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PCI-34051 is a valuable tool to interrogate the therapeutic effects of selective inhibition of HDAC8. However, it has not advanced to clinical trials, perhaps due to poor PK or off-target effects. We hypothesized that the presence of a hydroxamic acid (HA) group in PCI-34051 contributed to its lack of advancement. Therefore, we replaced the HA in the PCI-34051 scaffold with a series of moieties that have the potential to bind to Zn and evaluated their activity in a HDAC8 assay. Surprisingly, none of the replacements effectively mimicked the HA, and analogs lost significant potency. Evaluation of the analogs' affinity to Zn indicated that none had affinity for Zn within the same range as the HA. These studies point to the difficulty in the application of bioisosteric replacements for Zn binding motifs.
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Inibidores de Histona Desacetilases , Histona Desacetilases , Ácidos Hidroxâmicos , Proteínas Repressoras , Histona Desacetilases/metabolismo , Ácidos Hidroxâmicos/química , Ácidos Hidroxâmicos/farmacologia , Ácidos Hidroxâmicos/síntese química , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/síntese química , Humanos , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/metabolismo , Relação Estrutura-Atividade , Zinco/química , Zinco/farmacologia , Estrutura Molecular , Sulfonamidas/química , Sulfonamidas/farmacologia , Sulfonamidas/síntese química , Relação Dose-Resposta a Droga , IndóisRESUMO
In the adult mammalian brain, neurons are produced from neural stem cells (NSCs) residing in two niches-the subventricular zone (SVZ), which forms the lining of the lateral ventricles, and the subgranular zone in the hippocampus. Epigenetic mechanisms contribute to maintaining distinct cell fates by suppressing gene expression that is required for deciding alternate cell fates. Several histone deacetylase (HDAC) inhibitors can affect adult neurogenesis in vivo. However, data regarding the role of specific HDACs in cell fate decisions remain limited. Herein, we demonstrate that HDAC8 participates in the regulation of the proliferation and differentiation of NSCs/neural progenitor cells (NPCs) in the adult mouse SVZ. Specific knockout of Hdac8 in NSCs/NPCs inhibited proliferation and neural differentiation. Treatment with the selective HDAC8 inhibitor PCI-34051 reduced the neurosphere size in cultures from the SVZ of adult mice. Further transcriptional datasets revealed that HDAC8 inhibition in adult SVZ cells disturbs biological processes, transcription factor networks, and key regulatory pathways. HDAC8 inhibition in adult SVZ neurospheres upregulated the cytokine-mediated signaling and downregulated the cell cycle pathway. In conclusion, HDAC8 participates in the regulation of in vivo proliferation and differentiation of NSCs/NPCs in the adult SVZ, which provides insights into the underlying molecular mechanisms.
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Células-Tronco Adultas , Células-Tronco Neurais , Intervenção Coronária Percutânea , Animais , Camundongos , Ventrículos Laterais , Inibidores de Histona Desacetilases , Proliferação de Células , MamíferosRESUMO
Various diseases are deeply associated with aberrations in HDAC8 functions. These aberrations can be assigned to either structural functions or catalytic functions of HDAC8. Therefore, development of HDAC8 degradation inducers might be more promising than HDAC8 inhibitors. We employed the proteolysis targeting chimera (PROTAC) strategy to develop a selective and potent HDAC8 degradation inducer CT-4 with single-digit nanomolar DC50 values and over 95% Dmax in both triple-negative breast cancer MDA-MB-231 cells and T-cell leukemia cells. Notably, CT-4 demonstrated potent anti-migration activity and limited anti-proliferative activity in MDA-MB-231 cells. In contrast, CT-4 effectively induced apototic cell death in Jurkat cells, as assessed by a caspase 3/7 activity assay and flow cytometry. Our findings suggest that the development of HDAC8 degradation inducers holds great potential for the treatment of HDAC8-related diseases.
