Discovering red cell proteome biomarkers for identification of recombinant human erythropoietin misuse and hypoxic exposure
By
Investigateur principal
N.
Nordsborg
University of Copenhagen
Danemark
―
2021
―
En vigueur
Sommaire
Code: 21T07NN
Detection of blood doping remain a challenge, and novel detection methods are warranted. In the present project, the aim is to evaluate the potential of changes in red blood cell proteins for detection of recombinant human erythropoietin (rhEpo) misuse at sea level and at altitude in males and females. The red cell protein content is expected to change with rhEpo treatment, as the transcription of genes coding for red cell proteins change during blood doping practices. Indeed, changes in a few proteins of red blood cells are known to change with rhEpo misuse. However, whether the changes in these proteins are sensitive and specific to rhEpo misuse and whether more sensitive and more specific proteins exist remains unknown. Additionally, it remains unknown whether hypoxic exposure or gender are confounding factors to these changes. To answer these questions, the present project will investigate more than 1000 different red blood cell proteins using mass spectrometry and identify the most sensitive and specific ones. Further, the present project will develop and validate a model for rhEpo detection based on the detected proteins. Our goal is to develop a ready-to-use robust method that will allow precise quantification of our novel biomarkers within approximately 4 h, from receiving the sample to the result.
Ergogenic effects of new-generation beta2-agonists vilanterol and indacaterol
By
Investigateur principal
V.
Backer
Danemark
―
2021
―
En vigueur
Sommaire
Code: 21C20VB
This project aims to determine whether new-generation ultra long-acting beta2-agonist (U-LABA) formulations containing vilanterol and indacaterol can enhance performance when inhaled at therapeutic or supratherapeutic doses in highly-trained males and females. The project will also examine whether U-LABA can induce muscle hypertrophy when used chronically.
Objectives: This project will investigate the potential performance-enhancing effects of vilanterol and indacaterol at therapeutic and supratherapeutic inhaled doses. THe formulations tested are Relvar® (22 µg/dose vilanterol+fluticasone 55 µg/dose) and Atectura® (indacaterol 125 µg/dose+mometasone 62 µg/dose) as these are U-LABA formulations of relevance for asthma treatment.
The project deal with the synthesis and analytical characterization of an epimeric long-term metabolite of the anabolic steroid Dehydrochloromethyltestosterone (4a-chloro-18-nor-17b-hydroxymethyl-17a-methyl-5a-androst-13-en-3a-ol)
Synthesis of 17-epi-Oxymesterone long term metabolite
By
Investigateur principal
G.
Gmeiner
Seibersdorf Labor GmbH
Autriche
―
2023
―
En vigueur
Sommaire
Code: T23A03GG
The project deal with the synthesis and analytical characterization of a long term metabolite of the anabolic steroid oxymesterone (17β-methyl-4,17α-dihydroxy-androst-4-en-3-one)
The project deal with the synthesis and analytical characterization of the glucuronide phase II metabolite of the stimulant Adrafinil (Diphenylmethyl)sulfinyl]-N-hydroxyacetamide)
The project deal with the synthesis and analytical characterization of the glucuronide phase II metabolite of the anabolic agent Andarine (2S)-3-(4-acetamidophenoxy)-2-hydroxy-2-methyl-N-[4-nitro-3-(trifluoromethyl)phenyl]propanamide)
Detection of genetic polymorphisms of the human Erythropietin gene in urine, whole blood and dried blood spots
By
Investigateur principal
F.
Donati
Laboratorio Antidoping FMSI
Italie
―
2023
―
En vigueur
Sommaire
Code: 23E05FD
Several polymorphisms/variants of the human EPO gene could be the cause of confounding effects in the application of the analytical methods that are routinely applied for the detection of the abuse of recombinant erythropoietin (recEPO). The most known of them is the c.577del (rs369859204) variant, a single nucleotide deletion in the position 577 of the human EPO gene exon 5. The deletion causes a shifting of the sequence within the terminal exon 5 with consequent loss of the stop codon. At translational level, this results in the formation of a longer mature protein with a higher molecular weight of 3.3 kDa. In doping control, this results in the visualization of a “double band” during the application of the routinary SAR-Page method for the detection of the abuse of recEPO. The double band may be then erroneously interpreted as a false positive sample. As stated in the WADA technical document TDEPO2022, it is suggested to screen for the presence of the c.577del variant whenever a suspected double band is detected by the SAR-Page analysis, in the final aim to avoid the possibility of a false positive finding. Moreover, at least 12 other polymorphisms/variants have been described in the literature that are potentially capable of generating confounding effects in the interpretations of the antidoping analyzes. By following the input given by WADA TDEPO2022, this project involves the development and validation of a genetic test, based on the genomic DNA Sanger sequencing method, that is capable to identify the c.577del variant and all the other polymorphisms/variants present on the human EPO gene (namely: rs1554393458, rs1554393463, rs1419397684, rs773895305, rs763035217, rs1562901775, rs71517124, rs1562902091, rs893404064, rs752755372, rs951322017, rs369859204), and the study of their incidence in determining confounding effects in the interpretation of the routinary tests performed for the purpose of detecting the abuse of recombinant EPO by athletes.
