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DOI: 10.3389/fmolb.2023.1138594; PMCID: PMC10140755


Ewing Sarcoma (EWS) is the second most common osseous malignancy in children and young adults after osteosarcoma, while it is the fifth common osseous malignancy within adult age population. The clinical presentation of EWS is quite often non-specific, with the most common symptoms at presentation consisting of pain, swelling or general discomfort. The dearth of clinically relevant diagnostic or predictive biomarkers continues to remain a pressing clinical challenge. Identification of tumor specific biomarkers can lend towards an early diagnosis, expedited initiation of therapy, monitoring of therapeutic response, and early detection of recurrence of disease. We carried-out a complex analysis of cell lines and cell line derived small extracellular vesicles (sEVs) using label-free-based Quantitative Proteomic Profiling with an intent to determine shared and distinct features of these tumor cells and their respective sEVs. We analyzed EWS cells with different EWS-ETS fusions (EWS-FLI1 type I, II, and III and EWS-ERG) and their corresponding sEVs. Non-EWS controls included osteosarcoma, rhabdomyosarcoma, and benign cells, i.e., osteoid osteoma and mesenchymal stem cells. Proteomic profiling identified new shared markers between cells and their corresponding cell-derived sEVs and markers which were exclusively enriched in EWS-derived sEVs. These exo-biomarkers identified were validated by in silico approaches of publicly available protein databases and by capillary electrophoresis based western analysis (Wes). Here, we identified a protein biomarker named UGT3A2 and found its expression highly specific to EWS cells and their sEVs compared to control samples. Clinical validation of UGT3A2 expression in patient tumor tissues and plasma derived sEV samples demonstrated its specificity to EWS, indicating its potential as a EWS biomarker.

Journal Title

Front Mol Biosci



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EWS-ETS fusions; ewing sarcoma; exosomes; liquid biopsy; mass-spectrometry; proteomics; sEV-associated protein biomarkers; small extracellular vesicles


Grant support

This study was supported in part by a grant from NIH National Cancer Institute (R33 CA214333 to AG), the Kansas Institute for Precision Medicine (GM130423 to AG), the KU Cancer Center’s Support Grant (P30 CA168524), the MCA Partners Advisory Board from Children’s Mercy Hospital (to GS) and The University of Kansas Cancer Center (to AG), Alex Lemonade Stand (to GS), Braden’s Hope for Childhood Cancer Foundation (to GS and AG), Noah’s Bandage Project (to GS and AG), and Hyundai Hope on Wheels (to GS). This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Publisher's Link: