Characterization of a new potent, in vivo neutralizing monoclonal antibody to human vascular endothelial growth factor

• Published in: Journal of cancer research and clinical oncology Vol. 125; no. 6; pp. 336 - 342
• Main Authors: SCHLAEPPI, J.-M, SIEMEISTER, G, WEINDEL, K, SCHNELL, C, WOOD, J
• Format: Journal Article
• Language: English
• Published: Berlin Springer 1999; Springer Nature B.V
• Subjects: Animals; Antibodies, Monoclonal - therapeutic use; Antigen-Antibody Reactions; Antineoplastic agents; Biological and medical sciences; Endothelial Growth Factors - immunology; Endothelium, Vascular - cytology; Endothelium, Vascular - immunology; Epitope Mapping; Humans; Immunotherapy; Lymphokines - immunology; Medical sciences; Mice; Neovascularization, Pathologic - therapy; Pharmacology. Drug treatments; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors; Human; Animal model; Antigenic determinant; Rodentia; Binding site; Blocking antibody; Monoclonal antibody; In vitro; Angiogenesis; In vivo; Vertebrata; Mammalia; Vascular endothelium growth factor; Mouse; Animal; Immunotherapy; Neovascularization
• ISSN: 0171-5216; 1432-1335
• DOI: 10.1007/s004320050283

Vascular endothelial growth factor (VEGF) is an important mediator of tumor-induced angiogenesis and represents a potential target for anticancer therapy. Therefore, we prepared a panel of monoclonal antibodies (mAb) against both the VEGF121 and VEGF165 isoforms. Three of them completely neutralized the mitogenic stimulation by VEGF of human umbilical vein endothelial cells at mAb concentrations below 0.1 microg/ml. The most potent one, with a dissociation constant (Kd) of 8 pM, inhibited, in a dose-dependent manner, VEGF-induced angiogenesis in a growth factor implant model in mice. A complete inhibition of the angiogenic response was obtained by daily intraperitoneal injections of 10 microg mAb/mouse. Angiogenesis induced by basic fibroblast growth factor was not inhibited by the mAb. Epitope mapping of the mAb, performed by competitive enzyme-linked immunosorbent assay and Western blot analysis, showed that it did not bind to the reduced and denatured monomer of VEGF. Substitutions of three residues (Q87R, G88K, Q89K), located on the major surface loop beta5 to beta6 of VEGF, resulted in the complete loss of binding (more than 400-fold reduction). The results suggest that the mAb binds primarily to a conformation-dependent epitope on the VEGF dimeric form, encompassing one of the loop regions involved in KDR receptor binding. The mAb with its strong neutralizing properties represents a useful agent for effective blocking of VEGF-mediated tumor neovascularization.