Protection of nonhuman primates against two species of Ebola virus infection with a single complex adenovirus vector

Abstract

Ebola viruses are highly pathogenic viruses that cause outbreaks of hemorrhagic fever in humans and other primates. To meet the need for a vaccine against the several types of Ebola viruses that cause human diseases, we developed a multivalent vaccine candidate (EBO7) that expresses the glycoproteins of Zaire ebolavirus (ZEBOV) and Sudan ebolavirus (SEBOV) in a single complex adenovirus-based vector (CAdVax). We evaluated our vaccine in nonhuman primates against the parenteral and aerosol routes of lethal challenge. EBO7 vaccine provided protection against both Ebola viruses by either route of infection. Significantly, protection against SEBOV given as an aerosol challenge, which has not previously been shown, could be achieved with a boosting vaccination. These results demonstrate the feasibility of creating a robust, multivalent Ebola virus vaccine that would be effective in the event of a natural virus outbreak or biological threat.

FIG. 1.

Vaccination and challenge schedule. Cynomolgus macaques were divided into two groups of five per group, and each group was vaccinated on days 0 and 65 (group 1) or days 0 and 120 (group 2) with CAdVax-EBO7 or a control CAdVax vector. Group 1 was challenged with 1,000 PFU of ZEBOV on day 106, and group 2 was challenged with 1,000 PFU of SEBOV on day 162 (both groups were challenged 6 weeks post-boosting vaccination). After a 10-week recovery period, group 2 was subsequently back-challenged with 1,000 PFU of ZEBOV on day 233. Filled arrows, vaccination; open arrows, virus challenge.

FIG. 2.

Humoral immune responses to ZEBOV and SEBOV before and after challenges. Geometric mean titers (plus standard deviations) of total immunoglobulin (Ig) in response to ZEBOV (A) and SEBOV (B) were measured by ELISA using inactivated filovirus preparations as immune targets. Group 1 animals were challenged with ZEBOV only. Group 2 animals were challenged with SEBOV and then back-challenged with ZEBOV. The control NHPs (n = 3) were evaluated on day zero before challenge.

FIG. 3.

Ad5 antibody responses after repeated exposure. Three monkeys were injected i.m. three times (arrows) with 10⁶ PFU of wild-type Ad5. Animal serum samples were used to measure Ad5-reactive antibodies by ELISA. After each injection, anti-Ad5 antibody titers increased rapidly and then began to drop, but they remained well above the preinjection level, indicating a sustained response.

FIG. 4.

Increasing the CAdVax vaccine dose is sufficient to overcome preexisting Ad5 immunity. Rhesus macaques were vaccinated twice at weeks 0 and 8 with an unrelated CAdVax vector to induce Ad5 immunity (gray arrows). At week 52 (black arrow), the animals were separated into three groups (n = 3) and vaccinated with 1 × 10⁸ PFU (low dose, open diamonds), 1 × 10⁹ PFU (medium dose, open squares), or 1 × 10¹⁰ PFU (high dose, open triangles) of EBO7. As a control, a group of Ad5-naïve animals was vaccinated with a low dose of EBO7 (shaded circles). Antibody titers were determined for Ad5 (black squares) and ZEBOV (open or shaded symbols) GP. Error bars show standard deviations.