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Omega-3 fatty acids from fish oil reduce triglyceride levels in mammals, yet the mechanisms underlying this effect have not been fully clarified, despite the clinical use of omega-3 ethyl esters to treat severe hypertriglyceridemia and reduce cardiovascular disease risk in humans. Here, we identified in bile a class of hypotriglyceridemic omega-3 fatty acid-derived N-acyl taurines (NATs) that, after dietary omega-3 fatty acid supplementation, increased to concentrations similar to those of steroidal bile acids. The biliary docosahexaenoic acid-containing (DHA-containing) NAT C22:6 NAT was increased in human and mouse plasma after dietary omega-3 fatty acid supplementation and potently inhibited intestinal triacylglycerol hydrolysis and lipid absorption. Supporting this observation, genetic elevation of endogenous NAT levels in mice impaired lipid absorption, whereas selective augmentation of C22:6 NAT levels protected against hypertriglyceridemia and fatty liver. When administered pharmacologically, C22:6 NAT accumulated in bile and reduced high-fat diet-induced, but not sucrose-induced, hepatic lipid accumulation in mice, suggesting that C22:6 NAT is a negative feedback mediator that limits excess intestinal lipid absorption. Thus, biliary omega-3 NATs may contribute to the hypotriglyceridemic mechanism of action of fish oil and could influence the design of more potent omega-3 fatty acid-based therapeutics., Introduction Omega-3 fatty acids are essential to many cellular functions, but are not made by mammals and instead are found in highest abundance in fatty fish and fish oil supplements [...]

Dietary vitamin B3 components, such as nicotinamide and nicotinic acid, are precursors to the ubiquitous redox cofactor nicotinamide adenine dinucleotide (NAD+). NAD+ levels are thought to decline with age and disease. While the drivers of this decline remain under intense investigation, strategies have emerged seeking to functionally maintain NAD+ levels through supplementation with NAD+ biosynthetic intermediates. These include marketed products, such as nicotinamide riboside (NR) and its phosphorylated form (NMN). More recent developments have shown that NRH (the reduced form of NR) and its phosphorylated form NMNH also increases NAD+ levels upon administration, although they initially generate NADH (the reduced form of NAD+). Other means to increase the combined levels of NAD+ and NADH, NAD(H), include the inhibition of NAD+-consuming enzymes or activation of biosynthetic pathways. Multiple studies have shown that supplementation with an NAD(H) precursor changes the profile of NAD(H) catabolism. Yet, the pharmacological significance of NAD(H) catabolites is rarely considered although the distribution and abundance of these catabolites differ depending on the NAD(H) precursor used, the species in which the study is conducted, and the tissues used for the quantification. Significantly, some of these metabolites have emerged as biomarkers in physiological disorders and might not be innocuous. Herein, we review the known and emerging catabolites of the NAD(H) metabolome and highlight their biochemical and physiological function as well as key chemical and biochemical reactions leading to their formation. Furthermore, we emphasize the need for analytical methods that inform on the full NAD(H) metabolome since the relative abundance of NAD(H) catabolites informs how NAD(H) precursors are used, recycled, and eliminated. [ABSTRACT FROM AUTHOR]

Isotope labelling of otherwise endogenous metabolites has emerged as a powerful approach to study metabolism-related biological processes, when used in conjunction with nuclear magnetic resonance or chromatography-supported mass spectrometry. Given the advantages of metabolite tracing in uncovering metabolic pathways, there is always a need to develop new methods to generate isotopically labelled compounds. In this direction, we developed a new synthetic route to access the labelled agmatine. To access labelled agmatine, we developed a two-step method that includes the treatment of labelled cyanamide with N-Boc-1,4-butanediamine, followed by a BOC deprotection. Structural confirmation was achieved by 1DNMR, 2DNMR and IR spectroscopy. This isotopologue of agmatine can be very helpful to study the pharmacokinetics and bio-distribution of this neurotransmitter and its metabolites in vitro and in vivo or used as an internal standard in mass spectrometry measurements. [ABSTRACT FROM AUTHOR]

