乌苯美司抑制肿瘤细胞侵袭与诱导凋亡的研究  

Inhibition of tumor cell invasion and induction of apoptosis by ubenimex

作　　者：郑艳波[1] 弓建华[1] 李毅[1] 甄永苏[1] 

机构地区：[1]中国医学科学院,北京协和医学院医药生物技术研究所,北京100050

出　　处：《药学学报》2012年第12期1593-1598,共6页Acta Pharmaceutica Sinica

基　　金：国家"重大新药创制"科技重大专项(2010ZX09401-407)

摘　　要：探讨乌苯美司对肿瘤细胞侵袭及凋亡的影响及其剂量关系,探讨其作用机制。采用免疫荧光法检测CD13/APN在HT-1080细胞中的表达;MTT法检测乌苯美司对HT-1080细胞增殖的影响;Annexin V-EGFP/PI双染法检测乌苯美司对HT-1080细胞凋亡的影响;流式细胞术检测乌苯美司对HT-1080细胞周期的影响;运用Ala-pNA为底物,检测乌苯美司对细胞表面氨肽酶活性的抑制作用;Transwell法检测乌苯美司对HT-1080细胞侵袭及运动能力的影响;明胶酶谱法检测乌苯美司对基质金属蛋白酶活性的影响;Western blotting法检测CD13的表达变化。研究结果显示,高浓度的乌苯美司可抑制HT-1080细胞的增殖(IC50:3.8 mg.mL-1),诱导HT-1080细胞的凋亡,将细胞周期阻滞于G1期;在低浓度时乌苯美司即可显著抑制HT-1080细胞的氨肽酶活性(IC50:8.3μg.mL-1),抑制细胞的侵袭能力,但对细胞的运动能力及增殖无影响;乌苯美司对CD13的表达及基质金属蛋白酶的活性无明显影响。以上研究结果表明,乌苯美司在低浓度时可通过直接抑制细胞表面的氨肽酶活性而抑制肿瘤细胞的侵袭;在高浓度时可通过非CD13依赖的途径抑制肿瘤细胞的增殖,诱导细胞凋亡。This study is to investigate the effects of ubenimex on tumor cell invasion and apoptosis, dose relationship and mechanism. Immunofluorescence staining was performed to detect the expression of CD13 in HT-1080 cells. MTT assay was used to analyze the effect of ubenimex on cell proliferation. Annexin V-EGFP/PI was used to detect apoptotic cells by flow cytometry. Cell cycle was analyzed using flow cytometry. Ala-pNA was used as substrate to evaluate the effect of ubenimex on the aminopeptidase activity. Transwell assay was used to analyze the effect of ubenimex on cell invasion and migration ability. Western blotting was used to detect the expression level of CD13. MMP activity was analyzed using gelatin zymography. The results showed that ubenimex at high concentration inhibited the proliferation of HT-1080 cells （IC50： 3.8 mg·mL-1）, and induced cell apoptosis. Cell cycle was blocked at G1 phase. Ubenimex at low concentration inhibited the aminopeptidase activity of HT-1080 cells （IC50：8.3 μg·mL-1） and inhibited cell invasion, but it had no effects on the cell migration and proliferation. Ubenimex had no effects on CD13 expression and MMP activity. In conclusion, ubenimex at low concentration can inhibit the invasion ability of tumor cells by directly inhibiting the aminopeptidase activity; ubenimex at high concentration can inhibit the proliferation of tumor cells and induce cell apoptosis by a CD 13-independent pathway.

关 键 词：乌苯美司 CD13 APN 侵袭 细胞凋亡 HT-1080细胞 

分 类 号：R963[医药卫生—微生物与生化药学]