Recombinant hepatitis C virus genotype 5a infectious cell culture systems expressing minimal JFH1 NS5B sequences permit polymerase inhibitor studies

TY - JOUR

T1 - Recombinant hepatitis C virus genotype 5a infectious cell culture systems expressing minimal JFH1 NS5B sequences permit polymerase inhibitor studies

AU - Humes, Daryl

AU - Ramirez, Santseharay

AU - Jensen, Tanja B.

AU - Li, Yi Ping

AU - Gottwein, Judith M.

AU - Bukh, Jens

PY - 2018

Y1 - 2018

N2 - The six major epidemiologically important hepatitis C virus (HCV) genotypes differ in global distribution and antiviral responses. Full-length infectious cell-culture adapted clones, the gold standard for HCV studies in vitro, are missing for genotypes 4 and 5. To address this challenge for genotype 5, we constructed a consensus full-length clone of strain SA13 (SA13fl), which was found non-viable in Huh7.5 cells. Step-wise adaptation of SA13fl-based recombinants, beginning with a virus encoding the NS5B-thumb domain and 3´UTR of JFH1 (SA13/JF372-X), resulted in a high-titer SA13 virus with only 41 JFH1-encoded NS5B-thumb residues (SA13/JF470-510cc); this required sixteen cell-culture adaptive substitutions within the SA13fl polyprotein and two 3´UTR-changes. SA13/JF372-X and SA13/JF470-510cc were equally sensitive to nucleoside polymerase inhibitors, including sofosbuvir, but showed differential sensitivity to inhibitors targeting the NS5B palm or thumb. SA13/JF470-510cc represents a model to elucidate the influence of HCV RNA elements on viral replication and map determinants of sensitivity to polymerase inhibitors.

AB - The six major epidemiologically important hepatitis C virus (HCV) genotypes differ in global distribution and antiviral responses. Full-length infectious cell-culture adapted clones, the gold standard for HCV studies in vitro, are missing for genotypes 4 and 5. To address this challenge for genotype 5, we constructed a consensus full-length clone of strain SA13 (SA13fl), which was found non-viable in Huh7.5 cells. Step-wise adaptation of SA13fl-based recombinants, beginning with a virus encoding the NS5B-thumb domain and 3´UTR of JFH1 (SA13/JF372-X), resulted in a high-titer SA13 virus with only 41 JFH1-encoded NS5B-thumb residues (SA13/JF470-510cc); this required sixteen cell-culture adaptive substitutions within the SA13fl polyprotein and two 3´UTR-changes. SA13/JF372-X and SA13/JF470-510cc were equally sensitive to nucleoside polymerase inhibitors, including sofosbuvir, but showed differential sensitivity to inhibitors targeting the NS5B palm or thumb. SA13/JF470-510cc represents a model to elucidate the influence of HCV RNA elements on viral replication and map determinants of sensitivity to polymerase inhibitors.

KW - DAA

KW - Direct acting antivirals

KW - Genotype 5a

KW - HCV

KW - Hepatitis C virus

KW - Infectious cell-culture

KW - Liver Cancer

KW - Polymerase inhibitors

KW - Replication

U2 - 10.1016/j.virol.2018.05.020

DO - 10.1016/j.virol.2018.05.020

M3 - Journal article

C2 - 30032031

AN - SCOPUS:85050079569

SN - 0042-6822

VL - 522

SP - 177

EP - 192

JO - Virology

JF - Virology

ER -