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SUMMARY: Among the three active aldehyde oxidases in Arabidopsis thaliana leaves (AAO1‐3), AAO3, which catalyzes the oxidation of abscisic‐aldehyde to abscisic‐acid, was shown recently to function as a reactive aldehyde detoxifier. Notably, aao2KO mutants exhibited less senescence symptoms and lower aldehyde accumulation, such as acrolein, benzaldehyde, and 4‐hydroxyl‐2‐nonenal (HNE) than in wild‐type leaves exposed to UV‐C or Rose‐Bengal. The effect of AAO2 expression absence on aldehyde detoxification by AAO3 and/or AAO1 was studied by comparing the response of wild‐type plants to the response of single‐functioning aao1 mutant (aao1S), aao2KO mutants, and single‐functioning aao3 mutants (aao3Ss). Notably, aao3Ss exhibited similar aldehyde accumulation and chlorophyll content to aao2KO treated with UV‐C or Rose‐Bengal. In contrast, wild‐type and aao1S exhibited higher aldehyde accumulation that resulted in lower remaining chlorophyll than in aao2KO leaves, indicating that the absence of active AAO2 enhanced AAO3 detoxification activity in aao2KO mutants. In support of this notion, employing abscisic‐aldehyde as a specific substrate marker for AAO3 activity revealed enhanced AAO3 activity in aao2KO and aao3Ss leaves compared to wild‐type treated with UV‐C or Rose‐Bengal. The similar abscisic‐acid level accumulated in leaves of unstressed or stressed genotypes indicates that aldehyde detoxification by AAO3 is the cause for better stress resistance in aao2KO mutants. Employing the sulfuration process (known to activate aldehyde oxidases) in wild‐type, aao2KO, and molybdenum‐cofactor sulfurase (aba3‐1) mutant plants revealed that the active AAO2 in WT employs sulfuration processes essential for AAO3 activity level, resulting in the lower AAO3 activity in WT than AAO3 activity in aao2KO. Significance Statement: The current study demonstrates that impairment of AAO2 causes enhancement of AAO3 activity and carbonyl aldehyde detoxification increase in Arabidopsis. [ABSTRACT FROM AUTHOR]

Unexpected metabolism could lead to the failure of many late-stage drug candidates or even the withdrawal of approved drugs. Thus, it is critical to predict and study the dominant routes of metabolism in the early stages of research. We describe the development and validation of a 'WhichEnzyme' model that accurately predicts the enzyme families most likely to be responsible for a drug-like molecule's metabolism. Furthermore, we combine this model with our previously published regioselectivity models for Cytochromes P450, Aldehyde Oxidases, Flavin-containing Monooxygenases, UDP-glucuronosyltransferases and Sulfotransferases – the most important Phase I and Phase II drug metabolising enzymes – and a 'WhichP450' model that predicts the Cytochrome P450 isoform(s) responsible for a compound's metabolism. The regioselectivity models are based on a mechanistic understanding of these enzymes' actions and use quantum mechanical simulations with machine learning methods to accurately predict sites of metabolism and the resulting metabolites. We train heuristics based on the outputs of the 'WhichEnzyme', 'WhichP450', and regioselectivity models to determine the most likely routes of metabolism and metabolites to be observed experimentally. Finally, we demonstrate that this combination delivers high sensitivity in identifying experimentally reported metabolites and higher precision than other methods for predicting in vivo metabolite profiles. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) plays a crucial role in plant defense mechanisms under adverse environmental conditions, but its metabolism and perception in response to heavy metals are largely unknown. In Pisum sativum exposed to CdCl2, an accumulation of free ABA was detected in leaves at different developmental stages (A, youngest, unexpanded; B1, youngest, fully expanded; B2, mature; C, old), with the highest content found in A and B1 leaves. In turn, the content of ABA conjugates, which was highest in B2 and C leaves under control conditions, increased only in A leaves and decreased in leaves of later developmental stages after Cd treatment. Based on the expression of PsNCED2, PsNCED3 (9-cis-epoxycarotenoid dioxygenase), PsAO3 (aldehyde oxidase) and PsABAUGT1 (ABA-UDP-glucosyltransferase), and the activity of PsAOγ, B2 and C leaves were found to be the main sites of Cd-induced de novo synthesis of ABA from carotenoids and ABA conjugation with glucose. In turn, β-glucosidase activity and the expression of genes encoding ABA receptors (PsPYL2, PsPYL4, PsPYL8, PsPYL9) suggest that in A and B1 leaves, Cd-induced release of ABA from inactive ABA-glucosyl esters and enhanced ABA perception comes to the forefront when dealing with Cd toxicity. The distinct role of leaves at different developmental stages in defense against the harmful effects of Cd is discussed. [ABSTRACT FROM AUTHOR]

Selective chemical inhibitors are critical for reaction phenotyping to identify drug-metabolizing enzymes that are involved in the elimination of drug candidates. Although relatively selective inhibitors are available for the major cytochrome P450 enzymes (CYP), they are quite limited for the less common CYPs and non-CYPs. To address this gap, we developed a multiplexed high throughput screening (HTS) assay using 20 substrate reactions of multiple enzymes to simultaneously monitor the inhibition of enzymes in a 384-well format. Four 384-well assay plates can be run at the same time to maximize throughput. This is the first multiplexed HTS assay for drug-metabolizing enzymes reported. The HTS assay is technologically enabled with state-of-the-art robotic systems and highly sensitive modern LC-MS/MS instrumentation. Virtual screening is utilized to identify inhibitors for HTS based on known inhibitors and enzyme structures. Screening of ~4600 compounds generated many hits for many drug-metabolizing enzymes including the two time-dependent and selective aldehyde oxidase inhibitors, erlotinib and dibenzothiophene. The hit rate is much higher than that for the traditional HTS for biological targets due to the promiscuous nature of the drug-metabolizing enzymes and the biased compound selection process. Future efforts will focus on using this method to identify selective inhibitors for enzymes that do not currently have quality hits and thoroughly characterizing the newly identified selective inhibitors from our screen. We encourage colleagues from other organizations to explore their proprietary libraries using a similar approach to identify better inhibitors that can be used across the industry. [ABSTRACT FROM AUTHOR]

Chiral pollutants often pose significant differential environmental health risks. In this study, the biotransformation of chiral dinotefuran (DIN) and its enantioselective metabolic toxicity mechanisms have been systemically investigated. Firstly, reversed-phase chromatography-high resolution mass spectrometry was developed to quantify the content of DIN R/S chiral enantiomer with pg level sensitivity, revealing a lower elimination rate constant (Ke) of S-DIN (0.730 h−1) than R-DIN (0.746 h−1). Secondly, the interaction mechanism between DIN metabolism and important endogenous bioactive molecules, such as aldehyde oxidase (AOX) and neurotransmitters, was revealed. The DIN nitro-group was converted into a guanidine group by the reducing site of nearby flavin adenine dinucleotide (FAD) in AOX with the preferred higher affinity of S-configuration. Meanwhile, the endogenous tryptophan (Trp) aldehyde metabolic intermediate, 5-hydroxyindoleacetaldehyde (5-HIAL), provides a persistent electron donor for DIN reduction via the oxidation-catalyzed site in AOX, resulting in remarkable up-regulation of monoamine neurotransmitters such as serotonin and dopamine. Thirdly, the higher level of neurotransmitters further mediated dysregulation of oxylipin homeostasis via the serotonergic pathway, where S-DIN exhibited more pronounced liver lipid damage and environmental health risk with the accumulated lipid biomarkers, oxidized triglyceride (OxTG) and oxidized sphingomyelin (OxSM). This study elucidates the AOX-mediated enantioselectivity metabolic pathway of DIN, providing a new analytical method for chiral pollutants and paves the way for their health risk assessments. [ABSTRACT FROM AUTHOR]

Aldehyde oxidase (AOX) is a molybdoenzyme that is primarily expressed in the liver and is involved in the metabolism of drugs and other xenobiotics. AOX-mediated metabolism can result in unexpected outcomes, such as the production of toxic metabolites and high metabolic clearance, which can lead to the clinical failure of novel therapeutic agents. Computational models can assist medicinal chemists in rapidly evaluating the AOX metabolic risk of compounds during the early phases of drug discovery and provide valuable clues for manipulating AOX-mediated metabolism liability. In this study, we developed a novel graph neural network called AOMP for predicting AOX-mediated metabolism. AOMP integrated the tasks of metabolic substrate/non-substrate classification and metabolic site prediction, while utilizing transfer learning from 13C nuclear magnetic resonance data to enhance its performance on both tasks. AOMP significantly outperformed the benchmark methods in both cross-validation and external testing. Using AOMP, we systematically assessed the AOX-mediated metabolism of common fragments in kinase inhibitors and successfully identified four new scaffolds with AOX metabolism liability, which were validated through in vitro experiments. Furthermore, for the convenience of the community, we established the first online service for AOX metabolism prediction based on AOMP, which is freely available at https://aomp.alphama.com.cn.

