Histone Deacetylase 9 Expression Research Articles

HDAC9 and miR-512 Regulate CAGE-Promoted Anti-Cancer Drug Resistance and Cellular Proliferation

Histone deacetylase 9 (HDAC9) is known to be upregulated in various cancers. Cancer-associated antigens (CAGEs) are cancer/testis antigens that play an important role in anti-cancer drug resistance. This study aimed to investigate the relationship between CAGEs and HDAC9 in relation to anti-cancer drug resistance. AGSR cells with an anti-cancer drug-resistant phenotype showed higher levels of CAGEs and HDAC9 than normal AGS cells. CAGEs regulated the expression of HDAC9 in AGS and AGSR cells. CAGEs directly regulated the expression of HDAC9. Rapamycin, an inducer of autophagy, increased HDAC9 expression in AGS, whereas chloroquine decreased HDAC9 expression in AGSR cells. The downregulation of HDAC9 decreased the autophagic flux, invasion, migration, and tumor spheroid formation potential in AGSR cells. The TargetScan analysis predicted that miR-512 was a negative regulator of HDAC9. An miR-512 mimic decreased expression levels of CAGEs and HDAC9. The miR-512 mimic also decreased the autophagic flux, invasion, migration, and tumor spheroid forming potential of AGSR cells. The culture medium of AGSR increased the expression of HDAC9 and autophagic flux in AGS. A human recombinant CAGE protein increased HDAC9 expression in AGS cells. AGSR cells displayed higher tumorigenic potential than AGS cells. Altogether, our results show that CAGE–HDAC9–miR-512 can regulate anti-cancer drug resistance, cellular proliferation, and autophagic flux. Our results can contribute to the understanding of the molecular roles of HDAC9 in anti-cancer drug resistance.

Histone deacetylase 9-mediated phenotypic transformation of vascular smooth muscle cells is a potential target for treating aortic aneurysm/dissection

Aortic aneurysm/dissection (AAD) is a serious cardiovascular condition characterized by rapid onset and high mortality rates. Currently, no effective drug treatment options are known for AAD. AAD pathogenesis is associated with the phenotypic transformation and abnormal proliferation of vascular smooth muscle cells (VSMCs). However, endogenous factors that contribute to AAD progression remain unclear. We aimed to investigate the role of histone deacetylase 9 (HDAC9) in AAD pathogenesis. HDAC9 expression was considerably increased in human thoracic aortic dissection specimens. Using RNA-sequencing (RNA-seq) and chromatin immunoprecipitation, we demonstrated that HDAC9 transcriptionally inhibited the expression of superoxide dismutase 2 and insulin-like growth factor-binding protein-3, which are critically involved in various signaling pathways. Furthermore, HDAC9 triggered the transformation of VSMCs from a systolic to synthetic phenotype, increasing their proliferation and migration abilities and suppressing their apoptosis. Consistent with these results, in vivo experiments revealed that TMP195, a pharmacological inhibitor of HDAC9, suppressed the formation of the β-aminopropionitrile-induced AAD phenotype in mice. Our findings indicate that HDAC9 may be a novel endogenous risk factor that promotes the onset of AAD by mediating the phenotypic transformation of VSMCs. Therefore, HDAC9 may serve as a potential therapeutic target for drug-based AAD treatment. Furthermore, TMP195 holds potential as a therapeutic agent for AAD treatment.

HDAC9 Inhibition as a Novel Treatment for Stroke.

The identification of a variant in the HDAC9 gene as a risk factor for large-artery atherosclerotic stroke, and subsequently coronary artery disease, has opened novel treatment pathways for stroke and more widely atherosclerotic disease. This article describes the pathway from gene discovery to novel therapeutic approaches that are now entering man. HDAC9 expression is elevated in human atherosclerotic plaque, while in animal and cellular models, reducing HDAC9 (histone deacetylase 9) protein is associated with reduced disease. Several mechanisms have been proposed to account for the association between HDAC9 and atherosclerosis including alterations in the inflammatory response and cholesterol efflux and endothelial-mesenchymal transition. The association raises the possibility that inhibiting HDAC9 may provide a novel treatment approach for atherosclerotic cardiovascular disease. This is supported by intervention studies demonstrating HDAC9 inhibition reduces atherosclerosis in animal and cellular models. Indirect data support such an approach in man. The antiseizure drug sodium valproate, which has nonspecific HDAC inhibitory properties, both inhibits atherosclerosis in animal models and is epidemiologically associated with reduced stroke and myocardial infarction risk in man. It is now being trailed in phase 2 studies in large-artery stroke, while more specific HDAC9 inhibitors are being developed.

