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OBJECTIVES: Although guidelines recommend antiviral therapy for outpatients with COVID-19 who are at high risk of progressing to severe conditions, such as older adults, many patients do not receive appropriate treatment. Little is known, however, about the physician factors associated with the prescription of guideline-recommended antiviral therapy for patients with COVID-19. DESIGN: A cross-sectional study. SETTING: Data including outpatient visits in primary care clinics in Japan from April to August 2023. PARTICIPANTS: We analysed 30 953 outpatients aged ≥65 years treated with COVID-19 (mean (SD) age, 75.0 (7.6) years; 17 652 women (57.0%)) in 1394 primary care clinics. OUTCOME MEASURES: The primary outcome was the prescription of guideline-recommended antivirals (ie, nirmatrelvir-ritonavir or molnupiravir), adjusted for patient characteristics, months of visits and regions. RESULTS: Antiviral prescriptions were concentrated among a small proportion of physicians; for example, the top 10% of physicians that had the largest number of nirmatrelvir-ritonavir prescriptions accounted for 92.4% of all nirmatrelvir-ritonavir prescriptions. After adjusting for potential confounders, physicians with higher patient volumes were more likely to prescribe guideline-recommended antivirals to their patients (adjusted OR (aOR) for high vs low volume, 1.76; 95% CI 1.31 to 2.38; adjusted p

Nucleoside derivatives are essential to medicinal chemistry because they provide biologically active drugs. A 5´-O-palmitoyl derivative (2) was obtained by directly treating cytidine (1) with palmitoyl chloride. New antimicrobial compounds were developed by transforming the 5´-O-acyl derivative into 2´,3´-di-O-acyl derivatives (3-7) with several functionalities. Physicochemical, spectroscopic, and elemental investigations were used to determine the structures of the synthesized compounds. XRD confirmed the crystalline structure of the synthesized compounds. Compounds 3 and 5 exhibited good antibacterial and antifungal activity against bacteria and fungi in vitro. MIC and MBC investigations were performed on compounds 3 and 5 on the basis of their effectiveness. Most of the compounds resulted in >77% fungal mycelial growth. Compound 6 had antiproliferative effects on EAC cells in vitro, with an IC50 value of 1001.11 µg/ml. A DFT study was used to calculate the FMO and MEP parameters, whereas molecular docking identified microbial pathogen prescription drug possibilities. In silico docking studies of cytidine derivatives against the 4URO and 6COX receptors revealed that compounds 3 and 6 had the best docking. In a stimulating environment, a 100-ns MD simulation revealed stable conformation and binding patterns. MD simulation and MM-PBSA analysis of the 3-4URO and 6-6COX complexes indicated good receptor-best-docked molecule interactions. Finally, in vitro and in silico, SAR studies, the acyl chains, (CH3(CH2)10CO-) and (C6H5CH=CHCO-) incorporated into sugar moieties were shown to have the most promising antimicrobial/anticancer drug-targeting potential.

Cytidine-5’-diphosphocholine (CDP-choline) is a key precursor for the intracellular synthesis of phosphatidylcholine and other phospholipids. Following either intravenous or oral application citicoline (CDP-choline of exogenous origin) undergoes quick decomposition to cytidine and choline, and for this reason it is frequently considered a prodrug. However, upon acute intravenous application in mice citicoline is, on a molar basis, 20 times less toxic than choline. To find out whether cytidine may attenuate toxicity of choline, in the present experiments we compared maximum tolerated doses of single intravenous injections of choline and equimolar mixture of choline and cytidine. We assumed that, if after oral intake a substantial part of citicoline is catabolised already in the intestine and its catabolites enter blood separately, intravenously applied equimolar mixture of cytidine and choline will be markedly less toxic than an equivalent molar dose of choline. However, the maximum tolerated single doses determined in our experiment for choline and equimolar mixture of choline and cytidine were similar. These data suggest that citicoline taken orally is not significantly decomposed in the intestinal lumen, but absorbed to blood as the intact molecule.

The course of pathophysiological mechanisms involved in fragile X-associated tremor/ataxia syndrome (FXTAS) remains largely unknown. Previous proteomics and metabolomics studies conducted in blood samples collected from FMR1 premutation carriers with FXTAS reported abnormalities in energy metabolism, and precursors of gluconeogenesis showed significant changes in plasma expression levels in FMR1 premutation carriers who developed FXTAS. We conducted an analysis of postmortem human brain tissues from 44 donors, 25 brains with FXTAS, and 19 matched controls. We quantified the metabolite relative abundance in the inferior temporal gyrus and the cerebellum using untargeted mass spectrometry (MS)-based metabolomics. We investigated how the metabolite type and abundance relate to the number of cytosine-guanine-guanine (CGG) repeats, to markers of neurodegeneration, and to the symptoms of FXTAS. A metabolomic analysis identified 191 primary metabolites, the data were log-transformed and normalized prior to the analysis, and the relative abundance was compared between the groups. The changes in the relative abundance of a set of metabolites were region-specific with some overlapping results; 22 metabolites showed alterations in the inferior temporal gyrus, while 21 showed differences in the cerebellum. The relative abundance of cytidine was decreased in the inferior temporal gyrus, and a lower abundance was found in the cases with larger CGG expansions; oleamide was significantly decreased in the cerebellum. The abundance of 11 metabolites was influenced by changes in the CGG repeat number. A histological evaluation found an association between the presence of microhemorrhages in the inferior temporal gyrus and a lower abundance of 2,5-dihydroxypyrazine. Our study identified alterations in the metabolites involved in the oxidative-stress response and bioenergetics in the brains of individuals with FXTAS. Significant changes in the abundance of cytidine and oleamide suggest their potential as biomarkers and therapeutic targets for FXTAS.

