title: Clinical and analytical relevance of NNRTIs minority mutations on viral failure in HIV-1 infected patients.
abstract: The objective of this study was to assess the analytical and clinical relevance of minority variants using a new pyrosequencing (PSQ) assay and to detect minor variants with frequencies below the current 20% clinical setting limit. A PSQ approach for detecting and quantifying mutations was developed for the analysis of 14 codons of the human immunodeficiency virus (HIV) reverse transcriptase (RT) gene below the limit of conventional sequencing. Ten patients who experienced virological failure (VF) after a first-line regimen of lamivudine, tenofovir, and either efavirenz, nevirapine, or etravirine, as well as 10 controls patients without VF, were included in this retrospective study. Baseline plasma and plasma from the time of VF were assessed using Sanger sequencing and PSQ methods. The analytical sensitivity for the detection of minor sequence variants is 5%. At baseline, no minority variant was detected in 10/10 patient controls using both the Sanger sequencing and PSQ assays, whereas, two patients who failed therapy had baseline non-nucleoside reverse transcriptase inhibitor (NNRTI) mutations that were not detected by the standard genotyping. At the time of VF, standard genotyping detected mutations in four out of the 10 VF patients, whereas, PSQ detected mutations in five out of the 10 VF patients. Clinically, minority mutations at a 10% level of detection can be assessed efficiently by pyrosequencing and used as a suitable predictor of the evolution of viral populations. These traits allow for a better interpretation of data analysis, which can help clinicians in providing a suitable treatment for HIV.
authors.affiliation: Laboratoire Alphabio, Marseille, France; Laboratoire d'Immunologie des Tumeurs et Centre INSERM de Recherche en Cancérologie, Institut Paoli Calmettes, Marseille, France.
authors.forename: Sofiane
authors.initials: S
authors.lastname: Mohamed
authors.forename: Sophie
authors.initials: S
authors.lastname: Ravet
authors.forename: Claire
authors.initials: C
authors.lastname: Camus
authors.forename: Hacène
authors.initials: H
authors.lastname: Khiri
authors.forename: Daniel
authors.initials: D
authors.lastname: Olive
authors.forename: Philippe
authors.initials: P
authors.lastname: Halfon
mesh_terms: D019380:Anti-HIV Agents / Q000627:therapeutic use*; D024882:Drug Resistance, Viral; D005260:Female; D005838:Genotype; D015658:HIV Infections / Q000821:virology*; D054303:HIV Reverse Transcriptase / Q000235:genetics*; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D008297:Male; D020125:Mutation, Missense; D017211:Treatment Failure
chemical_list: D019380:Anti-HIV Agents; D054303:HIV Reverse Transcriptase
keywords: HIV; NNRTIs; minority mutations; pyrosequencing