CA2157215A1 - New 4-aminopyridines - processes for their production as well as pharmaceutical agents containing them - Google Patents

Abstract

Compounds of formula I

(I), in which R1 denotes an aryl, a heteroaryl or a cycloalkyl group which can be substituted if desired, AS denotes an amino acid n denotes the numbers 0 or 1, R2 and R3 are the same or different and denote hydrogen atoms, alkyl, carboxyalkyl or alkoxycarbonyl-alkyl groups, or R2 and R3 together with the nitrogen atom to which they are bound, form a heterocyclyl ring which, if desired, can in addition contain a second heteroatom and can be substituted by alkyl, carboxy or alkoxycarbonyl groups, R4 and R5 are the same or different and denote hydrogen atoms or alkyl groups, m denotes the numbers 0, 1 or 2, R6, R7, R8 and R9 are the same or different and denote hydrogen atoms or halogen atoms, as well as hydrates, solvates and their physiologically tolerated salts thereof.

The invention also concerns optically active forms, racemates and diastereomer mixtures of these compounds and processes for their production and pharmaceutical agents which contain these compounds in particular for the production of pharmaceutical agents for the treatment of thromboembolic diseases.

Description

Boehringer Mannheim GmbH

New 4-aminopyridine~ - proce~e~ for their production as well a~ pharmaceutical agent~ containing them The invention concerns new 4-aminopyridines of the general formula I

~'N
Rl-SO2-(AS)n-NH~ R3 (CjR4Rs)m (I), HN
E~R6 in which R1 denotes an aryl, a heteroaryl or a cycloalkyl group which can be substituted if desired, AS denotes an amino acid n denotes the numbers O or 1,

2 and R3 are the same or different and denote hydrogen atoms, alkyl, carboxyalkyl or alkoxycarbonyl-alkyl groups, or R2 and R3 together with the nitrogen atom to which they are bound, form a heterocyclyl ring that can additionally contain a second heteroatom if desired and can be substituted by alkyl, carboxy or alkoxycarbonyl groups, 4 and R5 are the same or different and denote hydrogen atoms or alkyl groups, m denotes the numbers 0, 1 or 2, 6, R7, R8 and Rg are the same or different and denote hydrogen atoms or halogen atoms, as well as hydrates, solvates and physiologically tolerated salts thereof. The invention also concerns the optically active forms, racemates and mixtures of diastereomers of these compounds.

The invention also concerns processes for the production of the above-mentioned compounds, pharmaceutical agents that contain such compounds as well as the use of these compounds in the production of pharmaceutical agents.

The aminopyridines of the general formula I, their solvates and their salts inhibit thrombin-induced coagulation of fibrinogen in blood as well as thrombin-induced aggregation of blood platelets. Thus they prevent formation of hyaline thrombi and platelet-rich thrombi and can be used to combat and prevent diseases such as thrombosis, apoplexy, coronary infarction, inflammations and arteriosclerosis. Furthermore these compounds have an effect on tumour cells and prevent formation of metastases. As a result they can be used as anti-tumour agents.

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Thrombin, the last enzyme of the coagulation cascade, cleaves fibrinogen to form fibrin which is cross-linked by factor XIIIa and becomes an insoluble gel which forms the matrix for a thrombus. Thrombin activates platelet aggregration by proteolysis of its receptor on the blood platelets and in this way also contributes to thrombus formation. When a blood vessel is damaged these processes are necessary in order to stop bleeding. No measurable thrombin concentrations are present in blood plasma under normal conditions. Increases in the thrombin concentration can lead to the formation of thrombi and hence to thromboembolic diseases which occur very frequently above all in industrial countries.

Thrombin in plasma is kept ready in the form of prothrombin and is released from it by factor Xa.
Thrombin activates factor VIII which together with factor IXa, then converts factor X into factor Xa. By this means thrombin catalyzes its own release which is why very rapid increases in thrombin concentrations can occur.

Thrombin inhibitors can therefore inhibit the release of thrombin, the platelet-induced and plasmatic blood coagulation.

There is a whole series of serine proteases apart from thrombin that cleave peptide substrates next to a basic amino acid. In order to limit side-effects, the thrombin inhibitors should be selective i.e. they should inhibit other serine proteases only slightly or not at all.
Trypsin in particular being the least specific serine protease, can be easily inhibited by the various inhibitors. Trypsin inhibition can lead to pancreatic stimulation and to pancreatic hypertrophy (J.D. Geratz, Am. J. Physiol. 216, (1969) p. 812).

Plasma contains the protein plasminogen which is converted into plasmin by activators. Plasmin is a proteolytic enzyme whose activity is similar to that of trypsin. It serves to dissolve thrombi by degrading fibrin. Inhibition of plasmin would thus have the opposite effect to that which one would like to achieve by inhibiting thrombin.

Synthetic thrombin inhibitors have already been known for a long time. Substances of the (D)-Phe-Pro-Arg type were synthesized from fibrinogen the natural substrate of thrombin. Such tripeptides imitate the amino acid sequence before the cleavage site on fibrinogen. In order to obtain good inhibitors the carboxylate group of the arginine was changed in such a way that the hydroxy group of serine 195 in the active site of thrombin can react with it. This can for example be achieved by replacing the carboxylate group by an aldehyde group.
Corresponding (D)-Phe-Pro-arginals are described in the Patent Application EP-A 185390.

Benzamidine, a known trypsin inhibitor, was used as the basis for a second type of thrombin inhibitors. The inhibitors obtained in this way do not only differ from the (D)-Phe-Pro-Arg types in their chemical structure but also in the way they inhibit: serine 195 of thrombin does not bind to these inhibitors. This clearly follows from X-ray examinations of the structure (W. Bode, D.
Turk, J. St~rzebecher, Eur. J. Biochem. 193, 175-182 (1990)). N-(2-naphthylsulfonylglycyl)-4-amidino-(R,S)-phenylalanine-piperidide ("NAPAP", DD 235866) belongs to this second class of thrombin inhibitors.

A disadvantage of inhibitors of the (D)-Phe-Pro-Arg class compared to other serine proteases is the lack of selectivity (J. C. Powers, C.-M. Kam, in Thrombin, Structure and Function, (L. J. Berliner, publisher), Plenum, New York 1992, p. 117). The selectivity of NAPAP
is somewhat better. The inhibition constants of NAPAP
are as follows (J. Sturzebecher et al., Pharmazie 34 (1988), p. 782): thrombin 6 nM, trypsin 0.69 ~M, plasmin 30 ~M. The selectivity of this inhibitor between thrombin and trypsin, expressed as a quotient of the inhibition constants is therefore about 1:100. The biggest disadvantage of these inhibitors is that when administered orally, they do not reach their site of action, i.e. the bloodstream, or only inadequately. The strong basicity of the inhibitors is regarded as being mainly responsibility for this deficiency. Since thrombin selectively recognizes the amino acid arginine, it is not surprising that the inhibitors also contain groups with a similar basicity to that of the guanidino group of arginine. The pKa value of the side chain of arginine is 12.5 (D. Voet, J.G. Voet, Biochemie, "VCH-Verlag Weinheim 1992", p. 60), the pKa value of benzamidine is 11.8 (Albert, J. Chem. Soc. 1948, 2240).

There has been no lack of attempts to develop thrombin inhibitors with fewer basic ~ou~ in order to improve oral availability. St~rzebecher et al (loc.cit.) for example produced a compound which only differs from NAPAP by the replacement of the benzamidino group by a benzylamino group. Benzylamine has a significantly reduced basicity compared to benzamidine: pKa = 9.35 ~Robinson, Trans. Faraday Soc. 52 (1956), 327). However, this modification of NAPAP led to a fall in the inhibitory effect towards thrombin by several powers of ten to Ki = 19 ~M and the selectivity thrombin/trypsin was only 1:4, i.e. it nearly completely disappeared.

It was now surprisingly found that 4-aminopyridines of the general formula I are strong and selective thrombin inhibitors although 4-aminopyridine with a pKa of 9.29 (J. M. Essen, K. Schofield, J. Chem. Soc. 1961, 3939) has a similarly low basicity to that of benzylamine. Not only the oral availability is improved by this means, but it also improves the tolerance and reduces the fall in blood pressure which is observed with NAPAP
derivatives.

