Design of universal Ebola virus vaccine candidates via immunofocusing

Ebola virus causes hemorrhagic fever in humans and poses a significant threat to global public health. Although two viral vector vaccines have been approved to prevent Ebola virus disease, they are distributed in the limited ring vaccination setting and only indicated for prevention of infection from orthoebolavirus zairense (EBOV) - one of three orthoebolavirus species that have caused previous outbreaks. Ebola virus glycoprotein GP mediates viral infection and serves as the primary target of neutralizing antibodies. Here we describe a universal Ebola virus vaccine approach using structure-guided design of candidates with hyperglycosylation that aims to direct antibody responses away from variable regions and toward conserved epitopes of GP. We first determined the hyperglycosylation landscape on Ebola virus GP and used that to generate hyperglycosylated GP variants with two to four additional glycosylation sites to mask the highly variable glycan cap region. We then created vaccine candidates by displaying wild-type or hyperglycosylated GP variants on ferritin nanoparticles (Fer). Immunization with these antigens elicited potent neutralizing antisera against EBOV in mice. Importantly, we observed consistent cross-neutralizing activity against Bundibugyo virus and Sudan virus from hyperglycosylated GP-Fer with two or three additional glycans. In comparison, elicitation of cross-neutralizing antisera was rare in mice immunized with wild-type GP-Fer. These results demonstrate a potential strategy to develop universal Ebola virus vaccines that confer cross-protective immunity against existing and emerging filovirus species. SIGNIFICANCE STATEMENTEbola virus outbreaks cause hemorrhagic fever with high mortality rates. Current viral vaccines require cold-chain storage and are distributed in limited ring vaccination settings. They are only indicated for protection against orthoebolavirus zairense (EBOV), one of three human-pathogenic Ebola virus species. Here we harness hyperglycosylation as an immunofocusing approach to design universal Ebola virus vaccine candidates based on Ebola virus glycoprotein (GP) displayed on ferritin nanoparticles (Fer). Compared with wild-type GP-Fer, immunization with hyperglycosylated GP-Fer elicited potently neutralizing antisera against EBOV, and more importantly, consistent cross-neutralizing activity against the other two orthoebolavirus species. Our work shows that immunofocusing antibody responses toward conserved and neutralizing epitopes of GP represents a promising strategy for vaccine design against antigenically diverse Ebola virus species.