RESUMO
Cannabis sativa is known for its therapeutic benefit in various diseases including pain relief by targeting cannabinoid receptors. The primary component of cannabis, Δ9-tetrahydrocannabinol (THC), and other agonists engage the orthosteric site of CB1, activating both Gi and ß-arrestin signaling pathways. The activation of diverse pathways could result in on-target side effects and cannabis addiction, which may hinder therapeutic potential. A significant challenge in pharmacology is the design of a ligand that can modulate specific signaling of CB1. By leveraging insights from the structure-function selectivity relationship (SFSR), we have identified Gi signaling-biased agonist-allosteric modulators (ago-BAMs). Further, two cryoelectron microscopy (cryo-EM) structures reveal the binding mode of ago-BAM at the extrahelical allosteric site of CB1. Combining mutagenesis and pharmacological studies, we elucidated the detailed mechanism of ago-BAM-mediated biased signaling. Notably, ago-BAM CB-05 demonstrated analgesic efficacy with fewer side effects, minimal drug toxicity and no cannabis addiction in mouse pain models. In summary, our finding not only suggests that ago-BAMs of CB1 provide a potential nonopioid strategy for pain management but also sheds light on BAM identification for GPCRs.
Assuntos
Microscopia Crioeletrônica , Receptor CB1 de Canabinoide , Transdução de Sinais , Receptor CB1 de Canabinoide/metabolismo , Receptor CB1 de Canabinoide/genética , Receptor CB1 de Canabinoide/química , Animais , Regulação Alostérica/efeitos dos fármacos , Camundongos , Humanos , Transdução de Sinais/efeitos dos fármacos , Proteínas de Ligação ao GTP/metabolismo , Proteínas de Ligação ao GTP/genética , Células HEK293 , Relação Estrutura-Atividade , Dronabinol/farmacologia , Dronabinol/química , Dronabinol/análogos & derivados , Cannabis/química , Cannabis/metabolismoRESUMO
Cannabis sativa L. has been cultivated and used around the globe for its medicinal properties for millennia1. Some cannabinoids, the hallmark constituents of Cannabis, and their analogues have been investigated extensively for their potential medical applications2. Certain cannabinoid formulations have been approved as prescription drugs in several countries for the treatment of a range of human ailments3. However, the study and medicinal use of cannabinoids has been hampered by the legal scheduling of Cannabis, the low in planta abundances of nearly all of the dozens of known cannabinoids4, and their structural complexity, which limits bulk chemical synthesis. Here we report the complete biosynthesis of the major cannabinoids cannabigerolic acid, Δ9-tetrahydrocannabinolic acid, cannabidiolic acid, Δ9-tetrahydrocannabivarinic acid and cannabidivarinic acid in Saccharomyces cerevisiae, from the simple sugar galactose. To accomplish this, we engineered the native mevalonate pathway to provide a high flux of geranyl pyrophosphate and introduced a heterologous, multi-organism-derived hexanoyl-CoA biosynthetic pathway5. We also introduced the Cannabis genes that encode the enzymes involved in the biosynthesis of olivetolic acid6, as well as the gene for a previously undiscovered enzyme with geranylpyrophosphate:olivetolate geranyltransferase activity and the genes for corresponding cannabinoid synthases7,8. Furthermore, we established a biosynthetic approach that harnessed the promiscuity of several pathway genes to produce cannabinoid analogues. Feeding different fatty acids to our engineered strains yielded cannabinoid analogues with modifications in the part of the molecule that is known to alter receptor binding affinity and potency9. We also demonstrated that our biological system could be complemented by simple synthetic chemistry to further expand the accessible chemical space. Our work presents a platform for the production of natural and unnatural cannabinoids that will allow for more rigorous study of these compounds and could be used in the development of treatments for a variety of human health problems.
Assuntos
Vias Biossintéticas , Canabinoides/biossíntese , Canabinoides/química , Cannabis/química , Engenharia Metabólica , Saccharomyces cerevisiae/metabolismo , Acil Coenzima A/biossíntese , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Benzoatos/metabolismo , Vias Biossintéticas/genética , Canabinoides/metabolismo , Cannabis/genética , Dronabinol/análogos & derivados , Dronabinol/metabolismo , Fermentação , Galactose/metabolismo , Ácido Mevalônico/metabolismo , Fosfatos de Poli-Isoprenil/biossíntese , Fosfatos de Poli-Isoprenil/metabolismo , Saccharomyces cerevisiae/genética , Salicilatos/metabolismoRESUMO
The diversity of cannabinoid isomers and complexity of Cannabis products pose significant challenges for analytical methodologies. In this study, we developed a method to analyze 14 different cannabinoid isomers in diverse samples within milliseconds by leveraging the unique adduct-forming behavior of silver ions in advanced cyclic ion mobility spectrometry-mass spectrometry. The developed method achieved the separation of isomers from four groups of cannabinoids: Δ3-tetrahydrocannabinol (THC) (1), Δ8-THC (2), Δ9-THC (3), cannabidiol (CBD) (4), Δ8-iso-THC (5), and Δ(4)8-iso-THC (6) (all MW = 314); 9α-hydroxyhexahydrocannabinol (7), 9ß-hydroxyhexahydrocannabinol (8), and 8-hydroxy-iso-THC (9) (all MW = 332); tetrahydrocannabinolic acid (THCA) (10) and cannabidiolic acid (CBDA) (11) (both MW = 358); Δ8-tetrahydrocannabivarin (THCV) (12), Δ8-iso-THCV (13), and Δ9-THCV (14) (all MW = 286). Moreover, experimental and theoretical traveling wave collision cross section values in nitrogen (TWCCSN2) of cannabinoid-Ag(I) species were obtained for the first time with an average error between experimental and theoretical values of 2.6%. Furthermore, a workflow for the identification of cannabinoid isomers in Cannabis and Cannabis-derived samples was established based on three identification steps (m/z and isotope pattern of Ag(I) adducts, TWCCSN2, and MS/MS fragments). Afterward, calibration curves of three major cannabinoids were established with a linear range of 1-250 ng·ml-1 for Δ8-THC (2) (R2 = 0.9999), 0.1-25 ng·ml-1 for Δ9-THC (3) (R2 = 0.9987), and 0.04-10 ng·ml-1 for CBD (4) (R2 = 0.9986) as well as very low limits of detection (0.008-0.2 ng·ml-1). Finally, relative quantification of Δ8-THC (2), Δ9-THC (3), and CBD (4) in eight complex acid-treated CBD mixtures was achieved without chromatographic separation. The results showed good correspondence (R2 = 0.999) with those obtained by gas chromatography-flame ionization detection/mass spectrometry.
