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A high-throughput screen against human DGAT-1 led to the identification of a core structure that was subsequently optimized to afford the potent, selective, and orally bioavailable compound 14. Oral administration at doses ≥0.03 mg/kg significantly reduced postprandial triglycerides in mice following an oral lipid challenge. Further assessment in both acute and chronic safety pharmacology and toxicology studies demonstrated a clean profile up to high plasma levels, thus culminating in the nomination of 14 as clinical candidate ABT-046.

Mounting evidence suggests excessive glucocorticoid activity may contribute to Alzheimer's disease (AD) and age-associated memory impairment. 11β-hydroxysteroid dehydrogenase type-1 (HSD1) regulates conversion of glucocorticoids from inactive to active forms. HSD1 knock-out mice have improved cognition, and the nonselective inhibitor carbenoxolone improved verbal memory in elderly men. Together, these data suggest that HSD1 inhibition may be a potential therapy for cognitive deficits, such as those associated with AD. To investigate this, we characterized two novel and selective HSD1 inhibitors, A-918446 and A-801195. Learning, memory consolidation, and recall were evaluated in mouse 24 h inhibitory avoidance. Inhibition of brain cortisol production and phosphorylation of cAMP response element-binding protein (CREB), a transcription factor involved in cognition, were also examined. Rats were tested in a short-term memory model, social recognition, and in a separate group cortical and hippocampal acetylcholine release was measured via in vivo microdialysis. Acute treatment with A-801195 (10-30 mg/kg) or A-918446 (3-30 mg/kg) inhibited cortisol production in the ex vivo assay by ∼ 35-90%. Acute treatment with A-918446 improved memory consolidation and recall in inhibitory avoidance and increased CREB phosphorylation in the cingulate cortex. Acute treatment with A-801195 significantly improved short-term memory in rat social recognition that was not likely due to alterations of the cholinergic system, as acetylcholine release was not increased in a separate set of rats. These studies suggest that selective HSD1 inhibitors work through a novel, noncholinergic mechanism to facilitate cognitive processing.

A highly potent and selective DGAT-1 inhibitor was identified and used in rodent models of obesity and postprandial chylomicron excursion to validate DGAT-1 inhibition as a novel approach for the treatment of metabolic diseases. Specifically, compound 4a conferred weight loss and a reduction in liver triglycerides when dosed chronically in DIO mice and depleted serum triglycerides following a lipid challenge in a dose-dependent manner, thus, reproducing major phenotypical characteristics of DGAT-1(-/-) mice.

A series of novel cyanoguanidine derivatives was designed and synthesized. Condensation of N-(1-benzotriazol-1-yl-2,2-dichloropropyl)-substituted benzamides with N-(substituted-pyridin-3-yl)-N'-cyanoguanidines furnished N-{2,2-dichloro-1-[N'-(substituted-pyridin-3-yl)-N''-cyanoguanidino]propyl}-substituted benzamide derivatives. These agents were glyburide-reversible potassium channel openers and hyperpolarized human bladder cells as assessed by the FLIPR membrane potential dye (KATP-FMP). These compounds were also potent full agonists in relaxing electrically stimulated pig bladder strips, an in vitro model of overactive bladder. The most active compound 9 was evaluated for in vivo efficacy and selectivity in a pig model of bladder instability. Preliminary pharmacokinetic studies in dog demonstrated excellent oral bioavailability and a t1/2 of 15 h. The synthesis, SAR studies, and biological properties of these agents are discussed.

Background and Purpose: ATP-sensitive K(+) channels (K(ATP)) play a pivotal role in contractility of urinary bladder smooth muscle. This study reports the characterization of 4-methyl-N-(2,2,2-trichloro-1-(3-pyridin-3-ylthioureido)ethyl)benzamide (A-251179) as a K(ATP) channel opener., Experimental Approach: Glyburide-sensitive membrane potential, patch clamp and tension assays were employed to study the effect of A-251179 in vitro. The in vivo efficacy of A-251179 was characterized by suppression of spontaneous contractions in obstructed rat bladder and by measuring urodynamic function of urethane-anesthetized rat models., Key Results: A-251179 was about 4-fold more selective in activating SUR2B-Kir6.2 derived K(ATP) channels compared to those derived from SUR2A-Kir6.2. In pig bladder smooth muscle strips, A-251179 suppressed spontaneous contractions, about 27- and 71-fold more potently compared to suppression of contractions evoked by low-frequency electrical stimulation and carbachol, respectively. In vivo, A-251179 suppressed spontaneous non-voiding bladder contractions from partial outlet-obstructed rats. Interestingly, in the neurogenic model where isovolumetric contractions were measured by continuous transvesical cystometry, A-251179 at a dose of 0.3 micromol kg(-1), but not higher, was found to increase bladder capacity without affecting either the voiding efficiency or changes in mean arterial blood pressure., Conclusions and Implications: The thioureabenzamide analog, A-251179 is a potent novel K(ATP) channel opener with selectivity for SUR2B/Kir6.2 containing K(ATP) channels relative to pinacidil. The pharmacological profile of A-251179 is to increase bladder capacity and to prolong the time between voids without affecting voiding efficiency and represents an interesting characteristic to be explored for further investigations of K(ATP) channel openers for the treatment of overactive bladder.

The incorporation of constrained tertiary amines into an existing class of N-benzyl-4-aminopiperidinyl chromone-based MCHr1 antagonists led to the identification of a series of chiral racemic compounds that displayed good to excellent functional potency, binding affinity, and selectivity over the hERG channel. Further separation of two distinct chiral racemic compounds into their corresponding pairs of enantiomers revealed a considerable selectivity for MCHr1 for one configuration, in addition to a striking difference in oral exposure between one pair of enantiomers in diet-induced obese mice. Oral administration of the most potent compound in this class in the same animal model led to significant reduction of fat mass in a semi-chronic model for weight loss.

Starting from a rapidly metabolized adamantane 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) inhibitor 22a, a series of E-5-hydroxy-2-adamantamine inhibitors, exemplified by 22d and (+/-)-22f, was discovered. Many of these compounds are potent inhibitors of 11beta-HSD1 and are selective over 11beta-HSD2 for multiple species (human, mouse, and rat), unlike other reported species-selective series. These compounds have good cellular potency and improved microsomal stability. Pharmacokinetic profiling in rodents indicated moderate to large volumes of distribution, short half-lives, and a pharmacokinetic species difference with the greatest exposure measured in rat with 22d. One hour postdose liver, adipose, and brain tissue 11beta-HSD1 inhibition was confirmed with (+/-)-22f in a murine ex vivo assay. Although 5,7-disubstitued-2-adamantamines provided greater stability, a single, E-5-position, polar functional group afforded inhibitors with the best combination of stability, potency, and selectivity. These results indicate that adamantane metabolic stabilization sufficient to obtain short-acting, potent, and selective 11beta-HSD1 inhibitors has been discovered.

Structure-activity relationships were investigated on the tricyclic dihydropyridine (DHP) KATP openers 9-(3-bromo-4-fluorophenyl)-5,9-dihydro-3H,4H-2,6-dioxa-4-azacyclopenta[b]naphthalene-1,8-dione (6) and 10-(3-bromo-4-fluorophenyl)-9,10-dihydro-1H,8H-2,7-dioxa-9-azaanthracene-4,5-dione (65). Substitution off the core of the DHP, absolute stereochemistry, and aromatic substitution were evaluated for KATP channel activity using Ltk- cells stably transfected with the Kir6.2/SUR2B exon 17- splice variant and in an electrically stimulated pig bladder strip assay. A select group of compounds was evaluated for in vitro inhibition of spontaneous bladder contractions. Several compounds were found to have the unique characteristic of partial efficacy in both the cell-based and electrically stimulated bladder strip assays but full efficacy in inhibiting spontaneous bladder strip contractions. For compound 23b, this profile was mirrored in vivo where it was fully efficacious in inhibiting spontaneous myogenic bladder contractions but only partially able to reduce neurogenically mediated reflex bladder contractions.

