Between bedside and bench Tumor strengths and frailties

Cancer cells thrive owing to different means of survival and proliferation. But despite growing understanding of the biology of and the mechanism of tumorigenesis, complete knowledge of what causes cancer is still lacking. There are multiple hypotheses as to what drives cells to become malignant. One of them is the Warburg effect, which supports that an increase in over oxidative respiration, even in the presence of , may be the cause of cancer. But this premise has not yet been confirmed. In ‘Bench to Bedside’, Michael Ohh peruses a recent study showing a common in people with renal carcinoma and melanoma that may rekindle the debate as to whether a metabolic switch is a major driver in cancer and whether it has potential as a therapeutic target. Every so often, an ‘old’ drug seems to work for a condition that was not previously known. Two recent human studies show that aspirin can prevent colorectal cancer in people genetically predisposed to this disease after several years after treatment. In ‘Bedside to Bench’, Patrick Maxwell discusses the possible mechanisms of action of aspirin in decreasing the risk of developing colorectal cancer.

■ bench to bedside Cancer SUMmOns Otto’s metabolism

Michael Ohh

Cancer cells generate by increas- riers of the E318K variant had at least fivefold a human melanoma cell line identified 22,157 ing nonoxidative breakdown of — increased risk of developing melanoma, RCC sites occupied by MITF(E318K), whereas only glycolysis—instead of the more energy-effi- or both2. 9,107 sites were found for wild-type MITF2. cient oxidative respiration. This preference MITF is a member of the Myc supergene Notably, among the sites that were uniquely for anaerobic over aerobic metabolism in family of basic helix-loop-helix zipper tran- or preferentially occupied by MITF(E318K) cancer cells, even in the presence of oxygen, scription factors that has been proposed to were the promoters for HIF1A, implicated is referred to as the Warburg effect, so named act as a lineage survival oncogene amplified in numerous , and HMOX1, a gene after its discoverer, , in malignant melanoma3, but no previous implicated in both kidney cancer and mela- who over 80 years ago argued the conversion association has been made between MITF noma cell growth. These results suggest that from oxidative respiration to of and RCC. In melanocyte lineage, MITF sumoylation regulates MITF-dependent pro- sugar to be the prime cause of cancer1. Despite upregulates genes that promote develop- moter activity and specificity. the requirement of glycolysis in generating cel- ment, proliferation and invasion, includ- HIF-1a was first identified as a DNA- lular energy and metabolites to support cancer ing CDKN2A and MET. E318K substitution binding protein that activates the transcription © 2012 Nature America, Inc. All rights reserved. America, Inc. © 2012 Nature cell proliferation, scientists have struggled to occurs within a consensus motif (YKXE) of the gene encoding erythropoietin under substantiate Warburg’s hypothesis that the for covalent binding of a small ubiquitin-like hypoxia5. In normal cells under , metabolic switch is required for cancer devel- modifier (SUMO), previously discovered as HIF-1a remains unmodified and escapes the npg opment. A recent study2 reporting the identi- an interacting protein to the death domain destructive recognition of the tumor suppres- fication of a rare germline mutation in people of Fas and tumor necrosis factor receptor 1 sor von Hippel-Lindau (VHL) to form a tran- with coexisting melanoma and renal cell carci- (ref. 4). Bertolotto et al.2 show that MITF is scription factor with its constitutively stable noma (RCC) rekindles this debate on whether covalently modified by SUMO on Lys182 and partner HIF-1b (also known as ARNT) and the Warburg effect is essential for the genesis Lys316, the latter of which was determined coactivator p300 (Fig. 1). HIF transactivates of cancer. to be the major SUMO acceptor site. E318K numerous hypoxia-inducible genes to trigger No common environmental or phenotypic mutation alters a crucial residue within the adaptive cellular responses to compromised risk factors have been associated with mela- SUMO conjugation motif, which markedly oxygen tension such as angiogenesis, eryth- noma and RCC. However, there are individu- impaired MITF sumoylation. ropoiesis and anaerobic metabolism (glycoly- als who develop both melanoma and RCC, Although MITF(E318K) increased the sis)5,6. Cancer cells exploit these otherwise suggesting a possible genetic predisposition activity of the hypoxia-inducible factor 1a normal hypoxic responses to increase the pro- underlying the association between these (HIF1A) promoter compared to wild-type duction or delivery of metabolites, nutrients two seemingly unrelated cancers. A germline MITF; it did not affect the activity of MITF on and oxygen for their survival, proliferation missense substitution, E318K, in microph- other physiological target genes, such as MET, and metastasis. The rare oncogenic mutation thalmia-associated transcription factor TYR and CDKN2A, suggesting that only a sub- E318K in MITF transcription factor increases (MITF) was detected at a significantly higher set of MITF target gene transcription is altered HIF1A transcription, which is predicted to frequency in individuals with melanoma, by this mutation2. Moreover, genomic expres- initiate glycolysis, even under normoxia, and RCC or both cancers, and heterozygote car- sion profiling of RCC cell lines expressing increase the risk of developing melanoma and adenovirus-driven wild-type or MITF(E318K) RCC (Fig. 1). Although increased HIF-1a Michael Ohh is at the University of Toronto, showed differential transcriptional signatures, protein expression or nonoxidative breakdown Toronto, Ontario, Canada. in particular for genes involved in prolifera- of glucose was not shown2, the prediction e-mail: [email protected] tion and inflammation. ChIP-seq analysis of would be that E318K mutation–dependent

