ITMI951688A1 - BICYCLIC LACTAM DERIVATIVES AS THROMBIN INHIBITORS - Google Patents

Abstract

Bicyclic lactams containing an arginine residue that can be used in therapy as thrombin inhibitors are described.

Description

Description of the industrial invention entitled: "DERIVATIVES OF BICYCLIC LACTAMES AS THROMBIN INHIBITORS"

The present invention relates to bicyclic lactam derivatives with antithrombotic activity, to processes for their preparation, to pharmaceutical compositions containing them and to their use as therapeutic agents.

Object of the invention are new derivatives of bicyclic lactams bearing an arginine residue, their salts and hydrates, in pure diastereomeric form or as mixtures of stereoisomers, endowed with inhibitory activity towards some serino-proteases. In particular, the compounds were found to be active in inhibiting the action of the thrombin enzyme and are therefore usable as potential antithrombotic agents, such as antiplatelet or anticoagulants. The new derivatives are characterized by the fact that they present a bicyclic lactam residue, capable of acting as a conformationally constrained analogue of a peptide sequence, such as that constituted by Phe-Pro-Arg, present in the fibrinogen structure and considered important in the recognition of the active site of thrombin. Numerous examples of thrombin inhibitors based on structural modifications of the Phe-Pro-Arg sequence are known, see for example US patents n ° 4478745, US n ° 4399065, EP 526877, US n ° 697987 and Bajusz et al., J. Med. Chem., 1990, 33, 1729-1735 and Kettner et al., Thromb. Res., 1979, 14, 969-973.

The compounds of the invention have general formula I:

in which:

- m is 0, 1, 2 or 3;

is a group of formula -

independently represent hydrogen, COOR7, alkyl, benzyl,

they are independently hydrogen,, linear or branched alkyl, cycloalkyl or an arylachyl or heteroarylalkyl group, optionally substituted on the ring with one or more substituents such as halogen (Cl, Br, I), methoxyl, trifluoromethyl, linear or branched alkyl;

-

it is hydrogen, linear or branched alkyl, cycloalkyl

or an aryl, heteroaryl, arylalkyl or heteroarylalkyl group, optionally substituted on the ring with one or more substituents such as halogen (Cl, Br, I), methoxy, trifluoromethyl, linear or branched alkyl;

is benzyl alkyl;

are independently hydrogen, linear or branched alkyl or a group of general formula -W-Q where:

W can be a group

Q can be a phenyl, benzyl, naphthyl, quinolyl, naphthylmethyl, tetrahydroquinolyl, tetrahydroisoquinolyl group, optionally substituted with one or more groups such as halogen (Cl, Br, I), linear or branched alkyl, methoxy, trifluoromethyl.

The compounds of the invention form with various acids both inorganic and organic salts which are also the object of this invention. Such salts include, for example, hydrochlorides, hydrobromides, sulphates, phosphates, maleates, fumarates.

Examples of alkyl groups are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl.

Examples of cyclo alkyl groups are cyclopropyl, cyclopentyl, cyclohexyl.

An aryl group is preferably phenyl. A heteroaryl group is preferably thienyl or furyl. Examples of arylalkyl groups include benzyl and phenethyl, preferably benzyl. Examples of heteroarylkyl groups include furylmethyl and thienylmethyl.

Examples of and / or groups are benzyl, thienylmethyl, amino, acetylamino, methylamino, dimethylamino, t-butoxycarbonylamino, benzyloxycarbonylamino, naphthylsulfonylamino, quinolylsulfonylamino, benzylsulfonylamino, naphthylmethylsulfonylamino, ethylaminoid.

Examples of Rg groups are phenyl, thienyl, methyl, ethyl.

Preferred compounds of formula I are those in which m is equal to 2,

equals have meanings

previously reported.

Other preferred compounds of formula I are those in which m is equal to 1, is equal to have the previously reported meanings.

