IE48130B1 - Synergistic mixtures of antibiotics and processes for their manufacture - Google Patents

Description

The present invention relates to new synergistic mixtures of antibiotic compounds having properties superior to those of the individual components, pharmaceutical preparations containing said mixtures and processes for their manufacture, and the use of said preparations for combating infections.

The present invention comprises a synergistic mixture of antibiotics consisting of cefsulodin sodium and a second antibiotic selected from the group consisting of clavulanic acid, penicillanic acid 4,4-dioxide, azlocillin, mezlocillin, piperacillin, mecillinam, cephacetril, cefoxitin, cefotiam, amikacin, dibekacin, tobramycin, sisomicin or gentamicin, or a salt thereof.

Cefsulodin sodium is the international non-proprietory name for the sodium salt of Ν-/7β-(α-sulfophenyiacetamido)ceph-3-em — 3-yl-methyl/r-4'-carbamoylpyridinium-4-carboxylate. The compound is known from German Offenlegungsschrift 2,234,280. Its antibiotic action is directed against Gram-negative cocci, Gram-positive cocci and bacteria, and especially against Pseudonomas strains, both carbenicillinsensitive and carbenicillin-resistant. Cefsulodin sodium - 3 has a relatively insignificant action against enterobacteria, such as Escherichia coli and Klebsiella pneumoniae, and anaerobei, for example Bacteroides.

The present invention is based on the surprising observation that the antibiotic action of said antibiotics and that of cefsulodin sodium, intensify in synergistic manner in the mixtures of the invention, and that in some cases there is even an increase in the tolerance to the individual components. This synergism has also surprisingly been observed in the case of enterobacteria, anaerobes and polyinfections.

The advantages of synergistic mixtures over the individual components of which they consist are well known and can be utilised in various ways. The advantages of the antibiotic mixtures of this invention reside both in a reduction of the dose while achieving the same effect (which means imposing less of a strain on the organism with foreign substances and consequently diminishing the side-effects) and in a broadening of the activity spectrum (which is of importance in the treatment of polyinfections, especially of secondary infections In Pseudomonas infections). Furthermore, a reduced tendency to develop resistant strains is to be expected. By means of a suitable combination it is also possible to combat strains which are not inhibited by the individual components. - 4 Depending on their solubilities and on the presence of basic and/or acid groups, the antibiotics suitable for the synergistic mixtures can be used as free compounds or as salts.

Antibiotics containing acid and basic groups can be used in the form of inner salts, i.e. in zwitter-ion form Antibiotics of predominantly basic character can be used as stable acid addition salts, for example with - 5 inorganic acids, such as hydrochloric acid, sulfuric acid or phosphoric acid, or with suitable organic carboxylic or sulfonic acids, as with aliphatic mono-, di- or tricarboxylic acids, for example acetic acid, malonic acid, tartaric acid, citric acid, 4-(N, N-dipropylsulfamoyl)benzoic acid (Probenecid), with p-toluenesulfonic acid, a- or β-naphthalenesulfonic acid or naphthalene-disulfonic acid, in particular naphthalene-1,5-disulfonic acid. Antibiotics of predominantly acid character can form stable salts with bases and be used in this form. Preferred salts of this kind are in particular pharmaceutically acceptable non-toxic salts, such as alkali metal salts or alkaline earth metal salts, for example sodium, potassium, magnesium or calcium salts, and also ammonium salts with ammonia or suitable organic amines, in which latter case suitable amines for the salt formation are chiefly aliphatic, cycloaliphatic and araliphatic, primary, secondary or tertiary mono-, di- or poiyamines, as well as heterocyclic bases, for example triethylamine, hydroxylower alkyl amines, for example 2-hydroxyethylamine, bis(2-hydroxyethyl)amine or tris-(2-hydroxyethyl)amine, basic aliphatic esters of carboxylic acids, for example (2diethylaminoethyl)-4-aminobenzoate, lower alkyleneamines, for example 1-ethylpiperidine, cycloalkylamines, for example dicyclohexylamine, or benzylamines, for example N,N'-dibenzylethylene diamine, and bases of the pyridine type, for example pyridine, collidine or quinoline.

In a synergistic mixture of this invention, the ratio of cefsulodin sodium to the second antibiotic can vary within wide limits.

