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Introduction. Tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis (M. tb), remains a significant global public health concern. It is crucial to develop more effective vaccines for TB in order to achieve global control of the disease. Extracellular vesicles (EVs) are spherical membrane-bound structures released by pathogens and host cells. During the course of an infection, both pathogen- and host-derived EVs are produced and play important roles in determining the course of the infection. EVs offer intriguing tools as potential vaccines due to their ability to deliver multiple pathogen or host antigens.Hypothesis /Gap Statement. We hypothesized that EVs derived from M. tb and EVs from M. tb-infected macrophages may serve as potential vaccine candidates against M. tb infection.Aim. This study aims to compare the immunogenicity and immune protection between M. tb EVs and M. tb-infected macrophage-derived EVs.Methodology. In this study, EVs were extracted from culture supernatants of M. tb and M. tb-infected macrophages, respectively. Mass spectrometry was employed to explore the antigen composition of H37Rv-Mφ-EVs and H37Rv-EVs. Cytokine profiling and antibody response assays were used to analyse the immunogenicity offered by EVs. Additionally, we used histological examination to evaluate and protective efficacy of the EVs.Results. Our results demonstrated that mice immunized by EVs released from M. tb-infected macrophages induced stronger inflammatory cytokine response than M. tb. Moreover, EVs from M. tb-infected macrophages reinforced T-cell activation and antibody response compared to M. tb EVs. Proteomic analysis revealed that EVs from M. tb-infected macrophages containing immunodominant cargos have stronger binding ability with major histocompatibility complex molecules, which may contribute to the protection from M. tb infection. Indeed, immunization of EVs released from M. tb-infected macrophages significantly reduced the bacterial load and better protection against M. tb infection than EVs from M. tb. Importantly, the selected antigens (Ag85B, ESAT-6 and the Rv0580c) from EVs of M. tb-infected macrophages exhibited effective immunogenicity.Conclusion. Our results suggested that EVs derived from M. tb-infected macrophages might serve as a proper antigenic library for vaccine candidates against M. tb challenge.
Assuntos
Antígenos de Bactérias , Vesículas Extracelulares , Macrófagos , Mycobacterium tuberculosis , Vacinas contra a Tuberculose , Tuberculose , Vesículas Extracelulares/imunologia , Mycobacterium tuberculosis/imunologia , Animais , Antígenos de Bactérias/imunologia , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Camundongos , Macrófagos/imunologia , Macrófagos/microbiologia , Tuberculose/prevenção & controle , Tuberculose/imunologia , Tuberculose/microbiologia , Citocinas/metabolismo , FemininoRESUMO
Tuberculosis (TB), caused by Mycobacterium tuberculosis, remains the leading cause of mortality by a single infectious agent in the world. M. tuberculosis infection could also result in clinical chronic infection, known as latent TB infection (LTBI). Compared to the current limited treatment, several subunit vaccines showed immunotherapeutic effects and were included in clinical trials. In this study, a subunit vaccine of Ag85B with a novel mucosal adjuvant c-di-AMP (Ag85B:c-di-AMP) was delivered intranasally to a persistent M. tuberculosis H37Ra infection mouse model, which also presented the asymptomatic characteristics of LTBI. Compared with Ag85B immunization, Ag85B:c-di-AMP vaccination induced stronger humoral immune responses, significantly higher CD4+ T cells recruitment, enhanced Th1/Th2/Th17 profile response in the lung, decreased pathological lesions of the lung, and reduced M. tuberculosis load in mice. Taken together, Ag85B:c-di-AMP mucosal route immunization provided an immunotherapeutic effect on persistent M. tuberculosis H37Ra infection, and c-di-AMP, as a promising potential mucosal adjuvant, could be further used in therapeutic or prophylactic vaccine strategies for persistent M. tuberculosis infection as well as LTBI.
Assuntos
Adjuvantes Imunológicos , Modelos Animais de Doenças , Mycobacterium tuberculosis , Vacinas contra a Tuberculose , Animais , Adjuvantes Imunológicos/administração & dosagem , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Mycobacterium tuberculosis/imunologia , Camundongos , Feminino , Antígenos de Bactérias/imunologia , Aciltransferases/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Proteínas de Bactérias/imunologia , Tuberculose/imunologia , Tuberculose/prevenção & controle , Tuberculose Latente/imunologia , Camundongos Endogâmicos BALB C , Administração IntranasalRESUMO
Tuberculosis (TB) is one of the infectious diseases caused by the pathogen Mycobacterium tuberculosis that continuously threatens the global human health. Bacillus Calmette-Guérin (BCG) vaccine is the only vaccine that has been used clinically to prevent tuberculosis in recent centuries, but its limitations in preventing latent infection and reactivation of tuberculosis do not provide full protection. In this study, we selected the membrane-associated antigen Rv1513 of Mycobacterium. In order to achieve stable expression and function of the target gene, the prokaryotic expression recombinant vector pET30b-Rv1513 was constructed and expressed and purified its protein. Detection of IFN- γ levels in the peripheral blood of TB patients stimulated by whole blood interferon release assay (WBIA) and multi-microsphere flow immunofluorescence luminescence (MFCIA) revealed that the induced production of cytokines, such as IFN-γ and IL-6, was significantly higher than that in the healthy group. Rv1513 combined with adjuvant DMT (adjuvant system liposomes containing dimethyldioctadecylammonium bromide (DDA), monophospholipid A (MPL), and trehalose-660-dibenzoic acid (TDB)) was used to detect serum specific antibodies, cytokine secretion from splenic suprasplenic cell supernatants, and multifunctional T-cell levels in splenocytes in immunised mice. The levels of IFN-γ, TNF-α, and IL-2 secreted by mouse splenocytes were found in the Rv1513+DMT group and the BCG+Rv1513+DMT group. The serum levels of IgG and its subclasses and the number of IFN-γ+T cells, TNF-α+T and IFN-γ+TNF-α+T cells in the induced CD4+/CD8+T cells in mice were significantly higher than those in the BCG group, and the highest levels were found in the BCG+Rv1513+DMT group. These findings suggest that Rv1513/DMT may serve as a potential subunit vaccine candidate that may be effective as a booster vaccine after the first BCG vaccination.
