BACKGROUND: Class III b-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 b-tubulin expression, avoiding the toxicity associated with chronic overexpression of class III b-tubulin, and study the efficacy of a panel of clinical and pre-clinical drugs in this model. METHODS: MCF-7 (ER þ ve) and MDA-MB-231 (ERÀve) 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 b-tubulin mRNA and protein expression. RESULTS: The model allowed for subtle changes in class III b-tubulin expression to be achieved, which had no direct effect on the viability of the cells. Class III b-tubulin overexpression conferred resistance to paclitaxel and vinorelbine, whereas downregulation of class III b-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 b-tubulin expression. CONCLUSION: These data indicate that the effect of class III b-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.

A chimeric approach is used to discover microtubule disruptors with excellent in vitro activity and oral bioavailability; a ligand-protein interaction with carbonic anhydrase that enhances bioavailability is characterised by protein X-ray crystallography. Dosing of a representative chimera in a tumour xenograft model confirms the excellent therapeutic potential of the class.

The synthesis, SAR, and preclinical evaluation of 17-cyanated 2-substituted estra-1,3,5(10)-trienes as anticancer agents are discussed. 2-Methoxy-17beta-cyanomethylestra-1,3,5(10)-trien-3-ol ( 14), but not the related 2-ethyl derivative 7, and the related 3- O-sulfamates 8 and 15 display potent antiproliferative effects (MCF-7 GI 50 300, 60 and 70 nM, respectively) against human cancer cells in vitro. Investigation of the SAR reveals that a sterically unhindered hydrogen bond acceptor attached to C-17 is most likely key to the enhanced activity. Compound 8 displayed significant in vitro antiangiogenic activity, and its ability to act as a microtubule disruptor was confirmed. Inhibitory activity of the sulfamate derivatives against steroid sulfatase and carbonic anhydrase II (hCAII) was also observed, and the interaction between 15 and hCAII was investigated by protein crystallography. The potential of these multimechanism anticancer agents was confirmed in vivo, with promising activity observed for both 14 and 15 in an athymic nude mouse MDA-MB-231 human breast cancer xenograft model.