P300 Histone Acetyltransferase Inhibitor Research Articles

Curcumin, an Inhibitor of p300-HAT Activity, Suppresses the Development of Hypertension-Induced Left Ventricular Hypertrophy with Preserved Ejection Fraction in Dahl Rats.

We found that curcumin, a p300 histone acetyltransferase (HAT) inhibitor, prevents cardiac hypertrophy and systolic dysfunction at the stage of chronic heart failure in Dahl salt-sensitive rats (DS). It is unclear whether curcumin suppresses the development of hypertension-induced left ventricular hypertrophy (LVH) with a preserved ejection fraction. Therefore, in this study, we randomized DS (n = 16) and Dahl salt-resistant (DR) rats (n = 10) at 6 weeks of age to either curcumin or vehicle groups. These rats were fed a high-salt diet and orally administrated with 50 mg/kg/d curcumin or its vehicle for 6 weeks. Both curcumin and vehicle treatment groups exhibited similar degrees of high-salt diet-induced hypertension in DS rats. Curcumin significantly decreased hypertension-induced increase in posterior wall thickness and LV mass index, without affecting the systolic function. It also significantly reduced hypertension-induced increases in myocardial cell diameter, perivascular fibrosis and transcriptions of the hypertrophy-response gene. Moreover, it significantly attenuated the acetylation levels of GATA4 in the hearts of DS rats. A p300 HAT inhibitor, curcumin, suppresses the development of hypertension-induced LVH, without affecting blood pressure and systolic function. Therefore, curcumin may be used for the prevention of development of LVH in patients with hypertension.

Histone Deacetylase 1 Reduces Lipogenesis by Suppressing SREBP1 Transcription in Human Sebocyte Cell Line SZ95.

Proper regulation of sebum production is important for maintaining skin homeostasis in humans. However, little is known about the role of epigenetic regulation in sebocyte lipogenesis. We investigated histone acetylation changes and their role in key lipogenic gene regulation during sebocyte lipogenesis using the human sebaceous gland cell line SZ95. Sebocyte lipogenesis is associated with a significant increase in histone acetylation. Treatment with anacardic acid (AA), a p300 histone acetyltransferase inhibitor, significantly decreased the lipid droplet number and the expression of key lipogenic genes, including sterol regulatory-binding protein 1 (SREBP1), fatty acid synthase (FAS), and acetyl-CoA carboxylase (ACC). In contrast, treatment with trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, increased the expression of these genes. Global HDAC enzyme activity was decreased, and HDAC1 and HDAC2 expression was downregulated during sebaceous lipogenesis. Interestingly, HDAC1 knockdown increased lipogenesis through SREBP1 induction, whereas HDAC1 overexpression decreased lipogenesis and significantly suppressed SREBP1 promoter activity. HDAC1 and SREBP1 levels were inversely correlated in human skin sebaceous glands as demonstrated in immunofluorescence images. In conclusion, HDAC1 plays a critical role in reducing SREBP1 transcription, leading to decreased sebaceous lipogenesis. Therefore, HDAC1 activation could be an effective therapeutic strategy for skin diseases related to excessive sebum production.

3,4-dihydroxytoluene, a metabolite of rutin, suppresses the progression of nonalcoholic fatty liver disease in mice by inhibiting p300 histone acetyltransferase activity.

