BCG-induced Immune Responses Research Articles

Macrophages exploit the mannose receptor and JAK-STAT1-MHC-II pathway to drive antigen presentation and the antimycobacterial immune response after BCG vaccination.

Tuberculosis (TB), caused by Mycobacterium tuberculosis ( M. tb), remains one of the leading causes of fatal infectious diseases worldwide. The only licensed vaccine, Mycobacterium bovis Bacillus Calmette-Guérin (BCG), has variable efficacy against TB in adults. Insufficiency of immune cell function diminishes the protective effects of the BCG vaccine. It is critical to clarify the mechanism underlying the antimycobacterial immune response during BCG vaccination. Macrophage mannose receptor (MR) is important for enhancing the uptake and processing of glycoconjugated antigens from pathogens for presentation to T cells, but the roles of macrophage MR in the BCG-induced immune response against M. tb are not yet clear. Here, we discover that macrophage MR deficiency impairs the antimycobacterial immune response in BCG-vaccinated mice. Mechanistically, macrophage MR triggers JAK-STAT1 signaling, which promotes antigen presentation via upregulated MHC-II and induces IL-12 production by macrophages, contributing to CD4 + T cell activation and IFN-γ production. MR deficiency in macrophages reduces the vaccine efficacy of BCG and increases susceptibility to M. tb H37Ra challenge in mice. Our results suggest that MR is critical for macrophage antigen presentation and the antimycobacterial immune response to BCG vaccination and offer valuable guidance for the preventive strategy of BCG immunization.

BCG immunotherapy promotes tumor-derived T-cell activation through the FLT3/FLT3LG pathway in bladder cancer.

Bladder instillation therapy is a common treatment for superficial or nonmuscle invasive bladder cancer. After surgery or reresection, chemotherapy drugs (epirubicin) or medications such as Bacillus Calmette-Guérin (BCG) are used for bladder instillation therapy, which can reduce the risk of bladder cancer recurrence and progression. However, the specific mechanism by which BCG stimulates the antitumor response has not been thoroughly elucidated. Additionally, although BCG immunotherapy is effective, it is difficult to predict which patients will have a positive response. In this study, we explored the BCG-induced immune response and found that high levels of Fms-related receptor tyrosine kinase 3 ligand (FLT3LG) were expressed after BCG treatment. This FLT3LG can directly act on CD8+ T cells and promote their proliferation and activation. The use of FLT3 inhibitors can neutralize the antitumor effects of BCG. In vitro experiments showed that FLT3LG can synergize with T-cell receptor activators to promote the activation of tumor-derived T cells. This study partially elucidates the mechanism of CD8+ T-cell activation in BCG immunotherapy and provides a theoretical basis for optimizing BCG instillation therapy in bladder cancer.

A CD4+ TNF+ monofunctional memory T-cell response to BCG vaccination is associated with Mycobacterium tuberculosis infection in infants exposed to HIV.

SummaryBackgroundThe immunologic correlates of risk of Mycobacterium tuberculosis (Mtb) infection after BCG vaccination are unknown. The mechanism by which BCG influences the tuberculin skin test (TST) remains poorly understood. We evaluated CD4+ T-cell responses in infants exposed to HIV and uninfected (HEU) who received BCG at birth and examined their role in susceptibility to Mtb infection and influence on TST induration.MethodsHEU infants were enrolled in a randomised clinical trial of isoniazid (INH) to prevent Mtb infection in Kenya. We measured mycobacterial antigen-specific Th1 and Th17 cytokine responses at 6–10 weeks of age prior to INH randomisation and compared responses between Mtb infected and uninfected infants. Outcomes at 14 months of age included TST, QuantiFERON-Plus (QFT-Plus), and ESAT-6/CFP-10-specific non-IFN-γ cytokines measured in QFT-Plus supernatants.FindingsA monofunctional mycobacterial antigen-specific TNF+ CD4+ effector memory (CCR7-CD45RA-) T-cell response at 6–10 weeks of age was associated with Mtb infection at 14 months of age as measured by ESAT-6/CFP-10-specific IFN-γ and non-IFN-γ responses (Odds Ratio 2.26; Confidence Interval 1.27–4.15; P = 0.006). Mycobacterial antigen-specific polyfunctional effector memory Th1 responses at 6–10 weeks positively correlated with TST induration in infants without evidence of Mtb infection at 14 months, an association which was diminished by INH therapy.InterpretationInduction of monofunctional TNF+ CD4+ effector memory T-cell responses may be detrimental in TB vaccine development. This study also provides mechanistic insight into the association of BCG-induced immune responses with TST induration and further evidence that TST-based diagnoses of Mtb infection in infants are imprecise.FundingThrasher Research Fund.

