Background/Purpose: Rheumatoid arthritis (RA) is a systemic autoimmune disease associated with MHC polymorphisms. The shared epitope polymorphism in MHC class II genes is by far the strongest genetic association with ACPA⁺ RA, and the shared epitope polymorphisms have been demonstrated to facilitate the binding and presentation of citrullinated peptides. MHC class I gene polymorphisms are also associated with the susceptibility to seropositive RA. To further investigate the potential role of MHC class I polymorphisms in the pathogenesis of RA, we characterized CD8⁺ T cell phenotypes and clonality in ACPA⁺ RA blood using flow cytometric and single cell RNA and TCR sequencing analysis. We also performed in vitro assays to characterize the antigen specificity and functional properties of clonally expanded CD8⁺ T cells in ACPA⁺ RA blood.

Methods: We used flow cytometric and single-cell RNA profiling analyses including VDJ sequencing to analyze the effector phenotypes and clonal expansion of CD8⁺ T cells in ACPA⁺ RA blood. To investigate the antigen targets of the expanded CD8⁺ T cells in ACPA⁺ RA, we performed in vitro protein stimulation in ACPA⁺ RA peripheral blood mononuclear cells (PBMCs) or whole blood with candidate citrullinated RA antigens. To assess the cytotoxic potential of citrullinated antigen-activated cytotoxic of CD8⁺ T cells, we used an in vitro MHC-independent cytotoxicity assay.

Results: We identified GZMB⁺CD8⁺ subpopulations in ACPA⁺ RA blood containing large clonal lineage expansions expressing cytotoxic and tissue homing transcriptional programs, and a GZMK⁺CD8⁺ memory subpopulation comprised of smaller clonal expansions expressing effector T cell transcriptional programs. We demonstrated citrullinated RA autoantigens presented by MHC class I activate ACPA⁺ RA blood-derived GZMB⁺CD8⁺ T cells to expand, express cytotoxic mediators, and mediate killing of target cells. We also demonstrated that these activated GZMB⁺CD8⁺ T cells are present in RA synovium.

Conclusion: Our findings demonstrated that clonally expanded GZMB⁺ CD8⁺ T cells are present in ACPA⁺ RA blood. We further demonstrated that these clonally expanded cytotoxic CD8⁺ T cells are specific for citrullinated antigens and can mediate cytolytic cell killing. Our findings suggest that cytotoxic CD8⁺ T cells targeting citrullinated antigens could contribute to synovitis and joint tissue destruction in ACPA⁺ RA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/clonally-expanded-cytotoxic-cd8-t-cells-recognize-citrullinated-antigens-in-acpa-rheumatoid-arthritis/