trichostatin-a and apicidin

Eosinophils are one of the cells that play a critical role in the pathogenesis of allergic diseases. The increase in the number of eosinophils in such diseases is regulated by interleukin-5 (IL-5). The author have prepared recombinant rat IL-5 using a baculovirus expression system and examined its biological activities in rat eosinophils. It was demonstrated that recombinant rat IL-5 prolongs the survival of mature eosinophils and differentiates immature eosinophils into mature eosinophils, suggesting that rat IL-5 is a factor for eosinophilia in rats. Recombinant rat eosinophil-associated ribonuclease (Ear)-1 and Ear-2 were also prepared. Eosinophil granule proteins are thought to cause tissue damage due to their cytotoxic activity, but using recombinant rat Ear-1 and Ear-2, it was found that rat Ear-1 and Ear-2 have strong RNase A activity and bactericidal activity, suggesting that these proteins play critical roles in host defense. Finally, the important role of acetylation of histones was clarified in the differentiation of HL-60 clone 15 cells into eosinophils using the histone deacetylase inhibitors sodium n-butyrate, apicidin, and trichostatin A. These findings would be useful for further investigations of the role of eosinophils in allergic inflammation.

Topics: Acetylation; Animals; Butyrates; Cell Differentiation; Cell Division; Enzyme Inhibitors; Eosinophil Cationic Protein; Eosinophil Granule Proteins; Eosinophils; Histone Deacetylase Inhibitors; Histones; HL-60 Cells; Humans; Hydroxamic Acids; Hypersensitivity; Inflammation; Interleukin-5; Peptides, Cyclic; Rats; Recombinant Proteins

Eosinophilic cell lines, HL-60 clone 15 cells and EoL-1 cells, have contributed to clarifying the mechanisms responsible for differentiation into eosinophils. These cells differentiate into eosinophils by continuous histone acetylation. Histone deacetylase inhibitors, sodium butyrate and apicidin, promote the transactivation of various genes in these cells by causing the hyperacetylation of histones, resulting in the differentiation of cells into eosinophils. In contrast, transient acetylation by histone deacetylase inhibitors such as trichostatin A does not induce eosinophilic differentiation. This chapter describes the maintenance of HL-60 clone 15 cells and EoL-1 cells and induction of the differentiation of these cell lines into eosinophils by the histone deacetylase inhibitor sodium butyrate.

Topics: Acetylation; Butyrates; Cell Differentiation; Cell Line; Eosinophils; Histone Deacetylase Inhibitors; Histones; HL-60 Cells; Humans; Hydroxamic Acids; Peptides, Cyclic

Dynamic histone acetylation, catalyzed by lysine acetyltransferases and HDACs, is critical to IEG expression. Expression of IEGs, such as FOSL1, is induced by several signal transduction pathways resulting in activation of the protein kinase MSK and phosphorylation of histone H3 at serine 10 of nucleosomes (the nucleosome response) at the upstream promoter and regulatory region of target genes. HDAC inhibitors prevent FOSL1 gene induction and the association of HDAC1, 2 and 3 with the gene body. However, HDAC inhibitors did not prevent the nucleosome response. Thus HDAC inhibitors perturb events downstream of the nucleosome response required for FOSL1 transcription initiation.

Topics: Gene Expression Regulation, Neoplastic; Genes, Immediate-Early; HCT116 Cells; Histone Deacetylase 1; Histone Deacetylase 2; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Hydroxamic Acids; Nucleosomes; Peptides, Cyclic; Proto-Oncogene Proteins c-fos; Tetradecanoylphorbol Acetate; Transcriptional Activation

