Nucleic acid-based infectious and pseudoinfectious flavivirus vaccines

The genus Flavivirus contains a number of important pathogens of humans including yellow fever virus (YFV), dengue virus (DENV), Japanese encephalitis virus (JEV), tick-borne encephalitis virus (TBEV), and West Nile virus (WNV). Despite causing significant morbidity and mortality worldwide, commercially available vaccines only exist for YFV (live-attenuated), TBEV, and JEV (inactivated). Flavivirus vaccine research has been driven by the need for cheap, safe, thermally stable, and efficacious preparations amenable to use in developing nations. The creation of infectious cDNA clones of various flaviviruses has led to the development of genetically engineered, nucleic acid-delivered, attenuated live vaccine candidates. These provide effective immunity from a single immunisation, however share the same safety concerns as traditional live-attenuated vaccines. The generation of large internal deletions in the capsid gene of flavivirus genomes create; a vaccine that secretes large amounts of immunogenic prM/E particles from;elf-replicating RNA but does not form a spreading infection. Packaging of these capsid-deleted RNAs into virus-like particles (VLPs) using a cell line that produces capsid gene from another expression vector creates a pseudo-infectious vaccine that elicits a highly efficient immune response from a single dose and is safer than infectious virus. However, production of these VLPs is cumbersome and the resulting product is heat labile. Providing the capsid gene in trans from another promoter but within the same plasmid DNA as the capsid-deleted viral genome creates a DNA vaccine capable of producing VLPs in vivo. Uptake of this plasmid DNA results in the generation of self-replicating, capsid-deleted RNA and the capsid protein in the same cell, leading to production of secreted single-round infectious particles (SRIPs). These SRIPs then deliver capsid-deleted RNA to adjacent cells where it replicates to produce more prM/E particles. As functional capsid cannot be produced in SRIP-infected cells, further spread does not occur. SRIP-producing DNA was shown to be highly effective in mice and horses and provides an easier to manufacture and thermally stable alternative to other vaccine candidates currently being developed