DD241909A5 - GnRH Antagonists. VII - Google Patents

Abstract

The invention relates to a process for the preparation of peptides having antagonistic properties to gonadotropic hormones. The peptides should have the property of preventing the release of steroids by the germs in mammals. The peptides prepared according to the invention have the structure XR1 (W) DPheR3R4R5R6R7ArgProR10, wherein X is a hydrogen atom or an acyl group of not more than 7 carbon atoms R1 dehydroPro, DpGlu, DPhe, 4ClDPhe, DTrp, Pro or bDNAL; W is Cl, F, NO2, CaMe / 4Cl, Cl2 or BrR3 D3PAL, bDNAL or (Y) DTrp with YH, NO2, NH2, OCH3, F, Cl, Br, CH3, NinHCO or NinAcR4 Ser, Orn, AAL or aBu; R5 Tyr, Arg, (3F) Phe, (2F) Phe, (3I) Tyr, (3CH3) Phe, (2CH3) Phe, (3Cl) Phe or (2Cl) Phe; or (2Cl) Phe; R6 is a D-isomer of a lipophilic amino acid or 4NH2DPhe, DLys, DOrn, DHar, DHis, 4guaDPhe, DPAL or DArg R7 Nle, Leu, NML, Phe, Met, Nva, Tyr, Trp, Cys, PAL or 4FDPhe and R10 GlyNH2, DAlaNH2, NHY with Y is lower alkyl, cycloalkyl, fluoro-substituted lower alkyl or NHCONHQ with QH or lower alkyl, R3 is DPAL, then R5 is Arg, and when R3 is bDNAL or DTrp, then R7 is Tyr, PAL, Phe or 4FDPhe. The peptides prepared according to the invention inhibit the secretion of gonadotropins by the pituitary and the release of steroids by the germ cells. Administration of an appropriate dose will prevent ovulation of female mammals and / or the release of steroids by the germs.

Description

Field of application of the invention

The invention relates to a process for the production of peptides which inhibit the release of gonadotropins by the pituitary gland in mammals, including humans, as well as prevent ovulation and / or inhibit the release of steroids.

Characteristic of the known state of the art

The pituitary gland is attached with a pedicle to the hypothalamic region of the brain base. Specifically, the pituitary gland releases the follicololotropic hormone (FSH) and the luteogenic hormone (LH), both of which are also called gonadotropins or gonadotropic hormones. These hormones regulate the function of the gonads in the production of testosterone in the testicles and progesterone and estrogen in the ovaries and also regulate the formation and maturation of germ cells.

The release of a hormone by the anterior pituitary usually requires the prior release of another hormone class produced by the hypothalamus. One of the hypothalamic hormones acts as a factor that triggers the release of gonadotropic hormones, specifically LH. This hormone is referred to below as GnRH. It is also referred to in the literature as LH-RH and LRF. GnRH was isolated and analyzed as a decapeptide with the following structure: p-Glu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH ₂ . Peptides are compounds containing two or more amino acids with the carboxy group of one acid attached to the amino group of the other acid. The above formula of GnRH corresponds to the usual representation of peptide formulas with the amino-terminus on the left and the carboxy-terminus on the right. The position of each amino acid residue is indicated by numbering the amino acid residues from left to right. In the case of GnRH, the hydroxy group of glycine is replaced by an amino group (NH ₂ ). The abovementioned abbreviations for the individual amino acid residues are generally known and mean:

p-Glu is pyroglutamic acid,

His-histidine,

Trp tryptophan,

Ser serine,

Type tyrosine,

Gyl Gylcin,

Leucine leucine,

Orn ornithine,

Arg arginine,

Pro proline, -.

SarSarcosin,

Phe phenylalanine,

and Ala Alanine.

The amino acids, as well as valine, isoleucine, threonine, lysine, aspartic acid, asparagine, glutamine, cysteine, methionine, phenylalanine and proline are referred to as ordinary, natural or peptide-derived amino acids.

Object of the invention

The object of the invention is to obtain _% improved peptides which are potent antagonists of endogenous GnRH and prevent the secretion of lutogenic hormones and the release of steroids by the gonads in mammals.

Explanation of the essence of the invention

The invention has for its object to provide a process for the preparation of peptides that show strong antagonistic properties towards endogenous GnRH.

The peptides of the present invention are represented by the formula I X-R rlWlD-Phe-Rs-R ^ Rs -Re -Ry-Arg-Pro-Rio, where X is hydrogen or an acyl group of at most 7 carbon atoms, R ₁ dehydro-Pro ^ -pGlu.D-Phe ^ CI-D-Phe, D-Trp, Pro or β-D-NAL, WCI, F, NO ₂ , C ^a Me / 4CI, Cl ₂ or Br, R ₃ D-3PAL, β D-NAL or (Y) D-Trp with Y = H, NO ₂ , NH ₂ , OCH ₃ , F, Cl, Br, CH ₃ , N ⁱⁿ HCO or N _in Ac, R ₄ Ser, Orn, AAL or aBu, R ₅ Tyr, Arg, (3F) Phe, (2F) Phe, (3I) Tyr, (3CH ₃ ) Phe, (2CH ₃ ) Phe, (3CI) Phe or (2CI) Phe, R ₆ a D- Isomer of a lipophilic amino acid or 4-NH ₂ -D-Phe, D-Lys, D-Orn, D-Har, D-His, 4-gua-D-Phe, D-PAL or D-Arg, R ₇ Never, Leu, NML, Phe, Met, Nva, Tyr, Trp, Cys, PAL or 4F-D-Phe and R ^{10 is} Gly-NH ₂ , D-Ala-NH ₂ or NH-Y 'with Y' = lower alkyl, cycloalkyl , fluorine-substituted lower alkyl or NHCONH-Q with Q = H or lower alkyl. When R _{3 is} D-3PAL, R _{5 is} Arg; when R _{3 is} β-D-NAL or D-Trp, R _{7 is} Tyr, PAL, Phe or 4F-D-Phe; when R ₁ β-D-NAI and R _{5 is} not Arg, R _{6 is} preferably 4-NH ₂ -D-Phe, D-Lys, D-Orn, D-Har, D-His, 4-gua-D Phe, D-PaI or D-Arg. By β-D-NAL is meant D-alanine that is substituted at the β-carbon atom by naphthyl, ie 3-D-NAL. Preferably, β-D-2NAL is used in which the naphthalene is attached via the 2-position of the ring structure, but also β-D-1 NAL can be used. PAL denotes alanine that is substituted by pyridyl on the β-carbon atom, preferably the bonding takes place via the 3-position of the pyridine ring. In D-Trp in the 3-position, a hydrogen atom in the 5- or 6-position by chlorine, fluorine, bromine, methyl. Amino, methoxy or nitro may be substituted, with chlorine, fluorine and nitro are preferred. Alternatively, the indole nitrogen atom may be acylated, e.g. B. with formyl (N ^'n HC0- or 1HCO-) or acetyl. The preferred substituted residues are N ^'n HCO-D-Trp-D-Trp and 6NO _2. When unsubstituted D-Trp is used in the 3-position, a specific residue is introduced in the 7-position, namely Phe or 4F-D-Phe or preferably Tyr or PAL. NML refers to N ° CH ₃ -L-Leu. AAL denotes β-amino-Ala, aBu denotes α, γ-diamino-butyric acid. These two or Orn can be in the 4 position. If Ser is not in the 4-position, Dehydro-Pro is preferably present in the 1-position. 4-gua-D-Phe means the p-position substituted with guanidino D-Phe radical.

