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Class B G-protein-coupled receptors (GPCRs) remain an underexploited target for drug development. The calcitonin receptor (CTR) family is particularly challenging, as its receptors are heteromers comprising two distinct components: the calcitonin receptor-like receptor (CLR) or calcitonin receptor (CTR) together with one of three accessory proteins known as receptor activity-modifying proteins (RAMPs). CLR/RAMP1 forms a CGRP receptor, CLR/RAMP2 forms an adrenomedullin-1 (AM1) receptor, and CLR/RAMP3 forms an adrenomedullin-2 (AM2) receptor. The CTR/RAMP complexes form three distinct amylin receptors. While the selective blockade of AM2 receptors would be therapeutically valuable, inhibition of AM1 receptors would cause clinically unacceptable increased blood pressure. We report here a systematic study of structure–activity relationships that has led to the development of first-in-class AM2 receptor antagonists. These compounds exhibit therapeutically valuable properties with 1000-fold selectivity over the AM1 receptor. These results highlight the therapeutic potential of AM2 antagonists.

[Image: see text] The hormone adrenomedullin has both physiological and pathological roles in biology. As a potent vasodilator, adrenomedullin is critically important in the regulation of blood pressure, but it also has several roles in disease, of which its actions in cancer are becoming recognized to have clinical importance. Reduced circulating adrenomedullin causes increased blood pressure but also reduces tumor progression, so drugs blocking all effects of adrenomedullin would be unacceptable clinically. However, there are two distinct receptors for adrenomedullin, each comprising the same G protein-coupled receptor (GPCR), the calcitonin receptor-like receptor (CLR), together with a different accessory protein known as a receptor activity-modifying protein (RAMP). The CLR with RAMP2 forms an adrenomedullin-1 receptor, and the CLR with RAMP3 forms an adrenomedullin-2 receptor. Recent research suggests that a selective blockade of adrenomedullin-2 receptors would be therapeutically valuable. Here we describe the design, synthesis, and characterization of potent small-molecule adrenomedullin-2 receptor antagonists with 1000-fold selectivity over the adrenomedullin-1 receptor, although retaining activity against the CGRP receptor. These molecules have clear effects on markers of pancreatic cancer progression in vitro, drug-like pharmacokinetic properties, and inhibit xenograft tumor growth and extend life in a mouse model of pancreatic cancer. Taken together, our data support the promise of a new class of anticancer therapeutics as well as improved understanding of the pharmacology of the adrenomedullin receptors and other GPCR/RAMP heteromers.

Orexin receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on Orexin receptors [42]) are activated by the endogenous polypeptides orexin-A and orexin-B (also known as hypocretin-1 and -2; 33 and 28 aa) derived from a common precursor, preproorexin or orexin precursor, by proteolytic cleavage and some typical peptide modifications [109]. Currently the only orexin receptor ligands in clinical use are suvorexant and lemborexant, which are used as hypnotics. Orexin receptor crystal structures have been solved [134, 133, 54, 117, 46].

The NLRP3 inflammasome is a component of the innate immune system involved in the production of proinflammatory cytokines. Aberrant activation by a wide range of exogenous and endogenous signals can lead to chronic, low-grade inflammation. It has attracted a great deal of interest as a drug target due to the association with diseases of large unmet medical need such as Alzheimer's disease, Parkinson's disease, arthritis, and cancer. To date, no drugs specifically targeting inhibition of the NLRP3 inflammasome have been approved. In this work, we used the known NLRP3 inflammasome inhibitor CP-456,773 (aka CRID3 or MCC 950) as our starting point and undertook a Structure-Activity Relationship (SAR) analysis and subsequent scaffold-hopping exercise. This resulted in the rational design of a series of novel ester-substituted urea compounds that are highly potent and selective NLRP3 inflammasome inhibitors, as exemplified by compounds 44 and 45. It is hypothesized that the ester moiety acts as a highly permeable delivery vehicle and is subsequently hydrolyzed to the carboxylic acid active species by carboxylesterase enzymes. These molecules are greatly differentiated from the state-of-the-art and offer potential in the treatment of NLRP3-driven diseases, particularly where tissue penetration is required.

