SK282467B6 - Peptides, pharmaceutical preparation containing them and their use - Google Patents

Abstract

Peptides of formula R1R2N-CHX-CO-A-B-D-E-(F)t-K (I) and their salts with acids are new. In the formula, R1 = Me, Et or iPr; R2 = H, Me or Et; or NR1R2 = a pyrrolidine ring; A = a valyl, isoleucyl, leucyl, 2-tert-butylglycyl, 2-ethylglycyl, norleucyl or norvalyl residue; B = a N-methyl-valyl, -leucyl, -isoleucyl, -norvalyl, -norleucyl, -2-tert-butylglycyl, -3-tert-butylalanyl or -2-ethylglycyl residue; D = a prolyl, 3,4-dehydroprolyl, 4-fluoroprolyl, 4,4-difluoroprolyl, azetidine-2-carbonyl, homoprolyl, thiazolidine-4-carbonyl or 3-, 4- or 5-methylprolyl residue; E = a 3,4-dehydroprolyl, 4-fluoroprolyl, 3- or 4-methylprolyl, 4,4-difluoroprolyl or azetidine-2-carbonyl residue; F = a valyl, 2-tert-butylglycyl, isoleucyl, leucyl, 2-cyclohexylglycyl, norleucyl, norvalyl, neopentylglycyl, alanyl, ~b-alanyl or aminoisobutyroyl residue; X = alkyl (pref. 2-5C alkyl), cyclopropyl or cyclopentyl; t = 0 or 1; and K = alkoxy (pref. 1-4C alkoxy), benzyloxy or opt. substd. amino.

Description

Technical field

The present invention relates to peptides, to a pharmaceutical composition containing them, and to their use in the treatment of cancer and mammalian tumors.

BACKGROUND OF THE INVENTION

Peptides isolated from marine sources such as Dolastatin-10 (U.S. Pat. No. 4,816,444) and Dolastatin-15 (European Patent Specification No. EP-A-398558) are known to have great activity as cell growth inhibitors (Biochem Pharmacology 40; 8, pp. 1859 to 1864, 1990; J. Natl.Cancer Inst. 85, pp. 483-488, 1993 and references therein). Based on interesting results in experiments with tumor systems in vivo, further preclinical evaluations of these natural products are currently underway to initiate clinical trials for cancer patients.

However, natural products have the drawback that they are poorly soluble in aqueous solvents and the investment costs for their synthesis are high.

The invention relates to novel peptides and derivatives thereof which represent an improved therapeutic potential for the treatment of neoplastic diseases compared to Dolastatin-15. In addition, the novel peptides of the invention can be readily synthesized in the manner described in detail below.

SUMMARY OF THE INVENTION

The present invention provides peptides of formula (I)

R'R ² N-CHX-CO-A-B-D-E- (F) _t -K (I) where

R ^{1 is} methyl, ethyl or isopropyl,

R ² is hydrogen, methyl or ethyl,

^R'-NR2 together pyroilidínový ring,

A residues valyl, isoleucyl, leucyl, 2-tert-butylglycyl, 2-ethylglycyl, norleucyl or norvaiyl,

B residues N-methylvalyl, N-methylleucyl, N-methylisoleucyl, N-methylnorvalyl, N-methylnorleucyl, N-methyl-2-tert-butylglycyl, N-methyl-3-tert-butylalanyl or N-methyl-2-ethylglycyl,

D residues prolyl, 3,4-dehydroprolyl, 4-fluorprolyl, 4,4-difluoropolyl, azetidine-2-carbonyl, homoprolyl, 3-methylprolyl, 4-methylprolyl, 5-methylprolyl or thiazolidine-4-carbonyl,

E 3,4-dehydroprolyl, 4-fluorprolyl, 3-methylprolyl, 4-methylprolyl, azetidine-2-carbonyl or 4,4-difluoroprolyl,

F residues valyl, 2-tert-butylglycyl, isoleucyl, leucyl, 2-cyclohexylglycyl, norleucyl, norvaiyl, neopentylglycyl, alanyl, β-alanyl or aminoisobutyroyl,

X is preferably alkyl of 2 to 5 carbon atoms, cyclopropyl or cyclopentyl, t is 0 or 1;

The alkoxy group preferably having 1 to 4 carbon atoms, benzyloxy or an amino group -NR ⁵ R ^6, wherein

R ^{5 is} hydrogen or hydroxy or C _1-4 alkoxy or benzyloxy optionally substituted with up to two substituents independently selected from the group consisting of CF ₃₎ nitro, C _1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl, cyano, hydroxy, N (CH) ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or R ⁵ represents a phenyloxy group, optionally substituted with up to two substituents independently selected from the group consisting of CF ₃ , nitro, C ₁ . ₄ alkylsulfonyl, C _1-4 alkoxy, halogen, C _1-4 alkyl, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ ,

R ^{6 is} hydrogen or C 1-4 alkyl, optionally substituted with at least one fluorine atom, or - (CH ₃ ) ₂ CN, -C (CH ₃ ) ₂ CCH, -C (CH ₃ ) ₂ C = CH ₂ , -C (CH ₃ ) ₂ CH ₂ OH, -C (CH ₃ ) ₂ CH ₂ CH ₂ OH, or - (CH ₂ ) n C ₃ . ₇ cycloalkyl wherein v is 0, 1 or 2 and which is optionally substituted with methyl, or is norephedryl, norpseudoephedryl, quinolyl, pyrazyl, adamantyl, -CH ₂ -benzimidazolyl, -CH ₂ -adamantyl, α-methylbenzyl, α-dimethylbenzyl, - (CH ₂₎ phenyl, wherein v is 0, 1, 2 or 3, and optionally substituted with one or two substituents each independently selected from: CF _3, nitro, C _{i. 4} alkylsulfonyl, C 1-4 alkoxy, halogen, C _{1 (} alkyl which can form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr and COONH _Z , or is - (CH 2 n 1) phthyl where m is 0 or 1 and which is optionally substituted by one or two substituents independently selected from the group consisting of CF ₃ , nitro, C _{1 J (} alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, N (CH _{3) 12} , COOMe, COOEt, COOiPr or COONH ₂ , or - (CH ₂ ) _w benzhydryl, wherein w is 0, 1 or 2, and which is optionally substituted with one or two substituents independently selected from CF ₃ , nitro , C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is biphenyl which is optionally subs substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ or is pyridyl optionally substituted by one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 -alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is a picolyl optionally substituted by one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is -CH ₂ -CH ₂ -pyr idyl optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-6 alkyl which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is benzothiazolyl, which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is benzoisothiazazolyl optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano , hydroxy, N (CH _{3) 2,}

