EP2403521A1

EP2403521A1 - Novel uses of fibrinogen

Info

Publication number: EP2403521A1
Application number: EP10715301A
Authority: EP
Inventors: Caroline Teboul; Bruno Padrazzi
Original assignee: LFB SA
Current assignee: LFB SA
Priority date: 2009-03-06
Filing date: 2010-03-05
Publication date: 2012-01-11
Also published as: FR2942718A1; US20110319331A1; BRPI1010251A2; CA2754399A1; CN102369020A; JP2012519674A; AU2010219520A1; IL214958A0; KR20120022717A; WO2010100385A1

Abstract

The invention relates to the use of fibrinogen for producing a drug to be used in the prevention or treatment of severe acute haemorrhage, wherein said drug is to be delivered in an amount of at least 4.5g of fibrinogen per single dose.

Description

TITLE OF THE INVENTION

New uses of fibrinogen

FIELD OF THE INVENTION The present invention relates to the field of treatment of severe acute hemorrhages, including but not limited to postpartum haemorrhage, severe post-traumatic and surgical (perioperative) acute hemorrhage

PRIOR ART Severe Acute Hemorrhages (SAH) are defined as the rapid loss - in a few hours - of at least 20 to 30% of an individual's blood volume. The main HAS situations are encountered in particular in obstetrics (mainly postpartum haemorrhage), traumatology and surgery, without limiting themselves to these situations.

Coagulation disorders (coagulopathies) are common in HAS and are directly or indirectly related to bleeding. Among the coagulation factors, the plasma fibrinogen level is chronologically the first to fall to reach critical values (Hippala et al., 1995, Anesth Analg, Vol 81: 360-365, Torrielli et al., 1988, Rev Fr Gynecol Obstet, Vol 83: 7-9). These threshold values are conventionally situated between 0.5 and 1 g / l and correspond to the concentrations making it possible to obtain a satisfactory blood coagulation, in a stable non-haemorrhagic situation. The mechanisms involved in lowering the level of fibrinogen are: 1 / direct losses, 2 / dilution by vascular filling solutes, and 3 / fibrinogen consumption for clot formation.

In addition to the fact that fibrinogen is the first coagulation factor whose blood level falls, it has been shown that the fall in fibrinogen level is a factor of poor clinical prognosis, and this decline is associated with a change in patients' the more severe the fibrinogen concentration is low (Charbit et al., 2006, Journal of Thrombosis and Haemostasis, Vol 5: 266-273, Karlsson et al., 2008, Transfusion, Vol 48 (10): 2152- 2158).

The conditions of management of an HAS, in emergency, are adapted to each individual case but include common objectives, including the treatment of coagulation disorders when they exist, which is frequent. Currently, the administration of fibrinogen is recommended when the plasma fibrinogen assay reaches the critical values mentioned above. This surrogate approach aims to correct situations of coagulopathies formed, said coagulopathies often being at the stage of disseminated intravascular coagulation (DIC). This substitutionary approach is late because it is dependent on the time required to obtain the results of repeated laboratory tests, sometimes long to obtain (nearly 1 hour) to diagnose an acquired deficit in fibrinogen and document its evolution during the time. Moreover, according to this substitutionary approach, administration of fibrinogen is repeated over time as it is conditioned to the results of repeated fibrinogen assays. The dose of fibrinogen can be administered in several infusions, each being conditioned by the ratio of the quantized deficit of fibrinogen to the blood fibrinogen threshold value to be reached. Fibrinogen is administered at a recommended slow rate of less than 5ml / minute, which is inappropriate for an emergency.

When the circulating level of fibrinogen is less than 1 g / L, exogenous fibrinogen is administered to allow the formation of a normal clot. The contribution of exogenous fibrinogen varies from 2 to 4 grams, the required intake being calculated on a case-by-case basis and in real time according to the results of successive balances of the coagulation parameters described above. More specifically, the amount of fibrinogen to be injected is conventionally calculated according to the following formula: Amount of exogenous fibrinogen (in grams) = [circulating fibrinogen level to be reached (in g / L)] [circulating fibrinogen level before treatment (in g / L)] x 0.04 x [patient weight (in kg)]. Thus, the amounts of fibrinogen to be administered successively during the course of treatment are calculated according to the balance sheets of the coagulation parameters that are also successive in time. It should be noted that the precise calculation of the quantity of exogenous fibrinogen to be administered is considered essential, in order to avoid an excess of circulating fibrinogen, for example a circulating fibrinogen level greater than 5 g / L, capable of causing thromboses.

We can also mention the administration of a platelet concentrate as a means of additional treatment to the treatment means described above, in particular in situations in which the bleeding persists despite the provision of fibrinogen.

The treatment protocols for postpartum haemorrhage that are currently practiced are generally satisfactory. However, given the significant risk of mortality or morbidity related to these severe acute hemorrhages, there is a constant need in the state of the art for alternative or improved technical solutions compared to existing solutions.

The need to find alternative or improved treatment options for other types of severe acute bleeding, including severe hemorrhages caused by surgical procedures or severe physical trauma. Other severe acute haemorrhages share similarities with postpartum hemorrhages, including homeostatic disorders.

SUMMARY OF THE INVENTION

The present invention relates to the use of fibrinogen for the manufacture of a medicament used in the treatment of severe acute bleeding, said drug providing an early and rapid sufficient amount of fibrinogen to restore the coagulant capacity of blood, independently initial fibrinogen level. By for example, an amount of at least 4.5 g can be administered with a duration of administration of less than 30 minutes.

According to the invention, severe acute hemorrhages include, but are not limited to, postpartum haemorrhage, perioperative haemorrhage and post-traumatic hemorrhage.

The fibrinogen that is administered concerns any fibrinogen, regardless of its origin and its nature.

The invention also relates to pharmaceutical compositions, preventive or curative, comprising fibrinogen, which are specially adapted for the implementation of the uses and methods above.

DESCRIPTION OF THE FIGURES

Figure 1 illustrates the succession of time steps of the in vivo experimental protocol in pigs. The horizontal line represents time. The numbered vertical bars represent the successive steps of the protocol. 1: Stage of anesthesia and instrumentation of the animals. 2: Perform baseline baseline measurements of HR parameters (Heart Rate), MAP (Average Arterial Pressure), PAP (Pulmonary Pulmonary Pressure) Arterial Pressure "), PCWP (Pulmonary Central Wedge Pressure), CVP (central venous pressure), ROTEM®, standard coagulation tests, hemoglobin, hematocrit, platelets. 3: Step of carrying out the hemodilution, during which 60% of the blood volume is taken, which is replaced by a composition of HES 130 / 0.4 (Voluven®), with the aim of achieving a MCF value less than 40 mm. 4: Step of re-transfusion of washed red blood cells, with the objective of achieving a hemoglobin value of 5 to 6 g / dl. 5: Realization of the bone wound step. 7: Realization of HR, MAP, PAP, PCWP, ROTEM® parameters, standard coagulation tests, hemoglobin, hematocrit, platelets and blood loss at 15 minutes, 1 hour, respectively 2 hours and 4 hours after administration of the drug (fibrinogen at different doses and placebo composition). 9: Achievement of the hepatic injury step. 9: End of the experiment, 2 hours after the liver injury, during which the parameters HR, MAP, PAP, CVP, PCWP, ROTEM, standard blood coagulation tests, hemoglobin, hematocrit, platelets and blood loss.