Assuntos
Canabinoides , Cannabis , Dronabinol , Espectrometria de Mobilidade Iônica , Espectrometria de Massas , Cannabis/química , Canabinoides/análise , Canabinoides/química , Dronabinol/análise , Dronabinol/análogos & derivados , Espectrometria de Mobilidade Iônica/métodos , Espectrometria de Massas/métodos , Extratos Vegetais/química , Extratos Vegetais/análise , IsomerismoRESUMO
Cultivation of industrial low-Δ9-tetrahydrocannabinol (Δ9-THC) hemp has created an oversupply of cannabidiol (CBD)-rich products. The fact that phytocannabinoids, including CBD, can be used as precursors to synthetically produce a range of THC variants-potentially located in a legal loophole-has led to a diversification of cannabis recreational drug markets. 'Hemp-compliant', 'hemp-derived' and 'semisynthetic' cannabinoid products are emerging and being advertised as (legal) alternatives for Δ9-THC. This study included a large panel (n = 30) of THC isomers, homologs, and analogs that might be derived via semisynthetic procedures. As a proxy for the abuse potential of these compounds, we assessed their potential to activate the CB1 cannabinoid receptor with a ß-arrestin2 recruitment bioassay (picomolar-micromolar concentrations). Multiple THC homologs (tetrahydrocannabihexol, THCH; tetrahydrocannabiphorol, THCP; tetrahydrocannabinol-C8, THC-C8) and THC analogs (hexahydrocannabinol, HHC; hexahydrocannabiphorol, HHCP) were identified that showed higher potential for CB1 activation than Δ9-THC, based on either higher efficacy (Emax) or higher potency (EC50). Structure-activity relationships were assessed for Δ9-THC and Δ8-THC homologs encompassing elongated alkyl chains. Additionally, stereoisomer-specific differences in CB1 activity were established for various THC isomers (Δ7-THC, Δ10-THC) and analogs (HHC, HHCP). Evaluation of the relative abundance of 9(S)-HHC and 9(R)-HHC epimers in seized drug material revealed varying epimeric compositions between batches. Increased abundance of the less active 9(S)-HHC epimer empirically resulted in decreased potency, but sustained efficacy for the resulting diastereomeric mixture. In conclusion, monitoring of semisynthetic cannabinoids is encouraged as the dosing and the relative composition of stereoisomers can impact the harm potential of these drugs, relative to Δ9-THC products.
Assuntos
Canabinoides , Cannabis , Dronabinol , beta-Arrestina 2 , Cannabis/química , Humanos , Dronabinol/análogos & derivados , Dronabinol/toxicidade , Dronabinol/química , Canabinoides/toxicidade , Canabinoides/química , beta-Arrestina 2/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Drogas Ilícitas/toxicidade , Drogas Ilícitas/química , Canabidiol/toxicidade , Canabidiol/química , Células HEK293RESUMO
Natural and semi-synthetic cannabinoid analogs are getting increasing media attention for their recreative use as an alternative to traditional cannabis, in Sweden as well as internationally. To investigate an increasing number of urine samples incoming to our clinical laboratory that were screening positive, using a CEDIA THC-COOH immunoassay from ThermoFisher Scientific, but then testing negative using GC-MS based verification analysis, we developed an LC-MS/MS-method for verification of hexahydrocannabinol (HHC) and Δ8-tetrahydrocannabinol. Assessment of HHC intake was based on identification of the following four metabolites: 11-nor-9(R)-carboxy-hexahydrocannabinol (R-HHC-COOH), 11-nor-9(S)-carboxy-hexahydrocannabinol (S-HHC-COOH), 11-hydroxy-9(R)-hexahydrocannabinol (R-HHC-OH) and 11-hydroxy-9(S)-hexahydrocannabinol (S-HHC-OH). Out of 46 urine samples analysed in this study, 44 showed presence of HHC-metabolites, which indicate HHC as the main explanation for an increased number of negative verifications for THC-COOH. In these samples, the HHC-OH metabolites occurred at a higher concentration than R-HHC-COOH while S-HHC-COOH was only detected in few samples at low concentrations. R-HHC-COOH and S-HHC-COOH can easily be added to a pre-existing verification method for THC-COOH, and still show acceptable results, while HHC-OH requires an enzyme capable of hydrolysing the ether glucuronide bond.
