EP0502874A1

EP0502874A1 - Novel peptides derived from neuropeptide y

Info

Publication number: EP0502874A1
Application number: EP19900917028
Authority: EP
Inventors: Johann-Christian Zechel; Sabine Schult; Liliane Unger
Original assignee: BASF SE
Current assignee: BASF SE
Priority date: 1989-12-01
Filing date: 1990-11-22
Publication date: 1992-09-16
Also published as: WO1991008223A1; DE3939797A1

Abstract

Il est décrit des peptides ayant la formule Z-Ak-Bl-Cm-Dn-Eo-Fp-Gq-H-Thr-I-Gln-J-K-NH2, dans laquelle A - K, Z et k - q ont la notation donnée dans la description, ainsi que leur procédé de fabrication. Ces peptides conviennent pour combattre des maladies.Peptides having the formula Z-Ak-Bl-Cm-Dn-Eo-Fp-Gq-H-Thr-I-Gln-JK-NH2 are described, in which A - K, Z and k - q have the notation given in the description, as well as their manufacturing process. These peptides are suitable for fighting diseases.

Description

New peptides derived from neuropeptide Y

The invention relates to new peptides derived from neuropeptide Y (NPY), their preparation and their use as medicaments.

NPY was isolated from pig brain in 1982 (Nature 296 (1982) 659) and has now been detected in a number of mammalian species. Human NPY has the following sequence:

H-Tyr-Pro-Ser-Lys-Pro-Asp-Ala-Pro-Gly-Glu-Asp-Ala-Pro-Ala-Glu-Asp-Met-Ala

Arg-Tyr-Tyr-Ser-Ala-Leu-Arg-His-Tyr-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-

NH ₂

The peptide, which is widely used in the peripheral and central nervous system, is one of the most potent known vasoconstrictors. It also increases the blood pressure-increasing effects of norepinephrine and is able to cause cardiac and cerebral vasospasm. All of these observations indicate that NPY is significantly involved in the neuronal regulation of blood pressure.

Peptides have now been found with which NPY can be antagonized. The invention relates to peptides of the formula

ZA _k -B ₁ -C _m -D _n -E _o -F _p -G _q -H-Thr-I-Gln-JK-NH ₂ wherein

A Phe or Tyr,

B Pro, Ser, Abu, Gly, Ala, Cys, Pro-Ser, Cys-Ser, Abu-Ser, Gly-Ser, Ala- Ser or Pro-Ala,

C an amino acid with a basic side chain,

D Pro or Cys,

E the grouping -NH- (CH ₂ ) t_CO_ (with t meaning an integer from 0 to 10), F Cys, Abu, Gly or Ala,

G Ile-Asn, Leu-Asn, Val-Asn or Asn,

H is a dipeptide residue composed of lipophilic amino acids,

I an amino acid with a basic side chain, J an amino acid with a basic side chain,

K Phe or Tyr and

Z represents a hydrogen atom, an amino protecting group, a benzyl radical, a C _2-10 acyl radical or a C _1-5 alkyl radical, k, l, m, n, o, p and q denote the numbers 0 or 1, but k + 1 + m + n + o + p + q> l and two possibly present Cys residues can be connected to one another via a disulfide bridge, as well as their salts with physiologically compatible acids.

In the new peptides the partially structurally modified N- and C-terminal fragments of the NPY are connected to each other via a spacer (E) which replaces the central sequence section of the NPY. The lengths of the N- and C-terminal ends and the spacer are variable.

Arg, hArg, His, Lys and Orn come into consideration as basic amino acids for C, I and J. Ile, Leu, Val, Phe, Trp, Nie, Pro are to be mentioned as lipophilic amino acids (see H). Suitable protecting groups (see Z) are the following: t-butyloxycarbonyl, benzyloxycarbonyl, fluorenylmethyloxycarbonyl.

