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. 1999 Apr;73(4):2832-40.
doi: 10.1128/JVI.73.4.2832-2840.1999.

Lentivirus vectors using human and simian immunodeficiency virus elements

S M White  1 , M Renda, N Y Nam, E Klimatcheva, Y Zhu, J Fisk, M Halterman, B J Rimel, H Federoff, S Pandya, J D Rosenblatt, V Planelles
Affiliations

Lentivirus vectors using human and simian immunodeficiency virus elements

S M White et al. J Virol. 1999 Apr.

Abstract

Lentivirus vectors based on human immunodeficiency virus (HIV) type 1 (HIV-1) constitute a recent development in the field of gene therapy. A key property of HIV-1-derived vectors is their ability to infect nondividing cells. Although high-titer HIV-1-derived vectors have been produced, concerns regarding safety still exist. Safety concerns arise mainly from the possibility of recombination between transfer and packaging vectors, which may give rise to replication-competent viruses with pathogenic potential. We describe a novel lentivirus vector which is based on HIV, simian immunodeficiency virus (SIV), and vesicular stomatitis virus (VSV) and which we refer to as HIV/SIVpack/G. In this system, an HIV-1-derived genome is encapsidated by SIVmac core particles. These core particles are pseudotyped with VSV glycoprotein G. Because the nucleotide homology between HIV-1 and SIVmac is low, the likelihood of recombination between vector elements should be reduced. In addition, the packaging construct (SIVpack) for this lentivirus system was derived from SIVmac1A11, a nonvirulent SIV strain. Thus, the potential for pathogenicity with this vector system is minimal. The transduction ability of HIV/SIVpack/G was demonstrated with immortalized human lymphocytes, human primary macrophages, human bone marrow-derived CD34(+) cells, and primary mouse neurons. To our knowledge, these experiments constitute the first demonstration that the HIV-1-derived genome can be packaged by an SIVmac capsid. We demonstrate that the lentivirus vector described here recapitulates the biological properties of HIV-1-derived vectors, although with increased potential for safety in humans.

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Figures

FIG. 1
FIG. 1
Production of an RNA pseudotype lentivirus vector. (A) SIVpack is a packaging construct based on SIVmac1A11. SIVpack was constructed by subcloning a subgenomic fragment of SIVmac1A11 into an SV40-derived expression vector and subsequently deleting gp120 envelope sequences; SIVmac1A11 contains a frameshift mutation which inactivates vpr. (B) Transfer vectors. HIV-GFP and HIV-thy are transfer vectors based on HIV-1 and contain all the cis-acting elements needed for reverse transcription, integration, and expression. PBS, primer binding site; SD, splice donor; SA, splice acceptor; RRE, Rev-responsive element. (C) Production of lentivirus vectors. HIV-GFP or HIV-thy, SIVpack, and a VSV-G expression construct (7) were transfected by electroporation into COS cells, and supernatants were harvested and frozen at 48 h. Infection was quantitated on the basis of GFP or Thy-1 expression, depending on the transfer vector used. (D) Flow cytometric analysis of HeLa cells infected with lentivirus vectors. HeLa cells were infected with the indicated lentivirus vectors at a dilution of 1:10. At 48 h postinfection, cells were analyzed by flow cytometry for expression of GFP or Thy-1. See Table 2 for the resulting titers.
FIG. 2
FIG. 2
HIV-GFP/SIVpack/G transduces nondividing cells with a high efficiency. MAGI cells (23) were treated with 0 (Non-irrad.), 2,000, or 5,000 rads of gamma radiation. After 24 h, cells were infected with the indicated retrovirus vectors. Infections were analyzed on the basis of GFP expression. Infections were performed in triplicate. Mean values are reported. Error bars represent standard deviations.
FIG. 3
FIG. 3
Infection of CD4+ lymphocytes and primary neurons with HIV-GFP/SIVpack/G. Cells were infected with HIV-GFP/SIVpack/G at an MOI of 0.2 and visualized by fluorescence microscopy at day 3 postinfection. (A and B) CEMX174 cells. (C and D) Mouse neuronal cells. (A and C) Phase-contrast micrographs. (B and D) Fluorescence micrographs. Mouse neuronal cells were positive for reactivity to the following antibodies: mouse anti-neurofilament 200, rabbit anti-tau, mouse anti-MAP2, and rabbit anti-rat neuron-specific enolase.
FIG. 4
FIG. 4
HIV-thy/SIVpack/G does not induce cell cycle arrest. HeLa cells were mock infected or infected with HIV-thy/SIVpack/G or HIV-1NL4-3-thyenv(−)/G (50) as indicated. At 48 h, cells were stained with fluorescein isothiocyanate-conjugated anti-Thy-1 antibody, fixed, permeabilized, and stained for DNA contents with propidium iodide. Histograms show the cell cycle profiles of Thy-1-positive cells for HIV-thy/SIVpack/G and HIV-1NL4-3-thy env(−)/G infections and untreated cells for mock infection. Cell cycle analysis was performed with Multicycle AV software. The G1, S, and G2 peaks (left, middle, and right shaded areas, respectively) are shown below the DNA profile (dotted line).
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References

    1. Akkina R K, Walton R M, Chen M L, Li Q-X, Planelles V, Chen I S Y. High-efficiency gene transfer into CD34+ cells with a human immunodeficiency virus type 1-based retroviral vector pseudotyped with vesicular stomatitis virus envelope glycoprotein G. J Virol. 1996;70:2581–2585. - PMC - PubMed
    1. Banapour B, Marthas M L, Munn R J, Luciw P A. In vitro macrophage tropism of pathogenic and nonpathogenic molecular clones of simian immunodeficiency virus (SIVmac) Virology. 1991;183:12–19. - PubMed
    1. Banapour B, Marthas M L, Ramos R A, Lohman B L, Unger R E, Gardner M B, Pedersen N C, Luciw P A. Identification of viral determinants of macrophage tropism for simian immunodeficiency virus SIVmac. J Virol. 1991;65:5798–5805. - PMC - PubMed
    1. Berchtold S, Hornung U, Aepinus C. The activation domain of simian immunodeficiency virus SIVmac239 Rev protein is structurally and functionally analogous to the HIV-1 Rev activation domain. Virology. 1995;211:285–289. - PubMed
    1. Brewer G J, Torricelli J R, Evege E K, Price P J. Optimized survival of hippocampal neurons in B27-supplemented Neurobasal, a new serum-free medium combination. J Neurosci Res. 1993;35:567–576. - PubMed

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