A monoclonal antibody recognizes a von Willebrand factor domain within the amino-terminal portion of the subunit that modulates the function of the glycoprotein IB- and IIB/IIIA-binding domains.
We developed a monoclonal antibody, 1C1E7, against vWf that increases ristocetin-induced platelet aggregation in a dose-dependent manner and lowers the threshold concentration of ristocetin needed to obtain a full aggregatory response. The platelet aggregatory effect of asialo vWf (ASvWf) also is enhanced by 1C1E7, in the presence or absence of glycoprotein (GP) IIb/IIIa receptor antagonism. In the presence of ristocetin, both intact 1C1E7 and its Fab fragments enhance specific binding of 125I-vWf to platelets. With 1C1E7, the intermediate and higher molecular weight multimers of vWf are preferentially bound to both GP Ib and GP IIb/IIIa. Thrombin-induced 125I-vWf binding to GP IIb/IIIa also is increased by 1C1E7. Maximal binding of 1C1E7 to vWf corresponds to 0.97 mol/mol vWf monomer with a Kd of 4.7 x 10(-10) M. 1C1E7 reacts with a 34/36-kD tryptic fragment (III-T4) and a 34-kD plasmic fragment (P34), which localizes the epitope between amino acid residues 1 and 272; this was confirmed by NH2-terminal amino acid sequencing. Finally, platelet aggregation by ASvWf was associated with a sharp rise in intracellular Ca2+ only in the presence of 1C1E7. An antibody-mediated conformational change of vWf may result in an improved presentation of the GP Ib- and GP IIb/IIIa-binding domains of mainly the larger multimers; the increased density of vWf on the platelet surface leads to platelet activation. The antibody may thus recognize a domain of relevance for vWf physiology.
Tornai
I
I
Center for Thrombosis and Vascular Research, University of Leuven, Belgium.
Arnout
J
J
Deckmyn
H
H
Peerlinck
K
K
Vermylen
J
J
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001665:Binding Sites; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000097:blood; D003094:Collagen / Q000378:metabolism; D010450:Endopeptidases; D006493:Heparin / Q000378:metabolism; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000302:isolation & purification; D010974:Platelet Aggregation / Q000187:drug effects; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011487:Protein Conformation; D012310:Ristocetin / Q000494:pharmacology; D014841:von Willebrand Factor / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D046911:Macromolecular Substances; D010446:Peptide Fragments; D010980:Platelet Membrane Glycoproteins; D014841:von Willebrand Factor; D012310:Ristocetin; D006493:Heparin; D003094:Collagen; D010450:Endopeptidases; D002118:Calcium