一种新的吲哚咔唑类化合物(ZWM233)的体外抗肿瘤作用及机制探讨

目的探讨ZWM233体外对多种肿瘤细胞株的增殖抑制及对K562细胞的凋亡诱导作用。方法采用MTT法检测ZWM233对K562、HL-60、A549、HeLa及HCT-116等10种肿瘤细胞株的增殖抑制作用;流式细胞仪检测对K562细胞周期的影响;AnnexinV/PI双染法考察对K562细胞的凋亡诱导作用;Western blot检测K562细胞中凋亡相关蛋白bax、bcl-2、Caspase-3、Caspase-9及PARP的变化,同时检测蛋白激酶C(PKC)各亚型、GSK-3β、ERK1/2、JNK及其磷酸化水平的变化。结果 ZWM233体外能有效抑制多种肿瘤细胞株的增殖,其中对K562细胞作用明显,IC50为2.81μmol.L-1;流式细胞仪检测K562细胞被明显阻滞在G2/M期,并诱导其凋亡;Western blot结果显示ZWM233作用K562细胞24 h后,可明显下调抗凋亡蛋白bcl-2的表达,引起Caspase-9、Caspase-3及下游底物PARP的剪切,诱导K562细胞凋亡。Western blot检测结果进一步显示药物作用24 h后PKCδ的表达有所降低,其下游分子p-GSK-3β及p-ERK的表达水平降低,而p-JNK蛋白的表达水平升高。结论ZWM233体外能有效抑制K562细胞生长,并通过激活线粒体途径诱导其凋亡,其机制可能是通过下调PKCδ,进一步抑制p-GSK-3β及p-ERK1/2的表达来发挥凋亡诱导作用。

Anticancer effects of ZWM233 in vitro and its mechanisms

Aim To investigate the antiproliferation effects of ZWM233 in a variety of cancer cell lines and the apoptosis mechanisms of K562 cells in vitro.Methods Inhibitory effects of ZWM233 on proliferation of a variety of cancer cell lines were analyzed by MTT method.The changes of K562 cell cycle was detected by flow cytometry.Apoptosis was determined by AnnexinV / PI dual staining.The expressions of bax,bcl2,Caspase-3,Caspase-9,PARP,ERK,p-ERK,JNK,pJNK,GSK-3β,p-GSK-3β and PKC isotypes were detected by Western blotting assay.Results ZWM233 potentially inhibited proliferation of a variety of cancer cell lines,among which K562 cells were the most significant(IC50 was 2.81 μmol.L-1).FCM analysis revealed that ZWM233 induced k562 cells toward apoptosis in a dose-dependent manner and the cell cycle was blocked at G2 / M phase.After treated with ZWM233 for 24 h,ZWM233 could down-regulate bcl2 and activate Caspase-9 and Caspase-3 to cleave the downstream substrate PARP to induce K562 cells apoptosis.Western blotting also showed that ZWM233 could down-regulate PKCδ,p-ERK1 /2 and p-GSK-3β and reversely regulate p-JNK in a dose-dependent manner.Conclusion ZWM233 potentially inhibits proliferation in K562 leukemia cells in vitro and induces apoptosis via activating the mitochondrial pathways.Its mechanism is involved in the down-regulation of pkcδ as well as its downstream p-GSK-3β and p-ERK1 /2.