WO2001077095A2

WO2001077095A2 - Derivatives of alpha-mercaptoacetamide

Info

Publication number: WO2001077095A2
Application number: PCT/EP2001/003848
Authority: WO
Inventors: Stéphane De Lombaert; Fariborz Firooznia; Denton Wade Hoyer
Original assignee: Novartis Ag; Novartis-Erfindungen Verwaltungsgesellschaft M.B.H.
Priority date: 2000-04-06
Filing date: 2001-04-04
Publication date: 2001-10-18
Also published as: AU2001254776A1; WO2001077095A3

Abstract

Novel thiol derivatives of formula (I) wherein variables R1, R2, R3, R4, A, X, and Y have the meanings as defined hereinbefore; disulfide derivatives derived from said compounds wherein R1 is hydrogen; and pharmaceutically acceptable salts thereof; said compounds being useful as endothelin converting enzyme inhibitors.

Description

Organic Compounds

The endothelins (ET-1 , ET-2, ET-3) constitute a family of potent vasoconstrictor and mitogenic peptides produced by various cells, including vascular endothelial, epithelial, and smooth muscle cells. Elevated levels of ET-1 have been measured in a variety of human disease states.

Endothelins are produced by a unique proteolytic cleavage at the Trp²¹ -Val²² or Trp²¹ -lie²² bond of the corresponding and biologically inactive propeptides, termed big ETs, by one or more endothelin-converting enzymes (ECE).

Since the blockade of ECE constitutes a means to prevent or decrease endothelin (e.g. ET-1 ) production, inhibitors of this enzyme offer an attractive therapeutic potential in the treatment of associated disorders.

The aim of the present invention is to provide the compounds of formula I below which are useful as endothelin-converting enzyme (ECE) inhibitors in mammals, including human beings.

The compounds of the invention thus inhibit the formation of endothelin, reduce the plasma and tissue levels of eηdothelin and inhibit the biological effects of endothelin activity in mammals. The compounds of the invention are thus also useful for the treatment and/or prevention of endothelin dependent conditions and diseases, e.g. cardio- and cerebro- vascular disorders such as essential hypertension, congestive heart failure, pulmonary hypertension, cerebral ischemia (stroke), subarachnoid hemorrhage, traumatic brain injury, acute and chronic renal failure, atherosclerosis, cerebral vasospasm, arterial hypertrophy, restenosis, Raynaud's disease, myocardial infarction, obesity; also respiratory disorders such as bronchial asthma; gastrointestinal disorders such as inflammatory bowel disease, pancreatitis, emesis; also prostate hyperplasia, migraine, diabetes mellitus (diabetic nephropathy), preeclampsia, glaucoma and transplantation rejection, such as in aorta or solid organ transplantation in either allo- or xeno- transplantation.

The present invention relates to the novel thiol derivatives of formula I

wherein

^ represents hydrogen or acyl;

R₂ represents hydrogen, lower alkyl, carbocyclic or heterocyclic aryl, carbocyclic or heterocyclic aryl-lower alkyl, cycloalkyl, cycloalkyl-iower alkyl, biaryi, biaryl-lower alkyl,

(hydroxy, lower alkoxy or acyloxy)-lower alkyl, X or lower alkyl-(thio, sulfinyl or sulfonyl)- lower alkyl;

R₃ represents hydrogen or lower alkyl; or R₂ and R₃ together with the carbon atom to which they are attached represent cycloalkylidene or benzo-fused cycloalkylidene;

A together with the carbon atom to which it is attached forms a ring and represents 3 to 10 membered cycloalkylidene or 5 to 10 membered cycloalkenylidene radical which may be substituted by lower alkyl or aryl-lower alkyl or may be fused to a saturated or unsaturated carbocyclic 5-7-membered ring; or A together with the carbon to which it is attached represents 5 to 6 membered oxacycloalkylidene, thiacycioalkylidene or azacycloalkylidene optionally substituted by lower alkyl or aryl-lower alkyl; or A together with the carbon atom to which it is attached represents 2,2-norbonylidene;

X represents O, CH₂, C=O, N-R₄ or S;

R is hydrogen, acyl, lower alkyl or aryl-lower alkyl;

Y represents 5-tetrazolyl, carboxyl or carboxyl derivatized in form of a pharmaceutically acceptable ester; disulfide derivatives derived from said compounds wherein Ri is hydrogen; and pharmaceutically acceptable salts thereof; pharmaceutical compositions comprising said compounds; methods for preparation of said compounds; intermediates; and methods of treating disorders in mammals which are responsive to ECE inhibition by administration of said compounds to mammals in need of such treatment.

Pharmaceutically acceptable esters are preferably prodrug ester derivatives, such being convertible by solvolysis or under physiological conditions to the free carboxylic acids of formula I. Encompassed by the instant invention are any prodrug derivatives of compounds of the invention having a free carboxyl, sulfhydryl or hydroxyl group, said prodrug derivatives being convertible by solvolysis or under physiological conditions to the free carboxyl, sulfhydryl and/or hydroxyl compounds. Prodrug derivatives are e.g. the esters of free carboxylic acids and S-acyl and O-acyl derivatives of thiols, or alcohols, wherein acyl has meaning as defined herein.

Pharmaceutically acceptable prodrug esters of carboxylic acids are preferably e.g. lower alkyl esters, cycloalkyl esters, lower alkenyl esters, aryl-lower alkyl esters, α-(lower alkanoyloxy)-lower alkyl esters such as the pivaloyioxy-methyl ester, and α-(lower alkoxycarbonyl- or di-lower alkylamino carbonyl-)-lower alkyl esters.

Pharmaceutically acceptable salts are salts derived from pharmaceutically acceptable bases for any acidic compounds of the invention, e.g. those wherein Y represents carboxyl. Such are e.g. alkali metal salts (e.g. sodium, potassium salts), alkaline earth metal salts (e.g. magnesium, calcium salts), amine salts (e.g. tromethamine salts).

Compounds of formula I, depending on the nature of substituents, possess one or more asymmetric carbon atoms. The resulting diastereomers and optical antipodes are encompassed by the instant invention. Preferred is the configuration wherein the asymmetric carbon with the substituent Y has the S-configuration.

Preferred are the compounds of formula I wherein R, represents hydrogen or acyl derived from a carboxylic acid; R₂ represents hydrogen, lower alkyl, hydroxy-iower alkyl, or carbocyclic or heterocyclic aryl-lower alkyl; R₃ represents hydrogen; A represents C₂-C₆- straight chain alkylene optionally substituted by lower alkyl, or C₂-C₄-straight chain alkylene interrupted by 1 ,2-phenyIene or by 1 ,2-C₅-or C₆-cycloalkylene, or C₃- or C₄-straight chain alkylene interrupted by oxygen, sulfur or by NR₄ wherein R₄ is hydrogen, acyl, aryl-lower alkyl or lower alkyl; X is O, S, CH₂ or C=O; Y represents 5-tetrazolyl, carboxyl or carboxyl derivatized in form of a pharmaceutically acceptable ester; disulfide derivatives derived from said compounds wherein Ri is hydrogen; and pharmaceutically acceptable salts thereof.

Preferred are the compounds with the S-configuration of formula II

wherein

Ri represents hydrogen or carboxyl derived acyl; R₂ represents hydrogen, lower alkyl, hydroxy-lower alkyl, carbocyclic or heterocyclic aryl, carbocyclic or heterocyclic aryl-lower alkyl, cycloalkyl, cycloalkyl-lower alkyl, biaryl or biaryl-lower alkyl; X is O, S, CH₂ or C=O; Y represents 5-tetrazolyl, carboxyl or carboxyl derivatized in form of a pharmaceutically acceptable ester; n represents 2-6, preferably 2, 4 or 5; disulfide derivatives derived from said compounds wherein Ri is hydrogen; and pharmaceutically acceptable salts thereof.

Further preferred are said compounds of formula II wherein n represents 2, 4 or 5.

Further preferred are said compounds of formula II wherein R^ represents hydrogen, aryl-lower alkanoyl, lower alkanoyl, lower alkoxy-lower alkanoyl, or heterocyclic or carbocyclic aroyl; R₂ represents hydrogen, lower alkyl or carbocyclic aryl-lower alkyl; X is O; Y represents 5-tetrazolyl, carboxyl, lower alkoxycarbonyl, carbocyclic or heterocyclic aryl-lower alkoxycarbonyl, oc-(lower alkanoyloxy-, lower alkoxycarbonyl- or di-lower alkylaminocarbonyl-)- lower alkoxycarbonyl; n is 2, 4 or 5; and pharmaceutically acceptable salts thereof.

Particularly preferred are said compounds with the S-configuration of formula III

and of formula Ilia

wherein

X is O, S, CH₂ or C=O, preferably O;

Ri represents hydrogen, lower alkanoyl, methoxy-lower alkanoyl, benzoyl or pyridylcarbonyl;

R₂ represents hydrogen, lower alkyl, hydroxy-lower alkyl, benzyl, or benzyl substituted by hydroxy, halo, lower alkyl, acyloxy, lower alkoxy or trifluoromethyl;

Y represents 5-tetrazolyl, carboxyl, lower alkoxycarbonyl, benzyloxycarbonyl, pyridylmethoxycarbonyl, α-(lower alkanoyloxy-, lower alkoxycarbonyl- or di-lower alkylaminocarbonyl-) lower alkoxycarbonyl; and pharmaceutically acceptable salts thereof.

A further embodiment of the invention relates to the compounds with the S- configuration of formula 1Mb

wherein

X is O, S, CH₂ or C=O, preferably O;

W represents CH₂, O, S or NR₄ in which R₄ is hydrogen, acyl, lower alkyl or aryl-lower alkyl; Ri represents hydrogen, lower alkanoyl, methoxy-lower alkanoyl, benzoyl or pyridylcarbonyl;

Further preferred are said compounds of formula III, Ilia or lllb wherein X is O; Ri represents hydrogen, or lower alkanoyl; R₂ represents lower alkyl, benzyl, or benzyl substituted by hydroxy, halo, lower alkyl, lower alkanoyloxy, lower alkoxy or trifluoromethyl; Y represents 5-tetrazolyl, carboxyl, lower alkoxycarbonyl, benzyloxycarbonyl, pyridylmethoxycarbonyl, α-(lower alkanoyloxy-, lower alkoxycarbonyl- or di-lower alkylaminocarbonyl-) lower alkoxycarbonyl; and pharmaceutically acceptable salts thereof.

A particular preferred embodiment relates to compounds of any of the above formulae wherein X is O;

is hydrogen, or lower alkanoyl; R₂ is Cι-C₄-alkyl; and Y is 5-tetrazolyl, carboxyl or lower alkoxycarbonyl.

Particularly preferred are said compounds of formula II, III, Ilia and lllb wherein X is O; Ri represents hydrogen; R₂ represents Crd-alky!; Y represents carboxyl; ester and S-acyl prodrug derivatives thereof; and pharmaceutically acceptable salts thereof.

The definitions as such or in combination as used herein, unless denoted otherwise, have the following meanings within the scope of the present invention.

Aryl represents carbocyclic or heterocyclic aryl, either monocyclic or bicyclic.

Monocyclic carbocyclic aryl represents optionally substituted phenyl, being preferably phenyl or phenyl substituted by one to three substituents, such being advantageously lower alkyl, hydroxy, lower alkoxy, acyloxy, halogen, cyano, trifluoromethyl, amino, lower alkanoylamino, lower alkyl-(thio, sulfinyl or sulfonyl), lower alkoxycarbonyl, mono- or di-lower alkylcarbamoyl, or mono- or di-lower alkyiamino.

Bicyclic carbocyclic aryl represents 1- or 2-naphthyl or 1- or 2-naphthyl preferably substitued by lower alkyl, lower alkoxy or halogen. Monocyclic heterocyclic aryl represents preferably optionally substituted thiazolyl, thienyl, furanyl or pyridyl.

Optionally substituted furanyl represents 2- or 3-furanyl or 2- or 3-furanyl preferably substituted by lower alkyl.

Optionally substituted pyridyl represents 2-, 3- or 4-pyridyl or 2-, 3- or 4-pyridyl preferably substituted by lower alkyl, halogen or cyano.

Optionally substituted thienyl represents 2- or 3-thienyl or 2- or 3-thienyl preferably substituted by lower alkyl.

Optionally substituted thiazolyl represents e.g. 4-thiazolyl, or 4-thiazolyl substituted by lower alkyl.

Bicyclic heterocyclic aryl represents preferably indolyl or benzothiazolyl optionally substituted by hydroxy, lower alkyl, lower alkoxy or halogen, advantageously 3-indolyl or 2- benzothiazolyl.

Aryl in aryl-lower alkyl is preferably phenyl or phenyl substituted by one or two of lower alkyl, lower alkoxy, hydroxy, lower alkanoyloxy, halogen, trifluoromethyl, cyano, lower alkanoylamino or lower alkoxycarbonyl; also, optionally substituted naphthyl.

