Coadministered cannabidiol and clobazam: Preclinical evidence for both pharmacodynamic and pharmacokinetic interactions

Abstract

Objective: Cannabidiol (CBD) has been approved by the US Food and Drug Administration (FDA) to treat intractable childhood epilepsies, such as Dravet syndrome and Lennox-Gastaut syndrome. However, the intrinsic anticonvulsant activity of CBD has been questioned due to a pharmacokinetic interaction between CBD and a first-line medication, clobazam. This recognized interaction has led to speculation that the anticonvulsant efficacy of CBD may simply reflect CBD augmenting clobazam exposure. The present study aimed to address the nature of the interaction between CBD and clobazam.

Methods: We examined whether CBD inhibits human CYP3A4 and CYP2C19 mediated metabolism of clobazam and N-desmethylclobazam (N-CLB), respectively, and performed studies assessing the effects of CBD on brain and plasma pharmacokinetics of clobazam in mice. We then used the Scn1a^+/- mouse model of Dravet syndrome to examine how CBD and clobazam interact. We compared anticonvulsant effects of CBD-clobazam combination therapy to monotherapy against thermally-induced seizures, spontaneous seizures and mortality in Scn1a^+/- mice. In addition, we used Xenopus oocytes expressing γ-aminobutyric acid (GABA)_A receptors to investigate the activity of GABA_A receptors when treated with CBD and clobazam together.

Results: CBD potently inhibited CYP3A4 mediated metabolism of clobazam and CYP2C19 mediated metabolism of N-CLB. Combination CBD-clobazam treatment resulted in greater anticonvulsant efficacy in Scn1a^+/- mice, but only when an anticonvulsant dose of CBD was used. It is important to note that a sub-anticonvulsant dose of CBD did not promote greater anticonvulsant effects despite increasing plasma clobazam concentrations. In addition, we delineated a novel pharmacodynamic mechanism where CBD and clobazam together enhanced inhibitory GABA_A receptor activation.

Significance: Our study highlights the involvement of both pharmacodynamic and pharmacokinetic interactions between CBD and clobazam that may contribute to its efficacy in Dravet syndrome.

Keywords: Dravet syndrome; cannabidiol; cannabis; clobazam; epilepsy.

Figure 1

Cannabidiol (CBD) inhibits the metabolism of clobazam and its active metabolite, N‐desmethylclobazam (N‐ CLB). Substrate‐velocity curves of (A) CYP3A4‐mediated metabolism of clobazam and (B) CYP2C19‐mediated metabolism of N‐ CLB in the presence of varying concentrations of CBD. Error bars represent standard error of the mean (SEM), with n = 3. Clobazam (5 mg/kg, i.p. injection) was administered in the absence (black) and presence (green‐blue) of 12 mg/kg CBD (i.p.). Concentration‐time curves for clobazam (solid lines) in mouse (C) plasma and (D) brain and N‐ CLB (dashed lines) in (E) plasma and (F) brain. Error bars represent SEM, with n = 4 per time point. Curves represent fit of data using a two‐compartment model

Figure 2

Cannabidiol (CBD) and clobazam combination therapy improved survival of Scn1a ^+/− mice. A, Spontaneous generalized tonic‐clonic seizure (GTCS) frequency of individual untreated and drug‐treated mice. Treatment began after the induction of a single hyperthermia‐induced seizure and spontaneous GTCS were subsequently quantified over a 60‐h recording period. Clobazam in both the absence and presence of CBD significantly reduced the proportion of mice experiencing GTCS (blue, open symbols), with n = 15‐19 per group (**P < 0.005, ****P < 0.0001; Fisher's exact test). B, Survival curves comparing untreated and drug‐treated mice. Clobazam in the presence of CBD significantly improved survival (blue line), with n = 15‐19 per group (**P < 0.005; log‐rank Mantel‐Cox)

Figure 3

Increased anticonvulsant effect of co‐administered clobazam and cannabidiol (CBD) on hyperthermia‐induced seizures in Scn1a ^+/− mice. CBD (12 mg/kg) did not increase the anticonvulsant effects of clobazam on hyperthermia‐induced seizures in Scn1a ^+/− mice despite the presence of a pharmacokinetic interaction. A, Temperature threshold for generalized tonic‐clonic seizures (GTCS) in individual mice treated with varying doses of clobazam in the absence (white bars) or presence (gray bars) of CBD. Clobazam treatment significantly increased body temperature of GTCS compared to vehicle. Concurrent CBD treatment did not affect the response of clobazam to thermally‐induced seizures. Error bars represent standard error of the mean (SEM), with n = 13‐17 per treatment group (**P < 0.005, ****P < 0.0001 compared to vehicle; log‐rank Mantel‐Cox). B, Average clobazam plasma concentrations in Scn1a ^+/− mice treated with clobazam only (white bars) or clobazam and CBD (12 mg/kg; grey bars). Combination treatment resulted in significantly higher plasma clobazam concentrations (P < 0.001; two‐way analysis of variance [ANOVA] followed by Sidak's post hoc; *P < 0.05). Error bars represent SEM, with n = 5‐8 per group. C, Threshold temperature of individual mice for GTCS induced by hyperthermia following acute treatment with clobazam (0.1 or 1 mg/kg) or CBD at a higher, anticonvulsant dose (100 mg/kg) administered individually or as a combination. Low‐dose clobazam (0.1 mg/kg) had no effect of GTCS threshold. All other treatments resulted in a significantly improved response to thermal seizure induction compared to vehicle (blue, open symbols, P < 0.05). Combination clobazam (1 mg/kg) and CBD (100 mg/kg) treatment was significantly more effective than either treatment alone, with n = 16‐17 per group (**P < 0.005, ****P < 0.0001; log‐rank Mantel‐Cox). [Note: vehicle and clobazam alone are replotted from 3A for clarity.] D, Average clobazam plasma concentrations in Scn1a ^+/− mice from the hyperthermia‐induced seizure experiment treated with clobazam (0.1 or 1 mg/kg) alone (white bars) or clobazam and CBD (100 mg/kg; gray‐striped bars). Combination treatment resulted in significantly higher clobazam levels (P < 0.001; two‐way ANOVA followed by Sidak's post hoc; **P < 0.005). Error bars represent SEM, with n = 5‐7 per group

Figure 4

Interaction between cannabidiol (CBD) and clobazam at α1β2γ2 γ‐aminobutyric acid (GABA)_A receptors. A, Concentration‐response curves for clobazam (black) and CBD (gray) at GABA_A receptors. Clobazam and CBD modulate currents evoked by 15 μmol/L GABA. Data are shown as mean ± standard error of the mean (SEM) fit to the Hill equation, with n = 4 per group. B, Representative trace of currents evoked by GABA applied alone or in the presence of CBD, clobazam, or N‐desmethylclobazam (N‐ CLB) individually or in combination in oocytes expressing the GABA_A receptor. C, Paired modulation of 15 μmol/L GABA by CBD (gray, open symbols), clobazam (black, open symbols), N‐ CLB (red, open symbols), CBD and clobazam (gray, closed circles) and CBD and N‐ CLB (red, closed circles). Error bars represent SEM, with n = 7 (*P < 0.05; paired one‐way analysis of variance (ANOVA) followed by Dunnett's post hoc test)