Induction Of Histone Acetylation Research Articles

Acute depletion of BRG1 reveals its primary function as an activator of transcription

The mammalian SWI/SNF-like BAF complexes play critical roles during animal development and pathological conditions. Previous gene deletion studies and characterization of human gene mutations implicate that the complexes both repress and activate a large number of genes. However, the direct function of the complexes in cells remains largely unclear due to the relatively long-term nature of gene deletion or natural mutation. Here we generate a mouse line by knocking in the auxin-inducible degron tag (AID) to the Smarca4 gene, which encodes BRG1, the essential ATPase subunit of the BAF complexes. We show that the tagged BRG1 can be efficiently depleted by osTIR1 expression and auxin treatment for 6 to 10 h in CD4 + T cells, hepatocytes, and fibroblasts isolated from the knock-in mice. The acute depletion of BRG1 leads to decreases in nascent RNAs and RNA polymerase II binding at a large number of genes, which are positively correlated with the loss of BRG1. Further, these changes are correlated with diminished accessibility at DNase I Hypersensitive Sites (DHSs) and p300 binding. The acute BRG1 depletion results in three major patterns of nucleosome shifts leading to narrower nucleosome spacing surrounding transcription factor motifs and at enhancers and transcription start sites (TSSs), which are correlated with loss of BRG1, decreased chromatin accessibility and decreased nascent RNAs. Acute depletion of BRG1 severely compromises the Trichostatin A (TSA) -induced histone acetylation, suggesting a substantial interplay between the chromatin remodeling activity of BRG1 and histone acetylation. Our data suggest BRG1 mainly plays a direct positive role in chromatin accessibility, RNAPII binding, and nascent RNA production by regulating nucleosome positioning and facilitating transcription factor binding to their target sites.

Abstract 6031: Fasting associated microbiota contributes to small intestinal radioprotection by modulating the epigenome

Abstract Normal tissue toxicity often restricts the administration of effective radiation doses for optimal tumor cell elimination. We identified a novel host-protective effect conferred by short-term fasting when treating mice with high dose radiation. We showed that fasting mice for 24h induces epigenetic and transcriptional changes in small intestinal epithelial cells (siECs) and confers protection to small intestinal stem cells from high dose radiation. Here we report that fasting induces changes in the gut microbiome favoring bacteria producing metabolites, specifically short-chain fatty acids, that induce epigenetic changes in siECs. 16S rRNA sequencing of feces isolated from the small intestines of fed and fasted mice revealed a marked enrichment of Akkermansia muciniphila (AKK) in fasted relative to fed mice. Specific depletion of AKK through antibiotic treatment impaired the ability of fasting to provide organismal radioprotection whereas both organismal and OLFM4+ stem cell protection was restored in fasted animals upon reintroduction of AKK. AKK produces short chain fatty acids (SCFAs) including propionate which is able to induce histone modifications and our metabolomic analysis of feces isolated from the ileums of mice demonstrated a significant increase in propionic acid levels in fasted mice compared to fed mice. Importantly, we showed that AKK conditioned medium as well as its metabolite (propionate) are able to induce histone acetylation in Lgr5+ stem cell enriched epithelial spheroids cultured in vitro. We are currently employing high-dimensional analysis techniques, including single cell analysis, Cut&Tag and RNAseq to determine how the epigenetic and transcriptional changes induced by fasting provide organismal and SI radioprotection. Citation Format: Praveen Barrodia, Emre Arslan, Sabrina Jeter-Jones, Robert R. Jenq, Kunal Rai, Helen Piwnica-Worms. Fasting associated microbiota contributes to small intestinal radioprotection by modulating the epigenome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6031.

NEAT1 repression by MED12 creates chemosensitivity in p53 wild-type breast cancer cells.

