A functional study of EGFR and Notch signaling in brain cancer stem-like cells from glioblastoma multiforme (Ph.d.)

Karina Kristoffersen

    Abstract

    Glioblastoma Multiforme (GBM) is the most common and aggressive brain tumor in adults with a median survival for newly diagnosed GBM patients at less than 1.5 year. Despite intense treatment efforts the vast majority of patients will experience relapse and much research today is therefore searching for new molecular and cellular targets that can improve the prognosis for GBM patients. One such target is the brain cancer stem-like cells (bCSC) that are believed to be responsible for tumor initiation, progression, treatment resistance and ultimately relapse. bCSC are identified based on their resemblance to normal neural stem cells (NSC) and their tumorigenic potential. Like for NSC, the epidermal growth factor receptor (EGFR) and Notch receptor signaling pathways are believed to be important for the maintenance of bCSC. These pathways as such present promising targets in a future anti-bCSC GBM treatment.
    The overall aim of the present PhD project has been to study the functional role of EGFR and Notch activity in bCSCs stem cell-like features and tumorigenic potential with the purpose of deepen our knowledge about the significance of these pathways in the bCSC population in GBM.
    By establishing and culturing human derived GBM xenograft cells under NSC conditions we obtained neurosphere cultures that contained cells with stem cell-like and tumorigenic properties. We moreover characterized the different cultures based on their expression level of the EGFR and Notch receptor as well as the expression of the mutant receptor EGFRvIII, an expression that was maintained from patient material to the xenograft tumors and cell cultures. In a culture expressing EGFR and EGFRvIII we found that EGFR inhibition induced differentiation, while forced differentiation led to down-regulation of EGFR and EGFRvIII. In addition, we showed that EGFR/EGFRvIII down regulation either as a result of induced differentiation or EGFR inhibition led to decreased in vitro tumorigenic and stem cell-like potential. In cultures expressing high levels of the Notch-1 receptor we found that Notch inhibition decreased the in vitro tumorigenic potential while, of the stem cell features, only the primary sphere forming potential was inhibited. Cultures with low Notch expression were not affected by Notch inhibition. In opposite, we found that artificial Notch activation resulted in increased in vitro tumorigenic potential along with indications of increased stem cell-like potential in all cultures. Taken together, these in vitro results suggest that EGFR and Notch activity are important for maintaining the stem cell-like and tumorigenic potential of the bCSC population. When we tested the effect of Notch inhibition on intracranial tumor growth, we did not observe increased survival for mice injected with Notch inhibited cells regardless of the cells initial Notch activity. However, we found that tumors grown from high Notch expressing cultures treated with a Notch inhibitor displayed augmented angiogenic potential and a tendency to increased differentiation. Finally, we found that the neurosphere cultures used in this project could be further characterized based on their global gene expression profile and that this profile, to some degree could be correlated with response to Notch inhibitory treatment.
    Based on the results obtained throughout this thesis project, we suggest that targeting a bCSC population by EGFR and/or Notch inhibition is feasible and future studies might prove if anti-bCSC therapy in combination with conventional therapy can improve the prognosis for GBM patients displaying a specific gene expression profile
    OriginalsprogEngelsk
    Antal sider170
    StatusUdgivet - 7 okt. 2013

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