Organic solvents are extensively used in in vitro drug metabolism studies to overcome the solubility issues of lipophilic substrate/new chemical entities. The effects of ten common water miscible organic solvents on cytochrome P450s and non-cytochrome P450 enzymes, namely, flavin monooxygense, aldehyde oxidase, xanthine oxidase, esterases and glutathione S-transferases were evaluated. Rat liver microsomes and rat liver cytosol were used for cytochrome P450, flavin monooxygenase and glutathione S-transferase assays. Partially purified guinea pig liver aldehyde oxidase and rat liver xanthine oxidase fractions were used for molybdenum hydroxylase activity assays. Human plasma was used for esterase activity. Para-nitro phenol, metoprolol, imipramine, methyl para-tolyl sulfide, para-nitrophenyl acetate, vanillin, xanthine and chloro dinitrobenzene, were used as substrates for evaluating CYP2E1, CYP2D6, CYP1A2, flavin monooxyenase, esterase, aldehyde oxidase, xanthine oxidase and glutathione S-transferase activities, respectively. Ten solvents (acetonitrile, acetone, dioxane, dimethyl formamide, dimethyl sulfoxide, ethanol, methanol, polyethylene glycol 400, n-propanol, and isopropanol) were evaluated at four concentrations (0.1, 0.25, 0.5 and 1 % v/v). The following results were obtained. Dioxane (1 % v/v) was found to a potent inhibitor inhibiting all the tested CYP's (>90 % inhibition) and flavin monooxygenase (71 % inhibition). Seven of ten organic solvents inhibited esterases in a concentration dependent manner out of which n-propanol showed the maximal inhibition (79 % inhibition). Methanol was the only solvent that inhibited aldehyde oxidase (78 % inhibition) and xanthine oxidase mediated metabolism (64 % inhibition). Minimal inhibition effect was observed on glutathione S-transferase mediated activity by all solvents. Polyethylene glycol 400 showed least inhibition of enzyme activities. Among the 10 solvents studies, acetonitrile overall appears to be the safest organic solvent at concentrations of 1 % v/v or less. The results of this study indicate that many water miscible organic solvents must be used very judiciously in drug metabolism studies. [ABSTRACT FROM AUTHOR]

Aldehyde oxidase (AOX) is a molybdoenzyme that is primarily expressed in the liver and is involved in the metabolism of drugs and other xenobiotics. AOX-mediated metabolism can result in unexpected outcomes, such as the production of toxic metabolites and high metabolic clearance, which can lead to the clinical failure of novel therapeutic agents. Computational models can assist medicinal chemists in rapidly evaluating the AOX metabolic risk of compounds during the early phases of drug discovery and provide valuable clues for manipulating AOX-mediated metabolism liability. In this study, we developed a novel graph neural network called AOMP for predicting AOX-mediated metabolism. AOMP integrated the tasks of metabolic substrate/non-substrate classification and metabolic site prediction, while utilizing transfer learning from 13C nuclear magnetic resonance data to enhance its performance on both tasks. AOMP significantly outperformed the benchmark methods in both cross-validation and external testing. Using AOMP, we systematically assessed the AOX-mediated metabolism of common fragments in kinase inhibitors and successfully identified four new scaffolds with AOX metabolism liability, which were validated through in vitro experiments. Furthermore, for the convenience of the community, we established the first online service for AOX metabolism prediction based on AOMP, which is freely available at https://aomp.alphama.com.cn. A transfer learning enhanced graph neural network was established to predict the AOX-mediated metabolism. The proposed model achieved state-of-the-art performance and was applied to identify new scaffolds with AOX metabolism liability. [Display omitted] [ABSTRACT FROM AUTHOR]

Crop production is the basis of agriculture in Kazakhstan. Soils in Kazakhstan suffer from molybdenum deficiency. Molybdenum deficiency in plants is characterized by the stunted growth, poor seed setting, fruit deformation and reduced plant resistance to diseases. Molybdenum catalyzes key steps in the metabolism of nitrogen, carbon and sulfur. It promotes the biosynthesis of the phytohormones of abscisic acid and indole-3-butyric acid. An increase in molybdenum concentration leads to an increase in aldehyde oxidase activity. Aldehyde oxidase generates hydrogen peroxide during viral infection; hydrogen peroxide triggers a hypersensitive response, strengthens cell walls and prevents the spread of viral infection. Molybdenum is biologically inactive until a special complex called molybdenum cofactor is formed. For the formation of the latter it is necessary for molybdenum to be in the state of molybdate anion. There are five known plant molybdoenzymes: aldehyde oxidase, sulfite oxidase, nitrate reductase, xanthine dehydrogenase and mitochondrial amidoxime resuctase. The aim of this work is to find the optimal concentration of molybdenum for aldehyde oxidase activation. The objectives of the conducted experiment are: - to apply an efficient and cost-effective method to replenish molybdenum deficiency in the experimental plant; - to demonstrate the effect of molybdenum on the development of virus infection of the infected plant. As a result, it was proved that with the increase of concentration from 0.1 mM to 1 mM, the activity of aldehyde oxidase increases, which contributes to the increase of seed germination. The optimal concentration for seed germination is 1 Mm molybdenum concentration. As the concentration of molybdenum increases, the resistance of adult plants to virus infection also increases. Thus, 1 mМ molybdenum concentration is optimal for seed germination and resistance to virus infection. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) plays a crucial role in plant defense mechanisms under adverse environmental conditions, but its metabolism and perception in response to heavy metals are largely unknown. In Pisum sativum exposed to CdCl2, an accumulation of free ABA was detected in leaves at different developmental stages (A, youngest, unexpanded; B1, youngest, fully expanded; B2, mature; C, old), with the highest content found in A and B1 leaves. In turn, the content of ABA conjugates, which was highest in B2 and C leaves under control conditions, increased only in A leaves and decreased in leaves of later developmental stages after Cd treatment. Based on the expression of PsNCED2, PsNCED3 (9-cis-epoxycarotenoid dioxygenase), PsAO3 (aldehyde oxidase) and PsABAUGT1 (ABA-UDP-glucosyltransferase), and the activity of PsAOγ, B2 and C leaves were found to be the main sites of Cd-induced de novo synthesis of ABA from carotenoids and ABA conjugation with glucose. In turn, β-glucosidase activity and the expression of genes encoding ABA receptors (PsPYL2, PsPYL4, PsPYL8, PsPYL9) suggest that in A and B1 leaves, Cd-induced release of ABA from inactive ABA-glucosyl esters and enhanced ABA perception comes to the forefront when dealing with Cd toxicity. The distinct role of leaves at different developmental stages in defense against the harmful effects of Cd is discussed.

