JPH10109999A - Novel cyclic peptide and cardiotonic agent containing it - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a new cyclic peptide having a cyclic depsipeptide skeleton, and useful as an active ingredient for cardiotonic medicines having a strong cardiotonic action and an antiarrhythmic action and not causing side effects such as tachycardia and arrhythmia, etc. SOLUTION: The cyclic peptide of formula I [R<1> is CH2 , CH2 CH2 ; R<2> is H, an alkyl, phenyl, benzyl; R<3> is NH, N(CH3 ), CH2 , O] is useful as an active ingredient for cardiotonic medicines having a strong cardiotonic action and not causing side effects such as tachycardia and arrhythmia, etc., and obtained by condensing a t-butoxycarbonyl-carboxylic acid derivative with proline benzyl ester hydrochloride, removing t-butoxycarbonyl group, reacting the product with a tetrapeptide of formula II (Z is benzyloxycarbonyl; Me is methyl), removing the protecting group, and subsequently subjecting the obtained cyclic peptide of formula III (A.A is an amino acid) to a cyclization reaction.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a novel cyclic peptide and a cardiotonic agent having an antiarrhythmic action containing the same as an active ingredient.

[0002]

2. Description of the Related Art Conventionally, digitalis has been used as a remedy for heart failure, but it has a weak cardiotonic effect, and
Side effects were extremely numerous, including those affecting the heart, and the safety margin was narrow (Hoffman BF and Bigger JT
Jr .: Digitalis and allied cardiac glycosides. In
Goodman and Gilman's "The Pharmacological basis o
f therapeutics. "eds .: Gilman A. G, et al. Pergamo
n Press, pp. 814-839, 1990 .; Poole-Whilson, PA a
nd Robinson K .: Digoxin-A redundant drug incongest
ive cardiac failure.Cardiovasc.Drugs Ther. 2 , 73
3-741, 1989.).

In recent years developed beta ₁ receptor agonists and cyclic AMP enzyme inhibitor, has the effect of increasing the heart rate, there is a drawback that cause arrhythmias as a side effect. Furthermore, the latter drugs have been reported to exacerbate the underlying disease when used in long-term treatment (Pa
cker M. et al .: Hemodynamic and clinical limitatio
n of long-term inotropic therapy with amrinone in
patients with severe chronic heart failure.Circul
aton 70 , 1038-1047, 1984 .; Packer M. et al .: Effect
of oral milrinone on mortality in severe chronic
heart failure.New Engl. J. Med., 325 , 1468-1475, 1
991.).

Therefore, the present inventors have a strong cardiotonic effect,
In addition, they searched for cardiotonic agents with few side effects, and found that compounds having a cyclic depsipeptide skeleton had a strong cardiotonic effect and had few side effects.
7-138290, JP-A-7-188286).

[0005]

Accordingly, an object of the present invention is to provide a cardiotonic agent which has a strong cardiotonic effect and does not cause side effects such as tachycardia and arrhythmia.

[0006]

Means for Solving the Problems The present inventors have synthesized various compounds having a cyclic depsipeptide skeleton and made an intensive search to obtain a cardiotonic agent having few side effects from the compounds.

[0007]

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Wherein R ¹ is —CH ₂ — or —CH ₂ CH ₂ —, R ² is a hydrogen atom or an alkyl group or a phenyl or benzyl group, and R ³ is —NH— or —N (CH ₃ )-or -CH ₂ -or an oxygen atom,
The present inventors have found that the cyclic peptides represented by the following formulas are novel in the literature, and that these compounds have a strong cardiotonic effect having an antiarrhythmic effect, and completed the present invention.

The compound represented by the general formula (1) has a plurality of asymmetric carbon atoms and therefore has stereoisomers, and the present invention includes any of the stereoisomers. The cyclic peptide of the present invention also includes hydrates and solvates.

[0010]

BEST MODE FOR CARRYING OUT THE INVENTION The compounds represented by the general formula (1) can be exemplified by the following structural formulas, but the following compounds can be mentioned, but are not limited thereto. R ¹ is -CH _2- , R
Assuming that ² is an isobutyl group (D form) and R ³ is -NH-,

[0011]

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R ¹ is —CH ₂ —, R ² is an isobutyl group (L-form), R ³
Is -NH-,

[0013]

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R ¹ is —CH ₂ —, R ² is an isobutyl group, and R ³ is —N (CH
₃ )-

[0015]

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In the chemical formula (5), the asymmetric carbon atom represented by * has the configuration of L-form TK2-6A, and the asymmetric carbon atom has the configuration of TK2-TK.
Call it 6B.

R ¹ is —CH ₂ —, R ² is an isopropyl group, and R ³ is —N
(CH ₃ )-

[0018]

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R ¹ is —CH ₂ —, R ² is a methyl group, and R ³ is —N (CH ₃ ) —
As

[0020]

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R ¹ is —CH ₂ —, R ² is a hydrogen atom, and R ³ is —N (CH ₃ ) —
As

[0022]

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Assuming that R ¹ is —CH ₂ —, R ² is an isobutyl group, and R ³ is an oxygen atom,

[0024]

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Assuming that R ¹ is —CH ₂ —CH ₂ —, R ² is an isobutyl group, and R ³ is an oxygen atom,

[0026]

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R ¹ is —CH ₂ —, R ² is a butyl group, and R ³ is —N (CH ₃ ) —
As

[0028]

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R ¹ is —CH ₂ —, R ² is a sec-butyl group, and R ³ is —N (CH
₃ )-

[0030]

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R ¹ is —CH ₂ —, R ² is a benzyl group, and R ³ is —N (CH ₃ )
-

[0032]

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R ¹ is —CH ₂ —CH ₂ —, R ² is an isobutyl group, and R ³ is —
N (CH ₃ )-

[0034]

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R ¹ is —CH ₂ —, R ² is an isobutyl group, and R ³ is —CH ₂ —
As

[0036]

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In formula (15), the asymmetric carbon atom represented by * has the L-configuration as the TK16-22A and the D-configuration as the asymmetric carbon atom.
Called TK16-22B.

Among the compounds represented by the general formula (1) of the present invention, the compounds wherein R ³ is —NH or —N (CH ₃ ) — can be produced, for example, according to the following reaction formula. Can be. On the other hand, compounds in which R ³ is —CH ₂ — or an oxygen atom, that is, Example 8 for TK3-8, Example 9 for TK11-3, and Example 14 for TK16-22A and TK16-22B.
Shows an example of each manufacturing method.

[0039]

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(Where AA represents an amino acid, Me represents a methyl group, R ¹ represents —CH ₂ or —CH ₂ CH ₂ —, R ⁴ represents a side chain of the amino acid,
R ⁵ represents a hydrogen atom or a methyl group, respectively)

That is, the t-butoxycarbonyl-carboxylic acid derivative (2) is condensed with proline benzyl ester hydrochloride (3) to obtain t-butoxycarbonyl-AA-proline benzyl ester (4), from which a protecting group is added. After removal to give -AA-proline benzyl ester hydrochloride (5), (5) was condensed with a separately prepared intermediate benzyloxycarbonyl-isoleucyl-methylvalyl-methylalanyl-β-alanine (6). Benzyloxycarbonyl-isoleucyl-methylvalyl-
Methylalanyl-β-alanyl-AA-proline benzyl ester (7) is obtained. By removing the protecting group of (7), isoleucyl-methylvalyl-methylalanyl-β-alanyl-AA-proline (8) is obtained, and a cyclization reaction is performed on (8). R ³ is -NH or-
Compound (1 ′), which is N (CH ₃ ) —, is obtained.

First, a t-butoxycarbonyl-carboxylic acid derivative (2) and proline benzyl ester hydrochloride (3)
Condensation reaction with water-soluble carbodiimide (WSCD)
Is carried out in the presence of an additive such as 1-hydroxybenzotriazole (hereinafter referred to as HOBt). Deprotection of t-butoxycarbonyl-AA-proline benzyl ester (4) (removal of t-butoxycarbonyl group) can be carried out by using trifluoroacetic acid, hydrochloric acid / acetic acid,
The reaction is performed by reacting hydrochloric acid / dioxane or the like.
The obtained AA-proline benzyl ester hydrochloride (5)
And production intermediate benzyloxycarbonyl-isoleucyl-methylvalyl-methylalanyl-β-alanine (6)
Is carried out, for example, by reacting WSCD in the presence of HOBt. Deprotection (cleavage of the benzyloxycarbonyl group and the benzyl group) of the obtained compound benzyloxycarbonyl-isoleucyl-methylvalyl-methylalanyl-β-alanyl-AA-proline benzyl ester (7) is carried out by using hydrogen gas in the presence of a palladium catalyst. The reaction is carried out by catalytic reduction, anhydrous hydrogen fluoride, trifluoromethanesulfonic acid or the like. The condensation / cyclization reaction of the resulting compound isoleucyl-methylvalyl-methylalanyl-β-alanyl-AA-proline (8)
-(7-Azabenzotriazole) -1,1,3,3-tetramethyluronium The condensation reaction is carried out in the presence of hexafluorophosphate (HATU) and diisopropylethylamine (DIEA).

The production intermediate benzyloxycarbonyl-isoleucyl-methylvalyl-methylalanyl-β
-Alanine (6) is produced, for example, according to the following reaction formula.

[0044]

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That is, t-butoxycarbonyl-β-alanine (9) was converted to t-butoxycarbonyl-β-alanine phenacyl ester (10) in the presence of phenacyl bromide and trimethylamine to obtain t-butoxy of (10). The carbonyl group is removed with trifluoroacetic acid, hydrochloric acid / acetic acid, hydrochloric acid / dioxane or the like to obtain β-alanine phenacyl ester hydrochloride (11). A commercially available t-butoxycarbonyl-methylalanine is condensed with (11) to obtain t-butoxycarbonyl-methylalanyl-β-alanine phenacyl ester (12), from which the t-butoxycarbonyl group is removed to remove methylalanyl-β-alanine. Alanine phenacyl ester hydrochloride (13) is obtained. Subsequently, (13) is subjected to a condensation reaction with t-butoxycarbonyl-methylvaline, and the t-butoxycarbonyl group is removed to remove methylvalyl-methylalanyl-.
β-alanine phenacyl ester hydrochloride (14) is obtained. (14) is condensed with benzyloxycarbonyl-isoleucine to give benzyloxycarbonyl-isoleucyl-methylvalyl-methylalanyl-β-alaninephenacyl ester (15), which is deprotected and benzyloxycarbonyl-isoleucyl-methylvalyl-methylalanyl -Β-alanine (6) is obtained.

