Growth of tumors and metastasis are processes known to require neovascularization. To ascertain the participation of the endogenous angiogenic inhibitor thrombospondin-1 (TSP1) in tumor progression, we generated mammary tumor-prone mice that either lack, or specifically overexpress, TSP1 in the mammary gland. Tumor burden and vasculature were significantly increased in TSP1-deficient animals, and capillaries within the tumor appeared distended and sinusoidal. In contrast, TSP1 overexpressors showed delayed tumor growth or lacked frank tumor development (20% of animals); tumor capillaries showed reduced diameter and were less frequent. Interestingly, absence of TSP1 resulted in increased association of vascular endothelial growth factor (VEGF) with its receptor VEGFR2 and higher levels of active matrix metalloproteinase-9 (MMP9), a molecule previously shown to facilitate both angiogenesis and tumor invasion. In vitro, enzymatic activation of proMMP9 was suppressed by TSP1. Together these results argue for a protective role of endogenous inhibitors of angiogenesis in tumor growth and implicate TSP1 in the in vivo regulation of metalloproteinase-9 activation and VEGF signaling.C ancer is a multistep process that includes deregulation of cell cycle, transformation, invasion of stroma, and metastasis (1). In addition, successful establishment of solid tumors depends on neovascularization (2, 3). More recently, the contribution of angiogenesis has been further supported by the demonstration that angiogenic inhibitors suppress tumor growth. Our present understanding of the mechanism of action of most angiogenic inhibitors is limited, and further efforts are required to evaluate their effects in vivo.Thrombospondin-1 (TSP1) was the first protein to be recognized as a potential endogenous suppressor of capillary morphogenesis in vivo (4). Most likely, modulation of vasculature is not its only function; this was well illustrated by the fact that mice lacking TSP1 developed epithelial lung hyperplasia and pneumonia with multifocal inflammatory sites, but no significant vascular phenotype was observed (5). Inflammation and several pathological conditions have been shown to induce rapid and robust TSP1 expression (6, 7). Its role in these situations remains elusive; however, it has been shown that wound healing is aberrant in TSP1-null mice (8). With respect to antiangiogenic capabilities, TSP1 inhibits endothelial cell proliferation (9) and migration (10) and can induce endothelial apoptosis (11,12). In addition, xenograft implants of tumor cells transfected with TSP1 developed smaller tumors than the parental cell lines in a variety of assays (13-16), and fusion proteins and peptides from specific domains of TSP1 have been shown to inhibit proliferation of melanoma cells and reduce tumor growth in immunosuppressed animals (17). In clinical studies, expression of TSP1 has been inversely correlated with malignant progression of breast cancer, melanoma, and lung carcinomas (18,19). A hypothesis consistent with these data is t...