ABSTRACT
BACKGROUND: This study assessed the safety and immunogenicity of a new booster vaccine against tetanus, diphtheria, and pertussis manufactured by Serum Institute of India Pvt. Ltd (SIIPL Tdap). METHODS: The Phase II/III trial was randomized (2:1), observer blinded and active controlled. Healthy subjects aged 4-65 years received a single dose of either SIIPL Tdap or comparator Tdap vaccine (Boostrix®, GlaxoSmithKline, Belgium), and were followed-up for 30 days. Blood samples for safety and immunogenicity assessments were collected pre-vaccination and on day 30 post-vaccination. The study assessed safety and reactogenicity of SIIPL Tdap compared to the comparator Tdap as well as the co-primary immunogenicity outcomes: (i) seroprotection rates against diphtheria toxoid (DT) and tetanus toxoid (TT) and (ii) the booster response rates against pertussis toxoid (PT), filamentous hemagglutinin (FHA) and pertactin (PRN) 30 days post-vaccination in all study subjects. A margin of -10 % was used for non-inferiority testing. Secondary outcomes included the booster response rates against DT and TT, seropositivity rates against pertussis antigens, and antibody geometric mean concentrations (GMCs) for all vaccine components. RESULTS: At Day 30 post-vaccination, SIIPL Tdap was assessed as non-inferior to the comparator Tdap in terms of: i) seroprotection rates against DT (94.4 % vs. 94.9 %) and TT (99.9 % vs. 100 %) and ii) pertussis booster response rates (93.8 % vs. 88.4 % anti-PT, 89.7 % vs. 90.9 % anti-FHA and 86.3 % vs. 84.4 % anti-PRN), for SIIPL Tdap versus comparator Tdap, respectively. GMCs for anti-PT and anti-PRN were higher in subjects vaccinated with SIIPL Tdap compared to comparator Tdap. All other secondary outcomes were comparable. The overall frequency of local and systemic solicited AEs was comparable; no treatment related SAEs were reported. CONCLUSIONS: Booster vaccination with SIIPL Tdap was non-inferior to comparator Tdap with respect to the immunogenicity of the vaccine components and was equally well tolerated. EudraCT number: 2019-002706-46.
ABSTRACT
A more effective vaccine against tuberculosis than Bacille Calmette-Guérin (BCG) is urgently needed. BCG derived recombinant VPM1002 has been found to be more efficacious and safer than the parental strain in mice models. Newer candidates, such as VPM1002 Δpdx1 (PDX) and VPM1002 ΔnuoG (NUOG), were generated to further improve the safety profile or efficacy of the vaccine. Herein, we assessed the safety and immunogenicity of VPM1002 and its derivatives, PDX and NUOG, in juvenile goats. Vaccination did not affect the goats' health in regards to clinical/hematological features. However, all three tested vaccine candidates and BCG induced granulomas at the site of injection, with some of the nodules developing ulcerations approximately one month post-vaccination. Viable vaccine strains were cultured from the injection site wounds in a few NUOG- and PDX- vaccinated animals. At necropsy (127 days post-vaccination), BCG, VPM1002, and NUOG, but not PDX, still persisted at the injection granulomas. All strains, apart from NUOG, induced granuloma formation only in the lymph nodes draining the injection site. In one animal, the administered BCG strain was recovered from the mediastinal lymph nodes. Interferon gamma (IFN-γ) release assay showed that VPM1002 and NUOG induced a strong antigen-specific response comparable to that elicited by BCG, while the response to PDX was delayed. Flow cytometry analysis of IFN-γ production by CD4+, CD8+, and γδ T cells showed that CD4+ T cells of VPM1002- and NUOG-vaccinated goats produced more IFN-γ compared to BCG-vaccinated and mock-treated animals. In summary, the subcutaneous application of VPM1002 and NUOG induced anti-tuberculous immunity, while exhibiting a comparable safety profile to BCG in goats.