The peptides of the present invention can be prepared by classical synthesis in solution or by a solid phase method on a chloromethylated resin, a methyl benzhydrylamine resin (MBHA), a benzhydrylamine resin (BHA), or other suitable conventional resin. Solid phase synthesis is accomplished by sequential attachment of the amino acids in the chain in the manner described in detail in US Pat. No. 4,211,693. Side chain protecting groups are preferably added in the usual manner to existing Ser, Tyr, and Arg, as well as certain substituents, and may also be added to Trp before these amino acids are attached to the chain built on the resin. This method gives the fully protected peptide-resin intermediate. The intermediates of the process according to the invention can be represented by the following formula: Xi-R 1 -Wl-D-Phe-RafX 1 -RifX 1 -RsiX ⁴ or X ⁵ ) -R ₆ (X ⁵ ) -R ₇ (X ⁶ ) -Arg (X ⁶ ) -Pro-X ⁷ ,

wherein X ^{1 is} an a-amino protecting group suitable for the stepwise synthesis of polypeptides. When X in the desired peptide composition is a certain acyl group, this group can be used as a protecting group. As a-amino-protecting groups X ¹ , the following classes of compounds can be used:

1. Acyl groups such as formyl, trifluoroacetyl, phthalyl, p-toluenesulfonyl (Tos), benzoyl (Bz), benzenesulfonyl, o -nitrophenylsulfenyl (Nps), tritylsulfenyl, o-nitro-phenoxyacetyl, acryloyl (Acr), chloroacetyl, acetyl (Ac) and γ-chlorobutyryl;

2. aromatic urethane protecting group, e.g. Benzyl oxycarbonyl (Z) and substituted benzyloxycarbonyl such as p-chlorobenzyloxycarbonyl (CIZ), p-nitrobenzyloxycarbonyl, p-bromobenzyloxycarbonyl and p-methoxybenzyloxycarbonyl;

3. aliphatic urethane protecting groups such as tert-butoxycarbonyl (Boc), diisopropylmethoxycarbonyl, isopropoxycarbonyl, ethoxycarbonyl and allyloxycarbonyl;

4. cycloalkyl urethane protecting groups such as cyclopentyloxycarbonyl, adamantyloxycarbonyl and cyclohexyloxycarbonyl;

5. Thiourethane protecting groups, such as (phenylthio) carbonyl;

6. alkyl protecting groups such as allyl (Al y), triphenylmethyl (trityl) and benzyl (Bzl);

7. Trialkylsilane protecting groups, such as trimethylsilane.

When X is hydrogen, the preferred a-amino protecting group is Boc.

X ₂ is hydrogen or a protecting group for the indole nitrogen of Trp, such as benzyl. In many syntheses, however, Trp does not need to be protected.

X ³ is hydrogen or a protecting group for the alcoholic hydroxy group of Ser, such as acetyl / benzoyl, tetrahydropyranyl, tert-butyl, trityl, benzyl or 2,6-dichlorobenzyl, with benzyl being preferred. When Ser is substituted by another amino acid, X ³ may be a protective group for an amino group in the side chain, such as Tos, Z or CIZ.

X ⁴ is hydrogen or a protecting group for the phenolic hydroxy group of Tyr (if present), namely tetrahydropyranyl, tert -butyl, trityl, benzyl, benzyloxycarbonyl, 4-bromo-benzyloxycarbonyl or 2,6-dichloro-benzyl, wherein 2,6-dichloro-benzyl (DCB) is preferred.

X ⁶ is a protective group for the guanidino group or the side-chain amino group or the imidazole group of Arg, Lys, His etc., such as nitro, tos, trityl, benzyloxycarbonyl, adamantyloxycarbonyl, Z and Boc, a protecting group for Tyr, such as X ⁴ or a protecting group for Trp such as X ² , or H, ie that there is no protecting group on the atoms of the side chain, Tos being generally preferred.

X ⁶ is hydrogen, a protecting group for Tyr, such as X ⁴ , or a protecting group for Cys, preferably p-methoxybenzyl (MeOBzI), p -methylbenzyl, acetylaminomethyl, trityl and BzI. The most preferred protecting group is p-methoxybenzyl.

X ⁷ represents a Gly-O-CH ₂ - (carrier resin), O-CH ₂ - (carrier resin), D-Ala-O-CH ₂ - (carrier resin), Gly-NH- (carrier resin) or D-Ala NH (carrier resin), but may also be OH, ester, amide or hydrazide of Gly or D-Ala or be directly bonded to Pro.

The criterion for the choice of side chain protecting group X ² -X ₆ is that the protecting group must be stable to the reagent chosen under the particular reaction conditions for cleaving the α-aminoprotection group at each stage of the synthesis. The protecting group should not be cleaved under the coupling conditions, but should be cleavable upon completion of the synthesis of the desired amino acid sequence under reaction conditions under which the peptide chain is not altered.

When X ^{7 is} Gly-O-CH ₂ - (carrier resin), D-Ala-O-CH ₂ - (carrier resin) or O-CH ₂ - (carrier resin), the ester component is one of the many functional groups of the polystyrene carrier resin shown. When X ^{7 is} Gly-NH- (carrier resin) or D-Ala-NH- (carrier resin), GIy or D-Ala is attached to a BHA resin or MBHA resin through an amiol bond.

