Cell-lineage controlled epigenetic regulation in glioblastoma stem cells determines functionally distinct subgroups and predicts patient survival.

There is ample support for developmental regulation of glioblastoma stem cells. To examine how cell lineage controls glioblastoma stem cell function, we present a cross-species epigenome analysis of mouse and human glioblastoma stem cells. We analyze and compare the chromatin-accessibility landscape of nine mouse glioblastoma stem cell cultures of three defined origins and 60 patient-derived glioblastoma stem cell cultures by assay for transposase-accessible chromatin using sequencing. This separates the mouse cultures according to cell of origin and identifies three human glioblastoma stem cell clusters that show overlapping characteristics with each of the mouse groups, and a distribution along an axis of proneural to mesenchymal phenotypes. The epigenetic-based human glioblastoma stem cell clusters display distinct functional properties and can separate patient survival. Cross-species analyses reveals conserved epigenetic regulation of mouse and human glioblastoma stem cells. We conclude that epigenetic control of glioblastoma stem cells primarily is dictated by developmental origin which impacts clinically relevant glioblastoma stem cell properties and patient survival.

A molecularly distinct subset of glioblastoma requires serum-containing media to establish sustainable bona fide glioblastoma stem cell cultures.

Glioblastoma (GBM) is the most frequent and deadly primary malignant brain tumor. Hallmarks are extensive intra-tumor and inter-tumor heterogeneity and highly invasive growth, which provide great challenges for treatment. Efficient therapy is lacking and the majority of patients survive less than 1 year from diagnosis. GBM progression and recurrence is caused by treatment-resistant glioblastoma stem cells (GSCs). GSC cultures are considered important models in target identification and drug screening studies. The current state-of-the-art method, to isolate and maintain GSC cultures that faithfully mimic the primary tumor, is to use serum-free (SF) media conditions developed for neural stem cells (NSCs). Here we have investigated the outcome of explanting 218 consecutively collected GBM patient samples under both SF and standard, serum-containing media conditions. The frequency of maintainable SF cultures (SFCs) was most successful, but for a subgroup of GBM specimens, a viable culture could only be established in serum-containing media, called exclusive serum culture (ESC). ESCs expressed nestin and SOX2, and displayed all functional characteristics of a GSC, that is, extended proliferation, sustained self-renewal and orthotopic tumor initiation. Once adapted to the in vitro milieu they were also sustainable in SF media. Molecular analyses showed that ESCs formed a discrete group that was most related to the mesenchymal GBM subtype. This distinct subgroup of GBM that would have evaded modeling in SF conditions only provide unique cell models of GBM inter-tumor heterogeneity.

Human Mesenchymal glioblastomas are characterized by an increased immune cell presence compared to Proneural and Classical tumors.

Glioblastoma (GBM) is the most aggressive malignant primary brain tumor in adults, with a median survival of 14.6 months. Recent efforts have focused on identifying clinically relevant subgroups to improve our understanding of pathogenetic mechanisms and patient stratification. Concurrently, the role of immune cells in the tumor microenvironment has received increasing attention, especially T cells and tumor-associated macrophages (TAM). The latter are a mixed population of activated brain-resident microglia and infiltrating monocytes/monocyte-derived macrophages, both of which express ionized calcium-binding adapter molecule 1 (IBA1). This study investigated differences in immune cell subpopulations among distinct transcriptional subtypes of GBM. Human GBM samples were molecularly characterized and assigned to Proneural, Mesenchymal or Classical subtypes as defined by NanoString nCounter Technology. Subsequently, we performed and analyzed automated immunohistochemical stainings for TAM as well as specific T cell populations. The Mesenchymal subtype of GBM showed the highest presence of TAM, CD8+, CD3+ and FOXP3+ T cells, as compared to Proneural and Classical subtypes. High expression levels of the TAM-related gene AIF1, which encodes the TAM-specific protein IBA1, correlated with a worse prognosis in Proneural GBM, but conferred a survival benefit in Mesenchymal tumors. We used our data to construct a mathematical model that could reliably identify Mesenchymal GBM with high sensitivity using a combination of the aforementioned cell-specific IHC markers. In conclusion, we demonstrated that molecularly distinct GBM subtypes are characterized by profound differences in the composition of their immune microenvironment, which could potentially help to identify tumors amenable to immunotherapy.

Mouse Models of Pediatric Supratentorial High-grade Glioma Reveal How Cell-of-Origin Influences Tumor Development and Phenotype.

High-grade glioma (HGG) is a group of primary malignant brain tumors with dismal prognosis. Whereas adult HGG has been studied extensively, childhood HGG, a relatively rare disease, is less well-characterized. Here, we present two novel platelet-derived growth factor (PDGF)-driven mouse models of pediatric supratentorial HGG. Tumors developed from two different cells of origin reminiscent of neural stem cells (NSC) or oligodendrocyte precursor cells (OPC). Cross-species transcriptomics showed that both models are closely related to human pediatric HGG as compared with adult HGG. Furthermore, an NSC-like cell-of-origin enhanced tumor incidence, malignancy, and the ability of mouse glioma cells (GC) to be cultured under stem cell conditions as compared with an OPC-like cell. Functional analyses of cultured GC from these tumors showed that cells of NSC-like origin were more tumorigenic, had a higher rate of self-renewal and proliferation, and were more sensitive to a panel of cancer drugs compared with GC of a more differentiated origin. These two mouse models relevant to human pediatric supratentorial HGG propose an important role of the cell-of-origin for clinicopathologic features of this disease. Cancer Res; 77(3); 802-12. ©2016 AACR.

Glioblastoma Cell Malignancy and Drug Sensitivity Are Affected by the Cell of Origin.

The identity of the glioblastoma (GBM) cell of origin and its contributions to disease progression and treatment response remain largely unknown. We have analyzed how the phenotypic state of the initially transformed cell affects mouse GBM development and essential GBM cell (GC) properties. We find that GBM induced in neural stem-cell-like glial fibrillary acidic protein (GFAP)-expressing cells in the subventricular zone of adult mice shows accelerated tumor development and produces more malignant GCs (mGC1GFAP) that are less resistant to cancer drugs, compared with those originating from more differentiated nestin- (mGC2NES) or 2,'3'-cyclic nucleotide 3'-phosphodiesterase (mGC3CNP)-expressing cells. Transcriptome analysis of mouse GCs identified a 196 mouse cell origin (MCO) gene signature that was used to partition 61 patient-derived GC lines. Human GC lines that clustered with the mGC1GFAP cells were also significantly more self-renewing, tumorigenic, and sensitive to cancer drugs compared with those that clustered with mouse GCs of more differentiated origin.