A common pharmacophore for epothilone and taxanes: Molecular basis for drug resistance conferred by tubulin mutations in human cancer cells

The epothilones are naturally occurring antimitotic drugs that share with t he taxanes a similar mechanism of action without apparent structural simila rity. Although photoaffinity labeling and electron crystallographic studies have identified the taxane-binding site on beta-tubulin, similar data are not available for epothilones. To identify tubulin residues important for e pothilone binding, we have isolated two epothilone-resistant human ovarian carcinoma sublines derived in a single-step selection with epothilone A or B, These epothilone-resistant sublines exhibit impaired epothilone- and tax ane-driven tubulin polymerization caused by acquired beta-tubulin mutations (beta 274(Thr-->lle) and beta 282(Arg-->Gln)) located in the atomic model of alpha beta-tubulin near the taxane-binding site. Using molecular modelin g, we investigated the conformational behavior of epothilone, which led to the identification of a common pharmacophore shared by taxanes and epothilo nes. Although two binding modes for the epothilones were predicted, one mod e was identified as the preferred epothilone conformation as indicated by t he activity of a potent pyridine-epothilone analogue. In addition, the stru cture-activity relationships of multiple taxanes and epothilones in the tub ulin mutant cells can be fully explained by the model presented here, verif ying its predictive value. Finally, these pharmacophore and activity data f rom mutant cells were used to model the tubulin binding of sarcodictyins, a distinct class of microtubule stabilizers, which in contrast to taxanes an d the epothilones interact preferentially with the mutant tubulins, The uni fication of taxane, epothilone, and sarcodictyin chemistries in a single ph armacophore provides a framework to study drug-tubulin interactions that sh ould assist in the rational design of agents targeting tubulin.