JP2000191616A - New peptidylaldehyde derivative and medicine containing the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a new compound, represented by a specific formula, having a free radical capturing activity and a calpain inhibiting activity and useful as a medicine useful for preventing and treating cytopathy. SOLUTION: This compound is represented by formula I (A is an organic compound residue having a free radical capturing activity; R1 and R2 are each H or a lower alkyl) or its pharmacologically acceptable salt, e.g. 2-((6- hydroxy-2,5,7,8-tetramethylchroman-2-yl)carbonylamino)-3-methyl-N-(1-(2- methylpropyl)-2-oxoethyl)butanamide. Furthermore, the compound is obtained by passing a reaction for reacting an aldehyde derivative represented by formula II (B is a substituent group used as a protecting group of amino group) with an alcohol under acidic conditions and then providing an acetal derivative represented by formula III (R3 and R4 are each a lower alkyl, benzyl or the like).

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a novel compound having a free radical scavenging activity and a calpain inhibitory activity. Furthermore, the present invention relates to a medicament comprising a compound having a free radical scavenging activity and a calpain inhibitory activity as an active ingredient.

[0002]

2. Description of the Related Art Among proteases that degrade proteins, those having a cysteine residue in the active center of the enzyme are called cysteine proteases. A typical example is calpain present in the cytoplasm. Calpain exists widely in the body, is activated by calcium ions, and has an optimum pH near neutrality. Although the physiological role of this enzyme is often unknown, it has been revealed to date that the degradation of cytoskeletal proteins, activation of inactive cell precursors such as protein kinase C, and degradation of receptor proteins And so on. Furthermore, it has become clear that abnormalities in the activity of this enzyme are involved in many diseases. For example,
Stroke, subarachnoid hemorrhage, Alzheimer's disease, ischemic disease, muscular dystrophy, cataract, platelet aggregation, arthritis,
Involvement in intractable diseases such as osteoporosis has been suggested.
[Trends in Pharmacological Sciences, 15, 412 (1994)].

Heretofore, peptide aldehyde derivatives (Japanese Patent Laid-Open No. 6-28)
7167), peptide diazomethane derivative [Biochem.
J., 253, 751-758 (1988), J. Med. Chem., 35
Volume, 216-220 (1992)], peptide disulfide derivatives [Chem. Lett., 191-194 (1990)], and isocoumarin derivatives (WO 92/118).
50), and non-peptidic compounds such as KP-1241 (JP-A-6-41067).

In recent years, the involvement of calpain in brain and neuronal cell damage due to ischemia has been revealed.
The efficacy of the above inhibitors has been demonstrated [Raymond T., Ba
rtus, Stroke, 25, 2265-2270 (1994)].

[0005] Causes of brain cell and neuronal damage include trauma, inflammation, light damage, degenerative degeneration and the like in addition to the above-mentioned ischemia. Activation and oxidation of intracellular proteases due to stress induced by these ischemia. Sexual disorders trigger cell death in the brain and nerves, causing a loss of cellular function.
This destroys cells, causing tissue damage, organ damage,
This results in loss of organ function. Their loss of function is caused by both acute and chronic disorders [Prog. Neuro-Psycopharmacol. And Biol., 16, 23.
30 (1993), Pysch., 17, 21-70 (1993
Year)〕.

Many papers have discussed that organic compounds having free radical scavenging activity have a protective effect against oxidative damage caused by cellular stress [Arch. P
harmacol., Vol. 325, pp. 129-146 (1992), Free Rad.
Biol. Med., 11, 215-232 (1991), Eur. J. Ph.
armacol., 210, 85-90 (1992)]. In addition, particularly as a protective substance against ischemic neuropathy, for example, an enzyme inhibitor relating to arachidonic acid cascade, an ion channel blocker, a PAF antagonist, neurotrophin,
Many classes of compounds are known, such as growth factors, calpain inhibitors and antioxidants [J. Clin. Neurosci.,
4 (3), 290-310 (1997)]. As mentioned above, protective substances against many ischemic neuropathies have been found so far, but simultaneously administering them in duplicate to patients can be a major instability factor in drug metabolism and distribution. For this reason, we have conducted research on compounds that combine these advantages in one molecule. In the past research on calpain inhibitors, no knowledge about its antioxidant ability was obtained, and creation of an excellent drug having a potent antioxidant action while maintaining calpain inhibitory activity was desired.

[0007]

An object of the present invention is to develop a novel compound having calpain inhibitory activity and free radical scavenging activity and useful for prevention and treatment of various cell disorders.

[0008]

Means for Solving the Problems The present inventors have intensively studied to solve the above problems. As a result, the following general formula:

[0009]

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Wherein A is a residue of an organic compound having free radical scavenging activity, and R ₁ and R ₂ each represent hydrogen or a lower alkyl group. The dipeptidyl aldehyde derivative or a pharmaceutically acceptable salt thereof has a calpain inhibitory activity and a free radical scavenging activity, and has completed the present invention.

