Glutamine as an Essential Amino Acid for KRas-Driven Cancer Cells.

Cancer cells consume glutamine, a nonessential amino acid (NEAA), at exceedingly high rates to fulfill their energetic and biosynthetic requirements for proliferation. Glutamine plays distinct roles from essential amino acids in cell cycle progression and in the activation of mammalian target of rapamycin (mTOR). Furthermore, the need of cancer cells for glutamine can be exploited therapeutically - especially those driven by KRas. In this review we explore several distinct cellular roles for glutamine that contribute to glutamine addiction in KRas-driven cancer cells and discuss opportunities for therapeutic intervention created by glutamine addiction.

Phospholipase D-dependent mTOR complex 1 (mTORC1) activation by glutamine.

Glutamine is a key nutrient required for sustaining cell proliferation, contributing to nucleotide, protein, and lipid synthesis. The mTOR complex 1 (mTORC1) is a highly conserved protein complex that acts as a sensor of nutrients, relaying signals for the shift from catabolic to anabolic metabolism. Although glutamine plays an important role in mTORC1 activation, the mechanism is not clear. Here we describe a leucine- and Rag-independent mechanism of mTORC1 activation by glutamine that depends on phospholipase D and the production of phosphatidic acid, which is required for the stability and activity of mTORC1. The phospholipase D-dependent activation of mTORC1 by glutamine depended on the GTPases ADP ribosylation factor 1 (Arf1), RalA, and Rheb. Glutamine deprivation could be rescued by α-ketoglutarate, a downstream metabolite of glutamine. This mechanism represents a distinct nutrient input to mTORC1 that is independent of Rag GTPases and leucine.

Lipid sensing by mTOR complexes via de novo synthesis of phosphatidic acid.

mTOR, the mammalian target of rapamycin, integrates growth factor and nutrient signals to promote a transformation from catabolic to anabolic metabolism, cell growth, and cell cycle progression. Phosphatidic acid (PA) interacts with the FK506-binding protein-12-rapamycin-binding (FRB) domain of mTOR, which stabilizes both mTOR complexes: mTORC1 and mTORC2. We report here that mTORC1 and mTORC2 are activated in response to exogenously supplied fatty acids via the de novo synthesis of PA, a central metabolite for membrane phospholipid biosynthesis. We examined the impact of exogenously supplied fatty acids on mTOR in KRas-driven cancer cells, which are programmed to utilize exogenous lipids. The induction of mTOR by oleic acid was dependent upon the enzymes responsible for de novo synthesis of PA. Suppression of the de novo synthesis of PA resulted in G1 cell cycle arrest. Although it has long been appreciated that mTOR is a sensor of amino acids and glucose, this study reveals that mTOR also senses the presence of lipids via production of PA.

Aspartate Rescues S-phase Arrest Caused by Suppression of Glutamine Utilization in KRas-driven Cancer Cells.

During G1-phase of the cell cycle, normal cells respond first to growth factors that indicate that it is appropriate to divide and then later in G1 to the presence of nutrients that indicate sufficient raw material to generate two daughter cells. Dividing cells rely on the "conditionally essential" amino acid glutamine (Q) as an anaplerotic carbon source for TCA cycle intermediates and as a nitrogen source for nucleotide biosynthesis. We previously reported that while non-transformed cells arrest in the latter portion of G1 upon Q deprivation, mutant KRas-driven cancer cells bypass the G1 checkpoint, and instead, arrest in S-phase. In this study, we report that the arrest of KRas-driven cancer cells in S-phase upon Q deprivation is due to the lack of deoxynucleotides needed for DNA synthesis. The lack of deoxynucleotides causes replicative stress leading to activation of the ataxia telangiectasia and Rad3-related protein (ATR)-mediated DNA damage pathway, which arrests cells in S-phase. The key metabolite generated from Q utilization was aspartate, which is generated from a transaminase reaction whereby Q-derived glutamate is converted to α-ketoglutarate with the concomitant conversion of oxaloacetate to aspartate. Aspartate is a critical metabolite for both purine and pyrimidine nucleotide biosynthesis. This study identifies the molecular basis for the S-phase arrest caused by Q deprivation in KRas-driven cancer cells that arrest in S-phase in response to Q deprivation. Given that arresting cells in S-phase sensitizes cells to apoptotic insult, this study suggests novel therapeutic approaches to KRas-driven cancers.