Discovery of 4-(phenoxymethyl)-1H-1,2,3-triazole derivatives as novel xanthine oxidase inhibitors.

A series of 4-(phenoxymethyl)-1H-1,2,3-triazole derivatives were designed, synthesized, and evaluated for their xanthine oxidase (XO) inhibitory activities. Among these compounds, 9m emerged as the most effective XO inhibitor with an IC50 value of 0.70 µM, which was approximately 14-fold more potent than allopurinol. Additionally, compound 9m displayed favorable drug-like properties with ligand efficiency (LE) and lipophilic ligand efficiency (LLE) values of 0.33 and 3.41, respectively. We further explored the binding mode of 9m in complex with XO by molecular docking and molecular dynamics studies. In vivo hypouricemic studies also suggested that 9m could effectively lower the serum uric acid levels of rat. In summary, compound 9m could be a promising lead for further development of XO inhibitors.

Febuxostat does not delay progression of carotid atherosclerosis in patients with asymptomatic hyperuricemia: A randomized, controlled trial.

BACKGROUND: An elevated level of serum uric acid (SUA) is associated with an increased risk of cardiovascular disease. Pharmacological intervention with urate-lowering agents, such as the conventional purine analogue xanthine oxidase (XO) inhibitor, allopurinol, has been used widely for a long period of time in clinical practice to reduce SUA levels. Febuxostat, a novel non-purine selective inhibitor of XO, has higher potency for inhibition of XO activity and greater urate-lowering efficacy than conventional allopurinol. However, clinical evidence regarding the effects of febuxostat on atherosclerosis is lacking. The purpose of the study was to test whether treatment with febuxostat delays carotid intima-media thickness (IMT) progression in patients with asymptomatic hyperuricemia. METHODS AND FINDINGS: The study was a multicenter, prospective, randomized, open-label, blinded-endpoint clinical trial undertaken at 48 sites throughout Japan between May 2014 and August 2018. Adults with both asymptomatic hyperuricemia (SUA >7.0 mg/dL) and maximum IMT of the common carotid artery (CCA) ≥1.1 mm at screening were allocated equally using a central web system to receive either dose-titrated febuxostat (10-60 mg daily) or as a control-arm, non-pharmacological lifestyle modification for hyperuricemia, such as a healthy diet and exercise therapy. Of the 514 enrolled participants, 31 were excluded from the analysis, with the remaining 483 people (mean age 69.1 years [standard deviation 10.4 years], female 19.7%) included in the primary analysis (febuxostat group, 239; control group, 244), based on a modified intention-to-treat principal. The carotid IMT images were recorded by a single sonographer at each site and read in a treatment-blinded manner by a single analyzer at a central core laboratory. The primary endpoint was the percentage change from baseline to 24 months in mean IMT of the CCA, determined by analysis of covariance using the allocation adjustment factors (age, gender, history of type 2 diabetes, baseline SUA, and baseline maximum IMT of the CCA) as the covariates. Key secondary endpoints included changes in other carotid ultrasonographic parameters and SUA and the incidence of clinical events. The mean values (± standard deviation) of CCA-IMT were 0.825 mm ± 0.173 mm in the febuxostat group and 0.832 mm ± 0.175 mm in the control group (mean between-group difference [febuxostat - control], -0.007 mm [95% confidence interval (CI) -0.039 mm to 0.024 mm; P = 0.65]) at baseline; 0.832 mm ± 0.182 mm in the febuxostat group and 0.848 mm ± 0.176 mm in the control group (mean between-group difference, -0.016 mm [95% CI -0.051 mm to 0.019 mm; P = 0.37]) at 24 months. Compared with the control group, febuxostat had no significant effect on the primary endpoint (mean percentage change 1.2% [95% CI -0.6% to 3.0%] in the febuxostat group (n = 207) versus 1.4% [95% CI -0.5% to 3.3%] in the control group (n = 193); mean between-group difference, -0.2% [95% CI -2.3% to 1.9%; P = 0.83]). Febuxostat also had no effect on the other carotid ultrasonographic parameters. The mean baseline values of SUA were comparable between the two groups (febuxostat, 7.76 mg/dL ± 0.98 mg/dL versus control, 7.73 mg/dL ± 1.04 mg/dL; mean between-group difference, 0.03 mg/dL [95% CI -0.15 mg/dL to 0.21 mg/dL; P = 0.75]). The mean value of SUA at 24 months was significantly lower in the febuxostat group than in the control group (febuxostat, 4.66 mg/dL ± 1.27 mg/dL versus control, 7.28 mg/dL ± 1.27 mg/dL; mean between-group difference, -2.62 mg/dL [95% CI -2.86 mg/dL to -2.38 mg/dL; P < 0.001]). Episodes of gout arthritis occurred only in the control group (4 patients [1.6%]). There were three deaths in the febuxostat group and seven in the control group during follow-up. A limitation of the study was the study design, as it was not a placebo-controlled trial, had a relatively small sample size and a short intervention period, and only enrolled Japanese patients with asymptomatic hyperuricemia. CONCLUSIONS: In Japanese patients with asymptomatic hyperuricemia, 24 months of febuxostat treatment did not delay carotid atherosclerosis progression, compared with non-pharmacological care. These findings do not support the use of febuxostat for delaying carotid atherosclerosis in this population. TRIAL REGISTRATION: University Hospital Medical Information Network Clinical Trial Registry UMIN000012911.

