Histone Deacetylase Activity Research Articles

109 (PB099) - A novel dual PARP-HDAC inhibitor for treatment of Ewing sarcoma

Background: Inhibition of poly-adenosine diphosphate-ribose polymerase (PARP) is an effective therapy against cancers with DNA damage repair (DDR) deficiencies, such as BRCA1 and BRCA2 defects. In preclinical studies, PARP inhibitors demonstrated potential therapeutic value in Ewing sarcoma (ES), but clinical trials with olaparib failed to show significant clinical benefit. A key regulatory event in DNA damage repair is acetylation and deacetylation of histones, controlled by histone acetyltransferases (HATs) and histone deacetylases (HDACs). Increased expression of HDACs correlate with more malignant phenotypes in sarcomas and inhibition of HDAC in pre-clinical models of ES inhibits tumor growth. HDAC inhibition combined with PARP inhibition has promising results in vitro, however clinically, combination therapies often require complicated sequential dosing protocols due to different pharmacokinetic profiles and overlapping toxicities, that severely limits clinical utility. Here, we report the efficacy of a novel bifunctional small-molecule compound designed to have both PARP and HDAC inhibition activity. Methods: PARP1 and PARP2 activity was measured using the Trevigen Universal Colorimetric PARP Assay Kit, the BPS Bioscience PARP2 Colorimetric PARP2 Assay Kit, and by PARylation assays. HDAC activity was measured using HeLa cell nuclear extracts and a fluorogenic peptide-based biochemical assay. Cell survival EC50 s were determined using live cell imaging with an Incucyte S3 system and the CellTiter Glo viability assay. DNA damage was detected by western blot, immuno-fluorescence, and comet assays. Results: A representative compound from the kt-3000 series showed potent inhibition of PARP1 and PARP2 with IC50 values in the low nM range, comparable to FDA-approved PARP inhibitors. The compound also showed inhibition of HDAC enzymes with IC50 values in the low μM range, slightly lower than the FDA-approved HDAC inhibitor, vorinostat. Cell survival EC50 values were superior to olaparib in ES cell lines in vitro. Treatment with the kt-3000 compound also resulted in increased DNA damage and S and G2/M cell cycle arrest as compared to olaparib. Conclusion: Our kt-3000 compound shows potent inhibition of PARP1, PARP2, and HDAC, as well as induction of DNA damage and cell cycle arrest. Further development of these bifunctional single molecule inhibitors may offer a novel treatment opportunity for Ewing sarcoma and other solid tumors with limited responses to PARPi monotherapies. Conflict of interest: Board of Directors: J. Bacha and D. Brown are directors of Rakovina Therapeutics. Corporate-sponsored Research: S. Truong, B. Zhai, F. Ghaidi, L. Ramos, J. Joshi are employees of Rakovina Therapeutics. J. Langlands, M. Daugaard and W. Shen are consultants to Rakovina Therapeutics.

