雷公藤红素与凋亡蛋白突变体通过强化Nur77诱发凋亡通路发挥协同抗肿瘤作用  

Celastrol and Apoptin Mutant Exert Synergistic Anti-tumor Effects by Enhancing Nur77-induced Apoptosis Pathway

作　　者：尹晓梦 曹雪玮 王富军 赵健[1] 张惠展[1] YIN Xiao-meng;CAO Xue-wei;WANG Fu-jun;ZHAO Jian;ZHANG Hui-zhan(State Key Laboratory of Bioreactor Engineering,East China University of Science and Technology,Shanghai 200237;Zhejiang Fonow Medicine Co.Ltd,Dongyang 322100;Institute of Chinese Materia Medica,Shanghai University of Traditional Chinese Medicine,Shanghai 201203)

机构地区：[1]华东理工大学生物反应器工程重点实验室,上海200237 [2]浙江孚诺医药股份有限公司,东阳322100 [3]上海中医药大学中药研究所,上海201203

出　　处：《生物技术通报》2020年第7期119-129,共11页Biotechnology Bulletin

基　　金：国家自然科学基金项目(81571795)。

摘　　要：雷公藤红素(Celastrol)和凋亡蛋白(Apoptin)均可通过Nur77信号通路介导肿瘤细胞凋亡,Celastrol与含有商陆皂苷甲(EsA)的Apoptin突变体tApoptin(简称为tApoptinE)联用是否具有高效的抗肿瘤活性及其协同作用的分子机制值得研究。通过MTT法检测药物联用后对肿瘤细胞增殖的抑制活性,流式细胞术和免疫印迹(Western blot)技术分析药物联用的分子机制。结果显示,Celastrol浓度≥300 nmol/L时可以显著提高tApoptinE对肝癌细胞SMMC-7721生长的抑制活性;上述联用条件下两种药物联合作用指数(Combined index)均小于1,其中,Celastrol(600 nmol/L)与tApoptinE联用的协同效果最显著;流式凋亡分析揭示联合用药强化了对细胞的凋亡效应;Western blot分析表明两种药物协同作用于共同的Nur77通路,促进更多的Nur77被磷酸化,磷酸化的Nur77显著调控Caspase和Bcl-2家族蛋白的表达,强化了细胞凋亡途径。上述两种药物联合使用后通过强化Nur77诱发的细胞凋亡通路,进一步诱导肿瘤细胞凋亡,从而发挥协同抗肿瘤作用。Both celastrol and apoptin can exert anti-tumor effect through the Nur77 pathway.We named a truncated variant of apoptin as t Apoptin.tApoptin co-administered with esculentoside A(EsA)is known simply as tApoptinE.Combination of cleastrol with t ApoptinE is applied to enhance the inhibiting tumor cell growth activity of these two drugs,and the molecular mechanism for these two drugs synergistically exerting anti-tumor ability is studied.The cytotoxicity of the combination of celastrol with tApoptinE on tumor cells was detected by MTT assay,and the molecular mechanism of cell apoptosis was observed by flow cytometry and Western blot analysis.The results showed that celastrol whose concentration is above 300 nmol/L could effectively increase the anti-tumor activity of tApoptinE in cell line SMMC-7721 in comparison with the free tApoptinE.Under the combined conditions above,the combined index values of all combined groups were<1,which showed that celastrol and tApoptinE synergistically inhibit the growth of tumor cells in vitro in this case.In addition,600 nmol/L celastrol in combination with t ApoptinE had the best synergistic effect.Flow cytometry analysis revealed that the synergistic effect was due to cell apoptosis enhancement.Western blot analysis showed that after combined administration,celastrol and tApoptinE synergistically targeted their common Nur77 pathway which will have Nur77 phosphorylated.And the phosphorylated Nur77 can significantly regulate the expression of Caspase and Bcl-2 family proteins,hence enhancing the induction of apoptosis.Therefore,the combination of celastrol and tApoptinE can further induce tumor cell apoptosis by enhancing the apoptosis pathway induced by Nur77,so as to exert synergistic anti-tumor effect.

关 键 词：雷公藤红素 凋亡蛋白 Nur77 协同抗肿瘤 细胞凋亡 

分 类 号：R285.5[医药卫生—中药学]