mTOR信号通路在克唑替尼诱导的EML4-ALK融合基因阳性肺癌细胞株H2228凋亡中的作用  

Role of mTOR signaling pathway in crizotinib-induced apoptosis of EML4-ALK fusion gene-positive lung adenocarcinoma cell line H2228

作　　者：戴辉[1] 宋向群[2] 潘星辰[1] 彭海燕[1] 韦江[1] 周韶璋[2] 

机构地区：[1]广西医科大学研究生院,广西南宁530021 [2]广西医科大学附属肿瘤医院化疗二科,广西南宁530021

出　　处：《中国病理生理杂志》2014年第6期1103-1109,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81060188);广西科技厅项目(No.科201017)

摘　　要：目的:探讨以磷脂酰肌醇3-激酶相关激酶蛋白家族成员哺乳动物雷帕霉素靶蛋白(mTOR)为中心的信号通路在克唑替尼(crizotinib)诱导的棘皮动物微管结合蛋白样蛋白4-间变性淋巴瘤激酶(EML4-ALK)融合基因阳性的非小细胞肺癌细胞株H2228凋亡中的作用。方法:根据不同的实验目的处理H2228细胞后,荧光定量PCR检测基因状态,MTT法检测细胞抑制率;流式细胞术检测细胞凋亡和细胞周期;Western blotting检测细胞mTOR信号通路中关键蛋白的表达及活化水平。结果:Crizotinib对H2228细胞有促凋亡作用,呈时间和剂量依赖性,且能使H2228细胞阻滞在G1期。在使用crizotinib处理的凋亡细胞株中发现mTOR活化水平降低,mTOR上、下游关键蛋白的活化水平都呈下降趋势,肺癌细胞株H2228中特殊表达的融合蛋白EML4-ALK变异体3表达量未受影响,但其活化形式p-ALK明显受到抑制。结论:初步证实mTOR信号通路在crizotinib诱导含有EML4-ALK融合基因的肺癌细胞H2228凋亡中有一定作用,为crizotinib的作用机制提供了依据。AIM：To investigate the mammalian target of rapamycin （ mTOR） signaling pathway as the center playing a role in the crizotinib-induced apoptosis of non-small cell lung cancer （NSCLC） cell line H2228, which represents positive echinoderm microtubule-associated protein-like 4 （EML4）-anaplastic lymphoma kinase （ALK） fusion gene. METHODS：H2228 cells were processed according to different purposes .Fluorescence quantitative PCR is used to observe the gene states .MTT assay is used to detect the cell inhibition rates .The cell apoptosis and cell cycle were analyzed by flow cytometry .The expression and activation levels of the key proteins in the mTOR signaling pathway were determined by Western blotting .RESULTS：Crizotinib promoted the apoptosis of H 2228 cells in a time-and dose-dependent manner . Crizotinib blocked the H2228 cells staying at the G1 phase.In apoptotic H2228 cells processed with crizotinib, the activation level of mTOR was decreased , and the activation levels of the key proteins in upstream and downstream of mTOR path -way were both declined .The expression level of the fusion protein EML 4-ALK variant 3 was not affected , but its active form of p-ALK was significantly suppressed .CONCLUSION：mTOR signaling pathway has a certain relationship with the crizotinibinduced apoptosis of lung cancer cell H 2228, which represents positive EML4-ALK fusion gene.

关 键 词：EML4-ALK融合基因 H2228细胞 克唑替尼 细胞凋亡 哺乳动物雷帕霉素靶蛋白 

分 类 号：R73-3[医药卫生—肿瘤]