Uric acid induced hepatocytes lipid accumulation through regulation of miR-149-5p/FGF21 axis.

BACKGROUND: Hyperuricemia is a major risk for non-alcoholic fatty liver disease. However, the mechanisms for this phenomenon are not fully understood. This study aimed to investigate whether microRNAs mediated the pathogenic effects of uric acid on non-alcoholic fatty liver disease. METHODS: Microarray was used to determine the hepatic miRNA expression profiles of male C57BL/6 mice fed on standard chow diet, high fat diet (HFD), and HFD combined with uric acid-lowering therapy by allopurinol. We validated the expression of the most significant differentially expressed microRNAs and explored its role and downstream target in uric acid-induced hepatocytes lipid accumulation. RESULTS: Microarray analysis and subsequent validation showed that miR-149-5p was significantly up-regulated in the livers of HFD-fed mice, while the expression was down-regulated by allopurinol therapy. MiR-149-5p expression was also significantly up-regulated in uric acid-stimulated hepatocytes. Over-expression of miR-149-5p significantly aggregated uric acid-induced triglyceride accumulation in hepatocytes, while inhibiting miR-149-5p ameliorated the triglyceride accumulation. Luciferase report assay confirmed that FGF21 is a target gene of miR-149-5p. Silencing FGF21 abolished the ameliorative effects of miR-149-5p inhibitor on uric acid-induced hepatocytes lipid accumulation, while overexpression of FGF21 prevented the lipid accumulation induced by miR-149-5p mimics. CONCLUSIONS: Uric acid significantly up-regulated the expression of miR-149-5p in hepatocytes and induced hepatocytes lipid accumulation via regulation of miR-149-5p/FGF21 axis.

Uric acid regulates NLRP3/IL-1ß signaling pathway and further induces vascular endothelial cells injury in early CKD through ROS activation and K+ efflux.

BACKGROUND: Chronic kidney disease (CKD) has been considered as a major health problem in the world. Increasing uric acid (UA) could induce vascular endothelial injury, which is closely related to microinflammation, oxidative stress, and disorders of lipids metabolism. However, the specific mechanism that UA induces vascular endothelial cells injury in early CKD remains unknown. METHODS: Human umbilical vein endothelial cells (HUVECs) were cultured and subjected to different concentrations of UA for different periods. Early CKD rat model with elevated serum UA was established. Western blotting and quantitative real-time PCR (qPCR) were applied for measuring protein and mRNA expression of different cytokines. The animals were sacrificed and blood samples were collected for measurement of creatinine, UA, IL-1ß, TNF-α, and ICAM-1. Renal tissues were pathologically examined by periodic acid-Schiff (PAS) or hematoxylin-eosin (HE) staining. RESULTS: The expression of IL-1ß, ICAM-1, NLRP3 complexes, and activation of NLRP3 inflammasome could be induced by UA, but the changes induced by UA were partially reversed by siRNA NLRP3 or caspase 1 inhibitor. Furthermore, we identified that UA regulated the activation of NLRP3 inflammasome by activating ROS and K+ efflux. In vivo results showed that UA caused the vascular endothelial injury by activating NLRP3/IL-1ß pathway. While allopurinol could reduce UA level and may have protective effects on cardiovascular system. CONCLUSIONS: UA could regulate NLRP3/IL-1ß signaling pathway through ROS activation and K+ efflux and further induce vascular endothelial cells injury in early stages of CKD.

