JPH06192293A - Cyclic pentapeptide, its production and use thereof - Google Patents

Abstract

PURPOSE:To obtain the cyclic pentapeptide having a strong endothelin receptor- antagonizing action and useful as circulatory function-improving agents or as medicines for preventing or treating cardiac infraction, acute renal failure or asthma. CONSTITUTION:A cyclic pentapeptide of the general formula: cyclo(-A-B-C-D-E-) [A is D-acidic alpha-amino acid residue; B and D are L-alpha-amino acid residus; C is D-neutral-alpha-amino acid residue; E is aromatic ring group-having D-alpha-amino acid residue, but when A is D-Glu, B is L-Ala, C is D-aIle, and D is L-Leu, L-Val or L-Nva; or when A is D-Glu, B is L-Ala, C is D-Val or D-Leu, and D is L-Leu or L-Val; or when A is D-Glu, B is L-Ala, C is D-Nva, and D is L-Val or L-Nva, E is not D-Trp, and when A is D-Glu, B is L-Ala, C is D-aIle and D is L-Leu, E is not D-mTrp] or its pharmacologically acceptable salt.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a novel cyclic pentapeptide having an endothelin antagonism which is expected to be useful as a medicine such as a therapeutic agent for hypertension, a therapeutic agent for cardio-cerebral circulation disease, a therapeutic agent for renal disease, a therapeutic agent for asthma and the like. , Its manufacturing method and its use.

[0002]

BACKGROUND OF THE INVENTION Endothelin (ET) is a vasoconstrictor peptide consisting of 21 amino acids, which was isolated and structurally determined from the culture supernatant of porcine aortic endothelial cells by Yanagisawa et al. In 1988 (Yanagisawa et al., Nature. (Nature), Volume 332,
411-412). After that, the study of the gene encoding endothelin revealed the existence of peptides having a structure similar to endothelin, and endothelin-1 (ET-1) and endothelin-2 (ET-, respectively).
2), which is named endothelin-3 (ET-3), and its structure is as follows (the constituent amino acids in ET-1, 2, 3 are all L-forms).

[Chemical 1] [Inoue et al., Proc. Nat of the National Academy of Sciences USA
Acad. Sci. USA), 86, 2863-2867.
page〕. The endothelin family peptides are present in the body and are expected to be an endogenous factor involved in circulatory system regulation because they have a vasoconstrictor action, hypertension, cardio-cerebral circulation disease (for example, myocardial infarction). ), And the relationship with renal disease (for example, acute renal failure) is estimated. It also has a bronchial smooth muscle contraction effect, and its relationship with asthma has been estimated.

[0003]

It is considered that if an antagonist of the above endothelin family peptide is obtained, it will be useful not only for elucidating the action mechanism of the peptide, but also as an effective therapeutic drug for these diseases. To be
So far, a cyclic pentapeptide TAN-1 having an endothelin antagonistic activity obtained from a fermentation product.
462A, B, TAN-1477C, D, E, F, G,
H, I, J and K have been filed (Japanese Patent Application No. 2-413828 and Japanese Patent Application No. 3-126160).
It is an object of the present invention to provide a novel cyclic pentapeptide derivative having a similar or higher effect.

[0004]

[Means for Solving the Problems] The inventors of the present invention have made extensive studies using the action of suppressing the strong vascular smooth muscle contractile activity of endothelin as an index, and have synthesized various cyclic pentapeptides. The inventors have found that the derivative has a remarkable endothelin antagonism and completed the present invention. That is, the present invention is: General formula Cyclo (-A-B-C-D-E-) (I) [In formula, A is D-acidic-alpha-amino acid residue, B and D are L-alpha-amino acid residue, C Indicates a D-neutral-α-amino acid residue, and E indicates a D-α-amino acid residue having an aromatic ring group. However, A is D-Glu, B is L-Ala, and C is Da.
Ile is D is L-Leu, L-Val or L-Nva, or A is
D-Glu, B is L-Ala, C is D-Val or D-Leu, D
Is L-Leu or L-Val, or A is D-Glu and B is
When L-Ala is C-D-Nva and D is L-Val or L-Nva, E is not D-Trp but A is D-Glu and B is L-Ala.
When C is D-aIle and D is L-Leu, E is D-mTrp
Not. ] The cyclic pentapeptide represented by or its pharmacologically acceptable salt,

2. General formula H- (X) -OH (II) [In formula, X is -A-B-C-D-E-, -B-C-D-.
E-A-, -C-D-E-A-B-, -D-E-A-B
Represents -C- or -E-A-B-C-D-, A, B,
C, D and E have the same meaning as described in claim 1. ]
A cyclic pentapeptide represented by the general formula (I), which comprises treating the peptide represented by or a reactive derivative of the peptide with a cyclizing agent, and subjecting the peptide to a removal reaction of a protecting group, if necessary, or A method for producing a pharmacologically acceptable salt thereof, and

3. The present invention relates to an endothelin receptor antagonist containing a cyclic pentapeptide represented by the general formula (I) or a pharmacologically acceptable salt thereof. In the above general formula (I), examples of the amino acid constituting the D-acidic-α-amino acid residue represented by A include amino acids having an acidic group such as a carboxyl group or a sulfonyl group in the side chain. , And more specifically, glutamic acid,
Examples thereof include aspartic acid and cysteic acid. In the general formula (I), the amino acids constituting the L-α-amino acid residues represented by B and D include, for example, glycine, alanine, valine, norvaline, leucine, isoleucine, t-leucine, norleucine, methionine. , Α
-Aminobutyric acid, serine, threonine, phenylalanine, aspartic acid, glutamic acid, lysine, arginine, tyrosine, proline and the like, and commonly known amino acids are listed. In the general formula (I), as the amino acid constituting the D-neutral-α-amino acid residue represented by C, for example, valine, leucine, isoleucine, alloisoleucine, norleucine, phenylglycine, t-
Examples thereof include leucine and γ-methylleucine. In the above general formula (I), D having an aromatic cyclic group represented by E
Examples of the amino acids constituting the -α-amino acid residue include amino acids having an aromatic ring group in the side chain, and specific examples thereof include tryptophan, 5-methyltryptophan, phenylalanine, tyrosine, 1-naphthylalanine, 2 - naphthylalanine, N ⁱⁿ - formyl tryptophan, and the like.

In this specification, when amino acids and peptides are represented by abbreviations, IUPAC-IUB Co
It is based on the abbreviations used in mmision on Biochemical Nomenclature or the abbreviations commonly used in this field, and examples are given below. Gly: glycine Ala: alanine Val: valine Nva: norvaline Ile: isoleucine aIle: alloisoleucine Leu: leucine tLeu: tertiary leucine γMeLeu: gammamethylleucine Met: methionine Arg: arginine Hys Lys: lysine lysine: lysine: lysine: lysine: lysine: lysine: lysine: lysine: lysine: lysine: lysine: isys: Glutamic acid Ser: Serine Thr: Threonine Phe: Phenylalanine Tyr: Tyrosine Trp: Tryptophan mTrp: 5-Methyltryptophan Phg: Phenylglycine Nal (1): 1-naphthylalanine Nal (2): 2-naphthylalanine Also commonly used in the text. The protecting groups and reagents used are indicated by the following abbreviations. Boc: t-butoxycarbonyl Bzl: benzyl BrZ: 2-bromobenzyloxycarbonyl ClZ: 2-chlorobenzyloxycarbonyl Tos: p-toluenesulfonyl For: formyl OBzl: benzyl ester OPac: phenacyl ester ONB: HONB ester TFA: tri. Fluoroacetic acid TEA: triethylamine IBCF: isobutylchloroformate DMF: N, N-dimethylformamide DCC: N, N'-dicyclohexylcarbodiimide DCU: N, N'-dicyclohexylurea HONB: N-hydroxy-5-norbornene-2,
3-dicarboximide HOBt: 1-hydroxybenzotriazole DCM: dichloromethane THF: tetrahydrofuran

As the pharmacologically acceptable salt of the cyclic pentapeptide (I) of the present invention, sodium salt, potassium salt,
Calcium salts, magnesium salts, inorganic acid addition salts such as hydrochlorides, sulfates, phosphates, acetates, propionates,
Examples thereof include organic acid salts such as citrate, tartrate, malate, and oxalate. Cyclic pentapeptide (I) of the present invention
Can be produced by conventional means of peptide synthesis. That is, either the liquid phase synthesis method or the solid phase synthesis method may be used, but the liquid phase synthesis method may be preferable in some cases. The means for synthesizing such a peptide may be according to any known method, for example, M.
Bodansky and MA Ondetti, Peptide Synthesis, Interscience, New York, 1966; FMFinn and K. Hofman.
n, The Proteins, Volume 2, H. N
Enrath, edited by RL Hill, Academic Press, Inc., New York, 1976; Nobuo Izumiya et al., "Basics and Experiments of Peptide Synthesis," Maruzen Co., Ltd. 1985; Haruaki Yajima,
Shunpei Sakakibara et al., Biochemistry Experiment Course 1, edited by The Biochemical Society of Japan, Tokyo Chemistry Doujin 1977; Shun Kimura et al.
Stewart and JD Young, Solid Phase Peptide Synthes.
is), Pierce Chemical Company, Illinois, 1984
Years, etc., such as azide method, chloride method, acid anhydride method, mixed acid anhydride method, DCC method, active ester method, method using Woodward reagent K, carbonyl imidazole method, redox method, DCC / HONB method, B
Examples thereof include a method using an OP reagent.

