Inhibition of vascular endothelial growth factor-induced receptor activation with anti-kinase insert domain-containing receptor single-chain antibodies from a phage display library.
A single-chain antibody phage display library was constructed from spleen cells of mice immunized with a soluble form of a human vascular endothelial growth factor (VEGF) receptor, kinase insert domain-containing receptor (KDR). After two rounds of biopanning, >90% of the clones recovered were specifically reactive to KDR. Subsequent selection identified two clones that blocked VEGF binding to KDR. The clones were expressed in Escherichia coli and purified as soluble single-chain Fv (scFv) antibodies. The affinities of the scFv for binding to KDR were determined by BIAcore analysis (2.1 x 10(-9)-5.9 x 10(-9) M). One scFv, p1C11, was shown to inhibit VEGF-induced KDR phosphorylation and VEGF-stimulated DNA synthesis in human umbilical vein endothelial cells. There is much experimental evidence to suggest that the VEGF/KDR/Flk-1 pathway plays an important role in tumor angiogenesis, a process that is essential for tumor growth and metastasis. The antibodies discussed here, which block VEGF binding to KDR, have potential clinical application in the treatment of cancer and other diseases where pathological angiogenesis is involved.
Department of Molecular and Cell Biology, ImClone Systems Inc., New York, New York 10014, USA. zhenping@imclone.com
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Zhu
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Rockwell
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Lu
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Kotanides
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Pytowski
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DJ
Hicklin
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Bohlen
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L
Witte
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001435:Bacteriophages / Q000235:genetics; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D016228:Endothelial Growth Factors / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology*; D008222:Lymphokines / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D017978:Receptors, Growth Factor / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D040262:Receptors, Vascular Endothelial Growth Factor; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D007128:Immunoglobulin Fragments; D008222:Lymphokines; D012333:RNA, Messenger; D017978:Receptors, Growth Factor; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D004247:DNA; D020794:Receptor Protein-Tyrosine Kinases; D040262:Receptors, Vascular Endothelial Growth Factor