Doping Test Results Dependent on Genotype of Uridine Diphospho-Glucuronosyl Transferase 2B17, the Major Enzyme for Testosterone Glucuronidation

Clinical Pharmacology C1:68, Karolinska University Hospital, Huddinge, Stockholm, Sweden.
Journal of Clinical Endocrinology & Metabolism (Impact Factor: 6.21). 08/2008; 93(7):2500-6. DOI: 10.1210/jc.2008-0218
Source: PubMed


Testosterone abuse is conventionally assessed by the urinary testosterone/epitestosterone (T/E) ratio, levels above 4.0 being considered suspicious. The large variation in testosterone glucuronide (TG) excretion and its strong association with a deletion polymorphism in the uridine diphospho-glucuronosyl transferase (UGT) 2B17 gene challenge the accuracy of the T/E ratio test.
Our objective was to investigate whether genotype-based cutoff values will improve the sensitivity and specificity of the test.
This was an open three-armed comparative study.
A total of 55 healthy male volunteers with either two, one, or no allele [insertion/insertion, insertion/deletion, or deletion/deletion (del/del)] of the UGT2B17 gene was included in the study. Intervention: A single im dose of 500 mg testosterone enanthate was administered.
Urinary excretion of TG after dose and the T/E ratio during 15 d were calculated.
The degree and rate of increase in the TG excretion rate were highly dependent on the UGT2B17 genotype with a 20-fold higher average maximum increase in the insertion/insertion group compared with the del/del group. Of the del/del subjects, 40% never reached the T/E ratio of 4.0 on any of the 15 d after the dose. When differentiated cutoff levels for the del/del (1.0) and the other genotypes (6.0) were applied, the sensitivity increased substantially for the del/del group, and false positives in the other genotypes were eliminated.
Consideration of the genetic variation in disposition of androgens will improve the sensitivity and specificity of the testosterone doping test. This is of interest not only for combating androgen doping in sports, but also for detecting and preventing androgen abuse in society.

