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A Phase I, open-label, randomized, crossover study in three parallel groups to evaluate the effect of Rifampicin, Ketoconazole, and Omeprazole on the pharmacokinetics of THC/CBD oromucosal spray in healthy volunteers

A Phase I, open-label, randomized, crossover study in three parallel groups to evaluate the... This Phase I study aimed to assess the potential drug-drug interactions (pharmacokinetic [PK] and safety profile) of Δ9-tetrahydrocannabinol (THC)/cannabidiol (CBD) oromucosal spray (Sativex®, nabiximols) in combination with cytochrome P450 (CYP450) inducer (rifampicin) or inhibitors (ketoconazole or omeprazole). Thirty-six healthy male subjects were divided into three groups of 12, and then randomized to one of two treatment sequences per group. Subjects received four sprays of THC/CBD (10.8/10 mg) alongside single doses of the CYP3A and 2C19 inducer rifampicin (600 mg), CYP3A inhibitor ketoconazole (400 mg) or CYP2C19 inhibitor omeprazole (40 mg). Plasma samples were analyzed for CBD, THC and its metabolite 11-hydroxy-THC (11-OH-THC). A single dose of four sprays of THC/CBD spray (10.8/10 mg) following repeated doses of rifampicin (600 mg) reduced the C and AUC of all analytes. C reduced from 2.94 to 1.88 ng/mL (-36%), 1.03 to 0.50 ng/mL (-52%) max max and 3.38 to 0.45 ng/mL (-87%) for THC, CBD and 11-OH-THC, respectively compared to single dose administration of THC/CBD spray alone. Ketoconazole co-administration with THC/CBD spray had the opposite effect, increasing the C of the respective analytes from 2.65 to 3.36 ng/mL (+27%), 0.66 to 1.25 ng/mL (+89%) and 3.59 to 10.92 max ng/mL (+204%). No significant deviations in C or AUC for any analyte were observed when THC/CBD spray was max co-administered with omeprazole. THC/CBD spray was well tolerated by the study subjects both alone and in combination with rifampicin, ketoconazole and omeprazole. Evaluation of the PKs of THC/CBD spray alone and in combination with CYP450 inhibitors/inducers suggests that all analytes are substrates for the isoenzyme CYP3A4, but not CYP2C19. On the basis of our findings, there is likely to be little impact on other drugs metabolized by CYP enzymes on the PK parameters of THC/CBD spray, but potential effects should be taken into consideration when co-administering THC/CBD spray with compounds which share the CYP3A4 pathway such as rifampicin or ketoconazole. Trials registration: NCT01323465 † † Keywords: Cannabidiol, Cytochrome P450, Delta-9-tetrahydrocannabinol, Sativex® , Nabiximols , THC/CBD spray * Correspondence: cgs@gwpharm.com GW Pharma Ltd, Porton Down Science Park Salisbury, Wiltshire SP4 0JQ, UK Full list of author information is available at the end of the article © 2013 Stott et al.; licensee Springer. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Stott et al. SpringerPlus 2013, 2:236 Page 2 of 15 http://www.springerplus.com/content/2/1/236 Introduction clinically significant interactions from occurring. More- The endocannabinoid system modulator Δ9-tetrahydro- over, avoiding co-administration or adjusting a patient's cannabinol (THC)/cannabidiol (CBD) oromucosal spray drug regimen early in the course of therapy can provide (Sativex®, nabiximols) has been reported to be effective optimal response with minimal AEs (Ogu & Maxa 2000). in relieving a number of multiple sclerosis (MS) symp- Many different CYP450 isoenzymes have been identified toms including spasticity, central neuropathic pain and to-date, including six which play important roles in drug bladder dysfunction (Rog et al. 2005; Johnson et al. 2010; metabolism (DiPiro 1999; Cupp & Tracy 1998): CYP1A2, Rog et al. 2007), and has recently been approved in vari- CYP2C19, CYP2C9, CYP2D6, CYP2E1, and CYP3A4. ous European countries and abroad (i.e. in Canada, CYP450 inhibitors and inducers are known to affect Israel, New Zealand) as add-on treatment for spasticity the metabolism of THC. Previous literature reports have in MS patients. Other potential indications for this com- indicated that CBs, especially THC, are metabolized by pound include pain relief in advanced cancer (Johnson CYP3A4, 2C9, 2C19 and possibly 2D6 in humans et al. 2010; Porteney et al. 2012), as well as peripheral (Huestis 2007), and that the primary metabolites of neuropathic pain in MS (Nurmikko et al. 2007). Derived THC and CBD are 11-hydroxy-THC (11-OH-THC) and from proprietary cannabis plant varieties bred to exhibit 7-hydroxy-CBD, respectively (Huestis 2007). The forma- a pre-determined content of cannabinoids (CBs), THC/ tion of 11-OH-THC has been reported to be primarily CBD spray is fully standardized and contains two princi- catalysed by CYP2C19 and 2C9 (Bland et al. 2005). pal CBs, THC and CBD at an approximately 1:1 ratio as In vitro studies of THC and CBD on CYP450 induc- well as minor amounts of other CBs and non-CB com- tion and inhibition indicate that both inhibit CYP1A1, ponents. The specified CBs constitute at least 90% of the 1A2 and 1B1 enzymes (Yamaori et al. 2010). CBD also total CB content of the extract, however, the minor CBs has an inhibitory effect on CYP3A4 and CYP2C19. and other constituents also contribute to the therapeutic However, this effect only occurred at high concentra- profile of THC/CBD spray (Russo 2011), and may be in- tions (IC = 6-9 μM) of CBD (GW unpublished data), volved in stabilizing the extract (Whittle et al. 2001). and in normal dosing, peak plasma concentrations of CBs are thought to act primarily via activation of spe- CBD are approximately 5 ng/mL or less, 400-fold lower cific CB receptors, CB and CB (Howlett et al. 2002). than the levels at which CYP inhibition may be antici- 1 2 CB is predominantly expressed in the central nervous pated. As such, it is unlikely that THC/CBD spray would system (CNS), while CB is primarily expressed in the cause a relevant inhibition of CYP450s. However, to in- periphery, especially in immune cells (Pertwee 2007). vestigate the potential interactions of THC and CBD Endogenous ligands (“endocannabinoids”) produced in with drugs which also interact with the CYP450s mammalian tissues target these receptors, and together CYP3A4 and CYP2C19, various known inducers/inhi- with the catabolic and metabolic enzymes and transporter bitors of these isoenzymes were employed and the systems they constitute the endocannabinoid system. pharmacokinetics (PKs) of their co-administration with Multiple drug therapy is often used with a single pa- THC/CBD spray evaluated. tient. As THC/CBD spray is indicated for MS and po- Rifampicin is an antibiotic drug, a strong inducer of tentially advanced cancer pain, the likelihood is high CYP3A4 and moderate inducer of CYPs 2C19, 2B6, 2C8 that patients would be receiving different concomitant and 2C9, and has been extensively used in clinical stud- medications. As such, drug-drug interactions could ies as a prototypical inducer of these enzymes (Division occur which affect the bioavailability of THC/CBD spray of Clinical Pharmacology 2012; Federal Drug Association through absorption, metabolism or disposition. In turn 2012). Ketoconazole is a synthetic antifungal drug, is a this could affect the treatment and adverse events (AEs) strong inhibitor of CYP3A4 (Federal Drug Association experienced by the patient (Chen & Raymond 2006). In 2012), and a weak inhibitor of CYPs 2C8 and 2C19 some incidences, AEs experienced due to drug-drug inter- (Federal Drug Association 2012). Omeprazole is a actions can be life-threatening, therefore understanding proton-pump inhibitor which is primarily metabolized the mechanisms of these interactions is important so that by, and demonstrates high affinity for CYP2C19 (Furuta dosing and safety information can be adjusted accordingly. et al. 2005), and is also a moderate inhibitor of 2C19 Cytochrome P450 (CYP450) is a family of isoenzymes (Federal Drug Association 2012). This study investigated responsible for the biotransformation of several drugs, the potential interaction of these CYP450 inhibitors/in- and drug metabolism via this system has emerged as an ducers on the PK and safety profile of THC/CBD spray important determinant of the occurrence of several in healthy male subjects. drug-drug interactions that can result in toxicity, re- duced pharmacological effect and AEs (Guengerich Methods 2008). Determining whether the drugs involved act as This open-label, randomized, crossover, drug-interaction enzyme substrates, inducers, or inhibitors can prevent study took place at one study site in the UK (Quintiles Stott et al. SpringerPlus 2013, 2:236 Page 3 of 15 http://www.springerplus.com/content/2/1/236 Drug Research Unit at Guys Hospital), was approved by Research (Quintiles Limited) labels. The aliquots were Guy's Research Ethics Committee, and was conducted stored in clearly labelled containers in a freezer set at or according to the International Conference on Harmon- below -20°C, until shipped for assay. Samples were isation guidelines on Good Clinical Practice and the eth- shipped on dry ice at the appropriate time-points. ical principles stated in the Declaration of Helsinki and local UK regulations. All participants gave written in- Analysis method formed consent. The assay validation was undertaken by Advanced Bioanalytical Service Laboratories (London, UK), who Study design and treatment groups developed the technique after reviewing the literature, A total of 36 healthy males subjects enrolled and were which was based on the methodology adopted by three divided into three groups of 12. Within each group par- different groups (Foltz et al. 1983; Goodall & Basteyns ticipants were randomized to one of two treatment se- 1995; Kemp et al. 1995), and designed with reference to quences with six subjects receiving each sequence. FDA guidelines for industry (FDA Guidance for Industry Subjects received four sprays of THC/CBD spray (10.8/ 2012). The method utilised protein precipitation, solvent 10 mg) alongside single usual daily doses of either rifam- extraction and derivatisation for the sample preparation picin (600 mg), ketoconazole (400 mg) or omeprazole and then sample analysis by capillary gas chromatog- (40 mg) according to the following sequences, designed raphy and detection by a mass spectrometer (GC-MS). with a time-frame that was standard and fitting to the The validation procedure investigated the calibration aims of this study: model with the best regression fit over the concentration range 0.1 - 100 ng/mL for CBD, THC and 11-OH-THC, Sequence 1A. Subjects received a single dose of 4 as well as precision and accuracy of the method, stabil- sprays THC/CBD on Day 1 and once daily ity, carry-over, and specificity. rifampicin on Days 2-10. Subjects then received Human plasma from healthy volunteers was used to both THC/CBD spray and rifampicin on Day 11. prepare the standards and quality control (QC) samples, Sequence 1B. Subjects received rifampicin on Days 1-9. with analytes extracted using hexane/ethyl acetate (7:1 Subjects received THC/CBD spray and rifampicin ratio), derivatised with N,O-Bis(trimethylsilyl)trifluoroa- on Day 10 and then a single dose of 4 sprays THC/ cetamide. Analytic grade THC (Sigma, UK), CBD CBD on Day 18. (Sigma, UK) and 11-OH-THC (Radian International and Sequence 2C. Subjects received a single dose of 4 Cerillant, UK) were obtained, and three sets of CBD, sprays THC/CBD on Day 1 and once daily THC and 11-OH-THC were used to support the study. ketoconazole on Days 2-5. Subjects then received Deuterated THC-d (Sigma, UK) was used as the in- both THC/CBD spray and ketoconazole on Day 6. ternal standard. The GC-MS equipment was a Hewlett Sequence 2D. Subjects received ketoconazole on Days Packard 6890 Gas Chromatograph attached to a Hewlett 1-4. Subjects received THC/CBD spray and once Packard 5973 Mass Selective Detector. Data handling daily ketoconazole on Day 5. Subjects then received was carried out using an MS Chemstation System and a single dose of 4 sprays THC/CBD on Day 10. the peak area ratio of the analytes to the internal stand- Sequence 3E. Subjects received a single dose of 4 ard was calculated in Excel (2000). The concentrations sprays THC/CBD on Day 1. Subjects received once were calculated from the ratio data using least squares daily omeprazole on Days 2-6, and then THC/CBD ln(y) on ln(x) regression performed in Excel (2000), and spray and omeprazole on Day 7. were then checked manually. Regression analysis was Sequence 3F. Subjects received a once daily dose of undertaken to find the regression model that best de- omeprazole on Days 1-5. Subjects received both scribed the calibration data (for details, see (Miller & THC/CBD spray and omeprazole on Day 6. Subjects Miller 1992)). then received a single dose of 4 sprays THC/CBD Intra-assay precision and bias was examined using on Day 9. spiked control samples analysed in replicates of five. Inter-assay precision and accuracy were analysed in Blood sampling procedure and plasma preparation quintuplet at three concentrations and on three separate Blood samples were collected at specified