Doping Prevalence among U.S. Elite Athletes Subject to Drug Testing under the World Anti-Doping Code.

BACKGROUND: Determining the prevalence of doping within an elite athlete population is challenging due to the extreme sensitivity of the topic; however, understanding true doping prevalence is important when designing anti-doping programs and measuring their effectiveness. The objective of this study was to estimate the prevalence of doping among Olympic, Paralympic, World, and National-level competitive athletes in the United States subject to the World Anti-Doping Code. All athletes who were subject to the U.S. Anti-Doping Agency's Protocol for Olympic and Paralympic Movement Testing, a World Anti-Doping Code ("Code")-compliant anti-doping program, were invited to complete a web-delivered survey. Using a direct questioning approach, the survey items asked athletes whether they had used each specific category of banned substance / method on the World Anti-Doping Agency's Prohibited List. Multiple strategies to encourage honest reporting (e.g., protecting anonymity by collecting minimal demographic information; using an outside organization to administer the survey) and to detect inconsistent responses were used. RESULTS: Depending on the method of calculation, 6.5-9.2% of the 1,398 respondents reported using one or more prohibited substances or methods in the 12 months prior to survey administration. Specific doping prevalence rates for each individual substance / method categories ranged from 0.1% (for both diuretics / masking agents and stem cell / gene editing) to 4.2% for in-competition use of cannabinoids. CONCLUSION: Determining the prevalence of doping within different athlete populations is critical so that sport governing bodies can evaluate their anti-doping efforts and better tailor their programming. By measuring doping prevalence of specific categories of substances and methods, rather than just the overall prevalence of doping, this study also highlights where sport governing bodies should focus their future educational and detection efforts.

Application of micro capillary blood sampling in an anti-doping setting.

Blood collection is an important facet of anti-doping testing, forming the basis of the Athlete Biological Passport (ABP). Traditional blood collection via venipuncture can be uncomfortable for athletes, especially those who are tested frequently or close to competition. Athletes may also have negative perceptions of venipuncture due to past experiences or the risks of adverse health events such as bruising, hematomas, syncope, and general discomfort that has the potential to affect performance. Advances in capillary whole blood collection technology now affords the ability to collect micro-volumetric capillary whole blood from the upper arm (or other suitable vascular location such as the abdomen) that is "needle-free" and virtually painless using devices such as the Tasso+. The present study extends previous work, by collecting microliter capillary whole blood samples via the Tasso+ EDTA device in an official anti-doping setting prior to competition, as well as requiring temperature-monitored cold chain shipping by air to the laboratory before analysis. Fifty-eight matched capillary and venous blood samples were collected under official doping control conditions by certified Doping Control Officers. No impact of sample shipment by air under cool conditions was observed on sample integrity. Provided that no visible clots were identified prior to analysis, capillary and venous blood samples showed excellent laboratory agreement for all CBC parameters, with the exception of platelets. Micro capillary blood collection provides a valid alternative to venous blood collection for ABP purposes, with the advantage of providing a more athlete-friendly experience, particularly close to competition.

Detection of capromorelin in urine following oral and dermal routes of administration.

Capromorelin is a growth hormone secretagogue. Despite promising results to alleviate muscle-wasting in the elderly, it has not advanced further in human development. Subsequent studies demonstrated capromorelin's ability to increase food intake in animals, leading to approval in the United States and Europe as an appetite stimulant for cats (Elura) and dogs (Entyce). Capromorelin is prohibited in sports due to its ability to stimulate growth hormone production and enhance performance. However, given that its veterinary preparation is formulated as a highly concentrated solution (20 or 30 mg/mL) delivered orally, incidental ingestion or dermal absorption may result in an adverse analytical finding (AAF) by way of direct exposure during oral administration to a pet. An administration study was conducted by either oral or transdermal application of capromorelin solution to mimic the scenario of inadvertent exposure to the drug. Ingestion of 30 µg of capromorelin orally (equivalent to 1 µL of Entyce) resulted in detectable amounts of capromorelin in urine for up to 48 h after administration with a maximum urinary concentration of 7 ng/mL. Importantly, when applied directly to the skin on the hands in larger quantities mimicking a pet administration exposure scenario (30 mg or 1 mL of Entyce), capromorelin was also detected reaching a maximum urinary concentration of 0.7 ng/mL. Athletes and testing authorities should be aware of the risk of an AAF arising due to incidental exposure to veterinary preparations of capromorelin. To our knowledge, before 2022, no positive test for capromorelin had ever been reported.

δ13 C values of urinary 19-norandrosterone in antidoping samples and potential for adverse findings from boar offal consumption.

