Publications by authors named "Martin J Gunthorpe"

23 Publications

  • Page 1 of 1

A novel series of benzimidazole NR2B-selective NMDA receptor antagonists.

Bioorg Med Chem Lett 2012 Apr 6;22(7):2620-3. Epub 2012 Feb 6.

GlaxoSmithKline R&D, Medicines Research Centre, Gunnels Wood Road, Stevenage, Hertfordshire SG1 2NY, UK.

A series of novel benzimidazoles are discussed as NR2B-selective N-methyl-d-aspartate (NMDA) receptor antagonists. High throughput screening (HTS) efforts identified a number of potent and selective NR2B antagonists such as 1. Exploration of the substituents around the core of this template identified a number of compounds with high potency for NR2B (pIC(50) >7) and good selectivity against the NR2A subunit (pIC(50) <4.3) as defined by FLIPR-Ca(2+) and radioligand binding studies. These agents offer potential for the development of therapeutics for a range of nervous system disorders including chronic pain, neurodegeneration, migraine and major depression.
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http://dx.doi.org/10.1016/j.bmcl.2012.01.108DOI Listing
April 2012

The spectrum of anticonvulsant efficacy of retigabine (ezogabine) in animal models: implications for clinical use.

Epilepsia 2012 Mar 5;53(3):425-36. Epub 2012 Jan 5.

Medicines Research Centre, GlaxoSmithKline S.p.A., Verona, Italy.

Retigabine [RTG (international nonproprietary name); ezogabine (EZG; U.S. adopted name)] is a first-in-class antiepileptic drug (AED) that reduces neuronal excitability by enhancing the activity of KCNQ (K(v)7) potassium (K(+)) channels. RTG/EZG has recently been approved by the European Medicines Agency and the U.S. Food and Drug Administration as adjunctive therapy in adults with partial-onset seizures. In this review we discuss the activity that RTG/EZG has demonstrated across a broad spectrum of in vitro/in vivo animal models of seizures, including generalized tonic-clonic, primary generalized (absence), and partial seizures, in addition to the compound's ability to resist and block the occurrence of seizures induced by a range of stimuli across different regions of the brain. The potency of RTG/EZG in models refractory to several conventional AEDs and the work done to assess antiepileptogenesis and neuroprotection are discussed. Studies that have evaluated the central nervous system side effects of RTG/EZG in animals are reviewed in order to compare these effects with adverse events observed in patients with epilepsy. Based on its demonstrated effect in a number of animal epilepsy models, the synergistic and additive activity of RTG/EZG with other AEDs supports its potential use in therapeutic combinations for different seizure types. The distinct mechanism of action of RTG/EZG from those of currently available AEDs, along with its broad preclinical activity, underscores the key role of KCNQ (K(v)7) K(+) channels in neuronal excitability, and further supports the potential efficacy of this unique molecule in the treatment of epilepsy.
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http://dx.doi.org/10.1111/j.1528-1167.2011.03364.xDOI Listing
March 2012

The mechanism of action of retigabine (ezogabine), a first-in-class K+ channel opener for the treatment of epilepsy.

Epilepsia 2012 Mar 5;53(3):412-24. Epub 2012 Jan 5.

New Frontiers Science Park, GlaxoSmithKline plc, Harlow, Essex, United Kingdom.

The pharmacologic profile of retigabine [RTG (international nonproprietary name); ezogabine, EZG (U.S. adopted name)], is different from all currently approved antiepileptic drugs (AEDs). Its primary mechanism of action (MoA) as a positive allosteric modulator of KCNQ2-5 (K(v) 7.2-7.5) ion channels defines RTG/EZG as the first neuronal potassium (K(+)) channel opener for the treatment of epilepsy. KCNQ2-5 channels are predominantly expressed in neurons and are important determinants of cellular excitability, as indicated by the occurrence of human genetic mutations in KCNQ channels that underlie inheritable disorders including, in the case of KCNQ2/3, the syndrome of benign familial neonatal convulsions. In vitro pharmacologic studies demonstrate that the most potent action of RTG/EZG is at KCNQ2-5 channels, particularly heteromeric KCNQ2/3. Furthermore, mutagenesis and modeling studies have pinpointed the RTG/EZG binding site to a hydrophobic pocket near the channel gate, indicating how RTG/EZG can stabilize the open form of KCNQ2-5 channels; the absence of this site in KCNQ1 also provides a clear explanation for the inbuilt selectivity RTG/EZG has for potassium channels other than the KCNQ cardiac channel. KCNQ channels are active at the normal cell resting membrane potential (RMP) and contribute a continual hyperpolarizing influence that stabilizes cellular excitability. The MoA of RTG/EZG increases the number of KCNQ channels that are open at rest and also primes the cell to retort with a larger, more rapid, and more prolonged response to membrane depolarization or increased neuronal excitability. In this way, RTG/EZG amplifies this natural inhibitory force in the brain, acting like a brake to prevent the high levels of neuronal action potential burst firing (epileptiform activity) that may accompany sustained depolarizations associated with the initiation and propagation of seizures. This action to restore physiologic levels of neuronal activity is thought to underlie the efficacy of RTG/EZG as an anticonvulsant in a broad spectrum of preclinical seizure models and in placebo-controlled trials in patients with partial epilepsy. In this article, we consider the pharmacologic characteristics of RTG/EZG at the receptor, cellular, and network levels as a means of understanding the novel and efficacious MoA of this new AED as defined in both preclinical and clinical research.
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http://dx.doi.org/10.1111/j.1528-1167.2011.03365.xDOI Listing
March 2012

Ototrauma induces sodium channel plasticity in auditory afferent neurons.

