"The purposes of the World Anti-Doping Code and the World Anti-Doping Program which supports it are: i) to protect the Athletes' fundamental right to participate in doping-free sport and thus promote health, fairness and equality for Athletes worldwide, and ii) to ensure harmonized, coordinated and effective anti-doping programs at the international and national level with regard to detection, deterrence and prevention of doping" [1
]. Current doping control is primarily based on enforcement, complemented by deterrence strategies. Meticulously regulated testing protocols, including regulated analytical testing in accredited laboratories and international standards are in place to identify those who violate the anti-doping rules worldwide and to deter others from doing so [2
]. The use of performance-enhancing substances and methods deemed to be prohibited is an anti-doping rule violation that is established by the presence of a prohibited substance or its metabolites or markers in the athlete's sample, typically urine or blood [1
Aside from an ongoing ethical debate on various facets of doping [4
], including moral reasoning [5
] and medical ethics [6
], recent critical analyses of the current anti-doping approach have claimed that the current anti-doping is not fit for purpose [10
]. Reasons for this criticism encompassed ethical issues around constant surveillance [14
], analytical difficulties and costs [16
], as well as marked inter-individual differences [18
Anabolic androgenic steroids (AASs) are synthetic derivatives of the endogenously produced male sex hormone, testosterone, which exhibits both anabolic (protein synthesizing) and androgenic (masculinising) effects. Steroids are one of the most potent and the most widely used performance-enhancing substances both amongst Olympic athletes [21
] and also those outside of the auspices of the World Anti-Doping Agency (WADA), such as competitive and recreational body builders, professional players or even non-athlete adolescent boys [24
]. The use of AASs is widespread particularly amongst athletes because such drugs can improve their performance in sports by accelerating muscle growth, increasing aggressiveness and enhancing a sense of well-being. Chronic use of AASs has been known to cause serious adverse effects such as virilization, feminization, liver disorders, neuropsychiatric disorders, adverse blood lipid profiles (increased LDL and decreased HDL), cardiovascular disorders and renal complications [26
]. Among these, renal diseases have received less attention, most likely because renal disorders are infrequent among AAS users in comparison to other, more prevalent diseases. Throughout the last decade, the literature sporadically presented cases of severe renal disorders among AASs users, especially with elevated and prolonged use [29
]. The number of incidents presented is well below the estimated number of AAS users, however, the user profiles described in these case studies do not differ significantly from those AASs abusers who do not develop renal complications. Thus, there may be a connection between deletion mutation in a steroid conjugating enzyme and occurrence of renal diseases with chronic use of AAS.
AASs are commonly excreted in urine mainly as glucuronide conjugates, the formation of which is catalyzed by various uridine diphosphate-glucuronosyltransferase (UGT) enzymes. The UGTs 2B7, 2B15 and 2B17 are found to be the principal enzymes involved in glucuronidation of androgens and their metabolites in humans. Glucuronidation of steroids and their phase I metabolites is an important detoxification and deactivation metabolic pathway which is catalyzed mainly by UGT2B17
and to a minor extent by UGT2B15
]. In 2008, a few months before the Beijing Olympics, Schultze and colleagues discovered that the occurrence of deletion polymorphism in the gene coding of UGT2B17
enzyme affected the urinary excretion patterns of testosterone [18
]. As the current doping testing regime relies on detecting the steroid metabolites in urine, UGT2B17
deficient drug abusers are likely to test negative despite the use of the drug. Although the enzyme deficient athletes who are aware of their own genetic profile may benefit from evading doping testing, they may be vulnerable to serious health consequences due to inadequate deactivation and elimination of steroids. The potential consequences of this discovery has triggered the WADA and sport governing bodies to examine their testing regimes.
However, it is inevitable that this phenomenon has significant implications beyond the sporting arena. Juul et al. [39
] observed in pubertal boys, that the homozygous deletion in the UGT2B17
gene, in line with Schulze et al. [18
], affected the urinary excretion pattern of androgen metabolites, but not circulating androgen levels. A deficiency of the UGT2B17
enzyme decreases the rate of steroid deactivation and elimination, and owing to this, the bioavailability of steroids may be enhanced to a certain extent. Hence, elevated and prolonged use of AAS may predispose the enzyme deficient individuals to detrimental effects, particularly relating to kidney damage.
Therefore, in this paper we hypothesise that the observed renal disorders among AAS users is connected to the genetic profiles of these users and functional polymorphic deletion of the UGT2B17 gene significantly increases the chance of developing kidney complications.
Inter-individual variations in the frequency of UGT2B17 gene deletion polymorphism
The occurrence of the UGT2B17
gene varies amongst individuals of various ethnic groups. Wilson et al. carried out a study on African Americans and the Caucasian population, and found that the occurrence of deletion mutation in the UGT2B17
gene was five time more frequent in Caucasians than in African Americans [40
Several comparative studies of urinary steroid concentrations amongst various ethnic groups have been reported and it has been observed that individuals lacking this enzyme have negligible excretion of steroids. Sjöqvist et al. examined the association of androgen excretion with UGT2B17
deletion in a population based study comprising of Korean and Swedish participants [22
]. It was found that the absence of the UGT2B17
gene was seven times more frequent in Koreans than in the Swedish population. On examining the association between deletion polymorphism and urinary levels of androgens it was revealed that testosterone excretion was 16 times higher in Swedish people compared to the Koreans. These findings indicate the importance of this gene in the excretion of steroids.
Baume et al. reported inter-individual variations in the excretion patterns and kinetics of nandrolone and its metabolites after administration of [13
C] nandrolone to volunteers and noted the possible natural production of nandrolone and its metabolites [20
]. However, it can be postulated that the variations in nandrolone excretion could be due to variations in UGT2B17
genotypes amongst individuals.
According to WADA guidelines, if the ratio of concentrations of testosterone to epitestosterone glucuronide in urine is greater than 4, then drug doping is suspected [41
]. Since deletion polymorphism in the gene coding for the UGT2B17
enzyme affects the urinary level of testosterone, the accuracy of the T/E ratio test is challenged. A study carried out on a heterogeneous group of healthy volunteers with different UGT2B17
genotypes (ins/ins, ins/del
) reported that administration of exogenous testosterone to individuals lacking the UGT2B17
gene did not yield T/E values above the population based threshold of 4 for all individuals [18
]. This is because the testosterone glucuronide excretion rate in individuals with del/del
genotype was found to be significantly less than those carrying the ins/del
genotype. No significant effect on epitestosterone excretion was observed. The excretion of unconjugated steroid was found to be a minor elimination pathway even in individuals devoid of the gene. This indicates the possibility of an increase in serum levels of biologically active steroids. It has been reported previously that individuals of Asian ethnicity excrete less testosterone which complements the surveillance that the del/del
genotype is more frequent amongst the Asian population [18
]. Many researchers have observed similar difficulty in testing testosterone abuse pertaining to the UGT2B17
deletion, and have suggested making use of genotype based cut-off levels to overcome inter-individual and inter-ethnic variations [42
These results indicate that a deletion polymorphism in the gene coding is associated with urinary levels of steroid glucuronide conjugates. Owing to an impaired glucuronidation pathway, deactivation and elimination of steroids is reduced. This may result in elevated serum levels of active steroids which can be harmful over a long period of time. Thus, specific groups of individuals who are devoid of the UGT2B17 gene are more susceptible to adverse health conditions due to chronic exposure to elevated doses of steroids, compared to individuals having the gene.
Concomitant use of drugs
UGTs not only contribute to AAS glucuronidation but they also act as conjugating enzymes for various other pharmaceutical drugs [35
]. It has been reported that athletes use AASs in combination with other medications which may enhance the AAS effects and decrease the side effects associated with such performance-enhancing drugs. Co-administration of steroids along with other UGT substrates may lead to competitive inhibition of steroid glucuronidation. Sten et al. discovered that commonly used over-the-counter (OTC) non steroidal anti-inflammatory drugs (NSAIDs) like diclofenac and ibuprofen inhibited the testosterone glucuronidation activity of UGT2B17
and some other UGTs that have previously shown low but detectable activity [43
]. Compared to UGT2B17
was found to be more sensitive to both the NSAIDs, particularly ibuprofen. Since, UGT2B17
shares 96% homology with UGT2B15
, in a UGT2B17
deficient individual, UGT2B15
may be the major contributor to glucuronidation. However, use of steroids together with NSAIDs will competitively inhibit the steroid glucuronidation activity of UGT2B15
. The inhibitory effect was also found to be dependent on the UGT2B17
genotype. Regular use of such painkillers is common amongst athletes in order to overcome the pain associated with extensive exercise regimes [44
]. Thus, athletes devoid of the UGT2B17
gene are susceptible to having complications in steroid elimination, as well as to other medications administered simultaneously. Also, such drug combinations can have direct deleterious effects on renal health.
The occurrence of renal disorders among AAS users
Herlitz and colleagues reported a study on a cohort of bodybuilders (white and hispanic) and observed that long term use of anabolic steroids combined with high protein intake was associated with proteinuria and secondary focal segmental glomerulosclerosis (FSGS). The pattern of glomerular injury observed, involves scarring of the glomerules and is mediated by elevated glomerular filtration rate (hyperfiltration), glomerular pressure and other adaptive structural-functional responses within the kidneys [29
]. One of the bodybuilders with a history of occasional urinary tract infection progressed to end stage renal disease (ESRD) after prolonged use of AASs. It has been reported that non obese individuals with increased body mass index (BMI) owing to elevated muscle mass are susceptible to developing secondary FSGS [29
]. This indicates the possibility of developing serious renal conditions after prolonged use of steroids by the gene deficient individuals. Many researchers have also reported other adverse renal conditions such as acute kidney injury (AKI), nephropathy, diffuse type 1 membranoproliferative glomerulonephritis and also renal failure owing to long term use of AASs [30
]. In all these case studies, improvements in renal conditions have been observed in patients who discontinued the use of anabolic steroids. Also, acute renal failure has been reported along with cholestatic liver damage after long term use of AASs [32
]. Disorders in the liver, the major site for detoxification of steroids and other xenobiotics, can hinder its performance and hence worsen the renal conditions due to elevated level of circulating toxins.