JPH0832704B2 - Prolyl endopeptidase inhibitor - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] (In the formula, Ar is a phenyl group or a naphthyl group, Y is a saturated or unsaturated alkylene group having 1 to 4 carbon atoms which may pass through an oxygen atom, provided that when Ar is a phenyl group, Y is a carbon atom. (Not a saturated alkylene group of the numbers 1 to 4)
The present invention relates to a prolyl endopeptidase inhibitor, which comprises a compound represented by as an active ingredient.

[Conventional technology]

As the population ages, issues related to medical care for the elderly are becoming more important. Among them, senile dementia is a serious social problem, and urgent development of effective therapeutic agents is desired.

Until now, as therapeutic agents for amnesia and dementia, cerebral circulation improving agents due to actions such as cerebral vasodilation and cerebral metabolism activating agents due to actions such as oxygen consumption enhancement have been used exclusively.

In recent years, a drug acting on the cholinergic system, thyrotropin-releasing hormone (Thyrotropin-Releasing Hormone,
Various anti-dementia agents with new actions such as TRH-like acting substances have been found.

Prolyl Endo-peptidas
e, hereinafter referred to as PEP) is known as an enzyme that acts on a physiologically active peptide containing proline and a synthetic substrate to specifically cleave the carboxyl side of proline. This enzyme is associated with memory, vasopressin (Vasopressi
Various studies have been conducted on the relationship between the inhibitory activity of this enzyme and the anti-amnestic effect by degrading n) and TRH, and as a result, it was suggested that PEP inhibitors could be therapeutic agents for dementia and amnesia. Chemistry, Vol. 55, No. 8, page 831, 1983), has been attracting attention.

So far, the compound that inhibits PEP is 2 at the C-terminus.
Amino acid derivatives having -formylpyrrolidine, 2-chloromethylcarbonylpyrrolidine and 2-diazomethylcarbonylpyrrolidine are known, but none of them has been put to practical use yet (Japanese Patent Publication Sho 60-18831).
No. 7, 60-172929).

[Problems to be Solved by the Invention]

Most compounds with conventional PEP inhibitory activity have 2 at the C-terminal.
It is an amino acid derivative having -formylpyrrolidine, 2-chloromethylcarbonylpyrrolidine, 2-diazomethylcarbonylpyrrolidine, etc., and these are difficult to put into practical use in terms of safety.

Therefore, the development of safer PEP inhibitors has been desired.

[Means for solving the problem]

As a result of investigations to find out a safer and more potent inhibitory PEP inhibitor, the present inventors have found that a certain cyclic amide derivative of proline has PEP inhibitory activity and can achieve the object. The present invention is based on such knowledge.

The compound represented by the general formula (I) of the present invention exhibits inhibitory activity against bovine brain-derived PEP, has low toxicity, and is useful as a therapeutic agent for amnesia and the like.

Examples of the acyl group in the compound of the general formula (I) of the present invention include 1-naphthylacetyl group, 2- (1-naphthyl) propionyl group, 3- (1-naphthyl) propionyl group, 4-
Examples thereof include (1-naphthyl) butanoyl group, cinnamoyl group, phenoxyacetyl group, 2-phenoxypropionyl group, 3-phenoxypropionyl group and benzyloxycarbonyl group. The most preferred of these acyl groups is a benzyloxycarbonyl group.

The compound of the general formula (I) of the present invention is a novel compound and can be produced as follows.

For example, the general formula It can be produced by reacting N-acylproline represented by the formula (wherein Ar and Y have the same meaning as described above) or a reactive functional derivative thereof with thiazolidine.

In addition, a carboxylic acid represented by the general formula Ar-Y-COOH (III) (wherein Ar and Y have the same meaning as described above) or a reactive functional derivative thereof, It can also be produced by reacting with prolylthiazolidine represented by

The compounds of the general formulas (II), (III) and (IV) used as starting materials in the production method of the present invention are all known compounds and are commercially available or easily produced by the method described in the literature. can do.

