ANGI-14. GLIOBLASTOMA PROGRESSION FROM TRANSCRIPTOMIC REGULATION TO BRAIN FUNCTIONAL CONNECTIVITY

Abstract Glioblastoma progression features extensive dissemination of cancer cells throughout the brain. Limited by neuroimaging resolution and lack of specific markers, malignant cell infiltration is clinically undetectable. This makes complete tumor removal impossible and relapse inevitable. During their exodus from the tumor core, glioblastoma cells are traversing distinct brain territories influencing their phenotypic cell state. To better understand the cellular and molecular dynamics of glioblastoma invasion, we profiled glioblastoma patient avatars and clinical samples using total, single-cell and spatial transcriptomics. In addition to gene expression data, active intracellular pathways of glioblastoma invasion were screened with high throughput activated kinase assays (Pamgene). We identified that human glioblastoma stem cells exhibiting proneural (PN) and neuron-progenitor like (NPC) transcriptomic traits in vitro formed highly infiltrative and connected xenografts in patient avatars. Consistent with this, PN/NPC glioblastoma stem cells cultured on primary hippocampal neurons exhibited higher connectivity and motility compared to mesenchymal-like cells. Leveraging single-cell and spatial transcriptome analytics of avatar and patient samples, we performed cell clustering, annotation, pseudotime and RNA velocity analyses to associate glioblastoma phenotypic cell states to cellular functions. We found that invasive glioblastoma cells re-activated genes and pathways related to neuron progenitor maturation when transitioning from the tumor core to the brain microenvironment. To verify whether invasive PN/NPC glioblastoma cell integration would alter brain functionality, we performed preclinical live cerebral blood volume recordings using noninvasive brain functional ultrasounds (fUS). As glioblastomas progressed, we detected a gradual alteration of brain functional connectivity. Modifications happened in brain areas featuring minimal, yet consistent glioblastoma cell colonization, identified by post-mortem pathology analyses. Altogether, our data shed light on the importance of glioblastoma cell adaptability and brain integration during the metastatic progression. They also provide a framework for detecting invisible clinical features such as tumor invasion, focusing on brain functional connectivity assessments.