Assuntos
Dronabinol , Detecção do Abuso de Substâncias , Humanos , Dronabinol/urina , Dronabinol/análogos & derivados , Cromatografia Gasosa-Espectrometria de Massas/métodos , Espectrometria de Massa com Cromatografia Líquida , Detecção do Abuso de Substâncias/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
This study investigated the effects of hexahydrocannabinol (HHC) and other unclassified cannabinoids, which were recently introduced to the recreational drug market, on cannabis drug testing in urine and oral fluid samples. After the appearance of HHC in Sweden in 2022, the number of posts about HHC on an online drug discussion forum increased significantly in the spring of 2023, indicating increased interest and use. In parallel, the frequency of false positive screening tests for tetrahydrocannabinol (THC) in oral fluid, and for its carboxy metabolite (THC-COOH) in urine, rose from <2% to >10%. This suggested that HHC cross-reacted with the antibodies in the immunoassay screening, which was confirmed in spiking experiments with HHC, HHC-COOH, HHC acetate (HHC-O), hexahydrocannabihexol (HHC-H), hexahydrocannabiphorol (HHC-P), and THC-P. When HHC and HHC-P were classified as narcotics in Sweden on 11 July 2023, they disappeared from the online and street shops market and were replaced by other unregulated variants (e.g. HHC-O and THC-P). In urine samples submitted for routine cannabis drug testing, HHC-COOH concentrations up to 205 (mean 60, median 27) µg/L were observed. To conclude, cannabis drug testing cannot rely on results from immunoassay screening, as it cannot distinguish between different tetra- and hexahydrocannabinols, some being classified but others unregulated. The current trend for increased use of unregulated cannabinols will likely increase the proportion of positive cannabis screening results that need to be confirmed with mass spectrometric methods. However, the observed cross-reactivity also means a way to pick up use of new cannabinoids that otherwise risk going undetected.
Assuntos
Drogas Ilícitas , Detecção do Abuso de Substâncias , Humanos , Detecção do Abuso de Substâncias/métodos , Drogas Ilícitas/urina , Drogas Ilícitas/análise , Suécia , Dronabinol/urina , Dronabinol/análise , Dronabinol/análogos & derivados , Cannabis/química , Saliva/química , Canabinoides/urina , Canabinoides/análise , Canabinol/análise , Canabinol/urina , Reações Cruzadas , Imunoensaio/métodosRESUMO
Concerns about health hazards associated with the consumption of trans-delta-8-tetrahydrocannabinol products were highlighted in public health advisories from the U.âS. Food and Drug Administration and U.âS. Centers for Disease Control and Prevention. Simple and rapid quantitative methods to determine trans-delta-8-tetrahydrocannabinol impurities are vital to analyze such products. In this study, a gas chromatography-flame ionization detection method was developed and validated for the determination of delta-8-tetrahydrocannabinol and some of its impurities (recently published) found in synthesized trans-delta-8-tetrahydrocannabinol raw material and included olivetol, cannabicitran, Δ 8-cis-iso-tetrahydrocannabinol, Δ 4-iso-tetrahydrocannabinol, iso-tetrahydrocannabifuran, cannabidiol, Δ 4,8-iso-tetrahydrocannabinol, Δ 8-iso-tetrahydrocannabinol, 4,8-epoxy-iso-tetrahydrocannabinol, trans-Δ 9-tetrahydrocannabinol, 8-hydroxy-iso-THC, 9α-hydroxyhexahydrocannabinol, and 9ß-hydroxyhexahydrocannabinol. Validation of the method was assessed according to the International Council for Harmonization guidelines and confirmed linearity with R2 ≥ 0.99 for all the target analytes. The limit of detection and limit of quantitation were 1.5 and 5 µg/mL, respectively, except for olivetol, which had a limit of detection of 3 µg/mL and a limit of quantitation of 10 µg/mL. Method precision was calculated as % relative standard deviation and the values were less than 8.4 and 9.9% for the intraday precision and inter-day precision, respectively. The accuracy ranged from 85 to 118%. The method was then applied to the analysis of 21 commercially marketed vaping products claiming to contain delta-8-tetrahydrocannabinol. The products analyzed by this method have various levels of these impurities, with all products far exceeding the 0.3% of trans-Δ 9-tetrahydrocannabinol limit for hemp under the Agriculture Improvement Act of 2018. The developed gas chromatography-flame ionization detection method can be an important tool for monitoring delta-8-tetrahydrocannabinol impurities in commercial products.
