Immune response characterization in a human challenge study with a Shigella flexneri 2a bioconjugate vaccine

Abstract

Background: Diarrheal diseases are a leading cause of global morbidity and mortality affecting all ages, but especially children under the age of five in resource-limited settings. Shigella is a leading contributor to diarrheal diseases caused by bacterial pathogens and is considered a significant antimicrobial resistance threat. While improvements in hygiene, and access to clean water help as control measures, vaccination remains one of the most viable options for significantly reducing morbidity and mortality.

Methods: Flexyn2a is a bioconjugate vaccine manufactured using novel conjugation methodologies enzymatically linking the O-polysaccharide of S. flexneri 2a to exotoxin A of Pseudomonas aeruginosa. The protective capacity of Flexyn2a was assessed in a controlled human infection model after two intramuscular immunizations. Immune responses pre- and post-immunization and/or infection were investigated and are described here.

Findings: Flexyn2a induced lipopolysaccharide (LPS)-specific serum IgG responses post-immunization which were associated with protection against shigellosis. Additionally, several other immune parameters, including memory B cell responses, bactericidal antibodies and serum IgA, were also elevated in vaccinees protected against shigellosis. Immunization with Flexyn2a also induced gut-homing, LPS-specific IgG and IgA secreting B cells, indicating the vaccine induced immune effectors functioning at the site of intestinal infection.

Interpretation: Collectively, the results of these immunological investigations provide insights into protective immune mechanisms post-immunization with Flexyn2a which can be used to further guide vaccine development and may have applicability to the larger Shigella vaccine field.

Funding: Funding for this study was provided through a Wellcome Trust grant.

Keywords: Bioconjugate vaccine; Gut-homing responses; Human challenge; Immunogenicity; Parenteral immunization; Shigella.

Fig. 1

S. flexneri 2a LPS-Specific Serum IgG and IgA. (a) S. flexneri 2a LPS-specific serum IgG responses grouped by vaccinated subjects with or without per protocol shigellosis, and placebo subjects with or without per protocol shigellosis. (b) S. flexneri 2a LPS-specific serum IgG responses grouped by vaccinated subjects with or without consensus shigellosis, and placebo subjects with or without consensus shigellosis. (c) S. flexneri 2a LPS-specific serum IgA responses grouped by vaccinated subjects with or without consensus shigellosis, and placebo subjects with or without consensus shigellosis. * = significant difference as compared to baseline titres within the same treatment group/shigellosis outcome. Significance determined by repeated measures ANOVA of log-transformed titres with Bonferroni post-hoc test. (d) Percent efficacy against consensus shigellosis post-challenge in vaccinated subjects across increasing serum IgG and IgA ELISA endpoint titres.

Fig. 2

S. flexneri 2a LPS-Specific Serum IgM, IgG1 and IgG2. S. flexneri 2a LPS-specific serum IgM (a), IgG1 (b) and IgG2 (c) geometric mean ELISA endpoint titres grouped by vaccinated subjects with or without consensus shigellosis, and placebo subjects with or without consensus shigellosis. * = significant difference as compared to baseline titres within the same treatment group/shigellosis outcome. Significance determined by repeated measures ANOVA of log-transformed titres with Bonferroni post-hoc test. (d) Percent efficacy against consensus shigellosis post-challenge in vaccinated subjects across increasing serum IgG1 and IgG2 ELISA endpoint titres measured at time of challenge (day 56).

Fig. 3

Bactericidal responses. (a) S. flexneri 2a-specific geometric mean SBA endpoint titres grouped by vaccinated or placebo subjects. * = significant difference as compared to baseline titres within treatment group. Significance determined by repeated measures ANOVA of log-transformed titres with Bonferroni post-hoc test. (b) S. flexneri 2a-specific geometric mean SBA endpoint titres grouped by vaccinated subjects with or without consensus shigellosis, and placebo subjects with or without consensus shigellosis. * = significant difference as compared to baseline titres within the same treatment group/shigellosis outcome. Significance determined by repeated measures ANOVA of log-transformed titres with Bonferroni post-hoc test. (c) Percent efficacy against consensus shigellosis post-challenge in vaccinated subjects across increasing serum bactericidal titres measured at time of challenge (day 56).

Fig. 4

α4β7+ and α4β7- ALS IgG and IgA Responses. Individual S. flexneri 2a LPS-specific α4β7+ and α4β7- ALS IgG and IgA ELISA endpoint titres with group geometric mean and 95% confidence intervals either 7 days post-first immunization or 7 days post-challenge (day 63) grouped by treatment and consensus shigellosis outcome. (a) α4β7+ ALS IgG; (b) α4β7+ ALS IgA; (c) α4β7- ALS IgG; (d) α4β7- ALS IgA.

Fig. 5

Memory B cell responses. S. flexneri 2a LPS-specific memory B cell ALS IgG (a) and IgA (b) geometric mean ELISA endpoint titres with 95% confidence intervals at baseline (day 0), day of challenge (day 56) and 28 days post-challenge (day 84), grouped by vaccinated or placebo subjects. * = significant difference as compared to baseline titres within treatment group as determined by repeated measures ANOVA of log-transformed titres with Bonferroni post-hoc test.

Fig. 6

Spearman correlation of immune parameters in vaccinated subjects. Spearman correlation heat map of immune parameters in vaccinated subjects either 7 days post-first immunization (α4β7+ and α4β7- responses) or on day of challenge/day 56 (all other immune parameters).