Studies on the in vivo metabolism of the SARM YK11: Identification and characterization of metabolites potentially useful for doping controls

Abstract

A steroidal compound was recently detected in a seized black market product and was identified as (17α,20E)-17,20-[(1-methoxyethylidene) bis (oxy)]-3-oxo-19-norpregna-4,20-diene-21-carboxylic acid methyl ester (YK11). This compound is described to possess selective androgen receptor modulator-and myostatin inhibitor-like properties. As YK11 is an experimental drug candidate and a non-approved substance for humans, scientific data on its metabolism is scarce. Due to its steroidal backbone and the arguably labile orthoester-derived moiety positioned at the D-ring, substantial metabolic conversion in vivo was anticipated. To unambiguously detect urinary metabolites of YK11, an elimination study with six-fold deuterated YK11 was conducted. Post-administration specimens were analyzed using hydrogen isotope ratio mass spectrometry coupled to single quadrupole mass spectrometry to identify metabolites alongside basic mass spectrometric data. Further characterization of those metabolites relevant to sports drug testing was accomplished using gas chromatography-high resolution-high accuracy mass spectrometry. Fourteen deuterated urinary metabolites were detected comprising unconjugated, glucuronidated, and sulfoconjugated metabolites. As expected, no intact YK11 was observed in the elimination study urine samples. While the unconjugated metabolites disappeared within 24 hours post-administration, both glucuronidated and sulfated metabolites were traceable for more than 48 hours. The chemical structures of the two most promising glucuronidated metabolites (5β-19-nor-pregnane-3α,17β,20-triol and 5β-19-norpregnane-3α,17β-diol-20-one) were verified by in-house synthesis of both metabolites and confirmed by nuclear magnetic resonance analysis. In order to elucidate their potential in sports drug testing, both were successfully implemented into the currently applied analytical method for the detection of anabolic agents. KEYWORDS high resolution-high accuracy mass spectrometry, human metabolism, hydrogen isotope ratio mass spectrometry, SARM, YK11

Annual banned‐substance review – Analytical approaches in human sports drug testing

“…No drug candidate of the class of SARMs, however, has yet received clinical approval; nevertheless, the number of AAFs as the result of the presence of SARMs (or respective metabolites) in routine doping controls has continued to increase. The basis of the sensitive and specific detection of a subset of the enormous variety of SARM‐like drug candidates has been supported by sophisticated ITPs such as those reported by Ventura et al but also by identifying most adequate urinary metabolites . A comprehensive analytical assay covering a total of 15 SARMs was reported, validated for the determination of the target analytes in animal and human urine .…”

Identification of Trenbolone Metabolites Using Hydrogen Isotope Ratio Mass Spectrometry and Liquid Chromatography/High Accuracy/High Resolution Mass Spectrometry for Doping Control Analysis

“…Therefore, a GC Orbitrap system with an alternative ion source design favoring higher abundances of ions with larger m/z values was used, and the measured TMS derivatives further exhibited a reduced in-source fragmentation compared to acetates (Piper et al, 2018(Piper et al, , 2019. In addition, trimethylsilylation of steroidal analytes is routinely used, thus offering a platform for implementing potential new target analytes into sports drug testing methods.…”

Improved Synthesis and Determination of the Biologically Active Diastereomer of YK11

“…ARE-luciferase reporter assay in HEK293 cells suggested that the active constituent of YK11 was 2a. Although YK11 is sold via the internet as an anabolic steroid, it is an experimental drug candidate and a non-approved substance for humans [10][11][12]. It should be noted that YK11 has not been studied in humans or even animals.…”