TRANSDUCTION OF NONDIVIDING CELLS USING PSEUDOTYPED DEFECTIVE HIGH-TITER HIV TYPE-1 PARTICLES

The use of Moloney marine leukemia virus (Mo-MLV)-based vectors to del iver therapeutic genes into target cells is limited by their inability to transduce nondividing cells. To test the capacity of HIV-based vec tors to deliver genes into nondividing cells, we have generated replic ation-defective HIV type 1 (HIV-1) reporter vectors carrying neomycin phosphotransferase or mouse heat stable antigen, replacing the HIV-1 s equences encoding gp160. These vectors also harbor inactive vpr, vpu, and nef coding regions, Pseudotyped HIV-1 particles carrying either th e ecotropic or the amphotropic Mo-MLV envelope proteins or the vesicul ar stomatitis virus G protein were released after single or double tra nsfections of either human 293T or monkey COS-7 cells with titers of u p to 10(7) colony-forming units per milliliter, A simple ultrafiltrati on procedure resulted in an additional 10- to 20-fold concentration of the pseudotyped particles. These vectors along with Mo-MLV-based vect ors were used to transduce primary human skin fibroblasts and human pe ripheral blood CD34(+) cells. The HIV-1 vector system was significantl y more efficient than its Mo-MLV-based counterpart in transducing huma n skin fibroblasts arrested al the G(0)/G(1) stage of the cell. cycle by density-dependent inhibition of growth. Human CD34(+) cells were tr ansduced efficiently using HIV-1 pseudotype particles without prior st imulation with cytokines.