Research Area:	Cell and Molecular Biology
Technology Exchange:	Drug discovery, ES cell / homologous recombination, Immunohistochemistry, Mass spectrometry, Microscopy (Confocal), Protein interaction, Transcript profiling and Transgenesis
Scientific Themes:	Cancer Biology and Endocrinology & Metabolic Medicine
Keywords:	HLRCC, cancer metabolism, fumarate hydratase, hypoxia, kidney, smooth muscle and renal cancer
Web Links:	NDM website Podcast 'Cancer metabolism'

Background

The drive to understand how altered cellular metabolism and cancer are linked has caused a paradigm shift in the focus of cancer research. The discovery of a mutated metabolic enzyme, isocitrate dehydrogenase 1, that leads to accumulation of the oncometabolite 2-hydroxyglutarate, provided significant direct evidence that dysfunctional metabolism plays an important role in oncogenesis. Striking parallels exist with the Krebs cycle enzyme fumarate hydratase (FH), a tumour suppressor, whose mutation is associated with the development of leiomyomata, renal cysts, and tumors. Loss of FH enzymatic activity results in accumulation of intracellular fumarate which has been proposed to act as a competitive inhibitor of 2-oxoglutarate-dependent oxygenases including the hypoxia-inducible factor (HIF) hydroxylases, thus activating oncogenic HIF pathways. Interestingly, our studies have questioned the role of HIF and have highlighted other candidate mechanisms, in particular the non-enzymatic modification of cysteine residues (succination) that could lead to disruption or loss of protein functions, dysfunctional cell metabolism and cell signalling. 

Research Goals

• The role of fumarate as an oncometabolite
• The contribution of mitochondrial dysfunction to FH-associated neoplasia
• The potential role of protein succination  as a novel mechanism of oncogenesis
• Alternative metabolic pathways utilised by cancer cells
• Investigation of metabolic links between diabetes and cancer

Name	Department	Institution	Country
Prof Tomoyoshi Soga	Institute for Advanced Biosciences	Keio University	Japan
Dr Nicola Ternette	Ubiquitin Proteolysis Group	Oxford University	UK
Dr Benedikt M Kessler	Centre for Cellular and Molecular Physiology	Oxford University	UK
Prof Patrik Rorsman	Oxford Centre for Diabetes, Endocrinology & Metabolism	Oxford University	UK
Prof Christopher Schofield	Chemistry	Oxford University	UK
Dr Reshma Ramracheya	Oxford Centre for Diabetes, Endocrinology & Metabolism	Oxford University	UK
Prof Patrick Maxwell	School of Clinical Medicine	University of Cambridge	UK
Dr Norma Frizzell	Department of Exercise Science	University of South Carolina	USA
Dr Mona El-Bahrawry	Department of Histopathology	Imperial College London	UK
Prof Kamil Kranc	MRC Centre for Regenerative Medicine	University of Edinburgh	UK
Prof Chris W Pugh	Centre for Cellular and Molecular Physiology	Oxford University	UK
Dr Houman Ashrafian	Cardiovascular Medicine	Oxford University	UK
Prof Peter J Ratcliffe FRS	Centre for Cellular and Molecular Physiology	Oxford University	UK
Prof Ian Tomlinson	Wellcome Trust Centre for Human Genetics	Oxford University	UK
Dr Daniel Tennant	School of Cancer Sciences	University of Birmingham	UK
Dr Christian Frezza	MRC Cancer Cell Unit	University of Cambridge	UK

Ternette N, Yang M, Laroyia M, Kitagawa M, O'Flaherty L, Wolhulter K, Igarashi K, Saito K, Kato K, Fischer R, Berquand A, Kessler BM, Lappin T, Frizzell N, Soga T, Adam J, Pollard PJ et al. 2013. Inhibition of Mitochondrial Aconitase by Succination in Fumarate Hydratase Deficiency Cell Reports, 3 (3), pp. 689-700.

