Characterization of a non‐approved selective androgen receptor modulator drug candidate sold via the Internet and identification of <i>in vitro</i> generated phase‐I metabolites for human sports drug testing

Selective androgen receptor modulators (SARMs) are an emerging class of therapeutics targeted to cachexia, sarcopenia, and hypogonadism treatment. LGD-4033 is a SARM which has been included on the Prohibited List annually released by the World Anti-Doping Agency (WADA). The aim of the present work was the investigation of the metabolism of LGD-4033 in a human excretion study after administration of an LGD-4033 supplement, the determination of the metabolites' excretion profiles with special interest in the determination of its long-term metabolites, and the comparison of the excretion time of the phase I and phase II metabolites. The results were also compared to those derived from previous LGD-4033 studies concerning both in vitro and in vivo experiments. Supplement containing LGD-4033 was administered to one human male volunteer and urine samples were collected up to almost 21 days. Analysis of the hydrolyzed (with β-glucuronidase) as well as of the non-hydrolyzed samples was performed using liquid chromatography-high resolution mass spectrometry (LC-HRMS) in negative ionization mode and revealed that, in both cases, the two isomers of the dihydroxylated metabolite (M5) were preferred target metabolites. The glucoconjugated parent LGD-4033 and its gluco-conjugated metabolites M1 and M2 can be also considered as useful target analytes in non-hydrolyzed samples. The study also presents two trihydroxylated metabolites (M6) identified for the first time in human urine; one of them was recently reported in an LGD-4033 metabolism study in horse urine and plasma. KEYWORDS human doping, LGD-4033, liquid chromatography-mass spectrometry, SARM, selective androgen receptor modulator

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confidence: 99%

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confidence: 99%

Human in vivo metabolism study of LGD‐4033

Fragkaki

Sakellariou

Kiousi

et al. 2018

“…Due to the ease of availability over the internet, the misuse of LGD-4033 has already been reported in the human sports arena, 8 highlighting the need for routine methods that allow the best possible detection of these novel substances in doping control laboratories. 9,10 Numerous other SARMs have also been studied in the human. As a result, studies have been carried out utilising human liver microsomes to investigate the human metabolism of LGD-4033 products in vitro.…”

Section: -Yl)-2-(trifluoromethyl)-benzonitrile) Is a Member Of A CLmentioning

confidence: 99%

Investigation of the metabolism of the selective androgen receptor modulator LGD‐4033 in equine urine, plasma and hair following oral administration

Cutler¹,

Viljanto²,

Hincks³

et al. 2020

LGD-4033 is one of a number of selective androgen receptor modulators (SARMs) that are being developed by the pharmaceutical industry to provide the therapeutic benefits of anabolic androgenic steroids, without the less desirable side effects.Though not available therapeutically, SARMs are available for purchase online as supplement products. The potential for performance enhancing effects associated with these products makes them a significant concern with regards to doping control in sports. The purpose of this study was to investigate the metabolism of LGD-4033 in the horse following oral administration, in order to identify the most appropriate analytical targets for doping control laboratories. LGD-4033 was orally administered to two Thoroughbred horses and urine, plasma and hair samples were collected and analysed for parent drug and metabolites. LC-HRMS was used for metabolite identification in urine and plasma. Eight metabolites were detected in urine, five of which were excreted only as phase II conjugates, with the longest detection time being observed for di-and tri-hydroxylated metabolites. The parent compound could only be detected in urine in the conjugated fraction. Seven metabolites were detected in plasma along with the parent compound where mono-hydroxylated metabolites provided the longest duration of detection. Preliminary investigations with hair samples using LC-MS/MS analysis indicated the presence of trace amounts of the parent compound and one of the mono-hydroxylated metabolites. In vitro incubation ofLGD-4033 with equine liver microsomes was also performed for comparison, yielding 11 phase I metabolites. All of the metabolites observed in vivo were also observed in vitro.

“…Cases of illicit use of SARMs in human sports have been reported. [66,67] SARMs are also of great importance in equine doping control, and some studies concerning the in vivo metabolism pattern has not been referred in this species. [68] Hansson et al [69,70] described the urinary metabolites of the SARMs S1, S4, and S22 in horses after intravenous injection using UHPLCquadrupole Time-of-Flight-MS (UHPLC-QToF-MS), reporting a significant number of metabolites for each of the above substances.…”

Section: Nickelmentioning

confidence: 99%

Challenges in detecting substances for equine anti‐doping

Fragkaki

Kioukia‐Fougia

Kiousi

et al. 2017

The artificial increase of the physical capability of horses using drugs is well known in racing and other equine sports. Both illicit and therapeutic substances are regarded as prohibited substances in competition in most countries. Some countries make distinctions for a few, specific drugs which are, however, allowed for use in other countries. The primary objective in the case of doping control is the detection of any trace of drug exposure, either parent drug or any of its metabolites, using the most powerful analytical methods which are generally based on chromatographic/mass spectrometric techniques. Of major concern in horseracing is the absence of a single organization regulating the anti-doping framework; instead of this, individual racing authorities provide rules and regulations often resulting in variations in the applied doping control programmes of different countries. The aim of this paper is to review the recent literature (approximately from 2012 to mid-2016) to highlight the numerous and diverse challenges faced in doping control of racing and equestrian sports, including the detection of designer drugs (anabolic steroids or stimulants) and of other emerging prohibited substances, such as peptides and noble gases in horse urine and plasma. Moreover, the application of 'omics' techniques (especially of metabolomics) deserves attention for establishing possible fingerprints of drug abuse as well as the evolution of instrumental analysis resulting a powerful ally in the fight against doping in equine sports.