Abstract
Selective Androgen Receptor Modulators (SARMs) have anabolic properties but less adverse effects than anabolic androgenic steroids. They are prohibited in both equine and human sports and there have been several cases of SARMs findings reported over the last few years. The aim of this study was to investigate the metabolite profile of the SARM ACP-105 (2-chloro-4-[(3-endo)-3-hydroxy-3-methyl-8-azabicyclo[3.2.1]oct-8-yl]-3-methylbenzonitrile) in order to find analytical targets for doping control. Oral administration of ACP-105 was performed in horses, where blood and urine samples were collected over a time period of 96 h. The in vivo samples were compared with five in vitro incubation models encompassing Cunninghamella elegans, microsomes and S9 fractions of both human and equine origin. The analyses were performed using ultra-high performance liquid chromatography coupled to high resolution Q ExactiveTM OrbitrapTM mass spectrometry (UHPLC-HRMS). A total of 21 metabolites were tentatively identified from the in vivo experiments, of which several novel glucuronides were detected in plasma and urine. In hydrolyzed urine, hydroxylated metabolites dominated. The in vitro models yielded several biotransformation products, including a number of monohydroxylated metabolites matching the in vivo results. The suggested analytical target for equine doping control in plasma is a dihydroxylated metabolite with a net loss of two hydrogens. In urine, the suggested targets are two monohydroxylated metabolites after hydrolysis with β-glucuronidase, selected both due to prolongation of the detection time and the availability of reference material from the in vitro models.
Highlights
Selective Androgen Receptor Modulators (SARMs) are a novel class of compounds that are androgen receptor ligands with anabolic properties similar to those of anabolic steroids in muscle and bone, but with a milder profile of adverse effects, since they are more tissue selective [1,2,3]
The identification and time profile analysis of metabolites formed from ACP-105 in both the in vivo and in vitro experiments were performed by UHPLC-HRMS full scan and MS/MS product ion scan modes
RAD-140 (2-chloro-4-[[(1R,2S)-1-[5-(4-cyanophenyl)-1,3,4-oxadiazol-2yl]-2-hydroxypropyl]amino]-3-methylbenzonitrile) and ACP-105 for analysis were purchased from MedChemExpress (Monmouth Junction, NJ, USA) and LGD-4033 (4-[(2R)-2-[(1R)-2,2,2-trifluoro1-hydroxyethyl]-1-pyrrolidinyl]-2-(trifluoromethyl)-benzonitrile) was purchased from Toronto Research Chemicals (North York, ON, Canada). β-nicotinamide adenine dinucleotide phosphate sodium salt hydrate (β-NADPH), uridine 5 -diphosphoglucuronic acid trisodium salt (UDPGA) and sabouraud dextrose broth were purchased from Sigma-Aldrich
Summary
Selective Androgen Receptor Modulators (SARMs) are a novel class of compounds that are androgen receptor ligands with anabolic properties similar to those of anabolic steroids in muscle and bone, but with a milder profile of adverse effects, since they are more tissue selective [1,2,3]. The pharmacological effects and limited side effects make them tempting to use as doping agents and they have been on the World Anti-Doping Agency’s Prohibited List since 2008 [6,7]. Many SARMs are readily available to purchase online and there have been several doping cases involving this substance class in both humans and horses in the past few years [9,10,11,12,13]. In vitro generated metabolites from microsome incubations may be used as reference material in both equine and human doping control in accordance with the International Laboratory Accreditation Cooperation document ILAC-G7 Accreditation Requirements and Operating Criteria for Horseracing Laboratories and the World Anti-Doping Agency’s International Standard for Laboratories [16,17]. It has been possible to scale up the method to purify metabolites for use as reference substances [21]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