Breast cancer is often treated with chemotherapy. However, the development of chemoresistance results in treatment failure. Long non-coding RNA nuclear paraspeckle assembly transcript 1 (NEAT1) has been shown to contribute to chemoresistance in breast cancer cells. In studying the transcriptional regulation of NEAT1 using multi-omics approaches, we showed that NEAT1 is up-regulated by 5-fluorouracil in breast cancer cells with wild-type cellular tumor antigen p53 but not in mutant-p53-expressing breast cancer cells. The regulation of NEAT1 involves mediator complex subunit 12 (MED12)-mediated repression of histone acetylation marks at the promoter region of NEAT1. Knockdown of MED12 but not coactivator-associated arginine methyltransferase 1 (CARM1) induced histone acetylation at the NEAT1 promoter, leading to elevated NEAT1 mRNAs, resulting in a chemoresistant phenotype. The MED12-dependent regulation of NEAT1 differs between wild-type and mutant p53-expressing cells. MED12 depletion led to increased expression of NEAT1 in a wild-type p53 cell line, but decreased expression in a mutant p53 cell line. Chemoresistance caused by MED12 depletion can be partially rescued by NEAT1 knockdown in p53 wild-type cells. Collectively, our study reveals a novel mechanism of chemoresistance dependent on MED12 transcriptional regulation of NEAT1 in p53 wild-type breast cancer cells.

Tamoxifen exerts anti-peritoneal fibrosis effects by inhibiting H19-activated VEGFA transcription

BackgroundPeritoneal dialysis (PD) remains limited due to dialysis failure caused by peritoneal fibrosis. Tamoxifen (TAM), an inhibitor of estrogen receptor 1 (ESR1), has been reported to treat fibrosis, but the underlying mechanism remains unknown. In this study, we sought to explore whether tamoxifen played an anti-fibrotic role by affecting transcription factor ESR1.MethodsESR1 expression was detected in the human peritoneum. Mice were daily intraperitoneally injected with 4.25% glucose PD dialysate containing 40 mM methylglyoxal for 2 weeks to establish PD-induced peritoneal fibrosis. Tamoxifen was administrated by daily gavage, at the dose of 10 mg/kg. Chromatin immunoprecipitation (ChIP) and dual‐luciferase reporter assay were performed to validate ESR1 bound H19 promoter. Gain-of-function and loss-of-function experiments were performed to investigate the biological roles of H19 on the mesothelial-mesenchymal transition (MMT) of human peritoneal mesothelial cells (HPMCs). Intraperitoneal injection of nanomaterial-wrapped 2′-O-Me-modified small interfering RNA was applied to suppress H19 in the mouse peritoneum. RNA immunoprecipitation and RNA pull-down assays demonstrated binding between H19 and p300. Exfoliated peritoneal cells were obtained from peritoneal dialysis effluent to analyze the correlations between ESR1 (or H19) and peritoneal solute transfer rate (PSTR).ResultsESR1 was increased significantly in the peritoneum after long-term exposure to PD dialysate. Tamoxifen treatment ameliorated high glucose-induced MMT of HPMCs, improved ultrafiltration rate, and decreased PSTR of mouse peritoneum. Tamoxifen reduced the H19 level by decreasing the ESR1 transcription of H19. Depletion of H19 reversed the pro-fibrotic effect of high glucose while ectopic expression of H19 exacerbated fibrotic pathological changes. Intraperitoneal injection of nanomaterial-wrapped 2′-O-Me-modified siRNAs targeting H19 mitigated PD-related fibrosis in mice. RNA immunoprecipitation (RIP) and RNA pull-down results delineated that H19 activated VEGFA expression by binding p300 to the VEGFA promoter and inducing histone acetylation of the VEGFA promoter. ESR1 and H19 were promising targets to predict peritoneal function.ConclusionsHigh glucose-induced MMT of peritoneal mesothelial cells in peritoneal dialysis via activating ESR1. In peritoneal mesothelial cells, ESR1 transcribed the H19 and H19 binds to transcription cofactor p300 to activate the VEGFA. Targeting ESR1/H19/VEGFA pathway provided new hope for patients undergoing peritoneal dialysis.Graphic

Repurposing of triamterene as a histone deacetylase inhibitor to overcome cisplatin resistance in lung cancer treatment.

