CA1246449A - Drug kit or drug composition for ischaemic damage - Google Patents

Abstract

ABSTRACT

A drug kit or drug composition for use in preventing and treating ischaemic cell damage comprises:

a) at least one plasma volume expander;
b) at least one low molecular, physiologically acceptable hydroxyl radical scavenger;
c) at least one physiologically acceptable and water-soluble magnesium salt; and d) at least one organic compound active as a calcium blocking agent dissolved in a carrier, either per se or in one or several combinations.

Description

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A drug kit or drug composition The present invention relates to a drug kit or drug composition for use in preventing and treating ischaemic cell damage.

When circulation of the blood collapses and ischaemia occurs in peripheral body organs, particularly in the brain, a large number of pathophysiological changes take place. In present clinical practice it is only possible to treat measurable pathophysiological changes, for example changes in blood volume, impaired-cardiac function, central acidosis, etc.
In such cases each change has been treated individually and it can be sP~id'generally that present day therapy for the resuscitation of an organ is mainly directed towards re-establishing blood circulation.

The present invention is based on the concept that incurabletissue damage can be caused as a result of unfavourable con-ditions created when re-establishing the blood circulation to a body organ.

According to one aspect of this concept the transportation of calcium into and out of,a cell is of great significance. The transportation of calcium into and out of a cell normally takes place while maintaining externally of the cell a calcium concentration which is 1000 times greater than the calcium concentration inside the cell. When a deficiency in energy occurs as a result of ischaemia, the calcium gradient cann~t be maintained, and calcium will consequently leak into the cell.
Calcium is taken up in the cell by the mitochondria, resulting in serious disturbances in energy production. Wh~n blood again starts to flow, calcium will enter the cell in still greater quantities, while transportation of calcium from the cell is impaired due to the fact that the build-up of energy in the cell is inhibited by the high calcium content thereof. This greatly increases the load on the mitochondria, which can lead to incurable cell damage and cell death.

6~9 According to another aspect of the concept there occurs during the ischaemic period a gathering of degradation products, such as hypoxanthine, which when oxygen is supplied in connec-tion with the re-establishment of circul~tion are converted by certain enzymes, such as xanthine oxidase, in processes which produce free hydroxyl radicals as a secondary product, possibly via O2-radicals. Those enzyme systems which protect the tissue from the hydroxyl radicals are not able herewith to deal with the radicals at the rate in which they are formed, which can lead to damage of blood vessels for example.

On the basis of these concepts concerning incurable tissue damage, there is now provided a drug kit or drug composition which provides a better result and enables persons who are subjected to the risk of ischaemic cell damage to be treated in a simplified manner.

The drug kit or drug composition according to the invention is characterized in that it comprises a) at least one plasma volume expander;
b) at least one low molecular, physiologically - acceptable hydroxyl radical scavenger;
c) at Ieast one physiologically acceptable and water soluble magnesium salt; and d) at least one calcium blocking organic compound dissolved in a carrier, either per se or in one or more com-bina~ions.

The invention is described hereinafter with reference to a drug kit intended for-single-unit administration, although the invention also relates to-different stock solutions which might come into question.

The plasma volume expander used may be a physiologically acceptable high molecular substance known per se in the expan-sion of blood plasma volume. These substances have an average molecular weight Mw (weight average value) which is higher than 10,000 Daltons, e.g. higher than 15,000 and preferably higher ~an 30,000 and lower than 400,000 and preferably 3 ~-z,~

lower than 300,000 Daltons. It is well known in the art that the average molecular weight Mw chosen depends on the high molecular substance used. Examples of such plasma expanders ' are plasma-albumin and substances based on dextran, starch derivatives or gelatine derivatives. The dextran products normally have an average molecular weight Mw within the range of 30,000 to 80,000 Daltons. Examples of starch derivatives for this purpose include hydroxyethyl starch ha~ing an average molecular weight Mw within the range of 40,000 - 400,000 Daltons, e.g. in the order of 200,000 Daltons. A number of different gelatine derivatives of varying average molecular weights ~ are also used for this purpose. (A review of some plasma volume expanders is found, for example, in the book "Blood Replacement" by U.F. Gruber, Springer Verlag, Berlin-Heidelberg-New Yo~k 1969). Of these plasma volume expanders, those based on dextran are primarily preferred.

