Characterization of NS5A and NS5B resistance-associated substitutions from genotype 1 HCV infected patients in a Portuguese cohort

Hepatitis C virus (HCV) is considered to be the leading cause of hepatocellular carcinoma (HCC). During the last years, several highly efficacy regimens of direct-acting antivirals (DAAs) with excellent rates of success became available. However, therapeutic failure may occur in up to 10% of treated individuals. Our aim was to study the profile of NS5 coding region RASs in DAA-naive genotype 1 HCV infected patients, as well as to ascertain an association between treatment failure and the presence of baseline NS5 RASs. A comparison between LiPA and Sanger sequencing genotyping methods was also assessed. Plasma RNA from 81 DAA-naïve GT1 HCV infected patients was extracted, followed by an in-house nested RT-PCR of the NS5 coding region. PCR products were purified, leading to Sanger population sequencing on the 3130xl ABI Genetic Analyzer. Sequences were aligned using ChromasPro v1.7.6, and analyzed online in hcv.geno2pheno.org. NS5A RASs were present in 28,4% (23/81) of all GT1 infected patients. The most commonly detected NS5A RAS was Y93C/H with a prevalence of 9,9% (8/81) in all GT1 infected patients. NS5B RASs showed a prevalence of 14,8% (12/81) in all GT1 infected patients, and were only detected in GT1b, being mainly represented by C316N accounting for 40% (10/25). The combined Q30H+Y93H NS5A RASs, were detected at baseline in one HIV/HCV GT1a co-infected patient who later failed a treatment with sofosbuvir/ledipasvir (SOF/LDV) for 12 weeks. An isolated Y93H mutation was also detected at baseline in a relapsing GT1b mono-infected patient. Overall 38,3% (31/81) of all GT1 HCV infected patients presented NS5 RASs at baseline, in which 58% (18/31) were co-infected with HIV/HCV. The obtained data supports the usefulness of resistance testing prior to treatment initiation, as a statistical significant association was found between treatment failure and the baseline presence of specific NS5 RASs, namely Y93C/H (p = 0.04)