PURINE-DERIVED PHENYLHYDROXAMATE COMPOUNDS INDUCES CELL DEATH IN HEMATOLOGICAL NEOPLASM MODELS

The histone acetylation process is the best-studied type of post-translational modification, playing a key role in chromatin remodeling regulated by the activity of two enzymes: histone acetyltransferases and histone deacetylases (HDAC). High levels of HDAC have been observed in patients with hematological and solid cancers and are associated with poor prognosis. The present study aimed to evaluate the cellular effects of three novel potential HDAC inhibitors in hematological neoplasm models. A panel containing myeloid (OCI-AML3, Kasumi 1, HL60, THP1, MOLM13, MV4-11, U937, NB4, NB4-R2, K562, KU812, SET2, and HEL) and lymphoid (Jurkat, CEM, Namalwa, NALM6, Daudi, Raji, SUP-B15, REH, U266, MM1.S, MM1.R, and Karpas 422) neoplasm cells were used. Peripheral blood mononuclear cells (normal leukocytes) from healthy donors were used for toxicity evaluation (n = 4). Three novel purine-derived phenylhydroxamate with potential HDAC inhibitor activity (named here as 82, 83, and 84) were used. Vorinostat was used as a reference drug. Cell viability was assessed by MTT assay, apoptosis by annexin V/propidium iodide labeling and flow cytometry (FC), clonogenicity by autonomous colony formation, cell cycle by propidium iodide and FC, and protein expression by Western Blot. IC 50 values were obtained by non-linear regression. Statistical analysis was performed by ANOVA and Bonferroni post-test. A p < 0.05 was considered significant. All compounds tested reduced cell viability at submicromolar concentrations being compound 82 more potent than the others exhibiting less concentration necessary to reduce cell viability by 50% (IC 50 ranged from 0.09 to 2.04 μM). THP1, MOLM13, NB4-R2, and HEL leukemia cells were more sensitive to these compounds and were selected for the next experiments. In normal leukocytes, vorinostat and compound 82 did not reduce cell viability at the same extension as observed for leukemia cells, indicating a selective antineoplastic activity. All compounds induced time- and dose-dependent manner apoptosis in THP1, MOLM13, NB4-R2, and HEL cells (all p < 0.05). Vorinostat and compound 82 strongly reduced colony formation, being compound 82 more efficient, especially in MOLM13 and NB4-R2 cells (all p < 0.05). Low concentration of vorinostat and compound 82 lead to reduced cell cycle progression (all p < 0.05). THP1, MOLM13, NB4-R2, and HEL cells were exposed to vorinostat or compound 82 and the expression of proteins involved in apoptosis and HDAC activity was modulated. Our results indicate that the three purine-derived phenylhydroxamate tested presented antineoplastic effects being a future candidate to compose the list of HDAC inhibitors in the antineoplastic arsenal. CNPq, CAPES, and FAPESP.

O.13 Givinostat in DMD: results of the Epidys Study

Givinostat is a novel orally active histone deacetylase (HDAC) inhibitor developed for the treatment of DMD. The Epidys Study is a randomized, double blind, placebo controlled, multicenter Phase 3 study to evaluate the efficacy and safety of givinostat in ambulant DMD patients. One-hundred and seventy-nine DMD boys aged ≥6 years at baseline were enrolled and 95% of them completed the study. The primary efficacy assessment was the time to climb 4 standard stairs (4SC). The primary endpoint was complemented by six key secondary efficacy endpoints: time to rise from floor, distance walked in 6 minutes using 6-Minute Walk Test, physical function assessed by North Star Ambulatory Assessment (NSAA), knee extension and elbow flexion muscle strength, and fat fraction of vastus lateralis muscle (VL MFF) evaluated by magnetic resonance spectroscopy. Treatment duration was 18 months. Epidys Study met the primary endpoint with a significant difference between treatment and placebo arm in change from baseline at 18 months in 4SC (GLSmean ratio [SD] = 0.86 [0.071]; p=0.0345). Overall, treatment effect estimates relating to the key secondary endpoints also consistently favoured givinostat over placebo supporting the primary endpoint result, with treatment effects on total NSAA, cumulative loss of function, and VL MFF reaching nominal statistical significance with p <0.05. Safety and tolerability profile was acceptable. In conclusion, Epidys Study met its primary endpoint with consistent results in the key secondary endpoints. Safety and tolerability profile in Epidys Study was consistent with previous studies of givinostat.

Selenium-sensitive histone deacetylase 2 is required for forkhead box O3A and regulates extracellular matrix metabolism in cartilage.

Selenium (Se) as well as selenoproteins are vital for osteochondral system development. Se deficiency (SeD) has a definite impact on the expression and activity of histone deacetylases (HDACs). Abnormal expression of some HDACs affects cartilage development. This current study aims to explore the relationship between differentially expressed HDACs and cartilage development, especially extracellular matrix (ECM) homeostasis maintenance, under SeD conditions. Dark Agouti rats and C28/I2 cell line under SeD states were used to detect the differently expressed HDAC by RT-qPCR, western blotting and IHC staining. Meanwhile, the biological roles of the above HDAC in cartilage development and homeostasis maintenance were confirmed by siRNA transfection, western blotting, RNA sequence and inhibitor treatment experiments. HDAC2 exhibited lower expression at protein level in both animals and chondrocytes during SeD condition. The results of cell-level experiments indicated that forkhead box O3A (FOXO3A), which was required to maintain metabolic homeostasis of cartilage matrix, was reduced by HDAC2 knockdown. Meanwhile, induced HDAC2 was positively associated with FOXO3A in rat SeD model. Meanwhile, knockdown of HDAC2 and FOXO3A led to an increase of intracellular ROS level, which activated NF-κB pathway. Se supplementary significantly inhibited the activation of NF-κB pathway with IL-1β treatment. Our results suggested that low expression of HDAC2 under SeD condition increased ROS content by decreasing FOXO3A in chondrocytes, which led to the activation of NF-κB pathway and ECM homeostasis imbalance.

