Gliomas are a diverse group of brain tumors comprised of malignant cells (‘tumor’ cells) and non-malignant ‘normal’ cells, including neural (neurons, glia), inflammatory (microglia, macrophage) and vascular cells. Tumor heterogeneity arises in part because, within the glioma mass, both ‘tumor’ and ‘normal’ cells secrete factors that form a unique microenvironment to influence tumor progression. Extracellular vesicles (EVs) are critical mediators of intercellular communication between immediate cellular neighbors and distantly located cells in healthy tissues/organs and in tumors, including gliomas. EVs mediate cell–cell signaling as carriers of nucleic acid, lipid and protein cargo, and their content is unique to cell types and physiological states. EVs secreted by non-malignant neural cells have important physiological roles in the healthy brain, which can be altered or co-opted to promote tumor progression and metastasis, acting in combination with glioma-secreted EVs. The cell-type specificity of EV content means that ‘vesiculome’ data can potentially be used to trace the cell of origin. EVs may also serve as biomarkers to be exploited for disease diagnosis and to assess therapeutic progress. In this review, we discuss how EVs mediate intercellular communication in glioma, and their potential role as biomarkers and readouts of a therapeutic response.

Direct neuronal reprogramming, the process whereby a terminally differentiated cell is converted into an induced neuron without traversing a pluripotent state, has tremendous therapeutic potential for a host of neurodegenerative diseases. While there is strong evidence for astrocyte-to-neuron conversion in vitro, in vivo studies in the adult brain are less supportive or controversial. Here, we set out to enhance the efficacy of neuronal conversion of adult astrocytes in vivo by optimizing the neurogenic capacity of a driver transcription factor encoded by the proneural gene Ascl1. Specifically, we mutated six serine phospho-acceptor sites in Ascl1 to alanines (Ascl1^SA6) to prevent phosphorylation by proline-directed serine/threonine kinases. Native Ascl1 or Ascl1^SA6 were expressed in adult, murine cortical astrocytes under the control of a glial fibrillary acidic protein (GFAP) promoter using adeno-associated viruses (AAVs). When targeted to the cerebral cortex in vivo, mCherry⁺ cells transduced with AAV8-GFAP-Ascl1^SA6-mCherry or AAV8-GFAP-Ascl1-mCherry expressed neuronal markers within 14 days post-transduction, with Ascl1^SA6 promoting the formation of more mature dendritic arbors compared to Ascl1. However, mCherry expression disappeared by 2-months post-transduction of the AAV8-GFAP-mCherry control-vector. To circumvent reporter issues, AAV-GFAP-iCre (control) and AAV-GFAP-Ascl1 (or Ascl1^SA6)-iCre constructs were generated and injected into the cerebral cortex of Rosa reporter mice. In all comparisons of AAV capsids (AAV5 and AAV8), GFAP promoters (long and short), and reporter mice (Rosa-zsGreen and Rosa-tdtomato), Ascl1^SA6 transduced cells more frequently expressed early- (Dcx) and late- (NeuN) neuronal markers. Furthermore, Ascl1^SA6 repressed the expression of astrocytic markers Sox9 and GFAP more efficiently than Ascl1. Finally, we co-transduced an AAV expressing ChR2-(H134R)-YFP, an optogenetic actuator. After channelrhodopsin photostimulation, we found that Ascl1^SA6 co-transduced astrocytes exhibited a significantly faster decay of evoked potentials to baseline, a neuronal feature, when compared to iCre control cells. Taken together, our findings support an enhanced neuronal conversion efficiency of Ascl1^SA6 vs. Ascl1, and position Ascl1^SA6 as a critical transcription factor for future studies aimed at converting adult brain astrocytes to mature neurons to treat disease.

Isocitrate dehydrogenase (IDH) mutant gliomas, including oligodendroglioma (IDH-O) and astrocytoma (IDH-A), have signature slow-growth rates that are poorly understood. Here, we reveal that SMPD3, a ceramide-producing sphingomyelinase implicated as a tumor suppressor gene and involved in extracellular vesicle biogenesis, suppresses IDH-mutant tumor growth via autocrine and paracrine actions. In patients with IDH-mutant gliomas, higher SMPD3 expression levels correlate with longer survival, consistent with ceramide acting as an anti-oncometabolite. SMPD3 knock-down in patient-derived IDH-O cells enhances proliferation cell-autonomously in 2D-culture and 3D-human cerebral organoids, and accelerates tumor growth in mouse orthotopic xenografts. Supporting paracrine actions, IDH-O-derived extracellular vesicles, enriched in ribosomal proteins, induce astrocytic death in vitro . Furthermore, non-neoplastic glia in IDH-O tumors proliferate abnormally yet undergo apoptosis, concomitant with the acquisition of a translation-enriched transcriptional signature by tumor-associated oligodendrocytes. SMPD3 thus suppresses IDH-mutant glioma growth cell-autonomously and phenotypically alters normal glia via extracellular vesicle biogenesis and paracrine actions.