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Quimera de Direcionamento de Proteólise , Proteínas Repressoras , Humanos , Linhagem Celular Tumoral , Histona Desacetilases/metabolismo , Células Jurkat , Proteólise , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/químicaRESUMO
Purpose: The aim of this study was to evaluate the dynamic change in staining of Class I HDACs and Hdac6 in lesions harvested serially from different time points in mice with induced endometriosis. In addition, the effect of Hdac8 activation as well as Hdac8 and Hdac6 inhibition on lesional progression and fibrogenesis was evaluated. Methods: Immunohistochemistry analysis of Class I HDACs and Hdac6 in serially harvested lesion samples in mouse. Hdac8 activation, as well as Hdac6/8 inhibition, was evaluated in mice with induced endometriosis. Results: We found a progressive increase in lesional staining of Hdac1, Hdac8, and Hdac6 and gradual decrease in Hdac2 staining and consistently reduced staining of Hdac3 during the course of lesional progression. The stromal Hdac8 staining correlated most prominently with all markers of lesional fibrosis. Hdac8 activation significantly accelerated the progression and fibrogenesis of endometriotic lesions. In contrast, specific inhibition of Hdac8 or Hdac6, especially of Hdac8, significantly hindered lesional progression and fibrogenesis. Conclusions: Hdac8 is progressively and aberrantly overexpressed as endometriotic lesions progress. This, along with the documented HDAC1 upregulation in endometriosis and the overwhelming evidence for the therapeutic potentials of HDACIs, calls for further and in-depth investigation of epigenetic aberrations of endometriosis in general and of HDACs in particular.
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Purpose: To screen Zn2+-dependent histone deacetylase (HDAC) 1-11 in endometriotic cells and then evaluated the HDACs identified from the screening in ovarian endometrioma (OE) and deep endometriotic (DE) lesions, and to evaluate the therapeutic potential of HDAC8 inhibition in mice. Methods: Quantification of gene and protein expression levels of HDAC1-11 in endometriotic cells stimulated by TGF-ß1, and immunohistochemistry analysis of Class I HDACs and HDAC6 in OE/DE lesion samples. The therapeutic potential of HDAC8 inhibition was evaluated by a mouse model of deep endometriosis. Results: The screening identified Class I HDACs and HDAC6 as targets of interest. Immunohistochemistry analysis found a significant elevation in HDAC8 immunostaining in both OE and DE lesions, which was corroborated by gene and protein expression quantification. For other Class I HDACs and HDAC6, their lesional expression was more subtle and nuanced. HDAC1 and HDAC6 staining was significantly elevated in DE lesions while HDAC2 and HDAC3 staining was reduced in DE lesions. Treatment of mice with induced deep endometriosis with an HDAC8 inhibitor resulted in significantly longer hotplate latency, a reduction of lesion weight by nearly two-thirds, and significantly reduced lesional fibrosis. Conclusions: These findings highlight the progression-dependent nature of specific HDAC aberrations in endometriosis, and demonstrate, for the first titme, the therapeutic potential of suppressing HDAC8.
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Histone deacetylase 8 (HDAC8) is overexpressed in multiple cancers and lack of effective chemical probes which could detect and visualize HDAC8 in tumor cells and tissues remains unsolved. In this work, three novel turn-on HDAC8 fluorescent probes 17-19 derived from solvatochromic fluorophore 4-sulfamonyl-7-aminobenzoxadiazole (SBD) conjugating with a potent HDAC8 inhibitor PCI-34051 (IC50 = 10 nM) as the recognition group were fabricated. The probes exhibited much stronger fluorescence when they transfer from hydrophilic environment (Φ < 8%) to hydrophobic environment (Φ > 46%). Compared with PCI-34051 (KD = 9.16 × 10-6 M), probes 17 (KD = 5.37 × 10-6 M), 18 (KD = 3.57 × 10-6 M) and 19 (KD = 8.89 × 10-6 M) possessed slightly better affinity for HDAC8. Probe 19 was selected for cell imaging and it showed significantly enhanced fluorescence only after binding into the cavity of HDAC8 in SH-SY5Y and MDA-MB-231 tumor cells. Co-localization results demonstrated that HDAC8 is expressed in cytoplasm and nucleus. Furthermore, probe 19 was successfully utilized to distinguish the expression level of HDAC8 in SH-SY5Y tumor and normal tissue slices.