Adenosine is important messenger molecule that is involved in signal transmission in central nervous, cardiovascular and immunological systems. The immunosuppressive role of adenosine attract attention of many researchers recently and reversal of that effect is addressed by several different approaches i.e.: inhibition of enzymes that are producing the adenosine in tumor microenvironment like CD39 and CD73 but also the antagonization of adenosine receptors mainly A2A and A2B subtypes. Here we present a novel series of unsurmountable, dual A2A/A2B antagonists that retain its nanomolar potency in tumor-like adenosine-rich environment. This advantageous profile comes mainly from long residence time in adenosine receptors which transfers into high receptor occupancy even after full clearance of the compound from plasma. We are demonstrating how our inhibitors restore adenosine depleted immune functionality in the series of functional in vitro assays in primary cells. This includes the cytokine release by activated CD4+ and CD8+ human T-lymphocytes (driven by A2A) and dendritic cells (driven by A2B). Most importantly presented compounds show improved in vivo pharmacological profile in comparison to adenosine inhibitors currently tested in clinical trials manifested by high inhibition of CREB phosphorylation in mouse model. Citation Format: Michal Galezowski, Paulina Węgrzyn, Aneta Bobowska, Katarzyna Dziedzic, Joanna Szeremeta-Spisak, Marcin Nowogrodzki, Grzegorz Satala, Alicja Obara, Iwona Lozinska-Raj, Marcelina Dudek, Anita Janiga, Jacek Reus, Marek Wronowski, Magdalena Zastawna, Grzegorz Statkiewicz, Maciej Rogacki, Magdalena Ziembik, Karolina Grycuk, Foteini Soukou, Kinga Michalik, Agnieszka Adamus, Karolina Wiatrowska, Natalia Lewandowska, Aniela Golas, Olga Haberkiewicz, Rod Porter, Krzysztof Brzozka, Mateusz Nowak. Novel dual A2A A2B adenosine receptor antagonists for cancer immunotherapy: in vitro and in vivo characterization [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4135.

A screening hit was used as the basis for the core structure of a new series of acylglycinamide GlyT-1 inhibitors. Investigation of the SAR around four areas of diversity used facile chemistry to prepare compounds quickly. By focussing on reducing the lipophilicity and improving the aqueous solubility in the series we were able to prepare a compound (17e) with a good level of activity at GlyT-1, selectivity over GlyT-2 and moderate oral bioavailability.

The type-1 glycine transporter (GlyT1) is an important target for the development of new medications for schizophrenia. A specific and selective positron emission tomography (PET) GlyT1 ligand would facilitate drug development studies to determine whether a drug reaches this target and help establish suitable doses for clinical trials. This article describes the evaluation of three candidate GlyT1 PET radioligands (GSK931145, GSK565710, and GSK991022) selected from a library of compounds based on favorable physicochemical and pharmacological properties. Each candidate was successfully labeled using [11C]methyl iodide or [11C]methyl triflate and administered to a pig pre- and postadministration with a pharmacological dose of a GlyT1 inhibitor to determine their suitability as PET ligands in the porcine brain in vivo. All three candidate ligands were analyzed quantitatively with compartment analyses employing a plasma input function. [11C]GSK931145 showed good brain penetration and a heterogeneous distribution in agreement with reported GlyT1 localization. Following pretreatment with GSK565710, uptake of [11C]GSK931145 was reduced to homogeneous levels. Although [11C]GSK565710 also showed good brain penetration and a heterogeneous distribution, the apparent level of specific binding was reduced compared to [11C]GSK931145. In contrast, [11C]GSK991022 showed a much lower brain penetration and resultant signal following pretreatment with GSK565710. Based on these findings [11C]GSK931145 was identified as the most promising ligand for imaging GlyT1 in the porcine brain, possessing good brain penetration, specific signal, and reversible kinetics. [11C]GSK931145 is now being progressed into higher species. Synapse 64:542–549, 2010. © 2010 Wiley-Liss, Inc.

High throughput screening led to the discovery of a novel series of 1,3-diaminopropan-2-ol sulfonamides as selective GlyT-1 inhibitors. Structure-activity relationships of this novel series and optimisation of the initial hit that led to the identification of (2), a potent and selective GlyT-1 inhibitor, are also presented.

Acute systemic treatment with the selective orexin-1 receptor antagonist SB-334867 (30 mg/kg, i.p.) has been reported not only to inhibit food intake and to accelerate behavioural satiety in rats, but also to produce a significant loss of bodyweight over the 24 h period post-dosing. The present studies were designed to test the hypothesis that the inhibition of weight gain following acute treatment with SB-334867 is due to a persistent anorectic action of the compound. In Experiment 1, the acute effects of SB-334867 (30 mg/kg, i.p.) on food intake and behaviour in a 1 h test with palatable mash were assessed as a function of injection-test interval. Results confirmed that, when administered 30 min prior to testing, SB-334867 significantly suppressed mash intake and accelerated behavioural satiety. More importantly, significant anorexia and behavioural change were also observed when animals were tested 24 h, but not 48 h, post-dosing. As previously reported, all animals treated with the orexin-1 receptor antagonist lost bodyweight over the 24 h period following acute treatment. The generality of these findings was confirmed in Experiment 2, where acute treatment with SB-334867 (30 mg/kg, i.p.) significantly suppressed home cage chow consumption over the 24 h period post-dosing, an effect also accompanied by a significant loss of bodyweight. The results of Experiment 3 showed that, following i.p. administration of 30 mg/kg, SB-334867 has good CNS penetration, reaches peak plasma and brain concentrations at 30 min, and maintains good exposure over 4 h post-dosing. Overall, current data support the hypothesis that a persistent anorectic action contributes to the significant loss of bodyweight observed 24 h following acute dosing with SB-334867. As the compound is virtually undetectable in plasma or brain beyond 8 h post-dosing, and since nothing is known about potentially active metabolites, we consider the possibility that single dose treatment with SB-334867 results in enduring alterations to the orexin-1 receptor and/or downstream signalling pathways.