Assuntos
Mycobacterium tuberculosis , Células Th1 , Vacinas contra a Tuberculose , Tuberculose , Animais , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/genética , Camundongos , Humanos , Células Th1/imunologia , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/genética , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose/imunologia , Tuberculose/prevenção & controle , Tuberculose/microbiologia , Feminino , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Citocinas/metabolismo , Citocinas/imunologia , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/genética , Interferon gama/imunologia , Interferon gama/metabolismo , Camundongos Endogâmicos BALB C , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Adjuvantes Imunológicos/administração & dosagem , AdultoRESUMO
The eradication of tuberculosis remains a global challenge. Despite being the only licensed vaccine, Bacillus Calmette-Guérin (BCG) confers limited protective efficacy in adults and individuals with latent tuberculosis infections (LTBI). There is an urgent need to develop novel vaccines that can enhance the protective effect of BCG. Protein subunit vaccines have garnered significant research interest due to their safety and plasticity. Based on previous studies, we selected three antigens associated with LTBI (Rv2028c, Rv2029c, Rv3126c) and fused them with an immunodominant antigen Ag85A, resulting in the construction of a multistage protein subunit vaccine named A986. We evaluated the protective effect of recombinant protein A986 adjuvanted with MPL/QS21 as a booster vaccine for BCG against Mycobacterium tuberculosis (Mtb) infection in mice. The A986 + MPL/QS21 induced the secretion of antigen-specific Th1 (IL-2+, IFN-γ+ and TNF-α+) and Th17 (IL-17A+) cytokines in CD4+ and CD8+ T cells within the lung and spleen of mice, while also increased the frequency of central memory and effector memory T cells. Additionally, it also induced the enhanced production of IgG antibodies. Compared to BCG alone, A986 + MPL/QS21 boosting significantly augmented the proliferation of antigen-specific multifunctional T cells and effectively reduced bacterial load in infected mice. Taken together, A986 + MPL/QS21 formulation induced robust antigen-specific immune responses and provided enhanced protection against Mtb infection as a booster of BCG vaccine.
Assuntos
Antígenos de Bactérias , Vacina BCG , Citocinas , Mycobacterium tuberculosis , Tuberculose , Vacinas de Subunidades Antigênicas , Animais , Vacinas de Subunidades Antigênicas/imunologia , Mycobacterium tuberculosis/imunologia , Vacina BCG/imunologia , Antígenos de Bactérias/imunologia , Feminino , Tuberculose/prevenção & controle , Tuberculose/imunologia , Camundongos , Citocinas/metabolismo , Imunização Secundária , Modelos Animais de Doenças , Adjuvantes Imunológicos/administração & dosagem , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos BALB C , Aciltransferases/imunologia , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/patologia , Proteínas de Bactérias/imunologia , HumanosRESUMO
BACKGROUND: A BCG booster vaccination administered via the respiratory mucosa may establish protective immune responses at the primary site of Mycobacterium tuberculosis infection. The primary objective of this trial was to compare the safety and immunogenicity of inhaled versus intramuscular administered ChAdOx1-85A. METHODS: We conducted a single-centre, randomised, double-blind, controlled phase 1 study (Swiss National Clinical Trials Portal number SNCTP000002920). After a dose-escalation vaccination in nine BCG-vaccinated healthy adults, a dose of 1 × 1010 vp of ChAdOx1-85A was administered to twenty BCG-vaccinated adults that were randomly allocated (1:1) into two groups: aerosol ChAdOx1-85A with intramuscular saline placebo or intramuscular ChAdOx1-85A with aerosol saline placebo, using block randomisation. A control group of ten BCG-naïve adults received aerosol ChAdOx1-85A at the same dose. Primary outcomes were solicited and unsolicited adverse events (AEs) up to day 16 post-vaccination and Serious AEs (SAEs) up to 24 weeks; secondary outcomes were cell-mediated and humoral immune responses in blood and bronchoalveolar lavage (BAL) samples. FINDINGS: Both vaccination routes were well tolerated with no SAEs. Intramuscular ChAdOx1-85A was associated with more local AEs (mostly pain at the injection site) than aerosol ChAdOx1-85A. Systemic AEs occurred in all groups, mainly fatigue and headaches, without differences between groups. Respiratory AEs were not different between BCG-vaccinated groups. Aerosol ChAdOx1-85A vaccination induced Ag85A BAL and systemic cellular immune responses with compartmentalisation of the immune responses: aerosol ChAdOx1-85A induced stronger BAL cellular responses, particularly IFNγ/IL17+CD4+ T cells; intramuscular ChAdOx1-85A induced stronger systemic cellular and humoral responses. INTERPRETATION: Inhaled ChAdOx1-85A was well-tolerated and induced lung mucosal and systemic Ag85A-specific T-cell responses. These data support further evaluation of aerosol ChAdOx1-85A and other viral vectors as a BCG-booster vaccination strategy.
