Enhanced host defense after gene transfer in the murine p47(phox)-deficient model of chronic granulomatous disease

The p47(Phox-/-) mouse exhibits a phenotype similar to that of human chronic granulomatous disease (CGD) and, thus, is an excellent model for the study of gene transfer technology. Using the Moloney murine leukemia virus-based retroviral vector MFG-S encoding the human form of p47(phox), We performed ex vivo gene transfer into Sca-1(+) p47(phox-/-) marrow progenitor cells without conditioning of donors with 5-fluorouracil. Transduced progenitors were transplanted into moderately irradiated (500 cGy), G-CSF preconditioned sibling p47(Phox-/-) mice. Using the fluorescent probe dihydrorhodamine 123 (DHR), in vivo biochemical correction of the superoxide-generating NADPH oxidase system was detected by flow cytometry in 12.3% +/- 0.9% of phorbol myristate acetate-stimulated peripheral blood neutrophils at 4 weeks and 2.6% +/- 1.0% at 14 weeks after transplantation. Following gene therapy, mice were challenged with the CGD pathogen Burkholderia (formerly Pseudomonas) cepacia and bacteremia levels were assessed at 24 hours and 7 days after inoculation, At both time points, bacteremia levels in gene corrected p47(phox-/-) mice were significantly lower than untreated p47(phox-/-) mice (0.89 +/- 0.30 colonies v 237.7 +/- 83.6 colonies at 24 hours, P<.02; 4.0 +/- 2.0 colonies v 110.2 +/- 26.5 colonies at 7 days, P<.0014). More importantly, Kaplan-Meier survival analysis showed a significant survival advantage of gene corrected versus untreated p47(Phox-/-) mice (P<.001). Thus, stem-cell-directed ex vivo gene therapy is capable of restoring phagocyte oxidant-dependent host-defense function in this mouse model of a human immune-system disorder.