GENE THERAPY Third-generation, self-inactivating gp91phox lentivector corrects the oxidase defect in NOD/SCID mouse–repopulating peripheral blood–mobilized CD34 cells from patients with X-linked chronic granulomatous disease

HIV-1–derived lentivectors are promising for gene transfer into hematopoietic stem cells but require preclinical in vivo evaluation relevant to specific human diseases. Nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice accept human hematopoietic stem cell grafts, providing a unique opportunity for in vivo evaluation of therapies targeting human hematopoietic diseases. We demonstrate for the first time that hematopoietic stem cells from patients with X-linked chronic granulomatous disease (X-CGD) give rise to X-CGD–phenotype neutrophils in the NOD/SCID model that can becorrectedusingVSV-G–pseudotyped,3rdgeneration, self-inactivating (SIN) lentivector encoding gp91phox. We transduced X-CGD patient-mobilized CD34 peripheral blood stem cells (CD34 PBSCs) with lentivector– gp91phox or amphotropic oncoretrovirus MFGS–gp91phox and evaluated correction ex vivo and in vivo in NOD/SCID mice. Only lentivector transduced CD34 PBSCs under ex vivo conditions nonpermissive for cell division, but both vectors performed best under conditions permissive for proliferation (multiple growth factors). Under the latter conditions, lentivector and MFGS achieved significant ex vivo correction of X-CGD CD34 PBSCs (18% and 54% of cells expressing gp91phox, associated with 53% and 163% of normal superoxide production, respectively). However, lentivector, but not MFGS, achieved significant correction of human X-CGD neutrophils arising in vivo in NOD/SCID mice that underwent transplantation (20% and 2.4%, respectively). Thus, 3rd-generation SIN lentivector–gp91phox performs well as assessed in human X-CGD neutrophils differentiating in vivo, and our studies suggest that the NOD/SCID model is generally applicable for in vivo study of therapies evaluated in human blood cells expressing a specific disease phenotype. (Blood. 2002;100:4381-4390)