SK3893A3 - Bombesine antagonists - Google Patents

Abstract

The present invention provides a polypeptide of the formula (I): XX<1>TrpX<2>X<3>X<4>X<5>X<6>X<7>NH2, wherein X is a group X<8>Arg(or D-Arg)X<9>X<10> and X<8> is des NH2Pro,TyrPro,des NH2TyrPro, Ada, Pro, D-Pro or is deleted; X<9> is Gly, Ala, D-Ala or is deleted; X<10> is Asn, Phe, D-Phe, or Phe or D-Phe substituted by one or more halo atoms; or X is a group A-(CH2)n-CO- in which A is a group containing 1 to 3 rings of which at least one ring is aromatic, each ring system being optionally substituted; and the alkylene group is optionally substituted by one to four groups selected from amino, hydroxy, C1-4 alkoxy and C1-4 alkyl optionally substituted by halo and n is 0 to 4; or X is cyclopentyl carbonyl substituted by a group X<8>Arg(or D-Arg)X<9>X<10> as hereinbefore defined; X<1> is His, ThiAla or is deleted; X<2> is Ala, D-Ala, CPenc, D-tBuGly or Pro; X<3> is Val or Val substituted by one or more halo atoms; X<4> is Gly, Ala, D-Ala, Sarcosine, Pro, D-Pro or D-Phe; X<5> is His or ThiAla; X<6> is D-Pro psi , Pro psi , 2-pyrrolidinyl-3-hydroxypropionyl or D-Pro; X<7> is Nle, Leu, Phe, Val, Mox, D-Phe, Phe, or D-Phe substituted by one or more halo atoms or naphthylAla or naphthyl D-Ala or a hydrophobic, substituted aromatic amino acid or aralkylamine or is deleted, and the pharmaceutically acceptable salts thereof. The polypeptides possess antagonist properties against bombesin-like peptides and are of value in the treatment of malignant disease.

Description

Technical field

Ohio / Ull '. ·. ·

1 'avto: · ·.

!

Ϊ CO E9.r0.82 33200 fr

H ....

The invention relates to polypeptide compounds having antagonistic properties against bombesin or bombesinoid peptides and useful in the treatment of diseases, in particular human small cell fungal cancer, Zollinger-Ellison syndrome or pancreatic cancer. Thus, the invention encompasses polypeptides, processes for their manufacture, pharmaceutical compositions containing them, and their use in medicine.

BACKGROUND OF THE INVENTION

Bombesin is a tetradecapeptide, originally isolated from the skin of a frog. It has a formula

Glp-Gln-Arg-Gly-Asn-Gln-Trp-Ala-Val-Gl, γ-His-Leu-Met-NH ₂ .

The gastrin-releasing peptide is a 27 amino acid peptide isolated from porcine gut. The last ten amino acids at the C-terminus of the gastrin releasing peptide correspond to the amino acid variation (3) in the last ten amino acids of bombesin, namely:

H-Gly-Asn-His-Trp-Ala-Val-Gly-His-Leu-Met-NH _second

It has been reported (JHWalsh and JRReeve, Peptides 6, (3), 63-68, (1985) that bombesin and bombesinoid peptides such as gastrin releasing peptide are secreted by human cells in small cell lung cancer. PJWoll and E Rosengurt, PNAS 85, 1859-1863 (1988)) that gastrin releasing factor antagonists bind competitively to the bombesin receptors in animals and are therefore useful in the treatment of small cell lung cancer (SCLC) and / or in the management of clinical symptoms associated with Bombesin analogs have been shown to inhibit the binding of gastrin releasing peptide to the SCLC cell line and inhibit the growth of SCLC cells in vitro and in vivo (S. Mahmoud et al., Cancer Research, 51, 1798-1802). 1991 /).

The different been published bombesin antagonists, e.g., / Leu ¹³ - Ψ (CH2 -NH) Leu ^{1 4} _7bombesin and / ^_ -Φ ⁹ Al (CH2-NH) Leu-Val ^{1 0 14} _ / bombesin (Coy, et al. J (Chem., 1988, 263, 5056) and 4-pyridyl-CO-His-Trp-Ala-Val-D-Ala-His-Leu-0Me,

4-pyridyl-CO-His-Trp-Ala-Val-D

4-pyridyl-CO-His-Trp-Ala-Val-D

3-Pyridyl-CO-His-Trp-Ala-Val-D

4-Pyridyl-CO-His-Trp-Ala-Val-D

3-Indolyl-Co-His-Trp-Ala-Val-D

4-Pyridyl-CO-His-Trp-Ala-Val-D 4-Pyridyl-CO-His-Trp-Ala-Val-D 4-Pyridyl-CO-His-Trp-Ala-Val-D 4-Pyridyl-CO His-Trp-Ala-Val-D

Ala-His-Leu-NHMe,

Ala-His-MeLeu-OMe,

Ala-His-MeLeu-OMe,

-Ala-Lys (Z) -MeLeu-OMe, -Ala-His-Leu-OMe, -Ala-His-MeLeu-NHMe, -Ala-Lys (Z) -Leu-NHMe, Ala-Lys (COCH ₂ Ph) -Leu-NHMe and Ala-Lys (COCH ₂ CH ₂ Ph) -Leu-NHMe.

European patent application no. 34599OA /.

In formula I below and throughout this specification, amino acid moieties are designated by their standard abbreviations (Pure and Applied Chemistry, 1974, 40, 317-331; European Journal of Biochemistry, 1984, 138, 9-37).

For the avoidance of doubt, amino acid symbols indicate the L-configuration unless otherwise indicated by the D or DL appearing before the symbol and separated from this symbol by a dash. (R) and / S / are standard designations for molecular alignment.

- 3 Use! with the following abbreviations:

Τ '= canine (CH ₂ NH)

Ada = 1-Adamantanecarboxylic acid

CPenc - aminoeyclopentanecarboxylic acid

Mox = methoxinin

Des-NH ₂ Pro = 1-cyclopentanecarboxylic acid

Des-NH ₂ Tyr = (4'-hydroxy) -3-phenylpropionic acid ThiAla with 3- (2-thienyl) -alanine

D-tBuGly = D-tertiary butylglycine (tertiary-D-leucine).

When Ada, CPenc, Mox, Des-NH ₂ Pro, Des-NH ₂ Tyr are groups in the polypeptide chain, they are in carbonyl form.

SUMMARY OF THE INVENTION

It has now been found that another group of polypeptides has potent antagonistic activity against bombesin.

The compounds of the invention inhibit the formation of gastrin-releasing peptide in mammalian cells and are therefore useful in suppressing the clinical symptoms of diseases that induce gastrin. secretion or hypersecretion of gastrin-releasing peptide (e.g., SCLC).

Accordingly, the present invention provides a polypeptide of Formula I:

XX ¹ TrpX ² X ³ X ⁴ X ⁵ X ⁶ X ⁷ NH ₂ '

X is X ⁸ Arg (or 1'-Arg) X 1 X X a

^X8 is des-NHgPro, TyrPro, Des-BH ₂ TyrPro, Ada, Pro, D-Pro or is deleted;

- 4 ο

X is Gly, Ala, D-Ala or is deleted;

Χ ¹ θ is Asn, Phe, D-Phe or Phe or D-Phe substituted with one or more halogen atoms; the sky

X is A (CH ₂ ) _n -CO- wherein A is a group containing 1 to 3 rings, of which at least one ring is aromatic, each ring system optionally substituted; and the alkylene group is optionally substituted with one or four groups selected from amino, hydroxyC 1-6 -alkoxy and C 1-6 alkyl optionally substituted with halogen and n is 0 to 4; the sky

X is ΑΑ-)COCO wherein A is an optionally substituted 1 to 3 ring aromatic radical and the alkylene group is optionally substituted with one or four groups selected from amino, C 1-4 alkoxy and C 1-4 alkyl. alkyl, optionally substituted with halogen and n is 1 to 4, or g

X is cyclopentylcarbonyl substituted with X; Arg (or D-Arg) X 1 X ¹⁰ are as defined above;

χ1 is Hie, ThiAla, or is deleted;

X ² is Ala, D-Ala, CPenc, D-t BuGly or Pro;

X 1 is Val or Val substituted with one or more halogen atoms;

X 1 is Gly, Ala, D-Ala, sarcosine, Pro, D-Pro or D-Phe;

X is His or ThiAla;

Χ 6 is D-Pro 3 Z, Pro Ψ, 2-pyrrolidinyl-3-hydroxypropionyl or D-Pro;

X? is Nie, Leu, Phe, Val, Mox, D-Phe or Phe, or

D-Phe substituted with one or more halogen atoms or nontytylAla or naphthyl-D-Ala or a hydrophobic substituted atomatic amino acid or aralkylamine or is deleted.

Preferably A is phenyl, naphthyl, phenothiazinyl, indolyl. A is phenyl or naphthyl.

Suitable substituents for the aromatic ring Ar include hydroxy, phenyl, halogen, C ₁ _ alkyl or C ^^ alkoxy optionally substituted by halogen.

Preferably n is 2.

Preferred X ends with DesNH ₂ counts.

Preferred X ⁸ is DesNHgTyr or DesNH ₂ Pro. Preferably X 1 is Gly or Ala-Ala. Preferred is D-Phe. Preferred C is DesNH ₂ Phe, LesNH ₂ Tyr, DesNH ₂ TyrPro (or D-Pro) Arg (or D-Arg).

Preferably X 1 is His or ThiAla, preferred X 1 is Ala, Pro, preferred X is Val or hexafluorvalm, and preferred X is D-Ala or D-Phe,

Preferably X 1 is His or ThiAla, preferred X ⁶ is D-Pro Ψ, Pro ¥, Pro, D-Pro, preferably X 1 is N nebo or Phe, Leu, methoxinin, 2-naphthyl -2-2-alanine.

