Vascular endothelial growth factor 165 (VEGF165) activities are inhibited by carboxymethyl benzylamide dextran that competes for heparin binding to VEGF165 and VEGF165•KDR complexes

We have previously shown that carboxymethyl dextran benzylamide (CMDB7), a heparin-like molecule, inhibits the growth of tumors xenografted in nude mice, angiogenesis, and metastasis by altering the binding of angiogenic growth factors, including platelet-derived growth factor, transforming growth factor beta, and fibroblast growth factor 2, to their specific receptors. In this study, we explore the effect of CMDB7 on the most specific angiogenic growth factor, vascular endothelial growth factor 165 (VEGF(165)). We demonstrate here that CMDB7 inhibits the mitogenic effect of VEGF(165) on human umbilical vein endothelial cells (HUV-ECs) by preventing the VEGF(165)-induced VEGF receptor-2 (KDR) autophosphorylation and consequently a specific intracellular signaling. In competition experiments, the binding of I-125-VEGF(165) to HUV-ECs is inhibited by CMDB7 with an IC50 of 2 muM. Accordingly, CMDB7 inhibits the cross-linking of I-125-VEGF(165), to the surface of HUV-ECs, causing the disappearance of both labeled complexes, 170-180 and 240-250 kDa. We show that CMDB7 increases the electrophoretic mobility of VEGF165, thus evidencing formation of a stable complex with this factor. Moreover, CMDB7 reduces the I-125-VEGF(165) binding to coated heparin-albumin and prevents a heparin-induced increase in iodinated VEGF(165) binding to soluble I-125-KDR-Fc chimera. Concerning KDR, CMDB7 has no effect on I-125-KDR-Fc electrophoretic migration and does not affect labeled KDR-Fc binding to coated heparin-albumin. In the presence of VEGF(165), I-125-KDR-Fc binding to heparin is enhanced, and under these conditions, CMDB7 interferes with KDR binding. These data indicate that CMDB7 effectively inhibits the VEGF(165) activities by interfering with heparin binding to VEGF(165) and VEG(165).KDR complexes but not by direct interactions with KDR.