Impact of housing temperature on adipose tissue HDAC9 expression and adipogenic differentiation in high fat-fed mice.

Impaired adipogenic differentiation exacerbates metabolic disease in obesity. This study reported that high-fat diet (HFD)-fed mice housed at thermoneutrality exhibited impaired adipogenic differentiation, attributed to increased expression of histone deacetylase 9 (HDAC9). However, the impact of HFD on adipogenic differentiation is reportedly variable, possibly reflecting divergent environmental conditions such as housing temperature. C57BL/6J (wild-type [WT]) mice were housed at either thermoneutral (28-30°C) or ambient (20-22°C) temperature and fed HFD or chow diet (CD) for 12 weeks. For acute exposure experiments, WT or transient receptor potential cation channel subfamily M member 8 (TRPM8) knockout mice housed under thermoneutrality were acutely exposed to ambient temperature for 6 to 24 h. WT mice fed HFD and housed at thermoneutrality, compared with ambient temperature, gained more weight despite reduced food intake. They likewise exhibited increased inguinal adipose tissue HDAC9 expression and reduced adipogenic differentiation in vitro and in vivo compared with CD-fed mice. Conversely, HFD-fed mice housed at ambient temperature exhibited minimal change in adipose HDAC9 expression or adipogenic differentiation. Acute exposure of WT mice to ambient temperature reduced adipose HDAC9 expression independent of sympathetic β-adrenergic signaling via a TRPM8-dependent mechanism. Adipose HDAC9 expression is temperature sensitive, regulating adipogenic differentiation in HFD-fed mice housed under thermoneutrality.

Sex-Dependent Role of Adipose Tissue HDAC9 in Diet-Induced Obesity and Metabolic Dysfunction.

Obesity is a major risk factor for both metabolic and cardiovascular disease. We reported that, in obese male mice, histone deacetylase 9 (HDAC9) is upregulated in adipose tissues, and global deletion of HDAC9 protected against high fat diet (HFD)-induced obesity and metabolic disease. Here, we investigated the impact of adipocyte-specific HDAC9 gene deletion on diet-induced obesity in male and female mice. The HDAC9 gene expression was increased in adipose tissues of obese male and female mice and HDAC9 expression correlated positively with body mass index in humans. Interestingly, female, but not male, adipocyte-specific HDAC9 KO mice on HFD exhibited reduced body weight and visceral adipose tissue mass, adipocyte hypertrophy, and improved insulin sensitivity, glucose tolerance and adipogenic differentiation gene expression. Furthermore, adipocyte-specific HDAC9 gene deletion in female mice improved metabolic health as assessed by whole body energy expenditure, oxygen consumption, and adaptive thermogenesis. Mechanistically, compared to female mice, HFD-fed male mice exhibited preferential HDAC9 expression in the stromovascular fraction, which may have offset the impact of adipocyte-specific HDAC9 gene deletion in male mice. These results suggest that HDAC9 expressed in adipocytes is detrimental to obesity in female mice and provides novel evidence of sex-related differences in HDAC9 cellular expression and contribution to obesity-related metabolic disease.

Abstract P3029: Role Of Histone Deacetylase 9 In The Development Of Adipose Tissue Senescence And Mitochondrial Dysfunction

