METHYLATION AND MUTATION PROFILES IN MENINGIOMA CELL-DERIVED EXTRACELLULAR VESICLE DNA REFLECT EPIGENETIC AND GENOMIC ALTERATIONS IN ORIGINAL TUMORS

Abstract The majority of meningiomas are benign but approximately 20% of display an aggressive behavior, resulting in significant patient morbidity and mortality. Standard monitoring after meningioma resection relies on serial MRI examinations, which are time-consuming, expensive and provide no information on molecular alterations that may indicate progression towards a more aggressive tumor. Extracellular vesicles (EVs) are released by tumor cells and contain high molecular weight DNA, rendering circulating EVs a potential biomarker source for non-invasive disease monitoring and for obtaining information on genetic and epigenetic alterations. We quantified EVs in plasma of 46 meningioma patients (n = 29 M1, 12 M2, 5 M3) by nanoparticle tracking analysis and detected significantly higher levels compared to age-matched healthy donors (n = 18). EV concentrations correlated with malignancy grade (p = 0.0049) and with the extent of peritumoral edema (p = 0.0031). Comparisons between paired pre- and postoperative samples revealed that EV levels counts dropped significantly the day after tumor resection and were reduced to normal levels after about one week. Completely resected patients (Simpson grade I) displayed a greater reduction of postoperative EV concentrations than incompletely resected patients. DNA methylation profiling was performed on EVs secreted by cultured meningioma cells, as well as matched cells and original tumors using 850k arrays (n = 7 M1, 5 M2, 3 M3). All EV samples were correctly identified as meningiomas by the Heidelberg classifier, and methylation subclasses were also correctly assigned in almost all cases. t-SNE analysis showed that EVs mapped in close proximity to their corresponding parental cells and tumor tissue. Tumor specific mutations and copy number variations were detected in EV-DNA with high accuracy. Differential quantitative proteomic analysis of EVs, cells and tumors identified shared proteins that could potentially be useful for enriching tumor-derived circulating EVs from biofluids.