Murine anti type VII collagen, complement fixing recombinant monoclonal IgG2b antibodies to induce inflammatory experimental epidermolysis bullosa acquisita in mice
SCHMIDT-JIMÉNEZ L. 1, THEOCHARIS T. 2, KROHN S. 2, BIEBER K. 1, LUDWIG R. 1,3, PEIPP M. 2, VERSCHOOR A. 3
1 Lübeck Institute for Experimental Dermatology, University of Lübeck, Lübeck, Germany; 2 Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig- Holstein, Kiel, Germany; 3 Department of Dermatology, Allergology and Venerology, University of Lübeck, Lübeck, Germany
Epidermolysis bullosa acquisita (EBA) is a prototypic autoantibody-mediated disease in which autoantibodies target the dermal-epidermal junction (DEJ) anchoring fibril type VII collagen (COL7) and mediate tissue inflammation and destruction. The complement system was shown to play a crucial role in the pathogenesis of EBA: C3 deposition at the DEJ serves as diagnostic hallmark of disease and preclinical models demonstrated a critical role of the C5a/C5aR-axis in autoantibody-mediated tissue damage in EBA.
Current antibody transfer models of EBA carry limitation due to the xenogenic nature of the transferred (auto)antibodies. To establish a wholly murine antibody transfer model of EBA, we used phage display selection of two immune libraries - generated from the spleen and inguinal lymph nodes of a mouse that developed active EBA upon immunization with murine COL7- to identify and generate a panel of autoantibodies targeting mCOL7.
We thoroughly characterized six distinct mCOL7 binding antibody clones from our selection, which were cloned into the complement fixing IgG2b backbone, as this represents the pre-dominant antibody isotype in sera from immunized mice with active EBA. All clones bound to recombinant mCOL7 and to the DEJ of murine skin sections. 4/6 of the clones displayed higher individual binding avidity than the overall polyclonal anti-mCOL7 serum pool. All tested clones were able to induce reactive oxygen species release from neutrophils (representing the dominant immune cell population driving EBA) upon binding to mCOL7. Combined, our in vitro findings suggest that we succeeded in generating murine autoantibodies that should also display EBA-related pathogenicity in vivo.