CA2107476C - Pharmaceutical packaging unit containing plasminogen activators for multiple bolus administration - Google Patents
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- CA2107476C CA2107476C CA002107476A CA2107476A CA2107476C CA 2107476 C CA2107476 C CA 2107476C CA 002107476 A CA002107476 A CA 002107476A CA 2107476 A CA2107476 A CA 2107476A CA 2107476 C CA2107476 C CA 2107476C
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Abstract
The subject of the present invention is a pharmaceutical packaging unit containing plasminogen activators for multiple bolus administration.
The invention especially concerns a pharma-ceutical packaging unit for the treatment of thrombo-embolic diseases, whereby the packaging unit essent-ially comprises two components and the first component is a pharmaceutically usual form of administration of a protein with plasminogen activator-like action with half life prolonged in comparison with t-PA and the second component consist of an instruction of the administration of this protein in the form of a fractionated admin-istration of two or more bolus injections.
The invention especially concerns a pharma-ceutical packaging unit for the treatment of thrombo-embolic diseases, whereby the packaging unit essent-ially comprises two components and the first component is a pharmaceutically usual form of administration of a protein with plasminogen activator-like action with half life prolonged in comparison with t-PA and the second component consist of an instruction of the administration of this protein in the form of a fractionated admin-istration of two or more bolus injections.
Description
2107~~7~
3oehrin;er Mannheim Umbi: 3496/OA/
Pharmaceutical DackaminS unit containin-, plasminomen activators for multipla bolus administration The subject of the present invention is a pharma-ceutical packaging unit containing plasminogen activators for multiple bolus administration.
The invention concerns a pharmaceutical packaging unit especially for the treatment of thromboembolic diseases, whereby the packa~ing unit essentially comprises two components and the first component consists of a pharmaceutically usual form of admin-istration of a protein with plasminogen activator-like action with a half life prolonged in comparison with t-PA and the second component is an instruction for the administration of this protein in the form of a fractionated administration of two or more bolus injections.
The thrombolytic therapy of myocardial infarction is an effective, medically well tested and proved therapy for the removal of occlusive. thrombi in the coronary arteries of the heart. t-PA (tissue-type plasminogen activator)produced by DNA technology is used worldwide for the therapy of this disease and has proved to be effective for the dissolution of coronary thrombi (Verstraete et al., Lancet II, 1985; 965-969)v By early lysis therapy of the myocardial infarction the residual heart function after myocardial infarction can be imprnved in comparison with other therapies (Armstrong et al., J. Hm. Col1. :;ardiola, 1989; 13: 1469-1476) and a hi~hP:- survival rate can be achieved in comparison with otiier therapies (Wilcox et a1., Lancet II, 1988; 525-539).
Large thrombolysis studies with, in all, several thousand patients show that a rapid induction of throznbolysis with early reperfusion of the -mycocardial tissue leads to a rescuing of'myocardial tissue and thus to an increase of the survival rate (GISSI study group, Lancet I, 1986; 397-401). For this reason, it is necessary to induce thrombo-lytic therapy at a point of time at which the tissue lying behind the coronary occlusion is not yet irreversibly damaged.
In principle, in the case of the treatment of a coronary occlusion, the problem exists of avoiding a possibly occurring reocclusion of the infarct blood vessel after successful initial thrombolysis. This dzsadvantageous effect could be observed for t-PA
(A1teplase)(Cbesebro et al., Circulation, 198?; 76s 142-154). The reocclusion of the inf'arct blood vessel leads to increased morbidity and mortality. For the prevention of reocciusions, substances are used of differing pharmacological working principlesy such as heparin (Bleich et a1., Am. J. Cardiol ., 1990;
66: 1412-1417) and acetylsalicylic acid (Hsia et al., N. En;1. J. Med,, 1990; 323: 1433-1437). The 21074t6 _4_ prolonged infusion of t-PA (Alteplase) is also said to prevent the reocclusiori of the infarct blood vessel due to a longer period of thrombolysis (sold et al,., Circulation, 1986; 73: 347-352;
Verstraete et al., Am. J. Cardiol., 1987; 60: 231-237;
Johns et a1o, Circulation, 1988; 78: 546-556). However, after ending of the infusion, the appearance of reocciusions is frequently observed.
-t-PA (rt-PA) produced by recombinant DNA technology has already been administered as double or multiple bolus to a few patients in the scope of clinical preliminary invest-igations. Admittedly, briefly, i.e, up to 90 minutes after the first bolus administration, high dissolution rates of the thrombi were found but, because of the high doses, an extensive, systemic plasminogen activation with subsequent almost complete reducing of fibrinogen was observed (only 15.5 to 5.2% of the initial value)(J. Am. Co21, Cardioll., 1991, 17(2), 152A). Thus undesired reduction of the fibrinogen level represents a disadvantage insofar.as, in the emergency operations, a high tendency to haemorrhage exists and an intensive supervision of the patient is necessary..Furthermore, in the case of double and triple bolus administration of rt-PA, there was disadvantageously observed a reocclusion tendency, i.e. a reocclusion of the blood vessel (Circulation, 1990, 82(4), Suppl. III, 538, abstract 2137; Br. J.
Haematol., 1991, 77, Suppl. 1, 47, abstract P080).
_5_ A study was also carried out for A1.teplase in which the multiple bolus administration was investigated in more detail (Coronary Artery Disease, 199C, 1.(1), 83-88).
Because of the above-mentioned disadvantageous effects, the administration of rt-PA in the form of a double or multiple bolus has found no practical use.
Furthermore, the bolus administration of rt-PA has been regarded as beina inexpedient alone from the fact that the half life of rt-PA is relatively short and only amounts to 3 - 6 minutes (Garabedian et al., J. Am. Coll, Cardiol., 1987; 9: 599 60?)m This means that, for the ensuring of the thrombolysis success, a relatively long-lasting infusion is necessary for the maintenance of effective plasma levels.
However, in emergency situations this long-lasting infusion of the substance (30 minutes to 6 hours) represents a di-stinct treatment disadvantage.
Furthermore, there then exists an increased danger of the appearance of tiaemorrhages (Marder and Sherry, Ne$ngls J. i1Cd=.' 1988; 318: 1512-1520 ) s 'Surpriaingly, it has now been found that proteins with plasminogen activator~'-like action and a half life prolonged in comparsion with t-PA can be used successfully in the treatment of coronary occlusions when the proteins used are administered in the form of a fractionated administration of two or more bolus injections. In this way, a rapid and simple administ-ration of proteins with plasminogen activator-like properties is made possible for the effective treatment of thromboembolic diseases.
SUMMARY OF THE PRESENT INVENTION
In accordance with one aspect of the present invention there is use of a plasminogen activator for the production of a pharmaceutical agent for extending the cumulative patency time in the treatment of a thromboembolic disease, said plasminogen activator being in the form of two or several bolus injections.
In accordance with another aspect of the present invention there is a pharmaceutical packaging unit for extending the cumulative patency time in the treatment of a thromboembolic disease, comprising essentially a pharmaceutical form of administration of a plasminogen activator in two or several separate containers.
BRIEF DESCRIPTION OF THE FIGURES
FIG. 1 compares the thrombolytic dose-effect range of thrombolytically active protein BM 06.022 and r-tPA
(Alteplase), following an intravenous injection extending over one minute, in a dog model where the animals have coronary thrombosis. Reperfusion rate (o) is given as the percent of reperfused dogs with a dosage group (each group contains 6 dogs). The curves were produced using semi-logarithmic regression analysis.
FIG. 2 compares plasma concentration time curves, pharmacokinetically, using BM 06.022 and rt-PA "Alteplase"
in anesthetized doges. Either 140 KU/kg (1 mg/kg) of Alteplase were used, in groups of six dogs. What is presented is the arithmetic mean value. Starting concentration was subtracted from subsequent measurements.
- 6a -FIG. 3 displays time course of coronary blood flow after a single intravenous bolus injection, using either 140 or 280 KU/kg of BM 06.022, at time point t=0 minutes.
Data are shown in mean values ±SEM, where four dogs were used in each group. "Pre" indicates initial blood flow.
FIG. 4 also shows time course of coronary blood flow as in FIG. 3, (single bolus, t=o), and double bolus (140 KU/kg each bolus, at t=0 and t=44 minutes).
FIG. 5 shows time course of coronary blood flow before and after double bolus administration (140 KU/kg each bolus, for six dogs), or of 1.40 and 50 KU/kg administrations (for five dogs), of BM 06.022 in both cases, again at t=0 and t=44 minutes. Data are calculated and presented as explained in FIG. 3.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
In the meantime of the present invention, the term "fractionated administration" means the administration of a therapeutically effective amount of a thrombolytically active protein in two or more partial amounts in the form of a double or multiple bolus, "therapeutically effective amount" referring to sufficient protein to inhibit reocclusion.
