Elsevier

Toxicology Letters

Volume 164, Issue 1, 20 June 2006, Pages 16-23
Toxicology Letters

Tetrahydrogestrinone is a potent but unselective binding steroid and affects glucocorticoid signalling in the liver

https://doi.org/10.1016/j.toxlet.2005.11.006Get rights and content

Abstract

Tetrahydrogestrinone (THG) is a steroid recently identified to be misused as doping agent. However, the knowledge on functions of this substance in humans or animal models is rather limited. Therefore, it was our aim to further characterize the pharmacological profile of THG and identify potential adverse side effects. THG was synthesized, the purity was confirmed and its biological activity was tested. The potency of THG to transactivate AR dependent reporter gene expression was two orders of magnitude lower compared to dihydrotestosterone. THG binds with high affinity but unselective to the androgen (AR), progesterone (PR), glucocorticoid (GR) and mineralocorticoid (MR) receptor. Treatment of orchiectomised rats with THG resulted in a stimulation of prostate, seminal vesicle and levator ani muscle, indicating androgenic and anabolic properties. In the liver THG, in contrast to testosteronepropionate (TP), down regulates the expression of the GR dependent tyrosine aminotransferase gene (TAT).

In summary, our results demonstrate that THG is not a specific AR agonist. THG exhibits a high binding affinity to all tested steroid hormone receptors and binds with highest affinity to the GR. Our in vivo data are indicative of an anabolic and androgenic potency of THG, but the repression of TAT demonstrates that THG also interferes with the glucocorticoid hormone system. Therefore, it is conceivable that an intake will result in adverse side effects.

Introduction

Anabolic-androgenic steroids (AAS) have been banned in sport since 1980. According to the worldwide doping control statistics they are the most frequently detected substances (WADA, 2004). Since the ban of AAS, athletes and their entourage have always tried to find new products and applications to evade the doping controls. Rumours of new steroids, produced and used specifically to escape doping tests in sport have been spread since more than 15 years. In 2002, Catlin et al. found a never-marketed steroid called norbolethone in two urine samples. In 2003, a track and field coach sent a syringe with an oily substance to the United States Anti-Doping Agency. The substance proved to be tetrahydrogestrinone (THG, Fig. 1), the first true “designer steroid”, designed, synthesized and distributed solely as an undetectable doping agent (Catlin et al., 2004). Even more recently, a new “designer steroid”, desoxymethyltestosterone (DMT) or “madol” was discovered, and its detection in urine described (Sekera et al., 2005). This reflects an alarmingly sophisticated, illicit manufacturing facility and an underground network to distribute these substances. There exists an unknown number of abusers of such substances in sport, even though their anabolic-androgenic potency and hormonal properties are not yet resolved. For doping prevention and control it is essential to know the effects and side effects of these “designer steroids” to plan informational and educational campaigns for athletes.

Therefore, the aim of our study was to further characterize the pharmacological profile of THG and identify potential adverse side effects. THG was synthesized and its identity, purity and biological activity was tested. The specificity of a steroid for the binding to distinct steroid hormone receptors is a general issue with respect to potential side effects. Therefore, the binding affinity of THG to the androgen (AR) but also to the mineralocorticoid (MR), glucocorticoid (GR) and progesterone receptor (PR) was determined. To verify if the detected binding preferences to the distinct steroid hormone receptors result in a biological activity in vivo, orchiectomised rats were treated for 10 days with equimolar doses of THG and the reference compound testosteronepropionat (TP). Effects on prostate, seminal vesicle, heart and the levator ani muscle were investigated. Furthermore the regulation of the GR dependent tyrosine aminotransferase gene expression (TAT) was assessed in the liver.

Section snippets

Substances

Androstendione and testosteroneproprionate were provided by the Institute of Biochemistry, German Sport University Cologne. Purity of the substances was verified by mass spectrometry. Dihydrotestosterone (DHT) was obtained by Sigma–Aldrich (Deisenhofen, Germany). R1881, Aldosterone, Dexametasone and Progesterone were provided by the Schering AG (Berlin, Germany).