R1 in the general formula 1 is understood as a phenyl, naphthyl and anthryl group which, if desired, can be substituted by 1-5 identical or different substituents such as halogen, nitro, nitrile, phenyl, cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, trifluoromethyl, trifluoromethoxy, C1-C8 alkyl, C1-C8 alkenyl, C1-C8 alkinyl, hydroxy, C1-C8 alkyloxy, C1-C8 alkenyloxy, C1-C8 alkinyloxy, amino, C1-C8 alkylamino, C1-C8 alkenylamino, C1-C8 alkinylamino, di-(C1-C8 alkyl)amino, benzylamino, dibenæylamino, carboxyl, C1-C8 alkyloxy-carbonyl, aminocarbonyl, C1-C8 alkylaminocarbonyl, di-(C1-C8 alkyl)aminocarbonyl, C1-C8 alkylthio, C1-C8 alkylsulfinyl, C1-C8 alkylsulfonyl or C1-C8 alkylsulfonylamino. The phenyl groups can be condensed with a cycloalkyl or heterocyclyl group in which case tetrahydronaphthyl, indanyl, chromanyl, methylene-dioxyphenyl, ethylenedioxyphenyl and tetrahydro-quinolinyl groups are particularly preferred.

Heteroaryl for R1 is understood as five- and six-membered aromatics with 1-4 heteroatoms such as nitrogen, oxygen or sulphur which can be condensed with one or two phenyl groups and the carbon atoms thereof can, if desired, carry substituents such as halogen, nitro, nitrile, phenyl, trifluoromethyl, C1-C8 alkyl, Cl-C8 alkenyl, C1-C8 alkinyl, hydroxy, C1-C8 alkyloxy, C1-C8 alkenyloxy, C1-C8 alkinyloxy, amino, C1-C8 alkyl-amino, Cl-C8 alkenyl-amino, Cl-C8 alkinylamino, di-(Cl-C8 alkyl)amino, benzylamino, carboxyl, Cl-C8 alkyloxycarbonyl, amino-carbonyl, Cl-C8 alkylamino-carbonyl, di-(Cl-C8 alkyl)-aminocarbonyl, Cl-C8 alkyl-thio, Cl-C8 alkylsulfinyl, C1-C8 alkylsulfonyl or Cl-C8 alkylsulfonylamino. Preferred aromatics are furan, thiophene, pyrrole, oxazole, isoxazole, thiazole, isothiazole, imidazole, pyrazole, triazole, tetrazole, pyridine, pyrazine, pyrimidine, pyridazine, triazine, tetrazine, benzothiophene, dibenzothiophene, benzimidazole or carbazole.

The aforementioned Cl-C8 components can be straight-chained or branched. These are preferably understood to be methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert.butyl, pentyl, vinyl, allyl and propargyl residues.
Cycloalkyl groups for Rl are understood to be rings with

3-8 C atoms and preferably a cyclopentyl, cyclohexyl and cycloheptyl group.

In the general formula I, AS is understood to be glycine, azaglycine and the amino acids alanine, valine, leucine, isoleucine, methionine, proline, phenylalanine, tryptophan, serine, threonine, asparagine, aspartic acid, glutamine, glutamic acid, tyrosine, cysteine, lysine, arginine und histidine, which can be present in the D or L form or as mixtures of both forms.

If R2 und R3 together with the nitrogen atom to which they are bound form a heterocyclyl ring then this is preferably understood to be pyrrolidine, piperidine, homopiperidine, piparazine, morpholine and thiomorpholine. These rings can carry one or two C1-C8 alkyl, carboxyl or Cl-C8 alkyloxycarbonyl groups.

Rl denotes in particular a phenyl, naphthyl, tetrahydronaphthyl, pyridinyl, thienyl, cyclohexyl or chromanyl ring which can be substituted once or several times by Cl-C6 alkyl, Cl-C6 alkoxy or halogen groups.

AS denotes in particular glycine, azaglycine, alanine, glutamine, glutamate, asparagine or aspartate.

n denotes in particular the numbers O or 1.

R2 and R3 can be the same or different and denote in particular a Cl-C6 alkyl group such as for example the ethyl group; a Cl-C6-alkoxycarbonyl-C1-C6-alkyl group such as for example ethoxycarbonylmethyl; or a carboxy-Cl-C6-alkyl group such as for example carboxymethyl; or together with the N atom to which they are bound, they form a pyrrolidine, piperidine, homopiperidine, morpholine, thiomorpholine or piperazine ring which is, substituted if desired by one or two Cl-C6 alkyl groups such as for example methyl, ethyl, propyl or butyl;
carboxyl; or by Cl-C6 alkoxycarbonyl such as for example methoxycarbonyl, ethoxycarbonyl or tert. butyloxy-carbonyl.

R4 and R5 can be the same or different and in particular represent hydrogen atoms or C1-C6 alkyl groups preferably methyl groups.

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g R6, R7, R8, Rg can be the same or different and in particular represent hydrogen, fluorine or chlorine atoms.

Compounds of the general formula I are particularly preferred in which R1 denotes phenyl, 4-methylphenyl, 4-chloro-phenyl, 4-methoxyphenyl, 1-naphthyl, 2-naphthyl, 5,6,7,8-tetrahydro-2-naphthyl, 3-pyridinyl, 2-thienyl, cyclohexyl, 2,2,5,7,8-pentamethylchroman-6-yl or

4-methoxy-2,3,6-trimethylphenyl, AS denotes glycine, azaglycine or alanine, glutamine, glutamate, asparagine or aspartate, n can be O or 1, R2 and R3 are the same or different and denote ethyl, ethoxycarbonylmethyl or carboxymethyl or together with the N atom to which they are bound form a pyrrolidine, piperidine, homo-piperidine, morpholine, thiomorpholine or piperazine ring which, if desired, can carry one or two methyl, ethyl, propyl, butyl, carboxyl, methoxycarbonyl, ethoxycarbonyl or tert.butyloxycarbonyl groups, R4 and R5 are the same or different and denote hydrogen atoms or methyl groups, R6, R7, R8, Rg are the same or different and denote hydrogen, fluorine or chlorine atoms.

The physiologically tolerated salts of compounds of the general formula I are understood to be formates, acetates, caproates, oleates, lactates, or salts of carboxylic acids with up to 16 C atoms, hydrochlorides, hydrobromides, hydroiodides, alkanesulfonates with up to 10 C atoms, salts of dicarboxylic and tricarboxylic acids such as citrates, malonates und tartrates.

The compounds of the general formula I in which R1-R5, AS, n and m have the aforementioned meanings and R6-Rg denote halogen are prepared by reacting a compound of the general formula II, ,R2 Rl-S02-(AS)n - NH ~ R3 (CR4R5)m (II), in which R1-R5, AS, n and m have the aforementioned ~e~n;ngs with a compound of the general formula III, R~ ~ R6 (III), in which R6-Rg denote halogen.

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The compounds of the general formula I in which R1-R5, AS, n and m have the aforementioned meanings and R6-Rg denote hydrogen are prepared by subjecting a compound of the general formula I in which R1-R5, AS, n and m have the aforementioned meanings and R6-Rg denote halogen to a dehalogenation.

The compounds of the general formula II in which R1-R5, AS, n and m have the aforementioned meanings are prepared by reacting a compound of the general formula IV, Rl-SO2-(AS)n - NH ~ R3 (CR4R5)m (IV), ,.~ HNJ
Sch in which R1-R5, AS, n and m have the aforementioned meanings and Sch represents a protecting group such as a benzyloxycarbonyl, t-butoxycarbonyl or phthalimido group with a reagent that cleaves protecting groups.

The compounds of the general formula IV can be prepared by reacting a compound of the general formula V, 0~,0 HN ~ (V), (<R4R5)m NH
Sch in which m, n and Sch have the stated meanings, with an amine of the general formula VI, HN,R2 (VI), in which R2 and R3 have the stated meanings and the compounds of the general fomula VII that are formed ~ N (VII), H2N ~ ~R3 (CR4R5)m HN
Sch in which R2, R3, Sch and m have the stated meanings are reacted with a compound of the general formula VIII, ~15721~

Rl s-x (VIII), in which R1 has the stated meaning and X represents either a halogen atom or a residue of the general formula IX, HN A~o y (IX), in which Y denotes a halogen atom or an activated residue commonly used in peptide chemistry and A denotes a nitrogen atom or an atomic group of the general formula X, Rlo (X), ~ CH

in which Rlo represents one of the common amino acid side chains.

The compounds of the general formula V are prepared according to processes known from the literature.

The compounds of the general formula I can also be prepared by reacting a compound of the general formula XI, ~ N
H2N ~ R3 (CR4Rs)m HN (XI), R9~R6 R8l NJ~R7 in which R2-Rg and m have the stated meanings, with a compound of the general formula VIII.

The compounds of the general formula XI are produced by reacting a compound of the general formula XII, ~ N'R~
SchNH~ R3 (CR4Rs)m (XII), HN
R9~R6 in which R2-Rg have the stated meanings and Sch represents a common protecting group in peptide chemistry such as a benzyloxycarbonyl, t-butoxycarbonyl or phthalimido residue, with a reagent conventionally used in peptide chemistry to cleave protecting groups.

The compounds of the general formula XII are obtained by reacting a compound of the general formula XIII, ~ N
Sch NH~ R3 ( XI I I ), (CR4R5)m in which R2-R5, m and Sch have the stated meanings with a compound of the general formula III.

The compounds of the general formula XIII are prepared by subjecting the amide group of a compound of the general formula XIV, O R, ~ N ( XIV ), Sch NH~ ` R3 (CR4R5)m H2N~J

in which R2-R5, Sch and m have the stated meanings to a Hofmann degradation.

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The compounds of the general formula XIV are prepared by reacting a compound of the general formula XV, ~ OH
SchNH ~

(CR4R5)m ( xV), H2N ~

in which R4, R5, Sch and m have the stated meanings, with a compound of the general formula VI.

The compounds of the general formula XV are prepared from compounds of the general formula XVI, ~ OH
H2N ~
(CR4R5)m (XVI), H2N ~

in which R4 and R5 have the stated meanings by reaction with a conventional protecting group reagent in peptide chemistry.

The compounds of the general formula XVI are known from the literature.

The reactions of a compound of the general formula II
with a compound of the general formula III to form a compound of the general formula I are carried out in an inert solvent such as e.g. dimethylformamide, dioxane, dimethylsulfoxide or toluene at temperatures between O degrees Celsius and the boiling point of the solvent, and preferably at room temperature in the presence of an auxiliary base such as e.g. triethylamine, N-methyl-morpholine, pyridine or N-ethyldiisopropylamine.

The compounds of the general formula I, in which R6-Rg denote hydrogen, are obtained from the compounds of the general formula I in which R6-Rg denote halogen by catalytic hydrogenation in an inert solvent such as e.g.
methanol or ethanol in the presence of an acid-binding agent such as sodium methylate or sodium ethylate and preferably at room temperature and normal pressure with platinum or palladium as a catalyst.

The production of compounds of the general formula II by cleaving a protecting group from the compounds of the general formula IV is carried out according to methods common in peptide chemistry by acid reagents such as e.g. hydrogen bromide in glacial acetic acid, trifluoro-acetic acid or by means of hydrogenolysis or by cleavage with hydrazine.

The compounds of the general formula V are prepared according to methods known from the literature e.g. from an amino acid precursor with phosgene in an inert solvent such as dioxane.

The reaction of compounds of the general formula V to form compounds of the general formula IV is also carried out according to methods known from the literature in an inert solvent such as dimethylformamide at temperatures between -50 and +50 degrees Celsius.

The reaction of compounds of the general formula XI with compounds of the general formula VIII to form compounds of the general formula I is carried out in an inert solvent such as dimethylformamide, methylene chloride or dioxane at temperatures between O and 50 degrees Celsius and preferably at room temperatures in the presence of an auxiliary base such as e.g. triethylamine, N-methyl-morpholine or N-ethyldiisopropyl-amine.

Cleavage of the amino protecting group from a compound of the general formula XII to form a compound of the general formula XI is carried out hydrolytically e.g.
using a solution of hydrogen bromide in glacial acetic acid, trifluoroacetic acid or by means of hydrogenolysis or by reaction with hydrazine according to conventional methods known in peptide chemistry.

The reaction of a compound of the general formula XIII
with a compound of the general formula III to form a compound of the general formula XII is carried out in an inert solvent such as dimethylformamide, dioxane, dimethylsulfoxide or toluene at temperatures between o degrees Celsius and the boiling temperature of the solvent and preferably at room temperature in the presence of an auxiliary base such as e.g. triethyl-amine, N-methylmorpholine, pyridine or N-ethyldiiso-propylamine.

The conversion of compounds of the general formula XIV
into compounds of the general formula XIII is carried out by means of Hofmann degradation preferably using [bis(trifluoro-acetoxy)-iodo]benzene in a mixture of an inert solvent with water, preferably in an acetonitrile/water mixture and preferably at room temperature.

The production of compounds of the general formula XIV
from compounds of the general formula XV is carried out according to conventional methods in peptide chemistry.

The preparation of compounds of the general formula XV
from compounds of the general formula XVI is also carri-ed out according to conventional methods in peptide chemistry.

The compounds of the general formula XVI are known from the literature.

Examples of salts of compounds of formula I which can be used physiologically are salts with physiologically tolerated mineral acids such as hydrochloric acid, sulphuric acid, sulphurous acid or phosphoric acid; or with organic acids such as methanesulfonic acid, p-toluenesulfonic acid, acetic acid, trifluoroacetic acid, citric acid, fumaric acid, maleic acid, tartaric acid, succinic acid or salicylic acid. The compounds of formula I with a free carboxy group can also form salts with physiologically tolerated bases. Examples of such salts are alkaline metal, alkaline-earth metal, ammonium and alkylammonium salts such as a sodium, potassium, calcium or tetramethylammonium salt.

The compounds of formula I can be solvated and in particular hydrated. The hydration can be achieved in the course of the production process or gradually occur as a result of hygroscopic properties of a compound of formula I which is firstly anhydrous.

Pure enantiomers of compounds of formula I can either be obtained by racemate resolution (by formation of salts with optically active bases) or by using optically active starting materials in the synthesis.

For the production of pharmaceutical agents, the substances of the general formula I are mixed with suitable pharmaceutical carrier substances, aromatics, flavourings and dyes and are for example formed into tablets or coated tablets or are suspended or dissolved in water or oil e.g. olive oil with the addition of appropriate auxiliary substances.

The substances of the general formula I and their salts can be administered enterally or parenterally in a liquid or solid form. Water is preferably used as an injection medium which contains the usual additives in injection solutions such as stabilizers, solubilizers or buffers. Such additives are e.g. tartrate and citrate buffer, complexing agents (such as ethylenediaminetetra-acetic acid and their non-toxic salts) and high molecular polymers such as liquid polyethyloxide in order to regulate viscosity. Solid carrier materials are e.g. starch, lactose, mannitol, methylcellulose, talcum, highly dispersed silicic acids, high molecular fatty acids (such as stearic acid), animal and vegetable fats and solid high molecular polymers (such as polyethylene glycols). Preparations suitable for oral administration can, if desired, contain flavourings and sweeteners.

The compounds are usually administered in amounts of 10-1500 mg per day in relation to 75 kg body weight. It is preferable to administer 1-2 tablets with a content of active substance of 5-500 mg, 2-3 times per day. The 215721~

tablets can also be retarded as a result of which only 1-2 tablets have to be administered per day with 20-700 mg active substance. The active substance can also be administered by injection 1-8 times per day or by continuous infusion in which case 50-2000 mg per day are usually sufficient.

The following compounds are preferred within the sense of the invention in addition to those mentioned in the examples:

1. (R)-N-a-(2-naphthylsulfonylglycyl)-N-y-(4-pyridinyl)-a,y-diaminobutyric acid-homopiperidide 2. (R)-N-a-(2-naphthylsulfonylglycyl)-N-y-(4-pyridinyl)-a,y-diaminobutyric acid-4-methylpiperazide 3. (R)-N-a-(2-naphthylsulfonylglycyl)-N-y-(4-pyridinyl)-a,y-diaminobutyric acid-thiomorpholide 4. (R)-N-a-(2-naphthylsulfonylglycyl)-N--(4-pyridinyl)-a,y-diaminobutyric acid-(N-ethyl-N-ethoxycarbonyl-methyl)amide

5. (R)-N-a-(2-naphthylsulfonylglycyl)-N-y-(4-pyridinyl)-a,y-diaminobutyric acid-(N-ethyl-N-carboxymethyl)amide

6. (R)-N-a-(2-naphthylsulfonylazaglycycl)-N-y-(4-pyridinyl)-a,y-diaminobutyric acid-homopiperidide

7. (R)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-glycyl)-N-y-(4-pyridinyl)-a,y-diaminobutyric acid-homopiperidide

8. (R)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-azaglycyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-piperidide

9. (R)-N-a-(2,2,5,7,8-pentamethyl-6-chromanylsulfonyl-glycyl-N-~-(4-pyridinyl)-a,y-diaminobutyric acid-homopiperidide

10. (R)-N-a-(2,2,5,7,8-Pentamethyl-6-chromanylsulfonyl-aza-glycyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-piperidide