Abstract A024: In vitro efficacy of a novel dual PARP-HDAC inhibitor in ewing sarcoma

Abstract Introduction: Inhibition of poly-adenosine diphosphate-ribose polymerase (PARP) is an effective therapy against cancers with DNA damage repair (DDR) deficiencies, such as BRCA1 and BRCA2 defects. In preclinical studies, PARP inhibitors demonstrated potential therapeutic value in Ewing sarcoma (ES), though clinical trials with olaparib failed to show significant clinical benefit. While single agent therapy proved inefficacious in the clinical treatment of ES, combination therapies may show anti-tumour activity. A key regulatory event in DNA damage repair is acetylation and deacetylation of histones, controlled by histone acetyltransferases (HATs) and histone deacetylases (HDACs). Increased expression of HDACs have been correlated to more malignant phenotypes in sarcomas and inhibition of HDAC in ES has been shown to be effective in inhibiting tumor growth. HDAC inhibition combined with PARP inhibition has been shown to sensitize cells to treatment in vitro, however clinically, combination therapies often require sequential administration due to different pharmacokinetic profiles and overlapping toxicities, severely limiting clinical utility. Here, we evaluate the activity and efficacy of a novel bifunctional small-molecule compound designed to have both PARP and HDAC inhibiting activity. Methods: PARP1 activity was measured using the Trevigen Universal Colorimetric PARP Assay Kit and PARP2 activity was measured using the BPS Bioscience PARP2 Colorimetric PARP2 Assay Kit. HDAC activity was measured using HeLa nuclear extracts and a fluorogenic peptide-based biochemical assay. Cell survival EC50s were determined using live cell imaging with an Incucyte S3 system and the CellTiter Glo viability assay. Accumulation of phospho-histone H2AX (pH2AX) was detected by western blot using anti-phospho histone H2AX (Ser139) antibody from Cell Signaling Technologies. Results: A representative compound from the kt-3000 series showed potent inhibition of PARP1 and PARP2 with IC50 values in the low nM range, comparable to FDA-approved PARP inhibitors. The compound also showed inhibition of HDAC enzymes with IC50 values in the low µM range, slightly lower than the FDA-approved HDAC inhibitor, vorinostat. Cell survival EC50 values were superior to olaparib in ES cell lines in vitro. Treatment with the kt-3000 compound also resulted in the increased accumulation of pH2AX by western blot and increased S and G2/M cell cycle arrest compared to olaparib. Conclusion: Our kt-3000 compound shows potent inhibition of PARP1, PARP2, and HDAC, as well as induction of DNA damage and cell cycle arrest. Further development of these bifunctional single molecule inhibitors may result in a novel treatment opportunity for Ewing sarcoma. Citation Format: Sarah Truong, Beibei Zhai, Fariba Ghaidi, Louise Ramos, Jay Joshi, Dennis Brown, Neil Sankar, John Langlands, Jeffrey Bacha, Wang Shen, Poul Sorensen, Mads Daugaard. In vitro efficacy of a novel dual PARP-HDAC inhibitor in ewing sarcoma [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr A024.

From HDAC to Voltage-Gated Ion Channels: What's Next? The Long Road of Antiepileptic Drugs Repositioning in Cancer.

