Pharmacological characterization of AC-90179 [2-(4-methoxyphenyl)-N-(4-methyl-benzyl)-N-(1-methyl-piperidin-4-yl)-acetamide hydrochloride]: a selective serotonin 2A receptor inverse agonist

The primary purpose of the present series of experiments was to characterize the in vitro and in vivo pharmacology profile of 2-(4-methoxy-phenyl)-N-(4-methyl-benzyl)-N-(1-methyl-piperidin-4-yl)-acetamide hydrochloride (AC-90179), a selective serotonin (5-HT2A) receptor inverse agonist, in comparison with the antipsychotics haloperidol and clozapine. The secondary purpose was to characterize the pharmacokinetic profile of AC-90179. Like all atypical antipsychotics, AC-90179 shows high potency as an inverse agonist and competitive antagonist at 5HT2A receptors. In addition, AC-90179 exhibits antagonism at 5HT2C receptors. In contrast, AC-90179 does not have significant potency for D2 and H1 receptors that have been implicated in the dose-limiting side effects of other antipsychotic drugs. The ability of AC-90179 to block 5-HT2A receptor signaling in vivo was demonstrated by its blockade of the rate-decreasing effects of the 5-HT2A agonist, (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride, under a fixed ratio schedule of reinforcement. Similar to clozapine and haloperidol, AC-90179 attenuated phencyclidine-induced hyperactivity. Although haloperidol impaired acquisition of a simple autoshaped response and induced cataleptic-like effects at behaviorally efficacious doses, AC-90179 and clozapine did not. Furthermore, unlike haloperidol and clozapine, AC-90179 did not decrease spontaneous locomotor behavior at efficacious doses. Limited oral bioavailability of AC-90179 likely reflects rapid metabolism rather than poor absorption. Taken together, a compound with a similar pharmacological profile as AC-90179 and with increased oral bioavailability may have potential for the treatment of psychosis.     

ACP-103, a 5-hydroxytryptamine 2A receptor inverse agonist, improves the antipsychotic efficacy and side-effect profile of haloperidol and risperidone in experimental models

Dopamine D(2) receptor antagonism contributes to the therapeutic action of antipsychotic drugs (APDs) but also produces undesirable side effects, including extrapyramidal motor deficits, cognitive dulling, and prolactinemia. The introduction of atypical APDs was a significant advancement in the treatment of schizophrenia. Whereas these agents are D(2) receptor antagonists, they are also potent 5-hydroxytryptamine (5-HT)(2A) receptor inverse agonists, a feature that may explain their improved efficacy and tolerability. Recently, we reported that N-(4-fluorophenylmethyl)-N-(1-methylpiperidin-4-yl)-N'-(4-(2-methylpropyloxy)phenylmethyl) carbamide (2R,3R)-dihydroxybutanedioate (2:1) (ACP-103), a novel selective 5-HT(2A) receptor inverse agonist that fails to bind D(2) receptors, is active in several models predictive of antipsychotic activity. Using ACP-103, we tested the hypothesis that combining high levels of 5-HT(2A) inverse agonism with low levels of D(2) antagonism would result in a favorable interaction, such that antipsychotic efficacy could be achieved with reduced D(2) receptor-related adverse effects. Here we show that ACP-103 1) potently inhibited head-twitching produced by the 5-HT(2A/2C) receptor agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine, 2) increased the potency of haloperidol against amphetamine-induced hyperactivity, 3) interacted synergistically with haloperidol or risperidone to suppress hyperactivity induced by the N-methyl-d-aspartate receptor antagonist (5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801), and, by contrast, 4) attenuated haloperido-l- or risperidone-induced prolactinemia. ACP-103 also attenuated catalepsy produced by haloperidol or risperidone. However, the doses that were required for this effect were higher than would be expected for a 5-HT(2A) receptor-mediated mechanism. These data indicate that utilizing ACP-103 as an adjunctive therapy to currently used APDs may result in enhanced antipsychotic efficacy while reducing adverse effects including those attributable to D(2) receptor antagonism.     

Pharmacological profile of (2R-trans)-4-[1-[3,5-bis(trifluoromethyl)benzoyl]-2-(phenylmethyl)-4-piperidinyl]-N-(2,6-dimethylphenyl)-1-acetamide (S)-Hydroxybutanedioate (R116301), an orally and centrally active neurokinin-1 receptor antagonist

In comparison with a series of reference compounds, (2R-trans)-4-[1-[3,5-bis(trifluoromethyl)benzoyl]-2-(phenylmethyl)-4-piperidinyl]-N-(2,6-dimethylphenyl)-1-acetamide (S)-Hydroxybutanedioate (R116301) was characterized as a specific, orally, and centrally active neurokinin-1 (NK(1)) receptor antagonist with subnanomolar affinity for the human NK(1) receptor (K(i): 0.45 nM) and over 200-fold selectivity toward NK(2) and NK(3) receptors. R116301 inhibited substance P (SP)-induced peripheral effects (skin reactions and plasma extravasation in guinea pigs) and a central effect (thumping in gerbils) at low doses (0.08-0.16 mg/kg, s.c. or i.p.), reflecting its high potency as an NK(1) receptor antagonist and excellent brain disposition. Higher doses blocked various emetic stimuli in ferrets, cats, and dogs (ED(50) values: 3.2 mg/kg, s.c.; 0.72-2.5 mg/kg, p.o.). Even higher doses (11-25 mg/kg, s.c.) were required in mice (capsaicin-induced ear edema) and rats (SP-induced extravasation and salivation), consistent with lower affinity for the rodent NK(1) receptor and known species differences in NK(1) receptor interactions. R116301 inhibited the ocular discharge (0.034 mg/kg) but not the dyspnoea, lethality, or cough (>40 mg/kg, s.c.) induced by [betaALA(8)]-neurokinin A (NKA) (4-10) in guinea pigs, attesting to NK(1) over NK(2) selectivity. R116301 did not affect senktide-induced miosis (>5 mg/kg, s.c.) in rabbits, confirming the absence of an interaction with the NK(3) receptor. R116301 was inactive in guinea pigs against skin reactions induced by histamine, platelet-aggregating factor, bradykinin, or Ascaris allergens (>10 mg/kg, s.c.). In all species, R116301 showed excellent oral over parenteral activity (ratio, 0.22-2.7) and a relatively long duration (6.5-16 h, p.o.). The data attest to the specificity and sensitivity of the animal models and support a role of NK(1) receptors in various diseases.     

