CS228546B2 - Method of preparing pentapeptides - Google Patents

Description

The invention relates to a process for the preparation of pentapeptides of the general formula I,

A — B — Gly — D — E (I) in which

A is a radical of formula (II),

(II) where

Z ¹ represents a hydrogen atom, a methyl group or an amidino group,

R 1 represents a hydrogen atom or a C 1 -C 4 alkyl group;

R2 is hydrogen or

R 1 and R 2 together form an ethylene bridge and

R3 represents a hydroxy or alkoxy group having 1 to 4 carbon atoms attached to the 3, 4 or 5 position,

B represents a residue (Dj-isomer of a natural N-amino acid,

D is a radical of formula III,

Z ²

AND

R ₄ CH 2

I - N - CH - CO - (IIP in which

R ₄ represents a hydrogen atom or a (C 1 -C 4) alkyl group;

Z ² is phenyl optionally mono- or disubstituted with halogen, nitro, C 1 -C 4 alkyl and / or C 1 -C 4 alkoxy; and

E is a radical of formula (IV),

R 5 R 6 I-N-CH-CH 2 OH (IV) wherein

R 5 represents a hydrogen atom or a C 1 -C 4 alkyl group;

R e represents a hydrogen atom, a substituent corresponding to the residue attached to the α-carbon of the natural α-amino acid, a residue of the formula - (CH ₂ j _m - CH ₂ OH where m is a number of 1 to 6 or jp — R7 where

R 7 represents an alkyl group of β 1 to 5 carbon atoms and, for example, independently of each other, represents a number of 0, 1 or 2, provided that if n has a lap of 0, then R 2 is not a methyl group, (L 1 -configuration, residue E has the (Dj- or (L 1 -configuration and residue B) has (D 1 -coinfiguration, with the restriction that if Z ¹ is a hydrogen atom or a methyl group, then R ₄ represents an alkyl group having 3 to 4 carbon atoms, as well as their physiologically hydrolyzable, physiologically acceptable esters and salts and complexes of these pentapeptides and esters.

When describing the polypeptides of the invention as pentapeptides, it should be understood that the amino alcohol moiety of E is considered to be a zia derivative of an amino acid.

The term "amidinoskiupina" refers to a group of the formula

NH

NH — C—

When Z ¹ is an amido group, R 4 is a hydrogen atom or a C 1 -C 4 alkyl group. When Z ¹ represents a hydrogen atom or a methyl group, then R 4 represents a C 3 -C 4 alkyl group.

Compounds of the formula I according to the invention in which Z ¹ represents a hydrogen atom or a methyl group belonging to the class of compounds described in DOS No. 2 702 711 and the equivalent GB patent No. 1 578 873. All the compounds produced by the process according to the invention are novel.

As mentioned above, the compounds produced by the process of the invention exhibit advantageous biological properties, in particular luteinizing hormone secretion inhibiting activity, and can be used as pharmaceutically active substances.

The residue in the meaning of the symbols E may have either (L) - or (Dj-configuration). In the text of the present invention and in the definition of the invention the terms "(L) - β-aminoalcohol residue" and "residue (Dj-β-aminoalcohol)" "Means those residues in which the relative configuration corresponds to that of the corresponding (L 1 -a-amino acid or (L 1 -a-amino acid) [wherein the carboxyl group in that residue (L 1 - or (L 1 -a amino acid is replaced in each case) It is to be understood that these radicals may carry an alkyl substituent of 1 to 4 carbon atoms on the nitrogen atom in the .beta.-position.

In the pentapeptides of formula I, the following meanings of the individual symbols or combinations thereof are preferred:

lj Both Rt and Rz are hydrogen,

2] R 3 represents hydroxy. Most preferably, R 3 occupies position 4 and is especially 4-hydroxy,

3] B represents the residue of the (D) -isomer of the natural non-aromatic α-amino acid. Most preferably, B is - (D) -Ala-,

4] R ₄ represents a C 1 -C 4 alkyl group, for example a methyl group. Where R4 represents an alkyl group having 3-4 carbon atoms (and in case is when Z ¹ is hydrogen or methyl and where it constitutes the Z ¹ amidimoskupinu) group this is preferably n-propyl, isopropyl or n-butyl, especially n-propyl,