Assuntos
Vacinas contra a Tuberculose , Humanos , Masculino , Injeções Intramusculares , Adulto , Feminino , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/efeitos adversos , Método Duplo-Cego , Administração por Inalação , Adulto Jovem , Aerossóis , ChAdOx1 nCoV-19/administração & dosagem , Vacinação/métodos , Mycobacterium tuberculosis/imunologia , Tuberculose/prevenção & controle , Tuberculose/imunologia , Pessoa de Meia-Idade , Imunização Secundária/métodos , Vacina BCG/administração & dosagem , Vacina BCG/imunologia , Vacina BCG/efeitos adversos , Imunidade Celular , Imunidade Humoral , Anticorpos Antibacterianos/sangue , Imunogenicidade da VacinaRESUMO
Selenium nanoparticles (SeNPs) enhance the immune response as adjuvants, increasing the efficacy of viral vaccines, including those for COVID-19. However, the efficiency of mucosal SeNPs in boosting vaccine-induced protective immunity against tuberculosis remains unclear. Therefore, this study aims to investigate whether the combination of SeNPs with the AH antigen (Ag85A-HspX) can boost respiratory mucosal immunity and thereby enhance the protective effects against tuberculosis. We synthesized SeNPs and assessed their impact on the immune response and protection against Mycobacterium bovis (M. bovis) as a mucosal adjuvant in mice, administered intranasally at a dose of 20 µg. SeNPs outperformed polyinosinic-polycytidylic acid (Poly IC) in stimulating the maturation of bone marrow-derived dendritic cells (BMDCs), which enhanced antigen presentation. SeNPs significantly activated and proliferated tissue-resident memory T cells (TRMs) and effector CD4+ T cells in the lungs. The vaccines elicited specific antibody responses in the respiratory tract and stimulated systemic Th1 and Th17 immune responses. Immunization with AH and SeNPs led to higher levels of mucosal secretory IgA in bronchoalveolar lavage fluid (BALF) and secretory IL-17 in splenocytes. Moreover, SeNPs immunized mice showed reduced M. bovis infection loads and inflammatory lesions in the lungs post-challenge. Notably, immunization with AH and SeNPs significantly reduced bacterial load in the lungs, achieving the lowest levels compared to all other tested groups. This study calls for pre-clinical investigation of AHB-SeNPs as an anti-bovine tuberculosis vaccine and for exploring its human vaccine potential, which is anticipated to aid in the development of innovative vaccines or adjuvants.
Assuntos
Adjuvantes Imunológicos , Antígenos de Bactérias , Imunidade nas Mucosas , Mycobacterium bovis , Nanopartículas , Selênio , Animais , Mycobacterium bovis/imunologia , Imunidade nas Mucosas/efeitos dos fármacos , Nanopartículas/administração & dosagem , Camundongos , Adjuvantes Imunológicos/administração & dosagem , Feminino , Antígenos de Bactérias/imunologia , Camundongos Endogâmicos C57BL , Tuberculose/imunologia , Tuberculose/prevenção & controle , Células Dendríticas/imunologia , Células Dendríticas/efeitos dos fármacos , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Pulmão/imunologia , Pulmão/microbiologia , Proteínas de Bactérias/imunologiaRESUMO
Identifying and evaluating potential vaccine candidates has become one of the main objectives to combat tuberculosis. Among them, mannosylated Apa antigen from Mycobacterium tuberculosis and the non-mannosylated protein expressed in Escherichia coli, have been studied. Although both proteins can induce a protective response in mice, it has been considered that native protein can be dispensed. In this work, we study the protective response induced by Apa expressed in E. coli and in Streptomyces lividans. The latter, like native is secreted as a double band of 45/47 kDa, however, only its 47 kDa band is mannosylated. Both antigens and BCG were intranasal administrated in mice, and animals were then challenged by aerosol with M. tuberculosis H37Rv. The results showed that both, Apa from S. lividans and E. coli conferred statistically significantly protection to animals compared to controls. The cytokine immune response was studied by an immunoassay after animals' immunization, revealing that Apa from S. lividans induced a statistically significant proliferation of T cell, as well as the expression of IFN-γ, IL-1ß, IL-17 and IL-10. In contrast, non-proliferation was obtained with non-mannosylated protein, but induction of IL-12 and IL-17 was observed. Together, these results demonstrate that both proteins were able to modulate a specific immune response against M. tuberculosis, that could be driven by different mechanisms possibly associated with the presence or not of mannosylation. Furthermore, stimulation of cells from BCG-vaccinated animals with the proteins could be an important tool, to help define the use of a given subunit-vaccine after BCG vaccination.
