Wild-type p53 suppresses angiogenesis in human leiomyosarcoma and synovialsarcoma by transcriptional suppression of vascular endothelial growth factor expression

Our recent studies (R. Pollock et al,, Clin, Cancer Res., 4: 1985-1994, 199 8; M, Milas et al, Cancer Gene Ther., in press, 2000) have shown that the r estoration of wild type (wt) p53 enhances cell cycle control in vitro and i nhibits the growth of human soft-tissue sarcoma in severe combined immunode ficient mice. We hypothesized that the antitumor effect of wt p53 overexpre ssion in sarcoma cells is attributable not only to enhanced cell cycle cont rol but also to inhibition of angiogenesis. We evaluated the effect of rest oring wt p53 function on angiogenesis in human soft-tissue sarcoma harborin g mutant p53, Restoration of wt p53 expression in human leiomyosarcoma SKLM S-1 cells that contain mutant p53 markedly inhibited angiogenesis induced b y tumor cells in vivo, Angiogenesis assays using an in vivo Matrigel plug a ssay demonstrated that less neovascularization in severe combined immunodef icient mice was observed with conditioned medium (CM) from human synovial s arcoma cells expressing wt p53 compared with CM from human synovial sarcoma cells expressing mutant p53. Microvessel density and microvessel counts we re lower in tumor xenografts from cells containing wt p53 than in tumor xen ografts from cells containing mutant p53. The growth and migration of murin e lung endothelial cells were decreased when cells were treated with CM fro m sarcoma cells expressing wt p53 compared with CM from sarcoma cells expre ssing mutant p53, The introduction of wt p53 into sarcoma cells containing mutant p53 significantly reduced the expression of vascular endothelial gro wth factor (VEGF), which is a key mediator of tumor angiogenesis. Stimulati on of endothelial cell migration by CM from cells expressing mutant p53 was significantly reduced after anti-VEGF neutralrzing antibody was added to t he CM. Using luciferase as the reporter of VEGF promoter activity, we found that wt p53 inhibited VEGF promoter activity in SKLMS-1 cells. Deletion an alysis defined an 87-bp region (bp -135 to -48) in the VEGF promoter that i s necessary for inhibiting VEGF promoter activity by wt p53. The transcript ion factor Spl may be involved in the repression of VEGF promoter activity by wt p53 in SKLMS-1 cells. These data indicated that wt p53 can suppress a ngiogenesis in human soft-tissue sarcomas by transcriptional repression of VEGF expression.