HCV genotype 1-6 NS3 residue 80 substitutions impact protease inhibitor activity and promote viral escape

Long V. Pham; Sanne Brun Jensen; Ulrik Fahnøe; Martin Schou Pedersen; Qi Tang; Lubna Ghanem; Santseharay Ramirez; Daryl Humes; Stéphanie B.N. Serre; Kristian Schønning; Jens Bukh; Judith M. Gottwein

doi:10.1016/j.jhep.2018.10.031

HCV genotype 1-6 NS3 residue 80 substitutions impact protease inhibitor activity and promote viral escape

Long V. Pham, Sanne Brun Jensen, Ulrik Fahnøe, Martin Schou Pedersen, Qi Tang, Lubna Ghanem, Santseharay Ramirez, Daryl Humes, Stéphanie B.N. Serre, Kristian Schønning, Jens Bukh, Judith M. Gottwein^*

^*Corresponding author for this work

Department of Clinical Medicine

17 Citations (Scopus)

Abstract

Background & Aims: Protease inhibitors (PIs) are of central importance in the treatment of patients with chronic hepatitis C virus (HCV) infection. HCV NS3 protease (NS3P) position 80 displays polymorphisms associated with resistance to the PI simeprevir for HCV genotype 1a. We investigated the effects of position-80-substitutions on fitness and PI-resistance for HCV genotypes 1-6, and analyzed evolutionary mechanisms underlying viral escape mediated by pre-existing Q80K. Methods: The fitness of infectious NS3P recombinants of HCV genotypes 1-6, with engineered position-80-substitutions, was studied by comparison of viral spread kinetics in Huh-7.5 cells in culture. Median effective concentration (EC50) and fold resistance for PIs simeprevir, asunaprevir, paritaprevir, grazoprevir, glecaprevir and voxilaprevir were determined in short-term treatment assays. Viral escape was studied by long-term treatment of genotype 1a recombinants with simeprevir, grazoprevir, glecaprevir and voxilaprevir and of genotype 3a recombinants with glecaprevir and voxilaprevir, next generation sequencing, NS3P substitution linkage and haplotype analysis. Results: Among tested PIs, only glecaprevir and voxilaprevir showed pan-genotypic activity against the original genotype 1-6 culture viruses. Variants with position-80-substitutions were all viable, but fitness depended on the specific substitution and the HCV isolate. Q80K conferred resistance to simeprevir across genotypes but had only minor effects on the activity of the remaining PIs. For genotype 1a, pre-existing Q80K mediated accelerated escape from simeprevir, grazoprevir and to a lesser extent glecaprevir, but not voxilaprevir. For genotype 3a, Q80K mediated accelerated escape from glecaprevir and voxilaprevir. Escape was mediated by rapid and genotype-, PI- and PI-concentration-dependent co-selection of clinically relevant resistance associated substitutions. Conclusions: Position-80-substitutions had relatively low fitness cost and the potential to promote HCV escape from clinically relevant PIs in vitro, despite having a minor impact on results in classical short-term resistance assays. Lay summary: Among all clinically relevant hepatitis C virus protease inhibitors, voxilaprevir and glecaprevir showed the highest and most uniform activity against cell culture infectious hepatitis C virus with genotype 1-6 proteases. Naturally occurring amino acid changes at protease position 80 had low fitness cost and influenced sensitivity to simeprevir, but not to other protease inhibitors in short-term treatment assays. Nevertheless, the pre-existing change Q80K had the potential to promote viral escape from protease inhibitors during long-term treatment by rapid co-selection of additional resistance changes, detected by next generation sequencing.

Original language	English
Journal	Journal of Hepatology
Volume	70
Issue number	3
Pages (from-to)	388-397
ISSN	0168-8278
DOIs	https://doi.org/10.1016/j.jhep.2018.10.031
Publication status	Published - Mar 2019

Keywords

Asunaprevir
Direct-acting antiviral
Glecaprevir
Grazoprevir
Linkage analysis
Paritaprevir
Q80K
Resistance
Resistance associated substitutions
Simeprevir
Voxilaprevir, Hepatitis C virus

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1016/j.jhep.2018.10.031

Cite this

@article{bcebd52dadb5418195dfb9c5195fd026,

title = "HCV genotype 1-6 NS3 residue 80 substitutions impact protease inhibitor activity and promote viral escape",