Treat to target in gout.

The treat-to-target (T2T) approach has been successfully implemented in a number of diseases. T2T has been proposed for rheumatic diseases such as RA, spondyloarthritis, lupus, and recently for gout. The level of evidence for such approaches differs from one condition to the other (moderate to high for hyperlipidaemia, for example). Practice is based on the best available evidence at any time, and in absence of good evidence for T2T in gout, some suggest a conservative only-treat-symptoms approach. Evidence suggests that not treating gout to target in the long term is overall associated with worsening outcomes, such as flares, tophi and structural damage, which is associated to loss of quality of life and mortality. Different targets have been proposed for hyperuricaemia in gout; lower than 6 mg/dl (0.36 mmol/l) for all patients, at least <5 mg/dl (0.30 mmol/l) for patients with severe-polyarticular or tophaceous-gout.

Novel uricosurics.

Objective: According to recent guidelines, the mainstay of urate-lowering therapies remains xanthine oxidase inhibition. However, some patients with gout show failure to achieve the predefined target of 5-6 mg/dl with xanthine oxidase inhibitors alone, so alternative drugs are needed. The aim of this study was to review studies of novel drugs targeting uric acid transporter 1 (URAT1) and/or other urate transporters for the management of gout. Methods: MeSH terms were used to identify phase 2/3 trials assessing the efficacy of novel uricosurics. A narrative review of novel drugs targeting URAT1 and/or other urate transporters for the management of gout is provided. Results: Lesinurad is a recently approved uricosuric that inhibits URAT1 and the organic ion transporter organic anion transporter 4 (OAT4). Phase 3 trials showed that lesinurad, combined with allopurinol or febuxostat, is a potent urate-lowering therapeutic with an acceptable safety profile. Arhalofenate, another emerging uricosuric, also interacts with URAT1 and organic anion transporter 4. Phase 2 trials suggested that it can both lower serum UA levels and reduce the risk of flares. Conclusions: New drugs inhibiting URAT1 should cover the unmet need for patients with failure to respond or with contraindications to xanthine oxidase inhibitors.

Cardiac and renal protective effects of urate-lowering therapy.

Patients with gout often have co-morbidities such as cardiovascular disease, renal failure and metabolic syndrome components. Some studies, but not all, have suggested that hyperuricaemia and gout are associated with increased risk of myocardial infarction, renal failure and death primarily because of increased risk of cardiovascular events. Therefore, knowledge of the effects of urate-lowering therapy (ULT) on co-morbidities, in particular cardiovascular events and chronic kidney disease, is crucial. Randomized controlled trials (RCTs) have suggested that allopurinol, a xanthine oxidase inhibitor, could improve exercise capacity in patients with chronic stable angina and could decrease blood pressure in adolescents. In contrast, a well-designed RCT found no effect of allopurinol in patients with heart failure. The impact of ULT in patients with chronic kidney disease is unclear. Some RCTs found that allopurinol could slow the decline in kidney function, whereas a recent controlled trial found no benefit of febuxostat. Large randomized placebo-controlled trials are warranted to confirm or not the benefit of ULT on co-morbidities.

Pharmacophore modeling, molecular docking and molecular dynamics studies on natural products database to discover novel skeleton as non-purine xanthine oxidase inhibitors.

