Diallyltrisulfide誘導人類口腔癌細胞凋亡機轉之研究

Abstract

In Taiwan, Oral cancer is the fourth leading cause of cancer-related deaths in male population. Despite recent advances in radiotherapy and chemotherapy, the overall 5-year survival rate for patients with oral squamous cell carcinoma (OSCC) has not changed during the past two decades. More researches are needed to develop new therapertic and prevention methodologies to increase overall survival and improve patient’s living quality. Continuous investigation of new chemotherapeutic agents is thus needed. Epidemiological studies support the premise that dietary intake of Allium vegetables (e.g., garlic, onions and so forth) may lower the risk of varies types of cancer. Garlic-derived compounds have been shown to offer protection against oral tumor in animal models induced by carcinogens. Garlic-derived organosulfur compounds (OSCs) including diallyl sulfide (DAS), diallyl disulfide (DADS) and diallyl trisulfide (DATS) have the ability to suppress proliferation of human cancer cells by causing apoptosis. The different cell lines will be probably end up same or different mechanism of proliferation inhibition and apoptosis. However, the effects of OSCs on oral cancer proliferation is not clearly confirmed. In our studies, the viability of oral cancer SAS and Ca9-22 cells were reduced upon a 24h exposure to DADS and DATS, however, DATS has better efficiency in inhibiting OSCC proliferation. The data indicated that SAS and Ca9-22 cells were reduced significantly on exposure to DATS in a concentration-dependent manner with each IC50 about 34 μM and 21 μM respectively. Per cell cycle distribution via flow cytometric analysis process, a 3h exposure of SAS cells to DATS (20μM) resulted increasing G2/M fraction. A 12h treatment of SAS cells with DATS resulted in a increase in the fraction of sub-G0/G1 cells. Western blot data showed that DATS treatment increased the protein level of p-Cdk1Tyr15 and Wee1, on the contrary, the protein level of Cdc25C and Cdk1 decreased. Our studies defined the mechanism of DATS-mediated G2 phase arrest, not mitotic arrest. In addition, the protein level alteration of PARP indicated that DATS-induced apoptosis in human oral cancer cells. Previous studies illustrated that DATS stimulated the intrinsic apoptotic pathway. In our study, DATS-induced apoptosis was decreased in the presence of caspase 8 or caspase 9 inhibitors (Z-LEHD-FMK, Z-IETD-FMK) in SAS and Ca9-22 cells, and the result was more effective when both of caspase 8 and capase 9 inhibitors treated. The data showed that DATS-induced apoptosis by activating intrinsic- and extrinsic-apoptosis pathway. Western blot data confirmed that DATS activates both apoptosis pathway in oral cancer cells with up-regulation of cytochrome c, DR5 and FADD. DATS-induced cell death was inhibited by N-acetyl cysteine in SAS and Ca9-22 cells, indicated that involvement of ROS was an important mechanism for DATS-induced apoptosis in human oral cancer cells.