The cyclic pentapeptide (I) of the present invention corresponds to a raw material having a reactive carboxyl group corresponding to one of two kinds of fragments that are bisected at any position of the peptide bond, and the other fragment. A raw material having a reactive amino group is condensed by a conventional method for peptide synthesis, and then the protecting groups of the C-terminal α-carboxyl group and the N-terminal α-amino group of the product are removed simultaneously or stepwise, and then both of them are subjected to known condensation. By a method, the compound is condensed in the molecule to obtain a cyclic compound, and when the product has a protecting group, it can be produced by removing the protecting group by a conventional method.

Protection of a functional group which should not be involved in the reaction of the starting material, protection of the protective group, elimination of the protective group, activation of the functional group involved in the reaction and the like can be appropriately selected from known ones or means. Examples of the protecting group for the amino group of the raw material include carbobenzoxy, t-butyloxycarbonyl, t-amyloxycarbonyl, isobornyloxycarbonyl, 4-methoxybenzyloxycarbonyl, 2-chlorobenzyloxycarbonyl, adamantyloxycarbonyl. , Trifluoroacetyl, phthalyl, formyl, 2-nitrophenylsulfenyl, diphenylphosphinothioyl, 9-fluorenylmethyloxycarbonyl and the like. Examples of the protecting group for the carboxyl group include alkyl ester (eg, ester group such as methyl, ethyl, propyl, butyl, t-butyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, 2-adamantyl), benzyl ester, 4- Examples thereof include nitrobenzyl ester, 4-methoxybenzyl ester, 4-chlorobenzyl ester, benzhydryl ester, phenacyl ester, carbobenzoxyhydrazide, t-butyloxycarbonylhydrazide and tritylhydrazide.

The hydroxyl group of serine can be protected by, for example, esterification or etherification. Suitable groups for this esterification include, for example, lower alkanoyl groups such as acetyl group, aroyl groups such as benzoyl group,
Examples thereof include groups derived from carbonic acid such as a benzyloxycarbonyl group and an ethyloxycarbonyl group. Further, as a group suitable for etherification, for example, a benzyl group,
Examples include a tetrahydropyranyl group and a t-butyl group. However, the hydroxyl group of serine does not necessarily need to be protected. Examples of the protective group for the phenolic hydroxyl group of tyrosine include benzyl, 2,6-dichlorobenzyl,
2-nitrobenzyl, 2-bromobenzyloxycarbonyl, t-butyl and the like can be mentioned, but they are not necessarily protected. Methionine may be protected in the form of sulfoxide. Examples of the protecting group for imidazole of histidine include p-toluenesulfonyl, 4-methoxy-
2,3,6-trimethylbenzenesulfonyl, 2,4-dinitrophenyl, benzyloxymethyl, t-butoxymethyl, t-butoxycarbonyl, trityl, t-butoxycarbonyl, trityl, 9-fluorenylmethyloxycarbonyl, etc. Although it can be given, it does not necessarily have to be protected. The protecting group for the indole of tryptophan includes formyl, 2,4,6-trimethylbenzenesulfonyl, 2,4,6-trimethoxybenzenesulfonyl, 4
-Methoxy-2,3,6-trimethylbenzenesulfonyl, β, β, β-trichloroethyloxycarbonyl,
Examples thereof include diphenylphosphinothi oil, but they do not necessarily need to be protected.

The activated carboxyl group of the raw material includes, for example, corresponding acid anhydrides, azides, active esters [alcohols (eg, pentachlorophenol, 2,
4,5-trichlorophenol, 2,4-dinitrophenol, cyanomethyl alcohol, p-nitrophenol, N-hydroxy-5-norbornene-2,3-dicarboximide, N-hydroxysuccinide, N-
Esters with hydroxyphthalimide, N-hydroxybenztriazole), and the like. Examples of the activated amino group of the raw material include the corresponding phosphoric acid amide. The condensation reaction can be carried out in the presence of a solvent. The solvent can be appropriately selected from those known to be usable in the peptide condensation reaction. For example, anhydrous or water-containing dimethylformamide, dimethylsulfoxide, pyridine, chloroform, dioxane, dichloromethane, tetrahydrofuran, acetonitrile, ethyl acetate, N-methylpyrrolidone or an appropriate mixture thereof can be used. The reaction temperature is
It is appropriately selected from the range known to be used in the peptide bond-forming reaction, and is usually appropriately selected from the range of about -20 ° C to 30 ° C.

The intramolecular cyclization reaction can be carried out at any position of the peptide by a known method. For example, first, the C-α-carboxy protecting group of the C-terminal amino acid of the protected pentapeptide is eliminated by a known method and then activated by a known method, and then the N-α-amino protecting group of the N-terminal amino acid is removed. It can be eliminated by a known method and cyclized in the molecule. Alternatively, the protected pentapeptide C
The C-α-carboxyl protecting group of the terminal amino acid and the N-α-amino protecting group of the N-terminal amino acid may be simultaneously eliminated and then cyclized in the molecule by a known condensation reaction. In some cases, it may be preferable to carry out the intramolecular cyclization reaction under high dilution. Examples of the method for removing the protective group include catalytic reduction in a hydrogen stream in the presence of a catalyst such as Pd black or Pd carbon, or anhydrous hydrogen fluoride, methanesulfonic acid, trifluoromethanesulfonic acid, trifluoro Examples thereof include acid treatment with acetic acid or a mixed solution thereof, and reduction with sodium in liquid ammonia. The leaving group reaction by the acid treatment is generally carried out at an appropriate temperature of -20 ° C to 40 ° C, but in the acid treatment, anisole, phenol, thioanisole, m-cresol, p-cresol, dimethyl sulfide, 1,4 -The addition of cation scavengers such as butanedithiol and 1,2-ethanedithiol is effective. Further, the 2,4-dinitrophenyl group used as the imidazole protecting group of histidine is removed by thiophenol treatment, and the formyl group used as the indole protecting group of tryptophan is the above-mentioned 1,2-ethanedithiol, 1,4-butane group. In addition to deprotection by acid treatment in the presence of dithiol or the like, it can be removed by alkali treatment with dilute sodium hydroxide or dilute ammonia.

After the reaction, the cyclic pentapeptide (I) thus produced is separated by means of peptide separation, for example,
It is collected by extraction, partitioning, reprecipitation, recrystallization, column chromatography, high performance liquid chromatography and the like. The cyclic pentapeptide (I) of the present invention can also be obtained as a salt such as sodium salt, potassium salt, calcium salt, magnesium salt or the like, or an acid addition salt, particularly a pharmaceutically acceptable acid addition salt, by a method known per se. Inorganic acids (eg hydrochloric acid, sulfuric acid, phosphoric acid) or organic acids
Examples thereof include salts such as acetic acid, propionic acid, citric acid, tartaric acid, malic acid, oxalic acid, and methanesulfonic acid.

Next, the pharmacological action of the cyclic pentapeptide of the present invention will be described. (1) Measurement of contractile inhibitory effect on smooth muscle of porcine coronary artery Produced from right circumflex branch of coronary artery from which adventitial connective tissue was removed
Fix a x15 mm spiral strip on a 4 ml Magnus device,
The tension was detected by a variable displacement / displacement converter UL-10GR (Minebea Co.) and recorded by a polygraph (NEC Sanei). Magnus device kept at 37 ℃, O ₂ 95%,
Krebs-Henseleit solution (composition [mM]: NaCl 118, KCl 4.7, CaCl ₂ saturated with 5% CO ₂ mixed gas)
2.5, KH ₂ PO ₄ 1.2, NaHCO ₃ 25.0, MgS
O ₄ 1.2, Glucose 10.0). Strips are 1.
A tension of 25-1.5 g is applied, equilibration is performed for 1.5 hours, 60 mM KCl administration is repeated at 30-minute intervals until the contraction response becomes constant, and equilibration is performed for another 1.5 hours, and then a measurement sample is administered. It was The strip shrinkage is 60m for each strip
The contraction response to M KCl was normalized and statistically processed. The inhibitory effect was measured at about 1 of the given concentration of the compound.
Five minutes later, 10 ^-9 M endothelin-1 was administered, and the contraction was compared with the control. The results are shown in Table 1.