Background: The prevalence of obesity increases with age and is a major risk factor for the development of cardiovascular disease. Our previous studies showed that histone deacetylase 9 (HDAC9) gene deletion protected mice against obesity-associated metabolic disorder. Here, we hypothesized that HDAC9 expression during aging plays an important role in the development of adipose tissue senescence and mitochondrial dysfunction. Methods and Results: Visceral adipose tissue from 20-month-old wild type (WT) mice showed increased expression of HDAC9 (2-fold), p16 (5-fold) and p21 (2-fold) compared to 3-month-old mice. In addition, HDAC9 expression in human subcutaneous adipose tissue positively correlated with age (r 2 =0.088, p=0.034). Furthermore, 1-year-old HDAC9 knockout (KO) mice were protected against the development of age-associated weight gain compared to age-matched WT, in conjunction with reduced levels of senescence-associated beta-galactosidase (SABG) staining and lower expression of p16 (0.7-fold) and p21 (0.5-fold). Interestingly, preadipocytes, which reside in adipose tissue and replenish aged adipocytes, were protected against H2O2 and UV-light induced senescence as evaluated by SABG staining, p16 and p21 expression. Next, we measured adipose tissue mitochondrial function, which is mechanistically linked to senescence, using the Seahorse analyzer. HDAC9 KO significantly increased adipose tissue mitochondrial oxygen consumption rate (e.g., increased basal respiration, proton leak and ATP production). To gain further mechanistic insight, we performed RNA-sequencing of WT and KO adipose tissues and identified thiosulfate sulfurtransferase (TST), an enzyme that is reduced in human obesity and modulates mitochondrial function, as one of the potential mediators for adipose tissue senescence and metabolism. Interestingly, TST expression was reduced in 1-year-old compared to 3-month-old mice (0.2-fold) and enriched in KO compared to WT (4-fold) adipose tissue by qPCR analysis. Conclusion: HDAC9 plays a crucial role in the development of adipose tissue senescence, potentially through regulation of TST and mitochondrial function and may be a promising target to maintain healthy adipose tissue during aging.

MiR-383-5p Regulated by the Transcription Factor CTCF Affects Neuronal Impairment in Cerebral Ischemia by Mediating Deacetylase HDAC9 Activity.

Stroke, the leading cause of long-term disability worldwide, is caused by the blockage or hemorage of cerebral arteries. The resultant cerebral ischemia causes local neuronal death and brain injury. Histone deacetylase 9 (HDAC9) has been reported to be elevated in ischemic brain injury, but its mechanism in stroke is still enigmatic. The present study aimed to unveil the manner of regulation of HDAC9 expression and the effect of HDAC9 activation on neuronal function in cerebral ischemia. MicroRNAs (miRNAs) targeting HDAC9 were predicted utilizing bioinformatics analysis. We then constructed the oxygen glucose deprivation (OGD) cell model and the middle cerebral artery occlusion (MCAO) rat model, and elucidated the expression of CCCTC binding factor (CTCF)/miR-383-5p/HDAC9. Targeting between miR-383-5p and HDAC9 was verified by dual-luciferase reporter assay and RNAi. After conducting an overexpression/knockdown assay, we assessed neuronal impairment and brain injury. We found that CTCF inhibited miR-383-5p expression via its enrichment in the promoter region of miR-383-5p, whereas the miR-383-5p targeted and inhibited HDAC9 expression. In the OGD model and the MCAO model, we confirmed that elevation of HDAC9 regulated by the CTCF/miR-383-5p/HDAC9 pathway mediated apoptosis induced by endoplasmic reticulum stress, while reduction of HDAC9 alleviated apoptosis and the symptoms of cerebral infarction in MCAO rats. Thus, the CTCF/miR-383-5p/HDAC9 pathway may present a target for drug development against ischemic brain injury.

The Histone Deacetylase 9 Stroke-Risk Variant Promotes Apoptosis and Inflammation in a Human iPSC-Derived Smooth Muscle Cells Model.

A common variant in the Histone Deacetylase 9 (HDAC9) gene is the strongest genetic risk for large-vessel stroke, and HDAC9 offers a novel target for therapeutic modulation. However, the mechanisms linking the HDAC9 variant with increased stroke risk is still unclear due to the lack of relevant models to study the underlying molecular mechanisms. We generated vascular smooth muscle cells using human induced pluripotent stem cells with the HDAC9 stroke risk variant to assess HDAC9-mediated phenotypic changes in a relevant cells model and test the efficacy of HDAC inhibitors for potential therapeutic strategies. Our human induced pluripotent stem cells derived vascular smooth muscle cells show enhanced HDAC9 expression and allow us to assess HDAC9-mediated effects on promoting smooth muscle cell dysfunction, including proliferation, migration, apoptosis and response to inflammation. These phenotypes could be reverted by treatment with HDAC inhibitors, including sodium valproate and small molecules inhibitors. By demonstrating the relevance of the model and the efficacy of HDAC inhibitors, our model provides a robust phenotypic screening platform, which could be applied to other stroke-associated genetic variants.