Aims Cytidine, as an important commercial precursor in the chemical synthesis of antiviral and antitumor drugs, is in great demand in the market. Therefore, the purpose of this study is to build a microbial cell factory with high cytidine production. Methods and results A mutant E. coli NXBG-11-F34 with high tolerance to uridine monophosphate structural analogs and good genetic stability was obtained by atmospheric room temperature plasma (ARTP) mutagenesis combined with high-throughput screening. Then, the udk and rihA genes involved in cytidine catabolism were knocked out by CRISPR/Cas9 gene editing technology, and the recombinant strain E. coli NXBG-13 was constructed. The titer, yield, and productivity of cytidine fermented in a 5 l bioreactor were 15.7 g l−1, 0.164 g g−1, and 0.327 g l−1 h−1, respectively. Transcriptome analysis of the original strain and the recombinant strain E. coli NXBG-13 showed that the gene expression profiles of the two strains changed significantly, and the cytidine de novo pathway gene of the recombinant strain was up-regulated significantly. Conclusions ARTP mutagenesis combined with metabolic engineering is an effective method to construct cytidine-producing strains. [ABSTRACT FROM AUTHOR]

Cytarabine (Ara-C) is a synthetic isomer of cytidine that differs from cytidine and deoxycytidine only in the sugar. The use of arabinose instead of deoxyribose hinders the formation of phosphodiester linkages between pentoses, preventing the DNA chain from elongation and interrupting the DNA synthesis. The minor structural alteration (the inversion of hydroxyl at the 2′ positions of the sugar) leads to change of the biological activity from anti-depressant and DNA/RNA block builder to powerful anti-cancer. Our study aimed to determine the molecular nature of this phenomenon. Three 1H-14N NMR-NQR experimental techniques, followed by solid-state computational modelling (Quantum Theory of Atoms in Molecules, Reduced Density Gradient and 3D Hirshfeld surfaces), Quantitative Structure–Property Relationships, Spackman's Hirshfeld surfaces and Molecular Docking were used. Multifaceted analysis—combining experiments, computational modeling and molecular docking—provides deep insight into three-dimensional packing at the atomic and molecular levels, but is challenging. A spectrum with nine lines indicating the existence of three chemically inequivalent nitrogen sites in the Ara-C molecule was recorded, and the lines were assigned to them. The influence of the structural alteration on the NQR parameters was modeled in the solid (GGA/RPBE). For the comprehensive description of the nature of these interactions several factors were considered, including relative reactivity and the involvement of heavy atoms in various non-covalent interactions. The binding modes in the solid state and complex with dCK were investigated using the novel approaches: radial plots, heatmaps and root-mean-square deviation of the binding mode. We identified the intramolecular OH···O hydrogen bond as the key factor responsible for forcing the glycone conformation and strengthening NH···O bonds with Gln97, Asp133 and Ara128, and stacking with Phe137. The titular butterfly effect is associated with both the inversion and the presence of this intramolecular hydrogen bond. Our study elucidates the differences in the binding modes of Ara-C and cytidine, which should guide the design of more potent anti-cancer and anti-viral analogues. [ABSTRACT FROM AUTHOR]

Cytidine-5’-diphosphocholine (CDP-choline) is a key precursor for the intracellular synthesis of phosphatidylcholine and other phospholipids. Following either intravenous or oral application citicoline (CDP-choline of exogenous origin) undergoes quick decomposition to cytidine and choline, and for this reason it is frequently considered a prodrug. However, upon acute intravenous application in mice citicoline is, on a molar basis, 20 times less toxic than choline. To find out whether cytidine may attenuate toxicity of choline, in the present experiments we compared maximum tolerated doses of single intravenous injections of choline and equimolar mixture of choline and cytidine. We assumed that, if after oral intake a substantial part of citicoline is catabolised already in the intestine and its catabolites enter blood separately, intravenously applied equimolar mixture of cytidine and choline will be markedly less toxic than an equivalent molar dose of choline. However, the maximum tolerated single doses determined in our experiment for choline and equimolar mixture of choline and cytidine were similar. These data suggest that citicoline taken orally is not significantly decomposed in the intestinal lumen, but absorbed to blood as the intact molecule. [ABSTRACT FROM AUTHOR]

The irradiation of water solution of cytidine with UVC light showed that the compound participates in a tautomerism following the kinetics of a first‐order reaction. The PIDA mechanism showed that the proton detachments in the tautomers of cytidine occur through the repulsive 1πσ* excited states and lead to conical intersections S0/S1, whose structures were optimized. More probable is the IRC mechanism, which proceeds along 1nπ* excited state reaction paths. This mechanism includes a water molecule as a catalyst, which assists the phototransformation of amino‐oxo tautomer of cytidine into imino‐hydroxy one. [ABSTRACT FROM AUTHOR]

Here, we synthesized a cytidine-based surfactant and anchored it onto the surface of liquid crystal (LC) droplets to enable the specific detection of silver ions. The synthesized surfactant could induce LC droplets into a radial configuration, and the introduction of silver ions changed the configuration of LC droplets in an intermediate (including escaped-radial and axial) configuration within 2 min. The LC droplet-based sensor developed here can specifically respond to the silver ions as low as 0.6 μM, making it suitable for the detection of Ag+ in environmental and food samples. [Display omitted] Liquid crystal (LC) droplets exhibit unique and sensitive response behaviors to surface absorptions, making them promising candidates for sensing aplications. Here, we have developed a label-free, portable, and cost-effective sensor for the specific and rapid detection of silver ions (Ag+) in drinking-water samples. To achieve this, we have modified cytidine into a surfactant (denoted as C10–M–C) and anchored it onto the surface of LC droplets. The specific binding ability between cytidine and Ag+ enables LC droplets anchored with C10–M–C to respond rapidly and specifically to Ag+ ions. Furthermore, the sensitivity of the response meets requirements for the harmless concentration of Ag+ in drinking-water. The sensor we developed is label-free, portable, and cost-effectively. We believe that the sensor reported here can be applied to the detection of Ag+ in drinking-water and environmental samples. [ABSTRACT FROM AUTHOR]

BackgroundCoronavirus Disease 2019 (Covid-19) continues to challenge the limits of our knowledge and our healthcare system. Here we sought to define the host immune response, a.k.a, the "cytokine storm" that has been implicated in fatal COVID-19 using an AI-based approach.MethodOver 45,000 transcriptomic datasets of viral pandemics were analyzed to extract a 166-gene signature using ACE2 as a 'seed' gene; ACE2 was rationalized because it encodes the receptor that facilitates the entry of SARS-CoV-2 (the virus that causes COVID-19) into host cells. An AI-based approach was used to explore the utility of the signature in navigating the uncharted territory of Covid-19, setting therapeutic goals, and finding therapeutic solutions.FindingsThe 166-gene signature was surprisingly conserved across all viral pandemics, including COVID-19, and a subset of 20-genes classified disease severity, inspiring the nomenclatures ViP and severe-ViP signatures, respectively. The ViP signatures pinpointed a paradoxical phenomenon wherein lung epithelial and myeloid cells mount an IL15 cytokine storm, and epithelial and NK cell senescence and apoptosis determine severity/fatality. Precise therapeutic goals could be formulated; these goals were met in high-dose SARS-CoV-2-challenged hamsters using either neutralizing antibodies that abrogate SARS-CoV-2•ACE2 engagement or a directly acting antiviral agent, EIDD-2801. IL15/IL15RA were elevated in the lungs of patients with fatal disease, and plasma levels of the cytokine prognosticated disease severity.InterpretationThe ViP signatures provide a quantitative and qualitative framework for titrating the immune response in viral pandemics and may serve as a powerful unbiased tool to rapidly assess disease severity and vet candidate drugs.FundingThis work was supported by the National Institutes for Health (NIH) [grants CA151673 and GM138385 (to DS) and AI141630 (to P.G), DK107585-05S1 (SD) and AI155696 (to P.G, D.S and S.D), U19-AI142742 (to S.C, cchiCooperative Centers for Human Immunology)]; Research Grants Program Office (RGPO) from the University of California Office of the President (UCOP) (R00RG2628 & R00RG2642 to P.G, D.S and S.D); the UC San Diego Sanford Stem Cell Clinical Center (to P.G, D.S and S.D); LJI Institutional Funds (to S.C); the VA San Diego Healthcare System Institutional funds (to L.C.A). GDK was supported through The American Association of Immunologists Intersect Fellowship Program for Computational Scientists and Immunologists.One sentence summaryThe host immune response in COVID-19.

