Depolymerization of SUMO chains induces slender to stumpy differentiation in T. brucei bloodstream parasites.

Trypanosoma brucei are protozoan parasites that cause sleeping sickness in humans and nagana in cattle. Inside the mammalian host, a quorum sensing-like mechanism coordinates its differentiation from a slender replicative form into a quiescent stumpy form, limiting growth and activating metabolic pathways that are beneficial to the parasite in the insect host. The post-translational modification of proteins with the Small Ubiquitin-like MOdifier (SUMO) enables dynamic regulation of cellular metabolism. SUMO can be conjugated to its targets as a monomer but can also form oligomeric chains. Here, we have investigated the role of SUMO chains in T. brucei by abolishing the ability of SUMO to polymerize. We have found that parasites able to conjugate only SUMO monomers are primed for differentiation. This was demonstrated for monomorphic lines that are normally unable to produce stumpy forms in response to quorum sensing signaling in mice, and also for pleomorphic cell lines in which stumpy cells were observed at unusually low parasitemia levels. SUMO chain mutants showed a stumpy compatible transcriptional profile and better competence to differentiate into procyclics. Our study indicates that SUMO depolymerization may represent a coordinated signal triggered during stumpy activation program.

A Gamma-adapted subunit vaccine induces broadly neutralizing antibodies against SARS-CoV-2 variants and protects mice from infection.

In the context of continuous emergence of SARS-CoV-2 variants of concern (VOCs), one strategy to prevent the severe outcomes of COVID-19 is developing safe and effective broad-spectrum vaccines. Here, we present preclinical studies of a RBD vaccine derived from the Gamma SARS-CoV-2 variant adjuvanted with Alum. The Gamma-adapted RBD vaccine is more immunogenic than the Ancestral RBD vaccine in terms of inducing broader neutralizing antibodies. The Gamma RBD presents more immunogenic B-cell restricted epitopes and induces a higher proportion of specific-B cells and plasmablasts than the Ancestral RBD version. The Gamma-adapted vaccine induces antigen specific T cell immune responses and confers protection against Ancestral and Omicron BA.5 SARS-CoV-2 challenge in mice. Moreover, the Gamma RBD vaccine induces higher and broader neutralizing antibody activity than homologous booster vaccination in mice previously primed with different SARS-CoV-2 vaccine platforms. Our study indicates that the adjuvanted Gamma RBD vaccine is highly immunogenic and a broad-spectrum vaccine candidate.

Regulation of human neutrophil IL-1ß secretion induced by Escherichia coli O157:H7 responsible for hemolytic uremic syndrome.

Shiga-toxin producing Escherichia coli (STEC) infections can cause from bloody diarrhea to Hemolytic Uremic Syndrome. The STEC intestinal infection triggers an inflammatory response that can facilitate the development of a systemic disease. We report here that neutrophils might contribute to this inflammatory response by secreting Interleukin 1 beta (IL-1ß). STEC stimulated neutrophils to release elevated levels of IL-1ß through a mechanism that involved the activation of caspase-1 driven by the NLRP3-inflammasome and neutrophil serine proteases (NSPs). Noteworthy, IL-1ß secretion was higher at lower multiplicities of infection. This secretory profile modulated by the bacteria:neutrophil ratio, was the consequence of a regulatory mechanism that reduced IL-1ß secretion the higher were the levels of activation of both caspase-1 and NSPs, and the production of NADPH oxidase-dependent reactive oxygen species. Finally, we also found that inhibition of NSPs significantly reduced STEC-triggered IL-1ß secretion without modulating the ability of neutrophils to kill the bacteria, suggesting NSPs might represent pharmacological targets to be evaluated to limit the STEC-induced intestinal inflammation.

Safety and immunogenicity of a SARS-CoV-2 Gamma variant RBD-based protein adjuvanted vaccine used as booster in healthy adults.