An HDAC9-associated immune-related signature predicts bladder cancer prognosis.

BackgroundThe close relationship between histone deacetylase 9 (HDAC9) and immunity has attracted attention. We constructed an immune signature for HDAC9, a vital epigenetic modification, to predict the survival status and treatment benefits in bladder cancer (BC).MethodsAn exhaustive analysis of HDAC9 and immunology via the tumor and immune system interaction database (TISIDB) was performed, and an immune prognostic risk signature was developed based on genes enriched in the top five immune-related pathways under high HDAC9 status. Comprehensive analysis of survival curves and Cox regression were used to estimate the effectiveness of the risk signature. The relationship between immunological characteristics and the risk score was evaluated, and the mechanisms were also explored.ResultsIn the TISIDB, HDAC9 was closely related to various immunological characteristics. The risk signature was obtained based on genes related to prognosis enriched in the top five immune-related pathways under high HDAC9 status. The survival rate of the high-risk BC patients was poor. The risk score was closely related to multiple immunological characteristics, drug sensitivity, immunotherapy benefits and biofunctions.ConclusionAn immune-related prognostic signature established for HDAC9 expression status could independently predict the prognosis of BC patients. The use of this signature could help clinicians make personalized treatment decisions.

Hsa_circ_0000527 Downregulation Suppresses the Development of Retinoblastoma by Modulating the miR-27a-3p/HDAC9 Pathway

ABSTRACT Background Accumulating evidence indicates that the progression of retinoblastoma (RB) may involve circRNA dysfunction. We aimed to disclose the role of hsa_circ_0000527 and its potential functional mechanism in RB. Methods The expression of hsa_circ_0000527, miR-27a-3p and histone deacetylase 9 (HDAC9) mRNA was monitored using quantitative real-time polymerase chain reaction (qPCR). Functional assays, including cell proliferation and apoptosis, were investigated using cell counting kit-8 (CCK-8) assay, colony formation assay and flow cytometry assay. The expression of apoptosis-associated proteins and HDAC9 protein was detected by western blot. The targeting relationship between miR-27a-3p and hsa_circ_0000527 or HDAC9 was verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. Besides, Xenograft models were constructed to confirm the effect of hsa_circ_0000527 in vivo. Results Hsa_circ_0000527 and HDAC9 were upregulated, while miR-27a-3p was downregulated in RB tissues and cells. Hsa_circ_0000527 downregulation repressed RB cell proliferation and induced RB cell apoptosis. MiR-27a-3p was a target of hsa_circ_0000527, and hsa_circ_0000527 suppressed the expression of miR-27a-3p. MiR-27a-3p inhibition reversed the role of hsa_circ_0000527 downregulation. In addition, HDAC9 was a target of miR-27a-3p, and hsa_circ_0000527 indirectly regulated HDAC9 expression by targeting miR-27a-3p. MiR-27a-3p restoration inhibited RB cell proliferation and promoted apoptosis, which was reversed by HDAC9 overexpression. Hsa_circ_0000527 downregulation could inactivate the PI3K/AKT pathway. Moreover, hsa_circ_0000527 downregulation blocked tumor growth rate in vivo. Conclusion hsa_circ_0000527 downregulation blocked the progression of RB by regulating the miR-27a-3p/HDAC9 pathway, which might be associated with the inactivation of the PI3K/AKT pathway.

HDAC9 rs11984041 polymorphism is associated with diabetic retinopathy in Slovenian patients with type 2 diabetes mellitus.

