RECOMBINANT HUMAN NATURAL AUTOANTIBODIES AGAINST GPIIB IIIA INHIBIT BINDING OF AUTOANTIBODIES FROM PATIENTS WITH AITP/

Autoimmune thrombocytopenic purpura (AITP) is caused by autoantibodies predominantly against platelet membrane glycoproteins (GP) IIb/IIIa a nd GPIb/IX. Naturally occurring autoantibodies have been described aga inst a variety of autoantigens; it has been suggested that perturbatio n of their regulation may be associated with autoimmune diseases. Usin g a combinatorial Fab phagemid library from an individual immunized wi th human RhD+ red blood cells, we evaluated the presence of natural an ti-GPIIb/IIIa autoantibodies as well as their relation to AITP-associa ted anti-GPIIb/IIIa autoantibodies. Selection on native GPIIb/IIIa and characterization of positive clones by inhibition studies against mur ine monoclonal anti-GPIIb/IIIa antibodies and by DNA analysis revealed the presence of two distinct recombinant anti-GPIIb/IIIa autoantibodi es, which partially inhibited binding of affinity-purified platelet-as sociated autoantibodies from 8/12 AITP patients. Our results demonstra ted that GPIIb/IIIa-specific Fab directed against conformational epito pes within the GPIIb/IIIa complex may be cloned from the genome of an individual immunized with RhD+ red blood cells, who was not affected b y AITP. The partial inhibition of binding of platelet-associated autoa ntibodies from AITP patients to GPIIb/IIIa by the recombinant anti-GPI Ib/IIIa phage clones suggests recognition of closely related antigenic epitopes. These phage clones may represent down-regulated, potentiall y pathological autoantibodies and could be used as new tools for inves tigation of AITP.