Mesenchymal-like glioma cells are enriched in the gelatin methacrylate hydrogels

Glioblastoma is the most lethal primary malignant brain tumor in adults. Simplified two-dimensional (2D) cell culture and neurospheres in vitro models fail to recapitulate the complexity of the tumor microenvironment, limiting its ability to predict therapeutic response. Three-dimensional (3D) scaffold-based models have emerged as a promising alternative for addressing these concerns. One such 3D system is gelatin methacrylate (GelMA) hydrogels, which can be used for modeling the glioblastoma microenvironment. We characterized the phenotype of patient-derived glioma cells cultured in GelMA hydrogels (3D-GMH) for their tumorigenic properties using invasion and chemoresponse assays. In addition, we used integrated single-cell and spatial transcriptome analysis to compare cells cultured in 3D-GMH to cells in vivo . Finally, we assessed tumor-immune cell interactions with a macrophage infiltration assay and a cytokine array. We show that cells cultured in 3D-GMH develop a mesenchymal-like cellular phenotype found in perivascular and hypoxic regions present in the core of the tumor, and recruit macrophages by secreting cytokines in contrast to the cells grown as neurospheres that match the phenotype of cells of the infiltrative edge of the tumor.