查尔酮衍生物H175促结肠癌细胞凋亡及其机制研究

Effects and mechanisms of a novel chalcone analogue-H175 on inducing colon cancer cells＇ apoptosis

目的 通过对经初步筛选证明具有生物活性的查尔酮类似物H175可能涉及的信号通道进行检测,进一步明确其抗肿瘤活性及作用机制.方法 对于以查尔酮为先导物设计合成类似物,用MTT法检测其对小鼠结肠癌细胞（CT26）的杀伤作用,筛选出具有较好生物活性的化合物H175.通过流式细胞技术检测H175诱导CT26细胞凋亡和抑制细胞周期的情况.Western blot法检测CT26细胞中H175对于活性转录因子4（activating transcriptional factor 4,ATF-4）及DNA损伤诱导基因（DNA damage-inducible gene 153,CHOP）表达的影响.利用siRNA技术敲除CHOP后,用免疫荧光和流式细胞技术检测H175对于细胞凋亡的影响.结果 通过对113个自行设计合成查尔酮类似物进行筛选,发现H175对CT26细胞具有较强的增殖抑制作用（IC50=1.9 μmol/L）.流式细胞技术检测发现H175通过诱导CT26细胞发生凋亡并抑制细胞周期发挥其生物活性.Western blot检测发现H175剂量依赖性激活内质网应激信号通路中的关键蛋白CHOP,并通过siRNA基因沉默技术进一步确证H175是通过刺激CHOP的过表达发挥促进细胞凋亡的作用.结论 查尔酮类似物H175通过激活内质网应激信号通道中的关键蛋白CHOP发挥对结肠癌CT26细胞的增殖抑制作用.

Objective To investigate the anti-tumor activity of a novel chalcone analogue-H175.Methods We synthesized chalcone analogues and further tested the cytotoxicity on colon cancer cell line （CT26） through MTT method,and identified H175 with powerful biological activity.Flow cytometry was used to assay the ability of H175 to induce CT26＇s apoptosis and inhibit its cell cycle.Western blot was used to detect the effect of H175 on ATF4 and CHOP expression in CT26.After knockdown CHOP by siRNA technology,further investigate the influence of H175 on cell＇s apoptosis through immunofluorescence （IF） and FCM.Results H175 inhibits the proliferation of CT26 （ICs0 =1.9 μmol/L）,by inducing CT26＇s apoptosis inhibiting its cell cycle,on a dose-dependent manner of key protein CHOP of endocytoplasmic reticulum stress （ERS） signal path.siRNA technology confirmed H175 induce apoptosis by stimulating the CHOP over-expression.Conclusions Novel chalcone analogue-H175 show inhibitory effects on the proliferation of CT26 by activating the ERS signal path＇s key protein CHOP.