Simple SummaryAlthough in the last decades the clinical outcome of cancer patients considerably improved, the major drawbacks still associated with chemotherapy are the unwanted side effects and the development of drug resistance. Therefore, a continuous effort in trying to discover new tumor markers, possibly of diagnostic, prognostic and therapeutic value, is being made. This review is aimed at highlighting the anti-tumor activity that several antiepileptic drugs (AEDs) exert in breast, prostate and other types of cancers, mainly focusing on their ability to block the voltage-gated Na+ and Ca++ channels, as well as to inhibit the activity of histone deacetylases (HDACs), all well-documented tumor markers and/or molecular targets. The existence of additional AEDs molecular targets is highly suspected. Therefore, the repurposing of already available drugs as adjuvants in cancer treatment would have several advantages, such as reductions in dose-related toxicity CVs will be sent in a separate mail to the indicated address of combined treatments, lower production costs, and faster approval for clinical use.Cancer is a major health burden worldwide. Although the plethora of molecular targets identified in the last decades and the deriving developed treatments, which significantly improved patients’ outcome, the occurrence of resistance to therapies remains the major cause of relapse and mortality. Thus, efforts in identifying new markers to be exploited as molecular targets in cancer therapy are needed. This review will first give a glance on the diagnostic and therapeutic significance of histone deacetylase (HDAC) and voltage gated ion channels (VGICs) in cancer. Nevertheless, HDAC and VGICs have also been reported as molecular targets through which antiepileptic drugs (AEDs) seem to exert their anticancer activity. This should be claimed as a great advantage. Indeed, due to the slowness of drug approval procedures, the attempt to turn to off-label use of already approved medicines would be highly preferable. Therefore, an updated and accurate overview of both preclinical and clinical data of commonly prescribed AEDs (mainly valproic acid, lamotrigine, carbamazepine, phenytoin and gabapentin) in breast, prostate, brain and other cancers will follow. Finally, a glance at the emerging attempt to administer AEDs by means of opportunely designed drug delivery systems (DDSs), so to limit toxicity and improve bioavailability, is also given.

Valeric acid delays aril breakdown of longan (Dimocarpus longan Lour.) fruit in relation to the regulation of histone deacetylase activity

Aril breakdown results in the loss of nutrition and commodity value, thereby limits the postharvest storage of longan fruit. Histone deacetylases (HDACs) are important epigenetic regulators in many biological processes. Results in the current study showed that application of valeric acid (VA), which belongs to HDAC inhibitors, suppressed longan aril breakdown during postharvest storage. Meanwhile, lower contents of malondialdehyde, H2O2, and O2−., but higher activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione peroxidase (GPX) were observed in VA-treated fruit. Western blotting (WB) assay confirmed enhancement of histone H3 and H4 acetylation in VA-treated fruit, suggesting the possible inhibition of HDAC activity by VA. Furthermore, VA regulated DlHDACs expressions, with DlHDA5 and DlHDA8 expressions apposite of WB results. These results revealed that VA inhibited longan aril breakdown through enhancing the reactive oxygen species scavenging ability in relation to HDAC inhibitor activity. Our findings suggested that VA treatment delayed aril breakdown of longan fruit in relation to histone deacetylase-mediated epigenetic regulation and highlighted DlHDA5 and DlHDA8 potentially be important candidates for further research.

Mangifera zeylanica and Mangifera indica Leaf Extracts Exert Anticancer Effects Possibly by Inhibiting Histone Deacetylases

ABSTRACT Mangifera indica L. is a popular and widely available plant. Mangifera zeylanica Hook.f. is a plant endemic to Sri Lanka. Both plants bare edible fruits and possess medicinal properties. In the present study, potential anticancer effects of M. zeylanica and M. indica leaves were investigated in lung cancer cells. Hexane, chloroform, ethyl acetate, and methanol extracts of leaves from both plants were obtained using sequential extraction. Chloroform extract of M. zeylanica leaves (MZC) and ethyl acetate extract of M. indica leaves (MIEA) inhibited the proliferation, migration, and colony formation. These extracts showed low cytotoxicity in MRC-5 normal lung fibroblast cells. Furthermore, DNA fragmentation analysis and ethidium bromide/acridine orange (AO/EB) staining showed that MZC and MIEA can induce apoptosis. Gene expression analysis by real-time PCR identified that MZC and MIEA can regulate the expression of apoptosis-associated genes p53, Bax, and survivin. In addition, treatment of both MZC and MIEA decreased histone deacetylase (HDAC) activity, indicating the HDAC inhibitory potentials of leaf extracts. Liquid chromatography-mass spectrometry (LC-MS) identified the presence of mangiferin, 6-geranylnaringenin, nodakenin, and pyrogallol in both leaf extracts as major phytochemicals. Overall, our findings indicate that M. zeylanica and M. indica leaf extracts exert anticancer activity by decreasing the proliferation of non-small lung cancer cells possibly through the inhibition of the activity of HDACs, a major class of enzymes implicated in epigenetic modifications. This investigation provides a rationale to isolate compounds with anticancer and HDAC inhibitory potentials from M. zeylanica and M. indica leaves.

