Proteomic Exploration of Plasma Exosomes and Other Small Extracellular Vesicles in Pediatric Hodgkin Lymphoma: A Potential Source of Biomarkers for Relapse Occurrence

Abstract

Exosomes and other small extracellular vesicles (EVs) are potential sources of cancer biomarkers. Plasma-derived EVs have not yet been studied in pediatric Hodgkin lymphoma (HL), for which predictive biomarkers of relapse are greatly needed. In this two-part proteomic study, we used two-dimensional difference gel electrophoresis (2D-DIGE) followed by liquid chromatography–tandem mass spectrometry (LC–MS/MS) to analyze EV proteins of plasma collected at diagnosis from children with nodular sclerosis HL, relapsed or not. EVs isolated using membrane affinity had radii ranging from 20 to 130 nm and contained the programmed cell death 6-interacting (ALIX) and the tumor susceptibility gene 101 (TSG101) proteins, whereas calnexin (CANX) was not detected. 2D-DIGE identified 16 spots as differentially abundant between non-relapsed and relapsed HL (|fold change| ≥ 1.5, p < 0.05). LC–MS/MS identified these spots as 11 unique proteins, including five more abundant in non-relapsed HL (e.g., complement C4b, C4B; fibrinogen γ chain, FGG) and six more abundant in relapsed HL (e.g., transthyretin, TTR). Shotgun LC–MS/MS on pooled EV proteins from non-relapsed HL identified 161 proteins, including 127 already identified in human exosomes (ExoCarta data). This EV cargo included 89 proteins not yet identified in exosomes from healthy plasma. Functional interrogation by the Database for Annotation, Visualization and Integrated Discovery (DAVID) revealed that the EV proteins participate in platelet degranulation and serine-type endopeptidase activity as the most significant Gene Ontology (GO) biological process and molecular function (p < 0.01).

The updated role of exosomal proteins in the diagnosis, prognosis, and treatment of cancer

“…He et al 68 identified a novel function of endoplasmic reticulum (ER) stress-associated exosomes in mediating macrophage cytokine (i.e., IL-10) secretion in the liver cancer microenvironment and indicated the potential to treat liver cancer by targeting an ER stress-exosomal-STAT3 pathway. Hodgkin lymphoma-derived exosomes can facilitate the transformation of normal fibroblasts into tumor-specific fibroblasts, which causes the release of proinflammatory cytokines (e.g., IL-1α, IL-6, and TNF-α), growth factors (e.g., G-CSF and GM-CSF) and proangiogenic factors (e.g., VEGF) 28 , 69 . This finding is supported by research on metastatic melanoma-derived exosomes with surface expression of PD-L1 12 .…”

“…(3) Due to advances in mass spectrometry-based detection technology and the continuous optimization of data collection and analysis methods, the protein coverage and sensitivity of comparative proteomics have improved, and specific exosomal proteins in urine from patients with cancer can be detected 14 . The application of proteomics tools to analyze posttranslational modifications in exosome research has also increased the depth of exosome proteomics data in the context of cancer pathogenesis, function, and disease associations 28 . In recent years, researchers have paid increasing attention to exosomal proteins, and numerous studies on exosomal proteins have been recently published.…”

AuNP Aptasensor for Hodgkin Lymphoma Monitoring

“…Small extracellular vesicles (SEVs) are lipid-bilayer-bound vesicles released from living cells into the extracellular environment, which mediate cell-to-cell communication and are present in most of the biological fluids including plasma [ 4 ]. For instance, SEVs secreted by HRS cells into interstitial spaces have been detected in circulating plasma and could present a potential source of biomarkers [ 5 ].…”

Profile of circulating extracellular vesicles microRNA correlates with the disease activity in granulomatosis with polyangiitis

“…Based on their size and biogenesis, EVs are classified as apoptotic bodies (large EVs, released by apoptotic cells), microvesicles (medium EVs, released from cells plasma membrane), and exosomes (small EVs, produced in multivesicular bodies) [ 10 ]. EVs can communicate molecular signals due to their cargo of proteins [ 11 , 12 ], lipids [ 13 , 14 ], and nucleic acids (DNA, RNA) [ 15 , 16 ] delivered to target cells [ 9 ]. MicroRNAs (miRNA) transported by EVs are frequently studied for their impact on the expression of genes.…”