Cellular Metabolism and Anticancer Effects

Cancer

Cancer cells rapidly take in large amounts of nutrients in order to maintain active cell growth. They metabolize these nutrients to synthesize and nucleic acids and to produce energy such as ATP. Even under unfavorable conditions, such as hypoxia or low nutrition, cancer cells can survive by altering their metabolic systems. Therefore, the metabolic systems of cancer cells have attracted many researchers' attention.

A recurring characteristic of cancer cell metabolism is that cancer cells generally prefer to produce ATP via the glycolytic system despite that system being less efficient than mitochondrial oxidative phosphorylation (Warburg effect). As a result, cancer cells take up large amounts of glucose. They also produce a large amount of lactate due to increased glycolytic activity. This method of ATP production allows cancer cells to proliferate even under hypoxia, because the glycolytic system does not require oxygen. Meanwhile, the mitochondria of cancer cells use amino acids and fats to produce NADH. It is commonly recognized that NADH in cancer cell mitochondria is mainly used for redox regulation in addition to ATP production. The abnormal functions of mitochondria in cancer cells result in increased mitochondrial membrane potential (hyperpolarization) and excessive ROS production. Consequently, they produce large amounts of glutathione to maintain the redox balance. Since glutamine and cysteine are essential nutrients for glutathione production, cancer cells take up large amounts of these amino acids. Additionally, since NADPH is required to maintain the reduction of glutathione, the pentose phosphate pathway (downstream from the glycolytic system) and NADH in mitochondria are used to maintain high NADPH levels.

Note: the above information represents the general metabolic characteristics of cancer cells and may vary depending on the type of cancer cell and its environment.

• • Glucose Metabolism Inhibition and Anticancer Effects

    • 

      Cancer cells mainly use the glycolytic system to produce ATP. Thus, the glycolytic system is the most important pathway to understand in the metabolism of cancer cells. Consequently, the glycolytic system has long been a target of anticancer drug development. Although effective anticancer drugs have not yet been developed, the glycolytic system remains a major drug target.

      One of the drug discovery target proteins of the glycolytic system is the glucose transporter (GLUT). Since cancer cells take up large amounts of sugar via glucose transporters, it is possible to suppress the glycolytic system by directly inhibiting glucose transporters. Inhibition of the enzymes responsible for glucose starvation and glycolysis [hexokinase (HK), lactate dehydrogenase (LDH), etc.] as well as inhibition of the efflux of lactate (the end product of glycolysis) are also effective.

      Cell Line	Inhibitor / Inducer	Changes to Cellular Metabolism	Publication
      KO99L	LAT1 inhibitor: JPH203	Mitochondrial MP↓ Autophagy↑	Leukemia, 2015, 29, 1253
      LS174T A549	LAT1 inhibitor: JPH203	Glutamine uptake↓ Leucine uptake↓	J. Biol. Chem., 2018, 293 (8), 2877
      HeLa	LAT1 inhibitor: BCH	Tryptophan uptake↓	J. Immunol., 2011, 187 (4), 1617
      A549	ASCT2 inhibitor: GPNA	Glutamine uptake↓ ROS↑	Clin. Cancer Res., 2013, 19 (3), 560
      MG63.3	GLS inhibitor: CB839	Glutamine↑ Glutamate↓, GSH↓	Cancer&Metab., 2020, 8:4
      OCI-AML3	GLS inhibitor: CB839	ATP↓, NADH/NAD↓ GSH/GSSG↓, Alanine↓ Glutamate↓	Mol. Cancer Ther., 2019, 18 (11), 1937
      H1299 MDA-MB231	GDH inhibitor: R162	ROS↑ GPx activity↓	Cancer Cell, 2015, 27, 257
      A2780	xCT inhibitor: Erastin	Cystine uptake↓ GSH↓	Sci. Rep., 2018, 8 (1), 968
      B16-F10	xCT inhibitor: Sulfasalazine	GSH↓ ROS↑	PLoS One., 2018, 13 (4), e0195151.
      HT-1080	xCT inhibitor: Sorafenib	Cystine uptake↓, GSH↓ Lipid peroxidation↑	Elife, 2014, 3, e02523
      PANC-1	GCL inhibitor: BSO	GSH↓ Lipid peroxidation↑	Oncol. Lett., 2018, 15 (6), 8735
      A549	GCL inhibitor: BSO	Cystine uptake↓, GSH↓	Toxicol. Appl. Pharmacol., 1985, 381

      
      

      Products

      Objective	Product Name	CAT. No.
      Glucose Metabolism Assay	Glucose Assay Kit-WST	G264
      Glucose Uptake Assay	Glucose Uptake Assay Kit-Green	UP02
      Lactic Acid Measurement	Lactate Assay Kit-WST	L256
      NAD+/NADH Assay	NAD/NADH Assay Kit-WST	N509
      NADP+/NADPH Assay	NADP/NADPH Assay Kit-WST	N510
      JC-1 Mitochondrial Membrane Potential Detection	JC-1 MitoMP Detection Kit	MT09
      MT-1 Mitochondrial Membrane Potential Detection	MT-1 MitoMP Detection Kit	MT13

      
      
  • Amino Acid Metabolism Inhibition and Anticancer Effects

    • 

      In cancer cells, which are actively proliferating, amino acids are essential for the synthesis of proteins and nucleic acids. Furthermore, many cancer cells downregulate acetyl CoA production from pyruvate, requiring cancer cells to also use amino acids as nutrient sources for the TCA cycle. This explains why cancer cells have been shown to increase expression of amino acid transporters to take up large amounts of amino acids.