3,3',4',5,7-Pentahydroxyflavone-3-rhamnoglucoside (rutin) is a flavonoid with a wide range of pharmacological activities. Dietary rutin is hardly absorbed because the microflora in the large intestine metabolize rutin into a variety of compounds including quercetin and phenol derivatives such as 3,4-dihydroxyphenolacetic acid (DHPAA), 3,4-dihydroxytoluene (DHT), 3,4-hydroxyphenylacetic acid (HPAA) and homovanillic acid (HVA). We examined the potential of rutin and its metabolites as novel histone acetyltransferase (HAT) inhibitors. DHPAA, HPAA and DHT at the concentration of 25 μM significantly inhibited in vitro HAT activity with DHT having the strongest inhibitory activity. Furthermore, DHT was shown to be a highly efficient inhibitor of p300 HAT activity, which corresponded with its high degree of inhibition on intracellular lipid accumulation in HepG2 cells. Docking simulation revealed that DHT was bound to the p300 catalytic pocket, bromodomain. Drug affinity responsive target stability (DARTS) analysis further supported the possibility of direct binding between DHT and p300. In HepG2 cells, DHT concentration-dependently abrogated p300-histone binding and induced hypoacetylation of histone subunits H3K9, H3K36, H4K8 and H4K16, eventually leading to the downregulation of lipogenesis-related genes and attenuating lipid accumulation. In ob/ob mice, administration of DHT (10, 20 mg/kg, iv, every other day for 6 weeks) dose-dependently improved the NAFLD pathogenic features including body weight, liver mass, fat mass, lipid accumulation in the liver, and biochemical blood parameters, accompanied by the decreased mRNA expression of lipogenic genes in the liver. Our results demonstrate that DHT, a novel p300 histone acetyltransferase inhibitor, may be a potential preventive or therapeutic agent for NAFLD.

Field-based rational design of p300 histone acetyltransferase inhibitor and systematic evaluation as an anti-fibrotic agent

(E)-3-(3-(4-((3-Carbamoylbenzyl)oxy)-3-iodo-5-methoxyphenyl) acryloyl)benzamide (A6) was found to be a potent p300 inhibitor (IC50 = 870 nM) showing a similar binding mode to that of acetyl-CoA, a p300 substrate, and effective anti-fibrotic activity in both TGF-β1-stimulated lung fibroblast cells and bleomycin-induced in vivo lung fibrosis mice.

Expression of DHA-Metabolizing Enzyme Alox15 is Regulated by Selective Histone Acetylation in Neuroblastoma Cells

The omega-3 polyunsaturated fatty acid, docosahexaenoic acid (DHA) is enriched in neural membranes of the CNS, and recent studies have shown a role of DHA metabolism by 15-lipoxygenase-1 (Alox15) in prefrontal cortex resolvin D1 formation, hippocampo-prefrontal cortical long-term-potentiation, spatial working memory, and anti-nociception/anxiety. In this study, we elucidated epigenetic regulation of Alox15 via histone modifications in neuron-like cells. Treatment of undifferentiated SH-SY5Y human neuroblastoma cells with the histone deacetylase (HDAC) inhibitors trichostatin A (TSA) and sodium butyrate significantly increased Alox15 mRNA expression. Moreover, Alox15 expression was markedly upregulated by Class I HDAC inhibitors, MS-275 and depsipeptide. Co-treatment of undifferentiated SH-SY5Y cells with the p300 histone acetyltransferase (HAT) inhibitor C646 and TSA or sodium butyrate showed that p300 HAT inhibition modulated TSA or sodium butyrate-induced Alox15 upregulation. Differentiation of SH-SY5Y cells with retinoic acid resulted in increased neurite outgrowth and Alox15 mRNA expression, while co-treatment with the p300 HAT inhibitor C646 and retinoic acid modulated the increases, indicating a role of p300 HAT in differentiation-associated Alox15 upregulation. Increasing Alox15 expression was found in primary murine cortical neurons during development from 3 to 10 days-in-vitro, reaching high levels of expression by 10 days-in-vitro—when Alox15 was not further upregulated by HDAC inhibition. Together, results indicate regulation of Alox15 mRNA expression in neuroblastoma cells by histone modifications, and increasing Alox15 expression in differentiating neurons. It is possible that one of the environmental influences on the immature brain that can affect cognition and memory, may take the form of epigenetic effects on Alox15 and metabolites of DHA.