The discovery and pharmacological evaluation of potent, selective, and orally bioavailable growth hormone secretagogue receptor (GHS-R) antagonists are reported. Previously, 2,4-diaminopyrimidine-based GHS-R antagonists reported from our laboratories have been shown to be dihydrofolate reductase (DHFR) inhibitors. By comparing the X-ray crystal structure of DHFR docked with our GHS-R antagonists and GHS-R modeling, we designed and synthesized a series of potent and DHFR selective GHS-R antagonists with good pharmacokinetic (PK) profiles. An amide derivative 13d (Ca2+ flux IC50 = 188 nM, [brain]/[plasma] = 0.97 @ 8 h in rat) showed a 10% decrease in 24 h food intake in rats, and over 5% body weight reduction after 14-day oral treatment in diet-induced obese (DIO) mice. In comparison, a urea derivative 14c (Ca2+ flux IC50 = 7 nM, [brain]/[plasma] = 0.0 in DIO) failed to show significant effect on food intake in the acute feeding DIO model. These observations demonstrated for the first time that peripheral GHS-R blockage with small molecule GHS-R antagonists might not be sufficient for suppressing appetite and inducing body weight reduction.

The aim of this study was to validate melanin-concentrating hormone (MCH)-1 receptor antagonism as a potential treatment of mood disorders. We attempted to replicate the effects previously reported with SNAP-7941 and expanded the investigation to three other orally bioavailable MCH-1 receptor antagonists with good brain penetration. SNAP-7941 (3-30 mg/kg, i.p.) and T-226296 (5-60 mg/kg, p.o.) (+/- racemate), were evaluated in the rat forced swim and mouse tail suspension tests. (+)SNAP-7941 (3-10 mg/kg, p.o.) was also tested in a modified 5-min rat forced swim protocol as previously reported. A-665798 (3-30 mg/kg, p.o.) and A-777903 (3-30 mg/kg, p.o.) were tested in mouse tail suspension and rat Vogel tests. None of the compounds showed meaningful efficacy in the paradigms tested. The lack of efficacy with four structurally different MCH-1 receptor antagonists does not support a role for therapeutic treatment of depression/anxiety via this mechanism of action.

Ghrelin, a gut-derived orexigenic hormone, is an endogenous ligand of the growth hormone secretagogue receptor (GHS-R). Centrally administered ghrelin has been shown to cause hunger and increase food intake in rodents. Inhibition of ghrelin actions with ghrelin antibody, peptidyl GHS-R antagonists, and antisense oligonucleosides resulted in weight loss and food intake decrease in rodents. Here we report the effects of GHS-R antagonists, some of which were potent, selective, and orally bioavailable. A structure-activity relationship study led to the discovery of 8a, which was effective in decreasing food intake and body weight in several acute rat studies.

Dexfenfluramine (DEX) and sibutramine (SIB) are effective antiobesity agents. Their effects on weight control and hormone profile have not been previously studied in diet-switched diet-induced obese (DIO) mice, in which treatment is initiated upon cessation of a low-fat diet and resumption of a high-fat diet. Furthermore, their effects on circulating ghrelin in obese humans or in animal models of obesity have not yet been reported. Male C57Bl/6J DIO mice after 16 wk on a high-fat diet (HF, 60 kcal% fat) were switched to a low-fat diet (LF, 10 kcal% fat) for 50 d. HF diet resumed concurrently with treatment for 28 d with DEX 3 and 10 mg/kg, twice a day (BID); SIB 5 mg/kg BID; or vehicle. Rapid weight regain ensued in vehicle-treated DIO mice. DEX or SIB treatment significantly blunted the body weight gain. Caloric intake was decreased acutely by DEX or SIB vs vehicle during the first 2 d treatment, but returned to control after 5 d. At the end of study, epididymal fat weight and whole body fat mass determined by DEXA scan were decreased by DEX 10 mg/kg, and whole body lean mass decreased with DEX 3 mg/kg treatment. Circulating ghrelin on d 28 was increased with either DEX 3 or 10 mg/kg treatment, while growth hormone and insulin were decreased. Leptin was also decreased in the DEX 10 mg/kg group. SIB did not significantly affect fat mass, ghrelin, growth hormone, insulin, or leptin. Mice chronically fed LF diet maintained a lower caloric intake, gained less weight and fat mass than diet-switched mice, and had higher ghrelin and lower insulin and leptin. In summary, weight regain in diet-switched DIO mice is delayed with either DEX or SIB treatment. DEX treatment of diet-switched DIO mice decreased growth hormone, insulin, leptin, fat mass, lean mass, and increased ghrelin, while SIB only decreased body weight.

Partial bladder outlet obstruction of the pig is considered as a valuable preclinical model for evaluating the profile of compounds for the treatment of bladder overactivity. In this study, we characterized the pharmacological properties of isolated bladder smooth muscle from pigs following partial outlet obstruction and its sensitivity to potassium channel openers. Bladder strips from obstructed animals showed significantly lower maximal efficacy (E(max)) and sensitivity to stimulation by ATP and carbachol, but not to those evoked by serotonin, compared to age-matched controls. Tissue strips from obstructed animals also showed a 2.5-fold increase in the potency and significantly reduced maximum response following K+ depolarization. With respect to spontaneous activity, bladder strips from control strips demonstrated little spontaneous phasic activity at all preloads examined. In contrast, bladder strips from obstructed animals showed large preload-dependent increases in spontaneous phasic activity at preload values of 16-32 g. The potencies of K(ATP) channel openers to relax carbachol-evoked contractions showed a good 1:1 correlation (r(2)=0.90) between obstructed and control bladder strips. These studies demonstrate that obstructed pig bladders show enhanced spontaneous phasic activity especially at elevated preloads, which may underlie unstable myogenic bladder contractions reported in cystometrographic measurements in vivo. The impaired responses to electrical field stimulation could be attributed to reduced efficacies and/or lower sensitivities of muscarinic and purinergic signaling pathways. K(ATP) channel sensitivities remain essentially unimpaired in the obstructed bladder and could be effectively modulated by openers with potential for the treatment of overactive bladder secondary to outlet obstruction.

Deficiency in Vitamin D and its metabolites leads to a failure in bone formation primarily caused by dysfunctional mineralization, suggesting that Vitamin D analogs might stimulate osteoblastic bone formation and mineralization. In this study, we compare the effect of selected Vitamin D analogs and active metabolite, 1alpha,25-dihydroxyvitamin D(3), 19-nor-1alpha, 25-dihydroxyvitamin D(2), and 1alpha-hydroxyvitamin D(2) or 1alpha,25-dihydroxyvitamin D(2) on bone formation and resorption. In a mouse calvariae bone primary organ culture system, all Vitamin D analogs and metabolite tested-stimulated collagen synthesis in a dose-dependent manner and 19-nor-1alpha, 25-dihydroxyvitamin D(2) was the most efficacious among three. 19-nor-1alpha, 25-dihydroxyvitamin D(2) and 1alpha,25-dihydroxyvitamin D(2) showed similar potencies and 1alpha,25-dihydroxyvitamin D(3) was less potent than others. Osteocalcin was also up-regulated in a dose-dependent manner, suggesting that the three Vitamin D analogs have the equal potencies on bone formation. 25-Hydroxyvitamin D-24-hydroxylase expression was induced in a dose-dependent manner and 19-nor-1alpha, 25-dihydroxyvitamin D(2) was less potent than other two compounds. In a mouse calvariae organ culture, all induced a net calcium release from calvariae in a dose-dependent manner, but the potency is in the order of 1alpha,25-dihydroxyvitamin D(2) congruent with1alpha,25-dihydroxyvitamin D(3)>19-nor-1alpha, 25-dihydroxyvitamin D(2). In a Vitamin D/calcium-restricted rat model, all caused an elevation in serum calcium in a dose-dependent manner. There is no significant difference between 1alpha,25-dihydroxyvitamin D(3) and 1alpha-hydroxyvitamin D(2) in potencies, but 19-nor-1alpha, 25-dihydroxyvitamin D(2) is at least 10-fold less potent than the other two compounds. Our results suggest that Vitamin D analogs have direct effects on bone resorption and formation, and 19-nor-1alpha, 25-dihydroxyvitamin D(2) may be more effective than 1alpha,25-dihydroxyvitamin D(3) and 1alpha-hydroxyvitamin D(2) on stimulating anabolic bone formation.

