定靶突变温敏酵母模型在Pin1抑制剂筛选中的应用  

Application of temperature sensitive yeast models with definite target in the screening of potential human Pin1 inhibitors

作　　者：张晶[1] 韩小敏[2] 胡文辉[3] 郭宗儒[4] 贺晓波[1] 司书毅[1] 

机构地区：[1]中国医学科学院医药生物技术研究所,北京100050 [2]国家食品安全风险评估中心卫生部食品安全风险评估重点实验室,北京100021 [3]中国科学院广州生物医药与健康研究院,广东广州510530 [4]中国医学科学院药物研究所,北京100050

出　　处：《药学学报》2014年第6期854-860,共7页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金海外及港澳学者资助研究基金资助项目(81128014)

摘　　要：利用定靶突变温敏酵母模型寻找Pin1的新型抑制剂,并探讨阳性化合物的体内外抗肿瘤活性。采用MTT法、流式细胞仪法、免疫印迹法、划痕实验和虚拟对接检测阳性化合物的体外活性。利用移植瘤裸鼠模型检测阳性化合物的体内活性。结果发现,酵母模型初筛得到的阳性化合物8-11在体外,对Pin1的IC50值为(10.40±1.68)μmol·L-1,能抑制多种肿瘤细胞的增殖,诱导HeLa细胞G1期阻滞和凋亡,降低Cyclin D1的蛋白水平表达,降低肿瘤细胞迁移能力,虚拟对接中与Pin1的活性结构域呈现出较高的亲和性;在体内,8-11抑制裸鼠移植瘤生长。本研究首次证实8-11可通过抑制Pin1发挥抗肿瘤作用。This study is to explore new lead compounds by inhibition of Pinl for anticancer therapy using temperature sensitive mutants. As Pinl is conserved from yeast to human, we established a high-throughput screening method for Pinl inhibitors, which employed yeast assay. This method led to the identification of one potent hits, 8-11. In vitro, 8-11 inhibited purified Pin1 enzyme activity with IC50 of （10.40±1.68）μmol·L-1, induced G1 phase arrest and apoptosis, showed inhibitory effects on a series of cancer cell proliferation, reduced Cyclin D1 expression, was defined as reciprocally matched for protein-ligand complex in virtual docking analysis and reduced cell migration ability. In vivo, we could observe reduction of tumor volume after treatment with 8-11 in xenograft mice compared with vehicle DMSO treatment. Altogether, these results provide for the first time the involvement of 8-11 in the anticancer activity against Pinl.

关 键 词：PIN1 抗肿瘤 周期阻滞 细胞凋亡 裸鼠 

分 类 号：R963[医药卫生—微生物与生化药学]