Assuntos
Administração Intranasal , Citocinas , Mycobacterium tuberculosis , Streptomyces lividans , Tuberculose , Animais , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/genética , Camundongos , Citocinas/metabolismo , Tuberculose/prevenção & controle , Tuberculose/imunologia , Streptomyces lividans/genética , Streptomyces lividans/imunologia , Aerossóis , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/administração & dosagem , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/administração & dosagem , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Feminino , Camundongos Endogâmicos BALB C , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/genética , Antígenos de Bactérias/administração & dosagemRESUMO
Tools to evaluate and accelerate tuberculosis (TB) vaccine development are needed to advance global TB control strategies. Validated human infection studies for TB have the potential to facilitate breakthroughs in understanding disease pathogenesis, identify correlates of protection, develop diagnostic tools, and accelerate and de-risk vaccine and drug development. However, key challenges remain for realizing the clinical utility of these models, which require further discussion and alignment among key stakeholders. In March 2023, the Wellcome Trust and the International AIDS Vaccine Initiative convened international experts involved in developing both TB and bacillus Calmette-Guérin (BCG) human infection studies (including mucosal and intradermal challenge routes) to discuss the status of each of the models and the key enablers to move the field forward. This report provides a summary of the presentations and discussion from the meeting. Discussions identified key issues, including demonstrating model validity, to provide confidence for vaccine developers, which may be addressed through demonstration of known vaccine effects (eg, BCG vaccination in specific populations), and by comparing results from field efficacy and human infection studies. The workshop underscored the importance of establishing safe and acceptable studies in high-burden settings, and the need to validate >1 model to allow for different scientific questions to be addressed as well as to provide confidence to vaccine developers and regulators around use of human infection study data in vaccine development and licensure pathways.
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Vacinas contra a Tuberculose , Tuberculose , Humanos , Tuberculose/prevenção & controle , Tuberculose/imunologia , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Desenvolvimento de Vacinas , Vacina BCG/imunologia , Vacina BCG/administração & dosagem , Mycobacterium tuberculosis/imunologia , AnimaisRESUMO
Introduction: Mycobacteria are known to exert a range of heterologous effects on the immune system. The mycobacteria-based Freund's Complete Adjuvant is a potent non-specific stimulator of the immune response used in immunization protocols promoting antibody production, and Mycobacterium bovis Bacille Calmette Guérin (BCG) vaccination has been linked with decreased morbidity and mortality beyond the specific protection it provides against tuberculosis (TB) in some populations and age groups. The role of heterologous antibodies in this phenomenon, if any, remains unclear and under-studied. Methods: We set out to evaluate antibody responses to a range of unrelated pathogens following infection with Mycobacterium tuberculosis (M.tb) and vaccination with BCG or a candidate TB vaccine, MTBVAC, in non-human primates. Results: We demonstrate a significant increase in the titer of antibodies against SARS-CoV-2, cytomegalovirus, Epstein-Barr virus, tetanus toxoid, and respiratory syncytial virus antigens following low-dose aerosol infection with M.tb. The magnitude of some of these responses correlated with TB disease severity. However, vaccination with BCG administered by the intradermal, intravenous or aerosol routes, or intradermal delivery of MTBVAC, did not increase antibody responses against unrelated pathogens. Discussion: Our findings suggest that it is unlikely that heterologous antibodies contribute to the non-specific effects of these vaccines. The apparent dysregulation of B cell responses associated with TB disease warrants further investigation, with potential implications for risk of B cell cancers and novel therapeutic strategies.
Assuntos
Vacina BCG , Mycobacterium tuberculosis , Tuberculose , Vacinação , Animais , Vacina BCG/imunologia , Vacina BCG/administração & dosagem , Tuberculose/imunologia , Tuberculose/prevenção & controle , Mycobacterium tuberculosis/imunologia , Anticorpos Antibacterianos/imunologia , Anticorpos Antibacterianos/sangue , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/sangue , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Feminino , Macaca mulatta , SARS-CoV-2/imunologia , COVID-19/imunologia , COVID-19/prevenção & controle , Imunidade Heteróloga , MasculinoRESUMO
Bacille Calmette-Guérin (BCG) remains the sole licensed vaccine against tuberculosis (TB), despite its variable efficacy in protecting against pulmonary TB. The development of effective TB vaccines faces significant challenges, marked by the absence of validated correlates of protection and predictive animal models. Strategic approaches to enhance TB vaccines and augment BCG efficacy include utilising prime-boost strategies with viral-vectored vaccines and exploring innovative delivery techniques, such as mucosal vaccine administration. Viral vectors offer numerous advantages, including the capacity to accommodate genes encoding extensive antigenic fragments and the induction of robust immune responses. Aerosol delivery aligns with the route of Mycobacterium tuberculosis infection and holds the potential to enhance protective mucosal immunity. Aerosolised viral-vectored vaccines overcome anti-vector immunity, facilitating repeated aerosol deliveries.
