The
epothilones are naturally occurring
antimitotic drugs that share with the
taxanes a similar mechanism of action without apparent structural similarity. 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
epothilone 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
taxane-driven
tubulin polymerization caused by acquired
beta-tubulin mutations (beta274(Thr-->Ile) and beta282(
Arg-->Gln)) located in the atomic model of alphabeta-
tubulin near the
taxane-binding site. Using molecular modeling, we investigated the conformational behavior of
epothilone, which led to the identification of a common pharmacophore shared by
taxanes and
epothilones. Although two binding modes for the
epothilones were predicted, one mode was identified as the preferred
epothilone conformation as indicated by the activity of a potent
pyridine-
epothilone analogue. In addition, the structure-activity relationships of multiple
taxanes and
epothilones in the
tubulin mutant cells can be fully explained by the model presented here, verifying its predictive value. Finally, these pharmacophore and activity data from mutant cells were used to model the
tubulin binding of sarcodictyins, a distinct class of microtubule stabilizers, which in contrast to
taxanes and the
epothilones interact preferentially with the mutant tubulins. The unification of
taxane,
epothilone, and sarcodictyin chemistries in a single pharmacophore provides a framework to study
drug-
tubulin interactions that should assist in the rational design of agents targeting
tubulin.