INHIBITION OF ANGIOGENESIS-- PROTEINS BINDING TO TSP2

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The extracellular matrix protein thrombospondin (TSP) 2, like TSP1, interacts with a number of cell surface receptor and extracellular matrix proteins. The immediate goals of this grant are to focus on and analyze the TSP structural features and protein partners relevant to the function of inhibition of angiogenesis. The pursuit of these goals is facilitated by the stimulating, well-equipped and intellectually enriched research environment available for the performance of the work. The findings will lay the framework for long-term goals aimed at understanding the molecular basis of vascular biological functions in vivo. It is widely recognized nowadays that in many pathological conditions a common underlying process is a derangement in angiogenesis. Inhibitors of angiogenesis have potential clinical application in disease states where abnormal blood vessel formation is related to disease progression e.g. solid tumors, hemangiomas and inflammatory diseases. On the other hand, enhancers of angiogenesis would be useful in conditions resulting from insufficient blood supply e.g. wound healing, coronary artery disease and stroke. TSP1 and TSP2 have been of particular interest in the medical sciences recently because of their ability to inhibit angiogenesis. The importance of the TSP1 and TSP2 proteins in the regulation of angiogenesis has been underscored by recent in vivo studies that demonstrated TSP-dependent induction of concomitant tumor resistance, suppression of tumor growth, and inhibition of vascularization of polydimethylsiloxane implants. However, the mechanism(s) of action remains unknown. This gap in our knowledge is reflected in the low potency of thrombospondin-based anti-angiogenic synthetic peptides, which was an order of magnitude less than that of intact TSP1. Since inhibition assays using synthetic peptides and knockout studies suggested the possibility of multiple signal pathways elicited or matrix reactions initiated, we wanted to characterize proteins that interact with TSP2. There is very little known about TSP2 for most studies have been on TSP1. Use of the TSP2 type 1 repeats in a yeast two-hybrid system led to the isolation of 36 independent clones. The aims of the proposed investigation are to study the TSP structural determinants involved in regulation of angiogenesis, analyze the significance of the interaction of TSP1 and TSP2 with the proangiogenic MMP2, and identify and functionally characterize novel TSP-binding proteins. It is hoped that the studies will serve as the basis for designing more potent antiangiogenic reagents.