A Gamma Variant RBD-based aluminum hydroxide adjuvanted vaccine called ARVAC CG was selected for a first in human clinical trial. Healthy male and female participants (18-55 years old) with a complete COVID-19-primary vaccine scheme were assigned to receive two intramuscular doses of either a low-dose or a high-dose of ARVAC CG. The primary endpoint was safety. The secondary objective was humoral immunogenicity. Cellular immune responses were studied as an exploratory objective. The trial was prospectively registered in PRIISA.BA (Registration Code 6564) and ANMAT and retrospectively registered in ClinicalTrials.gov (NCT05656508). Samples from participants of a surveillance strategy implemented by the Ministry of Health of the Province of Buenos Aires that were boosted with BNT162b2 were also analyzed to compare with the booster effect of ARVAC CG. ARVAC CG exhibits a satisfactory safety profile, a robust and broad booster response of neutralizing antibodies against the Ancestral strain of SARS-CoV-2 and the Gamma, Delta, Omicron BA.1 and Omicron BA.5 variants of concern and a booster effect on T cell immunity in individuals previously immunized with different COVID-19 vaccine platforms.

Bm Delta-pgm, a vaccine for the control of Brucella melitensis with cross-species protective properties.

Brucellosis remains one of the most worldwide distributed zoonosis inflicting serious economical and human health problems in many areas of the world. The disease is caused by different species of the genus Brucella that have different tropisms towards different mammals being the most relevant for human health Brucella abortus, Brucella melitensis and Brucella suis that infect cows, goats/sheep, and swine respectively. For B. melitensis, considered the species with more zoonotic potential and highly aggressive for animals, only one vaccine is available to date in the market: Rev 1. This attenuated strain has the disadvantage that is has a very high residual virulence for animals and humans and, for this reason, it is applied by ocular instillation which is technically challenging in many productive settings. For this reason, the search for new vaccines for caprine and ovine brucellosis is an active topic of research. We describe here the construction of a novel highly attenuated vaccine strain (Bm Delta-pgm) that confers excellent levels of protection against B. melitensis in the mouse model of infection. This strain is a clean deletion of the phosphoglucomutase (pgm) gene that codes for a protein that catalyzes the conversion of glucose-6-P to glucose-1-P, which is used as a precursor for the biosynthesis of many polysaccharides, including the O-antigen of the lipopolysaccharide and cyclic beta glucans. Our results indicate that vaccination with Bm Delta-pgm induces a robust memory cellular immune response but no antibody production against the O-antigen. Cross protection experiments show that this new vaccine protects against B. abortus and B. suis raising the possibility that Bm Delta-pgm could be used as a universal vaccine for the most important Brucella species.

Design and characterization of chimeric Rabies-SARS-CoV-2 virus-like particles for vaccine purposes.

Due to the high number of doses required to achieve adequate coverage in the context of COVID-19 pandemics, there is a great need for novel vaccine developments. In this field, there have been research approaches that focused on the production of SARS-CoV-2 virus-like particles. These are promising vaccine candidates as their structure is similar to that of native virions but they lack the genome, constituting a biosafe alternative. In order to produce these structures using mammal cells, it has been established that all four structural proteins must be expressed. Here we report the generation and characterization of a novel chimeric virus-like particle (VLP) that can be produced by the expression of a single novel fusion protein that contains SARS-CoV-2 spike (S) ectodomain fused to rabies glycoprotein membrane anchoring region in HEK293 cells. This protein is structurally similar to native S and can autonomously bud forming enveloped VLPs that resemble native virions both in size and in morphology, displaying S ectodomain and receptor binding domain (RBD) on their surface. As a proof of concept, we analyzed the immunogenicity of this vaccine candidate in mice and confirmed the generation of anti-S, anti-RBD, and neutralizing antibodies. KEY POINTS: • A novel fusion rabies glycoprotein containing S ectodomain was designed. • Fusion protein formed cVLPs that were morphologically similar to SARS-CoV-2 virions. • cVLPs induced anti-S, anti-RBD, and neutralizing antibodies in mice.

