Investigation of Synthetic DNA-based Viral Particles for Spatially Controlled Antigen Presentation - PROJECT SUMMARY HIV and influenza pose major health burdens in the US and worldwide, claiming hundreds of thousands of lives annually, with the latter additionally posing a significant pandemic threat. Protein subunit vaccines formulated as nanoparticles with multivalent display of pathogenic immunogens are a proven, successful strategy to safely and effectively induce long lasting humoral, antibody-based immunity. Indeed, this strategy may be particularly important for challenging-to-neutralize pathogens such as HIV, which still does not have an approved preventative vaccine, and for designing future-proofed vaccines against the rapidly evolving pathogen influenza that poses an ongoing pandemic threat. However, to date it remains unclear what the optimal characteristics of nanoparticulate vaccines are. In addition, because most particulate subunit vaccines are composed of protein virus-like particles (P-VLPs) that multivalently display pathogenic antigens that are the target of broadly neutralizing antibody (bnAb) responses, it remains unclear whether the protein scaffolds used to formulate these VLPs are in themselves eliciting humoral immune responses that distract the immune system away from the pathogenic antigens of interest. In this project, we previously discovered that DNA-based VLPs (D-VLPs) can be used to spatially organize antigens multivalently in a manner similar to P-VLPs to elicit humoral antibody responses across multiple pathogens, yet without the protein-based scaffold VLP and resulting off-target antibody response. Here, we investigate how D-VLPs can be used to optimize priming, shepherding, and polishing phases of HIV vaccination using sequential introduction of engineered immunogens to drive the generation of target bnAbs. Specifically, we test the relative roles of antigen copy number, spacing, VLP size and geometry, glycosylation, and passivation, on lymph node targeting, germinal center formation, and on-target B cell activation. We investigate how these principles can be used together with controlled presentation of T cell helper epitopes and cytokine co-delivery to promote robust, long-lived GCs that result in bnAbs for HIV. Finally, we explore how the D-VLP platform generalizes to the distinct, challenging-to-neutralize pathogen influenza, towards identifying future-proofed vaccines to avert pandemic threats. Taken together, these results will help elucidate optimal VLP design rules for HIV and influenza, and may eventually also offer a clinically relevant new vaccine platform in future work.