30 volume 18 | number 1 | january 2012 nature medicine between bedside and bench

Normal cell Cancer cell energy through oxidation. Tumors deficient in SDH or FH accumulate succinate or fuma- Normoxia Hypoxia Normoxia/hypoxia rate, respectively, which inhibits the activ- SUMO ? ity of PHD, leading to HIF-1a stabilization MITF MITF SUMO MITF E318K SUMO and the induction of various HIF-dependent ? responses, including glycolysis. These obser- ABCB5 vations have provided a direct link between HIF1A HIF1A HIF1A GADD45G TRIM63 mitochondrial defect and tumor development, supporting Warburg’s notion that cancer may SUMO HIF-1α HIF-1α HIF-1α be a disease of the mitochondria. 2 ARNT p300 Although Bertolotto et al. did not unequiv- PHD + O SENP1 2 ocally support or refute Warburg’s prime cause OH SUMO VHL HIF-1α HIF-1α Transcriptional activation of of cancer, the study has provided compelling Cul2eBC + E2 ARNT p300 HIF-responsive genes evidence implicating the SUMO conjugation Rbx1 cascade in cancer. Sumoylation is analogous to OH Transcriptional activation of Pseudohypoxic response ubiquitination and neddylation, which in con- HIF-1 α HIF-responsive genes or aerobic glycolysis Ub cert with the 26S proteasome, form the ubiq- Ub (Warburg effect) Ub uitin-proteasome system (UPS). Inhibitors Ub that alter the UPS have moved from the bench Proteasome-mediated Anaerobic metabolism Cell survival, growth and degradation or glycolysis metastasis to the clinic—bortezomib, a highly selective, reversible inhibitor of the 26S proteasome with Figure 1 MITF-directed regulation of HIF in normal and cancer cells. Left, in normal cells under antitumor activity in multiple myeloma and normoxia, SUMO binds tMITF, decreasing HIF1A transcription. In addition, HIF-1a is hydroxylated mantle cell lymphoma, and MLN4924 (ref. 12), by prolyl-hydroxylase domain–containing enzymes (PHDs). This enables direct binding to VHL, a a selective inhibitor of general NEDD8 conju- component of an E3 ubiquitin ligase called ECV (elongins, Cul2, VHL) that ubiquitinates prolyl- gation of substrates with antitumor properties hydroxylated HIF-1a for subsequent 26S proteasome–mediated destruction. Under hypoxia, SUMO in acute myeloid leukemia (AML), possibly is released, allowing the transcription of and anaerobic metabolism (glycolysis). Right, the HIF1A because NEDD8 regulates proteins vital for MITF(E318K) mutant in melanoma and renal carcinoma cells impairs sumoylation of MITF, resulting 13 in specific increased transcription of HIF1A and other genes compared to wild-type MITF. Even under AML cell survival —but they lack specific- normoxic conditions, this mutation allows cancer cells to activate a pseudohypoxic response or aerobic ity. For instance, MLN4924 suppresses indis- glycolysis, supporting the premise that this Warburg effect may predispose cells to malignancy and criminately the E3 ubiquitin ligase activity of cancer progression. eBC, elongins B and C; Rbx1, RING-box protein 1. all cullin-based RING ligases (CRLs), which target numerous tumor suppressors and onco- desumoylation of MITF would trigger the the relevant melanoma- or RCC-initiating proteins for degradation. Thus inhibition Warburg effect via the upregulation of its prin- cells is sufficient for cellular transformation. of Cul2 scaffold protein neddylation would cipal driver, HIF-1a. Notably, in numerous tumor sup- attenuate the E3 activity of Cul2-CRL, such as Remarkably, HIF-1a itself is also subjected pressor genes, including TP53, PTEN, TSC1 