Correlation of Nabiximols Dose to Steady-State Concentrations of Cannabinoids in Urine Samples from Patients with Multiple Sclerosis.

Therapeutic drug monitoring of Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD) is based on a complex procedure and is therefore not possible in most laboratories, especially in emergency cases. This work addresses the question of whether therapeutic drug monitoring of nabiximols can be performed using an immunological urine-based test system for cannabinoid abuse. Seventeen patients with multiple sclerosis were included in this study. Administered doses of nabiximols were correlated with immunologically determined urine concentrations of cannabinoids using the DRITM Cannabinoid (THC) Assay. Significant correlations with the administered nabiximols doses were found for creatinine-normalized urine concentrations of cannabinoids without (r = 0.675; p = 0.0015) and after (r = 0.650; p = 0.0044) hydrolysis, as well as for gas-chromatography-coupled mass spectrometry (GC/MS)-measured concentrations of the THC metabolite 11-nor-9-carboxy-Δ9-THC (THC-COOH) in urine samples (r = 0.571; p = 0.0084) by Pearson's correlation. In addition, doses were significantly correlated with plasma THC-COOH concentrations (r = 0.667; p = 0.0017) measured by GC/MS. Simple immunological cannabinoid measurements in urine samples could provide an estimate of nabiximols dosage, although the correlations obtained here were weak because of the small number of patients observed. Longitudinal monitoring of individual patients is expected to exhibit good results of therapeutic drug monitoring of nabiximols.

Metabolism of TEGDMA and HEMA in human cells.

Previous in vivo studies have shown that the comonomers triethylene glycol dimethacrylate (TEGDMA) and 2-hydroxyethyl methacrylate (HEMA) from dental materials can be metabolised to CO(2) by two postulated pathways: an epoxide and a valine pathway. In the epoxide pathway the formation of pyruvate is postulated and in valine pathway the formation of l-malate. The aim of this investigation was to quantify the formation of the intermediates pyruvate and l-malate to show which pathway may be preferred in A549 cells. Therefore A549 cells were incubated with TEGDMA or HEMA (with a tracer dose 14C-TEGDMA or 14C-HEMA) and afterwards 14C-TEGDMA or 14C-HEMA, 14C-methacrylate, 14C-l-malate and 14C-pyruvate were identified and quantified by thin layer chromatography at different time intervals from the extracellular and intracellular fluid. Our results show that in the metabolism of both comonomers more 14C-pyruvate was formed compared to 14C-l-malate for 14C-HEMA metabolisation during 0.5 up to 6h after 14C-HEMA exposure and for 14C-TEGDMA metabolisation >4h after 14C-TEGDMA exposure. Therefore the epoxide pathway with formation of the epoxy-intermediate 2,3-epoxymethacrylic acid is the main route of metabolisation of HEMA and TEGDMA.

Glutathione synthesis against oxidant injury by peroxides in two alveolar epithelial cell lines.

The D- and L-forms of N-acetylcysteine (NADC, NAC) were tested in antagonizing the toxicity mediated by hydrogen peroxide (H(2)O(2)) or tertiary butyl hydroperoxide (tBHP) in two lung cell lines to assess the effectivity of glutathione synthesis against peroxides. Toxicity was assessed by methionine incorporation, total glutathione content, and glutathione disulfide to glutathione ratio. NAC or NADC, at 2 mmol/L, increased cellular glutathione to about 1.5- or 3-fold (NAC) and 1.1- or 1.2-fold (NADC) in A549 or L2 cells, respectively, as compared to naive cells. H(2)O(2)-mediated toxicity was decreased by NADC (as compared to controls), but increased slightly with NAC, whereas tBHP-mediated toxicity was decreased both by NAC and NADC. However, when compared to controls, NADC was an effective antidote against tBHP in L2 cells only. Dexamethasone pretreatment increased toxicity of H(2)O(2) and tBHP in L2 cells, but did not affect the antioxidative efficacy of NAC/NADC. Antidotal properties of NAC/NADC were similar in both cell lines, despite significant differences of the glutathione redox system in both situations. Hence, it is concluded that direct antioxidative properties of NAC and NADC is a main antagonizing factor in H(2)O(2)-based toxicity but not in tBHP-mediated toxicity. Enhancement of glutathione biosynthesis decreased toxicity of tBHP, but not of H(2)O(2) in 2 pulmonary cell lines.

Glucocorticoid pretreatment increases toxicity due to peroxides in alveolar epithelial-like cell lines.

In previous experiments an increase in zinc-mediated toxicity was found after pretreatment of alveolar epithelial type II-like cells with glucocorticoids. In this work toxicity of two peroxides (tertiary butyl hydroperoxide [tBHP], hydrogene peroxide [HP]) was assessed in L2 and A549 cells compared to dexamethasone (DEX) pretreated cells. Pretreatment of cells with 7.5micromol/l DEX for 72h decreased cellular glutathione content in both cell lines. Furthermore compared to not pretreated cells toxicity of both peroxides was increased in A549 cells, while in L2 cells only toxicity of tBHP was significantly increased by the glucocorticoid pretreatment. HP toxicity only showed a tendency to be increased in L2 cells after DEX pretreatment. The results point to a glucocorticoid-dependent increased oxidative stress of alveolar epithelial type II cells as antagonised by antioxidative enzymes such as catalase and/or preferentially by the glutathione system. This furthermore should be considered for all glucocorticoid applications in vivo as well.

Changes in the glutathione system of lung cell lines after treatment with hydrocortisone.

Administration of anti-inflammatory glucocorticoids is a drug option in the therapy of acute respiratory distress syndrome (ARDS), according to present pathophysiological concepts. Surprisingly, glucocorticoids failed to show beneficial effects. This failure is not understood. In this investigation changes in the glutathione system due to hydrocortisone were found to consist of glutathione depletion and lowered glutathione reductase activities in alveolar epithelial type II cells, contrasted with unchanged activities in a fibroblast-like lung cell line. The glutathione system is thought to be the most important cellular antioxidative system and therefore alveolar epithelial type II cells might be more susceptible to oxidative stress after glucocorticoid treatment. As alveolar epithelial type II cells may be important targets in ARDS, because of their functions (stem cells of type I epithelial cells; surfactant synthesis), these changes might provide an explanation for the failure of glucocorticoids. In the present experiments the capability of hydrocortisone-treated alveolar epithelial type II cells to synthesise glutathione was found to be cysteine dependent at physiological concentrations. Transposing this observation to the in vivo situation, it might be expected that glucocorticoid efficacy in ARDS therapy requires co-administration of substances that increase glutathione synthesis, e.g. N-acetylcysteine.