The compound represented by the general formula (1) obtained as described above can be administered by any of oral and parenteral methods.
Various dosage forms such as powders, granules, tablets, dragees, capsules, ampoules and the like; oral administration; subcutaneous, intramuscular or intravenous injections; suppositories and the like. The above-mentioned preparation comprises the compound represented by the general formula (1) alone or the compound represented by the general formula (1) and an excipient, a bulking agent, a binder, a wetting agent, a disintegrant, a surfactant, a lubricant. It can be produced by formulating an appropriate combination of agents, dispersants, buffering agents, preservatives, flavoring agents, flavors, coating agents and the like.

The dosage of the thus-obtained cardiotonic agent of the present invention varies depending on symptoms, administration route, etc. In general, in adults, the compound represented by the general formula (1) is used in an amount of 0.5 to 5%.
00 mg / day, preferably 1-200 mg / day, usually 1
It is preferable to administer the drug in 3 to 4 times a day.

[0048]

Industrial Applicability The present invention provides a novel cyclic peptide and a useful cardiotonic agent containing the same as an active ingredient and having an antiarrhythmic effect.

[0049]

EXAMPLES The present invention will be described more specifically with reference to examples and test examples, but the present invention is not limited to these examples. The configuration of the amino acid is L-form unless otherwise specified.

Boc represents a t-butoxycarbonyl group,
ac represents a phenacyl group, Z represents a benzyloxycarbonyl group, Bzl represents a benzyl ester group, and OLeu represents leucic acid.

Further, ¹ H-NMR was measured by JNM-EX27 manufactured by JEOL Ltd.
270 MHz in deuterated chloroform solution using FT NMR SYSTEM
The z ¹ H-NMR spectrum was measured. The reference substance used was tetramethylsilane.

In the HPLC analysis, a capsule pack was used as a column.
Using C18 (4.6φx150mm, manufactured by Shiseido), a solvent having the following composition (A: B = 70: 30 mixed solution, pH 5.
6) and eluted at a flow rate of 1 ml / min. Column temperature is
The temperature was 35 ° C., and detection was performed at UV 220 nm.

[0053]

[Table 1]

Example 1 (Synthetic intermediate Z-Ile-MeVal-MeAla-
Synthesis of β-Ala-OH (6)) (a) Synthesis of Boc-β-Ala-OPac Boc-β-Ala-OH (10.0 g) was dissolved in acetone (100 ml), and triethylamine (8.1 ml) was added under ice-cooling. ) Was added dropwise, followed by the addition of phenacyl bromide (11.57 g), and the mixture was stirred under ice-cooling for 5 minutes and further at room temperature for 24 hours. After acetone was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. After evaporating the solvent under reduced pressure, the residue was crystallized with hexane to obtain Boc-β-Ala-OPac (14.7 g).

(B) Synthesis of HCl-H-β-Ala-OPac 5.0N, HCl / dioxane (189 ml) was added to Boc-β-Ala-OPac (14.54 g), and the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The precipitate is washed with ether and dried under reduced pressure to obtain HClH-β-Ala-O
Pac (11.2 g) was obtained.

(C) Synthesis of Boc-MeAla-β-Ala-OPac HCl / H-β-Ala-OPac (5.00 g) was dissolved in DMF (25 ml) and Boc-MeAla-OH (4.37 g) under ice-cooling. , HOAt (3.07g)
WSCD (4.94 ml) was added dropwise. The mixture was stirred for 30 minutes under ice cooling and further for 18 hours at room temperature. DMF was distilled off under reduced pressure, and the residue was dissolved in ethyl acetate. The extract was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. This was purified by silica gel column chromatography (silica gel 288 g, toluene: ethyl acetate = 2: 1) to give Boc-MeAla-
β-Ala-OPac (8.13 g) was obtained.

(D) Synthesis of HCl.H-MeAla-β-Ala-OPac 4.5N HCl / dioxane (101 ml) was added to Boc-MeAla-β-Ala-OPac (8.92 g), and then at room temperature for 1 hour. I left it. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The precipitate was washed with ether and dried under reduced pressure to obtain HClH-MeAl
a-β-Ala-OPac (7.32 g) was obtained.

(E) Synthesis of HCl.H-MeVal-MeAla-β-Ala-OPac HCl.H-MeAla-β-Ala-OPac (7.00 g), Boc-MeVal-OH (5.
18g) and HOAt (3.20g) dissolved in DMF (35ml)
SCD (5.15 ml) was added dropwise. 1 hour under ice cooling and at room temperature
Stirred for 16 hours. DMF was distilled off under reduced pressure, and the residue was dissolved in ethyl acetate. The extract was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure to obtain 11.2 g of a residue. After adding 4.5N HCl / dioxane (95.1 ml) to the residue, the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The precipitate is washed with ether and dried under reduced pressure to obtain HClH-MeVal-MeAla-β-Ala-OPa.
c (8.71 g) was obtained.

(F) Synthesis of Z-Ile-MeVal-MeAla-β-Ala-OPac HCl · H-MeVal-MeAla-β-Ala-OPac (8.50 g), Z-Ile-OH
(5.37 g) and HOAt (2.89 g) were dissolved in DMF (45 ml), and WSCD (4.64 ml) was added dropwise under ice cooling. The mixture was stirred for 1 hour under ice cooling and further for 17 hours at room temperature. DMF was distilled off under reduced pressure, and the residue was dissolved in ethyl acetate. The extract was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. Purified by silica gel column chromatography (silica gel 480 g, chloroform: methanol = 100: 1), Z-Ile-MeV
al-MeAla-β-Ala-OPac (10.7 g) was obtained.

(G) Z-Ile-MeVal-MeAla-β-Ala-OH
Synthesis of (6) Z-Ile-MeVal-MeAla-β-Ala-OPac (10.7 g) was dissolved in a 90% acetic acid aqueous solution (200 ml), and zinc powder (53.7 g) was added.
Ultrasonic stirring for hours. After filtering the zinc powder, acetic acid was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, dried over magnesium sulfate, and the solvent was distilled off under reduced pressure. The residue was washed twice with hexane and purified by silica gel column chromatography (450 g of silica gel, chloroform: methanol = 19: 1). The residue was crystallized from hexane to obtain Z-Ile-MeVal-MeAla-β-Ala-OH (6.05 g).

Example 2 (Production of TK1-19: AA is D-leucine in the production process described in Chemical formula 16) (a) Synthesis of HCl.H-Pro-OBzl Boc-Pro-OH (2.00 g) was dissolved in acetone (20 ml), and triethylamine (1.42 ml) and benzyl bromide (1.22 ml) were added under ice-cooling. 5 minutes under ice-cooling and 23 at room temperature
Stirred at 40 degrees for 4 hours. After acetone was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, and 10% citric acid, saturated saline,
Wash with saturated aqueous sodium bicarbonate and saturated saline in that order,
After drying over anhydrous magnesium sulfate, the solvent was distilled off under reduced pressure to obtain a residue. 5.0N HCl / dioxane (55.6ml)
Was added and left at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The precipitate was washed with ether, dried under reduced pressure, and treated with HCl.H-Pro-OBzl (2.32
g) was obtained.

(B) Synthesis of Boc-D-Leu-Pro-OBzl HCl / H-Pro-OBzl (1.00 g) was dissolved in DMF (5 ml), and Boc-D-Leu (1.08 g) and HOBt were dissolved under ice cooling. (0.62 g) was added. WSCD
(1.00 ml) was added dropwise, and the mixture was stirred for 1 hour under ice-cooling and further for 17 hours at room temperature. DMF was distilled off under reduced pressure, and the residue was dissolved in ethyl acetate. The extract was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. Purification was performed by silica gel column chromatography (silica gel 54 g, toluene: ethyl acetate = 10: 1), and Boc-D-Leu-Pro-OBzl (1.70 g) was used.
g) was obtained.

(C) Synthesis of HCl-HD-Leu-Pro-OBzl 5.0N HCl / dioxane (16.3 ml) was added to Boc-D-Leu-Pro-OBzl (1.7 g) and left at room temperature for 1 hour. did. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure to obtain HCl-HD-Leu-
Pro-OBzl (1.20 g) was obtained.

(D) Z-Ile-MeVal-MeAla-β-Ala-D-Leu-P
ro-OBzl Z-Ile-MeVal-MeAla-β-Ala-OH (0.4
03g) was dissolved in DMF (25ml). HCl ・ HD-Leu-
After adding Pro-OBzl (0.282 g) and HOBt (112 mg), WSCD
(0.182 ml) was added dropwise. 2 hours under ice-cooling, then 20 at room temperature
Stirred for hours. After DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. To give a residue. Purified by silica gel column chromatography (silica gel 26 g, eluted with chloroform only), Z-Ile-MeVal-MeAla-
β-Ala-D-Leu-Pro-OBzl (0.485 g) was obtained.

(E) H-Ile-MeVal-MeAla-β-Ala-D-Leu-P
Synthesis of ro-OH Z-Ile-MeVal-MeAla-β-Ala-D-Leu-Pro-OBzl (360 mg) was dissolved in methanol: acetic acid: water = 8: 2: 1 (30 ml) and palladium black (spartel small) was dissolved. 2 spoons), and hydrogen gas was blown for 1 hour with stirring at room temperature. The palladium was filtered off, the mother liquor was concentrated under reduced pressure, dissolved in water and freeze-dried to give H-Ile-MeVa
1-MeAla-β-Ala-D-Leu-Pro-OH (256 mg) was obtained.

(F) Synthesis of TK1-19 H-Ile-MeVal-MeAla-β-Ala-D-Leu-Pro-OH (49.7 mg)
Was dissolved in dichloromethane (25 ml) and DIEA (85.2 μl
l) was added. Put this in a 25 ml syringe and use HATU (93.
(0 mg) was added dropwise over 2 hours to a dichloromethane solution (57 ml) containing the solution, followed by stirring at room temperature for 17 hours. After dichloromethane was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated brine, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by silica gel thin-layer chromatography (chloroform: methanol = 9: 1) to obtain TK1-19 (8.82 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ8.17 (t, 1
H, β-Ala NH), 7.25 (d, 1H, Ile NH), 5.78 (d, 1H, Le
u NH), 5.20 (q, 1H, MeAla α), 5.02 (d, 1H, Pro
α), 4.82 (q, 1H, Ile α), 4.64 (d, 1H, MeVal α),
4.38 (m, 1H, Leu α), 4.10 (t, 1H, Pro δ ₁ ), 3.95 (m,
1H, β-Ala β ₁ ), 3.49 (m, 1H, Pro δ ₂ ), 3.22 (s, 3H,
N-Me), 3.17 (d, 1H, β-Ala β ₂ ), 2.73 (s, 3H, NM
e), 2.36-2.26 (m, 4H, Proββ-Alaα), 2.08-1.96
(m, 4H, Ileβ Pro γMeVal β), 1.56-1.38 (m, 5H, Ile
γ ₁ Leu βγ), 1.01-0.82 (m, 18H, Ile γ ₂ δ MeVal
γLeuδ) PD-MS: m / z 593.7 (M + H) + C 30 calculated for H ₅₂ O ₆ N _6: 592.4

Example 3 (Production of TK4-5: AA is L-leucine in the production process described in Chemical formula 16) (a) Synthesis of Boc-L-Leu-Pro-OBzl HCl.H-Pro-OBzl (0.50 g) was dissolved in DMF (3 ml), and Boc-L-Leu (0.541 g) and HOBt (0.31 g) were added under ice cooling. WSCD
(0.50 ml) was added dropwise, and the mixture was stirred for 1 hour under ice cooling and further for 17 hours at room temperature. DMF was distilled off under reduced pressure, and the residue was dissolved in ethyl acetate. The extract was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. Purified by silica gel column chromatography (silica gel 40 g, chloroform: methanol = 100: 1), Boc-L-Leu-Pro-OBzl
(0.65 g) was obtained.

