title: Analysis of the PKR-eIF2alpha phosphorylation homology domain (PePHD) of hepatitis C virus genotype 1 in HIV-coinfected patients by ultra-deep pyrosequencing and its relationship to responses to pegylated interferon-ribavirin treatment.
abstract: Chronic coinfection with hepatitis C virus (HCV) and human immunodeficiency virus (HIV) is among the greatest challenges facing public health worldwide. In this population, the response to hepatitis C therapy by treatment with pegylated interferon plus ribavirin (PEG-IFN+RBV) is lower than in HCV-monoinfected patients, particularly in those infected by HCV genotype 1. A PKR/eIF-2α phosphorylation homology domain (PePHD) within the E2 protein has been found to interact with PKR and inhibit PKR in vitro, suggesting a possible mechanism for HCV to evade the antiviral effects of IFN. The aim of this work was to analyze the amino acid conservation in the HCV-E2-PePHD and quasispecies diversity among HCV-HIV-coinfected patients exhibiting sustained virological response, non-response, or partial response with viral relapse to PEG-IFN+RBV by ultra-deep pyrosequencing. For this purpose, HCV-E2-PePHD PCR products were generated and sequenced directly for four patients with a sustained response, seven patients with no virological response, and four patients with viral relapse before and after treatment with PEG-IFN+RBV. HCV-E2-PePHD amino acid sequences were obtained for isolates from serum collected before and during treatment (24 h, 4 weeks, and 12 weeks). Quasispecies analysis of the HCV-E2-PePHD and flanking genomic regions was performed using 454/Roche pyrosequencing, analyzing 39,364 sequence reads in total. The HCV-E2-PePHD sequence at the amino acid and nucleotide level was highly conserved among HCV genotype 1 strains, irrespective of the PEG-IFN+RBV response. This high degree of amino acid conservation and sporadic mutations in the HCV-E2-PePHD domain do not appear to be associated with treatment outcome. The HCV-E2-PePHD sequence before or during treatment cannot be used to predict reliably the outcome of treatment in patients coinfected with HCV genotype 1 and HIV.
authors.affiliation: Departamento de Microbiología, Facultad de Medicina (UBA), Centro Nacional de Referencia para el SIDA, Paraguay 2155, Piso 11, C1121ABG, Buenos Aires, Argentina.
authors.forename: F
authors.initials: F
authors.lastname: Bolcic
authors.forename: M
authors.initials: M
authors.lastname: Sede
authors.forename: F
authors.initials: F
authors.lastname: Moretti
authors.forename: G
authors.initials: G
authors.lastname: Westergaard
authors.forename: M
authors.initials: M
authors.lastname: Vazquez
authors.forename: N
authors.initials: N
authors.lastname: Laufer
authors.forename: J
authors.initials: J
authors.lastname: Quarleri
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000627:therapeutic use; D016000:Cluster Analysis; D017124:Conserved Sequence; D005260:Female; D014644:Genetic Variation; D005838:Genotype; D015658:HIV Infections / Q000150:complications*; D016174:Hepacivirus / Q000187:drug effects* / Q000235:genetics; D019698:Hepatitis C, Chronic / Q000145:classification / Q000188:drug therapy / Q000821:virology*; D006801:Humans; D007372:Interferons / Q000627:therapeutic use*; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010802:Phylogeny; D012254:Ribavirin / Q000627:therapeutic use*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D044967:Serum / Q000821:virology; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000378:metabolism*
chemical_list: D000998:Antiviral Agents; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus; D012254:Ribavirin; D007372:Interferons; D019892:eIF-2 Kinase