11. (R)-N-a-(5,6,7,8-tetrahydronaphthylsulfonylglycyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-homo-piperidide

12. (R)-N-a-(5,6,7,8-tetrahydronaphthylsulfonyl-azaglycyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-piperidide

13. (R)-N-a-(2-naphthylsulfonyl-(S)-asparagyl)-N-y-(4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

14. (R)-N-a-(5,6,7,8-tetrahydronaphthylsulfonyl-(S)-asparagyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-piperidide

15. (R)-N-a-(2,2,5,7,8-pentamethyl-6-chromanylsulfonyl-(S)-asparagyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-piperidide

16. (R)-N-a-( 4-methoxy-2,3,6-trimethylphenylsulfonyl-(S)-asparagyl)-N-~-( 4-pyridinyl)-a,~-diaminobutyric acid-piperidide

17. (R)-N-a-(S, 6, 7, 8-tetrahydronaphthylsulfonyl-(S)-asparagyl)-N-~-( 4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

18. (R)-N-a-( 2,2,5,7,8-pentamethyl-6-chromanylsulfonyl-(S)-asparagyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

19. (R) -N-a- (4-methoxy-2,3,6-trimethylphenylsulfonyl-(S)-asparagyl)-N-~-( 4-pyridinyl)-a, y-diaminobutyric acid-homopiperidide

20. (R) -N-a-( 2-naphthylsulfonyl-(S)-glutamyl)-N-~-(4-pyridinyl)-a, y-diaminobutyric acid-piperidide

21. (R) -N-a-( 2-naphthylsulfonyl-(S)-glutamyl)-N-~-( 4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

22. (R) -N-a-( 5,6,7,8-tetrahydronaphthylsulfonyl-(S)-glutamyl)-N-~-( 4-pyridinyl)-a, y-diaminobutyric acid-piperidide

23. (R) -N-a-( 2,2,5,7,8-pentamethyl-6-chromanylsulfonyl-(S)-glutamyl)-N-~-( 4-pyridinyl)-a,~-diaminobutyric acid-piperidide

24. (R) -N-a- (4-methoxy-2,3,6-trimethylphenylsulfonyl-(S)-glutamyl)-N-~-( 4-pyridinyl)-a,~-diaminobutyric acid-piperidide

25. (R)-N-a-(5,6,7,8-tetrahydronaphthylsulfonyl-(S)-glutamyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

26. (R)-N-a-(2,2,5,7,8-pentamethyl-6-chromanylsulfonyl-(S)-glutamyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

27. (R)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-(S)-glutamyl)-N-~-(4-pyridinyl)-a,~-diaminobutyric acid-homopiperidide

28. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(4-pyridinyl)- a,~-diaminopropionic acid-homopiperidide

29. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(4-pyridinyl)- a,~-diaminopropionic acid-4-methyl-piperidide

30. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(4-pyridinyl)- a,~-diaminopropionic acid-2-carboxy-piperidide

31. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(4-pyridinyl)- a,~-diaminopropionic acid-3-carboxy-piperidide

32. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(4-pyridinyl)- a,~-diaminopropionic acid-4-carboxy-piperidide

33. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(4-pyridinyl)- a,~-diaminopropionic acid-2-carboxy-pyrrolidide

34. (R,S)-N-a-(5,6,7,8-tetrahydronaphthylsulfonylglycyl)-N-~-(4-pyridinyl)-a, ~-diaminopropionic acid-piperidide

35. (R,S)-N-a-(5,6,7,8-tetrahydronaphthylsulfonylglycyl)-N-~-(4-pyridinyl) -a, ~-diaminopropionic acid-4-methyl-piperidide

36. (R,S)-N-a-(5,6,7,8-tetrahydronaphthylsulfonylglycyl)-N-~-(4-pyridinyl)-a, ~-diaminopropionic acid-2-carboxy-piperidide

37. (R)-N-a-(5,6,7,8-tetrahydronaphthylsulfonylglycyl)-N-~-(4-pyridinyl)-a,~-diaminopropionic acid-2-carboxypyrrolidide

38. (R,S)-N-a-(2,2,5,7,8-pentamethyl-6-chromanyl-sulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diamino-propionic acid-piperidide

39. (R,S)-N-a- (2, 2, 5,7,8-pentamethyl-6-chromanyl-sulfonyl-glycyl)-N-~-(4-pyridinyl) -a, ~-diamino-propionic acid-4-methylpiperidide

40. (R,S)-N-a-( 2, 2, 5,7,8-pentamethyl-6-chromanyl-sulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diamino-propionic acid-2-carboxypiperidide 215721~

41. (R,S)-N-a-(2,2,5,7,8-pentamethyl-6-chromanyl-sulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diamino-propionic acid-2-carboxypyrrolidide

42. (R,S)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diaminopropionic acid-piperidide

43. (R,S)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diaminopropionic acid-4-methylpiperidide

44. (R,S)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diaminopropionic acid-2-carboxypiperidide

45. (R,S)-N-a-(4-methoxy-2,3,6-trimethylphenylsulfonyl-glycyl)-N-~-(4-pyridinyl)-a,~-diaminopropionic acid-2-carboxypyrrolidide BeiSDi~l 1 (R,S)-N-a-(2-naphthylsulfonylq,lycyl)-N-~-(2,3,5,6-tetra-chloropyridin-4-yl)-a,r-diaminobutYric acid-pi~eridide 1. (R.S)-N-a-(2-na~hthylsulfonylqlycyl~-y-~hthalimido-a-aminobutyric acid-pi~eridide A solution of 4.6 g (R,S)-4-B-phthalimido-ethyloxazolidine-2,5-dione in 22 ml absolute dimethylformamide is cooled to -50 degrees Celsius.
A solution of 1.7 ml piperidine and 2 ml N-methyl-morpholine in 22 ml absolute dimethyl-formamide is ~_ 21~721~

added dropwise to this during 15 minutes while stirring. It is stirred for a further 30 minutes and then the reaction mixture is heated for 30 minutes to 60 degrees Celsius. It is cooled to room temperature and a solution of 4.5 g 2-naphthyl-sulfonylglycyl chloride in 20 ml absolute methylene chloride is added dropwise while stirring further.
It is stirred for a further 2 hours at room temperature and then evaporated in a vacuum. The residue is dissolved in methylene chloride and the solution is washed with water. The methylene chloride phase is dried over sodium sulfate and concentrated by evaporation. The residue is chromatographed on a silica gel column (mobile solvent: acetone/toluene 1:1) for purification.
After concentrating the appropriate column fractions by evaporation, 4.5 g of the title compound is obtained as an amorphous substance.

2. (R,S)-N-a-(2-na~hthYlsulfonYlglYcyl)-a.y-diaminobutYric acid-pi~eridide 4.5 g (R,S)-N-a-(2-naphthylsulfonylglycyl)-~-phthalimido-a-aminobutyric acid-piperidide is dissolved in 30 ethanol. 4 ml 2M ethanolic hydrazine hydrate solution is added and the mixture is stirred overnight at room temperature. The reaction mixture is acidified with 10 ml 2 N
hydrochloric acid, heated for a short time and the precipitate is removed by filtration. The filtrate is concentrated by evaporation and the residue is dissolved in water. A dilute soda solution is added to the aqueous solution and it is extracted with methylene chloride. The methylene chloride solution is dried over sodium sulfate and concentrated by _.

evaporation. 2.5 g of the title compound is obtained as a yellow, amorphous residue.

3. (R,S)-N-a-(2-naPhthylsulfonylqlycyl)-N-~-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminobutyric acid-piperidide 2 g (R,S)-N-a-(2-naphthylsulfonylglycyl)-a,~-diaminobutyric acid-piperidide is dissolved in 20 ml absolute dioxane. A solution of 1.4 g 4-nitro-2,3,5,6-tetrachloropyridine and 0.6 ml triethylamine in lO ml absolute dioxane is added to this while cooling on ice and while stirring. It is stirred for a further 3 hours at room temperature and then the reaction mixture is evaporated in a vacuum. The residue is chromatographed on a silica gel column (mobile solvent: isohexane/ethyl acetate 1:2) for purification. After evaporating the column fractions, 1.1 g crystals is obtained with a melting point of 205 degrees Celsius. FAB-MS: M+H
648.

kxampl~ 2 (R,S)-N-a-(2-naphthylsulfonylqlycyl)-N-~-(pYridin-4-yl)-a, ~-diaminobutyric acid-piDeridide 1 g (R,s)-N-a-(2-naphthylsulfonylglycyl-N-~-~2,3,s,6-tetrachloropyridin-4-yl)-a, ~-diaminobutyric acid-piperidide is dissolved in 20 ml methanol and after addition of 5 ml 1 M sodium methylate solution it is hydrogenated in the presence of lO0 mg Pd/C (10 %) catalyst. When the calculated amount of hydrogen has been taken up, it is filtered from the catalyst and 215721~
_ concentrated by evaporation. The residue is chromatographed on a silica gel column (mobile solvent:
methylene chloride/methanol 8:2). The column fractions are evaporated and the residue is triturated with ether.
350 mg of the title compound is obtained. Melting point:
135 degrees Celsius. FAB-MS: M+H 510.