(B) Synthesis of HCl-HL-Leu-Pro-OBzl 4.5N HCl / dioxane (6.90 ml) was added to Boc-L-Leu-Pro-OBZl (0.65 g), and then left at room temperature for 1 hour. did. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure to obtain HCl-HL-Leu-
Pro-OBzl (0.493 g) was obtained.

(C) Z-Ile-MeVal-MeAla-β-Ala-L-Leu-P
Synthesis of ro-OBzl Z-Ile-MeVal-MeAla-β-Ala-OH (0.2
11g) was dissolved in DMF (3ml). HCl ・ HL-Leu-P under ice cooling
After adding ro-OBzl (0.147g) and HATU (0.225g), DIEA
(0.206 ml) was added dropwise. 2 hours under ice-cooling, then 20 at room temperature
Stirred for hours. After DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. To give a residue. Purified by silica gel column chromatography (silica gel 25 g, chloroform: methanol = 30: 1), Z-Ile-MeVa
1-MeAla-β-Ala-L-Leu-Pro-OBzl (0.237 g) was obtained.

(D) H-Ile-MeVal-MeAla-β-Ala-L-Leu-P
Synthesis of ro-OH Z-Ile-MeVal-MeAla-β-Ala-L-Leu-Pro-OBzl (0.237g)
Was dissolved in methanol: acetic acid: water = 8: 2: 1 (30 ml), palladium black (2 teaspoons of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. Filter the palladium,
After concentrating the mother liquor under reduced pressure, dissolve in water and freeze-dry
eVal-MeAla-β-Ala-L-Leu-Pro-OH (0.175 g) was obtained.

(E) Synthesis of TK4-5 H-Ile-MeVal-MeAla-β-Ala-L-Leu-Pro-OH (50.0 mg) was dissolved in dichloromethane (25 ml), and DIEA (85.6 μm) was added thereto.
l) was added. Put this in a 25 ml syringe and use HATU (93.
The solution was added dropwise over 2 hours to a dichloromethane solution (57 ml) containing 4 mg), and then stirred at room temperature for 18 hours. After dichloromethane was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate.
The solvent was distilled off under reduced pressure to obtain a residue. Purification by silica gel thin layer chromatography (chloroform: methanol = 9: 1) gave TK4-5 (5.43 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ8.56 (d, 1H,
Ile NH), 7.90 (d, 1H, β-Ala NH), 5.86 (s, 1H, Leu N
H), 5.05 (d, 1H, MeVal α), 4.98 (q, 1H, MeAla α),
4.56 (t1H, Ile α), 4.28 (d, 1H, Leu α), 4.19 (d, 1
H, Pro α), 3.99 (m, 1H, β-Ala β ₁ ), 3.65-3.59 (m,
2H, Pro δ), 3.24 (s, 3H, N-Me), 3.13 (m, 1H, β-Ala
β ₂ ), 2.72 (s, 3H, N-Me), 2.42 (m, 2H, β-Ala α),
2.33 (m, 1H, MeVal β), 2.27-2.17 (m, 3H, Ile β Pro
β), 1.92 (m, 2H, Pro γ), 1.33 (d, 3H, MeAla β),
1.28-1.25 (m, 3H, Leu β Leu γ), 1.00-0.75 (m, 18H,
Ile γ ₂ δ MeVal γ Leu δ) PD-MS: m / z 594.0 (M + H) ⁺ C ₃₀ H ₅₂ O ₆ N ₆ Calculated: 592.4

Example 4 (Production of TK2-6-A and TK2-6-B: In the production process described in Chemical formula 16, AA is L-methylleucine and D-methylleucine, respectively). (A) Boc-DL-MeLeu Synthesis of Boc-D-Leu-OH (4.63g) in anhydrous THF under argon atmosphere
(10 ml) and dissolve it in potassium hydride (2.23
g) and 18-crown-6 (0.246 g) in THF (40 ml). After dropwise addition of methyl iodide (2.32 ml), the mixture was stirred in a water bath (20 ° C.) for 15 hours. Glacial acetic acid (2 ml) was added to the reaction solution to decompose excess base, and then 2N sodium hydroxide was added to adjust the pH to 8.5, followed by washing with ethyl acetate. The separated aqueous layer was adjusted to pH 2.5 with 2N HCl and extracted with ethyl acetate. After drying over anhydrous magnesium sulfate, the solvent was distilled off under reduced pressure to obtain a residue. Silica gel column chromatography (400 g silica gel, chloroform: methanol =
19: 1) to obtain Boc-DL-MeLeu-OH (1.37 g). N
α- (2,4-dihydro-5-fluorophenyl) -L /
D-alanine amide (FDAA) derivatization and analysis by HPLC confirmed that racemization had occurred during this methylation reaction. The D and L isomers were used without separation in the following synthesis.

(B) Synthesis of Boc-DL-MeLeu-L-Pro-OBzl HCl · H-Pro-OBzl (0.500 g), Boc-DL-MeLeu-OH (0.533)
g), HOAt (0.310g) dissolved in DMF (3ml)
D (0.498 ml) was added dropwise, and the mixture was cooled for 1 hour under ice-cooling and then at room temperature for 18 hours.
Stirred for hours. After DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. A residue was obtained. Purified by silica gel column chromatography (silica gel 40 g, chloroform: methanol = 50: 1), Boc-DL-MeLeu
-L-Pro-OBzl (0.520 g) was obtained.

(C) Synthesis of HCl.H-DL-MeLeu-L-Pro-OBzl After adding 4.5N HCl / dioxane (5.4 ml) to Boc-DL-MeLeu-L-Pro-OBzl (0.52 g). And left at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure to obtain HClH-DL
-MeLeu-L-Pro-OBzl (0.368 g) was obtained.

(D) Z-Ile-MeVal-MeAla-β-Ala-DL-MeLe
Synthesis of u-Pro-OBzl Z-Ile-MeVal-MeAla-β-Ala-OH (0.219 g) in DMF (3 ml)
Melted into. HCl ・ H-DL-MeLeu-Pro-OBzl (0.159
g) and HATU (0.234 g) were added, and then DIEA (0.214 ml) was added dropwise. The mixture was stirred for 1 hour under ice cooling and further for 17 hours at room temperature. DMF
Was distilled off under reduced pressure, dissolved in ethyl acetate, washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated brine, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. A residue was obtained. Purification by silica gel column chromatography (silica gel 25 g, chloroform: methanol = 100: 1), Z-Ile-MeVal-MeAla-β-Ala
-DL-MeLeu-Pro-OBzl (0.227 g) was obtained.

(E) H-Ile-MeVal-MeAla-β-Ala-DL-MeLe
Synthesis of u-Pro-OH Z-Ile-MeVal-MeAla-β-Ala-DL-MeLeu-Pro-OBzl (0.227
g) in methanol: acetic acid: water = 8: 2: 1 (30 ml),
Palladium black (2 teaspoons of spatula) was added, and hydrogen gas was blown for 1 hour with stirring at room temperature. Filter the palladium, concentrate the mother liquor under reduced pressure, dissolve in water and freeze-dry
le-MeVal-MeAla-β-Ala-DL-MeLeu-Pro-OH (0.171 g) was obtained.

(F) Synthesis of Synthesis of TK2-6-A and TK2-6-B H-Ile-MeVal-MeAla-β-Ala-DL-MeLeu-Pro-OH (0.50 g)
Was dissolved in dichloromethane (25 ml), and DIEA (83.8
μl) was added. Put this in a 25 ml syringe and add HATU (9
The solution was added dropwise to a dichloromethane solution (1.4 ml) (55 ml) over 2 hours, and then stirred at room temperature for 17 hours. After the dichloromethane was distilled off under reduced pressure, the precipitate was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel thin layer chromatography (chloroform: methanol = 19:
TK2-6-A (6.48 mg) and TK2-6-B (5.1
0 mg).

¹ H-NMR of TK2-6-A (270 MHz, CDCl ₃ ): δ9.
03 (d, 1H, Ile NH), 7.83 (1H, β-Ala NH), 4.99 (q, 1
H, MeAla α), 4.94 (d, 1H, Pro α), 4.82 (q, 1H, Ile
α), 4.50 (q, 1H, MeVal α), 4.29 (d, 1H, MeLeu α),
3.98-3.90 (m, 1H, β-Alaβ ₁ ), 3.59-3.30 (m, 2H, Pro
δ), 3.14 (s, 3H, N-Me), 3.07-3.04 (m, 1H, β-Ala β
₂ ), 3.02 (s, 3H, N-Me), 2.66 (s, 3H, N-Me), 2.62-2.3
0 (m, 2H, β-Ala α), 2.00-1.80 (m, 4H, Ile β MeVal
β MeLeu β), 1.66-1.52 (m, 3H, MeLeuγProγ), 1.3
3-1.18 (m, 4H, Ile γ ₁ Pro β), 1.25 (d, 3H, MeAla
β), 0.95-0.72 (m, 18H, Ile γ ₂ δ MeVal γ MeLeu
δ) PD-MS: m / z calcd for 607.9 (M + H) ⁺ C ₃₁ H ₅₄ O ₆ N ₆ : 606.4 HPLC analysis: retention time 11.19 min

¹ H-NMR of TK2-6-B (270 MHz, CDCl ₃ ): δ8.
12 (d, 1H, β-Ala NH), 7.18 (d, 1H, Ile NH), 5.22 (q,
1H, MeAla α), 5.01 (d, 1H, MeVal α), 4.84 (q, 1H,
Ileα), 4.75 (q, 1H, Proα), 4.70 (d, 1H, MeLeu
α), 4.03-3.94 (m, 2H, β-Ala β ₁ Pro δ ₁ ), 3.50 (q,
1H, Pro δ ₂ ), 3.28-3.20 (m, 1H, β-Ala β ₂ ), 3.21
(s, 3H, N-Me), 3.12 (s, 3H, N-Me), 2.73 (s, 3H, NM
e), 2.65-2.42 (m, 2H, β-Ala α), 2.37-2.28 (m, 1H,
MeVal β), 2.07-1.88 (m, 7H, Ile β MeLeu β Pro β
γ), 1.46-1.33 (m, 3H, Ile γ ₁ MeLeu γ), 1.30 (d,
3H, MeAla β), 1.02-0.81 (m, 18H, Ile γ ₂ δ MeVal
γ MeLeu δ) PD-MS: m / z Calculated as 607.8 (M + H) ⁺ C ₃₁ H ₅₄ O ₆ N ₆ : 606.4 HPLC analysis: retention time 15.49 min

The method for confirming the D-form and the L-form is as follows. TK2-6-A and TK2-6-B (1 μmol) each in 6N H
After hydrolyzing with Cl at 150 ° C. for 1 hour, hydrochloric acid was distilled off. The obtained precipitate is dissolved in acetone: water = 1: 1 (1.2 ml) and FD
AA (4.2μmol) and 0.2M sodium carbonate aqueous solution (60μ
l) was added and reacted at 40 ° C. for 1 hour. After adding 1N hydrochloric acid to stop the reaction, the configuration of the constituent amino acids was analyzed by HPLC (YMCPack, ODS). As a result, TK2-6-A is [L-MeLeu ^5] - a derivative, TK2-6-B is [D-MeLeu ^5] - proved to be derivatives.

