RESUMO
Multimodal T cell profiling can enable more precise characterization of elusive cell states underlying disease. Here, we integrated single-cell RNA and surface protein data from 500,089 memory T cells to define 31 cell states from 259 individuals in a Peruvian tuberculosis (TB) progression cohort. At immune steady state >4 years after infection and disease resolution, we found that, after accounting for significant effects of age, sex, season and genetic ancestry on T cell composition, a polyfunctional type 17 helper T (TH17) cell-like effector state was reduced in abundance and function in individuals who previously progressed from Mycobacterium tuberculosis (M.tb) infection to active TB disease. These cells are capable of responding to M.tb peptides. Deconvoluting this state-uniquely identifiable with multimodal analysis-from public data demonstrated that its depletion may precede and persist beyond active disease. Our study demonstrates the power of integrative multimodal single-cell profiling to define cell states relevant to disease and other traits.
Assuntos
Memória Imunológica , Mycobacterium tuberculosis/imunologia , Células Th17/imunologia , Tuberculose Pulmonar/imunologia , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Criança , Progressão da Doença , Feminino , Seguimentos , Predisposição Genética para Doença , Técnicas de Genotipagem , Humanos , Masculino , Pessoa de Meia-Idade , Mycobacterium tuberculosis/isolamento & purificação , Peru , RNA-Seq , Fatores Sexuais , Análise de Célula Única , Fatores Socioeconômicos , Tuberculose Pulmonar/sangue , Tuberculose Pulmonar/genética , Tuberculose Pulmonar/microbiologia , Adulto JovemRESUMO
Mammalian genomes encode up to four Notch receptors (Notch1-4) and five Notch ligands of the DSL (Delta/Serrate/Lag-2) family, and Notch signaling controls a wide spectrum of developmental processes. Intrathymic Notch1 signaling is essential for several distinct aspects of early T cell development. Notch signaling has also been implicated as a key regulator of peripheral T cell activation and effector cell differentiation, but its functions in these processes remain poorly understood. Notch signaling is dispensable for B cell development in the bone marrow, but it is required to generate the innate-like marginal zone B cell subset in the spleen and may also regulate plasma cell functions. Modification of Notch receptors by fringe glycosyltransferases influences many Notch-dependent aspects of hematopoiesis by altering Notch responsiveness to Delta-like versus Jagged DSL ligands. Here we review recent advances in general aspects of Notch signaling, as well as studies probing Notch functions in these immunological processes.
Assuntos
Sistema Imunitário/imunologia , Sistema Imunitário/metabolismo , Receptores Notch/imunologia , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Diferenciação Celular , Humanos , Sistema Imunitário/citologia , Ativação Linfocitária , Polissacarídeos/imunologiaRESUMO
The hallmark function of αß T cell antigen receptors (TCRs) involves the highly specific co-recognition of a major histocompatibility complex molecule and its carried peptide. However, the molecular basis of the interactions of TCRs with the lipid antigen-presenting molecule CD1c is unknown. We identified frequent staining of human T cells with CD1c tetramers across numerous subjects. Whereas TCRs typically show high specificity for antigen, both tetramer binding and autoreactivity occurred with CD1c in complex with numerous, chemically diverse self lipids. Such extreme polyspecificity was attributable to binding of the TCR over the closed surface of CD1c, with the TCR covering the portal where lipids normally protrude. The TCR essentially failed to contact lipids because they were fully seated within CD1c. These data demonstrate the sequestration of lipids within CD1c as a mechanism of autoreactivity and point to small lipid size as a determinant of autoreactive T cell responses.
Assuntos
Antígenos CD1/imunologia , Autoantígenos/imunologia , Autoimunidade/imunologia , Glicoproteínas/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T/imunologia , Apresentação de Antígeno/imunologia , Humanos , Lipídeos/imunologia , Ativação Linfocitária/imunologiaRESUMO
Non-coding genetic variants may cause disease by modulating gene expression. However, identifying these expression quantitative trait loci (eQTLs) is complicated by differences in gene regulation across fluid functional cell states within cell types. These states-for example, neurotransmitter-driven programs in astrocytes or perivascular fibroblast differentiation-are obscured in eQTL studies that aggregate cells1,2. Here we modelled eQTLs at single-cell resolution in one complex cell type: memory T cells. Using more than 500,000 unstimulated memory T cells from 259 Peruvian individuals, we show that around one-third of 6,511 cis-eQTLs had effects that were mediated by continuous multimodally defined cell states, such as cytotoxicity and regulatory capacity. In some loci, independent eQTL variants had opposing cell-state relationships. Autoimmune variants were enriched in cell-state-dependent eQTLs, including risk variants for rheumatoid arthritis near ORMDL3 and CTLA4; this indicates that cell-state context is crucial to understanding potential eQTL pathogenicity. Moreover, continuous cell states explained more variation in eQTLs than did conventional discrete categories, such as CD4+ versus CD8+, suggesting that modelling eQTLs and cell states at single-cell resolution can expand insight into gene regulation in functionally heterogeneous cell types.
