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RESEARCH HIGHLIGHT

Cell Research (2012) 22:443-446. 2012 IBCB, SIBS, CAS All rights reserved 1001-0602/12 $ 32.00www.nature.com/cr

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Correspondence: Lewis C CantleyTel: +1 617 735 2601; Fax: +1 617 735 2646E-mail: lewis_cantley@hms.harvard.edu

Breathless cancer cells get fat on glutamineDimitrios Anastasiou 1, Lewis C Cantley 1

1 Beth Israel Deaconess Medical Center, Department of Medicine-Division of Signal Transduction; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USACell Research (2012) 22 :443-446. doi:10.1038/cr.2012.5; published online 3 January 2012

Many cancer cells depend on glu-tamine as a fuel for proliferation, yetthe mechanisms by which glutaminesupports cancer metabolism are notfully understood. Two recent studieshighlight an important role for glu-tamine in the synthesis of lipids andprovide novel insights into how glu-tamine metabolism could be targetedfor cancer therapy.

As a consequence of rapid prolif-eration cancer cells require a constantsupply of building blocks to generatedaughter cells. It is increasingly appreci-ated that most oncogenic pathways leadto changes in the cancer proteome thatfacilitate nutrient uptake and reprogrammetabolic processes to promote theutilization of nutrients for anabolism[1]. A boost in lipid synthesis is criticalfor supporting proliferation because, inaddition to serving signaling functions,lipids are essential structural compo-nents of cellular membranes. Cancercells prefer to generate lipids de novo rather than use them directly from theirenvironment [2]. It has been proposedthat glucose can provide carbon atoms

for lipid synthesis in cancer cells. Twostudies by Metallo et al .[3] and Mullenet al .[4] shed fresh light into how an-other key nutrient for cancer, glutamine,also contributes to lipogenesis.

Glucose is predominantly metabo-lized through a series of enzymaticreactions collectively known as glyco-lysis to generate pyruvate. Pyruvate isimported into mitochondria where it isconverted to acetyl-coenzyme A (acetyl-CoA) by the pyruvate dehydrogenase(PDH) enzyme complex (Figure 1A).Acetyl-CoA then condenses with ox-aloacetate to enter the tricarboxylic acidcycle (TCA cycle, also known as theKrebs cycle or citric acid cycle), which

provides reducing power to fuel energygeneration in the form of ATP via theoxidative phosphorylation (OXPHOS)chain. Alternatively, pyruvate is con-verted to lactate by cytoplasmic lactatedehydrogenase (LDH). In cancer cellsthe relative amount of pyruvate oxidizedto lactate versus that metabolized via theTCA cycle is higher compared to normalcells. However, in many cancer cells, afraction of glucose-derived acetyl-CoAis also incorporated into lipids. This is

because mitochondrial citrate exits intothe cytoplasm, where the enzyme ATPcitrate lyase (ACL) converts it to acetyl-CoA and oxaloacetate [5]. Cytoplasmic

acetyl-CoA can then be utilized for lipidsynthesis.Glutamine has long been recognized

as an essential nutrient for the prolifera-tion of most cancer cells [6] but also un-transformed cells in culture. In additionto being utilized for protein synthesis,glutamine also serves as a precursorfor other amino acids [7]. Further-more, glutamine provides nitrogen for

the synthesis of nucleotide bases, anddeoxyribonucleotide supplementationcan rescue a cell cycle arrest caused byglutamine deprivation in transformedcells [8]. However, in some cancer celllines, glutamine can be consumed atrates that exceed the need for nitrogen

provision [9]. One route of glutaminecatabolism involves its conversion toglutamate by glutaminase (GLS) inmitochondria. Glutamate is, in turn,deaminated to generate -ketoglutarate(-KG). -KG can then enter the TCAcycle and keep it going when glucose-derived citrate is re-routed to the cy-toplasm. This function of glutamineis referred to as anaplerosis (whichmeans to top up).

Under hypoxic conditions, whichare frequently encountered in thetumor environment, pyruvate entersmitochondria at reduced rates. This is,in part, because hypoxia induces thestabilization of the transcription factorHIF-1a (hypoxia-inducible factor-1a).One of the transcriptional targets ofHIF-1a is pyruvate dehydrogenasekinase 1 (PDK1), which limits conver-

sion of pyruvate into acetyl-CoA inmitochondria by phosphorylating theE1 subunit of PDH and thus inactivat-ing the enzyme. Limiting pyruvateentry into mitochondria will also reduceglucose-derived carbons available foracetyl-CoA and ultimately lipid syn-thesis. This raises the question: what isthe source of carbon for lipid synthesisunder hypoxia?

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even under normoxic conditions. Incontrast to cells with intact VHL wheremost of lipogenic acetyl-CoA originatedfrom glucose, in VHL-mutated RCCcells up to 70% of lipid carbons werederived from glutamine. When VHLexpression was restored in these cells,lipid synthesis switched back to glucoseas the main carbon source.