The limiting ratios of the mixture components at which synergism occurs depend on the micro-organism, the test system employed, and, naturally, on the second antibiotic. The invention provides in particular synergistic mixtures in which the weight ratio of cefsulodin sodium to the second antibiotic, with retention of the synergistic effect, is between about 1:0.001 and 1:4000. The invention provides in particular mixtures of cefsulodin sodium (=1) with mecillinam in the weight ratio of 1:0.04 to 1:128, with azlocillin, piperacillin or mezlocillin in the weight ratio of 1:0.04 to 1:3125, with amikacin, dibekacin, tobramycin, sisomycin or gentamicin in the weight ratio of 1:0.004 to 1:5, in particular 1:0.06 to 1:0.09, and with cephacetril, cefoxitin or cefotiam in the weight ratio of 1:0.0155 to 1:5.

Most particularly, the invention provides the mixtures described in the Examples.

The synergistic mixtures of the present invention are used for the manufacture of pharmaceutical preparations which contain an effective amount of the above active substance mixtures by themselves or in admixture with customary inorganic or organic, solid or liquid pharmaceutically acceptable carriers or adjuvants, for example preservatives, stabilisers, wetting agents and/or emulsifiers, solubilisers, salts for regulating the osmotic pressure and/or buffers. Such adjuvants are, for example, carbohydrates, such as saccharose, lactose, dextrose, sucrose, - 7 sorbitol, cellulose or cellulose derivatives, such as methyl cellulose, sodium carboxymethylcellulose, polyethylene glycol, polyvinyl pyrrolidone and, in particular, D-mannitol.

The pharmaceutical preparations contain 0.1 to 100%, in particular 1 to 90%, of the active substance mixture, and for convenient dosage are packed in dosage unit forms of 0.1 g to 10 g, for example in ampoules or vials. A preferred dosage form consists of dry-filled ampoules from which an injection, infusion or drip solution can be prepared, before use, by addition of the required amount of pyrogen-free water or another physiologically acceptable solvent, such as a physiological sodium chloride solution or of a plasma-containing solvent.

The pharmaceutical preparations are manufactured in the conventional manner known in the art, usually by nonchemical means, for example by conventional mixing, solution, drying or lyophilising methods.

The synergistic mixtures, or the pharmaceutical preparations which can be obtained therefrom, are administered preferably parenterally, especially intravenously, intramuscularly or subcutaneously. Administration is made either once in high doses (shock therapy), in smaller successive doses or in long-term treatment (e.g. continuous infusion). The dosage depends on the nature and degree of severity of the infection, the weight and general condition of the patient and on the mode of administration, and must be determined by the physician from case to case. In general, the dosage is between 1 and 100 mg/kg in parenteral administration. - 8 When using synergistic mixtures containing cefsulodin sodium and an aminoglycoside, the dosage is between about 3 to 63 mg/kg, the ratio of cefsulodin sodium to aminoglycoside, for example to amikacin, being preferably about 5 1:0.06 to 1:0.22. When using mixtures with the more potent aminoglycosides, for example with gentamycin, the dosage is between about 3.5 to 36 mg/kg and the ratio of the components is between about 1:0.06 and 1:0.09.

The invention is illustrated by the following experimental part and the Examples. - 9 Experimental Part The synergistic, i.e. more than additive, antibiotic effects were determined by means of the following experimental procedures: 1. Demonstration of the synergistic action in vitro: The combination substances A and B are diluted in DST broth OXOID in chessboard arrangement with the aid of microtitre system in Ito 2 steps. All the concentrations of substance A are combined with all the concentrations of substance B and then inoculated with a standardised baceteria suspension (final concentration about 10 bacilli/ml). After incubation for 18 hours at 37°C, the inhibition of the bacterial growth is determined. The inhibiting concentrations are converted by the method of Kerry et al. (J. Antimicrob. Chemotherapy 1, 417-427, 1975) into fractional inhibition concentrations (PIC) MIC of A in combination with B -=FIC of A; (according to the formula MIC of A alone MIC of B in combination with A c _ MIC of B alone FIC of A + FIC of B = £FIC) wherein a STIC of ζθ.7 is taken as proof of the synergistic interaction of the combination components.