Correlating efficacy in rodent cognition models with in vivo 5-hydroxytryptamine1a receptor occupancy by a novel antagonist, (R)-N-(2-methyl-(4-indolyl-1-piperazinyl)ethyl)-N-(2-pyridinyl)-cyclohexane carboxamide (WAY-101405)

5-Hydroxytryptamine (5-HT)(1A) receptors play an important role in multiple cognitive processes, and compelling evidence suggests that 5-HT(1A) antagonists can reverse cognitive impairment. We have examined the therapeutic potential of a potent (K(i) = 1.1 nM), selective (>100-fold), orally bioavailable, silent 5-HT(1A) receptor antagonist (K(B) = 1.3 nM) (R)-N-(2-methyl-(4-indolyl-1-piperazinyl)-ethyl)-N-(2-pyridinyl)-cyclohexane carboxamide (WAY-101405). Oral administration of WAY-101405 was shown to be effective in multiple rodent models of learning and memory. In a novel object recognition paradigm, 1 mg/kg enhanced retention (memory) for previously learned information, and it was able to reverse the memory deficits induced by scopolamine. WAY-101405 (1 mg/kg) was also able to reverse scopolamine-induced deficits in a rat contextual fear conditioning model. In the Morris water maze, WAY-101405 (3 mg/kg) significantly improved learning in a paradigm of increasing task difficulty. In vivo microdialysis studies in the dorsal hippocampus of freely moving adult rats demonstrated that acute administration of WAY-101405 (10 mg/kg) increased extracellular acetylcholine levels. The selective radioligand [(3)H]WAY-100635, administered i.v., was used for in vivo receptor occupancy studies, where WAY-101405 occupied 5-HT(1A) receptors in the rat cortex, with an ED(50) value of 0.1 mg/kg p.o. Taken together, these studies demonstrate that WAY-101405 is a potent and selective, brain penetrant, orally bioavailable 5-HT(1A) receptor "silent" antagonist that is effective in preclinical memory paradigms at doses where approximately 90% of the postsynaptic 5-HT(1A) receptors are occupied. These results further support the rationale for use of this compound class in the treatment of cognitive dysfunction associated with psychiatric and neurological conditions.     

Antidepressant-like behavioral effects in 5-hydroxytryptamine(1A) and 5-hydroxytryptamine(1B) receptor mutant mice

The development of serotonin receptor knockout mice has provided an opportunity to study antidepressant drug effects in animals with targeted genetic deletion of receptors involved in antidepressant responses. In the current study, the effects of two types of antidepressant drugs, the selective serotonin reuptake inhibitors fluoxetine and paroxetine and the selective norepinephrine reuptake inhibitor desipramine, were examined in 5-hydroxytryptamine (5-HT)(1A) and 5-HT(1B) receptor mutant mice using the tail suspension test (TST). Under baseline conditions, the immobility of 5-HT(1A) receptor mutant mice, but not 5-HT(1B) receptor mutant mice, was significantly lower than that of wild-type mice. The decreased baseline immobility in 5-HT(1A) receptor mutant mice was reversed by pretreatment with alpha-methyl-para-tyrosine, but not by para-chlorophenylalanine, suggesting mediation by enhanced catecholamine function. In wild-type mice, fluoxetine (10.0--20.0 mg/kg i.p.) and desipramine (5.0--20.0 mg/kg i.p.) both significantly decreased immobility in the TST. In 5-HT(1A) receptor mutant mice, desipramine (20.0 mg/kg i.p.) significantly decreased immobility, whereas fluoxetine (20.0 mg/kg i.p.) and paroxetine (20.0 mg/kg i.p.) had no effect. The immobility of 5-HT(1B) receptor mutant mice was decreased similarly by desipramine (5.0--20.0 mg/kg i.p.). However, the effect of low doses of fluoxetine were significantly augmented in the 5-HT(1B) receptor mutant mice (2.5--20.0 mg/kg i.p.) compared with wild-type mice. Administration of selective 5-HT receptor antagonists in wild-type mice partially reproduced the phenotypes of the mutant mice. These results suggest that 5-HT(1A) and 5-HT(1B) receptors have different roles in the modulation of the response to antidepressant drugs in the TST.     