Nitric oxide (NO) plays numerous essential physiological functions in terrestrial animals. In mammals, NO production from l-arginine is catalysed by the enzyme NO synthase (NOS). In recent years, data have begun to emerge on NOS expression and the physiological significance of NO in ectothermic vertebrates such as fish. However, there are relatively fewer data compared to the mammalian system. Although it is already well known that animal molybdoenzymes can convert nitrate and nitrite into NO, there is almost no information on the content and properties of molybdoenzymes in fish organs in the scientific literature. In this regard, the objectives of the present work were to detect the activity of classical molybdoenzymes xanthine oxidase (XO) and aldehyde oxidase (AO) in the liver and to study their possible activity to reduce nitrate and nitrite to nitrogen monoxide. In this work, the intrinsic activity of XO and AO was examined by using their substrates. At the same time, their nitrate (NR) - and nitrite reductase (NiR) activity were determined. It was determined that XO and AO in the fish liver are mainly represented by the molybdenum-free forms. The presence of an additional source of molybdenum can activate both the intrinsic and the NR and NiR activities. The NiR activity of XO and AO was higher than their NR activity. The data shows that treatment at a concentration of sodium molybdate and glutathione of 1.0 mM increases all activities of XO and AO. The optimal conditions for maximum activation of exogenous molybdate for XO and AO were reached by heating at 70 °C for 5 minutes. The activity of XO increased almost 4.7 times, and the activity of AO 7.7 times compared with its intrinsic activity without heat treatment. NO is formed from nitrite by the enzymes XO and AO much more than from nitrate.

SGX523 is a c-Met tyrosine kinase inhibitor that failed in clinical trials because of renal toxicity caused by crystal deposits in renal tubules. SGX523 is metabolized by aldehyde oxidase (AOX) in a species-dependent manner to the considerably less soluble 2-quinolinone-SGX523, which is likely involved in the clinically observed obstructive nephropathy. This study investigated the metabolism and renal toxicity of SGX523 in chimeric mice with humanized livers (humanized-liver mice). The 2-quinolinone-SGX523 formation activity was higher in humanized-liver mouse and human hepatocytes than in mouse hepatocytes. Additionally, this activity in the liver cytosolic fraction from humanized-liver mice was inhibited by the AOX inhibitors raloxifene and hydralazine. After oral SGX523 administration, higher maximum concentrations, larger areas under the plasma concentration versus time curves, and higher urinary concentrations of 2-quinolinone-SGX523 were observed in humanized-liver mice than in non-humanized mice. Serum creatinine and blood urea nitrogen levels were elevated in humanized-liver mice following repeated oral SGX523 administration. The accumulation of amorphous material in the tubules and infiltration of inflammatory cells around tubules were observed in the kidneys of humanized-liver mice after repeated oral SGX523 administration. These findings demonstrate that humanized-liver mice are useful for understanding the metabolism and toxicity of SGX523. [Display omitted] • Human AOX1 metabolizes SGX523 into 2-quinolinone SGX523 in humanized-liver mice. • The less soluble 2-quinolinone SGX523 may be involved in SGX523-induced nephropathy. • Humanized-liver mice can help predict metabolism-dependent drug toxicities. [ABSTRACT FROM AUTHOR]

Drug metabolism is critical to many aspects of drug safety and efficacy. Thus, understanding the various intricacies of drug metabolism helps to avoid possible pitfalls and contributes to drug design during the discovery stage and to drug characterization during the development stage. Drug metabolism expertise was mainly taught at the universities and prominent professors have contributed to the field and here, we like to acknowledge Professor Thomas Baillie contribution to the field. Over the years, we have deployed our learnings in both drug discovery and development at Genentech. Here, we describe case studies from novel reaction mechanisms to novel types of metabolites. Taken together, we describe our lessons in a manner that allows the reader to understand the significance of the lessons and the thinking behind the experimental design. Here, we describe 10 case studies that highlight our experience in the past decade with the first 5 case studies capturing non-cytochrome P450 enzymatic reactions such as the discovery of a novel hydrolytic reaction by aldehyde oxidase (Case Study 1). The remaining 5 case studies describe cytochrome P450-derived reactions including various types of cytochrome P450-mediated reactions that resulted in ring formation or cleavage (Case Studies 6–9). The last case study (Case Study 10), emphasizes the importance of exploring ways to understand binding and enzyme kinetics, and explores co-incubation of 1-aminobenzotriazole (ABT) with warfarin in CYP2C9 metabolism. [ABSTRACT FROM AUTHOR]

Molybdenum-containing enzymes of the xanthine oxidase (XO) family are well known to catalyse oxygen atom transfer reactions, with the great majority of the characterised enzymes catalysing the insertion of an oxygen atom into the substrate. Although some family members are known to catalyse the "reverse" reaction, the capability to abstract an oxygen atom from the substrate molecule is not generally recognised for these enzymes. Hence, it was with surprise and scepticism that the "molybdenum community" noticed the reports on the mammalian XO capability to catalyse the oxygen atom abstraction of nitrite to form nitric oxide (NO). The lack of precedent for a molybdenum- (or tungsten) containing nitrite reductase on the nitrogen biogeochemical cycle contributed also to the scepticism. It took several kinetic, spectroscopic and mechanistic studies on enzymes of the XO family and also of sulfite oxidase and DMSO reductase families to finally have wide recognition of the molybdoenzymes' ability to form NO from nitrite. Herein, integrated in a collection of "personal views" edited by Professor Ralf Mendel, is an overview of my personal journey on the XO and aldehyde oxidase-catalysed nitrite reduction to NO. The main research findings and the path followed to establish XO and AO as competent nitrite reductases are reviewed. The evidence suggesting that these enzymes are probable players of the mammalian NO metabolism is also discussed. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) plays numerous essential physiological functions in terrestrial animals. In mammals, NO production from l-arginine is catalysed by the enzyme NO synthase (NOS). In recent years, data have begun to emerge on NOS expression and the physiological significance of NO in ectothermic vertebrates such as fish. However, there are relatively fewer data compared to the mammalian system. Although it is already well known that animal molybdoenzymes can convert nitrate and nitrite into NO, there is almost no information on the content and properties of molybdoenzymes in fish organs in the scientific literature. In this regard, the objectives of the present work were to detect the activity of classical molybdoenzymes xanthine oxidase (XO) and aldehyde oxidase (AO) in the liver and to study their possible activity to reduce nitrate and nitrite to nitrogen monoxide. In this work, the intrinsic activity of XO and AO was examined by using their substrates. At the same time, their nitrate (NR) - and nitrite reductase (NiR) activity were determined. It was determined that XO and AO in the fish liver are mainly represented by the molybdenum-free forms. The presence of an additional source of molybdenum can activate both the intrinsic and the NR and NiR activities. The NiR activity of XO and AO was higher than their NR activity. The data shows that treatment at a concentration of sodium molybdate and glutathione of 1.0 mM increases all activities of XO and AO. The optimal conditions for maximum activation of exogenous molybdate for XO and AO were reached by heating at 70 °C for 5 minutes. The activity of XO increased almost 4.7 times, and the activity of AO 7.7 times compared with its intrinsic activity without heat treatment. NO is formed from nitrite by the enzymes XO and AO much more than from nitrate. [ABSTRACT FROM AUTHOR]