The pharmacological profile of the novel dihydropyridine K channel opener (KCO), (9R)-9-(3-iodo-4-methylphenyl)-5,9-dihydro-3H-furo[3,4-b]pyrano[4,3-e]pyridine-1,8(4H,7H)-dione (A-325100), is described in numerous in vitro assays. Furthermore, the cardiovascular effects of A-325100 are characterized in both the anesthetized and conscious dog. In vitro, A-325100 selectively activated KATP currents and potently relaxed vascular smooth muscle (IC50 between 7.69x10 M and 7.78x10 M), an effect that was abolished by glyburide. Moreover, A-325100 did not interact with L-type Ca2+ channels at concentrations up to 30 microM. In anesthetized dogs A-325100 produced a dose-dependent reduction in systemic vascular resistance and mean arterial pressure concomitant with dose-dependent increases in dP/dtmax and heart rate. In conscious telemetry-instrumented dogs oral administration of A-325100 produced a similar response profile, including dose-dependent reductions in MAP and increases in heart rate and dP/dtmax. When concentration-dependent changes in MAP, heart rate, and dP/dtmax were compared relative to circulating plasma concentrations, A-325100 produced similar effects in both the anesthetized and conscious dog. In conclusion, the present study provides the first pharmacological description of the novel and selective tricyclic dihydropyridine KCO, A-325100. When studied in vivo, A-325100 produced similar concentration-dependent cardiovascular effects in both models consistent with its mode of action and independent of route of administration. Thus, these data demonstrate that the hemodynamic effects of vasoactive compounds, such as KCOs, can be effectively profiled in both the conscious and anesthetized dog.

A series of urea-based N-1-(2-aminoethyl)-indazoles was synthesized and evaluated for melanin-concentrating hormone receptor 1 (MCHr1) antagonism in both binding and functional assays. Several compounds that acted as MCHr1 antagonists were identified, and optimization afforded a compound with excellent binding affinity, good functional potency, and oral efficacy in a chronic model for weight loss in diet-induced obese mice.

Histamine is a key neurotransmitter that alters central nervous system functions in both behavioural and homeostatic contexts through its actions on the histamine (H) subreceptors H(1), H(2) and H(3) G-protein-coupled receptors. H(3)receptors have a diverse central nervous system distribution where they function as both homo- and hetero-receptors to modulate the synthesis and/or release of several neurotransmitters. H(3) receptors are constitutively active, which implies that antagonists of H(3) receptors may also function as inverse agonists to alter the basal state of the receptor and uncouple constitutive receptor-G-protein interactions. Reference H(3) antagonists such as thioperamide and ciproxifan, administered either centrally or systemically, have been shown to cause changes in food consumption and/or body weight in proof-of-concept studies. More recently, several non-imidazole-based H(3) antagonists/inverse agonists have also been described with efficacy in at least one animal model of human obesity. Considerable preclinical effort remains necessary before such compounds achieve therapeutic success or failure. Moreover, ongoing research in a number of laboratories has shed new insights into the effects of H(3) ligands in the control of feeding, appetite and body weight, which offer different results and conclusions. The goal of this review is to appraise these findings and forecast whether any H(3) antagonists/inverse agonists will provide clinical utility to treat human obesity.

1. Openers of ATP-sensitive K(+) channels are of interest in several therapeutic indications including overactive bladder and other lower urinary tract disorders. This study reports on the in vitro and in vivo characterization of a structurally novel naphthylamide N-[2-(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl)-naphthalen-1-yl]-acetamide (A-151892), as an opener of the ATP-sensitive potassium channels. 2. A-151892 was found to be a potent and efficacious potassium channel opener (KCO) as assessed by glibenclamide-sensitive whole-cell current and fluorescence-based membrane potential responses (-log EC(50)=7.63) in guinea-pig bladder smooth muscle cells. 3. Evidence for direct interaction with KCO binding sites was derived from displacement of binding of the 1,4-dihydropyridine opener [(125)I]A-312110. A-151892 displaced [(125)I]A-312110 binding to bladder membranes with a -log Ki value of 7.45, but lacked affinity against over 70 neurotransmitter receptor and ion channel binding sites. 4. In pig bladder strips, A-151892 suppressed phasic, carbachol-evoked and electrical field stimulus-evoked contractility in a glibenclamide-reversible manner with -log IC(50) values of 8.07, 7.33 and 7.02 respectively, comparable to that of the potencies of the prototypical cyanoguanidine KCO, P1075. The potencies to suppress contractions in thoracic aorta (-log IC(50)=7.81) and portal vein (-log IC(50)=7.98) were not substantially different from those observed for suppression of phasic contractility of the bladder smooth muscle. 5. In vivo, A-151892 was found to potently suppress unstable bladder contractions in obstructed models of unstable contractions in both pigs and rats with pED(35%) values of 8.05 and 7.43, respectively. 6. These results demonstrate that naphthylamide analogs exemplified by A-151892 are novel K(ATP) channel openers and may serve as chemotypes to exploit additional analogs with potential for the treatment of overactive bladder and lower urinary tract symptoms.

Structure-activity relationships were investigated on a novel series of sulfonyldihydropyridine-containing K(ATP) openers. Ring sizes, absolute stereochemistry, and aromatic substitution were evaluated for K(ATP) activity in guinea pig bladder cells using a fluorescence-based membrane potential assay and in a pig bladder strip assay. The inhibition of spontaneous bladder contractions in vitro was also examined for a select group of compounds. All compounds studied showed greater potency to inhibit spontaneous bladder contractions relative to their potencies to inhibit contractions elicited by electrical stimulation. In an anesthetized pig model of myogenic bladder overactivity, compound 14 and (-)-cromakalim 1 were found to inhibit spontaneous bladder contractions in vivo at plasma concentrations lower than those that affected hemodynamic parameters. Compound 14 showed approximately 5-fold greater selectivity than 1 in vivo and supports the concept that bladder-selective K(ATP) channel openers may have utility in the treatment of overactive bladder.

Structure-activity studies were performed on the alpha(1A)-adrenoceptor (AR) selective agonist N-[5-(1H-imidazol-4-yl)-5,6,7,8-tetrahydro-1-naphthalenyl]methanesulfonamide (4). Compounds were evaluated for binding activity at the alpha(1A), alpha(1b), alpha(1d), alpha(2a), and alpha(2B) subtypes. Functional activity in tissues containing the alpha(1A) (rabbit urethra), alpha(1B) (rat spleen), alpha(1D) (rat aorta), and alpha(2A) (rat prostatic vas deferens) was also evaluated. A dog in vivo model simultaneously measuring intraurethral pressure (IUP) and mean arterial pressure (MAP) was used to assess the uroselectivity of the compounds. Many of the compounds that were highly selective in vitro for the alpha(1A)-AR subtype were also more uroselective in vivo for increasing IUP over MAP than the nonselective alpha(1)-agonists phenylpropanolamine (PPA) (1) and ST-1059 (2, the active metabolite of midodrine), supporting the hypothesis that greater alpha(1A) selectivity would reduce cardiovascular side effects. However, the data also support a prominent role of the alpha(1A)-AR subtype in the control of MAP.

