Development of subtype selective GABAA modulators. CNS Spectr

Merck Sharp & Dohme Research Laboratories, Terlings Park, Eastwick Road, Essex CM20 1QR, England.
CNS spectrums (Impact Factor: 2.71). 02/2005; 10(1):21-7.
Source: PubMed


Drugs modulating gamma-aminobutyric acid (GABA) transmission via the benzodiazepine (BZ) site on the gamma-aminobutyric acid type A (GABAA) receptor have been in widespread use for more than 40 years to treat anxiety, epilepsy, and sleep disorders. These drugs have been shown to be safe, well tolerated, and effective although the mechanism by they produce a myriad of pharmacologic effects remains elusive. In recent years it has been discovered that, although the GABAA receptor is widely distributed in the brain, the substructure and composition of the receptor differs from between brain regions. Termed "GABAA receptor subtypes" their discovery leads to speculation that different subtypes may mediate specific effects of BZs such as anxiety or sedation. The phenotypic analysis of transgenic knock-in and knock-out mice in which particular GABAA receptors were rendered insensitive to the effects of BZ while others were unaffected confirmed this speculation. Subsequently, subtype-specific GABAA ligands were developed that, for example, retained the anxiolytic effects of BZs but were devoid of their sedative effects. Therefore, it may be possible to develop effective anxiolytic compounds that have a much reduced side-effect profile compared with existing drugs.