Pyridone adenine dinucleotides (ox-NADs) are redox inactive derivatives of the enzyme cofactor and substrate nicotinamide adenine dinucleotide (NAD) that have a carbonyl group at the C2, C4, or C6 positions of the nicotinamide ring. These aberrant cofactor analogs accumulate in cells under stress and are potential inhibitors of enzymes that use NAD(H). We studied the conformational landscape of ox-NADs in solution using molecular dynamics simulations. Compared to NAD+ and NADH, 2-ox-NAD and 4-ox-NAD have an enhanced propensity for adopting the anti conformation of the pyridone ribose group, whereas 6-ox-NAD exhibits greater syn potential. Consequently, 2-ox-NAD and 4-ox-NAD have increased preference for folding into compact conformations, whereas 6-ox-NAD is more extended. ox-NADs have distinctive preferences for the orientation of the pyridone amide group, which are driven by intramolecular hydrogen bonding and steric interactions. These conformational preferences are compared to those of protein-bound NAD(H). Our results may help in identifying enzymes targeted by ox-NADs. [ABSTRACT FROM AUTHOR]

Simple Summary: Glioblastoma is the most prevalent and lethal brain tumor type, often treated with the DNA alkylating agent temozolomide (TMZ). The cytotoxic DNA lesion O6-methylguanine only accounts for about 9% of the DNA lesions induced by TMZ. The other DNA lesions (>80%) are quickly repaired by the base excision repair (BER) pathway. However, resistance to cytotoxicity of the O6-methylguanine lesion is common in cancer due to defects in the mismatch repair pathway or overexpression of the MGMT repair protein. Therefore, the aim of this study was to find approaches to inhibit the BER pathway to overcome TMZ resistance. We found that combining TMZ with an NAD+ precursor (dihydronicotinamide riboside) and a PARG inhibitor strongly inhibited BER and overcame TMZ resistance. This combination treatment regimen provides a novel approach to consider for glioblastoma. Glioblastoma multiforme (GBM) is an incurable brain cancer with an average survival of approximately 15 months. Temozolomide (TMZ) is a DNA alkylating agent for the treatment of GBM. However, at least 50% of the patients treated with TMZ show poor response, primarily due to elevated expression of the repair protein O6-methylguanine-DNA methyltransferase (MGMT) or due to defects in the mismatch repair (MMR) pathway. These resistance mechanisms are either somatic or arise in response to treatment, highlighting the need to uncover treatments to overcome resistance. We found that administration of the NAD+ precursor dihydronicotinamide riboside (NRH) to raise cellular NAD+ levels combined with PARG inhibition (PARGi) triggers hyperaccumulation of poly(ADP-ribose) (PAR), resulting from both DNA damage-induced and replication-stress-induced PARP1 activation. Here, we show that the NRH/PARGi combination enhances the cytotoxicity of TMZ. Specifically, NRH rapidly increases NAD+ levels in both TMZ-sensitive and TMZ-resistant GBM-derived cells and enhances the accumulation of PAR following TMZ treatment. Furthermore, NRH promotes hyperaccumulation of PAR in the presence of TMZ and PARGi. This combination strongly suppresses the cell growth of GBM cells depleted of MSH6 or cells expressing MGMT, suggesting that this regimen may improve the efficacy of TMZ to overcome treatment resistance in GBM. [ABSTRACT FROM AUTHOR]

Through evolution, eukaryote organisms have developed the ability to use different molecules as independent precursors to generate nicotinamide adenine dinucleotide (NAD+), an essential molecule for life. However, whether these different precursors act in an additive or complementary manner is not truly well understood. Here, we have evaluated how combinations of different NAD+ precursors influence intracellular NAD+ levels. We identified dihydronicotinic acid riboside (NARH) as a new NAD+ precursor in hepatic cells. Second, we demonstrate how NARH, but not any other NAD+ precursor, can act synergistically with nicotinamide riboside (NR) to increase NAD+ levels in cultured cells and in mice. Finally, we demonstrate that the large increase in NAD+ prompted by the combination of these two precursors is due to their chemical interaction and conversion to dihydronicotinamide riboside (NRH). Altogether, this work demonstrates for the first time that NARH can act as a NAD+ precursor in mammalian cells and how different NAD+ precursors can interact and influence each other when co-administered. [ABSTRACT FROM AUTHOR]

