Preclinical safety and efficacy of cannabidivarin for early life seizures

Abstract

A significant proportion of neonatal and childhood seizures are poorly controlled by existing anti-seizure drugs (ASDs), likely due to prominent differences in ionic homeostasis and network connectivity between the immature and mature brain. In addition to the poor efficacy of current ASDs, many induce apoptosis, impair synaptic development, and produce behavioral deficits when given during early postnatal development. There is growing interest in new targets, such as cannabidiol (CBD) and its propyl analog cannabidivarin (CBDV) for early life indications. While CBD was recently approved for treatment of refractory childhood epilepsies, little is known about the efficacy or safety of CBDV. Here, we addressed this gap through a systematic evaluation of CBDV against multiple seizure models in postnatal day (P) 10 and 20 animals. We also evaluated the impact of CBDV on acute neurotoxicity in immature rats. CBDV (50-200 mg/kg) displayed an age and model-specific profile of anticonvulsant action. In P10 rats, CBDV suppressed seizures only in the pentylenetetrazole model. In P20 rats, CBDV suppressed seizures in the pentylenetetrazole, DMCM, and maximal electroshock models. Between P10 and P20, we identified significant increases in mRNA expression of TRPV1 in multiple brain regions; when CBDV was tested in P20 TRPV1 knockout mice, anticonvulsant effects were attenuated. Finally, CBDV treatment generally avoided induction of neuronal degeneration in immature rats. Together, the efficacy and safety profile of CBDV suggest it may have therapeutic value for early life seizures.

Keywords: Cannabidivarin; Cannabinoid; Cell death; Neonatal; Seizure.

Figure 1.. CBDV attenuates the severity of DMCM-evoked seizures in postnatal day 20, but not postnatal day ten old rats.

Mean (+standard error of the mean [SEM]) seizure score as a function of drug treatment and dose in P10 (A) and P20 rats (C). Mean (+SEM) latency (in seconds) to first behavioral seizure manifestation in P10 (B) and P20 (D) rats. Asterisks indicate significant difference from vehicle control group at *p<0.05, ^p=0.085, Dunn’s tests for multiple comparisons.

Figure 2.. CBDV does not alter KA-evoked seizures in immature rats.

Mean (+SEM) seizure score as a function of drug treatment and dose in P10 (A) and P20 (C) rats. Mean (+SEM) latency to first behavioral seizure manifestation in P10 (B) and P20 (D) rats.

Figure 3.. CBDV attenuates the severity of PTZ-evoked seizures in immature rats.

Mean (+SEM) seizure score as a function of drug treatment and dose in P10 (A) and P20 (D) rats. Asterisks indicate significant difference compared to vehicle control, *p<0.05, **p<0.01, Dunn’s test for multiple comparisons. Mean (+SEM) latency (in seconds) to first behavioral clonic seizure manifestation in P10 (B) and P20 (E) rats. Mean (+SEM) latency (in seconds) to first behavioral tonic or tonic-clonic seizure manifestation in P10 (C) and P20 (F) and rats.

Figure 4.. CBDV protects against MES-evoked tonic seizures in postnatal day 20 rats.

Percent of P10 animals (A) and P20 (B) exhibiting tonic extension as a function of drug treatment. Asterisks indicate significant reduction in tonic seizures in P20 rats compared to vehicle control, ****p<0.0001, Fisher’s exact test.

Figure 5.. CBDV does not protect against hypoxia-induced seizures or NMDA evoked spasms in postnatal day 10 rats.

(A) Mean (+SEM) hypoxic seizure score as a function of drug treatment and dose. (B) Mean (+SEM) number of spasms (C) and latency (in seconds) to first flexion spasm as a function of drug treatment.

Figure 6.. mRNA expression levels of TRPV1, TRPV3, and GPR55 increase as a factor of age from postnatal day 10 to postnatal day 20 in rats.

Violin plot illustrating the fold change in TRPV1 (A), TRPV2 (B), TRPV3 (C), TRPA1 (D), and GPR55 (E) mRNA expression in P20 rats over P10 expression. *p<0.05, ^p=0.052 over P10 levels, one-sample t-test, Holm-Sidak corrected.

Figure 7.. TRPV1 expression mediates anticonvulsant effects of CBDV against MES seizures in postnatal day 20 mice.

Proportion of tonic seizures as a function of drug treatment and genotype. 100% of vehicle treated wild-type (8 of 8) and TRPV1 knockout (9 of 9) mice displayed tonic hindlimb extension. 100% of CBDV treated TRPV1 knockout (10/10) mice displayed tonic hindlimb extension. 37.5% of CBDV treated wild-type (3/8) mice displayed tonic hindlimb extension. The difference between CBDV-treated wild-type and TRPV1 mice differed significantly; *p<0.05, Chi-Square followed by Fisher’s exact test, Holm-Sidak corrected for multiple comparisons.

Figure 8.. CBDV increases cell death in the lateral thalamus, but not other regions in the postnatal day 7 rat brain.

Quantification of cell death as indicated by Fluoro-Jade B positive cells in: (A) cingulate cortex, (B) motor cortex, (C) somatosensory cortex, (D) striatum, (E) lateral septum, and (F) lateral thalamus from P7 rat pups treated with SAL, PB 75 mg/kg, VEH, or CBDV (100 or 200 mg/kg). Values are expressed as mean (+SEM) of 3 sequential 40 μm-thick, 1.18 mm² tissue samples through each region of interest, averaged per animal. *p<0.05, ***p=0.0001, ****p<0.0001; significantly different from SAL or VEH control group. Scale bar = 200 μm. Red boxes indicate the area of interest imaged for each brain region.