19-Norandrosterone (19NA) is the preferred urinary target compound to identify doping with nandrolone or related 19-norsteroids. At concentrations between 2.5 and 15 ng/mL, isotope ratio mass spectrometry (IRMS) is required to establish exogenous origin of urinary 19NA. An absolute difference of 3‰ between urinary 19NA and an endogenous reference compound (ERC) constitutes a finding for exogenous origin of 19NA. Over the last 3 years, 77 samples containing urinary 19NA between 2.5 and 15 ng/mL were analyzed at our laboratory. The measured δ13 C values for 19NA ranged from -29.5‰ to -16.8‰. In comparison, the δ13 C values for the corresponding urinary ERCs ranged from -22.4‰ to -16.2‰. Due to the considerable overlap in values between the target compound and the natural range of urinary ERCs, it can be challenging to distinguish between endogenous and exogenous origins of urinary 19NA. In addition, it is well known that consumption of offal from non-castrated pigs can produce 19NA in urine. To determine whether this could cause a positive IRMS finding under the current IRMS positivity criteria, meat from non-castrated boars fed a mixture of corn and soy was consumed by 13 volunteers. Two volunteers produced 19NA findings above 2.5 ng/mL, and the measured isotope values, while inconsistent with documented 19-norsteroid preparations, did meet IRMS positivity criteria. However, these increases in 19NA urinary concentrations were short-lived due to rapid elimination. Timely follow-up collections may help support a claim for dietary exposure when low urinary concentrations of 19NA with pseudo-endogenous isotope values are observed.

Feasibility of microvolumetric capillary whole blood collections for usage in Athlete Biological Passport analysis.

The hematological module of the Athlete Biological Passport (ABP) represents an important tool in the pursuit to detect blood doping in athletes. Currently, collecting blood samples for ABP analysis can be cumbersome, invasive, and expensive, involving a venous blood draw performed by a trained phlebotomist followed by cold-chain monitored shipping to the analysis laboratory. Developing innovative methods to collect and transport ABP blood samples while adhering to strict preanalytical and analytical requirements has the potential to greatly increase testing frequency and, consequently, the effectiveness of the ABP program globally. The focus of this study was to compare venous blood collections with capillary blood collections to determine if capillary samples could be used for ABP analysis without sacrificing the analytical integrity required for antidoping testing procedures. In this study, capillary blood was collected using the Tasso+ EDTA device (Tasso, Inc.), a novel microvolumetric device that collects liquid, whole blood from skin capillaries on the upper arm. Excellent laboratory agreement was observed between venous and capillary blood samples for the three main ABP parameters: HGB, RET%, and OFF-Score. Additionally, the stability of capillary samples after storage at 4°C, similar to what would be required during transport, was acceptable for up to 72 h following collection. Finally, we generated individual ABP profiles using the adaptive model for 10 participants and observed excellent agreement between venous and capillary profiles. These results indicate capillary blood collection is a viable alternative to venous blood collections for ABP analysis.

Generic Pharmaceuticals as a Source of Diuretic Contamination in Athletes Subject to Sport Drug Testing.

This paper describes nine instances of positive anti-doping tests that could be accounted for by the use of permitted generic prescription drugs contaminated with diuretics, which are prohibited in sport at all times under the WADA Prohibited List. The contamination levels found in the medications are reported and were below FDA limits for manufacturers that are based primarily on safety considerations. These cases demonstrate that great care must be taken to identify the source of low-level anti-doping positives for diuretics reported by WADA-accredited laboratories, and possibly other prohibited substances as well, in order to avoid sanctioning innocent athletes. An evaluation of the cases in this paper supports an approach which establishes a laboratory minimum reporting level (MRL) for diuretics found most commonly in medications. A global consensus after extensive review of similar anti-doping cases has resulted in implementation of a recently announced solution regarding potential diuretic contamination cases.

Sports drug testing and the athletes' exposome.

Similar to the general population, elite athletes are exposed to a complex set of environmental factors including chemicals and radiation and also biological and physical stressors, which constitute an exposome that is, unlike for the general population, subjected to specific scrutiny for athletes due to applicable antidoping regulations and associated (frequent) routine doping controls. Hence, investigations into the athlete's exposome and how to distinguish between deliberate drug use and different contamination scenarios has become a central topic of antidoping research, as a delicate balance is to be managed between the vital and continually evolving developments of sensitive analytical techniques on the one hand, and the risk of the athletes' exposome potentially causing adverse analytical findings on the other.

Chromatographic-mass spectrometric analysis of the urinary metabolite profile of chlorphenesin observed after dermal application of chlorphenesin-containing sunscreen.

