Lentiviral-mediated gene transfer into human lymphocytes: role of HIV-1 accessory proteins

Resting lymphocytes are refractory to gene transfer using Moloney murine le ukemia virus (MMLV)-based retroviral vectors because of their quiescent sta tus, Recently, it has been shown that lentiviral Vectors are capable of tra nsferring genes into nondividing and terminally differentiated cells, We us ed human immunodeficiency virus type-1 (HIV-1)-based vectors expressing enh anced green fluorescent protein (EGFP) driven by different promoters (CMV, MPSV, or PGK) and investigated their ability to transduce human T- and B-ce ll lines, as well as resting or activated primary peripheral and umbilical cord blood lymphocytes, The effects of the presence or the absence of HIV-I accessory proteins (Vif, Vpr, Vpu, and Nef) in the vector system were also assessed, Flow cytometry analysis showed no differences in the ability of these vectors of transferring the reporter gene into lymphocytic lines and mitogen-stimulated primary lymphocytes in the presence or the absence of HI V-1 accessory proteins (APs), Similarly, viral supernatants generated in th e presence of accessory genes could efficiently transduce various subsets o f resting lymphocytes and provide long-term expression of the transgene, No significant transduction-induced changes in cell activation or cycling sta tus were observed and Alu-HIV-1 long terminal repeat polymerase chain react ion (LTR PCR) analysis demonstrated integration of the vector sequences at the molecular level. In contrast, in the absence of HIV-1 APs, lentiviral v ectors failed to integrate and express the transgene in resting lymphocytes , These results show that transduction of primary resting lymphocytes with HIV-1-based vectors requires the presence of viral accessory proteins.(Bloo d. 2000;96:1309-1316) (C) 2000 by The American Society of Hematology.