Acquired Immune Deficiency Syndrome (AIDS) is a current major health problem. This deadly, disease is caused by the human immune deficiency virus (HIV-1). As of August 1993 about 255,000 AIDS cases have been reported to the Center for Disease control. The targets of the virus are the cells such as human peripheral blood lymphocytes, monocytes/macrophages with virus specific attachment surface proteins (CD4+). The long-term objective of this renewal application is to elucidate the mechanism of action and metabolism of a new series of acyclic nucleoside, phosphonates which have demonstrated broad spectrum antiviral activity both in vitro and in vivo. The phosphonylmethoxy ethyl adenine (PMEA), the lead member of the purine series of compounds, has been shown to be a chain terminator of proviral DNA replication. Phase 1/11 trials of PMEA in HIV-infected patients are underway at various sites in the U.S. The present renewal application proposes basic biochemical studies of PMEA and several other members of the phosphonate analogs, particularly the phosphonylmethoxy propyl (PMP) derivatives which demonstrate increased selectivity against HIV.
The specific aims of the proposed work are to: 1)use variants from the human T cell line CCRF-CEM that have a deficiency in the anabolic phosphorylation of PMEA as model for identification and detailed characterization of the enzyme pathway(s) responsible for the activation of the phosphonate analogs, 2) purify and characterize the enzyme(s) from CEM cells and their variants to define the substrate specificity of analog metabolizing enzyme(s), 3) determine the mechanism of activation that is operative for PMEA and its analogs in primary human mononuclear cells, including lymphocytes and monocytes/ macrophages , that are the primary targets for HIV infection, 4) investigate the potential of HIV to develop resistance to the phosphonate analogs and determine the cross-resistance, if any, with HIV strains that carry mutations to existing antiretroviral drugs such as the dideoxynucleosides and non-nucleoside RT inhibitors. The work proposed in this application will address several deficiencies in our under-standing of the cellular metabolism and action of acyclic phosphonate analogs in human lymphoid cells and will contribute to the long-term goal of developing selective chemotherapy against AIDS and associated opportunistic infections using different drug combinations.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI027652-13
Application #
2063954
Study Section
AIDS and Related Research Study Section 2 (ARRB)
Project Start
1982-07-01
Project End
1997-06-30
Budget Start
1995-07-01
Budget End
1996-06-30
Support Year
13
Fiscal Year
1995
Total Cost
Indirect Cost
Name
St. Jude Children's Research Hospital
Department
Type
DUNS #
067717892
City
Memphis
State
TN
Country
United States
Zip Code
38105
Robbins, B L; Greenhaw, J; Fridland, A (2000) Cellular phosphorylation of 2',3'-dideoxyadenosine-5'-monophosphate, a key intermediate in the activation of the antiviral agent DDI, in human peripheral blood mononuclear cells. Nucleosides Nucleotides Nucleic Acids 19:405-13
Srinivas, R V; Fridland, A (1998) Antiviral activities of 9-R-2-phosphonomethoxypropyl adenine (PMPA) and bis(isopropyloxymethylcarbonyl)PMPA against various drug-resistant human immunodeficiency virus strains. Antimicrob Agents Chemother 42:1484-7
Robbins, B L; Srinivas, R V; Kim, C et al. (1998) Anti-human immunodeficiency virus activity and cellular metabolism of a potential prodrug of the acyclic nucleoside phosphonate 9-R-(2-phosphonomethoxypropyl)adenine (PMPA), Bis(isopropyloxymethylcarbonyl)PMPA. Antimicrob Agents Chemother 42:612-7
Robbins, B L; Tran, T T; Pinkerton Jr, F H et al. (1998) Development of a new cartridge radioimmunoassay for determination of intracellular levels of lamivudine triphosphate in the peripheral blood mononuclear cells of human immunodeficiency virus-infected patients. Antimicrob Agents Chemother 42:2656-60
Srinivas, R V; Middlemas, D; Flynn, P et al. (1998) Human immunodeficiency virus protease inhibitors serve as substrates for multidrug transporter proteins MDR1 and MRP1 but retain antiviral efficacy in cell lines expressing these transporters. Antimicrob Agents Chemother 42:3157-62
Tanaka, M; Srinivas, R V; Ueno, T et al. (1997) In vitro induction of human immunodeficiency virus type 1 variants resistant to 2'-beta-Fluoro-2',3'-dideoxyadenosine. Antimicrob Agents Chemother 41:1313-8
Srinivas, R V; Connely, M; Fridland, A (1997) (S)-1-(3-hydroxy-2-phosphonylmethoxypropyl)cytosine (HPMPC) inhibits HIV-1 replication in epithelial cells, but not T-lymphocytes. Antiviral Res 35:23-7
Robbins, B L; Waibel, B H; Fridland, A (1996) Quantitation of intracellular zidovudine phosphates by use of combined cartridge-radioimmunoassay methodology. Antimicrob Agents Chemother 40:2651-4
Robbins, B L; Greenhaw, J; Connelly, M C et al. (1995) Metabolic pathways for activation of the antiviral agent 9-(2-phosphonylmethoxyethyl)adenine in human lymphoid cells. Antimicrob Agents Chemother 39:2304-8
Robbins, B L; Connelly, M C; Marshall, D R et al. (1995) A human T lymphoid cell variant resistant to the acyclic nucleoside phosphonate 9-(2-phosphonylmethoxyethyl)adenine shows a unique combination of a phosphorylation defect and increased efflux of the agent. Mol Pharmacol 47:391-7

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