Abstract 1031: Fumarate hydratase deficiency redirects glucose metabolism of hypoxic cancer cells into the pentose phosphate pathway
Authors: Schito Luana , Rey Sergio , Pawling Judy , Dennis James W. , Wouters Bradly G. , Koritzinsky Marianne 
Published: July 2016

Abstract
Hypoxia is a common feature of all solid cancers and strongly correlated with poor prognosis. As an adaptive response to hypoxia, cancer cells reprogram their metabolism by increasing glycolysis and reductive carboxylation at the expense of mitochondrial respiration, a phenomenon orchestrated by the transcription factor hypoxia inducible factor (HIF) -1. Mutations in the gene encoding for the mitochondrial enzyme fumarate hydratase (FH), found in the hereditary leiomyomatosis and renal cell carcinoma (HLRCC) syndrome, lead to a similar phenotype despite the presence of O2, a phenomenon due to normoxic stabilization of HIF-1 (pseudohypoxia). Here, we report for the first time that FH loss-of-function (LOF) redirects glucose metabolism into the pentose phosphate pathway (PPP) in non-RCC cells subjected to severe hypoxia (O2&lt; .02%). We show that this metabolic shift favors the buildup of biosynthetic precursors supporting hypoxic cell growth and proliferation.
HCT-116 (colon), HeLa (cervix) and H460 (lung) adenocarcinoma cells were transfected with lentiviral vectors encoding for shRNAs targeting FH. Immunoblot analysis showed that FH LOF did not induce pseudohypoxia in these cells. In contrast, HLRCC-derived UOK262 cells showed accumulation of HIF-1 under normoxia which was reversed upon FH re-introduction. A comprehensive analysis utilizing a RT-qPCR array to profile the mRNA expression of 84 HIF-1 target genes, further confirmed that FH LOF did not result in a pseudohypoxic phenotype in HCT-116 cells. An unbiased analysis of 250 metabolites detected by liquid chromatography-tandem mass spectrometry followed by quantitative enrichment analysis, identified glycolysis and the PPP among the most enriched metabolic pathways in hypoxic FH knockdown cells (P&lt; 5×10-8). Since the PPP provides precursors for synthesis of nucleic acids, we analyzed the effect of FH LOF on cell cycle progression and found an inhibition of hypoxia-induced cell cycle arrest in HCT-116 and HeLa cells.
Our study reveals novel insights into the effect of FH loss-of-function in cancer cells and indicates a stark contrast between the pseudohypoxic phenotype described in kidney cancer cell lines obtained from HLRCC patients (i.e, UOK-262) and a HIF- independent mechanism of metabolic rerouting in colon, lung and cervix cancer cell lines. Our data show that FH LOF promotes an anabolic phenotype in hypoxic cancer cells that could be exploited to enhance the therapeutic response targeting this resistant subset of cancer cells.
Citation Format: Luana Schito, Sergio Rey, Judy Pawling, James W. Dennis, Bradly G. Wouters, Marianne Koritzinsky. Fumarate hydratase deficiency redirects glucose metabolism of hypoxic cancer cells into the pentose phosphate pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1031.