Nuclear receptors use lysine acetyltransferases and lysine deacetylases (KDACs) in regulating transcription through histone acetylation. Lysine acetyltransferases interact with steroid receptors upon binding of an agonist and are recruited to target genes. KDACs have been shown to interact with steroid receptors upon binding to an antagonist. We have shown previously that KDAC inhibitors (KDACis) potently repress the mouse mammary tumor virus promoter through transcriptional mechanisms and impair the ability of the glucocorticoid receptor (GR) to activate it, suggesting that KDACs can play a positive role in GR transactivation. In the current study, we extended this analysis to the entire GR transcriptome and found that the KDACi valproic acid impairs the ability of agonist-bound GR to activate about 50% of its target genes. This inhibition is largely due to impaired transcription rather than defective GR processing and was also observed using a structurally distinct KDACi. Depletion of KDAC1 expression mimicked the effects of KDACi in over half of the genes found to be impaired in GR transactivation. Simultaneous depletion of KDACs 1 and 2 caused full or partial impairment of several more GR target genes. Altogether we found that Class I KDAC activity facilitates GR-mediated activation at a sizable fraction of GR-activated target genes and that KDAC1 alone or in coordination with KDAC2 is required for efficient GR transactivation at many of these target genes. Finally, our work demonstrates that KDACi exposure has a significant impact on GR signaling and thus has ramifications for the clinical use of these drugs.

Topics: Acetylation; Amidohydrolases; Animals; Cell Line, Tumor; Dexamethasone; Enzyme Inhibitors; Glucocorticoids; Histones; HSP90 Heat-Shock Proteins; Hydroxamic Acids; Lysine; Mice; Peptides, Cyclic; Protein Binding; Receptors, Glucocorticoid; Transcription, Genetic; Transcriptional Activation; Valproic Acid

The 5-lipoxygenase (5-LO) is the key enzyme in the formation of leukotrienes. We have previously shown that the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) activates 5-LO transcription via recruitment of Sp1, Sp3 and RNA polymerase II to the proximal promoter. To identify the HDACs involved in the regulation of 5-LO promoter activity isoform-specific HDAC inhibitors were applied. 5-LO promoter activity and mRNA expression were up-regulated by the class I HDAC inhibitors apicidin and MS-275 but not by class II inhibitors. Knockdown of HDAC 1, 2 and 3 revealed that HDAC2 and HDAC3 but not HDAC1 is involved in the up-regulation of 5-LO mRNA expression. To analyse the chromatin modifications at the 5-LO promoter associated with HDAC inhibition, the time course of 5-LO mRNA induction by trichostatin A was investigated and the concomitant changes in histone modifications at the 5-LO promoter in HL-60, U937 and Mono Mac6 cells were determined. Chromatin immunoprecipitation analysis revealed that trichostatin A increases acetylation of histones H3 and H4 at the 5-LO core promoter in HL-60 and U937 cells whereas no significant changes were observed in Mono Mac6 cells. The appearance of H3 and H4 acetylation preceded the 5-LO mRNA induction whereas in all three cell lines, induction of 5-LO mRNA expression correlated with histone H3 lysine 4 trimethylation (H3K4me3), a marker for transcriptional activity of gene promoters.

Topics: Acetylation; Arachidonate 5-Lipoxygenase; Benzamides; Chromatin Immunoprecipitation; Histone Deacetylase 1; Histone Deacetylase 2; Histone Deacetylase Inhibitors; Histone Deacetylases; Histones; HL-60 Cells; Humans; Hydroxamic Acids; Peptides, Cyclic; Promoter Regions, Genetic; Pyridines; RNA, Messenger; Transcription Factors; Transcriptional Activation; U937 Cells; Up-Regulation