Preferred polypeptides of the invention include:

N- ((R) -2- (6-methyl-2-t.hoxy nef tyl) propyl they 1) -HisTrpAlaValD-AlaHisĽ the _W-Nle-NH2,

N - ((S) -2- (6-methoxy-2-naphthyl) propionyl) -HisTrpAlaValD-AlaHisD-ProWle-NHg,

N - ((S) -3-phenyl-butyryl) -HisTrpAlaValD-AlaHisD-Pro-Nle Φ NH _2,

N - ((R) -3-phenyl-butyryl) -HisTrpAlaValD-AlaHisD-Pro _Nle-NH2,

N - ((3-phenyl) propyl ony1) sTŕpAlaValD -His-Ala (3- (2-Thi) -Ala) Ψ DPro _Nle-NH2 ^y '

N - ((S) -3,3,3-trifluoro-2-methoxy-2-phenyl-propionyl) -HisTrpAlaValD-Pro Ψ _Nle-NH2,

- N 6 - ((R) -3,3,3-trifluoroacetyl-2-methoxy-2-methoxy-2-phenyl-propionyl) -HisTrpAlaValD-Pro Ύ _Nle-NH2,

N-3 - (((4'-hydroxy) phenyl) propyl sulfonyl) -PROL-ArgGlyD-PheHi sTrpAlaValGlyHisD to _¥-Nle-NH2,

N - (((4'hydroxy) -3-phenyl) propionyl) -Prop-ArgHisTrpAlaVaipAlaHisD-Pro ¥ Leu-NHg

N - ((3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro SP mox-NHg

N - ((3-ŕenyl) propionyl) HisTrpAlaVaip SPPH-Pro-NH _2,

N - ((3-phenyl) propionyl) -TrpAlaVal 4 -AlaHisD-Pro SP Leu-NH ₂ ,

N - ((3-phenyl) propionyl) HisTrpProValD-ProHisD-Pro SP Leu> NH ₂ ,

N-3 - (((3 &apos; triŕluormethyl) phenyl) propionyl) -HisTrpAlaValD-AlaHisD to ¥ Leu-NH _2,

N - ((3-ŕenyl) propiony1) - (3- (2-Thi) -Ala) TrpAlaValD AlaHisDPr.o Ϋ-Leu-NH _2,

N - ((Deamino-Pro) -D-ArgD-AlaD-PheHisTrpAlaValGlyHisD-ProΨ Nle-NH ₂

N - ((3-ŕenyl) propionyl) -HisTrpAlaValGlyHisD _Pro-Nle-NH2,

N - ((deamino-Pro) -D-RGD-ALAD-PheHisTrpAlaValD _{AlaHisDPro-Nle-NH2,}

N '- ((3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro-Nle-NH 4 N - ((3-phenyl) propionyl) -HisTrpAlaVal'-AlaHisD-Pro SP Nle-NH ₉ TyrProD-ArgGlyD-PheHisTrpAlaValGly Ϋ _Nle-NH2 D-ArgGlyD-PheHisTrpAlaValGlyHisD _SP-Nle-NH2,

N - ((3-ŕenyl) Propionyl) -HisTrpAlaValD-AlaHisD-prophy-NH _2,

N - ((3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro SP (2-naphthyl) -D-Ala) -NH ₂ ,

N - ((3-phenyl) -propionyl) -HisTrpAlaValD-PheHisD-Pro SP Phe-NH. _? D-PheHisTrpAlaValD-AlaHisD-Pro SP Phe-NH _2,

- 7 NC (3-Phenyl) propionyl) -L-ProArgGlyD-PheHi sTrpAlaValDAlaHisDProT Phe-NH ₂ ,

N- ((3-Phenyl) propionyl) - (3- (2-Thi) -Ala) TrpAlaValD-AlaHisDΡγοΨΨ Phe-NH ₂ y

N - ((3-phenyl) propionyl) -HisTrpAlaVal- (sarcosine) -HisD Why on-Phe-NH _2,

N-3 - (((4'-hydroxy) phenyl) propionyl) -HisTrpAlaValD-AlaHisDProPhe-NHgj

N - (((2 ', 6'-dichloro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisDProSf Nle-NH ₂ ,

N (((3 ', 4'-dichloro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisDPro-Nle-NH ₂ ,

N - (((4'-Hydroxy) -2-Phenyl) Acetyl) -HisTrpAlaValD-AlaHisDPro _Nle-NH2,

N- (1-naphthyl) -HisTrpAlaVaip-AlaHisD-ProNle-NH _2,

N - ((3,7-dihydroxy) -2-naphthyl) -HisTrpAlaValD-Pro _Nle-NH2,

N - (((3,4-dihydroxy) -2-phenyl) acetyl) -HisTrpAlaVaip-AlaHisD-Pro Ϋ _Nle-NH2,

N- (2- (3-pyridyl) acetyl) -HisTrpAlaValD-Pro SF _Nle-NH2,

N- (2- (2-thienyl) acetyl) -HisTrpAlaValD-AlaHisL-Pro SF Nle-NH ₂

N- (((3-fluoro) -3-phenyl) propyl-benzenesulfonyl) -His-sTrpAlaValD AlaHisDPro * _Nle-NH2,

N - (((4-hydroxy-3-methoxy) -2-phenyl) acetyl) -HisTrpÁlaValDAlaHisD The _Ψ-Nle-NH2,

N - (((R) - (-) - 2-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-ProΨ Phe-NH ₂

N - (((S) - (+) - 2-phenyl) propyl ony1) -HisTrpAlaValD-Allah with D-Phe-NH ProΨ _second, θ _ _

N - (((trans) -2-phenyl) -cyclopropanoyl) -HisTrpAlaVaip-AlaHishD-Pro-Phe-NYHg,

N- (3-O-Renothiazinyl) propionyl-HisTrpAlaValD-AlaHisDPro & Phe-NH ₂

N - ((3-methyl-3-phenyl) butyl) -HisTrpAlaValD-AlaHisD-Pro SF Phe-NH _2,

N - (((2'-trifluoromethyl) 2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro # _Phe-NH2,

N - (((3 '-trifluoromethyl) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro SF Phe-NBL ·,,

N - (((4'-trifluoromethyl) -2-Phenyl) Acetyl) -HisTrpAlaValD-AlaHisD-Pro Phe-NH _2,

N - (C (2 ', 3'-Difluoro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisPro for Phe-NH ₂

N - (((2 ', 4'-difluoro) -2-phenyl) acetyl) HisTrpAlaValD-AlaHisDPro-Phe-NH ₂ ,

N - (((2 ', 6'-difluoro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisDPro Phe-NH ₂ ,

N - (((2-amino) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro SF Phe-NH _2,

N) 1-naŕtoyl) -HisTrpAlaValD-AlaHisD-ProΎ Phe-NH _2,

N - (((3 ', 4', 5'-trimethoxy) 3-phenyl) propionyl) -HisTrpAlaVaipAlaHisD-ProPhe-NH ₂ ,

N - ((6'-Methoxy) -2- (2-naŕtoyl) Propionyl) -HisTrpAlaValD-AlaHisD-Pro T Phe-NH _2,

N - (((3'-trifluoromethyl) -3-phenyl) propionyl) -HisTrpAlaValDAlaHisD-Pro SF Phe-NH ₂ ,

N - (((S) -3-phenyl) butyryl) -HisTrpAlaValD-AlaHisD-Pro Phe-NH ₂

N - (((4'-Methoxy) -3-Phenyl) Propionyl) -HisTrpAlaVaip-AlaHisDPro ^ Phe-NH _2,

- 9 N - ((((S) -2-hydroxy) -2-Phenyl) Acetyl) -HisTrpAlaValD-AlaHisD_Pro Phe-NH _2,

N - ((3-Phenyl) Propionyl) -HisTrpAlaValD-AlaHisProΨ Phe-NH _2,

N - ((2-methy1-2-phenyl) -propionyl) -HisTrpAlaValD-AlaHisD-Pro # _Phe-NH2,

N- (3-0-naphthyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro Ψ Phe-NH _2,

N - (((R) -3-phenyl) butyl) -HisTrpAlaValD-AlaHisD-Pro 'L -Phe-NH _2,

N - ((9-fluorenoyl) 1-carbonyl) -HisTrpAlaValD-AlaHisD-Pro S / Phe-NH _2,

N - (((2'-methoxy) -3-phenyl) propionyl) -Hi sTrpAlaValD-AlaHi with DPro-Phe-NH ₂ ,

N - (((2 ', 5'-dimethoxy) -3-phenyl) propionyl) -HisTrpAlaValDAlaHisD-Pro'5' Phe-NH ₂ ,

N - ((3-Phenyl) Propionyl) -HisTrpAlaValD-AlaHisD-Pro "» y Tyr-NH _2,

N - (((2 ', 3'-dimethoxy) -3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-ProΤ' Phe-NH ₂

N - ((3-phenyl) propionyl) -HisTrpAlaValD-AlaHis (3- (2-pyrrolinyl-3-hydroxy) propionyl) -Phe-NH ₂ ((Isoquinolylcarbonyl) -HisTrpAlaValD-AlaHisD-Pro 'Phe-NH ₂ , N - ((3-phenyl) propionyl) -HisTrpAla 't' Valle AlaHisD-Pro-Phe-Ψ NH _2,

Pharmaceutically acceptable salts or prodrugs of the polypeptides of Formula I are also included within the scope of this invention. Suitable pharmaceutically acceptable salts are acid addition salts when the polypeptide is sufficiently basic, for example when it contains one or more basic moieties such as histidine.

