Inhibition of bone resorption by the cathepsin K inhibitor odanacatib is fully reversible.

The cathepsin K (CatK) inhibitor odanacatib (ODN) is currently being developed for the treatment of osteoporosis. In clinical trials, efficacy and resolution of effect of ODN treatment on bone turnover biomarkers and accrued bone mass have been demonstrated. Here, we examine the effects of continuing treatment and discontinuation of ODN versus alendronate (ALN) on osteoclast (OC) function. First, accessibility and reversible engagement of active CatK in intracellular vesicles and resorption lacunae of actively resorbing OCs were demonstrated by the selective and reversible CatK inhibitors, BODIPY-L-226 (IC50=39nM) and L-873,724 (IC50=0.5nM). Next, mature human OCs on bone slices were treated with vehicle, ODN, or ALN for 2days, followed by either continuing with the same treatment, or replacement of the inhibitors by vehicle for additional times as specified per experimental conditions. Maintaining OCs on ODN or ALN significantly reduced CTx-I release compared to vehicle controls. However, only the treatment of OCs with ODN resulted in the formation of small shallow discrete resorption pits, retention of intracellular vesicles enriched with CatK and other lysosomal enzymes, increase in 1-CTP release and number of TRAP(+) OCs. Upon discontinuation of ODN treatment, OCs rapidly resumed bone resorption activity, as demonstrated by a return of OC functional markers (CTx-I, 1-CTP), cell number and size, morphology and number of resorption pits, and vesicular secretion of CatK toward the respective vehicle levels. As expected, discontinuation of ALN did not reverse the treatment-related inhibition of OC activity in the time frame of the experiment. In summary, this study demonstrated rapid kinetics of inhibition and reversibility of the effects of ODN on OC bone resorption, that differentiated the cellular mechanism of CatK inhibition from that of the bisphosphate antiresorptive ALN.

Synthesis, characterization, and activity of metabolites derived from the cyclooxygenase-2 inhibitor rofecoxib (MK-0966, Vioxx).

Metabolites of the COX-2 inhibitor rofecoxib (MK-0966, Vioxx) were prepared by synthetic or biosynthetic methods. Metabolites include products of oxidation, glucuronidation, reduction and hydrolytic ring opening. Based on an in vitro whole blood assay, none of the known human metabolites of rofecoxib inhibits COX-1 nor contributes significantly to the inhibition of COX-2.

Description of a 96-well plate assay to measure cytochrome P4503A inhibition in human liver microsomes using a selective fluorescent probe.

The standard method to evaluate CYP3A inhibition is to study the conversion of the specific CYP3A probe testosterone to its 6 beta-hydroxy metabolite in human liver microsomes, in the absence and presence of potential inhibitors. Quantification of the 6 beta-hydroxy metabolite is achieved by HPLC resulting in a tedious and time-consuming assay. In order to increase the P450 inhibition throughput, efforts were made to find a CYP3A probe that would produce a fluorescent metabolite. This paper reports the discovery of DFB as a potential CYP3A fluorescent probe. DFB was significantly metabolized in human microsomes (approximately 1-2 nmol/(min. mg protein)) to give the fluorescent compound DFH. The involvement of CYP3A in the metabolism of DFB was determined using multiple approaches. First, incubations conducted with microsomes made from cell lines expressing single CYPs (Gentest Supersomes) indicated that CYP3A played a major role in the metabolism of DFB. Secondly, immunoinhibition studies conducted with CYP3A antibody resulted in >95% inhibition of DFB metabolism in HLM. Thirdly, inhibition studies with specific CYP1A1, 1A2, 2C8/9, 2C19, 2D6, and 2E1 chemical inhibitors did not suppress DFB activity in HLM. However, ketoconazole, miconazole, nicardipine, and nifedipine, all known CYP3A inhibitors, completely abolished the formation of DFH in HLM. The potency of several inhibitors determined using DFB and testosterone as CYP3A probes was consistent (R = 0.98). Finally, a good agreement was obtained for the formation of DFH and production of 6 beta-hydroxytestosterone when DFB and testosterone were incubated separately with various human liver microsome preparations (R = 0.94, N = 11). In order to use DFH as a fluorescent CYP3A marker in a 96-well plate format, it was important to remove the excess of NADPH at the end of the incubation because the fluorescence of NADPH interferes with DFH detection. This was achieved by adding oxidized glutathione and glutathione reductase to convert NADPH to NADP(+) which is not fluorescent. The liquid-handling steps were fully automated in a 96-well plate format and a template was designed to generate IC(50) curves and to address potential fluorescent interferences from the test compounds. The assay was found to be reproducible (intraday variability <10% and interday variability indicated less than a 2-fold variation in the IC(50) values) and is now routinely used in our laboratory to evaluate CYP3A inhibition of NCEs.

