Tracking immature reticulocyte proteins for improved detection of recombinant human erythropoietin (rhEPO) abuse

Abstract

Athletes abuse recombinant human erythropoietin (rhEPO) and erythropoiesis stimulating agents to increase hemoglobin mass and improve performance. To evade detection, athletes have developed sophisticated blood doping regimens, which often include rhEPO micro-dosing. Detection of these methods requires biomarkers with increased sensitivity and a sample matrix that is more amenable to frequent testing in the field. We have developed a method to measure two immature reticulocyte proteins, CD71 and ferrochelatase (FECH), and one total erythrocyte protein, Band 3, in dried blood spots (DBS). This method was tested in response to rhEPO administration after low doses, 40 IU/kg, micro-doses, 900 IU, or saline injection in 20 healthy subjects. During administration of low-dose rhEPO, the mean CD71/Band 3 and FECH/Band 3 ratio increased by 412 ± 197% and 250 ± 44%, respectively. The mean response for the current biomarker, RET%, increased by 195 ± 35%. During administration of rhEPO micro-doses, CD71/Band 3 increased to 127 ± 25% on day 35 and 139 ± 36% on day 39, while no increase was observed in RET%. After rhEPO administration, during the washout phase, mean values decreased to a minimum of 64 ± 4% and 64 ± 11% for CD71/Band 3 and RET%, respectively. However, CD71/Band 3 remained below 75% of base-

Supra‐physiological doses of anabolic androgenic steroids impact erythropoietin and blood parameters

“…Additionally, IRF can be used as a marker for rEPO 7 and Cox et al showed that immature reticulocyte proteins measured in dried blood spots (DBS) showed a larger response to rEPO than RET%. 8 Since androgens can also be quantified in DBS, 30 co-monitoring of blood parameters and steroids in DBS may be possible in the future.…”

“…Since, in the future, more samples with both blood parameters and serum hormones may be available, it is of continuous interest to understand the interactions between the different ABP modules. Additionally, IRF can be used as a marker for rEPO 7 and Cox et al showed that immature reticulocyte proteins measured in dried blood spots (DBS) showed a larger response to rEPO than RET% 8 . Since androgens can also be quantified in DBS, 30 co‐monitoring of blood parameters and steroids in DBS may be possible in the future.…”

“…6 Recent studies showed that monitoring of immature reticulocytes (IRF) improved detection of rEPO use. 7,8 Anabolic androgenic steroids (AAS) are forbidden in sports and considered a health problem in society. 9 Testosterone (T) is one of the most reported and detected steroids among AAS users and in sports.…”

“…The hematological biomarkers in the ABP include hemoglobin concentration (HB) and reticulocyte percentage (RET%), 5 of which RET% is particularly sensitive to rEPO use 6 . Recent studies showed that monitoring of immature reticulocytes (IRF) improved detection of rEPO use 7,8 …”

Changes in Immature Reticulocytes Aid the Indirect Detection of Microdose Recombinant Erythropoietin Use in Men and Women

“…A valuable alternative to the expensive and labor-intensive (9) direct detection of rHuEPO is the indirect approach of the Athlete Biological Passport (ABP). The ABP hematological module longitudinally monitors hemoglobin concentration ([Hb]), reticulocyte percentage (RET%), a combined score based on [Hb] and RET% also known as the OFF-hr score defined as [Hb]-60 Â √[RET%] (10), and the multiparametric Abnormal Blood Profile Score (ABPS) (11).…”