If X in the final formula z. Acetyl, may be useful as the protective group X ¹ for the α-amino group of D-NAL or any other amino acid used in the 1-position by introducing it to the peptide chain prior to the coupling of this last amino acid. Preferably, however, a reaction with the peptide on the resin (after deblocking the α-amino group leaving the groups protected in the side chain) is carried out, e.g. Example by reaction with acetic acid in the presence of dicyclohexylcarbodiimide (DCC) or preferably with acetic anhydride or by another known suitable reaction.

As is known from the literature, the fully protected peptide can be cleaved from a chloromethylated carrier resin by ammonolysis and then provides the fully protected amide intermediate. The deprotection of the.

Peptide and cleavage of the peptide from a benzhydrylamine resin can be carried out at 0 ° C with hydrofluoric acid (HF).

Anisole is preferably added to the peptide prior to treatment with HF. After removal of HF in vacuo, the cleaved, unprotected peptide is best treated with ether, decanted, taken up in dilute acetic acid and lyophilized.

According to the inventive method one, X receives a peptide of the formula X-Ri (W) D-Phe-R3-R ₄ -R ₅ -R 6 -R 7-Arg-Pro-R _10, or a non-toxic salt thereof wherein Hydrogen atom or an acyl group of not more than 7 carbon atoms, R ₁ dehydro-Pro, D-pGlu, D-Phe, 4CI-D-Phe, D-Trp, Pro, or β-D-NAL, WCI, F, NO ₂ , C ^a Me / 4CI, Cl ₂ or Br, R ₃ D-3-PAL, β-D-NAL or (Y) D-Trp with Y = H, NO ₂ , NH ₂ , OCH ₃ , F, Cl, Br, CH ₃ , N ⁱⁿ HCO or N ⁱⁿ Ac, R ₄ Ser, Orn, AAL or aBu,

R _{5 is} Tyr, Arg, (3F) Phe, (2F) Phe, (3I) Tyr, (3CH ₃ ) Phe, (2CH ₃ ) Phe, (3CI) Phe or (2CI) Phe, R _{6 is} a D-isomer of one lipophilic amino acid or 4-NH ₂ -D-Phe, D-Lys, D-Om, D-Har, D-His, 4-gua-D-Phe, D-PAL or D-Arg, R ₇ NLe, Leu, NML, Phe, Met, Nva, Tyr, Trp, Cys, PAL or 4F-D-Phe and

R _{10 is} Gly-NH ₂ , D-Ala-NH ₂ or NH-Y 'with Y' = lower alkyl, cycloalkyl, fluoro-substituted lower alkyl or NHCONH-Q with Q = H or lower alkyl, but where R _{3 is} D 3PAL, R _{5 is} Arg, and when R _{3 is} β-D-NAL or D-Trp, R _{7 is} Tyr, PAL, Phe or 4F-D-Phe; to form an intermediate compound of the formula X ¹ -R _r (W) D-Phe-R ₃ (X ² ) -R ₄ (X ³ ) -R ₅ (X ⁴ or X ⁵ IR ₆ (X ⁵ ) -R ₇ ( X ⁶ ) -Arg (X ⁵ ): Pro-X ⁷ ,

where X ^{1 is} a hydrogen atom or an α-amino protective group,

X ^{2 is} a hydrogen atom or a protective group for the indole nitrogen,

X ^{3 is} hydrogen or a protective group for the alcoholic hydroxy group of Ser or for an amino group in the side chain,

X ^{4 is} a hydrogen atom or a protecting group for the phenolic hydroxy group of Tyr, X ⁵ and X ^{6 are} each either a hydrogen atom or a protecting group for each side chain present, and R ^{7 is} Gly-O-CH ₂ - (carrier resin), O-CH ₂ - (carrier resin), D-Ala-O-CH ₂ - (carrier resin), Gly-NH- (carrier resin), D-Ala-NH- (carrier resin), GIy-NH ₂ or an ester, amide or hydrazide

Cleavage of one or more of the groups X ¹ to X ⁶ and / or cleavage of a carrier resin contained in X ⁷ , and if desired, by conversion of the resulting peptide into a non-toxic salt.

Purification of the peptide is carried out by ion exchange chromatography on a carboxymethylcellulose column followed by partition chromatography with the elution system butane-i-ol / 0.1 N acetic acid (1: 1 by volume) on a Sephadex G-25 filled column or by high pressure chromatography as in in the literature and specifically described in J. Rivier et al., J. Chromatography, 288 (1984), pp. 303-328. Except glycine, the amino acids of the peptides prepared according to the invention have the L-configuration, unless stated otherwise.

The peptides obtained according to the invention have a substitution of the 1-position, preferably by dehydropoline or ß- (naphth-1 or 2-yl) -D-analin, hereinafter referred to as ß-D-1 NAL or ß-D-2NAL , the 2-position in the form of a modified D-Phe, the 3-position preferably in the form of substituted D-Trp, D-3PAL or β-D-NAL, a possible substitution by a diamino acid having not more than 5 carbon atoms in FIG Position, a possible 5-position substitution in the form of (a) a halogenated or methylated L-Phe or L-Tyr or (b) Arg, a substitution of the 6-position and a possible substitution of the 7-position by Nie, NML , Phe, Nva, Met, Tyr, Trp, Cys, PAL, 4F-D-Phe and so on. A modified D-Phe in the 2-position provides an increased antagonistic effect by specific modifications in the benzene ring.

Simple substitution in the ring takes place in the para or 4 position by chlorine, fluorine, bromine and nitro, preference being given to chlorine, fluorine and nitro. Dichloro substitution occurs in the 2,4 or 3,4 position of the ring. The α-carbon atom may be methylated at the same time, for. B. (C ^a Me / 4CI) Phe. When D-Trp or β-D-NAL is present in the 3-position, R _{7 is} preferably Tyr, PAL, Phe or 4F-D-Phe; is in the 3-position D-3PAL, R _{5 is} Arg. The substituent in the 1-position can be modified such that its a-amino group is an acyl group such as formyl (HCO), acetyl, acryloyl, but-3-enyl ( Vinyl acetyl, Vac) or benzoyl (Bz), with acetyl (Ac) and acryloyl (Acr) being preferred. The symbols PAL and D-PAL represent the L- or D-isomers of pyridylalanine, wherein preferably the 3-position of the pyridine ring is bonded to the β-carbon atom of the alanine. When β-D-NAL is present in the 1-position and R _{6 is} not Arg, it is preferable in the 6-position to have a hydrophilic D-amino acid residue, such as 4-NH ₂ -D-Phe, 4-guanidino-D-Phe, D- His, D-Lys, D-Om, D-Arg, D-Har (D-Homoarginine) or D-PAL. When Dehydro-Pro is present at the 1-position, a D-isomer of a lipophilic amino acid, such as D-Trp, D-Phe, HCO-D-Trp, NO ₂ -D-Trp, D-Leu, is preferably in the 6-position. D-IIe, D-NIe, D-Tyr, D-VaI, D-Ala, D-Ser (OtBu), β-D-NAL or ( ⁱⁿ Bzl) D-His, but D-PAL can also be used become. In addition, a substitution of GIy by D-Ala in the 10-position, as well as other mentioned substitutions.