abstract = "Background & Aims: Protease inhibitors (PIs) are of central importance in the treatment of patients with chronic hepatitis C virus (HCV) infection. HCV NS3 protease (NS3P) position 80 displays polymorphisms associated with resistance to the PI simeprevir for HCV genotype 1a. We investigated the effects of position-80-substitutions on fitness and PI-resistance for HCV genotypes 1-6, and analyzed evolutionary mechanisms underlying viral escape mediated by pre-existing Q80K. Methods: The fitness of infectious NS3P recombinants of HCV genotypes 1-6, with engineered position-80-substitutions, was studied by comparison of viral spread kinetics in Huh-7.5 cells in culture. Median effective concentration (EC50) and fold resistance for PIs simeprevir, asunaprevir, paritaprevir, grazoprevir, glecaprevir and voxilaprevir were determined in short-term treatment assays. Viral escape was studied by long-term treatment of genotype 1a recombinants with simeprevir, grazoprevir, glecaprevir and voxilaprevir and of genotype 3a recombinants with glecaprevir and voxilaprevir, next generation sequencing, NS3P substitution linkage and haplotype analysis. Results: Among tested PIs, only glecaprevir and voxilaprevir showed pan-genotypic activity against the original genotype 1-6 culture viruses. Variants with position-80-substitutions were all viable, but fitness depended on the specific substitution and the HCV isolate. Q80K conferred resistance to simeprevir across genotypes but had only minor effects on the activity of the remaining PIs. For genotype 1a, pre-existing Q80K mediated accelerated escape from simeprevir, grazoprevir and to a lesser extent glecaprevir, but not voxilaprevir. For genotype 3a, Q80K mediated accelerated escape from glecaprevir and voxilaprevir. Escape was mediated by rapid and genotype-, PI- and PI-concentration-dependent co-selection of clinically relevant resistance associated substitutions. Conclusions: Position-80-substitutions had relatively low fitness cost and the potential to promote HCV escape from clinically relevant PIs in vitro, despite having a minor impact on results in classical short-term resistance assays. Lay summary: Among all clinically relevant hepatitis C virus protease inhibitors, voxilaprevir and glecaprevir showed the highest and most uniform activity against cell culture infectious hepatitis C virus with genotype 1-6 proteases. Naturally occurring amino acid changes at protease position 80 had low fitness cost and influenced sensitivity to simeprevir, but not to other protease inhibitors in short-term treatment assays. Nevertheless, the pre-existing change Q80K had the potential to promote viral escape from protease inhibitors during long-term treatment by rapid co-selection of additional resistance changes, detected by next generation sequencing.",

keywords = "Asunaprevir, Direct-acting antiviral, Glecaprevir, Grazoprevir, Linkage analysis, Paritaprevir, Q80K, Resistance, Resistance associated substitutions, Simeprevir, Voxilaprevir, Hepatitis C virus",

author = "Pham, {Long V.} and Jensen, {Sanne Brun} and Ulrik Fahn{\o}e and Pedersen, {Martin Schou} and Qi Tang and Lubna Ghanem and Santseharay Ramirez and Daryl Humes and Serre, {St{\'e}phanie B.N.} and Kristian Sch{\o}nning and Jens Bukh and Gottwein, {Judith M.}",

year = "2019",

month = mar,

doi = "10.1016/j.jhep.2018.10.031",

language = "English",

volume = "70",

pages = "388--397",

journal = "Journal of Hepatology",

issn = "0168-8278",

publisher = "Elsevier",

number = "3",

}

TY - JOUR

T1 - HCV genotype 1-6 NS3 residue 80 substitutions impact protease inhibitor activity and promote viral escape

AU - Pham, Long V.

AU - Jensen, Sanne Brun

AU - Fahnøe, Ulrik

AU - Pedersen, Martin Schou

AU - Tang, Qi

AU - Ghanem, Lubna

AU - Ramirez, Santseharay

AU - Humes, Daryl

AU - Serre, Stéphanie B.N.

AU - Schønning, Kristian

AU - Bukh, Jens

AU - Gottwein, Judith M.

