Structure-activity studies of novel cyanoguanidine ATP-sensitive potassium channel openers for the treatment of overactive bladder.

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.

Effects of substitution on 9-(3-bromo-4-fluorophenyl)-5,9-dihydro-3H,4H-2,6-dioxa-4- azacyclopenta[b]naphthalene-1,8-dione, a dihydropyridine ATP-sensitive potassium channel opener.

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.

Pharmacological characterization of urinary bladder smooth muscle contractility following partial bladder outlet obstruction in pigs.

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.

Pharmacological characterization of the novel dihydropyridine potassium channel opener, (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), and the regulation of cardiovascular function in conscious and anesthetized beagle dogs.

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.

In vitro and in vivo characterization of a novel naphthylamide ATP-sensitive K+ channel opener, A-151892.

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.

Synthesis and structure-activity relationships of a novel series of 2,3,5,6,7,9-hexahydrothieno[3,2-b]quinolin-8(4H)-one 1,1-dioxide K(ATP) channel openers: discovery 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 potent K(ATP) opener that selectively inhibits spontaneous bladder contractions.

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.

Synthesis and structure-activity relationships of a novel series of tricyclic dihydropyridine-based KATP openers that potently inhibit bladder contractions in vitro.

Structure-activity relationships were investigated on a novel series of tricyclic dihydropyridine-containing K(ATP) openers. This diverse group of analogues, comprising a variety of heterocyclic rings fused to the dihydropyridine nucleus, was designed to determine the influence on activity of hydrogen-bond-donating and -accepting groups and their stereochemical disposition. Compounds 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 subset of compounds. All compounds studied showed greater potency to inhibit spontaneous bladder contractions relative to their potencies to inhibit contractions elicited by electrical stimulation.

Synthesis and structure-activity studies on N-[5-(1H-imidazol-4-yl)-5,6,7,8-tetrahydro-1-naphthalenyl]methanesulfonamide, an imidazole-containing alpha(1A)-adrenoceptor agonist.

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.

Functional characterization of large conductance calcium-activated K+ channel openers in bladder and vascular smooth muscle.

Calcium activated K(+) channels (K(Ca) channels) are found in a variety of smooth muscle tissues, the most characterized of which are the large conductance K(Ca) channels (BK(Ca) or maxi-K(+) channels). Recent medicinal chemistry efforts have identified novel BK(Ca) openers including 2-amino-5-(2-fluoro-phenyl)-4-methyl-1H-pyrrole-3-carbonitrile (NS-8), BMS-204352 and its analog 3-(5-chloro-2-hydroxy-phenyl)-3-hydroxy-6-trifluoromethyl-1,3-dihydro-indol-2-one (compound 1), and 5,7-dichloro-4-(5-chloro-2-hydroxy-phenyl)-3-hydroxy-1H-quinolin-2-one (compound 2). Although these compounds are effective BK(Ca) openers as shown by electrophysiological methods, little is known about their effects on smooth muscle contractility. In this study, the responsiveness of structurally diverse BK(Ca) openers-NS-8, compounds 1 and 2 and the well characterized nonselective NS-1619-was assessed using segments of endothelium denuded rat aorta, rat and guinea pig detrusor precontracted with extracellular K(+), and Landrace pig detrusor stimulated by electrical field. In all preparations, the compounds tested inhibited or completely abolished contractions with similar potencies (-logIC(50) values: 3.8 to 5.1). In rat aorta, in the presence of 80 mM K(+), the compounds significantly shifted the concentration-response curve to the right compared with those obtained in 30 mM K(+). These data are consistent with K(+) channel (BK(Ca) channel) activation as the underlying mechanism of relaxation by compounds that share the electrophysiological property of BK(Ca) current activation. The similar potencies at detrusor and vascular smooth muscle suggest that the achievement of smooth muscle selectivity in vitro with the representative compounds examined in this study may prove to be a challenge when targeting BK(Ca) channels for smooth muscle indications such as overactive bladder.

Structure-activity studies for a novel series of tricyclic dihydropyridopyrazolones and dihydropyridoisoxazolones as K(ATP) channel openers.

In search of a novel chemotype of K(ATP) channel openers a series of tricyclic dihydropyridopyrazolones and dihydropyridoisoxazolones was synthesized. It was found that cyclopentanone in the left hand portion of the molecule was 4-fold more potent than cyclohexanone. Introduction of gem-dimethyl groups as well as incorporation of oxygen in the cyclohexanone ring in the left hand portion of the molecule increased the potency 10-fold. In the right hand portion of the molecule, the NH-group of the pyrazolone can be effectively substituted by oxygen increasing the activity 5-fold. Incorporation of a methyl group adjacent to the dihydropyridine (DHP) nitrogen not only significantly boosted activity, but also provided an additional benefit of increased metabolic stability. In vitro tests on the tissue from pig bladder strips provided further confirmation of K(ATP) activity of these compounds.