[Table 1] ──────────────────────────────── Example Inhibitory effect on sample ET-1 (Control-Sample) / Control × 100 ──────────────────────────────── 1 [D-Asp ¹ ] -TAN -1462A 58.1% 2 [Asp ² ] -TAN-1462A 45.2% 6 [D-Asp ¹ , Asp ² ] -TAN-1477D 63.8% ────────────── ─────────────────── Control: Contraction by 10 ^-9 M ET-1 Sample: 10 ^-9 M ET in the presence of sample 3 × 10 ^-5 M
-1 Contraction The novel cyclic pentapeptide [I] of the present invention and some of its salts exhibited an action of suppressing endothelin-induced contraction in porcine coronary artery smooth muscle, but such an example has not been reported yet. Not not. Therefore, some of the peptide derivatives [I] or salts thereof of the present invention are
It can be used for treating hypertension, myocardial infarction, acute renal failure, etc. (eg, mouse, rat, rabbit, dog, cat, pig, human etc.).

(2) The results of measuring the binding activity of the cyclic pentapeptide of the present invention to the endothelin receptor are shown in Table 2 together with Comparative Examples. The method for measuring the binding activity of the receptor is as follows. Receptor binding assay Membrane fraction prepared from pig heart was diluted to 0.15 mg / ml with assay buffer, and 100 μl of this was dispensed into assay tubes for use in the assay. To this membrane fraction suspension, 2 μl of endothelin-1 solution labeled with 5 μM radioactive iodine, and 3 μl of test peptide solution were added.
In addition, the temperature was kept at 25 ° C for 1 hour. Then, the membrane fraction suspension was diluted with 900 μl of an ice-cooled assay buffer, and then centrifuged at 12,000 × g for 10 minutes to separate a supernatant and a precipitate. The sediment contains the cell membrane and the endothelin receptor embedded therein, and the radioiodinated endothelin bound to the receptor is also collected in the sediment. Therefore, the amount of radioiodine-labeled endothelin bound to the endothelin receptor was quantified by measuring the radioactive iodine in this sink with a gamma ray counter.

[Table 2] ────────────────────────── Examples Compounds Binding activity (* 1) No. Specific activity ──── ────────────────────── TAN-1462A 1.00 (* 2) 1 [D-Asp ¹ ] -TAN-1462A 26.6 2 [Asp ² ] -TAN- 1462A 2.70 3 [D-tLeu ³ ] -TAN-1462A 2.44 4 [D-γMeLeu ³ ] -TAN-1462A 1.54 5 [D-Phg ³ ] -TAN-1462A 1.27 6 [D-Asp ¹ , Asp ² ] -TAN -1477D 22.8 ────────────────────────────────── * 1 Porcine myocardial fraction * 2 IC ₅₀ = 1.10 × 10 ^-6 M Compounds of the invention, such as the compound Cyclo (-D-Asp-Ala
-D-aIle-Leu-D-Trp-) was 200 mg / kg by oral administration in an acute toxicity test (LD ₅₀ ) using mice.
As mentioned above, the intraperitoneal administration was 100-200 mg / kg.
As described above, the novel cyclic pentapeptide [I] of the present invention or a salt thereof has a property as an endothelin antagonist and can be used as a circulatory function improving agent, a vasodilator or a therapeutic agent for asthma.

[0017]

[Operation / Effect] The novel cyclic pentapeptide of the present invention is
As an endothelin antagonist, it exerts a remarkable effect in suppressing the vasoconstrictor activity of endothelin. Therefore, the novel cyclic pentapeptide of the present invention or a salt thereof can be used as a circulatory function improving agent, a therapeutic agent for myocardial infarction, acute renal failure or the like, or an asthma therapeutic agent. When the peptide of the present invention is used as the above-mentioned therapeutic agent, as it is or mixed with a pharmacologically acceptable carrier, excipient, diluent, powder, granule, tablet, capsule, injection, suppository, ointment It can be safely administered orally or parenterally in a dosage form such as a sustained-release preparation. The derivative of the present invention is mainly administered parenterally (eg, intravenous or subcutaneous injection, intraventricular or intraspinal injection, nasal administration, rectal administration),
In some cases, it may be administered orally.

Since the derivative of the present invention is stable as a substance, it can be stored as a solution of physiological saline, but it can also be dissolved at the time of use by adding mannitol and sorbitol to form a freeze-dried ampoule. The peptide derivative of the present invention can be administered as a free form or as an alkali addition salt or acid addition salt thereof. The dosage of the derivative [I] is generally 1 μg to 100 mg per kg of body weight of the free form, the alkali addition salt and the acid addition salt of the derivative [I].
It is a proper amount in the range of. More specifically, the dose varies depending on the target disease, symptom, administration subject, administration method, etc., but when administered by injection to an adult hypertensive patient, for example, the medicinal component [Compound (1)] 1 1μ as a dose
It is convenient to administer about g / kg to 100 mg / kg body weight once to three times a day. In addition, it is effective even with infusion,
The total dose for infusion is the same as for injection. When this peptide is used as a therapeutic agent, it must be carefully purified and care should be taken to avoid the presence of bacteria and pyrogens.

[0019]

EXAMPLES The present invention will be described in more detail with reference to the following examples. In addition, all amino acids other than glycine in the examples are in the L-configuration except where specifically noted. The following [Table 3] shows the fermentation products TAN-1462A, B,
1 shows the chemical structures of TAN-1477C-K. The nomenclature order of the constituent amino acids is D-Glu at position 1.

[Table 3] ─────────────────────────── 1 2 3 4 5 ─────────────── ───────────── TAN-1462A Cyclo (-D-Glu-Ala-D-aIle-Leu-D-Trp-) TAN-1462B Cyclo (-D-Glu-Ala-D- Val- Leu-D-Trp-) TAN-1477C Cyclo (-D-Glu-Ala-D-aIle-Val-D-Trp-) TAN-1477D Cyclo (-D-Glu-Ala-D-Leu- Leu- D-Trp-) TAN-1477E Cyclo (-D-Glu-Ala-D-Leu- Val-D-Trp-) TAN-1477F Cyclo (-D-Glu-Ala-D-Val- Val-D-Trp- ) TAN-1477G Cyclo (-D-Glu-Ala-D-Nva- Val-D-Trp-) TAN-1477H Cyclo (-D-Glu-Ala-D-Nva- Nva-D-Trp-) TAN-1477J Cyclo (-D-Glu-Ala-D-aIle-Nva-D-Trp-) TAN-1477K Cyclo (-D-Glu-Ala-D-aIle-Leu-D-mTrp-) ─────── ───────────────────── The thin-layer chromatography plate used in the examples is a product of Merck (SILICAGEL60F-254), and the developing solvent is Rf1: Chloroform-methanol (19: 1), Rf
2: Chloroform-methanol-acetic acid (9: 1: 0.5)
Was used.

