We propose a collaborative project to assess the ability of a novel, highly effective gene delivery system to deliver TAR decoy constructs (polyTAR) to inhibit human and simian immunodeficiency virus (HIV-1 and SIV) replication in vitro and SIV in vivo in rhesus macaque monkeys. Effective use of genetic therapy for prophylaxis and treatment of HIV-1 infection remains an elusive goal. This project will address that goal using a series of unique tools. These include an in vitro CD34+ cell differentiation culture system developed by Dr. Paul Johnson, in which progenitor cells cultured on thymic stroma differentiate to CD3+4+8-T lymphocytes, susceptible to SIV infection. This system will test protection of differentiated cells from SIV by progenitor cell gene transfer. PolyTAR delivered by retroviral vector to selected progenitor cells inhibits SIV and HIV-1 replication in derivative T cells and macrophages. We will also apply a very effective gene delivery vehicle, SV40. We have shown that SV40 can transduce normal human and monkey bone marrow progenitor cells in vitro with very high efficiency without selection. As SV40-derived viruses are made in titers greater than 10/10ml and as they are not immunogenic, SV40 delivery of polyTAR may allow in vivo gene therapy of SIV and -HIV-1. We hypothesize that SV40 can deliver polyTAR efficiently to T cells and to bone marrow progenitor cells, to effectively inhibit HIV-1 and SIV in vitro and in vivo. To test this hypothesis we will generate SV40 containing polyTAR (SV(polyTAR)) and will test both its delivery of polyTAR and inhibition of SIV and HIV-1 in susceptible cell lines. PolyTAR expression and cellular susceptibility to SIV will be measured in both macrophages and T lymphocytes derived from SV(polyTAR)- transduced progenitor cells. SV(polyTAR) will be tested for inhibition of SIV in rhesus mecaque monkeys following progenitor cell transduction ex vivo and reinfusion in vivo, and following direct transduction in vivo. Based on these data, vector design will be modified to enhance polyTAR expression and lentivirus resistance of SIV- and HIV-1- susceptible cell lines and normal simian CD34+ cells, compared to retrovirus-transduced polyTAR. These modified SV(polyTAR) vectors will be designed for longer and greater polyTAR expression. The efficacy of these optimized SV(polyTAR) vectors in inhibiting SIV and HIV-1 infection in vitro and SIV infection in vivo will be assessed similarly. The combined approach of SV40 delivery, polyTAR activity, and stem cell transduction offers great promise for the therapy of HIV-1 infection. Studies proposed here in nonhuman primates will help to define the utility of this approach to AIDS therapy.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Research Project (R01)
Project #
1R01RR013156-01A1
Application #
2719232
Study Section
AIDS and Related Research Study Section 4 (ARRD)
Program Officer
Robinson, Jerry
Project Start
1998-09-01
Project End
2002-08-31
Budget Start
1998-09-01
Budget End
1999-08-31
Support Year
1
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Thomas Jefferson University
Department
Pathology
Type
Schools of Medicine
DUNS #
061197161
City
Philadelphia
State
PA
Country
United States
Zip Code
19107
Strayer, David S; Mitchell, Christine; Maier, Dawn A et al. (2010) Production of SV40-derived vectors. Cold Spring Harb Protoc 2010:pdb.prot5436
Mueller, C; Strayer, M S; Sirninger, J et al. (2010) In vitro and in vivo functional characterization of gutless recombinant SV40-derived CFTR vectors. Gene Ther 17:227-37
Strayer, David S; Mitchell, Christine; Maier, Dawn A et al. (2010) Titering replication-defective rSV40 vectors. Cold Spring Harb Protoc 2010:pdb.prot5437
Strayer, David S; Feitelson, Mark; Sun, Bill et al. (2005) Paradigms for conditional expression of RNA interference molecules for use against viral targets. Methods Enzymol 392:227-41
Strayer, David S; Cordelier, Pierre; Kondo, Rumi et al. (2005) What they are, how they work and why they do what they do? The story of SV40-derived gene therapy vectors and what they have to offer. Curr Gene Ther 5:151-65
Cordelier, P; Kulkowsky, J W; Ko, C et al. (2004) Protecting from R5-tropic HIV: individual and combined effectiveness of a hammerhead ribozyme and a single-chain Fv antibody that targets CCR5. Gene Ther 11:1627-37
Cordelier, Pierre; Morse, Barry; Strayer, David S (2003) Targeting CCR5 with siRNAs: using recombinant SV40-derived vectors to protect macrophages and microglia from R5-tropic HIV. Oligonucleotides 13:281-94
Matskevich, Alexey A; Cordelier, Pierre; Strayer, David S (2003) Conditional expression of IFN-alpha and IFN-gamma activated by HBV as genetic therapy for hepatitis B. J Interferon Cytokine Res 23:709-21
Cordelier, P; Zern, M A; Strayer, D S (2003) HIV-1 proprotein processing as a target for gene therapy. Gene Ther 10:467-77
Matskevich, A A; Strayer, D S (2003) Exploiting hepatitis C virus activation of NFkappaB to deliver HCV-responsive expression of interferons alpha and gamma. Gene Ther 10:1861-73

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