Abstract
Hepatitis C virus (HCV) frequently establishes persistent infections that can develop into severe liver disease. The HCV NS3/4A serine protease is not only essential for viral replication but also cleaves multiple cellular targets that block downstream interferon activation. Therefore, NS3/4A is an ideal target for the development of anti-HCV drugs and inhibitors. In the current study, we generated a novel NS3/4A/Lap/LC-1 triple-transgenic mouse model that can be used to evaluate and screen NS3/4A protease inhibitors. The NS3/4A protease could be conditionally inducibly expressed in the livers of the triple-transgenic mice using a dual Tet-On and Cre/loxP system. In this system, doxycycline (Dox) induction resulted in the secretion of Gaussia luciferase (Gluc) into the blood, and this secretion was dependent on NS3/4A protease-mediated cleavage at the 4B5A junction. Accordingly, NS3/4A protease activity could be quickly assessed in real time simply by monitoring Gluc activity in plasma. The results from such monitoring showed a 70-fold increase in Gluc activity levels in plasma samples collected from the triple-transgenic mice after Dox induction. Additionally, this enhanced plasma Gluc activity was well correlated with the induction of NS3/4A protease expression in the liver. Following oral administration of the commercial NS3/4A-specific inhibitors telaprevir and boceprevir, plasma Gluc activity was reduced by 50% and 65%, respectively. Overall, our novel transgenic mouse model offers a rapid real-time method to evaluate and screen potential NS3/4A protease inhibitors.
Highlights
At least 150 million people are chronically infected with hepatitis C virus (HCV) worldwide
Mouse Model for Rapid Real-Time Detection of HCV NS3/4A Protease subsequently cleaved into three structural proteins and seven nonstructural (NS) proteins (p7, NS2, NS3, NS4A, NS4B, NS5A and NS5B) through the actions of two host proteases and two viral proteases (NS2 and NS3) [4]
After the administration of Dox, the reverse tetracycline-controlled transactivator (rtTA) in the vector undergoes nuclear translocation and activates Ptetbi-1, which induces the expression of Cre and the firefly luciferase (Fluc) reporter gene
Summary
At least 150 million people are chronically infected with hepatitis C virus (HCV) worldwide. The majority of these individuals are at significant risk of developing severe liver diseases, including liver fibrosis, liver cirrhosis, and hepatocellular carcinoma [1,2]. Mouse Model for Rapid Real-Time Detection of HCV NS3/4A Protease subsequently cleaved into three structural proteins (core, E1 and E2) and seven nonstructural (NS) proteins (p7, NS2, NS3, NS4A, NS4B, NS5A and NS5B) through the actions of two host proteases and two viral proteases (NS2 and NS3) [4]. Using NS4A as a co-factor, the NS3/4A serine protease is vital for viral replication [3] and cleaves multiple cellular targets that block downstream interferon activation [3,6]. No small animal model is available to test NS3/4A inhibitors in vivo that provides rapid, real time, and reproducible results
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