PC Deficiency Testing: Thrombin-Thrombomodulin as PC Activator and Aptamer-Based Enzyme Capturing Increase Diagnostic Accuracy

Abstract

Protein C (PC) activity tests are routinely performed in a thrombophilia workup to screen for PC deficiency. Currently used tests combine conversion of PC to activated PC (APC) by the snake venom Protac with subsequent APC detection through hydrolysis of a chromogenic peptide substrate or prolongation of a clotting time. In this prospective cohort study, we analyzed how different modes of PC activation and subsequent APC determination influence the diagnostic accuracy of PC activity testing in a cohort of 31 patients with genetically confirmed PC deficiency. In addition to chromogenic and clot-based measurement, an oligonucleotide-based enzyme capture assay utilizing a basic exosite-targeting aptamer was used for APC detection. To study the influence of the PC activation step on diagnostic sensitivity, PC activation through Protac and through the thrombin-thrombomodulin (TM) complex were compared. Twenty-six (84%) and 24 (77%) PC deficient patients were identified as true-positive using the chromogenic and the clot-based PC activity assay, respectively. True-positive results increased to 27 (87%) when the basic exosite-targeting aptamer approach was used for APC measurement. Additional replacement of the PC activator Protac by thrombin-TM gave true-positive results in all patients. These data indicate that the mode of PC activation is crucial in determining the accuracy of PC activity testing and that diagnostic sensitivity can be significantly improved by replacing the PC activator Protac with thrombin-TM. APC detection using a basic exosite-targeting aptamer achieves high sensitivity toward mutations outside the active center while being less subject to interfering factors than clot-based PC activity assays.

Effect of hemocoagulase on the coagulation parameters, blood transfusion volume and bleeding volume of patients with severe traumatic fractures

“…Hemocoagulase plays both "thrombin-like" and "thromboplastin-like" roles in clinical practice [24,25]. Hemocoagulase acts similarly to human thrombin in that it causes platelet aggregation and white thrombus formation at the bleeding site without affecting the patient's other healthy sites [26,27]. On the other hand, hemocoagulase can also act as thromboplastin in the degradation of fibrin to fibrinopeptide A, which forms soluble fibrin I with fibrin to achieve coagulation in patients with injury-induced bleeding [28,29].…”

Compound heterozygous protein C deficiency with pulmonary embolism caused by a novel PROC gene mutation: Case report and literature review

“…[ 3 ] PC is a vitamin K-dependent plasma zymogen that is synthesized mainly in the liver and plays an important roles in anticoagulation and fibrinolysis by inactivating the blood coagulation factors Va and VIIIa. [ 4 ] Protein C is encoded by the Protein C, Inactivator Of Coagulation Factors Va And VIIIa (PROC) gene on chromosomes 2q13–q14 and is composed of 9 exons and 8 introns. [ 5 ] Several studies have reported the spectrum of PROC mutation, and at least 300 mutations have been reported to date.…”