This study characterises the binding of a novel nonpeptide antagonist radioligand, [3H]SB-674042 (1-(5-(2-fluoro-phenyl)-2-methyl-thiazol-4-yl)-1-((S)-2-(5-phenyl-(1,3,4)oxadiazol-2-ylmethyl)-pyrrolidin-1-yl)-methanone), to the human orexin-1 (OX1) receptor stably expressed in Chinese hamster ovary (CHO) cells in both a whole cell assay and in a cell membrane-based scintillation proximity assay (SPA) format. Specific binding of [3H]SB-674042 was saturable in both whole cell and membrane formats. Analyses suggested a single high-affinity site, with Kd values of 3.76±0.45 and 5.03±0.31 nM, and corresponding Bmax values of 30.8±1.8 and 34.4±2.0 pmol mg protein−1, in whole cell and membrane formats, respectively. Kinetic studies yielded similar Kd values. Competition studies in whole cells revealed that the native orexin peptides display a low affinity for the OX1 receptor, with orexin-A displaying a ∼five-fold higher affinity than orexin-B (Ki values of 318±158 and 1516±597 nM, respectively). SB-334867, SB-408124 (1-(6,8-difluoro-2-methyl-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) and SB-410220 (1-(5,8-difluoro-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) all displayed high affinity for the OX1 receptor in both whole cell (Ki values 99±18, 57±8.3 and 19±4.5 nM, respectively) and membrane (Ki values 38±3.6, 27±4.1 and 4.5±0.2 nM, respectively) formats. Calcium mobilisation studies showed that SB-334867, SB-408124 and SB-410220 are all functional antagonists of the OX1 receptor, with potencies in line with their affinities, as measured in the radioligand binding assays, and with approximately 50-fold selectivity over the orexin-2 receptor. These studies indicate that [3H]SB-674042 is a specific, high-affinity radioligand for the OX1 receptor. The availability of this radioligand will be a valuable tool with which to investigate the physiological functions of OX1 receptors. British Journal of Pharmacology (2004) 141, 340–346. doi:10.1038/sj.bjp.0705610

The treatment of schizophrenia has long been dominated by aminergic receptor antagonist-based therapeutics largely founded on the dopamine hypothesis of schizophrenia. More recently the glutamatergic theory has come to the fore which may potentially address some of the deficiencies of current therapies. While there are many approaches to manipulating the glutamatergic system, the most advanced approach is to increase synaptic concentrations of the NMDA receptor co-agonist glycine via inhibition of the glycine transporter 1 (GlyT-1). Here we will describe the background biological rationale for this approach and review the diverse classes of compounds which have been identified as GlyT-1 inhibitors with particular focus on the identification of those molecules which have entered the clinical stages of development. The role of target kinetics in drug action, a review of the rich vein of PET ligand development and their use in clinical development and the status of clinical-stage compounds will be addressed. Finally there is a discussion of some of the issues that have arisen with the discovery and development of GlyT-1 inhibitors and the prospects for the future of this mechanistic approach.

This communication reports SARs for the first orexin-1 receptor antagonist series of 1-aryl-3-quinolin-4-yl and 1-aryl-3-naphthyridin-4-yl ureas. One of these compounds, 31 (SB-334867), has excellent selectivity for the orexin-1 receptor, blood-brain barrier permeability and shows in vivo activity following ip dosing.

Intracerebroventricular (i.c.v.) administration of the novel hypothalamic neuropeptide orexin-A stimulates food intake in rats, and delays the onset of behavioural satiety (i.e. the natural transition from feeding to resting). Furthermore, preliminary findings with the selective orexin-1 receptor antagonist, SB-334867, suggest that orexin-A regulation of food intake is mediated via the orexin-1 receptor. At present, however, little is known about either the intrinsic effects of SB-334867 on the normal structure of feeding behaviour, or its effects upon orexin-A-induced behavioural change. In the present study, we have employed a continuous monitoring technique to characterize the effects of SB-334867 (3-30 mg/kg, i.p.) on the microstructure of rat behaviour during a 1-h test with palatable wet mash. Administered alone, SB-334867 (30 mg/kg, but not lower doses) significantly reduced food intake and most active behaviours (eating, grooming, sniffing, locomotion and rearing), while increasing resting. Although suggestive of a behaviourally nonselective (i.e. sedative) action, the structure of feeding behaviour was well-preserved at this dose level, with the reduction in behavioural output clearly attributable to an earlier onset of behavioural satiety. As previously reported, orexin-A (10 microg per rat i.c.v.) stimulated food intake, increased grooming and delayed the onset of behavioural satiety. Pretreatment with SB-334867 dose-dependently blocked these effects of orexin-A, with significant antagonism evident at dose levels (3-10 mg/kg) below those required to produce intrinsic behavioural effects under present test conditions. Together, these findings strongly support the view that orexin-A is involved in the regulation of feeding patterns and that this influence is mediated through the orexin-1 receptor.

