CPTH6, a Thiazole Derivative, Induces Histone Hypoacetylation and Apoptosis in Human Leukemia Cells

Abstract

Purpose: We previously identified novel thiazole derivatives able to reduce histone acetylation and histone acetyltransferase (HAT) activity in yeast. Among these compounds, 3-methylcyclopentylidene-[4-(4 0 -chlorophenyl)thiazol-2-yl]hydrazone (CPTH6) has been selected and used throughout this study.Experimental Design: The effect of CPTH6 on histone acetylation, cell viability and differentiation, cellcycle distribution, and apoptosis in a panel of acute myeloid leukemia and solid tumor cell lines has been evaluated.Results: Here, we showed that CPTH6 leads to an inhibition of Gcn5 and pCAF HAT activity. Moreover, it inhibits H3/H4 histones and a-tubulin acetylation of a panel of leukemia cell lines. Concentration-and time-dependent inhibition of cell viability, paralleled by accumulation of cells in the G 0 /G 1 phase and depletion from the S/G 2 M phases, was observed. The role of mitochondrial pathway on CPTH6-induced apoptosis was shown, being a decrease of mitochondrial membrane potential and the release of cytochrome c, from mitochondria to cytosol, induced by CPTH6. Also the involvement of Bcl-2 and Bcl-xL on CPTH6-induced apoptosis was found after overexpression of the two proteins in leukemia cells. Solid tumor cell lines from several origins were shown to be differently sensitive to CPTH6 treatment in terms of cell viability, and a correlation between the inhibitory efficacy on H3/H4 histones acetylation and cytotoxicity was found. Differentiating effect on leukemia and neuroblastoma cell lines was also induced by CPTH6.Conclusions: These results make CPTH6 a suitable tool for discovery of molecular targets of HAT and, potentially, for the development of new anticancer therapies, which warrants further investigations.

Histone acetyltransferase inhibitor CPTH6 preferentially targets lung cancer stem-like cells

“…Consistent with the HAT inhibitory activity of CPTH6 [12], decreased acetylation of both histone H3 and α-tubulin was observed in H1299 cells, among the most sensitive cell lines, by Western blot analysis after 24h treatment with CPTH6 ( Figure 1B). In order to investigate whether CPTH6 inhibition of cell viability was associated with cell death in NSCLC cells, H1299 cells were treated with CPTH6 for 24h at concentrations ranging from 20 to 100μM, and cell survival was assessed.…”

“…To evaluate the specific functional significance of HAT inhibition in human NSCLC, we explored cell proliferation of nine commercially available established NSCLC cell lines exposed to increasing concentrations of CPTH6, a novel Gcn5 and pCAF HAT inhibitor [12].…”

“…Western blot analyses of total protein extracts were performed as previously described [12]. Immunodetection was performed using antibodies directed to: H3 histone (abcam), H3 acetylated histone (Millipore, Billerica, MA, USA), α-tubulin (DM1A) (Santa Cruz Biotechnology, Santa Cruz, CA, USA), acetyl-α-tubulin (K40) (SigmaAldrich), γH2AX (Ser139) (Millipore), PARP cleaved (Millipore), PARP (Santa Cruz Biotechnology) β-actin (Sigma-Aldrich), HSP72/73 (Calbiochem, San Diego, CA, USA), anti-mouse or anti-rabbit immunoglobulin G (IgG)-horseradish peroxidase conjugated antibodies (Cell Signaling; Amersham Biosciences, Freiburg, Germany).…”

“…Several molecules with HAT inhibitory activity have been identified and some of them showed to induce cell death preferentially in cancer cells when compared to normal ones [6][7][8][9][10]. Among them, the thiazole derivative 3-methyl-cyclopentylidene-[4-(4'-chlorophenyl)thiazol-2-yl)]hydrazone (CPTH6), has been characterized by our group as a novel Gcn5 and pCAF HAT inhibitor, able to activate the apoptotic program and to modulate the autophagic flux in a panel of tumor cell lines [11,12]. Cancer stem cells (CSCs) have been proposed as potential culprits not only of the tumor initiation and progression, but also of therapeutic failure and tumor relapse, mainly because of their intrinsic resistance to conventional drugs [13].…”

Affinity purification-mass spectrometry analysis of bcl-2 interactome identified SLIRP as a novel interacting protein

“…Flow cytometric analysis (BD Accuri™ C6, BD Biosciences, Franklin Lakes, NJ, USA) was performed to evaluate apoptosis by Annexin V-FITC/PI staining 35 and ROS level. 36 Cells were transfected by using siRNA against SLIRP or scramble siRNA and after 24 h medium was replaced with medium containing 0.5 μM CPT or 20 μM DDP.…”