Lgg-15. single-nucleus and bulk rna sequencing of pediatric pilocytic astrocytomas reveals cns location-associated tumor cell and microenvironmental heterogeneity

Abstract BACKGROUND Pilocytic astrocytomas are the most common pediatric central nervous system (CNS) tumors. While the overall survival is generally favorable, a substantial part of the patients relapse, resulting in long-term complications. Additionally, pilocytic astrocytomas show limited genetic heterogeneity and are often driven by a single molecular event. The role of the tumor microenvironment (TME) in disease progression is yet unknown. Here, we aimed to delineate the composition of the tumor in pilocytic astrocytomas originating from different CNS locations. METHODS We collected 112 tumor samples, out of which a subset (n = 10) was used for single-nucleus RNA-sequencing (10X Genomics). These tumor samples originated from various CNS locations: posterior fossa (n = 64), supratentorial (n = 38) and spinal (n = 10). Clinical characteristics, DNA methylation profiles, whole exome/genome sequencing and bulk RNA sequencing data were available for most of these samples. RESULTS Single-nucleus RNA-sequencing revealed substantial heterogeneity amongst both tumor and TME cells, which mainly associated with tumor location. While a large proportion of tumor cells expressed gene signatures related to oligodendrocyte precursor cells (OLIG1/2, PDGFRA), a subset of tumor cells was characterized by high expression of astrocyte genes (AQP4, GFAP). Moreover, we defined multiple populations of lymphocytes and myeloid cells, with the latter constituting the vast majority of non-neoplastic cells. Deconvolution of the bulk RNA sequencing data showed heterogeneous involvement of the TME. Additionally, the relative abundances of TME cells varied considerably across CNS locations. We are currently correlating the relative proportions of tumor and TME cell populations with clinicopathological data. CONCLUSION Single-nucleus RNA-sequencing demonstrated transcriptional differences in both tumor cell states and TME cell populations, which associated with CNS location. Future research should elucidate the importance of the cellular composition of the tumor in pilocytic astrocytomas.