Characterization of Live-Attenuated Powassan Virus Vaccine Candidates Identifies an Efficacious Prime-Boost Strategy for Mitigating Powassan Virus Disease in a Murine Model

Abstract

Powassan virus (POWV) is an emerging tick-borne virus and cause of lethal encephalitis in humans. The lack of treatment or prevention strategies for POWV disease underscores the need for an effective POWV vaccine. Here, we took two independent approaches to develop vaccine candidates. First, we recoded the POWV genome to increase the dinucleotide frequencies of CpG and UpA to potentially attenuate the virus by raising its susceptibility to host innate immune factors, such as the zinc-finger antiviral protein (ZAP). Secondly, we took advantage of the live-attenuated yellow fever virus vaccine 17D strain (YFV-17D) as a vector to express the structural genes pre-membrane (prM) and envelope (E) of POWV. The chimeric YFV-17D-POWV vaccine candidate was further attenuated for in vivo application by removing an N-linked glycosylation site within the nonstructural protein (NS)1 of YFV-17D. This live-attenuated chimeric vaccine candidate significantly protected mice from POWV disease, conferring a 70% survival rate after lethal challenge when administered in a homologous two-dose regimen. Importantly, when given in a heterologous prime-boost vaccination scheme, in which vaccination with the initial chimeric virus was followed by a protein boost with the envelope protein domain III (EDIII), 100% of the mice were protected without showing any signs of morbidity. Combinations of this live-attenuated chimeric YFV-17D-POWV vaccine candidate with an EDIII protein boost warrant further studies for the development of an effective vaccine strategy for the prevention of POWV disease.

Establishment of a CPER Reverse Genetics System for Powassan Virus Defines Attenuating NS1 Glycosylation Sites and an Infectious NS1-GFP11 Reporter Virus

“…Reverse genetics has also been used to generate chimeric FVs with prM-Env specific neutralization determinants, and NS protein backgrounds derived from attenuated FVs or the Yellow Fever Virus (YFV) 17D vaccine (31,45,51,67) . Insertion of the POWV prM-Env in a YFV-17D background retained lethal neurovirulence with only 43% of inoculated mice surviving this virulent chimera (50) .…”

“…Reverse genetics systems are needed to genetically modify POWVs, define determinants of POWV pathogenesis and create live attenuated recombinant POWV vaccines. Typical reverse genetics approaches are based on cloning viral RNA genomes into bacterial plasmids or artificial chromosome vectors with the toxicity and stability issues of large viral inserts that magnify the complexity of genetic virus modification (50)(51)(52)(53)(54)(55)(56) . The circular polymerase extension reaction (CPER) was originally devised to clone complex genes (57,58) and subsequently used to bypass the plasmid instability of large viral inserts and to rapidly generate mutant and reporter viruses (36,(59)(60)(61)(62)(63)(64) .…”

Immunoinformatics and computational approaches driven designing a novel vaccine candidate against Powassan virus

“…However, it's worth noting that one of the patients experienced considerable neurological complications following their recovery and discharge 22 . Recently, a vaccine candidate that uses the yellow fever virus vaccine strain as a vector expresses the prM/M and envelope proteins of POWV protected mice following lethal challenge and conferred a survival rate of 70% 23 .…”

“…Hence, in this study, a vaccine candidate has been developed using immunoinformatics approaches by targeting the structural proteins of POWV. Previously, several studies have targeted the Powassan structural proteins, mainly prM/M and envelope proteins, for developing vaccine candidates using strategies 19 , 23 , 24 . Lately, there has been a notable rise in the adoption of immunoinformatics in creating vaccine candidates.…”

Recapitulation of the Powassan virus life cycle in cell culture

“…This has raised the possibility that POWV, similar to other pathogens transmitted by Ixodes scapularis ticks, may be considered an emerging human disease in areas where enzootic cycles of POWV, and high populations of I. scapularis ticks coexist ( 4 – 6 ). To date, no prophylactic vaccine or direct antivirals are available to prevent or treat POWV infections ( 5 , 7 ). Therefore, it is urgent to understand the POWV viral life cycle and develop antiviral strategies.…”