JPH11246436A - Connective tissue disease medicine and peptide derivative - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a medicine for preventing or treating connective tissue diseases, exhibiting more excellent effects than those in a case where the inhibitors against the enzymes are separately used, by allowing to have inhibiting actions against matrix metaprotease and cathepsin. SOLUTION: This medicine for preventing or treating connective tissue diseases has inhibitory actions against both the enzymes of matrix metaprotease(MMP) and cathepsin. Concretely, the medicine for preventing or trating connective tissue diseases contains, for example, a MMP inhibitor and a cathepsin inhibitor as active ingredients. The MMP inhibitor is preferably [4-(N- hydroxyamino)-2(R)-isobutyl-3(S)-methylsuccinyl]-L-phenylglycine-N-met hylamide, etc., and the cathepsin inhibitor is preferably L-trans-epoxysuccinyl-leucylamide(4- guanidino)butane. When a compound having inhibitory activities against both the enzymes is used as an active ingredient, a new compound of formula I [A is a group of formula II or III (Ar is phenyl); R<1> is a 1-6C alkyl; R<2> and R<3> are each H, a natural amino acid side chain or the like] is preferable.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION Matrix metalloproteases (MMPs) and cathepsins are known as enzymes that degrade protein components constituting the extracellular matrix. Excessive elevation of these enzymes is thought to be the cause of diseases involving connective tissue destruction such as osteoarthritis, rheumatoid arthritis, and osteoporosis (referred to as connective tissue diseases). Is considered to be effective in preventing or treating the above-mentioned diseases (referred to as MMP inhibitor and cathepsin inhibitor, respectively). The present invention
The present invention relates to a drug for preventing or treating a disease associated with the destruction of connective tissue, which is characterized by inhibiting the action of both enzymes.
The present invention further relates to compounds having an inhibitory effect on both enzymes of MMP and cathepsin.

[0002]

2. Description of the Related Art Cathepsins are a kind of proteolytic enzymes contained in intracellular lysosomes. In particular, it is known that cysteine proteases called cathepsin B and cathepsin L are involved in the degradation of connective tissues [Essert]. Arthritis & Rheumatism, 37 (2), 236 (1994)].
In the present specification, unless otherwise specified, these cysteine proteases involved in the degradation of connective tissue are referred to as cathepsins.

[0003] On the other hand, MMP is an enzyme classified as a Zn-dependent metalloprotease, and is also known to be involved in the degradation of connective tissue. It is not clear what role each of MMP and cathepsin plays in the degradation of connective tissue, but the optimal pH for the expression of the enzyme activity of cathepsin is more acidic than that of MMP,
Cathepsins can degrade non-collagenous parts (proteoglycans) in which MMP does not work, and it is thought that each plays a different role [Evert et al., J. Cell. Physiol, 150, 221 (1992)]. Therefore, when preventing or treating a connective tissue disease, it is preferable to inhibit both of these enzymes. However, the effect of the combination of an MMP inhibitor and a cathepsin inhibitor was reported by Battle et al. [Battle et al., Arthritis & Rheumatism, 36 (12), 170
According to 9 (1993)], in in vitro experiments, the effect of the combination of both inhibitors was not significantly different from the case of using each inhibitor alone, The effect of the combination of both inhibitors has not been clarified.

Many compounds are already known as MMP inhibitors and cathepsin inhibitors. Also, JP-A-3-86
No. 852 discloses a fluoroethyl ketone derivative having an inhibitory effect on serine protease, carboxy protease and metalloprotease.
However, a compound having an inhibitory effect on both metalloprotease and cathepsin enzymes is not known.

[0005]

SUMMARY OF THE INVENTION An object of the present invention is to provide an MM
It is an object of the present invention to provide a preventive or therapeutic agent for connective tissue disease, characterized by inhibiting both P and cathepsin enzymes.

[0006]

Means for Solving the Problems The inventors of the present invention inhibit both MMP and cathepsin enzymes to prevent or prevent connective tissue diseases by using inhibitors of each enzyme alone. As a result, the present invention has been completed.

[0007]

BEST MODE FOR CARRYING OUT THE INVENTION One of the embodiments of the present invention is a drug (mixture) containing two components of an MMP inhibitor and a cathepsin inhibitor as active ingredients.
Examples of the MMP inhibitors that can be used in this form include those described in the following gazettes. JP-A-7-101925 U.S. Pat. No. 4,743,587 International Publication WO92 / 09556 US Pat. No. WO93 / 09090 US Pat. No. WO93 / 24449 US Pat. No. WO95 / 04033 European Patent Publication 489579 US Pat. Publication No. 575844 International Publication WO90 / 05719 Publication No. WO94 / 21625 Publication No. WO94 / 24140 British Publication GB2268934 Japanese Patent Laid-Open No. 6-145148 Patent Publication No. 7-157470 International Publication WO95 / 47535 Patent Publication WO97 / No. 20824 No. WO97 / 18194 No. WO96 / 33172 Unexamined Japanese Patent Publication No. Hei 9-188631 Unexamined Japanese Patent Publication No. Hei 6-256293

Among them, the following compounds are preferred. [4- (N-hydroxyamino) -2 (R) -isobutyl-3 (S) -methylsuccinyl] -L-phenylglycine-N-methylamide (JP-A-7-101925) [4- (N-hydroxyamino ) -2 (R) -Isobutyl-3-methylsuccinyl] -L-3- (5,6,7,
8-Tetrahydro-1-naphthyl) alanine-N-methylamide (Japanese Patent Application Laid-Open No. Hei 9-188633) [4- (N-hydroxyamino) -2 (R) -isobutyl-3 (S)-(2-thienylthiomethyl) Succinyl] -L-phenylalanine-N-methylamide (WO 90/05719) 3 (S) -N-hydroxy-4- (4-((pyrido-4
- yl) oxy) benzenesulfonyl) -2,2-dimethyl - tetrahydro-2H-1,4-thiazine-3-carboxamide (International Publication WO97 / No. ^{20824) N 2 - [3 (} S) - hydroxy-4- (N-hydroxyamino) -2 (R) -isobutylsuccinyl] -L-te
rt-leucine-N ¹ -methylamide (GB GB
Further, in the present embodiment, examples of the cathepsin inhibitor that can be used in combination with the MMP inhibitor include those described in the following documents or patent publications. JP-A-55-153778 JP-A-63-253061 JP-A-2-268145 JP-A-5-178758 JP-A-5-345754 JP-A-5-230004 JP-A-5-246968 Arthritis & Rheumatism, 37 ( 2), 236 (1994) Among them, the following compound (E-64) is preferable.

L-trans-epoxysuccinyl-leucylamido (4-guanidino) butane (JP-A-55-15)
No. 3778) Further, as another embodiment of the present invention, the following general formula (I)

[0010]

Embedded image (Wherein A represents the following formula (II) or (III)

[0011]

Embedded image R ¹ represents a C ₁ -C ₆ alkyl group; R ² and R ^{3 each} represent a natural α-amino acid side chain or a hydrogen atom whose functional group may be protected; ₁
-C ₆ alkyl group or C ₆ -C ₁₂ aryl (C ₁ -C ₆
An alkyl) group; Ar represents a phenyl group which may be mono-, di-, tri-, tetra- or penta-substituted by chlorine, fluorine and / or bromine atoms. ) Or a pharmaceutically acceptable salt thereof as an active ingredient.
Here, the C ₁ -C ₆ alkyl group means a linear or branched alkyl group having 1 to 6 carbon atoms, such as a methyl group, an ethyl group, a propyl group, an isopropyl group, an n-butyl group, Examples thereof include an isobutyl group, a sec-butyl group, a tert-butyl group, an n-pentyl group, an isopentyl group, an n-hexyl group, and an isohexyl group. Examples of the natural α-amino acid side chain include a methyl group (alanine), a carboxymethyl group (aspartic acid), a carbamoylmethyl group (asparagine), a carboxyethyl group (glutamic acid), a carbamoylethyl group (glutamine), and a guanidinopropyl group (arginine ), Mercaptomethyl group (cysteine), 4-imidazoylmethyl group (histidine), sec-butyl group (isoleucine), isobutyl group (leucine), 4-aminobutyl group (lysine), 2-methylthioethyl group (methionine) , Hydroxymethyl group (serine), 1-hydroxyethyl group (threonine), p-hydroxybenzyl group (tyrosine),
Examples thereof include an isopropyl group (valine), a benzyl group (phenylalanine), and a 3-indolylmethyl group (tryptophan). [Corresponding amino acids are shown in parentheses. ]. Further, functional groups such as a hydroxyl group, a carboxy group, a mercapto group, an amino group, and a guanidino group present in these side chains may be protected groups usually used in peptide chemistry (for example, Nobuo Izumiya et al., Basics and Experiments on Peptide Synthesis, Maruzen
Issued by Co., Ltd., 1985]. C ₆
And a C ₁₂ aryl group may be substituted with a hydroxyl group,
It means a monocyclic or bicyclic group having 6 to 12 carbon atoms having aromatic properties, and examples thereof include a phenyl group, a p-hydroxyphenyl group, a 1-naphthyl group, and a 2-naphthyl group.