Assuntos
Predisposição Genética para Doença , Células T de Memória , Locos de Características Quantitativas , Regulação da Expressão Gênica , Predisposição Genética para Doença/genética , Humanos , Células T de Memória/imunologia , Células T de Memória/metabolismo , Peru , Locos de Características Quantitativas/genéticaRESUMO
A quarter of humanity is estimated to have been exposed to Mycobacterium tuberculosis (Mtb) with a 5-10% risk of developing tuberculosis (TB) disease. Variability in responses to Mtb infection could be due to host or pathogen heterogeneity. Here, we focused on host genetic variation in a Peruvian population and its associations with gene regulation in monocyte-derived macrophages and dendritic cells (DCs). We recruited former household contacts of TB patients who previously progressed to TB (cases, n = 63) or did not progress to TB (controls, n = 63). Transcriptomic profiling of monocyte-derived DCs and macrophages measured the impact of genetic variants on gene expression by identifying expression quantitative trait loci (eQTL). We identified 330 and 257 eQTL genes in DCs and macrophages (False Discovery Rate (FDR) < 0.05), respectively. Four genes in DCs showed interaction between eQTL variants and TB progression status. The top eQTL interaction for a protein-coding gene was with FAH, the gene encoding fumarylacetoacetate hydrolase, which mediates the last step in mammalian tyrosine catabolism. FAH expression was associated with genetic regulatory variation in cases but not controls. Using public transcriptomic and epigenomic data of Mtb-infected monocyte-derived dendritic cells, we found that Mtb infection results in FAH downregulation and DNA methylation changes in the locus. Overall, this study demonstrates effects of genetic variation on gene expression levels that are dependent on history of infectious disease and highlights a candidate pathogenic mechanism through pathogen-response genes. Furthermore, our results point to tyrosine metabolism and related candidate TB progression pathways for further investigation.
Assuntos
Células Dendríticas , Macrófagos , Mycobacterium tuberculosis , Locos de Características Quantitativas , Tuberculose , Humanos , Peru , Tuberculose/genética , Tuberculose/microbiologia , Macrófagos/metabolismo , Macrófagos/microbiologia , Mycobacterium tuberculosis/patogenicidade , Mycobacterium tuberculosis/genética , Feminino , Células Dendríticas/metabolismo , Masculino , Adulto , Predisposição Genética para Doença , Variação Genética , Regulação da Expressão Gênica , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Perfilação da Expressão GênicaRESUMO
AIM: Tirzepatide is a glucose-dependent insulinotropic polypeptide and glucagon-like peptide-1 (GLP-1) dual receptor agonist (RA) that reduces glycated haemoglobin (HbA1c) and weight in patients with type 2 diabetes. We assessed the effectiveness of tirzepatide in real-world use in an Arab population. METHODS: Review of clinical data from a specialist outpatient diabetes centre; study time points and outcome measures were pre-specified. RESULTS: Tirzepatide was initiated in 8945 patients between 24 October 2022 and 31 December 2023. Of these, 3686 individuals reached 40 weeks of follow-up. At initiation, the mean ± SD age was 54.1 ± 11.5 years, body mass index 34.6 ± 6.0 kg/m2 and HbA1c 7.3 ± 1.5% (56 ± 17 mmol/mol); 2296 (62%) were switched to tirzepatide from another GLP-RA and 317 (8.6%) reported previous bariatric surgery. The maximum dose dispensed was ≥12.5 mg/week in 1087, 7.5-10.0 mg/week in 1688 and 2.5-5.0 mg/week in 911. The mean 40-week reduction in HbA1c was 0.6 ± 1.2% (8 ± 13 mmol/mol) and the reduction in weight was 4.5 ± 6.9 kg (4.8 ± 7.3%). GLP-RA-naïve patients experienced a significantly greater reduction in HbA1c [1.0 ± 1.3% (11 ± 14 mmol/mol) versus 0.5 ± 1.2% (6 ± 13 mmol/mol), p < .0001] and weight (7.2 ± 8.6 vs. 4.2 ± 6.6 kg, p < .0001) compared with previously exposed individuals. Post-metabolic bariatric surgery patients lost significantly more weight (7.8 ± 9.4 vs. 4.5 ± 7.0 kg, p < .0001). Improvements in blood pressure, lipid profile, and liver transaminases were noted at 40 weeks. Tirzepatide was well tolerated, with 288 (7.8%) of patients discontinuing treatment because of adverse effects, predominantly gastrointestinal. CONCLUSION: In real-world use, tirzepatide significantly reduced HbA1c levels and weight and was well tolerated. Previous GLP-RA use was associated with significantly lesser HbA1c and weight reduction, and previous metabolic bariatric surgery was associated with greater weight loss.