[0011]

DETAILED DESCRIPTION OF THE INVENTION That is, the present invention provides a compound represented by the following general formula:

[0012]

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Wherein A is an organic compound residue having a free radical scavenging activity, and R ₁ and R ₂ each represent hydrogen or a lower alkyl group. The dipeptidyl aldehyde derivative or a pharmaceutically acceptable salt thereof, or the derivative or the pharmaceutically acceptable salt thereof according to the above (1), wherein the organic compound residue is a phenolic compound; (3) the derivative according to the above (2), wherein the phenolic compound is a compound selected from a phenol derivative, a benzofuran derivative or a benzopyran derivative, or a pharmaceutically acceptable salt thereof; 6-bis (tert-butyl) phenol, 2,3,5-trimethylbenzene-1,4-diol, 2,3-dimethoxy-5-methyl-1,4-diol, 2,3-dimethoxy-5-methyl The derivative according to the above (3), which is a compound selected from -6-vinylbenzene-1,4-diol, or a pharmaceutically acceptable salt thereof. (5) benzofuran derivatives 2,4,6,7
Tetramethyloxaindan-5-ol, 3,4
5,6-tetramethyloxaindan-5-ol, 6
-Methyl-1,3-dihydroisobenzofuran-4,
The derivative according to (3) above, which is a compound selected from 5,6-triol, or a pharmaceutically acceptable salt thereof, and (6) the benzopyran derivative is 2,5,7,8-tetramethylchroman The derivative according to the above (3) or a pharmaceutically acceptable salt thereof, (7) the general formula (1)

[0014]

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Wherein A is a residue of an organic compound having free radical scavenging activity, and R ₁ and R ₂ each represent hydrogen or a lower alkyl group. And a pharmaceutically acceptable salt thereof as an active ingredient; and (8) the drug according to the above (7), which is an agent for preventing or treating cell damage.

In the general formula (I), the organic compound residue having free radical scavenging activity represented by A may be any compound as long as it has free radical scavenging activity. Phenolic compounds having activity are preferred. Phenol derivatives, benzofuran derivatives and benzopyran derivatives are preferred as phenolic compounds. Examples of the phenol derivative include 2,6-bis (tert-butyl) phenol, 2,3,5-trimethylbenzene-1,4-diol, 2,3-dimethoxy-5-methyl-1,4-diol, , 3-dimethoxy-5-methyl-6-vinylbenzene-1,4-diol and the like. Examples of the benzofuran derivative include 2,4,6,7-tetramethyloxaindan-5-ol, 3,4 , 5,6-Tetramethyloxaindan-5-ol, 6-methyl-
Examples of 1,3-dihydroisobenzofuran-4,5,6-triol and the like, and examples of the benzopyran derivative include 2,5,7,8-tetramethylchroman and the like.

In the above formula (I), lower alkyl groups represented by R ₁ and R ₂ include, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, s
ec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, tert-pentyl, hexyl,
4-methylpentyl, 1,1-dimethylbutyl, 2,2
-Dimethylbutyl, 3,3-dimethylbutyl, 2-ethylbutyl and the like. Preferred are methyl, isopropyl and isobutyl. The lower alkyl group represented by R ₁ and R ₂ may have a substituent, and examples of the substituent include an aryl group, a cycloalkyl group, and an aromatic hetero residue. Examples of the aryl group include phenyl, 1-naphthyl, 2-naphthyl and the like, and phenyl is preferable. Examples of the cycloalkyl group include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like, and preferably cyclohexyl. As the aromatic heterocyclic residue, oxygen,
Short-chain heterocyclic and fused-type heterocyclic residues substituted with nitrogen and sulfur atoms are included. Examples of the short heterocyclic residue include pyrrolyl, furanyl, thiophenyl,
Oxazolyl, thiazolyl, imidazolyl, pyrazolyl, pyridine, and the like. . Preferred specific examples of the optionally substituted lower alkyl group represented by R ₁ and R ₂ are isopropyl, isobutyl, benzyl, cyclohexylmethyl and indol-3-ylmethyl.

The salt of the compound represented by formula (I) in the present invention is preferably a physiologically acceptable salt, for example, a salt with an inorganic base (for example, an alkali metal salt such as a sodium salt or a potassium salt, Alkaline earth metal salts such as calcium salts and magnesium salts), salts with inorganic acids (eg, hydrochloride, hydrobromide, sulfate, nitrate, phosphate, etc.) and salts with organic acids (eg, acetic acid) Salts, citrates, toluenesulfonates, etc.), but are not limited thereto. Furthermore, the present invention also includes various solvates and polymorphs of the compound represented by the general formula (I) and salts thereof.

The compound represented by the general formula (I) of the present invention can be produced by a method combining a known peptide synthesis method with acetalization and deacetalization of an aldehyde group. )

[0020]

Embedded image [In the formula (II), B represents a substituent usually used as a protecting group for an amino group, and R ₁ and R ₂ have the same meaning as described above. The aldehyde derivative represented by the general formula (III) is reacted with an alcohol under acidic conditions.

