Tuberculosis alters immune-metabolic pathways resulting in perturbed IL-1 responses

Tuberculosis (TB) remains a major public health problem and we lack a comprehensive understanding of how Mycobacterium tuberculosis (M. tb) infection impacts host immune responses. We compared the induced immune response to TB antigen, BCG and IL-1β stimulation between latently M. tb infected individuals (LTBI) and active TB patients. This revealed distinct responses between TB/LTBI at transcriptomic, proteomic and metabolomic levels. At baseline, we identified a novel immune-metabolic association between pregnane steroids, the PPARγ pathway and elevated plasma IL-1ra in TB. We observed dysregulated IL-1 responses after BCG stimulation in TB patients, with elevated IL-1ra responses being explained by upstream TNF differences. Additionally, distinct secretion of IL-1α/IL-1β in LTBI/TB after BCG stimulation was associated with downstream differences in granzyme mediated cleavage. Finally, IL-1β driven signalling was dramatically perturbed in TB disease but was completely restored after successful treatment. This study improves our knowledge of how immune responses are altered during TB disease, and may support the design of improved preventive and therapeutic tools, including host-directed strategies

Live-attenuated Mycobacterium tuberculosis vaccine MTBVAC versus BCG in adults and neonates: a randomised controlled, double-blind dose-escalation trial

Background: Infants are a key target population for new tuberculosis vaccines. We assessed the safety and immunogenicity of the live-attenuated Mycobacterium tuberculosis vaccine candidate MTBVAC in adults and infants in a region where transmission of tuberculosis is very high. Methods: We did a randomised, double-blind, BCG-controlled, dose-escalation trial at the South African Tuberculosis Vaccine Initiative site near Cape Town, South Africa. Healthy adult community volunteers who were aged 18–50 years, had received BCG vaccination as infants, were HIV negative, had negative interferon-¿ release assay (IGRA) results, and had no personal history of tuberculosis or current household contact with someone with tuberculosis were enrolled in a safety cohort. Infants born to HIV-negative women with no personal history of tuberculosis or current household contact with a person with tuberculosis and who were 96 h old or younger, generally healthy, and had not yet received routine BCG vaccination were enrolled in a separate infant cohort. Eligible adults were randomly assigned (1:1) to receive either BCG Vaccine SSI (5 × 105 colony forming units [CFU] of Danish strain 1331 in 0·1 mL diluent) or MTBVAC (5 × 105 CFU in 0·1 mL) intradermally in the deltoid region of the arm. After favourable review of 28-day reactogenicity and safety data in the adult cohort, infants were randomly assigned (1:3) to receive either BCG Vaccine SSI (2·5 × 105 CFU in 0·05 mL diluent) or MTBVAC in three sequential cohorts of increasing MTBVAC dose (2·5 × 103 CFU, 2·5 × 104 CFU, and 2·5 × 105 CFU in 0·05 mL) intradermally in the deltoid region of the arm. QuantiFERON-TB Gold In-Tube IGRA was done on days 180 and 360. For both randomisations, a pre-prepared block randomisation schedule was used. Participants (and their parents or guardians in the case of infant participants), investigators, and other clinical and laboratory staff were masked to intervention allocation. The primary outcomes, which were all measured in the infant cohort, were solicited and unsolicited local adverse events and serious adverse events until day 360; non-serious systemic adverse events until day 28 and vaccine-specific CD4 and CD8 T-cell responses on days 7, 28, 70, 180, and 360. Secondary outcomes measured in adults were local injection-site and systemic reactions and haematology and biochemistry at study day 7 and 28. Safety analyses and immunogenicity analyses were done in all participants who received a dose of vaccine. This trial is registered with ClinicalTrials.gov, number NCT02729571. Findings: Between Sept 29, 2015, and Nov 16, 2015, 62 adults were screened and 18 were enrolled and randomly assigned, nine each to the BCG and MTBVAC groups. Between Feb 12, 2016, and Sept 21, 2016, 36 infants were randomly assigned—eight to the BCG group, nine to the 2·5 × 103 CFU MTBVAC group, nine to the 2·5 × 104 CFU group, and ten to the 2·5 × 105 CFU group. Mild injection-site reactions occurred only in infants in the BCG and the 2·5 × 105 CFU MTBVAC group, with no evidence of local or regional injection-site complications. Systemic adverse events were evenly distributed across BCG and MTBVAC dose groups, and were mostly mild in severity. Eight serious adverse events were reported in seven vaccine recipients (one adult MTBVAC recipient, one infant BCG recipient, one infant in the 2·5 × 103 CFU MTBVAC group, two in the 2·5 × 104 CFU MTBVAC group, and two in the 2·5 × 105 CFU MTBVAC group), including one infant in the 2·5 × 103 CFU MTBVAC group treated for unconfirmed tuberculosis and one in the 2·5 × 105 CFU MTBVAC group treated for unlikely tuberculosis. One infant died as a result of possible viral pneumonia. Vaccination with all MTBVAC doses induced durable antigen-specific T-helper-1 cytokine-expressing CD4 cell responses in infants that peaked 70 days after vaccination and were detectable 360 days after vaccination. For the highest MTBVAC dose (ie, 2·5 × 105 CFU), these responses exceeded responses induced by an equivalent dose of the BCG vaccine up to 360 days after vaccination. Dose-related IGRA conversion was noted in three (38%) of eight infants in the 2·5 × 103 CFU MTBVAC group, six (75%) of eight in the 2·5 × 104 CFU MTBVAC group, and seven (78%) of nine in the 2·5 × 105 CFU MTBVAC group at day 180, compared with none of seven infants in the BCG group. By day 360, IGRA reversion had occurred in all three infants (100%) in the 2·5 × 103 CFU MTBVAC group, four (67%) of the six in the 2·5 × 104 CFU MTBVAC group, and three (43%) of the seven in the 2·5 × 105 CFU MTBVAC group. Interpretation: MTBVAC had acceptable reactogenicity, and induced a durable CD4 cell response in infants. The evidence of immunogenicity supports progression of MTBVAC into larger safety and efficacy trials, but also confounds interpretation of tests for M tuberculosis infection, highlighting the need for stringent endpoint definition. Funding: Norwegian Agency for Development Cooperation, TuBerculosis Vaccine Initiative, UK Department for International Development, and Biofabri