By way of example a number of the MIC values (minimum inhibitory concentrations) for the action of cefsulodin sodium, mecillinam, mezlocillin and gentamicin alone against a number of micro-organisms, as well as the MIC values for a number of mixtures in which synergism occurs, are reported in Table 1. - 10 Table 1 Mi cro-o^anisn MIC (scg/al! Anti biotic A alone Antibiotic A in coabination •ith 3 Antibiotic B alone Antibiotic 3 in coabination eitfj A 2?ic Cefsulodia-Na (A Hecillinan (3) Pseudomonas aeruginosa 7S9A I 0.Z5 1000 32 0.23 Escherichia coli 12—44 25 12.5 2 0.12 0.55 Klebsiella pr.s’.aomae 1132 25 6.2 1 0.25 0.50 Klecsiei la pneusoniae 1136 50 12.5 1 0.25 0.:0 Proteus rettgeri 1121 12.5 0.4 3.2 1.5 0.53 Cefsulodin-Sa (A) Mezlocillin (3) Pseudosonas aeruginosa S 12! 1.2 0.04 25 12.5 0.53 Staphylococcus aurous 3.2 0.2 3.2 1.6 0.55 Klebsiella pneusoniae 1 50 25 1.25 0.16 - 0.53 Proteus oiraoilis 1077 50 12.5 1 0.25 0.50 Cefsulodin-Na (A) Oentaaicin (3) Proteus rettgeri 1121 12.5 0.3 1 0.5 0.56 Proteus aeruginosa 410 2 0.06 0.5 0.25 0.53 Pseudosonas aeruginosa 1140 3 2 32 4 0.23 Escherichia coli 575 64 32 2 0.25 0.53 2. Demonstration of the synergistic action in vivo by means of systemic infections in mice: The combination substances A and B are diluted in 1:3 5 to 1:3.3 steps such that the average concentration contains the respective amount of substance A and substance B corresponding to the Εϋ^θ Female MF 2 mice weighing 18 to 22 g are divided at random into groups (n=20) and infected intraperitoneally 1° with a standardised bacteria suspension which, - 11 according to strain, represents 5 to>20 ΙΛ^θ.

Immediately after the infection, and 3 hours later, the mice are treated in groups subcutaneously with dilutions of substances A and B as well as with combinations thereof. These combinations are prepared shortly before administration by mixing each dilution of substance A with each dilution of substance B. The survival rates are determined on the fifth day after the infection. The survival rates of mice which were determined with each of substances A and B alone are compared with those obtained with the combinations of these substances. A combination which has protected more animals from death than would be expected from the independent action of the substances, is designated as having synergistic action.

The survival rates, expressed as percentages of surviving animals, found in infections with a number of pure micro-organisms and in polyinfections is reported in Table 2 for cefsulodin sodium, mecillinam, mezlocillin, gentamicin and CGP 14 221/E, as well as for corresponding mixtures. - 12 Table 2 Infective aicro-organisn Antibiotic Dose s.c. ag/kg surviving animals (in Antibiotic alone coobination Escherichia coll 205 Cefsulodin-Na MecHlinam 25 1 5 0 90 Ce£ealodin-Ha Mecillinam 20 2 5 0 75 1 Cefsulodin-Na Mezlocillin 20 50 20 0 SO Cefsulodin-Na CGP 14221/E 40 0.62 10 0 95 Cefsulodin-Nai CGP 14221/E 20 2.5 0 0 SO Escherichia coli 205 R * TEA Cefsulodin-Na CGP 14221/E1 40 1.2 0 10 70 Klebsiella jneucnniae 329 Cefsulodin-Na Mecillinam 40 300 0 0 85 CefsulodinrNa Mecillinam ao 300 5 0 100 Cefsulodin-Na Mezlocillin 40 300 0 0 90 Cefsulodin-Na Gentamicin 50 0.2 0 30 35 Klebsiella pneuaoniae 327 Cefsulodin-Na CG? 14221/E 100 20 0 0 85 Cefsulodin-Na CGP 14221/S 50 40 0 0 85 Klebsiella pneuaoniae 329 + Escherichia coli 205 Cefsulodin-Na CGP 14221/E 100 20 0 0 70 7jJ-{2-(2-l«jno-l,3-H!l3Zi>lin-4-yl)acit3aiA)]-3-[]-(UiMtiiyla»jno)-tetrazol-5-iflthio»ethpl]-3-cephee-4-carDoxylic acid hydrochloride (Cefotian) 1) - 13 3. Demonstration of the synergistic action in vivo by means of experimental pyelonephritis (polyinfections)in mice: Mice are treated intravenously with 10 mg/kg of carrageenin. Seven days later they are infected with a standardised suspension of bacteria. One hour after the infection, 6 hours later and twice daily on each of 4 subsequent days, the mice are treated subcutaneously with the test substances alone and their combinations.

Two days after the last treatment, the mice are sacrificed, dissected and examined for the number of germs in the kidneys.

The number of animals (in 7.) which exhibit complete clearance of the kidneys from both micro-organisms on 15 treatment with cefsulodin sodium and gentamycin alone and in combination, is reported in Table 3.

Table 3 Infecting micro-organism Antibiotic Bose s.c. ig/kg Clearance of the kidnejo from both micro-organisms (percentage of animals) antibiotic alone combination Pseudomonas aeruginosa Cefsulodin-Na 25 0 + Escherichia coli Gentamicin I 0 70 Pseudoaoras aeruginosa Cefsulodin-Na 25 10 Klebsiella pneuaoniae Gentamicin 2.5 40 SO Cefsulodin-Na Gentamicin 25 0.5 10 0 50 - 14 Pharmaceutical preparations ampoule(s) Example 1: Dry-filled / or vials containing 0.50 g of cefsulodin sodium and 0.02 g of mecillinam (1:0.04) are prepared as follows: Composition (for 1000 ampuls or vials): cefsulodin sodium 500 g mecillinam 20 g mannitol 6 g 526 g The components are homogeneously mixed and an ampoule(s) or vial is filled with 0.526 g of the mixture under aseptic conditions. The ampuls or vials are sealed and tested.

Ampoule(s)or vials containing corresponding amounts of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner.

Example 2: Dry-filled ampoule(s)or vials containing 1 g of cefsulodin sodium and 0.01 g of mecillinam (1:0.01) are prepared as follows: Composition (for 1000 ampuls or vials): cefsulodin sodium 1000 g mecillinam 100 g mannitol 100 g 1200 g The components are homogeneously mixed and an aijipoule(s) or vial is filled with 1.,2, g of the mixture under aseptic ampoule(s) r conditions. The / or vials are sealed and tested. - 15 Ampoule(s)or vials containing corresponding amounts of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner. ampoule(s) Example 3; Dry-filled / or vials containing 2 g of 5 cefsulodin sodium and 0.2 g of mecillinam (1:0.01) are prepared as follows: ampoule(s) Composition (for 1000 / or vials): cefsulodin sodium 2000 g mecillinam 200 g IQ mannitol 200 g 2400 g The components are homogeneously mixed and an ampoule(s) or vial is filled with ,2.4 g of the mixture under aseptic ampoule(s) conditions. The / or vials are sealed and tested.

Atnpoule(s) or vials containing corresponding amounts 15 of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner. ampoule(s) Example 4: Dry-filled - / or vials containing 2 g of cefsulodin sodium and 1 g of mecillinam (1:0.05) are prepared as follows: Composition (for 1000 ampuls or vials): cefsulodin sodium 2000 g mecillinam 1000 g mannitol 300 g 3300 g - 16 The components are homogeneously mixed and an ampoule(s) or vial is filled with 3.3 g of the mixture under aseptic ampouιe(ε) conditions. The / or vials are sealed and tested.

Ampoule(s) or vials containing corresponding amounts 5 of mezlocillin, gentamicin CGP 14221/E as second active component are prepared in the same manner. ampoule(s) Example 5: Dry-filled / or vials containing 1 g of cefsulodin sodium and 1 g of mecillinam (1:1) are prepared as follows: ampoule(s) Composition (for 1000 / or vials): cefsulodin sodium 1000 g mecillinam 1000 g mannitol 200 g 2200 g ampoule(s) The components are homogeneously mixed and an / or vial is filled with 2.2 g of the mixture under aseptic ampoule(s) conditions. The / or vials are sealed and tested.

Ampoule(s) or vials containing corresponding amounts of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner. ampoule(s) Example 6: Dry-filled / or vials containing 1 g of cefsulodin sodium and 2.5 g of mecillinam (1:2.5) are prepared as follows: - 17 Composition (for 1000 ampuls or vials): cefsulodin sodium 1000 g mecillinam 2500 g mannitol 350 g 3850 g The components are homogeneously mixed and an ampoule(s) or vial is filled with 3.85 g of the mixture under aseptic ampoule(s) conditions. The / or vials are sealed and tested.

Ampoule(s) or vials containing corresponding amounts of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner. ampoule(s) Example 7: Dry-filled / or vials containing 0.25 g of cefsulodin sodium and 0.937 g of mecillinam (1:3.75) are prepared as follows: ampoule(s) Composition (for 1000 / or vials): cefsulodin sodium 250 g mecillinam 937 g mannitol 100 g 1287 g The components are homogeneously mixed and an ampoule(s) or vial is filled with 1.287 g of the mixture under aseptic ampoule(s) conditions. The / or vials are sealed and tested.