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Neuroblastoma , Intervenção Coronária Percutânea , Corantes Fluorescentes , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Humanos , Proteínas RepressorasRESUMO
Many epigenetic modifications occur in glioma, in particular the histone-deacetylase class proteins play a pivotal role in glioma development, driving the proliferation rate and the invasiveness of tumor cells, and modulating the tumor microenvironment. In this study, we evaluated the role of the histone deacetylase HDAC8 in the regulation of the immune response in glioma and tumor growth. We found that inhibition of HDAC8 by the specific inhibitor PCI-34051 reduces tumor volume in glioma mouse models. We reported that HDAC8 modulates the viability and the migration of human and murine glioma cells. Interestingly, HDAC8 inhibition increases the acetylation of alpha-tubulin, suggesting this epigenetic modification controls glioma migration. Furthermore, we identify HDAC8 as a key molecule that supports a poorly immunogenic tumor microenvironment, modulating microglial phenotype and regulating the gene transcription of NKG2D ligands that trigger the Natural Killer cell-mediated cytotoxicity of tumor cells. Altogether, these results identify HDAC8 as a key actor in glioma growth and tumor microenvironment, and pave the way to a better knowledge of the molecular mechanisms of immune escape in glioma.
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Glioma , Histona Desacetilases , Intervenção Coronária Percutânea , Animais , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/genética , Histona Desacetilases/imunologia , Histona Desacetilases/metabolismo , Histonas/metabolismo , Imunidade , Camundongos , Microambiente TumoralRESUMO
Mantle Cell Lymphoma (MCL) is a non-Hodgkin lymphoma with a median survival rate of five years. Standard treatment with high-dose chemotherapy plus rituximab (anti-CD20 antibody) has extended overall survival although, the disease remains incurable. Histone deacetylases (HDAC) are a family of enzymes that regulate multiple proteins and cellular pathways through post-translational modification. Broad spectrum HDAC inhibitors have shown some therapeutic promise, inducing cell cycle inhibition and apoptosis in leukemia and non-Hodgkin's lymphoma. However, the therapeutic effects of these broad-spectrum HDAC inhibitors can detrimentally dampen Natural Killer (NK) cell cytotoxicity, reduce NK viability, and downregulate activation receptors important for NK mediated anti-tumor responses. Impairment of NK function in MCL patients during therapy potentially limits therapeutic activity of rituximab. Thus, there is an unmet need to decipher specific roles of individual HDACs in order to preserve and/or enhance NK function, while, directly impairing MCL viability. We investigated the impact of HDAC8 in MCL cell lines. Inhibition or genetic loss of HDAC8 caused MCL cells to undergo apoptosis. In contrast, exposure of primary human NK cells to an HDAC8 inhibitor does not alter viability, receptor expression, or antibody dependent cellular cytotoxicity (ADCC). However, an increase in effector cytokine interferon-gamma (IFNγ) producing NK cells was observed in response to HDAC8 inhibition. Taken together these data suggest that selective HDAC8 inhibitors may simultaneously preserve NK functional activity, while impairing MCL tumor growth, establishing a rationale for future clinical evaluation.
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Células Matadoras Naturais/efeitos dos fármacos , Linfoma de Célula do Manto/tratamento farmacológico , Proteínas Repressoras/antagonistas & inibidores , Antineoplásicos Imunológicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/genética , Humanos , Ácidos Hidroxâmicos/farmacologia , Indóis/farmacologia , Interferon gama/metabolismo , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Linfoma de Célula do Manto/imunologia , Linfoma de Célula do Manto/patologia , Proteínas Repressoras/genética , Rituximab/farmacologiaRESUMO
Histone deacetylases (HDACs) have been shown to alleviate renal fibrosis, however, the role of individual HDAC isoforms in this process is poorly understood. In this study, we examined the role of HDAC8 in the development of renal fibrosis and partial epithelial-mesenchymal transitions (EMT). In a murine model of renal fibrosis induced by unilateral ureteral obstruction (UUO), HDAC8 was primarily expressed in renal tubular epithelial cells and time-dependently upregulated. This occurred in parallel with the deacetylation of cortactin, a nonhistone substrate of HDAC8, and increased expression of three fibrotic markers: α-smooth muscle actin, collagen 1, and fibronectin. Administration of PCI34051, a highly selective inhibitor of HDAC8, restored acetylation of contactin and reduced expression of those proteins. PCI34051 treatment also reduced the number of renal tubular epithelial cells arrested at the G2/M phase of the cell cycle and suppressed phosphorylation of Smad3, STAT3, ß-catenin, and expression of Snail after ureteral obstruction. In contrast, HDAC8 inhibition reversed UUO-induced downregulation of BMP7 and Klotho, two renoprotective proteins. In cultured murine proximal tubular cells, treatment with PCI34051 or specific HDAC8 siRNA was also effective in inhibiting transforming growth factor ß1 (TGFß1)-induced deacetylation of contactin, EMT, phosphorylation of Smad3, STAT3, and ß-catenin, upregulation of Snail, and downregulation of BMP7 and Klotho. Collectively, these results suggest that HDAC8 activation is required for the EMT and renal fibrogenesis by activation of multiple profibrotic signaling and transcription factors, and suppression of antifibrotic proteins. Therefore, targeting HDAC8 may be novel therapeutic approach for treatment of renal fibrosis.
