Glioblastoma biomarkers in urinary extracellular vesicles reveal the potential for a 'liquid gold' biopsy.

BACKGROUND: Biomarkers that reflect glioblastoma tumour activity and treatment response are urgently needed to help guide clinical management, particularly for recurrent disease. As the urinary system is a major clearance route of circulating extracellular vesicles (EVs; 30-1000 nm nanoparticles) we explored whether sampling urinary-EVs could serve as a simple and non-invasive liquid biopsy approach for measuring glioblastoma-associated biomarkers. METHODS: Fifty urine specimens (15-60 ml) were collected from 24 catheterised glioblastoma patients immediately prior to primary (n = 17) and recurrence (n = 7) surgeries, following gross total resection (n = 9), and from age/gender-matched healthy participants (n = 14). EVs isolated by differential ultracentrifugation were characterised and extracted proteomes were analysed by high-resolution data-independent acquisition liquid chromatography tandem mass spectrometry (DIA-LC-MS/MS). RESULTS: Overall, 6857 proteins were confidently identified in urinary-EVs (q-value ≤ 0.01), including 94 EV marker proteins. Glioblastoma-specific proteomic signatures were determined, and putative urinary-EV biomarkers corresponding to tumour burden and recurrence were identified (FC ≥ | 2 | , adjust p-val≤0.05, AUC > 0.9). CONCLUSION: In-depth DIA-LC-MS/MS characterisation of urinary-EVs substantiates urine as a viable source of glioblastoma biomarkers. The promising 'liquid gold' biomarker panels described here warrant further investigation.

Understanding the Epitranscriptome for Avant-Garde Brain Tumour Diagnostics.

RNA modifications are diverse, dynamic, and reversible transcript alterations rapidly gaining attention due to their newly defined RNA regulatory roles in cellular pathways and pathogenic mechanisms. The exciting emerging field of 'epitranscriptomics' is predominantly centred on studying the most abundant mRNA modification, N6-methyladenine (m6A). The m6A mark, similar to many other RNA modifications, is strictly regulated by so-called 'writer', 'reader', and 'eraser' protein species. The abundance of genes coding for the expression of these regulator proteins and m6A levels shows great potential as diagnostic and predictive tools across several cancer fields. This review explores our current understanding of RNA modifications in glioma biology and the potential of epitranscriptomics to develop new diagnostic and predictive classification tools that can stratify these highly complex and heterogeneous brain tumours.

Understanding the extracellular vesicle surface for clinical molecular biology.

Extracellular vesicles (EVs) are lipid-membrane enclosed nanoparticles that play significant roles in health and disease. EVs are abundant in body fluids and carry an array of molecules (proteins, lipids, nucleic acids and glycans) that reflect the identity and activity of their cell-of-origin. While the advent of high throughput omics technologies has allowed in-depth characterisation of EV compositions, how these molecular species are spatially distributed within EV structures is not well appreciated. This is particularly true of the EV surface where a plethora of molecules are reported to be both integral and peripherally associated to the EV membrane. This coronal layer or 'atmosphere' that surrounds the EV membrane contributes to a large, highly interactive and dynamic surface area that is responsible for facilitating EV interactions with the extracellular environment. The EV coronal layer harbours surface molecules that reflect the identity of parent cells, which is likely a highly valuable property in the context of diagnostic liquid biopsies. In this review, we describe the current understanding of the mechanical, electrostatic and molecular properties of the EV surface that offer significant biomarker potential and contribute to a highly dynamic interactome.

Coordinated activation of a cluster of MMP genes in response to UVB radiation.

Ultraviolet (UV) B radiation is a dangerous environmental stressor, which can lead to photoaging, inflammation, immune suppression and tumour formation. A recent report has shown the transcriptional activation of several skin-specific genes including matrix metalloproteases (MMPs) in response to UV irradiation. Here, we use a novel human keratinocyte model, HKerE6SFM, to demonstrate that UVB activates the transcription of most members of the 11q22.3 MMP gene cluster including MMP13, MMP12, MMP3, MMP1 and MMP10. Curiously, the expression of the well-characterized UVB-inducible MMP9, which is located outside of the cluster, remains unchanged. In accordance with the increased expression of the MMP gene cluster upon UVB irradiation, RNA polymerase II showed increased occupancy at their promoters following UVB irradiation. The results also demonstrate increased acetylated histone H3K9 levels at the promoters of the MMP13, MMP12, MMP3, MMP1 and MMP10 genes. These findings suggest a coordinated transcriptional activation of genes in the MMP cluster at 11q22.3 and that acetylation of histone H3 at lysine 9 has an important role in the UVB-dependent enhancement of transcription of MMP genes in this region.

Pediatric brain tumor cells release exosomes with a miRNA repertoire that differs from exosomes secreted by normal cells.

High-grade gliomas (HGGs) are very aggressive brain tumors with a cancer stem cell component. Cells, including cancer stem cells, release vesicles called exosomes which contain small non-coding RNAs such as microRNAs (miRNAs). These are thought to play an important role in cell-cell communication. However, we have limited knowledge of the types of exosomal miRNAs released by pediatric HGG stem cells; a prerequisite for exploring their potential roles in HGG biology. Here we isolated exosomes released by pediatric glioma stem cells (GSCs) and compared their repertoire of miRNAs to genetically normal neural stem cells (NSCs) exosomes, as well as their respective cellular miRNA content. Whereas cellular miRNAs are similar, we find that the exosomal miRNA profiles differ between normal and tumor cells, and identify several differentially expressed miRNAs. Of particular interest is miR-1290 and miR-1246, which have previously been linked to 'stemness' and invasion in other cancers. We demonstrate that GSC-secreted exosomes influence the gene expression of receiving NSCs, particularly targeting genes with a role in cell fate and tumorigenesis. Thus, our study shows that GSCs and NSCs have similar cellular miRNA profiles, yet differ significantly in the repertoire of exosomal miRNAs and these could influence malignant features of HGG.