Cisplatin is the core chemotherapeutic drug used for first-line treatment of advanced non-small cell lung cancer (NSCLC). However, drug resistance is severely hindering its clinical efficacy. This study investigated the circumvention of cisplatin resistance by repurposing non-oncology drugs with putative histone deacetylase (HDAC) inhibitory effect. A few clinically approved drugs were identified by a computational drug repurposing tool called "DRUGSURV" and evaluated for HDAC inhibition. Triamterene, originally indicated as a diuretic, was chosen for further investigation in pairs of parental and cisplatin-resistant NSCLC cell lines. Sulforhodamine B assay was used to evaluate cell proliferation. Western blot analysis was performed to examine histone acetylation. Flow cytometry was used to examine apoptosis and cell cycle effects. Chromatin immunoprecipitation was conducted to investigate the interaction of transcription factors to the promoter of genes regulating cisplatin uptake and cell cycle progression. The circumvention of cisplatin resistance by triamterene was further verified in a patient-derived tumor xenograft (PDX) from a cisplatin-refractory NSCLC patient. Triamterene was found to inhibit HDACs. It was shown to enhance cellular cisplatin accumulation and potentiate cisplatin-induced cell cycle arrest, DNA damage, and apoptosis. Mechanistically, triamterene was found to induce histone acetylation in chromatin, thereby reducing the association of HDAC1 but promoting the interaction of Sp1 with the gene promoter of hCTR1 and p21. Triamterene was further shown to potentiate the anti-cancer effect of cisplatin in cisplatin-resistant PDX in vivo. The findings advocate further clinical evaluation of the repurposing use of triamterene to overcome cisplatin resistance.

Inhibition of histone deacetylases attenuates tumor progression and improves immunotherapy in breast cancer.

Breast cancer is one of the common malignancies with poor prognosis worldwide. The treatment of breast cancer patients includes surgery, radiation, hormone therapy, chemotherapy, targeted drug therapy and immunotherapy. In recent years, immunotherapy has potentiated the survival of certain breast cancer patients; however, primary resistance or acquired resistance attenuate the therapeutic outcomes. Histone acetyltransferases induce histone acetylation on lysine residues, which can be reversed by histone deacetylases (HDACs). Dysregulation of HDACs via mutation and abnormal expression contributes to tumorigenesis and tumor progression. Numerous HDAC inhibitors have been developed and exhibited the potent anti-tumor activity in a variety of cancers, including breast cancer. HDAC inhibitors ameliorated immunotherapeutic efficacy in cancer patients. In this review, we discuss the anti-tumor activity of HDAC inhibitors in breast cancer, including dacinostat, belinostat, abexinostat, mocetinotat, panobinostat, romidepsin, entinostat, vorinostat, pracinostat, tubastatin A, trichostatin A, and tucidinostat. Moreover, we uncover the mechanisms of HDAC inhibitors in improving immunotherapy in breast cancer. Furthermore, we highlight that HDAC inhibitors might be potent agents to potentiate immunotherapy in breast cancer.

Histone acetylation modifications: A potential targets for the diagnosis and treatment of papillary thyroid cancer.

Thyroid cancer is a common malignancy of the endocrine system, with papillary thyroid cancer (PTC) being the most common type of pathology. The incidence of PTC is increasing every year. Histone acetylation modification is an important part of epigenetics, regulating histone acetylation levels through histone acetylases and histone deacetylases, which alters the proliferation and differentiation of PTC cells and affects the treatment and prognosis of PTC patients. Histone deacetylase inhibitors induce histone acetylation, resulting in the relaxation of chromatin structure and activation of gene transcription, thereby promoting differentiation, apoptosis, and growth arrest of PTC cells.

First-in-Class Histone Acetyltransferase (HAT) Activator, YF2, Modulates Immune Evasion in DLBCL, Enhancing the Effects of Immune Checkpoint Blockade