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Fibrose/metabolismo , Histona Desacetilases/metabolismo , Nefropatias/metabolismo , Rim/metabolismo , Acetilação/efeitos dos fármacos , Animais , Linhagem Celular , Regulação para Baixo/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Fibrose/tratamento farmacológico , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Indóis/farmacologia , Rim/efeitos dos fármacos , Nefropatias/tratamento farmacológico , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima/efeitos dos fármacos , Obstrução Ureteral/tratamento farmacológico , Obstrução Ureteral/metabolismoRESUMO
P19 pluripotent embryonic carcinoma (EC) stem cells are derived from pluripotent germ cell tumours and can differentiate into three germ layers. Treatment of these cells in suspension culture with retinoic acid induces their differentiation into neurons and glial cells. Hence, these cells are an excellent in vitro model to study the transition from the upper blastoderm to the neuroectoderm. However, because of the complex nature of the techniques involved, the results are highly dependent on the skills of the experimenter. Herein, we developed a simple method to induce neuronal differentiation of adherent P19 EC cells in TaKaRa NDiff® 227 serum-free medium (originally N2B27 medium). This medium markedly induced neuronal differentiation of P19 EC cells. The addition of retinoic acid to the NDiff® 227 medium further enhanced differentiation. Furthermore, cells differentiated by the conventional method, as well as the new method, showed identical expression of the mature neuronal marker, neuronal nuclei. To determine whether our approach could be applied for neuronal studies, we measured histone deacetylase 8 (HDAC8) activity using an HDAC8 inhibitor and HDAC8-knockout P19 EC cells. Inhibition of HDAC8 activity suppressed neuronal maturation. Additionally, HDAC8-knockout cell lines showed immature differentiation compared to the wild-type cell line. These results indicate that HDAC8 directly regulates the neuronal differentiation of P19 EC cells. Thus, our method involving P19 EC cells can be used as an experimental system to study the nervous system. Moreover, this method is suitable for screening drugs that affect the nervous system and cell differentiation.
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Histona Desacetilases/metabolismo , Neurogênese , Animais , Linhagem Celular Tumoral , Corpos Embrioides , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Camundongos Endogâmicos C3H , Neurogênese/efeitos dos fármacos , Neurônios/citologia , Neurônios/efeitos dos fármacos , Triazóis/farmacologiaRESUMO
Histone deacetylase 8 (HDAC8) is a class 1 histone deacetylase and a member of the cohesin complex. HDAC8 is expressed in smooth muscles, but its expression in skeletal muscle has not been described. We have shown for the first time that HDAC8 is expressed in human and zebrafish skeletal muscles. Using RD/12 and RD/18 rhabdomyosarcoma cells with low and high differentiation potency, respectively, we highlighted a specific correlation with HDAC8 expression and an advanced stage of muscle differentiation. We inhibited HDAC8 activity through a specific PCI-34051 inhibitor in murine C2C12 myoblasts and zebrafish embryos, and we observed skeletal muscles differentiation impairment. We also found a positive regulation of the canonical Wnt signaling by HDAC8 that might explain muscle differentiation defects. These findings suggest a novel mechanism through which HDAC8 expression, in a specific time window of skeletal muscle development, positively regulates canonical Wnt pathway that is necessary for muscle differentiation.