A COVID-19 vaccine candidate based on SARS-CoV-2 spike protein and immune-stimulating complexes.

Spike protein from SARS-CoV-2, the etiologic agent of the COVID-19 pandemic disease, constitutes a structural protein that proved to be the main responsible for neutralizing antibody production. Thus, its sequence is highly considered for the design of candidate vaccines. Animal cell culture represents the best option for the production of subunit vaccines based on recombinant proteins since they introduce post-translational modifications that are important to mimic the natural antigenic epitopes. Particularly, the human cell line HEK293T has been explored and used for the production of biotherapeutics since the products derived from them present human-like post-translational modifications that are important for the protein's activity and immunogenicity. The aim of this study was to produce and characterize a potential vaccine for COVID-19 based on the spike ectodomain (S-ED) of SARS-CoV-2 and two different adjuvants: aluminum hydroxide (AH) and immune-stimulating complexes (ISCOMs). The S-ED was produced in sHEK293T cells using a 1-L stirred tank bioreactor operated in perfusion mode and purified. S-ED characterization revealed the expected size and morphology. High N-glycan content was confirmed. S-ED-specific binding with the hACE2 (human angiotensin-converting enzyme 2) receptor was verified. The immunogenicity of S-ED was evaluated using AH and ISCOMs. Both formulations demonstrated the presence of anti-RBD antibodies in the plasma of immunized mice, being significantly higher for the latter adjuvant. Also, higher levels of IFN-γ and IL-4 were detected after the ex vivo immune stimulation of spleen-derived MNCs from ISCOMs immunized mice. Further analysis confirmed that S-ED/ISCOMs elicit neutralizing antibodies against SARS-CoV-2. KEY POINTS: Trimeric SARS-CoV-2 S-ED was produced in stable recombinant sHEK cells in serum-free medium. A novel S-ED vaccine formulation induced potent humoral and cellular immunity. S-ED formulated with ISCOMs adjuvant elicited a highly neutralizing antibody titer.

A disordered region retains the full protease inhibitor activity and the capacity to induce CD8+ T cells in vivo of the oral vaccine adjuvant U-Omp19.

U-Omp19 is a bacterial protease inhibitor from Brucella abortus that inhibits gastrointestinal and lysosomal proteases, enhancing the half-life and immunogenicity of co-delivered antigens. U-Omp19 is a novel adjuvant that is in preclinical development with various vaccine candidates. However, the molecular mechanisms by which it exerts these functions and the structural elements responsible for these activities remain unknown. In this work, a structural, biochemical, and functional characterization of U-Omp19 is presented. Dynamic features of U-Omp19 in solution by NMR and the crystal structure of its C-terminal domain are described. The protein consists of a compact C-terminal beta-barrel domain and a flexible N-terminal domain. The latter domain behaves as an intrinsically disordered protein and retains the full protease inhibitor activity against pancreatic elastase, papain and pepsin. This domain also retains the capacity to induce CD8+ T cells in vivo of U-Omp19. This information may lead to future rationale vaccine designs using U-Omp19 as an adjuvant to deliver other proteins or peptides in oral formulations against infectious diseases, as well as to design strategies to incorporate modifications in its structure that may improve its adjuvanticity.

A Novel Bacterial Protease Inhibitor Adjuvant in RBD-Based COVID-19 Vaccine Formulations Containing Alum Increases Neutralizing Antibodies, Specific Germinal Center B Cells and Confers Protection Against SARS-CoV-2 Infection in Mice.