SK 282467 Β6

COOMe, COOEt, COOiPr, or COONHj, or is benzopyrazolyl, KTOI is optionally substituted by one or two substituents each independently selected from: CF _3, nitro, C _{alkylsulfonyl,} C _M alkoxy, halogen, C _M alkyl which may form a cyclic a system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is benzooxyzolyl optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C _1-4 alkylsulfonyl, CMalkoxyskupinu, halogen, _C-alkyl which may form a cyclic system, cyano, hydroxy, N (CH _{3) 2,} COOMe, COOEt, COOiPr, or COONH _2, or is - (CH _{2) m} fluorenyl, wherein m is 0, or 1, which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C _1-4 alkoxy groups u, halogen, C _1-4 alkyl, which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is pyrimidyl optionally substituted by one or two substituents independently selected from one another from the group consisting of CF _3, nitro, C _w alkylsulfonyl, C _M alkoxy, halogen, _C-alkyl which may form a cyclic system, cyano, hydroxy, N (CH _{3) 2,} COOMe, COOEt, COOiPr, or COONH _2, or is - (CH ₂ ) _m -indanyl, wherein m is 0 or 1, which is optionally substituted with one or two substituents independently selected from the group consisting of CF 3, nitro, C _1-4 alkylsulfonyl, C _1-4 alkoxy, halogen, C _1-4 alkyl which can form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is - (CH ₂ CH ₂ O) _y CH ₃ , where y is 0, 1, 2, 3, 4 or 5, or - (CH ₂ CH ₂ O) _y CH ₂ CH ₃ , wherein y is 0, 1, 2, 3, 4 or 5, or is -NH-C ₆ H ₃ , which is optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C _1-4 alkoxy, halogen, C _1-4 alkyl, which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is -NCH ₃ -C ₆ H ₃ optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or -NH-CH ₂ -C ₅ H ₅ , which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form c a cyclo system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is -NCH ₃ -CH ₂ -C 6 H 5 optionally substituted by one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy , halogen, C 1-4 alkyl, which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is a five-membered heteroaryl optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl , C 1-4 alkoxy, thiomethyl, thioethyl, picolyl, acetyl, C ₃ . ₆ cycloalkyl, thiophenyl, CH ₂ COOEt, C ₃ . _{A (4)} alkylene forming a bicyclic system with a heterocycle, phenyl optionally substituted with one or two substituents independently selected from the group consisting of nitro, CF ₃ , cyano, halogen, or C 1-4 alkyl, and further comprising benzyl optionally substituted with one or two substituents from independently selected from the group consisting of nitro, CF ₃ , halogen, C 1-4 alkyl, C 1-4 alkylsulfonyl, cyano, hydroxy, C 1-4 dialkylamino, or is a five-membered -CHR ⁷ -heteroaryl optionally substituted by one or two substituents independently selected from each other CF ₃ , nitro, cyano, halogen, COOMe, COOEt, COOiPr and COONH ₂ , C 1-4 alkyl, C 1-4 alkoxy, phenyl, benzyl, naphthyl or C 1-4 alkylsulfonyl, wherein R ⁷ is hydrogen, straight-chain or branched C 1-4 alkyl or benzyl or a salt thereof with a physiologically acceptable acid. The invention also relates to a pharmaceutical composition comprising these peptides of formula (I) together with pharmaceutically acceptable carriers, and their use in the treatment of cancer and mammalian tumors.

For example, the five-membered -CHR ⁷ -heteroaryl moieties include:

For example, the five-membered heteroaryl is as follows:

As a group -NR ^{6 R} is, for example, include the following groups

-Ό / “Λ r ~ \ ~ \ _7 ^S -Ν __ / = Ο r ~ ₄ o ~ ^N vJ

Preference is given to peptides of the general formula (I), in which:

R ^{1 is} methyl or ethyl,

R ² is hydrogen, methyl or ethyl,

And the residues valyl, isoleucyl or 2-tert-butylglycyl,

B residues N-methylvalyl, N-methylisoleucyl or N-methyl-2-tert-butylglycyl,

D residues prolyl, 3,4-dehydroprolyl, 4-fluorprolyl, 3-methylprolyl or azetidine-2-carbonyl,

E residues 3,4-dehydroprolyl, 4-fluorprolyl, 3-methylprolyl or azetidine-2-carbonyl,

F residues D-valyl, 2-tert-butylglycyl, D-isoleucyl, D-leucyl or aminoisobutyroyl,

X-CH (CH ₃ ) ₂ , -C (CH ₃ ) ₃ , -CH (CH ₃ ) CH ₂ CH ₃ ;

t is 0 or 1; and

K is preferably

-OC (CH ₃ ) ₃ , -NH ₂ , -NH-C 1-4 -alkyl, -NH-C _1-4 -cycloalkyl, -NH- [3,3,0] -bicyclooctyl, norephedryl, norpseudoephedryl, -NH-quinolyl, - NH-pyrazyl, _-NH-CH2 -benzimidazolyl, -NH-adamantyl, _{-NH-CH2-adamantyl,} -NH-CH (CH ₃₎ phenyl,

-NH-C _{(CH3) 2-phenyl,} -N (C _M alkoxy) C _M-alkyl, N (C _{first 4} -alkoxy) _-CH2 -phenyl, -N (C _M alkoxy) - phenyl, -N (CH ₃ ) OBzl, -NH- (CH ₂ ) _v -phenyl (v = 0.1, 2.3), -NH- (CH ₂ ) _m naphthyl (m = 0 or 1), - NH- (CH ₂ ) _n -benzhydryl (w = 0, 1 or 2),

NH-biphenyl, -NH-pyridyl, -NH-CH ₂ -pyridyl, _{-NH-CH2 -CH2} -pyridyl, -NH-benzothiazolyl, -NH-benzoisothiazolyl, -NH-benzopyrazolyl, -NH-benzoxazolyl, - NH- (CH ₂ ) _m -fluorenyl (m = 0, 1), -NH-pyrimidyl, -NH- (CH ₂ ) _m -indanyl (m = 0, 1), -NH- (CH ₂ CH ₂ O) _y -CH ₃ (y = 0.1, 2,3,4,5), -NH- (CH ₂ CH ₂ O) _y -CH ₂ CH ₃ (y = 0,1,2,4,4,5) -NH-five-membered heteroaryl, as previously described, -NH-CHR ⁷ -five-membered heteroaryl, as described above, wherein R ⁷ is hydrogen, C 1. ₅ alkyl, straight or branched chain, or C is a residue

In particular, K represents a residue

-NHCH, -NHCHjCHj, -NHÍCHiKHj, -NHÍCHjjjCHj, -NH (CH _2> CH, -NH _(CH2) SCH3, NH (CH _{2) r} CH _3, -NH _{(CH2> CH3,} -NHCH ( CH ₃ ), -NHCH (CH ₃ ) CH ₂ CH ₃ , -NHCHCCH ₃ CH ₂ CH ₂ CH ₃ , -NHCH 2 CH ₃ CH ₃ , -NHCH ₃ CH ₃ CH ₃ , -NHC (CH ₃ ) ₃ , -NHCH ₃ CH ₃ CH ₃ CH ₃ CH (CH ₃ h, - NHCH (CH ₂ CH ₃ ) CH (CH ₃ ) 2, -NHCHCCH ₃ HCH 3),