Aryl-lower alkyl is advantageously benzyl or 1- or 2-phenethyl optionally substituted on phenyl by one or two of lower alkyl, lower alkoxy, hydroxy, lower alkanoyloxy, halogen or trifluoromethyl.

The term "lower" referred to herein in connection with organic radicals or compounds respectively defines such with up to and including 7, preferably up to and including 4 and advantageously one or two carbon atoms. Such may be straight chain or branched.

A lower alkyl group preferably contains 1-4 carbon atoms and represents for example ethyl, propyl, butyl or advantageously methyl.

A lower alkoxy group preferably contains 1-4 carbon atoms and represents for example methoxy, propoxy, isopropoxy or advantageously ethoxy. Cycloalkyl represents a saturated cyclic hydrocarbon radical which preferably contains 5 to 7 ring carbons, preferably cyclopentyl or cyclohexyl.

The term cycloalkyl(lower) alkyl represents preferably 1- or 2-(cyclopentyl or cyclohexyl)ethyl, 1-, 2- or 3-(cyclopentyl or cyclohexyl)propyl, or 1-, 2-, 3- or 4-(cyclopentyl or cyclohexyl)-butyl.

A lower alkoxycarbonyl group preferably contains 1 to 4 carbon atoms in the alkoxy portion and represents, for example, methoxycarbonyl, propoxycarbonyl, isopropoxycarbonyl or advantageously ethoxycarbonyl.

Cycloalkylidene is 3 to 10 membered, preferably 3, 5 or 6-membered, and represents a cycloalkane linking group e.g. cyclopropylidene, cyclopentylidene, cyclohexylidene, cycloheptylidene or cycloctylidene, in which the two attached groups are attached to the same carbon of the cycloalkane ring.

Cycloalkenylidene is 5 to 10 membered, prefereably 5 or 6-membered, and represents a cycloalkene linking group in which the two attached groups are attached to the same carbon atom of the cycloalkene ring.

Cycloalkylidene fused to a saturated carbocyclic ring represents e.g. perhydronaphthylidene.

Cycloalkylidene fused to an unsaturated carbocyclic ring represents e.g. 1 ,1- or 2,2- tetralinylidene or 1 ,1 - or 2,2-indanylidene.

5 or 6 Membered oxacycloalkylidene represents preferably a tetrahydrofuran or tetrahydropyran linking group, e.g. tetrahydrofuranylidene or tetrahydropyranylidene, in which the two attached groups are attached to the same carbon atom of the respective rings, e.g. at the 3 or 4 position thereof. -

5 or 6 Membered thiacycloalylidene represents preferably a tetrahydrothiophene or tetrahydrothiopyran linking group in which the two attached groups are attached to the same carbon atom of the respective rings, e.g. at the 3 or 4 position thereof. 5 or 6 Membered azacyloalkyllidene represents preferably a pyrrolidine or piperidine linking groups in which the two attached groups are attached to the same carbon atom of the respective rings, e.g. at the 3 or 4 position thereof, and the nitrogen may be substituted by lower alkyl, e.g. methyl, or by aryl-lower alkyl, e.g. benzyl.

Halogen (halo) preferably represents fluoro or chloro, but may also be bromo or iodo.

Acyl is derived from a carboxylic acid and represents preferably optionally substituted lower alkanoyl, cycloalkylcarbonyl, carbocyclic aryl-lower alkanoyl, aroyl, lower alkoxycarbonyl or aryl-lower alkoxycarbonyl, advantageously optionally substituted lower alkanoyl or aroyl.

Lower alkanoyl is preferably acetyl, propionyl, butanoyl* pentanoyl, or pivaloyl.

Optionally substituted lower alkanoyl for example represents lower alkanoyl or lower alkanoyl substituted by lower alkoxycarbony, lower alkanoyloxy, lower alkanoylthio, lower alkoxy, or by lower alkylthio; also lower alkanoyl substituted by e.g. hydroxy, di-lower alkylamino, lower alkanoylamino, morpholino, pipeidino, pyrrolidino or 1 -lower alkylpiperazino.

Aroyl is carbocyclic or heterocyclic aroyl, preferably monocyclic carbocyclic or monocyclic heterocyclic aroyl.

Monocyclic carbocyclic aroyl is preferably benzoyl or benzoyl substituted by lower alkyl, lower alkoxy, halogen or trifluoromethyl.

Monocyclic heterocyclic aroyl is preferably pyridylcarbonyl or thienylcarbonyl.

Acyloxy is preferably optionally substituted lower alkanoyloxy, lower alkoxycarbonyloxy, monocyclic carbocyclic aroyloxy or monocyclic heterocyclic aroyloxy; also carbocyclic or heterocyclic aryl-lower alkanoyloxy.

Optionally substituted lower alkanoyloxy is preferably lower alkanoyloxy, such as acetyloxy, substituted by any group indicated above under optionally substituted alkanoyl. Aryl-lower alkoxycarbonyl is preferably monocyclic carbocyclic-lower alkoxycarbonyl, advantageously benzyloxycarbonyl.

Biaryl represents monocarbocyclic aryl substituted by monocyclic carbocyclic or monocyclic heterocyclic aryl, and preferably represents biphenylyl, advantageous 4- biphenylyl optionally substituted on one or both benzene rings by lower alkyl, lower alkoxy, halogen or trifluoromethyl.

Biaryl-lower alkyl is preferably 4-biphenylyl-lower alkyl, advantageously 4-biphenylyl- methyl.

The novel compounds of the invention are pharmacologically potent endothelin converting enzyme inhibitors which inhibit the formation of endothelin in mammals. They thus inhibit the biological effects of endothelin in mammals.

The compounds of the invention are thus particularly useful in mammals for the treatment of e.g. hypertension and heart failure, cerebral vasospasm and stroke, bronchial asthma, and complications associated with organ transplantations.

The above-cited properties are demonstrable in vitro and in vivo tests, using advantageously mammals, e.g. mice, rats, dogs, monkeys or isolated organs, tissues and preparations thereof. Said compounds can be applied in vitro in the form of solutions, e.g. preferably aqueous solutions, and in vivo either enterally, parenterally, advantageously intravenously, e.g. as a suspension or in aqueous solution. The dosage in vitro may range between about 10^'5 molar and 10^'9 molar concentrations. The dosage in vivo may range depending on the route of administration, between about 0.1 and 50 mg/kg, advantageously between about 1.0 and 25 mg/kg.

The in vitro inhibition of endothelin-converting enzyme can be determined as follows:

The test compound is dessolved in dimethyl sulfoxide or 0.25 M sodium bicarbonate solution, and the solution is diluted with a pH 7.4 buffer to the desired concentration.

Endothelin converting enzyme (ECE) is partially purified from porcine primary aortic endothelial cells by DE52 anion exchange column chromatography and its activity is quantified by radioimmunoassay as described in Anal. Biochem.212, 434-436 (1993). Alternatively, the native enzyme can be substituted by a recombinant form of ECE, as described, for example in Cell 78, 473-485 (1994). Human ECE-1 has been described by several groups (Schmidt, M. et al. FEBS Letters, 1994, 356, 238-243; Kaw, S.; Emoto, N.; Jeng, A.; Yanagisawa, M. 4th Int. Conf. on Endothelin; April 23-25, London (UK), 1995; C6; Valdenaire, O. et al. J. Biol. Chem. 1995, 270, 29794-29798; Shimada, K. et al. Biochem. Biophys. Res. Commun., 1995, 207, 807-812). The ECE inhibiton can be determined as described in Biochem. Mol. Biol. Int. 31, (5), 861-867 (1993), by radioimmunoassay to measure ET-1 formed from big ET-1.

Alternatively, recombinant human ECE-1 (rhECE-1 ) can be used, as follows:

Chinese hamster ovary cells expressing recombinant human endothelin converting enzyme-1 (rhECE-1; Kaw, S.; Emoto, N.; Jeng, A.; Yanagisawa, M. 4th Int. Conf. on Endothelin; April 23-25, London (UK), 1995; C6) are cultured in DMEM/F12 medium containing 10% fetal bovine serum and 1x antibiotic-antimycotic. Cells are harvested by scraping, pelleted by centrifugation, and homogenized at 4 °C in a buffer containing 5 mM MgCI₂, 1 μM pepstatin A, 100 μM leupeptin, 1 mM PMSF, and 20 mM Tris, pH 7.0, with a ratio of 2 mL of buffer/mL of cells. The cell debris is removed by brief centrifugation, and the supernatant is centrifuged again at 100,000 x g for 30 minutes. The resulting pellet is resuspended in a buffer containing 200 mM NaCl and 50 mM Tes, pH 7.0, at a protein concentration about 15 mg/mL and stored in aliquots at -80°C.

To assess the effect of an inhibitor on ECE-1 activity, 10 μg of protein is pre- incubated with the compound at a desired concentration for 20 min at room temperature in 50 mM TES, pH 7.0, and 0.005% Triton X-100 in a volume of 10 μL. Human big ET-1 (5 μL) is then added to a final concentration of 0.2 μM, and the reaciton mixture is further incubated for 2 h at 37 °C. The reaction is stopped by adding 500 μL of radioimmunoassay (RIA) buffer containing 0.1% Triton X-100, 0.2% bovine serum albumin, and 0.02% NaN₃ in phosphate-buffered saline.

Diluted samples (200 μL) obtained from the above enzyme assay are incubated at 4 °C overnight with 25 μL each of [¹²⁵I]ET-1 (10,000 cpm/tube) and 1 :20,000-fold diluted rabbit antibodies that recognize specifically the carboxyl terminal tryptophan of ET-1. Goat anti-rabbit antibodies coupled to magnetic beads (70 μg) are then added to each tube, and the reaction mixture is further incubated for 30 min at room temperature. The beads are pelleted using a magnetic rack. The supernatant is decanted, and the radioactivity in the pellet is counted in a gamma counter. Total and nonspecific binding are measured in the absence of non radioactive ET-1 and anti-ET antibodies, respectively. Under these conditions, ET-1 and big ET-1 displace [¹²⁵I]ET-1 binding to the antibodies with IC₅₀ values of 21 ± 2 and 260,000 + 66,000 fmol (mean ± SEM, n = 3 - 5), respectively.

In order to determine the IC₅₀ value of an inhibitor, a concentration-response curve of each inhibitor is determined. An IBM-compatible version of ALLFIT program is used to fit data to a one-site model.

In vitro testing is most appropriate for the compounds wherein Y is 5-tetrazolyl or carboxyl.

Illustrative of the invention, the compound of Example 1 demonstrates an IC₅₀ of about 11 nM in the in vitro assay for rh-ECE-1 inhibition.

Endothelin converting enzyme inhibition can also be determined in vivo by measuring the inhibition of big ET-1 -induced pressor response in the anesthesized or conscious rat, as described below. The effect of the inhibitors on the pressor response resulting from big ET- 1 challenge is measured in Sprague-Dawley rats as described in Biochem. Mol. Biol. Int. 31, (5), 861-867 (1993). Results are expressed as percent inhibition of the big ET-1 -induced pressor response as compared to vehicle.

Male Sprague-Dawley rats are anesthetized with Inactin (100 mg/kg i.p.) and instrumented with catheters in the femoral artery and vein to record mean arterial pressure (MAP) and administer compounds, respectively. A tracheostomy is performed and a cannula inserted into the trachea to ensure airway patency. The body temperature of the animals is maintained at 37 ± 1 °C by means of a heating blanket. Following surgery, MAP is allowed to stabilize before interrupting autonomic neurotransmission with chlorisondamine (3 mg/kg i.v.). Rats are then treated with the test compound at 10 mg/kg i.v. or vehicle and challenged with big ET-1 (1 nmol/kg i.v.) 15 min and 90 min later. Generally, the data are reported as the maximum increase in MAP produced by big ET-1 in animals treated with the test compound or vehicle.

Male Sprague-Dawley rats are anesthetized with methohexital sodium (75 mg/kg i.p.) and instrumented with catheters in the femoral artery and vein to measure mean arterial pressure (MAP) and administer drugs, respectively. The catheters are threaded through a swivel system that enables the rats to move freely after regaining consciousness. The rats are allowed to recover from this procedure for 24 h before initiating the study. On the following day, MAP is recorded via the femoral artery catheter and a test compound or vehicle is adminstered via the femoral vein. Animals are challenged with big ET-1 at 1 nmol/kg i.v. at various times after dosing. After an adequate washout period, depending upon the dose and regimen, animals can be re-tested at another dose of test compound or vehicle. Generally, the data are reported as the change in MAP produced by big ET-1 at 2- minute intervals in animals treated with the test compound as compared to vehicle.

Illustrative of the invention, the compound of Example 1 inhibits the big ET-1 -induced pressor response by about 50% at 90 minutes.

ECE inhibition can also be determined in vivo by measuring the inhibition of the big ET-1 induced pressor response in conscious spontaneously hypertensive rats (SHR), e.g. as described in Biochem. Biophys. Res. Commun.204, 407-412 (1994).