Exam~l~ 3 (R,S)-N-a-(4-toluene-sulfonYlglycyl~-N-~-(pyridin-4-yl)-a,~-diaminobutyric acid-piperidide The preparation was analogous to examples 1 and 2 with the only exception that in step 1 p-toluenesulfonyl-glycyl chloride was used instead of 2-naphthylsulfonyl-glycyl chloride. Melting point of the title compound:
160 degrees Celsius (decomp.). FAB-MS: M+H 474.

Exampl~ 4 (R,S)-N-a-(2-naphthylsulfonyl)-N-~-(pyridin-4-yl)-a,~-diaminobutyric acid-pi~eridide The preparation was analogous to Examples 1 and 2 with the only exception that in step 1 2-naphthylsulfonyl chloride was used instead of 2-naphthylsulfonylglycyl chloride. The title compound was obtained as an amorphous substance. FAB-MS: M+H 453.

Ex~mDle 5 (R,S)-N-a-(2-na~hthylsulfonylqlycvl)-N-~-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminovaleric acid-piperidide 1. (R,S)-a-amino-~-phthalimido-valeric acid 29.6 g phthalic anhydride is heated together with 23.4 g 5-aminovaleric acid for 30 minutes to 190 degrees Celsius. The residue of ~-phthalimido-valeric acid obtained after cooling is recrystallized from aqueous ethanol. Melting point:
117 degrees Celsius. 47.3 g of this compound is mixed with 2 g red phosphorus. 20.7 g bromine is slowly added dropwise to this mixture while stirring. After completion of the bromine addition it is heated for 2 hours to 100 degrees Celsius and then allowed to cool. The residue is admixed with 200 ml ice water and the mixture is stirred for a further hour. Methylene chloride is added to the mixture, the water phase is separated and discarded. The methylene chloride phase is dried over sodium sulfate and concentrated by evaporation. The crude a-bromo-~-phthalimido-valeric acid obtained (53 g) is dissolved in 300 ml dimethylformamide without further purification.
20.8 g sodium azide is added to this solution and the mixture is stirred for 24 hours at room temperature. Then the solution is evaporated in a vacuum, the residue is dissolved in ethyl acetate, the solution is washed with water, dried over sodium sulfate and evaporated. The a-azido-~-phthalimido-valeric acid obtained is dissolved in a mixture of 240 ml glacial acetic acid and 30 ml concentrated hydrochloric acid and hydrogenated after addition of 2 g platinum oxide as a catalyst.
After the uptake of hydrogen is complete, it is filtered from the catalyst by suction and the filtrate is evaporated. The residue is dissolved in 50 ml water and the solution is admixed with 12 ml pyridine. The precipitate is filtered by suction, washed with water and dried. 25 g (R,S)-a-amino-~-phthalimido-valeric acid is obtained. Melting point: 230 degrees Celsius.

2. (R,S)-4-(~-phthalimidopropyl)-oxazolidine-2,5-dione 12 g (R,S)-a-amino-~-phthalimido-valeric acid is suspended in 100 ml absolute dioxane. Phosgene is passed into this suspension while stirring at 70 degrees Celsius until a clear solution is formed.
It is allowed to cool and the precipitate which forms is filtered by suction. Yield: 9.5 g; melting point: 235 degree Celsius (decomp.).

3. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-phthalimido-a-aminovaleric acid-piperidide The preparation was carried out analogously to step 1 in example 1 with the only exception that (R,S)-4-(~-phthalimido-propyl)-oxazolidine-2,5-dione was used instead of (R,S)-4-(~-phthalimidoethyl)-oxazolidine-2,5-dione. The title compound was obtained as a white amorphous solid.

4. (R,S)-N-a-(2-naphthylsulfonylglycyl)-a,~-diaminovaleric acid-piperidide -The preparation was carried out analogously to step 2 in example 1 using (R,S)-N-a-(2-naphthylsulfonyl-glycyl)-~-phthalimido-a-aminovaleric acid-piperidide as the starting material. The titlecompound was obtained as an amorphous solid.

5. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminovaleric acid-piperidide The preparation was carried out analogously to step 3 in example 1 using (R,S)-N-a-(2-naphthylsulfonyl-glycyl)-a,~-diaminovaleric acid-piperidide as the starting material. Melting point of the title compound: 187 degrees Celsius (from methanol). FAB-MS: M+H 662.

kxampl~ 6 (R.S)-N-a-(2-naphthylsulfonylglYcYl)-N-~-(pyridin-4-yl)-a,~-diaminovaleric acid-pi~eridide The preparation was carried out analogously to example 2 using (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-~-(2,3,5,6-tetrachloro-pyridin-4-yl)-a,~-diaminovaleric acid as the starting material. The title compound was obtained as a white amorphous solid. FAB-MS: M+H 524 ExamPle 7 (R.S)-N-a-(2-naphthylsulfonylglycyl)-N-B-(2.3 5 6-tetrachloropyridin-4-Yl)-a.B-diaminoproionic acid-~i~eridide 1. (R,S)-N-a-N-B-dibenzyloxycarbonyl-a,B-diamino-propionic acid 17.5 g (R,S)-a,B-diaminopropionic acid is suspended in 170 ml water. The pH value is adjusted to 9 by addition of dilute sodium hydroxide solution during which the substance dissolves. A solution of 53.5 ml chloroformic acid benzyl ester in 300 ml toluene is added dropwise while stirring. The pH
value is kept at 9 by simultaneous addition of dilute sodium hydroxide solution. After the addition of chloroformic acid benzyl ester is completed, it is stirred for a further 4 hours at room temperature and then the solution is acidified to pH 1 with dilute hydrochloric acid. The precipitate is filtered by suction, washed with water and dried. Yield: 38.3 g; melting point: 122 degrees Celsius.

2. (R,S)-4-(benzyloxycarbonylaminomethyl)-oxazolidine-2,5-dione 25 g (R,S)-N-a-N-B-dibenzyloxycarbonyl-a,B-diamino-propionic acid is dissolved in 250 ml absolute chloroform. 20 ml thionyl chloride is added to this and the mixture is heated for 1 hour to 50 degrees Celsius. It is then evaporated to dryness, the residue is taken up in 100 ml ethyl acetate and it is heated for 1 hour uder reflux. It is evaporated to half its volume, 250 ml absolute hexane is added and it is allowed to crystallize. The crystals are filtered by suction, washed with absolute hexane and dried. Yield: 17.5 g; melting point: 128 degrees Celsius.

3. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-B-benzyloxy-carbonyl-a,B-diaminopropionic acid-piperidide 1.2 g (R,S)-4-benzyloxycarbonylaminomethyl-oxazolidine-2,5-dione is dissolved in 10 ml absolute dimethylformamide and cooled to -50 degrees Celsius. A solution of 0.45 ml piperidine and 0.5 ml N-methylpiperidine in 10 ml absolute methylene chloride is added to this while stirring.
It is allowed to reach room temperature and stirred for a further 30 minutes and then the reaction mixture is heated for 30 minutes to 60 degrees Celsius. It is cooled to room temperature and a solution of 1.31 g 2-naphthylsulfonylglycyl chloride in 10 ml methylene chloride is added dropwise while stirring. It is stirred for a further 4 hours at room temperature, the solution is diluted with 100 ml methylene chloride and washed with water. The methylene chloride phase is dried over sodium sulfate and evaporated. The residue is chromatographed on a silica gel column (mobile solvent: ethyl acetate/isohexane 1.5:1) for purification. After evaporation of the appropriate column fractions, 1.45 g of a white solid is obtained. Melting point: 148 degrees Celsius.

4. (R,S)-N-a-(2-naphthylsulfonylglycyl)-a,B-diaminopropionic acid-piperidide hydrobromide 1.1 g (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-B-benzyloxycarbonyl-a,B-diaminopropionic acid is dissolved in 4 ml of a 33 % solution of hydrogen bromide in glacial acetic acid. It is stirred for a further 1 hour and then evaporated in a vacuum. The - 215721~

residue is triturated with ether, filtered by suction and dried. 0.9 g of the title compound is obtained as a white amorphous solid.