times and occasions. stored on ice (except rifampicin PK samples which were The lowest and upper Limits of Quantification (LOQ) stored in iced water) prior to processing and storage. were investigated by looking at five (500 μL) plasma Plasma samples were separated by centrifugation (ap- samples containing 0.10 ng/mL and 100.0 ng/mL proximately 2500 rpm × 15 minutes at 4°C). Samples (the lowest and highest calibrators) of THC, CBD and were stored in 4 mL amber glass screw top glass vials 11-OH-THC, assayed in one batch, possessing accept- with PTFE lined screw caps labelled with Guys Drug able precision and accuracy. As such, these lowest and Stott et al. SpringerPlus 2013, 2:236 Page 4 of 15 http://www.springerplus.com/content/2/1/236 upper LOQ were deemed suitable for the measurement abnormal findings upon physical examination, 12-lead of these analytes in human plasma over these concentra- electrocardiogram (ECG), medical history, clinical la- tion ranges. boratory at screening, or renal and hepatic function. The inter- and intra-assay accuracy of the assay calcu- Subjects were non-users of tobacco products and were lated for THC was -0.53, -0.45, -1.72% and -0.50, -2.10, - negative for Human Immunodeficiency Viruses I and II, 0.86% at plasma THC concentrations of 2.0, 20.0 and Hepatitis B surface antigen, and antibodies to the Hepa- 80.0 ng/mL, respectively. The inter- and intra-assay pre- titis C virus. Eligible subjects had a negative urine screen cision of the assay calculated for THC was 1.88, 2.51, for alcohol, drugs of abuse (screening only) and cotinine, 2.41% and 2.64, 1.09, 1.29% at plasma THC concentra- and were using an appropriate barrier method of contra- tions of 2.0, 20.0 and 80.0 ng/mL, respectively. ception in addition to a second method of barrier The inter- and intra-assay accuracy of the assay calcu- contraception being used by their partner for the study lated for CBD was -2.90, 0.81, 1.78% and -2.00, -0.75, duration and for three months following administration 2.38% at plasma CBD concentrations of 2.0, 20.0 and of THC/CBD spray. 80.0 ng/mL, respectively. The inter- and intra-assay pre- cision of the assay calculated for CBD was 4.05, 2.28, Exclusion criteria 2.31% and 6.67, 1.43, 1.08% at plasma CBD concentra- Subjects with a history of significant cardiovascular, pul- tions of 2.0, 20.0 and 80.0 ng/mL, respectively. monary, hepatic, renal, haematologic, gastrointestinal, The inter- and intra-assay accuracy of the assay calcu- endocrine, immunologic, dermatologic, neurologic, or lated for 11-OH-THC was 0.00, 0.20, -3.44% and -0.46, - psychiatric disorder were excluded. Those with a history 0.05, -1.43% at plasma 11-OH-THC concentrations of of alcohol or drug abuse within two years of the study 2.17, 21.70 and 86.80 ng/mL, respectively. The inter- were also excluded; however, those with a history of pre- and intra-assay precision of the assay calculated for vious cannabis use were not excluded if willing to ab- 11-OH-THC was 3.45, 3.44, 2.57% and 6.04, 1.96, 1.24% stain for the study duration, unless they had used at plasma 11-OH-THC concentrations of 2.17, 21.70 and cannabis or CB-based medicine within 30 days prior to 86.80 ng/mL, respectively. receiving study medication. Subjects with an abnormal diet, who had made substantial changes to eating habits Extraction procedure in the 30 day period prior to the study, or who had par- A 0.5 mL aliquot of test sample, QC or blank plasma ticipated in another clinical trial in the 90 day period was placed into a test tube. The blank plasma was spiked prior to study entry were also excluded. Subjects who with 50 μL of the appropriate standard solution to pro- used any prescription or over the counter medication duce the calibration standards. 50 μL of the internal within 14 and seven days of study onset, or during the standard was added to each test tube. The samples were study, respectively, were also excluded, as were subjects then diluted by the addition of 500 μL of 0.1% (w/v) as- who had treatment with any known enzyme-altering corbic acid and the protein precipitated by the addition agents within 30 days prior to or during the study. In of 1.0 mL of acetonitrile. The proteins were removed by addition, subjects who had a postural drop of 20 mmHg centrifugation and the supernatant concentrated to 1 or more in systolic blood pressure at screening were ex- mL using nitrogen at 50ºC. The samples were then basi- cluded, as were subjects who had donated blood or fied by the addition of 300 μL of 5M sodium hydroxide plasma within 90 days of study initiation. Subjects with a and the analytes extracted by the addition of 2.0 mL of known history of hypersensitivity or idiosyncratic reac- 7:1 hexane:ethyl acetate. After mixing for 16 minutes the tion to the study drug or related compounds were also tubes were centrifuged and the top layer transferred into excluded. clean 3 dram vials. The solvent was removed using ni- trogen at 70ºC and the dried extract resuspended in 100 Concomitant medication μL of BSTFA and transferred to microvials, capped and If concomitant medication was taken during the study, a placed at 70ºC for 30 minutes to derivatise the analytes. joint decision was be made by the investigator and spon- The microvials were then cooled and loaded onto the sor if the subject should continue in the study. No sub- autosampler tray for analysis where 1 μL was injected ject was permitted to take medication during the time of onto the GC-MS system. sample collection. Inclusion and exclusion criteria Dietary restrictions Inclusion criteria Xanthines and alcohol were prohibited 48 hours prior to Eligible subjects were healthy males between 18 and 45 dosing days and throughout each period of sample col- years of age with a Body Mass Index (BMI) of between lection. Grapefruit was prohibited 10 days prior to initial 18 and 30 Kg/m . Subjects had no clinically significant dosing and throughout the study. Stott et al. SpringerPlus 2013, 2:236 Page 5 of 15 http://www.springerplus.com/content/2/1/236 Study endpoints was concluded if the 90% CI of the ratio of the (0-inf) Pharmacokinetic endpoints geometric means fell within the range of 0.80 to 1.25. The PK endpoints were mean peak plasma concentra- The summaries and descriptive statistics were calculated tion (C ), area under the plasma concentration versus using WinNonlin® Professional, version 4.1b and SAS®, max time curve (AUC), from time 0 to the last measurable version 9.1. concentration (AUC ), AUC to infinite time (AUC (0-t) (0- ), time to peak plasma concentration (T ), half-time inf) max Results (t ), elimination rate constant (Kel), oral clearance 1/2 Mean participant ages for sequences 1A, 1B, 2C, 2D, 3E, (CL/F) and apparent volume of distribution following and 3F were 28.8, 25.8, 32.5, 23.7, 26.5 and 27.0 years of oral administration (Varea/F; THC and CBD only) of age, respectively. Mean BMIs for the same respective se- THC, 11-OH-THC and CBD following administration of quences were 26.2, 24.2, 26.8, 25.2, 23.3 and 24.3 Kg/m , THC/CBD spray alone or THC/CBD spray concomi- giving a similar demographic profile across treatment tantly with rifampicin, ketoconazole, and omeprazole. groups. Safety endpoints Plasma concentrations and exposure The safety endpoints were blood pressure, heart rate, Mean plasma concentration versus time curves for ECG, clinical laboratory data (haematology and bio- THC, CBD and 11-OH-THC following administration of chemistry), urinalysis, AEs and concomitant medica- THC/CBD sprays alone and in combination with rifam- tions, recorded at each visit. picin, ketoconazole or omeprazole, are presented in Figures 1, 2 and 3, respectively. Statistical methods Based on group mean C and AUC, the plasma ex- max Sample size posure of THC, CBD and 11-OH-THC decreased for all A total of 36 subjects were planned and analyzed, with three analytes following a single dose (4 sprays) of THC/ 12 subjects in each treatment group. However, there was CBD administered at the end of a 10-day dosing period no formal sample size power calculation for this study. (repeated dosing over 10 days) with rifampicin, com- pared with the PK parameters following a single dose of PK parameters THC/CBD alone (4 sprays, no rifampicin). The decrease Summary statistics of PK parameters and concentrations was general, occurring in 82-100% of subjects (Table 1). included all treated subjects. Only subjects completing For the ketoconazole treatment group, mean C and max the study (i.e. PK data available for THC/CBD spray and AUC increased for all three analytes following a single THC/CBD spray plus interacting drug), were included in dose of THC/CBD (4 sprays) administered at the end of the statistical analyses of the interaction effects of rifam- a 5-day dosing period with ketoconazole compared with picin (Group 1), ketoconazole (Group 2), and omepra- the PK parameters following a single dose of THC/CBD zole (Group 3). Data from the 3 groups were analyzed alone (4 sprays, no ketoconazole). The increase was also separately and no comparisons were made between general, occurring in 63-100% of the subjects (Table 2). groups. For each group, the PK parameters C ,AUC max (0- Following single dose administration of THC/CBD and AUC were statistically analyzed using an t) (0-inf) spray (4 sprays) at the end of a 6-day dosing period with analysis of variance model (ANOVA, SAS PROC omeprazole, the plasma concentrations and PK parame- MIXED). The traditional two-period crossover design ters were similar for THC, marginally higher for CBD was implemented. The model included effects of treat- and marginally lower for 11-OH-THC compared with ment, period, sequence, and subject within sequence. the PK parameters following a single dose of THC/CBD The log-transformed AUC and C data was analyzed max alone (4 sprays, no omeprazole) (Table 3). using a general linear mixed model. The model included fixed terms for treatment, sequence, period and a ran- dom term for subject within sequence. Point estimates Oral clearance and 90% confidence intervals (CIs) for the ratios of the The mean CL/F of THC/CBD spray increased from treatment means were calculated. The two one-sided hy- 1207 L/h to 1595 L/h (+32%) for THC and from 2817 L/ potheses were tested at a 5% level for C ,AUC and h to 5966 L/h (+112%) for CBD after multiple dose ri- max (0-t) AUC by constructing 90% CIs for the ratio of the fampicin treatment (Table 1). Conversely, the mean CL/ (0-inf) treatment means. The 90% CIs were obtained from the F of THC/CBD spray decreased from 1504 L/h to 920 antilogarithms of the lower and upper bounds of the L/h (-39%) for THC and from 2998 L/h to 1731 L/h 90% CIs for the differences in the least-squares means of (-42%) for CBD when co-administered with ketocona- the log-transformed data. No significant interaction with zole (Table 2). No significant differences in CL/F for respect to the log-transformed C ,AUC and AUC THC or CBD were seen when THC/CBD spray was co- max (0-t) Stott et al. SpringerPlus 2013, 2:236 Page 6 of 15 http://www.springerplus.com/content/2/1/236 THC pre-Rifampicin THC post-Rifampicin 0 3 6 9 12 15 18 21 24 Time (h) 2.0 CBD pre-Rifampicin 1.5 CBD post-Rifampicin 1.0 0.5 0.0 0 3 6 9 12 15 18 21 24 Time (h) 11-OH-THC pre-Rifampicin 11-OH-THC post-Rifampicin 0 3 6 9 12 15 18 21 24 Time (h) Figure 1 Mean (+SD) plasma concentrations of THC (a), CBD (b) and 11-OH-THC (c) over time after administration of a single dose (4 sprays) of THC/CBD (n= 11) or THC/CBD spray in combination with multiple dose (2 x 300 mg) administration of Rifampicin (n = 12). Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Stott et al. SpringerPlus 2013, 2:236 Page 7 of 15 http://www.springerplus.com/content/2/1/236 THC pre-Ketoconazole THC post-Ketoconazole 0 3 6 9 12 15 18 21 24 Time (h) 2.0 1.5 CBD pre-Ketoconazole CBD post-Ketoconazole 1.0 0.5 0.0 0 3 6 9 12 15 18 21 24 Time (h) 11-OH-THC pre-Ketoconazole 11-OH-THC post-Ketoconazole 0 3 6 9 12 15 18 21 24 Time (h) Figure 2 Mean (+SD) plasma concentrations of THC (a), CBD (b) and 11-OH-THC (c) over time after administration of a single dose (4 sprays) of THC/CBD (n= 11) or THC/CBD spray in combination with multiple dose (2 x 200 mg) administration of Ketoconazole (n = 11). Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Stott et al. SpringerPlus 2013, 2:236 Page 8 of 15 http://www.springerplus.com/content/2/1/236 THC pre-Omeprazole THC post-Omeprazole 0 3 6 9 12 15 18 21 24 Time (h) 1.0 CBD pre-Omeprazole CBD post-Omeprazole 0.5 0.0 0 3 6 9 12 15 18 21 24 Time (h) 11-OH-THC pre-Omperazole 11-OH-THC post-omeprazole 0369 12 15 18 21 24 Time (h) Figure 3 Mean (+SD) plasma concentrations of THC (a), CBD (b) and 11-OH-THC (c) over time after administration of a single dose (4 sprays) of THC/CBD (n= 11) or THC/CBD spray in combination with multiple dose (2 x 20 mg) administration of Omeprazole (n = 12). Plasma concentrations (ng/mL) Plasma concentrations (ng/mL) Plasma concentrations (ng/mL) Stott et al. SpringerPlus 2013, 2:236 Page 9 of 15 http://www.springerplus.com/content/2/1/236 Table 1 Summary of pharmacokinetic parameters of THC, was no change in the t of THC or CBD, but a general 1/2 CBD and 11-OH-THC after a single dose of THC/CBD decrease for 11-OH-THC from 9.51 (THC/CBD spray (4 sprays) alone or in combination with rifampicin alone) to 7.48 h (THC/CBD spray and ketoconazole) Parameter THC/CBD spray alone THC/CBD spray and was observed (respective % changes for THC, CBD and (n= 12) rifampicin (n= 12) 11-OH-THC of +44%, -16% and -21%) (Table 2). When THC THC/CBD spray was co-administered with omeprazole, AUC (h*ng/mL) 9.10 (2.98) 6.53 (2.70) (0-t) the t and T for all analytes were similar to respect- 1/2 max AUC (h*ng/mL) 9.86 (3.35) 7.53 (2.99) ive THC/CBD spray alone values (respective % changes (0-inf) in t for THC, CBD and 11-OH-THC of -11%, +5% 1/2 C (ng/mL) 2.94 (1.21) 1.88 (1.07) max and +9%; respective % changes in T for THC, CBD max T (h) 1.01 (0.50-6.02) 1.75 (1.25-2.57) max and 11-OH-THC of 0%, +9% and -24%%) (Table 3). -1 a Kel (h ) 0.197 (0.093) 0.189 (0.071) t (h) 4.68 (3.42) 4.93 (3.91) Elimination rate constant CL/F (L/h) 1207 (373) 1595 (473) Kel was estimated over a short sampling period (less Varea/F (L) 7625 (4326) 10297 (5888) than two t s) for the majority of CBD and 11-OH-THC 1/2 profiles, and the regression had low precision (rsq ad- CBD justed <0.7) in a few profiles. Furthermore, AUC had (0-inf) AUC (h*ng/mL) 3.23 (2.13) 1.31 (0.89) (0-t) an extrapolated area larger than 20% for several CBD pro- AUC (h*ng/mL) 5.10 (3.06) 2.15 (0.94) (0-inf) files (Table 1). C (ng/mL) 1.03 (0.81) 0.50 (0.37) max T (h) 1.00 (0.50-4.00) 1.50 (1.00-6.00) max 90% confidence intervals -1 b Kel (h ) 0.148 (0.108) 0.196 (0.092) Following co-administration of THC/CBD spray with ri- fampicin or ketoconazole, the 90% CIs for the geometric t (h) 10.86 (12.71) 4.13 (1.65) mean ratios of C ,AUC and AUC did not fall max (0-t) (0-inf) CL/F (L/h) 2817 (1913) 5966 (3713) within the pre-defined “no interaction” range of 0.80 to Varea/F (L) 28312 (19355) 34790 (20036) 1.25 for any of the analytes (Table 4). 11-OH-THC Following co-administration of THC/CBD spray with AUC (h*ng/mL) 18.61 (7.81) 1.84 (0.79) (0-t) omeprazole, for THC and CBD the upper limit of the AUC (h*ng/mL) 21.59 (8.76) 2.78 (0.68) (0-inf) 90% CI of the geometric mean ratio of C was above max C (ng/mL) 3.38 (1.95) 0.45 (0.18) 1.25. The AUC and AUC 90% CI upper limits max (0-t) (0-inf) were above 1.25 and the lower limits were below 0.80 T (h) 1.38 (0.50-6.02) 1.75 (1.50-2.57) max for THC and CBD (Table 4). For 11-OH-THC the lower -1 c Kel (h ) 0.083 (0.042) 0.189 (0.074) limits of the 90% CI of C ,AUC and AUC max (0-t) (0-inf) t (h) 9.88 (3.89) 4.19 (1.62) were below 0.80, did not fall within the pre-defined “no Data presented are mean and (SD) except for T where median and range max interaction” range of 0.80 to 1.25 (Table 4). a b c are shown, n=11; n=10; n=11. Inter-subject variability administered with omeprazole compared to THC/CBD The C and AUC % Co-efficient of Variation (CV%) max spray alone (Table 3). ranges were similar for all analytes in all treatment groups with the exception of a higher CV% for THC T and t½ when THC/CBD spray was administered alone in the max A small increase in median T for all analytes was ob- THC/CBD spray alone or in combination with omepra- max served when THC/CBD spray was co-administered with zole treatment group. rifampicin (respective % changes for THC, CBD and THC/CBD spray in combination with rifampicin had a 11-OH-THC of +73%, +50% and +27%) (Table 1). There CV% of 25-74% and for THC/CBD spray alone the CV% was no change in the mean t of THC, but a general was 33-79% for all analytes. THC/CBD spray in combin- 1/2 decrease for CBD and 11-OH-THC was observed (re- ation with ketoconazole had a CV% of 34-87% and for spective % changes for THC, CBD and 11-OH-THC of THC/CBD spray alone this was 23-69% for all analytes. +5%, -62% and -58%) (Table 1). When THC/CBD spray THC/CBD spray in combination with omeprazole had a was co-administered with ketoconazole, there was a CV% range of 40-67% for combination treatment and 48- small increase in mean T for 11-OH-THC and CBD, 69% for THC/CBD spray alone for CBD and 11-OH-THC. max but no trend was observed for THC when looking at in- The CV% range for THC was higher at 74-98% for THC/ dividual data (respective % changes for THC, CBD and CBD spray alone compared to 43-64% when administered 11-OH-THC of +17%, +27% and +50%) (Table 2). There in combination with omeprazole. Stott et al. SpringerPlus 2013, 2:236 Page 10 of 15 http://www.springerplus.com/content/2/1/236 Table 2 Summary of pharmacokinetic parameters of THC, TEAEs reported by subjects receiving THC/CBD spray CBD and 11-OH-THC after a single dose of THC/CBD plus omeprazole were also nervous system disorders, the (4 sprays) alone or in combination with ketoconazole most common being dizziness. Parameter THC/CBD spray THC/CBD spray and No other clinically significant abnormalities for labora- alone (n= 12) ketoconazole (n= 11) tory safety measurements or ECG parameters were THC reported during the study. AUC (h*ng/mL) 8.19 (5.67) 15.38 (13.43) (0-t) AUC (h*ng/mL) 9.22 (5.94) 16.76 (13.80) Discussion (0-inf) This study investigated the effects of the known CYP450 C (ng/mL) 2.65 (1.32) 3.36 (1.65) max inhibitors or inducers rifampicin, ketoconazole and omep- T (h) 1.50 (0.75-6.00) 1.75 (1.00-3.00) max razole, on the PK and safety profiles of THC/CBD spray. -1 a Kel (h ) 0.258 (0.092) 0.189 (0.078) t (h) 3.07 (1.31) 4.43 (2.19) Rifampicin and THC/CBD spray CL/F (L/h) 1504 (688) 920 (450) When prescribing drugs that share the CYP3A4 path- Varea/F (L) 6328 (4164) 5111 (2221) way, plasma levels should be periodically monitored, otherwise it is possible that drug levels may reach a toxic CBD state that can manifest as serious medical events if one AUC (h*ng/mL) 1.82 (1.03) 4.83 (2.01) (0-t) of them is a CYP3A4 inhibitor (such as ketoconazole) b c AUC (h*ng/mL) 3.54 (0.80) 6.50 (2.23) (0-inf) (Ogu & Maxa 2000). If one of the drugs is a CYP3A4 in- C (ng/mL) 0.66 (0.37) 1.25 (0.51) max ducer, such as rifampicin, then the effectiveness of the T (h) 1.38 (0.75-6.00) 1.75 (1.00-2.52) max other drug might be compromised following a more -1 b c Kel (h ) 0.122 (0.111) 0.143 (0.066) rapid reduction than usual of the plasma level. As such, b c investigating the PKs of THC/CBD spray in combination t (h) 7.81 (3.00) 6.54 (4.59) b c with a CYP3A4 inducer was of high clinical importance. CL/F (L/h) 2998 (896) 1731 (650) Rifampicin was chosen as it has been extensively used in b c Varea/F (L) 31994 (12794) 14349 (7076) clinical studies as a prototypical inducer of CYP3A4 11-OH-THC (Division of Clinical Pharmacology 2012), but it is also a AUC (h*ng/mL) 21.78 (11.34) 84.34 (40.18) (0-t) moderate CYP2C19 inducer (Federal Drug Association AUC (h*ng/mL) 27.13 (13.34) 95.26 (48.93) (0-inf) 2012). Overall exposure to repeated daily doses of rifam- C (ng/mL) 3.59 (1.67) 10.92 (3.83) picin for 10 days, followed by a single dose of THC/CBD max spray (4 sprays), reduced the mean plasma levels of T (h) 1.50 (1.00-6.00) 2.25 (1.50-4.13) max THC and CBD, and most of all, 11-OH-THC, as com- -1 d Kel (h ) 0.076 (0.014) 0.095 (0.016) pared with levels observed when THC/CBD spray alone t (h) 9.51 (2.18) 7.48 (1.39) was administered. While the magnitude of the reduction Data presented are mean and (SD) except for T where median and range max in the plasma levels of the three analytes was within the a b c d are shown, n=11; n=9; n=10; n=11. range of inter-individual variation observed with THC/ CBD spray alone, suggesting that significant induction of Safety and tolerability CYP3A4 would lead to only a slight reduction in expos- A summary of all treatment-emergent AEs (TEAEs) oc- ure to CBs, intra-subject variability must also be consid- curring in one or more subject is presented in Table 5. ered. For example, should a subjects whose symptoms With the exception of the THC/CBD spray plus ketoco- are controlled by a daily dose of seven sprays of THC/ nazole study group, all study medication was generally CBD spray experience a reduction in plasma levels of well tolerated by subjects. CBs of 1/3 in the presence of a CYP3A4 inducer, they The most common TEAEs with THC/CBD spray alone would need to increase their dose to experience the were somnolence and headache. For THC/CBD spray same efficacy of THC/CBD spray. The effect of rifampi- plus rifampicin, these were rhinitis, headache and mal- cin on the group mean C and AUC of CBD and THC max aise; but none was considered related to THC/CBD was consistent with the inducing effects of rifampicin on spray, all were of mild severity, and none was reported the CYP3A4 isoenzyme (Michalets 1998). Both CBD and with a subject incidence greater than one. Subjects re- THC exposure decreased when rifampicin was adminis- ceiving THC/CBD spray plus ketoconazole reported the tered with THC/CBD spray and the apparent clearance greatest subject incidence (100%) of TEAEs in the study, for these analytes increased after multiple dose adminis- with a total of 35 TEAEs reported in 11 subjects. The tration of rifampicin. Although an increase in exposure majority of these were of the system organ class (SOC) of the 11-OH-THC metabolite was expected due to an- nervous system disorders. Similarly, the majority of ticipated conversion of THC to 11-OH-THC via Stott et al. SpringerPlus 2013, 2:236 Page 11 of 15 http://www.springerplus.com/content/2/1/236 Table 3 Summary of pharmacokinetic parameters of THC, administration of ketoconazole. These increases in mean CBD and 11-OH-THC after a single dose of THC/CBD values were considerably less than the range of inter- (4 sprays) alone or in combination omeprazole subject variability for the same parameters. However, Parameter THC/CBD spray alone THC/CBD spray and intra-subject variability must also be considered, in that (n= 12) omeprazole (n= 12) should an individual experienced enhanced plasma levels THC of CBs in the presence of a CYP3A4 inhibitor, then their AUC (h*ng/mL) 8.76 (8.62) 7.41 (4.75) (0-t) daily dose of THC/CBD spray would need to be reduced accordingly in order to balance the efficacy of the com- AUC (h*ng/mL) 9.39 (8.81) 8.10 (4.78) (0-inf) pound against any adverse effects that may occur at C (ng/mL) 2.50 (1.85) 2.48 (1.06) max higher doses. T (h) 1.25 (0.77-3.02) 1.25 (1.00-1.75) max For 11-OH-THC, the C was 3.1-times higher after max -1 Kel (h ) 0.305 (0.109) 0.357 (0.218) CYP3A4 inhibition by ketoconazole, and the AUC was t (h) 2.65 (1.25) 2.37 (0.92) 3.8-times higher, confirming that 11-OH-THC is likely CL/F (L/h) 2161 (1990) 2284 (2520) to be metabolized predominantly by CYP3A4. The effect of ketoconazole on the group mean C and AUC of Varea/F (L) 6889 (5296) 6052 (3384) max CBD and THC was consistent with the inhibitory effects CBD of ketoconazole on the CYP3A4 isoenzyme. Both CBD AUC (h*ng/mL) 1.83 (1.19) 2.25 (1.51) (0-t) and THC exposure increased when THC/CBD spray AUC (h*ng/mL) 3.00 (1.43) 3.33 (1.77) (0-inf) was administered with ketoconazole and the apparent C (ng/mL) 0.63 (0.43) 0.73 (0.30) max clearance for CBD and THC decreased after multiple T (h) 1.15 (0.50-3.02) 1.25 (0.48-1.75) max dose administration of ketoconazole. Although a de- -1 a Kel (h ) 0.224 (0.158) 0.210 (0.114) crease in exposure of the 11-OH-THC metabolite was expected due to inhibition of the conversion of THC to t (h) 5.22 (4.51) 5.46 (6.13) a 11-OH-THC via CYP3A4, a significant increase in me- CL/F (L/h) 4741 (3845) 4772 (4550) tabolite exposure was observed which again suggests Varea/F (L) 26298 (14532) 24757 (16311) CYP3A4 is involved in the metabolism of the metabolite, 11-OH-THC 11-OH-THC. AUC (h*ng/mL) 21.52 (14.76) 17.69 (9.05) (0-t) AUC (h*ng/mL) 24.17 (16.47) 19.80 (9.74) (0-inf) Omeprazole and THC/CBD spray To investigate the potential interaction of THC/CBD C (ng/mL) 3.48 (2.27) 2.84 (1.40) max spray with a CYP2C19 inhibitor, omeprazole was emplo- T (h) 2.00 (1.25-3.02) 1.52 (1.23-2.50) max yed. In contrast to rifampicin and ketoconazole, co- -1 Kel (h ) 0.114 (0.062) 0.095 (0.029) administration of THC/CBD spray with omeprazole had t (h) 7.30 (2.87) 7.98 (2.82) no apparent affect the group mean C or AUC results max Data presented are mean and (SD) except for T where median and range max for any of the analytes. Any slight differences group are shown, n=11. mean AUC and C values for all three analytes were max non-significant and well within the range of inter- CYP3A4, instead a significant decrease in the metabolite individual variation observed with THC/CBD spray exposure was observed, suggesting that CYP3A4 is also alone. Furthermore, the apparent clearance for THC and involved in the further metabolism of the primary me- CBD was not changed between THC/CBD spray alone tabolite, 11-OH-THC. A suggested mechanism for this versus THC/CBD spray and omeprazole treatments. effect is that increased CYP3A4 activity induced by ri- However, a decrease in exposure to 11-OH-THC was fampicin induced further metabolism of 11-OH-THC, observed following co-administration