Ampoule(s) or vials containing corresponding amounts of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner. - 18 arapoule(s) Example 8: Dry-filled / or vials containing 0.25 g of cefsulodin sodium and 1.875 g of mecillinam (1:7.5) are prepared as follows: ampoule(s) Composition (for 1000 / or vials): cefsulodin sodium 250 g mecillinam 1875 g mannitol 200 g 2125 g The components are homogeneously mixed and an ampoule(s) or vial is filledkwi^h^.125 g of the mixture under aseptic conditions. The / or vials are sealed and tested.

Ampoule(s) or vials containing corresponding amounts of mezlocillin, gentamicin or CGP 14221/E as second active component are prepared in the same manner.

Example 9: In accordance with Examples 1 to 8, and using the same ratios of mixture components, dry-filled atnpoule(s) or vials can be prepared which, in addition to containing cefsulodin sodium and mannitol, contain a corresponding amount of clavulanic acid, penicillanic acid 4,4-dioxide, 2o carbenicillin, ticarcillin, sulfocillin, azlocillin, Bay K4999, piperazillin, cephacetril, cefoxitin, cefuroxime, cefazolin, cefamandole, CGP 11481, cefotiam, SCE-1365, cefotaxime, CGP-17845, kanamycin A, amikacin, dibekacin, tobramycin, sisomycin, netilmicin, SCH-22591, SCH-21420, or a pharmaceutically acceptable salt thereof.

Claims (9)

1. CLAIMS:1. A synergistic mixture of antibiotics consisting of cefsulodin sodium and a second antibiotic selected from the group consisting of clavulanlc acid, penicillanic acid 4,45 dioxide, azlocillin', mezlocillin, piperacillin, mecillinam, cephacetril, cefoxitin, cefotiam, amikacin, dibekacin, tobramycin, sisomicin or gentamicin, or a salt thereof.

2. A synergistic mixture according to claim 1, wherein the weight ratio of cefsulodin sodium to the second antibiotic 10 is between 1:0.001 and 1:4000.

3. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and clavulanic acid, or a salt thereof, in a weight ratio between 1:0.001 and 1:4000. 15 4. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and penicillanic acid 4,4-dioxide, or a salt thereof, in a weight ratio between 1:0.001 and 1:4000. 5. A synergistic mixture according to either of claims 1 20 or 2, consisting of cefsulodin sodium and azlocillin, or a salt thereof, in a weight ratio between 1:0.04 and 1:3125. - 20 6. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and mezlocillin, or a salt thereof, in a weight ratio between 1:0.04 and 1:3125. 7. A synergistic mixture according to either of claims 5 1 or 2, consisting of cefsulodin sodium and piperacillin, or a salt thereof, in a weight ratio between 1:0.04 and 1:3125. 8. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and mecillinam, or a 10 salt thereof, in a weight ratio between 1:0.04 and 1:128. 9. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and cephacetril, or a salt thereof, in a weight ratio between 1:0.0155 and 1:5. 10. A synergistic mixture according to either of claims 15 1 or 2, consisting of cefsulodin sodium and cefoxitin, or a salt thereof, in a weight ratio between 1:0.0155 and 1:5. 11. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and cefotiam, or a salt thereof, in a weight ratio between 1*.0.0155 and 1:5 20 12. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and amikacin, or a salt thereof, in a weight ratio between 1:0.004 and 1:5.

4. -8130 - 21 13. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and dibekacin, or a salt thereof, in a weight ratio between 1:0.004 and 1:5. 14. A synergistic mixture according to either of claims

5. 1 or 2, consisting of cefsulodin sodium and tobramycin, or a salt thereof, in a weight ratio between 1:0.004 and 1:5. 15. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and sisomicin, or a salt thereof, in a weight ratio between 1:0.004 and 1:5.

6. 10 16. A synergistic mixture according to either of claims 1 or 2, consisting of cefsulodin sodium and gentamicin, or a salt thereof, in a weight ratio between 1:0.004 and 1:5. 17. A pharmaceutical preparation containing a synergistic mixture as claimed in any one of claims 1 to 16.

7. 15 18. A process for the manufacture of the pharmaceutical preparation of claim 17 hy non-chemical means.

8. 19. A method of combatting infections in a non-human animal, which comprises administering a therapeutically effective dose or series of doses of a composition claimed in

9. 20 claim 17 to an animal in need of such treatment.