mTORC1抑制剂增强KRAS突变型肺癌细胞对化疗药物敏感性药效及机制研究

Effect and Mechanism of mTORC1 Inhibitor Enhancing the Sensitivity of KRAS-mutant Lung Cancer Cells to Chemotherapy Drugs

目的:探讨Kirsten大鼠肉瘤病毒基因同源物(KRAS)突变型肺癌细胞对化疗药物敏感性降低的机制及哺乳动物雷帕霉素靶蛋白1(m TORC1)抑制剂增强KRAS突变型肺癌细胞对化疗药物敏感性药效及机制。方法:选取在本院就诊并确诊为KRAS突变型非小细胞肺癌的患者53例,通过Q-PCR及Western blot检测患者肿瘤病变部位m RNA及蛋白水平m TORC1相关因子的表达,对比产生耐药及未产生耐药患者体内以上因子的差异。构建KRAS突变型肺癌细胞,采用MTT及Western blot检测不同浓度m TORC1抑制剂或m TORC1抑制剂与顺铂联合使用对KRAS突变型肺癌细胞的抑制增殖、m TORC1活性底物S6K和4EBP1的磷酸化(p)水平的作用。结果:与未耐药KRAS突变型肺癌患者相比,耐药患者体内m RNA及蛋白水平m TORC1相关因子的表达均显著上升(P<0.05);低浓度m TORC1抑制剂即可对KRAS突变型肺癌细胞有明显抑制增殖作用(P<0.05),呈剂量依赖性;且同浓度m TORC1抑制剂与顺铂联合使用时抑制细胞增殖作用显著增加(P<0.05)。KRAS突变型肺癌细胞给予顺铂刺激时,P-4EBP及P-S6K水平显著提高(P<0.05),m TORC1被显著激活,这一升高可被m TORC1抑制剂降低(P<0.05)。结论:KRAS突变型肺癌患者对化疗药物敏感性的产生可能与m TORC1的激活有关,m TORC1抑制剂可以通过抑制m TORC1的激活增强KRAS突变型肺癌患者对化疗药物敏感性;且顺铂与m TORC1抑制剂联合使用具有更好的抑制KRAS突变型肺癌细胞增殖的作用。

Objective:To investigate the mechanism of resistance to chemotherapy in patients with KRAS mutated lung cancer and study the mechanism of m TORC1 inhibitor enhancing the sensitivity to chemotherapy in KRAS mutated lung cancer cells.Methods:Totally 53 patients who were diagnosed as KRAS-mutant lung cancer were selected to detect the expressions of m TORC1 related factors in tumor lesions by Q-PCR and Western blot.And then KRAS-mutant cells were constructed,and the proliferation of KRAS-mutant cancer cells and the phosphorylation of m TORC1 active substrates S6 K and 4 EBP1 were compared by using MTT or Western blot when m TORC1 inhibitor singly or combined with cisplatin was added to the medium.Results:Both the expressions of m RNA and protein of m TORC1 related factors in the patients with drug resistance were significantly increased(P<0.05)when compared with the patients without drug resistance.m TORC1 inhibitor could significantly inhibit the proliferation of KRAS-mutant lung cancer cells(P<0.05)even at a low dose,and m TORC1 inhibitor could inhibit the proliferation of KRAS-mutant lung cancer cells in a dose-dependent manner.In addition,the inhibiting efficacy of cell proliferation was significantly increased when cisplatin was combined with m TORC1 inhibitor(P<0.05).When KRAS-mutant lung cancer cells were stimulated by cisplatin,the expressions of P-4 EBP and P-S6 K were significantly increased(P<0.05),and m TORC1 was significantly activated.However,the increase was reduced by m TORC1 inhibitors(P<0.05).Conclusion:Hyperactivated mammalian target of rapamycin(m TOR)pathway is a characteristic hallmark of KRAS-mutant lung adenocarcinoma after chemotherapy treatment,and KRAS-mutant lung cancer cells rely on persistent m TOR signaling to resist chemotherapeutic drugs.m TORC1 inhibitors can enhance the sensitivity of KRAS mutated lung cancer cells to chemotherapy.In addition,cisplatin combined with m TORC1 inhibitor has a better efficacy in inhibiting the proliferation of KRAS-mutant lung cancer cells.