Why tocotrienols work better: insights into the in vitro anti-cancer mechanism of vitamin E

The selective constraint of liver uptake and the sustained metabolism of tocotrienols (T3) demonstrate the need for a prompt detoxification of this class of lipophilic vitamers, and thus the potential for cytotoxic effects in hepatic and extra-hepatic tissues. Hypomethylated (γ and δ) forms of T3 show the highest in vitro and in vivo metabolism and are also the most potent natural xenobiotics of the entire vitamin E family of compounds. These stimulate a stress response with the induction of detoxification and antioxidant genes. Depending on the intensity of this response, these genes may confer cell protection or alternatively they stimulate a senescence-like phenotype with cell cycle inhibition or even mitochondrial toxicity and apoptosis. In cancer cells, the uptake rate and thus the cell content of these vitamers is again higher for the hypomethylated forms, and it is the critical factor that drives the dichotomy between protection and toxicity responses to different T3 forms and doses. These aspects suggest the potential for marked biological activity of hypomethylated “highly metabolized” T3 that may result in cytoprotection and cancer prevention or even chemotherapeutic effects. Cytotoxicity and metabolism of hypomethylated T3 have been extensively investigated in vitro using different cell model systems that will be discussed in this review paper as regard molecular mechanisms and possible relevance in cancer therapy.     

Lactate promotes glutamine uptake and metabolism in oxidative cancer cells

Oxygenated cancer cells have a high metabolic plasticity as they can use glucose, glutamine and lactate as main substrates to support their bioenergetic and biosynthetic activities. Metabolic optimization requires integration. While glycolysis and glutaminolysis can cooperate to support cellular proliferation, oxidative lactate metabolism opposes glycolysis in oxidative cancer cells engaged in a symbiotic relation with their hypoxic/glycolytic neighbors. However, little is known concerning the relationship between oxidative lactate metabolism and glutamine metabolism. Using SiHa and HeLa human cancer cells, this study reports that intracellular lactate signaling promotes glutamine uptake and metabolism in oxidative cancer cells. It depends on the uptake of extracellular lactate by monocarboxylate transporter 1 (MCT1). Lactate first stabilizes hypoxia-inducible factor-2α (HIF-2α), and HIF-2α then transactivates c-Myc in a pathway that mimics a response to hypoxia. Consequently, lactate-induced c-Myc activation triggers the expression of glutamine transporter ASCT2 and of glutaminase 1 (GLS1), resulting in improved glutamine uptake and catabolism. Elucidation of this metabolic dependence could be of therapeutic interest. First, inhibitors of lactate uptake targeting MCT1 are currently entering clinical trials. They have the potential to indirectly repress glutaminolysis. Second, in oxidative cancer cells, resistance to glutaminolysis inhibition could arise from compensation by oxidative lactate metabolism and increased lactate signaling.     

Pharmacokinetics and metabolism of dietary flavonoids in humans

Flavonoids are components of fruit and vegetables that may be beneficial in the prevention of disease such as cancer and cardiovascular diseases. Their beneficial effects will be dependent upon their uptake and disposition in tissues and cells. The metabolism and pharmacokinetics of flavonoids has been an area of active research in the last decade. To date, approximately 100 studies have reported the pharmacokinetics of individual flavonoids in healthy volunteers. The data indicate considerable differences among the different types of dietary flavonoids so that the most abundant flavonoids in the diet do not necessarily produce the highest concentration of flavonoids or their metabolites in vivo. Small intestinal absorption ranges from 0 to 60% of the dose and elimination half-lives (T_1/2) range from 2 to 28 h. Absorbed flavonoids undergo extensive first-pass Phase II metabolism in the small intestine epithelial cells and in the liver. Metabolites conjugated with methyl, glucuronate and sulfate groups are the predominant forms present in plasma. This review summarizes the key differences in absorption, metabolism and pharmacokinetics between the major flavonoids present in the diet. For each flavonoid, the specific metabolites that have been identified so far in vivo are indicated. These data should be considered in the design and interpretation of studies investigating the mechanisms and potential health effects of flavonoids.     