Assuntos
Dronabinol , Dronabinol/análogos & derivados , Resorcinóis , Vaping , Dronabinol/análise , Cromatografia Gasosa-Espectrometria de Massas/métodos , Cromatografia GasosaRESUMO
INTRODUCTION: The presence of cannabinoid products in the home may increase the likelihood of unintended adverse consequences for children and adolescents. Secure storage of these products is one prevention method to decrease the risk of diversion and use of cannabinoid products among youth. We sought to examine cannabis, delta-8 tetrahydrocannabinol (THC), and cannabidiol (CBD) storage practices among a sample of adults 18-64 years old residing in the USA. METHODS: In December 2021, we conducted an online cross-sectional survey of 1042 current (past 30 day) users of cannabinoid products (88.3% cannabis, 49.0% delta-8 THC, and 67.2% CBD). Participants were asked about where they typically keep products in their home (ie, in a locked container, unlocked container, or out in the open). We conducted multinomial regression analyses to examine the relationship between sociodemographic characteristics and cannabinoid use behaviours with home storage practices. RESULTS: For all products, participants more frequently reported locking, followed by storing the product in an unlocked but not visible location. Storing the product in an unlocked and visible location was endorsed the least across all three products. Participants reported more frequent endorsement of locking cannabis products as compared with delta-8-THC and CBD. Storage practices varied by biological sex, sexual orientation, ethnicity, educational attainment, having a child who lives in the home, frequency of use, possession of a medical cannabis card and exposure to advertising. CONCLUSIONS: Increasing the prevalence of secure storage practices of cannabinoid products may facilitate prevention of unanticipated consequences associated with diversion of these products.
Assuntos
Canabinoides , Humanos , Masculino , Feminino , Adulto , Estudos Transversais , Estados Unidos , Pessoa de Meia-Idade , Adolescente , Adulto Jovem , Dronabinol/análise , Dronabinol/análogos & derivados , Cannabis , Canabidiol , Inquéritos e QuestionáriosRESUMO
Importance: Gummies, flavored vaping devices, and other cannabis products containing psychoactive hemp-derived Δ8-tetrahydrocannabinol (THC) are increasingly marketed in the US with claims of being federally legal and comparable to marijuana. National data on prevalence and correlates of Δ8-THC use and comparisons to marijuana use among adolescents in the US are lacking. Objective: To estimate the self-reported prevalence of and sociodemographic and policy factors associated with Δ8-THC and marijuana use among US adolescents in the past 12 months. Design, Setting, and Participants: This nationally representative cross-sectional analysis included a randomly selected subset of 12th-grade students in 27 US states who participated in the Monitoring the Future Study in-school survey during February to June 2023. Exposures: Self-reported sex, race, ethnicity, and parental education; census region; state-level adult-use (ie, recreational) marijuana legalization (yes vs no); and state-level Δ8-THC policies (regulated vs not regulated). Main Outcomes and Measures: The primary outcome was self-reported Δ8-THC and marijuana use in the past 12 months (any vs no use and number of occasions used). Results: In the sample of 2186 12th-grade students (mean age, 17.7 years; 1054 [48.9% weighted] were female; 232 [11.1%] were Black, 411 [23.5%] were Hispanic, 1113 [46.1%] were White, and 328 [14.2%] were multiracial), prevalence of self-reported use in the past 12 months was 11.4% (95% CI, 8.6%-14.2%) for Δ8-THC and 30.4% (95% CI, 26.5%-34.4%) for marijuana. Of those 295 participants reporting Δ8-THC use, 35.4% used it at least 10 times in the past 12 months. Prevalence of Δ8-THC use was lower in Western vs Southern census regions (5.0% vs 14.3%; risk difference [RD], -9.4% [95% CI, -15.2% to -3.5%]; adjusted risk ratio [aRR], 0.35 [95% CI, 0.16-0.77]), states in which Δ8-THC was regulated vs not regulated (5.7% vs 14.4%; RD, -8.6% [95% CI, -12.9% to -4.4%]; aRR, 0.42 [95% CI, 0.23-0.74]), and states with vs without legal adult-use marijuana (8.0% vs 14.0%; RD, -6.0% [95% CI, -10.8% to -1.2%]; aRR, 0.56 [95% CI, 0.35-0.91]). Use in the past 12 months was lower among Hispanic than White participants for Δ8-THC (7.3% vs 14.4%; RD, -7.2% [95% CI, -12.2% to -2.1%]; aRR, 0.54 [95% CI, 0.34-0.87]) and marijuana (24.5% vs 33.0%; RD, -8.5% [95% CI, -14.9% to -2.1%]; aRR, 0.74 [95% CI, 0.59-0.94]). Δ8-THC and marijuana use prevalence did not differ by sex or parental education. Conclusions and Relevance: Δ8-THC use prevalence is appreciable among US adolescents and is higher in states without marijuana legalization or existing Δ8-THC regulations. Prioritizing surveillance, policy, and public health efforts addressing adolescent Δ8-THC use may be warranted.