The ongoing pandemic caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), necessitates strategies to identify prophylactic and therapeutic drug candidates for rapid clinical deployment. Here, we describe a screening pipeline for the discovery of efficacious SARS-CoV-2 inhibitors. We screen a best-in-class drug repurposing library, ReFRAME, against two high-throughput, high-content imaging infection assays: one using HeLa cells expressing SARS-CoV-2 receptor ACE2 and the other using lung epithelial Calu-3 cells. From nearly 12,000 compounds, we identify 49 (in HeLa-ACE2) and 41 (in Calu-3) compounds capable of selectively inhibiting SARS-CoV-2 replication. Notably, most screen hits are cell-line specific, likely due to different virus entry mechanisms or host cell-specific sensitivities to modulators. Among these promising hits, the antivirals nelfinavir and the parent of prodrug MK-4482 possess desirable in vitro activity, pharmacokinetic and human safety profiles, and both reduce SARS-CoV-2 replication in an orthogonal human differentiated primary cell model. Furthermore, MK-4482 effectively blocks SARS-CoV-2 infection in a hamster model. Overall, we identify direct-acting antivirals as the most promising compounds for drug repurposing, additional compounds that may have value in combination therapies, and tool compounds for identification of viral host cell targets.

Adenosine Deaminases Acting on RNA (ADARs) convert adenosine to inosine in double stranded RNA. Human ADARs can be directed to predetermined target sites in the transcriptome by complementary guide strands, allowing for the correction of disease-causing mutations at the RNA level. Here we use structural information available for ADAR2-RNA complexes to guide the design of nucleoside analogs for the position in the guide strand that contacts a conserved glutamic acid residue in ADARs (E488 in human ADAR2), which flips the adenosine into the ADAR active site for deamination. Mutating this residue to glutamine (E488Q) results in higher activity because of the hydrogen bond donating ability of Q488 to N3 of the orphan cytidine on the guide strand. We describe the evaluation of cytidine analogs for this position that stabilize an activated conformation of the enzyme-RNA complex and increase catalytic rate for deamination by the wild-type enzyme. A new crystal structure of ADAR2 bound to duplex RNA bearing a cytidine analog revealed a close contact between E488, stabilized by an additional hydrogen bond and altered charge distribution when compared to cytidine. In human cells and mouse primary liver fibroblasts, this single nucleotide modification increased directed editing yields when compared to an otherwise identical guide oligonucleotide. Our results show that modification of the guide RNA can mimic the effect of hyperactive mutants and advance the approach of recruiting endogenous ADARs for site-directed RNA editing.

Feline calicivirus (FCV) causes upper respiratory tract disease (URTD) and sporadic outbreaks of virulent systemic disease (FCV-VSD). The basis for the increased pathogenicity of FCV-VSD viruses is incompletely understood, and antivirals for FCV-VSD have yet to be developed. We investigated the clinicoepidemiology and viral features of three FCV-VSD outbreaks in Australia and evaluated the in vitro efficacy of nitazoxanide (NTZ), 2'-C-methylcytidine (2CMC) and NITD-008 against FCV-VSD viruses. Overall mortality among 23 cases of FCV-VSD was 39%. Metagenomic sequencing identified five genetically distinct FCV lineages within the three outbreaks, all seemingly evolving in situ in Australia. Notably, no mutations that clearly distinguished FCV-URTD from FCV-VSD phenotypes were identified. One FCV-URTD strain likely originated from a recombination event. Analysis of seven amino-acid residues from the hypervariable E region of the capsid in the cultured viruses did not support the contention that properties of these residues can reliably differentiate between the two pathotypes. On plaque reduction assays, dose-response inhibition of FCV-VSD was obtained with all antivirals at low micromolar concentrations; NTZ EC50, 0.4-0.6 µM, TI = 21; 2CMC EC50, 2.7-5.3 µM, TI > 18; NITD-008, 0.5 to 0.9 µM, TI > 111. Investigation of these antivirals for the treatment of FCV-VSD is warranted.

Cytarabine (Ara-C) is a synthetic isomer of cytidine that differs from cytidine and deoxycytidine only in the sugar. The use of arabinose instead of deoxyribose hinders the formation of phosphodiester linkages between pentoses, preventing the DNA chain from elongation and interrupting the DNA synthesis. The minor structural alteration (the inversion of hydroxyl at the 2′ positions of the sugar) leads to change of the biological activity from anti-depressant and DNA/RNA block builder to powerful anti-cancer. Our study aimed to determine the molecular nature of this phenomenon. Three 1H-14N NMR-NQR experimental techniques, followed by solid-state computational modelling (Quantum Theory of Atoms in Molecules, Reduced Density Gradient and 3D Hirshfeld surfaces), Quantitative Structure–Property Relationships, Spackman’s Hirshfeld surfaces and Molecular Docking were used. Multifaceted analysis—combining experiments, computational modeling and molecular docking—provides deep insight into three-dimensional packing at the atomic and molecular levels, but is challenging. A spectrum with nine lines indicating the existence of three chemically inequivalent nitrogen sites in the Ara-C molecule was recorded, and the lines were assigned to them. The influence of the structural alteration on the NQR parameters was modeled in the solid (GGA/RPBE). For the comprehensive description of the nature of these interactions several factors were considered, including relative reactivity and the involvement of heavy atoms in various non-covalent interactions. The binding modes in the solid state and complex with dCK were investigated using the novel approaches: radial plots, heatmaps and root-mean-square deviation of the binding mode. We identified the intramolecular OH···O hydrogen bond as the key factor responsible for forcing the glycone conformation and strengthening NH···O bonds with Gln97, Asp133 and Ara128, and stacking with Phe137. The titular butterfly effect is associated with both the inversion and the presence of this intramolecular hydrogen bond. Our study elucidates the differences in the binding modes of Ara-C and cytidine, which should guide the design of more potent anti-cancer and anti-viral analogues.

