Study Participants

Table 1. Table 1. Characteristics of the Participants at Enrollment.

A total of 78 volunteers (55 men [71%] and 23 women [29%]), with a mean age of 36 years (range, 20 to 64), were enrolled in a consecutive manner; injections were administered between October 10, 2014, and January 6, 2015. A total of 60 volunteers were randomly assigned to receive rVSV-ZEBOV, and 18 volunteers were randomly assigned to receive saline placebo. At the NIH site, all the participants received a second identical dose of vaccine 28 days after the initial dose. All the volunteers completed the follow-up visits that were scheduled during the 28-day windows after vaccination; however, 4 volunteers were lost to follow-up by the conclusion of the trial (Fig. S1 in the Supplementary Appendix). Additional details regarding the demographic characteristics of the volunteers are provided in Table 1.

Safety

Figure 1. Figure 1. Frequency of Solicited Adverse Events According to Cohort and Grade. Cohort 1 received a dose of 3 million plaque-forming units (PFU) of the vaccine, Cohort 2 a dose of 20 million PFU, and Cohort 3 a dose of 100 million PFU. All adverse events were assessed for relatedness to the vaccine; events that were judged by the investigating physicians not to be related to the vaccine are not shown. Adverse events were graded for severity on the basis of Food and Drug Administration toxicity grading.15 Unsolicited adverse events and laboratory adverse events are shown in the Supplementary Appendix.

There were no deaths, serious adverse events, or adverse events resulting in withdrawal from the study. There was no association between vaccine dose and the frequency or severity of adverse events (Figure 1, and Figs. S2 and S3 and Table S10 in the Supplementary Appendix). After a single inoculation of vaccine, mild-to-moderate injection-site pain was observed in the majority of participants. Systemic reactogenicity was transient and, in the majority of volunteers, mild to moderate in severity. Objective fever was noted in 20 of the 60 vaccinees: 11 (18%) had grade 1 fever (temperature range, 38.0 to 38.4°C), 7 (12%) had grade 2 fever (temperature range, 38.5 to 38.9°C), and 2 (3%) had grade 3 fever (temperature range, 39.0 to 40.0°C). Fever onset and frequency did not appear to be dose-dependent (Figure 1, and Fig. S2 in the Supplementary Appendix); fever typically developed 12 to 24 hours after vaccination and resolved by the end of postvaccination day 1. One volunteer who received a dose of 3 million PFU had grade 1 fever 7 days after vaccination that resolved within 24 hours without development of other symptoms.

Other commonly reported systemic symptoms among vaccinees were headache, myalgia, and fatigue, with typical onset 12 to 24 hours after vaccination. Notable adverse events were unilateral conjunctivitis that developed in one volunteer 1 day after inoculation and oral ulcers that developed in five vaccinated volunteers 4 to 16 days after vaccination. PCR analysis of swabs of the affected areas (a conjunctival swab and swabs of three of the five oral ulcers) was negative for the Ebola glycoprotein gene insert. Three vaccinated participants had cervical lymphadenopathy; one of the three also reported an oral ulcer. Infectious colitis developed in one participant 21 days after vaccination; symptoms included severe abdominal pain on the left side and four episodes of mild diarrhea with blood. Computed tomography of the abdomen at an outside hospital showed mild thickening of the descending colon. All conditions resolved without complications. Among participants who received a second dose of the vaccine, reactogenicity at the injection site and systemic reactogenicity were less severe after the second dose than after the first dose. A complete list of solicited and unsolicited adverse events is provided in Table S10 in the Supplementary Appendix.

Safety laboratory values were generally unremarkable, with the majority of adverse events occurring after the first dose of vaccine. Transient mild-to-moderate lymphopenia occurred in 24 of 60 participants, typically on day 1, with abatement by day 3 after vaccination. Mild-to-moderate neutropenia, which occurred in 14 of 60 participants, was most notable on day 3 after vaccination and typically abated within 2 to 4 days. An asymptomatic grade 2 thrombocytopenia, associated with grade 1 lymphopenia, was noted on day 1 after vaccination in one volunteer who received a dose of 20 million PFU; the condition resolved by day 7.

