Successful post-exposure prophylaxis of Ebola infected non-human primates using Ebola glycoprotein-specific equine IgG

Abstract

Herein we describe production of purified equine IgG obtained from horses immunized with plasmid DNA followed by boosting with Kunjin replicon virus-like particles both encoding a modified Ebola glycoprotein. Administration of the equine IgG over 5 days to cynomolgus macaques infected 24 hours previously with a lethal dose of Ebola virus suppressed viral loads by more than 5 logs and protected animals from mortality. Animals generated their own Ebola glycoprotein-specific IgG responses 9-15 days after infection, with circulating virus undetectable by day 15-17. Such equine IgG may find utility as a post-exposure prophylactic for Ebola infection and provides a low cost, scalable alternative to monoclonal antibodies, with extensive human safety data and WHO-standardized international manufacturing capability available in both high and low income countries.

Figure 1. Production and purification of anti-EBOV GP equine IgG.

(a) Schedule for horse immunization, serum sample collection and plasmapheresis. DNA - phCMV-GP/D637L. VLP - KUN-VLP-GP/D637L. (b) Anti-EBOV antibody titers of equine serum samples determined by ELISA. Inserts show virus neutralization titers; PRNT - plaque reduction neutralizing titers. (c) Workflow of IgG purification and concentration from equine plasma collected by plasmapheresis. TFF - tangential flow filtration. (d) Analysis of purified IgG product. Polyacrylamide gel electrophoreses of purified IgG product stained with Coomassie blue (left) and analysis of purified IgG product by ELISA and virus-neutralization assays (right).

Figure 2. Post-exposure treatment of EBOV-infected NHPs with purified anti-EBOV GP equine IgG.

Cynomolgus macaques (Macaca fascicularis) were infected with 1000 PFU of EBOV. The first equine IgG treatment (20 ml intravenously) was given 24 h post-infection (day 1) followed by daily treatments for 4 days (20 ml intravenously), with NHP2 and NHP3 receiving an additional treatment (5 ml intravenously) on day 13 post-infection. (a) EBOV RNA (RNAemia) levels in the Control and IgG-treated NHPs expressed in PFU equivalents per ml of serum (see Materials and Methods). Red cross in the Control shows time of death. (b) Body temperature of Control and IgG-treated NHPs. Red boxes represent ≥1.5 °C fever. (c) ALT and AST levels in Control and IgG-treated NHPs. Red boxes represent values above the normal range. (d) Analysis of anti-GP antibody responses by ELISA assays; total anti-GP antibody titers (top panels) and NHP anti-GP titers (bottom panels). Yellow boxes illustrate periods when equine GP-specific IgG, but not NHP GP-specific IgG, were detected.