PY - 2019/3

Y1 - 2019/3

N2 - Background & Aims: Protease inhibitors (PIs) are of central importance in the treatment of patients with chronic hepatitis C virus (HCV) infection. HCV NS3 protease (NS3P) position 80 displays polymorphisms associated with resistance to the PI simeprevir for HCV genotype 1a. We investigated the effects of position-80-substitutions on fitness and PI-resistance for HCV genotypes 1-6, and analyzed evolutionary mechanisms underlying viral escape mediated by pre-existing Q80K. Methods: The fitness of infectious NS3P recombinants of HCV genotypes 1-6, with engineered position-80-substitutions, was studied by comparison of viral spread kinetics in Huh-7.5 cells in culture. Median effective concentration (EC50) and fold resistance for PIs simeprevir, asunaprevir, paritaprevir, grazoprevir, glecaprevir and voxilaprevir were determined in short-term treatment assays. Viral escape was studied by long-term treatment of genotype 1a recombinants with simeprevir, grazoprevir, glecaprevir and voxilaprevir and of genotype 3a recombinants with glecaprevir and voxilaprevir, next generation sequencing, NS3P substitution linkage and haplotype analysis. Results: Among tested PIs, only glecaprevir and voxilaprevir showed pan-genotypic activity against the original genotype 1-6 culture viruses. Variants with position-80-substitutions were all viable, but fitness depended on the specific substitution and the HCV isolate. Q80K conferred resistance to simeprevir across genotypes but had only minor effects on the activity of the remaining PIs. For genotype 1a, pre-existing Q80K mediated accelerated escape from simeprevir, grazoprevir and to a lesser extent glecaprevir, but not voxilaprevir. For genotype 3a, Q80K mediated accelerated escape from glecaprevir and voxilaprevir. Escape was mediated by rapid and genotype-, PI- and PI-concentration-dependent co-selection of clinically relevant resistance associated substitutions. Conclusions: Position-80-substitutions had relatively low fitness cost and the potential to promote HCV escape from clinically relevant PIs in vitro, despite having a minor impact on results in classical short-term resistance assays. Lay summary: Among all clinically relevant hepatitis C virus protease inhibitors, voxilaprevir and glecaprevir showed the highest and most uniform activity against cell culture infectious hepatitis C virus with genotype 1-6 proteases. Naturally occurring amino acid changes at protease position 80 had low fitness cost and influenced sensitivity to simeprevir, but not to other protease inhibitors in short-term treatment assays. Nevertheless, the pre-existing change Q80K had the potential to promote viral escape from protease inhibitors during long-term treatment by rapid co-selection of additional resistance changes, detected by next generation sequencing.

AB - Background & Aims: Protease inhibitors (PIs) are of central importance in the treatment of patients with chronic hepatitis C virus (HCV) infection. HCV NS3 protease (NS3P) position 80 displays polymorphisms associated with resistance to the PI simeprevir for HCV genotype 1a. We investigated the effects of position-80-substitutions on fitness and PI-resistance for HCV genotypes 1-6, and analyzed evolutionary mechanisms underlying viral escape mediated by pre-existing Q80K. Methods: The fitness of infectious NS3P recombinants of HCV genotypes 1-6, with engineered position-80-substitutions, was studied by comparison of viral spread kinetics in Huh-7.5 cells in culture. Median effective concentration (EC50) and fold resistance for PIs simeprevir, asunaprevir, paritaprevir, grazoprevir, glecaprevir and voxilaprevir were determined in short-term treatment assays. Viral escape was studied by long-term treatment of genotype 1a recombinants with simeprevir, grazoprevir, glecaprevir and voxilaprevir and of genotype 3a recombinants with glecaprevir and voxilaprevir, next generation sequencing, NS3P substitution linkage and haplotype analysis. Results: Among tested PIs, only glecaprevir and voxilaprevir showed pan-genotypic activity against the original genotype 1-6 culture viruses. Variants with position-80-substitutions were all viable, but fitness depended on the specific substitution and the HCV isolate. Q80K conferred resistance to simeprevir across genotypes but had only minor effects on the activity of the remaining PIs. For genotype 1a, pre-existing Q80K mediated accelerated escape from simeprevir, grazoprevir and to a lesser extent glecaprevir, but not voxilaprevir. For genotype 3a, Q80K mediated accelerated escape from glecaprevir and voxilaprevir. Escape was mediated by rapid and genotype-, PI- and PI-concentration-dependent co-selection of clinically relevant resistance associated substitutions. Conclusions: Position-80-substitutions had relatively low fitness cost and the potential to promote HCV escape from clinically relevant PIs in vitro, despite having a minor impact on results in classical short-term resistance assays. Lay summary: Among all clinically relevant hepatitis C virus protease inhibitors, voxilaprevir and glecaprevir showed the highest and most uniform activity against cell culture infectious hepatitis C virus with genotype 1-6 proteases. Naturally occurring amino acid changes at protease position 80 had low fitness cost and influenced sensitivity to simeprevir, but not to other protease inhibitors in short-term treatment assays. Nevertheless, the pre-existing change Q80K had the potential to promote viral escape from protease inhibitors during long-term treatment by rapid co-selection of additional resistance changes, detected by next generation sequencing.

KW - Asunaprevir

KW - Direct-acting antiviral

KW - Glecaprevir

KW - Grazoprevir

KW - Linkage analysis

KW - Paritaprevir

KW - Q80K

KW - Resistance

KW - Resistance associated substitutions

KW - Simeprevir

KW - Voxilaprevir, Hepatitis C virus

U2 - 10.1016/j.jhep.2018.10.031

DO - 10.1016/j.jhep.2018.10.031

M3 - Journal article

C2 - 30395912

AN - SCOPUS:85059647622

SN - 0168-8278

VL - 70

SP - 388

EP - 397

JO - Journal of Hepatology

JF - Journal of Hepatology

IS - 3

ER -

HCV genotype 1-6 NS3 residue 80 substitutions impact protease inhibitor activity and promote viral escape

Abstract

Keywords

UN SDGs

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