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. 2001 Feb;8(3):190-8.
doi: 10.1038/sj.gt.3301378.

Lentivirus-mediated gene transfer in primary T cells is enhanced by a central DNA flap

V Dardalhon  1 , B Herpers, N Noraz, F Pflumio, D Guetard, C Leveau, A Dubart-Kupperschmitt, P Charneau, N Taylor
Affiliations

Lentivirus-mediated gene transfer in primary T cells is enhanced by a central DNA flap

V Dardalhon et al. Gene Ther. 2001 Feb.

Abstract

Retroviral vectors have become the primary tool for gene delivery into hematopoietic cells, including T lymphocytes. Lentiviral vectors offer an advantage over Moloney murine leukemia virus (MuLV) vectors because of their ability to translocate across an intact nuclear membrane and integrate into the genome of nonproliferating cells. We have recently demonstrated that a central strand displacement event, controlled by the central polypurine tract (cPPT) and the central termination sequence (CTS), results in the formation of a central DNA flap which acts as a cis-determinant of HIV-1 genome nuclear import. Here, we show that insertion of this DNA determinant in a classical lentiviral vector resulted in a significantly higher level of transduction in activated T cells (51 +/- 12.7% versus 15 +/- 1.4%). CD4(+) and CD8(+) T cells were transduced at equivalent levels. Importantly, freshly isolated T cells stimulated only during the 12-h transduction period could be efficiently transduced with this new flap-containing lentiviral vector, but not with the parental lentiviral vector nor an MuLV vector. Transgene expression in the flap-containing lentiviral vector, under the control of either an internal cytomegalovirus or the elongation factor-1 alpha (EF1 alpha) promoter, was significant and expression remained elevated in resting T cells. Thus, this system allows stable expression of transgenes in T lymphocytes following a short ex vivo transduction protocol.

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