VHL complex ECV (Fig. 1), which may sta- © 2012 Nature America, Inc. All rights reserved. America, Inc. © 2012 Nature to SUMO modification and requires the activ- and TSC2 can promote HIF activation, the bilize HIF-1a and promote a pseudohypoxic ity of SENP1, a deSUMOylase, during hypoxia most heuristic association being the loss of response, including glycolysis. to desumoylate and activate HIF-1a7 (Fig. 1). or inactivating mutations in VHL and the Whereas innovations that control UPS npg This would support the original hypothesis by direct stabilization of HIF-1a, leading to inhibitor specificity will undoubtedly enhance Warburg, who believed that the cause of can- increased glycolysis5. Yet HIF upregulates not their efficacy for a wider spectrum of cancers, cer lies in the metabolic switch from oxidative only glycolysis but also other crucial adaptive a ubiquitin-like SUMO seems to be able to respiration to glycolysis and, thus, in essence responses to hypoxia. Thus, given the lack of direct gene promoter specification of tran- that cancer is a disease of the mitochondria1. complete understanding of the role of HIF- scription factors, suggesting targeting of However, MITF(E318K) increases not only mediated glycolysis in cancer initiation and specific sumoylation events as a promising the transcription of HIF1A but also the tran- the complexity of cancer biology, the Warburg alternative avenue for anticancer therapy. scription of many other genes such as ABCB5, effect has been widely regarded as a conse- a well-known marker of melanoma-initiating quence rather than a cause of cancer. COMPETING FINANCIAL INTERESts The author declares no competing financial interests. cells; GADD45G, a stress sensor that links However, the discovery of mutations in NF-kB to the MAPK pathway; and TRIM63, mitochondrial enzymes causing hereditary 1. Warburg, O. Science 123, 309–314 (1956). one of the most upregulated genes in tumors of cancers has countered this general belief. 2. Bertolotto, C. Nature 480, 94-98 (2011). 3. Garraway, L.A. et al. Nature 436, 117–122 (2005). 2 people with TFE3/TFEB translocation RCC . Germline mutations in genes encoding succi- 4. Okura, T. et al. J. Immunol. 157, 4277–4281 (1996). Thus, it is unclear whether the activation of nate dehydrogenase (SDH) and SDH assem- 5. Semenza, G.L. Science 318, 62–64 (2007). 6. Kaelin, W.G. Jr. & Ratcliffe, P.J. Mol. Cell 30, 393–402 HIF pathway via MITF(E318K) is essential bly factor 2 (SDHAF2) have been identified (2008). for causing melanoma or RCC, leaving the in people with hereditary paraganglioma and 7. Cheng, J., Kang, X., Zhang, S. & Yeh, E.T. Cell 131, debate on Warburg’s prime cause of cancer phaeochromocytoma8–10. In addition, muta- 584–595 (2007). 8. Baysal, B.E. et al. Science 287, 848–851 (2000). unresolved. tions in fumarate hydratase (FH) have been 9. Hao, H.X. et al. Science 325, 1139–1142 (2009). The resolution of this question will first detected in hereditary leiomyomatosis and 10. Niemann, S. & Muller, U. Nat. Genet. 26, 268–270 require elucidation of how this MITF target RCC syndrome11. SDH and FH are enzymes (2000). 11. Tomlinson, I.P. et al. Nat. Genet. 30, 406–410 (2002). gene specification occurs and whether isolated of the tricarboxylic acid (TCA) cycle, which 12. Soucy, T.A. et al. Nature 458, 732–736 (2009). modulation of HIF1A expression via MITF in occurs in the mitochondria to generate 13. Swords, R.T. et al. Blood 115, 3796–3800 (2010).

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