Strategies for next-generation flavivirus vaccine development

Dengue virus (DENV; DV) is the most common mosquito-borne virus in the world, with 4 billion people living in regions that put them at risk for infection by any of its four serotypes (DV1-4). Infection by DV and its related flavivirus Zika virus (ZIKV; ZV) may clinically manifest as mild flu-like symptoms, rash, and arthralgia. Severe DV cases can progress into hemorrhagic fever and shock syndrome, while severe complications of ZV infection include Guillain-Barre syndrome in adults and neonatal microcephaly. Severe dengue is associated with antibody-dependent enhancement (ADE) of infection, a phenomenon seen as potentially fatal. In response to a primary DV infection, the immune system produces antibodies that can bind and neutralize that serotype to resolve infection. However, during a subsequent infection by a heterotypic DV serotype or ZV, some antibodies from the primary infection are re-elicited but may poorly neutralize the heterotypic infection; thus, instead of eliminating infection, these antibodies promote uptake of active virus into Fcγ receptor-expressing immune cells and increase viremia and levels of pro-inflammatory cytokines. Currently, there is a need for vaccines and anti- viral therapies for DV and ZV that do not carry the risk of inducing ADE. To avoid ADE, a significant goal for DV and ZV vaccine development is to elicit broadly protective antibodies, as opposed to broadly reactive, non- neutralizing ones. We determined two glycoproteins expressed by DV and ZV as favorable candidates for immunogen design. Neutralizing, protective antibodies against DV and ZV have shown to target domain III (DIII) of E glycoprotein, which is critical for viral entry and cell attachment. However, non-neutralizing, ADE-inducing antibodies were also identified to bind to other motifs on WT DIII. Targeting WT NS1 glycoprotein, which plays a key role in viral replication and endothelial dysfunction, may reduce disease severity without ADE, but it may elicit non-neutralizing, even autoreactive antibodies. To address these challenges, structure-based approaches may be used for immunogen mutagenesis and multivalent display of these immunogens. This proposal will investigate methods to increase the immunodominance of heterologous epitopes found on the lateral ridge (LR) of DIII and the β-ladder of NS1, while masking certain epitopes on these immunogens, to induce broad humoral protection and mitigate severity of disease. We previously demonstrated the strong immunoprotecting potential of a homotypic ZV immunization strategy, which involved the multivalent display of a LR-focused DIII mutant onto nanoparticles. We will study the effectiveness of various heterotypic nanoparticles as cross-immunization strategies against ZV and DV and characterize underlying humoral responses to the immunizations in mice (Aim 1). We will then explore the design of miniβ, a novel NS1 mutant which may be employed to curb disease progression without eliciting ADE or autoreactive antibodies (Aim 2). These studies will further our understanding of the adaptive immune response to flavivirus infections and may provide a framework for developing novel vaccines and antibody-based therapeutics.