Data mining-guided alleviation of hyperuricemia by Paeonia veitchii Lynch through inhibition of xanthine oxidase and regulation of renal urate transporters.

BACKGROUND: Hyperuricemia (HUA) is a metabolic disease characterized by a high level of uric acid (UA). The extensive historical application of traditional Chinese medicine (TCM) offers a range of herbs and prescriptions used for the treatment of HUA-related disorders. However, the core herbs in the prescriptions and their mechanisms have not been sufficiently explained. PURPOSE: Our current investigation aimed to estimate the anti-HUA effect and mechanisms of Paeonia veitchii Lynch, an herb with high use frequency identified from data mining of TCM prescriptions. METHODS: Prescriptions for HUA/gout treatment were statistically analyzed through a data mining approach to determine the common nature and use frequency of their composition herbs. The chemical constituents of Paeonia veitchii extract (PVE) were analyzed by UPLC-QTOF-MS/MS, while its UA-lowering effect was further evaluated in adenosine-induced liver cells and potassium oxonate (PO) and hypoxanthine (HX)-induced HUA mice. RESULTS: A total of 225 prescriptions involving 246 herbs were sorted out. The properties, flavors and meridians of the appearing herbs were mainly cold, bitter and liver, respectively, while their efficacy was primarily concentrated on clearing heat and dispelling wind. Further usage frequency analysis yielded the top 20 most commonly used herbs, in which PVE presented significant inhibitory activity (IC50 = 131.33 µg/ml) against xanthine oxidase (XOD), and its constituents showed strong binding with XOD in a molecular docking study and further were experimentally validated through XOD enzymatic inhibition and surface plasmon resonance (SPR). PVE (50 to 200 µg/ml) dose-dependently decreased UA levels by inhibiting XOD expression and activity in BRL 3A liver cells. In HUA mice, oral administration of PVE exhibited a significant UA-lowering effect, which was attributed to the reduction of UA production by inhibiting XOD activity and expression, as well as the enhancement of UA excretion by regulating renal urate transporters (URAT1, GLUT9, OAT1 and ABCG2). Noticeably, all doses of PVE treatment did not cause any liver injury, and displayed a renal protective effect. CONCLUSIONS: Our results first comprehensively clarified the therapeutic effect and mechanisms of PVE against HUA through suppressing UA production and promoting UA excretion with hepatic and renal protection, suggesting that PVE could be a promising UA-lowering candidate with a desirable safety profile for the treatment of HUA and prevention of gout.

Identification of natural xanthine oxidase inhibitors: Virtual screening, anti-xanthine oxidase activity, and interaction mechanism.

Xanthine oxidase (XO) is a crucial target for hyperuricemia treatment(s). Naturally occurred XO inhibitors with minimal toxicity and high efficacy have attracted researchers' attention. With the goal of quickly identifying natural XO inhibitors, an integrated computational screening strategy was constructed by molecular docking and calculating the free energy of binding. Twenty-seven hits were achieved from a database containing 19,377 natural molecules. This includes fourteen known XO inhibitors and four firstly-reported inhibitors (isolicoflavonol, 5,7-dihydroxycoumarin, parvifolol D and clauszoline M, IC50 < 40 µM). Iolicoflavonol (hit 8, IC50 = 8.45 ± 0.68 µM) and 5,7-dihydroxycoumarin (hit 25, IC50 = 10.91 ± 0.71 µM) displayed the great potency as mixed-type inhibitors. Docking study and molecular dynamics simulation revealed that both hits could interact with XO's primarily active site residues ARG880, MOS1328, and ASN768 of XO. Fluorescence spectroscopy studies showed that hit 8 bound to the active cavity region of XO, causing changes in XO's conformation and hydrophobicity. Hits 8 and 25 exhibit favorable Absorption, Distribution, Metabolism, and Excretion (ADME) properties. Additionally, no cytotoxicity against human liver cells was observed at their median inhibition concentrations against XO. Therefore, the present study offers isolicoflavonol and 5,7-dihydroxycoumarin with the potential to be disease-modifying agents for hyperuricemia.

Unveiling the inhibitory mechanism of aureusidin targeting xanthine oxidase by multi-spectroscopic methods and molecular simulations.

Xanthine oxidase (XO) is a key target for gout treatment. Great efforts have been made towards the discovery and development of new XO inhibitors. Aureusidin (AUR), a natural compound, emerges as the second reported XO inhibitor with an aurone skeleton with an IC50 value of 7.617 ± 0.401 µM in vitro. The inhibitory mechanism of AUR against XO was explored through enzyme kinetic studies, multi-spectroscopic methods, computer simulation techniques, and ADME prediction. The results showed that AUR acts as a rapid reversible and mixed-type XO inhibitor and its binding to XO was driven by hydrogen bonding and hydrophobic interaction. Moreover, AUR presented a strong fluorescence quenching effect through a static quenching process and induced a conformation change of XO. Its binding pattern with XO was revealed through molecular docking, and its affinity toward XO was enhanced through interactions with key amino acid residues in the active pocket of XO. Further, AUR demonstrated good stability and pharmacokinetic behavior properties in molecular dynamics simulation and ADME prediction. In short, the current work clarified in depth the inhibitory mechanism of AUR on XO firstly and then provided fresh insights into its further development as a natural potent XO inhibitor with aurone skeleton.

Chalcone derivatives as xanthine oxidase inhibitors: synthesis, binding mode investigation, biological evaluation, and ADMET prediction.

Xanthine oxidase (XO) is a crucial target for the treatment of hyperuricemia and gout. A series of derivatives based on natural 3,4-dihydroxychalcone, obtained from Carthamus tinctorious and Licorice, were designed and synthesized. Nine derivatives (9a-e, 10b,c, and 15a,b) exhibited apparent XO inhibitory activity in vitro (IC50 values varied from 0.121 to 7.086 µM), 15b presented the most potent inhibitory activity (IC50 = 0.121 µM), which was 27.47-fold higher than that of allopurinol (IC50 = 3.324 µM). The SAR analysis indicated that introducing hydroxyl groups at 3'/4'/5'-position on ring A was more beneficial to the inhibition of XO than at 2'/6'-position; the removal of 3­hydroxyl group on ring B could weaken the inhibitory potency of hydroxychalcones on XO, but it was beneficial to the XO inhibitory potency of methoxychalcones. Molecule modeling studies afforded insights into the binding mode of 15b with XO and supported the findings of SAR analysis. Additionally, kinetics studies demonstrated that 15b presented a reversible and competitive XO inhibitor, which spontaneously combined with XO through hydrophobic force, and finally changed the secondary conformation of XO. Furthermore, the acute hyperuricemia model was employed to investigate the hypouricemic effect of 15b, which could effectively reduce the serum uric acid levels of rats at an oral dose of 10 mg/kg. ADMET prediction suggested that compound 15b possessed good pharmacokinetic properties. Briefly, compound 15b emerges as an interesting XO inhibitor for the treatment of hyperuricemia and gout with beneficial effects on serum uric acid levels regulating. Meanwhile, the XO inhibitors with chalcone skeleton will deserve further attention and discussion.