Figure 2 illustrates the results of the INTEM coagulation time (CT) measurement using the ROTEM® method. On the upper part of Figure 2 are shown the results obtained with the animals that were administered respectively: (i) a placebo composition, represented by solid diamonds, (ii) 37.7 mg / kg of human fibrinogen, represented by filled squares, (iii) 75 mg / kg of human fibrinogen, represented solid black triangles and (iv) 150 mg / kg fibrinogen, represented by gray inverted triangles. In the lower part of Figure 3 are shown the results obtained with the animals that were administered respectively: (i) a placebo composition, represented by solid diamonds, (ii) 300 mg / kg of human fibrinogen, represented by squares solid, (iii) 450 mg / kg of human fibrinogen, represented by solid gray triangles and (iv) 600 mg / kg of fibrinogen, represented by gray inverted triangles. On the ordinates: coagulation time (CT), expressed in seconds. On the abscissa, the successive stages of the in vivo experiment, from left to right respectively: (i) after instrumentation and before treatment of the animals, (ii) after hemodilution, (iii) 15 minutes after administration of the drug, (iv) 1 hour after administration of the drug, (v) 2 hours after drug administration, (vi) 4 hours after drug administration and (vii) 2 hours after liver injury or before death.

Figure 3 illustrates the results of the measurement of maximum clot strength (MCF) INTEM using the ROTEM® method. On the upper part of Figure 3 are shown the results obtained with the animals to which were respectively administered: (i) a placebo composition, represented by solid diamonds, (ii) 37.7 mg / kg of human fibrinogen, represented by solid squares, (iii) 75 mg / kg of human fibrinogen, represented by solid black triangles and (iv) 150 mg / kg of fibrinogen, represented by gray inverted triangles. In the lower part of Figure 3 are shown the results obtained with the animals that were administered respectively: (i) a placebo composition, represented by solid diamonds, (ii) 300 mg / kg of human fibrinogen, represented by squares solid, (iii) 450 mg / kg of human fibrinogen, represented by solid gray triangles and (iv) 600 mg / kg of fibrinogen, represented by gray inverted triangles. On the ordinate: maximum clot strength (MCF), expressed in millimeters. On the abscissa, the successive stages of the in vivo experiment, from left to right respectively: (i) after instrumentation and before treatment of the animals, (ii) after hemodilution, (iii) 15 minutes after administration of the drug, (iv) 1 hour after administration of the drug, (v) 2 hours after drug administration, (vi) 4 hours after drug administration and (vii) 2 hours after liver injury or before death. Figure 4 illustrates the results of maximum clot strength (MCF) measurement

PLASMA EXTEM (modified FibTEM) according to the ROTEM® method. The upper part of FIG. 4 shows the results obtained with the animals administered respectively: (i) a placebo composition, represented by solid diamonds, (ii) 37.7 mg / kg of human fibrinogen, represented by solid squares, (iii) 75 mg / kg of human fibrinogen, represented by solid black triangles and (iv) 150 mg / kg of fibrinogen, represented by gray inverted triangles. In the lower part of Figure 4 are shown the results obtained with the animals that were administered respectively: (i) a placebo composition, represented by solid diamonds, (ii) 300 mg / kg of fibrinogen human, represented by solid squares, (iii) 450 mg / kg of human fibrinogen, represented by solid gray triangles and (iv) 600 mg / kg of fibrinogen, represented by gray inverted triangles. On the ordinate: maximum clot strength (MCF), expressed in millimeters. On the abscissa, the successive stages of the in vivo experiment, from left to right respectively: (i) after instrumentation and before treatment of the animals, (ii) after hemodilution, (iii) 15 minutes after administration of the drug, (iv) 1 hour after administration of the drug, (v) 2 hours after drug administration, (vi) 4 hours after drug administration and (vii) 2 hours after liver injury or before death. Figure 5 illustrates the results of measurements of plasma fibrinogen concentration. The curves represent the results obtained with the animals administered respectively: (i) a placebo composition, represented by solid diamonds, on the lower curve of FIG. 5, (ii) 37.5 mg / kg of human fibrinogen, represented by solid squares on the curve immediately above the previous curve, (iii) 75 mg / kg of human fibrinogen, represented by solid black triangles on the curve immediately above the previous curve, (iv) 150 mg / kg of fibrinogen, represented by gray inverted triangles on the curve immediately above the previous curve, (v) 300 mg / kg of human fibrinogen, represented by solid squares on the curve immediately above the previous curve, (vi) 450 mg / ml kg of human fibrinogen, represented by solid gray triangles on the curve immediately above the previous curve, and (vii) 600 mg / kg of fibrinogen, representing are entered by gray triangles on the curve immediately above the previous curve and which is also the upper curve of Figure 5. On the ordinate: plasma concentration of fibrinogen, expressed in mg / dl. On the abscissa, the successive stages of the in vivo experiment, from left to right respectively: (i) after instrumentation and before treatment of the animals, (ii) after hemodilution, (iii) 15 minutes after administration of the drug, (iv) 1 hour after administration of the drug, (v) 2 hours after drug administration, (vi) 4 hours after drug administration and (vii) 2 hours after liver injury or before death. Figure 6 illustrates the results of maximum clot strength (MCF) measurements

INTEM according to the ROTEM® method according to the dose of fibrinogen administered to the animals. On the ordinate: Clot strength maximum value (MCF), expressed in millimeters. On the abscissa, human fibrinogen concentration, expressed in mg / dl.

Figure 7 illustrates the measures of blood loss and coagulation capacity. On the ordinate: blood loss value and clot size value, expressed in ml / kg. In Fig. 7, the respective values of clot size ("closed") and blood loss ("liquid") are represented by adjacent bars, for each of the increasing concentrations of human fibrinogen administered to the animals and for the placebo composition. From the left to the right of Figure 7, each pair of adjacent bars (clot / liquid) represents the respective values for (i) 37.5 mg / kg of fibrinogen, (ii) 75 mg / kg of fibrinogen, (iii) 150 mg / kg fibrinogen, (iv) 300 mg / kg fibrinogen, (v) 450 mg / kg fibrinogen, (vi) 600 mg / kg fibrinogen and (vii) the placebo composition.

DETAILED DESCRIPTION OF THE INVENTION

Surprisingly, it has been shown according to the invention that in the treatment of severe acute haemorrhage, even in the absence of a known deficit in fibrinogen, the early and rapid administration of a single high dose of fibrinogen is able to control severe acute bleeding and prevent its progression to uncontrollable haemorrhage.

According to the invention, a high dose of exogenous fibrinogen, administered in an early and rapid manner, makes it possible to interrupt, in the event of severe acute hemorrhage, the course of a beginning and occult coagulopathy, constituted for the most part by a gradual decrease in the coagulante capacity of the blood, itself caused mainly by the reduction of endogenous fibrinogen, to a coagulopathy that can cause uncontrollable haemorrhage.

