We are investigating the mechanisms by which the adhesive glycoproteins TSP1 and TSP2 regulate tumor growth, metastasis, and angiogenesis. Data from tumor xenograft and transgenic mouse models demonstrated that TSP1 and TSP2 are suppressors of tumor progression. Although the suppressive activity of TSP1 was initially ascribed to inhibition of angiogenesis, we have recently obtained evidence for significant direct effects of TSP1 on both tumor cells and the host immune response. Using synthetic peptides, recombinant TSP1 fragments, and mutagenesis, we defined domains and specific amino acid sequences that mediate activities of TSP1 toward each of these cell types. We identified peptide sequences in TSP1 that mimic the anti-angiogenic activities of the whole molecule. Stable analogs of these peptides inhibited angiogenesis in several animal models and are being developing for therapeutic applications. We also identified a pro-angiogenic sequence in TSP1 and its endothelial cell receptor. The specific responses of tumor cells, T lymphocytes, and endothelial cells to TSP1 arise from the utilization of distinct combinations of cell surface TSP1 receptors and result in different intracellular signals in each cell type. Recognition of TSP1 by the integrin a3b1 on breast carcinoma cells is specifically modulated by IGF1 and CD98. Activation by IGF1 is associated with externalization of HSP60, and exogenous HSP60 is sufficient to activate a3b1. The same integrin is stimulated by EGF, but not by IGF1, in small cell lung carcinoma and by VE-cadherin in endothelial cells. In addition to the regulation of this TSP1 receptor, we found that the a3b1 recognition site on TSP1 is conformationally regulated by TSP1 binding to another matrix protein, fibronectin. Most T lymphocytes lack a3b1 integrin, and their TSP1 responses are mediated by CD47, heparan sulfate proteoglycan, and a4b1 integrin. a4b1 integrin also recognizes TSP2, whereas a3b1 is specific for TSP1. An amino acid sequence in the N-terminal domain of TSP1 was identified that is recognized by a4b1 integrin. We are defining the cell-specific signal transduction pathways for each TSP receptor using TSP1-null transgenic mice and by identifying genes that are regulated by these TSP1-initiated signals in specific cell types. Based on DNA microarray analysis, TSP1 acts globally to suppress T cell receptor signaling. Two TSP1 receptors, CD47 and a heparan sulfate proteoglycan, mediate this activity. In contrast, adhesion on TSP1 and TSP2 required only the a4b1 integrin, and regulation of T cell chemotaxis and matrix metalloproteinase gene expression required the integrin and indirectly required CD47.
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