IMMU-18. GLIOBLASTOMA-DERIVED EXTRACELLULAR VESICLES INDUCE DRAMATIC CHANGES IN THE TRANSCRIPTOMIC LANDSCAPE OF MONOCYTES

Abstract

Abstract Glioblastoma (GBM) is the most common and deadly primary brain tumor. Novel therapeutic strategies are urgently needed to improve outcomes, but a number of disease-specific barriers pose challenges to innovation. Tumor-mediated immunosuppression is one such hurdle, and a growing body of evidence suggests that GBM-derived extracellular vesicles (EVs) play an important role in host immunosuppression. GBM-derived EVs have been shown to induce the formation of immunosuppressive monocytes, including myeloid-derived suppressor cells. Work by our group and others has increasingly shown that these immunosuppressive monocytes are a heterogenous group, and that many constellations of surface markers are inadequate to capture the changes wrought by EVs. In order to better understand the effects of GBM-EVs on monocytes, we conducted RNA-seq analysis on monocytes collected from four healthy donors treated with GBM-derived EVs harvested by ultracentrifugation from the patient-derived BT116 cell line. Following 72h of EV conditioning, total RNA was harvested from treated monocytes and untreated controls. RNA-seq was performed using the Illumina HiSeq4000 platform with paired end index reads. Analysis was performed using RNA STAR and the hg19 ENCODE reference sequence. Differential expression analysis was performed using DESeq2. Genes with a false-discovery rate (FDR)-corrected P value < 0.05 and a log2 fold-change value of >|1| were considered significantly different between groups. Pathway analysis was performed using ClueGO in Cytoscape (GO term fusion on, p< 0.05). Unsupervised clustering analysis of the top 500 most differentially-expressed genes demonstrated grouping of BT116 EV-treated monocytes together versus untreated monocytes. Pathway analysis upregulated genes in pathways important for heparan sulfate proteoglycan synthesis and cholesterol synthesis, which could potentially point to positive regulation of EV uptake. Downregulated pathways included regulation of T cell differentiation and chemoattractant activity, underscoring the induction of a potentially immunosuppressive phenotype by GBM-derived EVs.