Identification of Novel Adjuvants for Ebola Virus-Like Particle Vaccine

Abstract

Ebola virus disease is a severe disease, often fatal, with a mortality rate of up to 90%. Presently, effective treatment and safe prevention options for Ebola virus disease are not available. Therefore, there is an urgent need to develop control measures to prevent or limit future Ebola virus outbreaks. Ebola virus protein-based virus-like particle (VLP) and inactivated whole virion vaccines have demonstrated efficacy in animal models, and the addition of appropriate adjuvants may provide additional benefits to these vaccines, including enhanced immune responses. In this study, we screened 24 compounds from injectable excipients approved for human use in Japan and identified six compounds that significantly enhanced the humoral response to Ebola VLP vaccine in a murine model. Our novel adjuvant candidates for Ebola VLP vaccine have already been demonstrated to be safe when administered intramuscularly or subcutaneously, and therefore, they are closer to clinical trials than adjuvants whose safety profiles are unknown.

Keywords: Ebola vaccine; adjuvants; virus-like particle.

Figure 1

Generation and characterization of Ebola Makona virus-like particles (VLPs). (A) Budding of Ebola VLPs from insect cells. High Five Cells were infected with recombinant Baculovirus expressing GP protein (rBV-GP) and recombinant Baculovirus expressing VP40 protein (rBV-VP40). At 24 h post-infection, the cells were fixed with 2.5% glutaraldehyde. Ultrathin (50-nm-thick) sections were stained with 2% uranyl acetate and Reynold’s lead and were observed under a transmission electron microscope. (B) Western blot analysis of the purified Ebola VLPs subjected to SDS-PAGE followed by Western blot analysis with rabbit polyclonal antibodies against the Ebola VP40 and GP proteins. Purified VLP ×10, ×100, and ×1000 indicate the purified VLP diluted to 1:10, 1:100, and 1:1000. (C) Electronic microscopic analysis of the purified VLPs. The purified VLPs were immunogold stained for Ebola GP with a rabbit anti-GP polyclonal antibody as the primary antibody and a goat anti-rabbit IgG 10 nm gold as the secondary antibody, fixed, cut, and analyzed by means of electron microscopy. Arrows indicate 10 nm gold particles conjugated with goat anti-rabbit IgG. Scale bar indicates 50 nm.

Figure 2

Screening adjuvant candidates for Ebola Makona VLP vaccine Three groups of six-week-old BALB/c mice (n = 4) were intramuscularly immunized with PBS, Ebola VLPs alone, or Ebola VLPs plus compound three times. Ebola VLPs plus AddaVax was used as a positive control. Blood was collected two weeks after the third immunization. Ebola GP-specific antibodies were assessed by using an enzyme-linked immunosorbent assay (ELISA) using purified His-tagged GP mutant as the coating antigen. Asterisks indicate that the antibody titer was significantly higher in mice immunized with the Ebola VLP vaccine plus the respective compound compared with the vaccine alone group (* p < 0.05; ** p < 0.01).