Example 5 (Production of TK6-5: AA is L-methylvaline in the production process described in Chemical formula 16) (a) Synthesis of HCl.HL-MeVal-L-Pro-OBzl HCl.H-Pro-OBzl (0.551 g), Boc-L-MeVal-OH (0.502
g) and HOAt (0.324 g) in DMF (5 ml), and
SCD (0.521 ml) was added dropwise. The mixture was stirred for 1 hour under ice cooling and further for 18 hours at room temperature. After DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. A residue (0.972 g) was obtained. After 4.5N HCl / dioxane (10.1 ml) was added to the residue, the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The precipitate was washed with ether, dried under reduced pressure, and used for HCl / HL-MeVal-L-Pro-OBzl.
(0.664 g).

(B) H-Ile-MeVal-MeAla-β-Ala-L-MeVal
Synthesis of -Pro-OH Z-Ile-MeVal-MeAla-β-Ala-OH (0.200g), HCl ・ HL-MeV
al-Pro-OBzl (0.139 g) and HOAt (0.056 g) were added to DMF (2 m
l), and WSCD (0.090 ml) was added dropwise under ice cooling. The mixture was stirred for 1 hour under ice cooling, and further for 16 hours at room temperature. After the DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. To give a residue. Silica gel column chromatography (silica gel 20 g, chloroform: methanol = 50: 1)
And purified. This was converted to methanol: acetic acid: water = 8: 2:
Dissolve in 1 (30ml) and palladium black (2 teaspoons spatula)
Cup), and hydrogen gas was blown for 1 hour with stirring at room temperature. After filtering the palladium, concentrating the mother liquor under reduced pressure, dissolving in water and freeze-drying, H-Ile-MeVal-MeAla-β-Ala-L-MeVal
-Pro-OH (0.252 g) was obtained.

(C) Synthesis of TK6-5 H-Ile-MeVal-MeAla-β-Ala-L-MeVal-Pro-OH (0.100 g)
Was dissolved in dichloromethane (25 ml) and DIEA (85.7 μl)
Was added. Put this in a 25ml syringe and use HATU (93.5m
The mixture was added dropwise over 2 hours to a dichloromethane solution (139 ml) containing g), and then stirred at room temperature for 18 hours. After distilling off dichloromethane under reduced pressure, the precipitate was dissolved in ethyl acetate,
The extract was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel thin layer chromatography (chloroform: methanol = 19: 1).
To give TK6-5 (3.61 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ 8.84 (d, 1H,
Ile NH), 8.33 (d, 1H, β-Ala NH), 5.20 (q, 1H, MeAl
a α), 5.06-5.02 (2H, Ile α MeVal α), 4.75 (d, 1H,
Proα), 4.66 (d, 1H, MeVal α), 3.94-3.80 (m, 2H,
β-Ala β ₁ Pro δ ₁ ), 3.28-3.23 (m, 2H, β-Ala β ₂
Pro δ ₂ ), 3.19 (s, 3H, N-Me), 3.14 (s, 3H, N-Me), 2.7
8 (s, 3H, N-Me), 2.58-2.54 (m, 1H, Pro γ ₁ ), 2.40-
2.23 (m, 3H, Ile βMeVal β), 1.94-1.83 (m, 3H, β-A
la α Pro γ ₂ ), 1.77-1.43 (m, 2H, Pro β), 1.32 (d,
3H, MeAla β), 1.19-1.06 (m, 2H, Ile γ ₁ ), 1.03-0.7
2 (m, 18H, Ile γ ₂ δ MeVal γ). PD-MS: m / z 594.0 (M + H) + Molecular formula C ₃₀ calculated for _{H 52 O 6 N 6: 592.4} HPLC analysis: retention time 7.93 min

Example 6 (Production of TK7-5: In the production process described in Chemical formula 16, AA is L-methylalanine) (a) Synthesis of HCl.HL-MeAla-L-Pro-OBzl HCl.H-Pro- OBzl (0.624g), Boc-L-MeAla-OH (0.500g)
And HOAt (0.368g) in DMF (5ml)
(0.593 ml) was added dropwise, and the mixture was stirred for 1 hour under ice cooling and further for 18 hours at room temperature. After DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. A residue (1.05 g) was obtained. After 4.5N HCl / dioxane (10.9 ml) was added to the residue, the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure to obtain HCl · HL-MeAla-L-Pro-OBzl (0.653 g).

(B) H-Ile-MeVal-MeAla-β-Ala-L-MeAla
Synthesis of -Pro-OH Z-Ile-MeVal-MeAla-β-Ala-OH (0.200g), HCl ・ HL-MeA
la-Pro-OBzl (0.128 g) and HOAt (0.056 g) were added to DMF (2 m
l), and WSCD (0.090 ml) was added dropwise under ice cooling. The mixture was stirred for 1 hour under ice cooling, and further for 16 hours at room temperature. After DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline, dried over anhydrous magnesium sulfate, and then the solvent was distilled off under reduced pressure. To give a residue. This was subjected to silica gel column chromatography (silica gel 20 g, chloroform: methanol = 5
0: 1). This is methanol: acetic acid: water =
The mixture was dissolved in 8: 2: 1 (30 ml), palladium black (2 spoons of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. After filtering the palladium, the mother liquor was concentrated under reduced pressure, dissolved in water and freeze-dried to obtain H-Ile-MeVal-MeAla-β-Ala-L-MeAla
-Pro-OH (0.186 g) was obtained.

(C) Synthesis of TK7-5 H-Ile-MeVal-MeAla-β-Ala-L-MeAla-Pro-OH (58.2 mg)
Was dissolved in dichloromethane (25 ml) and DIEA (105 μl) was added. Put this in a 25ml syringe, HATU (114mg)
Was added dropwise over 2 hours to a dichloromethane solution (75 ml) containing, and then stirred at room temperature for 18 hours. Dichloromethane was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1) to obtain TK7-5 (1.5 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ9.15 (d, 1H,
Ile NH), 7.83 (d, 1H, β-Ala NH), 5.10 (q, 1H, MeAl
a α), 5.03-4.96 (2H, Ile α MeVal α), 4,47 (q, 1H,
MeAla α), 4.40 (d, 1H, Pro α), 4.08-3.97 (m, 1H,
β-Ala β ₁ ), 3.73-3.62 (m, 1H, Pro δ ₁ ), 3.48-3.40
(m, 1H, Pro δ ₂ ), 3.21 (s, 3H, N-Me), 3.16-3.11 (m, 1
H, β-Ala β ₂ ), 3.08 (s, 3H, N-Me), 2.72 (s, 3H, NM
e), 2.37-2.31 (m, 1H, MeVal β), 2.61-2.52 (m, 2H,
β-Ala α), 2.06-1.71 (m, 5H, Proγβ Ile β), 1.45
-1.41 (m, 2H, Ileγ ₁ ), 1.38 (d, 3H, MeAla β), 1.30
(d, 3H, MeAla β), 0.94-0.77 (m, 12H, Ile γ ₂ δ MeVa
lγ) PD-MS measurement: m / z 565.9 (M + H) ⁺ molecular formula Calculated as C ₂₈ H ₄₈ O ₆ N ₆ : 564.3 HPLC analysis: retention time 4.15 minutes

Example 7 (Synthesis of TK8-5: In the production process described in Chemical formula 16, AA is Sar (sarcosine)) (a) Synthesis of HCl.H-Sar-L-Pro-OBzl HCl.H-Pro- OBzl (0.670g), Boc-Sar-OH (0.500g) and
Dissolve HOAt (0.396 g) in DMF (5 ml) and add WSCD (0.
637 ml), and the mixture was stirred for 1 hour under ice-cooling and further for 18 hours at room temperature. After distilling off DMF under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to obtain a residue (1.07 g). After 4.5N HCl / dioxane (11.7 ml) was added to the residue, the mixture was allowed to stand at room temperature for 1 hour. Dioxane is distilled off under reduced pressure,
Crystallization was obtained by adding ether. The residue was washed with ether, dried under reduced pressure, and treated with HCl.H-Sar-L-Pro-OBzl (0.575
g) was obtained.

(B) H-Ile-MeVal-MeAla-β-Ala-Sar-Pro
Synthesis of -OH Z-Ile-MeVal-MeAla-β-Ala-OH (0.200 g), HCl-H-Sar-P
ro-OBzl (0.123 g) and HOAt (0.056 g) were dissolved in DMF (2 ml), and WSCD (0.090 ml) was added dropwise under ice cooling. The mixture was stirred for 1 hour under ice cooling, and further for 16 hours at room temperature. DMF is distilled off under reduced pressure, the residue is dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated brine in this order, dried over anhydrous magnesium sulfate, and then the solvent is distilled off under reduced pressure. And a precipitate was obtained. This was purified by silica gel column chromatography (silica gel 20 g, chloroform: methanol = 50: 1). This was dissolved in methanol: acetic acid: water = 8: 2: 1 (30 ml), palladium black (2 spoons of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. After filtering the palladium, concentrating the mother liquor under reduced pressure, dissolving in water and freeze-drying, H-Ile-MeVal-MeAla-β-Ala-Sar-P
ro-OH (0.174 g) was obtained.