Lactate increases stemness of CD8 + T cells to augment anti-tumor immunity

Lactate is a key metabolite produced from glycolytic metabolism of glucose molecules, yet it also serves as a primary carbon fuel source for many cell types. In the tumor-immune microenvironment, effect of lactate on cancer and immune cells can be highly complex and hard to decipher, which is further confounded by acidic protons, a co-product of glycolysis. Here we show that lactate is able to increase stemness of CD8+ T cells and augments anti-tumor immunity. Subcutaneous administration of sodium lactate but not glucose to mice bearing transplanted MC38 tumors results in CD8+ T cell-dependent tumor growth inhibition. Single cell transcriptomics analysis reveals increased proportion of stem-like TCF-1-expressing CD8+ T cells among intra-tumoral CD3+ cells, a phenotype validated by in vitro lactate treatment of T cells. Mechanistically, lactate inhibits histone deacetylase activity, which results in increased acetylation at H3K27 of the Tcf7 super enhancer locus, leading to increased Tcf7 gene expression. CD8+ T cells in vitro pre-treated with lactate efficiently inhibit tumor growth upon adoptive transfer to tumor-bearing mice. Our results provide evidence for an intrinsic role of lactate in anti-tumor immunity independent of the pH-dependent effect of lactic acid, and might advance cancer immune therapy.

Intra-mPFC injection of sodium butyrate promotes BDNF expression and ameliorates extinction recall impairment in an experimental paradigm of post-traumatic stress disorder.

Objective(s):Therapeutic strategies that facilitate extinction are promising in the treatment of post-traumatic stress disorder (PTSD). Brain-derived neurotrophic factor (BDNF) has a crucial role in neural plasticity, a process needed for the retention of fear extinction. In this study, we investigated the effects of local administration of a histone deacetylase (HDAC) inhibitor, sodium butyrate (NaBu), on BDNF transcription and behavioral markers of extinction in the single prolonged stress (SPS) model of PTSD. Materials and Methods:NaBu was infused into the infralimbic (IL) subregion of the medial prefrontal cortex (mPFC) of male rats. The freezing response was recorded as the criterion to assess fear strength on the day of extinction as well as 24 hr later in the retention test. Other behavioral tests were also measured to evaluate the anxiety level, locomotor activity, and working memory on the retention day. HDAC activity and BDNF mRNA expression were evaluated after the behavioral experiments. Results: NaBu facilitated the recall of fear extinction in SPS rats (P<0.0001). SPS rats had higher HDAC activity (P<0.0001) and lower BDNF expression (P<0.05) than non-SPS animals. Also, anxiety was higher in the SPS group (P<0.0001), but locomotor activity (P=0.61) and working memory (P=0.36) were not different between SPS and Non-SPS groups.Conclusion:Our findings provide evidence that the mechanism of action of NaBu in the improvement of extinction recall is mediated, in part, by enhancing histone acetylation and reviving BDNF expression in IL.

Synthetic Study on Bicyclic Depsipeptides Containing an Intramolecular Disulfide Bond

Bicyclic depsipeptide natural products containing an intramolecular disulfide bond are potent histone deacetylase (HDAC) inhibitors. Among them, FK228 (romidepsin) is approved for treating cutaneous T-cell lymphoma and peripheral T-cell lymphoma. This study focused on developing a new synthesis method for producing this class of natural products for use as HDAC inhibitors with high efficacy and low toxicity. In this paper, the total syntheses of FK228 as well as spiruchostatins A and B are described. The synthesis routes include a convergent way to assemble seco-acids via the amide condensation of amine segments with carboxylic acid segments. The syntheses of C4- and C7-modified FK228 analogs (FK-A1 to FK-A8) are also described. The evaluation of HDAC and cell growth inhibitory activities of the synthesized analogs revealed novel aspects of their structure-activity relationship. Potent and highly isoform-selective HDAC1 inhibitors were identified. Furthermore, the analogs showed phosphatidylinositol 3-kinase (PI3K) inhibitory activity. Structural optimization of the analogs as HDAC/PI3K dual inhibitors led to the identification of FK-A11 as the most potent analog.