Particularly preferred compounds are those in which

they are an arylalkyl, heteroarylalkyl group optionally substituted as indicated above, or a group and is an aryl, heteroaryl, arylalkyl or heteroarylalkyl group, optionally substituted as indicated above, or a group containing a protected amino group. Even more particularly preferred compounds are the following:

According to the invention, compounds of general formula I can be obtained starting from intermediates of general formula VIII

in which and m have the previously reported meanings, by removal of the protective group (or groups) on the guanidine residue. The deprotection reaction, just in case

represents a benzyl, nitro or benzyloxycarbonyl group, it can be carried out in solvents such as ethyl acetate alcohols, tetrahydrofuran, in the presence of a Pd or Pt catalyst supported on carbon and in a hydrogen atmosphere, or, if

represents a t-butoxycarbonyl group, by treatment with strong organic or inorganic acids in non-polar solvents such as dioxane or tetrahydrofuran.

The intermediates of general formula VIII can be obtained from compounds of general formula VII.

in which and m have the previously reported meanings, for example by reduction with metal hydrides, such as

in both non-polar and

polar such as ethyl ether, tetrahydrofuran or alcohols at temperatures between -20 ° C and room temperature. Intermediates of general formula VII can be obtained by coupling compounds of general formula VI, the preparation of which is described in the literature (see for example Balasubramanian N. et. Al, J. Med. Chem., 1993, 36, 300- 303) and in which R2 and R3 have the meanings previously reported,

with compounds of general formula V, in which and m have the previously reported meanings and is equal to hydrogen.

The coupling reaction can be carried out according to one of the methods known from the literature, used for the formation of peptide or amide bonds (see for example Bodanszky M., Peptide Chemistry, chapter V, pages 55-72, Publisher Springer-Verlag) .

For example, the dicyclohexylcarbodiimide carboxylic group, diphenyl phosphorylazide can be used as activators, or mixed anhydrides can be formed by reaction with alkylchloroformates; subsequently the following intermediates can be reacted with the amino derivative of general formula VI, in solvents such as dichloromethane, chloroform, tetrahydrofuran, dimethylformamide, at temperatures generally comprised between 0 ° C and room temperature.

Compounds of general formula V can be obtained by cyclization starting from intermediates of general formula IV, in which

and m have the meanings previously reported, can

be alkyl, X can be Br, I or phenylselenyl.

The cyclization reaction can be carried out in the presence of radical initiators, such as a, a '-azabisisobutyronitrile or dibenzoyl peroxide, by adding trialkyl- or triphenylstannylhydrides, in apolar aprotic solvents such as ethyl ether, tetrahydrofuran, benzene, toluene, carbon tetrachloride, at temperatures generally between the room temperature and the boiling temperature of the solvent. Generally the cyclization reaction is stereoselective and among the various possible stereoisomers, there is the predominant formation of a diastereomer. In the case of obtaining a possible mixture of stereoisomers, it is possible by crystallization and / or chromatographic techniques to separate and purify the single diastereomers.

The compounds of general formula IV can be easily obtained starting from compounds of formula IV ', in which and m have the previously reported meanings and X is equal to hydroxyl, by conversion of the alcohol into the corresponding mesylate, trifluoromethanesulfonate or tosylate and subsequent reaction with sodium or potassium bromides and iodides, in dipolar aprotic solvents such as acetone, dimethylformamide, dimethylsulfoxide at the boiling temperature of the solvent. If X represents a phenylselenyl group, the transformation can be carried out starting from the corresponding alcohol, by treatment with phenylselenophthalimide, in the presence of tributylphosphine, in solvents such as tetrahydrofuran or dichloromethane at temperatures between 0 "C and room temperature .

The intermediates of general formula IV can be obtained starting from intermediates of general formula III, the preparation of which is described in the literature, (see for example, J. Am. Chem. Soc., 1984, 106, 4439-4547), where X, and m have the previously reported meanings

by coupling with compounds of general formula II, in which and R6 have the previously reported meanings.

The coupling reaction can be carried out in the presence of condensing agents, for example dicyclohexylcarbodiimide, in apolar aprotic solvents such as tetrahydrofuran or dichloromethane, at temperatures between 0 ° C and room temperature.

