Small Extracellular Vesicles Isolated from Serum May Serve as Signal-Enhancers for the Monitoring of CNS Tumors

Abstract

Liquid biopsy-based methods to test biomarkers (e.g., serum proteins and extracellular vesicles) may help to monitor brain tumors. In this proteomics-based study, we aimed to identify a characteristic protein fingerprint associated with central nervous system (CNS) tumors. Overall, 96 human serum samples were obtained from four patient groups, namely glioblastoma multiforme (GBM), non-small-cell lung cancer brain metastasis (BM), meningioma (M) and lumbar disc hernia patients (CTRL). After the isolation and characterization of small extracellular vesicles (sEVs) by nanoparticle tracking analysis (NTA) and atomic force microscopy (AFM), liquid chromatography -mass spectrometry (LC-MS) was performed on two different sample types (whole serum and serum sEVs). Statistical analyses (ratio, Cohen’s d, receiver operating characteristic; ROC) were carried out to compare patient groups. To recognize differences between the two sample types, pairwise comparisons (Welch’s test) and ingenuity pathway analysis (IPA) were performed. According to our knowledge, this is the first study that compares the proteome of whole serum and serum-derived sEVs. From the 311 proteins identified, 10 whole serum proteins and 17 sEV proteins showed the highest intergroup differences. Sixty-five proteins were significantly enriched in sEV samples, while 129 proteins were significantly depleted compared to whole serum. Based on principal component analysis (PCA) analyses, sEVs are more suitable to discriminate between the patient groups. Our results support that sEVs have greater potential to monitor CNS tumors, than whole serum.

Raman Spectral Signatures of Serum-Derived Extracellular Vesicle-Enriched Isolates May Support the Diagnosis of CNS Tumors

“…For this purpose, 138 serum samples obtained from four patient groups were analyzed. Serum samples were collected from three brain tumor groups considered as the most common malignant, benign and metastatic brain tumors (GBM, BM, M) and from a control group (CTRL) [ 20 , 50 , 51 ]. sEV-sized particles from serum samples were isolated by differential centrifugation.…”

“…Despite these difficulties, we have revealed in a previously published article that EV isolation from the serum samples of the same patient groups significantly improves the signal-to-noise ratio, even in the case of GBM with an elevated procoagulant activity [ 20 ]. Although abundant serum proteins and LPs were still present in EV isolates, isolation depleted their concentration and enriched the EV and tumor-specific protein markers.…”

“…Secreted EVs are stably present in body fluids, and represent a concentrated sample of the cytosolic milieu (proteins, nucleic acids, and lipids) of the donor cells [ 16 , 17 , 18 ]. It has been shown that EVs isolated from the serum and plasma offer a useful tool to improve the signal-to-noise ratio in analytics by assuring to abundant protein depletion (such as albumin and lipoproteins) and enriching the tumor-specific molecular composition [ 19 , 20 ]. Moreover, EVs can cross various biological barriers, such as the blood–brain barrier (BBB), and easily enter the peripheral blood [ 21 , 22 ].…”

“…The serum samples were collected from patients diagnosed with the two most common types of brain tumors, namely malignant glioblastoma multiforme (GBM) and the typically benign meningioma (M), as well as from patients with a prevalent brain metastasis originating from non-small-cell lung cancer (BM). Patients with lumbar disc herniation without evidence of neurological cancer served as controls (CTRL) [ 20 , 50 , 51 ]. Particles within the size range of small EVs (sEVs) were isolated from serum samples via differential centrifugation, and were assessed by a Raman microscope.…”

Small Extracellular Vesicles Released from Glioma Cells Under Hypoxic Conditions Induce Pericyte-Phenotype Transition of Glioma Stem Cells.

“…TGF-β1 signaling pathway mediates this phenotype transition. It is well known that sEVs constitute an outstanding communication system in GBM, they can deliver not only a wide variety of coding and noncoding RNAs, but soluble proteins that can alter gene expression and cellular function in recipient cells [35,36]. Evidence from the last few years show that GBMs interact with surrounding non-cancer cells through sEVs to maintain a microenvironment that favors tumor proliferation, invasion of the brain, angiogenesis and immunosuppression [37].…”