Physiological target, molecular evolution and pathogenic functions of a monoclonal ACPA obtained from an RA patient.

OBJECTIVES We previously isolated a human monoclonal anti-citrullinated protein/peptide antibody (ACPA), CCP-Ab1, which recognizes various citrullinated antigens, from a patient with rheumatoid arthritis (RA). In this study, we aimed to explore the physiological target for CCP-Ab1 and the role of molecular evolution, through affinity maturation of CCP-Ab1, in the onset and the exacerbation of RA. METHODS We identified and purified the target protein of CCP-Ab1 in plasma, under native conditions. Then, we generated germline-reverted CCP-Ab1 (GL-rev CCP-Ab1) and compared its reactivity to mature CCP-Ab1. Finally, we analyzed the function of CCP-Ab1 and GL-rev CCP-Ab1 in the onset or the exacerbation of autoimmune arthritis using autoimmune arthritis-prone SKG mice. RESULTS CCP-Ab1 bound citrullinated fibrinogen under native conditions. GL-rev CCP-Ab1 drastically reduced the binding affinity to citrullinated fibrinogen (p < 0.05). Also, the elements implicated in GL-rev CCP-Ab1 binding to citrullinated-peptide, cfc1-cyc were almost identical to those bound by CCP-Ab1. CCP-Ab1, but not GL-rev CCP-Ab1, induced significant exacerbation of experimental arthritis when administered into SKG mice (p < 0.05). IL-6 production, in both joint tissue and serum, was observed in the CCP-Ab1-treated SKG mice but not in the GL-rev CCP-Ab1-treated group (p < 0.05). Furthermore, the immune complex formed by CCP-Ab1 and fibrinogen was detected in the synovial tissues in CCP-Ab1-administered mice (p < 0.05). CONCLUSIONS Our data show that germline-encoded CCP-Ab1, which binds weakly to citrullinated fibrinogen, undergoes hypermutation through activation of naive B cells by citrullinated peptides/proteins, stimulating high reactivity to citrullinated fibrinogen. These findings deepen our understanding of the role of molecular evolution of ACPAs in the onset/exacerbation of RA.