Novel Approach to Oral Gene Therapy for Chronic Granulomatous Disease

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DESCRIPTION (provided by applicant): Chronic granulomatous disease (CGD) is an inherited primary immunodeficiency disorder characterized by severe recurrent infections by catalase positive bacteria and fungi. The molecular defects causing CGD result in the absence, low expression, or malfunction of one of the phagocyte NADPH oxidase components responsible for the generation of microbicidal reactive oxygen species. X-linked CGD results from mutations in the CYBB gene encoding the cytochrome b heavy chain glycoprotein, gp91-phox. The novel yeast cell wall particle (YCWP) DNA delivery technology traps DNA in the form of cationic polymer nano-complexes within porous cell wall "ghosts," providing for controlled release of the DNA upon particle internalization in macrophages. The micron-sized YCWP can be administered orally in order to deliver a therapeutic gene to peritoneal and gut wall macrophages, which then migrate to locations throughout the body. We hypothesize that this innovative approach will result in expression of the wild type gene encoding normal gp91-phox and correction of the functional defect in CGD phagocytes. Specifically, we propose to: 1. Test YCWP delivery and cDNA expression in an in vitro system using elicited peritoneal phagocytes from Cybb knock-out mice and transfection with cDNA encoding gp91-phox for transient replacement of gene function and correction of the CGD phenotype. Measurements will include fluorescence microscopy for particle uptake; molecular, histochemical and flow cytometric assays of gene expression and peroxide generation; and bacterial killing assays for evaluation of microbicidal function. 2. Test intraperitoneal and oral YCWP delivery for functional expression of cDNA encoding gp91-phox in an in vivo system using a murine Cybb knock-out model of CGD. Assays will include those used in Aim 1, as well as in vivo assays of host defense. The results of these studies, if successful, should demonstrate the feasibility of this novel gene therapy system for CGD and would provide a strong basis for large animal studies and translational research to bridge the gap from animal to human gene therapy. We hope that the proposed exploratory/developmental investigations will eventually lead to a safe, effective alternative method for gene therapy of CGD. PUBLIC HEALTH RELEVANCE: The proposed research, if successful, would demonstrate the feasibility of a novel oral gene therapy system for chronic granulomatous disease, an important primary immune deficiency disorder. The findings would provide a strong basis for translational research culminating in clinical trials for human gene therapy for the disease.