5] Z ² is preferably a phenyl group, optionally mono- or substituted with one or two of fluoro, chloro and nitro, especially fluoro. When Z ^{2 has a} substituted phenyl group, it is preferably an inosubstituted phenyl group. Most preferably, Z ² is phenyl or m- or p-fluorophenyl group, especially a phenyl or p-fluorophenyl,

6.1} R 5 preferably represents a hydrogen atom,

6.2) As a substituent attached to the .alpha.-carbon of the natural .alpha.-amino acid, R @ 6 preferably represents a group of the formula

CH3 — S — CH2 — CH2—, HO — CH2—,

CH 3 —CH (OH) - or

H2NCO — CHa — CH2—,

6.3] When R 6 has a meaning other than that mentioned in 6.2 j, it preferably represents the remainder of the formula

CH3-SO-CH2-CH2- or

CH3 — SO2 — CH2 — CHa—,

6.4} Most preferably R6 is a radical of formula

CH3-SO-CH2-CH2-a

6.5) The radical E, including the preferred radicals of the symbol E as defined in paragraphs 6.1) to 6.4) above, preferably has the (L) -configuration (as defined above). Most preferably, E is the -Met (O) -ol residue, i.e. the (L) residue of methionlnol sulfoxide,

The hydroxyl groups in the pentapeptides of the formula I, in particular the hydroxy group of the β-amino alcohol radical E, can be esterified to give the corresponding physiologically hydrolyzable, physiologically acceptable esters, i.e. esters which give a physiologically acceptable acid by hydrolysis.

Suitable esters include, for example, esters of mono- and dicarboxylic acids, in particular esters of carboxylic acids having 2 to 5 carbon atoms, such as acetates, as well as aromatic carboxylic acids, such as benzoates.

The pentapeptides of formula I and esters thereof may be in the form of acid addition salts or complexes. The addition of salts with acids can also be formed, for example, with organic acids, polymeric acids and inorganic acids. Such acid addition salts include, for example, hydrochlorides and acetates. The term "complexes" refers to compounds of a known type prepared from the pentapeptide of formula I and their esters by addition of inorganic compounds, for example inorganic salts or hydroxides such as salts of calcium, magnesium, aluminum, cobalt and especially zinc, or addition of polymeric organic substances.

One group of compounds of the invention are the pentapeptides of formula I wherein:

Z ¹ is an amidino group,

B represents the remainder - (Dj-Alla-,

Z ² is phenyl. optionally mono- or di-substituted with halogen, nitro, C 1 -C 4 alkyl or C 1 -C 4 alkoxy; and

R 1, R 2, R 3, R a and E are as defined in formula I, and their acid addition salts and complexes.

A second group of compounds of the invention are the pentapeptides of formula I wherein:

Z ¹ represents a hydrogen atom or a methyl group,

R 1 and R 2 either represent hydrogen atoms or together form an ethylene bridge,

R 5 represents a hydroxyl group at the 4-position,

B is - (D) -Ala-,

R 4 is C 3 -C 4 alkyl (especially n-propyl, n-butyl and isobutyl),

Z ² represents a phenyl group optionally mono- or disubstituted by one or two substituents selected from the group consisting of halogen atoms, nitroskiupine of a C 1 -C 4 alkyl group and a C 1 -C 4 alkoxy group, and

Ξ represents the radical -Met (O) -ol, and their acid addition salts and complexes.

The present invention provides a process for the preparation of the compounds described above. The compounds may be prepared by amidic linkage of two peptide units, each containing at least one amino acid or amimoalcohol residue, the peptide units being of a chiarakterurakteir to form a pentapeptide or ester thereof having the sequence shown in Formula I, and the thus obtained pentapeptide or ester thereof is isolated in free form or in the form of an acid addition salt or complex.

The above reaction can be carried out analogously to the procedure described below in the example. If the preparation of the starting materials is not specifically described, these compounds are either known or can be prepared and purified by methods known per se.

The pentapeptides and esters of the invention, as well as their physiologically acceptable acid addition salts and complexes, exhibit valuable pharmacological properties as evidenced by animal testing. In particular, these compounds inhibit luteinizing hormone secretion and have central nervous system depressant activity, as exemplified by the following tests.