We report the synthesis of vitamin B1, B2, and B3 derived nucleotides and dinucleotides generated either through mechanochemical or solution phase chemistry. Under the explored conditions, adenosine and thiamine proved to be particularly amenable to milling conditions. Following optimization of the chemistry related to the formation pyrophosphate bonds, mixed dinucleotides of adenine and thiamine (vitamin B1), riboflavin (vitamin B2), nicotinamide riboside and 3-carboxamide 4-pyridone riboside (both vitamin B3 derivatives) were generated in good yields. Furthermore, we report an efficient synthesis of the MW+4 isotopologue of NAD+ for which deuterium incorporation is present on either side of the dinucleotidic linkage, poised for isotopic tracing experiments by mass spectrometry. Many of these mixed species are novel and present unexplored possibilities to simultaneously enhance or modulate cofactor transporters and enzymes of independent biosynthetic pathways. [ABSTRACT FROM AUTHOR]

Nicotinamide adenine dinucleotide (NAD) metabolism plays an important role in the regulation of immune function. However, a complete picture of how NAD, its metabolites, precursors, and metabolizing enzymes work together in regulating immune function and inflammatory diseases is still not fully understood. Surprisingly, few studies have compared the effect of different forms of vitamin B3 on cellular functions. Therefore, we investigated the role of NAD boosting in the regulation of macrophage activation and function using different NAD precursors supplementation. We compared nicotinamide mononucleotide (NMN), nicotinamide riboside (NR), and nicotinamide (NAM) supplementation, with the recently described potent NAD precursor NRH. Our results show that only NRH supplementation strongly increased NAD+ levels in both bone marrow-derived and THP-1 macrophages. Importantly, NRH supplementation activated a pro-inflammatory phenotype in resting macrophages, inducing gene expression of several cytokines, chemokines, and enzymes. NRH also potentiated the effect of lipopolysaccharide (LPS) on macrophage activation and cytokine gene expression, suggesting that potent NAD+ precursors can promote inflammation in macrophages. The effect of NRH in NAD+ boosting and gene expression was blocked by inhibitors of adenosine kinase, equilibrative nucleoside transporters (ENT), and IκB kinase (IKK). Interestingly, the IKK inhibitor, BMS-345541, blocked the mRNA expression of several enzymes and transporters involved in the NAD boosting effect of NRH, indicating that IKK is also a regulator of NAD metabolism. In conclusion, NAD precursors such as NRH may be important tools to understand the role of NAD and NADH metabolism in the inflammatory process of other immune cells, and to reprogram immune cells to a pro-inflammatory phenotype, such as the M2 to M1 switch in macrophage reprogramming, in the cancer microenvironment. [ABSTRACT FROM AUTHOR]

Research was conducted to understand the mechanics of adenosine 5'-diphosphate (ADP) ribosyl cyclase substrate recognition and effect of the inhibitors on the process. Results point out to the importance of nicotinamide ribose and the nicotinamide aromatic ring for the enzyme-substrate recognition and modifications in the former inhibits this reaction.

E-Phenylitaconic acid has been isolated as a metabolite produced by Azoarcus tolulyticus Tol-4 along with benzylsuccinic acid during anaerobic degradation of toluene. The new metabolite was identified based on samples of benzylfumaric acid, benzylmaleic acid, E-phenylitaconic acid, Z-phenylitaconic acid and corresponding dimethyl diesters. E-phenylitaconic acid could have major implications for defining the pathway of anaerobic toluene metabolism.

