WO1997024342A1 - Thiol derivatives with metallopeptidase inhibitory activity - Google Patents

Thiol derivatives with metallopeptidase inhibitory activity Download PDF

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Publication number
WO1997024342A1
WO1997024342A1 PCT/EP1996/005663 EP9605663W WO9724342A1 WO 1997024342 A1 WO1997024342 A1 WO 1997024342A1 EP 9605663 W EP9605663 W EP 9605663W WO 9724342 A1 WO9724342 A1 WO 9724342A1
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WO
WIPO (PCT)
Prior art keywords
phenylalanme
mercapto
group
methyl ester
compound
Prior art date
Application number
PCT/EP1996/005663
Other languages
French (fr)
Inventor
Franco Pellacini
Mario Fantucci
Gabriele Norcini
Stefano Romagnano
Francesco Santangelo
Claudio Semeraro
Original Assignee
Zambon Group S.P.A.
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Filing date
Publication date
Priority to AU13018/97A priority Critical patent/AU713156B2/en
Priority to EA199800544A priority patent/EA000991B1/en
Application filed by Zambon Group S.P.A. filed Critical Zambon Group S.P.A.
Priority to RO98-01109A priority patent/RO119882B1/en
Priority to SK889-98A priority patent/SK282977B6/en
Priority to HU9903636A priority patent/HUP9903636A3/en
Priority to JP09524001A priority patent/JP2000502687A/en
Priority to UA98074090A priority patent/UA62923C2/en
Priority to DE69634543T priority patent/DE69634543D1/en
Priority to EP96944583A priority patent/EP0877740B1/en
Priority to EE9800192A priority patent/EE03766B1/en
Priority to IL12491596A priority patent/IL124915A/en
Priority to PL96327580A priority patent/PL188151B1/en
Priority to APAP/P/1998/001264A priority patent/AP1186A/en
Priority to BR9612373A priority patent/BR9612373A/en
Priority to AT96944583T priority patent/ATE292123T1/en
Priority to KR1019980704934A priority patent/KR19990076809A/en
Priority to NZ325785A priority patent/NZ325785A/en
Publication of WO1997024342A1 publication Critical patent/WO1997024342A1/en
Priority to BG102553A priority patent/BG63942B1/en
Priority to IS4780A priority patent/IS1885B/en
Priority to NO19982977A priority patent/NO310914B1/en
Priority to HK99103110A priority patent/HK1018008A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/26—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/425—Thiazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/24—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with substituted hydrocarbon radicals attached to ring carbon atoms
    • C07D213/54—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
    • C07D213/55—Acids; Esters
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D241/00—Heterocyclic compounds containing 1,4-diazine or hydrogenated 1,4-diazine rings
    • C07D241/02—Heterocyclic compounds containing 1,4-diazine or hydrogenated 1,4-diazine rings not condensed with other rings
    • C07D241/10—Heterocyclic compounds containing 1,4-diazine or hydrogenated 1,4-diazine rings not condensed with other rings having three double bonds between ring members or between ring members and non-ring members
    • C07D241/12—Heterocyclic compounds containing 1,4-diazine or hydrogenated 1,4-diazine rings not condensed with other rings having three double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/02—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings
    • C07D277/20—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
    • C07D277/22—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • C07D277/30—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
    • C07D307/02—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings
    • C07D307/34—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
    • C07D307/38—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with substituted hydrocarbon radicals attached to ring carbon atoms
    • C07D307/54—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D333/00—Heterocyclic compounds containing five-membered rings having one sulfur atom as the only ring hetero atom
    • C07D333/02—Heterocyclic compounds containing five-membered rings having one sulfur atom as the only ring hetero atom not condensed with other rings
    • C07D333/04—Heterocyclic compounds containing five-membered rings having one sulfur atom as the only ring hetero atom not condensed with other rings not substituted on the ring sulphur atom
    • C07D333/06—Heterocyclic compounds containing five-membered rings having one sulfur atom as the only ring hetero atom not condensed with other rings not substituted on the ring sulphur atom with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to the ring carbon atoms
    • C07D333/24—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • the present invention relates to thiol denvatives with metallopeptidase inhibitory activity and, more particularly, it relates to N-mercaptoacyl phenylalanine denvatives useful in the treat ⁇ ment of cardiovascular diseases
  • ECE endothelin converting enzyme
  • NEP neutral endopeptidase
  • enkephalinase compounds with inhibitory activity of the neutral endopeptidase (NEP) enzyme, also called enkephalinase, are useful as vasodilators since the NEP enzyme is responsible for the inactivation, not only of endogenous enkephaline, but also of some natriuretic factors such as, for instance, the atnal factor (ANF), a hormone secreted by heart which increases the vaso- dilatation and, on the renal level, increases diuresis and natnuresis
  • NAF atnal factor
  • the compounds with metallopeptidase inhibitory activity are generally used, alone or in combmation, in the treatment of hypertension, renal failure, congestive heart failure and lschemic cardiopathoiogies
  • Thiorphan (DL-(3-mercapto-2- benzyipropanoyl)glyc ⁇ ne]
  • Captop ⁇ l (The Merck Index, XI ed , No 1773, page 267) are considered the parent compounds for NEP-inhibitors and ACE-inhibitors, respectively
  • Other molecules having thiol structure endowed with metallopep ⁇ dase inhibitory activity are desc ⁇ bed in the literature
  • the European patent application No. 0524553 (Thstitut National de la Sante et de la Recher ⁇ che Medicale (INSERM)] describes acylmercaptoalkanoyldipepudes endowed with neutral endopeptidase and peptidyldipeptidase A inhibitory activity.
  • the European patent application No. 0566157 (Schering Co ⁇ oration) describes thiol deriva- tives such as, for instance, N-mercaptoacyl amino acids, endowed with NEP-inhibitory activ ⁇ ity.
  • ⁇ -Mercaptoacyl dipeptides endowed with ACE-inhibitory and NEP-inhibitory activity are also described by S.S. Bhagwat et al.
