Recombinant semaphorin 6A-1 ectodomain inhibits in vivo growth factor and tumor cell line-induced angiogenesis.

The Semaphorins are a large family of transmembrane, GPI-anchored and secreted proteins that play an important role in neuronal and endothelial cell guidance. A human gene related to the class 6 Semaphorin family, Semaphorin 6A-1 (Sema 6A-1) was identified by homology-based genomic mining. Recent implication of Sema 3 family members in tumor angiogenesis and our expression analysis of Sema 6A-1 suggested that class 6 Semaphorin might effect tumor neovascularization. The mRNA expression of Sema 6A-1 was elevated in several renal tumor tissue samples relative to adjacent nontumor tissue samples from the same patient. Sema 6A-1 transcript was also expressed in the majority of renal clear cell carcinoma (RCC) cell lines and to a lesser extent in endothelial cells. To test the role of Sema 6A-1 in tumor angiogenesis, we engineered, expressed and purified the Sema 6A-1 soluble extracellular domain (Sema-ECD). The purified Sema-ECD was screened in a variety of endothelial cell-based assays both in vitro and in vivo. In vitro, Sema-ECD blocked VEGF-mediated endothelial cell migration. These effects were explained in part by our observation in endothelial cells that Sema-ECD inhibited VEGF-mediated Src, FAK and ERK phosphorylation. In vivo, mouse Matrigel assays demonstrated that the intraperitoneal administration of recombinant Sema-ECD inhibited both bFGF/VEGF and tumor cell line-induced neovascularization. These findings reveal a novel therapeutic utility for Sema 6A-1 (Sema-ECD) as an inhibitor of growth factor as well as tumor-induced angiogenesis.

Angioarrestin: an antiangiogenic protein with tumor-inhibiting properties.

The angiopoietins comprise a family of proteins that have pro or antiangiogenic activities. Through a proprietary technology designed to identify transcripts of all expressed genes, we isolated a cDNA encoding an angiopoietin-related protein that we designate angioarrestin. The mRNA expression profile of angioarrestin was striking in that it was down-regulated in many tumor tissues when compared with adjacent nontumor tissue, suggesting a role for this protein in tumor inhibition. To test this hypothesis, we ectopically expressed angioarrestin in HT1080 tumor cells and measured pulmonary tumor nodule formation in nude mice. HT1080 cells expressing angioarrestin showed a marked reduction in the number and size of tumor nodules. In vitro, the recombinant protein was systematically tested in a number of endothelial cell assays and found to block critical processes involved in the angiogenic cascade, such as vascular endothelial growth factor/basic fibroblast growth factor-mediated endothelial cell proliferation, migration, tubular network formation, and adhesion to extracellular matrix proteins. These findings reveal a novel function for angioarrestin as an angiogenesis inhibitor and indicate that the molecule may be a potential cancer therapeutic.