494 Multi-Omics Integration and Mapping of Invasion Markers Reveal Tumor-Specific Druggable Targets in Primary Human Glioblastoma

INTRODUCTION: Glioblastoma (GBM) recurrence is the rule rather than the exception. GBM cells invading healthy brain and the tumor microenvironment are believed to be responsible for tumor recurrence. However, little is known about molecular alterations in the primary tumor that drive this progression. Multi-omics studies have revealed specific tissue domains of GBM that could have higher invasive profiles in primary (pGBM) and recurrent (rGBM) tumors. METHODS: Paraffin-embedded sample blocks from matched primary (n = 8) and recurrent (n = 10) GBM were utilized. Spatial transcriptomics (10x-Visium) and in situ molecular imaging (10x-Xenium) were performed. Matching fresh-frozen samples were processed for single-nucleus RNAseq (10x-Chromium). Visium spots were clustered together, and cells were annotated based on gene expression. Differential gene expression analysis, pathway enrichment analysis, and downstream analyses were performed. RESULTS: Spatial transcriptomics spots were grouped by transcriptomic similarity. In pGBM we identified that cluster 2 is significantly enriched with high expression of Neurocan (NCAN) (p < 0.001), a proteoglycan member of the brain extracellular matrix (ECM) that is only expressed during development and is absent in the adult brain. NCAN is known to increase proliferation and migration in glioma cells via the RHOA/ROCK pathway, representing an important target for GBM. Comparing spatial transcriptomic spots with high NCAN vs. Low NCAN shows upregulation of RHOA/ROCK and cell motility genes. In-situ molecular imaging confirms the distribution of ECM and cell motility gene transcripts. CONCLUSIONS: Patient-matched GBM samples can differ substantially within and between patients, but molecular signatures associated with tumor progression remain constant in subpopulations of cells. NCAN is a brain extracellular matrix component overexpressed in distinct tissue domains in pGBM with implications in cell invasion, representing a potential target that could hinder tumor recurrence.