ABT-737增强Mcl-1小分子抑制剂UMI-77诱导的胃癌细胞凋亡

ABT-737 enhances apoptosis of gastric cancer cells induced by Mcl-1 small-molecule inhibitor UMI-77

目的探讨在对Mcl-1小分子抑制剂UMI-77不敏感的胃癌细胞中,将Bcl-2/Bcl-xL抑制剂ABT-737与UMI-77联用,增加胃癌细胞凋亡的分子机制。方法 MTS法检测不同浓度UMI-77处理胃癌细胞MGC-803和HGC-27的细胞存活率。在对UMI-77抵抗的HGC-27细胞中,将UMI-77和ABT-737联用,MTS法检测细胞存活情况。Annexin V-FITC/PI染色,流式细胞术分析细胞凋亡情况;JC-1染色,流式细胞术分析线粒体膜电位的变化;Western blot检测caspase-9、caspase-3、PARP-1的裂解,以及Bcl-2家族和IAP家族的表达水平。结果与MGC-803相比,HGC-27细胞对UMI-77较为抵抗,同时它对ABT-737也不太敏感,但是当ABT-737与UMI-77联用后,能明显增加细胞死亡,表现为Annexin V(+)、线粒体膜电位下降、caspases裂解,说明两者联用是通过线粒体途径诱导细胞凋亡。在此过程中,XIAP、cIAP1和cIAP2的表达水平下降,NOXA、Bcl-2升高及PUMA、Mcl-1降低可能参与了增敏作用。结论在对UMI-77不敏感的胃癌细胞中,将UMI-77和ABT-737联用能明显增加胃癌细胞凋亡。

Aim To investigate the promoting effect of Bcl-2/Bcl-xL inhibitor ABT-737 on apoptosis of gastric cancer cells induced by small molecule Mcl-1 inhibitor UMI-77,and to explore its possible mechanism.Methods The response of gastric cancer MGC-803 and HGC-27 cells to different concentrations of UMI-77 was detected by MTS assay.In the UMI-77-resistant cell lines,the effect of treatment with UMI-77/ABT-737 alone or in combination on cell viability was detected by MTS assay.The apoptotic rate and the changes of the mitochondrial membrane potential were analyzed by flow cytometry.The cleavage of caspase-9,caspase-3 and PARP-1,as well as the expression level of Bcl-2 family members and IAP proteins,were determined by Western blot.Results Compared with MGC-803 cells,HGC-27 cells were resistant to UMI-77.Treatment with ABT-737 alone in HGC-27 cells also induced minimal level of cell death.While treatment with both agents induced much greater decreased cell viability.All the dead cells were positive for Annexin V and mitochondrial membrane potential collapsed.Caspase-9,caspase-3 and its substrate PARP-1 were cleaved.All of these proved that the sensitization effect was achieved by activating the mitochondrial apoptotic pathway.Protein levels of XIAP,cIAP1and cIAP2 decreased after treatment with UMI-77 plus ABT-737.It also resulted in the increase of NOXA and Bcl-2 along with the decline of PUMA and Mcl-1.Conclusions The combination of UMI-77 and ABT-737 could significantly increase the sensitivity of gastric cancer cells to the Mcl-1 small molecule inhibitor UMI-77.