IE911387A1 - Peptides with a bradykinin-antagonistic action - Google Patents

Abstract

Peptides of the formula I A-B-C-E-F-K-(D)-Tic-G-M-F'-I (I> in which A is hydrogen, alkyl, alkanoyl, alkoxycarbonyl, alkylsulphonyl, cycloalkyl, aryl, arylsulphonyl, heteroaryl or an amino acid, each of which can optionally be substituted, B is a basic amino acid, C is a di- or tripeptide, E is the residue of an aliphatic or alicyclic-aliphatic amino acid, F is, independently of one another, an amino acid which is optionally substituted in the side chain or is a direct bond, G is an amino acid, F' is defined as F, can be -NH-(CH2)2-8 or a direct bond, I is -OH, -NH2 or -NHC2H5, and K is a radical -NH-(CH2)1-4-CO-, or is a direct bond, have a bradykinin-antagonistic action. Their therapeutic uses comprise all pathological states which are mediated, induced or assisted by bradykinin and bradykinin-related peptides. The peptides of the formula I are prepared by known methods of peptide synthesis.

Description

HOECHST AKTIENGESELLSCHAFT HOE 90/F 131 Dr.WI/je Description Peptides with a bradykinin-antagonistic action The invention relates to novel peptides with a brady5 kinin-antagonistic action and to a process for the preparation thereof.

Bradykinin-antagonistic peptides are described in WO 86/07263, where, inter alia, L-Pro in position 7 of the peptide hormone bradykinin or other bradykinin analogs is replaced by a D-amino acid such as D-Phe, D-Thia, D-Pal, CDF, D-Nal, MDY, D-Phg, D-His, D-Trp, D-Tyr, D-hPhe, D-Val, D-Ala, D-His, D-Ile, D-Leu and DOMT.

The object of the invention is to find novel active peptides with a bradykinin-antagonistic action.

This object is achieved by the peptides of the formula I A-B-C-E-F-K-(D)-Tic-G-M-F'-I (I) in which A is ax) hydrogen, (Cj-Ce)-alkyl, (Ci-Cg) -alkanoyl, (ci_ce)-alkoxycarbonyl or (Ci-C8) -alkylsulfonyl, in each of which 1, 2 or 3 hydrogen atoms are optionally replaced by 1, 2 or identical or different radicals from the series comprising carboxyl, amino, (Ci-CJ-alkyl, IE 911387 (Ci-C*) -alkylamino, hydroxyl, (Ci-C*)-alkoxy, halogen, di- (Ci-CJ -alkylamino, carbamoyl, sulfamoyl, (CX-CA)-alkoxycarbonyl, (C6-C12)-aryl and (C6-C12)-aryl-(Ci-Cj)-alkyl, or in each of which 1 hydrogen atom is optionally replaced by a radical from the series comprising (C3-C8) -cycloalkyl, (Ci-CJ-alkylsulfonyl, (Ci-C*) -alkylsulf inyl, (C6-C12) -aryl- (Cj-C^) -alkylsulfonyl, (c6-ci2) -aryl- (Cj-C*) -alkylsulf inyl, (C6-C12)-aryloxy, (C3-C8)-heteroaryl and (C3—C0) -heteroaryloxy and or 2 hydrogen atoms are replaced by 1 or 2 identical or different radicals from the series comprising carboxyl, amino, (Cx-C4) -alkylamino, hydroxyl, (Ci-C*)-alkoxy, halogen, di- (Ci-C*) -alkylamino, carbamoyl, sulfamoyl, (Ci-C*)-alkoxycarbonyl, (C5-C12)-aryl and (C6-C12) -aryl- (C^-Cs) -alkyl, a2) (C3-C8)-cycloalkyl, carbamoyl which can optionally be substituted - 3 10 on the nitrogen by (Ci-Cg)-alkyl or (C6-C12)aryl, (C6-C12)-aryl, (C7-C13)-aroyl, (C6-C12) -arylsul fonyl, (C3-C8) -heteroaryl or (C3-C8) -heteroaroyl, where in each of the radicals defined under ax) and a2) aryl, heteroaryl, aroyl, arylsulfonyl and heteroaroyl is optionally substituted by 1, 2, 3 or 4 identical or different radicals from the series comprising carboxyl, amino, nitro, (Ci-C4) -alkylamino, hydroxyl, (Ci-CJ-alkyl, (Ci-CJ-alkoxy, halogen, cyano, di- (Ci-C4) -alkylamino, carbamoyl, sulfamoyl and (Ci-C4) -alkoxycarbonyl or a3) a radical of the formula II (II) in which R1 R2 R3 is defined as A under ax) or a2), is hydrogen or methyl, is hydrogen or (Ci-C6)-alkyl, preferably (Cx-C4)-alkyl, which is optionally monosubstituted by amino, substituted amino, hydroxyl, carboxyl, carbamoyl, guanidino, substituted guanidino, ureido, mercapto, methylmercapto, phenyl, 4-chlorophenyl, 4-fluoropheny1, 4-nitrophenyl, 4-methoxyphenyl, 4-hydroxyphenyl, phthalimido, 4-imidazolyl, 3-indolyl, 2- thienyl, 3- thienyl, 2- pyridyl, 3- pyridyl or cyclohexyl, where substituted amino is -NH-A and substituted guanidino is -NH-C(NH)-NH-A in which A is as defined under ax) or a2); B is a basic amino acid which is in the L or D configuration and can be substituted in the side chain; C is a linkage of the formula Ilia or Illb G'-G'-Gly (Ilia) G'-NH-(CH2) n-CO (Illb) in which G' is, independently of one another, a radical of the formula IV (IV) - 5 R4 R5 0 ι I fl - N - CH - C in which R* and R5 form, together with the atoms carrying them, a heterocyclic mono-, bi- or tricyclic ring system with 2 to 15 carbon atoms, and n is 2 to 8; is the residue of a neutral, acidic or basic aliphatic or alicyclic-aliphatic amino acid; is, independently of one another, the residue of a neutral, acidic or basic, aliphatic or aromatic amino acid which can be substituted in the side chain, or is a direct bond; (D)-Tic is the radical of the formula V; H 0 ccr (V) G is defined as G' above or is a direct bond; F' is defined as F, is a radical -NH-(CH2)n- with n = 2 to 8, or can be a direct bond if G is not a direct bond, and I is -OH, -NH2 or -NHC2H5, K is the radical -NH-(CH2) X-CO- with x = 1-4 or is a direct bond, and M is defined as F, and the physiologically tolerated salts thereof.