Assuntos
Dronabinol , Alucinógenos , Uso da Maconha , Transtornos Relacionados ao Uso de Substâncias , Adolescente , Adulto , Feminino , Humanos , Masculino , Cannabis , Estudos Transversais , Fumar Maconha/epidemiologia , Fumar Maconha/legislação & jurisprudência , Uso da Maconha/epidemiologia , Uso da Maconha/legislação & jurisprudência , Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Estados Unidos/epidemiologia , Prevalência , Estudantes/estatística & dados numéricos , Autorrelato , Grupos Raciais/etnologia , Grupos Raciais/estatística & dados numéricos , Dronabinol/análogos & derivadosRESUMO
Phytocannabinoids with seven-carbon alkyl chains (phorols) have gained a lot of attention, as they are commonly believed to be more potent versions of typical cannabinoids with shorter alkyl chains. At the time of this article, cannabidiphorol (CBDP) and tetrahydrocannabiphorol (THCP) can both be purchased in the North American market, even though their biological activities are nearly unknown. To investigate their relative potency, we conducted in vitro receptor-binding experiments with CBDP (cannabinoid CB1/CB2 receptor antagonism, serotonin 5HT-1A agonism, dopamine D2S (short form) agonism, and mu-opioid negative allosteric modulation) and compared the observed activity with that of CBD. To our knowledge, this is the first publication to investigate CBDP's receptor activity in vitro. A similar activity profile was observed for both CBD and CBDP, with the only notable difference at the CB2 receptor. Contrary to common expectations, CBD was found to be a slightly more potent CB2 antagonist than CBDP (p < 0.05). At the highest tested concentration, CBD demonstrated antagonist activity with a 33% maximum response of SR144528 (selective CB2 antagonist/inverse agonist). CBDP at the same concentration produced a weaker antagonist activity. A radioligand binding assay revealed that among cannabinoid and serotonin receptors, CB2 is likely the main biological target of CBDP. However, both CBD and CBDP were found to be significantly less potent than SR144528. The interaction of CBDP with the mu-opioid receptor (MOR) produced unexpected results. Although the cannabidiol family is considered to be a set of negative allosteric modulators (NAMs) of opioid receptors, we observed a significant increase in met-enkephalin-induced mu-opioid internalization when cells were incubated with 3 µM of CBDP and 1 µM met-enkephalin, a type of activity expected from positive allosteric modulators (PAMs). To provide a structural explanation for the observed PAM effect, we conducted molecular docking simulations. These simulations revealed the co-binding potential of CBDP (or CBD) and met-enkephalin to the MOR.
Assuntos
Receptor CB2 de Canabinoide , Humanos , Receptor CB2 de Canabinoide/metabolismo , Canabidiol/farmacologia , Canabidiol/metabolismo , Canabidiol/química , Receptores Opioides mu/metabolismo , Receptores Opioides mu/agonistas , Receptor CB1 de Canabinoide/metabolismo , Receptor CB1 de Canabinoide/antagonistas & inibidores , Ligação Proteica , Canabinoides/metabolismo , Canabinoides/farmacologia , Canabinoides/química , Dronabinol/farmacologia , Dronabinol/análogos & derivados , Dronabinol/química , Dronabinol/metabolismo , Receptores de Dopamina D2/metabolismo , AnimaisRESUMO
Recently, hexahydrocannabinol (HHC) was posed under strict control in Europe due to the increasing HHC-containing material seizures. The lack of analytical methods in clinical laboratories to detect HHC and its metabolites in biological matrices may result in related intoxication underreporting. We developed and validated a comprehensive GC-MS/MS method to quantify 9(R)-HHC, 9(S)-HHC, 9αOH-HHC, 9ßOH-HHC, 8(R)OH-9(R)-HHC, 8(S)OH-9(S)HHC, 11OH-9(R)HHC, 11OH-9(S)HHC, 11nor-carboxy-9(R)-HHC, and 11nor-carboxy-9(S)-HHC in whole blood, urine, and oral fluid. A novel QuEChERS extraction protocol was optimized selecting the best extraction conditions suitable for all the three matrices. Urine and blood were incubated with ß-glucuronidase at 60 °C for 2 h. QuEChERS extraction was developed assessing different ratios of Na2SO4:NaCl (4:1, 2:1, 1:1, w/w) to be added to 200 µL of any matrix added with acetonitrile. The chromatographic separation was achieved on a 7890B GC with an HP-5ms column, (30 m, 0.25 mm × 0.25 µm) in 12.50 min. The analytes were detected with a triple-quadrupole mass spectrometer in the MRM mode. The method was fully validated following OSAC guidelines. The method showed good validation parameters in all the matrices. The method was applied to ten real samples of whole blood (n = 4), urine (n = 3), and oral fluid (n = 3). 9(R)-HHC was the prevalent epimer in all the samples (9(R)/9(S) = 2.26). As reported, hydroxylated metabolites are proposed as urinary biomarkers, while carboxylated metabolites are hematic biomarkers. Furthermore, 8(R)OH-9(R)HHC was confirmed as the most abundant metabolite in all urine samples.
Assuntos
Dronabinol , Cromatografia Gasosa-Espectrometria de Massas , Espectrometria de Massas em Tandem , Humanos , Cromatografia Gasosa-Espectrometria de Massas/métodos , Espectrometria de Massas em Tandem/métodos , Dronabinol/urina , Dronabinol/sangue , Dronabinol/análogos & derivados , Saliva/química , Saliva/metabolismo , Reprodutibilidade dos TestesRESUMO
Cannabis sativa is one of the oldest plants utilized by humans for both economic and medical purposes. Although the use of cannabis started millennia ago in the Eastern hemisphere, its use has moved and flourished in the Western nations in more recent centuries. C. sativa is the source of psychoactive cannabinoids that are consumed as recreational drugs worldwide. The C21 aromatic hydrocarbons are restricted in their natural occurrence to cannabis (with a few exceptions). Delta-9-tetrahydrocannabinol (Δ9-THC) is the main psychoactive component in cannabis, with many pharmacological effects and various approved medical applications. However, a wide range of side effects are associated with the use of Δ9-THC, limiting its medical use. In 1966, another psychoactive cannabinoid, Delta-8-tetrahydrocannabinol (Δ8-THC) was isolated from marijuana grown in Maryland but in very low yield. Δ8-THC is gaining increased popularity due to its better stability and easier synthetic manufacturing procedures compared to Δ9-THC. The passing of the U.S. Farm Bill in 2018 led to an increase in the sale of Δ8-THC in the United States. The marketed products contain Δ8-THC from synthetic sources. In this review, methods of extraction, purification, and structure elucidation of Δ8-THC will be presented. The issue of whether Δ8-THC is a natural compound or an artifact will be discussed, and the different strategies for its chemical synthesis will be presented. Δ8-THC of synthetic origin is expected to contain some impurities due to residual amounts of starting materials and reagents, as well as side products of the reactions. The various methods of analysis and detection of impurities present in the marketed products will be discussed. The pharmacological effects of Δ8-THC, including its interaction with CB1 and CB2 cannabinoid receptors in comparison with Δ9-THC, will be reviewed.
