On the trail of blood doping—microRNA fingerprints to monitor autologous blood transfusions in vivo

Abstract

Autologous blood doping refers to the illegal re‐transfusion of any quantities of blood or blood components with blood donor and recipient being the same person. The re‐transfusion of stored erythrocyte concentrates is particularly attractive to high‐performance athletes as this practice improves their oxygen capacity excessively. However, there is still no reliable detection method available. Analyzing circulating microRNA profiles of human subjects that underwent monitored autologous blood transfusions seems to be a highly promising approach to develop novel biomarkers for autologous blood doping. In this exploratory study, we randomly divided 30 healthy males into two different treatment groups and one control group and sampled whole blood at several time points at baseline, after whole blood donation and after transfusion of erythrocyte concentrates. Hematological variables were recorded and analyzed following the adaptive model of the Athlete Biological Passport. microRNA profiles were examined by small RNA sequencing and comprehensive multivariate data analyses, revealing microRNA fingerprints that reflect the sampling time point and transfusion volume. Neither individual microRNAs nor a signature of transfusion‐dependent microRNAs reached superior sensitivity at 100% specificity compared to the Athlete Biological Passport (≤11% 6 h after transfusion versus ≤44% 2 days after transfusion). However, the window of autologous blood doping detection was different. Due to the heterogenous nature of doping, with athletes frequently combining multiple medications in order to both gain a competitive advantage and interfere with known testing methods, the true applicability of the molecular signature remains to be validated in real anti‐doping testings.

A panel of blood-derived miRNAs with a stable expression pattern as a potential pan-cancer detection signature

“…Our input criteria only include datasets that obtained blood. The RNA seq raw data for healthy individuals ( n = 90) were obtained from the study of ( Haberberger et al, 2018 ; Mussack et al, 2021 ), including whole blood samples of ten healthy male individuals at nine time-points (0 h, 3 h, 6 h, 24 h, 48 h, 72 h, Day7, Day30, and day60). A table of clinical information of 10 healthy individuals was provided in Supplementary Table S1 ( Supplementary File S1 ).…”

“…Eleven different cancer datasets with their specific controls in nine different tissues. One time series dataset obtained from ten healthy individuals including 90 samples (Mussack et al, 2021). All the samples were obtained from blood in each cancer.…”

Indirect biomarkers of blood doping: A systematic review

“…34,98,103 Nevertheless, a recent study comparing the effectiveness of miRNA to ABP biomarkers in the monitoring of ABT tempered the absolute effectiveness of the transcriptomic approach. 38 Indeed, despite an interesting response of miRNA markers following blood transfusion, the detection window did not appear to be significantly longer in comparison with traditional markers. Therefore, if the heterogeneity in individual responses seems to be the major limitation when using a single marker, 37 a combination of multiple markers in a mathematical model completed with existing variables would certainly be the most promising approach.…”

Annual banned‐substance review: Analytical approaches in human sports drug testing 2020/2021

“…Investigations into options to further optimize the ABP by implementing additional biomarkers have continued, for instance by monitoring microRNA (miRNA) fingerprints indicative for autologous blood doping. Mussack et al 149 simulated blood doping scenarios with 20 study volunteers, where either one or two units of blood were donated, processed to erythrocyte concentrates, and re‐infused after cold storage 28 or 42 days after withdrawal, respectively. Circulating microRNA profiles were prepared, which yielded promising biomarker candidates such as miR144‐3p and miR‐320d that orthogonally complement the established hematological markers.…”