Hepatitis C virus (HCV) genotype 1 subtype identification in new HCV drug development and future clinical practice.
BACKGROUND
With the development of new specific inhibitors of hepatitis C virus (HCV) enzymes and functions that may yield different antiviral responses and resistance profiles according to the HCV subtype, correct HCV genotype 1 subtype identification is mandatory in clinical trials for stratification and interpretation purposes and will likely become necessary in future clinical practice. The goal of this study was to identify the appropriate molecular tool(s) for accurate HCV genotype 1 subtype determination.


RESULTS
A large cohort of 500 treatment-naïve patients eligible for HCV drug trials and infected with either subtype 1a or 1b was studied. Methods based on the sole analysis of the 5' non-coding region (5'NCR) by sequence analysis or reverse hybridization failed to correctly identify HCV subtype 1a in 22.8%-29.5% of cases, and HCV subtype 1b in 9.5%-8.7% of cases. Natural polymorphisms at positions 107, 204 and/or 243 were responsible for mis-subtyping with these methods. A real-time PCR method using genotype- and subtype-specific primers and probes located in both the 5'NCR and the NS5B-coding region failed to correctly identify HCV genotype 1 subtype in approximately 10% of cases. The second-generation line probe assay, a reverse hybridization assay that uses probes targeting both the 5'NCR and core-coding region, correctly identified HCV subtypes 1a and 1b in more than 99% of cases.


CONCLUSIONS
In the context of new HCV drug development, HCV genotyping methods based on the exclusive analysis of the 5'NCR should be avoided. The second-generation line probe assay is currently the best commercial assay for determination of HCV genotype 1 subtypes 1a and 1b in clinical trials and practice.
Chevaliez
Stéphane
S
French National Reference Center for Viral Hepatitis B, C and delta, Department of Virology, Hôpital Henri Mondor, Université Paris 12, Créteil, France.
Bouvier-Alias
Magali
M
Brillet
Rozenn
R
Pawlotsky
Jean-Michel
JM
D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D015510:Clinical Medicine; D021901:DNA, Intergenic / Q000235:genetics; D015195:Drug Design; D005814:Genes, Viral / Q000235:genetics; D005838:Genotype; D016174:Hepacivirus / Q000145:classification* / Q000187:drug effects / Q000235:genetics*; D008828:Microbiological Techniques / Q000379:methods*; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D011933:Reagent Kits, Diagnostic; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017361:Viral Nonstructural Proteins / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase
D000998:Antiviral Agents; D021901:DNA, Intergenic; D011933:Reagent Kits, Diagnostic; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase