NO760707L - - Google Patents

Description

"Procedure for the manufacture of

new organic compounds."

The present invention relates to a method for the production of new compounds of formula I

wherein R means (i) H

(ii) NH2,

(iii) H-Ala-NH-; H/.Tyr-NH-; H-Cys-NH-; H H -Val-NHT; H-Lys-NH-,' "H-(α-amino-isobutyryl)-NH-; H-Phe-NH-; (iv) NH2-NH-CO-NH; (v) CgHg.NH-CO-Gly-NH-; H-Ala- Gly-NH-; H-Val-Gly-NH-; H-α-phenyl-Gly-NH-; (vi) R2-A-CO-NH-; wherein R2 stands for lower alkyl, phenyl or phenyl substituted with halogen, hydroxy, amino, lower alkyl or lower alkoxy,

A stands for a direct bond, an alkylene radical with in each case up to 6 carbon atoms or for -NH-(vii) R2-Ala-NH-;

R 2 -Ala-Gly-NH-;

in which R^ has the above meaning,

-iviii) CcH!-.AI.CO-M-NH-,

' j stay

in which A stands for an alkylene radical with up to 06 carbon atoms and

-M- means (mono- or di-Hal)-Phe-Gly- or -Ala-,

(ix) T- means (mono- or di-Hal)-Phe-NH-, where T stands for H-, H-Val- or benzoyl,

wherein A"<1>" has the above meaning and L- stands for -Val-, -Val-(mono- or di-Hal)-Phe-,.-a-phenyl-Gly- and n stands for a whole numbers between I and 10,

B stands for H or both B stands for a direct bond between the S atoms,

X stands for -Lys-, -Nie- or -Cys- and

Y stands for -Asn-, -Gin- or -Thr- and

Z stands for the residues (i) H; (ii) COOH; (lii) C00R3 (wherein R3 is a lower alkyl group); (iv) -CO-NR^R,- (wherein R^ and Rj- independently of each other mean hydrogen or a lower alkyl group); (v). -CON CCH^) , (where n has the above meaning); (vi) CH 2 OH; (vii) -CO-asparaginol; -CO-1 eucinol; -CO-isoleucinol; -CO-valinol; -CO-norleucinol; -CO-glutaminol; -CO-theoninol; (viii) -CO-leu;

-CO-ser; -CO-iso-leu; -CO selection and

their corresponding amides, in which the amide means the residue -CO-NR.R,-

or

(wherein n, R₂ and R₂- have the above-mentioned meaning), or (ix) -CO-ThrOR₃ or -CO-SerOR₂ (wherein R₃ has the above-mentioned meaning); (x) -CO-NH-CCH^jc-OH (wherein k means an integer between 2 and 6), or (wherein 1^ and 1^ independently of each other means an integer between 1 and 6); (xii)

(xii) -CO-NH-Rg (wherein Rg means a 5- or 6-membered saturated heterocyclic ring containing an oxygen atom as a ring member) where in formula I the radicals R^, X, Y and Z as also with the other units in the peptide sequence at least one unit can be in the D form,

when X means Lys and Y Asn and all other units in the peptide sequence 3 to 14 exist in the D or L form, and when

(i) R means H-Ala-Gly-NH- or H-(D)-Ala-Gly-NH, Z cannot be COOH, COOR3, CH2OH or CONR4R5or, when (ii) R means H, NH^ or H -(D)-Ala-Gly-NH-, Z. cannot stand for H, COOH, CONR„R,- or

„ ■'■ ,, -'. c ell is , when 4- or 5 '

tmj R 2 means H-toc-amino-isobutyryl; -NH- or H-(D)-Ala-NH-,

cannot Z stand for H or COOH or, when

(iv) R means H-Ala-Gly-NH-, '

cannot Z stand for H or, when

(v) R 1 is H-Ala-NH- or.

R2" -CO-NH-, (where R2<1>does not stand for lower alkyl or unsubstituted phenyl) Z cannot stand for COOH, and

in that when R means H-Cys-NH or N-(D)-Cys-NH and/or X means

-Cys- or -(D)-Cys, B-B cannot account for a direct bond.