Design and synthesis of novel cyanoguanidine ATP-sensitive potassium channel openers for the treatment of overactive bladder.

Thiourea derivatives were identified as glyburide-reversible potassium channel openers through high-throughput screening. Based on these findings, a number of novel cyanoguanidines were designed and synthesized, which hyperpolarized human bladder K(ATP) channels. These agents are potent full agonists in relaxing electrically-stimulated pig bladder strips. The synthesis, SAR and biological properties of these agents are discussed.

[125I]A-312110, a novel high-affinity 1,4-dihydropyridine ATP-sensitive K+ channel opener: characterization and pharmacology of binding.

Although ATP-sensitive K+ channels continue to be explored for their therapeutic potential, developments in high-affinity radioligands to investigate native and recombinant KATP channels have been less forthcoming. This study reports the identification and pharmacological characterization of a novel iodinated 1,4-dihydropyridine KATP channel opener, [125I]A-312110 [(9R)-9-(4-fluoro-3-125iodophenyl)-2,3,5,9-tetrahydro-4H-pyrano[3,4-b]thieno[2,3-e]pyridin-8(7H)-one-1,1-dioxide]. Binding of [125I]A-312110 to guinea pig cardiac (KD = 5.8 nM) and urinary bladder (KD = 4.9 nM) membranes were of high affinity, saturable, and to a single set of binding sites. Displacement of [125I]A-312110 by structurally diverse potassium channel openers (KCOs) indicated a similar rank order of potency in both guinea pig cardiac and bladder membranes (Ki, heart): A-312110 (4.3 nM) > N-cyano-N'-(1,1-dimethylpropyl)-N"-3-pyridylguanidine (P1075) > (-)-N-(2-ethoxyphenyl)-N'-(1,2,3-trimethylpropyl)-2-nitroethene-1,1-diamine (Bay X 9228) > pinacidil > (-)-cromakalim > N-(4-benzoyl phenyl)-3,3,3-trifluro-2-hydroxy-2-methylpropionamine (ZD6169) > 9-(3-cyanophenyl)-3,4,6,7,9,10-hexahydro-1,8-(2H,5H)-acridinedione (ZM244085) >> diazoxide (16.7 microM). Displacement by KATP channel blockers, the sulfonylurea glyburide, and the cyanoguanidine N-[1-(3-chlorophenyl)cyclobutyl]-N'-cyano-N"-3-pyridinyl-guanidine (PNU-99963) were biphasic in the heart but monophasic in bladder with about a 100- to 500-fold difference in Ki values between high- and low-affinity sites. Good correlations were observed between cardiac or bladder-binding affinities of KCOs with functional activation as assessed by their respective potencies to either suppress action potential duration (APD) in Purkinje fibers or to relax electrical field-stimulated bladder contractions. Collectively, these results demonstrate that [125I]A-312110 binds with high affinity and has an improved activity profile compared with other radiolabeled KCOs. [125I]A-312110 is a useful tool for investigation of the molecular and functional properties of the KATP channel complex and for the identification, in a high throughput manner, of both novel channel blockers and openers that interact with cardiac/smooth muscle-type KATP channels.

The discovery of a new class of large-conductance Ca2+-activated K+ channel opener targeted for overactive bladder: synthesis and structure-activity relationships of 2-amino-4-azaindoles.

2-Amino-4-azaindoles have been identified as a structurally novel class of BK(Ca) channel openers. Their synthesis from 2-chloro-3-nitropyridine is described together with their in vitro properties assessed by 86Rb(+) efflux and whole-cell patch-clamp assays using HEK293 cells stably transfected with the BK(Ca) alpha subunit. In vitro functional characterization of BK(Ca) channel opening activity was also assessed by measurement of relaxation of smooth muscle tissue strips obtained from Landrace pig bladders. The preliminary SAR data indicate the importance of steric bulk around the 2-amino substituent.

Structure-activity relationship of a novel class of naphthyl amide KATP channel openers.

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

Pharmacological characterization of a 1,4-dihydropyridine analogue, 9-(3,4-dichlorophenyl)-3,3,6,6-tetramethyl-3,4,6,7,9,10-hexahydro-1,8(2H,5H)-acridinedione (A-184209) as a novelK(ATP) channel inhibitor.

