Glioblastoma (GBM) is a deadly disease with a need for deeper understanding and new therapeutic approaches. The microenvironment of glioblastoma has previously been shown to guide glioblastoma progression. In this study, astrocytes were investigated with regard to their effect on glioblastoma proliferation through correlative analyses of clinical samples and experimental in vitro and in vivo studies. Co‐culture techniques were used to investigate the GBM growth enhancing potential of astrocytes. Cell sorting and RNA sequencing were used to generate a GBM‐associated astrocyte signature and to investigate astrocyte‐induced GBM genes. A NOD scid GBM mouse model was used for in vivo studies. A gene signature reflecting GBM‐activated astrocytes was associated with poor prognosis in the TCGA GBM dataset. Two genes, periostin and serglycin, induced in GBM cells upon exposure to astrocytes were expressed at higher levels in cases with high “astrocyte signature score”. Astrocytes were shown to enhance glioblastoma cell growth in cell lines and in a patient‐derived culture, in a manner dependent on cell–cell contact and involving increased cell proliferation. Furthermore, co‐injection of astrocytes with glioblastoma cells reduced survival in an orthotopic GBM model in NOD scid mice. In conclusion, this study suggests that astrocytes contribute to glioblastoma growth and implies this crosstalk as a candidate target for novel therapies.

The microenvironment and architecture of peritumoral tissue have been suggested to affect permissiveness for infiltration of malignant cells. Astrocytes constitute a heterogeneous population of cells and have been linked to proliferation, migration, and drug sensitivity of glioblastoma (GBM) cells. Through double‐immunohistochemical staining for platelet‐derived growth factor receptor α (PDGFRα) and glial fibrillary acidic protein (GFAP), this study explored the intercase variability among 45 human GBM samples regarding density of GFAP+ peritumoral astrocytes and a subset of GFAP+ peritumoral astrocyte‐like cells also expressing PDGFRα. Large intercase variability regarding the total peritumoral astrocyte density and the density of PDGFRα+/GFAP+ peritumoral astrocyte‐like cells was detected. DNA fluorescence in situ hybridization analyses for commonly altered genetic tumor markers supported the interpretation that these cells represented a genetically unaffected host cell subset referred to as PDGFRα+/GFAP+ peritumoral astrocytes. The presence of PDGFRα+/GFAP+ peritumoral astrocytes was significantly positively correlated to older patient age and peritumoral astrocyte density, but not to other established prognostic factors. Notably, presence of PDGFRα+/GFAP+ peritumoral astrocytes, but not peritumoral astrocyte density, was associated with significantly shorter patient overall survival. The prognostic association of PDGFRα+/GFAP+ peritumoral astrocytes was confirmed in multivariable analyses. This exploratory study thus demonstrates previously unrecognized intercase variability and prognostic significance of peritumoral abundance of a novel PDGFRα+ subset of GFAP+ astrocytes. Findings suggest clinically relevant roles of the microenvironment of peritumoral GBM tissue and encourage further characterization of the novel astrocyte subset with regard to origin, function, and potential as biomarker and drug target.

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Glioblastoma expression subtypes have been previously been associated with genomic abnormalities, treatment response, and differences in tumor microenvironment. We leveraged IDH wild-type glioblastomas, derivative neurospheres, and single cell gene expression profiles to define three tumorintrinsic transcriptional subtypes designated as proneural, mesenchymal, and classical, a revision of the previously reported TCGA subtypes. Transcriptomic subtype multiplicity correlated with increased intratumoral heterogeneity and the presence of tumor microenvironment. In silico cell sorting identified macrophages/microglia, CD4 + T lymphocytes, and neutrophils in the glioma microenvironment. NF1 deficiency resulted in increased tumorassociated macrophages/microglia infiltration. Comparison of matching primary and recurrent gliomas elucidated treatment-induced phenotypic tumor evolution, including expression subtype switching, in 45% of our cohort as well as associations between microenvironmental components and treatment response. Gene signature-based tumor microenvironment inference revealed a decrease in invading monocytes and a subtype-dependent increase in macrophages/microglia cells upon disease recurrence. Hypermutation at diagnosis or at recurrence was associated with CD8 + T cell enrichment. Frequency of M2 macrophage detection was associated with short-term relapse after radiation therapy. Our study provides a comprehensive transcriptional and cellular landscape of IDH wild-type glioblastoma during treatment modulated tumor evolution. Characterization of the evolving glioblastoma transcriptome and tumor microenvironment aids in designing more effective immunotherapy trials.