嵌合分子靶向降解前列腺癌LNCaP细胞表达的雄激素受体

Targeting androgen receptors for degradation by chimeric molecules in prostate cancer LNCaP cells

目的观察嵌合分子双氢睾酮-蛋白靶向降解嵌合分子（DHT—PROTAC）通过泛素蛋白酶体途径易化降解前列腺癌LNCaP细胞表达的雄激素受体以及雄激素受体降解后该细胞增殖和活力的变化。方法细胞爬片免疫组织化学和Western blot技术检测DHT—PROTAC处理后LN—CaP细胞中雄激素受体蛋白的改变。DHT-PROTAC分别处理前列腺癌LNCaP细胞、PC-3细胞、肾癌786-O细胞，噻唑蓝（MTY）比色法检测细胞活力，细胞计数并绘制生长曲线观察细胞增殖能力。结果随着DHT—PROTAC浓度升高，LNCaP细胞表达雄激素受体减少，并呈剂量依赖性，Western blot检测结果表现为信号减弱，细胞免疫组织化学见阳性细胞明显减少，随着处理浓度升高，细胞阳性率分别为93．22％、85．65％、61．94％、1．71％。DHT、ALAPYIP-（Arg）8肽段能拮抗DHT—PRO—TAC作用，蛋白酶体抑制剂抑制雄激素受体降解；噻唑蓝（MTT）细胞活力试验及细胞计数显示LNCaP细胞活力和增殖受抑制。结论嵌合分子DHT-PROTAC能易化降解LNCaP细胞表达的雄激素受体，并抑制该细胞的生长和增殖。

Objective To investigate the degradation of androgen receptor （AR） by chimeric molecules （DHT-PROTAC） via ubiquitin-proteasome pathway, and explore the proliferation and viability of prostate cancer LNCaP cells. Methods LNCaP cells were treated with different concentrations of DHT-PROTAC. AR levels in LNCaP cells were detected by immunohistochemistry （IHC） and Western blot. Both LNCaP and PC-3 cells were treated with DHT-PROTAC, then cell counting and growth curve were done to analyze the proliferation of cells. The cell viability was evaluated by MTT assay. Results Band signals of the DHT-PROTAC-treated group were obviously attenuated compared to the DMSO-or PBS-treated controls, and degradation of AR was concentration dependent. IHC also depicted that AR-positive cells were notably decreased, and the positive-rate was reduced with the increases of DHT-PROTAC. Either DHT or ALAPYIP-（Arg） 8 peptide could antognize the effects of DHT-PROTAC. Inhibition of cell proliferation and decreased viability were observed in LNCaP cells. Conclusion Chimeric molecule （DHT-PROTAC） can facilitate the degradation of AR and repress the growth of LNCaP cells.