In particular, the following can be mentioned as physiologically acceptable acids:

Hydrochloric acid, citric acid, tartaric acid, lactic acid, phosphoric acid, methanesulfonic acid, acetic acid, formic acid, maleic acid, fumaric acid, malic acid, succinic acid, malonic acid, sulfuric acid, L-glutamic acid,

L-aspartic acid, pyruvic acid, mucic acid, benzoic acid, glucuronic acid, oxalic acid, ascorbic acid, acetylglycine. Of the new peptides, those are preferred in which C Lys, Orn, Arg or hArg, G Ile-Asn, Leu-Asn, Val-Asn or Asn, H Leu-Ile, Leu-Val, Leu-Leu, Val-Leu , Val-Val, Val-Ile, Ile-Ile, Ile-Leu or Ile-Val, I orn, Lys, Arg or hArg, J Orn, Lys, Arg or hArg and Z is a hydrogen atom, an amino protecting group, an acetyl, p-hydroxybenzoyl-,

p-Hydroxyphenylacetyl- or p-Hydroxyphenylpropionylrest mean.

The peptides in which

C Lys or Arg,

E the grouping -NH- (CH ₂ ) t-CO- (with t meaning an integer from 1 to 7)

F Cys or Abu,

G Ile-Asn or Asn H Leu-Ile

I Arg

J Arg

Z is a hydrogen atom, an acetyl, p-hydroxybenzoyl,

p-Hydroxyphenylacetyl-, p-Hydroxyphenylpropionylrest mean and A, B, D, K and k-q have the meanings given.

The new compounds can be prepared by methods known in peptide chemistry.

So you can build the peptides sequentially from amino acids or by fragment linking suitable small peptides. In the sequential construction, the peptide chain is gradually extended by one amino acid each, starting at the C terminus. In the fragment coupling, fragments of different lengths can be linked to one another, the fragments in turn being obtained by sequential construction from amino acids or, in turn, by fragment coupling. After the synthesis of the open-chain peptides, the cyclic peptides are obtained by a cyclization reaction carried out in high dilution. Both in the sequential structure and in the fragment coupling, the building blocks must be linked by forming an amide bond. Enzymatic and chemical methods are suitable for this. Chemical methods for forming amide bonds are dealt with in detail by Müller, Methods of Organic Chemistry Vol XV / 2, pp 1 - 364, Thieme Verlag, Stuttgart, 1974; Stewart, Young, Solid Phase Peptide Synthesis, pp 31-34, 71-82, Pierce Chemical Company, Rockford, 1984; Bodanszky, Klausner, Ondetti, Peptide Synthesis, pp 85 - 128, John Wiley & Sons, New York, 1976 and other standard works of peptide chemistry. The azide method, the symmetrical and mixed anhydride method, in situ generated or preformed active esters and the amide bond formation with the aid of coupling reagents (activators), in particular dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), 1-ethoxycarbonyl-2-ethoxy, are particularly preferred -1,2-dihydroquinoline (EEDQ), 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI), n-propanephosphonic anhydride (PPA), N, N- bis (2-oxo-3-oxazolidinyl) amidophosphoric acid chloride ( BOP-Cl), diphenylphosphoryl azide (DPPA), Castro's reagent (BOP), 0-benzotriazolyl-N, N, N ', N'-tetramethyluronium salts (HBTU), 2,5-diphenyl-2,3-dihydro -3-oxo-4-hydroxy-thiophene dioxide (Steglich reagent; HOTDO) and 1, 1'-carbonyl-diimidazole (CDI). The coupling reagents can be used alone or in combination with additives such as N, N'-dimethyl-4-aminopyridine (DMAP), N-hydroxybenzotriazole (HOBt), N-hydroxybenzotriazine (HOOBt), N-hydroxysuccinimide (HOSu) or 2-hydroxypyridine .

While protective groups can normally be dispensed with in enzymatic peptide synthesis, reversible protection of the reactive functional groups of the two reactants which are not involved in the formation of the amide bond is required for chemical synthesis. In chemical peptide synthesis, three protecting group techniques known from the literature are preferred: the benzyloxycarbonyl (Z), the t-butyloxycarbonyl (Boc) and the 9-fluorenylmethyloxycarbonyl (Fmoc) protective group technique. The protective group of the α-amino function of the chain-extending building block is designated in each case. The side chain protecting groups of the trifunctional amino acids are chosen so that they are not necessarily split off together with the α-amino protecting group. Müller, Methods of Organic Chemistry Vol XV / 1, pp 20 - 906, Thieme Verlag, Stuttgart, 1974 provides a detailed overview of amino acid protecting groups. The building blocks which serve to build up the peptide chain can be in solution, in suspension or by a similar process as described by Merrifield in 3rd Amer. Chem. Soc. 85, 2149, 1963. Methods in which peptides are sequential are particularly preferred or by fragment coupling using the Z, Boc or Fmoc protective group technique, the reactants being reacted in solution, and processes in which, similar to the Merrifield technique mentioned, a reactant to an insoluble polymeric support (hereinafter also called resin) bound to react. The peptide is typically built up sequentially on the polymeric support using the Boc or Fmoc protective group technique, the growing peptide chain at the C-terminus being covalently linked to the insoluble resin particles (see Figs. 1 and 2). This procedure allows reagents and by-products to be removed by filtration, making recrystallization of intermediate products unnecessary.