In this work, we evaluated recombinant receptor binding domain (RBD)-based vaccine formulation prototypes with potential for further clinical development. We assessed different formulations containing RBD plus alum, AddaS03, AddaVax, or the combination of alum and U-Omp19: a novel Brucella spp. protease inhibitor vaccine adjuvant. Results show that the vaccine formulation composed of U-Omp19 and alum as adjuvants has a better performance: it significantly increased mucosal and systemic neutralizing antibodies in comparison to antigen plus alum, AddaVax, or AddaS03. Antibodies induced with the formulation containing U-Omp19 and alum not only increased their neutralization capacity against the ancestral virus but also cross-neutralized alpha, lambda, and gamma variants with similar potency. Furthermore, the addition of U-Omp19 to alum vaccine formulation increased the frequency of RBD-specific geminal center B cells and plasmablasts. Additionally, U-Omp19+alum formulation induced RBD-specific Th1 and CD8+ T-cell responses in spleens and lungs. Finally, this vaccine formulation conferred protection against an intranasal severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) challenge of K18-hACE2 mice.

Stability Studies of the Vaccine Adjuvant U-Omp19.

Unlipidated outer membrane protein 19 (U-Omp19) is a novel mucosal adjuvant in preclinical development to be used in vaccine formulations. U-Omp19 holds two main properties, it is capable of inhibiting gastrointestinal and lysosomal peptidases, increasing the amount of co-administered antigen that reaches the immune inductive sites and its half-life inside cells, and it is able to stimulate antigen presenting cells in vivo. These activities enable U-Omp19 to enhance the adaptive immune response to co-administrated antigens. To characterize the stability of U-Omp19 we have performed an extensive analysis of its physicochemical and biological properties in a 3-year long-term stability study, and under potentially damaging freeze-thawing and lyophilization stress processes. Results revealed that U-Omp19 retains its full protease inhibitor activity, its monomeric state and its secondary structure even when stored in solution for 36 months or after multiple freeze-thawing cycles. Non-enzymatic hydrolysis resulted the major degradation pathway for storage in solution at 4 °C or room temperature which can be abrogated by lyophilization yet increasing protein tendency to form aggregates. This information will play a key role in the development of a stable formulation of U-Omp19, allowing an extended shelf-life during manufacturing, storage, and shipping of a future vaccine containing this pioneering adjuvant.

The Trypanosoma cruzi TcTASV-C protein subfamily administrated with U-Omp19 promotes a protective response against a lethal challenge in mice.

The development of a Chagas disease vaccine has yet the need for the identification of novel combinations of antigens and adjuvants. Here, the performance of TcTASV-C proteins that are virulence factors of trypomastigotes and belong to a novel surface protein family specific for T. cruzi, have been evaluated as antigens for a prophylactic vaccine. Several immunization schemes in which TcTASV-C was combined with aluminum hydroxide, saponin and/or U-Omp19 were assayed. Aluminum hydroxide and saponin were assayed together to trigger different pathways of the immune response simultaneously. U-Omp19 is a promising novel adjuvant able to promote a Th1 immune response with IFNg production, thus an interesting molecule to be tested as adjuvant for the control of T. cruzi infection. Therefore, U-Omp19 was added to the aluminum hydroxide-saponin formulation as well as assayed individually with TcTASV-C. The immunization with TcTASV-C and U-Omp19 had the best performance as a prophylactic vaccine. Mice presented the lowest parasitemias and improved survival by 40% after being challenged with a highly virulent T. cruzi strain, which promoted 100% mortality in all other immunized groups. Immunization with TcTASV-C and U-Omp19 triggered cellular responses with IFN-γ and IL-17 production and with lytic antibodies that could explain the protection achieved by this vaccination scheme. To our knowledge, this is the first time that U-Omp19 is tested with a defined T. cruzi antigen in a vaccine formulation.

U-Omp19 from Brucella abortus increases dmLT immunogenicity and improves protection against Escherichia coli heat-labile toxin (LT) oral challenge.