Characterization of vabicaserin (SCA-136), a selective 5-hydroxytryptamine 2C receptor agonist

The 5-hydroxytryptamine 2C (5-HT(2C)) receptor subtype has received considerable attention as a target for drug discovery, having been implicated in a wide variety of disorders. Here, we describe the in vitro pharmacological profile of the novel 5-HT(2C) receptor-selective agonist vabicaserin [(-)-4,5,6,7,9,9a,10,11,12,12a-decahydrocyclopenta[c] [1,4]diazepino[6,7,1-ij]quinoline hydrochloride] (SCA-136), including a comprehensive strategy to assess 5-HT(2B) receptor selectivity using diverse preparations and assays of receptor activation. Vabicaserin displaced (125)I-(2,5-dimethoxy)phenylisopropylamine binding from human 5-HT(2C) receptor sites in Chinese hamster ovary cell membranes with a K(i) value of 3 nM and was >50-fold selective over a number of serotonergic, noradrenergic, and dopaminergic receptors. Binding affinity determined for the human 5-HT(2B) receptor subtype using [(3)H]5HT was 14 nM. Vabicaserin was a potent and full agonist (EC(50), 8 nM; E(max), 100%) in stimulating 5-HT(2C) receptor-coupled calcium mobilization and exhibited 5-HT(2A) receptor antagonism and 5-HT(2B) antagonist or partial agonist activity in transfected cells, depending on the level of receptor expression. In rat stomach fundus and human colonic longitudinal muscle endogenously expressing 5-HT(2B) receptors, vabicaserin failed to induce a 5-HT(2B) receptor-dependent contraction and produced a rightward shift of the 5-HT and α-methyl-5-HT concentration-response curves in these preparations, respectively, consistent with 5-HT(2B) competitive antagonism. Likewise, vabicaserin failed to induce a 5-HT(2B) receptor-mediated contraction in arteries from deoxycorticosterone acetate-salt-treated rats, a model of hypersensitized 5-HT(2B) receptor function, and produced a rightward shift in the 5-HT-induced response that was consistent with 5-HT(2B) receptor antagonism. In summary, vabicaserin is a novel, potent, and selective 5-HT(2C) receptor agonist.     

N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-nitrophenyl) cyclohexanecarboxamide: a novel pre- and postsynaptic 5-hydroxytryptamine(1A) receptor antagonist active on the lower urinary tract.

N-[2-[4-(2-Methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-nitrophenyl) cyclohexanecarboxamide (Rec 15/3079) was synthesized with the aim of obtaining a novel compound with 5-hydroxytryptamine (5-HT)(1A) antagonistic properties and activity in controlling bladder function at the level of the central nervous system. Rec 15/3079 showed a selective high affinity for the 5-HT(1A) receptor (K(i) = 0.2 nM). At the human recombinant 5-HT(1A) receptor, Rec 15/3079 acted as a competitive, neutral antagonist in that it did not modify basal [(35)S]guanosine-5'-O-(3-thio)triphosphate binding to HeLa cell membranes but shifted the activation isotherm to 5-HT to the right, in a parallel manner, with a pK(b) value of 10.5. Accordingly, Rec 15/3079 (i.v.) potently antagonized 8-hydroxy-2-dipropylaminotetralin (8-OH-DPAT)-induced hypothermia in mice (ID(50) = 20 microg/kg) and 8-OH-DPAT-induced forepaw treading in rats (ID(50) = 36 microg/kg). In vitro Rec 15/3079 was poorly active in antagonizing carbachol-induced bladder (pD'(2) = 5.03) and norepinephrine-induced urethral (apparent pK(b) = 6) contractions. However, in anesthetized rats, Rec 15/3079 (10-100 microg/kg i.v.) blocked isovolumic bladder contractions with no effect on their amplitude. In conscious rats and guinea pigs with bladders filled with saline, Rec 15/3079 (300-1000 microg/kg i.v.) increased bladder volume capacity (BVC) without affecting bladder contractility. In conscious rats with bladders filled with dilute acetic acid, Rec 15/3079 (300 microg/kg i.v.) reversed the decrease of BVC induced by the acid. To evaluate apparent selective effect on lower urinary tract reflexes, Rec 15/3079 was tested in experimental models for sedative, analgesic, anxiolytic, and antidepressant activity. Rec 15/3079 showed only a slight decrease in the duration of immobility in the behavioral despair test (antidepressant activity) at 1 mg/kg i.v. No anxiolytic activity was observed at 10 mg/kg i.v. No effect was observed in the hot plate test, but Rec 15/3079 increased tail-flick latencies after 3 to 10 mg/kg i.v. In conclusion, these studies demonstrate that Rec 15/3079 is endowed with favorable effects on bladder function, and it is devoid of unwanted side effects at the level of central nervous system at doses at least 10-fold higher than those active on the bladder.     

Antihyperalgesic effects of (R,E)-N-(2-hydroxy-2,3-dihydro-1H-inden-4-yl)-3-(2-(piperidin-1-yl)-4-(trifluoromethyl)phenyl)-acrylamide (AMG8562), a novel transient receptor potential vanilloid type 1 modulator that does not cause hyperthermia in rats

