This proposal focuses on an in-depth investigation into the mechanism of action of the vinca alkaloids. The vinca alkaloids are a group of chemically related indole-dihydroindole drugs that are currently used in the treatment of a number of forms of cancer. They appear to inhibit cell growth at least in part by the disruption of microtubules. Although a considerable amount has been learned about how the vinca alkaloids inhibit microtubule assembly in vitro, the precise mechanism of action in quantitative kinetic terms is almost completely unknown, as is an understanding of the relationship between the ability of the vinca alkaloids to inhibit the polymerization of microtubules, and to destroy preformed microtubules directly. Further, the antitumor specificities of the individual vinca alkaloids are different, and it is conceivable that the different vinca congeners exert their chemotherapeutic activities on cells by a combination of distinct mechanisms. Our research strategies fall into three specific areas. One major aim is to continue studies begun previously in our laboratory on the quantitative characterization of the mechanism of microtubule assembly inhibition by the vinca alkaloids. These studies will involve the kinetic characterization of tubulin addition and loss at the opposite ends of steady-state microtubules in vitro, using microtubules from several sources and of different molecular compositions. The second goal is to characterize the interactions of the vinca alkaloids that result in the direct depolymerization of microtubules (the peeling of protofilament strands at microtubule ends). The relationship between this interaction of vinblastine with microtubules and the interaction that results in assembly inhibition will be determined. The third goal is to investigate the structure-activity-relationship of the vinca alkaloid derivatives in terms of the abilities of individual congeners to inhibit microtubule polymerization in cells, and the abilities of the congeners to inhibit cell growth, using a human cell line (Hela) whose microtubule properties in vitro have been characterized. The purpose is to determine whether the action of a particular vinca alkaloid derivative in inhibiting cell growth is due exclusively to the disruption of microtubules in cells, or occurs through a concerted ability to inhibit microtubule function and other cellular processes.
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