厄洛替尼对非小细胞肺癌H1299细胞放射增敏研究

Mechanism of radiosensitization of erlotinib to non-small cell lung cancer cell line H1299

目的探讨表皮生长因子受体酪氨酸激酶抑制剂对人非小细胞肺癌细胞放射敏感性的影响及其可能的机制。方法体外培养人非小细胞肺癌细胞H1299。CCK-8检测厄洛替尼对H1299的毒性作用,并计算IC50及IC20,将IC20作为后续试验的药物作用浓度。克隆形成实验检测射线联合厄洛替尼对H1299的作用,计算放射敏感性参数,绘制细胞存活曲线。流式细胞术检测细胞周期分布及凋亡。Western blot检测细胞EGFR/PI3K/AKT通路及凋亡相关蛋白表达。结果厄洛替尼对H1299具有一定增殖抑制作用,IC50为27.3μmol/L、IC20为3.3μmol/L。X射线联合IC20浓度厄洛替尼能够降低H1299的克隆能力,使G0/G1期、G2/M期比例增加,S期减少比例,细胞凋亡增加;抑制pEGFR及pAKT蛋白表达,增加凋亡相关蛋白Active Caspase 3、Cleaved PARP表达。结论厄洛替尼对H1299具有放射增敏作用,其可能的机制是厄洛替尼联合射线抑制EGFR/PI3K/AKT通路,降低细胞损伤修复能力,改变细胞生长周期,诱导细胞凋亡。

Objective To investigate the effect of epidermal growth factor receptor tyrosine kinase inhibitor on the radiosensitivity of human non-small cell lung cancer cells and its possible mechanism.Methods Human non-small cell lung cancer cells H1299 were cultured in vitro.CCK-8 was used to detect the toxic effects of erlotinib on H1299 cells,IC50 and IC20 were calculated,and IC20 was utilized as the drug concentration for subsequent experiments.The colony formation assay was performed to identifiy the effect of X-ray combined with erlotinib on H1299 cells,the radiosensitivity parameters were calculated,and the cell survival curves were delineated.Flow cytometry was conducted to detect the cell cycle distribution and apoptosis.Western blot analysis was used to detect the expression of EGFR/PI3K/AKT pathway and apoptosis-related proteins.Results Erlotinib exerted inhibitory effect upon the proliferation of H1299 cells,IC50 was calculated as 27.3μmol/L,and 3.3μmol/L for IC20.X-ray combined with IC20 concentration of erlotinib could reduce the cloning ability of H1299,increased the proportion of G0/G1 phase and G2/M phase,decreased the proportion of S phase,aggravated cell apoptosis,down-regulated the expression of pEGFR and pAKT proteins,and up-regulated the expression of apoptosis-related proteins including Active Caspase 3 and Cleaved PARP.Conclusions Erlotinib exerts a radiosensitizing effect on H1299.The possible mechanism is that erlotinib combined with radiation can suppress the EGFR/PI3K/AKT pathway,reduce the ability of repairing cell damage,change cell growth cycle and induce cell apoptosis.