Bardella C, Olivero M, Lorenzato A, Geuna M, Adam J, O'Flaherty L, Rustin P, Tomlinson I, Pollard PJ, Di Renzo MF. 2012. Cells lacking the fumarase tumor suppressor are protected from apoptosis through a hypoxia-inducible factor-independent, AMPK-dependent mechanism. Mol Cell Biol, 32 (15), pp. 3081-3094. Read abstract | Read more

Loss-of-function mutations of the tumor suppressor gene encoding fumarase (FH) occur in individuals with hereditary leiomyomatosis and renal cell cancer syndrome (HLRCC). We found that loss of FH activity conferred protection from apoptosis in normal human renal cells and fibroblasts. In FH-defective cells, both hypoxia-inducible factor 1α (HIF-1α) and HIF-2α accumulated, but they were not required for apoptosis protection. Conversely, AMP-activated protein kinase (AMPK) was activated and required, as evidenced by the finding that FH inactivation failed to protect AMPK-null mouse embryo fibroblasts (MEFs) and AMPK-depleted human renal cells. Activated AMPK was detected in renal cysts, which occur in mice with kidney-targeted deletion of Fh1 and in kidney cancers of HLRCC patients. In Fh1-null MEFs, AMPK activation was sustained by fumarate accumulation and not by defective energy metabolism. Addition of fumarate and succinate to kidney cells led to extracellular signal-regulated kinase 1/2 (ERK1/2) and AMPK activation, probably through a receptor-mediated mechanism. These findings reveal a new mechanism of tumorigenesis due to FH loss and an unexpected pro-oncogenic role for AMPK that is important in considering AMPK reactivation as a therapeutic strategy against cancer. Hide abstract

Ashrafian H, Czibik G, Bellahcene M, Aksentijević D, Smith AC, Mitchell SJ, Dodd MS, Kirwan J, Byrne JJ, Ludwig C, Isackson H, Yavari A, Støttrup NB, Contractor H, Cahill TJ, Sahgal N, Ball DR, Birkler RI, Hargreaves I, Tennant DA, Land J, Lygate CA, Johannsen M, Kharbanda RK, Neubauer S, Redwood C, de Cabo R, Ahmet I, Talan M, Günther UL, Robinson AJ, Viant MR, Pollard PJ, Tyler DJ, Watkins H et al. 2012. Fumarate is cardioprotective via activation of the Nrf2 antioxidant pathway. Cell Metab, 15 (3), pp. 361-371. Read abstract | Read more

The citric acid cycle (CAC) metabolite fumarate has been proposed to be cardioprotective; however, its mechanisms of action remain to be determined. To augment cardiac fumarate levels and to assess fumarate's cardioprotective properties, we generated fumarate hydratase (Fh1) cardiac knockout (KO) mice. These fumarate-replete hearts were robustly protected from ischemia-reperfusion injury (I/R). To compensate for the loss of Fh1 activity, KO hearts maintain ATP levels in part by channeling amino acids into the CAC. In addition, by stabilizing the transcriptional regulator Nrf2, Fh1 KO hearts upregulate protective antioxidant response element genes. Supporting the importance of the latter mechanism, clinically relevant doses of dimethylfumarate upregulated Nrf2 and its target genes, hence protecting control hearts, but failed to similarly protect Nrf2-KO hearts in an in vivo model of myocardial infarction. We propose that clinically established fumarate derivatives activate the Nrf2 pathway and are readily testable cytoprotective agents. Hide abstract

Yang M, Soga T, Pollard PJ, Adam J. 2012. The emerging role of fumarate as an oncometabolite. Front Oncol, 2 pp. 85. Read abstract | Read more

The drive to understand how altered cellular metabolism and cancer are linked has caused a paradigm shift in the focus of cancer research. The discovery of a mutated metabolic enzyme, isocitrate dehydrogenase 1, that leads to accumulation of the oncometabolite 2-hydroxyglutarate, provided significant direct evidence that dysfunctional metabolism plays an important role in oncogenesis. Striking parallels exist with the Krebs cycle enzyme fumarate hydratase (FH), a tumor suppressor, whose mutation is associated with the development of leiomyomata, renal cysts, and tumors. Loss of FH enzymatic activity results in accumulation of intracellular fumarate which has been proposed to act as a competitive inhibitor of 2-oxoglutarate-dependent oxygenases including the hypoxia-inducible factor (HIF) hydroxylases, thus activating oncogenic HIF pathways. Interestingly, our studies have questioned the role of HIF and have highlighted other candidate mechanisms, in particular the non-enzymatic modification of cysteine residues (succination) that could lead to disruption or loss of protein functions, dysfunctional cell metabolism and cell signaling. Here, we discuss the evidence for proposing fumarate as an onco-metabolite. Hide abstract

Adam J, Ratcliffe PJ, Pollard PJ. 2011. Novel insights into FH-associated disease are KEAPing the lid on oncogenic HIF signalling. Oncotarget, 2 (11), pp. 820-821.