The administration of exogenous fibrinogen, under the conditions specified in this invention, is in this way both a treatment of severe acute bleeding and prevention of its adverse evolution to uncontrollable hemorrhage.

According to the invention, a predetermined quantity of exogenous fibrinogen makes it possible to control a severe acute haemorrhage without the need to perform a preliminary measurement of the circulating fibrinogen level. Indeed, the principle of the treatment according to the invention is not to fill a deficit in fibrinogen, that is to say to provide a quantity of fibrinogen from a predetermined blood threshold value as being critical to achieve a presumed effective blood target value, but to restore as quickly as possible the coagulating capacity of the blood.

However, taking into account the fibrinogen dosage to determine the need to administer fibrinogen, and to determine the amount of fibrinogen to be administered, requires that the blood fibrinogen level measurement results are available to the medical team. It should be noted that the results of measuring blood levels of fibrinogen are generally available only after at least 45 minutes to 1 hour after blood sampling, even in emergency situations. This loss of time in the administration of treatment of a therapeutic emergency may result in the value of the patient's blood fibrinogen level, at the moment when the measurement results are available, is less than the above-mentioned threshold value, which may in some cases significantly affect the patient's chances of survival.

However, it has been shown in the invention that the administration of fibrinogen was effective without the need to know beforehand the initial blood level of fibrinogen. So we showed that the treatment of an HAS can be achieved by an early administration of fibrinogen, without prior measurement of blood levels, as soon as a clinical diagnosis of severe acute hemorrhage is made

It has thus been shown according to the invention that the administration of fibrinogen to patients suffering from severe acute hemorrhage can be carried out with a single quantity of fibrinogen, in order to immediately restore the coagulating capacity of the blood, thus without requiring a adaptation of the amount of fibrinogen according to the patient to be treated.

In particular, it has been shown according to the invention that severe acute haemorrhage can be successfully treated by administration of a predetermined amount of exogenous fibrinogen, without the need to perform a prior assay of the circulating fibrinogen level.

Surprisingly, it has been shown according to the invention that the rapid and rapid administration to a patient affected by severe acute hemorrhage, a high single dose of fibrinogen without assaying the level of fibrinogen circulating prior to administration, it makes it possible to improve or prevent the aggravation of irreversible haemostasis disorders that may be experienced by said patient, without causing thrombosis.

In addition, it has also been shown according to the invention that the treatment of severe acute hemorrhage can be achieved by early and rapid administration of a single large dose of fibrinogen, an amount of at least 4.5 g , that is to say without requiring the realization of multiple sequential administrations of exogenous fibrinogen during the entire period of treatment and / or monitoring of the patient, related to the adaptation of the amount of fibrinogen as a function of fibrinogen levels.

The examples illustrate a study in animals administered a range of fibrinogen doses ranging from 37.5 mg / kg to 600 mg / kg, according to a model of hemodilution caused by a traumatic shock with a loss of 60 % of the blood volume, which is then replaced by a 6% solution of hydroxyethyl starch (HES). The results show that coagulation time and maximum clot firmness are significantly affected by hemodilution. The administration of fibrinogen induces an increase in all measures of clot firmness, in a dose-dependent manner. The results of the examples show that a treatment of the animals having undergone a haemorrhage with a dose of fibrinogen of 50 mg / kg completely restores the maximum values of clot firmness 15 minutes after the end of the infusion of fibrinogen, this effect being maintained until the end of the experiment. With larger doses of fibrinogen, a plateau is reached for the EXTEM MCF values (ROTEM® test). In contrast, with increasing doses of fibrinogen, the values of INTEM MCF (ROTEM® test: platelet independent coagulation) continue to increase, as shown in Figure 6. The results of the examples show that thrombin production in the animals administered fibrinogen did not differ from the thrombin production in the control group of animals. A statistical analysis of blood loss shows a dose effect significant response (p = 0.02), indicating a reduction in total blood loss with increasing doses of fibrinogen up to 400 mg / kg, a plateau being achieved for a fibrinogen dose of 600 mg / kg, as is The results of the examples show that the concentration of fibrinogen obtained in the animals that received 300 mg / kg of fibrinogen was close to the base value obtained in control animals without hemorrhage. The concentration of fibrinogen in animals treated with 400 mg / kg of fibrinogen was even greater than the same group of control animals.

Importantly, it has been shown in the examples that even with high doses of fibrinogen, no evidence of hypercoagulation has been observed after analysis of biological or histological parameters.

The results of the examples confirm that clot firmness is affected by low levels of fibrinogen, low levels of fibrinogen being caused in the experimental model used by dilution coagulopathy. The results of the examples confirm the key role of fibrinogen in this type of coagulopathy. The role of fibrinogen is further confirmed by the fact that the single administration of fibrinogen has an intrinsic potential to correct the effects of coagulopathy, in a dose-dependent manner.

Also, the results of the examples show that at least the complete restoration of the fibrinogen concentration to normal levels (ROTEM® test, EXTEM MCF) or even to slightly above normal levels (INTEM MCF and weight of clot) is necessary for optimal coagulation to be restored.

As already mentioned, it is remarkable that even at high doses of fibrinogen, no sign of potential hypercoagulation could be demonstrated.

Thus, the results of the examples show that the administration of a high single dose of fibrinogen, without assaying the level of circulating fibrinogen prior to administration, makes it possible to prevent the aggravation of haemostasis disorders, and this in all safety for the patient since, quite surprisingly, no potential effect of hypercoagulation is observed, even with high doses of fibrinogen.

In addition, it has been shown that administration of a single high dose of fibrinogen according to the invention does not induce a hypersensitivity reaction (measurement of plasma histamine levels). Plasma histamine levels remained stable for all fibrinogen doses (37.5 mg / kg to 600 mg / kg).

In addition, plasma concentrations of TNF-α were reduced or remained stable after fibrinogen administration. Finally, endothelin-1 has never been detected.

All of these results confirm the patient safety character of administering fibrinogen in a large single dose and rapidly for the treatment of severe acute bleeding.

The present invention relates to the use of fibrinogen for the manufacture of a medicament for use in the treatment of severe acute bleeding, said drug for administering an amount of at least 4.5 g fibrinogen in a single dose.

The invention also relates to fibrinogen for its use in the treatment of severe acute hemorrhage, intended for parenteral administration, preferably intravenously.

The invention also relates to a method for the prevention or treatment of severe acute hemorrhage, comprising a step of administering fibrinogen in an amount of at least 4.5 g.

It has been shown that a single step of administering the predetermined amount of at least 4.5 g of fibrinogen was sufficient for the prevention or treatment of severe acute hemorrhages.

It has been shown in the examples that the administration of a predetermined amount of at least 4.5 g of fibrinogen is effective and does not cause an adverse effect when the duration of fibrinogen administration is rapid. In particular, in the porcine experimental model illustrated in the examples, an amount of 600 mg / kg of fibrinogen could be administered in a duration of the 30 minute administration step. Transposed to humans, the results of the examples show that a dose of 6 g of fibrinogen can be administered very rapidly, with a duration of the administration step of less than 30 minutes. According to the invention, a duration of the fibrinogen administration step of less than 30 minutes, includes durations of less than 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, or 6 minutes.

