title: Low Mass Blood Peptides Discriminative of Inflammatory Bowel Disease (IBD) Severity: A Quantitative Proteomic Perspective.
abstract: Breakdown of the protective gut barrier releases effector molecules and degradation products into the blood stream making serum and plasma ideal as a diagnostic medium. The enriched low mass proteome is unexplored as a source of differentiators for diagnosing and monitoring inflammatory bowel disease (IBD) activity, that is less invasive than colonoscopy. Differences in the enriched low mass plasma proteome (<25 kDa) were assessed by label-free quantitative mass-spectrometry. A panel of marker candidates were progressed to validation phase and "Tier-2" FDA-level validated quantitative assay. Proteins important in maintaining gut barrier function and homeostasis at the epithelial interface have been quantitated by multiple reaction monitoring in plasma and serum including both inflammatory; rheumatoid arthritis controls, and non-inflammatory healthy controls; ulcerative colitis (UC), and Crohn's disease (CD) patients. Detection by immunoblot confirmed presence at the protein level in plasma. Correlation analysis and receiver operator characteristics were used to report the sensitivity and specificity. Peptides differentiating controls from IBD originate from secreted phosphoprotein 24 (SPP24, p = 0.000086, 0.009); whereas those in remission and healthy can be differentiated in UC by SPP24 (p = 0.00023, 0.001), α-1-microglobulin (AMBP, p = 0.006) and CD by SPP24 (p = 0.019, 0.05). UC and CD can be differentiated by Guanylin (GUC2A, p = 0.001), and Secretogranin-1 (CHGB p = 0.035). Active and quiescent disease can also be differentiated in UC and CD by CHGB (p ≤ 0.023) SPP24 (p ≤ 0.023) and AMBP (UC p = 0.046). Five peptides discriminating IBD activity and severity had very little-to-no correlation to erythrocyte sedimentation rate, C-reactive protein, white cell or platelet counts. Three of these peptides were found to be binding partners to SPP24 protein alongside other known matrix proteins. These proteins have the potential to improve diagnosis and evaluate IBD activity, reducing the need for more invasive techniques. Data are available via ProteomeXchange with identifier PXD002821.
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia; v.wasinger@unsw.edu.au.
authors.forename: Valerie C
authors.identifier: http://orcid.org/0000-0001-5338-0869
authors.initials: VC
authors.lastname: Wasinger
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; ¶Gastroenterology Department, Concord Repatriation General Hospital, Hospital Rd, Concord, NSW, Australia;
authors.forename: Yunki
authors.initials: Y
authors.lastname: Yau
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia;
authors.forename: Xizi
authors.initials: X
authors.lastname: Duo
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia;
authors.forename: Ming
authors.initials: M
authors.lastname: Zeng
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia;
authors.forename: Beth
authors.initials: B
authors.lastname: Campbell
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia;
authors.forename: Sean
authors.initials: S
authors.lastname: Shin
authors.affiliation: From the ‡Bioanalytical Mass Spectrometry Facility, Mark Wainwright Analytical centre, The University of New South Wales, Australia; §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia;
authors.forename: Raphael
authors.initials: R
authors.lastname: Luber
authors.affiliation: ‖Department of Gastroenterology, Bankstown-Lidcombe Hospital, Eldridge Rd, Bankstown, NSW, Australia.
authors.forename: Diane
authors.initials: D
authors.lastname: Redmond
authors.affiliation: §School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia; ¶Gastroenterology Department, Concord Repatriation General Hospital, Hospital Rd, Concord, NSW, Australia; ‖Department of Gastroenterology, Bankstown-Lidcombe Hospital, Eldridge Rd, Bankstown, NSW, Australia.
authors.forename: Rupert W L
authors.initials: RW
authors.lastname: Leong
mesh_terms: D000328:Adult; D000368:Aged; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000175:diagnosis / Q000378:metabolism; D003093:Colitis, Ulcerative / Q000097:blood / Q000175:diagnosis / Q000378:metabolism; D003424:Crohn Disease / Q000097:blood / Q000175:diagnosis / Q000378:metabolism; D005260:Female; D006801:Humans; D015151:Immunoblotting; D015212:Inflammatory Bowel Diseases / Q000097:blood / Q000175:diagnosis / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D008875:Middle Aged; D008970:Molecular Weight; D010455:Peptides / Q000097:blood / Q000737:chemistry / Q000378:metabolism*; D020543:Proteome / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D012372:ROC Curve; D015203:Reproducibility of Results; D012720:Severity of Illness Index; D055815:Young Adult
chemical_list: D010455:Peptides; D020543:Proteome