Potent Cross-Serotype Dengue Neutralization with Low ADE by AG-Strand-Targeting Anti-EDIII Antibodies

Dengue virus (DENV) envelope protein domain III (EDIII) is a premier target for vaccine design because EDIII-specific antibodies can potently neutralize DENV with a low propensity for inducing antibody-dependent enhancement (ADE). We previously identified a cross-reactive antibody, R3N_2D3, which targets the EDIII AG-strand epitope and neutralizes all four DENV serotypes with low ADE, a desirable combination of activities among anti-DENV antibodies. In this study, we expanded this AG-strand-targeting antibody panel by identifying four additional anti-EDIII antibodies from our dengue-immune single-chain variable fragment (scFv)-phage libraries. Despite variations in their binding footprints, these new antibodies converged on key residues within the AG-strand epitope. When expressed as full IgG1, these candidates showed potent cross-neutralization with subnanomolar focus reduction neutralization titers (FRNT₅₀ values) and consistently low ADE compared to the prototypical fusion-loop antibody 4G2 when tested in an in vitro U937 monocytic cell model. Collectively, these findings demonstrate that the AG-strand epitope can elicit broadly protective antibodies with reduced ADE risk, highlighting its potential as a strategic epitope for pan-DENV vaccine development.