Benny Sørensen

Fibrinogen concentrate substitution therapy in patients with massive haemorrhage and low plasma fibrinogen concentrations

BACKGROUNDnPatients experiencing massive haemorrhage are at high risk of developing coagulopathy through loss, consumption, and dilution of coagulation factors and platelets. It has been reported that plasma fibrinogen concentrations may reach a critical low level relatively early during bleeding, calling for replacement fibrinogen therapy. Cryoprecipitate has been widely used in the past, but more recently, a pasteurized fibrinogen concentrate has become available. We audited the effects of fibrinogen concentrate therapy on laboratory and clinical outcome in patients with massive haemorrhage.nnnMETHODSnWe identified 43 patients over the previous 2 yr to whom a fibrinogen concentrate had been administered as treatment for hypofibrinogenaemia during serious haemorrhage. Platelet count, P-fibrinogen, activated partial thromboplastin time (APTT), prothrombin time (PT), D-dimer, and volume of blood lost were obtained from medical and laboratory records. Numbers of units of red blood cells (RBC), fresh frozen plasma (FFP), and pooled platelet concentrates were recorded before and after fibrinogen substitution.nnnRESULTSnA significant increase in plasma fibrinogen concentration was observed after fibrinogen concentrate therapy. Platelet counts and fibrin D-dimer values remained unchanged, whereas the APTT and PT improved significantly. Requirements for RBC, FFP, and platelets were significantly reduced. Blood loss decreased significantly.nnnCONCLUSIONSnOff-label substitution therapy with a fibrinogen concentrate generally improved global laboratory coagulation results and as supplementary intervention, appeared to diminish the requirements for RBC, FFP, and platelet substitution in this patient cohort.

Fibrinogen substitution improves whole blood clot firmness after dilution with hydroxyethyl starch in bleeding patients undergoing radical cystectomy: a randomized, placebo- controlled clinical trial

Summary.u2002 Background:u2002Infusion of artificial colloids such as hydroxyethyl starch (HES) induces coagulopathy to a greater extent than simple dilution. Several studies have suggested that the coagulopathy could be corrected by substitution with a fibrinogen concentrate. Objectives:u2002The aims of the present prospective, randomized, placebo‐controlled trial were to investigate the hemostatic effect of a fibrinogen concentrate after coagulopathy induced by hydroxyethyl starch in patients experiencing sudden excessive bleeding during elective cystectomy. Methods:u2002Twenty patients were included. Blood loss was substituted 1:1 with HES 130/0.4. At a dilution level of 30%, patients were randomly selected for intra‐operative administration of a fibrinogen concentrate or placebo. The primary endpoint was maximum clot firmness (MCF), as assessed by thromboelastometry. Secondary endpoints were blood loss and transfusion requirements, other thromboelastometry parameters, thrombin generation and platelet function. Results:u2002Whole‐blood MCF was significantly reduced after 30% dilution in vivo with HES. The placebo resulted in a further decline of the MCF, whereas randomized administration of fibrinogen significantly increased the MCF. Furthermore, only 2 out of 10 patients randomly chosen to receive fibrinogen substitution required postoperative red blood cell transfusions, compared with 8 out of 10 in the placebo group (Pu2003=u20030.023). Platelet function and thrombin generation were reduced after 30% hemodilution in vivo, and fibrinogen administration caused no significant changes. Conclusions:u2002During cystectomy, fluid resuscitation with HES 130/0.4 during sudden excessive bleeding induces coagulopathy that shows reduced whole‐blood maximum clot firmness. Randomized administration of fibrinogen concentrate significantly improved maximum clot firmness and reduced the requirement for postoperative transfusion.

Mechanisms of hydroxyethyl starch‐induced dilutional coagulopathy

Summary.u2002 Background: The biochemical mechanisms causing dilutional coagulopathy following infusion of hydroxyethyl starch 130/0.4 (HES) are not known in detail. Objectives: To give a detailed biochemical description of the mechanism of coagulopathy following 30%in vivo dilution with HES, to present a systematic ex vivo test of various hemostatic agents, and to investigate the hypothesis that acquired fibrinogen deficiency constitutes the most important determinant of the coagulopathy. Methods: Dynamic whole blood clot formation assessed by thromboelastometry, platelet count, thrombin generation, and the activities of von Willebrand factor, coagulation factor II, FVII, FVIII, FIX, FX and FXIII were measured in 20 bleeding patients enrolled in a prospective clinical study investigating in vivo substitution of blood loss with HES up to a target level of 30%. Thromboelastometry parameters were further evaluated after ex vivo spiking experiments with fibrinogen, prothrombin complex concentrate (PCC), FXIII, activated recombinant FVIIa (rFVIIa), fresh frozen plasma, and platelets. Results: Hemodilution reduced maximum clot firmness (MCF), whereas whole blood clotting time (CT) and maximum velocity remained unaffected. All coagulation factor activities were reduced. Fibrinogen, FII, FXIII and FX activities decreased significantly below the levels expected from dilution. The endogenous thrombin potential was unchanged. Ex vivo addition of fibrinogen normalized the reduced MCF and increased the maximum velocity, whereas PCC, rFVIIa and platelets shortened the CT but showed no effect on the reduced MCF. Conclusions: Acquired fibrinogen deficiency seems to be the leading determinant in dilutional coagulopathy, and ex vivo addition corrected the coagulopathy completely.

