摘要
目的：构建靶向K-ras的siRNA，研究K—rassiRNA对K-ras基因突变型肺癌细胞A549及K-ras野生型小细胞肺癌细胞NCI—H446生长和迁移的抑制作用。方法：设计并人工合成4条K．rassiRNA（针对野生型K-ras基因的K—rassiRNA1-K—rassiRNA3；针对突变型K-ras基因的K—rassiRNA4），并分别转入A549和NCI—H446细胞。RT—PCR和Westernblotting检测不同K—rassiRNA对K—rasmRNA和蛋白表达的影响，MTr法检测不同K—rassiRNA对A549和NCI—H446细胞增殖的抑制作用。Transwell实验和Hoechst33258染色检测K—rassiRNA对细胞迁移和凋亡的影响。结果：靶向突变型K-ras的K．rassiR—NA4能特异性抑制A549细胞中K-ras的表达，但对N-ras和H-ras的表达没有影响。K—rassiRNA4抑制A549细胞的增殖，但不影响含野生型K-ras基因的NCI—H446细胞的增殖。K—rassiRNA4还能诱导A549细胞凋亡、抑制A549细胞迁移。结论：针对突变型K-ras基因的siRNA可特异性抑制K-ras突变型肺癌细胞的增殖和迁移，并诱导该细胞凋亡，K—rassiRNA可望用于K-ras突变型肿瘤特别是肺癌的个体化治疗。
Objective：To construct K-ras-targeted siRNAs （K-ras siRNA） and to investigate the inhibitory effects of K-ras siRNAs on the growth and migration of lung cancer A549 cells （ containing mutant K-ras gene） and NCI-H446 cells （ containing wild-type K-ras gene）. Methods ： Four K-ras siRNAs （ K-ras siRNA1 - K-ras siRNA3 targeting wild-type K-ras and K-ras siRNA4 targeting mutant K-ras） were designed and artificially synthesized; they were used to transfect A549 cells and NCI-H446 cells. The expressions of Ras mRNA and protein were examined by RT-PCR and Western blotting. The inhibitory effects of K-ras siRNAs on the proliferations of A549 and NCI-H446 cells were determined by MTT assay. The effects of K-ras siRNAs on the cell migration and apoptosis were observed by Transwell assay and Hoechst 33258 staining, respectively. Results： Mutant K-ras-targeted siRNA （ K-ras siRNA4） specifically inhibited the K-ras expression but had no influence on H-ras and N-ras expression in A.549 cells. K-ras siRNA4 inhibited the proliferation of A549 cells but did not inhibit that of NCI-H446 cells, which contained wild type K-ras gene. K-ras siRNA4 also induced apoptosis and inhibited migration of A549 cells. Conclusion： Mutant K-ras-targeted siRNA4 can inhibit the proliferation, migration and induce apoptosis of A549 cells. It may be a potential and personalized drug for the treatment against lung cancer containing mutant K-ras gene.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2009年第6期564-569,共6页
Chinese Journal of Cancer Biotherapy
基金
国家高技术研究发展计划(863计划)课题资助项目(No.2008AA02Z135)
福建省发改委课题资助项目(2008第3批26号)~~