A new structural variation on the methanesulfonylphenyl class of selective cyclooxygenase-2 inhibitors.

By inserting an oxygen link between the 3-fluorophenyl and the lactone ring of 5,5-dimethyl-3-(3fluorophenyl)-4-(4-methanesulfonylphenyl)-2 (5H)-furanone 1 (DFU), analogs with enhanced in vitro COX-2 inhibitory potency as well as in vivo potency in models of inflammation were obtained.

Synthesis and biological evaluation of 3-heteroaryloxy-4-phenyl-2(5H)-furanones as selective COX-2 inhibitors.

A series of 3-heteroaryloxy4-phenyl-2-5H)-furanones were prepared and evaluated for their potency and selectivity as COX-2 inhibitors. This led to the identification of L-778,736 as a potent, orally active and selective inhibitor of the COX-2 enzyme.

Rofecoxib [Vioxx, MK-0966; 4-(4'-methylsulfonylphenyl)-3-phenyl-2-(5H)-furanone]: a potent and orally active cyclooxygenase-2 inhibitor. Pharmacological and biochemical profiles.

The discoveries that cyclooxygenase (COX)-2 is an inducible form of COX involved in inflammation and that COX-1 is the major isoform responsible for the production of prostaglandins (PGs) in the gastrointestinal tract have provided a rationale for the development of specific COX-2 inhibitors as a new class of anti-inflammatory agents with improved gastrointestinal tolerability. In the present study, the preclinical pharmacological and biochemical profiles of rofecoxib [Vioxx, also known as MK-0966, 4-(4'-methylsulfonylphenyl)-3-phenyl-2-(5H)-furanone], an orally active COX-2 inhibitor, are described. Rofecoxib is a potent inhibitor of the COX-2-dependent production of PGE(2) in human osteosarcoma cells (IC(50) = 26 +/- 10 nM) and Chinese hamster ovary cells expressing human COX-2 (IC(50) = 18 +/- 7 nM) with a 1000-fold selectivity for the inhibition of COX-2 compared with the inhibition of COX-1 activity (IC(50) > 50 microM in U937 cells and IC(50) > 15 microM in Chinese hamster ovary cells expressing human COX-1). Rofecoxib is a time-dependent inhibitor of purified human recombinant COX-2 (IC(50) = 0.34 microM) but caused inhibition of purified human COX-1 in a non-time-dependent manner that could only be observed at a very low substrate concentration (IC(50) = 26 microM at 0.1 microM arachidonic acid concentration). In an in vitro human whole blood assay, rofecoxib selectively inhibited lipopolysaccharide-induced, COX-2-derived PGE(2) synthesis with an IC(50) value of 0.53 +/- 0.02 microM compared with an IC(50) value of 18.8 +/- 0.9 microM for the inhibition of COX-1-derived thromboxane B(2) synthesis after blood coagulation. Using the ratio of the COX-1 IC(50) values over the COX-2 IC(50) values in the human whole blood assay, selectivity ratios for the inhibition of COX-2 of 36, 6.6, 2, 3, and 0.4 were obtained for rofecoxib, celecoxib, meloxicam, diclofenac, and indomethacin, respectively. In several in vivo rodent models, rofecoxib is a potent inhibitor of carrageenan-induced paw edema (ID(50) = 1.5 mg/kg), carrageenan-induced paw hyperalgesia (ID(50) = 1.0 mg/kg), lipopolysaccharide-induced pyresis (ID(50) = 0.24 mg/kg), and adjuvant-induced arthritis (ID(50) = 0.74 mg/kg/day). Rofecoxib also has a protective effect on adjuvant-induced destruction of cartilage and bone structures in rats. In a (51)Cr excretion assay for detection of gastrointestinal integrity in either rats or squirrel monkeys, rofecoxib has no effect at doses up to 200 mg/kg/day for 5 days. Rofecoxib is a novel COX-2 inhibitor with a biochemical and pharmacological profile clearly distinct from that of current nonsteroidal anti-inflammatory drugs and represents a new therapeutic class of anti-inflammatory agents for the treatment of the symptoms of osteoarthritis and rheumatoid arthritis with improved gastrointestinal tolerability.

