PROJECT

“An integrative biomarker discovery and validation approach to aid the development of point-of-care tests for tuberculosis”

January 2024 - December 2026

Objectives:

1) To validate the performance of individual as well as combinations previously identified soluble protein, miRNA and metabolite biomarkers for predicting risk of progression to active TB disease (predictive biomarkers).
2) To evaluate the diagnostic accuracy of a previously identified miRNA and mRNA signature in blood and circulating cell-free Mycobacterium tuberculosis DNA and to identify novel monocyte-specific miRNAs for detection or triaging for active TB (diagnostic biomarkers).
3) To explore the usefulness of soluble complement proteins and neutrophil extracellular trap markers in plasma, and non-replicating persister (NRP) bacteria in sputum of TB patients undergoing anti-TB treatment, for monitoring response to treatment and predicting treatment outcome (prognostic biomarkers).
4) To capture and catalogue the landscape of transcriptomic interplay between the host and pathogen in different manifestations of TB to help in the identification of unique signatures for different forms of extrapulmonary TB, that may serve as novel drug targets/diagnostic biomarkers.

Study Overview:

Millions of TB cases still go undiagnosed each year. While traditional TB diagnostic methods such as culture or smear microscopy are slow or low in sensitivity, newer molecular techniques such as GeneXpert MTB/RIF and Line Probe Assay are costly and often unavailable in primary-care settings because of their infrastructure needs. Providing high-quality patient-centered diagnostics and care when patients first present to the health care facility would increase the number of TB cases detected and reduce patient loss at the initial stage of the care cascade, which is key to reaching the End-TB target. The scope of this study is to undertake rigorous evaluation of the diagnostic accuracy of the predictive, diagnostic and prognostic biomarkers already identified by our group in earlier exploratory studies with the aim of identifying the most promising biomarkers and biomarker categories that can be taken forward for the development of a POC test.

Study Approach:

The study will use ELISA/bioanalytical methods to measure soluble proteomic biomarkers including cytokines, chemokines, inflammatory molecules, complement factors, acute phase proteins, nutritional factors and neutrophil extracellular trap (NET) markers in plasma/QuantiFERON supernatants. qRT-PCR will be used to measure levels of mRNA and miRNA biomarkers in blood and plasma respectively. Flowcytometry will be employed to analyze NET markers on neutrophils and for monocyte sorting for identification of monocyte-specific miRNAs. Droplet digital PCR will be used to quantify circulating cell-free Mycobacterium tuberculosis DNA. Dual RNAseq analysis will be used to identify novel mRNA signatures characterizing different stages in the spectrum of TB pathogenesis including latent, incipient and sub-clinical disease, and those specific for various forms of extrapulmonary TB.

Expected Public Health Impact:

The study will provide evidence on the diagnostic performance of various types of host and and pathogen-associated cellular and molecular biomarkers and help in the identification of the most potential biomarker/biomarker category that can be exploited for the developing of a POC diagnostic/triaging test for TB which in turn will contribute to better control and management of tuberculosis.