Acute diarrhea disease caused by bacterial infections is a major global health problem. Enterotoxigenic Escherichia coli (ETEC) is one of the top causes of diarrhea-associated morbidity and mortality in young children and travelers to low-income countries. There are currently no licensed vaccines for ETEC. Induction of immunity at the site of entry of the bacteria is key to prevent infection. Current approaches to ETEC vaccines include a less toxic mutant form of E. coli heat-labile toxin (double-mutant heat-labile enterotoxin -dmLT-) with both antigenic and immunostimulatory properties. U-Omp19 is a protease inhibitor from Brucella spp. with immunostimulatory properties that has been used as oral adjuvant. In this work, we use U-Omp19 as adjuvant in an oral vaccine formulation against ETEC containing dmLT in outbred and inbred mice. To evaluate antigen dose sparing by U-Omp19 three different immunization protocols with three different doses of dmLT were evaluated. We demonstrated that U-Omp19 co-delivery increases anti-LT IgA in feces using a mid-dose of dmLT following a prime-boost protocol (after one or two boosts). Oral immunization with U-Omp19 induced protection against LT challenge when co-formulated with dmLT in CD-1 and BALB/c mice. Indeed, there was a significant increase in anti-LT IgG and IgA avidity after a single oral administration of dmLT plus U-Omp19 in comparison with dmLT delivered alone. Interestingly, sera from dmLT plus U-Omp19 vaccinated mice significantly neutralize LT effect on intestine inflammation in vivo compared with sera from the group immunized with dmLT alone. These results demonstrate the adjuvant capacity of U-Omp19 to increase dmLT immunogenicity by the oral route and support its use in an oral subunit vaccine formulation against ETEC.

Sublingual Omp16-driven redirection of the allergic intestinal response in a pre-clinical model of food allergy.

BACKGROUND: IgE-mediated food allergy remains a significant and growing worldwide problem. Sublingual immunotherapy (SLIT) shows an excellent safety profile for food allergy, but the clinical efficacy needs to be improved. This study assessed the effects of the Toll-like receptor 4 agonist outer membrane protein (Omp) 16 from Brucella abortus combined with cow´s milk proteins (CMP) through the sublingual route to modulate cow's milk allergy in an experimental model. METHODS: Mice sensitized with cholera toxin and CMP were orally challenged with the allergen to elicit hypersensitivity reactions. Then, mice were treated with a very low amount of CMP along with Omp16 as a mucosal adjuvant, and finally, animals were re-exposed to CMP. Systemic and mucosal immune parameters were assessed in vivo and in vitro. RESULTS: We found that the sublingual administration of Omp16 + CMP induced a buccal Th1 immune response that modulated the intestinal allergic response with the suppression of symptoms, reduction of IgE and IL-5, and up-regulation of IgG2a and IFN-γ. The adoptive transfer of submandibular IFN-γ-producing α4ß7+ CD4+ and CD8+ cells conferred protection against allergic sensitization. The use of Omp16 + CMP promoted enhanced protection compared to CMP alone. CONCLUSION: In conclusion, Omp16 represents a promising mucosal adjuvant that can be used to improve the clinical and immune efficacy of SLIT for food allergy.

Omp25-dependent engagement of SLAMF1 by Brucella abortus in dendritic cells limits acute inflammation and favours bacterial persistence in vivo.

The strategies by which intracellular pathogenic bacteria manipulate innate immunity to establish chronicity are poorly understood. Here, we show that Brucella abortus outer membrane protein Omp25 specifically binds the immune cell receptor SLAMF1 in vitro. The Omp25-dependent engagement of SLAMF1 by B. abortus limits NF-κB translocation in dendritic cells (DCs) with no impact on Brucella intracellular trafficking and replication. This in turn decreases pro-inflammatory cytokine secretion and impairs DC activation. The Omp25-SLAMF1 axis also dampens the immune response without affecting bacterial replication in vivo during the acute phase of Brucella infection in a mouse model. In contrast, at the chronic stage of infection, the Omp25/SLAMF1 engagement is essential for Brucella persistence. Interaction of a specific bacterial protein with an immune cell receptor expressed on the DC surface at the acute stage of infection is thus a powerful mechanism to support microbe settling in its replicative niche and progression to chronicity.