Subject(s)
BCG Vaccine , Tuberculosis , Animals , Mice , Goats , Tuberculosis/prevention & control , T-Lymphocytes , Vaccination/adverse effectsABSTRACT
BACKGROUND: Bacille Calmette-Guérin (BCG) vaccination can potentially reduce the rate of respiratory infections in vulnerable populations. This study evaluates the safety and efficacy of VPM1002 (a genetically modified BCG) as prophylaxis against severe respiratory tract infections including coronavirus disease 2019 (COVID-19) in an elderly population. METHODS: In this phase 3, randomized, double-blind, placebo-controlled, multicenter clinical trial, healthy elderly volunteers (N = 2064) were enrolled, randomized (1:1) to receive either VPM1002 or placebo, and followed up remotely for 240 days. The primary outcome was the mean number of days with severe respiratory infections at hospital and/or at home. Secondary endpoints included the incidence of self-reported fever, number of hospital and intensive care unit (ICU) admissions, and number of adverse events. RESULTS: A total of 31 participants in the VPM1002 group reported at least 1 day with severe respiratory disease and a mean number of days with severe respiratory disease of 9.39 ± 9.28 while in the placebo group; 38 participants reported a mean of 14.29 ± 16.25 days with severe respiratory disease. The incidence of self-reported fever was lower in the VPM1002 group (odds ratio, 0.46 [95% confidence interval, .28-.74]; P = .001), and consistent trends to fewer hospitalization and ICU admissions due to COVID-19 were observed after VPM1002 vaccination. Local reactions typical for BCG were observed in the VPM1002-vaccinated group, which were mostly of mild intensity. CONCLUSIONS: Vaccination with VPM1002 is well tolerated and seems to have a prophylactic effect against severe respiratory disease in the elderly. CLINICAL TRIALS REGISTRATION: NCT04435379.
Subject(s)
BCG Vaccine , COVID-19 , Aged , Humans , BCG Vaccine/immunology , BCG Vaccine/standards , COVID-19/prevention & control , Double-Blind Method , Hospitalization/statistics & numerical data , SARS-CoV-2 , Respiratory Tract Diseases/prevention & control , Middle Aged , Aged, 80 and over , Male , Female , Time Factors , Patient AcuityABSTRACT
Tuberculous granulomas are highly dynamic structures reflecting the complex host-mycobacterium interactions. The objective of this study was to compare granuloma development at the site of vaccination with BCG and its recombinant derivatives in goats. To characterize the host response, epithelioid cells, multinucleated giant cells (MNGC), T cell subsets, B cells, plasma cells, dendritic cells and mycobacterial antigen were labelled by immunohistochemistry, and lipids and acid-fast bacteria (AFB) were labelled by specific staining. Granulomas with central caseous necrosis developed at the injection site of most goats though lesion size and extent of necrosis differed between vaccine strains. CD4+ T and B cells were more scarce and CD8+ cells were more numerous in granulomas induced by recombinant derivatives compared to their parental BCG strain. Further, the numbers of MNGCs and cells with lipid bodies were markedly lower in groups administered with recombinant BCG strains. Microscopic detection of AFB and mycobacterial antigen was rather frequent in the area of central necrosis, however, the isolation of bacteria in culture was rarely successful. In summary, BCG and its recombinant derivatives induced reproducibly subcutaneous caseous granulomas in goats that can be easily monitored and surgically removed for further studies. The granulomas reflected the genetic modifications of the recombinant BCG-derivatives and are therefore suitable models to compare reactions to different mycobacteria or TB vaccines.
Subject(s)
BCG Vaccine , Mycobacterium , Tuberculosis , Animals , BCG Vaccine/adverse effects , Goats , Granuloma/etiology , Lipids , Mycobacterium/genetics , NecrosisABSTRACT
BACKGROUND: Tuberculosis is a major public health problem worldwide. Immunisation with Mycobacterium bovis BCG vaccine is partially effective in infants, reducing the incidence of miliary and tuberculosis meningitis, but is less effective against pulmonary tuberculosis. We aimed to compare safety and immunogenicity of VPM1002-a recombinant BCG vaccine developed to address this gap-with BCG in HIV exposed and HIV unexposed newborn babies. METHODS: This double-blind, randomised, active controlled phase 2 study was conducted at four health centres in South Africa. Eligible neonates were aged 12 days or younger with a birthweight of 2·5-4·2 kg, and could be HIV exposed (seropositive mothers) or unexposed (seronegative mothers). Newborn babies were excluded if they had acute or chronic illness, fever, hypothermia, sepsis, cancer, or congenital malformation, or if they received blood products or immunosuppressive therapy. Participants were excluded if their mothers (aged ≥18 years) had active tuberculosis disease, diabetes, a history of immunodeficiency except for HIV, hepatitis B or syphilis seropositivity, received blood products in the preceding 6 months, any acute