Advances in the isolation, characterization, and culture of human hematopoietic CD34+CD38- cells with extensive in vivo repopulating potential have been recently described. These developments have facilitated their evaluation as suitable targets for bone marrow transplantation and human gene therapy approaches to treat AIDS, cancer, hematological abnormalities, and inborn errors of metabolism. Unfortunately, the development of murine retrovirus-based gene transfer vectors has not kept up with these advances and progress towards clinical implementation of gene therapy protocols has lagged. Here we demonstrate that lentivirus-based vectors transduce human CD34+ and CD34+CD38- cells with high efficiency under conditions that maintain their in vivo repopulating potential. We also show that transduced CD34+CD38- cells can be induced to differentiate in culture resulting in sustained expression of the tgransgene in differentiated cell types. This grant proposal is intended to further these exciting observations in a human/mouse xenograft model. Our hypothesis is that lentivirus-based vectors are an excellent alternative for gene therapy with promise for clinical implementation. We therefore focus this proposal on the in vivo analysis of genetically modified CD34+ and CD34+CD38- cells and propose to show that after transduction these cells maintain their in vivo repopulating potential. The applicant believes that progress towards these goals will bring lentivirus-based vectors significantly closer to clinical implementation and further their utility as tools for discovery in basic research.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA082055-02
Application #
6377274
Study Section
Special Emphasis Panel (ZRG1-AARR-3 (01))
Program Officer
Evans, Gregory
Project Start
2000-05-05
Project End
2003-03-31
Budget Start
2001-04-01
Budget End
2002-03-31
Support Year
2
Fiscal Year
2001
Total Cost
$351,000
Indirect Cost
Name
University of Texas Sw Medical Center Dallas
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
City
Dallas
State
TX
Country
United States
Zip Code
75390
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Denton, Paul W; Estes, Jacob D; Sun, Zhifeng et al. (2008) Antiretroviral pre-exposure prophylaxis prevents vaginal transmission of HIV-1 in humanized BLT mice. PLoS Med 5:e16
Sun, Zhifeng; Denton, Paul W; Estes, Jacob D et al. (2007) Intrarectal transmission, systemic infection, and CD4+ T cell depletion in humanized mice infected with HIV-1. J Exp Med 204:705-14
Melkus, Michael W; Estes, Jacob D; Padgett-Thomas, Angela et al. (2006) Humanized mice mount specific adaptive and innate immune responses to EBV and TSST-1. Nat Med 12:1316-22
Cravens, Petra D; Melkus, Michael W; Padgett-Thomas, Angela et al. (2005) Development and activation of human dendritic cells in vivo in a xenograft model of human hematopoiesis. Stem Cells 23:264-78
Islas-Ohlmayer, Miguel; Padgett-Thomas, Angela; Domiati-Saad, Rana et al. (2004) Experimental infection of NOD/SCID mice reconstituted with human CD34+ cells with Epstein-Barr virus. J Virol 78:13891-900
Gatlin, Joel; Melkus, Michael W; Padgett, Angela et al. (2003) In vivo fluorescent labeling of corneal wound healing fibroblasts. Exp Eye Res 76:361-71
Gatlin, J; Unett, D J; Lerner, M R et al. (2001) Efficient, long-term transgene expression in Xenopus laevis dermal melanophores. Pigment Cell Res 14:275-82
Gatlin, J; Melkus, M W; Padgett, A et al. (2001) Engraftment of NOD/SCID mice with human CD34(+) cells transduced by concentrated oncoretroviral vector particles pseudotyped with the feline endogenous retrovirus (RD114) envelope protein. J Virol 75:9995-9
Gatlin, J; Padgett, A; Melkus, M W et al. (2001) Long-term engraftment of nonobese diabetic/severe combined immunodeficient mice with human CD34+ cells transduced by a self-inactivating human immunodeficiency virus type 1 vector. Hum Gene Ther 12:1079-89

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