Structural basis of HPA-1a alloimmunization in FNAIT and allosteric regulation of integrin conformation.

abstractFetal and neonatal alloimmune thrombocytopenia (FNAIT) can be caused by maternal immunoglobulin G alloantibodies against the human platelet antigen 1a (HPA-1a), a Leu33Pro polymorphism at the plexin-semaphorin-integrin (PSI) domain of the β3 subunit shared by integrins αIIbβ3 and αVβ3. Although serologic detection of anti-HPA-1a alloantibodies has long aided FNAIT diagnosis, the disease remains a major clinical challenge due to unpredictable severe outcomes, particularly intracranial hemorrhage and fetal loss. To define the molecular basis of HPA-1a immunogenicity and antibody pathogenicity, we determined crystal structures of the HPA-1a antigen bound to Fabs from 4 anti-HPA-1a monoclonal antibodies, including 3 clinically relevant maternal alloantibodies (26.4, D-204, and M-204) and 1 murine antibody (SZ21). The structures reveal that all antibodies recognize an HPA-1a epitope centered on Leu33, spanning both PSI and integrin-epidermal growth factor (I-EGF) 1 domains of β3 integrin. Variations in antibody-binding interfaces account for differential affinities to both αIIbβ3 and αVβ3, whereas distinct binding orientations determine whether antibody engagement stabilizes the bent inactive β3 conformation. Functional assays demonstrated antibody-dependent inhibition of αIIbβ3-mediated platelet aggregation, thrombus formation, and clot retraction, as well as αVβ3-mediated endothelial adhesion and spreading. These results establish the structural framework of HPA-1a alloantigenicity, explain the conformational selectivity of anti-HPA-1a antibodies, and elucidate how they block integrin activation. The findings provide mechanistic insight into FNAIT pathogenesis and suggest that antibody heterogeneity in affinity and function may contribute to clinical variability. Furthermore, these findings demonstrate antibody-mediated stabilization of the PSI/I-EGF1 domain as a potential strategy for allosteric modulation of integrin structure and function.