  • object ofthe present invention are the compounds of formula 1
  • R is a mercapto group or a R4COS group convertible in the organism to mercapto group
  • R j is a straight or branched C2-C4 alkyl group or an aryl or arylalkyl group having from 1 to 6 carbon atoms in the alkyl moiety wherein the aryl is a phenyl or a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consisting of mtrogen, oxygen and sulphur, optionally substituted with one or more substituents, the same or different, selected from the group consisting of halogen atoms, hydroxy groups, alkoxy, alkyl, alkylthio, alkylsulphonyl or alkoxycarbonyl groups having from 1 to 6 carbon atoms in the alkyl moiety, C1-C3 alkyl groups containing one or more fluorine atoms, carboxy groups, nitro groups, ammo or aminocarbonyl groups, acylarruno groups, aminosulphonyl groups, mono- or -alkylamino or mono- or di-al
  • R 2 is a hydrogen atom, a straight or branched C1-C4 alkyl or a benzyl group
  • R3 is a phenyl group substituted with a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consisting of nitrogen, oxygen and sulphur, being the phenyl and the heterocyclic groups optionally substituted with one or more substituents, the same or different, as indicated for R ]
  • R3 is a phenyl group substituted with a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consisting of nitrogen, oxygen and sulphur, being the phenyl and the heterocyclic groups optionally substituted with one or more substituents, the same or different, as indicated for R ]
  • R4 is a straight or branched C1-C4 alkyl group or a phenyl group, d e carbon atoms marked with an astensk are stereogenic centers, and pharmaceutically acceptable salts thereof
  • object of the present mvenUon are the compounds of formula I in the form of stereoisome ⁇ c mixture as well as in the form of single stereoisomers
  • straight or branched alkyl group we intend an alkyl such as methyl, ethyl, n propyl, isopropyl, n butyl, sec-butyl, tert- butyl, isobutyl, n pentyl, 2-pentyl, 3-pentyl, isopentyl, tert-pentyl, n hexyi and isohexyl group
  • straight or branched alkoxy group we intend an alkoxy such as methoxy, ethoxy, n propoxy and isopropoxy group
  • halogen atom we intend a fluorine, chlorine, bromme or iodine atom
  • acyl we intend an acyl group de ⁇ vmg from an aliphatic or aromatic carboxylic acid such as acetic, propionic,
  • Examples of pharmaceutically acceptable salts of the compounds of formula I are the salts with alkali or alkali-earth metals and the salts with pharmaceutically acceptable organic bases
  • Preferred compounds of formula I are the compounds wherem R. is a phenyl group substi ⁇ tuted in position 4 with a heterocyclic group
  • More preferred compounds, in this class are the compounds of formula I wherein R repre ⁇ sents an arylalkyl group optionally substituted with one or more substituents, the same or dif ⁇ ferent, selected from the group consisting of halogen atoms, hydroxy, alkyl or alkoxy
  • Still more preferred compounds, in this class are the compound of formula I wherein R ] rep ⁇ resents a phenylalkyl group optionally substituted with one or more substitutents, the same or different, selected from the group consisting of halogen atoms, hydroxy, alkyl or alkoxy
  • Preferred examples of pharmaceutically acceptable salts of the compounds of formula I are the salts with alkali metals such as sodium, lithium and potassium
  • preferred compounds of formula I object of die present mvention, are N-(3-phenylcarbonyllh ⁇ o-2-phenylme ⁇ ylprop ⁇ onyl)-4-(2-th ⁇ azolyl)-phenylalamne methyl es- ter,
  • condensation reaction is earned out accordmg to conventional techniques of the chemistry of peptides Before carrymg out the reaction it can be useful to properly protect the optional functional groups which could interfere in the reaction
  • the compounds of formula II are known or easily prepared accordmg to conventional methods as descnbed, for instance, m Bntish patent No 1576161 in die name of E R Squibb & Sons Inc Altematively, tiie compounds of formula II can be prepared accordmg to the synd etic meth ⁇ ods descnbed by M C Foumie-Zalusky et al. in J Med Chem 1994, 37, 1070-1083 Also the intermediate compounds of formula HI are known or easily prepared accordmg to known methods For instance, the compounds of formula UJ can be prepared accordmg to the synthetic meth- ods descnbed by W C Shieh et al m J Org Chem 1992, 57, 379-381
  • die compounds of formula HI can be prepared by coupling methods (cross- coupling) starting from halogenated heterocyclic compounds and stannyl phenylalanine de ⁇ ⁇ vatives, as descnbed by D.S Wilbur et al in Bioconjugate Chem , 1993, 4, 574-580
  • the compounds of formula I in the form of single stereoisomers are prepared by stereoselec- tive synthesis or by separation of the stereoisomenc mixture accordmg to conventional tech ⁇ niques
  • the inhibitory activity ofthe compounds of formula I was evaluated m compan- son to the aforementioned Thio ⁇ han and Captop ⁇ l
  • the vitro inhibitory activity of the compounds of formula I expressed as IC50 value, re ⁇ sulted at nM concentrations
  • the compounds of formula I can be formulated in solid or liquid pharmaceutical compositions, suitable to oral or parenteral administration
  • compositions containing a therapeutically effective amount of a compound of formula I admixture with a earner for pharmaceutical use are a further object ofthe present mvention
  • compositions accordmg to the present mvention are tab- lets, coated tablets, capsules, granulates, solutions and suspensions suitable to oral admini ⁇ stration, solutions and suspensions suitable to parenteral administration
  • the pharmaceutical compositions object of me present mvention are prepared according to conventional techniques
  • the daily dose ofthe compound of formula I or of me correspondmg pro-drug will depend on several factors such as the se ⁇ ousness of the ⁇ sease, the mdividual response of the patient or the land of formulation but it is usually comp ⁇ sed between 0 1 mg and 10 mg per kg of body weight divided mto a single dose or mto more daily doses
  • reaction mixture was then treated with petroleum ether 40-60°C (100 ml) and cooled at
  • reaction mixture was kept under stirring for 20 hours, then dicyclohexyl urea was filtered off and the solvent was evaporated at reduced pressure The residue was collected with ethyl acetate and the solution was washed with an aqueous solution of sodium chlonde at 20%, sodium bicarbonate at 5% and sodium chlonde at 20% agam
  • N-[(2S)-3-phenylcarbonyld ⁇ o-2-phenylmethylprop ⁇ onyl]-4-(2-th ⁇ azolyl)-L-phenylalanme ethyl ester (1.4 g; 2.57 mmoles), prepared as described in example 6, was suspended in ethanol (30 ml), degassed by nitrogen bubbling to eliminate the oxygen.
  • reaction mixture was kept under stirring for 4 hours at room temperature, then cooled at
  • the NEP-inhibitory activity was evaluated m rat kidney cortex membranes prepared ac- cording to the procedure descnbed by T Maeda et al in Biochim Biophys Acta 1983,
  • Kidneys were removed from male Sprague-Dawley rats weighing approximately 300 g and kept at 4 C C
  • Kidney cortex was carefully dissected, finely minced and suspended m a homogenization buffer (10 mM sodium phosphate pH 7 4 containmg 1 mM MgCl 2 , 30 mM NaCl, 002%
  • the tissue was then cold homogenized for 30 seconds usmg an Ultra-Turrax homogenizer