Induction of Functional Specific Antibodies, IgG-Secreting Plasmablasts and Memory B Cells Following BCG Vaccination.

Tuberculosis (TB) is a major global health problem and the only currently-licensed vaccine, BCG, is inadequate. Many TB vaccine candidates are designed to be given as a boost to BCG; an understanding of the BCG-induced immune response is therefore critical, and the opportunity to relate this to circumstances where BCG does confer protection may direct the design of more efficacious vaccines. While the T cell response to BCG vaccination has been well-characterized, there is a paucity of literature on the humoral response. We demonstrate BCG vaccine-mediated induction of specific antibodies in different human populations and macaque species which represent important preclinical models for TB vaccine development. We observe a strong correlation between antibody titers in serum versus plasma with modestly higher titers in serum. We also report for the first time the rapid and transient induction of antibody-secreting plasmablasts following BCG vaccination, together with a robust and durable memory B cell response in humans. Finally, we demonstrate a functional role for BCG vaccine-induced specific antibodies in opsonizing mycobacteria and enhancing macrophage phagocytosis in vitro, which may contribute to the BCG vaccine-mediated control of mycobacterial growth observed. Taken together, our findings indicate that the humoral immune response in the context of BCG vaccination merits further attention to determine whether TB vaccine candidates could benefit from the induction of humoral as well as cellular immunity.

Novel personalized cancer vaccine platform based on Bacillus Calmette-Guèrin

BackgroundIntratumoral BCG therapy, one of the earliest immunotherapies, can lead to infiltration of immune cells into a treated tumor. However, an increase in the number of BCG-induced tumor-specific T cells...

A century of BCG: Impact on tuberculosis control and beyond.

BCG turns 100 this year and while it might not be the perfect vaccine, it has certainly contributed significantly towards eradication and prevention of spread of tuberculosis (TB). The search for newer and better vaccines for TB is an ongoing endeavor and latest results from trials of candidate TB vaccines such as M72AS01 look promising. However, recent encouraging data from BCG revaccination trials in adults combined with studies on mucosal and intravenous routes of BCG vaccination in non-human primate models have renewed interest in BCG for TB prevention. In addition, several well-demonstrated non-specific effects of BCG, for example, prevention of viral and respiratory infections, give BCG an added advantage. Also, BCG vaccination is currently being widely tested in human clinical trials to determine whether it protects against SARS-CoV-2 infection and/or death with detailed analyses and outcomes from several ongoing trials across the world awaited. Through this review, we attempt to bring together information on various aspects of the BCG-induced immune response, its efficacy in TB control, comparison with other candidate TB vaccines and strategies to improve its efficiency including revaccination and alternate routes of administration. Finally, we discuss the future relevance of BCG use especially in light of its several heterologous benefits.