Mol Cell Neurosci 2011 Sep 25;48(1):51-61. Epub 2011 Jun 25.

Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, LE1 9HN UK.

Exposure to intense sound can cause damage to the delicate sensory and neuronal components of the cochlea leading to hearing loss. Such damage often causes the dendrites of the spiral ganglion neurons (SGN), the neurons that provide the afferent innervation of the hair cells, to swell and degenerate thus damaging the synapse. In models of neuropathic pain, axotomy, another form of afferent nerve damage, is accompanied by altered voltage-gated sodium channel (VGSC) expression, leading to neuronal hyperactivity. In this study, adult Wistar rats were exposed to noise which produced a mild, 20 dB hearing threshold elevation and their VGSC expression was investigated. Quantitative PCR showed decreased Na(V)1.1 and Na(V)1.6 mRNA expression in the SGN following noise exposure (29% and 56% decrease respectively) while Na(V)1.7 mRNA expression increased by approximately 20% when compared to control rats. Immunohistochemistry extended these findings, revealing increased staining for Na(V)1.1 along the SGN dendrites and Na(V)1.7 in the cell bodies after noise. These results provide the first evidence for selective changes in VGSC expression following moderate noise-induced hearing loss and could contribute to elevated hearing thresholds and to the generation of perceptual anomalies commonly associated with cochlear damage, such as tinnitus and hyperacusis.
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http://dx.doi.org/10.1016/j.mcn.2011.06.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3176910PMC
September 2011

The efficacy of sodium channel blockers to prevent phencyclidine-induced cognitive dysfunction in the rat: potential for novel treatments for schizophrenia.

J Pharmacol Exp Ther 2011 Jul 12;338(1):100-13. Epub 2011 Apr 12.

Neuroscience Centre of Excellence for Drug Discovery, Medicines Research Centre, GlaxoSmithKline SpA., Verona, Italy.

Sodium channel inhibition is a well precedented mechanism used to treat epilepsy and other hyperexcitability disorders. The established sodium channel blocker and broad-spectrum anticonvulsant lamotrigine is also effective in the treatment of bipolar disorder and has been evaluated in patients with schizophrenia. Double-blind placebo-controlled clinical trials found that the drug has potential to reduce cognitive symptoms of the disorder. However, because of compound-related side-effects and the need for dose titration, a conclusive evaluation of the drug's efficacy in patients with schizophrenia has not been possible. (5R)-5-(4-{[(2-Fluorophenyl)methyl]oxy}phenyl)-l-prolinamide (GSK2) and (2R,5R)-2-(4-{[(2-fluorophenyl)methyl]oxy}phenyl)-7-methyl-1,7-diazaspiro[4.4]nonan-6-one (GSK3) are two new structurally diverse sodium channel blockers with potent anticonvulsant activity. In this series of studies in the rat, we compared the efficacy of the two new molecules to prevent a cognitive deficit induced by the N-methyl-d-aspartic acid receptor antagonist phencyclidine (PCP) in the reversal-learning paradigm in the rat. We also explored the effects of the drugs to prevent brain activation and neurochemical effects of PCP. We found that, like lamotrigine, both GSK2 and GSK3 were able to prevent the deficit in reversal learning produced by PCP, thus confirming their potential in the treatment of cognitive symptoms of schizophrenia. However, higher doses than those required for anticonvulsant efficacy of the drugs were needed for activity in the reversal-learning model, suggesting a lower therapeutic window relative to mechanism-dependent central side effects for this indication.
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http://dx.doi.org/10.1124/jpet.110.178475DOI Listing
July 2011

Targeting gut T cell Ca2+ release-activated Ca2+ channels inhibits T cell cytokine production and T-box transcription factor T-bet in inflammatory bowel disease.

J Immunol 2009 Sep 31;183(5):3454-62. Epub 2009 Jul 31.

Centre for Infectious Disease, Institute of Cell and Molecular Science, Barts and the London School of Medicine and Dentistry, London, UK.

Prolonged Ca(2+) entry through Ca(2+) release-activated Ca(2+) (CRAC) channels is crucial in activating the Ca(2+)-sensitive transcription factor NFAT, which is responsible for directing T cell proliferation and cytokine gene expression. To establish whether targeting CRAC might counteract intestinal inflammation, we evaluated the in vitro effect of a selective CRAC inhibitor on T cell cytokine production and T-bet expression by lamina propria mononuclear cells (LPMC) and biopsy specimens from inflammatory bowel disease (IBD) patients. The inhibitory activity of the CRAC blocker was investigated through patch-clamp experiments on rat basophilic leukemia cells and fluorometric imaging plate reader intracellular Ca(2+) assays using thapsigargin-stimulated Jurkat T cells and its detailed selectivity profile defined using a range of in vitro radioligand binding and functional assays. Anti-CD3/CD28-stimulated LPMC and biopsy specimens from 51 patients with IBD were cultured with a range of CRAC inhibitor concentrations (0.01-10 microM). IFN-gamma, IL-2, IL-8, and IL-17 were analyzed by ELISA. T-bet was determined by immunoblotting. We found that the CRAC blocker concentration-dependently inhibited CRAC current in rat basophilic leukemia cells and thapsigargin-induced Ca(2+) influx in Jurkat T cells. A concentration-dependent reduction in T-bet expression and production of IFN-gamma, IL-2, IL-17, but not IL-8, was observed in IBD LPMC and biopsy specimens treated with the CRAC inhibitor. In conclusion, we provide evidence that the suppression of CRAC channel function may dampen the increased T cell response in the inflamed gut, thus suggesting a promising role for CRAC inhibitor drugs in the therapeutic management of patients with IBD.
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http://dx.doi.org/10.4049/jimmunol.0802887DOI Listing
September 2009

Allosteric modulators of NR2B-containing NMDA receptors: molecular mechanisms and therapeutic potential.

Br J Pharmacol 2009 Aug 8;157(8):1301-17. Epub 2009 Jul 8.

Laboratoire de Neurobiologie, Ecole Normale Supérieure, CNRS, Paris, France.

N-methyl-D-aspartate receptors (NMDARs) are ion channels gated by glutamate, the major excitatory neurotransmitter in the mammalian central nervous system (CNS). They are widespread in the CNS and are involved in numerous physiological and pathological processes including synaptic plasticity, chronic pain and psychosis. Aberrant NMDAR activity also plays an important role in the neuronal loss associated with ischaemic insults and major degenerative disorders including Parkinson's and Alzheimer's disease. Agents that target and alter NMDAR function may, thus, have therapeutic benefit. Interestingly, NMDARs are endowed with multiple extracellular regulatory sites that recognize ions or small molecule ligands, some of which are likely to regulate receptor function in vivo. These allosteric sites, which differ from agonist-binding and channel-permeation sites, provide means to modulate, either positively or negatively, NMDAR activity. The present review focuses on allosteric modulation of NMDARs containing the NR2B subunit. Indeed, the NR2B subunit confers a particularly rich pharmacology with distinct recognition sites for exogenous and endogenous allosteric ligands. Moreover, NR2B-containing receptors, compared with other NMDAR subtypes, appear to contribute preferentially to pathological processes linked to overexcitation of glutamatergic pathways. The actions of extracellular H+, Mg2+, Zn2+, of polyamines and neurosteroids, and of the synthetic compounds ifenprodil and derivatives ('prodils') are presented. Particular emphasis is put upon the structural determinants and molecular mechanisms that underlie the effects exerted by these agents. A better understanding of how NR2B-containing NMDARs (and NMDARs in general) operate and how they can be modulated should help define new strategies to counteract the deleterious effects of dysregulated NMDAR activity.
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http://dx.doi.org/10.1111/j.1476-5381.2009.00304.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2765303PMC
August 2009

Clinical development of TRPV1 antagonists: targeting a pivotal point in the pain pathway.

Drug Discov Today 2009 Jan 26;14(1-2):56-67. Epub 2008 Dec 26.

Neurology Centre of Excellence for Drug Discovery, GlaxoSmithKline, New Frontiers Science Park (North), Third Avenue, Harlow, Essex CM19 5AW, UK.

TRPV1 is a noxious heat, capsaicin (vanilloid) and acid receptor for which the development of antagonists represents a novel therapeutic approach for the treatment of pain. TRPV1 antagonists have entered early clinical development and initial reports indicate that they have demonstrated pharmacodynamic effects consistent with TRPV1 antagonist activity and anti-hyperalgesic action in humans. Should these effects extend to the relief of symptoms experienced by patients with chronic pain then this class of compounds may offer one of the first novel mechanisms of action for the treatment for pain for many years. In this article we will discuss recent progress and challenges in the field in this highly competitive area of drug discovery.
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http://dx.doi.org/10.1016/j.drudis.2008.11.005DOI Listing
January 2009

Characterisation of 5-HT3C, 5-HT3D and 5-HT3E receptor subunits: evolution, distribution and function.

J Neurochem 2009 Jan 29;108(2):384-96. Epub 2008 Nov 29.

Molecular Discovery Research, GlaxoSmithKline, Harlow, Essex, UK.

