Gene doping detection: evaluation of approach for direct detection of gene transfer using erythropoietin as a model system.
As clinical gene therapy has progressed toward realizing its potential, concern over misuse of the technology to enhance performance in athletes is growing. Although 'gene doping' is banned by the World Anti-Doping Agency, its detection remains a major challenge. In this study, we developed a methodology for direct detection of the transferred genetic material and evaluated its feasibility for gene doping detection in blood samples from athletes. Using erythropoietin (EPO) as a model gene and a simple in vitro system, we developed real-time PCR assays that target sequences within the transgene complementary DNA corresponding to exon/exon junctions. As these junctions are absent in the endogenous gene due to their interruption by introns, the approach allows detection of trace amounts of a transgene in a large background of the endogenous gene. Two developed assays and one commercial gene expression assay for EPO were validated. On the basis of ability of these assays to selectively amplify transgenic DNA and analysis of literature on testing of gene transfer in preclinical and clinical gene therapy, it is concluded that the developed approach would potentially be suitable to detect gene doping through gene transfer by analysis of small volumes of blood using regular out-of-competition testing.
Baoutina
A
A
Bioanalysis, National Measurement Institute, New South Wales, Australia. anna.baoutina@measurement.gov.au
Coldham
T
T
Bains
G S
GS
Emslie
K R
KR
D056352:Athletes; D054874:Athletic Performance; D004300:Doping in Sports / Q000517:prevention & control*; D004921:Erythropoietin / Q000235:genetics*; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D019076:Transgenes
D004921:Erythropoietin