[0021]

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[In the formula (III), B, R ₁ and R ₂ have the same meanings as described above, and R ₃ and R _{4 each} represent a lower alkyl group or a benzyl group, or may be linked to form a ring. Good. And an acetal derivative represented by the following formula: In the general formula (II), examples of the substituent used as a protecting group for the amino group represented by B include benzyloxycarbonyl, tert-butoxycarbonyl, 9-fluorenylmethoxycarbonyl and the like. Examples of the acidic conditions include acetic acid, p-toluenesulfonic acid, camphorsulfonic acid, pyridinium p-toluenesulfonic acid, organic acids such as trimethylsilyl chloride, inorganic acids such as hydrochloric acid and sulfuric acid, and the presence of an anion exchange resin. But
Preferably, it is in the presence of p-toluenesulfonic acid. As the alcohol used in the above reaction, methanol,
Ethanol, benzyl alcohol, ethane-1,2-diol, propane-1,3-diol, 2,2-dimethylpropane-1,3-diol and the like can be mentioned, but preferably ethane-1,2-diol is used. is there. Examples of the reaction solvent include conventional solvents that do not adversely affect the reaction, such as benzene and toluene, and mixed solvents thereof, and benzene is preferred. The reaction temperature is usually in a range from cooling to heating, and preferably in a range from room temperature to reflux temperature.

Next, by deprotecting the amino protecting group of the compound represented by the general formula (III), the compound represented by the general formula (IV)

[0024]

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[In the formula (IV), R ₁ , R ₂ , R ₃ and R
₄ has the same meaning as described above. Is synthesized. Examples of the deprotection conditions include acid hydrolysis and catalytic reduction, and preferably catalytic reduction.

Next, a compound represented by the general formula (IV) or a reactive derivative at the amino group thereof or a salt thereof,
General formula (V)

[0027]

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[In the formula (V), A is as defined above. ]
By condensing a reactive derivative or a salt thereof at the carboxylic acid or its carboxyl group represented by the general formula (VI)

[0029]

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[In the formula (VI), A and R ₁ , R ₂ ,
R ₃ and R ₄ are as defined above. An N-substituted dipeptidyl acetal derivative represented by the following formula is synthesized.

In the above reaction, as the suitable reactive derivative at the amino group of the compound represented by the general formula (IV), a compound represented by the general formula (IV) and a carbonyl compound such as an aldehyde and a ketone may be used. Or a compound represented by the general formula (IV) and a bis (trimethylsilyl) acetamide, a mono (trimethylsilyl) acetamide, a bis (trimethylsilyl) urea, etc. A silyl derivative formed by a reaction with a silyl compound,
Alternatively, a derivative or the like generated by a reaction between the compound represented by the general formula (IV) and phosphorus trichloride or phosgene can be given. Suitable salts of the compound represented by the general formula (IV) and a reactive derivative thereof include, for example, inorganic salts such as hydrochloride, hydrobromide, nitrate, sulfate and phosphate, for example, formate, Acetate, trifluoroacetate, fumarate,
Organic salts such as oxalate, tartrate, maleate, citrate, succinate, malate, methanesulfonate, benzenesulfonate, p-toluenesulfonate and the like can be mentioned. Salts of these reactive derivatives can be arbitrarily selected depending on the kind of the compound represented by the general formula (IV) to be used.

In the above reaction, suitable reactive derivatives at the carboxyl group of the compound represented by formula (V) include acid halides, acid anhydrides, activated amides, and the like.
Activated esters and the like. Acid halides include acid chlorides, and acid anhydrides include, for example, substituted phosphoric acids (dialkyl phosphoric acid, phenyl phosphoric acid,
Diphenylphosphoric acid, dibenzylphosphoric acid, halogenated phosphoric acid, etc.), dialkyl phosphorous acid, sulfurous acid, thiosulfuric acid, sulfuric acid, sulfonic acid (methanesulfonic acid, etc.), aliphatic carboxylic acid (acetic acid, propionic acid, butyric acid, isobutyric acid, Pivalic acid, pentanoic acid, isopentanoic acid, trichloroacetic acid, etc.)
Or a mixed acid anhydride or a symmetric acid anhydride with an acid such as an aromatic carboxylic acid (such as benzoic acid). Suitable examples of activated amides include, for example, imidazole,
-Substituted imidazole, dimethylpyrazole, triazole or tetrazole and the like. Preferred examples of the activated ester include, for example, cyanomethyl ester, methoxymethyl ester, dimethyliminomethyl ester, vinyl ester, propargyl ester, p-nitrophenyl ester, trichlorophenyl ester, pentachlorophenyl ester, methylphenyl ester, phenylazo Phenyl ester, phenyl thioester, p-nitrophenyl thioester, p-cresyl thioester, carboxymethyl thioester, pyranyl ester, pyridyl ester, 8-quinolyl thioester, or N, N-dimethylhydroxyamine, 1-
Examples thereof include esters with N-hydroxy compounds such as hydroxy-2- (1H) -pyridone, N-hydroxysuccinimide, N-hydroxyphthalimide, and 1-hydroxy-1H-benzotriazole. Suitable salts of the compound represented by the general formula (V) and its reactive derivative include, for example, alkali metal salts such as sodium salt and potassium salt, alkaline earth metal salts such as calcium salt and magnesium salt, and aluminum salt And ammonium salts such as organic base salts such as trimethylamine salt, triethylamine salt, pyridine salt, picoline salt, ethanolamine salt, diethanolamine salt, triethanolamine salt, dicyclohexylamine salt, N, N-dibenzylethylenediamine salt and the like. Base salts are mentioned. These reactive derivatives can be arbitrarily selected depending on the kind of the compound represented by the general formula (V) to be used.