Gout is a common inflammatory arthritis caused by the deposition of urate crystals within joints. It is increasingly in prevalence during the past few decades as shown by the epidemiological survey results. Xanthine oxidase (XO) is a key enzyme to transfer hypoxanthine and xanthine to uric acid, whose overproduction leads to gout. Therefore, inhibiting the activity of xanthine oxidase is an important way to reduce the production of urate. In the study, in order to identify the potential natural products targeting XO, pharmacophore modeling was employed to filter databases. Here, two methods, pharmacophore based on ligand and pharmacophore based on receptor-ligand, were constructed by Discovery Studio. Then GOLD was used to refine the potential compounds with higher fitness scores. Finally, molecular docking and dynamics simulations were employed to analyze the interactions between compounds and protein. The best hypothesis was set as a 3D query to screen database, returning 785 and 297 compounds respectively. A merged set of the above 1082 molecules was subjected to molecular docking, which returned 144 hits with high-fitness scores. These molecules were clustered in four main kinds depending on different backbones. What is more, molecular docking showed that the representative compounds established key interactions with the amino acid residues in the protein, and the RMSD and RMSF of molecular dynamics results showed that these compounds can stabilize the protein. The information represented in the study confirmed previous reports. And it may assist to discover and design new backbones as potential XO inhibitors based on natural products.

Effects of Discontinuation of Urate-Lowering Therapy: A Systematic Review.

BACKGROUND: Urate-lowering therapy (ULT) is associated with low rates of adherence, leading to a potential risk of relapse of gouty arthritis, tophi, or urolithiasis. Our main aim was to identify the recurrence of gouty arthritis, tophi, or urolithiasis after discontinuation of ULT. Secondary aims included an assessment of ULT reintroduction rates and factors associated with relapse. METHODS: We conducted a systematic literature review of clinical studies investigating the effect of discontinuing any ULT (allopurinol, febuxostat, probenecid, sulfinpyrazone, benzbromarone) in adults on long-term therapy. We searched The Cochrane Central Register of Controlled Trials, MEDLINE, EMBASE, Science Citation Index, and ClinicalTrials.gov from inception to March 2016. Conference abstracts of the ACR/ARHP and EULAR annual conferences were hand-searched. Study quality was assessed using the first eight items of the methodological index for non-randomized studies (MINORS) tool. The review protocol is registered with PROSPERO (CRD42016042048). RESULTS: A total of 4640 articles were identified, eight of which were ultimately included. Most of these studies predated 2000. MINORS scores ranged from 5 to 10 out of a possible 16. Mean follow-up duration after discontinuation ranged from 12 to 96 months. Five studies focused on discontinuation of ULT in gouty arthritis and tophi, two in urolithiasis, and one in asymptomatic hyperuricemia. Relapse rates were high in gout (36-81%) and lower in urolithiasis (15%). Relapses occurred 1-4.5 years after ULT discontinuation. In one study, a low serum urate level before and after ULT discontinuation was associated with lower gout recurrence. DISCUSSION: Relapse of gout is common although delayed after discontinuation of ULT. Short-term prognosis after ULT discontinuation appears favorable if the serum urate level was low before ULT discontinuation. The results of this review are limited by the paucity of existing studies and their low quality. Further comparative studies should consider larger primary care populations and discontinuation of febuxostat.

Hypouricemic and Nephroprotective Effects of an Active Fraction from Polyrhachis Vicina Roger On Potassium Oxonate-Induced Hyperuricemia in Rats.