Intermediates of general formula II, in which Rg has the previously reported meanings and R4 (R5) can be an acetylamino, t-butoxycarbonylamino or benzyloxycarbonylamino group, are generally available commercially or easily obtainable starting from the corresponding amino acids, as described in the literature ( see for example Kolar A. J. et al, Synthesis, 1997, 457-459 and Ranganathan D. et al., J. Chem. Soc. Chem. Commun., 1992, (16), 1145-1147).

The compounds disclosed in the present invention act as thrombin inhibitors. For the characterization and evaluation of their efficacy, an in vitro human thrombin inhibition test was chosen (in the presence of tosyl-glycyl-prolyl-arginine-4-nitroaniline acetate as substrate).

The compounds of the invention proved to be active in the above test, showing that they have IC50 values lower than 5 μΜ. The compounds of the invention can therefore be used as active ingredients of pharmaceutical compositions with antithrombotic activity. The compositions of the invention, which can be prepared using conventional methods or excipients, will typically contain from 1 to 1000 mg of compounds I, and can be administered from 1 to 4 times a day by oral, parenteral, transdermal or other convenient route of administration. .

The following examples illustrate the invention without limiting it. The melting points are not correct, the identity of the compounds and their purity have been established by elemental analysis (C, H, N), NMR, IR and mass spectroscopy.

Example 1

(2S, 5R) -1- [2- (acetylamino) propenoyl] -2- (t-butoxycarbonyl) -5- (2-hydroxyethyl) pyrrolidine

DCC (14.4 g, 70 mmol). When solubilization is complete, 2-acetylaminoacrylic acid (6 g, 46.5 mmoles) is added and the mixture is stirred for 18 h at room temperature. The solution, diluted with to decrease the solubility of the urea formed, is filtered and concentrated to dryness. After purification by flash chromatography (AcOEt-MeOH 85:15) 12.1 g of product are obtained, as a yellow spongy solid (yield 80%).

Example 2

(2S, 5R) -1- [2- (acetylamino) propenoyl] -2- (t-butoxycarbonyl) -5- [2- (phenylselenyl) ethyl] pyrrolidine

To a solution of (2S, 5R) -1- [2- (acetylamino) propenoyl] -2- (tbutoxycarbonyl) -5- (2-hydroxyethyl) -pyrrolidine (3.5 g, 10.8 limols) in 50 ml of anhydrous THF in N2 atmosphere and at 0 ° C, tributylphosphine (4.4 g, 21.6 nmoles) and N-phenylselenophthalimide (6.5 g, 21.6 mmoles) are added. The mixture is stirred for 2h at 0 ° C, after which the solvent is removed at reduced pressure and purified by flash chromatography (EtOAcesane 6: 4). 3.2 g of product are obtained, as a pale yellow solid (yield 54%).

Similarly, they prepare:

a) (2S, 5R) -1- [2- (acetylamino) -3-phenylpropenoi1] -2-t-butoxycarbonyl-5- [2- (phenylselenyl) ethyl] pyrrolidine,

<1>

4.31-4.52 (ra, 2H); 6.85 (d, 1H); 7.10-7.80 (m, 11H)

i) 2- (t-butoxycarbonyl) -5- [2- (phenylselenyl) ethyl] -1- [3- (thiophen-2-yl) propenoyl] -pyrrolidine.

Example 3

{2S, 5R) —1— [2- (acetylamino) propenoyl] 2- {t-butoxycarbonyl) —5— (2-iodoethyl) -pyrrolidine

To a solution of (2S, 5R) -1- [2- (acetylamino) propenoyl] -2- (t-butoxycarbonyl) -5- (2-hydroxyethyl) pyrrolidine (10.3 g, 31.5 mmoles) in 200 ml of anhydrous, under an atmosphere of and cooled to 0 ° C, triethylamine (6.1 ml, 44.1 mmol) is added and subsequently a solution of methanesulfonyl chloride (3.1 ml, 39.7, mmol) in 70 ml from

It is stirred at 0'C until the initial product disappears, after which

it is washed several times with water, anhydrified on N and the solvent is removed at reduced pressure. The crude mesylate thus obtained is redissolved in 300 ml of acetone, sodium iodide (22.8 g, 150 mmoles) is added to the solution, finally it is refluxed and after 3 hours the solvent is removed under reduced pressure. The residue is redissolved in

it is washed with water, dried and concentrated a

dryness. After purification by flash chromatography (EtOAc-MeOH 95: 5) 11.0 g of yellowish solid are obtained.

