Abstract 1784: NOX-mediated chronic oxidative stress is inherent in human papillomavirus positive head and neck cancer cells and is implicated in radiosensitivity.
Authors: Marullo Rossella , Zhang Hongzheng , Chen Georgia Z. , Hunt William D. , Shin Dong M. , Doetsch Paul, W. 
Published: August 2014

Abstract
An increasing subgroup of oropharyngeal carcinomas is caused by infection with high risk types of human papillomavirus (HPV). Recent clinical data suggest that HPV status is an important prognostic factor associated with a favorable outcome in head and neck cancer patients treated with radiation therapy alone or in combination with chemotherapy. Although such data suggest an intrinsic sensitivity of HPV-positive tumors to radio/chemotherapy, no specific mechanism(s) has been shown to explain this feature. As DNA damage repair and/or tolerance capacity is one of the major determinants of sensitivity to DNA damaging agents, we interrogated a panel of five HPV positive (HPV+) and five HPV negative (HPV-) head and neck cancer cell lines for endogenous levels of DNA damage. HPV+ cells display an approximately 100% increase in the level of DNA damage compared to HPV- cells, as revealed by alkaline comet assays. As reactive oxygen species (ROS) represent the major endogenous source of DNA damage, we investigated whether HPV+ cells harbor high levels of ROS. We observed that HPV+ cells have approximately 100% higher levels of ROS compared to HPV- cells, measured by flow cytometry analysis. Treatment with the antioxidant N-acetyl-cysteine resulted in a concomitant reduction of ROS levels (∼35%) and endogenous DNA damage (∼41%) in HPV+ cells, indicating that chronic oxidative stress may contribute to genomic instability in HPV+ head and neck cancer cells. In addition, we found that treatment with the NOX inhibitor diphenylen iodonium reduces the levels of ROS in HPV+ cells (∼50%), indicating that NADPH oxidases are the source of the higher levels of ROS in these cells. Consistent with this mechanism, we observed that NOX-2, an inducible member of the NOXs family, is expressed only in HPV+ cells, suggesting that HPV infection may induce the transcription/translation of NOX-2 in head and neck cells. Collectively these data suggest that HPV infection may induce a state of chronic oxidative stress in head and neck cancer cells that may render HPV+ cells more susceptible to certain types of DNA damage. To test this hypothesis we evaluated the amount of DNA damage induced by exposure to a dose range of H2O2 or ionizing radiation (IR) in HPV+ and HPV- head and neck cancer cells. We found that each dose of H2O2 and IR induced a significantly greater amount of DNA damage in HPV+ cells compared to HPV- cells, indicating that HPV+ head and neck cells may be more prone to ROS-mediated DNA damage. This study provides new insights into HPV+ head and neck cancer cell biology and its sensitivity to IR.
Pilot funding for this project was provided by a grant from the V Foundation.
Citation Format: Rossella Marullo, Hongzheng Zhang, Georgia Z. Chen, William D. Hunt, Dong M. Shin, Paul, W. Doetsch. NOX-mediated chronic oxidative stress is inherent in human papillomavirus positive head and neck cancer cells and is implicated in radiosensitivity. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1784. doi:10.1158/1538-7445.AM2013-1784