Brefeldin A通过内质网应激途径协同增强顺铂抗肺癌细胞GLC-82作用  

The Synergistic Anti-cancer Effect of BFA and CDDP on Human Lung Cancer GLC-82 Cells

作　　者：吴明松[1,2] 郑翔[1] 耿娜娜[1] 张志敏[1] 赵彦禹[1] 王哲[1] 李学英[1] 

机构地区：[1]遵义医学院医学遗传学教研室,贵州遵义563000 [2]贵州省高等学校口腔疾病研究特色重点实验室,医学与生物学研究中心,贵州遵义563000

出　　处：《中国生物化学与分子生物学报》2016年第5期587-593,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：贵州省科技厅资助项目(黔科合J字[2013]2319号;黔科合LH字[2014]7548号);贵州省教育厅特色药用资源研发创新团队(黔科合人才团队字[2013]15);遵义医学院博士启动基金(F-655);遵义医学院遗传学扶持学科经费(2012-2016)资助

摘　　要：已有研究证明,内质网应激(endoplasmic reticulum stress,ERS)启动的未折叠蛋白质反应(unfolded protein response,UPR)信号途径有助细胞存活;然而,长时间剧烈ERS可诱导细胞凋亡,因此,ERS是肿瘤治疗的新靶点。抗癌药物具有严重的毒副作用,而药物的协同作用是减少抗癌药物使用剂量、减少毒副作用的重要手段之一。本研究证明布雷菲德菌素A(brefeldin A,BFA)与顺铂(cis-dichlorodiamine platinum,CDDP)的协同抗肺癌细胞的作用。MTT试验显示,单独应用BFA和顺铂时,它们抑制肺癌细胞GLC-82生长的半数有效浓度(IC50)分别是100 ng/m L和4μg/m L;而采用半量的BFA和CDDP联合处理GLC-82细胞24 h后,其抑制生长作用进一步加强;等效线图解法及联合作用指数分析进一步证明二者具有协同作用。两药的这种协同作用进一步被形态学检查证明——与单独用药比较,除了细胞皱缩,出现了更多的染色质固缩,细胞质及细胞核裂解碎片,乃至形成凋亡小体,提示细胞凋亡的发生。实时定量PCR及蛋白质印迹证明,与单独用药比较,联合用药导致内质网应激标志分子GRP78、CHOP表达水平明显增加,提示有内质网应激通路激活。上述结果证明,BFA与CDDP联合用药可增强对肺癌细胞GLC-82的生长抑制作用,其协同机制可能与内质网应激通路的激活有关。我们的结果支持"ERS是肿瘤治疗新靶点"学说,对肺癌的新化疗方案有一定的指导意义。Endoplasmic reticulum stress （ERS）, which triggers unfolded protein response （UPR） to support cell survival, is indicated as a new target for the development of cancer therapy, despite long- term ERS initiates apoptosis. Combination of anti-cancer drugs can reduce side effects and synergistically enhance drug effects. In this study, we investigated the synergistic anti-cancer effect of brefeldin A （BFA） and cis-dichlorodiamine platinum （CDDP） in human lung cancer cells. MTT assays revealed that BFA or CDDP alone inhibited the proliferation of GLC-82 human lung cancer cells with an IC50 of 100 ng/mL or 4 μg/mL, respectively. At half doses of BFA and CDDP in combination showed more inhibitory effect at 24 hours with an observed synergism shown in the isobologram of cell proliferation index. With combined exposure of drugs, more shrunk cells or abnormal cells with chromatin condensation were observed, together with increased numbers of apoptotic bodies with nucleus and cytoplasm fragmentation. Real-time quantitative PCR （RT-qPCR） and Western blotting results showed that the levels of GRP78 and CHOP, the endoplasmic reticulum stress markers, were markedly increased, as compared to single drug exposures. These data suggested that combination of BFA and CDDP could enhance the inhibitory effect on human lung cancer cells, where the ERS pathway might be involved.

关 键 词：布雷菲德菌素A 顺铂 人肺癌细胞系GLC-82 内质网应激 协同作用 细胞凋亡 

分 类 号：R73-34[医药卫生—肿瘤]