Assuntos
Árabes , Diabetes Mellitus Tipo 2 , Hemoglobinas Glicadas , Hipoglicemiantes , Humanos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/sangue , Pessoa de Meia-Idade , Masculino , Feminino , Hipoglicemiantes/uso terapêutico , Adulto , Hemoglobinas Glicadas/análise , Hemoglobinas Glicadas/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/agonistas , Redução de Peso/efeitos dos fármacos , Idoso , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Resultado do Tratamento , Estudos Retrospectivos , Receptor do Peptídeo Semelhante ao Glucagon 2 , Polipeptídeo Inibidor GástricoRESUMO
Mucosal-associated invariant T (MAIT) cells are innate-like T cells that are highly abundant in human blood and tissues. Most MAIT cells have an invariant TCRα-chain that uses T cell receptor α-variable 1-2 (TRAV1-2) joined to TRAJ33/20/12 and recognizes metabolites from bacterial riboflavin synthesis bound to the Ag-presenting molecule MHC class I related (MR1). Our attempts to identify alternative MR1-presented Ags led to the discovery of rare MR1-restricted T cells with non-TRAV1-2 TCRs. Because altered Ag specificity likely alters affinity for the most potent known Ag, 5-(2-oxopropylideneamino)-6-d-ribitylaminouracil (5-OP-RU), we performed bulk TCRα- and TCRß-chain sequencing and single-cell-based paired TCR sequencing on T cells that bound the MR1-5-OP-RU tetramer with differing intensities. Bulk sequencing showed that use of V genes other than TRAV1-2 was enriched among MR1-5-OP-RU tetramerlow cells. Although we initially interpreted these as diverse MR1-restricted TCRs, single-cell TCR sequencing revealed that cells expressing atypical TCRα-chains also coexpressed an invariant MAIT TCRα-chain. Transfection of each non-TRAV1-2 TCRα-chain with the TCRß-chain from the same cell demonstrated that the non-TRAV1-2 TCR did not bind the MR1-5-OP-RU tetramer. Thus, dual TCRα-chain expression in human T cells and competition for the endogenous ß-chain explains the existence of some MR1-5-OP-RU tetramerlow T cells. The discovery of simultaneous expression of canonical and noncanonical TCRs on the same T cell means that claims of roles for non-TRAV1-2 TCR in MR1 response must be validated by TCR transfer-based confirmation of Ag specificity.
Assuntos
Células T Invariantes Associadas à Mucosa , Antígenos de Histocompatibilidade Classe I/genética , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/metabolismo , Mucosa , Receptores de Antígenos de Linfócitos T/metabolismoRESUMO
Whereas proteolytic cleavage is crucial for peptide presentation by classical major histocompatibility complex (MHC) proteins to T cells, glycolipids presented by CD1 molecules are typically presented in an unmodified form. However, the mycobacterial lipid antigen mannosyl-ß1-phosphomycoketide (MPM) may be processed through hydrolysis in antigen presenting cells, forming mannose and phosphomycoketide (PM). To further test the hypothesis that some lipid antigens are processed, and to generate antigens that lead to defined epitopes for future tuberculosis vaccines or diagnostic tests, we aimed to create hydrolysis-resistant MPM variants that retain their antigenicity. Here, we designed and tested three different, versatile synthetic strategies to chemically stabilize MPM analogs. Crystallographic studies of CD1c complexes with these three new MPM analogs showed anchoring of the lipid tail and phosphate group that is highly comparable to nature-identical MPM, with considerable conformational flexibility for the mannose head group. MPM-3, a difluoromethylene-modified version of MPM that is resistant to hydrolysis, showed altered recognition by cells, but not by CD1c proteins, supporting the cellular antigen processing hypothesis. Furthermore, the synthetic analogs elicited T cell responses that were cross-reactive with nature-identical MPM, fulfilling important requirements for future clinical use.