In the present reaction, when the compound represented by the general formula (V) is used in a free form or a salt thereof, N, N'-dicyclohexylcarbodiimide, N-cyclohexyl-N'-morpholinoethylcarbodiimide , N
-Cyclohexyl-N '-(4-diethylaminocyclohexyl) carbodiimide, N, N'-diethylcarbodiimide, N, N'-diisopropylcarbodiimide, N-
Ethyl-N ′-(3-dimethylaminopropyl) carbodiimide, N, N′-carbonylbis (2-methylimidazole), pentamethyleneketene-N-cyclohexylimine, diphenylketene-N-cyclohexylimine, ethoxyacetylene, 1- Alkoxy-1-chloroethylene, trimethyl phosphite, ethyl polyphosphate, isopropyl polyphosphate, phosphorous oxychloride, diphenylphosphoryl azide, thionyl chloride, oxalyl chloride, for example, lower alkyl haloformate such as ethyl chloroformate, isopropyl chloroformate, Triphenylphosphine, 2-ethyl-7-hydroxybenzisoxazolium salt, 2
-Ethyl-5- (m-sulfophenyl) isoxazolium hydroxide inner salt, N-hydroxybenzotriazole, 1- (p-chlorobenzenesulfonyloxy) -6-chloro-1H-benzotriazole, N, N
It is desirable to carry out the reaction in the presence of a conventional condensing agent such as the so-called Vilsmeier reagent prepared by the reaction of dimethylformamide with thionyl chloride, phosgene, trichloromethyl chloroformate, phosphorus oxychloride and the like.

In the above reaction, compounds (IV) and (V)
The reaction of the compound represented by is usually water, for example, alcohols such as methanol and ethanol, acetone, dioxane, acetonitrile, chloroform, methylene chloride, ethylene chloride, tetrahydrofuran, ethyl acetate, N, N
The reaction is carried out in a conventional solvent such as dimethylformamide and pyridine, but the reaction can be carried out in any other organic solvent which does not adversely affect the reaction. These conventional solvents may be used as respective mixtures. The reaction can also be carried out in the presence of an inorganic or organic base such as alkali metal bicarbonate, tri (lower) alkylamine, pyridine, N- (lower) alkylmorpholine, N, N-di (lower) alkylbenzylamine and the like. May go. The reaction temperature is not particularly limited, but the reaction is usually performed under cooling, at room temperature or under heating. Finally, by hydrolyzing the acetal site of the compound represented by the general formula (VI), the desired compound represented by the general formula (I)

[0035]

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[In the formula (I), A, R ₁ and R ₂ are as defined above. To synthesize a peptidyl aldehyde derivative represented by the following formula: As the conditions for the hydrolysis of the acetal moiety, trifluoroacetic acid, acetic acid, p-toluenesulfonic acid, organic acids such as oxalic acid, hydrochloric acid, inorganic acids such as sulfuric acid, or
Examples of the method include acid hydrolysis in the presence of a Lewis acid such as ferric chloride, zinc, trimethylsilyl iodide, and bisacetonitrile dichloropalladium. Preferably, the hydrolysis is performed in the presence of bisacetonitrile dichloropalladium.

Since the compound of the present invention has both free radical-scavenging activity and calpain inhibitory activity, a medicament containing the compound is advantageously used as an agent for preventing and treating cell damage induced by free radicals and calpain. it can. Various diseases caused by cell damage induced by free radicals and calpain, for example, cataract, retinal disorder, genetic degenerative disease, macular degeneration, ocular ischemia, angiogenic disease, glaucoma, ischemia reperfusion injury, It is useful as a preventive and therapeutic agent for optical damage, damage due to eye surgery, ischemic neuronal damage, and the like.

The medicament of the present invention is appropriately used orally or parenterally. As the form of the preparation, any of solid preparations such as tablets, granules, powders, capsules, and ointments and liquid preparations such as injections and eye drops can be appropriately prepared by a known method. These preparations usually contain excipients, binders, disintegrants, dispersants, absorption enhancers, buffers, surfactants, solubilizers, preservatives, emulsifiers, isotonic agents, stabilizers and pH. Various additives such as a regulator may be appropriately used.