infectious disease, or any suspected substance abuse. Participants were randomly assigned to VPM1002 or BCG vaccination in a 3:1 ratio, stratified by HIV status using the random number generator function in SAS, using a block size of eight paticipants. The primary outcome was non-inferiority (margin 15%) of VPM1002 to BCG vaccine in terms of incidence of grade 3-4 adverse drug reactions or ipsilateral or generalised lymphadenopathy of 10 mm or greater in diameter by 12 months. The primary outcome was assessed in all vaccinated participants (safety population) at regular follow-up visits until 12 months after vaccination. Secondary immunogenicity outcomes were interferon-γ levels and percentages of multifunctional CD4+ and CD8+ T cells among all lymphocytes across the 12 month study period. The study was registered with ClinicalTrials.gov, NCT02391415. FINDINGS: Between June 4, 2015 and Oct 16, 2017, 416 eligible newborn babies were randomly assigned and received study vaccine. Seven (2%) of 312 participants in the VPM1002 group had a grade 3-4 vaccine-related adverse reaction or lymphadenopathy of 10 mm or greater in diameter compared with 34 (33%) of 104 participants in the BCG group (risk difference -30·45% [95% CI -39·61% to -21·28%]; pnon-inferiority<0·0001); VPM1002 was thus non-inferior to BCG for the primary outcome. Incidence of severe injection site reactions was lower with VPM1002 than BCG: scarring occurred in 65 (21%) participants in the VPM1002 group versus 77 (74%) participants in the BCG group (p<0·0001); ulceration occurred in one (<1%) versus 15 (14%; p<0·0001); and abscess formation occurred in five (2%) versus 23 (22%; p<0·0001). Restimulated IFNγ concentrations were lower in the VPM1002 group than the BCG group at week 6, week 12, month 6, and month 12. The percentage of multifunctional CD4+ T cells was higher in the VPM1002 group than the BCG group at day 14 but lower at week 6, week 12, month 6, and month 12. The percentage of multifunctional CD8+ T cells was lower in the VPM1002 group than the BCG group at week 6, week 12, and month 6, but did not differ at other timepoints. INTERPRETATION: VPM1002 was less reactogenic than BCG and was not associated with any serious safety concern. Both vaccines were immunogenic, although responses were higher with the BCG vaccine. VPM1002 is currently being studied for efficacy and safety in a multicentric phase 3 clinical trial in babies in sub-Saharan Africa. FUNDING: Serum Institute of India.
Subject(s)
HIV Infections , Lymphadenopathy , Tuberculosis , Adolescent , Adult , BCG Vaccine , CD8-Positive T-Lymphocytes , Double-Blind Method , HIV Infections/drug therapy , Humans , Immunogenicity, Vaccine , Infant , Infant, Newborn , Interferon-gamma , South Africa , Tuberculosis/drug therapyABSTRACT
Bovine tuberculosis (bTB) not only poses a zoonotic threat to humans but also has a significant economic impact on livestock production in many areas of the world. Effective vaccines for humans, livestock, and wildlife are highly desirable to control tuberculosis. Suitable large animal models are indispensable for meaningful assessment of vaccine candidates. Here, we describe the refinement of an animal model for bTB in goats. Intrabronchial inoculation procedure via video-guided endoscopy in anesthetized animals, collection of lungs after intratracheal fixation in situ, and imaging of lungs by computed tomography (CT) were established in three goats using barium sulfate as surrogate inoculum. For subsequent infection experiments, four goats were infected with 4.7 × 102 colony-forming units of M. bovis by intrabronchial inoculation using video-guided endoscopy with spray catheters. Defined amounts of inoculum were deposited at five sites per lung. Four age-matched goats were mock-inoculated. None of the goats developed clinical signs until they were euthanized 5 months post infection, but simultaneous skin testing confirmed bTB infection in all goats inoculated with M. bovis. In tissues collected at necropsy, M. bovis was consistently re-isolated from granulomas in lymph nodes, draining the lungs of all the goats infected with M. bovis. Further dissemination was observed in one goat only. Pulmonary lesions were quantified by CT and digital 2D radiography (DR). CT revealed mineralized lesions in all the infected goats ranging from <5 mm to >10 mm in diameter. Small lesions <5 mm predominated. The DR failed to detect small lesions and to determine the exact location of lesions because of overlapping of pulmonary lobes. Relative volume of pulmonary lesions was low in three but high in one goat that also had extensive cavitation. CT lesions could be correlated to gross pathologic findings and histologic granuloma types in representative pulmonary lobes. In conclusion, video-guided intrabronchial inoculation with spray catheters, mimicking the natural way of infection, resulted in pulmonary infection of goats with M. bovis. CT, but not DR, presented as a highly sensitive method to quantify the extent of pulmonary lesions. This goat model of TB may serve as a model for testing TB vaccine efficacy.
