A novel series of 5-HT(7) receptor ligands has been identified and evaluated, providing compounds showing a broad spectrum of functional activities and good selectivity over selected receptors and ion channels.

The affinity of several antidepressant and antipsychotic drugs for the 5-HT7 receptor and its CNS distribution suggest potential in the treatment of psychiatric diseases. However, there is little direct evidence of receptor function in vivo to support this. We therefore evaluated 5-HT7 receptors as a potential drug target by generating and assessing a 5-HT7 receptor knockout mouse. No difference in assays sensitive to potential psychotic or anxiety states was observed between the 5-HT7 receptor knockout mice and wild type controls. However, in the Porsolt swim test, 5-HT7 receptor knockout mice showed a significant decrease in immobility compared to controls, a phenotype similar to antidepressant treated mice. Intriguingly, treatment of wild types with SB-258719, a selective 5-HT7 receptor antagonist, did not produce a significant decrease in immobility unless animals were tested in the dark (or active) cycle, rather than the light, adding to the body of evidence suggesting a circadian influence on receptor function. Extracellular recordings from hypothalamic slices showed that circadian rhythm phase shifts to 8-OH-DPAT are attenuated in the 5-HT7 receptor KO mice also indicating a role for the receptor in the regulation of circadian rhythms. These pharmacological and genetic knockout studies provide the first direct evidence that 5-HT7 receptor antagonists should be investigated for efficacy in the treatment of depression.

The 5-HT7 receptor is a recent addition to the 5-HT receptor family and to date there is no clear idea as to its potential role in the CNS. The receptor has been mapped by in situ hybridization and 5-HT7-like immunoreactivity and has been detected in discrete areas of the brain including the hypothalamus (Oliver et al., 1999). This suggests the receptor may be involved in temperature regulation and Hagan et al. (2000) have shown that a selective 5-HT7 receptor antagonist reverses the hypothermic effect of 5-CT in guinea-pigs. The current study confirmed that the 5-HT7 receptor antagonists, SB-269970 (1–30 mg/kg, i.p.) and SB-258719 (5–20 mg/kg, i.p.), but not the 5-HT1A receptor antagonist, WAY 100635(0.1–1 mg/kg, s.c.), or the 5-HT1B/D antagonist, GR127935 (1.25–5 mg/kg, i.p.), reversed the hypothermic effect of 5-CT in mice. In addition the effect of 5-CT on body temperature was examined on 5-HT7 receptor null mutant mice. 5-CT (0.1–1 mg/kg, i.p.) significantly reduced rectal temperature in wildtype but not 5-HT7 receptor knockout mice. This suggests that the hypothermic effects of 5-CT are mediated through the 5-HT7 receptor. All procedures were carried out in accordance with the UK Animals (Scientific Procedures) Act (1986).

A bstract : EDG receptors are a family of closely related G‐protein‐coupled receptors, so‐called since the first family member to be cloned is encoded by an e ndothelial d ifferentiation g ene. Of the six family members identified, five use lysophospholipids as their endogenous ligands. The sixth receptor, EDG‐6, remains an orphan. These receptors activate multiple secondary‐messenger pathways involving coupling to Gi, Gq/11, and G12/13 trimeric guanine nucleotide‐binding proteins and are thought to play an important role in cell growth, development and maintenance, and cytoskeletal‐dependent changes. EDG receptors are expressed in most mammalian cells and tissues, each subtype having a distinct distrubution pattern, raising the possibility of tissue‐specific biological roles that could be explored in drug‐discovery programs. In this study the distribution of EDG‐receptor mRNA within the nervous system has been investigated. As seen in peripheral tissues, these receptors appear to be discretely localized within specific brain regions and cell types. For example, EDG‐1, ‐3, ‐4 receptors are confined to neuronal cells, EDG‐2 receptors to white matter tracts, while EDG‐5 receptors appear to be expressed in various cell types, including neuronal cells, white matter tracts, and ependymal cells. EDG‐6‐receptor mRNA was not detected in the nervous system. Speculation as to the role of these receptors in physiological/pathophysiological processes, particularly those involving cell development, proliferation, maintenance, migration, differentiation, plasticity, and apoptosis can be made from such distribution studies. EDG receptors located in brain neuronal cells might, for example, influence apoptosis and be involved in cell rescue following ischemic damage or during the early stages of progressive neurodegenerative diseases. Those restricted to oligodendrocytes might play a crucial role in myelination and offer a potential target in the treatment of demyelinating diseases, such as multiple sclerosis. In order to explore the role of these receptors, it is necessary to identify selective compounds. To this end we have developed an agonist‐induced [ 35 S]GTPγS binding assay using an HEK cell line expressing a pertussis‐toxin‐insensitive human‐EDG‐2‐receptor‐rat‐Giα1‐fusion protein. Such as assay system ovecomes the problems associated with the almost ubiquitous responsiveness of mammalian cells to lysophospholipid. This assay lends itself to high throughput application, opening up the possibility of identifying compounds to further probe the therapeutic potential of EDG receptor manipulation

Recombinant receptor cell lines are widely used in G-protein-coupled receptor selectivity studies. To unequivocally interpret the results of such studies, it is essential that the host cell line does not endogenously express the receptor of interest and in addition is unresponsive to the receptor9s natural ligand. Here we describe an approach to overcome such difficulties associated with orphan receptors or, as in the present case, receptors whose endogenous ligand ubiquitously affects mammalian cells. The functional heterologous assay system described is for the hEdg2 receptor, which uses lysophosphatidic acid as its endogenous ligand. Once activated, this receptor mediates its effects via multiple secondary messenger pathways, including a Gi-coupled pathway. We have transiently expressed a pertussis toxin-insensitive hEdg2 receptor-ratGiα1 fusion protein into human embryonic kidney cells and have monitored the ability of compounds to stimulate [35S]GTPγS binding in membranes prepared from these cells after pretreatment with toxin. Because the assay conditions used favor Gi-mediated responses and because endogenous Giα subunits are rendered inactive, the response measured is, by definition, fusion protein-mediated. Consequently, we have developed an assay that monitors definitively Edg2 receptor-mediated responses in a mammalian cell line. A limited structure activity relationship study suggests that the lysophospholipid carbon chain has a role in receptor activation and in addition indicates that certain modifications to the phosphate group are tolerated.

Several 5-HT(1D/1B) receptor agonists are now entering the marketplace as treatments for migraine. This paper describes the development of selective h5-HT(1D) receptor agonists as potential antimigraine agents which may produce fewer side effects. A series of 3-[3-(piperidin-1-yl)propyl]indoles has been synthesized which has led to the identification of 80 (L-772,405), a high-affinity h5-HT(1D) receptor full agonist having 170-fold selectivity for h5-HT(1D) receptors over h5-HT(1B) receptors. L-772,405 also shows very good selectivity over a range of other serotonin and nonserotonin receptors and has excellent bioavailability following subcutaneous administration in rats. It therefore constitutes a valuable tool to delineate the role of h5-HT(1D) receptors in migraine. Molecular modeling and physical properties have been utilized to postulate the binding conformation of these compounds in the receptor cavity.

The conformational restriction of a (benzylamino)methyl substituted pyrrolidine to form 2,7-diazabicyclo[3.3.0]octanes has led to a series of compounds with high affinity at the h5-HT1D receptor as well as dramatically increased concentrations in the hepatic portal vein following oral administration.

Clinically effective antimigraine drugs such as Sumatriptan have similar affinity at h5-HT1D and h5-HT1B receptors. In the search for a h5-HT1D-selective agonist as an antimigraine agent, a novel series of 3-(propylpiperazinyl)indoles have been synthesized and evaluated at h5-HT1D and h5-HT1B receptors. This class of compounds has provided subnanomolar, fully efficacious h5-HT1D agonists with up to 200-fold selectivity for the h5-HT1D receptor over the h5-HT1B receptor. Unlike other h5-HT1D-selective series, several propylpiperazines demonstrate good oral bioavailability. The optimum compound was 1-(3-[5-(1,2,4-triazol-4-yl)-1H-indol-3-yl]-4-(2-(3-fluorophenyl)ethyl)piperazine (7f) which has excellent selectivity for h5-HT1D receptors over other 5-HT receptor subtypes and good oral bioavailability in three species. Compound 7f has been selected for further investigation as a potential development candidate in the treatment of migraine.

A series of 4-hydroxy-1-[3-(5-(1,2,4-triazol-4-yl)-1H-indol-3-yl)propyl] piperidines was investigated as potential selective h5-HT1D agonists for the treatment of migraine. The 4-[(N-benzyl-N-methyl)amino]methyl analog 12a was found to be a full agonist at the h5-HT1D receptor with good binding selectivity over the h5-HT1B receptor.

The design, synthesis, and biological evaluation of a novel series of 3-[2-(pyrrolidin-1-yl)ethyl]indoles with excellent selectivity for h5-HT1D (formerly 5-HT1Dalpha) receptors over h5-HT1B (formerly 5-HT1Dbeta) receptors are described. Clinically effective antimigraine drugs such as Sumatriptan show little selectivity between h5-HT1D and h5-HT1B receptors. The differential expression of h5-HT1D and h5-HT1B receptors in neural and vascular tissue prompted an investigation of whether a compound selective for the h5-HT1D subtype would have the same clinical efficacy but with reduced side effects. The pyrrolidine 3b was initially identified as having 9-fold selectivity for h5-HT1D over h5-HT1B receptors. Substitution of the pyrrolidine ring of 3b with methylbenzylamine groups gave compounds with nanomolar affinity for the h5-HT1D receptor and 100-fold selectivity with respect to h5-HT1B receptors. Modification of the indole 5-substituent led to the oxazolidinones 24a,b with up to 163-fold selectivity for the h5-HT1D subtype and improved selectivity over other serotonin receptors. The compounds were shown to be full agonists by measurement of agonist-induced [35S]GTPgammaS binding in CHO cells expressed with h5-HT receptors. This study suggests that the h5-HT1D and h5-HT1B receptors can be differentiated by appropriate substitution of the ligand in the region which binds to the aspartate residue and reveals a large binding pocket in the h5-HT1D receptor domain which is absent for the h5-HT1B receptor. The compounds described herein will be important tools to delineate the role of h5-HT1D receptors in migraine.