      Glutamine is a raw material for glutathione and a source of α-ketoglutarate, which is essential for the TCA cycle. For these reasons, glutamine uptake and glutaminolysis (glutamine metabolism) have attracted attention as drug targets. As well, the amino acid transporter LAT (L-type amino acid transporter), which is involved in the uptake of many essential amino acids, was found to be overexpressed in many cancer cells. LAT is expected to a be future drug target.

      Because cancer cells produce a large amount of reactive oxygen species, they maintain redox balance by increasing the production of glutathione, an antioxidant. Thus, inhibition of the pathways involved in glutathione production can change the intracellular redox balance and induce methods of cell death such as ferroptosis. In addition to the reason just stated, glutathione also contributes to drug resistance, which is why the pathway involved in glutathione production has become a major target for drug development.

      Cysteine is an amino acid also required for redox regulation and is mainly taken up into cells by the cystine transporter (xCT). Sulfasalazine, long used as an anti-inflammatory drug, and sorafenib, a molecular targeted therapy for cancer, have recently been shown to inhibit xCT. For the same cell death-mediated anticancer effect as glutathione inhibition, xCT has also attracted attention as a target for drug development.

      Changes in Intracellular Metabolism by Each Inhibitor / References

      Cell Line	Inhibitor / Inducer	Changes to Cellular Metablism	Publication
      KO99L	LAT1 inhibitor: JPH203	Mitochondrial MP↓	Leukemia, 2015, 29, 1253
      		Autophagy↑
      LS174T	LAT1 inhibitor: JPH203	Glutamine uptake↓	J. Biol. Chem., 2018, 293 (8), 2877
      A549		Leucine uptake↓
      HeLa	LAT1 inhibitor: BCH	Tryptophan uptake↓	J. Immunol., 2011, 187 (4), 1617
      A549	ASCT2 inhibitor: GPNA	Glutamine uptake↓	Clin. Cancer Res., 2013, 19 (3), 560
      		ROS↑
      MG63.3	GLS inhibitor: CB839	Glutamine↑	Cancer&Metab., 2020, 8:4
      		Glutamate↓, GSH↓
      OCI-AML3	GLS inhibitor: CB839	ATP↓, NADH/NAD↓	Mol. Cancer Ther., 2019, 18 (11), 1937
      		GSH/GSSG↓, Alanine↓
      		Glutamate↓
      H1299	GDH inhibitor: R162	ROS↑	Cancer Cell, 2015, 27, 257
      MDA-MB231		GPx activity↓
      A2780	xCT inhibitor: Erastin	Cystine uptake↓	Sci. Rep., 2018, 8 (1), 968
      		GSH↓
      B16-F10	xCT inhibitor: Sulfasalazine	GSH↓	PLoS One., 2018, 13 (4), e0195151.
      		ROS↑
      HT-1080	xCT inhibitor: Sorafenib	Cystine uptake↓, GSH↓	Elife, 2014, 3, e02523
      		Lipid peroxidation↑
      PANC-1	GCL inhibitor: BSO	GSH↓	Oncol. Lett., 2018, 15 (6), 8735
      		Lipid peroxidation↑
      A549	GCL inhibitor: BSO	Cystine uptake↓, GSH↓	Toxicol. Appl. Pharmacol., 1985, 381

      
      

      Products

      Objective	Product Name	CAT. No.
      NAD+/NADH Assay	NAD/NADH Assay Kit-WST	N509
      JC-1 Mitochondrial Membrane Potential Detection	JC-1 MitoMP Detection Kit	MT09
      MT-1 Mitochondrial Membrane Potential Detection	MT-1 MitoMP Detection Kit	MT13
      Total ROS Detection	ROS Assay Kit-Highly Sensitive DCFH-DA-	R252
      Glutamine Assay	Glutamine Assay Kit-WST	G268
      Glutamate Assay	Glutamate Assay Kit-WST	G269
      Glutathione Quantification	GSSG/GSH Quantification Kit	G257
      Lipid Peroxide Detection	Liperfluo	L248
      Mitochondrial Lipid Peroxide Detection	MitoPeDPP	M466
      Autophagosome Detection	DAPGreen - Autophagy Detection	D676
      	DAPRed - Autophagy Detection	D677

      
      
  • Fatty Acid Metabolism Inhibition and Anticancer Effects

    • Cancer cells, which are actively proliferating, naturally require a large amount of lipids. Thus, intracellular fatty acid synthesis and extracellular fatty acid uptake are both very active. Therefore, many cancer cells have enhanced lipid droplet accumulation. Consequently, the pathway involved in fatty acid production is a very popular therapeutic target for cancer, and many inhibitors have been developed. Additionally, cancer cells perform β-oxidation of fatty acids for efficient energy production in order to compensate for inefficient energy production by the glycolytic system. Thus, drugs targeting the β-oxidation of fatty acids are also being developed.