BACKGROUND/AIMS: The objective of this study is to evaluate the hypouricemic and nephroprotective effects of an active fraction from Polyrhachis vicina Roger (AFPR) in potassium oxonate-induced hyperuricemic rats. METHODS: Hyperuricemia was induced by potassium oxonate in male rats. AFPR was orally administered to hyperuricemic rats for 12 consecutive weeks. Serum, liver and kidney samples were collected for effects and mechanism analysis. The levels of serum uric acid (SUA) were measured by the phosphotungstic acid method, xanthine oxidase (XOD) activity in the hepatic and serum samples were measured by ultraviolet spectrophotometry, serum levels of interleukin-1 (IL-1ß), interleukin-1 (IL-6) and tumor necrosis factor-α (TNF-α) were measured by ELISA, the levels of serum creatinine (SCr), blood urea nitrogen (BUN), super oxide dismutase (SOD) and malondialdehyde (MDA) in serum were determined by colorimetric method. Protein expression of renal URAT1, GLUT9, and OAT1 were analyzed by Western blot. RESULTS: AFPR significantly decreased the levels of SUA, serum and hepatic XOD, SCr, BUN, and MDA as well as increased SOD. In addition, AFPR treatment significantly reduced the levels of proinflammatory cytokines in serum, including IL-1ß, IL-6 and TNF-α. Moreover, we found the significant decrease in protein expression of URAT1 and GLUT9, and the significant increase in protein expression of OAT1 in the kidney in AFPR treated groups compared to the model groups of hyperuricemia. CONCLUSION: These findings suggest that AFPR has anti-hyperuricemic activity attributed to the inhibition of uric acid generation in the liver and probably to the enhancement of urate excretion in the kidney, and possess nephroprotective effect in hyperuricemic rats due to its anti-inflammatory and antioxidant activities.

Rational use of rasburicase for the treatment and management of tumor lysis syndrome.

Purpose There is a lack of high-level evidence identifying meaningful outcomes and the optimal place in therapy of rasburicase in patients with, or at high risk for tumor lysis syndrome. The primary objective of this study was to evaluate and characterize outcomes resulting from an institution-specific guideline emphasizing supportive care, xanthine oxidase inhibitors, and lower doses of rasburicase. Methods In this retrospective chart review, we compared conservative rasburicase dosing, in accordance with newly developed UMHS tumor lysis syndrome guidelines, with aggressive rasburicase in adult patients (≥ 18 years of age) with hematological or solid tumor malignancies, and a uric acid level between 8 and 15 mg/dL. The primary efficacy outcome assessed the difference in the proportion of patients achieving a uric acid level <8 mg/dL within 48 h using a one-sided noninferiority test. The principle safety outcomes analyzed included incidence of acute kidney injury and hemodialysis requirement. Results One hundred sixty-one patients met inclusion criteria and were included in the study. Within 48 h of an elevated uric acid level, treatment was successful in 97.03% of patients in the conservative group, as compared with 98.33% in the aggressive group (difference, 1.3 percentage points; 95% confidence interval [CI], -3.33 to 5.93). Furthermore, there was no difference in the proportion of patients requiring hemodialysis (2.97% vs. 10.0%, p-value 0.079), or incidence of acute kidney injury (4.0% vs. 12.5%, p-value 1.00) between the treatment group and control group, respectively. Conclusions Conservative rasburicase use was noninferior to aggressive rasburicase use in patients with or at high risk for tumor lysis syndrome.

The Antioxidant and Xanthine Oxidase Inhibitory Activity of Plumeria rubra Flowers.

Plumeria rubra Linn of the family Apocynaceae is locally known in Malaysia as "Kemboja". It has been used by local traditional medicine practitioners for the treatment of arthritis-related disease. The LCMS/MS analysis of the methanol extract of flowers (PR-ME) showed that it contains 3-O-caffeyolquinic acid, 5-caffeoquinic acid, 1,3-dicaffeoquinic acid, chlorogenic acid, citric acid, 3,3-di-O-methylellagic acid, kaempferol-3-O-glucoside, kaempferol-3-rutinoside, kaempferol, quercetin 3-O-α-l-arabinopyranoside, quercetin, quinic acid and rutin. The flower PR-ME contained high amounts of phenol and flavonoid at 184.632 mg GAE/g and 203.2.2 mg QE/g, respectively. It also exhibited the highest DPPH, FRAP, metal chelating, hydrogen peroxide, nitric oxide superoxide radical scavenging activity. Similarly, the XO inhibitory activity in vitro assay possesses the highest inhibition effects at an IC50 = 23.91 µg/mL. There was no mortality or signs of toxicity in rats at a dose of 4 g/kg body weight. The administration of the flower PR-ME at doses of 400 mg/kg to the rats significantly reduced serum uric acid 43.77%. Similarly, the XO activity in the liver was significantly inhibited by flower PR-ME at doses of 400 mg/kg. These results confirm that the flower PR-ME of P. rubra contains active phytochemical compounds as detected in LCMS/MS that contribute to the inhibition of XO activity in vitro and in vivo in reducing acid uric level in serum and simultaneously scavenging the free radical to reduce the oxidative stress.