Adam J, Hatipoglu E, O'Flaherty L, Ternette N, Sahgal N, Lockstone H, Baban D, Nye E, Stamp GW, Wolhuter K, Stevens M, Fischer R, Carmeliet P, Maxwell PH, Pugh CW, Frizzell N, Soga T, Kessler BM, El-Bahrawy M, Ratcliffe PJ, Pollard PJ et al. 2011. Renal cyst formation in Fh1-deficient mice is independent of the Hif/Phd pathway: roles for fumarate in KEAP1 succination and Nrf2 signaling. Cancer Cell, 20 (4), pp. 524-537. Read abstract | Read more

The Krebs cycle enzyme fumarate hydratase (FH) is a human tumor suppressor whose inactivation is associated with the development of leiomyomata, renal cysts, and tumors. It has been proposed that activation of hypoxia inducible factor (HIF) by fumarate-mediated inhibition of HIF prolyl hydroxylases drives oncogenesis. Using a mouse model, we provide genetic evidence that Fh1-associated cyst formation is Hif independent, as is striking upregulation of antioxidant signaling pathways revealed by gene expression profiling. Mechanistic analysis revealed that fumarate modifies cysteine residues within the Kelch-like ECH-associated protein 1 (KEAP1), abrogating its ability to repress the Nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-mediated antioxidant response pathway, suggesting a role for Nrf2 dysregulation in FH-associated cysts and tumors. Hide abstract

Frezza C, Zheng L, Folger O, Rajagopalan KN, MacKenzie ED, Jerby L, Micaroni M, Chaneton B, Adam J, Hedley A, Kalna G, Tomlinson IPM, Pollard PJ, Watson DG, Deberardinis RJ, Shlomi T, Ruppin E, Gottlieb E et al. 2011. Haem oxygenase is synthetically lethal with the tumour suppressor fumarate hydratase NATURE, 477 (7363), pp. 225-U132. | Read more

Bardella C, El-Bahrawy M, Frizzell N, Adam J, Ternette N, Hatipoglu E, Howarth K, O'Flaherty L, Roberts I, Turner G, Taylor J, Giaslakiotis K, Macaulay VM, Harris AL, Chandra A, Lehtonen HJ, Launonen V, Aaltonen LA, Pugh CW, Mihai R, Trudgian D, Kessler B, Baynes JW, Ratcliffe PJ, Tomlinson IP, Pollard PJ et al. 2011. Aberrant succination of proteins in fumarate hydratase-deficient mice and HLRCC patients is a robust biomarker of mutation status. J Pathol, 225 (1), pp. 4-11. Read abstract | Read more

Germline mutations in the FH gene encoding the Krebs cycle enzyme fumarate hydratase predispose to hereditary leiomyomatosis and renal cell cancer (HLRCC) syndrome. FH-deficient cells and tissues accumulate high levels of fumarate, which may act as an oncometabolite and contribute to tumourigenesis. A recently proposed role for fumarate in the covalent modification of cysteine residues to S-(2-succinyl) cysteine (2SC) (termed protein succination) prompted us to assess 2SC levels in our existing models of HLRCC. Herein, using a previously characterized antibody against 2SC, we show that genetic ablation of FH causes high levels of protein succination. We next hypothesized that immunohistochemistry for 2SC would serve as a metabolic biomarker for the in situ detection of FH-deficient tissues. Robust detection of 2SC was observed in Fh1 (murine FH)-deficient renal cysts and in a retrospective series of HLRCC tumours (n = 16) with established FH mutations. Importantly, 2SC was undetectable in normal tissues (n = 200) and tumour types not associated with HLRCC (n = 1342). In a prospective evaluation of cases referred for genetic testing for HLRCC, the presence of 2SC-modified proteins (2SCP) correctly predicted genetic alterations in FH in every case. In two series of unselected type II papillary renal cancer (PRCC), prospectively analysed by 2SCP staining followed by genetic analysis, the biomarker accurately identified previously unsuspected FH mutations (2/33 and 1/36). The investigation of whether metabolites in other tumour types produce protein modification signature(s) that can be assayed using similar strategies will be of interest in future studies of cancer. Hide abstract