Cytidine and uridine are endogenous metabolites in the pyrimidine metabolism pathway, and cytidine is a substrate that can be metabolized into uridine via cytidine deaminase. Uridine has been widely reported to be effective in regulating lipid metabolism. However, whether cytidine could ameliorate lipid metabolism disorder has not yet been investigated. In this research, ob/ob mice were used, and the effect of cytidine (0.4 mg/mL in drinking water for five weeks) on lipid metabolism disorder was evaluated in terms of an oral glucose tolerance test, serum lipid levels, liver histopathological analysis and gut microbiome analysis. Uridine was used as a positive control. Our findings reveal that cytidine could alleviate certain aspects of dyslipidemia and improve hepatic steatosis via modulating the gut microbiota composition in ob/ob mice, especially increasing the abundance of short-chain fatty acids-producing microbiota. These results suggest that cytidine supplementation could be a potential therapeutic approach for dyslipidemia. [ABSTRACT FROM AUTHOR]

Fluorinated nucleosides are well-established as anticancer and antiviral medications. As with many pharmaceuticals, the effects of fluorine modifications are often only partially understood. In this work, the effects of 5-fluorination of the cytosine nucleobase on the structures and glycosidic bond stabilities of the protonated canonical DNA and RNA cytidine nucleosides (dCyd and Cyd) are examined. Infrared multiple photon dissociation action spectroscopy experiments and electronic structure calculations are employed to probe the structural influences of 5-fluorination. Spectral signatures in the IR fingerprint and hydrogen-stretching regions indicate that 5-fluorination heavily directs for and solely produces O2 protonation para to the 5-fluoro substituent. This differs from the canonical cytidine nucleosides where roughly equal populations of O2 and N3 protonated structures were observed. Energy-resolved collision-induced dissociation experiments combined with survival yield analyses are performed to probe the influence of 5-fluorination on glycosidic bond stability. Trends in the energy-dependence of the survival yield curves indicate that 5-fluorination weakens the glycosidic bond, and that the influence of 5-fluorination on glycosidic bond stability is much greater for dCyd than Cyd. Theory also finds that 5-fluorination weakens the glycosidic bond but predicts that the influence of 5-fluorination on N-glycosidic bond stability is roughly the same for dCyd and Cyd. Oh, what a difference the 2′-hydroxy substituent makes? [Display omitted] • 5-Fluorination favors O2 protonation of DNA and RNA cytidine nucleosides. • 5-Fluorination shuts down N3 protonation of DNA and RNA cytidine nucleosides. • 5-Fluorination weakens glycosidic bonds of DNA and RNA cytidine nucleosides. • 2′-Hydroxy substituent stabilizes glycosidic bonds of cytidine nucleoside analogues. • Influence of 5-fluorination on glycosidic bond stability greater for DNA than RNA? [ABSTRACT FROM AUTHOR]

Cytidine is an antiviral and anticancer drug intermediate, its primary method of manufacture being fermentation. Uridine-cytidine kinase (UCK) catalyzes the reverse process of phosphorylation of cytidine to produce cytidylic acid, which influences cytidine accumulation in the Escherichia coli cytidine biosynthesis pathway. The cytidine-producing strain E. coli NXBG-11 was used as the starting strain in this work; the udk gene coding UCK was knocked out of the chromosomal genome using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology. The mutant strain E. coli NXBG-12 was obtained; its transcriptomics were studied to see how udk gene deletion affected cytidine synthesis and cell-wide transcription. The mutant strain E. coli NXBG-12 generated 1.28 times more cytidine than the original strain E. coli NXBG-11 after 40 h of shake-flask fermentation at 37 °C. The udk gene was knocked out, and transcriptome analysis showed that there were 1168 differentially expressed genes between the mutant and original strains, 559 upregulated genes and 609 downregulated genes. It was primarily shown that udk gene knockout has a positive impact on the cytidine synthesis network because genes involved in cytidine synthesis were significantly upregulated (p < 0.05) and genes related to the cytidine precursor PRPP and cofactor NADPH were upregulated in the PPP and TCA pathways. These results principally demonstrate that udk gene deletion has a favorable impact on the cytidine synthesis network. The continual improvement of cytidine synthesis and metasynthesis is made possible by this information, which is also useful for further converting microorganisms that produce cytidine. [ABSTRACT FROM AUTHOR]

Altogether six tautomers of cytosine and three tautomers of cytidine and deoxycytidine are studied theoretically in the gas phase and in a microhydrated environment. Their structures are optimized at MP2/6–31 + G(d,p) level. The relative energies of the isolated and the hydrated tautomers included the correction to higher correlation energy terms evaluated at the SCS‐MP2, MP4, and CCSD(T) levels. The free energies at 298 K and higher temperatures are based on the above‐mentioned relative energies and temperature‐dependent enthalpy terms and entropies evaluated at the MP2/6–31 + G(d,p) level. Theoretical predictions for the coexistence of five species for cytosine in the gas phase (canonical, trans‐ and cis‐enol‐amino, trans‐ and cis‐imino‐oxo forms) fully agree with recent experimental results. Five‐hydrated cytosine tautomeric forms of are investigated at CPCM/SCS‐MP2/6–31 + G(d,p) level to evaluate the interconversion barriers between them and to explain the coexistence, experimentally proven, of two amino‐oxo and one imino‐oxo tautomers of cytosine in aqueous solution. The presence of coordinated water molecules acting as a catalyst make the tautomerization processes quite easier. It appeared clear from the obtained data that the influence of the hydrogen‐bonded water molecules as well as the introduction of solvent effects in reducing the height of the tautomerization barriers for these five‐hydrated systems is quite substantial. Similar results are obtained for the tri‐hydrated cytidine and the 2‐deoxycytidine tautomeric forms. In aqueous solution of cytidine, syn‐ and syn‐clinal‐conformers оf the amino‐oxo tautomer should coexist with small amounts of syn‐ and syn‐clinal conformers оf imino‐oxo tautomer. For 2‐deoxycytidine an equilibrium has been found to exist between the syn‐ and anti‐conformers of the amino‐oxo and the imino‐oxo tautomers. The water‐assisted proton transfer reactions released through asynchronous concerted mechanism and the conformation of the sugar ring in nucleosides does not change during the tautomerization. [ABSTRACT FROM AUTHOR]

