Zidovudine (AZT) Monotherapy Selects for the A360V Mutation in the Connection Domain of HIV-1 Reverse Transcriptase

Jessica H Brehm,Scott Hammer,Nicolas Sluis-Cremer,John W Mellors,David Katzenstein,Dianna L Koontz,Yanille Scott,Steven Perry,Gilda Tachedjian

doi:10.1371/journal.pone.0031558

Abstract

BackgroundWe previously demonstrated in vitro that zidovudine (AZT) selects for A371V in the connection domain and Q509L in ribonuclease H (RNase H) domain of HIV-1 reverse transcriptase (RT) which, together with the thymidine analog mutations D67N, K70R and T215F, confer greater than 100-fold AZT resistance. The goal of the current study was to determine whether AZT monotherapy in HIV-1 infected patients also selects the A371V, Q509L or other mutations in the C-terminal domains of HIV-1 RT.Methodology/Principal FindingsFull-length RT sequences in plasma obtained pre- and post-therapy were compared in 23 participants who received AZT monotherapy from the AIDS Clinical Trials Group study 175. Five of the 23 participants reached a primary study endpoint. Mutations significantly associated with AZT monotherapy included K70R (p = 0.003) and T215Y (p = 0.013) in the polymerase domain of HIV-1 RT, and A360V (p = 0.041) in the connection domain of HIV-1 RT. HIV-1 drug susceptibility assays demonstrated that A360V, either alone or in combination with thymidine analog mutations, decreased AZT susceptibility in recombinant viruses containing participant-derived full-length RT sequences or site-directed mutant RT. Biochemical studies revealed that A360V enhances the AZT-monophosphate excision activity of purified RT by significantly decreasing the frequency of secondary RNase H cleavage events that reduce the RNA/DNA duplex length and promote template/primer dissociation.ConclusionsThe A360V mutation in the connection domain of RT was selected in HIV-infected individuals that received AZT monotherapy and contributed to AZT resistance.

Highlights

Zidovudine (39-azido-39-deoxythymidine, AZT) was the first antiviral drug approved by the U.S Food and Drug Administration for the treatment of HIV infection
The A360V mutation in the connection domain of reverse transcriptase (RT) was selected in HIV-infected individuals that received AZT monotherapy and contributed to AZT resistance
Each of the known thymidine analog mutations (TAM) was identified in one or more on-treatment samples, only the K70R and T215Y mutations were significantly associated with AZT monotherapy

Summary

Introduction

Zidovudine (39-azido-39-deoxythymidine, AZT) was the first antiviral drug approved by the U.S Food and Drug Administration for the treatment of HIV infection. Biochemical studies have demonstrated that RT containing TAMs shows an increased capacity to unblock AZT-monophosphate (MP) terminated primers in the presence of physiological concentrations of ATP [6,7]. In this regard, structural studies have shown that TAMs enhance the binding and/or placement of ATP in the HIV-1 RT active site [8]. We previously demonstrated in vitro that zidovudine (AZT) selects for A371V in the connection domain and Q509L in ribonuclease H (RNase H) domain of HIV-1 reverse transcriptase (RT) which, together with the thymidine analog mutations D67N, K70R and T215F, confer greater than 100-fold AZT resistance. The goal of the current study was to determine whether AZT monotherapy in HIV-1 infected patients selects the A371V, Q509L or other mutations in the Cterminal domains of HIV-1 RT

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