Abstract A41: Leveraging metabolic dependencies in cancer: Cysteine addiction in pancreatic cancer cells

Field: Biochemistry, Genetics and Molecular Biology (Life Sciences)

Abstract Pancreatic cancer is a devastating and lethal malignancy, estimated to kill over 40,000 Americans per year. Little has changed in the treatment of this disease over the past half-century, highlighting the need for an improved understanding of the molecular mechanisms driving tumor development and maintenance. The malignant behavior of tumor cells is underwritten by alterations to their cellular metabolism, which is optimized to meet the catabolic needs of a rapidly dividing cell. One of the consequences of this oncogenic growth is increased generation of reactive oxygen species (ROS), a potentially toxic byproduct of many metabolic reactions. Interestingly, oncogenes such as Kras, the dominant driver of pancreatic cancer, can counteract ROS toxicity through activation of an antioxidant program, creating a highly reducing intracellular environment. Cysteine is the rate-limiting precursor for the synthesis of glutathione (GSH), the preeminent antioxidant of the cell, and as such may represent a key metabolite in the maintenance of redox balance in tumor cells. In this study we explore the importance of cysteine metabolism in the viability of pancreatic cancer cells and test the effects of cysteine deprivation and the pharmacological inhibition of cysteine uptake on a variety of pancreatic cancer cell lines. To explore the role of cysteine in pancreatic cancer growth, we cultured a panel of human and murine pancreatic cancer cell lines in the absence of cysteine. We found that cysteine withdrawal rapidly induced a peculiar form of cell death called ferroptosis, which can result from the catastrophic accumulation of certain lipid peroxides. Cystine, the oxidized form of cysteine, is the predominant extracellular source of cysteine in most cells, and it is normally imported into the cell by a specific amino acid transporter known as system xc-. We show that pharmacological inhibition of sytem xc- causes rapid cell death in most pancreatic cancer cells through ferroptosis. We find, however, that some cells do exhibit baseline resistance to cysteine deprivation or system xc- inhibition. We hypothesized that these cells utilize methionine to synthesize cysteine through transsulfuration. We show that these cells can be sensitized to system xc- inhibition when co-treated with propargyglycine (PPG), an irreversible inhibitor of gamma-cystathionase, an enzyme required for transsulfuration. These findings imply that cysteine is an important substrate for appropriate redox homeostasis in these cells. To further explore if GSH synthesis is the key mediator of cysteine-deprivation induced cell death, we treated pancreatic tumor cells with buthionine sulfoxamine (BSO), a potent and specific inhibitor of GSH synthesis. To our surprise, cells treated with BSO do not exhibit any of the ill-effects we see in cysteine-deprived pancreatic cancer cells. Therefore, it is unlikely that the cell death seen in cells deprived of cysteine is due solely to GSH depletion, but possibly due to the activity of other antioxidant pathways that require cysteine. In conclusion, we find that pancreatic cancer cells are highly sensitive to cysteine-deprivation and to treatment with system xc- inhibitors, and that the cell death caused by this stress is an iron-dependent, oxidative form of cell death called ferroptosis. Furthermore, it appears that GSH, the primary antioxidant molecule of the cell, is not solely responsible for mediating this cell death, arguing that cysteine may play other important roles in maintaining redox homeostasis in these cells. Given that previous studies show that global system xc- knockout mice do not exhibit any changes in cellular GSH content, these findings may represent a targetable metabolic dependency in pancreatic cancer cells. Citation Format: Michael A. Badgley, Carmine F. Palermo, Stephen A. Sastra, Brent R. Stockwell, Kenneth P. Olive. Leveraging metabolic dependencies in cancer: Cysteine addiction in pancreatic cancer cells. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr A41.