Single-component multilayered self-assembling nanoparticles presenting rationally designed glycoprotein trimers as Ebola virus vaccines

Abstract

The Ebola virus (EBOV) glycoprotein (GP) can be recognized by neutralizing antibodies (NAbs) and is the main target for vaccine design. We first investigated the contribution of the stalk and the heptad repeat 1-C (HR1C) region to GP metastability. Specific stalk and HR1C modifications in a mucin-deleted form (GPΔmuc) increased trimer yield, while alteration of HR1C exerted a more complex effect on thermostability. Crystal structures were determined to validate these rationally designed GPΔmuc trimers in their unliganded state. We then displayed a modified GPΔmuc trimer on engineered nanoparticles with encapsulated locking domains (LD) and a cluster of helper T-cell epitopes. In mice and rabbits, GP trimers and nanoparticles elicited cross-ebolavirus NAbs as well as non-NAbs that enhance in-vitro infection. Next-generation sequencing (NGS) revealed B-cell profiles that are specific to vaccine platforms. This study provides insight into GP metastability and paves the way for development of an effective, pan-ebolavirus nanoparticle vaccine.

Single-component, self-assembling, protein nanoparticles presenting the receptor binding domain and stabilized spike as SARS-CoV-2 vaccine candidates

“…1J). Previously, we genetically fused a small LD protein and a T-cell epitope to each subunit of the E2p and I3-01v9 60-mers, resulting in "multilayered" SApNPs (60). PADRE, a 13-aa pan-DR epitope that activates CD4 + T cells (74), was used to promote B cell development toward NAbs.…”

“…Recent cryo-EM and cryo-ET studies revealed that the SARS-CoV-2 spikes could adopt diverse orientations on native virions due to the highly flexible HR2 stalk (54)(55)(56). Previously, we identified an HR1 bend as the cause of HIV-1 Env metastability (58,81) and examined the role of an equivalent HR1 bend and the HR2 stalk in EBOV GP metastability (60). This understanding of metastability proved critical for designing stable trimers and trimer-presenting SApNP vaccines for both viruses (58)(59)(60)81).…”

“…Using the gene fusion approach, we previously designed single-component SApNPs displaying stabilized HIV-1 Env trimers (58,59) and optimized HCV E2 cores (57). Recently, we reengineered E2p and I3-01v9 60-mers to incorporate an inner layer of LDs to stabilize the non-covalently formed NP shell, in addition to a cluster of T-cell epitopes (60). This multilayered design may be essential to the stability of resulting SApNP vaccines when they are used to display large, complex viral antigens such as the SARS-CoV-2 spike.…”

“…3D). The varying shape of these profusions may correspond to S2GΔHR2 spikes with "open" RBDs, in contrast to an array of "closed" HIV-1 and EBOV trimers on the SApNP surface (58)(59)(60). Such open conformation may facilitate induction of a strong RBD-specific NAb response in vivo.…”

“…In this study, we designed and optimized SARS-CoV-2 antigens for multivalent display on self-assembling protein nanoparticles (SApNPs) (57)(58)(59), including a ferritin (FR) 24-mer and two 60-mers (E2p and I3-01) containing an inner layer of locking domains (LD) and a cluster of T-cell epitopes (60). To facilitate tag-free vaccine purification, we developed an immunoaffinity column based on antibody CR3022 that binds to both SARS-CoV-1/2 RBDs (34,50).…”