Camellia sinensis in asymptomatic hyperuricemia: A meta-analysis of tea or tea extract effects on uric acid levels.

Flavanols of Camellia sinensis exhibit uric acid (UA) lowering effect, through the modulation of both xanthine oxidase and urate excretion. In order to investigate the potential benefit of Camellia Sinenis products in asymptomatic hyperuricemia, a meta-analysis of long-term Randomized Controlled Trials (RCT) with tea or tea extract has been conducted. From 20 human intervention studies selected only 5 RCT (13 interventions) were suitable for meta-analysis (n = 472). The current "normal" range set for hyperuricemia fails to identify patients with potential metabolic disorders. Therefore on the basis of the literature data, we fixed cut-off limits for UA baseline levels of 4.5 mg/dl for women, 6.1 mg/dl for men, and 5.5 mg/dl for studies involving mixed populations. Statistically significant effects were not found, but subgroup analysis revealed that the Pooled Estimate effect was different in subjects with baseline levels under [MD (95% CI): 0.1078 (-0.0528 to 0.2684)] and over the cut-off [MD (95% CI): -0.0239 (0.3311 to 0.2833)]. However, due to the low number of RCT and to the lack of data on bioavailability, it is difficult to draw any firm conclusion and more studies are needed to establish if tea flavanols could be useful in asymptomatic hyperuricemia treatment.

Comparative effects of green and black tea extracts on lowering serum uric acid in hyperuricemic mice.

CONTEXT: Tea (Camellia sinensis (L.) Kuntze [Theaceae]) is used to induce urination and inducing nervous excitation. Green and black teas have multifarious physiological functions. The different effects of green and black tea aqueous extracts (GTEs and BTEs) on hyperuricemia are not definitely reported. OBJECTIVE: The different effects of GTEs and BTEs on lowering serum uric acid (UA) in hyperuricemic mice were determined. MATERIALS AND METHODS: Kunming mice were divided into nine groups (n = 6/each group). GTEs and BTEs at the doses of 0.5, 1 and 2 g/kg were orally administrated to mice for seven days, respectively. Hepatic xanthine oxidase (XOD) and adenosine deaminase (ADA) activities as mechanisms of actions were assessed. RESULTS: Research indicated that the LD50 of tea extract is greater than 2 g/kg in mice. UA levels were suppressed significantly with dose-dependent treatment of 0.5, 1 and 2 g/kg BTEs (up to 25.5%, 28.7% and 29.8%, respectively); the serum UA levels were decreased by GTEs but not significant. The activities of XOD and ADA in high dose (2 g/kg) groups of both GTEs and BTEs were notably lower than those of the model group. DISCUSSION AND CONCLUSIONS: The results suggested that both GTEs and BTEs have hypouricaemic and renal protective effects on hyperuricemic mice and the latter one was better. Our study sheds light on the research and development of anti-hyperuricemic functional foods and drugs from tea.