The peptides of the invention are often administered in the form of pharmaceutically acceptable non-toxic salts, such as those described in U.S. Pat. As salts with acids or metal complexes, for. With zinc, barium, calcium, magnesium, aluminum, etc. (these are considered to be addition salts in the teaching of the invention), or a combination of both types. Examples of such salts with acids are hydrochloride, hydrobromide, sulfate, phosphate, nitrate, oxalate, fumarate, gluconate, tannate, meleate, acetate, citrate, benzoate, succinate, alginate, malate, ascorbate, tartrate, etc. For example, an aqueous solution of the Peptides are treated several times with 1 N acetic acid and then lyophilized to give the acetate of the peptide. When the active ingredient is to be administered in tablet form, the tablet may contain a pharmaceutically acceptable diluent, which may also be or contain a binder, such as e.g. B. tragacanth, corn starch or geiantine; a release agent such as alginic acid and a lubricant such as magnesium stearate. When desired to be administered in liquid form, a sweetener and / or flavoring agent may be included in the pharmaceutically acceptable diluent; Intravenous administration may be in isotonic saline, phosphate buffer, etc.

The pharmaceutical preparation is usually intended to contain the peptide in association with a conventional pharmaceutically acceptable carrier. Usually, when intravenously administered, the dose should be from about 1 to 10 C-1 g of peptide per kg of body weight; oral doses may be higher. Generally, treatment with these peptides will be in the same manner as clinical treatment with other GnRH antagonists.

These peptides may be administered to mammals intravenously, subcutaneously, intramuscularly, orally, percutaneously (eg, intranasally or intravaginally) to achieve inhibition and / or control of fertility, as well as for reversible suppression of gonadal activity, e.g. As for the treatment of prematurity or during radio or chemotherapy. Effective doses vary with the mode of administration and the type of mammal being treated. An example of a typical dosage is a bacteriostatic aqueous solution containing the peptide, which is administered to achieve a dose of 0.1 to 2.5 mg / kg of body weight. Oral administration of the peptide may be in both solid and liquid form.

Although the invention has been described in terms of its preferred embodiments, also changes and modifications are included. For example, other substituents known in the literature that do not significantly affect the effectiveness of the peptides may be used in the peptides of the invention. The substitution in the phenyl ring of the D-Phe ² radical can also take place in the 3-position or in the 2,4-position, which are considered to be equivalent; Similarly, D-2PAL and D-4PAL are considered to be the equivalent of D-3PAL. At the C-terminus, Pro ⁹ may be attached to one of the following structures generally considered to be its equivalents: Gly-OCH ₃ , Gly-OCH ₂ CH ₃ , Sar-NH _2, or NH-Y 'with Y' = lower alkyl , especially ethyl, cycloalkyl, fluoro-substituted lower alkyl or NHCONHQ with Q = H or lower alkyl. Sar means S'arcosin.

The peptides produced by the method of the invention strongly inhibit the secretion of gonadotropins by the pituitary gland of mammals including humans and / or the release of steroids by the gonads. There are reasons to want to prevent the ovulation of female mammals. The administration of GnRH analogs, which are antagonists of normal GnRH function, can be used to suppress or delay ovulation. For this reason, GnRH analogs, which are GnRH antagonists, are used as contraceptives or to regulate the periods of conception. GnRH antagonists can also be used to treat prematurity and endometriosis. These antagonists have also been shown to be useful for regulating the release of gonadotropins in male mammals and may be useful for cessation of spermatogenesis, i. H. used as male contraceptives, and for the treatment of prostate hypertrophy.

The peptides are very effective in inhibiting LH release and are therefore often referred to as GnRH antagonists. When administered at very low doses during proestrus, the peptides inhibit the ovulation of female mammals and, when administered shortly after conception, also cause the resorption of fertilized eggs. Peptides are also effective in the contraceptive treatment of male mammals.

When administered around noon on the day of the proestrus, the peptides of the invention are effective in preventing doses of less than 100μg per kg of body weight in preventing ovulation of female rats. For longer suppression of ovulation, it may be necessary to use dosages of about 0.1 to about 2.5 mg per kg of body weight. These antagonists also cause an interruption of spermatogenesis when administered in a regular manner to male mammals, and thus can be used as contraceptives. Because these compounds lower testosterone levels (an undesirable consequence in the normal, sexually active male), it is advantageous to administer replacement doses of testosterone with the GnRH antagonist. These antagonists can also be used to regulate the production of gonadotropins and sex steroids for purposes other than those listed above.

embodiment example 1

Peptides, as indicated in Table 1, having the formula Ac-Ri- (4F) D-Phe-R ₃ -Ser-Tyr-R ₆ -Leu-Arg-Pro-Ri ₀ are prepared by the solid-phase method given above.

Table I

Ri R ₃ R6 Rio

1 ß-D-2NAL (6NO ₂ ) D-Trp D-Arg Gly-NH ₂ 2 ß-D-2NAL (6NO ₂ ) D-Trp D-Arg D-Ala-NH ₂ 3 Dehydro-Pro (6NO ₂ ) D-Trp ß-D-2NAL Gly-NH ₂ 4 ß-D-2NAL (6NH ₂ ) D-Trp D-Arg Gly-NH ₂ 5 ß-D-2NAL (5OCH ₃ ) D-Trp D-Arg Gly-NH ₂ 6 ß-D-2NAL (5Br) D-Trp D-Arg Gly-NH ₂ 7 ß-D-2NAL (5F) D-Trp D-Arg Gly-NH ₂ 8th ß-D-2NAL (5Cl) D-Trp D-Arg Gly-NH ₂ 9 ß-D-2NAL (5CH ₃ ) D Trp D-Arg Gly-NH ₂ 10 ß-D-2NAL (N ⁱⁿ HCO) D-Trp D-Arg Gly-NH ₂ 11 ß-D-2NAL (5F) D-Trp (4-gua) D-Phe Gly-NH ₂ 12 ß-D-2NAL (5 Cl), D-Trp D-His Gly-NH ₂ 13 Dehydro-Pro (6NO ₂ ) D-Trp D-Leu NHCH ₂ CH ₃ 14 D-Trp (5F) D-Trp D-Phe NHCH ₂ CH ₃ 15 D-pGlu (5F) D-Trp D-IIe D-Ala-NH ₂ 16 D-Phe (6NO ₂ ) D-Trp D-Val NHNHCONH ₂

As an example, a representative solid-phase synthesis of the above peptide No. 1, [Ac-β-D-2NAL ¹ , (4F) D-Phe ² , will be described below.

