Inactivated Japanese encephalitis virus vaccination imprints fusion loop-biased antibody responses that are attenuated by repeated live-attenuated dengue vaccination.

Immune imprinting, also known as immune history, is a core aspect of adaptive immunity that influences antibody responses to future antigen exposures. Nevertheless, the impact of sequential flavivirus vaccinations on epitope targeting and antibody activity in humans remains incompletely understood. This question is particularly important in regions where the inactivated Japanese encephalitis virus (JEV) vaccines and the live-attenuated dengue virus (DENV) vaccines are used, as both have been associated with an increased risk of symptomatic dengue infection and severe illness. We studied the impact of prior inactivated JEV IXIARO vaccination and simultaneous vaccination on humoral immunity following live-attenuated dengue CYD-TDV vaccination. Long-term analysis showed that JEV IXIARO priming guides the dengue vaccine-induced antibody response toward conserved fusion loop epitopes (FLEs) of the DENV envelope protein, as indicated by 4G2 FLE-bias. This imprinting was characterized by higher levels of 4G2 FLE-like antibodies, rapid recall responses after dengue vaccination, and broad but low-potency neutralization across dengue serotypes and Zika virus. Notably, 4G2 FLE-focused responses correlated with higher FcγRIIa-mediated antibody-dependent enhancement relative to neutralization potency, suggesting functional effects beyond neutralization. To better understand epitope dominance within the native envelope, we used a structurally defined fusion loop epitope mutant (FLE-mut) envelope dimer assay. Disrupting fusion loop accessibility significantly decreased antibody binding, confirming that FLE-specific antibodies are a major component of the response after sequential vaccination. Importantly, a complete series of live-attenuated dengue vaccine reduced 4G2 FLE bias, encouraged the recruitment of non-fusion-loop epitopes, and lessened FcγRIIa-biased antibody activity. Overall, these results show that vaccination platform, timing, and regimen are critical determinants of epitope dominance and antibody quality following flavivirus vaccination.