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P2043
Multiplexing HCV Replicon Luciferase Assay With Cell-Titer Blue Toxicity Assay In HTS Format
Presenter Sony Agrawal, Schering-Plough, USA
Additional Authors: Boris Feld
Hepatitis C virus is a major cause of chronic liver disease. Current treatments for hepatitis caused by HCV include interferon combination with ribavirin. Approximately, 50 to 60% of individuals still are not able to resolve infection. Even though, there are HCV Protease, Polymerase, and novel inhibitors in clinical trials at this time, there is still an unmet medical need to develop more effective therapies to treat HCV infection.
Basic research on HCV has been difficult because a lack of a traditional robust cell culture system. Although, recently a new HCV infectious particle system has been developed, it cannot be used as a screening tool. Currently an HCV subgenomic replicon cell cultured system is used as a surrogate cell-based model to study HCV replication. The replicon cell line that constitutively expresses an HCV subgenomic replicon as well as luciferase reporter enables the quantification of replication level by measuring not only RNA level (Taqman), but luciferase as well. Therefore, it makes a suitable tool for HTS screening. Accessing toxicity for establishing therapeutic window (activity vs. cytotoxicity) is required in cell-based screen. We were able to adapt HCV replicon luciferase assay to the 384 well format as well as multiplex with Cell-Titer Blue cytotoxicity assay for HTS. Assay validation and screening data will be presented.