Detection of rEPO administration using deep learning on blood smears
By
Investigateur principal
S.
Voss
IDAS Dresden
Allemagne
―
2023
―
En vigueur
Sommaire
Code: 23D05SV
The aim of this study is to use artificial intelligence (AI) to evaluate blood smears as a potential matrix to detect doping with recombinant EPO (rEPO). Previous publications reported changes in the Red blood Cell Morphology after rEPO administration (e.g. Macrocytes, Stomatocytes). During these times automated haematology analysers and manual microscopy were used for the estimation of these parameters. Recently, digital Morphology is a developing field in Haematology which enables the automated analysis of blood smears by artificial intelligence. With this technology it is possible to evaluate morphological changes with an increased precision, based on a higher number of cells and to discover even minor changes in cellular shapes. AI and deep learning is capable of revealing new insights which conventional approaches were lacking so far, like predicting molecular changes on cytomorphology. Therefore, the goal of this study is to identify relevant changes in cell morphology during rEPO administration which are not addressed using current state of the art techniques. In the long term, a blood smear-based athlete blood passport providing an individual erythrocyte signature might be a prospective application of monitoring athletes by using artificial intelligence based on this postulated deep learning model.
Investigations into the metabolism and elimination of flmodafinil and fladrafinil as well as their prevalence in elite sports
By
Investigateur principal
M.
Thevis
German Sport University
Allemagne
―
2023
―
En vigueur
Sommaire
Code: 23C03MT
Stimulants are substances with wake-promoting effects, whose misuse in sports is only prohibited in-competition. The high analytical sensitivity of anti-doping laboratories allows for comprehensive screening procedures, which cannot only reveal the misuse of stimulants, but also unintended doping due to the use of contaminated/adulterated nutritional supplements. Therefore, stimulants constantly account for more than 10% of all adverse analytical findings (AAF) reported by the World Anti-Doping Agency (WADA). Due to the easy detectability of prohibited stimulants, especially novel drugs such as flmodafinil and fladrafinil have an enormous potential for being misused in sports. These bisfluoro analogs of modafinil and its pro-drug adrafinil are supposed to be significantly more effective than the original compounds, but currently not explicitly mentioned in the WADA Prohibited List. Consequently, the aim of this research project is to investigate the urinary metabolism of flmodafinil and fladrafinil, as well as their elimination behavior in urine and blood. The acquired data will lay the foundation for the implementation of both drugs as well as their metabolites into doping control routine procedures. Moreover, a retrospective evaluation of in-competition routine samples will be conducted to determine the prevalence of these stimulants in sports.
Inhibitors of the activin receptor signaling pathways (IASPs) in sports drug testing
By
Investigateur principal
M.
Thevis
German Sport University
Allemagne
―
2023
―
En vigueur
Sommaire
Code: 23A13MT
Members of the transforming growth factor beta (TGF-β) superfamily such as myostatin (MSTN), activin A, and GDF-11, are dimeric cytokines signaling through activin receptors. They play important regulative roles in different biological processes as for example the formation of muscle and red blood cells. Therefore, inhibitors of the activin receptor signaling pathways (IASPs) can be considered as potential performance-enhancing agents in sports, which are included in sections S2 (“Peptide hormones, growth factors, related substances and mimetics”) and S4 (“Hormone and metabolic modulators”) of the WADA Prohibited List. They comprise targeted (e.g. anti-MSTN antibodies) as well as multi-targeting approaches (e.g. receptor competitors and anti-ActRII antibodies), and for several drug candidates, doping control detection methods based on electrophoretic techniques and Western blotting or LC-HRMS/MS have been proactively developed within the last years. This is of utmost importance, as, even without clinical approval, reference material for such emerging drugs is often available for research purposes and/or distributed on the black market. Therefore, the aim of this research project is to develop a combined mass spectrometric detection assay for the novel anti-activin A antibody Garetosmab and other IASPs such as Sotatercept, Domagrozumab, and Luspatercept. A combined extraction of the target analytes from plasma/serum can be achieved by using NHS magnetic sepharose beads coupled to different TGF-β cytokines such as activin A, MSTN, and GDF-11. Subsequent LC-HRMS/MS analysis will not only allow for an unambiguous identification of the target analytes based on the amino acid level, but also retrospective sample evaluation for the presence of novel IASPs binding to one of the TGF-β cytokines employed for affinity purification.