RATIONALE: Chlorphenesin is an approved biocide frequently used in cosmetics, and its carbamate ester is an approved skeletal muscle relaxant in certain countries for the treatment of discomfort related to skeletal muscle trauma and inflammation. A major urinary metabolite is 4-chlorophenoxy acetic acid (4-CPA), also known as para-chlorophenoxyacetate, which is also employed as a target analyte in sports drug testing to detect the use of the prohibited nootropic stimulant meclofenoxate. To distinguish between 4-CPA resulting from chlorphenesin, chlorphenesin carbamate, and meclofenoxate, urinary metabolite profiles of chlorphenesin after legitimate use were investigated. METHODS: Human administration studies with commercially available sunscreen containing 0.25% by weight of chlorphenesin were conducted. Six study participants dermally applied 8 g of sunscreen and collected urine samples before and up to 7 days after application. Another set of six study participants applied 8 g of sunscreen on three consecutive days, and urine samples were also taken for up to 5 days after the last dosing. Urine specimens were analyzed using liquid chromatography-high resolution (tandem) mass spectrometry, and urinary metabolites were identified in accordance with literature data by accurate mass analysis of respective precursor and characteristic product ions. RESULTS: In accordance with literature data, chlorphenesin yielded the characteristic urinary metabolites, chlorphenesin glucuronide, chlorphenesin sulfate, and 3-(4-chlorophenoxy)-2-hydroxypropanoic acid (4-CPP), as well as the common metabolite 4-CPA. 4-CPA and 4-CPP were observed at similar abundances, with urinary concentrations of 4-CPA reaching up to ~1500 and 2300 ng/mL after single and multiple sunscreen applications, respectively. CONCLUSION: 4-CPA is a common metabolite of meclofenoxate, chlorphenesin, and chlorphenesin carbamate. Monitoring the diagnostic urinary metabolites of chlorphenesin provides conclusive supporting evidence of whether chlorphenesin or the prohibited nootropic meclofenoxate was administered.

Investigating oral fluid and exhaled breath as alternative matrices for anti-doping testing: Analysis of 521 matched samples.

Current anti-doping testing is primarily conducted in urine and blood. Recently, due to confounding factors with urine and blood collections such as invasiveness, cost, and stringent shipping conditions, there has been a push for the use of alternative sample matrices to ameliorate these issues. Gaining support within the anti-doping field is the use of oral fluid, and more recently exhaled breath, as viable alternative or complementary matrices to traditional urine and blood for drug testing. Thus, we designed a first-in-field study with the purpose of investigating the utility of oral fluid and exhaled breath testing, and the preference of athlete participants, comparative to conventional anti-doping methods of urine testing. To accomplish this, 521 total matched samples, consisting of exhaled breath, oral fluid, and urine samples, were collected and analyzed, and the results compared across matrices. Participants in this study preferred the exhaled breath collection (rated 4.90 ±â€¯0.34 out of 5, mean ±â€¯SD) over the oral fluid collection procedure (4.29 ±â€¯0.85), and most preferred both over urine collections. Exhaled breath resulted in the shortest collection time (2.58 ±â€¯1.00 min, mean ±â€¯SD), followed by urine (3.08 ±â€¯1.50 min), and finally oral fluid (4.14 ±â€¯1.94 min). Prohibited substances from the drug categories of stimulants, narcotics, cannabinoids, diuretics, glucocorticoids, beta-blockers, and others, were analyzed in this study for a comparison of testing efficacy. Of the total findings 49% were detectable in only urine, 38% in urine + oral fluid, and 9% in all three matrices. Of the unique findings 3% were detectable in only oral fluid, 1% in oral fluid + breath, and 0% of unique findings were present only in exhaled breath. The findings from this study provide a strong foundation for the future use of oral fluid and exhaled breath as viable alternative or complementary matrices for in-competition anti-doping testing.

Essential Features of Third-Party Certification Programs for Dietary Supplements: A Consensus Statement.

The presence of performance-enhancing drugs in dietary supplements poses serious anti-doping and health risks to athletes and military service members. A positive drug test, suboptimal health, or adverse event can ruin a career in either setting. These populations need to be certain in advance that a product is of high quality and free from performance-enhancing drugs and other banned substances. However, no regulatory authority conducts or mandates a quality review before dietary supplements are sold. Under the Food Drug and Cosmetic Act, the Food and Drug Administration does not have a role in the premarket safety review of dietary supplements. Due to the increasing demand for high-quality, properly labeled dietary supplements, multiple companies have stepped into this void by offering testing and quality review programs for dietary supplements. Each of these third-party programs has its own quality assurance program with varying testing components. It is difficult for consumers in the sport and military settings to assess whether a particular certification program reduces the risks enough so that they can use a product with confidence. This article puts forward the consensus of the authors on current best practices for third-party certification programs for dietary supplements consumed by athletes and military service members. Also discussed are important ways that third-party programs can develop in the future to improve access to safe, high-quality dietary supplements for these populations.