The discovery of rofecoxib, [MK 966, Vioxx, 4-(4'-methylsulfonylphenyl)-3-phenyl-2(5H)-furanone], an orally active cyclooxygenase-2-inhibitor.

The development of a COX-2 inhibitor rofecoxib (MK 966, Vioxx) is described. It is essentially equipotent to indomethacin both in vitro and in vivo but without the ulcerogenic side effect due to COX-1 inhibition.

A series of non-quinoline cysLT1 receptor antagonists: SAR study on pyridyl analogs of Singulair.

The structure-activity relationship of a series of styrylpyridine analogs of MK-0476 (montelukast, Singulair) is described. This work has led to the identification of a number of potent and orally active cysLT1 receptor (LTD4 receptor) antagonists including 2ab (L-733,321) as an optimized candidate.

2-Pyridinyl-3-(4-methylsulfonyl)phenylpyridines: selective and orally active cyclooxygenase-2 inhibitors.

A series of novel 2-pyridinyl-3-(4-methylsulfonyl)phenylpyridines has been synthesized and evaluated with respect to their ability to inhibit the isozymes of cyclooxygenase, COX-1, and COX-2. Optimum COX-2 activity is observed by introduction of a substituent at C5 of the central pyridine. 5- Chloro-3-(4-methylsulfonyl)phenyl-2-(2-methyl-5-pyridinyl)pyridine 33 was identified as the optimum compound in this series.

Cyclooxygenase-2 inhibitors. Synthesis and pharmacological activities of 5-methanesulfonamido-1-indanone derivatives.

The recent discovery of an alternative form cyclooxygenase (cyclooxygenase-2, COX-2), which has been proposed to play a significant role in inflammatory conditions, may provide an opportunity to develop anti-inflammatory drugs with fewer side effects than existing non-steroidal anti-inflammatory drugs (NSAIDs). We have now identified 6-[(2,4-difluorophenyl)-thio]-5-methanesulfonamido-1-indanone++ + (20) (L-745,337) as a potent, selective, and orally active COX-2 inhibitor. The structure-activity relationships in this series have been extensively studied. Ortho- and para-substituted 6-phenyl substitutents are optimal for in vitro potency. Replacement of this phenyl ring by a variety of heterocycles gave compounds that were less active. The methanesulfonamido group seems to be the optimal group at the 5-position of the indanone system. Compound 20 has an efficacy profile that is superior or comparable to that of the nonselective COX inhibitor indomethacin in animal models of inflammation, pain, and fever and appears to be nonulcerogenic within the dosage ranges required for functional efficacy. Although 20 and its oxygen linkage analog 2 (flosulide) are equipotent in the in vitro assays, compound 20 is more potent in the rat paw edema assay, has a longer t1/2 in squirrel monkeys, and seems less ulcergenic than 2 in rats.

Pharmacology of montelukast sodium (Singulair), a potent and selective leukotriene D4 receptor antagonist.

Montelukast sodium (Singulair), also known as MK-0476 (1-(((1(R)-(3-(2-(7-chloro-2-quinolinyl)-(E)-ethenyl)phenyl)(3-2-(1- hydroxy-1-methylethyl)phenyl)propyl)thio)methyl)cyclopropane) acetic acid sodium salt, is a potent and selective inhibitor of [3H]leukotriene D4 specific binding in guinea pig lung (Ki 0.18 +/- 0.03 nM), sheep lung (Ki 4 nM), and dimethylsulfoxide-differentiated U937 cell plasma membrane preparations (Ki 0.52 +/- 0.23 nM), but it was essentially inactive versus [3H]leukotriene C4 specific binding in dimethylsulfoxide-differentiated U937 cell membranes (IC50 10 microM) and [3H]leukotriene B4 specific binding in THP-1 cell membranes (IC50 40 microM). Montelukast also inhibited specific binding of [3H]leukotriene D4 to guinea pig lung in the presence of human serum albumin, human plasma, and squirrel monkey plasma with Ki values of 0.21 +/- 0.08, 0.19 +/- 0.02, and 0.26 +/- 0.02 nM, respectively. Functionally, montelukast antagonized contractions of guinea pig trachea induced by leukotriene D4 (pA2 value 9.3; slope 0.8). In contrast, montelukast (16 microM) failed to antagonize contractions of guinea pig trachea induced by leukotriene C4 (45 mM serine-borate), serotonin, acetylcholine, histamine, prostaglandin D2, or U-44069. Intravenous montelukast antagonized bronchoconstriction induced in anesthetized guinea pigs by i.v. leukotriene D4 but did not block bronchoconstriction to arachidonic acid, histamine, serotonin, or acetylcholine. Oral administration of montelukast blocked leukotriene D4 induced bronchoconstriction in conscious squirrel monkeys, ovalbumin-induced bronchoconstriction in conscious sensitized rats (ED50 0.03 +/- 0.001 mg/kg; 4 h pretreatment), and also ascaris-induced early and late phase bronchoconstriction in conscious squirrel monkeys (0.03-0.1 mg/kg; 4 h pretreatment). A continuous i.v. infusion of montelukast (8 micrograms.kg-1.min-1) resulted in a 70% decrease in the peak early response and a 75% reduction of the late response to ascaris aerosol in allergic conscious sheep. Montelukast, a potent and selective leukotriene D4 receptor antagonist with excellent in vivo activity is currently in clinical development for the treatment of asthma and related diseases.