In the case specifically illustrated in the examples, the step of administering an amount of 6 g of fibrinogen is five minutes, without causing any adverse effect to the patient.

According to the invention, a duration of the fibrinogen administration step of less than 30 minutes includes a duration of the administration step greater than 1 minute.

With the use of fibrinogen according to the invention, it is avoided the compulsory use of an assay of the circulating fibrinogen level in the patient prior to the administration of fibrinogen, which constitutes an important technical advantage for the prevention or the treatment of situations clinical urgency of patients, patients for whom a rapid preventive or therapeutic intervention can be decisive regarding the patient's chances of survival.

Thus, the use of fibrinogen according to the invention is further characterized in that the medicament is intended for the administration of an amount of at least 4.5 g of fibrinogen in a single dose, without control or dosage of circulating rate of fibrinogen prior to administration to the patient. Consequently, the method of prevention or treatment of the invention can be further characterized in that it does not include a control step or for assaying the level of circulating fibrinogen, prior to the fibrinogen administration step.

Also, the use of fibrinogen according to the invention is further characterized in that the medicament is intended for the administration of an amount of at least 4.5 g of fibrinogen in a single dose, without control or dosage of circulating rate of fibrinogen after administration to the patient. Consequently, the method of prevention or treatment of the invention can be further characterized in that it does not comprise a step of controlling or assaying the level of circulating fibrinogen, after the step of administration of the fibrinogen. In the alternative, the absence of a circulating fibrinogen level assay also represents an economic advantage over the overall cost of the preventive or curative medical procedure.

The amount of fibrinogen can be at least 4.5 g, 4.6 g, 4.7 g, 4.8 g, 4.9 g, 5 g, 5.1 g, 5.2 g, 5.3 g. g, 5.4 g, 5.5 g, 5.6 g, 5.7 g, 5.8 g or 5.9 g.

Although the maximum amount of fibrinogen to be administered is not an essential characteristic, said preferred amount of fibrinogen is preferably at most 15 g and most preferably at most 12 g.

Preferentially, the amount of fibrinogen is about 6 g, which includes an amount of fibrinogen ranging from 5.5 g to 6.5 g, including from 5.8 g to 6.2 g.

The use of fibrinogen according to the invention is suitable both for the prevention and the treatment of certain severe acute hemorrhages.

By way of example, the use of fibrinogen according to the invention is suitable for the treatment of postpartum hemorrhages.

In the conventional medical sense, postpartum haemorrhage is defined as bleeding with a volume of more than 500 mL during the 24-hour period following delivery.

There are various risk factors for postpartum hemorrhage, including persistent anemia, congenital hemostasis, and uterine atony (eg multiparity). , uterine over-distention, long or fast work, existence of chorioamnionitis), situations at risk of abnormal retraction of the uterus (eg retention of placental or clot, fibromatous or malformed uterus , placenta accreta), acquired haemostatic disorders, genital lesions including episiotomy, uterine inversion, uterine rupture or caesarean section.

Thus, in patients in whom there is a significant risk of triggering a postpartum haemorrhage, the use of fibrinogen according to the invention can be carried out as a preventive measure.

However, the use of fibrinogen according to the invention is mainly carried out curatively, in patients who have actually already triggered a postpartum haemorrhage. Similarly, the use of fibrinogen according to the invention is suitable for the prevention and treatment of hemorrhages caused during a surgical procedure. Indeed, the surgeon practitioner can determine in advance, from his experience, surgical procedures for which the risk of causing severe acute hemorrhage are significant. This is the reason why the use of fibrinogen according to the invention is also suitable for the prevention or treatment of severe acute haemorrhages caused during a surgical act, that is to say during a surgical procedure. .

Another situation of severe acute hemorrhage of great urgency is represented by traumatic hemorrhagic shock, caused by major physical trauma, which is often associated with a sudden drop in blood volume and acute anemia. In the case of traumatic hemorrhagic shock, the patient's vital prognosis is directly related to the volume of blood loss and the speed of treatment. This is another situation of acute bleeding for which a primary aspect of the therapeutic strategy is to restore hemostasis. Thus, according to another aspect, the use of fibrinogen according to the invention is suitable for the treatment of severe acute hemorrhages caused by physical trauma, also called traumatic hemorrhagic shocks.

Most preferably, fibrinogen is human fibrinogen. In certain embodiments of the use according to the invention, fibrinogen consists of a purified fibrinogen of natural origin.

In other embodiments of the use according to the invention, fibrinogen is a recombinant fibrinogen. For example, a recombinant fibrinogen prepared according to any one of the processes described in the PCT international applications published under the numbers WO-207/103447, WO-2005/010178, WO-1996/07728 or WO-1995 / may be used. 022,249. Said recombinant fibrinogen may be in the form of a pharmaceutical composition in which the recombinant fibrinogen molecules are combined with one or more pharmaceutically acceptable excipients.

In the embodiments of the use according to the invention in which the fibrinogen consists of purified fibrinogen of natural origin, said fibrinogen is purified from human plasma and is optionally combined with one or more pharmaceutically acceptable excipients.

One can for example use of purified fibrinogen of natural origin prepared as disclosed in European Patent Application Publication No. EP ⁰ 739 093. This 1 is a purified fibrinogen of natural origin is in the form of a fibrinogen concentrate obtained by a purification process from a solubilized human plasma fraction which comprises the successive steps of: a) chromatographic purification comprising the steps of i) charging a weak base type anion exchanger in said solubilized fraction, previously equilibrated by a predetermined ionic strength buffer of basic pH, ii) elution of a biological glue by increasing the ionic strength of said buffer, b) separation of Factor XIII from fibrinogen by addition to at least a part of the biological glue eluate at least one Factor XIII precipitating chemical, and recovering the resulting solution of purified fibrinogen supernatant, and c) diafiltering the solutions of fibrinogen, biological glue and remediated FXIII, followed by lyophilization of said solutions. .

It is also possible to use purified natural fibrinogen prepared as described in PCT Application No. WO-2005/004901. It is a purified fibrinogen of natural origin in the form of a fibrinogen concentrate obtained by a purification process comprising a viral inactivation step by heat treatment of a lyophilizate of cryoprecipitable proteins from human plasma. . The method described in PCT Application No. WO-2005/004901 is characterized in particular by the addition, before the cryoprecipitable proteins are put in the form of a lyophilizate, of a stabilizing and solubilizing formulation comprising a mixture of arginine, at least one hydrophobic amino acid and tri-sodium citrate.

One can also use the purified fibrinogen prepared from natural origin according to the general method described in European Patent Application No. EP ⁰ 1739093 and comprises a viral inactivation step of the type described for the method disclosed in the application PCT No. WO-2005/004901.

Fibrinogen purified of natural origin obtained in the form of a human plasma fibrinogen concentrate by a method selected from the methods described in European Patent Application No. EP ⁰ 1739093 or in the PCT Application No. WO-2005 / 004901 has the advantage of comprising a high concentration of human fibrinogen, of the order of 15g / l to 20g / l.

