With the increasing number of licensed direct-acting antivirals (DAA) for the treatment of chronic HCV infection, choosing the right treatment regimen for the right patient has become paramount. We believe baseline sequencing for the presence of resistance-associated substitutions (RAS) is an important contributor to this decision-making process. In the present study in an Irish cohort, we performed a retrospective analysis on all HCV samples received for drug resistance testing at the Irish National Virus Reference Laboratory between September 2014 and May 2016. Particular attention was paid to patients who experienced virological failure in an attempt to identify predictors of failure. Sanger sequence data covering the HCV NS3 protease coding region were obtained for 682 samples received during the study period. These were analysed using PAUP phylogenetic software. Sequence data for the NS5A and NS5B regions in some samples were also obtained. The rs12989860 single nucleotide polymorphism site was examined by allelic discrimination real-time PCR. Analysis of the NS3 viral sequences demonstrated that 85.5% (583/682) were HCV subtype 1a, 14.2% (97/682) subtype 1b and 0.3% (2/682) subtype 1c infections, subtype 1a was further differentiated into 76% clade 1 (443/583) and 24% clade 2 (140/583). RAS proven to reduce susceptibility to NS3 inhibitor treatment were detected in 45.9% of cases (313/682). Although the vast majority of all RAS detected were found in subtype 1a viruses, 7.2% (7/97) subtype 1b samples also contained one or more RAS. The Q80K polymorphism was found in 313/583 (57.3%) of HCV subtype 1a, and almost exclusively in clade 1 (242/443; 54.6%) versus clade 2 viruses (2/140; 1.4%). This distribution is reflected in the neighbour joining tree. Among the cohort of patients who experienced virological failure whilst on treatment, RAS could be detected in 11/17 (64.7%) patients for whom sequence could be generated. These included V36M/L (6/11; 54.5%), Q80K (5/11; 45.5%), R155K/T (3/11; 27.3%) and T54S (1/11; 9.1%). The majority of these patients were found to possess the deleterious “T” single nucleotide polymorphism (SNP) at the rs12989860 site within the Interferon lambda 4 (INFλ 4) gene locus. Nine of eleven patients with detected RAS were found to also be either CT or TT at rs12989860, one patient was CC at this SNP. Preliminary data from patients experiencing treatment failure on NS5A/B inhibitors also indicate the presence of RAS in 4 of 7 individuals. The high incidence of RAS within HCV NS3 protease sequences, the detection of RAS in NS5A sequences, and the apparent risk of treatment failure, albeit in a small number of patients, when the RAS are present, highlights the importance of sequencing these viruses prior to commencing treatment with protease inhibitors, and the need to identify additional predictors of failure.
Contributor Information
J. Dean, National Virus Reference Laboratory, University College Dublin, Belfield, Dublin 4, Ireland.
M. Neary, National Virus Reference Laboratory, University College Dublin, Belfield, Dublin 4, Ireland.
S. Coughlan, National Virus Reference Laboratory, University College Dublin, Belfield, Dublin 4, Ireland.
C.F. De Gascun, National Virus Reference Laboratory, University College Dublin, Belfield, Dublin 4, Ireland.
on Behalf of the ICORN Group, ICORN - Irish Hepatitis C Outcomes Research Network.