Synthesis of THG

THG was prepared from 500 mg (1.6 mmol) of gestrinone, which was dissolved in 250 ml of methanol in a round-bottomed flask. 10 mg of PtO2

Results

THG was synthesized as described in material and methods. Purity and authenticity was determined by HPLC-UV, and characterization of THG was performed by means of nuclear magnetic resonance spectroscopy as well as electrospray ionization mass spectrometry (Thevis et al., 2005). THG was obtained at a purity of >97% and a yield of 78% of theory.

To test the biological activity of the synthesized THG a yeast-based androgen receptor assay was performed as described in material and methods. THG

Discussion

The discovery and possible widespread use of the steroid THG has unleashed concerns over the public health implications of abuse of such substances (Handelsman et al., 2004). THG represent a new class of anabolic steroids, which are synthesized in underground labs and never have been characterized toxicologically and pharmacologically before it was used in humans. As a strategy for future doping prevention and control the aim of this study was to pharmacologically characterize this substance

Acknowledgments

We thank Perter Muhn, Schering AG Berlin for his support, realising the receptor binding assay. We also thank J. Seibel and T. Hertrampf for their assistance performing the animal experiments. This study was founded by the Federal Office of Sports, Magglingen, Switzerland.

References (17)

  • M.S. Miller et al.

    In vivo inhibition of glucocorticoid-inducible gene expression by dimethylnitrosamine in rat liver

    Biochem. Pharmacol.

    (1993)
  • L.D. Boada et al.

    Evaluation of acute and chronic hepatotoxic effects exerted by anabolic-androgenic steroid stanozolol in adult male rats

    Arch. Toxicol.

    (1999, Nov)
  • P.B. Becker et al.

    In vivo protein-DNA interactions in a glucocorticoid response element require the presence of the hormone

    Nature

    (1986)
  • D.H. Catlin et al.

    Tetrahydrogestrinone: discovery, synthesis, and detection in urine

    Rapid Commun. Mass Spectrom.

    (2004)
  • D.H. Catlin et al.

    Detection of norbolethone, an anabolic steroid never marketed, in athletes’ urine

    Rapid Commun. Mass Spectrom.

    (2002)
  • A.K. Death et al.

    Tetrahydrogestrinone is a potent androgen and progestin

    J. Clin. Endocrinol. Metab.

    (2004, May)
  • R. Jasuja et al.

    Tetrahydrogestrinone is an androgenic steroid that stimulates androgen receptor-mediated, myogenic differentiation in C3H10T1/2 multipotent mesenchymal cells and promotes muscle accretion in orchidectomized male rats

    Endocrinology

    (2005)
  • M. Johansson et al.

    Xenobiotics and the glucocorticoid receptor: additive antagonistic effects on tyrosine aminotransferase activity in rat hepatoma cells

    Basic Clin. Pharmacol. Toxicol.

    (2005)
There are more references available in the full text version of this article.

Cited by (25)

  • Anabolic-Androgenic Steroids

    2013, Disease-a-Month
    Citation Excerpt :

    Tetrahydrogestrinone (THG;13-ethyl-17 hydroxy-18,19-dinor-17α-pregn-4,9,11-trien-3-one), norbolethone (CAS RN: 1235-15-0; 13-ethyl-17 hydroxy-18,19-dinor-17α-pregn-4-en-3-one), and madol (17α-methyl-5α-androst-2-en-17β-ol) are designer anabolic steroids, which were synthesized to avoid detection during use.15,16 However, THG is a nonspecific androgen agonist that binds many steroid hormone receptors, particularly the glucocorticoid receptors.17 Data on the actual prevalence of AAS use are limited by recall and reporting biases associated with illicit drug use.

  • Micronucleus induction in V79 cells by the anabolic doping steroids desoxymethyltestosterone (madol) and 19-norandrostenedione

    2008, Toxicology Letters
    Citation Excerpt :

    It is known to be metabolized by 17β-hydroxy steroid dehydrogenase to 19-nortestosterone (NT, nandrolon), which itself is also frequently used as doping substance (Schrader et al., 2006; Uralets and Gillette, 1999). Unlike other prohormones showing strong anabolic and androgenic potency (Friedel et al., 2006), NA was recently characterized to have more pronounced anabolic activity with relatively weak androgenic activity (Diel et al., 2008). Fig. 1 shows the structural formulae of these compounds.

View all citing articles on Scopus
View full text