To investigate the ability of patient derived glioma stem cells organoids to model the intrinsic glioblastoma cells immune program in the dish to define the molecular, metabolic, and genetic organization of the initial tumor without contaminant immune cells and blood vessels.

Glioblastoma stem cells (GSCs) are highly self-renewing, resistant to therapy, and can form lethal tumors. Tumor organoids have been developed to study tumor evolution, and while GSCs can form organoids from glioblastoma multiforme, our understanding of their intrinsic immune, metabolic, genetic, and molecular programs is limited.

Generation of glioblastoma organoids (GBMOs) from patient’s resected GBM surgical tissue and iPSC control n=6. A total of 314 organoids were generated for the study. Deep characterization of genetic and molecular programs was performed with RNA microarrays. Genomic microarrays were used to determine copy number variants and mutational landscape of GBMOs. Bioinformatic analysis was used for mutation and gene expression analysis.  Metabolic analysis of GBMOs was performed with the Seahorse assay. Confocal microscopy was used for confirmation of molecular pathways.

GBMOs develop into complex 3D tissues with unique self-organization similar to in vivo GBM and recapitulate the presence of two cell populations thought to drive GBM progression, SATB2⁺ and HOPX⁺ progenitors. Despite being devoid of immune cells, transcriptomic analysis across GBMOs revealed an immune-like molecular program, enriched in cytokine, antigen presentation and processing, T-cell receptor inhibitors, and interferon genes. We determined that SATB2⁺ and HOPX⁺ populations contribute to this immune and interferon landscape in GBM in vivo and GBMOs.

Our work deepens our understanding of the intrinsic molecular and cellular architecture of GSC-derived GBMOs and defines a novel GBMOs intrinsic immune-like program, this work will enable future translational and mechanistic of intrinsic immune program of GBM in patient specific manner.