Vascular endothelial growth factor (VEGF) is one of the most important angiogenic growth factors for tumor angiogenesis. Here, we sought to explore whether RNA interference (RNAi) targeting matrix metalloproteinase-2 (MMP-2) could disrupt VEGF-mediated angiogenesis in lung cancer. MMP-2 siRNA inhibited lung cancer cell-induced tube formation of endothelial cells in vitro; addition of recombinant human-MMP-2 restored angiogenesis. MMP-2 transcriptional suppression decreased VEGF, phosphatidylinositol 3-kinase (PI3K) protein levels and AKT phosphorylation in lung cancer cells. In addition, MMP-2 suppression decreased hypoxia inducible factor-1alpha (HIF-1alpha), a transcription factor for VEGF, as determined by electrophoretic mobility shift assay (EMSA). We also show that MMP-2 suppression disrupted PI3K dependent VEGF expression; ectopic expression of myr-AKT restored VEGF inhibition. Further, MMP-2 suppression decreased the interaction of integrin-alphaVbeta3 and MMP-2 as confirmed by immunoprecipitation analyses. Studies with either function blocking integrin-alphaVbeta3 antibody or MMP-2 specific inhibitor (ARP-100) indicate that suppression of MMP-2 decreased integrin-alphaVbeta3-mediated induction of PI3K/AKT leading to decreased VEGF expression. Moreover, A549 xenograft tissue sections from mice that treated with MMP-2 siRNA showed reduced expression of VEGF and the angiogenic marker, factor-VIII. The inhibition of tumor angiogenesis in MMP-2 suppressed tumor sections was associated with decreased co-localization of integrin-alphaVbeta3 and MMP-2. In summary, these data provide new insights into the mechanisms underlying MMP-2-mediated VEGF expression in lung tumor angiogenesis.