蛋白激酶Cβ抑制剂Enzastaurin逆转肿瘤多药耐药机制的研究

Reversing mechanism of protein kinase Cβ inhibitor enzastaurin on tumor multidrug resistance

目的探讨蛋白激酶Cβ抑制剂Enzastaurin逆转多药耐药(MDR)的作用及机制。方法 Enzastaurin处理高表达ABCB1、ABCC1及ABCG2的MDR基因细胞株后,通过MTT法测定细胞毒作用,流式细胞仪测定阿霉素及罗丹明123在细胞内累积,Western blotting测定蛋白表达,应用半定量RT-PCR测定m RNA表达水平及采用化学发光法测定ATP酶活性。结果 Enzastaurin可逆转ABCB1介导MDR,对ABCC1及ABCG2介导MDR无逆转作用;Enzastaurin抑制ABCB1药物外排功能而逆转肿瘤MDR,但与ABCB1表达及阻断Akt及Erk1/2磷酸化无关;Enzastaurin通过抑制ATP酶激活而抑制ABCB1药物外排泵功能。结论 Enzastaurin在体外可通过抑制药物外排泵ATP酶活性逆转ABCB1介导MDR作用。

Objective To investigate the reversing effect and mechanism of protein kinase Cβ inhibitor enzastaurin on tumor multidrug resistance（MDR）. Methods After MDR cell lines with ABCB1, ABCC1 and ABCG2 overexpression were treated with enzastaurin, the cytotoxicity, intracellular accumulation of doxorubicin and rhodamine 123, protein expression, ABCB1 m RNA, and ATPase activity were determined by MTT, flow cytometry, Western blotting, RT-PCR and chemiluminescence, respectively. Results ABCB1-mediated MDR was reversed by enzastaurin, but ABCC1 and ABCG2-mediated MDR was not. The reversing effect of enzastaurin was related to the suppresion of ABCB1 drug efflux via inhibiting ATPase activity, but not to ABCB1 expression and Akt and ERK1/2 phosphorylation inhibition. Conclusion Enzastaurin can in vitro reverse ABCB1-mediated MDR by inhibiting ATPase activity on the drug efflux pump.