Kobidze Giorgi, Möller Tristan, Wen Hui-Chung, Busardò Francesco Paolo, Thevis Mario
Department of Excellence-Biomedical Sciences and Public Health, Università Politecnica Delle Marche, Ancona, Italy.
Center for Preventive Doping Research, Institute of Biochemistry, German Sport University Cologne, Cologne, Germany.
Rapid Commun Mass Spectrom. 2025 Nov 30;39(22):e10124. doi: 10.1002/rcm.10124.
2-Chloro-4-[[(1R,2R)-2-hydroxy-2-methyl-cyclopentyl]amino]-3-methyl-benzonitrile (LY305) is a transdermal selective androgen receptor modulator (SARM) that has been under development as a potential therapeutic for conditions involving muscle wasting, osteoporosis, or hypogonadism. Due to its proven anabolic effects, its potential (and illicit) use in sport must be taken into consideration, necessitating information on its biotransformation for the implementation of adequate target analytes into routine doping control analytical procedures. In this study, the synthesis of LY305 and in vitro-derived metabolites of the SARM are described.
LY305 was synthesized by a Buchwald-Hartwig amination using 2-chloro-4-iodo-3-methylbenzonitrile and (1R,2R)-2-amino-1-methyl-cyclopentanol. The in vitro metabolism experiments were conducted using human liver microsomes (HLM) and the S9 fraction to allow for studying phase-I and phase-II biotransformations. For the detection and separation of LY305 and its metabolites, liquid chromatography-high-resolution tandem mass spectrometry was used.
Overall, 18 metabolites were detected, nine of which were identified as phase-I metabolites and an additional nine were attributed to phase-II conjugates. Metabolic reactions were as follows: hydroxylation, dehydrogenation, oxidation, oxidation and hydroxylation, O-glucuronidation, hydroxylation with subsequent glucuronidation, and bis-hydroxylation with subsequent glucuronidation.
The transdermal SARM LY305 was successfully synthesized and subjected to in vitro metabolic studies, yielding chromatographic and mass spectral data that support improving comprehensive anti-doping tests. To the best of our knowledge, no published experimental data exist that report on the in vitro metabolic profile of LY305, a substance that might undergo further (pre)clinical evaluation.
2-氯-4-[[(1R,2R)-2-羟基-2-甲基环戊基]氨基]-3-甲基苯甲腈（LY305）是一种经皮选择性雄激素受体调节剂（SARM），已被开发作为治疗肌肉萎缩、骨质疏松或性腺功能减退等病症的潜在药物。由于其已被证实的合成代谢作用，必须考虑其在体育领域的潜在（和非法）用途，这就需要了解其生物转化情况，以便在常规兴奋剂检测分析程序中确定合适的目标分析物。在本研究中，描述了LY305的合成及其体外衍生代谢物。
使用2-氯-4-碘-3-甲基苯甲腈和(1R,2R)-2-氨基-1-甲基环戊醇通过布赫瓦尔德-哈特维希胺化反应合成LY305。使用人肝微粒体（HLM）和S9组分进行体外代谢实验，以研究I相和II相生物转化。使用液相色谱-高分辨率串联质谱法检测和分离LY305及其代谢物。
总共检测到18种代谢物，其中9种被鉴定为I相代谢物，另外9种归因于II相结合物。代谢反应如下：羟基化、脱氢、氧化、氧化和羟基化、O-葡萄糖醛酸化、羟基化后葡萄糖醛酸化、双羟基化后葡萄糖醛酸化。
经皮SARM LY305成功合成并进行了体外代谢研究，获得了色谱和质谱数据，有助于改进全面的反兴奋剂检测。据我们所知，尚无已发表的实验数据报道LY305的体外代谢谱，该物质可能会进行进一步的（临床前）评估。