-NH-cyclopropyl, -NH-cyclobutyl, -NH-cyldopentyl, -NH-cycloxyL -NH-cycloheptyl, -NH-cyclooctyl, -NH-bicyclo [33.0] octyl, -NfCH3JOCHa, -NfCH3JOCH3, -NjCH3jOCHiCH3CH3, -N (CH ₃ CH ₃ ) -N (CH ₃ CH ₃ ) -N (CH ₃ CH ₃ ) -N (CH (CH ₃ );) OCH 3, -N (CH ₃ CH ₃₎ -N (CH ₃ CH 2 ₎ , -N (CH ₃ X 2 H, _and -NH-CH 2 Celk -NH ₃ CH ₃ ); -NHfCH3JjCeHs -NHCH3CHjjCsH ,, -NHCfCH3jjJfs, -NHCiCl2jJjCIIjCIIj, -NHQCH3XCH3H4, -NHCHfCH3jCHfOHjCÄ

-NHCHcyclohexyl, -NH-CH ₃ CF ₃ , -NHCH (CH ₂ Fh, -NHQCH 2 H ₃ CH ₃ OH, -N 1 -NHC (CH ₃ h CH (CH ₃ ) 2),

-NHC (CH3CN, -NHQCHaCCH, -NHQCH3hC-CH3, norephedryl, norpseudoephediyl, -NH-quinolyl, -NH-pyrazyl,

-NH-adamantyl (l), -NH-adamantyl (2), -NH-CH _r adamantyl, -NH-CHrnaítyi, -NH-benzhydryl, -NH-biphenyl, -NH-pyridyl, -NH-picolyl, -NH -CH 3 -CH 3 -pyridyl, -NH-benzothiazolyl, -NH-benzoisothiazolyl, -NH-benzopyrazolyl, -NH-benzoxaznlyl, -NH-fluorenyl, -NH-pyrimidyl, -NH-thiazolyl (2), -NH-isoxazolyl ( 3), -NH-CH 2 -firanyl (2), -NH- (3-methyl) 12Oxazolyl (5), -NH- (3-methyl) 12-thio 2 Dlyl (5), -NH- (2-trifluoromethyl) thiadiazolyl ( 5), -OC (CH ₃ , -NH-CH 2 (4-methyl) thiazolyl (2), -NH-CH 2 -thienyl (2), -NH-CH 2 (5-n-ethyl) ticyl (2), -NH - (2-methyl) thiadiazoles (5), -NH- (2-cyclopropyl) thiadiazolyl (5), or K represents a radical of formula

The examples given above merely illustrate the invention but do not limit it.

The peptides of formula (I) are composed of L-amino acids, but F may also be D-amino acids.

The peptides of formula (I) may be in the form of a salt with physiologically acceptable acids such as hydrochloric, citric, tartaric, lactic, phosphoric, methanesulfonic, acetic, formic, maleic, fumaric, malic, succinic, malonic, sulfuric, L- glutamic, L4

SK 282467 Β6

-aspartic, pyruvic, mucosal, benzoic, glucuronic, oxalic, ascorbic and acetylglycine.

The peptides of formula (I) may be prepared by methods known in the art from peptide chemistry. For example, they may sequentially consist of amino acids, or may be obtained by binding suitable small peptide fragments. Gradual folding starts from the C-terminus of the peptide chain, which is gradually extended by one amino acid each. In coupling the fragments, it is possible to bind fragments of varying length together, such fragments being obtained by sequential amino acid binding or also by coupling the fragments. In both of these methods, it is necessary to bind the units to each other by forming an amide bond. Both chemical and enzymatic methods are suitable for this purpose.

Chemical methods for amide bond formation have been described in detail by Mueller (Methoden der organischen Chemie, Vol. XV / 2, pp. 1 to 364, Thieme Verlag, Stuttgart, 1974; Stewart, Young, Solid Phase Peptide Synthesis, pp. 31-34, 71-71). 82, Pierce Chemical Company, Rockford, 1984; Bodanszky, Klausner, Ondetti, Peptide Synthesis, pp. 85-128, John Wiley & Sons, New York, 1976) and also described in other basic publications on peptide chemistry. Particularly preferred are the azide process, symmetrical and mixed anhydride processes, in situ generated and preformed active esters, use of urethane protected with N-carboxyanhydrides of amino acids, and amide bonding using a coupling / activator system, mentioning in particular dicyclohexylcarbodiimide (DCC), diisopropylcarbodi DIC), 1-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI), n-propanephosphonic anhydride (PPA), N, N-bis (2 -oxo-3-oxazolidinyl iamidophosphoryl chloride (ΒΟΡ-Cl), bromo-trispyrrolidinophosphonium hexafluorophosphate (PyBrop), diphenylphosphoryl azide (DPPA) of Castro reagent (BOP, PyBop), O-baisotriazolyl-N, N, Tamet, , 0-azabenzotriazolyl-N, N, N ', N'-tetramethyluronium salt (HATU)> diethylphosphoryl cyanide (DEPCN), 2,5-diphenyl-2,3-dihydro-3-oxo-4-hydroxythiophene dioxide (Steglich reagent, HOTDO) α1,1'-carbonyldiimidaz The coupling reagents may be used alone or in combination with additives such as N, N-dimethyl-4-aminopyridine (DMAP), N-hydroxybenzotriazole (HOBt), N-hydroxysuccinimide (HOSu) or 2-hydroxypyridine.

While it is generally possible to dispense with protecting groups in the enzymatic synthesis of peptides, transient protection of reactive groups not involved in the amide bond is needed for the reactants in chemical synthesis. Three conventional methods of protecting groups are preferred for chemical peptide synthesis: the method using benzyloxycarbonyl (Z), t-butoxycarbonyl (Boe) and 9-fluorenylmethoxycarbonyl (Fmoc). In each case, a protecting group on the α-amino group of the chain extender is used. A detailed review of the amino protecting groups is given by Mueller (Methoden der organischen Chemie, Vol. XV / 1, pp. 20-906, Thieme Verlag, Stuttgart, 1974).

The units used to form the peptide chain can be reacted in solution, suspension or the like, as described by Murrifield (J. Amer. Chem. Soc. 85, 2149, 1963). Particularly suitable are methods in which the peptides are formed sequentially or by coupling the fragments using Z, Boe or Fmoc protecting groups. One of the reactants in the method described by Murrifield binds to the insoluble polymeric carrier (also referred to herein as resin). As a rule, the peptide is formed sequentially on a polymeric support using Boe or Fmoc protecting groups, the growing peptide chain is covalently linked by a C terminus to insoluble resin particles (Figs. 1 and 2). This process makes it possible to remove the reagents and by-products by filtration, and thus recrystallization of the intermediates is unnecessary.

The protected amino acids may bind to any suitable polymer, which must only be insoluble in the solvent used and must be in a stable physical form that facilitates filtration. The polymer must contain a functional group to which the first protected amino acid can be firmly bound by a covalent bond. A wide variety of polymers are suitable for this purpose: for example, cellulose, polyvinyl alcohol, polymethacrylate, sulfonated polystyrene, a copolymer of chloromethylated styrene and divinylbenzene (Murrifield resin), 4-methylbenzhydrylamine resin (MBHA resin), phenylacetamidomethyl resin, benzhydrylamine resin (BHA resin), 4- (hydroxymethyl) benzoyloxymethyl resin, the resin described by Breipohl et al. (Tetrahedron Letters 28, p. 565, 1987, commercial product of BACHEM), 4- (2,4-dimethoxyphenylaminomethyl) phenoxy resin (commercial product of Novabiochem) or o-chlorotrityl resin (commercial product of Biohellas).

