Androgen abuse remains the most potent and prevalent form of sports doping. It is particularly sought in power sports where the potential of androgens to increase muscle mass and strength may improve performance.1 Although antidoping initiatives have become increasingly stringent over the past 4 decades, the temptation to seek an unfair advantage by drug cheating still prevails. In nearly 270 000 laboratory tests performed in 2013 by World Anti-Doping Agency (WADA)-approved antidoping laboratories, hormones accounted for 75% of all adverse analytical findings with androgens accounting for 84% of hormone (or 63% of all) positives. Originally, androgen abuse involved the surreptitious use of marketed synthetic androgens, exemplified by the secret national sports doping program of the former German Democratic Republic2 and by individual athletes and their coaches. Because these marketed synthetic androgens become easily detected by their characteristic nonnatural chemical signatures in mass spectrometry (MS)-based urine detection tests, the pursuit of illicit ergogenic benefits from androgen doping has turned to other means, including administration of “designer” or “nutraceutical” synthetic androgens, to indirect androgen doping methods, or use of exogenous natural androgens. The use of “designer” or “nutraceutical” androgens3–5 draws on the legacy of largely forgotten patents of the 1950s-1970's, the “Golden Age” of steroid pharmacology, when thousands of potent synthetic androgens were developed but rarely underwent clinically testing or were marketed. Such synthetic androgens can now be readily synthesized by modern technology available affordably from chemical manufacturers and can be detected by yeast or mammalian cell-based in vitro androgen bioassays6–8 until their chemical structure is identified as required for MS-based detection. Indirect androgen doping methods consists of using nonsteroidal drugs such as human chorionic gonadotropin (hCG),9,10 gonadotropin-releasing hormone analogs,11 and antiestrogens,12 which aim to increase endogenous testosterone secretion. Finally, administration of exogenous natural androgens and their precursors [testosterone, dihydrotestosterone (DHT), androstenedione, dehydroepiandrosterone (DHEA)] have to be distinguished from their endogenous counterparts through analysis by isotope ratio MS.13–15
WADA code mandates strict athlete liability in prohibiting drug use, attempted use, possession, trade, and sample tampering.16 Furthermore, the revised 2015 Code extends sanctions to any member of the “athlete entourage” complicit in drug misuse. A positive test for a prohibited substance or method constitutes an antidoping rule violation, regardless of intent, fault, negligence, or knowing use. However, an important caveat is the provision for athletes who suffer coincidentally from any proven medical condition for which a prohibited substance is required. Where this involves pathological hormone deficiency, effective safe treatment must demonstrate that replacement therapy at standard doses confers no ergogenic advantage to the athlete. Such circumstances are embodied in the WADA provision for Therapeutic Use Exemption (TUE), whereby certification authorizes time-limited use of a prohibited substance based on legitimate medical need and validated by independent expert scrutiny. The requirements necessary to satisfy a TUE are that (1) withholding treatment would cause significant harm to health, (2) that the use of the prohibited substance will confer no ergogenic benefit beyond the return to normal health, (3) that no nonbanned alternative treatment exists, and (4) that the therapeutic need is not due to the previous use of any prohibited substance without a TUE.17
A TUE is justified for testosterone replacement therapy for male athletes suffering from pathological disorders of the hypothalamo-pituitary testicular axis such as congenital anorchia, bilateral orchidectomy, idiopathic hypogonadotropic hypogonadism (including Kallmann syndrome), hypopituitarism because of pituitary tumor and Klinefelter syndrome.18 Such authorization is not considered appropriate for functional disorders including age-related low serum testosterone related to age (“andropause,” “LowT,” “late-onset hypogonadism”), suppression of endogenous testosterone by previous androgen abuse, or where serum testosterone is low secondary to systemic nonreproductive disorders and their treatment (obesity, chronic systemic illness, diabetes, HIV) or major stressful states including overtraining.19
Another potential reason for TUE application for (pro) androgen therapy is DHEA treatment of confirmed primary adrenal insufficiency in females. However, given the status of DHEA as a WADA-prohibited androgen precursor, any application for its therapeutic use would require a robust, validated argument. This review considers the evidence required for issuing such a TUE.
