Functional RNA structures as targets in emerging arboviruses

Abstract

Emerging arthropod-borne viruses, such as Zika virus (ZIKV) and Chikungunya virus (CHIKV), represent a significant and increasing threat to global health. Despite the expanding prevalence of their vectors, the Aedes sp.mosquitoes, and the potential for major epidemics, there are currently no specific antiviral compounds or vaccines available for either viral pathogen. The positive-strand genomes of ZIKV and CHIKV, members of the flavivirus and alphavirus genera respectively, contain functional, structured cis-acting RNA elements. We are investigating a range of approaches for targeting these RNA elements and analysing the effect on virus replication at different stages of their life cycle. Gaining high resolution structural data is essential prior to targeting RNA elements, and consequently, we mapped RNA structural elements within the ZIKV 5’ genome region using a combination of biochemical SHAPE probing, thermodynamic predictions and phylogenetic analysis. We are currently validating our structural data by analysis of mutant phenotypes in a reverse genetic system. Using antisense locked nucleic acid oligonucleotides (antisense-LNA), we demonstrated that functional RNA elements in CHIKV can be specifically targeted – inhibiting replication in both sub-genomic replicon and infectious virus systems. Surface plasmon resonance confirmed that the antisense-LNA binds to a specific stem-loop target with aKd of 310 nM and has an IC50 of 35 nM in the sub-genomic replicon system. In future work, we aim to investigate selection of RNA-aptamers against CHIKV and target ZIKV genomic stem-loops using antisense-LNAs.