GB2067999A

GB2067999A - Epoxysuccinic Acid Derivatives

Info

Publication number: GB2067999A
Application number: GB8102291A
Authority: GB
Original assignee: Nippon Chemiphar Co Ltd
Current assignee: Nippon Chemiphar Co Ltd
Priority date: 1980-01-25
Filing date: 1981-01-26
Publication date: 1981-08-05
Also published as: IT8119331A0; JPS56104816A; FR2474497A1; ES8201567A1; BR8100350A; ES498734A0; JPS6365644B2; IT1141956B; DE3102486A1

Abstract

Novel epoxysuccinic acid derivatives are provided of the formula:- <IMAGE> and non-toxic salts thereof, where R represents a lower alkyl group. These epoxysuccinic acid derivatives and their non-toxic salt are useful for inhibiting myocardial infarction. The derivatives may be made by reacting a leucine lower alkyl ester with a trans- epoxysuccinic acid mono-ester, or trans-epoxysuccinic acid itself.

Description

SPECIFICATION Epoxysuccinic Acid Derivatives The present invention relates to epoxysuccinic acid derivatives, and more particularly to N-(dl-3trans-carboxyoxirane-2-carbonyl)-L-leucine lower alkyl esters and non-toxic salts thereof. The invention further relates to processes for the production of such derivatives, and to anti-myocardial infarct agents containing such esters or salts.

Myocardial infarction is caused by partial death of the myocardium due to a decrease of blood supply, as described in "Terms Used in Cardiovascular Diseases" (U.S. Department of Health, Education and Welfare, Public Health Service, 1964).

Myocardial infarction, like cancer and hypertension, is a typical geriatric disease and its medical treatment is an important problem. Currently, no medicine is available which directly cures myocardial infarction, although there are medicines capable of suppressing the accompanying diseases, such as cardiac insufficiency, arrhythmia, and ischemic heart disease. There is therefore a continuous need for the development of an effective anti-myocardial infarct agent.

As a result of extensive research for an effective anti-myocardial infarct agent, the present applicants have found that certain novel N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine lower alkyl esters and non-toxic salts thereof have extremely superior inhibiting effects against myocardial infarction, and upon this finding, the present invention is based.

Certain N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine derivatives have previously been reported in Japanese Patent Application No. 108923/78 as open to inspection. However, the only medicinal effects of these compounds are stated to be in the suppression of the activity of proteolytic enzymes, and there is no reference to their use as anti-myocardial infarction agents. -Among the compounds disclosed in this prior Specification, N-(dI-3-trans-ethoxycarbonyloxirane-2-carbonyl)-L- leu cine ethyl ester and N-(d l-3-trans-carboxyoxirane-2-carbonyl)-L-leucine, which have structures which are most closely similar to those of the compounds of the present invention.It has been confirmed, however, that despite the structural similarity, the known compounds are less effective in curing myocardial infarction.

It is accordingly, an object of the present invention to provide certain novel N-(dl-3-trans carboxyoxirane-2-carbonyl)-L-Ieuci ne lower alkyl esters and non-toxic salts thereof.

Another object of the present invention is to provide a process for producing such novel N-(dl-3 tra ns-carboxyoxirane-2-ca rbonyl)-L-leucine lower alkyl esters and non-toxic salts thereof.

A further object of the present invention is to provide novel anti-myocardial infarct agents containing N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine lower alkyl esters or non-toxic salts thereof.

According to the invention, there is provided an epoxy-succinic acid derivative in the form of an N-(dl-3-trnns-carboxyoxirane-2-carbonyl)-L-leucine lower alkyl ester represented by the formula (I),

where R represents a lower alkyl group selected from the methyl, ethyl, n-propyl, n-butyl and iso-butyl groups, or a non-toxic salt thereof. Such derivatives may be prepared by protecting one of the carboxyl groups of trans-epoxysuccinic acid, reacting a trans-epoxysuccinic acid monoester so formed with an L-leucine lower alkyl ester represented by the formula (II),

wherein R has the same meanings as defined above, and removing the protecting group from the carboxyl group after the reaction.