Inhibition of Class I Histone Deacetylase Activity Blocks the Induction of TNFAIP3 Both Directly and Indirectly via the Suppression of Endogenous TNF-α

Histone deacetylase inhibitors (HDIs) are promising drugs for the treatment of inflammatory diseases. However, their therapeutical exploitation is slowed down by severe adverse manifestations that can hardly be foreseen, mainly due to incomplete knowledge of how HDIs impact the delicate balance of inflammatory mediators. In this work, we characterized the effects of the HDI trichostatin A (TSA) on the expression of TNFAIP3, which is a crucial inhibitor of the classical NF-kB pathway and an LPS-induced negative feedback regulator. The accumulation of TNFAIP3 mRNA after LPS stimulation showed biphasic behavior, with one wave within the first hour of stimulation and a second wave several hours later, which were both reduced by TSA. By using inhibition and knockdown approaches, we identified two temporally and mechanistically distinct modes of action. The first wave of TNAIP3 accumulation was directly blunted by the histone deacetylase (HDAC) blockade. By contrast, the second wave was decreased mainly because of the lack of endogenous TNF-α induction, which, in turn, depended on the intact HDAC activity. In both cases, class I HDACs appeared to play a nonredundant role, with HDAC3 required, but not sufficient, for TNF-α and TNFAIP3 induction. In addition to TNFAIP3, TNF-α is known to induce many response genes that orchestrate the inflammatory cascade. Thus, suppression of TNF-α may represent a general mechanism through which HDIs regulate a selected set of target genes.

Roles and regulation of histone acetylation in hepatocellular carcinoma.

Hepatocellular Carcinoma (HCC) is the most frequent malignant tumor of the liver, but its prognosis is poor. Histone acetylation is an important epigenetic regulatory mode that modulates chromatin structure and transcriptional status to control gene expression in eukaryotic cells. Generally, histone acetylation and deacetylation processes are controlled by the opposing activities of histone acetyltransferases (HATs) and histone deacetylases (HDACs). Dysregulation of histone modification is reported to drive aberrant transcriptional programmes that facilitate liver cancer onset and progression. Emerging studies have demonstrated that several HDAC inhibitors exert tumor-suppressive properties via activation of various cell death molecular pathways in HCC. However, the complexity involved in the epigenetic transcription modifications and non-epigenetic cellular signaling processes limit their potential clinical applications. This review brings an in-depth view of the oncogenic mechanisms reported to be related to aberrant HCC-associated histone acetylation, which might provide new insights into the effective therapeutic strategies to prevent and treat HCC.

Ibrutinib combined with low-dose histone deacetylases inhibitor chidamide synergistically enhances the anti-tumor effect in B-cell lymphoma.

Aberrant activity of histone deacetylases (HDACs) is frequently detected in B-cell lymphomas, which indicated the therapeutic implications of HDAC inhibitors for B-cell malignancies. We have discovered that lymphoma cells treated with HDAC inhibitor presented with activation of Bruton tyrosine kinase (BTK) which played an important role in the development of B-cell malignancies. Therefore, our study intended to explore whether the addition of ibrutinib (BTK inhibitor) to chidamide (HDAC inhibitor) could generate combined anti-tumor effects in B-cell lymphomas. Using cell viability assay, cell cycle and apoptosis kit, we demonstrated an evident synergistic action of ibrutinib and chidamide in inhibiting tumor cell proliferation and motility. Consistent with in vitro data, the synergistic anti-tumor effects were also observed in multiple tumor-bearing mice models. By performing RNA-seq and flow cytometry of tumor tissue, the enhancement of anti-tumor immunity was observed with the co-treatment of chidamide and ibrutinib. Together, these mechanistic insights indicated that simultaneously targeting BTK and HDAC could be a promising clinical therapy for B-cell lymphomas.

Sulforaphane inhibits angiotensin II-induced cardiomyocyte apoptosis by acetylation modification of Nrf2.

