Abstract 1210: Biopterin metabolism drives tumor progression

Abstract

Abstract Previous studies demonstrate that nitric oxide synthases (NOS’s) can become uncoupled during inflammatory disease states. A hallmark of this uncoupling is superoxide production from NOS in lieu of nitric oxide (NO) resulting from a reduced tetrahydrobiopterin:dihydrobiopterin ratio (BH4:BH2 ratio). As BH4 is a necessary cofactor for NO production from NOS, when this ratio is reduced, NOS becomes a peroxynitrite synthase enhancing the pro-inflammatory/pro-survival tumor environment. Our lab demonstrated that NOS is uncoupled in a variety of tumor cells both in vitro and in vivo, as well as in paired human colorectal samples due to a reduced BH4:BH2 ratio and that recoupling NOS in via exogenous BH4 both in vitro and in vivo leads to decreased tumor cell proliferation. To examine the mechanism behind the reduced BH4:BH2 ratio we used the DSS/AOM model of colitis and colon tumorigenesis. Here we demonstrate that BH4:BH2 ratio is reduced in mouse colon tumors vs mouse normal colon tissue, 1.36±0.35 vs 7.1±0.6. We determined that this reduced ratio is due at least in part to direct oxidation of BH4 to BH2, as treatment of colon cells with an NADPH Oxidase inhibitor increased the BH4:BH2 two-fold; however, this is unable to account for the degree of uncoupling associated with tumor cells. Given that inflammation appears to drive NOS uncoupling, we extracted colons from DSS/AOM treated animals and preferentially harvested colonic epithelial cells and colonic tumor cells at different stages of inflammation to assess the molecular and biochemical changes in molecules important in the regulation and production of BH4. The molecules we focused on were: GTP cyclohydrolase-1 (GCH1, rate limiting enzyme in de novo BH4 synthesis), GTP cyclohydrolase 1 feedback regulatory protein (GCHFRP, regulates GCH1 expression), quinoiddihydropteridine reductase (QDPR, necessary for salvage pathway) and sepiapterin reductase (SR, necessary for both de novo synthesis and salvage of BH4). Transcript levels of these molecules in DSS treated epithelial cells showed that GCH1 and SR were upregulated but QDPR was significantly downregulated when compared with untreated cells, potentially leading to an increase in total BH4 but once oxidized to BH2, a decrease in the salvage back to BH4. Our current research investigates the protein levels as well biochemical activity of these molecules in DSS treated epithelial cells, as well as tumor cells compared with normal cells to further understand the molecular component of the decrease in BH4:BH2 ratio in tumors. Note: This abstract was not presented at the meeting. Citation Format: Christopher Rabender, Asim Alam, Mike Waters, Ross Mikkelsen. Biopterin metabolism drives tumor progression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1210. doi:10.1158/1538-7445.AM2015-1210