Discovery and optimization of piperidyl benzamide derivatives as a novel class of 11β-HSD1 inhibitors

Discovery and optimization of a piperidyl benzamide series of 11β-HSD1 inhibitors is described. This series was derived from a cyclohexyl benzamide lead structures to address PXR selectivity, high non-specific protein binding, poor solubility, limited in vivo exposure, and in vitro cytotoxicity issues observed with the cyclohexyl benzamide structures. These efforts led to the discovery of piperidyl benzamide 15 which features improved properties over the cyclohexyl benzamide derivatives.     

Design and synthesis of novel imidazoline derivatives with potent antihyperglycemic activity in a rat model of type 2 diabetes

Imidazoline derivatives have been reported to show antihyperglycemic activity in vivo. In the present study, we first showed that there was no correlation between the in vivo antidiabetic activity and the in vitro affinities for the I1/I2 binding sites for several substituted aryl imidazolines. Among these compounds, 2-(alpha-cyclohexyl-benzyl)-4,5-dihydro-1H-imidazole 2 exhibited potent antihyperglycemic properties. It was then chosen as lead compound. Thirty-six new derivatives were synthesized by replacing the cyclohexyl/benzyl group by various cyclic systems or the imidazoline ring by isosteric heterocycles. These compounds were evaluated in vivo for their antihyperglycemic activity using an oral glucose tolerance test (OGTT) in a rat model of type-2 diabetes obtained by giving a single intravenous (iv) injection of a low dose of streptozotocin to rats (STZ rats) and in normal rats. Nine compounds with an imidazoline moiety, possibly substituted by a methyl group, had a potent effect on the glucose tolerance in normal or STZ-diabetic rats, after an oral (po) administration of the test compound at a dose of 30 or 10 mg kg(-1), without any hypoglycemia. Replacement of the imidazoline ring by isosteric heterocycles resulted in a total loss of activity.     

Discovery of a highly potent orally bioavailable imidazo-[1, 2-a]pyrazine Aurora inhibitor

Imidazo-[1, 2-a]pyrazine 1 is a potent inhibitor of Aurora A and B kinase in vitro and is effective in in vivo tumor models, but has poor oral bioavailbility and is unsuitable for oral dosing. We describe herein our effort to improve oral exposure in this class, resulting ultimately in the identification of a potent Aurora inhibitor 16, which exhibited good drug exposure levels across species upon oral dosing, and showed excellent in vivo efficacy in a mouse xenograft tumor model when dosed orally.     

Compelling P1 substituent affect on metalloprotease binding profile enables the design of a novel cyclohexyl core scaffold with excellent MMP selectivity and HER-2 sheddase inhibition

A serendipitous discovery that the metalloprotease binding profile of a novel class of 2-carboxamide-3-hydroxamic acid piperidines could be significantly attenuated by the modification of the unexplored P1 substituent enabled the design and synthesis of a novel 2-carboxamide-1-hydroxamic acid cyclohexyl scaffold core that exhibited excellent HER-2 potency and unprecedented MMP-selectivity that we believe would not have been possible via conventional P1′ perturbations.     

Design and synthesis of novel tetrahydronaphthyl azoles and related cyclohexyl azoles as antileishmanial agents

A novel series of trans-2-aryloxy-1,2,3,4,-tetrahydronaphthyl azoles and related cyclohexyl azoles were synthesized and evaluated in vitro against Leishmania donovani. Compound 9 identified as most active analog with IC₅₀ value of 0.64 μg/mL and SI value of 34.78 against amastigotes, and is several folds more potent than the reference drugs sodium stilbogluconate and paromomycin. It also exhibited significant in vivo inhibition of 83.33%, and provided a new structural scaffold for antileishmanials.     

Identification of Saccharomyces cerevisiae Tub1 alpha-tubulin as a potential target for NKH-7, a cytotoxic 1-naphthol derivative compound

In a screening for small-molecule compounds that alleviate the deleterious effects of external CaCl(2) on zds1 Delta strain yeast, we found 2-((1-(hydroxymethyl) cyclohexyl) methyl) naphthalen-1-ol (NKH-7) to be an active compound. NKH-7 also inhibited cell growth at higher concentrations. To identify its target in growth inhibition, we isolated NKH-7-resistant mutants and selected those mutants that exhibited dominant or semi-dominant resistance specifically to NKH-7. By gene cloning, a TUB1 mutant gene encoding alpha-tubulin with a Ser248Pro mutation was identified. Deletion of the TUB3 gene, a minor gene encoding alpha-tubulin, led to supersensitivity to NKH-7. Cellular tubulin-containing arrays as visualized by green fluorescent protein (GFP)-labeled alpha-tubulin diminished rapidly on exposure to the inhibitor. The mutation was situated proximal to the alpha-beta interface of alpha-tubulin in microtubule protofilaments, suggesting the possibility that NKH-7 affects the hydrolysis of GTP bound to beta-tubulin. A functional connection perhaps exists between the tubulin inhibition and Ca(2+)-dependent cell-cycle regulation.     

Discovery of imidazo[1,2-a]pyridines as potent MCH1R antagonists

A series of imidazo[1,2-a]pyridine derivatives was identified and evaluated for MCH1R binding and antagonistic activity. Introduction of a methyl substituent at the 3-position of imidazo[1,2-a]pyridine provided compounds with a significant improvement in MCH1R affinity. Representative compounds in this series exhibited good potency and brain exposure in rats.     

The discovery of potent antagonists of NPBWR1 (GPR7)

The synthesis and evaluation of small molecule antagonists of the G protein-coupled receptor NPBWR1 (GPR7) are reported for the first time. [4-(5-Chloropyridin-2-yl)piperazin-1-yl][(1S,2S,4R)-4-{[(1R)-1-(4-methoxyphenyl)ethyl]amino}-2-(thiophen-3-yl)cyclohexyl]methanone (1) emerged as a hit from a high-throughput screen. Examination of substituents that focused on replacing the 5-chloropyridine and 4-methoxybenzylamino groups of 1 led to the identification of compounds that exhibited subnanomolar potencies as low as 660pM (9k) in the functional assay and 200pM in the binding assay (9i).     

Lead optimization of 4-(dimethylamino)quinazolines, potent and selective antagonists for the melanin-concentrating hormone receptor 1

The optimization of a series of 4-(dimethylamino)quinazoline antagonists of the melanin-concentrating hormone receptor 1 (MCH-R1) is described. The combination of the elaboration of both the linker portion and the terminal phenyl ring provided N-(cis-4-{[4-(dimethylamino)quinazolin-2-yl]amino}cyclohexyl)-3,4-difluorobenzamide hydrochloride 28 (ATC0175), which showed excellent antagonist activity at the MCH-R1 (IC50 = 3.4 nM) as well as good selectivity over the Y5 and the alpha2A receptors.     

Stereoselectivity of (R)- and (S)-hexahydro-difenidol binding to neuroblastoma M1, cardiac M2, pancreatic M3, and striatum M4 muscarinic receptors

(R)-Hexahydro-difenidol has a higher affinity for M1 receptors in NB-OK 1 cells, pancreas M3 and striatum M4 receptors (pKi 7.9 to 8.3) than for cardiac M2 receptors (pKi 7.0). (S)-Hexahydro-difenidol, by contrast, is nonselective (pKi 5.8 to 6.1). Our goal in the present study was to evaluate the importance of the hydrophobic phenyl, and cyclohexyl rings of hexahydro-difenidol for the stereoselectivity and receptor selectivity of hexahydro-difenidol binding to the four muscarinic receptors. Our results indicated that replacement of the phenyl ring of hexahydro-difenidol by a cyclohexyl group (----dicyclidol) and of the cyclohexyl ring by a phenyl moiety (----difenidol) induced a large (4- to 80-fold) decrease in binding affinity for all muscarinic receptors. Difenidol had a significant preference for M1, M3, and M4 over M2 receptors; dicyclidol, by contrast, had a greater affinity for M1 and M4 than for M2 and M3 receptors. The binding free energy decrease due to replacement of the phenyl and the cyclohexyl groups of (R)-hexahydro-difenidol by, respectively, a cyclohexyl and a phenyl moiety was almost additive in the case of M4 (striatum) binding sites. In the case of the cardiac M2, pancreatic M3, or NB-OK 1 M1 receptors the respective binding free energies were not completely additive. These results suggest that the four (R)-hexahydro-difenidol "binding moieties" (phenyl, cyclohexyl, hydroxy, and protonated amino group) cannot simultaneously form optimal interactions with the M1, M2, and M3 muscarinic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

1,2,4-triazolo[3,4-a]pyridine as a novel, constrained template for fibrinogen receptor (GPIIb/IIIa) antagonists

Conformationally constrained analogues of the GPIIb/IIIa antagonist elarofiban (RWJ-53308) have been synthesized and biologically evaluated. The 1,2,4-triazolo[3,4-a]pyridine scaffold provided potent antagonists with favorable pharmacodynamic and pharmacokinetic attributes in dogs. Compounds 12a and 13a exhibited enhancements in oral bioavailability, t(1/2), and ex vivo duration of action (inhibition of ADP-induced platelet aggregation) relative to elarofiban.     

Rational design, synthesis and structure-activity relationships of a cyclic succinate series of TNF-alpha converting enzyme inhibitors. Part 2: lead optimization

Modifications of the lead TACE inhibitor 1 (N-hydroxy-trans-2-[[4-(4-quinolinyloxymethyl)anilinyl]carbonyl]-1-cyclohexanecarboxamide) at the cyclohexyl ring and the quinoline moiety led to the identification of a series of piperidine containing TACE inhibitors with potent activity in the inhibition of TNF-alpha release in the whole blood assay (WBA). The most potent analogue IM491 [N-hydroxy-(5S,6S)-1-methyl-6-[[4-(2-methyl-4-quinolinylmethoxy)anilinyl]carbonyl]-5-piperidinecarboxamide] exhibited an IC(50) value of 20 nM in WBA with excellent selectivity over MMP-1, -2 and -9 and is orally bioavailable with an F value of 43% in beagle dogs.     

IgG Expression upon Oral Sensitization in Association with Maternal Exposure to Ovalbumin

The role of maternal allergen exposure in the allergenicity of the offspring remains controversial. Some studies have shown that maternal exposure is a risk factor for allergy in the offspring, whereas other studies have shown that maternal exposure induces immune tolerance and protects offspring from allergy disease. Therefore, we utilized maternal rat allergen exposure model to evaluate the offspring immune reactions to ovalbumin protein and to determine whether the Brown Norway (BN) rat model is a suitable animal model for studying the allergenicity of food proteins. For three generations, rats received an allergens or non-allergens by gavage during the pregnancy and lactation periods. After weaning, the offspring rats were used for oral sensitization experiment. In the sensitization experiment, the control rat, which had maternal exposure to phosphate-buffered saline (PBS), exhibited full response of IgG to oral exposure to OVA. The IgG level was significantly lower in F1 rats that were sensitized by maternal exposure to ovalbumin(OVA). Moreover, the lowest IgG level was found for the F3b sensitized by maternal rats exposed to OVA allergen for three continuous generations. Compared with maternal OVA exposure prior to postnatal sensitization, the sensitization via maternal PBS led to a higher serum level of OVA-specific IgG. However, the OVA-specific IgG levels for the two generations of maternal PBS exposure prior to postnatal sensitization was not higher than that for the one generation of maternal rats exposed to PBS prior to postnatal sensitization. Our studies demonstrate that maternal OVA exposure during the pregnancy and lactation can affect the results of oral sensitization studies using ovalbumin protein. BN rats must be bred in non-allergen conditions for at least one generation to avoid problems in rat models for studying the allergenicity of food proteins.     

Studies toward the discovery of the next generation of antidepressants. Part 6: Dual 5-HT1A receptor and serotonin transporter affinity within a class of arylpiperazinyl-cyclohexyl indole derivatives

Based on the previously reported discovery lead, 3-(cis-4-(4-(1H-indol-4-yl)piperazin-1-yl)cyclohexyl)-5-fluoro-1H-indole (2), a series of related arylpiperazin-4-yl-cyclohexyl indole analogs were synthesized then evaluated as 5-HT transporter inhibitors and 5-HT(1A) receptor antagonists. The investigation of the structure-activity relationships revealed the optimal pharmacophoric elements required for activities in this series. The best example from this study, 5-(piperazin-1-yl)quinoline analog (trans-20), exhibited equal binding affinities at 5-HT transporter (K(i)=4.9nM), 5-HT(1A) receptor (K(i)=6.2nM) and functioned as a 5-HT(1A) receptor antagonist.     

Fused bicyclic Gly-Asp beta-turn mimics with potent affinity for GPIIb-IIIa. Exploration of the arginine isostere

6-[4-Amidinobenzoyl]amino]-tetralone-2-acetic acid is a potent antagonist of GPIIb-IIIa. Substitution in the meta position of the benzamidine, or replacement with a heteroaryl amidine was tolerated in this series. Use of an acyl-linked 4-alkyl piperidine as an arginine isostere also provided active compounds. Compounds from this series provided substantial systemic exposure in the rat following oral administration.     

The Ketamine Analogue Methoxetamine and 3- and 4-Methoxy Analogues of Phencyclidine Are High Affinity and Selective Ligands for the Glutamate NMDA Receptor

In this paper we determined the pharmacological profiles of novel ketamine and phencyclidine analogues currently used as ‘designer drugs’ and compared them to the parent substances via the resources of the National Institute of Mental Health Psychoactive Drug Screening Program. The ketamine analogues methoxetamine ((RS)-2-(ethylamino)-2-(3-methoxyphenyl)cyclohexanone) and 3-MeO-PCE (N-ethyl-1-(3-methoxyphenyl)cyclohexanamine) and the 3- and 4-methoxy analogues of phencyclidine, (1-[1-(3-methoxyphenyl)cyclohexyl]piperidine and 1-[1-(4-methoxyphenyl)cyclohexyl]piperidine), were all high affinity ligands for the PCP-site on the glutamate NMDA receptor. In addition methoxetamine and PCP and its analogues displayed appreciable affinities for the serotonin transporter, whilst the PCP analogues exhibited high affinities for sigma receptors. Antagonism of the NMDA receptor is thought to be the key pharmacological feature underlying the actions of dissociative anaesthetics. The novel ketamine and PCP analogues had significant affinities for the NMDA receptor in radioligand binding assays, which may explain their psychotomimetic effects in human users. Additional actions on other targets could be important for delineating side-effects.     

Mechanistic pathways of antioxidant cytoprotection by a novel IH636 grape seed proanthocyanidin extract

To understand the bioavailability and mechanistic pathways of cytoprotection by IH636 grape seed proanthocyanidin extract (GSPE, commercially known as ActiVin) a series of in vitro and in vivo studies were conducted. Comparative protective abilities of GSPE, and vitamins C and E, singly and in combination, were assessed against smokeless tobacco extract (STE)-induced oxidative stress, DNA fragmentation and apoptotic cell death in a primary culture of normal human oral keratinocytes. GSPE protected against STE-induced oxidative stress, DNA damage and apoptotic cell death, and provided better protection as compared to vitamins C and E, singly and in combination. The bioavailability and protective ability of GSPE were examined against acetaminophen (AP)-induced hepato- and nephrotoxicity, amiodarone (AM)-induced lung toxicity, doxorubicin (DX)-induced cardiotoxicity and dimethylnitrosamine (DM)-induced spleenotoxicity in mice. GSPE-fed animals were compared with GSPE-untreated mice to evaluate the protective ability of GSPE against these structurally diverse drugs/chemicals. Serum chemistry changes, histopathology and DNA damage were evaluated. Results indicate that GSPE preexposure prior to the drugs/chemicals such as AP, AM, DX or DM treatment, provided near complete protection in terms of serum chemistry changes and inhibition of both forms of cell death, e.g., apoptosis and necrosis. DNA damage in various tissues triggered by these agents was significantly reduced in GSPE-fed animals. Histopathological examination of multiple target organs provided similar data. The results suggest that GSPE exposure is bioavailable and provides significant multiorgan protection against structurally diverse drug- and chemical-induced toxic assaults. Further, these studies exhibited a series of mechanistic information including free radical scavenging ability, anti-endonucleolytic activity, cytochrome P450 2E1 inhibitory activity, anti-necrotic, anti-apoptotic and anti-carcinogenic activities, modulatory effects on antioxidative and apoptotic regulatory genes such as Bcl2, c-myc and p53, which may be responsible for the novel chemoprotective properties exhibited by GSPE.     

Evaluation of intrinsic spectroscopic properties of chromophore assemblies by shielding with cyclohexyl base pairs within a DNA duplex

Here, we investigated spectroscopic behaviors of tetramethylrhodamine (TMR) homo- and hetero-dimers within DNA duplex. In order to shield the chromophores from natural base pairs, we used cyclohexyl base pairs as ‘insulators’; these pairs were inserted between the chromophores and nucleobases. When a single TMR moiety was sandwiched between cyclohexyl base pairs, the emission intensity increased by fivefold relative to a TMR between natural base pairs, because electron transfer from nucleobases was suppressed. Next, we inserted two TMRs between the cyclohexyl base pairs and found that they facilitated H-dimer formation of TMR; a distinct hypsochromic shift was induced only when cyclohexyl base pairs were inserted. We further examined quenching behavior of a TMR paired with a quencher dye between cyclohexyl base pairs. Interestingly, fluorescence from TMR was quenched by nitro methyl red more efficiently in the presence of cyclohexyl base pairs than in their absence. This suggests that neighboring natural base pairs disturbed electron or hole transfer between the fluorophore and the quencher. The cyclohexyl base pairs shielded the chromophore pair from the natural base pairs and allowed intrinsic electron transfer.     

Synthesis and phosphodiesterase 5 inhibitory activity of new 5-phenyl-1,6-dihydro-7H-pyrazolo[4,3-d]pyrimidin-7-one derivatives containing an N-acylamido group phenyl ring.

New sildenafil analogues with an N-acylamido group at the 5'-position of the phenyl ring, 6a--e, were prepared from the readily available starting compound 2 in four straightforward steps. Enzyme assays demonstrated that all the target compounds 6a-e showed higher PDE5 inhibitory activities than sildenafil. It was observed that the PDE5 inhibitory activity was enhanced as the chain length of R group increased, but introduction of the branched alkyl groups such as isopropyl (6d) and cyclohexyl (6e) resulted in the drop of potency compared with 6c. In particular the N-butyrylamido derivative 6c exhibited the highest PDE5 inhibitory activity, and was about 6-fold more potent than sildenafil. However, all the compounds exhibited somewhat weak selectivity (1--3-fold) over PDE6, indicating that the compounds 6a--e have intrinsically lower selectivity than sildenafil.