(6NO ₂ ) D-Trp ³ , D-Arg ⁶ ] -GnRH. This peptide has the following formula:

Ac-β-D-2-NAL- (4F) D-Phe- (6NO ₂ ) D-Trp-Ser-Tyr-D-Arg-Leu-Arg-Pro-Gly-NH ₂ . The other peptides will work the same way

shown and cleaned.

Boc-protected Gly is coupled to a BHA resin using a threefold excess in CH ₂ Cl ₂ and in the presence of DCC as the activating reagent within 2 h. The glycine residue is linked to the BHA residue via an amine linkage

bound.

Subsequent coupling of each amino acid residue, washing, deblocking, and coupling of the next amino acid residue is done by the following schedule with an automatic machine and starts with about 5 g of resin.

step

Reagents and operations

Mixing time in min.

Washing with 80 ml CH ₂ Cl ₂ (2 χ)

Wash with 30 ml of methanol (2 χ)

Washing with 80 ml CH ₂ Cl ₂ (3 χ)

70 ml 50%. TFA with 5% ethane-1,2-dithiol in CH ₂ Cl ₂ (2 χ)

Wash with 80 ml of isopropyl alcohol with 1% ethane-1,2-dithiol (2 χ)

70 ml 12.5% TEA in CH ₂ CI ₂ (2 x)

Wash with 40 ml MeOH (2 x)

Wash with 80 ml CH ₂ Cl ₂ (3x)

Boc-amino acid (1 OmMoI) in 30 ml of DMF or CH ₂ Cl ₂ , depending on the solubility of the

corresponding protected amino acid,

and DCC (10mMoI) in CH ₂ Cl ₂ (I x)

Wash with 40 ml MeOH (2 x)

70 ml 12.5%. TEA in CH ₂ Cl ₂ (I x)

Wash with 30 ml MeOH (2 x)

Wash with 80 ml CH ₂ CL ₂ (2 χ)

3 3 3 10 3 5 2 3

30-300 3 3 3 3

After step 13, a ninhydrin test can be performed on a sample. If the sample is negative, step 1 can proceed to coupling the next amino acid; if the sample is positive, or slightly negative, the levels become 9 to 13

repeated.

The above scheme is used for the coupling of each amino acid of the peptide prepared according to the invention after the first amino acid has been attached. Throughout the synthesis, N ^a -Boc is used as a protecting group for all remaining amino acids. N ^a -Boc-β-D-2NAL is prepared in a known manner, e.g. B. according to the teaching of the US patent

4234571. The side chain of Arg is protected with Tos. The protective group for the hydroxy group of Ser is OBzI. (6NO ₂ ) D-Trp remains unprotected. The last amino acid introduced is N Boc-β-D-NAL. Boc-Arg (Tos) and Boc- (6N0 ₂ ) D-Trp, the small ones

Solubility in CH ₂ CI ₂ have be coupled ₂ mixtures in DMF-CH ₂ CI.

After blocking the a-amino group at the N-terminus, acetylation succeeds with a large excess of acetic anhydride in dichloromethane. The cleavage of the peptide from the resin and the complete elimination of the protective groups on the side chains is very easy at 0 ° C with HF. Anisole is added as a rinse before treatment with HF. After removal of the HF in vacuo, the resin is extracted with 50% acetic acid and the wash is lyophilized to give a crude

Peptide powder.

Purification of the peptide is then carried out by ion exchange chromatography on carboxymethylcellulose (Whatman CM with a gradient of 0.05 to 0.3 mol of NH ₄ OAc in a 50:50 methanol-water mixture) followed by vertex chromatography in a gel filtration column with the elution system butane. i-ol / 0.1 N acetic acid (1: 1 by volume).

The peptide is considered to be homogeneous by thin layer chromatography with various solvent systems and reverse phase high pressure liquid chromatography and aqueous triethylammonium phosphate plus acetonitrile. The amino acid analysis of the resulting purified peptide corresponds to the formula for the depicted structures and provides integer values for each amino acid in the chain. The optical rotation is carried out on a photoelectric polarimeter

[α] "= -31.8 ± 1 ⁰ C ⁰ C (c = 1.50% ig. acetic acid) were measured.

The peptides are tested in vivo and can also be tested in vitro. In vitro assays are performed on dissociated rat pituitary cells maintained in culture for 4 days prior to testing. The LH level obtained in response to the administration of the peptides is determined by specific radioimmunoassay for rat LH. The control dishes of cells receive only one dose that is 3 nanomolar to GnRH; the test dishes are given a 3nanomolar dose of GnRH 'and either one dose of the current standard antagonist [Ac-dehydro-Pro ¹ , (4F) D-Phe ² , D-Trp ^3l6 ] -GnRH for comparison purposes or the test peptide in concentrations of 0, 01 to 10 nanomolar. The amount of LH released in the GnRH treated samples is compared to that in the peptide and GnRH treated samples. The potency of the test peptide to reduce the amount of LH replaced by 3nanomolar GnRH is compared to that of the current standard peptide. The in vivo tests determine efficacy for preventing ovulation in female rats. In this test, a certain number of mature female Sprague-Dawley rats, namely 6, each weighing 225-250 g, are injected with a given microgram dosage of the peptide in corn oil at noon on the day of the proestrus. Proöstrus is the afternoon of ovulation. Another group of female rats to which the peptide is not administered serves as a control group. In each of the control rats ovulation occurred on the evening of the proestrus; in the treated rats, the number of ovulations is registered. Each of the peptides is considered to be significantly effective in preventing ovulation of female rats at a very low dose, and each peptide is considered to be absolutely effective at a dose of about 5μ9. The results of further tests at lower dosages are given in Table A below.