(C) Synthesis of TK8-5 H-Ile-MeVal-MeAla-β-Ala-Sar-Pro-OH (56.8 mg) was dissolved in dichloromethane (25 ml), and DIEA (105 μl) was added. This was put into a 25 ml syringe, dropped into a dichloromethane solution (75 ml) containing HATU (114 mg) over 2 hours, and then stirred at room temperature for 18 hours. After dichloromethane was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate.
The solvent was distilled off under reduced pressure to obtain a residue. This was subjected to silica gel thin layer chromatography (chloroform: methanol = 19:
Separation by 1) gave TK8-5 (2.91 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ 8.05 (d, 1H,
β-Ala NH), 7.42 (d, 1H, Ile NH), 5.29 (q, 1H, MeAl
a α), 4.99 (q, 1H, MeVal α), 4.86 (q, 1H, Ile α),
4.74 (d, 1H, Pro α), 4.51 (d, 1H, Sar α ₁ ), 4.08-
3.97 (m, 1H, β-Ala β ₁ ), 3.91-3.85 (m, 1H, Pro
δ ₁ ), 3.48-3.45 (m, 1H, Pro δ ₂ ), 3.27 (d, 1H, Sar
α ₂ ), 3.22-3.13 (m, 1H, β-Ala β ₂ ), 3.21 (s, 3H, N-
Me), 3.20 (s, 3H, N-Me), 2.74 (s, 3H, N-Me), 2.71-2.
65 (m, 1H, β-Ala α ₁ ), 2.47 (m, 1H, β-Ala α ₂ ), 2.4
1-2.31 (m, 1H, MeVal β), 2.07-1.95 (m, 5H, Ile β P
ro γβ), 1.47-1.36 (m, 2H, Ile γ ₁ ), 1.30 (d, 3H, M
eAla β), 0.95-0.80 (m, 12H, Ile γ ₂ δ MeVal γ) PD-MS measurement: m / z 551.7 (M + H) ⁺ calculated value as molecular formula C ₂₇ H ₄₆ O ₆ N ₆ : 550.3 HPLC analysis: retention time 3.77 minutes

Example 8 (Production of TK3-8) This compound was produced according to the following production method.

[0096]

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[0097]

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(A) Synthesis of L-leucine phenacyl ester L-leucine (3.00 g) was dissolved in a solvent (150 ml) of water: 1N hydrochloric acid: acetic acid = 4.5: 1: 2, and the solution was added under ice cooling. An aqueous solution of sodium nitrate (16.1 g / 28 ml of water) was added dropwise over 40 minutes, and the mixture was further stirred at room temperature for 1 hour. After evaporating the solvent under reduced pressure, the residue was dissolved in water, adjusted to pH 3 with hydrochloric acid, and extracted with ether. The ether layer was washed with a saturated saline solution and then distilled off under reduced pressure to obtain 2.53 g of a precipitate. Dissolve (2.00 g) of this precipitate in ethyl acetate,
After adding triethylamine (2.32 ml) under ice-cooling, phenacyl bromide (3.31 g) was further added, and the mixture was stirred at room temperature for 41 hours. After evaporating the solvent under reduced pressure, the residue was dissolved in ethyl acetate, washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated brine, dried over anhydrous magnesium sulfate, and then ethyl acetate was distilled off under reduced pressure. . Dissolve the precipitate in chloroform and use silica gel column chromatography (silica gel).
(180 g, eluted with chloroform only) to obtain L-leucinate phenacyl ester (1.86 g).

(B) Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-
Synthesis of OPac Z-Ile-MeVal-MeAla-β-Ala-OH (0.500 g), phenacyl L-leucinate (0.257 g) and 4-pyrrolidinopyridine (41.6 mg) were dissolved in DMF (2 ml), DCC under ice cooling (213
mg), and the mixture was stirred under ice-cooling for 30 minutes and further at room temperature for 15 hours. Ethyl acetate was added to the reaction solution, and the insolubles were removed from the furnace. This was washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. The residue was dissolved in chloroform: methanol = 100: 1, purified by silica gel column chromatography (silica gel 40 g, chloroform: methanol = 100: 1), and purified by Z-Ile-MeVal-MeAla-β-Ala-LO.
Leu-OPac (378 mg) was obtained.

(C) Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-
Synthesis of OH Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-OPac (0.378 g)
Dissolve in 20% aqueous acetic acid (20 ml), add zinc powder (1.61 g),
Sonication for 2 hours. After filtering the zinc powder, acetic acid was distilled off under reduced pressure. The residue was dissolved in ethyl acetate, washed with 10% citric acid, and dried over magnesium sulfate. The residue obtained by evaporating the solvent under reduced pressure was dissolved in chloroform: methanol = 19: 1, and the mixture was subjected to silica gel column chromatography (silica gel).
40 g, purified with chloroform: methanol = 19: 1)
le-MeVal-MeAla-β-Ala-L-OLeu-OH (250 mg) was obtained.

(D) Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-
Synthesis of Pro-OBzl Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-OH (100mg), HCl ・ H
-Pro-OBzl (39.2 mg) and HOAt (23.1 mg) were dissolved in DMF (2 ml), WSCD (37.2 μl) was added dropwise under ice cooling, and the mixture was stirred for 1 hour under ice cooling and further at room temperature for 16 hours. After distilling off DMF under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, the residue was dissolved in chloroform: methanol = 100: 1, and silica gel column chromatography (silica gel)
10 g, purified by chloroform: methanol = 100: 1)
Ile-MeVal-MeAla-β-Ala-L-OLeu-Pro-OBzl (86.9 mg) was obtained.

(E) H-Ile-MeVal-MeAla-β-Ala-L-OLeu-
Synthesis of Pro-OH Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-Pro-OBzl (86.9mg)
Was dissolved in methanol: acetic acid: water = 8: 2: 1 (30 ml), palladium black (2 teaspoons of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. Filter the palladium,
After concentrating the mother liquor under reduced pressure, dissolve in water and freeze-dry to give H-Ile-M
eVal-MeAla-β-Ala-L-OLeu-Pro-OH (64.0 mg) was obtained.

(F) Synthesis of TK3-8 H-Ile-MeVal-MeAla-β-Ala-L-OLeu-Pro-OH (50.0 mg) was dissolved in dichloromethane (25 ml), and DIEA (42.7 μl) was added. . Put this in a 25ml syringe, HATU (46.9mg)
Was added dropwise over 2 hours to a dichloromethane solution (57 ml) containing. Then, after stirring at room temperature for 16 hours, the solvent was distilled off under reduced pressure. The residue was dissolved in ethyl acetate, washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline, dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure to obtain a residue. This was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1) to obtain TK3-8 (5.90 mg).

TK3-8 has the following physicochemical properties. (1) Shape and color White powder (2) Molecular formula C ₃₀ H ₅₁ N ₅ O ₇ (3) Molecular weight Measured value of PD-MS m / z 594.6 (M + H) ⁺ (4) Elemental analysis FISONS EA1108 The experimental values measured with a type element analyzer were
C59.33, H8.99, N11.73. Calculated for _{C 30 H 51 N 5 O 7} · H 2 O is, C58.90, H8.73, N1
1.45 (5) Melting point The temperature was 177.0 to 180.0 ° C. as measured by a Buch-type B-530 thermometer with immersion wire. (6) Specific rotation Measured by Horiba SEPA-300 type optical rotation,
[α] ²⁵ _{D was} −235.6 ° (c 1.0, methanol). (7) Ultraviolet absorption spectrum Measured with an ultraviolet-visible spectrophotometer (UV-160A, manufactured by Shimadzu Corporation). λmax (methanol) nm (ε): 208 (21300 ± 900) (8) Solubility Soluble: methanol, ethanol, chloroform Insoluble: water (9) Infrared absorption spectrum FT-IR (Perkin Elmer System2000) by KBr tablet method
Was used to measure the infrared absorption spectrum. Figure 1 shows the results
Shown in (10) ¹ H-NMR spectrum ¹ H-NMR (270 MHz, CDCl ₃ ): δ 7.92 (d, 1 H, Ile NH),
7.84 (d, 1H, β-Ala NH), 5.03 (d, 1H, MeVal α), 4.9
9-4.89 (m, 2H, MeAla α, OLeu α), 4.58 (t, 1H, Ile
α), 4.15 (d, 1H, Pro α), 4.08-4.00 (m, 1H, β-Ala
β ₁ ), 3.65-3.60 (m, 2H, Pro δ), 3.42-3.40 (m, 1H,
β-Ala β ₂ ), 3.25 (s, 3H, N-Me), 2.71 (s, 3H, N-Me),
2.60 (m, 2H, β-Ala α), 2.49-2.31 (m, 2H, Pro β),
2.28-2.07 (m, 1H, MeVal β), 1.96-1.82 (m, 1H, Ile
β), 1.80-1.67 (m, 4H, OLeu β, Pro γ), 1.34 (d, 3
H, MeAla β), 1.28-1.12 (m, 1H, OLeu γ), 1.28-1.12
(m, 2H, Ile γ ₁ ), 0.98-0.80 (m, 18H, Ile γ ₂ δ, MeV
al γ, OLeu δ). (11) HPLC analysis: retention time 14.59 minutes

Example 9 (Production of TK11-3) This compound was produced according to the following production method.

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[0107]

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(A) Synthesis of N- (t-butoxycarbonyl) -L-pipecolic acid (hereinafter referred to as Boc-L-Pip-OH) L-pipecolic acid (2.00) was converted to dioxane: water = 2: 1 ( 50m
l), sodium carbonate (3.91 g), (Boc) ₂ under ice-cooling
After O (3.92 ml) was added, the mixture was stirred at room temperature for 2 hours. to this
1M NaOH (15 ml) and (Boc) ₂ O (3.92 ml) were added, and the mixture was further stirred at room temperature for 13 hours. After evaporating the solvent under reduced pressure, the residue was dissolved in water and washed with ether. Citric acid was added to the aqueous layer, and extracted with ethyl acetate. After concentrating the washed ethyl acetate, it was crystallized and washed with hexane to obtain Boc-L-Pip-OH (3.14 g).

(B) Synthesis of HCl.HL-Pip-OBzl Boc-L-Pip-OH (2.50 g) was dissolved in methanol aqueous solution (30 ml), cesium carbonate aqueous solution (2.0 g) was added, and the mixture was concentrated under reduced pressure. . DMF was added and azeotroped twice to remove water. DM the precipitate
Dissolved in F (30 ml), benzyl bromide (1.43 ml) was added under ice cooling, and the mixture was stirred for 10 minutes under ice cooling and further at room temperature for 20 hours. DMF
Was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with water and concentrated. 4.5N to the obtained residue (3.48g)
After adding HCl / dioxane (48.4 ml), the mixture was left at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure.
HCl.HL-Pip-OBzl (2.39 g) was obtained.

(C) Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-
Synthesis of L-Pip-OBzl Z-Ile- synthesized by steps (a) to (c) of Example 8
MeVal-MeAla-β-Ala-L-OLeu-OH (87.0mg), HCl ・ HL-Pi
p-OBzl (36.0 mg) and HOAt (20.0 mg) were dissolved in DMF (2 ml), and WSCD (32.3 μl) was added dropwise under ice-cooling.
The mixture was further stirred at room temperature for 16 hours. After distilling off DMF under reduced pressure,
The residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to obtain a residue. This is chloroform: methanol = 10
0: 1, silica gel column chromatography (silica gel 6.5 g, chloroform: methanol = 100:
Purify in 1), Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-L-Pip
-OBzl (110 mg) was obtained.