ABSTRACT
Background: VPM1002BC is a modified mycobacterium Bacillus Calmette Guérin (BCG) for the treatment of non-muscle invasive bladder cancer (NMIBC). The genetic modifications are expected to result in better immunogenicity and less side effects. We report on patient safety and immunology of the first intravesical application of VPM1002BC in human. Methods: Six patients with BCG failure received a treatment of 6 weekly instillations with VPM1002BC. Patients were monitored for adverse events (AE), excretion of VPM1002BC and cytokines, respectively. Results: No DLT (dose limiting toxicity) occurred during the DLT-period. No grade ≥3 AEs occurred. Excretion of VPM1002BC in the urine was limited to less than 24 hours. Plasma levels of TNFα significantly increased after treatment and blood-derived CD4+ T cells stimulated with PPD demonstrated significantly increased intracellular GM-CSF and IFN expression. Conclusion: The intravesical application of VPM1002BC is safe and well tolerated by patients and results in a potential Th1 weighted immune response.
Subject(s)
BCG Vaccine , Mycobacterium bovis , Urinary Bladder Neoplasms , Administration, Intravesical , Aged , Aged, 80 and over , BCG Vaccine/administration & dosage , Humans , Male , Mycobacterium bovis/genetics , Mycobacterium bovis/immunology , Urinary Bladder Neoplasms/drug therapyABSTRACT
Infections with the human cytomegalovirus (HCMV) are associated with severe clinical manifestations in children following prenatal transmission and after viral reactivation in immunosuppressed individuals. The development of an HCMV vaccine has long been requested but there is still no licensed product available. Subviral dense bodies (DB) are immunogenic in pre-clinical models and are thus a promising HCMV vaccine candidate. Recently, we established a virus based on the laboratory strain Towne that synthesizes large numbers of DB containing the pentameric protein complex gH/gL/UL128-131 (Towne-UL130repΔGFP). The work presented here focuses on providing strategies for the production of a safe vaccine based on that strain. A GMP-compliant protocol for DB production was established. Furthermore, the DB producer strain Towne-UL130rep was attenuated by deleting the UL25 open reading frame. Additional genetic modifications aim to abrogate its capacity to replicate in vivo by conditionally expressing pUL51 using the Shield-1/FKBP destabilization system. We further show that the terminase inhibitor letermovir can be used to reduce infectious virus contamination of a DB vaccine by more than two orders of magnitude. Taken together, strategies are provided here that allow for the production of a safe and immunogenic DB vaccine for clinical testing.
ABSTRACT
The only licensed vaccine against tuberculosis (TB), bacille Calmette-Guérin (BCG), protects against severe extrapulmonary forms of TB but is virtually ineffective against the most prevalent form of the disease, pulmonary TB. BCG was genetically modified at the Max Planck Institute for Infection Biology to improve its immunogenicity by replacing the urease C encoding gene with the listeriolysin encoding gene from Listeria monocytogenes. Listeriolysin perturbates the phagosomal membrane at acidic pH. Urease C is involved in neutralization of the phagosome harboring BCG. Its depletion allows for rapid phagosome acidification and promotes phagolysosome fusion. As a result, BCGΔureC::hly (VPM1002) promotes apoptosis and autophagy and facilitates release of mycobacterial antigens into the cytosol. In preclinical studies, VPM1002 has been far more efficacious and safer than BCG. The vaccine was licensed to Vakzine Projekt Management and later sublicensed to the Serum Institute of India Pvt. Ltd., the largest vaccine producer in the world. The vaccine has passed phase I clinical trials in Germany and South Africa, demonstrating its safety and immunogenicity in young adults. It was also successfully tested in a phase IIa randomized clinical trial in healthy South African newborns and is currently undergoing a phase IIb study in HIV exposed and unexposed newborns. A phase II/III clinical trial will commence in India in 2017 to assess efficacy against recurrence of TB. The target indications for VPM1002 are newborn immunization to prevent TB as well as post-exposure immunization in adults to prevent TB recurrence. In addition, a Phase I trial in non-muscle invasive bladder cancer patients has been completed, and phase II trials are ongoing. This review describes the development of VPM1002 from the drawing board to its clinical assessment.