After a report from a phase 1 study in Geneva of the onset of arthritis in 22% of the participants starting the second week after injection,17,18 volunteers were specifically queried about the development of new arthralgia, arthritis, or rash during the second week or later after vaccination. A total of 19 participants reported arthralgia, typically soon after vaccination. Five participants had an onset of arthralgia 7 to 14 days after vaccination, and 3 participants had arthralgia that began after the second vaccination. No clinical cases of arthritis were diagnosed.

rVSV-ZEBOV on PCR Assay

Table 2. Table 2. Vaccine Virus Detection by Means of Qualitative Reverse-Transcriptase–Polymerase-Chain-Reaction Assay.

PCR results are shown in Table 2. All the vaccinated volunteers had detectable vaccine viremia at the first visit after vaccination (day 1 at the WRAIR and day 3 at the NIH). Twelve of the 60 vaccinated volunteers (20%) had viremia on day 7 after vaccination. Viremia was undetectable by day 14 in all vaccinees tested at that time point (30 volunteers at the WRAIR). In the group that received a dose of 3 million PFU, there was one positive urine sample on day 3 and one on day 7. Across the vaccine groups, a small number of saliva samples were PCR-positive on days 1, 3, 7, and 14. Two subsequent saliva samples were PCR-negative in the single volunteer who had a positive PCR saliva sample on day 14. Cycle-threshold values for the positive urine sample on day 7 and saliva sample on day 14 were near the lower limit of detection for the assay.

After administration of a second vaccine dose at the NIH site, a single volunteer in the group that received a dose of 100 million PFU had viremia 3 days later. PCR results were otherwise negative in blood, urine, and saliva.

ELISA for Ebola Glycoprotein

Figure 2. Figure 2. Antibody Responses to Ebola Glycoprotein. Individual antibody titers as assessed at 14 and 28 days after vaccination are shown according to vaccine dose group, as measured by an enzyme-linked immunosorbent assay (ELISA) against the Zaire–Kikwit strain glycoprotein (Panel A) and a pseudovirion neutralization assay (Panel B). Geometric mean titers (horizontal lines) are shown for each group and time point. Geometric mean titers from 28 days after initial vaccination through 180 days after initial vaccination are shown for the glycoprotein ELISA (Panel C) and the pseudovirion neutralization assay (Panel D). Solid lines indicate groups that received a second dose at day 28, and dashed lines indicate groups that did not receive a second dose at day 28. In all panels, I bars indicate 95% confidence intervals.

Table 3. Table 3. Geometric Mean Antibody Titers.

ELISA results are shown in Figure 2 and Table 3, and Tables S1 through S4 in the Supplementary Appendix. After a single dose of vaccine, IgG responses were observed. A total of 16 of 20 volunteers (80%) who received a dose of 3 million PFU, 19 of 20 volunteers (95%) who received a dose of 20 million PFU, and 18 of 20 who received a dose of 100 million PFU had undergone seroconversion by day 14. All 60 vaccinated volunteers (100%) had undergone seroconversion by day 28. The groups that received a dose of 20 million PFU or 100 million PFU had higher geometric titers against the Zaire–Kikwit strain than the group that received a dose of 3 million PFU, both on day 14 (857 and 888 vs. 283; P=0.008 and P=0.02, respectively) and on day 28 (4079 and 4079 vs. 1300; P=0.001 and P<0.001, respectively). All vaccinated cohorts showed increases in titers from day 14 to day 28; titers increased from 283 on day 14 to 1300 on day 28 in the group receiving a dose of 3 million PFU (P<0.001), from 857 to 4079 in the group receiving a dose of 20 million PFU (P<0.001), and from 888 to 4079 in the group receiving a dose of 100 million PFU (P=0.01)). There was no significant difference in the geometric mean titer between the group that received a dose of 20 million PFU and the group that received a dose of 100 million PFU.

