Impact of dietary supplementation with N-carbamoyl-aspartic acid on serum metabolites and intestinal microflora of sows.

BACKGROUND: N-Carbamoyl-aspartic acid (NCA) is a critical precursor for de novo biosynthesis of pyrimidine nucleotides. To investigate the cumulative effects of maternal supplementation with NCA on the productive performance, serum metabolites and intestinal microbiota of sows, 40 pregnant sows (∼day 80) were assigned into two groups: (1) the control (CON) and (2) treatment (NCA, 50 g t-1 NCA). RESULTS: Results showed that piglets from the NCA group had heavier birth weight than those in the CON group (P < 0.05). In addition, maternal supplementation with NCA decreased the backfat loss of sows during lactation (P < 0.05). Furthermore,16S-rRNA sequencing results revealed that maternal NCA supplementation decreased the abundance of Cellulosilyticum, Fournierella, Anaerovibrio, and Oribacterium genera of sows during late pregnancy (P < 0.05). Similarly, on the 14th day of lactation, maternal supplementation with NCA reduced the diversity of fecal microbes of sows as evidenced by significantly lower observed species, Chao1, and Ace indexes, and decreased the abundance of Lachnospire, Faecalibacterium, and Anaerovorax genera, while enriched the abundance of Catenisphaera (P < 0.05). Untargeted metabolomics showed that a total of 48 differentially abundant biomarkers were identified, which were mainly involved in metabolic pathways of arginine/proline metabolism, phenylalanine/tyrosine metabolism, and fatty acid biosynthesis, etc. CONCLUSION: Overall, the results indicated that NCA supplementation regulated intestinal microbial composition of sows and serum differential metabolites related to arginine, proline, phenylalanine, tyrosine, and fatty acids metabolism that may contribute to regulating the backfat loss of sows, and the birth weight and diarrhea rate of piglets. © 2022 Society of Chemical Industry.

Coordination Chemistry of Phosphate Groups in Systems Including Copper(II) Ions, Phosphoethanolamine and Pyrimidine Nucleotides.

The activity of phosphate groups of phosphoethanolamine and pyrimidine nucleotides (thymidine 5-monophosphate, cytidine 5-monophosphate and uridine 5'monophosphate) in the process of complexation metal ions in aqueous solution was studied. Using the potentiometric method with computer calculation of the data and spectroscopic methods such as UV-Vis, EPR, 13C and 31P NMR as well as FT-IR, the overall stability constants of the complexes as well as coordination modes were obtained. At lower pH, copper(II) ions are complexed only by phosphate groups, whereas the endocyclic nitrogen atom of nucleotides has been identified as a negative center interacting with the -NH3+ groups of phosphoethanolamine.

Structure-activity relationships of pyrimidine nucleotides containing a 5'-α,ß-methylene diphosphonate at the P2Y6 receptor.

The Gq-coupled P2Y6 receptor (P2Y6R) is a component of the purinergic signaling system and functions in inflammatory, cardiovascular and metabolic processes. UDP, the native P2Y6R agonist and P2Y14R partial agonist, is subject to hydrolysis by ectonucleotidases. Therefore, we have synthesized UDP/CDP analogues containing a stabilizing α,ß-methylene bridge as P2Y6R agonists and identified compatible affinity-enhancing pyrimidine modifications. A distal binding region on the receptor was explored with 4-benzyloxyimino cytidine 5'-diphosphate analogues and their potency determined in a calcium mobilization assay. A 4-trifluoromethyl-benzyloxyimino substituent in 25 provided the highest human P2Y6R potency (MRS4554, 0.57 µM), and a 5-fluoro substitution of the cytosine ring in 28 similarly enhanced potency, with >175- and 39-fold selectivity over human P2Y14R, respectively. However, 3-alkyl (31-33, 37, 38), ß-d-arabinofuranose (39) and 6-aza (40) substitution prevented P2Y6R activation. Thus, we have identified new α,ß-methylene bridged N4-extended CDP analogues as P2Y6R agonists that are highly selective over the P2Y14R.

cCMP and cUMP occur in vivo.