(D) H-Ile-MeVal-MeAla-β-Ala-L-OLeu-
Synthesis of L-Pip-OH Z-Ile-MeVal-MeAla-β-Ala-L-OLeu-L-Pip-OBzl (110m
g) was dissolved in methanol: acetic acid: water = 8: 2: 1 (30 ml), palladium black (2 spoonfuls of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. The palladium was filtered off, the mother liquor was concentrated under reduced pressure, dissolved in water and freeze-dried.
-Ile-MeVal-MeAla-β-Ala-L-OLeu-L-Pip-OH (75.4mg)
I got

(E) Synthesis of TK11-3 H-Ile-MeVal-MeAla-β-Ala-L-OLeu-L-Pip-OH (50.0 mg)
Was dissolved in dichloromethane (25 ml), and DIEA (83.6
μl) was added. Put this in a 25 ml syringe and add HATU (9
1.3 mg) was added dropwise over 2 hours to a dichloromethane solution (55 ml) containing the solution, and after stirring at room temperature for 16 hours, the solvent was distilled off under reduced pressure. The residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1).
K11-3 (13.6 mg) was obtained.

¹ H-NMR (270 MHz, CDCl ₃ ): δ 8.40 (d, 1
H, Ile NH), 7.82 (d, 1H, β-Ala NH), 5.08-4.99 (m, 3
H, Ile α, MeAla α, OLeu α), 4.89 (d, 1H, MeVal
α), 4.67 (d, 1H, Pip α), 4.38 (t, 2H, Pip ε), 4.1
2-4.02 (m, 1H, β-Ala β ₁ ), 3.23 (s, 3H, N-Me), 3.0
1-2.96 (m, 1H, β-Ala β ₂ ), 2.70 (s, 3H, N-Me), 2.6
6-2.52 (m, 6H, β-Ala α, Pip βγ), 2.38-2.29 (m, 1
H, MeVal β), 2.02-1.85 (m, 3H, Ile β, OLeu β),
1.47-1.22 (m, 5H, Ile γ 1, OLeu γ, Pip δ), 1.35 (d,
3H, MeAla β), 1.00-0.77 (m, 18H, Ile γ ₂ δ, MeVal
γ, OLeu δ) PD-MS measurement: m / z 608.4 (M + H) ⁺ molecular formula Calculated as C ₃₁ H ₅₃ O ₇ N ₅ : 607 HPLC analysis: retention time 27.31 minutes

Example 10 (Synthesis of TK14-6: In the production process described in Chemical formula 16, AA was L-methylnorleucine (MeNl
e)) (a) Synthesis of Boc-L-MeNle-OH Boc-L-Nle-OH (2.50 g) was dissolved in anhydrous THF (30 ml), and NaH (0.78 g) and CH ₃ I (5.38 ml) were cooled under ice-cooling. ) And add at room temperature 29
Stirred for hours. After THF was distilled off, the residue was dissolved in ethyl acetate, and washed with 10% citric acid and saturated saline. Ethyl acetate was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 400 g, chloroform: methanol = 9:
Purification according to 1) gave Boc-L-MeNle-OH (0.237 g).

(B) Synthesis of HCl.HL-MeNle-L-Pro-OBzl HCl.H-Pro-OBzl (0.228 g), Boc-L-MeNle-OH (0.220 g)
And HOAt (0.134g) in DMF (5ml)
(0.216 ml) was added dropwise. 1 hour under ice cooling, then 7 at room temperature
Stirred for hours. After distilling off DMF under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue (0.367 g)
I got After 4.5N HCl / dioxane (3.8 ml) was added to the residue, the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether, dried under reduced pressure, and used for HCl / HL-MeNle-L-Pro-OBzl.
(0.266 g) was obtained.

(C) H-Ile-MeVal-MeAla-β-Ala-L-MeNle
Synthesis of -Pro-OH Z-Ile-MeVal-MeAla-β-Ala-OH (0.150g), HCl ・ HL-MeN
le-Pro-OBzl (0.109 g) and HOAt (0.042 g) in DMF (2 m
l), and WSCD (67.7 μl) was added dropwise under ice-cooling. The mixture was stirred for 1 hour under ice cooling, and further for 16 hours at room temperature. After distilling off DMF under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 10 g, chloroform: methanol = 50: 1).
And purified. Methanol: acetic acid: water = 8: 2:
Dissolve in 1 (30ml) and palladium black (2 teaspoons spatula)
Cup), and hydrogen gas was blown for 1 hour with stirring at room temperature. After filtering the palladium, the mother liquor was concentrated under reduced pressure, dissolved in water and freeze-dried, and H-Ile-MeVal-MeAla-β-Ala-L-MeNle
-Pro-OH (0.156 g) was obtained.

(D) Synthesis of TK14-6 H-Ile-MeVal-MeAla-β-Ala-L-MeNle-Pro-OH (50.0 mg)
Was dissolved in dichloromethane (25 ml), and DIEA (83.6
μl) was added. Put this in a 25 ml syringe and add HATU (9
1.3 mg) was added dropwise over 2 hours to a dichloromethane solution (55 ml) containing the solution, followed by stirring at room temperature for 18 hours. After distilling off dichloromethane under reduced pressure, the residue was dissolved in ethyl acetate, and 10
% Citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate,
The extract was washed with a saturated saline solution and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1) to obtain TK14-6 (18.3 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ9.21 (d, 1H,
Ile NH), 7.94 (d, 1H, β-Ala NH), 5.11 (q, 1H, MeAla
α), 5.02-4.96 (2H, Ile α MeNle α), 4.46-4.41 (2H,
MeVal α Pro α), 4.02 (m, 2H, β-Ala β), 3.70-3.
67 (m, 1H, Pro δ ₁ ), 3.40 (m, 1H, Pro δ ₂ ), 3.21 (s, 3
H, N-Me), 3.09 (s, 3H, N-Me), 2.73 (s, 3H, N-Me), 2.5
8-2.54 (m, 2H, β-Ala α), 2.37-2.33 (m, 1H, Ile
β), 2.07-1.81 (m, 7H, MeVal β MeNle β Pro βγ),
1.35-1.14 (m, 2H, MeNle γ), 1.31 (d, 3H, MeAla β),
0.94-0.76 (m, 19H, Ile γδ MeVal γ MeNle δε) PD-MS measurement: 607.9 Calculated as molecular formula C ₃₁ H ₅₄ O ₆ N ₆ : 606.4 HPLC analysis: retention time 14.01 min

Example 11 (Production of TK15-5: AA is L-methylisoleucine in the production process described in Chemical formula 16) (a) Synthesis of HCl.HL-MeIle-L-Pro-OBzl HCl.H-Pro- OBzl (0.500 g), Boc-L-MeIle-OH (0.533 g)
And HOAt (0.310g) in DMF (5ml)
(0.498 ml) was added dropwise. 1 hour under ice-cooling, then 16 at room temperature
Stirred for hours. After distilling off DMF under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to obtain a residue. After adding 4.5N HCl / dioxane (9.2 ml) to this residue,
Left at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether, dried under reduced pressure, and HCl / HL-MeIle-L-Pro-OBzl (0.618 g)
I got

(B) H-Ile-MeVal-MeAla-β-Ala-L-MeIle
-Pro-OH Synthesis Z-Ile-MeVal-MeAla-β-Ala-OH (80.20g), HCl ・ HL-MeI
le-Pro-OBzl (0.145 g) and HOAt (56.1 mg) in DMF (2 m
l), and WSCD (90.2 μl) was added dropwise under ice cooling. The mixture was stirred for 1 hour under ice cooling and further for 5 hours at room temperature. After distilling off DMF under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 10 g, chloroform: methanol = 50: 50).
Purified according to 1). This is methanol: acetic acid: water = 8:
The mixture was dissolved in 2: 1 and palladium black was added. The mixture was stirred at room temperature for 1 hour while blowing hydrogen gas. Filter the palladium,
The solvent was distilled off under reduced pressure. After dissolving in water and freeze-drying, H-Ile-M
eVal-MeAla-β-Ala-L-MeIle-Pro-OH (0.201 g) was obtained.

(C) Synthesis of TK15-5 H-Ile-MeVal-MeAla-β-Ala-L-MeIle-Pro-OH (100 mg) was dissolved in dichloromethane (25 ml), and DIEA (167.4) was added thereto.
μl) was added. Put this in a 25 ml syringe and use HATU (1
83 mg) was added dropwise over 2 hours to a dichloromethane solution (135 ml), followed by stirring at room temperature for 18 hours. Dichloromethane was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1) to obtain TK15-5 (11.7 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ 8.84 (d, 1H,
Ile NH), 8.31 (d, 1H, β-Ala NH), 5.19 (q, 1H, MeAl
a α), 5.06-5.01 (2H, Ile α MeIle α), 4.73-4.69
(d, 2H, MeVal α Pro α), 3.90-3.80 (m, 2H, β-Ala
β ₁ Pro δ ₁ ), 3.28-3.21 (m, 2H, β-Ala β ₂ Pro
δ ₂ ), 3.18 (s, 3H, N-Me), 3.12 (s, 3H, N-Me), 2.77
(s, 3H, N-Me), 2.58-2.54 (m, 2H, Pro γ), 2.35-2.30
(m, 1H, MeIle β), 2.07-1.83 (m, 4H, Ile β MeVal
β β-Ala α), 1.77-1.67 (m, 2H, Pro β), 1.32 (d, 3
H, MeAla β), 1.19-1.04 (m, 4H, Ile γ ₁ MeIle γ ₁ ),
0.97-0.72 (m, 18H, Ile γ ₂ δ MeVal γ MeIle γ ₂ δ) PD-MS measurement: m / z 608.1 (M + H) ⁺ calculated value as molecular formula C ₃₁ H ₅₄ O ₆ N ₆ : 606.4 HPLC analysis: retention time 11.64 minutes

Example 12 (Production of TK17-5: In the production process described in Chemical formula 16, AA is L-methylphenylalanine) (a) Synthesis of HCl.HL-MePhe-L-Pro-OBzl HCl.H-Pro- OBzl (0.362g), Boc-L-MePhe-OH (0.400g)
And HOAt (0.214 g) in DMF (5 ml)
(0.344 ml) was added dropwise. 1 hour under ice-cooling, then 16 at room temperature
Stirred for hours. DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to obtain a residue. After 4.5N HCl / dioxane (6.4 ml) was added to the residue, the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure to obtain HCl · HL-MePhe-L-Pro-OBzl (0.545 g).

(B) H-Ile-MeVal-MeAla-β-Ala-L-MePhe
Synthesis of -Pro-OH Z-Ile-MeVal-MeAla-β-Ala-OH (0.150g), HCl ・ HL-MeP
He-Pro-OBzl (0.119 g) and HOAt (42.1 mg) were added to DMF (2 m
l), and WSCD (67.7 μl) was added dropwise under ice-cooling. The mixture was stirred for 1 hour under ice cooling and further for 18 hours at room temperature. DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was purified by silica gel column chromatography (silica gel 10 g, chloroform: methanol = 50: 1). Methanol: acetic acid: water = 8: 2: 1 (3
0 ml), palladium black (2 spoonfuls of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. The palladium was filtered, the mother liquor was concentrated under reduced pressure, dissolved in water and freeze-dried, and H-Ile-MeVal-MeAla-β-Ala-L-MePhe-Pro-OH
(0.152 g) was obtained.

(C) Synthesis of TK17-5 H-Ile-MeVal-MeAla-β-Ala-L-MePhe-Pro-OH (50 mg) was dissolved in dichloromethane (25 ml), and DIEA (79.3 μm) was added thereto.
l) was added. Put this in a 25 ml syringe and use HATU (86.
The solution was added dropwise to a dichloromethane solution (5 ml) containing 5 mg) over 2 hours, and then stirred at room temperature for 18 hours. Dichloromethane was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate.
The solvent was distilled off under reduced pressure, and the residue was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1) to obtain TK17-5 (13.3 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ9.13 (d, 1H,
Ile NH), 8.01 (d, 1H, β-Ala NH), 7.35-7.20 (m, 5H,
MePhe aromatic), 5.14 (q, 1H, MeAla α), 5.07 (d, 1
H, MeVal α), 4.93-4.90 (m, 1H, Ile α), 4.82 (t, 1
H, MePhe α), 4.06-3.96 (m, 1H, β-Ala β ₁ ), 3.81 (d,
1H, Pro α), 3.72-3.55 (2H, MePhe β ₁ Pro δ ₁ ), 3.4
5-3.24 (m, 1H, Pro δ ₂ ), 3.19 (s, 3H, N-Me), 3.14 (s,
3H, N-Me), 3.12-3.05 (m, 2H, β-Ala β ₂ MePhe
β ₂ ), 2.73 (s, 3H, N-Me), 2.57-2.55 (m, 2H, β-Ala
α), 2.40-2.31 (m, 1H, MeVal β), 2.29-2.15 (m, 2H,
Pro β), 2.03-1.95 (m, 1H, Ile β), 1.58-1.51 (m, 2
H, Pro γ), 1.35-1.25 (m, 2H, Ile γ ₁ ), 1.32 (d, 3H,
MeAla β), 0.94-0.90 (m, 6H, MeVal γ), 0.82-0.73
(m, 6H, Ile γ ₂ δ) PD-MS measurement: m / z 642.1 (M + H) ⁺ calculated as molecular formula C ₃₄ H ₅₁ O ₆ N ₆ : 639.5 HPLC analysis: retention time 12.31 min

Example 13 (Production of TK18-5: In the production process described in Chemical formula 16, AA is L-methylleucine) (a) Synthesis of HCl · HL-MeLeu-L-Pip-OBzl HCl · Pip-OBzl ( 0.219g), Boc-L-MeLeu-OH (0.200g),
Dissolve HOAt (0.122g) in DMF (2ml) and add WSCD (0.
196 ml) was added dropwise. The mixture was stirred for 1 hour under ice cooling, and further for 16 hours at room temperature. DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to obtain a residue. To this residue
After adding 4.5N HCl / dioxane (3.6 ml), the mixture was left at room temperature for 1 hour. Dioxane was distilled off under reduced pressure, and ether was added for crystallization. The residue was washed with ether and dried under reduced pressure to obtain HCl · HL-MeLeu-L-Pip-OBzl (0.302 g).

(B) H-Ile-MeVal-MeAla-β-Ala-L-MeLeu
Synthesis of -Pip-OH Z-Ile-MeVal-MeAla-β-Ala-OH (0.150g), HCl ・ HL-MeL
eu-Pip-OBzl (0.113 g) and HOAt (42.1 mg) were added to DMF (2 m
l), and WSCD (67.7 μl) was added dropwise under ice-cooling. The mixture was stirred for 1 hour under ice cooling and further for 18 hours at room temperature. DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure to obtain a residue. This was purified by silica gel column chromatography (silica gel 10 g, chloroform: methanol = 50: 1). This is methanol: acetic acid:
Water was dissolved in 8: 2: 1 (30 ml), palladium black (2 teaspoons of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. After filtering the palladium and concentrating the mother liquor under reduced pressure,
Dissolve in water and freeze-dry, then add H-Ile-MeVal-MeAla-β-Ala-L
-MeLeu-Pip-OH (0.121 g) was obtained.

(C) Synthesis of TK18-5 H-Ile-MeVal-MeAla-β-Ala-L-MeLeu-Pip-OH (50 mg) was dissolved in dichloromethane (25 ml), and DIEA (81.9 μl) was added thereto.
l) was added. Put this in a 25 ml syringe and use HATU (89.
3 mg) was added dropwise over 2 hours to a dichloromethane solution (55 ml) containing the solution, followed by stirring at room temperature for 18 hours. After distilling off dichloromethane under reduced pressure, the residue was dissolved in ethyl acetate, and 10
% Citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate,
The extract was washed with a saturated saline solution and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was separated by silica gel thin layer chromatography (chloroform: methanol = 19: 1) to obtain TK18-5 (24.7 mg).

¹ H-NMR (270 MHz, CDCl ₃ ): δ 9.07 (d, 1
H, Ile NH), 8.23 (d, 1H, β-Ala NH), 5.19-5.13 (2H,
Ile α MeAla α), 4.98 (d, 1H, MeVal α), 4.69-4.5
6 (m, 2H, Pip αε ₁ ), 4.90 (q, 1H, MeLeu α), 4.05-3.
76 (m, 1H, β-Ala β ₁ ), 3.90-3.81 (m, 1H, Pip ε ₂ ),
3.06 (m, 1H, β-Ala β ₂ ), 2.61-2.43 (m, 4H, β-Ala
α, Pip β), 2.39 (m, 2H, Ile β, MeVal β), 1.99-
1.82 (q, 2H, MeLeu β), 1.50-1.10 (m, 7H, Ile γ ₁ , Me
Leu γ, Pip γδ), 1.32 (d, 3H, MeAla β), 1.01-0.7
1 (m, 18H, Ile γ ₂ δ, MeVal γ, MeLeu δ) PD-MS measurement: m / z 621.8 (M + H) ⁺ molecular formula Calculated as C ₃₁ H ₅₄ O ₆ N ₆ : 620.4 HPLC Analysis: retention time 24.09 minutes

Example 14 (Production of TK16-22A and TK16-22B) This compound was produced according to the following production procedure.

[0132]

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[0133]

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[0134]

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[0135]

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Since the intermediate compound produced is unstable, TK
The reaction was carried out in the form of a mixture without purification until 16-9. (A) Synthesis of TK16-1 Boc-β-Ala-OH (5.00 g) was dissolved in ethyl acetate (80 ml), and triethylamine (3.67 ml) and isobutyl chloroformate (3.43 ml) were added dropwise under cooling at -20 ° C. After stirring for 10 minutes, the reaction solution was filtered. The filtrate was ice-cooled, a diazomethane ether solution (400 ml) was added, and the mixture was stirred under ice-cooling for 18 hours. Thereafter, argon gas was blown into the reaction solution for 1 hour to remove unreacted diazomethane. The reaction solution was washed with water and distilled off under reduced pressure to obtain a residue (5.32 g). Of which, 80m
g was separated by silica gel thin layer chromatography (benzene: ethyl acetate = 5: 1), and it was confirmed that TK16-1 was formed.

(B) Synthesis of TK16-7 and TK16-8 TK16-1 (3.3 g) was dissolved in ether (30 ml), and stirred for 2 minutes while blowing hydrogen bromide. After distilling ether,
It was azeotroped three times with chloroform. The residue was suspended in chloroform. (Boc) ₂ O (4.3 ml) and triethylamine (2.59 ml) were added dropwise thereto under ice cooling, and the mixture was stirred for 5 minutes under ice cooling and further for 1 hour at room temperature. The reaction solution was washed sequentially with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline, and dried over anhydrous magnesium sulfate. The solvent is distilled off under reduced pressure and TK
16-8 (6.52 g) was obtained.

(C) Synthesis of TK16-9 Sodium hydride (470 mg) was suspended in anhydrous THF (50 ml), and dibenzylmalonic acid (3.86 ml) was added at room temperature, followed by stirring for 1 hour. Thereafter, a THF solution (5 ml) of TK16-8 was added dropwise under ice cooling, and the mixture was stirred for 30 minutes under ice cooling and further for 1 hour at room temperature. THF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 400 g, hexane: ethyl acetate = 4:
Purification in 1) gave TK16-9 (1.61 g).

(D) Synthesis of TK16-12 TK16-9 (0.957 g) was dissolved in anhydrous DMSO (3 ml), sodium hydride (56.7 mg) was added, the mixture was stirred at room temperature for 1 hour, and isobutyl iodide (0.47 ml) was added. ) Was added dropwise, and the mixture was further stirred at room temperature for 22 hours. Ethyl acetate was added to the reaction solution, and the mixture was washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate, and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 80 g, hexane: ethyl acetate = 4: 1).
To obtain TK16-12 (1.19 g).

(E) Synthesis of TK16-13 TK16-12 (0.969 g) was dissolved in methanol (30 ml), palladium black (2 spoonfuls of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 2 hours. Filter the palladium,
After concentration of the mother liquor under reduced pressure, a residue was obtained. Add toluene (30m
l) was added and the mixture was stirred at 130 ° C for 2 hours. The toluene was distilled off under reduced pressure to obtain TK16-13 (0.474 g).

(F) Synthesis of TK16-14 TK16-13 (0.474 g), HCl.H-Pro-OBzl (0.379 g), HOBt
(233 mg) was dissolved in DMF (10 ml), and the WSCD (0.379 m
l, 1.73 mmol) was added dropwise, and the mixture was cooled for 1 hour under ice cooling, and
Stirred for hours. DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was purified by silica gel column chromatography (silica gel 65 g, chloroform: methanol = 100: 1) to give TK16-14 (0.44%).
4g) obtained.

(G) Synthesis of TK16-15 4.5N HCl / dioxane (3.66 ml) was added to TK16-14, and the mixture was allowed to stand at room temperature for 1 hour. Dioxane is distilled off under reduced pressure, and T
K16-15 was obtained (0.332 g).