Omp19 Enables Brucella abortus to Evade the Antimicrobial Activity From Host's Proteolytic Defense System.

Pathogenic microorganisms confront several proteolytic events in the molecular interplay with their host, highlighting that proteolysis and its regulation play an important role during infection. Microbial inhibitors, along with their target endogenous/exogenous enzymes, may directly affect the host's defense mechanisms and promote infection. Omp19 is a Brucella spp. conserved lipoprotein anchored by the lipid portion in the Brucella outer membrane. Previous work demonstrated that purified unlipidated Omp19 (U-Omp19) has protease inhibitor activity against gastrointestinal and lysosomal proteases. In this work, we found that a Brucella omp19 deletion mutant is highly attenuated in mice when infecting by the oral route. This attenuation can be explained by bacterial increased susceptibility to host proteases met by the bacteria during establishment of infection. Omp19 deletion mutant has a cell division defect when exposed to pancreatic proteases that is linked to cell-cycle arrest in G1-phase, Omp25 degradation on the cell envelope and CtrA accumulation. Moreover, Omp19 deletion mutant is more susceptible to killing by macrophage derived microsomes than wt strain. Preincubation with gastrointestinal proteases led to an increased susceptibility of Omp19 deletion mutant to macrophage intracellular killing. Thus, in this work, we describe for the first time a physiological function of B. abortus Omp19. This activity enables Brucella to better thrive in the harsh gastrointestinal tract, where protection from proteolytic degradation can be a matter of life or death, and afterwards invade the host and bypass intracellular proteases to establish the chronic infection.

Oral co-administration of a bacterial protease inhibitor in the vaccine formulation increases antigen delivery at the intestinal epithelial barrier.

The study of capture and processing of antigens (Ags) by intestinal epithelial cells is very important for development of new oral administration systems. Efficient oral Ag delivery systems must resist enzymatic degradation by gastric and intestinal proteases and deliver the Ag across biological barriers. The recombinant unlipidated outer membrane protein from Brucella spp. (U-Omp19) is a protease inhibitor with immunostimulatory properties used as adjuvant in oral vaccine formulations. In the present work we further characterized its mechanism of action and studied the interaction and effect of U-Omp19 on the intestinal epithelium. We found that U-Omp19 inhibited protease activity from murine intestinal brush-border membranes and cysteine proteases from human intestinal epithelial cells (IECs) promoting co-administered Ag accumulation within lysosomal compartments of IECs. In addition, we have shown that co-administration of U-Omp19 facilitated the transcellular passage of Ag through epithelial cell monolayers in vitro and in vivo while did not affect epithelial cell barrier permeability. Finally, oral co-delivery of U-Omp19 in mice induced the production of Ag-specific IgA in feces and the increment of CD103+ CD11b- CD8α+ dendritic cells subset at Peyer's patches. Taken together, these data describe a new mechanism of action of a mucosal adjuvant and support the use of this rationale/strategy in new oral delivery systems for vaccines.

RomA, A Periplasmic Protein Involved in the Synthesis of the Lipopolysaccharide, Tunes Down the Inflammatory Response Triggered by Brucella.

Brucellaceae are stealthy pathogens with the ability to survive and replicate in the host in the context of a strong immune response. This capacity relies on several virulence factors that are able to modulate the immune system and in their structural components that have low proinflammatory activities. Lipopolysaccharide (LPS), the main component of the outer membrane, is a central virulence factor of Brucella, and it has been well established that it induces a low inflammatory response. We describe here the identification and characterization of a novel periplasmic protein (RomA) conserved in alpha-proteobacteria, which is involved in the homeostasis of the outer membrane. A mutant in this gene showed several phenotypes, such as membrane defects, altered LPS composition, reduced adhesion, and increased virulence and inflammation. We show that RomA is involved in the synthesis of LPS, probably coordinating part of the biosynthetic complex in the periplasm. Its absence alters the normal synthesis of this macromolecule and affects the homeostasis of the outer membrane, resulting in a strain with a hyperinflammatory phenotype. Our results suggest that the proper synthesis of LPS is central to maximize virulence and minimize inflammation.