Circulating DHEA and its sulfated form DHEA sulfate (DHEAS) are primarily synthesized and secreted from the adrenals of higher primates. The major known roles of both are in pregnancy and during adrenarche. In pregnancy, the fetal adrenal cortex secretes large amounts of DHEAS that undergoes desulfation by placental sulfatases during passage into the mother's circulation with conversion to potent sex steroid, as well as exerting other local tissue effects.20,21 In childhood, DHEA(S) is believed to be important in initiating adrenarche.22,23 The interrelationship between DHEA and DHEAS is complex but the more hydrophilic sulfated form does not cross lipophilic membranes or bind to steroid receptors and, on infusion in humans, is not converted into DHEA in the circulation.24 This suggests that, in adults, DHEAS may primarily be a biologically inactive reservoir and/or excretory form of DHEA which becomes bioactive only after excision of the sulfate moiety by steroid sulfatase (arylsulfatase C).25,26 Steroid sulfatase is an X-linked, membrane-bound microsomal enzyme which is strongly expressed in placenta and, to a much lower extent, in other reproductive and nonreproductive (skin, kidney, bone, brain) tissues25,26 with the tissue-specific variations determined by alternate splicing.27
DHEA lacks any known receptor, so is not presently considered a hormone.28 Rather, it is a proandrogen or androgen precursor that requires steroidogenic enzyme metabolism for conversion to a potent androgen (testosterone or DHT) or estrogen (estradiol) necessary to exert any hormonal bioactivity by activating their cognate androgen or estrogen receptors. Nevertheless, there remain members of the nuclear receptor superfamily, the so-called orphan receptors,29 which still lack an identified ligand. Orphan receptors continue to have their unexpected ligands identified, including small molecules such as retinoic acid, bile acids, heme, fatty acids, oxysterols, phospholipids, and xenobiotics.30 DHEA is reported to have modulating effects on neurotransmitter action31 and ion channels,32 especially in the brain. These effects may depend on the synthesis of DHEA within the brain to act as a local neurosteroid33 rather than systemic effects of adrenal DHEA, so has no clear clinical significance for antidoping at present. Therefore, although it is premature to conclude definitively that DHEA has no hormone receptor and is therefore not a hormone, for antidoping purposes its conversion to potent androgens is its most relevant physiological feature.
The adrenal produces three classes of steroid, each from anatomically and functionally distinct regions of the adrenal cortex and subject to independent physiological regulation21,34 (Figure). Mineralocorticoids, notably aldosterone, are secreted from the zona glomerulosa, the small subcortical region, comprising ∼5% of the adrenal cortex and distributed in discontinuous nests of cells. The zona glomerulosa is characterized functionally by expression of the angiotensin II receptor and the 18-hydroxylase enzyme (P450c11AS, CYP11B2), which allows for regulation of aldosterone secretion through the renin/angiotensin system.35 Glucocorticoids, primarily cortisol in humans, are secreted from zona fascicuata, the mid-region comprising ∼70% of the adrenal cortex organized as columns of cells which characteristically expresses 17-hydroxylase/lyase (P450c17, CYP17A1) and the adrenocorticotropic hormone (ACTH) (melanocortin type 2) receptor and which allows for cortisol secretion under regulation through negative feedback regulating hypothalamus corticotropin-releasing hormone (CRH) and pituitary ACTH secretion. Finally, DHEA and other adrenal androgen precursors are secreted from the innermost region, the zona reticularis, comprising ∼25% of the adrenal cortex and characterized functionally by expression of the ACTH (melanocortin type 2) receptor, of cytochrome b5 (CYB5), which accentuates the 17,20 lyase activity of 17-hydroxylase/lyase (P450c17, CYP17A1) and by the relatively low expression of type 2 3β-hydroxysteroid dehydrogenase together with high expression of DHEA sulfotransferase which are, in concert, responsible for predominant production of DHEA and DHEAS by this region of the adrenal cortex. DHEA secretion is mainly responsive to ACTH but regulation by additional pituitary-dependent factors has long been postulated but still remains unproven.
