Identification of Human UDP-Glucuronosyltransferase and Sulfotransferase as Responsible for the Metabolism of Dotinurad, a Novel Selective Urate Reabsorption Inhibitor.

Dotinurad, a novel selective urate reabsorption inhibitor, is used to treat hyperuricemia. In humans, orally administered dotinurad is excreted mainly as glucuronide and sulfate conjugates in urine. To identify the isoforms of UDP-glucuronosyltransferase (UGT) and sulfotransferase (SULT) involved in dotinurad glucuronidation and sulfation, microsome and cytosol fractions of liver, intestine, kidney, and lung tissues (cytosol only) were analyzed along with recombinant human UGT and SULT isoforms. Dotinurad was mainly metabolized to its glucuronide conjugate by human liver microsomes (HLMs), and the glucuronidation followed the two-enzyme Michaelis-Menten equation. Among the recombinant human UGT isoforms expressed in the liver, UGT1A1, UGT1A3, UGT1A9, and UGT2B7 catalyzed dotinurad glucuronidation. Based on inhibition analysis using HLMs, bilirubin, imipramine, and diflunisal decreased glucuronosyltransferase activities by 45.5%, 22.3%, and 22.2%, respectively. Diflunisal and 3'-azido-3'-deoxythymidine, in the presence of 1% bovine serum albumin, decreased glucuronosyltransferase activities by 21.1% and 13.4%, respectively. Dotinurad was metabolized to its sulfate conjugate by human liver cytosol (HLC) and human intestinal cytosol (HIC) samples, with the sulfation reaction in HLC samples following the two-enzyme Michaelis-Menten equation and that in HIC samples following the Michaelis-Menten equation. All eight recombinant human SULT isoforms used herein catalyzed dotinurad sulfation. Gavestinel decreased sulfotransferase activity by 15.3% in HLC samples, and salbutamol decreased sulfotransferase activity by 68.4% in HIC samples. These results suggest that dotinurad glucuronidation is catalyzed mainly by UGT1A1, UGT1A3, UGT1A9, and UGT2B7, whereas its sulfation is catalyzed by many SULT isoforms, including SULT1B1 and SULT1A3. SIGNIFICANCE STATEMENT: The identification of enzymes involved in drug metabolism is important to predicting drug-drug interactions (DDIs) and interindividual variability for safe drug use. The present study revealed that dotinurad glucuronidation is catalyzed mainly by UGT1A1, UGT1A3, UGT1A9, and UGT2B7 and that its sulfation is catalyzed by many SULT isoforms, including SULT1B1 and SULT1A3. Therefore, dotinurad, a selective urate reabsorption inhibitor, is considered safe for use with a small risk of DDIs and low interindividual variability.

Hypouricemic agents reduce indoxyl sulfate excretion by inhibiting the renal transporters OAT1/3 and ABCG2.

Indoxyl sulfate (IS) accumulates in the body in chronic kidney disease (CKD). In the renal proximal tubules, IS excretion is mediated by OAT1/3 and ABCG2. These transporters are inhibited by some hypouricemic agents; OATs by probenecid and benzbromarone, ABCG2 by febuxostat and benzbromarone. Thus, we evaluated whether hypouricemic agents including dotinurad, a novel selective urate reabsorption inhibitor with minimal effect on OATs or ABCG2, affect IS clearance in rats. Intact and adenine-induced acute renal failure rats were orally administered hypouricemic agents, and both endogenous IS and exogenously administered stable isotope-labeled d4-IS in the plasma and kidney were measured. Our results demonstrated that OATs inhibitors, such as probenecid, suppress IS uptake into the kidney, leading to increased plasma IS concentration, whereas ABCG2 inhibitors, such as febuxostat, cause renal IS accumulation remarkably by suppressing its excretion in intact rats. The effects of these agents were reduced in adenine-induced acute renal failure rats, presumably due to substantial decrease in renal OAT1/3 and ABCG2 expression. Dotinurad did not significantly affected the clearance of IS under both conditions. Therefore, we suggest that hypouricemic agents that do not affect OATs and ABCG2 are effective therapeutic options for the treatment of hyperuricemia complicated by CKD.

Ideal pharmacokinetic profile of dotinurad as a selective urate reabsorption inhibitor.

Dotinurad, a novel selective urate reabsorption inhibitor (SURI), has potent inhibitory effects at low doses on the uptake of urate by urate transporter 1 (URAT1, solute carrier family 22 member 12 [SLC22A12]), localized at the apical membrane of renal proximal tubular cells. This study sought to clarify the pharmacokinetic (PK) profile of dotinurad. In rats, monkeys, and humans, the apparent distribution volume (0.257, 0.205, and 0.182 L/kg, respectively) and oral clearance (0.054, 0.037, and 0.013 L·h-1·kg-1, respectively) of dotinurad were very low, whereas plasma and luminal concentrations were adequately maintained at high levels. In addition, species differences were scarcely observed with plasma protein binding of 99.4%. The main metabolite was dotinurad glucuronide (no specific metabolites in humans), and percentage excretion of unchanged dotinurad was low in all the investigated species. The risk of drug interaction with dotinurad was expected to be low, because it weakly inhibits metabolic enzymes such as cytochrome P450 (CYP). In conclusion, low-dose dotinurad exhibited excellent pharmacological effects as well as ideal PK properties as a SURI.

