LAUSR

Abstract Pediatric Glioblastoma (pGBM) and Diffuse Intrinsic Pontine Glioma (DIPG) are highly heterogeneous brain tumors which we demonstrated are comprised by distinct sub-clones interacting in a functional network. Exosomes are known to mediate the crosstalk between tumor and its microenvironment. Based on this, we aimed to investigate the role of exosomes in mediating pGBM and DIPG inter-clonal communication. By using optical barcoding for single cell-tracking, we generated two bulk multicolor patient derived-cell lines (one DIPG H3.3K27M and one pGBM histone WT) from which we obtained two and five single cell-derived clones respectively. The sub-clones demonstrated significantly phenotypic differences in terms of morphology, growth, adhesion, migration and invasion properties. In particular, co-culture experiments, with the two most different clones for both cell-lines, confirmed the cell-cell interaction key role in driving their more aggressive phenotype. Furthermore, we found that pGBM and DIPG sub-clones release exosomes which are actively and differentially up-taken by individual clones. Analysis of the exosomal microRNAs showed a different profile between the two selected clones in each cell-line. In particular, we found a pool of five upregulated microRNAs in 1C5 clone (DIPG cell-line) strongly associated to Wnt-signaling and PI3K-AKT pathway. Similarly, a pool of five upregulated microRNAs for 5E2 clone (pGBM cell-line) were found associated with focal adhesion and PI3K-AKT pathway. Our study may provide novel therapeutic strategies by interfering with the exosome-mediated inter-clonal communication in pGBM and DIPG.