Abstract Background Allopurinol is used to treat hyperuricemia and gout. It is metabolized to oxypurinol by xanthine oxidase (XO), and aldehyde oxidase (AO). Allopurinol and oxypurinol are potent XO inhibitors that reduce the plasma uric acid levels. Although oxypurinol levels show large inter-individual variations, high concentrations of oxypurinol can cause various adverse effects. Therefore, it is important to understand allopurinol metabolism by XO and AO. In this study we aimed to estimate the role of AO and XO in allopurinol metabolism by pre-administering Crl:CD and Jcl:SD rats, which have known strain differences in AO activity, with XO inhibitor febuxostat. Methods Allopurinol (30 or 100 mg/kg) was administered to Crl:CD and Jcl:SD rats with low and high AO activity, respectively, after pretreatment with or without febuxostat. The serum concentrations of allopurinol and oxypurinol were measured, and the area under the concentration-time curve (AUC) was calculated from the 48 h serum concentration-time profile. In vivo metabolic activity was measured as the ratio AUCoxypurinol /AUCallopurinol. Results Although no strain-specific differences were observed in the AUCoxypurinol/AUCallopurinol ratio in the allopurinol (30 mg/kg)-treated group, the ratio in Jcl:SD rats was higher than that in Crl:CD rats after febuxostat pretreatment. Contrastingly, the AUC ratio of allopurinol (100 mg/kg) was approximately 2-fold higher in Jcl:SD rats than that in Crl:CD rats. These findings showed that Jcl:SD rats had higher intrinsic AO activity than Crl:CD rats did. However, febuxostat pretreatment substantially decreased the activity, as measured by the AUC ratio using allopurinol (100 mg/kg), to 46 and 63% in Crl:CD rats and Jcl:SD rats, respectively, compared to the control group without febuxostat pretreatment. Conclusions We elucidated the role of XO and AO in allopurinol metabolism in Crl:CD and Jcl:SD rats. Notably, AO can exert a proportionately greater impact on allopurinol metabolism at high allopurinol concentrations. AO’s impact on allopurinol metabolism is meaningful enough that individual differences in AO may explain allopurinol toxicity events. Considering the inter-individual differences in AO activity, these findings can aid to dose adjustment of allopurinol to avoid potential adverse effects.

In the treatment of COVID-19, the antiviral medication Favipiravir has proven to be quite successful. Its metabolism is mediated by the enzymes aldehyde oxidase (AO) and xanthine oxidase (XO). This research investigated the potential drug-drug interaction between favipiravir and amlodipine in hypertensive rabbits. Twenty local adult male rabbits (aged between 10 and 12 months and weighing between 2 and 2.5 kg) were induced with hypertension by 20 mg/kg BW of desoxycorticosterone acetate subcutaneously for three weeks and then divided randomly into two groups of ten. The first group received a single oral dose of 40 mg/kg BW of favipiravir, while the second group received 5 mg/kg of amlodipine orally for 14 consecutive days to inhibit AO before receiving a single oral dose of 40 mg/kg BW of favipiravir. Blood samples were collected from the marginal ear vein at 15, 30, 45 min, and 1, 2, 4, 8, 12, 24, 48, 36, 48, and 72 h. High-performance liquid chromatography (HPLC) was used to determine the concentration of favipiravir in the plasma. The results showed that co-administration of amlodipine prolonged the time taken for favipiravir (Tmax) to reach maximum plasma concentration (Cmax) and decreased its elimination half-life, while increasing the area under the curve (AUC). Amlodipine also prolonged the elimination of favipiravir by reducing the clearance per unit time (Cl/f). Additionally, hypertension potentiated the effect of amlodipine on the absorption, distribution, metabolism, and excretion of favipiravir. In conclusion, concomitant use of favipiravir with other drugs that affect AO enzyme activity may alter the pharmacokinetic profile of the drug. Therefore, adjusting the dose of favipiravir administered to hypertensive patients receiving amlodipine is recommended.

Lactiplantibacillus plantarum‎, one of lactic acid bacteria (LAB), is found in various foods, ‎‎including dairy products, meat, and vegetables, and most of these bacteria offer beneficial ‎‎effects to humans and animals as potential probiotics with broad-spectrum antimicrobial ‎‎activities. The aim of this study was evaluating the antibacterial efficacy of L. ‎plantarum‎ ‎against ‎some foodborne bacteria isolated from dairy products. This research involved 34 dairy ‎products, ‎including local and imported milk, cheese, and yogurt sold locally in Baghdad ‎province, Iraq, during May ‎‎2022. For the isolation of L. ‎plantarum‎, a special medium called ‎MRS (de Man Rogosa and ‎Sharpe) was applied. Colonies were purified and identified by routine ‎bacteriological methods, ‎Vitek2 system, and confirmed by the polymerase chain reaction (PCR) ‎targeting the ‎16S rRNA‎ ‎gene followed by the amplicon sequencing. Other aerobic bacteria ‎contaminating dairy products ‎were also isolated onto sterile selective media specific for each ‎microorganism, and the isolates ‎were identified by routine diagnostics tests followed by ‎verification with Vitek2 system. Then, ‎the culture supernatant of L. ‎plantarum‎ was tested for its ‎antagonistic activity toward foodborne ‎bacteria by the use of agar well diffusion assay. The ‎findings showed the isolation of 2 L. ‎plantarum‎‎, 3 Pseudomonas aeruginosa‎, 4 Escherichia coli, ‎one isolate of Bacillus subtilis, and ‎another Staphylococcus hominis. The filtered supernatant of ‎L. ‎plantarum‎ was significantly ‎efficient in inhibiting the growth of the above bacteria. Each of E. ‎coli and B. subtilis‎ revealed ‎zones of inhibition of 36 and 38 mm in diameter, respectively, ‎while P. aeruginosa‎ and S. ‎hominis had inhibition zones diameters of 27 and 29 mm, ‎respectively. This suggests that the L. ‎plantarum‎‎ supernatant possesses a broad-spectrum ‎activity against foodborne bacteria. To ‎conclude, locally made dairy products can hold different ‎contaminating bacteria, which can be ‎eliminated by using probiotics, such as L. ‎plantarum‎, to ‎avoid foodborne diseases onset‎.

Copyright of Iraqi Journal of Veterinary Medicine is the property of Republic of Iraq Ministry of Higher Education & Scientific Research (MOHESR) and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Iraqi Journal of Veterinary Medicine is the property of Republic of Iraq Ministry of Higher Education & Scientific Research (MOHESR) and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Background: Allopurinol is used to treat hyperuricemia and gout. It is metabolized to oxypurinol by xanthine oxidase (XO), and aldehyde oxidase (AO). Allopurinol and oxypurinol are potent XO inhibitors that reduce the plasma uric acid levels. Although oxypurinol levels show large inter-individual variations, high concentrations of oxypurinol can cause various adverse effects. Therefore, it is important to understand allopurinol metabolism by XO and AO. In this study we aimed to estimate the role of AO and XO in allopurinol metabolism by pre-administering Crl:CD and Jcl:SD rats, which have known strain differences in AO activity, with XO inhibitor febuxostat. Methods: Allopurinol (30 or 100 mg/kg) was administered to Crl:CD and Jcl:SD rats with low and high AO activity, respectively, after pretreatment with or without febuxostat. The serum concentrations of allopurinol and oxypurinol were measured, and the area under the concentration-time curve (AUC) was calculated from the 48 h serum concentration-time profile. In vivo metabolic activity was measured as the ratio AUCoxypurinol /AUCallopurinol. Results: Although no strain-specific differences were observed in the AUCoxypurinol/AUCallopurinol ratio in the allopurinol (30 mg/kg)-treated group, the ratio in Jcl:SD rats was higher than that in Crl:CD rats after febuxostat pretreatment. Contrastingly, the AUC ratio of allopurinol (100 mg/kg) was approximately 2-fold higher in Jcl:SD rats than that in Crl:CD rats. These findings showed that Jcl:SD rats had higher intrinsic AO activity than Crl:CD rats did. However, febuxostat pretreatment substantially decreased the activity, as measured by the AUC ratio using allopurinol (100 mg/kg), to 46 and 63% in Crl:CD rats and Jcl:SD rats, respectively, compared to the control group without febuxostat pretreatment. Conclusions: We elucidated the role of XO and AO in allopurinol metabolism in Crl:CD and Jcl:SD rats. Notably, AO can exert a proportionately greater impact on allopurinol metabolism at high allopurinol concentrations. AO's impact on allopurinol metabolism is meaningful enough that individual differences in AO may explain allopurinol toxicity events. Considering the inter-individual differences in AO activity, these findings can aid to dose adjustment of allopurinol to avoid potential adverse effects. [ABSTRACT FROM AUTHOR]