T-cell biomarkers for diagnosis of tuberculosis: candidate evaluation by a simple whole blood assay for clinical translation

Conflict of interest: T.J. Scriba reports grants received from BMGF by University of Cape Town, during the conduct of the study.Conflict of interest: E. Nemes received grants from the Bill and Melinda Gates Foundation, during the conduct of the study.International audienceWe compared candidate T-cell-based TB biomarkers and identified HLA-DR on Mtb-specific T-cells as a priority marker http://ow.ly/TyHa30iwVb

Newborn bacille Calmette-Guérin vaccination induces robust infant interferon-γ-expressing natural killer cell responses to mycobacteria

Objectives: The bacille Calmette-Guérin (BCG) vaccine is usually administered at birth to protect against severe forms of tuberculosis in children. BCG also confers some protection against other infections, possibly mediated by innate immune training. We investigated whether newborn BCG vaccination modulates myeloid and natural killer (NK) cell responses to mycobacteria. Methods: BCG vaccination was either administered at birth or delayed to 6 or 10 weeks of age in 130 South African infants. Whole blood was stimulated with BCG and clusters of differentiation (CD)4+ T, myeloid, and NK cell responses were measured by flow cytometry; the levels of secreted cytokines were measured by a multiplex bead array. Results: Newborn BCG vaccination was associated with significantly higher frequencies of BCG-reactive, cytokine-expressing CD4+ T cells, and interferon (IFN)-γ-expressing NK cells than in unvaccinated infants but no differences in cytokine-expressing CD33+ myeloid cells were observed. The induction of BCG-reactive IFN-γ-expressing NK cells was not associated with the markers of NK cell maturation, differentiation, or cytokine receptor expression. BCG-reactive NK cell responses correlated directly with the levels of secreted interleukin (IL)-2 and IFN-γ and the innate pro-inflammatory cytokines IL-6, IL-1β, and tumor necrosis factor (TNF) in BCG-vaccinated infants only. Conclusion: We showed that BCG-reactive IFN-γ-expressing NK cells are strongly induced by BCG vaccination in infants and are likely amplified through bystander cytokines

Tuberculosis alters immune-metabolic pathways resulting in perturbed IL-1 responses

Tuberculosis (TB) remains a major public health problem with host-directed therapeutics offering potential as novel treatment strategies. However, their successful development still requires a comprehensive understanding of how Mycobacterium tuberculosis ( M.tb ) infection impacts immune responses. To address this challenge, we applied standardised immunomonitoring tools to compare TB antigen, BCG and IL-1β induced immune responses between individuals with latent M.tb infection (LTBI) and active TB disease, at diagnosis and after cure. This revealed distinct responses between TB and LTBI groups at transcriptomic, proteomic and metabolomic levels. At baseline, we identified pregnane steroids and the PPARγ pathway as new immune-metabolic drivers of elevated plasma IL-1ra in TB. We also observed dysregulated induced IL-1 responses after BCG stimulation in TB patients. Elevated IL-1 antagonist responses were explained by upstream differences in TNF responses, while for IL-1 agonists it was due to downstream differences in granzyme mediated cleavage. Finally, the immune response to IL-1β driven signalling was also dramatically perturbed in TB disease but was completely restored after successful antibiotic treatment. This systems immunology approach improves our knowledge of how immune responses are altered during TB disease, and may support design of improved diagnostic, prophylactic and therapeutic tools

Immune profiling enables stratification of patients with active TB disease or M. tuberculosis infection

International audienceBackgroundTuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb) infection and is a major public health problem. Clinical challenges include the lack of a blood-based test for active disease. Current blood-based tests, such as QuantiFERON (QFT) do not distinguish active TB disease from asymptomatic Mtb infection.MethodsWe hypothesized that TruCulture, an immunomonitoring method for whole blood stimulation, could discriminate active disease from latent Mtb infection (LTBI). We stimulated whole blood from active TB patients and compared to LTBI donors. Mtb- specific antigens and live bacillus Calmette-Guerin (BCG) were used as stimuli, with direct comparison to QFT. Protein analyses were performed using conventional and digital ELISA, as well as Luminex.ResultsTruCulture showed discrimination of active TB cases from LTBI (p < 0.0001 AUC = 0.81) as compared to QFT (p = 0.45 AUC = 0.56), based on an IFNγ readout after Mtb antigen stimulation. This result was replicated in an independent cohort (AUC = 0.89). In exploratory analyses, TB stratification could be further improved by the Mtb Ag/BCG IFNγ ratio (p < 0.0001 AUC = 0.91). Finally, the combination of digital ELISA and transcriptional analysis showed that LTBI donors with high IFNγ clustered with TB patients, suggesting the possibility to identify sub-clinical disease.ConclusionsTruCulture offers a next-generation solution for whole blood stimulation and immunomonitoring with the possibility to discriminate active and latent infection

Functional, Antigen-Specific Stem Cell Memory (TSCM) CD4+ T Cells Are Induced by Human Mycobacterium tuberculosis Infection

BackgroundMaintenance of long-lasting immunity is thought to depend on stem cell memory T cells (TSCM), which have superior self-renewing capacity, longevity and proliferative potential compared with central memory (TCM) or effector (TEFF) T cells. Our knowledge of TSCM derives primarily from studies of virus-specific CD8+ TSCM. We aimed to determine if infection with Mycobacterium tuberculosis (M. tb), the etiological agent of tuberculosis, generates antigen-specific CD4+ TSCM and to characterize their functional ontology.MethodsWe studied T cell responses to natural M. tb infection in a longitudinal adolescent cohort of recent QuantiFERON-TB Gold (QFT) converters and three cross-sectional QFT+ adult cohorts; and to bacillus Calmette–Guerin (BCG) vaccination in infants. M. tb and/or BCG-specific CD4 T cells were detected by flow cytometry using major histocompatibility complex class II tetramers bearing Ag85, CFP-10, or ESAT-6 peptides, or by intracellular cytokine staining. Transcriptomic analyses of M. tb-specific tetramer+ CD4+ TSCM (CD45RA+ CCR7+ CD27+) were performed by microfluidic qRT-PCR, and functional and phenotypic characteristics were confirmed by measuring expression of chemokine receptors, cytotoxic molecules and cytokines using flow cytometry.ResultsM. tb-specific TSCM were not detected in QFT-negative persons. After QFT conversion frequencies of TSCM increased to measurable levels and remained detectable thereafter, suggesting that primary M. tb infection induces TSCM cells. Gene expression (GE) profiling of tetramer+ TSCM showed that these cells were distinct from bulk CD4+ naïve T cells (TN) and shared features of bulk TSCM and M. tb-specific tetramer+ TCM and TEFF cells. These TSCM were predominantly CD95+ and CXCR3+, markers typical of CD8+ TSCM. Tetramer+ TSCM expressed significantly higher protein levels of CCR5, CCR6, CXCR3, granzyme A, granzyme K, and granulysin than bulk TN and TSCM cells. M. tb-specific TSCM were also functional, producing IL-2, IFN-γ, and TNF-α upon antigen stimulation, and their frequencies correlated positively with long-term BCG-specific CD4+ T cell proliferative potential after infant vaccination.ConclusionHuman infection with M. tb induced distinct, antigen-specific CD4+ TSCM cells endowed with effector functions, including expression of cytotoxic molecules and Th1 cytokines, and displayed chemokine receptor profiles consistent with memory Th1/17 cells. Induction of CD4+ TSCM should be considered for vaccination approaches that aim to generate long-lived memory T cells against M. tb

Tuberculosis alters immune-metabolic pathways resulting in perturbed IL-1 responses

Tuberculosis (TB) remains a major public health problem with host-directed therapeutics offering potential as novel treatment strategies. However, their successful development still requires a comprehensive understanding of how Mycobacterium tuberculosis ( M.tb ) infection impacts immune responses. To address this challenge, we applied standardised immunomonitoring tools to compare TB antigen, BCG and IL-1β induced immune responses between individuals with latent M.tb infection (LTBI) and active TB disease, at diagnosis and after cure. This revealed distinct responses between TB and LTBI groups at transcriptomic, proteomic and metabolomic levels. At baseline, we identified pregnane steroids and the PPARγ pathway as new immune-metabolic drivers of elevated plasma IL-1ra in TB. We also observed dysregulated induced IL-1 responses after BCG stimulation in TB patients. Elevated IL-1 antagonist responses were explained by upstream differences in TNF responses, while for IL-1 agonists it was due to downstream differences in granzyme mediated cleavage. Finally, the immune response to IL-1β driven signalling was also dramatically perturbed in TB disease but was completely restored after successful antibiotic treatment. This systems immunology approach improves our knowledge of how immune responses are altered during TB disease, and may support design of improved diagnostic, prophylactic and therapeutic tools