Assuntos
Canabinoides , Cannabis , Dronabinol/análogos & derivados , Alucinógenos , Humanos , Dronabinol/farmacologia , Canabinoides/farmacologia , Agonistas de Receptores de Canabinoides/farmacologia , Alucinógenos/farmacologiaRESUMO
The cannabinoid receptor 1 (CB1) is the principal target of the psychoactive constituent of marijuana, the partial agonist Δ9-tetrahydrocannabinol (Δ9-THC). Here we report two agonist-bound crystal structures of human CB1 in complex with a tetrahydrocannabinol (AM11542) and a hexahydrocannabinol (AM841) at 2.80 Å and 2.95 Å resolution, respectively. The two CB1-agonist complexes reveal important conformational changes in the overall structure, relative to the antagonist-bound state, including a 53% reduction in the volume of the ligand-binding pocket and an increase in the surface area of the G-protein-binding region. In addition, a 'twin toggle switch' of Phe2003.36 and Trp3566.48 (superscripts denote Ballesteros-Weinstein numbering) is experimentally observed and appears to be essential for receptor activation. The structures reveal important insights into the activation mechanism of CB1 and provide a molecular basis for predicting the binding modes of Δ9-THC, and endogenous and synthetic cannabinoids. The plasticity of the binding pocket of CB1 seems to be a common feature among certain class A G-protein-coupled receptors. These findings should inspire the design of chemically diverse ligands with distinct pharmacological properties.
Assuntos
Agonistas de Receptores de Canabinoides/química , Dronabinol/análogos & derivados , Droperidol/análogos & derivados , Receptor CB1 de Canabinoide/agonistas , Receptor CB1 de Canabinoide/química , Sítios de Ligação , Agonistas de Receptores de Canabinoides/síntese química , Agonistas de Receptores de Canabinoides/farmacologia , Cristalografia por Raios X , Dronabinol/síntese química , Dronabinol/química , Dronabinol/farmacologia , Droperidol/síntese química , Droperidol/química , Droperidol/farmacologia , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Humanos , Ligantes , Simulação de Acoplamento Molecular , Ligação Proteica , Conformação Proteica , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptor CB1 de Canabinoide/metabolismoRESUMO
BACKGROUND: Lenabasum is a synthetic cannabinoid receptor type-2 (CB2) agonist able to exert potent anti-inflammatory effects, but its role on T cells remains unknown. OBJECTIVES: The present study was undertaken to investigate anti-inflammatory mechanisms of lenabasum in T lymphocyte subsets and its in vivo therapeutic efficacy in experimental autoimmune encephalomyelitis (EAE). METHODS: Mononuclear cells from 17 healthy subjects (HS) and 25 relapsing-remitting multiple sclerosis (RRMS) patients were activated in presence or absence of lenabasum and analysed by flow cytometry and qRT-PCR. EAE mice were treated with lenabasum, and clinical score and neuroinflammation were evaluated. RESULTS: Lenabasum significantly reduced TNF-a production from CD4+ T cells and CD8+ T cells in a dose-dependent manner in both HS and RRMS patients. In MS patients, lenabasum also reduced activation marker CD25 and inhibited IL-2 production from both T cell subsets and IFN-γ and IL-17 from committed Th1 and Th17 cells, respectively. These effects were blocked by the pretreatment with selective CB2 inverse agonist SR144528. In vivo treatment of EAE mice with lenabasum significantly ameliorated disease severity, reduced neuroinflammation and demyelination in spinal cord. CONCLUSION: Lenabasum exerts potent T cell-mediated immunomodulatory effects, suggesting CB2 as a promising pharmacological target to counteract neuroinflammation in MS.