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    • "They are pentameric oligomers assembled with several subunit classes that may have multiple isoforms [1]–[3]. In adult rat brain, the most abundant subunits are α1, β2, and γ2 typical for 60% to 90% of GABAAR [4]. The α1 subunit is highly expressed throughout most brain regions especially in the cortex [5]. "
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    ABSTRACT: Background GABAA receptor (GABAAR) function is maintained by an endogenous phosphorylation mechanism for which the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is the kinase. This phosphorylation is specific to the long intracellular loop I2 of the α1 subunit at two identified serine and threonine residues. The phosphorylation state is opposed by an unknown membrane-bound phosphatase, which inhibition favors the phosphorylated state of the receptor and contributes to the maintenance of its function. In cortical nervous tissue from epileptogenic areas in patients with drug-resistant epilepsies, both the endogenous phosphorylation and the functional state of the GABAAR are deficient. Methodology/Principal Findings The aim of this study is to characterize the membrane-bound phosphatases counteracting the endogenous phosphorylation of GABAAR. We have developed a new analytical tool for in vitro detection of the phosphatase activities in cortical washed membranes by liquid chromatography coupled to mass spectrometry. The substrates are two synthetic phosphopeptides, each including one of the identified endogenous phosphorylation sites of the I2 loop of GABAAR α1 subunit. We have shown the presence of multiple and atypical phosphatases sensitive to zinc ions. Patch-clamp studies of the rundown of the GABAAR currents on acutely isolated rat pyramidal cells using the phosphatase inhibitor okadaic acid revealed a clear heterogeneity of the phosphatases counteracting the function of the GABAAR. Conclusion/Significance Our results provide new insights on the regulation of GABAAR endogenous phosphorylation and function by several and atypical membrane-bound phosphatases specific to the α1 subunit of the receptor. By identifying specific inhibitors of these enzymes, novel development of antiepileptic drugs in patients with drug-resistant epilepsies may be proposed.
    PLoS ONE 06/2014; 9(6):e100612. DOI:10.1371/journal.pone.0100612 · 3.23 Impact Factor
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    • "The studies with point-mutated mice, made insensitive to diazepam at only one of four populations of BZ-sensitive GABA A receptors, have attributed the sedative and ataxic properties of this standard BZ to the α 1 subunit-containing GABA A receptor, the anxiolytic and muscle relaxant actions to the α 2 /α 3 subtype-containing receptors, and anticonvulsant activity, in part, to all the α 1 /α 2 /α 3 GABA A receptors (reviewed in Rudolph and Möhler 2004). Based on these results, inactivity/low-activity at α 1 GABA A receptors has evolved as the key concept for development of new BZ-like drugs, purportedly devoid of many adverse effects of standard BZs, such as sedation or abuse liability (Dawson et al. 2005; Rudolph and Knoflach 2011). However, generalization of this concept in the quest for anxioselective anxiolytics has been questioned by the findings in humans that several ligands, reportedly possessing properties of either null or low efficacy at α 1 GABA A receptors, still exerted the effects reflective of sedation (reviewed in Skolnick 2012). "
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    ABSTRACT: Rationale: Synthesis of ligands inactive or with low activity at α₁ GABA(A) receptors has become the key concept for development of novel, more tolerable benzodiazepine (BZ)-like drugs. WYS8, a remarkably (105 times) α₁-subtype selective partial positive modulator, may serve as a pharmacological tool for refining the role of α₁ GABA(A) receptors in mediation of BZs' effects. Objectives: Here, the effects of WYS8 on GABA-induced currents and on diazepam-induced potentiation of recombinant BZ-sensitive GABA(A) receptors were studied in more detail. In addition, the behavioral profile of WYS8 (0.2, 1, and 10 mg/kg i.p.), on its own and in combination with diazepam, was tested in the spontaneous locomotor activity, elevated plus maze, grip strength, rotarod, and pentylenetetrazole tests. Results: WYS8, applied at an in vivo attainable concentration of 100 nM, reduced the stimulation of GABA currents by 1 μM diazepam by 57 % at α₁β₃γ₂, but not at α₂β₃γ₂, α₃β₃γ₂, or α₅β₃γ₂ GABA(A) receptors. The administration of WYS8 alone induced negligible behavioral consequences. When combined with diazepam, WYS8 caused a reduction in sedation, muscle relaxation, and anticonvulsant activity, as compared with this BZ alone, whereas ataxia was preserved, and the anxiolytic effect of 2 mg/kg diazepam was unmasked. Conclusions: Hence, a partial instead of full activation at α₁ GABA(A) receptors did not necessarily result in the attenuation of the effects assumed to be mediated by activation of these receptors, or in the full preservation of the effects mediated by activation of other GABA(A) receptors. Thus, the role of α₁ GABA(A) receptors appears more complex than that proposed by genetic studies.
    Psychopharmacology 05/2013; 230(1). DOI:10.1007/s00213-013-3143-4 · 3.88 Impact Factor
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    • "GABA A receptors are heteropentameric, and the majority of those present in the CNS contain two α, two β, and a single γ subunit [1]. Benzodiazepines are allosteric ligands, that is, they exhibit no intrinsic activity of their own, but potentiate or inhibit the effects of GABA at receptors that contain either an α1, 2, 3, or 5 subunit [2]. GABA activation of GABA A receptors leads to the opening of their integrated chloride channels. "
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    ABSTRACT: GABA(A) receptors containing α2/3 subunits are current targets in the battle to develop new pain medications, as they are expressed in the spinal cord where increasing inhibitory drive should result in analgesia. However, this approach is prone to a range of side effects including sedation, cognitive impairment, and abuse as a consequence of the widespread influence of GABA. The ability to make subtype selective low-efficacy benzodiazepine compounds, which potentiate the action of GABA at specific α subunits, has the potential to reduce this side effect profile. In this study, we have investigated the effects of the medium-efficacy positive allosteric modulator (PAM) L-838,417 and the low-efficacy PAM TPA023 in a number of preclinical inflammatory and neuropathic pain models. We conclude that either the higher level of efficacy at α2/3 or efficacy at α5 is required for compounds to have a significant analgesic effect in a range of models, and, therefore, although the side-effect profile of compounds can be reduced compared to typical benzodiazepines, it is unlikely that it can be completely eliminated.
    Advances in Pharmacological Sciences 11/2011; 2011:608912. DOI:10.1155/2011/608912
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