INK4基因座中反义非编码RNA介导肺腺癌厄洛替尼抵抗的发生机制研究  

Occurrence mechanism of erlotinib resistance in lung adenocarcinoma mediated by antisense non-coding RNA in the INK4 locus

作　　者：姚彪 罗谣琴 姚海文 YAO Biao;LUO Yao-qin;YAO Hai-wen(Department of Oncology,Tongren People’s Hospital,Tongen 554309,Guizhou Province,China)

机构地区：[1]铜仁市人民医院肿瘤科,贵州铜仁554309

出　　处：《中国临床药理学杂志》2020年第19期3061-3065,共5页The Chinese Journal of Clinical Pharmacology

基　　金：铜仁市科技计划基金资助项目(2017-47-14号)。

摘　　要：目的探究长链非编码RNA(LncRNA)的INK4基因座中反义非编码RNA(ANRIL)对肺腺癌细胞厄洛替尼抵抗(ER)的影响及相关机制。方法将培养的人肺腺癌PC9细胞分为5组:对照组、厄洛替尼处理(ET)组、ER组、转染1组和转染2组。对照组用二甲基亚砜同步处理后转染GV248对照载体;ET组用0.2μmol·L^-1厄洛替尼处理24 h后转染GV248对照载体;ER组用梯度递增的厄洛替尼处理至6.4μmol·L^-1,维持2周后转染GV248对照载体;转染1组用梯度递增的厄洛替尼处理至GV248对照载体维持2周后,转染siANRIL载体;转染2组用转染1组细胞进一步转染过表达SLUG载体。用实时定量聚合酶链反应法检测各细胞系中LncRNA ANRIL、转录因子SLUG基因的表达,蛋白质印迹法检测各细胞系中各蛋白表达,CCK-8实验检测各细胞系厄洛替尼半抑制浓度(IC50)。结果对照组、ET组、ER组、转染1组和转染2组细胞中ANRIL基因表达分别为1.00±0.10,0.63±0.05,3.25±0.59,0.43±0.06和0.45±0.05;这5组的SLUG基因表达分别为1.00±0.08,0.55±0.07,3.83±0.24,3.36±0.10和8.18±0.03;ER组、转染1组和转染2组与对照组和ET组比较,ANRIL、SLUG基因表达的差异均有统计学意义(P<0.05,P<0.01)。对照组、ER组、转染1组和转染2组的厄洛替尼IC50分别为(0.26±0.04),(5.75±0.79),(3.09±0.52)和(7.57±0.94)μmol·L^-1;ER组、转染1组和转染2组与对照组相比,或转染1组与ER组比较,或转染2组与转染1组比较,差异均有统计学意义(均P<0.05)。蛋白结果的趋势与基因一致。结论肺腺癌ER与LncRNA ANRIL的重新表达密切相关,ANRIL可通过SLUG途径促进肺腺癌细胞上皮间充质转化(EMT)进程和ER。Objective To investigate the effect of long non-coding RNA(lncRNA)antisense non-coding RNA in the INK4 locus(ANRIL)on erlotinib resistance(ER)in lung adenocarcinoma cells and its mechanism.Methods PC9 cells were cultured and divided into 5 groups:control group,erlotinib treatment group(ET),ER group,transfection-1 group and transfection-2 group.The ER group was treated with erlotinib to 6.4μmol·L^-1 for 2 weeks and then transfected with GV248 control vector;control group was treated with DMSO and then transfected with GV248 control vector;ET group was treated with erlotinib to 0.2μmol·L^-1 for 24 h and then transfected with GV248 control vector;transfection-1 group was treated with erlotinib to 6.4μmol·L^-1 for 2 weeks and then transfected with si ANRIL vector;transfection 2 group further transfected SLUG vector in PC9 ER si ANRIL cells.The mRNA expressions of lncRNA ANRIL,transcription factors(SLUG)in each cell line were detected by real time quantitative-polymerase chain reaction.Western blot was used to detect the expression of the proteins in each cell line,and the semi-inhibitory concentration(IC50)of erlotinib in each cell line was detected by CCK-8.Results The expressions of ANRIL mRNA in control group,ET group,ER group,transfection-1 group and transfection-2 group were respectively1.00±0.10,0.63±0.05,3.25±0.59,0.43±0.06 and 0.45±0.05;the expressions of SLUG mRNA in the 5 groups were 1.00±0.08,0.55±0.07,3.83±0.24,3.36±0.10 and 8.18±0.03;compared between ER group,transfection-1 group,transfection-2 group and control group,ET group,the differences of ANRIL and SLUG genes expressions were significant(P<0.05,P<0.01).Erlotinib IC50 in control group,ER group,transfection-1 group and transfection-2 group were(0.26±0.04),(5.75±0.79),(3.09±0.52)and(7.57±0.94)μmol·L^-1;compared between ER group,transfection-1 group,transfection-2 group and control group,or between transfection-1 group and ER group,or between transfection-2 group and transfection-1 group,the differences were significant(all P<0.0

关 键 词：肺腺癌 厄洛替尼 药物抵抗 长链非编码RNA 上皮间充质转化 

分 类 号：R979.1[医药卫生—药品]