Cellular fatty acid uptake: the contribution of metabolism

PURPOSE OF REVIEW: The aim of this review is to highlight the importance of fatty acid metabolism as a major determinant in fatty acid uptake. In particular, we emphasize how the activation, intracellular transport and downstream metabolism of fatty acids influence their uptake into cells. RECENT FINDINGS: Studies examining fatty acid entry into cells have focused primarily on the roles of plasma membrane proteins or the question of passive diffusion. Recent studies, however, strongly suggest that a driving force governing fatty acid uptake is the metabolic demand for fatty acids. Both gain and loss-of-function experiments indicate that fatty acid uptake can be modulated by activation at both the plasma membrane and internal sites, by intracellular fatty acid binding proteins, and by enzymes in synthetic or degradative metabolic pathways. Although the mechanism is not known, it appears that converting fatty acids to acyl-CoAs and downstream metabolic intermediates increases cellular fatty acid uptake, probably by limiting efflux. SUMMARY: Altered fatty acid metabolism and the accumulation of triacylglycerol and lipid metabolites has been strongly associated with insulin resistance and diabetes, but we do not fully understand how the entry of fatty acids into cells is regulated. Future studies of cellular fatty acid uptake should consider the influence of fatty acid metabolism and the possible interactions between fatty acid metabolism or metabolites and fatty acid transport proteins.     

Biological significance of circulating polyamines in oncology.

Malignant cell proliferation is associated with an increase intracellular polyamine metabolism which itself appears to be in equilibrium with the extracellular circulating polyamine compartments. Erythrocyte polyamine contents may be used clinically as an index of cell proliferation, but the exact biological roles of circulating polyamines, considered as physio(patho)logical parameters involved in the homeostatic(dys)regulation of cell proliferation, remain obscure. It is known that circulating polyamines help promote malignant cell proliferation and metastatic dissemination, but their ultimate targets are not yet completely understood. Either produced by actively proliferating normal or cancer cells, or absorbed from the gastro-intestinal tract (food and colonic microfloral population), circulating polyamines could favour in vivo malignant cell proliferation. 1) Since these organic polycations are more rapidly internalized by cancer cells than by normal ones, do they join and facilitate the malignant intracellular polyamine metabolism? 2) Does binding of polyamines to specific acceptor sites at the surface of cancer cells, thereby modulating endocytosis of biological factors present in the extracellular spaces, modify the homeostatic control of cell proliferation and differentiation? 3) Do modifications of blood polyamine compartmentalization, observed in cancerous organisms, responsible for new enzyme and/or immune capacities, contribute to tumor progression? Answering the above-mentioned questions would lead to new therapeutic approaches in human oncology.     

Rapid assessment of drug resistance of cancer cells to gefitinib and carboplatin using optical imaging

The overall goal of this study was to evaluate optical molecular imaging approaches to determine the drug response of chemotherapy and molecular targeted agents in drug sensitive and drug resistant cell lines. The optical molecular imaging approaches selected in this study were based on changes in intracellular uptake and retention of choline and glucose molecules. The breast cancer cell lines were treated with a molecular targeted anti-EGFR therapy. The bladder cancer cell lines were treated with a conventional chemotherapy approach. Sensitivity of optical molecular imaging approach was also compared with conventional cell viability and cell growth inhibition assays. Results demonstrate that optical molecular imaging of changes in intracellular uptake of metabolites was effective in detecting drug susceptibility for both molecular targeted therapy in breast cancer cells and chemotherapy in bladder cancer cells. Between the selected metabolites for optical molecular imaging, changes in glucose metabolic activity showed higher sensitivity in discrimination between the drug sensitive and drug resistant cell lines. The results demonstrated that optical molecular imaging approaches more significantly sensitive as compared to the conventional cell viability and growth assays. Overall, the results demonstrate potential of optical molecular imaging of metabolic activity to improve sensitivity of in-vitro drug response assays.     

