Relative Neutralization of Heparin from Different Origins by Protamine, Polybrene, Platelet Factor 4, and synthetic heparin antagonist PMX

Abstract

IntroductionHeparin is the most widely used anticoagulant, which is derived from mammalian tissues. While porcine mucosal heparin is most commonly used clinically, sheep mucosal heparin and bovine mucosal heparin are also currently developed. Protamine Sulfate is the most commonly used antagonist of heparin, which is used clinically. Platelet Factor 4 is a platelet‐derived protein which is capable of neutralizing heparin endogenously. Polybrene and PMX are synthetic agents which can neutralize heparin. These agents are not used clinically. The purpose of this study is to compare the neutralization profile of various heparins by protamine and other heparin‐neutralizing agents.Materials and methodPorcine heparin was obtained from Medefill Inc. (Glendale Heights, IL). Bovine Heparins were obtained from Extrasul (Sao Paolo, Brazil). Ovine Heparin was obtained from Ronssi Co. (Souzhou, China). Protamine sulfate was obtained from Sanofi Avantis Co. (Paris, France). Platelet Factor 4 was obtained from Enzyme Research (South Bend, IN). Polybrene and PMX were of synthetic origin. Pool‐citrated plasma was obtained from Loyola University Medical Center Blood Bank. Blood‐clotting tests were performed using aPPT and TT reagents. Chromogenic substrate assays were carried out on the ACL‐Elite instrument. Plasma samples were supplemented with each of the pooled heparin preparations at a concentration of 0–10 mg/ml. Protamine neutralization studies were carried out at a fixed concentration of 10 mg/ml. Neutralization profiles were determined graphically and by calculating percent neutralization at a fixed concentration of 5 mg/ml.ResultsAt 5 ug/ml the protamine, the neutralization index for the different heparins were comparable in the TT, anti‐Xa and anti‐IIa assays (> 90%) however in the APTT it was around 75% for porcine and bovine pool and 66% for ovine pool. In the polybrene neutralization the all heparins comparable neutralization index in the APTT (>80%), TT (>90%) and anti‐IIa (>90%) however in the anti‐Xa the porcine and bovine were greater than 80% and bovine pool was greater than 90%.ConclusionsProtamine sulfate was found to be the most effective neutralizing agent for the bovine, ovine and porcine heparins. Assay based variations were noted in the neutralization profiles. With polybrene and platelet factor 4. These studies suggest that titration with the heparin neutralizing agent may be useful in demonstration the biosimilarities of heparins obtained from different sources.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.