活性氧在导致吉非替尼耐药细胞凋亡中的作用

摘要

Objective: To investigate the specific cytotoxicity of the anticancer agent NSC-741909 on the growth of a non-small cell lung cancer ( NSCLC ) cell line primarily resistant to gefitinib, and to explore the role of reactive oxygen species ( ROS ) in the process.Methods: Two NSCLC cell lines with different sensitivities to gefitinib were selected, and the effects of gefitinib on cell growth, and induction of apoptosis induction by NSC-741909 were observed.The fluorescent probe 2',7'-dichlorofluorescein diacetate ( DCFH-DA ) was used to label the intracellular ROS and the intracellular fluorescence intensity was detected using flow cytometry ( FCM ).Jun N-terminal kinase ( JNK ) activation was detected using Western blot analysis.Results: NSC-741909 induced the apoptosis of the NSCLC cells primarily resistant to gefitinib.ROS was detected in this cell line.There was an increase in P-JNK expression, whereas the total JNK protein expression remained unchanged.The MKP1 expression was suppressed.Conclusion: NSC-741909, which suppresses mutant K-Ras expression, induced the apoptosis of NSCLC cells that are primarily resistant to gefitinib.This inhibition was mediated by increased ROS production and subsequent JNK activation.%目的:研究针对K-ras突变小分子NSC-741909是否可特异性杀伤吉非替尼原发耐药细胞,并对活性氧(reactive oxygen species,ROS)在其中的作用进行探讨.方法:选取H322(K-ras野生)及A549(K-ras突变)人源非小细胞肺癌细胞系,观察吉非替尼对不同细胞系生长的影响;NSC-741909作用于吉非替尼耐药细胞,观察细胞增殖与凋亡的变化;DCFH-DA荧光探针标记细胞内ROS,FCM检测细胞内ROS水平的变化,Western blot检测细胞内JNK通路蛋白变化.结果:K-ras突变型细胞对吉非替尼原发耐药,但NSC-741909可使这种原发耐药细胞出现凋亡;细胞内产生ROS,p-JNK表达增加而总JNK无改变,MKP1表达受抑制.结论:针对K-ras突变的小分子NSC-741909可使吉非替尼耐药细胞产生凋亡,NSC-741909通过使细胞内产生ROS持续激活JNK通路,是其导致吉非替尼耐药细胞凋亡的可能机制之一.