of omeprazole causing a reduced plasma level of 11-OH-THC. This is with THC/CBD spray. Nevertheless, examining the indi- the first investigation in humans to identify CYP3A4 as a vidual exposure data, no clear conclusion could be significant mediator of 11-OH-THC metabolism, an inter- drawn with respect to any effect of omeprazole. These esting finding for this psychoactive THC metabolite. findings suggest that THC, CBD are not substrates for the CYP2C19 isoenzyme. Ketoconazole and THC/CBD spray Similarly, the effects of the potent CYP3A4 inhibitor ke- Safety and tolerability toconazole on the PKs of THC/CBD spray were also in- THC/CBD spray was generally well tolerated when given vestigated. For both THC and CBD, there was an alone, illustrated by the fact that there were no serious increase in mean C and AUC following THC/CBD AEs during the study. The majority of TEAEs were mild max spray administration after 5-day period of daily in severity and, as expected, of the SOC of nervous Stott et al. SpringerPlus 2013, 2:236 Page 12 of 15 http://www.springerplus.com/content/2/1/236 Table 4 Point estimate and 90% confidence interval for the geometric mean ratios of C , AUC and AUC for max (0-t) (0-inf) THC/CBD spray in combination with rifampicin, ketoconazole and omeprazole 90% confidence limits Analyte/PK variable Number of subjects Estimate Lower Upper Rifampicin and THC/CBD spray THC/C 12 0.614 0.525 0.719 max THC/AUC 12 0.711 0.617 0.820 (0-t) THC/AUC 11 0.761 0.667 0.868 (0-inf) CBD/C 12 0.480 0.420 0.550 max CBD/AUC 12 0.381 0.274 0.529 (0-t) CBD/AUC 10 0.422 0.265 0.673 (0-inf) 11-OH-THC/C 12 0.140 0.120 0.163 max 11-OH-THC/AUC 12 0.099 0.088 0.113 (0-t) 11-OH-THC/AUC 11 0.131 0.115 0.150 (0-inf) Ketoconazole and THC/CBD spray THC/C 11 1.252 1.043 1.503 max THC/AUC 11 1.770 1.385 2.263 (0-t) THC/AUC 10 1.840 1.446 2.342 (0-inf) CBD/C 11 1.961 1.497 2.569 max CBD/AUC 11 2.715 2.047 3.601 (0-t) CBD/AUC 9 1.923 1.560 2.370 (0-inf) 11-OH-THC/C 11 3.074 2.705 3.493 max 11-OH-THC/AUC 11 3.823 3.405 4.292 (0-t) 11-OH-THC/AUC 10 3.616 3.181 4.111 (0-inf) Omeprazole and THC/CBD spray THC/C 12 1.125 0.842 1.501 max THC/AUC 12 0.964 0.643 1.445 (0-t) THC/AUC 12 0.962 0.668 1.386 (0-inf) CBD/C 12 1.320 0.938 1.859 max CBD/AUC 12 1.326 0.756 2.325 (0-t) CBD/AUC 11 1.096 0.745 1.611 (0-inf) 11-OH-THC/C 12 0.866 0.700 1.073 max 11-OH-THC/AUC 12 0.869 0.629 1.201 (0-t) 11-OH-THC/AUC 12 0.869 0.641 1.179 (0-inf) system disorders, with somnolence being the most reported AEs (incidence >10%) were dizziness (28%), commonly reported. While the exposure to THC/CBD diarrhoea (13%), fatigue (11%) and nausea (11%) spray was lower than usual in chronic patients, the (Constantinescu & Sarantis 2006). Recent publications administration procedure did not follow the slow up- also demonstrate that AEs the SOC of nervous systems titration process that takes 4-5 days to reach a dose disorders were among the commonly occurring AEs of 4 sprays/day (and goes on for 7-8 days to reach reported with THC/CBD spray use (Johnson et al. 2012; the average 6-7 sprays/day dose, and up to 14 days to Langford et al. 2013). reach the maximum dose of 12 sprays/day). As such, No difference was observed in the proportions of sub- the incidence of AEs in the SOC of nervous disorders jects reporting AEs upon co-administration of THC/ in the current study is not surprising. A review of CBD spray with rifampicin or omeprazole compared collated results from initial randomized-controlled with THC/CBD spray alone. Again, all AEs were of mild clinical trials with THC/CBD spray involving 930 severity, with headache and dysguesia being the most patients demonstrated that the most frequently commonly reported for THC/CBD spray and rifampicin Stott et al. SpringerPlus 2013, 2:236 Page 13 of 15 http://www.springerplus.com/content/2/1/236 Table 5 Treatment emergent adverse events with an incidence >1 Primary system organ class Preferred term* No. of subjects (%) No. of subjects (%) No. of subjects (%) THC/CBD spray (n = 36) Psychiatric disorders Euphoric mood 2 (6) - - Nervous system disorders Dizziness 2 (6) - - Dizziness postural 3 (8) - - Headache 5 (14) - - Somnolence 11 (31) - - Gastrointestinal disorders Nausea 2 (6) - - Rifampicin (n = 12) THC/CBD spray and Rifampicin (n = 12) THC/CBD spray (n = 12) Nervous system disorders Dysgeusia 1 (8) 0 1 (8) Headache 0 1 (8) 1 (8) Somnolence 0 0 2 (17) Ketoconazole (n = 12) THC/CBD spray and Ketoconazole (n = 11) THC/CBD spray (n = 12) Nervous system disorders Somnolence 0 4 (36) 4 (33) Dizziness 0 3 (27) 2 (17) Lethargy 0 3 (27) 0 Dysgeusia 0 2 (18) 0 Headache 0 2 (18) 0 Somnolence 0 1 (9) 1 (8) Psychiatric disorders Euphoric mood 0 7 (64) 0 Anxiety 0 1 (9) 1 (8) Disorientation 0 1 (9) 1 (8) Omeprazole (n = 12) THC/CBD spray and Omeprazole (n = 12) THC/CBD spray (n = 12) Nervous system disorders Dizziness 0 3 (25) 0 Dizziness postural 0 0 2 (17) Headache 1 (8) 0 4 (33) Somnolence 0 2 (17) 5 (42) Gastrointestinal disorders Dry mouth 0 2 (17) 1 (8) *MedDRA version 10.0. (one subject), and dizziness in three subjects taking spray alone. However, there was an increased incidence THC/CBD spray and omeprazole. of euphoric mood, lethargy, dygeusia and headache Although all subjects receiving THC/CBD spray and when THC/CBD spray was given in combination with ketoconazole experienced AEs, this combination was still ketoconazole. Only one event was classed as moderate relatively well tolerated, and all but one AE was of mild in terms of severity (anxiety) and the event resolved severity. The majority of TEAEs were classed as nervous without intervention. All other events were of mild system disorders, including somnolence and dizziness severity. Notably in this group, all subjects had which occurred at the same incidence as with THC/CBD increases in C for 11-OH-THC, and seven had an max Stott et al. SpringerPlus 2013, 2:236 Page 14 of 15 http://www.springerplus.com/content/2/1/236 increase in THC C , with nine subjects also having Conclusions max an increase in AUC. Taking into account the increase In conclusion, inhibition of CYP2C19 by omeprazole did in exposure after combined administration compared not significantly alter the PK of THC/CBD spray to THC/CBD spray alone, it is possible that the difference suggesting that THC, CBD and 11-OH-THC are not in PKs may account for the increase in CNS-type substrates for this isoenzyme at the clinically relevant AEs observed. dose of THC/CBD spray investigated. The CYP3A4 in- ducer rifampicin caused a decrease in exposure to all Study limitations three analytes, although not extreme and within the nat- There were a number of limitations to this study. Kel ural range of variation between subjects. Conversely, the was estimated over a short sampling period (less than CYP3A4 inhibitor ketoconazole caused increased expos- two t s) for the majority of profiles in this study and ure to all analytes, suggesting that THC, CBD and 1/2 the regression had low precision in some profiles. AUC 11-OH-THC all are substrates for this isoenzyme. More- had an extrapolated area which was larger than over, the findings with rifampicin and ketoconazole sug- (0-inf) 20% for some profiles. However, despite these limita- gest that CYP3A4 is involved in the metabolism of tions, the AUC and the AUC were in agreement, 11-OH-THC. On the basis of these findings, there is (0-inf) (0-t) and the statistical results are thus considered reliable. likely to be little impact on other drugs metabolized by Further consistency and validity of the study is demon- CYP enzymes on the PK parameters of THC/CBD spray, strated by the similarity of the PK exposure parameters but potential effects should be taken into consideration following a single dose of THC/CBD spray in the ab- when co-administering THC/CBD spray with com- sence of any inducer or inhibitor, which are in good pounds which share the CYP3A4 pathway. THC/CBD agreement with the PK data provided in a previous clin- spray was also well tolerated in healthy subjects both ical study performed by the authors (ref PK study). The alone and in combination with rifampicin, ketoconazole inter-subject variability was substantial and greater than and omeprazole. the difference in exposure means before and after rifampi- cin, ketoconazole, or omeprazole treatment, suggesting Ethical standards that any effect of other medications metabolized by rele- The current study was approved by Guy's Research Eth- vant CYP enzymes on THC/CBD spray is likely to be ics Committee, and was conducted in accordance with within the normal range of variation. However, there was the International Conference on Harmonisation guide- generally a similar variability in the group mean primary lines on Good Clinical Practice and the ethical principles PK parameters between the THC/CBD spray alone and stated in the Declaration of Helsinki and local UK regu- the THC/CBD spray plus inducer or inhibitor treatments. lations. All participants gave written informed consent. Doses of THC/CBD (4 sprays instead the average 6-8 † Sativex®, a THC/CBD oromucosal spray, does not sprays/day) and intake patterns (4 sprays in a row in- have an INN. Nabiximols is the US Adopted Name stead of evenly distributed through the day) were not (USAN). equivalent to those in the approved label. However, dur- ing a phase IIb dose ranging study in cancer patients Competing interests with pain, in the low dose THC/CBD spray (1-4 sprays) GW Pharmaceutical Ltd produces THC/CBD spray and is licensed for the treatment of spasticity in multiple sclerosis in several European countries, as group, over 90% of patients titrated to a dose of 3 or 4 well as Canada and New Zealand. This study was funded by GW sprays, leading to the conclusion that a minimal effective Pharmaceuticals Ltd. Darren Wilbraham is an employee of Quintiles Ltd who dose was 3 sprays per day (Porteney et al. 2012). were contracted to perform the clinical study. Colin Stott, Linda White, Stephen Wright and Geoffrey Guy are all employees of GW Pharmaceuticals Additionally, efficacy was observed in the low dose Ltd, and are shareholders in the company. group, reaching statistical significance for the continuous response analysis (pain 0-10 Numerical Rating Scale Authors’ contributions score) and for the mean change from baseline in score, CGS, LW, SW and GWG made a substantial contribution to the conception demonstrating that 4 daily sprays is a clinically relevant and design of the study. DW made a substantial contribution to the acquisition of the data, and CGS, LW, SW and GWG were involved in the dose (Porteney et al. 2012). Distributing the dose through- analysis of the data. All authors had a hand in the preparation of this out the day in the current study would have given a very manuscript based on their interpretation of the data, and reviewed, different PK profile, and would not have been a suitable approved and agreed upon the manuscript in its final format. approach for the current study. Additionally, during a Author details previous Phase I pharmacokinetics study, doses of 2 GW Pharma Ltd, Porton Down Science Park Salisbury, Wiltshire SP4 0JQ, UK. daily sprays of THC/CBD spray gave low C values Quintiles Drug Research Unit at Guy's Hospital, 6 Newcomen Street, max London, UK. (Stott et al. 2012). As such, a dose of 4 daily sprays was chosen to give good plasma concentration over Received: 26 February 2013 Accepted: 3 May 2013 time curves. Published: 24 May 2013 Stott et al. 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J Neurol 260(4):984–97 Michalets EL (1998) Update: clinically significant cytochrome P-450 drug Submit your manuscript to a interactions. Pharmacotherapy 18:84–112 journal and benefi t from: Miller JC, Miller JN (1992) Statistics for analytical Chemistry. Ellis Horwood Nurmikko TJ, Serpell MG, Hoggart B, Toomey PJ, Morlion BJ, Haines D (2007) 7 Convenient online submission Sativex successfully treats neuropathic pain characterised by allodynia: A 7 Rigorous peer review randomised, double-blind, placebo-controlled clinical trial. Pain 133:210–20 Ogu CC, Maxa JL (2000) Drug interactions due to cytochrome P450. Proc (Bayl 7 Immediate publication on acceptance Univ Med Cent) 13(4):421–3 7 Open access: articles freely available online Pertwee RG (2007) Cannabinoids and Multiple Sclerosis. Mol Neurobiol 36:45–59 7 High visibility within the fi eld Porteney R, Ganae-Motan ED, Allende S et al (2012) Nabiximols for opioid-treated 7 Retaining the copyright to your article cancer patients with poorly-controlled chronic pain: a randomized, placebo-controlled, graded-dose trial. J Pain 13(5):438–49 Submit your next manuscript at 7 springeropen.com http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png SpringerPlus Springer Journals

A Phase I, open-label, randomized, crossover study in three parallel groups to evaluate the effect of Rifampicin, Ketoconazole, and Omeprazole on the pharmacokinetics of THC/CBD oromucosal spray in healthy volunteers

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Copyright © 2013 by Stott et al.; licensee Springer.