Halogen means bromine, fluorine and preferably chlorine. The meaning of lower alkyl or lower' alkoxy includes up to 6 carbon atoms,

but preferably means 2 or 1 carbon atom.

If R 2 means substituted phenyl, then this is preferably mono- or di-substituted, but especially mono-substituted. A substituent is preferably located in the para position.

When A"1" or when A denotes alkyl, these preferably contain

up to 3 carbon atoms. With respect to meaning (viii), A X preferably means trimethylene. With regard to the meaning (x), A preferably means methyl enr -

Examples of -(mono- or di-Hal)-Phe moieties are -(2Cl)Phe; -(3Cl)Phe-;

-(4Cl)Phe- or -(3,4-diCl)Phe-.

n preferably means an integer 5 or 4.

means preferred e.g. H-(mono or di-Hal)-Phe-NH-, phenyl CO-NH, H-Val-(mono- or di-Hal)-Phe-NH-, phenylbutyryl-NH-

or a residue containing -(D)-Ala, benzoyl-(mono- or di-Hal )-Phe-NH-, cyclohexylacetyl-α-phenyl-Gly-NH-, cyclohexylacetyl-Val-(mono- or di-Hal )-Phe-NH-, cyclohexylacetyl-Val-NH-, H-Val-NH, H, H-(cc-am.ino-isobutyryl )-NH-, H-(mono- or jdi-Hal)-Phe- NH-, X means preferably Lys and especially Nie,

Y means preferably Gin and especially Asn.

k means preferably 2 or 3.

When Z has the meaning (x) or (xii), these preferably comprise a -CO-NH-CH-CH2-0 sequence and mean e.g. -CO.NH.CH(CH2vOH)CH2. CH2-CH2.OH;

Z preferably does not mean H, CH2OH, COOH or COOR3.

In the preferred sense given above, the amino acids are present both in the D-form and also in the L-form.

The compounds can be prepared according to the synthesis of compounds

of this type of commonly known methods or similar chemical equivalents.

E.g. the compounds with the above formula can be prepared by a) at least one protecting group, which is present in a protected compound of formula I with the specified sequence, is removed, or b) by two peptide units, each of which contains at least one amino acid in protected or unprotected form, connect with each other

by means of an amide bond, the peptide bond must take place in such a way that the amino acid sequence contained in formula I is produced

and then optionally the process step a) is carried out, or c) while transferring a group Z into another group Z with the above definition, whereby an unprotected or protected

compound with formula I is obtained and in the latter case process step a) is carried out, or

d) as a compound of formula I, in which B stands for hydrogen, is oxidized, whereby a compound of formula I is obtained in which

B-B means a direct bond.

This involves methods known in and of themselves in peptide chemistry and these can be carried out analogously to the methods described in the following examples.

To the extent that the preparation of the starting products is not specifically described, these compounds are known or can be prepared and purified according to methods known per se. Likewise, the starting product can be produced analogously to the methods described in the examples.

The compounds can be present in salt form or in the form of their complexes.

As acid addition salts, the salt of hydrochloric acid and acetic acid are mentioned in particular.

Complexes are understood to mean the complex-like compounds known in the field with not yet elucidated structure, which are formed by the addition of certain inorganic or organic substances to the compounds of formula I. Such inorganic substances are compounds derived from metals such as calcium, magnesium, aluminum, cobalt and especially zinc, above all sparingly soluble salts, such as phosphates, pyrophosphates and polyphosphates as well as hydroxides of these metals, especially alkali metal polyphosphates. Organic substances are e.g. non-antigenic gelatins, e.g. oxypolygeliatins, polyvinylpyrrolidone and carboxymethylcellulose, further sulphonic acid or phosphoric acid esters of alginic acid, dextran, polyphenols and polyalcohols, above all polyphloretin phosphate and phytic acid, as well as polymers of amino acids, e.g. protamine, polyglutamic acid or polyaspartic acid.

