Inhibition of Notch signaling alters the phenotype of orthotopic tumors formed from glioblastoma multiforme neurosphere cells but does not hamper intracranial tumor growth regardless of endogene Notch pathway signature

Abstract

Background

Brain cancer stem-like cells (bCSC) are cancer cells with neural stem cell (NSC)-like properties found in the devastating brain tumor glioblastoma multiforme (GBM). bCSC are proposed a central role in tumor initiation, progression, treatment resistance and relapse and as such present a promising target in GBM research. The Notch signaling pathway is often deregulated in GBM and we have previously characterized GBM-derived bCSC cultures based on their expression of the Notch-1 receptor and found that it could be used as predictive marker for the effect of Notch inhibition. The aim of the present project was therefore to further elucidate the significance of Notch pathway activity for the tumorigenic properties of GBM-derived bCSC.

Methods

Human-derived GBM xenograft cells previously established as NSC-like neurosphere cultures were used. Notch inhibition was accomplished by exposing the cells to the gamma-secretase inhibitor DAPT prior to gene expression analysis and intracranial injection into immunocompromised mice.

Results

By analyzing the expression of several Notch pathway components, we found that the cultures indeed displayed different Notch pathway signatures. However, when DAPT-treated neurosphere cells were injected into the brain of immunocompromised mice, no increase in survival was obtained regardless of Notch pathway signature and Notch inhibition. We did however observe a decrease in the expression of the stem cell marker Nestin, an increase in the proliferative marker Ki-67 and an increased number of abnormal vessels in tumors formed from DAPT-treated, high Notch-1 expressing cultures, when compared with the control.

Conclusion

Based on the presented results we propose that Notch inhibition partly induces differentiation of bCSC, and selects for a cell type that more strongly induces angiogenesis if the treatment is not sustained. However, this more differentiated cell type might prove to be more sensitive to conventional therapies.

Keywords: :

• glioblastoma multiforme
• neurosphere culture
• orthotopic xenograft
• brain cancer stem-like cells
• Notch signaling
• Notch activity
• DAPT

Disclosure of Potential Conflict of Interest

The authors declare that they have no competing interests.

Acknowledgments

We thank Babloo Lukram for performing the immunohistochemistry and the Department of Neuropathology for technical assistance and advice. Financial support was kindly provided by: The Faculty of Health Sciences, University of Copenhagen (211-0610/09-3012); Dansk Kraeftforsknings Fond; Kong Christian den Tiendes Fond; Civilingenioer Frode V. Nyegaard og Hustrus Fond, Harboefonden, Danish National Advanced Technology Foundation, and The AP Møller Foundation.