Immunogenicity of a West Nile virus DIII-cholera toxin A2/B chimera after intranasal delivery.

Juliette Tinker,Panos Panayiotou,Jie Yan,Kenneth Cornell,Reece Knippel

doi:10.3390/toxins6041397

Abstract

West Nile virus (WNV) causes potentially fatal neuroinvasive disease and persists at endemic levels in many parts of the world. Despite advances in our understanding of WNV pathogenesis, there remains a significant need for a human vaccine. The domain III (DIII) region of the WNV envelope protein contains epitopes that are the target of neutralizing antibodies. We have constructed a chimeric fusion of the non-toxic cholera toxin (CT) CTA2/B domains to DIII for investigation as a novel mucosally-delivered WNV vaccine. Purification and assembly of the chimera, as well as receptor-binding and antigen delivery, were verified by western blot, GM1 ELISA and confocal microscopy. Groups of BALB/c mice were immunized intranasally with DIII-CTA2/B, DIII, DIII mixed with CTA2/B, or CTA2/B control, and boosted at 10 days. Analysis of serum IgG after 14 and 45 days revealed that mucosal immunization with DIII-CTA2/B induced significant DIII-specific humoral immunity and drove isotype switching to IgG2a. The DIII-CTA2/B chimera also induced antigen-specific IgM and IgA responses. Bactericidal assays indicate that the DIII-CTA2/B immunized mice produced DIII-specific antibodies that can trigger complement-mediated killing. A dose escalation resulted in increased DIII-specific serum IgG titers on day 45. DIII antigen alone, in the absence of adjuvant, also induced significant systemic responses after intranasal delivery. Our results indicate that the DIII-CTA2/B chimera is immunogenic after intranasal delivery and merits further investigation as a novel WNV vaccine candidate.

Highlights

West Nile virus (WNV) is a positive, single-stranded RNA virus belonging to the arthropod-borne genus Flavivirus
This plasmid, constructed from the parental vector pARLDR19, utilizes E. coli LTIIB leader sequences to direct expression of the domain III (DIII)-CTA2 fusion protein and monomeric CTB to the periplasm
We have demonstrated that the DIII-CTA2/B chimera can be expressed and purified efficiently from E. coli and that this molecule retains the ability to bind to mammalian host cells in vitro and internalize antigen

Summary

Introduction

West Nile virus (WNV) is a positive, single-stranded RNA virus belonging to the arthropod-borne genus Flavivirus. In 2012, there was a dramatic upsurge in WNV transmission in the U.S resulting in the highest mortality rate recorded in this country, with 2873 cases of neuroinvasive disease and 286 deaths [3,4]. This resurgence is compelling evidence that the virus will remain endemic in the U.S and continue to trigger sporadic epidemics. Humoral immunity is essential for viral clearance, and neutralizing IgG and IgM antibodies against WNV surface envelope (E) and premembrane (prM) proteins constitute a majority of the protective response [10,14,16,17,18]. E protein is subdivided into three domains with the surface-exposed domain III (DIII) producing the most potent neutralizing antibodies [21,22,23]

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