Positive allosteric modulation of GABA_A receptors by a novel antiepileptic drug cenobamate

Cenobamate is a novel antiepileptic drug under investigation for use in patients with focal (partial-onset) seizures. To understand its potential molecular mechanism of action, the effects of cenobamate on GABA_A-mediated currents and GABA_A receptors in rodent hippocampal neurons were examined. Cenobamate potentiated GABA-induced currents (I_GABA) in acutely isolated CA3 pyramidal cells in a concentration-dependent manner (EC₅₀, 164 μM), which was not affected by flumazenil, a benzodiazepine receptor antagonist. Cenobamate enhanced tonic GABA_A currents (I_tonic), which is defined as a holding current shift by the GABA_A receptor antagonist bicuculline (EC₅₀, 36.63 μM). At therapeutically relevant concentrations, cenobamate induced minimal changes in the frequency, amplitudes, and decay time of spontaneous inhibitory postsynaptic currents in the CA1 neurons. Flumazenil failed to affect cenobamate-potentiated I_tonic and I_phasic in CA1 neurons. Cenobamate showed positive allosteric modulation of GABA-induced I_GABA mediated by GABA_A receptors. This effect was similar for all tested hGABA_A receptors containing six different alpha subunits (α₁β₂γ₂ or α_2-6β₃γ₂), with EC₅₀ values ranging from 42 to 194 μM. Cenobamate did not displace the binding of flunitrazepam, a benzodiazepine derivative, or flumazenil to GABA_A receptors. The results showed that cenobamate, a novel antiepileptic drug, acts as a positive allosteric modulator of high-affinity GABA_A receptors, activated by GABA at a site independent of the benzodiazepine binding site and efficiently enhances I_tonic inhibition in hippocampal neurons, which could be an underlying molecular mechanism stabilizing neural circuits of the epileptic hippocampus.