Long-term high-level reconstitution of NADPH oxidase activity in murine X-linked chronic granulomatous disease using a bicistronic vector expressing gp91phox and a Delta LNGFR cell surface marker.
A murine model of X-linked chronic granulomatous disease (X-CGD), an inherited immune deficiency with absent phagocyte NADPH oxidase activity caused by defects in the gp91(phox) gene, was used to evaluate a bicistronic retroviral vector in which expression of human gp91(phox) and a linked gene for Delta LNGFR, a truncated form of human low-affinity nerve growth factor receptor, are under the control of a spleen focus-forming virus long-terminal repeat (LTR). Four independent cohorts of 11-Gy irradiated X-CGD mice (total, 22 mice) were transplanted with or without preselection of transduced X-CGD bone marrow (BM). Transplanted mice had high-level correction of neutrophil gp91(phox) expression and reconstitution of NADPH oxidase activity. Expression lasted for at least 14 months in primary transplants, and persisted in secondary and tertiary transplants. Both gp91(phox) and Delta LNGFR were detected on circulating granulocytes, lymphocytes, lymphoid, and (for Delta LNGFR) red blood cells. Mice receiving transduced bone marrow [BM] preselected ex vivo for Delta LNGFR expression had high-level (= 80%) reconstitution with transduced cells, with an improved fraction of oxidase-corrected neutrophils posttransplant. Analysis of secondary and tertiary CFU-S showed that silencing of individual provirus integrants can occur even after preselection for Delta LNGFR prior to transplantation, and that persistent provirus expression was associated with multiple integration sites in most cases. No obvious adverse consequences of transgenic protein expression were observed.
Sadat
Mohammed A
MA
Herman B. Wells Center for Pediatric Research, Department of Pediatrics (Hematology/Oncology), James Whitcomb Riley Hospital for Children, Indiana University School of Medicine, Indianapolis, IN 46202, USA.
Pech
Nancy
N
Saulnier
So
S
Leroy
Brendan A
BA
Hossle
Johann P
JP
Grez
Manuel
M
Dinauer
Mary C
MC
D000818:Animals; D000954:Antigens, Surface / Q000032:analysis; D015415:Biomarkers / Q000032:analysis; D001772:Blood Cell Count; D016026:Bone Marrow Transplantation; D002460:Cell Line; D015870:Gene Expression; D005822:Genetic Vectors; D006105:Granulomatous Disease, Chronic / Q000201:enzymology* / Q000378:metabolism / Q000628:therapy; D006801:Humans; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D042844:L-Lactate Dehydrogenase (Cytochrome); D008562:Membrane Glycoproteins / Q000235:genetics*; D026901:Membrane Transport Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D009252:NADPH Dehydrogenase / Q000235:genetics; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000235:genetics* / Q000378:metabolism; D009504:Neutrophils / Q000201:enzymology; D010750:Phosphoproteins / Q000235:genetics; D020800:Receptor, Nerve Growth Factor / Q000235:genetics* / Q000378:metabolism; D016897:Respiratory Burst; D012190:Retroviridae / Q000235:genetics; D013155:Spleen Focus-Forming Viruses / Q000235:genetics*; D014161:Transduction, Genetic
D000954:Antigens, Surface; D015415:Biomarkers; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D010750:Phosphoproteins; D020800:Receptor, Nerve Growth Factor; D007770:L-Lactate Dehydrogenase; D042844:L-Lactate Dehydrogenase (Cytochrome); C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; C086986:CYBA protein, human; D009252:NADPH Dehydrogenase