Fibrinogen concentrate – a potential universal hemostatic agent

Background: Human fibrinogen concentrates have been commercially available for decades for substitution therapy in hypofibrinogenemia, dysfibrinogenemia and afibrinogenemia. Accumulating new data suggest that fibrinogen plays a critical role in achieving and maintaining hemostasis, particularly in patients suffering from acquired fibrinogen deficiency during massive bleeding, where benefit from early intervention with fibrinogen concentrate appears to be important. Objective/methods: This work focuses on pasteurized fibrinogen concentrate, with special emphases on product characteristics, pharmacodynamics, pharmacokinetics, laboratory monitoring, dosing, clinical efficacy, safety and tolerability. Future clinical and laboratory perspectives on fibrinogen are discussed and outlined. Results/conclusion: Pasteurized fibrinogen concentrate is derived from human plasma. Its half life is 2.7 days in patients with congenital fibrinogen deficiency. For congenital and acquired deficiency in vivo recovery rates vary from 60% to 109%. Reportedly, administration of pasteurized fibrinogen in patients with congenital deficiency is efficacious. Acquired deficiency of fibrinogen appears to be an early event in seriously bleeding patients, preceding critical levels of platelets or other coagulation factors. Experimental animal studies, as well as clinical observations suggest a beneficial role of early substitution with fibrinogen in management of critical traumatic and surgical bleeds. Pasteurized fibrinogen concentrate is well tolerated and associated with a low incidence of adverse thrombo-embolic events.

Parallel use of by-passing agents in haemophilia with inhibitors: a critical review

In the absence of new outbreaks of transfusion‐related infections, the occurrence of neutralizing antibodies currently remains the most prominent complication in haemophilia. Coagulation factor products that may circumvent the inadequate activation of factor X in classical haemophilia, often referred to as bypassing agents, have demonstrated a high degree of efficacy. A smaller number of patients have been described in whom either bypassing agent, or both, demonstrate diminished efficacy. In those cases, the use of both bypassing agents in parallel was attempted, either using simultaneous (combined) or alternating (sequential) infusion of the two drugs, reportedly with successful haemostasis. We speculated whether such treatment might cause thromboembolism. A thorough literature search disclosed 17 reports regarding the parallel use of bypassing agents in the same bleeding episode in 49 patients; reporting nine patients with acquired haemophilia and forty patients with congenital haemophilia with inhibitors. Notable incidences of thromboembolic manifestations were observed: in nine patients with acquired haemophilia, five and in 40 patients with congenital haemophilia five suffered from significant thrombotic complications, and overall four cases were fatal. Although efficacy of parallel treatment was reported excellent in most cases, thromboembolism is rare in haemophilia and parallel treatment with activated prothrombin complex concentrate and activated recombinant human factor VII appears to increase the risk of thrombosis in these patients.

The combination of recombinant factor VIIa and fibrinogen correct clotting ex vivo in patient samples obtained following cardiopulmonary bypass surgery.

Cardiac surgery involving cardio pulmonary bypass (CPB) may be associated with development of a coagulopathy that increases risk of bleeding. In the present ex vivo study we investigated the influence of fibrinogen and rFVIIa, alone or in combination, on whole blood coagulation thromboelastometry using pre- and postoperative blood samples from 18 consecutive adult patients undergoing CPB surgery. Dynamic thromboelastometric clotting profiles were recorded using citrated whole blood activated with trace amounts of tissue factor (Innovin, final dilution 1:17000). Blood samples were collected before surgery (control) and postoperative samples were obtained following in vivo neutralization of heparin with protamine sulphate. All blood samples were treated with heparinase to ensure neutralization of possible residual heparin effect. The post-operative blood samples were spiked with buffer, rFVIIa (2 microg/mL), fibrinogen (1 mg/mL), or the combination of rFVIIa and fibrinogen. Despite neutralization of heparin, CPB surgery left a measurable coagulopathy that was thromboelastometrically characterized by prolonged onset of clotting, reduced maximum velocity of clot formation (MaxVel), and decreased maximum clot firmness (MCF). Ex vivo spiking of the postoperative samples with rFVIIa shortened the clotting time. Fibrinogen also shortened the clotting time and, in addition, improved the MaxVel, and MCF. Finally, adding the combination of rFVIIa and fibrinogen to the postoperative samples corrected all thromboelastometric parameters to the preoperative range. In conclusion, the correction of whole blood clotting abnormalities that occurs with rFVIIa and/or fibrinogen suggests that future clinical trials on treatment of bleeding during CPB surgery should study the haemostatic effect of fibrinogen or possibly the combination of rFVIIa and fibrinogen.

