Resistance to histone deacetylase inhibitors confers hypersensitivity to oncolytic reovirus therapy

Abstract Despite the promising antilymphoma activity of histone deacetylase (HDAC) inhibitors as a drug class, resistance is a significant clinical issue. Elucidating the molecular mechanisms driving HDAC inhibitor resistance and/or the specific targets that are altered in drug-resistant cells may facilitate the development of strategies that overcome drug resistance and are more effective for refractory patients. We generated novel T-cell lymphoma (TCL) cell line models of acquired resistance to the HDAC inhibitor belinostat to identify potential effective therapies. Belinostat-resistant cells displayed significant cross-resistance to other HDAC inhibitors including romidepsin, panobinostat, and vorinostat. Consistent with a lack of sensitivity to HDAC inhibitors, the resistant cells failed to induce increased acetylated histones. Drug-resistant cells featured significantly decreased expression of the key antiviral mediators IRF1 and STAT1. On the basis of these findings, we investigated the efficacy of the clinical formulation of reovirus (Reolysin) in parental and drug-resistant models. Our investigation revealed that HDAC inhibitor–resistant cells displayed enhanced vulnerability to reovirus replication and cell death in both in vitro and in vivo models compared with their parental counterparts. Importantly, Reolysin also significantly increased the antilymphoma activity of belinostat in HDAC inhibitor–resistant cells. Our data demonstrate that Reolysin alone or in combination with belinostat is a novel therapeutic strategy to treat TCL patients who develop resistance to HDAC inhibitors.

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Oncolytic Reovirus Is an Effective Treatment for Histone Deacetylase Inhibitor Resistant T-Cell Lymphoma

Blood ◽

10.1182/blood-2018-99-119152 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 2941-2941 ◽

Cited By ~ 1

Author(s):

Shariful Islam ◽

Claudia M Espitia ◽

Ning Qu ◽

Daniel Persky ◽

Jennifer S. Carew ◽

...

Keyword(s):

Gene Expression ◽

T Cell ◽

Hdac Inhibitor ◽

Research Funding ◽

Cell Lymphoma ◽

Hdac Inhibitors ◽

T Cell Lymphoma ◽

Drug Resistant ◽

Inhibitor Resistance ◽

Resistant Cells

Abstract Aberrant gene expression plays a pivotal role during tumorigenesis and cancer progression. The acetylation status of histones is an important determinant of gene expression and is controlled by two opposing classes of enzymes: histones acetyl transferases (HATs) and histone deacetylases (HDACs). The deacetylation of histones is associated with repression of key tumor suppressor genes and has been linked to HDAC overexpression in multiple forms of cancer including lymphomas. Several HDAC inhibitors have been FDA approved for T-cell lymphoma (TCL) therapy including belinostat, vorinostat, and romidepsin. Despite the promising anti-lymphoma activity of HDAC inhibitors as a drug class, resistance is a significant clinical issue. Identification of new strategies that are more effective in the drug resistant patient population is a high priority, but the mechanisms underlying HDAC inhibitor-induced cell death and the development of drug resistance are not completely understood. Elucidating the molecular mechanisms driving HDAC inhibitor resistance and/or the specific targets that are altered in drug-resistant cells may facilitate the development of strategies that overcome drug resistance and are effective for refractory patients. To pursue this goal, we generated novel TCL cell line models of acquired HDAC inhibitor resistance through repeated exposure to belinostat. The sensitivity of parental and resistant TCL cells to belinostat and other clinically relevant HDAC inhibitors was initially characterized using cell viability and flow cytometric assays. Notably, belinostat-resistant cells displayed significant cross-resistance to other HDAC inhibitors including vorinostat, romidepsin, panobinostat, and ricolinostat. This indicates that TCL patients that fail one HDAC inhibitor regimen may not benefit significantly from subsequent treatment with other drugs of this class. Consistent with a lack of sensitivity to HDAC inhibitors, the resistant cells failed to induce increased acetylated histones, tubulin acetylation (an HDAC6 target), and exhibited reduced upregulation of the CDK inhibitor p21 following belinostat treatment. In agreement with the absence of these hallmark characteristics of HDAC inhibition, belinostat also failed to cleave caspase-3 in the belinostat-resistant cells. Comprehensive transcriptome analysis was conducted to further characterize these new drug-resistant models and identify potential mechanisms of resistance and targets for second-line treatment. Drug resistant cells featured significantly increased basal levels of the reovirus receptor junctional adhesion molecule-A (JAM-A) as well as decreased JAK/STAT activity, a key antiviral response pathway. Based on these findings, we investigated the efficacy of the proprietary clinical formulation of reovirus (Reolysin) in parental and drug-resistant models. Our investigation revealed that belinostat-resistant cells displayed enhanced sensitivity to oncolytic reovirus-induced cell death compared to their parental counterparts. The increased benefit of Reolysin in resistant models was linked to elevated viral loads and more efficient endoplasmic reticular stress-mediated apoptosis. The heightened sensitivity of HDAC-resistant TCL cells to Reolysin was further validated in parental and belinostat-resistant T-cell lymphoma xenograft models. Collectively, these data demonstrate that oncolytic reovirus may be a novel therapeutic approach to treat T-cell lymphoma patients that are relapsed/refractory to HDAC inhibitors. We are currently planning an early phase clinical trial to further test the safety and benefit of this new approach. Disclosures Persky: Morphosys (IDMC): Consultancy; Merck: Research Funding; Spectrum: Research Funding; Genentech: Honoraria.