A suitable pharmaceutically acceptable acid addition salt of the invention may be formed with an inorganic acid such as hydrochloric acid, hydrobromic acid, sulfuric acid or phosphoric acid or an organic acid such as acetic, citric, maleic, fumaric, succinic, tartaric or trifluoroacetic acid.

As a further feature, the invention encompasses any one or more preferred compounds together with their pharmaceutically acceptable acid addition salts.

The polypeptides of the invention can be prepared by any of the methods well known in the art. peptide chemistry that is acceptable for the synthesis of analogous compounds. For example, a polypeptide of the invention may be obtained by procedures analogous to those published in Solid Phase Peptide.Synthesis by Stewart and Xoung (published by Pierce Chemical Company, Illinois, 1984), Principles of Peptide Synthesis (published by Springer-Verlag, Berlin, 1984) ), practice of Peptide Synthesis (published by Springer ^ Verlag, Berlin, 1984) and The Synthesis of a Tetrapeptide (J. Am. Chem. Soc., 83 2149 (1963)).

Preferred processes for making the polypeptides of the invention include, for example:

(a) removing one or more conventional peptide protecting groups from the protected polypeptide to obtain a polypeptide of the invention of Formula I;

(b) forming an amide bond by linking two peptide units, one containing a carboxylic acid group or a reactive derivative thereof and the other containing an amino group, so that a protected or unprotected polypeptide having the sequence shown in Formula I is thereby formed, and then, if necessary, deprotected by the above procedure.

In process (a) there may be as many protecting groups in the starting material as there are groups that may need protection, for example some or all of those groups that exist in the product as free hydroxy or basic amino groups (either primary or secondary amino groups). The protecting group or groups may be selected from those described in the standard peptide chemistry textbooks set forth above. Erosion methods for removing the protecting group or groups are also described in other books.

In process (a) a suitable protecting group for a basic amino group (either at the N-terminus or in an amino acid side chain) is, for example, an arylmethoxycarbonyl group which can be removed by hydrogenation over a catalyst, for example palladium on charcoal or can be removed by inorganic treatment an acid such as anhydrous hydrogen fluoride or hydrogen bromide.

In process (a), a particularly suitable protecting group for a basic amino group is, for example, an alkoxycarbonyl group, for example a Boc group, which can be removed by treatment with an organic acid such as trifluoroacetic acid or can be removed by treatment with an inorganic acid such as anhydrous chloroform or hydrogen bromide; or, for example, a 9-fluorenylmethoxycarbonyl group which may be removed by treatment with an organic base, for example piperidine.

In process (a), a suitable amino-protecting group in the histidine side chain is, for example, an arylsulfonyl group, for example a tosyl group, which can be removed by treatment with a hydroxylamine such as N-hydroxytriazole, especially 1-hydroxybenzotriazole, benzyloxymethyl or t-butyloxymethyl.

In process (a), a suitable hydroxy protecting group is, for example, an arylmethyl group, for example, a benzyl group which can be removed by treatment with an inorganic acid, for example anhydrous hydrogen fluoride, or can be removed by hydrogenation over a catalyst, for example palladium on charcoal; or it can be, for example, an esterification group, for example an acetyl or benzoyl group, which can be removed by hydrolysis with a base, for example sodium hyfiroxide.

In process (a), a suitable carboxy protecting group is, for example, an esterifying group, for example an arylmethyl group, for example a benzyl group, which can be removed by treatment with an inorganic group, for example anhydrous hydrogen fluoride or can be removed by hydrogenation over a catalyst, for example palladium on charcoal; or an alkyl group, a C 1-4 alkyl group, for example a tert-butyl group, which can be removed by treatment with an organic acid, for example trifluoroacetic acid.

In process (a), a particularly suitable protection for the carboxyl group at the C-terminus is the formation of, for example, an ester, for example an ester formed by combining the C-terminus of an amino acid and a resin, for example hydroxymethylated styrene-divinylbenzene crosslinked resins; or by forming, for example, an amide, for example, an amide formed by combining the C-terminus of an amino acid and a resin, for example, methylbenzhydrylaminatedyrene-divinylbenzene crosslinked resin.

In process (b), any standard peptide binding reaction may be used, for example one that is described together with others in the standard peptide chemistry textbooks mentioned above.

In process (b) it will be appreciated that the peptide unit may contain exactly one protected or unprotected amino acid.

In process (b), a suitable coupling reaction is, for example, a solvent phase coupling reaction, for example an active ester bond, an azide bond or a bond comprising N, N'-dicyclohexylcarbodiimide, 1-hydroxybenzotriazole and BOP (benzotriazol-1-yl-oxy-tris- ( dimethylamino) phosphonium.

In process (b), a suitable reactive derivative of a carboxylic acid group containing peptide unit is, for example, an acyl halide, for example an acyl chloride formed by the reaction of an acid and an inorganic acid chloride, for example thionyl chloride; a mixed anhydride, for example an anhydride formed by the reaction of an acid and a haloformate, for example isobutyl chloroformate; or an acyl azide, for example an azide formed by the reaction of an oxygen and an azide such as diphenylphosphoryl azide.

In process (b) it is a particularly suitable reactive derivative

A peptide unit containing a carboxylic acid group, for example, the reaction product of an acid and a carbodiimide, for example N, N'-dicyclohexylcarbodiimide or N, N'-diisopropylcarbodimide, or is the product of the reaction of an acid and N-hydroxytriazole, for example. 1-hydroxybenzotriazole and carbodiimide, for example N, N'-dicyclohexylcarbodiimide or N, N'-diisopropylcarbodiimide.

In process (b), a preferred strategy is, for example, to use solid phase synthesis wherein the amino acid to be G-terminal amino acid of the polypeptide of the invention is protected in the alpha-amino group and when needed in a side chain and attached to the solid support e.g. a resin, for example, a hydroxymethylated or methylbenzhydrylaminatedyrene-divinylbenzene crosslinking resin using an ester or amide resin »

The protecting group on the alpha-amino group is then removed. The amino acid to be attached to the C-terminus of the amino acid is protected in the alpha-amino group, and when needed in the side chain, and attached to the C-terminus of the amino acid that remains attached to the solid support. The sequential method of alpha-amino deprotection and attachment to another amino acid is repeated to provide a protected or unprotected polypeptide coupled to a solid support.

The protected or unprotected polypeptide may be released from a hydroxymethyl resin solid resin, for example by hydrolysis, for example by acid hydrolysis, for example by an organic acid, for example trifluoroacetic acid or for example by an inorganic acid or for example anhydrous hydrogen fluoride or hydrogen bromide; or the polypeptide is released, for example, by alcoholysis, e.g. by methanolysis in the presence of a base, for example an organic base, for example diisopropylethylamine, and then, if necessary, deprotecting with the aid of process (a) above.

When methylbenzhydrylamine resin is used, the protected or unprotected polypeptide can be released from the solid support by, for example: treatment with an inorganic acid such as hydrogen fluoride, whereupon, if necessary, protecting groups are removed as described above (a).

In process (b), another preferred strategy is, for example, to use solid phase synthesis wherein the amino acid to be linked to the amino acid chain of the polypeptide of the invention is protected in the alpha-amino group and when needed in the side chain and attached to the solid support. for example to a resin as described above, whereupon the alpha-amino protecting group is removed. Amino acid that l

it is to be attached and to the amino group which is bound to the solid support, is protected in the alpha-amino group and, when needed in the side chain, and joined to the amino acid which is attached to the solid support. The sequential method of alpha-amino deprotection and attachment to another amino acid is repeated to provide a protected or unprotected polypeptide attached to a solid flame retardant.

The protected or unprotected polypeptide can be released from the solid support, for example, using one of the methods described above, whereupon another peptide unit can be coupled using a solution phase coupling reaction as described for process (b) and then, if necessary, protecting groups is removed using the above procedure (a).

The polypeptides of the invention have an antagonistic effect against bombesin, which can be demonstrated by their ability to inhibit bombesin C-stimulated mitogenesis of mouse Swiss 3T3 fibroplastic cells as determined by the capture of (1 H) -thymidine.

According to another feature of the present invention there is provided a pharmaceutical composition comprising a polypeptide of Formula I or a pharmaceutically acceptable salt thereof in association with a pharmaceutically acceptable diluent or carrier.

The composition may be in a form suitable for oral use, for example, in the form of a tablet, capsule, aqueous or oily solution, suspension or emulsion; for nasal use, for example, as a snuff, nasal spray or nasal drops; for vaginal or rectal use, for example, a suppository; for administration by inhalation, for example, as a finely divided powder or a liquid aerosol; for sublingual or buccal use, for example, tablets or capsules, and for parenteral use (including intravenous, subcutaneous, intramuscular, intravascular or infusion), for example, as a sterile aqueous or oily solution or suspension.

In general, the above compositions may be prepared in a conventional manner using conventional bases. However, in the case of a composition for oral administration, it may be appropriate for the composition to include a coating to protect the polypeptide active ingredient from the action of enzymes in the stomach.

The composition of the invention may also contain, in addition to the polypeptide of the invention, one or more known antitumor agents such as mitotic inhibitors, for example vinblastine; alkylating agents such as cis-platin, carboplatin and cyclophosphamide; antimetabolites such as 5-fluorouracil, cytosine arabinoside and hydroxyurea; intercalating antibiotics such as adriamycin and bleomycin; enzymes such as aspartaginase, topoisomerase inhibitors such as etoposide, and biological response modifiers such as interferon.

A preferred composition of the invention is, for example, a composition suitable for oral administration in a unit dosage form, for example tablets or capsules containing from 2.5 to 500 mg and preferably 10 to 100 mg of polypeptide per unit dose, or a composition suitable for parenteral administration comprising from 0.5 to 100 mg of polypeptide per ml and preferably 1 to 10 mg of polypeptide per ml of solution.