The compounds are distinguished by pharmacological activity. In particular, they act as an inhibition of sex hormone secretion. They therefore find use in Diabetes Mellitus, in the treatment of acromegaly, angiopathy, as well as the use of these compounds for diagnostic purposes.

The compounds produced by the method according to the invention can be supplied in a pharmaceutically acceptable salt form, especially in the form of their complexes. Such salt or complex forms can be prepared according to methods known per se from the free bases and vice versa. The compounds of formula I can be contained in pharmaceutical preparations in a pharmaceutically acceptable salt or complex form and in admixture with a carrier or a solvent. Such pharmaceutical preparations can be available as tablets or as injection or infusion solutions. A pharmaceutical solution

is preferably buffered at a pH between 5 and 8.

Pharmaceutical preparations can e.g. contain compounds of formula I in which H-(α-amino)-isobutyryl-NH— or H-(D)-Ala-NbkX means Lvs, Y means Asn, Z means -CONR^R,. or

4 or 5<*><F>overstretched, each B stands for hydrogen.

In the following examples, all temperature indications are in degrees Celsius and are uncorrected.

The following abbreviations are used:

Cbo = carbobenzyloxy

MBzl = p-methoxybenzyl

Iabu = H-(α-amino-isobutyryl)-

(4Cl)Phe = P-chlorophenylalanyl

ACOH = acetic acid

OCP = trichlorophenyl ester

Trp = L-tryptof a.nyl

OSu = o-succinimide

All final compounds are characterized in hydrochloride salt form. Unless otherwise stated, the amino acids are in the L form. In the following table /pc/D 20 refers to ACOH 1% volume/volume (c = l), when not stated otherwise.

(1) refers to ACOH 20% v/v (c = l)

(2) refers to ACOH 1% v/v (c = 0.25)

(3) refers to ACOH 30% v/v (c = 1)

(4) refers to ACOH 40% v/v (c = l)

(5) refers to ACOH 50% v/v (c = l).

Example 1: H-(4Cl2§he-Cys-Lys-Asn-Ph

Ser-Cys-OH

480 mg Cbo-(4C1)Phe-Cys(MBzL)-Lys(BOC)-Asn-Phe-Phe-Trp-Lys(Cbo)-Thr-Phe-Thr-Ser-Cys(MBzl)-OBzl, 410 mg H -Try-OH, 320 mg H-Cys-OH. HCl and 1.2 ml of thioanisole are dissolved at 0°C in 5 ml of trifluoroacetic acid and 10 ml of boron rice (trifluoroacetate) 2 molar in trifluoroacetic acid are added.

The mixture is shaken for 1 hour at room temperature, cooled and 50 ml of ibert.-butyl alcohol is added, the irin is evaporated in vacuo and the residue is added to a solution of 0.01 M 2-mercapto-ethanol in ether. Then centrifuged. The solid residue is dissolved in a little 10% acetic acid and purified by chromatography on "Sefadex" G 25 - in the system 0.01 M mercaptoethanol in 10% acetic acid. - The residue can also be dissolved in 30% acetic acid or in a mixture of n-butanol /glacial vinegar/water (e.g. 4:2:1) and purified by chromatography on "Sefadex" G 25 in the system 0.01 M mercaptoethanol in 30% acetic acid or in the butanol/glacial acetic acid/water mixture. The fractions containing the desired product are combined, dissolved in 0.1 n hydrochloric acid and freeze-dried, whereby the compound mentioned in the title is obtained.

Temp. 195°C (decomposition); /q/^° = -20.5° in AcOH (c = l)

The starting material is produced in the following way:

a) Cbo-Lys_(B0C2 zAsn-Phe-0Me

74.8 g of Cbo-Lys(BOC)-ONP are reacted in a solution of 51 g of H-Asn-Phe-

OMe » HC1 and 21 ml of triethylamine in 300 ml of dimethylformamide.. After work-up, the compound mentioned above is obtained.