We reported the discovery of YF2, a EP300 (p300) and CREBBP (CBP) HAT activator which demonstrates selective cytotoxicity in HAT-mutated DLBCL cell lines and induces histone acetylation in vitro and in vivo. Recently, we have focused on the role of HAT induction in regulating immune response in DLBCLs. CBP has been shown to function as an enhancer that regulates GC B-cell fate by controlling antigen presentation and terminal differentiation. HAT mutations are linked to reduced MHC-II HLA transcripts due to decreased acetylation at the promoter regions which leads to decreased GC B:T cell entanglement and derangements in the GC reaction. Numerous studies have reported the effect of epigenetic drugs on antigen presentation. This response is caused by disruption of the BLC6/SMRT/HDAC3 repressor complex which co-occupies enhancers in the MHC Class II loci. Previously, we demonstrated YF2 induced p300 mediated acetylation of BCL6; abrogating its activity. More recently, we demonstrated through RNA-seq that prolonged treatment of YF2 upregulated the expression of several MHC Class I-II genes resulting in the upregulation of the immune regulation signaling pathway. Further, treatment with YF2 up-regulated MHC-I and -II expression in a dose dependent manner. In addition to regulating immune response, BCL6 is a master regulator of GC formation. We hypothesize that induction of HATs will modulate immune response in splenic and circulating B and T-cells. Immunized BALB/c mice were treated with saline or 50mg/kg YF2 for 10 days. Spleen and blood samples were analyzed by flow cytometry. We observed a significant reduction in splenic weight, B-cell frequency, and GCs formation in YF2 treated mice. YF2 treatment increased the frequency of mature and transitional B-cells in the spleen. Further, YF2 treatment reduced light zone and increased dark zone cells in the spleen. YF2 treatment also increased the frequency of plasma cells and plasmablast. MHC-I and -II expression was significantly upregulated in YF2 treated mice. YF2 exposure resulted in an increase of PD-L1hi B-cells. Lastly, circulating B-cells showed significant increase in MHC-II expression following YF2 treatment. In splenic T-cells, we observed a reduction in CD4+ helper T-cells and an increase in CD8+ cytotoxic T-cells in YF2 treated mice. YF2 caused a reduction in naïve CD4+ cells and an increase in the frequency of central and effector memory cells Lastly, we examined the frequency of inhibitory molecules PD1, TIGIT, CD226, Lag3, and VISTA. PD-1, TIGIT, and CD226 were upregulated with YF2 treatment in CD4+ cells and CD226 was upregulated in CD8+ cells. Together, these results demonstrate YF2 is able to modulate immune response in B and T-cells. DLBCLs are known to have limited sensitivity to checkpoint blockade with PD-1 and PD-L1 inhibitors due to low PD-L1 and PD-1 expression which is in part driven by HAT mutations and overexpression of BCL6; which regulates PD-L1. We hypothesize that combination treatment of YF2 and PD-1 inhibitor in the immunocompetent syngeneic A20 mouse tumor xenograft model will result in decreased tumorigenicity and improved survival in mice. Xenografted and immunized BALBc mice were treated with saline, YF2, PD-1 inhibitor, or a combination of YF2 and PD-1 inhibitor for 28 days. We observed, a significant reduction in tumor volume in both the PD-1 and combination treated groups. Kaplan-Meier analysis showed an increase in median survival for all treatment with the combination treatment resulting in the greatest increase in median survival vs. control (Fig A). Spatial analysis was performed using the 10x Visium platform and sc-RNA-seq to examine the tumor microenvironment in control and YF2 treated mouse tumor samples (Fig B). We observed a more aggressive phenotype in the untreated tumors as well as an increase in the number of distinct clusters indicating a more heterogenous tumor population. Lastly, PBMCs from patient samples were isolated and treated with YF2. Compared to healthy donor patients, YF2 induced cytotoxicity across various lymphoma subtype. Normal PBMCs were not affected by treatment. Based on the success of YF2 in the preclinical setting, the use of this first-in-class drug for treatment of lymphoma was approved to proceed in clinical studies in patients with relapsed/refractory lymphoma. Patient samples will be collected and analyzed for immune response. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