When the medicament of the present invention is used as a prophylactic / therapeutic agent for cytotoxicity, the dose depends on the kind of the disease to be treated, the kind of the compound to be used, the age and weight of the patient, the indications and the dosage form thereof. For example, in the case of an oral preparation, the dosage is about 1 mg to 100 mg once a day for an adult several times a day.
It is better to administer to the extent. In the case of injection, adult 1
It is preferable to administer about 0.1 mg to 30 mg once a day.
Further, in the case of eye drops, the present compound is added in an amount of about 0.01 w / v%.
~ 1.0 w / v%, preferably about 0.05 w / v% ~
The eye drop containing 0.5% w / v is preferably administered in a dose of 1 to several drops, about 1 to 4 times a day depending on the symptoms.

The medicament of the present invention may appropriately contain other components for preventing or treating cell damage or other medicinal components as long as the object of the present invention is not violated.

[0041]

The present invention will now be described in detail with reference to examples, test examples and preparation examples, but the present invention is not limited to these examples. In the physical properties of the compounds of the examples,
Proton nuclear magnetic resonance spectrum (NMR) is measured by Varian Gemin
i 2000 (300 MHz), and melting points were measured using Shibata MEL-270, respectively.

Example 1 2-((6-hydroxy-2,
5,7,8-Tetramethylchroman-2-yl) carbonylamino) -3-methyl-N- (1- (2-methylpropyl) -2-oxoethyl) butanamide (Compound 1) Step 1 N-benzyloxycarbonyl To 10 mL of a benzene solution of -L-valinyl-L-leucinal (2.00 g, 5.74 mmol) was added ethylene glycol (1.07 g, 17.22).
mmol) and pyridinium p-toluenesulfonate (0.43 g, 1.72 mmol) were added and the mixture was heated under reflux for about 2 hours. The resulting water was trapped using a Dean-Stark tube. After evaporating the solvent of the reaction solution, the reaction solution was dissolved in 100 mL of ethyl acetate, and twice with 100 mL of saturated aqueous sodium hydrogen carbonate solution.
Washed once with 00 mL of saturated saline. After the obtained organic layer was dried over magnesium sulfate, the solvent was distilled off and N
-(1- (2,5-dioxolanyl) -3-methylbutyl)
-3-Methyl-2-((phenylmethoxy) carbonylamino) butanamide (1.87 g, 83%) was obtained as a white solid. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 0.76-0.84 (12H, m), 1.1
6-1.38 (2H, m), 1.55 (1H, m), 1.87-1.97 (1H, m), 3.
70-4.01 (6H, m), 4.68 (1H, d, J = 3.1 Hz), 5.00 (2
H, dd, J = 21.5, 12.5 Hz), 7.17 (1H, d, J = 9.2 H
z), 7.28-7.34 (5H, s), 7.52 (1H, d, J = 9.5 Hz).

Step 2 50 ml of N- (1- (2,5-dioxolanyl) -3-methylbutyl) -3-methyl-2-((phenylmethoxy) carbonylamino) butanamide (1.85 g, 4.71 mmol) in 50 mL of ethanol To the solution, 5% palladium carbon (0.50 g) was added under a nitrogen atmosphere, and catalytic reduction was carried out overnight at room temperature and normal pressure. After the reaction solution was filtered, the filtrate was evaporated to dryness to give 2-amino-N- (1- (2,5-dioxolanyl)-
3-methylbutyl) -3-methylbutanamide (1.20
g, 98%) as a white solid. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 0.72-0.86 (12H, m), 1.
16-1.64 (5H, m), 1.84-1.90 (1H, m), 2.93 (1H, d, J
= 4.7 Hz), 3.72-3.89 (4H, m), 3.94-4.03 (1H, m),
4.70 (1H, d, J = 3.1 Hz), 7.58 (1H, d, J = 9.6 H
z).

Step 3 (S)-(+)-6-Hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (5.0 g, 19.98
mmol) and hydroxysuccinimide (2.99 g, 2
5.97 mmol) in tetrahydrofuran (100 m
L) and cooled on ice. 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (4.98 g,
25.97 mmol) in 100 mL of methylene chloride and triethylamine (2.63 g, 25.97 mmol).
l) was added to the reaction solution together with the mixture, followed by stirring at room temperature overnight. After the reaction, the reaction solution was dissolved in 100 mL of chloroform, and washed three times with 200 mL of 1N hydrochloric acid, once with 100 mL of a saturated aqueous solution of sodium hydrogencarbonate, and once with 100 mL of saturated saline. After the obtained organic layer was dried over magnesium sulfate, the solvent was evaporated. Recrystallize using 40 mL of hexane: ethyl acetate = 3: 1 and add another 180
The washing was repeated with mL of the above solvent, and 2,5-dioxopyrrolidinyl-6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylate was converted into white crystals (3.54 g, 51%). As obtained. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 1.69 (3H, s), 1.89-2.0
3 (7H, m), 2.39-2.47 (1H, m), 2.64-2.66 (2H, m), 2.
75 (4H, s), 7.53 (0.5H, s).