High-altitude long-term hypoxia (LTH) is known to induce pulmonary arterial smooth muscle cell (PASMC) proliferation in the fetus, leading to pulmonary arterial remodeling and pulmonary hypertension of the newborn. The mechanisms underlying these conditions remain enigmatic however. We hypothesized that epigenetic alterations in fetal PASMC induced by high-altitude LTH may play an important role in modulating their proliferation during pulmonary arterial remodeling. To test this hypothesis, we have analyzed epigenetic alterations in the pulmonary vasculature of fetal lambs exposed to high-altitude LTH [pregnant ewes were kept at 3,801 m altitude from ~40 to 145 days gestation] or to sea level atmosphere. Intrapulmonary arteries were isolated, and fetal PASMC were cultured from both control and LTH fetuses. Compared with controls, in LTH fetus pulmonary arteries measurements of histone acetylation and global DNA methylation demonstrated reduced levels of global histone 4 acetylation and DNA methylation, accompanied by the loss of the cyclin-dependent kinase inhibitor p21. Treatment of LTH fetal PASMCs with histone deacetylase (HDAC) inhibitor trichostatin A decreased their proliferation rate, in part because of altered expression of p21 at both RNA and protein level. In PASMC of LTH fetuses, HDAC inhibition also decreased PDGF-induced cell migration and ERK1/2 activation and modulated global DNA methylation. On the basis of these observations, we propose that epigenetic alterations (reduced histone acetylation and DNA methylation) caused by chronic hypoxia leads to fetal PASMC proliferation and vessel remodeling associated with vascular proliferative disease and that this process is regulated by p21.

Topics: Acclimatization; Acetylation; Altitude; Animals; Cell Cycle Checkpoints; Cell Movement; Cell Proliferation; Cell Survival; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p21; DNA Methylation; Epigenesis, Genetic; Female; Fetal Hypoxia; Fetus; Gene Expression; Histone Deacetylase Inhibitors; Histones; Hydroxamic Acids; MAP Kinase Signaling System; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Peptides, Cyclic; Platelet-Derived Growth Factor; Pregnancy; Protein Processing, Post-Translational; Pulmonary Artery; Sheep; Time Factors; Tumor Suppressor Protein p53

The existence of stable, transcriptionally silent human immunodeficiency virus (HIV) in latently infected cells represents a major obstacle to acquired immune deficiency syndrome (AIDS) therapy. Histone deacetylase (HDAC) can inhibit histone acetylation, resulting in HIV-1 provirus transcription silence. Apicidin, a widely used antiparasitic drug, exhibits antiparasitic activity by inhibiting HDAC. Using the latently infected A10.6 cell line, we describe the dose- and time-dependent manner in which Apicidin reverses HIV-1 latency. We found that Apicidin can synergize with trichostatin A (TSA) to activate HIV-1 gene expression. Chromatin immunoprecipitation (ChIP) assay further indicates that Apicidin induces HIV-1 reactivation by increasing the acetylation levels of H3 and H4 at nucleosome 1 in HIV-1 long terminal repeats (LTR). Our research reveals a potent activator for reactivating latent HIV-1 and shows promise for HIV-1 therapy.

Topics: Cells, Cultured; Chromatin Immunoprecipitation; Drug Synergism; Gene Expression Profiling; Histone Deacetylase Inhibitors; Histones; HIV Long Terminal Repeat; HIV-1; Humans; Hydroxamic Acids; Jurkat Cells; Peptides, Cyclic; Virus Activation; Virus Latency

Overexpression of histone deacetylases has been reported in various human malignancies; however, the expression of histone deacetylases in endometrial tissue is not fully understood. In the present study, the expression of histone deacetylase 1, histone deacetylase 2, and Ki-67 was examined immunohistochemically in 30 normal and 66 malignant endometrial tissue samples. The results were expressed as a positivity index and compared with the positivity index for Ki-67 and rates of patient survival. The effect of 2 histone deacetylase inhibitors, trichostatin A and apicidine, on cell proliferation and the expression of cell cycle regulators such as cyclins (D1, E, and A), p21, p27, and p16 were investigated using 6 endometrial carcinoma cell lines. The positivity index for histone deacetylase 1 (79.8 +/- 33.0, mean +/- SD) and histone deacetylase 2 (106.3 +/- 41.9) was higher in endometrial carcinoma than the normal endometrium, with a significant difference for histone deacetylase 2. The positivity index for histone deacetylase 2 was significantly increased in higher-grade carcinomas (positivity index for grade 3, 124.9 +/- 28.4) compared with grade 1 tumors (86.0 +/- 41.0) and was positively correlated with that for Ki-67. In addition, patients with histone deacetylase 2-positive carcinomas had a poor prognosis compared with those with histone deacetylase 2-negative carcinoma (P = .048). Treatment with trichostatin A or apicidine suppressed the proliferation in all cell lines examined, in association with increased expression of p21 and down-regulation of cyclin D1 and cyclin A expression. These results indicated that increased histone deacetylase 2 expression is involved in the acquisition of aggressive behavior by endometrial carcinoma and suggest histone deacetylase inhibitor to be a promising anticancer drug for this carcinoma.

