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

Abstract

T-cell biomarkers for diagnosis of tuberculosis: candidate evaluation by a simple whole blood assay for clinical translation To the Editor: Diagnosis of pulmonary tuberculosis (TB) disease, the most common cause of death due to infection globally [1], depends on direct detection of Mycobacterium tuberculosis, typically from a sputum specimen. However, up to half of individuals with microbiologically proven TB do not have a prolonged productive cough [2], required to produce sputum. To address this challenge, investigators have identified candidate blood-based host biomarkers for TB diagnosis, based on M. tuberculosis-specific T-cell memory phenotypes, activation or cytokine expression profiles [3-9]. Such biomarkers have also shown potential as surrogates to monitor TB treatment response [3, 10]. Here, we aimed to: 1) compare the diagnostic performance of several blood-based T-cell biomarkers in TB patients and controls; 2) determine if combinations of these biomarkers improve diagnostic performance; 3) identify the minimal number of flow cytometry parameters required to distinguish active TB patients from persons with latent M. tuberculosis infection (LTBI); and 4) translate measurement of these biomarkers from peripheral blood mononuclear cells to whole blood to enable a simplified application in field studies. We enrolled 25 healthy adults with LTBI, defined by QuantiFERON-TB Gold In-Tube assay (QFT; Qiagen, Hilden, Germany), and 25 HIV-negative adults with TB disease (XpertMTB/RIF+) (Cepheid, Sunnyvale, CA, USA) from a region endemic for TB in South Africa. Blood was collected prior to treatment initiation in persons with active TB and 12-18 months later, after they were declared cured (n=19). In controls, blood was also collected at 12-18 months after enrolment (n=20). Whole blood was stimulated with whole mycobacteria (BCG, Connaught), ESAT-6/CFP-10 peptides from QFT tubes, and TB122 peptide pool [11] using a standardised protocol [12]. Cells were stained with the following antibodies: anti-CD3 (Beckman Coulter, Krefeld, Germany; clone UCHT1; ECD), anti-CD4

Can we predict tuberculosis cure? What tools are available?

“…Therefore, late effector T-cells exhibit low to no CD27 expression [58,59]. Several studies have demonstrated that significantly higher proportions of M. tuberculosis-specific IFN-γ-producing CD4 + T-cells do not express CD27 (CD27 − IFN-γ + CD4 + ) in persons with active TB disease when compared with healthy controls or cured TB patients [60][61][62][63][64][65][66]. In addition, it has been shown that frequencies of CD27 − IFN-γ + CD4 + cells strongly correlate with the degree of lung pathology and matrix destruction [67,68], providing a good biomarker of TB treatment success [63,69].…”

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

“…For the analysis of donors in a non-endemic setting, non-parametric ANOVA tests were performed due to the smaller sample size (n=10). Dot plot graphs were compiled with GraphPad Prism and heat maps were generated using Qlucore v.3.4 (11). Receiver operating characteristic (ROC) curves were calculated and compared with R Studio v.3.3.1 pROC package and results drawn with graphical package ggplot2 v.2.1.0.…”

Diagnostic Challenge of Tuberculosis Heterogeneity

“…46,47 This concept has been further validated and translated to a simplified whole blood assay which is more suitable for future large field and pediatric studies. 48 Moreover, proportions of activated Mtb-specific T cells decrease during TB treatment, suggesting potential to monitor treatment response. 45,48 Whether this assay could also identify asymptomatic individuals with subclinical disease, who are at high risk of progressing to active disease, is likely but remains to be established.…”

“…48 Moreover, proportions of activated Mtb-specific T cells decrease during TB treatment, suggesting potential to monitor treatment response. 45,48 Whether this assay could also identify asymptomatic individuals with subclinical disease, who are at high risk of progressing to active disease, is likely but remains to be established. TB disease has also been associated with increased differentiation of Mtb-specific T cells, measured by loss of CD27 expression.…”

“…49 While this finding has been validated in children, 49 the utility of this biomarker in adults appears more limited 47 and does not seem to yield additional diagnostic accuracy over HLA-DR expression. 48 Cytokine expression profiles of Mtb-specific T cells are also affected by TB disease status, 50 but findings have been inconsistent across different studies, 47 perhaps due to different assay protocols used or heterogeneity in clinical phenotypes, including antigen load. 43 Mounting evidence suggests that the frequency of Mtbspecific CD8 þ T cells can serve as a reflection of in vivo antigen load and disease burden.…”