(H) Synthesis of TK16-16 TK16-15 (332 mg), Boc-MeAla-OH (158 mg), HOAt (117
mg) was dissolved in DMF (4 ml), WSCD (188 μl) was added dropwise under ice cooling, and the mixture was stirred for 1 hour under ice cooling and further at room temperature for 15 hours. DMF
Was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated brine in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 80 g, chloroform: methanol = 10).
Purification was carried out according to 0: 1) to obtain TK16-16 (347 mg).

(I) Synthesis of TK16-17 4.5N HCl / dioxane (12.1 ml) was added to TK16-16 (347 mg).
Was added and left at room temperature for 1 hour. Dioxane was distilled off under reduced pressure to obtain TK16-17 (308 mg).

(J) Synthesis of TK16-18 TK16-17 (308 mg), Boc-MeVal-OH (140 mg) and HOAt (9
0.6 mg) in DMF (3 ml) and WSCD (146 μl) under ice-cooling
Was added dropwise, and the mixture was stirred for 1 hour under ice cooling and further for 6 hours at room temperature. DMF was distilled off under reduced pressure, and the residue was dissolved in ethyl acetate.
The extract was washed with citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was purified by silica gel column chromatography (silica gel 40 g, chloroform: methanol = 50: 1) to obtain TK16-18 (347 mg).

(K) Synthesis of TK16-19 To TK16-18 (347 mg) was added 4.5N HCl / dioxane (10.1 ml), and the mixture was allowed to stand at room temperature for 1 hour. Dioxane was distilled off under reduced pressure to obtain TK16-19 (312 mg).

(L) Synthesis of TK16-20 TK16-19 (312 mg), Z-Ile-OH (133 mg) and HOAt (74.9 m
g) was dissolved in DMF (3 ml), WSCD (120 μl) was added dropwise under ice cooling, and the mixture was stirred for 1 hour under ice cooling and further at room temperature for 18 hours. DMF was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The solvent was distilled off under reduced pressure, and the residue was subjected to silica gel column chromatography (silica gel 80 g, chloroform: methanol = 10).
Purification by 0: 1) gave TK16-20 (341 mg).

(M) Synthesis of TK16-21 TK16-20 (280 mg) was dissolved in methanol: acetic acid: water = 8: 2: 1 (3
0 ml), palladium black (2 spoonfuls of spatula) was added, and hydrogen gas was blown in at room temperature with stirring for 1 hour. The palladium was filtered off, the mother liquor was concentrated under reduced pressure, dissolved in water and freeze-dried to obtain TK16-21 (140 mg).

(N) Synthesis of TK16-22 TK16-21 (50.0 mg) was dissolved in dichloromethane (25 ml), DIEA (85.7 μl) was added thereto, and this was put into a 25 ml syringe, and HATU (93.5 mg) was added. ) Was added dropwise over 2 hours to a dichloromethane solution (57 ml) containing. Thereafter, the mixture was stirred at room temperature for 18 hours. The solvent was distilled off under reduced pressure, the residue was dissolved in ethyl acetate, washed with 10% citric acid, saturated saline, saturated aqueous sodium hydrogen carbonate and saturated saline in this order, and dried over anhydrous magnesium sulfate. The residue is subjected to silica gel thin layer chromatography (chloroform: methanol = 19: 1).
TK16-22-A (L form) 17.8mg, TK16-22-B (D
4.8 mg).

¹ H-NMR of TK16-22-A (270 MHz, CDCl ₃ ): δ
8.29 (d, 1H, Ile NH), 7.80 (m, 1H, β-Ala NH), 5.03
(d, 1H, MeVal α), 4.91 (q, 1H, MeAla α), 4.71 (t,
1H, Ile α), 4.34 (d, 1H, Pro α), 3.96-3.88 (m, 1H,
β-Ala β ₁ ), 3.66-3.50 (m, 2H, Pro δ), 3.25 (s, 3H,
N-Me), 3.17-3.02 (m, 1H, β-Ala β ₂ ), 2.85-2.75 (m,
3H, β-Ala α, cLeu H-2), 2.70 (s, 3H, N-Me), 2.50
(q, 2H, cLeu H-1), 2.41-2.28 (m, 1H, MeVal β), 2.1
3-1.83 (m, 5H, Ile β, Pro βγ), 1.66-1.51 (m, 2H,
_{Ile γ 1), 1.47-1.37 (m} , 1H, cLeu H-3 1), 1.34 (d, 3H,
MeAla β), 1.21-1.12 (m, 2H, cLeu H-3 2, H-4), 0.93-
0.80 (m, 18H, Ile γ ₂ δ, MeVal γ, cLeu H-5) SIMS: m / z 592.0 (M + M) ⁺ Calculated as C ₃₁ H ₅₃ O ₆ N ₅ : 591.4 HPLC analysis: retention time 13.80 minutes

¹ H-NMR of TK16-22-B (270 MHz, CDCl ₃ ) δ8.
04 (d, 1H, β-Ala NH), 7.42 (d, 1H, Ile NH), 5.13 (q,
1H, MeAla α), 5.00-4.90 (m, 2H, Ile α, MeVal
α), 4.69 (d, 1H, Pro α), 3.93-3.84 (m, 2H, β-Ala
β ₁ , Pro δ ₁ ), 3.65-3.48 (m, 1H, Pro δ ₂ ), 3.22 (s,
3H, N-Me), 3.16-3.02 (m, 3H, β-Ala β ₂ , cLeu H-1),
2.86-2.73 (m, 2H, β- Ala α 2, cLeu H-2), 2.71 (s, 3
H, N-Me), 2.57-2.42 (m, 1H, β-Ala α ₁ ), 2.38-2.25
(m, 1H, MeVal β), 2.03-1.96 (m, 5H, Ile β, Pro β
γ), 1.62-1.46 (m, 3H, Ile γ ₁ , cLeu H-4), 1.40-1.3
2 (m, 2H, cLeu H-3), 1.29 (d, 3H, MeAla β), 0.95-0.
85 (m, 18H, Ile γ ₂ δ, MeVal γ, cLeu H-5) SIMS: m / z 591.9 (M + M) ⁺ Calculated as C ₃₁ H ₅₃ O ₆ N ₅ : 591.4 HPLC analysis: retention time 22.43 minutes

Test Example 1 The effect of TK3-8 was described in the literature (Kumkawa M. and Tsunoo A .: Paras
ympathetic depression of vas deferens contraction
in the guinea-pig involves adenosine receptors. J.
Physiol., 407, 135-153, 1988, Tsunoo A. Kurokawa: M.
and TakahashiK .: Neurally evoked potentiation of t
onic contractions in the guinea-pigvas deferens in
volves adenosine receptors. J. Physiol., 433, 163-
181, 1991).

That is, the isolated myocardium of an SD rat (male, 200 to 350 g) was fixed in a 0.8-ml horizontal perfusion tank, and the contractile force was recorded isometrically. The perfusate has a composition consisting of Krebs solution equilibrated with 95% oxygen and 5% carbon dioxide, or a composition consisting of 140 mM sodium chloride, 5 mM potassium chloride, 2.6 mM calcium chloride, 1.3 mM magnesium chloride, 10 mM glucose, and 5 mM HEPES. The salt solution was used. The perfusate was maintained at 36-37 ° C and the flow rate was 3-4 ml / min. The test substance was dissolved in the perfusate and applied to the specimen. Papillary muscle specimens were electrically stimulated at 2 Hz through a platinum wire in the perfusion tank.

FIG. 2 shows the relationship between the TK3-8 concentration in the perfusion tank and the contractile force in the right atrium of the rat, and FIG.
Shown in The values are shown as ratios (relative values) to control values before drug application.

TK3-8 dose-dependently enhanced the contraction of the right atrial muscle by automatic activity in the concentration range of 2 μM to 200 μM (FIG. 2). Further, as shown in FIG. 3, the contraction interval was hardly affected or tended to increase. TK3-8 also enhanced contraction of right ventricular papillary muscle by electrical stimulation in a dose-dependent manner, as in right atrial muscle.

Test Example 2 Cyclic peptides of the present invention other than TK3-8 (TK2-6-A, TK11-3, TK
14-6, TK16-22-A, TK17-5 and TK18-5) were measured in accordance with the method described in Test Example 1 to increase the contractile force on the isolated rat right atrial muscle (all drug concentrations were 20 μM). .
Table 2 shows the results. The values in Table 2 were expressed as ratios to control values before drug administration.

[0157]

[Table 2]

Test Example 3 In order to investigate the inhibitory effect on abnormal contraction induced by digitalis, an experiment was conducted in which 1 μM of digoxin was administered to the isolated right atrium of a guinea pig (male, 100 to 200 g) to induce abnormal contraction. . Then, as shown in FIG. 4A, the amplitude and rhythm of the right atrial automatic contraction were both significantly irregular. Thus, when TK3-8 was administered at 20 μM, the amplitude and rhythm became constant as shown in FIG. 4B, abnormal contraction by digoxin was suppressed, and sinus rhythm was restored. in this way,
TK3-8 was found to have an effect of suppressing abnormal contraction induced by digitalis.

Test Example 4 Whether or not the compound of the present invention is effective was examined not only in the isolated organ but also in the living body. The left ventricular pressure of male SD rats was measured by cardiac catheterization under thiopental anesthesia, and the effect of TK3-8 on the time derivative was examined. TK3-8 is 10% s
Dissolve in ulfolane and bolus from right femoral vein at time 0
Was administered. FIG. 5 shows the results.

As shown in FIG. 5, TK3-8 (1 mg / kg)
The administration caused a sustained increase in the differential value of the left ventricular pressure with time, which revealed that the cyclic peptide of the present invention also has a cardiotonic effect in a living body.

[Brief description of the drawings]

FIG. 1 is an infrared absorption spectrum of TK3-8.

FIG. 2 is a graph showing the relationship between TK3-8 concentration in a perfusion tank and contractile force in the right atrium of a rat.

FIG. 3 is a graph showing the relationship between TK3-8 concentration in a perfusion tank and contraction interval time in the right atrium of a rat.

FIG. 4 is a graph showing the inhibitory effect of TK3-8 on abnormal contraction induced by digitalis.

FIG. 5 is a graph showing the effect of TK3-8 on the sustained increase in the time derivative of left ventricular pressure in rats.

 ──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor Tetsuo Shiba 4-1-2, Ina, Minoh-shi, Osaka Within the Protein Research Promotion Foundation

Claims (2)

[Claims] 1. A compound of the general formula (1) (Wherein, R ¹ represents —CH ₂ — or —CH ₂ CH ₂ —, R ² represents a hydrogen atom or an alkyl group or a phenyl group or a benzyl group, and R ³ represents —NH—
Or —N (CH ₃ ) — or —CH ₂ — or an oxygen atom, respectively). 2. A cardiotonic agent comprising the cyclic peptide according to claim 1 as an active ingredient.