Adrenal failure may be complete, typically in primary adrenal disorders such as bilateral adrenalectomy, adrenal hemorrhage or autoimmune Addison's disease or partial, typically caused by secondary or central (hypothalamic-pituitary) disorders or use of exogenous glucocorticoid therapy that suppresses the ACTH-glucocorticoid axis alone. Although any clinically significant adrenal failure requires lifelong glucocorticoid replacement therapy, typically using cortisone acetate, hydrocortisone (cortisol), prednisone or dexamethasone, the distinction between complete and partial (or alternatively primary vs secondary) adrenal failure has important clinical significance.
Complete adrenal failure, involving anatomical loss or severe structural damage to all 3 adrenal cortical zones, carries a risk of potentially fatal mineralocorticoid deficiency because of extracellular fluid volume loss and circulatory collapse, which usually necessitates mineralocorticoid replacement therapy. The latter is typically achieved by treatment with the mineralocorticoid, fludrocortisone, if the modest mineralocorticoid activity of cortisone acetate is not sufficient or using pure glucocorticoids like prednisone or dexamethasone that have negligible mineralocorticoid activity.
In partial or secondary (central) adrenal failure, the zone fasciculata is structurally preserved but functionally suppressed by reduced ACTH secretion.36,37 Secondary adrenal failure is mostly frequently due to prolonged use of exogenous glucocorticoids that suppress endogenous hypothalamic CRH and pituitary ACTH secretion. Less frequently, partial adrenal insufficiency may be due to structural defects of hypothalamus or pituitary (eg, tumors and their treatment) that reduces ACTH secretion but this is nearly always in conjunction with other pituitary-dependent hormonal axes (gonadal and thyroidal). Secondary adrenal failure involving a combination of adrenal and ovarian insufficiency may have more pronounced general symptomatic effects than isolated secondary adrenal failure because of exogenous glucocorticoid use. In contrast to primary adrenal failure, partial or secondary adrenal failure usually does not usually require mineralocorticoid replacement therapy.36 This reflects the fact that the adrenocortical zones that secrete mineralocorticoids (aldosterone) and adrenal androgen precursors (DHEA) remain structurally intact and independently regulated. By contrast, in primary adrenal failure, significant structural damage severely reduces or abolishes steroidal output of all 3 adrenocortical zones. Hence structural defects, which may abolish DHEA secretion can be inferred from the status of the mineralocorticoid axis, corresponding with the clinical experience that neither mineralocorticoid nor DHEA replacement are required in secondary adrenal failure. Nevertheless, reductions in serum DHEA may be difficult to interpret given that prolonged or supraphysiological treatment with exogenous glucocorticoids reduces pituitary ACTH secretion and ACTH-dependent adrenal DHEA secretion. Thus, blood DHEA(S) concentrations may be reduced irrespective of the clinical purpose of glucocorticoid use. In summary, there may be an a priori rationale for DHEA replacement therapy in primary adrenal failure, but little physiological justification in partial or secondary adrenal failure, especially that due to exogenous glucocorticoid use.