Step 4 2-amino-N- (1- (2,5-dioxolanyl) -3-
Methylbutyl) -3-methylbutanamide (0.75 g,
2.17 mmol) was dissolved in 50 mL of methylene chloride and 2,5-dioxopyrrolidinyl-6-hydroxy-2,
5,7,8-Tetramethylchroman-2-carboxylate (0.56 g, 2.17 mmol) was added to the reaction solution together with triethylamine (0.66 g, 6.51 mmol), and the mixture was stirred at room temperature overnight. After the solvent was distilled off under reduced pressure, the solvent was distilled off, and the residue was dissolved in 100 mL of ethyl acetate and washed twice with 100 mL of 1N hydrochloric acid, once with 100 mL of a saturated aqueous solution of sodium hydrogencarbonate and once with 100 mL of a saturated saline solution. After the obtained organic layer was dried over magnesium sulfate, the solvent was removed under reduced pressure to obtain N- (1-
(2,5-dioxolanyl) -3-methylbutyl) -2-
((6-Hydroxy-2,5,7,8-tetramethylchroman-2-yl) carbonylamino) -3-methylbutanamide (1.11 g, quantitative) was obtained. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 0.30 (3H, d, J = 6.9 H
z), 0.49 (3H, d, J = 6.7 Hz), 0.78-0.86 (8H, m), 1.
22-1.36 (2H, m), 1.38 (3H, s), 1.48-1.51 (1H, m),
1.68-1.72 (1H, m), 1.95 (3H, s), 2.05 (3H, s), 2.1
0 (3H, s), 2.21-2.60 (2H, m), 3.71-3.83 (4H, m),
3.96-4.02 (1H, m), 4.18 (1H, dd, J = 9.1, 5.2 Hz),
4.65 (1H, d, J = 3.2 Hz), 6.91 (1H, d, J = 9.2 H
z), 7.52 (1H, s), 7.82 (1H, d, J = 9.3 Hz).

Step 5 N- (1- (2,5-dioxolanyl) -3-methylbutyl) -2-((6-hydroxy-2,5,7,8-tetramethylchroman-2-yl) carbonylamino) -3-Methylbutanamide (1.11 g, 2.26 mmol) was dissolved in 110 mL of acetone, and bis (acetonitrile) dichloropalladium (II) (0.03 g, 0.11
mmol) and stirred at room temperature overnight. After the solvent was distilled off under reduced pressure, the reaction solution was dissolved in 200 mL of ethyl acetate, and dissolved twice in 100 mL of 1 N hydrochloric acid, once with 100 mL of a saturated aqueous solution of sodium hydrogen carbonate, and saturated saline 10 times.
Washed once with 0 mL. After the obtained organic layer was dried over magnesium sulfate, the solvent was removed under reduced pressure, and 1.18 g of the resulting mixture was obtained as a viscous oil. This is hexane:
Purification by silica gel column chromatography using ethyl acetate = 1: 1, 2-((6-hydroxy-2,
5,7,8-Tetramethylchroman-2-yl) carbonylamino) -3-methyl-N- (1- (2-methylpropyl) -2-oxoethyl) butanamide (0.2 g, 2
0%; compound 1) was obtained as white crystals. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 0.33 (3H, d, J = 6.7 H
z), 0.53 (3H, d, J = 6.7 Hz), 0.83 (3H, d, J = 6.3
Hz), 0.86 (3H, d, J = 6.3 Hz), 1.32-1.36 (1H, m),
1.40 (3H, s), 1.49-1.88 (4H, m), 1.95 (3H, s), 2.0
4 (3H, s), 2.09 (3H, s), 2.25-2.56 (3H, m), 4.07-
4.14 (1H, m), 4.23 (1H, dd, J = 9.9,4.9 Hz), 6.87
(1H, d, J = 9.2 Hz), 7.52 (1H, s), 8.44 (1H, d, J
= 6.9 Hz), 9.36 (1H, s). Melting point: 165-166C.

Example 2 2-((3,5-bis (tert-
(Butyl) -4-hydroxyphenyl) carbonylamino)
-3-Methyl-N- (1- (2-methylpropyl) -2-
Oxoethyl) butanamide (Compound 2) Step 1 Instead of (S)-(+)-6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid, 3,5-
Di-tert-butyl-4-hydroxybenzoic acid (5.0 g, 1
The same operation as in Step 3 of Example 1 was carried out using 9.97 mmol) to give 2,5-dioxopyrrolidinyl 3,5-bis (tert-butyl) -4-hydroxybenzoate (2.43 g, 35 %) As white crystals. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 1.39 (18H, s), 2.84 (4
H, s), 7.82 (2H, s), 8.27 (1H, s).