Download full-text


Available from: Jenny J Schulze
  • Source
    • "We have also shown that the ethnic disparity in the T/E ratio is strongly associated with a deletion polymorphism in the UGT2B17 gene (10). Individuals homozygous for this deletion (del/del) may not reach a T/E ratio of 4 when doped with testosterone (11). The deletion polymorphism is much more common in East Asian populations as compared to Caucasians and Africans (12). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The steroid module of the Athlete Biological Passport, the newest innovation in doping testing, is currently being finalized for implementation. Several factors, other than doping, can affect the longitudinal steroid profile. In this study, we investigated the effect of hormonal contraceptives (HC) as well as the effect of three polymorphisms on female steroid profiles in relation to doping controls. The study population consisted of 79 female elite athletes between the ages of 18 and 45. HC were used by 32% of the subjects. A full urinary steroid profile was obtained using World Anti-Doping Agency accredited methods. In addition all subjects were genotyped for copy number variation of UGT2B17 and SNPs in UGT2B7 and CYP17. Subjects using HC excreted 40% less epitestosterone as compared to non-users (p = 0.005) but showed no difference in testosterone excretion. When removing individuals homozygous for the deletion in UGT2B17, the testosterone to epitestosterone (T/E) ratio was 29% higher in the HC group (p = 0.016). In agreement with previous findings in men, copy number variation of UGT2B17 had significant effect on female urinary testosterone excretion and therefore also the T/E ratio. Subjects homozygous for the T allele of CYP17 showed a lower urinary epitestosterone concentration than the other CYP17 genotypes. It is of great importance that the athlete's steroidal passport can compensate for all possible normal variability in steroid profiles from women. Therefore, considering the large impact of HC on female steroid profiles, we suggest that the use of HC should be a mandatory question on the doping control form.
    Full-text · Article · Apr 2014 · Frontiers in Endocrinology
  • Source
    • "[2] However, due to the recent identification of long-term metabolites of many synthetic anabolic steroids (metandienone, [3] dehydrochlormethyltestosterone) [4] and the resulting long detection windows, a return to endogenous steroids was observed. Any detection of manipulations with endogenous steroids requires a complex discrimination from the state of 'normality', which is potentially affected by age, gender, inter-individual and diurnal variations, genotype (UGT2B17 [5] and ethnicity) as well as nutrition, medication (5α-reductase inhibitors, e.g. finasteride) [6] or alcoholic beverages (ethanol). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The legally defensible proof of the abuse of endogenous steroids in sports is currently based on carbon isotope ratio mass spectrometry (IRMS), i.e. a comparison between (13) C/(12) C ratios of diagnostic precursors and metabolites of testosterone. The application of this technique requires a chromatographic baseline separation of respective steroids prior to IRMS detection and hence laborious sample pre-processing of the urinary steroid extracts including clean up by solid-phase extraction and/or liquid chromatography. Consequently, an efficient pre-selection of suspicious control urine samples is essential for appropriate follow up confirmation by IRMS and effective doping control. Two single transdermal administration studies of testosterone (50 mg Testogel® and Testopatch® at 3.8 mg in 16 h, respectively) were conducted and resulting profiles of salivary testosterone and urinary steroid profiles and corresponding carbon isotope ratios were determined. Conventional doping control markers (testosterone/epitestosterone ratio, threshold concentrations of androsterone, etiocholanolone, or androstanediols) did not approach or exceed critical thresholds. In contrast to these moderate variations, the testosterone concentration in oral fluid increased from basal values (30-142 pg/mg) to peak concentrations above 1000 pg/mg. It is likely that this significant increase in oral fluid is due to a pulsatile elevation of free (protein unbound) circulating testosterone after transdermal administration and may be assumed to represent a more diagnostic marker for transdermal testosterone administration. Copyright © 2013 John Wiley & Sons, Ltd.
    Full-text · Article · Nov 2013 · Drug Testing and Analysis
  • Source
    • "Since the comparison is almost always made with the reference population and not yet based on the individual longitudinal profile, a great number of athletes' samples requiring further GC/C/IRMS analysis are those with a T/E value greater than 4, normally found in approximately 2 % of the male athletes' samples received for routine testing in our laboratory. [6] [36] When considering the wide inter-individual variation reported following the administration of testosterone itself with frequent observations of T/E values not exceeding 4, this populationbased initial test is generally considered to be inefficient. [5,6,13,18,26,36–39] From our experience, the administration of testosterone, androstenedione or DHEA could be detected in samples presenting relatively minor alterations of the steroid profile (T/E values between 1 and 4). "
    [Show abstract] [Hide abstract]
    ABSTRACT: The analysis of urinary metabolites of testosterone-related steroids through the measurement of their carbon isotopic signature (δ(13) C) by gas chromatography/combustion/mass spectrometry (GC/C/IRMS) is a confirmation method employed in doping control analyses. Stringent analytical conditions are essential to an accurate and precise analysis as well as the proper selection of the metabolites, which forms the basis of the refined method presented in this paper. In a simplified approach, following enzymatic hydrolysis and extraction from a relatively low volume of urine sample, a one-step high-performance liquid chromatography (HPLC) purification was developed for seven diagnostic urinary metabolites (TS) including testosterone itself, dehydroepiandrosterone, 5α- and 5β-androstanediol, epitestosterone, androsterone, etiocholanolone and two endogenous reference compounds (ERC), 5β-pregnanediol and 5α-androst-16-en-3β-ol. These steroids were pooled in three fractions and analyzed as such. With regards to the GC/C/IRMS analysis, a multi-level isotopic calibration using the 'identical treatment' principle was created. The proposed isotopic calibration yielded results for purified reference steroids with a precision ≤0.15 and accuracy of ≤0.30 ‰ (between-assay, n = 26). Compared to other common endogenous reference compounds, those selected in this study had δ(13) C values close to the target metabolites which, along with the proposed isotopic calibration, produced narrow reference intervals within ± 3‰ for most diagnostic TS-ERC pairs, in compliance with the requirements of the World Anti-Doping Agency. These carefully controlled analytical conditions are compatible with routine operations, affording accurate and precise results for the more diagnostically relevant metabolites such as testosterone itself and the 5α- and 5β-androstanediols. The values of the TS-ERC pairs measured in reference populations are described and the results from the routine testing of several hundreds of athletes' samples are discussed. Robust, this technique permitted the detection of adverse findings that would have been missed had these low level metabolites not been analyzed. Copyright © 2013 John Wiley & Sons, Ltd.
    Full-text · Article · Aug 2013 · Rapid Communications in Mass Spectrometry
Show more