A parenteral composition is preferably a solution of isotonic saline or isotonic dextrose buffered when needed at a pH of 5 to 9. Alternatively, the parenteral composition may be a slow release composition in which case the amount of polypeptide per unit dose is generally greater than the amount required when using conventional injectable injectables. formulation. A preferred slow release formulation is, for example, a continuous release formulation, for example a formulation of the type described in US 4767628 and US 5004602. A preferred slow release parenteral formulation comprises 10 to 100 mg of polypeptide per unit dose. Another preferred slow release formulation is a microcapsulated polypeptide using a biodegradable biocompatible copolymer.

These preparations are preferably administered intravenously, although administration may also be by subcutaneous, intramuscular or intradermal injection.

Polypeptides are therapeutic compositions requiring specialized pharmaceutical formulations for effective, safe and appropriate use in patients. Since even small oligopeptides such as thyrotropin releasing (TRH) have very low oral activity and larger molecules are inactivated by endopeptidases in the gastrointestinal tract, long-term treatment requires daily injection or administration from small portable infusion pumps.

On the other hand, there are enough potential procedures for self-administration routes (e.g. nasal administration, lace), or for controlled release formulations such as injectable microcapsule suspensions or implants.

Drugs suitable for transdermal administration may be in the form of an optional buffered aqueous solution of a compound of Formula III

and can be delivered by passive diffusion or electrically stimulated transport, for example by iontophoresis (see, for example, Pharmaceutical Research J (6), 318 (1966)).

The composition of the invention will normally be administered so that the daily oral dose will be from 0.1 mg / kg to 50 mg / kg and most preferably from 0.1 mg / kg to 25 mg / kg and the daily parenteral dose will be from 20yu / kg to 10 mg. mg / kg, more preferably from 100 yy / kg to 10 mg / kg

According to another feature of the present invention, there is provided a method of producing an antagonist effect against bombesin in a warm-blooded animal such as a human being in a treatment comprising administering to the animal or a pharmaceutically acceptable salt thereof an effective amount of a polypeptide of Formula I. The invention also provides the use of the polypeptide of formula I or a pharmaceutically acceptable salt thereof in the manufacture of a novel medicament for use in the treatment of a disease or medical condition mediated by a bombesin or a bombesinoid peptide.

The compounds of the invention are useful for inhibiting the binding of gastrin-releasing peptide in cells. The compounds of the invention also inhibit the growth of cancer cells,

The polypeptide of the invention is useful in the treatment of malignant diseases, for example malignant lung diseases such as human small cell lung cancer, for example malignant diseases

- 19 pituitary gland, adrenal gland, pancreas or skin.

In particular, the polypeptide of the invention is useful for symptomatic granulation and / or treatment of exocrine pancreatic adenocarcinoma. The polypeptide of the invention is useful in the art. treating conditions associated with overproduction of bombesin or bornbesinoid peptides (such as gastrin releasing factor), for example, overproduction of gastrin in the intestines.

Gastrin production in animals is associated with inhibition of growth hormone and prolactin release. Thus, the polypeptides of the invention can be used to assist the availability of growth hormone in a human or animal in need of such treatment. The polypeptides of the invention may also be used in the treatment of conditions associated with the failure of normal physiological control of gastric acid secretion control.

According to further aspects of the invention, there are provided:

(a) polypeptides of the invention for use in therapy;

(b) pharmaceutical formulations comprising the polypeptides of the invention, at least one pharmaceutical carrier or base, and optionally one or more other therapeutic ingredients;

(c) use of the polypeptides of the invention in the manufacture of a medicament for the treatment of cancer;

(d) use of a polypeptide of the invention in the manufacture of a medicament for the treatment of:

(i) entercutaneous fistula (ii) type II diabetes, type I diabetes (iii) zollinger-ellison syndrome (iv) pancreatic islet carcinoma (v) acromegaly (vi) psoriasis, such as chronic scab psoriasis $ vii) postoperative small-bowel fistula (viii) dumping syndrome (ix) malignant ihsuloma (x) pituitary adenoma growth hormone cells (xi) pituitary adenoma thyrotropin secretion and (xii) small cell cancer mold.

(e) a method of inhibiting cell growth that is sensitive to the growth promoter activity of a gastrin-releasing peptide in a mammal (such as a human) in need of such treatment comprising administering to said mammal a growth inhibiting amount of a polypeptide of the invention.

DETAILED DESCRIPTION OF THE INVENTION

The following examples serve to illustrate the preparation of the polypeptides of the invention and their biological properties

Materials:

The following abbreviations, which are used hereinafter:

BOC = tertiary butyloxycarbonyl

TLC = thin layer chromatography

NMR = nuclear magnetic resonance

MS = mass spectrometry

The following compounds were obtained from Advanced Chemtech, Tne. PlO.Box 1403, Louisville, KY 40201

p-methylbenzhydrylamine resin.HCl (substitution range from 0.56 to 0.94 meq / g), trifluoroacetic acid N-Boc-D-alanine, N-Boc-L-alanine, N-Boc-N-tosylarginine, N- Boc-glycine, N-Boc-L-leucine, N-Boc-L-leucinol, N-Boc-L-norleucine, N-Boc-D-phenylalanine, N-Boc-L-phenylalanine, N-Boc-D- proline, N-Boc-L-proline, N-Boc-D-prolinol, N-3oc-sarcosine, N-Boc-L-tryptophan and N-Boc-L-valine.

N-Boc-L-im-CBZ-L-histidine, N-Boc-beta-thienyl-L-alanine and 3- (2-naphthyl) -L-alanine were purchased from Bachem, Inc., 3132 Kashiwa Street , Torrance, CA 90505 USA.

L-methoxinin (O-methyl-L-homoserine) was obtained from Biohellas S.A. 10. Parnithos Street, 154 52 P. Psychike, Athens, Greece.

Leucine methyl ester hydrochloride, (+) - 6-methoxy- [beta], -methyl-2-naphthaleneacetic acid and (S) -6-methoxy-4-methyl-2-naphthaleneacetic acid were purchased from Sigma Chemical Company, P.O. Box 14508, St. St. Louis, MO 63178, USA.

1'-tert-Butyldicarbonate, 4-fluorophenylalanine, 3- (4-hydroxyphenyl) propionic acid, N-hydroxysuccinimide ester and both (R) - and (S) -3-phenylbutyric acids were obtained from Fluka Chemical Corp., 980 South Street, Ronkonkoma,

NY 11779, USA.

The following were purchased from Aldrich Chemical Co., Inc., 1001 West Saint Paul Avenue, Milwaukee, WI 53233, USA:

j '

3-phenylpropionic acid (hydrocinnamic acid), (R) and (S) -3,3,3-trifluoro-2-methoxy, γ-2-phenylpropionic acid, dicyclohexylamine, 1-cyclopentanecarboxylic acid,

4-methylmorpholine, 1-methylimidazole, sodium cyanoborohydride and diisopropylethylamine.

Dichloromethane (CH 2 Cl 2), Ν, Ν-dimethylformamide (DMF), and N-methylpyrrolidone were used as solvents for solid phase peptide synthesis and were obtained from Baxter Healthcare Corp. Burdick and Jackson Division, Muskegon,

MI 49442, USA.

The Bop binder, benzotriazol-1-yl-oxy-tris (dimethylamino) phosphonium hexafluorophosphate, was obtained from Richelieu Biotechnologies, Inc., 5726 Laurier Blvd., St-Hyacinthe, QC., J2S, 3V8 Canada.

Hydrofluoric acid was obtained. lähde: Matheson Gas Products, P.O. Box 85, 932 Paterson Plank Road, East Rutherford, NJ 07073

Preparation of Peptides of the Invention

Step I, preparation of Boc-amino acids:

Amino acids which were obtained without the tert-butyloxycarbonyl protecting group on the amino group (L-methoxynin, 4-fluorophenylalanine) were protected as follows.

One equivalent of the amino acid was dissolved in 5% aqueous Na ₂ CO 4 / 1,4-dioxane and cooled to 0 ° C. Two equivalents of di-tert-butyl dicarbonate in 1,4-dioxane were added dropwise to the cooled amino acid mixture. After the addition, a ^mixture allowed to warm to 25 and stirred for 12 hours. After completion of the reaction as determined by TLC, dioxane was evaporated. The aqueous mixture was acidified with 1N HCl to pH = 2, after which crystals formed, in most cases with a 60% yield.

- 23 The solid was filtered and characterized by TLC,

NMR and MS, as generally pure, could be continued without further purification.

In those extreme cases, such as L-methoxinine, in which the product that precipitated from the acidic aqueous solution was liquid, the product was removed from the aqueous solution with ethyl acetate. The organics were then dried over anhydrous sodium sulfate and the ethyl acetate was evaporated.

The liquid residue was dissolved in a small portion of ethyl acetate. One equivalent of dicyclohexylamine was added to the mixture, which was then diluted with a large excess of diethyl ether. After cooling, the Boc-amino acid-dicyclohexylamine salt crystallized. The solid was filtered off (20-30% yield), characterized (TLC, NMR, MS) and used without further purification.

Stage II, Boc-D-prolinal preparation:

The following procedure is a variation of the reaction performed by Mancus et al. in J. Org. Chem. 43, 2481 (1978).