Table A

Peptide No. in vivo

Dose ^ g) number of ovulations

1.1 0/5

C 0.5 4/14

2 1 1/10

3 2.5 2/10 2 0/6

4 1 5/14

5 1 7/10

6 2 > 0/10 1 6/18

7 5 2/10 1 3/9

8 1 7/14 0,5 2/7

9 2.5 0/10 1 10716

10 1 0/10

0.5 9/17

All of the peptides listed in Table I are considered to be effective for blocking GnRH-induced LH release in vitro at an appropriate concentration. Many of the peptides are much more potent in vivo than the current standard. All peptides are considered to be effective in preventing ovulation in female mammals at very low doses, and some selected peptides are considered to be at least twice as effective as any previously known and tested GnRH antagonists.

Example Il

According to the above-described solid phase method, the peptides indicated in Table II having the formula Ac-β-D-2NAL- (W) D-Phe-R ₃ -R 4 -R 5 -D-Arg-R ₇ -Arg-Pro-D- Ala-NH2 shown.

Table II W R ₃ R ₄ R ₅ R ₇ 4br (6 NO ₂ ) D-Trp Ser Type Never 17 4br (6 NO ₂ ) D-Trp Ser (2F) Phe Leu 18 4cl (1 HCO) D-Trp Ser Tyr Nva 19 4cl (1 HC0) D-Trp Ser Tyr Leu 20 4cl (1 HCOID Trp Ser Tyr Never 21 4CI (1 HCO) D-Trp Ser (2CH ₃ ) Phe Leu 22 4cl (1HCO) -D-Trp _x Ser Tyr NML 23 4cl ' (1 HCO) D-Trp Ser Tyr NML (AIa ⁹ ) 24 4cl (1 HCO) D-Trp Ser (2CI) Phe NML 25 4NO ₂ (5CH ₃ ) D Trp Ser (3CH ₃ ) Phe NML ? 6 4NO ₂ (5 F) D Trp Orn Tyr Never 27 2.4Cl ₂ (5 Cl), D-Trp Ser (3F) Phe Never (acetate) 28 2.4Cl ₂ (6NO ₂ ) D-Trp EEL (3F) Phe Nva 29 CMe / 4CI (5 F) D Trp aBu (3l) Tyr NML 30 3,4Cl ₂ (5 F) D Trp Orn (3CI) Phe Leu 31

Testing in vitro and / or in vivo of the peptides listed in Table II shows that the peptides, at appropriate levels, are considered to be effective in blocking GnRH-induced LH release. Many of these peptides are more effective in vivo than the current standard. All peptides are considered to be effective in preventing ovulation in female mammals at very low doses. ,

Example III

According to the above-described solid phase method, the peprides given in Table III with the formula X-β-D-2NAL- (4CI) D-Phe- (6NO ₂ ) D-Trp-Ser-R ₅ -R ₆ -NML- Arg-Pro-R ₁₀ shown.

Table III X - R ₅ R ₆ Rio ac Tyr D-Arg Gly-NH ₂ 32 acr bad D-Tyr D-Ala-NH ₂ 33 HCO bad D-Tyr NHCH ₂ CH ₃ 34 Bz (3F) Phe D-Arg NHCH ₃ 35 ac (2F) Phe D-Lys NHCF ₃ 36 Vac (2CI) Phe D-Har NHCH ₂ CH ₂ CH ₃ 37 acr (3CI) Phe (4gua) D-Phe NHCF ₂ CF ₃ 38 ac (3F) Phe Mandrel D-Ala-NH ₂ 39 acr (3) Tyr D-His D-Ala-NH ₂ 40 ac Tyr D-Arg D-Ala-NH ₂ (Pro ⁸ , Arg ⁹ ) 41 ac (3CI) Phe D-Arg Gly-NH ₂ 42 Vac (3CI) Phe (4NH ₂ ) D-Phe NHNHCONH ₂ 43 Bz (3CI) Phe (4NH ₂ ) D-Phe NHNHCONHCh ₃ 44

Testing of the peptides shown in Table III in vivo and / or in vitro shows that these peptides are considered to be effective in blocking GnRH-induced LH release at appropriate concentration. Many of these peptides are more effective in vivo than the current standard. All peptides are considered to be effective in preventing ovulation in female mammals at very low doses.

Example IV

According to the solid phase method described above, the peptides shown in Table IV are represented by the formula Ac-R _r 4CI-D-Phe-D-Trp-Ser-Tyr-R ₆ -R 7 -R-Pro-R ₁₀ -NH ₂ .

Table IV

45 46 47 48 49 50 51 52 53 54 55 56 57 58 59

β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β- D-2 NAL Dehydro-Pro β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL Dehydro-Pro

D-Arg D-Arg D-Arg D-Arg D-Arg D-Arg D-Arg D-Arg D-Arg D-Trp D-Arg D-Har D-Lys D-Arg D-Leu

Never Met Tyr Never Met Tyr Phe 4F-D-Phe Cys Never Trp Cys Nva 3PAL Tyr

D-Ala

D-Ala

D-Ala

Gly

Gly

Gly

D-Ala

D-Ala

D-Ala

D-Ala

D-Ala

D-Ala

D-Ala

D-Ala

NHCH ₂ CH ₃ ¹ '

¹¹ R ₁₀ -NH ₂

Peptide # 45, referred to as [Ac-β-D-2 NAL ¹ , 4CI-D-Phe ² , D-Trp ³ , D-Arg ⁶ , Never ⁷ , D-Ala ¹⁰ J-GnRH the following

Formula:

Ac-.beta.-D-2 NAL-> CI-D-Phe-D-Trp-Ser-Tyr-D-Arg-Nle-Arg-Pro-D-Ala-NHz.

All peptides listed in Table IV are considered to be effective in blocking GnRH-induced LH release, with adequate concentration in vitro. Many of these peptides are much more potent in vivo than the current standard.

All peptides are considered to be effective in preventing ovulation in female mammals at very low doses, and some are considered to be at least as effective as any previously known and tested GnRH antagonists

considered.

Example V

According to the above-described solid phase method, the peptides represented by Table V are represented by the formula Ac-R ₁ - (W) D-Phe-R ₃ -Ser-Tyr-R ₆ -R ₇ -Arg-Pro-D-Ala-NH ₂ shown.