The 5-HT(3) receptor is a member of the 'Cys-loop' family of ligand-gated ion channels that mediate fast excitatory and inhibitory transmission in the nervous system. Current evidence points towards native 5-HT(3) receptors originating from homomeric assemblies of 5-HT(3A) or heteromeric assembly of 5-HT(3A) and 5-HT(3B). Novel genes encoding 5-HT(3C), 5-HT(3D), and 5-HT(3E) have recently been described but the functional importance of these proteins is unknown. In the present study, in silico analysis (confirmed by partial cloning) indicated that 5-HT(3C), 5-HT(3D), and 5-HT(3E) are not human-specific as previously reported: they are conserved in multiple mammalian species but are absent in rodents. Expression profiles of the novel human genes indicated high levels in the gastrointestinal tract but also in the brain, Dorsal Root Ganglion (DRG) and other tissues. Following the demonstration that these subunits are expressed at the cell membrane, the functional properties of the recombinant human subunits were investigated using patch clamp electrophysiology. 5-HT(3C), 5-HT(3D), and 5-HT(3E) were all non-functional when expressed alone. Co-transfection studies to determine potential novel heteromeric receptor interactions with 5-HT(3A) demonstrated that the expression or function of the receptor was modified by 5-HT(3C) and 5-HT(3E), but not 5-HT(3D). The lack of distinct effects on current rectification, kinetics or pharmacology of 5-HT(3A) receptors does not however provide unequivocal evidence to support a direct contribution of 5-HT(3C) or 5-HT(3E) to the lining of the ion channel pore of novel heteromeric receptors. The functional and pharmacological contributions of these novel subunits to human biology and diseases such as irritable bowel syndrome for which 5-HT(3) receptor antagonists have major clinical usage, therefore remains to be fully determined.
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http://dx.doi.org/10.1111/j.1471-4159.2008.05775.xDOI Listing
January 2009

Design and synthesis of 6-phenylnicotinamide derivatives as antagonists of TRPV1.

Bioorg Med Chem Lett 2008 Oct 31;18(20):5609-13. Epub 2008 Aug 31.

Neurology CEDD, GlaxoSmithKline, New Frontiers Science Park, Third Avenue, Harlow, Essex CM19 5AW, UK.

6-Phenylnicotinamide (2) was previously identified as a potent TRPV1 antagonist with activity in an in vivo model of inflammatory pain. Optimization of this lead through modification of both the biaryl and heteroaryl components has resulted in the discovery of 6-(4-fluorophenyl)-2-methyl-N-(2-methylbenzothiazol-5-yl)nicotinamide (32; SB-782443) which possesses an excellent overall profile and has been progressed into pre-clinical development.
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http://dx.doi.org/10.1016/j.bmcl.2008.08.105DOI Listing
October 2008

Peripheral TRPV1 receptors as targets for drug development: new molecules and mechanisms.

Curr Pharm Des 2008 ;14(1):32-41

Neurology Centre of Excellence for Drug Discovery, GlaxoSmithKline, Harlow, Essex, CM19 5AW, UK.

Based on the painful effects of exposure to capsaicin, TRPV1 (transient receptor potential vanilloid subfamily member 1) localization is most readily associated with peripheral sensory neurons, however, TRPV1 is now known to be expressed, albeit at lower levels, in the spinal cord, brain and a wide-range of non-neuronal cells. The latter includes epithelial cells (e.g. keratinocytes, urothelium, gastric epithelial cells, enterocytes, and pneumocytes) through vascular endothelium and cells of the immune system (e.g. T-cells and mast cells) to smooth muscle, fibroblasts and hepatocytes. Despite extensive research, the physiological function of TRPV1 in the brain and in non-neuronal tissues remains elusive. The preliminary results are exciting, but many are unconfirmed and/or contradictory. As yet, studies with TRPV1 knock-out mice have proven unhelpful in clarifying such biological roles. Now that a range of potent and selective TRPV1 antagonists are available in this rapidly expanding research field, further understanding of the biological roles of TRPV1 throughout the body is within reach. In this article, we will summarize the known roles of peripheral TRPV1 receptors in physiology and disease and review the current perspectives for the therapeutic potential of TRPV1 agonists and antagonists in the treatment of a wide range of conditions such as pain, cancer, migraine, chronic cough, asthma, rectal hypersensitivity, inflammatory bowel disease, obesity, overactive bladder and diabetes. New applications of targeting central TRPV1 receptors are reviewed in the accompanying article by Starowicz et al. (in this issue).
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http://dx.doi.org/10.2174/138161208783330754DOI Listing
February 2008

Characterization of SB-705498, a potent and selective vanilloid receptor-1 (VR1/TRPV1) antagonist that inhibits the capsaicin-, acid-, and heat-mediated activation of the receptor.

J Pharmacol Exp Ther 2007 Jun 28;321(3):1183-92. Epub 2007 Mar 28.

Neurology and Gastrointestinal Centre of Excellence for Drug Discovery, GlaxoSmithKline, Harlow, Essex, UK.

