The highly conserved 5′ untranslated region as an effective target towards the inhibition of Enterovirus 71 replication by unmodified and appropriate 2′-modified siRNAs

Abstract

Background: Enterovirus 71 (EV71) is a highly infectious agent that plays an etiological role in hand, foot, and mouth disease. It is associated with severe neurological complications and has caused significant mortalities in recent large-scale outbreaks. Currently, no effective vaccine or specific clinical therapy is available against EV71. Methods: Unmodified 21 nucleotide small interfering RNAs (siRNAs) and classic 2′-modified (2′-O-methylation or 2′-fluoro modification) siRNAs were designed to target highly conserved 5′ untranslated region (UTR) of the EV71 genome and employed as anti-EV71 agents. Real-time TaqMan RT-PCR, western blot analysis and plaque assays were carried out to evaluate specific viral inhibition by the siRNAs. Results: Transfection of rhabdomyosarcoma (RD) cells with siRNAs targeting the EV71 genomic 5′ UTR significantly delayed and alleviated the cytopathic effects of EV71 infection, increased cell viability in EV71-infected RD cells. The inhibitory effect on EV71 replication was sequence-specific and dosage-dependent, with significant corresponding decreases in viral RNA, VP1 protein and viral titer. Appropriate 2′-modified siRNAs exhibited similar RNA interference (RNAi) activity with dramatically increased serum stability in comparison with unmodified counterparts. Conclusion: Sequences were identified within the highly conserved 5′ UTR that can be targeted to effectively inhibit EV71 replication through RNAi strategies. Appropriate 2′-modified siRNAs provide a promising approach to optimizing siRNAs to overcome barriers on RNAi-based antiviral therapies for broader administration. © 2012 Deng et al.; licensee BioMed Central Ltd.