Whole blood coagulation in children with thrombocytopenia and the response to platelet replacement, recombinant factor VIIa, and a potent factor VIIa analogue

The present study evaluated dynamic coagulation profiles, platelet aggregation, and thrombin generation in whole blood (WB) from eight children with thrombocytopenia during chemotherapy, and the haemostatic potential of platelets (+60u2003×u2003109/l), recombinant factor VIIa (rFVIIa – NovoSeven®), and a potent rFVIIa analogue (NN1731) both at 1 and 4u2003μg/ml. Dynamic WB coagulation profiles were recorded by thrombelastometry employing activation with tissue factor (TF – Innovin®) at low concentrations. The baseline WB coagulation patterns were characterised by a prolonged clotting time (CT) and a pronounced reduction in clot propagation (MaxVel). WB platelet aggregation signal was five times lower in the study group compared with measurements in modelled thrombocytopenic WB from healthy volunteers. In vitro addition of fresh platelets reversed the coagulopathy. Addition of rFVIIa induced no significant changes in the thrombelastographic profile, whereas spiking with NN1731 shortened the CT significantly. The changes in WB thrombin generation reflected the changes in the MaxVel. In modeled thrombocytopenic WB from healthy individuals, both rFVIIa and NN1731 exhibited a pronounced haemostatic effect with NN1731 showing greater potency than rFVIIa. Compromised platelet function in the study group was assumingly responsible for the weakened haemostatic potential of rFVIIa as well as that of NN1731.

Recombinant factor VIIa and fibrinogen display additive effect during in vitro haemodilution with crystalloids

Background: Major blood loss requires fluid resuscitation for maintaining hemodynamic stability. Excessive volume infusions predispose to dilutional coagulopathy through loss, consumption and dilution of cells and proteins involved in haemostasis. Further treatment with fibrinogen concentrate and/or recombinant activated factor VII (rFVIIa) may be initiated, although the haemostatic effects in a situation with haemodilution are not fully detailed. The present study evaluates haemostatic effect of fibrinogen and rFVIIa and their combination in an in vitro model of haemodiluted whole blood with two commonly used crystalloids.

Reduced clot-stability during the first 6 hours after aneurysmal subarachnoid haemorrhage - a prospective case-control study

INTRODUCTIONnEarly rebleeding is an important cause of death and disability following aneurysmal subarachnoid haemorrhage (SAH). Recent studies have shown that 50-90% of the rebleedings occurred within the first 6 hours after the primary bleeding. The mechanism leading to rebleeding remains to be established. In the present prospective case-control study we hypothesize that patients with SAH develop a coagulopathy characterized by reduced clot stability during the early period after the initial bleeding.nnnMETHODSnPatients with aneurysmal SAH was studied with a dynamic clot lysis assay and markers of fibrinolysis and clot stabilizers in blood samples taken within and after 6 hours after onset of bleeding. Results were compared with blood samples from age and gender matched healthy controls.nnnRESULTSn36 patients were enrolled, 26 patients had blood samples collected within 6 hours after the initial bleeding whereas 10 patients had blood samples taken later than 6 hours after the initial bleeding. Patients demonstrated significantly reduced clot stability during the first 6 hours after initial bleeding. Fibrinolytic activity was increased during the first 6 hours along with the inhibitors of fibrinolysis whereas the modulators of fibrinolysis were reduced or inactivated.nnnCONCLUSIONnDuring the first 6 hours after SAH patients exhibit reduced clot-stability. Probably a consequence of activated fibrinolysis in combination with reduced or inactivated factor XIII and thrombin-activable fibrinolysis inhibitor.

Artificial contact pathway activation masks the haemostatic potential of rFVIIa and NN1731 in thrombocytopenic whole blood

The authors would like to thank Prof. Roger Pain for critical reading of the manuscript. We would also like to thank Bojana Mirkovic, M.Pharm. (Faculty of Pharmacy, University of Ljubljana) for the assistance with kinetic analysis. Surface plasmon resonance experiments were performed in the Infrastructural Centre for Surface Plasmon Resonance at the Department of Biology, University of Ljubljana. This work was supported by a grant from the Research Agency of the Republic of Slovenia (grant P4-0127 to JK) and partially by the Seventh EU Framework IP project NanoPhoto (JK).