PO-127 Investigating the response of normal and cancer bladder cells to radiotherapy

Introduction Bladder cancer is the fourth most commonly diagnosed cancer among males worldwide. Recent advances in radiation delivery have increased the importance of bladder preserving strategies using radiotherapy, while also improving patient’s quality of life. However, the outcome of radiotherapy depends on the radiosensitivity of both the patient and his/her tumour. Recent developments in radiobiology have highlighted the importance of the assessment of radiation-induced DNA double-strand breaks (DSB) repair kinetics in predicting radiosensitivity of both normal and cancerous tissues. Indeed, after irradiation, several proteins (ATM, γH2AX, etc.) involved in the signalling and repair of DNA DSB relocalize as nuclear foci. The purpose of this study is to characterise the radiosensitivity of bladder cell lines, based on their capacity to repair radio-induced DNA damage. Material and methods Four human bladder cell lines were used in this study: T24, UM-UC3, RT4 and SVHUC. Clonogenic assay was performed to study cell survival after irradiation: cells were irradiated with doses ranging from 0 to 10 Gy and their capacity to form colonies was assessed. Immunofluorescent analysis using anti-pATM and anti-γH2AX antibodies was then performed to assess DNA DSB signalling and repair kinetics after a 2Gy irradiation. Finally, we performed the three-dimensional (3D) sphere formation assay to assess the effect of irradiation on cancer/stem progenitor cells. Irradiated cells with doses ranging from 0 to 10 Gy were embedded in Matrigel and their capacity to form spheres was assessed. Results and discussions RT4 was found to have the highest survival rate after irradiation, followed by T24, UM-UC3 and SVHUC according to the clonogenic assay results. Immunofluorescence results were consistent with cell survival as RT4 showed the fastest DSB recognition and repair kinetics while SVHUC had a much slower rate. Furthermore, the capacity of cells to recognise radio-induced DNA damage was found to have the best correlation with their survival. Lastly, the sphere formation ability of the different cell lines showed a differential response to increasing irradiation doses. Conclusion We were able to radiobiologically characterise 4 human bladder cell lines by assessing their survival and capacity to repair radio-induced DNA damage. Moreover, we showed the differential effects of radiation on cancer stem/progenitor cells. The results highlighted the importance of DNA DSB signalling through the ATM protein and its role in cell survival after irradiation.