Major histocompatibility complex-peptidome in granulomatosis with polyangiitis

Granulomatosis with polyangiitis (GPA) is a potentially life-threatening chronic inflammatory autoimmune disease. The disease is characterized by systemic necrotizing vasculitis and ex-travascular granulomatous inflammation. It is associated with highly specific autoantibodies against proteinase 3 (PR3-ANCA). Our own previous studies suggest an autoantigen- and pathogen-driven pathogenesis of GPA. The aim of the investigations applied for in this grant application is the identification of antigenic triggers of GPA by analyzing the MHC-peptidome, i.e. peptides that are presented to the cells of the adaptive immune system via the major his-tocompatibility complex (MHC) on the cell surface of mononuclear and polymorphonuclear cells from the peripheral blood. Using bioinformatic algorithms, peptides of the MHC-peptidome from patients will be assigned to their source proteins, e.g. to the autoantigen PR3 and other endogenous proteins as well as pathogen-derived proteins, and differences in the MHC-peptidome between patients and healthy persons will be analyzed. In addition, peptides of the GPA-specific MHC-peptidome signature will be tested for their immunogenicity, i.e. the ability to trigger an immune response after stimulating T-cells with these peptides. Analysis of the MHC-peptidome is performed by means of peptide elution from purified MHC-class I and II complexes and subsequent analysis of the eluted peptides by means of liquid chromatog-raphy-coupled tandem mass spectrometry (LC-MS/MS). T-cell responses following peptide stimulation are screened using ELISpot, and reactive T-cells will be phenotypically and func-tionally characterized using flow cytometry. Analysis of the MHC-peptidome will for the first time enable the identification of peptide sequences presented in vivo to the adaptive immune system in GPA. These investigations will lead to the development of future targeted therapeu-tic interventions at the level of the antigen-triggered T-cell response.