers Sep 2019Pancreatic ductal adenocarcinoma (PDAC) has a five-year survival rate of <10% due in part to a lack of effective therapies. Pan-histone deacetylase (HDAC) inhibitors...
Pancreatic ductal adenocarcinoma (PDAC) has a five-year survival rate of <10% due in part to a lack of effective therapies. Pan-histone deacetylase (HDAC) inhibitors have shown preclinical efficacy against PDAC but have failed in the clinic due to toxicity. Selective HDAC inhibitors may reduce toxicity while retaining therapeutic efficacy. However, their use requires identification of the specific HDACs that mediate the therapeutic effects of HDAC inhibitors in PDAC. We determined that the HDAC1/2/3 inhibitor Mocetinostat synergizes with the HDAC4/5/6 inhibitor LMK-235 in a panel of PDAC cell lines. Furthermore, while neither drug alone synergizes with gemcitabine, the combination of Mocetinostat, LMK-235, and gemcitabine showed strong synergy. Using small interfering (si)RNA-mediated knockdown, this synergy was attributed to inhibition of HDACs 1, 2, and 6. Pharmacological inhibition of HDACs 1 and 2 with Romidepsin and HDAC6 with ACY-1215 also potently synergized with gemcitabine in a panel of PDAC cell lines, and this drug combination potentiated the antitumor effects of gemcitabine against PDAC xenografts in vivo. Collectively, our data show that inhibition of multiple HDACs is required for therapeutic effects of HDAC inhibitors and support the development of novel strategies to inhibit HDACs 1, 2, and 6 for PDAC therapy.
PubMed: 31500290
DOI: 10.3390/cancers11091327 -
Communications Biology 2019Cancer cells exhibit phenotypic plasticity during epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) involving intermediate states. To...
Cancer cells exhibit phenotypic plasticity during epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) involving intermediate states. To study genome-wide epigenetic remodeling associated with EMT plasticity, we integrate the analyses of DNA methylation, ChIP-sequencing of five histone marks (H3K4me1, H3K4me3, H3K27Ac, H3K27me3 and H3K9me3) and transcriptome profiling performed on ovarian cancer cells with different epithelial/mesenchymal states and on a knockdown model of EMT suppressor Grainyhead-like 2 (GRHL2). We have identified differentially methylated CpG sites associated with EMT, found at promoters of epithelial genes and GRHL2 binding sites. GRHL2 knockdown results in CpG methylation gain and nucleosomal remodeling (reduction in permissive marks H3K4me3 and H3K27ac; elevated repressive mark H3K27me3), resembling the changes observed across progressive EMT states. Epigenetic-modifying agents such as 5-azacitidine, GSK126 and mocetinostat further reveal cell state-dependent plasticity upon GRHL2 overexpression. Overall, we demonstrate that epithelial genes are subject to epigenetic control during intermediate phases of EMT/MET involving GRHL2.
Topics: Cell Line, Tumor; CpG Islands; DNA Methylation; DNA-Binding Proteins; Epigenesis, Genetic; Epithelial-Mesenchymal Transition; Female; Gene Knockdown Techniques; Histones; Humans; Ovarian Neoplasms; Transcription Factors
PubMed: 31372511
DOI: 10.1038/s42003-019-0506-3 -
Stem Cell Research Jul 2019Here we utilized the chromatin in vivo assay (CiA) mouse platform to directly examine the epigenetic barriers impeding the activation of the CiA:Oct4 allele in mouse...
Here we utilized the chromatin in vivo assay (CiA) mouse platform to directly examine the epigenetic barriers impeding the activation of the CiA:Oct4 allele in mouse embryonic fibroblasts (MEF)s when stimulated with a transcription factor. The CiA:Oct4 allele contains an engineered EGFP reporter replacing one copy of the Oct4 gene, with an upstream Gal4 array in the promoter that allows recruitment of chromatin modifying machinery. We stimulated gene activation of the CiA:Oct4 allele by binding a transcriptional activator to the Gal4 array. As with cellular reprograming, this process is inefficient with only a small percentage of the cells re-activating CiA:Oct4 after weeks. Epigenetic barriers to gene activation potentially come from heavy DNA methylation, histone deacetylation, chromatin compaction, and other posttranslational marks (PTM) at the differentiated CiA:Oct4 allele in MEFs. Using this platform, we performed a high-throughput chemical screen for compounds that increased the efficiency of activation. We found that Azacytidine and newer generation histone deacetylase (HDAC) inhibitors were the most efficient at facilitating directed transcriptional activation of this allele. We found one hit form our screen, Mocetinostat, improved iPSC generation under transcription factor reprogramming conditions. These results separate individual allele activation from whole cell reprograming and give new insights that will advance tissue engineering.