The compound of the general formula (I) of the present invention is prepared by reacting the carboxylic acid of the general formula (II) or (III) with thiazolidine or the formula (IV).
In the case of producing with prolylthiazolidine, the reaction is generally performed in the presence of a condensing agent. As such a condensing agent, a condensing agent generally used in peptide synthesis,
For example, N, N'-dicyclohexylcarbodiimide or the like is used.

In addition, the compound of the general formula (I) of the present invention can be converted to the general formula (II)
Alternatively, when it is produced using a reactive functional derivative of the compound of (III), examples of such a derivative include acid halides, acid anhydrides, mixed acid anhydrides and active esters.

The compound of the general formula (I) of the present invention can be made into various pharmaceutical preparations according to a conventional method. That is, if necessary, excipients, disintegrating agents, condensing agents, pharmaceutical additives such as lubricants, etc. are added, and various formulations are prepared by preparing in accordance with a conventional method,
For example, tablets, powders, granules, capsules and the like can be used.

When the compound of the general formula (I) of the present invention is used for the treatment of amnesia, etc., its dose is the age, body weight, sex,
Depending on the degree of symptoms, etc., it is generally 50-1000 mg for oral administration per day for adults, 1-for parenteral administration.
Used within the range of 500mg.

〔The invention's effect〕

The compound of the general formula (I) of the present invention is derived from bovine brain using N-carbobenzoxy-L-glycyl-L-proline-β-naphthylamide (hereinafter referred to as Z-Gly-Pro-β-NA) as a substrate. In a test for measuring the inhibitory activity against prolyl endopeptidase, it shows 50% inhibitory activity at a molar concentration of 2 × 10 ^{−6 to} 5 × 10 ⁻⁹ . For example, 3- (N-carbobenzoxy-L-prolyl) thiazolidine has an IC ₅₀ value of 4.57 × 10 ⁻⁹ mol.

As described above, the compound of the general formula (I) of the present invention exhibits a strong PEP inhibitory activity and low toxicity, and is therefore useful as a safe and excellent amnesia therapeutic agent.

〔Example〕

The present invention will be described in more detail with reference to the following reference examples and examples. The melting points of the compounds in Reference Examples and Examples are all uncorrected.

Reference Example 1 N-Cinnamoyl-L-proline L-prolinic acid 2.30 g and 2N-sodium hydroxide aqueous solution 10 ml
Dissolve in cinnamoyl chloride 5.
4 g and 15 ml of 2N-sodium hydroxide aqueous solution were slowly added at the same time. After the dropping, the mixture was further stirred at room temperature for 2 hours, washed with diethyl ether, and acidified by adding concentrated hydrochloric acid under ice cooling. After standing for 30 minutes, the mixture was extracted with ethyl acetate, the organic layer was washed with saturated saline and dried over anhydrous sodium sulfate.
The solvent was distilled off under reduced pressure, and the residue was recrystallized from ethyl acetate-methanol to give N-cinnamoyl-L-proline (3.68 g, 75.1%).
I got

Melting point: 180 to 182 ° C IR (KBr): ν _co 1740,1650 cm ^-1 ▲ [α] ¹⁴ _D ▼ = −64.0 ° (c = 1.0, methanol) MS (FAB): C ₁₄ H ₁₅ O ₃ N = 245.27 m / z 246 (M ⁺ +1) Elemental analysis value: (as C ₁₄ H ₁₅ O ₃ N) C% H% N% Calculated value 68.55 6.16 5.71 Actual value 68.67 6.23 5.71 Reference example 2 Reference example using corresponding raw material The reaction was performed in the same manner as in 1 to synthesize the following compounds.