Tumor Immune Microenvironment in Response to Radiotherapy vs BCG in a Murine Model of Bladder Cancer

The role of radiotherapy combined with immunotherapy in patients with non-muscle invasive bladder cancer (NMIBC) is currently being investigated. Radiation induced DNA damage triggering immunogenic tumor cell death and consequent immune responses in bladder tumor microenvironment is not fully understood. We compared local immune response following radiotherapy with BCG-induced immune response using a BBN murine model of urothelial carcinoma (UC). Female C57BL/6 control(no tumor, n=15) or BBN exposed mice(tumor, n=13) received radiotherapy(single external beam dose,15 Gy,n=10), intravesical BCG instillation (3 × 100μL of 1 × 10 8 CFU,n=9) or saline treatment (n=9). Tumor involvement was assessed by pathologic grading of bladder tissue and immune responses quantified using Nanostring Myeloid Innate Immunity Panel. Infiltrating Tcells were evaluated by CD4, CD8 and FOXP3 immunohistochemistry (IHC) and tissue macrophages quantified as M1 or M2 phenotype based on CD86 and CD206 IHC. No tumors were seen in BBN+BCG bladder tissues compared to BBN+saline or BBN+radiotherapy. Intravesical BCG instillation in BBN mice elevated cytosolic DNA sensing pathway as seen by increased mRNA levels of tmem173 (STING) that was consistent with elevated levels of downstream cytokines tnfa, infg, il6 and chemokine cxcl10 . mRNA expression of 3’exonuclease trex1 , that degrades 3′ ends of nicked DNA during immunogenic cell death, was elevated in BBN+BCG as well as BBN+radiation samples. BBN+BCG samples had increased CD8/FOXP3 and CD4/FOXP3 ratios assessed by IHC and increased M1 macrophage shift validated by mRNA expression and IHC. BCG instillation caused a robust proinflammatory immune response compared to radiation in murine UC possibly via STING-dependent induction of proinflammatory cytokines and chemokines concomitant with shift toward M1 macrophage polarization and increased Teff/Treg ratios. Elevated trex1 may degrade DNA damage as a direct response to radiation thereby inhibiting STING activation and cytokine response in radiated samples. These findings highlight the need to uncover mechanisms of radiation induced immune response in NMIBC to optimize utilization of immunotherapy with radiotherapy.

Evidence for the Effect of Vaccination on Host-Pathogen Interactions in a Murine Model of Pulmonary Tuberculosis by Mycobacterium tuberculosis.

The global control of Tuberculosis remains elusive, and Bacillus Calmette-Guérin (BCG) -the most widely used vaccine in history—has proven insufficient for reversing this epidemic. Several authors have suggested that the mass presence of vaccinated hosts might have affected the Mycobacterium tuberculosis (MTB) population structure, and this could in turn be reflected in a prevalence of strains with higher ability to circumvent BCG-induced immunity, such as the recent Beijing genotype. The effect of vaccination on vaccine-escape variants has been well-documented in several bacterial pathogens; however the effect of the interaction between MTB strains and vaccinated hosts has never been previously described. In this study we show for the first time the interaction between MTB Beijing-genotype strains and BCG-vaccinated hosts. Using a well-controlled murine model of progressive pulmonary tuberculosis, we vaccinated BALB/c mice with two different sub-strains of BCG (BCG-Phipps and BCG-Vietnam). Following vaccination, the mice were infected with either one of three selected MTB strains. Strains were selected based on lineage, and included two Beijing-family clinical isolates (strains 46 and 48) and a well-characterized laboratory strain (H37Rv). Two months after infection, mice were euthanized and the bacteria extracted from their lungs. We characterized the genomic composite of the bacteria before and after exposure to vaccinated hosts, and also characterized the local response to the bacteria by sequencing the lung transcriptome in animals during the infection. Results from this study show that the interaction within the lungs of the vaccinated hosts results in the selection of higher-virulence bacteria, specifically for the Beijing genotype strains 46 and 48. After exposure to the BCG-induced immune response, strains 46 and 48 acquire genomic mutations associated with several virulence factors. As a result, the bacteria collected from these vaccinated hosts have an increased ability for immune evasion, as shown in both the host transcriptome and the histopathology studies, and replicates far more efficiently compared to bacteria collected from unvaccinated hosts or to the original-stock strain. Further research is warranted to ascertain the pathways associated with the genomic alterations. However, our results highlight novel host-pathogen interactions induced by exposure of MTB to BCG vaccinated hosts.