Histone deacetylase 9 (HDAC9) is an important regulator of transcription that has also been investigated as a candidate gene in some pathologies. Our aim was to investigate the association between rs2107595 and rs11984041 HDAC9 gene polymorphisms and diabetic retinopathy (DR) in Slovenian patients with type 2 diabetes mellitus (T2DM). We also investigated HDAC9 expression in the fibrovascular membranes (FVMs) of patients with proliferative DR (PDR). Our study involved 1290 unrelated Slovenian patients with T2DM: 542 of them with DR as the study group, and 748 without DR as the control group. The investigated polymorphisms were genotyped using KASPar genotyping assay. The expression of HDAC9 was examined by immunohistochemistry in human FVM from 25 patients with PDR. The T allele and TT genotype frequencies of the rs11984041 polymorphism were significantly higher in the study group compared to the controls. The logistic regression analysis showed that the carriers of the TT genotype of this polymorphism have a 3.76-fold increase (95% CI 1.04-11.67) in the risk of developing DR. The T allele of rs11984041 was associated with increased HDAC9 expression in FVMs, obtained from T2DM patients with PDR. Patients with the T allele of rs11984041 compared to the homozygotes for the wild type C allele exhibited higher density of HDAC9-positive cells (35±10/mm2 vs. 12±6/mm2, respectively). We observed a notable association between the TT genotype of rs11984041 and DR, indicating its possible role as a genetic risk factor for the development of this diabetic complication.

Role of histone deacetylase 9 in the development of adipose tissue senescence

Background Senescence is a state associated with aging and obesity in which cells stop replicating. It has recently been shown that elimination of senescent cells by senolytic therapy can extend health and lifespan in mice. Our previous studies showed that global histone deacetylase 9 (HDAC9) gene deletion protected mice against obesity-associated metabolic disorder. Here, we hypothesized that HDAC9 expression during aging plays an important role in the development of adipose tissue senescence. Methods and Results Adipose tissue from 20-month old wild type mice showed increased expression of HDAC9 (2 fold) and senescence markers, p16 (5 fold) and p21 (2 fold) compared to 3-month old mice, as examined by qRT-PCR. Furthermore, HDAC9 expression in human subcutaneous adipose tissue positively correlated with age (r2=0.088, p=0.034). Intriguingly, adipose tissues from HDAC9 knockout (KO) mice exhibited reduced levels of senescence-associated beta-galactosidase (SABG) staining and lower expression of p16 and p21 (0.4 fold and 0.2 fold, respectively). Preadipocytes reside in adipose tissue and replenish aged or dying adipocytes, thus playing an important role in regulating adipose tissue health and senescence. Interestingly, reduced SABG staining was also observed in early passage primary preadipocyte culture derived from HDAC9 KO adipose tissue as compared to WT (17% vs 30% SABG+). Furthermore, we found that HDAC9 KO primary preadipocytes were resistant to senescence-inducing stimuli such as H2O2 treatment or UV light exposure as evaluated by senescent marker expression. Next, we measured adipose tissue mitochondrial function which is mechanistically linked to cellular senescence. Interestingly, HDAC9 gene deletion significantly increased adipose tissue mitochondrial oxygen consumption rate (e.g. increased basal and maximal respiration, greater proton leak and ATP production) in adipose tissue, as measured by the Seahorse Analyzer. Conclusion HDAC9 plays a crucial role in the development of adipose tissue senescence, possibly through regulating mitochondrial function, and thus, HDAC9 may be a promising target to combat the development of a senescent phenotype and to maintain healthy adipose tissue.

Knockdown of long non‑coding RNA HCP5 suppresses the malignant behavior of retinoblastoma by sponging miR‑3619‑5p to target HDAC9.

A number of studies have verified the vital effects of long non‑coding RNAs on the malignant behaviour of retinoblastoma (RB). The objective of the present study was to examine the specific role and mechanisms of HLA complex P5 (HCP5) in RB. For this purpose, reverse transcription‑quantitative polymerase chain reaction was used to determine the expression of HCP5, microRNA (miRNA/miR)‑3619‑5p and histone deacetylase 9 (HDAC9). A 3‑(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2‑H-tetrazolium bromide assay was conducted to detect cell viability. Transwell assays were used to evaluate the abilities of cell migration and invasion. A mouse tumor model was established to explore the functions of HCP5 in RB invivo. The interactions between HCP5, miR‑3619‑5p and HDAC9 were confirmed by a dual‑luciferase reporter assay. The protein expression of HDAC9 was determined by western blot analysis. The results revealed that the expression levels of HCP5 and HDAC9 were upregulated, whereas those of miR‑3619‑5p were downregulated in RB tissues and cell lines. The downregulation of HCP5 or the overexpression of miR‑3619‑5p suppressed RB cell proliferation, migration and invasion invitro. Simultaneously, the knockdown of HCP5 suppressed tumor growth in mice invivo. In addition, HCP5 was directly bound to miR‑3619‑5p and inversely correlated with miR‑3619‑5p. HDAC9 was found to be a target gene of and negatively regulated by miR‑3619‑5p. HCP5 expression also positively correlated with HDAC9 expression. Rescue experiments revealed that the overexpression of HDAC9 or the inhibition of miR‑3619‑5p reversed the inhibition of RB cell viability, migration and invasion induced by the knockdown of HCP5. On the whole, the present study demonstrates that the silencing of HCP5 exerts an anti‑tumor effect in RB by sponging miR‑3619‑5p to target HDAC9.

