Anticancer activity of Δ^(9)-tetrahydrocannabinol and cannabinol in vitro and in human lung cancer xenograft

Objective:To investigate the effects of Δ^(9)-tetrahydrocannabinol,the principal psychoactive compound of Cannabis sativa,and cannabinol,a Δ^(9)-tetrahydrocannabinol degradative product,on human non-small cell lung cancer cells.Methods:Δ^(9)-Tetrahydrocannabinol and cannabinol were tested for anticancer activity in human non-small cell lung cancer(A549)cells.The effects on cell proliferation,apoptosis,and phosphorylation profiles were examined.The effects of Δ^(9)-tetrahydrocannabinol and cannabinol on tumor growth were also investigated using a xenograft nude mouse model.Apoptosis and targeted phosphorylation were verified by immunohistochemistry.Results:Δ^(9)-Tetrahydrocannabinol and cannabinol significantly inhibited cell proliferation and increased the number of apoptotic cells in a concentration-dependent manner.The Δ^(9)-tetrahydrocannabinol-and cannabinol-treated cells had lower levels of phosphorylated protein kinase B[AKT(S473)],glycogen synthase kinase 3 alpha/beta,and endothelial nitric oxide synthase compared to the controls.The study of xenograft mice revealed that tumors treated with 15 mg/kg Δ^(9)-tetrahydrocannabinol or 40 mg/kg cannabinol were significantly smaller than those of the control mice.The tumor progression rates in mice treated with 15 mg/kg Δ^(9)-tetrahydrocannabinol or 40 mg/kg cannabinol were significantly slower than in the control group.Conclusions:These findings indicate that Δ^(9)-tetrahydrocannabinol and cannabinol inhibit lung cancer cell growth by inhibiting AKT and its signaling pathways,which include glycogen synthase kinase 3 alpha/beta and endothelial nitric oxide synthase.

大麻二酚类药物在我国面临的问题与探讨

目的:大麻二酚是大麻植物中存在的一种大麻素类物质。欧美国家已经批准其用于癫痫的治疗。在我国,大麻二酚作为四氢大麻酚的同分异构体,按照第一类精神药品进行监管,尚未有此类药品上市。本研究旨在探讨大麻二酚类药物研发、生产、上市过程中存在的监管问题,提出可行性监管途径,促进此类药品在国内的研发和应用。方法:通过梳理《麻醉药品和精神药品管理条例》《关于麻醉药品和精神药品实验研究管理规定的通知》,以及云南、黑龙江两省的地方规定,结合大麻二酚成瘾性和医疗价值,探讨工业大麻种植、大麻二酚类药物实验研究立项和生产中存在的监管问题,并提出针对性建议。结果与结论:大麻二酚类药物不具有成瘾性,在神经精神疾病中具有重要临床价值。目前监管法规对大麻二酚类药物实验研究立项存在一定的限制。工业大麻种植监管存在一定的空白。建议在借鉴国内已有经验的基础上,平衡好药品的可及性和滥用风险,有计划、有步骤地推动大麻二酚类药物的发展,促进其在国内的研发和临床应用,助力神经精神疾病的临床治疗。

S4A-1 Cognitive and Pharmacological Assays to Characterize both the Deleterious and Medicinal Effects of Cannabinoids in Monkeys