Such a human fibrinogen concentrate, which may also be designated "FGT1" in the present description, is particularly well suited to the uses of fibrinogen according to the invention which require the administration of a large amount of fibrinogen at least equal to 4, 5 g, preferably in a single dose, because of a high rate of fibrinogen recovery of this type of concentrate. Another advantage of this type of plasma-derived human fibrinogen concentrate is the low content of said concentrate in the other plasma proteins, which considerably reduces the total amount of plasma proteins that are administered to the patient.

An additional advantage is the high degree of viral safety of this type of human fibrinogen concentrate.

In certain preferred embodiments, the fibrinogen is in the form of a lyophilizate, optionally in combination with one or more pharmaceutically acceptable excipients. In certain preferred embodiments, the fibrinogen is in the form of a lyophilizate, optionally in combination with one or more pharmaceutically acceptable excipients, contained in a container whose inner atmosphere is maintained at a pressure below the pressure. atmospheric. Advantageously, the interior volume of the container is maintained under a partial vacuum vacuum.

Preferably, said lyophilized fibrinogen concentrate is contained in a vacuum reconstitution device. The fibrinogen solution to be administered to the patient can be prepared extemporaneously by adding to the vacuum vial the appropriate volume of a suitable solvent, for example water or a saline solution, sterile and pyrogen-free. In general, the fibrinogen solution obtained after reconstitution from the freeze-dried fibrinogen concentrate is stored at most for 7 days, preferably at most 24 hours, at 25 ^° C. or preferably at most 6 hours, at 25 ^° C.

In some embodiments of the use according to the invention, the purified fibrinogen or recombinant fibrinogen is in the form of a pharmaceutical composition in which said fibrinogen is combined with one or more excipients selected from lysine hydrochloride, trometamol, glycine, sodium citrate and sodium chloride. In some embodiments, said fibrinogen is combined with all of the following excipients: lysine hydrochloride, trometamol, glycine, sodium citrate and sodium chloride. Preferably, for the reconstitution of the liquid pharmaceutical composition to be administered, the solvent used consists of water for injection.

In other embodiments of the use according to the invention, the purified fibrinogen or the recombinant fibrinogen is in the form of a pharmaceutical composition in which said fibrinogen is combined with one or more excipients chosen from hydrochloride. arginine, isoleucine, lysine hydrochloride, glycine, and sodium citrate. In some embodiments, said fibrinogen is combined with all of the following excipients: arginine hydrochloride, isoleucine, lysine hydrochloride, glycine, and sodium citrate. Preferably, for the reconstitution of the liquid pharmaceutical composition to be administered, the solvent used consists of water for injection.

One flask of fibrinogen concentrate "FGT1" to be reconstituted with 100 ml of PPI water consists of 1.5 g of fibrinogen, 4 g of Arginine, 1 g of Isoleucine, 0.2 g of Lysine hydrochloride, 0, 2 g of Glycine and 0.25 g of sodium citrate

In still other embodiments of the use according to the invention, the purified fibrinogen or recombinant fibrinogen is in the form of a pharmaceutical composition in which said fibrinogen is combined with one or more excipients selected from albumin human, sodium chloride, arginine hydrochloride, sodium citrate, and sodium hydroxide. In some embodiments, said fibrinogen is combined with all of the following excipients: human albumin, sodium chloride, arginine hydrochloride, sodium citrate, and sodium hydroxide. Preferably, for the reconstitution of the liquid pharmaceutical composition to be administered, the solvent used consists of water for injections.

Preferably, the step of administering fibrinogen in an amount of at least 4.5 g is carried out with a medicament adapted for parenteral injection, of the type of the pharmaceutical compositions described above.

In the preferred embodiments of the use of fibrinogen according to the invention, it is carried out with a medicament adapted for intravenous administration.

(IV). Thus, in preferred embodiments of a method of preventive or curative treatment according to the invention, the step of administering fibrinogen in an amount of at least 4.5 g consists of a step of administration by route. intravenous.

In some embodiments of the use according to the invention, vacuum containers containing a lyophilized fibrinogen concentrate comprising an amount of 1.5 g of fibrinogen are used, which means that the contents of four containers are used to prepare administering the desired amount of fibrinogen in a single dose of about 6 g.

In certain preferred embodiments, the reconstituted liquid composition containing human fibrinogen is administered intravenously at a delivery rate of from 5 mL / minute to 30 mL / minute. Preferably, the reconstituted human fibrinogen liquid composition is administered intravenously at a flow rate of about 20 mL / minute, which ranges from 15 mL / minute to 25 mL / minute. A high rate of administration of this type is justified by the urgency to reduce homeostatic disorders in patients with severe acute bleeding.

The present invention also relates to a pharmaceutical composition comprising, as active ingredient, fibrinogen, said pharmaceutical composition being characterized in that it is adapted for the parenteral administration of a single dose comprising a quantity of fibrinogen at least 4.5 g, preferably about 6 g, of fibrinogen.

The present invention is further illustrated, but not limited to, the following examples.

EXAMPLES

Example 1 Treatment of Postpartum Hemorrhage

A. Materials and methods

Patients The present study included 320 patients who gave birth vaginally or by caesarean section at more than 27 weeks of pregnancy and had severe postpartum hemorrhage. Severe postpartum haemorrhage is characterized by a haemorrhage volume greater than or equal to 1000 ml and resistant to 2 uterotonic lines of treatment.

Human Plasma Fibrinoaene Concentrate Patients are treated with a single intravenous bolus injection of FGT1 (6 g) immediately after reconstitution. Human fibrinogen concentrate contains 1.5g of fibrinogen

Treatment Protocols The bolus administration of 6 g of fibrinogen is carried out without prior determination of the circulating fibrinogen level.

Clinical observations after treatment

The volume of blood lost after administration of FGT1 is monitored following the administration of 6g FGT1 administered intravenously and the time between FGT1 administration and the end of bleeding. The success of the therapy will be evaluated on the absence of recourse to ultimate resources, massive transfusion and deaths after 12 hours.

Example 2 Effectiveness in a Hemorrhagic Pig Model

A. Materials and methods

A.1. OBJECTIVE OF THE STUDY The study of Example 2 was to compare the efficacy of different doses (from 37.5 mg / kg to 600 mg / kg) of fibrinogen compositions (human fibrinogen concentrate compositions designated FGTT) in pigs The effect on hemostasis and blood loss was measured in animals after the application of standardized bone and liver injury.

The objectives of the study in Example 2 were to determine:

- whether the efficacy of a pro-coagulant treatment with different concentrations of fibrinogen induced different effects on hemostasis and blood loss;

- if a single administration of increasing doses of fibrinogen concentrate induced distinct plasma fibrinogen levels and / or resulted in distinct plasma half-life times for fibrinogen; and - if the pro-coagulant treatment with fibrinogen concentrate induced a hypercoagulation situation associated with subsequent thromboembolic complications.

A.2. Animals

This study was performed in forty-two healthy pigs aged 12 to 14 weeks and having a body weight of 25 to 35 kg. The animal model was used to determine the clot strength and blood loss dynamics in a situation where fibrinogen is administered after induction of dilution coagulopathy.