MiR-211-5p is down-regulated and a prognostic marker in bladder cancer.

The micro RNA (miRNA)/histone deacetylase 9 (HDAC9) signaling axis has been reported to be involved in initiating and developing multiple malignant tumors. In the present study, we aimed to determine whether miR-211-5p serves as a post-transcriptional regulator in bladder cancer (BCa) cell proliferation and apoptosis by targeting HDAC9. miRNA expression profiling of BCa tissues and para-carcinoma tissues was screened by miRNA microarray. After transfection with miR-211-5p mimics or short hairpin RNA of HDAC9 (sh-HDAC9), mRNA and protein expression was evaluated using a quantitative reverse transcription-polymerase chain reaction and western blotting, respectively. A bioinformatics algorithm was used, and a dual-luciferase reporter assay was performed to validate HDAC9 as a direct target of miR-211-5p. Cell proliferation was analyzed by the 3-(4, 5-dimethylthiazl2-yl)-2,5-diphenyltetazolium bromide (MTT) assay. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) detection was used to evaluate apoptosis in 5637 and T24 cells. A transwell assay was used to assess migration and invasion. miR-211-5p is down-regulated in BCa tumor tissues and cell lines. miR-211-5p is identified as an independent biomarker for predicting overall survival. HDAC9 is a direct target of miR-211-5p, and overexpression of miR-211-5p represses HDAC9 protein expression in vitro. Overexpression of miR-211-5p or HDAC9 knockdown significantly inhibits proliferation, migration and invasion of 5637 and T24 cells, and also induces cell apoptosis. miR-211-5p may play a role as a tumor suppressor and as a favourable prognostic marker in BCa.

Long noncoding RNA CBR3 antisense RNA 1 promotes the aggressive phenotypes of non‑small‑cell lung cancer by sponging microRNA‑509‑3p and competitively upregulating HDAC9 expression.

Long noncoding RNA CBR3 antisense RNA 1 (CBR3-AS1) plays significant roles in the initiation and progression of osteosarcoma. The aim of the present study was to investigate the involvement of CBR3-AS1 in the development of non-small cell lung cancer (NSCLC). Reverse transcription-quantitative PCR was performed to detect CBR3-AS1 expression in NSCLC tissues and cell lines. The impacts of CBR3-AS1 on cellular proliferation, apoptosis, migration and invasiveness in vitro, and tumor growth in vivo, were investigated using the Cell Counting Kit-8 assay, flow cytometry, Transwell migration and invasion assays, and tumor xenograft model-based analysis, respectively. The results indicated that CBR3-AS1 was markedly upregulated in NSCLC tissues and cell lines. High CBR3-AS1 expression was correlated with larger tumor size, advanced TNM stage, increased incidence of lymph node metastasis and shorter overall survival times in patients with NSCLC. Furthermore, CBR3-AS1-knockdown notably suppressed cellular proliferation, migration and invasiveness in vitro, and also promoted apoptosis and suppressed tumorigenicity in vivo. Mechanistic investigation demonstrated that CBR3-AS1 functions as a competing endogenous RNA for microRNA-509-3p (miR-509-3p) in NSCLC cells. Furthermore, miR-509-3p exerted tumor-suppressive effects in NSCLC, and histone deacetylase 9 (HDAC9) was identified as a direct target of miR-509-3p. HDAC9 expression was suppressed by CBR3-AS1 depletion, which was abolished by miR-509-3p inhibition. Further rescue experiments revealed that increasing the output of the miR-509-3p/HDAC9 axis counteracted the CBR3-AS1 depletion-induced inhibitory effects on NSCLC cells. Collectively, the results of the present study indicate that the CBR3-AS1/miR-509-3p/HDAC9 pathway exerts tumor-promoting actions in NSCLC oncogenesis and progression, suggesting that this pathway is an effective target for the management of NSCLC.