Assuntos
Aerossóis , Vetores Genéticos , Mycobacterium tuberculosis , Vacinas contra a Tuberculose , Tuberculose , Humanos , Animais , Vacinas contra a Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/genética , Vetores Genéticos/imunologia , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/genética , Tuberculose/prevenção & controle , Tuberculose/imunologia , Administração por Inalação , Vacina BCG/imunologia , Vacina BCG/administração & dosagem , Vacina BCG/genética , Vacinação/métodos , Tuberculose Pulmonar/prevenção & controle , Tuberculose Pulmonar/imunologiaRESUMO
Given the encouraging clinical results of both candidate subunit vaccines and revaccination with Bacillus Calmette-Guérin (BCG) against tuberculosis (TB), there is support for combining BCG and subunit vaccination for increased efficacy. BCG and Mycobacterium tuberculosis (Mtb) share ~98% of their genome and current subunit vaccines are almost exclusively designed as BCG boosters. The goal of this study is to design a TB subunit vaccine composed of antigens not shared with BCG and explore the advantages of this design in a BCG + subunit co-administration vaccine strategy. Eight protective antigens are selected to create an Mtb-specific subunit vaccine, named H107. Whereas traditional vaccines containing BCG-shared antigens exhibit in vivo cross-reactivity to BCG, H107 shows no cross-reactivity and does not inhibit BCG colonization. Instead, co-administering H107 with BCG leads to increased adaptive responses against both H107 and BCG. Importantly, rather than expanding BCG-primed T cells, H107 broadens the overall vaccine repertoire with new T cell clones and introduces 'adjuvant-imprinted' qualities including Th17 responses and less-differentiated Th1 cells. Collectively, these features of H107 are associated with a substantial increase in long-term protection.
Assuntos
Mycobacterium tuberculosis/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose/prevenção & controle , Vacinação/métodos , Animais , Antígenos de Bactérias/imunologia , Vacina BCG/administração & dosagem , Vacina BCG/imunologia , Humanos , Imunogenicidade da Vacina , Camundongos , Mycobacterium bovis/imunologia , Células Th1/imunologia , Células Th17/imunologia , Tuberculose/imunologia , Vacinas contra a Tuberculose/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
Interleukin (IL)-17A-producing T helper (Th)17 cells are increasingly being acknowledged to be associated with protective immunity to Mycobacterium tuberculosis (Mtb). Subunit vaccines potently promote protective immune responses against Mtb infection that correlate with an expansion of IL-23-dependent Th17 cells. Previous studies revealed that after vaccination, IL-23 is required for protection against challenge with Mtb but the underlying IL-23-dependent-and possibly IL-17A-mediated-mechanisms remain elusive. Therefore, we here analyzed the early outcome of Mtb infection in C57BL/6, IL-23p19-deficient (-/-), and IL-17A-/- mice after vaccination with the subunit vaccine H1-DDA/TDB to investigate the role of the IL-23-Th17 immune axis for the instruction of vaccine-induced protection. While in IL-23p19-/- mice the protective effect was reduced, protection after vaccination was maintained in IL-17A-/- animals for the course of infection of 6 weeks, indicating that after vaccination with H1-DDA/TDB early protection against Mtb is-although dependent on IL-23-not mediated by IL-17A. In contrast, IL-17A deficiency appears to have an impact on maintaining long-term protection. In fact, IL-23 instructed the vaccine-induced memory immunity in the lung, in particular the sustained expansion of tumor necrosis factor (TNF)+IL-2+ multifunctional T cells, independently of IL-17A. Altogether, a targeted induction of IL-23 during vaccination against Mtb might improve the magnitude and quality of vaccine-induced memory immune responses. KEY MESSAGES: After subunit Mtb vaccination with H1-DDA/TDB, IL-23 but not IL-17A contributes to vaccine-induced early protection against infection with Mtb. IL-17F does not compensate for IL-17A deficiency in terms of H1-DDA/TDB-induced protection against Mtb infection. IL 23 promotes the H1-DDA/TDB-induced accumulation of effector memory T cells independently of IL 17A. IL-23 arbitrates the induction of H1-specific IFN-γ-TNF+IL-2+ double-positive multifunctional CD4 T cells after subunit Mtb vaccination in an IL-17A-independent manner.