Unless stated otherwise, the abbreviation of an amino 25 acid residue without a stereodescriptor represents the residue in the L form (cf. Schrdder, Liibke, The Peptides, Volume I, New York 1965, pages XXII-XXIII; Houben-Weyl, - 6 Methoden der Organischen Chemie (Methods of Organic Chemistry), Volume XV/1 and 2, Stuttgart 1974), such as, for example, Aad, Abu, 7 Abu, ABz, 2ABz, eAca, Ach, Acp, Adpd, Ahb, 5 Aib, 0Aib, Ala, BAla, AAla, Alg, All, Ama, Amt, Ape, Apm, Apr, Arg, Asn, Asp, Asu, Aze, Azi, Bai, Bph, Can, Cit, Cys, Cyta, Daad, Dab, Dadd, Dap, Dapm, Dasu, Djen, Dpa, Dtc, Fel, Gin, Glu, Gly, Guv, hAla, hArg, hCys, hGln, hGlu, His, hlle, hLeu, hLys, hMet, hPhe, hPro, hSer, hThr, hTrp, hTyr, Hyl, Hyp, 3Hyp, lie, Ise, Iva, Kyn, Lant, Lcn, Leu, Lsg, Lys, 0Lys, ALys, Met, Mim, Min, nArg, Nle, Nva, Oly, Orn, Pan, Pec, Pen, Phe, Phg, Pic, Pro, APro, Pse, Pya, Pyr, Pza, Qin, Ros, Sar, Sec, Sem, Ser, Thi, 0Thi, Thr, Thy, Thx, Tia, Tie, Tly, Trp, Trta, Tyr, Val.

Suitable as radical of a heterocyclic ring system of the formula IV are, in particular, radicals of heterocycles from the following group: pyrrolidine-2-carboxylic acid; piperidine-2-carboxylic acid; l,2,3,4-tetrahydroisoquinoline-3-carboxylic acid; decahydroisoquinoline-3-carboxylic acid; octahydroindole2-carboxylic acid; decahydroquinoline-2-carboxylic acid; octahydrocyclopenta[b]pyrrole-2-carboxylic acid; 2-azabicyclo[2.2.2]octane-3-carboxylic acid; 2-azabicyclo25 [2.2.l]heptane-3-carboxylic acid; 2-azabicyclo[3.1.0]hexane-3-carboxylic acid; 2-azaspiro[4.4]nonane-3carboxylic acid; 2-azaspiro[4.5]decane-3-carboxylic acid; spiro[(bicyclo[2.2.1]heptane )-2,3-pyrrolidine-5carboxylic acid]; spiro[(bicyclo[2.2.2]octane)-2,330 pyrrolidine-5-carboxylic acid]; 2-azatricyclo[4.3.0. l6 8]decane-3-carboxylic acid; decahydrocyclohepta[b]pyrrole2-carboxylic acid; decahydrocycloocta[b]pyrrole-2carboxylic acid; octahydrocyclopenta[c]pyrrole-2carboxylic acid; octahydroisoindole-l-earboxylic acid; 2,3,3a,4,6a-hexahydrocyclopenta[b]pyrrole-2-carboxylic - 7 acid; 2,3,3a,4,5,7a-hexahydroindole-2-carboxylic acid; tetrahydrothiazole-4-carboxylic acid; isoxazolidine-3carboxylic acid; pyrazolidine-3-carboxylic acid; hydroxyproline-2-carboxylic acid; all of which can optionally be substituted: co The heterocycles which form the basis for the above15 mentioned radicals are disclosed, for example, in US-A 4,344,949, US-A 4,374,847, US-A 4,350,704, EP-A 29 488, EP-A 31 741, EP-A 46 953, EP-A 49 605, EP-A 49 658, EP-A 50 800, EP-A 51 020, EP-A 52 870, EP-A 79 022, EP-A 84 164, EP-A 89 637, EP-A 90 341, EP-A 90 362, EP-A 105 102, EP-A 109 020, EP-A 111 873, EP-A 271 865 and EP-A 344 6i B2.

Unless otherwise stated in the specific case, alkyl can be straight-chain or branched. The corresponding statement applies to radicals derived therefrom, such as alkoxy, aralkyl or alkanoyl.

(C6-C12) -Aryl is preferably phenyl, naphthyl or biphenylyl. Radicals derived therefrom, such as aryloxy, aralkyl or aroyl, are to be formulated correspondingly.

Halogen is fluorine, chlorine, bromine or iodine, preferably chlorine.

Particularly suitable salts are alkali metal or alkaline earth metal salts, salts with physiologically tolerated amines and salts with inorganic or organic acids such as, for example, HCl, HBr, H2SO4, H3PO4, maleic acid, fumaric acid, citric acid, tartaric acid and acetic acid.