Some of the compounds represented by the general formula (I) have stereoisomers, but in the present specification, unless otherwise specified, all possible stereoisomers or a mixture thereof are included. .

Specifically, the following compounds can be exemplified. Compound 1. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Alaninal compounds2. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-leucyl-L-alaninal compound3. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Leucinal compounds 4. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-glycinal compound5. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Alanyl [(2,6-dichlorobenzoyl) oxy] methyl ketone compound6. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-glycyl [(2,6-dichlorobenzoyl) oxy] methyl ketone compound 7. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-1-naphthylalanyl-glycyl [(2,6-dichlorobenzoyl) oxy] methyl ketone compound 8. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-glycyl [pentafluorobenzoyloxy] methyl ketone compound9. [4- (N-hydroxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Asparagyl [(2,6-dichlorobenzoyl) oxy] methyl ketone

These are compounds having an inhibitory effect on both the cathepsin and MMP enzymes, and can be produced, for example, by the following method. The types of protecting groups for amino acids or peptides used in the following reactions, conditions for introduction and deprotection, etc. are described in literatures [for example, Nobuo Izumiya et al., Basics and Experiments of Peptide Synthesis, Maruzen
1985). Among the compounds represented by the general formula (I), the compound (Ia) in which A is represented by the atomic group (II) can be synthesized according to the following scheme 1.

[0015]

Embedded image (Wherein Bzl represents a benzyl group, and R ¹ , R ² and R
³ is the same as above. That is, compound (IV) is converted to tetrahydrofuran (hereinafter referred to as T
Abbreviated as HF. ), In a solvent such as ether 0.5 to
Using 5.0 moles of lithium aluminum hydride, -1
The compound (V) is obtained by stirring at 0 ° C. to room temperature for 5 minutes to 2 hours. Next, the compound (Ia) can be produced by performing hydrogenolysis in the presence of a catalyst such as palladium carbon (hereinafter abbreviated as Pd-C) under a hydrogen stream or under pressure. Hydrocracking, methanol,
In lower alcohols such as ethanol, if necessary, water, hydrochloric acid, acetic acid, N, N-dimethylformamide (hereinafter referred to as DM
Abbreviated as F. ) May be added. When R ² or R ³ is a side chain of a natural amino acid protected with a protecting group, the protecting group is cleaved and removed according to a conventional method, if necessary, before the final step or the above-mentioned hydrogenolysis. The compound (Ia) of the present invention obtained by the above method can be converted into a pharmaceutically acceptable salt thereof by a conventional method. Next, a method for producing a starting compound will be described. The starting compound (IV) in Scheme 1 can be produced by the following method.

[0016]

Embedded image (In the formula, Z represents a benzyloxycarbonyl group,
R ¹ , R ² , R ³ and Bzl are the same as described above. First, 1-hydroxybenzotriazole (hereinafter, H
Abbreviated as OBt. The dipeptide (VI) is reacted with N, O-dimethylhydroxylamine in the presence of a condensing agent to obtain a compound (VII). The reaction is DMF, T
In an aprotic solvent such as HF or dichloromethane,
Condensing agents used for peptide synthesis, for example, dicyclohexylcarbodiimide (hereinafter abbreviated as DCC), 1
The reaction is usually performed at 0 ° C. to room temperature for 2 to 24 hours using -ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (hereinafter abbreviated as WSC) or the like. The molar ratio of the compound to be subjected to the reaction is usually 1.0 mol of N, O-dimethylhydroxylamine to 1 mol of dipeptide (VI).
To 1.5 mol, condensing agent 1.0 to 1.5 mol, HOBt
1.0 to 1.5 mol. Then, compound (VII)
To remove the amino-protecting group (Z group) from the compound (VII)
I) is obtained. The removal of the Z group can be carried out by a method usually used for peptide bonding, that is, hydrogenolysis or acidolysis using a hydrogen bromide acetic acid solution or the like.

Next, compound (VIII) and compound (I)
X) is condensed to give compound (IV). The condensation reaction is
The reaction can be performed using DCC or WSC in an aprotic solvent such as DMF, THF, or dichloromethane. The reaction is usually performed at 0 ° C. to room temperature for 2 to 24 hours,
The molar ratio of the compound to be subjected to the reaction is generally 1.0 to 1.5 mol of (IX) and 1.0 to 1.5 mol of the condensing agent, relative to 1 mol of compound (VIII). Compound (IX) can be produced according to the method described in JP-A-7-101925. 2. The compound (Ib) in which A is represented by the atomic group (III) among the compounds represented by the general formula (I) can be synthesized according to the following scheme 2.

[0018]

Embedded image (In the formula, Ar, Bzl, R ¹ , R ² and R ³ are the same as described above.) That is, the compound (X) is aromatic in a solvent such as DMF, THF and dioxane in the presence of a base such as potassium fluoride. The compound (XI) is obtained by stirring with an aromatic carboxylic acid at room temperature to 60 ° C for 2 to 24 hours. The molar ratio of the compound to be subjected to the reaction is usually 1.0 to 1.5 mol of the aromatic carboxylic acid and 1.0 to 5.0 mol of the base per 1 mol of the compound (X).
Is a mole. Next, the compound (XI) is hydrogenolyzed to produce the compound (Ib). Hydrocracking can be performed by the method described above. still,
When R ² or R ³ is a side chain of a natural amino acid protected by a protecting group, the protecting group is cleaved and removed according to a conventional method, if necessary, before the final step or the above-mentioned hydrogenolysis. Next, a method for producing a starting compound will be described. The starting compound (X) can be produced by the following method.

[0019]

Embedded image (In the formula, R represents a tert-butoxycarbonyl group or a benzyloxycarbonyl group, and Et represents an ethyl group. Bzl, R ¹ , R ² and R ³ are the same as described above.) Amino group of compound (XII) The compound (XIII) is obtained by removing the protecting group (R). Removal of the protecting group can be performed by a method usually used for peptide synthesis. That is, in the case of a tert-butoxycarbonyl group, an acid such as trifluoroacetic acid is used, and in the case of a benzyloxycarbonyl group, hydrogenation or acid decomposition is used.

Next, compound (XIII) and compound (I
After condensing X) to obtain compound (XIV), compound (XV) is obtained by alkaline hydrolysis. The alkali hydrolysis is performed at 0 ° C. to room temperature for 1 to 5 hours by adding an alkali such as sodium hydroxide using water or dioxane as a solvent. The molar ratio used for the reaction is 1.0 to 3.0 mol of alkali relative to 1 mol of compound (XIV). Then, a chloroformate such as ethyl chlorocarbonate is added to the compound (XV) in a solvent such as THF in the presence of a base such as triethylamine, and the mixture is added at -10 ° C to 0 ° C for 5 to 1 hour.
After reacting for 5 minutes, a diazomethane / ether solution is added, and the mixture is reacted at 0 ° C. to room temperature for 5 to 30 minutes. Then, a hydrogen bromide acetic acid solution is added, and the mixture is stirred at 0 ° C. to room temperature for 5 minutes to 1 hour to obtain a compound ( X) is obtained. The molar ratio for the reaction is 1.0 to 1.5 mol of triethylamine and 1.0 mol of chlorocarbonate ethyl ester per 1 mol of compound (XV).
0 to 1.5 moles of diazomethane and hydrogen bromide are required in excess.

