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Abstract
Despite the World Anti-Doping Agency (WADA) ban on gene doping in the context of advancements in gene therapy, the risk of EPO gene-based doping among athletes is still present. To address this and similar risks, gene-doping tests are being developed in doping control laboratories worldwide. In this regard, the present study was performed with two objectives: to develop a robust gene-doping mouse model with the human EPO gene (hEPO) transferred using recombinant adenovirus (rAdV) as a vector and to develop a detection method to identify gene doping by using this model. The rAdV including the hEPO gene was injected intravenously to transfer the gene to the liver. After injection, the mice showed significantly increased whole-blood red blood cell counts and increased expression of hematopoietic marker genes in the spleen, indicating successful development of the gene-doping model. Next, direct and potentially indirect proof of gene doping were evaluated in whole-blood DNA and RNA by using a quantitative PCR assay and RNA sequencing. Proof of doping could be detected in DNA and RNA samples from one drop of whole blood for approximately a month; furthermore, the overall RNA expression profiles showed significant changes, allowing advanced detection of hEPO gene doping.
Highlights
Doping is the act of using prohibited substances and/or methods in sports to enhance athletic performance and success by improving physical performance [1]
We considered that the detection of viral vector-specific DNA fragments by a TaqMan quantitative real-time PCR (qPCR) assay would be the most reasonable approach because the principle underlying this assay has already been successfully used in testing for SARS-CoV-2
As indirect proof, we considered that monitoring fluctuations of RNA expression in whole-blood samples could be applied to the parameters of the Athlete Biological Passport (ABP) because the fundamental principle underlying
Summary
Doping is the act of using prohibited substances and/or methods in sports to enhance athletic performance and success by improving physical performance [1]. The International Standard Prohibited List [3] stipulated in the World Anti-Doping Code 2021 [4], which is published with annual revisions by the WADA, describes various formulations and methods used in doping. This list includes “gene doping” as a form of abuse of gene therapy technology. Our research group has been conducting research for more than five years to establish an examination method for gene doping [5,6,7]
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