Immunoglobulin A nephropathy (IgAN) is the most common form of glomerulonephritis worldwide. The disease is characterized by progressive deposition of abnormal immune complexes of polyclonal IgA in the mesangia of the glomeruli, causing chronic inflammation and progressive damage to the kidneys, evolving into end-stage renal disease in up to 40% of cases, for which dialysis or transplantation is required. There are currently no effective treatments for incipient IgAN, and advanced disease is treated primarily with anti-inflammatory and anti-hypertensive regimens prior to dialysis or transplantation. However, several lines of evidence have indicated that reduction of IgA deposits in the kidney is typically associated with a clinical and pathological resolution of glomerulonephritis, and vice versa. Furthermore, removal of IgA complexes from the circulation by plasmapheresis or transplantation of an IgAN kidney into a non-IgAN recipient has lead to disappearance of kidney deposits, and restoration of kidney function. Thus, any treatment aimed at removing and/or preventing pathogenic IgA deposits in the kidneys could lead to substantial recovery of kidney function, and would therefore be highly desirable. One strategy has sought to exploit IgA-cleaving proteases, which are secreted by mucosal bacteria. The protease from H influenzae was effective in removing circulating and mesangial IgA deposits in mice injected with immune complexes containing human IgA1 in amounts comparable to the IgA1 concentration in the human circulation. Thus, such a strategy seems promising. However, a serious drawback of the bacterial protease is its potential immunogenicity. The risks of anaphylaxis and loss of efficacy will probably be unacceptable, since large doses will likely be required to deal with the abundance of IgA in the human circulation, and since IgAN patients will continue to produce "IgAN-ogenic" IgA, and will therefore need periodic IgA protease treatments for the rest of their lives. Thus, the ultimate success of this strategy will depend on engineering an IgA1-specific proteolytic activity on a human protease scaffold that is stable in the human circulation, which is the aim of the present proposal. To facilitate the engineering of target-cleaving activities on human protease scaffolds, a proprietary proteolytic activity sensor has been developed, which, when co-expressed in cells with mutagenic gene libraries of a protease scaffold, confers a selectable phenotype on cells expressing target-cleaving variants of the scaffold in quantitative proportion to their activities. We will use this system to engineer human IgA-cleaving activity int human complement factor D (hFD), a circulatory protease that is uniquely suited to serve as a scaffold for engineering proteases for human therapy. It is hoped that treatment with such a protease will facilitate the clearance of IgA immune complexes from the glomerular mesangium and allow the restoration of normal kidney function in IgAN patients.
Public Health Relevance Statement: Public Health Relevance: The goal of this project is to develop a novel effective treatment for Immunoglobulin A (IgA) nephropathy, the most common form of glomerulonephritis worldwide. The disease is characterized by progressive deposition of abnormal IgA immune complexes in the kidneys, causing chronic inflammation and progressive damage, and evolving into end-stage renal disease in up to 40% of cases, for which dialysis or transplantation is required. We will attempt to engineer a human protease that can facilitate the clearance of IgA immune complexes from the kidneys and allow the restoration of normal kidney function in IgAN patients without the need for dialysis or transplantation.
Project Terms: advanced disease; Affect; Anaphylaxis; Animal Model; Anti-inflammatory; Anti-Inflammatory Agents; Antigen-Antibody Complex; Antigens; Antihypertensive Agents; Bacteria; Blood Circulation; Cells; Chronic; chymotrypsin; Cleaved cell; Clinical; Clostridium difficile; Complement Factor B; Complement Factor D; Complex; Deposition; design; Dialysis procedure; Disease; Dose; effective therapy; End stage renal failure; Engineering; Enzymes; Epitopes; Escherichia coli; Excision; Exotoxins; Family; flu; Funding; Galium; Gene Library; Glomerulonephritis; Goals; Haemophilus influenzae; Human; Human Engineering; IgA1; Immune; immunogenic; immunogenicity; Immunoglobulin A; Immunoglobulin G; Inflammation; Injection of therapeutic agent; Kidney; Kidney Diseases; Lead; Libraries; Mediating; Modeling; Mus; Mutagenesis; novel; Patients; Peptide Hydrolases; periplasm; Phagocytosis; Phase; Phenotype; Plasmapheresis; Positioning Attribute; prevent; Process; public health relevance; Reagent; Recovery; Regimen; Renal Circulation; Renal function; research clinical testing; Resolution; response; Rest; restoration; Risk; scaffold; screening; sensor; Serine Protease; Specificity; Structure of glomerular mesangium; Substrate Specificity; success; System; Therapeutic; Therapeutic Uses; Time; Toxic effect; Transplantation; Validation; Variant; Work