Alternatively, the compound of the formula (I) may be prepared by a direct condensation reaction of trans-epoxysuccinic acid with an L-leucine lower alkyl ester.

The ester residue of the trans-epoxysuccinic acid mono-ester, may be a t-butyl group, or a benzhydryl group. This mono-ester residue must be a group which can be removed by means of trifluoroacetic acid or the like, because the mono-ester residue must be removable without removing the L-leucine lower alkyl ester group or the lower alkyl group thereof. The lower alkyl group may be a methyl, ethyl, n-propyl, n-butyl or iso-butyl group.

The trans-epoxysuccinic acid mono-ester may be prepared by reacting trans-epoxysuccinic acid with a t-butyl-activated derivative, such as O-(t-butyl)-N,N'-diisopropyl pseudo urea, in an organic solvent such as tetrahydrofuran, methylene chloride, ethylene chloride, chloroform or benzene, at a temperature of from 0 to 500C, preferably at room temperature, for from 5 to 20 hours.

The reaction of the trans-epoxysuccinic acid mono-ester with an L-leucine lower alkyl ester may be carried out in the presence of a condensation agent such as N-hydroxy succinimide and N,N'dicyclohexyl carbodiimide, or N-hydroxyphthalimide and N,N'-dicyclohexyl carbodiimide, in an organic solvent such as tetrahydrofuran, methylene chloride, ethylene chloride, chloroform or benzene, at a temperature of from 0 to 500C, preferably at room temperature, for from 5 to 20 hours.

The direct condensation reaction of the trans-epoxysuccinic acid with the L-leucine lower alkyl ester may similarly be carried out in the presence of a condensation agent, such as Nhydroxysuccinimide and N,N'-dicyclohexyl carbodiimide, or N-hydroxyphthalimide and N,N'dicyclohexyl carbodiimide, in an organic solvent, such as tetrahydrofuran or dimethylformamide, at a temperature of from 0 to 500C, preferably at room temperature, for a period of from 5 to 20 hours.

The removal of the mono-ester residue, such as a t-butyl group or a benzhydryl group from the starting compound, or trans-epoxysuccinic acid mono-ester, may be carried out by reacting the condensation reaction product with, for example, trifluoroacetic acid, at a temperature of from -10 to 500C for from 0.5 to 5 hours.

The N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine lower alkyl ester, thus obtained, may be converted to a non-toxic salt in known manner, if desired. Preferred non-toxic salts are alkali metal salts, such as sodium and potassium salts, and calcium salts.

The anti-myocardial infarction effectiveness and acute toxicity of certain compounds of the present invention were evaluated by the following methods. The anti-myocardial infarntion effectiveness was studied by means of an experimental myocardial infarction model of rabbits. At the same time, the effectiveness of verapamil hydrochloride and propranolol hydrochloride, which are known to be medicines for healing isohemic heart disease, was also evaluated. The acute toxicity was evaluated using mice.

The myocardial infarction model, the experimental procedure for the toxicity determination and the results obtained, are given below.

Experiment 1 White male rabbits weighing about 2 kg each were anesthetized with pentobarbital sodium (3.5 mg/kg, i.v.). The thorax was opened while giving artificial respiration, and the left anterior descending coronary artery (about 7 mm below the origin) was ligated. After 24 hours, the heart was taken out and the myocardium was sliced from the cardiac apex to the ligated portion to obtain a specimen of a thickness of 2 mm. The dead portion was dyed by the method of Lake et al. [Histochemical J., 2441 (1 970)j and the ratio thereof to the area of the left ventricle was calculated.Medicinal substances were administered 3 minutes prior to the ligature (5 mg/kg, i.v.), at three intervals of one hour starting immediately after the ligature (10 mg/kg/hr, drip infusion), 8 hours after the ligature (5 mg/kg, i.p.) and 20 hours after the ligature (5 mg/kg, i.p.), respectively. To a group of control animals, a 0.9% sodium chloride solution was administered at the same intervals as in the case of the group of animals to which the medicinal substances were administered.