Inhibitory activity on luteinizone hormone secretion; inhibition of ovulation in rats

Adult female rats weighing 200-250 g, with a regular four-day estrous cycle, are maintained under standard conditions [14 hours of light daily from 04.00 to 18.00, 24 ° C and 55 to 60% relative humidity], while not restricting food and water intake. The test compound is administered subcutaneously in the preest stage, with the test doses administered only at 13.00 h and then again at 16.00 h. 20 hours after the first dose, the rats are sacrificed, their fallopian tubes are removed and the total number is determined using a microscope. eggs in both fallopian tubes. The evaluation is carried out by judging the inhibition of ovulation only when the presence of the desired egg cannot be detected. Each dose was tested in five rats.

2. Depressive activity on the central nervous system

This assay is performed by monitoring changes in the spontaneous behavior of mice and rats in accordance with the principles described by J. H. Nodid et al. in Animal and Clinical Pharmacological Techniques in Drug Evaluation, Chicago, 1964, and in "Psychopharmacologia", 13, 222-257 (Berlin, 1968, p.

The pentapeptides, esters, salts and complexes described above can be used accordingly

(a) for the treatment of patients with disorders whose etiology is related to or modulated by luteinizing hormone secretion, or whose etiology includes physiological regulation of luteinizing hormone secretion, for example in the treatment of prostate hypertrophy or menopausal syndrome, in particular post-menopausal hot flashes studies described by Tataryn et al. ("Thermo-regulatory Mechanism and Their Therapeutic Implacatioms", 4th International Symposium on Pharmacology and Thermoregulation, Oxford, 1979; publ. Karger, Basel 1980, pp. 202-207) and Casper et al. ["Science", 205, 823-825 (1979)]

b] as psychopharmaceuticals, for example in the treatment of psychotic conditions.

For both of these uses a) and b], the recommended dose is in the range of 0.1 to 10 mg of peintapeptide or its ester, the total dose being administered in several divided doses two to four times daily in unit dosage form or in prolonged form. Thus, suitable unit dosage forms, for example, for oral administration, contain from 0.025 to 5.0 mg of the pentapeptide or ester mentioned above, or an equivalent amount of a pharmaceutically acceptable salt or complex, together with a solid or liquid pharmaceutical diluent or carrier. These compositions may take the form of, for example, solutions or tablets.

The invention further describes

(1) a method of inhibiting luteinizing hormone secretion, in particular a method of treating prostate hypertrophy or menopausal syndrome, in particular post-menopausal hot flashes, and a method of treating psychotic disorders in patients in need of such treatment, comprising administering to said patient an effective amount of pentapeptide or a physiologically hydrolyzable, physiologically acceptable ester, or a physiologically acceptable acid addition salt or complex of such a pentapeptide or ester, and

2} pentapeptides, esters, acid addition salts and complexes mentioned in paragraph 1j above as pharmaceuticals, ie for use in the treatment of human or animal, in particular for use as described in paragraph 1 above

1) a

3) pharmaceutical compositions containing the pentapeptides, esters, acid addition salts or complexes mentioned in paragraph 1j above as active ingredients.

The following common abbreviations are used in the specification, in the examples, and in the definition of the invention:

Met-ol = (L) -methioninol residue of formula CH3SCH2CH2CH (CH2OH) NH-,

Met (O) -ol = residue of (L 1 -methionine sulfoxide of formula)

CH 3 -S (O) -CH 2 CH 2 CH (CHaOH) -NH-,

MeTyr = -N-methyl-Tyr-

MeTyr (Butj - N-methyl-O-tert-butyl-Tyr-,

MePhe = N-methyl-Phe-, (N-propyl-J-Phe = N-n-propyl-Phe, (N-isobutyl JPhe = N-isobutyl-Phe-, (N-butyl) Phe = -N-n-butyl-Phe-,

N-propyl is Phe (p-F) = -N-n-propyl-p-fluoro-Phe,

Boc = t-butoxycarbonyl,

Z - Benzyloxycarbonyl.

In accordance with the standard nomenclature, all amino acid residues denoted by the three-letter abbreviations are considered to be L-configurations unless otherwise specified. Example 1

N ^and -amidino-Tyr- (Dj-Ala-Gly- (N-propyl Phe-Met-ol hydrochloride)

To a solution of 1.04 g of N ^K -amidino-Tyr- (D) -Ala-NH-NH2-hydrochloride in 20 ml of dimethylformamide, at -18 DEG C., is added first 1.3 ml of 8N hydrogen chloride in dioxane and then 0, 35 ml of t-butyl nitrite. After standing for 10 minutes at -8 ° C, the reaction mixture is neutralized with 2.3 ml of N-ethyl-diisopropylamine and mixed with the cooled solution.

