Research Papers:

Dual effects of radiation bystander signaling in urothelial cancer: purinergic-activation of apoptosis attenuates survival of urothelial cancer and normal urothelial cells

Malgorzata A. Bill, Kirtiman Srivastava, Conor Breen, Karl T. Butterworth, Stephen J. McMahon, Kevin M. Prise and Karen D. McCloskey _

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Oncotarget. 2017; 8:97331-97343. https://doi.org/10.18632/oncotarget.21995

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Malgorzata A. Bill1, Kirtiman Srivastava1, Conor Breen1, Karl T. Butterworth1, Stephen J. McMahon1, Kevin M. Prise1 and Karen D. McCloskey1

1Centre for Cancer Research and Cell Biology, School of Medicine, Dentistry and Biomedical Sciences, Queen’s University Belfast, Belfast, BT9 7AE, Northern Ireland, UK

Correspondence to:

Karen D. McCloskey, email: [email protected]

Keywords: urothelial cancer; radiation; bystander response; apoptosis; purinergic signalling

Abbreviations: CM: conditioned media; T24: Grade 3 urothelial cancer cells; HT1376: Grade 3 urothelial cancer cells; HUC: primary urothelial cells; SV-HUC: immortalised urothelial cells

Received: March 01, 2017    Accepted: August 26, 2017    Published: October 24, 2017


Radiation therapy (RT) delivers tumour kill, directly and often via bystander mechanisms. Bladder toxicity is a dose limiting constraint in pelvic RT, manifested as radiation cystitis and urinary symptoms. We aimed to investigate the impact of radiation-induced bystander signaling on normal/cancer urothelial cells. Human urothelial cancer cells T24, HT1376 and normal urothelial cells HUC, SV-HUC were used. Cells were irradiated and studied directly, or conditioned medium from irradiated cells (CM) was transferred to naïve, cells. T24 or SV-HUC cells in the shielded half of irradiated flasks had increased numbers of DNA damage foci vs non-irradiated cells. A physical barrier blocked this response, indicating release of transmitters from irradiated cells. Clonogenic survival of shielded T24 or SV-HUC was also reduced; a physical barrier prevented this phenomenon. CM-transfer increased pro-apoptotic caspase-3 activity, increased cleaved caspase-3 and cleaved PARP expression and reduced survival protein XIAP expression. This effect was mimicked by ATP. ATP or CM evoked suramin-sensitive Ca2+-signals. Irradiation increased [ATP] in CM from T24. The CM-inhibitory effect on T24 clonogenic survival was blocked by apyrase, or mimicked by ATP. We conclude that radiation-induced bystander signaling enhances urothelial cancer cell killing via activation of purinergic pro-apoptotic pathways. This benefit is accompanied by normal urothelial damage indicating RT bladder toxicity is also bystander-mediated.

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