  • NEP-inhibitory activity was evaluated by usmg the method desc ⁇ bed by C Llorens et al , Eur J Pharmacol , 69, (1981), 113-116, as hennafter reported Aliquots of die membrane suspension prepared as above descnbed (concentration 5 ⁇ g/ml of proteins) were preincubated m the presence of an aminopeptidase inhibitor (Bestatin - 1 mM) for 10 minutes at 30°C
  • IC50 (nM) value a percentage of inhibition of the metabolite formation m the membrane preparations treated with the compounds of formula I and wi the comparative compounds with re- spect to the untreated membrane preparations was expressed as IC50 (nM) value b) ACE-inhibitorv activity
  • the ACE-inhibitory activity was evaluated according to die method reported in die litera ⁇ ture by B Holmquist et al., m Analytical Biochemistry 95, 540-548 (1979) 50 ⁇ M of ACE (250 mU/ml punfied by lung rabbit, EC 3 4 15 1 SIGMA) were premcu- bated with 50 ⁇ l of the compound of formula I or of die reference compound or related vehicle in mermostated cuvettes at 37°C
  • the reaction was started by add g furylacryloylphenylalanylglycylglycine 0 8 mM (FAPGG-SIGMA) Contemporaneously, by usmg a Beckman DU-50 spectrophotometer provided with a pro- gram for calculating delta A/minutes and regression coeffi ⁇ ents of the enzyme kinetics curves, the absorbance at 340 nm was recorded in continuo for 5 minutes
  • the percentage ofthe enzyme inhibition in the preparations treated with the compounds of formula I or with the comparative compounds with respect to the untreated preparations was expressed as IC,- (nM) value
  • the IC50 (nM) values related to die ACE-inhibitory activity and NEP-inhibitory activity of the compounds 16, 18-25, 27-33 and of the compa ⁇ son compounds Thio ⁇ han and Captop ⁇ l are reported m the following table 1
  • IC,_ (nM) value of compounds
  • Captopril 4.6 not active The data reported in table 1 show that d e compounds of formula I, object of the present in ⁇ vention, are endowed wrth a significant mixed ACE/NEP inhibitory activity Said activity is comparable to that of Captop ⁇ l, to what it concerns the ACE inhibitory activ- ity, and greater dian that of Thio ⁇ han, to what it concerns the NEP inhibitory activity
  • the inhibitory activity of the compounds of formula I was evaluated kidneys of sponta ⁇ neously hypertensive rats (SHR), 5 mmutes after 1 v injection (0 6 and 21 ⁇ moles/kg) and 30 mmutes, 60 mmutes and 4 hours after oral administration (30 ⁇ moles/kg) of the tested compounds
  • SHR sponta ⁇ neously hypertensive rats
  • the renal tissue was homogenized and incu ⁇ bated for 15 mmutes at 37°C m the presence of Glutaryl-Ala-Ala-Phe-2-naphthylam ⁇ de (GAAP), as a substrate, and arrunopeptidase M at pH 7 6
  • the reaction was stopped by addmg an aqueous solution at 10% of tnchloroacetic a ⁇ d
  • the released 2-naphthylamme was determmed by addmg fast garnet dye (2 ml)
  • Enzyme reaction rates were determmed by measuring the mcrease m the optical density at
  • the NEP-inhibitory activity of the tested compounds was expressed as percentage of NEP-inhibition m SHR kidneys b) ACE-inhibitorv activity
  • the ex vivo ACE-inhibitory activity was evaluated by usmg a radiomet ⁇ c assay method, as reported m the literature by J W Ryan et al in Biochem J (1977), 167, 501-504
  • the inhibitory activity ofthe compounds of formula I was evaluated lungs of spontane ⁇ ously hypertensive rats (SHR), 5 mmutes after l v injection (0 6 and 21 ⁇ moles/kg) and 30 mmutes, 60 minutes and 4 hours after oral administration (30 ⁇ moles/kg) of die tested compounds
  • SHR spontane ⁇ ously hypertensive rats
  • 5 mmutes after l v injection (0 6 and 21 ⁇ moles/kg)
  • 30 mmutes 60 minutes and 4 hours after oral administration (30 ⁇ moles/kg) of die tested compounds
  • the lung tissue was homogenized and incubated for 2 hours at 37°C m the presence of [ H]Hyp-Gly-Gly, as a substrate
  • the released radiolabeUed hyppu ⁇ c a ⁇ d was extracted with ethyl acetate and counted by liquid scintillation spectrometry, accordmg to conventional methods
  • the ACE-inhibttory activity of the tested compounds was expressed as percentage of

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Abstract

Compounds of formula (I), wherein R, R1, R2, and R3 have the meanings reported in the description, processes for their preparation and pharmaceutical compositions which contain them as active ingredients are described. The compounds of formula (I) are endowed with a mixed ACE-inhibitory and NEP-inhibitory activity and are useful in the treatment of cardiovascular diseases.

Description

"Thiol deπvatives with metallopeptidase inhibitory activity"
**********************
The present invention relates to thiol denvatives with metallopeptidase inhibitory activity and, more particularly, it relates to N-mercaptoacyl phenylalanine denvatives useful in the treat¬ ment of cardiovascular diseases
The pharmacologic interest towards the study of metallopeptidase inhibitory molecules deπves from the role that said enzymes exert on the level ofthe cardio rculatory system It is well-known in fact that compounds with angiotensin converting enzyme (ACE) inhibitory activity are mainly useful m the treatment of hypertension, heart failure and post-infarct m that they inhibit die formauon of angiotensin π, a substance which produces several effects among which the increase ofthe blood pressure
Compounds with endothelin converting enzyme (ECE) inhibitory activity are useful as anti- vasoconstnctors because they inhibit the formation of endothelin, a 21 amino acid peptide with vasoconstrictor activity
Instead, compounds with inhibitory activity of the neutral endopeptidase (NEP) enzyme, also called enkephalinase, are useful as vasodilators since the NEP enzyme is responsible for the inactivation, not only of endogenous enkephaline, but also of some natriuretic factors such as, for instance, the atnal factor (ANF), a hormone secreted by heart which increases the vaso- dilatation and, on the renal level, increases diuresis and natnuresis
Therefore, even exerting their action on the cardiovascular system with different mechanisms of action, the compounds with metallopeptidase inhibitory activity are generally used, alone or in combmation, in the treatment of hypertension, renal failure, congestive heart failure and lschemic cardiopathoiogies Among the thiol deπvatives inhibitors of metallopeptidases, Thiorphan [(DL-(3-mercapto-2- benzyipropanoyl)glycιne], descπbed for the first time by Roques et al in Nature, Vol 288, pages 286-288, (1980), and Captopπl (The Merck Index, XI ed , No 1773, page 267) are considered the parent compounds for NEP-inhibitors and ACE-inhibitors, respectively Other molecules having thiol structure endowed with metallopepϋdase inhibitory activity are descπbed in the literature
The US patents No 4,401,677 and No 4,199,512 (both in the name of E R Squibb & Sons, Inc.) describe mercaptoalkanoyl amino acids endowed with enkephalinase inhibitory activity and ACE-inhibitory activity, respectively.
The European patent application No. 0419327 (Societe Civile Bioproject) describes amino acid derivatives endowed with enkephalinase and ACE inhibitory activity, respectively.
The European patent application No. 0449523 (E.R. Squibb & Sons, Inc.) describes mercapto or acylthio trifluoromethylarnides with NEP-inhibitory activity.
The intemational patent application No. WO 93/08162 [Rhone-Poulenc Rorer S A. - Institut National de la Sante et de la Recherche Medicale (ENSERM)] describes β,β-disubstituted α- mercaptomethylpropionylarnides endowed with mixed ACE/NEP inhibitory activity.
