Oncology Drug Discovery and Women's Health Group, Faculty of Medicine, Imperial College London, St Mary's Hospital, London W2 1NY, UK.
Br J Cancer. 2010 Jan 19;102(2):316-24. doi: 10.1038/sj.bjc.6605489. Epub 2009 Dec 22.
Class III beta-tubulin overexpression is a marker of resistance to microtubule disruptors in vitro, in vivo and in the clinic for many cancers, including breast cancer. The aims of this study were to develop a new model of class III beta-tubulin expression, avoiding the toxicity associated with chronic overexpression of class III beta-tubulin, and study the efficacy of a panel of clinical and pre-clinical drugs in this model.
MCF-7 (ER+ve) and MDA-MB-231 (ER-ve) were either transfected with pALTER-TUBB3 or siRNA-tubb3 and 24 h later exposed to test compounds for a further 96 h for proliferation studies. RT-PCR and immunoblotting were used to monitor the changes in class III beta-tubulin mRNA and protein expression.
The model allowed for subtle changes in class III beta-tubulin expression to be achieved, which had no direct effect on the viability of the cells. Class III beta-tubulin overexpression conferred resistance to paclitaxel and vinorelbine, whereas downregulation of class III beta-tubulin rendered cells more sensitive to these two drugs. The efficacy of the colchicine-site binding agents, 2-MeOE2, colchicine, STX140, ENMD1198 and STX243 was unaffected by the changes in class III beta-tubulin expression.
These data indicate that the effect of class III beta-tubulin overexpression may depend on where the drug's binding site is located on the tubulin. Therefore, this study highlights for the first time the potential key role of targeting the colchicine-binding site, to develop new treatment modalities for taxane-refractory breast cancer.
III 类微管蛋白β表达过度是许多癌症（包括乳腺癌）在体外、体内和临床中对微管破坏剂产生耐药性的标志物。本研究旨在开发一种新的 III 类微管蛋白表达模型，避免 III 类微管蛋白慢性过表达相关的毒性，并在该模型中研究一组临床前和临床药物的疗效。
MCF-7（ER+ve）和 MDA-MB-231（ER-ve）用 pALTER-TUBB3 或 siRNA-tubb3 转染，24 小时后用测试化合物进一步处理 96 小时进行增殖研究。RT-PCR 和免疫印迹用于监测 III 类微管蛋白β mRNA 和蛋白表达的变化。
该模型允许实现 III 类微管蛋白表达的微妙变化，而对细胞活力没有直接影响。III 类微管蛋白过表达导致对紫杉醇和长春瑞滨的耐药性，而 III 类微管蛋白下调使细胞对这两种药物更敏感。秋水仙碱结合位点结合剂 2-MeOE2、秋水仙碱、STX140、ENMD1198 和 STX243 的疗效不受 III 类微管蛋白表达变化的影响。
这些数据表明，III 类微管蛋白过表达的效果可能取决于药物结合位点在微管上的位置。因此，本研究首次强调了靶向秋水仙碱结合位点的潜在关键作用，以开发治疗紫杉醇耐药性乳腺癌的新治疗方法。