The results obtained are given in Table 1: Table I Animals Ratio of the dead portion { /OJ Nos. A /25 mg/kg) B 63.5 mg/kgJ C 5 mg/kg) Controls 1 11.8 19.6 18.3 19.8 2 5.5 11.2 16.7 19.3 3 0 10.9 9.1 9.1 4 3.8 15.1 11.9 15.5 5 4.9 9.2 18.5 18.9 Average +S.E. 5.2+1.9 13.2+1.9 14.9+1.9 16.5+2.0 A: Potassium salt of N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine ethyl ester (dissolved in a physiological sodium chloride solution) B: Verapanil hydrochloride (dissolved in a physiological sodium chloride solution) C: Propranolol hydrochloride (Inderalt) Injection was used) Experiment 2 Acute Toxicity Test: Mice of ddN type weighing from 20 to 28 g were used.The medicinal substances were intravenously administered from the tails.

As shown in Table 2, the medicinal agent of the present invention was administered up to 1 g/kg.

The results obtained are shown in Table 2.

Table 2 Tested LD50 Compounds (mg/kg, iv.) A MLD > 1000 B 15 C 28 A, B and C are the same as before.

It can be seen from the results of Experiments 1 and 2 that when 25 mg/kg were administered, the compound of the present invention in the form of a non-toxic salt thereof, exhibited an extremely good inhibiting effectiveness against the experimental myocardial infarction of rabbits, as much as 70% as compared to the case where no administration was made, and that no instance of death was observed in the acute toxicity test of mice, even when as much as 1000 mg/kg were administered, and no other adverse symptoms were observed. Thus, it is to be expected that the compounds of the present invention and their non-toxic salts will provide highly safe and useful anti-myocardial infarct agents.

The dosage for a compound of the present invention may vary, depending upon the symptoms of the myocardinal infarction. However, it may usually be administered in an amount of from about 100 mg to about 1 g per day to a patient. The method of administration may be by any one of intravenous, drop infusion, hypodermic, intramuscular and oral administration. It may be administered in any suitable form, for instance, in the form of injections, powders, capsules, tablets, or granules. In the preparation of compositions, it may be combined with an inorganic vehicle, such as magnesium carbonate, silicic acid anhydride, synthetic aluminum silicate, or calcium phosphate, or with an organic vehicle, such as lactose, corn starch, or cellulose, or with any other normally used vehicle.Further, in the case of injections, it may be dissolved in a medium normally used for injections, such as distilled water, a physiological sodium chloride solution, or a lidocaine solution.

The invention is illustrated by the following examples:- Example 1 N-(dl-3-trans-Carboxyoxirane-2-carbonyl-L-leucine Ethyl Ester Into a tetrahydrofuran solution (1 5 ml) containing 709 mg of trans-epoxysuccinic acid mono-tbutyl ester and 434 mg of N-hydroxysuccinimide, 777 mg of N,N-dicyclohexyl carbodiimide was added and the resulting solution stirred at room temperature for 3 hours. Separately, into a tetrahydrofuran suspension (15 ml) of 737 mg of L-leucine ethyl ester hydrochloride, 0.53 ml of triethylamine was added. To this mixture, the above tetrahydrofuran solution was added while filtering off insoluble substances. The resulting mixture was stirred overnight at room temperature. The solvent was then removed by distillation, and ethyl acetate was added.The resulting organic solution was washed with a saturated aqueous sodium bicarbonate solution and with 1 N hydrochloric acid, and dried over sodium sulfate. The solvent was removed and the residue was purified by means of silica gel column chromatography (developer: benzene/ethyl acetate=10/1), whereupon 472 mg of N-(dI-3-trans-5- butoxy carbonyloxirane-2-carbonyl)-L-leucine ethyl ester was obtained as a colourless oily substance (yield: 38%).