Example 1 Production of [D-Asp ¹ ] -TAN-1462A (I) Production of Boc-D-aIle-Leu-OBzl Boc-D-aIle-OH.CHA 16.5 g of AcOEt, water and 1N. H ₂ S
60 ml of O ₄ was added and shaken, and the AcOEt layer was washed with water and then Na ₂ SO ₄ was added.
After drying and concentration, a residue was obtained. This and H-Leu-OBzl ・ pT
Dissolve 21.6 g of os in 100 ml of DMF and cool with ice.
EA 11.6ml, HOBt 7.4g, DCC 11.4g
And stir overnight. The generated DCU is filtered off,
The filtrate was concentrated, the residue was dissolved in AcOEt and the residue was dissolved in 4% NaH.
Washed with CO ₃ water, 10% citric acid water, washed with water, and then Na ₂ S
It was dried over O ₄ , concentrated, and ether was added to the residue to obtain a precipitate, which was collected by filtration. Yield 21.1 g (Yield 97.1%) mp 109-110 ° C. Rf2 0.82 [α] _D ²⁵ -10.4 ° (c = 0.93 in DMF) Elemental analysis C ₂₄ H ₃₈ N ₂ O Calculated as ₅ : C, 66.33; H, 8.81; N, 6.45 Experimental value: C, 66.54; H, 9.03; N, 6.49

(II) Production of Boc-D-aIle-Leu-OPac 21.1 g of Boc-D-aIle-Leu-OBzl was added to 500 m of methanol.
It was dissolved in 1 and catalytically reduced in a hydrogen stream using 10% Pd-carbon as a catalyst. The catalyst was filtered off and then concentrated to remove the residue and Cs ₂ C.
7.9 g of O ₃ was dissolved in 90% MeOH in water, concentrated, and the residue was dissolved in 200 ml of DMF. Phenacyl Bromide 1
0.6 g was added and stirred overnight. The produced TEA hydrochloride was filtered off, the filtrate was concentrated, the residue was dissolved in AcOEt, this was washed with 4% NaHCO ₃ water, 10% citric acid water, washed with water, dried over Na ₂ SO ₄ and concentrated. Then, ether was added to the residue and the precipitate was collected by filtration. Yield 22.2 g (Yield 98.7%) mp 116-117 ° C. Rf2 0.79 [α] _D ²⁵ -17.6 ° (c = 0.93 in DMF) Elemental analysis C ₂₅ H ₃₈ N ₂ O Calculated as ₆ : C, 64.91; H, 8.28; N, 6.06 Experimental value: C, 65.17; H, 8.68; N, 6.13

(III) Preparation of Boc-Ala-D-aIle-Leu-OPac 80 ml of TFA was added to 7.40 g of Boc-D-aIle-Leu-OPac, dissolved and concentrated, and the residue was dissolved in AcOEt. 4% Na
It was washed with HCO ₃ water, dried over Na ₂ SO ₄ and concentrated. Dissolve this in 80 ml of DMF, cool with ice, add 2.2 ml of TEA, and then add Boc-Ala-ONB (3.33 g of Boc-Ala-OH, HONB
3.48 g and DCC 4.00 g) were added and the mixture was stirred overnight. (CH ₃ ) ₂ N (CH ₂ ) ₃ NH ₂ 0.66 ml was added to this, and 3
Shake for 0 minutes and then concentrate, dissolve the residue in AcOEt,
This was washed with 4% NaHCO ₃ water, 10% citric acid water,
After washing with water, it was dried over Na ₂ SO ₄ and concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 7.80 g (Yield 91.4%) mp 154-155 ° C. Rf2 0.69 [α] _D ²⁵ -33.1 ° (c = 1.01 in DMF) Elemental analysis C ₂₈ H ₄₃ N ₃ O Calculated as ₇ : C, 63.01; H, 8.12; N, 7.87 Experimental value: C, 63.33; H, 8.24; N, 7.69

(IV) Boc-D-Asp (OBzl) -Ala-D-aIle-Leu-OP
Production of ac To 1.71 g of Boc-Ala-D-aIle-Leu-OPac, 20 ml of TFA was added, dissolved and concentrated, and 8N-HCl / dioxane 1.
0 ml was added, ether was added, and the precipitated crystals were collected by filtration,
Dried. Dissolve this in 20 ml of DMF and chill with ice.
0.67 ml of EA was added. To this, Boc-D-Asp (OBzl) -ONB
(Boc-D-Asp (OBzl) -OH 1.14 g, HONB 0.69
g, prepared from 0.80 g of DCC), stirred overnight and concentrated. Water was added to the residue and the precipitate was collected by filtration. This was dissolved in DMF, the insoluble matter was filtered off, and the mixture was concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 2.00 g (yield 84.6%) mp 154-155 ° C. Rf2 0.68 [α] _D ²⁵ -2.1 ° (c = 0.94 in DMF) Elemental analysis C ₃₉ H ₅₄ N ₄ O Calculated as ₁₀ : C, 63.39; H, 7.37; N, 7.58 Experimental value: C, 63.51; H, 7.45; N, 7.52

(V) Boc-D-Trp-D-Asp (OBzl) -Ala-D-aIle
-Production of Leu-OPac Boc-D-Asp (OBzl) -Ala-D-aIle-Leu-OPac 1.70 g of T
After adding 20 ml of FA to dissolve and concentrate, 8N-HCl /
Dioxane (0.7 ml) was added, ether was added, and the precipitated crystals were collected by filtration and dried. This was dissolved in DMF (15 ml), ice-cooled, and TEA (0.48 ml) was added. To this, Boc-
D-Trp-ONB (Boc-D-Trp-OH 0.84g, HONB 0.54)
g, prepared from 0.63 g of DCC), stirred overnight and concentrated. Water was added to the residue and the precipitate was collected by filtration. This was dissolved in DMF, the insoluble matter was filtered off, and the mixture was concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 1.72 g (yield 80.8%) mp 189-190 ° C. Rf2 0.66 [α] _D ²⁵ 3.9 ° (c = 0.90 in DMF) Elemental analysis C ₅₀ H ₆₄ N ₆ O ₁₁ Calculated value: C, 64.91; H, 6.97; N, 9.08 Experimental value: C, 64.84; H, 6.78; N, 9.13

(VI) Boc-D-Trp-D-Asp (OBzl) -Ala-D-aIle-
Production of Leu-OH Boc-D-Trp-D-Asp (OBzl) -Ala-D-aIle-Leu-OPac 1.48
g was dissolved in 40 ml of AcOH, 5.23 g of Zn powder was added, and the mixture was stirred for 3 hours. The Zn powder was filtered off and the filtrate was concentrated. AcOEt was added to the residue to dissolve it, washed with 10% citric acid, washed with water, dried over Na ₂ SO ₄ , concentrated, and ether was added to the residue to obtain a precipitate by filtration. Yield 1.14 g (yield 88.3%) mp 128-130 ° C. Rf2 0.64 [α] _D ²⁵ 11.6 ° (c = 1.02 in DMF) Elemental analysis C ₄₂ H ₆₆ N ₆ O ₁₀ Calculated value: C, 62.51; H, 7.24; N, 10.41 Experimental value: C, 62.68; H, 7.33; N, 10.27

(VII) Cyclo (-D-Asp-
Ala-D-aIle-Leu-D-Trp-)
Preparation of ([D-Asp ¹ ] -TAN-1462A) Boc-D-Trp-D-Asp (OBzl) -Ala-D-aIle-Leu-OH 0.81 g
Was dissolved in 20 ml of DCM and cooled with ice.
g and DCC 0.41 g were added and the mixture was stirred for 3 hours, then the DCU formed was separated by filtration, concentrated, and ether was added to the residue to collect a precipitate. Under ice-cooling, ethanedithiol (0.17 ml) and 8N-HCl / dioxane (20 ml) were added and dissolved. The mixture was stirred for 10 minutes and concentrated. Ether was added to the residue, and the precipitate was collected by filtration and dried. This is D
DMF 2 containing 1.4 ml TEA dissolved in 20 ml MF
After adding dropwise to 00 ml over 30 minutes, the mixture was stirred overnight and concentrated. Acetonitrile was added to the residue and the precipitate was collected by filtration and dried. 0.50 g of this is DMF 20
It was dissolved in ml and catalytically reduced in a hydrogen stream using Pd-carbon as a catalyst. The catalyst was filtered off and concentrated, the residue was dissolved in a small amount of AcOH, water was added, and the mixture was freeze-dried. Furthermore, this is a YMC-made YMC-D-OD.
Purification by preparative liquid chromatography using an S-5 (2 cm x 25 cm) column gave the desired product. Yield 19.4 mg (yield 4.0%) Amino acid analysis value (hydrolysis at 110 ° C. for 24 hours; values in parentheses indicate theoretical value): Asp 1.00 (1); Ala 0.99 (1); Leu
0.83 (1) LSIMS (M + H ⁺ ) = 599 (theoretical value) = 599

Example 2 Production of [Asp ² ] -TAN-1462A (I) Production of Boc-Asp (OBzl) -D-aIle-Leu-OPac Boc-D-aIle-prepared in Example 1 (II) Leu-OPac 1.85
15 g of TFA was added to g to dissolve, then the mixture was concentrated, the residue was dissolved in AcOEt, and the mixture was washed with 4% NaHCO ₃ water, and Na ₂ S was added.
It was dried over O ₄ and concentrated. This was dissolved in DMF (15 ml) and cooled with ice, and Boc-Asp (OBzl) -ONB (Boc-Asp (OBzl) -OH 1.42
g, HONB 0.87 g, DCC 1.00 g) were added and the mixture was stirred overnight. (CH ₃ ) ₂ N (CH ₂ ) ₃ NH ₂ 0.17
Add ml and shake for 30 minutes, then concentrate and concentrate the residue with AcOEt.
It was dissolved in water, washed with 4% NaHCO ₃ water, 10% citric acid water, washed with water, dried over Na ₂ SO ₄ and concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 2.42 g (yield 90.6%) mp 116-117 ° C. Rf2 0.75 [α] _D ²⁵ -40.1 ° (c = 1.02 in DMF) Elemental analysis C ₃₆ H ₄₉ N ₃ O Calculated as ₉ : C, 64.75; H, 7.40; N, 6.29 Experimental value: C, 64.86; H, 7.63; N, 6.55