The medicament of the present invention is orally or parenterally administered to a human. Oral dosage forms include tablets, granules,
It includes liquid preparations such as syrups and soft capsules in addition to solid preparations such as powders, fine granules and hard capsules. Such preparations can be manufactured in a conventional manner, and tablets, granules, powders, or fine granules may be mixed with MMP inhibitors and cathepsin inhibitors, for example, lactose, starch, crystalline cellulose, magnesium stearate, hydroxypropylcellulose, talc. Manufactured by mixing with commonly used pharmaceutical additives such as
Hard capsules are prepared by appropriately filling the above-mentioned fine granules or powders into capsules. Syrups are prepared by dissolving or suspending an MMP inhibitor and a cathepsin inhibitor in an aqueous solution containing sucrose, carboxycellulose, and the like. Soft capsules are prepared using a lipid excipient, such as vegetable oil, oily emulsion, or glycol. And an MMP inhibitor and a cathepsin inhibitor are dissolved or suspended therein and filled into soft capsules.

[0022] Parenteral dosage forms include injections,
Suppositories such as suppositories and vaginal suppositories, and nasal administration agents such as sprays are exemplified. These preparations can be manufactured by conventional methods,
For example, injections are produced by dissolving or emulsifying MMP inhibitors and cathepsin inhibitors in physiological saline or lipid excipients, for example, vegetable oils, oily emulsions, glycols and the like, and aseptically encapsulating them in ampules or vials. . The dose of the drug of the present invention depends on its active ingredient, namely, M
In the case of a mixture consisting of an MP inhibitor and a cathepsin inhibitor and in the case of a compound having inhibitory activity on both enzymes,
Although it varies depending on the dosage form, age, sex or weight of the patient, or symptoms, generally, the active ingredients of the MMP inhibitor and the cathepsin inhibitor are each 0.1 to 50%.
0 mg / kg body weight / day, preferably 1 to 200 mg / k
g body weight per day, with both inhibitors once or twice daily
Administer in 4 divided doses. In the case of the compound of the present invention, since it exerts an inhibitory effect on MMP and cathepsin alone, it differs depending on the inhibitory effect on both enzymes.
0.1-500 mg / kg body weight / day, preferably 1-2
An appropriate amount is 00 mg / kg body weight / day, which is administered once a day or divided into 2 to 4 times.

[0023]

EFFECT OF THE INVENTION The MMP inhibitor and the cathepsin inhibitor show an effect of inhibiting bone resorption alone in an experiment using mice, but an additive effect of inhibiting bone resorption is observed when both inhibitors are used in combination ( Test Example 1), no toxicity was observed when used in combination. The compound of the present invention alone has an inhibitory effect on both MMP and cathepsin enzymes (see Test Example 2). Therefore, a drug containing both the MMP inhibitor and the cathepsin inhibitor as active ingredients and a drug containing the compound of the present invention as an active ingredient are both useful as preventive or therapeutic drugs for connective tissue diseases. Hereinafter, the effects of the drug and the compound of the present invention will be described with reference to test examples. <Test Example 1> Bone resorption inhibitory effect When breeding with a Ca-deficient feed is continued, bone collagen degradation is promoted to compensate for the lack of Ca, and bone loss is induced (experimental osteoporosis model). As a result, the inhibitory effect on increasing urinary hydroxyproline excretion was tested. Test compound Known compound X: [4- (N-hydroxyamino) -2
(R) -isobutyl-3 (S) -methylsuccinyl]-
L-phenylglycine-N-methylamide (Japanese Unexamined Patent Publication No.
No. 101925) Known compound Y: L-trans-epoxysuccinyl-leucylamide (4-guanidino) butane (E-64; JP-A-55-153778) Test method Four-week-old ddY male mice were grouped into 5 mice. Except for the untreated group, on the third day after starting breeding on a Ca-deficient feed (Oriental Yeast Co., Ltd .; calcium 0.0105% to 0.012%, phosphorus 0.62%), the test compounds and / or
Alternatively, a placebo was administered and urine was collected overnight.

[0024]

[Table 1]

The amount of urine, creatinine and hydroxyproline were quantified by the following method, the amount of urinary hydroxyproline was calculated as the ratio of creatinine, and the significance of each group was tested by Tukey's method. Creatinine Test _Wako (Wako Pure Chemical _Industries ) was used for quantification of creatinine.
Hydroxyproline is obtained by hydrolyzing urine with 6N hydrochloric acid and then using an ion exchange column (Hitachi Gel #) with a high performance liquid chromatograph.
2619), reacted with o-phthalaldehyde to form a hydroxyproline derivative, and measured using a fluorometer. Test results are shown in FIG.

Compound X and Compound Y each alone increase the amount of excreted hydroxyproline,
It is clear that the combined use of both compounds further suppresses the excretion of hydroxyproline. <Test Example 2> Inhibitory effect on MMP and cathepsin The inhibitory effect on MMP-1, cathepsin B and L was tested. 1. Test compound Compound of the present invention A [4- (N-hydroxyamino) -2
R-isobutylsuccinyl] -L-phenylalanyl-
L-alaninal Compound of the present invention B [4- (N-hydroxyamino) -2
R-isobutylsuccinyl] -L-leucyl-L-alaninal Compound of the present invention C ... [4- (N-hydroxyamino) -2
R-isobutylsuccinyl] -L-phenylalanyl-
L-Leucinal Compound of the present invention D ... [4- (N-hydroxyamino) -2
R-isobutylsuccinyl] -L-phenylalanine-
Alanyl [(2,6-dichlorobenzoyl) oxy] methyl ketone Control compound Known compound X, Known compound Y (Test Example 1
Refer to) 2. Test method The measuring method of MMP-1 inhibition is as follows. MMP
-1 used was obtained and purified by the method described in JP-A-3-103178. The test compound was dissolved in a 50 mM Tris-HCl buffer containing a buffer for measurement (0.2 M salt, 5 mM calcium chloride, 0.05% by volume bridge-35 (polyoxyethylene lauryl ether) and 0.02% by volume sodium azide). , PH 7.5) and a known amount of MMP-1 solution were mixed, and fluorescein isothiocyanate-labeled type I collagen (manufactured by Cosmo Bio) was used as a substrate. Japanese Journal of Inflamation, Vol. 4, 123, 198
MMP-1 activity (A) was measured according to 4 years). On the other hand, MMP-
1) The activity (B) was measured, the inhibition rate was calculated by the following formula: inhibition rate (%) = (1−A / B) × 100, and the IC ₅₀ value was determined from the relationship between the concentration of the test compound and the inhibition rate. . The method for measuring cathepsins B and L is as follows. 1 mM ethylenediaminetetraacetic acid, 2 mM dithiothreitol and 0.05% by volume
Synthetic substrate (carbobenzoxy-phenylalanyl-arginyl-7-amide-4-methylcoumarin) and cathepsin B (human) in 100 mM acetate buffer (pH 5.5) containing Bridge-35 (polyoxyethylene lauryl ether) Liver-derived; purchased from Cosmo Bio or Cathepsin L (derived from human kidney; purchased from Cosmo Bio) was reacted at 30 ° C. for 10 minutes. The reaction was stopped by adding an acetate buffer (pH 5.5) containing monochloroacetic acid, and the enzyme activity was measured from the fluorescence intensity (excitation wavelength Ex = 375 nm, fluorescence measurement wavelength Em = 445 nm) in the supernatant.
The IC ₅₀ value was determined in the same manner as in P-1. (2) Test results Table 2 shows the measurement results (IC ₅₀ ) of the inhibitory activities of MMP-1, cathepsin B and L.

