Patent Application: US-201214345968-A

Abstract:
this invention relates to a pharmaceutical composition comprising or consisting essentially of the phytocannabinoids cannabidivarin and cannabidiol . the composition is particularly safe and efficacious for use in the treatment of neurological conditions , characterized by hyper - excitability of the central nervous system , convulsions or seizures such as occur in epilepsy . preferably the cbdv and the cbd are present with at least one non - cannabinoid component of cannabis such as one or more terpenes or a terpene fraction . more particularly the composition further comprises one or more cannabichromene type compounds . particularly cannabichromene propyl variant and / or cannabichromene . more particularly still the composition is absent or substantially absent of other cannabinoids , including in particular tetrahydrocannabinol and tetrahydrocannabivarin , which would normally be present in significant amounts in cannabis chemotypes bred to contain a significant amount of cbdv and / or cbd .

Description:
example 1 below describes the use of a cbdv botanical drug substance ( bds ) from which the cannabinoids thcv and thc have been selectively removed , hereinafter cbdv (−/−) bds . the ptz model of generalized seizures in epilepsy was used to determine the anti - convulsant activity of the test article . use of a composition comprising cbdv and cbd in the ptz model of generalised seizures male wistar rats ( p24 - 29 ; 75 - 110 g ) were used to assess the combined effect of a composition comprising the phytocannabinoids cbdv and cbd in the ptz model of generalised seizures . animals were habituated to the test environment , cages , injection protocol and handling prior to experimentation . animals were housed in a room at 21 ° c . on a 12 hour light : dark cycle ( lights on 0900 ) in 50 % humidity , with free access to food and water . the human dose equivalent ( hed ) can be estimated using the following formula : the k m for a rat is 6 and the k m for a human is 37 . thus , for a human of approx 60 kg a 200 mg / kg dose in rat would equate to a human daily dose of about 2000 mg . a composition was prepared using a cbdv botanical drug substance ( bds ) that had been further prepared by centrifugal partition chromatography to remove the cannabinoids thcv and thc , such that the cannabinoids consisted essentially of cbdv and cbd , and lesser amounts of cbcv and cbc . this bds is termed cbdv (−/−) bds for the purpose of this application . five 6 l perspex tanks with lids were placed on a single bench with dividers between them . closed - circuit television ( cctv ) cameras were mounted onto the dividers to observe rat behaviour . sony topica ccd cameras ( bluecherry , usa ) were linked via bnc cables to a low - noise pc via brooktree digital capture cards ( bluecherry , usa ). zoneminder ( http :// www . zoneminder . com ) software was used to monitor rats , start and end recordings and manage video files . in - house linux scripts were used to encode video files into a suitable format for further offline analysis using the observer ( noldus technologies ). a range of doses of ptz ( 50 - 100 mg / kg body weight ) were used to determine the best dose for induction of seizures . as a result , a dose of 85 mg / kg injected intra - peritoneally ( ip ; stock solution 50 mg / ml in 0 . 9 % saline ) were used to screen the cbdv (−/−) bds test article . on the day of testing , the cbdv (−/−) bds was administered via intra - peritoneal ( i . p .) injection at doses of 50 , 100 , 200 , 275 and 346 mg / kg alongside animals that were injected with a matched volume of the cannabinoid vehicle ( 2 : 1 : 17 ethanol : cremophor : saline ), which served as the negative control group , ( giving defined doses of cbdv and cbd as set out in table 1 . 1 below ). animals were then observed for 1 hour , after which time they received an ip injection of 85 mg / kg ptz . negative vehicle controls were performed in parallel with cannabinoid - dosed subjects . after receiving a dose of ptz , animals were observed and videoed to determine the severity of seizure and latency to several seizure behaviour types ( see in vivo analysis , below ). animals were filmed for half an hour after last sign of seizure , and then returned to their cage . table 1 . 1 below demonstrates the respective content of the cannabinoids cbdv and cbd in the different dose groups of the cbdv (−/−) bds . animals were observed during experimental procedures , but all analysis was performed offline on recorded video files using the observer behavioural analysis software ( noldus , netherlands ). a seizure severity scoring system was used to determine the levels of seizure experienced by subjects ( pohl & amp ; mares , 1987 ). all signs of seizure were detailed for all animals . table 1 . 