An animal model for granulomatosis with polyangiitis

Proteinase 3 (PR3) is a serine protease of neutrophil granules and a major target of autoantibodies in Granulomatosis with PolyAngiitis (GPA). The disease starts in the respiratory tract and progresses to a generalized life-threatening vasculitis of small blood vessels in multiple essential organs. Anti-neutrophil cytoplasmic antibodies (ANCA) against PR3 are a laboratory biomarker for the diagnosis of GPA and are believed to be involved in disease progression. Due to the lack of interaction of hPR3 antibodies with murine PR3, wild type animals cannot be used for ANCA transfers in mice. Thus a human PR3 knock-in mouse line expressing the human autoantigen in peripheral blood neutrophils was developed enabling both the transfer of ANCA and mouse monoclonal antibodies (mAb) to human PR3. To confirm the expression and functionality of hPR3 in the knock-in mice, neutrophils were isolated from bone marrow and analyzed by ELISA, activity assays and flow cytometry. Comparison of the expression of hPR3 in knock-in mice with murine PR3 in wild type mice revealed that the expression levels were similar in both lines. Furthermore we found that the hPR3 expressed from the endogenous mouse PR3 promoter was active. Upon activation of freshly isolated neutrophils, surface expression of hPR3 could be confirmed. The newly generated hPR3 knock-in mouse line is able to store functional hPR3 in neutrophils. Upon activation hPR3 is expressed on the surface of these cells, making the hPR3 accessible to antibodies. Thus antibody binding to the neutrophil surface could lead to neutrophil activation and inflammatory burst. To investigate the pathogenicity of autoantibodies in GPA, transfer of mAb will give us first insights into the pathogenicity these.