Bolus administration is an intravenous rapid injection and is, therefore, especially advantageous because the time from the beginning of the clinical symptoms up to treatment and, also the time from the commencement of treatment up to the dissolving of the coronary thrombus is shortened. Thus, more myocardial tissue can be rescued from irreversible destruction. The double or multiple bolus administration according to the invention brings about a higher thrombolytic potency of the thrombolytically active protein use. In this way, it is possible to reduce dosages as ~ ~. , . u..~~. .. ~ ~ . . ,,.~ ,. .-,m.~ . ._ . ...... - . _ .-..~.. .-.m_ - ---. _ õ ~
- 6b -compared to rt-PA. Surprisingly, after bolus injection, more rapid reperfusion as compared to administration in the form of an infusion is found. Furthermore, double or multiple bolus administration brings about a significant prolongation of the cumulative patency time (sum of the time intervals after reperfusion in which coronary blood flow is present), as well as a ~. *
distinct increase of the coronary blood flow, whereby this also remains stable at a relatively high level for a longer time after the administration. Further-more, the double or multiple bolus injection has the surprising advantage of a smaller decrease of the plasmd.fibrinogen in contradistinction to a single bolus injection However, these advantageous properties of a double or multiple bolus administration are not achieved when the administration of the corresponding total amount of the plasminogen activator took place by a single bolus injection.
Thromboembolic diseases in the meaning of the present invention are especially those diseases which are to be attributed to a myocardial infarction or to a reocciusion of the coronary arteriese whereby re-occl:usions occur above all in the case of the use of thrombolytics for the treatment of the heart infarct.
The cause for a myocardial infarction is the formation of a thrombus in the coronary arteries. This thrombus consists of a combination of .'-~fibrin and thrombocytes.
TO&primar9= aim in the case of the treatmant of the heart infarct is the rapid dissolution of this throinbus and the recreation of the blood flow (reperf'usion).
A successful therapy must dissolve the thrombus as quickly as possible and prevent its reformation (reocclusion). The double or multiple bolus admin-istration according to the invention is especially advantageous for the treatment of the brain stroke.
21074"16 The pharmaceutical packaging unit according to the invention consists, on the one hand, of a form of administration which contains the plasminogen activator and, on the other hand, of an instruction, for example in the form of a package leaflet prescribed for medicaments, from which it follows that the administration of a therapeutically effective amount of the plasminogen activator advantageously takes place by double or multiple bolus administration. This information regarding the manner of the use can also be given as packaging overprint on the medicinal prep.arati.on ready for sale or can be taken from an information leaflet which can be brought together with medicinal preparations which contain the plasminogen activators in the meaning of the present invehtiona As suitable forms of administration for plasminogen activators, from the prior art there come into question known galenical formulations, for example lyophilisates or solutions in appropriate containers, such as e.g. in ampoules. As a rule, these formulations contain usual pharmaceutical adjuvants which are suitable for the prepaxation of isotonic solutionsf and possibly further stabilising and solubilising agents.
As proteins with plasminogen activator-like action (in the meaning of the present invention also briefly called "plasminogen activators"), those proteins are used which, in comparison with t-PA, possess a prolonged half life time. Plasminogen activators bring about the dissolving of thrombi and thereby make possible that blood again flows through the blood vessels concerned. As already mentioned above, the half life of t-PA in the human blood is about 3 - 6 minutes. In the meaning of the present invention, those plasminogen activators preferably come into question which, in comparison with t-PA, possess a half life prolonged by at least the factor 2, preferably 3 - 7 and especially 3 - 5. In particular, the half life should amount to at least about 10 minutes, for example 10 - 90, 10 - 40 or 10 - 20 minutes. Those proteins are preferably employed, the half life time of which is longer than the half life time of t-PA by the factor 3 - 30, especially 5 - 20 times.
In the meaning of the present invention, as proteins with plasminogen activatoralike action, there come into question, for example, the following plasminogen activatorsc LY 210825 (.K2P from "Syrian hamster cell 1ine:=" , Cira.I.990, 82, 930-940) ;-DeltaMx and-DeltaFElx (K1K2P from "Chinese hamster ovarycelisn, B1ood,'1988; 71, 216-219); DeltaFEKl (K2P from "mouse C127 cells", J. Cardibvasc. Pharmacol., 1990, 16, 197-203); E-6010 (Jap. Circ. J., 1989, 53, p. 918); t-PA variants.(Thromb. Haemost,, 1989, 62, P. 542); K2P and D-K2P (Thromb. Haemost., 1989, 62,.
p. 393); MB-1018 (FK2K2P, Thromb. Haemost., 1989, 62, -lc-pe 543); FK2P (FASEB J., 1989, 3, A1031, abstract 4791); Deltalx (Circulation, 1988, 4, 11-15);
i{1K2P (Thromb. Res., 1988, 5C, 33-41); FK1K2P
(J. Bio1. Chem., 1988, 263, 1599-1602). There are especially used recombinant plasminogen activators of the type K2P, such as e.g. BM 06.022 (known from EP-A-0 382 174). Further t-PA muteins of this kind are described in the following Patent Applications:
EP-A-o,J96,920, EP-A-0,207,589; AU 61804/86;
EP-A-0,231,624; EP-A- 0,289,508; JP 63133988;
EP-A-0,234,051; EP-A-0,263,172; EP-A-0,241,208;
EP-A-0,292,009; EP-A-0,297,066; EP-A-09302,4569 EP-A-0,379,890.
The pharmabbki.tietic profile of rt-PA (A1teplase) 15. is typically characterised by a two or even three compartment model (Thromb. Haemost., 1989, 61, 497-501), The most important-is thereby'the,initial half life since, with 66%, it contributes to the total area under the curve (area under the curve = AUC). Therefore, this half time is designated as the dominant half life. Tn the case of plasmi.nogen activators der.ived from t-PA
with prolonged half life, the pharmacokinetic propi1e is -naturally also changed. The designation "prolonged half life" thereby refers to the dominant half life, which is characteristic for the pharmacokinetic profile because it contributes quantitatively the most to the total AUC. The determination of the half life takes place accordin; to the processes known frnm the prior art (Pharmacokinetics, Ch. 2, Viarcel Dekker, New York, 1982).
As plasminogen activators, those especially derivatives of t-PA produced by recombinant DNA technology come into question which essentially include those protein repions of the natural protein which are responsible for the fibrinolysis of the thrombi. Those derivatives I0 of t-PA can thereby also be-used which display deletions or substitutions of individual or several amino acids in the sequence of the t-PA as long as the half life of such derivatives is prolonged in the above meaning with regard to t-PA. These are especially t-PA derivatives of the type K2P.
By way of example for the plasminogen activators coming into question in the meaning of the present invention, there was used the plasminogen activator K2P (BM 06.002) described in more detail in European Patent Application EP4A-0,382,174. It consists of the kringle 2(K) and -p~rotiease (P) domains of human t-PA and, because of its expression in Escherichia coli cells, is present in unglycosilated form. The specific activity of K2P amounts to 550,000 +/-IU/mg (= 550 +/- 200 KU/mg). K2P is used for the thrombolytic therapy of thromboembolic diseases (myocardial infarction, lung embolism, stroke and other occlusive vessel diseases) ':'tie missin~ ~1;~c osilatic~n leads ;, p a reduced clearance and results in a half life prolonged iith ra:;a~d to t-PA by at least the factor 3- 4.
The prolonged half life makes possible the use rfl the substance as clinically desirable multiple bolus administration. The thrombolytic effect of the bolus administration of the substance was investigated in various animal models. There was thereby shown a thrombolytic effectiveness increased approximately by the factor 5 compared with Alteplase. Especially impressive was thereby the rapid opening of thrombotically occluded vessels, in comparsion with Alteplase and other thrombolytics, which means a reduced time for the blood vessel opening in comparison with other thrombolytics and thus a more rapid reperfusion of the infarcted myocardial tissue.
On the basis of the improved pharmacokinetic properties of K2P, the effective dose to be admin-istered can be reduced and K2P can be administered as i.vs multiple bolus injection. A very rapid maximum reperl'nsion is therewith achinved. Furthermore, by the administration of a double or multiple bolus, the impairment of perfusion in the coronary arteries after reperfusion (tendency to reocclusion) can be significantly reduced or prevented. In addition, after double or multiple boli, the decrease of the plasma fibrinogen is smaller than after the single bolus 210 7d 7 G
-lj-in,jection nf the same total dose.
Besides the rapid opening of thrombotically occluded blood vessels, the prevention of the rencclusion of the npened binod vessels is of ~reat clinical importance since a reocclusion of the infarct blood vessel can lead to a clinically important reinfarct with the clinical complications resulting therefrom, The administration of K2P as bolus in equi-effective dose in comparison to the infusion of Alteplase leads, in the animal model, to a surprisingly rapid opening of the infarct blood vessel and thus makes possible a rapid reperfusion of the infarcted mycocardial tissue. In view of the present 1.5 knowledge regarding thrombolysis, this observed effect is to be evaluated very positively from the clinical point of va.ew.
