Glioblastomas (GBM) are highly aggressive primary brain tumors. Complex and dynamic tumor microenvironment (TME) plays a crucial role in the sustained growth, proliferation, and invasion of GBM. Several means of intercellular communication have been documented between glioma cells and the TME, including growth factors, cytokines, chemokines as well as extracellular vesicles (EVs). EVs carry functional genomic and proteomic cargo from their parental cells and deliver that information to surrounding and distant recipient cells to modulate their behavior. EVs are emerging as crucial mediators of establishment and maintenance of the tumor by modulating the TME into a tumor promoting system. Herein we review recent literature in the context of GBM TME and the means by which EVs modulate tumor proliferation, reprogram metabolic activity, induce angiogenesis, escape immune surveillance, acquire drug resistance and undergo invasion. Understanding the multifaceted roles of EVs in the niche of GBM TME will provide invaluable insights into understanding the biology of GBM and provide functional insights into the dynamic EV-mediated intercellular communication during gliomagenesis, creating new opportunities for GBM diagnostics and therapeutics.

Tumour cells release diverse populations of extracellular vesicles (EVs) ranging in size, molecular cargo, and function. We sought to characterize mRNA and protein content of EV subpopulations released by human glioblastoma (GBM) cells expressing a mutant form of epidermal growth factor receptor (U87EGFRvIII) in vitro and in vivo with respect to size, morphology and the presence of tumour cargo. The two EV subpopulations purified from GBM U87EGFRvIII cancer cells, non-cancer human umbilical vein endothelial cells (HUVEC; control) and serum of U87EGFRvIII glioma-bearing mice using differential centrifugation (EVs that sediment at 10,000 × g or 100,000 × g are termed large EVs and small EVs, respectively) were characterized using transmission electron microscopy (TEM), confocal microscopy, nanoparticle tracking analysis (NTA), flow cytometry, immunofluorescence (IF), quantitative-polymerase chain reaction (qPCR), droplet digital polymerase chain reaction (ddPCR) and micro-nuclear magnetic resonance (μNMR). We report that both U87EGFRvIII and HUVEC release a similar number of small EVs, but U87EGFRvIII glioma cells alone release a higher number of large EVs compared to non-cancer HUVEC. The EGFRvIII mRNA from the two EV subpopulations from U87EGFRvIII glioma cells was comparable, while the EGFR protein (wild type + vIII) levels are significantly higher in large EVs. Similarly, EGFRvIII mRNA in large and small EVs isolated from the serum of U87EGFRvIII glioma-bearing mice is comparable, while the EGFR protein (wild type + vIII) levels are significantly higher in large EVs. Here we report for the first time a direct comparison of large and small EVs released by glioma U87EGFRvIII cells and from serum of U87EGFRvIII glioma-bearing mice. Both large and small EVs contain tumour-specific EGFRvIII mRNA and proteins and combining these platforms may be beneficial in detecting rare mutant events in circulating biofluids.

The past decade has witnessed a rapid growth in the field of extracellular vesicle (EV) based biomarkers for the diagnosis and monitoring of cancer. Several studies have reported novel EV based biomarkers, but the technical and clinical validation phase has been hampered by general challenges common to biomedical research field as well as specific challenges inherent to the nanoparticle field. This has led to more common failures than success stories in the biomarker discovery pipeline. As a result, more attention must be focused on the process of biomarker discovery, verification, and validation to allow for translation and application of novel EV based research to patient care. Herein, we briefly discuss the hurdles and potential solutions in EV biomarker discovery and verification and validation, and clinical translation.