A New Uricosuric Diuretic, S-8666, in Rats and Chimpanzees Yukio YONETANI, Kazumi IWAKI, Toshihiro SHINOSAKI, Atsuko KAWASE-HANA

A New Uricosuric Diuretic, S-8666, in Rats and Chimpanzees Yukio YONETANI, Kazumi IWAKI, Toshihiro SHINOSAKI, Atsuko KAWASE-HANAFUSA, Hiroshi HARADA and Arthur A. van ES* ShionogiResearch Laboratories, Shionogi & Co., Ltd., Fukushima-ku,Osaka 553, Japan *Primate Center TNO , Lange Kleiweg151, 2288 GJ Rijswijk,The Netherlands AcceptedJanuary 6, 1987 Abstract-5 Dimethylsulfamoyl-6,7-dichloro-2,3-dihydrobenzofuran-2-carboxylic acid (S-8666) was studied as a possible new uricosuric diuretic agent using rats and chimpanzees. Various new compounds belonging to the 5-sulfamoyl-6,7 dichloro-2,3-dihydrobenzofuran-2-carboxylic acids were clearly diuretic with uricosuric activity in intraperitoneally oxonate-treated rats. S-8666 was chosen as a favorable candidate because its uricosuric activity due to the effects of tubular transport of uric acid were apparently more marked than those of known uricosuric agents such as probenecid, benzbromarone, tienilic acid and indacrinone in oxonate treated rats. S-8666 was also uricosuric in rats not given urate oxidase inhibitor. The diuretic effect of S-8666 in oxonate-treated rats was as high-ceilinged as that of furosemide, while those of tienilic acid, indacrinone and a known compound of a 5-carbonyl-6,7-dichloro-2,3-dihydrobenzofuran-2-carboxylic acid were rather low-ceilinged. These uricosuric and diuretic activities of S-8666 were manifested by two enantiomers, of which the (+)-enantiomer displayed predominantly uricosuric activity and the (-)-enantiomer, diuretic activity like furosemide. The new compound was also uricosuric and diuretic in chimpanzees, although the potency of the uricosuric activity was similar to that of probenecid and less than that of indacrinone. Thus, it seems that S-8666 is a different type of uricosuric diuretic from known agents which have already been tried in humans. Diuretic thiazides and various loop excretion of uric acid. To help alleviate this diuretics have been generally utilized as problem, we recently devised a practical useful antihypertensive agents, but the method employing a commonly used experi hyperuricemic effect incident to their major mental animal, the rat, which was treated effects has sometimes necessitated their with an inhibitor of urate oxidase, potassium withdrawal (1-3). Uricosuric (generally used oxonate (11). Rats are different from primates to mean 'hyperuricosuric') diuretics have in how they metabolize uric acid to allantoin been studied over the past decade to im in the liver, but the net flux of uric acid in prove diuretic antihypertensives. the renal tubules is clearly reabsorptive (12). At first, tienilic acid (4) was considered to Therefore, the animals have sometimes been be an excellent uricosuric diuretic, which used to support the uricosuric activity of could lower the blood level of uric acid test compounds (13, 14). However, the during medication, but its hepatotoxicity results do not always demonstrate the effect forced it to be banned from clinical use (5). of test compounds on the renal clearance of This accelerated further studies on a new uric acid under experimental conditions using generation of diuretic antihypertensives (6 animals displaying both hepatic metabolism 10), but their development as drugs has been and renal excretion of uric acid, and hence delayed because the evaluation of uricosuric the animals are not feasible for evaluating the activity requires tests on primates, due to uricosuric activity of new compounds. On species differences in the metabolism and the other hand, oxonate-treated rats also had a reabsorptive net flux of uric acid in the renal collected. Plasma was separated immediately tubules and responded well to various known after the blood collection using a Beckman agents which had already been demonstrated Microfuge. Plasma concentrations of uric to have effects on uric acid excretion in acid and inulin paired with those in the urine humans, although all responses were not sample were assessed as the average of their always identical to those in primates. In values at the beginning and end of the urine addition, an experiment using rats is much collection. Test compounds were adminis easier than that using primates. It thus seems tered intraperitoneally just after the first urine that the clearance experiment using oxonate collection as a suspension in 1 % gum arabic treated rats is feasible for the first screening solution. Experiments using oxonate of favorable candidates from various new untreated rats were done according to the compounds. In the present study, 5-dimethyl method described previously (15). Fourteen sulfamoyl-6,7-dichloro-2,3-dihydrobenzo week-old male SIc-Wistar rats were infused fu ran -2 carboxyl ic acid (S-8666) was with 0.26% sodium pentobarbital-4% studied as a favorable candidate for a mannitol-1.5% inulin-0.25% sodium chloride uricosuric diuretic after