Using a Human Challenge Model of Infection to Measure Vaccine Efficacy: A Randomised, Controlled Trial Comparing the Typhoid Vaccines M01ZH09 with Placebo and Ty21a

Abstract

Background: Typhoid persists as a major cause of global morbidity. While several licensed vaccines to prevent typhoid are available, they are of only moderate efficacy and unsuitable for use in children less than two years of age. Development of new efficacious vaccines is complicated by the human host-restriction of Salmonella enterica serovar Typhi (S. Typhi) and lack of clear correlates of protection. In this study, we aimed to evaluate the protective efficacy of a single dose of the oral vaccine candidate, M01ZH09, in susceptible volunteers by direct typhoid challenge.

Methods and findings: We performed a randomised, double-blind, placebo-controlled trial in healthy adult participants at a single centre in Oxford (UK). Participants were allocated to receive one dose of double-blinded M01ZH09 or placebo or 3-doses of open-label Ty21a. Twenty-eight days after vaccination, participants were challenged with 104CFU S. Typhi Quailes strain. The efficacy of M01ZH09 compared with placebo (primary outcome) was assessed as the percentage of participants reaching pre-defined endpoints constituting typhoid diagnosis (fever and/or bacteraemia) during the 14 days after challenge. Ninety-nine participants were randomised to receive M01ZH09 (n = 33), placebo (n = 33) or 3-doses of Ty21a (n = 33). After challenge, typhoid was diagnosed in 18/31 (58.1% [95% CI 39.1 to 75.5]) M01ZH09, 20/30 (66.7% [47.2 to 87.2]) placebo, and 13/30 (43.3% [25.5 to 62.6]) Ty21a vaccine recipients. Vaccine efficacy (VE) for one dose of M01ZH09 was 13% [95% CI -29 to 41] and 35% [-5 to 60] for 3-doses of Ty21a. Retrospective multivariable analyses demonstrated that pre-existing anti-Vi antibody significantly reduced susceptibility to infection after challenge; a 1 log increase in anti-Vi IgG resulting in a 71% decrease in the hazard ratio of typhoid diagnosis ([95% CI 30 to 88%], p = 0.006) during the 14 day challenge period. Limitations to the study included the requirement to limit the challenge period prior to treatment to 2 weeks, the intensity of the study procedures and the high challenge dose used resulting in a stringent model.

Conclusions: Despite successfully demonstrating the use of a human challenge study to directly evaluate vaccine efficacy, a single-dose M01ZH09 failed to demonstrate significant protection after challenge with virulent Salmonella Typhi in this model. Anti-Vi antibody detected prior to vaccination played a major role in outcome after challenge.

Trial registration: ClinicalTrials.gov (NCT01405521) and EudraCT (number 2011-000381-35).

Conflict of interest statement

I have read the journal's policy and the authors of this manuscript have the following competing interests: ZH is a named inventor of a patent (US7887816 B2) now owned by Prokarium Ltd, UK. ZH and SL were paid employees of Emergent BioSolutions at the time this trial was performed. All other authors have declared that no competing interests exist.

Figures

Fig 1. Study profile.

During the screening process, participant exclusions were made either during initial telephone screening or at/after the study centre screening visit. ‘Other’ reasons for exclusions were: contact with young children (3), contact with vulnerable individuals (9), food-related occupation (5), previously resident in typhoid-endemic area for >6 months (15), unable to contact (6) & unknown (8).

Fig 2. Cumulative incidence of typhoid infection after S. Typhi challenge at Time = 0.

Time to infection, measured from challenge agent ingestion to development of first fever ≥38°C or first positive blood culture sampling. Non-diagnosed participants censored at 348 hours (dashed line). P value from log-rank test comparing all three groups.

Fig 3. Cumulative incidence of bacteraemia or fever after S. Typhi challenge at Time = 0.

(A) Time to bacteraemia, measured from challenge agent ingestion to time of first positive blood culture sampling. Non-bacteraemic participants censored at time of diagnosis or at 348 hours (dashed line). P value from log-rank test comparing all three groups and comparing M01ZH09 and Ty21a to placebo, respectively. (B) Time to fever, measured from challenge agent ingestion to first recording of fever (oral temperature ≥38°C). Afebrile participants censored at time of diagnosis or at 336 hours (dashed line). P value from log-rank test comparing all three groups.