Assuntos
Anti-Inflamatórios/farmacologia , Agonistas de Receptores de Canabinoides/farmacologia , Dronabinol/análogos & derivados , Esclerose Múltipla Recidivante-Remitente/imunologia , Receptor CB2 de Canabinoide/agonistas , Subpopulações de Linfócitos T/efeitos dos fármacos , Adulto , Animais , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Dronabinol/farmacologia , Encefalomielite Autoimune Experimental/imunologia , Feminino , Humanos , Masculino , Camundongos , Subpopulações de Linfócitos T/imunologiaRESUMO
(-)-Δ9-Tetrahydrocannabinol (THC) is the psychoactive constituent of cannabis, a drug recreationally consumed orally or by inhalation. Physiologically based pharmacokinetic (PBPK) modeling can be used to predict systemic and tissue exposure to THC and its psychoactive metabolite, (±)-11-hydroxy-Δ9-THC (11-OH-THC). To populate a THC/11-OH-THC PBPK model, we previously characterized the depletion clearance of THC (by CYP2C9) and 11-OH-THC (by UDP-glucuronosyltransferase (UGT), CYP3A, and CYP2C9) in adult human liver microsomes. Here we focused on quantifying extrahepatic depletion clearance of THC/11-OH-THC, important after oral (intestine) and inhalational (lung) consumption of THC as well as prenatal THC use (placenta and fetal liver). THC (500 nM) was metabolized in adult human intestinal microsomes (n = 3-5) by CYP2C9 [Vmax: 1.1 ± 0.38 nmol/min/mg; Michaelis-Menten constant (Km): 70 nM; intrinsic clearance (CLint): 15 ± 5.4 ml/min/mg; fraction metabolized (fm): 0.89 ± 0.31 at concentration ⪠70 nM] and CYP3A (CLint: 2.0 ± 0.86 ml/min/mg; fm: 0.11 ± 0.050). 11-OH-THC (50 nM) was metabolized by CYP3A (CLint: 0.26 ± 0.058 ml/min/mg; fm: 0.51 ± 0.11) and UGT2B7 (CLint: 0.13 ± 0.027 ml/min/mg; fm: 0.25 ± 0.053). THC at 500 nM (CLint: 4.7 ± 0.22 ml/min/mg) and 11-OH-THC at 50 nM (CLint: 2.4 ± 0.13 ml/min/mg) were predominately (fm: 0.99 and 0.80, respectively) metabolized by CYP3A in human fetal liver microsomes (n = 3). However, we did not observe significant depletion of THC/11-OH-THC in adult lung, first trimester, second trimester, or term placentae microsomes. Using PBPK modeling and simulation, these data could be used in the future to predict systemic and tissue THC/11-OH-THC exposure in healthy and special populations. SIGNIFICANCE STATEMENT: This is the first characterization and quantification of (-)-Δ9-tetrahydrocannabinol (THC) and (±)-11-hydroxy-Δ9-THC (11-OH-THC) depletion clearance by cytochrome P450 and UDP-glucuronosyltransferase enzymes in extrahepatic human tissues: intestine, fetal liver, lung, and placenta. These data can be used to predict, through physiologically based pharmacokinetic modeling and simulation, systemic and tissue THC/11-OH-THC exposure after inhalational and oral THC use in both healthy and special populations (e.g., pregnant women).
Assuntos
Citocromo P-450 CYP3A , Dronabinol , Adulto , Citocromo P-450 CYP2C9/metabolismo , Citocromo P-450 CYP3A/metabolismo , Dronabinol/análogos & derivados , Dronabinol/metabolismo , Feminino , Glucuronosiltransferase/metabolismo , Humanos , Microssomos Hepáticos/metabolismo , Gravidez , Difosfato de Uridina/metabolismoRESUMO
Cannabis e-cigarettes containing Δ8-tetrahydrocannabinol (Δ8-THC) produced synthetically from hemp-derived cannabidiol (CBD) have recently risen in popularity as a legal means of cannabis consumption, but questions surrounding purity and unlabeled additives have created doubts of their safety. Herein, NMR, GC-MS, and ICP-MS were used to analyze major components of 27 products from 10 brands, and it was determined none of these had accurate Δ8-THC labeling, 11 had unlabeled cutting agents, and all contained reaction side-products including olivetol, Δ4(8)-iso-tetrahydrocannabinol, 9-ethoxyhexahydrocannabinol, Δ9-tetrahydrocannabinol (Δ9-THC), heavy metals, and a novel previously undescribed cannabinoid, iso-tetrahydrocannabifuran.
Assuntos
Dronabinol/síntese química , Metais Pesados/química , Dronabinol/análogos & derivados , Dronabinol/química , Estrutura Molecular , Nebulizadores e VaporizadoresRESUMO
Objectives. To assess the popularity of an emergent drug, delta-8 tetrahydrocannabinol (THC), and compare interest levels between US states with or without legalized recreational cannabis. Methods. We used Google Trends to assess the growth of interest among delta-8 THC-related search terms from May 17, 2020, to May 9, 2021. We examined differences between states with or without legalized cannabis using state-level Google Trends data from February 13 to May 13, 2021, and policy data from the National Conference of State Legislatures. Results. Interest in delta-8 THC increased starting in mid-June 2020, with search volumes for delta-8 THC queries currently at 35% of the "marijuana" query. States where recreational cannabis is illegal had higher relative queries than did states with legalized recreational cannabis (52.3 vs 14.8; t = 40.9; P < .001). Conclusions. There has been rapid growth in interest in delta-8 THC. Findings between state policy contexts likely indicate delta-8 THC's role as a substitute good for delta-9 THC. Public Health Implications. Digital signals such as search volumes may point to an emergent use trend in the substance delta-8 THC. Further studies are needed to assess potential harms and correlates of delta-8 THC use. (Am J Public Health. 2022;112(2):296-299. https://doi.org/10.2105/AJPH.2021.306586).
