We thank Oncothyreon Inc. to the lung and brain and with decreased overall patient survival. Our studies provided insight into redox-based mechanisms underlying tumor growth and metastasis and suggest that TXNL2 could be a target for treatment of breast cancer. == Introduction == ROS, including superoxide O2, hydroxyl radical OH, and H2O2, are constantly generated during intracellular metabolism and in response to environmental stimuli (1). Generally, ROS are regarded as Klf1 host-defending molecules that destroy exogenous pathogens (2) and act as secondary messengers in signal JNJ-64619178 transduction (1,3). However, increased production of ROS is involved in committing cells to apoptosis (3,4). Although ROS are involved in tumorigenesis and progression, as reflected by ROS activation of tumor-promoting signaling pathways (5), excess oxidative stress, due to further elevated ROS levels beyond a threshold or weakened antioxidative defense, can damage macromolecules vital for cellular functions (6,7). This in turn results in pathophysiological changes, such as apoptosis, cell cycle disruption, and necrosis (8). As such, induction of ROS-mediated damage in cancer cells by proper pharmacological agents that either promote ROS generation or disable the cellular antioxidant system has been considered as a radical therapeutic strategy to preferentially kill cancer cells (9). The redox state in the normal cell is balanced by the cellular antioxidant capacity to maintain a viable steady-state environment that is predominantly reducing (10). A key mechanism by which cells regulate redox processes is the reversible formation of disulfides through the oxidation of thiol groups in cysteine residues (11). To maintain the cellular thiol-disulfide redox balance, living cells possess 2 major regulatory systems: the thioredoxin/thioredoxin (Trx/Trx) reductase system and the glutaredoxin/glutathione/glutathione (Grx/GSH/GSH) reductase system (12). Trx-1 (12 kDa) is a well-documented member of the Trx regulatory system that reduces disulfide bonds and thus regulates the activity of transcriptional factors like AP-1, NF-B, and p53 (13,14). Overexpression of Trx-1 inhibits apoptosis (15). Grxs can readily reduce S-glutathionylated protein (protein-SSG) mixed disulfide and can be regenerated JNJ-64619178 JNJ-64619178 by the reduced form of GSH (16). Grxs protect cells against oxidative stress by catalyzing protein de-glutathionylation and has therefore been implicated in various cellular processes, including regulation of transcription factor binding activities and redox regulation (1719). For example, Grx-1 regulates intracellular and extracellular homeostasis of protein glutathionylation (2022). Elevated oxidative status has been observed in many types of cancer cells, due in part to their high metabolic rate. On the other hand, many tumor cells possess stronger antioxidative defense mechanisms to counterbalance excessive ROS, maintain their redox status, and thus suppress apoptosis (23). This phenomenon may be a consequence of JNJ-64619178 cellular adaption to ROS stress and may play an important role in the development of highly malignant behaviors and drug resistance (9). Overexpression of Trx-1 in MCF-7 human breast cancer cells enhances JNJ-64619178 cell growth (24). Increased Trx-1 protein levels are found in several human cancers (25,26). Interestingly, Grx-1 expression can be induced by oxidative stress in breast cancer cells and thus inhibits apoptosis (27). During a recent microarray analysis of the IGF-regulated genes in breast cancer cells, we found that a novel Trx-related protein, Trx-like 2 (TXNL2; also known as Grx3 and PICOT), is significantly induced by IGFs (28). The 38-kDa TXNL2 protein is much larger than typical Trx proteins and has a unique protein structure consisting of an N-terminal Trx homology region, followed by 2 tandem repeats of Grx domains (22,29,30). Grx3/4, the yeast homolog of TXNL2, was implicated in the regulation of the oxidative stress response (31). Although TXNL2 is conserved in eukaryotes, the physiological function in mammalian cells is still poorly understood (30). Recent reports showed that it can inhibit cardiac hypertrophy through enhancing ventricular function and cardiomyocyte contractility and can regulate FcRI-mediated mast cell activation (32,33). Deletion of TXNL2 in mice causes embryonic lethality (34), indicating its role in protecting cells against oxidative stress during embryogenesis. We hypothesized that TXNL2 may play an important role in antagonizing oxidative stress in cancer cells. This study was designed to determine how TXNL2 contributes to the regulation of the cellular redox state in cancer cells and redox-mediated signaling pathways. We characterized the molecular actions of TXNL2 and its involvement in tumor development and metastasis and.