Background: Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor in adults, with a median survival of 15 months for newly diagnosed GBM patients. Brain cancer stem-like cells (bCSC) are cancer cells with neural stem cell (NSC)-like properties found in GBM and they are proposed a central role in tumor initiation, progression, treatment resistance and relapse. The Notch receptor is expressed in the normal NSC population and is important for NSC maintenance and cell fate decision. The Notch signaling pathway is often deregulated in GBM and recent results suggest that this pathway plays a significant role in bCSC as well. The aim of the present project was therefore to further elucidate the significance of Notch expression and activation in GBM derived bCSC. Materials and methods: Human derived GBM xenograft cells were cultured in serum-free media in order to establish NSC-like neurosphere cultures. Notch modulation in the established GBM neurosphere cultures was accomplished either by blocking the Notch pathway using the γ-secretase inhibitor DAPT or by transfecting the cells with the intracellular constitutive active Notch-1 domain. Results: The established GBM neurosphere cultures expressed different levels of the Notch-1 receptor and the Notch downstream target Hes-1, which implies that the cultures have different Notch activation level. Cultures with high Notch activation showed sensitivity towards DAPT treatment. This was demonstrated by hampered cell viability, increase in the G0/G1 population, suggesting cell cycle arrest, and reduced ability to form colonies in soft-agar, together proposing that Notch inhibition targets the in vitro tumorigenic potential of these GBM neurosphere culture cells. When examining the effect of Notch inhibition on a variety of stem cell-like characteristics, only the primary sphere forming potential was affected. As such, the number of spheres formed in primary culture was reduced upon DAPT treatment while no effect was observed on the self-renewing capacity or the level of differentiation. In opposite, when the GBM neurosphere cells were transfected with the active part of the Notch-1 receptor a decrease in the G0/G1 population and an enhanced capability of soft-agar colony formation was observed as well as an increase in the self-renewing potential and de-differentiation. All cultures showed sensitivity towards artificial Notch-1 activation. These data implies that constitutive active Notch signaling increases the in vitro tumorigenic potential and affects the stem cell-like population in the GBM neurosphere cultures. Conclusion: Based on the observed results we propose that active Notch signaling is pivotal for the tumorigenic potential but not for the bCSC characteristics in GBM neurosphere cultures with elevated endogenous Notch activity.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3339. doi:1538-7445.AM2012-3339