At day 28, there were no significant differences in the geometric mean titer between the groups that were to receive a second vaccine dose and those that were not. All three groups that received a second dose had increases in titers from day 28 to day 56; titers increased from 1300 on day 28 to 4222 on day 56 in the group that received a dose of 3 million PFU (P<0.001), from 5198 to 7352 in the group that received a dose of 20 million PFU (P=0.27), and from 3676 to 11,143 in the group that received a dose of 100 million PFU (P<0.001). Among participants who received a second dose, the geometric mean titer was higher at day 84 than at day 28 in the group that received a dose of 3 million PFU (1300 at day 28 vs. 2986 at day 84 [P=0.02]) and in the group that received a dose of 100 PFU (3676 at day 28 vs. 7352 at day 84 [P=0.02]). Among participants who did not receive a second dose, only the group that received a dose of 3 million PFU had significant increases in the geometric mean titer from day 28 through day 84 (1300 at day 28 vs. 2599 at day 56 [P<0.001] and 1300 at day 28 vs. 2263 at day 84 [P=0.003]). In all three vaccinated groups, participants who received a second vaccination had higher geometric mean titers on day 56 than those who did not (4222 vs. 2599 in the group that received a dose of 3 million PFU [P=0.16], 7352 vs. 3733 in the group that received a dose of 20 million PFU [P=0.04], and 11,143 vs. 4525 in the group that received a dose of 100 million PFU [P=0.04]). At the 180-day follow-up, there was no significant difference in geometric mean titers between the groups that received a second dose of vaccine and the groups that received a single dose.

PsVNA Titers

Results with respect to neutralizing antibody titers against the Zaire–Kikwit strain glycoprotein are shown in Figure 2 and Table 3, and Table S5 through S8 in the Supplementary Appendix. After a single vaccination, all groups had neutralizing antibodies by day 28, in a dose-dependent manner. The geometric mean titer in the group that received a dose of 100 million PFU was significantly higher than in the group that received a dose of 3 million PFU both on day 14 (127 vs. 39 [P=0.004]) and on day 28 (461 vs. 223 [P=0.01]). All three dose groups had significant increases in the geometric mean titer from day 14 to day 28; the titer increased from 39 on day 14 to 223 on day 28 in the group that received a dose of 3 million PFU (P<0.001), from 47 to 441 in the group that received a dose of 20 million PFU (P<0.001), and from 127 to 461 in the group that received a dose of 100 million PFU (P<0.001).

At day 28, there were no significant differences in the geometric mean titer between the groups that were to receive a second vaccine dose and those that were not. Groups that received a second dose of vaccine had an initial trend of increased geometric mean titers during the month after revaccination (222 at day 28 vs. 344 at day 56 in the group that received a dose of 3 million PFU [P=0.08], 415 vs. 653 in the group that received a dose of 20 million PFU [P=0.33], and 476 vs. 669 in the group that received a dose of 100 million PFU [P=0.19]). However, this trend was reversed in titers measured 2 months after revaccination (222 at day 28 vs. 33 at day 84 in the group that received a dose of 3 million PFU [P<0.001], 415 vs. 47 in the group that received a dose of 20 million PFU [P=0.003], and 476 vs. 90 in the group that received a dose of 100 million PFU [P<0.001]). Vaccine groups that did not receive a second dose had a decrease in neutralizing antibody responses from day 28 to 56 (223 vs. 138 in the group that received a dose of 3 million PFU [P=0.06], 468 vs. 170 in the group that received a dose of 20 million PFU [P=0.008], and 447 vs. 219 in the group that received a dose of 100 million PFU [P=0.02]). At the 180-day follow-up, there was no significant difference in neutralizing antibody responses between the groups that received a second dose of vaccine and the groups that received a single dose.