The protected amino acids can be bound to any suitable polymer, which are only insoluble in the solvents used and must have a stable physical form which enables easy filtration. The polymer must contain a functional group to which the first protected amino acid can be bound by a covalent bond. A wide variety of polymers are suitable for this purpose, e.g. Cellulose, polyvinyl alcohol, polymethacrylate, sulfonated polystyrene, chloromethylated copolymer of styrene and divinylbenzene (Merrifield resin), 4-methylbenzhydrylamine resin (MBHA resin), phenylacetamidomethyl resin (Pam resin), p-benzyloxybenzyl alcohol resin , Benzhydrylamine resin (BHA resin), 4- (hydroxymethyl) benzoyl oxymethyl resin, resin according to Breipohl et al. (Tetrahedron Lett. 28, 565, 1987; BACHEM), HYCRAM resin (ORPEGEN) or SASRIN resin

(BACHEM).

All solvents which prove inert under the reaction conditions are particularly suitable for peptide synthesis in solution, in particular water, N, N'-dimethylformamide (DMF), dimethyl sulfoxide (DMSO), acetonitrile, dichloromethane (DCM), 1,4- Dioxane, tetrahydrofuran (THF), N-methyl-2-pyrrolidone (NMP) and mixtures of the solvents mentioned. The peptide synthesis on the polymeric support can be carried out in all inert organic solvents in which the amino acid derivatives used are soluble; however, solvents which additionally have resin-swelling properties, such as DMF, DCM, NMP, acetonitrile and DMSO, and mixtures of these solvents are preferred. After successful synthesis, the peptide is split off from the polymeric carrier. The conditions under which the peptides can be split off from the various types of resin are known from the literature. Acid and palladium-catalyzed cleavage reactions are most commonly used, in particular the cleavage in liquid anhydrous hydrogen fluoride, in anhydrous trifluoromethanesulfonic acid, in dilute or concentrated trifluoroacetic acid or the palladium-catalyzed cleavage in THF or THF-DCM mixture in the presence of a weak base such as, for example, morpholine. Depending on the choice of the protective groups, these can be retained under the cleavage conditions or can also be split off. Partial deprotection of the peptide can also be useful if certain derivatization reactions or a cyclization are to be carried out. The new peptides bind with high affinity to NPY receptors, but without triggering the physiological effect of the NPY, are therefore competitive antagonists and suppress the effect of the endogenous NPY. The new peptides have hypotensive properties and are valuable medicinal products that are suitable for the treatment of circulatory diseases, in particular high blood pressure, and vascular spasms. They can be combined with ACE inhibitors, Ca antagonists or diuretics.

The new peptides are also valuable diagnostic and analytical reagents for biochemical and medical research, for researching hypertension and especially for studying the biochemistry and pathophysiology of NPY.

The biological characterization of the new peptides was carried out in the following test systems:

1. Receptor binding assay (rabbit kidney cortex)

2. Functional tests on the despinalized and anesthetized rats.

1. Receptor binding assay

Rabbit renal cortex was prepared immediately after the kidney was removed and homogenized in 0.32 M sucrose solution (4 ° C.). The homogenate was filtered and then centrifuged at 480 g (5 min, 4 ° C), the supernatant collected and centrifuged at 45,000 g (10 min, 4 ° C). The resulting residue was washed by resuspension and recentrifugation twice with incubation buffer (25 mM Tris, 5 mM MgCl ₂ , 0.5% bovine serum albumin and 0.1% bacitracin, pH 7.4) and taken up in incubation buffer. The test batches (1 ml) were composed of: 100 μg membrane protein (BCA protein assay, Pierce, Rockford, Illinois, USA), 0.5 nM 3H-NPY (Amersham, Braunschweig, specific radioactivity 3 TBq / mmol) in incubation buffer (total binding) or a) with an additional 0.3 μM NPY (non-specific binding) or b) with test substance. The batches were made as triplicates.

