組蛋白去乙醯化抑制劑Suberoylanilide Hydroxamic Acid (SAHA)誘導人類口腔癌細胞凋亡機轉之研究

Abstract

Oral cancer is the fourth leading cause of cancer-related deaths in male population in Taiwan. Despite recent advances in radiotherapy and chemotherapy, the survival of patients with oral cancer has not improved significantly. Continued investigation of new chemotherapeutic agents is thus needed. Our recent studies have shown that histone deacetylase 2 (HDAC2) is overexpressed in 70% of oral cancer specimens. Furthermore, recent studies have shown that inhibitors of HDACs (HDACIs) possess antitumor activity and are well tolerated, supporting the idea that their use might develop as a specific strategy for cancer treatment. In this study, we investigated the effects and mechanisms of suberoylanilide hydroxamic acid (SAHA, one of the most potent HDAC inhibitors) on OSCC cell lines SAS and Ca9-22. Here, we demonstrated that SAHA induces apoptosis in human oral cancer cell lines SAS and Ca9-22 as evidenced by nuclear condensation, TUNEL labeling and cleavage of PARP. Apoptosis induced by SAHA was both time- and dose-dependent; however, the mechanisms are different in these two cells. In SAS cells, SAHA treatment induced DR5, FAS/FASL, FADD, caspase-8, -9 activation and Bid cleavage. In addition, SAHA treatment induced reactive oxygen species (ROS) production as detected by H2DCFDA fluorescence. Pretreatment of cells with N-acetyl cysteine (NAC) reduced the up-regulation of DR5, FAS, FADD and completely inhibited SAHA-induced apoptosis. These results indicated that ROS was an important mechanism for SAHA-induced apoptosis in SAS cells. SAHA-induced apoptosis was also completely inhibited in the presence of caspase 8 or caspase 9 inhibitors (Z-LEHD-FMK, Z-IETD-FMK). Taking together, SAHA induced apoptosis via subsequent induction of ROS, DR5, FADD, FAS/FASL, caspase-8 activation, Bid cleavage and then activation of mitochondrial pathway. In Ca9-22 cells, SAHA induced Bax protein expression, caspase 9 activation. In addition, we found SAHA down-regulated the expression of Bcl-2. Treatment caspase 9 inhibitor (Z-IETD-FMK) decreased SAHA induced apoptosis and the result was not more effective when both of caspase 8 and capase 9 inhibitors were treated. These data showed that SAHA-induced apoptosis by activating intrinsic- apoptosis pathway. We further evaluated the potential combinative effect of TRAIL and SAHA in OSCC cell lines. Compared with either TRAIL (20ng/ml) or SAHA (1