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Histona Desacetilases/metabolismo , Desenvolvimento Muscular/fisiologia , Músculo Esquelético/metabolismo , Proteínas Repressoras/metabolismo , Via de Sinalização Wnt/fisiologia , Animais , Diferenciação Celular/fisiologia , Humanos , Camundongos , Músculo Esquelético/citologia , Mioblastos/metabolismo , Peixe-ZebraRESUMO
Histone deacetylases (HDACs) catalyze the hydrolysis of acetylated lysine side chains in histone and non-histone proteins, and play a critical role in the regulation of many biological processes, including cell differentiation, proliferation, senescence, and apoptosis. Aberrant HDAC activity is associated with cancer, making these enzymes important targets for drug design. In general, HDAC inhibitors (HDACi) block the proliferation of tumor cells by inducing cell differentiation, cell cycle arrest, and/or apoptosis, and comprise some of the leading therapies in cancer treatments. To date, four HDACi have been FDA approved for the treatment of cancers: suberoylanilide hydroxamic acid (SAHA, Vorinostat, Zolinza®), romidepsin (FK228, Istodax®), belinostat (Beleodaq®), and panobinostat (Farydak®). Most current inhibitors are pan-HDACi, and non-selectively target a number of HDAC isoforms. Six previously reported HDACi were rationally designed, however, to target a unique sub-pocket found only in HDAC8. While these inhibitors were indeed potent against HDAC8, and even demonstrated specificity for HDAC8 over HDACs 1 and 6, there were no structural data to confirm the mode of binding. Here we report the X-ray crystal structure of Compound 6 complexed with HDAC8 to 1.98Å resolution. We also describe the use of molecular docking studies to explore the binding interactions of the other 5 related HDACi. Our studies confirm that the HDACi induce the formation of and bind in the HDAC8-specific subpocket, offering insights into isoform-specific inhibition.
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Inibidores de Histona Desacetilases/química , Histona Desacetilases/química , Ácidos Hidroxâmicos/química , Proteínas Repressoras/química , Domínio Catalítico , Cristalografia por Raios X , Humanos , Isoenzimas/química , Simulação de Acoplamento Molecular , Ligação Proteica , Proteínas Repressoras/antagonistas & inibidoresRESUMO
Histone deacetylases catalyze the hydrolysis of an acetyl group from post-translationally modified acetyl-lysine residues in a wide variety of essential cellular proteins, including histones. Because these lysine modifications can alter the activity and properties of affected proteins, aberrant acetylation/deacetylation may contribute to disease states. Many fundamental questions regarding the substrate specificity and regulation of these enzymes have yet to be answered. Here, we optimize an enzyme-coupled assay to measure low micromolar concentrations of acetate, coupling acetate production to the formation of NADH (nicotinamide adenine dinucleotide, reduced form) that is measured by changes in either absorbance or fluorescence. Using this assay, we measured the steady-state kinetics of peptides representing the H4 histone tail and demonstrate that a C-terminally conjugated methylcoumarin enhances the catalytic efficiency of deacetylation catalyzed by cobalt(II)-bound histone deacetylase 8 [Co(II)-HDAC8] compared with peptide substrates containing a C-terminal carboxylate, amide, and tryptophan by 50-, 2.8-, and 2.3-fold, respectively. This assay can be adapted for a high-throughput screening format to identify HDAC substrates and inhibitors.
