Background/Purpose: Diagnostic biomarkers are decision-making tools in clinical lab routine and are of growing importance for clinical management of patients. In autoimmune diseases, one important class of biomarkers are autoantibodies (AAB) directed against human autoantigens. Apart from diagnostic antigens used in clinical routine, additional AAB reactivities to more than 100 human antigens relevant for disease are described in literature. Obviously, the autoimmune profile of humans covers a huge number of AABs, which display an enormous resource to identify novel marker candidates. Here, we describe a new screening platform SeroTag for discovery of novel AABs in healthy controls and autoimmune diseases. By selecting 100+ AABs a comprehensive autoimmune landscape will be outlined to define diseases and disease subgroups according to their intrinsic, highly differentiated AAB pattern.

Methods: SeroTag utilizes over 8,000 human proteins as antigen collection in bead-based suspension arrays (Luminex FlexMap 3D) to allow for high-throughput (HTS) serum sample processing with high accuracy, followed by standard and advanced data mining procedures. Recombinant antigens were covalently coupled to magnetic, color-coded beads and serum samples were incubated with several multiplex bead mixes each representing 400 different antigens. We screened over 6,000 serum samples from patients with autoimmune diseases such as SLE (n= >1,000), SSc (n= >450), RA (n= >1,500), SjS (n= >100) and over 1,000 healthy individuals to confirm known and to discover novel AABs.

Results: In SLE, SjS, SSc and RA, and also in healthy controls, novel autoantigens were discovered in several independent discovery studies and subsequently validated in separate validation studies. Antigens showing reproducible, significant reactivity compared to active and passive controls were selected in a stepwise marker refinement approach. Examples include BICD2 and KDM6B/JMJD3 as novel antigens in SSc with 20-30% prevalence, TMPO and MVP in SLE with 15-25% prevalence, and several novel protein targets of anti-citrullinated peptide antibodies.

Conclusion: HTS AAB screening is a valuable tool for “omics”-type biomarker discovery and verification in autoimmune diseases. Utilizing a technical platform capable to analyze thousands of antigens in thousands of patients, a high-resolution landscape of autoimmunity can be drawn. Over 50 novel autoantigens were discovered and validated in RA, SLE, SSc, and a set of novel autoantigens with high prevalence in healthy controls is presented. In combination with known AABs our findings show potential for improved and earlier diagnosis, differential diagnosis, and disease subgrouping. Distinct sets of AABs are utilized for biomarker support in the development of novel medicines for autoimmune diseases.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/high-throughput-screening-discovers-multiple-novel-autoantibodies-in-rheumatoid-arthritis-systemic-lupus-systemic-sclerosis-and-sjogrens-syndrome/