Immunological Interaction of HLA-DPB1 and Proteinase 3 in ANCA Vasculitis is Associated with Clinical Disease Activity

Significance Statement In a longitudinal, prospective cohort study, we observed that patients with PR3-ANCA vasculitis and HLA-DPB1*04:01 are more likely to experience disease flares, which informed our hypothesis that HLA has an immunopathogenic role. We found that an epitope of PR3 (PR3 225-239 ) has high affinity for HLA-DPB1*04:01. By examining patient peripheral blood mononuclear cells, we demonstrated that PR3 225-239 presentation by HLA-DPB1*04:01 stimulates PR3 225-239 –specific autoreactive T cells. This may explain the associated increased relapse risk. However, in patients who are in long-term remission off therapy, HLA-DPB1 + cells bind PR3 225-239 at levels seen in healthy controls. The diminished interaction between HLA-DPB1 and autoantigen in long-term remission signals immunological nonresponsiveness, creating a foundation to define immunological remission. Background PR3-ANCA vasculitis has a genetic association with HLA-DPB1. We explored immunologic and clinical features related to the interaction of HLA-DPB1*04:01 with a strongly binding PR3 peptide epitope (PR3 225–239 ). Methods Patients with ANCA vasculitis with active disease and disease in remission were followed longitudinally. Peripheral blood mononuclear cells from patients and healthy controls with HLA-DPB1*04:01 were tested for HLA-DPB1*04:01 expression and interaction with a PR3 peptide identified via in silico and in vitro assays. Tetramers (HLA/peptide multimers) identified autoreactive T cells in vitro. Results The HLA-DPB1*04:01 genotype was associated with risk of relapse in PR3-ANCA (HR for relapse 2.06; 95% CI, 1.01 to 4.20) but not in myeloperoxidase (MPO)-ANCA or the combined cohort. In silico predictions of HLA and PR3 peptide interactions demonstrated strong affinity between ATRLFPDFFTRVALY (PR3 225–239 ) and HLA-DPB1*04:01 that was confirmed by in vitro competitive binding studies. The interaction was tested in ex vivo flow cytometry studies of labeled peptide and HLA-DPB1*04:01-expressing cells. We demonstrated PR3 225–239 specific autoreactive T cells using synthetic HLA multimers (tetramers). Patients in long-term remission off therapy had autoantigenic peptide and HLA interaction comparable to that of healthy volunteers. Conclusions The risk allele HLA-DPB1*04:01 has been associated with PR3-ANCA, but its immunopathologic role was unclear. These studies demonstrate that HLA-DPB1*04:01 and PR3 225–239 initiate an immune response. Autoreactive T cells specifically recognized PR3 225–239 presented by HLA-DPB1*04:01. Although larger studies should validate these findings, the pathobiology may explain the observed increased risk of relapse in our cohort. Moreover, lack of HLA and autoantigen interaction observed during long-term remission signals immunologic nonresponsiveness.