Table V

Ri

R ₆

R ₇

60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75

β-D-2 NAL β-D-2 NAL β-D-2 NAL Dehydro-Pro Dehydro-Pro β-D-2 NAL β-D-2 NAL Dehydro-Pro Dehydro-Pro Dehydro-Pro Dehydro-Pro Dehydro- Proβ-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL

4br

4F

4F

4F

4NO ₂

4cl

4cl

4br

4br

C ° Me / 4CI

C ^a Me / 4CI

4cl

4NO ₂

4NO ₂

3,4Cl ₂

4cl

(6NO ₂ ) D-Trp D-Arg Never (6NO ₂ ) D-Trp D-His Tyr D-Trp 4-gua-D-Phe Tyr D-Trp D-Trp Phe β-D-1 NAL D-Val mead N ⁱⁿ HCO-D-Trp D-Arg Never N _in HCO-D-Trp D-Arg Nva β-D-2 NAL D-Tyr 4F-D-Phe β-D-2 NAL D-Nle Trp β-D-2 NAL D-Phe Never D-PAL β-D-2 NAL Phe D-PAL β-D-2 NAL Leu D-PAL D-O rn 4F-D-Phe P-PAL 4NH ₂ -D-Phe mead D-Trp 4NH ₂ -D-Phe Tyr D-PAL D-PAL Leu

Testing the peptides indicated in Table V in vitro and / or in vivo shows that, with adequate concentration in vitro, they are effective in blocking the GnRH-induced release of LH. Many of these peptides are more effective in vivo than the current standard. All peptides are considered to be effective in preventing ovulation in female mammals at very low doses. ,

Example Vl

After the above solid phase method, the peptides of the formula given in Table VI are XR _r 4F-D-Phe-D-Trp-Ser-Tyr-R ₆ -R ₇ -Arg-Pro-D-Ala-NH _{2 is} shown. ,

Table VI X acr 76 acr 77 acr 78 acr 79 H 80 Bz 81 Bz 82 HCO 83 HCO 84 Vac 85 Vac 86 H 87

Dehydro-Pro D-Trp Tyr Dehydro-Pro D-IIe PAL Dehydro-Pro D-Val Nle (3F-Phe ⁵ ) Per D-Ser (OtBu) Phe Dehydro-Pro (ImBzl) D-His Cys (0rn ⁴ ) D-Phe D-Trp mead D-pGlu D-Trp Nile (3l-Tyr ⁵ ) β-D-1 NAL D-Arg Phe (acetate) Dehydro-Pro D-Har Tyr (aBu ⁴ ) ß-D-2NAL D-Lys Nva (acetate) D-Phe D-Nle Cys Dehydro-Pro D-Ala Trp

Testing the peptides indicated in Table VI in vitro and / or in vivo indicates that these peptides, when appropriately concentrated in vitro, are considered to be effective in blocking GnRH-induced LH release. Many of these peptides are more effective in vivo than the current standard. All peptides are considered to be effective in preventing ovulation in female mammals at very low doses.

Example VII

According to the solid-phase method described above, the peptides of the formula Ac-Ri- (W) D-Phe-R ₃ -R ₄ -Tyr-R ₆ -Leu-Arg-Pro-D-Ala-NH ₂ are shown in Table VII.

Table VII Ri W R ₃ R ₄ R ₆ Dehydro-Pro 4cl (6NO ₂ ) D-Trp Orn β-D-2 NAL 88 Dehydro-Pro 4F (6N0 ₂ ) D-Trp Orn D-Val 89 Dehydro-Pro 4F (6 F) D Trp EEL 4gua-D-Phe 90 Dehydro-Pro 4F (6 F) D Trp EEL Mandrel 91 Dehydro-Pro 4NO ₂ (5OCH ₃ ) D-Trp EEL D-Lys 92 Dehydro-Pro 4NO ₂ (5OCH ₃ ) D-Trp EEL D-PAL 93 β-D-2 NAL 4NO ₂ 1 Ac-D Trp Ser D-Har 94 β-D-2 NAL 4NO ₂ 1 HCO-D Trp Ser D-Trp 95 Dehydro-Pro 4NO ₂ (6Br) D-Trp aBu D-Nle 96 Dehydro-Pro C ^a Me / 4CI (6 Br) D-Trp aBu D-Leu 97 Dehydro-Pro C ^a Me / 4CI (6CH ₃ ) D Trp aBu β-D-2 NAL 98 Dehydro-Pro 4br (6NH ₂ ) D-Trp Orn 4NH ₂ -D-Phe 99 Dehydro-Pro 3,4Cl ₂ (5NH ₂ ) D-Trp Om D-Lys 100

Testing of the peptides indicated in Table VII in vitro and / or in vivo indicates that these peptides, when appropriately concentrated in vitro, are considered to be effective in blocking GnRH-induced LH release. Many of these peptides are more effective in vivo than the current standard. All peptides are considered to be effective in preventing ovulation in female mammals at very low doses.

Testing in vivo is performed on different peptides with variable doses. The test results are included in Table B. The optical rotation of the prepared peptides is measured with a photoelectric polarimeter in 50% acetic acid (c = 1) and reported in Table B.

Table B

Peptide no.

[a] g ² dose ^ g)

Number of ovulations in vivo

17 19 20 21 22 23 25 25 45

46 47 51

52

53 55 58

88

-28, O ⁰ C -22.4 ° C -18.8 ⁰ C -22.5 ⁰ C

-23.8

-22.0 -17.2

1.10

2.7

6.7

4.11

7.9

1.5

5.6

1.5

3/16

9.10

2.15

3.9

0/10

4.7

0/8

6.15

2.3

0/10

4.7

6.8

0/8

0/10

2/10 ~

0/10

6.9

Example VIII

After the above solid phase method, the peptide of the formula Ac-D-3PAL-RRR2-Ser-Arg-R shown in Table VIII are shown -R7 _{6-Arg-Pro-Rio.}

Table VIII

101 102 103 104

105 106 107 108 109 110 111 112 .113 114 115 116 117 118 119 120 121 122 123 124 125

β-D-2 NAL β-D-2NAL β-D-2 NAL β-D-2 NAL

Dehydro-Pro Dehydro-Proβ-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β-D-2 NAL β- D-2 NAL Pro