5. (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-B-(2,3,5,6-tetrachloropyridin-4-yl)-a,B-diaminopropionic acid-piperidide 0.6 g (R,S)-N-a-(2-naphthylsulfonylglycyl)-a,B-diaminopropionic acid-piperidide hydrobromide is dissolved in 5 ml absolute dioxane. 0.4 g 4-nitro-2,3,5,6-tetrachloropyridine and 0.2 ml triethylamine is added to this and the mixture is stirred overnight at room temperature. It is then evaporated and the residue is chromatographed on a silica gel column (mobile solvent: methylene chloride/methanol 97.5:2.5). The residue obtained after evaporation of the column fractions is recrystallized from methanaol. Yield: 350 mg, melting point: 189 degrees Celsius. FAB-MS: M+H
634.

Exampl~ 8 (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-B-~yridin-4-yl)-a,B-diaminopropionic acid-~iperidide 300 mg (R,S)-N-a-(2-naphthylsulfonylglycyl)-N-B-2,3,s,6-chloropyridin-4-yl is dissolved in 15 ml methanol and hydrogenated after addition of 7 ml 1 M sodium methylate solution and 200 mg Pd/C (10 %) catalyst. After the calculated amount of hydrogen has been taken up, it is filtered from the catalyst and the filtrate is evaporated. The residue is chromatographed on a silica -- 36 _ 215721~

gel column (mobile solvent: methylene chloride/methanol 8:2). The column fractions are evaporated and the residue is triturated with ether. 200 mg of the title compound is obtained as colourless crystals. Melting point: 95 degrees Celsius. FAB-MS: M+H 496.

Example 9 (R)-N-a-(2-na~hthylsulfonylqlycyl~-N-y-(2.3.5,6-tetra-chloropYridin-4-yl~-a,y-diaminobutyric acid-pi~eridide 1. (R)-Z-glutamine-piperidide 6 g (R)-Z-glutamine is dissolved in 60 ml absolute dioxane. 2.S g N-hydroxysuccinimide and 5 g dicyclohexyldi-carbodiimide is added to this and the mixture is subsequently stirred for 20 hours at room temperature. The precipitate which formed is removed by filtration and discarded. The filtrate is admixed with 2.1 ml piperidine and the mixture is stirred for a further 24 hours at room temperature. It is separated from turbidity which may have formed by suction filtration and the filtrate is concentrated by evaporation. The residue is triturated with ethyl acetate and the crystals are filtered by suction. 3.7 g (R)-Z-glutamine-piperidide is obtained. Melting point: 95 degrees Celsius.

2. (R)-N-a-benzyloxycarbonyl-a,y-diaminobutyric acid-piperidide 3.4 g (R)-Z-glutamine-piperidide is dissolved in a mixture of 30 ml acetonitrile and 30 ml water.
6.5 g tbis(trifluoroacetoxy)iodo]-benzene is added to this and the mixture is stirred overnight at room temperature. It is diluted with 200 ml water, acidified to pH 1 with dilute hydrochloric acid and shaken out with ether. The aqueous phase is made alkaline with 10 N sodium hydoxide solution while cooling and extracted with ethyl acetate. The ethyl acetate phase is dried over sodium sulfate and evaporated. 2.1 g of the title compound is obtained as an amorphous substance.

3. (R)-N-a-(benzyloxycarbonyl-N-y-(2,3,5,6-tetrachloropyridin-4-yl)-a,r-diaminobutyric acid-piperidide 1.9 g (R)-N-a-benzyloxycarbonyl-a,r-diaminobutyric acid-piperidide is dissolved in 20 ml absolute dioxane. 1.7 g 4-nitro-2,3,5,6-tetrachloropyridine is added to this and the mixture is stirred for 3 hours at room temperature. Subsequently the reaction mixture is diluted with 300 ml water and extracted with ethyl acetate. The ethyl acetate phase is dried over sodium sulfate and evaporated.
The residue is chromatographed on a silica gel column (mobile solvent: ethyl acetate:isohexane 3:1) for purification. After evaporation of the column fractions, 1.9 g of the title compound is obtained as an oily substance.

4. (R)-N-y-(2,3,5,6-tetrachloropyridin-4-yl)-a, r-diaminobutyric acid-piperidide hydrobromide 950 mg (R)-N-a-benzyloxycarbonyl-N-y-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminobutyric acid-piperidide is dissolved in 2 ml of a 33 % solution of hydrogen bromide in glacial acetic acid. The mixture is stirred overnight at room temperature and then evaporated in a vacuum. The residue is triturated with ether. 630 mg of the title compound is obtained with a melting point of 180 degrees Celsius.

5. (R)-N-a-(2-naphthylsulfonylglycyl)-N-~-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminobutyric acid-piperidide 325 mg (R)-N-y-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminobutyric acid-piperidide hydrobromide is dissolved in 4 ml absolute methylene chloride.
0.2 ml N-methylmorpholine and 200 mg 2-napthyl-sulfonylglycyl chloride is added to this. The mixture is stirred for 2 hours at room temperature and then evaporated. The residue is chromatographed on a silica gel column (mobile solvent: ethyl acetate/isohexane 2:1). After evaporation of the column fractions, 220 mg of the title compound is obtained with a melting point of 130 degrees Celsius. FAB-MS: M+H 648.

Exampl~ 10 (R)-N-a-(2-naphthylsulfonylqlycyl)-N-~-(pyridin-4-Yl)-a.~-diaminobutyric acid-piperidide 140 mg (R)-N-a-(2-naphthylsulfonylglycyl)-N-~-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminobutyric acid-piperidide is dissolved in 5 ml methanol and hydrogenated after addition of 2 ml 1 M sodium methylate solution and 100 mg Pd/C (10 %) catalyst. After the calculated amount of hydrogen has been taken up, it is filtered from the catalyst and evaporated. The residue is chromatographed on a silica gel column (mobile solvent: methylene chloride/methanol 8:2). The residue obtained after evaporation of the column fractions is triturated with ether and filtered by suction. 50 mg of the title compound is obtained with a melting point of 148 degrees Celsius. FAB-MS: M+H 510 [a]D = +2.40 [ethanol].

Ex~mple 11 (S)-N-a-(2-naPhthylsulfonylqlycyl~-N-y-(pYridin-4-yll-a.y-diaminobutyric acid-Piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that (L)-Z-glutamine was used instead of (R)-Z-glutamine as the starting material. Melting point: 148 degrees Celsius. FAB-MS: M+H 510.[a]D = -2.40 [ethanol].

Ex~mDle 12 (R)-N-a-(phenylsulfonylglYcyl)-N-~-(Pyridin-4-yl)-a.y-diaminobutyric acid-piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that phenylsulfonylglycyl chloride was used in step 5 instead of 2-naphthylsulfonylglycyl chloride. Melting point: 130 degrees Celsius. FAB-MS:
M+H 460.

215721~
_ Ex~mPle 13 (R)-N-a-(4-methoxY-phenYlsulfonylglycyl)-N-r-(pYridin-4-yl)-a.~-diaminobutyric acid-Piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 4-methoxyphenylsulfonylglycyl chloride was used in step 5 instead of 2-naphthyl-sulfonylglycyl chloride. Melting point: 140 degrees Celsius. FAB-MS: M+H 490.

Examle 14 (R)-N-a-(4-trifluoromethylphenylsulfonylglYcYl)-N-r-(pYridin-4-Yl)-a.r-diaminobutYric acid-piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 4-trifluoromethylphenylsulfonyl-glycyl chloride was used in step 5 instead of 2-naphthylsulfonylglycyl chloride. Melting point: 180 degrees Celsius. FAB-MS: M+H 528. ~a]D = +10.7.

Ex~mpl~ 15 (R)-N-a-(5,6.7,8-tetrahYdronaphthalino-2-sulfonyl-glYcyl~-N-r-(pYridin-4-yl)-a,~-diaminobutyric acid-piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 5,6,7,8-tetrahydronaphthalino-2-21~7215 sulfonylglycyl chloride was used in step 5 instead of 2-naphthylsulfonylglycyl chloride. Melting point: 120 degrees Celsius. FAB-MS: M+H 514.

Example 16 (R)-N-a-(2-N-naPhthylsulfonyl-(R)-alanyl)-N-~-(pyridin-4-yl)-a,~-diaminobutYric acid-piPeridide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 2-naphthylsulfonyl-(R)-alanyl chloride was used in step 5 instead of 2-naphthyl-sulfonylglycyl chloride. Melting point: 195 degrees Celsius. FAB-MS: M+H 524. [a]D = + 77.6.