After the incubation had ended (60 min, 30 ° C.), the batches were filtered through glass fiber filters (Whatman GF / B) and washed with ice-cold washing buffer (50 mM Tris, 0.5 mM EDTA, 0.2% bovine serum ibumin, pH 7.4) . The radioactivity retained on the filters was determined by means of liquid scintillation measurement. The nonspecific binding was approx. 20 to 30% of the total binding at 0.5 nM 3H-NPY. The competition curves are evaluated using iterative nonlinear regression analysis based on the "Ligand" program (Munson and Rodbard: Anal. Biochem. 107, 220, 1980).

2. a) Functional test for NPY antagonism in the despinalized normotonic rat

Male Sprague-Dawley rats of 250 to 300 g body weight were used for the experiments. The feeding was 16 to

Suspended 24 hours before the start of the test, water was available ad libitum. The animals were included 30 to 40 minutes before the start of the experiment

Amobarbital 120 mg / kg i.p. anesthetized. Ventilation was carried out artificially using a breathing pump with a breathing volume of 3 ml / stroke and 50 breaths / min. The substance application was carried out i.v. via the jugular vein with an injection volume of 1 ml / kg and an injection duration of 20 sec.

The trachea, carotid artery and jugular vein were prepared 10 min after the initiation of post-coughing, and the despinalization and vagotomy were then carried out. NPY injections (3 μg / kg IV) were given 15 min before and 5 min after substance administration. The blood pressure in the carotid artery was measured with Statham pressure transducers, the amplifiers and recording devices were from Hellige. The relative inhibition (Δ%) of the increase in blood pressure caused by NPY (Δ mmHg) was measured.

2. b) Blood pressure measurement on the normotonic rat

Male Sprague-Dawley rats of 250 to 300 g body weight were used for the experiments. The feeding was 16 to Suspended 24 h before the start of the experiment, water was available ad libitum. The animals were anesthetized with urethane 1.7 g / kg ip 30 to 40 min before the start of the experiment. Breathing was spontaneous via tracheal cannula.

The trachea, carotid artery and jugular vein were dissected 20 minutes after induction of anesthesia. Subcutaneous electrode needles were used to record the ECG. The mean carotid pressure MAP (mmHg) was measured via Statham transducer P23Db. The heart rate HR (min ^-1 ) was determined from the blood pressure amplitude. Both parameters were registered with Heilige writers.

The dose-response relationship was described by a linear regression analysis between log dose (mg / kg) and the relative change in blood pressure (y = a + b. Log x). Animals with ECG changes were registered.

The following examples are intended to explain the invention in more detail. The proteogenic amino acids are abbreviated in the examples with the three-letter code (according to Eur. J. Biochem. 138, 1984, 9-37). Furthermore: Abs = 4-aminobutyric acid, Abu = 2-aminobutyric acid, Ac = acetic acid, Ahx = aminohexanoic acid, Aoc = 8-aminooctanoic acid, Ape = 5-aminopentanoic acid, HBz = p-hydroxybenzoic acid, Hpac = p-hydroxyphenylacetic acid,

Hpp = p-hydroxyphenylpropionic acid.

A. General working instructions

The synthesis of the peptides according to claim 1 was carried out using the standard methods of solid phase peptide synthesis on a fully automatic device model 430 A from APPLIED BIOSYSTEMS.

The following synthesis cycle for the Boc protection group technology was used:

1. 30% trifluoroacetic acid in DCM 1 x 3 min

2. 50% trifluoroacetic acid in DCM 1 x 17 min

3. DCM washing step 5 x 1 min

4. 5% diisopropylethylamine in DCM 1 x 1 min. 5% diisopropylethylamine in NMP 1 x 1 min

6. NMP wash 5 x 1 min 7. Add the preactivated protected amino acid (activation by 1 equivalent of DCC and

1 equivalent of HOBt in NMP / DCM);

Peptide coupling (1st part) 1 x 30 min 8. Add DMSO to the reaction mixture up to

a volume share of 20% DMSO

9. Peptide coupling (2nd part) 1 x 16 min

10. Add 3.8 equivalents of diisopropylethylamine

to the reaction mixture

11. Peptide coupling (3rd part) 1 x 7 min

12. DCM washing step 3 x 1 min

13. Repetition of incomplete sales

Clutch (back to 5.)

