Deciphering the peptidome of urine from ovarian cancer patients and healthy controls

Abstract

BackgroundOvarian cancer (OvCa) is the most lethal gynecological malignancy. The emergence of high-throughput technologies, such as mass spectrometry, has allowed for a paradigm shift in the way we search for novel biomarkers. Urine-based peptidomic profiling is a novel approach that may result in the discovery of noninvasive biomarkers for diagnosing patients with OvCa. In this study, the peptidome of urine from 6 ovarian cancer patients and 6 healthy controls was deciphered.ResultsUrine samples underwent ultrafiltration and the filtrate was subjected to solid phase extraction, followed by fractionation using strong cation exchange chromatography. These fractions were analyzed using an Orbitrap mass spectrometer. Over 4600 unique endogenous urine peptides arising from 713 proteins were catalogued, representing the largest urine peptidome reported to date. Each specimen was processed in triplicate and reproducibility at the protein (69-76%) and peptide (58-63%) levels were noted. More importantly, over 3100 unique peptides were detected solely in OvCa specimens. One such promising biomarker was leucine-rich alpha-2-glycoprotein (LRG1), where multiple peptides were found in all urines from OvCa patients, but only one peptide was found in one healthy control urine sample.ConclusionsMining the urine peptidome may yield highly promising novel OvCa biomarkers.

Discovery and Validation of Novel Protein Biomarkers in Ovarian Cancer Patient Urine

“…In recent studies, expression analysis and immunohistochemical staining of ovarian tumor tissues identified no significant difference in uteroglobin levels between ovarian cancer tissue and normal ovarian tissue . Our study and a previous analysis identified the increased abundance of endogenous uteroglobin peptides in ovarian cancer patient urine samples, strongly supporting its potential as a urinary biomarker candidate for further investigation. Insulin‐like growth factor‐binding protein 3 (IGFBP3) is expressed in a wide variety of cancer types (including lung, liver, and breast), and methylation of the IGFBP3 gene has been identified as a prognostic marker of ovarian cancer progression .…”

“…Label‐free quantitative (LFQ) mass spectrometry, whilst well established as a proteomic discovery tool, has seen only limited application in the analysis of ovarian cancer–related proteomic changes to date . Only a single study has applied LFQ to examine the urine peptidome of ovarian cancer patients versus healthy controls . In previous work, we used isotope‐coded protein labeling (ICPL) and MALDI MS/MS to identify urinary biomarkers of ovarian cancer .…”

Syringe-push membrane absorption as a simple rapid method of urine preparation for clinical proteomics

“…Ultrafiltration was performed as described previously [6, 9]. Briefly, 5-ml urine was spun at 12,000 × g using an ultrafiltration column at 4 °C until approximately 1/30 of initial volume remained.…”

Stress Urinary Incontinence: A Proteomics Overview

“…Another protein, the leucine-rich alpha-2-glycoprotein, was also found to be increased in samples of SUI patients. This protein is secreted and normally present in plasma; however, it was also described to be involved in nonspecific inflammatory and cancer processes [25][26][27][28]. It has recently been described in the context of ulcerative colitis activity, pediatric, invasive bladder cancer, biliary tract cancer, lung cancer, pancreatic cancer, heart failure, neutrophilic granulocyte differentiation, and autoimmune diseases [29,30].…”