trichostatin-a and Leukemia--Myelogenous--Chronic--BCR-ABL-Positive

Histone deacetylase (HDAC) is a validated target for pursuing anticancer agents. However, obtaining a selective inhibitor against a given HDAC member remains a significant challenge. We report here the use of 1-hydroxypyridine-2-thione (1HPT) as a key pharmacophore for zinc-binding can result in highly selective HDAC inhibitors. 1HPT-6-carboxylic acid exhibits selective inhibition of HDAC6 with an IC50 of 150 nM that corresponds to a remarkable 0.9 ligand efficiency. Two analogs with simple amino acids shows nearly 600-fold selectivity among the eleven zinc-dependent HDACs. At low micromolar concentration these compounds inhibit the growth of HDAC8-overexpressing chronic myelogenous leukemia cells and specific form of acute myelogenous leukemia cells. Their potential mode of binding was examined by molecular docking and their stability was assessed in mouse and human plasma. Together the results suggest 1HPT analogs exhibit promising therapeutic potential for further development as anticancer agents to treat leukemia.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; Cell Survival; Crystallography, X-Ray; Dose-Response Relationship, Drug; Drug Discovery; Drug Screening Assays, Antitumor; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Mice; Molecular Docking Simulation; Molecular Structure; Pyridines; Structure-Activity Relationship; Thiones

An exciting recent approach to targeting transcription factors in cancer is to block formation of oncogenic complexes. We investigated whether interfering with the interaction of the transcription factor SALL4, which is critical for leukemic cell survival, and its epigenetic partner complex represents a novel therapeutic approach. The mechanism of SALL4 in promoting leukemogenesis is at least in part mediated by its repression of the tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN) through its interaction with a histone deacetylase (HDAC) complex. In this study, we demonstrate that a peptide can compete with SALL4 in interacting with the HDAC complex and reverse its effect on PTEN repression. Treating SALL4-expressing malignant cells with this peptide leads to cell death that can be rescued by a PTEN inhibitor. The antileukemic effect of this peptide can be confirmed on primary human leukemia cells in culture and in vivo, and is identical to that of down-regulation of SALL4 in these cells using an RNAi approach. In summary, our results demonstrate a novel peptide that can block the specific interaction between SALL4 and its epigenetic HDAC complex in regulating its target gene, PTEN. Furthermore, targeting SALL4 with this approach could be an innovative approach in treating leukemia.

Topics: Animals; Carcinoma, Hepatocellular; Drug Design; Endometrial Neoplasms; Epigenesis, Genetic; Female; Gene Expression Regulation, Leukemic; Histone Deacetylase 1; Histone Deacetylase 2; Histone Deacetylase Inhibitors; HL-60 Cells; Humans; Hydroxamic Acids; Leukemia, Monocytic, Acute; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Leukemia, Myeloid, Acute; Liver Neoplasms; Male; Mice; Mice, Inbred NOD; Mice, SCID; Peptide Fragments; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Transcription Factors; Xenograft Model Antitumor Assays

Epigenetic therapy has had a significant impact on the management of haematologic malignancies. The aim of this study was to assess whether 5-aza-CdR and TSA inhibit the growth of leukaemia cells and induce caspase-3-dependent apoptosis by upregulating RUNX3 expression.. K562 and Reh cells were treated with 5-aza-CdR, TSA or both compounds. RT-PCR and Western blot analyses were used to examine the expression of RUNX3 at the mRNA and protein levels, respectively. Immunofluorescence microscopy was used to detect the cellular location of RUNX3. Additionally, after K562 cells were transfected with RUNX3, apoptosis and proliferation were studied using Annexin V staining and MTT assays.. The expression of RUNX3 in leukaemia cell lines was markedly less than that in the controls. Demethylating drug 5-aza-CdR could induce RUNX3 expression, but the combination of TSA and 5-aza-CdR had a greater effect than did treatment with a single compound. The combination of 5-aza-CdR and TSA induced the translocation of RUNX3 from the cytoplasm into the nucleus. TSA enhanced apoptosis induced by 5-aza-CdR, and Annexin V and Hoechst 33258 staining showed that the combination induced apoptosis but not necrosis. Furthermore, apoptosis was dependent on the caspase-3 pathway. RUNX3 overexpression in K562 cells led to growth inhibition and apoptosis and potentiated the effects of 5-aza-CdR induction.. RUNX3 plays an important role in leukaemia cellular functions, and the induction of RUNX3-mediated effects may contribute to the therapeutic value of combination TSA and 5-aza-CdR treatment.