An Overview: The Diversified Role of Mitochondria in Cancer Metabolism

Mitochondria are intracellular organelles involved in energy production, cell metabolism and cell signaling. They are essential not only in the process of ATP synthesis, lipid metabolism and nucleic acid metabolism, but also in tumor development and metastasis. Mutations in mtDNA are commonly found in cancer cells to promote the rewiring of bioenergetics and biosynthesis, various metabolites especially oncometabolites in mitochondria regulate tumor metabolism and progression. And mutation of enzymes in the TCA cycle leads to the unusual accumulation of certain metabolites and oncometabolites. Mitochondria have been demonstrated as the target for cancer treatment. Cancer cells rely on two main energy resources: oxidative phosphorylation (OXPHOS) and glycolysis. By manipulating OXPHOS genes or adjusting the metabolites production in mitochondria, tumor growth can be restrained. For example, enhanced complex I activity increases NAD⁺/NADH to prevent metastasis and progression of cancers. In this review, we discussed mitochondrial function in cancer cell metabolism and specially explored the unique role of mitochondria in cancer stem cells and the tumor microenvironment. Targeting the OXPHOS pathway and mitochondria-related metabolism emerging as a potential therapeutic strategy for various cancers.     

Measurement of homocysteine and related metabolites in human plasma and urine by liquid chromatography electrospray tandem mass spectrometry

The sulfur amino acids, methionine and cysteine play crucial roles in cells as a substrate for protein synthesis, as a methyl donor, and for the synthesis of sulfur-containing compounds, including the key intracellular tripeptide, glutathione. Homocysteine is an intermediary metabolite formed during the metabolism of methionine to cysteine. Dysregulation of homocysteine metabolism is implicated in adverse clinical outcomes such as increased risk of cardiovascular disease, stroke, Alzheimer's disease dementia and osteoporosis. While hyperhomocysteinemia is commonly observed in those conditions, the impact on other related metabolites is condition-specific. Therefore, there exists a need to establish precise and sensitive analytical techniques that allow for the simultaneous measurement of homocysteine and related metabolites in biological samples. The current review outlines the development and use of liquid chromatography electrospray tandem mass spectrometry (LC–MS/MS) to simultaneously measure metabolites involved in sulfur amino acid metabolism. Additionally, extensions of the technique in relation to the measurement of sulfur amino acid and one-carbon kinetics in vivo are discussed. The LC–MS/MS technique has the capacity for unambiguous analyte identification and confirmation, due to its high specificity and sensitivity. It has the greatest potential of being accepted and utilized as a dedicated homocysteine and its related metabolite Standard reference method (SRM).     

Cellular mechanisms of vitamin E uptake: relevance in alpha-tocopherol metabolism and potential implications for disease

alpha-Tocopherol is an essential micronutrient involved in various oxidative stress-related processes. Because of its hydrophobic nature, alpha-tocopherol is transported in plasma lipoproteins, and the pathways involved in its cellular uptake are closely related to the lipoprotein metabolism. alpha-Tocopherol transfer from plasma to cells can occur by different mechanisms such as uptake facilitated by lipid transfer proteins and lipases, receptor-mediated lipoprotein endocytosis, and selective lipid uptake. Here we discuss recent progress in understanding the physiological and pathophysiological relevance of these different pathways for cellular uptake of vitamin E in vivo. This review is mainly focused on the role of the scavenger receptor class B type I (SR-BI) on alpha-tocopherol metabolism and its potential implications for disease conditions.     

