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Adjuvanted soluble protein vaccines have been used extensively in humans for protection against various viral infections based on their robust induction of antibody responses. Here, soluble prefusion-stabilized spike protein trimers (preS dTM) from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were formulated with the adjuvant AS03 and administered twice to nonhuman primates (NHPs). Binding and functional neutralization assays and systems serology revealed that the vaccinated NHP developed AS03-dependent multifunctional humoral responses that targeted distinct domains of the spike protein and bound to a variety of Fc receptors mediating immune cell effector functions in vitro. The neutralizing 50% inhibitory concentration titers for pseudovirus and live SARS-CoV-2 were higher than titers for a panel of human convalescent serum samples. NHPs were challenged intranasally and intratracheally with a high dose (3 × 106 plaque forming units) of SARS-CoV-2 (USA-WA1/2020 isolate). Two days after challenge, vaccinated NHPs showed rapid control of viral replication in both the upper and lower airways. Vaccinated NHPs also had increased spike protein-specific immunoglobulin G (IgG) antibody responses in the lung as early as 2 days after challenge. Moreover, passive transfer of vaccine-induced IgG to hamsters mediated protection from subsequent SARS-CoV-2 challenge. These data show that antibodies induced by the AS03-adjuvanted preS dTM vaccine were sufficient to mediate protection against SARS-CoV-2 in NHPs and that rapid anamnestic antibody responses in the lung may be a key mechanism for protection.

Enterotoxigenic E. coli (ETEC) is a leading cause of moderate-to-severe diarrhoea. ETEC colonizes the intestine through fimbrial tip adhesin colonization factors and produces heat-stable and/or heat-labile (LT) toxins, stimulating fluid and electrolyte release leading to watery diarrhoea. We reported that a vaccine containing recombinant colonization factor antigen (CfaEB) targeting fimbrial tip adhesin of the colonization factor antigen I (CFA/I) and an attenuated LT toxoid (dmLT) elicited mucosal and systemic immune responses against both targets. Additionally, the toll-like receptor 4 ligand second-generation lipid adjuvant (TLR4-SLA) induced a potent mucosal response, dependent on adjuvant formulation. However, a combination of vaccine components at their respective individual optimal doses may not achieve the optimal immune profile. We studied a subunit ETEC vaccine prototype in mice using a response surface design of experiments (DoE), consisting of 64 vaccine dose-combinations of CfaEB, dmLT and SLA in four formulations (aqueous, aluminium oxyhydroxide, squalene-in-water stable nanoemulsion [SE] or liposomes containing the saponin Quillaja saponaria-21 [LSQ]). Nine readouts focusing on antibody functionality and plasma cell response were selected to profile the immune response of parenterally administered ETEC vaccine prototype. The data were integrated in a model to identify the optimal dosage of each vaccine component and best formulation. Compared to maximal doses used in mouse models (10 µg CfaEB, 1 µg dmLT and 5 µg SLA), a reduction in the vaccine components up to 37%, 60% and 88% for CfaEB, dmLT and SLA, respectively, maintained or even maximized immune responses, with SE and LSQ the best formulations. The DoE approach can help determine the best vaccine composition with a limited number of experiments and may accelerate development of multi-antigen/component ETEC vaccines.

Many pathogens establish infection at mucosal surfaces such as the enteric pathogen Enterotoxigenic E. coli (ETEC). Thus, there is a pressing need for effective vaccination strategies that promote protective immunity at mucosal surfaces. Toll-like receptor (TLR) ligands have been extensively developed as vaccine adjuvants to promote systemic immunity, whereas attenuated bacterial toxins including cholera toxin and heat-labile toxin (LT) have initially been developed to promote mucosal immunity. Here we evaluate the ability of the TLR4 agonist second-generation lipid adjuvant formulated in a stable emulsion (SLA-SE) to augment functional mucosal antibodies elicited by intramuscular immunization with a recombinant ETEC vaccine antigen. We find that, in mice, parenterally delivered SLA-SE is at least as effective as the double-mutant LT (LTR192G/L211A, dmLT) adjuvant in promoting functional antibodies and eliciting intestinal IgA responses to the vaccine antigen. In addition, SLA-SE enhanced both the IgG2a response in the mucosa and serum, and the production of LT neutralizing serum antibodies elicited by dmLT four to eightfold. These results reveal unexpected mucosal adjuvant properties of this TLR4 agonist adjuvant when delivered intramuscularly. This may have a substantial impact on the development of vaccines against enteric and other mucosal pathogens., Mucosal immunity: parenteral vaccination hits the gut Although offering great potential for generating intestinal immunity, vaccination by the oral route suffers from several barriers such as the breakdown of protein vaccines in the stomach and/or the induction of oral tolerance. To investigate whether these barriers can be circumvented, Mark T. Orr and colleagues at the Infectious Disease Research Institute use a parenteral (intramuscular) vaccination protocol in mice. Intramuscular immunization with an enterotoxigenic E. coli (ETEC) vaccine plus a Toll-like receptor 4 adjuvant in stable emulsion (SLA-SE) elicits a functional antibody response in both the gut and serum. Importantly, this intramuscular vaccination triggers robust production of IgA in the gut. These findings suggest that with the right adjuvant combination it might possible to generate potent protective mucosal immunity following parenteral immunization.

Dendritic cells (DCs) play an essential role in the induction of immune responses to pathogen infections. Native DCs are difficult to obtain in large numbers and consequently the vast majority of DCs employed in all experiments are derived from bone marrow progenitors. In an attempt to solve this problem, we have established a novel CD8alpha(+) DC line (H-2(k)) from spleen, which we have named SRDC line, and which is easy to culture in vitro. These cells display similar morphology, phenotype and activity to CD4(-)CD8alpha(+)CD205(+)CD11b(-) DCs purified ex vivo. Toxoplasma gondii antigen was shown to be taken up by these cells and to increase class I and class II major histocompatibility complex (MHC), CD40, CD80 and CD86 surface expression. We report that vaccination with T. gondii antigen-pulsed SRDCs, which synthesize large amounts of interleukin-12, induced protective immune responses against this intracellular pathogen in syngeneic CBA/J mice. This protection was associated with strong cellular and humoral immune responses at systemic and intestinal levels. Spleen and mesenteric lymph node cell proliferations were correlated with a Th1/Th2-type response and a specific SRDC homing to spleen and intestine was observed. The SRDC or CD4(-)CD8alpha(+)CD205(+)CD11b(-) DC line can be expected to be a very useful tool for immunobiology studies of DC.

Our results show that exosomes secreted by SRDC pulsed in vitro with Toxoplasma gondii-derived antigens (Exo-TAg) induced protective responses against infection with the parasite in both syngeneic and allogeneic mice. After oral infection, syngeneic CBA/J mice exhibited significantly fewer cysts in their brains and allogeneic C57BL/6 mice survived. This protection was associated with strong humoral responses in vivo in serum from both CBA/J and C57BL/6 mice, and with high levels of anti-TAg IgA antibodies in intestinal secretions from CBA/J mice alone. Furthermore, strong cellular responses in vivo were observed in both mouse models. Cellular proliferation was associated with cytokines production by spleen and mesenteric lymph node cells. The results presented here show that exosomes are nucleic acid free vesicles that are able to induce immune responses correlated with protection against parasitic infections in both syngeneic and allogeneic mice. They could constitute an efficient tool for use in vaccination and antitumor strategies based on exosomes.