Assuntos
Antígenos de Bactérias/química , Antígenos CD1/química , Glicolipídeos/química , Glicoproteínas/química , Mycobacterium tuberculosis/química , Fosfolipídeos/química , Linfócitos T/química , Antígenos de Bactérias/imunologia , Antígenos CD1/imunologia , Linhagem Celular Transformada , Cristalografia por Raios X , Glicolipídeos/imunologia , Glicoproteínas/imunologia , Humanos , Mycobacterium tuberculosis/imunologia , Fosfolipídeos/imunologia , Linfócitos T/imunologiaRESUMO
AIM: To evaluate the effectiveness and safety of bariatric surgery in patients with coexisting type 1 diabetes and obesity who choose to undergo bariatric surgery for weight management. MATERIALS AND METHODS: We conducted a retrospective, longitudinal review of the clinical data of patients with type 1 diabetes, followed up at our centre after bariatric surgery had been performed elsewhere. RESULTS: Sixty-one patients were included, of whom 51% were women, and 57 (93%) were Emirati Arab, three (5%) were other Arab, and one (2%) was White in ethnic origin. The mean age at surgery was 31 years. A total of 42 patients (69%) underwent sleeve gastrectomy, 17 (28%) gastric bypass, and two (3%) gastric banding. In 48 patients with complete follow-up data, improvements were observed at 12 months in the median (interquartile range [IQR]) values for body mass index (BMI; 38.5 [34.9-40.9] to 26.1 [24.2-29.6] kg/m2 , P <0.001), glycated haemoglobin concentration (8.6 [7.8-9.2]% to 7.8 [7.2-8.5]%; P <0.001), daily insulin dose (1.0 [0.7-1.2] to 0.8 [0.6-0.9] units/kg/d; P <0.001), systolic blood pressure (127 [116-136] to 116 [110-120] mmHg; P <0.001), total cholesterol: high-density lipoprotein cholesterol ratio (3.4 [2.9-4.3] to 3.0 [2.6-3.5]; P <0.001), and albuminuria. In 32 patients followed up at 3 years, the median (IQR) reduction in BMI was 10.4 (5.9-11.7) kg/m2 , 50% reduced or stopped antihypertensive medication and 58% reduced or stopped lipid-modifying medication. Three patients each reported a single episode of diabetic ketoacidosis; in one case this was due to cessation of insulin treatment. CONCLUSIONS: In our cohort of patients with obesity and type 1 diabetes, bariatric surgery led to significant improvements in weight and cardiometabolic variables, with modest improvements in glycaemia. Few adverse events were reported.
Assuntos
Cirurgia Bariátrica , Diabetes Mellitus Tipo 1 , Derivação Gástrica , Obesidade Mórbida , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/epidemiologia , Feminino , Gastrectomia , Humanos , Obesidade/complicações , Obesidade/cirurgia , Obesidade Mórbida/complicações , Obesidade Mórbida/epidemiologia , Obesidade Mórbida/cirurgia , Estudos Retrospectivos , Resultado do TratamentoRESUMO
Tuberculosis (TB) is the leading cause of mortality from a single infectious agent, Mycobacterium tuberculosis Relevant immune targets of the partially efficacious TB vaccine bacille Calmette-Guérin (BCG) remain poorly defined. Mucosal-associated invariant T (MAIT) cells are MHC-related protein 1 (MR1)-restricted T cells, which are reactive against M. tuberculosis, and underexplored as potential TB vaccine targets. We sought to determine whether BCG vaccination activated mycobacteria-specific MAIT cell responses in humans. We analyzed whole blood samples from M. tuberculosis-infected South African adults who were revaccinated with BCG after a six-month course of isoniazid preventative therapy. In vitro BCG stimulation potently induced IFN-γ expression by phenotypic (CD8+CD26+CD161+) MAIT cells, which constituted the majority (75%) of BCG-reactive IFN-γ-producing CD8+ T cells. BCG revaccination transiently expanded peripheral blood frequencies of BCG-reactive IFN-γ+ MAIT cells, which returned to baseline frequencies a year following vaccination. In another cohort of healthy adults who received BCG at birth, 53% of mycobacteria-reactive-activated CD8 T cells expressed CDR3α TCRs, previously reported as MAIT TCRs, expressing the canonical TRAV1-2-TRAJ33 MAIT TCRα rearrangement. CD26 and CD161 coexpression correlated with TRAV1-2+CD161+ phenotype more accurately in CD8+ than CD4-CD8- MAIT cells. Interestingly, BCG-induced IFN-γ expression by MAIT cells in vitro was mediated by the innate cytokines IL-12 and IL-18 more than MR1-induced TCR signaling, suggesting TCR-independent activation. Collectively, the data suggest that activation of blood MAIT cells by innate inflammatory cytokines is a major mechanism of responsiveness to vaccination with whole cell vaccines against TB or in vitro stimulation with mycobacteria (Clinical trial registration: NCT01119521).
Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Menor/imunologia , Células T Invariantes Associadas à Mucosa/imunologia , Mycobacterium tuberculosis/imunologia , Adolescente , Criança , Estudos de Coortes , Citocinas/imunologia , Humanos , Receptores de Antígenos de Linfócitos T/imunologiaRESUMO
High-throughput TCR sequencing allows interrogation of the human TCR repertoire, potentially connecting TCR sequences to antigenic targets. Unlike the highly polymorphic MHC proteins, monomorphic Ag-presenting molecules such as MR1, CD1d, and CD1b present Ags to T cells with species-wide TCR motifs. CD1b tetramer studies and a survey of the 27 published CD1b-restricted TCRs demonstrated a TCR motif in humans defined by the TCR ß-chain variable gene 4-1 (TRBV4-1) region. Unexpectedly, TRBV4-1 was involved in recognition of CD1b regardless of the chemical class of the carried lipid. Crystal structures of two CD1b-specific TRBV4-1+ TCRs show that germline-encoded residues in CDR1 and CDR3 regions of TRBV4-1-encoded sequences interact with each other and consolidate the surface of the TCR. Mutational studies identified a key positively charged residue in TRBV4-1 and a key negatively charged residue in CD1b that is shared with CD1c, which is also recognized by TRBV4-1 TCRs. These data show that one TCR V region can mediate a mechanism of recognition of two related monomorphic Ag-presenting molecules that does not rely on a defined lipid Ag.
Assuntos
Motivos de Aminoácidos , Antígenos CD1d/química , Antígenos CD1d/metabolismo , Sítios de Ligação , Receptores de Antígenos de Linfócitos T alfa-beta/química , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Apresentação de Antígeno , Sequência Conservada , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/metabolismo , Rearranjo Gênico , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunofenotipagem , Lipídeos/química , Modelos Moleculares , Mutação , Ligação Proteica , Conformação Proteica , Multimerização Proteica , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Relação Estrutura-Atividade , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
BACKGROUND: In human blood, mucosal-associated invariant T (MAIT) cells are abundant T cells that recognize antigens presented on non-polymorphic major histocompatibility complex-related 1 (MR1) molecules. The MAIT cells are activated by mycobacteria, and prior human studies indicate that blood frequencies of MAIT cells, defined by cell surface markers, decline during tuberculosis (TB) disease, consistent with redistribution to the lungs. METHODS: We tested whether frequencies of blood MAIT cells were altered in patients with TB disease relative to healthy Mycobacterium tuberculosis-exposed controls from Peru and South Africa. We quantified their frequencies using MR1 tetramers loaded with 5-(2-oxopropylideneamino)-6-D-ribitylaminouracil. RESULTS: Unlike findings from prior studies, frequencies of blood MAIT cells were similar among patients with TB disease and latent and uninfected controls. In both cohorts, frequencies of MAIT cells defined by MR1-tetramer staining and coexpression of CD161 and the T-cell receptor alpha variable gene TRAV1-2 were strongly correlated. Disease severity captured by body mass index or TB disease transcriptional signatures did not correlate with MAIT cell frequencies in patients with TB. CONCLUSIONS: Major histocompatibility complex (MHC)-related 1-restrictied MAIT cells are detected at similar levels with tetramers or surface markers. Unlike MHC-restricted T cells, blood frequencies of MAIT cells are poor correlates of TB disease but may play a role in pathophysiology.