Inhibition of DNA Methylation in the COL1A2 Promoter by Anacardic Acid Prevents UV-Induced Decrease of Type I Procollagen Expression

UV radiation decreases type I procollagen production mainly by inhibiting the transforming growth factor-β/Smad signaling pathway. Because further epigenetic regulatory mechanisms are unclear, we investigated the roles of DNA methylation and histone acetylation in UV-induced regulation of COL1A2 transcription in human dermal fibroblasts. Anacardic acid, a p300 histone acetyltransferase inhibitor, rescued the UV-induced decrease of type I procollagen expression in human dermal fibroblasts. Although UV irradiation induced global histone acetylation, it reduced the local recruitment of histone H3 acetylation as well as p300, and Smad2/3 to the p300 binding site (-1406/-1393), in the COL1A2 promoter as shown by chromatin immunoprecipitation. This effect was reversed by anacardic acid treatment. In contrast, pyrosequencing analysis showed that UV irradiation induced DNA methylation in the same region of the COL1A2 promoter, which was reversed by anacardic acid and a DNA methyltransferase inhibitor (5-AZA-2'-deoxycytidine). Inhibition of UV-induced DNA methylation led to an increase of UV-induced histone acetylation in the COL1A2 promoter and increased the recruitment of transcription factors, leading to up-regulation of type I collagen after UV irradiation. Collectively, our findings indicate that the epigenetic crosstalk between DNA methylation and histone acetylation plays a crucial role in COL1A2 transcription induced by UV irradiation.

Application of Curcumin to Heart Failure Therapy by Targeting Transcriptional Pathway in Cardiomyocytes

Heart failure is one of the leading causes of death throughout the world. During the development and deterioration processes of heart failure, cardiomyocytes undergo maladaptive hypertrophy by altering hypertrophy-related gene expression. The zinc finger protein GATA4 is one of the transcription factors involved in the regulation of cardiomyocyte hypertrophy. In response to hypertrophic stimuli such as the synaptic nervous and rennin-angiotensin systems, GATA4 forms a large complex with various functional proteins including an intrinsic histone acetyltransferase, p300, and the disruption of this complex results in the inhibition of hypertrophic responses in cardiomyocytes. While such a transcriptional signal pathway is recognized as a critical event during cardiomyocyte hypertrophy, pharmacological heart failure therapy that targets this pathway has not been established. In order to develop novel heart failure therapy targeting the pathway in cardiomyocytes, we have studied the potential of curcumin, a p300 histone acetyltransferase inhibitor, as an agent for novel heart failure therapy. In this review, we describe a recent study on the cardiac transcriptional signal pathway, especially p300/GATA4 pathway, and a novel heart failure therapy using curcumin.

Quercetin Suppresses Cyclooxygenase-2 Expression and Angiogenesis through Inactivation of P300 Signaling

Quercetin, a polyphenolic bioflavonoid, possesses multiple pharmacological actions including anti-inflammatory and antitumor properties. However, the precise action mechanisms of quercetin remain unclear. Here, we reported the regulatory actions of quercetin on cyclooxygenase-2 (COX-2), an important mediator in inflammation and tumor promotion, and revealed the underlying mechanisms. Quercetin significantly suppressed COX-2 mRNA and protein expression and prostaglandin (PG) E(2) production, as well as COX-2 promoter activation in breast cancer cells. Quercetin also significantly inhibited COX-2-mediated angiogenesis in human endothelial cells in a dose-dependent manner. The in vitro streptavidin-agarose pulldown assay and in vivo chromatin immunoprecipitation assay showed that quercetin considerably inhibited the binding of the transactivators CREB2, C-Jun, C/EBPβ and NF-κB and blocked the recruitment of the coactivator p300 to COX-2 promoter. Moreover, quercetin effectively inhibited p300 histone acetyltransferase (HAT) activity, thereby attenuating the p300-mediated acetylation of NF-κB. Treatment of cells with p300 HAT inhibitor roscovitine was as effective as quercetin at inhibiting p300 HAT activity. Addition of quercetin to roscovitine-treated cells did not change the roscovitine-induced inhibition of p300 HAT activity. Conversely, gene delivery of constitutively active p300 significantly reversed the quercetin-mediated inhibition of endogenous HAT activity. These results indicate that quercetin suppresses COX-2 expression by inhibiting the p300 signaling and blocking the binding of multiple transactivators to COX-2 promoter. Our findings therefore reveal a novel mechanism of action of quercetin and suggest a potential use for quercetin in the treatment of COX-2-mediated diseases such as breast cancers.