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Abstract

This Phase I study aimed to assess the potential drug-drug interactions (pharmacokinetic [PK] and safety profile) of Δ9-tetrahydrocannabinol (THC)/cannabidiol (CBD) oromucosal spray (Sativex®, nabiximols) in combination with cytochrome P450 (CYP450) inducer (rifampicin) or inhibitors (ketoconazole or omeprazole). Thirty-six healthy male subjects were divided into three groups of 12, and then randomized to one of two treatment sequences per group. Subjects received four sprays of THC/CBD (10.8/10 mg) alongside single doses of the CYP3A and 2C19 inducer rifampicin (600 mg), CYP3A inhibitor ketoconazole (400 mg) or CYP2C19 inhibitor omeprazole (40 mg). Plasma samples were analyzed for CBD, THC and its metabolite 11-hydroxy-THC (11-OH-THC). A single dose of four sprays of THC/CBD spray (10.8/10 mg) following repeated doses of rifampicin (600 mg) reduced the C and AUC of all analytes. C reduced from 2.94 to 1.88 ng/mL (-36%), 1.03 to 0.50 ng/mL (-52%) max max and 3.38 to 0.45 ng/mL (-87%) for THC, CBD and 11-OH-THC, respectively compared to single dose administration of THC/CBD spray alone. Ketoconazole co-administration with THC/CBD spray had the opposite effect, increasing the C of the respective analytes from 2.65 to 3.36 ng/mL (+27%), 0.66 to 1.25 ng/mL (+89%) and 3.59 to 10.92 max ng/mL (+204%). No significant deviations in C or AUC for any analyte were observed when THC/CBD spray was max co-administered with omeprazole. THC/CBD spray was well tolerated by the study subjects both alone and in combination with rifampicin, ketoconazole and omeprazole. Evaluation of the PKs of THC/CBD spray alone and in combination with CYP450 inhibitors/inducers suggests that all analytes are substrates for the isoenzyme CYP3A4, but not CYP2C19. On the basis of our findings, there is likely to be little impact on other drugs metabolized by CYP enzymes on the PK parameters of THC/CBD spray, but potential effects should be taken into consideration when co-administering THC/CBD spray with compounds which share the CYP3A4 pathway such as rifampicin or ketoconazole. Trials registration: NCT01323465 † † Keywords: Cannabidiol, Cytochrome P450, Delta-9-tetrahydrocannabinol, Sativex® , Nabiximols , THC/CBD spray * Correspondence: cgs@gwpharm.com GW Pharma Ltd, Porton Down Science Park Salisbury, Wiltshire SP4 0JQ, UK Full list of author information is available at the end of the article © 2013 Stott et al.; licensee Springer. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Stott et al. SpringerPlus 2013, 2:236 Page 2 of 15 http://www.springerplus.com/content/2/1/236 Introduction clinically significant interactions from occurring. More- The endocannabinoid system modulator Δ9-tetrahydro- over, avoiding co-administration or adjusting a patient's cannabinol (THC)/cannabidiol (CBD) oromucosal spray drug regimen early in the course of therapy can provide (Sativex®, nabiximols) has been reported to be effective optimal response with minimal AEs (Ogu & Maxa 2000). in relieving a number of multiple sclerosis (MS) symp- Many different CYP450 isoenzymes have been identified toms including spasticity, central neuropathic pain and to-date, including six which play important roles in drug bladder dysfunction (Rog et al. 2005; Johnson et al. 2010; metabolism (DiPiro 1999; Cupp & Tracy 1998): CYP1A2, Rog et al. 2007), and has recently been approved in vari- CYP2C19, CYP2C9, CYP2D6, CYP2E1, and CYP3A4. ous European countries and abroad (i.e. in Canada, CYP450 inhibitors and inducers are known to affect Israel, New Zealand) as add-on treatment for spasticity the metabolism of THC. Previous literature reports have in MS patients. Other potential indications for this com- indicated that CBs, especially THC, are metabolized by pound include pain relief in advanced cancer (Johnson CYP3A4, 2C9, 2C19 and possibly 2D6 in humans et al. 2010; Porteney et al. 2012), as well as peripheral (Huestis 2007), and that the primary metabolites of neuropathic pain in MS (Nurmikko et al. 2007). Derived THC and CBD are 11-hydroxy-THC (11-OH-THC) and from proprietary cannabis plant varieties bred to exhibit 7-hydroxy-CBD, respectively (Huestis 2007). The forma- a pre-determined content of cannabinoids (CBs), THC/ tion of 11-OH-THC has been reported to be primarily CBD spray is fully standardized and contains two princi- catalysed by CYP2C19 and 2C9 (Bland et al. 2005). pal CBs, THC and CBD at an approximately 1:1 ratio as In vitro studies of THC and CBD on CYP450 induc- well as minor amounts of other CBs and non-CB com- tion and inhibition indicate that both inhibit CYP1A1, ponents. The specified CBs constitute at least 90% of the 1A2 and 1B1 enzymes (Yamaori et al. 2010). CBD also total CB content of the extract, however, the minor CBs has an inhibitory effect on CYP3A4 and CYP2C19. and other constituents also contribute to the therapeutic However, this effect only occurred at high concentra- profile of THC/CBD spray (Russo 2011), and may be in- tions (IC = 6-9 μM) of CBD (GW unpublished data), volved in stabilizing the extract (Whittle et al. 2001). and in normal dosing, peak plasma concentrations of CBs are thought to act primarily via activation of spe- CBD are approximately 5 ng/mL or less, 400-fold lower cific CB receptors, CB and CB (Howlett et al. 2002). than the levels at which CYP inhibition may be antici- 1 2 CB is predominantly expressed in the central nervous pated. As such, it is unlikely that THC/CBD spray would system (CNS), while CB is primarily expressed in the cause a relevant inhibition of CYP450s. However, to in- periphery, especially in immune cells (Pertwee 2007). vestigate the potential interactions of THC and CBD Endogenous ligands (“endocannabinoids”) produced in with drugs which also interact with the CYP450s mammalian tissues target these receptors, and together CYP3A4 and CYP2C19, various known inducers/inhi- with the catabolic and metabolic enzymes and transporter bitors of these isoenzymes were employed and the systems they constitute the endocannabinoid system. pharmacokinetics (PKs) of their co-administration with Multiple drug therapy is often used with a single pa- THC/CBD spray evaluated. tient. As THC/CBD spray is indicated for MS and po- Rifampicin is an antibiotic drug, a strong inducer of tentially advanced cancer pain, the likelihood is high CYP3A4 and moderate inducer of CYPs 2C19, 2B6, 2C8 that patients would be receiving different concomitant and 2C9, and has been extensively used in clinical stud- medications. As such, drug-drug interactions could ies as a prototypical inducer of these enzymes (Division occur which affect the bioavailability of THC/CBD spray of Clinical Pharmacology 2012; Federal Drug Association through absorption, metabolism or disposition. In turn 2012). Ketoconazole is a synthetic antifungal drug, is a this could affect the treatment and adverse events (AEs) strong inhibitor of CYP3A4 (Federal Drug Association experienced by the patient (Chen & Raymond 2006). In 2012), and a weak inhibitor of CYPs 2C8 and 2C19 some incidences, AEs experienced due to drug-drug inter- (Federal Drug Association 2012). Omeprazole is a actions can be life-threatening, therefore understanding proton-pump inhibitor which is primarily metabolized the mechanisms of these interactions is important so that by, and demonstrates high affinity for CYP2C19 (Furuta dosing and safety information can be adjusted accordingly. et al. 2005), and is also a moderate inhibitor of 2C19 Cytochrome P450 (CYP450) is a family of isoenzymes (Federal Drug Association 2012). This study investigated responsible for the biotransformation of several drugs, the potential interaction of these CYP450 inhibitors/in- and drug metabolism via this system has emerged as an ducers on the PK and safety profile of THC/CBD spray important determinant of the occurrence of several in healthy male subjects. drug-drug interactions that can result in toxicity, re- duced pharmacological effect and AEs (Guengerich Methods 2008). Determining whether the drugs involved act as This open-label, randomized, crossover, drug-interaction enzyme substrates, inducers, or inhibitors can prevent study took place at one study site in the UK (Quintiles Stott et al. SpringerPlus 2013, 2:236 Page 3 of 15 http://www.springerplus.com/content/2/1/236 Drug Research Unit at Guys Hospital), was approved by Research (Quintiles Limited) labels. The aliquots were Guy's Research Ethics Committee, and was conducted stored in clearly labelled containers in a freezer set at or according to the International Conference on Harmon- below -20°C, until shipped for assay. Samples were isation guidelines on Good Clinical Practice and the eth- shipped on dry ice at the appropriate time-points. ical principles stated in the Declaration of Helsinki and local UK regulations. All participants gave written in- Analysis method formed consent. The assay validation was undertaken by Advanced Bioanalytical Service Laboratories (London, UK), who Study design and treatment groups developed the technique after reviewing the literature, A total of 36 healthy males subjects enrolled and were which was based on the methodology adopted by three divided into three groups of 12. Within each group par- different groups (Foltz et al. 1983; Goodall & Basteyns ticipants were randomized to one of two treatment se- 1995; Kemp et al. 1995), and designed with reference to quences with six subjects receiving each sequence. FDA guidelines for industry (FDA Guidance for Industry Subjects received four sprays of THC/CBD spray (10.8/ 2012). The method utilised protein precipitation, solvent 10 mg) alongside single usual daily doses of either rifam- extraction and derivatisation for the sample preparation picin (600 mg), ketoconazole (400 mg) or omeprazole and then sample analysis by capillary gas chromatog- (40 mg) according to the following sequences, designed raphy and detection by a mass spectrometer (GC-MS). with a time-frame that was standard and fitting to the The validation procedure investigated the calibration aims of this study: model with the best regression fit over the concentration range 0.1 - 100 ng/mL for CBD, THC and 11-OH-THC, Sequence 1A. Subjects received a single dose of 4 as well as precision and accuracy of the method, stabil- sprays THC/CBD on Day 1 and once daily ity, carry-over, and specificity. rifampicin on Days 2-10. Subjects then received Human plasma from healthy volunteers was used to both THC/CBD spray and rifampicin on Day 11. prepare the standards and quality control (QC) samples, Sequence 1B. Subjects received rifampicin on Days 1-9. with analytes extracted using hexane/ethyl acetate (7:1 Subjects received THC/CBD spray and rifampicin ratio), derivatised with N,O-Bis(trimethylsilyl)trifluoroa- on Day 10 and then a single dose of 4 sprays THC/ cetamide. Analytic grade THC (Sigma, UK), CBD CBD on Day 18. (Sigma, UK) and 11-OH-THC (Radian International and Sequence 2C. Subjects received a single dose of 4 Cerillant, UK) were obtained, and three sets of CBD, sprays THC/CBD on Day 1 and once daily THC and 11-OH-THC were used to support the study. ketoconazole on Days 2-5. Subjects then received Deuterated THC-d (Sigma, UK) was used as the in- both THC/CBD spray and ketoconazole on Day 6. ternal standard. The GC-MS equipment was a Hewlett Sequence 2D. Subjects received ketoconazole on Days Packard 6890 Gas Chromatograph attached to a Hewlett 1-4. Subjects received THC/CBD spray and once Packard 5973 Mass Selective Detector. Data handling daily ketoconazole on Day 5. Subjects then received was carried out using an MS Chemstation System and a single dose of 4 sprays THC/CBD on Day 10. the peak area ratio of the analytes to the internal stand- Sequence 3E. Subjects received a single dose of 4 ard was calculated in Excel (2000). The concentrations sprays THC/CBD on Day 1. Subjects received once were calculated from the ratio data using least squares daily omeprazole on Days 2-6, and then THC/CBD ln(y) on ln(x) regression performed in Excel (2000), and spray and omeprazole on Day 7. were then checked manually. Regression analysis was Sequence 3F. Subjects received a once daily dose of undertaken to find the regression model that best de- omeprazole on Days 1-5. Subjects received both scribed the calibration data (for details, see (Miller & THC/CBD spray and omeprazole on Day 6. Subjects Miller 1992)). then received a single dose of 4 sprays THC/CBD Intra-assay precision and bias was examined using on Day 9. spiked control samples analysed in replicates of five. Inter-assay precision and accuracy were