Assuntos
Células T Invariantes Associadas à Mucosa/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculose/epidemiologia , Tuberculose/imunologia , Adulto , Biomarcadores , Estudos de Casos e Controles , Feminino , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Células T Invariantes Associadas à Mucosa/metabolismo , Prevalência , Vigilância em Saúde Pública , Medição de Risco , Fatores de Risco , Tuberculose/microbiologiaRESUMO
RATIONALE: Global tuberculosis (TB) control requires effective vaccines in TB-endemic countries, where most adults are infected with Mycobacterium tuberculosis (M.tb). OBJECTIVES: We sought to define optimal dose and schedule of H56:IC31, an experimental TB vaccine comprising Ag85B, ESAT-6, and Rv2660c, for M.tb-infected and M.tb-uninfected adults. METHODS: We enrolled 98 healthy, HIV-uninfected, bacillus Calmette-Guérin-vaccinated, South African adults. M.tb infection was defined by QuantiFERON-TB (QFT) assay. QFT-negative participants received two vaccinations of different concentrations of H56 in 500 nmol of IC31 to enable dose selection for further vaccine development. Subsequently, QFT-positive and QFT-negative participants were randomized to receive two or three vaccinations to compare potential schedules. Participants were followed for safety and immunogenicity for 292 days. MEASUREMENTS AND MAIN RESULTS: H56:IC31 showed acceptable reactogenicity profiles irrespective of dose, number of vaccinations, or M.tb infection. No vaccine-related severe or serious adverse events were observed. The three H56 concentrations tested induced equivalent frequencies and functional profiles of antigen-specific CD4 T cells. ESAT-6 was only immunogenic in QFT-negative participants who received three vaccinations. CONCLUSIONS: Two or three H56:IC31 vaccinations at the lowest dose induced durable antigen-specific CD4 T-cell responses with acceptable safety and tolerability profiles in M.tb-infected and M.tb-uninfected adults. Additional studies should validate applicability of vaccine doses and regimens to both QFT-positive and QFT-negative individuals. Clinical trial registered with www.clinicaltrials.gov (NCT01865487).
Assuntos
Vacinas contra a Tuberculose/uso terapêutico , Tuberculose/prevenção & controle , Aciltransferases/imunologia , Aciltransferases/uso terapêutico , Adolescente , Adulto , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/uso terapêutico , Proteínas de Bactérias/imunologia , Proteínas de Bactérias/uso terapêutico , Relação Dose-Resposta a Droga , Método Duplo-Cego , Combinação de Medicamentos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Oligodesoxirribonucleotídeos/imunologia , Oligodesoxirribonucleotídeos/uso terapêutico , Oligopeptídeos/imunologia , Oligopeptídeos/uso terapêutico , África do Sul , Resultado do Tratamento , Tuberculose/imunologia , Vacinas contra a Tuberculose/imunologia , Adulto JovemRESUMO
BACKGROUND: A nonsputum blood test capable of predicting progression of healthy individuals to active tuberculosis (TB) before clinical symptoms manifest would allow targeted treatment to curb transmission. We aimed to develop a proteomic biomarker of risk of TB progression for ultimate translation into a point-of-care diagnostic. METHODS AND FINDINGS: Proteomic TB risk signatures were discovered in a longitudinal cohort of 6,363 Mycobacterium tuberculosis-infected, HIV-negative South African adolescents aged 12-18 years (68% female) who participated in the Adolescent Cohort Study (ACS) between July 6, 2005 and April 23, 2007, through either active (every 6 months) or passive follow-up over 2 years. Forty-six individuals developed microbiologically confirmed TB disease within 2 years of follow-up and were selected as progressors; 106 nonprogressors, who remained healthy, were matched to progressors. Over 3,000 human proteins were quantified in plasma with a highly multiplexed proteomic assay (SOMAscan). Three hundred sixty-one proteins of differential abundance between progressors and nonprogressors were identified. A 5-protein signature, TB Risk Model 5 (TRM5), was discovered in the ACS training set and verified by blind prediction in the ACS test set. Poor performance on samples 13-24 months before TB diagnosis motivated discovery of a second 3-protein signature, 3-protein pair-ratio (3PR) developed using an orthogonal strategy on the full ACS subcohort. Prognostic performance of both signatures was validated in an independent cohort of 1,948 HIV-negative household TB contacts from The Gambia (aged 15-60 years, 66% female), longitudinally followed up for 2 years between March 5, 2007 and October 21, 2010, sampled at baseline, month 6, and month 18. Amongst these contacts, 34 individuals progressed to microbiologically confirmed TB disease and were included as progressors, and 115 nonprogressors were included as controls. Prognostic performance of the TRM5 signature in the ACS training set was excellent within 6 months of TB diagnosis (area under the receiver operating characteristic curve [AUC] 0.96 [95% confidence interval, 0.93-0.99]) and 6-12 months (AUC 0.76 [0.65-0.87]) before TB diagnosis. TRM5 validated with an AUC of 0.66 (0.56-0.75) within 1 year of TB diagnosis in the Gambian validation cohort. The 3PR signature yielded an AUC of 0.89 (0.84-0.95) within 6 months of TB diagnosis and 0.72 (0.64-0.81) 7-12 months before TB diagnosis in the entire South African discovery cohort and validated with an AUC of 0.65 (0.55-0.75) within 1 year of TB diagnosis in the Gambian validation cohort. Signature validation may have been limited by a systematic shift in signal magnitudes generated by differences between the validation assay when compared to the discovery assay. Further validation, especially in cohorts from non-African countries, is necessary to determine how generalizable signature performance is. CONCLUSIONS: Both proteomic TB risk signatures predicted progression to incident TB within a year of diagnosis. To our knowledge, these are the first validated prognostic proteomic signatures. Neither meet the minimum criteria as defined in the WHO Target Product Profile for a progression test. More work is required to develop such a test for practical identification of individuals for investigation of incipient, subclinical, or active TB disease for appropriate treatment and care.