Antagonists of the vanilloid receptor TRPV1 (transient receptor potential vanilloid type 1) have been reported to produce antihyperalgesic effects in animal models of pain. These antagonists, however, also caused concomitant hyperthermia in rodents, dogs, monkeys, and humans. Antagonist-induced hyperthermia was not observed in TRPV1 knockout mice, suggesting that the hyperthermic effect is exclusively mediated through TRPV1. Since antagonist-induced hyperthermia is considered a hurdle for developing TRPV1 antagonists as therapeutics, we investigated the possibility of eliminating hyperthermia while maintaining antihyperalgesia. Here, we report four potent and selective TRPV1 modulators with unique in vitro pharmacology profiles (profiles A through D) and their respective effects on body temperature. We found that profile C modulator, (R,E)-N-(2-hydroxy-2,3-dihydro-1H-inden-4-yl)-3-(2-(piperidin-1-yl)-4-(trifluoromethyl)phenyl)acrylamide (AMG8562), blocks capsaicin activation of TRPV1, does not affect heat activation of TRPV1, potentiates pH 5 activation of TRPV1 in vitro, and does not cause hyperthermia in vivo in rats. We further profiled AMG8562 in an on-target (agonist) challenge model, rodent pain models, and tested for its side effects. We show that AMG8562 significantly blocks capsaicin-induced flinching behavior, produces statistically significant efficacy in complete Freund's adjuvant- and skin incision-induced thermal hyperalgesia, and acetic acid-induced writhing models, with no profound effects on locomotor activity. Based on the data shown here, we conclude that it is feasible to modulate TRPV1 in a manner that does not cause hyperthermia while maintaining efficacy in rodent pain models.     

Pharmacological characterization of (3S)-3-(hydroxymethyl)-4-(5-methylpyridin-2-yl)-N-[6-(2,2,2-trifluoroethoxy)pyridin-3-yl]-3,4-dihydro-2H-benzo[b][1,4]oxazine-8-carboxamide (JTS-653), a novel transient receptor potential vanilloid 1 antagonist

Transient receptor potential vanilloid 1 (TRPV1) activation in peripheral sensory nerve is known to be associated with various pain-related diseases, thus TRPV1 has been the focus as a target for drug discovery. In this study, we characterized the pharmacological profiles of (3S)-3-(hydroxymethyl)-4-(5-methylpyridin-2-yl)-N-[6-(2,2,2-trifluoroethoxy)pyridin-3-yl]-3,4-dihydro-2H-benzo[b][1,4]oxazine-8-carboxamide (JTS-653), a novel TRPV1 antagonist. JTS-653 displaced [(3)H]resiniferatoxin binding to human and rat TRPV1. JTS-653 competitively antagonized the capsaicin-induced activation of human TRPV1 with pA(2) values of 10.1. JTS-653 also inhibited proton-induced activation of human and rat TRPV1 with IC(50) values of 0.320 and 0.347 nM, respectively. Electrophysiological studies indicated that JTS-653 blocked heat-induced inward currents in rat TRPV1 with IC(50) values of 1.4 nM. JTS-653 showed weak or no inhibitory effects on other TRP channels, receptors, and enzymes. JTS-653 significantly prevented capsaicin-induced mechanical hyperalgesia at 1 mg/kg p.o. and attenuated carrageenan-induced mechanical hyperalgesia at 0.3 mg/kg p.o. JTS-653 significantly attenuated carrageenan-induced thermal hyperalgesia at 0.1 mg/kg p.o. and fully reversed at 0.3 mg/kg p.o. without affecting the volume of the carrageenan-treated paw. JTS-653 showed a transient increase of body temperature at 0.3 mg/kg p.o. These results indicated that JTS-653 is a highly potent and selective TRPV1 antagonist in vitro and in vivo and suggested that JTS-653 is one of the most potent TRPV1 antagonists. The profiles of JTS-653, high potency in vivo and transient hyperthermia, seem to be associated with polymodal inhibition of TRPV1 activation.     

Pharmacological properties of (2R)-N-[1-(6-aminopyridin-2-ylmethyl)piperidin-4-yl]-2-[(1R)-3,3-difluorocyclopentyl]-2-hydroxy-2-phenylacetamide: a novel mucarinic antagonist with M(2)-sparing antagonistic activity

We evaluated the pharmacological profiles of (2R)-N-[1-(6- aminopyridin-2-ylmethyl)piperidin-4-yl]-2-[(1R)-3,3-difluorocyclopentyl]-2-hydroxy-2-phenylacetamide(compound A), which is a novel muscarinic receptor antagonist with M(2)-sparing antagonistic activity. Compound A inhibited [(3)H]NMS binding to cloned human muscarinic m1, m2, m3, m4, and m5 receptors expressed in Chinese hamster ovary cells with K(i) values (nM) of 1.5, 540, 2.8, 15, and 7.7, respectively. In isolated rat tissues, compound A inhibited carbachol-induced responses with 540-fold selectivity for trachea (K(B) = 1.2 nM) over atria (K(B) = 650 nM). In in vivo rat assays, compound A inhibited acetylcholine-induced bronchoconstriction and bradycardia with intravenous ED(50) values of 0.022 mg/kg and >/=10 mg/kg, respectively. Furthermore, in dogs, compound A (0.1-1 mg/kg p.o.) dose dependently shifted the methacholine concentration-respiratory resistance curves. In mice, compound A (10 mg/kg i.v.) did not inhibit oxotremorine-induced tremor. The brain/plasma ratio (K(p)) of compound A (3 mg/kg i.v.) was 0.13 in rats; this K(p) was less than that of scopolamine (1.7) and darifenacin (0.24). The inhibition of compound A (3 mg/kg i.v.) on ex vivo binding in rat cerebral cortex was almost similar to that of NMS. These findings demonstrate that compound A has high selectivity for M(3) receptors over M(2) receptors, displays a potent, oral M(3) antagonistic activity without inhibition of central muscarinic receptors because of low brain penetration. It is well known that central muscarinic antagonists may have diverse CNS effects, and M(2) receptors regulate cardiac pacing and act as autoreceptors in the lung and bladder. Thus, compound A may have fewer cardiac or CNS side effects than nonselective compounds.     

