Local and Systemic Immunity against Respiratory Syncytial Virus Induced by a Novel Intranasal Vaccine. A Randomized, Double-Blind, Placebo-controlled Clinical Trial

Abstract

Rationale: Needle-free intranasal vaccines offer major potential advantages, especially against pathogens entering via mucosal surfaces. As yet, there is no effective vaccine against respiratory syncytial virus (RSV), a ubiquitous pathogen of global importance that preferentially infects respiratory epithelial cells; new strategies are urgently required.Objectives: Here, we report the safety and immunogenicity of a novel mucosal RSV F protein vaccine linked to an immunostimulatory bacterium-like particle (BLP).Methods: In this phase I, randomized, double-blind, placebo-controlled trial, 48 healthy volunteers, aged 18-49 years, were randomly assigned to receive placebo or SynGEM (low or high dose) intranasally by prime-boost administration. The primary outcome was safety and tolerability, with secondary objectives assessing virus-specific immunogenicity.Measurements and Main Results: There were no significant differences in adverse events between placebo and vaccinated groups. SynGEM induced systemic plasmablast responses and significant, durable increases in RSV-specific serum antibody in healthy, seropositive adults. Volunteers given low-dose SynGEM (140 μg F, 2 mg BLP) required a boost at Day 28 to achieve plateau responses with a maximum fold change of 2.4, whereas high-dose recipients (350 μg F, 5 mg BLP) achieved plateau responses with a fold change of 1.5 after first vaccination that remained elevated up to 180 days after vaccination, irrespective of further boosting. Palivizumab-like antibodies were consistently induced, but F protein site ∅-specific antibodies were not detected, and virus-specific nasal IgA responses were heterogeneous, with the strongest responses in individuals with lower pre-existing antibody levels.Conclusions: SynGEM is thus the first nonreplicating intranasal RSV subunit vaccine to induce persistent antibody responses in human volunteers.Clinical trials registered with www.clinicaltrials.gov (NCT02958540).

Keywords: clinical trial; immunology; mucosal; respiratory; virus.

Figure 1.

Intranasal SynGEM induces significant increases in F-specific serum IgG. Volunteers were given SynGEM or placebo and serum IgG was measured by ELISA using unstabilized F protein as coating antigen at time points up to 180 days after “prime.” Titers (scatter plots) and fold changes (box-and-whisker plots) compared with baseline are shown after (A and B) placebo, (C and D) low dose, and (E and F) high dose. Geometric means are shown in red. Wilcoxon signed-rank test was used to test statistically significant rises compared with prevaccination; *P < 0.05, ***P < 0.001, and ****P < 0.0001. For box-and-whisker plots, boxes indicate the median and interquartile range and whiskers the range of the fold changes for each time point after vaccination. Vaccinations are indicated by red triangles.

Figure 2.

SynGEM induces palivizumab-like, but not prefusion F-specific (pre-F), antibodies. Participants were given SynGEM or placebo. (A, D, and G) Serum neutralizing antibody titers were measured by classical plaque-reduction neutralization assay up to 28 days after boost. (B, C, E, F, H, and I) Palivizumab-like (B, E, and H) and D25-like (C, F, and I) antibodies were measured by competition ELISA up to 28 days after boost. Geometric means are shown in red. Wilcoxon signed-rank test was used to test statistically significant rises compared with prevaccination; **P < 0.01, ***P < 0.001, and ****P < 0.0001. Vaccinations are indicated by red triangles.

Figure 3.

Intranasal SynGEM protein induces heterogeneous mucosal IgA responses. Subjects were given SynGEM or placebo and nasal wash IgA was measured by ELISA using unstabilized F protein as coating antigen at time points up to 180 days after “prime.” (A–C) Individuals in the placebo (A), low-dose (B), and high-dose (C) groups were clustered if they displayed a greater than twofold rise in nasal IgA titers according to the time point of maximal increase. (D–F) Cluster analysis of individual participant-level data was carried out by Fisher's exact test on placebo (D), low-dose (E), and high-dose (F) groups at the indicated time points after vaccination. **P < 0.01. Vaccinations are indicated by red triangles.

Figure 4.

Intranasal SynGEM protein stimulates IgG⁺ and IgA⁺ antibody-secreting cells (ASCs) in peripheral blood from volunteers administered low or high doses of SynGEM. (A–F) ASCs from peripheral blood at time points up to 28 days after boost were enumerated by B cell ELISpot. (G and H) There was no correlation between IgG⁺ ASC frequencies and F-specific serum IgG titers (G) or IgA⁺ ASC frequencies and F-specific nasal IgA titers (H). Median values are shown in red. *P < 0.05, ***P < 0.001, and ****P < 0.0001. PBMCs = peripheral blood mononuclear cells. Vaccinations are indicated by red triangles.

Figure 5.

SynGEM (bacterium-like particle [BLP]-F) stimulation of adenotonsillar cells provokes dose-dependent antibody and T cell responses. Tonsil cells from (A–C) adult and (D–F) pediatric donors were cultured with SynGEM (BLP-F) containing 5 μg/ml and 1 μg/ml F protein, BLP alone (25 μg/ml), F protein alone (1 μg/ml), and medium only. F-specific IgG, IgA, and IgM in resulting supernatant were measured by ELISA. (G–I) Cytokines were measured in culture supernatant by cytometric bead array. Tonsil cells were cultured with SynGEM (BLP-F), BLP alone (25 μg/ml), or F protein alone (1 μg/ml). (J) CD4⁺ and (K) CD8⁺ T cell proliferation was then measured by analysis of CFSE dilution and expressed as a percentage of dividing cells in the CD3⁺ CD4⁺ or CD3⁺ CD8⁺ populations. Mann-Whitney U test and Wilcoxon signed-rank test were used to test significant differences. *P < 0.05 and **P < 0.005. NALT = nasal-associated lymphoid tissue.