In vitro characterization of the activity of PF-05095808 a novel biological agent for Hepatitis C Virus therapy

PF-05095808 is a novel biological agent for chronic Hepatitis C Virus (HCV) therapy. It comprises a recombinant Adeno-associated virus (AAV) DNA vector packaged into an AAV serotype 8 capsid. The vector directs expression of 3 short hairpin (sh) RNAs targeted to conserved regions of the HCV genome. These shRNAs are processed by the host cell into the small interfering RNAs which mediate sequence specific cleavage of target regions. For small molecule inhibitors the key screens needed to assess in vitro activity are well defined, we developed new assays to assess this RNA interference agent and so understand its therapeutic potential. Following administration of PF-05095808 or corresponding synthetic shRNAs, sequence specific antiviral activity was observed in HCV replicon and infectious virus systems. To quantify the numbers of shRNA molecules required for antiviral activity in vitro and potentially also in vivo, a universal qPCR assay was developed. The number of shRNA molecules needed to drive antiviral activity proved independent of the vector delivery system used for PF-05095808 administration. The emergence of resistant variants at the target site of one shRNA was characterised. A novel RNA cleavage assay was developed to confirm the spectrum of activity of PF-05095808 against common HCV clinical isolates. In summary, our data supports both antiviral activity consistent with an RNA interference mechanism and demonstrates the potential of PF-05095808 as a therapeutic agent for chronic HCV infection.