(Yield 80%)

Similarly, they prepare:

Example 4

(3S, 6S, 9aS) -6-acetylamino-3- (t-butoxycarbonyl) -octahydropyrrole [1, 2-a] azepin-5-one

To a solution of (2S, 5R) -2- (t-butoxycarbonyl) -5- (2-iodoethyl) -1- [2- (acetylamino) propenoyl] -pyrrolidine (8.5 g, 19.5 mmol) in 1500 ml of reflux anhydrous benzene and a solution of (6.5 ml, 23.1 mmoles) and AIBN (0.65 g, 3.9 mmoles) in 350 ml of benzene.

At the end of the reaction, it is cooled to room temperature, the volume is reduced to about 300 ml and a saturated solution of KF is added. it is kept under stirring for 8 h, the salts are filtered on a celite bed, the phases are separated and the organic phase is anhydrated on

After elimination of the solvent under reduced pressure and purification by flash chromatography (EtOAc-MeOH 95: 5) 3.7 g of white foamy solid are obtained (yield 62%).

carboxylic

To a solution of (3S, 6S, 9aS) -3- (t-butoxycarbonyl) -6-acetylamino-octahydropyrrole [1,2-a] azepin-5-one (0.88 g, 2.9 mmol) in 10 ml of anhydrous at room temperature, add 10 ml of

The mixture is stirred until the starting terbutyl ester disappears and the solvent is removed under reduced pressure. The residue redissolves in and extracted with 2M. The alkaline aqueous phase is acidified to pH = 2 with 2N HCl and the solvent is evaporated under reduced pressure.

The crude acid is purified by basic ion exchange resin chromatography (DOWEX) eluting with 2N HCl. After elimination of the solvent under reduced pressure and drying, 0.48 g of white solid are obtained (yield 67%).

Similarly, they prepare:

a) (3S, 6R, 8aR) -6-acetylamino-6-benzyl-octahydroindolizin-5-one-3-carboxylic acid,

mp 170-174 ° C (dec)

b) 6-benzyl-octahydroindolizin-5-one-3-carboxylic acid,

c) 6-acetylamino-7-methyl-octahydropyrrole [1,2-a] azepin-5-one-3-carboxylic acid,

d) 6-acetylamino-octahydroindolizin-5-one-3-carboxylic acid,

e) 6 - [(benzyloxycarbonyl) amino] -octahydropyrrole [1,2-a] azepin-5 acid

Example 6

(3S, 6S, 9aS) -N <a> - [(6-acetylamino-octahydropyrrole [1,2-a] azepin-5-one-3-yl] carbons1] -N <w> -benzyloxycarbonyl-L-arginine lactate

To a solution of {3S, 6S, 9aS) -6- (acetylamino) -octahydropyrrole [1,2-a] azepin-5-one-3-carboxylic acid (0.41 g, 1.62 mmol) in 10 ml of anhydrous DMF under an N2 atmosphere and cooled to -15 ° C, N-methylmorpholine (0.18 ml, 1.62 mmol) and isobutyl chloroformate (0.23 ml, 1.62 mmoles). The reaction mixture is stirred for 30 min at -15 ° C, after which a solution of N-methylmorpholine (0.36 ml, 3.24 mmol) and N <w> -benzyloxycarbonyl-L-arginine lactam hydrochloride (0 , 6 g, 1.62 mmol) dissolved in 10 ml of DMF.

The suspension obtained is stirred for 2 hours, allowing it to reach room temperature and at the end the solvent is removed under reduced pressure. The residue is taken up with washing with water and with a saturated solution of NaCl, it is anhydrified and the solvent is removed at reduced pressure. After purification by flash chromatography and trituration with Et20-hexane 0.63 g of product are obtained, as a white amorphous solid (yield 54%).