(II) Boc-D-Glu (OBzl) -Asp (OBzl) -D-aIle-
Production of Leu-OPac 2.27 g of Boc-Asp (OBzl) -D-aIle-Leu-OPac with TFA 2
0 ml was added to dissolve and then concentrated. 8N-HCl / dioxane 1.1 ml was added, ether was added, and the precipitated crystals were collected by filtration and dried. This was dissolved in DMF (15 ml), ice-cooled, and TEA (0.62 ml) was added. To this, Boc-D-Glu
(OBzl) -ONB (Boc-D-Glu (OBzl) -OH 1.26g, HONB
0.74 g and DCC 0.85 g) was added and the mixture was stirred overnight and concentrated. Water was added to the residue and the precipitate was collected by filtration. This was dissolved in DMF, the insoluble matter was filtered off, and the mixture was concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 2.79 g (yield 92.5%) mp 155-156 ° C. Rf2 0.71 [α] _D ²⁵ −24.4 ° (c = 1.01 in DMF) Elemental analysis C ₄₈ H ₆₂ N ₄ O Calculated as ₁₂ : C, 64.99; H, 7.05; N, 6.32 Experimental value: C, 65.02; H, 7.14; N, 6.42

(III) Boc-D-Trp-D-Glu (OBzl) -Asp (OBzl)-
Production of D-aIle-Leu-OPac Boc-D-Glu (OBzl) -Asp (OBzl) -D-aIle-Leu-OPac 2.40
20 ml of TFA was added to and dissolved in g, and the mixture was concentrated. Ether was added, and the precipitated crystals were collected by filtration and dried. This is D
It was dissolved in 15 ml of MF, cooled with ice, and 0.57 ml of TFA was added. In addition, Boc-D-Trp-ONB (Boc-D-Trp-OH 0.90
g, HONB 0.59 g, DCC 0.67 g)
Was added and the mixture was stirred overnight and then concentrated. Water was added to the residue, and the precipitate was collected by filtration. This was dissolved in DMF, the insoluble matter was filtered off, and the mixture was concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 2.35 g (yield 81.1%) mp 164-165 ° C. Rf2 0.67 [α] _D ²⁵ -18.8 ° (c = 0.99 in DMF) Elemental analysis C ₅₉ H ₇₂ N ₆ O Calculated as ₁₃ : C, 66.03; H, 6.76; N, 7.83 Experimental value: C, 65.90; H, 6.72; N, 7.77

(IV) Boc-D-Trp-D-Glu (OBzl) -Asp (OBzl) -D
-aIle-Leu-OH Production Boc-D-Trp-D-Glu (OBzl) -Asp (OBzl) -D-aIle-Leu-OPac
2.15 g was dissolved in 50 ml AcOH, and Zn powder 6.54
g and stirred for 3 hours. The Zn powder was filtered off and the filtrate was concentrated. AcOEt was added to the residue to dissolve it, and the residue was washed with 10% citric acid, washed with water, dried over Na ₂ SO ₄ , concentrated, and ether was added to the residue to collect a precipitate. Yield 1.74 g (Yield 91.1%) mp 175-176 ° C. Rf2 0.62 [α] _D ²⁵ -19.1 ° (c = 1.13 in DMF) Elemental analysis C ₅₁ H ₆₆ N ₆ O Calculated as ₁₂ : C, 64.13; H, 6.97; N, 8.80 Experimental value: C, 64.36; H, 7.09; N, 8.89

(V) Cyclo (-D-Glu-Asp-D-aIle-Leu-D-Trp
-) Production of ([Asp ² ] -TAN-1462A) Boc-D-Trp-D-Glu (OBzl) -Asp (OBzl) -D-aIle-Leu-OH 0.
Dissolve 95 g in 20 ml DCM and cool with ice, then use HONB
0.36 g and 0.41 g of DCC were added and the mixture was stirred for 3 hours. The DCU formed was filtered off and concentrated, and ether was added to the residue to collect a precipitate. Under ice cooling, 0.17 ml of ethanedithiol, 8N-HCl / dioxane 20 was added.
After adding ml to dissolve it, the mixture was stirred for 10 minutes and concentrated, and ether was added to the residue, and the precipitate was collected by filtration and dried.
Dissolve this in 20 ml DMF and add 1.4 ml TEA D
After adding dropwise to 200 ml of MF over 30 minutes, the mixture was stirred overnight and concentrated. Acetonitrile was added to the residue and the precipitate was collected by filtration and dried. 0.50 g of this is D
It was dissolved in 20 ml of MF and subjected to catalytic reduction under a hydrogen stream using Pd-carbon as a catalyst. The catalyst was filtered off and concentrated, the residue was dissolved in a small amount of AcOH, water was added, and the mixture was freeze-dried. Yield 328 mg (yield 63.7%) Amino acid analysis value (hydrolysis at 110 ° C. for 24 hours; values in parentheses indicate theoretical value): Asp 1.00 (1); Glu 1.06 (1); Leu
0.86 (1) LSIMS (M + H ⁺ ) = 657 (theoretical value) = 657

Example 3 Manufacture of [D-tLeu ³ ] -TAN-1462A (I) Manufacture of Boc-Ala-OPac 90% of 28.4 g of Boc-Ala-OH and 24.5 g of Cs ₂ CO ₃
Dissolve in MeOH water, concentrate and concentrate the residue with DMF 450 ml.
The mixture was dissolved in water, Phenacyl Bromide (32.9 g) was added, and the mixture was stirred overnight. The TEA hydrochloride formed was filtered off, the filtrate was concentrated, the residue was dissolved in AcOEt and this was dissolved in 4% NaHCO ₃ water,
It was washed with 10% citric acid water, washed with water, dried over Na ₂ SO ₄ and concentrated. Ether was added to the residue and the precipitate was collected by filtration. Yield 42.4 g (Yield 91.9%) mp 123 ° C. Rf1 0.72 Rf2 0.74 [α] _D ²⁸ -46.1 ° (c = 1.36 in DMF) Elemental analysis C ₁₆ H ₂₁ NO Calculated as ₅ : C, 62.53; H, 6.89; N, 4.56 Experimental value: C, 62.94; H, 6.87; N, 4.63

(II) Preparation of Boc-D-Glu (OBzl) -Ala-OPac Boc-D-Glu (OBzl) -OH (2.02 g) was dissolved in THF and cooled to -15 ° C with stirring. 0.66 ml of N-methylmorpholine was added, followed by 0.80 ml of IBCF. After 2 minutes, HCI-H-Ala-OPac and N-methylmorpholine (0.66 ml of DMF solution) were added (HCI-H-Ala-OP).
ac was obtained by adding TFA (20 ml) to Boc-Ala-OPac (1.84 g), dissolving and concentrating the mixture, adding 8N-HCl / dioxane (1.95 ml), adding ether and filtering the precipitated crystals, and drying the crystals. ). After stirring at -15 ° C for 30 minutes,
Return to room temperature. After 30 minutes, the insoluble matter was filtered off and then concentrated, the residue was dissolved in AcOEt, and this was dissolved in water, 10% aqueous citric acid solution, and 4%.
% NaHCO ₃ water, washed with water, dried with Na ₂ SO ₄ ,
Concentrate and recrystallize from AcOEt-petroleum ether. Yield 2.63 g (Yield 83.2%) mp 128 ° C. Rf1 0.54 Rf2 0.65 [α] _D ²⁸ -13.0 ° (c = 1.19 in DMF) Elemental analysis C ₂₈ H ₃₄ N Calculated as ₂ O ₈ : C, 63.87; H, 6.51; N, 5.32 Experimental value: C, 63.88; H, 6.32; N, 5.18