Male SHR (16-18 weeks of age) are administered either test compound or vehicle (1 M NaHCO₃) via an osmotic minipump implanted subcutaneously. On day 5 femoral arterial and venous catheters are placed in anesthetized rats for the measurement of MAP and for test compound administration, respectively. After a 48 hour recovery period, MAP is recorded (day 7) through the arterial catheter connected to a pressure transducer. Blood pressure and heart rate are allowed to stabilize for 30 min before ganglion blockade is performed using chlorisondamine (10 mg/kg i.v.). Approximately 15 min later, a bolus dose of big ET-1 (0.25 nmol/kg i.v.) is administered to both vehicle- and test compound treated rats. The change in blood pressure in response to big ET-1 is then compared between the two groups of rats at 1 , 5, 10, 15, 30 and 60 min after dosing using a two-way ANOVA.

The compounds of the invention inhibit cerebrovascular constriction and are useful for the treatment and alleviation of cerebral spasm. They are thus in turn useful for the treatment and alleviation of conditions in which cerebral vasospasm occurs. Such conditions include stroke, cerebral ischemia, acute and traumatic brain injury, brain hemorrhage, in particular aneurysmal subarachnoid hemorrhage, as well as migraine.

The inhibition of cerbral vasospasm is demonstrated by measuring the inhibition of experimentally induced constriction of basilar cerebral arteries in the rabbit (Caner et al., J. Neurosurg., 1996, 85, 917-922). Bronchial effects can be determined by measuring the effect in a model of ET-1 induced bronchoconstriction.

The compounds of the invention may also possess neutral endopeptidase (NEP) inhibitory activity. Tests for determination thereof are described e.g. in U.S. patent 5,506,244 which is incorporated herein by reference.

The combined effect is beneficial for e.g. the treatment of cardiovascular disorders in mammals such as hypertension, congestive heart failure and renal failure.

The compounds of the invention (as represented by formula I) can be prepared using the processes described and illustrated below, e.g.

(a) by condensing a compound of formula IV

wherein the symbols A and X have the meaning as defined above and Y represents N- protected 5-tetrazolyl or esterified carboxyl, with a carboxylic acid of the formula V

or a reactive funcitonal derivative thereof, wherein R₂ and R₃ have meaning as defined above, Ri' represents a labile S-protecting group, e.g. acyl, t-butyl or optionally substituted benzyl; or

(b) by condensing a compound of the formula VI

or a reactive functional derivative thereof wherein the symbols A, Ri', R₂ and R₃ have meaning as defined above, with a compound of the formula VII

wherein X and Y' have meaning as defined above; or

(c) by condensing under basic conditions a compound of the formula

wherein the symbols A, R₂, R₃, and Y' have meaning as defined above and Z represents a reactive esterified hydroxyl group (e.g. halo such as chloro or bromo) as a leaving group, with a compound of the formula

wherein Ri' represents a labile S-protecting group, e.g. acyl, t-butyl or optionally substituted benzyl; and converting a resulting product wherein Ri' is optionally substituted benzyl to a compound of formula I wherein Ri is hydrogen; and in above said process, if temporarily protecting any interfering reactive group(s), removing said protecting group(s), and then isolating the resulting compound of the invention; and, if desired, converting any resulting compound of the invention into another compound of the invention; and/or, if desired, converting a free carboxylic acid function into a pharmaceutically acceptable ester derivative, or converting a resulting ester into the free acid or into another ester derivative; and/or, if desired, converting a resulting free compound into a salt or a resulting salt into the free compound or into another salt, and/or, if desired, separating a mixture of isomers or racemates obtained into the single isomers or racemates, and/or, if desired, resolving a racemate obtained into the optical antipodes.

In starting compounds and intermediates which are converted to the compounds of the invention in manner described herein, functional groups present, such as thiol, carboxyl, amino and hydroxyl groups, are optionally protected by conventional protecting groups that are common in preparative organic chemistry. Protected thiol, carboxyl, amino and hydroxyl groups are those that can be converted under mild conditions into free thiol, carboxyl, amino and hydroxyl groups without other undesired side reactions taking place.

The purpose of introducing protecting groups is to protect the functional groups from undesired reactions with reaction components and under the conditions used for carrying out a desired chemical transformation. The need and choice of protecting groups for a particular reaction is known to those skilled in the art and depends on the nature of the functional group to be protected (thiol, carboxyl, amino group, etc.), the structure and stability of the molecule of which the substituent is a part, and the reaction conditions.

Well-known protecting groups that meet these conditions and their introduction and removal are described, for example, in J. F. W. McOmie, "Protective Groups in Organic Chemistry", Plenum Press, London, N.Y. 1973, T. W. Greene and P.G.M. Woots, "Protective Groups in Organic Synthesis", Wiley, N.Y. 1991, 'The Peptides", Vol. I, Schroeder and Luebke, Academic Press, London, N.Y., 1965, and also in P. J. Kocienski, "Protecting Groups", Thieme, N.Y. 1994. Suitable protecting groups for the preparation of the 5-tetrazolyl compounds are the protecting groups customarily used in tetrazole chemistry, especially triphenylmethyl, unsubstituted or substituted, (for example nitro-substituted), benzyl such as 4-nitrobenzyl, lower alkoxymethyl such as methoxy- and ethoxymethyl, also 1-ethoxyethyl, lower alkylthiomethyl such as methylthiomethyl, silyl such as tri-lower alkylsilyl, for example dimethyl-tert-butyl- and triisopropyl-silyl, and also 2-cyanoethyl, also lower alkoxy-lower alkoxy-methyl, such as 2-methoxyethoxymethyl, benzyloxymethyl and phenacyl.

The removal of the protecting groups is carried out in accordance with known methods. For example, the triphenylmethyl group is customarily removed by hydrolysis, especially in the presence of an acid, or by hydrogenolysis in the presence of a hydrogenation catalyst; 4-nitrobenzyl is removed, for example, by hydrogenolysis in the presence of a hydrogenation catalyst; methoxy- or ethoxy-methyl is removed, for example, by treatment with a tri-lower alkyl-, such as triethyl- or tributyl-tin bromide; methylthiomethyl is removed, for example, by treatment with trifluoroacetic acid; silyl radicals are removed, for example, by treatment with fluorides, such as tetra-lower alkyl-ammonium fluorides, for example tetrabutylammonium fluoride, or alkali metal fluorides, for example sodium fluoride; 2-cyanoethyl is removed, for example, by hydrolysis, for example with sodium hydroxide solution; 2-methoxyethoxymethyl is removed, for example, by hydrolysis, for example with hydrochloric acid; and benzyloxymethyl and phenacyl are removed, for example, by hydrogenolysis in the presence of a hydrogenation catalyst.

A tetrazole protecting group, which is preferably introduced by conversion of a similarly protected amide to the corresponding N-substituted tetrazole, is e.g. cyanoethyl, p-nitrophenylethyl, lower alkoxycarbonylethyl, phenylsulfonylethyl and the like. Such tetrazole protecting groups can be removed by a retro-Michael deblocking reaction with a base such as DBN (1 ,5-diazabicyclo[4.3.0]non-5-ene), an amidine, an alkali metal carbonate or alkoxide, e.g. potassium carbonate, potassium t-butoxide, sodium methoxide in an inert solvent.

An amino protecting group is preferably t-butoxycarbonyl or benzyloxycarbonyl.

A sulfhydryl protecting group is preferably lower alkanoyl, e.g. acetyl. The preparation of compounds of the invention according to process (a) involving the condensation of an amine of formula IV with the acid of formula V or a functional reactive derivative thereof, is carried out by methodology well-known for peptide synthesis.

The condensation according to process (a) of a compound of formula IV with a free carboxylic acid of formula V is carried out advantageously in the presence of a condensing agent such as dicyclohexylcarbodiimide or N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide, and hydroxybenzotriazole, 1 -hydroxy-7-azabenzotriazole, chlorodimethoxytriazine or benzotriazol-1 -yloxy-tris-(dimethylamino)-phosphonium hexaf luorophosphate (BOP Reagent), and triethylamine or N-methylmorpholine, in an inert polar solvent such as dimethylformamide or methylene chloride, preferably at room temperature.

The condensation of a compound of formula IV with a reactive functional derivative of an acid of formula V in the form of an acid halide, advantageously an acid chloride, or mixed anhydride, is carried out in an inert solvent such as toluene or methylene chloride, advantageously in the presence of a base, e.g. an inorganic base such as potassium carbonate or an organic base such as triethylamine, N-methylmorpholine or pyridine, preferably at room temperature.

Reactive functional derivatives of carboxylic acids of formula V are preferably acid halides (e.g. the acid chloride) and mixed anhydrides, such as the pivaloyl or isobutyl- oxycarbonyl anhydride, or activated esters such as benzotriazole, 7-azabenzotriazole or hexafluorophenyl ester.

The starting materials of formula IV can be prepared according to methods described herein and illustrated in the examples.

The preparation of a starting material of formula IV involves the acylation of an ester of the amino acid of formula X

wherein X and Y' have meaning as defined hereinabove with an appropriately N-protected cyclic amino acid (or a reactive functional derivative) of formula XI

wherein A has meaning as defined hereinabove and R₅ is a labile amino protecting group, e.g. t-butoxycarbonyl, to obtain the corresponding N-protected compound of formula IV.

The condensation of a compound of formula X with a compound of formula XI is carried out by methodology well known in peptide synthesis, e.g. as described above for the condensation of a compound of formula IV with a compound of formula V. The N-protecting group is removed according to methods well-known in the art, e.g. the t-butoxycarbonyl is removed with anhydrous acid such as trifluoroacetic acid.

The starting amino acids and esters of compounds of formula X and XI are either known in the art or if new can be prepared according to methods well-known in the art, e.g. from the corresponding aldehyde or ketone. The amino acids of formula X are preferably obtained as the -S- enantiomers.

The starting materials of formula V are known or if new may be prepared according to conventional methods. The starting materials are prepared e.g. from the corresponding racemic or optically active oc-amino acids, by conversion thereof to the α-bromo derivative followed by displacement thereof with the appropriate thio acids or optionally substituted benzylthiol, under basic conditions, for example as illustrated in European Patent application No. 524,553 published January 27, 1993. S-Debenzylation of the resulting final products is carried out by reductive cleavage, e.g. with sodium in ammonia. S-Deacylation is carried out by e.g. base catalyzed hydrolysis with dilute aqueous sodium hydroxide or lithium hydroxide.

The preparation of the compounds of the invention according to process (b) involving the condensation of an acid of formula VI with a compound of formula VII is carried out in a similar fashion to process (a). Similarly the starting materials of formula VI are prepared by condensation of an acid of formula V with an ester corresponding to cyclic amino acids of formula XI (R₇ being hydrogen) under conditions similar to those described above, followed by removal of the carboxyl or tetrazolyl protecting group.

The preparation of the compounds of the invention according to process (c) involving the displacement of a leaving group Z in a compound of formula VIII with a sulfhydryl derivative Ri'-SH is carried out according to methods well-known in the art.

A reactive esterified hydroxyl group, represented by Z, is a hydroxyl group esterified by a strong inorganic or organic acid. Corresponding Z groups are in particular halo, for example chloro, bromo or iodo, also sulfonyloxy groups, such as lower alkyl- or arylsulfonyloxy groups, for example (methane-, ethane-, benzene- or toluene-) sulfonyloxy groups, also the trifluoromethylsulfonyloxy group.

The displacement is carried out in an inert solvent, such as dimethylformamide, methylene chloride or THF in the presence of a base such as potassium carbonate, triethylamine, diisopropylethylamine, N-methylmorpholine, and the like at room or elevated temperatures.

Similarly, the starting materials of formula VIII can be prepared by reacting the amide derivative of formula IV with an acid of the formula

Z-C-COOH (XII)

/

R* \

wherein R₂ and R₃ and Z have meaning as defined above, under conditions described for process (a). Acids of formula XII e.g. wherein Z is bromo, can be prepared from the corresponding α-aminoacids according to methods well known in the art. Optionally active acids of formula XII can be obtained from optically active α-aminoacids as illustrated herein.

The following sequence of reactions is illustrative of process (c).

The compounds of the invention wherein Y represents 1 H-5-tetrazolyl are similarly prepared, but starting with a tetrazole derivative of formula X" (instead of amino acid ester X')

wherein R_p is a tetrazolyl protecting group (such as 2-cyanoethyl).

The tetrazole starting materials of formula X" are prepared from the corresponding N- acyl amino acids by first converting such to the N-R_p-substituted amides. The resulting amides are then treated under conditions known in the art for tetrazole ring formation, e.g. under conditions described in Tetrahedron Letters 1979. 491 and J. Org. Chem. 56, 2395 (1991 ), e.g. with trimethylsilyl azide in the presence of diisopropyl azodicarboxylate and triphenylphosphine. Removal of the N- acyl group leads to the starting materials of formula X".

In the above illustrated sequence of reactions for process (c) the tetrazole protecting groups is preferably removed after formation of the bromo intermediate and prior to reaction with potassium thioacetate.

