DIPG-83. RBPJ-SIX1 AS A TUMOR SUPPRESSOR AXIS IN DIFFUSE INTRINSIC PONTINE GLIOMA

Abstract BACKGROUND Diffuse intrinsic pontine glioma (DIPG) is a rare, incurable pediatric brain tumor that arises in the brainstem. Despite decades of clinical trials, no treatments have proven effective at prolonging survival for children with this disease. A deeper insight into the epigenetic and genetic changes driving DIPG are imperative for identifying therapeutic targets. A defining feature of DIPG is the histone H3 lysine to methionine mutation at residue 27 (H3K27M), which prevents H3K27 trimethylation. The H3.1K27M mutation, specifically, co-segregates with mutations in the Activin A receptor ACVR1. Tumor cells with both mutations have significantly higher levels of the transcripts and proteins that inhibit proneural gene expression in the NOTCH signaling pathway. METHODS We used the replication-competent avian sarcoma-leukosis virus long terminal repeat with splice acceptor/tumor virus A (RCAS/Tv-a) system to evaluate the role of various NOTCH signaling proteins in a genetically engineered mouse model of DIPG. We also performed RNA sequencing of specific NOTCH signaling protein knockout DIPG models. RESULTS Knocking out Hairy of Enhancer Split-1 (Hes-1) in combination with H3K27M and ACVR1 mutations did not significantly reduce survival compared to control. However, knocking out Recombination Signal Binding Protein for Immunoglobulin kappa J region (RBPjk)—a transcriptional regulator upstream of Hes-1—did significantly reduce survival compared to wild-type RBPjk expression. RNA sequencing of RBPjk knockout tumors revealed that Sine Oculis-related Homeobox 1 (Six1) was the most top most differentially expressed gene. To assess whether Six1 downregulation drives DIPG tumorigenesis in RBPjk knockout tumors, we overexpressed Six1 in a murine model of DIPG with H3.3K27M and p53 loss. Six1 overexpression did not significantly prolong survival or decrease tumor grade but trended toward significance. However, Six1 overexpression significantly increased tumor cell proliferation. CONCLUSIONS These results indicate that Six1 could function as a potential tumor suppressor in DIPG, necessitating further exploration.