Rationale:Marijuana is the most commonly used illicit drug in the United States with recent surveys estimating over 22 million current users.However,there is growing acceptance of its recreational use,evident by successful efforts to decriminalize and,in some states,legalize use.In addition,although the full medicinal value of cannabis is not yet understood,such cannabinergic effects are of known benefit in the palliative care of anorectic patients undergoing chemotherapy or suffering debilitating conditions such as AIDS or Alzheimer’s disease and,as well,may be an effective analgesic under certain painful conditions.Objective:This current state of affairs has led to a broadening interest in the clinical utility of drugs that target the endocannabinoid system.In this regard,however,delta-9-tetrahydrocannabinol(THC),the primary psychoactive ingredient in marijuana,is generally acknowledged to produce some unwanted effects in humans.These include deleterious effects on several types of complex behavior,especially related to learning,memory,and vigilance.Unfortunately,synthetic cannabinoid agonists appear to share these adverse side effect profiles.Recently,however,a greater understanding has developed of the two primary endocannabinoid neurotransmitters,anandamide and 2-arachidonoylglycerol(2-AG)and,as well,ligands that inhibit the activity of fatty acid amide hydrolase(FAAH)and monoacylglycerol lipase(MGL),which can indirectly increase the concentrations of,respectively,anandamide and 2-AG.Methods:Employing operant techniques in nonhuman primates such as drug discrimination,emetic observation,nociception assays,and touchscreen-based models of learning and memory,this presentation will highlight recent advances in the understanding of THC’s effects on complex behavioral processes and,as well,efforts to develop drugs that engage the cannabinergic system and retain medicinal value,yet produce lesser adverse psychoactive effects.Results:Synthetic and phyto-cannabinoids produce highly consistent effects on most behavioral endpoints despite efficacy differences identified in vitro.However,engaging the endogenous cannabinoid system,either via exogenous administration of endocannabinoids or indirect endocannabinoid elevation via inhibition of catabolic enzyme inhibition,produced lesser disruptive effects on cognitive behavior with similar effectiveness of medicinal endpoints(e.g.,analgesia and emesis).Conclusion:Engagement of the endogenous cannabinoid system via endocannabinoid administration or indirect elevation via FAAH and/or MGL inhibition may represent a therapeutic advantage in clinical settings relative to THC or other typical cannabinoid agonists.

Effects of Gibberellic Acid on Primary Terpenoids and △^9 -Tetrahydrocannabinol in Cannabis sativa at Flowering Stage

Plants synthesize an astonishing diversity of isoprenoids, some of which play essential roles in photosynthesis, respiration, and the regulation of growth and development. Two independent pathways for the biosynthesis of isoprenoid precursors coexist within the plant cell： the cytosolic mevalonic acid （MVA） pathway and the plastidial methylerythritol phosphate （MEP） pathway. However, little is known about the effects of plant hormones on the regulation of these pathways. In the present study we investigated the effect of gibberellic acid （GA3） on changes in the amounts of many produced terpenoids and the activity of the key enzymes, 1-deoxy-D-xylulose 5-phosphate synthase （DXS） and 3-hydroxy-3-methylglutaryl coenzyme A reductase （HMGR）, in these pathways. Our results showed GA3 caused a decrease in DXS activity in both sexes that it was accompanied by a decrease in chlorophylls, carotenoids and Ag-tetrahydrocannabinol （THC） contents and an increase in α-tocopherol content. The treated plants with GA3 showed an increase in HMGR activity. This increase in HMGR activity was followed by accumulation of stigmasterol and β-sitosterol in male and female plants and campestrol in male plants. The pattern of the changes in the amounts of sterols was exactly similar to the changes in the HMGR activity. These data suggest that GA3 can probably influence the MEP and MVA pathways oppositely, with stimulatory and inhibitory effects on the produced primary terpenoids in MVA and DXS pathways, respectively.

Role of cannabinoids and the endocannabinoid system in modulation of diabetic cardiomyopathy

Diabetic complications,chiefly seen in long-term situations,are persistently deleterious to a large extent,requiring multi-factorial risk reduction strategies beyond glycemic control.Diabetic cardiomyopathy is one of the most common deleterious diabetic complications,being the leading cause of mortality among diabetic patients.The mechanisms of diabetic cardiomyopathy are multi-factorial,involving increased oxidative stress,accumulation of advanced glycation end products(AGEs),activation of various pro-inflammatory and cell death signaling pathways,and changes in the composition of extracellular matrix with enhanced cardiac fibrosis.The novel lipid signaling system,the endocannabinoid system,has been implicated in the pathogenesis of diabetes and its complications through its two main receptors:Cannabinoid receptor type 1 and cannabinoid receptor type 2,alongside other components.However,the role of the endocannabinoid system in diabetic cardiomyopathy has not been fully investigated.This review aims to elucidate the possible mechanisms through which cannabinoids and the endocannabinoid system could interact with the pathogenesis and the development of diabetic cardiomyopathy.These mechanisms include oxidative/nitrative stress,inflammation,accumulation of AGEs,cardiac remodeling,and autophagy.A better understanding of the role of cannabinoids and the endocannabinoid system in diabetic cardiomyopathy may provide novel strategies to manipulate such a serious diabetic complication.

