Chiral Plasma Pharmacokinetics of 3,4-Methylenedioxymethamphetamine and Its Phase I and II Metabolites Following Controlled Administration to Humans
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1521-009X/43/12/1864–1871$25.00 http://dx.doi.org/10.1124/dmd.115.066340 DRUG METABOLISM AND DISPOSITION Drug Metab Dispos 43:1864–1871, December 2015 Copyright ª 2015 by The American Society for Pharmacology and Experimental Therapeutics Chiral Plasma Pharmacokinetics of 3,4-Methylenedioxymethamphetamine and its Phase I and II Metabolites following Controlled Administration to Humans Andrea E. Steuer, Corina Schmidhauser, Yasmin Schmid, Anna Rickli, Matthias E. Liechti, and Thomas Kraemer Department of Forensic Pharmacology and Toxicology, Zurich Institute of Forensic Medicine, University of Zurich, Switzerland (A.E.S, C.S., T.K.); Division of Clinical Pharmacology and Toxicology, Department of Biomedicine and Department of Clinical Research, University Hospital Basel, Switzerland (Y.S., A.R., M.E.L.) Received July 16, 2015; accepted September 21, 2015 Downloaded from ABSTRACT Generally, pharmacokinetic studies on 3,4-methylenedioxymetham- dihydroxymethamphetamine (DHMA), and sulfate and glucuronide phetamine (MDMA) in blood have been performed after conjugate of 4-hydroxy-3-methoxymethamphetamine (HMMA) were identified, cleavage, without taking into account that phase II metabolites whereas free phase I metabolites were not detected. Stereoselec- represent distinct chemical entities with their own effects and tive differences in Cmax and AUC24 were observed with the following stereoselective pharmacokinetics. The aim of the present study was preferences: R>S for MDMA and DHMA 4-sulfate; S>R for 3,4- dmd.aspetjournals.org to stereoselectively investigate the pharmacokinetics of intact methylenedioxyamphetamine (MDA), DHMA 3-sulfate, and HMMA glucuronide and sulfate metabolites of MDMA in blood plasma after glucuronide; and no preference in Cmax for HMMA sulfate. R/S ratios a controlled single MDMA dose. Plasma samples from 16 healthy were >1 for all analytes after 24 hours, independent of the initial participants receiving 125 mg of MDMA orally in a controlled study chiral preference. These are the first data on chiral pharmacokinet- were analyzed using liquid chromatography–tandem mass spec- ics of MDMA phase II metabolites in human plasma in vivo after troscopy after chiral derivatization. Pharmacokinetic parameters controlled administration. The main human MDMA metabolites were of R-andS-stereoisomers were determined. Sulfates of 3,4- shown to be sulfate and glucuronide conjugates. at ASPET Journals on September 26, 2021 Introduction interaction with dopamine and/or serotonin metabolism, and for- 3,4-Methylenedioxymethamphetamine (MDMA, Ecstasy) is a rec- mation of reactive oxygen species (Carvalho et al., 2012). MDMA reational drug that acts by releasing dopamine, norepinephrine, and metabolites were also discussed to play a role in acute cardiovas- serotonin in the brain from presynaptic terminals via the respec- cular effects observed after MDMA consumption (Schindler et al., tive monoamine transporter (Hysek et al., 2012). In addition to 2014). amphetamine-like stimulant effects, MDMA increases empathy and As shown in Fig. 1, the main metabolic pathways of MDMA prosociality (Rietjens et al., 2012; Hysek et al., 2014) and exhibits observed in humans include O-demethylation to 3,4-dihydroxymeth- some hallucinogenic-like effects (Liechti et al., 2001). However, amphetamine (DHMA) mainly via the cytochrome P450 2D6 also proposed were severe acute poisonings, including tachycardia, (CYP2D6), followed by O-methylation mainly to 4-hydroxy-3- hypertension, hyperthermia, hyponatremia, and serotonin syn- methoxymethamphetamine (HMMA). DHMA is further sulfated by drome (Fallon et al., 1999; Kalant, 2001). Furthermore, MDMA sulfotransferases (SULT) to DHMA 3-sulfate and DHMA 4-sulfate. has been described as exhibiting neurotoxicity to serotonergic HMMA can be further conjugated by UDP-glucuronosyltransferases neurons (Monks et al., 2004; Easton and Marsden, 2006; McCann (UGT) or by SULT. A minor pathway includes the formation of 3,4- et al., 2008). methylenedioxyamphetamine (MDA) by N-demethylation followed MDMA metabolism may be responsible for neurotoxicity, presum- again by O-demethylation, O-methylation, and conjugation (Maurer, ably through the formation of glutathione adducts (Hiramatsu et al., 1996; Maurer et al., 2000; de la Torre et al., 2004). Analysis of urine 1990; Miller et al., 1997; Bai et al., 1999; Capela et al., 2009; Mueller samples after recreational MDMA consumption revealed MDMA and et al., 2009; Antolino-Lobo et al., 2010; Carvalho et al., 2012), primarily phase II metabolites (DHMA 3-sulfate, HMMA 4-sulfate, and HMMA 4-glucuronide) as major excretion products (Schwaninger et al., 2011a). Other