Level of Notch activation determines the effect on growth and stem cell-like features in glioblastoma multiforme neurosphere cultures

Background: Brain cancer stem-like cells (bCSC) are cancer cells with neural stem cell (NSC)-like properties found in glioblastoma multiforme (GBM) and they are assigned a central role in tumor initiation, progression and relapse. The Notch pathway is important for maintenance and cell fate decisions in the normal NSC population. Notch signaling is often deregulated in GBM and recent results suggest that this pathway plays a significant role in bCSC as well. We therefore wished to further elucidate the role of Notch activation in GBM-derived bCSC.   Methods: Human-derived GBM xenograft cells were cultured as NSC-like neurosphere cultures. Notch modulation was accomplished either by blocking the pathway using the γ-secretase inhibitor DAPT or by activating it by transfecting the cells with the constitutive active Notch-1 receptor. Results: GBM neurosphere cultures with high endogenous Notch activation displayed sensitivity toward Notch inhibition with regard to tumorigenic features as demonstrated by increased G(0)/G(1) population and reduced colony formation capacity. Of the NSC-like characteristics, only the primary sphere forming potential was affected, while no effect was observed on self-renewal or differentiation. In contrast, when Notch signaling was activated a decrease in the G(0)/G(1) population and an enhanced capability of colony formation was observed, along with increased self-renewal and de-differentiation. Conclusion: Based on the presented results we propose that active Notch signaling plays a role for cell growth and stem cell-like features in GBM neurosphere cultures and that Notch-targeted anti-bCSC treatment could be feasible for GBM patients with high endogenous Notch pathway activation

Differentiation of glioblastoma multiforme stem-like cells leads to downregulation of EGFR and EGFRvIII and decreased tumorigenic and stem-like cell potential

Glioblastoma multiforme (GBM) is the most common and devastating primary brain tumor among adults. Despite recent treatment progress, most patients succumb to their disease within 2 years of diagnosis. Current research has highlighted the importance of a subpopulation of cells, assigned brain cancer stem-like cells (bCSC), to play a pivotal role in GBM malignancy. bCSC are identified by their resemblance to normal neural stem cells (NSC), and it is speculated that the bCSC have to be targeted in order to improve treatment outcome for GBM patients. One hallmark of GBM is aberrant expression and activation of the epidermal growth factor receptor (EGFR) and expression of a deletion variant EGFRvIII. In the normal brain, EGFR is expressed in neurogenic areas where also NSC are located and it has been shown that EGFR is involved in regulation of NSC proliferation, migration, and differentiation. This led us to speculate if EGFR and EGFRvIII are involved in the regulation of bCSC. In this study we use GBM neurosphere cultures, known to preserve bCSC features. We demonstrate that EGFR and EGFRvIII are downregulated upon differentiation and moreover that when EGFR signaling is abrogated, differentiation is induced. Furthermore, we show that differentiation leads to decreased tumorigenic and stem cell-like potential of the neurosphere cultures and that by specifically inhibiting EGFR signaling it is possible to target the bCSC population. Our results suggest that differentiation therapy, possibly along with anti-EGFR treatment would be a feasible treatment option for patients with GBM, by targeting the bCSC population

Clinical variables serve as prognostic factors in a model for survival from glioblastoma multiforme:an observational study of a cohort of consecutive non-selected patients from a single institution

BACKGROUND: Although implementation of temozolomide (TMZ) as a part of primary therapy for glioblastoma multiforme (GBM) has resulted in improved patient survival, the disease is still incurable. Previous studies have correlated various parameters to survival, although no single parameter has yet been identified. More studies and new approaches to identify the best and worst performing patients are therefore in great demand. METHODS: This study examined 225 consecutive, non-selected GBM patients with performance status (PS) 0–2 receiving postoperative radiotherapy with concomitant and adjuvant TMZ as primary therapy. At relapse, patients with PS 0–2 were mostly treated by reoperation and/or combination with bevacizumab/irinotecan (BEV/IRI), while a few received TMZ therapy if the recurrence-free period was >6 months. RESULTS: Median overall survival and time to progression were 14.3 and 8.0 months, respectively. Second-line therapy indicated that reoperation and/or BEV/IRI increased patient survival compared with untreated patients and that BEV/IRI was more effective than reoperation alone. Patient age, ECOG PS, and use of corticosteroid therapy were significantly correlated with patient survival and disease progression on univariate analysis, whereas p53, epidermal growth factor receptor, and O(6)-methylguanine-DNA methyltransferase expression (all detected by immunohistochemistry), tumor size or multifocality, and extent of primary operation were not. A model based on age, ECOG PS, and corticosteroids use was able to predict survival probability for an individual patient. CONCLUSION: The survival of RT/TMZ-treated GBM patients can be predicted based on patient age, ECOG PS, and corticosteroid therapy status

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

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