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Acetatos/metabolismo , Ensaios Enzimáticos/métodos , Histona Desacetilases/metabolismo , Sequência de Aminoácidos , Biocatálise , Inibidores de Histona Desacetilases/farmacologia , NAD/metabolismo , Oligopeptídeos/química , Oligopeptídeos/metabolismo , Especificidade por SubstratoRESUMO
SCOPE: Histone deacetylases (HDACs) play a crucial role in the transcriptional regulation of various genes which can contribute to metabolic disorders. Although sulforaphane (SFN), a natural HDAC inhibitor, has been reported to alleviate obesity in humans and mice, the specific mechanisms and how HDACs contribute to SFN's anti-obesity effects remain unclear. METHODS AND RESULTS: Oral administration of SFN in mice fed high-fat diet increases peroxisome proliferator activating receptor γ coactivator (PGC1α)-induced mitochondrial biogenesis in skeletal muscle. Among HDACs, SFN specifically inhibits HDAC8 activity. SFN enhances mitochondrial DNA and adenosine triphosphate (ATP) production in C2C12 myotubes, similar to the action of PCI34051, a synthetic HDAC8-specific inhibitor. These effects are mediated by increased expression of PGC1α via upregulation of cAMP response element binding (CREB, Ser133 ) phosphorylation and p53 (Lys379 ) acetylation. These SFN-induced effects are not observed in cells with a genetic deletion of HDAC8, suggesting the existence of a regulatory loop between HDAC8 and PGC1α in SFN's action. CONCLUSION: SFN prevents obesity-related metabolic dysregulation by enhancing mitochondrial biogenesis and function via targeting the HDAC8-PGCα axis. These results suggest SFN as a beneficial anti-obesity agent providing new insight into the role of HDAC8 in the PGC1α-mediated mitochondrial biogenesis, which may be a novel and promising drug target for metabolic diseases.
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Dieta Hiperlipídica , Biogênese de Organelas , Humanos , Camundongos , Animais , Dieta Hiperlipídica/efeitos adversos , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Obesidade/tratamento farmacológico , Obesidade/etiologia , Obesidade/metabolismo , Músculo Esquelético/metabolismo , Histona Desacetilases/metabolismo , Proteínas Repressoras/metabolismoRESUMO
Macroautophagy/autophagy has been utilized by many viruses, including foot-and-mouth disease virus (FMDV), to facilitate replication, while the underlying mechanism of the interplay between autophagy and innate immune responses is still elusive. This study showed that HDAC8 (histone deacetylase 8) inhibits FMDV replication by regulating innate immune signal transduction and antiviral response. To counteract the HDAC8 effect, FMDV utilizes autophagy to promote HDAC8 degradation. Further data showed that FMDV structural protein VP3 promotes autophagy during virus infection and interacts with and degrades HDAC8 in an AKT-MTOR-ATG5-dependent autophagy pathway. Our data demonstrated that FMDV evolved a strategy to counteract host antiviral activity by autophagic degradation of a protein that regulates innate immune response during virus infection.Abbreviations: 3-MA: 3-methyladenine; ATG: autophagy related; Baf-A1: bafilomycin A1; CCL5: C-C motif chemokine ligand 5; Co-IP: co-immunoprecipitation; CQ: chloroquine phosphate; DAPI: 4",6-diamidino-2-phenylindole; FMDV: foot-and-mouth disease virus; HDAC8: histone deacetylase 8; ISG: IFN-stimulated gene; IRF3: interferon regulatory factor 3; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MOI: multiplicity of infection; MAVS: mitochondria antiviral signaling protein; OAS: 2"-5'-oligoadenylate synthetase; RB1: RB transcriptional corepressor 1; SAHA: suberoylanilide hydroxamic acid; TBK1: TANK binding kinase 1; TCID50: 50% tissue culture infectious doses; TNF/TNF-α: tumor necrosis factor; TSA: trichostatin A; UTR: untranslated region.