We have discovered a novel series of N-[2-(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl)-naphthalen-1-yl] amides that are potent openers of K(ATP) channels and investigated structure-activity relationships (SAR) around the 1,2-disubstituted naphthyl core. A-151892, a prototype compound of this series, was found to be a potent and efficacious potassium channel opener in vitro in transfected Kir6.2/SUR2B cells and pig bladder strips. Additionally, A-151892 was found to selectively inhibit unstable bladder contractions in vivo in an obstructed rat model of myogenic bladder function

Symptoms of urinary frequency and urgency secondary to benign prostatic obstruction are common in elderly men. In many patients, these symptoms correspond to the urodynamic finding of involuntary detrusor contractions during filling cystometry (i.e., detrusor instability). Spontaneous non-voiding contractions during filling can be modeled in animals by subchronic, partial urethral obstruction. However, many investigators remove the obstructive ligature a few days prior to cystometrical evaluation (which may not be an ideal representation of the clinical situation where obstruction is still present), and all perform cystometry within 3 days post-bladder catheterization surgery (i.e., while considerable wound healing is present). In the current study, we evaluated the effects, after oral dosing, of three structurally diverse ATP-sensitive potassium channel openers (KCOs) on spontaneous contractions secondary to obstruction in rats with an intact obstructive ligature at the time of testing and 2 weeks post-bladder catheterization. ZD6169, WAY-133537 and a novel dihydropyridine KCO, A-278637, all significantly decreased spontaneous bladder contractions at 30 min post-dosing (p.o.). However, only ZD6169 (10 micromol/kg) and A-278637 (3 micromol/kg) attenuated such bladder contractions at doses that did not concurrently, significantly affect mean arterial blood pressure and heart rate. These data confirm the efficacy of KCOs to inhibit unstable contractions in obstructed rats, and they further demonstrate the positive effect of a novel, bladder-selective KCO, A-278637, in an animal model with potentially less artifact than in previous such models.

Objective: To compare in vivo the potency and bladder-vascular selectivity of ATP-sensitive potassium channel openers (KCOs) (-)-cromakalim, WAY-133537 and ZD6169 and a muscarinic antagonist, tolterodine in rats., Materials and Methods: Bladder and arterial pressures were monitored simultaneously, before and after increasing intravenous doses of compounds, in each of two urethane-anaesthetized rat bladder hyperactivity models: spontaneous non-voiding myogenic contractions secondary to partial outlet obstruction and volume-induced neurogenic contractions., Results: (-)-Cromakalim, WAY-133537 and ZD6169 caused a dose-dependent suppression of spontaneous contractions in the obstructed model, with a 50% inhibition of the contraction area under the curve at doses of 0.06, 0.14 and 2.4 micro mol/kg (intravenous), respectively. Corresponding decreases in mean arterial pressure at these effective doses were 24%, 15% and 15%, respectively. The KCO potency rank order was the same and their relative potency highly comparable in the neurogenic model. There was complete inhibition of spontaneous contractions in obstructed rats at doses corresponding to approximately 50% inhibition of the neurogenic contractions. While tolterodine caused a dose-dependent inhibition of contractions in the neurogenic model, it was ineffective at inhibiting non-voiding contractions in obstructed rats., Conclusions: All KCOs tested caused significant decreases in arterial pressure at doses effective on the bladder in the model of obstructive instability, suggesting a lack of bladder-vascular selectivity. Similar KCO potency in both assays suggests no appreciable changes in KATP channel function as a result of partial outlet obstruction.

Aims: To compare in vivo the efficacy, potency, and bladder-vascular selectivity of ATP-sensitive potassium channel openers (KCOs), YM934 and (-)-cromakalim to a muscarinic antagonist, tolterodine in a novel partial outlet obstructed pig model., Methods: Partially obstructed female Landrace pigs were implanted with telemetry transmitters to allow the continuous measurement of intravesical, abdominal and arterial pressures. A subcutaneous port catheter was used to adjust bladder volume. Bladder and arterial pressure were simultaneously monitored under isoflurane anesthesia before and after increasing i.v. doses of test compounds., Results: Under anesthesia, voiding was completely inhibited, but spontaneous, nonvoiding bladder contractions were observed with mean amplitude of 16 +/- 1 cm H(2)O, duration of 35 +/- 2 seconds, and intercontraction interval of 43 +/- 4 seconds (n = 25). YM934 and (-)-cromakalim both caused dose-dependent decreases in bladder contraction area under the curve (AUC) with effective doses to inhibit AUC by 35% of 3.6 and 14.9 nmol/kg, i.v., respectively. However, concomitant reductions in mean arterial pressure of 12 and 13% were also observed. Tolterodine did not inhibit spontaneous bladder contractions at doses up to 100 nmol/kg, i.v. corresponding to plasma concentrations up to 41 ng/mL., Conclusions: The superior efficacy of KCOs to inhibit spontaneous bladder contractions relative to tolterodine support the hypothesis that KCOs may provide an alternate therapeutic mechanism to treat symptoms of overactive bladder if bladder-vascular selectivity can be sufficiently improved. The minimally invasive model described herein appears useful in the preclinical evaluation of potential therapeutics targeted to treat the overactive bladder., (Copyright 2003 Wiley-Liss, Inc.)

ATP-sensitive potassium (K(ATP)) channel openers (KCOs) have been shown to inhibit spontaneous myogenic contractile activity of the urinary bladder, a mechanism hypothesized to underlie detrusor instability and symptoms of overactive bladder. However, the therapeutic utility of KCOs has been limited by a lack of differentiation of bladder versus vascular effects. In this study, we evaluated the in vivo potency and bladder selectivity of (-)-(9S)-9-(3-bromo-4-fluorophenyl)-2,3,5,6,7,9-hexahydrothieno[3,2-b]quinolin-8(4H)-one 1,1-dioxide (A-278637), a novel dihydropyridine KCO, in a pig model of detrusor instability secondary to partial bladder outlet obstruction. For comparison, we profiled two KCOs, ((R)-4-[3,4-dioxo-2-(1,2,2-trimethyl-propylamino)-cyclobut-1-enylamino]-3-ethyl-benzonitrile (WAY-133537) and (S)-N-(4-benzoylphenyl)-3,3,3-trifluro-2-hydroxy-2-methyl-propionamide (ZD6169), reported previously to have improved bladder selectivity in vivo and a calcium channel blocker, nifedipine. Effective doses of A-278637, WAY-133537, ZD6169, and nifedipine to inhibit unstable contraction area under the curve by 35% and to decrease mean arterial pressure by 10% were 4.2 and 12, 109 and 51, 661 and 371, and 136 and 30 nmol/kg i.v., yielding corresponding bladder selectivity ratios of 3, 0.5, 0.6, and 0.2. Therefore, A-278637 was approximately 5- to 6-fold more bladder-selective than the other KCOs and 15-fold more selective than nifedipine, the latter approximately 4.5-fold vascular-selective. The potency of KCOs to inhibit unstable contraction in vivo was accurately predicted by their potency to inhibit spontaneous contractile activity of pig detrusor strips in vitro. These results indicate that A-278637, with enhanced potency and bladder selectivity compared with the other compounds evaluated, could serve as a useful tool in the investigation of smooth muscle K(ATP) channel openers as novel therapeutic agents for the treatment of overactive bladder.

N-[3-(1H-Imidazol-4-ylmethyl)phenyl]ethanesulfonamide (ABT-866, 1) is a novel alpha(1) agent having the unique profile of alpha(1A) (rabbit urethra, EC(50) = 0.60 microM) agonism with alpha(1B) (rat spleen, pA(2) = 5.4) and alpha(1D) (rat aorta, pA(2) = 6.2) antagonism. An in vivo dog model showed 1 to be more selective for the urethra over the vasculature than A-61603 (2), ST-1059 (3, the active metabolite of midodrine), and phenylpropanolamine (4).

