Targeted neuronal delivery of a single antiviralsiRNA protects mice against encephalitis caused by two distinct flaviviruses

Abstract

Japanese encephalitis (JE) and West Nile (WN) viruses are neurotropic flaviviruses that can cause acute encephalitis with a high degree of fatality. Infections by diverse neurotropic flaviviruses are clinically indistinguishable and currently there are no effective drugs to treat these infections. Thus it is important to develop therapeutic approaches that are effective against multiple viruses within and across the flaviviral species. In this study, we employed RNA interference (RNAi) targeting a cross-species conserved sequence in the flaviviral genome as a broad-based intervention against flavivirus infection. By using a lentiviral vector pseudotyped with the rabies virus glycoprotein (RVG) that enabled specific and targeted delivery of antiviral siRNA to neurons we were able to completely abrogate flaviviral replication in the neuroblastoma cell line, Neuro 2a. Moreover, a single administration of vector-delivered short-hairpin RNA was able to completely protect mice against a lethal intracranial challenge with as high as 50LD50 of virus. RVG-mediated targeting also resulted in better levels of protection compared to the conventional methods of delivery using the Vesicular Stomatitis virus glycoprotein, possibly due to better retro-axonal spread attributed to RVG. As a further demonstration of the feasibility of an RNAi-based therapy for viral encephalitis, we used cationic lipid formulations that mediate effective brain delivery of short interfering (si)RNA. A single intracranial application of lipid-complexed siRNA targeting conserved viral sequences before viral challenge and more importantly, as late as 18 hours after viral challenge was sufficient for protection of mice against lethal encephalitis induced by both JEV and WNV. These results demonstrate the practicality of using lipid-based delivery of single siRNA targeting a conserved sequence as a broad-based therapeutic strategy against multiple related viruses. Efforts are currently underway towards developing a nanoparticle-mediated approach for targeted brain delivery following peripheral administration of this common siRNA.