      

      Changes in Intracellular Metabolism by Each Inhibitor / References

      Cell Line	Inhibitor / Inducer	Changes to Cellular Metabolism	Publication
      CD36 Highly expressed MCF7, SUM159	CD36 inhibitor: SSO	Fatty acid uptake↓ Lipid droplet↓	Cancers, 2019, 11, 2012
      A375, SKMel28	FATP inhibitor: Lipofermata	Fatty acid uptake↓ Lipid droplet↓	Cancer Discov., 2018, 8 (8), 1006
      BT474, MCF7, T47D	ACACA inhibitor: TOFA	Lipid droplet↓ FA oxidation↓	J. Clin. Med., 2020, 9, 87
      ccRCC	SREBP inhibitor: Betulin ACACA inhibitor: TOFA FASN inhibitor: C75	Lipid droplet↓	Mol. Cell Biol., 2017, 37 (22), e00265-17
      NCI-H1703	ACSL inhibitor: Triacsin C	Lipid droplet↓	Int. J. Cancer, 2020, 147, 1680
      UM-UC-3	FAO inhibitor: Etomoxir	Lipid droplet↑ ATP↓, NADPH↓	Clin. Sci., 2019, 133 (15), 1745
      SF188	FAO inhibitor: Etomoxir	ATP↓, NADPH↓ GSH↓ ROS↑	Biochim. Biophys. Acta, 2011, 1807 (6), 726

      
      

      Products

      Objective	Product Name	CAT. No.
      Lipid Droplet Assay	Lipid Droplet Assay Kit-Blue	LD05
      	Lipid Droplet Assay Kit-Deep Red	LD06
      Lipid Droplet Staining	Lipi-Blue	LD01
      	Lipi-Green	LD02
      	Lipi-Red	LD03
      	Lipi-Deep Red	LD04
      NADP+/NADPH Assay	NADP/NADPH Assay Kit-WST	N510
      Glutathione Quantification	GSSG/GSH Quantification Kit	G257
      Total ROS Detection	ROS Assay Kit -Highly Sensitive DCFH-DA-	R252

      
      
  • Cancer Immunity and Cellular Metabolism

    • T cells play a central role in the immune system by eliminating cancer cells. In recent years, it has become clear that metabolism is also involved in the regulation of T cell functions such as differentiation and activation, inspiring more active research on metabolism in cancer immunity.

      Cancer cells take in large amounts of nutrients to maintain their proliferative activity. Activated T cells also require large amounts of nutrients to eliminate cancer cells. Cancer cells and activated T cells thus compete for nutrients - particularly glucose.

      Activated T cells express the PD-1 immune checkpoint receptor on their surface. Cancer cells, in turn, express PD-L1, as the interaction will suppress T cell glucose uptake. It is in this way that cancer cells will regulate the metabolism of immune cells in order to evade the immune system. Therefore, it is important to understand the metabolism of not only cancer cells but also immune cells for the sake of cancer immunology.

      

      Changes in Intracellular Metabolism by Each Inhibitor / References

      Cell Line	Antigen Activation	Changes to Cellular Metabolism	Publication
      T cell	Activation	Glucose uptake ↑	J. Immunol., 2008, 180, 4476
      CD8 (+) T cell	Activation	Glucose uptake ↑ FAO ↓	J. Clin. Invest., 2013, 123, 4479
      CD4 (+) T cell	Activation	Glucose uptake ↑ Lactate ↑, ATP ↑	eLife, 2018, 7, e30938
      CD4 (+) T cell	Activation	Glucose uptake ↑, Lactate ↑ Glutamine uptake ↑ FAO ↓	Nat. Commun., 2015, 6, 6692
      CD4 (+) T cell	Activation (Interactions between PD-1 & PDL1)	Glucose uptake ↓, Lactate ↓ Glutamine uptake ↓ FAO ↑

      
      

      Products

      Objective	Product Name	CAT. No.
      Glucose Metabolism Assay	Glucose Assay Kit-WST	G264
      Glucose Uptake Assay	Glucose Uptake Assay Kit-Green	UP02
      Lactate Detection	Lactate Assay Kit-WST	L256
      Glutamine Detection	Glutamine Assay Kit-WST	G268
      Glutamate Detection	Glutamate Assay Kit-WST	G269
      ATP Measurement	ATP Assay Kit-Luminescence	A550