Fiduxosin is a new alpha(1)-adrenoceptor antagonist targeted for the treatment of symptomatic benign prostatic hyperplasia. The purpose of this study was to determine and compare the potencies of the alpha(1)-adrenoceptor antagonists terazosin, doxazosin, tamsulosin, and fiduxosin, based on relationships between plasma drug concentrations and blockade of phenylephrine (PE)-induced intraurethral (IUP) and mean arterial pressure (MAP) responses after single oral dosing in conscious male beagle dogs. Magnitude of blockade and plasma concentrations were evaluated at selected time points over 24 h. All drugs produced dose-dependent antagonism of PE-induced IUP and MAP responses. When IUP and MAP blockade effects were plotted against drug plasma concentrations, direct relationships were observed that were well described by the sigmoidal maximal effect model. IUP IC(50) values for terazosin, doxazosin, tamsulosin, and fiduxosin were 48.6, 48.7, 0.42, and 261 ng/ml, respectively. MAP IC(50) values were 12.2, 13.8, 1.07, and 1904 ng/ml, respectively. Uroselectivity index values, defined as MAP IC(50)/IUP IC(50), were 0.25, 0.28, 2.6, and 7.3, respectively. These results extend previous observations with terazosin in this model, showing that doxazosin exhibits a uroselectivity index comparable to terazosin, consistent with the lack of alpha(1)-adrenoceptor subtype selectivity or uroselectivity of these drugs. Tamsulosin, an alpha(1a)-/alpha(1d)-subtype selective agent, had an index value approximately 10-fold greater than the nonselective drugs. Based on its pharmacokinetic profile and a relative uroselectivity 29-fold greater than the nonselective drugs, fiduxosin is expected to exhibit greater selectivity for urethral compared with vascular alpha(1)-adrenoceptors in human and should be a novel, long-acting, uroselective alpha(1)-adrenoceptor antagonist.

Fiduxosin is an alpha(1)-adrenoceptor antagonist with higher affinity for alpha(1A)-adrenoceptors and for alpha(1D)-adrenoceptors than for alpha(1B)-adrenoceptors. Our hypothesis is that such a compound with higher affinity for subtypes implicated in the control of lower urinary tract function and lower affinity for a subtype implicated in the control of arterial pressure could result in a superior clinical profile for the treatment of lower urinary tract symptoms suggestive of benign prostatic obstruction. The purpose of this study was to evaluate the potency and selectivity of fiduxosin for effects on prostatic intraurethral pressure (IUP) versus mean arterial pressure (MAP) relative to current clinical standards, terazosin and tamsulosin, in conscious dogs. Phenylephrine (PE)-induced increases in IUP and MAP were determined before and at various time points after an oral dose of each antagonist. Hypotensive potency was also determined. All three antagonists caused dose- and time-dependent blockade of the IUP and MAP pressor effects of PE. The IUP ED(50) values of fiduxosin, tamsulosin, and terazosin were 0.24, 0.004, and 0.23 mg/kg p.o., respectively. The corresponding MAP ED(50) values were 1.79, 0.006, and 0.09 mg/kg p.o. The rank order of IUP selectivity (ratio) was fiduxosin (7.5-fold), tamsulosin (1.5-fold), and terazosin (0.4 = 2.5-fold MAP-selective). Tamsulosin and terazosin caused dose-dependent hypotension, whereas no change in arterial pressure was seen after fiduxosin. These data, illustrating a superior selectivity profile of fiduxosin, are consistent with our hypothesis.

Benign prostatic hyperplasia (BPH), common in aging males, is often treated with alpha(1)-adrenoceptor antagonists. To minimize hypotensive and other side effects, compounds with selective antagonist activity at alpha(1A)- and alpha(1D)- (compared with alpha(1B)-) adrenoceptors were evaluated that would block lower urinary tract alpha(1)-adrenoceptors in preference to cardiovascular alpha(1B)-adrenoceptors. Fiduxosin (3-[4-((3aR,9bR)-cis-9-methoxy-1,2,3,3a,4,9b-hexahydro-[1]-benzopyrano[3,4-c]pyrrol-2-yl)butyl]-8-phenyl-pyrazino[2',3':4,5] thieno-[3,2-d]pyrimidine-2,4(1H,3H)-dione; ABT-980) was tested in radioligand binding assays, isolated tissue bioassays, intraurethral pressure (IUP) tests in isoflurane-anesthetized dogs, and blood pressure analyses in spontaneously hypertensive rats (SHR). Fiduxosin had higher affinity for cloned human alpha(1a)- (0.16 nM) and alpha(1d)-adrenoceptors (0.92 nM) in radioligand binding studies compared with alpha(1b)-adrenoceptors (25 nM) or in isolated tissue bioassays [pA(2) values of 8.5-9.6 for alpha(1A)-receptors in rat vas deferens or canine prostate strips, 8.9 at alpha(1D)-adrenoceptors (rat aorta), compared with 7.1 at alpha(1B)-adrenoceptors (rat spleen)]. Furthermore, the compound antagonized putative alpha(1L)-adrenoceptors in the rabbit urethra (pA(2) value of 7.58). Fiduxosin blocked epinephrine-induced increases in canine IUP (pseudo-pA(2) value of 8.12), eliciting only transient decreases in mean arterial blood pressure (MAP) in SHR. The area under the curve (AUC(0-->60) min) for the hypotensive response was dose related with a log index value for fiduxosin of 5.23, indicating a selectivity of 770-fold comparing IUP to MAP effects. Preferential antagonism of alpha(1A)- and alpha(1D)- versus alpha(1B)-adrenoceptors in vitro, blockade of putative alpha(1L)-sites in vitro, and selective effects on lower urinary tract function versus blood pressure in vivo by fiduxosin suggest the potential utility of this compound for the treatment of BPH.

A-204176 (N-[5-(1H-imidazol-4-y1)-5,6,7,8-tetrahydro-1-naphthalenyl]methanesulfonamide) is a potent and selective alpha1A adrenoceptor agonist that binds with 17-fold and 9-fold greater affinity to the alpha1A (Ki=176 nM) than the alpha1b and alpha1d subtypes, respectively. In functional studies A-204176 is potent (pD2=6.4) and efficacious (83% of maximum control phenylephrine response) at rabbit urethra alpha1A receptors, with weaker potency and greatly reduced efficacy at rat spleen alpha1B (pD2=5.3, 11%) and rat aorta alpha1D (pD2=4.4, 10%) subtypes. In anesthetized female dogs, A-204176 is more potent than the non-selective alpha1 adrenoceptor agonist phenylpropanolamine (PPA) to increase measures of urethral tone and is more efficacious to increase pressure in the proximal region of the urethra. Significant increases on parameters of the urethral pressure profilometry were induced at 100 and 300 nmol/kg, i.v., by A-204176 and PPA, respectively. A-204176 was more potent than PPA to increase the abdominal pressure required to produce leakage. In the simultaneous measurement of intraurethral pressure and mean arterial blood pressure, A-204176 displays enhanced urethral selectivity relative to PPA. However, despite its selectivity for alpha1A versus alpha1B and alpha1D adrenoceptors in vitro, A-204176 did not display the degree of urethral selectivity in vivo that would have been expected. The observed effect of A-204176 on blood pressure may be due to the presence of extra-synaptic alpha1A adrenoceptors in the vasculature or to activation of spinal and supraspinal alpha1A adrenoceptors. These data indicate that A-204176 may represent a useful pharmacological tool to investigate the functional role of the alpha1A adrenoceptor in the urethra and to elucidate the lack of uroselectivity observed in vivo.

Purpose: We describe and compare the usefulness of 3 minimally invasive dog urethral function models to demonstrate the efficacy potential of alpha 1 agonists for stress urinary incontinence. From this overall composite dataset the efficacy profiles of the alpha 1A selective agonist A-61603 and the active metabolite of midodrine ST-1059 were specifically compared., Materials and Methods: Isoflurane anesthetized multiparous female beagle dogs were used in all studies. Intraurethral pressure was measured using a 7Fr balloon catheter. Profilometry was performed using an 8Fr Millar transducer catheter. Bladder pressure required to produce leakage in response to external abdominal ballottements of increasing intensity was measured using a 5Fr transurethral catheter. Agonist responses were measured before and after increasing cumulative intravenous doses of each compound in each test., Results: Agonist induced increases in maximal urethral closure and leak point pressure strongly correlated in linear fashion (R(2) = 0.94), as did measurements of agonist induced increases in proximal intraurethral pressure using the microtransducer or balloon catheter technique (R(2) = 0.87). A dose of 0.01 to 1 nmol./kg. A-61603 and 10 to 1,000 nmol./kg. ST-1059 intravenously each caused dose dependent increases in maximum urethral closure, leak point and intraurethral pressure., Conclusions: While the dose range for which alpha 1 agonists affect urethral pressure was adequately predicted by any of the 3 methods used, the leak point pressure assay described has the advantage of being a dynamic test that directly evaluates efficacy to protect against leakage caused by increases in abdominal pressure. This leak point pressure test appears be useful for the preclinical evaluation of compounds used to treat stress urinary incontinence.