HDAC9 deficiency promotes tumor progression by decreasing the CD8+ dendritic cell infiltration of the tumor microenvironment

BackgroundThe tumor microenvironment (TME) contains a variety of immune cells, which play critical roles during the multistep development of tumors. Histone deacetylase 9 (HDAC9) has been reported to have either...

Decreased expression and hypomethylation of HDAC9 lead to poor prognosis and inhibit immune cell infiltration in clear cell renal cell carcinoma

Introduction & ObjectivesTo evaluate the expression and methylation levels of histone deacetylase 9 (HDAC9) in clear cell renal cell carcinoma (ccRCC), investigate the correlations between the prognosis of ccRCC patients and HDAC9, and examine the associations between immunological parameters and HDAC9. Materials & MethodsThe differences between HDAC9 expression in ccRCC tissues and normal tissues were evaluated with reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR), western blotting and immunohistochemistry. HDAC9 methylation levels in ccRCC tissues and normal tissues were evaluated via The Cancer Genome Atlas and University of California, Santa Cruz Xena. Kaplan-Meier curves were also obtained from University of California, Santa Cruz Xena. Then, cell proliferation was examined via colony formation assays and 5-ethynyl-2′-deoxyuridine assays, and cell metastasis was investigated via transwell assays. Gene set enrichment analysis was conducted to examine the biofunctions of HDAC9. The relationships between HDAC9 expression and immunological parameters were assessed via the Tumor Immune Estimation Resource database. ResultsHDAC9 down-regulation and hypomethylation were observed in ccRCC tissues and led to poor prognosis. HDAC9 up-regulation inhibited the proliferation and metastasis of ccRCC cells. A subset of the pathways identified by gene set enrichment analysis were associated with the immune response and inflammation, which were significantly positively correlated with HDAC9 expression in ccRCC. We confirmed that HDAC9 significantly promoted immune cell infiltration but was positively correlated with the expression of immune checkpoint molecules. ConclusionDecreased expression and hypomethylation of HDAC9 lead to poor prognosis and inhibit immune cell infiltration in ccRCC.

MicroRNA‑936 inhibits the malignant phenotype of retinoblastoma by directly targeting HDAC9 and deactivating the PI3K/AKT pathway.

MicroRNA-936 (miR-936) has been reported to play important roles in the progression of non-small cell lung cancer and glioma. However, the expression and functions of miR-936 in retinoblastoma (RB) remain elusive and need to be further elucidated. Herein, the aims were to measure miR-936 expression in RB, identify the functional importance of miR-936 in the oncogenicity of RB, and investigate the underlying molecular mechanisms. Reverse-transcription quantitative PCR was carried out to determine miR-936 expression in RB tissues and cell lines. Cell proliferation, colony formation, apoptosis, migration, and invasion in vitro and tumor growth in vivo were examined respectively by Cell Counting Kit-8, colony formation, flow cytometric, and Transwell migration and invasion assays and a subcutaneous heterotopic xenograft experiment. The potential target of miR-936 was predicted by bioinformatic analysis and was subsequently validated by luciferase reporter assay, reverse-transcription quantitative PCR, and western blotting. miR-936 expression was weak in both RB tissues and cell lines and was correlated with differentiation, lymph node metastasis and TNM staging in RB. RB cell proliferation, colony formation, migration, and invasion in vitro and tumor growth in vivo were attenuated by exogenous miR-936, whereas apoptosis was enhanced by miR-936 overexpression. Further molecular investigation identified histone deacetylase 9 (HDAC9) as a direct target gene of miR-936 in RB cells. HDAC9 depletion had effects similar to those of miR-936 overexpression in RB cells. Recovery of HDAC9 expression counteracted the tumor-suppressive action of miR-936 on the oncogenicity of RB cells. Ectopic miR-936 expression deactivated the PI3K/AKT pathway in RB cells in vitro and in vivo by decreasing HDAC9 expression. Downregulated miR-936 is related to poor prognosis in RB, and its upregulation inhibits RB aggressiveness via direct targeting of HDAC9 mRNA and thereby inactivation of the PI3K/AKT pathway.