HDAC AND NFKB INHIBITORS REDUCE CANCER STEM CELLS AND SENSITIZE MUCOEPIDERMOID CARCINOMA CELL LINES TO CISPLATIN

Objectives To evaluate the potential of histone deacetylase (HDAC) suberoylanilide hydroxamic acid (SAHA) and nuclear factor kappa-light-chain-enhancer of activated B cells (NFkB) (emetine) inhibitors, isolated and associated with cisplatin (CDDP) in salivary gland mucoepidermoid carcinoma (MEC) cell lines, specifically through cancer stem cell (CSC) modulation. Study Design Cell viability was determined by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay. Immunofluorescence staining was performed to identify the acetylation of lysine 9 from histone H3 protein (H3k9ac) and phosphorylated protein p65 (NFkB pathway). CSC was evaluated by quantifying the enzymatic activity of aldehyde dehydrogenases (ALDH) using flow cytometry and by in vitro tumorsphere formation. Results A single administration of SAHA (1 μM) induced histone acetylation, whereas 0.05 μM of emetine resulted in inhibition of the NF-kB pathway. We showed a disruption of CSC, illustrated by a reduction of enzymatic activity of ALDH and inhibition of tumorsphere formation. Furthermore, we observed that MEC tumor cells were sensitized to CDDP after the administration of a combination of SAHA and emetine. Conclusions Our findings suggest that HDAC and NF-kB inhibitors might be an alternative approach to treat MEC through disruption of CSC and sensitizing cells to conventional therapy. To evaluate the potential of histone deacetylase (HDAC) suberoylanilide hydroxamic acid (SAHA) and nuclear factor kappa-light-chain-enhancer of activated B cells (NFkB) (emetine) inhibitors, isolated and associated with cisplatin (CDDP) in salivary gland mucoepidermoid carcinoma (MEC) cell lines, specifically through cancer stem cell (CSC) modulation. Cell viability was determined by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) assay. Immunofluorescence staining was performed to identify the acetylation of lysine 9 from histone H3 protein (H3k9ac) and phosphorylated protein p65 (NFkB pathway). CSC was evaluated by quantifying the enzymatic activity of aldehyde dehydrogenases (ALDH) using flow cytometry and by in vitro tumorsphere formation. A single administration of SAHA (1 μM) induced histone acetylation, whereas 0.05 μM of emetine resulted in inhibition of the NF-kB pathway. We showed a disruption of CSC, illustrated by a reduction of enzymatic activity of ALDH and inhibition of tumorsphere formation. Furthermore, we observed that MEC tumor cells were sensitized to CDDP after the administration of a combination of SAHA and emetine. Our findings suggest that HDAC and NF-kB inhibitors might be an alternative approach to treat MEC through disruption of CSC and sensitizing cells to conventional therapy.

Comprehensive Transcriptomic Analysis of Novel Class I HDAC Proteolysis Targeting Chimeras (PROTACs).

The class I histone deacetylase (HDAC) enzymes;HDAC1,2 and 3 form the catalytic engine of at least seven structurally distinct multiprotein complexes in cells. These molecular machines play a vital role in the regulation of chromatin accessibility and gene activity via the removal of acetyl moieties from lysine residues within histone tails. Their inhibition via small molecule inhibitors has beneficial effects in a number of disease types, including the clinical treatment of hematological cancers. We have previously reported a library of proteolysis targeting chimeras (PROTACs) incorporating a benzamide-based HDAC ligand (from CI-994), with an alkyl linker and ligand for the von Hippel-Lindau (VHL) E3 ubiquitin ligase that degrade HDAC1-3 at submicromolar concentrations. Here we report the addition of two novel PROTACs (JPS026 and JPS027), which utilize a ligand for the cellular inhibitor of apoptosis (IAP) family of E3 ligases. We found that both VHL (JPS004)- and IAP (JPS026)-based PROTACs degrade HDAC1-3 and induce histone acetylation to a similar degree. However, JPS026 is significantly more potent at inducing cell death in HCT116 cells than is JPS004. RNA sequencing analysis of PROTAC-treated HCT116 cells showed a distinct gene expression signature in which cell cycle and DNA replication machinery are repressed. Components of the mTORC1 and -2 complexes were also reduced, leading to an increase in FOXO3 and downstream target genes that regulate autophagy and apoptosis. In summary, a novel combination of HDAC and IAP ligands generates a PROTAC with a potent ability to stimulate apoptosis and differential gene expression in human cancer cells.

Angiotensin II Mediates Cardiomyocyte Hypertrophy in Atrial Cardiomyopathy via Epigenetic Transcriptional Regulation.