Topics: Adult; Aged; Apoptosis; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Endometrial Neoplasms; Endometrium; Female; Histone Deacetylase 1; Histone Deacetylase 2; Humans; Hydroxamic Acids; Immunohistochemistry; Ki-67 Antigen; Middle Aged; Peptides, Cyclic; Prognosis; Survival Analysis

Mitosis is a highly regulated process in which errors can lead to genomic instability, a hallmark of cancer. During this phase of the cell cycle, transcription is silent and RNA translation is inhibited. Thus, mitosis is largely driven by post-translational modification of proteins, including phosphorylation, methylation, ubiquitination, and sumoylation. Here, we show that protein acetylation is prevalent during mitosis. To identify proteins that are acetylated, we synchronized HeLa cells in early prometaphase and immunoprecipitated lysine-acetylated proteins with antiacetyl-lysine antibody. The immunoprecipitated proteins were identified by LC-ESI-MS/MS analysis. These include proteins involved in RNA translation, RNA processing, cell cycle regulation, transcription, chaperone function, DNA damage repair, metabolism, immune response, and cell structure. Immunoprecipitation followed by Western blot analyses confirmed that two RNA processing proteins, eIF4G and RNA helicase A, and several cell cycle proteins, including APC1, anillin, and NudC, were acetylated in mitosis. We further showed that acetylation of APC1 and NudC was enhanced by apicidin treatment, suggesting that their acetylation was regulated by histone deacetylase. Moreover, treating mitotic cells with apicidin or trichostatin A induced spindle abnormalities and cytokinesis failure. These studies suggest that protein acetylation/deacetylation is likely an important regulatory mechanism in mitosis.

Topics: Acetylation; Cell Cycle Proteins; Cytokinesis; DNA; HeLa Cells; Humans; Hydroxamic Acids; Immunoblotting; Mass Spectrometry; Microscopy, Fluorescence; Mitosis; Peptides, Cyclic; Proteomics; RNA Processing, Post-Transcriptional; Subcellular Fractions; Tubulin

In this study, we investigated the effects of various histone deacetylase (HDAC) inhibitors on adipocyte differentiation. Treatment of 3T3-L1 cells with HDAC inhibitors such as apicidin, trichostatin A, or suberoylanilide hydroxamic acid, under conditions that normally promote differentiation led to a dramatic attenuation of adipocyte differentiation. In contrast, sodium butyrate (NaB) treatment increased adipocyte differentiation. Accordingly, the expression of adipogenic marker genes such as FAS, aP2, PPARgamma, resistin, C/EBPalpha, ADD1/SREBP1c, and adiponectin were inhibited by apicidin treatment but not NaB, indicating that the adipocyte differentiation process could be differentially regulated depending on the type of HDAC inhibitor utilized. In addition, this differential effect seemed not to be due to disruption of the insulin- signaling pathway. Interestingly, our data showed that apicidin treatment could induce dedifferentiation of fully differentiated adipocytes, as evident by the fact that apicidin treatment led to a decrease of Oil Red O-stained adipocytes with a concomitant reduction in the expression levels of adipogenic marker genes. Collectively, our results suggest that adipocyte differentiation and dedifferentiation may be regulated by HDAC inhibitors.