The problem of the reocclusion of thrombolysed infarct blood vessels occurd, according to the atiimal ox perimental data, also with the single bolus of K2P
to an extent which is compaxable. wi.th other tbrombo-lytics Surprisingly, it is shown that a dividing up of the total dose of K2P into two or more chrono-logically staggered injected boli reduces or even prevents the appearance of perfusion impair-ments in the coronary arteries after reperfusion.
The time integral between beginning of the first and a subsequently following bolus injection is 2107417b indication-dependent and can amrun't to between 1.C
minutes and 6 hours, preferably 20 minutes and 2 - 3 hours, especially 30 - 9G minutes. The injection period is itself relatively short and amoun-us, dependinb upon the volume to be administered, to about 0 5 - 3 minutes, whereby an injection rate of about 5 - 10 ml/min, is advantageous. In the meaning of the present invention, by the term multiple balus administration one also understands the cumulated administration of two or more individual bolus injections which are administered in the scope of an interval therapy. Two or more, preferably three, bolus injections are carried out daily, whereby the therapy can eictend over the course of several successive days.
This treatment scheme can possibly also be interrupted 1+
by one or two days so that the administration in the form of bolus injections can take place, for example, on every second day during the whole treatment period Essentially, this -cumulated adn3nistratieu of single boli difi'e.rs from the known single bolus administration in that the administration is repeated daily several times-o If tha time interval between the first and a subsequently following bolus injection is relatively short, about 10 minutes up to one hour,one speaks of a double bolus or multiple bolus administration. If the time intervals are of the order of magnitude of several hours,then one can also speak of a cumulated administration of single boli.
210'7476 _15_ Fnr the production of the pharmaceutic3l forms of administration with regard to the prrtein with plasminogen activator-like action, there are used the usual pharmaceutical adjuvant and additive materia3.s Furthermore, there can be added thereto stabilising or the known solubilising agents, such as e.g. basic amino acids (arginine, lysine or ornithine).
Suitable galenical forms of administration are known from the prior art or can be produced according to the generally usual methods (cf. EP 0,217,379;
EP 0,228,862; WO 91/08763; WO 91/08764; WO 91/08765;
W0 91/08766; WO 91/08767 or WO 90/01334. The form of administration ca-n be present in lyophilised form or as injection solution ready for use. The bolus injection can take place intravenously, intramuscularly or also subcutaneously, whereby the i.v. injection is preferred.
In the case_ of the production of the pharma-ceutical packaging unit, the amounts of the protein to be admini.stsred for the two or multiple bolus injection can be made available in one or more, preferably in two separate containers, such as eog.
ampoules, It is aIlso possible to make available a multidose form of administration from which the particularly desired amount of the protein can be removed by withdrawal into a suitable'syringe. The ampoules can contain the solution already ready for injection or, alternatively, a lyophilisate oz the protein and possibly further pharmaceutically usual adjuvant or carrier materials. The lyophilisates are i-nixed with usual injection solutions shortly before the administration so that a solution results which can be administered directl.y.
The amount of the protein with plasminogen activator-like action can be the same or also different in the first and second container, depending upon the particular requirements. As a rule, amounts of 3 - 50 MU per container are used, For the first bolus injection, there is preferably administered a larger amount of the protein than for the second bolus injection. In particular, amounts of 5 - 20 MU, especially of 5 - 15 MU, are used for the first and 3 - 15 MU, especiall9 3- 10 MU for the second bolus injection. Especially preferably, one uses about 10 MU f or the first.- and about 5 MU for t he second injections The cumulative dose preferably lies in the range of 15 - 40 MQw The invention is acpla.ined in more detail on the basis of the following Examples:
Example 1 For the acperiments, a dog model was used'for the simulation of the myocardial infarction. Adult beagles of both sexes were narcotised with barbiturate, intubated and artificially respirated. Arterial and venous catheters were applied in order to monitor the blood pressure, tc administer substances or to take blood samples. The chest was opened and the heart exposed. A short segment of the ramus circumf].exus of the arteria coronaria sinistra was isolated and prepared. Subsequently, the coronary artery was provided with the following instruments from proximal to distal: an electromagnetic flow probefor the measurement of the blood flow in the coronary artery, a stimulation electrode, an adjustable screw and a thread,. The tip of the stimulation electrode was passed through the wall of the coronary artery and so placed within the blood vessel that the needle tip came into contact with the inner surface of the blood vessel. The screw'Was so adjusted that 90% of the reactive hyperaemia were eliminitad as the result of a short-term interruption of perfusion of the coronary artery.
The thrombus in the coronary artery for the initiation of the myocardial infarction was formed according to the method originally-deaoribed by Romson at a1.
(Thromb. Res,., 1980; 17s= 841-853). The method was employed in the modUfied form of 'Shebusk.i at a1., (J. Pharmac-ol. Exp. Ther.., 1988; 246: 790-796). Via the. stimulation'electrode, a 150 microamVere direct current was applied to the coronary artery and maintained until the blood flow in the coronary artery fell to 0 ml/min and remained there for at least 3 minutes. Subsequently, the thrombus should age for 30 minuteso In this time, the animals were --heparinised in a dosage of lCu(, units/animal/hour.
30 minutes after thrombus fcrmation, the fibrino-lytic or the solvent was administered as a one-minute i.v. bolus injection to six animals per group.
BM 06sC22 was administered in four doses: 50, 100, 14C and 200 KU/kg. Alteplase was also given in four doses: 200, 800' 1130 and 160C KU/kg (= 2 mg/kg).
The specific activity of Alteplase amounted to 800,000 IU/mg (= 800 kU/mg). Plasma samples were obtained before and repeatedly after injection of the fibrinolytic in order to determine the concent-ration of the activity of BM 06.022 or of Alteplase according to the method of Verheijen et al. (Thromb.
Haemostas., 1982; 48: 266-269). A reperfusion was assumed if at least 33% of the initial blood flow in the coronary artery was again achieved. Accordingly, the time to reperfusion from the beginning of the injection up to the achievement of this blood flow was defined, In each of a further experimental group,,. BM 06,022 waa administered in a dosage of 140 KU/kg and ' Alteplase i.n a dosage of 800 KU/kg l nng/kg) intravenoualy over 90 minutes as continuous"'infusion (10% of the totaldosage as initial i.v. bolus). ' Figure 1 shows the dose-effect relationship for the reperfusion rate after bolus injection of BM 06.022 or of Alteplaseo A 1CC% reperfusion rate (6 out of 6 animals) was achieved with i.v. bolus 21~7 d 7 6 injection of 2CG ItiU/kg in the case of BM 06.022. In the case of the same dose, none of the animals injected with Alteplase reperfused. For the achievement of the same maximum effect, an injection of 1600 KU/kg (= 2 mg/kg) of Alteplase was necessary. This injection dose is approximately twice as high as the dose of Alteplase at present used in the clinic as infusion (about 1 mg/kg). The higher thrombolqtic potency of BM 06.022, as seen by the shift to the left of the dose effect curve, can be explained by the improved pharmacokinetic properties of BM 06.022=(see Fig. 2).
The total plasma clearance of BM C6.022 amounts to 4.4 + 0.4 and that of Alteplase to 20.4 2,0 ml/
min- x kg Thus the total plasma clearance (a measure for the removal of a substance from the plasma) of BM 06.022 is about 4.6 times slower than that of Alteplase. Furthermore, it was found that Altepl.ase, after infusion, achieves a twice as high reperfusion rate thaa after single bolus injection of the same dose (800 KU/kg). BM 06.022 in a dose of 140 KU/kg shows the same reperfusion rate (4 out of 6 aniinals) after injection and after infusion but the time to reperfusion was significantly shorter after injection (Table 1).
The results show that Alteplase displays more favourable thrombolytic success after infusion than after injection, 'Lhis finding agrees with the clinical _. practice in which Alteplase is usually administered ..2C-as infusion (Chesebro et al., Circulaticn, 1987; '~J6e 142-254) .'1'he slower elimination of BM C6.C122 in compara.snn with Alteplase from the plasma led to a hioher thrombolytic potency of BM C6.022. Not only is dosage reduction thereby possible but, after single i.v. bolus injection, surprisingly a more rapid reperfusion takes place than after infusion.
Example 2 For the experiments in Example 2, there was used 1C the same dog model of coronary artery thrombosis as in Example 1. However, in contradistinction to the experiments in Example 1, the thrombus ageing in the case of administration of the substances was one hour instead of only 30 minutes. BM 06.022 was administered either as a single i.v. bolus injection or as a double bolus injection, in each case over one minute. The single bolus dose amounted to 140 or 280 KU/kg. In both groups with the double bolus administration, the first bolus dose amounted to 140 KU/kg in each case, follow by the second bolus in a dose of 140 or 50 KU/kg of BM 06.022 (44 minutes later). Each of the four experimen~al groups consisted bf six dogs. Additional parameters for the evaluation were the maximum of the coronary blood flow which was measured after reperfusion and the coronary blood flow at termination of the experiment three hours after injection. Further-more, the cumulative patency time was calculated;
thereunder one understands the sum of the time 210"14 76 intervals after reperfusion in which a coronary blood flow was present. The animal model is so designed that typically after ~eperfusinn cyclic flow vari.ations occur with reocclusion. Nt the end of the experiment, the residual thrombus was removed and its wet weight measured.