ABSTRACT
Tuberculosis is a global threat to which infants are especially vulnerable. Effective vaccines are required to protect infants from this devastating disease. VPM1002, a novel recombinant Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccine previously shown to be safe and immunogenic in adults, was evaluated for safety in its intended target population, namely, newborn infants in a region with high prevalence of tuberculosis. A total of 48 newborns were vaccinated intradermally with VPM1002 (n = 36) or BCG Danish strain (n = 12) in a phase II open-labeled, randomized trial with a 6-month follow-up period. Clinical and laboratory measures of safety were evaluated during this time. In addition, vaccine-induced immune responses to mycobacteria were analyzed in whole-blood stimulation and proliferation assays. The safety parameters and immunogenicity were comparable in the two groups. Both vaccines induced interleukin-17 (IL-17) responses; however, VPM1002 vaccination led to an increase of CD8+ IL-17+ T cells at the week 16 and month 6 time points. The incidence of abscess formation was lower for VPM1002 than for BCG. We conclude that VPM1002 is a safe, well-tolerated, and immunogenic vaccine in newborn infants, confirming results from previous trials in adults. These results strongly support further evaluation of the safety and efficacy of this vaccination in larger studies. (This study has been registered at ClinicalTrials.gov under registration no. NCT01479972.).
Subject(s)
BCG Vaccine/adverse effects , BCG Vaccine/immunology , Tuberculosis/prevention & control , Abscess/epidemiology , Abscess/pathology , BCG Vaccine/administration & dosage , BCG Vaccine/genetics , CD8-Positive T-Lymphocytes/immunology , Drug-Related Side Effects and Adverse Reactions/epidemiology , Drug-Related Side Effects and Adverse Reactions/pathology , Female , Humans , Incidence , Infant, Newborn , Male , South Africa , Treatment Outcome , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/adverse effects , Vaccines, Synthetic/genetics , Vaccines, Synthetic/immunologyABSTRACT
Tuberculosis remains a major health threat and vaccines better than bacillus Calmette-Guérin (BCG) are urgently required. Here we describe our experience with a recombinant BCG expressing listeriolysin and deficient in urease. This potential replacement vaccine has demonstrated superior efficacy and safety over BCG in Mycobacterium tuberculosis aerosol-challenged mice and was safe in numerous animal models including immune-deficient mice, guinea pigs, rabbits and nonhuman primates. Phase I clinical trials in adults in Germany and South Africa have proven safety and a current Phase IIa trial is under way to assess immunogenicity and safety in its target population, newborns in a high tuberculosis incidence setting, with promising early results. Second-generation candidates are being developed to improve safety and efficacy.
Subject(s)
BCG Vaccine/administration & dosage , Clinical Trials as Topic/trends , Mycobacterium tuberculosis/drug effects , Tuberculosis/prevention & control , Animals , Humans , Mycobacterium tuberculosis/physiology , Tuberculosis/epidemiologyABSTRACT
BACKGROUND: Current vaccination using Mycobacterium bovis bacillus Calmette-Guérin (BCG), fails to prevent pulmonary tuberculosis (TB). New vaccination strategies are essential for reducing the global incidence of TB. We assessed the safety and immunogenicity of VPM1002, a recombinant BCG vaccine candidate. EudraCT (2007-002789-37) and ClinicalTrials.gov (NCT00749034). METHODS: Healthy volunteers were enrolled in a phase 1 open-label, dose escalation randomized clinical trial, and received one intradermal dose of VPM1002 (Mycobacterium bovis BCG ΔureC::hly Hm(R)) or BCG. Immunogenicity was assessed by interferon-gamma (IFN-γ) production, cellular immune response markers by flow cytometry and serum antibodies against mycobacterial antigens. RESULTS: Eighty volunteers were randomized into two groups according to previous BCG vaccination and mycobacterial exposure (BCG-naïve, n=40 and BCG-immune, n=40). In each group, 30 individuals were vaccinated with VPM1002 (randomized to three escalating doses) and 10 with BCG. VPM1002 was safe and stimulated IFN-γ-producing and multifunctional T cells, as well as antibody-producing B cells in BCG-naïve and BCG-immune individuals. CONCLUSIONS: VPM1002 was safe and immunogenic for B-cell and T-cell responses and hence will be brought forward through the clinical trial pipeline.