Clinical and molecular insights into BCG immunotherapy for melanoma.

The incidence of cutaneous melanoma and the mortality rate of advanced melanoma patients continue to rise globally. Despite the recent success of immunotherapy including ipilimumab and pembrolizumab checkpoint inhibitors, a large proportion of patients are refractory to such treatment modalities. The application of mycobacteria such as Bacillus Calmette-Guérin (BCG) in the treatment of various malignancies, including cutaneous melanoma, has been clearly demonstrated after almost a century of observations and experimentation. Intralesional BCG (IL-BCG) immunotherapy is a highly efficient and cost-effective treatment option for inoperable stage III in-transit melanoma, as recommended in the National Comprehensive Cancer Network Guidelines. IL-BCG has shown great efficacy in the regression of directly injected metastatic melanoma lesions, as well as distal noninjected nodules in immunocompetent patients. Clinical and preclinical studies have shown that BCG serves as a strong immune modulator, inducing the recruitment of various immune cells that contribute to antitumour immunity. However, the specific mechanism of BCG-mediated tumour immunity remains poorly understood. Comparative genome analyses have revealed that different BCG strains exhibit distinct immunological activity and virulence, which might impact the therapeutic response and clinical outcome of patients. In this review, we discuss the immunostimulatory potential of different BCG substrains and highlight clinical studies utilizing BCG immunotherapy for the treatment of cutaneous melanoma. Furthermore, the review focuses on the cellular and molecular mechanisms of the BCG-induced immune responses of both the innate and adaptive arms of the immune system. Furthermore, the review discussed the administration of BCG as a monotherapy or in combination with other immunotherapeutic or chemotherapeutic agents.

Licensed Bacille Calmette-Guérin (BCG) formulations differ markedly in bacterial viability, RNA content and innate immune activation

BackgroundBacille Calmette-Guérin (BCG), the live attenuated tuberculosis vaccine, is manufactured under different conditions across the globe generating formulations that may differ in clinical efficacy. Innate immune recognition of live BCG contributes to immunogenicity suggesting that differences in BCG viability may contribute to divergent activity of licensed formulations. MethodsWe compared BCG-Denmark (DEN), -Japan (JPN), -India (IND), -Bulgaria (BUL) and -USA in vitro with respect to a) viability as measured by colony-forming units (CFU), mycobacterial membrane integrity, and RNA content, and b) cytokine/chemokine production in newborn cord and adult peripheral blood. ResultsUpon culture, relative growth was BCG-USA > JPN ≫ DEN > BUL = IND. BCG-IND and -BUL demonstrated >1000-fold lower growth than BCG-JPN in 7H9 medium and >10-fold lower growth in commercial Middlebrook 7H11 medium. BCG-IND demonstrated significantly decreased membrane integrity, lower RNA content, and weaker IFN-γ inducing activity in whole blood compared to other BCGs. BCG-induced whole blood cytokines differed significantly by age, vaccine formulation and concentration. BCG-induced cytokine production correlated with CFU, suggesting that mycobacterial viability may contribute to BCG-induced immune responses. ConclusionsLicensed BCG vaccines differ markedly in their content of viable mycobacteria possibly contributing to formulation-dependent activation of innate and adaptive immunity and distinct protective effects.

Absolute basophil count is associated with time to recurrence in patients with high-grade T1 bladder cancer receiving bacillus Calmette-Guérin after transurethral resection of the bladder tumor.