The dimerization of ⊿~9-tetrahydrocannabinolic acid A(THCA-A)

The renewed interest in dimeric salicylates as broad-spectrum anti-inflammatory and antidiabetic agents provided a rationale to investigate the dimerization of the substituted salicylate D9-tetrahydrocannabinolic acid(THCA-A, 3 a) as a strategy to solve its instability to decarboxylation and to generate analogues and/or pro-drugs of this native pre-cannabinoid. Activation of the carboxylic group with the DCC-HOBt-DMAP protocol afforded a high yield of the OBt ester 4, that was next converted into the highly crystalline di-depsidic dimer 5 upon treatment with DMAP. The mono-depsidic dimer 6 was also formed when the reaction was carried out with partially decarboxylated THCA-A samples. The structure of the depsidic dimers was established by spectroscopic methods and by aminolysis of 5 into the pre-cannabinoid amide 7. Both dimers showed excellent shelf stability and did not generate significant amounts of D9-THC upon heating. However, only the didepsidic dimer 5 activated PPAR-g, the major target of pre-cannabinoids, but strong binding to serum proteins abolished this activity, also shielding it from the action of esterases.

The Detection of THCA Using 2-Dimensional Gas Chromatography-Tandem Mass Spectrometry in Human Fingernail Clippings: Method Validation and Comparison with Head Hair

Marijuana use as well as abuse is a significant public health and public safety concern in the United States and using hair to identify marijuana users and abusers has been gaining acceptance in a number of venues including workplace, court ordered, and substance abuse treatment monitoring. After the presentation of a fully validated 2-dimensional gas chromatography-tandem mass spectrometry method for the detection of 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THCA), the chief metabolite of the main psychoactive compound in marijuana, Δ9-tetrahydrocannabinol (THC), we evaluated the usefulness of fingernail clippings as an alternative specimen type to hair by the analysis of a set of 60 matched pairs of head hair and fingernail clippings. The limit of detection was 10 fg/mg, the limit of quantitation was 20 fg/mg, and the assay was linear from 20 fg/mg to 500 fg/mg. The intra- and inter-assay imprecision and bias studies at 4 different concentrations (50, 100, 500, and 1000 fg/mg) were acceptable where all % Target observations were within 16% of their expected concentrations and all %CV calculations were less than 13.5%. THCA was detectable in more fingernail specimens (53.3%) than hair specimens (46.7%) and the mean concentrations in nails were on average 4.9 times higher than in hair (1813 fg/mg and 364 fg/mg, respectively). The THCA concentrations in hair and nail were strongly associated (r = 0.974, P < 0.01, n = 60) and the association was significant. The study demonstrated that fingernail clippings are a suitable alternative specimen type to hair to monitor for marijuana use and abuse.

Chemical and physical variations of cannabis smoke from a variety of cannabis samples in New Zealand

Studies have compared the chemical properties of tobacco smoke to those of cannabis smoke,with the objective of identifying the chemical attributes responsible for the mutagenicity and carcinogenicity of cannabis smoke.Comparative studies have included small sample sizes and produced conflicting results.The aim of this study was to assess the major chemical and physical variations of cannabis smoke across a range of cannabis samples of different potencies and origins,sourced from the illegal market in New Zealand.Twelve cannabis samples were studied ranging from 1.0%to 13.4%delta-9-tetrahydrocannabinol(D9THC)content.A smoking machine was used to smoke“joints”(cannabis cigarettes)and the chemical/physical properties of the smoke assessed.The chemical constituents of the smoke extracts were analysed by gas chromatography/mass spectrometry.A range of different chemical constituents(in addition to D9THC)were identified and their concentrations estimated.Terpenoids were identified as the major variable in cannabis smoke,showing a 40-fold range in total terpenoid content.Analysis of the total particulate matter showed that significantly different levels of particulate matter were produced between the different cannabis samples,ranging from 14.6 to 66.3 mg/g of cannabis smoked.The D9THC delivery efficiency during smoking was also investigated and produced consistent results showing a mean and median of 12.6%and 10.8%,respectively,of the theoretically available D9THC(ranging from 7.2%to 28.0%).