Interplay of Amino Acid Residues at Positions 28 and 31 in NS5A Defines Resistance Pathways in Hepatitis C Virus Genotype 2

Abstract

Hepatitis C virus (HCV) genotype 2 (GT2) represents approximately 9% of all viral infections globally. While treatment outcomes for GT2-infected patients have improved substantially with direct-acting antiviral agents (DAAs) compared to alpha interferon, the presence of polymorphisms in NS5A can impact the efficacy of NS5A inhibitor-containing regimens. Thus, pathways of NS5A resistance were explored in GT2 subtypes using elbasvir, an NS5A inhibitor with broad genotype activity. Resistance selection studies, resistance analysis in NS5A-inhibitor treated virologic failures, and analyses of antiviral activities in replicons bearing a panel of GT2 subtype sequences and amino acid substitutions introduced by site-directed mutagenesis were performed to define determinants of inhibitor susceptibility. Elbasvir showed differential antiviral activities in replicons bearing GT2 sequences. The 50% effective concentration (EC50) values for replicons bearing reference NS5A sequences for GT2a and GT2b were 0.003 and 3.4 nM, respectively. Studies performed with recombinant replicons demonstrated cross talk between amino acid positions 28 and 31. The combination of phenylalanine and methionine at positions 28 and 31, respectively, conferred the highest potency reduction for elbasvir in GT2a and GT2b. This combination was observed in failures seen in the C-SCAPE trial. Addition of grazoprevir, an NS3/4A protease inhibitor, to elbasvir more effectively suppressed the emergence of resistance in GT2 at modest inhibitor concentrations (3× EC90). Ruzasvir, a potent, pan-genotype NS5A inhibitor, successfully inhibited replicons bearing GT2 resistance-associated substitutions (RASs) at positions 28 and 31. The results demonstrate that cross talk between amino acids at positions 28 and 31 in NS5A modulated inhibitor potency and may impact treatment outcomes in some HCV GT2-infected patients.

Cell Culture Studies of the Efficacy and Barrier to Resistance of Sofosbuvir-Velpatasvir and Glecaprevir-Pibrentasvir against Hepatitis C Virus Genotypes 2a, 2b, and 2c

“…Moreover, in this study, the T9(2a) strain harbored the double F28/M31 polymorphism at the baseline. This combined polymorphism has been shown to confer a significant reduction in the susceptibility to elbasvir in the replicon system (57); however, this combined polymorphism did not decrease the efficacy of the sofosbuvir-velpatasvir or uprifosbuvirelbasvir regimen in cell culture. Thus, virological determinants of the response to DAA combinations appear to be rather complex and might depend on multiple combinations of polymorphisms across drug targets.…”

Synthesis of Indole-Fused Six-, Seven-, or Eight-Membered N,O-Heterocycles via Rhodium-Catalyzed NH-Indole-Directed C–H Acetoxylation/Hydrolysis/Annulation

“…Indole-fused N,O-heterocycles are important fused heterocyclic motifs found in many pharmaceutically active compounds . For examples, MK-8742, 13-cyclohexyl-6,7-dihydrobenzo­[6,7]­[1,4]­oxazepino­[4,5- a ]­indole-10-carboxylic acid, and MK-3281 have been proven promising therapeutic agents for the treatment of hepatitis C virus (HCV), which has infected ∼3% of the world’s population (Scheme ). Therefore, efficient and concise synthesis of such skeletal analogues, which can offer a meaningful lead for the development of new molecular candidates for bioactivity studies, has attracted the attention of the synthetic community.…”