Objectives Autoantibodies against post-translationally modified proteins (anti-modified protein antibodies or AMPAs) are a hallmark of rheumatoid arthritis (RA). A variety of classes of AMPAs against different modifications on proteins, such as citrullination, carbamylation and acetylation, have now been described in RA. At present, there is no conceptual framework explaining the concurrent presence or mutual relationship of different AMPA responses in RA. Here, we aimed to gain understanding of the co-occurrence of AMPA by postulating that the AMPA response shares a common ‘background’ that can evolve into different classes of AMPAs.
Methods Mice were immunised with modified antigens and analysed for AMPA responses. In addition, reactivity of AMPA purified from patients with RA towards differently modified antigens was determined.
Results Immunisation with carbamylated proteins induced AMPAs recognising carbamylated proteins and also acetylated proteins. Similarly, acetylated proteins generated (autoreactive) AMPAs against other modifications as well. Analysis of anti-citrullinated protein antibodies from patients with RA revealed that these also display reactivity to acetylated and carbamylated antigens. Similarly, anti-carbamylated protein antibodies showed cross-reactivity against all three post-translational modifications.
Conclusions Different AMPA responses can emerge from exposure to only a single type of modified protein. These findings indicate that different AMPA responses can originate from a common B-cell response that diversifies into multiple distinct AMPA responses and explain the presence of multiple AMPAs in RA, one of the hallmarks of the disease.
- rheumatoid arthritis
- anti-ccp antibodies
- post-translationally modified proteins
- anti-modified protein antibodies
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ASBK and JSD contributed equally.
Handling editor Prof Josef S Smolen
Contributors ASBK, JSD and RT have designed the experiments. ASBK and JSD have done the animal experiments. ASBK, JSD, MV and ALD have performed the ELISAs (murine and human). LH, ACK and MvD have performed the AMPA purification from patients with RA. GMCJ and PAvV have done the mass spectrometry analysis of the modified antigens. ASBK, JSD, MV, HB, TWJH, LAT, DvdW and RT were involved in critically revising the manuscript for intellectual improvement. ASBK, JSD, MV, ALD, TK, SR, LAT, DvdW and RT have been extensively involved in the interpretation and analysis of the results. All authors have contributed to the writing and editing of the manuscript.
Funding This work has been financially supported by the EU/EFPIA Innovative Medicines Initiative 2 Joint Undertaking RTCure grant no. 777357, by ReumaNederland (13-3-401) and by Target to B! (grant no. LSHM18055-SGF). In addition, this work is part of the research programme Investment Grant NWO Medium, project no. 91116004, which is (partly) financed by ZonMw. L.A.T. has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovationprogramme (grant agreement no. 724517).
Competing interests None declared.
Patient consent for publication Obtained.
Ethics approval All animal experiments were approved by the Ethical Committee for Animal Experimentation of the LUMC, Leiden. The study with human material was conducted with the approval of the regional ethics committee at Leiden University Medical Center.
Provenance and peer review Not commissioned; externally peer reviewed.
Data sharing statement All data relevant to the study are included in the article or uploaded as supplementary information.