Subject(s)
BCG Vaccine/adverse effects , BCG Vaccine/immunology , Adolescent , Adult , Antibodies, Bacterial/blood , BCG Vaccine/administration & dosage , BCG Vaccine/genetics , Flow Cytometry , Healthy Volunteers , Humans , Injections, Intradermal , Interferon-gamma/metabolism , Leukocytes, Mononuclear/immunology , Male , Middle Aged , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/adverse effects , Vaccines, Synthetic/genetics , Vaccines, Synthetic/immunology , Young AdultABSTRACT
The failure of current Mycobacterium bovis bacille Calmette-Guérin (BCG) vaccines, given to neonates to protect against adult tuberculosis and the risk of using these live vaccines in HIV-infected infants, has emphasized the need for generating new, more efficacious and safer replacement vaccines. With the availability of genetic techniques for constructing recombinant BCG (rBCG) strains containing well-defined gene deletions or insertions, new vaccine candidates are under evaluation at both the preclinical and clinical stages of development. Since most BCG vaccines in use today were evaluated in clinical trials decades ago and are produced by outdated processes, the development of new BCG vaccines offers a number of advantages that include a modern well-defined manufacturing process along with state-of-the-art evaluation of safety and efficacy in target populations. We provide a description of the preclinical development of two novel rBCGs, VPM1002 that was constructed by adding a modified hly gene coding for the protein listeriolysin O (LLO) from Listeria monocytogenes and AERAS-422, which carries a modified pfoA gene coding for the protein perfringolysin O (PFO) from Clostridium perfringens, and three genes from Mycobacterium tuberculosis. Novel approaches like these should be helpful in generating stable and effective rBCG vaccine candidates that can be better characterized than traditional BCG vaccines.
ABSTRACT
BACKGROUND: New vaccines against tuberculosis (TB) are urgently needed because the only available vaccine, Mycobacterium bovis bacillus Calmette-Guérin (BCG), fails to protect against pulmonary TB in adults. The recombinant ΔureC hly+ BCG (rBCG) is more efficient than parental BCG (pBCG) against pulmonary TB in preclinical studies and has proven safe and immunogenic in phase I clinical trials. METHODS: In an attempt to identify the mechanisms underlying the superior protection of rBCG, we compared the immune responses elicited after vaccination and subsequent aerosol infection with Mycobacterium tuberculosis (MTB) in mice. RESULTS: We demonstrate that both rBCG and pBCG induce marked type 1 cytokine responses, whereas only rBCG elicits a profound type 17 cytokine response in addition. We observed earlier recruitment of antigen-specific T lymphocytes to the lung upon MTB infection of rBCG-vaccinated mice. These T cells produced abundant type 1 cytokines after restimulation, resulting in 10-fold reduced bacterial burden 90 days after infection. CONCLUSIONS: Our findings identify a general immunologic pathway for improved vaccination strategies against TB that can also be harnessed by other vaccine candidates.
Subject(s)
BCG Vaccine/immunology , CD4-Positive T-Lymphocytes/metabolism , Cytokines/metabolism , Interleukin-17/metabolism , Mycobacterium tuberculosis/immunology , Tuberculosis/immunology , Animals , Female , Mice , Mice, Inbred BALB C , Tuberculosis/prevention & control , Vaccines, Synthetic/immunologyABSTRACT
Although tuberculosis remains a substantial global threat, the mechanisms that enable mycobacterial persistence and replication within the human host are ill defined. This study represents the first genome-wide expression analysis of Mycobacterium tuberculosis from clinical lung samples, which has enabled the identification of M. tuberculosis genes actively expressed during pulmonary tuberculosis. To obtain optimal information from our DNA array analyses, we analyzed the differentially expressed genes within the context of computationally inferred protein networks. Protein networks were constructed using functional linkages established by the Rosetta stone, phylogenetic profile, conserved gene neighbor, and operon computational methods. This combined approach revealed that during pulmonary tuberculosis, M. tuberculosis actively transcribes a number of genes involved in active fortification and evasion from host defense systems. These genes may provide targets for novel intervention strategies.
