Synthesis and biological activities of 4-phenyl-5-pyridyl-1,3-thiazole derivatives as selective adenosine A3 antagonists.

To investigate the potency of an adenosine A3 receptor (A3AR) antagonist as an anti-asthmatic drug, a novel series of 4-phenyl-5-pyridyl-1,3-thiazole derivatives was synthesized and evaluated in human adenosine A1, A2A and A3 receptor and rat adenosine A3 receptor binding assays. From investigation of the SAR study, compound 7af was identified as a highly potent human and rat A3AR antagonist. This compound inhibited IB-MECA-induced plasma protein extravasation in the skin of rats and showed good oral absorption. Also, compound 7af significantly inhibited antigen-induced hyper-responsiveness to acetylcholine in actively sensitized Brown Norway rats. These results show that 4-phenyl-5-pyridyl-1,3-thiazole derivatives are potential candidates to enable the evaluation of A3AR antagonists. Further evaluation of this class of compounds may afford a novel anti-inflammatory agent such as an anti-asthmatic drug.

Novel inhibitor of p38 MAP kinase as an anti-TNF-alpha drug: discovery of N-[4-[2-ethyl-4-(3-methylphenyl)-1,3-thiazol-5-yl]-2-pyridyl]benzamide (TAK-715) as a potent and orally active anti-rheumatoid arthritis agent.

The p38 mitogen-activated protein (MAP) kinase has been implicated in the proinflammatory cytokine signal pathway, and its inhibitors are potentially useful for the treatment of chronic inflammatory diseases such as rheumatoid arthritis (RA) and inflammatory bowel disease. To develop a new drug for RA, we synthesized a novel series of 4-phenyl-5-pyridyl-1,3-thiazoles and evaluated their inhibition of p38 MAP kinase, lipopolysaccharide (LPS)-stimulated release of tumor necrosis factor-alpha (TNF-alpha) from human monocytic THP-1 cells in vitro, and LPS-induced TNF-alpha production in vivo in mice. During the course of the study, we found that these compounds risk the inhibition of cytochrome P450 (CYP) isoforms by coordination of the 4-pyridyl nitrogen with heme iron. We therefore investigated the effects of substitution at the 2-position of the pyridyl ring on the inhibitory activity of p38 MAP kinase and CYPs in more detail. As a result, N-[4-[2-ethyl-4-(3-methylphenyl)-1,3-thiazol-5-yl]-2-pyridyl]benzamide (8h, TAK-715) exhibited potent inhibitory activity in these assays (inhibition of p38alpha, IC50 = 7.1 nM; LPS-stimulated release of TNF-alpha from THP-1, IC50 = 48 nM; LPS-induced TNF-alpha production in mice, 87.6% inhibition at 10 mg/kg, po) and no inhibitory activity for major CYPs, including CYP3A4. This compound also showed good bioavailability in mice and rats and significant efficacy in a rat adjuvant-induced arthritis model. Compound 8h was selected as a clinical candidate and is now under clinical investigation for the treatment of RA.

Synthesis and biological activities of 4-phenyl-5-pyridyl-1,3-thiazole derivatives as p38 MAP kinase inhibitors.

A novel series of 4-phenyl-5-pyridyl-1,3-thiazole analogues possessing potent in vitro inhibitory activity against p38 mitogen-activated protein kinase and the release of tumor necrosis factor-alpha (TNF-alpha) from human monocytic THP-1 cells stimulated by lipopolysaccharide has been identified. Subsequent structure-activity relationship (SAR) studies and optimization for absorption, distribution, metabolism, and elimination (ADME) profiles led to the identification of compounds 7 g and 10b as orally active lead candidates that block the in vivo production of proinflammatory cytokine (TNF-alpha). In pharmacokinetic studies, compound 10b showed good oral administration in mice and demonstrated significant in vivo anti-inflammatory activity in an anti-collagen monoclonal antibody-induced arthritis mouse model (minimum effective dose (MED)=30 mg/kg). Further elucidation of this class of compounds may provide novel anti-inflammatory agents, such as anti-rheumatoid arthritis drugs.

Neurochemical properties of ramelteon (TAK-375), a selective MT1/MT2 receptor agonist.

Ramelteon (TAK-375) is a novel melatonin receptor agonist currently under investigation for the treatment of insomnia. This study describes the neurochemical and receptor binding characteristics of ramelteon in vitro. Ramelteon showed very high affinity for human MT1 (Mel1a) and MT2 (Mel1b) receptors (expressed in Chinese hamster ovary [CHO] cells), and chick forebrain melatonin receptors (consisting of Mel1a and Mel1c receptors) with Ki values of 14.0, 112, and 23.1 pM, respectively, making the affinities of ramelteon for these receptors 3-16 times higher than those of melatonin. The affinity of ramelteon for hamster brain MT3 binding sites was extremely weak (Ki: 2.65 microM) compared to melatonin's affinity for the MT3 binding site (Ki: 24.1 nM). In addition, ramelteon showed no measurable affinity for a large number of ligand binding sites (including benzodiazepine receptors, dopamine receptors, opiate receptors, ion channels, and transporters) and no effect on the activity of various enzymes. Ramelteon inhibited forskolin-stimulated cAMP production in the CHO cells that express the human MT1 or MT2 receptors. Taken together, these results indicate that ramelteon is a potent and highly selective agonist of MT1/MT2 melatonin receptors.

Ramelteon (TAK-375) accelerates reentrainment of circadian rhythm after a phase advance of the light-dark cycle in rats.

In vivo pharmacological effects of ramelteon (TAK-375), a novel, highly MT1/MT2-selective receptor agonist, were studied in rats to determine ramelteon's ability to reentrain the circadian rhythm after an abrupt phase advance. Experiments were also conducted to assess the potential cognitive side effects of ramelteon and its potential to become a drug of abuse. After an abrupt 8-h phase shift, ramelteon (0.1 and 1 mg/kg, p.o.) and melatonin (10 mg/kg, p.o.) accelerated reentrainment of running wheel activity rhythm to the new lightdark cycle. Ramelteon (3-30 mg/kg, p.o.) and melatonin (10-100 mg/kg, p.o.) did not affect learning or memory in rats tested by the water maze task and the delayed match to position task, although diazepam and triazolam impaired both of the tasks. Neither ramelteon (3-30 mg/kg, p.o.) nor melatonin (10-100 mg/kg, p.o.) demonstrated a rewarding property in the conditioned place-preference test, implying that MT1/MT2 receptor agonists have no abuse potential. In contrast, benzodiazepines and morphine showed rewarding properties in this test. The authors' results suggest that ramelteon may be useful for treatment of circadian rhythm sleep disorders without adverse effects typically associated with benzodiazepine use, such as learning and memory impairment, and drug dependence.

The sleep-promoting action of ramelteon (TAK-375) in freely moving cats.

INTRODUCTION: Ramelteon (TAK-375) is an MT1/MT2 receptor agonist being studied for the treatment of insomnia and circadian rhythm sleep disorders. We compared the behavioral effects of ramelteon and exogenous melatonin in freely moving cats. METHODS: Ramelteon and melatonin were each suspended in a 0.5% (weight per volume) methylcellulose solution and administered orally to freely moving cats. In the control trial, each cat was given vehicle. Each dose of ramelteon or melatonin was compared with the vehicle control in a crossover design. Electroencephalogram, electromyogram, and electrooculogram recordings were assessed. RESULTS: Ramelteon significantly decreased wakefulness at doses of 0.001,0.01, and 0.1 mg/kg, increased slow-wave sleep at doses of 0.001, 0.01, and 0.1 mg/kg, and increased rapid eye movement sleep at a dose of 0.1 mg/kg, compared with the vehicle controls, as assessed by analysis of variance. The effects of ramelteon lasted for up to 6 hours when evaluated by reduction of wakefulness. Exogenous melatonin (0.01-1 mg/kg) significantly increased slow-wave sleep, but the effect was weaker than that of ramelteon and lasted for only 2 hours. The lowest doses of ramelteon (0.0001 mg/kg) and melatonin (0.001 mg/kg) had no significant effect on sleep-wakefulness stage. CONCLUSIONS: Ramelteon was more effective than exogenous melatonin in promoting and maintaining sleep in freely moving cats. Based on its unique mechanism of action, ramelteon should be studied further to evaluate its potential for the treatment of sleep disorders.

Effects of ramelteon (TAK-375) on nocturnal sleep in freely moving monkeys.

We investigated the effects of (S)-N-[2-(1,6,7,8-tetrahydro-2H-indeno-[5,4]furan-8-yl)ethyl]propionamide (ramelteon, TAK-375), a novel MT1/MT2 receptor agonist, on nocturnal sleep in freely moving monkeys and compared these results with those of melatonin and zolpidem. Treatment with ramelteon (0.03 and 0.3 mg/kg, p.o.) significantly shortened latency to sleep onset and significantly increased total duration of sleep. Treatment with melatonin (0.3, 1, and 3 mg/kg, p.o.) also decreased sleep latency, but the effect was weak; the only significant reduction was seen with the 0.3 mg/kg dose on latency to light sleep. Melatonin had no significant effects on the duration of sleep. Zolpidem had no significant effects on latency to sleep onset in this study at any dose (1, 3, 10, and 30 mg/kg, p.o.). The highest dose (30 mg/kg) of zolpidem had a tendency to increase slow wave sleep; however, it also induced apparent sedation and myorelaxation. Treatment with ramelteon and melatonin had no evident effect on the general behavior of the monkeys. Spectral analysis (fast Fourier transform, FFT) of both ramelteon and melatonin revealed sleep patterns that were indistinguishable from those of naturally occurring sleep. The EEG power spectra of zolpidem were qualitatively different from that of naturally occurring physiological sleep. Results of the present study support the investigation of ramelteon as a sleep-promoting agent in humans.