Vanilloid receptor-1 (TRPV1) is a nonselective cation channel, predominantly expressed by sensory neurons, which plays a key role in the detection of noxious painful stimuli such as capsaicin, acid, and heat. TRPV1 antagonists may represent novel therapeutic agents for the treatment of a range of conditions including chronic pain, migraine, and gastrointestinal disorders. Here we describe the in vitro pharmacology of N-(2-bromophenyl)-N'-[((R)-1-(5-trifluoromethyl-2-pyridyl)pyrrolidin-3-yl)]urea (SB-705498), a novel TRPV1 antagonist identified by lead optimization of N-(2-bromophenyl)-N'-[2-[ethyl(3-methylphenyl)amino]ethyl]urea (SB-452533), which has now entered clinical trials. Using a Ca(2+)-based fluorometric imaging plate reader (FLIPR) assay, SB-705498 was shown to be a potent competitive antagonist of the capsaicin-mediated activation of the human TRPV1 receptor (pK(i) = 7.6) with activity at rat (pK(i) = 7.5) and guinea pig (pK(i) = 7.3) orthologs. Whole-cell patch-clamp electrophysiology was used to confirm and extend these findings, demonstrating that SB-705498 can potently inhibit the multiple modes of receptor activation that may be relevant to the pathophysiological role of TRPV1 in vivo: SB-705498 caused rapid and reversible inhibition of the capsaicin (IC(50) = 3 nM)-, acid (pH 5.3)-, or heat (50 degrees C; IC(50) = 6 nM)-mediated activation of human TRPV1 (at -70 mV). Interestingly, SB-705498 also showed a degree of voltage dependence, suggesting an effective enhancement of antagonist action at negative potentials such as those that might be encountered in neurons in vivo. The selectivity of SB-705498 was defined by broad receptor profiling and other cellular assays in which it showed little or no activity versus a wide range of ion channels, receptors, and enzymes. SB-705498 therefore represents a potent and selective multimodal TRPV1 antagonist, a pharmacological profile that has contributed to its definition as a suitable drug candidate for clinical development.
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http://dx.doi.org/10.1124/jpet.106.116657DOI Listing
June 2007

Effects of pan- and subtype-selective N-methyl-D-aspartate receptor antagonists on cortical spreading depression in the rat: therapeutic potential for migraine.

J Pharmacol Exp Ther 2007 May 31;321(2):564-72. Epub 2007 Jan 31.

MRI Group, Neurology and GI Centre of Excellence in Drug Discovery, GlaxoSmithKline, New Frontiers Science Park (North), Third Ave., Harlow, Essex CM19 5AW, UK.

Spreading depression (SD) has long been associated with the underlying pathophysiology of migraine. Evidence that the N-methyl-D-aspartate (NMDA) glutamate receptor (NMDA-R) is implicated in the generation and propagation of SD has itself been available for more than 15 years. However, to date, there are no reports of NMDA-R antagonists being developed for migraine therapy. In this study, an uncompetitive, pan-NMDA-R blocker, memantine, approved for clinical use, and two antagonists with selectivity for NMDA-R containing the NR2B subunit, (1S,2S)-1-(4-hydroxyphenyl)-2-(4-hydroxy-4-phenylpiperidino)-1-propanol (CP-101,606) and (+/-)-(R*,S*)-alpha-(4-hydroxyphenyl)-beta-methyl-4-(phenylmethyl)-1-piperidine propanol (Ro 25-6981), were investigated to assess their protective effects against SD in the rat. Under isoflurane anesthesia, d.c. potential and the related cortical blood flow and partial pressure of O2 (pO2) were recorded simultaneously at separate cortical sites. Drugs (1, 3, and 10 mg/kg i.p.) were given 1 h or 30 min before KCl application to the brain surface. Core temperature and arterial pCO2,pO2, and pH measurements confirmed physiological stability. KCl induced 7.7+/-1.8 (mean+/-S.D.) SD events with d.c. amplitude of 14.9+/-2.8 mV. Memantine and CP-101,606 dose-dependently decreased SD event number (to 2.0+/-1.8 and 2.3+/-2.9, respectively) and SD amplitude at doses relevant for therapeutic use. Ro 25-6981 also decreased SD events significantly, but less effectively (to 4.5+/-1.6), without affecting amplitude. These results indicate that NR2B-containing NMDA receptors are key mediators of SD, and as such, memantine- and NR2B-selective antagonists may be useful new therapeutic agents for the treatment of migraine and other SD-related disorders (e.g., stroke and brain injury). Whether chronic, rather than acute, treatment may improve their efficacy remains to be determined.
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http://dx.doi.org/10.1124/jpet.106.117101DOI Listing
May 2007

The voltage-gated sodium channel Na(v)1.9 is an effector of peripheral inflammatory pain hypersensitivity.

J Neurosci 2006 Dec;26(50):12852-60

Neural Plasticity Research Group, Department of Anesthesia and Critical Care, Massachusetts General Hospital and Harvard Medical School, Charlestown, Massachusetts 02129, USA.