It has previously been reported that a 3-(3-(piperazin-1-yl)propyl)indole series of 5-HT1D receptor ligands have pharmacokinetic advantages over the corresponding 3-(3-(piperidin-1-yl)propyl)indole series and that the reduced pKa of the piperazines compared to the piperidines may be one possible explanation for these differences. To investigate this proposal we have developed versatile synthetic strategies for the incorporation of fluorine into these ligands, producing novel series of 4-fluoropiperidines, 3-fluoro-4-aminopiperidines, and both piperazine and piperidine derivatives with one or two fluorines in the propyl linker. Ligands were identified which maintained high affinity and selectivity for the 5-HT1D receptor and showed agonist efficacy in vitro. The incorporation of fluorine was found to significantly reduce the pKa of the compounds, and this reduction of basicity was shown to have a dramatic, beneficial influence on oral absorption, although the effect on oral bioavailability could not always be accurately predicted.

In this study, the binding of [3H]5-HT to the cloned dog 5-hydroxytryptamine1B (dog 5-HT1B) receptor, stably expressed in Chinese hamster ovary cells (ATCC CCL 61)(CHO-K1), was characterised and its pharmacology compared with that of the cloned human and rat 5-HT1B receptors. [3H]5-HT specifically labeled, with high affinity, an apparently homogeneous population of binding sites in the dog 5-HT1B receptor cell line yielding a pKd of 8.1. [3H]5-HT inhibition and agonist-induced [35S] guanosine 5'[gamma-thio] triphosphate ([35S]GTPgammaS) binding studies revealed comparable results with the human but not the rat 5-HT1B receptor. In all three recombinant receptor cell lines, methiothepin displayed inverse agonism and GR127935 (N-[4-methoxy-3-(4-methyl-1-piperizinyl)phenyl]-2'-methyl-4'-(5-me thyl-1,2,4-oxadiazole-3-yl)[1,1'-biphenyl]-carboxamide) weak partial agonism.

Annals of the New York Academy of SciencesVolume 861, Issue 1 p. 265-265 In Vitro Visualization of 5-HT Receptors in Rat Brain Using [35S]GTPγS Binding M. S. BEER, Corresponding Author M. S. BEER Merck, Sharp & Dohme Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, Essex, CM20 2QR, UK Corresponding author: Tel: 44.1279.44.04.43; Fax: 44.1279.44.03.90; http://[email protected]Search for more papers by this authorJ. A. STANTON, J. A. STANTON Merck, Sharp & Dohme Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, Essex, CM20 2QR, UKSearch for more papers by this author M. S. BEER, Corresponding Author M. S. BEER Merck, Sharp & Dohme Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, Essex, CM20 2QR, UK Corresponding author: Tel: 44.1279.44.04.43; Fax: 44.1279.44.03.90; http://[email protected]Search for more papers by this authorJ. A. STANTON, J. A. STANTON Merck, Sharp & Dohme Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, Essex, CM20 2QR, UKSearch for more papers by this author First published: 07 February 2006 https://doi.org/10.1111/j.1749-6632.1998.tb10215.xRead the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat No abstract is available for this article. REFERENCES 1 Sim L. J., D. E. Selley, & S. R. Childers. 1995. Proc. Natl. Acad. Sci. USA 92: 7242–7246. Volume861, Issue1ADVANCES IN SEROTONIN RECEPTOR RESEARCH: MOLECULAR BIOLOGY, SIGNAL TRANSDUCTION, AND THERAPEUTICSa December 1998Pages 265-265 ReferencesRelatedInformation

ADVERTISEMENT RETURN TO ISSUEPREVLetterNEXTEnhancement of Oral Absorption in Selective 5-HT1D Receptor Agonists: Fluorinated 3-[3-(Piperidin-1-yl)propyl]indolesJosé L. Castro, Ian Collins, Michael G. N. Russell, Alan P. Watt, Bindi Sohal, Denise Rathbone, Margaret S. Beer, and Josephine A. StantonView Author Information Chemistry and Biochemistry Departments, Merck Sharp and Dohme Research Laboratories, Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, Essex CM20 2QR, U.K. Cite this: J. Med. Chem. 1998, 41, 15, 2667–2670Publication Date (Web):June 30, 1998Publication History Received1 April 1998Published online30 June 1998Published inissue 1 July 1998https://pubs.acs.org/doi/10.1021/jm980204ehttps://doi.org/10.1021/jm980204erapid-communicationACS PublicationsCopyright © 1998 American Chemical SocietyRequest reuse permissionsArticle Views1204Altmetric-Citations28LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-AlertscloseSupporting Info (1)»Supporting Information Supporting Information SUBJECTS:Absorption,Agonists,Piperidines,Receptors,Rodent models Get e-Alerts