In the USA, DHEA has had a tortuous regulatory status. Originally considered as a food (dietary supplement) available for over-the-counter (OTC) sales, in 1985 the Food and Drug Administration prohibited OTC until 1994 when the passage of the Dietary Supplement Health and Education Act again interpreted DHEA as a food rather than a drug. In 2004, the Anabolic Steroid Control Act re-regulated many steroidal “dietary supplements” but excluded DHEA, which remains regarded as a food rather than a drug subject to FDA regulation. In other countries, DHEA is often formally regulated as a drug requiring a prescription (Canada, Australia, UK, EU), but remains relatively freely available at health food shops or through internet. In addition, dietary supplements are known to contain DHEA without declaration on the label.38
Well-controlled clinical studies establish that DHEA administration to women significantly increases serum T compared with placebo39–48 when serum T is measured by reliable methods such as immunoassays using preassay extraction40,43 or MS.48 Similar findings were reported in most39,41,42,44–47 but not every49 study using direct (nonextracted) immunoassays that are less accurate and insensitive at the low serum T concentrations prevailing in females.50,51 In men, exogenous DHEA administration increases blood estradiol in some52 but not all53 studies; these discrepancies being probably due to the unreliability of direct estradiol immunoassays at the low circulating estradiol concentrations in male serum.54 Administration of DHEA to men does not change serum T or DHT41,44,46,47,52,53 when measured by direct T immunoassays.50,51 These findings presumably reflect the much higher (>20-fold) endogenous blood T concentration in men compared with women so that additional conversion of DHEA to T and/or DHT is quantitatively minimal and difficult to detect in men, unlike women. Therefore, for antidoping purposes, the potential ergogenic effects of DHEA administration are of primary concern in women where this amounts to an indirect form of androgen doping. Although the increase in serum T after administration of DHEA in women may not reach male serum T concentrations, it is nevertheless, a definite effect to increase a potent ergogenic agent. Although it may be argued that prohibition of DHEA treatment may not be strictly necessary in men, DHEA remains prohibited for both sexes.55,56
In women with complete adrenal failure, daily DHEA treatment with 25–50 mg achieves restoration of blood DHEA concentrations to levels comparable with age-matched controls with normal adrenal function.39,57,58 In several studies, however, the 50 mg DHEA dose in women with adrenal failure causes androgenic side effects consistent with an excessive dose.41,42,47,59,60 Such variability in androgenic response to DHEA dose may arise because OTC DHEA products, categorized as foods or “nutritional supplements” rather than pharmaceutical products, require no proof of bioequivalence and may differ substantially in oral bioavailability. Hence, even with nominally standard doses of DHEA sourced as OTC or pharmaceutical products, there is a risk of excessive DHEA dose leading to supraphysiological serum DHEA, as well as T and DHT concentrations and consequent potential ergogenic effects. Hence, any permitted DHEA use for female athletes, would necessitate individual titration of optimal DHEA replacement doses so as to maintain steady state, physiological blood T and DHT concentrations.
There remains an objection that the available clinical studies are underpowered, neither long nor large enough, to fully exclude subtle or small benefits from DHEA replacement in adrenal failure.61 On that basis, an argument could be envisaged for the use of physiological replacement doses of DHEA accompanied by monitoring of blood T. This may be feasible for the supervised use of DHEA in the rare elite female athlete with proven primary adrenal failure seeking a TUE. Under this scenario, approval could be justified with a recommended commencing oral DHEA dose of 25 mg daily monitored by regular unannounced serum DHEA (or DHEAS) and T concentrations. This monitoring for serum T must use validated MS-based methods such as those available in WADA-accredited laboratories where these are used in conjunction with established MS-based reference ranges for healthy premenopausal women.62 The widely available direct (unextracted) T immunoassays used by most clinical/chemical pathology laboratories are inadequate for monitoring such physiological DHEA usage. The method-specific bias, insensitivity, and inaccuracy of direct T immunoassays for female serum would create random false negatives and positives.50,63,64 Serum T assays based on validated MS or immunoassays using preassay extraction are available from US commercial pathology laboratories. It is noted that the so-called “free” T (free analog) assays, which purports to measure the free T is notoriously unsatisfactory50,65–68 with results corresponding to neither serum T nor “free” T as measured by reference dialysis methods and is not suitable for use in this context.
