MLN4924与5-氟尿嘧啶协同杀伤人结肠癌HCT116细胞的研究

Synergistic anticancer effects of MLN4924 and 5-FU on human colon cancer cell line HCT116

目的探讨MLN4924与5-氟尿嘧啶(5-FU)单用及联用对人结肠癌HCT116细胞活力的影响及其发挥作用的可能途径。方法细胞活力实验检测两药单用及联用对HCT116细胞活力的影响;应用中效原理计算两药联合应用的联合指数,判断药物联用的效果;将HCT116细胞分为对照组、MLN4924组、5-FU组及联用组,药物作用后,流式AnnexinⅤ-FITC/PI双标法检测各组细胞凋亡情况;蛋白质印迹实验检测各组细胞Cleaved PARP、Cleavedcaspase3的表达水平。结果随着MLN4924和5-FU浓度的增加,HCT116细胞活力逐渐降低; MLN4924与5-FU联用时,抑制效果明显强于单药作用(P <0. 001),各联合用药组的联合指数均<1,提示两药联用起协同效果;与对照组相比,单用及联用均可显著促进HCT116细胞的凋亡(P <0. 001),且联用效果显著优于单用(P <0. 01);与流式AnnexinⅤ-FITC/PI双标法检测细胞凋亡的结果相一致,联用组Cleaved PARP、Cleaved caspase3蛋白表达较对照组及单用组明显升高。结论 MLN4924与5-FU通过促进细胞凋亡而发挥协同抑制人结肠癌HCT116细胞活力的作用。

Objective To explore the effects of a combination of MLN4924 and 5-fluorouracil( 5-FU) on the cell viability of human colon cancer cell HCT116 and the relevant mechanisms involved. Methods Cytotoxicity of the agents used singly and in combination was evaluated by cell viability assay. The joint effect of the two drugs was further analyzed using the median-effect method of Chou and Talalay. HCT116 cells were divided into control group,MLN4924 group,5-FU group,and combination group. Cell apoptosis was quantified by flow-cytometry after double-staining of the cells with AnnexinⅤ-FITC/PI. The expression levels of Cleaved PARP and Cleaved caspase3 were evaluated by Western blotting.Results Cell viability assays revealed that MLN4924 or 5-FU alone inhibited cell viability in a dose-dependent manner.When MLN4924 was combined with 5-FU,the inhibitory effect was significantly stronger than that of the single drug( P <0. 001). The combination index( CI) of each combination group was < 1,indicating synergism regarding HCT116 cell viability. The levels of apoptosis induction in HCT116 cells treated with MLN4924 or 5-FU were significantly increased compared with the untreated control cells( P < 0. 001),and the combined drug effect was significantly better than that of the drug alone( P < 0. 01). The results of Western blotting analyses revealed an increase in Cleaved PARP and Cleaved caspase3 protein expressions in the combination group compared with control group and single drug group. Conclusion The combination of MLN4924 and 5-fluorouracil can significantly inhibit the cell viability and enhance the apoptosis of human colon cancer cell HCT116.