The mechanism of inhibition of myo-inositol 1-phosphate (MIP) synthase by myo-2-inosose 1-phosphate at pH 7.2 was determined by examining the structural features for active site binding in myo-2-inosose 1-phosphate and the contribution of its keto form to these interactions. 2-Deoxy-myo-inositol 1-phosphate (dMIP) was used to evaluate the impact of removing the oxidized reaction center of the enzyme. dMIP reduced its inhibitor potency by 47-fold. Meanwhile, dihydroxyacetone phosphate inhibited MIP synthase activity at 700 muMolar at pH 7.2. These results suggest that active site interactions are essential in inhibiting MIP synthase.

The standard methodology for the synthesis of oligonucleotides is the solid‐phase method. Here, we report the first case of mechanochemistry being directly applied to synthesising a dinucleotide on a soluble support. While mechanochemistry eliminates the need to solubilise a supported oligomer in the solution phase for phosphoramidite coupling, the support still confers the benefit of being able to purify the product by precipitation or by simple liquid‐liquid extractions as needed. Furthermore, this study also explores the use of mechanochemistry in the initial functionalisation of the ionic liquid support with a nucleoside monomer. [ABSTRACT FROM AUTHOR]

An analysis of two approaches to phosphonomethylate cyclohexene oxide and substituted cyclohexene oxides obtained from quinic acid and myoinositol involved a study of the tradeoff between synthetic simplicity and regioselectivity during oxirane ring opening. Direct phosphonomethylation of the cyclohexene oxides involves the application of boron trifluoride in conjunction with diisopropyl lithiomethanephosphonate.

Dihydroxyacetone (DHA) is a three‐carbon sugar that is the active ingredient in sunless tanning products and a by‐product of electronic cigarette (e‐cigarette) combustion. Increased use of sunless tanning products and e‐cigarettes has elevated exposures to DHA through inhalation and absorption. Studies have confirmed that DHA is rapidly absorbed into cells and can enter into metabolic pathways following phosphorylation to dihydroxyacetone phosphate (DHAP), a product of fructose metabolism. Recent reports have suggested metabolic imbalance and cellular stress results from DHA exposures. However, the impact of elevated exposure to DHA on human health is currently under‐investigated. We propose that exogenous exposures to DHA increase DHAP levels in cells and mimic fructose exposures to produce oxidative stress, mitochondrial dysfunction, and gene and protein expression changes. Here, we review cell line and animal model exposures to fructose to highlight similarities in the effects produced by exogenous exposures to DHA. Given the long‐term health consequences of fructose exposure, this review emphasizes the pressing need to further examine DHA exposures from sunless tanning products and e‐cigarettes. [ABSTRACT FROM AUTHOR]

Nicotinamide adenine dinucleotide (NAD+), the essential cofactor derived from vitamin B3, is both a coenzyme in redox enzymatic processes and substrate in non-redox events; processes that are intimately implicated in all essential bioenergetics. A decrease in intracellular NAD+ levels is known to cause multiple metabolic complications and age-related disorders. One NAD+ precursor is dihydronicotinamide riboside (NRH), which increases NAD+ levels more potently in both cultured cells and mice than current supplementation strategies with nicotinamide riboside (NR), nicotinamide mononucleotide (NMN) or vitamin B3 (nicotinamide and niacin). However, the consequences of extreme boosts in NAD+ levels are not fully understood. Here, we demonstrate the cell-specific effects of acute NRH exposure in mammalian cells. Hepatocellular carcinoma (HepG3) cells show dose-dependent cytotoxicity when supplemented with 100–1000 μM NRH. Cytotoxicity was not observed in human embryonic kidney (HEK293T) cells over the same dose range of NRH. PUMA and BAX mediate the cell-specific cytotoxicity of NRH in HepG3. When supplementing HepG3 with 100 μM NRH, a significant increase in ROS was observed concurrent with changes in the NAD(P)H and GSH/GSSG pools. NRH altered mitochondrial membrane potential, increased mitochondrial superoxide formation, and induced mitochondrial DNA damage in those cells. NRH also caused metabolic dysregulation, altering mitochondrial respiration. Altogether, we demonstrated the detrimental consequences of an extreme boost of the total NAD (NAD+ + NADH) pool through NRH supplementation in HepG3. The cell-specific effects are likely mediated through the different metabolic fate of NRH in these cells, which warrants further study in other systemic models. [ABSTRACT FROM AUTHOR]