The concentration of plasma volume expander in the solution in which it is present-is'chosen so that subse~uent to being optionally mixed wi~h one or more solutions incorporated in the kit, the soiution injected into the patient will have a plasma-volume-expander concentration which is normal in the use of the substance in question. The plasma volume expander solution of the invention usu~lly has a concentration of 1-15 g/
100 ml, su-ch as'2-12 g/100 ml, for exam~le 3-10 g/100 ml. ' ' A common requirement of the hydroxyl radical scavengers which can be used in accordance with the invention is that they are physiologically acceptable and have a molecular weight beneath 10,000 Daltons, preferably beneath 1,000 Daltons.
Hydroxyl radical scavengers which have a molecular weight above 10,000 Daltons as a rule have a poor effect. A suitable hydroxyl radical scavenger is soluble in water at physio-logical pH and ion strengths. It normally includes a func-tional structure selected from-~aromatic o~ aliphatic thiol (-SH~, alcoholic and phenolic hydroxyl t-oH) and nitrogen-containing structures, such as primary amine ! -NH2) secondary amine l-NH-) and imine (=NH). The hydroxyl radical scavenger is advantageously selected from the group comprising physio-logically acceptable sugar alcohols, monosaccharides, oligo-saccharides, amino acids which contain mercapto groups, and methionine and histidine. Among the group of sugar alcohols, mannitol -is the primary choice, because it is able to function simultaneously as a diuretic and an anti-oedema agent. Other sugar alcohols of interest in this context are sorbitol and xylitol. Examples of physiologically acceptable monosaccha-rides are glucose and fructose, and of oligosaccharides malto-oligosaccharides and isomalto-oligosaccharides (which can be obtained by means o partial hydrolysis of starch and dextran respectively), e.g. maltose. Cysteine is an example of amino acids which contain mercapto groups.

The hydroxyl scavenger used is preferably a combination of at least one sugar alcohol and at least one amino acid according to the above, particularly a combination of mannitol and L-methionine, or of mannitol, L-methionine and histidine.

The concentration of hydroxyl radical scavenger is determined by-the specific substance in question and by the amount it is desired to administer. It is always so high as to enable a therapeutically active ~uantity to be administered when the kit is used. The drug kit or drug composition according to the invention may thus contain from 1 g up to 150 g hydroxyl radical scavenger~ The range of 1-10 g is particularly appli-cable in the case of methionine and histidine and a range of 5-150 g in the case of-sugar alcohols, calculated per occasion of treatment.
Magnesium salt present in the compQsition comprises one or more salts from the group water-soluble, pharmaceutically acceptable magnesium salts. Examples of magnesium salts which are thus comtemplated are magnesium sulphate and magnesium chloride. ~a:gnesium chloride is particularly preferred. Water-soluble magnesium salts are present in the composition according to the invention in quantities corresponding to 5-100 mmol Mg2+, calculted per occasion of treatment.

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The organic compounds acting as calcium blockers are normally of low molecular weight, with a molecular weight beneath 2000 Daltons. They are deined by their ahility to prevent the migra~ion of calcium ions into cells. Cf. "Calcium Blockers" (edited by Flaim~ S.F. et al; Urban and Scharzen-berg. Baltimore-Munich, 1983). The compounds in question may be of highly different structure, niphedipine, nimodipine, verapamil, diltiazem, lidoflazine, flunarazine and analogous compounds can be mentioned by way of example. The calcium blockers used in accordance with the invention may be soluble in water and/or in fat. Verapamil(5- ~(3,4-dimethoxyphenylethyl) methylamino~ -2-(3,4-dimethoxyphenyl)-2-isopropylvaleronitrile) is an example of a water-soluble calcium blocker, while an example of a fat-soluble calcium blocker is lidoflazine(4-[4,4-bis(4-fluorophenyl)butyl~ -N-(2,6-dimethylphenyl)-1-piper-azine acetamide). When a fat soluble Galcium blocker is used in accordance with the invention, it is advantageously in-cluded in the kit as a component separate from the plasma volume expander. According to one aspect of the invention, this enables lidoflazine to be administered in a separate in-jection when using the drug kit. In this variant of the in-vention, the fat-soluble calcium klocker may be dissolved in, for example,:

I. A mixture of water and ethanol in an amount which is physiologically acceptable for the purpose. When the calcium blocker has the nature of an amine, the mixture can be acidified, to increase solubility. It is essential in this respect-that acidification of the mixture is adapted to the pH and buffer capacity o~ the remaining kit components to be used on the occasion-of the treatment. The mixture is advantageously acidified with aceti~ acid, hydrochloric acid, or some other physiologically acceptable acid. The mixture may also contain glycerol.
II. Physiologically acceptable fat emulsions used for paren-teral nutrition (a number of such emulsions are described, inter alia, in patent literature; cf. for example the U.S.

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Patent Specification No. 4,168,308).

A usable p~oduct in this connection is retailed under the name Intralipid~by Apoteksvarucentralen Vitrurn AB, Stockholm, Sweden. This product contains fractionated soya oil in an amount of 100 or 200 mg/ml, fractionated egg-phospholipides (as stabilizer) in an amount of 12 mg/ml, and glycerol in an amount of'25 mg/ml, with the remainder sterile water.

III. Physiologically acceptable emulsions of fluorinated hydrocarbons, which are administered parenterally due to their ability to dissolve and transport oxygen gas.

The amount of-calcium blocker included in the kit varies from substance to substance. Calculated per occasion of treatment it is normally included in amounts of from 1 to 300 mg; a particular value for lidoflazin0 is from 10 to 200 mg.

The carrier or vehicle in which the active kit somponents can be dissolved is physiologically acceptable and contains water. It may optionally be buffered'with a physiologically acceptable bu'ffer substance to a pH-value and an ion strength such that the total efect of that inténded to be administered is physiologically acceptable. This means that in the selection of a suitable-buffer systemj attention is paid to all compo-nents included in ~e kit or the composition according to the invention. Examples of buffer systems include trometamol buffers, carbonate buffers,' phosphate buffers, histidine buffers, acetate buffers and combinations thereof. According to the invention a buffer system may be included as a solutlon separate from the solution containing the plasma volume expander, hydroxyl radical scavenger, magnesium salt. A se-- parate buffer system shall be used when acidose is present.
It shall be capable of restoring the blood of the patient in question to a pH-value of from 7.0 to 8.0, preferably the physiological pH-value 7.4. The buffer capacity lies in the region of 25-300 mmol, preferably 50-200 mmol. In practice this means that a separate buffer system shall have a pH-value 7 ~ 2~6~

in the range of 7.0 - 10.0, preferably 7.4 - 9.2.

The drug kit or composition according to the invention pre-ferably also include's a diuretic agent, particularly an osmotic diuretic agent, and/or an anti-oedema substance.
Since in addition to being an hydroxyl radical scavenger, mannitol is also able to fulfil the function of both a diuretic and an anti-oedema substance, mannitol is a pre-ferred substance in the present context. Sorbitol or gly-cerol can be used as a diuretic agent, either instead of ortogether with mannitol. The quantities in which a diuretic agent and anti-oedema substance is used is dependent on the substance utilized, and may thus vary within wide limits.
In the case o~ an osmotic diuretic agent, the quantities used may lie within the range 5-150 g, otherwise 0.1-200 mg. In the case of the anti-oedema substance a corresponding range may be 5-150 g.

It may also be of advantage~to incorporate in the kit or the composition according to the invention an xanthine oxidase inhibitor, such as allopurinol for example, (50 mg - 5 g, de-pending on which is chosen), and/or a superoxide radical scavenger, such as superoxide dismutase for example, and/or an hydrogen peroxide inactivator,such as catalase for example, and/or a substance which binds iron in a solid complex, such as desferrioxamine or diethylenetriamine-pentaacetic acid or ethylenediamine-di(o-hydroxyphenylacetic acid)~ or a phytic acid derivative.

The quantities quoted above in respect of the diuretic agent, anti-oedema substance and-xanthine-oxidase inhibitor apply to each occasion of treatment.

The active components included in the drug kit or drug compo-sition are present in the form of a single solution or a plu-rality of solutions. Precisely how they are combined is de-termined, inter alia, on the grounds of solubility and sta-bility, even though for practical reasons the aim is to place ~z~

them in a common solution. For example, in accordance ~7ith one advantageous variant, the plasma volume expander hydroxyl radical scavenger, magnesium salt and calcium blocker are selected so as to be compatible with one another in solubilized form and with the desired pH-value of the solution to be administered. Similar considerations are applicable to remaining active components such as the anti-oedema substance, diuretic agent, xanthine-oxidase inhibitor, superoxide radical scavenger, hydrogen peroxide inactivator and iron-binding substance.

On the basis of those studies carried out hitherto, the embodiment most preferred has a solution (A) which contains plasma volume expander, hydroxyl radical scavenger and magnesium salt; a solution (B) which contains a buffer system and a solution or dispersion (C) which contains a fat-soluble calcium blocker. In this embodiment, the remaining active components are placed in one of the solutions A, B or C. For example, if allopurinol is chosen as the xanthine-oxidase inhibitor, it can be added to the buffer solution B for reasons of solubility. If the kit does not include such a solution, it may be necessary to choose another xanthine-oxidase inhibitor.

The various solutions included in a drug kit according to the inven-tion (and in certain cases the dispersion of organic calcium blocker) may have the form of sterile storage solutions from which a suitable quantity of the separate solutions or dispersion is taken on each treatment occasion; preferably, however, the kit is made up with dosages suited to the purpose, each dosage containing therapeutically active quantities of the substances in question. In this latter case a solution (A) according to the aforegoing can be packed into units of 100-1000 ml, normally 500 ml, a solution (B) packed in units of 10-100 ml, preferably 25-100 ml, and a solution or dispersion (C) packed in units of 5-50 ml, preferably 10-30 ml. The units can be poured into plastic sachets, glass or plastic bottles, ampoules, syringes etc..
The exact choice varies from case to case, and is determined, in~er alia, by practical considerations. It can be mentioned by way of example that the solution C is advantageously placed in an ampoule or disposable . . . . . . . . . . . . . . . . . . . . . . . . .

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syringe.

The concentration in which the active components are present are selected so as to maintain the mutual proportions between the aforementioned quantities. In the aforesaid preferred embodiments, the concentration of hydroxyl radical scavenger and magnesium salt corresponds to the aforementioned quantities per 500 ml of solution. The same applies to the calcium blocker, when it is incorporated in the same solution as these two substances. When it is present in a separate solution (C), the calcium blocker concentration may be from 10 to 100 times greater than in the previous case, due among other things to the solubility conditions.

When calculated on the basis of a patient weighing 70 kg, the kit components are normally administered to the patient in a total solution volume of 500-600 ml.

When using a drug kit according to the invention in which the calcium blocker is included as a separate unit (C), this unit is the first to be injected into the patient. It is desirable that this injection can be given relatively quickly. In those cases where métabolic acidose prevails, as with a cardiac arrest for example, solution (B3 i5 used to corrent the pH of the patient. The solution (B) may be mixed with the solution (A) immediately or shortly before being used. The mixture, or the solutions (A) and (B) each per se, is or are then adminis-tered to the patient as soon as possible after having injectedthat patient with (C). In the absence of metabolic acidose, only solution (A) is administered.

When using a drug composition according to the invention in which a plasma volume expander, an hydroxyl radical scavenger, magnesium salt and a calcium blocker are present in a common solution separate from a buffer solution (B), this common solu-tion is injected into the patient separately or in mixture with (B). The solution (B) is only used in the case of metabolic acidose.

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The drug kit accordiny to the invention is intended for use primarily in acute resuscitation, such as in the event of a cardiac arrest or in other situations in which blood circulation collapses and ~he brain is subjected to ischae-mia. The drug kit can also be used in various kinds oftrauma in the central ner~ous system, cerebral haemorrhage, apoplectic strokesl su~arachnoidal bleeding, or in the case of intracranial vessel surgery, where blood vessels must be temporarily closed. The drug kit can also be used with ischaemic conditions in other body organs, such as the heart, kidneyst intestines and skeleton muscle, in conjunction with shock, trauma, embolies and heart attacks, and also in sur-gical operations, such-as heart surgery, vessel reconstruction and organ transplantation.
The drug kit can also be used as a~perfusion solution and preserving solution for body organs in, for example, cardioplegy or-organ transplantation.

The invention also relates to a-method of treating the afore-said conditions. In such a treatment the components of the kit are administered in any of the ways described above.

The invention will now be described with reference to a number o working examples.

EXEMPLE
Preparation of a drug kit Solution A
15 g dextran having an average molecular weight (Mw) o~ about 60 000, 4.0 g MgCl2 (anhydrous), 25 g mannitol, 5 g L-methio-nine, and 5 g L-histidine were dissolved in 500 ml distilled water. The resultant solution was sterilized by sterile fil-tration and poured into a 500 ml sterile plastics bag, whichwas then sealed under aseptic conditions.

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Solution B
There were used for this solution 50 ml of a conventional commercial buffer solution having a pH of 9.2 and containing 20 g trometamol with a buffer capacity of 150 mmol (Adde THAM form Pharmacia Infusion AB, Uppsala, Sweden).

Solution C
-80 mg lidoflazine were dissolved in 1.0 g ethanol (99.5%), 0.1 g concentrated acetic acid and 1.5 g glycerol, and was diluted up to 10 ml with distilled water. The solution was sterilized by sterile filtration ana poured into a 1~ ml ampoule lmder aseptic conditions.

The solutions A, B and C were then packed in a box, as a unit.

Pharmacological tests The tests were carried out with a rat model, which gives an incomplete cerebral ischaemia with a cortical flow < 5% of the normal flow, and a flow in the brain stem which is about 30% of the normal flow.;This is effected by squeezing the two carotid arteries while simultaneously lowering the blood pressure to 50 mm Hgj by bleeding. The method has been de-~5 scribed by Nordstrom C.I-I. and Siesjo B.K., Stroke 9, 327-335 (1978).

Wistar-rats weighing 300-400 g and fasted overnight were used in the tests. The rats were anaesthetized with 4% Fluothane ~
(ICI-Pharma AB, Gothenburg, Sweden), 30% 02/70% N~O, intubated and connected to a respirator. The vena jugularis externa was uncovered. Celocurine ~5 mg/kg) was injected and a cathe-ter was placed in vena cava supe~ior. Catheters were also placed in the tail artery and in a tail vein for measuring blood pressure and infusion, respectively. EEG-electrodes were applied and finally 5 ml 0.9% NaCl were administered intra-peritoneally and 100 IU heparin~ intravenously. The supply of Fluothane ~ was cut-off, whereafter blood gases, pH and the sugar content of the blood were measured for a period of a-t least 30 minutes. It was endeavoured to obtain a pH in the region of 7.35 - 7.40, PCO2 Of 4.67 - 5.50 kPa, and PO2 of ' 11.0 - 18.0 kPa, and a bloodsugar content in the region 3.0 - 8.0 mmol/l. If these criteria were not attained, the animal was excluded.

The following procedure was undertaken in order to create ischaemia:

A solution of trimethaphan-D-cam~harsulphcnate in sterile water (15 mg/ml, Arfonad ~ from Hoffmann-La Roche & Co AG, Basle, Switzerland) was administered intravenously, until the average blood pressure was 80 mm Hg, whereafter the two carotid arteries of respective animals were shut-off and blood was drained from the animals through respective catheters in vena cava superior until an average blood pressure of 50 mm Hg was reached.

~he EEG was recorded contin~usly during this time period, and the ischaemic period was taken to commence when an isoelec-tric EEG was obtained. Subsequent to an ischaemic period of 10 mins, the infusion of lidoflazine in the treatment group was commenced. Of a total dosage of 1.0 mg in one ml of a ~5 physiological sodium ~h~or~db solution, half was administered during the ischaemia and the remainder after 5 minutes re-circulation. A corresponding volume of physiological sodium chloride solution was administered to a control group.
Infusion of an aqueous solution containing 3.5~ albumin, 10% mannitol, 2~ L-methionine, 92.2 mM`magnesium chloride and 0.5 M Tris (percentages given in w/v~, was commenced during the last two minutes of the ischaemic period and was continued for two minutes during the recirculation phase. A total of 2 ml were injected. The blood pressure 3~ was monitored during the infusion period and adjusted when necessary, by bleeding the animal or infusing blood there-into. The rats were left in the respirator until they began to waken, whereupon they were ventilated for two minutes with 100% axygen gas and the respiratQ~ then disconnected.
Tracheal tubes and oxygen masks were left in position until ~246449 stable breathing was observed.

Of 10 test animals in each group, the mortality of the con-trol group was 60%. The cor~esponding figure in the group treated with a drug kit according to the invention was 20%.
No significant differences were observed with regard to average arterial blood pressure, blood gas or blood sugar.
With regard to the pH of the blood, it was observed that the blood-pH of the animals in the group treated with a drug kit according to the invention fell after the ischaemic period to a lesser extent than that of the animals in the control group, this being attributed to the buffer capacity of the drug kit accordin~ to the invention.

Claims (11)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:

1. A drug kit or drug composition for use in preventing and treating ischaemic cell damage, characterized in that it contains:
a) at least one plasma volume expander;
b) at least one low molecular, physiologically accept-able hydroxyl radical scavenger;
c) at least one pbysiologically acceptable and water-soluble magnesium salt; and d) at least one organic compound active as a calcium blocking agent.
dissolved in a carrier, either per se or in one or several combinations.

2. A drug kit or composition according to Claim 1, charac-terized in that the plasma volume expander is plasma-albumin or is based on dextran, a starch derivative or gelatin derivative.

3. A drug kit or composition according to Claim 1, charac-terized in that the hydroxyl radical scavenger comprises one or more substances from the group physiologically acceptable sugar alcohols, monosaccharides, oligosaccharides, amino acids which contain mercapto groups, methionine and histidine.

4. A drug kit or composition according to Claim 1, charac-terized in that the magnesium salt is magnesium sulphate or magnesium chloride.

5. A drug kit or composition according to Claim 1, charac-terized in that the calcium blocker is lidoflazine.

6. A drug kit or composition according to Claim 1, charac-terized in that it also includes a diuretic agent and/or anti-oedema substance.

7. A drug kit or composition according to Claim 6, charac-terized in that the diuretic agent is mannitol and/or sorbitol.

8. A drug kit or composition according to claim 6, characterized in that the anti-oedema substance is mannitol.

9. A drug kit or composition according to either of claims 1 and 6, characterized in that it also includes an xanthine oxidase inhibitor and/or a superoxide radical scavenger and/or a hydrogen peroxide inactivator and/or an iron-binding substance.

10. A drug kit or composition according to either of claims 1 and 6, characterized in that it also includes a physiologically acceptable buffer system.

11. A drug kit or composition according to either of claims 1 and 6, characterized in that it further includes a physiologically acceptable buffer system and an xanthine oxidase inhibitor and/or a superoxide radical scavenger and/or a hydrogen peroxide inactivator and/or an iron-binding substance.