Project #1 of this IP/CP application proposes exploit the high transduction efficiency of recombinant SV40-derived gene therapy vectors, to test combination genetic therapies to inhibit HIV-1 infection in vitro in cultured cell lines and primary peripheral blood mononuclear cells (pbmc). rSV40 vectors are replication-defective viruses derived from Tag-deleted simian virus-40 genomes that integrate very rapidly in to cellular genomes, and infect almost all cell types. They have very high production titers (up to 10/11 infectious units (IU)/ml) and transduction efficiencies (>98%). These vectors deliver several different anti-lentiviral transgenes, or in sequence, to cultured cell lines, human pbmc and human and simian CD34+ cells. These inhibit HIV-1 in transduced cells. Our overall hypotheses for this project are: 1. SV40 can deliver multiple transgenes to the same cells to inhibit lentivirus replication with high efficiency 2. Simultaneous expression of combinations of transgenes, each targeting a different phase of the lentivirus replicative cycle, will provide enhanced protection from lentiviral challenge This project proposes to develop and evaluate rSV40 delivery of combinations of anti-lentiviral transgenes, each targeting a different phase of the HIV-1 replicative cycle: RT#3, single chain antibody vs. HIV-1 reverse transcriptase (RT); Rev410, dominant negative mutant Rev protein; and MIP-1beta-KDEL, a ligand for the CCR5 co-receptor,, targeted to the endoplasmic reticulum. Combinations of these transgenes, delivered in sequence by their respective rSV40 vectors, will be studied for levels and duration of expression in transduced cell lines and pbmc, cytotoxicity and effects on cell survival. Protection conferred by these transgenes, administered in the proposed combinations, will be measured using both cell lines and normal pbmc as targets, and using and T- tropic strains of HIV-1, including at least one clinical isolate of HIV-1. In the event that combinations of these transgenes prove either ineffective or toxic, alternative transgenes are on hand that will be used in their steads. Toxicity and efficacy data from Projects #2 and #3 will be collated with data from this project to determine the need (if any) for revising transduction protocols and/or transgenes used. Finally, as new and promising genetic therapies for lentiviral infection are reported and become available, rSV40 vectors incorporating these will be developed and studied in the same fashion. Therefore, in this project we propose to take advantage of the strengths of the rSV40 vector system and, working in tandem with investigators in Projects #2 and #3, will develop combinations gene therapy to inhibit HIV-1.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Program Projects (P01)
Project #
1P01AI048244-01
Application #
6400325
Study Section
Special Emphasis Panel (ZAI1)
Project Start
2000-09-30
Project End
2004-08-31
Budget Start
Budget End
Support Year
1
Fiscal Year
2000
Total Cost
Indirect Cost
Name
Thomas Jefferson University
Department
Type
DUNS #
061197161
City
Philadelphia
State
PA
Country
United States
Zip Code
19107
Louboutin, Jean-Pierre; Strayer, David S (2012) Blood-brain barrier abnormalities caused by HIV-1 gp120: mechanistic and therapeutic implications. ScientificWorldJournal 2012:482575
Louboutin, J-P; Chekmasova, A A; Reyes, B A S et al. (2011) Bone marrow-derived cells migrate to line the vessels of the CNS: opportunities for gene delivery to CNS vasculature. Neuroscience 195:215-23
Strayer, David S; Mitchell, Christine; Maier, Dawn A et al. (2010) Titering replication-defective rSV40 vectors. Cold Spring Harb Protoc 2010:pdb.prot5437
Louboutin, Jean-Pierre; Agrawal, Lokesh; Reyes, Beverly A S et al. (2010) HIV-1 gp120-induced injury to the blood-brain barrier: role of metalloproteinases 2 and 9 and relationship to oxidative stress. J Neuropathol Exp Neurol 69:801-16
Louboutin, Jean-Pierre; Reyes, Beverly A S; Agrawal, Lokesh et al. (2010) Blood-brain barrier abnormalities caused by exposure to HIV-1 gp120--protection by gene delivery of antioxidant enzymes. Neurobiol Dis 38:313-25
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
Agrawal, Lokesh; Maxwell, Christina R; Peters, Paul J et al. (2009) Complexity in human immunodeficiency virus type 1 (HIV-1) co-receptor usage: roles of CCR3 and CCR5 in HIV-1 infection of monocyte-derived macrophages and brain microglia. J Gen Virol 90:710-22
Louboutin, Jean-Pierre; Agrawal, Lokesh; Reyes, Beverly A S et al. (2009) A rat model of human immunodeficiency virus 1 encephalopathy using envelope glycoprotein gp120 expression delivered by SV40 vectors. J Neuropathol Exp Neurol 68:456-73
Louboutin, J-P; Agrawal, L; Reyes, B A S et al. (2007) Protecting neurons from HIV-1 gp120-induced oxidant stress using both localized intracerebral and generalized intraventricular administration of antioxidant enzymes delivered by SV40-derived vectors. Gene Ther 14:1650-61

Showing the most recent 10 out of 31 publications