Thalhammer A, Bencokova Z, Poole R, Loenarz C, Adam J, O'Flaherty L, Schödel J, Mole D, Giaslakiotis K, Schofield CJ, Hammond EM, Ratcliffe PJ, Pollard PJ et al. 2011. Human AlkB homologue 5 is a nuclear 2-oxoglutarate dependent oxygenase and a direct target of hypoxia-inducible factor 1α (HIF-1α). PLoS One, 6 (1), pp. e16210. Read abstract | Read more

Human 2-oxoglutarate oxygenases catalyse a range of biological oxidations including the demethylation of histone and nucleic acid substrates and the hydroxylation of proteins and small molecules. Some of these processes are centrally involved in regulation of cellular responses to hypoxia. The ALKBH proteins are a sub-family of 2OG oxygenases that are defined by homology to the Escherichia coli DNA-methylation repair enzyme AlkB. Here we report evidence that ALKBH5 is probably unique amongst the ALKBH genes in being a direct transcriptional target of hypoxia inducible factor-1 (HIF-1) and is induced by hypoxia in a range of cell types. We show that purified recombinant ALKBH5 is a bona fide 2OG oxygenase that catalyses the decarboxylation of 2OG but appears to have different prime substrate requirements from those so far defined for other ALKBH family members. Our findings define a new class of HIF-transcriptional target gene and suggest that ALKBH5 may have a role in the regulation of cellular responses to hypoxia. Hide abstract

Frezza C, Pollard PJ, Gottlieb E. 2011. Inborn and acquired metabolic defects in cancer Journal of Molecular Medicine, pp. 1-8.

Ashrafian H, O'Flaherty L, Adam J, Steeples V, Chung YL, East P, Vanharanta S, Lehtonen H, Nye E, Hatipoglu E, Miranda M, Howarth K, Shukla D, Troy H, Griffiths J, Spencer-Dene B, Yusuf M, Volpi E, Maxwell PH, Stamp G, Poulsom R, Pugh CW, Costa B, Bardella C, Di Renzo MF, Kotlikoff MI, Launonen V, Aaltonen L, El-Bahrawy M, Tomlinson I, Pollard PJ et al. 2010. Expression profiling in progressive stages of fumarate-hydratase deficiency: the contribution of metabolic changes to tumorigenesis. Cancer Res, 70 (22), pp. 9153-9165. Read abstract | Read more

Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) is caused by mutations in the Krebs cycle enzyme fumarate hydratase (FH). It has been proposed that "pseudohypoxic" stabilization of hypoxia-inducible factor-α (HIF-α) by fumarate accumulation contributes to tumorigenesis in HLRCC. We hypothesized that an additional direct consequence of FH deficiency is the establishment of a biosynthetic milieu. To investigate this hypothesis, we isolated primary mouse embryonic fibroblast (MEF) lines from Fh1-deficient mice. As predicted, these MEFs upregulated Hif-1α and HIF target genes directly as a result of FH deficiency. In addition, detailed metabolic assessment of these MEFs confirmed their dependence on glycolysis, and an elevated rate of lactate efflux, associated with the upregulation of glycolytic enzymes known to be associated with tumorigenesis. Correspondingly, Fh1-deficient benign murine renal cysts and an advanced human HLRCC-related renal cell carcinoma manifested a prominent and progressive increase in the expression of HIF-α target genes and in genes known to be relevant to tumorigenesis and metastasis. In accord with our hypothesis, in a variety of different FH-deficient tissues, including a novel murine model of Fh1-deficient smooth muscle, we show a striking and progressive upregulation of a tumorigenic metabolic profile, as manifested by increased PKM2 and LDHA protein. Based on the models assessed herein, we infer that that FH deficiency compels cells to adopt an early, reversible, and progressive protumorigenic metabolic milieu that is reminiscent of that driving the Warburg effect. Targets identified in these novel and diverse FH-deficient models represent excellent potential candidates for further mechanistic investigation and therapeutic metabolic manipulation in tumors. Hide abstract

Adam J, Pollard PJ. 2011. In the ring with polycystic kidney disease--avoiding the knockout punch. J Pathol, 223 (1), pp. 1-3. Read abstract | Read more

