Targeting Cell Cycle and Signaling Pathways by Anticancer Agents in Prostate Cancer

• Main Authors: Yao-Ting Huang, 黃耀霆
• Other Authors: 鄧哲明
• Format: Others
• Language: zh-TW
• Published: 2005
• Online Access: http://ndltd.ncl.edu.tw/handle/36574069010650692607

博士 === 國立臺灣大學 === 藥理學研究所 === 93 === In the present studies we elucidate the anti-tumour mechanisms of Alisol B acetate, CIL-102 and YC-1 in the androgen-refractory human prostate adenocarcinoma cell line PC-3 cells. The anti-tumor potential of components from Chinese herbal medicines has been greatly concerned. In the first part, alisol B acetate, a triterpene from Alismatis rhizoma, induced apoptotic cell death in PC-3 cells in a time- and concentration-dependent manner. A good correlation between loss of mitochondrial membrane potential and apoptotic cell death was apparent, indicating the participation of mitochondria-related mechanism. Alisol B acetate induced Bax up-regulation and nuclear translocation; it also induced the activation of initiator caspase-8 and caspase-9, and executor caspase-3, suggesting the involvement of both extrinsic and intrinsic apoptosis pathways. Taken together, it is suggested that alisol B acetate induces apoptosis in PC-3 cells via a mitochondria-mediated mechanism with activation of caspase-8, -9 and -3. Furthermore, the Bax activation and translocation from the cytosol to nucleus might be a crucial response to the apoptotic effect. In the second part, CIL-102, a 4-anilinofuro[2,3-b]quinoline derivative, displayed a potent anti-tumor effect in PC-3 cells by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenylte-trazolium bromide and sulforhodamine B assays. Immunofluorescence microscopy and in vitro tubulin assembly assays indicated that CIL-102 bound to tubulin and disrupted microtubule organization. The data from flow cytometric analysis showed that CIL-102 caused cells to accumulate in G2/M phase and hypodiploid sub-G1 phase of the cell cycle. CIL-102-induced apoptosis was also characterized by immunofluorescence microscopy. Additionally, CIL-102 exposure induced up-regulation of cyclin B1 and p34cdc2 kinase activity while olomoucine, a p34cdc2 inhibitor, profoundly reduced the number of cells accumulated in mitotic phase. Moreover, CIL-102 also induced the increase of Bcl-2 phosphorylation, Cdc25C phosphorylation, and survivin expression. Although CIL-102-induced apoptosis was associated with activation of caspase-3, benzyloxycarbonyl-VAD-fluoromethyl ketone, a pancaspase inhibitor, only partially inhibited the apoptosis, and apoptosis inducing factor（AIF） was translocated from mitochondria to cytosol, suggesting the involvement of a caspase-independent pathway. Taken together, it is suggested that CIL-102 induces mitotic arrest and apoptosis by binding to tubulin and inhibiting tubulin polymerization. CIL-102 causes mitotic arrest, at least partly, by modulating cyclin-dependent kinases and then apoptosis executed by caspase and noncaspase pathways. Although the indazole compound, YC-1, is reported to exert anticancer activities in several cancer cell types, its target and mechanism of action have not been well explored. The objectives of this study were to ascertain whether YC-1 directly induces apoptosis in prostate cancer cells and to explore the anti-tumor mechanism. YC-1 suppressed growth of PC-3 cells in a concentration- and time-dependent manner. Apoptosis was determined using 4’,6-diamidino-2-phe-nylindole staining and cell cycle progression was examined by FACScan flow cytometry. YC-1 treatment showed chromatin condensation and increased the percentage of PC-3 cells in the hypodiploid sub-G1 phase, indicative of apoptosis. Additionally, cells exposed to YC-1 were found to induce activation of caspase-3 and cleavage of PARP. Translocation and activation of NF-κB were determined by immunofluorescent staining and ELISA, respectively. The results demonstrated that YC-1 abolished constitutive nuclear translocation and activation of NF-κB/p65. Furthermore, inhibition of IκBα phosphorylation and accumulation of IκBα were observed. The anti-tumor effects of YC-1 were evaluated by measuring the growth of tumor xenografts in SCID mice. The data demonstrated that the increase of tumor size was significantly inhibited by YC-1. It is suggested that the YC-1 displays potential anti-tumor activity in both in vitro and in vivo models and the suppression of NF-κB activation might explain YC-1-induced anti-tumor effect in PC-3 cells.