Métabolomique et imagerie TEP-FDG des cancers du sein

Cancer metabolism is an essential aspect of tumorogenesis, as cancer cells have increased energy requirements in comparison to normal cells. Metabolomic techniques can provide quantitative data for a large number of small molecules in tissues and enable the analysis of multiple intricate metabolic pathways. Positron emission tomography (PET) using ¹⁸F-Fluorodeoxyglucose (FDG) enables in vivo analysis of glycolysis and is widely used in oncology. High tumor FDG uptake is a prognostic factor in breast cancer and has been associated with tumor aggressively. Seventy breast cancer samples obtained from untreated patients who had underwent FDG-PET imagery were analyzed through an untargeted metabolomic approach using liquid chromatography-mass spectroscopy (LC-MS) to study possible correlations between metabolomic data and FDG uptake. Tumors were split into two groups depending on avidity for FDG as measured with PET. The Compound Discoverer 4.0 software enabled identification of 854 metabolites. PLSDA based models predicted FDG uptake with an accuracy ranging from 0,73 to 0,77. Selected metabolites varied depending on the use of scaling or log transformation. Metabolites correlated with tumor FDG uptake were, among others, glutathione, amino-acids such as glutamate, proline or tyrosine, L-acetyl-carnitine, metabolites from the kynurenine pathway such as L-kynurenine or formyl-kynurenine and polyamines such as N1,N12-diacetylspermine or N1-acetylspermine. These metabolites have been previously shown to reflect cancer aggressivity. The correlation between the glycolytic pathway activation and tumor FDG uptake could not be directly assessed but indirect signs showed a higher glycolytic activity in tumours presenting a higher FDG uptake. Studying new metabolites identified through this process could enable a better understanding of tumor metabolism and identification of new biomarkers.     

Effects of deferasirox and deferiprone on cellular iron load in the human hepatoma cell line HepaRG

Two oral chelators, CP20 (deferiprone) and ICL670 (deferasirox), have been synthesized for the purpose of treating iron overload diseases, especially thalassemias. Given their antiproliferative effects resulting from the essential role played by iron in cell processes, such compounds might also be useful as anticancer agents. In the present study, we tested the impact of these two iron chelators on iron metabolism, in the HepaRG cell line which allowed us to study proliferating and differentiated hepatocytes. ICL670 uptake was greater than the CP20 uptake. The iron depletion induced by ICL670 in differentiated cells increased soluble transferrin receptor expression, decreased intracellular ferritin expression, inhibited ⁵⁵Fe (III) uptake, and reduced the hepatocyte concentration of the labile iron pool. In contrast, CP20 induced an unexpected slight increase in intracellular ferritin, which was amplified by iron-treated chelator exposure. CP20 also promoted Fe(III) uptake in differentiated HepaRG cells, thus leading to an increase of both the labile pool and storage forms of iron evaluated by calcein fluorescence and Perls staining, respectively. In acellular conditions, compared to CP20, iron removing ability from the calcein-Fe(III) complex was 40 times higher for ICL670. On the whole, biological responses of HepaRG cells to ICL670 treatment were characteristic of expected iron depletion. In contrast, the effects of CP20 suggest the potential involvement of this compound in the iron uptake from the external medium into the hepatocytes from the HepaRG cell line, therefore acting like a siderophore in this cell model.     

Resveratrol inhibits cancer cell metabolism by down regulating pyruvate kinase M2 via inhibition of mammalian target of rapamycin

Metabolism of cancer cells with pyruvate kinase M2 (PKM2) at its centre stage has assumed a prime significance in cancer research in recent times. Cancer cell metabolism, characterized by enhanced glucose uptake, production of lactate and anabolism is considered an ideal target for therapeutic interventions. Expression of PKM2 switches metabolism in favor of cancer cells, therefore, the present study was designed to investigate the hitherto unknown effect of resveratrol, a phytoalexin, on PKM2 expression and resultant implications on cancer metabolism. We observed that resveratrol down-regulated PKM2 expression by inhibiting mTOR signaling and suppressed cancer metabolism, adjudged by decreased glucose uptake, lactate production (aerobic glycolysis) and reduced anabolism (macromolecule synthesis) in various cancer cell lines. A contingent decrease in intracellular levels of ribose-5-phosphate (R5P), a critical intermediate of pentose phosphate pathway, accounted for a reduced anabolism. Consequently, the state of suppressed cancer metabolism resulted in decreased cellular proliferation. Interestingly, shRNA-mediated silencing of PKM2 inhibited glucose uptake and lactate production, providing evidence for the critical role of PKM2 and its mediation in the observed effects of resveratrol on cancer metabolism. Further, an over-expression of PKM2 abolished the observed effects of resveratrol, signifying the role of PKM2 downregulation as a critical function of resveratrol. The study reports a novel PKM2-mediated effect of resveratrol on cancer metabolism and provides a new dimension to its therapeutic potential.     