Preferred peptides of the formula I are those in which B is Arg, Lys, Orn, 2,4-diaminobutyryl or an L-homoarginine residue where, in each case, the amino or the guanidino group of the side chain can be substituted by A as described under a:) or a2); E is the residue of an aliphatic or alicyclic-aliphatic amino acid which is in the L or D configuration and which contains 1 to 14 carbon atoms in the side chain, such as alanine, serine, threonine, 0-(Ci-Cg)-alkyl- or O-(C6-Ci0)-aryl-protected serine or threonine, valine, norvaline, leucine,isoleucine, norleucine, neopentylglycine, tert-butylglycine or (C3-C7) -cycloalkyl- (Ci-C3) -alkylglycine; - 9 F' is the residue of a basic amino acid in the L or D configuration, such as Arg or Lys, where the guanidino group or amino group of the side chain can be substituted by A as described under ax) or a2), or is a radical -NH-(CH2)n - with n = 2 to 8; K is the radical -NH- (CH2)X-CO- with x = 2-4 or is a direct bond.

Particularly preferred peptides of the formula I are those in which B is Arg, Orn or Lys, where the guanidino group or the amino group of the side chain is unsubstituted or can be substituted by (0χ-08)-alkanoyl, (C7-C13)aroyl, (C3-C8)-heteroaroyl, (Cx-C8)-alkylsulfonyl or (06-0χ2)-arylsulfonyl, where the aryl, heteroaryl, aroyl, arylsulfonyl and heteroaroyl radicals can be substituted with, where appropriate, 1, 2, 3 or 4 identical or different radicals as described under a2) ; E is leucine, isoleucine, norleucine, tert-butyl20 glycine, serine, threonine or cyclohexylalanine; K is a direct bond, and M is a direct bond.

Very particularly preferred peptides of the formula I are those in which A is hydrogen, (D)- or (L)-H-Arg, (D)- or (L)-H-Lys or (D) — or (L)-H-Orn, B is Arg, Orn or Lys, where the guanidino group or the amino group in the side chain can be substituted by (Cx-C8)-alkanoyl, (C7-Cx3)-aroyl, (C3-C9)-heteroaroyl, (Cx-C8)-alkylsulfonyl or (C6-Cx2)-arylsulfonyl, where - 10 the aryl, heteroaryl, aroyl, arylsulfonyl and heteroaroyl radicals can optionally be substituted with 1, 2, 3 or 4 identical or different radicals from the series comprising methyl, methoxy and halogen; c is Pro-Pro-Gly, Hyp-Pro-Gly or Pro-Hyp-Gly, E is Leu, Ile, Tbg or Cha, F is Ser, Hser, Lys, Leu, Val, Nle, Ile or Thr K is a direct bond, M is a direct bond, G is the radical of a heterocyclic ring system of the formula IV, where the radicals of the heterocycles pyrrolidine-2-carboxylic acid; piperidine-2carboxylic acid; tetrahydroisoquinoline-3-carboxylic acid, cis- and trans-decahydroisoquinoline-3carboxylic acid; cis-endo-, cis-exo-, trans-octahydroindole-2 -carboxylic acid; cis-endo-, cis-exo-, trans-octahydrocyclopenta[b]pyrrole-2-carboxylic acid or hydroxyproline-2-carboxylic acid are pre20 ferred, F' is Arg and I is OH.

Examples of very particularly preferred peptides of the formula I are: H-(D)-Arg-Arg-Pro-Hyp-Gly-Leu-Ser-(D)-Tic-Oic-Arg-OH H-(D)-Arg-Arg-Pro-Hyp-Gly-Cha-Ser-(D)-Tic-Oic-Arg-OH H-(D)-Arg-Arg-Pro-Hyp-Gly-Tbg-Ser-(D)-Tic-Oic-Arg-OH The invention also relates to a process for preparing peptides of the formula I, which comprises - 11 a) reacting a fragment with a C-terminal free carboxyl group or its activated derivative with a corresponding fragment with an N-terminal free amino group, or b) synthesizing the peptide stepwise, eliminating one or more protective groups temporarily introduced into the compound obtained as in (a) or (b) to protect other functions where appropriate, and converting the compounds of the formula I which have been obtained in this way into their physio10 logically tolerated salt where appropriate.

The peptides of the present invention were prepared by generally known methods of peptide chemistry, see, for example, Houben-Weyl, Methoden der organischen Chemie, Volume 15/2, preferably by solid-phase synthesis as described, for example, by B. Merrifield, J. Am. Chem.

Soc. 85, 2149 (1963) or R.C. Sheppard, Int. J. Peptide Protein Res. 21, 118 (1983) or by equivalent known methods. Used as α-amino protective groups are urethane protective groups such as, for example, the 0 tert-butyloxycarbonyl(Boc) or fluorenylmethy1oxycarbonyl(Fmoc) protective group.

If necessary to prevent side reactions or for synthesizing specific peptides, the functional groups in the side chain of amino acids are additionally protected by suitable protective groups (see, for example, T.W. Greene, Protective Groups in Organic Synthesis), employing primarily Arg(Tos), Arg(Mts), Arg(Mtr), Arg(PMC), Asp(OBzl), Asp(OBut), Cys(4-MeBzl), Cys(Acm), Cys(SBut), Glu(OBzl), Glu(OBut), His(Tos), His(Fmoc), His(Dnp), His(Trt), Lys(Cl-Z), Lys(Boc), Met(O), Ser(Bzl), Ser(But), Thr(Bzl), Thr(But), Trp(Mts), Trp(CHO), Tyr(Br-Z), Tyr(Bzl) or Tyr(But).

The solid-phase synthesis starts at the C-terminal end of the peptide with the coupling of a protected amino acid onto an appropriate resin. Starting materials of this - 12 type can be obtained by linking a protected amino acid, via an ester or amide linkage, to a polystyrene or polyacrylamide resin modified with a chloromethyl, hydroxymethyl, benzhydrylamino(BHA) or methylbenzhydryl5 amino (MBHA) group. The resins used as support material are commercially available. BHA- and MBHA-resins are usually employed when the synthesized peptide is to contain a free carbamoyl group at the C terminus. If the peptide is to contain a secondary carbamoyl group at the C-terminal end, a chloromethyl- or hydroxymethyl-resin is used and the cleavage off is carried out with the appropriate amines. If, for example, it is wished to obtain the ethylamide, the peptide can be cleaved off the resin with ethylamine, in which case the side-chain protective groups are subsequently eliminated by other suitable reagents. If the tert-butyl protective groups in the amino acid side chain are to remain in the peptide, the synthesis is carried out with the Fmoc protective group for temporary blocking of the α-amino group of the amino acid using the method described, for example, by R.C. Sheppard, J. Chem. Soc., Chem. Comm. 1982. 587, in which case the guanidino group of the arginine is protected by protonation with pyridinium perchlorate, and the other amino acids functionalized in the side chain are protected with benzyl protective groups which can be eliminated by catalytic transfer hydrogenation (A. Felix et al. J. Org. Chem. 13, 4194 (1978)) or by sodium in liquid ammonia (W. Roberts, J. Am. Chem. Soc. 76. 6203 (1954)).

After elimination of the amino protective group of the amino acid coupled to the resin using a suitable reagent such as, for example, trifluoroacetic acid in methylene chloride in the case of the Boc protective group or a 20% strength solution of piperidine in dimethylformamide in the case of the Fmoc protective group, the subsequent protected amino acids are coupled on successively in the required sequence. The peptide-resins with N-terminal protection which are the intermediate products are - 13 deblocked by the reagents described above for the linkage to the subsequent amino acid derivative.

It is possible to use as coupling reagent all possible activating reagents used in peptide synthesis, see, for example, Houben-Weyl, Methoden der organischen Chemie, Volume 15/2, but especially carbodiimides such as, for example, Ν,Ν'-dicyclohexylcarbodiimide, N,N'-diisopropylcarbodiimide or N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide. The coupling can, moreover, be carried out directly by addition of amino acid derivative with the activating reagent and, where appropriate, with an additive suppressing racemization, such as, for example, 1-hydroxybenzotriazole (HOBt) (W. Kdnig, R. Geiger, Chem. Ber. 103. 708 (1970)) or 3-hydroxy-4-oxo-3,4-dihydro15 benzotriazine (HOObt) (W. Konig, R. Geiger, Chem. Ber. 103. 2054 (1970)) to the resin, or else the preactivation of the amino acid derivative as symmetrical anhydride or HOBt or HOObt ester can take place separately, and the solution of the activated species in a suitable solvent can be added to the peptide-resin ready for coupling.

The coupling or activation of the amino acid derivatives with one of the abovementioned activating reagents can be carried out in dimethylformamide, N-methylpyrrolidone or methylene chloride or a mixture of the said solvents. The activated amino acid derivative is normally employed in a 1.5- to 4-fold excess. In cases where incomplete coupling occurs, the coupling reaction is repeated without previously carrying out the deblocking, which is necessary for the coupling of the amino acid next in sequence, of the α-amino group of the peptide-resin.

The success of the coupling reaction can be checked using the ninhydrin reaction as described, for example, by E. Kaiser et al. Anal. Biochem. 34 595 (1970). Automated synthesis is also possible, for example using a model 430A peptide synthesizer from Applied Biosystems, in which case it is possible to use either the synthesis - 14 programs provided by the manufacturer of the apparatus or else those drawn up by the user himself. The latter are employed especially when amino acid derivatives protected by the Fmoc group are used.

After the peptides have been synthesized in the manner described above, the peptide can be cleaved off the resin using reagents such as, for example, liquid hydrogen fluoride (preferably for the peptides prepared by the Boc method) or trifluoroacetic acid (preferably for the peptides synthesized by the Fmoc method). These reagents cleave not only the peptide off the resin but also the other side-chain protective groups of the amino acid derivatives. The peptide is obtained in the form of the free acid in this way, except when BHA- and MBHA-resins are used. In the case of the BHA- and MBHA-resins, the cleavage with hydrogen fluoride or trifluoromethanesulfonic acid yields the peptide as amide. Further processes for preparing peptide amides are described in EP-A 287 882 and EP-A 322 348. In this case the peptide amides are cleaved off the resin by treatment with moderately strong acids conventionally used in peptide synthesis (for example trifluoroacetic acid), adding as cation traps substances such as phenol, cresol, thiocresol, anisole, thioanisole, ethanedithiol, dimethyl sulfide, ethyl methyl sulfide or similar cation traps conventional in solid-phase synthesis, singly or a mixture of two or more of these aids. The trifluoroacetic acid can also be used diluted by suitable solvents such as, for example, methylene chloride in this case.

If the tert-butyl or benzyl side-chain protective groups on the peptides are to be retained, the peptide which has been synthesized on a specially modified support resin is cleaved off with 1% trifluoroacetic acid in methylene chloride as described, for example, by R.C. Sheppard J. Chem. Soc., Chem. Comm. 1982. 587. If individual tertbutyl or benzyl side-chain protective groups are to be retained, a suitable combination of the synthesis and It 911387 * ' · ’«Λ - 15 cleavage off methods is used.

The modified support resin described by Sheppard is also used for synthesizing peptides with a C-terminal carbamoyl group or an ω-amino- or ω-guanidinoalkyl group.