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Background: Peritoneal dialysis (PD) is an effective replacement therapy for end-stage renal disease patients. However, long-term exposure to peritoneal dialysate will lead to the development of peritoneal fibrosis. Epigenetics has been shown to play an important role in peritoneal fibrosis, but the role of histone deacetylases 8 (HDAC8) in peritoneal fibrosis have not been elucidated. In this research, we focused on the role and mechanisms of HDAC8 in peritoneal fibrosis and discussed the mechanisms involved. Methods: We examined the expression of HDAC8 in the peritoneum and dialysis effluent of continuous PD patients. Then we assessed the role and mechanism of HDAC8 in peritoneal fibrosis progression in mouse model of peritoneal fibrosis induced by high glucose peritoneal dialysis fluid by using PCI-34051. In vitro, TGF-ß1 or IL-4 were used to stimulate human peritoneal mesothelial cells (HPMCs) or RAW264.7 cells to establish two cell injury models to further explore the role and mechanism of HDAC8 in epithelial-mesenchymal transition (EMT) and macrophage polarization. Results: We found that HDAC8 expressed highly in the peritoneum from patients with PD-related peritonitis. We further revealed that the level of HDAC8 in the dialysate increased over time, and HDAC8 was positively correlated with TGF-ß1 and vascular endothelial growth factor (VEGF), and negatively correlated with cancer antigen 125. In mouse model of peritoneal fibrosis induced by high glucose dialysate, administration of PCI-34051 (a selective HDAC8 inhibitor) significantly prevented the progression of peritoneal fibrosis. Treatment with PCI-34051 blocked the phosphorylation of epidermal growth factor receptor (EGFR) and the activation of its downstream signaling pathways ERK1/2 and STAT3/HIF-1α. Inhibition of HDAC8 also reduced apoptosis. In vitro, HDAC8 silencing with PCI-34051 or siRNA inhibited TGF-ß1-induced EMT and apoptosis in HPMCs. In addition, continuous high glucose dialysate or IL-4 stimulation induced M2 macrophage polarization. Blockade of HDAC8 reduced M2 macrophage polarization by inhibiting the activation of STAT6 and PI3K/Akt signaling pathways. Conclusions: We demonstrated that HDAC8 promoted the EMT of HPMCs via EGFR/ERK1/2/STAT3/HIF-1α, induced M2 macrophage polarization via STAT6 and PI3K/Akt signaling pathways, and ultimately accelerated the process of peritoneal fibrosis.
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Intervenção Coronária Percutânea , Fibrose Peritoneal , Animais , Humanos , Camundongos , Soluções para Diálise/farmacologia , Transição Epitelial-Mesenquimal , Receptores ErbB , Glucose/farmacologia , Histona Desacetilases , Interleucina-4/farmacologia , Macrófagos/metabolismo , Fibrose Peritoneal/metabolismo , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Fator de Crescimento Transformador beta1/metabolismo , Fator A de Crescimento do Endotélio Vascular/farmacologiaRESUMO
HDAC8 catalyzes the deacetylation of both histones and nonhistone proteins. The abnormal expression of HDAC8 is associated with various pathological conditions causing cancer and other diseases like myopathies, Cornelia de Lange syndrome, renal fibrosis, and viral and parasitic infections. The substrates of HDAC8 are involved in diverse molecular mechanisms of cancer such as cell proliferation, invasion, metastasis and drug resistance. Based on the crystal structures and the key residues at the active site, HDAC8 inhibitors have been designed along the canonical pharmacophore. This article details the importance, recent advancements, and the structural and functional aspects of HDAC8 with special emphasis on the medicinal chemistry aspect of HDAC8 inhibitors that will help in developing novel epigenetic therapeutics.
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Química Farmacêutica , Neoplasias , Humanos , Histona Desacetilases/metabolismo , Descoberta de Drogas , Neoplasias/tratamento farmacológico , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/química , Proteínas Repressoras/metabolismoRESUMO
Histone deacetylase 8 (HDAC8) is a verified biomolecular target associated with diverse diseases including cancer. Though several HDAC inhibitors emerged effective against such diseases, no selective HDAC8 inhibitor is approved to date. Therefore, the development of potent HDAC8-selective inhibitors is inevitable to combat such diseases. Here, some benzothiazine-derived HDAC8 inhibitors were considered for a comparative QSAR analysis which may elucidate the prime structural components responsible for modulating their efficacy. Several outcomes from these diverse modelling techniques justified one another and thus validated each other. The ligand-based pharmacophore modelling study identified ring aromatic, positive ionizable, and hydrophobic features as essential structural attributes for HDAC8 inhibition. Besides, MLR, HQSAR and field-based 3D-QSAR studies signified the utility of the positive ionizable and hydrophobic features for potent HDAC8 inhibition. Again, the field-based 3D-QSAR study provided useful insight regarding the substitution in the fused phenyl ring. Moreover, the current observations also validated the previously reported molecular docking observations. Based on the outcomes, some new molecules were designed and predicted. Therefore, this comparative structural analysis of these HDAC8 inhibitors will surely assist in the development of potent HDAC8 inhibitors as promising anticancer therapeutics in the future.