All solvents which are inert under the reaction conditions, in particular water, Ν, Ν-dimethylformamide (DMF), dimethylsulfoxide (DMSO), acetonitrile, dichloromethane (DCM), 1,4-dioxane, tetrahydrofuran (THF), are suitable for solution peptide synthesis. ), N-methyl-2-pyrrolidone (NMP) and mixtures of these solvents. The polymer-supported peptide synthesis can be carried out in all inert organic solvents in which the amino acid derivatives used are soluble. Preferred solvents such as N, N-dimethylformamide, dichloromethane, N-methyl-2-pyrrolidone, acetonitrile and dimethylsulfoxide and mixtures thereof, however, cause swelling of the resins. When the synthesis is complete, the peptide is cleaved from the polymeric carrier. The conditions under which cleavage from various resins is possible are described in the literature. The most commonly used cleavage reactions are catalyzed by acids and palladium, in particular suitable cleavage in liquid anhydrous hydrogen fluoride, in anhydrous trifluoromethanesulfonic acid, in dilute or concentrated trifluoroacetic acid, palladium catalyzed cleavage in tetrahydrofuran or in a mixture of tetrahydrofuran, morpholine or cleavage in mixtures of acetic acid and dichloromethane / trifluoromethanol. Depending on the protecting groups selected, these groups may be maintained or optionally cleaved under cleavage conditions. Partial deprotection of the peptide may also be valuable if certain derivatization reactions are to be performed. N-terminally alkylated peptides can be prepared by:

(a) by coupling the appropriate Ν, dial-dialkylamino acids in solution or on a polymeric support;

b) reductive alkylation of the resin-bound peptide in dimethylformamide / 1% acetic acid with NaCNBH ₃ and the corresponding aldehyde or ketone,

c) hydrogenating the peptides in solution in the presence of an aldehyde or ketone and palladium on carbon.

The various non-naturally occurring amino acids of the invention are commercially available or can be synthesized from commercially available materials by methods known in the art. Commercially available starting materials are azetidine-2-carboxylic acid, 3-methyl-L-proline, 5-methyl-L-proline and the Boe or Fmoc protected 3,4-dehydroproline (ACROS, NOVABIOCHEM,

Bachem). Cis- and trans-4-fluorproline can be prepared as described by Panasik et al. (N. Panasik, E. S. Eberhardt, A. S. Edison, D. R. Powell, R. T. Raines Int. J. Peptide Protein Res. 44, pp. 262-269, 1994) from hydroxyproline.

The compounds of the invention may serve to inhibit or otherwise treat solid tumors (e.g., lung, breast, colon, prostate, gall bladder, rectum or endometrial tumors) or hematological diseases (e.g., leukemia, lymphomas), by administering the compounds to mammals.

A particular advantage of the novel compounds of the invention is that they are more resistant to enzymatic degradation compared to Dolastatin-15.

Administration can be by conventional pharmaceutical, preferably oncological agents, including oral and parenteral agents, subcutaneously, intravenously, intramuscularly and intraperitoneally.

The compounds may be administered alone or in the form of pharmaceutical compositions comprising a compound of formula (I) together with a pharmaceutically acceptable carrier for a given route of administration. Such pharmaceutical compositions may be combination products and thus may contain other therapeutically active ingredients.

The dose administered to the mammal comprises a tumor inhibiting active ingredient which depends on common factors such as the biological activity of the compound used, the route of administration, the age, health and weight of the patient, the nature and type of symptoms, the frequency of treatment, other drugs and the desired effect. A typical daily dose is about 0.5 to 50 mg per kg body weight for oral administration and about 0.05 to 20 mg for parenteral administration.

The novel compounds of the invention may be administered in the form of conventional solid or liquid pharmaceutical compositions, i.e., coated or uncoated tablets of capsules, powders, granules, suppositories or solutions. These are produced in the usual way. The active ingredients can be combined with conventional pharmaceutical auxiliaries, such as tablet binders, fillers, preservatives, tablet disintegrants, flow regulators, emollients, solvents, delaying agents, antioxidants and / or carrier gases (H. Sucker and co-gases). Pharmazeutische Technologie, Tliieme Verlag, Stuttgart, 1978). The drug forms thus administered contain normally from 1 to 90% by weight of active ingredient.

The following examples illustrate the invention without limiting it. In the examples, protein amino acids are abbreviated in the conventional 3-letter code. Other abbreviations are: Me ₂ Val = Ν, Ν-dimethylvaline, MeVal-N-methylvaline.

DETAILED DESCRIPTION OF THE INVENTION

A. General procedures

I. The peptides of the invention of formula (I) are either synthesized by classical solution synthesis by standard methods described using Z and Boe, or by standard solid-state synthesis methods using Boe and Fmoc protecting groups.

a) Synthesis cycle for the Fmoc protecting group technique.

1. Wash DMF 1 x 1 min.

2. 20% piperidine in DMF 1 x 4 min.

3. 20% piperidine in DMF 1x16 min.

4. Wash DMF 5 x 1 min.

5. Adding a pre-activated guard

1x61 min. 3 x 1 min.

x 1 min.

amino acid (activated with 1 equivalent of TBTU and 5 equivalents of DIPEA in DMF) peptide coupling

6. Wash DMF

7. if the conversion is not complete, repeat the copulation (back to 5th)

8. Wash DMF

9. back to 2.

Reagents ΒΟΡ-Cl and PyBrop are used to coupling amino acids to N-amino acids. The reaction times are increased accordingly by the inclusion of double bonds. In solution synthesis, either Boc-protected (N-carboxy anhydrides) amino acids NCA, Z-protected (N-carboxyanhydrides) amino acids NCA, or pivaloyl chloride as a condensing agent are most preferred for this coupling method.

II. Reductive alkylation of the N-terminus

The Ala resin peptide is deprotected at the N-terminus (steps 2-4 in Ala) and then reacted with a 3-fold molar excess of aldehyde or ketone in dimethylformamide / 1% acetic acid, with 3 equivalents of NaCNBH 3 added. After completion of the reaction (negative Kaiser test), the resin was washed several times with water, isopropanol, dimethylformamide and dichloromethane. For example, reductive alkylation in solution can be achieved by reacting N-terminally deprotected peptides, peptide fragments, or amino acids with the corresponding aldehydes or ketones using NaCNBH ₃ or a hydrogen palladium on carbon system.

III. Treatment of the peptide resins obtained according to paragraphs Ib and II

The peptide resin is dried under reduced pressure and then treated with TFA / water (95: 5) for 1.5 hours (Wade, Tregear, Howard Florey Fmoc Workshop Manual, Melboume, 1985). The resin was filtered off and washed with TFA and dichloromethane. The filtrate and washings were concentrated and the peptide was precipitated by the addition of diethyl ether. After cooling in an ice bath, the precipitate is filtered off, taken up in 30% acetic acid and lyophilized.