Synthesis of a novel series of tricyclic indan derivatives as melatonin receptor agonists.

To develop a new therapeutic agent for sleep disorders, we synthesized a novel series of tricyclic indan derivatives and evaluated them for their binding affinity to melatonin receptors. In our previous paper, we proposed a conformation of the methoxy group favorable for the binding of the MT(1) receptor. To fix the methoxy group in an active conformation, we decided to synthesize conformationally restricted tricyclic indan analogues with the oxygen atom in the 6-position incorporated into a furan, 1,3-dioxane, oxazole, pyran, morpholine, or 1,4-dioxane ring system. Among these compounds, indeno[5,4-b]furan analogues were found to be the most potent and selective MT(1) receptor ligands and to have superior metabolic stability. The optimization of substituents led to (S)-(-)-22b, which showed very strong affinity for human MT(1) (K(i) = 0.014 nM), but no significant affinity for hamster MT(3)() (K(i) = 2600 nM) or other neurotransmitter receptors. The pharmacological effects of (S)-(-)-22b were studied in experimental animals, and it was found that a dose of 0.1 mg/kg, po promoted a sleep in freely moving cats, as demonstrated by a decrease in wakefulness and increases in slow wave sleep and rapid eye movement sleep, which lasted for 6 h after administration. Melatonin (1 mg/kg, po) also had a sleep-promoting effect, though it lasted only 2 h. A new chiral method for the synthesis of (S)-(-)-22b starting from 60, which was prepared from 59 employing asymmetric hydrogenation with the (S)-2,2'-bis(diphenylphosphino)-1,1'-binaphthyl-Ru complex, was developed. (S)-(-)-22b (TAK-375) is currently under clinical trial for the treatment of insomnia and circadian rhythm disorders.

Synthesis of a novel series of benzocycloalkene derivatives as melatonin receptor agonists.

We synthesized a novel series of benzocycloalkene derivatives and evaluated their binding affinities to melatonin receptors. To control the spatial position of the amide group, one of the important pharmacophores, we incorporated an endo double bond, an exo double bond (E- and Z-configurations), and a chiral center (R- and S-configurations) at position 1. The indan derivatives with the S-configuration at position 1 were the most promising in terms of potency and selectivity for the human melatonin receptor (MT(1) site), while compounds with the R-configuration showed little potential. Our next attempt was to investigate the most favorable conformation of the methoxy group, the other important pharmacophore for binding to the MT(1) receptor. The introduction of a methyl group at position 5 of the indene ring conserved affinity; however, at position 7, it caused a decrease in affinity. These results suggested that the substitution at position 7 forced the methoxy group to adopt an unfavorable orientation. The optimization of the condensed ring size and substituents led to (S)-8d [(S)-N-[2-(2,3-dihydro-6-methoxy-1H-inden-1-yl)ethyl]propionamide], which had high affinity for the human MT(1) receptor (K(i) = 0.041 nM) but no significant affinity for the hamster MT(3)receptor (K(i) = 3570 nM). In addition, a practical synthetic method of chiral N-[2-(2,3-dihydro-1H-inden-1-yl)ethyl]alkanamides employing asymmetric hydrogenation with (S)-2,2'-bis(diphenylphosphino)-1,1'-binaphthyl-Ru has been established.

Kinetic resolutions of indan derivatives using bacteria.

Racemic indan derivatives have been resolved by the hydrolysis of amide bonds using Corynebacterium ammoniagenes IFO12612 to produce (S)-amine and (R)-amides. In the kinetic resolution of 1 (N-12-(6-methoxy-indan-1-yl)ethyl]acetamide), it was possible to run the reaction to 44% conversion on a 10-g scale, obtaining (S)-amine 4 ((S)-2-(6-methoxy-indan-1-yl)ethylamine) at >99% enantiomeric excess (ee) and (R)-1 at 98% ee.

Kinetic resolution of an indan derivative using Bacillus sp. SUI-12: synthesis of a key intermediate of the melatonin receptor agonist TAK-375.

The chiral indan derivative (S)-2 (2-[(8S)-1,6,7,8-tetrahydro-2H-indeno[5,4-b]furan-8-yl]ethyl-amine) was synthesized by enzyme-catalyzed asymmetric hydrolysis of the racemic acetamide 1 (N-[2-(1,6,7,8-tetrahydro-2H-indeno[5,4-b]furan-8-yl)ethyl]acetamide). The reaction was carried out using Bacillus sp. SUI-12 screened for the ability to hydrolyze 1 to give (S)-2 with high enantioselectivity. In a scaled-up experiment, a low reaction rate was observed. However, by changing the culture medium and the reaction conditions, it became possible to run the reaction to 40% conversion on a 10-g or more scale, obtaining (S)-2 at >;99% enantiomeric excess (ee). The (S)-2 obtained was available for the synthesis of the melatonin receptor agonist TAK-375 (N-[2-[(8S)-1,6,7,8-tetrahydro-2H-indeno[5,4-b]furan-8-yl]ethyl]propanamide).