Temp. 175°Cj r& Q0- -4.4° in dimethylformamide (c =1.0)

b^ ^zkysi?OC2 zAsn-Phe-OMe

92 g of Cbo-Lys(B€>C)-Asn-Phe-OMe are suspended in a solution of 2 1 dioxane and 400 ml of water. Palladium charcoal is added and hydrogenated under normal pressure. After processing, the compound mentioned in the title is obtained.

Temp. 122°C; /a/^<0>= -8.3° in dimethylformamide (c = 1.0)

c) 2^2zi å?i2E!}®z9Ysi^?2i2z^!jz^^3

20 g of H-Cys(MBzlO-OMe.CH3S03H and 7 ml of triethylamine are dissolved in 120 ml

dimethylformamide and 18.2 g of Cbo-(4Cl)Phe-0NP are added. The mixture is set aside for 16 hours and evaporated at room temperature, with CBo-(4Cl)Phe-Cys(MBzl)-0Me (m.p. 115°C) subsequently obtained. processing. The latter compound is dissolved in methanol and hydrazine hydrate is added. The mixture is set aside for a day at room temperature.

The connection mentioned in the heading is obtained after processing.

Temp. 158°C -; /a7^° = -9.9° in dimethylformamide (c = l)

<d>) 9°2zL^Cl)Phe-Cys^MBzl) =Lys (§0C^-Asn-Phe-NH-NH2

Dissolve 6 g of Cbo-(4Cl)Phe-Cys(MBzl)-NH-NH2 in 80 ml of dimethylformamide

cooled to -20°C, 6 ml of 5.3 N hydrochloric acid in dioxane and then 1.3 ml of tert-butyl nitrite are added and stirred for 10 minutes at

-20°C. After adding 5.5 ml of triethylamine at -20°C, 8.4 g of H-Lys(BOC-)--Asn-Phe-OMe are added, the mixture is set aside overnight at 0°C, evaporated, water is added at 8° C to pH-2.. The precipitate is filtered off and washed with water. The Cbo-(4Cl)Phe-Cys (MBzl)-Lys(BOC)-Asn-Phe-OMe formed is recrystallized from methanol, dissolved in dimethylformamide and 7 ml of hydrazine hydrate are added. The mixture is allowed to stand for a day at room temperature, evaporated and the residue recrystallized

from methanol, whereby the compound mentioned in the title is obtained.

in

Temp. 260°C; /a7p° = -23° in dimethylformamide (c = 1,'io)

e ) ?29=tYsiCbo2 ::Thr ::Phe-Thr-Ser-CYS_( MBzl 2-OBzl

In an analogous manner to under section d), 2.3 g of BOC-Lys(Cbo)-Thr-Phe-Thr-Ser-NH-NH2 are reacted with 1.86 g of H-Cys(MBzl)-OBzl, whereby the

the connection mentioned in the heading is obtained.

Temp. 169°C; Z"a7p° = -16° in dimethylformamide (c = l)

<f>) ?22z?£Ez^y 9^°2z^]}£zEt}Éz^£z§S£z2^i??5^i2z2?5i 2.3 g BOC-Lys(Cbo)-Thr -Phe-Thr-Ser-Cys(MBzl)-OBzl resolves in a

mixture of 10 ml of methylene chloride and 6 ml of trifluoroacetic acid and set aside for 25 minutes at room temperature. H-Lys(cbo)-Thr-Phe-Thr-Ser-Cys(MBzl)-OBzl.trif}uoroacetate is then precipitated with ether, filtered off and washed well with ether. The residue is dissolved in 7 ml of dimethylformamide, 1.5 g of BOC-Trp-OCP is added and 0.3 ml of triethylamine is added and left overnight at room temperature. The product is precipitated with ether/ethyl acetate (1:1) and filtered. It is dried and this heading newnte fdrbindelse.obtained.

M.p. 167°C; Zo/d° = -17.5° in dimethylformamide (c = l)

9) 222zEI}<®>z5,EPzt:Yfi2E22z^£zEl}2^II}£é§^£z2Y£i!!}§^i2z2?5i

36 g of BOC-Trp-Eys(Cbo)-Thr-Phe-Thr-Ser-Cys(MBzl)-OBzl are reacted with 20 g of BOC-Phe-ONP in an analogous manner as under section f) and the compound mentioned in the title is obtained .