Basophils, eosinophils and monocytes may be involved in BCG-induced immune responses and be associated with outcomes of bladder cancer patients receiving intravesical BCG. Our objective was to explore the association of baseline counts of basophils, eosinophils and monocytes with outcomes of patients with high-grade T1 bladder cancer receiving a standard course of intravesical BCG. We retrospectively reviewed medical records of patients with primary T1 HG/G3 bladder cancer. After re-TURBT, patients were treated with a 6-week course of intravesical BCG induction followed by intravesical BCG every week for 3weeks given at 3, 6, 12, 18, 24, 30 and 36months from initiation of therapy The analysis of potential risk factors for recurrence, muscle invasion and cancer-specific and overall survival was performed using univariable Cox regression models. Those factors that presented, at univariate analysis, an association with the event at a liberal p < 0.1, have been selected for the development of a multivariable model. A total of 1045 patients with primary T1 HG/G3 were included. A total of 678 (64.9%) recurrences, 303 (29.0%) progressions and 150 (14.3%) deaths were observed during follow-up. Multivariate analysis showed that logarithmic transformation of basophils count was associated with a 30% increment in the hazard of recurrence per unit increase of logarithmic basophils count (HR 1.30; 95% confidence interval 1.09-1.54; p = 0.0026). Basophil count modeled by quartiles was also significantly associated with time to recurrence [second vs. lower quartile HR 1.42 (1.12-1.79); p = 0.003, third vs. lower quartile HR 1.26 (1.01-1.57); p = 0.041; upper vs. lower quartile HR 1.36 (1.1-1.68); p = 0.005]. The limitations of a retrospective study are applicable. Baseline basophil count may predict recurrence in BCG-treated HG/G3 T1 bladder cancer patients. External validation is warranted.

Differential DNA methylation of potassium channel KCa3.1 and immune signalling pathways is associated with infant immune responses following BCG vaccination

Bacillus Calmette–Guérin (BCG) is the only licensed vaccine for tuberculosis (TB) and induces highly variable protection against pulmonary disease in different countries. We hypothesised that DNA methylation is one of the molecular mechanisms driving variability in BCG-induced immune responses. DNA methylation in peripheral blood mononuclear cells (PBMC) from BCG vaccinated infants was measured and comparisons made between low and high BCG-specific cytokine responders. We found 318 genes and 67 pathways with distinct patterns of DNA methylation, including immune pathways, e.g. for T cell activation, that are known to directly affect immune responses. We also highlight signalling pathways that could indirectly affect the BCG-induced immune response: potassium and calcium channel, muscarinic acetylcholine receptor, G Protein coupled receptor (GPCR), glutamate signalling and WNT pathways. This study suggests that in addition to immune pathways, cellular processes drive vaccine-induced immune responses. Our results highlight mechanisms that require consideration when designing new TB vaccines.

Using Data from Macaques To Predict Gamma Interferon Responses after Mycobacterium bovis BCG Vaccination in Humans: a Proof-of-Concept Study of Immunostimulation/Immunodynamic Modeling Methods.

ABSTRACTMacaques play a central role in the development of human tuberculosis (TB) vaccines. Immune and challenge responses differ across macaque and human subpopulations. We used novel immunostimulation/immunodynamic modeling methods in a proof-of-concept study to determine which macaque subpopulations best predicted immune responses in different human subpopulations. Data on gamma interferon (IFN-γ)-secreting CD4+ T cells over time after recent Mycobacterium bovis BCG vaccination were available for 55 humans and 81 macaques. Human population covariates were baseline BCG vaccination status, time since BCG vaccination, gender, and the monocyte/lymphocyte cell count ratio. The macaque population covariate was the colony of origin. A two-compartment mathematical model describing the dynamics of the IFN-γ T cell response after BCG vaccination was calibrated to these data using nonlinear mixed-effects methods. The model was calibrated to macaque and human data separately. The association between subpopulations and the BCG immune response in each species was assessed. The macaque subpopulations that best predicted immune responses in different human subpopulations were identified using Bayesian information criteria. We found that the macaque colony and the human baseline BCG status were significantly (P < 0.05) associated with the BCG-induced immune response. For humans who were BCG naïve at baseline, Indonesian cynomolgus macaques and Indian rhesus macaques best predicted the immune response. For humans who had already been BCG vaccinated at baseline, Mauritian cynomolgus macaques best predicted the immune response. This work suggests that the immune responses of different human populations may be best modeled by different macaque colonies, and it demonstrates the potential utility of immunostimulation/immunodynamic modeling to accelerate TB vaccine development.