A3. Anesthesia

Animal pre-treatment was performed with azaperone (4 mg / kg, intramuscular injection-Stresnil ™, Janssen, Vienna, Austria) and atropine (0.1 mg / kg by intravenous injection). one hour before the start of the experiment. Induction and maintenance of anesthesia were performed with propofol (1-2 mg / kg by intravenous injection). For induction of analgesia, piritramide (30 mg, opioid with a half-life time of about 4-8 hours-dipidolor ™, Janssen, Vienna, Austria) was injected. Muscle relaxation was achieved by using 0.6 mg / kg per hour of rocuronium after endotracheal anesthesia. After obtaining endotracheal anesthesia, the femoral artery, the two femoral veins and the subclavian artery were anatomically prepared. The basic requirement for fluid replacement (4 mg / kg body weight) was achieved during the test with crystalloids (Ringer's lactate solution). The following invasive catheters were placed in these vessels: - a large-diameter catheter (double lumen venous access with a length of 20 cm),

- a Swan-Ganz catheter, and

- a means of invasive measurement of arterial blood pressure.

Then, the initial measurements of the basic values were carried out (time reference No. 2 in FIG. 1).

A.4. hemodilution

After being instrumented as described above, the animals underwent normovolemic hemodilution with a 6% solution of HES 130 / 0.4 (Voluven®, Fresenius Co., Bad Homburg, Germany).

The blood was taken from animals, step by step, using wide catheters and the blood was replaced with the colloidal solution in a ratio of 1: 1. For obtaining an estimated total blood loss of about 60%, the animals with a weight of for example 30 kg were infused with 1700 ml of 6% HES solution: 130 / 0.4 (Voluven®, Fresenius co ., Bad Homburg, Germany). After completion of the hemodilution, the collected blood was processed in a "cell saver" type system (Cats®, Fresenius), was concentrated and transfused again, to prevent anemia related to hemodynamics. The normovolemic hemodilution was achieved when the resulting coagulopathy reached a peak clot strength (MCF) value of less than 40 mm, as measured by thromboelastometry (time markers # 3 and 4 in Figure 1).

AT 5. Standardized bone injury

A standardized bone injury was achieved by drilling a 3 mm hole in the tibia head to a depth required to penetrate the bone marrow five minutes prior to administration of the test drug. Five minutes after the bone wound, a fibrinogen measurement was made (time mark # 5 in Figure 1), followed by administration of the test drug (time mark # 6 in Figure 1). Excess blood was removed by sucking the wound surface between bone and muscle and collecting it in a collection vessel. The duration to hemostasis was also determined. In the case where blood loss due to bone injury exceeded 500 ml, the bleeding was stopped by compression using a standard gas bandage, to ensure that the pig was stable from a hemodynamic point of view. for the next four hours of observation.

Fifteen minutes after administration of the test drug, additional determinations of all measured parameters were made (time parameter # 4). In addition, one hour, two hours and four hours after administration of the test drug, measurements of all parameters were made (time mark No. 7 in FIG. 1).

A.6. Standardized liver injury

Four hours after fibrinogen administration, standardized liver injury was achieved by making a central incision of the falciform ligament above the central liver lobe, using a template. The resulting hepatic incision is about 8 cm long and about 2 cm deep (time mark # 8 in Figure 1). No effect on the catecholamine-mediated circulation occurred. Achievement of a standardized hepatic injury was chosen to demonstrate that impaired coagulation negatively affected mortality.

Two hours after performing the liver injury or immediately before the suspected death, measurements were made of all parameters (time mark # 9 in Figure 1). Two hours after the liver injury, the surviving animals were euthanized by administering a potassium infusion. In accordance with the results of a previous pilot study, a two-hour follow-up evaluation was considered sufficient (Fries et al., 2006, Br. J. Anaesth., Vol 97 (4): 460-467; Fries et al., 2005, Br. J. Anaesth., Vol 95 (2): 172- 177).

It is specified that the study was carried out blind.

A7. Random distribution of animals ("randomization")

Using an appropriate computer program, the animals were randomized (randomized) to groups 1-6 (1: 37.5 mg / kg, 2: 75 mg / kg, 3: 150 mg / kg, 4: 300 mg / kg, 5: 450 mg / kg and 6: 600 mg / kg).

AT 8. Experimental protocol

The experimental protocol is illustrated in particular in FIG. 1 and in the Table below, in which the details of the treatment of the 42 pigs included in the study are described.

Table 1

for a 30 kg pork.

Fibrinogen administration was by infusion for 30 minutes.

A.9. Blood sampling and methods of analysis

All blood samples were collected from the femoral artery and the first volume of 5 ml of blood was removed. Blood samples for the ROTEM® study and coagulation analysis were collected in 3 ml tubes containing 0.3 ml (0.106 mol / L sodium citrate buffer pH 5.5) (Sarstedt, Nuermbrecht , Germany) Blood samples for the purpose of counting blood cells were collected in 2.7 ml tubes containing 1.6 mg DTA / ml (Sarstedt, Nuermbrecht, Germany) All tests were performed by the same experimenter, the prothrombin time (PT), the thromboplastin time (PTT-LA1), fibrinogen concentration, antithrombin (AT), and thrombin-antithrombin (TAT) were determined by standard laboratory methods using the appropriate tests of Dade Behring, Marburg, Germany and the Amelung coagulometry device (Baxter, UK). For D-dimer measurements, the D-dimer-0020008500® test (Instrumentation Laboratory Company, Lexington, USA) was used. Blood cell counts were performed using the Sysmex Poch-100i® counter (Sysmex, Lake Zurich, Illinois, USA).

A.10. Rotational thromboelastometry (ROTEM®) ROTEM® is a point of care diagnostic tool that allows a coagulation test on whole blood without the need for a time-consuming laboratory test. Moreover, and unlike standard coagulation tests, this method makes it possible to evaluate information on the quality of the clot, and especially on the clot firmness.

ROTEM® consists of a modification of the concept of thromboelastography that was originally developed by Hartert in 1948. The method consists of a cylindrical sensor that is immersed in a cuvette containing the blood sample. The sensor is rotating at a degree of 4.75 ° along its longitudinal axis. This movement is altered as the fibrin strands begin to form between the bowl and the sensor. This inhibition is detected by a change in the light reflection factor which is detected continuously and converted into a typical typical ROTEM® curve in which the following parameters are determined:

- CT, which means the clotting time, expressed in seconds, which is the time until the beginning of coagulation,

- CFT, which means the clot formation time ("Clôt Formation Time"), expressed in seconds, and the time elapsed between the start of coagulation and the moment when an amplitude of 20 mm is reached,

- MCF, which means the maximum clot firmness, which is expressed in millimeters, and which is the maximum amplitude, and

- the percentage of maximum lysis ("Maximum Lysis (%)"), which is the percentage of amplitude (MCF), 60 minutes after the onset of coagulation.

ROTEM® tests

Various tests of coagulation activating agents are available, as detailed below. The tests used in Example 2 are the INTEM test and the modified FIBTEM test. - INTEM: when performing an INTEM test, coagulation is activated via the intrinsic pathway. The INTEM reagent contains ellagic acid which is a strong activator of the intrinsic coagulation pathway.