In both groups with single bolus injection, the reperfusion rate amounted to four of six animals. In the double bolus group, it was higher and amounted to six of six or five of six animals (Table 2). The results shown in Table 3 for the cumulative patency time, the coronary blood flow and the residual thrombus weight show that a double bolus administration of BM 06.022 significantly prolongs the cumulative patency time, significantly increases the coronary blood flow, this also" remains significantly increased at the end of the experiment and the residual thrombus weight has-decreased significantly in comparison with the single bolus injection of 140 KU/kg. Figure 3 shows that an increase of the single bolus injection from 140 to 280 KU/kg cannot markedly improve the corftaxy, blood flow. 9tr4ngly cyclic flow variations are still observed'. Cn ih.e other hand=,. in Pigure 4 it can be seen that the double bolus administration of 140 and 140 KU/kg of BY, 06.022 clearly improves the coronary blood flow in comparison with the single bolus injection of 140 ItU/kg of BM 06,G22. The same effect on the coronary blood flow can also be achieved by "-he double bolus administration of 14C and 5(. KU/krg of 3i4I C6.L22 (Figure 5). As Table 4 shows, the dividing up of a total drse of 28C KU/kg into 140 and 140 KU/kg instead of the sin;;le bolus injection of the same total dose of Bi1 C6,c.22 prevents the significant decrease of the plasma fibrinogen.
The results show that the deterioration of the coronary blood flow after reperfusion, which in this model intrinsically occurs in simulation of patients with reocclusion tendency, cannot be prevented by the single bolus injection even at the higher dosage. On the other hand, the administration of the double bolus is surprisingly able not only to prolong the cumulative patency time but also quantitatively to increase the flow and to maintain it increased up to the end of the experiment. In addition, the double bolus injection has the surprisingly advantage of the lower decrease of the plasmafibrinogen in contradistinction to the single bolus injection of the same total dosage. The size of the second bolus in the double bolus administ-ration can thereby be made variable without limiting the successd Description of the Figures Figure 1 shows the thrombolytic dose-effect range of BM 06.022 and of Alteplase after I minute i.v, injection in dogs with coronary thrombosis. The reperfusion rate (%) signifies the percentage of reperfused dogs within a dosal-e group (n = 6 per dose). The dose effect curves were produced with the help of a semi-logarithmic re;;ression analysis.
The Alteplase group without reperfusinn (at 20U KU/kg) was excluded from the regression analysis.
Figure 2 shows the pharmacokinetic comparison of the plasma concentration time curves for the activity of BM 06.022 and Alteplase in anaesthetised dogs.
A one minute i.v, injection of 140 FCU/kg of BM C6.022 or of 800 KU/kg (= 1 mg/kg) of Alteplase,was administered to six dogs per substance. The data represent the arithmetic mean value. The starting concentration was substracted from the subsequently measured concent-rations.
Figure 3 shows the time-course of the coronary blood flow before and after a single i.v. bolus injection with 140 or 280 KU/kg of BM 06.022 at the time point t = 0 minutes. The data are mean values SEM; n = 4 reperf'used dogs per group. Pro represents the initial blood flow.
Figure 4 shows the time-course of the coronary blood flow before and af'ter a single i.v. bolus injection of 140 RU/kg of BM 06.022 *(n = 4) at the time point t = 0 minutes or of a double bolus injection of 140 and 140 KfJ/kg of BM 06.022 (n = 6) at the time point t= 0 and t = 44 minutes. The data are average values + SEM; Pre is the initial blood flow.
_?L}_ :r,isure 5 shorrs the course ~f time of the coronary blood flow befoi=e and after double bolus administration of 14C and 140 KU/kg (n = 6) or of 140 and 50 KU/kg of BM 06.022 (n = 5) at the time point t = 0 and t= 44 minutes. The data are avera;e values + Sai;
Pre is the initial blood flows
3oehrin;er Mannheim Umbi: 3496/OA/
Pharmaceutical DackaminS unit containin-, plasminomen activators for multipla bolus administration The subject of the present invention is a pharma-ceutical packaging unit containing plasminogen activators for multiple bolus administration.
The invention concerns a pharmaceutical packaging unit especially for the treatment of thromboembolic diseases, whereby the packa~ing unit essentially comprises two components and the first component consists of a pharmaceutically usual form of admin-istration of a protein with plasminogen activator-like action with a half life prolonged in comparison with t-PA and the second component is an instruction for the administration of this protein in the form of a fractionated administration of two or more bolus injections.
The thrombolytic therapy of myocardial infarction is an effective, medically well tested and proved therapy for the removal of occlusive. thrombi in the coronary arteries of the heart. t-PA (tissue-type plasminogen activator)produced by DNA technology is used worldwide for the therapy of this disease and has proved to be effective for the dissolution of coronary thrombi (Verstraete et al., Lancet II, 1985; 965-969)v By early lysis therapy of the myocardial infarction the residual heart function after myocardial infarction can be imprnved in comparison with other therapies (Armstrong et al., J. Hm. Col1. :;ardiola, 1989; 13: 1469-1476) and a hi~hP:- survival rate can be achieved in comparison with otiier therapies (Wilcox et a1., Lancet II, 1988; 525-539).
Large thrombolysis studies with, in all, several thousand patients show that a rapid induction of throznbolysis with early reperfusion of the -mycocardial tissue leads to a rescuing of'myocardial tissue and thus to an increase of the survival rate (GISSI study group, Lancet I, 1986; 397-401). For this reason, it is necessary to induce thrombo-lytic therapy at a point of time at which the tissue lying behind the coronary occlusion is not yet irreversibly damaged.
In principle, in the case of the treatment of a coronary occlusion, the problem exists of avoiding a possibly occurring reocclusion of the infarct blood vessel after successful initial thrombolysis. This dzsadvantageous effect could be observed for t-PA
(A1teplase)(Cbesebro et al., Circulation, 198?; 76s 142-154). The reocclusion of the inf'arct blood vessel leads to increased morbidity and mortality. For the prevention of reocciusions, substances are used of differing pharmacological working principlesy such as heparin (Bleich et a1., Am. J. Cardiol ., 1990;
66: 1412-1417) and acetylsalicylic acid (Hsia et al., N. En;1. J. Med,, 1990; 323: 1433-1437). The 21074t6 _4_ prolonged infusion of t-PA (Alteplase) is also said to prevent the reocclusiori of the infarct blood vessel due to a longer period of thrombolysis (sold et al,., Circulation, 1986; 73: 347-352;
Verstraete et al., Am. J. Cardiol., 1987; 60: 231-237;
Johns et a1o, Circulation, 1988; 78: 546-556). However, after ending of the infusion, the appearance of reocciusions is frequently observed.
-t-PA (rt-PA) produced by recombinant DNA technology has already been administered as double or multiple bolus to a few patients in the scope of clinical preliminary invest-igations. Admittedly, briefly, i.e, up to 90 minutes after the first bolus administration, high dissolution rates of the thrombi were found but, because of the high doses, an extensive, systemic plasminogen activation with subsequent almost complete reducing of fibrinogen was observed (only 15.5 to 5.2% of the initial value)(J. Am. Co21, Cardioll., 1991, 17(2), 152A). Thus undesired reduction of the fibrinogen level represents a disadvantage insofar.as, in the emergency operations, a high tendency to haemorrhage exists and an intensive supervision of the patient is necessary..Furthermore, in the case of double and triple bolus administration of rt-PA, there was disadvantageously observed a reocclusion tendency, i.e. a reocclusion of the blood vessel (Circulation, 1990, 82(4), Suppl. III, 538, abstract 2137; Br. J.
Haematol., 1991, 77, Suppl. 1, 47, abstract P080).
_5_ A study was also carried out for A1.teplase in which the multiple bolus administration was investigated in more detail (Coronary Artery Disease, 199C, 1.(1), 83-88).
Because of the above-mentioned disadvantageous effects, the administration of rt-PA in the form of a double or multiple bolus has found no practical use.
Furthermore, the bolus administration of rt-PA has been regarded as beina inexpedient alone from the fact that the half life of rt-PA is relatively short and only amounts to 3 - 6 minutes (Garabedian et al., J. Am. Coll, Cardiol., 1987; 9: 599 60?)m This means that, for the ensuring of the thrombolysis success, a relatively long-lasting infusion is necessary for the maintenance of effective plasma levels.
However, in emergency situations this long-lasting infusion of the substance (30 minutes to 6 hours) represents a di-stinct treatment disadvantage.
Furthermore, there then exists an increased danger of the appearance of tiaemorrhages (Marder and Sherry, Ne$ngls J. i1Cd=.' 1988; 318: 1512-1520 ) s 'Surpriaingly, it has now been found that proteins with plasminogen activator~'-like action and a half life prolonged in comparsion with t-PA can be used successfully in the treatment of coronary occlusions when the proteins used are administered in the form of a fractionated administration of two or more bolus injections. In this way, a rapid and simple administ-ration of proteins with plasminogen activator-like properties is made possible for the effective treatment of thromboembolic diseases.
