siRNA-mediated suppression of Japanese encephalitis virus replication in cultured cells and mice

Background: Japanese encephalitis virus (JEV) is a major cause of viral encephalitis in South-East Asia and there is a pressing need to develop novel therapeutic options against it. Methods: Gene silencing by RNA interference has therapeutic potential by way of degrading the RNA genome of JEV. Four small hairpin RNAs (shRNAs) targeting different locations in the JEV genome were evaluated for antiviral activity against JEV in different cell lines and the mouse model of disease. Results: shN8010, an shRNA targeting the NS5-coding sequence of JEV, had significant antiviral activity in cultured cells. JEV titres were suppressed by 99% in human embryonic kidney cells at 24 h post-infection (p.i.) when shN8010 was delivered using a plasmid. Further, shN8010 delivered using recombinant adenovirus caused 99% and 95% suppression of JEV titres at 24 h p.i. in porcine stable kidney and Vero cells, respectively. In Neuro-2a cells, JEV titres at 24 h p.i. were suppressed by 90% when shN8010 was delivered using a recombinant adenovirus or retrovirus. Four-week-old FvB/J mice treated intracerebrally with recombinant adenovirus-delivered shN8010 1 week prior to lethal intraperitoneal JEV challenge showed no protection, although the mean survival time was prolonged. In a similar experiment, retrovirus-delivered shN8010 provided 100% protection to mice following the lethal JEV challenge. Conclusions: NS5-targeting shRNA (shN8010) had very significant antiviral activity in both cultured cells and the mouse model of JEV infection.