Virtual Ligand Screening of the p300/CBP Histone Acetyltransferase: Identification of a Selective Small Molecule Inhibitor

The histone acetyltransferase (HAT) p300/CBP is a transcriptional coactivator implicated in many gene regulatory pathways and protein acetylation events. Although p300 inhibitors have been reported, a potent, selective, and readily available active-site-directed small molecule inhibitor is not yet known. Here we use a structure-based, in silico screening approach to identify a commercially available pyrazolone-containing small molecule p300 HAT inhibitor, C646. C646 is a competitive p300 inhibitor with a K(i) of 400 nM and is selective versus other acetyltransferases. Studies on site-directed p300 HAT mutants and synthetic modifications of C646 confirm the importance of predicted interactions in conferring potency. Inhibition of histone acetylation and cell growth by C646 in cells validate its utility as a pharmacologic probe and suggest that p300/CBP HAT is a worthy anticancer target.

Curcumin Exerts Synergistic Effects With ACE Inhibitor During Systolic Function Restoration After Myocardial Infarction in Rats

Purpose: We found that curcumin, a p300 histone acetyltransferase inhibitor, prevents deterioration of the systolic function in rat heart failure models in vivo. To clinically apply this novel therapy to humans, it should be clarified whether or not Cur exhibits additional effects on conventional HF therapy involving angiotensin-converting enzyme inhibitors (ACEI). Methods: Rats were subjected to a sham operation or MI. One week later, 32 rats were randomly assigned to solvents (control), enalapril (ACEI, 10mg/kg/day) alone, curcumin (50mg/kg/day) alone, or curcumin plus enalapril for 6 weeks.

Identification and characterization of a novel p300 HAT inhibitor

p300 is a histone acetyltransferase (HAT) with important roles in chromatin remodeling and transcriptional coactivation. Along with its homolog CBP, p300 is involved in many cellular functions, including cell cycle regulation and differentiation, and has been implicated in many diseases, including heart disease, diabetes, HIV, neurodegenerative disorders, and several forms of cancer. Despite its importance, few inhibitors of p300's HAT activity have been found to date. A recent X‐ray crystal structure of p300 HAT in complex with the bisubstrate analog Lys‐CoA served as a template for computational docking analysis. We utilized virtual ligand screening (VLS) to identify novel inhibitors of the p300 HAT domain. After an iterative screening process of the VLS hits using spectrophotometric and radioactive assays, a new p300 HAT inhibitor was identified. This compound was found to be a potent competitive inhibitor of p300 vs. acetyl‐CoA and noncompetitive vs. peptide substrate, consistent with prediction. Furthermore, this small molecule p300 HAT inhibitor demonstrated high selectivity versus other acetyltransferases tested. Preliminary analysis with this compound supports its ability to block p300/CBP HAT activity in cell culture. Ongoing studies are directed at characterizing the structural basis of inhibition and the range of cellular actions of this compound and related analogs. Funding: NIH, FAMRI.

Synthesis and Evaluation of a Potent and Selective Cell-Permeable p300 Histone Acetyltransferase Inhibitor

This paper describes the first potent and selective p300 histone acetyltransferase (HAT) inhibitor which is effective in live cells. This compound 7 is a coenzyme A analogue conjugated to a cell permeabilizing oligoArg peptide via disulfide linkage. This compound was shown to block cellular histone acetylation and transcription using a p300-sensitive reporter. It should thus be broadly useful for dissecting the role of p300 HAT activity in physiologic and disease states.