As to the amino acid starting materials of formula X, such can be prepared by one of the following methods (as illustrated for compounds wherein X is O):

Method A:

b. Pd(OAc)₂ TBACI / DMF

AcNH X COOMe

Method B:

The oxidative cyclization to form the dibenzofuran tricyclic ring system is carried out according to the general procedure described in J. Org. Chem.40, 1365 (1975).

The enzymatic resolution of the N-acyl amino acid ester can be performed by hydrolysis with an esterase, e.g. alcalase (substilisin) as illustrated below.

NaHCO,

Certain compounds of the invention and intermediates can be converted to each other according to general reactions well known in the art.

The free mercaptans may be converted to the S-acyl derivatives by reaction with a reactive derivative of a carboxylic acid (corresponding to Ri being acyl in formula I), such as an acid anhydride or said chloride, preferably in the presence of cobalt chloride (CoCI₂) in an inert solvent such as acetonitrile or methylene chloride.

Free alcohols can be converted to the corresponding acyl derivatives e.g. by reaction with a corresponding acid chloride in the presence of a base, such as triethylamine.

The free mercaptans, wherein Ri represents hydrogen, may be oxidized to the corresponding disulfides, e.g. by air oxidation or with the use of mild oxidizing agents such as iodine in alcoholic solution. Conversely, disulfides may be reduced to the corresponding mercaptans, e.g. with reducing agents such as sodium borohydride, zinc and acetic acid or tributylphosphine.

Carboxylic acid esters may be prepared from a carboxylic acid by condensation with e.g. the halide corresponding to the esterifying alcohol in the presence of a base, or with an excess of the alcohol, in the presence of an acid catalyst, according to methods well-known in the art.

Carboxylic acid esters and S-acyl derivatives may be hydrolyzed, e.g. with aqueous alkali such as alkali metal carbonates or hydroxides.

In case mixtures of stereoisomers (e.g. diastereomers) are obtained, these can be separated by known procedures such as fractional crystallization and chromatography (e.g. thin layer, column, flash chromatography). Racemic free acids can be resolved into the optical antipodes by fractional crystallization of d- or I- (α-methylbenzylamine, cinchonidine, cinchonine, quinine, quinidine, dehydroabietylamine, brucine or strychnine) salts and the like. Racemic products, if not diastereoisomers, can first be converted to diastereoisomers with optically active reagents (such as optically active alcohols to form esters) which can then be separated as described above, and e.g. hydrolyzed to the individual enantiomer. Racemic products can also be resolved by chiral chromatography, e.g. high pressure liquid chromatography using a chiral adsorbent; also by enzymatic resolution, e.g. of esters with alcalase.

The above-mentioned reactions are carried out according to standard methods, in the presence or absence of diluents, preferably such as are inert to the reagents and are solvents thereof, of catalysts, alkaline or acidic condensing or said other agents respectively and/or inert atmospheres, at low temperatures, room temperature or elevated temperatures, preferably near the boiling point of the solvents used, at atmospheric or superatmospheric pressure.

The invention further includes any variant of said processes, in which an intermediate product obtainable at any stage of the process is used as a starting material and any remaining steps are carried out, or the process is discontinued at any stage thereof, or in which the starting materials are formed under the reaction conditions, or in which the reaction components are used in the form of their salts or optically pure antipodes. Mainly those starting materials should be used in said reactions, that lead to the formation of those compounds indicated above as being preferred.

The present invention additionally relates to the use in mammals of the compounds of the invention and their pharmaceutically acceptable, non-toxic acid addition salts, or pharmaceutical compositions thereof, as medicaments, for inhibiting endothelin converting enzyme, and e.g. for the treatment of endothelin dependent disorders such as those mentioned hereinabove, e.g. cardiovascular disorders.

The present invention also relates to the use of the compounds of the invention for the preparation of medicaments, e.g. pharmaceutical compositions, especially pharmaceutical compositions having endothelin converting enzyme inhibiting activity. The pharmaceutical compositions according to the invention are those suitable for enteral, such as oral or rectal, transdermal and parenteral administration to mammals, including man, for the treatment of endothelin dependent disorders, comprising an effective amount of a pharmacologically active compound of the invention or a pharmaceutically acceptable salt thereof, alone or in combination with one or more pharmaceutically acceptable carriers.

The pharmacologically active compounds of the invention are useful in the manufacture of pharmaceutical compositions comprising an effective amount thereof in conjunction or admixture with excipients or carriers suitable for either enteral or parenteral application. Preferred are tablets and gelatin capsules comprising the active ingredient together with a) diluents, e.g. lactose, dextrose, sucrose, mannitol, sorbitol, cellulose and/or glycine; b) lubricants, e.g. silica, talcum, stearic acid, its magnesium or calcium salts and/or polyethyleneglycol; for tablets also c) binders, e.g. magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and/or polyvinylpyrrolidone; if desired, d) disintegrants, e.g. starches, agar, alginic acid or its sodium salt, or effervescent mixtures; and/or e) absorbents, colorants, flavors and sweeteners. Injectable compositions are preferably aqueous isotonic solutions or suspensions, and suppositories are advantageously prepared from fatty emulsions or suspensions. Said compositions may be sterilized and/or contain adjuvants, such as preserving, stabilizing, wetting or emulsifying agents, solution promoters, salts for regulating the osmotic pressure and/or buffers. In addition, the compositions may also contain other therapeutically valuable substances. Said compositions are prepared according to conventional mixing, granulating or coating methods, respectively, and contain about 0.1 to 75%, preferably about 1 to 50%, of the active ingredient.

Suitable formulations for transdermal application include an effective amount of a compound of the invention with carrier. Advantageous carriers include absorbable pharmacologically acceptable solvents to assist passage through the skin of the host. Characteristically, transdermal devices are in the form of a bandage comprising a backing member, a reservoir containing the compound, optionally with carriers, optionally a rate controlling barrier to deliver the compound to the skin of the host at a controlled and predetermined rate over a prolonged period of time, and means to secure the device to the skin. A unit dosage for a mammal of about 50 to 70 kg may contain between about 5 and 100 mg of the active ingredient. The dosage of active compound is dependent on the species of warm-blooded animal (mammal), the body weight, age and individual condition, and on the form of administration.

The following examples are intended to illustrate the invention and are not to be construed as being limitations thereof. Temperatures are given in degrees Centigrade. If not mentioned otherwise, all evaporations are performed under reduced pressure, preferably between about 15 and 100 mm Hg. Optical rotations are measured at room temperature at 589 nm (D line of sodium) or other wavelengths as specified in the examples.

The prefixes R and S are used to indicate the absolute configuration at each asymmetric center. L-Amino acids as used herein correspond to the S-configuration.

Abbreviations used are those standard in the art, e.g. "BOP" reagent is the abbreviation for benzotriazol -1 -yloxy-tris (dimethylamino) phosphonium hexafluorophosphate, HOAT is the abbreviation for 1-hydroxy-7-azabenzotriazole, HOBT is the abbreviation for 1 -hydroxybenzotriazole, EDCI is the abbreviation for 1-ethyl-3-(3- dimethylaminopropyl) carbodiimide hydrochloride.

EXAMPLES

(A) Preparation of (SVdibenzofuran-3-ylalanine - Method A

To a suspension of 3-aminodibenzofuran (15g, 81.9 mmol) in H₂O (80 mL) is added concentrated HCI (18.6 mL, 0.2 mol) and the mixture is cooled to 0 ^SC (internal temperature). A solution of NaNO₂ (6.3 g, 91.2 mmol) in H₂O (30 mL) is added dropwise over 10 minutes and stirred at 0⁹ C for 15 minutes. The cold solution is transferred into an addition funnel and added dropwise over 15 minutes to an ice-cold mechanically stirred solution of Kl (20.4 g, 123 mmol) in H₂O (200 mL). To the viscous mixture is added ice (200 g) and Et₂O (50 mL). Stirring is continued at 0⁹C for 90 minutes, then the reaction mixture is allowed to warm up slowly to room temperature and stirred for 14 hours under nitrogen. The brown mixture is heated to 50⁹ C for 35 minutes then cooled to room temperature. The precipitate is filtered off and washed with H₂O. The dark brown solid is dissolved in a mixture of EtOAc and toluene (95:5, 700 mL), washed successively with 0.1 M Na₂S₂O₃ solution and brine, dried (Na₂SO₄), and treated with charcoal. Filtration through a bed of silica gel and concentration in vacuo gives a solid residue which is dissolved in boiling isopropyl ethyl ether (400 mL). The solution is filtered through glass micro fiber paper and cooled to -5⁹C for 18 hours . The resulting solid is filtered off and dried under high vacuum at 50⁹C for 2 hours, affording 3-iododibenzofuran, 61.4%). mp 139-141 ⁹C. IR(CH₂CI₂) 1588, 1597, 1229, 1193, 877 cm^"1. ¹H NMR (CDCI₃, 300 MHz) δ 7.35 (1 H, m), 7.42-7.60 (2 H, m), 7.66-7.72 (2 H, m), 7.83-7.98 (2 H, m). MS (DCI/ CH₄) m/z 295 (M+1).

To a stirred solution of 3-iododibenzofuran (10.0 g, 34 mmol) in anhydrous DMF (260 mL) is added methyl 2-acetamidoacrylate (6.8 g, 47.26 mmol), followed by Pd(OAc)₂ (0.30 g, 1.36 mmol), n-Bu₄NCI (9.45 g, 34 mmol), and triethylamine (4.7 mL, 34 mmol). The solution is placed in a pre-heated oil bath (85 ⁹C) and stirred for 15 hours The reaction mixture is cooled to room temperature and poured into ice H₂O (300 L) and 1 N HCI (60 mL) and stirred for 30 minutes. The precipitate is collected and rinsed with H₂O. The filtrate is then taken up in Et₂O, stirred and filtered. The solid is dried under high vacuum at 50⁹C for 1 hour and at room temperature overnight to give 2-acetylamino-3-dibenzofuran-3-yl-acrylic acid methyl ester, mp 217-218⁹C. ¹H NMR (DMSO-d₆: 300 MHz) δ 2.05 (3 H, s), 3.34 (1 H, s), 3.73 (3 H, s), 7.34 (1 H, s), 7.42 (1 H, t), 7.53 - 7.58 (1 H, m), 7.66 - 7.73 (2 H, m), 8.00 (1 H, s), 8.17 (2 H, d) ; IR (KBr) 1715, 1656, 1523, 1243, 1204, 755 cm^"1.

To a solution of 2-acetylamino-3-dibenzofuran-3-yl acrylic acid methyl ester (8.72 g, 28.19 mmol) in MeOH (350 mL) and toluene (310 mL) is added 10% Pd/C (5.5 g) under N₂ atmosphere. The reaction mixture is shaken under 50 psi H₂ overnight. The reaction mixture is filtered through a bed of celite and rinsed with 25% MeOH/ CH₂CI₂ and concentrated in vacuo. The residue is triturated with a 50/50 mixture of Et₂O and hexanes. The light tan solid is filtered and dried under high vacuum at 50⁹C for 2 hours to give 2- acetylamino-3-dibenzofuran-3-yl-propionic acid methyl ester; ¹H NMR (CDCI₃, 250 MHz) δ 1.99 (3 H, s), 3.2-3.35 (2 H, m), 3.73 (3 H, s), 4.90 - 4.98 (1 H, m), 5.98 - 6.02 (1 H, m), 7.04-7.08 (1 H, m), 7.28-7.35 (2 H, m), 7.40-7.46 (1 H, m), 7.52-7.55 (1 H, m), 7.83 - 7.91 (2 H, m).

To a mechanically stirred solution of 2-acetylamino-3-dibenzofuran-3-yl propionic acid methyl ester (11.5 g, 30.50 mmol) in MeCN (385 mL) and 0.2 M NaHCO₃ (510 mL) is added alcalase (1.2 mL; Novo Nordisk Bioindustrials Inc.) and the reaction is monitered by HPLC, until 50% conversion (YMC-Pack ODS-A Cι₈, 70% MeCN/ 30% H₃PO₄). CH₂CI₂ is added to the reaction mixture after 2 hours, the mixture is stirred for 10 minutes, and the layers are separated. The organic layer is washed once with H₂O. The combined aqueous layers are washed several times with CH₂CI₂. The combined organic layers are dried (MgSO₄), filtered and concentrated to give (R)-2-acetylamino-3-dibenzofuran-3-y! propionic acid methyl ester. The aqueous layer is acidified (pH=1 ) with 1 N HCI, the precipitate is filtered and dried under high vacuum at 50⁹C for 4 hours to give (S)-2-acetylamino-3-dibenzofuran-3-yl propionic acid; [α]²⁵ _D = +25.102, 8.55 mg/mL DMSO. ¹H NMR (DMSO - d₆, 300 MHz) δ 1.77 (3 H, s), 2.96-3.04 (1 H, m), 3.22 (1 H, dd), 4.46-4.54 (1 H, m), 7.27 (1 H, d), 7.35 - 7.40 (1 H, m), 7.46 - 7.52 (1 H, m), 7.56 (1 H, s), 7.67 (1 H, d), 8.03 (1 H, d), 8.09 (1 H, d), 8.26 (1 H, d).

A mechanically stirred mixture of (S)-2-acetylamino-3-dibenzofuran-3-yl propionic acid (6.27 g, 21.09 mmol) in concentrated HCI (211 mL) and AcOH (71 mL) is heated to 115⁹C overnight. The reaction mixture (white slurry) is cooled to room temperature, then cooled in an ice bath, the white solid is filtered and dried under high vacuum at 55 ⁹C for 4 hours to give (S)-2-amino-3-dibenzofuran-3-yl-propionic acid hydrochloride; [α] ⁵ _D = +15.09; 8.5 mg/mL DMSO + 2% TFA. ¹H NMR (DMSO-d₆-TFA, 300 MHz) δ 3.23-3.37 (2 H, m), 4.3 - 4.31 (1 H, br m), 7.29-7.32 (1 H, m), 7.36-7.42 (1 H, m), 7.48-7.53 (1 H, m), 7.62 (1 H, s), 7.69 (1 H, d), 8.1-8.13 (2 H, m), 8.35-8.5 (1 H, br m).

(B) Preparation of (S)-dibenzofuran-3-ylalanine Synthesis - Method B

To a mechanically stirred solution of 2-acetylamino-2-(3-phenoxybenzyl) malonic acid diethyl ester (EP application 284,632; 363.86 g, 0.91 mol) in DMF (1.8 L) is added LiBr (79 g, 0.91 mol) and H₂O (33 mL, 1.82 mol). The solution is heated to 138 ^SC (internal temperature) and stirred for 16 hours. The heterogenous mixture is cooled to 60⁹C and concentrated in vacuo at 70 ^eC. The residue is partioned between ether (1 L), EtOAc (500 mL) and ice cold H₂O (500 mL). The insoluble material is filtered off. The organic layer is washed with H₂O (2X500 mL) and brine (300 mL), dried (MgSO₄), and filtered. The solvent is evaporated in vacuo and the residue is crystallized from f-butyl methyl ether and hexane. The solid is filtered and dried under high vacuum at 40⁹C to give 2-acetylamino-3-(3- phenoxybenzyl) propionic acid ethyl ester as a white crystalline solid; ¹H NMR (CDCI₃, 300 MHz) δ 1.22 (3 H, t), 1.97 (3 H, s), 3.01-3.18 (2 H, m), 4.02-4.21 (2 H, m), 4.84 (1 H, m), 6.0 (1 H, d), 6.71 (1 H, m), 6.84 (1 H, d), 6.92 (1 H, dd), 6.94-7.02 (2 H, m), 7.11 (1 H, m), 7.25 (1 H, m), 7.30-7.39 (2 H, m).

To a stirred solution of 2-acetylamino-3-(3-phenoxybenzyl) propionic acid ethyl ester (50 g, 153 mmol) in AcOH (500 mL) is added Pd(OAc)₂ (51.4 g, 230 mmol). The reaction mixture is heated to reflux temperature (130⁹C internal) for 19 hours, then cooled to room temperature, and filtered through Celite. The filtrate is concentrated under reduced pressure, and the residue is dissolved in CH₂CI₂ (50 mL) and diluted with ether (500 mL). The insoluble material is removed by filtration and washed with ether. The filtrate is then concentrated in vacuo. Acetic acid is removed by co-evaporation with toluene. The oily residue is purified by flash chromatography on silica gel (900 g, gradient elution with 40- 50% EtOAc/hexane). The pure fractions, according to HPLC analysis (Waters symmetry C8 column; eluent: MeCN - pH 2.4 buffer; isocratic 50-50; flowrate: 1.5 mL/min; Rt = 10 min) are combined and concentrated under reduced pressure. The residue is recrystallized from EtOAc-hexane and dried at 40⁹C under high vacuum to give 2-acetylamino-3- dibenzofuran-3-yl propionic acid ethyl ester contaminated with a minor amount of regioisomeric 2-acetylamino-3-dibenzofuran-1-yl propionic acid ethyl ester.

To a stirred solution of 2-acetylamino-3-dibenzofuran-3-yl propionic acid ethyl ester (25 g, 77 mmol) (containing a minor amount of 2-acetylamino-3-dibenzofuran-1-yl propionic acid ethyl ester which is not hydrolyzed by alcalase) in MeCN (990 mL) is added 0.2M NaHCO₃ (1.3 L), followed by alcalase (2.6 mL; Novo Nordisk Bioindustrials Inc.). After 3 hours, EtOAc (1.75 L) and H₂O (750 mL) are added. The organic layer is washed with H₂O (500 mL) and set aside for subsequent optional epimerization (see below). The aqueous layer is acidified with 2N HCI (140 mL) under stirring. The white precipitate is filtered, washed with H₂O (2X500 mL), and dried under high vacuum at 60⁹C for 3 hours and at room temperature for 15 hours to give (S)-2-acetylamino-3-dibenzofuran-3-yl propionic acid.

The above organic layer containing the remaining (R)-unhydrolyzed ester is dried (Na₂SO₄), filtered, and concentrated in vacuo. The resulting (R)-2-acetylamino-3-dibenzofuran-3-yl- propionic acid ethyl ester (containing some racemic regioisomer 2-acetylamino-3- dibenzofuran-1 -yl propionic acid ethyl ester) (53 g, 160 mmol) is dried by azeotropic removal of H₂O with toluene-ethanol, then dissolved in dry ethanol (450 mL). A solution of sodium ethoxide, prepared from sodium (0.2 g, 8 mmol) and ethanol (50 mL), is added at room temperature. The reaction mixture is stirred at 60 ^eC for 3 hours, then at room temperature for 12 hours. The solution is saturated with HCI gas and stirred for 1.5 hours at room temperature. Ethanol is removed by evaporation under reduced pressure. The residue is partitioned between EtOAc (500 mL) and H₂O (500 mL). The organic layer is separated, washed successively with saturated NaHCO₃ (250 mL), H₂O (250 mL), and brine (250 mL), then dried (Na₂SO₄), and filtered. The solution is concentrated in vacuo and the residue is recrystallized from EtOAc-hexane to give racemic 2-acetylamino-3-dibenzofuran- 3-yl propionic acid ethyl ester, which can again be used as starting material in the above mentioned enzymatic resolution.

HCI (g) is bubbled through a stirred solution of (S)-2-amino-3-dibenzofuran-3-yl propionic acid hydrochloride (2.58g, 8.84 mmol) in MeOH (90 mL) at 0 ^eC, until the solution is saturated. The solution is warmed to room temperature and then placed in a pre-heated oil bath (70⁹C) for 2 hours, cooled to room temperature and stirred overnight. Some solid precipitates overnight, the mixture is concentrated in vacuo and the residue is treated with Et₂O, filtered and dried under high vacuum at 60⁹C for 1 hour to give (S)-2-amino-3- dibenzofuran-3-yl-propionic acid methyl ester hydrochloride (also called (S)-dibenzofuran-3- yl-alanine methyl ester hydrochloride).

[α]²⁵ _D = +13.982; 8.35 mg/mL MeOH. ¹H NMR (CD₃OD, 300 MHz) δ 3.3 - 3.37 (1 H, m), 3.46 (1 H, dd,), 3.83 (3 H, s), 4.43 (1 H, dd), 7.26 - 7.29 (1 H, m), 7.34 - 7.47 (1 H, m), 7.49 - 7.6 (3 H, m), 8.02 - 8.05 (2 H, m).

(C) Preparation of α-bromocarboxylic acids

(a) 5.00 g (38.1 mmol) of L-norleucine and 22.7 g (191 mmol) of potassium bromide are dissolved in 50 mL of water at room temperature. 10.8 mL (95.5 mmol) of aqueous 48 % hydrobromic acid is then added and the mixture is cooled to -12 °C in an ice/NaCI bath. Next, the flask is equipped with an addition funnel containing 3.16 g (45.7 mmol) of sodium nitrite dissolved in 20 mL of water. The sodium nitrite solution is allowed to drip into the reaction mixture over the course of 30 minutes. After the addition of sodium nitrite is complete, the mixture is stirred for an additional 45 minutes, transferred to a separatory funnel, and diluted with ethyl acetate. The aqueous phase is separated and extracted two times with ethyl acetate. The combined ethyl acetate phases are washed three times with aqueous sodium bisulfite (removing the yellow color), dried over sodium sulfate, and concentrated to dryness to afford a clear colorless oil, which is dried under high vacuum to give 2S-bromohexanoic acid. ¹H NMR (250 MHz, CDCI₃) δ 10.4 (s, 1 H), 4.24 (t, 1 H), 1.92- 2.17 (m, 2 H), 1.32-1.55 (m, 4 H, 0.93 (t, 3 H).

Similarly prepared are:

(b) 2S-bromo-3R-methylpentanoic acid;¹H NMR (250 MHz, CDCI₃) δ 10.88 (s, 1 H), 4.29 (d, 1 H), 1.86-2.09 (m, 0.5 H), 1.43-1.68 (m, 0.5 H), 1.24-1.43 (m, 2 H), 1.07 (d, 3 H), 0.95 (t, 3 H).

(c) 2S-bromo-3S-methylpentanoic acid; ¹H NMR (250 MHz, CDCI₃) δ 10.35 (s, 1 H), 4.12 (d, 1 H), 1.98-2.10 (m, 0.5 H), 1.67-1.83 (m, 0.5 H), 1.24-1.48 (m, 2 H), 1.05 (d, 3 H), 0.92 (t, 3 H).

(d) 2R-bromo-3R-methylpentanoic acid; ¹H NMR (300 MHz, CDCI₃) δ 10.65 (s, 1 H), 4.11 (d, 1 H), 1.99-2.10 (m, 0.5 H), 1.67-1.80 (m, 0.5 H), 1.22-1.44 (m, 2 H), 1.04 (d, 3 H), 0.91 (t, 3 H).

(e) 2R-bromo-3S-methylpentanoic acid; ¹H NMR (300 MHz, CDCI₃) δ 10.15 (s, 1 H), 4.27 (d, 1 H), 1.90-2.06 (m, 0.5 H), 1.43-1.54 (m, 0.5 H), 1.22-1.38 (m, 2 H), 1.03 (d, 3 H), 0.93 (t, 3 H).

(f) 2R-bromo-4-methylpentanoic acid; ¹H NMR (250 MHz, CDCI₃) δ 9.81 (s, 1 H), 4.29 (d, 1 H), 1.92 (t, 2 H), 1.72-1.89 (m, 1 H), 0.97 (d, 3 H), 0.92 (d, 3 H).

(g) 2R-bromo-4-methylthiobutanoic acid; ¹H NMR (250 MHz, CDCI₃) δ 9.56 (s, 1 H), 4.50 (dd, 1 H), 2.57-2.76 (m, 2 H), 2.22-2.43 (m, 2 H), 2.11 (s, 3 H).

(h) 2R-bromopentanoic acid; H NMR (250 MHz, CDCI₃) δ 10.06 (s, 1 H), 4.25 (dd, 1 H), 1.91-2.15 (m, 2 H), 1.34-1.62 (m, 2 H), 0.97 (t, 3 H). Example 1

2-{[1-(2S-(acetylthio-3-R-methyl-pentanoylamino)-cyclopentanecarbonyl]-amino}-3- dibenzofuran-3-yl-propionic acid methyl ester:

90 mg (0.161 mmol) of 2-{[1-(2R-bromo-3R-methyl-pentanoylamino)-cyclopentanecarbonyl]- amino}-3-dibenzofuran-3-yl-propionic acid methyl ester is dissolved in methanol, and treated with 55 mg (0.483 mmol) of potassium thioacetate for 7 hours. The crude mixture is diluted with ether, washed with brine, dried over sodium sulfate, and concentrated to dryness to afford a yellow solid. The product is purified by chromatography on silica gel with 30% ethyl acetate/hexane to produce 2-{[1-(S-(acetylthio-3-R-methyl-pentanoylamino)- cyclopentanecarbonyl]-amino}-3-dibenzofuran-3-yl-propionic acid methyl ester as a white solid with R,= 0.15 (30 % ethyl acetate/hexane). ¹H NMR (250 MHz, CDCI₃) δ 7.88 (d, 1 H); 7.86 (d, 1 H); 7.49 (d, 1 H); 7.42 (t, 1 H); 7.25-7.40 (m, 2 H); 7.13-7.20 (m, 2 H); 6.29 (s, 1 H); 4.80 (dd, 1 H); 3.64-3.72 (m, 4 H); 3.30 (t, 2 H); 2.27 (s, 3 H); 2.10-2.32 (m, 2 H); 1.84- 2.09 (m, 2 H); 1.51-1.82 (m, 6 H); 1.00-1.15 (m, 1 H); 0.90 (t, 3 H); 0.80 (d, 3 H).