HSP90B1 Regulates TLR-dependent Monocyte Signaling and its Common Variants are Associated with BCG-specific T-cell Responses and Protection from Pediatric TB Disease

Abstract Introduction HSP90B1 is a chaperone for multiple Toll-like receptors (TLRs), regulates innate immune signaling in murine myeloid cells and is expressed in T-cells. We hypothesized human HSP90B1 regulates monocyte responses to Mycobacterium tuberculosis (Mtb) and that its variants are associated with T-cell responses to the BCG vaccine and susceptibility to TB disease. Methods &amp; Results We screened 17 haplotype-tagging HSP90B1 gene region SNPs for association with BCG-induced immune responses in South African infants vaccinated with BCG at birth. We stimulated peripheral blood drawn at 10 weeks of age with BCG in a whole blood cytokine assay. Three HSP90B1 SNPs (rs10507172, 10507173, and 2164747) were associated with increased IL-2 secretion (P&amp;lt;0.05, generalized linear model). SNPrs10507173 was associated with increased IL-2 production in the CD4+ T-cell subset (p=0.028, ANOVA) based on intracellular cytokine staining and flow cytometry. The same three alleles were associated with protection from pediatric TB disease in a gene association study of 226 children who contracted tuberculosis compared with 626 controls who remained healthy (dominant model, unadjusted p&amp;lt;0.05, OR 0.58–0.72; after adjusting for ethnicity, p&amp;lt;0.05 for 2/3 SNPs). We used CRISPR/Cas9 gene editing to reduce HSP90B1 expression in U937 monocyte cells. Relative to controls, HSP90B1-deficient cells secreted less TNF when exposed to TLR1/2/6 (p=0.02) and TLR4 (p=0.01) ligands and Mtb whole cell lysate (p=0.001). Conclusion These data suggest HSP90B1 deficiency is associated with an impaired monocyte response to Mtb ligands. In infants, HSP90B1 variants are associated with increased BCG-specific IL-2 responses T-cells and protection from TB disease.

The HyVac4 Subunit Vaccine Efficiently Boosts BCG-Primed Anti-Mycobacterial Protective Immunity

BackgroundThe current vaccine against tuberculosis (TB), BCG, has failed to control TB worldwide and the protective efficacy is moreover limited to 10–15 years. A vaccine that could efficiently boost a BCG-induced immune response and thus prolong protective immunity would therefore have a significant impact on the global TB-burden.Methods/FindingsIn the present study we show that the fusion protein HyVac4 (H4), consisting of the mycobacterial antigens Ag85B and TB10.4, given in the adjuvant IC31® or DDA/MPL effectively boosted and prolonged immunity induced by BCG, leading to improved protection against infection with virulent M. tuberculosis (M.tb). Increased protection correlated with an increased percentage of TB10.4 specific IFNγ/TNFα/IL-2 or TNFα/IL-2 producing CD4 T cells at the site of infection. Moreover, this vaccine strategy did not compromise the use of ESAT-6 as an accurate correlate of disease development/vaccine efficacy. Indeed both CD4 and CD8 ESAT-6 specific T cells showed significant correlation with bacterial levels.Conclusions/SignificanceH4-IC31® can efficiently boost BCG-primed immunity leading to an increased protective anti-M.tb immune response dominated by IFNγ/TNFα/IL-2 or TNFα/IL2 producing CD4 T cells. H4 in the CD4 T cell inducing adjuvant IC31® is presently in clinical trials.