The European patent application No. 0524553 (Thstitut National de la Sante et de la Recher¬ che Medicale (INSERM)] describes acylmercaptoalkanoyldipepudes endowed with neutral endopeptidase and peptidyldipeptidase A inhibitory activity. The European patent application No. 0566157 (Schering Coφoration) describes thiol deriva- tives such as, for instance, N-mercaptoacyl amino acids, endowed with NEP-inhibitory activ¬ ity. α-Mercaptoacyl dipeptides endowed with ACE-inhibitory and NEP-inhibitory activity are also described by S.S. Bhagwat et al. in Bioorganic & Medicinal Chemistry Letters, 7, 735-738, 1995. In this last work the authors conclude that, while the presence of a biphenylmethyl group con¬ fers an interesting mixed ACE/NEP-inhibitory activity at the molecules having an α-mercap- toacyl dipeptide structure, the substitution ofthe biphenyl group with groups such as α- or β- naphthyl causes a significant loss of activity. Now we have found N-mercaptoacyl phenylalanine derivatives which are endowed with a remarkable inhibitory activity on the angiotensin converting enzyme as well as on the neutral endopeptidase enzyme (mixed or dual ACE/NEP inhibitory activity) which makes them par¬ ticularly useful in the therapy of cardiovascular pathologies. Therefore, object ofthe present invention are the compounds of formula 1
R-CH2-CH-CONH-CH-COOR2 (I) * I
CH2-R3 wherem
R is a mercapto group or a R4COS group convertible in the organism to mercapto group,
Rj is a straight or branched C2-C4 alkyl group or an aryl or arylalkyl group having from 1 to 6 carbon atoms in the alkyl moiety wherein the aryl is a phenyl or a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consisting of mtrogen, oxygen and sulphur, optionally substituted with one or more substituents, the same or different, selected from the group consisting of halogen atoms, hydroxy groups, alkoxy, alkyl, alkylthio, alkylsulphonyl or alkoxycarbonyl groups having from 1 to 6 carbon atoms in the alkyl moiety, C1-C3 alkyl groups containing one or more fluorine atoms, carboxy groups, nitro groups, ammo or aminocarbonyl groups, acylarruno groups, aminosulphonyl groups, mono- or -alkylamino or mono- or di-alkylarrunocar- bonyl groups havmg from 1 to 6 carbon atoms in the alkyl moiety,
R2 is a hydrogen atom, a straight or branched C1-C4 alkyl or a benzyl group, R3 is a phenyl group substituted with a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consisting of nitrogen, oxygen and sulphur, being the phenyl and the heterocyclic groups optionally substituted with one or more substituents, the same or different, as indicated for R] ,
R4 is a straight or branched C1-C4 alkyl group or a phenyl group, d e carbon atoms marked with an astensk are stereogenic centers, and pharmaceutically acceptable salts thereof
The compounds of formula I contam two stereogenic centers and can thus exist in the form of stereoisomers
Therefore, object of the present mvenUon are the compounds of formula I in the form of stereoisomeπc mixture as well as in the form of single stereoisomers
The compounds of formula I object of the present invention are endowed with a mixed
ACE NEP-inhibitory activity and are useful in the treatment of cardiovascular diseases
Although both terms mixed and dual are indifferently used, the term dual is the one more commonly accepted in the literature for compounds endowed contemporaneously with ACE- and iNEP-inhibitory activity
In the present context, the terms mixed and dual are to be considered equivalent In the present descπption, unless otherwise specified, with the term straight or branched alkyl group we intend an alkyl such as methyl, ethyl, n propyl, isopropyl, n butyl, sec-butyl, tert- butyl, isobutyl, n pentyl, 2-pentyl, 3-pentyl, isopentyl, tert-pentyl, n hexyi and isohexyl group, with the term straight or branched alkoxy group we intend an alkoxy such as methoxy, ethoxy, n propoxy and isopropoxy group, with the term halogen atom we intend a fluorine, chlorine, bromme or iodine atom, with the term acyl we intend an acyl group deπvmg from an aliphatic or aromatic carboxylic acid such as acetic, propionic, butyπc and benzoic aαd, with the term 5 or 6 membered aromaϋc heterocycle corrtaining 1 or 2 heteroatoms selected among nitrogen, oxygen and sulphur we intend a group such as thiazole, isoxazole, oxazoie, isothiazole, pyra- zole, imidazole, thiophene, pyrrole, pyndine, pyπmidme, pyrazme, pyndaz e and furan, op- tionally benzocondensed
Examples of pharmaceutically acceptable salts of the compounds of formula I are the salts with alkali or alkali-earth metals and the salts with pharmaceutically acceptable organic bases Preferred compounds of formula I are the compounds wherem R. is a phenyl group substi¬ tuted in position 4 with a heterocyclic group More preferred compounds, in this class, are the compounds of formula I wherein R repre¬ sents an arylalkyl group optionally substituted with one or more substituents, the same or dif¬ ferent, selected from the group consisting of halogen atoms, hydroxy, alkyl or alkoxy Still more preferred compounds, in this class, are the compound of formula I wherein R] rep¬ resents a phenylalkyl group optionally substituted with one or more substitutents, the same or different, selected from the group consisting of halogen atoms, hydroxy, alkyl or alkoxy
Preferred examples of pharmaceutically acceptable salts of the compounds of formula I are the salts with alkali metals such as sodium, lithium and potassium
It is clear to the man skilled in the art that the compounds of formula I wherem R is a R4COS group convertible in the organism to mercapto group, as well as the compounds of formula I wherem R is an alkyl or benzyl group, are biologic precursors (pro-drugs) ofthe correspond¬ ing compounds of formula I wherem R is a mercapto group (R=-SH) or R is a hydrogen atom (R =H), respectively
Specific examples of preferred compounds of formula I, object of die present mvention, are N-(3-phenylcarbonyllhιo-2-phenylme ιylpropιonyl)-4-(2-thιazolyl)-phenylalamne methyl es- ter,
