Added Value of Deep Sequencing Relative to Population Sequencing in Heavily Pre-Treated HIV-1-Infected Subjects

Francisco M Codoñer,David Dalmau,Christian Pou,Miguel Álvarez-Tejado,Alexander Thielen,Federico García,Rafael Delgado,Lidia Ruiz,Bonaventura Clotet,Roger Paredes,Esper Georges Kallas

doi:10.1371/journal.pone.0019461

Abstract

ObjectiveTo explore the potential of deep HIV-1 sequencing for adding clinically relevant information relative to viral population sequencing in heavily pre-treated HIV-1-infected subjects.MethodsIn a proof-of-concept study, deep sequencing was compared to population sequencing in HIV-1-infected individuals with previous triple-class virological failure who also developed virologic failure to deep salvage therapy including, at least, darunavir, tipranavir, etravirine or raltegravir. Viral susceptibility was inferred before salvage therapy initiation and at virological failure using deep and population sequencing genotypes interpreted with the HIVdb, Rega and ANRS algorithms. The threshold level for mutant detection with deep sequencing was 1%.Results7 subjects with previous exposure to a median of 15 antiretrovirals during a median of 13 years were included. Deep salvage therapy included darunavir, tipranavir, etravirine or raltegravir in 4, 2, 2 and 5 subjects, respectively. Self-reported treatment adherence was adequate in 4 and partial in 2; one individual underwent treatment interruption during follow-up. Deep sequencing detected all mutations found by population sequencing and identified additional resistance mutations in all but one individual, predominantly after virological failure to deep salvage therapy. Additional genotypic information led to consistent decreases in predicted susceptibility to etravirine, efavirenz, nucleoside reverse transcriptase inhibitors and indinavir in 2, 1, 2 and 1 subject, respectively. Deep sequencing data did not consistently modify the susceptibility predictions achieved with population sequencing for darunavir, tipranavir or raltegravir.ConclusionsIn this subset of heavily pre-treated individuals, deep sequencing improved the assessment of genotypic resistance to etravirine, but did not consistently provide additional information on darunavir, tipranavir or raltegravir susceptibility. These data may inform the design of future studies addressing the clinical value of minority drug-resistant variants in treatment-experienced subjects.

Highlights

The rate of virological failure of the 3 original drug classes is low, but not negligible, and does not appear to diminish over time from starting antiretroviral therapy.[1]
Deep salvage therapy included darunavir, tipranavir, etravirine or raltegravir in 4, 2, 2 and 5 subjects, respectively. (Table S1) Treatment adherence was adequate in 4 individuals (Subjects 1 to 4) (Table S1) and partial in 2 (Subjects 5 and 6), one individual (Subject 7) (Table S1) interrupted antiretroviral therapy during follow-up
Deep sequencing detected all mutations found by population sequencing in all subjects, and found additional mutations in 6 out of 7 individuals. (Table S1) Additional mutations were congruent with the treatment history and modified 5.2% of phenotypic susceptibility predictions overall (Table S2), with no significant differences between interpretation algorithms

Summary

Introduction

The rate of virological failure of the 3 original drug classes is low, but not negligible, and does not appear to diminish over time from starting antiretroviral therapy.[1]. Studies have shown that pre-existing minority drug-resistant mutants increase the risk of virological failure to first-line antiretroviral therapy with non-nucleoside reverse transcriptase inhibitors (NNRTIs) [3,4,5,6,7]. Low-frequency drugresistant variants do not affect virological outcomes of first-line therapy including drugs with high genetic barrier, like ritonavirboosted protease inhibitors (PIr) [8]. Whereas most studies addressing the role of minority variants have been performed in antiretroviral-naıve subjects [2,3,4,5,6,7,9,10,11,12,13], less information exists on the clinical significance of minority mutants in antiretroviral-experienced individuals. Whereas most studies addressing the role of minority variants have been performed in antiretroviral-naıve subjects [2,3,4,5,6,7,9,10,11,12,13], less information exists on the clinical significance of minority mutants in antiretroviral-experienced individuals. [14,15,16]

Methods

Results

Discussion

Conclusion