EGFR TKIs 联合化疗的给药顺序对NSCLC 细胞促凋亡的影响

Effect of EGFR-TKIs and chemotherapy combinations and administration sequence on apoptosis of NSCLC cells

摘要:

目的 探究表皮生长因子受体络氨酸激酶抑制剂（EGFR TKIs）与化疗药物单用、联用，以及顺 序给药时，对获得性TKI 耐药的非小细胞肺癌（NSCLC）细胞系凋亡的影响。方法 选用存在TKI 耐药基 因突变（T790M，cMET）的NSCLC 细胞系PC9ER、H1975 和HCC827GR，以及TKI 敏感基因突变（19 外 显子突变）的细胞系PC9 和HCC827。采用MTS 法检测化疗药物顺铂和紫杉醇，以及TKI 厄洛替尼单用、 联用，或者顺序给药干预各NSCLC 细胞系时的细胞活性状态，并利用集落形成试验加以验证。采用Western blot 检测各处理组蛋白裂解液中细胞凋亡标志物cPARP 含量变化。结果 MTS 法和集落形成试验结果发 现，EGFR TKIs 联合化疗能协同增加NSCLC 细胞系的细胞毒性。同时给予EGFR TKIs 和化疗或先化疗再用 EGFR TKIs，可获得最佳干预效果。Western bolt 检测结果表明，联合用药后cPARP 蛋白表达升高。而且先 化疗后EGFR TKIs 干预比两者顺序颠倒的促凋亡作用更强。结论 先化疗后EGFR TKIs 干预的最优给药顺 序可使对获得性TKI 耐药的NSCLC 细胞系产生最强的促凋亡作用。

Abstract:

Objective To explore the effect of epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) and chemotherapeutic drug alone, concurrent and sequential administration on apoptosis of non-small cell lung cancer (NSCLC) cells with acquired TKI-resistance. Methods NSCLC cell lines with TKI-resistance gene mutations (T790M and cMET) such as PC9ER, H1975 and HCC827GR as well as TKI-sensitive gene mutation (exon 19) cell lines PC9 and HCC827 were selected. Cell viability was detected by MTS assay when chemotherapy drugs Cisplatin, Paclitaxel and TKI Erlotinib were used alone, in combination or in sequential administration for intervention of NSCLC cell lines, and verified using colony formation assay. The content of apoptosis marker cPARP in cell lysate was detected in each treatment group by Western blot. Results MTS assay and colony formation revealed that the combination of EGFR-TKIs and chemotherapeutic drugs could synergistically increase the cytotoxicity to NSCLC cell lines. Simultaneous use of EGFR-TKIs and chemotherapeutic drugs or chemotherapy followed by EGFR-TKIs provided the maximum intervention. Western blot showed that the apoptotic marker cPARP expression was significantly increased when two drugs were simultaneously used. Moreover, chemotherapy then EGFR-TKIs intervention could obtain a more powerful pro-apoptotic performance than the reverse order intervention. Conclusions The sequence of chemotherapy followed by EGFR-TKI is likely to be the most active strategy, and could have the greatest effect on apoptosis in cell lines with acquired TKI-resistance.