2-[(1-methylpropyl)dithio]-1H-imidazole inhibits tubulin polymerization through cysteine oxidation

被引:68
作者
Huber, Kelly [1 ]
Patel, Poulam [1 ]
Zhang, Lei [1 ]
Evans, Helen [1 ]
Westwell, Andrew D. [4 ]
Fischer, Peter M. [2 ]
Chan, Stephen [3 ]
Martin, Stewart [1 ]
机构
[1] Univ Nottingham, Univ Nottingham Hosp, Div Clin Oncol, Sch Mol Med Sci, Nottingham NG5 1PB, England
[2] Univ Nottingham, Sch Pharm, Ctr Biomol Sci, Nottingham NG7 2RD, England
[3] Univ Nottingham, Dept Clin Oncol, NHS Trust, Nottingham NG7 2RD, England
[4] Cardiff Univ, Welsh Sch Pharm, Cardiff, Wales
关键词
D O I
10.1158/1535-7163.MCT-07-0486
中图分类号
R73 [肿瘤学];
学科分类号
100214 ;
摘要
2-[(1-Methylpropyl)dithio]-1H-imidazole (IV-2) is a known inhibitor of the thioredoxin system. It causes the oxidation of cysteine residues from both thioredoxin reductase and thioredoxin, with only the latter leading to irreversible inhibition of protein function. Although IV-2 is considered to be the first specific inhibitor of thioredoxin to undergo evaluation in cancer patients (under the name PX-12), it is unclear whether the oxidative ability of IV-2 is limited to proteins of the thioredoxin family. The current study investigated the specificity of IV-2 by examining its interaction with tubulin, a protein in which cysteine oxidation causes loss of polymerization competence. The cellular effects of IV-2 were examined in MCF-7 breast cancer and endothelial cells (human umbilical vein endothelial cells). Immunocytochemistry revealed a loss of microtubule structure with Western blot analysis confirming that treated cells contained a higher proportion of unpolymerized tubulin. Cell-free tubulin polymerization assays showed a dose-dependent inhibition of tubulin polymerization and depolymerization of preformed microtubules, confirming a direct interaction between IV-2 and tubulin. Further investigation of the tubulin interaction, through analysis of sulfhydryl reactivity and disulfide bond formation, suggested that IV-2 acts through the oxidation of cysteines in tubulin. Biochemical assays indicated that the oxidative properties of IV-2 are not limited to thioredoxin and tubulin, as cysteine-dependent proteases were also inhibited. Breast cancer cells with thioredoxin silenced by short interfering RNA remained sensitive to IV-2, albeit at higher antiproliferative GI(50) values than in cells with normal thioredoxin function. These findings show that modulation of targets other than thioredoxin contribute to the effects of IV-2 on proliferating cells.
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收藏
页码:143 / 151
页数:9
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