íR meat cm~1: 1735,1670,1520,1150,895 NMR (CDCí3)S: 0.95 (6H, d, J=5Hz, CH3) 1.28 (3H, t, J=7Hz, CH2CH3) 1.52 (9H, s, L-butyl)

3.32 (0.5H, d, J=2Hz, 3.42 (0.5H, d, J=2Hz, 3.61 (1H,d,J=2Hz,

4.16 (1 H, q,J=7Hz, O--CH,) 4.18 (1H, q, J=7Hz, O--CH,) 4.58 (1 H, m, NHCHOO) 5.37 (1H, m, NH) To 472 mg of the obtained N-(dl-3-trans-t-butoxy-carbonyloxirane-2-carbonyl)-L-leucine ethyl ester, 5 ml of trifluoroacetic acid was added, and stirred for one hour while cooling on ice. The trifluoroacetic acid was removed by distillation, and a saturated sodium bicarbonate aqueous solution was added.The aqueous solution was washed with chloroform, acidified with 1 N hydrochloric acid, extracted with ethyl acetate, dried over sodium sulfate, and the solvent removed, whereupon 375 mg of N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine ethyl ester was obtained as a colourless oily substance (yield: 95%).

IR meaaxt cm~1: 1730,1660,1540,1200,895 NMR (CDCI3): 0.94 (6H, d, J=5Hz, CH3) 1.27 (3H, t, J=7Hz, CH2CH3)

3.54 (0.5H, s, 3.60 (0.5H, s 3.78 (1 H, s,

4.16 (2H, q, J=7Hz, C-CH2) 4.56 (1H, m, NCHCO) 6.88 (0.5H, d, J=8Hz, NH) 7.09 (0.5H, d, J=8Hz, NH) 10.18 (1H, brood, COOH) Example 2 To a solution of 39.6 g of trans-epoxysuccinic acid in dimethyíformamide (1 on ml) and tetrahydrofuran (300 ml), 34.5 g of N-hydroxysuccinimide was then added to which a tetrahydrofuran solution (100 ml) of 61.8 g of dicyclohexylcarbodiimide was then added slowly, and the mixture was stirred at room temperature for 2 hours.To this mixture 48.8 g of an L-leucine ethyl ester was added and the resulting mixture was stirred overnight at room temperature. Any insoluble substances were removed by filtration, and the filtrate was concentrated. To the residue was added ethyl acetate, and the resulting organic solution was extracted with a saturated sodium bicarbonate aqueous solution.

The aqueous extract was acidified with a 1 N hydrochloric acid solution and extracted further with ethyl acetate. The extraction from the ethyl acetate solution with an aqueous sodium bicarbonate solution was repeated as the extract was acidified with 1 N hydrochloric acid and extracted again with ethyl acetate. This ethyl acetate solution was washed twice with an aqueous sodium chloride solution and dried over sodium sulfate. Removal of the solvent gave 30 g of an N-(dl-3-trans-carboxyoxirane-2- carbonyl)-L-leucine ethyl ester as a colorless oily substance (yield: 37%).

Example 3 Potassium Salt of N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine Ethyl Ester To an aqueous solution (1 ml) containing 1 37 mg of potassium hydrogen carbonate, an ethyl acetate solution (10 ml) containing 375 mg of the N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine ethyl ester, was added. The resulting solution was stirred for 10 minutes, and completely dried under reduced pressure, to give 462 mg of a potassium salt of N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L- ecine ethyl ester as a hygroscopic white powder (quantitatively).

IRuKBxcm~': 1730,1670,1610,1540, 1380,1200,895 NMR (DMSO-d6)S: 0.84 (6H, m, CH3) 1.15(3H, t, J=7Hz, CH2ChZ3)

2.94(1 H, m, 3.24(1 H, m,

4.01 (2H, q, J=7Hz, O--CH,) 4.20 (1 H, m, NHCHCO) 8.25 (1 H, d, J=8Hz, NH) Example 4 Sodium Salt of N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine Ethyl Ester To an aqueous solution (2 ml) containing 27 mg of sodium hydrogen carbonate, an ethyl acetate solution (5 ml) containing 87 mg of N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine ethyl ester, was added.The resultant solution was stirred for 10 minutes at room temperature, and completely dried under reduced pressure, to give 94 mg of a sodium salt of N-(dl-3-trans-carboxyoxirane-2carbonyl)-L-leucine ethyl ester as a hygroscopic white powder (quantitatively).