Similarly, they prepare:

a) [(6-acetylamino-6-benzyl-octahydroindolizin-5-one-3-yl) carbonyl] -N <w> -benzyloxycarbonyl-L-arginine lactam,

b) -N <a> - [[(6-acetamido-7-methyloctahydro [1, 2-a] azepin-5-one-3-yl] carbonyl] -N <w> -benzyloxycarbonyl-L-arginine lactam,

c) -Ν <a> - [[6-acetamido-6-benzyl-hexahydropyrrolizin-5-one-3-yl] carbonyl] -N <w> -benzyloxycarbonyl-L-arginine lactam,

d) -N <a> - [[6-acetamido-6-benzyl-hexahydropyrrolizin-5-one-3-yl] carbonyl] -N <w> -benzyloxycarbon 1-L-arginine lactam,

e) -N <a> - [[6-acetamido-6-ethyl-hexahydropyrrolizin-5-one-3-yl] carbonyl] -N <w> -benzyloxycarbonyl-L-arginine lactam,

f) -Ν <a> - [[6-acetamido-6-methyl-hexahydropyrrolizin-5-one-3-yl] carbonyl] -N <w> -benzyloxycarbonyl-L-arginine lactam,

g) Ν <α> - [[6 - [[(3-methylquinolin-8-yl) sulfonyl] amino] -octahydropyrrole [1,2-a] azepin-5-one-3-yl] carbonyl] -N < w> -benzyloxycarbonyl-L-arginine lactam, h) N <a> - [[6 - [[[(naphthalen-1-yl) methyl] sulfonyl] amino] -octahydro-pyrrole [1,2-a] azepin- 5-one-3-yl] carbonyl] -L-arginine lactam,

Example 7

(3S, 6S, 9aS) -N <a> - [[{6-acetylamino-octahydropyrrole [1,2-a] azepin-5-one) -3-yl] carbonyl] -N <w> -benzyloxycarbonyl-L -arginine aldehyde

To a solution of (3S, 6S, 9aS) -N <a> - [[(6-acetylamino-octahydropyrrole [1,2-a] azepin-5-one) -3-yl] carbonyl] -N <w> benzyloxycarbonyl-L-arginine lactam aldehyde (0.2 g, 0.38 mmol) in 5 ml of anhydrous THF, cooled to -20 ° C and under an atmosphere of N2, a suspension of

(0.010 g, 0.23 mmol) in 4 ml of anhydrous THF. After 2 hours of stirring at -20 ° C, 1N HCl is added up to pH 7, it is left to rise to room temperature and the suspension is filtered on a Si bed, the solvent is removed at reduced pressure and the residue obtained is purified by flash chromatography) . 0.18 g of white amorphous solid are obtained (yield 92%).

Similarly, they prepare:

a) [[(6-acetylamino-6-benzyl-octahydroindolizin-5-one) 3-yl] carbonyl] -N <w> -benzyloxycarbonyl-L-arginine aldehyde

Example 8

(3S, 6S, 9aS) -N <a> - [[[6-acetylamino-octahydropyrrole [1,2-a] -azepin-5-one] -3-yl] carbonyl] -L-arginine aldehyde hydrochloride

To a solution {3S, 6S, 9aS) -of Na - [[(6-acetylamino-octahydropyrrole [1,2-a] azepin-5-one] -3-yl] carbonyl] -N <w> -benzyloxycarbonyl- L-arginine aldehyde (0.18 g, 0.34 mmoles) palladium on carbon (10%, 35 mg) and 0.35 ml of 1N HCl are added in 10 ml of THF. it is left to react under stirring until the initial product disappears. It is filtered on a celite bed and the solvent is removed under reduced pressure. After trituration with 0.13 g of amorphous white solid are obtained.

