Drug-selected complete restoration of superoxide generation in Epstein-Barr virus-transformed B cells from p47(phox)-deficient chronic granulomatous disease patients by using a bicistronic retrovirus vector encoding a human multi-drug resistance gene (MDR1) and the p47(phox) gene

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 p47(phox), 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 p47(phox). 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 p47(phox)-deficient CGD patients, as an in vitro model of gene therapy for p47(phox)-deficient CGD. The transduced cells expressed both P-glycoprotein and p47(phox) 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 p47(phox)-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.