Many αIIbβ3 autoepitopes in chronic immune thrombocytopenic purpura are localized to αIIb between amino acids L1 and Q459

In chronic immune thrombocytopenic purpura (ITP), autoantibodies bind to platelet surface proteins, particularly alpha(IIb) , resulting in platelet destruction by the reticulo-endothelial system. In order to better localize the autoepitopes on alpha(IIb) , we studied the binding of antibodies to Chinese hamster ovary (CHO) cells expressing either alpha(IIb) beta(3) or alpha(IIb) -alpha(v) beta(3) chimaeras in which a segment of alpha(IIb) (either amino acids L1-Q459, L1-F223 or F223-Q459) was substituted for that portion of alpha(v) . We evaluated platelet-associated autoantibodies from 14 ITP patients with alpha(IIb) -dependent antibodies. Ten of 14 bound to alpha(IIb (L1-Q459)) -alpha(v) beta(3) , showing that autoepitopes were often localized to this region of alpha(IIb) . In addition, each of the autoantibodies binding to alpha(IIb (L1-Q459)) -alpha(v) beta(3) , also bound to CHO cells expressing either alpha(IIb(L1-F223)) -alpha(v) beta(3) or alpha(IIb(F223-Q459)) -alpha(v) beta(3) ). In two of the three eluates tested, > 95% of the autoantibody binding to alpha(IIb) could be adsorbed using CHO cells expressing any of the three chimaeras, showing that the epitope(s) have contact points on either side of amino acid F223; in the third eluate, only a portion (similar to40%) could be adsorbed by the chimaeric cell lines showing that, in this patient, an additional antibody was also present, directed to a site distal to amino acid Q459. The remaining four eluates bound to CHO cells expressing alpha(IIb) beta(3) but to none of the chimaeras, suggesting that these epitopes are also distal to amino acid Q459. We conclude that the binding of many anti-alpha(IIb) beta(3) autoantibodies is dependent on the presence of alpha(IIb) amino acids L1-Q459.