Mammalian cells contain the cyclic pyrimidine nucleotides cCMP and cUMP. It is unknown whether these tentative new second messenger molecules occur in vivo. We used high performance liquid chromatography quadrupole tandem mass spectrometry to quantitate nucleoside 3',5'-cyclic monophosphates. cCMP was detected in all organs studied, most notably pancreas, spleen and the female reproductive system. cUMP was not detected in organs, probably due to the intrinsically low sensitivity of mass spectrometry to detect this molecule and organ matrix effects. Intratracheal infection of mice with recombinant Pseudomonas aeruginosa harboring the nucleotidyl cyclase toxin ExoY massively increased cUMP in lung. The identity of cCMP and cUMP in organs was confirmed by high performance liquid chromatography quadrupole time of flight mass spectrometry. cUMP also appeared in serum, urine and faeces following infection. Taken together, this report unequivocally shows for the first time that cCMP and cUMP occur in vivo.

A Patent Review of Human Dihydroorotate Dehydrogenase (hDHODH) Inhibitors as Anticancer Agents and their Other Therapeutic Applications (1999-2022).

Highly proliferating cells, such as cancer cells, are in high demand of pyrimidine nucleotides for their proliferation, accomplished by de novo pyrimidine biosynthesis. The human dihydroorotate dehydrogenase (hDHODH) enzyme plays a vital role in the rate-limiting step of de novo pyrimidine biosynthesis. As a recognised therapeutic target, hDHODH plays a significant role in cancer and other illness. In the past two decades, small molecules as inhibitors hDHODH enzyme have drawn much attention as anticancer agents, and their role in rheumatoid arthritis (RA), and multiple sclerosis (MS). In this patent review, we have compiled patented hDHODH inhibitors published between 1999 and 2022 and discussed the development of hDHODH inhibitors as anticancer agents. Therapeutic potential of small molecules as hDHODH inhibitors for the treatment of various diseases, such as cancer, is very well recognised. Human DHODH inhibitors can rapidly cause intracellular uridine monophosphate (UMP) depletion to produce starvation of pyrimidine bases. Normal cells can better endure a brief period of starvation without the side effects of conventional cytotoxic medication and resume synthesis of nucleic acid and other cellular functions after inhibition of de novo pathway using an alternative salvage pathway. Highly proliferative cells such as cancer cells do not endure starvation because they are in high demand of nucleotides for cell differentiation, which is fulfilled by de novo pyrimidine biosynthesis. In addition, hDHODH inhibitors produce their desired activity at lower doses rather than a cytotoxic dose of other anticancer agents. Thus, inhibition of de novo pyrimidine biosynthesis will create new prospects for the development of novel targeted anticancer agents, which ongoing preclinical and clinical experiments define. Our work brings together a comprehensive patent review of the role of hDHODH in cancer, as well as various patents related to the hDHODH inhibitors and their anticancer and other therapeutic potential. This compiled work on patented DHODH inhibitors will guide researchers in pursuing the most promising drug discovery strategies against the hDHODH enzyme as anticancer agents.

De novo pyrimidine synthesis is a targetable vulnerability in IDH mutant glioma.

Mutations affecting isocitrate dehydrogenase (IDH) enzymes are prevalent in glioma, leukemia, and other cancers. Although mutant IDH inhibitors are effective against leukemia, they seem to be less active in aggressive glioma, underscoring the need for alternative treatment strategies. Through a chemical synthetic lethality screen, we discovered that IDH1-mutant glioma cells are hypersensitive to drugs targeting enzymes in the de novo pyrimidine nucleotide synthesis pathway, including dihydroorotate dehydrogenase (DHODH). We developed a genetically engineered mouse model of mutant IDH1-driven astrocytoma and used it and multiple patient-derived models to show that the brain-penetrant DHODH inhibitor BAY 2402234 displays monotherapy efficacy against IDH-mutant gliomas. Mechanistically, this reflects an obligate dependence of glioma cells on the de novo pyrimidine synthesis pathway and mutant IDH's ability to sensitize to DNA damage upon nucleotide pool imbalance. Our work outlines a tumor-selective, biomarker-guided therapeutic strategy that is poised for clinical translation.

Formyl-Peptide Receptor 2 Signaling Redirects Glucose and Glutamine into Anabolic Pathways in Metabolic Reprogramming of Lung Cancer Cells.