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153 DIFFERENT MOLECULAR MECHANISMS FOR HISTONE DEACETYLASE INHIBITOR-INDUCED APOPTOSIS IN DOG FIBROBLASTS AND MESENCHYMAL STEM CELLS

Reproduction Fertility and Development ◽

10.1071/rdv28n2ab153 ◽

2016 ◽

Vol 28 (2) ◽

pp. 206

Author(s):

M. J. Kim ◽

H. J. Oh ◽

G. A. Kim ◽

Y. K. Jo ◽

Y. B. Choi ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Stem Cell ◽

Histone Deacetylase ◽

Hdac Inhibitor ◽

Molecular Mechanisms ◽

Hdac Inhibitors ◽

Normal Cells ◽

Apoptotic Genes ◽

Induced Apoptosis

Histone deacetylase (HDAC) inhibitors have been applied to cancer research for a therapeutic purpose or somatic cell nuclear transfer for improvement of embryonic reprogramming. Considering the ubiquitous expression of HDAC in normal cells, effects of HDAC inhibitors on normal cells need to be evaluated in detail. Therefore, we aimed to investigate molecular mechanisms of HDAC inhibitor-induced apoptosis in mesenchymal stem cells and compare with the results from fibroblasts. Beagle skin fibroblasts (BF) and adipose-derived mesenchymal stem cell (MSC) lines were established from a 7-year-old beagle. Dulbecco’s modified Eagle medium supplemented with 10% fetal bovine serum and RCMEp (K-stem Cell Ltd, Seoul, Korea) were used as culture media for BF and MSC, respectively. A Food and Drug Administration-approved HDAC inhibitor, suberoylanilide hydroxamic acid (SAHA), was dissolved in dimethyl sulfoxide (DMSO). On passage 4, cells were subcultured to reach 50% of confluency, and cultured with 5 μM of SAHA. Culture medium containing the same volume of DMSO used in the SAHA group was used for the control. After 24 h, all cells were harvested, RNAs were extracted, and cDNA were synthesised. Transcript expression of anti-apoptotic genes (BFL, MCL, BCLxl, BCL2) and pro-apoptotic genes (BAX, BID, BIM) were analysed using RT-PCR. Two-way ANOVA using GraphPad Prism 5.0 (GraphPad Software, San Diego, CA, USA) with Bonferroni post-tests was performed for statistical analysis. Expression of MCL and BCLxl was significantly decreased in both groups. However, although BFL expression was remarkably increased only in BF (16.7-fold), BCL2 was significantly increased in MSC (7.2-fold) after SAHA treatment. Also, transcript of BAX was significantly increased in MSC (1.5-fold), and BID was significantly decreased in BF (0.3-fold). These results would be helpful to understand different molecular mechanisms of apoptosis induced by HDAC inhibitor on fibroblasts and mesenchymal stem cells. This study was supported by RDA (#PJ010928032015), IPET (#311062-04-3SB010), NRF (#2014R1A1A2059928), Research Institute for Veterinary Science, Nestle Purina PetCare, and the BK21 plus program.