(III) Boc-D-Trp-D-Glu (OBzl) -Ala-OPac
Preparation of Boc-D-Glu (OBzl) -Ala-OPac 1.05g TFA 20ml
Was added to dissolve and then concentrated, 0.58 ml of 8N-HCl / dioxane was added, ether was added, and the precipitated crystals were collected by filtration and dried. This was dissolved in DMF (15 ml), ice-cooled, and TEA (0.20 ml) was added. To this, Boc-D-Trp-ON
B (Boc-D-Trp-OH 0.67g, HONB 0.43g, DC
C 0.50 g) was added and the mixture was stirred overnight. The resulting insoluble matter was filtered off and concentrated, and the residue was dissolved in AcOEt, washed with water, 10% citric acid water, 4% NaHCO ₃ water, washed with water, dried over Na ₂ SO ₄ and concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 1.39 g (Yield 97.8%) mp 127-129 ° C. Rf1 0.36 Rf2
0.62 [α] _D ²⁸ -4.04 ° (c = 1.04 in DMF) Elemental analysis Calculated as C ₃₉ H ₄₄ N ₄ O ₉ : C, 65.72; H, 6.22; N, 7.86 Experimental value : C, 65.62; H, 6.46; N, 7.78

(IV) Boc-Leu-D-Trp-D-Glu (OBzl) -Ala-OPa
Preparation of c 21.4 g of Boc-D-Trp-D-Glu (OBzl) -Ala-OPac was suspended in dioxane, 5.1 ml of ethanedithiol was added and ice-cooled, 8N-HCl / dioxane was added, and ice-cooled The mixture is stirred for 1 hour below, then concentrated, ether is added, and the precipitated crystals are collected by filtration and dried. This is dissolved in 150 ml of DMF, cooled with ice, and 8.42 ml of TEA is added. To this, Boc-Leu-ONB
(Boc-Leu-OH.H ₂ O 8.22 g, HONB 6.51 g,
DCC (prepared from 7.50 g) was added and the mixture was stirred overnight. The resulting insoluble matter was filtered off and concentrated, the residue was dissolved in AcOEt, and this was washed with water, 10% citric acid water, 4% NaHCO ₃ water, washed with water, dried over Na ₂ SO ₄ and concentrated, Ether was added to the residue and the precipitate was collected by filtration. Yield 23.0 g (Yield 92.8%) mp 127-128 ° C. Rf1 0.35 Rf2
0.63 [α] _D ²⁸ −9.91 ° (c = 1.15, in DMF) Elemental analysis Calculated as C ₄₅ H ₅₅ N ₅ O ₁₀ : C, 65.44; H, 6.71; N, 8.48 Experimental value : C, 65.39; H, 6.92; N, 8.30

(V) Boc-D-tLeu-Leu-D-Trp-D-Glu (OBzl)-
Preparation of Ala-OPac 2.07 g of Boc-Leu-D-Trp-D-Glu (OBzl) -Ala-OPac was suspended in dioxane, 0.42 ml of ethanedithiol was added and ice-cooled to obtain 8N-HCl / dioxane. In addition, under ice cooling 30
After stirring for minutes, the mixture was concentrated, ether was added, and the precipitated crystals were collected by filtration and dried. This was dissolved in DMF (15 ml), ice-cooled, and TEA (0.70 ml) was added. To this, Boc-D-tLeu
-OH 0.64g, HONB 0.41g, DCC 0.63
g and stirred overnight. The resulting insoluble matter was filtered off and concentrated, the residue was dissolved in AcOEt, and this was washed with water, 10% citric acid water, 4% NaHCO ₃ water, washed with water, and then Na ₂ S.
It was dried with O ₄ , concentrated, and ether was added to the residue to obtain a precipitate, which was collected by filtration. Yield 2.13 g (yield 92.1%) mp 162-164 ° C. Rf1 0.27 Rf2
0.64 [α] _D ²⁸ -4.77 ° (c = 0.78 in DMF) Elemental analysis Calculated as C ₅₁ H ₆₆ N ₆ O ₁₁ : C, 65.23; H, 7.08; N, 8.95 Experimental value : C, 65.15; H, 7.31; N, 9.18

(VI) Boc-D-tLeu-Leu-D-Trp-D-Glu (OBzl)-
Production of Ala-OH Boc-D-tLeu-Leu-D-Trp-D-Glu (OBzl) -Ala-OPac 1.41
g was dissolved in 20 ml of 90% AcOH water, and Zn powder 4.91 was added.
Add g and stir for 3 hours. The Zn powder is filtered off and the filtrate is concentrated. AcOEt was added to the residue to dissolve it, and the residue was washed with 10% citric acid, washed with water, dried over Na ₂ SO ₄ , concentrated, and ether was added to the residue to collect a precipitate. Yield 1.08 g (Yield 87.7%) mp 140-142 ° C. Rf1 0.04 Rf2
0.61 [α] _D ²⁸ + 9.09 ° (c = 0.99 in DMF) Elemental analysis Calculated as C ₄₃ H ₆₀ N ₆ O ₁₀ : C, 62.91; H, 7.37; N, 10.24 Experimental value: C, 62.70; H, 7.64; N, 10.24

(VII) Cyclo (-D-Glu-Ala-D-tLeu-Leu-D-Al
a-) ([D-tLeu ³ ] -TAN-1462A) Production Boc-D-tLeu-Leu-D-Trp-D-Glu (OBzl) -Ala-OH 0.41 g
Was dissolved in 20 ml of DCM and cooled with ice.
g and DCC (0.21 g) were added and the mixture was stirred for 3 hours. The DCU formed was filtered off and concentrated. Ether was added to the residue and the precipitate was collected by filtration. Under ice-cooling, 0.09 ml of ethanedithiol and 20 ml of 8N-HCl / dioxane were added and dissolved, and the mixture was stirred for 10 minutes and concentrated. Ether was added to the residue, and the precipitate was collected by filtration and dried. DM this
DMF 90 containing 0.7 ml TEA dissolved in 10 ml F
The mixture was added dropwise to ml over 30 minutes, then stirred overnight and concentrated. Acetonitrile was added to the residue and the precipitate was collected by filtration and dried. 50 mg of this was dissolved in 15 ml of DMF, and catalytic reduction was carried out in a hydrogen stream using Pd-carbon as a catalyst. The catalyst was filtered off and concentrated, and the residue was mixed with a small amount of A
After dissolving in cOH, water was added and freeze-dried. Finally, this is YMC-D-ODS-5 (2 cm
The product was purified by preparative liquid chromatography using a column having a size of 25 cm. Yield 9.3 mg (yield 6.8%) Amino acid analysis value (hydrolysis at 110 ° C. for 24 hours; values in parentheses indicate theoretical value): Glu 1.00 (1); Ala 0.98 (1); Leu
0.79 (1) LSIMS (M + H ⁺ ) = 613 (theoretical value) = 613

Example 4 Production of [D-γMeLeu ³ ] -TAN-1462A (I) Boc-D-γMeLeu-Leu-D-Trp-D-Glu (OBzl) -Ala-OPac
Preparation of Boc-Leu-D-Trp-D-Glu (OBzl) -Al prepared in Example 3 (IV)
2.07 g of a-OPac was suspended in dioxane, 0.42 ml of ethanedithiol was added, and the mixture was ice-cooled, 8N-HCl / dioxane was added, and the mixture was stirred for 30 minutes under ice-cooling and then concentrated, and ether was added to precipitate crystals. Was collected by filtration and dried. DMF this
It was dissolved in 15 ml and cooled with ice, and 0.70 ml of TEA was added. Boc-D-γMeLeu-OH 0.68g, HONB
0.41 g and DCC 0.63 g were added and stirred overnight.
The resulting insoluble matter was filtered off and concentrated, the residue was dissolved in AcOEt, and this was washed with water, 10% citric acid water, 4% NaHCO ₃ water, washed with water, dried over Na ₂ SO ₄ and concentrated, Ether was added to the residue and the precipitate was collected by filtration. Yield 2.09 g (yield 87.7%) mp 121-124 ° C. Rf1 0.28 Rf2
0.64 [α] _D ²⁸ + 7.66 ° (c = 1.01 in DMF) Elemental analysis Calculated as C ₅₂ H ₆₈ N ₆ O ₁₁ : C, 65.53; H, 7.19; N, 8.82 Experimental value: C, 65.53; H, 7.33; N, 9.18

(II) Boc-D-γMeLeu-Leu-D-Trp-D-Glu (OBz
l) -Production of Ala-OH Boc-D-γMeLeu-Leu-D-Trp-D-Glu (OBzl) -Ala-OPac 1.4
3 g was dissolved in 20 ml of 90% AcOH water, and Zn powder 4.9
1 g was added and stirred for 3 hours. The Zn powder was filtered off and the filtrate was concentrated. Add AcOEt to the residue to dissolve, 10%
The extract was washed with citric acid, washed with water, dried over Na ₂ SO ₄ , concentrated, and ether was added to the residue to collect a precipitate by filtration. Yield 1.13 g (Yield 90.2%) mp 135-136 ° C. Rf1 0.07 Rf2
0.61 [α] _D ²⁸ + 22.1 ° (c = 1.07 in DMF) Elemental analysis Calculated as C ₄₄ H ₆₂ N ₆ O ₁₀ : C, 63.29; H, 7.48; N, 10.06 Experimental value: C, 63.29; H, 7.56; N, 9.96