Topics: 3T3-L1 Cells; Adipocytes; Adipogenesis; Animals; Butyrates; Cell Dedifferentiation; Enzyme Inhibitors; Gene Expression Regulation; Histone Deacetylase Inhibitors; Histone Deacetylases; Hydroxamic Acids; Insulin; Mice; Peptides, Cyclic; Triglycerides; Vorinostat

In inflammation, nitric oxide (NO) is produced by inducible nitric oxide synthase (iNOS) induced by bacterial products and cytokines, and NO acts as a regulatory and pro-inflammatory mediator. Glucocorticoids are powerful anti-inflammatory agents that inhibit the expression of iNOS and various other inflammatory factors. Histone deacetylation has been recently described as a novel mechanism how glucocorticoids down-regulate transcriptional activation of some inflammatory genes. The aim of the present study was to investigate the effects of inhibitors of histone deacetylation on the suppressive effects of glucocorticoids on NO production and iNOS expression. Dexamethasone and a dissociated glucocorticoid RU24858 inhibited NO production, and iNOS protein and mRNA expression in macrophages exposed to bacterial lipopolysaccharide (LPS). In the presence of a glucocorticoid receptor (GR) antagonist mifepristone, dexamethasone and RU24858 had no effect on NO production. The role of histone deacetylation in the glucocorticoid effect was studied by using three structurally different inhibitors of histone deacetylases (HDACs): trichostatin A, apicidin and MC1293. HDAC inhibitors reversed the effects of dexamethasone and RU24858 on iNOS expression and NO production. Stably transfected A549/8 cells containing luciferase gene under the control of human iNOS promoter were used in promoter-activity studies. iNOS promoter activity induced by IL-1beta was inhibited by dexamethasone and the inhibitory effect was reversed by HDAC inhibitor trichostatin A. The results suggest that glucocorticoids inhibit iNOS expression and NO production by a GR-mediated and GRE-independent manner through histone deacetylation and transcriptional silencing.

Topics: Animals; Anti-Inflammatory Agents; Blotting, Western; Cell Line; Desoximetasone; Dexamethasone; Glucocorticoids; Histone Deacetylase Inhibitors; Hydroxamic Acids; Hydroxycorticosteroids; Lipopolysaccharides; Macrophages; Mice; Nitric Oxide; Nitric Oxide Synthase Type II; Peptides, Cyclic; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

EoL-1 cells differentiate into eosinophils in the presence of n-butyrate, but the mechanism has remained to be elucidated. Because n-butyrate can inhibit histone deacetylases, we hypothesized that the inhibition of histone deacetylases induces the differentiation of EoL-1 cells into eosinophils. In this study, using n-butyrate and two other histone deacetylase inhibitors, apicidin and trichostatin A, we have analyzed the relationship between the inhibition of histone deacetylases and the differentiation into eosinophils in EoL-1 cells. It was demonstrated that apicidin and n-butyrate induced a continuous acetylation of histones H4 and H3, inhibited the proliferation of EoL-1 cells without attenuating the level of FIP1L1-PDGFRA mRNA, and induced the expression of markers for mature eosinophils such as integrin beta7, CCR1, and CCR3 on EoL-1 cells, while trichostatin A evoked a transient acetylation of histones and induced no differentiation into eosinophils. These findings suggest that the continuous inhibition of histone deacetylases in EoL-1 cells induces the differentiation into mature eosinophils.

Topics: Acetylation; Butyrates; Cell Differentiation; Cell Proliferation; Dose-Response Relationship, Drug; Enzyme Inhibitors; Eosinophils; Gene Expression Regulation, Enzymologic; Histone Deacetylase Inhibitors; Histone Deacetylases; Histones; HL-60 Cells; Humans; Hydroxamic Acids; Hypereosinophilic Syndrome; mRNA Cleavage and Polyadenylation Factors; Peptides, Cyclic; Platelet-Derived Growth Factor; RNA, Messenger

Histone deacetylase inhibitors (HDACIs) are currently in clinical trials partly due to their potent anti-angiogenic effects. However, the detailed mechanism of their action is unclear. Here, we observed that several HDACIs (TSA, SB, Apicidin, and VPA) dramatically decreased HIF-1alpha protein level and transcriptional activity of HIF-1 in human and mouse tumor cell lines. Furthermore, class I HDACs, HDAC1 and 3 enhanced HIF-1alpha stability and HIF-1 transactivation function in hypoxic conditions. In addition, immunoprecipitation and in vitro binding assays revealed that HDAC1 and 3 directly bind to the oxygen-dependent degradation domain of HIF-1alpha. Collectively, these results suggest that HDAC1 and 3 are considered as a positive regulator of HIF-1alpha stability via direct interaction and may play an important role in HIF-1-induced tumor angiogenesis.