Assuntos
Dronabinol/análogos & derivados , Internet , Legislação de Medicamentos , Antieméticos/uso terapêutico , Antagonistas de Dopamina/uso terapêutico , Dronabinol/uso terapêutico , HumanosRESUMO
Answer questions and earn CME/CNE Marijuana has been used for centuries, and interest in its medicinal properties has been increasing in recent years. Investigations into these medicinal properties has led to the development of cannabinoid pharmaceuticals such as dronabinol, nabilone, and nabiximols. Dronabinol is best studied in the treatment of nausea secondary to cancer chemotherapy and anorexia associated with weight loss in patients with acquired immune deficiency syndrome, and is approved by the US Food and Drug Administration for those indications. Nabilone has been best studied for the treatment of nausea secondary to cancer chemotherapy. There are also limited studies of these drugs for other conditions. Nabiximols is only available in the United States through clinical trials, but is used in Canada and the United Kingdom for the treatment of spasticity secondary to multiple sclerosis and pain. Studies of marijuana have concentrated on nausea, appetite, and pain. This article will review the literature regarding the medical use of marijuana and these cannabinoid pharmaceuticals (with emphasis on indications relevant to oncology), as well as available information regarding adverse effects of marijuana use.
Assuntos
Analgésicos não Narcóticos/uso terapêutico , Antieméticos/uso terapêutico , Dronabinol/análogos & derivados , Dronabinol/uso terapêutico , Maconha Medicinal/uso terapêutico , Neoplasias/tratamento farmacológico , Dor/tratamento farmacológico , Canadá , Canabidiol/uso terapêutico , Ensaios Clínicos como Assunto , Combinação de Medicamentos , Medicina Baseada em Evidências , Humanos , Esclerose Múltipla/tratamento farmacológico , Neoplasias/complicações , Dor/etiologia , Resultado do Tratamento , Reino Unido , Estados UnidosRESUMO
A first-in-class cannabinoid analog called lenabasum that is a CB2 agonist is being developed as an inflammation-resolving drug candidate. Thus far, specific therapeutic targets include scleroderma, cystic fibrosis, dermatomyositis, and lupus, all of which represent unmet medical needs. Two somewhat-independent molecular mechanisms for this type of action are here proposed. Both pathways initially involve the release of free arachidonic acid after activation of the CB2 receptor and phospholipase A2 by lenabasum. The pathways then diverge into a cyclooxygenase 2-mediated and a lipoxygenase-mediated route. The former leads to increased levels of the cyclopentenone prostaglandin 15-deoxy-Δ12,14-prostaglandin-J2 that can activate the NLPR3 inflammasome, which in turn releases caspase-3, leading to apoptosis and the resolution of chronic inflammation. The lipoxygenase-mediated pathway stimulates the production of lipoxin A4 as well as other signaling molecules called specialized proresolving mediators. These also have inflammation-resolving actions. It is not well understood under which conditions each of these mechanisms operates and whether there is crosstalk between them. Thus, much remains to be learned about the mechanisms describing the actions of lenabasum. SIGNIFICANCE STATEMENT: The resolution of chronic inflammation is a major unmet medical need. The synthetic nonpsychoactive cannabinoid lenabasum could provide a safe and effective drug for this purpose. Two putative molecular mechanisms are suggested to better understand how lenabasum produces this action. In both, different metabolites of arachidonic acid act as mediators.
Assuntos
Anti-Inflamatórios/farmacologia , Agonistas de Receptores de Canabinoides/farmacologia , Dronabinol/análogos & derivados , Animais , Ácido Araquidônico/metabolismo , Dronabinol/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Fosfolipases A2/metabolismo , Receptor CB2 de Canabinoide/metabolismo , Receptores de Formil Peptídeo/metabolismo , Receptores de Lipoxinas/metabolismoRESUMO
OBJECTIVE: The objective of this study was to assess the efficacy and safety of nabilone, a synthetic tetrahydrocannabinol analogue, as a treatment for non-motor symptoms (NMS) in Parkinson's disease (PD). METHODS: This was a phase II placebo-controlled, double-blind, parallel-group, enriched enrollment randomized withdrawal trial conducted at the Medical University Innsbruck. A random sample of 47 patients with PD with stable motor disease and disturbing NMS defined by a score of ≥4 points on the Movement Disorder Society - Unified PD Rating Scale-I (MDS-UPDRS-I) underwent open-label nabilone titration (0.25 mg once daily to 1 mg twice daily, phase I). Responders were randomized 1:1 to continue with nabilone or switch to placebo for 4 weeks (phase II). The primary efficacy criterion was the change of the MDS-UPDRS-I between randomization and week 4. Safety was analyzed in all patients who received at least one nabilone dose. RESULTS: Between October 2017 and July 2019, 19 patients received either nabilone (median dose = 0.75 mg) or placebo. At week 4, mean change of the MDS-UPDRS-I was 2.63 (95% confidence interval [CI] 1.53 to 3.74, p = 0.002, effect size = 1.15) in the placebo versus 1.00 (95% CI -0.16 to 2.16, p = 0.280, effect size = 0.42) in the nabilone-group (difference: 1.63, 95% CI 0.09 to 3.18, p = 0.030, effect size = 0.66). Seventy-seven percent of patients had adverse events (AEs) during open-label titration, most of them were transient. In the double-blind phase, similar proportions of patients in each group had AEs (42% in the placebo group and 32% in the nabilone group). There were no serious AEs. INTERPRETATION: Our results highlight the potential efficacy of nabilone for patients with PD with disturbing NMS, which appears to be driven by positive effects on anxious mood and night-time sleep problems. TRIAL REGISTRY: ClinicalTrials.gov (NCT03769896) and EudraCT (2017-000192-86). ANN NEUROL 2020;88:712-722.