title: Preparation and functional evaluation of RGD-modified proteins as alpha(v)beta(3) integrin directed therapeutics.
abstract: Tumor blood vessels can be selectively targeted by RGD-peptides that bind to alpha(v)beta(3) integrin on angiogenic endothelial cells. By inhibiting the binding of these integrins to its natural ligands, RGD-peptides can serve as antiangiogenic therapeutics. We have prepared multivalent derivatives of the cyclic RGD-peptide c(RGDfK) by covalent attachment of the peptide to side chain amino groups of a protein. These RGDpep-protein conjugates inhibited alpha(v)beta(3)-mediated endothelial cell adhesion in vitro, while conjugates prepared with a control RAD-peptide showed no activity. Radiobinding and displacement studies with endothelial cells demonstrated an increased affinity of the RGDpep-protein conjugates compared to the free peptide, with IC(50) values ranging from 23 to 0.6 nM, depending on the amount of coupled RGDpep per protein. Compared to the parental RGD-peptide and the related RGD-peptide ligand c(RGDfV), the RGDpep-protein conjugates showed a considerable increase in affinity (IC(50) parent RGDpep: 818 nM; IC(50) c(RGDfV): 158 nM). We conclude that the conjugation of RGD-peptides to a protein, resulting in products that can bind multivalently, is a powerful approach to increase the affinity of peptide ligands for alpha(v)beta(3)/alpha(v)beta(5) integrins.
authors.affiliation: Groningen University Institute for Drug Exploration, Department of Pharmacokinetics and Drug Delivery, University Centre for Pharmacy, A. Deusinglaan 1, 9713 AV Groningen, The Netherlands. r.j.kok@farm.rug.nl
authors.forename: Robbert J
authors.initials: RJ
authors.lastname: Kok
authors.forename: Astrid J
authors.initials: AJ
authors.lastname: Schraa
authors.forename: Erwin J
authors.initials: EJ
authors.lastname: Bos
authors.forename: Henk E
authors.initials: HE
authors.lastname: Moorlag
authors.forename: Sigridur A
authors.initials: SA
authors.lastname: Asgeirsdóttir
authors.forename: Maaike
authors.initials: M
authors.lastname: Everts
authors.forename: Dirk K F
authors.initials: DK
authors.lastname: Meijer
authors.forename: Grietje
authors.initials: G
authors.lastname: Molema
mesh_terms: D020533:Angiogenesis Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology*; D002448:Cell Adhesion / Q000187:drug effects; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry; D009842:Oligopeptides / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D011506:Proteins / Q000737:chemistry* / Q000494:pharmacology*; D019030:Receptors, Vitronectin / Q000187:drug effects*
chemical_list: D020533:Angiogenesis Inhibitors; D007074:Immunoglobulin G; D009842:Oligopeptides; D010455:Peptides; D011506:Proteins; D019030:Receptors, Vitronectin; C047981:arginyl-glycyl-aspartic acid