Dehydro-Pro Dehydro-Pro Dehydro-Pro Dehydro-Pro Dehydro-Pro Dehydro-Pro Pro

β-D-2 NAL β-D-2 NAL 4CI-D-Phe

4CI-D-Phe D-Trp Leu 4CI-D-Phe D-3PAL Leu 4-CI-D-Phe β-D-2 NAL Leu 4CI-D-Phe D-3PAL Leu 4CI-D-Phe β-D-2 NAL 3PAL 4CI-D-Phe β-D-2 NAL Tyr 4CI-D-Phe D-3PAL NML 4CI-D-Phe β-D-2 NAL 3PAL 4CI-D-Phe (ImBzl) D-His Leu 4CI-D-Phe 6 NO ₂ -D Trp Leu 4CI-D-Phe D-Tyr Leu 4CI-D-Phe (HCO) D-Trp Leu 4 NO ₂ -D-Phe D-Trp NML 4br-D-Phe D-Tyr Never 4br-D-Phe (ImBzl) D-His mead 4br-D-Phe D-Trp Nva C ^a Me / 4CI-D-Phe D-Trp Nva 4F-D-Phe D-Trp 4F-Phe 4F-D-Phe D-Trp NML 4F-D-Phe D-Trp Never 4F-D-Phe β-D-2 NAL Trp 4F-D-Phe β-D-2 NAL Nva 3,4CI ₂ -D-Phe .. β-D-1 NAL Tyr 3,4CI ₂ -D-Phe D-Trp mead 3,4CI ₂ -D-Phe D-Tyr 3PAL

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

(4guaPhe ⁵ )

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

NHCH ₂ CH ₃

NHCH ₂ CH ₂

NHNHCONH ₂

NHCH ₂ CH ₃

D-Ala-NH ₂

NHNHCONH ₂

NHCH ₂ CH ₃

Gly-NH ₂

Gly-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

D-Ala-NH ₂

(acetate)

The peptides described in Table VIII are tested in vivo to determine their effectiveness in preventing ovulation in female rats. All are considered to be effective in preventing ovulation of female rats at a very low dose and are fully effective at a dose of about 10 μg. Some of these peptides were also tested specifically at lower doses. The results are listed in Table C below.

Table C

Peptide no. [Α] "

101 -30.3 ⁰ C ± 1

in vivo Dose (μρ) Number of ovulations 1 0/6 0.5 2.10 1 1.16 0.5 5.10 1 0/5 0.5 6.10 0.25 10.7 1 9.10 0.5 0/6 0.5 0/5

Claims (10)

Invention claim: 1. A process for the preparation of a peptide or one of its non-toxic salts having antagonistic properties to gonadotropic hormones, characterized in that compounds of the formula -X-Ri- (W) -D-Phe-R ₃ -R ₄ -R ₅ -R ₆ -R ₇ -Arg-Pro-Rio, wherein X is a hydrogen atom or an acyl group having a maximum of 7 carbon atoms; Ri Dehydro-Pro, D-pGlu, D-Phe, 4CI-D-Phe, D-Trp, Pro or β-D-NAL; W is Cl, F, NO ₂ , C ^a Me / 4 Cl, Cl ₂ or Br; R ₃ D-3PAL, β-D-NAL or (Y) D-Trp with Y = H, NO ₂ , NH ₂ , OCH ₃ , F, Cl, Br, CH ₃ , N ⁱⁿ HCO or N ⁱⁿ Ac; R _{4 is} Ser, Orn, AAL or aBu; R _{5 is} Tyr, Arg, (3F) Phe, (2F) Phe, (3I) Tyr, (3CH ₃ ) Phe, (2CH ₃ ) Phe, (3Cl) Phe or (2CI) Phe; R _{6 is} a D-isomer of a lipophilic amino acid or 4-Nh ₂ -D-Phe, D-Lys, D-Har, D-Orn, D-His, 4-gua-D-Phe, D-PAL or D-Arg ; R ₇ Never, Leu, NML, Phe, Met, Nva, Tyr, Trp, Cys, PAL or 4F-D-Phe and R ₁₀ Gly-NH ₂ , D-Ala-NH ₂ , NH-Y 'with Y' = niederesAlky ^ CycloalkyUiuorsubstituiertesniederesAlkyl.oderNHCONH-QmitQ = H or lower alkyl, but when R ₃ is D-3PAL, _R5 is Arg, and when R ₃ is ß-D-NAL or D-Trp, _R7 is Tyr, PAL, Phe , or 4F-D-Phe from an intermediate of the formula X ^ RHWJD-Phe-R ^ lR ^^ iX ⁴ or X ⁵ ) -R ₆ (X ⁵ ) -R ₇ (X ⁶ ) -Arg (X ⁵ ) -Pro -X ⁷ , wherein X ^{1 is} hydrogen or an a-amino protecting group; X ^{2 is} hydrogen or a protecting group for the indole nitrogen; X ^{3 is} hydrogen or a protective group for the alcoholic hydroxy group of Ser or for an amino group of the side chain; X ^{4 is} hydrogen or a protecting group for the phenolic hydroxy group of Tyr; X ⁵ and X ^{6 are} each either hydrogen or a protecting group for the particular side chain present; and X ^{7 is} Gly-O-CH ₂ - (carrier resin), O-CH ₂ - (carrier resin), D-Ala-O-CH ₂ - (carrier resin), Gly-NH ₂ - (carrier resin), D-Ala-NH - (Carrier resin), GIy NH ₂ , an ester, an amide or a hydrazide by splitting off one or more of the groups X ¹ to X ⁶ and / or cleavage of a carrier resin contained in X ⁷ with hydrofluoric acid at 0 ⁰ C with addition are prepared by anisole, which are optionally converted into one of their non-toxic salts. 2. The method according to item 1, characterized in that that means W4CL or 4F. 3. The method according to item 1 or 2, characterized in that Ri ß-D-2NAL and R ₆ 4-NH ₂ -D-Phe, D-Lys, D-Om, D-Har, D-His, 4-gua Mean -D-Phe, D-PAL or D-Arg. 4. The method according to item 3, characterized in that R _{6 is} D-Arg. 5. The method according to item 3, characterized in that R _{7 is} Tyr or 3PAL. 6. The method according to any one of items 1 to 5, characterized in that R _{3 is} (Y) D-Trp and Y6NO ₂ or N ^ln CH0. 7. The method according to item 1 or 2, characterized in that Ri ß-D-2NAL, R _{3 is} D-PAL and R _{5 is} Arg. 8. The method according to item 7, characterized in that R _{6 is} D-Trp, D-PAL, ß-D-NAL, (in BzI) D-HiS or (6NO ₂ ) -D-Trp. 9. The method according to item 8, characterized in that R ₆ (imbzl) is D-His or (6NO ₂ ) D-Trp and R _{7 is} Leu. 10. The method according to any one of items 1 to 9, characterized in that R _{4 is} Ser, X is Ac or Acr and Ri is ₀ D-Ala-NH ₂ GIy-NH ₂ .