In search of a uroselective alpha1A subtype selective antagonist, a novel series of 6-OMe hexahydrobenz[e]isoindoles attached to a bicyclic heterocyclic moiety via a two-carbon linker was synthesized. It was found that in contrast to the previously described series of tricyclic heterocycles,(1) this bicyclic series has very specific requirements for the heterocyclic attachments. The most important structural features contributing to the alpha1A/alpha1B selectivity of these compounds were identified. In vitro functional assays for the alpha1 adrenoceptor subtypes were used to further characterize the most selective compounds, and in vivo models of vascular vs prostatic tone were used to assess uroselectivity. Compound 48 showed the highest degree of selectivity in the radioligand binding assays (56-fold), in the in vitro functional tests (80-fold), and for in vivo prostate selectivity (960-fold).

In search of a uroselective agent that exhibits a high level of selectivity for the alpha(1A) receptor, a novel series of tricyclic hexahydrobenz[e]isoindoles was synthesized. A generic pharmacophoric model was developed requiring the presence of a basic amine core and a fused heterocyclic side chain separated by an alkyl chain. It was shown that the 6-OMe substitution with R, R stereochemistry of the ring junction of the benz[e]isoindole and a two-carbon spacer chain were optimal. In contrast to the highly specific requirements for the benz[e]isoindole portion and linker chain, a wide variety of tricyclic fused heterocyclic attachments were tolerated with retention of potency and selectivity. In vitro functional assays for the alpha(1) adrenoceptor subtypes were used to further characterize these compounds, and in vivo models of vascular vs prostatic tone were used to assess uroselectivity.

A-131701 (3-[2-((3aR,9bR)-cis-6-methoxy-2,3,3a,4,5,9b, hexahydro-[1H]-benz[e]isoindol-2-yl)ethyl]pyrido [3',4': 4,5]thieno[3,2-d]pyrimidine-2,4(1H,3H)-dione) is a novel compound previously shown to be selective for alpha-1a sites compared with alpha-1b adrenoceptors in radioligand binding studies and isolated tissue bioassays and to block canine urethral pressure (IUP) responses to exogenous alpha-1 adrenergic agonists to a greater extent than blood pressure responses. In conscious dogs in which IUP and mean arterial blood pressure (MABP) responses were measured periodically up to 24 hr, A-131701 blocked phenylephrine (PHE)-induced increases in IUP to a greater extent than MABP responses, and the blockade of the IUP effects of PHE was significantly different from control for up to 12 hr after doses greater than 0.3 mg/kg p.o., whereas blood pressure effects were of a lesser extent and duration. In addition to the weak antagonism of PHE-induced blood pressure responses, A-131701 also exhibited minimal effects on basal blood pressure in the dog, unlike terazosin, doxazosin or tamsulosin. Pharmacokinetic analysis of plasma samples from dogs indicated that A-131701 had a half-life of 0.4 to 0.8 hr and a bioavailability of 30 to 50% in dogs. Somewhat longer half-lives were observed in rat and monkey, with bioavailability values in the 25 to 30% range. Evidence of nonlinearity of pharmacokinetics was obtained in dogs and monkeys. Pharmacodynamic analysis revealed differences between A-131701 and nonselective alpha-1 adrenoceptor antagonists in selectivity for prostatic versus vascular alpha-1 adrenoceptors based on either extent or duration of blockade, which were either similar to or superior to compounds such as tamsulosin or REC 15/2739. These data demonstrate that A-131701 selectively blocks canine prostatic alpha-1 adrenoceptors for prolonged periods compared with MABP responses in vivo. Therefore, A-131701 should have clinical utility in the pharmacotherapy of benign prostatic hyperplasia.

A-74283, (+,-)trans-2-(4-(3a,4,4a,6a,7,7a-hexahydro-4,7-etheno-1 H cyclobut [f] isoindol-1,3-dionyl)-butyl)-9-methoxy-2,2,2a,4,5,9b-hexahydr o-1 H-benz[e]isoindol HC1, was studied in receptor binding assays and in the spontaneously hypertensive rat (SHR). In radioligand binding to rat cortex, A-74283 had high affinity (equipotent to 8-OH-DPAT) and high selectivity for 5HT1A receptors compared to 5HT1B sites. In conscious SHR, A-74283 lowered mean arterial pressure (MAP) in a dose-related fashion with a prolonged effect after oral administration of higher doses, but heart rate (HR) was not changed. In anesthetized SHR, i.v. administration of A-74283 decreased MAP and total peripheral resistance, but not cardiac output. Pretreatment of conscious SHR with the selective 5TH1A receptor antagonists spiroxatrine or BMY 7378 reduced the hypotensive effect of A-74283 significantly, but pretreatment with adrenergic antagonists phenoxybenzamine or idazoxan or the 5HT2 receptor blocker ketanserin did not alter the effect of A-74283. Intracisternal administration of A-74283 also decreased MAP; however, A-74283 had no effect on blood pressure in pithed SHR in which blood pressure was supported with vasopressin, in contrast to nitroprusside. These data demonstrate that A-74283 exerts a potent hypotensive effect in SHR via systemic vasodilation originating from a central 5HT1A receptor mechanism. A-74283 may be useful for studying 5HT1A receptors and cardiovascular function.

The purpose of this study was to determine the potency and selectivity of the alpha-1 adrenergic receptor antagonist terazosin based on relationships between plasma concentrations and blockade of intraurethral pressure (IUP) and mean arterial pressure (MAP) responses after single dosing and to determine cumulative effects after repeated dosing. To this end, the relationships between plasma concentrations and blockade effects of terazosin on phenylephrine (PE)-induced IUP and MAP were evaluated in conscious male beagle dogs after single (0.1, 0.3 and 1 mg/kg) and repeated (0.3 and 1 mg/kg) daily oral dosing of terazosin. Blockade effects and plasma concentrations were evaluated at selected times for periods of < or = 24 hr. Terazosin produced dose-dependent antagonism of PE-induced IUP and MAP responses. When IUP and MAP blockade effects were plotted against terazosin plasma concentration, direct relationships were observed that were well described by the sigmoidal maximal effect model and resulted in IUP and MAP IC50 values of 48.6 and 12.2 ng/ml, respectively. Repeated daily dosing resulted in little accumulation of terazosin in plasma and demonstrated consistent blockade responses over 7 days. MAP blockade was observed up to 23 hr after terazosin administration, whereas IUP blockade returned to control levels before 23 hr. Combined pharmacokinetic/pharmacodynamic analysis showed no selective antagonism of IUP by terazosin but may provide a useful way to show uroselectivity of novel agents.

In this study, we used a novel conscious dog model to evaluate the uroselectivity of selected alpha 1-antagonists either approved for human use or in clinical development for the treatment of symptomatic benign prostatic hyperplasia (BPH) and compared those results to their in vitro binding and functional affinities at alpha 1A, alpha 1B and alpha 1D receptor subtypes. Conscious dogs were instrumented acutely with a balloon catheter for the measurement of changes in prostatic intraurethral pressure (IUP) and chronically with implantable telemetry devices for the measurement of arterial pressure. The pressor effects of the alpha 1-agonist phenylephrine (PE) on IUP and mean arterial pressure (MAP) were compared before and at various time points after oral doses of either terazosin, doxazosin, tamsulosin or Rec 15/2739 (SB 216469). At submaximal doses, terazosin and doxazosin blocked PE-induced increases in MAP to a greater extent than increases in IUP. Tamsulosin blocked both parameters equally at the lowest and highest doses; however, at the intermediate dose, IUP was blocked more than MAP. Rec 15/2739 at each dose always blocked IUP to a greater extent than MAP. While the in vivo uroselectivity of these agents was predicted by radioligand binding and in vitro functional selectivity for the alpha 1A subtype over alpha 1B and alpha 1D subtypes, results from conscious dog experiments indicate that estimates of in vivo uroselectivity also depend upon dose and the time after administration. Our conscious canine model provides the basis for frequent and repeated evaluation of uroselectivity parameters over many hours, thus providing a pharmacological profile of compound effects perhaps more relevant to clinical practice.

