Circulating nucleosome immunoassay: Evaluating a clinically-applicable test to risk stratify COVID-19 and target anticoagulation

Background : Early therapeutic anticoagulation may decrease thrombotic and non-thrombotic complications in infection with SARSCoV-2 (COVID-19), improving outcomes. Accurately targeting only high risk patients would improve safety of this approach;bloodbased biomarkers to accurately risk-stratify would be of value. Dysregulated neutrophil extracellular traps (NETs) drive pulmonary inflammation, thrombosis and mortality and may therefore represent a useful biomarker. NETs determined via free DNA quantification are elevated in severe COVID-19 and correlate with outcome, but the technical requirements of these tests preclude their clinical use. Aims : We assessed the potential of a clinically-applicable immunoassay for the quantification of cell free H3.1-nucleosome as a NETs biomarker and predictor of thrombosis and mortality. Methods : Plasma samples on admission, day 3, 7 and 10 were evaluated from 20 patients with severe COVID-19 requiring organ support (severe cohort) and compared with 28 samples from COVID-19 patients requiring hospitalization, but not organ support (non-severe cohort). Circulating H3.1-nucleosomes were measured using Nu.Q™ H3.1-nucleosome ELISA (Belgian Volition SRL, Isnes, Belgium) as per manufacturer's instruction. Results : Patient demographics: Severe cohort: Age (median/ range) = 63 (43-84). Gender distribution: F = 5, M = 15. Non-severe cohort: Age = 51.5 (28-80). F = 11, M = 17. H3.1 nucleosome levels were significantly elevated in the severe versus non-severe cohort (Figure 1a). H3.1 nucleosome levels could not be used to predict thrombotic outcome, but there was an association with 28 day mortality, with significantly higher admission values recorded in ITU patients who died (Mann-whitney P = 0.014, n = 6) and comparatively higher H3.1-nucleosome values maintained during day 1-7 of ITU admission (Figure 1b). Conclusions : The immunoassay NETs biomarker replicated findings of free DNA quantification studies;NETs correlate with disease severity. Although of no value in predicting thrombosis in this study, there is an indication that elevated values predict poor outcomes in patient admitted to ITU and may be of value in risk stratifying to treatments such as therapeutic anticoagulation and tracking response to treatment.