SUMMARY OF THE PRESENT INVENTION
In accordance with one aspect of the present invention there is use of a plasminogen activator for the production of a pharmaceutical agent for extending the cumulative patency time in the treatment of a thromboembolic disease, said plasminogen activator being in the form of two or several bolus injections.
In accordance with another aspect of the present invention there is a pharmaceutical packaging unit for extending the cumulative patency time in the treatment of a thromboembolic disease, comprising essentially a pharmaceutical form of administration of a plasminogen activator in two or several separate containers.
BRIEF DESCRIPTION OF THE FIGURES
FIG. 1 compares the thrombolytic dose-effect range of thrombolytically active protein BM 06.022 and r-tPA
(Alteplase), following an intravenous injection extending over one minute, in a dog model where the animals have coronary thrombosis. Reperfusion rate (o) is given as the percent of reperfused dogs with a dosage group (each group contains 6 dogs). The curves were produced using semi-logarithmic regression analysis.
FIG. 2 compares plasma concentration time curves, pharmacokinetically, using BM 06.022 and rt-PA "Alteplase"
in anesthetized doges. Either 140 KU/kg (1 mg/kg) of Alteplase were used, in groups of six dogs. What is presented is the arithmetic mean value. Starting concentration was subtracted from subsequent measurements.
- 6a -FIG. 3 displays time course of coronary blood flow after a single intravenous bolus injection, using either 140 or 280 KU/kg of BM 06.022, at time point t=0 minutes.
Data are shown in mean values ±SEM, where four dogs were used in each group. "Pre" indicates initial blood flow.
FIG. 4 also shows time course of coronary blood flow as in FIG. 3, (single bolus, t=o), and double bolus (140 KU/kg each bolus, at t=0 and t=44 minutes).
FIG. 5 shows time course of coronary blood flow before and after double bolus administration (140 KU/kg each bolus, for six dogs), or of 1.40 and 50 KU/kg administrations (for five dogs), of BM 06.022 in both cases, again at t=0 and t=44 minutes. Data are calculated and presented as explained in FIG. 3.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
In the meantime of the present invention, the term "fractionated administration" means the administration of a therapeutically effective amount of a thrombolytically active protein in two or more partial amounts in the form of a double or multiple bolus, "therapeutically effective amount" referring to sufficient protein to inhibit reocclusion.
Bolus administration is an intravenous rapid injection and is, therefore, especially advantageous because the time from the beginning of the clinical symptoms up to treatment and, also the time from the commencement of treatment up to the dissolving of the coronary thrombus is shortened. Thus, more myocardial tissue can be rescued from irreversible destruction. The double or multiple bolus administration according to the invention brings about a higher thrombolytic potency of the thrombolytically active protein use. In this way, it is possible to reduce dosages as ~ ~. , . u..~~. .. ~ ~ . . ,,.~ ,. .-,m.~ . ._ . ...... - . _ .-..~.. .-.m_ - ---. _ õ ~
- 6b -compared to rt-PA. Surprisingly, after bolus injection, more rapid reperfusion as compared to administration in the form of an infusion is found. Furthermore, double or multiple bolus administration brings about a significant prolongation of the cumulative patency time (sum of the time intervals after reperfusion in which coronary blood flow is present), as well as a ~. *
distinct increase of the coronary blood flow, whereby this also remains stable at a relatively high level for a longer time after the administration. Further-more, the double or multiple bolus injection has the surprising advantage of a smaller decrease of the plasmd.fibrinogen in contradistinction to a single bolus injection However, these advantageous properties of a double or multiple bolus administration are not achieved when the administration of the corresponding total amount of the plasminogen activator took place by a single bolus injection.
Thromboembolic diseases in the meaning of the present invention are especially those diseases which are to be attributed to a myocardial infarction or to a reocciusion of the coronary arteriese whereby re-occl:usions occur above all in the case of the use of thrombolytics for the treatment of the heart infarct.
The cause for a myocardial infarction is the formation of a thrombus in the coronary arteries. This thrombus consists of a combination of .'-~fibrin and thrombocytes.
TO&primar9= aim in the case of the treatmant of the heart infarct is the rapid dissolution of this throinbus and the recreation of the blood flow (reperf'usion).
A successful therapy must dissolve the thrombus as quickly as possible and prevent its reformation (reocclusion). The double or multiple bolus admin-istration according to the invention is especially advantageous for the treatment of the brain stroke.
21074"16 The pharmaceutical packaging unit according to the invention consists, on the one hand, of a form of administration which contains the plasminogen activator and, on the other hand, of an instruction, for example in the form of a package leaflet prescribed for medicaments, from which it follows that the administration of a therapeutically effective amount of the plasminogen activator advantageously takes place by double or multiple bolus administration. This information regarding the manner of the use can also be given as packaging overprint on the medicinal prep.arati.on ready for sale or can be taken from an information leaflet which can be brought together with medicinal preparations which contain the plasminogen activators in the meaning of the present invehtiona As suitable forms of administration for plasminogen activators, from the prior art there come into question known galenical formulations, for example lyophilisates or solutions in appropriate containers, such as e.g. in ampoules. As a rule, these formulations contain usual pharmaceutical adjuvants which are suitable for the prepaxation of isotonic solutionsf and possibly further stabilising and solubilising agents.
As proteins with plasminogen activator-like action (in the meaning of the present invention also briefly called "plasminogen activators"), those proteins are used which, in comparison with t-PA, possess a prolonged half life time. Plasminogen activators bring about the dissolving of thrombi and thereby make possible that blood again flows through the blood vessels concerned. As already mentioned above, the half life of t-PA in the human blood is about 3 - 6 minutes. In the meaning of the present invention, those plasminogen activators preferably come into question which, in comparison with t-PA, possess a half life prolonged by at least the factor 2, preferably 3 - 7 and especially 3 - 5. In particular, the half life should amount to at least about 10 minutes, for example 10 - 90, 10 - 40 or 10 - 20 minutes. Those proteins are preferably employed, the half life time of which is longer than the half life time of t-PA by the factor 3 - 30, especially 5 - 20 times.
In the meaning of the present invention, as proteins with plasminogen activatoralike action, there come into question, for example, the following plasminogen activatorsc LY 210825 (.K2P from "Syrian hamster cell 1ine:=" , Cira.I.990, 82, 930-940) ;-DeltaMx and-DeltaFElx (K1K2P from "Chinese hamster ovarycelisn, B1ood,'1988; 71, 216-219); DeltaFEKl (K2P from "mouse C127 cells", J. Cardibvasc. Pharmacol., 1990, 16, 197-203); E-6010 (Jap. Circ. J., 1989, 53, p. 918); t-PA variants.(Thromb. Haemost,, 1989, 62, P. 542); K2P and D-K2P (Thromb. Haemost., 1989, 62,.
p. 393); MB-1018 (FK2K2P, Thromb. Haemost., 1989, 62, -lc-pe 543); FK2P (FASEB J., 1989, 3, A1031, abstract 4791); Deltalx (Circulation, 1988, 4, 11-15);
i{1K2P (Thromb. Res., 1988, 5C, 33-41); FK1K2P
(J. Bio1. Chem., 1988, 263, 1599-1602). There are especially used recombinant plasminogen activators of the type K2P, such as e.g. BM 06.022 (known from EP-A-0 382 174). Further t-PA muteins of this kind are described in the following Patent Applications:
EP-A-o,J96,920, EP-A-0,207,589; AU 61804/86;
EP-A-0,231,624; EP-A- 0,289,508; JP 63133988;
EP-A-0,234,051; EP-A-0,263,172; EP-A-0,241,208;
EP-A-0,292,009; EP-A-0,297,066; EP-A-09302,4569 EP-A-0,379,890.
The pharmabbki.tietic profile of rt-PA (A1teplase) 15. is typically characterised by a two or even three compartment model (Thromb. Haemost., 1989, 61, 497-501), The most important-is thereby'the,initial half life since, with 66%, it contributes to the total area under the curve (area under the curve = AUC). Therefore, this half time is designated as the dominant half life. Tn the case of plasmi.nogen activators der.ived from t-PA
with prolonged half life, the pharmacokinetic propi1e is -naturally also changed. The designation "prolonged half life" thereby refers to the dominant half life, which is characteristic for the pharmacokinetic profile because it contributes quantitatively the most to the total AUC. The determination of the half life takes place accordin; to the processes known frnm the prior art (Pharmacokinetics, Ch. 2, Viarcel Dekker, New York, 1982).
As plasminogen activators, those especially derivatives of t-PA produced by recombinant DNA technology come into question which essentially include those protein repions of the natural protein which are responsible for the fibrinolysis of the thrombi. Those derivatives I0 of t-PA can thereby also be-used which display deletions or substitutions of individual or several amino acids in the sequence of the t-PA as long as the half life of such derivatives is prolonged in the above meaning with regard to t-PA. These are especially t-PA derivatives of the type K2P.
