17-Allylamino-17-demethoxygeldanamycin induces downregulation of critical Hsp90 protein clients and results in cell cycle arrest and apoptosis of human urinary bladder cancer cells
17-Allylamino-17-demethoxygeldanamycin induces downregulation of critical Hsp90 protein clients and results in cell cycle arrest and apoptosis of human urinary bladder cancer cells
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DOI：
10.1186/1471-2407-10-481
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发表时间：
2010-09-09
期刊：
影响因子：
3.8
通讯作者：
Voutsinas, Gerassimos E.
中科院分区：
文献类型：
--
作者：
Karkoulis, Panagiotis K.;Stravopodis, Dimitrios J.;Voutsinas, Gerassimos E.
背景资料：17-烯丙基氨基-17-去甲氧基格尔德霉素（17-AAG）是一种苯醌安莎霉素抗生素，特异性靶向热休克蛋白90（Hsp 90）并干扰其作为分子伴侣的功能，该分子伴侣维持参与细胞信号传导的各种蛋白质客户的结构和功能完整性。在这项研究中，我们研究了17-AAG对Hsp 90依赖性信号通路的调节作用，该信号通路直接涉及人膀胱癌细胞系的细胞周期进展、存活和运动。我们在RT 4中使用了基于MTT的测定、FACS分析、Western印迹、半定量RT-PCR、免疫细胞化学和划痕试验，RT 112和T24人膀胱癌细胞系。结果：我们已经证明，在17-AAG治疗，膀胱癌细胞被逮捕在G1期的细胞周期，并最终经历凋亡细胞死亡的剂量依赖性的方式。此外，17-AAG给药显示诱导多种Hsp 90蛋白客户和其他下游效应物（如IGF-IR、Akt、IKK-α、IKK-β、FOXO 1、ERK 1/2和c-Met）的显著下调，导致螯合介导的NF-κ B失活、细胞增殖减少和细胞运动性下降。总之，我们已经清楚地证明了17-AAG对人膀胱癌细胞的细胞周期进展、存活和运动性的剂量依赖性和细胞类型特异性作用，这是由于多个Hsp 90客户的下调和随后的信号传导完整性的破坏。
Background: 17-Allylamino-17-demethoxygeldanamycin (17-AAG), a benzoquinone ansamycin antibiotic, specifically targets heat shock protein 90 (Hsp90) and interferes with its function as a molecular chaperone that maintains the structural and functional integrity of various protein clients involved in cellular signaling. In this study, we have investigated the effect of 17-AAG on the regulation of Hsp90-dependent signaling pathways directly implicated in cell cycle progression, survival and motility of human urinary bladder cancer cell lines.Methods: We have used MTT-based assays, FACS analysis, Western blotting, semi-quantitative RT-PCR, immunocytochemistry and scratch-wound assay in RT4, RT112 and T24 human urinary bladder cancer cell lines.Results: We have demonstrated that, upon 17-AAG treatment, bladder cancer cells are arrested in the G1 phase of the cell cycle and eventually undergo apoptotic cell death in a dose-dependent manner. Furthermore, 17-AAG administration was shown to induce a pronounced downregulation of multiple Hsp90 protein clients and other downstream effectors, such as IGF-IR, Akt, IKK-alpha, IKK-beta, FOXO1, ERK1/2 and c-Met, resulting in sequestration-mediated inactivation of NF-kappa B, reduced cell proliferation and decline of cell motility.Conclusions: In total, we have clearly evinced a dose-dependent and cell type-specific effect of 17-AAG on cell cycle progression, survival and motility of human bladder cancer cells, due to downregulation of multiple Hsp90 clients and subsequent disruption of signaling integrity.