Replication stress drives constitutive activation of the DNA damage response and consequent radioresistance in glioblastoma cancer stem cells

INTRODUCTION Glioblastoma (GBM)is a lethal primary brain tumour characterised by treatment resistance and inevitable recurrence, both of which are driven by a subpopulation of glioblastoma cancerstem cells (GSCs) with... [ view full abstract ]

INTRODUCTION Glioblastoma (GBM)is a lethal primary brain tumour characterised by treatment resistance and inevitable recurrence, both of which are driven by a subpopulation of glioblastoma cancerstem cells (GSCs) with tumorigenic and self-renewal properties. Radiation resistance of GSCs has been shown to be associated with enhanced DNA double strand break (DSB) repair proficiency caused by preferential activation of the DNA damage response (DDR), however the underlying reasons for activation of DDR in GSCs remains enigmatic. Uncertainty over the origin of GSCs also persists,with the original hypothesis that they derive from neural stem cells remaining unproven. METHODS AND RESULTS In this study we show that constitutive DDR activation in GSCs is driven by high levels of DNA replication stress (RS). We demonstrate increased RS in GSC compared to non-stem GBM cells in multiple primary cell cultures by DNA fibre assay and show that RS generates DSBs at DNA replication factories in GSCs. From a therapeutic perspective we demonstrate that radioresistance of GSC is reversed by dual inhibition of the key RS and DDR proteins ATR and PARP. Finally we identify increased expression of long neural genes as a likely mechanism for RS in GSC, an observation that correlates closely with recently published studies in neural progenitor cells which demonstrate that DSB arising from replication stress are preferentially located in long neural genes. Several of these genes are overexpressed in our GSC cultures. CONCLUSION Together these findings support the controversial hypothesis that GSC derive from neural stem cells. Overall, our observations elucidate the mechanism underlying DDR activation and radioresistance in GSC, shed new light on gliomagenesis and cancer stem cell biology, and identify novel therapeutics with potential to improve clinical outcomes by overcoming the inherent radioresistance of GBM.