Antigenic binding sites of anti‐protein S autoantibodies in patients with recurrent pregnancy loss

ESSENTIALS: Anti‐protein S autoantibodies (anti‐PS) may lead to a PS deficiency and recurrent pregnancy loss. We performed epitope mapping of anti‐PS in patients with recurrent pregnancy loss. Anti‐PS recognized epidermal growth factor (EGF)‐like domains in PS and recombinant human EGF. Anti‐PS may be associated with not only thrombophilia but also the disruption of the EGF system. BACKGROUND: Protein S (PS) deficiency is a risk factor for adverse pregnancy outcomes including recurrent pregnancy loss. Several studies have shown that the presence of anti‐PS autoantibodies (anti‐PS) leads to an acquired PS deficiency. Hence, an epitope mapping study was conducted to know the pathogenesis of anti‐PS in patients with recurrent pregnancy loss. METHODS: PS was treated with thrombin to divide the protein into γ‐carboxyglutamic acid (Gla) domain and Gla‐domain free PS. For the preparation of fragments of epidermal growth factor (EGF)‐like domains (EGF1‐4), PS was subjected to proteolysis using lysyl endopeptidase. The epitopes were identified in immunoblot. Whether anti‐PS recognized EGF family proteins in anti‐PS‐positive patients was also examined. RESULTS: Anti‐PS recognized Gla‐domain free PS, especially the three fragments of EGF‐like domains, EGF1‐2, EGF3‐4, and EGF1‐4. Anti‐PS recognized recombinant human EGF. Anti‐PS and polyclonal antibodies to recombinant human EGF recognized PS in the absence of Ca(2+) but not in the presence of Ca(2+). In competitive inhibition studies, polyclonal antibodies to recombinant mouse EGF blocked anti‐PS binding to PS in a concentration‐dependent manner. CONCLUSIONS: These results suggest that anti‐PS in patients with recurrent pregnancy loss recognize EGF‐like domains in PS. Interestingly, anti‐PS also recognized EGF family proteins. Anti‐PS in patients with recurrent pregnancy loss may be associated with not only thrombophilia but also the disruption of the EGF system.