A new class of leukotriene biosynthesis inhibitor: the development of MK-0591.

The evolution of MK-0591 (3-[1-(4-chlorobenzyl)-3-(t-butylthio)-5-(quinolin-2-ylmethoxy+ ++)indol-2-yl]- 2,2-dimethylpropanoic acid), 12, a potent, orally active leukotriene biosynthesis inhibitor is described. MK-0591 is currently undergoing clinical evaluation as a potential agent for the treatment of asthma and inflammatory bowel disease. It acts through a novel mechanism by a specific interaction with a membrane protein, 5-lipoxygenase activating protein (FLAP), which has been shown to be essential for LT synthesis in inflammatory cells. A brief comparison of its biological activity with that of its progenitors MK-886 and L-674,636 is described.

Development of a novel series of styrylquinoline compounds as high-affinity leukotriene D4 receptor antagonists: synthetic and structure-activity studies leading to the discovery of (+-)-3-[[[3-[2-(7-chloro-2-quinolinyl)-(E)-ethenyl]phenyl][[3- (dimethylamino)-3-oxopropyl]thio]methyl]thio]propionic acid.

Based on LTD4 receptor antagonist activity of 3-(2-quinolinyl-(E)-ethenyl)pyridine (2) found in broad screening, structure-activity studies were carried out which led to the identification of 3-[[[3-[2-(7-chloro-2-quinolinyl)-(E)-ethenyl]phenyl][[3- (dimethylamino)-3-oxopropyl]thio]methyl]thio]propionic acid (1, MK-571) as a potent and orally active LTD4 receptor antagonist. These studies demonstrated that a phenyl ring could replace the pyridine in 2 without loss of activity, that 7-halogen substitution in the quinoline group was optimal for binding, that the (E)-ethenyl linkage was optimal, that binding was enhanced by incorporation of a polar acidic group or groups in the 3-position of the aryl ring, and that two acidic groups could be incorporated via a dithioacetal formed from thiopropionic acid and the corresponding styrylquinoline 3-aldehyde to yield compounds such as 20 (IC50 = 3 nM vs [3H]LTD4 binding to the guinea pig lung membrane). It was found that one of the acidic groups could be transformed into a variety of the amides without loss of potency and that the dimethylamide 1 embodied the optimal properties of intrinsic potency (IC50 = 0.8 nM on guinea pig lung LTD4 receptor) and oral in vivo potency in the guinea pig, hyperreactive rat, and squirrel monkey. The evolution of 2 to 1 involves the increase of > 6000-fold in competition for [3H]LTD4 binding to guinea pig lung membrane and a > 40-fold increase in oral activity as measured by inhibition of antigen-induced dyspnea in hyperreactive rats.

Pharmacology of the leukotriene antagonist verlukast: the (R)-enantiomer of MK-571.

