Abstract 1592: Mitochondrial dysfunction and radioresistance in esophageal cancer.
Authors: Lynam-Lennon Niamh , Maher Stephen G. , Reynolds John V. , O'Sullivan Jacintha 
Published: August 2014

Introduction: Radiation therapy is fundamental to the treatment of esophageal cancer. However, radioresistance is a significant clinical problem. The elucidation of molecular mechanisms underlying radioresistance would be of substantial clinical benefit. Mitochondria are a critical cellular target of reactive oxygen species (ROS), the generation of which accounts for ∼65% of radiation-induced damage. We hypothesize that mitochondrial dysfunction may play a significant role in driving radioresistance in esophageal cancer.

Materials and Methods: An isogenic model of radioresistance in esophageal adenocarcinoma (EAC) was established by chronically irradiating OE33 cells with fractionated, clinically-relevant doses of 2 Gy X-ray radiation (cumulative dose, 50 Gy). Radiosensitivity was assessed by clonogenic assay. ROS levels were assessed by fluorimetry and high content screening. Mitochondrial mutation frequency was determined using the qPCR-based random mutation capture assay. Oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) were measured using the Seahorse XF Extracellular Flux Analyzer (Seahorse Bioscience. Expression of 84 genes involved in mitochondrial energy metabolism was assessed using qPCR-based arrays. Mitochondrial number was assessed by transmission electron microscopy (TEM).

Results: Chronic exposure of OE33 cells to fractionated doses of radiation resulted in a radioresistant subline, OE33 R. Characterization of this model revealed that, relative to the age- and passage-matched parent control (OE33 P), radioresistant cells had altered ROS levels following radiation and increased basal frequencies of random mitochondrial mutations (p = 0.01). Basal mitochondrial metabolism was also altered in these cells, with OE33 R demonstrating increased basal OCR (p &lt; 0.05) and increased expression of 7 genes involved in regulating mitochondrial energy metabolism. TEM analysis of these radioresistant cells demonstrated increased basal numbers of mitochondria, which were darker and denser in appearance, when compared to OE33 P cells. Radiation did not alter the mitochondrial number in OE33 R cells, which was in contrast to radiosensitive OE33 P cells.

Conclusion: Whilst a role for mitochondrial dysfunction is strongly implicated in carcinogenesis, its potential role in modulating the tumor response to anti-cancer therapy is unknown. We have generated a novel radioresistant subline that differs from its parent only in terms of radiosensitivity. We have demonstrated alterations in mitochondrial function, mitochondrial mutagenesis, mitochondrial metabolism, mitochondrial number and gene expression in these radioresistant cells. This strongly supports a role for mitochondrial dysfunction in modulating radioresistance in EAC and suggests the mitochondria as a potential therapeutic target for improving the tumor response to radiation therapy.

Citation Format: Niamh Lynam-Lennon, Stephen G. Maher, John V. Reynolds, Jacintha O'Sullivan. Mitochondrial dysfunction and radioresistance in esophageal cancer. [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 1592. doi:10.1158/1538-7445.AM2013-1592