14. 10% acetic anhydride, 5% diisopropylethylamine

in DCM 1 X 2 min

15. 10% acetic anhydride in DCM 1 X 4 ml

16. DCM washing step 4 x 1 min

17. Back to 1. B. Special work regulations

example 1

H-Tyr-Abu-Ser-Lys-Aoc-Abu-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

0.5 g of Boc-Tyr (Br-Z) -p-methylbenzhydrylamine resin with a substitution of 0.49 mmol / g, corresponding to a batch size of 0.25 mmol, were, according to A, with 1 mmol of Boc-Arg (Tos) - OH Boc-Ile-OH

Boc-Gln-OH Boc-Abu-OH

Boc-Arg (Tos) -OH Boc-Aoc-OH

Boc-Thr (Bzl) -OH Boc-Lys- (Cl-Z) -OH

Boc-Ile-OH Boc-Ser (Bzl) -OH

Boc-Leu-OH Boc-Abu-OH

Boc-Asn-OH Boc-Tyr (Br-Z) -OH implemented. After completion of the synthesis, the resin was deprotected at the N-terminal. (Execution of steps 1-3 according to A).

The peptide resin obtained (1.06 g) was subjected to HF cleavage (10 ml HF / 1 ml anisole; 0 ° C, 45 min). HF and most of the anisol were removed in vacuo, the resin was washed with ethyl acetate, and extracted the peptide with 10% acetic acid. The extract was lyophilized. The crude peptide was purified by medium pressure chromatography (HD-SIL® reversed phase C ₁₈ material, 100 Å, 25-45 μ grain size) with a gradient of 50-70% B in 80 min

(A = 0.1% TFA in H ₂ O; B = 0.1% TFA in MeOH). Yield: 93 mg.

Analyte. HPLC: retention time 14.3 min (VYDAC ^R 218TP54-C ₁₈ RP column; flow 2 ml / min; t _o = 1.8 min; linear gradient from 0% - 50% B in 50 min; A = 0, 1% TFA in H ₂ O; B = 0.1% TFA in acetonitrile). FAB-MS: M = 1864.2; calculated monoisotopic molecular mass: 1864.1.

Preparation was carried out analogously to Example 1 (steps 1 - 6 and 14 - 16 according to A were carried out for the N-terminally acetylated peptides after the synthesis had ended):

2. H-Tyr-Pro-Ser-Lys-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

3. H-Tyr-Pro-Ser-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂ 4. H-Tyr-Pro-Aoc-Ile-Asn-Leu-Ile- Thr-Arg-Gln-Arg-Tyr-NH ₂

5. H-Tyr-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

6. H-Tyr-Pro-Lys-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

7. H-Tyr-Lys-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

8. H-Tyr-Pro-Arg-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂ 9. H-Gly-Aoc-Ile-Asn-Leu-Ile-Thr- Arg-Gln-Arg-Tyr-NH ₂

10. H-Tyr-Arg-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

11. H-Phe-Pro-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

12. H-Phe-Pro-Ser-Lys-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

13. H-Tyr-Pro-Ser-Arg-Aoc-Ile-Asn-Leu-lle-Thr-Arg-Gln-Arg-Tyr-NH ₂ 14. H-Phe-Pro-Ser-Arg-Aoc-Ile- Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

15. H-Tyr-Pro-Ahx-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

16. H-Tyr-Pro-Abs-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂ 17. Ac-Tyr-Pro-Aoc-Ile-Asn-Leu-lle-Thr-Arg-Gln-Arg-Tyr-NH ₂

18. Hpp-Pro-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

19. Hpac-Pro-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

20. Hbz-Pro-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

21. H-Tyr-Aoc-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

22. H-Tyr-Lys-Aoc-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

23. Ac-Tyr-Aoc-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

24. Hpp-Pro-Ser-Lys-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

25. H-Tyr-Lys-Ahx-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

26. H-Tyr-Lys-Abs-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

27. H-Tyr-Lys-Aoc-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

28. H-Tyr-Lys-Ahx-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

29. H-Tyr-Lys-Abs-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

30. H-Tyr-Lys-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

31. H-Tyr-Pro-Aoc-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH2

32. H-Tyr-Pro-Ser-Lys-Aθc-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

33. Ac-Phe-Pro-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂ 34. H-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln- Arg-Tyr-NH ₂