Assuntos
Biomarcadores/sangue , Proteoma/análise , Tuberculose/diagnóstico , Adolescente , Biomarcadores/análise , Biomarcadores/metabolismo , Criança , Estudos de Coortes , Testes Diagnósticos de Rotina/métodos , Progressão da Doença , Feminino , Humanos , Estudos Longitudinais , Masculino , Testes Imediatos , Prognóstico , Estudos Prospectivos , Proteoma/metabolismo , Proteômica , Tuberculose/sangue , Tuberculose/patologiaRESUMO
[This corrects the article DOI: 10.1371/journal.pmed.1002781.].
RESUMO
Our understanding of mechanisms underlying progression from Mycobacterium tuberculosis infection to pulmonary tuberculosis disease in humans remains limited. To define such mechanisms, we followed M. tuberculosis-infected adolescents longitudinally. Blood samples from forty-four adolescents who ultimately developed tuberculosis disease ("progressors") were compared with those from 106 matched controls, who remained healthy during two years of follow up. We performed longitudinal whole blood transcriptomic analyses by RNA sequencing and plasma proteome analyses using multiplexed slow off-rate modified DNA aptamers. Tuberculosis progression was associated with sequential modulation of immunological processes. Type I/II interferon signalling and complement cascade were elevated 18 months before tuberculosis disease diagnosis, while changes in myeloid inflammation, lymphoid, monocyte and neutrophil gene modules occurred more proximally to tuberculosis disease. Analysis of gene expression in purified T cells also revealed early suppression of Th17 responses in progressors, relative to M. tuberculosis-infected controls. This was confirmed in an independent adult cohort who received BCG re-vaccination; transcript expression of interferon response genes in blood prior to BCG administration was associated with suppression of IL-17 expression by BCG-specific CD4 T cells 3 weeks post-vaccination. Our findings provide a timeline to the different immunological stages of disease progression which comprise sequential inflammatory dynamics and immune alterations that precede disease manifestations and diagnosis of tuberculosis disease. These findings have important implications for developing diagnostics, vaccination and host-directed therapies for tuberculosis. TRIAL REGISTRATION: Clincialtrials.gov, NCT01119521.
Assuntos
Mycobacterium tuberculosis , Linfócitos T/imunologia , Tuberculose/microbiologia , Tuberculose/terapia , Adolescente , Criança , Progressão da Doença , Humanos , Inflamação/complicações , Inflamação/imunologia , Inflamação/terapia , Vacinas/uso terapêuticoRESUMO
Rationale: Contacts of patients with tuberculosis (TB) constitute an important target population for preventive measures because they are at high risk of infection with Mycobacterium tuberculosis and progression to disease.Objectives: We investigated biosignatures with predictive ability for incident TB.Methods: In a case-control study nested within the Grand Challenges 6-74 longitudinal HIV-negative African cohort of exposed household contacts, we employed RNA sequencing, PCR, and the pair ratio algorithm in a training/test set approach. Overall, 79 progressors who developed TB between 3 and 24 months after diagnosis of index case and 328 matched nonprogressors who remained healthy during 24 months of follow-up were investigated.Measurements and Main Results: A four-transcript signature derived from samples in a South African and Gambian training set predicted progression up to two years before onset of disease in blinded test set samples from South Africa, the Gambia, and Ethiopia with little population-associated variability, and it was also validated in an external cohort of South African adolescents with latent M. tuberculosis infection. By contrast, published diagnostic or prognostic TB signatures were predicted in samples from some but not all three countries, indicating site-specific variability. Post hoc meta-analysis identified a single gene pair, C1QC/TRAV27 (complement C1q C-chain / T-cell receptor-α variable gene 27) that would consistently predict TB progression in household contacts from multiple African sites but not in infected adolescents without known recent exposure events.Conclusions: Collectively, we developed a simple whole blood-based PCR test to predict TB in recently exposed household contacts from diverse African populations. This test has potential for implementation in national TB contact investigation programs.
