Production and characterization of high-titer serum-free cell culture grown hepatitis C virus particles of genotype 1-6

TY - JOUR

T1 - Production and characterization of high-titer serum-free cell culture grown hepatitis C virus particles of genotype 1-6

AU - Mathiesen, Christian K

AU - Jensen, Tanja B

AU - Prentoe, Jannick

AU - Krarup, Henrik

AU - Nicosia, Alfredo

AU - Law, Mansun

AU - Bukh, Jens

AU - Gottwein, Judith M

N1 - Copyright © 2014 Elsevier Inc. All rights reserved.

PY - 2014/6

Y1 - 2014/6

N2 - Recently, cell culture systems producing hepatitis C virus particles (HCVcc) were developed. Establishment of serum-free culture conditions is expected to facilitate development of a whole-virus inactivated HCV vaccine. We describe generation of genotype 1-6 serum-free HCVcc (sf-HCVcc) from Huh7.5 hepatoma cells cultured in adenovirus expression medium. Compared to HCVcc, sf-HCVcc showed 0.6-2.1 log10 higher infectivity titers (4.7-6.2 log10 Focus Forming Units/mL), possibly due to increased release and specific infectivity of sf-HCVcc. In contrast to HCVcc, sf-HCVcc had a homogeneous single-peak density profile. Entry of sf-HCVcc depended on HCV co-receptors CD81, LDLr, and SR-BI, and clathrin-mediated endocytosis. HCVcc and sf-HCVcc were neutralized similarly by chronic-phase patient sera and by human monoclonal antibodies targeting conformational epitopes. Thus, we developed serum-free culture systems producing high-titer single-density sf-HCVcc, showing similar biological properties as HCVcc. This methodology has the potential to advance HCV vaccine development and to facilitate biophysical studies of HCV.

AB - Recently, cell culture systems producing hepatitis C virus particles (HCVcc) were developed. Establishment of serum-free culture conditions is expected to facilitate development of a whole-virus inactivated HCV vaccine. We describe generation of genotype 1-6 serum-free HCVcc (sf-HCVcc) from Huh7.5 hepatoma cells cultured in adenovirus expression medium. Compared to HCVcc, sf-HCVcc showed 0.6-2.1 log10 higher infectivity titers (4.7-6.2 log10 Focus Forming Units/mL), possibly due to increased release and specific infectivity of sf-HCVcc. In contrast to HCVcc, sf-HCVcc had a homogeneous single-peak density profile. Entry of sf-HCVcc depended on HCV co-receptors CD81, LDLr, and SR-BI, and clathrin-mediated endocytosis. HCVcc and sf-HCVcc were neutralized similarly by chronic-phase patient sera and by human monoclonal antibodies targeting conformational epitopes. Thus, we developed serum-free culture systems producing high-titer single-density sf-HCVcc, showing similar biological properties as HCVcc. This methodology has the potential to advance HCV vaccine development and to facilitate biophysical studies of HCV.

KW - Cell Line, Tumor

KW - Cell Survival

KW - Culture Media, Serum-Free

KW - Gene Expression Regulation, Viral

KW - Genotype

KW - Hepacivirus

KW - Humans

KW - Virus Cultivation

U2 - 10.1016/j.virol.2014.03.021

DO - 10.1016/j.virol.2014.03.021

M3 - Journal article

C2 - 24928051

SN - 0042-6822

VL - 458-459

SP - 190

EP - 208

JO - Virology

JF - Virology

ER -