By way of example for the plasminogen activators coming into question in the meaning of the present invention, there was used the plasminogen activator K2P (BM 06.002) described in more detail in European Patent Application EP4A-0,382,174. It consists of the kringle 2(K) and -p~rotiease (P) domains of human t-PA and, because of its expression in Escherichia coli cells, is present in unglycosilated form. The specific activity of K2P amounts to 550,000 +/-IU/mg (= 550 +/- 200 KU/mg). K2P is used for the thrombolytic therapy of thromboembolic diseases (myocardial infarction, lung embolism, stroke and other occlusive vessel diseases) ':'tie missin~ ~1;~c osilatic~n leads ;, p a reduced clearance and results in a half life prolonged iith ra:;a~d to t-PA by at least the factor 3- 4.
The prolonged half life makes possible the use rfl the substance as clinically desirable multiple bolus administration. The thrombolytic effect of the bolus administration of the substance was investigated in various animal models. There was thereby shown a thrombolytic effectiveness increased approximately by the factor 5 compared with Alteplase. Especially impressive was thereby the rapid opening of thrombotically occluded vessels, in comparsion with Alteplase and other thrombolytics, which means a reduced time for the blood vessel opening in comparison with other thrombolytics and thus a more rapid reperfusion of the infarcted myocardial tissue.
On the basis of the improved pharmacokinetic properties of K2P, the effective dose to be admin-istered can be reduced and K2P can be administered as i.vs multiple bolus injection. A very rapid maximum reperl'nsion is therewith achinved. Furthermore, by the administration of a double or multiple bolus, the impairment of perfusion in the coronary arteries after reperfusion (tendency to reocclusion) can be significantly reduced or prevented. In addition, after double or multiple boli, the decrease of the plasma fibrinogen is smaller than after the single bolus 210 7d 7 G
-lj-in,jection nf the same total dose.
Besides the rapid opening of thrombotically occluded blood vessels, the prevention of the rencclusion of the npened binod vessels is of ~reat clinical importance since a reocclusion of the infarct blood vessel can lead to a clinically important reinfarct with the clinical complications resulting therefrom, The administration of K2P as bolus in equi-effective dose in comparison to the infusion of Alteplase leads, in the animal model, to a surprisingly rapid opening of the infarct blood vessel and thus makes possible a rapid reperfusion of the infarcted mycocardial tissue. In view of the present 1.5 knowledge regarding thrombolysis, this observed effect is to be evaluated very positively from the clinical point of va.ew.
The problem of the reocclusion of thrombolysed infarct blood vessels occurd, according to the atiimal ox perimental data, also with the single bolus of K2P
to an extent which is compaxable. wi.th other tbrombo-lytics Surprisingly, it is shown that a dividing up of the total dose of K2P into two or more chrono-logically staggered injected boli reduces or even prevents the appearance of perfusion impair-ments in the coronary arteries after reperfusion.
The time integral between beginning of the first and a subsequently following bolus injection is 2107417b indication-dependent and can amrun't to between 1.C
minutes and 6 hours, preferably 20 minutes and 2 - 3 hours, especially 30 - 9G minutes. The injection period is itself relatively short and amoun-us, dependinb upon the volume to be administered, to about 0 5 - 3 minutes, whereby an injection rate of about 5 - 10 ml/min, is advantageous. In the meaning of the present invention, by the term multiple balus administration one also understands the cumulated administration of two or more individual bolus injections which are administered in the scope of an interval therapy. Two or more, preferably three, bolus injections are carried out daily, whereby the therapy can eictend over the course of several successive days.
This treatment scheme can possibly also be interrupted 1+
by one or two days so that the administration in the form of bolus injections can take place, for example, on every second day during the whole treatment period Essentially, this -cumulated adn3nistratieu of single boli difi'e.rs from the known single bolus administration in that the administration is repeated daily several times-o If tha time interval between the first and a subsequently following bolus injection is relatively short, about 10 minutes up to one hour,one speaks of a double bolus or multiple bolus administration. If the time intervals are of the order of magnitude of several hours,then one can also speak of a cumulated administration of single boli.
210'7476 _15_ Fnr the production of the pharmaceutic3l forms of administration with regard to the prrtein with plasminogen activator-like action, there are used the usual pharmaceutical adjuvant and additive materia3.s Furthermore, there can be added thereto stabilising or the known solubilising agents, such as e.g. basic amino acids (arginine, lysine or ornithine).
Suitable galenical forms of administration are known from the prior art or can be produced according to the generally usual methods (cf. EP 0,217,379;
EP 0,228,862; WO 91/08763; WO 91/08764; WO 91/08765;
W0 91/08766; WO 91/08767 or WO 90/01334. The form of administration ca-n be present in lyophilised form or as injection solution ready for use. The bolus injection can take place intravenously, intramuscularly or also subcutaneously, whereby the i.v. injection is preferred.
In the case_ of the production of the pharma-ceutical packaging unit, the amounts of the protein to be admini.stsred for the two or multiple bolus injection can be made available in one or more, preferably in two separate containers, such as eog.
ampoules, It is aIlso possible to make available a multidose form of administration from which the particularly desired amount of the protein can be removed by withdrawal into a suitable'syringe. The ampoules can contain the solution already ready for injection or, alternatively, a lyophilisate oz the protein and possibly further pharmaceutically usual adjuvant or carrier materials. The lyophilisates are i-nixed with usual injection solutions shortly before the administration so that a solution results which can be administered directl.y.
The amount of the protein with plasminogen activator-like action can be the same or also different in the first and second container, depending upon the particular requirements. As a rule, amounts of 3 - 50 MU per container are used, For the first bolus injection, there is preferably administered a larger amount of the protein than for the second bolus injection. In particular, amounts of 5 - 20 MU, especially of 5 - 15 MU, are used for the first and 3 - 15 MU, especiall9 3- 10 MU for the second bolus injection. Especially preferably, one uses about 10 MU f or the first.- and about 5 MU for t he second injections The cumulative dose preferably lies in the range of 15 - 40 MQw The invention is acpla.ined in more detail on the basis of the following Examples:
Example 1 For the acperiments, a dog model was used'for the simulation of the myocardial infarction. Adult beagles of both sexes were narcotised with barbiturate, intubated and artificially respirated. Arterial and venous catheters were applied in order to monitor the blood pressure, tc administer substances or to take blood samples. The chest was opened and the heart exposed. A short segment of the ramus circumf].exus of the arteria coronaria sinistra was isolated and prepared. Subsequently, the coronary artery was provided with the following instruments from proximal to distal: an electromagnetic flow probefor the measurement of the blood flow in the coronary artery, a stimulation electrode, an adjustable screw and a thread,. The tip of the stimulation electrode was passed through the wall of the coronary artery and so placed within the blood vessel that the needle tip came into contact with the inner surface of the blood vessel. The screw'Was so adjusted that 90% of the reactive hyperaemia were eliminitad as the result of a short-term interruption of perfusion of the coronary artery.
The thrombus in the coronary artery for the initiation of the myocardial infarction was formed according to the method originally-deaoribed by Romson at a1.
(Thromb. Res,., 1980; 17s= 841-853). The method was employed in the modUfied form of 'Shebusk.i at a1., (J. Pharmac-ol. Exp. Ther.., 1988; 246: 790-796). Via the. stimulation'electrode, a 150 microamVere direct current was applied to the coronary artery and maintained until the blood flow in the coronary artery fell to 0 ml/min and remained there for at least 3 minutes. Subsequently, the thrombus should age for 30 minuteso In this time, the animals were --heparinised in a dosage of lCu(, units/animal/hour.
30 minutes after thrombus fcrmation, the fibrino-lytic or the solvent was administered as a one-minute i.v. bolus injection to six animals per group.
BM 06sC22 was administered in four doses: 50, 100, 14C and 200 KU/kg. Alteplase was also given in four doses: 200, 800' 1130 and 160C KU/kg (= 2 mg/kg).
The specific activity of Alteplase amounted to 800,000 IU/mg (= 800 kU/mg). Plasma samples were obtained before and repeatedly after injection of the fibrinolytic in order to determine the concent-ration of the activity of BM 06.022 or of Alteplase according to the method of Verheijen et al. (Thromb.
Haemostas., 1982; 48: 266-269). A reperfusion was assumed if at least 33% of the initial blood flow in the coronary artery was again achieved. Accordingly, the time to reperfusion from the beginning of the injection up to the achievement of this blood flow was defined, In each of a further experimental group,,. BM 06,022 waa administered in a dosage of 140 KU/kg and ' Alteplase i.n a dosage of 800 KU/kg l nng/kg) intravenoualy over 90 minutes as continuous"'infusion (10% of the totaldosage as initial i.v. bolus). ' Figure 1 shows the dose-effect relationship for the reperfusion rate after bolus injection of BM 06.022 or of Alteplaseo A 1CC% reperfusion rate (6 out of 6 animals) was achieved with i.v. bolus 21~7 d 7 6 injection of 2CG ItiU/kg in the case of BM 06.022. In the case of the same dose, none of the animals injected with Alteplase reperfused. For the achievement of the same maximum effect, an injection of 1600 KU/kg (= 2 mg/kg) of Alteplase was necessary. This injection dose is approximately twice as high as the dose of Alteplase at present used in the clinic as infusion (about 1 mg/kg). The higher thrombolqtic potency of BM 06.022, as seen by the shift to the left of the dose effect curve, can be explained by the improved pharmacokinetic properties of BM 06.022=(see Fig. 2).