N-(3-phenylv-*arbonyluucH2-phenylmet ylpropiOT ester,
N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyl)-4-(3-pyπdyl)-phenylalanme benzyl ester, N-(3-phenylcarbonylthιo-2-phenylme ιylpropιonyl)-4-(2-furyl)-phenyialanme methyl ester, N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyl 4-(5-pyπrmdmyl)phenylalarune ethyl es- ter,
N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyl)-4-(2-pyrazmyl)-phenylalamne methyl es¬ ter,
N-(3-phenylcarbonylu^o-2-phenylmethylprcφιonyl)-4-(2-thjenyl)-phenylalanme methyl ester, N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyl)-4-(3-thιenyl)-phenylalanme methyl ester, N-(3-phenylcarbonylthiCr-2-phenylmethylpropionyl)-4-(3-furyl)-phenylalanme methyl ester,
N-[3-phenylcarbonylthιo-2-(3-pyπdylmeΛyl)propιonyl]-4-(2-thιazolyl)-phenylalan e methyl ester,
N-[3-acetylthιo-2-(3-memoxyphenyl)memyl-propιonyl]-4-(2-thιazolyl)-phenylalan e methyl ester, N-[3-acetyltmo-2-(2-fluorophenyl)memyl-propιonyl]-4-(2-thιazolyl)-phenylalanme methyl es¬ ter,
N-[3-phenylcarbonyltdτio-2-(2-ti^enyl)methyl-propιonyl]-4-(2-tlτjazolyl)-phenylalanme methyl ester, N-[2-(2-furyl)methyl-3-phenyl(»rbonylthιo-propιonyl]-4-(2-thιazolyl)-phenylalanme methyl ester,
N-[2-(3-methyl-5-ιsoxazolyl)meuιyl-3-phenylcarbonylthιo-propιonyl]-4-(2-thιazolyl)-phenyl- alanine methyl ester,
N-(3-mercapto-2-phenylmethyipropιonyl)-4-(2-thιazolyl phenylalarune, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-pyπdyl)-phenylalanme, N-(3-mercapto-2-phenylmethylpropιony])-4-(3-pyridyl)-phenylalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-furyl)-phenylalamne, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(5-pyπn-ud yl)-phenylalanme,
N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-pyrazmyl)-phenylalanme,
N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-thιenyl)-phenylalamne, N-(3-mercapto-2-phenylmethylprcφιonyl)-4-(3-thιenyl)-phenylalaι*ιine,
N-(3-mercapto-2-phenylmethylpropιonyl)-4-(3-furyl)-phenylalanme,
N-[3-mercapto-2-(3-pyπdylmethyl)propιonyl]-4-(2-thιazolyl)-phenylalanme,
N-[3-mercaptcκ2-(3-methoxyphenyl)methyl-propιonyl]-4-(2-lhιazolyl)-phenylalarune,
N-[3-mercapto-2-(2-tiuenyl)methyl-propιonyl]-4-(2-thιazolyl)-phenylalanme, N-[3-mercapto-2-(3-memyl-5-ιsoxazolyl)methyl-propιonyI]-4-(2-thιazolyl)-phenylalanme,
N-[2-(2-fluorophenyl)methyl-3-mercapto-propιonyl]-4-(2-thιazolyl)-phenylalanme,
N-[2-(2-fuιyl)methyl-3-mercapto-propιonyl]-4-(2-thιazolyl)-phenylalanme
The preparation of the compounds of formula I, object ofthe present mvention, is earned out according to a synthetic process compπsmg the reaction between a compound of formula
R-CH2-
Figure imgf000008_0001
* wherem R and Ri have the above reported meanings, and a phenylalanine deπvative of formula
H2N-CH-COOR2 (IH) CH2-R3 wherem
R2 and R3 have d e above reported meanings
The condensation reaction is earned out accordmg to conventional techniques of the chemistry of peptides Before carrymg out the reaction it can be useful to properly protect the optional functional groups which could interfere in the reaction
The optional protection is earned out accordmg to conventional techniques In this respect the compounds wherem R is a R4COS group are preferably used as intermedi¬ ates of formula II, thus obtaining the correspondmg compounds of formula I wherem R=R4COS from which, by hydrolysis, the compounds of formula I wherem R=SH can be obtained
The evaluation of the usefulness of the optional protection as well as the selection ofthe kind of adopted protection, according to the reaction to be earned out and to the functional groups to be protected, are within the normal knowledge ofthe man skilled m the art T e removal of the optional protective groups is earned out according to conventional tech¬ niques For a general reference to the use of protective groups in organic chemistry see Theodora W Greene and Peter G M Wuts "Protective Groups Organic Synthesis", John Wiley & Sons, Inc , π Ed , 1991
The compounds of formula II are known or easily prepared accordmg to conventional methods as descnbed, for instance, m Bntish patent No 1576161 in die name of E R Squibb & Sons Inc Altematively, tiie compounds of formula II can be prepared accordmg to the synd etic meth¬ ods descnbed by M C Foumie-Zalusky et al. in J Med Chem 1994, 37, 1070-1083 Also the intermediate compounds of formula HI are known or easily prepared accordmg to known methods For instance, the compounds of formula UJ can be prepared accordmg to the synthetic meth- ods descnbed by W C Shieh et al m J Org Chem 1992, 57, 379-381
Alternatively, die compounds of formula HI can be prepared by coupling methods (cross- coupling) starting from halogenated heterocyclic compounds and stannyl phenylalanine de¬ πvatives, as descnbed by D.S Wilbur et al in Bioconjugate Chem , 1993, 4, 574-580 The compounds of formula I in the form of single stereoisomers are prepared by stereoselec- tive synthesis or by separation of the stereoisomenc mixture accordmg to conventional tech¬ niques
Also the preparation of the salts of the compounds of formula I, object of the present mven¬ tion, is earned out accordmg to conventional techniques The compounds of formula I, object of the present invention, are endowed with a mixed ACE/NEP-inhibitory activity and are useful m the treatment of cardiovascular diseases
The inhibitory activity ofthe compounds of formula I was evaluated by means of in vitro tests (example 8)
In particular, the inhibitory activity ofthe compounds of formula I was evaluated m compan- son to the aforementioned Thioφhan and Captopπl The vitro inhibitory activity of the compounds of formula I, expressed as IC50 value, re¬ sulted at nM concentrations
Said activity resulted to be comparable to that of Captopnl, to what it concerns me ACE-in- hibitory activity, and greater man that of Thioφhan, to what it concerns the NEP-inhibitory activity The inhibitory activity of me compounds of formula I, moreover, was also evaluated by means of ex- vivo tests in compaπson to known compounds (example 9)
In particular, N-[3-mercapto-2-(3,4-methylenedιoxyphenyl)methyl-propιonyl]-(S)-phenylala- mne and N-[3-merv**apto-2-(3,4-medιylenedιoxyphenyl)methyl-propιonyl]-(S)-tyrosme, de¬ scnbed as enkephalinase and ACE-inhibitors in me aforementioned European patent applica- tion No 0419327 and hereinafter referred to as compounds R-l and R-2 respectively, as well as N-(3-mercapto-2-methylpropanoyl)-L-tyrosιne and N-(3-mercapto-2-methylpropanoyl)-L- tπptophan, descnbed as ACE-inhibitors U S 4,199,512 and as enkephalinase inhibitors in U S 4,401,677 and hereinafter referred to as compounds R-3 and R-4 respectively, were used as compaπson compounds The ex-vivo ACE/NEP-inhibitory activity, in particular, was determined by evaluating the en¬ zymatic activity m tissue homogenates (lung and kidney for ACE and NEP activity, respec¬ tively) from spontaneously hypertensive rats (SHR) treated with the tested compounds by in¬ travenous or oral route It is worth noting that the activity shown by the compounds of formula I m the ex vivo tests confirms the mixed activity (dual activity) shown in the in vitro tests, also after oral admini¬ stration
Said activity, moreover, resulted to be significantly higher than that of the compaπson com¬ pounds For a practical use in therapy, the compounds of formula I can be formulated in solid or liquid pharmaceutical compositions, suitable to oral or parenteral administration
Therefore, the pharmaceutical compositions containing a therapeutically effective amount of a compound of formula I admixture with a earner for pharmaceutical use are a further object ofthe present mvention
Specific examples of pharmaceutical compositions accordmg to the present mvention are tab- lets, coated tablets, capsules, granulates, solutions and suspensions suitable to oral admini¬ stration, solutions and suspensions suitable to parenteral administration The pharmaceutical compositions object of me present mvention are prepared according to conventional techniques The daily dose ofthe compound of formula I or of me correspondmg pro-drug will depend on several factors such as the seπousness of the ώsease, the mdividual response of the patient or the land of formulation but it is usually compπsed between 0 1 mg and 10 mg per kg of body weight divided mto a single dose or mto more daily doses
With the aim of illustrating the present invention the following examples are now given Unless otherwise specified, the flash chromatographies were earned out by usmg flash chro- matography silica gel from Baker company (code 7024-00)
Example 1 Preparation of N-tert-butoxycarbonyl-4-(5-pynmιdmyl)-L-phenylalamne ethyl ester A mixture of 5-pyπmιdιnylboronjc aαd (850 mg, 2 mmoles), N-tert-butoxycarbonyl-4-tπ- fluoromethylsulphonyl-L-phenylalanme ethyl ester (450 mg, 2 2 mmoles), a solution of so- dium carbonate (530 mg) in water (2 59 ml) and a mixture of toluene ethanol=10 4 5 (20 ml) was degassed with nitrogen for 30 mmutes
Subsequently, palladium(0)tetrakis(tnphenylphosprune) (120 mg, 0 06 mmoles) was therein added and the reaction mixture was heated at 90°C and kept under stirring for 3 hours The mixture was then kept at room temperature and N-tert-butoxycarbonyl-4-tnfluorometh- ylsulphonyl-L-phenylalanme ethyl ester (112 mg) and palladium(0)tetrakis- (tπphenylphosphine) (30 mg) were added agam
The mixture was further heated at the temperature of 90°C and kept under stirring for other 18 hours After cooling the reaction mixture at room temperature, ethyl acetate (100 ml) and water (40 ml) were added
The phases were separated and the organic phase was dπed on sodium sulphate and evapo- rated under vacuum The obtamed residue was punfied by flash chromatography (silica gel, eluent hexane ethyl acetate=7 3, pressure of nitrogen 0 1 atm) thus affording N-tert-butoxy- carbcιnyl-4^5-pyπrradmyl)-L-phenylalarune ethyl ester (130 mg, 14% yield) as a colourless oil
^-NMR (200 MHz, CDCy δ (ppm) 1 24 (t, 3H, CH2-CH3), 1 40 [s, 9H, C(CH3)3], 3 01-3 25 (m, 2H, CH2-CH), 4 19 (q, 2H, CH2-CH3), 4 52-465 (m, IH, CH-COO), 5 06 (d, IH, NH), 7 25-7 52 (m, 4H, phenylene), 8 91 (s, 2H, N-CH-C-CH-N), 9 19 (s, IH, N-CH- N) Example 2
Preparation of N-tert-butoxycarbonyl-4-(2-thiazolylVL-phenylalamne methyl ester N-tert-butoxycarbonyl-4-(tπfluoromethylsulphonyl)-L-phenylalanme methyl ester (8 g, 32 3 mmoles) and palladιum-bιs(tnphenylphosphme) chloπde (2 3 g) were added to a solution of 2- tnmethylstannyl-thiazole (13 8 g, 32 3 mmoles) in a mixture of tetrahydrofuran toluene=10 1 (50 ml), previously degassed with nitrogen
The mixture was refluxed for 24 hours and, subsequendy, 2-tnmethylstannyl-thιazole (2 g) was added agam
After 6 hours at reflux N-tert-butoxycarbonyl-4-(tπfluoromethylsulphonyl)-L-phenylalanme methyl ester (2 g) and palladιum-bιs(tnphenylphosphιne) chlonde (700 mg) were added The resultant reaction mixture was kept under stirnng at 70°C for 16 hours and, subsequently, cooled at room temperature
Water (200 ml) was men added to me mixture which was extracted with methylene chlonde (4x200 ml) The collected organic phases were dned on sodium sulphate and evaporated under vacuum The obtamed residue was punfied by flash chromatography (silica gel, eluent methylene chlo¬ nde ethyl acetate=9 1, pressure of nιtrogen=0 1 atm) thus affording N-tert-butoxycarbonyl-4- (2-thιazolyl)-L-phenylalanme methyl ester (2 3 g, 20% yield)
^-NMR (200 MHz, CDC13) δ (ppm) 1 40 [s, 9H, C(CH3)3], 3 00-3 21 (m, 2H, CH2), 3 70 (s, 3H, COOCH3), 4 42-4 65 (m, IH, CH-COO), 5 02 (bd, IH, NH), 7 30 (d, IH, S- CH-CH-N), 7 81 (d, IH, S-CH-CH-N), 7 15-7 90 (m, 4H, phenylene)
Example 3 Preparation of N-tert-butoxycarbonyl-4-(3-pyπdylVL-phenylalan e methyl ester 3-Bromopyπdme (1 67 g, 10 mmoles) and pallaώum(0 etrakιs(tnphenylphosphιne) (370 mg, 0 219 mmoles) were added to a solution of N-tert-butoxycarbonyl-4-(tπbutylstannyl)-L- phenylalanine methyl ester (4 g, 7 03 mmoles), prepared as descπbed by D S Wilbur et al in Bioconjugate Chem , 1973, 4, 574-580, m anhydrous dimethylformamide (30 ml), previously degassed with nitrogen
The reaction mixture was kept under stirring for 10 mmutes at room temperature and, subse¬ quently, heated at 105°C for 6 hours Palladium(0)tetrakis(tnphenylphospliine) (0 0035 mmoles) was agam added and the mixture was kept under stirring at 105°C for 8 hours and then cooled at room temperature Water (100 ml) was mere added and me reaction mixture was extracted with hexane (3x150 ml) The collected organic phases were washed with a saturated aqueous solution of potassium fluonde, dπed on sodium sulphate and evaporated under vacuum The obtamed residue was collected with ethyl acetate and filtered
The resultant solution was evaporated under vacuum and the residue was punfied by flash chromatography (silica gel, eluent hexane ethyl acetate=8 2, pressure of nitrogen 0 1 atm) af¬ fording N-tert-butoxycarbonyl-4-(3-pyndyl)-L-phenylalanme methyl ester (1 5 g, 60% yield) as a colourless oil
Mass (C I ) (M+H)+=357
^-NMR (200 MHz, CDC13) δ (ppm) 1 40 [s, 9H, C(CH3)3], 3 00-3 20 (m, 2H, CH2), 3 71 (s, 3H, COOCH3), 4 55 (m, IH, CH-COO), 5 05 (bd, IH, NH), 7 19-7 51 (m, 4H, phenylene), 7 30-8 82 (bm, 4H, pyndyl) By working in an analogous way the following compounds were prepared N-tert-butoxycarbonyl-4-(2-pyndyl)-L-phenylalanme methyl ester Mass (C I ) (M+H)+=357
]H-NMR (200 MHz, CDC13) δ (ppm) 1 40 [s, 9H, C(CH3)3], 3 03-3 21 (m, 2H, CH CH), 3 70 (s, 3H, COOCH3), 4 54-4 65 (m, IH, CH-COO), 4 98 (d, IH, CONH), 7 16-7 21 (m, IH, N-C-CH-CH), 7 65-7 77 (m, 2H, N-CH-CH-CH), 7 19-7 93 (m, 4H, phenylene), 8 63-8 68 (m, IH, N-CH), N-tert-butoxycarbonyl-4-(2-p azmylVL-phenylalarιιne methyl ester
^-NMR (200 MHz, CDC13) δ (ppm) 1 40 [s, 9H, C(CH3)3], 3 02 (m, 2H, CH2), 3 70 (s, 3H, COOCH3), 4 53-4 70 (m, IH, CH-COO), 5 03 (bd, IH, NH), 7 21-7 98 (m, 4H, phenylene), 8 49 and 8 62 [2(bs, 2H, N-CH-CH-N)], 9 00 (s, IH, CH-N-CH-CH), N-tert-butoxycarbonyl-4-(2-thιenyl)-L-phenylalarι ne methyl ester
!H-NMR (200 MHz, CDC13) δ (ppm) 1 41 [m, 9H, C(CH3)3], 2 98-3 18 (m, 2H, CH2), 3 71 (s, 3H, COOCH3), 4 53-4 64 (m, IH, CH-COO), 4 98 (bd, IH, NH), 7 02-7 28 (m, 3H, thienyl), 7 10-7 54 (m, 4H, phenylene) Example 4
Preparation of 4-(3-pyπdyl)-L-phenylalanιne methyl ester dihvdrochloπde Thionyl chlonde (0 85 ml, 4 78 mmoles) was added dropwise to a solution of N-tert-butoxy- carbonyl-4-(3-pyndyl)-L-phenylalarune methyl ester (1 4 g, 3 93 mmoles) in methanol (30 ml), prepared as descπbed in example 3, keeping the temperature at 0°C At the end of me addition, the reaction mixture was brought at room temperature and kept under stirring for 8 hours
The solvent was then evaporated under vacuum affording 4-(3-pyndyl)-L-phenylalanme methyl ester dihydrochloπde (820 mg, 71% yield) used as such in d e following reactions Mass (C I ) (M+H)+= 257 (free base) !H-NMR (200 MHz, D20) δ (ppm) 3 11-3 32 ( , 2H, CH ), 3 67 (s, 3H, CH3), 4 31-4 37 (m, IH, CH), 7 30-7 63 (m, 4H, phenylene), 7 97 (dd, IH, CH-N-CH-CH-CH), 8 59 (d, IH, CH-N-CH-CH-CH), 8 65-8 71 (m, IH, CH-N-CH-CH), 8 89 (d, IH, CH-N-CH-CH-CH) By working m an analogous way the following compounds were prepared 4-(2-pyndyl)-L-phenylalanιne methyl ester dihvdrochlonde Mass (C I ) (M+H)+= 257 (free base)
1H-NMR (200 MHz, D20) δ (ppm) 3 12-3 33 (m, 2H, CH2), 3 64 (s, 3H, CH3), 4 30-4 37 (m, IH, CH), 7 36-7 73 (m, 4H, phenylene), 7 78-8 58 (m, 4H, pyπdyl), 4-(5-pynmιdmyl)-L-phenylalanme ethyl ester dihvdrochloπde ^-NMR (200 MHz, DMSO-dg) δ (ppm) 1 11 (t, 3H, CH2-CH3), 3 10-3 35 (m, 2H, CH2- CH), 4 04-4 20 (m, 2H, CH2-CH3), 4 22-4 35 (m, IH, CH), 7 40-7 84 (m, 4H, phenylene), 9 15 (s, 2H, N-CH-C-CH-N), 9 19 (s, IH, N-CH-N), 4-(2-pyrazmyl)-L-phenylalan ne methyl ester dihydrochlonde
Mass (C I ) (M+H)+= 258 (free base)
^-NMR (200 MHz, DC1 IN) δ (ppm) 3 45-3 67 (m, 2H, CH2), 3 98 (s, 3H, CH3), 4 65- 4 72 (m, IH, CH), 7 68-8.26 (m, 4H, phenylene), 9 02 (d, IH, CH-N-CH-CH), 9 44 (dd, IH,
CH-N-CH-CH), 9 54 (d, IH, CH-N-CH-CH),
4-(2-thιenyl)-L-ρhenylalanme methyl ester hvdrochloπde
Mass (C I ) (M+H)+= 262 (free base)
!H-NMR (200 MHz, D20) δ (ppm) 2 98-3 19 (m, 2H, CH2), 3 65 (s, 3H, CH3), 4 21-4 28 (m, IH, CH), 6 96-7 28 (m, 3H, thienyl), 7 08-7 52 (m, 4H, phenylene),
4-(2-thιazolviyL-phenylalanιne methyl ester dihydrochlonde
^-NMR (200 MHz, D20) δ (ppm) 3 10-3 32 (m, 2H, CH2-CH), 3 68 (s, 3H, CH3), 4 30-
4 38 (m, IH, CH), 7 30-7 80 (m, 4H, phenylene), 7 70-7 91 (m, 2H, thiazolyl)
Example 5 Preparation of 2-ιsobutyl-3-phenylcarbonylthιo-propιonιc aαd
A mixture of 2-ιsobutyl-acrylιc aαd (6 34 g, 49 mmoles) and thiobenzoic aαd (5 96 ml, 51 mmoles) was heated at 100°C under stirring for 2 hours
The reaction mixture was then treated with petroleum ether 40-60°C (100 ml) and cooled at
-70°C in a dry ice acetone bath By filtration and washmg with petroleum ether at -70°C a residue was collected which, dπed at reduced pressure, furnished 2-ιsobutyl-3-phenylcarbonylthιo-propιonιc aαd (11 12 g, 85% yield) as a white sohde
^-NMR (200 MHz, CDC13) δ (ppm) 0 90-1 00 (m, 6H), 1 40-1 90 (m, 3H), 2 70-2 90 (m,
IH), 3 10-3 40 (m, 2H), 7 35-7 62 (m, 3H), 7 90-8 00 (d, 2H) By working m an analogous way the following compounds were prepared
2-(3-memoxyphenyl)methyl-3-phenylcarbonylthιo-propιonιc aαd lH-NMR (200 MHz, CDC13) δ (ppm) 2 32 (s, 3H), 2 80-3 15 (m, 5H), 3 77 (s, 3H), 6 70-
6 80 (m, 3H), 7 14-7 25 (m, IH)
3-acetylthιo-2-(2-fluorophenyl)methyl-propιonιc aαd ^^--NNMMRR ((220000 MMHHzz,, CCDDCC1133)) δδ ((ppppmm)) 22 3311 ((ss,, 33HH)),, 2 90-3 20 (m, 5H), 6 95-7 30 (m, 4H) 3-phenylcarbonylthιo-2-(2-thιenyl)methyl-propιonιc aαd benzylamme salt H-NMR (200 MHz, DMSO-d6) δ (ppm) 2 45-3 25 (m, 5H, S-CH2-CH-CH2), 3 90 (s, 2H,
CH2-phenyl), 6 85-7 91 (m, 13H, aryl)
Example 6 Preparation of N-r(2SV3-phenyl(arbonylthιo-2-phOTylmemylpropιonyl1-4-f2-thιazolyl'r-L- phenylalanine methyl ester (compound 1)