Autosomal dominant polycystic kidney disease (PKD) is an inherited disease that results from mutations in either polycystin (PKD1) or polycystin 2 (PKD2), both of which are large, complex, and multifunctional proteins whose loss results in the development of numerous fluid-filled cysts and fibrosis that compromise renal function. A number of spontaneous and engineered mouse models of PKD have provided some understanding of many aspects of cyst development, modifier genes, and mechanistic pathways, but fall short of reproducing the human disease accurately. Two recent papers in The Journal of Pathology set out new models using miRNA, or inducible and targeted recombination, that achieve partial or timed suppression of Pkd1. Instead of knocking out Pkd1 immediately, or completely, these more subtle approaches may help deliver more faithful models of this significant human renal disease. Hide abstract

Pollard PJ, Kranc KR. 2010. Hypoxia signaling in hematopoietic stem cells: a double-edged sword. Cell Stem Cell, 7 (3), pp. 276-278. | Read more

O'Flaherty L, Adam J, Heather LC, Zhdanov AV, Chung YL, Miranda MX, Croft J, Olpin S, Clarke K, Pugh CW, Griffiths J, Papkovsky D, Ashrafian H, Ratcliffe PJ, Pollard PJ et al. 2010. Dysregulation of hypoxia pathways in fumarate hydratase-deficient cells is independent of defective mitochondrial metabolism. Hum Mol Genet, 19 (19), pp. 3844-3851. Read abstract | Read more

Mutations in the gene encoding the Krebs cycle enzyme fumarate hydratase (FH) predispose to hereditary leiomyomatosis and renal cell cancer in affected individuals. FH-associated neoplasia is characterized by defective mitochondrial function and by upregulation of transcriptional pathways mediated by hypoxia-inducible factor (HIF), although whether and by what means these processes are linked has been disputed. We analysed the HIF pathway in Fh1-/- mouse embryonic fibroblasts (MEFs), in FH-defective neoplastic tissues and in Fh1-/- MEFs re-expressing either wild-type or an extra-mitochondrial restricted form of FH. These experiments demonstrated that upregulation of HIF-1alpha occurs as a direct consequence of FH inactivation. Fh1-/- cells accumulated intracellular fumarate and manifested severe impairment of HIF prolyl but not asparaginyl hydroxylation which was corrected by provision of exogenous 2-oxoglutarate (2-OG). Re-expression of the extra-mitochondrial form of FH in Fh1-/- cells was sufficient to reduce intracellular fumarate and to correct dysregulation of the HIF pathway completely, even in cells that remained profoundly defective in mitochondrial energy metabolism. The findings indicate that upregulation of HIF-1alpha arises from competitive inhibition of the 2-OG-dependent HIF hydroxylases by fumarate and not from disruption of mitochondrial energy metabolism. Hide abstract

Pollard P, Deheragoda M, Segditsas S, Lewis A, Rowan A, Howarth K, Willis L, Nye E, McCart A, Mandir N, Silver A, Goodlad R, Stamp G, Cockman M, East P, Spencer-Dene B, Poulsom R, Wright N, Tomlinson I et al. 2009. The Apc 1322T mouse develops severe polyposis associated with submaximal nuclear beta-catenin expression. Gastroenterology, 136 (7), pp. 2204-2213.e1-13. Read abstract | Read more

We previously demonstrated that the 2 APC mutations in human colorectal tumors are coselected, because tumorigenesis requires an optimal level of Wnt signaling. We and others subsequently showed that the truncated APC proteins in colorectal tumors usually retain a total of 1-2 beta-catenin binding/degradation repeats (20AARs); very few intestinal tumors have proteins with no 20AARs. The coselection of the "2 hits" at APC makes it difficult to undertake further mechanistic studies in this area in humans. In mice, however, second hits appear to vary with the strain or genetic background used. This suggested the possibility of creating suboptimal Apc genotypes in the mouse. Hide abstract

Pollard PJ, Ratcliffe PJ. 2009. Cancer. Puzzling patterns of predisposition. Science, 324 (5924), pp. 192-194. | Read more

Pollard PJ, Loenarz C, Mole DR, McDonough MA, Gleadle JM, Schofield CJ, Ratcliffe PJ. 2008. Regulation of Jumonji-domain-containing histone demethylases by hypoxia-inducible factor (HIF)-1alpha. Biochem J, 416 (3), pp. 387-394. Read abstract | Read more