Sucrose metabolism and indoleglucosinolate production of immobilized horseradish cells

The problem of transient primary or secondary metabolism remaining a persisting problem in plant cell cultures is discussed. Since secondary metabolites occurred mainly in differentiated tissues, an effort was made to mimic the cell-to-cell contact of multicellular organisms. Sucrose metabolism and indoleglucosinolate production from immobilized cells of Armoracia rusticanawere investigated. Immobilization acted by reducing the assimilation of the hexoses released into the culture medium. Although sucrose hydrolysis occurred prior to uptake, the decrease of acid invertase activity in immobilized cells was accompanied by an increased yield (2–3-fold) of the intracellular sucrose. Glucosinolates accumulated as indolic forms only during the stationary stage of cell growth. Their amount in immobilized cells may be increased 2-fold compared to the control cultures. On the other hand intracellular sucrose concentration declined whilst the cleavage activity of sucrose synthase increased simultaneously with production of indole-3-methyl- and 4-hydroxy-indole-3-methyl-glucosinolates. Thus, the role of cell immobilization in the biosynthesis of indoleglucosinolates is discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

Red Blood Cells Protein Profile Is Modified in Breast Cancer Patients

Metastasis is the primary cause of death for most breast cancer (BC) patients who succumb to the disease. During the hematogenous dissemination, circulating tumor cells interact with different blood components. Thus, there are microenvironmental and systemic processes contributing to cancer regulation. We have recently published that red blood cells (RBCs) that accompany circulating tumor cells have prognostic value in metastatic BC patients. RBC alterations are related to several diseases. Although the principal known role is gas transport, it has been recently assigned additional functions as regulatory cells on circulation. Hence, to explore their potential contribution to tumor progression, we characterized the proteomic composition of RBCs from 53 BC patients from stages I to III and IV, compared with 33 cancer-free controls. In this work, we observed that RBCs from BC patients showed a different proteomic profile compared to cancer-free controls and between different tumor stages. The differential proteins were mainly related to extracellular components, proteasome, and metabolism. Embryonic hemoglobins, not expected in adults’ RBCs, were detected in BC patients. Besides, lysosome-associated membrane glycoprotein 2 emerge as a new RBCs marker with diagnostic and prognostic potential for metastatic BC patients. Seemingly, RBCs are acquiring modifications in their proteomic composition that probably represents the systemic cancer disease, conditioned by the tumor microenvironment.     

Hydrogen sulfide and nitric oxide metabolites in the blood of free-ranging brown bears and their potential roles in hibernation

During winter hibernation, brown bears (Ursus arctos) lie in dens for half a year without eating while their basal metabolism is largely suppressed. To understand the underlying mechanisms of metabolic depression in hibernation, we measured type and content of blood metabolites of two ubiquitous inhibitors of mitochondrial respiration, hydrogen sulfide (H₂S) and nitric oxide (NO), in winter-hibernating and summer-active free-ranging Scandinavian brown bears. We found that levels of sulfide metabolites were overall similar in summer-active and hibernating bears but their composition in the plasma differed significantly, with a decrease in bound sulfane sulfur in hibernation. High levels of unbound free sulfide correlated with high levels of cysteine (Cys) and with low levels of bound sulfane sulfur, indicating that during hibernation H₂S, in addition to being formed enzymatically from the substrate Cys, may also be regenerated from its oxidation products, including thiosulfate and polysulfides. In the absence of any dietary intake, this shift in the mode of H₂S synthesis would help preserve free Cys for synthesis of glutathione (GSH), a major antioxidant found at high levels in the red blood cells of hibernating bears. In contrast, circulating nitrite and erythrocytic S-nitrosation of glyceraldehyde-3-phosphate dehydrogenase, taken as markers of NO metabolism, did not change appreciably. Our findings reveal that remodeling of H₂S metabolism and enhanced intracellular GSH levels are hallmarks of the aerobic metabolic suppression of hibernating bears.     