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The Effect of Vicinal Difluorination on the Conformation and Potency of Histone Deacetylase Inhibitors

Molecules ◽

10.3390/molecules26133974 ◽

2021 ◽

Vol 26 (13) ◽

pp. 3974

Author(s):

A. Daryl Ariawan ◽

Flora Mansour ◽

Nicole Richardson ◽

Mohan Bhadbhade ◽

Junming Ho ◽

...

Keyword(s):

Histone Deacetylase ◽

Histone Deacetylase Inhibitors ◽

Hdac Inhibitors ◽

Molecular Conformations ◽

Fluorinated Analogs ◽

Selective Agents ◽

Isoform Selectivity

Histone deacetylase enzymes (HDACs) are potential targets for the treatment of cancer and other diseases, but it is challenging to design isoform-selective agents. In this work, we created new analogs of two established but non-selective HDAC inhibitors. We decorated the central linker chains of the molecules with specifically positioned fluorine atoms in order to control the molecular conformations. The fluorinated analogs were screened against a panel of 11 HDAC isoforms, and minor differences in isoform selectivity patterns were observed.

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Synergism of Heat Shock Protein 90 and Histone Deacetylase Inhibitors in Synovial Sarcoma

Sarcoma ◽

10.1155/2009/794901 ◽

2009 ◽

Vol 2009 ◽

pp. 1-10 ◽

Cited By ~ 15

Author(s):

Anne Nguyen ◽

Le Su ◽

Belinda Campbell ◽

Neal M. Poulin ◽

Torsten O. Nielsen

Keyword(s):

Heat Shock ◽

Heat Shock Protein ◽

Synovial Sarcoma ◽

Histone Deacetylase ◽

Histone Deacetylase Inhibitors ◽

Hdac Inhibitors ◽

Heat Shock Protein 90 ◽

Hsp90 Inhibitor ◽

Therapeutic Benefit ◽

Multiple Time

Current systemic therapies have little curative benefit for synovial sarcoma. Histone deacetylase (HDAC) inhibitors and the heat shock protein 90 (Hsp90) inhibitor 17-AAG have recently been shown to inhibit synovial sarcoma in preclinical models. We tested combinations of 17-AAG with the HDAC inhibitor MS-275 for synergism by proliferation and apoptosis assays. The combination was found to be synergistic at multiple time points in two synovial sarcoma cell lines. Previous studies have shown that HDAC inhibitors not only induce cell death but also activate the survival pathway NF-κB, potentially limiting therapeutic benefit. As 17-AAG inhibits activators of NF-κB, we tested if 17-AAG synergizes with MS-275 through abrogating NF-κB activation. In our assays, adding 17-AAG blocks NF-κB activation by MS-275 and siRNA directed against histone deacetylase 3 (HDAC3) recapitulates the effects of MS-275. Additionally, we find that the NF-κB inhibitor BAY 11-7085 synergizes with MS-275. We conclude that agents inhibiting NF-κB synergize with HDAC inhibitors against synovial sarcoma.

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Design, synthesis and evaluation of belinostat analogs as histone deacetylase inhibitors

Future Medicinal Chemistry ◽

10.4155/fmc-2018-0587 ◽

2019 ◽

Vol 11 (21) ◽

pp. 2765-2778

Author(s):

Jie-Huan Zhang ◽

Madhusoodanan Mottamal ◽

Hai-Shan Jin ◽

Shanchun Guo ◽

Yan Gu ◽

...