IV. When o-chlorotrityl resin (commercial product from Biohellas) is used, the peptide resin suspension in acetic acid / / trifluoroethanol / dichloromethane (1: 1: 3) is stirred at room temperature for 1 hour. The resin is filtered off under suction and washed thoroughly with cleavage solution. The combined filtrates were concentrated in vacuo and treated with ether. The precipitated solid is removed by filtration or centrifugation, washed with diethyl ether and dried under reduced pressure.

V. Purification and characterization of peptides

Purification is by gel chromatography (SEPHADEX G-10, G-15 (10% HOAc, SEPHADEX LH 2 O / MeOH) and / or medium pressure chromatography (stationary phase: HD-SIL C-8, 20-45 micron, 100 Angstrom; mobile phase) gradient with A = 0.1% TFA / water system, B = 0.1% TFA / MeOH system, or preparative HPLC (stationary phase: Waters Delta-Pak C-18, 15 micrometer 100 Angstrom; mobile phase: gradient with A = 0.1% TFA / water system, B = 0.1% TFA / MeOH system).

The purity of the resulting products was determined by analytical HPLC (stationary phase: 100 2.1 mm)

ISSUE C-18 300 Angstrom; mobile phase: acetonitrile / water gradient, buffered with 0.1% TFA, 40 ° C). Characterization of fast atom bombardment by mass spectroscopy and NMR spectroscopy.

B. Specific procedures

Example 1 (SEQ ID NO: 1) Me ₂ Val-Val-MeVal-Pro-L-azetidinyl-2-carboxamide

0.53 g of Fmoc-RINK resin (0.46 mmol / g substitution), corresponding to a dose size of 0.25 mmol as in Ala, is reacted with 0.4 mmol of Fmoc-azetidine-2-carboxylic acid each,

Fmoc-Pro-OH, Fmoc-Val-OH, Fmoc-Val-OH, Fmoc-Val-OH.

The amino acid following the N-methylamino acid is linked by a double bond with the ByProp coupling reagent. After completion of the etheric synthetic cycles, the N-terminal protecting groups (steps 2-4 in Ala) are removed from the peptide resin and further reacted with an aqueous formaldehyde solution as in AII and dried under reduced pressure. The resulting resin was cleaved by TFA as in AIII. The crude product was purified by preparative medium pressure chromatography to give 5 mg of the desired pure peptide (10 to 40% A for 10 '; 40 to 90% A for 200'). The compound is further characterized by fast atom bombardment mass spectroscopy and NMR spectroscopy. [M + H] < ^{+ &gt};.

Example 2 (SEQ ID NO: 1)

Me ₂ V al-V al-MeV al-Pro-3,4-dehydroprolylbenzylamide

a) Z-MeVal-Pro-OMe

66.25 g (250 mmol) of Z-Val-OH are dissolved in 250 ml of dry dichloromethane. After addition of 36.41 ml (262.5 mmol) of triethylamine, the reaction mixture is cooled to -25 ° C and 32.27 ml (262.5 mmol) of pivaloyl chloride are added. After stirring for 2.5 hours, 41.89 g (250 mmol) of H-Pro-OMe x HCl in 250 ml of dichloromethane are added to the reaction mixture and neutralized at 0 ° C with 36.41 ml (262.5 mmol) of triethylamine. . Stirring was continued for 2 hours at -25 ° C and overnight at room temperature. The reaction mixture was diluted with dichloromethane and washed thoroughly with saturated aqueous sodium bicarbonate solution (3x), water (1x), 5% citric acid (3x) and saturated sodium chloride solution. The organic phase is dried over sodium sulphate and evaporated to dryness. The residue (91.24 g) was stirred overnight with petroleum ether and filtered. 62.3 g of product are obtained.

b) H-MeVal-Pro-OMe

48.9 g (130 mmol) of Z-MeVal-Pro-OMe are dissolved in 490 ml of methanol. After addition of 10.9 ml (130 mmol) of concentrated hydrochloric acid and 2.43 g of 10% palladium on carbon, the reaction mixture is hydrogenated. Filtration and evaporation to dryness yielded 36.43 g of product.

c) Z-Val-MeVal-Pro-OMe

In 110 ml of dimethylformamide, 18.1 g of H-MeVal-Pro-OMe, 21.6 g (78 mmol) of Z-Val-N-carboxyanhydride and 22.8 ml (130 mmol) of diisopropylethylamine are stirred at 40 ° C for two days. for two days. After evaporating the dimethylformamide, dichloromethane is added and the organic phase is washed with saturated aqueous sodium bicarbonate solution (3x), water (1x), 5% citric acid (3x) and saturated sodium chloride solution. The organic phase is dried over sodium sulphate and evaporated to dryness. 29.3 g of product are obtained in the form of a viscous oil.

d) H-Val-MeVal-Pro-OMe

29.3 g (61.6 mmol) of Z-Val-MeVal-Pro-OMe are dissolved in 230 ml of methanol. After the addition of 1.15 g of 10% palladium on carbon, the reaction mixture is hydrogenated. Filtration and evaporation to dryness gave 21.96 g of product.

e) Z-Val-Val-MeVal-Pro-OMe

15.29 g (61 mmol) of Z-Val-OH and 21.96 g (61 mmol) of H-Val-MeVal-Pro-OMe are dissolved in 610 ml of dichloromethane and cooled to 0 ° C. After addition of 8.16 ml (73.2 mmol) of N-methylmorpholine, 2.77 g (20.3 mmol) of HOBt and 11.74 g (61 mmol) of EDCI, the reaction mixture is stirred overnight at room temperature, diluted with dichloromethane. and washed thoroughly with saturated aqueous sodium bicarbonate (3x), water (1x), 5% citric acid (3x) and saturated sodium chloride solution. The organic phase is dried over sodium sulphate and evaporated to dryness. 31.96 g of product are obtained.

f) Z-Val-Val-MeVal-Pro-OH

31.96 g (57 mmol) of Z-Val-Val-MeVal-Pro-OMe are dissolved in 250 ml of methanol and 102.6 ml (102.6 mmol) of a 1 M solution of lithium hydroxide in water are added. After stirring at room temperature for 24 hours, water was added and the methanol was evaporated under reduced pressure. The aqueous phase is extracted (3x) with ethyl acetate, adjusted to pH 2 at 4 ° C and extracted (3x) with ethyl acetate. The combined organic extracts were dried over sodium sulfate and the solvent was evaporated under reduced pressure. 30.6 g of product are obtained in the form of a white solid.

g) Me ₂ Val-Val-MeVal-Pro-OH

Ν, Ν-Dimethylated tetrapeptide acid is prepared from a Z-protected tetrapeptide acid by hydrogenation of the Z-protecting group followed by addition of aqueous formaldehyde solution to the hydrogenation mixture to give the desired product in almost quantitative yield.