Aims European Heart Rhythm Association established an expert consensus to define, characterize, and classify atrial cardiomyopathy into four subgroups based on their histopathological features. The predominant pathological feature of classes I and III is the hypertrophy of atrial cardiomyocytes. Here, we aim to investigate the mechanism of epigenetic transcriptional regulation of cardiomyocyte hypertrophy in atrial cardiomyopathy. Methods and Results Compared with that of sinus rhythm control individuals, the myocardium of patients with atrial fibrillation exhibited increased levels of angiotensin II (AngII), chromatin-bound myocyte enhancer factor 2 (MEF2), acetylated histone H4 (H4ac), and H3K27ac; upregulation of hypertrophy-related genes; and decreased levels of histone deacetylase (HDAC) 4 and HDAC5 bound to the promoters of hypertrophy-related genes. Furthermore, incubation of atrial cardiomyocytes with AngII increased their cross-sectional area and improved the expression of hypertrophy-related genes. AngII also promoted the phosphorylation of HDAC4 and HDAC5 and induced their nuclear export. RNA sequencing analyses revealed that AngII significantly upregulated genes associated with cardiac hypertrophy. Chromatin immunoprecipitation showed that this correlated with increased levels of chromatin-bound MEF2, H4ac, and H3K27ac and decreased HDAC4 and HDAC5 enrichment in the promoters of hypertrophy-related genes. Moreover, these AngII-induced prohypertrophic effects could be partially reverted by treatment with the AngII receptor blocker losartan. Conclusions AngII had a prohypertrophic effect on atrial cardiomyopathy which was epigenetic-dependent. Patients with atrial fibrillation manifest an increased susceptibility to hypertrophy and exhibit epigenetic characteristics that are permissive for the transcription of hypertrophy-related genes. AngII induces histone acetylation via the cytoplasmic-nuclear shuttling of HDACs, which constitutes a novel mechanism of atrial hypertrophy regulation and might provide a promising therapeutic strategy for atrial cardiomyopathy.

HBO1 induces histone acetylation and is important for non-small cell lung cancer cell growth

We examined the expression and the potential biological function of HBO1 in non-small cell lung cancer (NSCLC). TCGA and Oncomine databases showed that HBO1 transcripts were elevated in NSCLC. Furthermore, in local NSCLC tumor tissues HBO1 expression was higher than that in matched adjacent lung tissues. In primary and immortalized NSCLC cells, HBO1 shRNA robustly inhibited cell viability, proliferation and migration. Moreover, HBO1 knockout by CRISPR/Cas9 induced significant anti-tumor activity in NSCLC cells. Conversely, ectopic HBO1 overexpression in primary NSCLC cells increased proliferation and migration. H3-H4 histone acetylation and expression of several potential oncogenic genes (CCR2, MYLK, VEGFR2 and OCIAD2) were significantly decreased in NSCLC cells with HBO1 silencing or knockout. They were however increased after HBO1 overexpression. Intratumoral injection of HBO1 shRNA-expressing adeno-associated virus hindered the growth of A549 cell xenografts and primary NSCLC cell xenografts in nude mice. H3-H4 histone acetylation as well as expression of HBO1 and HBO1-dependent genes were decreased in HBO1-silenced NSCLC xenograft tissues. An HBO1 inhibitor WM-3835 potently inhibited NSCLC cell growth. Together, HBO1 overexpression promotes NSCLC cell growth.

Synthesis and biological evaluation of aminobenzamides containing purine moiety as class I histone deacetylases inhibitors

The aminobenzamide is selective to class I histone deacetylases (HDACs) and displays unique tight-binding/slow-off HDAC-binding mechanism. Herein, we report a series of 9-substituted purine aminobenzamides that selectively inhibit class I HDACs. The activities in vitro showed compound 9d exhibited 12 folds more potent than MS-275 against HDAC1 isoform and showed excellent inhibitory activity on cancer cells, including HCT-116, MDA-MB-231, K562 cell lines. The metabolic stability of 9d was much better than that of the well-known HDAC inhibitor SAHA. Pulse exposure test of western blot assay demonstrated that 9a, 9d induced histone acetylation in a similar manner to MS-275. Further biological validation demonstrated that 9d prevented cell transition from G1 phase to S phase by reducing Cyclin D1, CDK2 and lifting p21, induced early apoptosis by upregulating BAX and downregulating Bcl-2 in HCT-116 cells.

Remodeling “cold” tumor immune microenvironment via epigenetic-based therapy using targeted liposomes with in situ formed albumin corona

