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目的探讨甲氧基-(S)-三苯甲基-L-半胱氨酸[S(MeO)TLC]靶向抑制有丝分裂驱动蛋白(KSP)在体外试验中对多西紫杉醇耐药前列腺癌的治疗效果。方法培养建立多西紫杉醇耐药前列腺癌细胞株(PC3R),Western blotting检测不同细胞株(PC3、DU145及PC3R)KSP蛋白表达水平;实验分空白对照组、PC3R组及PC3组,MTT及台盼兰染色细胞活性实验观察S(MeO)TLC抑制耐药细胞增殖效果;以耐药细胞株PC3R为研究对象,实验分为S(MeO)TLC组及对照组,Heochst染色、流式细胞仪及RT-PCR技术检测S(MeO)TLC诱导耐药细胞凋亡的效果。结果PC3、DU145、PC3R细胞KSP蛋白表达量差异无统计学意义(P>0.05)。PC3R细胞S(MeO)TLC半数抑制浓度(IC50)为120 nmol/L,与PC3细胞(IC50:106 nmol/L)比较,差异无统计学意义(P>0.05)。PC3R细胞给予S(MeO)TLC作用24 h后,87.9%细胞停滞在有丝分裂期;给药72 h后,有丝分裂期停滞细胞显著凋亡。与对照组相比,S(MeO)TLC组PC3R细胞Caspase-3(t=13.445,P=0.000 2)、Caspase-8(t=9.494,P=0.000 7)、Caspase-9(t=5.198,P=0.007)、PARP(t=19.097,P=0.000 04)及自噬标志指标LC3(t=22.609,P=0.000 02)和Beclin1(t=61.266,P=0.000 000 4)的mRNA量均明显升高。结论前列腺癌多西紫杉醇耐药与KSP蛋白表达无关,KSP蛋白靶向抑制剂S(MeO)TLC能够有效抑制多西紫杉醇耐药前列腺癌细胞增殖并诱导其凋亡。在此过程中,内源性和外源性Caspase依赖性的凋亡途径均发挥了重要作用,且自噬可能发挥协同作用。
OBJECTIVE: To investigate the inhibitory effect of methoxy-S-trityl-L-cysteine (S (MeO) TLC] on the inhibition of mitotic kinesin (KSP) against docetaxel-resistant prostate cancer in vitro treatment effect. Methods To establish a multidrug-resistant prostate cancer cell line (PC3R) and detect the expression of KSP protein in different cell lines (PC3, DU145 and PC3R) by Western blotting. The experiment was divided into blank control group, PC3R group and PC3 group, (MeO) TLC was used to inhibit the proliferation of drug-resistant cells. The drug-resistant cell line PC3R was used as experimental object. The experiment was divided into S (MeO) TLC group and control group. Heochst staining, flow cytometry and RT -PCR technique was used to detect the effect of S (MeO) TLC on drug-resistant cell apoptosis. Results The expression of KSP protein in PC3, DU145 and PC3R cells showed no significant difference (P> 0.05). The IC50 of S (MeO) TLC for PC3R cells was 120 nmol / L, which was not significantly different from that of PC3 cells (IC50: 106 nmol / L) (P> 0.05). After treated with S (MeO) for 24 h, 87.9% of the cells in the PC3R cells were arrested in mitosis; 72 h after the administration, the arrested cells in mitosis were significantly apoptotic. Compared with the control group, Caspase-3 (t = 13.445, P = 0.0002), Caspase-8 (t = 9.494, P = 0.007), PARP (t = 19.097, P = 0.000 04) and autophagy markers LC3 (t = 22.609, P = 0.000 02) and Beclin1 Rise. Conclusions Docetaxel resistance in prostate cancer has nothing to do with the expression of KSP protein. S (MeO) TLC, a KSP protein inhibitor, can effectively inhibit the proliferation and induce the apoptosis of docetaxel-resistant prostate cancer cells. In this process, both endogenous and exogenous caspase-dependent apoptotic pathways play an important role, and autophagy may play a synergistic role.