Topics: Animals; Blotting, Western; Cell Line, Tumor; Enzyme Inhibitors; HeLa Cells; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Hydroxamic Acids; Hypoxia-Inducible Factor 1, alpha Subunit; Immunoprecipitation; Mice; Peptides, Cyclic; Reverse Transcriptase Polymerase Chain Reaction; Transcriptional Activation; Valproic Acid

EoL-1 cells have a FIP1L1-PDGFRA fusion gene which causes the transformation of eosinophilic precursor cells into leukemia cells. Recently, we suggested that the induction of differentiation of EoL-1 cells into eosinophils by the HDAC inhibitors apicidin and n-butyrate is due to the continuous inhibition of HDACs. However, neither apicidin nor n-butyrate inhibited the expression of FIP1L1-PDGFRA mRNA, although both these inhibitors suppressed cell proliferation. Therefore, in this study, we analyzed whether the levels of FIP1L1-PDGFRalpha protein and phosphorylated-Stat5 involved in the signaling for the proliferation of EoL-1 cells are attenuated by HDAC inhibitors.. EoL-1 cells were incubated in the presence of apicidin, TSA or n-butyrate. FIP1L1-PDGFRalpha and phosphorylated-Stat5 were detected by Western blotting.. Treatment of EoL-1 cells with apicidin at 100 nM or n-butyrate at 500 microM decreased the levels of FIP1L1-PDGFRalpha protein and phosphorylated-Stat5, while that with trichostatin A at 30 nM did not.. The decrease in the level of FIP1L1-PDGFRalpha protein caused by apicidin and n-butyrate might be one of the mechanisms by which EoL-1 cells are induced to differentiate into eosinophils by these HDAC inhibitors.

Topics: Butyrates; Cell Differentiation; Cell Line, Tumor; Eosinophils; Gene Expression Regulation, Leukemic; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Hypereosinophilic Syndrome; mRNA Cleavage and Polyadenylation Factors; Neoplasm Proteins; Oncogene Proteins, Fusion; Peptides, Cyclic; Phosphorylation; Protein Processing, Post-Translational; Receptor, Platelet-Derived Growth Factor alpha; RNA, Messenger; STAT5 Transcription Factor

Histone deacetylase inhibitors (HDI) increase gene expression through induction of histone acetylation. However, it remains unclear whether increases in specific gene expression events determine the apoptotic response following HDI administration. Herein, we show that a variety of HDI trigger in hematopoietic cells not only widespread histone acetylation and DNA damage responses but also actual DNA damage, which is significantly increased in leukemic cells compared with normal cells. Thus, increase in H2AX and ataxia telangiectasia mutated (ATM) phosphorylation, early markers of DNA damage, occurs rapidly following HDI administration. Activation of the DNA damage and repair response following HDI treatment is further emphasized by localizing DNA repair proteins to regions of DNA damage. These events are followed by subsequent apoptosis of neoplastic cells but not normal cells. Our data indicate that induction of apoptosis by HDI may result predominantly through accumulation of excessive DNA damage in leukemia cells, leading to activation of apoptosis.

