Are EPB41 and alpha-synuclein diagnostic biomarkers of sport-related concussion? Findings from the NCAA and Department of Defense CARE Consortium

Background: Current protein biomarkers are only moderately predictive at identifying individuals with mild traumatic brain injury or concussion. Therefore, more accurate diagnostic markers are needed for sport-related concussion.Methods: This was a multicenter, prospective, case-control study of athletes who provided blood samples and were diagnosed with a concussion or were a matched non-concussed control within the National Collegiate Athletic Association -Department of Defense Concussion Assessment, Research, and Education Consortium conducted between 2015 and 2019. The blood was collected within 48 h of injury to identify protein abnor-malities at the acute and subacute timepoints. Athletes with concussion were divided into 6 h post-injury (0-6 h post-injury) and after 6 h post -injury (7-48 h post-injury) groups. We applied a highly multiplexed proteomic technique that used a DNA aptamers assay to target 1305 proteins in plasma samples from athletes with and without sport-related concussion.Results: A total of 140 athletes with concussion (79.3% males; aged 18.71 +/- 1.10 years, mean +/- SD) and 21 non-concussed athletes (76.2% males; 19.14 +/- 1.10 years) were included in this study. We identified 338 plasma proteins that significantly differed in abundance (319 upregu-lated and 19 downregulated) in concussed athletes compared to non-concussed athletes. The top 20 most differentially abundant proteins discriminated concussed athletes from non-concussed athletes with an area under the curve (AUC) of 0.954 (95% confidence interval: 0.922- 0.986). Specifically, after 6 h of injury, the individual AUC of plasma erythrocyte membrane protein band 4.1 (EPB41) and alpha-synuclein (SNCA) were 0.956 and 0.875, respectively. The combination of EPB41 and SNCA provided the best AUC (1.000), which suggests this combi-nation of candidate plasma biomarkers is the best for diagnosing concussion in athletes after 6 h of injury.Conclusion: Our data suggest that proteomic profiling may provide novel diagnostic protein markers and that a combination of EPB41 and SNCA is the most predictive biomarker of concussion after 6 h of injury.