metabolites could only be detected at negligible This work was supported financially by the Emma Louise Kessler Fund of the concentrations. Data on disposition and concentrations of phase II Zurich Institute of Forensic Medicine and the Swiss National Science Foundation metabolites in plasma relative to their unconjugated primary metabo- [320030_149493]. lites are not available so far. Generally, pharmacokinetic studies have dx.doi.org/10.1124/dmd.115.066340. used different conjugate cleavage procedures prior to analysis (Helmlin ABBREVIATIONS: AUC, Area under the curve; DHMA, 3,4-dihydroxymethamphetamine; HMMA, 4-hydroxy-3-methoxymethamphetamine; LOQ, limit of quantification; MDA, 3,4-methylenedioxyamphetamine; MDMA, 3,4-methylenedioxymethamphetamine; SULT, sulfotransferase; tmax, time of maximum concentration; UGT, UDP-glucuronosyltransferase. 1864 Plasma Pharmacokinetics of Chiral MDMA Metabolites 1865 et al., 1996; Fallon et al., 1999; Segura et al., 2001; Pizarro et al., R/S-DHMA sulfates, R/S-HMMA 4-sulfate, and diastereomers of HMMA 2002; Kolbrich et al., 2008; Shen et al., 2013) only allowing quantifi- 4-glucuronides were synthesized as described (Schwaninger et al., 2009, 2011c). cation of sum values for unconjugated and conjugated metabolites. Water was purified with a Millipore filtration unit and acetonitrile and methanol of However, the different conjugates represent individual chemical entities high-performance liquid chromatography grade were obtained from Fluka (Buchs, with their own potential risks of adverse effects, toxicity, or drug Switzerland). All other chemicals used were from Merck (Zug, Switzerland) and of the highest grade available. interactions. Controlled Oral MDMA Administration. We used plasma samples from Chemically, MDMA possesses a chiral center with pharmacody- a double-blind, placebo-controlled, crossover study with four experimental test namic and pharmacokinetic differences for the R-and S-enantiomers sessions (placebo-placebo, bupropion–placebo, placebo-MDMA, and bupropion- (Fallon et al., 1999; Kalant, 2001; Kraemer and Maurer, 2002; Peters MDMA) that were performed in a counterbalanced order according to a Latin et al., 2003; Pizarro et al., 2004; Peters et al., 2005). Whereas the square randomization design as described in detail by Schmid et al. (2015). The S-MDMA enantiomer primarily causes the described stimulating clinical study was conducted at the University Hospital of Basel in accordance effects, R-MDMA produces more hallucinogenic effects (de la Torre with the Declaration of Helsinki and International Conference on Harmonization et al., 2004). Elimination of S-MDMA was shown to be faster Guidelines in Good Clinical Practice and with approval by the Ethics Commit- compared with the R-enantiomer (Fallon et al., 1999; Kalant, 2001; tee of the Canton of Basel, Switzerland, and the Swiss Agency for Therapeu- Kraemer and Maurer, 2002; Peters et al., 2003, 2005; Pizarro et al., tic Products (Swissmedic). The study was registered at ClinicalTrials.gov (NCT01771874). All subjects provided written informed consent and were 2004) most probably owing to stereoselective metabolism that has paid for their participation. been extensively studied in vitro (Meyer et al., 2008; Meyer and Plasma samples from 16 healthy Caucasian subjects (eight men and eight Downloaded from Maurer, 2009; Schwaninger et al., 2009, 2011b). As not only women, age 20–27, body mass index of 22.7 6 2.1 kg/m2) from the placebo- stereoselectivity of MDMA itself but also of its primary metabolite MDMA session were analyzed in the Institute of Forensic Pharmacology and DHMA is discussed in terms of (neuro-)toxicity (Felim et al., 2010; Toxicology Zurich as described in Sample Preparation and Analysis following. Martinez et al., 2012), the further metabolic fate of DHMA and Placebo was administered at 8:00 AM and MDMA (125 mg p.o.) at 10:00 AM. resulting stereoselectivities of metabolites is of interest. Comprehen- Blood samples were collected at 0, 0.33, 0.67, 1, 1.5, 2, 2.5, 3, 4, 6, 8, and sive controlled pharmacokinetic studies on the stereoselectivity of 24 hours after MDMA administration. MDMA and all relevant metabolites—including phase II metabolites— Sample Preparation and Analysis. Blood plasma samples were analyzed dmd.aspetjournals.org are still missing but are needed to understand the entire stereoselective using a Thermo Fisher Ultimate 3000 UHPLC system coupled to an ABSciex disposition of MDMA and its differences in pharmacological effects and QTRAP 5500 in positive electrospray ionization (ESI) mode. Chromatography kinetics. was performed on a Phenomenex Kinetex C18 column after chiral derivatization with Marfey’s reagent (N-(2,4-dinitro-5-fluorophenyl) L-valinamide) as de- Therefore, the aim of this study was first to assess the extent of scribed in detail by Steuer et al. (2015). Briefly,