Combination of V106I and V179D polymorphic mutations in human immunodeficiency virus type 1 reverse transcriptase confers resistance to efavirenz and nevirapine but not etravirine

Abstract

Etravirine (ETV) is a second-generation nonnucleoside reverse transcriptase (RT) inhibitor (NNRTI) introduced recently for salvage antiretroviral treatment after the emergence of NNRTI-resistant human immunodeficiency virus type 1 (HIV-1). Following its introduction, two naturally occurring mutations in HIV-1 RT, V106I and V179D, were listed as ETV resistance-associated mutations. However, the effect of these mutations on the development of NNRTI resistance has not been analyzed yet. To select highly NNRTI-resistant HIV-1 in vitro, monoclonal HIV-1 strains harboring V106I and V179D (HIV-1(V106I) and HIV-1(V179D)) were propagated in the presence of increasing concentrations of efavirenz (EFV). Interestingly, V179D emerged in one of three selection experiments from HIV-1(V106I) and V106I emerged in two of three experiments from HIV-1(V179D). Analysis of recombinant HIV-1 clones showed that the combination of V106I and V179D conferred significant resistance to EFV and nevirapine (NVP) but not to ETV. Structural analysis indicated that ETV can overcome the repulsive interactions caused by the combination of V106I and V179D through fine-tuning of its binding module to RT facilitated by its plastic structure, whereas EFV and NVP cannot because of their rigid structures. Analysis of clinical isolates showed comparable drug susceptibilities, and the same combination of mutations was found in some database patients who experienced virologic NNRTI-based treatment failure. The combination of V106I and V179D is a newly identified NNRTI resistance pattern of mutations. The combination of polymorphic and minor resistance-associated mutations should be interpreted carefully.

FIG. 1.

Selected amino acid substitutions under selective pressure from EFV. HIV-1_V106I (A to C) and HIV-1_V179D (D to F) were propagated in MT-2 cells in the presence of increasing concentrations of EFV. The selected amino acid substitutions were analyzed at several passages by sequencing the proviral HIV-1 RT coding region in MT-2 cells. Amino acid substitutions compared with wild-type strain BH 10 are shown.

FIG. 2.

Replication kinetics of recombinant HIV-1 clones in the absence and presence of NNRTIs. Recombinant HIV-1 clones were propagated in MT-2 cells in the absence (A) and presence of 10 nM EFV (B), 100 nM NVP (C), or 10 nM ETV (D). The concentration of p24 in the culture medium was measured every other day. The assay was performed in triplicate and repeated three times (nine experiments). The data are logarithmic mean p24 concentrations ± standard deviations (days 6 and 8 in panels A to C).

FIG. 3.

Interactions of NNRTIs with the 106th and 179th residues of RTs. Interaction sites of NNRTIs and RTs in the models are shown. (A) Wild-type RT (RT_WT) with EFV. (B) RT_V106I/V179D with EFV. (C) RT_WT with NVP. (D) RT_V106I/V179D with NVP. (E) RT_WT with ETV. (F) RT_V106I/V179D with ETV. In the RT_WT and RT_V106I/V179D models, carbon atoms appear gray and cyan, respectively. NNRTIs and the 106th and 179th residues are highlighted by the stick configuration. Blue sticks, nitrogen; red sticks, oxygen; light blue, fluorine; light green, chlorine; pink sticks, bromine atoms.

FIG. 4.

Superposition of RT_WT and five mutant models. (A) RTs with ETV. (B) RTs with EFV. (C) RTs with NVP. NNRTIs and the 106th and 179th residues are highlighted by the stick configuration. White sticks, RT_WT; yellow, RT_V106A; green, RT_V106I; orange, RT_V179D; purple, RT_V106A/V179D; cyan, RT_V106I/V179D.