Antinociceptive pharmacology of N-(4-chlorobenzyl)-N'-(4-hydroxy-3-iodo-5-methoxybenzyl) thiourea, a high-affinity competitive antagonist of the transient receptor potential vanilloid 1 receptor

The transient receptor potential vanilloid 1 receptor (TRPV1) is expressed predominantly in a subset of primary afferent nociceptors. Due to its specific anatomical location and its pivotal role as a molecular integrator for noxious thermal and chemical stimuli, there is considerable interest to develop TRPV1 antagonists for the treatment of pain. Recently, N-(4-chlorobenzyl)-N'-(4-hydroxy-3-iodo-5-methoxybenzyl) thiourea (IBTU) was synthesized, and it was found in vitro to be a high-affinity competitive antagonist of cytoplasmic, but not intracellular, TRPV1. In this study, we examined the in vivo antinociceptive activity of IBTU in several acute and inflammatory pain models in mice. Our emphasis was on nociceptive pathways that are likely mediated by TRPV1, including capsaicin-, noxious heat-, and proton (including inflammation)-induced nociception tests. Capsazepine was used as a positive control in these experiments. IBTU dose-dependently blocked the capsaicin-induced nociception, confirming its antagonism at TRPV1 in vivo. By itself, IBTU produced significant antinociception, because it significantly prolonged the tail-flick latency in a dose-dependent manner. IBTU also blocked both early and late phases of the formalin-induced flinching response as well as acetic acid-induced writhing behavior. Moreover, IBTU inhibited the complete Freund's adjuvant-induced persistent hyperalgesia. Taken together, these data demonstrate that IBTU acts as a TRPV1 antagonist in vivo, and they suggest that it may be of therapeutic use for the treatment of pain.     

2-[4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy]-N-(methylsulfonyl)acetamide (NS-304), an orally available and long-acting prostacyclin receptor agonist prodrug

Prostacyclin (PGI(2)) and its analogs are useful for the treatment of various vascular disorders, but their half-lives are too short for widespread clinical application. To overcome this drawback, we have synthesized a novel diphenylpyrazine derivative, 2-[4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy]-N-(methylsulfonyl)acetamide (NS-304), a prodrug of the active form [4-[(5,6-diphenylpyrazin-2-yl)(isopropyl)amino]butoxy]acetic acid (MRE-269). NS-304 is an orally available and potent agonist for the PGI(2) receptor (IP receptor). The inhibition constant (K(i)) of MRE-269 for the human IP receptor was 20 nM; in contrast, the K(i) values for other prostanoid receptors were >2.6 microM. MRE-269 was therefore a highly selective agonist for the IP receptor. The plasma concentrations of MRE-269 remained near peak levels for more than 8 h after oral administration of NS-304 to rats and dogs, and NS-304 increased femoral skin blood flow in rats in a long-lasting manner without affecting the hemodynamics. These findings indicate that NS-304 acts as a long-acting IP receptor agonist in vivo. The continuous vasodilation evoked by NS-304 was not attenuated by repeated treatment, indicating that NS-304 is unlikely to cause severe desensitization of the IP receptor in rats. Moreover, a microdose pharmacokinetic study in which NS-304 was orally administered to healthy male volunteers showed conversion of NS-304 to MRE-269 and a long plasma elimination half-life for MRE-269 (7.9 h). In conclusion, NS-304 is an orally available and long-acting IP receptor agonist prodrug, and its active form, MRE-269, is highly selective for the IP receptor. Therefore, NS-304 is a promising drug candidate for various vascular diseases, especially pulmonary arterial hypertension and arteriosclerosis obliterans.     

Pharmacological characterization of the novel histamine H3-receptor antagonist N-(3,5-dichlorophenyl)-N'-[[4-(1H-imidazol-4-ylmethyl)phenyl]-methyl]-urea (SCH 79687)

We present the pharmacological and pharmacokinetic profiles of a novel histamine H3 receptor antagonist, N-(3,5-dichlorophenyl)-N'-[[4-(1H-imidazol-4-ylmethyl)phenyl]-methyl]-urea (SCH 79687). The H3-receptor binding Ki values for SCH 79687 were 1.9 and 13 nM in the rat and guinea pig (GP), respectively. The Ki values for SCH 79687 at histamine H1 and H2 receptors were greater than 1 microM. SCH 79687 showed a 41- and 82-fold binding selectivity for the H3 receptor over alpha 2A-adrenoceptors and imidazoline I2, and >500-fold H3 selectivity compared with over 60 additional receptors. The pA2 value for SCH 79687 in the GP ileum electrical field-stimulated (EFS) contraction was 9.6 +/- 0.3. Similar H3 antagonist activity was observed in the EFS cryopreserved and fresh tissue isolated human saphenous vein (HSV) assays (pKb = 9.4 +/- 0.3 and 10.1 +/- 0.4). SCH 79687 (30 nM) did not block clonidine-induced inhibition of EFS-induced contractions in HSV. SCH 79687 (ED50 = 0.3 mg/kg i.v.) attenuated (R)-alpha-methylhistamine inhibition of sympathetic hypertensive responses in the GP. At the time of activity evaluation, the GP plasma SCH 79687 concentration was 25 ng/ml at the dose of 0.3 mg/kg i.v. In feline nasal studies, combined administration of SCH 79687 (3 mg/kg i.v.) and the H1-antagonist loratadine (3 mg/kg i.v.), at individual doses that do not produce decongestion, inhibited the compound 48/80-induced congestion by 47%. The alpha-adrenergic agonist phenylpropanolamine (PPA; 1 mg/kg i.v.) also attenuated compound 48/80 nasal responses by 42%. Unlike the H3/H1 combination that did not affect blood pressure (BP), PPA (1 mg/kg i.v.) significantly increased BP compared with control animals by a maximum of 31 mm Hg. Orally, SCH 79687 (10 mg/kg) plus loratadine (10 mg/kg) also produced decongestion without effects on BP. In pharmacokinetic studies, oral dosing with SCH 79687 in the rat (10 mg/kg) and monkey (3 mg/kg) achieved plasma Cmax and area under the curve values greater than 1.5 and 12.1 microg. h/ml, respectively. SCH 79687 is an orally active H3 antagonist with a good pharmacokinetic profile that, in combination with an H1 antagonist, demonstrates decongestant efficacy comparable with oral sympathomimetic decongestants but without hypertensive liabilities.     