Curcumin Lowers Serum Lipids and Uric Acid in Subjects With Nonalcoholic Fatty Liver Disease: A Randomized Controlled Trial.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is one of the most common hepatic diseases in the general adult population. Dyslipidemia, hyperuricemia, and insulin resistance are common risk factors and accompanying features of NAFLD. Curcumin is a dietary natural product with beneficial metabolic effects relevant to the treatment of NAFLD. AIM: To assess the effects of curcumin on metabolic profile in subjects with NAFLD. METHODS: Patients diagnosed with NAFLD (grades 1-3; according to liver sonography) were randomly assigned to curcumin (1000 mg/d in 2 divided doses) (n = 50) or control (n = 52) group for a period of 8 weeks. All patients received dietary and lifestyle advises before the start of trial. Anthropometric measurements, lipid profile, glucose, insulin, glycated hemoglobin, and uric acid concentrations were measured at baseline and after 8 weeks of follow-up. RESULTS: Eighty-seven subjects (n = 44 and 43 in the curcumin and control group, respectively) completed the trial. Supplementation with curcumin was associated with a reduction in serum levels of total cholesterol (P < 0.001), low-density lipoprotein cholesterol (P < 0.001), triglycerides (P < 0.001), non-high-density lipoprotein cholesterol (P < 0.001), and uric acid (P < 0.001), whereas serum levels of high-density lipoprotein cholesterol and glucose control parameters remained unaltered. Curcumin was safe and well tolerated during this study. CONCLUSION: Results of the present trial suggest that curcumin supplementation reduces serum lipids and uric acid concentrations in patients with NAFLD.

A new sensitive and quantitative chemiluminescent assay to monitor intracellular xanthine oxidase activity for rapid screening of inhibitors in living endothelial cells.

Xanthine oxidase (XO) is an important enzyme, expressed at high levels in the vasculature in endothelial cells, that catalyzes the hydroxylation of hypoxanthine to xanthine and xanthine to uric acid. Excessive production of uric acid results in hyperuricemia linked to gout and cardiovascular diseases. Testing inhibition of XO is important for detection of potentially effective drugs or natural products that could be used to treat diseases caused by increased XO activity. In the present study, for the first time, we developed an in vitro chemiluminescent bioassay to determine XO activity in living endothelial cells and the IC50 value of oxypurinol, the active metabolite of the inhibitor drug allopurinol. Intracellular XO activity was measured in less than 20 min with a luminol/catalyst-based chemiluminescence assay able to measure XO with a limit of 0.4 µU/mL. Oxypurinol addition to 5 × 103 cells (ranging from 5.0 to 0.0 µM) caused a linear decrease in XO activity, with an IC50 of 1.0 ± 0.5 µM. The detection system developed was low-cost, rapid, reproducible, and easily miniaturizable so suitable to be used on small quantities of cells.

Inhibition of sterile danger signals, uric acid and ATP, prevents inflammasome activation and protects from alcoholic steatohepatitis in mice.

BACKGROUND & AIMS: The inflammasome is a well-characterized inducer of inflammation in alcoholic steatohepatitis (ASH). Inflammasome activation requires two signals for mature interleukin (IL)-1ß production. Here we asked whether metabolic danger signals trigger inflammasome activation in ASH. METHODS: Wild-type mice, ATP receptor 2x7 (P2rx7)-KO mice, or mice overexpressing uricase were fed Lieber-DeCarli ethanol or control diet. We also implemented a pharmacological approach in which mice were treated with probenecid or allopurinol. RESULTS: The sterile danger signals, ATP and uric acid, were increased in the serum and liver of alcohol-fed mice. Depletion of uric acid or ATP, or lack of ATP signaling attenuated ASH and prevented inflammasome activation and its major downstream cytokine, IL-1ß. Pharmacological depletion of uric acid with allopurinol provided significant protection from alcohol-induced inflammatory response, steatosis and liver damage, and additional protection was achieved in mice treated with probenecid, which depletes uric acid and blocks ATP-induced P2rx7 signaling. We found that alcohol-damaged hepatocytes released uric acid and ATP in vivo and in vitro and that these sterile danger signals activated the inflammasome in LPS-exposed liver mononuclear cells. CONCLUSIONS: Our data indicate that the second signal in inflammasome activation and IL-1ß production in ASH results from the endogenous danger signals, uric acid and ATP. Inhibition of signaling triggered by uric acid and ATP may have therapeutic implications in ASH.

Design and synthesis of novel 2-(indol-5-yl)thiazole derivatives as xanthine oxidase inhibitors.