Step 2 2,5-Dioxopyrrolidinyl-6-hydroxy-2,
Instead of 5,7,8-tetramethylchroman-2-carboxylate, 2,5-dioxopyrrolidinyl-3,5
Perform the same operation as in step 4 of Example 1 using -bis (tert-butyl) -4-hydroxybenzoate (0.85 g, 2.44 mmol) to obtain 2-((3,5
-Bis (tert-butyl) -4-hydroxyphenyl)
Carbonylamino) -N- (1- (2,5-dioxolanyl) -3-methylbutyl) -3-methylbutanamide (0.50 g, 42%) was obtained as a white solid. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 0.76-0.90 (12H, m), 1.
13-1.32 (2H, m), 1.38 (18H, s), 1.54-1.59 (1H, m),
2.04-2.11 (1H, m), 3.84-3.71 (4H, m), 3.97-4.04 (1
H, m), 4.22 (1H, t, J = 8.6 Hz), 4.70 (1H, d, J =
3.4 Hz), 5.93 (0.5H, s), 7.36 (0.5H, s), 7.51 (2H,
s), 7.59 (1H, d, J = 9.5 Hz), 7.99 (1H, d, J = 8.
7 Hz).

Step 3 N- (1- (2,5-dioxolanyl) -3-methylbutyl) -2-((6-hydroxy-2,5,7,8-tetramethylchroman-2-yl) carbonylamino) Instead of -3-methylbutanamide, 2-((3,5-bis (te
rt-butyl) -4-hydroxyphenyl) carbonylamino) -N- (1- (2,5-dioxolanyl) -3-methylbutyl) -3-methylbutanamide (0.50 g,
1.02 mmol), and the same operation as in step 5 of Example 1 was carried out, and the resulting mixture was used as an oil in 0.1 ml.
50 g were obtained. This was purified by silica gel column chromatography using chloroform: methanol = 9: 1 to give 2-((3,5-bis (tert-butyl) -4-hydroxyphenyl) carbonylamino) -3 as a white powder.
-Methyl-N- (1- (2-methylpropyl) -2-oxoethyl) butanamide (0.07 g, 15%; compound 2) was obtained. ¹ H-NMR (DMSO-d ₆ , 300 MHz): 0.81-0.94 (12H, m), 1.
20-1.63 (21H, m), 2.11 (1H, m), 4.10 (1H, m), 4.27
(1H, t, J = 8.4 Hz), 7.54 (2H, s), 8.09 (1H, d, J
= 7.3 Hz), 8.40 (1H, d, J = 7.6 Hz), 9.39 (1H,
s). Melting point: 213-215 C.

Test Example 1 Measurement of Calpain Inhibitory Activity (Test Method) The activity of μ-calpain (Nacalai Tesque) was described in the literature [Anal. Biochem., 208, 387-392 (1993).
Year)). That is, 200 μL of a reaction solution containing casein (0.5 mg / mL), 50 mM tris (hydroxymethyl) aminomethane hydrochloride (pH 7.4), dithiothreitol (10 mM), and μ-calpain (0.03 enzyme unit) are used. 2.5 μL of a dimethyl sulfoxide solution containing the test drug at a concentration of 20 mM and 20 mM
The reaction was started by adding 50 μL of an aqueous solution of calcium chloride. After reacting for 30 minutes at 30 C, 100 μL of the reaction solution
Was transferred to another container, 50 μL of purified water and 100 μL of a 50% Coomassie brilliant blue solution were added, the mixture was left at room temperature for 15 minutes, and the absorbance was measured at 595 nm. 2.5 μL of a dimethyl sulfoxide solution not containing the test drug was added, and the same treatment was performed.
Using 1 mM disodium ethylenediaminetetraacetate in place of 20 mM calcium chloride as a blank value, the inhibition rate was calculated by the following formula, and the amount required for 50% inhibition (IC ₅₀ ) was determined. Compounds 1 and 2 were used as test drugs, and leupeptin was used as a control drug. Inhibition rate = {1− (measured value−blank value) / (control value−blank value) × 100}

(Test results) Calpain inhibitory activity (IC
₅₀ ) are shown in Table 1. This test result shows that the peptidyl derivative of the present invention has calpain inhibitory activity.

[0052]

[Table 1]

Test Example 2 Free Radical Scavenging Activity (Test Method) The method of Blois et al. [Blios. MS, Nat.
ure (London), 181: 1199 (1967))
1,1-diphenyl-2-picrylhydrazyl (0.1
mM; hereinafter abbreviated as DPPH) in ethanol 0.9m
0.1 mL of the test drug dissolving solution was added to
The absorbance at 17 nm was measured. The extinction activity was determined from the rate of decrease in the absorbance in the absence of the example compound. In addition,
The test drugs used Compounds 1 and 2, and the control drug used α-tocopherol.