lRv D KBx cm-': 1730,1650,1615,1540, 1380,1200,895 NMR(DMSO-d6)S: 0.86 (6H, m, CH3) 1.16 (3H, t, J=7Hz, CH2CH3)

2.99 (1H, m, 3.28 (1H, m,

4.01 (2H, q, J=7Hz, O--CH,) 4.20 H, m, NHCHCO) 8.28 (1 H, d, J=7Hz, NH) Example 5 Composition (Tablets) Film coating tablets were prepared to contain the following ingredients per tablet (220 mg): :- Potassium salt of an N-(dl-3-trans-carboxyoxirane-2-carbonyl-L-leucine ethyl ester 50 mg Lactose 100 mg Crystalline cellulose 50 mg Magnesium stearate 1 mg Hydroxypropylmethyl cellulose 15 mg Hydroxypropyl cellulose 4 mg Example 6 Composition (Granules) The following ingredients are contained in 1 g of the granular composition N-(dl-3-trans-carboxyoxirane-2-carbonyl)-L-leucine ethyl ester 200 mg Lactose 500 mg Corn starch 300 mg

Claims

1. An epoxysuccinic acid derivative represented by the formula:

where R represents a lower alkyl group selected from the methyl, ethyl, n-propyl, n-butyl and iso-butyl groups, or a non-toxic salt thereof.

2. An epoxysuccinic acid derivative as claimed in Claim 1, said derivative being an N-(dl-3-trans- carboxyoxirane-2-carbonyl)-L-leucine lower alkyl ester, the lower alkyl group of which is selected from the methyl, ethyl, n-propyl, n-butyl and iso-butyl groups, or a non-toxic salt thereof.

3. An epoxysuccinic acid derivative as claimed in Claim 1 or Claim 2, wherein the lower alkyl group is an ethyl group.

4. An epoxysuccinic acid derivative as claimed in Claim 1 substantially as hereinbefore described with reference to any of Examples 1 to 4.

5. A process for producing an epoxysuccinic acid derivative of the formula

where R represents a lower alkyl group selected from the methyl, ethyl, n-propyl, n-butyl and iso-butyl groups, or a non-toxic salt thereof, said process comprising reacting a leucine lower alkyl ester of the formula

where R is the same as in formula (I), with a transepoxysuccinic acid mono-ester, and if desired, converting the resulting derivative to a corresponding non-toxic salt in known manner.

6. A process as claimed in Claim 5, wherein an L-leucine lower alkyl ester is reacted with a dltrans-epoxysuccinic acid mono-esten

7. A process as claimed in Claim 5 or Claim 6, wherein L-leucine ethyl ester is reacted with a dltrans-epoxysuccinic acid mono-ester.

8. A process as claimed in Claim 5 substantially as hereinbefore described with reference to any one of Examples 1,3 and 4.

9. A process for producing an epoxysuccinic acid derivative of the formula

where R is a lower alkyl group selected from the methyl, ethyl, n-propyl, n-butyl and iso-butyl groups, or a non-toxic salt thereof, said process comprising reacting a leucine lower alkyl ester of the formula:

where R is the same as in formula (I), with trans-epoxysuccinic acid, and, if desired, subsequently converting the resulting derivative to a corresponding non-toxic salt in known manner.

10. A process as claimed in Claim 9 substantially as hereinbefore described with reference to Example 2.

11. An anti-myocardial infarct agent comprising an epoxysuccinic derivative as claimed in Claim 1.

12. An anti-myocardial infarct agent as claimed in Claim 11, wherein the derivative is an N-(dl-3trans-carboxyoxirane-2-carbonyl)-L-leucine lower alkyl ester, in which the lower alkyl group is selected from the methyl, ethyl, n-propyl, n-butyl and iso-butyl groups, or a non-toxic salt thereof.

13. An anti-myocardial infarct agent as claimed in Claim 11 or Claim 12, wherein the derivative is N-(dl-3-trans-carboxyoxirane-2-ca rbonyl)-L-Ieucine ethyl ester, or a non-toxic salt thereof.

14. An anti-myocardial infarct agent as claimed in Claim 10, substantially as hereinbefore described with reference to Example 5 or Example 6.