Carbamylation of β2-GPI generates new autoantigens for antiphospholipid syndrome. a new tool for diagnosis of "seronegative" patients

Objectives: Antiphospholipid syndrome (APS) is a prothrombotic condition defined by recurrent thrombosis, pregnancy complications and circulating antiphospholipid antibodies (aPL), including anti-β2-Glycoprotein I (β2-GPI). In clinical practice it is possible to find patients with APS persistently negative for the aPL tests according to Sydney criteria ("seronegative APS", SN-APS). Recently, several autoimmune responses have been described as a consequence of post-translational modifications of their target autoantigens. This study was undertaken to test carbamylated-β2-GPI (Carb-β2-GPI) as a new autoantigen of APS. Methods: β2-GPI was carbamylated by potassium cyanate and used to investigate its effect on monocyte-derived DC (moDC) phenotype and function. Sera from 114 SN-APS patients, 60 APS, 20 patients with Rheumatoid Arthritis, 20 NON-APS thrombosis and 50 healthy donors were analyzed for anti-Carb-β2-GPI by ELISA. Results: Carb-β2-GPI is able to activate moDCs, inducing up-regulation of CD80, CD86, and CD40, activation of ERK, p38 MAPK and NF-κB and IL-12p70 release.Serological results showed that both 37/114 SN-APS (32.45%) and 23/60 APS patients (38.33%) resulted positive for anti-Carb-β2-GPI. Interestingly, SN-APS patients tested positive for anti-Carb-β2-GPI showed a higher prevalence of thrombocytopenia (p= 0.04, likelihood positive ratio of 3.9). Conclusion: Data obtained from both functional tests on moDCs and from immunologic approaches prompted identification of Carb-β2-GPI as a "new" antigenic target in APS. In particular, anti-Carb-β2-GPI revealed a potential usefulness in identification of a significant proportion of SN-APS patients. Moreover, since patients tested positive for anti-Carb-β2-GPI reported a high risk of thrombocytopenia, this test may be considered a suitable approach in the clinical evaluation of SN-APS.