Ex~mple 17 (R)-N-a-(2-naPhthylsulfonyl)-(S)-alanyl-N-~-(pyridin-4-Yl)-a,~-diaminobutyric acid-piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 2-naphthylsulfonyl-(S)-alanyl chloride was used in step 5 instead of 2-naphthylsulfonylglycyl chloride. Melting point:
amorphous substance. [a]D = -44.5 [ethanol]. FAB-MS:
M+H 524.

Ex~mpl~ 18 (R)-N-a-(4-nitrophenylsulfonyl)-glYcyl-N-r-(pyridin-4-yl)-a,~-diaminobutYric acid-pi~eridide 1. (R)-N-~-(pyridin-4-yl)-a,y-diaminobutyric acid-piperidide 960 mg (R)-N-y-(2,3,5,6-tetrachloropyridin-4-yl)-a,~-diaminobutyric acid-piperidide hydrobromide is dissolved in 30 ml methanol and hydrogenated after addition of 25 ml 1 M sodium methylate solution and 200 mg Pd/C (10 %). After the calculated amount of hydrogen has been taken up, it is filtered from the catalyst and the filtrate is evaporated. The residue is dissolved in water, the solution is adjusted to pH 10 by addition of dilute sodium hydroxide solution and extracted with methylene chloride. The methylene chloride phase is dried over sodium sulfate and evaporated. 480 mg of the title compound is obtained as an amorphous residue.

2. (R)-N-a-(4-nitrophenylsulfonylglycyl)-N-~-(pyridin-4-yl)-a,y-diaminobutyric acid-piperidide 220 mg N-~-(pyridin-4-yl)-a,y-diaminobutyric acid-piperidide is dissolved in 5 ml absolute dimethylformamide. o. 25 ml N-methylmorpholine and 250 mg 4-nitrophenylsulfonylglycyl chloride is added to this. The mixture is stirred for 2 hours at room temperature and then evaporated in a vacuum. The residue is chromatographed on a silica gel column (mobile solvent: methylene chloride/methanol 8:2) for purification. After evaporation of the column fractions, 180 mg of the title compound is obtained as an amorphous substance. FAB-MS: M+H 505.

Ex~mple 19 (R)-N-a-(3-nitrophenylsulfonylglycvl)-N-~-(pyridin-4-yl)-a.~-diaminobutyric acid-piPeridide The title compound was prepared analogously to step 2 in example 18 with the only exception that 3-nitrophenyl-sulfonylglycyl chloride was used instead of 4-nitro-phenylsulfonylglycyl chloride. Amorphous substance. FAB-MS: 505.

Ex~mple 20 (R)-N-a-(4-chloroPhenylsulfonylglycYl)-N-y-(pyridin-4-yl)-a.~-diaminobutyric acid-PiPeridide The title compound was prepared analogously to step 2 in example 18 with the only exception that 4-chlorophenyl-sulfonylglycyl chloride was used instead of 4-nitro-phenylsulfonylglycyl chloride. Amorphous substance. FAB-MS: M+H 495.

~x~mple 21 (R)-N-a-(cYclohexYlsulfonylglycyl)-N-~-(pyridin-4-yl)-a,~-diaminobutYric acid-piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that cyclohexylsulfonyl chloride was used in step 5 instead of 2-naphthylsulfonylglycyl chloride. Melting point: 110 degrees Celsius. FAB-MS:
M+H 466.

Example 22 (R)-N-a-(2-na~hthYlsulfonylqlycyl)-N-~ Yridin-4-yl)-a,~-diaminobutyric acid-(4-methyl-~i~eridide) The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that homopiperidine was used instead of piperidine. Melting point: 150 degrees Celsius. FAB-MS: M+H 524.

Ex~mple 23 (R)-N-a-(2.2,5,7,8-pentamethyl-6-chromanylsulfonyl-glycYl)-N-y-(pyridin-4-yl)-a,Y-diaminobutyric acid-piperidide) The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 2,2,5,7,8-pentamethyl-6-chromanyl-sulfonylglycyl chloride was used instead of 2-naphthyl-sulfonylglycyl chloride. FAB-MS: M+H 529.

-Example 24 (R)-N-a-(4-methoxY-2,3,6-trimethyl~henYlsulfonylglycyl)-N-~-(pyridin-4-yl)-a,~-diaminobutYric acid-piPeridide) The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 4-methoxy-2,3,6-trimethyl-sulfonylglycyl chloride was used instead of 2-naphthylsulfonylglycyl chloride. FAB-MS: M+H 532.

Ex~mple 25 (R)-N-a-(2-naphthYlsulfonyl-(S)-methyl-asparagyl)-N-y-(~yridin-4-Yl)-a.~-diaminobutyric acid-piperidide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that 2-naphthylsulfonyl-(S)-methyl-asparaginyl chloride was used instead of 2-naphthyl-sulfonylglycyl chloride. FAB-MS: M+H 582.

~mple 26 (R)-N-a-(2-naphthylsulfonyl-(S)-asparagyl)-N-r-(pyridin-4-yl)-a,~-diaminobutyric acid-piperidide The title compound was prepared by alkaline hydrolysis of example 25. FAB-MS: M+H 568.

- 46 -~x~mpl~ 27 (R)-N-a-(2-naphthylsulfonylglycyl)-N-r-(pYridin-4-yl)-a.r-diaminobutYric acid-morpholide The title compound was prepared analogously to the reaction sequence described in examples 9 and 10 with the only exception that morpholine was used instead of piperidine. FAB-MS: M+H 512.

Ex~mpl~ 28 (R)-N-a-(2-naphthYlsulfonylazaqlycyl)-N-~-(pyridin-4-Yl)-a,y-diaminobutyric acid-~iperidide 1.5 g (R)-N-y-(2,3,5,6-tetrachloropyridin-4-yl)-a,y-diaminobutyric acid-piperidide is dissolved in 15 ml absolute dimethylformamide. 1 g N-tert.butoxycarbonyl-N-(4-nitrophenoxycarbonyl)-hydrazine and 0.2 ml diisopropylamine is added to this. The mixture is stirred for 1 hour at room temperature, 200 ml water is added and the precipitate is filtered by suction. The precipitate is chromatographed on a silica gel column (mobile solvent: ethyl acetate/isohexane 2:1) for purification. 1.4 g crystals are obtained with a melting point of 120 Celsius. These are dissolved in 30 ml of a 1.2 molar solution of hydrobromic acid in glacial acetic acid and the solution is stirred for 2 hours at room temperature. It is evaporated in a vacuum, the residue is triturated with ether and filtered by suction. The filter residue is dissolved in 10 ml pyridine without further purification. 1.2 g 2-naphthalinesulfonic acid chloride is added to this and the mixture is stirred for 4 hours at room temperature. It is diluted with 50 ml

- 47 -water and filtered by suction. The residue is chromatographed on a silica gel column (mobile solvent:
ethyl acetate/isohexane 2:1) for purification. The oily residue obtained (1.1 g) is dissolved in 50 ml methanol and hydrogenated after addition of 20 ml of a 0.4 molar sodium methylate solution and 200 mg Pd/C (10 %) catalyst. After the uptake of hydrogen has been completed, it is separated from the catalyst by suction filtration and evaporated. The residue is taken up in water and extracted with ethyl acetate. The ethyl acetate phase is dried over sodium sulfate and evaporated. The residue is chromatographed on a silica gel column (mobile solvent: ethyl acetate). 400 mg of the title compound is obtained as an amorphous substance. FAB-MS: M+H 511.

Ex~mple 29 DescriDtion of pharmaGological experiments Thrombin time A conventional test in clinical coagulation diagnostics is the thrombin time. This parameter measures the action of thrombin on fibrinogen and the formation of clots.
Inhibitors of thrombin result in an extended thrombin time.

In order to obtain plasma 9 parts of fresh blood from healthy donors was mixed with one part of sodium citrate solution (0.11 mol/l) and it was centrifuged for 10 minutes at room temperature at ca. 3000 r.p.m.. The plasma was pipetted off and can be stored at room temperature for ca. 8 hours.

~ 215721S

- 48 -200 ~1 citrate plasma was incubated for 2 minutes at 37C in a ball coagulometer (KC10 from the Amelung Company). 10 ~1 dimethylsulfoxide (DMSO) or a solution of the active substance in DMSO was added to 190 ~1 pre-heated thrombin reagent (Boehringer Mannheim GmbH;
contains ca. 3 U/ml horse thrombin and 0.0125 M Ca++).
On addition of 200 ~1 of this solution to the plasma a stopwatch was started and the time at which coagulation starts was determined. The thrombin time was ca. 24 sec.
in control measurements and was substantially increased by the active substances.

