HIV-1 Drug Resistance. HIV-1 drug resistance plays an important role in limiting the efficacy of antiretroviral therapy. One of our goals is to characterize the impact of resistance mutations to non-nucleoside reverse transcriptase inhibitors (NNRTIs) on HIV-1 replication competence and reverse transcriptase function. We have found that 5 different mutants conferring NNRTI resistance demonstrate alterations in RNase H cleavage, and that greater reductions in RNase H activity correlate with reduced replication fitness. We are investigating the effect that these mutants have on specific cleavage events mediated by RNase H, including primer processing and removal, and are studying potential mechanisms for how these mutations, which are remote from the RNase H active site, affect RNase H cleavage.

HIV-1 Recombination. Another focus in my laboratory is the development of an assay to study HIV-1 recombination. We have constructed 2 molecular clones of HIV-1 that differ in a 4 kB region of the genome encompassing gag and part of pol. These 2 clones each contain deletions of env, and contain one of 2 reporter genes that are expressed on the cell surface and can be detected by flow cytometry. Co-transfection of each of these constructs with a plasmid that expresses the VSV G protein results in infectious virions that can complete only a single round of replication. Sequence analysis of sorted dually-infected cell populations will allow us to determine whether certain regions of the viral genome are more prone to recombination and whether recombination is an error-prone process.

Dykes C, Najjar J, Bosch RJ, Wantman M, Furtado M, Hart S, Hammer SM, Demeter LM. Detection of drug-resistant minority variants of HIV-1 during virologic failure of indinavir, lamivudine, and zidovudine. J Infect Dis. 189:1091-6, 2004.

Hirsch MS, Brun-Vezinet F, Clotet B, Conway B, Kuritzkes DR, D'Aquila RT, Demeter LM, .... Richman DD. Antiretroviral drug resistance testing in adults infected with human immunodeficiency virus type 1: 2003 recommendations of an International AIDS Society-USA Panel. Clin Infect Dis. 37:113-28, 2003.

Hammer SM, Bassett R, Squires KE, Fischl MA, Demeter LM, ..... DeGruttola V; ACTG 372B/D Study Team. A randomized trial of nelfinavir and abacavir in combination with efavirenz and adefovir dipivoxil in HIV-1-infected persons with virological failure receiving indinavir. Antivir Ther. 8:507-18, 2003.

Koval CE, Gigliotti F, Nevins D, Demeter LM. Immune reconstitution syndrome after successful treatment of Pneumocystis carinii pneumonia in a man with human immunodeficiency virus type 1 infection. Clin Infect Dis. 35:491-3, 2002.

Demeter LM, Bosch RJ, Coombs RW, Fiscus S, Bremer J, Johnson VA, Erice A, Jackson JB, Spector SA, Squires KM, Fischl MA, Hughes MD, Hammer SM. Detection of replication-competent human immunodeficiency virus type 1 (HIV-1) in cultures from patients with levels of HIV-1 RNA in plasma suppressed to less than 500 or 50 copies per milliliter. J Clin Microbiol. 40:2089-94, 2002.

Demeter LM, Hughes MD, Coombs RW, Jackson JB, Grimes JM, Bosch RJ, Fiscus SA, Spector SA, Squires KE, Fischl MA, Hammer SM. Predictors of virologic and clinical outcomes in HIV-1-infected patients receiving concurrent treatment with indinavir, zidovudine, and lamivudine. AIDS Clinical Trials Group Protocol 320. Ann Intern Med. 135:954-64, 2001.

Dykes C, Fox K, Lloyd A, Chiulli M, Morse E, Demeter LM. Impact of clinical reverse transcriptase sequences on the replication capacity of HIV-1 drug-resistant mutants. Virology. 285:193-203, 2001.

Evans TG, Bonnez W, Rose RC, Koenig S, Demeter L, Suzich JA, O'Brien D, Campbell M, White WI, Balsley J, Reichman RC. A Phase 1 Study of a Recombinant Viruslike Particle Vaccine against Human Papillomavirus Type 11 in Healthy Adult Volunteers. J Infect Dis. 183:1485-1493, 2001.

Archer RH, Wisniewski M, Bambara RA, Demeter LM. The y181c mutant of hiv-1 reverse transcriptase resistant to nonnucleoside reverse transcriptase inhibitors alters the size distribution of rnase h cleavages. Biochemistry 40:4087-95, 2001.

Archer RH, Dykes C, Gerondelis P, Lloyd A, Fay P, Reichman RC, Bambara RA, Demeter LM. Mutants of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase resistant to nonnucleoside reverse transcriptase inhibitors demonstrate altered rates of RNase H cleavage that correlate with HIV-1 replication fitness in cell culture. J Virol. 74:8390-401, 2000.

Dykes C, Mootsikapun P, Dexter A, Berrios L, Chiulli M, Reichman RC, Demeter LM. Analysis of env sequence evolution in human immunodeficiency virus-infected patients receiving therapy with nonnucleoside reverse-transcriptase inhibitors. J Infect Dis. 182:316-20, 2000.