analysed in Blood sampling procedure and plasma preparation quintuplet at three concentrations and on three separate Blood samples were collected at specified times and occasions. stored on ice (except rifampicin PK samples which were The lowest and upper Limits of Quantification (LOQ) stored in iced water) prior to processing and storage. were investigated by looking at five (500 μL) plasma Plasma samples were separated by centrifugation (ap- samples containing 0.10 ng/mL and 100.0 ng/mL proximately 2500 rpm × 15 minutes at 4°C). Samples (the lowest and highest calibrators) of THC, CBD and were stored in 4 mL amber glass screw top glass vials 11-OH-THC, assayed in one batch, possessing accept- with PTFE lined screw caps labelled with Guys Drug able precision and accuracy. As such, these lowest and Stott et al. SpringerPlus 2013, 2:236 Page 4 of 15 http://www.springerplus.com/content/2/1/236 upper LOQ were deemed suitable for the measurement abnormal findings upon physical examination, 12-lead of these analytes in human plasma over these concentra- electrocardiogram (ECG), medical history, clinical la- tion ranges. boratory at screening, or renal and hepatic function. The inter- and intra-assay accuracy of the assay calcu- Subjects were non-users of tobacco products and were lated for THC was -0.53, -0.45, -1.72% and -0.50, -2.10, - negative for Human Immunodeficiency Viruses I and II, 0.86% at plasma THC concentrations of 2.0, 20.0 and Hepatitis B surface antigen, and antibodies to the Hepa- 80.0 ng/mL, respectively. The inter- and intra-assay pre- titis C virus. Eligible subjects had a negative urine screen cision of the assay calculated for THC was 1.88, 2.51, for alcohol, drugs of abuse (screening only) and cotinine, 2.41% and 2.64, 1.09, 1.29% at plasma THC concentra- and were using an appropriate barrier method of contra- tions of 2.0, 20.0 and 80.0 ng/mL, respectively. ception in addition to a second method of barrier The inter- and intra-assay accuracy of the assay calcu- contraception being used by their partner for the study lated for CBD was -2.90, 0.81, 1.78% and -2.00, -0.75, duration and for three months following administration 2.38% at plasma CBD concentrations of 2.0, 20.0 and of THC/CBD spray. 80.0 ng/mL, respectively. The inter- and intra-assay pre- cision of the assay calculated for CBD was 4.05, 2.28, Exclusion criteria 2.31% and 6.67, 1.43, 1.08% at plasma CBD concentra- Subjects with a history of significant cardiovascular, pul- tions of 2.0, 20.0 and 80.0 ng/mL, respectively. monary, hepatic, renal, haematologic, gastrointestinal, The inter- and intra-assay accuracy of the assay calcu- endocrine, immunologic, dermatologic, neurologic, or lated for 11-OH-THC was 0.00, 0.20, -3.44% and -0.46, - psychiatric disorder were excluded. Those with a history 0.05, -1.43% at plasma 11-OH-THC concentrations of of alcohol or drug abuse within two years of the study 2.17, 21.70 and 86.80 ng/mL, respectively. The inter- were also excluded; however, those with a history of pre- and intra-assay precision of the assay calculated for vious cannabis use were not excluded if willing to ab- 11-OH-THC was 3.45, 3.44, 2.57% and 6.04, 1.96, 1.24% stain for the study duration, unless they had used at plasma 11-OH-THC concentrations of 2.17, 21.70 and cannabis or CB-based medicine within 30 days prior to 86.80 ng/mL, respectively. receiving study medication. Subjects with an abnormal diet, who had made substantial changes to eating habits Extraction procedure in the 30 day period prior to the study, or who had par- A 0.5 mL aliquot of test sample, QC or blank plasma ticipated in another clinical trial in the 90 day period was placed into a test tube. The blank plasma was spiked prior to study entry were also excluded. Subjects who with 50 μL of the appropriate standard solution to pro- used any prescription or over the counter medication duce the calibration standards. 50 μL of the internal within 14 and seven days of study onset, or during the standard was added to each test tube. The samples were study, respectively, were also excluded, as were subjects then diluted by the addition of 500 μL of 0.1% (w/v) as- who had treatment with any known enzyme-altering corbic acid and the protein precipitated by the addition agents within 30 days prior to or during the study. In of 1.0 mL of acetonitrile. The proteins were removed by addition, subjects who had a postural drop of 20 mmHg centrifugation and the supernatant concentrated to 1 or more in systolic blood pressure at screening were ex- mL using nitrogen at 50ºC. The samples were then basi- cluded, as were subjects who had donated blood or fied by the addition of 300 μL of 5M sodium hydroxide plasma within 90 days of study initiation. Subjects with a and the analytes extracted by the addition of 2.0 mL of known history of hypersensitivity or idiosyncratic reac- 7:1 hexane:ethyl acetate. After mixing for 16 minutes the tion to the study drug or related compounds were also tubes were centrifuged and the top layer transferred into excluded. clean 3 dram vials. The solvent was removed using ni- trogen at 70ºC and the dried extract resuspended in 100 Concomitant medication μL of BSTFA and transferred to microvials, capped and If concomitant medication was taken during the study, a placed at 70ºC for 30 minutes to derivatise the analytes. joint decision was be made by the investigator and spon- The microvials were then cooled and loaded onto the sor if the subject should continue in the study. No sub- autosampler tray for analysis where 1 μL was injected ject was permitted to take medication during the time of onto the GC-MS system. sample collection. Inclusion and exclusion criteria Dietary restrictions Inclusion criteria Xanthines and alcohol were prohibited 48 hours prior to Eligible subjects were healthy males between 18 and 45 dosing days and throughout each period of sample col- years of age with a Body Mass Index (BMI) of between lection. Grapefruit was prohibited 10 days prior to initial 18 and 30 Kg/m . Subjects had no clinically significant dosing and throughout the study. Stott et al. SpringerPlus 2013, 2:236 Page 5 of 15 http://www.springerplus.com/content/2/1/236 Study endpoints was concluded if the 90% CI of the ratio of the (0-inf) Pharmacokinetic endpoints geometric means fell within the range of 0.80 to 1.25. The PK endpoints were mean peak plasma concentra- The summaries and descriptive statistics were calculated tion (C ), area under the plasma concentration versus using WinNonlin® Professional, version 4.1b and SAS®, max time curve (AUC), from time 0 to the last measurable version 9.1. concentration (AUC ), AUC to infinite time (AUC (0-t) (0- ), time to peak plasma concentration (T ), half-time inf) max Results (t ), elimination rate constant (Kel), oral clearance 1/2 Mean participant ages for sequences 1A, 1B, 2C, 2D, 3E, (CL/F) and apparent volume of distribution following and 3F were 28.8, 25.8, 32.5, 23.7, 26.5 and 27.0 years of oral administration (Varea/F; THC and CBD only) of age, respectively. Mean BMIs for the same respective se- THC, 11-OH-THC and CBD following administration of quences were 26.2, 24.2, 26.8, 25.2, 23.3 and 24.3 Kg/m , THC/CBD spray alone or THC/CBD spray concomi- giving a similar demographic profile across treatment tantly with rifampicin, ketoconazole, and omeprazole. groups. Safety endpoints Plasma concentrations and exposure The safety endpoints were blood pressure, heart rate, Mean plasma concentration versus time curves for ECG, clinical laboratory data (haematology and bio- THC, CBD and 11-OH-THC following administration of chemistry), urinalysis, AEs and concomitant medica- THC/CBD sprays alone and in combination with rifam- tions, recorded at each visit. picin, ketoconazole or omeprazole, are presented in Figures 1, 2 and 3, respectively. Statistical methods Based on group mean C and AUC, the plasma ex- max Sample size posure of THC, CBD and 11-OH-THC decreased for all A total of 36 subjects were planned and analyzed, with three analytes following a single dose (4 sprays) of THC/ 12 subjects in each treatment group. However, there was CBD administered at the end of a 10-day dosing period no formal sample size power calculation for this study. (repeated dosing over 10 days) with rifampicin, com- pared with the PK parameters following a single dose of PK parameters THC/CBD alone (4 sprays, no rifampicin). The decrease Summary statistics of PK parameters and concentrations was general, occurring in 82-100% of subjects (Table 1). included all treated subjects. Only subjects completing For the ketoconazole treatment group, mean C and max the study (i.e. PK data available for THC/CBD spray and AUC increased for all three analytes following a single THC/CBD spray plus interacting drug), were included in dose of THC/CBD (4 sprays) administered at the end of the statistical analyses of the interaction effects of rifam- a 5-day dosing period with ketoconazole compared with picin (Group 1), ketoconazole (Group 2), and omepra- the PK parameters following a single dose of THC/CBD zole (Group 3). Data from the 3 groups were analyzed alone (4 sprays, no ketoconazole). The increase was also separately and no comparisons were made between general, occurring in 63-100% of the subjects (Table 2). groups. For each group, the PK parameters C ,AUC max (0- Following single dose administration of THC/CBD and AUC were statistically analyzed using an t) (0-inf) spray (4 sprays) at the end of a 6-day dosing period with analysis of variance model (ANOVA, SAS PROC omeprazole, the plasma concentrations and PK parame- MIXED). The traditional two-period crossover design ters were similar for THC, marginally higher for CBD was implemented. The model included effects of treat- and marginally lower for 11-OH-THC compared with ment, period, sequence, and subject within sequence. the PK parameters following a single dose of THC/CBD The log-transformed AUC and C data was analyzed max alone (4 sprays, no omeprazole) (Table 3). using a general linear mixed model. The model included fixed terms for treatment, sequence, period and a ran- dom term for subject within sequence. Point estimates Oral clearance and 90% confidence intervals (CIs) for the ratios of the The mean CL/F of THC/CBD spray increased from treatment means were calculated. The two one-sided hy- 1207 L/h to 1595 L/h (+32%) for THC and from 2817 L/ potheses were tested at a 5% level for C ,AUC and h to 5966 L/h (+112%) for CBD after multiple dose ri- max (0-t) AUC by constructing 90% CIs for the ratio of the fampicin treatment (Table 1). Conversely, the mean CL/ (0-inf) treatment means. The 90% CIs were obtained from the F of THC/CBD spray decreased from 1504 L/h to 920 antilogarithms of the lower and upper bounds of the L/h (-39%) for THC and from 2998 L/h to 1731 L/h 90% CIs for the differences in the least-squares means of (-42%) for CBD when co-administered with ketocona- the log-transformed data. No significant interaction with zole (Table 2). No significant differences in CL/F for respect to the log-transformed C ,AUC and AUC THC or CBD were seen when THC/CBD spray was co- max (0-t) Stott et al. SpringerPlus 2013, 2:236 Page 6 of 15 http://www.springerplus.com/content/2/1/236 THC pre-Rifampicin THC post-Rifampicin 0 3 6 9 12 15 18 21 24 Time (h) 2.0 CBD pre-Rifampicin 1.5 CBD post-Rifampicin 1.0 0.5 0.0 0 3 6 9 12 15 18 21 24 Time (h) 11-OH-THC pre-Rifampicin 11-OH-THC post-Rifampicin 0 3 6 9 12 15 18 21 24 Time (h) Figure 1 Mean (+SD) plasma concentrations of THC (a), CBD (b) and 11-OH-THC (c) over time after administration of a single dose (4 sprays) of THC/CBD (n= 11) or THC/CBD spray in combination with multiple dose (2 x 300 mg) administration of Rifampicin (n = 12). Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Stott et al. SpringerPlus 2013, 2:236 Page 7 of 15 http://www.springerplus.com/content/2/1/236 THC pre-Ketoconazole THC post-Ketoconazole 0 3 6 9 12 15 18 21 24 Time (h) 2.0 1.5 CBD pre-Ketoconazole CBD post-Ketoconazole 1.0 0.5 0.0 0 3 6 9 12 15 18 21 24 Time (h) 11-OH-THC pre-Ketoconazole 11-OH-THC post-Ketoconazole 0 3 6 9 12 15 18 21 24 Time (h) Figure 2 Mean (+SD) plasma concentrations of THC (a), CBD (b) and 11-OH-THC (c) over time after administration of a single dose (4 sprays) of THC/CBD (n= 11) or THC/CBD spray in combination with multiple dose (2 x 200 mg) administration of Ketoconazole (n = 11). Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Plasma concetrations (ng/mL) Stott et al. SpringerPlus 2013, 2:236 Page 8 of 15 http://www.springerplus.com/content/2/1/236 THC pre-Omeprazole THC post-Omeprazole 0 3 6 9 12 15 18 21 24 Time (h) 1.0 CBD pre-Omeprazole CBD post-Omeprazole 0.5 0.0 0 3 6 9 12 15 18 21 24 Time (h) 11-OH-THC pre-Omperazole 11-OH-THC post-omeprazole 0369 12 15 18 21 24 Time (h) Figure 3 Mean (+SD) plasma concentrations of THC (a), CBD (b) and 11-OH-THC (c) over time after administration of a single dose (4 sprays) of THC/CBD (n= 11) or THC/CBD spray in combination with multiple dose (2 x 20 mg) administration of Omeprazole (n = 12). Plasma concentrations (ng/mL) Plasma concentrations (ng/mL) Plasma concentrations (ng/mL) Stott et al. SpringerPlus 2013, 2:236 Page 9 of 15 http://www.springerplus.com/content/2/1/236 Table 1 Summary of pharmacokinetic parameters of THC, was no change in the t of THC or CBD, but a general 1/2 CBD and 11-OH-THC after a single dose of THC/CBD decrease for 11-OH-THC from 9.51 (THC/CBD spray (4 sprays) alone or in combination with rifampicin alone) to 7.48 h (THC/CBD spray and ketoconazole) Parameter THC/CBD spray alone THC/CBD spray and was observed (respective % changes for THC, CBD and (n= 12) rifampicin (n= 12) 11-OH-THC of +44%, -16% and -21%) (Table 2). When THC THC/CBD spray was co-administered with omeprazole, AUC (h*ng/mL) 9.10 (2.98) 6.53 (2.70) (0-t) the t and T for all analytes were similar to respect- 1/2 max AUC (h*ng/mL) 9.86 (3.35) 7.53 (2.99) ive THC/CBD spray alone values (respective % changes (0-inf) in t for THC, CBD and 11-OH-THC of -11%, +5% 1/2 C (ng/mL) 2.94 (1.21) 1.88 (1.07) max and +9%; respective % changes in T for THC, CBD max T (h) 1.01 (0.50-6.02) 1.75 (1.25-2.57) max and 11-OH-THC of 0%, +9% and -24%%) (Table 3). -1 a Kel (h ) 0.197 (0.093) 0.189 (0.071) t (h) 4.68 (3.42) 4.93 (3.91) Elimination rate constant CL/F (L/h) 1207 (373) 1595 (473) Kel was estimated over a short sampling period (less Varea/F (L) 7625 (4326) 10297 (5888) than two t s) for the majority of CBD and 11-OH-THC 1/2 profiles, and the regression had low precision (rsq ad- CBD justed <0.7) in a few profiles. Furthermore, AUC had (0-inf) AUC (h*ng/mL) 3.23 (2.13) 1.31 (0.89) (0-t) an extrapolated area larger than 20% for several CBD pro- AUC (h*ng/mL) 5.10 (3.06) 2.15 (0.94) (0-inf) files (Table 1). C (ng/mL) 1.03 (0.81) 0.50 (0.37) max T (h) 1.00 (0.50-4.00) 1.50 (1.00-6.00) max 90% confidence intervals -1 b Kel (h ) 0.148 (0.108) 0.196 (0.092) Following co-administration of THC/CBD spray with ri- fampicin or ketoconazole, the 90% CIs for the geometric t (h) 10.86 (12.71) 4.13 (1.65) mean ratios of C ,AUC and AUC did not fall max (0-t) (0-inf) CL/F (L/h) 2817 (1913) 5966 (3713) within the pre-defined “no interaction” range of 0.80 to Varea/F (L) 28312 (19355) 34790 (20036) 1.25 for any of the analytes (Table 4). 11-OH-THC Following co-administration of THC/CBD spray with AUC (h*ng/mL) 18.61 (7.81) 1.84 (0.79) (0-t) omeprazole, for THC and CBD the upper limit of the AUC (h*ng/mL) 21.59 (8.76) 2.78 (0.68) (0-inf) 90% CI of the geometric mean ratio of C was above max C (ng/mL) 3.38 (1.95) 0.45 (0.18) 1.25. The AUC and AUC 90% CI upper limits max (0-t) (0-inf) were above 1.25 and the lower limits were below 0.80 T (h) 1.38 (0.50-6.02) 1.75 (1.50-2.57) max for THC and CBD (Table 4). For 11-OH-THC the lower -1 c Kel (h ) 0.083 (0.042) 0.189 (0.074) limits of the 90% CI of C ,AUC and AUC max (0-t) (0-inf) t (h) 9.88 (3.89) 4.19 (1.62) were below 0.80, did not fall within the pre-defined “no Data presented are mean and (SD) except for T where median and range max interaction” range of 0.80 to 1.25 (Table 4). a b c are shown, n=11; n=10; n=11. Inter-subject variability administered with omeprazole compared to THC/CBD The C and AUC % Co-efficient of Variation (CV%) max spray alone (Table 3). ranges were similar for all analytes in all treatment groups with the exception of a higher CV% for THC T and t½ when THC/CBD spray was administered alone in the max A small increase in median T for all analytes was ob- THC/CBD spray alone or in combination with omepra- max served when THC/CBD spray was co-administered with zole treatment group. rifampicin (respective % changes for THC, CBD and THC/CBD spray in combination with rifampicin had a 11-OH-THC of +73%, +50% and +27%) (Table 1). There CV% of 25-74% and for THC/CBD spray alone the CV% was no change in the mean t of THC, but a general was 33-79% for all analytes. THC/CBD spray in combin- 1/2 decrease for CBD and 11-OH-THC was observed (re- ation with ketoconazole had a CV% of 34-87% and for spective % changes for THC, CBD and 11-OH-THC of THC/CBD spray alone this was 23-69% for all analytes. +5%, -62% and -58%) (Table 1). When THC/CBD spray THC/CBD spray in combination with omeprazole had a was co-administered with ketoconazole, there was a CV% range of 40-67% for combination treatment and 48- small increase in mean T for 11-OH-THC and CBD, 69% for THC/CBD spray alone for CBD and 11-OH-THC. max but no trend was observed for THC when looking at in- The CV% range for THC was higher at 74-98% for THC/ dividual data (respective % changes for THC, CBD and CBD spray alone compared to 43-64% when administered 11-OH-THC of +17%, +27% and +50%) (Table 2). There in combination with omeprazole. Stott et al. SpringerPlus 2013, 2:236 Page 10 of 15 http://www.springerplus.com/content/2/1/236 Table 2 Summary of pharmacokinetic parameters of THC, TEAEs reported by subjects receiving THC/CBD spray CBD and 11-OH-THC after a single dose of THC/CBD plus omeprazole were also nervous system disorders, the (4 sprays) alone or in combination with ketoconazole most common being dizziness. Parameter THC/CBD spray THC/CBD spray and No other clinically significant abnormalities for labora- alone (n= 12) ketoconazole (n= 11) tory safety measurements or ECG parameters were THC reported during the study. AUC (h*ng/mL) 8.19 (5.67) 15.38 (13.43) (0-t) AUC (h*ng/mL) 9.22 (5.94) 16.76 (13.80) Discussion (0-inf) This study investigated the effects of the known CYP450 C (ng/mL) 2.65 (1.32) 3.36 (1.65) max inhibitors or inducers rifampicin, ketoconazole and omep- T (h) 1.50 (0.75-6.00) 1.75 (1.00-3.00) max razole, on the PK and safety profiles of THC/CBD spray. -1 a Kel (h ) 0.258 (0.092) 0.189 (0.078) t (h) 3.07 (1.31) 4.43 (2.19) Rifampicin and THC/CBD spray CL/F (L/h) 1504 (688) 920 (450) When prescribing drugs that share the CYP3A4 path- Varea/F (L) 6328 (4164) 5111 (2221) way, plasma levels should be periodically monitored, otherwise it is possible that drug levels may reach a toxic CBD state that can manifest as serious medical events if one AUC (h*ng/mL) 1.82 (1.03) 4.83 (2.01) (0-t) of them is a CYP3A4 inhibitor (such as ketoconazole) b c AUC (h*ng/mL) 3.54 (0.80) 6.50 (2.23) (0-inf) (Ogu & Maxa 2000). If one of the drugs is a CYP3A4 in- C (ng/mL) 0.66 (0.37) 1.25 (0.51) max ducer, such as rifampicin, then the effectiveness of the T (h) 1.38 (0.75-6.00) 1.75 (1.00-2.52) max other drug might be compromised following a more -1 b c Kel (h ) 0.122 (0.111) 0.143 (0.066) rapid reduction than usual of the plasma level. As such, b c investigating the PKs of THC/CBD spray in combination t (h) 7.81 (3.00) 6.54 (4.59) b c with a CYP3A4 inducer was of high clinical importance. CL/F (L/h) 2998 (896) 1731 (650) Rifampicin was chosen as it has been extensively used in b c Varea/F (L) 31994 (12794) 14349 (7076) clinical studies as a prototypical inducer of CYP3A4 11-OH-THC (Division of Clinical Pharmacology 2012), but it is also a AUC (h*ng/mL) 21.78 (11.34) 84.34 (40.18) (0-t) moderate CYP2C19 inducer (Federal Drug Association AUC (h*ng/mL) 27.13 (13.34) 95.26 (48.93) (0-inf) 2012). Overall exposure to repeated daily doses of rifam- C (ng/mL) 3.59 (1.67) 10.92 (3.83) picin for 10 days, followed by a single dose of THC/CBD max spray (4 sprays), reduced the mean plasma levels of T (h) 1.50 (1.00-6.00) 2.25 (1.50-4.13) max THC and CBD, and most of all, 11-OH-THC, as com- -1 d Kel (h ) 0.076 (0.014) 0.095 (0.016) pared with levels observed when THC/CBD spray alone t (h) 9.51 (2.18) 7.48 (1.39) was administered. While the magnitude of the reduction Data presented are mean and (SD) except for T where median and range max in the plasma levels of the three analytes was within the a b c d are shown, n=11; n=9; n=10; n=11. range of inter-individual variation observed with THC/ CBD spray alone, suggesting that significant induction of Safety and tolerability CYP3A4 would lead to only a slight reduction in expos- A summary of all treatment-emergent AEs (TEAEs) oc- ure to CBs, intra-subject variability must also be consid- curring in one or more subject is presented in Table 5. ered. For example, should a subjects whose symptoms With the exception of the THC/CBD spray plus ketoco- are controlled by a daily dose of seven sprays of THC/ nazole study group, all study medication was generally CBD spray experience a reduction in plasma levels of well tolerated by subjects. CBs of 1/3 in the presence of a CYP3A4 inducer, they The most common TEAEs with THC/CBD spray alone would need to increase their dose to experience the were somnolence and headache. For THC/CBD spray same efficacy of THC/CBD spray. The effect of rifampi- plus rifampicin, these were rhinitis, headache and mal- cin on the group mean C and AUC of CBD and THC max aise; but none was considered related to THC/CBD was consistent with the inducing effects of rifampicin on spray, all were of mild severity, and none was reported the CYP3A4 isoenzyme (Michalets 1998). Both CBD and with a subject incidence greater than one. Subjects re- THC exposure decreased when rifampicin was adminis- ceiving THC/CBD spray plus ketoconazole reported the tered with THC/CBD spray and the apparent clearance greatest subject incidence (100%) of TEAEs in the study, for these analytes increased after multiple dose adminis- with a total of 35 TEAEs reported in 11 subjects. The tration of rifampicin. Although an increase in exposure majority of these were of the system organ class (SOC) of the 11-OH-THC metabolite was expected due to an- nervous system disorders. Similarly, the majority of ticipated conversion of THC to 11-OH-THC via Stott et al. SpringerPlus 2013, 2:236 Page 11 of 15 http://www.springerplus.com/content/2/1/236 Table 3 Summary of pharmacokinetic parameters of THC, administration of ketoconazole. These increases in mean CBD and 11-OH-THC after a single dose of THC/CBD values were considerably less than the range of inter- (4 sprays) alone or in combination omeprazole subject variability for the same parameters. However, Parameter THC/CBD spray alone THC/CBD spray and intra-subject variability must also be considered, in that (n= 12) omeprazole (n= 12) should an individual experienced enhanced plasma levels THC of CBs in the presence of a CYP3A4 inhibitor, then their AUC (h*ng/mL) 8.76 (8.62) 7.41 (4.75) (0-t) daily dose of THC/CBD spray would need to be reduced accordingly in order to balance the efficacy of the com- AUC (h*ng/mL) 9.39 (8.81) 8.10 (4.78) (0-inf) pound against any adverse effects that may occur at C (ng/mL) 2.50 (1.85) 2.48 (1.06) max higher doses. T (h) 1.25 (0.77-3.02) 1.25 (1.00-1.75) max For 11-OH-THC, the C was 3.1-times higher after max -1 Kel (h ) 0.305 (0.109) 0.357 (0.218) CYP3A4 inhibition by ketoconazole, and the AUC was t (h) 2.65 (1.25) 2.37 (0.92) 3.8-times higher, confirming that 11-OH-THC is likely CL/F (L/h) 2161 (1990) 2284 (2520) to be metabolized predominantly by CYP3A4. The effect of ketoconazole on the group mean C and AUC of Varea/F (L) 6889 (5296) 6052 (3384) max CBD and THC was consistent with the inhibitory effects CBD of ketoconazole on the CYP3A4 isoenzyme. Both CBD AUC (h*ng/mL) 1.83 (1.19) 2.25 (1.51) (0-t) and THC exposure increased when THC/CBD spray AUC (h*ng/mL) 3.00 (1.43) 3.33 (1.77) (0-inf) was administered with ketoconazole and the apparent C (ng/mL) 0.63 (0.43) 0.73 (0.30) max clearance for CBD and THC decreased after multiple T (h) 1.15 (0.50-3.02) 1.25 (0.48-1.75) max dose administration of ketoconazole. Although a de- -1 a Kel (h ) 0.224 (0.158) 0.210 (0.114) crease in exposure of the 11-OH-THC metabolite was expected due to inhibition of the conversion of THC to t (h) 5.22 (4.51) 5.46 (6.13) a 11-OH-THC via CYP3A4, a significant increase in me- CL/F (L/h) 4741 (3845) 4772 (4550) tabolite exposure was observed which again suggests Varea/F (L) 26298 (14532) 24757 (16311) CYP3A4 is involved in the metabolism of the metabolite, 11-OH-THC 11-OH-THC. AUC (h*ng/mL) 21.52 (14.76) 17.69 (9.05) (0-t) AUC (h*ng/mL) 24.17 (16.47) 19.80 (9.74) (0-inf) Omeprazole and THC/CBD spray To investigate the potential interaction of THC/CBD C (ng/mL) 3.48 (2.27) 2.84 (1.40) max spray with a CYP2C19 inhibitor, omeprazole was emplo- T (h) 2.00 (1.25-3.02) 