We used a mouse with deletion of exons 4, 5, and 6 of the SCN11A (sodium channel, voltage-gated, type XI, alpha) gene that encodes the voltage-gated sodium channel Na(v)1.9 to assess its contribution to pain. Na(v)1.9 is present in nociceptor sensory neurons that express TRPV1, bradykinin B2, and purinergic P2X3 receptors. In Na(v)1.9-/- mice, the non-inactivating persistent tetrodotoxin-resistant sodium TTXr-Per current is absent, whereas TTXr-Slow is unchanged. TTXs currents are unaffected by the mutation of Na(v)1.9. Pain hypersensitivity elicited by intraplantar administration of prostaglandin E2, bradykinin, interleukin-1beta, capsaicin, and P2X3 and P2Y receptor agonists, but not NGF, is either reduced or absent in Na(v)1.9-/- mice, whereas basal thermal and mechanical pain sensitivity is unchanged. Thermal, but not mechanical, hypersensitivity produced by peripheral inflammation (intraplanatar complete Freund's adjuvant) is substantially diminished in the null allele mutant mice, whereas hypersensitivity in two neuropathic pain models is unchanged in the Na(v)1.9-/- mice. Na(v)1.9 is, we conclude, an effector of the hypersensitivity produced by multiple inflammatory mediators on nociceptor peripheral terminals and therefore plays a key role in mediating peripheral sensitization.
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http://dx.doi.org/10.1523/JNEUROSCI.4015-06.2006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6674969PMC
December 2006

TRPV1 antagonist, SB-366791, inhibits glutamatergic synaptic transmission in rat spinal dorsal horn following peripheral inflammation.

Eur J Pharmacol 2006 Jul 6;540(1-3):73-81. Epub 2006 May 6.

Neurology and GI CEDD, GlaxoSmithKline, New Frontiers Science Park North, Third Avenue, Harlow, Essex, CM19 5AW, UK.

The anti-hyperalgesic effects of TRPV1 receptor antagonists are well documented in animal models of pain, however, the precise site of their action is not known. Here we have examined the effects of the selective TRPV1 antagonist SB-366791 on glutamatergic synaptic transmission in substantia gelatinosa using spinal cord slices from either control rats or animals that had undergone a peripheral inflammation induced by intraplantar injection of Freund's complete adjuvant (FCA). In control animals, SB-366791 (30 microM) had no effect on spontaneous excitatory post-synaptic currents (sEPSC) or evoked EPSCs. In slices from FCA-inflamed animals, SB-366791 decreased sEPSC frequency to 66+/-8% of control in 5/10 neurones, and decreased miniature glutamatergic EPSCs (mEPSC) frequency to 63+/-4% of control, in 6/7 neurones; with no significant effect on sEPSC or mEPSC amplitude. Dorsal root evoked EPSCs at C-fibre intensity were reduced to 72+/-6% of control by SB-366791 (30 microM) in 3/4 neurones from FCA-treated animals. In conclusion, SB-366791 inhibited glutamatergic transmission in a subset of neurones via a pre-synaptic mechanism following peripheral inflammation. We hypothesise that during peripheral inflammation spinal TRPV1 becomes tonically active, promoting the synaptic release of glutamate. These results provide evidence for a mechanism by which TRPV1 contributes to inflammatory pain and provides a basis for the understanding of the efficacy of TRPV1 antagonists.
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http://dx.doi.org/10.1016/j.ejphar.2006.04.046DOI Listing
July 2006

Discovery of SB-705498: a potent, selective and orally bioavailable TRPV1 antagonist suitable for clinical development.

Bioorg Med Chem Lett 2006 Jun 31;16(12):3287-91. Epub 2006 Mar 31.

Neurology and GI CEDD, New Frontiers Science Park, GlaxoSmithKline, Third Avenue, Harlow, Essex CM19 5AW, UK.

Small molecule antagonists of the vanilloid receptor TRPV1 (also known as VR1) are disclosed. Pyrrolidinyl ureas such as 8 and 15 (SB-705498) emerged as lead compounds following optimisation of the previously described urea SB-452533. Pharmacological studies using electrophysiological and FLIPR-Ca2+-based assays showed that compounds such as 8 and 15 were potent antagonists versus the multiple chemical and physical modes of TRPV1 activation (namely capsaicin, acid and noxious heat). Furthermore, 15 possessed suitable developability properties to enable progression of this compound into in vivo studies and subsequently clinical development.
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http://dx.doi.org/10.1016/j.bmcl.2006.03.030DOI Listing
June 2006

An animal model of chronic inflammatory pain: pharmacological and temporal differentiation from acute models.

Eur J Pain 2006 Aug 30;10(6):537-49. Epub 2005 Sep 30.

Department of Pain Research, Neurology and Gastrointestinal CEDD, GlaxoSmithKline Research and Development Ltd., Harlow, Essex, UK.

Clinically, inflammatory pain is far more persistent than that typically modelled pre-clinically, with the majority of animal models focussing on short-term effects of the inflammatory pain response. The large attrition rate of compounds in the clinic which show pre-clinical efficacy suggests the need for novel models of, or approaches to, chronic inflammatory pain if novel mechanisms are to make it to the market. A model in which a more chronic inflammatory hypersensitivity phenotype is profiled may allow for a more clinically predictive tool. The aims of these studies were to characterise and validate a chronic model of inflammatory pain. We have shown that injection of a large volume of adjuvant to the intra-articular space of the rat knee results in a prolonged inflammatory pain response, compared to the response in an acute adjuvant model. Additionally, this model also results in a hypersensitive state in the presence and absence of inflammation. A range of clinically effective analgesics demonstrate activity in this chronic model, including morphine (3mg/kg, t.i.d.), dexamethasone (1mg/kg, b.i.d.), ibuprofen (30mg/kg, t.i.d.), etoricoxib (5mg/kg, b.i.d.) and rofecoxib (0.3-10mg/kg, b.i.d.). A further aim was to exemplify the utility of this chronic model over the more acute intra-plantar adjuvant model using two novel therapeutic approaches; NR2B selective NMDA receptor antagonism and iNOS inhibition. Our data shows that different effects were observed with these therapies when comparing the acute model with the model of chronic inflammatory joint pain. These data suggest that the chronic model may be more relevant to identifying mechanisms for the treatment of chronic inflammatory pain states in the clinic.
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http://dx.doi.org/10.1016/j.ejpain.2005.08.003DOI Listing
August 2006