Trading Health Risks for Glory: A Reformulation of the Goldman Dilemma.

BACKGROUND: The Goldman dilemma presented athletes with a Faustian bargain that guaranteed winning an Olympic gold medal in their sport but resulted in certain death 5 years later. Athletes' responses to Goldman's bargain were reported from 1982 to 1995. Several studies subsequently evaluated people's willingness to accept the bargain proposed in the Goldman question. Our study updates Goldman's question using contingent-behavior questions, a preference-elicitation method widely applied in economics, marketing and psychology to understand people's choice behavior. Contingent-behavior questions ask people to evaluate hypothetical tradeoffs between outcomes when real-world decisions are unobservable, nonexistent, or unreliable. METHODS: A web-enabled survey was conducted with athletes in 50 sports between June, 2012 and April, 2013. Athletes were invited by their sport governing bodies in the United States to complete the online survey. Responses from 2888 athletes were collected. Our reformulation elicited athletes' willingness to accept a performance-enhancing drug (PED) associated with the risk of a realistic fatal event, not certain death. A double-bounded dichotomous-choice question format was used to elicit athletes' maximum acceptable mortality risk (MAMR) for winning an Olympic gold medal. Data were analyzed using an interval regression model to estimate the implicit probability of accepting a continuous risk level. MAMR was defined as the mortality risk level with a 0.50 probability of acceptance. RESULTS: Estimated mean MAMRs varied between 7 and 14% across athletes in different ranks and sports. Elite athletes were generally the most willing to accept a fatal cardiovascular risk to win a gold medal in the Olympics. This range was similar to the levels of risk that patients accept for life-changing interventions. CONCLUSIONS: Results suggest that very few athletes would be expected to accept a PED in the bargain postulated by the Goldman dilemma. Risk tolerance among elite athletes suggest they may be more aware of the potential financial and nonfinancial benefits of such a win, and/or less optimistic about their potential to move up in the level of competition without the use of PEDs.

Treating the elite athlete: anti-doping information for the health professional.

Physicians and health professionals are a vital component in preserving the integrity of competition and the core principles of true sport. When treating an athlete, health professionals need to be cognizant of the anti-doping rules of the relevant sport organization. This review aims to provide an overview of the World Anti-Doping Agency Prohibited List, Therapeutic Use Exemptions, roles and responsibilities of the health professional, as well as provide resources that will guide their work with athletes.

Human genetic variation: new challenges and opportunities for doping control.

Sport celebrates differences in competitors that lead to the often razor-thin margins between victory and defeat. The source of this variation is the interaction between the environment in which the athletes develop and compete and their genetic make-up. However, a darker side of sports may also be genetically influenced: some anti-doping tests are affected by the athlete's genotype. Genetic variation is an issue that anti-doping authorities must address as more is learned about the interaction between genotype and the responses to prohibited practices. To differentiate between naturally occurring deviations in indirect blood and urine markers from those potentially caused by doping, the "biological-passport" program uses intra-individual variability rather than population values to establish an athlete's expected physiological range. The next step in "personalized" doping control may be the inclusion of genetic data, both for the purposes of documenting an athlete's responses to doping agents and doping-control assays as well facilitating athlete and sample identification. Such applications could benefit "clean" athletes but will come at the expense of risks to privacy. This article reviews the instances where genetics has intersected with doping control, and briefly discusses the potential role, and ethical implications, of genotyping in the struggle to eliminate illicit ergogenic practices.

The effects of exercise in hypoxic and normoxic conditions on endothelin-1 and arterial compliance.

The purpose of this study was to determine the effects of short-term normoxic and hypoxic exercise on plasma endothelin-1 and nitric oxide levels, and the relationship of arterial compliance and pulmonary artery pressure to endothelin-1. Seven endurance-trained males completed two incremental and two steady-state exercise tests performed at ventilatory threshold in normoxia and hypoxia (fraction of inspired oxygen = 0.14). Plasma endothelin-1was measured throughout steady-state tests. Arterial compliance using applanation tonometry, plasma nitric oxide and pulmonary artery pressure using Doppler echocardiography were measured before and after exercise. Small arterial compliance and pulmonary artery pressure significantly increased following exercise. There were no main effects of condition or time for plasma endothelin-1and nitric oxide levels. There were no significant relationships between plasma endothelin-1 and arterial compliance or pulmonary artery pressure. In conclusion, mechanisms other than the endothelial system may play a role in the exercise-induced changes in small artery compliance in this study population. Moderate hypoxia and a 30-minute steady-state exercise have limited effects on plasma endothelin-1 in endurance-trained males.

