Safety and Immunogenicity of a Recombinant Tetanus Vaccine in Healthy Adults in China: A Randomized, Double‐Blind, Dose Escalation, Placebo‐ and Positive‐Controlled, Phase 1/2 Trial

Abstract Tetanus is a fatal but vaccine‐preventable disease. The currently available tetanus vaccines are tetanus toxoid (TT)‐based. Although these vaccines are generally effective, challenges in vaccine development and access remain. A randomized, double‐blind, dose escalation, placebo‐ and positive‐controlled, phase 1/2 trial (ChiCTR1800015865) is performed to evaluate the safety and immunogenicity of an alternative recombinant tetanus vaccine based on the Hc domain of tetanus neurotoxin (TeNT‐Hc) in healthy adult volunteers. The primary outcome is the safety profile of the recombinant tetanus vaccine, and immunogenicity is the secondary outcome. 150 eligible participants were enrolled and randomly assigned to receive one of the three doses of recombinant tetanus vaccine (TeNT‐Hc 10/20/30 µg), TT vaccine, or placebo. The recombinant tetanus vaccine shows a good safety profile. The frequency of any solicited and unsolicited adverse events after each vaccination does not differ across the vaccine and placebo recipients. No serious treatment‐related adverse events occur. The recombinant tetanus vaccine shows strong immune responses (seroconversion rates, geometric mean titer, and antigen‐specific CD4+/CD8+ T‐cell responses), which are roughly comparable to those of the TT vaccine. In conclusion, the findings from this study support that recombinant tetanus vaccine is safe and immunogenic; thereby, it represents a novel vaccine candidate against tetanus.


Supporting Information
Inclusion/exclusion criteria Inclusion criteria 1. Men or women aged >18 and <60 years 2. Ability to comprehend the full nature and purpose of the study, including possible risks and side effects; ability to co-operate with the investigator and to comply with the requirements of the entire study 3. Volunteers can comply with the requirements of the clinical trial program and complete all follow-up studies; 4. Ear temperature ≤ 37.5 degree C; 5. BMI index 19-32; 6. Signed informed consent obtained before any trial-related activities 7. Normal findings in the medical history, by physical examination and normal laboratory tests Exclusion criteria for first vaccination 1. Women who are pregnant or breastfeeding, or who have a childbirth plan during the first three months of the study; 2. Infected with tetanus; 3. Accepted inoculation of tetanus vaccine in past 5 years, tetanus-diphtheria combined vaccine or polysaccharide-binding vaccine with TT as carrier protein; 4. allergies, such as those who are allergic to two or more drugs or food allergies, or are known to be allergic to the vaccine component (aluminum hydroxide adjuvant, sodium chloride); 5. Serious adverse reactions to any vaccine, such as allergies, urticaria, dyspnea, angioneurotic edema or abdominal pain; 6. Autoimmune diseases, immunodeficiency or HIV positive, important organs are primary diseases; 7. Systemic chemotherapy in the past 5 years, systemic immunosuppressive therapy, radiation therapy, cytotoxic therapy, high-dose inhaled corticosteroids in the past 6 months (excluding corticosteroid spray therapy for allergic rhinitis, acute non-Treatment of dermatitis with topical corticosteroids); prescription and/or non-prescription patients used within 2 months; physician-diagnosed coagulation disorders (such as clotting factor deficiency, coagulopathy, platelet abnormalities), or significant bruising or clotting disorders; 8. Received other study drugs within the first 3 months of receiving the vaccine for this study; 9. Received blood products within the first 3 months of receiving the vaccine for this study; 10. Live attenuated vaccines received within 30 days of receiving the vaccine for this study; 11. Received a subunit or inactivated vaccine, such as a pneumococcal vaccine, or antiallergic therapy within the first 14 days of receiving the vaccine for this study; 12. Ongoing anti-TB prevention or treatment; 13. According to the researcher's judgment, due to various medical, psychological, social, occupational, or other conditions, it is contrary to the trial program or affects the subject's signing of informed consent.