The serotonin1A receptor partial agonist S15535 [4-(benzodioxan-5-yl)1-(indan-2-yl)piperazine] enhances cholinergic transmission and cognitive function in rodents: a combined neurochemical and behavioral analysis

These studies examined the influence of the selective 5-hydroxytryptamine (serotonin) (5-HT)(1A) receptor partial agonist S15535 [4-(benzodioxan-5-yl)1-(indan-2-yl)piperazine] upon cholinergic transmission and cognitive function in rodents. In the absence of acetylcholinesterase inhibitors, S15535 dose-dependently (0.04-5.0 mg/kg s.c.) elevated dialysis levels of acetylcholine in the frontal cortex and dorsal hippocampus of freely moving rats. In the cortex, the selective 5-HT(1A) receptor antagonist WAY100,635 [(N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclo-hexanecarboxamide) fumarate] dose-dependently (0.0025-0.63) blocked this action of S15535. By contrast, in dorsal hippocampus, WAY100,635 mimicked the induction of acetylcholine release by S15535. In a social recognition paradigm, S15535 dose-dependently (0.16-10.0) improved retention, an action blocked by WAY100,635 (0.16), which was ineffective alone. Furthermore, S15535 dose-dependently (0.04-2.5) and WAY100,635 reversibly abolished amnesic properties of the muscarinic antagonist scopolamine (0.63) in this procedure. Cognitive deficits provoked by scopolamine in autoshaping and Morris water-maze procedures were likewise blocked by S15535 at doses of 0.63 to 10.0 and 0.16 to 2.5, respectively. In a two-platform spatial discrimination task, in which S15535 similarly abrogates cognitive deficits elicited by scopolamine, injection of S15535 (1.0 and 10.0 microg) into dorsal hippocampus blocked amnesic effects of the 5-HT(1A) agonist 8-hydroxy-2-dipropylaminotetralin (0.5 microg). Finally, S15535 (0.16-0.63) improved performance in a spatial, delayed nonmatching to sample model in mice, and in an operant delayed nonmatching to sample model in old rats, S15535 (1.25-5.0 mg/kg p.o.) increased response accuracy and reduced latency to respond. In conclusion, S15535 reinforces frontocortical and hippocampal release of acetylcholine and displays a broad-based pattern of procognitive properties. Its actions involve both blockade of postsynaptic 5-HT(1A) receptors and engagement of 5-HT(1A) autoreceptors.     

Characterization of N-(1-Acetyl-2,3-dihydro-1H-indol-6-yl)-3-(3-cyano-phenyl)-N-[1-(2-cyclopentyl-ethyl)-piperidin-4yl]acrylamide (JNJ-5207787), a small molecule antagonist of the neuropeptide Y Y2 receptor

The in vitro pharmacological properties of N-(1-Acetyl-2,3-dihydro-1H-indol-6-yl)-3-(3-cyano-phenyl)-N-[1-(2-cyclopentyl-ethyl)-piperidin-4yl]-acrylamide (JNJ-5207787), a novel neuropeptide Y Y(2) receptor (Y(2)) antagonist, were evaluated. JNJ-5207787 inhibited the binding of peptide YY (PYY) to human Y(2) receptor in KAN-Ts cells (pIC(50) = 7.00 +/- 0.10) and to rat Y(2) receptors in rat hippocampus (pIC(50) = 7.10 +/- 0.20). The compound was >100-fold selective versus human Y(1),Y(4), and Y(5) receptors as evaluated by radioligand binding. In vitro receptor autoradiography data in rat brain tissue sections confirmed the selectivity of JNJ-5207787. [(125)I]PYY binding sites sensitive to JNJ-5207787 were found in rat brain regions known to express Y(2) receptor (septum, hypothalamus, hippocampus, substantia nigra, and cerebellum), whereas insensitive binding sites were observed in regions known to express Y(1) receptor (cortex and thalamus). JNJ-5207787 was demonstrated to be an antagonist via inhibition of PYY-stimulated guanosine 5'-O-(3-[(35)S]thio)triphosphate binding ([(35)S]GTPgammaS) in KAN-Ts cells (pIC(50) corrected = 7.20 +/- 0.12). This was confirmed auto-radiographically in rat brain sections where PYY-stimulated guanosine 5'-O-(3-[(35)S]thio)triphosphate binding was inhibited by JNJ-5207787 (10 microM) in hypothalamus, hippocampus, and substantia nigra. After intraperitoneal administration in rats (30 mg/kg), JNJ-5207787 penetrated into the brain (C(max) = 1351 +/- 153 ng/ml at 30 min) and occupied Y(2) receptor binding sites as revealed by ex vivo receptor autoradiography. Hence, JNJ-5207787 is a potent and selective pharmacological tool available to establish the potential role of central and peripheral Y(2) receptors in vivo.     

