title: Drug-selected complete restoration of superoxide generation in Epstein-Barr virus-transformed B cells from p47phox-deficient chronic granulomatous disease patients by using a bicistronic retrovirus vector encoding a human multi-drug resistance gene (MDR1) and the p47phox gene.
abstract: Chronic granulomatous disease (CGD) is a group of disorders characterized by the failure of phagocytes to produce superoxide. One-third of the cases of CGD in the USA and Europe results from defects in the gene encoding p47phox, a cytoplasmic component of NADPH oxidase for superoxide generation. In this study, we constructed the bicistronic retrovirus vector Ha-MDR-IRES-p47, which carries cDNAs for a human multi-drug-resistance gene (MDR1) and p47phox. The amphotropic retroviral producer cells were generated, and the supernatant of the producer cells was used to transduce Epstein-Barr virus-transformed B (EBV-B) cells, established from B cells of p47phox-deficient CGD patients, as an in vitro model of gene therapy for p47phox-deficient CGD. The transduced cells expressed both P-glycoprotein and p47phox protein, and the expression levels were increased after appropriate vincristine selection. The levels of superoxide production in the vincristine-selected cells were increased to a level similar to normal EBV-B cells. This result suggests that it is possible to achieve 100% correction of the CGD defect in p47phox-deficient EBV-B cells by using the bicistronic vector. This strategy could be employed not only in vitro, but also in vivo, in the gene therapy of a number of inherited diseases.
authors.affiliation: Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Iwata
authors.forename: H
authors.initials: H
authors.lastname: Nunoi
authors.forename: I
authors.initials: I
authors.lastname: Matsuda
authors.forename: S
authors.initials: S
authors.lastname: Kanegasaki
authors.forename: T
authors.initials: T
authors.lastname: Tsuruo
authors.forename: Y
authors.initials: Y
authors.lastname: Sugimoto
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D001402:B-Lymphocytes / Q000378:metabolism*; D002461:Cell Line, Transformed; D002469:Cell Separation; D004686:Encephalomyocarditis virus / Q000235:genetics; D005434:Flow Cytometry; D019450:Genes, MDR; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006105:Granulomatous Disease, Chronic / Q000235:genetics*; D004854:Herpesvirus 4, Human / Q000235:genetics; D006801:Humans; D019255:NADPH Oxidases; D010750:Phosphoproteins / Q000172:deficiency* / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*; D014750:Vincristine / Q000494:pharmacology
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D010750:Phosphoproteins; D011994:Recombinant Proteins; D013481:Superoxides; D014750:Vincristine; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1