being chosen from solution at a flow rate of 0.03 ml/min after a screening with the animals, and then it was the cannulation. After the equilibration for confirmed to also be uricosuric and diuretic 60 min, continuous 20-min urine samples in chimpanzees*. were collected four times, and blood was collected at the midpoint of each urine Materials and Methods collection period. In these experiments, uric Chemicals: Probenecid (Sigma), furose acid was determined by the fluorometric mide (Sigma) and furosemide solution for method, basically according to the procedure injection (LASIX, Hoechst) were com of Sum] et al. (16), while inulin was estimated mercial preparations. The other compounds according to the method of Vurek and Pegram used were prepared in our laboratories. (17). Experiments in rats: Uricosuric activity in Data are given as the mean±standard oxonate-treated rats was determined by the error, and the significance of the difference method described previously (11); ten-week from the level before dosing was evaluated old male SIc-Wistar rats which had received by Student's t-test. 250 mg/kg potassium oxonate i.p. were The abbreviations used here are UuaV, cannulated in the right femoral artery, left urine-excreted amounts of uric acid; Fua, femoral vein and urinary bladder under glomerular filtered amounts of uric acid; anesthesia with sodium pentobarbital within FEua, fractional excretion of uric acid; and 2 hr after dosing with oxonate. The same dose Cin, inulin clearance. of oxonate, i.p.; 2 ml/kg of 60% urethane, Experiments in chimpanzees: Three 10 s.c.; and 4 ml/kg of 15% inulin, s.c., were year-old male laboratory-born chimpanzees given just 2 hr after the first administration (Pan troglocytes) weighing 55 to 65 kg were of oxonate, and then the animals were infused used. Clearance experiments were performed with 4% mannitol-1.5% inulin-0.9% sodium basically according to the commonly used chloride solution at the flow rate of 0.1 ml/ procedure under anesthesia (18); the animals min on a hot plate at 30°C. After the were anesthetized by the premedication of equilibration for 40 min, 0.2 ml of arterial ketamine hydrochloride, 10 mg/kg, 1.m., and blood was collected six times at 20-min atropine sulfate, 0.05 mg/kg, i.m., and then intervals, and five 20-min urine samples were the introductory and maintenance anesthesia with oxygen-nitrous oxide-halothane mixture. * The clearance experiments using chimpanzees The clearance experiments were started with were performed in the Netherlands Organization an intravenous infusion of Ringer-lactate for Applied Scientific Research, TNO, supervised solution at a flow rate of 3 ml/min, a bolus by Dr. A.A. van Es and directed by Dr. P.M.C.A. administration of inulin, 50 mg/kg, i.v., and van Eerd under the sponsorship of Shionogi an intravenous infusion of 1 mg/kg/min Research Laboratories. inulin in saline as a sustaining dose. Blood and urine samples were collected every 30 treated rats ranged from 0.5 to 0.7, which min. After equilibration for 120 min, the test was apparently higher than those in humans compound dissolved into 1.26% sodium and chimpanzees, and meant that the net bicarbonate solution was administered orally flux of uric acid in the renal tubules in the by a stomach tube; and thereafter, the animals was definitely reabsorptive. Therefore, infusion volume of the Ringer-lactate solution it should be possible to evaluate the increase was adjusted to the urine flow to prevent of the FEua value in the animals due to volume loss in the animals after the adminis inhibition of the tubular transport of uric tration of diuretic agents. acid. The uricosuric and diuretic effects of Determination of S-8666 and the metab various compounds which have been well olite, 5 monomethylsulfamoyl-6,7-dichloro known for their effects on uric acid excretion 2,3 dihydrobenzofuran 2 carboxylic acid: in primates are summarized in Table 1. Part of the plasma and urine samples in the Experiments on each drug were performed clearance studies using chimpanzees was with 8 to 10 animals, of which part of the used to determine the concentrations of S experimental results had been reported 8666 and the metabolite to confirm the previously (11). The increases of urine reliability of the clearance experiments. The excreted amounts of uric acid (UuaV), plasma sample (0.5 ml) was mixed and glomerular-filtered amounts of uric acid shaken well with 1.0 ml of 1 N NaOH and (Fua), FEua and urine volume were calculated 2.0 ml of dichloromethane containing 20 /cg using the mean values in each experimental of 5-(pyrrolidin-1-yl-sulfonyl)-6,7-dichloro period, when the values were significantly 2,3-dihydrobenzofuran-2-carboxylic acid as different from the level before dosing with an internal standard. the drug at P<0.05, and then the increases After centrifuging, 1.0 ml of the aqueous were shown as the average value for 80 min phase was mixed and shaken with 1.0 ml after the dosing.

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