Department of Neurosurgery

Human organotypic brain slice culture: a novel framework for environmental research in neuro-oncology

When it comes to the human brain, models that closely mimic in vivo conditions are lacking. Living neuronal tissue is the closest representation of the in vivo human brain outside of a living person. Here, we present a method that can be used to maintain therapeutically resected healthy neuronal tissue for prolonged periods without any discernible changes in tissue vitality, evidenced by immunohistochemistry, genetic expression, and electrophysiology. This method was then used to assess glioblastoma (GBM) progression in its natural environment by microinjection of patient-derived tumor cells into cultured sections. The result closely resembles the pattern of de novo tumor growth and invasion, drug therapy response, and cytokine environment. Reactive transformation of astrocytes, as an example of the cellular nonmalignant tumor environment, can be accurately simulated with transcriptional differences similar to those of astrocytes isolated from acute GBM specimens. In a nutshell, we present a simple method to study GBM in its physiological environment, from which valuable insights can be gained. This technique can lead to further advancements in neuroscience, neuro-oncology, and pharmacotherapy.

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

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

Reactive astrocytes evolve after brain injury, inflammatory and degenerative diseases, whereby they undergo transcriptomic re-programming. In malignant brain tumors, their function and crosstalk to other components of the environment is poorly understood. Here we report a distinct transcriptional phenotype of reactive astrocytes from glioblastoma linked to JAK/STAT pathway activation. Subsequently, we investigate the origin of astrocytic transformation by a microglia loss-of-function model in a human organotypic slice model with injected tumor cells. RNA-seq based gene expression analysis of astrocytes reveals a distinct astrocytic phenotype caused by the coexistence of microglia and astrocytes in the tumor environment, which leads to a large release of anti-inflammatory cytokines such as TGFß, IL10 and G-CSF. Inhibition of the JAK/STAT pathway shifts the balance of pro- and anti- inflammatory cytokines towards a pro-inflammatory environment. The complex interaction of astrocytes and microglia cells promotes an immunosuppressive environment, suggesting that tumor-associated astrocytes contribute to anti-inflammatory responses.

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

In recent years, PD-1/PD-L1 immune checkpoint inhibitors have improved cancer therapy in many tumor types, but no benefit of immune checkpoint therapy has been found in glioblastoma multiforme (GBM). Based on the results of our earlier work, which showed a reduction of PD-L1 expression in patients treated with temozolomide (TMZ), we aimed to investigate the link between TMZ therapy and the immune control point target PD-L1. RNA-sequencing data from de-novo and recurrent glioblastoma were analyzed by AutoPipe algorithm. Results were confirmed either in a cell model by two primary and one established GBM cell line and specimens of de-novo and recurrent GBM. PD-L1 and pathway activation of the JAK/STAT pathway was analyzed by quantitative real-time PCR and western blot. We found a significant downregulation of the JAK/STAT pathway and immune response in recurrent tumors. The cell model showed an upregulation of PD-L1 after IFN? treatment, while additional TMZ treatment lead to a reduction of PD-L1 expression and JAK/STAT pathway activation. These findings were confirmed in specimens of de-novo and recurrent glioblastoma.Our results suggest that TMZ therapy leads to a down-regulation of PD-L1 in primary GBM cells. These results support the clinical findings where PD-L1 is significantly reduced in recurrent GBMs. If the target is diminished, it may also lead to impaired efficacy of PD-1/PD-L1 inhibitors such as nivolumab.

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

The integrative metabolomic-transcriptomic landscape of glioblastome multiforme