Oxidative stress is the central cause of angiotensin II (Ang II)-induced myocardial injury, and nuclear factor erythroid 2-related factor (Nrf2) is the core molecule of the anti-oxidant defense system. We have previously demonstrated that sulforaphane (SFN) can prevent Ang II-induced myocardial injury by activating Nrf2; however, the underlying molecular mechanism is still unclear. This study aimed to evaluate whether SFN prevents Ang II-induced cardiomyocyte apoptosis through acetylation modification of Nrf2. Wild-type and Nrf2 knockdown embryonic rat cardiomyocytes (H9C2) were exposed to Ang II to induce apoptosis, oxidative stress, and inflammatory responses. SFN treatment significantly reduced Ang II-induced cardiomyocyte apoptosis, inflammation and oxidative stress. Activation of Nrf2 played a critical role in preventing cardiomyocyte apoptosis. After Nrf2 was knockdown, the anti-inflammatory, antioxidant stress of SFN were eliminated. Furthermore, Nrf2 activation by SFN was closely related to the decreased activity of histone deacetylases (HDACs) and increased histone-3 (H3) acetylation levels in Nrf2 promoter region. These findings confirm that Nrf2 plays a key role in SFN preventing Ang II-induced cardiomyocyte apoptosis. SFN activates Nrf2 by inhibiting HDACs expression and activation.

Do biological activities of selective histone deacetylase 6 (HDAC6) inhibitors rely on the modification of cap group?

Nowadays, significant progress has been made in the development of selective histone deacetylase 6 (HDAC6) inhibitors, exerting great potential in the treatment of various malignant tumors and neurodegenerative diseases. Previously, selective inhibitory activities of HDAC inhibitors were generally considered sensitive to the interactions between the Cap group and the binding site of HDAC6, and a large number of selective HDAC6 inhibitors have been designed and synthesized based on the strategy. However, some inhibitors without Cap group could also exhibit excellent potency and selective inhibition towards HDAC6, and in this study, BRD9757 and compound 8, as capless selective HDAC6 inhibitors, were selected as molecular probes to explore the difference of their binding interactions in HDAC1&6. Through the analysis of binding-free energies and conformational rearrangements after 1μs molecular dynamics simulation, it could be learned that although the residues in the binding site remained highly consistent, the binding mechanisms of BRD9757 and compound 8 in HDAC1&6 were different, which will provide valuable hints for the discovery of novel selective HDAC6 inhibitors.

COMBINATION OF HDAC AND NF-KB INHIBITORS DEPLET CANCER STEM CELL POPULATION AND SENSITIZE HNSCC TO CHEMOTHERAPY

<h3>Objectives</h3> Recent evidence suggests a major role for cancer stem cell (CSC) as responsible for tumor progression, recurrence, and chemotherapy and radiotherapy resistance in head and neck squamous cell carcinoma (HNSCC). Epigenetic modifications and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) activation have been associated with CSC behavior and represent promising targets for HNSCC treatment. The aim was to explore the potential of the histone deacetylase (HDAC) and NF-kB inhibitors (SAHA and emetine, respectively) as potential modulators of the CSC population in HNSCC. <h3>Study Design</h3> SAHA and emetine cytotoxic concentration were determined using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) cell viability assay for the SCC-4 and SCC9 HNSCC cell lines. The CSC population was estimated by aldehyde dehydrogenases activity using flow cytometry, tumosphere formation, and clonogenic assay. The drugs were tested isolated and in combination. <h3>Results</h3> Treatment with SAHA and emetine isolated decreased the CSC in SCC-4 and SCC-9 population in all assays. The combination of both drugs led to a more significant reduction in CSC population and sensitivity to chemotherapy with cisplatin. <h3>Conclusions</h3> Our findings suggests that the inhibition of HDAC activity combined with the depletion of NF-kB pathway effectively depleted CSC population and that the double inhibition can be a possible target for new adjuvant therapies in HNSCC.

Potential of histone deacetylase inhibitors in the control and regulation of prostate, breast and ovarian cancer.