- EXTEM: when performing an EXTEM test, coagulation is activated by tissue factor.

- FIBTEM: the FIBTEM test activates coagulation with tissue factor. At the same time, the thrombocyte function is altered by adding cytochalasin D. This results in the formation of a clot that is only induced by fibrinogen.

- The modified FIBTEM test was used because previous experiments showed that porcine thrombocytes can not be completely blocked by cytochalasin D. For this reason, thrombocytes were completely eliminated from this test by performing an EXTEM test. in a plasma sample instead of whole blood (modified FIBTEM).

A.1 1. Tissue sampling, preservation, slide preparation and microscopic examination Tissue samples from organs (lungs, heart, kidneys, intestines and liver) were collected at the end of each experiment after dissection of the animal. The samples were immediately immersed in 10% formalin solution. After dehydration with an ascending sequence of steps using alcohol samples, the samples were embedded in paraffin and cut into 7 μm thick slices. The assay slides were stained by the conventional hematoxylin / eosin staining technique and then examined.

AT 12. Statistical analysis

The Shapiro Wilks test was used to verify the normality of the distribution of study variables. The presumption of normality of the distribution was rejected for the following variables: ZVD, WEDGE, SPO2, CT, ALPHA.

For these variables, only non-parametric tests are valid.

Parametric tests: ANOVA was used for repeated measurements to evaluate the differences between the test groups. A significance value of 0.05 was used for the effect groups, the measures, and the group x measure.

Multiple comparisons were evaluated against the placebo group using the Dunnett protocol. A non-parametric test was performed using the Kruskal-Wallis test: all the groups were compared in one test. A level of statistical significance of 0.05 was retained.

The Wilcoxon test was compared for each test group, with the placebo group. Due to the Bonferroni multiple comparison protocol, only P values less than 0.0084 (0.05 divided by the number of comparisons (6)) were retained as statistically significant values.

A Jonckheere-Terpstra test was used to assess whether total blood loss decreased with increasing doses of fibrinogen. Since the distribution of total blood loss is not normal, this nonparametric test for ordered differences in treatment groups is appropriate (non parametric statistical methods, Hollander and

Wolfe, 1973). The Jonckheere-Terpstra test tests the null hypothesis that the distribution of total blood loss does not differ according to the groups that received separate doses of fibrinogen (respectively control, 37.5 mg / kg, 75 mg / kg, 150 mg / kg, 300 mg / kg, 450 mg / kg and 600 mg / kg).

The Student's test was used to compare the parameters specific to the baseline control values after hemodilution.

B. Results B.1. Coagulation parameters

1) ROTEM® parameters

ROTEM® parameters were significantly affected after hemodilution with Voluven®. Coagulation time increased and peak clot strength decreased significantly after Voluven® infusion (p <0.0001). In the same way, the angle α decreased significantly and the duration of clot formation significantly increased (p <0.001). Fibrinogen administration did not significantly shorten the prolonged duration of coagulation but significantly increased MCF (p <0.001 versus placebo for the corresponding 150 mg / kg to 600 mg / kg groups, 15 minutes after completion of fibrinogen infusion) and angle α (p <0.05 versus placebo for all dosing groups 15 minutes after completion of fibrinogen infusion). In the context of a 60% hemodilution, the results show that treatment with a dose of 150 mg / kg of fibrinogen is capable of completely restoring the initial baseline MCF values. Larger doses of fibrinogen induced an increase in INTEM MCF values to a maximum of 80 mm, at which point a plateau was reached (Figures 2-4 and 6).

2) Standard coagulation tests The PT value increased significantly from 1 1, 1 1 +/- 0.7 Is to 17.43 +/- 1, 9 Is after hemodilution (p <0.0001 vs baseline or "BL" for " Baseϋne "). Following fibrinogen administration, PT values increased significantly to 14.54

+/- 1, 44 s, compared with hemodilution (p <0.0001); however, there was no significant difference between the groups.

The aPTT values increased significantly from 10.94 +/- 3.34 s initially up to 21.7 +/- 2.72 sec after hemodilution (p <0.001).

After administration of fibrinogen, the value of aPTT remained high compared to the initial value. There was no difference between the groups, with the exception of the F group, which showed a significantly increased value of aPTT, compared with the placebo group and the other dosage groups, at all times after the administration of the drug. .

The value of D-dimer was significantly increased compared to placebo at four hours after fibrinogen administration for groups E and F (p <0.01). For groups A, D, E and F, there was a significant increase in the D-dimer value at the end of the experiment (p <0.05 for group A and p <0.001 for groups D, E and F).

The TAT and thrombin generation values (Calatzis method) were not different from the placebo group values.

3) Plasma Fibrinogen Concentration Plasma fibrinogen concentration significantly decreased after hemodilution, compared to baseline (p <0.0001) as shown in Figure 5. All animals treated with fibrinogen doses 150 mg / kg or greater had a significant increase in plasma fibrinogen levels compared to placebo at 15 minutes, 1 hour, 2 hours and 4 hours after treatment (p <0.001). Group B showed a significant increase in fibrinogen concentration only 15 minutes, 1 hour and 4 hours after treatment (p <0.05). All groups showed a reduction in fibrinogen levels 2 hours after hepatic injury or just before death. However, animals treated with doses of 300 mg / kg or more of fibrinogen exhibited significantly increased fibrinogen levels at 2 hours compared to placebo.

B.2. Blood count

After collecting 60% of the total blood volume and replacing the blood with Voluven®, the hemoglobin values dropped significantly to levels between 3-4 g / dl (p <0.0001, compared to baseline initial "BL"). The hematocrit significantly decreased in parallel to values between 11% and 13% (p <0.0001). After re-transfusion of red blood cells, hemoglobin and hematocrit values increased significantly up to 5-6 g / dl and up to 18-20% respectively (p <0.0001 versus the initial baseline "BL" for both parameters).

B.3. Haemodynamics and mixed oxygen saturation Mixed venous oxygen saturation significantly increased after haemodilution (p <0.0001 vs baseline baseline "BL") and increased again after re-transfusion of red blood cells (p <0.0001). <0.01 after hemodilution). There was a significant increase in mixed venous oxygen saturation (p <0.001), 2 hours after hepatic injury / or just before death, compared to time after red blood cell replenishment.

B.4. Blood loss

The total blood loss after bone injury and liver injury was:

- 42.12 +/- 18.792 ml / kg in the placebo group, - 41, 55 +/- 13.944 ml / kg at 37.5 mg / kg of fibrinogen,

- 34.30 +/- 13.593 ml / kg at 75 mg / kg of fibrinogen,

- 29.41 +/- 12,508 ml / kg to 150 mg / kg of fibrinogen,

29.79 +/- 10.155 ml / kg to 300 mg / kg of fibrinogen,

- 26.59 +/- 16.250 ml / kg at 450 mg / kg of fibrinogen, and - 28.02 +/- 10.325 ml / kg at 600 mg / kg of fibrinogen.

Statistical analysis showed a significant dose-response effect (p = 0.02), indicating a reduction in total blood loss for increasing doses of fibrinogen.

