维罗非尼联合爱必妥对结直肠癌细胞株的作用研究  

Effects of Cetuximab Combined with Vemurafenib on Colorectal Cancer Cell Lines

作　　者：马瑞[1] 徐玲[1] 曲秀娟[1] 车晓芳[1] 郭天舒[1] 刘云鹏[1] 

机构地区：[1]中国医科大学附属第一医院,辽宁沈阳110001

出　　处：《微生物学杂志》2017年第2期83-88,共6页Journal of Microbiology

基　　金：国家自然科学基金项目(81673025);辽宁省自然科学基金项目(2014021069)

摘　　要：探讨BRAF抑制剂维罗非尼联合EGFR单克隆抗体西妥昔单抗对BRAF突变型和野生型结直肠癌细胞株的作用。应用MTT法检测,将不同浓度西妥昔单抗、维罗非尼单药以及两药联合分别作用于BRAF突变型结直肠癌细胞株RKO、HT-29和BRAF野生型结直肠癌细胞株DIFI、CACO-2后,观察各细胞株增殖变化;通过集落形成实验,经过长时间培养细胞验证不同处理因素对细胞增殖的抑制作用;Western blot实验检测EGFR下游通路靶蛋白的活性在两药联合后是否出现明显下调。结果显示,单药治疗,西妥昔单抗能一定程度抑制BRAF野生型结直肠癌细胞株增殖,抑制率达26%~40%,而BRAF突变组两种细胞株对西妥昔单抗均耐药。维罗非尼对不同基因状态细胞的增殖均有一定的抑制作用,IC50均小于7μmol/mL,其中BRAF突变细胞更为敏感。对比单药西妥昔单抗,联合维罗非尼后更加明显地抑制了细胞增殖,4种细胞存活率均明显降低(P<0.05)。长时间集落形成实验证实两药联合形成集落的面积和数量均明显减少。Western Blot实验显示对比单药,两药联合明显下调了p-EGFR、BRAF、p-ERK的表达。结果显示,BRAF抑制剂维罗非尼无论在BRAF突变型还是野生型结直肠癌细胞株中,均能增强西妥昔单抗作用的敏感性。The effects of cetuximabin combined with vemurafenib on wild type and mutant BRAF colorectal cancer cell lines were investigated. The cell proliferation was measured using MTT assay. The inhibitory effects of single and combination on cell proliferation were detected. Colony formation experiment： The inhibitory effect of different treat- ment factors on cell proliferation was eonfirmed by long-term culture of cells. The expression of EGFR and phosphor- EGFR and downstream pathway protein were determined by Western blotting. The results showed that for single drug therapy, BRAF wild group is relatively sensitive to cetuximab treatment. The inhibition of 100 μg/mL is 26% -40% in 72 h, whereas the BRAF mutation group of two cell lines was resistant to eetuximab. Both BRAF wild or mutant group cell lines were sensitive to vemurafenib, IC50 was less than 7 μmol/mL. The cell proliferative inhibition test showed the effect of two drug combinations were significantly lower than single drug eetuximab （ P 〈 0.05 ）. Long time colony forming experiments confirmed that the combination in the area and the number of colony formation was significantly reduced. Western Blot experiment indicated that compared with single drug, two drug combination was significantly reduced p-EGFR, BRAF, p-ERK expression. The results showed that BRAF inhibitor vemurafenib can increase the sensitivity of cetuximab in either BRAF wild-type or mutant colorectal cancer cell lines.

关 键 词：西妥昔单抗 维罗非尼 EGFR BRAF 结直肠癌 

分 类 号：Q939.9[生物学—微生物学] R73[医药卫生—肿瘤]