DEHYDROEPIANDROSTERONE TREATMENT FOR ADRENAL INSUFFICIENCY
The role of DHEA replacement therapy for primary or secondary adrenal failure remains controversial and not well established.69,70 The rationale for DHEA use in females with adrenal failure is based on evidence from well-designed, randomized, placebo-controlled studies of quality of life (QoL) in women with adrenal failure. For such trials, the requirement for placebo controls and double-blinding are indispensable because the endpoints are subjective or potentially influenced by secondary gain. The most authoritative review is the 2006 US Endocrine Society Clinical Practice Guideline,70 updated in 2014,69 which recommends against making the diagnosis of androgen deficiency in women, specifically in those with primary or secondary (hypopituitary) adrenal failure. Meta-analyses of placebo-controlled clinical trials of DHEA in women with71 or without72 adrenal insufficiency with a focus on QoL endpoints reached similar conclusions.71 These conclusions are also consistent with WADA's TUE guidance document on adrenal insufficiency.73
The most salient rationale for justifying DHEA replacement is for primary adrenal failure because (1) DHEA may be only partly deficient in secondary adrenal failure especially after or during exogenous glucocorticoid use and (2) for women with hypopituitarism, the clinical features, notably quality of life, may be influenced by suboptimal hormone replacement for one or more other pituitary-dependent hormones such as estradiol, cortisol, thyroid hormones, and growth hormone.
Despite evidence from some early and prominent randomized, placebo-controlled clinical trials showing DHEA treatment improves some QoL features (well-being, mood, libido, sexual function) in women with primary adrenal failure,39,59 many subsequent studies produced no, small, or deleterious QoL and/or biochemical effects.43,44,47,49,60,74–83 Overall, most well-controlled clinical studies published (see Table, updated from Alkatib et al71) show little or no clinical quality of life benefit of DHEA treatment in women with adrenal failure although they all involve small numbers of women.
DEHYDROEPIANDROSTERONE TREATMENT AND WADA-PROHIBITED LIST
The pinnacle of evidence for drug effects in humans is data from well-controlled, prospective clinical trials featuring randomization, placebo controls and specific clinical endpoints. The definitive appraisal of the potential ergogenic effects of any substance in the sporting context remains an unsolved dilemma. Evidential certainty from definitive research would require the use of banned drugs by elite athletes in competition, and this is clearly fraught with moral and ethical arguments.1 Hence for antidoping science, it becomes necessary to draw the most plausible inference from the best available surrogate evidence on the ergogenic effects of DHEA.
Classical studies using suitable surrogate variables provide compelling evidence for ergogenesis linked with a number of banned hormonal drugs. For example, Bhasin et al84,85 provided a strong biological basis for androgen doping effects in power sports from a series of studies demonstrating a tight linear relationship between testosterone dose and muscle mass or strength. These androgenic effects extended from below to well above the physiological testosterone levels without any plateau even at 6 times the regular replacement dosage for testosterone. The data also displayed additive effects with physical exercise that were unaffected by age. Similarly, the strong linear relationship between acute changes in circulating hemoglobin and maximal oxygen consumption86 explains the effectiveness of hemoglobin doping whereby the circulating hemoglobin level is increased by blood transfusion or indirectly by erythropoiesis-stimulating agents. This also applies to indirect androgen or hemoglobin doping which employs nonhormonal drugs to increase endogenous hormones (T, erythropoietin) with consequential performance-enhancing properties. For example, the use of hCG, luteinizing hormone, gonadotropin-releasing hormone analogs, or antiestrogens, all of which are banned on the basis that androgens are banned.87 Similarly, indirect hemoglobin doping methods include erythropoietin and its analogs, hypoxic-mimetics and artificial oxygen carriers are banned on the basis that they are likely to increase circulating hemoglobin.88 It is therefore neither necessary nor feasible to evaluate explicit ergogenic effects for each of the growing list of putative doping substances. It is sufficient to show that for any substance in question, the key surrogate variable that may induce performance enhancement is increased (endogenous testosterone for indirect androgen doping; hemoglobin for indirect blood doping). It is also germane to consider that the doses and combinations of drugs accepted for safe use in demonstrative, ethically approved clinical trials are far lower than the doses used illicitly by athletes for doping purposes where exotic, massively supraphysiological dosing regimens (“stacking,” “pyramiding,” “cycling”) are commonplace.89,90
ANALYSIS OF DEHYDROEPIANDROSTERONE FOR ADRENAL FAILURE IN TERMS OF TUE CRITERIA
Based on the above considerations, there is equivocal evidence that DHEA administration may meet the formal requirement necessary to satisfy a TUE for female athletes with primary adrenal failure. However, the criteria are not met for secondary adrenal failure. The specific criteria, paraphrased from the WADA International Standard for TUEs,17 are addressed below.