Topics: Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Azacitidine; Blotting, Western; Caspase 3; Cell Line, Tumor; Cell Proliferation; Core Binding Factor Alpha 3 Subunit; CpG Islands; Fluorescent Antibody Technique; Gene Expression Regulation, Neoplastic; Humans; Hydroxamic Acids; K562 Cells; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Reverse Transcriptase Polymerase Chain Reaction; Up-Regulation

To identify novel proteins involved in multidrug resistance in chronic myeloid leukemia (CML).. Comparative proteomics was used to screen multidrug resistance-related proteins from K562 and K562/A02; the differently expressed proteins were further confirmed by western blot and real-time PCR. short hairpin RNA (shRNA) assay was applied to determine the relationship between candidate protein and adriamycin resistance. Bisulfite sequencing was carried out to assess methylation status of candidate multidrug resistance-related gene promoter. K562/A02 was treated with 5-azacytidine or trichostatin A (TSA); multidrug resistance phenotype and corresponding protein or gene changes were detected.. Seventeen proteins with altered abundances of more than twofold were detected, among which mitochondrial ATPase in K562/A02 was significantly down-regulated. Suppressing mitochondrial ATPase by shRNA could enhance adriamycin resistance and antiapoptosis activity of K562. The promoter hypermethylation in mitochondrial ATPase was found to be attributed to the adriamycin-resistant phenotype of both K562/A02 (methylated frequency 18.18%) and CML primary cells in accelerated phase (methylated frequency 7.95%) or blast crisis (methylated frequency 26.59%). Inhibition of hypermethylation increased adriamycin sensitivity of K562/A02. A synergistic effect on reversing adriamycin-resistant phenotype was obtained when 5-azacytidine was combined with TSA.. Down-regulation of mitochondrial ATPase can lead to adriamycin resistance in CML and the mechanism is associated with DNA methylation regulation.

Topics: Antibiotics, Antineoplastic; Antimetabolites, Antineoplastic; Apoptosis; ATP Binding Cassette Transporter, Subfamily B, Member 1; Azacitidine; Base Sequence; Blotting, Western; DNA Methylation; Down-Regulation; Doxorubicin; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Electrophoresis, Gel, Two-Dimensional; Flow Cytometry; Gene Expression Regulation, Enzymologic; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Mitochondrial Proton-Translocating ATPases; Molecular Sequence Data; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tumor Cells, Cultured

In a variety of malignant cells the prostate-apoptosis-response-gene-4 (Par-4) induces increased sensitivity towards chemotherapeutic agents by down-regulating anti-apoptotic B-cell lymphoma-gene 2 (Bcl-2). Hypothesizing that Par-4 also influences apoptosis in myeloid cell lines, we tested this hypothesis by stably transfecting bcr-abl transformed-K562 cells with a Par-4-expressing vector. Here we demonstrate that over-expression of Par-4 in K562 cells up-regulates expression levels of Bcl-2 and death-associated protein (Daxx). Upon treatment with different chemotherapeutic agents, Fas- or TRAIL agonistic antibodies, Par-4-positive cells did not exhibit an increased rate of apoptosis as compared to Par-4-negative control cells. However, incubation with histone deacetylase (HDAC)-inhibitors Trichostatin A (TSA) and LAQ824 or the tyrosinkinase inhibitor Imatinib (STI571) increased the rate of apoptosis in Par-4-positive K562 cells. Assessing the underlying molecular mechanisms for the Par-4-induced response to HDAC-inhibitors and STI571 we provide evidence, that these effects are associated with a down-regulation of Daxx, enforced activation of caspases and enhanced cleavage of cellular inhibitor of apoptosis (cIAP)-1 and -2.

Topics: Adaptor Proteins, Signal Transducing; Antibodies; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Benzamides; Carrier Proteins; Caspases; Co-Repressor Proteins; Down-Regulation; Drug Screening Assays, Antitumor; Enzyme Inhibitors; fas Receptor; Fusion Proteins, bcr-abl; Gene Expression; Histone Deacetylase Inhibitors; Humans; Hydroxamic Acids; Imatinib Mesylate; Inhibitor of Apoptosis Proteins; Intracellular Signaling Peptides and Proteins; K562 Cells; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Male; Membrane Glycoproteins; Molecular Chaperones; Myeloid Cells; Nuclear Proteins; Piperazines; Proteins; Proto-Oncogene Proteins c-bcl-2; Pyrimidines; TNF-Related Apoptosis-Inducing Ligand; Tumor Necrosis Factor-alpha; Up-Regulation