The overall goal of this National Cooperative Drug Discovery Group (NCDDG) application is to design, synthesize and evaluate potential anticancer drugs based on inhibition of farnesylation of the ras oncogene product, mutated forms of which are involved in about 30% of human cancers. Farnesylation is required for Ras membrane association and malignant transformation. Therefore, the molecular target of our rational approach to cancer therapeutics is the enzyme Ras farensyltransferase (FTase), which catalyzes the transfer of a farnesyl isoprenoid lipid to the cysteine contained in the carboxylterminal CAAX sequence of Ras (C=cysteine, A=aliphatic, X=methionine or serine). We and others have designed CAAX peptidomimetics that block Ras farnesylation, signaling and transformation. Although these peptidomimetics are potent inhibitors of FTase in vitro, their ability to block Ras function in whole cells is weak and an intense search for better inhibitors is underway. A better understanding of the inhibitors structure activity relationship with FTase, their mechanism of action and the basis of their selectivity at the biochemical, the cellular signaling and the in vivo level will be critical to the successful design of clinically useful FTase inhibitors. Our multidisciplinary approach to accomplish these goals consists of three distinct but complementary programs. Program #1 will design and synthesize CAAX peptidomimetics, transition state bisubstrate analogs of the farnesylpyrophosphate/CAAX complex and mechanism-based inhibitors. Using a variety of substrates, Program #2 will perform structure/function studies on these compounds to determine the requirements for selective inhibition of farnesylation relative to geranylgeranylation in vitro and in vivo and the effects of the compounds on cell growth. Program #3 will determine the effects of these compounds on Ras signaling and transformation in cell culture systems. Both Programs #2 and #3 will address selectivity issues in terms of, H-vs. K-vs. N-Ras, Ras vs. Ras- related proteins, and Ras vs. other oncogene products. Feedback from Programs #2 and #3 will be used by Program #1 to produce second and third generation leads. The best lead compounds to emerge from these combined studies will then be evaluated by Program #2 for their ability to specifically block the growth of tumors with aberrant Ras function, using a nude mouse xenograft model. Finally, advanced preclinical and clinical studies will be carried out in collaboration with our industrial and clinical partners. The work described in this NCDDG will enhance our understanding of the mechanism of action of Ras FTase inhibitors, and ultimately result in novel anticancer drugs that will broaden the spectrum of human tumors that can be treated successfully.
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