Because tumor cells are exposed to many nutrients simultaneously, achieving a comprehensive view of tumor metabolism requires understanding how cells integrate these pathways into an over-arching metabolic phenotype. Consequently, discussions about glutamine cannot ignore the rapid glucose utilization that also accompanies cell proliferation. In fact the rates of glucose and glutamine consumption far outpace the utilization of other nutrients available to the cell. Presumably this type of metabolism supports both bioenergetics and the production of precursor pools while sparing other energy-rich substrates like fatty acids and essential amino acids for direct incorporation into macromolecules (DeBerardinis et al., 2006
Warburg's work and many studies since then have focused on the production of lactate from glucose, on the low relative rate of glucose oxidation, and on the high apparent contribution of glycolysis to overall energetics in tumor cells. These have led to the generalization that tumors do not or cannot engage in oxidative metabolism, and that aerobic glycolysis (i.e. conversion of glucose to lactate in the presence of ample oxygen) is a sine qua non
for the tumor metabolic phenotype. A few points should be made about these assumptions. First, metabolism of glucose to lactate is not limited to tumor cells, but is a common feature of rapid cell proliferation (Brand, 1985
; Wang et al., 1976
). The potential advantages of this form of metabolism for proliferating cells have been reviewed (DeBerardinis et al., 2008a
). Second, while many tumor cell lines do exhibit high glycolytic rates, the contribution of glycolysis to total cellular ATP content varies widely, from over 50% as Warburg found to less than 5% in other cells (Zu and Guppy, 2004
). Thus oxidative phosphorylation is not universally impaired in tumor cells. Third, mitochondrial metabolism directly contributes to cell growth because many macromolecular precursors are produced in the TCA cycle (DeBerardinis et al., 2008b
). Even if the glycolytic rate is high enough to support most of the cell's need for ATP synthesis, growth requires that the cells produce lipids, proteins and nucleic acids, and building blocks for these molecules come from the TCA cycle. Therefore, the aerobic glycolysis discovered by Warburg is only one piece of the puzzle of anabolic metabolism in tumor cells.
Our growing understanding of glutamine metabolism promises to help fill in this picture, as the concurrent consumption of glucose and glutamine has obvious theoretical advantages (). The simultaneous metabolism of these two nutrients would support bioenergetics in cells exhibiting the Warburg effect by delivering glutamine-derived α-KG to the TCA cycle for oxidation. Moreover, the entry of α-KG offsets the export of intermediates used in biosynthetic pathways. The process of replenishing TCA cycle intermediates during cell growth (anaplerosis) is a keystone of biomass production and is much less active in quiescent cells. In rapidly proliferating cultured glioblastoma cells, for example, most of the acetyl-CoA pool comes from glucose, while essentially all of the anaplerotic carbon (i.e. the oxaloacetate) comes from glutamine (DeBerardinis et al., 2007
). This results in citrate molecules that contain carbon from both glucose and glutamine (purple arrows in ). After this citrate leaves the TCA cycle, the two glucose-derived carbons are released as acetyl-CoA and used in lipid synthesis. Protein and nucleic acid synthesis also require precursor molecules derived from glucose and glutamine metabolism. Thus during cell proliferation, glucose and glutamine are the two major nutrient inputs, and the primary outputs are biomass (nucleic acids, proteins and lipids) and the by-products secreted as a result of this type of metabolism: lactate, alanine and ammonia ().
