Abstract 5504: A potential source in oxidative cell death mediated by thymidylate synthase inhibitors: NADPH oxidase 2

Abstract

Abstract Thymidylate synthase (TS) catalyzes the reductive transfer of a methyl group from N5,N10-methylenetetrahydrofolate (CH2-THF) to dUMP, forming dTMP and dihydrofolate (DHF). It is generally accepted that inhibition of the enzyme leads to a deficiency of dTMP, followed by genome damage and programmed cell death. As such, TS has long been viewed as an important target of anti-neoplastic agents, such as 5-fluorouracil (5-FU) and raltitrexed (RTX), which lead to inhibition of the enzyme. Oxidative stress is well-recognized as having a central role in cellular response to a number of DNA damaging agents, and may be a proximate cause of therapy-induced cell death. Despite such recognition, little detail exists on the origin and regulation of reactive oxygen species (ROS) during TS-directed chemotherapy. We have undertaken a detailed examination of the origin, nature, and role of ROS in cell death mediated by TS inhibitors. We have previously identified NADPH oxidase (NOX) as the primary source of increased ROS following exposure to TS-directed agents, as indicated by the observation of decreased ROS production and apoptotic indices in presence of NOX inhibitors. We have found that in response to TS inhibitors, NOX enzyme activity increases in association with induction of the transcript for p67phox, which specifically regulates the NOX2 isoform. This result is also verified by silencing p67phox gene in HCT116 cells. Silencing p67phox gene inhibits drug-induced NOX activity and decreases drug-induced apoptotic indices. To substantiate and expand results obtained from HCT116, we used other colorectal cancer cells, including HCT15, SW480, DLD-1, LoVo, MOSER and LS180. We have determined that colon cancer cells exhibit diverse responses in both basal and drug-inducible levels of apoptosis, NOX activity and mRNA of NOX2 regulatory subunits, implicating the heterogeneity in colorectal cancer cells. Based on these results, we conclude that augmentation of NOX2 activity via induction of p67phox mRNA expression is the proximate cause of programmed cell death elicited by dTMP deficiency in HCT116 cells. Each colorectal cancer cells might have different mechanism to undergo ROS-mediated cell death. Citation Format: Ufuk Ozer, Franklin George Berger, Karen Wood Barbour. A potential source in oxidative cell death mediated by thymidylate synthase inhibitors: NADPH oxidase 2. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5504. doi:10.1158/1538-7445.AM2014-5504