Matthew Fedoruk’s research while affiliated with University of Colorado Colorado Springs and other places

Blood collection is an important facet of anti‐doping testing, forming the basis of the hematological module of the athlete biological passport (ABP). Presently, whole blood is collected via venepucture under standardized conditions, in accordance with the World Anti‐doping Agency's International Standard for Testing and Investigations (ISTI). Advances in capillary whole blood collection technology now afford the ability to collect microvolumetric capillary whole blood from the upper arm (or other suitable vascular location such as the abdomen) that is “needle‐free” and virtually painless. Previous work has demonstrated the suitability and feasibility of capillary whole blood compared with venous using the Tasso+ device. Recently, other microcapillary devices have received FDA approval increasing the options available to the anti‐doping community. The present study extends previous work, by collecting microliter capillary whole blood samples via two different FDA approved devices (RedDrop and the Tasso+ device) to compare with venous blood collected simultaneously. Ten participants provided three matched blood samples (2× capillary and 1× venous) in accordance with WADA ABP guidelines for blood sample collection, for a total of 30 samples. Capillary samples from both devices showed excellent laboratory agreement with venous blood for all CBC parameters, with the exception of platelets. Excellent laboratory agreement was also observed between the two microcapillary collection devices. Irrespective of the device, microcapillary blood collection provides a valid alternative to venous blood collection for ABP purposes.

Roxadustat (FG‐4592), an orally active hypoxia‐inducible factor prolyl hydroxylase stabilizer, has been shown to enhance erythropoiesis by increasing endogenous erythropoietin. It is indicated for the treatment of anemia and chronic kidney disease and is approved for clinical use in several countries, including the European Union, Japan and others. Due to its reasonably anticipated performance‐enhancing effect in athletes, roxadustat is prohibited for use in sports at all times. A few cases of adverse analytical findings in routine doping controls have been reported worldwide, some of which were claimed to be the result of contaminated dietary supplements. The present study offers new data demonstrating the long‐term excretion pattern of roxadustat. Even after a single‐dose administration, roxadustat can remain detectable in urine for 8 months, albeit at very low concentrations (<10 pg/mL). Following three times a week treatment with 70 to 100 mg of roxadustat, the drug was still detectable in the urine of anemic patients for between 9 and 18 months after treatment was discontinued. Lastly, an athlete who admitted use of roxadustat for almost a year (50 mg 3 to 5 times a week) has now tested positive multiple times over the course of 15 months (the first test being 12 months after the drug was discontinued), with an estimated concentration of roxadustat between 3 and 8 pg/mL. Altogether, these findings indicate the unusually prolonged terminal excretion kinetics of roxadustat, a property that testing authorities should consider in their results management process.

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.

Purpose Inadvertent and/or unknowing exposure to drugs and drug residues has been frequently debated in situations of so-called adverse analytical finding (AAF) in the context of sports drug testing programs. Transfer of drug residues via unprotected intercourse is a conceivable scenario but scientific data and authentic case reports are scarce. Herein, investigations into two AAFs with the peroxisome proliferator-activated receptor delta (PPARδ) agonist GW1516 are reported and discussed. Methods To probe for a contamination scenario involving sexual intercourse, two assays were used to determine semenogelin in human urine, with one employing an immunochromatographic lateral flow approach and another based on liquid chromatography–tandem mass spectrometry. Further, drug-residue testing using patients’ ejaculate was conducted by utilizing liquid chromatography in conjunction with a triple quadrupole mass spectrometer, followed by re-analysis of suspect samples (i.e., samples indicating the presence of relevant compounds) using high resolution/high mass accuracy mass spectrometry. Results In one case, but not the other, the possibility of intimate contact as the source of the AAF was confirmed after a thorough investigation of potential contamination scenarios. Subsequent research revealed analytical evidence for the presence of seminal fluid in one of the female athlete’s doping control urine samples, and the analysis of clinical ejaculate specimens provided first data on an authentic concentration level of GW1516 and its metabolites in human seminal fluid. Conclusions The combined facts substantiate the possibility of an AAF caused by unprotected sexual intercourse and the plausibility of the case-related arguments.

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.

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 Agency (WADA) Prohibited List. This study used a web-delivered survey and direct questioning approach to collect data from athletes in the United States Anti-Doping Agency (USADA) Registered Testing Pool or Clean Athlete Program. For the first time, a prevalence survey of elite athletes used the WADA Prohibited List to assess prevalence in each specific category of banned substance / method. Results: Of 1,425 respondents, overall doping prevalence among U.S. elite athletes in the 12 months prior to survey administration ranged from an estimated 6.7-9.2% depending on the method of calculation. Specific doping prevalence rates for each individual substance category ranged from 0.1% (diuretics or masking agents) to 4.1% for cannabinoids (in-competition). Conclusion: By measuring doping prevalence down to specific performance enhancing substances and methods, as well as coupling with individual athlete personality characteristics such as attitudes, motivators and norms, a clearer picture of the use of specific prohibited substances in- and out of-competition, attitudes toward doping, beliefs about testing and most effective deterrents can be identified. By understanding doping prevalence, anti-doping programs can be tailored to increase deterrence and overall program effectiveness.

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.

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. 2 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.

Similar to the general population, elite athletes are exposed to a complex set of environmental factors including chemicals, radiation, but 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 anti‐doping 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 anti‐doping 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.

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 (pCPA), which is also employed as target analyte in sports drug testing to detect the use of the prohibited nootropic stimulant meclofenoxate. In order 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 post-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 by 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 approximately 1500 ng/mL and 2300 ng/mL after single and multiple sunscreen application, 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 whether chlorphenesin or the prohibited nootropic meclofenoxate was administered.