Nucleobase building blocks have been demonstrated to be strong candidates when it comes to DNA/RNA-like materials by benefiting from hydrogen bond interactions as physical properties. Modifying at the 5′ position is the simplest way to develop nucleobase-based structures by transesterification using the lipase Novozym 435. Herein, we describe the optimization of the lipase-catalyzed synthesis of the monomer 5′-O-methacryloylcytidine with the assistance of microwave irradiation. Variable reaction parameters, such as enzyme concentration, molar ratio of the substrate, reaction temperature and reaction time, were investigated to find the optimum reaction condition in terms of obtaining the highest yield. [ABSTRACT FROM AUTHOR]

By the importance of exploring novel compounds for inhibiting the cancerous enzymes activities, this work was performed to recognize advantages of employing 4-amino modified derivatives of cytidine for participating in more efficient interactions with the methyltransferase (MTN) cancerous enzyme target. To this aim, four groups of modified models of cytidine were investigated in addition the original models to recognize the structural features and the corresponding activities. The 4-amino site of cytidine was functionalized by different carbon-based groups in linear and cyclic modes through a bridging peptide linkage. The models were optimized to reach the minimized energy structures by performing quantum chemical calculations and their interactions with the target were analyzed by performing molecular docking simulations. The obtained results of 4-amino modified derivatives of cytidine showed advantages of employing structural modifications to find structures with better molecular orbital based features. Formations of interacting complexes indicated that the additional of carbon-based groups helped to improve possibility of interactions between the substances in both of chemical and physical modes. As a remarkable achievement of this work, the model of cytidine with a phenyl group showed the best advantage of participating in interactions with the MTN target among all twenty five models of the investigated cytidine compounds. [ABSTRACT FROM AUTHOR]

We report the synthesis and characterization of three di-copper compounds, [Cu2phen2CMP(µ-OH)(µ-X)]·3H2O (with X = Cl− (1) and Br− (2)) and [Cu2phen2CMP(µ-OH)(µ-NO3)]·[Cu2phen2CMP(µ-OH)(NO3)]·7H2O (3). 1 and 3 present a dinuclear core assembled by an unusual bis-monodentate phosphate and µ2-hydroxo double bridge, further supported by bridging anions. The spontaneous self-assembly of an extended coordination structure of higher dimensionality, although possible in principle, has been hindered by the use of strongly coordinating counterions (Cl−, Br−) and, unexpectedly, even by the less coordinating nitrate ions. The neutral complexes feature unsaturated coordination sites, having both the N(3) and O(2) of the cytosine moieties of the CMP ligands still available for coordination. π···π Stacking and hydrogen bonding interactions established between the discrete chiral entities mainly drive the self-assembling of a 3D supramolecular network. Small hydrophilic voids are defined within the 3D net, which are filled by solvent water molecules. These small nuclearity complexes with coordinatively unsaturated CMP ligands are likely to act as tectons. They could be used as complex-as-ligands for further binding additional metal ions in a predictable manner, favoring a more rational synthetic approach aimed to a stricter structural and chemical control of the resulting material. [ABSTRACT FROM AUTHOR]

UnlabelledWe tested pairwise combinations of classical base analog mutagens in Escherichia coli to study possible mutagen synergies. We examined the cytidine analogs zebularine (ZEB) and 5-azacytidine (5AZ), the adenine analog 2-aminopurine (2AP), and the uridine/thymidine analog 5-bromodeoxyuridine (5BrdU). We detected a striking synergy with the 2AP plus ZEB combination, resulting in hypermutability, a 35-fold increase in mutation frequency (to 53,000 × 10(-8)) in the rpoB gene over that with either mutagen alone. A weak synergy was also detected with 2AP plus 5AZ and with 5BrdU plus ZEB. The pairing of 2AP and 5BrdU resulted in suppression, lowering the mutation frequency of 5BrdU alone by 6.5-fold. Sequencing the mutations from the 2AP plus ZEB combination showed the predominance of two new hot spots for A·T→G·C transitions that are not well represented in either single mutagen spectrum, and one of which is not found even in the spectrum of a mismatch repair-deficient strain. The strong synergy between 2AP and ZEB could be explained by changes in the dinucleoside triphosphate (dNTP) pools.ImportanceAlthough mutagens have been widely studied, the mutagenic effects of combinations of mutagens have not been fully researched. Here, we show that certain pairwise combinations of base analog mutagens display synergy or suppression. In particular, the combination of 2-aminopurine and zebularine, analogs of adenine and cytidine, respectively, shows a 35-fold increased mutation frequency compared with that of either mutagen alone. Understanding the mechanism of synergy can lead to increased understanding of mutagenic processes. As combinations of base analogs are used in certain chemotherapy regimens, including those involving ZEB and 5AZ, these results indicate that testing the mutagenicity of all drug combinations is prudent.

Nucleodyes, visibly colored chromophoric nucleoside analogues, are reported. Design criteria are outlined and the syntheses of cytidine and uridine azo dye analogues derived from 6-aminouracil are described. Structural analysis shows that the nucleodyes are sound structural analogues of their native nucleoside counterparts, and photophysical studies demonstrate that the nucleodyes are sensitive to microenvironmental changes. Quantum chemical calculations are presented as a valuable complementary tool for the design of strongly absorbing nucleodyes, which overlap with the emission of known fluorophores. Förster critical distance (R0) calculations determine that the nucleodyes make good FRET pairs with both 2-aminopurine (2AP) and pyrrolocytosine (PyC). Additionally, unique tautomerization features exhibited by 5-(4-nitrophenylazo)-6-oxocytidine (8) are visualized by an extraordinary crystal structure.

Current genome-editing technologies introduce double-stranded (ds) DNA breaks at a target locus as the first step to gene correction. Although most genetic diseases arise from point mutations, current approaches to point mutation correction are inefficient and typically induce an abundance of random insertions and deletions (indels) at the target locus resulting from the cellular response to dsDNA breaks. Here we report the development of 'base editing', a new approach to genome editing that enables the direct, irreversible conversion of one target DNA base into another in a programmable manner, without requiring dsDNA backbone cleavage or a donor template. We engineered fusions of CRISPR/Cas9 and a cytidine deaminase enzyme that retain the ability to be programmed with a guide RNA, do not induce dsDNA breaks, and mediate the direct conversion of cytidine to uridine, thereby effecting a C→T (or G→A) substitution. The resulting 'base editors' convert cytidines within a window of approximately five nucleotides, and can efficiently correct a variety of point mutations relevant to human disease. In four transformed human and murine cell lines, second- and third-generation base editors that fuse uracil glycosylase inhibitor, and that use a Cas9 nickase targeting the non-edited strand, manipulate the cellular DNA repair response to favour desired base-editing outcomes, resulting in permanent correction of ~15-75% of total cellular DNA with minimal (typically ≤1%) indel formation. Base editing expands the scope and efficiency of genome editing of point mutations.