After the synthesis, the peptide which is completely protected in the side chain is cleaved off the resin and then reacted in classical solution synthesis with the appropriate amine or ω-aminoalkylamine or ω-guanidinoalkylamine, in which case it is possible where approp10 riate to protect temporarily other functional groups which are present in a known manner.

Another process for preparing peptides with an ω-aminoalkyl group is described in EP-A 264 802.

The peptides of the present invention were preferably 15 synthesized using the solid-phase technigue with two general protective group tactics: The synthesis was carried out with a model 430 A automatic peptide synthesizer from Applied Biosystems using Boc and Fmoc protective groups for temporary blocking of the α-amino group.

When the Boc protective group was used, the synthesis cycles preprogramed by the manufacturer of the apparatus were used for the synthesis.

The peptides with a free carboxyl group at the C-terminal 25 end were synthesized on a 4-(hydroxymethyl)phenylacetamidomethylpolystyrene resin functionalized with the appropriate Boc-amino acid (R.B. Merrifield, J. Org.

Chem. 43, 2845 (1978)) from Applied Biosystems. An MBHAresin from the same company was used to prepare peptide amides. The activating reagents used were Ν,Ν'-dicyclohexylcarbodiimide or Ν,Ν'-diisopropylcarbodiimide. The activation took place as symmetrical anhydride, as HOBt ester or HOObt ester in - 16 CH2C12, CH2C12/DMF mixtures or NMP. 2-4 equivalents of activated amino acid derivative were employed for the coupling. In cases where the coupling was incomplete, the reaction was repeated.

When the Fmoc protective group was used for temporary protection of the α-amino group, our own synthesis programs were entered for the synthesis with the model 430A automatic peptide synthesizer from Applied Biosystems . The synthesis was carried out on a p-benzyloxy10 benzyl alcohol-resin (S. Wang, J.Am.Chem.Soc. 55., 1328 (1973)) from Bachem, which was esterified with the appropriate amino acid by a known method (E. Atherton et al. J.C.S.Chem. Comm. 1981. 336). The activation of the amino acid derivatives as HOBt or HOObt ester took place directly in the amino acid cartridges supplied by the manufacturer of the apparatus, by addition of a solution of diisopropylcarbodiimide in DMF to the mixture of amino acid derivative and HOBt or HOObt which had previously been weighed in. It is likewise possible to employ Fmoc20 amino acid OObt esters prepared in bulk, as are described in EP-A 247 573. The Fmoc protective group was eliminated using a 20% strength solution of piperidine in DMF in the reaction vessel. The excess of reactive amino acid derivative used was 1.5 to 2.5 equivalents. If the coupling was incomplete, it was repeated as for the Boc method.

The peptides according to the invention have, singly or in combination, a bradykinin-antagonistic action which can be tested in various models (see Handbook of Exp.

Pharmacol. Vol. 25, Springer Verlag, 1970, pp. 53 - 55), for example on the isolated rat uterus, on the guinea pig ileum or on the isolated guinea pig pulmonary artery.

To test the peptides according to the invention in the isolated pulmonary artery, guinea pigs (Dunkin Hartley) weighing 400 - 450 g are sacrificed by a blow to the back of the neck. - 17 The thoracic cavity is opened and the pulmonary artery is carefully dissected out. The surrounding tissue is carefully removed and the pulmonary artery is cut open helically at an angle of 45°.

The strip of vessel which is 2.5 cm long and 3 - 4 mm wide is fixed in an organ bath with a capacity of 10 ml which is filled with Ringer solution. Composition of the solution in mmol/1 NaCl 154 KC1 5.6 CaCl2 1.9 NaHCO3 2.4 Glucose 5.0 95% O2 and 5% C02 is bubbled through the solution, which is heated to 37*C. The pH is 7.4, and the load on the strip of vessel is 1.0 g.

The changes in the isometric contractions are detected with a lever attachment and an HF modem (path-measuring device) from Hugo Sachs and recorded on a potentiometric recorder (BEC, Goerz Metrawatt SE 460).

After equilibration for 1 hour the experiment is started. Once the strips of vessel have reached their maximum sensitivity to 2 x 10'7 mol/1 bradykinin - bradykinin causes the strips of vessel to contract - the peptides are allowed to act in the doses 5 x 10'8 -lx 10'5 mol/1 for 10 minutes each and, after renewed addition of bradykinin, the decrease in the effect of bradykinin is compared with the control.

To detect a partial agonistic effect, the peptides are used in the doses 1 x 10'5 -lx 103 mol/1.

The IC50 values of the peptides according to the invention - 18 calculated from the dose-effect plots are listed in Table 1.

Table 1: Compound IC50 [M] H-(D)-Arg-Arg-Pro-Hyp-Gly-Leu-Ser-(D)-Tic-Oic-Arg-OH 5.9xl0'8 H-(D)-Arg-Arg-Pro-Hyp-Gly-Cha-Ser-(D)-Tic-Oic-Arg-OH 3,7x10"® H-(D)-Arg-Arg-Pro-Hyp-Gly-Tbg-Ser-(D)-Tic-Oic-Arg-OH 6,0x10"6 The therapeutic uses of the peptides according to the invention embrace all pathological states which are mediated, induced or assisted by bradykinin and peptides related to bradykinin. These comprise, inter alia, traumata such as wounds, burns, rashes, erythemas, edemas, tonsillitis, arthritis, asthma, allergies, rhinitis, shock, inflammations, low blood pressure, pain, pruritus and altered sperm motility.

The invention therefore also relates to the use of peptides of the formula I as medicines and to pharmaceutical products which contain these compounds.

Pharmaceutical products contain an effective amount of the active compound of the formula I - singly or in combination - together with an inorganic or organic pharmaceutically utilizable excipient.

Administration can be enterally, parenterally - such as, for example, subcutaneously, i.m. or i.v. -, sublingu25 ally/ epidermally, nasally, rectally, intravaginally, intrabuccally or by inhalation. The dosage of the active compound depends on the warm-blooded species, the body weight, age and on the mode of administration.

The pharmaceutical products of the present invention are prepared by dissolution, mixing, granulation or coating processes known per se. - 19 For the form for oral administration or for administration onto the mucosa, the active compounds are mixed with the additives customary for this purpose, such as excipients, stabilizers or inert diluents, and converted by customary methods into suitable dosage forms such as tablets, coated tablets, hard gelatin capsules, aqueous, alcoholic or oily suspensions or aqueous, alcoholic or oily solutions. Examples of inert vehicles which can be used are gum arabic, magnesia, magnesium carbonate, potassium phosphate, lactose, glucose, magnesium stearyl fumarate or starch, especially corn starch. This preparation can take place both as dry and wet granules. Examples of suitable oily excipients or solvents are vegetable or animal oils, such as sunflower oil and fish liver oil.

A product for topical administration can be in the form of an aqueous or oily solution, lotion, emulsion or jelly, ointment or fatty ointment or, if possible, in spray form, it being possible where appropriate to improve adhesion by adding a polymer.

For the form for intranasal administration, the compounds are mixed with the additives customary for this purpose, such as stabilizers or inert diluents, and converted by the customary methods into suitable dosage forms such as aqueous, alcoholic or oily suspensions or aqueous, alcoholic or oily solutions. It is possible to add chelating agents, ethylenediamine-Ν,Ν,Ν',N'-tetraacetic acid, citric acid, tartaric acid or salts thereof to aqueous intranasal preparations. The nasal solutions can be administered using a metering atomizer or as nose drops with a viscosity-increasing content, or nasal gels or nasal creams.

It is possible to use for administration by inhalation atomizers or compressed gas packs employing inert carrier gases. - 20 For intravenous, subcutaneous, epicutaneous or intradermal administration, the active compounds or the physiologically tolerated salts thereof are converted, if required with the pharmaceutically customary auxiliaries, for example for isotonicity or adjusting the pH, and solubilizers, emulsifiers or other auxiliaries, in solution, suspension or emulsion.

The short half-lives of some of the described pharmaceutical substances in body fluids mean that it is worthwhile to use injectable depot preparations. Examples of pharmaceutical forms which can be used are oily crystal suspensions, microcapsules, rods or implants, it being possible for the latter to be composed of tissuecompatible polymers, especially biodegradable polymers such as, for example, based on polylactic acid/polyglycolic acid copolymers or human albumin.

A suitable dose range for forms for administration topically and by inhalation are solutions containing 0.01-5 mg/ml, while 0.01-10 mg/kg are suitable for systemic administration forms.

List of abbreviations: The abbreviations used for amino acids correspond to the three-letter code customary in peptide chemistry, as described in Europ. J. Biochem. 138. 9 (1984). Other abbreviations used are listed below.

Acm e-Ahx Aoc Boc But Bzl CDF Cha Chg acetamidomethyl £ -aminohexanoyl cis, endo-2-azabicyclo[3.3.0]octane-3-Scarbonyl tert-butyloxycarbonyl tert-butyl benzyl chloro-(D)-phenylalanyl eyelohexylalanyl eyelohexylglycyl IE 911387 - 21 10 Cl-Z 4-chlorobenzyloxycarbonyl DMF dimethylforrnamide DOMT O-methyl-(D)-threonyl Dnp 2,4-dinitrophenyl Fmoc 9-fluorenylmethyloxycarbonyl MDY O-methyl-(D)-tyrosyl Me methyl 4-Mebzl 4-methylbenzy1 Mtr 4-methoxy-2,3,6-trimethylphenylsulfonyl Mts mesitylene-2-sulfonyl Nal 1- or 2-naphthylalanyl NMP N-methylpyrrolidine Npg neopentylglycyl Oic cis-endo-octahydroindole-2-carbonyl Opr isoxazolidin-3-ylcarbonyl Pal 2- or 3-pyridylalanyl Pmc 2,2,5,7,8-pentamethylchroman-6-sulfonyl Tbg tert-butylglycyl TFA trifluoroacetic acid Thia 2-thienylalanyl Tcs 4-methylphenylsulfonyl Tic 1,2,3,4-tetrahydroisoquinolin-3-ylcarbonyl Trt trityl The examples which follow are intended to illustrate the 25 preferred methods for the solid-phase synthesis of the peptides according to the invention without restricting the invention thereto.

The following amino acid derivatives were used: Fmoc-Arg(Mtr)-OH, Boc-(D)-Arg-OH, Fmoc-Arg(Pmc)-OH, Fmoc-Hyp-OH, Fmoc-Pro-OObt, Fmoc-Gly-OObt, Fmoc-Phe-OObt, Fmoc-Ser(tBu)-OObt, Pmoc-(D)-Tic-OH, Fmoc-Gln-OH, Fmoc-Aoc-OH, Fmoc-Thia-OH, Fmoc-Opr-OH, Fmoc-(D)-Asn-OH, Fmoc-/9-Ala-OH, Fmoc-Oic-OH. - 22 Example 1: H-(D)-Arg-Arg-Hyp-Pro-Gly-Leu-Ser-(D)-Tic-Oic-Arg-OH was synthesized stepwise using a model 430 A peptide synthesizer from Applied Biosystems and using the Fmoc method on a p-benzyloxybenzyl alcohol-resin esterified with Fmoc-Arg(Mtr)-OH from Novabiochem (loading about 0.5 mmol/g of resin). 1 g of the resin was employed, and the synthesis was carried out using a synthesis program modified for the Fmoc method.

