A41 Characterization of NS5 coding region resistance associated substitutions from DAA-naïve GT1 HCV-infected patients in a Portuguese cohort

Abstract

Hepatitis C virus (HCV) is considered to be the leading cause of hepatocellular carcinoma (HCC) and other co-morbidities. During recent years, several highly effective regimens of directacting antivirals (DAAs) with excellent rates of success became available. However, therapeutic failure may occur in up to 10 per cent of treated individuals, and one of the main causes for this failure is the presence of resistance-associated substitutions (RASs) present before treatment initiation. Our aim was to study the profile and prevalence of baseline RASs in the NS5 coding region of DAA-naive GT1 HCV infected patients, and then understand the impact of the found RASs in the response to treatment by ascertaining an association between treatment failure and the presence of major NS5 RASs. Plasma RNA from eighty-one GT1 HCV infected patients was extracted using the NucliSensVR easyMAG system, followed by an in-house nested RT-PCR of the NS5 coding region. PCR products were purified and subsequently sequenced with the 3130xl ABI PRISM Genetic Analyzer. Sequences were finally aligned and edited using ChromasPro v1.7.6, and analyzed online with hcv.geno2pheno. org. NS5A RASs were present in 28.4 per cent (23/81) of all GT1 infected patients, with GT1a showing the highest prevalence followed by GT1b (17 vs. 11 per cent, respectively). Major NS5A RASs were detected in 23.6 per cent (13/55) of GT1a infected patients (M28V, Q30H/R, L31M, and Y93C/H) and in 15.4 per cent (4/26) of GT1b infected patients (L31M and Y93H). The most commonly detected NS5A RAS was Y93C/H with a prevalence of 9.9 per cent (8/81) in all GT1 infected patients, followed by L31M and Q30H/R with a prevalence of 8.6 per cent (7/81) and 6.2 per cent (5/81), respectively. Furthermore, Y93C/H showed a higher prevalence in GT1b patients than in GT1a, namely 11.5 per cent (3/26) vs. 9.1 per cent (5/55), respectively. NS5B RASs showed a prevalence of 14.8 per cent (12/81) in all GT1 infected patients, and were only detected in GT1b, being mainly represented by C316N with 38.5 per cent (10/26) of GT1b infected patients. The combined NS5A RASs (Q30H+Y93H), causing high level resistance to all NS5A inhibitors, were detected at baseline in one HIV/HCV GT1a co-infected patient who later failed a treatment with SOF+LDV for 12 weeks. Finally, an isolated Y93H mutation was also detected at baseline in a GT1b mono0-infected patient experiencing recurrence. Overall 38.3 per cent (31/81) of all GT1 HCV infected patients presented NS5 RASs at baseline, in which 58.1 per cent (18/31) were co-infected with HIV/HCV whereas only 38.7 per cent (12/31) of HCV monoinfected patients showed baseline RASs. Moreover, 27.3 per cent (15/55) of GT1a infected patients presented NS5 RASs at baseline, whereas patients infected with GT1b showed the highest prevalence of natural RASs, namely 61.5 per cent (16/26). These data support the usefulness of resistance testing prior to treatment initiation, thus preventing relapses associated to the presence of baseline RASs, as a statistical significant association was found between treatment failure and the presence of major NS5 RASs, namely Y93C/H (P=0.04). However, this reduced sampling can constitute a limiting factor since it may underestimate the statistical analysis, and lead to relatively higher RASs rates when comparing to other previous studies.