Identification and characterisation of small molecule inhibitors targeted to the hepatitis C virus NS2 autoprotease.

Hepatitis C virus (HCV) is a positive-strand RNA virus present in 2-3% of the global population and commonly establishing a chronic infection, leading to long term diseases such as liver cirrhosis and hepatocellular carcinoma. Recent advances have led to the development of a range of direct-acting anti-viral drugs (DAAs), some of which are already improving outcomes in the clinic. It is clear however, that effective therapy for the treatment of HCV will most likely require a combination of DAAs to overcome the rapid onset of viral resistance. In this regard additional inhibitors of the virus lifecycle, which act through a novel molecular target, are required.
The autoprotease activity encoded within the C-terminus of the non-structural 2 (NS2) protein is essential for processing of a precursor to the mature viral proteins, and as a consequence is also required for the onset of viral genome replication and the establishment of HCV infection. Despite representing an attractive target for anti-virals, no inhibitors of the NS2 autoprotease have been reported.
In order to identify small molecule inhibitors of the NS2 autoprotease, two independent assays were optimised as a measure of NS2-mediated proteolysis. These assays were employed to demonstrate that inhibitors of the NS2 autoprotease were able to block HCV genome replication. The assays were subsequently used to identify a lead-like small molecule inhibitor by screening an in silico enriched library. This compound was further characterised in the context of NS2 activity in vitro and cell culture models of the virus lifecycle. The resultant series represent the first documented inhibitors capable of exerting an anti-viral effect by targeting the NS2 autoprotease.