Aptamer-modified FXa generation assays to investigate hypercoagulability in plasma from patients with ischemic heart disease

High plasma levels of activated Factor VII-Antithrombin complex (FVIIa-AT) have been associated with an increased risk of cardiovascular mortality in patients with stable coronary artery disease (CAD). To investigate if FVIIa-AT levels are associated with activated factor X generation (FXaG) in modified assays. Forty CAD patients were characterized for FVIIa-AT levels by ELISA and for FXaG in plasma. Novel fluorogenic FXaG assays, based on aptamers inhibiting thrombin and/or tissue factor pathway inhibitor (TFPI), were set up. FXaG correlated with FVIIa-AT levels (RAUC = 0.393, P = 0.012). The combination of thrombin inhibition and FXaG potentiation by using anti-thrombin and anti-TFPI aptamers, respectively, favors the study of time parameters. The progressive decrease in lag time from the lowest to the highest FVIIa-AT quartile was magnified by combining TFPI and thrombin inhibitory aptamers, thus supporting increased FXaG activity in the coagulation initiation phase. By exploring FXaG rates across FVIIa-AT quartiles, the largest relative differences were detectable at the early times (the highest versus the lowest quartile; 5.0-fold, P = 0.005 at 45 s; 3.5-fold, P = 0.001 at 55 s), and progressively decreased over time (2.3-fold, P = 0.002 at 75 s; 1.8-fold, P = 0.008 at 95 s; 1.6-fold, P = 0.022 at 115 s). Association between high FVIIa-AT levels and increased FXaG was independent of F7 −323 A1/A2 polymorphism influencing FVIIa-AT levels. High FVIIa-AT plasma levels were associated with increased FXaG. Hypercoagulability features were specifically detectable in the coagulation initiation phase, which may have implications for cardiovascular risk prediction by either FVIIa-AT complex measurement or modified FXaG assays. [Display omitted] •High FVIIa-AT levels were associated with increased FXa generation in CAD patients.•FXa generation was modified by aptamer inhibition of TFPI and Thrombin.•Early time FXaG rates increased from the lowest to the highest FVIIa-AT quartile.•This relation was still detectable in patients grouped for F7 genotypes.•Hypercoagulability features were detectable in the coagulation initiation phase.