We characterize the in vitro and in vivo pharmacology of CHIR 2279, an N-substituted glycine peptoid previously identified from a combinatorial library as a novel ligand to alpha 1-adrenoceptors. Competitive receptor-binding assays with [3H]prazosin showed that CHIR 2279 was similar to prazosin in binding to alpha 1A (rat submaxillary), alpha 1a, alpha 1b, and alpha 1 d (cDNA expressed in LTK- cells) with high and approximately equipotent affinity. Ki values for CHIR 2279 ranged from 0.7 to 3 nM, and were 10-fold weaker than with prazosin. Functional assays for postsynaptic alpha 1-adrenoceptors showed CHIR 2279 was approximately equipotent in antagonizing agonist-induced contractile responses with rat was deferens (alpha 1A), canine prostate (alpha 1A), rat spleen (alpha 1B) and rat aorta (alpha 1D). The pA2 for CHIR 2279 averaged 7.07 in these assays, indicating a 10- to 100-fold lower in vitro potency than prazosin. In dogs, CHIR 2279 antagonized the epinephrine-induced increase in intraurethal pressure (pseudo pA2, 6.86) and in rats antagonized the phenylephrine-induced increase in mean arterial blood pressure. In rats and guinea pigs, CHIR 2279 induced a dose-dependent decrease in mean arterial blood pressure without eliciting the tachycardia commonly observed with other alpha 1-blockers. Pharmacokinetic/pharmacodynamic modeling showed the i.v. system clearance rate of CHIR 2279 was 60 and 104 ml/min/kg in rats and guinea pigs, respectively, and the in vivo potency for mean arterial blood pressure reduction was twice as great in guinea pigs (EC50, 520 ng/ml) than rats (EC50, 1170 ng/ml).

N-[5-(4,5-dihydro-1H-imidazol-2yl)-2-hydroxy-5,6,7,8-tetrahydro naphthalen-1-yl] methanesulfonamide hydrobromide (A-61603) is a novel and potent alpha-adrenoceptor agonist. In radioligand binding assays, the compound is at least 35-fold more potent at alpha 1A/a receptors than at alpha 1b or alpha 1d sites. In fibroblast cells transfected with alpha 1a receptors, A-61603 more potently stimulates phosphoinositide hydrolysis than norepinephrine, and is antagonized by prazosin. A-61603 is less potent in cells transfected with alpha 1b or alpha 1d receptors. A-61603 is a potent agonist at alpha 1A receptors in rat vas deferens (200- to 300-fold more potent than norepinephrine or phenylephrine, respectively) and in isolated canine prostate strips (130- to 165-fold more potent than norepinephrine or phenylephrine, respectively). In contrast, A-61603 is only 40-fold more potent than phenylephrine at alpha 1B sites in rat spleen and 35-fold less potent at rat aortic, alpha 1D sites. In an in vivo dog model, A-61603 raises intraurethral prostatic tone to a greater extent than mean arterial blood pressure. A-61603 induces a pressor response in conscious rats at doses 50- to 100-fold lower than phenylephrine, and the response is not attenuated by pretreatment with CEC, whereas YM-617 causes a 100-fold shift in the response. These results indicate that A-61603 is a potent adrenergic agonist, selective for alpha 1A/a receptors, and may prove a useful probe for studies of adrenergic function and alpha 1 adrenoceptor regulation of physiological functions.

(+/-)-(1'R*,3R*)-3-phenyl-1-[(1',2',3',4'-tetrahydro-5',6'-methylene- dioxy-1'-naphthalenyl) methyl] pyrrolidine methanesulfonate (ABT-200) was evaluated in a number of biological tests to establish its pharmacological profile of activity. ABT-200 antagonized the uptake of [3H]-norepinephrine into synaptosomes of rat hypothalamus (IC50 = 841 nM) and blocked 4,alpha-dimethyl-m-tyramine- induced hypermotility in rats. In addition, ABT-200 potently inhibited binding of [3H]-rauwolscine to alpha-2 adrenergic receptors with a Ki value in radioligand binding assays of approximately 1 nM in the rat cortex and was much less potent at other receptor binding sites. ABT-200 antagonized alpha-2 receptors in vitro in the rat vas deferens and dog saphenous vein, where pA2 values of 7.7 and 8.2, respectively, were obtained. ABT-200 also antagonized clonidine-induced mydriasis and increased the overflow of [3H]-norepinephrine in guinea pig hippocampal slices, manifestations of blockade of alpha-2 adrenoceptors in the central nervous system. ABT-200 was active in antagonizing nocturnal hyperactivity in olfactory bulbectomized rats, a test for putative antidepressant activity. In cardiovascular studies, ABT-200 exhibited negligible activity in affecting hemodynamic parameters and was free of postural hypotensive activity. In both in vitro and in vivo, ABT-200 was devoid of antihistaminic or anticholinergic activity. This profile of activity of moderate inhibition of norepinephrine uptake with blockade of alpha-2 adrenoceptors suggests potential dual-action effects for ABT-200, which may represent a putative antidepressant with minimal cardiovascular side-effect liability.

ABBOTT-81988 (A-81988), 2-(N-propyl-N[(2'-[1H-tetrazol-5-yl]biphenyl- 4yl)methyl] amino) pyridine-3-carboxylic acid, a nonpeptide angiotensin II (AII) antagonist was studied in the conscious spontaneously hypertensive rate (SHR) (male, 18 to 21 weeks) for cardiovascular effects of oral administration. Oral A-81988 at 0.3 to 3 mg/kg produced a dose-related 10 to 29% decrease in mean arterial pressure (MAP) in SHR (control, 161 to 177 mm Hg; n = 19) for 12 to 24 h without changing heart rate. Oral A-81988 at 3 mg/kg daily maintained MAP in SHR at normotensive levels (97 to 120 mm Hg) during a 5-day protocol with no rebound hypertension at termination of treatment. There was an increase in plasma renin activity in nanograms AI/milliliter/hour in SHR treated with A-81988 (32 +/- 3, n = 6 v 5 +/- 2, n = 6 for vehicle) during its antihypertensive action. The oral potency of A-81988 was enhanced about 10-fold in furosemide-treated SHR. The pressor response to AII was inhibited selectively in SHR even after an 8-day treatment with A-81988 (approximately 3 mg/kg/day orally). Total peripheral resistance was lowered and cardiac output unchanged in SHR administered A-81988 (3 mg/kg/day orally for 2 days). A-81988 (3 mg/kg orally) did not cause orthostatic hypotension in SHR.(ABSTRACT TRUNCATED AT 250 WORDS)

2-(N-Propyl-N[(2'-[1H-tetrazol-5-yl]biphenyl-4yl)methyl]amin o) pyridine-3-carboxylic acid (ABBOTT-81988), a novel nonpeptide angiotensin II (AII) antagonist, was evaluated to characterize its antihypertensive activity in the conscious renal hypertensive rat. Oral or i.v. administration of ABBOTT-81988 at 0.03 to 0.3 mg/kg produced a dose-dependent, sustained decrease in mean arterial pressure (MAP; control 162-173 mm Hg, n = 27) of approximately 20 to 70 mm Hg. At a dose of 0.3 mg/kg p.o., ABBOTT-81988 lowered MAP to a normotensive level for more than 24 hr and did not change heart rate. During its antihypertensive effect (delta MAP, -28% approximately -35%), ABBOTT-81988 (0.1-03 mg/kg i.v.) decreased total peripheral resistance (delta resistance, -31% approximately -43%), and cardiac output remained either unchanged or slightly elevated. ABBOTT-81988 (0.3 mg/kg i.v.) produced an additional antihypertensive effect (delta MAP, -12 +/- 2%, n = 5) in captopril-pretreated (10 mg/kg i.v.) hypertensive rats, but captopril (10 mg/kg i.v.) had no effect in ABBOTT-81988-pretreated (0.3 mg/kg i.v.) rats. In the normotensive rat, ABBOTT-81988 (0.3 mg/kg p.o.) had no effect on basal MAP, but it inhibited the AII-induced (0.1 microgram/kg i.v.) pressor response by 51% to 91% for 24 hr, whereas the responses to norepinephrine (0.3 microgram/kg i.v.), vasopressin (0.03 IU/kg i.v.) and bradykinin (3 micrograms/kg i.v.) were not affected. It is concluded that ABBOTT-81988 is a safe and efficacious AII antagonist that may have use in the treatment of human hypertension.