Aldehyde oxidase (AO) has garnered curiosity as a non-CYP metabolizing enzyme in drug development due to unexpected consequences such as toxic metabolite generation and high metabolic clearance resulting in the clinical failure of new drugs. Therefore, poor AO mediated clearance prediction in preclinical nonhuman species remains a significant obstacle in developing novel drugs. Various isoforms of AO, such as AOX1, AOX3, AOX3L1, and AOX4 exist across species, and different AO activity among humans influences the AO mediated drug metabolism. Therefore, carefully considering the unique challenges is essential in developing successful AO substrate drugs. The in vitro to in vivo extrapolation underpredicts AO mediated drug clearance due to the lack of reliable representative animal models, substrate-specific activity, and the discrepancy between absolute concentration and activity. An in vitro tool to extrapolate in vivo clearance using a yard-stick approach is provided to address the underprediction of AO mediated drug clearance. This approach uses a range of well-known AO drug substrates as calibrators for qualitative scaling new drugs into low, medium, or high clearance category drugs. So far, in vivo investigations on chimeric mice with humanized livers (humanized mice) have predicted AO mediated metabolism to the best extent. This review addresses the critical aspects of the drug discovery stage for AO metabolism studies, challenges faced in drug development, approaches to tackle AO mediated drug clearance's underprediction, and strategies to decrease the AO metabolism of drugs. [ABSTRACT FROM AUTHOR]

Molybdenum-containing enzymes of the xanthine oxidase (XO) family are well known to catalyse oxygen atom transfer reactions, with the great majority of the characterised enzymes catalysing the insertion of an oxygen atom into the substrate. Although some family members are known to catalyse the “reverse” reaction, the capability to abstract an oxygen atom from the substrate molecule is not generally recognised for these enzymes. Hence, it was with surprise and scepticism that the “molybdenum community” noticed the reports on the mammalian XO capability to catalyse the oxygen atom abstraction of nitrite to form nitric oxide (NO). The lack of precedent for a molybdenum- (or tungsten) containing nitrite reductase on the nitrogen biogeochemical cycle contributed also to the scepticism. It took several kinetic, spectroscopic and mechanistic studies on enzymes of the XO family and also of sulfite oxidase and DMSO reductase families to finally have wide recognition of the molybdoenzymes’ ability to form NO from nitrite. Herein, integrated in a collection of “personal views” edited by Professor Ralf Mendel, is an overview of my personal journey on the XO and aldehyde oxidase-catalysed nitrite reduction to NO. The main research findings and the path followed to establish XO and AO as competent nitrite reductases are reviewed. The evidence suggesting that these enzymes are probable players of the mammalian NO metabolism is also discussed.

Histamine (HIST) and other biogenic amines found in fish and fishery products accumulated by the action of bacterial amino acid decarboxylase cannot be decomposed and eliminated by heating or other chemical methods. A simple method for HIST elimination is proposed by a coupling reaction of the fungal amine oxidase (FAO) and bacterial aldehyde oxidase (ALOX) of acetic acid bacteria. As a model reaction, FAO oxidized benzylamine to benzaldehyde, which in turn was oxidized spontaneously to benzoic acid with ALOX. Likely, in HIST elimination, FAO coupled well with ALOX to produce imidazole 4-acetic acid from HIST with an apparent yield of 100%. Imidazole 4-acetaldehyde was not detected in the reaction mixture. In the absence of ALOX, the coupling reaction was incomplete given a number of unidentified substances in the reaction mixture. The proposed coupling enzymatic method may be highly effective to eliminate toxic amines from fish and fishery products. [ABSTRACT FROM AUTHOR]

Despite increased awareness of aldehyde oxidase (AO) as a major drug-metabolising enzyme, predicting the pharmacokinetics of its substrates remains challenging. Several drug candidates have been terminated due to high clearance, which were subsequently discovered to be AO substrates. Even retrospective extrapolation of human clearance, from models more sensitive to AO activity, often resulted in underprediction. The questions of the current work thus were: Is there an acceptable degree of in vitro AO metabolism that does not result in high in vivo human clearance? And, if so, how can this be predicted? We built an in vitro/in vivo correlation using known AO substrates, combining multiple in vitro parameters to calculate the blood metabolic clearance mediated by AO (CLbAO). This value was compared with observed blood clearance (CLb-obs), establishing cut-off CLbAO values, to discriminate between low and high CLb-obs. The model was validated using additional literature compounds, and CLb-obs was predicted in the correct category. This simple, categorical, semi-quantitative yet multi-factorial model is readily applicable in drug discovery. Further, it is valuable for high-clearance compounds, as it predicts the CLb group, rather than an exact CLb value, for the substrates of this poorly-characterised enzyme. [ABSTRACT FROM AUTHOR]

With the increasing use of vaping devices that deliver high levels of nicotine (NIC) to the lungs, sporadic lung injury has been observed. Commercial vaping solutions can contain high NIC concentrations of 150 mM or more. With high NIC levels, its metabolic products may induce toxicity. NIC is primarily metabolized to form NIC iminium (NICI) which is further metabolized by aldehyde oxidase (AOX) to cotinine. We determine that NICI in the presence of AOX is a potent trigger of superoxide generation. NICI stimulated superoxide generation from AOX with K m  = 2.7 μM and V max  = 794 nmol/min/mg measured by cytochrome-c reduction. EPR spin-trapping confirmed that NICI in the presence of AOX is a potent source of superoxide. AOX is expressed in the lungs and chronic e-cigarette exposure in mice greatly increased AOX expression. NICI or NIC stimulated superoxide production in the lungs of control mice with an even greater increase after chronic e-cigarette exposure. This superoxide production was quenched by AOX inhibition. Furthermore, e-cigarette-mediated NIC delivery triggered oxidative lung damage that was blocked by AOX inhibition. Thus, NIC metabolism triggers AOX-mediated superoxide generation that can cause lung injury. Therefore, high uncontrolled levels of NIC inhalation, as occur with e-cigarette use, can induce oxidative lung damage., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Immune response to SARS-CoV-2 and ensuing inflammation pose a huge challenge to the host's nicotinamide adenine dinucleotide (NAD+) metabolism. Humans depend on vitamin B3 for biosynthesis of NAD+, indispensable for many metabolic and NAD+-consuming signaling reactions. The balance between its utilization and resynthesis is vitally important. Many extra-pulmonary symptoms of COVID-19 strikingly resemble those of pellagra, vitamin B3 deficiency (e.g., diarrhoea, dermatitis, oral cavity and tongue manifestations, loss of smell and taste, mental confusion). In most developed countries, pellagra is successfully eradicated by vitamin B3 fortification programs. Thus, conceivably, it has not been suspected as a cause of COVID-19 symptoms. Here, the deregulation of the NAD+ metabolism in response to the SARS-CoV-2 infection is reviewed, with special emphasis on the differences in the NAD+ biosynthetic pathway's efficiency in conditions predisposing for the development of serious COVID-19. SARS-CoV-2 infection-induced NAD+ depletion and the elevated levels of its metabolites contribute to the development of a systemic disease. Acute liberation of nicotinamide (NAM) in antiviral NAD+-consuming reactions potentiates "NAM drain", cooperatively mediated by nicotinamide N-methyltransferase and aldehyde oxidase. "NAM drain" compromises the NAD+ salvage pathway's fail-safe function. The robustness of the host's NAD+ salvage pathway, prior to the SARS-CoV-2 infection, is an important determinant of COVID-19 severity and persistence of certain symptoms upon resolution of infection. [ABSTRACT FROM AUTHOR]

