Comprehensive Proteomic Characterization of Breast Cancer-Derived Small Extracellular Vesicles for Biomarker Discovery

Abstract

Breast cancer (BC) is the most prevalent cancer and the second leading cause of cancer-related mortality among Canadian women. Early detection and treatment significantly improve survival rates. The potential use of small extracellular vesicles (sEVs) as biomarkers for early BC diagnosis has gained increasing interest, mainly due to their promise as a non-invasive detection method. This study compared the sEVs proteome derived from a highly metastatic BC cell line, MDA-MB-231, with its parental cell line and exosomes derived from a non-cancerous breast epithelial MCF-10A cell line. sEVs isolation was performed using a less common approach involving precipitation with polyethylene glycol (PEG)-8000, separation via size exclusion chromatography (SEC), and concentration using filtration. Bioinformatic analyses revealed multiple enriched pathways in MDA-MB-231-derived sEVs contributing to cancer growth and proliferation. The proteins of these pathways can be a good source for BC biomarkers. In this study, three proteins were selected based on their unique presence in MDA-MB-231-derived sEVs and their contribution to the detected pathways related to BC: peroxidasin homolog (PXDN), glutathione hydrolase 5 proenzyme (GGT5), and plasminogen activator inhibitor 1 (SERPINE1). These proteins were validated through mass spectrometry-based parallel reaction monitoring, utilizing isotopically labelled synthetic peptides. This study highlights the promise of sEV-based proteins as non-invasive biomarkers for early BC diagnosis, potentially improving detection methods and patient outcomes.