The total plasma clearance of BM C6.022 amounts to 4.4 + 0.4 and that of Alteplase to 20.4 2,0 ml/
min- x kg Thus the total plasma clearance (a measure for the removal of a substance from the plasma) of BM 06.022 is about 4.6 times slower than that of Alteplase. Furthermore, it was found that Altepl.ase, after infusion, achieves a twice as high reperfusion rate thaa after single bolus injection of the same dose (800 KU/kg). BM 06.022 in a dose of 140 KU/kg shows the same reperfusion rate (4 out of 6 aniinals) after injection and after infusion but the time to reperfusion was significantly shorter after injection (Table 1).
The results show that Alteplase displays more favourable thrombolytic success after infusion than after injection, 'Lhis finding agrees with the clinical _. practice in which Alteplase is usually administered ..2C-as infusion (Chesebro et al., Circulaticn, 1987; '~J6e 142-254) .'1'he slower elimination of BM C6.C122 in compara.snn with Alteplase from the plasma led to a hioher thrombolytic potency of BM C6.022. Not only is dosage reduction thereby possible but, after single i.v. bolus injection, surprisingly a more rapid reperfusion takes place than after infusion.
Example 2 For the experiments in Example 2, there was used 1C the same dog model of coronary artery thrombosis as in Example 1. However, in contradistinction to the experiments in Example 1, the thrombus ageing in the case of administration of the substances was one hour instead of only 30 minutes. BM 06.022 was administered either as a single i.v. bolus injection or as a double bolus injection, in each case over one minute. The single bolus dose amounted to 140 or 280 KU/kg. In both groups with the double bolus administration, the first bolus dose amounted to 140 KU/kg in each case, follow by the second bolus in a dose of 140 or 50 KU/kg of BM 06.022 (44 minutes later). Each of the four experimen~al groups consisted bf six dogs. Additional parameters for the evaluation were the maximum of the coronary blood flow which was measured after reperfusion and the coronary blood flow at termination of the experiment three hours after injection. Further-more, the cumulative patency time was calculated;
thereunder one understands the sum of the time 210"14 76 intervals after reperfusion in which a coronary blood flow was present. The animal model is so designed that typically after ~eperfusinn cyclic flow vari.ations occur with reocclusion. Nt the end of the experiment, the residual thrombus was removed and its wet weight measured.
In both groups with single bolus injection, the reperfusion rate amounted to four of six animals. In the double bolus group, it was higher and amounted to six of six or five of six animals (Table 2). The results shown in Table 3 for the cumulative patency time, the coronary blood flow and the residual thrombus weight show that a double bolus administration of BM 06.022 significantly prolongs the cumulative patency time, significantly increases the coronary blood flow, this also" remains significantly increased at the end of the experiment and the residual thrombus weight has-decreased significantly in comparison with the single bolus injection of 140 KU/kg. Figure 3 shows that an increase of the single bolus injection from 140 to 280 KU/kg cannot markedly improve the corftaxy, blood flow. 9tr4ngly cyclic flow variations are still observed'. Cn ih.e other hand=,. in Pigure 4 it can be seen that the double bolus administration of 140 and 140 KU/kg of BY, 06.022 clearly improves the coronary blood flow in comparison with the single bolus injection of 140 ItU/kg of BM 06,G22. The same effect on the coronary blood flow can also be achieved by "-he double bolus administration of 14C and 5(. KU/krg of 3i4I C6.L22 (Figure 5). As Table 4 shows, the dividing up of a total drse of 28C KU/kg into 140 and 140 KU/kg instead of the sin;;le bolus injection of the same total dose of Bi1 C6,c.22 prevents the significant decrease of the plasma fibrinogen.
The results show that the deterioration of the coronary blood flow after reperfusion, which in this model intrinsically occurs in simulation of patients with reocclusion tendency, cannot be prevented by the single bolus injection even at the higher dosage. On the other hand, the administration of the double bolus is surprisingly able not only to prolong the cumulative patency time but also quantitatively to increase the flow and to maintain it increased up to the end of the experiment. In addition, the double bolus injection has the surprisingly advantage of the lower decrease of the plasmafibrinogen in contradistinction to the single bolus injection of the same total dosage. The size of the second bolus in the double bolus administ-ration can thereby be made variable without limiting the successd Description of the Figures Figure 1 shows the thrombolytic dose-effect range of BM 06.022 and of Alteplase after I minute i.v, injection in dogs with coronary thrombosis. The reperfusion rate (%) signifies the percentage of reperfused dogs within a dosal-e group (n = 6 per dose). The dose effect curves were produced with the help of a semi-logarithmic re;;ression analysis.
The Alteplase group without reperfusinn (at 20U KU/kg) was excluded from the regression analysis.
Figure 2 shows the pharmacokinetic comparison of the plasma concentration time curves for the activity of BM 06.022 and Alteplase in anaesthetised dogs.
A one minute i.v, injection of 140 FCU/kg of BM C6.022 or of 800 KU/kg (= 1 mg/kg) of Alteplase,was administered to six dogs per substance. The data represent the arithmetic mean value. The starting concentration was substracted from the subsequently measured concent-rations.
Figure 3 shows the time-course of the coronary blood flow before and after a single i.v. bolus injection with 140 or 280 KU/kg of BM 06.022 at the time point t = 0 minutes. The data are mean values SEM; n = 4 reperf'used dogs per group. Pro represents the initial blood flow.
Figure 4 shows the time-course of the coronary blood flow before and af'ter a single i.v. bolus injection of 140 RU/kg of BM 06.022 *(n = 4) at the time point t = 0 minutes or of a double bolus injection of 140 and 140 KfJ/kg of BM 06.022 (n = 6) at the time point t= 0 and t = 44 minutes. The data are average values + SEM; Pre is the initial blood flow.
_?L}_ :r,isure 5 shorrs the course ~f time of the coronary blood flow befoi=e and after double bolus administration of 14C and 140 KU/kg (n = 6) or of 140 and 50 KU/kg of BM 06.022 (n = 5) at the time point t = 0 and t= 44 minutes. The data are avera;e values + Sai;
Pre is the initial blood flows
Claims (16)
1. Use of a plasminogen activator for the production of a pharmaceutical agent for extending the cumulative patency time in the treatment of a thromboembolic disease, said plasminogen activator being in the form of two or several bolus injections.
2. Use of a plasminogen activator according to claim 1, wherein the plasminogen activator has a half-life which is increased by at least a factor of two compared to tPA.
3. Use of a plasminogen activator according to claim 1 or 2, for the production of a pharmaceutical agent for avoiding reocclusion tendencies in the treatment of coronary occlusions.
4. Use of a plasminogen activator according to claim 1, 2 or 3, wherein the plasminogen activator is selected from the group consisting of recombinantly-produced plasminogen activators of type K2P, K1K2P, D-K2P, FK2K2P, FK2P, DELTA1x, FK1K2P and tPA variants.
5. Use of a plasminogen activator according to claim 4, wherein the plasminogen activator is of recombinantly-produced type K2P.
6. Use of a plasminogen activator according to claim 1, 2, 3, 4 or 5, wherein the amount of plasminogen activator per bolus injection is 3-50 MU.
7. Use of a plasminogen activator according to claim 1, 2, 3, 4, 5 or 6, wherein a first bolus injection has an amount of 5-20 MU and a second bolus injection has an amount of 3-15 MU.
8. Use of a plasminogen activator according to claim 1, 2, 3, 4, 5 or 6, wherein the bolus injections provide a total amount of plasminogen activator of 15-40 MU.
9. Use of a plasminogen activator according to claim 1, 2, 3, 4, 5, 6, 7 or 8, wherein the bolus injections are adapted for administration with a time period between a first and second bolus of 10 to 90 minutes.
10. A pharmaceutical packaging unit for extending the cumulative patency time in the treatment of a thromboembolic disease, comprising essentially a pharmaceutical form of administration of a plasminogen activator in two or several separate containers.
11. A packaging unit according to claim 10, wherein the plasminogen activator has a half-life which is increased by at least a factor of two compared to tPA.
12. A packaging unit according to claim 10 or 11, wherein the plasminogen activator is selected from the group of recombinantly-produced plasminogen activators of type K2P, K1K2P, D-K2P, FK2K2P, FK2P, DELTA1x, FK1K2P and tPA variants.
13. A packaging unit according to claim 12, wherein the plasminogen activator is of recombinantly-produced type K2P.
14. A packaging unit according to claim 10, 11, 12 or 13, wherein the containers contain the plasminogen activators in an amount of 3-50 MU.
15. A packaging unit according to claim 10, 11, 12, 13 or 14, wherein a first container contains the plasminogen activator in an amount of 5-20 MU and the second container contains the plasminogen activator in an amount of 3-15 MU.
16. A packaging unit according to claim 10, 11, 12, 13, 14 or 15, wherein the containers have a total amount of plasminogen activator of 15-40 MU.
Applications Claiming Priority (5)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE4112398 | 1991-04-16 | ||
| DEP4112398.0 | 1991-04-16 | ||
| DE19914123845 DE4123845A1 (en) | 1991-07-18 | 1991-07-18 | Compsn. for treating thromboembolic disease with plasminogen activator |
| DEP4123845.1 | 1991-07-18 | ||
| PCT/EP1992/000851 WO1992018157A1 (en) | 1991-04-16 | 1992-04-15 | Pharmaceutical packaging unit containing plasminogen activators for multiple-bolus administration |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2107476A1 CA2107476A1 (en) | 1992-10-17 |
| CA2107476C true CA2107476C (en) | 2007-12-18 |
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ID=25902885
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002107476A Expired - Lifetime CA2107476C (en) | 1991-04-16 | 1992-04-15 | Pharmaceutical packaging unit containing plasminogen activators for multiple bolus administration |
Country Status (11)
| Country | Link |
|---|---|
| US (3) | US5500411A (en) |
| EP (2) | EP0580685B1 (en) |
| JP (1) | JPH06506459A (en) |
| AT (1) | ATE132373T1 (en) |
| CA (1) | CA2107476C (en) |
| DE (1) | DE59204945D1 (en) |
| DK (1) | DK0580685T3 (en) |
| ES (1) | ES2082465T3 (en) |
| GR (1) | GR3018740T3 (en) |
| HK (1) | HK117096A (en) |
| WO (1) | WO1992018157A1 (en) |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0580685B1 (en) * | 1991-04-16 | 1996-01-03 | Roche Diagnostics GmbH | Pharmaceutical packaging unit containing plasminogen activators for multiple-bolus administration |
| US5510330A (en) † | 1994-03-25 | 1996-04-23 | Boehringer Mannheim Gmbh | Combinations of thrombolytically active proteins and non-heparin anticoagulants, and uses thereof. |
| AU3765897A (en) * | 1996-08-14 | 1998-03-06 | Jurgen Brodersen Gram | Use of a plasminogen activator for the treatment of pulmonary disorders |
| WO2002052006A1 (en) * | 2000-12-26 | 2002-07-04 | Genox Research, Inc. | Method of examining allergic disease |
| DE10153601A1 (en) * | 2001-11-02 | 2003-05-22 | Paion Gmbh | DSPA for the treatment of stroke |
| US7084118B2 (en) * | 2002-02-22 | 2006-08-01 | Genentech, Inc. | Combination treatment with t-PA variant and low molecular weight heparin |
| US20070142272A1 (en) * | 2003-01-24 | 2007-06-21 | Zlokovic Berislav V | Neuroprotective activity of activated protein c independent of its anticoagulant activity |
| US20100260669A1 (en) * | 2004-05-13 | 2010-10-14 | Anthony Joonkyoo Yun | Treatment of Seasonal Conditions Through Modulation of the Autonomic Nervous System |
| JP2008507561A (en) * | 2004-07-23 | 2008-03-13 | ザ ユニバーシティ オブ ロチェスター | Inhibition of adverse effects of plasminogen activator in the brain by activated protein C |
| US8309063B2 (en) * | 2005-06-10 | 2012-11-13 | Amcol International Corporation | Stable sunscreen compositions containing zinc oxide |
| TWI482628B (en) * | 2007-10-18 | 2015-05-01 | Lundbeck & Co As H | Novel patient subgroups for thrombolysis |
| WO2024060167A1 (en) * | 2022-09-23 | 2024-03-28 | 庄伟哲 | Fusion protein targeting intergrin alpha(iib)beta3 and containing tissue plasminogen activator or variant thereof and use thereof |
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|---|---|---|---|---|
| US5234686A (en) * | 1985-04-03 | 1993-08-10 | Beecham Group P.L.C. | Human tissue plasminogen activator consisting essentially of t-PA residues to 160 to 527, pharmaceutical compositions and methods of treatment |
| GB8508717D0 (en) * | 1985-04-03 | 1985-05-09 | Beecham Group Plc | Composition |
| ZW14486A1 (en) * | 1985-07-29 | 1986-10-22 | Smithkline Beckman Corp | Pharmaceutical dosage unit |
| US4935237A (en) * | 1988-03-21 | 1990-06-19 | Genentech, Inc. | Processes for the preparation of t-PA mutants |
| WO1989000197A1 (en) * | 1987-06-30 | 1989-01-12 | Genentech, Inc. | Improved processes for the treatment of vascular disease |
| US5084274A (en) * | 1987-11-17 | 1992-01-28 | Scripps Clinic And Research Foundation | Inhibition of arterial thrombotic occlusion or thromboembolism |
| GB8814604D0 (en) * | 1988-06-20 | 1988-07-27 | Wellcome Found | Medicaments |
| US5342616A (en) * | 1988-06-20 | 1994-08-30 | The Wellcome Foundation Limited | Method of administering tissue plasminogen activator |
| DK176140B1 (en) * | 1988-07-20 | 2006-09-25 | Schering Ag Patentabteilung | Bat salivary plasminogen activators |
| US4980165A (en) * | 1989-01-27 | 1990-12-25 | Genetics Institute, Inc. | Pharmaceutical formulations of plasminogen activator proteins |
| DE3903581A1 (en) * | 1989-02-07 | 1990-08-16 | Boehringer Mannheim Gmbh | FABRIC PLASMINOGEN ACTIVATOR DERIVATIVE |
| DE3942143A1 (en) * | 1989-12-20 | 1991-06-27 | Boehringer Mannheim Gmbh | T-PA PRO STABILIZATION |
| US5366730A (en) * | 1989-12-20 | 1994-11-22 | Boehringer Mannheim Gmbh | Stabilized compositions having human tissue type plasminogen activator enzymatic activity |
| DE3942141A1 (en) * | 1989-12-20 | 1991-06-27 | Boehringer Mannheim Gmbh | K2P PRO STABILIZATION |
| US5242688A (en) * | 1990-12-24 | 1993-09-07 | Eli Lilly And Company | Method of treating thromboembolic disorders by administration of diglycosylated t-pa variants |
| EP0580685B1 (en) * | 1991-04-16 | 1996-01-03 | Roche Diagnostics GmbH | Pharmaceutical packaging unit containing plasminogen activators for multiple-bolus administration |
-
1992
- 1992-04-15 EP EP92908637A patent/EP0580685B1/en not_active Revoked
- 1992-04-15 US US08/137,116 patent/US5500411A/en not_active Expired - Lifetime
- 1992-04-15 DE DE59204945T patent/DE59204945D1/en not_active Revoked
- 1992-04-15 AT AT92908637T patent/ATE132373T1/en not_active IP Right Cessation
- 1992-04-15 DK DK92908637.9T patent/DK0580685T3/en active
- 1992-04-15 JP JP4507994A patent/JPH06506459A/en active Pending
- 1992-04-15 WO PCT/EP1992/000851 patent/WO1992018157A1/en not_active Application Discontinuation
- 1992-04-15 ES ES92908637T patent/ES2082465T3/en not_active Expired - Lifetime
- 1992-04-15 EP EP95106049A patent/EP0709096A2/en not_active Withdrawn
- 1992-04-15 CA CA002107476A patent/CA2107476C/en not_active Expired - Lifetime
-
1994
- 1994-03-25 US US08/217,616 patent/US5690931A/en not_active Expired - Lifetime
-
1996
- 1996-01-19 GR GR960400126T patent/GR3018740T3/en unknown
- 1996-07-04 HK HK117096A patent/HK117096A/en not_active IP Right Cessation
-
1997
- 1997-02-03 US US08/794,528 patent/US6235279B1/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| WO1992018157A1 (en) | 1992-10-29 |
| US6235279B1 (en) | 2001-05-22 |
| JPH06506459A (en) | 1994-07-21 |
| HK117096A (en) | 1996-07-12 |
| ATE132373T1 (en) | 1996-01-15 |
| EP0580685B1 (en) | 1996-01-03 |
| CA2107476A1 (en) | 1992-10-17 |
| US5500411A (en) | 1996-03-19 |
| ES2082465T3 (en) | 1996-03-16 |
| DK0580685T3 (en) | 1996-03-04 |
| EP0709096A2 (en) | 1996-05-01 |
| GR3018740T3 (en) | 1996-04-30 |
| DE59204945D1 (en) | 1996-02-15 |
| US5690931A (en) | 1997-11-25 |
| EP0709096A3 (en) | 1996-05-08 |
| EP0580685A1 (en) | 1994-02-02 |
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