1.5 g of H-Gly- (N-propyl) -Phe-Met-ol hydrochloride and 0.62 ml of N-ethyl-diisopropylamine in 15 ml of dimethylformamide, after one hour reaction at -15 ° C followed by three hours reaction at -15 ° C. The solvent was evaporated in vacuo at 10 ° C, the residue was taken up in water and the solution was washed several times with methylene chloride, the aqueous phase was evaporated to a volume of ca. after evaporating the eluate to dryness and precipitation from methanol-ethyl ether gave the title compound, with an optical rotation [a] _D ²⁰ = -41.0 ⁰⁴ (c = 1 in dimethylformiamidu extending at 90 ° C.

The starting material used in the above reaction was prepared as follows:

a) H- (N-piropropyl JPhe-NH-NH-Z-hydrochloride)

To a solution of 14.0 g of H-Phe-NH-NH-Z-hydrorld in 120 ml of methanol at 20 degrees Celsius was first added 10.0 g of sodium cyanoborohydride and, after 5 minutes, 3.2 ml of propionaldehyde. After reaction lasting

The solvent was removed in vacuo for 2.5 hours and the residue was added. 50 ml of water, the mixture was made strongly alkaline (pH ca. 12) by addition of 10N sodium hydroxide and the product was extracted with methylene chloride. The extract was washed thoroughly with water, dried over sodium sulfate and treated with excess hydrogen chloride in ethyl ether. Crystallization is then completed by addition of additional ether. The title compound is obtained, having an optical rotation of [α] D ²⁰ = + 42.5 ° (c = 1 in dimethylformamide).

b) Boc-Gly- (N-propyl) Phe-NH-NH-Z

To a solution of Boc-Gly-OH (16.8 g) in tetrahydrofuran (100 mL) was added pivaloyl chloride (11.8 mL) at -20 ° C, and the mixture was stirred at -15 ° C for 15 min. g of H- (N-propl) Phe-NH-NH-Z and 5.5 ml of N-ethyl-diisopiropylamine in 50 ml of tetrahydrofuran, the reaction mixture is stirred at -15 ° C for 1 hour, then at 0 ° 1 hour. C and finally 20 hours at 10 ° C. After addition of 150 ml of 1 N potassium hydrogen carbonate, the product is extracted with ethyl acetate, the extract is washed with dilute potassium hydrogen carbonate solution and potassium hydrogen sulphate and water, dried over sodium sulfate and the solvent is evaporated off under vacuum. The title compound is purified by chromatography using silica gel and a mixture of methyl chloride and methanol as eluent.

c) Boc-Gly -, (N-propyl) Phe-NH-NH 2

21.1 g of Boc-Gly- (N-propyl) Phe-NH-NH-Z in 100 ml of methanol are hydrogenated in the presence of a palladium catalyst for 1 hour under a hydrogen pressure of 0.2 to 0.3 MPa. The solvent was evaporated in vacuo and the amorphous residue was dried over phosphorus pentoxide and potassium hydroxide in vacuo to give the title compound, [.alpha.] D @ ²⁰ = -39.4 DEG (c = 1 in dimethylformamide j).

dj Boc-Gly- [N-propyl] Met-ol

To a solution of 7.8 g of Boc-Gly- (N-propyl) Phe-NH-NH2 in 50 ml of dimethylformamide at -18 ° C is added first with stirring

8.6 ml of 8 N hydrogen chloride in dioxane and then

2.1 ml of tert-butyl nitrite. After standing at -18 DEG C. for 10 minutes, the mixture is neutralized by the addition of 12.0 ml of N-ethyl-diisopropylamine and treated with a pre-cooled solution of 5.4 g of methioninol (CH3SCH2CH2CH (NH3 / CH3OH) in 30 ml of DMF). At 10 ° C, the reaction mixture was diluted with ethyl acetate, washed with dilute potassium hydrogen carbonate solution and potassium hydrogen sulfate, and water, dried over sodium sulfate, the solvent was evaporated in vacuo and the residue purified by silica gel chromatography eluting with methylene chloride / methanol. to give the title compound.