1.52 (1.23-2.50) max yed. In contrast to rifampicin and ketoconazole, co- -1 Kel (h ) 0.114 (0.062) 0.095 (0.029) administration of THC/CBD spray with omeprazole had t (h) 7.30 (2.87) 7.98 (2.82) no apparent affect the group mean C or AUC results max Data presented are mean and (SD) except for T where median and range max for any of the analytes. Any slight differences group are shown, n=11. mean AUC and C values for all three analytes were max non-significant and well within the range of inter- CYP3A4, instead a significant decrease in the metabolite individual variation observed with THC/CBD spray exposure was observed, suggesting that CYP3A4 is also alone. Furthermore, the apparent clearance for THC and involved in the further metabolism of the primary me- CBD was not changed between THC/CBD spray alone tabolite, 11-OH-THC. A suggested mechanism for this versus THC/CBD spray and omeprazole treatments. effect is that increased CYP3A4 activity induced by ri- However, a decrease in exposure to 11-OH-THC was fampicin induced further metabolism of 11-OH-THC, observed following co-administration of omeprazole causing a reduced plasma level of 11-OH-THC. This is with THC/CBD spray. Nevertheless, examining the indi- the first investigation in humans to identify CYP3A4 as a vidual exposure data, no clear conclusion could be significant mediator of 11-OH-THC metabolism, an inter- drawn with respect to any effect of omeprazole. These esting finding for this psychoactive THC metabolite. findings suggest that THC, CBD are not substrates for the CYP2C19 isoenzyme. Ketoconazole and THC/CBD spray Similarly, the effects of the potent CYP3A4 inhibitor ke- Safety and tolerability toconazole on the PKs of THC/CBD spray were also in- THC/CBD spray was generally well tolerated when given vestigated. For both THC and CBD, there was an alone, illustrated by the fact that there were no serious increase in mean C and AUC following THC/CBD AEs during the study. The majority of TEAEs were mild max spray administration after 5-day period of daily in severity and, as expected, of the SOC of nervous Stott et al. SpringerPlus 2013, 2:236 Page 12 of 15 http://www.springerplus.com/content/2/1/236 Table 4 Point estimate and 90% confidence interval for the geometric mean ratios of C , AUC and AUC for max (0-t) (0-inf) THC/CBD spray in combination with rifampicin, ketoconazole and omeprazole 90% confidence limits Analyte/PK variable Number of subjects Estimate Lower Upper Rifampicin and THC/CBD spray THC/C 12 0.614 0.525 0.719 max THC/AUC 12 0.711 0.617 0.820 (0-t) THC/AUC 11 0.761 0.667 0.868 (0-inf) CBD/C 12 0.480 0.420 0.550 max CBD/AUC 12 0.381 0.274 0.529 (0-t) CBD/AUC 10 0.422 0.265 0.673 (0-inf) 11-OH-THC/C 12 0.140 0.120 0.163 max 11-OH-THC/AUC 12 0.099 0.088 0.113 (0-t) 11-OH-THC/AUC 11 0.131 0.115 0.150 (0-inf) Ketoconazole and THC/CBD spray THC/C 11 1.252 1.043 1.503 max THC/AUC 11 1.770 1.385 2.263 (0-t) THC/AUC 10 1.840 1.446 2.342 (0-inf) CBD/C 11 1.961 1.497 2.569 max CBD/AUC 11 2.715 2.047 3.601 (0-t) CBD/AUC 9 1.923 1.560 2.370 (0-inf) 11-OH-THC/C 11 3.074 2.705 3.493 max 11-OH-THC/AUC 11 3.823 3.405 4.292 (0-t) 11-OH-THC/AUC 10 3.616 3.181 4.111 (0-inf) Omeprazole and THC/CBD spray THC/C 12 1.125 0.842 1.501 max THC/AUC 12 0.964 0.643 1.445 (0-t) THC/AUC 12 0.962 0.668 1.386 (0-inf) CBD/C 12 1.320 0.938 1.859 max CBD/AUC 12 1.326 0.756 2.325 (0-t) CBD/AUC 11 1.096 0.745 1.611 (0-inf) 11-OH-THC/C 12 0.866 0.700 1.073 max 11-OH-THC/AUC 12 0.869 0.629 1.201 (0-t) 11-OH-THC/AUC 12 0.869 0.641 1.179 (0-inf) system disorders, with somnolence being the most reported AEs (incidence >10%) were dizziness (28%), commonly reported. While the exposure to THC/CBD diarrhoea (13%), fatigue (11%) and nausea (11%) spray was lower than usual in chronic patients, the (Constantinescu & Sarantis 2006). Recent publications administration procedure did not follow the slow up- also demonstrate that AEs the SOC of nervous systems titration process that takes 4-5 days to reach a dose disorders were among the commonly occurring AEs of 4 sprays/day (and goes on for 7-8 days to reach reported with THC/CBD spray use (Johnson et al. 2012; the average 6-7 sprays/day dose, and up to 14 days to Langford et al. 2013). reach the maximum dose of 12 sprays/day). As such, No difference was observed in the proportions of sub- the incidence of AEs in the SOC of nervous disorders jects reporting AEs upon co-administration of THC/ in the current study is not surprising. A review of CBD spray with rifampicin or omeprazole compared collated results from initial randomized-controlled with THC/CBD spray alone. Again, all AEs were of mild clinical trials with THC/CBD spray involving 930 severity, with headache and dysguesia being the most patients demonstrated that the most frequently commonly reported for THC/CBD spray and rifampicin Stott et al. SpringerPlus 2013, 2:236 Page 13 of 15 http://www.springerplus.com/content/2/1/236 Table 5 Treatment emergent adverse events with an incidence >1 Primary system organ class Preferred term* No. of subjects (%) No. of subjects (%) No. of subjects (%) THC/CBD spray (n = 36) Psychiatric disorders Euphoric mood 2 (6) - - Nervous system disorders Dizziness 2 (6) - - Dizziness postural 3 (8) - - Headache 5 (14) - - Somnolence 11 (31) - - Gastrointestinal disorders Nausea 2 (6) - - Rifampicin (n = 12) THC/CBD spray and Rifampicin (n = 12) THC/CBD spray (n = 12) Nervous system disorders Dysgeusia 1 (8) 0 1 (8) Headache 0 1 (8) 1 (8) Somnolence 0 0 2 (17) Ketoconazole (n = 12) THC/CBD spray and Ketoconazole (n = 11) THC/CBD spray (n = 12) Nervous system disorders Somnolence 0 4 (36) 4 (33) Dizziness 0 3 (27) 2 (17) Lethargy 0 3 (27) 0 Dysgeusia 0 2 (18) 0 Headache 0 2 (18) 0 Somnolence 0 1 (9) 1 (8) Psychiatric disorders Euphoric mood 0 7 (64) 0 Anxiety 0 1 (9) 1 (8) Disorientation 0 1 (9) 1 (8) Omeprazole (n = 12) THC/CBD spray and Omeprazole (n = 12) THC/CBD spray (n = 12) Nervous system disorders Dizziness 0 3 (25) 0 Dizziness postural 0 0 2 (17) Headache 1 (8) 0 4 (33) Somnolence 0 2 (17) 5 (42) Gastrointestinal disorders Dry mouth 0 2 (17) 1 (8) *MedDRA version 10.0. (one subject), and dizziness in three subjects taking spray alone. However, there was an increased incidence THC/CBD spray and omeprazole. of euphoric mood, lethargy, dygeusia and headache Although all subjects receiving THC/CBD spray and when THC/CBD spray was given in combination with ketoconazole experienced AEs, this combination was still ketoconazole. Only one event was classed as moderate relatively well tolerated, and all but one AE was of mild in terms of severity (anxiety) and the event resolved severity. The majority of TEAEs were classed as nervous without intervention. All other events were of mild system disorders, including somnolence and dizziness severity. Notably in this group, all subjects had which occurred at the same incidence as with THC/CBD increases in C for 11-OH-THC, and seven had an max Stott et al. SpringerPlus 2013, 2:236 Page 14 of 15 http://www.springerplus.com/content/2/1/236 increase in THC C , with nine subjects also having Conclusions max an increase in AUC. Taking into account the increase In conclusion, inhibition of CYP2C19 by omeprazole did in exposure after combined administration compared not significantly alter the PK of THC/CBD spray to THC/CBD spray alone, it is possible that the difference suggesting that THC, CBD and 11-OH-THC are not in PKs may account for the increase in CNS-type substrates for this isoenzyme at the clinically relevant AEs observed. dose of THC/CBD spray investigated. The CYP3A4 in- ducer rifampicin caused a decrease in exposure to all Study limitations three analytes, although not extreme and within the nat- There were a number of limitations to this study. Kel ural range of variation between subjects. Conversely, the was estimated over a short sampling period (less than CYP3A4 inhibitor ketoconazole caused increased expos- two t s) for the majority of profiles in this study and ure to all analytes, suggesting that THC, CBD and 1/2 the regression had low precision in some profiles. AUC 11-OH-THC all are substrates for this isoenzyme. More- had an extrapolated area which was larger than over, the findings with rifampicin and ketoconazole sug- (0-inf) 20% for some profiles. However, despite these limita- gest that CYP3A4 is involved in the metabolism of tions, the AUC and the AUC were in agreement, 11-OH-THC. On the basis of these findings, there is (0-inf) (0-t) and the statistical results are thus considered reliable. likely to be little impact on other drugs metabolized by Further consistency and validity of the study is demon- CYP enzymes on the PK parameters of THC/CBD spray, strated by the similarity of the PK exposure parameters but potential effects should be taken into consideration following a single dose of THC/CBD spray in the ab- when co-administering THC/CBD spray with com- sence of any inducer or inhibitor, which are in good pounds which share the CYP3A4 pathway. THC/CBD agreement with the PK data provided in a previous clin- spray was also well tolerated in healthy subjects both ical study performed by the authors (ref PK study). The alone and in combination with rifampicin, ketoconazole inter-subject variability was substantial and greater than and omeprazole. the difference in exposure means before and after rifampi- cin, ketoconazole, or omeprazole treatment, suggesting Ethical standards that any effect of other medications metabolized by rele- The current study was approved by Guy's Research Eth- vant CYP enzymes on THC/CBD spray is likely to be ics Committee, and was conducted in accordance with within the normal range of variation. However, there was the International Conference on Harmonisation guide- generally a similar variability in the group mean primary lines on Good Clinical Practice and the ethical principles PK parameters between the THC/CBD spray alone and stated in the Declaration of Helsinki and local UK regu- the THC/CBD spray plus inducer or inhibitor treatments. lations. All participants gave written informed consent. Doses of THC/CBD (4 sprays instead the average 6-8 † Sativex®, a THC/CBD oromucosal spray, does not sprays/day) and intake patterns (4 sprays in a row in- have an INN. Nabiximols is the US Adopted Name stead of evenly distributed through the day) were not (USAN). equivalent to those in the approved label. However, dur- ing a phase IIb dose ranging study in cancer patients Competing interests with pain, in the low dose THC/CBD spray (1-4 sprays) GW Pharmaceutical Ltd produces THC/CBD spray and is licensed for the treatment of spasticity in multiple sclerosis in several European countries, as group, over 90% of patients titrated to a dose of 3 or 4 well as Canada and New Zealand. This study was funded by GW sprays, leading to the conclusion that a minimal effective Pharmaceuticals Ltd. Darren Wilbraham is an employee of Quintiles Ltd who dose was 3 sprays per day (Porteney et al. 2012). were contracted to perform the clinical study. Colin Stott, Linda White, Stephen Wright and Geoffrey Guy are all employees of GW Pharmaceuticals Additionally, efficacy was observed in the low dose Ltd, and are shareholders in the company. group, reaching statistical significance for the continuous response analysis (pain 0-10 Numerical Rating Scale Authors’ contributions score) and for the mean change from baseline in score, CGS, LW, SW and GWG made a substantial contribution to the conception demonstrating that 4 daily sprays is a clinically relevant and design of the study. DW made a substantial contribution to the acquisition of the data, and CGS, LW, SW and GWG were involved in the dose (Porteney et al. 2012). Distributing the dose through- analysis of the data. All authors had a hand in the preparation of this out the day in the current study would have given a very manuscript based on their interpretation of the data, and reviewed, different PK profile, and would not have been a suitable approved and agreed upon the manuscript in its final format. approach for the current study. Additionally, during a Author details previous Phase I pharmacokinetics study, doses of 2 GW Pharma Ltd, Porton Down Science Park Salisbury, Wiltshire SP4 0JQ, UK. daily sprays of THC/CBD spray gave low C values Quintiles Drug Research Unit at Guy's Hospital, 6 Newcomen Street, max London, UK. (Stott et al. 2012). As such, a dose of 4 daily sprays was chosen to give good plasma concentration over Received: 26 February 2013 Accepted: 3 May 2013 time curves. Published: 24 May 2013 Stott et al. 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