Verlukast (MK-679) (3-[(3-(2-(7-chloro-2-quinolinyl)-(E)-ethenyl)phenyl)[3-(dimethylamino)- 3- oxopropyl)thio)methyl)-thio)propionic acid) is a potent and selective inhibitor of [3H]leukotriene D4 binding in guinea-pig (IC50 = 3.1 +/- 0.5 nM) and human (IC50 = 8.0 +/- 3.0 nM) lung homogenates and dimethyl sulfoxide differentiated U937 cell membrane preparations (IC50 = 10.7 +/- 1.6 nM) but is essentially inactive versus [3H]leukotriene C4 binding in guinea-pig lung homogenates (IC50 values of 19 and 33 microM). Functionally, when tested at 60 nM, it antagonized contractions of guinea-pig trachea (GPT) induced by leukotriene C4, leukotriene D4, and leukotriene E4 (respective-log KB values of 8.6, 8.8, and 8.9) and contractions of human trachea (HT) induced by leukotriene D4 (-log KB value 8.3 +/- 0.2). In contrast, verlukast (20-200 nM) failed to antagonize contractions of GPT induced by leukotriene C4 in the presence of 45 mM L-serine borate. Intravenous (i.v.) and aerosol verlukast antagonized bronchoconstriction (BC) induced in anaesthetized guinea pigs by i.v. leukotriene D4 but did not block BC to arachidonic acid or histamine. Intraduodenal verlukast (0.25 mg/kg) antagonized leukotriene D4 (0.2 micrograms/kg) induced BC in guinea pigs. Oral and aerosol administration blocked leukotriene D4-induced BC in conscious squirrel monkeys. Orally administered compound also blocked ovalbumin-induced BC in conscious sensitized rats treated with methysergide (3 micrograms/kg). The pharmacological profile for verlukast is similar to that of the racemic compound, MK-571. Verlukast is currently in clinical development for the treatment of asthma and related diseases.

5-Lipoxygenase-activating protein is the target of a quinoline class of leukotriene synthesis inhibitors.

An indole class of leukotriene synthesis inhibitors, exemplified by MK-886, which does not directly inhibit 5-lipoxygenase, has been shown to bind to an 18-kDa leukocyte membrane protein and to inhibit 5-lipoxygenase membrane translocation. It was demonstrated that the 18-kDa protein is necessary for the cellular activation of leukotriene synthesis and was named 5-lipoxygenase-activating protein (FLAP). We describe here a class of leukotriene synthesis inhibitors based on a quinoline structure, which is structurally distinct from MK-886. However, similar to MK-886, several quinolines are potent inhibitors of cellular leukotriene synthesis but are poor inhibitors of soluble 5-lipoxygenase. To determine whether FLAP is the protein target of leukotriene synthesis inhibitors of the quinoline class, we investigated the ability of these compounds to inhibit photoaffinity labeling of FLAP and to elute FLAP from indole affinity gels. The abilities of the quinoline inhibitors to interact with FLAP correlated well with their abilities to inhibit leukotriene synthesis in human polymorphonuclear leukocytes. L-674,573, a potent quinoline leukotriene synthesis inhibitor, inhibited indole photoaffinity labeling of FLAP in a concentration-dependent manner. In addition, L-674,573 selectively eluted FLAP from indole affinity gels, in contrast to L-671,480, a quinoline that was inactive as an inhibitor of leukotriene synthesis. When human leukocyte membranes were labeled with the indole photoaffinity probe [125I]L-669,083 and immunoprecipitated with a FLAP antibody, the labeling of FLAP was inhibited by L-674,573 but not by L-671,480. These results suggest a direct binding site for the quinoline leukotriene synthesis inhibitors on FLAP and provide further evidence for the essential role of FLAP in cellular leukotriene synthesis.

Stereospecific synthesis, assignment of absolute configuration, and biological activity of the enantiomers of 3-[[[3-[2-(7-chloroquinolin-2-yl)-(E)-ethenyl]phenyl] [[3-(dimethylamino)-3-oxopropyl]thio]methyl]thio]propionic acid, a potent and specific leukotriene D4 receptor antagonist.

The enantiomers of the leukotriene D4 antagonist 3-[[[3-[2-(7-chloroquinolin-2-yl)-(E)-ethenyl]phenyl] [[3-(dimethylamino)-3-oxopropyl]thio]methyl]thio]propionic acid (L-660,711)(MK-571) have been prepared, their absolute stereochemistry has been assigned as S for (+)-1 and R for (-)-1 by X-ray analysis of a synthetic intermediate (5), and the biological activity of the enantiomers has been explored. Unexpectedly, the enantiomers are both comparably biologically active with (+)-1 slightly more intrinsically active at the LTD4 receptor in vitro.

