Abstract
Aim: Small extracellular vesicles (sEVs) are promising noninvasive biomarkers for several malignancies, including thyroid carcinoma (TC). However, their heterogeneity is frequently overlooked in bulk-level analyses. Methods: Plasma samples from TC and healthy controls (HC) were collected for a proximity-dependent barcoding assay (PBA) to identify plasma sEV biomarkers at the single-sEV level. We screened the potential biomarkers using the Panel260 (the panel that detects 260 proteins) of PBA in Cohort 1, and validated them using Panel550 (the panel that detects 550 proteins) in Cohort 2. Results: Plasma exosome counts were significantly elevated in TC compared with those in HC in both Cohort 1 and Cohort 2. Receiver operating characteristic curve analysis showed that sEV counts exhibited an area under the curve > 0.75 in both cohorts. The sEV proteomic analysis revealed that sEV epithelial cell adhesion molecule (EPCAM) levels were significantly increased, whereas claudin-11, integrin alpha X, and lymphocyte-activating 3 were significantly decreased in TC compared with HC. The increase in EPCAM in the plasma and tumor tissues was confirmed by enzyme-linked immunosorbent assay and immunohistochemistry analyses, respectively. The sEV subpopulation analysis further demonstrated that EPCAM(+) sEVs were significantly elevated in TC compared with HC in both cohorts. The reduction in sEV counts was observed in 18 out of 20 patients after the operation. The decrease in EPCAM(+) sEVs was observed in 20 patients with TC post-operatively, whereas the reduction in the conventional biomarker serum thyroglobulin (Tg) was observed in 14 patients. TC-derived plasma sEVs promoted TC cell proliferation, migration, invasion, and TC xenograft growth. Conclusion: EPCAM(+) sEVs could serve as a promising biomarker for the early diagnosis of TC and perform better in monitoring post-operative remission of TC than serum Tg.