[0027]

[Table 2]

[0028]

The present invention will be described in more detail with reference to the following Examples and Reference Examples. Reference Example 1 [4- (N-benzyloxyamino) -2 (R) -iso
Butylsuccinyl] -L-phenylalanyl-L-ara
Nyl-N-methyl-N-methoxyamide [general formula (I
In V), R ¹ is an isobutyl group, R ² is a benzyl group, R
³ is a methyl manufacture of, compound] group (1) N-benzyloxycarbonyl -L- phenylalanyl -L- alanine -N- methyl -N- methoxy amide In the general formula (VII) R ² is a benzyl group, Compound in which R ³ is a methyl group]; N-benzyloxycarbonyl-L-phenylalanyl-L-alanine 1.7 g
After dissolving 10 ml of MF, 890 mg of WSC, 630 mg of 1-hydroxybenzotriazole monohydrate (hereinafter abbreviated as HOBt), 450 mg of N, O-dimethylhydroxylamine hydrochloride, and 470 of triethylamine
mg was added and stirred at room temperature overnight. After acidification with dilute hydrochloric acid, the mixture was extracted with ethyl acetate. After washing the organic layer with water, an aqueous solution of sodium hydrogen carbonate, water and saturated saline,
Dried over magnesium sulfate. After removing magnesium sulfate, the residue obtained by evaporating the solvent was recrystallized from ethyl acetate to obtain 1.0 g of the title compound as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ; 1.23 (3H, d, J = 7.0
Hz), 2.68 (1H, dd, J = 11.2Hz, 13.6Hz), 3.00 (1H, dd, J = 3.3H
z, 13.6Hz), 3.12 (3H, s), 3.74 (3H, s), 4.24-4.35 (1H, m), 4.
68-4.80 (1H, m), 4.93 (2H, s), 7.15-7.36 (10H, m), 7.44 (1H,
d, J = 8.8 Hz), 8.29 (1 H, d, J = 7.5 Hz). (2) [4- (N-benzyloxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Alanine-N-methyl-N-methoxyamide; N-benzyloxycarbonyl-L-phenylalanyl-L-
15 ml of methanol was added to alanyl-N-methyl-N-methoxyamide, and the mixture was stirred under a hydrogen atmosphere using 100 mg of 10% Pd-C for 1.5 hours. After removing Pd-C, 5 ml of DMF was added to the residue obtained by evaporating the solvent, and 4- (N-benzyloxyamino) -2 (R) -isobutylsuccinic acid (produced according to JP-A-101925). 440 mg and WSC 305 mg were added, and the mixture was stirred at room temperature overnight. After acidification with dilute hydrochloric acid, the mixture was extracted with ethyl acetate. The organic layer was washed with water, an aqueous solution of sodium hydrogen carbonate, water and saturated saline, and then washed with magnesium sulfate. After removing magnesium sulfate, the solvent was distilled off to obtain 0.29 g of the title compound as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.2
Hz), 0.79 (3H, d, J = 6.2Hz), 0.80-1.00 (1H, m), 1.20 (3H, d, J
= 7.0Hz), 1.26-1.40 (2H, m), 1.83-1.90 (2H, m), 2.60-2.70
(1H, m), 2.78 (1H, dd, J = 10.5Hz, 13.9Hz), 3.05 (1H, dd, J = 4.
0Hz, 13.9Hz), 3.11 (3H, s), 3.72 (3H, s), 4.48-4.57 (1H, m),
4.68-4.78 (1H, m), 4.74 (2H, s), 7.10-7.28 (5H, m), 7.37 (5
H, s), 8.04 (1H, d, J = 7.2Hz), 8.10 (1H, d, J = 8.4Hz), 10.94 (1
H, s). Reference Example 2 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-leucyl-L-alanyl-N
Production of -methyl-N-methoxyamide [compound of the general formula (IV), wherein R ¹ is an isobutyl group, R ² is an isobutyl group, and R ³ is a methyl group] N-benzyloxycarbonyl-L-leucyl-L-alanine -N-methyl-N-methoxyamide [general formula (V
II) a compound in which R ² is an isobutyl group and R ³ is a methyl group], and reacts according to Reference Example 1 from 1.0 g;
0.5 g of the title compound was obtained as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ; 0.80 (6H, d, J = 6.4
Hz), 0.85 (6H, d, J = 6.4Hz), 0.98-1.10 (1H, m), 1.17 (3H, d, J
= 7.0Hz), 1.35-1.52 (4H, m), 1.56-1.70 (1H, m), 1.99 (1H, d
d, J = 7.0Hz, 14.4Hz), 2.14 (1H, dd, J = 7.4Hz, 14.4Hz), 2.70-
2.80 (1H, m), 3.10 (3H, s), 3.71 (3H, s), 4.24-4.34 (1H, m),
4.63-4.71 (1H, m), 4.73 (1H, d, J = 11.0Hz), 4.77 (1H, d, J = 1
1.0Hz), 7.32-7.40 (5H, m), 7.83 (1H, d, J = 7.4Hz), 8.01 (1H,
d, J = 8.3 Hz), 11.00 (1H, s). Reference Example 3 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-glycyl-N-methyl- Production of N-methoxyamide [compound of the general formula (IV), wherein R ¹ is an isobutyl group, R ² is a benzyl group, and R ³ is a hydrogen atom] N-benzyloxycarbonyl-L-phenylalanyl-L-glycyl- The reaction was carried out according to Reference Example 1 from 0.92 g of N-methyl-N-methoxyamide [compound of the general formula (VII), wherein R ² is a benzyl group and R ³ is a hydrogen atom] to obtain 485 mg of the title compound as a colorless solid. Was.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.2
Hz), 0.78 (3H, d, J = 6.2Hz), 0.87-0.99 (1H, m), 1.22-1.40 (2
H, m), 1.79-1.95 (2H, m), 2.60-2.70 (1H, m), 2.81 (1H, dd, J =
10.2Hz, 13.8Hz), 3.03-3.13 (1H, m), 3.10 (3H, m), 3.69 (3H,
s), 3.93-4.11 (2H, m), 4.51-4.60 (1H, m), 4.73 (2H, s), 7.11
-7.29 (5H, m), 7.37 (5H, s), 8.04 (1H, t, J = 5.5Hz), 8.13 (1H,
d, J = 8.5 Hz), 10.95 (1H, s). Reference Example 4 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alanyl [(2 , 6-Dichlorobenzoyl) oxy] methyl ketone [In the general formula (XI), R ¹ is an isobutyl group,
Production of compound in which R ² is a benzyl group, R ³ is a methyl group, and Ar is a 2,6-dichlorophenyl group] (1) [4- (N-benzyloxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Production of alanine ethyl ester [compound of the general formula (XIV), wherein R ¹ is an isobutyl group, R ² is a benzyl group, and R ³ is a methyl group] N-benzyloxycarbonyl-L-phenylalanyl-L-alanine ethyl A mixed solvent of 50 ml of methanol and 50 ml of THF was added to 5.0 g of the ester, and 10%
The mixture was stirred under a hydrogen atmosphere using 0.5 g of Pd-C. P
After removing dC, the residue obtained by distilling off the solvent was added to DMF.
30 ml, 4- (N-benzyloxyamino) -2R-
3.5 g of isobutyl succinic acid and 2.4 g of WSC were added, and the mixture was stirred overnight at room temperature. After acidification with dilute hydrochloric acid, the mixture was extracted with a mixed solvent of ethyl acetate and THF. The organic layer was washed with water, an aqueous potassium carbonate solution, water, and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate, the solvent was distilled off to obtain 5.0 g of the title compound.
Was obtained as a colorless solid.

¹ H-NMR (CDCl ₃ ) δ; 0.84 (6H, t, J = 6.2H
z), 1.26 (3H, t, J = 7.2Hz), 1.32 (3H, d, J = 7.2Hz), 1.10-1.30
(1H, m), 1.40-1.60 (2H, m), 2.10-2.30 (1H, m), 2.65-2.80 (1
H, m), 3.09 (2H, d, J = 7.0Hz), 4.15 (2H, q, J = 7.2Hz), 4.39-4.
52 (1H, m), 4.57-4.68 (1H, m), 4.86 (2H, s), 6.43 (1H, d, J = 7.
9 Hz), 6.51 (1H, d, J = 5.7 Hz), 7.16-7.40 (10H, m), 8.48 (1H,
s).