e] H-Gly- (N-propyl) Phe-Met-ol-hydr

To a solution of 9.9 g of Boc-Gly- (N-propyl) Phe-Metol in 20 mL of ethyl ether at 0 ° C was added 100 mL of 5 N hydrogen chloride in ethyl ether. After an additional hour at 0 ° C, the solvent was evaporated in vacuo and the residue was recrystallized from a mixture of ethyl acetate, methanol and ethyl ether to give the title compound, [α] _D ²⁰ = -72.5 ° (c) = 1 in dimethylformamide). The product decomposes at 60 ° C.

f) Boc-Tyr- (D) -Ala-NH-NH-Z

To a solution of 14.1 g of Boc-Tyr-OH and 5.6 ml of N-methylmorpholine in 100 ml of tetrahydrofuran at -20 ° C was added 6.6 ml of isobutyl chloroformate. After reaction at -20 ° C for 5 minutes, a solution of 13.7 g of H- (D) -Ala-NH-NH-Z-hydrochloride and 5.0 ml of N-methylmorpholimus in 50 ml of dimethylformamide is added. The mixture was allowed to react at -15 ° C for 2.5 hours, then 100 ml of 1 N potassium bicarbonate was added thereto, the reaction mixture was diluted with ethyl acetate, washed with dilute potassium bicarbonate solution and potassium hydrogen sulfate and water and dried over sodium sulfate. and the solvent was evaporated in vacuo. Recrystallization of the residue from a mixture of ethyl acetate, hexane and ethyl ether gave the title compound.

g) H-Tyr- (D) -Ala-NH-NH-Z-hydrochloride

To a solution of 25.0 g of Boc-Tyr- (D) -Ala-NH-NH-Z in 50 ml of ethyl acetate at 0 ° C was added 100 ml of a 5 N solution of hydrogen chloride in ether. After 1.5 hours, crystallization was completed by addition of additional ethyl ether. The title compound is isolated by filtration. The product has an optical rotation of [α] _D ²⁰ = + 32.8 ° (c = 1 in dimethylformamide) and decomposes at 200 ° C.

h) N-amido-Tyr- (D) -Ala-NH-NH-Z-hydrochloride

To a solution of 8.8 g of H-Tyr- (D) -Ala-NH-NH-Z-hydrochloride and 2.8 ml of triethylamine in 50 ml of water.

228 ml of dimethylformamide were added with 12.1 g of 1-amidino-3,5-dimethylpyrrole nitrate. The mixture was stirred at 50-60 ° C for 4 days and then 3.0 mL of triethylamine was added. The solvent was evaporated in vacuo, the residue was taken up in water and the solution was extracted several times with methylene chloride. The aqueous phase is concentrated to about 400 ml, the concentrate is adsorbed onto a polystyrene adsorption resin, and the title compound is eluted from the adsorbent as a nitrate using a mixture of methanol and water as eluent. The eluate was evaporated to dryness, the residue was re-dried in vacuo, dissolved in ethyl acetate in the presence of a small amount of ethanol, and the title compound precipitated as hydrochloride by addition of ethyl ether and an excess of 5N hydrogen chloride in ethyl acetate.

The product has an optical rotation of [α] _D ²⁰ = + 23.3 ^P (c = 1 in dimethylformamide) and decomposes at 50 ° C.

i) N-amidino-Tyr- (D) -Ala-NH-NHa-hydrochloride

1.6 g of N ' -amido-Tyr- (D) -Ala-NH-NH-Z-hydrochloride in 20 ml of methanol are hydrogenated under a hydrogen pressure of 0.2 to 0.3 MPa in the presence of a palladium catalyst. the catalyst is filtered off, the solvent is evaporated in vacuo and the residue is recrystallized from methanol / ethyl ether to give the title compound, [.alpha.] D @ ²⁰ = +13.2 DEG (c = 1 in dimethylformamide), decomposing at 80 ° C.

