Growth of new blood vessels (angiogenesis), required for all tumor growth, is stimulated by the expression of vascular endothelial growth factor (VEGF). VEGF is up-regulated in all known solid tumors but also in atherosclerosis, diabetic retinopathy, arthritis, and many other conditions. Conventional VEGF isoforms have been universally described as proangiogenic cytokines. Here, we show that an endogenous splice variant, VEGF165b, is expressed as protein in normal cells and tissues and is circulating in human plasma. We also present evidence for a sister family of presumably inhibitory splice variants. Moreover, these isoforms are down-regulated in prostate cancer. We also show that VEGF165b binds VEGF receptor 2 with the same affinity as VEGF165 but does not activate it or stimulate downstream signaling pathways. Moreover, it prevents VEGF165-mediated VEGF receptor 2 phosphorylation and signaling in cultured cells. Furthermore, we show, with two different in vivo angiogenesis models, that VEGF165b is not angiogenic and that it inhibits VEGF165-mediated angiogenesis in rabbit cornea and rat mesentery. Finally, we show that VEGF165b expressing tumors grow significantly more slowly than VEGF165- expressing tumors, indicating that a switch in splicing from VEGF165 to VEGF165b can inhibit tumor growth. These results suggest that regulation of VEGF splicing may be a critical switch from an antiangiogenic to a proangiogenic phenotype.