Authors: Amanda Muhs Saratsis, MD; Jin Qi, MD, PhD; Tina Huang, MS; Patrick Ozark, BA; Rintaro Hashizume, MD, PhD; Elizabeth Bartom, PhD; Charles James, PhD; Ali Shilatifard, PhD; Amanda Saratsis, MD (Chicago, IL)

INTRODUCTION: Pediatric diffuse midline glioma (DMG) and high-grade glioma (HGG) are the most morbid pediatric cancers. Tenascin-C (TNC) is an extracellular matrix glycoprotein that mediates cell-cell and cell-matrix interactions during central nervous system development. Our group recently found TNC overexpression in pediatric HGG and DMG CSF, tissue and cell lines. Here, we describe effects of modulated TNC expression in vitro and in vivo on HGG and DMG tumorigenesis.

METHODS: The effects of altered TNC expression in pediatric DMG (n=7), supratentorial HGG (n=3) cell lines were evaluated and compared to normal human astrocytes (n=1) and human stem cells (n=1) as controls. Lentiviral-mediated TNC shRNA and cDNA transfection was performed, and efficacy confirmed by qPCR and western blot. Cell proliferation, migration, adhesion, and viability were assessed. Comparative whole transcriptome analysis (RNA-Seq, Illumina Platform) was performed to determine effects of altered TNC on global gene expression in vitro (Ingenuity Pathways Analysis).

RESULTS: TNC knockdown was associated with decreased cell proliferation and migration in vitro, reversed with exogenous TNC administration, and differed by endogenous TNC expression, ACVR1 and histone H3 mutation status. TNC cDNA transfection increased tumor proliferative and migratory cell capacity. Functional pathways analysis implicated targetable signaling pathways associated with altered TNC expression, and suggest TNC may serve as a biomarker of EZH2 and BRD inhibition. Tumor formation was confirmed in mouse xenografts via bioluminescence imaging, with comparative analysis of tumor growth and animal survival currently underway.

CONCLUSION: Our findings suggest TNC expression levels are affected by ACVR1 and H3F3A mutation status, change in response to molecularly targeted therapies, and may contribute to DIPG tumorgenicity through activation of PDGF and VEGF signaling. In vitro studies animal xenografts are underway. Overall, these data suggest TNC may serve as a disease biomarker and potential therapeutic target for pediatric DMG.