HSP90抑制剂17-AAG经下调HIF-1α抑制甲状腺未分化癌侵袭  

HSP90 Inhibitor,17-AAG,Inhibits Invasion of ATC by Down-Regulating HIF-1α

作　　者：吴昌昊 孙威[1] 秦元 董文武[1] 张文谦 邵亮[1] 张浩[1] Wu Changhao;Sun Wei;Qin Yuan;Dong Wenwu;Zhang Wenqian;Shao Liang;Zhang Hao(Department of Thyroid Surgery,The First Hospital of China Medical University,Shenyang 110001,Liaoning,China)

机构地区：[1]中国医科大学附属第一医院甲状腺外科

出　　处：《肿瘤预防与治疗》2019年第12期1081-1087,共7页Journal of Cancer Control And Treatment

基　　金：辽宁省“百千万人才工程”(编号:2014921033);沈阳市科学技术计划项目(编号:F16-205-1-41)~~

摘　　要：目的:研究热休克蛋白90(heat shock protein 90,HSP90)抑制剂17-丙烯氨基-去甲氧基-格尔德霉素(17-Nallylamino-17-demethoxygeldanamycin,17-AAG)对甲状腺未分化癌(anaplastic thyroid carcinoma,ATC)侵袭能力的影响,并探究HSP90与低氧诱导因子-1α(hypoxia-inducible factor 1-alpha,HIF-1α)及其下游侵袭相关蛋白在ATC细胞中作用的分子机制。方法:本试验通过细胞计数试剂盒-8(cell counting kit 8,CCK8)试验明确17-AAG对ATC细胞系8505c的药物半抑制浓度(half maximal inhibitory concentration,IC50),并确定进一步试验的药物浓度。通过Transwell试验检测17-AAG对ATC细胞侵袭能力的影响,并通过Western blot和蛋白质免疫共沉淀(co-immunoprecipitation,CO-IP)探索潜在的作用机制。结果:CCK8试验提示,经过不同浓度的17-AAG处理ATC细胞系后,17-AAG表现出对ATC的生长有抑制作用,作用12h、24h、48h的IC50分别为26μM、23μM、5μM,效果呈浓度依赖。根据CCK8试验结果,选取浓度0. 5μM和1μM作用24h用于后续试验。Transwell试验结果显示17-AAG对ATC细胞系8505c的侵袭能力具有抑制作用,并且具有浓度依赖性。Western blot结果表明17-AAG使ATC细胞系HIF-1α与侵袭相关蛋白基质金属蛋白酶2(matrix metallopeptidase 2,MMP2)和血管内皮生长因子(vascular endothelial growth factor,VEGF)的表达降低。CO-IP结果提示,HIF-1α与HSP90在ATC细胞中形成蛋白质复合物。进一步通过挽救试验证明,过表达HIF-1α后,MMP2和VEGF的表达水平升高,从而减弱了17-AAG对ATC细胞侵袭能力的抑制。结论:17-AAG可能通过调控HSP90与HIF-1α形成的复合物,使HIF-1α下调,导致侵袭相关蛋白MMP2和VEGF表达降低,从而抑制ATC细胞的侵袭能力。Objective: To probe into the effects of 17-N-allylamino-17-demethoxygeldanamycin(17-AAG),a potent heat shock protein 90(HSP90) inhibitor,on invasion of anaplastic thyroid carcinoma(ATC), and investigate how HSP90,hypoxia inducible factor-1α(HIF-1α) and their downstream targets affect ATC cells. Methods: The half maximal inhibitory concentration of 17-AAG on 8505 c,a ATC cell line,was determined by cell counting kit 8(CCK8) assay for use in further experiments. How 17-AAG inhibit the invasion ability of ATC cells was tested by transwell cell invasion chamber experiments,and its mechanisms were assessed by western blot and co-immunoprecipitation(CO-IP). Results: In CCK8 assay,17-AAG in different concentrations inhibited the growth of ATC cells,the effects were said to be dose-dependent. Based on these results,17-AAG in 0. 5μM and 1μM for 24 h were chosen for subsequent assay. The transwell test showed that 17-AAG inhibited the invasion of 8505 c in a dose-dependent manner.Western blot results indicated that 17-AAG also decreased HIF-1α expression and expression of matrix metalloproteinase-2(MMP2) and vascular endothelial growth factor(VEGF) which are invasion-related proteins. CO-IP analysis suggested that HIF-1α and HSP90 formed protein complexes in ATC cells. Rescue experiment further proved that MMP2 and VEGF expression increased after HIF-1α overexpression,which weakened the inhibition of 17-AAG on the invasion ability of ATC cells.Conclusion: 17-AAG may downregulate HIF-1α by regulating the complex formed by HSP90 and HIF-1,resulting in lower expression of MMP2 and VEGF to inhibit the invasion ability of ATC cells.

关 键 词：甲状腺未分化癌 热休克蛋白90 17-丙烯氨基-去甲氧基-格尔德霉素 低氧诱导因子-1α 侵袭 

分 类 号：R736.1[医药卫生—肿瘤] R966[医药卫生—临床医学]