Second vaccination exclusion criteria
If one of the following (1) to (3) adverse events (AEs) occurs, the continuation of vaccination is prohibited, but other study steps may be continued at the discretion of the investigator; if one of the following (4) to (5) adverse events occurs The investigator determines whether to continue the vaccination; if one of the following (6) to (7) adverse events occurs, the vaccination may be postponed within the time window set by the plan (without the occurrence of (6) to (7) adverse events. Two-day vaccination is on the 28th day, postponed vaccination within the time window means vaccination within 3 days after 28 days). For the following AEs, subjects must be followed up to the event resolution: (1) any serious adverse event that has a causal relationship with the vaccination of the test vaccine; (2) Serious allergic reactions or hypersensitivity after vaccination (including urticaria/rashes occurring within 30 minutes of vaccination); (3) The occurrence of any first needle exclusion criteria after the election; (4) Acute or new chronic diseases occurred during this vaccination; (5) Grade 3 or above adverse reactions occurred within 72 hours of the previous vaccination; (6) Acute disease at the time of vaccination (acute disease refers to moderate or severe disease with or without fever); (7) Ear temperature >37.5°C at the time of vaccination.

Safety assessment
Vital sign checks including heart rate, blood pressure, respiratory rate, and ear temperature measurements were assessed at baseline. Blood samples for safety analysis were taken and assessed at baseline and at three predefined time points: 7 days, 28 days after first vaccination, and 7 days after second vaccination ( Table 1). Participants had a telephone visit for safety analysis within 0-7 days after each vaccination. Additionally, participants were required to keep a study diary to record any adverse events. Safety monitoring was carried out by Clinical Research Coordinators at the study site. Safety assessment at screening visit included: • Medical history • Concomitant medication • Body weight and height • Vital signs • Hematology: red blood cell count (RBC), white blood cell count (WBC), leukocyte differential count (neutrophils, basophils, eosinophils, monocytes and lymphocytes), platelet count, hemoglobin, hematocrit • Clinical chemistry (renal and hepatic function): Alanine transaminase (ALT), Aspartate transaminase (AST), albumin, Globulin, Bilirubin, total bile acid, Blood glucose, Urea, creatinine, Total Cholesterol, Triglyceride, High density liptein cholesterol, low density liptein cholesterol, blood urea nitrogen (BUN), potassium kalium, sodium, chlorine, phosphorus, calcium. Urine: PH, proportion, color, turbidity, urine creatinine, RBC, WBC, cast, bacteria, epithelial cells, crystal.

The detail of six serious adverse events
Three cases were reported in two participants from placebo group, including two cases of knee injury and one case of acute carbon monoxide poisoning. The other three cases occurred in the medium-dose vaccine group, including one case who had a spontaneous abortion 3 months after second vaccination, one case who reported 5 months after second vaccination that she was pregnant and had an abortion five days after diagnosis of pregnancy, and one case who had anal fissure. Unfortunately, one participant had suffered from acute carbon monoxide poisoning and died. Data are n (%). *p-values were generated from comparisons across the five groups. *p-values were generated from comparisons across the five groups.

Supplementary Table 6. Geometric mean titre of anti-TT IgG antibody, after initial and boost vaccinations.
Data are geometric mean titre (95% CI). Analysis of variance was used for the log-transformed antibody titres; *p values were generated from then comparison across five groups (placebo, TT vaccine, low-dose, medium-dose, high-dose).
†Both the low-dose and medium-dose groups showed lower GMT than did TT group. ‡The low-dose groups showed lower GMT than did TT group. §The high-dose, medium-dose and low-dose groups showed higher GMT than did the placebo group. Data are geometric mean titre (95% CI). Analysis of variance was used for the log-transformed antibody titres; *p values were generated from then comparison across five groups (placebo, TT vaccine, low-dose, medium-dose, high-dose). †The high-dose groups showed higher GMT than did TT group. ‡ The high-dose, medium-dose and low-dose groups showed higher GMT than did TT group. §The high-dose, medium-dose and low-dose groups showed higher GMT than did the placebo group. Data are geometric concentration ± SD. Analysis of variance was used for the log-transformed antibody titres. *p-values were generated from comparisons across the five groups (placebo, positive vaccine, low-dose, medium-dose, high-dose). †The high-dose, medium-dose, and low-dose groups showed lower GMC than did the positive group. ‡The medium-dose and low-dose groups showed lower GMC than did the positive group. §The high-dose, medium-dose, and low-dose groups showed higher GMC than did the placebo group.