1. Inhibition of simian/human immunodeficiency virus replication in CD4+ T cells derived from lentiviral-transduced CD34+ hematopoietic cells.
- Author
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Braun SE, Wong FE, Connole M, Qiu G, Lee L, Gillis J, Lu X, Humeau L, Slepushkin V, Binder GK, Dropulic B, and Johnson RP
- Subjects
- Adenosine Deaminase metabolism, Animals, Antigens, CD34 genetics, Blotting, Southern, Bone Marrow Cells metabolism, CD4-Positive T-Lymphocytes metabolism, Cell Differentiation, Cell Line, Dose-Response Relationship, Drug, Flow Cytometry, Gene Products, env metabolism, Gene Products, rev metabolism, Gene Products, tat metabolism, Genetic Therapy methods, Genetic Vectors, Green Fluorescent Proteins metabolism, Hematopoietic Stem Cells virology, Humans, Leukocytes, Mononuclear metabolism, Macaca mulatta, Models, Genetic, Oligonucleotides, Antisense chemistry, RNA chemistry, Retroviridae genetics, Stem Cells metabolism, T-Lymphocytes metabolism, Up-Regulation, rev Gene Products, Human Immunodeficiency Virus, tat Gene Products, Human Immunodeficiency Virus, Antigens, CD34 biosynthesis, CD4-Positive T-Lymphocytes immunology, HIV-1 genetics, Hematopoietic Stem Cells metabolism, Lentivirus genetics, Simian Immunodeficiency Virus genetics, Virus Replication
- Abstract
We examined the ability of a HIV-1-based vector (VRX494) encoding a 937-bp antisense HIV-1 envelope sequence to inhibit the replication of chimeric SIV/HIV-1 viruses encoding the HIV-1 envelope. Challenge of VRX494-transduced CEMx174 cells resulted in potent inhibition of HIV-1 and several SHIV strains. To evaluate the potential efficacy of the VRX494 vector for stem cell gene therapy, rhesus CD34(+) bone marrow cells were transduced with VRX494 and then cultured on thymus stroma to induce T cell differentiation. Transduction conditions for CD34(+) cells were optimized to yield high transduction efficiency with minimal effective multiplicity of infection. Purified CD4(+) GFP(+) T cells derived from VRX494-transduced CD34(+) cells strongly inhibited SHIV HXBC2P 3.2 and SHIV 89.6P replication compared to controls. Southern blot analysis of VRX494-transduced T cell clones revealed a subset of cells with multiple proviral copies per cell. Expression of GFP and the antisense inhibitor in VRX494-transduced cells was upregulated by Tat. Analysis of HIV-1 envelope sequences in VRX494-transduced cells revealed modifications consistent with those mediated by double-stranded RNA-dependent adenosine deaminase. These results indicate that the macaque/SHIV model should serve as a useful preclinical model to evaluate this lentiviral vector expressing an HIV-1 antisense inhibitor for stem cell gene therapy for AIDS.
- Published
- 2005
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