(2) [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alanyl bromomethyl ketone [In the general formula (X), R ¹ is an isobutyl group. , A compound wherein R ² is a benzyl group and R ³ is a methyl group]; [4- (N-benzyloxyamino) -2R-isobutylsuccinyl]-
To 1.0 g of L-phenylalanyl-L-alanine ethyl ester, 10 ml of dioxane was added, 5 ml of a 1N aqueous sodium hydroxide solution was added, and the mixture was stirred at room temperature for 3 hours.
After acidification with dilute hydrochloric acid, the mixture was extracted with ethyl acetate.
The organic layer was washed with water and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate, 12 ml of THF and 220 mg of triethylamine were added to the residue obtained by evaporating the solvent. Ethyl chlorocarbonate at 0 ° C 240
mg was added and stirred for 5 minutes. Five minutes after the addition of the excess diazomethane / ether solution, a 25% hydrogen bromide acetic acid solution was added, followed by stirring for 30 minutes. After adding water, the mixture was extracted with ethyl acetate. The organic layer was washed with water and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate,
The solvent was distilled off to obtain 0.48 g of the title compound.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.2
Hz), 0.79 (3H, d, J = 6.2 Hz), 0.90-1.02 (1 H, m), 1.16 (3H, d, J
= 7.0Hz), 1.25-1.40 (2H, m), 1.90 (1H, dd, J = 7.7Hz, 14.4H
z), 2.02 (1H, dd, J = 7.1Hz, 14.4Hz), 2.60-2.70 (1H, m), 2.85
(1H, dd, J = 9.4Hz, 13.9Hz), 3.03 (1H, dd, J = 5.8Hz, 13.9Hz),
4.25 (2H, s), 4.30-4.37 (1H, m), 4.42-4.51 (1H, m), 4.74 (2
H, s), 7.13-7.30 (5H, m), 7.37 (5H, s), 8.25 (1H, d, J = 7.8H
z), 8.49 (1H, d, J = 6.9 Hz), 11.01 (1H, s). (3) [4- (N-benzyloxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-L
-Alanyl [(2,6-dichlorobenzoyl) oxy]
Preparation of methyl ketone; [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alanyl bromomethyl ketone 0.4
8 g was dissolved in 10 ml of DMF, and potassium fluoride 150
mg, 2,6-dichlorobenzoic acid 190 mg,
Stirred overnight at room temperature. After adding ethyl acetate and THF,
The extract was washed with an aqueous solution of potassium carbonate, water, diluted hydrochloric acid, water, and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate, the solvent was distilled off to obtain 0.34 g of the title compound as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ; 0.72 (3H, d, J = 6.2
Hz), 0.79 (3H, d, J = 6.2 Hz), 0.90-1.02 (1 H, m), 1.21 (3H, d, J
= 7.1Hz), 1.25-1.40 (2H, m), 1.91 (1H, dd, J = 7.8Hz, 14.6H
z), 2.04 (1H, dd, J = 6.8Hz, 14.6Hz), 2.60-2.70 (1H, m), 2.86
(1H, dd, J = 9.2Hz, 13.8Hz), 3.06 (1H, dd, J = 5.5Hz, 13.8Hz),
4.33-4.43 (1H, m), 4.46-4.57 (1H, m), 4.74 (2H, s), 4.91 (1
H, d, J = 17.1Hz), 5.00 (1H, d, J = 17.1Hz), 7.12-7.29 (5H, m),
7.37 (5H, s), 7.53-7.64 (3H, m), 8.26 (1H, d, J = 8.0Hz), 8.48
(1H, d, J = 7.1 Hz), 11.02 (1H, s). Reference Example 5 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-glycyl [( Production of 2,6-dichlorobenzoyl) oxy] methyl ketone [compound of formula (XI) wherein R ¹ is an isobutyl group, R ² is a benzyl group, R ³ is a hydrogen atom, and Ar is a 2,6-dichlorophenyl group] 1) [4- (N-benzyloxyamino) -2 (R)
-Isobutylsuccinyl] -L-phenylalanyl-glycyl ethyl ester [in the general formula (XIV), R
¹ is an isobutyl group, R ² is a benzyl group and R ³ is a hydrogen atom] Preparation of 5.0 g of N-tert-butoxycarbonyl-L-phenylalanine-glycine ethyl ester with 15 ml of 90% aqueous trifluoroacetic acid The mixture was stirred under ice cooling for 40 minutes. After the trifluoroacetic acid was distilled off, the mixture was extracted with dichloromethane. The organic layer was washed with an aqueous solution of potassium carbonate, water, and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate, the solvent was distilled off, and the resulting residue was treated with 30 ml of DMF, 4.0 g of 4- (N-benzyloxyamino) -2 (R) -isobutylsuccinic acid, WSC
2.8 g was added, and the mixture was stirred at room temperature overnight. After adding dilute hydrochloric acid to make the pH acidic, the mixture was extracted with a mixed solvent of ethyl acetate and THF. The organic layer was washed with water, an aqueous potassium carbonate solution, water, and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate, the solvent was distilled off to obtain 5.2 g of the title compound.

(2) [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-glycyl bromomethyl ketone [In the formula (X), R ¹ is an isobutyl group, R Compound in which ² is a benzyl group and R ³ is a hydrogen atom]; [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alanine ethyl ester 1.1 g The reaction was carried out according to (2) of Reference Example 4 to obtain 0.47 g of the title compound. (3) [4- (N-benzyloxyamino) -2 (R)
Preparation of -isobutylsuccinyl] -L-phenylalanyl-glycyl [(2,6-dichlorobenzoyl) oxy] methyl ketone; [4- (N-benzyloxyamino)
-2 (R) -isobutylsuccinyl] -L-phenylalanyl-glycyl bromomethyl ketone The reaction was carried out according to (3) of Reference Example 4 from 0.47 g of the title compound 0.1.
13 g were obtained.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.2
Hz), 0.79 (3H, d, J = 6.2Hz), 0.89-1.02 (1H, m), 1.23-1.40 (2
H, m), 1.83-2.02 (2H, m), 2.62-2.70 (1H, m), 2.86 (1H, dd, J =
9.7Hz, 13.9Hz), 3.07 (1H, dd, J = 6Hz, 13.9Hz), 4.07 (2H, d, J
= 5.3Hz), 4.50-4.60 (1H, m), 4.74 (2H, s), 5.10 (2H, s), 7.12
-7.28 (5H, m), 7.37 (5H, m), 7.53-7.64 (3H, m), 8.23 (1H, d, J
= 8.2 Hz), 8.35 (1 H, t, J = 5.3 Hz), 10.98 (1 H, s). Reference Example 6 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-1 -Naphthylalanyl-glycyl [(2,6-dichlorobenzoyl) oxy] methyl ketone [in the general formula (XI), R ¹ is an isobutyl group,
R ² is a 1-naphthylmethyl group, R ³ is a hydrogen atom, and Ar is a 2,6-dichlorophenyl group] Production of N-tert-butoxycarbonyl-L-1-naphthylalanyl-glycine ethyl ester 1.65 g The reaction was carried out according to Reference Example 5 to obtain 0.48 g of the title compound.

¹ H-NMR (DMSO-d ₆ ) δ; 0.72 (3H, d, J = 6.2
Hz), 0.78 (3H, d, J = 6.2Hz), 0.90-1.02 (1H, m), 1.23-1.40 (2
H, m), 1.87 (1H, dd, J = 1.4Hz, 13.7Hz), 2.00 (1H, dd, J = 6.8H
z, 13.7Hz), 2.62-2.75 (1H, m), 3.30-3.40 (1H, m), 3.57 (1H,
dd, J = 5.2Hz, 14.2Hz), 4.05 (2H, s), 4.62-4.72 (1H, m), 4.74
(2H, s), 5.11 (2H, s), 7.30-7.40 (8H, m), 7.48-7.65 (5H, m),
7.73-7.80 (1H, m), 7.90 (1H, d, J = 7.7Hz), 8.20 (1H, d, J = 7.9
Hz), 8.33-8.40 (1H, m), 11.00 (1H, s). Reference Example 7 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-asparagyl (Tert-butyl) [(2,6-dichlorobenzoyl) oxy] methyl ketone [In the general formula (XI), R
¹ is an isobutyl group, R ² is a benzyl group, R ³ is a tert-butoxycarbonylmethyl group, and Ar is a 2,6-dichlorophenyl group] Production of N-benzyloxycarbonyl-L-phenylalanyl-L-aspara According to Reference Example 4, 0.12 g of the title compound was obtained from 1.8 g of gill (tert-butyl) ethyl ester.