h) Me ₂ V al-V al-Me N al-Pro-3,4-dehydroproline methyl ester

3.38 g of Me ₂ Val-Val-MeVal-Pro-OH (7.27 mmol) and 0.925 g of 3,4-dehydroproline methyl ester hydrochloride (7.27 mmol) are dissolved in 75 ml of dry dichloromethane. N-methylmorpholine (8.72 mmol), 0.332 HOBt (2.43 mmol) and 1.4 g EDCI (7.27 mmol) are added at 4 ° C. After stirring overnight at room temperature, dichloromethane is added and the organic phase is washed with saturated sodium bicarbonate solution (3x) and water (1x). The organic phase is extracted with 5% aqueous citric acid. The pH of the aqueous acid layer was adjusted to 8 with 1N sodium hydroxide solution and extracted 4 times with dichloromethane. After drying over sodium sulfate and evaporation under reduced pressure, 2.82 g of pentapeptide methyl ester is obtained.

k) Me ₂ Val-Val-MeVal-Pro-3,4-dehydroproline-benzylamide

A solution of 2.0 g of dehydroproline containing pentapeptide methyl ester in 20 ml of methanol is treated at room temperature with 4.6 ml of 1 N lithium hydroxide solution. After completion of the reaction (TLC), water was added, the methanol was evaporated and the compound freed from water by lyophilization. The white powder obtained is dissolved in 36 ml of dichloromethane and 0.388 ml (3.55 mmol) of benzylamide is added. After addition of 0.476 ml of N-methylmorpholine (4.26 mmol), 0.163 g of HOBt (1.19 mmol) and 0.684 g of EDCI (3.55 mmol) at 4 ° C, the reaction mixture is stirred overnight at room temperature. Dichloromethane is added and the organic phase is washed 3 times with saturated sodium bicarbonate solution and water (1x). The organic layer was extracted twice with 5% citric acid. The acidic aqueous layer was adjusted to pH 9 with 5N sodium hydroxide solution and extracted with dichloromethane (4x). After drying over sodium sulfate and evaporation of the solvent under reduced pressure, 600 mg of crude pentapeptide benzylamide is obtained. 300 mg of this crude peptide was purified by liquid medium pressure chromatography (0-10% A in 5 ', 10-25% A in 10', 25-90% A in 450 ') to give analytically pure product. The compound is further characterized by fast atom bombardment mass spectrography ([M + H] ⁺ = 639.4).

The following compounds were prepared and can be prepared according to Examples 1 and 2:

57. Xaa Val Xab Pro Xcd

58. Xaa Val Xab Pro Xca

59. Xaa Val Xab Pro Xcf

60. Xaa Val Xab Pro Xcj

61. Xaa Val Xab for Xeh

62. Xaa val xab Pro Xcl

63. Xaa Val Xab Pro Xck

64. xaa val Xab Xcn xco

65. Xaa Val Xab Xcn Xak

2. Xaa Val Xab Pro Xac

4. Xaa Val Xab Pro Xad

5. Xaa Val Xab for Xae ¢. Xaa Val Xab Pro Xaf 7. Xaa Val Xab Pro Xaj g. Xaa Val Xab Pro Xah

9. Xaa Val Xab Pro Xai

10. Xaa Val Xab Pro Xak

11th Xaa wall xab for Xai 12th Xaa wall xab for Xan 13th Xaa wall xab for Xan 14th Xaa wall xab for Xao 15th Xaa wall xab for XAP 16th Xaa wall xab for Xaq 17th Xaa wall xab for XAR 18th Xaa wall xab for xas 19th Xaa wall xab for Xat 20th Xaa wall xab for XAU 21st Xaa wall xab for Xav 22nd Xaa wall xab for xaw 23rd Xaa wall xab for XAX 24th Xaa wall xab for Xay 25th Xaa wall xab for x and z 26th Xaa wall xab for Xba 27th Xaa wall xab for xbb 28th Xaa wall xab For XBC 29th XBD wall xab for XAR 30th XBE wall xab for XAR 31st Xaa wall xab for xCX 32nd Xaa wall xab for xcy 33rd Xaa Ile xab for XAR 34th Xaa XBH xab for XAR 35th Xaa wall XBF for XAR 36th Xaa wall XBQ for XAR

66th xaa val Xab Xca Xaq 67th Xaa Val Xab Xcs Xar 68th Xaa Val Xab Xca Xay 69th xaa Val xab Xcp Xak 70th Xaa val Xab Xcp Xaq 71st Xcv Val Xab Xcp Xar 72nd Xcx Val Xab Xcp Xay 73rd Xaa val Xab Pro Xcn xer 74th Xaa Val Xab Xcn Xcs 7S. Xaa Val Xab Xcn Xct 76th Xaa Val Xab Xcn Xcn Xcn 77th Xaa val Xab Pro Xcn Xcv 78th Xaa val xab For Xcq Xcr 79th Xaa Val Xab Pro Xcq Xcs 80th Xaa Val Xab Xcq Xct 81st Xaa Val Xab For Xcq Xeti 82nd Xaa Val Xab Xcq xar 83rd xaa val xab for xgv xcx 84th Xaa Val Xab Xcv Xcs 85th Xaa Val Xab Pro Xcv Xct 86th Xaa Val Xab Xcv Xcq 87th Xaa Val Xab Xcv Xcv Xcv 88th Xaa Val Xab Pro Xcv OCSj 89th Xaa Val Xab Pro Xcl 90th Xaa Val Xab for Xat 91st xaa val xab Pro rez 92nd Xaa Val Xab Pro Xda

Examples of MS-characterization of synthesized novel compounds are shown in the following table.

Example

MS analysis in fast atom bombardment

Example Analysis of MS in fast atom bombardment

no. (Mol. Weight (measured))

61 627 64 546 88 552 B9 655

37. Xaa Val Xab Pro Xbi

38. Xaa Val Xab Pro xbk

39. Xaa val Xab Pro Xbl

40. Xaa Val Xab Pro Xbn

41. Xaa Val Xab for Xbn 43. Xaa Val Xab Pro Xbo

43. Xaa Val Xab Pro Xbp

44. Xaa Val Xab Pro Xbq

45. Xaa Val Xab Pro Xbr

46. Xaa Val Xab Pro Xbs

47. Xaa Val Xab Pro Xbt

48. Xaa Val Xab Pro Xbu

49. Xaa Val Xab Pro Xbv

50. Xaa Val Xab Pro Xbv

52. Xaa Val Xab Pro Xbx

52. Xaa Val Xab Pro Xby

53. Xaa Val Xab Pro Xbs

54. Xaa Val Xab Pro Xca

54. Xaa Val Xab Pro Xcb

56. Xaa Val Xab Pro Xcc

Table I - Sequence identification of compounds prepared according to Examples 1 and 2

Compound No.

Sequence ID number

1-32, 35-63, 88-321

332

343

64-724

74-875

The symbols Xaa in sum have the following meanings:

Xaa · N, N-Dimethylvaline

Xab: N-Methylvaline

XAC:

xad:

XAF:

xau:

Xags

Xaii:

Xai:

Xav:

xaw:

X & x:

Xak;

XAL:

Xay:

Xazs

* XM.