35. Ac-Tyr-Lys-Aoc-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂ Example 36

Hpp-Cys-Ser-Lys-Aoc-Cys-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂ 0.5 g of Boc-Tyr (Br-Z) -p-methylbenzhydrylamine resin with a substitution of 0.49 mmol / g, corresponding to a batch size of 0.25 mmol, were according to A with 1 mmol of Boc-Arg (Tos) -OH Boc-Ile-OH

Boc-Gln-OH BθC-Cys (pMB) -OH

Boc-Arg (Tos) -OH Boc-Aoc-OH

Boc-Thr (Bzl) -OH Boc-Lys- (Cl-Z) -OH

Boc-Ile-OH Boc-Ser (Bzl) -OH

Boc-Leu-OH Boc-Cys (pMB) -HO

Boc-Asn-OH Hpp-OH implemented. The peptide resin obtained (1.01 g) was subjected to HF cleavage (10 ml HF, 1 ml m-cresol; 0 ° C., 45 min). The HF was in a vacuum

stripped, the resin washed with ethyl acetate, and the peptide extracted with 10% acetic acid. The lyophilized crude peptide was dissolved in 1 liter of degassed water and the pH of the solution was adjusted to 8.4 with aqueous ammonia. 0.01 N aqueous K ₃ Fe (CN) ₆ solution was slowly added dropwise until the yellow color persisted. It was 1 h

stirred and acidified to pH 4.5 with glacial acetic acid. After adding BIORAD ^® AG3-X4A anion exchange resin, the mixture was stirred for 2 h, filtered and the filtrate lyophilized.

The crude peptide was purified by gel chromatography

(SEPHADEX ^® G-25, 10% HOAc) and subsequent medium pressure chromatography (HD-SIL ^® reversed phase C ₁₈ material, 100 A, 25-45 μ grain size) with a gradient of 50 - 70% B in 40 min (A = 0.1% TFA in H ₂ O; B = 0.1% TFA in MeOH). Yield: 87 mg.

Analyte. HPLC: retention time 13.9 min (VYDAC ^® 218TP54-C ₁₈ RP column; flow 2 ml / min; t _o = 1.8 min; linear gradient from 0% - 50% B in

50 min; A = 0.1% TFA in H ₂ O; B = 0.1% TFA in acetonitrile). FAB-MS: M = 1882.9; calculated monoisotopic molecular mass: 1883.0.

The following were prepared as in Example 36:

37. H-Tyr-Cys-Ser-Lys-Aoc-Cys-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

38. H-Tyr-Cys-Ser-Lys-Ahx-Cys-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

39. H-Tyr-Cys-Ser-Lys-Abs-Cys-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

40. H-Tyr-Cys-Ser-Lys-Cys-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

41. H-Tyr-Pro-Ser-Lys-Cys-Aoc-Cys-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr-NH ₂

Claims

1. Peptides of the formula ZA _K -B _l -C _m -D _n -E _o -F _p -G _q -H-Thr-I-Gln-JK-NH ₂ wherein

A Phe or Tyr,

B Pro, Ser, Abu, Gly, Ala, Cys, Pro-Ser, Cys-Ser, Abu-Ser, Gly-Ser, Ala-Ser or Pro-Ala,

C an amino acid with a basic side chain,

D Pro or Cys,

E the grouping -NH- (CH ₂ ) _t -CO- (with t meaning a

integer from 0 to 10),

F Cys, Abu, Gly or Ala, G Ile-Asn, Leu-Asn, Val-Asn or Asn, H a dipeptide residue composed of lipophilic amino acids, I an amino acid with a basic side chain,

J is an amino acid with a basic side chain, K Phe or Tyr and

Z represents a hydrogen atom, an amino protecting group, a benzyl radical, a C _2-10 acyl radical or a C _1-5 alkyl radical, k, l, m, n, o, p and q denote the numbers 0 or 1, but with k + l + m + n + o + p + q ≥ 1, and two possibly present Cys residues can be connected to one another via a disulfide bridge, as well as their salts with physiologically acceptable acids.

2. A process for the preparation of the peptides according to claim 1, characterized in that they are prepared by methods known in peptide chemistry.

3. Peptides according to claim 1 for use in combating diseases.

4. Use of the peptides according to claim 1 for combating high blood pressure

5. Use of the peptides according to claim 1 for the production of diagnostic and analytical reagents for biochemical and medical research.