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Heterogenous inhibition of platelet aggregation by monoclonal antibodies binding to multiple sites on GPIIIa
Author(s) -
Lid L.X.,
Nardi M. A.,
Nierodzik M. L.,
Karpatkin S.
Publication year - 1995
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1995.tb05422.x
Subject(s) - platelet , chemistry , epitope , monoclonal antibody , thrombin , microbiology and biotechnology , biochemistry , tyrosine , fibrinogen , binding site , antibody , platelet membrane glycoprotein , glycoprotein , biology , immunology
Summary. Six monoclonal IgGl‐k antibodies (LK2, LK3r, LK4‐55, LK5, LK6‐55, LK7r) were raised against platelet membrane GPIIIa in order to study the structure‐function relationship of this molecule. Antibodies were selected on their ability to react with GPIIIa by ELISA on adherent platelets, by immunoblot on platelet lysates and by fluorescence flow cytometry on intact platelets. Fluorescence reactivity varied from 3‐ to 202‐fold greater than isotype control fluorescence. Two MoAbs reacted on immunoblot under reduced conditions (LK7r and LK3r). Two reacted with a 55 kD chymotrypsin/subtilisin digest of GPIIIa which is likely to exclude amino acids 121‐348 (LK4‐55 and LK6‐55). Four of the MoAbs (LK5, LK3r, LK2 and LK4‐55) inhibited tyrosine phosphorylation of one to four distinct bands on immunoblot. LK4‐55 reacted with an N‐terminal 66 amino acid fusion protein of GPIIIa near the PLA epitope (Leu 33). LK7r reacted with a 212‐222 peptide reported to be an RGD fibrinogen binding site. LK2 reacted near a disintegrin‐RGD binding site. Except for LK5, all inhibited ADP, collagen and thrombin‐induced platelet aggregation in a heterogenous fashion. Percentage inhibition of 125 I‐fibrinogen binding to platelets varied from 18% to 98%. No correlation was noted between inhibition of fibrinogen binding, location of MoAb binding on GPIIIa, reactivity of MoAb binding with GPIIIa, inhibition of thrombin‐induced tyrosine phosphorylation or inhibition of platelet aggregation induced by ADP, collagen or thrombin. Thus MoAbs, binding to platelet GPIIIa at different sites, inhibit platelet aggregation in a heterogenous manner.