The pharmacology of various peptide and non-peptide ligands was studied in Chinese hamster ovary (CHO) cells stably expressing human orexin-1 (OX1) or orexin-2 (OX2) receptors by measuring intracellular calcium ([Ca2+]i) using Fluo-3AM. Orexin-A and orexin-B increased [Ca2+]i in CHO-OX1 (pEC50=8.38±0.04 and 7.26±0.05 respectively, n=12) and CHO-OX2 (pEC50=8.20±0.03 and 8.26±0.04 respectively, n=8) cells. However, neuropeptide Y and secretin (10 pM – 10 μM) displayed neither agonist nor antagonist properties in either cell-line. SB-334867-A (1-(2-Methyylbenzoxanzol-6-yl)-3-[1,5]naphthyridin-4-yl-urea hydrochloride) inhibited the orexin-A (10 nM) and orexin-B (100 nM)-induced calcium responses (pKB=7.27±0.04 and 7.23±0.03 respectively, n=8), but had no effect on the UTP (3 μM)-induced calcium response in CHO-OX1 cells. SB-334867-A (10 μM) also inhibited OX2 mediated calcium responses (32.7±1.9% versus orexin-A). SB-334867-A was devoid of agonist properties in either cell-line. In conclusion, SB-334867-A is a non-peptide OX1 selective receptor antagonist. British Journal of Pharmacology (2001) 132, 1179–1182; doi:10.1038/sj.bjp.0703953

Rationale: Orexins A and B have recently been discovered and shown to be derived from prepro-orexin, primarily expressed in the rat hypothalamus. Orexin-A has been ascribed a number of in vivo functions in the rat after intracerebroventricular (ICV) administration, including hyperphagia, neuroendocrine modulation and, most recently, evidence for a behavioural response characterised by an increase in grooming. Objectives: Here, we have investigated the orexin-receptor subtypes involved in the grooming response to orexin-A (3 μg, ICV) in the rat. Methods: Male rats, habituated to clear Perspex behavioural observation boxes, were pretreated with antagonists with mixed selectivity for OX1, OX2, 5-HT2B and 5-HT2C receptor subtypes prior to the administration of orexin-A and the intense grooming response elicited by this peptide assessed. Results: Pretreatment of rats with a mixed OX1/5-HT2B/2C receptor antagonist 1-(4-methylsulfanylphenyl)-3-quinolin-4-ylurea (SB-284422), revealed a significant, but incomplete, blockade of orexin-A-induced grooming. Despite the low potency of orexin-A at 5-HT2B and 5-HT2C receptors in vitro (pKi

Rationale: Orexin-A and orexin-B are hypothalamic neuropeptides derived from a 130-amino acid precursor, prepro-orexin, and are potent agonists at both the orexin-1 (OX1) and orexin-2 (OX2) receptors. Orexin-A has been ascribed a number of in vivo functions in the rat after intracerebroventricular (ICV) administration, including hyperphagia, neuroendocrine modulation and a role in the regulation of sleep-wake function. The in vivo role of orexin-B is not as clear. Objectives: To investigate the behavioural, endocrine and neurochemical effects of orexin-B in in-vivo tests. In a number of experiments, these effects were compared with those of orexin-A. Methods: Experiments were carried out in male, Sprague-Dawley rats with a guide cannula directed towards the lateral ventricle. The effects of orexin-B (ICV) upon grooming behaviour were compared with those of orexin-A. The effects of orexin-B upon the motor activity response to both novel and familiar environments were assessed in an automated activity monitor. Orexin-B was tested upon startle reactivity and body temperature. Further, plasma hormones and [DOPAC+HVA]/[DA] and [5-HIAA]/[5-HT] ratios in six brain areas were measured 40 min post-orexin-B or orexin-A. Results: The clearest behavioural response to orexin-B was increased motor activity in both novel and familiar environments. Orexin-B-induced hyperactivity was blocked by an OX1 receptor antagonist, SB-334867-A, implicating OX1 receptors in this behavioural response. In common with orexin-A, orexin-B reduced plasma prolactin and failed to influence startle reactivity. However, in contrast with orexin-A, orexin-B increased head grooming but failed to cause a robust whole body grooming response or increase plasma corticosterone levels. Further, orexin-B, but not orexin-A, increased plasma TSH and increased hypothalamic and striatal [5-HIAA]/[5-HT] ratios. Conclusions: The present study has demonstrated a number of behavioural, neuroendocrine and neurochemical effects of orexin-B that distinguish it from orexin-A. Further, we have demonstrated a role for OX1 receptors in the actions of orexin-B upon motor activity.

A variety of evidence implicates the orexins, especially orexin-A, in the regulation of food intake, but it has not been established whether this effect is mediated by the orexin-1 or orexin-2 receptor. In the present study, a selective orexin-1 receptor antagonist, 1-(2-methylbenzoxazol-6-yl)-3-[1,5]naphthyridin-4-yl urea hydrochloride (SB-334867-A), was administered intraperitoneally to rats under various conditions, and food consumption was subsequently measured over 24 h. In male rats, a single dose of SB-334867-A (30 mg/kg, i.p.) given during the light phase reduced both orexin-A-induced food intake (7 nmol, i.c.v.) and feeding stimulated by an overnight fast for 4 h. When given at the start of the dark phase, food consumption was reduced in both male and female rats over 24 h. Daily injections at the start of the dark phase for 3 days reduced natural feeding in male rats over 24 h on days one and three. These findings demonstrate direct inhibition of orexin-A induced food intake with a selective orexin-1 receptor antagonist. Furthermore, the suppression of nocturnal feeding and food intake stimulated by an overnight fast supports other evidence that orexin-A is involved in the regulation of natural feeding and suggests that orexin-1 receptor antagonists could be useful in the treatment of obesity.

From two related series of 2-(alkylthio)-pyrimidones, a novel series of 1-((amidolinked)-alkyl)-pyrimidones has been designed as nanomolar inhibitors of human lipoprotein-associated phopholipase A 2 . These compounds show greatly enhanced activity in isolated plasma. Selected derivatives such as compounds 51 and 52 are orally active with a good duration of action.

Simple and rapid single-pot preparations of substituted 2-mercaptobenzaldehydes 5 and 2-substituted benzo[ b ]thiophenes 3 based on ortho -lithiation methodology are described.

The neuropeptide NK3 receptor is expressed almost exclusively within the mammalian nervous system and its localization is commensurate with a role in modulating central monoaminergic neurotransmission. Following on from our previous work we review the rationale for NK3 receptor antagonists as wide spectrum antipsychotics and the recent scientific and patent literature that has highlighted new chemical strategies to identify selective NK3 and dual activity NK1/3 receptor ligands for the putative treatment of schizophrenia. We discuss the emerging structural biology and its use in the design of molecules with increased structural diversity and predictable receptor pharmacology. Particular attention is paid to the progress in improving ligand drug-like properties. The status of imaging and the development of translational technologies in the neurokinin field are also discussed. Finally, we summarize the available clinical information on the compounds that have progressed into psychiatric patient populations and evaluate the potential therapeutic utility of NK3 receptor targeted ligands.

The hypothalamic peptides orexin-A and orexin-B are potent agonists of two G-protein coupled receptors, namely the OX(1) and the OX(2) receptor. These receptors are widely distributed, though differentially, in the rat brain. In particular, the OX(1) receptor is highly expressed throughout the hypothalamus, whilst the OX(2) receptor is mainly located in the ventral posterior nucleus. A large body of compelling evidence, both pre-clinical and clinical, suggests that the orexin system is profoundly implicated in sleep disorders. In particular, modulation of the orexin receptors activation by appropriate antagonists was proven to be an efficacious strategy for the treatment of insomnia in man. A novel, drug-like bis-amido piperidine derivative was identified as potent dual OX(1) and OX(2) receptor antagonists, highly effective in a pre-clinical model of sleep.

Inhibitors of the glycine transporter GlyT-1 are being developed as potential treatments for schizophrenia. Here we report on the use of two novel radioligands, [ 3 H]-SB-733993 and [ 3 H]-GSK931145, for the characterisation of GlyT-1 in both cells and native tissue. Binding was evaluated in membranes either from HEK293 cells expressing recombinant human GlyT-1 (hGlyT-1) or from rat cerebral cortex. Specific binding of both [ 3 H]-SB-733993 and [ 3 H]-GSK931145 to hGlyT-1 HEK293 cell membranes and rat cerebral cortex membranes was saturable and comprised >90% of total binding. K d and B max values for the two radioligands were fairly similar, with K d values of 1–2 nM and B max values of around 7000 fmol/mg protein in hGlyT-1 membranes and 3000 fmol/mg protein in rat cortex membranes. Association of [ 3 H]-SB-733993 was faster, with binding reaching equilibrium within 30 min compared with 90 min for [ 3 H]-GSK931145. Dissociation was also much slower for [ 3 H]-GSK931145 than for [ 3 H]-SB-733993, with 50% of specific binding being dissociated by approximately 40 min and 5 min, respectively. Autoradiography studies with [ 3 H]-GSK931145 showed widespread distribution of binding in rat brain, with generally higher binding in caudal compared with rostral areas. Initial studies in human frontal cortex membranes showed clear specific binding of [ 3 H]-GSK931145, though with much lower density ( B max 570 fmol/mg protein) and slightly lower affinity (K d 4.5 nM) compared with rat cortex. A human brain autoradiography study showed higher specific binding in cerebellum compared with frontal cortex. All GlyT-1 inhibitors tested, as well as glycine itself, competed fully for the binding of both [ 3 H]-SB-733993 and [ 3 H]-GSK931145 in both hGlyT-1 and rat cortex membranes. Studies on the effect of varying NaCl concentration showed that [ 3 H]-SB-733993 binding was reduced by >90% in the absence of added Na + ions, whilst [ 3 H]-GSK931145 binding was unaffected. Glycine produced concentration-dependent decreases in binding affinity of both radioligands without major changes in B max values, suggesting that both [ 3 H]-SB-733993 and [ 3 H]-GSK931145 bind to sites on GlyT-1 that are orthosteric to the site at which glycine itself binds. Overall, these results show that both [ 3 H]-SB-733993 and [ 3 H]-GSK931145 are useful radioligands for studies on GlyT-1 in both cell lines and native tissues, with [ 3 H]-GSK931145 being the radioligand of choice for further studies on GlyT-1 expression and pharmacology.

SC-44368 (5-[6-(1-cyclohexyl-1H-tetrazol-5-y)hexyl]-1,8-naphthyridin-2(1H)-one) is a potent and selective competitive inhibitor of platelet cyclic AMP-dependent phosphodiesterase (cAMP-PDE) (Ki: 1.65 μM). For the phosphodiesterase isoenzyms from human platelets SC-44368 shows a 26-fold selectivity (IC50 ratio) for the inhibition of the cAMP-PDE over the cyclic GMP-dependent phosphodiesterase (cGMP-PDE). By comparison, 3-isobutyl-1-methyl-xanthine (IBMX) inhibited the cAMP-PDE and cGMP-PDE from human platelets with approximately equal efficacy. Broad inhibitory activity was evident against human platelet aggregatory responses in vitro. IC50 values of 18.1 ± 5.3 μM (25 nM platelet activating factor, PAF), 17.3 ± 3.0 μM (1.0 μg/ml collagen) and 24.2 ± 10.3 μM (1μM ADP) were obtained against maximum increases in platelet-rich plasma (PRP) light transmission achieved by each agonist. SC-44368 potentiated the prostacyclin-induced increase of intra-platelet cAMP levels but did not potentiate the sodium nitroprusside-induced increase of intraplatelet cGMP levels. In an ex vivo model of platelet aggregation SC-44368 (3 mg/kg, i.v.) produced a potent inhibition of collagen-induced platelet aggregation. SC-44368 produced only weak hypotensive activity in the rat. Thus, SC-44368 is a novel cAMP-PDE inhibitor which possesses potent, broad spectrum anti-aggregatory properties.

The pharmacology of the orexin-like peptides, hypocretin-1 and hypocretin-2, was studied in Chinese hamster ovary (CHO) cells stably expressing orexin-1 (OX1) or orexin-2 (OX2) receptors by measuring intracellular calcium ([Ca2+]i) using Fluo-3AM. Orexin-A and orexin-B increased [Ca2+]i in CHO-OX1 (pEC50=7.99±0.05 and 7.00±0.10 respectively, n=8) and CHO-OX2 (pEC50=8.30±0.05 and 8.21±0.07 respectively, n=5). However, hypocretin-1 and hypocretin-2 were markedly less potent, with pEC50 values of 5.31±0.04 and 5.41±0.04 respectively in CHO-OX2 cells (n=5). In CHO-OX1 cells 10 μM hypocretin-1 only elicited a 37.5±3.4% response whilst 10 μM hypocretin-2 elicited a 18.0±2.1% response (n=8). Desensitisation of OX1 or OX2 with orexin-A (100 nM) abolished the response to orexin-A (10 nM) and the hypocretins (10 μM), but not to UTP (3 μM). In conclusion, the hypocretins are only weak agonists at the orexin receptors. British Journal of Pharmacology (2000) 129, 1289–1291; doi:10.1038/sj.bjp.0703257

Glycine can act as either an inhibitory neurotransmitter or as a potentiator of NMDA-dependent excitatory neurotransmission. There is some evidence that glycine can have both pro- and anticonvulsant properties in various rodent models of epilepsy. In the present study we tested several glycine transporter 1 (GlyT1) inhibitors including NFPS, SSR 504734, Lu AA21279, Org 25935, SB-710622, GSK931145, as well as the glycine agonist d -serine, in the maximal electroshock threshold (MEST) test in the rat. In a series of experiments, male Sprague–Dawley rats (n = 12/group) were pre-treated with a compound of interest and then received an electric shock delivered via corneal electrodes. A cohort of satellite animals (n = 3/group) was also used to measure blood and brain levels of Org 25935. All GlyT1 inhibitors increased seizure thresholds dose-dependently, indicative of anticonvulsant activity. SB-710622 and GSK931145 had lower minimum effective doses (MEDs) in the MEST test than other GlyT1 inhibitors. At estimated tmax, increases in dose administered were paralleled by increases in blood and brain concentrations of Org 25935. Thus, increasing extracellular concentration of glycine via inhibition of its uptake protects from electroshock-induced seizures in the rat. Whether strychnine-sensitive or strychnine-insensitive glycine binding sites are involved in this effect remains to be determined.