Temp. 210°C; /a7p° = 15° in dimethylformamide (c =1.0)

h) £bo-(fCOPhe^Cy^ (MBzJ.^^ ^!}£zEl22z5'l}£z§2E~2y^l^?^i2z255i.

1.5 g of BOCiPhe-Trp-Lys(Cbo)-Thr-Phe-Thr-Ser-Cys(MBzl)-OBzl are dissolved in 30 ml of methylene chloride and 30 ml of trifluoroacetic acid. The mixture is allowed to stand for 25 minutes at room temperature, evaporated in vacuo and

.poured with ether. After filtration, washing with ether and drying

is held H-Phe-Tr p-Lys (Cbo)-Thr-Phe-Thr-Ser-Cys (MBzl)-OBzl-r trifluoroacetate.

Dissolve 0.9 g of Cbo-(4Cl)Phe-Cys(MBzl)-Lys(BOC)-Asn-Phe-NH-NH2 in 20 ml of dimethylformamide, cool to -20 C and add 0.6 ml of 5.3 N hydrochloric acid in dioxane, then Ojl ml of tert-butyl nitrite and stirred for 15 minutes at -20°C. 0.7 ml of triethylamine and 1.5 g of H^Phe-Trp-Lys(Cbo)-Thr-Phe-Thr-Ser-Cys (MBzl )-OBzl v trifluoroacetate are added. The mixture is allowed to stand overnight at 0°C, evaporated in vacuo, carried out with 100 ml of water, filtered, the residue is washed with water and then with methanol, heated in methanol, filtered and the title compound is obtained.

Temp. 270°C; C& 2^ = -19.3° in dimethylformamide (c = l)

In an analogous way, after building up the_corresponding protective compounds, the following compounds of formula A are obtained after cleavage of the protective groups

Example 2: 0.4 g of the compound from example A33 is dissolved in a mixture of methanol and 0.5% acetic acid and the pH is adjusted by adding ammonium hydroxide solution while stirring to pH 6. While stirring, 0.1 m potassium ferricyanide solution is slowly added until the yellow color becomes permanent. After a further 15 minutes of stirring, the pH is adjusted to 4-5 with acetic acid. It is filtered through a short column of "Bio Rad" AG 3/x4 (Cl ~ -form) £= weakly basic anion exchanger7.- The filtrate is then allowed to flow slowly at 0°C through a column of "Bio Rex" 70 ( -C00H- form) { = weakly acidic cation exchanger7. The column is then eluted with an acetic acid gradient of increasing acetic acid concentration or with an acetic acid-pyridine gradient. Fractions containing the desired product are combined and freeze-dried. The freeze-dried product is dissolved in a little n-butanol/glacial vinegar/water (2:1:1) or 30% acetic acid and purified by chromatography on "Sefadex) G 25 in the same mixture. The fractions containing the desired product are combined, freeze-dried and the connection mentioned in the heading is obtained.' In an analogous manner, the corresponding compounds with a free SH function (example 1, AI-7, A10-32, A34-50) are prepared from the corresponding compounds with an S-S bridge.

In an analogous way as described in example 1, the following compounds of formula A are prepared, in which:

The compounds are also converted into the corresponding ring-closed forms. It is noted that these ring-closed forms can be characterized by reduction back to the open chain forms.