Assuntos
Antígenos de Bactérias/administração & dosagem , Linfócitos T CD4-Positivos/efeitos dos fármacos , Interleucina-23/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas de Subunidades Antigênicas/administração & dosagem , Animais , Linfócitos T CD4-Positivos/imunologia , Feminino , Interleucina-17/genética , Interleucina-17/imunologia , Interleucina-23/genética , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium tuberculosis/imunologia , Tuberculose/prevenção & controleRESUMO
OBJECTIVES: To evaluate the immunogenicity of Mycobacterium intracellulare proteins and determine the cross-reactive proteins between M. intracellulare and M. tuberculosis. METHODS: Protein extracts from M. intracellulare were used to immunize BALB/c mice. The antigens were evaluated using cellular and humoral immunoassays. The common genes between M. intracellular and M. tuberculosis were identified using genome-wide comparative analysis, and cross-reactive proteins were screened using immunoproteome microarrays. RESULTS: Immunization with M. intracellulare proteins induced significantly higher levels of the cytokines interferon-γ (IFN-γ), interleukin-2 (IL-2), interleukin-12 (IL-12), interleukin-6 (IL-6) and immunoglobulins IgG, IgG1, IgM, and IgG2a in mouse serum. Bone marrow-derived macrophages isolated from mice immunized with M. intracellulare antigens displayed significantly lower bacillary loads than those isolated from mice immunized with adjuvants. Whole-genome sequence analysis revealed 396 common genes between M. intracellulare and M. tuberculosis. Microchip hybridization with M. tuberculosis proteins revealed the presence of 478 proteins in the serum of mice immunized with M. intracellulare protein extracts. Sixty common antigens were found using both microchip and genomic comparative analyses. CONCLUSION: This is the advanced study to investigate the immunogenicity of M. intracellulare proteins and the cross-reactive proteins between M. intracellulare and M. tuberculosis. The results revealed the presence of a number of cross-reactive proteins between M. intracellulare and M. tuberculosis. Therefore, this study provides a new way of identifying immunogenic proteins for use in tuberculosis vaccines against both M. intracellulare and M. tuberculosis in future.
Assuntos
Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Complexo Mycobacterium avium/imunologia , Mycobacterium tuberculosis/imunologia , Animais , Reações Cruzadas , Citocinas/imunologia , Feminino , Genoma Bacteriano , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Macrófagos/imunologia , Camundongos Endogâmicos BALB C , Complexo Mycobacterium avium/genética , Mycobacterium tuberculosis/genética , Vacinas contra a Tuberculose/administração & dosagem , Sequenciamento Completo do GenomaRESUMO
Background: Tuberculosis (TB) is still a global infectious disease that seriously threatens human beings. The only licensed TB vaccine Bacille Calmette-Guérin (BCG)'s protective efficacy varies significantly among populations and regions. It is very urgent to develop more effective vaccines. Methods: In this study, eleven candidate proteins of Mycobacterium tuberculosis were selected to predict peptides with high-affinity binding capacity for the HLA-DRB1*01:01 molecule. The immunodominant peptides were identified with the enzyme-linked immunospot assay (ELISPOT) and linked in silico to result in a novel polypeptide vaccine in Escherichia coli cells. The vaccine's protective efficacy was evaluated in humanized and wild-type C57BL/6 mice. The potential immune protective mechanisms were explored with Enzyme-linked Immunosorbent Assay (ELISA), flow cytometry, and ELISPOT. Results: Six immunodominant peptides screened from 50 predicted peptides were used to construct a new polypeptide vaccine named MP3RT. After challenge with M. tuberculosis, the colony-forming units (CFUs), lung lesion area, and the number of inflammatory cells in humanized mice rather than wild-type mice vaccinated with MP3RT were significantly lower than these in mice immunized with PBS. The humanized mice vaccinated with MP3RT revealed significant increases in IFN-γ cytokine production, IFN-γ+ T lymphocytes, CD3+IFN-γ+ T lymphocytes, and the MP3RT-specific IgG antibody. Conclusions: Taken together, MP3RT is a promising peptides-based TB vaccine characterized by inducing high levels of IFN-γ and CD3+IFN-γ+ T lymphocytes in humanized mice. These new findings will lay a foundation for the development of peptides-based vaccines against TB.
Assuntos
Mycobacterium tuberculosis/imunologia , Peptídeos/imunologia , Vacinas contra a Tuberculose/imunologia , Tuberculose/prevenção & controle , Animais , Anticorpos Antibacterianos/imunologia , Antígenos de Bactérias/química , Antígenos de Bactérias/imunologia , Modelos Animais de Doenças , Humanos , Epitopos Imunodominantes/química , Epitopos Imunodominantes/genética , Epitopos Imunodominantes/imunologia , Interferon gama/imunologia , Linfócitos/imunologia , Camundongos , Camundongos Transgênicos , Peptídeos/administração & dosagem , Peptídeos/química , Peptídeos/genética , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/química , Vacinas contra a Tuberculose/genética , Vacinação , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/química , Vacinas de Subunidades Antigênicas/genética , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
Spray drying is a technique that can be used to stabilize biopharmaceuticals, such as vaccines, within dry particles. Compared to liquid pharmaceutical products, dry powder has the potential to reduce costs associated with refrigerated storage and transportation. In this study, spray drying was investigated for processing an adjuvanted tuberculosis subunit vaccine, formulated as an oil-in-water nanoemulsion, into a dry powder composed of microparticles. Applying in-silico approaches to the development of formulation and processing conditions, successful encapsulation of the adjuvanted vaccine within amorphous microparticles was achieved in only one iteration, with high retention (>90%) of both the antigen and adjuvant system. Moisture-controlled stability studies on the powder were conducted over 26 months at temperatures up to 40 °C. Results showed that the powder was physically stable after 26 months of storage for all tested temperatures. Adjuvant system integrity was maintained at temperatures up to 25 °C after 26 months and after one month of storage at 40 °C. The spray-dried product demonstrated improved antigen thermostability when stored above refrigerated temperatures as compared to the liquid product. These results demonstrate the feasibility of spray drying as a method of encapsulating and stabilizing an adjuvanted vaccine.
Assuntos
Adjuvantes Imunológicos/química , Composição de Medicamentos/métodos , Secagem por Atomização , Vacinas contra a Tuberculose/química , Tuberculose/prevenção & controle , Adjuvantes Imunológicos/administração & dosagem , Química Farmacêutica , Estabilidade de Medicamentos , Armazenamento de Medicamentos , Emulsões , Excipientes , Humanos , Nanopartículas/química , Tamanho da Partícula , Pós , Vacinas contra a Tuberculose/administração & dosagem , Vacinas de Subunidades Antigênicas/administração & dosagemRESUMO
BCG vaccination can strengthen protection against pathogens through the induction of epigenetic and metabolic reprogramming of innate immune cells, a process called trained immunity. We and others recently demonstrated that mucosal or intravenous BCG better protects rhesus macaques from Mycobacterium tuberculosis infection and TB disease than standard intradermal vaccination, correlating with local adaptive immune signatures. In line with prior mouse data, here, we show in rhesus macaques that intravenous BCG enhances innate cytokine production associated with changes in H3K27 acetylation typical of trained immunity. Alternative delivery of BCG does not alter the cytokine production of unfractionated bronchial lavage cells. However, mucosal but not intradermal vaccination, either with BCG or the M. tuberculosis-derived candidate MTBVAC, enhances innate cytokine production by blood- and bone marrow-derived monocytes associated with metabolic rewiring, typical of trained immunity. These results provide support to strategies for improving TB vaccination and, more broadly, modulating innate immunity via mucosal surfaces.
Assuntos
Vacina BCG/administração & dosagem , Imunidade nas Mucosas , Mycobacterium tuberculosis/imunologia , Mucosa Respiratória/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose Pulmonar/prevenção & controle , Acetilação , Administração Intranasal , Animais , Medula Óssea/efeitos dos fármacos , Medula Óssea/imunologia , Medula Óssea/microbiologia , Reprogramação Celular/genética , Reprogramação Celular/imunologia , Feminino , Regulação da Expressão Gênica , Histonas/genética , Histonas/imunologia , Injeções Intravenosas , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/microbiologia , Macaca mulatta , Masculino , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/microbiologia , Mycobacterium tuberculosis/patogenicidade , Mucosa Respiratória/microbiologia , Tuberculose Pulmonar/genética , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/microbiologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
To fight tuberculosis, better vaccination strategies are needed. Live attenuated Mycobacterium tuberculosis-derived vaccine, MTBVAC, is a promising candidate in the pipeline, proven to be safe and immunogenic in humans so far. Independent studies have shown that pulmonary mucosal delivery of Bacillus Calmette-Guérin (BCG), the only tuberculosis (TB) vaccine available today, confers superior protection over standard intradermal immunization. Here we demonstrate that mucosal MTBVAC is well tolerated, eliciting polyfunctional T helper type 17 cells, interleukin-10, and immunoglobulins in the airway and yielding a broader antigenic profile than BCG in rhesus macaques. Beyond our previous work, we show that local immunoglobulins, induced by MTBVAC and BCG, bind to M. tuberculosis and enhance pathogen uptake. Furthermore, after pulmonary vaccination, but not M. tuberculosis infection, local T cells expressed high levels of mucosal homing and tissue residency markers. Our data show that pulmonary MTBVAC administration has the potential to enhance its efficacy and justifies further exploration of mucosal vaccination strategies in preclinical efficacy studies.