Keeping pace with ACE: are ACE inhibitors and angiotensin II type 1 receptor antagonists potential doping agents?

In the decade since the angiotensin-converting enzyme (ACE) gene was first proposed to be a 'human gene for physical performance', there have been numerous studies examining the effects of ACE genotype on physical performance phenotypes such as aerobic capacity, muscle function, trainability, and athletic status. While the results are variable and sometimes inconsistent, and corroborating phenotypic data limited, carriers of the ACE 'insertion' allele (the presence of an alu repeat element in intron 16 of the gene) have been reported to have higher maximum oxygen uptake (VO2max), greater response to training, and increased muscle efficiency when compared with individuals carrying the 'deletion' allele (absence of the alu repeat). Furthermore, the insertion allele has been reported to be over-represented in elite athletes from a variety of populations representing a number of endurance sports. The mechanism by which the ACE insertion genotype could potentiate physical performance is unknown. The presence of the ACE insertion allele has been associated with lower ACE activity (ACEplasma) in number of studies, suggesting that individuals with an innate tendency to have lower ACE levels respond better to training and are at an advantage in endurance sporting events. This could be due to lower levels of angiotensin II (the vasoconstrictor converted to active form by ACE), higher levels of bradykinin (a vasodilator degraded by ACE) or some combination of the two phenotypes. Observations that individuals carrying the ACE insertion allele (and presumably lower ACEplasma) have an enhanced response to training or are over-represented amongst elite athletes raises the intriguing question: would individuals with artificially lowered ACEplasma have similar training or performance potential? As there are a number of drugs (i.e. ACE inhibitors and angiotensin II type 1 receptor antagonists [angiotensin receptor blockers--ARBs]) that have the ability to either reduce ACEplasma activity or block the action of angiotensin II, the question is relevant to the study of ergogenic agents and to the efforts to rid sports of 'doping'. This article discusses the possibility that ACE inhibitors and ARBs, by virtue of their effects on ACE or angiotensin II function, respectively, have performance-enhancing capabilities; it also reviews the data on the effects of these medications on VO2max, muscle composition and endurance capacity in patient and non-patient populations. We conclude that, while the direct evidence supporting the hypothesis that ACE-related medications are potential doping agents is not compelling, there are insufficient data on young, athletic populations to exclude the possibility, and there is ample, albeit indirect, support from genetic studies to suggest that they should be. Unfortunately, given the history of drug experimentation in athletes and the rapid appropriation of therapeutic agents into the doping arsenal, this indirect evidence, coupled with the availability of ACE-inhibiting and ACE-receptor blocking medications may be sufficiently tempting to unscrupulous competitors looking for a shortcut to the finish line.

RNAi-mediated gene silencing in non-human primates.

The opportunity to harness the RNA interference (RNAi) pathway to silence disease-causing genes holds great promise for the development of therapeutics directed against targets that are otherwise not addressable with current medicines. Although there are numerous examples of in vivo silencing of target genes after local delivery of small interfering RNAs (siRNAs), there remain only a few reports of RNAi-mediated silencing in response to systemic delivery of siRNA, and there are no reports of systemic efficacy in non-rodent species. Here we show that siRNAs, when delivered systemically in a liposomal formulation, can silence the disease target apolipoprotein B (ApoB) in non-human primates. APOB-specific siRNAs were encapsulated in stable nucleic acid lipid particles (SNALP) and administered by intravenous injection to cynomolgus monkeys at doses of 1 or 2.5 mg kg(-1). A single siRNA injection resulted in dose-dependent silencing of APOB messenger RNA expression in the liver 48 h after administration, with maximal silencing of >90%. This silencing effect occurred as a result of APOB mRNA cleavage at precisely the site predicted for the RNAi mechanism. Significant reductions in ApoB protein, serum cholesterol and low-density lipoprotein levels were observed as early as 24 h after treatment and lasted for 11 days at the highest siRNA dose, thus demonstrating an immediate, potent and lasting biological effect of siRNA treatment. Our findings show clinically relevant RNAi-mediated gene silencing in non-human primates, supporting RNAi therapeutics as a potential new class of drugs.