The starting material is prepared as follows:

570 mg (1.86 mmol) of (S)-dibenzofuran-3-ylalanine methyl ester hydrochloride is suspended in 5 mL of chloroform in a vial. Ammonia gas is bubbled through the solution for 15 minutes at room temperature. The vial is then centrifuged to settle the ammonium chloride precipitate to the bottom of the vessel. The cloudy supernatant is extracted with ethyl acetate, and the organic phase is washed two times with aqueous saturated sodium bicarbonate, dried over sodium sulfate, concentrated to dryness, and the oil is dried under high vacuum to afford (S)-2-amino-3-dibenzofuran-3-yl-propionic acid methyl ester as an off white solid. ¹ NMR (300 MHz, DMSO) δ 8.09 (d, 1 H); 8.01 (d, 1 H); 7.67 (d, 1 H); 7.64- 7.53 (m, 2 H); 7.35 (t, 1 H); 7.20 (d, 1 H); 3.65 (t, 1 H); 3.58 (s, 3 H); 2.99 (dq, 2 H); 1.91 (s, 2 H). 493 mg (1.83 mmol) of (S)-2-Amino-3-dibenzofuran-3-yl-propionic acid methyl ester ; 504 mg (2.20 mmol) of N-.-Boc cycloleucine (U.S. patent 5,506,244) and 274 mg (2.01 mmol) of HOAt are suspended in 15 mL of dichloromethane at room temperature. Next 382 μL (2.75 mmol) of triethylamine is added with stirring for 5 minutes to give a clear brown solution. 702 mg (3.66 mmol) of EDCI coupling agent is then added and the clear brown mixture is stirred for 4 hours. The crude mixture is transferred to a separatory funnel containing 100 mL of ethyl acetate and 75 mL 1 M hydrochloric acid. After separating the phases, the organic phase is washed two times with 1 M hydrochloric acid, two times with saturated sodium bicarbonate, two times with brine, dried over sodium sulfate, and concentrated to dryness to afford a white solid. Drying under high vacuum yields 2-[(1 -tert- butoxycarbonylamino-cyclopentanecarbonyl)-amino]-3-dibenzofuran-3-yl-propionic acid methyl ester as a white powder. ¹H NMR (300 MHz, CDCI₃) δ 7.91 (d, 1 H); 7.85 (d, 1 H); 7.55 (d, 1 H); 7.41 (t, 1 H); 7.38-7.47 (m, 3 H); 7.11 (d, 1 H); 4.92 (dd, 1 H); 4.35 (broad s, 1 H); 3.70 (s, 3 H); 3.30 (d, 2 H); 2.08-2.38 (m, 2 H); 1.63-1.95 (m, 6 H); 1.38 (s, 9 H). Mass spec, m/z calcd for C₂₇H₃₂N₂O₆480.6, found 481.

1.065 g (1.83 mmol) of 2-[(1-terf-butoxycarbonylamino-cyclopentanecarbonyl)-amino]-3- dibenzofuran-3-yl-propionic acid methyl ester is dissolved in 20 mL of 3:1 dichloromethane/ether at room temperature. With rapid stirring, hydrogen chloride gas is bubbled through the solution for 10 minutes causing the solution to turn cloudy white. The solution is stirred at room temperature for three hours, concentrated to dryness and then placed on the high vacuum for 30 minutes to afford (S)-2-[(1 -aminocyclopentanecarbonyl)- amino]-3-dibenzofuran-3-yl-propionic acid methyl ester hydrochloride as a light yellow solid. ¹H NMR (250 MHz, DMSO) δ 8.68 (d, 1 H); 7.98-8.13 (m, 2 H); 7.66 (d, 1 H); 7.61 (s, 1 H); 7.50 (t, 1 H); 7.36 (t, 1 H); 7.28 (d, 1 H); 4.69 (dd, 1 H); 3.69 (s, 3 H); 3.17 (t, 2 H); 1.95-2.18 (m, 2 H); 1.68-1.95 (m, 6 H). Mass spec, m/z calcd for C₂₂H₂ N₂O₄ 380.5, found 381.

321 mg (0.770 mmol) of (S)-2-[(1-aminocyclopentanecarbonyl)-amino]-3-dibenzofuran-3-yl- propionic acid methyl ester hydrochloride, 165 mg (0.847 mmol) of 2R-bromo-3R- methylpentanoic acid, arid 115 mg (0.847 mmol) of HOAt are suspended in 7 mL of dichloromethane, and treated with 161 μL (1.15 mmol) of triethylamine causing the solution to turn clear. After stirring for five minutes, 295 mg (1.54 mmol) of EDCI coupling agent is added and the reaction mixture is stirred for five hours. The reaction mixture is diluted with ether and washed successively three times with 1 M hydrochloric acid, two times with saturated sodium bicarbonate, two times with brine, dried over sodium sulfate, and concentrated to dryness to afford a yellow residue. The residue is dissolved in dichloromethane and hexane is added to induce crystallizaion. Evaporation to dryness affords 2-{[1-(2R-bromo-3R-methyl-pentanoylamino)-cyclopentanecarbonyl]-amino}-3- dibenzofuran-3-yl-propionic acid methyl ester as a light yellow solid. ¹H NMR (300 MHz, CDCI₃) δ 7.92 (d, 1 H); 7.85 (d, 1 H); 7.54 (d, 1 H); 7.45 (t, 1 H); 7.27-7.41 (m, 2 H); 7.15 (d, 1 H) 6.97 (d, 1 H); 6.59 (s, 1 H); 4.19 (dd, 1 H); 4.20 (d, 1 H); 3.75 (s, 3 H); 3.35 (dd, 2 H); 2.14-2.40 (m, 2 H); 1.87-2.21 (m, 2 H); 1.68-1.85 (m, 4 H); 0.99-1.42 (m, 3 H); 0.92 (d, 3 H); 0.84 (t, 3 H). MS: m/z 557.

Example 2

(S)-3-Dibenzofuran-3-yl-2-{[1-(2S-mercapto-3R-methyl-pentanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid:

55 mg (0.0995 mmol) of 2-{[1-(2S-(acetylthio-3-R-methyl-pentanoylamino)- cyclopentanecarbonyl]-amino}-3-dibenzofuran-3-yl-propionic acid methyl ester is dissolved in methanol, and treated with 1 N sodium hydrioxide for two hours at room temperature. The mixture is then acidified to pH=1 with 1 M hydrochloric acid, the product extracted into ethyl acetate, the ethyl acetate solution is washed with brine, dried over sodium sulfate, and concentrated to dryness to afford a clear, colorless oil. The product is solidified by dissolving in dichloromethane and adding hexane and dried under high vacuum to afford (S)-3-dibenzofuran-3-yl-2-{[1-(2S-mercapto-3R-methyl-pentanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid: mp 191-195 °C; ¹H NMR (300 MHz, DMSO) δ 12.89 (s, 1 H); 8.09 (d, 2 H); 7.99 (d, 1 H); 7.68 (d, 1 H); 7.42-7.54 (m, 2 H); 7.38 (t, 3 H); 7.30 (d, 1 H); 7.26 (d, 1 H); 4.55 (dd, 1 H); 3.07-3.18 (m, 3 H); 2.41 (d, 1 H); 1.89-2.08 (m, 2 H); 1.77-1.88 (m, 2 H); 1.55-1.76 (m, 5 H); 1.20-1.40 (m, 1 H); 0.98-1.18 (m, 2 H); 0.87 (d, 3 H); 0.78 (t, 3 H).; Mass spec, m/z calcd for C₂₇H₃₂N₂SO₅ 496.7, found 497. Example 3

Prepared according to the general procedure described in Examples 1 and 2 are:

(a) (S)-3-Dibenzofuran-3-yl-2-{[1-(2S-mercapto-3S-methylpentanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid:

mp 202-205 °C; ¹H NMR (300 MHz, DMSO-d₆) δ 12.86 (s, 1 H); 8.09 (d, 2 H); 8.00 (d, 1 H); 7.66 (d, 1 H); 7.40-7.51 (m, 2 H); 7.36 (d, 1 H); 7.30 (d, 1 H); 7.20 (d, 1 H); 4.52 (dd, 1 H); 3.07-3.32 (m, 3 H); 2.42 (d, 1 H); 1.91-2.10 (m, 2 H); 1.75-1.90 (m, 2 H); 1.50-1.70 (m, 6 H); 1.08-1.20 (m, 1 H); 0.68-0.80 (m, 6 H). IR (KBr, cm^"1) 3419, 3292, 2963, 2928, 2550, 1733, 1652, 1500, 1457, 1203, 1127, 748. Mass spec, m/z calcd for C₂₇H₃₂N₂SO₅496.7, found 497.

(b) (S)-3-Dibenzofuran-3-yl-2-{[1-(2S-mercapto-4-methylpentanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid:

mp 170-172 °C; ¹H NMR (300 MHz, DMSO-d₆) δ 12.90 (s, 1 H); 8.06-8.15 (m, 2 H); 8.00 (d, 1 H); 7.65 (d, 1 H); 7.42-7.51 (m, 2 H); 7.37 (t, 1 H); 7.32 (d, 1 H); 7.28 (d, 2 H); 4.52 (dd, 1 H); 3.36 (dd, 1 H); 3.13 (dd, 2 H); 2.61 (d, 1 H); 1.90-2.05 (m, 2 H); 1.75-1.90 (m, 2 H); 1.50- 1.65 (m, 6 H); 1.27-1.40 (m, 1 H); 0.75 (dd, 6 H). IR (KBr, cm^*1) 3499, 3410, 3301, 2957, 2871, 2550, 1733, 1673, 1651, 1509, 1457, 1427, 1203. Mass spec, m/z calcd for C₂₇H₃₂N₂SO₅496.7, found 497.

(c) (S)-3-Dibenzofuran-3-yl-2-{[1 -(2S-mercaptopentanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid:

mp 160-163 °C; ¹H NMR (300 MHz, DMSO-d₆) δ 12.89 (s, 1 H); 8.09 (d, 2 H); 8.00 (d, 1 H); 7.66 (d, 1 H); 7.40-7.52 (m, 2 H); 7.34 (dd, 2 H); 7.21 (d, 1 H); 4.51 (dd, 1 H); 3.09-3.36 (m, 3 H); 2.60 (d, 1 H); 1.89-2.05 (m, 2 H); 1.76-1.89 (m, 2 H); 1.50-1.75 (m, 5 H); 1.31-1.50 (m, 1 H); 1.14-1.30 (m, 2 H); 0.80 (t, 3 H). Mass spec, m/z calcd for C₂₆H₃₀N₂SO₅482.6, found 483.

(d) (S)-3-Dibenzofuran-3-yl-2-{[1-(2(S)-mercapto-3-methylbutanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid:

mp 182-183 °C; H NMR (250 MHz, CD₃OD) δ 7.97 (d, 1 H), 7.91 (d, 1 H), 7.54 (br d, 1 H), 7.40-7.50 (m, 2 H), 7.33 (dt, 1 H), 7.24 (dd, 1 H), 4.70-4.80 (app t, 1 H), 3.15-3.45 (ABX m, 2 H), 2.91 (d, 1 H), 2.10-2.30 (m, 2 H), 1.80-2.05 (m, 3 H), 1.65-1.75 (m, 4 H), 0.97 (d, 3 H), 0.86 (d, 3 H); IR (KBr, cm^"1) 2550, 1733, 1652, 1515, 1203, 1127, 748. Mass spec, m/z calcd for C₂₆H₃₀N₂O₅S 482.6, found 482. (e) (S)-3-Dibenzofuran-3-yl-2-({1-[3-(4-hydroxyphenyl)-2(S)-mercaptopropionylamino]- cyclopentanecarbonyl}-amino)-propionic acid:

mp 236-240 °C; H NMR (500 MHz, CD_gOD) δ 7.80-8.00 (m, 2 H), 7.14-7.56 (m, 5 H), 6.85- 7.00 (m, 2 H), 6.55-6.70 (m, 2 H), 4.60-4.70 (m, 1 H), 3.40-3.50 (m, 1 H), 3.40-3.50 (m, 1 H), 3.25-3.38 (m, 1 H), 3.10-3.26 (m, 1 H), 2.80-3.00 (m, 1 H), 2.70-2.80 (m, 1 H), 2.00-2.14 (m, 1 H), 1.85-2.00 (m, 1 H), 1.65-1.85 (m, 2 H), 1.40-1.60 (m, 2 H), 1.20-1.40 (m, 2 H). IR (KBr, cm^"1) 1709, 1655, 1614, 1516, 1458, 1238, 1203, 1128, 748. Mass spec, m/z calcd for C₃₀H₃₀N₂O₆S 546.6, found 545.9.