1. This study reports on the identification and characterization of a 1,4-dihydropyridine analogue, 9-(3,4-dichlorophenyl)-3,3,6,6-tetramethyl-3,4,6,7,9,10-hexahydro-1,8(2H,5H)-acridinedione (A-184209) as a novel inhibitor of ATP-sensitive K(+) channels. 2. A-184209 inhibited membrane potential changes evoked by the prototypical cyanoguanidine ATP-sensitive K(+) channel opener (KCO) P1075 in both vascular (A10) and urinary bladder smooth muscle cells with IC(50) values of 1.44 and 2.24 micro M respectively. 3. P1075-evoked relaxation of 25 mM K(+) stimulated aortic strips was inhibited by A-184209 in an apparently competitive fashion with a pA(2) value of 6.34. 4. The potencies of A-184209 to inhibit P1075-evoked decreases in membrane potential responses in cardiac myocytes (IC(50)=0.53 micro M) and to inhibit 2-deoxyglucose-evoked cation efflux pancreatic RINm5F cells (IC(50)=0.52 micro M) were comparable to the values for inhibition of smooth muscle K(ATP) channels. 5. On the other hand, a structural analogue of A-184209 that lacked the gem-dimethyl substituent, 9-(3,4-dichlorophenyl)-3,4,6,7,9,10-hexahydro-1,8(2H,5H)-acridinedione (A-184208), was found to be a K(ATP) channel opener, evoking membrane potential responses in A10 smooth muscle cells (EC(50)=385 nM) and relaxing aortic smooth muscle strips (IC(50)=101 nM) in a glyburide-sensitive manner. 6. Radioligand binding studies demonstrated that A-184209 displaced SUR1 binding defined by [(3)H]glyburide binding to RINm5F cell membranes with a K(i) value of 0.11 micro M whereas A-184208 was ineffective. On the other hand, both A-184209 (K(i)=1.34 micro M) and A-184208 (K(i)=1.14 micro M) displaced binding of the KCO radioligand, [(125)I]A-312110 in guinea-pig bladder membranes with similar affinities. 7. These studies demonstrate that A-184209 is a novel and structurally distinct compound that inhibits K(ATP) channels in smooth muscle with potencies comparable to glyburide. The structural overlap between DHP openers and blockers, together with their differential interaction with ligand binding sites, support the notion that both openers and blockers bind to similar or very closely coupled sites on the sulfonylurea receptor and that subtle changes in the pharmacophore itself could switch functional properties from K(ATP) channel activation to inhibition.

(-)-(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 ATP-sensitive potassium channel opener efficacious in suppressing urinary bladder contractions. I. In vitro characterization.

Alterations in the myogenic activity of the bladder smooth muscle are thought to serve as a basis for the involuntary detrusor contractions associated with the overactive bladder. Activation of ATP-sensitive K(+) (K(ATP)) channels has been recognized as a potentially viable mechanism to modulate membrane excitability in bladder smooth muscle. In this study, we describe the preclinical pharmacology 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 1,4-dihydropyridine K(ATP) channel opener (KCO) that demonstrates enhanced bladder selectivity for the suppression of unstable bladder contractions in vivo relative to other reference KCOs. A-278637 activated K(ATP) channels in bladder smooth muscle cells in a glyburide (glibenclamide)-sensitive manner as assessed by fluorescence membrane potential assays using bis-(1,3-dibutylbarbituric acid)trimethine oxonol (EC(50) = 102 nM) and by whole cell patch clamp. Spontaneous (myogenic) phasic activity of pig bladder strips was suppressed (IC(50) = 23 nM) in a glyburide-sensitive manner by A-278637. A-278637 also inhibited carbachol- and electrical field-stimulated contractions of bladder strips, although the respective potencies were 8- and 13-fold lower compared with inhibition of spontaneous phasic activity. As shown in the accompanying article [Brune ME, Fey TA, Brioni JD, Sullivan JP, Williams M, Carroll WA, Coghlan MJ, and Gopalakrishnan M (2002) J Pharmacol Exp Ther 303:387-394], A-278637 suppressed myogenic contractions in vivo in a model of bladder instability with superior selectivity compared with other KCOs, WAY-133537 [(R)-4-[3,4-dioxo-2-(1,2,2-trimethyl-propylamino)cyclobut-1-enylamino]-3-ethyl-benzonitrile] and ZD6169 [(S)-N-(4-benzoylphenyl)3,3,3-trifluro-2hydroxy-2-methyl-priopionamide]. A-278637 did not interact with other ion channels, including L-type calcium channels or other neurotransmitter receptor systems. The pharmacological profile of A-278637 represents an attractive basis for further investigations of selective K(ATP) channel openers for the treatment of overactive bladder via myogenic etiology.

N-[3-(1H-imidazol-4-ylmethyl)phenyl]ethanesulfonamide (ABT-866, 1),(1) a novel alpha(1)-adrenoceptor ligand with an enhanced in vitro and in vivo profile relative to phenylpropanolamine and midodrine.

