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Clobazam and its active metabolite N-desmethylclobazam display significantly greater affinities for α₂- versus α₁-GABA(A)-receptor complexes - PubMed

  • ️Wed Jan 01 2014

Clobazam and its active metabolite N-desmethylclobazam display significantly greater affinities for α₂- versus α₁-GABA(A)-receptor complexes

Henrik Sindal Jensen et al. PLoS One. 2014.

Abstract

Clobazam (CLB), a 1,5-benzodiazepine (BZD), was FDA-approved in October 2011 for the adjunctive treatment of seizures associated with Lennox-Gastaut syndrome (LGS) in patients 2 years and older. BZDs exert various CNS effects through allosteric modulation of GABAA receptors. The structurally distinct, 1,4-BZD clonazepam (CLN) is also approved to treat LGS. The precise mechanisms of action and clinical efficacy of both are unknown. Data show that the GABAA α₁-subunit-selective compound zolpidem [ZOL] exhibits hypnotic/sedative effects. Conversely, data from knock-in mice carrying BZD binding site mutations suggest that the α₂ subunit mediates anticonvulsant effects, without sedative actions. Hence, the specific pattern of interactions across the GABAA receptor complexes of BZDs might be reflected in their clinical efficacies and adverse effect profiles. In this study, GABAA-receptor binding affinities of CLB, N-desmethylclobazam (N-CLB, the major metabolite of CLB), CLN, and ZOL were characterized with native receptors from rat-brain homogenates and on cloned receptors from HEK293 cells transfected with combinations of α (α₁, α₂, α₃, or α₅), β₂, and γ₂ subtypes. Our results demonstrate that CLB and N-CLB have significantly greater binding affinities for α₂- vs. α₁-receptor complexes, a difference not observed for CLN, for which no distinction between α₂ and α₁ receptors was observed. Our experiments with ZOL confirmed the high preference for α₁ receptors. These results provide potential clues to a new understanding of the pharmacologic modes of action of CLB and N-CLB.

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Conflict of interest statement

Competing Interests: The authors have read the journal's policy and have the following conflicts: This work was conducted and fully funded by Lundbeck. All authors were full-time Lundbeck employees at the time the analyses were conducted. Clobazam (Onfi) is a Lundbeck product. This does not alter the authors' adherence to all the PLoS ONE policies on sharing data and materials.

Figures

Figure 1
Figure 1. Saturation binding experiments of equilibrium binding of various concentrations of radio-ligand.

(A) Rat brain homogenate with 3H-flunitrazepam, and (B) recombinant human GABAA receptors with 3H-flumazenil.Total binding, non-specific binding, and specific binding are shown.

Figure 2
Figure 2

Distribution of individually determined pKi values for (A) clobazam, (B) N-desmethylclobazam, (C) clonazepam, and (D) zolpidem across GABAA-receptor subtypes.

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