Tu125 - Identification of Autoreactive Cytotoxic T Cells in Anca-associated Vasculitis

Laura van Dam – Stanford University; Shady Younis – Stanford University; Jae-Seung Moon – Stanford University; Shima Parsafar – Stanford University; Audra Horomanski – Stanford University; Orr Sharpe – Stanford University; Jolijn van Leeuwen – Stanford University; Cees van Kooten – Leiden University Medical Center; Y.K. Onno Teng – Leiden University Medical Center; William Robinson – Stanford University

Abstract Text: Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a rare and severe autoimmune disease, characterized by a pauci-immune necrotizing vasculitis. ANCAs targeting proteinase-3(PR3) or myeloperoxidase(MPO) and ANCA-producing B-cells play a central role in the pathogenesis. Autoreactive T-cells are less well-studied, but recent studies suggest their importance in the AAV pathogenesis. Here, we aimed to investigate autoreactive T-cells and their cytotoxic responses. AAV or healthy control (HC) PBMCs were cultured with ANCA antigens and anti-CD28 and analyzed with flow cytometry after 24h. In addition, T-cells were analyzed with single cell RNA sequencing. Anti-PR3+-AAV patients exhibited increased EMRA CD8+ T-cells and GZMB+CD8+ T-cells, whereas anti-MPO+-AAV patients had increased levels of GZMB+IFNy+CD4+ and GZMB+IFNy+CD8+ T-cells at baseline. MPO specifically induced expansion and proliferation of GZMK+IFNy+CD4+ and GZMK+IFNy+CD8+ T-cells in anti-MPO+-AAV patients. Single cell RNA sequencing demonstrated large populations of clonally expanded CD8+ T-cells in the GZMB+ and GZMK+CD8+T-cell compartment in AAV patients, which was not observed in HCs. The clonally expanded CD8+T-cells were characterized by expression of CD57, CD244 and CD314. Specifically, GZMB+CD8+T-cells expressed GPR56 and CX3CR1, whereas GZMK+CD8+ T-cells expressed CD56, CD161 and CD183. Moreover, both GZMB+ and GZMK+CD8+ T-cells of AAV patients displayed effector cytotoxic transcriptional programs, including different granzymes, IFN-y, perforin, CCL5, KLRK1 and NKG7, which was more abundant than in HCs.
This study demonstrated that clonally expanded cytotoxic CD8+T-cells are present in AAV patients, and we identified an autoreactive GZMK+IFNy+CD8+ subpopulation in anti-MPO+-AAV patients. The identification of pathogenic autoreactive T-cells could provide novel insights into next-generation therapeutic targets for AAV.