Fig 4. Changes in physiological signs in those participants developing typhoid by time after diagnosis according to vaccine group allocation.

(A) Temperature, (B) Heart rate and (C) Systolic blood pressure. Mean change from baseline and 95% confidence interval. Dashed black vertical line marks point of typhoid diagnosis; grey horizontal bar indicates all participant mean (95% CI) values pre-vaccination.

Fig 5. Proportion of participants reporting each solicited symptom during 14 days after challenge according to vaccine group allocation.

(A) Placebo, (B) M01ZH09, and (C) Ty21a vaccine recipient groups. Maximum severity score per participant for each symptom was used and graded according to criteria detailed in the study protocol: fever thresholds are Grade 1: 38.0–38.4°C; Grade 2: 38.5–38.9°C; Grade 3: 39.0–40.0°C; Grade 4: >40.0°C.

Fig 6. Group mean changes (95% confidence intervals) in haematological blood parameters compared to pre-challenge measurements according to vaccine allocation and challenge outcome.

(A) Haemoglobin, (B) platelets, (C) total white cell count, (D) neutrophils, (E) lymphocytes, and (F) eosinophils. TD, typhoid diagnosis; nTD, non-typhoid diagnosis.

Fig 7. Group mean changes (95% confidence intervals) in biochemistry blood parameters compared to individual baseline measurements according to vaccine allocation and challenge outcome.

(A) Sodium, (B) potassium, (C) urea, (D) creatinine, (E) Albumin, (F) amylase, (G) alkaline phosphatase, (H) alanine aminotransferase, (I) bilirubin, and (J) C-reactive protein. TD, typhoid diagnosis; nTD, non-typhoid diagnosis.

Fig 8. Blood quantification of Salmonella Typhi bacteria present at point of typhoid diagnosis by vaccine group.

10mL peripheral blood was collected at typhoid diagnosis (prior to antibiotic treatment) into an Isolator 10 tube (Wampole Laboratories). Lysis centrifugation was performed (30min x 3000G without brake) after which the deposit was plated to XLD and incubated aerobically at 37°C for 24 hours. After incubation, colony counts and slide agglutination tests were performed. P values calculated using Mann Whitney U test. Median [IQR] bacterial loads in CFU/mL were: M01ZH09 (n = 14): 0.13 [0.05–0.80]; Placebo (n = 15): 1.30 [0.30–5.40]; Ty21a (n = 12): 0.05 [0.05–0.99]. Lower limit of detection, 0.1 CFU/mL; zero values were substituted with LOD/2, i.e. 0.05 CFU/mL.

Fig 9. Proportion of participants reporting each solicited symptom during the 7-days after receipt of (first) vaccine dose.

(A) Placebo, (B) M01ZH09, and (C) Ty21a vaccine recipient groups. Maximum severity score per participant for each symptom was used; grade 1: symptom reported but no interference with daily activity; grade 2: some interference with normal daily activities; grade 3: significant symptoms preventing normal daily activity; grade 4: potentially life-threatening (see S1 Protocol).

Fig 10. Immunoglobulin G ASC and antibody responses to S. Typhi LPS and flagellin before (Day -28) and after vaccination (either Day -21 or Day 0 for ASC and ELISA assays, respectively).

(A) LPS and (B) flagellin specific IgG antibody secreting cell responses, respectively, measured pre-vaccination (Day -28) and 7 days later (Day -21). (C) Anti-LPS and (D) anti-flagellin antibody titres, respectively, measured pre-vaccination (Day -28) and 28 days later (Day 0, i.e. prior to S. Typhi challenge). Data are grouped according to vaccine allocation.

Fig 11. Pre-vaccination (Day -28) and pre-challenge (Day 0) IgG antibody titres according to outcome after challenge.

(A) Anti-LPS IgG, (B) anti-H IgG, and (C) anti-Vi IgG. Clear circles, non-typhoid diagnosed participants; black circles, typhoid diagnosed participants. Difference between Day -28 and Day 0 anti-LPS and anti-H antibodies analysed by ANCOVA adjusted for vaccine group and baseline titre; P values for anti-Vi antibody calculated by Wilcoxon Rank Sum. Lower limit-of Vi-antibody detection, 7.4EU/mL.