Objective To evaluate the pancreatic differentiation potential of α-1,3-galactosyltransferase knockout (GalTKO) pig-derived bone marrow-derived mesenchymal stem cells (BM-MSCs) using epigenetic modifiers with different pancreatic induction media. Methods The BM-MSCs have been differentiated into pancreatic β-like cells by inducing the overexpression of key transcription regulatory factors or by exposure to specific soluble inducers/small molecules. In this study, we evaluated the pancreatic differentiation of GalTKO pig-derived BM-MSCs using epigenetic modifiers, 5-azacytidine (5-Aza) and valproic acid (VPA), and two types of pancreatic induction media – advanced Dulbecco’s modified Eagle’s medium (ADMEM)-based and N2B27-based media. GalTKO BM-MSCs were treated with pancreatic induction media and the expression of pancreas-islets-specific markers was evaluated by real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. Morphological changes and changes in the 5′-C-phosphate-G-3′ (CpG) island methylation patterns were also evaluated. Results The expression of the pluripotent marker (POU class 5 homeobox 1 [OCT4]) was upregulated upon exposure to 5-Aza and/or VPA. GalTKO BM-MSCs showed increased expression of neurogenic differentiation 1 in the ADMEM-based (5-Aza) media, while the expression of NK6 homeobox 1 was elevated in cells induced with the N2B27-based (5-Aza) media. Moreover, the morphological transition and formation of islets-like cellular clusters were also prominent in the cells induced with the N2B27-based media with 5-Aza. The higher insulin expression revealed the augmented trans-differentiation ability of GalTKO BM-MSCs into pancreatic β-like cells in the N2B27-based media than in the ADMEM-based media. Conclusion 5-Aza treated GalTKO BM-MSCs showed an enhanced demethylation pattern in the second CpG island of the OCT4 promoter region compared to that in the GalTKO BM-MSCs. The exposure of GalTKO pig-derived BM-MSCs to the N2B27-based microenvironment can significantly enhance their trans-differentiation ability into pancreatic β-like cells.

Marco Antonio Naslausky Mibielli,1 Carlos Pereira Nunes,1,2 Henrique Goldberg,3 Luiz Buchman,2 Lisa Oliveira,4 Spyros GE Mezitis,5 Fernanda Wajnzstajn,6 Renato Kaufman,3 Rafael Nigri,7 Natasha Cytrynbaum,3 Karin Soares Cunha,8 Alessandra Santos,4 Stephanie Wrobel Goldberg,9 Natália Carvalho Platenik,1 Helio Rzetelna,10 Daniel Bertoluci Futuro,1 Adenilson de Souza Da Fonseca,1 Mauro Geller1,3,4 1UNIFESO Medical School, Teresópolis, Brazil; 2UERJ Medical School, Rio De Janeiro, Brazil; 3Instituto De Pós-Graduação Médica Carlos Chagas (ICC), Rio De Janeiro, Brazil; 4Federal University of Rio De Janeiro (UFRJ), Rio De Janeiro, Brazil; 5New York-Presbyterian Hospital/Weill-Cornell Medical Center, New York, NY, USA; 6Neurology Department, UConn Health, Farmington, CT, USA; 7Department of Medicine, Rutgers New Jersey Medical School, Newark, NJ, USA; 8Pathology Department, Universidade Federal Fluminense (UFF) Medical School, Niterói, Brazil; 9Tufts Medical Center, Boston, MA, USA; 10Santa Casa Da Misericórdia Do Rio De Janeiro, Rio De Janeiro, BrazilCorrespondence: Mauro Geller Av. Ataulfo de Paiva, 135/702 Rio de Janeiro, 22440-901, BrazilTel +55-21-3875-6660Email maurogeller@gmail.comPurpose: We report the results of low back pain treatment using a combination of nucleotides, uridine (UTP), cytidine (CMP) and vitamin B12, vs a combination of vitamins B1, B6, and B12.Patients and Methods: Randomized, double-blind, controlled trial, of a 60-day oral treatment: Group A (n=317) receiving nucleotides+B12 and Group B (n=317) receiving B vitamins. The primary endpoint was the percentage of subjects in each group presenting adverse events (AEs). Secondary endpoints were visual analog scale (VAS) pain scores at Visit 2 (day 30) and Visit 3 (day 60) in relation to pretreatment values, Roland–Morris Questionnaire (RMQ) scores and finger-to-floor distance (FFD) (percentage of subjects per group presenting improvement ≥ 5 points and ≥ 3cm, respectively).Results: Seventy-five (24%) and 105 (33%) subjects (P=0.21) presented 133 and 241 AEs, with 3159% of subjects presenting ≥ 2 AEs (P=0.0019) in Group A and Group B, respectively. Twenty-four subjects in Group B were discontinued due to AEs, while no AE-related discontinuations occurred in Group A (P< 0.0001). VAS score reduction after 30 and 60 days of treatment was statistically significant (P< 0.0001) in both groups, with Group A showing greater reduction at Visit 2 (P< 0.0001). RMQ score improvement ≥ 5 points occurred in 99% of subjects from each group, and FFD improvement ≥ 3 cm occurred in all subjects.Conclusion: Treatment with nucleotides+B12 was associated with a lower number of total AEs, fewer AEs per subject, and no AE-related treatment discontinuation. Pain intensity (VAS) reduction was superior at 30 days of treatment in the nucleotides+B12 group and equivalent between groups at 60 days of treatment. Improvements in efficacy measures RMQ and FFD were observed in both groups at treatment days 30 and 60.Keywords: low back pain, uridine, cytidine, vitamin B1, vitamin B6, vitamin B12

Orally delivered drugs offer significant benefits in the fight against viral infections, and cost-effective production is critical to their impact on pandemic response in low- and middle-income countries. One example, molnupiravir, a COVID-19 therapy developed by Emory, Ridgeback, and Merck & Co., had potential to benefit from significant cost of goods (COGs) reductions for its active pharmaceutical ingredient (API), including starting materials. A holistic approach to identifying, developing, and evaluating optimized synthetic routes, which includes detailed COGs modeling, provides a rapid means to increase the availability, uptake and application of molnupiravir and other antivirals in global markets. Identification and development of alternate processes for the synthesis of molnupiravir has been conducted by the Medicines for All Institute at Virginia Commonwealth University (M4ALL) and the Green and Turner Labs at the University of Manchester. Both groups developed innovative processes based on synthetic route design and biocatalysis aimed at lowering costs and improving global access. The authors then performed COGs modeling to assess cost saving opportunities. This included a focus on manufacturing environments and facilities amenable to global public health and the identification of key parameters using sensitivity analyses. While all of the evaluated routes provide efficiency benefits, the best options yielded 3-6 fold API COGs reductions leading to treatment COGs as low as