The measured thrombin times in seconds are given in the following table as a difference to the control. The concentrations of the active substances in the final volume were 250 ~M (TT250), 25 ~M (TT25) and 2.5 ~M
(TT2.5).

Thrombin inhibition The kinetic measurements were carried out in 0.1 M
phosphate buffer that contained 0.2 M sodium chloride and 0.5 % polyethylene glycol 6000 at a pH = 7.5 and 25C with the substrate H-(D)-Phe-Pro-Arg-pNA; Kabi and human a thrombin (Sigma, specific activity = 2150 NIH-units/mg) in polystyrene semi-microcuvettes in a total volume of 1 ml.

In a preliminary test each active substance was determined as to whether it inhibits thrombin rapidly or slowly. For this the reaction was firstly started by adding 0.03 NIH units thrombin to a 100 ~M solution of the substrate and the active substance. In a second experiment, substrate was added to a solution of 21~721~

- 49 -thrombin and the active substance which had been incubated for 5 minutes. The increase in the concentration of p-nitroaniline with time was monitored spectroscopically lW-VIS spectrophotometer Lambda-2 from the Perkin-Elmer Company) at 405 nm for 12 min.
Since the measured curves obtained in both experiments were linear and parallel, the active substances of the following table are rapid thrombin inhibitors. The inhibition constants Ki were then determined as follows.
The substrate was used at concentrations of 100 ~M,

50 ~M, 30 ~M, 20 ~M and at each substrate concentration a measurement was carried out without inhibitor and three measurements were carried out in the presence of various concentrations of the inhibitors listed in the following table. The reactions were started by addition of thrombin. The increase in absorbance at 405 nm due to the formation of p-nitroaniline was monitored over a time period of 12 minutes. Measurement points (time versus absorbance) were transferred to a PC at intervals of 20 seconds. The rates VO (ch~ngc in absorbance per second; measurements without inhibitor) and Vi (measurements with inhibitor) are determined by linear regression. Only that part of the measurement was used in which the substrate concentration had decreased by less than 15 %. Km and Vmax were determined from a measurement series (constant inhibitor concentration, variable substrate concentrations) by a non-linear fit to the equation Vmax* [S ]
V = _____ [S] + Km Finally Ki was calculated from the entire series of measurements by non-linear fitting to the equation VmaX* ts]
V = -- __ Km* (1 + [S] /Ki) + [S]

The Michaelis constant Km was 3.8 + 2 ~M in all measurements.
The inhibition constants Ki f the active substances are stated in the following table in units of ~M.

Inhibition of try~sin and plasmin 10 mg bovine pancreatic trypsin (Sigma) was dissolved in 100 ml 1 mM hydrochloric acid and stored in a refrigerator. 20 ~1 of this was admixed with 980 ~1 1 mM
hydrochloric acid. 25 ~1 thereof was used for each measurement. The measurement was carried out as described for thrombin. Km = 45 ~M. The substances listed in the following table do not inhibit trypsin (Ki > 400 ~M)-The measurements with human plasmin (Sigma, 10 units)were carried out as described for thrombin using the substrate S-2251 (H-(D)-Val-Leu-Lys-pNA, Kabi). 0.01 units plasmin were used for each measurement. Km =
250 ~M. The substances listed in the following table do not inhibit plasmin (Ki ~ 400 ~M).

_ 2157~15 Compoun~ TT250 TT25 TT2.5 Ki [~]
of ex~mpl~ thrombin 2 330 85 20 0.40 3 100 25 4 2.00 4 108 21 3 5.00 8 193 49 6 2.00 124 34 0.20 12 44 10 10.00 13 285 71 21 0.70 22 93 23 4 2.00

Claims (11)

C l a i m s

1. Compounds of formula I

(I), in which R1 denotes an aryl, a heteroaryl or a cycloalkyl group which can be substituted if desired, AS denotes an amino acid n denotes the numbers 0 or 1, R2 and R3 are the same or different and denote hydrogen atoms, alkyl, carboxyalkyl or alkoxycarbonylalkyl groups, or R2 and R3 together with the nitrogen atom to which they are bound, form a heterocyclyl ring that can additionally contain a second heteroatom if desired and can be substituted by alkyl, carboxy or alkoxycarbonyl groups, R4 and R5 are the same or different and denote hydrogen atoms or alkyl groups, m denotes the numbers 0, 1 or 2, R6, R7, R8 and R9 are the same or different and denote hydrogen atoms or halogen atoms, as well as their physiologically tolerated salts, hydrates, solvates, racemates and optically active isomers.

2. Compounds of formula I as claimed in claim 1, wherein R1 represents a phenyl, naphthyl, tetrahydronaphthyl, pyridinyl, thienyl, cyclohexyl or chromanyl ring which, if desired, is substituted once or several times by C1-C6 alkyl, C1-C6 alkoxy or halogen groups.

3. Compounds of formula I as claimed in claim 1 or 2, wherein AS denotes glycine, azaglycine, alanine, glutamine, glutamate, asparagine or aspartate.

4. Compounds of formula I as claimed in one of the claims 1-3, wherein n denotes the numbers 0 or 1.

5. Compounds of formula I as claimed in one of the claims 1-4, wherein R2 and R3 are the same or different and independently of one another denote a C1-C6 alkyl, C1-C6 alkoxycarbonyl-C1-C6 alkyl or carboxy-C1-C6 alkyl group or together with the N
atom to which they are bound form a pyrrolidine, piperidine, homopiperidine, morpholine, thiomorpholine or piperazine ring which can be substituted if desired by one or two C1-C6 alkyl, carboxyl or C1-C6 alkoxycarbonyl groups.

6. Compounds of formula I as claimed in one of the claims 1-5, wherein R4 and R5 are the same or different and represent hydrogen atoms or C1-C6 alkyl groups.

7. Compounds of formula I as claimed in one of the claims 1-6, wherein R6, R7, R8, R9 are the same or different and represent hydrogen, fluorine or chlorine atoms.

8. Compounds of formula I as claimed in one of the claims 1-7, wherein R1 denotes phenyl, 4-methylphenyl, 4-chloro-phenyl, 4-methoxyphenyl, 1-naphthyl, 2-naphthyl, 5,6,7,8-tetrahydro-2-naphthyl, 3-pyridinyl, 2-thienyl, cyclohexyl, 2,2,5,7,8-pentamethyl-chroman-6-yl or 4-methoxy-2,3,6-trimethylphenyl, AS denotes glycine, azaglycine or alanine, glutamine, glutamate, asparagine or aspartate, n can be the numbers 0 or 1, R2 and R3 are the same or different and denote ethyl, ethoxycarbonylmethyl or carboxy-methyl, or together with the N atom to which they are bound form a pyrrolidine, piperidine, homopiperidine, morpholine, thiomorpholine or piperazine ring which, if desired, can carry one or two methyl, ethyl, propyl, butyl, carboxyl, methoxy-carbonyl, ethoxycarbonyl or tert. butyl-oxycarbonyl groups, R4 and R5 are the same or different and denote hydrogen atoms or methyl groups, R6, R7, R8, R9 are the same or different and denote hydrogen, fluorine or chlorine atoms.

9. Process for the production of compounds of formula I as claimed in one of the claims 1-8, wherein one reacts in a known manner (a) a compound of the general formula II

( II), in which R1-R5, AS, n and m have the aforementioned meanings with a compound of the general formula III

(III), in which R6-R9 denote halogen;

or b) a compound of formula XI

(XI), in which R2-R9 and m have the stated meanings is reacted with a compound of the general formula VIII

(VIII), in which R1 has the stated meaning and X
represents either a halogen atom or a residue of the general formula IX

(IX), in which Y denotes a halogen atom or a conventional activated residue in peptide chemistry and A denotes a nitrogen atom or an atomic group of the general formula X

(X), in which R1 denotes one of the common amino acid side chains, and subsequently the compounds obtained are converted, if desired, into solvates, hydrates or physiologically tolerated salts and racemates are resolved into enantiomers.

10. Pharmaceutical agent containing at least one compound of formula I as claimed in one of the claims 1 to 8 in addition to pharmacologically acceptable carriers and auxiliary substances.

11. Use of compounds of formula I as claimed in claims 1 to 8 for the production of pharmaceutical agents for the treatment of thromboembolic diseases.