Metopimazine (MPZ) is a peripherally restricted, dopamine D2 receptor antagonist used for four decades to treat acute nausea and vomiting. MPZ is currently under clinical investigation for the treatment of gastroparesis (GP). MPZ undergoes high first‐pass metabolism that produces metopimazine acid (MPZA), the major circulating metabolite in humans. Despite a long history of use, the enzymes involved in the metabolism of MPZ have not been identified. Here we report a series of studies designed to identify potential MPZ metabolites in vitro, determine their clinical relevance in humans, and elucidate the enzymes responsible for their formation. The findings demonstrated that the formation of MPZA was primarily catalyzed by human liver microsomal amidase. Additionally, human liver cytosolic aldehyde oxidase (AO) catalyzes the formation of MPZA, in vitro, although to a much lesser extent. Neither cytochrome P450 enzymes nor flavin‐monooxygenases (FMO) were involved in the formation MPZA, although two minor oxidative pathways were catalyzed by CYP3A4 and CYP2D6 in vitro. Analysis of plasma samples from subjects dosed 60 mg of MPZ verified that these oxidative pathways are very minor and that CYP enzyme involvement was negligible compared to microsomal amidase/hydrolase in overall MPZ metabolism in humans. The metabolism by liver amidase, an enzyme family not well defined in small molecule drug metabolism, with minimal metabolism by CYPs, differentiates this drug from current D2 antagonists used or in development for the treatment of GP. [ABSTRACT FROM AUTHOR]

Aldehyde oxidases (AOXs) are common detoxifying enzymes in several organisms. In insects, AOXs act in xenobiotic metabolism and as odorant-degrading enzymes (ODEs). These last appear as crucial enzymes in the life cycle of insects, helping to reset their olfactory system, particularly in lepidopterans, which fulfill important ecological roles (e.g., pollination or destructive life cycles). A comprehensive understanding of their olfactory system has provided opportunities to study key chemosensory proteins. However, no significant advance has been made around lepidopteran AOXs research, and even less around butterflies, a recently evolved lineage. In this study we have identified novel AOX gene families in moths and butterflies in order to understand their role as ODEs. Eighteen genomes from both moths and butterflies were used for phylogenetics, molecular evolution and sequence analyses. We identified 164 AOXs, from which 91 are new. Their phylogeny showed two main clades that are potentially related to odorant-degrading function, where both moths and butterflies have AOXs. A first ODE-related clade seems to have a non-ditrysian origin, likely related to plant volatiles. A second ODE-related clade could be more pheromone-biased. Molecular evolution analysis suggests a slight purifying selection process, though a number of sites appeared under positive selection. ODE-related AOXs have changed a phenylalanine residue by proline in the active site. Finally, this study could serve as a reference for further evolutionary and functional studies around Lepidopteran AOXs.

Abstract Metopimazine (MPZ) is a peripherally restricted, dopamine D2 receptor antagonist used for four decades to treat acute nausea and vomiting. MPZ is currently under clinical investigation for the treatment of gastroparesis (GP). MPZ undergoes high first‐pass metabolism that produces metopimazine acid (MPZA), the major circulating metabolite in humans. Despite a long history of use, the enzymes involved in the metabolism of MPZ have not been identified. Here we report a series of studies designed to identify potential MPZ metabolites in vitro, determine their clinical relevance in humans, and elucidate the enzymes responsible for their formation. The findings demonstrated that the formation of MPZA was primarily catalyzed by human liver microsomal amidase. Additionally, human liver cytosolic aldehyde oxidase (AO) catalyzes the formation of MPZA, in vitro, although to a much lesser extent. Neither cytochrome P450 enzymes nor flavin‐monooxygenases (FMO) were involved in the formation MPZA, although two minor oxidative pathways were catalyzed by CYP3A4 and CYP2D6 in vitro. Analysis of plasma samples from subjects dosed 60 mg of MPZ verified that these oxidative pathways are very minor and that CYP enzyme involvement was negligible compared to microsomal amidase/hydrolase in overall MPZ metabolism in humans. The metabolism by liver amidase, an enzyme family not well defined in small molecule drug metabolism, with minimal metabolism by CYPs, differentiates this drug from current D2 antagonists used or in development for the treatment of GP.

SUMMARY: The Arabidopsis thaliana aldehyde oxidase 3 (AAO3) catalyzes the oxidation of abscisic aldehyde (ABal) to abscisic acid (ABA). Besides ABal, plants generate other aldehydes that can be toxic above a certain threshold. AAO3 knockout mutants (aao3) exhibited earlier senescence but equivalent relative water content compared with wild‐type (WT) during normal growth or upon application of UV‐C irradiation. Aldehyde profiling in leaves of 24‐day‐old plants revealed higher accumulation of acrolein, crotonaldehyde, 3Z‐hexenal, hexanal and acetaldehyde in aao3 mutants compared with WT leaves. Similarly, higher levels of acrolein, benzaldehyde, crotonaldehyde, propionaldehyde, trans‐2‐hexenal and acetaldehyde were accumulated in aao3 mutants upon UV‐C irradiation. Aldehydes application to plants hastened profuse senescence symptoms and higher accumulation of aldehydes, such as acrolein, benzaldehyde and 4‐hydroxy‐2‐nonenal, in aao3 mutant leaves as compared with WT. The senescence symptoms included greater decrease in chlorophyll content and increase in transcript expression of the early senescence marker genes, Senescence‐Related‐Gene1, Stay‐Green‐Protein2 as well as NAC‐LIKE, ACTIVATED‐BY AP3/P1. Notably, although aao3 had lower ABA content than WT, members of the ABA‐responding genes SnRKs were expressed at similar levels in aao3 and WT. Moreover, the other ABA‐deficient mutants [aba2 and 9‐cis‐poxycarotenoid dioxygenase3‐2 (nced3‐2), that has functional AAO3] exhibited similar aldehydes accumulation and chlorophyll content like WT under normal growth conditions or UV‐C irradiation. These results indicate that the absence of AAO3 oxidation activity and not the lower ABA and its associated function is responsible for the earlier senescence symptoms in aao3 mutant. Significance Statement: The current study demonstrates that AAO3, known to catalyze the last step of ABA biosynthesis by oxidizing ABal, is essential in preventing leaf damage by oxidizing toxic levels of aldehydes in rosette leaves. We provided credible evidences that premature senescence in this study was not because of low ABA content and its consequential function in aao3, but rather due to higher aldehyde accumulation, which is caused by AAO3 deficiency. [ABSTRACT FROM AUTHOR]

Despite the increasing importance of aldehyde oxidase (AO) in the drug metabolism of clinical candidates, ontogeny data for AO are limited. The objective of our study was to characterize the age-dependent AO content and activity in the human liver cytosolic fraction (HLC) and human hepatocytes (HH). HLC ( n = 121 donors) and HH ( n = 50 donors) were analyzed for (1) AO protein content by quantitative proteomics and (2) enzyme activity using carbazeran as a probe substrate. AO activity showed high technical variability and poor correlation with the content in HLC samples, whereas hepatocyte samples showed a strong correlation between the content and activity. Similarly, AO content and activity showed no significant age-dependent differences in HLC samples, whereas the average AO content and activity in hepatocytes increased significantly (∼20-40-fold) from the neonatal levels (0-28 days). Based on the hepatocyte data, the age at which 50% of the adult AO content is reached (age 50 ) was 3.15 years (0.32-13.97 years, 95% CI). Metabolite profiling of carbazeran revealed age-dependent metabolic switching and the role of non-AO mechanisms (glucuronidation and desmethylation) in carbazeran elimination. The content-activity correlation in hepatocytes improved significantly ( R 2 = 0.95; p < 0.0001) in samples showing <10% contribution of glucuronidation toward the overall metabolism, confirming that AO-mediated oxidation and glucuronidation are the key routes of carbazeran metabolism. Considering the confounding effect of glucuronidation on AO activity, AO content-based ontogeny data are a more direct reflection of developmental changes in protein expression. The comprehensive ontogeny data of AO in HH samples are more reliable than HLC data, which are important for developing robust physiologically based pharmacokinetic models for predicting AO-mediated metabolism in children.