¹ H-NMR (DMSO-d ₆ ) δ; 0.72 (3H, d, J = 6.2
Hz), 0.78 (3H, d, J = 6.2Hz), 0.90-1.00 (1H, m), 1.30-1.40 (2
H, m), 1.38 (9H, s), 1.90 (1H, dd, J = 8.3Hz, 14.6Hz), 2.01 (1
H, dd, J = 6.9Hz, 14.6Hz), 2.40-2.50 (1H, m), 2.62-2.72 (1H,
m), 2.73 (1H, m), 2.86 (1H, dd, J = 9.4Hz, 13.9Hz), 3.02 (1H, d
d, J = 5.8Hz, 13.9Hz), 4.48-4.56 (1H, m), 4.60-4.67 (1H, m),
4.74 (2H, s), 4.97 (2H, s), 7.10-7.28 (5H, m), 7.38 (5H, s),
7.52-7.62 (3H, m), 8.30 (1H, d, J = 7.8Hz), 8.69 (1H, d, J = 7.9
Hz), 11.01 (1H, s). Reference Example 8 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-glycyl [pentafluorobenzoyloxy] methyl ketone [general formula (XI) wherein R ¹ is an isobutyl group, R ² is a benzyl group, R ³ is a hydrogen atom, and Ar is a pentafluorophenyl group] [4- (N-benzyloxyamino) -2 (R)- Isobutylsuccinyl] -L-phenylalanyl-glycyl bromomethylketone (compound of Reference Example 5 (2))
29 g was dissolved in 3 ml of DMF, and potassium fluoride 90m
g and pentafluorobenzoic acid (140 mg) were added, and the mixture was stirred overnight at room temperature. After adding dilute hydrochloric acid, the mixture was extracted with ethyl acetate. The organic layer was washed with an aqueous solution of potassium carbonate, water, and saturated saline, and then dried over magnesium sulfate. After removing magnesium sulfate, the residue obtained by distilling off the solvent is
On LC (developing solution; chloroform / methanol = 20 /
Purification in 1) gave 0.12 g of the title compound. ¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.3 Hz), 0.78 (3H,
d, J = 6.3Hz), 0.88-1.00 (1H, m), 1.22-1.43 (2H, m), 1.85-2.
02 (2H, m), 2.61-2.69 (1H, m), 2.88 (1H, dd, J = 10.0Hz, 13.9H
z), 3.10 (1H, dd, J = 4.8Hz, 13.9Hz), 4.05 (2H, d, J = 5.4Hz),
4.47-4.57 (1H, m), 4.74 (2H, s), 5.15 (2H, s), 7.15-7.28 (5
H, m), 7.30-7.40 (5H, m), 8.24 (1H, d, J = 8.1Hz), 8.42 (1H, t,
J = 5.4 Hz), 8.77 (1H, s), 11.00 (1H, s). Reference Example 9 [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L -Leucine-N-methyl-N-methoxyamide [general formula (I
In V), R ¹ is an isobutyl group, R ² is a benzyl group, R
Production of a compound wherein ³ is an isobutyl group] N-benzyloxycarbonyl-L-phenylalanyl-L-leucine-N-methyl-N-methoxyamide [In the general formula (VII), R ² is a benzyl group, and R ³ is Compound as isobutyl group] was reacted according to Reference Example 1 from 1.5 g to obtain 0.3 g of the title compound as a colorless solid.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 5.2
Hz), 0.78 (3H, d, J = 5.2Hz), 0.86 (6H, t, J = 5.2Hz), 0.80-0.9
8 (2H, m), 1.25-1.65 (4H, m), 1.82-1.90 (2H, m), 2.62-2.72
(1H, m), 2.78 (1H, dd, J = 8.3Hz, 11.2Hz), 3.01 (1H, dd, J = 3.6
Hz, 11.2Hz), 3.09 (3H, s), 3.71 (2H, s), 4.48-4.58 (1H, m),
4.74 (1H, m), 4.77-4.89 (1H, m), 7.10-7.25 (5H, m), 7.30-7.
40 (5H, m), 7.95 (1H, d, J = 7.0Hz), 8.11 (1H, d, J = 7.1Hz), 10.
93 (1H, s).

Example 1[4- (N-hydroxyamino) -2 (R) -isobuty
Rusuccinyl] -L-phenylalanyl-L-alanina
[In general formula (Ia), R ¹ Is an isobutyl group, R ²
Is a benzyl group, R ³ In which is a methyl group] [4- (N-benzyloxyamino) -2 (R) -iso
Butylsuccinyl] -L-phenylalanyl-L-ara
Nin-N-methyl-N-methoxyamide [general formula (I
V)¹Is an isobutyl group, R^TwoIs a benzyl group, R
^ThreeIs a methyl group, the compound of Reference Example 1]
After adding 10 ml of THF, lithium hydride was added at -20 ° C.
35 mg of aluminum was added and stirred for 30 minutes. 2N
After acidification with aqueous HCl, extraction was performed using ethyl acetate.
Was. After the organic layer was washed with water and saturated saline,
Dried with calcium. Remove magnesium sulfate and remove solvent
The residue obtained by distillation is dissolved in 10 ml of methanol.
Was. Under a hydrogen atmosphere using 70 mg of 10% Pd-C,
Stir at room temperature for 1.5 hours. After removing Pd-C, the solvent was removed.
The solid obtained by evaporation is washed with ethyl acetate.
This gave 51 mg of the title compound as a colorless powder.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.2
Hz), 0.78 (3H, d, J = 6.2Hz), 0.90-1.30 (1H, m), 1.12 (3H, d, J
= 7.2Hz), 1.23-1.40 (2H, m), 1.85-2.02 (2H, m), 2.55-2.70
(1H, m), 2.87 (1H, dd, J = 9.6Hz, 13.7Hz), 3.05 (1H, dd, J = 5.3
Hz, 13.7Hz), 3.99-4.10 (1H, m), 4.45-4.56 (1H, m), 7.13-7.
30 (5H, m), 8.18 (1H, d, J = 8.2Hz), 8.40 (1H, d, J = 6.5Hz), 8.7
3 (1H, s), 9.25 (1H, s), 10.38 (1H, s). Example 2 [4- (N-hydroxyamino) -2 (R) -isobutyi
Rusuccinyl] -L-leucyl-L-alaninal
In the general formula (Ia), R ¹ is an isobutyl group, and R ² is an isobutyl group.
Production of a compound wherein tyl group and R ³ are methyl group] [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-leucyl-L-alanine-N
-Methyl-N-methoxyamide [the compound of Reference Example 2 in which R ¹ is an isobutyl group, R ² is an isobutyl group, and R ³ is a methyl group in the general formula (IV)] The reaction was performed to obtain 84 mg of the title compound as a colorless powder.

[0044]¹H-NMR (DMSO-d₆) δ; 0.75-0.92 (12H,
m), 0.90-1.10 (2H, m), 1.15 (3H, d, J = 7.2Hz), 1.35-1.70 (4
H, m), 1.90 (1H, dd, J = 1.5Hz, 14.3Hz), 2.16 (1H, dd, J = 7.0H
(z, 14.3Hz), 2.63-2.77 (1H, m), 4.00-4.10 (1H, m), 4.20-4.3
3 (1H, m), 8.04 (1H, d, J = 7.9Hz), 8.28 (1H, d, J = 6.6Hz), 8.70
(1H, s), 9.34 (1H, s), 10.39 (1H, s).[4- (N-hydroxyamino) -2 (R) -isobuty
Rusuccinyl] -L-phenylalanyl-L-leucine
[In general formula (Ia), R ¹ Is an isobutyl group, R ²
Is a benzyl group, R ³ In which is an isobutyl group]
Construction [4- (N-benzyloxyamino) -2R-isobutyi
Rusuccinyl] -L-phenylalanyl-L-leucine
-N-methyl-N-methoxyamide [in the general formula (IV)
Then R¹Is an isobutyl group, R^TwoIs a benzyl group, R^ThreeBut
Compound of Reference Example 9 which is a sobutyl group] from 500 mg
The reaction was conducted in the same manner as in Example 1, and 51 mg of the title compound was obtained.
Was obtained as a colorless powder.

¹ H-NMR (DMSO-d ₆ ) δ; 0.70-0.90 (12H,
m), 0.90-1.02 (2H, m), 1.22-1.62 (4H, m), 1.80-2.00 (2H, m
m), 2.60-2.70 (1H, m), 2.82-2.92 (1H, m), 2.96-3.10 (1H,
m), 4.00-4.12 (1H, m), 4.50-4.60 (1H, m), 7.13-7.30 (5H,
m), 8.22 (1H, d, J = 8.3Hz), 8.31 (1H, d, J = 7.4Hz), 8.72 (1H,
s), 9.22 (1H, s), 10.37 (1H, s). Example 4 [4- (N-hydroxyamino) -2 (R) -isobutyi
Rusuccinyl] -L-phenylalanyl-glycinal
[In the general formula (Ia), R ¹ is an isobutyl group, and R ² is
Njiru group, the production of R ³ compound is a hydrogen atom] [4- (N- benzyloxyamino) -2 (R) - isobutyl succinyl] -L- phenylalanyl - glycine -N- methyl -N- methoxy amide [Compound of Reference Example 3 wherein R ¹ is an isobutyl group, R ² is a benzyl group, and R ³ is a hydrogen atom in formula (IV)] From 325 mg, a reaction was carried out in the same manner as in the production of Example 1 to obtain 73 mg of the title compound. Obtained as a colorless powder.

¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.3
Hz), 0.77 (3H, d, J = 6.3Hz), 0.90-1.12 (1H, m), 1.22-1.40 (2
H, m), 1.83-2.01 (2H, m), 2.55-2.70 (1H, m), 2.86 (1H, dd, J =
10.0Hz, 13.8Hz), 3.08 (1H, dd, 4.9Hz, 13.8Hz), 3.78-3.93
(2H, m), 4.47-4.57 (1H, m), 7.10-7.30 (5H, m), 8.18 (1H, d, J
= 8.2Hz), 8.40 (1H, t, J = 5.4Hz), 8.75 (1H, bs), 9.35 (1H, s),
10.40 (1H, bs).

Example 5 [4- (N-hydroxyamino) -2 (R) -isobutyi
Rusuccinyl] -L-phenylalanyl-L-alanyl
[(2,6-dichlorobenzoyl) oxy] methylketo
[In the general formula (Ib), R ¹ is an isobutyl group, and R ² is
Benzyl group, R ³ is methyl group, Ar is 2,6-dichloro
Production of phenyl group compound] [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanine-L-alanyl [(2,6-dichlorobenzoyl) oxy] methyl ketone [general formula In (XI), R ¹ is an isobutyl group,
Compound in which R ² is a benzyl group, R ³ is a methyl group, and Ar is a 2,6-dichlorophenyl group; Reference Example 4] 300 mg
Was dissolved in 10 ml of methanol and 5 ml of THF, and 10%
The mixture was stirred for 2 hours under a hydrogen atmosphere using 100 mg of Pd-C. After removing Pd-C, the residue obtained by distilling off the solvent was separated and purified by HPLC to obtain 130 mg of the title compound eluted under the following conditions as a colorless solid.

HPLC conditions Column; YMC-Pack ODS-A A-312 150 × 6.0mm I.D. Moving layer; 0.05% aqueous trifluoroacetic acid / acetonitrile
= 3/2 flow rate; 1.0 ml / min Elution time: 10 minutes¹ H-NMR (DMSO-d₆) δ; 0.73 (3H, d, J = 6.2Hz), 0.78 (3H, d, J
= 6.2Hz), 0.91-1.03 (1H, m), 1.20 (3H, d, J = 7.1Hz), 1.25-1.
42 (2H, m), 1.86-2.04 (2H, m), 2.60-2.72 (1H, m), 2.86 (1H, d
d, J = 8.9Hz, 13.8Hz), 3.06 (1H, dd, J = 6.2Hz, 13.8Hz), 4.31-
4.41 (1H, m), 4.45-4.57 (1H, m), 4.91 (1H, d, J = 17.0Hz, 4.98
(1H, d, J = 17.0Hz), 7.13-7.30 (5H, m), 7.52-7.64 (3H, m), 8.
25 (1H, d, J = 8.1Hz), 8.47 (1H, d, J = 7.1Hz), 8.74 (1H, s), 10.
39 (1H, s). Example 6[4- (N-hydroxyamino) -2 (R) -isobuty
Rusuccinyl] -L-phenylalanyl-glycyl
[(2,6-dichlorobenzoyl) oxy] methylketo
[In general formula (Ib), R ¹ Is an isobutyl group, R ² But
Benzyl group, R ³ Is a hydrogen atom, Ar is 2,6-dichloro
Which is a phenyl group] [4- (N-benzyloxyamino) -2 (R) -iso
Butylsuccinyl] -L-phenylalanine-glycyl
[(2,6-dichlorobenzoyl) oxy] methylketo
[In general formula (XI), R¹Is an isobutyl group, R^TwoBut
Benzyl group, R ^ThreeIs a hydrogen atom, Ar is 2,6-dichloro
Compound which is a phenyl group; Reference Example 5]
Dissolved in a mixed solvent of 15 ml of ethanol and 5 ml of THF.
Using 75 mg of 0% Pd-C under a hydrogen atmosphere for 2 hours
Stirred. After removing the catalyst, the solvent
The separation and purification using HPLC
10mg of the title compound eluted as a colorless solid
Was.

HPLC conditions Column; YMC-Pack ODS-A A-312 150 × 6.0 mm ID moving bed; 0.05% aqueous trifluoroacetic acid / acetonitrile = 3/2 flow rate; 1.0 ml / min Elution time; 18 minutes ¹ H-NMR (DMSO-d ₆ ) δ; 0.73 (3H, d, J = 6.3 Hz), 0.78 (3H,
d, J = 6.3Hz), 0.90-1.00 (1H, m), 1.23-1.42 (2H, m), 1.83-2.
00 (2H, m), 2.60-2.70 (1H, m), 2.86 (1H, dd, J = 10.1Hz, 13.8H
z), 3.08 (1H, dd, J = 4.9Hz, 13.8Hz), 4.07 (2H, d, J = 5.4Hz),
4.49-4.59 (1H, m), 5.11 (2H, s), 7.14-7.30 (5H, m), 7.54-7.
66 (3H, m), 8.23 (1H, d, J = 8.1Hz), 8.37 (1H, t, J = 5.4Hz), 10.
40 (1H, s). Example 7 [4- (N-hydroxyamino) -2 (R) -isobutyi
Rusuccinyl] -L-1-naphthylalanyl-glycyl
[(2,6-dichlorobenzoyl) oxy] methylketo
[In the general formula (Ib), R ¹ is an isobutyl group, and R ² is
1-naphthylmethyl group, R ³ is a hydrogen atom, Ar is 2,6
Production of [4- (N-benzyloxyamino) -2R-isobutylsuccinyl] -L-phenylalanyl-glycyl [(2,6-dichlorobenzoyl) oxy] methyl ketone [general formula (XI )), R ¹ is an isobutyl group, R ² is a 1-naphthylmethyl group, R ³ is a hydrogen atom, and Ar is 2,6.
-A compound which is a dichlorophenyl group; Reference Example 6] 70 m
g in a mixed solvent of 10 ml of methanol and 5 ml of THF, and 20 mg of 10% Pd-C under a hydrogen atmosphere.
Stir for 4.5 hours. After removing Pd-C, the residue obtained by distilling off the solvent was separated and purified using HPLC to obtain 8 mg of the title compound eluted under the following conditions as a colorless solid.

HPLC conditions Column; YMC-Pack ODA-A A-312 150 × 6.0 mm ID moving bed; 0.05% aqueous trifluoroacetic acid / acetonitrile = 1/1 flow rate; 1.0 ml / min Elution time; 14 minutes ^{1 H-NMR (DMSO-d} 6) δ; 0.72 (3H, d, J = 6.3Hz), 0.77 (3H,
d, J = 6.3Hz), 0.90-1.02 (1H, m), 1.22-1.40 (2H, m), 1.87 (1
H, dd, J = 1.7Hz, 13.9Hz), 1.99 (1H, dd, J = 6.8Hz, 13.9Hz), 2.
60-2.70 (1H, m), 3.35-3.45 (1H, m), 3.61 (1H, dd, J = 4.7Hz, 1
4.0Hz), 4.05 (2H, s), 4.60-4.70 (1H, m), 5.12 (2H, s), 7.35-
7.44 (2H, m), 7.50-7.66 (5H, m), 7.75-7.81 (1H, m), 7.92 (1
(H, d, J = 8.0Hz), 8.20 (1H, d, J = 8.2Hz), 8.33-8.43 (1H, m), 1
0.4 (1H, s).