In each case, 1 mmol of the amino acid derivatives with free carboxyl group were weighed together with 0.95 mmol of HOObt into the synthesizer cartridges. These amino acids were preactivated directly in the cartridges by dissolving in 4 ml of DMF and adding 2 ml of a 0.55 mol/1 solution of diisopropylcarbodiimide in DMF. The HOObt esters of the other amino acids were dissolved in 6 ml of NMP and then coupled just like the in situ preactivated amino acids to the resin which had previously been deblocked with 20% piperidine in DMF. After the synthesis was complete, the peptide was cleaved off the resin with simultaneous removal of the side-chain protective groups with trifluoroacetic acid using thioanisole and ethanedithiol as cation traps. The residue obtained after stripping off the trifluoroacetic acid was digested several times with ethyl acetate and centrifuged. The remaining residue was chromatographed on *Sephadex LH 20 with 10% strength acetic acid. The fractions containing the pure peptide were combined and freeze-dried.

MS(FAB) : 1264 (M+H) The following examples were prepared in analogy to Example 1. - 23 Example 2: H-(D)-Arg-Arg-Hyp-Pro-Gly-Cha-Ser-(D)-Tic-Oic-Arg-OH MS (FAB): 1304 (M+H) Example 3: H-(D)-Arg-Arg-Hyp-Pro-Gly-Tbg-Ser-(D)-Tic-Oic-Arg-OH MS (FAB): 1264 (M+H)

Claims (12)

1. A peptide of the formula I A-B-C-E-F-K-(D)-Tic-G-M-F'-I (I) in which 5 A is a x ) hydrogen, (Ci-Cg)-alkyl, (C x -C 8 ) -alkanoyl, (Ci-Cg) -alkoxycarbonyl or (Ci-Cg) -alkylsulfonyl, 10 in each of which 1, 2 or 3 hydrogen atoms are optionally replaced by 1, 2 or 3 identical or different radicals from the series comprising carboxyl, 15 amino, (Ci-CJ-alkyl, (Ci-C 4 ) -alkylamino, hydroxyl, (Ci-CJ-alkoxy, 20 halogen, di- (Ci-C 4 ) -alkylamino, carbamoyl, sulfamoyl, (Ci-C 4 )-alkoxycarbonyl, 25 (C 6 -C 12 )-aryl and (C 6 -C 12 )-aryl-(Ci-C 5 )-alkyl, or in each of which 1 hydrogen atom is optionally replaced by a radical from the series comprising 30 (C 3 -C 8 )-cycloalkyl, (Ci-C 4 ) -alkylsulfonyl, (C1-C4) -alkylsulf inyl, (C 6 -Ci 2 ) -aryl- (Ci-C 4 ) -alkylsulfonyl, (C 6 “C 12 ) -aryl- (Ci-C 4 ) -alkylsulf inyl, 35 (C 6 -C 12 )-aryloxy, - 25 (C 3 -C 8 ) -heteroaryl and (C 3 -C e ) -heteroaryloxy and 1 or 2 hydrogen atoms are replaced by 1 5 or 2 identical or different radicals from the series comprising carboxyl, amino, (C x -C 4 ) -alkylamino, 10 hydroxyl, (C x -C 4 ) -alkoxy, halogen, di- (C x -C 4 ) -alkylamino, carbamoyl, 15 sulfamoyl, (C x -C 4 )-alkoxycarbonyl, (C 6 -C x2 )-aryl and (C 6 -C 12 ) -aryl- (C x -C 5 ) -alkyl, a 2 ) (C 3 -C 8 )-cycloalkyl, 20 carbamoyl which can optionally be substituted on the nitrogen by (C x -C 6 )-alkyl or (C 6 -C x2 )aryl, (C 6 -C 12 )-aryl, (C 7 -C 13 )-aroyl, 25 (C 6 -C X2 )-arylsulfonyl, (C 3 -C 9 )-heteroaryl or (C 3 -C 8 )-heteroaroyl, where in each of the radicals defined under a x ) and a 2 ) aryl, heteroaryl, aroyl, arylsulfonyl and heteroaroyl is optionally substituted by 1, 30 2, 3 or 4 identical or different radicals from the series comprising carboxyl, amino, nitro, 35 (C x -C 4 )-alkylamino, hydroxyl, (C x -C 4 ) -alkyl, (C x -C 4 )-alkoxy, halogen, - 26 cyano, di- (C^-C*) -alkylamino, carbamoyl, sulfamoyl and (Ci-04)-alkoxycarbonyl or a 3 ) a radical of the formula II R 1 - N - CH - C i - i _ a (II) in which 10 R 1 is defined as A under aj or a 2 ), R 2 is hydrogen or methyl, R 3 is hydrogen or (Ci-Cg)-alkyl, preferably (Ct-C*)-alkyl, which is optionally monosubstituted by 15 amino, substituted amino, hydroxyl, carboxyl, carbamoyl, 20 guanidino, substituted guanidino, ureido, mercapto, methylmercapto, 25 phenyl, 4-chlorophenyl, 4-fluorophenyl, 4-nitrophenyl, 4-methoxyphenyl, 30 4-hydroxyphenyl, phthalimido, 4-imidazolyl, 3-indolyl,

