Background Rheumatoid Factors (RF) and autoantibodies against citrullinated proteins (ACPA) are part of the diagnosis of rheumatoid arthritis (RA). Unfortunately these analytes lack sensitivity especially in early phases of RA. By way of screening proteins from a human protein library of over 7,000 proteins we discovered over 20 novel citrullinated antigens targeted by ACPAs in CCP positive as well as in 5–10% of CCP negative RA patients suggesting added diagnostic potential.
Objectives To challenge novel human proteins as biomarkers for early RA which exhibited reactivities in citrullinated format against early and late RA patient serum samples we set to analyze sufficiently large, new cohorts of patients with early and established RA against the candidate proteins both in their citrullinated and non-citrullinated form. We also planned to veify single antigen performance by comparison to established benchmark proteins and marketed assays.
Methods 34 novel citrullinated antigens were analyzed in serum samples from independent cohorts of 300 CCP positive and 151 CCP negative patients with early RA, 100 healthy volunteers and 50 patients with established RA. All samples were analyzed in a multiplexed, bead-based array on Luminex FlexMAP3D, compared to reactivities against known citrullinated proteins (vimentin, fibrinogen, enolase) and subsequently in 3 commercially available anti-CCP-assays.
Results We confirmed novel citrullinated antigens targeted by ACPAs as markers of RA. The citrullinated antigens are highly reactive compared to their non-citrullinated forms in mature RA and in CCP positive samples of patients with early RA. 4 single antigens reach high overlap to established CCP-Assays each with >75% sensitivity at 97% specificity. Increased reactivity of novel antigens was identified in up to 10% of CCP negative samples suggesting the presence of additional citrullinated epitopes not present in commercial CCP assays used in this study.
Conclusions Novel citrullinated antigens discovered by large scale “omics” screening via Luminex bead based arrays were confirmed in novel samples of early and established RA, especially in CCP positive samples. Reactivity in 10% CCP negative samples suggests added diagnostic value.
Disclosure of Interest P. Schulz-Knappe Employee of: Protagen AG, P. Budde Employee of: Protagen AG, A. Lueking Employee of: Protagen AG, P. Schriek Employee of: Protagen AG, H. Goehler Employee of: Protagen AG, H.-D. Zucht Employee of: Protagen AG, J. Detert: None declared, G.-R. Burmester: None declared, M. Schneider: None declared, J. Richter: None declared, S. Vordenbäumen: None declared
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