Despite early hypotheses that de-fective mitochondria underlie thecharacteristic glucose metabolismexhibited by cancer cells, in most can-cers mitochondrial function is intact[1]. However, some cancers harbormutations in genes encoding mito-chondrial proteins that lead to aberrantmitochondrial metabolism. In suchcancers, glucose-dependent acetyl-CoAgeneration and lipid synthesis wouldalso be compromised. To understandhow cancer cells with mitochondrialdefects generate biosynthetic precur-sors, Mullen et al . [4] generated a pair ofisogenic cell lines where mitochondriahad been previously depleted and sub-sequently re-populated using wild-typemitochondrial DNA or mitochondrialDNA containing a mutation in cyto-chrome c that abrogated mitochondrialrespiration. Indeed, glucose-dependentlipogenesis was severely impairedin cells with defective mitochondria.Interestingly, both cell lines requiredglutamine for anchorage-independentgrowth, suggesting that glutaminesupports proliferation by a respiration-independent pathway. Mullen et al . alsoused 13C-labeled glutamine to trace itsmetabolic fate with mass spectrometryand found that, similar to cells underhypoxia, cells with defective mitochon-

dria generate the majority of lipogenicacetyl-CoA via reductive carboxylationof -KG. This is also the case in RCCcell lines with mutations in the TCAcycle enzyme fumarate hydratase (FH),suggesting that this pathway is alsorelevant for tumors that naturally harbormitochondrial defects. Mullen et al .showed that pharmacologic impairmentof mitochondrial function by respiratory

chain inhibitors (metformin, rotenoneor antimycin) (Figure 1B) also inducedreductive glutamine metabolism in cellswith functional mitochondria, evidencein support of a more general role forreductive glutamine metabolism incancer cells.

Reductive glutamine metabolisminvolves the carboxylation of -KG byCO 2, a reaction that can be performed

by some isocit ra te dehydrogenase(IDH) isoforms by operating in thereverse direction compared to theclassical TCA cycle reaction that theycatalyze. In mammalian cells, thereare three known IDH isoforms: IDH1and IDH2 which are both dependenton NADP + and localize to the cytosoland mitochondria, respectively; andIDH3 which depends on NAD + andalso localizes to mitochondria. To shedlight into the pathway that mediatesreductive glutamine metabolism, bothMetallo et al . and Mullen et al . usedRNA interference to reduce the ex-

pression of all three IDHs individuallyand both groups showed that reductiveglutamine metabolism depends on the

presence of IDH1. Mullen et al . also ob-served a dependence of the pathway onIDH2. Given that they studied the roleof IDHs in respiration-defective cells,it is likely that, in this setting, the TCAcycle runs signi cantly slower and leadsto accumulation of -KG that pushesIDH2 to run in the reverse direction

by mass action (Figure 1B). While thisinterpretation could also be applicableto IDH3, this isoform is thought to oper-ate exclusively in the forward, oxidativedirection, hence its depletion does notaffect reductive glutamine metabolism.

Interestingly, an independent study byWise et al . [10] also pointed to IDH2as the enzyme that mediates reductiveglutamine metabolism under hypoxia.While Wise et al . did not compare theeffects of knocking down all three IDHisoforms to exclude a role for IDH1, it is

possible that, as they used an even loweroxygen concentration for their experi-ments than Metallo et al ., respiration

was limited to levels comparable withthose found in the cells used by Mullenet al ., hence the similarities in their

ndings with the latter study. In com - bination, however, these data suggestthat reductive glutamine metabolismcan take place in both the cytoplasmand mitochondria, possibly depend-ing on the genetic background or localmicroenvironment conditions, such asthe oxygen concentrations experienced

by individual tumors or within differentregions of the same tumor. Regardless,consistent with the idea that IDH1 and2 perform an essential function in pro-viding lipid precursors, knockdown ofeither isoform hindered cell prolifera-tion [3, 4, 10].

The studies by Mullen et al . andMetallo et al ., as well as that of Wise etal ., present compelling evidence thatunder conditions where glucose carbonin ux into mitochondria is limiting,a significant fraction of carbons forlipid synthesis is provided by glutaminethrough a previously underappreciated

pathway. As Metallo et al . propose, bytaking over the burden of providingcarbons for lipid synthesis, reductiveglutamine metabolism frees up glucosecarbons to enter into other anabolic

pathways. Interestingly, this metabolic pathway is not exclusive to cancer cellsas, under hypoxia, non-transformedlung broblasts or activated primary Tlymphocytes also exhibited enhanceddependence on reductive glutaminemetabolism for lipogenesis [3], sug-gesting a broader role for proliferativemetabolism.

Intriguingly, in addition to its roleas a metabolic intermediate in anabolic

reactions, acetyl-CoA is also a precursorfor protein acetylation. Glucose avail-ability has therefore been proposed toaffect histone acetylation and therebygene expression [11]. It is tempting tospeculate that modulation of reductiveglutamine metabolism by hypoxia mayalso have a role in protein acetylation,

but whether it is equivalent to that ofglucose in that it provides acetyl-CoA

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remains to be investigated. Idh1 and Idh2 mutations have re-

cently been identi ed in a variety oftumors. These mutations lead to a sub-stitution of an amino acid at the activesite of the protein that abrogates activityin the forward, oxidative direction, butallows the enzyme to operate in thereductive direction, albeit modifiedso that it produces 2-hydroxyglutarate(2-HG) instead of isocitrate [12, 13].Mutant IDH1/2 enzymes are thereforeattractive targets for developing noveltherapeutics for tumors that harbor thesemutations. The studies of Mullen et al .and Metallo et al . now suggest that, inaddition to other enzymes involved inthe pathway such as aconitase (ACO),wild-type IDH1 or IDH2 could alsoserve as potential candidates for target-ing hyperproliferation diseases. Giventheir role in the TCA cycle, inhibitingspeci cally the reductive direction ofthese enzymes would be essential toselectively hinder tumor cell prolif-eration and this may be challenging.Regardless, by elucidating yet anotheridiosyncrasy of proliferative metabo-lism, these studies provide a better graspof the cancer arsenal and bring us a stepcloser to understanding how metabolic

reprogramming contributes to cancerdevelopment.

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