Studies on the role of anti-citrullinated protein antibodies in rheumatoid arthritis

Anti-citrullinated protein antibodies (ACPA) are highly specific for rheumatoid arthritis (RA) and present in about two thirds of all patients at diagnosis. They can be detected already before disease onset and have direct pathogenic effect mediated partly through the Fc (fragment crystalizable) portion with attached Fc-glycan structure. On these grounds we aimed to further characterize ACPA’s role in the pathogenesis of RA both as a risk factor and a disease biomarker. To this end we investigated ACPA occurrence in a population-derived twin cohort and ACPA in a cohort of early-untreated RA patients in relation to disease outcomes. First we screened a large population-derived twin cohort (N= 12,590; median age 64, range 48-93 years) for occurrence of ACPA using an ACPA test used in clinical routine: the AntiCCP2 (IgG) test. Through linking the twin cohort with the Swedish National Patient Register we identified ACPA-positive individuals without RA (N=226) and ACPA-positive patients with RA (N=124). ACPA-positive individuals without RA had lower ACPA concentration and fewer different ACPA reactivities as compared to patients with ACPA-positive RA. Heritability estimates for having ACPA with or without RA were generally lower than expected (10% (95% CI: 0-43) for ACPA without RA; 23% (95% CI: 0-45) for ACPA; and 41% (95% CI: 0-74) for ACPA-positive RA). Heavy smoking and HLA-SE associated with ACPA occurrence with and without RA. Environmental factors (including smoking) appeared to be more important than genetic in determining which individuals develop ACPA, while genetic factors (and in particular HLA-SE) had a relatively larger impact in determining which ACPA-positive individuals that will ultimately develop arthritis. We also confirmed that presence of ACPA and especially high titers of ACPA have a high diagnostic accuracy for RA in a population based setting. Following this, we then screened a cohort of early-untreated RA patients (N=183) for occurrence of ACPA using either the Anti-CCP2 test or ELISA for detection of reactivities against specific citrullinated peptides. We demonstrated that ACPA (and especially anticitrullinated-vimentin antibodies) associated with markers of bone loss (as measured by ELISA detection of serum RANKL and/or presence of bone erosions on radiographs of hands and feet). Treatment with methotrexate (MTX) significantly lowered both ACPA and RANKL serum levels. In a subgroup of these patients (N=59) we investigated the Fcglycosylation patterns of serum IgG in relation to disease outcome further. A general low abundance of galactosylated glycans, partially restored by MTX treatment, was observed in the serum of early-untreated RA samples. This was more evident among future nonresponders as compared to responders to MTX treatment. The galactosylation status of the IgG-Fc had good predictive value for MTX response. In conclusion we showed that environmental as well as genetic factors are important for ACPA occurrence, which in turn has a high diagnostic accuracy for RA. In early-untreated RA, ACPA associate with bone loss and is modulated by methotrexate treatment. Further, Fcglycosylation patterns of antibodies are generally altered in early-untreated RA and might serve as a predictive factor for therapeutic response. LIST OF SCIENTIFIC PAPERS I. Environmental and genetic factors in the development of anticitrullinated protein antibodies (ACPAs) and ACPA-positive rheumatoid arthritis: an epidemiological investigation in twins A. Haj Hensvold, P. K. E. Magnusson, V. Joshua, M. Hansson, L. Israelsson, R. Ferreira, P-J. Jakobsson, R. Holmdahl, L. Hammarström, V. Malmström, J. Askling, L. Klareskog, A. I. Catrina Annuals of Rheumatic Diseases, 2015;74:375-380 II. How well do ACPA discriminate and predict RA in the general population – a study based on 12,590 population-representative Swedish twins A. Haj Hensvold, T. Frisell, P. K. E. Magnusson, R. Holmdahl, J. Askling, A. I. Catrina Recommended for publication, Annuals of Rheumatic Diseases III. Serum RANKL levels associate with anti-citrullinated protein antibodies in early untreated rheumatoid arthritis and are modulated following methotrexate A. Haj Hensvold, V. Joshua, W. Li, M. Larkin, F. Qureshi, L. Israelsson, L. Padyukov, K. Lundberg, N. Defranoux, S. Saevarsdottir, A. I. Catrina Arthritis Res. Ther. 2015;17:239 IV. IgG Fc galactosylation changes and predicts response to methotrexate in early rheumatoid arthritis S. L. Lundström*, A. Haj Hensvold*, D. Rutishauser, L. Klareskog, R. A. Zubarev, A. J. Ytterberg, A. I. Catrina Manuscript. *Equal contribution CONTENTS