2. -thienyl, - 27 3-thienyl, 2- pyridyl,

3. - pyridyl or cyclohexyl, 5 where substituted amino is -NH-A and substituted guanidino is -NH-C(NH)-NH-A in which A is as defined under a x ) or a 2 ); B is a basic amino acid which is in the L or D configuration and can be substituted in the side chain; C is a linkage of the formula Ilia or Illb G'-G'-Gly G'-NH-(CH 2 ) n -CO (Ilia) (Illb) in which G' is, independently of one another, a radical of 15 the formula IV in which R* and R 5 form, together with the atoms carrying them, a heterocyclic mono-, bi- or tricyclic ring 20 system with 2 to 15 carbon atoms, and n is 2 to 8; E is the residue of a neutral, acidic or basic aliphatic or alicyclic-aliphatic amino acid; F is, independently of one another, the residue of a 25 neutral, acidic or basic, aliphatic or aromatic amino acid which can be substituted in the side chain, or is a direct bond; (D)-Tic is the radical of the formula V; ccr (V) G is defined as G' above or is a direct bond; F' is defined as F, is a radical -NH-(CH 2 ) n - with n = 2 to 8, or can be a direct bond if G is not a direct bond, and 5 I is -OH, -NH 2 or -NHC 2 H 5 , K is the radical -NH-(CH 2 ) x -C0- with x = 1-4 or is a direct bond, and M is defined as F, and the physiologically tolerated salts thereof. 10 2. A peptide of the formula I as claimed in claim 1, in which B is Arg, Lys, Orn, 2,4-diaminobutyryl or an L-homoarginine residue where, in each case, the eunino or 15 the guanidino group of the side chain can be substituted by A as described under a x ) or a 2 ) in claim 1; E is the residue of an aliphatic or alicyclic-aliphatic amino acid which is in the L or D configuration and which contains 1 to 14 carbon atoms in the side chain; 20 F' is the residue of a basic amino acid in the L or D 25 configuration, such as Arg or Lys, where the guanidino group or amino group of the side chain can be substituted by A as described under a x ) or a 2 ) in claim 1, or is a radical -NH-(CH 2 ) n - with n = 2 to 8, and K is the radical -NH-(CH 2 ) X -CO- with x = 2-4 or is a direct bond. 3. A peptide of the formula I as claimed in claim 1 - 29 and/or 2 in which B is Arg, Orn or Lys, where the guanidino group or the amino group of the side chain is unsubstituted or can be substituted by (Cx-C 8 )-alkanoyl, (C 7 -C 13 )-aroyl, 5 (C 3 -C 8 )-heteroaroyl, (0χ-0 8 )-alkylsulfonyl or (C 8 -Cx 2 )arylsulfonyl, where the aryl, heteroaryl, aroyl, arylsulfonyl and heteroaroyl radicals can be substituted with, where appropriate, 1, 2, 3 or 4 identical or different radicals as described under a 2 ); E is leucine, isoleucine, norleucine, tert-butylglycine, serine, threonine or cyclohexylalanine, and K is a direct bond, and M is a direct bond.

4. A peptide of the formula I as claimed in one or more 15 of claims 1 to 3, in which A is hydrogen, (D)— or (L)-H-Arg, (D)— or (L)-H-Lys or (D)- or (L)-H-Orn, B is Arg, Orn or Lys, where the guanidino group or the amino group in the side chain can be substituted by 20 (Cx-C 8 )-alkanoyl, (C 7 -Cx 3 )-aroyl, (C 3 -C B )-heteroaroyl, (Cx-C 8 )-alkylsulfonyl or (C 6 -C 12 )-arylsulfonyl, where the aryl, heteroaryl, aroyl, arylsulfonyl and heteroaroyl radicals can optionally be substituted with 1, 2, 3 or 4 identical or different radicals from the 25 series comprising methyl, methoxy and halogen; C is Pro-Pro-Gly, Hyp-Pro-Gly or Pro-Hyp-Gly, E is Leu, lie, Tbg or Cha, F is Ser, Hser, Lys, Leu, Val, Nle, lie or Thr, K is a direct bond, - 30 M is a direct bond. G is the radical of a heterocyclic ring system of the formula IV, where the radicals of the heterocycles pyrrolidine-2-carboxylic acid; piperidine-25 carboxylic acid; l,2,3,4-tetrahydroisoquinoline-3carboxylic acid, cis- and trans-decahydroisoquinoline-3-carboxylic acid; cis-endo-, cis-exo-, trans-octahydroindole-2-carboxylic acid; cis-endo-, cis-exo-, trans-octahydrocyclopenta[b]pyrrole-210 carboxylic acid or hydroxyproline-2-carboxylic acid are preferred, F' is Arg and I is OH.

5. The preparation of a peptide of the formula I as 15 claimed in one or more of claims 1 to 4, which comprises a) reacting a fragment with a C-terminal free carboxyl group or its activated derivative with a corresponding fragment with an N-terminal free amino group, or b) synthesizing the peptide stepwise, eliminating one or more protective groups temporarily introduced into the compound obtained as in (a) or (b) to protect other functions where appropriate, and converting the compounds of the formula I which have been obtained in this way into their physiologically tolerated salt where appropriate.

6. A peptide of the formula I as claimed in one or more of claims 1 to 4 for use as medicine.

7. A peptide of the formula I as claimed in one or more 30 of claims 1 to 4 for the treatment of pathological states which are mediated, induced or assisted by

8. 8.

9. 9.

10. 10.

11. 11.

12. 12. bradykinin and bradykinin-related peptides. A pharmaceutical agent containing a peptide of the formula I as claimed in one or more of claims 1 to 4. A peptide as claimed in claim 1, substantially as hereinbefore described and exemplified. A process for preparing a peptide as claimed in claim 1, substantially as hereinbefore described and exemplified. A peptide as claimed in claim 1, whenever prepared by a process claimed in a preceding claim. A pharmaceutical agent according to claim 8, substantially as hereinbefore described.