The functional cofactors derived from vitamin B3 are nicotinamide adenine dinucleotide (NAD+), its phosphorylated form, nicotinamide adenine dinucleotide phosphate (NADP+) and their reduced forms (NAD(P)H). These cofactors, together referred as the NAD(P)(H) pool, are intimately implicated in all essential bioenergetics, anabolic and catabolic pathways in all forms of life. This pool also contributes to post-translational protein modifications and second messenger generation. Since NAD+ seats at the cross-road between cell metabolism and cell signaling, manipulation of NAD+ bioavailability through vitamin B3 supplementation has become a valuable nutritional and therapeutic avenue. Yet, much remains unexplored regarding vitamin B3 metabolism. The present review highlights the chemical diversity of the vitamin B3-derived anabolites and catabolites of NAD+ and offers a chemical perspective on the approaches adopted to identify, modulate and measure the contribution of various precursors to the NAD(P)(H) pool. [ABSTRACT FROM AUTHOR]

Axon degeneration, a hallmark of chemotherapy-induced peripheral neuropathy (CIPN), is thought to be caused by a loss of the essential metabolite nicotinamide adenine dinucleotide (NAD+) via the prodegenerative protein SARM1. Some studies challenge this notion, however, and suggest that an aberrant increase in a direct precursor of NAD+, nicotinamide mononucleotide (NMN), rather than loss of NAD+, is responsible. In support of this idea, blocking NMN accumulation in neurons by expressing a bacterial NMN deamidase protected axons from degeneration.We hypothesized that protection could similarly be achieved by reducing NMN production pharmacologically. To achieve this, we took advantage of an alternative pathway for NAD+ generation that goes through the intermediate nicotinic acid mononucleotide (NAMN), rather than NMN. We discovered that nicotinic acid riboside (NAR), a precursor of NAMN, administered in combination with FK866, an inhibitor of the enzyme nicotinamide phosphoribosyltransferase that produces NMN, protected dorsal root ganglion (DRG) axons against vincristineinduced degeneration as well as NMN deamidase. Introducing a different bacterial enzyme that converts NAMN to NMN reversed this protection. Collectively, our data indicate that maintaining NAD+ is not sufficient to protect DRG neurons from vincristineinduced axon degeneration, and elevating NMN, by itself, is not sufficient to cause degeneration. Nonetheless, the combination of FK866 and NAR, which bypasses NMN formation, may provide a therapeutic strategy for neuroprotection. [ABSTRACT FROM AUTHOR]

Research was conducted to examine the hypothesis that a single, active-site base and its conjugate acid mediate all of the non-redox steps separating the substrate from the product. Conformationally restricted (Z)- and (E)-vinylhomophosphonate analogues of the substrate D-glucose 6-phosphate and substrate analogues effectively locked in cyclic, pyranoside forms were synthesized and studied for binding to the active site of myo-inositol 1-phosphate (MIP). Results indicate that MIP synthase exploits the phosphate monoester of 5-keto-D-glucose 6-phosphate to catalyze the intramolecular aldol condensation of the reactive intermediate.

A study was conducted to analyze whether inhibition of myo-inositol 1-phosphate (MIP) synthase in the production of myo-inosotol can be used as alternative to lithium for treatment of manic depression. MIP was reduced through reoxidation of beta-nicotinamide adenine dinucleotide to NAD during the formation of an intermediate of myo-inositol. It is suggested that reduction in consumption of NAD inhibits production of the enzyme. The nature of the interaction of the proposed reaction coordinate intermediate and MIP synthase is discussed.