Xam:

Xant ⁰ H ₃ C

xbb:

Xao:

XBC:

XAP:

Xaq:

Xari

xas:

Xat;

Xbd: N, N-dimethylisoleucine

Xbe: N, N-dimethyl-2-tert-butyl-glycine

Xbf: N-methylisoleucine

Xbg: N-methyl-2-tert-butyl-glycine

Xbh: 2-tert-butylglycine

F

O HjG

XBK:

XBL:

Xbtn:

Xba:

xbo:

XBP:

XBQ:

XBR:

XBS:

XBT:

Xck: 3,4-dehydroproline-adamantyl (1) -amide

XCK:

FCL. '

3 ₁ < - &gt;-D'-hydropropanedanediyl (1) -amide

XCN: 3,4-dehydroproline XCO: 3,4-dehydroproline amide XCP: L-thiazolidine-4-carbonyl XCQ: 4-fluórprolín

XCR:

XCS:

XCT:

x Cu:

Xcvsam

Xcw: Xcx;

xcy:

XCZ:

ABOUT*.

"" (A / ^m ^ ^{of x} CH ₃ N ^{x 'and} y' _j ^N-CH ₃ OH ° C

Xda:

The compounds of the invention can be tested for anti-cancer activity by conventional methods including, but not limited to:

A. In vitro methodology

Cytotoxicity is measured by standard methodology for adherent cell lines, such as the microculture tetrazolium test (MTT). Details of this assay are described in the literature (Alley, MC et al. Cancer Research 48: 589-501, 1988). Exponentially growing cultures of tumor cells, such as HT-29 colon-carcinoma or LX-lung tumor cells, are used to obtain microtiter plate cultures. Cells are seeded at 5,000 to 20,000 per well in 96-well plates (in 150 μΐ medium) and cultured overnight at 37 ° C. Compounds are added at a 10-fold dilution of 10 ^-4 to 10 ^-1 M. The cells are then incubated for 48 hours. To determine the number of viable cells in each well, add MTT dye (50 μΐ of a 3 mg / ml solution of 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide in brine). The mixture is incubated at 37 ° C for 5 hours and then 50 μΐ 25% SDS, pH 2 is added to each well. After overnight incubation, the absorbance of each well at 550 nm is read by ELISA. Mean +/- SD data from replicated wells were calculated using the% T / C formula (% of treated live cells / control cells).

OD treated cells

------------------------ x 1OD - t T / c OD control cells

The concentration of test compound that gives a T / C growth inhibition value of 50% is referred to as an IC 50 value.

Example compound

I 1 x 1O ⁽ 2 1 X 10 '' 61 1 X 10 ' 64 1 x 10 ¹ 88 1 x 89 1 x 10 ⁱ

B. In vivo methodology

The compounds of the invention were further tested in preclinical assays for in vivo activity indicative of clinical use. Such assays are performed in nude mice transplanted with tumor tissues, preferably of human origin, as is well known in the art. Test compounds are evaluated for anti-tumor efficacy after administration to xenograft mice.

Human breast tumors (MX-1) that grew in athymic nude mice are transplanted into a new host mouse using tumor fragments of about 50 nm in size. The transplant day is day 0. Six to ten days later, mice are treated with test compounds administered as intravenous injections in groups of 5 to 10 mice at each dose. The compounds were administered every other day for three weeks at doses of 1 to 100 mg / kg body weight. Tumor diameters and body weight were measured twice a week. Tumor volumes were calculated from calliper diameters and tumor volumes were calculated for each treatment group according to the formula (length x width ² ) / 2 = mm ³ tumor volume

Mean tumor volumes are calculated for each group and T / C values for each group versus untreated control tumors are determined.

The novel compounds have good tumor inhibition properties.

Industrial usability

A peptide for the manufacture of pharmaceutical compositions for the treatment of mammalian cancer.

SK 282467 Β6

Sequence Listing (1) General Information

(i) Applicant

A. Addressee: BASF Aktiengesellschaft

B. Street: Carl-Bosch-Strasse 38

C. City: Ludwigshafen

E. Country: Federal Republic of Germany

F. ZIP: D-67056

G: Telephone: 0621/6048526

H: Telephone: 0621/6043123

1. Telex: 1762175170 ii) Title of the invention: Peptide and pharmaceutical composition containing it iii) Number of sequences: 5 iv) Form for computer

A. Media Type: 3.5 inch floppy disk, 2 DD

B. Computer: IBM AT-compatible, 80286 processor

C. Operating System: MS-DOS version 5.0

D. Software: WordPerfect

2. Information about SEQ ID NO: I:

i) Sequence characteristics

A. Length: 5 amino acids

B. Type: amino acid

D. Topology: linear ii) Type of molecule: peptide xi) Sequence description: SEQ ID. NO: 1:

Xaa Val Xaa For Xaa

2. SEQ ID. NO: 2:

i) Sequence characteristics

A. Length: 5 amino acids

B. Type: amino acid

D. Topology: linear ii) Type of molecule: peptide xi) Sequence description: SEQ ID. NO: 2:

Xaa White Xaa For Xaa

2. SEQ ID. NO: 3:

i) Sequence characteristics

A. Length: 5 amino acids

B. Type: amino acid

D. Topology: linear ii) Type of molecule: peptide xi) Sequence description: SEQ ID. NO: 3:

Xaa Xaa Xaa For Xaa

2. SEQ ID. NO: 4:

i) Sequence characteristics

A. Length: 5 amino acids

B. Type: amino acid

D. Topology: linear ii) Type of molecule: peptide xi) Sequence description: SEQ ID. NO: 4:

Xaa Val Xaa Xaa Xaa

2. SEQ ID. NO: 5:

i) Sequence characteristics

A. Length: 6 amino acids

B. Type: amino acid

D. Topology: linear ii) Type of molecule: peptide xi) Sequence description: SEQ ID. NO: 5: Xaa Val Xaa For Xaa Xaa

Claims (4)