This study was designed to investigate whether brain unbound concentration (C u,brain ) is a better predictor of dopamine D 2 receptor occupancy than total brain concentration, cerebrospinal fluid concentration (C CSF ), or blood unbound concentration (C u,blood ). The ex vivo D 2 receptor occupancy and concentration-time profiles in cerebrospinal fluid, blood, and brain of six marketed antipsychotic drugs were determined after oral administration in rats at a range of dose levels. The C u,brain was estimated from the product of total brain concentration and unbound fraction, which was determined using a brain homogenate method. In conclusion, the C u,brain of selected antipsychotic agents is a good predictor of D 2 receptor occupancy in rats. Furthermore, C u,brain seems to provide a better prediction of D 2 receptor occupancy than C CSF or C u,blood for those compounds whose mechanism of entry into brain tissue is influenced by factors other than simple passive diffusion.

Lead optimisation starting from the previously reported selective quinoline NK3 receptor antagonists talnetant 2 (SB-223412) and 3 (SB-222200) led to the identification of 3-aminoquinoline NK3 antagonist 10 (GSK172981) with excellent CNS penetration. Investigation of a structurally related series of sulfonamides with reduced lipophilicity led to the discovery of 20 (GSK256471). Both 10 and 20 are high affinity, potent NK3 receptor antagonists which despite having different degrees of CNS penetration produced excellent NK3 receptor occupancy in an ex vivo binding study in gerbil cortex.

The penetration of drugs into the central nervous system is a composite of both the rate of drug uptake across the blood-brain barrier and the extent of distribution into brain tissue compartments. Clinically, positron emission tomography (PET) is the primary technique for deriving information on drug biodistribution as well as target receptor occupancy. In contrast, rodent models have formed the basis for much of the current understanding of brain penetration within pharmaceutical Drug Discovery. Linking these two areas more effectively would greatly improve the translation of candidate compounds into therapeutic agents. This paper examines two of the major influences on the extent of brain penetration across species, namely plasma protein binding and brain tissue binding. An excellent correlation was noted between unbound brain fractions across species (R(2) > 0.9 rat, pig, and human, n = 21), which is indicative of the high degree of conservation of the central nervous system environment. In vitro estimates of human brain-blood or brain-plasma ratios of marketed central nervous system drugs and PET tracers agree well with in vivo values derived from clinical PET and post-mortem studies. These results suggest that passive diffusion across the blood-brain barrier is an important process for many drugs in humans and highlights the possibility for improved prediction of brain penetration across species.

The dispositions of 50 marketed central nervous system (CNS) drugs into the brain have been examined in terms of their rat in situ (P) and in vitro apparent membrane permeability (P(app)) alongside lipophilicity and free fraction in rat brain tissue. The inter-relationship between these parameters highlights that both permeability and brain tissue binding influence the uptake of drugs into the CNS. Hydrophilic compounds characterized by low brain tissue binding display a strong correlation (R(2) = 0.82) between P and P(app), whereas the uptake of more lipophilic compounds seems to be influenced by both P(app) and brain free fraction. A nonlinear relationship is observed between logP(oct) and P over the 6 orders of magnitude range in lipophilicity studied. These findings corroborate recent reports in the literature that brain penetration is a function of both rate and extent of drug uptake into the CNS.

Acute systemic treatment with the selective orexin-1 (OX1R) antagonist SB-334867 reduces food intake in rats, an effect associated with an acceleration in behavioural satiety and unrelated to gross behavioural disruption, alterations in palatability, or toxicity. However, as enhanced satiety is behaviourally indexed by an earlier-than-normal transition from eating to resting, and since orexin-A has been implicated in mechanisms of arousal, it remains possible that sedation contributes to the anorectic effect of acute OX1R blockade. Previous work has shown that, when treated with SB-334867 (30 mg/kg, i.p.) 30 min before a 1h test with palatable food, rats begin to show appreciable levels of resting 10-15 min earlier than under control conditions (i.e. around 20 min versus 30-35 min into the session). The present results demonstrate that a 20 min increase in the injection-test interval (i.e. 50 min) had no significant impact on the anorectic, behavioural or weight gain effects of SB-334867 in non-deprived male rats. Most importantly, this altered treatment regimen led to a temporal profile of resting virtually identical to that previously observed with the more conventional 30 min injection-test interval. Although parallel studies indicated that the OX1R antagonist accelerated the onset of resting (and suppressed most active behaviours) even in the absence of food, an equianorectic dose of the natural satiety-related signal cholescystokinin octapeptide (CCK-8S; 5 microg/kg, i.p.) also produced very similar behavioural effects regardless of the presence of food. Together with evidence that SB-334867 preserves the structural integrity of natural feeding behaviour, does not induce nausea/illness or alter taste/palatability and fails to influence EEG measures of arousal/sleep, the present findings are consistent with the view that acute OX1R antagonism selectively enhances satiety. However, unlike the immediate short-circuiting of the satiety sequence induced by CCK-8S, the slower response to SB-334867 implies a more indirect mechanism of action.

