Efficient Silencing of Hepatitis B Virus by Helper-dependent Adenovirus Vector-mediated Delivery of Artificial Antiviral Primary Micro RNAs

Mohube      Betty Mowa; Carol      Crowther; Abdullah      Ely; Patrick      Arbuthnot

doi:10.2174/2211536611201010019

Abstract

Hepatitis B virus (HBV) infection is endemic to southern Africa and parts of Asia where approximately 350 million individuals are chronically infected. Persistent infection increases risk for the serious complications of cirrhosis and hepatocellular carcinoma. Licensed HBV treatments rarely eradicate the virus, which makes developing new strategies for the treatment of chronic HBV a priority. Pol II-transcribed mono- and trimeric primary micro RNAs (primiRNAs) have previously been used to activate RNA interference (RNAi) and inhibit HBV gene expression, indicating that this approach holds promise for HBV therapy. Nevertheless, achieving safe and efficient delivery of anti-HBV RNAi expression cassettes remains an important objective before therapeutic application of this gene silencing technology is realized. Recombinant adenoviruses (Ads) are amongst the most efficient hepatotropic gene delivery vehicles, but a drawback of their use is transient transgene expression and toxicity that results from induction of host immune responses. To diminish immunostimulation of anti-HBV RNAi-activating vectors, helper-dependent (HD) Ads with all viral proteinencoding sequences removed from their genomes, were generated. A CMV Pol II promoter element was used to transcribe antiviral pri-miRNAs that target HBV. Processing of the anti-HBV pri-miRNA RNAi activators occurred according to intended design. Assessment in cultured cells and in a HBV transgenic model of the infection demonstrated that HD Ads delivered the silencing sequences efficiently and replication of the virus was inhibited without causing overt toxic effects. Collectively these data augur well for clinical use of HD Ads to deliver Pol II HBV-silencing cassettes to counter the persistent infection.

Keywords: HBV, Helper dependent adenovirus, Micro RNA, Pol II, RNAi, carcinoma, hepatocellular, adenoviruses, centrifugation, immunostimulatory.