(III) Cyclo (-D-Glu-Ala-D-γMeLeu-Leu-D
-Ala-) ([D-γMeLeu ³ ] -TAN-1462A) Production Boc-D-γMeLeu-Leu-D-Trp-D-Glu (OBzl) -Ala-OH 0.42
g in DCM 20 ml and chilled on ice, then HONB 0.1
After adding 8g and DCC 0.21g and stirring for 3 hours,
The DCU that formed was filtered off, concentrated, and ether was added to the residue to obtain a precipitate, which was collected by filtration. Under ice-cooling, 0.09 ml of ethanedithiol and 20 ml of 8N-HCl / dioxane were added and dissolved, and the mixture was stirred for 10 minutes and concentrated. Ether was added to the residue, and the precipitate was collected by filtration and dried. This is D
DMF 9 dissolved in 10 ml MF and containing 0.7 ml TEA
The mixture was added dropwise to 0 ml over 30 minutes, stirred overnight and concentrated. Acetonitrile was added to the residue and the precipitate was collected by filtration and dried. 51 mg of this was dissolved in 15 ml of DMF, and catalytic reduction was carried out in a hydrogen stream using Pd-carbon as a catalyst. After the catalyst was filtered off and concentrated, the residue was dissolved in a small amount of AcOH, water was added, and the mixture was freeze-dried. Finally, this was made by YMC YMC-D-ODS-5 (2
(cm × 25 cm) column was used for purification by preparative liquid chromatography to obtain the desired product. Yield 2.4 mg (yield 2.9%) Amino acid analysis value (hydrolysis at 110 ° C. for 24 hours; the value in parentheses indicates theoretical value): Glu 1.00 (1); Ala 1.00 (1); Leu
1.01 (1) LSIMS (M + H ⁺ ) = 627 (theoretical value) = 627

Example 5 Production of [D-Phg ³ ] -TAN-1462A (I) Production of Boc-D-Phg-Leu-D-Trp-D-Glu (OBzl) -Ala-OPac Example 3 (IV ) Boc-Leu-D-Trp-D-Glu (OBzl) -Al
2.07 g of a-OPac was suspended in dioxane, 0.42 ml of ethanedithiol was added, and the mixture was ice-cooled, 8.3N-HCl / dioxane was added, and the mixture was stirred under ice-cooling for 30 minutes and then concentrated, and ether was added to precipitate. The formed crystals were collected by filtration and dried. This is D
It was dissolved in 15 ml of MF, ice-cooled, and 0.70 ml of TEA was added. Boc-D-Phg-OH 0.69g, HONB 0.4
1 g and DCC 0.63 g were added and stirred overnight. The resulting insoluble matter was filtered off and concentrated, the residue was dissolved in AcOEt, and this was washed with water, 10% citric acid water, 4% NaHCO ₃ water, washed with water, dried over Na ₂ SO ₄ and concentrated, Ether was added to the residue and the precipitate was collected by filtration. Yield 2.19 g (Yield 91.3%) mp 158-159 ° C. Rf1 0.28 Rf2
0.64 [α] _D ²⁸ -15.2 ° (c = 0.95 in DMF) Elemental analysis Calculated as C ₅₃ H ₆₂ N ₆ O ₁₁ : C, 66.37; H, 6.52; N, 8.76 Experimental value : C, 66.24; H, 6.64; N, 8.95

(II) Boc-D-Phg-Leu-D-Trp-D-Glu (OBzl) -A
Production of la-OH Boc-D-Phg-Leu-D-Trp-D-Glu (OBzl) -Ala-OPac 1.44g
Was dissolved in 20 ml of 90% AcOH water, and Zn powder was 4.91 g.
Was added and stirred for 3 hours. The Zn powder was filtered off and the filtrate was concentrated. AcOEt was added to the residue to dissolve it, and the residue was washed with 10% citric acid, washed with water, dried over Na ₂ SO ₄ , concentrated, and ether was added to the residue to collect a precipitate. Yield 1.24 g (Yield 98.3%) mp 138-140 ° C. Rf1 0.05 Rf2
0.61 [α] _D ²⁸ -3.12 ° (c = 1.19 in DMF) Elemental analysis Calculated as C ₄₅ H ₅₆ N ₆ O ₁₀ : C, 64.27; H, 6.71; N, 9.99 Experimental value : C, 64.18; H, 6.99; N, 9.66

(III) Cyclo (-D-Glu-
Ala-D-Phg-Leu-D-Ala-) ([D
-Phg ^3] cooled -TAN-1462A) ice production Boc-D-Phg-Leu- D-Trp-D-Glu (OBzl) -Ala-OH 0.42g dissolved in DCM 20 ml of, HONB 0.18
g and DCC (0.21 g) were added and the mixture was stirred for 3 hours. The DCU formed was filtered off and concentrated. Ether was added to the residue and the precipitate was collected by filtration. Under ice-cooling, 0.09 ml of ethanedithiol and 20 ml of 8N-HCl / dioxane were added and dissolved, and the mixture was stirred for 10 minutes and concentrated. Ether was added to the residue, and the precipitate was collected by filtration and dried. DM this
DMF 90 containing 0.7 ml TEA dissolved in 10 ml F
The mixture was added dropwise to ml over 30 minutes, then stirred overnight and concentrated. Acetonitrile was added to the residue and the precipitate was collected by filtration and dried. 72 mg of this was dissolved in 15 ml of DMF, and catalytic reduction was carried out in a hydrogen stream using Pd-carbon as a catalyst. The catalyst was filtered off and concentrated, and the residue was mixed with a small amount of A
After dissolving in cOH, water was added and freeze-dried. Finally, this is YMC-D-ODS-5 (2 cm
The product was purified by preparative liquid chromatography using a column having a size of 25 cm. Yield: 7.7 mg (yield: 14.2%) Amino acid analysis value (hydrolysis at 110 ° C. for 24 hours; the value in parentheses indicates theoretical value): Glu 1.00 (1); Ala 1.04 (1); Leu
0.98 (1) LSIMS (M + H ⁺ ) = 633 (theoretical value) = 633

Example 6 Production of [D-Asp ¹ , Asp ² ] -TAN-1477D (I) Production of Boc-D-Leu-Leu-OBzl 21.6 g of H-Leu-OBzl.pTos was dissolved in 100 ml of DMF. Cool on ice, TEA 7.7 ml, Boc-D-Leu-ONB (Boc-D
-Leu-OH.H ₂ O 12.5 g, HONB 9.86 g, DCC1 1.4 g) was added and the mixture was stirred overnight. The DCU formed was filtered off, the filtrate was concentrated, and the residue was dissolved in AcOEt.
Wash with aHCO ₃ water, 10% citric acid water, wash with Na ₂ SO ₄
After drying and concentration with ether, ether was added to the residue and the precipitate was collected by filtration. Yield 19.8 g (Yield 91.3%) m.p. 94-95 ° C. Rf20.76 [α] _D ²⁵ + 3.6 ° (c = 1.06 in DMF) Elemental analysis C ₂₄ H ₃₆ N ₂ O ₅ calculated: C, 66.33; H, 8.81 ; N, 6.45 Found: C, 66.38; H, 8.87 ; N, 6.53

(II) Production of Boc-D-Leu-Leu-OPac 6.0 g of Boc-D-Leu-Leu-OBzl was dissolved in 20 ml of methanol, and catalytic reduction was carried out in a hydrogen stream using 10% Pd-carbon as a catalyst. I did. The catalyst was filtered off and then concentrated to remove the residue and C
2.1 g of s ₂ CO ₃ was dissolved in 90% MeOH water and concentrated. Dissolve the residue in 60 ml of DMF and use Phenacyl Bromi
de 2.8g was added and stirred overnight. Generated TEA
The hydrochloride was filtered off, the filtrate was concentrated, the residue was dissolved in AcOEt, washed with 4% NaHCO ₃ water, 10% citric acid water, washed with water, dried over Na ₂ SO ₄ and concentrated to leave the residue. Ether was added to the product, and the precipitate was collected by filtration. Yield 5.48 g (yield 85.8%) m.p. p. ^{_{98-99 ℃ Rf20.66 [α] D 25}} -3.9 ° (c = 1.09, in DMF) Elemental analysis C ₂₅ H ₃₈ N ₂ O ₆ Calculated: C, 64.91; H, 8 .28; N, 6.08 experimental value: C, 65.21; H, 8.54; N, 6.24

