A cell-based bicistronic lentiviral reporter system for identification of inhibitors of the hepatitis C virus internal ribosome entry site

作者：Lourenco Sofia; Boni Sebastien; Furling Denis; Cosset Francois Loic; Cahour Annie^*

来源：Journal of Virological Methods, 2009, 158(1-2): 152-159.

DOI：10.1016/j.jviromet.2009.02.007

• SIN lentiviral vector
• SIV
• HCV IRES inhibitors
• Reporters
• Gene therapy

This report describes the development, optimization and implementation of a persistent cell-based system to test inhibitors of hepatitis C (HCV) translation. The assay is based on a heterologous human immunodeficiency virus-1/simian immunodeficiency virus (HIV-1/SIV) lentiviral vector expressing the bicistronic cassette containing the firefly and renilla luciferase genes, respectively, as reporters, and the HCV internal ribosome entry site (IRES) inserted in between, under the control of the cytomegalovirus (CMV) promoter. The drug target in this assay is the HCV IRES, the activity of which leads to modulation of the renilla luciferase gene expression under its control, which is monitored by luminometry. The system has been validated using interferon (IFN), which is still the only consensual antiviral agent against HCV infection, associated with ribavirin. This bicistronic vector, extended to other viral IRESs and assayed in different cell lines, exhibited weak cell tropism, allowing its broad use in gene therapy, which frequently needs a multicistronic transfer vector to follow the expression of a gene of interest inside the target cells with the aid of a reporter, a drug selection marker, or a suicide gene, expressed from the same transcript.

• 出版日期2009-6