Conservation of functional and pharmacological properties in the distantly related temperature sensors TRVP1 and TRPM8.

Mol Pharmacol 2005 Aug 23;68(2):518-27. Epub 2005 May 23.

Neurology and GI-CEDD, GlaxoSmithKline, Harlow, Essex, CM19 5AW, UK.

Members of the transient receptor potential (TRP) superfamily of ion channels have now been defined as molecular transducers capable of reproducing the spectrum of temperature sensation exhibited by mammals. Because of their pivotal role in sensory transduction, many of these channels represent good targets for drug discovery. With a view to gaining further insight into the functional and pharmacological properties of these channels, we have used the whole-cell patch-clamp technique to study the human cold-sensitive menthol receptor transient receptor potential melastatin 8 (TRPM8) and compared its behavior with that of its distant relative, the heat-sensitive capsaicin-gated transient receptor potential vanilloid 1 (TRPV1). It is remarkable to find that TRPM8, in addition to its behavior as an outwardly rectifying, nonselective cation channel, shares many functional and pharmacological properties with TRPV1. TRPM8 exhibits prominent time- and voltage-dependent behavior, a property that may underlie the conserved rectification or gating mechanisms exhibited by these channels. We also show that TRPM8 is modulated by ethanol but unlike TRPV1 is insensitive to extracellular acidification. There is also significant overlap in the antagonist pharmacology of these channels with many TRPV1 antagonists such as capsazepine, N-(4-tertiarybutylphenyl)-4-(3-chloropyridin-2-yl) tetrahydropyrazine-1(2H)-carboxamide (BCTC), (2R)-4-(3-chloro-2-pyridinyl)-2-methyl-N-[4-(trifluoromethyl)phenyl]-1-piperazinecarboxamide (CTPC), and N-(2-bromophenyl)-N'-{2-[ethyl(3-methylphenyl)amino]ethyl}-urea (SB-452533) exhibiting similar activity at TRPM8. Overall, the degree of pharmacological overlap between TRPV1 and TRPM8 has implications for the interpretation of studies conducted with these ligands to date and highlights a clear challenge for the design of selective TRP channel antagonists. Our finding that N-(3-methoxyphenyl)-4-chlorocinnamide (SB-366791), at least, represents an apparently selective antagonist for TRPV1 suggests that this goal is attainable.
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http://dx.doi.org/10.1124/mol.105.012146DOI Listing
August 2005

Effects of piperine, the pungent component of black pepper, at the human vanilloid receptor (TRPV1).

Br J Pharmacol 2005 Mar;144(6):781-90

Neurology & GI-CEDD, GlaxoSmithKline, New Fronteirs Science Park, Third Avenue, Harlow, Essex CM19 5AW.

1. We have characterised the effects of piperine, a pungent alkaloid found in black pepper, on the human vanilloid receptor TRPV1 using whole-cell patch-clamp electrophysiology. 2. Piperine produced a clear agonist activity at the human TRPV1 receptor yielding rapidly activating whole-cell currents that were antagonised by the competitive TRPV1 antagonist capsazepine and the non-competitive TRPV1 blocker ruthenium red. 3. The current-voltage relationship of piperine-activated currents showed pronounced outward rectification (25+/-4-fold between -70 and +70 mV) and a reversal potential of 0.0+/-0.4 mV, which was indistinguishable from that of the prototypical TRPV1 agonist capsaicin. 4. Although piperine was a less potent agonist (EC50=37.9+/-1.9 microM) than capsaicin (EC50=0.29+/-0.05 microM), it demonstrated a much greater efficacy (approximately two-fold) at TRPV1. 5. This difference in efficacy did not appear to be related to the proton-mediated regulation of the receptor since a similar degree of potentiation was observed for responses evoked by piperine (230+/-20%, n=11) or capsaicin (284+/-32%, n=8) upon acidification to pH 6.5. 6. The effects of piperine upon receptor desensitisation were also unable to explain this effect since piperine resulted in more pronounced macroscopic desensitisation (t(1/2)=9.9+/-0.7 s) than capsaicin (t(1/2)>20 s) and also caused greater tachyphylaxis in response to repetitive agonist applications. 7. Overall, our data suggest that the effects of piperine at human TRPV1 are similar to those of capsaicin except for its propensity to induce greater receptor desensitisation and, rather remarkably, exhibit a greater efficacy than capsaicin itself. These results may provide insight into the TRPV1-mediated effects of piperine on gastrointestinal function.
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http://dx.doi.org/10.1038/sj.bjp.0706040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1576058PMC
March 2005

Discovery of small molecule antagonists of TRPV1.

