P02.30.A MODELING GLIOBLASTOMA-EXTRACELLULAR MATRIX INTERACTIONS IN VIVO USING ZEBRAFISH AND ATOMIC FORCE MICROSCOPY

Abstract BACKGROUND Glioblastoma (GB) remains the most aggressive adult brain tumor, marked by rapid invasion and poor patient prognosis. While research has traditionally focused on genetic drivers of GB, there is growing recognition that the extracellular matrix (ECM) alterations play a central role in tumor progression. GB is known to remodel its microenvironment, increasing tissue stiffness to as much as 35 kPa - conditions that have been shown in vitro to promote tumor cell migration and invasion. However, in vivo models that allow mechanistic investigation of ECM-GB interactions remain technically challenging. MATERIAL AND METHODS To address this, we established an orthotopic xenograft model in zebrafish (Danio rerio) larvae to study GB behavior within a live, developing brain. Human GB cells (U87, GG16, and GSC23) were transplanted into the midbrain of 3-day-old larvae, and their migration and invasion were monitored in real-time using confocal and light-sheet microscopy. Whilst tumor implantation had proved difficult, we in parallel used atomic force microscopy (AFM) to characterize the mechanical properties of healthy larval brains. RESULTS At 5 days post-fertilization, we found average stiffness values of 6.8 ± 2.8 kPa. High-resolution 3D reconstructions from AFM data revealed regional variation in brain stiffness and provided nanoscale insights into the brain’s microarchitecture regarding height and adhesion properties. CONCLUSION Our current focus involves experimentally manipulating multiple ECM-related genes in GB cells prior to xenografting (Fibronectin, collagen, and hyaluronic acid regulators). This integrative approach bridges biomechanics with tumor biology, with the ultimate aim of linking gene level ECM modulation to in vivo tumor stiffening and tumor behavior. This model will highlight zebrafish as a powerful and scalable model for investigating GB-ECM interaction and potentially uncover new therapeutic targets.