Cannabidiol Affects Extracellular Vesicle Release, miR21 and miR126, and Reduces Prohibitin Protein in Glioblastoma Multiforme Cells

Glioblastoma multiforme (GBM) is the most aggressive form of adult primary malignant brain tumour with poor prognosis. Extracellular vesicles (EVs) are a key-mediator through which GBM cells promote a pro-oncogenic microenvironment. Peptidylarginine deiminases (PADs), which catalyze the post-translational protein deimination of target proteins, are implicated in cancer, including via EV modulation. Pan-PAD inhibitor Cl-amidine affected EV release from GBM cells, and EV related microRNA cargo, with reduced pro-oncogenic microRNA21 and increased anti-oncogenic microRNA126, also in combinatory treatment with the chemotherapeutic agent temozolomide (TMZ). The GBM cell lines under study, LN18 and LN229, differed in PAD2, PAD3 and PAD4 isozyme expression. Various cytoskeletal, nuclear and mitochondrial proteins were identified to be deiminated in GBM, including prohibitin (PHB), a key protein in mitochondrial integrity and also involved in chemo-resistance. Post-translational deimination of PHB, and PHB protein levels, were reduced after 1 h treatment with pan-PAD inhibitor Cl-amidine in GBM cells. Histone H3 deimination was also reduced following Cl-amidine treatment. Multifaceted roles for PADs on EV-mediated pathways, as well as deimination of mitochondrial, nuclear and invadopodia related proteins, highlight PADs as novel targets for modulating GBM tumour communication.

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Serum Extracellular Vesicle-Derived circHIPK3 and circSMARCA5 Are Two Novel Diagnostic Biomarkers for Glioblastoma Multiforme

Pharmaceuticals ◽

10.3390/ph14070618 ◽

2021 ◽

Vol 14 (7) ◽

pp. 618

Author(s):

Michele Stella ◽

Luca Falzone ◽

Angela Caponnetto ◽

Giuseppe Gattuso ◽

Cristina Barbagallo ◽

...

Keyword(s):

Glioblastoma Multiforme ◽

Roc Analysis ◽

Characteristic Curve ◽

Area Under The Curve ◽

Digital Pcr ◽

Extracellular Vesicle ◽

Size Exclusion ◽

Circular Rnas ◽

Diagnostic Biomarkers ◽

Exclusion Chromatography

Glioblastoma multiforme (GBM) is the most frequent and deadly human brain cancer. Early diagnosis through non-invasive biomarkers may render GBM more easily treatable, improving the prognosis of this currently incurable disease. We suggest the use of serum extracellular vesicle (sEV)-derived circular RNAs (circRNAs) as highly stable minimally invasive diagnostic biomarkers for GBM diagnosis. EVs were isolated by size exclusion chromatography from sera of 23 GBM and 5 grade 3 glioma (GIII) patients, and 10 unaffected controls (UC). The expression of two candidate circRNAs (circSMARCA5 and circHIPK3) was assayed by droplet digital PCR. CircSMARCA5 and circHIPK3 were significantly less abundant in sEVs from GBM patients with respect to UC (fold-change (FC) of −2.15 and −1.92, respectively) and GIII (FC of −1.75 and −1.4, respectively). Receiver operating characteristic curve (ROC) analysis, based on the expression of sEV-derived circSMARCA5 and circHIPK3, allowed us to distinguish GBM from UC (area under the curve (AUC) 0.823 (0.667–0.979) and 0.855 (0.704 to 1.000), with a 95% confidence interval (CI), respectively). Multivariable ROC analysis, performed by combining the expression of sEV-derived circSMARCA5 and circHIPK3 with preoperative neutrophil to lymphocyte (NLR), platelet to lymphocyte (PLR) and lymphocyte to monocyte (LMR) ratios, three known diagnostic and prognostic GBM markers, allowed an improvement in the GBM diagnostic accuracy (AUC 0.901 (0.7912 to 1.000), 95% CI). Our data suggest sEV-derived circSMARCA5 and circHIPK3 as good diagnostic biomarkers for GBM, especially when associated with preoperative NLR, PLR and LMR.

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Cortactin and fascin-1 regulate extracellular vesicle release by controlling endosomal trafficking or invadopodia formation and function

Scientific Reports ◽

10.1038/s41598-018-33868-z ◽

2018 ◽

Vol 8 (1) ◽

Cited By ~ 15

Author(s):

Els Beghein ◽

Delphine Devriese ◽

Evy Van Hoey ◽

Jan Gettemans

Keyword(s):

Extracellular Vesicle ◽

Endosomal Trafficking ◽

Vesicle Release ◽

And Function ◽

Invadopodia Formation

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Genome-wide interrogation of extracellular vesicle biology using barcoded miRNAs

eLife ◽

10.7554/elife.41460 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 7

Author(s):

Albert Lu ◽

Paulina Wawro ◽

David W Morgens ◽

Fernando Portela ◽

Michael C Bassik ◽

...

Keyword(s):

Nucleic Acid ◽

Molecular Mechanisms ◽

Membrane Vesicles ◽

Extracellular Vesicle ◽

Biologically Active ◽

Vesicle Release ◽

Guide Rnas ◽

Disease States ◽

Genome Wide ◽

First Time

Extracellular vesicles mediate transfer of biologically active molecules between neighboring or distant cells, and these vesicles may play important roles in normal physiology and the pathogenesis of multiple disease states including cancer. However, the underlying molecular mechanisms of their biogenesis and release remain unknown. We designed artificially barcoded, exosomal microRNAs (bEXOmiRs) to monitor extracellular vesicle release quantitatively using deep sequencing. We then expressed distinct pairs of CRISPR guide RNAs and bEXOmiRs, enabling identification of genes influencing bEXOmiR secretion from Cas9-edited cells. This approach uncovered genes with unrecognized roles in multivesicular endosome exocytosis, including critical roles for Wnt signaling in extracellular vesicle release regulation. Coupling bEXOmiR reporter analysis with CRISPR-Cas9 screening provides a powerful and unbiased means to study extracellular vesicle biology and for the first time, to associate a nucleic acid tag with individual membrane vesicles.

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