Identification of a VWF peptide antagonist that blocks platelet adhesion under high shear conditions by selectively inhibiting the VWF-collagen interaction.
BACKGROUND
Because the collagen-VWF-GPIb/IX/V axis plays an important role in thrombus formation, it represents a promising target for development of new antithrombotic agents.


OBJECTIVE
We used phage display to identify potential small peptides that interfere with the VWF-collagen binding and might serve as lead products for the development of possible oral antithrombotic compounds.


METHODS
A random linear heptamer peptide library was used to select VWF-binding peptides.


RESULTS
We identified a phage clone, displaying the YDPWTPS sequence, further referred to as L7-phage, that bound to VWF in a specific and a dose-dependent manner. This L7-phage specifically inhibited the VWF-collagen interaction under both static and flow conditions. Epitope mapping using deletion mutants of VWF revealed that the L7-phage does not bind to the known collagen-binding A3 domain within VWF, but to the more carboxyterminal situated C domain. This inhibition was not due to steric hindrance of the A3 domain-collagen interaction by the L7-phage. Indeed, a tetrabranched multi-antigen peptide (MAP) presenting four copies of the peptide, but not the scrambled MAP, also inhibited VWF-collagen interaction under conditions of high shear stress at a concentration of 148 nmol L(-1).


CONCLUSIONS
Based on these results, we conclude that we have identified the first peptide antagonist that binds to the VWF C domain and by this specifically inhibits the VWF binding to collagen, suppressing platelet adhesion and aggregation under high shear conditions. As a consequence, this peptide and its future derivates are potentially interesting antithrombotic agents.
Szanto
T
T
Clinical Research Center, Medical and Health Science Center, University of Debrecen, Debrecen H-4012, Hungary.
Vanhoorelbeke
K
K
Toth
G
G
Vandenbulcke
A
A
Toth
J
J
Noppe
W
W
Deckmyn
H
H
Harsfalvi
J
J
D003094:Collagen / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D015195:Drug Design; D004353:Drug Evaluation, Preclinical; D018604:Epitope Mapping; D005343:Fibrinolytic Agents / Q000302:isolation & purification / Q000494:pharmacology*; D018056:Hemorheology; D006801:Humans; D009842:Oligopeptides / Q000302:isolation & purification / Q000494:pharmacology*; D019151:Peptide Library; D010973:Platelet Adhesiveness / Q000187:drug effects*; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D013314:Stress, Mechanical; D014841:von Willebrand Factor / Q000037:antagonists & inhibitors* / Q000276:immunology / Q000378:metabolism
D005343:Fibrinolytic Agents; D009842:Oligopeptides; D019151:Peptide Library; D010975:Platelet Aggregation Inhibitors; C547383:tyrosyl-aspartyl-prolyl-tryptophyl-threonyl-prolyl-serine; D014841:von Willebrand Factor; D003094:Collagen