Background: Methotrexate (MTX) is partially metabolized by aldehyde oxidase (AOX) in the liver and its clinical impact remains unclear. In this study, we aimed to demonstrate how AOX contributes to MTX-induced hepatotoxicity in vitro and clarify the relationship between concomitant AOX inhibitor use and MTX-associated liver injury development using the U.S. Food and Drug Administration Adverse Event Reporting System (FAERS)., Methods: We assessed intracellular MTX accumulation and cytotoxicity using HepG2 cells. We used the FAERS database to detect reporting odds ratio (ROR)-based MTX-related hepatotoxicity event signals., Results: AOX inhibition by AOX inhibitor raloxifene and siRNA increased the MTX accumulation in HepG2 cells and enhanced the MTX-induced cell viability reduction. In the FAERS analysis, the ROR for MTX-related hepatotoxicity increased with non-overlap of 95% confidence interval when co-administered with drugs with higher I max, u (maximum unbound plasma concentration)/IC 50 (half-maximal inhibitory concentration for inhibition of AOX) calculated based on reported pharmacokinetic data., Conclusion: AOX inhibition contributed to MTX accumulation in the liver, resulting in increased hepatotoxicity. Our study raises concerns regarding MTX-related hepatotoxicity when co-administered with drugs that possibly inhibit AOX activity at clinical concentrations.

The influence of drought stress on barley (Hordeum vulgare) has been investigated. The experiments were conducted with seedlings of barley—by the application of combined drought and temperature stresses. Here we showed that combination of drought and high temperature inflicted more severe damage to plants than the drought and low temperature stress. The temperature stress triggered more drastic changes in plant morphology/physiology and biochemistry than the drought stress. Interestingly, plants exposed to high temperature exhibited significant reduction of shoot and root aldehyde oxidase (AO) activity. Moreover, increased temperature resulted in lower levels of reactive oxygen species (ROS), while drought stress had an opposite effect on ROS accumulation and AO activity. This is the first demonstration of inhibition of plant AO and ROS in response to heat stress. The combined heat and drought stresses resulted in increased activity of catalase (CAT) and superoxide dismutase (SOD) in roots but not in shoots. Our findings indicate that heat and drought stresses may induce activation of different antioxidant enzymatic defense mechanisms and heat stress significantly affects enzymes responsible for the ROS accumulation in plants. [ABSTRACT FROM AUTHOR]

SUMMARY: Temperature passively affects biological processes involved in plant growth. Therefore, it is challenging to study the dedicated temperature signalling pathways that orchestrate thermomorphogenesis, a suite of elongation growth‐based adaptations that enhance leaf‐cooling capacity. We screened a chemical library for compounds that restored hypocotyl elongation in the pif4‐2–deficient mutant background at warm temperature conditions in Arabidopsis thaliana to identify modulators of thermomorphogenesis. The small aromatic compound 'Heatin', containing 1‐iminomethyl‐2‐naphthol as a pharmacophore, was selected as an enhancer of elongation growth. We show that ARABIDOPSIS ALDEHYDE OXIDASES redundantly contribute to Heatin‐mediated hypocotyl elongation. Following a chemical proteomics approach, the members of the NITRILASE1‐subfamily of auxin biosynthesis enzymes were identified among the molecular targets of Heatin. Our data reveal that nitrilases are involved in promotion of hypocotyl elongation in response to high temperature and Heatin‐mediated hypocotyl elongation requires the NITRILASE1‐subfamily members, NIT1 and NIT2. Heatin inhibits NIT1‐subfamily enzymatic activity in vitro and the application of Heatin accordingly results in the accumulation of NIT1‐subfamily substrate indole‐3‐acetonitrile in vivo. However, levels of the NIT1‐subfamily product, bioactive auxin (indole‐3‐acetic acid), were also significantly increased. It is likely that the stimulation of hypocotyl elongation by Heatin might be independent of its observed interaction with NITRILASE1‐subfamily members. However, nitrilases may contribute to the Heatin response by stimulating indole‐3‐acetic acid biosynthesis in an indirect way. Heatin and its functional analogues present novel chemical entities for studying auxin biology. Significance Statement: We identified the chemical compound Heatin that stimulates plant thermomorphogenesis and show that Heatin interferes with NIT1‐subfamily enzymatic activity and results in accumulation of auxin. The precise mechanism of action of Heatin requires further study but the investigations presented can contribute to knowledge‐based engineering of warm temperature‐resilient crops. Heatin is a promising agrochemical to steer plant growth and is a novel chemical tool for studying auxin biology. [ABSTRACT FROM AUTHOR]

Diquat (DQ) poisoning has garnered attention in recent years, primarily due to the rising incidence of cases worldwide, coupled with the absence of a viable antidote for its treatment. Despite the fact that diquat monopyridone (DQ-M) has been identified as a significant metabolite of DQ, the enzyme responsible for its formation remains unknown. In this study, we have identified aldehyde oxidase (AOX) as a vital enzyme involved in DQ oxidative metabolism. The metabolism of DQ to DQ-M was significantly inhibited by AOX inhibitors including raloxifene and hydralazine. The source of oxygen incorporated into DQ-M was proved to be from water through a H218O incubation experiment which further corroborated DQ-M formation via AOX metabolism. The product of DQ-M in vitro generated by fresh rat tissues co-incubation was consistent with its AOX expression. The result of the molecular docking analysis of DQ and AOX protein showed that DQ is capable of binding to AOX. Furthermore, the cytotoxicity of DQ was significantly higher than DQ-M at the same concentration tested in six cell types. This work is the first to uncover the involvement of aldehyde oxidase, a non-cytochrome P450 enzyme, in the oxidative metabolic pathway of diquat, thus providing a potential target for the development of detoxification treatment. [Display omitted] • AOX has been proved to be the key enzyme in the metabolism of DQ to DQ-M. • The cell viabilities of six types were all higher in DQ-M groups than DQ groups. • The metabolism of DQ to DQ-M by AOX is a detoxification metabolic process in vitro. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) regulates many aspects of plant growth and development and the responses to abiotic stresses. Arabidopsis aldehyde oxidase 3 (AAO3) catalyzes the final step of ABA biosynthesis. We cloned and functionally characterized a novel aldehyde oxidase gene, OsAO3 , the rice homolog of AAO3. OsAO3 was expressed in germinated seeds, roots, leaves, and floral organs, particularly in vascular tissues and guard cells, and its expression was significantly induced by exogenous ABA and mannitol. Mutation and overexpression of OsAO3 decreased and increased ABA levels, respectively, in seedling shoots and roots under both normal and drought stress conditions. The osao3 mutant exhibited earlier seed germination, increased seedling growth, and decreased drought tolerance compared to the wild-type, OsAO3 -overexpressing lines exhibited the opposite phenotype. Mutation and overexpression of OsAO3 increased and decreased grain yield, respectively, by affecting panicle number per plant, spikelet number per panicle, and spikelet fertility. Thus, OsAO3 may participate in ABA biosynthesis, and is essential for regulation of seed germination, seedling growth, grain yield, and drought tolerance in rice. • OsAO3 participates in ABA biosynthesis. • OsAO3 confers tolerance to drought stress. • OsAO3 is essential for regulation of seed germination and post-germination growth. • OsAO3 negatively regulates grain yield under well-watered conditions. [ABSTRACT FROM AUTHOR]

[Display omitted] • The pheromone gland transcriptome of Noorda blitealis was sequenced and analyzed. • 117 genes encoding pheromone biosynthesis and degradation enzymes were identified. • 79 pheromone biosynthesis and degradation genes exhibited a significant expression difference between PG and body. • Genome and transcriptome analyses reveal a novel AOX group in Pyraloidea. Noorda blitealis (Lepidoptera: Pyraloidea: Crambidae) is a major defoliating pest of Moringa trees. Focusing on its mating and reproduction, here we sequenced and analyzed the transcriptome of its pheromone glands (PGs) with a combination of Illumina sequencing, bioinformatics and phylogenetics approaches, coupled with a genome-based analysis. Transcriptome sequencing led to the yields of approximately 162 million clean reads, which were assembled into 60,578 unigenes and 121,692 transcripts, respectively. From the transcriptome, totally 117 genes encoding eight pheromone biosynthesis enzymes and one pheromone degradation enzyme were identified, 90 of which had complete open reading frames. A comparative analysis between PGs and bodies (removing PGs) revealed a large number of differentially expressed genes, including 79 pheromone biosynthesis and degradation related genes. Of the identified genes, NbliDES12 belonging to the △11 desaturase group was likely to a strong candidate for the desaturation of sex pheromones in N. blitealis , as implied by phylogenetic analyses and expression profiles. Finally and most notably, through genome and transcriptome analyses we discovered, for the first time, a novel aldehyde oxidase 6 (AOX6) group of the superfamily Pyraloidea that have been slightly expanded by gene duplications. Moreover, each orthologous AOX group shared highly conserved gene structure. Together, this current study has characterized the genes associated with sex pheromone biosynthesis and degradation from the PG transcriptome of N. blitealis , and more importantly, identifies a novel AOX group of the Pyraloidea. [ABSTRACT FROM AUTHOR]

