Patent ID: 7276373

Claim:
A surrogate cell-based reporter system to evaluate the protease activity of Hepatitis C virus NS3 protease comprising a mammalian cell transfected with: a) a first chimeric DNA molecule comprising: i) a non-cytopathic expression system for inducing expression of said first chimera upon transfection in said mammalian cell; ii) an HCV recombinant DNA molecule operably linked to said expression system, said HCV recombinant DNA molecule encoding an NS3–5 polyprotein comprising: an active NS3 protease, an NS4A protein, wherein the NS4A protein allows embedding of the polyprotein in the endoplasmic reticulum (ER) membrane and acts as a co-factor for the NS3 protease activity, NS4B and NS5A proteins of sufficient length to allow correct structural orientation of the active NS3 protease relative to an NS5A/5B target cleavage site, and NS5B protein or a fragment thereof sufficient to provide said NS5A/5B cleavage site for said active NS3 protease; and iii) a transactivator domain fused downstream of said HCV recombinant DNA molecule, said transactivator domain encoding a transactivator molecule capable of initiating expression of a reporter gene; wherein said first chimeric DNA molecule encodes the polyprotein of SEQ ID NO:2; and b) a second chimeric DNA molecule encoding said reporter gene co-joined to an operon responding to said transactivator molecule; wherein expression of said first recombinant molecule leads to the production of a fusion polyprotein anchored to the endoplasmic reticulum of said mammalian cell, said fusion polyprotein being cleaved by said active NS3 protease thereby allowing translocation of said transactivator domain for inducing expression of said reporter gene as a means to evaluate said active NS3 protease activity.