Claims (4)

1. New. compounds of formula I wherein R-^ means (i) H, (ii) NH2 , (iii) H-Ala-NH-; H-Tyr-NH-'; H-Cys-NH-; H-Val-NH-; H-Lys-NH-; H-(oc-amino-isobutyryl)-NH-; H-Phe-NH-; (iv) NH2 -NH-CO-NH-;■' ■(v) GgHg.NH-CO-Gly-NH-; H-Ala-Gly-NH-; H-Val-Gly-NH-;. H-cc-phenyl-Gly-NH-; (vi) Ro-A-C0-NH-, in which R 2 stands for lower alkyl, phenyl or phenyl substituted with halogen, hydroxy, amino, lower alkyl or lower alkoxy, A stands for a direct bond, an alkylene radical. with in the individual case up to 6 carbon atoms or for -NH-, (vii) R 2 -Ala-NH-; R 2 -Ala-Gly-NH-; wherein R 2 has therein the above meaning, (viii) C6 H5 .A <I> .CO-M-NH-, in which A1 stands for an alkylene radical with up to 6 carbon atoms and -M- means (mono- or di-Hal)-Phe-, Gly- or -Ala-, (ix) T-.(mono- or di-Hal) -Phe-NH-, where T stands for H-, H--Val- or benzoyl, wherein A has the above meaning and L stands for -val-, -val- (mono- or di-Hal)-Phe-, -cc-phenyl-Gly and n stands for an integer between 4 and 10, B stands for H or both B stands for a direct bond between the S atoms, - X stands for -Lys-, -Nie- or -Cys- and Y stands for -Asn-, Gin- or -Thr- and Z stands for the residues (i) H; (ii) COOH; (lii) COOR3 (wherein R3 means a lower alkyl group); (iv) CO-NR^R,- (in which R^ and R^ independently of each other mean hydrogen or a lower alkyl group) to (where n has the above meaning); (vi) CH 2 OH; (vii) -CO-Asparaginol; -CO-1 eucinol; -CO-isoleucinol• eO-valinol; -CO-norleucinel; -CO-glutaminol; -CO-threoninol; (viii) -CO-leu; -CO-Ser; -CO-isoléu; -CO selection and their corresponding amides, where amide means the residue -CO-NR.R,- ell is (wherein n, R^ and R,- have the above-mentioned meaning), or UxJ -CO-ThrOR3 or -CO-SerOR3 , Where R3 has the above-mentioned ■ meaning; (x) -C0-NH-(CH2)-OH (where k means an integer between 2 and 6), or each other (where 1^ and 12 regardless of each other means an integer between 1 and 6); (xi) (xii) -CO-NH-Rg (wherein Rg means a 5- or 6-membered saturated heterocyclic ring., which contains an oxygen atom as a ring member, or means a 5-' or 6-membered lactone ring), while in formula I radicals R , X, Y and Z as well as with the other units of the peptide sequence at least one unit can be present in the D form, when X means Lys and Y means Asn and all other units - in the peptide sequence 3 to 14 are present in The D. or L shape, and when (ij R1 means H-Ala-Gly-NH or H-(D)-Ala-Gly-NH, Z cannot stand for COOH, COOR.3 , CH2 0H or CONR4 R5 , or when (ii) . R. means H, NH0 or H-(D)-Ala/Gly-NH-, Z cannot stand for H, COOH, CONR4 R5 or or 5' or, where (iii) R means H-(α-amino-isobutyryl)-NH- or H_(D)-Alα-NH-, cannot Z stand for H-or COOH, or when (iv)' Ra means H-Ala-Gly-NH, can Z not stand for H or, when iy) R means H-Ala-NH, or R2 -CO-NH, (where R,-,' stands for lower alkyl or unsubstituted phenyl) can Z does not stand for COOH, and when R means H-Cys-NH or H-(D)-Cys-NH and/or X-Cys- or -(D)-Cys, B-B cannot account for a direct bond. 2. The process for the preparation of the compounds of formula I stated in claim 1, in which R , B, X, Y and Z have the above meaning, as well as acid addition salts and complexes of these compounds, characterized in that the compounds with the above formula are prepared using methods known in peptide chemistry and optionally then transferred in a manner known per se in their acid addition salts and complexes. 3. Compounds as set forth in claim 1, wherein when R 1 means H-(α-amino)-isobutyryl-NH- or H-(D)-Ala-NH, X means Lys, and Y means Asn, Z does not stand for -CO-NR^^ or or- 5 <*> 4. Method as specified in claim 2, characterized in that the compounds of formula I specified in claim 3 are prepared.