PATENT CLAIMS 1. Peptides of formula (I) R'R ² N-CHX-CO-A-B-D-E- (F) _t -K (I) where R ^{1 is} methyl, ethyl or isopropyl, R ² is hydrogen, methyl or ethyl, ^R'-NR2 together pyroilidínový ring, And the residues valyl, isoleucyl, leucyl, 2-tert-butylglycyl, 2-ethylglycyl, norleucyl or norvalyl, B residues N-methylvalyl, N-methylleucyl, N-methylisoleucyl, N-methylnorvalyl, N-methylnorleucyl, N-methyl-2-tert-butylglycyl, glycyl, N-methyl-3-tert-butylalanyl or N-methyl-2- ethylglycyl. D residues prolyl, 3,4-dehydroprolyl, 4-fluoropolyl, 4,4-difluoropolyl, azetidine-2-carbonyl, homoprolyl, 3-methylprolyl, 4-methylprolyl, 5-methylpropyl or thiazolidine-4-carbonyl, E 3,4-dehydroprolyl, 4-fluorprolyl, 3-methylprolyl, 4-methylprolyl, azetidine-2-carbonyl or 4,4-difluoroprolyl, F residues valyl, 2-tert-butylglycyl, isoleucyl, leucyl, 2-cyclohexylglycyl, norleucyl, norvalyl, neopentylglycyl, and -lanyl, β-alanyl or aminoisobutyroyl, X is preferably alkyl of 2 to 5 carbon atoms, cyclopropyl or cyclopentyl, t is 0 or 1; The alkoxy group preferably having 1 to 4 carbon atoms, the benzyloxy group or the amino group of the formula RNR ⁶ , where R ⁵ is hydrogen or hydroxy or C ^ -alkoxy or benzyloxy, optionally substituted with up to two substituents each independently selected from: CF _{3 J} nitro, alkylsulfonyl C _in, C _M alkoxy, by _halogen, alkyl, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or R ⁵ is phenyloxy optionally substituted with up to two substituents independently selected from CF ₃ , nitro, C ₁ . ₄ alkylsulfonyl, C _1-4 alkoxy, halogen, C _1-4 alkyl, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , R ^{6 is} hydrogen or Cn ₂ alkyl, optionally substituted with at least one fluorine atom, or - (CH ₃ ) ₂ CN, -C (CH ₃ ) ₂ CCH, -C (CH ₃ ) ₂ C = CH ₂ , -C (CH ₃ ) ₂ CH ₂ OH, -C (CH ₃ ) ₂ CH ₂ CH ₂ OH or - (CH ₂ ) _in C ₃ . ₇ cycloalkyl wherein v is 0, 1 or 2 and which is optionally substituted with methyl, or is norephedryl, norpseudoephedryl, quinolyl, pyrazyl, adamantyl, -CH ₂ -benzimidazolyl, -CH ₂ -adamantyl, d-methylbenzyl, d-dimethylbenzyl, - (CH ₂ ) _v phenyl, wherein v is 0, 1, 2 or 3 and which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, Cl. ₄ alkylsulfonyl, C |. ₄ alkoxy, halogen, _C-alkyl which may form a cyclic system, cyano, hydroxy, N (CH _{3) 2,} COOMe, COOEt, COOiPr, and COONH _2, or is - (CH ^ aífyl, m is 0 or 1, and which is optionally substituted by one or two substituents each independently selected from: CF _3, nitro, _{C1. 4} alkylsulfonyl, C |. ₄ alkoxy, halogen, C _M alkyl which may form a cyclic system, cyano, hydroxy, N ( CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or - (CH ₂ ) _w benzhydryl, wherein w is 0, 1 or 2, and is optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C _{l. 4} alkyl sulfonyl, CMalkoxyskupinu, halogen, _C-alkyl, KTOI may form a cyclic system, cyano, hydroxy, N (CH _{3) 2,} COOMe, COOEt, COOiPr, or COONH _2; or is biphenyl, optionally substituent One or two substituents independently selected from the group consisting of CF ₃ , nitro, C _1-4 alkylsulfonyl, C _1-4 alkoxy, halogen, C _1-4 alkyl, which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is pyridyl, which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 _{alkylsulfonyl} , C _1-6 -alkoxy, halogen, C 1-4 alkyl, which may form a cyclic system, cyano , hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is picolyl optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C _1-4 alkyl, which may form cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is -CH ₂ -CH ₂ -pyridyl, which is optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-6 alkyl; ₄ alkoxy, halo, C _M alkyl which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr, or COONH _2, or is a benzothiazolyl, KTOI is optionally substituted by one or two substituents each independently selected from: CF _3, nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl, which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is benzoisothiazolyl optionally substituted with one or two substituents each independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is benzopyrazolyl optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl, which may form a cyclic system, cyano, hydroxy , N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is benzooxyzolyl optionally substituted with one or two substituents independently selected from CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl, can form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is - (CH ₂ ) _m -fluorenyl, wherein m is 0 or 1, which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen , C _M alkyl, KTOI may form a cyclic system, cyano, hydroxy, N (CH _{3) 2,} COOMe, COOEt, COOiPr, or COONH _2, or is pyrimidyl, which is optionally substituted by one or two substituents each independently selected from: CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which can form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is - (CH ₂ ) _m -indanyl, wherein m is 0 or 1, which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, N (CH ₃ ) ₂ , COOMe, COOEt, COOiPr or COONH ₂ , or is - (CH ₂ CH ₂ O) _y CH ₃ , wherein y is 0, 1, 2, 3, 4, or 5, or - (CH ₂ CH ₂ O) _y CH ₂ CH ₃ , wherein y is 0, 1, 2, 3, 4 or 5, or is -NH-QHs, which is optionally substituted with one or two substituents independently selected from the group consisting of: including CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl, which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is -NCH ₃ -C ₆ H ₃ optionally substituted by one or two substituents independently selected from the group consisting of CF ₃ , nitro, C 1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or -NH-CH ₂ -C ₆ H ₅ , which is optionally substituted with one or two substituents independently selected from the group consisting of CF ₃ , nitro, C _1-4 alkylsulfonyl, C 1-4 alkoxy, halogen, C 1-4 alkyl, which may be to form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr or COONH ₂ , or is -NCH ₃ -CH ₂ -C ₆ H ₅ optionally substituted by one or two substituents independently selected from CF ₃ , nitro, C ₃ . ₄ alkylsulfonyl, CMalkoxyskupinu, halo, CMalkyl, which may form a cyclic system, cyano, hydroxy, COOMe, COOEt, COOiPr, or COONH _2, or is a five heteroaiyl, which is optionally substituted by one or two substituents each independently selected from: CF ₃ , nitro, C (. ₄ alkylsulfonyl, CMalkoxyskupinu, thiomethyl, thioethyl, picolyl, acetyl, C ₃ cycloalkyl, thiophenyl, CH ₂ COOEt, C _{3. 4} alkylene forming a bicyclic system with the heterocycle, phenyl which is optionally substituted by one or two substituents independently selected from nitro, CF ₃ , cyano, halogen or C 1-4 alkyl, and further comprising benzyl optionally substituted by one or two substituents independently selected from nitro, CF ₃ , halogen, C 1-4 alkylsulfonyl. , cyano, hydroxy, C 1-4 dialkylamino, or z general represents -CHR ⁷ membered heteroaryl, optionally substituted with one or two substituents each independently selected from: CF _3, nitro, cyano, halogen, COOMe, COOEt, COOiPr, and COONH _2, CMalkyl, CMalkoxyskupinu, phenyl, benzyl, naphthyl or C |. ₄ alkylsulfonyl, wherein R ⁷ is hydrogen, C] _salkyl straight or branched chain, or benzyl, or a salt thereof with physiologically tolerated acids. Peptides of formula (I) according to claim 1, or salts thereof, for use in medicine, in particular for the treatment of cancer. 3. A pharmaceutical composition comprising a therapeutically effective amount of a peptide of formula (I) according to claim 1 and a pharmaceutically acceptable carrier. Use of the peptides of formula (I) according to claim 1 for the preparation of a medicament for the treatment of mammalian tumors.