Assuntos
Vacina BCG/administração & dosagem , Imunidade nas Mucosas , Mycobacterium tuberculosis/imunologia , Mucosa Respiratória/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose Pulmonar/prevenção & controle , Administração Intranasal , Animais , Reprogramação Celular/genética , Reprogramação Celular/imunologia , Feminino , Regulação da Expressão Gênica , Injeções Intradérmicas , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-17/genética , Interleucina-17/imunologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/microbiologia , Macaca mulatta , Masculino , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/microbiologia , Mycobacterium tuberculosis/patogenicidade , Mucosa Respiratória/microbiologia , Células Th1/imunologia , Células Th1/microbiologia , Células Th17/imunologia , Células Th17/microbiologia , Tuberculose Pulmonar/genética , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/microbiologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Tuberculosis remains a serious global health problem. BCG is the only prophylactic TB vaccine and it shows variable protective efficacy. Chimeric protein subunit vaccines hold great potential as stand-alone vaccines or heterologous BCG prime boosters. We have designed a protein chimera, PP31, by combining Mtb ESAT-6 family antigen Rv1198 and MoCo biosynthesis family antigen Rv3111. Further, PP31 was extended by addition of latency antigen Rv1813c to yield PP43. Immunization of BALB/c mice with PP31 or PP43 with FIA adjuvant elicited strong humoral immune response. Restimulation of splenocytes of the immunized mice lead to significant proliferation of lymphocytes, secretion of cytokines IFN-γ, TNF, IL-2 of the Th1 class, IL-17A of the Th17 class, and IL-6. PP31 and PP43 also induced intracellular cytokine expression (IFN-γ, TNF, and IL-2) from both CD4+-CD44high and CD8+-CD44high T-cells. Antigen-specific IFN-γ+/IL-2+ double positive CD4+ T-cells were significantly higher in case of PP43 than PP31-immunized mice and control group. PP43 showed protection equivalent to heat-inactivated BCG in response to challenge of the immunized mice with Mtb H37Ra. Based on its immunogenicity and protective efficacy, PP43 appears to be a potential candidate for further development as a subunit vaccine against TB.
Assuntos
Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Epitopos , Imunogenicidade da Vacina , Mycobacterium tuberculosis/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose/prevenção & controle , Adjuvantes Imunológicos/administração & dosagem , Animais , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/administração & dosagem , Proteínas de Bactérias/administração & dosagem , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Feminino , Humanos , Imunidade Celular/efeitos dos fármacos , Imunidade Humoral/efeitos dos fármacos , Imunização , Ativação Linfocitária/efeitos dos fármacos , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/imunologia , Subpopulações de Linfócitos/metabolismo , Camundongos Endogâmicos BALB C , Mycobacterium tuberculosis/genética , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/imunologia , Tuberculose/sangue , Tuberculose/imunologia , Tuberculose/microbiologia , Vacinas contra a Tuberculose/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
BACKGROUND: Substantial recent advances in the comprehension of the molecular and cellular mechanisms behind asthma have evidenced the importance of the lung immune environment for disease outcome, making modulation of local immune responses an attractive therapeutic target against this pathology. Live attenuated mycobacteria, such as the tuberculosis vaccine BCG, have been classically linked with a type 1 response, and proposed as possible modulators of the type 2 response usually associated with asthma. METHODS: In this study we used different acute and chronic murine models of asthma to investigate the therapeutic efficacy of intranasal delivery of the live tuberculosis vaccines BCG and MTBVAC by regulating the lung immune environment associated with airway hyperresponsiveness (AHR). FINDINGS: Intranasal administration of BCG, or the novel tuberculosis vaccine candidate MTBVAC, abrogated AHR-associated hallmarks, including eosinophilia and lung remodeling. This correlated with the re-polarization of allergen-induced M2 macrophages towards an M1 phenotype, as well as with the induction of a strong allergen-specific Th1 response. Importantly, vaccine treatment was effective in a scenario of established chronic asthma where a strong eosinophil infiltration was already present prior to immunization. We finally compared the nebulization efficiency of clinical formulations of MTBVAC and BCG using a standard commercial nebulizer for potential aerosol application. INTERPRETATION: Our results demonstrate that pulmonary live tuberculosis vaccines efficiently revert established asthma in mice. These data support the further exploration of this approach as potential therapy against asthma. FUNDING: Spanish Ministry of Science [grant numbers: BIO2014-5258P, RTI2018-097625-B-I00], Instituto de Salud Carlos III, Gobierno de Aragón/Fondo Social Europeo, University of Zaragoza [grant number: JIUZ-2018-BIO-01].
Assuntos
Asma/imunologia , Asma/terapia , Vacinas contra a Tuberculose/uso terapêutico , Vacinas Atenuadas/uso terapêutico , Administração Intranasal , Remodelação das Vias Aéreas/imunologia , Alérgenos/imunologia , Animais , Vacina BCG , Biomarcadores , Microambiente Celular/imunologia , Citocinas/metabolismo , Modelos Animais de Doenças , Eosinófilos/imunologia , Eosinófilos/metabolismo , Eosinófilos/patologia , Feminino , Imunização , Camundongos , Ovalbumina/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas Atenuadas/administração & dosagemRESUMO
There have been notable advances in the development of vaccines against active tuberculosis (TB) disease for adults and adolescents. Using mathematical models, we seek to estimate the potential impact of a post-exposure TB vaccine, having 50% efficacy in reducing active disease, on global rifampicin-resistant (RR-) TB burden. In 30 countries that together accounted for 90% of global RR-TB incidence in 2018, a future TB vaccine could avert 10% (95% credible interval: 9.7-11%) of RR-TB cases and 7.3% (6.6-8.1%) of deaths over 2020-2035, with India, China, Indonesia, Pakistan, and the Russian Federation having the greatest contribution. This impact would increase to 14% (12-16%) and 31% (29-33%) respectively, when combined with improvements in RR-TB diagnosis and treatment relative to a scenario of no vaccine and no such improvements. A future TB vaccine could have important implications for the global control of RR-TB, especially if implemented alongside enhancements in management of drug resistance.