Immune response to SARS-CoV-2 and ensuing inflammation pose a huge challenge to the host’s nicotinamide adenine dinucleotide (NAD+) metabolism. Humans depend on vitamin B3 for biosynthesis of NAD+, indispensable for many metabolic and NAD+-consuming signaling reactions. The balance between its utilization and resynthesis is vitally important. Many extra-pulmonary symptoms of COVID-19 strikingly resemble those of pellagra, vitamin B3 deficiency (e.g., diarrhoea, dermatitis, oral cavity and tongue manifestations, loss of smell and taste, mental confusion). In most developed countries, pellagra is successfully eradicated by vitamin B3 fortification programs. Thus, conceivably, it has not been suspected as a cause of COVID-19 symptoms. Here, the deregulation of the NAD+ metabolism in response to the SARS-CoV-2 infection is reviewed, with special emphasis on the differences in the NAD+ biosynthetic pathway’s efficiency in conditions predisposing for the development of serious COVID-19. SARS-CoV-2 infection-induced NAD+ depletion and the elevated levels of its metabolites contribute to the development of a systemic disease. Acute liberation of nicotinamide (NAM) in antiviral NAD+-consuming reactions potentiates “NAM drain”, cooperatively mediated by nicotinamide N-methyltransferase and aldehyde oxidase. “NAM drain” compromises the NAD+ salvage pathway’s fail-safe function. The robustness of the host’s NAD+ salvage pathway, prior to the SARS-CoV-2 infection, is an important determinant of COVID-19 severity and persistence of certain symptoms upon resolution of infection.

Nonalcoholic fatty liver disease, which has been rapidly increasing in the world in recent years, is roughly classified into nonalcoholic fatty liver (NAFL) and nonalcoholic steatohepatitis. This study was based on our previous reports that stated that the combination treatment of N1-methylnicotinamide (MNA) and hydralazine (HYD) improves fatty liver in NAFL model rats. This finding was attributed to the MNA metabolism inhibition by HYD, which is a strong inhibitor of aldehyde oxidase (AO); this results in an increase in hepatic MNA and improved fatty liver. We hypothesized that orally administered nicotinamide (NAM), which is the precursor of MNA and is a form of niacin, would be efficiently metabolized by nicotinamide N-methyltransferase in the presence of exogenous S-adenosylmethionine (SAM) in NAFL rats. To address this issue, NAFL model rats were orally administered with NAM, SAM, and/or HYD. As a result, liver triglyceride (TG) and lipid droplet levels were barely altered by the administration of NAM, SAM, NAM+SAM, or NAM+HYD. By contrast, the triple combination of NAM+SAM+HYD significantly reduced hepatic TG and lipid droplet levels and significantly increased hepatic MNA levels. These findings indicated that the combination of exogenous SAM with AO inhibitors, such as HYD, has beneficial effects for improving fatty liver with NAM. [ABSTRACT FROM AUTHOR]

Background: Aldehyde oxidase 1 (AOX1) is associated with various pathophysiological processes, including cancer. Specifically, AOX1 has been demonstrated to have a close relationship with the progression of certain cancers. However, the expression, function, and mechanisms of action of AOX1 in gallbladder cancer (GBC) remain unclear., Methods: Utilizing immunohistochemistry, the study quantified the prevalence of AOX1 within tissues of gallbladder carcinoma and those of the surrounding non-cancerous regions. In vitro assays using gallbladder carcinoma cell lines with modulated AOX1 expression levels were performed to assess the protein's role in cell proliferation, migration, and invasion. Furthermore, flow cytometry techniques were harnessed to determine the influence of AOX1 on the content of reactive oxygen species (ROS) in these cells. Additionally, the expression of epithelial-mesenchymal transition (EMT) markers and the Wnt/β-catenin signaling pathway markersin cells with varied AOX1 expression, detected through Western blot analyses. An in vivo xenograft model involving athymic mice was implemented to explore the influence of AOX1 on gallbladder tumor growth, with Western blot analysis applied to measure EMT marker expression in the resulting tumours., Results: Elevated AOX1 protein levels have been observed in gallbladder carcinoma tissues, with such upregulation linked to a negative prognostic outlook for patients. In vitro analyses demonstrate that enhanced AOX1 expression facilitates gallbladder carcinoma cell proliferation, migration, and invasion, while AOX1 suppression yields an inhibitory effect on these cellular behaviors. Western blot results reveal an inverse relationship between AOX1 and E-cadherin levels, yet was positively correlation with N-cadherin, Vimentin, and Snail within both gallbladder cancer cells and in vivo xenograft tumours. Further mechanistic investigation indicates that AOX1 elevation augments reactive oxygen species (ROS) production and initiates the Wnt/β-catenin signaling pathway in these cells. The application of N-acetylcysteine (NAC) and/or KY1797K attenuates the proliferative, migratory, and invasive enhancements imparted by AOX1 overexpression and reinforces these effects when AOX1 is silenced-achieved through ROS mitigation and the obstruction of the Wnt/β-catenin pathway. In vivo studies corroborate these findings, showing AOX1 overexpression to amplify xenograft tumor growth and mesenchymal marker expression, whereas AOX1 interference did the opposite., Conclusions: The study indicates that AOX1 functions as a carcinogenic factor in gallbladder carcinoma, enhancing cell proliferation, migration, invasion, and the EMT. These effects are driven by the activation of the Wnt/β-catenin pathway mediated by reactive oxygen species (ROS). Therefore,AOX1 presents potential as a valuable prognostic and diagnostic marker as well as a target for therapeutic intervention in the gallbladder cancer., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: No conflict of interest If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Abstract Aldehyde Oxidase (hAOX1) is a cytosolic enzyme involved in the metabolism of drugs and xenobiotic compounds. The enzyme belongs to the xanthine oxidase (XO) family of Mo containing enzyme and is a homo‐dimer of two 150 kDa monomers. Nonsynonymous Single Nucleotide Polymorphisms (nsSNPs) of hAOX1 have been reported as affecting the ability of the enzyme to metabolize different substrates. Some of these nsSNPs have been biochemically and structurally characterized but the lack of a systematic and comprehensive study regarding all described and validated nsSNPs is urgent, due to the increasing importance of the enzyme in drug development, personalized medicine and therapy, as well as in pharmacogenetic studies. The objective of the present work was to collect all described nsSNPs of hAOX1 and utilize a series of bioinformatics tools to predict their effect on protein structure stability with putative implications on phenotypic functional consequences. Of 526 nsSNPs reported in NCBI‐dbSNP, 119 are identified as deleterious whereas 92 are identified as nondeleterious variants. The stability analysis was performed for 119 deleterious variants and the results suggest that 104 nsSNPs may be responsible for destabilizing the protein structure, whereas five variants may increase the protein stability. Four nsSNPs do not have any impact on protein structure (neutral nsSNPs) of hAOX1. The prediction results of the remaining six nsSNPs are nonconclusive. The in silico results were compared with available experimental data. This methodology can also be used to identify and prioritize the stabilizing and destabilizing variants in other enzymes involved in drug metabolism.