(f) 3-(9H-Fluoren-2-yl)-2-{[1-(2-(S)mercapto-3-methylbutanoylamino)- cyclopentanecarbonyl]-amino}-propionic acid

mp 203-204 °C; ¹H NMR (300 MHz, CDCI3) δ 7.62-7.78 (m, 2 H), 7.55 (br d, 1 H), 7.15-7.40 (m, 4 H), 6.80 (br s, 1 H), 4.76-4.84 (m, 1 H), 3.86 (br s, 2 H), 3.15-3.35 (ABX m, 2 H), 3.05 (app dd, 1 H), 2.22-2.30 (m, 1 H), 2.10-2.20 (m, 2 H), 1.88-1.98 (m, 2 H), 1.60-1.80 (m, 4 H), 0.94 (d, 3 H), 0.89 (d, 3 H); IR (KBr, cm"¹) 1734, 1668, 1602, 1509, 1248. Mass spec. m/z calcd for C27H32N2O4S 480.6, found 480.

The starting material, 2-amino-3-(9H-fluoren-2-yl)-propionic acid ethyl ester hydrochloride, is prepared according to the procedure reported by Stork etal{J. Org. Chem. 1976, 3491) for the synthesis of amino esters, from 2-bromomethyl-9H-fluorene. ¹ H NMR (250 MHz, CD3OD) δ 7.82 (dd, 2 H), 7.55 (d, 1 H), 7.47 (br s, 1 H), 7.26-7.39 (m, 3 H), 4.25-4.36 (m, 1 H), 4.26 (q, 2 H), 3.90 (s, 2 H), 1.20 (t, 3 H); IR (KBr, cm"¹) 1747, 1230, 1146, 1049, 763, 740.

Example 4

A solution of 170 mg (0.31) mmol) of 1-(2-acetylthio-3-methyl-butanoylamino)- cyclopentanecarboxylic acid [2-dibenzofurna-3-yl-1-(1H-tetrazol-5-yl)-ethyl]-amide in 5 mL of MeOH is degassed for 5 minutes. At this time, 1.24 mL of 1 N aqueous NaOH is added, and the reaction mixture is stirred at room temperature for 4 hours. The reaction mixture is then diluted with 2 mL of water, and then placed on the rotary evaporator to remove MeOH. The residue is partitioned between 5 mL of water and 10 mL of Et₂O. The aqueous phase is extracted with 10 mL of Et₂O, and acidified with 1.24 mL of 1 N aqueous HC1 solution, and then extracted twice with 10 mL of EtOAc. The combined EtOAc phases are concentrated to give 1 -(2-mercapto-3-methylbutanoylamino)-cyclopentanecarboxylic acid [(S)-2- dibenzofuran-3-yl-1-(1 W-tetrazol-5-yl)-ethyl]-amide as a white solid, mp 222-224 °C; H NMR (500 MHz, CD3OD) δ 7.97 (d, 1 H), 7.90 (d, 1 H), 7.54 (d, 1 H), 7.45 (dd, 1 H), 7.43 (s, 1 H), 7.34 (dd, 1 H), 7.17 (dd, 1 H), 5.56 (dd, 1 H), 3.48-3.57 (ABX m, 2 H), 3.00 (d, 1 H), 2.10-2.20 (m, 1 H), 2.02-2.09 (m, 1 H), 1.90-1.96 (m, 2 H), 1.80-1.85 (m, 1 H), 1.60-1.70 (m, 4 H), 0.98 (d, 3 H), 0.86 (t, 3 H); IR (KBr, cm"¹) 2549, 1660, 1511 , 1204, 751. Mass spec. m/z calcd for C26H30N6O3S 506.6, found 506.

The starting material is prepared as follows:

To a stirred mixture of (S)-2-amino-3-dibenzofuran-3-yl-propionic acid methyl ester hydrochloride (15.7 g, 51.4 mmol) in CH₂CI₂(300 mL) is added triethylamine (9.7 mL, 69 mmol) followed by a solution of di-f-butyl dicarbonate(14g, 64 mmol) in CH₂CI₂(40 mL). A clear solution results after 20 minutes, and the reaction mixture is stirred for 18 hours at room temperature. The solution is washed with ice-cold H₂O (2 x 100 mL), pH 4 (0.5M NaH₂PO₄) buffer (100 mL), H₂O (100 mL) dried (Na₂SO₄), and filtered through a plug of SiO₂ which is washed with EtOAc/ Hexanes (1:1) (500 mL). The filtrate is concentrated to give an oil, the residue is taken up in CH₂CI₂ and concentrated again. The oil slowly crystallizes to give (S)-2-tert-butoxycarbonylamino-3-dibenzofuran-3-yl-propionic acid methyl ester ; ¹H NMR (CDC1₃, 300 MHz) δ 1.53 (9 H, s), 3.2-3.33 (2 H, m), 3.74 (3 H, s), 4.66 (I H, m), 5.05 (1 H, m), 7.3-7.38 (2 H, m), 7.45 (1 H, m), 7.57 (1 H, m), 7.87 (1 H, d), 7.92 (1 H, d).

To a stirred solution of (S)-2-terf-butoxycarbonylamino-3-dibenzofuran-3-yl-propionic acid methyl ester (8.3 g, 22 mmol) in MeOH (100 mL) is added a solution of lithium hydroxide monohydrate (1.85 g, 44 mmol) in H₂O (40 mL) with adequate cooling to prevent the reaction temperature from rising above 20 °C. After 75 minutes, ice cold IN HCI (1 mL) is added and the solution is extracted with CH₂CI₂ (3X50 mL). The organic layer is dried (MgSO₄), filtered, and concentrated under reduced pressure. The residue is recrystallized from etheπhexane (1 :2), filtered, and dried at 50 °C under high vacuum for 2 hours and at room temperature for 12 hours to give (S)-2-tert-butoxycarbonylamino-3-dibenzofuran-3-yl propionic acid, mp 138-140 °C. IR(CH₂CI₂) 1726, 1652, 1233, 1165 cm^'1. ¹H NMR (CDC1₃, 300 MHz) δ 1.25 (9 H, s), 3.12 (2 H, ABX m), 4.41 (I H, m), 5.14 (I H, m), 7.02 (I H, dd), 7.15 (I H, m), 7.21-7.31 (2 H, m), 7.38 (I H, m), 7.70 (I H, d), 7.75 (I H, d). ¹³C NMR (CDC1₃, 300 MHz) δ 28.22, 38.17, 54.45, 79.40, 111.43, 112.44, 120.26, 120.42, 122.64, 122.71, 123.93, 124.30, 126.85, 136.30, 155.09, 156.06, 156.14, 173.34; [α]²⁵ _D= +47.56 (c=l, CH₂CL₂). MS (ES) m/z 354 (M- H). To a stirred solution of (S)-2-tert-butoxycarbonylamino-3-dibenzofuran-3-yl-propionic acid (17. g, 48.7 mmol) in anhydrous DMF (300 mL) under N₂ atmosphere is added a solution of 3- aminopropionitrile (3.93 g, 56 mmol) in anhydrous DMF (50 mL) followed by BOP reagent (26.! g, 60.9 mmol) and triethyl amine (17 mL, 121.8 mmol). The reaction is worked up after 2 houπ it is diluted with EtOAc (I L), added to ice-water (500 mL), and the layers are mixed and separated. The aqueous layer is extracted with EtOAc (2X400 mL), the combined organic layers are washed with H₂O (250 mL), 0.5M pH 4 Buffer (300 mL), half saturated NaCl (300 mL), ice-cold saturated NaHCO₃ (300 mL) and half saturated NaCl (300 mL), and dried (Na₂SO₄), filtered through a plug of SiO₂ (washed with EtOAc/Hexanes, 3:1, 1.5 L). The filtrate is concentrated to give a gelatinous solid, which is treated with f-butylmethyl ether (200 mL), heated to gentle reflux, then hexanes (75 mL) is added and the mixture is cooled to give a white solid, (S)-2-tert-butoxycarbonylamino-N-(2-cyanoethyl)-3-dibenzofuran-3-yl-propionamid( mp 165-166 °C. IR(KBr) 2245, 1673, 1658, 1528, 1176 cm^"1. ¹H NMR (CDC1₃,300 MHz) δ 1.38 (9 H, s), 2.38-2.6 (2 H, m), 3.17-3.27(2 H, m), 3.36-3.48 (2 H, m), 4.43(1 H, m), 5.21 (I H, m), 6.74 (1 H, m), 7.17 (I H, dd), 7.32(1 H, m), 7.38-7.48 (2 H, m), 7.54 (I H, m), 7.87 (I H, d), 7.91 (1 H, d). ¹³C NMR (CDCI₃, 75 MHz) δ 18.18, 28.25, 35.58, 38.75, 55.99, 80.63, 11.68, 112.35, 117.82, 120.58, 120.78, 122.82, 123.25, 123.91, 123.97, 127.15, 135.97, 155.55, 156.33, 156.43, 172.06 ppm. [α]²⁵ _D = +6.2778; 10.229 mg/mL CHC1₃.

To a mechanically stirred suspension of (S)-2-te/t-butoxycarbonylamino-N-(2-cyanoethyl)-3- dibenzofuran-3-yl-propionamide (18.2 g, 44.7 mmol) and triphenyl phosphine (29.3 g, 112 mmol) in ice cold anhydrous MeCN (350 mL) are added from 2 separate addition funnels, first, diisopropylazodicarboxylate (DIAD, 22 mL, 11.2 mmol) and, 2 minutes later, trimethylsilyl azide (16 mL, 11.8, mmol) over 20 minutes. The heterogeneous reaction mixture is allowed to warm up to room temperature over 30 minutes and stirred for 14 hours. To the mixture cooled to O °C is added, under stirring, a solution of NaNO₂ (3.1 g, 45 mmol) in H₂O (15 mL). After 30 minutes, a solution of eerie ammonium nitrate (25 g, 45 mmol) in H₂O (70 mL) is added over 10 minutes and the mixture is stirred for 20 minutes. The mixture is added to ice-H₂O (300 mL) and extracted with CH₂CI₂ (1 L then 2X250 mL). The combined organic layers are washed with H₂O (2X250 mL), dried (MgSO₄) filtered, and concentrated in vacuo. The yellow solid residue is recrystallized from isopropanol (350 mL). The solid is filtered, washed successively with cold isopropanol (100 mL) and etheπhexane (1:1, 200 mL), and dried under high vacuum at 60 °C to give (S)-3-[5-(l-fert- butoxycarbonyIamino-2-dibenzof uran-3-yl-ethyl)-tetrazol-l -yl] propionitrile mp 201 -202 °C. IR(KBr) 2203, 1680, 1508, 1249, 1233, 1163 cm^"1. ¹H NMR (DMSO-d₆, 300 MHz) δ 1.22 (9 H, s), 2.97-3.18 (2 H, m), 3.35-3.51 (2 H, m), 4.62-4.72 (2 H, m), 5.30 (I H, m), 7.29-7.42 (2 H, m), 7.50 (I H, m), 7.62-7.71 (2 H, m), 7.92 (I H, d), 8.05 (I H, d), 8.11 (I H, d). ¹³C NMR (DMSO-d₆, 75 MHz) δ 17.76, 27.91 , 42.33, 45.81 , 78.86, 111.6, 112.52, 117.63, 120.67, 120.98, 122.14, 123.03, 123.46, 124.62, 127.31 , 136.97, 155.36, 155.51 , 155.57, 156.29 ppm. [α]²⁵ _D= -5.41 (c=l, DMSO).

A mixture of (S)-3-[5-(I-tert-butoxycarbonylamino-2-dibenzofuran-3-yl-ethyl)-tetrazol-l-yl] propionitrile (15.6 g, 36.1 mmol) in formic acid (200 mL) is stirred at 48 °C for 50 minutes. The reaction mixture is concentrated in vacuo to remove most of the formic acid. Ether is added and the oil crystallizes. Additional ether is added and the mixture is cooled in an ice bath. The solid is filtered and dried under high vacuum at 50 °C to give (S)-3-[5-(l-amino-2- dibenzofuran-3-yl-ethyl)-tetrazol-1-yl]propionitriie (formate salt); mp 173-174 °C. IR (KBr) 2203, 1632,1557,1190 cm^"1. ¹H NMR (DMSO-d₆, 300 MHz) δ 3.0-3.21 (2 H, m), 4.58-4.73 (3 H, m), 7.24 (I H, dd), 7.37 (I H, m), 7.48 (I H, m), 7.5 8 (1 H, s), 7.66 (1 H, d), 8.02 (1 H, d), 8.09 (1 H, d), 8.15 (1 H, s). [α]²⁵ _D = +11.850; 10.124 mg/mL DMSO.

(S)-3-[5-(1- Amino-2-dibenzofuran-3-yl-ethyl)-tetrazol-1-yl]propionitrile is converted to 1-(2- bromo-3-methyl-butlanoylamino)-cyclopentanecarboxylic acid {2-dibenzofuran-3-yl-l-[l-(2- cyanoethyl-)-tetrazol-5-yl-ethyl}-amide using the general method described in example 1.

