Microenvironment-Derived Regulation of HIF Signaling Drives Transcriptional Heterogeneity in Glioblastoma Multiforme

Abstract

The evolving and highly heterogeneous nature of malignant brain tumors underlies their limited response to therapy and poor prognosis. In addition to genetic alterations, highly dynamic processes, such as transcriptional and metabolic reprogramming, play an important role in the development of tumor heterogeneity. The current study reports an adaptive mechanism in which the metabolic environment of malignant glioma drives transcriptional reprogramming. Multiregional analysis of a glioblastoma patient biopsy revealed a metabolic landscape marked by varying stages of hypoxia and creatine enrichment. Creatine treatment and metabolism was further shown to promote a synergistic effect through upregulation of the glycine cleavage system and chemical regulation of prolyl-hydroxylase domain. Consequently, creatine maintained a reduction of reactive oxygen species and change of the α-ketoglutarate/succinate ratio, leading to an inhibition of HIF signaling in primary tumor cell lines. These effects shifted the transcriptional pattern toward a proneural subtype and reduced the rate of cell migration and invasion Transcriptional subclasses of glioblastoma multiforme are heterogeneously distributed within the same tumor. This study uncovered a regulatory function of the tumor microenvironment by metabolism-driven transcriptional reprogramming in infiltrating glioma cells..

Tumor-associated reactive astrocytes aid the evolution of immunosuppressive environment in glioblastoma

“…Astrocytes (CRL-8621), Tumor cell lines (Glioma Stem-like Cells, GSC 1, 2, and 3, purified from surgical specimens) and microglia cells (T0251) were cultured in serum-free conditions as described previously 40 . For combining different cell types within a co-culture, we used fluorescence tagged cells and culture them under serum-free condition along a time period of 48 h. Cells were plated on laminin-coated dishes as described previously 40 . Validation of co-culture was done after 24 h and 48 h by fluorescence microscopy.…”

Astrogliosis Releases Pro-Oncogenic Chitinase 3-Like 1 Causing MAPK Signaling in Glioblastoma

“…In our preceding investigations, we demonstrated that the reactive transformation of astrocytes in our co-culture cell model closely mimics in-vivo circumstances, thereby allowing us to further map alterations in tumor cells caused by astrocytic crosstalk. By extracting gene signatures from all known transcriptional subtypes of glioblastoma (mesenchymal (Mes), proneural (PN), and classical (CL) tumors) as previous published [17], we were able to detect a subtype-switch of the BTSC#168 (classification in recent publication PN/CL [18,19]) cell line towards the mesenchymal phenotype. However, our results also showed that genes of the classical signature were partially up-regulated through astrocytic co-culture (Figure 2d).…”

Crosslink between Temozolomide and PD-L1 immune-checkpoint inhibition in glioblastoma multiforme

“…In a next step, we aimed to confirm our findings in cell culture models from one commercial (LN229) and two primary, patient-derived cell lines. To respect any possible effects of GBM subgroup properties, both primary cell lines were classified beforehand by RNA-sequencing analysis [22]. We noticed, that a baseline PD-L1 expression was only observable in the mesenchymal cell line.…”

“…For immunostaining, cells were cultivated on slides and fixed with 3% formaldehyde for 10 min at room temperature. Immunostaining was performed as recently published by our group [22]. A Fluoview FV10i confocal microscope from Olympus was used for fluorescence microscopy.…”