Pharmacology of L-660,711 (MK-571): a novel potent and selective leukotriene D4 receptor antagonist.

L-660,711 (3-(3-(2-(7-chloro-2-quinolinyl)ethenyl)phenyl) ((3-dimethyl amino-3-oxo propyl)thio)methyl)thio)propanoic acid is a potent and selective competitive inhibitor of [3H]leukotriene D4 binding in guinea pig (Ki value, 0.22 nM) and human (Ki value, 2.1 nM) lung membranes but is essentially inactive versus [3H]leukotriene C4 binding (IC50 value in guinea pig lung, 23 microM). Functionally it competitively antagonized contractions of guinea pig trachea and ileum induced by leukotriene (LT) D4 (respective pA2 values, 9.4 and 10.5) and LTE4 (respective pA2 values, 9.1 and 10.4) and contractions of human trachea induced by LTD4 (pA2 value, 8.5). L-660,711 (5.8 x 10(-8)M) antagonized contractions of guinea pig trachea induced by LTC4 in the absence (dose ratio = 28) but not in the presence of 45 mM L-serine borate (dose ratio less than 2). L-660,711 (1.9 x 10(-5)M) did not block contractions of guinea pig trachea induced by histamine, acetylcholine, 5-hydroxytryptamine, PGF2 alpha, U-44069, or PGD2. In the presence of atropine, mepyramine, and indomethacin, L-660,711 (1.9 x 10(-5)M) inhibited a small component of the response to antigen on guinea pig trachea but completely blocked anti-IgE-induced contractions of human trachea. L-660,711 (i.v.) antagonized bronchoconstriction induced in anesthetized guinea pigs by i.v. LTC4, LTD4, and LTE4 but did not block bronchoconstriction to arachidonic acid, U-44069, 5-hydroxytryptamine, histamine, or acetylcholine. Intraduodenal L-660,711 antagonized LTD4 (0.2-12.8 micrograms/kg)-induced bronchoconstriction in guinea pigs, and p.o. L-660,711 blocked LTD4- and Ascaris-induced bronchoconstriction in conscious squirrel monkeys and ovalbumin-induced bronchoconstriction in conscious sensitized rats treated with methysergide (3 micrograms/kg). The pharmacological profile of L-660,711 indicates that it is a potent, selective, orally active leukotriene receptor antagonist which is well suited to determine the role played by LTD4 and LTE4 in asthma and other pathophysiologic conditions.

L-649,923, sodium (beta S*, gamma R*)-4-(3-(4-acetyl-3-hydroxy-2-propylphenoxy)-propylthio)- gamma-hydroxy-beta-methylbenzenebutanoate, a selective, orally active leukotriene receptor antagonist.

L-649,923, Sodium (beta S*, gamma R*)-4-(3-(4-acetyl-3-hydroxy-2-propylphenoxy)propylthio)- gamma- hydroxy-beta-methylbenzenebutanoate is a selective and competitive inhibitor of [3H]leukotriene D4 (Ki value of 400 nM) and to a lesser extent [3H]leukotriene C4 (Ki value of 8.6 microM) binding in guinea-pig lung homogenates. Functionally, it selectively antagonized contractions of guinea pig trachea induced by leukotriene C4, D4, E4, and F4 but not those induced by acetylcholine, histamine, serotonin, prostaglandin F2 alpha, or U-44069 (stable endoperoxide analogue). Schild plot analysis indicated a competitive inhibition of contractions of guinea-pig ileum induced by leukotriene D4 (pA2 8.1) and contractions of guinea-pig trachea induced by leukotrienes E4 and F4 (pA2 7.1 and 6.9, respectively). In contrast, contractions of guinea-pig trachea induced by leukotrienes C4 (pA2 7.2; slope 0.6) and D4 (pA2 7.2; slope 0.7) were inhibited in a noncompetitive fashion. In vivo, intravenously administered L-649,923 selectively blocked bronchoconstriction induced in anesthetized guinea pigs by leukotriene C4 and D4 (ED50 values i.v. 0.38 and 0.26 mg/kg, respectively) but not that induced by histamine, arachidonic acid, serotonin, U-44069, or acetylcholine. Following intraduodenal administration, L-649,923, blocked leukotriene D4 induced bronchoconstriction (5 and 10 mg/kg). The present findings indicate that selective antagonists, such as L-649,923, may be useful for defining the role of leukotrienes in diseases such as bronchial asthma.