Cytidine is an antiviral and anticancer drug intermediate, its primary method of manufacture being fermentation. Uridine-cytidine kinase (UCK) catalyzes the reverse process of phosphorylation of cytidine to produce cytidylic acid, which influences cytidine accumulation in the Escherichia coli cytidine biosynthesis pathway. The cytidine-producing strain E. coli NXBG-11 was used as the starting strain in this work; the udk gene coding UCK was knocked out of the chromosomal genome using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology. The mutant strain E. coli NXBG-12 was obtained; its transcriptomics were studied to see how udk gene deletion affected cytidine synthesis and cell-wide transcription. The mutant strain E. coli NXBG-12 generated 1.28 times more cytidine than the original strain E. coli NXBG-11 after 40 h of shake-flask fermentation at 37 °C. The udk gene was knocked out, and transcriptome analysis showed that there were 1168 differentially expressed genes between the mutant and original strains, 559 upregulated genes and 609 downregulated genes. It was primarily shown that udk gene knockout has a positive impact on the cytidine synthesis network because genes involved in cytidine synthesis were significantly upregulated (p < 0.05) and genes related to the cytidine precursor PRPP and cofactor NADPH were upregulated in the PPP and TCA pathways. These results principally demonstrate that udk gene deletion has a favorable impact on the cytidine synthesis network. The continual improvement of cytidine synthesis and metasynthesis is made possible by this information, which is also useful for further converting microorganisms that produce cytidine.

Nucleoside analogs contribute in pharmaceutical and clinical fields as medicinal agents and approved drugs. This work focused to investigate the antimicrobial, anticancer activities, and structure–activity relationship (SAR) of cytidine and its analogs with computational studies. Microdilution was used to determine the antimicrobial activity, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) of the modified analogs against human and phytopathogenic strains. Compounds (7), (10), and (14) were the most potent against Escherichia coli and Salmonella abony strains with MIC and MBC values from 0.316 ± 0.02 to 2.50 ± 0.03 and 0.625 ± 0.04 to 5.01 ± 0.06 mg/ml, respectively. The highest inhibitory activity was observed against gram-positive bacteria. Numerous analogs (10), (13), (14), and (15) exhibited good activity against the tested fungi Aspergillus niger and Aspergillus flavus. Anticancer activity of the cytidine analogs was examined through MTT colorimetric assay against Ehrlich's ascites carcinoma (EAC) tumor cells whereas compound 6 showed the maximum antiproliferative activity with an IC50 value of 1168.97 µg/ml. To rationalize this observation, their quantum mechanical and molecular docking studies have been performed against urate oxidase of A. flavus 1R51 to investigate the binding mode, binding affinity, and non-bonding interactions. It was observed that most of the analogs exhibited better binding properties than the parent drug. In silico ADMET prediction was attained to evaluate the drug-likeness properties that revealed the improved pharmacokinetic profile with lower acute oral toxicity of cytidine analogs. Based on the in vitro and in silico analysis, this exploration can be useful to develop promising cytidine-based antimicrobial drug(s). [ABSTRACT FROM AUTHOR]

Cytidine is an important raw material for nucleic acid health food and genetic engineering research. In recent years, it has shown irreplaceable effects in anti-virus, anti-tumor, and AIDS drugs. Its biosynthetic pathway is complex and highly regulated. In this study, overexpression of uracil permease and a nucleoside transporter from Bacillus amyloliquefaciens related to cell membrane transport in Escherichia coli strain BG-08 was found to increase cytidine production in shake flask cultivation by 1.3-fold (0.91 ± 0.03 g/L) and 1.8-fold (1.26 ± 0.03 g/L) relative to that of the original strain (0.70 ± 0.03 g/L), respectively. Co-overexpression of uracil permease and a nucleoside transporter further increased cytidine yield by 2.7-fold (1.59 ± 0.05 g/L) compared with that of the original strain. These results indicate that the overexpressed uracil permease and nucleoside transporter can promote the accumulation of cytidine, and the two proteins play a synergistic role in the secretion of cytidine in Escherichia coli. [ABSTRACT FROM AUTHOR]

Nucleobase building blocks have been demonstrated to be strong candidates when it comes to DNA/RNA-like materials by benefiting from hydrogen bond interactions as physical properties. Modifying at the 5′ position is the simplest way to develop nucleobase-based structures by transesterification using the lipase Novozym 435. Herein, we describe the optimization of the lipase-catalyzed synthesis of the monomer 5′-O-methacryloylcytidine with the assistance of microwave irradiation. Variable reaction parameters, such as enzyme concentration, molar ratio of the substrate, reaction temperature and reaction time, were investigated to find the optimum reaction condition in terms of obtaining the highest yield.

We have previously shown that liquid sample desorption electrospray ionization-mass spectrometry (DESI-MS) is able to measure large proteins and noncovalently bound protein complexes (to 150 kDa) (Ferguson et al., Anal. Chem. 2011, 83, 6468-6473). In this study, we further investigate the application of liquid sample DESI-MS to probe protein-ligand interactions. Liquid sample DESI allows the direct formation of intact protein-ligand complex ions by spraying ligands toward separate protein sample solutions. This type of "reactive" DESI methodology can provide rapid information on binding stiochiometry, selectivity, and kinetics, as demonstrated by the binding of ribonuclease A (RNaseA, 13.7 kDa) with cytidine nucleotide ligands and the binding of lysozyme (14.3 kDa) with acetyl chitose ligands. A higher throughput method for ligand screening by liquid sample DESI was demonstrated, in which different ligands were sequentially injected as a segmented flow for DESI ionization. Furthermore, supercharging to enhance analyte charge can be integrated with liquid sample DESI-MS, without interfering with the formation of protein-ligand complexes.

