Functional Characterization of the 1,5-Benzodiazepine Clobazam and Its Major Active Metabolite N-Desmethylclobazam at Human GABA(A) Receptors Expressed in Xenopus laevis Oocytes

The 1,5-benzodiazepine clobazam is indicated for the adjunctive treatment of seizures associated with Lennox-Gastaut syndrome in patients 2 years of age or older in the United States, and for treatment of anxiety and various forms of epilepsy elsewhere. Clobazam has been reported to exhibit different in vivo adverse effects and addiction liability profile than the classic 1,4-benzodiazepines. In this study, it was investigated whether the in vitro pharmacological properties of clobazam and its major active metabolite N-desmethylclobazam could explain some of these clinical differences. The functional properties of the two 1,5-benzodiazepines were characterized at the human γ-aminobutyric acid type A receptor (GABA(A)R) subtypes α(1)β(2)γ(2S), α(2)β(2)γ(2S), α(3)β(2)γ(2S), α(5)β(2)γ(2S) and α(6)β(2)δ expressed in Xenopus laevis oocytes by use of two-electrode voltage-clamp electrophysiology and compared to those exhibited by the 1,4-benzodiazepine clonazepam. All three compounds potentiated GABA EC(20)-evoked responses through the α(1,2,3,5)β(2)γ(2S) GABA(A)Rs in a reversible and concentration-dependent manner, with each displaying similar EC(50) values at the four subtypes. Furthermore, the degrees of potentiation of the GABA EC(20) currents through the four receptors mediated by saturating modulator concentrations did not differ substantially for any of the three benzodiazepines. The three compounds were substantially less potent (200-3900 fold) as positive allosteric modulators at the α(6)β(2)δ GABA(A)R than at the α(1,2,3,5)β(2)γ(2S) receptors. Interestingly, however, clobazam and especially N-desmethylclobazam were highly efficacious potentiators of α(6)β(2)δ receptor signaling. Although this activity component is unlikely to contribute to the in vivo effects of clobazam/N-desmethylclobazam, the 1,5-benzodiazepine could constitute an interesting lead for novel modulators targeting this low-affinity binding site in GABA(A)Rs. In conclusion, the non-selective modulation exerted by clobazam, N-desmethylclobazam and clonazepam at the α(1)β(2)γ(2S), α(2)β(2)γ(2S), α(3)β(2)γ(2S) and α(5)β(2)γ(2S) GABA(A)Rs indicate that the observed clinical differences between clobazam and 1,4-benzodiazepines are likely to arise from factors other than their respective pharmacological properties at the GABA(A)Rs as investigated here.