Abstract 1138: The emerging role of thioredoxin-1 on ROS and metabolic reprogramming in cisplatin resistant lung cancer (2012)

Abstract Cisplatin resistance remains a major problem in treating lung cancer. We have discovered that all cisplatin resistant (CR) cells, regardless of their signaling mechanism, possess decreased levels of thioredoxin-1 (TRX1) in vitro and in vivo resulting in higher basal levels of ROS (Reactive Oxygen Species) accumulation. Although decreased TRX1 contributes to higher ROS levels in CR cells, we have found that these CR cells also possess elevate number and active mitochondria as evidence by increased oxygen consumption (4x higher than parental cells) and more intense mitochondria specific fluorescence (mitoTracker) as well as significantly higher mitochondrial membrane potential. These factors could also contribute to higher ROS production. Higher mitochondria activity and ROS also suggest that CR cells may rely more on oxidative metabolism (OXMET). We validated the involvement of specific OXMET components as well as the impact of TRX1 on CR cell metabolism. We examined key proteins in the TCA cycle in SCLC1 (parental), SR2 (resistance counterpart), and TRX1 over-expressing SR2 cells (SR2TRX+). We found that citrate synthetase, the first rate determining enzyme in the TCA cycle, is not changed but ATP-citrate lyase levels exhibit a 3fold increase. Fumarase (FH) which participates in the maintenance of succinate and fumarate equilibrium is increased in SR2 (1.8fold), but attenuates in TRX1 transfected cells, SR2TRX+. This finding implies that SR2 produces more fumarate, a known by-product from urea cycle. We then evaluated argininosuccinate synthetase (ASS) and argininosuccinate lyase (ASL), the two key enzymes in urea cycle which generate arginine. While ASL shows no changes, ASS mRNA is 30fold higher in SR2 compares to SCLC1. These results indicate that SCLC1 require an exogenous arginine supply for their growth while SR2 do not. Indeed, we found that SCLC1 could not withstand arginine free media supplement with citrulline (only 30% of cells survive after 48h). In contrast, SR2 which expressed ASS can survive in arginine free media with 80% of cells still viable at 48h. Importantly, over-expression of TRX1 suppresses ASS in SR2 which in turn sensitizes them to arginine deprivation. It is also known that HIF1α negatively regulates ASS and TRX1 is known to increase HIF1α. Thus lower TRX1 found in CR cells most likely increase ASS via decreased HIF1α expression. Here, we found that HIF1α is less in SR2 which may also contribute to decreased glycolysis. To consider this possibility, we assay lactate production and found lower amount of lactic acid in media from CR cells (0.013 vs. 0.009 nM/well/cell, SCLC1 vs. SR2; p=&amp;lt;0.05). Overall, our findings suggest that alteration in tumor metabolism in CR cells is most likely mediated by TRX1 through ROS. These findings may have future clinical application in treating CR patients using agents which generate ROS or target mitochondria. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1138. doi:1538-7445.AM2012-1138