In silico post-hoc analysis of a clinically tested recombinant West Nile virus envelope protein vaccine

The recombinant West Nile virus (WNV) envelope (Env) protein WN80E is one of the few WNV antigens that has been evaluated in humans and has demonstrated robust immunogenicity across animal models and in a Phase I clinical trial. Here, we performed a retrospective in silico analysis of WN80E and its variant containing a N-terminal methionine (WNM80E) to assess how accurately contemporary computational vaccinology tools can capture and differentiate their structural and immunological profiles. Physicochemical and sequence-based predictions classified both constructs as antigenic, non-allergenic, and containing stable disulfide patterns, although WNM80E exhibited markedly improved predicted in vitro and in vivo half-lives. Tertiary structure modeling and subsequent refinement produced high-confidence homodimeric structures for both antigens, with favorable stereochemical metrics and preserved quaternary organization. Conformational B cell epitope mapping identified hinge-proximal and domain II antigenic patches with high solvent accessibility and minimal glycan shielding. Molecular docking with the broadly neutralizing monoclonal antibody CR4354 yielded energetically favorable complexes for both constructs, with slightly enhanced interface complementarity for WNM80E. Immune simulations predicted strong and durable humoral and cellular responses for both antigens, dominated by a Th1 signature, sustained memory formation, and repeated antigen clearance following booster doses. These findings demonstrate that results from in silico vaccinology tools further support continued evaluation of the clinically tested WN80E antigen in clinical trials and identify WNM80E as a structurally and immunologically comparable variant with modestly improved predicted stability and immunogenicity. This work highlights the utility of integrated computational pipelines for antigen evaluation.