Pharmacokinetic/pharmacodynamic modeling and simulation of dotinurad, a novel uricosuric agent, in healthy volunteers.

This study aimed to investigate the pharmacokinetic and pharmacodynamic (PK/PD) profiles of dotinurad, a novel uricosuric agent, and to construct a PK/PD model to predict serum urate (SUA) levels after dotinurad administration in healthy men. PK/PD model was constructed using single-dose study data considering the physiological features of urate handling. Model validation was performed by comparing the predicted SUA levels with the SUA levels in a multiple-dose study. Dotinurad was absorbed rapidly, and its exposure increased proportionally in the tested dose ranges (0.5-20 mg) after a single-dose administration. The PK model after oral administration was described using a one-compartment model with first-order absorption. Effects on SUA and renal urate excretion of dotinurad increased with dose escalation but were apparently saturable at a dose >5 mg. The simple maximal effect (Emax) model was selected as the PD model of dotinurad on renal urate reabsorption, resulting in an estimated Emax of 0.51. The plasma concentration at the half-maximal effect of dotinurad was 196 ng/mL. Other PD parameters were calculated from the change in SUA level or urinary excretion of urate before and after dotinurad administration. The predicted SUA levels, using the PK/PD model, were well-fitted with the observed values. The constructed PK/PD model of dotinurad appropriately described the profiles of dotinurad plasma concentrations and SUA level in multiple administration study.

Characterization of N-glucuronidation of 4-(5-pyridin-4-yl-1H-[1,2,4]triazol-3-yl) pyridine-2-carbonitrile (FYX-051): a new xanthine oxidoreductase inhibitor.

In humans, orally administered 4-(5-pyridin-4-yl-1H-[1,2,4]triazol-3-yl) pyridine-2-carbonitrile (FYX-051) is excreted mainly as triazole N(1)- and N(2)-glucuronides in urine. It is important to determine the enzyme(s) that catalyze the metabolism of a new drug to estimate individual differences and/or drug-drug interactions. Therefore, the characterization and mechanism of these glucuronidations were investigated using human liver microsomes (HLMs), human intestinal microsomes (HIMs), and recombinant human UDP-glucuronosyltransferase (UGT) isoforms to determine the UGT isoform(s) responsible for FYX-051 N(1)- and N(2)-glucuronidation. FYX-051 was metabolized to its N(1)- and N(2)-glucuronide forms by HLMs, and their K(m) values were 64.1 and 72.7 microM, respectively; however, FYX-051 was scarcely metabolized to its glucuronides by HIMs. Furthermore, among the recombinant human UGT isoforms, UGT1A1, UGT1A7, and UGT1A9 catalyzed the N(1)- and N(2)-glucuronidation of FYX-051. To estimate their contribution to FYX-051 glucuronidation, inhibition analysis with pooled HLMs was performed. Mefenamic acid, a UGT1A9 inhibitor, decreased FYX-051 N(1)- and N(2)-glucuronosyltransferase activities, whereas bilirubin, a UGT1A1 inhibitor, did not affect these activities. Furthermore, in the experiment using microsomes from eight human livers, the N(1)- and N(2)-glucuronidation activity of FYX-051 was found to significantly correlate with the glucuronidation activity of propofol, a specific substrate of UGT1A9 (N(1): r(2) = 0.868, p < 0.01; N(2): r(2) = 0.775, p < 0.01). These results strongly suggested that the N(1)- and N(2)-glucuronidation of FYX-051 is catalyzed mainly by UGT1A9 in human livers.

Metabolic profile of FYX-051 (4-(5-pyridin-4-yl-1h-[1,2,4]triazol-3-yl)pyridine-2-carbonitrile) in the rat, dog, monkey, and human: identification of N-glucuronides and N-glucosides.

FYX-051, 4-(5-pyridin-4-yl-1H-[1,2,4]triazol-3-yl)pyridine-2-carbonitrile, is a novel xanthine oxidoreductase inhibitor that can be used for the treatment of gout and hyperuricemia. We examined the metabolism of FYX-051 in rats, dogs, monkeys, and human volunteers after the p.o. administration of this inhibitor. The main metabolites in urine were pyridine N-oxide in rats, triazole N-glucoside in dogs, and triazole N-glucuronide in monkeys and humans, respectively. Furthermore, N-glucuronidation and N-glucosidation were characterized by two types of conjugation: triazole N(1)- and N(2)-glucuronidation and N(1)- and N(2)-glucosidation, respectively. N(1)- and N(2)-glucuronidation was observed in each species, whereas N(1)- and N(2)-glucosidation was mainly observed in dogs. With regard to the position of conjugation, N(1)-conjugation was predominant; this resulted in a considerably higher amount of N(1)-conjugate in each species than N(2)-conjugate. The present results indicate that the conjugation reaction observed in FYX-051 metabolism is unique, i.e., N-glucuronidation and N-glucosidation occur at the same position of the triazole ring, resulting in the generation of four different conjugates in mammals. In addition, a urinary profile of FYX-051 metabolites in monkeys and humans was relatively similar; triazole N-glucuronides were mainly excreted in urine.