Histone deacetylases (HDACs) are enzymes that play a role in chromatin remodeling and epigenetics. They belong to a specific category of enzymes that eliminate the acetyl part of the histones’ -N-acetyl lysine, causing the histones to be wrapped compactly around DNA. Numerous biological processes rely on HDACs, including cell proliferation and differentiation, angiogenesis, metastasis, gene regulation, and transcription. Epigenetic changes, specifically increased expression and activity of HDACs, are commonly detected in cancer. As a result, HDACi could be used to develop anticancer drugs. Although preclinical outcomes with HDACs as monotherapy have been promising clinical trials have had mixed results and limited success. In both preclinical and clinical trials, however, combination therapy with different anticancer medicines has proved to have synergistic effects. Furthermore, these combinations improved efficacy, decreased tumor resistance to therapy, and decreased toxicity. In the present review, the detailed modes of action, classification of HDACs, and their correlation with different cancers like prostate, breast, and ovarian cancer were discussed. Further, the different cell signaling pathways and the structure-activity relationship and pharmaco-toxicological properties of the HDACi, and their synergistic effects with other anticancer drugs observed in recent preclinical and clinical studies used in combination therapy were discussed for prostate, breast, and ovarian cancer treatment.

Environmental enrichment mitigates PTSD-like behaviors in adult male rats exposed to early life stress by regulating histone acetylation in the hippocampus and amygdala

Early life stress (ELS) can cause long-term changes in gene expression, affect cognition, mood, and behavior, and increase susceptibility to post-traumatic stress disorder (PTSD) in adulthood, in which the histone acetylation plays a crucial role. Studies have found that environmental enrichment (EE) mitigated the unfavorable outcomes of ELS. However, the underlying mechanism of the histone acetylation is not yet completely clear. The purpose of this study was to explore the effect of EE on the histone acetylation after ELS. In this study, using single prolonged stress (SPS) paradigm in early adolescent rats explored the long-term effects of ELS on behavior, the activity of histone acetyltransferases (HATs) and histone deacetylases (HDACs), as well as the acetylation levels of the lysine 9 site of histone H3 (H3K9) and lysine 12 site of histone H4 (H4K12) in the hippocampus and amygdala. Meanwhile, the protective effects of EE intervention were examined. We found that adult male rats exposed to ELS showed behavioral changes, including reduced locomotor activity, increased anxiety-like behaviors, impaired spatial learning and memory, enhanced contextual and cued fear memory, and the HATs/HDACs ratio and acetyl H3K9 (Ac–H3K9) and acetyl H4K12 (Ac–H4K12) were increased in the hippocampus and decreased in the amygdala. Furthermore, EE attenuated the behavioral abnormalities from ELS, possibly through down-regulating the activity of HATs in the hippocampus and up-regulating HDACs activities in the amygdala. These finding suggested that EE could ameliorate ELS-induced PTSD-like behaviors by regulating histone acetylation in the hippocampus and amygdala, reducing the susceptibility to PTSD in adulthood.

Balanced control of thermogenesis by nuclear receptor corepressors in brown adipose tissue

Brown adipose tissue (BAT) is a key thermogenic organ whose expression of uncoupling protein 1 (UCP1) and ability to maintain body temperature in response to acute cold exposure require histone deacetylase 3 (HDAC3). HDAC3 exists in tight association with nuclear receptor corepressors (NCoRs) NCoR1 and NCoR2 (also known as silencing mediator of retinoid and thyroid receptors [SMRT]), but the functions of NCoR1/2 in BAT have not been established. Here we report that as expected, genetic loss of NCoR1/2 in BAT (NCoR1/2 BAT-dKO) leads to loss of HDAC3 activity. In addition, HDAC3 is no longer bound at its physiological genomic sites in the absence of NCoR1/2, leading to a shared deregulation of BAT lipid metabolism between NCoR1/2 BAT-dKO and HDAC3 BAT-KO mice. Despite these commonalities, loss of NCoR1/2 in BAT does not phenocopy the cold sensitivity observed in HDAC3 BAT-KO, nor does loss of either corepressor alone. Instead, BAT lacking NCoR1/2 is inflamed, particularly with respect to the interleukin-17 axis that increases thermogenic capacity by enhancing innervation. Integration of BAT RNA sequencing and chromatin immunoprecipitation sequencing data revealed that NCoR1/2 directly regulate Mmp9, which integrates extracellular matrix remodeling and inflammation. These findings reveal pleiotropic functions of the NCoR/HDAC3 corepressor complex in BAT, such that HDAC3-independent suppression of BAT inflammation counterbalances stimulation of HDAC3 activity in the control of thermogenesis.