Cooperativity between glucose and glutamine metabolism in growing tumors
The importance of glucose and glutamine metabolism to tumor cell biology is underscored by the fact that mutations in tumor suppressors and oncogenes allow cells to by-pass the normal, growth factor-dependent handling of these two nutrients. This has been extensively studied for glucose metabolism. As early as the 1980s it was demonstrated that over-expression of Ras or Myc was sufficient to drive glucose uptake in fibroblasts (Flier et al., 1987
). Subsequently, mutations in many other tumor suppressors and oncogenes have been implicated as drivers of the Warburg effect in tumor cells (reviewed in DeBerardinis, 2008
). While much less is known about the regulation of glutamine metabolism, several reports have recently implicated c-Myc as a major player. Enhanced c-Myc activity was sufficient to drive glutamine metabolism and to impair cell survival in low-glutamine conditions (Wise et al., 2008
; Yuneva et al., 2007
). c-Myc regulates glutamine metabolism in part by stimulating the expression of surface transporters (Wise et al., 2008
). Interestingly, c-Myc also indirectly regulates the protein expression of glutaminase through effects on the microRNAs miR23a and miR23b. Normally, these microRNAs bind to the GLS
3’-UTR and prevent translation of the message. However, c-Myc suppresses miR-23a/b expression, and thus enhanced c-Myc activity de-repressed glutaminase translation and facilitated glutamine oxidation in the mitochondria (Gao et al., 2009
The simplest mechanism to explain the enhanced utilization of both glutamine and glucose by tumor cells is that metabolism of the two nutrients is co-regulated. However, recent findings suggest that they can be regulated by independent signaling pathways within the same cells. In a glioblastoma cell line with genomic c-myc
amplification, inhibition of Akt signaling led to a decrease in glycolysis but had no effect on glutamine metabolism, which was only inhibited when c-Myc was suppressed to normal levels (Wise et al., 2008
). This raises the possibility that the complex metabolic phenotype observed in tumor cells is the result of multiple different signaling inputs, presumably through multiple mutations. This notion is consistent with the observation that serial transduction of human fibroblasts leads to step-wise changes in the cells’ metabolic profile and dependence on particular pathways to sustain ATP supply (Ramanathan et al., 2005
Tumors often display regional heterogeneity in oxygen availability, and this can significantly influence intermediary metabolism independently of tumor genetics. Stabilization of the transcription factor hypoxia-inducible factor-1α (HIF-1α) in hypoxic cells enhances the expression of glucose transporters, glycolytic enzymes, and inhibitory kinases for the pyruvate dehydrogenase complex, all of which serve to increase the production of lactate from glucose (Kim et al., 2006
; Papandreou et al., 2006
; Semenza, 2003
). Little is known about the effects of hypoxia on glutamine metabolism. Presumably in order to survive, cells would at least need to maintain some of the proximal steps of glutamine metabolism to satisfy homeostatic requirements for amino acid and nucleotide synthesis. One study showed that hypoxia could stimulate the import of glutamine in neuroblastoma cells, although downstream metabolism was not examined (Soh et al., 2007
). On the other hand, rat pheochromocytoma cells displayed increased rates of endogenous glutamine synthesis under hypoxia, perhaps reflecting a need for glutamine that exceeded transport capacity (Kobayashi and Millhorn, 2001
). Further work is needed to determine how hypoxia influences glutamine metabolism and whether glutamine influences cell survival during hypoxic stress.
Glucose and glutamine metabolism also have overlapping functions in redox homeostasis, since both can produce NADPH and glutamine has the additional role of supporting GSH biosynthesis. This is an important consideration during cell proliferation, because NADPH is required for biosynthetic reactions and because some production of reactive oxygen species is inevitable during rapid nutrient metabolism. Glutamine withdrawal led to decreased GSH pools in fibroblasts with enhanced c-Myc activity, and to frank oxidative damage in hybridoma cells (Guerin et al., 2006
; Yuneva et al., 2007
). In neither case, however, was the redox stress sufficient to explain the loss of cell viability. On the other hand, in P-493 B lymphoma cells and PC3 prostate cancer cells the loss of cell proliferation and viability triggered by glutamine deprivation or GLS knockdown could be partially reversed with antioxidants. Thus, while impaired glutamine metabolism limits the availability of GSH, this effect is not always responsible for the death of glutamine-dependent cells. It is likely that tumor cells differ in their use of glucose and glutamine to maintain redox balance, and exploiting these differences may be therapeutically useful, especially during therapies that induce oxidative stress.