RESUMO
One third of the global population is estimated to be latently infected with Mycobacterium tuberculosis We performed a phase I randomized controlled trial of isoniazid preventive therapy (IPT) before revaccination with bacillus Calmette-Guérin (BCG) in healthy, tuberculin skin test-positive (≥15-mm induration), HIV-negative South African adults. We hypothesized that preclearance of latent bacilli with IPT modulates BCG immunogenicity following revaccination. Frequencies and coexpression of IFN-γ, TNF-α, IL-2, IL-17, and/or IL-22 in CD4 T cells and IFN-γ-expressing CD8 T, γδ T, CD3(+)CD56(+) NKT-like, and NK cells in response to BCG were measured using whole blood intracellular cytokine staining and flow cytometry. We analyzed 72 participants who were revaccinated with BCG after IPT (n = 33) or without prior IPT (n = 39). IPT had little effect on frequencies or cytokine coexpression patterns of M. tuberculosis- or BCG-specific responses. Revaccination transiently boosted BCG-specific Th1 cytokine-expressing CD4, CD8, and γδ T cells. Despite high frequencies of IFN-γ-expressing BCG-reactive CD3(+)CD56(+) NKT-like cells and CD3(-)CD56(dim) and CD3(-)CD56(hi) NK cells at baseline, BCG revaccination boosted these responses, which remained elevated up to 1 y after revaccination. Such BCG-reactive memory NK cells were induced by BCG vaccination in infants, whereas in vitro IFN-γ expression by NK cells upon BCG stimulation was dependent on IL-12 and IL-18. Our data suggest that isoniazid preclearance of M. tuberculosis bacilli has little effect on the magnitude, persistence, or functional attributes of lymphocyte responses boosted by BCG revaccination. Our study highlights the surprising durability of BCG-boosted memory NKT-like and NK cells expressing antimycobacterial effector molecules, which may be novel targets for tuberculosis vaccines.
Assuntos
Antituberculosos/administração & dosagem , Vacina BCG/imunologia , Imunização Secundária/métodos , Isoniazida/administração & dosagem , Células Matadoras Naturais/imunologia , Tuberculose Latente/prevenção & controle , Adolescente , Adulto , Vacina BCG/administração & dosagem , Feminino , Citometria de Fluxo , Humanos , Tuberculose Latente/imunologia , Masculino , Adulto JovemRESUMO
BACKGROUND: Identification of blood biomarkers that prospectively predict progression of Mycobacterium tuberculosis infection to tuberculosis disease might lead to interventions that combat the tuberculosis epidemic. We aimed to assess whether global gene expression measured in whole blood of healthy people allowed identification of prospective signatures of risk of active tuberculosis disease. METHODS: In this prospective cohort study, we followed up healthy, South African adolescents aged 12-18 years from the adolescent cohort study (ACS) who were infected with M tuberculosis for 2 years. We collected blood samples from study participants every 6 months and monitored the adolescents for progression to tuberculosis disease. A prospective signature of risk was derived from whole blood RNA sequencing data by comparing participants who developed active tuberculosis disease (progressors) with those who remained healthy (matched controls). After adaptation to multiplex quantitative real-time PCR (qRT-PCR), the signature was used to predict tuberculosis disease in untouched adolescent samples and in samples from independent cohorts of South African and Gambian adult progressors and controls. Participants of the independent cohorts were household contacts of adults with active pulmonary tuberculosis disease. FINDINGS: Between July 6, 2005, and April 23, 2007, we enrolled 6363 participants from the ACS study and 4466 from independent South African and Gambian cohorts. 46 progressors and 107 matched controls were identified in the ACS cohort. A 16 gene signature of risk was identified. The signature predicted tuberculosis progression with a sensitivity of 66·1% (95% CI 63·2-68·9) and a specificity of 80·6% (79·2-82·0) in the 12 months preceding tuberculosis diagnosis. The risk signature was validated in an untouched group of adolescents (p=0·018 for RNA sequencing and p=0·0095 for qRT-PCR) and in the independent South African and Gambian cohorts (p values <0·0001 by qRT-PCR) with a sensitivity of 53·7% (42·6-64·3) and a specificity of 82·8% (76·7-86) in the 12 months preceding tuberculosis. INTERPRETATION: The whole blood tuberculosis risk signature prospectively identified people at risk of developing active tuberculosis, opening the possibility for targeted intervention to prevent the disease. FUNDING: Bill & Melinda Gates Foundation, the National Institutes of Health, Aeras, the European Union, and the South African Medical Research Council.