A novel chymase inhibitor, 2-(5-formylamino-6-oxo-2-phenyl-1,6-dihydropyrimidine-1-yl)-N-[[,4-dioxo-1-phenyl-7-(2-pyridyloxy)]2-heptyl]acetamide (NK3201), suppressed intimal hyperplasia after balloon injury

In this study, we investigated whether an orally active chymase inhibitor, 2-(5-formylamino-6-oxo-2-phenyl-1,6-dihydropyrimidine-1-yl)-N-[[3,4-dioxo-1-phenyl-7-(2-pyridyloxy)]-2-heptyl]acetamide (NK3201), prevents intimal hyperplasia in carotid arteries injured by a balloon catheter in dog. Each dog was administered NK3201 (1 mg/kg per day, p.o.) or placebo beginning 5 days before balloon injury and continuing through the experiments. Four weeks after balloon injury, NK3201 did not affect the plasma renin and angiotensin-converting enzyme activities. The chymase activity was significantly increased in the injured arteries, whereas the angiotensin-converting enzyme activity was not. NK3201 significantly reduced the chymase activity in the injured arteries. The intimal area in the placebo- and NK3201-treated group and was 0.46 +/- 0.06 and 0.24 +/- 0.04 mm2, respectively, and this difference was significant. In this study, we demonstrated for the first time that a chymase inhibitor prevented the development of intimal hyperplasia in the balloon-injured arteries.     

Pharmacology and antitussive efficacy of 4-(3-trifluoromethyl-pyridin-2-yl)-piperazine-1-carboxylic acid (5-trifluoromethyl-pyridin-2-yl)-amide (JNJ17203212), a transient receptor potential vanilloid 1 antagonist in guinea pigs

Transient receptor potential vanilloid 1 (TRPV1) plays an integral role in modulating the cough reflex, and it is an attractive antitussive drug target. The purpose of this study was to characterize a TRPV1 antagonist, 4-(3-trifluoromethyl-pyridin-2-yl)-piperazine-1-carboxylic acid (5-trifluoromethyl-pyridin-2-yl)-amide (JNJ17203212), against the guinea pig TRPV1 receptor in vitro followed by a proof-of-principle study in an acid-induced model of cough. The affinity of JNJ17203212 for the recombinant guinea pig TRPV1 receptor was estimated by radioligand binding, and it was functionally characterized by antagonism of low-pH and capsaicin-induced activation of the ion channel (fluorometric imaging plate reader and electrophysiology). The nature of antagonism was further tested against the native channel in isolated guinea pig tracheal rings. Following pharmacokinetic characterization of JNJ17203212 in guinea pigs, pharmacodynamic and efficacy studies were undertaken to establish the antitussive efficacy of the TRPV1 antagonist. The pK(i) of JNJ17203212 for recombinant guinea pig TRPV1 was 7.14 +/- 0.06. JNJ17203212 inhibited both pH (pIC(50) of 7.23 +/- 0.05) and capsaicin (pIC(50) of 6.32 +/- 0.06)-induced channel activation. In whole-cell patch clamp, the pIC(50) for inhibition of guinea pig TRPV1 was 7.3 +/- 0.01. JNJ17203212 demonstrated surmountable antagonism in isolated trachea, with a pK(B) value of 6.2 +/- 0.1. Intraperitoneal administration of 20 mg/kg JNJ17203212 achieved a maximal plasma exposure of 8.0 +/- 0.4 microM, and it attenuated capsaicin evoked coughs with similar efficacy to codeine (25 mg/kg). Last, JNJ17203212 dose-dependently produced antitussive efficacy in citric acid-induced experimental cough in guinea pigs. Our data provide preclinical support for developing TRPV1 antagonists for the treatment of cough.     

Inverse agonist actions of typical and atypical antipsychotic drugs at the human 5-hydroxytryptamine(2C) receptor

Atypical antipsychotic drugs, which are distinguished from typical antipsychotic drugs by a lower incidence of extra-pyramidal side effects and less propensity to elevate serum prolactin levels (e.g., clozapine, olanzapine, risperidone, quetiapine, ziprasidone), have become the most widely used treatments for schizophrenia, although their precise mechanism of action remains controversial. It has been suggested that this group of atypical antipsychotic drugs is characterized by preferentially high affinities for 5-hydroxytryptamine (5-HT)2A serotonin receptors and relatively low affinities for D2-dopamine receptors. It has recently been proposed that these atypical antipsychotic drugs may also be distinguished from typical antipsychotic drugs (e.g., haloperidol, fluphenazine, chlorpromazine, and so on) by inverse agonist actions at the 5-HT2C-INI RNA edited isoform of the human 5-HT2C receptor transiently expressed in COS-7 cells. We have examined the relationship among 5-HT2C inverse agonist potency, efficacy, and atypical antipsychotic drug status in HEK-293 cells of a large number of typical and atypical antipsychotic drugs using human embryonic kidney (HEK)-293 cells stably transfected with the h5-HT2C-INI receptor. Inverse agonist actions at h5-HT2C-INI receptors were measured for both typical and atypical antipsychotic drugs. Thus, some typical antipsychotic drugs (chlorpromazine, mesoridazine, fluphenazine, and loxapine) were efficient inverse agonists, whereas several clinically effective atypical antipsychotic drugs (remoxapride, quetiapine, sulpiride, melperone, amperozide) were not. Additionally, several drugs without significant antipsychotic actions (M100907, ketanserin, mianserin, ritanserin, and amitriptyline) were potent inverse agonists at the 5-HT2C-INI isoform expressed in HEK-293 cells. Taken together, these results demonstrate that both typical and atypical antipsychotic drugs may exhibit inverse agonist effects at the 5-HT2C-INI isoform of the human 5-HT2C receptor and that no relationship exists between inverse agonist actions and atypicality.     