The transcription factor HIF (hypoxia-inducible factor) mediates a highly pleiotrophic response to hypoxia. Many recent studies have focused on defining the extent of this transcriptional response. In the present study we have analysed regulation by hypoxia among transcripts encoding human Fe(II)- and 2-oxoglutarate-dependent oxygenases. Our results show that many of these genes are regulated by hypoxia and define two groups of histone demethylases as new classes of hypoxia-regulated genes. Patterns of induction were consistent across a range of cell lines with JMJD1A (where JMJD is Jumonji-domain containing) and JMJD2B demonstrating robust, and JMJD2C more modest, up-regulation by hypoxia. Functional genetic and chromatin immunoprecipitation studies demonstrated the importance of HIF-1alpha in mediating these responses. Given the importance of histone methylation status in defining patterns of gene expression under different physiological and pathophysiological conditions, these findings predict a role for the HIF system in epigenetic regulation. Hide abstract

Pollard PJ, Spencer-Dene B, Shukla D, Howarth K, Nye E, El-Bahrawy M, Deheragoda M, Joannou M, McDonald S, Martin A, Igarashi P, Varsani-Brown S, Rosewell I, Poulsom R, Maxwell P, Stamp GW, Tomlinson IP et al. 2007. Targeted inactivation of fh1 causes proliferative renal cyst development and activation of the hypoxia pathway. Cancer Cell, 11 (4), pp. 311-319. Read abstract | Read more

Germline mutations in the fumarate hydratase (FH) tumor suppressor gene predispose to leiomyomatosis, renal cysts, and renal cell cancer (HLRCC). HLRCC tumors overexpress HIF1alpha and hypoxia pathway genes. We conditionally inactivated mouse Fh1 in the kidney. Fh1 mutants developed multiple clonal renal cysts that overexpressed Hif1alpha and Hif2alpha. Hif targets, such as Glut1 and Vegf, were upregulated. We found that Fh1-deficient murine embryonic stem cells and renal carcinomas from HLRCC showed similar overexpression of HIF and hypoxia pathway components to the mouse cysts. Our data have shown in vivo that pseudohypoxic drive, resulting from HIF1alpha (and HIF2alpha) overexpression, is a direct consequence of Fh1 inactivation. Our mouse may be useful for testing therapeutic interventions that target angiogenesis and HIF-prolyl hydroxylation. Hide abstract

Pollard PJ, El-Bahrawy M, Poulsom R, Elia G, Killick P, Kelly G, Hunt T, Jeffery R, Seedhar P, Barwell J, Latif F, Gleeson MJ, Hodgson SV, Stamp GW, Tomlinson IP, Maher ER et al. 2006. Expression of HIF-1alpha, HIF-2alpha (EPAS1), and their target genes in paraganglioma and pheochromocytoma with VHL and SDH mutations. J Clin Endocrinol Metab, 91 (11), pp. 4593-4598. Read abstract | Read more

Activation of the hypoxia-inducible transcription factors HIF-1 and HIF-2 and a HIF-independent defect in developmental apoptosis have been implicated in the pathogenesis of pheochromocytoma (PCC) associated with VHL, SDHB, and SDHD mutations. Hide abstract

Pollard PJ, Brière JJ, Alam NA, Barwell J, Barclay E, Wortham NC, Hunt T, Mitchell M, Olpin S, Moat SJ, Hargreaves IP, Heales SJ, Chung YL, Griffiths JR, Dalgleish A, McGrath JA, Gleeson MJ, Hodgson SV, Poulsom R, Rustin P, Tomlinson IP et al. 2005. Accumulation of Krebs cycle intermediates and over-expression of HIF1alpha in tumours which result from germline FH and SDH mutations. Hum Mol Genet, 14 (15), pp. 2231-2239. Read abstract | Read more

The nuclear-encoded Krebs cycle enzymes, fumarate hydratase (FH) and succinate dehydrogenase (SDHB, -C and -D), act as tumour suppressors. Germline mutations in FH predispose individuals to leiomyomas and renal cell cancer (HLRCC), whereas mutations in SDH cause paragangliomas and phaeochromocytomas (HPGL). In this study, we have shown that FH-deficient cells and tumours accumulate fumarate and, to a lesser extent, succinate. SDH-deficient tumours principally accumulate succinate. In situ analyses showed that these tumours also have over-expression of hypoxia-inducible factor 1alpha (HIF1alpha), activation of HIF1alphatargets (such as vascular endothelial growth factor) and high microvessel density. We found no evidence of increased reactive oxygen species in our cells. Our data provide in vivo evidence to support the hypothesis that increased succinate and/or fumarate causes stabilization of HIF1alpha a plausible mechanism, inhibition of HIF prolyl hydroxylases, has previously been suggested by in vitro studies. The basic mechanism of tumorigenesis in HPGL and HLRCC is likely to be pseudo-hypoxic drive, just as it is in von Hippel-Lindau syndrome. Hide abstract

