TAXOL--MECHANISMS OF ACTION AND RESISTANCE

DESCRIPTION: (Applicant's Abstract) The long-term objectives of this application are to acquire a thorough understanding of the mechanisms of action and of resistance to Taxol, an antitumor agent that is known to be efficacious in the treatment of human cancer. Knowledge gained from the studies described in this application will improve the ability to design Taxol analogs with a better therapeutic index and to develop strategies to reverse or overcome drug resistance. The understanding of Taxol will be applied to other natural products such as the epothilones, whose mechanisms of action and resistance may be similar to that of Taxol. The specific objectives of this application are to: 1). Define the binding site for Taxol in the microtubule. Taxol analogs with radiolabeled photoreactive substituents at defined positions around the taxane nucleus and in the A-ring sidechain will be used to define the pocket in the microtubule in which Taxol fits. Each photoreactive substituent will provide another contact point between the drug and the microtubule. 2). Determine if modulating the levels of beta-tubulin isotypes in cells can alter sensitivity to Taxol. Two approaches will be used to analyze this problem; one involves inducible gene expression with the tetracycline and/or ecdysone regulated system and the other beta-tubulin antisense oligonucleotides. 3). Expand the applicant's studies on analyzing alterations in the levels of beta-tubulin isotypes in human ovarian tumors that acquire Taxol resistance. 4). Investigate endogenous molecules, such as p19, that regulate microtubule disassembly and may thereby antagonize the activity of Taxol and modulate low level resistance to the drug. 5). Perform structure activity relationships (SAR) studies on epothilone analogs to determine the structural requirements for biological activity. Examine the sensitivity of Taxol-resistant cells to epothilone and develop epothilone-resistant cells so that the two drugs can be carefully compared.