培美曲塞二钠与吉非替尼对肺腺癌细胞的放射增敏作用

Combined effects of pemetrexed disodium and gefitinib on radiation sensitivity in human lung adenocarcinoma cells

目的:研究培美曲塞二钠与吉非替尼单独及联合运用对肺腺癌细胞放射增敏作用及机制。方法:单独或联合使用培美曲塞二钠和吉非替尼处理人肺腺癌细胞株A549,使用噻唑蓝比色法(MTT)检测药物对细胞增殖的影响;采用克隆形成试验检测药物联合放射线对细胞的生长抑制作用;使用流式细胞术检测细胞周期和细胞凋亡的变化;Western blot检测细胞内Akt和p-Akt的表达变化。结果:培美曲塞二钠和吉非替尼对人肺腺癌细胞A549增殖具有抑制作用,体外培养克隆形成实验分析显示,放疗+两药联用组的准阈剂量(Dq)、平均致死剂量(D0)及2 Gy照射时的存活分数(SF2)均明显低于单纯放疗组、放疗+培美曲塞二钠组及放疗+吉非替尼组;培美曲塞二钠和吉非替尼放射增敏比(sensitivity enhanced ratio,SER)分别为1.17和1.48,两药联合SER为2.62。培美曲塞二钠和吉非替尼使细胞阻滞在G1/G0期,与放射线联合作用后,S期细胞比例进一步降低。培美曲塞二钠和吉非替尼均可以提高放射线诱导的凋亡,两药联用后细胞的凋亡率最高。药物作用前后、照射前后及药物增敏照射后Akt水平没有明显改变;培美曲塞二钠和吉非替尼均能抑制p-Akt表达;与单纯照射组相比,培美曲塞二钠和吉非替尼联合照射组p-Akt水平显著减低,p-Akt/Akt的比值最低。结论:培美曲塞二钠和吉非替尼各自均有放射增敏作用,两药联合使用可以明显提高放疗效果,其机制可能与减少S期细胞、增强放射线诱导凋亡和抑制PI3K/Akt信号转导通路有关。

Objective： To study the radiosensitization effects of pemetrexed disodium and gefitinib on human lung adenocarcinoma cells. Methods： Human lung adenocarcinoma A549 cells were treated with pemetrexed and/or gefinitib, and cell proliferation was examined by MTT. Colony formation was performed to detect growth inhibition effects of pemetrexed and/or gefinitib combined with radiation on A549 cells. Flow cytometry was performed to detect the changes of cell cycle and apoptosis. Expressions of intracellular Akt and p-Akt were detected by Western blot. Results： Pemetrexed and gefitinib inhibited cell proliferation of human lung adenocarcinoma A549 cells. Compared with cells treated with radiation only, radiation＋pemetrexed, and radiation＋gefitinib, the Dq, D0 and SF2 of cells treated with radiation＋pemetrexed＋gefitinib were significantly decreased. The SERs of cells treated with pemetrexed, gefitinib, and pemetrexed＋gefitinib were 1.17, 1.48 and 2.62, respectively. Both pemetrexed and gefitinib induced G1/G0 phase arrest. When combined with radiation, the proportion of S phase cells was further reduced. Both pemetrexed and gefitinib enhanced radiation-induced apoptosis by decreasing p-Akt level. The p-Akt level was lowest in the cells treated with radiation＋pemetrexed＋gefitinib. Conclusion： Both pemetrexed and gefitinib have radiosensitization effects on A549 cells. Combination of the two drugs significantly improves the inhibitory effect of radiation through reducing S phase cells, enhances radiation induced apoptosis, and inhibits PI3K/Akt signaling pathway. This combination regimen may provide a promising strategy for lung cancer therapy.