(1) N- (1-naphthylacetyl) -L-proline Melting point: 180-182 ° C (Ethyl acetate) IR (KBr): ν _co 1730,1600cm ^-1 ▲ [α] ²⁰ _D ▼ = -44.7 ° (c = 0.77, dimethyl sulfoxide) MS (EI): C ₁₇ H ₁₇ O ₃ N = 283.31 m / z 283 (M ⁺ ), 239,168,141,114,70 Elemental analysis value: (as C ₁₇ H ₁₇ O ₃ N) C% H% N% Calculated value 70.06 6.05 4.94 Found value 71.75 6.13 4.89 Example 1 3- (N-carbobenzoxy-L-prolyl) thiazolidine (Compound A) 2.49 g of N-carbobenzoxy-L-proline and N-hydroxysuccinic acid Dissolve 1.15 g of imide in 20 ml of dioxane,
A solution of 2.06 g of N, N'-dicyclohexylcarbodiimide and 5 ml of dioxane was added dropwise while stirring under cooling. After standing overnight in a cold place, the precipitated crystals were filtered off, and the filtrate was concentrated under reduced pressure. The residual oily substance was dissolved in 15 ml of dimethoxyethane, and 0.89 g of thiazolidine was added dropwise while stirring under ice cooling. After leaving it overnight, the solvent was distilled off under reduced pressure, 5 ml of water was added to the residue, and the mixture was stirred for 30 minutes and then extracted with ethyl acetate. The organic layer was washed successively with 1N-hydrochloric acid, 5% aqueous sodium hydrogen carbonate solution and saturated brine, dried over anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The residue was purified by silica gel column chromatography (eluting solvent: ethyl acetate / benzene = 6/4) to give colorless needle crystals of 3- (N-carbobenzoxy-L-prolyl) thiazolidine.

Melting point: 112-114 ° C IR (KBr): ν _co 1700,1650cm ^-1 ▲ [α] ²⁴ _D ▼ = -15.30 ° (c = 0.5, methanol) MS (EI): C ₁₆ H ₂₀ O ₃ N ₂ S = 320.39 m / z 320 (M ⁺ ), 204,160,91 Elemental analysis value: (as C ₁₆ H ₂₀ O ₃ N ₂ S) C% H% N% S% Calculated value 59.99 6.29 8.75 9.99 Measured value 59.93 6.27 8.72 10.08 Example 2 Using the corresponding starting materials, the same reactions as in Example 1 were carried out to synthesize the following compounds.

(1) 3- (N-Cinnamoyl-L-prolyl) thiazolidine (Compound B) Melting point: 150 to 152 ° C (ethyl acetate) IR (KBr): ν _co 1650cm ^-1 ▲ [α] ¹⁹ _D ▼ = +16.98 ° (c = 1.0, methanol) MS (FAB): C ₁₇ H ₂₀ O ₂ N ₂ S = 316.35 m / z 317 (M ⁺ +1), 228,185,131 Elemental analysis value: (As C ₁₇ H ₂₀ O ₂ N ₂ S ) C% H% N% S% Calculated value 64.54 6.37 8.86 10.12 Measured value 65.43 6.38 8.90 10.06 (2) 3- [N- (1-naphthylacetyl) -L-prolyl] thiazolidine (Compound C) IR (neat): ν _co 1740,1658cm ^-1 ▲ [α] ²⁰ _D ▼ ＝ -12.3 ° (c = 0.60, ethyl acetate) MS (EI): C ₂₀ H ₂₂ O ₂ N ₂ S = 354.40 m / z 354 (M ⁺ ) , 279,239,168,141,70 elemental _{analysis: (C 20 H 22 O 2} N 2 as _{S · 1 / 3H 2 O)} C% H% N% S% calculated 66.65 6.19 7.77 8.88 Found 66.76 6.45 7.52 8.50 example 3 PEP Inhibitory activity measurement experiment Using Z-Gly-Pro-β-NA as substrate for bovine brain-derived PEP The inhibitory activity was measured.