Characterization of nociceptin/orphanin FQ-induced pain responses by the novel receptor antagonist N-(4-amino-2-methylquinolin-6-yl)-2-(4-ethylphenoxymethyl) benzamide monohydrochloride

At the spinal level, nociceptin/orphanin FQ (Noc/OFQ) produces pronociceptive and allodynic effects at low doses (picogram range), while causing antinociceptive effects at high doses (microgram range). The discrepancy of pain modulation by Noc/OFQ at low and high doses raised a question whether Noc/OFQ exerted actions through the same Noc/OFQ receptor. In the present study, we examined the involvement of the Noc/OFQ receptor in pain responses with the novel nonpeptide antagonist N-(4-amino-2-methylquinolin-6-yl)-2-(4-ethylphenoxymethyl) benzamide monohydrochloride (JTC-801). Allodynia and hyperalgesia evoked by intrathecal administration of Noc/OFQ (50 pg/mouse) were dose dependently blocked by simultaneous administration of JTC-801 with IC(50) values of 32.2 and 363 pg, respectively. JTC-801 did not induce allodynia by itself. Subcutaneous injection of formalin into a hindpaw evoked biphasic pain behaviors such as flinching and biting in mice. Noc/OFQ at 10 pg increased the second-phase pain behaviors evoked by 1% formalin, whereas it strongly inhibited both the first-phase and second-phase pain evoked by 2% formalin at 1 microg. Although the pronociceptive effect by 10 pg of Noc/OFQ was dose dependently blocked by JTC-801 with an IC(50) value of 4.58 pg, the antinociceptive effects by 1 microg of Noc/OFQ were not antagonized by JTC-801. Furthermore, both phases of 2% formalin-induced pain behaviors were relieved by JTC-801. These results demonstrate that pronociceptive responses induced by a low dose of Noc/OFQ may be mediated through the Noc/OFQ receptor in the spinal cord and that JTC-801 can be a useful antagonist to examine the involvement of endogenous Noc/OFQ and mediation of the Noc/OFQ receptor under physiological and pathophysiological conditions including pain.     

Pharmacological and pharmacokinetic properties of a structurally novel, potent, and selective metabotropic glutamate 2/3 receptor agonist: in vitro characterization of agonist (-)-(1R,4S,5S,6S)-4-amino-2-sulfonylbicyclo[3.1.0]-hexane-4,6-dicarboxylic acid (LY404039)

Group II metabotropic glutamate (mGlu) receptor agonists, including (1S,2S,5R,6S)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylate monohydrate (LY354740) and (-)-2-oxa-4-aminobicyclo[3.1.0]hexane-4,6-dicarboxylate (LY379268), have demonstrated efficacy in animal models of anxiety and schizophrenia, and LY354740 decreased anxiety in human subjects. Herein, we report the in vitro pharmacological profile and pharmacokinetic properties of another potent, selective, and structurally novel mGlu2/3 receptor agonist, (-)-(1R,4S,5S,6S)-4-amino-2-sulfonylbicyclo[3.1.0]hexane-4,6-dicarboxylic acid (LY404039) and provide comparisons with LY354740. Similar to LY354740, LY404039 is a nanomolar potent agonist at recombinant human mGlu2 and mGlu3 receptors (K(i) = 149 and 92, respectively) and in rat neurons expressing native mGlu2/3 receptors (Ki = 88). LY404039 is highly selective for mGlu2/3 receptors, showing more than 100-fold selectivity for these receptors, versus ionotropic glutamate receptors, glutamate transporters, and other receptors targeted by known anxiolytic and antipsychotic medications. Functionally, LY404039 potently inhibited forskolin-stimulated cAMP formation in cells expressing human mGlu2 and mGlu3 receptors. Electrophysiological studies indicated that LY404039 suppressed electrically evoked excitatory activity in the striatum, and serotonin-induced l-glutamate release in the prefrontal cortex; effects reversed by LY341495. These characteristics suggest LY404039 modulates glutamatergic activity in limbic and forebrain areas relevant to psychiatric disorders; and that, similar to LY354740, it works through a mechanism that may be devoid of negative side effects associated with current antipsychotics and anxiolytics. Interestingly, despite the slightly lower potency (approximately 2-5-fold) of LY404039 versus LY354740 in binding, functional, and electrophysiological assays, LY404039 demonstrated higher plasma exposure and better oral bioavailability in pharmacokinetic experiments. Collectively, the current data indicate that LY404039 may be valuable in the treatment of neuropsychiatric disorders, including anxiety and psychosis.