Example 8 [4- (N-hydroxyamino) -2 (R) -isobutyi
Rusuccinyl] -L-phenylalanyl-asparagyl
[(2,6-dichlorobenzoyl) oxy] methylketo
[In the general formula (Ib), R ¹ is an isobutyl group, and R ² is
Benzyl group, R ³ is carboxymethyl group, Ar is 2,6
Production of [4- (N-benzyloxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-asparagyl (tert-butyl) [(2,6-dichlorobenzoyl) ) Oxy] methylketone [R in the general formula (XI)
Compound in which ¹ is an isobutyl group, R ² is a benzyl group, R ³ is a tert-butoxycarbonylmethyl group, and Ar is a 2,6-dichlorophenyl group; Reference Example 7] To 0.12 g, 5 ml of 90% aqueous trifluoroacetic acid was added. And stirred at room temperature for 50 minutes. The residue obtained by distilling off trifluoroacetic acid was extracted with chloroform. The organic layer was washed with water and saturated saline, and then dried over magnesium sulfate. The residue obtained by removing magnesium sulfate was dissolved in 10 ml of methanol, and 50 mg of 10% Pd-C was used under a hydrogen atmosphere to obtain 1 residue.
Stirred for hours. After removing Pd-C, the residue obtained by evaporating the solvent was separated and purified using HPLC to obtain 23 mg of the title compound eluted under the following conditions as a colorless solid.

[0052] HPLC Conditions Column; YMC-Pack ODS-A A -312 6.0 × 150mm ID mobile phase; 0.05% aqueous trifluoroacetic acid / acetonitrile = 3/2 flow rate; 1.0 ml / min elution time; 16 minutes ¹ H-NMR (DMSO-d ₆ ) δ; 0.72 (3H, d, J = 6.2 Hz), 0.78 (3H,
d, J = 6.2Hz), 0.90-1.00 (1H, m), 1.25-1.42 (2H, m), 1.83-2.
01 (2H, m), 2.45-2.55 (1H, m), 2.60-2.70 (1H, m), 2.75 (1H, m
m), 2.86 (1H, dd, J = 9.2Hz, 13.7Hz), 3.02 (1H, dd, J = 5.8Hz, 1
(3.7Hz), 4.48-4.62 (2H, m), 5.00 (2H, s), 7.15-7.29 (5H, m),
7.52-7.63 (3H, m), 8.29 (1H, d, J = 7.9Hz), 8.65 (1H, d, J = 7.7
Hz), 10.39 (1H, s), 12.46 (1H, s).

Example 9 [4- (N-hydroxyamino) -2 (R) -isobutyi
Rusuccinyl] -L-phenylalanyl-glycyl [pe
Nantafluorobenzoyloxy] methyl ketone [general formula
In (Ib), R ¹ is an isobutyl group, and R ² is benzyl
Group, R ³ is a hydrogen atom, Ar is a pentafluorophenyl group
Preparation of a compound] is [4-(N-benzyloxyamino) -2 (R) - isobutyl succinyl] -L- phenylalanyl - glycyl [R ¹ pentafluoro benzoyloxy] methyl ketone [general formula (XI) is Compound in which isobutyl group, R ² is benzyl group, R ³ is hydrogen atom, and Ar is pentafluorophenyl group; Reference Example 8] 0.12 g of methanol 1
The mixture was dissolved in 0 ml, and stirred for 1 hour under a hydrogen atmosphere using 50 mg of 10% Pd-C. After removing Pd-C, the residue obtained by evaporating the solvent was separated and purified using HPLC to obtain 26 mg of the title compound eluted under the following conditions as a colorless solid.

HPLC condition Column; YMC-Pack ODS-A A-312, 150 × 6.0 mm ID moving bed; 0.05% aqueous trifluoroacetic acid / acetonitrile = 1/1 flow rate; 1.0 ml / min Elution time; 0.5 min ¹ H-NMR (DMSO-d ₆ ) δ; 0.72 (3H, d, J = 6.3 Hz), 0.77 (3H,
d, J = 6.3Hz), 0.88-1.00 (1H, m), 1.22-1.42 (2H, m), 1.83-2.
00 (2H, m), 2.60-2.70 (1H, m), 2.86 (1H, dd, J = 10.2Hz, 13.8H
z), 3.08 (1H, dd, J = 4.7Hz, 13.8Hz), 4.05 (2H, d, J = 5.4Hz),
4.47-4.57 (1H, m), 5.15 (2H, s), 7.15-7.28 (5H, m), 8.24 (1
H, d, J = 8.1Hz), 8.42 (1H, t, J = 5.4Hz), 8.77 (1H, s), 10.39 (1
H, s).

Example 10 Production of tablets [4- (N-hydroxyamino)
-2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alaninal (Compound A) 100 mg is obtained. [Operation] The main drug, corn starch and microcrystalline cellulose are mixed, and hydroxypropylcellulose dissolved in 50 parts by weight of water is added thereto and kneaded sufficiently. The kneaded product is passed through a sieve, granulated on granules, and dried. Then, the obtained granules are mixed with magnesium stearate and compressed into 250 mg tablets.

Example 11 Preparation of Granules [4- (N-Hydroxyamino) -2
(R) -isobutylsuccinyl] -L-phenylalanyl-L-alaninal (compound A) -containing granules are obtained. [Operation] A main drug, lactose and corn starch are mixed, and hydroxypropylcellulose dissolved in 120 parts by weight of water is added and kneaded sufficiently. The kneaded product is passed through a 20-mesh sieve, granulated, dried and sized, and 500 mg
A granule containing 200 mg of the active ingredient is obtained.

Example 12 Production of Capsules [4- (N-Hydroxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alaninal (Compound A) in one capsule as follows 100m
A capsule containing g is obtained. [Operation] Thoroughly mix the above components, and mix 2
A capsule is obtained by filling the capsule with the amount of 00 mg.

Example 13 Preparation of Injection A mixture of 0.5 parts by weight of [4- (N-hydroxyamino) -2 (R) -isobutylsuccinyl] -L-phenylalanyl-L-alaninal and 5 parts by weight of sorbit Was dissolved in distilled water for injection to make up to 100 parts by weight, and this aqueous solution was filtered through a membrane filter. The filtrate was filled into ampoules purged with nitrogen in an amount of 5 g each, sealed and then sealed at 120 ° C.
[4- (N-hydroxyamino) -2 (R) -isobutylsuccinyl] in one ampoule
An injection containing 25 mg of -L-phenylalanyl-L-alaninal is obtained.

[Brief description of the drawings]

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 shows the effects of administration of compounds X and Y alone and in combination on the urinary hydroxyproline excretion (ratio to creatinine) of mice bred on a Ca-deficient diet (Test Example). 1) was indicated.

[Explanation of symbols]

The bar graphs a, b, c, d, and e show the urinary hydroxyproline excretion (ratio to creatinine) of the following groups, respectively. a: untreated group b: control group c: compound X administration group d: compound Y administration group e: compound X, Y combination group Further, a significant difference test between each group was performed by Tukey's method.
The results are indicated by *. * P <0.01

Claims (5)

[Claims] 1. A drug for preventing or treating a connective tissue disease, which has an inhibitory activity against both a matrix metalloprotease and a cathepsin enzyme. 2. The drug according to claim 1, which comprises two components, a matrix metalloprotease inhibitor and a cathepsin inhibitor, as active ingredients. 3. The drug according to claim 1, wherein a compound having an inhibitory activity against both a matrix metalloprotease and a cathepsin is used as an active ingredient. 4. A compound having an inhibitory activity on both a matrix metalloprotease and a cathepsin enzyme,
The following general formula (I): (Wherein A represents the following formulas (II) and (III): R ¹ represents a C ₁ -C ₆ alkyl group; R ² and R ^{3 each} represent a natural α-amino acid side chain or a hydrogen atom whose functional group may be protected; ₁
-C ₆ alkyl group or C ₆ -C ₁₂ aryl (C ₁ -C ₆
An alkyl) group; Ar represents a phenyl group which may be mono-, di-, tri-, tetra- or penta-substituted by chlorine, fluorine and / or bromine atoms. 4. The agent according to claim 3, which is a compound represented by the formula or a pharmaceutically acceptable salt thereof. 5. A compound represented by the following general formula (I): (Wherein A represents the following formulas (II) and (III): R ¹ represents a C ₁ -C ₆ alkyl group; R ² and R ^{3 each} represent a natural α-amino acid side chain whose functional group may be protected or a hydrogen atom; C
₁ -C ₆ alkyl or C ₆ -C ₁₂ aryl (C ₁ -C
_{6 represents} an alkyl) group; Ar represents a mono-, di-, tri-,
Represents a phenyl group which may be tetra- or penta-substituted. Or a pharmaceutically acceptable salt thereof.