Identifying the molecular mechanisms of the anticonvulsant drugs cannabidiol, cannabidivarin and cannabidiolic acid using the model organism Dictyostelium discoideum. / Perry, Christopher.

2019. 170 p.

Research output: ThesisDoctoral Thesis

Unpublished

Documents

  • 2019PerryCJPhD

    Other version, 4 MB, PDF document

    Embargo ends: 19/03/24

Abstract

Cannabis has been used for the treatment of seizures for many centuries. Recent medical interest has focused upon the non-psychotropic compounds found in cannabis plants, such as cannabidiol (CBD), cannabidivarin (CBDV) and cannabidiolic acid (CBDA). CBD has been demonstrated to provide effective control of seizures in recent phase III clinical trials as a treatment (Epidolex®) for specific refractory epilepsies such as Dravet syndrome. However, the molecular mechanism(s) in which CDB and other cannabinoids exert their therapeutic effects remain unknown. This study aimed to identify the molecular mechanism(s) of CBD, CBDV and CBDA using D. discoideum as a model system, translating any findings across to mammalian epilepsy models. Screening of a mutant library with each of these cannabinoids identified 4 genes of interest that show cannabinoid resistance when disrupted. Of these genes gcvH1 (DDB_G0287773) was investigated further due to its role in the metabolism of the neurotransmitter glycine and its close association with the one carbon cycle. The resistance to CBD seen in the mutant screen was recapitulated by independently ablating gcvH1 from wild-type cells (Ax3). When either gcvH1 or the human orthologue gcsH was replaced back into the gcvH1 null cell by use of an RFP tagged extra chromosomal vector, sensitivity to CBD was reinstated. Furthermore, we show that the D. discoideum and the H. sapiens RFP-fusion proteins both localize to the mitochondria. Using both GC-MS and NMR analysis we show that CBD treatment in D. discoideum significantly lowered cellular methionine levels, a key component of the one carbon cycle, whereas no change was detected in the gcvH1 null. These findings were translated across to rat primary hippocampal neurons treated with PTZ to induce seizure like activity. It was found that following PTZ treatment, cellular methionine levels were significantly elevated compared to untreated cells. However, if an intervention group was pre-treated with CBD prior to PTZ treatment, methionine levels remained stable. We further show the effects of CBD within an in vivo epilepsy model, where heterozygous mice for the Dravet syndrome linked Scn1a gene are prone to convulsive episodes. We show that brain methionine levels within this model are significantly elevated following an oral treatment regime with CBD. Thus, our research strongly suggests that CBD may exert its anticonvulsant properties by altering cellular methionine levels, by targeting the one carbon cycle through the mitochondrial protein GCVH1.
Original languageEnglish
QualificationPh.D.
Awarding Institution
Supervisors/Advisors
Thesis sponsors
  • GW Pharmaceuticals
Award date1 Mar 2019
Publication statusUnpublished - 2019

ID: 33508196