CSIG-36. NOTCH3 DRIVES MENINGIOMA TUMORIGENESIS AND RESISTANCE TO RADIOTHERAPY

Abstract

Abstract There are no approved targeted therapies for meningiomas and the cell types underlying meningeal tumorigenesis are incompletely understood. To address these limitations, we performed single-cell RNA sequencing of 57,114 cells from 8 human meningiomas and 54,607 cells from 3 canine meningiomas. Pseudotime, gene ontology, and copy number variant analyses revealed a population of pericyte-like meningioma cells that were conserved across human and canine tumors and were enriched in expression of Notch3 and other cancer stem cell genes. Deconvolution of cell types from bulk RNA sequencing and DNA methylation profiling of 200 human meningiomas integrated with immunohistochemistry (IHC), immunofluorescence (IF), and RNAScope demonstrated Notch3+ pericytes and Notch3 expression were enriched in high grade or Immune-enriched meningiomas, which were distinguished from other meningioma DNA methylation groups by genes driving vasculature development. IHC and IF of human meninges integrated with lineage tracing approaches using Notch3-CreERT2 ROSAmT/mG alleles in mice demonstrated Notch3 expression was restricted to the perivascular stem cell niche during meningeal development and homeostasis. Mice harboring Notch3-CreERT2 Nf2fl/fl alleles developed meningeal hyperproliferation. Overexpression of constitutively activated Notch3 (Notch3AICD) in Immune-enriched human meningioma cells increased the expression of cancer stem cell genes, driving clonogenic growth in vitro, limiting dilution tumor-initiating capacity in vivo, and resistance to radiotherapy in vivo. A selective Notch3 neutralizing antibody (αNRR3) blocked meningioma cell proliferation and expression of Notch3 target genes, inhibiting meningioma xenograft growth and prolonging overall survival. Single-cell RNA sequencing of 187,366 cells from meningioma xenografts after αNRR3 or radiotherapy treatment ± Notch3AICD overexpression revealed distinct meningioma cell-intrinsic or cell-extrinsic mechanisms driving responses to radiotherapy or αNRR3, respectively. Combined treatment with αNRR3 and radiotherapy additively blocked meningioma xenograft growth and extended survival benefit. In sum, these data shed light on a novel cell type, molecular mechanism, and therapeutic vulnerability in the most common primary intracranial tumor.