Topics: Animals; Apoptosis; Ataxia Telangiectasia Mutated Proteins; Butyrates; Cell Cycle Proteins; Chromatin Assembly and Disassembly; DNA Damage; DNA-Binding Proteins; Gamma Rays; Histone Deacetylase Inhibitors; Histones; HL-60 Cells; Humans; Hydroxamic Acids; K562 Cells; Mice; Mice, Transgenic; Organ Specificity; Peptides, Cyclic; Phosphorylation; Protein Serine-Threonine Kinases; RNA Interference; Staurosporine; Transfection; Tumor Suppressor Proteins

1 We have examined the effect of the histone deacetylase inhibitors apicidin, trichostatin A (TSA) and n-butyrate on the histone acetylation and the differentiation of human eosinophilic leukemia HL-60 clone 15 cells into eosinophils. 2 Viability of the cells incubated with apicidin (100 nm), TSA (30 nm) or n-butyrate (500 microm) did not change significantly, but higher concentrations of apicidin (> or =300 nm) or TSA (> or =100 nm) decreased the viability when examined at day 1. 3 Apicidin (100 nm) as well as n-butyrate (500 microm) induced continuous acetylations of histone H4 and lysine14 residue on histone H3, while TSA (30 nm) induced transient acetylations. 4 After 6 days incubation, eosinophilic cells stained by Luxol-fast-blue were generated by apicidin (100 nm) and n-butyrate (500 microm) but not by TSA (30 nm). Other markers for differentiation into eosinophils such as changes in intracellular structure, and expressions of integrin beta7 and major basic protein, and the inhibition of cell proliferation were also induced by apicidin and n-butyrate but not by TSA. 5 Continuous acetylation of histone H4 achieved by repeated treatment with TSA (30 nm) at an interval of 12 h for more than three times induced such changes when examined on day 6. In addition, the induction was impaired by shortening the period of incubation with apicidin (100 nm) or n-butyrate (500 microm). 6 CCAAT/enhancer binding protein was continuously activated by apicidin (100 nm) and n-butyrate (500 microm), but was transiently activated by TSA (30 nm). 7 These findings suggest that the continuous acetylation of histones H3 and H4 is necessary for the differentiation of HL-60 clone 15 cells into eosinophils.

Topics: Acetylation; Butyrates; CCAAT-Enhancer-Binding Proteins; Cell Differentiation; Cell Proliferation; Enzyme Inhibitors; Eosinophils; Histone Deacetylase Inhibitors; Histone Deacetylases; Histones; HL-60 Cells; Humans; Hydroxamic Acids; Immunoblotting; Peptides, Cyclic; Time Factors

Pharmacologic stimulation of fetal hemoglobin (HbF) expression may be a promising approach for the treatment of beta-thalassemia. In this study, we have investigated the HbF-inducing activity and molecular mechanisms of specific histone deacetylase (HDAC) inhibitors in human K562 erythroleukemia cells. Apicidin was the most potent agent compared with other HDAC inhibitors (trichostatin A, MS-275, HC-toxin, suberoylanilide hydroxamic acid [SAHA]) and previously tested compounds (butyrate, phenylbutyrate, isobutyramide, hydroxyurea, 5-aza-cytidine), leading to a 10-fold stimulation of HbF expression at nanomolar to micromolar concentrations. Hyperacetylation of histones correlated with the ability of HDAC inhibitors to stimulate HbF synthesis. Furthermore, analysis of different mitogen-activated protein (MAP) kinase signaling pathways revealed that p38 signaling was activated following apicidin treatment of cells and that inhibition of this pathway abolished the HbF-inducing effect of apicidin. Additionally, activation of the Agamma-globin promoter by apicidin could be inhibited by p38 inhibitor SB203580. In summary, the novel HDAC inhibitor apicidin was found to be a potent inducer of HbF synthesis in K562 cells. The present data outline the role of histone hyperacetylation and p38 MAP kinase signaling as molecular targets for pharmacologic stimulation of HbF production in erythroid cells.

Topics: Amides; Azacitidine; Benzamides; Butyrates; Enzyme Inhibitors; Fetal Hemoglobin; Gene Expression Regulation; Gene Expression Regulation, Leukemic; Globins; Histone Deacetylase Inhibitors; Histone Deacetylases; Histones; Humans; Hydroxamic Acids; Hydroxyurea; Imidazoles; K562 Cells; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; Neoplasm Proteins; p38 Mitogen-Activated Protein Kinases; Peptides, Cyclic; Phenylbutyrates; Pyridines; Vorinostat