A solution of 214 mg (0.353 mmol) of 1-(2-bromo-3-methyl-butanoylamino)- cyclopentanecarboxylic acid {2-dibenzofuran-3-yl-l-[l-(2-cyanoethyl-)-tetrazol-5-yl]-ethyl}- amide in 3 mL of MeOH is degassed for 5 minutes. At this time, 0.70 mL of IN aqueous NaOH is added, and the reaction mixture is stirred at room temperature for 3 hours. The reaction mixture is then concentrated to give 1-(2-bromo-3-methylbutanoylamino)- cyclopentanecarboxylic acid [(S)-2-dibenzofuran-3-yl-1-(IH-tetrazol-5-yl)-ethyl]-amide as a white solid; Η NMR (300 MHz, CD₃OD) δ 7.90 (d, I H), 7.84 (d, I H), 7.49 (d, I H), 7.37-7.43 (m, 2 H), 7.27 (t, I H), 7.18 (dd, I H), 5.63-5.70 (m, I H), 4.13 (d, I H), 3.62 (dd, I H), 3.45 (dd, I H), 2.00-2.20 (m, 2 H), 1.50-2.00 (m, 6 H), 1.25-1.45 (m, I H), 1.02 (d, 3 H), 0.91 (d, 3 H). The above bromide is converted to 1 -(2-acetylthio)-3-methylbutanoylamino)- cyclopentanecarboxylic acid [(S)-2-dibenzofuran-3-yl-1-(1 r -tetrazol-5-yl)-ethyl]-amide according to procedure described in example 1.

Example 5

Prepared similarly according to procedures described in the previous examples are:

(a) 2-{[I-(2S-Mercapto-3-methyl-pentanoylamino)-cyclopentanecarbonyl]-amino}-3-(9-oxo- 9W-fluoren-2-yl)-propionic acid from 2-amino-3-(9-oxo-9H-fluoren-2-yl)-propionic acid hydrochloride; mp 128-130 °C; ¹H NMR (250 MHz, CD₃ OD) δ 7.45-7.65 (m, 4 H), 7.35- 7.45 (m, 2 H), 7.29 (t, I H), 4,60-4.70 (m, 1 H), 3.15-3.25 (m, H), 3.00-3.12 (m, I H), 2.00-2.30 (m, 3 H), 1.85-2.00 (m, 2 H), 1.65-1.85 (m, 4 H), 1.35-11.50 (m, I H), 0.75-0.90 (m, 6 H); IR (KBr, cm^"1) 2560, 1710, 1650, 1612, 1519, 1181 Mass spec.mz calcd for C₂₈H₃₂N₂0₅ S 508.6, found 508.

The starting material, 2-amino-3-(9-oxo-9H-fluoren-2-yl)-propionic acid ethyl ester hydrochloride is prepared according to the procedure reported by Stork et al (J. Org. Chem. 1976, 3491) for the synthesis of amino esters, from 2-bromomethyl-fluorene-9- one. ¹H NMR (250 MHz, DMSO-d₆ ) δ 7.76 (t, 2 H), 7.56-7.62 (m, 2 H), 7.45-7.50 (m, 2 H), 7.35 (t, 1 H), 4.30 (br t 1 H), 4.10-4.20 (m, 2 H), 3.15 (t, 2 H), 1.10 (t, 3 H); IR (KBr, cm^"1) 1740, 1716, 1603, 1509, 1236, 857, 735. Mass Spec m/z calcd for C₁₈ Hι₇ NO₃ HCI 295.33, found 295.

2-Bromomethyl-fluorene-9-one is prepared from 2-hydroxymethyl-9W-fluorene by the method of Gannon and Krause (Gannon, S.M; Krause, J.G. Synthesis 1987, 915): Η NMR (CDCIg, 300 MHz) δ 7.64-7.67 (m, 2 H), 7.46-7.53 (m, 4 H), 7.30 (dt, I H), 4.48 (s, 2 H). ¹³C NMR (CDCI₃, 75 MHz): δ 193.2, 144.4. 143.9, 139,0, 135.3, 134.9, 134.8, 134.3, 129.4, 124.9, 124.5, 120.7, 120.5, 32.7. IR (KBr, cm^"1) 1723, 1600, 1459, 1180, 973, 743.

(b) 2-{[1-(2-Mercapto-4-methylthio-butanoylamino)-cyclopentanecarbonyl]animo}-3-(9-oxo- 9H-fluoren-2-yl)-propionic acid )from 2-amino-3-(9-oxo-9H-fluoren-2-yl)-propionic acid; ¹ H NMR (250 MHz, CD₃OD) δ 7.35-7.65 (m, 6 H), 7.25-7.35 (m.. I H). 4.60-4.75 (m, I H), 3.35-3.45 (m, I H), 3.00-3.15 (m, 1 H), 2.40-2.55 (m, 2 H), 1.65-2.20 (m, 11 H), 1.95, 1.99 (s each, 3 H); IR (KBr, cm^"1) 2545, 1746, 1715, 1672, 1608 1506, 1173, 743. Mass spec.m/ caled for C₂₇ H₃₀ N₂ 0₅ S₂526.68, found 526.

(c) 3-Dibenzofuran-4-yl-2-{[1-(2-mercapto-4-methylpentanoylamino)-cyclopentanecarbonyl]- aminoj-propionic acid from 2-amino-3-dibenzofuran-4-yl-propionic acid; ¹H NMR (500 MHz, CD₃OD) δ 8.03 (dd, 1 H), 7.86 (s, 1 H), 7.53 (dd, 1 H), 7.43-7.47 (m, 2 H), 7.31- 7.35 (m, 2 H), 4.70-4.75 (m, 1 H), 3.35-3.42 (m, 1 H), 3.15-3.28 (m, 2 H), 2.10-2.20 (m, 2 H), 1.85-1.98 (m, 2 H), 1.50-1.75 (m, 6 H), 1.28-1.40 (m, 1 H), 0.69-0.82 (m, 6 H); IR (KBr, cm^"1) 3283, 2956, 2870, 1742, 1652, 1516, 1196. Mass spec, m/z calcd for C₂₇H₃₂N₂0₅S 496.63, found 496.

The starting material, 2-amino-3-dibenzofuran-4-yl-propionic acid ethyl ester hydrochloride, is prepared according to the procedure reported by Stork et al (J. Org. Chem. 1976, 3491) for the synthesis of amino esters, from 4- bromomethyldibenzofuran. ¹H NMR (300 MHz, CD₃OD) δ 8.03 (d, 1 H), 7.96 (d, 1 H), 7.59 (d, 2 H), 7.50 (dt, 1 H), 7.35-7.38 (m, 2 H), 4.38 (t, 1 H), 4.26 (q, 2 H), 3.29-3.47 (ABX m, 2 H), 1.22 (t, 3 H); IR (KBr, cm^"1) 1735, 1239. 1199, 1060, 746. Mass Spec m/z calcd for C₁₇ H_n NO₃ HC1 283.32, found 283.

Claims

What is claimed is:

1. A compound of the formula

wherein

Ri represents hydrogen or acyl;

R₂ represents hydrogen, lower alkyl, carbocyclic or heterocyclic aryl, carbocyclic or heterocyclic aryl-lower alkyl, cycloalkyl, cycloalkyl-lower alkyl, biaryl, biaryl-lower alkyl,

(hydroxy, lower alkoxy or acyloxy)-lower alkoxy;

A together with the carbon atom to which it is attached forms a ring and represents 3 to 10 membered cycloalkylidene or 5 to 10 membered cycloalkenylidene radical which may be substituted by lower alkyl or aryl-lower alkyl or may be fused to a saturated or unsaturated carbocyclic 5-7-membered ring; or A together with the carbon to which it is attached represents 5 to 6 membered oxacycloalkylidene, thiacycloalkylidene or azacycloalkylidene optionally substituted by lower alkyl or aryl-lower alkyl; or A together with the carbon atom to which it is attached represents 2,2-norbonylidene;

X represents O, CH₂, C=O, N-R or S;

R₄ represents hydrogen, acyl, lower alkyl or aryl-lower alkyl;

Y represents 5-tetrazolyl, carboxyl or carboxyl derivatized in form of a pharmaceutically acceptable ester; a disulfide derivative derived from a said compound wherein Ri is hydrogen; or a pharmaceutically acceptable salt thereof.

2. A compound according to claim 1 wherein R^ represents hydrogen or acyl derived from a carboxylic acid; R₂ represents hydrogen, lower alkyl, hydroxy-lower alkyl, or carbocyclic or heterocyclic aryl-lower alkyl; A represents C₂-C₆-straight chain alkylene optionally substituted by lower alkyl, or C₂-C -straight chain alkylene interrupted by 1 ,2- phenylene or by 1 ,2-C₅-or C₆-cycloalkylene, or C₃- or C₄-straight chain alkylene interrupted by oxygen, sulfur or by NR₄ wherein R₄ is hydrogen, acyl, aryl-lower alkyl or lower alkyl; X is O, S, CH₂ or C=O; Y represents 5-tetrazolyl, carboxyl or carboxyl derivatized in form of a pharmaceutically acceptable ester; a disulfide derivative derived from a said compound wherein Ri is hydrogen; or a pharmaceutically acceptable salt thereof.

3. A compound according to claim 1 wherein the asymmetric carbon with the substituent Y has the S-configuration.

A compound according to claim 1 of the formula II

wherein Ri represents hydrogen or carboxyl derived acyl; R₂ represents hydrogen, lower alkyl, hydroxy-lower alkyl, carbocyclic or heterocyclic aryl, carbocyclic or heterocyclic aryl- lower alkyl, cycloalkyl, cycloalkyl-lower alkyl, biaryl or biaryl-lower alkyl; X is O, S, CH₂ or C=O; Y represents 5-tetrazolyl, carboxyl or carboxyl derivatized in form of a pharmaceutically acceptable ester; n represents 2-6; a disulfide derivative derived from a said compound wherein Ri is hydrogen; or a pharmaceutically acceptable salt thereof.

5. A compound according to claim 4 wherein Ri represents hydrogen, aryl-lower alkanoyl, lower alkanoyl, lower alkoxy-lower alkanoyl, or heterocyclic or carbocyclic aroyl; R₂ represents hydrogen, lower alkyl or carbocyclic aryl-lower alkyl; X is O; Y represents 5- tetrazolyl, carboxyl, lower alkoxycarbonyl, carbocyclic or heterocyclic aryl-lower alkoxycarbonyl, α-(lower alkanoyloxy-, lower alkoxycarbonyl- or di-lower alkylaminocarbonyl)-lower alkoxycarbonyl; n is 2, 4 or 5; or a pharmaceutically acceptable salt thereof.

6. A compound according to claim 5 of formula

or of formula Ilia

wherein

Y represents 5-tetrazolyl, carboxyl, lower alkoxycarbonyl, benzyloxycarbonyl, pyridylmethoxycarbonyl, α-(lower alkanoyloxy-, lower alkoxycarbonyl- or di-lower alkylaminocarbonyl-) lower alkoxycarbonyl; or a pharmaceutically acceptable salt thereof.

7. A compound according to claim 6 wherein Ri represents hydrogen, or lower alkanoyl; R₂ represents lower alkyl, benzyl, or benzyl substituted by hydroxy, halo, lower alkyl, lower alkanoyloxy, lower alkoxy or trifluoromethyl; Y represents 5-tetrazolyl, carboxyl, lower alkoxycarbonyl, benzyloxycarbonyl, pyridylmethoxycarbonyl, α-(lower alkanoyloxy-, lower alkoxycarbonyl- or di-lower alkylaminocarbonyl-) lower alkoxycarbonyl; or a pharmaceutically acceptable salt thereof.

8. A compound according to claim 7 wherein Ri is hydrogen or lower alkanoyl; R₂ is Cι- C₄-alkyl; and Y is 5-tetrazolyl, carboxyl or lower alkoxycarbonyl.

9. A compound according to claim 1 of the formula

wherein

W represents CH₂, O, S or NR₄ in which R is hydrogen, acyl, lower alkyl or aryl-lower alkyl;

10. A compound according to claim 9 wherein Ri is hydrogen or lower alkanoyl; R₂ is C C₄-alkyl; Y is 5-tetrazolyl, carboxyl or lower alkoxycarbonyl; and W is CH₂, or a pharmaceutically acceptable salt thereof.

11. A compound according to claim 7 which is (S)-3-dibenzofuran-3-yl-2-{[1-(2S-mercapto- 3R-methylpentanoylamino)-cycIopentanecarbonyl]-amino}-propionic acid, the compound of formula III wherein Ri is hydrogen, R₂ is 2-butyl, X is O and Y is carboxyl; or a pharmaceutically acceptable salt thereof.

12. A pharmaceutical composition comprising a compound according to claim 1 in combination with one or more pharmaceutically acceptable carriers.

13. Use of a compound of any one of claims 1-11 or a pharmaceutically acceptable salt thereof for the preparation of a medicament for inhibiting endothelin dependent disorders.

14. A method of treating endothelin dependent disorders in a mammal which comprises administering to a mammal in need thereof an effective endothelin converting enzyme inhibiting amount of a compound of claim 1.