Studies on the Interaction of Unfractionated Heparin and Sulodexide with Functional Antiheparin Platelet Factor 4 Antibodies as Studied in Platelet Aggregation Assays

Abstract

IntroductionSulodexide represents a glycosaminoglycan derived antithrombotic agent which is clinically used for the management of vascular disorders and is pharmacologically active after both parenteral and oral administration. The purpose of this study is to compare the relative effect of sulodexide in mediating heparin induced thrombocytopenia (HIT) antibodies induced aggregation of human platelets in a standardized test system which is developed to compare the effects of heparins and related drugs.Materials and MethodsActive pharmaceutical ingredients (API versions of sulodexide), fast moving heparin and dermatan sulfate were obtained from Alfa Sigma (Bologna, Italy). API versions of UFH were obtained from Medefil, Inc (Glendale Heights, IL, USA). All agents were dissolved in saline at a stock concentration of 1 mg/ml. Working dilutions were prepared at 100, 10 and 1 mg/ml. The source of functional HIT antibody represented a frozen pool of aphresis fluid (AF) collected from five clinically confirmed heparin induced thrombocytopenia patients. The functionality of the HIT antibodies in this pool were confirmed in the platelet aggregation and 14C serotonin release assay. All results were compiled in terms of percent aggregation of platelets and represented as mean ± standard deviation.ResultsUFH produced a concentration dependent aggregation from 0 – 10 ug/ml reaching a peak aggregation of 60 ± 18.76%. Sulodexide produced a relatively weaker aggregatory response and produced mild aggregation in the same concentration range exhibiting comparable platelet aggregation at 10 ug/ml. The fast moving heparin component of sulodexide produced a concentration dependent response comparable to heparin reaching a maximum aggregation of 66 ± 30.67% at 10 ug/ml. The dermatan sulfate component did not produce any significant aggregation of platelets and no concentration dependence was observed (<10% aggregation). Wide intra variations were observed in the HIT antibody aggregation response with both the heparin and sulodexide but not with dermatan sulfate.DiscussionThese results clearly show that in comparison to heparin, sulodexide produces a relatively weaker response in the functional HIT antibody screening studies. However, the fast moving heparin component of sulodexide produces comparable responses to UFH whereas dermatan sulfate has much weaker aggregation response. The presence of dermatan sulfate may modulate the interaction between platelet factor 4 and sulfated polysaccharide change present in sulodexide preparations.