A series of pyridines and other six-membered ring heterocycles connected to a biphenyltetrazole with a -CH2-NR'-link (1) were discovered to be potent angiotensin II antagonists. In the pyrimidine carboxylic acid series (W = CR, X = N, Y = CH, Z = COOH), compounds with an alkyl group (R') on the exocyclic nitrogen were much more potent than compounds with an alkyl group (R) on the heterocyclic ring. The corresponding pyridine, pyridazine, pyrazine, and 1,2,4-triazine carboxylic acids also showed potent in vitro angiotensin II antagonism. The pyridine (W, X, Y = CH, Z = COOH, R' = n-C3H7) demonstrated potent in vitro activity (pA2 = 10.10, rabbit aorta, and Ki = 0.61 nM, receptor binding in rat liver) as well as exceptional oral antihypertensive activity and bioavailability. Any nonacidic replacement for the carboxylic acid was detrimental for activity.

ABBOTT-81282, 4-(N-butyl-N-[(2-'-[1H-tetrazol-5yl]biphenyl-4-yl)methyl]ami no) pyrimidine-5-carboxylic acid is a novel nonpeptide angiotensin II (AII) antagonist. In vivo studies were performed to evaluate ABBOTT-81282 for its antihypertensive effect, pharmacological mechanism(s) of action, and cardiovascular safety. In the conscious renal artery-ligated (RAL) hypertensive rat, a model of high renin hypertension, ABBOTT-81282 (1-10 mg/kg p.o. and 0.1-1.0 mg/kg i.v.) lowered mean arterial pressure (MAP) in a dose-dependent manner with the ED30 values of 2.2 mg/kg for p.o. administration and 0.08 mg/kg for i.v. administration. At 10 mg/kg p.o., ABBOTT-81282 lowered blood pressure in the RAL rat (delta MAP 66 +/- 9 mm Hg from control MAP 167 +/- 7 mm Hg, n = 6) to a normotensive level (MAP, 115 +/- 5 mm Hg) for greater than 24 h and did not change heart rate. The i.v. administration of 1 mg/kg of ABBOTT-81282 also produced a sustained, long-lasting decrease (delta MAP 27-52 mm Hg) in blood pressure that was significantly different from the vehicle group at 8 h postdosing (143 +/- 3 mm Hg, n = 4 for ABBOTT-81282 vs. 181 +/- 3 mm Hg, n = 6 for vehicle group, p < 0.01). When blood pressure in the renal hypertensive rat was maximally lowered (delta MAP 72 +/- 9 mm Hg, n = 4) following the 1 mg/kg i.v. dose (cumulative) of ABBOTT-81282, additional administration of captopril (3 mg/kg i.v.) produced no further decline in blood pressure. In the conscious normotensive rat, 10 mg/kg p.o. of ABBOTT-81282 had no effect on basal MAP (119 +/- 3 vs. 115 +/- 4 mm Hg, pre- vs. 3.5 h postdosing, n = 4) and heart rate (364 +/- 18 vs. 363 +/- 14 beats/min, pre- vs. 3.5 h postdosing, n = 4) but inhibited the AII (0.1 micrograms/kg i.v.)-induced increase in MAP by 64-70%, while the MAP responses to norepinephrine (0.3 micrograms/kg i.v.), vasopressin (0.03 IU/kg i.v.) and bradykinin (3 micrograms/kg i.v.) remained intact. ABBOTT-81282 was also administered to conscious normotensive rats (n = 4) instrumented with ECG telemetry transmitters. At an i.v. dose of 10 mg/kg, which is 125 times greater than the i.v. ED30, ABBOTT-81282 caused a minimal decrease (< 14%) in MAP and had no effect on ECG waveforms. These data demonstrate that ABBOTT-81282 is a safe and efficacious antihypertensive agent with selective AII antagonism.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Studies were performed to evaluate whether chronic prazosin treatment alters the alpha 2-adrenoceptor function for orthostatic control of arterial blood pressure in conscious spontaneously hypertensive rats (SHR). 2. Conscious SHR (male 300-350 g) were subjected to 90 degrees head-up tilts for 60 s following acute administration of prazosin (0.1 mg kg-1 i.p.) or rauwolscine (3 mg kg-1 i.v.). Orthostatic hypotension was determined by the average decrease (%) in mean arterial pressure (MAP femoral) over the 60-s tilt period. The basal MAP of conscious SHR was reduced to a similar extent by prazosin (-23%(-)-26% MAP) and rauwolscine (-16%(-)-33% MAP). However, the head-up tilt induced orthostatic hypotension in the SHR treated with prazosin (-16% MAP, n = 6), but not in the SHR treated with rauwolscine (less than +2% MAP, n = 6). 3. Conscious SHR were treated for 4 days with prazosin at 2 mg kg-1 day-1 i.p. for chronic alpha 1-adrenoceptor blockade. MAP in conscious SHR after chronic prazosin treatment was 14% lower than in the untreated SHR (n = 8). Head-up tilts in these rats did not produce orthostatic hypotension when performed either prior to or after acute dosing of prazosin (0.1 mg kg-1 i.p.). Conversely, administration of rauwolscine (3 mg kg-1 i.v.) in chronic prazosin treated SHR decreased the basal MAP by 12-31% (n = 4), and subsequent tilts induced further drops of MAP by 19-23% in these rats. 4. The pressor responses and bradycardia to the alpha 1-agonist cirazoline (0.6 and 2 micrograms kg-1 i.v.), the alpha 2-agonist Abbott-53693 (1 and 3 micrograms kg-1 i.v.), and noradrenaline (0.1 and 1.0 micrograms kg-1 i.v.) were determined in conscious SHR with and without chronic prazosin pretreatment. Both the pressor and bradycardia effects of cirazoline were abolished in chronic prazosin treated SHR (n = 4) as compared to the untreated SHR (n = 4). On the other hand, the pressor effects of Abbott-53693 were similar in both groups of SHR, but the accompanying bradycardia was greater in SHR with chronic prazosin treatment than without such treatment. Furthermore, the bradycardia that accompanied the noradrenaline-induced pressor effect in SHR was similar with and without chronic prazosin treatment despite a 47-71% reduction of the pressor effect in chronic alpha 1-receptor blocked SHR.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Calcium mediated contractile effects were evoked in phenoxybenzamine (10(-7) M)-pretreated dog saphenous vein (DSV) rings under the continuing presence of norepinephrine (3 x 10(-5) M), a condition that selectively activates alpha 2 (alpha 2)-adrenergic receptors. 2. Calcium-induced contractions in this model were dose-dependently inhibited by the alpha 2 antagonists rauwolscine and SKF-86466, while the calcium channel blockers nifedipine and verapamil produced only small non-dose related inhibitions. The receptor antagonists sulpiride, buspirone, pyrilamine and atropine showed no inhibitory effect. 3. In contrast, rauwolscine and SKF-86466 produced no inhibition of the calcium-evoked contractions in potassium-depolarized DSV rings, in which nifedipine and verapamil exhibited potent calcium antagonist activity. 4. These data provide evidence to support the presence of alpha 2 related receptor-operated calcium channels in vascular smooth muscle, which are pharmacologically distinguishable from potential-dependent calcium channels.