In a 500 ml three-necked flask was dissolved 25 ml of a 2.0 M solution of oxalyl chloride in methylene chloride (50 mmol of a solution obtained from Aldrich) in 60 ml of dry CH 2 Cl 2 at -60 ° C under N 2. Dimethylsulfoxide (10 mL, 140 mmol, Aldrich) was dissolved in 25 mL dry CH ₂ Cl ₂ and added dropwise to a -60 ° C cold solution via a funnel. This mixture was then stirred for 15 minutes after the addition was complete. Boc-D-prolinol (10 g, 50 mmol) of Advanced Chemtech) was dissolved in 60 mL dry CH ₂ Cl ₂ e cooled to -6 ° C in a 500 mL round bottom flask below to facilitate the pooling of both cooled flasks. was N _; supply through the establishment of the seribvé · to serve both banks.

The Oxalyl chloride / DMCO mixture was bluntly added to the alcoholic solution of N ₂ via a two-point needle transfer. By manually controlling the N ₂ flow, the content was transferred from one flask to the other very slowly, but not dropwise. After the addition was complete, the mixture was stirred for 20 minutes. Finally, a slow 20 mL of triethylamine (140 mmol, Kodak) was added while maintaining the temperature at -60 ° C. The mixture was then allowed to warm to room temperature.

The mixture was washed with 50 ml of water. This aqueous layer was then extracted with 3 x 70 mL of CH ₂ Cl ₂ . The organics were washed successively with 50 ml of each of the following liquids, 5% N / NaHCO 3, 1N HB1 and water. The aqueous washings were extracted with 50 mL of CH ₂ Cl ₂ , which was then combined with other organic substances. After drying the organic drugs with anhydrous sodium sulfate, the solvent was removed by rotary evaporation. The residue was dried using a vacuum pump for 3 hours.

The NMR characteristic of the -OH peak for Boc-D-prolinol (delta 4.5 ppm) was no longer visible. A sharp peak at 9.4 ppm reported the presence of the aldehyde proton of Boc-D-prolinal TLC indicated a production mixture that contained no substantial by-products so that it could be used without further purification.

Step III, Preparation of Boc-D-pro Ψ (CH ₂ NH) X-MBHA resin:

i) p-Methylbenzhydrylamine resin.HCl (p-MBHA) from Advanced Chemtech was used. The resin substitution ranged from 0.90 to 0.97 meq / g.

A ‘g of β-MBHA resin (9.0-9.7 mmol) was loaded into a manual peptide shaker (Milligen).

The resin was washed twice (3 and 5 minutes) with a 10% solution of Ν, Ν-diisopropylethylamine (DIEA, Aldrich) in CH 2 Cl 2) with shaking. The resin was then washed twice with CH ₂ - The neutralization of HCl on the resin was indicated by the Kaiser ninhydrin test and was found to be a very dark blue color.

The resin was then treated with two equivalents as BOC-norleucine (Bachem, Torrance, CA), benzotriazol-1-yloxy-tris (dimethylamino) phosphonium hexafluorophosphate (BOP), Richelieu Biotechnologies of Canada) and 4-methylmorpholine, (Aldrich); all dissolved in 50 ml of N, N-dimethylformamide (DMF). The mixture was shaken for 1 hour and then washed twice with CH ₂ Cl ₂ . The binding was verified by the Kaiser test (colorless) compared to the previous Kaiser result (blue)

Deprotection Boe group on norleucine was undertaken by shaking the resin in the presence of 50% strength trifluoroacetic acid (TFA), and Advanced Chemtech, the CH ₂ C1 ₂ two times for 5 and 20 minutes. After washing the resin once CH Clg, excess TFA was neutralized with l0% sodium DIEA / CH., _O C 1 for 3 and 5 minutes with stirring. Two CH ₂ C1 ₂ washes, the resin was again qualitatively tested with ninhydrin (very deep blue).

The resin was then shaken for 2 hours with two equivalents of Boc-D-prolinal in 50 ml of 2% glacial acetic acid in DMF. 3 equivalents of sodium cyanoborohydride (Aldrich) were added slowly and gradually over two hours.

After washing twice with CH ₂ C1 ₂ was tested for the binding with ninhydrin (colorless). The resin was now ready for standard solid phase peptide synthesis.

- 26 ii) Alternative preparation of Boc-D-Pro 'Ψ (CH ₂ NH) X-MBHA resin

The following procedures can be used to prepare for any L-S / _(CH2 NH) X-MBHA resin, wherein X is any amino acid "with a primary amino group.

A. Boc-D-Pro Ψ (CH ₂ NH) X-OH can be prepared according to the methods of Martinez et al. in J. Med. Chem. 28, 1874 (1985) or D. Tourwe et al. in Peptides 1988:

Proceedings of the 2nd European Peptide Symposium. , Ed.

Jung, Bayer; Walter de Gruyter, ptr.

B. This resin can be prepared by binding Boc-D-Pro (CH ₂ NH) X-OH to MBHA resin by shaking with BOP and 4-methylmorpholine (or 1-methyliraidazole) in N-methylpyrrolidone for 2 hours.

Preparation of intermediates

Preparation of ethyl 3- (2 1- (tert-butoxycarbonyl) -2-pyrrolidinyl) -3-hydroxypropionate

Fresh activated zinc powder (C, 79 g, 12 ramol, Aldrich) and benzene (50 mL) were placed in a 250 ml two-necked round bottom flask under nitrogen. This flask was connected to a Bean-Stark apparatus and 25 ml of benzene was distilled off into a receiver. A solution of BOC-D-prolinal (1.90 g, 9.5 ml) and ethyl bromoacetate (2.0 g, mmol, Aldrich) was added dropwise under reilux. An iodine crystal was added to initiate the reaction after dropping half of the solution. After complete addition, it was refluxed for 3 hours, cooled and thoroughly washed with 0.5 N HCl. The aqueous solution was extracted with ether (25 mL) and then the combined organic solution was washed successively with water (30 mL) and saturated NaHCO 3 (30 mL). ³ mL) then dried over anhydrous sodium sulfate and concentrated in vacuo.

The yellow oil was chromatographed on silica (hexane / ethyl acetate) to isolate a colorless oil (1.12 g, 41%).

Mass spectral analysis gave results at 288 (m + 1, 20%), 232 (m-tBu + 1.40%) and 188 (m-CO ₂ Bu + 1, 100%). ¹ H-NMR (CDCl ₃ ): δ 4.15 (br m, 3H); 3.92 (m, 1H);

3.48 (m, 1H); 3.26 (m, 1H); 2.40 (m, 2 H); 1.65-2.00 (br m, 4H); 1.45 (s, 9H); 1.25 (t, 3 H).

Preparation of 3-O- (tert-butoxycarbonyl) -2-pyrrolidiethyl) -3-hydroxypropionic acid

A mixture of ethyl 3- (1- (tert-butoxycarbonyl) -2-pyrrolidinyl) -3-hydroxypropionate (3.06 g, 10.6 mmol) with NaOH (0.80 g, mmol) in H ₂ O: THF: methanol (3: 3: 1), 35 mL, was stirred at 25 ° C for 2 hours. The mixture was slightly concentrated (30 ° C) under vacuum. The aqueous solution was acidified with 1.0 N HCl at pH = 5 and was extracted with ethyl acetate. The organic washings were dried over MgSO 4, filtered and concentrated in vacuo. This residue was recrystallized from a mixture of hexane and ethyl acetate to give 1.19 g (47%) of a colorless solid (A, mp = 122-4 ° C). The mother liquor was concentrated to a viscous pale yellow oil (B).

Mass spectral analysis showed:

260 (m + 1, 60%); 204 (m-tBu + 1, 100%), 160 (m-100 + 1, 70%).

Elemental analysis: theory, 55.58% C, 8.16% H,

5.40% N; 55.39% C, 8.23% H, 5.37% N, HPLC:

one major peak for the NEW PAK with 50% methanol / H ₂ 0 / 0.1% TFA / 0.1% triethylamine (A = 93: 7 ratio, B = 1: 2 ratio found on extended HPLC) .

The two products were found to be different stereoisomers in the hydroxyl position. Both stereoisomers have been used in individual peptides; the more active of the two is N - ((3-phenyl) propionyl) -HisTrp-AlaValD-AlaHis (3- (2-pyrrolidinyl-3-hydroxy) propionyl) -Phe-NH2;

Step IV, peptide and purification synthesis

The peptides were synthesized using the improved version of the solid-phase method described by R. B. Merifield, Solid Phase Peptide Synthesis I. The Synthesis of a Tetrapeptide, J. Am. Chem. Soc., 83, 2149 (1963) using the Applied Biosystems Model 430A peptide synthesizer.

For the synthesis of the "Pseudopeptides", the appropriate resin, for example, a Boc-D-Pro ^ (CH ₂ NH) Phe-MBH was loaded into a synthesizer and a standard deprotection (TFA / CH ₂ Cl ₂ ) neutralization (diisopropylethylamine / CH ₂ Cl ₂ ) program as supplied Applied Biosystems (850 Lincoln Center Drive, Foster,

CA 94404 USA).

Boc-protected amino acids were attached to the resin using a modified program suitable for the BOP binding procedure as described: Dung Le-Nguyen, Annie Heik, and Bertrand Castro, J. Chem. Soc., Perkins Trans, 1, 1915 ('987). The binding protocol included dissolution of Immol Boc-protected amino acids, 1 mmol BOP and 1 mL 1M 1-methylimidazole in 7 mL DMF. The mixture was added to 0.5 mmol of resin, stirred for 1 hour and filtered, followed by washing with DMF untracked set and CH ₂ C1 _2nd

. · **

After the peptide was deposited on the resin, the peptide was deblocked and cleaved from the resin with liquid HF containing 10% anisole in a variation of the method described by S. Sakakibar et al.

- 29 at Bull. Chem. Bathroom. Jap., 40, 2164 (1967). The peptide and resin were then washed with ethyl acetate, and then the peptide was extracted from the resin with an aqueous 1% acetic acid solution. The peptide solution was then lyophilized to give a dry solid peptide.

