Safety and immunogenicity of an oral tablet norovirus vaccine, a phase I randomized, placebo-controlled trial

Abstract

Background: Noroviruses are the leading cause of epidemic acute gastroenteritis and foodborne diarrheal disease in humans. However, there are no approved vaccines for noroviruses. Potential correlates of protection identified through human challenge studies include mucosal IgA, memory B cells, and serum-blocking antibody titers (BT50).

Methods: We conducted a single-site, randomized, double-blind, placebo-controlled clinical trial of an oral norovirus vaccine to determine safety and immunogenicity. This tablet vaccine is comprised of a nonreplicating adenovirus-based vector expressing the VP1 gene from the GI.1 norovirus strain and a double-stranded RNA adjuvant. Sixty-six adult subjects meeting inclusion/exclusion criteria were randomized 2:1 to receive a single vaccine dose or placebo, respectively. Immunogenicity was primarily assessed by serum BT50. Additional outcomes included serum ELISA titers, fecal and saliva antibody titers, memory and antibody-secreting cell (ASC) frequency, and B cell phenotyping.

Results: The vaccine was well-tolerated, with no dose-limiting toxicities. Adverse events were mild or moderate. The primary immunological endpoint (increase in BT50 titers) was met in the high-dose group (P = 0.0003), with 78% showing a ≥2-fold rise in titers after a single immunization. Vaccine recipients also developed mucosally primed VP1-specific circulating ASCs, IgA+ memory B cells expressing gut-homing receptor (α4β7), and fecal IgA, indicating substantial and local responses potentially relevant to prevent norovirus infection.

Conclusion: This oral norovirus vaccine was well-tolerated and generated substantial immune responses, including systemic and mucosal antibodies as well as memory IgA/IgG. These results are a major step forward for the development of a safe and immunogenic oral norovirus vaccine.

Trial registration: ClinicalTrials.gov NCT02868073.

Funding: Vaxart.

Keywords: Adaptive immunity; Vaccines.

Conflict of interest statement

Conflict of interest: L. Kim, K. Lin, K. Kasparek, K. Gottlieb, D. Liebowitz, G. Trager, S.J. Garg, and S.N. Tucker are employees of Vaxart and have received stock options and compensation as part of their employment.

Figures

Figure 1. Trial profile, including enrollment.

Figure 2. Immune responses to norovirus.

Unless otherwise noted, horizontal bars represent average increase, with thinner lines indicating 95% confidence interval. n = 20 for placebo group, n = 23 for the vaccine groups. (A) IgA or IgG ELISA titers after immunization. Data represent fold increases in antibody levels 28 days after vaccination (compared with day 0) for all subjects divided by treatment group (each symbol represents an individual subject). Significance was assessed by Kruskal-Wallis. (B) Norovirus VP1-specific IgG and IgA ASC counts on day 7 after vaccination. Significance was assessed by Kruskal-Wallis. (C) Norovirus VP1-specific memory B cell counts. Longer black horizontal lines represent the geometric mean, and error bars indicate the 95% confidence interval. Significance was assessed by Kruskal-Wallis. (D) Norovirus VP-specific fecal responses. Samples were removed when total IgA was below the detection limit. Sample size of each group are as follows: high dose (day 0–28), n = 19; high dose (day 0–180), n = 21; low dose (day 0–28), n = 20; low dose (day 0–180), n = 16; placebo (day 0–28), n = 18; placebo (day 0–180), n = 16. Significance was assessed by Mann-Whitney or Fisher’s exact test. (E) Norovirus VP-specific saliva IgA responses. In D and E, data represent fold increase in specific IgA/total IgA for each group with each time pair point plotted.

Figure 3. Generation of intestinal homing plasmablasts and memory B cells after vaccination.

(A) A population of β7hi B cells was detected 7 days after vaccination. Note that CD19 expression is slightly downregulated in β7hi B cells. Based on the levels of β7, 3 populations were present on day 7 B cells: β7hi (noted as 1), β7intermediate (noted as 2), and β7– (noted as 3). (B) Expression of IgA and IgG in β7hi (noted as 1), β7intermediate (noted as 2), and β7– B cells (noted as 3). Higher frequencies of IgA-expressing B cells were present in the β7hi population than in other populations. (C) β7hi B cells (noted as 1) coexpress high levels of the activation marker CD38 and CCR9 (red). The light blue color represents CCR9 expression on β7– B cells (noted as 3). (D) β7hi B cells (noted as 1) contain both CD27hi plasmablasts and CD27intermediate memory B cells. Populations 4 and 5 are defined by CD27 expression level. (E) FSC and SSC plots gated from populations 4 and 5. (F) Surface expression of IgA and IgG from populations 4 and 5. (G) The gating strategy for the graph in H: CD27hi B cells appeared on day 7; IgA+ B cells were gated from CD27hi B cells; and then β7hi B cells were gated from CD27hiIgA+ B cells. (H) The percentage of β7hi cells gated from CD27hiIgA+ B cells in day 7 PBMCs. Subjects from low-dose (1 × 1010) and high-dose (1 × 1011) groups were examined. The black horizontal bars represent means, with error bars indicating the 95% confidence interval. For A–G, data are from one subject, but the analysis was performed on all 66 subjects, and the results are representative of the responding vaccine subjects (>70%). H shows the distribution of β7 on all vaccine-treated subjects.