Glucose and glutamine play a crucial role in the metabolic reprogramming of cancer cells. Proliferating cells metabolize glucose in the aerobic glycolysis for energy supply, and glucose and glutamine represent the primary sources of carbon atoms for the biosynthesis of nucleotides, amino acids, and lipids. Glutamine is also an important nitrogen donor for the production of nucleotides, amino acids, and nicotinamide. Several membrane receptors strictly control metabolic reprogramming in cancer cells and are considered new potential therapeutic targets. Formyl-peptide receptor 2 (FPR2) belongs to a small family of GPCRs and is implicated in many physiopathological processes. Its stimulation induces, among other things, NADPH oxidase-dependent ROS generation that, in turn, contributes to intracellular signaling. Previously, by phosphoproteomic analysis, we observed that numerous proteins involved in energetic metabolism are uniquely phosphorylated upon FPR2 stimulation. Herein, we investigated the role of FPR2 in cell metabolism, and we observed that the concentrations of several metabolites associated with the pentose phosphate pathway (PPP), tricarboxylic acid cycle, nucleotide synthesis, and glutamine metabolism, were significantly enhanced in FPR2-stimulated cells. In particular, we found that the binding of specific FPR2 agonists: (i) promotes NADPH production; (ii) activates the non-oxidative phase of PPP; (iii) induces the expression of the ASCT2 glutamine transporter; (iv) regulates oxidative phosphorylation; and (v) induces the de novo synthesis of pyrimidine nucleotides, which requires FPR2-dependent ROS generation.

Comparison of human erythrocyte purine nucleotide metabolism and blood purine and pyrimidine degradation product concentrations before and after acute exercise in trained and sedentary subjects.

This study aimed at evaluating the concentration of erythrocyte purine nucleotides (ATP, ADP, AMP, IMP) in trained and sedentary subjects before and after maximal physical exercise together with measuring the activity of purine metabolism enzymes as well as the concentration of purine (hypoxanthine, xanthine, uric acid) and pyrimidine (uridine) degradation products in blood. The study included 15 male elite rowers [mean age 24.3 ± 2.56 years; maximal oxygen uptake (VO2max) 52.8 ± 4.54 mL/kg/min; endurance and strength training 8.2 ± 0.33 h per week for 6.4 ± 2.52 years] and 15 sedentary control subjects (mean age 23.1 ± 3.41 years; VO2max 43.2 ± 5.20 mL/kg/min). Progressive incremental exercise testing until refusal to continue exercising was conducted on a bicycle ergometer. The concentrations of ATP, ADP, AMP, IMP and the activities of adenine phosphoribosyltransferase (APRT), hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and phosphoribosyl pyrophosphate synthetase (PRPP-S) were determined in erythrocytes. The concentrations of hypoxanthine, xanthine, uric acid and uridine were determined in the whole blood before exercise, after exercise, and 30 min after exercise testing. The study demonstrated a significantly higher concentration of ATP in the erythrocytes of trained subjects which, in part, may be explained by higher metabolic activity on the purine re-synthesis pathway (significantly higher PRPP-S, APRT and HGPRT activities). The ATP concentration, just as the ATP/ADP ratio, as well as an exercise-induced increase in this ratio, correlates with the VO2max level in these subjects which allows them to be considered as the important factors characterising physical capacity and exercise tolerance. Maximal physical exercise in the group of trained subjects results not only in a lower post-exercise increase in the concentration of hypoxanthine, xanthine and uric acid but also in that of uridine. This indicates the possibility of performing high-intensity work with a lower loss of not only purine but also pyrimidine.

Effects of Homoserine Dehydrogenase Deficiency on Production of Cytidine by Mutants of Bacillus subtilis.

A homoserine-requiring mutant, Bacillus subtilis strain No. 615, was isolated from a cytidine-producing mutant strain No. 515. The homoserine dehydrogenase activity of strain No. 615 was reduced to less than 1/50 of that of No. 515. Strain No. 615 accumulated 23.5 mg/ml cytidine in a medium containing 16% glucose, and strain No. 515 accumulated 18.8 mg/ml cytidine under the same conditions. The effects of glucose concentration on cytidine production were examined, and strain No. 615 accumulated 30.2 mg/ml cytidine in a medium containing 20% glucose.