Example 2

The following compounds were prepared in an analogous manner to Example 1:

2.1. H-MeTyr- (D 1 -Ala-Gly- (N-isohutyl) Pihe-Metf G 1 -ol-trifluoroacetate, optical rotation [α] D ²⁰ = -13.7 ° (c = 1 in 95% acetic acid) ,

2.2. H-MeTyr- (D) -Ala-Gly- (N-butyl) Phe-Met (O-ol-trifluoroacetate with optical rotation [a] p ^2Q - (c = 1 in 95% acetic acid),

2.3. H-MeTyr- (Dj-Ala-Gly- (N-propyl) Phe (p-Fj-Met (O) -ol-trifluoroacetate with optical rotation [α] _D ²⁰ = -9.0 ° (c - 1) in 95% acetic acid,

2.4. N '-amidino-Tyr- (D) -Ala-Gly-MePhe-Metf0j-ol acetate with optical rotation [ _.alpha. ] _D @ ²⁰ - -50.8 DEG (c = 0.7 in dimethylformamide);

2.5 N '-amidino-Tyr- (D) -Ala-Gly-MePihe (β-FJ-Met (ω-ol hydrochloride, optical rotation [α] d ²⁰ - 29.3 ° (c = 1 in 95%) acetic acid).

2.6. H-MeTy, .alpha .- (D) -Ala-Gly- (N-propyl) Phe-Met (O) -ol trifluoroacetate, [ _.alpha. ] _D @ 20 ₌ -19.3 DEG (c = 1 in 95) % acetic acid) and

^And 2.7 N-amidino-Tyr- (D) - Ala-Gly- (N-propyl) Phe-Met (O) -ol hydrochloride, optical rotation [a] ²⁰ _D - 33.6 ° (c = 1 in 95 % acetic acid).

Claims (3)

A process for the preparation of pentapeptides of the general formula I, A - B - Gly - D - E (1j in which A is a radical of formula (II), (C1-C4) group and R2 represents a hydrogen atom; R 1 and R 2 together form an ethylene bridge and R3 represents a hydroxy or alkoxy group having 1 to 4 carbon atoms attached to the 3, 4 or 5 position, B represents the residue of the (D) -isomer of the natural α-amino acid, D is a radical of formula III, Z ² AND R ₄ CH 2 I - N - CH - CO - (III) in which R a is hydrogen or C 1 -C 4 alkyl and Z ² represents a phenyl group, optionally Z ¹ represents a hydrogen atom, a methyl group or an amidino group, R1 is hydrogen or alkyl 228548 is mono- or di-substituted with halogen, nitro, C1-C4alkyl and / or C1-C4alkoxy, E is a radical of formula (IV), Rs Re N - CH - CH 2 OH (IV) wherein R 5 represents a hydrogen atom or a C 1 -C 4 alkyl group; R e represents a hydrogen atom, a substituent corresponding to the residue attached to the α-carbon of the natural α-amino acid, a radical of the formula - (CH 2) _m -CH 2, OH where m is a number 1 to 6, or a radical of the formula - (CH ₂ ) _n - CH ₂ -S (O) _p -R Z wherein R 7 is C 1 -C 5 alkyl and, independently of each other, is a number of 0, 1 or 2, provided that if n is 1 and p is 0, then R 7 is not methyl, and residues A and D have (Lj-configuration, the residue E has the (D) - or (L) -configuration and the residue B has the (Dj-configuration with the proviso that when Z ¹ is hydrogen or a methyl group, R4 represents an alkyl group And 3 to 4 carbon atoms, as well as their physiologically hydrolyzable, physiologically acceptable esters and salts, and complexes of these pentapeptides and esters, characterized in that two peptide units, each containing at least one amino acid or aminoalcohol residue, are amidically linked. the peptide units are of such a nature that a pentapeptide or ester thereof having the sequence shown in formula I is formed, and the pentapeptide or ester thereof thus formed is isolated in free form or as an acid addition salt or complex. 2. A process according to claim 1, wherein the corresponding starting materials are used to form compounds of formula (I) in which: B is a residue of D-Ala, Z ¹ is amidino a R1, R2, R3, D and E have the meanings given in 1. 3. A process according to claim 1, wherein the corresponding starting materials are used to form compounds of formula (I) in which: Z ¹ represents a hydrogen atom or a methyl group, R1 and R2 are either hydrogen or together are an ethylene bridge, R 3 is hydroxy at the 4-position, B is a residue of D-Ala, R 4 represents a C 3 -C 4 alkyl group, E represents a residue of Met (O) -ol- a Z ² is as in point 1.