Immunotherapy for Urothelial Carcinoma: Current Status and Perspectives

Intravesical instillation of bacillus Calmette Guérin (BCG) for the treatment of urothelial carcinoma (UC) of the bladder is based on the BCG-induced immune response, which eradicates and prevents bladder cancer. The results of recent studies have suggested that not only major histocompatibility complex (MHC)-nonrestricted immune cells such as natural killer cells, macrophages, neutrophils, etc., but also MHC-restricted CD8+ T cells play an important role and are one of the main effectors in this therapy. Better understanding of the mechanism of BCG immunotherapy supports the idea that active immunotherapy through its augmented T cell response can have great potential for the treatment of advanced UC. In this review, progress in immunotherapy for UC is discussed based on data from basic, translational and clinical studies. We also review the escape mechanism of cancer cells from the immune system, and down-regulation of MHC class I molecules.

Dose-Finding Study of the Novel Tuberculosis Vaccine, MVA85A, in Healthy BCG-Vaccinated Infants

BCG, the only licensed tuberculosis vaccine, affords poor protection against lung tuberculosis in infants and children. A new tuberculosis vaccine, which may enhance the BCG-induced immune response, is urgently needed. We assessed the safety of and characterized the T cell response induced by 3 doses of the candidate vaccine, MVA85A, in BCG-vaccinated infants from a setting where tuberculosis is endemic. Infants aged 5-12 months were vaccinated intradermally with either 2.5 × 10(7), 5 × 10(7), or 10 × 10(7) plaque-forming units of MVA85A, or placebo. Adverse events were documented, and T-cell responses were assessed by interferon γ (IFN-γ) enzyme-linked immunospot assay and intracellular cytokine staining. The 3 MVA85A doses were well tolerated, and no vaccine-related serious adverse events were recorded. MVA85A induced potent, durable T-cell responses, which exceeded prevaccination responses up to 168 d after vaccination. No dose-related differences in response magnitude were observed. Multiple CD4 T cell subsets were induced; polyfunctional CD4 T cells co-expressing T-helper cell 1 cytokines with or without granulocyte-macrophage colony-stimulating factor predominated. IFN-γ-expressing CD8 T cells, which peaked later than CD4 T cells, were also detectable. MVA85A was safe and induced robust, polyfunctional, durable CD4 and CD8 T-cell responses in infants. These data support efficacy evaluation of MVA85A to prevent tuberculosis in infancy. Clinical Trials Registration. NCT00679159.

Induction of Granulysin and Perforin Cytolytic Mediator Expression in 10-Week-Old Infants Vaccinated with BCG at Birth

Background. While vaccination at birth with Mycobacterium bovis Bacilli Calmette-Guérin (BCG) protects against severe childhood tuberculosis, there is no consensus as to which components of the BCG-induced immune response mediate this protection. However, granulysin and perforin, found in the granules of cytotoxic T lymphocytes and Natural Killer (NK) cells, can kill intracellular mycobacteria and are implicated in protection against Mycobacterium tuberculosis. Methods. We compared the cellular expression of granulysin and perforin cytolytic molecules in cord blood and peripheral blood from 10-week-old infants vaccinated at birth with either Japanese or Danish BCG, administered either intradermally or percutaneously. Results. In cord blood, only CD56+ NK cells expressed granulysin and perforin constitutively. These cytolytic mediators were upregulated in CD4+ and CD8+ cord blood cells by ex vivo stimulation with BCG but not with PPD. Following BCG vaccination of neonates, both BCG and PPD induced increased expression of granulysin and perforin by CD4+ and CD8+ T cells. There was no difference in expression of cytolytic molecules according to vaccination route or strain. Conclusions. Constitutive expression of perforin and granulysin by cord blood NK-cells likely provides innate immunity, while BCG vaccination-induced expression of these cytolytic mediators may contribute towards protection of the neonate against tuberculosis.

Reactive antibodies against bacillus Calmette‐Guerin heat‐shock protein‐65 potentially predict the outcome of immunotherapy for high‐grade transitional cell carcinoma of the bladder

Intravesical immunotherapy with Mycobacterium bovis (M. bovis) bacillus Calmette-Guerin (BCG) is the current standard of care against superficial, high-grade transitional cell carcinoma (TCC) of the urinary bladder (carcinoma in situ and pathologic T1, grade 3 disease). However, individual patient outcome is barely predictable because of the lack of serum markers. Consequently, progression to muscle-invasive bladder cancer and critical delay of treatments (such as neoadjuvant combination chemotherapy and/or radical cystectomy) often occur. The objectives of this study were to identify a marker for measuring the BCG-induced immune response and to predict the outcomes and potential improvements of BCG immunotherapy. Because host immunoresponse mediates BCG activity, the authors screened a combinatorial random peptide library on the circulating pool of immunoglobulins (Igs) purified from an index patient after successful BCG immunotherapy to identify the corresponding target antigen(s). An immunogenic peptide motif was selected, isolated, and validated from M. bovis BCG heat-shock protein 65 (HSP-65) as a dominant epitope of the humoral response to treatment. Increasing IgA and IgG anti-HSP-65 titers specifically predicted a positive patient outcome in a cohort of patients with bladder cancer relative to several cohorts of control patients. The current results indicated that antibody production against M. bovis BCG HSP-65 can serve as a serologic marker for the predictive outcome of BCG immunotherapy. Subsequent studies will determine the value of this candidate marker to modify BCG-based treatment for individual patients with bladder cancer.

Persistence of the immune response induced by BCG vaccination

BackgroundAlthough BCG vaccination is recommended in most countries of the world, little is known of the persistence of BCG-induced immune responses. As novel TB vaccines may be given to boost the immunity induced by neonatal BCG vaccination, evidence concerning the persistence of the BCG vaccine-induced response would help inform decisions about when such boosting would be most effective.MethodsA randomised control study of UK adolescents was carried out to investigate persistence of BCG immune responses. Adolescents were tested for interferon-gamma (IFN-γ) response to Mycobacterium tuberculosis purified protein derivative (M.tb PPD) in a whole blood assay before, 3 months, 12 months (n = 148) and 3 years (n = 19) after receiving teenage BCG vaccination or 14 years after receiving infant BCG vaccination (n = 16).ResultsA gradual reduction in magnitude of response was evident from 3 months to 1 year and from 1 year to 3 years following teenage vaccination, but responses 3 years after vaccination were still on average 6 times higher than before vaccination among vaccinees. Some individuals (11/86; 13%) failed to make a detectable antigen-specific response three months after vaccination, or lost the response after 1 (11/86; 13%) or 3 (3/19; 16%) years. IFN-γ response to Ag85 was measured in a subgroup of adolescents and appeared to be better maintained with no decline from 3 to 12 months. A smaller group of adolescents were tested 14 years after receiving infant BCG vaccination and 13/16 (81%) made a detectable IFN-γ response to M.tb PPD 14 years after infant vaccination as compared to 6/16 (38%) matched unvaccinated controls (p = 0.012); teenagers vaccinated in infancy were 19 times more likely to make an IFN-γ response of > 500 pg/ml than unvaccinated teenagers.ConclusionBCG vaccination in infancy and adolescence induces immunological memory to mycobacterial antigens that is still present and measurable for at least 14 years in the majority of vaccinees, although the magnitude of the peripheral blood response wanes from 3 months to 12 months and from 12 months to 3 years post vaccination. The data presented here suggest that because of such waning in the response there may be scope for boosting anti-tuberculous immunity in BCG vaccinated children anytime from 3 months post-vaccination. This supports the prime boost strategies being employed for some new TB vaccines currently under development.