Journal
CANCER SCIENCE
Volume 112, Issue 5, Pages 2033-2045Publisher
WILEY
DOI: 10.1111/cas.14881
Keywords
bladder cancer; exosome; extracellular vesicle; proteomics; urinary biomarker
Categories
Funding
- Japan Society for the Promotion of Science [JP17K16788]
- Japan Agency for Medical Research and Development [A102]
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Proteomic analysis of urinary EVs combined with Te-EVs allows for selective detection of urinary BCa biomarkers.
Proteomic analysis of urinary extracellular vesicles (EVs) is a powerful approach to discover potential bladder cancer (BCa) biomarkers, however urine contains numerous EVs derived from the kidney and normal urothelial epithelium, which can obfuscate information related to BCa cell-derived EVs. In this study, we combined proteomic analysis of urinary EVs and tissue-exudative EVs (Te-EVs), which were isolated from culture medium of freshly resected viable BCa tissues. Urinary EVs were isolated from urine samples of 11 individuals (7 BCa patients and 4 healthy individuals), and Te-EVs were isolated from 7 BCa tissues. We performed tandem mass tag (TMT)-labeling liquid chromatography (LC-MS/MS) analysis for both urinary EVs and Te-EVs and identified 1960 proteins in urinary EVs and 1538 proteins in Te-EVs. Most of the proteins identified in Te-EVs were also present in urinary EVs (82.4%), with 55 of these proteins showing upregulated levels in the urine of BCa patients (fold change > 2.0; P < .1). Among them, we selected 22 membrane proteins as BCa biomarker candidates for validation using selected reaction monitoring/multiple reaction monitoring (SRM/MRM) analysis on urine samples from 70 individuals (40 BCa patients and 30 healthy individuals). Six urinary EV proteins (heat-shock protein 90, syndecan-1, myristoylated alanine-rich C-kinase substrate (MARCKS), MARCKS-related protein, tight junction protein ZO-2, and complement decay-accelerating factor) were quantified using SRM/MRM analysis and validated as significantly upregulated in BCa patients (P < .05). In conclusion, the novel strategy that combined proteomic analysis of urinary EVs and Te-EVs enabled selective detection of urinary BCa biomarkers.
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