Keyword(s):

Histone Deacetylase ◽

Active Compound ◽

Histone Deacetylase Inhibitors ◽

Hdac Inhibitors ◽

Antitumor Therapy ◽

Design Synthesis ◽

Inhibitory Activities ◽

Antiproliferative Activities

Aim: Histone deacetylase (HDAC) is an attractive target for antitumor therapy. Therefore, the development of novel HDAC inhibitors is warranted. Materials & methods: A series of HDAC inhibitors based on N-hydroxycinnamamide fragment was designed as the clinically used belinostat analog using amide as the connecting unit. All target compounds were evaluated for their in vitro HDAC inhibitory activities and some selected compounds were tested for their antiproliferative activities. Conclusion: Among them, compound 7e showed an IC50 value of 11.5 nM in inhibiting the HDAC in a pan-HDAC assay, being the most active compound of the series.

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Development of hydroxamate-based histone deacetylase inhibitors of bis-substituted aromatic amides with antitumor activities

MedChemComm ◽

10.1039/c9md00306a ◽

2019 ◽

Vol 10 (10) ◽

pp. 1828-1837 ◽

Cited By ~ 2

Author(s):

Di Ge ◽

Lina Han ◽

Feifei Yang ◽

Na Zhao ◽

Yang Yang ◽

...

Keyword(s):

Histone Deacetylase ◽

Histone Deacetylase Inhibitors ◽

Hdac Inhibitors ◽

Antitumor Activities ◽

Aromatic Amide

Previously, we designed and synthesized a series of bis-substituted aromatic amide-based histone deacetylase (HDAC) inhibitors.

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Mechanism of Action for HDAC Inhibitors—Insights from Omics Approaches

International Journal of Molecular Sciences ◽

10.3390/ijms20071616 ◽

2019 ◽

Vol 20 (7) ◽

pp. 1616 ◽

Cited By ~ 11

Author(s):

Wenbo Li ◽

Zheng Sun

Keyword(s):

Clinical Trials ◽

Enzymatic Activity ◽

Histone Deacetylase ◽

Mechanism Of Action ◽

Histone Deacetylase Inhibitors ◽

Catalytic Domain ◽

Hdac Inhibitors ◽

Mechanistic Studies ◽

Low Toxicity ◽

Strict Specificity

Histone deacetylase inhibitors (HDIs) are a class of prominent epigenetic drugs that are currently being tested in hundreds of clinical trials against a variety of diseases. A few compounds have already been approved for treating lymphoma or myeloma. HDIs bind to the zinc-containing catalytic domain of the histone deacetylase (HDACs) and they repress the deacetylase enzymatic activity. The broad therapeutic effect of HDIs with seemingly low toxicity is somewhat puzzling when considering that most HDIs lack strict specificity toward any individual HDAC and, even if they do, each individual HDAC has diverse functions under different physiology scenarios. Here, we review recent mechanistic studies using omics approaches, including epigenomics, transcriptomics, proteomics, metabolomics, and chemoproteomics, methods. These omics studies provide non-biased insights into the mechanism of action for HDIs.

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Histone Deacetylase Inhibitors Down-Regulate bcl-2 Expression and Induce Apoptosis in t(14;18) Lymphomas

Molecular and Cellular Biology ◽

10.1128/mcb.25.5.1608-1619.2005 ◽

2005 ◽

Vol 25 (5) ◽

pp. 1608-1619 ◽

Cited By ~ 184

Author(s):

Hong Duan ◽

Caroline A. Heckman ◽

Linda M. Boxer

Keyword(s):