(Measurement method) Contains 10 mM EDTA and 10 mM 2-mercaptoethanol 20
mM Tris-HCl buffer (20 mM-Tris HCl Buffer, pH = 7.0)
100 μl of PEP (about 0.14 u / ml) in 0.7 ml and each concentration (0, 1
0 ^-9 to 10 ^-4 M) was added to the test compound solution (100 µl), and preincubation (Preincubat) was performed at 37 ° C for 5 minutes.
ion). Then, each concentration (5.0, 2.5, 1.25, 0.625, 0.3125 m) dissolved in 100 μl of 40% dioxane was added to this.
M) substrate was added, and incubation was again carried out at 37 ° C. for 15 minutes to allow the enzymatic reaction to proceed. Stop the reaction with 25% trichloroacetic acid, centrifuge at 3000 rpm for 10 minutes, collect 0.5 ml of the supernatant, add 0.5 ml of 0.1% nitrous acid, and after 3 minutes 0.05%. Of N- (1-naphthyl) ethylenediamine dihydrochloride in ethanol was added. After leaving the mixed solution at 37 ° C for 25 minutes, the absorbance at 570 nm was measured, and the oxygen activity at each concentration was calculated by the following formula,
The 50% inhibitory concentration (IC ₅₀ value) was determined from each activity value.

Enzyme activity unit (μmol / min / ml) = ΔOD × 0.42 × dilution ratio (result) Compound IC ₅₀ value Compound A 4.57 nM Compound B 1.8 μM Example 4 Preparation Various preparations are prepared according to the following formulations. In addition,
The type and formulation of the dosage form are not limited to those given as the preparation examples.

(A) Powder formulation Prescription compound A 25 g lactose 975 g Total amount 1000 g or more are well mixed to prepare a powder.

(B) Powder formulation Formula A Compound 5 g Lactose 495 g A total amount of 500 g or more is mixed well to prepare a powder.

(C) Tablets Formulation Compound A 25 g Lactose 140 g 6% HPC Lactose 110 g Vilechostarch 20 g Talc stearate 5 g Total amount 300 g or more are mixed well and tableted to produce 1000 tablets.

(D) Tablet Formulation Compound A 5 g Lactose 150 g 6% HPC Lactose 120 g Vilechostarch 20 g Talc stearate 5 g Total amount 300 g or more are mixed well and tableted to produce 1000 tablets.

(E) Capsule prescription Compound A 25g Lactose 220g Vilechostarch 50g Talc stearate 5g Total amount 300g or more well mixed and filled into hard capsules
Make 1000 capsules.

(E) Capsule formulation Compound A 25g Lactose 235g Vilechostarch 55g Talc stearate 5g Total amount 300g or more well mixed, filled into hard capsules, capsules
Make 1000 capsules.

 ─────────────────────────────────────────────────── ─── Continued Front Page (56) References JP 62-114957 (JP, A) JP 62-201877 (JP, A) JP 1-250370 (JP, A) JP 62- 148467 (JP, A) J. Neurochem. , 41 (1) 69-75 (1983)

Claims (4)

[Claims] 1. A general formula (In the formula, Ar is a phenyl group or a naphthyl group, Y is a saturated or unsaturated alkylene group having 1 to 4 carbon atoms which may pass through an oxygen atom, provided that when Ar is a phenyl group, Y is a carbon atom. (Not a saturated alkylene group of the numbers 1 to 4)
A prolyl endopeptidase inhibitor, which comprises a compound represented by as an active ingredient. 2. A formula The prolyl endopeptidase inhibitor according to claim 1, which comprises a compound represented by the formula (1) as an active ingredient. 3. General formula (In the formula, Ar is a phenyl group or a naphthyl group, Y is a saturated or unsaturated alkylene group having 1 to 4 carbon atoms which may pass through an oxygen atom, provided that when Ar is a phenyl group, Y is a carbon atom. (Not a saturated alkylene group of the numbers 1 to 4)
A prolyl thiazolidine derivative represented by. 4. A formula The prolyl thiazolidine derivative according to claim 3, which is represented by: