Tenascin-C is a mesenchymal glioma marker that regulates tumor plasticity

Introduction

Glioblastoma (GBM) is one of the deadliest human brain cancers, with an average survival of around one year from the time of diagnosis. Next generation sequencing has allowed the classification of GBM into subtypes based on gene expression signatures. Glioblastomas classified into the mesenchymal (MES) subtype exhibit worse prognosis and have been correlated with poor radiation responses, high expression of CD44, and NF-kB activation. We have developed a mouse model of GBM that resembles the MES subtype, which is reminiscent of a transition that has been linked to dedifferentiated and transdifferentiated tumors. In this study, we investigated the mechanism of dedifferentiation/reprogramming achieved by astrocytes and neurons upon transduction with oncogenic lentiviruses.

Materials and Methods

We performed transcriptome analysis of the dedifferentiated neurons and astrocytes along with parental cell controls and found among a list of differentially expressed genes, Tenascin C (TNC) to be at the top of the list. In order to assess the functional role of TNC in tumor reprogramming, stem cell maintenance and tumor growth, we knockdown TNC using lentiviral vectors expressing shRNAs. In vitro, we performed proliferation, cell cycle and apoptosis analysis, invasion/migration analysis (scratch assay), neurosphere formation and dedifferentiation and transdifferentiation assays. In vivo, we transplanted glioma cells with or without expression of TNC and followed tumor growth.

Results and Discussion

TNC is overexpressed in both murine and human MES glioblastoma, and negatively correlates with proneural subtype. Silencing TNC in murine glioma cells lead to reduced expression of MES markers, significantly decreased their proliferative capacity and induced apoptosis as well as decreased the S and G₂/M phase in the cell cycle. Loss of TNC in transformed astrocytes impaired their ability to dedifferentiate, evidenced by decreased ability to form characteristic neurospheres. Silencing TNC in 005 glioma stem cells (GSC) attenuated the ability to form spheres (smaller size) and reduced the proportion of self-renewing sphere-forming units. Interestingly, 005-shTNC GSC could not differentiate into functional tumor derived endothelial cells assessed by impaired tube formation capacity. Specific inhibition of TNC in both murine and human glioma tumors prolonged mice survival.

Conclusions

We conclude that TNC determines mesenchymal identity and is an important player in dedifferentiation of glioma cells, hence its inhibition can be a therapeutic target for this deadly type of cancer.