(III) Preparation of Boc-Asp (OBzl) -D-Leu-Leu-OPac 1.85 g of Boc-D-Leu-Leu-OPac was dissolved by adding 15 ml of TFA and then concentrated, and the residue was dissolved in AcOEt. 4% N
It was washed with aqCO ₃ water, dried over Na ₂ SO ₄ and concentrated.
This is dissolved in 20 ml of DMF and cooled with ice, then TEA0.6
After adding ml, Boc-Asp (OBzl) -ONB (Boc-Asp (OBzl)
-OH 1.42g, HONB 0.87g, DCC 1.00g
Prepared) was added and stirred overnight. Add to this (CH ₃ ) ₂ N (C
H ₂ ) ₃ NH _{2 (} 0.17 ml) was added, and the mixture was shaken for 30 minutes and then concentrated. The residue was dissolved in AcOEt, and this was dissolved in 4% NaHCO 3.
Wash with ₃ water, 10% citric acid water, wash with water, and then wash with Na ₂ SO _4.
After drying and concentration with ether, ether was added to the residue and the precipitate was collected by filtration. Yield 2.40 g (yield 89.9%) m. p. ^{_{121-122 ℃ Rf20.63 [α] D 25}} -22.8 ° (c = 1.17, in DMF) Elemental analysis C ₃₆ H ₄₉ N ₃ O ₉ Calculated: C, 64.74; H, 7 .40; N, 6.29 experimental value: C, 64.81; H, 7.58; N, 6.15.

(IV) Boc-D-Asp (OBzl) -Asp (OBzl) -D-Leu-
Preparation of Leu-OPac Boc-Asp (OBzl) -D-Leu-Leu-OPac 2.20 g of TFA20
After dissolving by adding ml, the mixture was concentrated, 1.0 ml of 8N-HCl / dioxane was added, ether was added, and the precipitated crystals were collected by filtration and dried. This was dissolved in 20 ml of DMF and cooled with ice, and 0.92 ml of TEA was added. To this, Boc-D-
Asp (OBzl) -ONB (Boc-D-Asp (OBzl) -OH 1.17g, HON
B 0.71 g and DCC 0.82 g) was added, and the mixture was stirred overnight and concentrated. Water was added to the residue, and the precipitate was collected by filtration. This was dissolved in DMF, the insoluble material was filtered off, and the mixture was concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 2.50 g (yield 87.0%) m.p. p. 118-119 ° Rf20.63 [α] _D ²⁵ -13.4 ° (c = 1.12 in DMF) Elemental analysis Calculated as C ₄₇ H ₆₈ N ₄ O ₁₂ : C, 64.66; H, 6 .93; N, 6.42 experimental value: C, 64.84; H, 7.03; N, 6.55.

(V) Boc-D-Trp-D-Asp (OBzl) -Asp (OBzl) -D
-Leu-Leu-OPac Production Boc-D-Asp (OBzl) -Asp (OBzl) -D-Leu-Leu-OPac 1.05g
TFA (10 ml) was added to dissolve and then concentrated to give 8N-H.
Cl / dioxane (0.36 ml) was added, ether was added, and the precipitated crystals were collected by filtration and dried. DMF 10m
It melt | dissolved in 1 and ice-cooled, TEA0.34ml was added. Boc-D-Trp-ONB (Boc-D-Trp-OH 0.40g, HON
B (0.26 g, prepared from 0.30 g of DCC) was added, and the mixture was stirred overnight and concentrated. Water was added to the residue and the precipitate was collected by filtration. This was dissolved in DMF, the insoluble material was filtered off, and the mixture was concentrated. Ether was added to the residue, and the precipitate was collected by filtration. Yield 0.79 g (yield 62.3%) m. p. 163-164 ° C. Rf20.58 [α] _D ²⁵ -4.9 ° (c = 0.73, in DMF) Elemental analysis Calculated as C ₅₈ H ₇₀ N ₆ O ₁₃ : C, 65.76; H, 6 .66; N, 7.93 experimental value: C, 65.59; H, 6.72; N, 7.81.

(VI) Boc-D-Trp-D-Asp (OBzl) -Asp (OBzl)-
Production of D-Leu-Leu-OH Boc-D-Trp-Asp (OBzl) -Asp (OBzl) -D-Leu-Leu-OPac 0.7
5 g was dissolved in 20 ml of AcOH, 2.31 g of Zn powder was added, and the mixture was stirred for 3 hours. The Zn powder was filtered off and the filtrate was concentrated. AcOEt was added to the residue to dissolve it, and the residue was washed with 10% citric acid, washed with water, dried over Na ₂ SO ₄ and concentrated. Ether was added to the residue and the precipitate was collected by filtration. Yield 0.59 g (yield 87.8%) m.p. p. 95-96 ° C. Rf20.56 [α] _D ²⁵ + 0.1 ° (c = 0.85 in DMF) Elemental analysis Calculated as C ₅₀ H ₆₄ N ₆ O ₁₂ : C, 63.81; H, 6. 85; N, 8.93 experimental value: C, 64.03; H, 6.99; N, 8.89.

(VII) cyclo (-D-Asp-Asp-D-Leu-Leu-D-Tr
Production of p-) ([D-Asp ¹ ,, Asp ² ] -TAN-1477D) Boc-D-Trp-D-Asp (OBzl) -Asp (OBzl) -D-Leu-Leu-OH 0.
Dissolve 49 g in 10 ml DCM and cool with ice, then use HONB
O, 19 g of DCC (0.21 g) was added and the mixture was stirred for 3 hours, then the formed DCU was filtered off and concentrated, and ether was added to the residue, and the precipitate was collected by filtration. Under ice cooling, 0.10 ml of ethanedithiol and 20 ml of 8N-HCl / dioxane were added and dissolved, followed by stirring for 10 minutes and concentration. Ether was added to the residue, and the precipitate was collected by filtration and dried. Dissolve this in DMF 10ml, TEA0.7
After adding dropwise to 100 ml of DMF containing 2 ml over 30 minutes, the mixture was stirred overnight and concentrated. Acetonitrile was added to the residue, and the precipitate was collected by filtration and dried. Of this, 0.20 g was dissolved in 20 ml of DMF, and catalytic reduction was carried out in a hydrogen stream using Pd-carbon as a catalyst. The catalyst was filtered off and concentrated, the residue was dissolved in a small amount of AcOH, water was added, and the mixture was freeze-dried. Yield 149 mg (yield 61.6%) Amino acid analysis value (hydrolysis at 110 ° C. for 24 hours; values in parentheses indicate theoretical value): Asp2.00 (2); Leu1.92 (2) LSIMS (M + H ⁺ ) = 643 (theoretical value) = 643

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁵ Identification code Office reference number FI technical display area C07K 1/02 // C07K 99:00

Claims (3)

[Claims] 1. A general formula Cyclo (-A-B-C-D-E-) [wherein A is a D-acidic-α-amino acid residue and B and D are L-α-amino acid residues. , C represents a D-neutral-α-amino acid residue, and E represents a D-α-amino acid residue having an aromatic ring group. However, A is D-Glu, B is L-Ala, and C is Da.
Ile is D is L-Leu, L-Val or L-Nva, or A is
D-Glu, B is L-Ala, C is D-Val or D-Leu, D
Is L-Leu or L-Val, or A is D-Glu and B is
When L-Ala is C-D-Nva and D is L-Val or L-Nva, E is not D-Trp but A is D-Glu and B is L-Ala.
When C is D-aIle and D is L-Leu, E is D-mTrp
Not. ] The cyclic pentapeptide represented by these, or its pharmacologically acceptable salt. 2. A general formula H- (X) -OH [wherein, X is -A-B-C-D-E-, -B-C-D-.
E-A-, -C-D-E-A-B-, -D-E-A-B
Represents -C- or -E-A-B-C-D-, A, B,
C, D and E have the same meaning as described in claim 1. ]
The cyclic pentapeptide according to claim 1 or a pharmacological agent thereof, which comprises treating the peptide represented by or a reactive derivative of the peptide with a cyclizing agent, and subjecting the peptide to elimination of a protecting group, if necessary. Acceptable salt production method. 3. An endothelin receptor antagonist comprising the cyclic pentapeptide according to claim 1 or a pharmacologically acceptable salt thereof.