Subject(s)
Bacterial Proteins/metabolism , Gene Expression Profiling , Genome, Bacterial , Mycobacterium tuberculosis/pathogenicity , Oligonucleotide Array Sequence Analysis , Tuberculosis, Pulmonary/microbiology , Bacterial Proteins/genetics , Computational Biology/methods , Gene Expression Regulation, Bacterial , Humans , Lung/microbiology , Lung/surgery , Mycobacterium tuberculosis/genetics , Mycobacterium tuberculosis/isolation & purification , Mycobacterium tuberculosis/metabolism , Proteome , Transcription, Genetic , Tuberculosis, Multidrug-Resistant/microbiology , Tuberculosis, Multidrug-Resistant/surgery , Tuberculosis, Pulmonary/surgeryABSTRACT
The tuberculosis vaccine Mycobacterium bovis bacille Calmette-Guérin (BCG) was equipped with the membrane-perforating listeriolysin (Hly) of Listeria monocytogenes, which was shown to improve protection against Mycobacterium tuberculosis. Following aerosol challenge, the Hly-secreting recombinant BCG (hly+ rBCG) vaccine was shown to protect significantly better against aerosol infection with M. tuberculosis than did the parental BCG strain. The isogenic, urease C-deficient hly+ rBCG (DeltaureC hly+ rBCG) vaccine, providing an intraphagosomal pH closer to the acidic pH optimum for Hly activity, exhibited still higher vaccine efficacy than parental BCG. DeltaureC hly+ rBCG also induced profound protection against a member of the M. tuberculosis Beijing/W genotype family while parental BCG failed to do so consistently. Hly not only promoted antigen translocation into the cytoplasm but also apoptosis of infected macrophages. We concluded that superior vaccine efficacy of DeltaureC hly+ rBCG as compared with parental BCG is primarily based on improved cross-priming, which causes enhanced T cell-mediated immunity.
Subject(s)
BCG Vaccine , Bacterial Toxins/metabolism , Heat-Shock Proteins/metabolism , Listeria monocytogenes/metabolism , Tuberculosis/prevention & control , Adult , Animals , Apoptosis , Bacterial Proteins/genetics , Bacterial Proteins/immunology , Cells, Cultured , Child , Hemolysin Proteins , Humans , Hydrogen-Ion Concentration , Listeria monocytogenes/immunology , Macrophages/cytology , Macrophages/metabolism , Mice , Mice, SCID , Mycobacterium tuberculosis/immunology , Mycobacterium tuberculosis/metabolism , Recombinant Proteins/immunology , Recombinant Proteins/therapeutic use , Survival Rate , Vaccines, SyntheticABSTRACT
The TB Vaccine Cluster project funded by the EU Fifth Framework programme aims to provide novel vaccines against tuberculosis that are suitable for evaluation in humans. This paper describes the studies of the protective efficacy of vaccines in a guinea pig aerosol-infection model of primary tuberculosis. The objective was to conduct comparative evaluations of vaccines that had previously demonstrated efficacy in other animal models. Groups of 6 guinea pigs were immunized with vaccines provided by the relevant EU Vaccine Cluster partners. Survival over 17 or 26 weeks was used as the principal measure of vaccine efficacy following aerosol challenge with H37Rv. Counts of mycobacteria in lungs and spleens, and histopathological changes in the lungs, were also used to provide evidence of protection. A total of 24 vaccines were evaluated in 4 experiments each of a different design. A heterologous prime-boost strategy of DNA and MVA, each expressing Ag85A and a fusion protein of ESAT-6 and Ag85B in adjuvant, protected the guinea pigs to the same extent as BCG. Genetically modified BCG vaccines and boosted BCG strategies also protected guinea pigs to the same extent as BCG but not statistically significantly better. A relatively high aerosol-challenge dose and evaluation over a protracted time post-challenge allowed superior protection over BCG to be demonstrated by BCG boosted with MVA and fowl pox vectors expressing Ag85A.
Subject(s)
Disease Models, Animal , Tuberculosis Vaccines/therapeutic use , Tuberculosis/prevention & control , Aerosols , Animals , BCG Vaccine/therapeutic use , Colony Count, Microbial/methods , Drug Evaluation, Preclinical/methods , European Union , Guinea Pigs , Humans , Lung/microbiology , Mycobacterium tuberculosis/immunology , Mycobacterium tuberculosis/isolation & purification , Spleen/microbiology , Survival Analysis , Tuberculosis/immunology , Tuberculosis Vaccines/immunology , Tuberculosis, Pulmonary/immunology , Tuberculosis, Pulmonary/prevention & control , Vaccination/methodsABSTRACT
BACKGROUND: Studies of different vaccine constructs have demonstrated variable efficacy against Mycobacterium tuberculosis in animal models. Despite the fact that these vaccines have used one or another of a very small number of immunodominant antigens, a direct comparison of the relative efficacy of the antigens and delivery systems has been difficult, because the studies have used different parameters for assessment. METHODS: We compared the efficacies of the most commonly used vaccine constructs--adjuvanted protein, plasmid DNA, and live bacterial vectors--bearing the immunodominant secreted antigens early secreted antigen target-6 and antigen 85B, either alone or as a fusion protein. Mice were vaccinated with these constructs, and the effects of different delivery systems on protective efficacy (as assessed by survival studies and by monitoring bacterial load) and antigen-specific responses (including the contribution of CD4 and CD8 T cells to these responses) were assayed by various methods. RESULTS: The relative efficacy of different vaccines is dependent on the delivery system, the antigen, and the animal model. Likewise, the relative immunodominance of individual antigens in the fusion molecule is altered by the choice of delivery system. CONCLUSION: These results clearly demonstrate the importance of assessing vaccine function by use of multiple parameters and indicate which parameters are most reliable for assessing vaccine efficacy.
Subject(s)
Tuberculosis Vaccines/immunology , Acyltransferases/immunology , Animals , Antigens, Bacterial/immunology , Bacterial Proteins/immunology , Female , Lymphocyte Activation/immunology , Male , Mice , Mice, Inbred C57BL , Mycobacterium tuberculosis/immunology , Recombinant Fusion Proteins/immunology , Specific Pathogen-Free Organisms , T-Lymphocytes/immunology , Time Factors , Tuberculosis/prevention & control , Vaccines, Synthetic/immunologyABSTRACT
Information from comparative proteome analysis of Mycobacterium tuberculosis and Mycobacterium bovis bacillus Calmette-Guerin (BCG) principally allows prediction of potential vaccine candidates. Thirty-six M. tuberculosis DNA vaccine candidates identified by comparative proteome analysis were evaluated in the mouse model for protection against low-dose aerosol M. tuberculosis infection. We identified the DNA vaccine candidate Rv3407 as a protective antigen and analyzed putative major histocompatibility complex class I epitopes by computational predictions and gamma interferon Elispot assays. Importantly, we discovered that the DNA vaccine Rv3407 improved the efficacy of BCG vaccination in a heterologous prime-boost vaccination protocol. Our data demonstrate the rationale of a combination of proteomics, epitope prediction, and broad screening of putative antigens for identification of novel DNA vaccine candidates. Furthermore, our experiments show that heterologous prime-boost vaccination with a defined antigen boost "on top" of a BCG primer provides superior protection against tuberculosis over vaccination with BCG alone.
Subject(s)
Antigens, Bacterial , BCG Vaccine , Immunization, Secondary , Proteome , Tuberculosis Vaccines , Tuberculosis/prevention & control , Vaccines, DNA , Animals , Antibodies, Bacterial/blood , Antigens, Bacterial/genetics , Antigens, Bacterial/immunology , Antigens, Bacterial/metabolism , BCG Vaccine/immunology , Bacterial Proteins/genetics , Bacterial Proteins/immunology , Bacterial Proteins/metabolism , Cattle , Computational Biology , Drug Design , Humans , Interferon-gamma/metabolism , Mice , Mice, Inbred BALB C , Mycobacterium bovis/immunology , Mycobacterium bovis/metabolism , Mycobacterium tuberculosis/immunology , Mycobacterium tuberculosis/metabolism , Tuberculosis Vaccines/immunology , Vaccination , Vaccines, DNA/immunologyABSTRACT
Immunization with plasmid DNA vectors represents a promising new approach to vaccination. It has been shown to elicit humoral and cellular immunity and protection in various infection models. Here, we assessed the immunogenicity and protective efficacy of a DNA vaccine vector encoding the antigen 85A (Ag85A) of Mycobacterium tuberculosis. Since intramuscular (i.m.) immunization with naked DNA requires considerable amounts of DNA in order to be effective, we evaluated a strategy to reduce the amount of DNA needed. To this end, we used Ag85A DNA adsorbed onto cationic poly(DL-lactide-co-glycolide) (PLG) microparticles and observed similar levels of protection against aerosol challenge in mice using doses of PLG-DNA two orders of magnitude lower than with naked DNA itself.