(a) …withholding treatment would cause significant impairment to health.
As the available evidence does not support that DHEA objectively improves health or subjective quality of life, there is no sufficient, reliable evidence that withholding DHEA treatment would cause significant harm to health.
(b) …that the use of the prohibited substance is highly unlikely to confer ergogenic benefit beyond the return to normal health.
As the administration of DHEA to women, with or without adrenal failure, will increase circulating T, it is plausible, or at least cannot be excluded, that this may confer an ergogenic benefit based on an increase in the circulating concentrations of a banned, potent ergogenic androgen on the background of the low serum T concentrations in normal women.
(c) …that no nonbanned alternative treatment exists.
There is no nonbanned alternative treatment. The only potential alternative treatment suggested for women with adrenal failure is T91 that is also banned.
(d) …that the therapeutic need is not due to the previous use (without a TUE) of any prohibited substance.
The use of supraphysiological doses of any synthetic glucocorticoids for antiinflammatory, immunosuppressive, or other indications will suppress serum DHEA(S) through negative feedback mechanisms regardless of underlying adrenal function. This criterion means that suppression of serum DHEA(S), with or without accompanying secondary adrenal failure, due to previous use of synthetic glucocorticoids without a TUE would not justify a TUE. Such circumstances might also be seen as “gaming” to justify exogenous DHEA use. This criterion is not relevant to primary adrenal failure.
There are significant policy concerns regarding permitting DHEA treatment for secondary adrenal insufficiency. Such precedent to permit the use of DHEA in women with secondary adrenal insufficiency would create the situation where athletes would be free, and in practice encouraged, to use exogenous systemic glucocorticoids for doping. An important related concern is that regulating of permitted DHEA dose is difficult, and there would be temptation to increase the dose and circulating serum T. A simplistic biochemical diagnosis based solely on suppressed serum DHEA and cortisol could encourage “gaming” to justify a TUE for DHEA.
Recent case law supports these interpretations as the Court for Arbitration in Sports (CAS), the ultimate legal authority in antidoping matters under the WADA code, upheld WADA's rejection of a TUE for DHEA use in athlete who had secondary adrenal insufficiency because of exogenous glucocorticoid use.
We conclude therefore that there is no convincing clinical evidence to support the use of DHEA replacement therapy in women with secondary adrenal failure and that the evidence to support its use in primary adrenal failure is scant and equivocal. Therefore, a TUE for DHEA is not justified for either secondary or primary adrenal failure without more definitive evidence of health benefit. For the management of primary adrenal failure, further randomized, placebo-controlled studies are needed using DHEA dosages that produce physiological levels of serum DHEA(S), T and DHT concentrations by reliable MS-based assays. These conclusions are consistent with the 2014 update69 and the original 200670 US Endocrine Society guidelines, meta-analyses of DHEA treatment in women with71 or without72 adrenal failure, the current WADA TUE guidance document for adrenal insufficiency73 and supported by CAS case law.
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