Dataset Information

Identification of candidate circulating glucocorticoid response biomarkers in humans using a proteomic approach

ABSTRACT: Glucocorticoids used in pharmacological doses for the treatment of a variety of medical conditions, and endogenous glucocorticoid excess – Cushing’s syndrome, may result in several adverse effects, but currently there is no clinically useful biomarker of glucocorticoid activity. A three-phase protein biomarker discovery strategy was used. Proteomic biomarker discovery and qualification was conducted on the secretome of ex vivo-stimulated peripheral blood mononuclear cells (PBMC) isolated from 6 volunteers, incubated ± dexamethasone 100 ng/mL for 4h and 24h. Untargeted proteomics with label-free quantification (LFQ) was conducted to discover candidate proteins which were quantified using targeted proteomics by a custom multiple reaction monitoring mass spectrometry (MRM-MS) assay. Five proteins were selected for serum measurement by immunoassay in 20 healthy volunteers, with blood drawn at baseline and 12h after 4 mg oral dexamethasone. Paired analysis of the discovery proteomics data (576 and 280 proteins for the 4h and 24h secretomes, respectively) generated a shortlist of candidates which were qualified using MRM-MS to obtain protein level intensity data for 39 proteins. In the validation cohort, 3/5 proteins were dexamethasone-responsive, two significantly decreased: lysozyme C (mean±SEM) – 101±5.5 vs 67±4.4 ng/mL, (P<0.0001); nucleophosmin-1 (median (interquartile range)) – 16.6 (14.4-18.4) vs 14.2 (11.1-17.4) ng/mL, (P<0.01), while high mobility group box 2 (mean±SEM) significantly increased – 819±34 vs 984±60 pg/mL (P<0.01). Using an ex vivo proteomic approach in PBMC, we have identified and verified circulating glucocorticoid-responsive proteins which may have potential as serum biomarkers.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Cell, Blood

SUBMITTER: Thomas Stoll

LAB HEAD: Michelle M. Hill

PROVIDER: PXD019945 | Pride | 2021-03-14

REPOSITORIES: Pride

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Source:

			Action	DRS
	20160727_JBarclay_1.mzML	Mzml
	20160727_JBarclay_1.raw	Raw
	20160727_JBarclay_10.mzML	Mzml
	20160727_JBarclay_10.raw	Raw
	20160727_JBarclay_11.mzML	Mzml

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Publications

Ex vivo glucocorticoid-induced secreted proteome approach for discovery of glucocorticoid-responsive proteins in human serum.

Inder Warrick J WJ Mohamed Ahmed A Keshvari Sahar S Barclay Johanna L JL Ruelcke Jayde E JE Stoll Thomas T Nolan Brendan J BJ Cesana-Nigro Nicole N Hill Michelle M MM

Proteomics. Clinical applications 20210312 2-3

<h4>Purpose</h4>To identify glucocorticoid-responsive proteins measurable in human serum that may have clinical utility in therapeutic drug monitoring and the diagnosis of cortisol excess or deficiency.<h4>Experimental design</h4>A phased biomarker discovery strategy was conducted in two cohorts. Secretome from peripheral blood mononuclear cells (PBMC) isolated from six volunteers after ex vivo incubation ± dexamethasone (DEX) 100 ng/mL for 4 h and 24 h was used for candidate discovery and quali ...[more]

PMID: 33641263

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