(Test Results) The results of the scavenging activity of the chemically stable free radical DPPH are shown in Table 2. This test result shows that the peptidyl derivative of the present invention has almost the same free radical-scavenging ability as α-tocopherol.

[0055]

[Table 2]

Formulation Example 1 Tablet Compound 1 50 mg Lactose 80 mg Starch 17 mg Magnesium stearate 3 mg Crystalline cellulose 10 mg The above ingredients were used as a material for one tablet to form a tablet by a conventional method. The tablet may be coated with a commonly used enteric coating (such as hydroxypropylmethylcellulose phthalate), sugar coating and a film (such as ethylcellulose) as necessary.

Formulation Example 2 Capsule Compound 2 75 mg Mannit 75 mg Starch 17 mg Calcium stearate 3 mg The above components are uniformly mixed as a material for one capsule,
Granules were formed by a conventional method, and filled into hard capsules. The granules to be filled may be coated with a commonly used enteric coating (eg, hydroxypropylmethylcellulose phthalate), sugar coating, and a film (eg, ethylcellulose) as necessary.

Formulation Example 3 Suspension Injection Compound 2 750 mg Sodium carboxymethylcellulose 500 mg Distilled water for injection A total of 100 mL The above components were aseptically mixed by a conventional method to prepare a suspension injection.

Formulation Example 4 Aqueous suspension ophthalmic solution Compound 1 0.5 g Hydroxypropyl methylcellulose 0.1 g Sodium chloride 0.9 g Sodium dihydrogen phosphate dihydrate 0.1 g Benzalkonium chloride 0.005 g 0.1 N Sodium hydroxide Appropriate amount (pH 7.2) Purified water Total 100 mL Hydroxypropyl methylcellulose was heated and dispersed in about 80 mL of purified water, then cooled to room temperature and dissolved. Sodium chloride and sodium dihydrogen phosphate were added to this solution. -Dihydrate and benzalkonium chloride were added and dissolved, and 0.1N sodium hydroxide was added to adjust the pH to 7.2. Compound 1 was added to this solution, and the mixture was uniformly suspended with a homogenizer. Add purified water, total volume 100mL
To prepare an aqueous suspension ophthalmic solution.

[0060]

Since the compound of the present invention has both free radical-scavenging activity and calpain inhibitory activity, a medicament containing the compound is advantageous as an agent for preventing and treating cell damage induced by free radicals and calpain. Available to Accordingly, various diseases caused by cytotoxicity induced by free radicals and calpain, such as cataract, retinal disorder, genetic degenerative disease, macular degeneration, ocular ischemia, angiogenic disease, glaucoma, ischemia reperfusion It is useful as an agent for preventing and treating damage, optical damage, damage due to eye surgery, ischemic neuronal damage, and the like.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61K 31/35 603 A61K 31/35 603 C07D 307/84 C07D 307/84 307/87 307/87 311/66 311/66 F term (reference) 4C037 QA10 QA15 4C062 FF73 4C086 AA01 AA02 AA03 BA05 BA06 BA08 ZB21 ZC20 ZC41 4C206 AA01 AA02 AA03 GA07 GA25 ZC20 ZC41 4H006 AA01 AA03 AB20 BJ50 BQ10 BV72

Claims (8)

[Claims] 1. A compound of the general formula (I) [In the formula, A is an organic compound residue having free radical scavenging activity, and R ₁ and R ₂ each represent hydrogen or a lower alkyl group. Or a pharmaceutically acceptable salt thereof. 2. The derivative according to claim 1, wherein the organic compound residue is a phenolic compound, or a pharmaceutically acceptable salt thereof. 3. The derivative according to claim 2, wherein the phenolic compound is a compound selected from a phenol derivative, a benzofuran derivative and a benzopyran derivative, or a pharmaceutically acceptable salt thereof. 4. The phenol derivative is 2,6-bis (ter
t-butyl) phenol, 2,3,5-trimethylbenzene-1,4-diol, 2,3-dimethoxy-5-methyl-1,4-diol, 2,3-dimethoxy-5-methyl-6-vinyl The derivative according to claim 3, which is a compound selected from benzene-1,4-diol, or a pharmaceutically acceptable salt thereof. 5. The method according to claim 5, wherein the benzofuran derivative is 2,4,6,7-
Tetramethyloxaindan-5-ol, 3,4
5,6-tetramethyloxaindan-5-ol, 6
-Methyl-1,3-dihydroisobenzofuran-4,
The derivative according to claim 3, which is a compound selected from 5,6-triol, or a pharmaceutically acceptable salt thereof. 6. The derivative according to claim 3, wherein the benzopyran derivative is 2,5,7,8-tetramethylchroman, or a pharmaceutically acceptable salt thereof. 7. A compound of the general formula (1) [In the formula, A is an organic compound residue having free radical scavenging activity, and R ₁ and R ₂ each represent hydrogen or a lower alkyl group. Or a pharmaceutically acceptable salt thereof as an active ingredient. 8. The medicament according to claim 7, which is an agent for preventing or treating cell damage.