Biosynthetic potentials of metabolites and their hierarchical organization

A major challenge in systems biology is to understand how complex and highly connected metabolic networks are organized. The structure of these networks is investigated here by identifying sets of metabolites that have a similar biosynthetic potential. We measure the biosynthetic potential of a particular compound by determining all metabolites than can be produced from it and, following a terminology introduced previously, call this set the scope of the compound. To identify groups of compounds with similar scopes, we apply a hierarchical clustering method. We find that compounds within the same cluster often display similar chemical structures and appear in the same metabolic pathway. For each cluster we define a consensus scope by determining a set of metabolites that is most similar to all scopes within the cluster. This allows for a generalization from scopes of single compounds to scopes of a chemical family. We observe that most of the resulting consensus scopes overlap or are fully contained in others, revealing a hierarchical ordering of metabolites according to their biosynthetic potential. Our investigations show that this hierarchy is not only determined by the chemical complexity of the metabolites, but also strongly by their biological function. As a general tendency, metabolites which are necessary for essential cellular processes exhibit a larger biosynthetic potential than those involved in secondary metabolism. A central result is that chemically very similar substances with different biological functions may differ significantly in their biosynthetic potentials. Our studies provide an important step towards understanding fundamental design principles of metabolic networks determined by the structural and functional complexity of metabolites.     

Endogenous catecholamine synthesis, metabolism, storage and uptake in human neutrophils

Evidence is presented that human neutrophils contain catecholamines and several of their metabolites. In vitro, incubation with alpha-methyl-p-tyrosine or pargyline affects intracellular dopamine, norepinephrine and their metabolites, suggesting catecholamine synthesis and degradation by these cells. Reserpine reduces intracellular dopamine and norepinephrine and desipramine reduces intracellular norepinephrine, suggesting the presence of storage and uptake mechanism. In view of the ability of catecholamines to affect neutrophil function, the present results support the hypothesis that autoregulatory adrenergic mechanisms may exist in these cells.     

Apoptotic cell-derived metabolites in efferocytosis-mediated resolution of inflammation

The resolution of inflammation, as part of standard host defense mechanism, is the process to guarantee timely termination of inflammatory responses and eventual restoration of tissue homeostasis . It is mainly achieved via efferocytosis, during which pro-resolving macrophages clear apoptotic neutrophils at the inflammatory site. Unfortunately, impaired resolution can be the leading cause of chronic inflammatory disorders and some autoimmune diseases. Existing studies have provided relatively comprehensive understandings about the recognition and uptake of apoptotic neutrophils by macrophages during early phases of efferocytosis. However, lack of information concerns macrophage metabolism of apoptotic cell-derived metabolites after being released from phagolysosomes or the relationship between such metabolism and efferocytosis. Notwithstanding, three recent studies have revealed macrophage metabolism of cholesterol, fatty acids and arginine, as well as their respective functions in the context of inflammation-resolution. This review provides an overview of the resolution of inflammation, efferocytosis and the key players involved, followed by a focus on the metabolism of apoptotic cell-derived metabolites within efferocytes. Hypotheses of more potential apoptotic cell-derived metabolites and their possible roles in the resolution are also formulated. Understanding the effect of these metabolites further advances the concept that apoptotic cells act as active players to regulate resolution, and also suggests novel therapeutic strategies for diseases driven by defective resolution and even cancer that may be treated through enhanced efferocytosis.