Lentivirus vector-mediated hematopoietic stem cell gene transfer of commongamma-chain cytokine receptor in rhesus macaques

Nonhuman primate model systems of autologous CD34(+) cell transplant are th e most effective means to assess the safety and capabilities of lentivirus vectors. Toward this end,,ve tested the efficiency of marking, gene express ion, and transplant of hone marrow and peripheral blood CD34(+) cells using a self-inactivating lentivirus vector (CS-Rh-MLV-E) bearing an internal mu rine leukemia virus long terminal repeat derived from a murine retrovirus a dapted to replicate in rhesus macaques. In vitro cytokine stimulation was n ot required to achieve efficient transduction of CD34(+) cells resulting in marking and gene expression of the reporter gene encoding enhanced green f luorescent protein (EGFP) following transplant of the CD34(+) cells. Monkey s transplanted with mobilized peripheral blood CD34(+) cells resulted in EG FP expression in 1 to 10% of multilineage peripheral blood cells, including red blood cells and platelets, stable for 15 months to date. The relative level of gene expression utilizing this vector is 2- to 10-fold greater tha n that utilizing a non-self-inactivating lentivirus vector bearing the cyto megalovirus immediate-early promoter. In contrast, in animals transplanted with autologous bone marrow CD34(+) cells, multilineage EGFP expression was evident initially but diminished over time. We further tested our lentivir us vector system by demonstrating gene transfer of the human common gamma-c hain cytokine receptor gene (gamma (c)), deficient in X-linked SCID patient s and recently successfully used to treat disease. Marking was 0.42 and .00 1 HIV-1 vector DNA copy per 100 cells in two animals. To date, all EGFP- an d gamma (c)-transplanted animals are healthy. This system may prove useful for expression of therapeutic genes in human hematopoietic cells.