NAEGER LK, MARGOT NA, MILLER MD; Interscience Conference on Antimicrobial Agents and Chemotherapy.

Abstr Intersci Conf Antimicrob Agents Chemother Intersci Conf Antimicrob Agents Chemother. 2000 Sep 17-20; 40: 308.

Gilead Sciences, Foster City, CA

BACKGROUND: Pyrophosphorolysis, the removal of nucleoside chain terminators by a pyrophosphate acceptor molecule, has been proposed as a mechanism of zidovudine (AZT) resistance. A similar mechanism (dinucleotide polyphosphate synthesis) which uses ATP as an acceptor molecule has also been proposed as a mechanism to remove nucleoside chain terminators. We have determined whether these two mechanisms may remove the chain terminating nucleotide analog tenofovir (PMPA).METHODS: Recombinant HIV-1 wild-type RT and mutant RT enzymes containing the thymidine analog resistance mutations T215Y or D67N/K70R were analyzed for pyrophosphorolysis and dinucleotide polyphosphate synthesis activities. These activities were determined in vitro using a heteropolymeric RNA template annealed to end-labelled primers terminated with either PMPA diphosphate (PMPApp) or AZT triphosphate (AZTTP) in the presence of 1 mM NaPPi or 3.2 mM ATP.RESULTS: Using an AZTTP-terminated primer and NaPPi, the T215Y and D67N/K70R RT mutants had increased pyrophosphorolysis activity compared to wild-type RT. In contrast, using a PMPApp-terminated primer and NaPPi, the T215Y and D67N/K70R RT mutants had equivalent pyrophosphorolysis activity as wild type RT. Moreover, analysis of these RT mutants using 3.2 mM ATP as an acceptor molecule demonstrated removal of AZTTP but no detectable removal of PMPApp. Conclusion: Our results suggest that wild-type RT and T215Y and D67N/K70R RT mutants remove tenofovir (PMPApp) less efficiently than AZTTP through pyrophosphorolysis or dinucleotide polyphosphate synthesis. These results are consistent with the minor changes in susceptibility to tenofovir of T215Y and D67N/K70R RT mutants and the corresponding resistance of these mutants to zidovudine.KEYWORDS: Dinucleotide polyphosphate synthesis; Pyrophosphorolysis; Tenofovir

Publication Types:

• Meeting Abstracts

Keywords:

• 3'-azido-3'-deoxythymidine 5'-triphosphate
• Adenine
• Diphosphates
• HIV-1
• In Vitro
• Mutation
• Nucleosides
• Phosphonic Acids
• Soman
• Thymine Nucleotides
• Zidovudine
• genetics
• pinacolyl methylphosphonic acid
• reverse transcriptase, Human immunodeficiency virus 1
• tenofovir

Other ID:

• GWAIDS0009656

UI: 102247154

From Meeting Abstracts