Pollard P, Wortham N, Barclay E, Alam A, Elia G, Manek S, Poulsom R, Tomlinson I. 2005. Evidence of increased microvessel density and activation of the hypoxia pathway in tumours from the hereditary leiomyomatosis and renal cell cancer syndrome. J Pathol, 205 (1), pp. 41-49. Read abstract | Read more

The Mendelian tumour syndromes hereditary leiomyomatosis and renal cell cancer (HLRCC) and hereditary paragangliomatosis with phaeochromocytomas (HPGL) result from mutations in nuclear genes (FH and SDHB/C/D, respectively) that encode Krebs cycle enzymes. HPGL tumours are highly vascular and there is evidence that inactivation of SDH leads to activation of the hypoxia/angiogenesis pathway. In contrast, uterine leiomyomas are not generally regarded as particularly vascular lesions. In order to test the possibility that activation of the hypoxia/angiogenesis pathway contributes to tumourigenesis in HLRCC, increased vascularity and hypoxia pathway activation were searched for in HLRCC tumours. Microvessel density was markedly higher in uterine leiomyomas from HLRCC than in the surrounding myometrium; it was notable that sporadic uterine leiomyomas were actually less vascular than normal myometrium. In HLRCC tumours, there was increased expression of transcripts from the hypoxia-responsive genes vascular endothelial growth factor (VEGF) and BNIP3; sporadic uterine leiomyomas did not show these changes. All uterine leiomyomas showed decreased expression of thrombospondin 1. Although sporadic and HLRCC uterine leiomyomas appear to have identical morphology, their pathways of tumourigenesis may be fundamentally different. As is the case in HPGL, it is probable that failure of the Krebs cycle in HLRCC tumours causes inappropriate signalling that the cell is in a hypoxic state, leading to angiogenesis and perhaps directly to clonal expansion and tumour growth through some uncharacterized, cell-autonomous effect. Hide abstract

Loss of fumarate hydratase in a novel model of diabetes-a metabolic link with cancer

Dysregulated metabolism is a common feature of two major diseases-cancer and diabetes. It is exhibited in many cancer cells by the production of energy predominantly through rapid glycolysis and has been identified as a “hallmark” of malignancy (Hanahan and Weinberg 2011), while diabetes is characterized by defective insulin secretion by the beta cells of the pancreatic islets and consequent elevated blood glucose (Ashcroft and Rorsman 2012). Evidence exists too that diabetes increases the ...

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The metabolism of renal cancer

Renal cell cancer (RCC) is a heterogeneous disease characterised by multiple pathological subtypes, each associated with unique genetic mutations, clinical features, and sensitivity to treatment. To date, seven genes have been identified as being involved in RCC: VHL, MET, FLCN, TSC1, TSC2, SDHB and FH, and interestingly, all have roles in metabolism (Linehan, Srinivasan et al. 2010).One of these genes encodes the Krebs cycle enzyme fumarate hydratase (FH) and is mutated in hereditary ...

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The role of fumarate in the epigenetic regulation of renal cancer

The Krebs cycle enzyme Fumarate hydratase (FH) is mutated in hereditary leiomyomatosis and renal cell cancer (HLRCC) (Tomlinson, Alam et al. 2002). Loss or mutation of FH causes an accumulation of fumarate which competes with 2-oxoglutarate (2OG) to inhibit 2OG–dependent enzymes such as the hypoxia inducible factor (HIF) hydroxylases, thus activating oncogenic HIF pathways (Pollard and Ratcliffe 2009). However, through the analysis of novel in vivo models, we have demonstrated recently that ...

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The role of protein succination in diabetes and oncogenesis

This proposal will study the role of a novel, irreversible chemical modification of proteins, S-(2-succinyl)-cysteine (2SC) associated with diabetes and hereditary leiomyomatosis and renal cell cancer syndrome (HLRCC) (Tomlinson, Alam et al. 2002; Alderson, Wang et al. 2006). HLRCC develops as a consequence of mutations in the gene encoding the Krebs cycle enzyme fumarate hydratase (FH) leading to fumarate accumulation, while diabetes is characterized by defective insulin secretion by the beta ...

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