Bioorg Med Chem Lett 2004 Jul;14(14):3631-4

Neurology and GI CEDD, New Frontiers Science Park, GlaxoSmithKline, Third Avenue, Harlow, Essex CM19 5AW, UK.

Small molecule antagonists of the vanilloid receptor 1 (TRPV1, also known as VR1) are disclosed. Ureas such as 5 (SB-452533) were used to explore the structure activity relationship with several potent analogues identified. Pharmacological studies using electrophysiological and FLIPR Ca(2+) based assays showed compound 5 was an antagonist versus capsaicin, noxious heat and acid mediated activation of TRPV1. Study of a quaternary salt of 5 supports a mode of action in which compounds from this series cause inhibition via an extracellularly accessible binding site on the TRPV1 receptor.
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http://dx.doi.org/10.1016/j.bmcl.2004.05.028DOI Listing
July 2004

TRPV channels as temperature sensors.

Cell Calcium 2003 May-Jun;33(5-6):479-87

Neurology, GlaxoSmithKline Research and Development Ltd., New Frontiers Science Park (North), Third Avenue, Harlow, Essex CM19 5AW, UK.

The past year has seen a doubling in the number of heat-sensitive ion channels to six, and four of these channels are from the TRPV family. These channels characteristically have Q(10) values of >10 above the thermal threshold, very different from the Q(10) values of 1.5-2.0 seen in most ion channels. Cells expressing TRPV1 show similar temperature sensitivity to small capsaicin-sensitive nociceptor neurons, consistent with these neurons expressing homomers of TRPV1. A-delta fibres exhibit properties that may be explained by TRPV2 containing channels which is present in large diameter sensory neurons that do not express TRPV1. TRPV3 has a lower temperature threshold and may contribute to warm-sensitive channels together with TRPV1. Warm sensation may also be transduced by TRPV4 expressing sensory neurons and hypothalamic neurons. We can now look forward to further work defining the properties of the recombinant channels in more detail and a re-analysis of endogenous i(heat) currents in thermosensitive neurons and other cells. Data from the study of mice in which TRPV2, TRPV3 or TRPV4 have been deleted are also eagerly awaited.
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http://dx.doi.org/10.1016/s0143-4160(03)00063-0DOI Listing
February 2004

The diversity in the vanilloid (TRPV) receptor family of ion channels.

Trends Pharmacol Sci 2002 Apr;23(4):183-91

Neurology-CEDD, GlaxoSmithKline, New Frontiers Science Park, Third Avenue, CM19 5AW, Harlow, UK.

Following cloning of the vanilloid receptor 1 (VR1) at least four other related proteins have been identified. Together, these form a distinct subgroup of the transient receptor potential (TRP) family of ion channels. Members of the vanilloid receptor family (TRPV) are activated by a diverse range of stimuli, including heat, protons, lipids, phorbols, phosphorylation, changes in extracellular osmolarity and/or pressure, and depletion of intracellular Ca2+ stores. However, VR1 remains the only channel activated by vanilloids such as capsaicin. These channels are excellent molecular candidates to fulfil a range of sensory and/or cellular roles that are well characterized physiologically. Furthermore, as novel pharmacological targets, the vanilloid receptors have potential for the development of many future disease treatments.
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http://dx.doi.org/10.1016/s0165-6147(02)01999-5DOI Listing
April 2002

Cloning and functional expression of a human orthologue of rat vanilloid receptor-1.

Pain 2000 Nov;88(2):205-215

Department of Biotechnology and Genetics, SmithKline Beecham Pharmaceuticals, New Frontiers Science Park North, Third Avenue, Harlow CM19 5AW, UK Department of Neuroscience Research, SmithKline Beecham Pharmaceuticals, New Frontiers Science Park North, Third Avenue, Harlow CM19 5AW, UK Department of Bioinformatics, SmithKline Beecham Pharmaceuticals, New Frontiers Science Park North, Third Avenue, Harlow CM19 5AW, UK.

Capsaicin, resiniferatoxin, protons or heat have been shown to activate an ion channel, termed the rat vanilloid receptor-1 (rVR1), originally isolated by expression cloning for a capsaicin sensitive phenotype. Here we describe the cloning of a human vanilloid receptor-1 (hVR1) cDNA containing a 2517 bp open reading frame that encodes a protein with 92% homology to the rat vanilloid receptor-1. Oocytes or mammalian cells expressing this cDNA respond to capsaicin, pH and temperature by generating inward membrane currents. Mammalian cells transfected with human VR1 respond to capsaicin with an increase in intracellular calcium. The human VR1 has a chromosomal location of 17p13 and is expressed in human dorsal root ganglia and also at low levels throughout a wide range of CNS and peripheral tissues. Together the sequence homology, similar expression profile and functional properties confirm that the cloned cDNA represents the human orthologue of rat VR1.
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http://dx.doi.org/10.1016/S0304-3959(00)00353-5DOI Listing
November 2000