The peptides were then purified by reverse-phase liquid chromatography using a Vydac 218TP1022 column on a Waters Delta Prep 3000 system equipped with an Oilson Model 116 ultraviolet detector. Purification was achieved by equilibrating the column with 0.1% TFA in water and developing with a linear gradient of acetonitrile from 1 to 40% in 20 minutes at a flow rate of 20 ml / min. Samples were manually pooled and checked for purity on a Spectra-Physics analytical HPLC system (including SP8700, SP8440, SP8780, and SP4200) using a Vydac 218TP54 column. Flow was used

1.5 ml per minute using a 0.1% TFA / acetonitrile gradient from 1 to 60% ACN in 10 minutes.

Synthesis of N - ((3-phenyl) propionyl) HisTrpAlaValD-AlaHisD-Pro Ψ (CH ₂ NH) Phe-NH ₂

Methylbenzhydrylamine resin (MBHA) (5.0 g, 4.7 mmol Advanced Chemtech) was washed twice (3 and 5 minutes) with 10% diisopropylethylamine (LIEA, Aldrich) in dichloromethane (DCM) on a Milligen peptide shaker. The resin was then washed with DCM and Ν, Ν-dimethylformamide (DMF). A solution containing 2.5 g Boc-Phe (9.4 mmol, Chemtech), 4.3 g BOP Reagent (9.4 mmol Richelieu Biotechnologies) and 0.95 mL 4-methylmorpholine (8.6 mmol, Aldrich) was shaken. the resin after a period _of one hour. the resin-then washed sequentially with methanol and DCM and coupling was checked using the Kaiser ninhydrin test qualitative.

A portion of Boe-Phe-MBHA (3.06 g, 2.8 mmol) was deprotected by treatment with 50% trifluoroacetic acid (TFA), Cheratech) in DCM for 5 and 20 minutes. The resin was washed twice with DCM. The acid was then neutralized by shaking for 3 and 5 minutes with 10% DIEA solution and washed with DCM and DMF. E, N-Phe-MBHA was shaken in a 1% acetic acid / N-methylpyrrolidone solution containing 1.2 g of Boc-Dprolinal (6.0 mmol, synthesis described earlier) for 2 hours. Sodium cyanoborohydride (0.54 g, 8.62 mmol, Alchrich) was added in four portions over 2 hours (approximately every 20 minutes). The resin was washed with methanol and DCM and tested with ninhydrin. Alternatively, the same resin may be prepared as described in the alternative preparation described above.

The following amino acids (Bqc-His (Z), Boc-P-Ala, Boc-Val, Boc-Ala, Boc-Trp, Boc-His (Z) and 3-phenylpropionic acid) were added respectively by an automated peptide synthesizer of useful biosystems (model 430A) using the same procedure described for binding, deprotection and neutralization involving Boc-Phe. (using 0.5 mmol of resin and 1 'pitch of each amino acid, BOP and 4-methylmorpholine). All amino acid linkages generated by the 430A model were checked by a qualitative Kaiser assay on resin samples formed by this load upon completion of each binding.

The peptide (approximately 1.0 g) was cleaved from the MBHA resin by dropping the resin bound peptide with hydrogen fluoride (approximately 10 mL at 0 ° C for 1 hour). The peptide was precipitated and filtered with MBHA resin with ethyl acetate. The peptide was then extracted from the resin. with 1% aqueous acetic acid and isolated by freeze-drying the extract.

A portion of the peptide (100 mg) was purified on a Vydac C-18 preparative column (Chemtech) using a 0.1% TFA / (0.1%) TFA / acetonitrile gradient. The pooled fractions were run on an analytical Vydac C-18 column (Chemtech) and these samples showing pure peptide were pooled and frozen. Approximately 15 mg of peptide was isolated. After characterization of PAB - mass spectrum (MH = 1081, 1 ') and amino acid analysis of Ala (2.19), His $ 1.68), Val (1.12), 12.2 mg remained (12% yield based on HPLC) .

Analytical methods

Purity was checked by analytical HPLC using a Spectra-Physics analytical HPLC system including SP8700, SP8440, SP8780 and SP4200. A Vydac 218TP54 column with a flow rate of 1.5 ml / min of a 10.1% TFA / acetonitrile gradient was used.

The correct mixture of each peptide was determined by amino acid analysis and (FAB) mass spectroscopy. Amino acid analysis was performed as follows. Approximately 1 to 10 nanomolar peptide was placed in an acid-washed pyrex test tube. The peptide was hydrolyzed under reduced nitrogen atmosphere of 6N HCl with 1% phenol for 1 hour at 150 ° C. The peptide was then dried with a 2: 2: 1 ethanol: water: triethylamine mixture and derivatized 7: 1: 1: 1 with ethanol: water: three ethylamine: phenylisothiocyanate. Derivatized amino acids were then analyzed by reverse phase chromatography.

Fast atom mass spectra (FAB) mass spectra were obtained on a VG 70SQ EBQQ geometry mass spectrometer using the VG 11-25OJ data acquisition system.

The mass spectrometer was operated at a 7 kV acceleration potential and a resolution of 1000 (10% saddle resolution). The FAB nozzle used in these experiments was an Ion Tech FAB 11N operating at 7 kilovolt potential and 1 milliamp current. The xenon was used as bombardment gas - ^c at a pressure of 1x10 &lt; -1 &gt; millibars of the source pressure. The test sample was dissolved in glycerol prior to FAB-MS analysis.

Biological results

Peptides were evaluated for their activity to inhibit GRP binding to Swiss 3T3 cells. Antagonist efficacy was measured by inhibiting mitogenic stimulation of immobile 3T3 ROZ cells. 3T3 ROZ cells were obtained from Enrique Rozengurt, Imperial Cancer Research Fund, P.O. Box 123, Lincoln`s Inn Field, London WC2A 3 PX, England. Cells were incubated for 18 hours with 10 ng / ml BN either alone or in the presence of an antheagonist. The cells were then pulsed labeled

3 for 2 hours H-thimidine and washed. The incorporated H was then calculated. Exposure to bombesin resulted in increased cpm above moderate control up to maximal response. Antagonist potency was measured by inhibiting this maximal response to baseline. IC 50 values were determined from titration curves. The analogues and IC 50 values achieved are summarized in Table I.

structure

- - 33 .. Table I ic _5Q (m)

MH &lt; ^{+ &gt;} (PAB) -MS

AAA

N - ((R) -2- (6-methoxy-2- 3x10 naphthyl) propionyl) -HisTrpAlaVa-D-AlaHiD-ProΨNe-KEL ·

1128

N - ((S) -2- (6-methoxy-2- 1.77.10 naphthyl) propionyl) -HisTrpAlaVal-D-AlaHisD-Pro-Nle-NH ₂

N - ((S) -3-phenylbutyryl) - 4.71.10

HisTrpAlaVal-D-AlaHis-B-Pro-Nle-NH _C

1128

1,061.6

N - ((R) -3-phenylbutyryl) -His- 7.54.10 ^-8 1061.6

TrpAlaVal-D-Ala-His-D £ please? _Nle-NH2

N - ((3-phenyl) propionyl-His-2,35.10 TrpAlaVal-D-Ala (3- (2-thienyl) -Ala) -L-Pro-Nle-NEL,

N - ((S) -3,3,3-trifluoro-2-methane 1132 thoxy-2-phenylpropionyl) -His-tested TrpAlaVal-B-AlaHis-D-Pro S? Nle-N & J

1,063.5

N - ((R) -3,3,3'-Trifluoro-2-methoxy-2-phenylpropionyl) -His · TrpAlaVal-D-AlaHis-D-Pro-Nle-NHg 11

N - (((4 ^x hydroxy) -3-phenyl) pro 1,99.10 pionyl) -Pro-D-ArgGly-D-PheHisTrpAlaValGlyHis-D-Pro 'P Nle-KL ·

Ala (1.85), His (1.79), Val (1.00)

Alad, 97), His / 1.81), Val (1.00)

Alad, 92), Hisd, 71), Val / 1.00)

Alad, 77), His (1.65), Val d, 00)

ON THE

Ala (2.40), Hisd (15), Val (1.00)

Alad, 90), Hisd, 19), Val d, 00)

Alad, 00),

Arg (1.05), Gly (2.15),

- 34 continued Table I Structure

ICl- _n (IV) 1 '1H ⁺ (PAB) -MS AAA 5U

His (1.78) _: Phed, 03), Pro (1.17), Val (0.91)

K - ((3-phenyl) propionyl) -HisTrp AlaVal-D-AlaHis-D-Pro Methoxininn ₂

K - ((3-phenyl) propionyl) -HisTrpAlaVal-D-AlaHis-D-Pro-Phe-NHg

B - ((3-phenyl) propionyl) -HisTrp-Ala-Val-D-AlaHis-D-Pro * 9? Leu-NHG

N - ((3-phenyl) propionyl) -HisTrpProVal-D-ProHis-D-Pro SE 'leu-NHg

N - (((3'-Trifluoroethyl) -3-phenyl) propionyl) -HisTrpAlaVal-D-AlaHis-D-Pro Leu-NH ₂

N - ((3-phenyl) propionyl) - (3- (2-thienyl) -Ala-TrpAlaVal-D-AlaHis-D-Pro'T Leu-RH ₂

J '

H - ((1-cyclopentyl) carboxyl) -DArgD-AlaD-PheHisTrpAlaValGlyHis-D-Pro'FNle-NHg

3.43-10 ⁸ 10-49.4 Ala (2.09),

His (1,84),

, 07)

8:30:10 ^{- 18} A.7a 1081.7 (2.19),

H 8 8 to 1, ó o), Vald, 12)

2.72.10 ^{- 8} 922.0 Ala (2.02),

Kis (0, 75) _r Vald, 00)

6.92.10 ^- 1099.9 Hisdal ^7, 73).

