Interrogation of the Coagulation Cascade Acute Coronary Syndrome Using Novel Elisa-Based Assays for Single Protease Quantification

Background: Acute coronary syndrome (ACS), the main contributor to myocardial infarction, is a thrombotic complication of atherosclerosis. Growing evidence supports a role for the intrinsic coagulation cascade in various thrombotic diseases. To gain more inside in the contribution of the intrinsic coagulation cascade in an ACS, we developed novel, ELISA-based assays for the quantification of Kallikrein, activated factor XII (FXIIa), FXIa, FXa, FIXa, and thrombin in complex with a physiological inhibitor.

Methods: During this prospective cohort study, blood from patients with a first ACS (n=61) was collected upon admission (before heparin injection(t=0), and after 1(t=1) and 6 (t=2) months. Plasma levels of Kallikrein-C1inhibitor (K-C1inh), FXIIa-C1inh, FXIa-C1inh, FXIa-α1 anti trypsin, FXIa-antithrombin (AT), FIXa-AT, FXa-AT, and thrombin-AT(TAT) were measured using newly developed ELISA's and compared to apparently healthy controls (n=60). Methods were validated according to the EP5-protocol for within- and -between run variability.

Results: The inter coefficients of variability for K-C1inh is 11.9%, FXIa-α1-AT 14.4%, FXIa-AT 13.7%, FIXa-AT 4.9%, FXa-AT 2.6% and TAT 8.6 %. Levels of FXIa-α1-AT, FXI- ΑΤ, FIXa-AT and TAT were all significantly elevated in plasma from ACS patients at time of the acute event compared to healthy control: FXIa-α1 anti trypsin respectively 202.8 (IQR 156.6 - 283.4) vs 29.0 pM (IQR 0.00 - 101.0); FXIa-AT levels were respectively 35.9 pM (IQR 23.9 - 43.4) vs 28.0 pM (IQR 23.3 - 34.0); FIXa-AT respectively 102.0pM (IQR 87.4 - 123.0) vs 90.5 pM (IQR 83.2 - 104.3); TAT respectively 5.59 (IQR 3.91 - 9.85) vs 2.26 pM (IQR 1.78 - 2.71)(Fig1). Plasma levels of FXIa-α1-AT, FXIa-AT FIXa-AT, and TAT decreased during follow up. Additionally, FXIa-α1-AT, FXIa-AT FIXa-AT, and TAT were higher in patients with ST-elevated myocardial infarction (STEMI) compared to subjects with non-STEMI or angina pectoris.

Conclusion: Elisa based profiling of the coagulation proteases provide a highly sensitive and reproducible method used to distinguish the different activation states of single proteases in the coagulation cascade. With these novel assays we show that FXIa-α1-AT, FXIa-AT FIXa-AT, and TAT are significantly elevated in patients with ACS. Additionally, patients with STEMI have a more pronounced hypercoagulable state than non-STEMI and angina pectoris patients.

Figure 1.

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Figure 1.

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