Inhibition of simian/human immunodeficiency virus replication in CD4+ T cells derived from lentiviral-transduced CD34+ hematopoietic cells

Mol Ther. 2005 Dec;12(6):1157-67. doi: 10.1016/j.ymthe.2005.07.698. Epub 2005 Sep 15.

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.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine Deaminase / metabolism
  • Animals
  • Antigens, CD34 / biosynthesis*
  • Antigens, CD34 / genetics
  • Blotting, Southern
  • Bone Marrow Cells / metabolism
  • CD4-Positive T-Lymphocytes / immunology*
  • 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
  • HIV-1 / genetics*
  • Hematopoietic Stem Cells / metabolism*
  • Hematopoietic Stem Cells / virology
  • Humans
  • Lentivirus / genetics*
  • Leukocytes, Mononuclear / metabolism
  • Macaca mulatta
  • Models, Genetic
  • Oligonucleotides, Antisense / chemistry
  • RNA / chemistry
  • Retroviridae / genetics
  • Simian Immunodeficiency Virus / genetics*
  • Stem Cells / metabolism
  • T-Lymphocytes / metabolism
  • Up-Regulation
  • Virus Replication*
  • rev Gene Products, Human Immunodeficiency Virus
  • tat Gene Products, Human Immunodeficiency Virus

Substances

  • Antigens, CD34
  • Gene Products, env
  • Gene Products, rev
  • Gene Products, tat
  • Oligonucleotides, Antisense
  • rev Gene Products, Human Immunodeficiency Virus
  • tat Gene Products, Human Immunodeficiency Virus
  • Green Fluorescent Proteins
  • RNA
  • Adenosine Deaminase