A Modular Design of a Cholera Toxin B Subunit-Scaffolded Sub-Virion Nanoparticle Vaccine Against West Nile Virus

Although veterinary vaccines against West Nile virus (WNV) have been developed, no approved human vaccine is currently available, highlighting the need for scalable and safer WNV vaccine candidates. In this study, a recombinant WNV subunit nanoparticle vaccine was developed by displaying the envelope protein domain III (ED3) on a cholera toxin B subunit (CTB) pentameric scaffold. The resulting recombinant protein comprising CTB-ED3 was expressed predominantly as soluble nanoparticles in Escherichia coli. Immunized mice produced strong humoral responses with balanced IgG1/IgG2a ratios, and some constructs achieved neutralizing titres comparable to those elicited by formalin-inactivated WNV. Importantly, no cross-reactivity with other flaviviruses was observed, alleviating potential concerns about ADE. These findings demonstrate that CTB-ED3 is assembled into multimeric nanoparticles in bacteria, offering a cost-effective, scalable, and biosafe platform for developing subunit nanoparticle vaccines against WNV and potentially other flaviviruses.