2 seizure severity scoring scale , adapted from pohl & amp ; mares , 1987 . seizure score behavioural expression righting reflex 0 no changes to behaviour preserved 0 . 5 abnormal behaviour ( sniffing , preserved excessive washing , orientation ) 1 isolated myoclonic jerks preserved 2 atypical clonic seizure preserved 3 fully developed bilateral forelimb preserved clonus 3 . 5 forelimb clonus with tonic component preserved and body twist 4 tonic - clonic seizure with suppressed lost tonic phase 5 fully developed tonic - clonic seizure lost 6 death the latency ( in seconds ) from injection of ptz to first myoclonic jerk ( fmj ; score of 1 ), and to the animal attaining “ forelimb clonus with tonic component and body twist ” ( score of 3 . 5 ) were recorded . fmj is an indicator of the onset of seizure activity , whilst & gt ; 90 % of animals developed scores of 3 . 5 , and so is a good marker of the development of more severe seizures . data are presented as the mean ± s . e . m . within an experimental group . this is given as the median value for each experimental group based on the scoring scale below . the percentage of animals within an experimental group that died as a result of ptz - induced seizures . note that the majority of animals that developed tonic - clonic seizures ( scores of 4 and 5 ) died as a result , and that a score of 6 ( death ) automatically denotes that the animal also experienced tonic - clonic seizures . the time ( in seconds ) from the first sign of seizure ( typically fmj ) to either the last sign of seizure or , in the case of subjects that died , the time of death — separated into animals that survived and those that did not . this is given as the mean ± s . e . m . for each experimental group . for measures of latency and severity , one way analysis of variance ( anova ) was performed on all the groups together in order to detect overall effects of the test article ( 00 . 05 considered significant ), and is denoted by a ‘*’ in the figures . significant anova results were followed by post hoc tests to test differences between vehicle and drug groups ( tukey &# 39 ; s test , 00 . 05 considered significant ), and is denoted by a ‘*’ in the figures . fig1 illustrates the maximum seizure severity , a significant effect of the cbdv (−/−) bds on the maximum seizure severity was observed at a dose of 275 mg / kg cbdv (−/−) bds . fig2 illustrates the percentage mortality of the animals dosed with the cbdv (−/−) bds . as can be observed the animals given the , 200 and 275 mg / kg cbdv (−/−) bds had a strongly statistical significance and the animals given the highest dose ( 346 mg / kg cbdv (−/−) bds had a less statistical significance but still resulted in a decrease in the percentage mortality . fig3 illustrates that although no significant effect of the cbdv (−/−) bds was observed on the percentage of animals that were seizure free , the 275 mg / kg dose resulted in 20 % of the animals becoming seizure free . fig4 illustrates the latency to seizure onset was statistically increased in all of the high dose groups ( 200 , 275 and 346 mg / kg ) of the cbdv (−/−) bds . fig5 illustrates the percentage of animals that experienced the severe tonic - clonic seizures decreased in the higher dose groups ( 200 , 275 and 346 mg / kg ) of the cbdv (−/−) bds ; however the decrease was not statistically significant . from the above data it would appear that the cbdv (−/−) bds composition will reduce seizure severity and mortality and increase latency to onset of seizures , making it a desirable composition for use in the treatment of epilepsy . the omission of the cannabinoids thcv and thc from a bds further obviates concerns associated with cb1 antagonism and psychosis . the cbdv (−/−) bds which was used in example 1 above can be obtained using centrifugal partition chromatography ( cpc ) of a cbdv (+/+) bds . a cbdv (−/−) bds has been produced and analysed as described in table 2 . 1 below : the total phytocannabinoid containing fraction of cbdv (−/−) bds comprises approximately 41 % of the total bds . according to variation this fraction may vary by ± 10 % up to ± 50 %. the amount of the principle phytocannabinoid in the cbdv (−/−) bds as a percentage of the phytocannabinoid containing fraction is approximately 61 %. according to variation this fraction may vary by ± 10 % up to ± 50 %. in this example it is intended that references be made to the principle or secondary components independently of the ‘ other ’ cannabinoids . the following example is included to provide details of the components of the cbdv (+/+) bds . the cbdv (+/+) bds was obtained by subcritical co 2 extraction . it comprises , as well as cbdv , the cannabinoids cbd , thcv and thc in significant quantities ( each greater than 1 % by weight as a percentage of total cannabinoid content ). thc has been ascribed a pro - convulsant and it can also have marked psychoactive effects in addition to other side effects such as anxiety which are not desired . thcv whilst showing anti - convulsant activity specific to generalized seizures in epilepsy is a cb1 antagonist and following evidence to suggest that the cb1 antagonist rimonabant may cause epilepsy and other undesired effects it may be desirable to remove these cannabinoids from a bds whilst still retaining the non - cannabinoid component ( s ) which may contribute to the activity of the bds . a cbdv (+/+) bds can be obtained from extraction of cbdv - rich plants . such chemovars are bred specifically to produce a significant proportion of their cannabinoids as cbdv . the cbdv chemotype results from the breeding of plants which carry both postulated b d and a pr genes . the b d gene instruct the plants to synthesize the cyclic part of the cbd molecule and the a pr gene instructs the plant to synthesize this molecule with a propyl side chain , as opposed to the usual pentyl chain found in cbd . a cbdv chemovar has been bred and the bds analysed as described in table 3 . 1 below : the total phytocannabinoid containing fraction of cbdv (+/+) bds comprises approximately 41 % of the total bds . according to variation this fraction may vary by ± 10 % up to ± 50 %. the amount of the principle phytocannabinoid in the cbdv (+/+) bds as a percentage of the phytocannabinoid containing fraction is approximately 58 %. according to variation this fraction may vary by ± 10 % up to ± 50 %. in this example it is intended that references be made to the principle or secondary components independently of the ‘ other ’ cannabinoids . this comparative example is included to demonstrate a typical terpene profile obtained from a cannabis plant that has been bred to produce a high quantity of cannabinoids . the non - cannabinoid components of a phytocannabinoid bds may play an important role in the bds &# 39 ; s pharmacology . as such the terpene profile is classified below . the following tables illustrate the terpene profile of a cbd chemovar which is representative of a high phytocannabinoid containing plant . five plants were freshly harvested and extracted using steam distillation . the principle monoterpene and sesquiterpene are highlighted in bold . the monoterpene containing fraction comprises approximately 52 - 64 % ( w / w ) of the total terpene fraction . the amount of the principle monoterpene myrcene in the monoterpene fraction as a percentage of the monoterpene fraction is approximately 61 - 75 % ( w / w ). the monoterpene fraction also has a secondary monoterpene pinene which is present at approximately 16 . 3 - 20 % ( w / w ) of the monoterpene fraction . the sesquiterpene containing fraction comprises approximately 27 - 32 % ( w / w ) of the total terpene fraction . this comparative example describes the terpene profile of a different cannabis plant to that described on example 4 above and is reproduced here for comparative purposes . patent application number pct / gb2008 / 001837 describes the production of a ‘ zero cannabinoid ’ plant . these plants were produced by selective breeding to produce a cannabis sativa l plant that contained a generally qualitatively similar terpene profile as a cannabis sativa l plant that produced cannabinoids yet it was devoid of any cannabinoids . these plants can be used to produce cannabinoid - free plant extracts which are useful control plants in experiments and clinical trials . a breakdown of the terpene profile produced in the plants can be found in the table below . the primary monoterpenes and sesquiterpene are highlighted in bold . the monoterpene containing fraction comprises approximately 65 - 79 % ( w / w ) of the total terpene fraction . the sesquiterpene containing fraction comprises approximately 25 - 31 % ( w / w ) of the total terpene fraction . the amount of the principle sesquiterpene caryophylene in the sesquiterpene fraction as a percentage of the sesquiterpene fraction is approximately 35 - 43 % ( w / w ). the sesquiterpene fraction also has a secondary sesquiterpene humulene which is present at approximately 16 - 20 % ( w / w ) of the sesquiterpene fraction . use of cbdv (+/+) bds in the ptz model of generalised seizures this comparative example was previously presented in gb1005364 . 3 ( unpublished ) patent application and is included here for representative purposes . cbdv (+/+) bds was administered at four doses that yielded a dose of cbdv of 50 and 100 mg / kg . table 6 . 1 below details the data obtained . as can be seen the cbdv (+/+) bds exhibited a trend to decrease seizure - related mortality . alger , b . e . 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