Cell Cycle ◽

Histone Deacetylase ◽

Chromatin Immunoprecipitation ◽

Histone Deacetylase Inhibitors ◽

Antitumor Agents ◽

Trichostatin A ◽

Hdac Inhibitors ◽

Transcriptional Level ◽

Cycle Arrest ◽

Induced Apoptosis

ABSTRACT Histone deacetylase (HDAC) inhibitors are promising antitumor agents, but they have not been extensively explored in B-cell lymphomas. Many of these lymphomas have the t(14;18) translocation, which results in increased bcl-2 expression and resistance to apoptosis. In this study, we examined the effects of two structurally different HDAC inhibitors, trichostatin A (TSA) and sodium butyrate (NaB), on the cell cycle, apoptosis, and bcl-2 expression in t(14;18) lymphoma cells. We found that in addition to potent cell cycle arrest, TSA and NaB also dramatically induced apoptosis and down-regulated bcl-2 expression, and overexpression of bcl-2 inhibited TSA-induced apoptosis. The repression of bcl-2 by TSA occurred at the transcriptional level. Western blot analysis and quantitative chromatin immunoprecipitation (ChIP) assay showed that even though HDAC inhibitors increased overall acetylation of histones, localized histone H3 deacetylation occurred at both bcl-2 promoters. TSA treatment increased the acetylation of the transcription factors Sp1 and C/EBPα and decreased their binding as well as the binding of CBP and HDAC2 to the bcl-2 promoters. Mutation of Sp1 and C/EBPα binding sites reduced the TSA-induced repression of bcl-2 promoter activity. This study provides a mechanistic rationale for the use of HDAC inhibitors in the treatment of human t(14;18) lymphomas.

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Cystathionine γ-lyase protects vascular endothelium: a role for inhibition of histone deacetylase 6

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00724.2016 ◽

2017 ◽

Vol 312 (4) ◽

pp. H711-H720 ◽

Cited By ~ 20

Author(s):

Thorsten M. Leucker ◽

Yohei Nomura ◽

Jae Hyung Kim ◽

Anil Bhatta ◽

Victor Wang ◽

...

Keyword(s):

Endothelial Cells ◽

Histone Deacetylase ◽

Vascular Endothelium ◽

High Fat Diet ◽

Histone Deacetylases ◽

Molecular Mechanisms ◽

Oxidized Ldl ◽

Hdac Inhibitors ◽

Histone Deacetylase 6 ◽

High Fat

Endothelial cystathionine γ-lyase (CSEγ) contributes to cardiovascular homeostasis, mainly through production of H2S. However, the molecular mechanisms that control CSEγ gene expression in the endothelium during cardiovascular diseases are unclear. The aim of the current study is to determine the role of specific histone deacetylases (HDACs) in the regulation of endothelial CSEγ. Reduced CSEγ mRNA expression and protein abundance were observed in human aortic endothelial cells (HAEC) exposed to oxidized LDL (OxLDL) and in aortas from atherogenic apolipoprotein E knockout (ApoE−/−) mice fed a high-fat diet compared with controls. Intact murine aortic rings exposed to OxLDL (50 μg/ml) for 24 h exhibited impaired endothelium-dependent vasorelaxation that was blocked by CSEγ overexpression or the H2S donor NaHS. CSEγ expression was upregulated by pan-HDAC inhibitors and by class II-specific HDAC inhibitors, but not by other class-specific inhibitors. The HDAC6 selective inhibitor tubacin and HDAC6-specific siRNA increased CSEγ expression and blocked OxLDL-mediated reductions in endothelial CSEγ expression and CSEγ promoter activity, indicating that HDAC6 is a specific regulator of CSEγ expression. Consistent with this finding, HDAC6 mRNA, protein expression, and activity were upregulated in OxLDL-exposed HAEC, but not in human aortic smooth muscle cells. HDAC6 protein levels in aortas from high-fat diet-fed ApoE−/− mice were comparable to those in controls, whereas HDAC6 activity was robustly upregulated. Together, our findings indicate that HDAC6 is upregulated by atherogenic stimuli via posttranslational modifications and is a critical regulator of CSEγ expression in vascular endothelium. Inhibition of HDAC6 activity may improve endothelial function and prevent or reverse the development of atherosclerosis. NEW & NOTEWORTHY Oxidative injury to endothelial cells by oxidized LDL reduced cystathionine γ-lyase (CSEγ) expression and H2S production, leading to endothelial dysfunction, which was prevented by histone deacetylase 6 (HDAC6) inhibition. Our data suggest HDAC6 as a novel therapeutic target to prevent the development of atherosclerosis.

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