Xanthine oxidase (XO) inhibitors have been widely used for the treatment of gout. Indole rings are frequently used as active scaffold in designing inhibitors for enzymes. Herein, we describe the structure-activity relationship for novel xanthine oxidase inhibitors based on indole scaffold. A series of novel tri-substituted 2-(indol-5-yl)thiazole derivatives were synthesized, and their in vitro inhibitory activities against xanthine oxidase and in vivo efficacy lowering uric acid level in blood were measured. Among them, 2-(3-cyano-2-isopropylindol-5-yl)-4-methylthiazole-5-carboxylic acid exhibits the most potent XO inhibitory activity (IC50 value: 3.5nM) and the excellent plasma uric acid lowering activity. Study of structure activity relationship indicated that hydrophobic moiety (e.g., isopropyl) at 1-position and electron withdrawing group (e.g., CN) at 3-position of indole ring and small hydrophobic group (CH3) at 4-position of the thiazole ring enhanced the XO inhibitory activity. Hydrophobic substitution such as isopropyl at 1-position of the indole moiety without any substitution at 2-position has an essential role for enhancing bioavailability and therefore for high in vivo efficacy.

Recent advances in gout drugs.

Gout is an autoinflammatory disease caused by the deposition of urate crystals. As the most common inflammatory arthritis, gout has a high incidence and can induce various severe complications. At present, there is no effective cure method in the world. With the deepening of medical research, gout treatment drugs continue to progress. In this review, we provide a landscape view of the current state of the research on gout treatment drugs, including the research progress of anti-inflammatory and analgesic drugs, drugs that promote uric acid excretion, and drugs that inhibit uric acid production. We mainly emphasize the understanding of gout as an auto-inflammatory disease and the discovery strategy of related gout drugs to provide a systematic and theoretical basis for the new exploration of gout drug discovery.

The efficacy and safety of febuxostat for urate lowering in gout patients ≥65 years of age.

BACKGROUND: The incidence of gout rises with increasing age. Management of elderly (≥65 years) gout patients can be challenging due to high rates of comorbidities, such as renal impairment and cardiovascular disease, and concomitant medication use. However, there is little data specifically addressing the efficacy and safety of available urate-lowering therapies (ULT) in the elderly. The objective of this post hoc analysis was to examine the efficacy and safety of ULT with febuxostat or allopurinol in a subset of elderly subjects enrolled in the CONFIRMS trial. METHODS: Hyperuricemic (serum urate [sUA] levels ≥ 8.0 mg/dL) gout subjects were enrolled in the 6-month, double-blind, randomized, comparative CONFIRMS trial and randomized, 1:1:1, to receive febuxostat, 40 mg or 80 mg, or allopurinol (200 mg or 300 mg based on renal function) once daily. Flare prophylaxis was provided throughout the study duration.Study endpoints were the percent of elderly subjects with sUA <6.0 mg/dL at the final visit, overall and by renal function status, percent change in sUA from baseline to final visit, flare rates, and rates of adverse events (AEs). RESULTS: Of 2,269 subjects enrolled, 374 were elderly. Febuxostat 80 mg was significantly more efficacious (82.0%) than febuxostat 40 mg (61.7%; p < 0.001) or allopurinol (47.3%; p < 0.001) for achieving the primary efficacy endpoint. Febuxostat 40 mg was also superior to allopurinol in this population (p = 0.029). In subjects with mild-to-moderate renal impairment, significantly greater ULT efficacy was observed with febuxostat 40 mg (61.6%; p = 0.028) and febuxostat 80 mg (82.5%; p < 0.001) compared to allopurinol 200/300 mg (46.9%). Compared to allopurinol 200/300 mg, the mean percent change in sUA from baseline was significantly greater for both febuxostat 80 mg (p < 0.001) and febuxostat 40 mg (p = 0.011) groups. Flare rates declined steadily in all treatment groups. Rates of AEs were low and comparable across treatments. CONCLUSIONS: These data suggest that either dose of febuxostat is superior to commonly prescribed fixed doses of allopurinol (200/300 mg) in subjects ≥65 years of age with high rates of renal dysfunction. In addition, in this high-risk population, ULT with either drug was well tolerated. TRIAL REGISTRATION: clinicaltrials.gov NCT#00430248.