Recent studies have shown that acute systemic administration of the selective orexin-1 receptor antagonist SB-334867 significantly reduces food intake in rats. Although this anorectic action of orexin-1 receptor blockade is associated with an acceleration in the transition from eating to resting, it is widely recognised that the behavioural indices of satiety are not dissimilar to those of illness. In this context, Experiment 1 confirmed a significant anorectic effect of 90 (but not 60) mg/kg lithium chloride (LiCl) in male rats presented with palatable mash in the home-cage environment. Experiment 2 employed a continuous monitoring technique to contrast the effects of LiCl (90 mg/kg) and SB-334867 (10 and 30 mg/kg) on food intake and behaviour during a 1-h test with palatable mash. SB-334867 dose-dependently inhibited food intake, with the higher dose producing a comparable degree of appetite suppression (approximately 40%) to that seen with LiCl. Despite equivalent anorectic action, the two compounds produced very different effects on behaviour. LiCl reduced active behaviours (locomotion, rearing, grooming and sniffing), slowed the rate of eating and disrupted the behavioural satiety sequence (BSS). In contrast, SB-334867 (30 mg/kg) decreased the duration of feeding and grooming, and modestly accelerated the transition between eating and resting. Furthermore, whereas LiCl failed to alter posttreatment bodyweight gain, SB-334867 (30 mg/kg) produced a significant weight loss in the 24-h period immediately following injection. Overall, the divergent profiles obtained with equianorectic doses of LiCl and SB-334867 provide convincing evidence for the behavioural selectivity of SB-334867-induced anorexia.

Truncated peptide analogues of orexin-A were prepared and their biological activity assesed at the orexin-1 receptor. Progressive N-terminal deletions identified the minimum C-terminal sequence required for maintaining a significant agonist effect, whilst an alanine scan and other pertinent substitutions identified key side-chain and stereochemical requirements for receptor activation.

Simple and rapid single-pot preparations of substituted 2-mercaptobenzaldehydes 5 and 2-substituted benzo[ b ]thiophenes 3 based on ortho -lithiation methodology are described.

A series of substituted omega-[2-(1H-imidazol-1-yl)ethoxy]alkanoic acid derivatives were synthesized and evaluated for their ability to inhibit thromboxane synthase both in vitro and in vivo. Compound 13 was identified as a potent and selective competitive inhibitor of human platelet thromboxane synthase having a Ki value of 9.6 X 10(-8) M. In collagen-treated human whole blood, 13 potentiated levels of 6-keto PGF1 alpha. Enantiospecific syntheses afforded the R and S enantiomers of 13, of which the S enantiomer 13b was the more potent. Compounds 13 and 13b were potent in vivo inhibitors of thromboxane synthase with good oral activity and duration of action.

The discovery and structure-activity of a new class of renal artery phosphodiesterase inhibitors is reported, some of which are highly selective for the guanosine cyclic 3',5'-monophosphate phosphodiesterase. One of these compounds, 5,6-dihydro-8,9,11,12-tetramethoxy-1,3-dioxo-1H-benz[ f ]-isoquino [8,1,2- hij ]quinazoline-2(3H)-carboxylic acid, ethyl ester ( 9 ), is amongst the most potent and selective compounds of this class yet identified. Furthermore, this compound demonstrates an antihypertensive effect in vivo which is presumably mediated through vascular smooth muscle relaxation.

A synthesis of the macrolactam tumour promoter indolactam V is described in 11 steps from 4-aminoindole.

A number of structurally related 5-substituted pyrimidine 2'-deoxyribonucleosides were synthesized and tested for antiviral activity against herpes simplex virus type 1 (HSV-1) in cell culture. A minimum inhibitory concentration was determined for each compound, and from a comparison of these values a number of conclusions were drawn with regard to those molecular features that enhance or reduce antiviral activity. Optimum inhibition of HSV-1 in cell culture occurred when the 5-substituent was unsaturated and conjugated with the pyrimidine ring, was not longer than four carbon atoms in length, had E stereochemistry, and included a hydrophobic, electronegative function but did not contain a branching point. Such features are contained in (E)-5-(2-bromovinyl)-2'-deoxyuridine, which was the most active of the compounds described.

A number of structurally related 5-substituted pyrimidine 2'-deoxyribonucleosides were synthesized and tested for antiviral activity against herpes simplex virus type 1 (HSV-1) in cell culture. A minimum inhibitory concentration was determined for each compound, and from a comparison of these values a number of conclusions were drawn with regard to those molecular features that enhance or reduce antiviral activity. Optimum inhibition of HSV-1 in cell culture occurred when the 5-substituent was unsaturated and conjugated with the pyrimidine ring, was not longer than four carbon atoms in length, had E stereochemistry, and included a hydrophobic, electronegative function but did not contain a branching point. Such features are contained in (E)-5-(2-bromovinyl)-2'-deoxyuridine, which was the most active of the compounds described.

Aminoindol (I) wird gemas Formelschema in die im Titel genannte Verbindung (Xa) umgewandelt.

4-Alkylaminoindoles rearrange in good yield to the corresponding 1-alkyl-4-aminoindoles in the presence of 10 mol% hydrated toluene-p-sulphonic acid in boiling toluene.

4-Alkylaminoindoles rearrange in good yield to the corresponding 1-alkyl-4-aminoindoles in the presence of 10 mol% hydrated toluene-p-sulphonic acid in boiling toluene.