There is a close connection between epigenetic regulation, cancer metabolism, and immunology. The combination of epigenetic therapy and immunotherapy provides a promising avenue for cancer management. As an epigenetic regulator of histone acetylation, panobinostat can induce histone acetylation and inhibit tumor cell proliferation, as well as regulate aerobic glycolysis and reprogram intratumoral immune cells. JQ1 is a BRD4 inhibitor that can suppress PD-L1 expression. Herein, we proposed a chemo-free, epigenetic-based combination therapy of panobinostat/JQ1 for metastatic colorectal cancer. A novel targeted binary-drug liposome was developed based on lactoferrin-mediated binding with the LRP-1 receptor. It was found that the tumor-targeted delivery was further enhanced by in situ formation of albumin corona. The lactoferrin modification and endogenous albumin adsorption contribute a dual-targeting effect on the receptors of both LRP-1 and SPARC that were overexpressed in tumor cells and immune cells (e.g., tumor-associated macrophages). The targeted liposomal therapy was effective to suppress the crosstalk between tumor metabolism and immune evasion via glycolysis inhibition and immune normalization. Consequently, lactic acid production was reduced and angiogenesis inhibited; TAM switched to an anti-tumor phenotype, and the anti-tumor function of the effector CD8+ T cells was reinforced. The strategy provides a potential method for remodeling the tumor immune microenvironment (TIME).

Involvement of 5-HT1A receptor-mediated histone acetylation in the regulation of depression.

Depression is one of the most common and disabling mental disorders. There is growing evidence that 5-HT1A receptor is involved in the regulation of depressive-related behaviors. However, the exact mechanism underlying the role of 5-HT1A receptor in depression remains unknown. Histone acetylation is associated with the pathophysiology and treatment of depression. In the current study, we investigated whether the epigenetic histone deacetylase (HDAC)-induced histone acetylation mediates the regulation of 5-HT1A receptor in depressive behaviors. We showed that 5-HT1A receptor selective agonist (±)-8-hydroxy-2-(dipropylamino) tetralin hydrobromide led to significant increase in acetylation of H3 at lysine 9 (Ac-H3K9) and H4 at lysine 5 (Ac-H4K5) and lysine 12 (Ac-H4K12) with obviously decreasing histone deacetylase 1 (HDAC1), histone deacetylase 2 (HDAC2), histone deacetylase 4 (HDAC4) and histone deacetylase 5 (HDAC5) expression in hippocampus of mice. Conversely, 5-HT1A receptor selective antagonist NAN-190 decreased the level of acetylation of H3 and H4 with increasing the expression of HDAC1, HDAC2, HDAC4 and HDAC5 in the hippocampus. Furthermore, we found that HDAC inhibitors, trichostatin A or suberoylanilide hydroxamic acid infusion to hippocampus prevented the depressive behaviors induced by NAN-190, as well as histone H3 and H4 acetylation in mice. Our results suggested that epigenetic histone acetylation coupled with 5-HT1A receptor may play vital role in the pathophysiology and treatment of depressive disorders.

Simvastatin-romidepsin combination kills bladder cancer cells synergistically

The HMG-CoA reductase inhibitor simvastatin activates AMP-activated protein kinase (AMPK) and thereby induces histone acetylation. We postulated that combining simvastatin with the histone deacetylase (HDAC) inhibitor romidepsin would kill bladder cancer cells by inducing histone acetylation cooperatively. The combination of romidepsin and simvastatin induced robust apoptosis and killed bladder cancer cells synergistically. In murine subcutaneous tumor models using MBT-2 cells, a 15-day treatment with 0.5 mg/kg romidepsin and 15 mg/kg simvastatin was well tolerated and inhibited tumor growth significantly. Mechanistically, the combination induced histone acetylation by activating AMPK. The combination also decreased the expression of HDACs, thus further promoting histone acetylation. This AMPK activation was essential for the combination's action because compound C, an AMPK inhibitor, suppressed the combination-induced histone acetylation and the combination's ability to induce apoptosis. We also found that the combination increased the expression of peroxisome proliferator-activated receptor (PPAR) γ, leading to reactive oxygen species production. Furthermore, the combination induced endoplasmic reticulum (ER) stress and this ER stress was shown to be associated with increased AMPK expression and histone acetylation, thus playing an important role in the combination's action. Our study also suggests there is a positive feedback cycle between ER stress induction and PPARγ expression.

Induction of the apoptosis, degranulation and IL-13 production of human basophils by butyrate and propionate via suppression of histone deacetylation.