Pro (2.06). Vald, 00) '

2.70.10 ^- 1116.0 Alad ^9, 92).

His (1,57), Vald (00)

4.70.10 ^{- 9} 1063.0 Ala (1.96),

His (0.75) • Vald, 00)

1.75.10 ^{- 8} 1371.6 Ala (1.93),

Arg (1.04), Glyd, 03), His (1.91), Phed, 04), Vald, 04) i

- 35 continued Table I

Structure ΊΟ ^ θίΜ)

MH &lt; ^{+ &gt;} (FAB) -MS AAA

N - ((3-phenyl) propionyl) -His-. TrpAlaValGlyHis-D-Pro-Nlenh ₂

N- ((1-cyclopentyl) carboxyl) D-Arg-D-Ala-D-PheHisTrpAlaval-D-Pro NIe-NHg

N - ((3-Phenyl) propionyl) -HisTrp AlaVla-D-AlaHis-D-Pro SIZNle-NH ₂

TyrPro-D-ArgGlylj-PheHisTrpAlaValGlyHisD-Pro ‘S'Nle-NHg

D-ArgGlyD-PheHisTrpAlaValGlyHiaD-Pro Ψ Nle-NHg

N - ((3-phenyl) propionyl) -HisTrpAlaValD-AlaHis (4-fluoroPhe) -NH ₂

2,00.10 ^-7 1034.1 Ala (O, 99),

Glyd, 11l, Hisd, 94), Val (0.96),

1.73.10 ^{- 8} 1386.7 Ala (2.90)

Arg (1.07), Hisd, 93), Phe (1.05), Val (1.05)

3.10.10 ^{- 9} 1048.2 Ala (2.02).,

Hisd, 90), Val d., 08)

8.54.10 ^{- 8} 1522.2 Alad 03)

Arg (0.97), Gly (2.07), Hi s (2,030-, Phed, 00), Pro (0.93), Tyr (0.87), Val d, 11)

2.00.10 ^-7 1262.5 Ala (0.97),

Argd, 05), Gly (1.96), Hisd, 94), Phed, 04), Val (1.04)

7.87.10 ^-8 1017.0 Alad 87)

His (0.86), Val d, 00)

- 36 continued Table I

structure

IC ₅₀ (M) ⁺ MH (PAB) -MS AAA

N - ((3-phenyl) propionyl) -HisTrp AlaValD-AlaHisD-ProPhe-NH ₂ 3,65.10 ⁸ 1096 Ala (2.13), His (2.31), Phed, 11), Pro (1.24), Vald, 00) N - ((3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro SI? (3- (2-naphthyl) -D-Ala) -NH ₂ 2.65.10 ^-8 1 1,132.4 Alad, 96), Hisd, 42), . Val (1.00) M Oígnjfi) ppoplonylX§ TrpAlaValD-PheHisD-Pro 'Ψ' Phenh ₂ d, 3S, 10 “ ^p * 115714 Λ1Μ1115) | Hisd, 76), Phe (0.99), Vald, 00) PheHisTrpAlaValD D-Phe-Ψ _{AlaHisDPro-NH2} x 5.47.1 O ^-10 ' 1,093.2 Alad, 98), Phe (O.92), Vald, 00) -i N- ((3-phenyl) propionyl) -DProArgGlyD-PheHisTrpAlaValD-AlaHisD-Pro 'Phe-NH ₂ -1 Q 3.90.10 ^IU 1540 Alad, 83), Arg (1.02), Gly (0.94), His (1.54), Phe (0.85), Pro (0.94), Val (1.00) N - ((3-phenyl) propionyl) -3- (2-thienyl) -Alai) -TrpAlaValD-AlaHisD-Pro SE / Phe-NI ^ 3,65.10 ^-8 Alad, 92), His (0.59), Vald, 00)

continuation of Table I structure IC ₅₀ (M) ^{+ -} (fab) -ms AAA N- ((3-Phenyl) propionyl) -HisTrpAlaVal-sarcoein) -HisDPro'S 'Phe-NH _{2 K} 5.55.10 ^-10 Ala (1.02), His (2.06), Val (1.00) N - (((4'-hydroxy) -3- $ enyl) propionyl) -HisTrpAlaValDAlaHisD-Pro Phe-Nl · ^ -1 0 K5,47.1A ^lu 1098 Ala (2.04), Hisd (96), Val d, 00) N - (((2 ', 6'-dichloro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro S &' Nle-NHg ' 9,07.10 ^-7 1103 Alad, 85), Hisd, 83), Val d, 00) N (((3 '»4'-dichloro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro SE ⁷ Nle-NHg <5.44.W ¹⁹ 1103.5 Ala (2.06), Hisd (79), Val d, 00) N - (((4'-hydroxy) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro Ψ Nle-NIL, 1,91.10 ^-9 1,050.1 W / A N- (1-naphthoyl) -HisTrpAlaValDAlaHisD-Pro-Nle-NH ₂ 9,35.10 ^-10 1,069.9 ON THE N- ((3,7-dihydroxy-2-naphthyl)) HisTrpAlaValD-AlaHisD-Pro StZ Nle-NH ₂ 2,36.10 ^-8 1101, 6 Alad, 34), His (2.05), Vald, 00) N - (((2,3-dihydoxy) -2-phenylacetyl) -HisTrpAlaValD-AlaHisD-Pro-Nle-NIj) 1,22.1ο ⁷ 1,065.5 Alad 23) Hisd 90) Vald 00) N- (2- (3-pyridyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro'S 'Nlenh ₂ 3,87.10 ^-7 1,034.6 Ala (2.26), His (2.00 (, Val (1.00))

- 38 continued Table I

structure ic ₅₀ (m) I WH ⁺ (PAB) -MS AAA N- (2- (2-thienyl) acetyl 2,98.10 ^-8 1040 Ala (2.15), -HisTrpAlaValD-AlaHisD- His (1.63), Pro-Nle-NHg Val (1/1) N - (((3-Fluoro) -3-phenyl) - 3,00.10 ^-9 1 066 Ala (2, L 4), propionyl) -HisTrpAlaValD-. HisD, 66). -AlaHisD-Pro'a'Nle-NHg Val (1.00) N - (((3,4-dihydroxy) -2-phe- 4,63.10 ^-8 1079 Ala (2, L 3), nylr) acetyl) -HisTrpAlaValD- Hisd, 43), AlaHisD-Pro Ϋ _Nle-NH2 Val d) The - (((R) - (-) - 2-phenyl) pro- -1 0 9.25.10 ^lu 1081, 2 Alak á 071 propionyl) -HisTrpAlaValD-AlaHisD- Hisd 51) Pro Ϋ PheNH ₂ Val d) N - (((S) - (+) - 2-methoxyphenyl) propionic 2.77.10 9 1,081.1 Al G. <2.0 µ), phenyl) -HisTrpAlaValD-AlaHisD- Hisd, 44) For S? Phe-H ₂ Val (1.00) H - (((trans) -2-phenyl) -cycloalkyl 1,46.10 • 9 1,093.1 Ala (1,999), propanoyl) -HisTrpAlaValD- Hisd, 44) AlaHisD-Pro Si? Phe-NH ₂ Val (1.00) N- (3- (10-phenothiazinyl) Pro 2,66.10 8 1,202.1 Ala (2,5) propionyl--HisTrpAlaValD-AlaHis- His (1.58) D-For -S? Phe-NH ₂ , 00) N- ((3-methyl-3-phenyl) butyl) 1,44.10 ' 8 1109.5 Ala (2.07), ryl) -HisTrpAlaValt> -AlaHis- His (1.63), D-Phe-Pro T _NH2 , 00)

- 39 continued Table I

structure , ic ₅₀ (m) MH ⁺ / PAB) MS AAA N - (((2'-Trifluoromethyl) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro-Phe-nh ₂ 3,50.10 ^-7 1,135.4 Ala (2.10), His (1.63), Vald, 00) N- (((3'-trifluoromethyl) 2-phenyljacetyl) -HisTrpAlaValD-AlaHisD-Pro '3? Phe-NHg 8.81, 10 ^-8 1,135.4 Ala (2.10), His (1.60), Vald, 00) N - (((4'-trifluoromethyl) -2-phenyl) acetyl) HisTrpAlaValD-AlaHisD-Pro-Phe-NH ₂ 1, 41.10 ^-7 1,135.5 Ala (2.24), Hisd, 68), Vald, 00) N - (((2 ', 3'-difluoro) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro Ψ Phe-NHg 3.61 .ΙΟ ' ⁷ 1,103.4 Ala (2,16) 'Hisd, 63), Vald, 00) N - (((2 ', 4'-difluoro) -2-phenyl) 1,8,80.10 ^-7 acetyl) -HisTrpAlaValD-AlaHisD-Pro T Phe-NHg 1,103.2 Ala (2.16), Hisd, 62), Vald, 00) N - (((2 ', 6'-difluoro) -2-phenyl) acetyl) HisTrpAlaValL-AlaHisD-Pro Ψ Phe-NHg 5,71.ΙΟ ' ⁷ 1,103.2 Ala (2.11), Hisd, 69), Vald, 00) N - (((2-Amino) 2-phenyl-acetyl) HisTrpAlaValD-AlaHisD-Pro T Phe-NHg 7,30.10 ^-9 1,082.2 ON THE N- d -naphthyl) -H'isTrpAlaValD-AlaHisD-Pro P Phe-NI ^ 1,81.10 ¹² 1,103.9 Ala (2.06), His (2.12), Vald, 00)

- 40 continued Table I

Structure - IC _Rf (M) MH ⁺ (PAB) - AAA ⁵⁰ MS

N - (((3 ', 4', 5'-trimethoxy) ~ 3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro SF Phe1¾