U-Omp19 from Brucella abortus Is a Useful Adjuvant for Vaccine Formulations against Salmonella Infection in Mice.

Most pathogens infect through mucosal surfaces, and parenteral immunization typically fails to induce effective immune responses at these sites. Development of oral-administered vaccines capable of inducing mucosal as well as systemic immunity while bypassing the issues of antigen degradation and immune tolerance could be crucial for the control of enteropathogens. This study demonstrates that U-Omp19, a bacterial protease inhibitor with immunostimulatory features, coadministered with Salmonella antigens by the oral route, enhances mucosal and systemic immune responses in mice. U-Omp19 was able to increase antigen-specific production of IFN-γ and IL-17 and mucosal (IgA) antibody response. Finally, oral vaccination with U-Omp19 plus Salmonella antigens conferred protection against virulent challenge with Salmonella Typhimurium, with a significant reduction in bacterial loads. These findings prove the efficacy of this novel adjuvant in the Salmonella infection model and support the potential of U-Omp19 as a suitable adjuvant in oral vaccine formulations against mucosal pathogens requiring T helper (Th)1-Th17 protective immune responses.

A Brucella spp. Protease Inhibitor Limits Antigen Lysosomal Proteolysis, Increases Cross-Presentation, and Enhances CD8+ T Cell Responses.

In this study, we demonstrate that the unlipidated (U) outer membrane protein (Omp) 19 from Brucella spp. is a competitive inhibitor of human cathepsin L. U-Omp19 inhibits lysosome cathepsins and APC-derived microsome activity in vitro and partially inhibits lysosomal cathepsin L activity within live APCs. Codelivery of U-Omp19 with the Ag can reduce intracellular Ag digestion and increases Ag half-life in dendritic cells (DCs). U-Omp19 retains the Ag in Lamp-2(+) compartments after its internalization and promotes a sustained expression of MHC class I/peptide complexes in the cell surface of DCs. Consequently, U-Omp19 enhances Ag cross-presentation by DCs to CD8(+) T cells. U-Omp19 s.c. delivery induces the recruitment of CD11c(+)CD8α(+) DCs and monocytes to lymph nodes whereas it partially limits in vivo Ag proteolysis inside DCs. Accordingly, this protein is able to induce CD8(+) T cell responses in vivo against codelivered Ag. Antitumor responses were elicited after U-Omp19 coadministration, increasing survival of mice in a murine melanoma challenge model. Collectively, these results indicate that a cysteine protease inhibitor from bacterial origin could be a suitable component of vaccine formulations against tumors.

Brucella abortus Omp19 recombinant protein subcutaneously co-delivered with an antigen enhances antigen-specific T helper 1 memory responses and induces protection against parasite challenge.

The discovery of effective adjuvants for many vaccines especially those with limited commercial appeal, such as vaccines to poverty-related diseases, is required. In this work, we demonstrated that subcutaneous co-administration of mice with the outer membrane protein U-Omp19 from Brucella spp. plus OVA as antigen (Ag) increases Ag-specific T cell proliferation and T helper (Th) 1 immune responses in vitro and in vivo. U-Omp19 treated dendritic cells promote IFN-γ production by specific CD4(+) T cells and increases T cell proliferation. U-Omp19 co-administration induces the production of Ag specific effector memory T cell populations (CD4(+) CD44(high) CD62L(low) T cells). Finally, subcutaneous co-administration of U-Omp19 with Trypanosoma cruzi Ags confers protection against virulent parasite challenge, reducing parasitemia and weight loss while increasing mice survival. These results indicate that the bacterial protein U-Omp19 when delivered subcutaneously could be a suitable component of vaccine formulations against infectious diseases requiring Th1 immune responses.