HGG-18. SINGLE CELL RNA-SEQ ANALYSIS OF PEDIATRIC HIGH-GRADE GLIOMA PATIENT SAMPLES IDENTIFIES COMMON GLIAL AND IMMUNE CELL TYPES THAT HELP TO EXPLAIN HETEROGENEITY AND TUMORIGENESIS

Abstract

Abstract BACKGROUND Pediatric high-grade gliomas (PHGG) are aggressive, undifferentiated central nervous system (CNS) tumors. PHGG comprises subtypes that differ phenotypically, histologically and by cellular composition. Despite intensive research, overall survival rates have remained for decades around 2% for diffuse midline glioma (DMG) and 20% for non-DMG PHGG. We hypothesized that PHGG’s heterogeneity, a key difficulty in devising therapies, results from functional and lineage differences in tumor cell types. METHODS Using 19 PHGG samples from our institution’s pediatric brain tumor bank, we constructed a single-cell RNA-Seq (scRNA-Seq) dataset that included tumor and immune cells, generated predicted cell types by mapping the scRNA-Seq data onto known cell types, and performed differential gene expression and geneset enrichment analyses. We acquired bulk RNA-Seq and DNA methylation data to characterize overall tumor properties. RESULTS Samples consisted of several principal tumor cell types, including cells with astrocyte characteristics, oligodendrocyte progenitor cell (OPC) characteristics, and cells that expressed OPC markers but had slight enrichment in mesenchymal and inflammatory gene expression (OPC-like/Mes cells). We also found a potential stemlike population that expressed neural stem cell markers. The predicted astrocytes, OPCs and OPC-like/Mes cells differed in their gene expression profiles, pathway enrichment, and tendency to proliferate as assessed from expression of cell cycle genes. A combined microglia/macrophage population, strongly enriched in mesenchymal and inflammatory gene expression, may serve to induce inflammatory gene expression in tumor cells. Key conclusions from the scRNA-Seq analysis were validated using multi-channel immunofluorescence staining. CONCLUSIONS PHGG comprises a small stemlike population and varying proportions of proliferating glial lineage tumor cells. CNS-resident microglia/macrophages may trigger mesenchymal gene expression in PHGG. We will use our results to investigate the roles of PHGG’s diverse tumor cell populations in resistance to radiotherapy and to develop treatments to target individual cell types comprising PHGG.