This proposal focuses on the application of a chemical approach to understanding and controlling the pathways and machinery of cell motility, processes for which small-molecule inhibitors are presently limited, with the goal of discovering new probes and proteins involved in cell migration. In addition to their great value as tools for basic biomedical research, small molecules that modulate cell migration may be of direct therapeutic importance. Anti-migratory compounds have great potential as anti-cancer agents, since both tumor angiogenesis and metastasis depend on cell migration. Using a combination of chemical and cellular approaches, we have previously shown that Rac, phosphoinositides and c-Jun N-terminal kinase are essential for cell migration during epithelial wound closure. We have discovered a number of new small-molecule inhibitors of cell migration, among the most potent inhibitors of cell motility known. The proposed research involves the elucidation of the mechanisms of action of four of these new inhibitors and the role of their cellular protein targets in the control of cell migration. Using binding assay-guided target purification, we have already identified the cellular protein targets of two of these compounds, and this proposal deals in part with characterization of these proteins as drug targets and modulators of cell migration. Specifically, we have discovered one compound that targets the signaling molecule Raf kinase inhibitor protein (RKIP) and acts by perturbing a critical protein-protein interaction, a relatively rare mode of action for a small-molecule inhibitor. In the process of validating RKIP as a relevant target, we have also found that RKIP positively regulated cell migration, and its overexpression results in a striking cancer-like transition of epithelial cells to a highly migratory fibroblast-like phenotype. Another of our new cell migration inhibitors specifically binds the membrane/cytoskeleton adaptor and signaling scaffold protein radixin. We will expand upon these findings. We will also synthesize analogs of the other two highly potent cell migration inhibitors we have discovered for structure-activity relationship studies, followed by preparation of affinity derivatives, isolation and identification of their specific cellular targets and determination of their molecular and cellular mechanisms of action. Cancer treatment has traditionally focused on trying to stop the ability of cancer cells to multiply. However, the development of malignant cancers also involves the migration of cells from one location to another, particularly when tumor cells migrate to form secondary tumor masses. We have discovered new drugs that prevent cells from migrating, and now we seek to find out how they do so in the hope of learning how cell migration is controlled and how to limit it. ? ? ?
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