N - ((6-methoxy) -2- (2-naphthoyl) propionyl) -HisTrpAla.alD-AlaHisD-Pro Phe-NHg

N - (((3-Trifluoromethyl (3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-ProSF Phe-NI)

N - (((S) -3-phenyl) butyryl) HisTrpAlaValD-AlaHisD-ProV Phe-NELp

N - (((4'-Methoxy) -3-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro-PheN - ((((S) -2-hydroxy) -2-phenyl) acetyl) -HisTrpAlaValD-AlaHisD-Pro & Phewh ₂

N- ((3-phenyl) propionyl)) HisTrpAlaValD-AlaHisProS? Phe-NH ₂

1,11.10 ^-11 1171,8 Ala (1,86)

His (1.71), Val (1.00)

3,44,10 ^-11 1161, 8 Ala (1.87),

His (1.87), Val (1.00)

3,47.10 ^-11 1149.8 Ala (2.17),

Hisd, 50), Val (1.00)

5:47:10 ^{- 11} 1095.9 Ala (1.83),

His (1,82), Vald (00)

5:40:10 ^{- 12} 1111.9 Ala (1.84),

His (1.76),

, 00)

5,53.10 ^-9 1083.8 Alad 92)

Hisd., 64), Val d, 00)

9,25.10 ^-9 1081.5 Ala (1.80),

Hisd, 75), Vald (00)

- 41 continued Table I

structure

IC _n (M) MH &lt; ^{+ &gt;} (PAB) - AAA

MS

N - ((2-methyl-2-phenyl) propionyl) -HisTrpAlaValD-AlaHisD-Pro Phe-NH ₂

N- (3-d-naphthyl) propionyl) HisTrpAlaValD-AlaHisDProPe Phe-NI ^

N- (((R) -3-phenyl) butyryl) HisTrpAlaValD-AlaHis-ProSP Phe-NI ^

ΙΊ - ((9-fluorenoyl) 1-carbonyl) -HisTrpAlaValD-AlaHisD-Pro S? PheNH ₂

N - (((2´-methoxy) -3-phenyl) propionyl) -HisTrpAlaValDAlaHisD-Pro * Phe-NHg

K- (((2 ', 5'-dimethoxy) -3-phenyl) propionyl) -HisTrpAlaValD-Pro' i 'Phe-NHg

N - ((3-phenyl) -piperonyl) -Hi with TrpAlaValD-AlaHisD-Propyr-NHg

K - (((2 *, 3'-dimethoxy) -3-phenyl) propionyl) HisTrpAlaValD-AlaHisD-Pro Phe-NH ₂

1:46:10 ^{- 11} 1095 Ala (2.00),

His (1.55), Val (1.00)

1.77.10 ^{- 8} 1131 Ala (2.22),

Hisd, 44), Val (1.00)

9/12/10 ^{- 9} 1095 Ala (2,68)

Hisd, 38), Val (1.00)

8.66.10 ^{- 9} 11 58 Ala (2.7D,

His (1.08),

Val (1.00)

9.00.10 ^-9 1111 Ala (2.07),

Hisd, 35), Val d (00)

8.76.10 ^{- 9} 1141.7 Ala (2.23),

Hisd, 67)

1.82.10 ^{- 8} 1097.7 H / A

3,50.10 ⁹ 1141/7 Als. (2,25),

Hisd, 66), Val d, 00),

Structure of IC, _n (M) MH ⁺ (PAB) ⁵ MS

AAA

N - ((3-phenyl) propionyl) HisTrpAlaValD-AlaHi (3- (2-pyrrolidinyl-3hydroxy) propionyl) -Phenh ₂ ((isoquinolylycarbonyl) -H is TrpAlaValD-AlaHi-DPro 'S' Phe-NH ₂

1.76.10 ^-8 1139.6

3,44.10 ⁹ 1104,5

N - ((3-phenyl) propionyl) - 7.50.10 ^-8 1067.1

HisTrpAla 'P ValD-rAlaHisDPro-Phe-NH ₂

ON THE

Ala (1,82), Hís (1,32), Val (1,00)

Ala (1.00), Hiss (3.17) · i

P1 / 33-33

-43- ‘JC-. EO E5JDJE 3SS00

Claims (12)

PATENT CLAIMS The polypeptide of formula I to XX ¹ TrpX ² X ³ X ⁴ X ⁵ X ⁶ X ⁷ NH _2/1 / wherein X is X®Arg (or D-Arg) X X ^ ^ and the Χθ _des-NH2 Pro, TyrPro , des-NH ₂ TyrPro, AdaProD-Pro or is deleted, g X is Gly ^ 1a, D-Ala or is deleted, χΐθ is Asn, Phe, D-Phe, or Phe or D-Phe substituted with at least one halogen atom, or X is A- (CH ₂ ) _n -COwhere A is a group containing 1 to 3 rings, of which at least one ring is aromatic, wherein each ring system is optionally substituted, and the alkylene group is optionally substituted with one to four groups selected from amino, hydroxy, C 1-6 alkoxy and C 1-6 alkoxy; C 1-6 alkyl optionally substituted by halogen atom and n is 0 to 4, Q or X is cyclopentylcarbonyl substituted by X with Arg (or D-Arg) X 1 X X 1 as defined above, X ¹ is His, ThiAla or is deleted, X ² is Ala, D-Ala, CPenc, D-tBuGly or Pro, X ³ is Val or Val substituted by at least one halogen atom, X is Gly, Ala, D-Ala, sarcosine, Pro, L-Pro or L-Phe, X 1 is His or ThiAla; - 44 y ^{Σ σ} is D-Pro Τ ', Pro Τ', 2-pyrrolidinyl-3-hydroxypropionyl or B-Pro, is No, Leu, Phe, Val, Mox, D-Phe, Phe or D-Phe substituted with at least one halo, a gene atom or naphthylAla or naphthyl D-Ala, or a hydrophobic substituted aromatic amino acid or aralkyl amine, or is deleted, or a pharmaceutically acceptable salt thereof. 2. The polypeptide of formula I according to claim 1, wherein X is a group A- _{(CH2) n-CO-} wherein A is phenyl, naphthyl, phenothiazinyl or indolyl, optionally substituted with substituents selected from hydroxy, phenyl, halogen, C ^ _ ^ alkyl or C 1-4 alkoxy, optionally substituted with halogen, n is 2, and X ¹ , X 4, X 4, χ ⁴ , χ 5, χ ⁷ _{has the} meaning given in point 1, or a pharmaceutically acceptable salt thereof. 3. The polypeptide of formula I according to item 1, wherein X is des-NH ₂ Phe, des-NH ₂ Tyr or des-NH ₂ TyrPro- (or D-Pro) Arg (or D-Arg), and X ¹ is His or ThiAla, X is Ala, Pro, X 1 is Val or hexafluorvaline, X ⁴ is D-Ala, D-Phe, X ^ is His, ThiAla, X ⁶ is D-Pro Τ ', Pro Τζ Pro, D-Pro v X is No or Phe, Leu, methoxinin, 2-naphthyl-2-alanine or a pharmaceutically acceptable salt thereof. 4. The polypeptide of formula (I) according to any one of items 1 to 3 which is N ((3-phenyl) propionyl) HisTrpAlaValD-AlaHisD-Pro S / Phe-NH ^ N - (((4'-hydroxy) -3-phenyl) propionyl) HisTrpAlaValD-AlaHisDPro '5 ^/ Phe-NH ₂ N- (1-naphthoyl) -HisTrpAlaValD-Pro 'S' Phe-NH ₂ N - (((4'-Methoxy) -3-phenyl) propinyl) -HisTrpAlaValD-AlaHisDProS / Phe-NH ₂ N - ((2-methyl-2-phenyl) pro'pionyl) -HisTrpAlaValD-Phe-Ψ AlaHisDPro NH _2, or a pharmaceutically acceptable salt thereof. 5. A polypeptide of formula I according to any one of items 1 to 4, or a pharmaceutically acceptable salt thereof, for use as a therapeutic agent. 6. Use of a polypeptide of formula I according to any one of items 1 to 4, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament suitable for inhibiting the growth of cells that are sensitive to the growth promoter activity of GRP. 7. Use of a polypeptide of formula I according to items 1 to 4, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for the treatment of cancer. 8. A method of inhibiting cell growth that is susceptible to the growth promoter activity of GRP in a mammal, comprising administering to said mammal a therapeutically effective amount of a polypeptide of formula I as defined in any one of items 1-4 or a pharmaceutically acceptable salt thereof. - 46 9. The method of claim 8, wherein the cancer prophylaxis is treated or performed. 10. A method according to any of the sides 8 to 9, wherein the mammal is a human. 1. A medicament characterized in that the content of the _/ .p'6lypeptič of formula I as described in any one of claims / to 4, or a pharmaceutically acceptable salt thereof, a pharmaceutically acceptable carrier and optionally at least one other pharmaceutically active agent. * _W 12. A medicament according to paragraph 11 characterized in that it is in the form of a tablet or capsule. 13. A process for preparing a polypeptide of formula (I) according to any one of Claims 1 to 4, which comprises condensing amino acids and / or derivatives of amino acids in the desired order and / or peptide fragments containing these amino acids or derivatives thereof in the desired order to obtain the desired peptide. wherein either the carboxylic acid end group or the amino end group is activated for the peptide bond and the remaining groups are protected and optionally the resulting peptide deprotected and / or converted to its pharmaceutically acceptable salt. 14. A method of inhibiting the binding of gastrin-releasing peptides in cells capable of binding, comprising exposing said cells to an inhibitory amount of a compound according to item 1, 2, 3 or 4.