(Yield 88%)

Similarly, they prepare:

a) [[(6-acetylamino-6-benzyloctahydroindolizin-5-one) -3-yl] carbonyl] -L-arginine aldehyde hydrochloride

b) -Ν <a> - [[(6-acetylamino-7-methyloctahydro [1,2-a] azepin-5-one) -3-yl] carbonyl] -L-arginine aldehyde hydrochloride,

c) -N <a> - [[(6-acetylamino-6-benzyl-hexahydropyrrolysin-5-one) -3-yl] carbo

Claims (8)

CLAIMS 1. Compounds of general formula I: in which: - m is 0, 1, 2 or 3; is a formula group independently represent hydrogen, benzyl alkyl, they are independently hydrogen, linear or branched alkyl, cycloalkyl or an arylalkyl or heteroarylalkyl group, optionally substituted on the ring with one or more substituents such as halogen (Cl, Br, I), methoxyl, trifluoromethyl, linear or branched alkyl; is hydrogen, linear or branched alkyl, cycloalkyl or an aryl, heteroaryl, arylalkyl or heteroarylalkyl group, optionally substituted on the ring with one or more substituents such as halogen (C1, Br, I), methoxy, trifluoromethyl, linear or branched alkyl; is benzyl alkyl; are independently hydrogen, linear or branched alkyl or a group of general formula -W-Q where: W could be a group Q can be a phenyl, benzyl, naphthyl, quinolyl, naphthylmethyl, tetrahydroquinolyl, tetrahydroisoquinolyl group, optionally substituted with one or more groups such as halogen (C1, Br, I), alkyl linear or branched, methoxy, trifluoromethyl, their stereoisomeric forms and their salts with pharmaceutically acceptable acids. Compounds according to claim 1, wherein m is equal to 2, R1 is equal to is equal to hydrogen and have the meanings reported above. 3. Compounds according to claim 2, which are: (3S, 6S, 9aS) -N <a> - [[6-acetylamino-octahydropyrrole [1,2-a] azepin-5-one-3-yl] carbonyl] -L-arginine aldehyde, (3S, 6S, 9aS) -N <α> - [[6-methylamino-octahydropyrrole [1,2-a] azepin-5-one-3-yl] -carbonyl] -L-arginine aldehyde, (3S, 6S, 9aS) -N <α> [[6 - [(benzylsulfonyl) amino] -octahydropyrrole- [1,2-a] azepin-5-one-3-yl] carbonyl] -L-arginine aldehyde, (3S, 6S, 9aS) -N <a> - [[6 - [[[naphthalen-1-yl) methyl] solionyl] amino] -octahydropyrrole [1,2-a] azepin-5-one-3-yl ] carbonyl] -L-arginine aldehyde, (3S, 6S, 9aS) -N <a> - [[6 - [[[(naphthalen-2-yl) methyl] sulfonyl] amino] -octahydropyrrole [1,2-a] azepin-5-one-3- the] carbonyl] -L-arginine aldehyde, (3S, 6S, 9aS) -N <a> - [[6 - [[(3-methylquinolin-8-yl) sulfonyl] amino] -octahydropyrrole [1,2-a] azepin-5-one-3-yl ] carbonyl] -L-arginine aldehyde 4. Compounds according to claim 1, wherein m is equal to 1, R1 is equal to have the meanings reported above. 5. Compounds according to claim 4, which are: 6. A process for the preparation of compounds of general formula I, which involves the coupling reaction of intermediates of general formula III in which m and have the previously reported meanings and X is equal to -OH, with compounds of general formula II, in which have the previously reported meanings to give compounds of general formula IV, which are transformed into compounds of general formula IV, where X is equal to which by cyclization are transformed into compounds of general formula V. that following hydrolysis of the carboxy group and coupling with intermediates of general formula VI are transformed into compounds of general formula VII which by reduction are transformed into compounds of general formula VIII which by removal of the protective groups and treatment with organic or inorganic acids, are transformed into the compounds of general formula I. 7. Use of the compounds of claims 1-5, as therapeutic agents to be used as antithrombotic, anticoagulant or antiplatelet agents. 8. Pharmaceutical compositions containing as active ingredients an effective amount of one or more compounds, of claims 1-5, in association with suitable excipients.