Comprehensive plasma proteomic profiling reveals biomarkers for active tuberculosis
Diana J. Garay-Baquero, Cory H. White, Naomi F. Walker, Marc Tebruegge, Hannah F. Schiff, Cesar Ugarte-Gil, Stephen Morris-Jones, Ben G. Marshall, Antigoni Manousopoulou, John Adamson, Andres F. Vallejo, Magdalena K. Bielecka, Robert J. Wilkinson, Liku B. Tezera, Christopher H. Woelk, Spiros D. Garbis, Paul Elkington
Diana J. Garay-Baquero, Cory H. White, Naomi F. Walker, Marc Tebruegge, Hannah F. Schiff, Cesar Ugarte-Gil, Stephen Morris-Jones, Ben G. Marshall, Antigoni Manousopoulou, John Adamson, Andres F. Vallejo, Magdalena K. Bielecka, Robert J. Wilkinson, Liku B. Tezera, Christopher H. Woelk, Spiros D. Garbis, Paul Elkington
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Clinical Research and Public Health Infectious disease

Comprehensive plasma proteomic profiling reveals biomarkers for active tuberculosis

Abstract

BACKGROUND Tuberculosis (TB) kills more people than any other infection, and new diagnostic tests to identify active cases are required. We aimed to discover and verify novel markers for TB in nondepleted plasma.METHODS We applied an optimized quantitative proteomics discovery methodology based on multidimensional and orthogonal liquid chromatographic separation combined with high-resolution mass spectrometry to study nondepleted plasma of 11 patients with active TB compared with 10 healthy controls. Prioritized candidates were verified in independent UK (n = 118) and South African cohorts (n = 203).RESULTS We generated the most comprehensive TB plasma proteome to date, profiling 5022 proteins spanning 11 orders-of-magnitude concentration range with diverse biochemical and molecular properties. We analyzed the predominantly low–molecular weight subproteome, identifying 46 proteins with significantly increased and 90 with decreased abundance (peptide FDR ≤ 1%, q ≤ 0.05). Verification was performed for novel candidate biomarkers (CFHR5, ILF2) in 2 independent cohorts. Receiver operating characteristics analyses using a 5-protein panel (CFHR5, LRG1, CRP, LBP, and SAA1) exhibited discriminatory power in distinguishing TB from other respiratory diseases (AUC = 0.81).CONCLUSION We report the most comprehensive TB plasma proteome to date, identifying novel markers with verification in 2 independent cohorts, leading to a 5-protein biosignature with potential to improve TB diagnosis. With further development, these biomarkers have potential as a diagnostic triage test.FUNDING Colciencias, Medical Research Council, Innovate UK, NIHR, Academy of Medical Sciences, Program for Advanced Research Capacities for AIDS, Wellcome Centre for Infectious Diseases Research.

Authors

Diana J. Garay-Baquero, Cory H. White, Naomi F. Walker, Marc Tebruegge, Hannah F. Schiff, Cesar Ugarte-Gil, Stephen Morris-Jones, Ben G. Marshall, Antigoni Manousopoulou, John Adamson, Andres F. Vallejo, Magdalena K. Bielecka, Robert J. Wilkinson, Liku B. Tezera, Christopher H. Woelk, Spiros D. Garbis, Paul Elkington

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Figure 1

Overview of the plasma proteomic discovery and validation strategy of potential TB biomarkers.

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Overview of the plasma proteomic discovery and validation strategy of po...
(A) Identification and quantification of plasma proteins were performed using a quantitative multidimensional protein identification approach, which comprises a series of fractionation steps at both protein (denaturing high-performance–size exclusion chromatography, HP-SEC) and peptide levels (offline high pH C4 HPLC followed by online low pH C18 ultrahigh-performance LC, UPLC). Initial plasma prefractionation using HP-SEC produces 5 segments depending on the molecular size. Only segments 1 to 4 were included in this study because these included most of the protein contents. (B) Bioinformatic processing prioritized markers, which were then measured by ELISA or Luminex in plasma or serum samples from 2 cohorts. Discovery and validation stages involved multiple ethnicities. iTRAQ, isobaric tags for relative and absolute quantitation; nESI-FTMS, nano-electrospray ionization Fourier-transform mass spectrometry.