The goal of this project is the development of new antineoplastic agents directed against tubulin, a protein critical for cell division and for the maintenance of cellular morphology. Detailed mechanistic and structure- activity studies were performed with many classes of compounds. [3H]Dolastatin 10 was found to bind avidly but reversibly to tubulin, with a dissociation constant of 26 nM. The drug was also shown to induce formation of complex ring and spiral aggregates, with the smallest detectable radiolabeled species containing two molecules each of tubulin and [3H]dolastatin 10. Inhibition of dolastatin 10 binding by maytansine and spongistatin l (noncompetitive) and vincristine and phomopsin A (competitive) was studied in detail. A new potent colchicine site inhibitor of extraordinary structure, curacin A, was examined. While inhibiting normal tubulin polymerization, it induces formation of complex alternate spiral polymers. The endogenous estrogen metabolite, 2- methoxyestradiol, was shown to bind in the colchicine site of tubulin, acting as a weak inhibitor of polymerization. Following preparation of [4-3H]2-methoxyestradiol, we showed that the drug is also incorporated into polymer in large amounts. Radiolabeled A-ring modified analogs of colchicine were shown to form a covalent bond with cys-239 of beta- tubulin, providing new information about the colchicine binding site. Structure-activity studies with a series of paclitaxel analogs showed that substituents at the meta position in the C(2)-benzoyl group substantially enhanced the interaction of this type of compound with tubulin. Extensive-structure activity studies were performed with phenylquinolone derivatives, and work continued with the highly potent spongistatin series.
