Characterizing the proteome of pathogenic IgA1-containing immune complexes in IgA Nephropathy

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Project Abstract IgA Nephropathy (IgAN) is an autoimmune glomerulonephritis that frequently results in kidney failure. IgAN is characterized by elevated production of galactose-deficient immunoglobulin A1 (Gd-IgA1) being bound by an immunoglobulin G (IgG) autoantibody specific for Gd-IgA1 to form IgA1-associated immune complexes (IgA1- ICs) with additional proteins in blood. These complexes deposit in the glomerulus and cause kidney injury by activating mesangial cell proliferation and breakdown of glomerular capillaries allowing protein or blood in the urine. There is no worldwide screening and can only be diagnosed by renal biopsy. We know that the immunoglobulins and the undefined blood proteins are necessary to the nephritogenic activity of the high molecular weight IgA1-ICs, but we do not know what concentration, and variability of each protein nor the ratio of immunoglobulins necessary for complex formation. The primary goal of my study is to quantify and establish the ratios of immunoglobulins and blood proteins in the composition of IgA1-ICs in the serum of patients with IgAN of opposing clinical spectrum in comparison to healthy controls. Our collaborative team has observed that the addition of serum to the immunoglobulin nucleating factors is necessary for the formation of pathogenic ICs that activate human cultured mesangial cells. The overarching goal of this proposal is that quantitative analysis of serum IgA1-ICs will determine stoichiometry of immunoglobulins and other blood proteins involved in the formation of IgA1-ICs in both progressing and stable disease patients with IgAN. In my recently submitted manuscript, I identified 21 high-quality blood protein targets specifically enriched in IgA1-ICs isolated from IgAN patient serum. When comparing the identification of proteins and their corresponding enrichment in the IgA1-ICs to the other uncomplexed forms of IgA1 control fractions (monomeric and polymeric IgA), we observed that each fraction clustered by molecular form, showing the value of our sample prep and innovative fractionation of serum. Based on preliminary data, I hypothesize the ratio of IgA1 to IgG autoantibody is discreet and can be determined by targeted LC-MS quantitative analysis, polymeric IgA1 is the predominant molecular form of IgA1 involved in complex formation and complement cascade activation factors are present in complex composition at a higher ratio than immunoglobulins before deposition into the kidney. Throughout the global pandemic, the world has learned of the importance of disease screening, the variability of disease presentation, and the daily fear that immunocompromised populations live in. Being a researcher in IgA Nephropathy, I have been aware of these realities in IgAN patient populations prior to COVID-19. This has fueled my dedication to answering important questions in the IgAN field and understanding this crucial part of pathogenesis of IgA1-IC formation for the sake of future development of less-invasive screening methods or therapies using blood biomarkers for this currently unpredictable disease.