High HDAC9 is associated with poor prognosis and promotes malignant progression in pancreatic ductal adenocarcinoma.

Histone deacetylase 9 (HDAC9) is involved in a variety of malignant tumors, and leads to malignant tumor development and poor prognosis. However, the association between HDAC9 expression, and the prognosis and clinicopathological features of patients with pancreatic ductal adenocarcinoma (PDAC) remains unclear. The present study used reverse transcription-quantitative PCR, western blotting and immunohistochemistry to detect the expression level of HDAC9 in PDAC tumors and cell lines. The Kaplan-Meier method and Pearson's χ2 test were applied to evaluate the prognostic impact of HDAC9. The present study investigated the effect of HDAC9 on the biological function of PDAC cells. The present results indicated that HDAC9 was highly expressed in PDAC tissue and PDAC cell lines (P<0.05). HDAC9 expression level in tumor tissues was negatively associated with tumor size (P=0.026), T stage (P=0.014) and N stage (P=0.004). Kaplan-Meier analysis suggested that patients with high HDAC9 had shorter recurrence-free survival (RFS; P=0.017) and disease-specific survival (DSS; P=0.022). Moreover, the present results suggested that T stage, N stage and HDAC9 expression level were independent predictive factors for RFS and DSS in patients with PDAC. In addition, silencing HDAC9 significantly inhibited the proliferation and migration of PDAC cells. The present results indicated that high expression levels of HDAC9 were associated with tumor progression and poor prognosis; thus, HDAC9 may serve as a prognostic predictor of PDAC.

The Atherosclerosis Risk Variant rs2107595 Mediates Allele-Specific Transcriptional Regulation of HDAC9 via E2F3 and Rb1.

Background and Purpose- Genome-wide association studies have identified the HDAC9 (histone deacetylase 9) gene region as a major risk locus for atherosclerotic stroke and coronary artery disease in humans. Previous results suggest a role of altered HDAC9 expression levels as the underlying disease mechanism. rs2107595, the lead single nucleotide polymorphism for stroke and coronary artery disease resides in noncoding DNA and colocalizes with histone modification marks suggestive of enhancer elements. Methods- To determine the mechanisms by which genetic variation at rs2107595 regulates HDAC9 expression and thus vascular risk we employed targeted resequencing, proteome-wide search for allele-specific nuclear binding partners, chromatin immunoprecipitation, genome-editing, reporter assays, circularized chromosome conformation capture, and gain- and loss-of-function experiments in cultured human cell lines and primary immune cells. Results- Targeted resequencing of the HDAC9 locus in patients with atherosclerotic stroke and controls supported candidacy of rs2107595 as the causative single nucleotide polymorphism. A proteomic search for nuclear binding partners revealed preferential binding of the E2F3/TFDP1/Rb1 complex (E2F transcription factor 3/transcription factor Dp-1/Retinoblastoma 1) to the rs2107595 common allele, consistent with the disruption of an E2F3 consensus site by the risk allele. Gain- and loss-of-function studies showed a regulatory effect of E2F/Rb proteins on HDAC9 expression. Compared with the common allele, the rs2107595 risk allele exhibited higher transcriptional capacity in luciferase assays and was associated with higher HDAC9 mRNA levels in primary macrophages and genome-edited Jurkat cells. Circularized chromosome conformation capture revealed a genomic interaction of the rs2107595 region with the HDAC9 promoter, which was stronger for the common allele as was the in vivo interaction with E2F3 and Rb1 determined by chromatin immunoprecipitation. Gain-of-function experiments in isogenic Jurkat cells demonstrated a key role of E2F3 in mediating rs2107595-dependent transcriptional regulation of HDAC9. Conclusions- Collectively, our findings imply allele-specific transcriptional regulation of HDAC9 via E2F3 and Rb1 as a major mechanism mediating vascular risk at rs2107595.