title: Dynamics of HCV genotype 4 resistance-associated variants during virologic escape with pIFN/RBV+daclatasvir: a case study using ultra deep pyrosequencing.
abstract: BACKGROUND
  Daclatasvir (DCV) is an approved NS5A inhibitor with potent anti-HCV activity and broad genotype coverage. DCV resistance-associated variants (RAVs) have been described for patients infected with genotype (GT) 1, but increased GT4 prevalence in European countries as a result of immigration has boosted interest in this genotype.
  
  
  OBJECTIVE
  Establishment of NS5A variability in treatment-naive patients with HCV genotype 4 infection and a case study of the dynamics of resistance-associated variants in a virologic failure receiving pIFN/RBV+DCV, as assessed by ultra-deep sequencing.
  
  
  METHODS
  Five treatment-naïve GT4 patients (GT4a [n = 1], GT4d [n = 3], GT4o [n = 1]) were evaluated for inclusion in the COMMAND-4 study and treatment with pIFN/RBV±DCV.
  
  
  RESULTS
  Patient (Pt) 1 received pIFN/RBV; Pts2-4 received pIFN/RBV + DCV; Pt5 was a screening failure. Pt1 relapsed; Pt2 experienced breakthrough at Wk4; Pts3 and 4 achieved a sustained virologic response. No substitutions associated with DCV-resistance were detected at baseline. In terms of viremic time points for Pts1 and 2, the extent of NS5A diversity pre-treatment was not significantly related to viral load (r = -0568; p = 0.035). In Pt2, multiple substitutions associated with DCV-resistance were observed after breakthrough at NS5A amino acid positions 28, 31 and 93. These substitutions were frequently observed on the same haplotype (L28S + M31I = 55.52, 82.50, and 99.36% at Wk4, 8 and 9; L28S + M31I + Y93H = 11.77, 5.01 and <0.6% at Wk4, 8 and 9).
  
  
  CONCLUSIONS
  This is the first report to describe DCV-resistance in patients infected with GT4d, supporting a possible role for a recently described RAV (L28S), and presenting the dynamics of HCV quasispecies during therapy failure, with indications of changes of diversity and association of mutations.
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Barbara
authors: B
authors: Bartolini
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Raffaella
authors: R
authors: Lionetti
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Emanuela
authors: E
authors: Giombini
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Catia
authors: C
authors: Sias
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Chiara
authors: C
authors: Taibi
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Marzia
authors: M
authors: Montalbano
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Gianpiero
authors: G
authors: D'Offizi
authors: D'Offizi
authors: D
authors: Gianpiero
authors: Research & Development, Bristol-Myers Squibb, Wallingford, CT, United States.
authors: Fiona
authors: F
authors: McPhee
authors: Research & Development, Bristol-Myers Squibb, Princeton, NJ, United States.
authors: Eric A
authors: EA
authors: Hughes
authors: Research & Development, Bristol-Myers Squibb, Wallingford, CT, United States.
authors: Nannan
authors: N
authors: Zhou
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Giuseppe
authors: G
authors: Ippolito
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy.
authors: Anna Rosa
authors: AR
authors: Garbuglia
authors: "L. Spallanzani" National Institute for Infectious Diseases, Rome, Italy. Electronic address: maria.capobianchi@inmi.it.
authors: Maria R
authors: MR
authors: Capobianchi
mesh_terms: D000293:Adolescent; D000328:Adult; D000998:Antiviral Agents / Q000627:therapeutic use*; D002219:Carbamates; D024882:Drug Resistance, Viral; D014644:Genetic Variation; D005838:Genotype; D016174:Hepacivirus / Q000145:classification / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D019698:Hepatitis C, Chronic / Q000188:drug therapy / Q000821:virology*; D006801:Humans; D007093:Imidazoles / Q000627:therapeutic use*; D007372:Interferons / Q000627:therapeutic use*; D008875:Middle Aged; D011759:Pyrrolidines; D012254:Ribavirin / Q000627:therapeutic use*; D017211:Treatment Failure; D014633:Valine / Q000031:analogs & derivatives; D019562:Viral Load; D017361:Viral Nonstructural Proteins / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase
chemical_list: D000998:Antiviral Agents; D002219:Carbamates; D007093:Imidazoles; D007372:Interferons; D011759:Pyrrolidines; D012254:Ribavirin; D014633:Valine; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase; C549273:daclatasvir
keywords: Daclatasvir; Genotype 4; Hepatitis C virus; NS5A; UDPS