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Desenho de Fármacos , Relação Quantitativa Estrutura-Atividade , Simulação de Acoplamento Molecular , Ligantes , Histona Desacetilases/química , Histona Desacetilases/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/químicaRESUMO
OBJECTIVE: Cancer stem-like cells (CSLCs) are closely associated with tumor recurrence, metastasis, and drug-resistance. PHD-finger domain protein 5A (PHF5A) is related to tumorigenesis and development of non-small cell lung cancer (NSCLC). The role and regulatory mechanism of PHF5A in CSLCs of NSCLC remain unclear. This study aimed to identify the biological characteristics of CSLCs and the role of PHF5A in maintaining stemness of NSCLC. METHODS: H1299-spheres and A549-spheres were obtained by oncosphere-forming assay and CSLCs by flow cytometry. Expression of CD133, aldehyde dehydrogenase isoform 1, E-cadherin, vimentin, PHF5A, and histone deacetylase 8 (HDAC8) was tested using immunofluorescence staining, qRT-PCR, and Western blotting. CCK-8 and Transwell assays were employed to determine proliferation, migration, and invasion ability of CSLCs and adherent monolayer cells in NSCLC. We regulated PHF5A expression and HDAC activity in CSLCs to explore the mechanism of PHF5A in stemness maintenance and analyzed expression of target proteins in NSCLC tissues. RESULTS: Compared with monolayer cells, CSLCs showed a decreased response to cisplatin-mediated inhibition of proliferation, increased migration and invasion, and high expression of PHF5A and HDAC8, accompanied by EMT marker alterations. Targeted knockdown of PHF5A in CSLCs of NSCLC resulted in diminished stemness phenotypes and HDAC8 expression, whereas inhibition of HDAC activity affected stemness maintenance. Moreover, the expression of target proteins showed consistent changes in NSCLC tissues. CONCLUSIONS: Compared with monolayer cells, cancer stem-like phenotype properties of NSCLC were altered, PHF5A was involved in stemness maintenance of CSLCs, and this process may be related to the activation of HDAC8.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Proteínas de Ligação a RNA , Transativadores , Carcinoma Pulmonar de Células não Pequenas/genética , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Humanos , Neoplasias Pulmonares/genética , Recidiva Local de Neoplasia , Fenótipo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transativadores/genética , Transativadores/metabolismoRESUMO
Histone deacetylase (HDAC) expression and enzymatic activity are dysregulated in cardiovascular diseases. Among Class I HDACs, HDAC2 has been reported to play a key role in cardiac hypertrophy; however, the exact function of HDAC8 remains unknown. Here we investigated the role of HDAC8 in cardiac hypertrophy and fibrosis using the isoproterenol-induced cardiac hypertrophy model system.Isoproterenol-infused mice were injected with the HDAC8 selective inhibitor PCI34051 (30 mg kg-1 body weight). Enlarged hearts were assessed by HW/BW ratio, cross-sectional area, and echocardiography. RT-PCR, western blotting, histological analysis, and cell size measurements were performed. To elucidate the role of HDAC8 in cardiac hypertrophy, HDAC8 knockdown and HDAC8 overexpression were also used. Isoproterenol induced HDAC8 mRNA and protein expression in mice and H9c2 cells, while PCI34051 treatment decreased cardiac hypertrophy in isoproterenol-treated mice and H9c2 cells. PCI34051 treatment also reduced the expression of cardiac hypertrophic markers (Nppa, Nppb, and Myh7), transcription factors (Sp1, Gata4, and Gata6), and fibrosis markers (collagen type I, fibronectin, and Ctgf) in isoproterenol-treated mice. HDAC8 overexpression stimulated cardiac hypertrophy in cells, whereas HDAC8 knockdown reversed those effects. HDAC8 selective inhibitor and HDAC8 knockdown reduced the isoproterenol-induced activation of p38 MAPK, whereas HDAC8 overexpression promoted p38 MAPK phosphorylation. Furthermore, p38 MAPK inhibitor SB203580 significantly decreased the levels of p38 MAPK phosphorylation, as well as ANP and BNP protein expression, induced by HDAC8 overexpression.Here we show that inhibition of HDAC8 activity or expression suppresses cardiac hypertrophy and fibrosis. These findings suggest that HDAC8 could be a promising target to treat cardiac hypertrophy and fibrosis by regulating p38 MAPK.