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).

ABT-866, a novel alpha(1A)-adrenoceptor agonist with antagonist properties at the alpha(1B)- and alpha(1D)-adrenoceptor subtypes.

N-[3-(1H-Imidazol-4-ylmethyl)phenyl]ethanesulfonamide, maleate (ABT-866) is a novel alpha(1)-adrenoceptor agent with mixed pharmacological properties in vitro. Compared to phenylephrine, ABT-866 demonstrates intrinsic activity at the alpha(1A)-adrenoceptor subtype present in the rabbit urethra (pD(2) = 6.22, with 80% of the phenylephrine response), reduced intrinsic activity at the alpha(1B)-adrenoceptor subtype in the rat spleen (pD(2)= 6.16, with 11% of the phenylephrine response), and no intrinsic activity at the rat aorta alpha(1D)-adrenoceptor subtype. ABT-866 also demonstrated antagonism at the rat spleen alpha(1B)-adrenoceptor (pA(2) = 5.39 +/- 0.08, slope = 1.20 +/- 0.12), and the rat aorta alpha(1D)-adrenoceptor (pA(2)= 6.18 +/- 0.09, slope = 0.96 +/- 0.13). This is in contrast to the weak non-selective activity seen with the alpha(1)-adrenoceptor agonist, phenylpropanolamine (2-amino-1-phenyl-1-propanol hydrochloride), and the alpha(1A/D)-adrenoceptor selective agonist 1-(2',5'-dimethoxyphenyl)-2-aminoethanol hydrochloride (ST-1059), the active metabolite of midodrine, that has been used clinically for the treatment of stress urinary incontinence. This study identifies a unique agent that may prove to be a valuable in vivo tool in testing the hypothesis that the alpha(1A)-adrenoceptor can be stimulated to contract the smooth muscle present in the urethra without evoking blood pressure elevations presumably caused by alpha(1B)- and alpha(1D)-adrenoceptor subtype involvements in the vasculature.

Structure-Activity studies for a novel series of tricyclic dihydropyrimidines as K(ATP) channel openers (KCOs).

A novel series of tricyclic dihydropyrimidines was synthesized and evaluated for activity as K(ATP) channel openers. The functional activity of several compounds, for example 6A (EC(50)=30nM) and its enantiomers exceeded cromakalim.

Functional characterization of adenosine receptors and coupling to ATP-sensitive K+ channels in Guinea pig urinary bladder smooth muscle.

Although multiple adenosine receptors have been identified, the subtype and underlying mechanisms involved in the relaxation response to adenosine in the urinary bladder remain unclear. The present study investigates changes in the membrane potential, as assessed by fluorescence-based techniques, of bladder smooth muscle cells by adenosine receptor agonists acting via ATP-sensitive potassium (K(ATP)) channels. Membrane hyperpolarization evoked by adenosine and various adenosine receptor subtype-selective agonists was attenuated or reversed by the K(ATP) channel blocker glyburide. Comparison of adenosine receptor agonist potencies eliciting membrane potential effects showed a rank order of potency 5'-N-ethyl-carboxamido adenosine (NECA; -log EC50 = 7.97) approximately 2-p-(2-carboxethyl)phenethyl-amino-5'-N-ethylcarboxamidoadenosine hydrochloride (CGS-21680; 7.65) > 2-chloro adenosine (5.90) approximately 2-chloro-N6-cyclopentyladenosine (CCPA; 5.51) approximately N6-cyclopentyladenosine approximately N6-(R)-phenylisopropyladenosine > 2-chloro- N6-(3-iodobenzyl)-adenosine-5'-N-methyl-carboxamide (2Cl-IBMECA; 4.78). Membrane potential responses were mimicked by forskolin, a known activator of adenylate cyclase, and papaverine, a phosphodiesterase inhibitor. The A(2A)-selective antagonist 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-yl-amino] ethyl)phenol (ZM-241385), and the adenylate cyclase inhibitor N-(cis-2-phenyl-cyclopentyl) azacyclotridecan-2-imine-hydrochloride (MDL-12330A) inhibited the observed change in membrane potential evoked by adenosine and adenosine-receptor agonists. The rank order potency for relaxation of K+-stimulated guinea pig bladder strips, NECA (-log EC50 = 6.41) approximately CGS-21680 (6.38) > 2-chloro adenosine (5.90) >> CCPA approximately 2Cl-IBMECA (>4.0) was comparable to that obtained from membrane potential measurements. Collectively, these studies demonstrate that adenosine-evoked membrane hyperpolarization and relaxation of bladder smooth muscle is mediated by A(2A) receptor-mediated activation of K(ATP) channels via adenylate cyclase and elevation of cAMP.