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Background: Heparanase is implicated in angiogenesis and tumor progression. We had earlier demonstrated that heparanase may also affect the hemostatic system in a non-enzymatic manner. It forms a complex and enhances the activity of the blood coagulation initiator- tissue factor (TF). Although increased heparanase antigen level in the plasma and biopsies of cancer patients was previously demonstrated, in the present study we evaluated, for the first time, the heparanase procoagulant activity in the plasma of patients with lung cancer., Materials and Methods: Sixty five patients with non-small cell lung cancer at presentation and twenty controls were recruited. Plasma was studied for TF / heparanase procoagulant activity, TF activity and heparanase procoagulant activity using chromogenic assay and heparanase antigen levels by ELISA., Results: Heparanase antigen levels were higher in the study group compared to control (P=0.05). TF / heparanase activity, and even more apparent, heparanase procoagulant activity were significantly higher in the study group compared to controls (P=0.008, P<0.0001, respectively). No significant difference was observed in the TF activity between the groups. Survival of patients with heparanase procoagulant activity higher than 31 ng/ml predicted a mean survival of 9 ± 1.3 months while heparanase procoagulant activity of 31 ng/ml or lower predicted a mean survival of 24 ± 4 months (P=0.001). Heparanase procoagulant activity was higher than 31 ng/ml in the four cases of thrombosis detected during the follow-up period., Conclusions: Elevated heparanase procoagulant activity in patients with lung cancer reveals a new mechanism of coagulation system activation in malignancy. Heparanase procoagulant activity can potentially be used as a predictor for survival., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Background: Heparanase that was cloned from and is abundant in the placenta is implicated in cell invasion, tumor metastasis and angiogenesis. Recently, we demonstrated that heparanase is directly involved in the regulation of the hemostatic system. Heparanase was shown to form a complex and enhance tissue factor (TF) activity, resulting in increased factor Xa production (Nadir et al. Haematologica, 2010). The present work suggests a novel assay to evaluate heparanase procoagulant activity., Methods: Heparanase procoagulant activity was studied using purified proteins of heparanase, TF, factor VIIa and factor X. The assay was verified in 55 plasma samples and compared to heparanase and tissue factor pathway inhibitor (TFPI) levels by ELISA and factor Xa, thrombin levels and antithrombin activity by chromogenic substrates. Thirty five samples were of third-trimester pregnant women (weeks 36-41) who were in labor or came for appointed elective cesarean section and 20 control samples were of non-pregnant healthy women., Results: Heparanase procoagulant activity assay was shown to differentiate heparanase procoagulant effect from TF activity, in purified proteins. Heparanase procoagulant activity was significantly higher in the plasma of pregnant women compared to non-pregnant (p < 0.005). Heparanase relative contribution to the TF / heparanase complex activity was significantly higher in the plasma of pregnant women compared to non-pregnant (29% increase, p < 0.0001). Differences in heparanase procoagulant activity were more prominent than changes in heparanase levels by ELISA, TF activity, factor Xa, thrombin and free TFPI levels., Conclusions: Heparanase procoagulant activity can be determined by the suggested assay. The assay revealed a significant contribution of heparanase to the procoagulant state in late third-trimester pregnancy and at delivery., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: Heparanase is an endo-β-D-glucuronidase dominantly involved in tumor metastasis and angiogenesis. Recently, we demonstrated that heparanase is involved in the regulation of the hemostatic system. Our hypothesis was that heparanase is directly involved in activation of the coagulation cascade., Design and Methods: Activated factor X and thrombin were studied using chromogenic assays, immunoblotting and thromboelastography. Heparanase levels were measured by enzyme-linked immunosorbent assay. A potential direct interaction between tissue factor and heparanase was studied by co-immunoprecipitation and far-western assays., Results: Interestingly, addition of heparanase to tissue factor and activated factor VII resulted in a 3- to 4-fold increase in activation of the coagulation cascade as shown by increased activated factor X and thrombin production. Culture medium of human embryonic kidney 293 cells over-expressing heparanase and its derivatives increased activated factor X levels in a non-enzymatic manner. When heparanase was added to pooled normal plasma, a 7- to 8-fold increase in activated factor X level was observed. Subsequently, we searched for clinical data supporting this newly identified role of heparanase. Plasma samples from 35 patients with acute leukemia at presentation and 20 healthy donors were studied for heparanase and activated factor X levels. A strong positive correlation was found between plasma heparanase and activated factor X levels (r=0.735, P=0.001). Unfractionated heparin and an inhibitor of activated factor X abolished the effect of heparanase, while tissue factor pathway inhibitor and tissue factor pathway inhibitor-2 only attenuated the procoagulant effect. Using co-immunoprecipitation and far-western analyses it was shown that heparanase interacts directly with tissue factor., Conclusions: Overall, our results support the notion that heparanase is a potential modulator of blood hemostasis, and suggest a novel mechanism by which heparanase increases the generation of activated factor X in the presence of tissue factor and activated factor VII.