Magnetic molecularly imprinted nanoparticles were fabricated for cytidine adsorption. Molecularly imprinted polymers were prepared by complexing N‐methacryloyl‐l‐histidine/Cu(II) and cytidine as a template molecule in a highly cross‐linked polymer. Magnetic molecularly imprinted nanoparticles were tested in batch experiments with aqueous samples of cytidine, and several parameters were investigated. The N‐methacryloyl‐l‐histidine/Cu(II) complex was selected to obtain a rebinding of cytidine on the magnetic nanoparticles. The template molecule was removed using 1.0 M NaCl as a desorption agent. The adsorption of cytidine was optimal in a buffer solution of pH 7.4 for 120 min with adsorption capacity of 39.77 mg/g. The imprinted polymer‐based magnetic nanoparticles were selective toward cytidine as compared to the analogs (uridine and thymidine). The adsorption fitted a Langmuir isotherm model with an R2 value of 0.9861 and the Qmax value of 43.86 mg/g for the magnetic molecularly imprinted nanoparticles. [ABSTRACT FROM AUTHOR]

Cytidine acetylation (ac4C) of RNA is a post-transcriptional modification catalyzed by Nat10. Recently, an approach termed RedaC:T was employed to map ac4C in human mRNA, relying on detection of C>T mutations in WT but not in Nat10-KO cells. RedaC:T suggested widespread ac4C presence. Here, we reanalyze RedaC:T data. We find that mismatch signatures are not reproducible, as C>T mismatches are nearly exclusively present in only one of two biological replicates. Furthermore, all mismatch types-not only C>T-are highly enriched in WT samples, inconsistent with an acetylation signature. We demonstrate that the originally observed enrichment in mutations in one of the WT samples is due to its low complexity, resulting in the technical amplification of all classes of mismatch counts. Removal of duplicate reads abolishes the skewed mismatch patterns. These analyses account for the irreproducible mismatch patterns across samples while failing to find evidence for acetylation of RedaC:T sites., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Cytidine is a well-known inhibitor of DNA methyltransferase (MTN) enzyme for preventing cancer cells growth. Based on therapeutic benefits, it could be considered as a "lead compound" to be optimized through structural modification for arising better binding affinity in this case. Halogenated derivatives of cytidine were investigated in this work to examine structural and biological features employing in silico approach. To this aim, geometries of the original cytidine and four of its halogenated derivatives were minimized to prepare ligands for interacting with MTN enzyme target in molecular docking simulations. The results for singular ligand structures introduced I-cytidine as an optimized lead compound for contributing to proper interactions with MTN enzyme; the trend was confirmed by molecular docking simulations. As a final remark, I-cytidine could be considered as better ligand for complexation with the MTN enzyme in comparison with the original cytidine. Halogenated derivatives of cytidine were investigated. Binding affinity of derivatives against DNA methyltransferase were investigated. The obtained in silico results indicated helpful information of ligand...target interactions. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the pancreatic differentiation potential of α-1,3-galactosyltransferase knockout (GalTKO) pig-derived bone marrow-derived mesenchymal stem cells (BM-MSCs) using epigenetic modifiers with different pancreatic induction media. Methods: The BM-MSCs have been differentiated into pancreatic β-like cells by inducing the overexpression of key transcription regulatory factors or by exposure to specific soluble inducers/small molecules. In this study, we evaluated the pancreatic differentiation of GalTKO pig-derived BM-MSCs using epigenetic modifiers, 5-azacytidine (5-Aza) and valproic acid (VPA), and two types of pancreatic induction media - advanced Dulbecco's modified Eagle's medium (ADMEM)-based and N2B27-based media. GalTKO BM-MSCs were treated with pancreatic induction media and the expression of pancreas-islets-specific markers was evaluated by real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. Morphological changes and changes in the 5'-C-phosphate-G-3' (CpG) island methylation patterns were also evaluated. Results: The expression of the pluripotent marker (POU class 5 homeobox 1 [OCT4]) was upregulated upon exposure to 5-Aza and/or VPA. GalTKO BM-MSCs showed increased expression of neurogenic differentiation 1 in the ADMEM-based (5-Aza) media, while the expression of NK6 homeobox 1 was elevated in cells induced with the N2B27-based (5-Aza) media. Moreover, the morphological transition and formation of islets-like cellular clusters were also prominent in the cells induced with the N2B27-based media with 5-Aza. The higher insulin expression revealed the augmented trans-differentiation ability of GalTKO BM-MSCs into pancreatic β-like cells in the N2B27-based media than in the ADMEM-based media. Conclusion: 5-Aza treated GalTKO BM-MSCs showed an enhanced demethylation pattern in the second CpG island of the OCT4 promoter region compared to that in the GalTKO BM-MSCs. The exposure of GalTKO pig-derived BM-MSCs to the N2B27-based microenvironment can significantly enhance their trans-differentiation ability into pancreatic β-like cells. [ABSTRACT FROM AUTHOR]

Trypanosomiasis is a major illness affecting camels in tropical and subtropical regions. Comparisons of camel and Trypanosoma evansi genomes can lead to the discovery of new drug targets for treating Trypanosoma infections. The synthesis pathways of cytosine, cytidine, cytidine monophosphate (CMP), cytidine diphosphate (CDP), cytidine triphosphate (CTP) deoxycytidine, deoxycytidine monophosphate (dCMP), deoxycytidine diphosphate (dCDP), and deoxycytidine triphosphate (dCTP) were compared in the dromedary camel (Camelus dromedarius) and T. evansi. None of the enzymes involved in cytosine pathway were detected in camels and T. evansi. Notably, cytidine kinase (CK) and 5′-nucleotidase, which interconverts cytidine to CMP, were not detected in T. evansi but were present in camels. UMP/CMP kinase was not predicted in T. evansi. Therefore, the presence of enzymes involved in the CTP synthesis cascade was not predicted in T. evansi. CMP synthesis might also be encoded by other enzymes, e.g., purine nucleotides kinases. Both camel and T. evansi share an efficient enzyme system for converting CDP to CTP. In conclusion, CTP synthase is important for homeostasis of cytosine nucleotides in T. evansi and could be a potential drug target against the parasite. In addition, the inhibition of UMP synthesis might contribute to parasite death as it is a shared source for CTP synthesis. [ABSTRACT FROM AUTHOR]

Summary: Targeted mutagenesis via genome‐editing technologies holds great promise in developing improved crop varieties to meet future demands. Point mutations or single nucleotide polymorphisms often determine important agronomic traits of crops. Genome‐editing‐based single‐base changes could generate elite trait variants in crop plants which help in accelerating crop improvement. Among the genome‐editing technologies, base editing has emerged as a novel and efficient genome‐editing approach which enables direct and irreversible conversion of one target base into another in a programmable manner. A base editor is a fusion of catalytically inactive CRISPR–Cas9 domain (Cas9 variants) and cytosine or adenosine deaminase domain that introduces desired point mutations in the target region enabling precise editing of genomes. In the present review, we have summarized the development of different base‐editing platforms. Then, we have focussed on the current advances and the potential applications of this precise technology in crop improvement. The review also sheds light on the limitations associated with this technology. Finally, the future perspectives of this emerging technology towards crop improvement have been highlighted. [ABSTRACT FROM AUTHOR]