HGG-17. SINGLE-CELL AND SPATIAL ANALYSES DECIPHER THE UNIQUE INVASIVE GROWTH PATTERN OF GLIOMATOSIS CEREBRI

Gliomatosis cerebri (GC) is a rare, lethal glioma that is radiologically diagnosed and characterised by its diffuse infiltration throughout the cerebral lobes of the brain. It is no longer recognised as a separate entity by the WHO classification, but its growth pattern and invasive phenotype differ from other types of glioma. In order to understand what underlies this unique presentation, we sought to unravel these complex tumours in children through single cell and spatial approaches. 26 paediatric cases (1.3-19 years, median=11.3) were collected for DNA methylation, whole exome sequencing and single cell RNA-sequencing. Of these, 16 cases were used for Imaging Mass Cytometry (IMC). 5 patient-derived cell lines were used to model migration and invasion in vitro, whereby we have also been able to explore gene expression differences associated with the invading cells by RNAseq. We analyzed the transcriptional landscape of all 26 GC patients at single-cell resolution, and identified shared tumor cellular programs of glial and neuronal lineage. We observed notable heterogeneity of neuronal lineage tumor cells that were composed of different states, such as metabolic active and synaptogenic states. We described tumor cell states associated with higher invasivity, as corroborated from integrated analysis with our in vitro invasion-associated gene expression profiles. Next, we stratified all tumors by their DNA methylation profiles, and defined 16 archetypal GC cases, for which we show specific transcriptional features. Furthermore, we analyzed multiple regions of interest within FFPE patient tissue sections by IMC and demonstrate spatial patterns in the interaction between the tumour and normal brain tissue interface, and inter-regional heterogeneity for stem, invasive and proliferative markers. Together, we dissect the characteristic invasive phenotype of GC at multiple molecular, spatial and functional layers, thereby providing the basis for modelling and therapeutic avenues much needed for this lethal disease.