Allergic diseases are caused by dysregulated Th2 immune responses involving multiple effector cells including basophils. Short chain fatty acids (SCFAs), mainly acetate, propionate and butyrate, exert immunomodulatory functions via activation of its receptors GPR41 and GPR43, and inhibition of the histone deacetylases (HDACs) activity. In allergic diseases, SCFAs suppress the activity of mast cells, eosinophils and type 2 innate lymphoid cells (ILC2) but enhance the function of Th2 cells. Here, we aimed to elucidate the function of SCFAs on human basophils. Human basophils were purified from healthy donors by flow cytometric sorting. The surface proteins, apoptosis and degranulation of basophils were analyzed by flow cytometric analysis. The mRNA expression was assayed using real-time PCR. Interleukin 4 (IL-4) and IL-13 were measured by ELISA. Histone acetylation was examined by western blot. GPR41 was expressed by basophils and was enhanced by IL-3. Acetate induced intracellular calcium influx in basophils which was suppressed by blocking GPR41. Propionate and butyrate, but not acetate, induced the expression of CD69 and IL-13. In addition, propionate and butyrate enhanced IgE-mediated basophil degranulation but inhibited basophil survival and IL-4secretion. Propionate and butyrate induced histone acetylation of basophils and suppression of HDACs activity mimicked the effects of propionate and butyrate on human basophils. Our findings demonstrate that propionate and butyrate may play a complex role in regulating basophil apoptosis, activation and degranulation via inhibiting HDACs activity. The in vivo effects of SCFAs on the regulation of basophil-associated allergic diseases need to be further explored.

Sodium Valproate-Induced Chromatin Remodeling.

Valproic acid/sodium valproate (VPA), a drug originally prescribed as an anticonvulsant, has been widely reported to act on epigenetic marks by inducing histone acetylation, affecting the DNA and histone methylation status, and altering the expression of transcription factors, thus leading to modulation of gene expression. All these epigenetic changes have been associated with chromatin remodeling effects. The present minireview briefly reports the main effects of VPA on chromatin and image analysis and Fourier transform infrared (FTIR) microspectroscopy in association with molecular biology methodological approaches to investigate the VPA-induced changes in chromatin structure and at the higher-order supraorganizational level.

\u0394Np63 is a pioneer factor that binds inaccessible chromatin and elicits chromatin remodeling

BackgroundΔNp63 is a master transcriptional regulator playing critical roles in epidermal development and other cellular processes. Recent studies suggest that ΔNp63 functions as a pioneer factor that can target its binding sites within inaccessible chromatin and induce chromatin remodeling.MethodsIn order to examine if ΔNp63 can bind to inaccessible chromatin and to determine if specific histone modifications are required for binding, we induced ΔNp63 expression in two p63-naïve cell lines. ΔNp63 binding was then examined by ChIP-seq and the chromatin at ΔNp63 targets sites was examined before and after binding. Further analysis with competitive nucleosome binding assays was used to determine how ΔNp63 directly interacts with nucleosomes.ResultsOur results show that before ΔNp63 binding, targeted sites lack histone modifications, indicating ΔNp63’s capability to bind at unmodified chromatin. Moreover, the majority of the sites that are bound by ectopic ΔNp63 expression exist in an inaccessible state. Once bound, ΔNp63 induces acetylation of the histone and the repositioning of nucleosomes at its binding sites. Further analysis with competitive nucleosome binding assays reveal that ΔNp63 can bind directly to nucleosome edges with significant binding inhibition occurring within 50 bp of the nucleosome dyad.ConclusionOverall, our results demonstrate that ΔNp63 is a pioneer factor that binds nucleosome edges at inaccessible and unmodified chromatin sites and induces histone acetylation and nucleosome repositioning.

Microbial colonization induces histone acetylation critical for inherited gut-germline-neural signaling.

The gut-neural axis plays a critical role in the control of several physiological processes, including the communication of signals from the microbiome to the nervous system, which affects learning, memory, and behavior. However, the pathways involved in gut-neural signaling of gut-governed behaviors remain unclear. We found that the intestinal distension caused by the bacterium Pseudomonas aeruginosa induces histone H4 Lys8 acetylation (H4K8ac) in the germline of Caenorhabditis elegans, which is required for both a bacterial aversion behavior and its transmission to the next generation. We show that induction of H4K8ac in the germline is essential for bacterial aversion and that a 14-3-3 chaperone protein family member, PAR-5, is required for H4K8ac. Our findings highlight a role for H4K8ac in the germline not only in the intergenerational transmission of pathogen avoidance but also in the transmission of pathogenic cues that travel through the gut-neural axis to control the aversive behavior.