Development of a Flow Cytometry Live Cell Assay for the Screening of Inhibitors of Hepatitis C Virus (HCV) Replication



Jose A Garcia-Rivera1, Kai Lin2, Sam Hopkins3, Matthew A Gregory4, Barrie Wilkinson4, Philippe A Gallay*, 1
1 Department of Immunology & Microbial Science, The Scripps Research Institute, La Jolla, California 92037, USA
2 Novartis Institutes for Biomedical Research, Inc., Cambridge, USA
3 SCYNEXIS, Inc., Research Triangle Park, USA
4 Biotica Technology Ltd, Chesterford Research Park, Cambridge, CB10 1XL, UK


Article Metrics

CrossRef Citations:
0
Total Statistics:

Full-Text HTML Views: 393
Abstract HTML Views: 228
PDF Downloads: 91
Total Views/Downloads: 712
Unique Statistics:

Full-Text HTML Views: 249
Abstract HTML Views: 133
PDF Downloads: 73
Total Views/Downloads: 455



© Garcia-Rivera et al.; Licensee Bentham Open.

open-access license: This is an open access article licensed under the terms of the Creative Commons Attribution Non-Commercial License (http: //creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted, non-commercial use, distribution and reproduction in any medium, provided the work is properly cited.

* Address correspondence to this author at the Department of Immunology & Microbial Science, IMM-9, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037, USA; Tel: (858) 784-8180; Fax: (858) 784-8831; E-mail: gallay@scripps.edu


Abstract

In this study, we established a flow cytometry live cell-based assay that permits the screening of hepatitis C virus (HCV) inhibitors. Specifically, we created a stable cell line, which harbors a subgenomic replicon encoding an NS5A-YFP fusion protein. This system allows direct measurement of YFP fluorescence in live hepatoma cells in which the HCV replicon replicates. We demonstrated that this stable fluorescent system permits the rapid and sensitive quantification of HCV replication inhibition by direct-acting antiviral agents (DAA) including protease and NS5A inhibitors and host-targeting antiviral agents (HTA) including cyclophilin inhibitors. This flow cytometry-based live cell assay is well suited for multiple applications such as the evaluation of HCV replication as well as antiviral drug screening.

Keywords: Direct-acting antiviral, FACS assay, HCV, hepatoma cells, host-targeting antiviral, inhibitors..