Animals that received 150 mg / kg of fibrinogen or larger doses showed a significant increase in clot formation capacity, as illustrated by a significant increase in clot size, formed on the surface of the liver after injury (150 mg / kg: p <0.05 versus placebo, 300-600 mg / kg: p <0.01 versus placebo), as shown in Figure 7.

B.6. Histological examination Histological examination of the kidneys, intestines, spleen, lungs, heart and liver of the animals did not show any microvascular thrombosis in the vessels.

Conclusion

It is known that, due to massive hemorrhage, the plasma fibrinogen concentration has reached critical levels before any other coagulation factor. The first line of treatment for severe haemorrhage is the administration of crystalloid and colloidal fluids to maintain normovolemia. However, colloids and especially hydroxyethylated starch (HES) are known to affect the polymerization of fibrin. The consequence is decreased clot resistance which can lead to subsequent blood loss.

The first main result of the study of Example 2 is that the administration of fibrinogen is capable of reversing the dilution coagulopathy in a dose-dependent manner. ROTEM® measurements clearly showed that maximum clot firmness (MCF) was increased and normalized after fibrinogen administration.

INTEM values showed that the administration of 150 mg / kg of fibrinogen was able to completely restore initial MCF values.

The modified FIBTEM test showed the same trend regarding MCF values; however, for 300, 450 and 600 mg / kg, MCF values increased well above baseline levels.

All animals exhibited a dose-dependent increase in plasma fibrinogen concentration, which was stable for the entire duration of the experiment.

It is important to note that, even though the plasma fibrinogen concentration increased beyond the baseline levels, the INTEM MCF values did not follow this increase linearly: at plasma fibrinogen concentrations of 350 mg / dl , MCF values reached a plateau; further increase in fibrinogen concentration did not lead to an increase in MCF values.

These results suggest the existence of a backup mechanism that prevents over-reaction of the coagulation process in a situation of excess bioavailability of fibrinogen. The presence of platelets seems to be crucial for the establishment of this mechanism, since plasma EXTEM data (or platelets are virtually absent) does not suggest such behavior: especially in high-dose groups (300-600 mg / kg) MCF values increase well beyond the baseline. Surprisingly, the results in Example 2 show that fibrinogen does not induce a hyper-coagulation state, nor does it induce thromboembolic events. In particular, no signs of thromboembolic events, either macroscopically or microscopically, were detected.

Fibrinogen did not affect thrombin production or TAT. To the Applicant's knowledge, the results presented in Example 2 are the first to show that doses of fibrinogen as high as 12 times the recommended fibrinogen dosage in humans (according to the Austrian Society of Anesthesiology, Reanimation and Intensive Care Medicine, available on the internet at the following address: http://www.oeagri.at/dateiarchiv/1 16 / Traumainduziertes ^o /o20Gerinnungsmanagement.pdf, can be administered in vivo.

The results of Example 2 also showed a reduction in blood loss after fibrinogen administration and a dose-dependent increase in clot size after hepatic injury.

These results are to be compared with the data regarding an increase in MCF values after fibrinogen treatment and clearly showed that clot formation capacity was improved after administration of fibrinogen. Notably, fibrinogen doses of 150mg / kg or greater significantly increased clot size.

In summary, the study of Example 2 confirms that the administration of a human fibrinogen concentrate (FGTW) is capable of reversing the dilution coagulopathy in a dose-dependent manner. Treatment at a dose of 150mg / kg is able to completely restore MCF values to baseline. The results of Example 2 also show that the INTEM MCF values plateau at plasma fibrinogen concentrations of 350 mg / dl. Higher concentrations of plasma fibrinogen do not increase the MCF values. Administration of fibrinogen significantly reduces blood loss after hepatic injury and increases coagulation capacity in a dose-dependent manner.

These results suggest that even larger doses of fibrinogen are completely safe in humans because hypercoagulability is not detected. Treatment at doses twelve times higher than the recommended human dose produces no adverse effects such as thrombosis or pulmonary embolism. The results of the examples also show that a large dose of fibrinogen can be administered according to a duration of the administration step which is very short, without causing any undesirable effect.

As detailed below, the results of Example 2, when transposed to human fibrinogen administration, indicate that a dose of 6 g of fibrinogen can be administered for a duration of 1 hour. a five-minute administration step, the administration of fibrinogen having the effect of preventing or stopping haemorrhage, without simultaneously causing an undesirable effect for the patient. More specifically, the results of Example 2 show that the expected beneficial effects on the recovery of haemodynamic parameters, without causing any undesirable effect, are obtained by administering to the experimental model of pork which is illustrated an amount of fibrinogen of 600 mg / ml. kg with a duration of the 30-minute administration step, i.e. a fibrinogen dose of 0.02 g / kg / min. It is specified that the fibrinogen concentrate composition which was used in Example 2 ("FGT1" composition) has a human fibrinogen concentration of 15 g / l, ie 0.015 g / ml. To obtain a fibrinogen dose of 0.02 g / kg / min, the pigs were administered a dose of the above-mentioned "FGT1" composition of 1.33 ml / kg / min. Transposed to a human patient having a weight of 60 kg, the administration of a dose of composition "FGT1" of 1.33 ml / kg / min consists in administering the composition "FGT1" to said patient according to a speed of administration of 80 ml / min. Thus, for a dose of fibrinogen of 6 g to be administered to said human patient, using the composition "FGT1", it is necessary to administer a volume of 400 ml of the composition "FGT1" to said patient, at the rate of administration. 80 ml / min. Accordingly, the dose of 6 g of fibrinogen, with the aforementioned rate of administration, can be administered to said human patient in five minutes.

Claims

1. Use of fibrinogen for the manufacture of a medicament for use in the prevention or treatment of severe acute bleeding, said medicament for administering an amount of at least 4.5 g of fibrinogen in a single, fast dose with a duration of administration of less than 30 minutes.

2. Use according to claim 1, characterized in that said medicament is suitable for parenteral administration, preferably intravenously.

3. Use according to one of claims 1 and 2, characterized in that said medicament is administered independently of the initial level of fibrinogen,

4. Use according to one of claims 1 to 3, characterized in that said drug can restore blood clotting capacity.

5. Use according to one of claims 1 to 3, characterized in that said drug can control severe acute bleeding and prevent its progression to uncontrollable haemorrhage.

6. Use according to one of claims 1 to 3 for the treatment of severe acute hemorrhage of the postpartum.

7. Use according to one of claims 1 to 3 for the prevention or treatment of severe acute hemorrhages during surgery.

8. Use according to one of claims 1 to 3, characterized in that said medicament is intended for the treatment of acute severe post-traumatic hemorrhages.

9. Use according to one of claims 1 to 3, characterized in that said medicament is intended for the prevention and treatment of other acute severe hemorrhages.

10. Use according to one of claims 1 to 9, characterized in that the fibrinogen is selected from a recombinant fibrinogen and a purified natural fibrinogen.

1. Use according to one of claims 1 to 9, characterized in that the fibrinogen consists of a fibrinogen concentrate with high viral safety.

12. Pharmaceutical composition comprising, as active ingredient, fibrinogen, characterized in that it is adapted for parenteral administration of a single dose comprising an amount of at least 4.5 g.