Preparation and functional evaluation of RGD-modified proteins as alpha(v)beta(3) integrin directed therapeutics.
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.
Kok
Robbert J
RJ
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
Schraa
Astrid J
AJ
Bos
Erwin J
EJ
Moorlag
Henk E
HE
Asgeirsdóttir
Sigridur A
SA
Everts
Maaike
M
Meijer
Dirk K F
DK
Molema
Grietje
G
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*
D020533:Angiogenesis Inhibitors; D007074:Immunoglobulin G; D009842:Oligopeptides; D010455:Peptides; D011506:Proteins; D019030:Receptors, Vitronectin; C047981:arginyl-glycyl-aspartic acid