Topics: Alleles; Animals; Chromatin; DNA Methylation; Epigenesis, Genetic; Histone Deacetylase Inhibitors; Induced Pluripotent Stem Cells; Mice; Octamer Transcription Factor-3; Transcriptional Activation
PubMed: 31170660
DOI: 10.1016/j.scr.2019.101470 -
The Protein Journal Oct 2019The Sonic hedgehog signalling is known to play a crucial role in regulating embryonic development, cancer stem cell maintenance and tissue patterning. Dysregulated...
The Sonic hedgehog signalling is known to play a crucial role in regulating embryonic development, cancer stem cell maintenance and tissue patterning. Dysregulated hedgehog signalling has been reported to affect tumorigenesis and drug response in various human malignancies. Epigenetic therapy relying on DNA methyltransferase and Histone deacetylase inhibitors are being proposed as potential drug candidates considering their efficiency in preventing development of cancer progenitor cells, killing drug resistant cells and also dictating "on/off" switch of tumor suppressor genes and oncogenes. In this docking approach, epigenetic modulators were virtually screened for their efficiency in inhibiting key regulators of SHH pathway viz., sonic hedgehog, Smoothened and Gli using polypharmacological approach. The control drugs and epigenetic modulators were docked with PDB protein structures using AutoDock vina and further checked for their drug-likeness properties. Further molecular dynamics simulation using VMD and NAMD, and MMP/GBSA energy calculation were employed for verifying the stability and entropy of the ligand-receptor complex. EPZ-6438 and GSK 343 (EZH2 inhibitors), CHR 3996 and Mocetinostat (HDAC inhibitors), GSK 126 (HKMT inhibitor) and UNC 1215 (L3MBTL3 antagonist) exhibited multiple-targeted approach in modulating HH signalling. This is the first study to report these epigenetic drugs as potential multi-targeted hedgehog pathway inhibitors. Thus, epigenetic polypharmacology approach can be explored as a better alternative to challenges of acute long term toxicity and drug resistance occurring due to traditional single targeted chemotherapy in the future.
Topics: Antineoplastic Agents; Databases, Pharmaceutical; Databases, Protein; Drug Discovery; Epigenesis, Genetic; Hedgehog Proteins; Humans; Molecular Docking Simulation; Molecular Targeted Therapy; Neoplasms; Signal Transduction; Thermodynamics
PubMed: 30993446
DOI: 10.1007/s10930-019-09832-9 -
Science Translational Medicine Mar 2019Mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitors have failed to show clinical benefit in Kirsten rat sarcoma () mutant lung cancer due to various...
Mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitors have failed to show clinical benefit in Kirsten rat sarcoma () mutant lung cancer due to various resistance mechanisms. To identify differential therapeutic sensitivities between epithelial and mesenchymal lung tumors, we performed in vivo small hairpin RNA screens, proteomic profiling, and analysis of patient tumor datasets, which revealed an inverse correlation between mitogen-activated protein kinase (MAPK) signaling dependency and a zinc finger E-box binding homeobox 1 (ZEB1)-regulated epithelial-to-mesenchymal transition. Mechanistic studies determined that MAPK signaling dependency in epithelial lung cancer cells is due to the scaffold protein interleukin-17 receptor D (IL17RD), which is directly repressed by ZEB1. Lung tumors in multiple mutant murine models with increased ZEB1 displayed low IL17RD expression, accompanied by MAPK-independent tumor growth and therapeutic resistance to MEK inhibition. Suppression of ZEB1 function with miR-200 expression or the histone deacetylase inhibitor mocetinostat sensitized resistant cancer cells to MEK inhibition and markedly reduced in vivo tumor growth, showing a promising combinatorial treatment strategy for mutant cancers. In human lung tumor samples, high ZEB1 and low IL17RD expression correlated with low MAPK signaling, presenting potential markers that predict patient response to MEK inhibitors.
Topics: Animals; Benzamides; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Drug Resistance, Neoplasm; Epithelial Cells; Humans; Lung Neoplasms; MAP Kinase Signaling System; Mesoderm; Mice; MicroRNAs; Mitogen-Activated Protein Kinase Kinases; Mutation; Neoplasms; Phosphorylation; Protein Kinase Inhibitors; Proto-Oncogene Proteins p21(ras); Pyrimidines; Receptors, Interleukin-17; Zinc Finger E-box-Binding Homeobox 1
PubMed: 30867319
DOI: 10.1126/scitranslmed.aaq1238