A solution of hydroxybenzotnazole (0 54 g, 4 mmoles) in tetrahydrofuran (30 ml) and, subse¬ quently, a solution of dicyclohexylcarbodnmide (0 825 g, 4 mmoles) m methylene chlonde (15 ml) were added, at 0°C under stirring, to a mixture of (2S)-3-phenylcarbonylthιo-2-phenyl- methylpropionic aαd (1 2 g, 4 mmoles), 4-(2-thιazolyl)-L-phenylalanιne methyl ester dihydro¬ chlonde (1 34 g, 4 mmoles), prepared as descπbed in example 4, tπethylam e (1 11 ml, 8 mmoles) in tetrahydrofuran (20 ml) and methylene chlonde (30 ml)
The reaction mixture was kept under stirring for 20 hours, then dicyclohexyl urea was filtered off and the solvent was evaporated at reduced pressure The residue was collected with ethyl acetate and the solution was washed with an aqueous solution of sodium chlonde at 20%, sodium bicarbonate at 5% and sodium chlonde at 20% agam
After separation of the phases and evaporation of die organic phase, die resultant white solid was punfied by flash chromatography (silica gel, eluent ethyl acetate hexane=40 60, pressure of nitrogen 0 1 atm) uius affordmg N-[(2S)-3-phenylcarbonylthιo-2-phenylmethylpropιonyl]-
4-(2-thιazolyl)-L-phenylalanιne methyl ester (1 5 g) m p 98-100°C
Mass (C I ) (M+H)+= 545
^-NMR (200 MHz, CDCl3) δ (ppm) 2 63-3 35 (m, 7H, CH2-CH-CH2, CH2-C6H4-thιa- zolyl), 3 68 (s, 3H, COOCH3), 4 75-4 85 (m, IH, CH-COO), 5 78 (d, IH, NH), 7 10-8 00
(m, 16H, aryl)
By working m an analogous way, starting from compounds known or prepared as descπbed m examples 4 and 5, the following compounds were obtamed
N-f(2SV3-phmylcarbonylthιo-2-phenylmemylpropιonyll-4-(2-furyl'r-L-phenylalanιne methyl ester (compound 2) m p 132-134°C Mass (C I ) (M+H)+= 528
^-NMR (200 MHz, CDC13) δ (ppm) 2 60-3 35 (m, 7H, CH2-CH-CH2, CH-j-C^- furyl), 3 58 (s, 3H, COOCH3), 4 71-4 81 (m, IH, CH-COO), 5 73 (d, IH, NH), 6 40-8 00 (m, 17H, aryl),
N- f (2S V3 -phenylcarbonylthιo-2-phenylmethylp ropi onyl ]-4-(5-p vnmidinyl VL-phenylalanine ethyl ester (compound 3) m p 117-119°C
Mass (C I ) (M+H)+= 554 !H-NMR (200 MHz, CDC13) δ (ppm) 1 18 (t, 3H, CH3-CH2), 2 65-3 35 (m, 7H, CH2- CH-CH2, CH2-C6H4-pynιτudιnyl), 3 95-4 20 (m, 2H, COOCH2), 4 70-4 80 (m, IH, CH- COO), 5 78 (d, IH, NH), 7 05-8 00 (m, 14H, phenyl, phenylene), 8 68 (s, 2H, CH-N-CH-N- CH), 9 11 (s, IH, N-CH-N), N-l(2S)-3-phenylcarbonylthio-2-phenylmethylpropionyll-4-(2-pyrazmyl)-L-phenylalanme methyl ester (compound 4) m p 145-147°C Mass (C I ) (M+H)+= 540
^-NMR (200 MHz, CDC13) δ (ppm) 2 65-3 35 (m, 7H, CH2-CH-CH2, C^-C^- pyrazmyl), 3 61 (s, 3H, COOCH3), 4 75-4 85 (m, IH, CH-COO), 5 78 (d, IH, NH), 7 10- 8 00 (m, 14H, phenyl, phenylene), 8 48-8 75 (m, 3H, CH-N-CH-CH-N),
N-r(2S)-3-phenylcarbonylthιo-2-phenylmethylpropιonvπ-4-(3-pyπdyl)-L-phenylalanme methyl ester (compound 5) m p 132-134°C
Mass (C I ) (M+H)+= 539 !H-NMR (200 MHz, CDC13) δ (ppm) 2 66-2 79 ( , IH, CH2-CH-CH2), 2 88-3 35 (m, 6H, CH2-CH-CH2, CH2-C6H4-pyndyi), 3 61 (s, 3H, COOCH3), 4 75-4 85 (m, IH, CH- COO), 5 77 (d, IH, NH), 7 07-7 99 (m, 16H, CH-CH-CH-N, phenyl, phenylene), 8 51-8 64 (m, 2H, CH-N-CH), N-r(2S)-3-phenylcarbonylthjo-2-phenylmethylpropιonyll-4-('2-pyπdyl')-L-phenylalanme methyl ester (compound 6) m p 123-125°C Mass (CI.) (M+H)+= 539
^-NMR (200 MHz, CDC13): δ (ppm): 2.63-2.77 (m, IH, CH2-CH-CH2); 2.85-3.35 (m,
6H, CH2-CH-CH2, CH2-C6H4-pyridyl); 3.56 (s, 3H, COOCH3); 4.75-4.85 (m, IH, CH- COO); 5.75 (d, IH, NH); 7.09-7.99 (m, 16H, CH-CH-CH-N, phenyl, phenylene); 8.61-8.65
(m, IH, N-CH-CH);
N-f(2SV3-phenylcarbonyluιio-2-phenylme ylpropionyll-4-(2-thien^ ester (compound 7)
Mass (CI.) (M+H)+= 544 !H-NMR (200 MHz, CDC13): δ (ppm): 2.64-3.36 (m, 7H, CH2-CH-CH2, CONH-CH-CH2);
3.58 (s, 3H, COOCH3); 4.74-4.83 ( , IH, CH-COO); 5 74 (d, IH, NH); 6.97-7 99 ( , 17H, aryl);
N-f(2S)-3-phenylcarbonylthio-2-phenylmethylpropionyll-4-(3-thienyl)-L-phenylalan e methyl ester (compound 8) Mass (CI.) (M+H)+= 544
^-NMR (200 MHz, CDC13): δ (ppm): 2.63-3.35 (m, 7H, CH2-CH-CH2, NH-CH-CH2);
3.58 (s, 3H, COOCH3); 4.73-4.85 (m, IH, CH-COO); 5.70-5.76 (bd, IH, NH); 7.00-7.62
(m, 15H, phenyl, phenylene, CH-CH-S); 7.93-8.00 (m, 2H, CH-S-CH);
N-[(2S)-3-phenylcarbonyldιio-2-phenylmemylpropionyl -4-(3-furyl)-L-phenylalanine methyl ester (compound 9) m.p. 115-117°C
Mass (CI.) (M+H)+= 528
^-NMR (200 MHz, CDC13): δ (ppm): 2.62-3.36 (m, 7H, CH2-CH-CH2, NH-CH-CH2);
3.58 (s, 3H, COOCH3); 4.71-4.82 (m, IH, CH-COO); 5.72 (bd, IH, NH); 6.50-8.00 (m, 17H, aryl);
N-f3-phenylcarbonylthio-2-(3-pyridylmemyl)propionyll-4-(2-thiazolyl)-L-phenylalanine methyl ester - stereoisomer A (compound 10)
Mass (CI.) (M+H)+= 546
TLC (ethyl acetate:petroleum ether=95:5), Rf = 0.33 ^^--NNMMRR ((220000 MMHHzz,, CCDDCC1133)):: δδ ((ppppmm)):: 22..6600--33..3388 (m, 7H, CH2-CH-CH2, NH-CH-CH2); 3.60 (s, 3H, COOCH3); 4.79-4.90 (m, IH, CH-COO); 6.21 (bd, IH, NH); 7.29-7 81 (m, 2H, thiazolyl), 6 75-8 48 (m, 13H, phenyl, phenylene, pyndyl),
N-[3-phenylcarbonylt o-2-f3-pyπdylmemyl)propιonyl]-4-f2-thιazolylVL-phenylalan e methyl ester - stereoisomer B (compound 11) Mass (C I ) (M+H)+= 546
TLC (ethyl acetate petroleum ether=95 5), Rf = 0 24 lH-NMR (200 MHz, CDC13) δ (ppm) 2 61-3 25 (m, 7H, CH2-CH-CH2, NH-CH-CH2),
3 60 (s, 3H, COOCH3), 4 80-4 91 (m, IH, CH-COO), 6 09 (bd, IH, NH), 7 27-7 80 (m, 2H, thiazoly), 7 10-8 40 (m, 13H, phenyl, phenylene, pyndyl), N-[2-ιsobutyl-3-phenylcarbonylthjc>-propιonyll-4-(2-thιazolyl)-L-phenylalamne methyl ester
(compound 12)
^-NMR (200 MHz, CDC13) δ (ppm) 0 80-0 95 (m, 6H), 1 30-1 80 (m, 3H), 2 40-2 60 (m,
IH), 3 00-3 30 (m, 4H), 3 70 (d, 3H), 4 90-5 05 (m, IH), 6 00-6 15 (bt, IH), 7 10-8 00 (m,
HH), N-r3-av^ylιdιιo-2-(3-memoxyphenyl)memyl-propιcmyll-4-(2-tmazolyl)-L-phenylalanme methyl ester (compound 13)
!H-NMR (200 MHz, CDC13) δ (ppm) 2 30 (d, 3H), 2 45-3 20 (m, 7H), 3 63 (d, 3H), 3 75
(d, 3H), 4 70-4 93 (m, IH), 5 70-5 90 (dd, IH), 6 60-6 85 (m, 4H), 7 17-7 32 (m, 3H), 7 68-
7 90 (m, 3H), N-[3-acetylthιo-2-(3-fluorophenyl)methyl-propιonyll-4-(2-thιazolyl)-L-phenylalanme methyl ester (compound 14)
]H-NMR (200 MHz, CDC13) δ (ppm) 2 29 (s, 3H), 2 55-3 20 (m, 7H), 3 65 (2s, 3H), 4 70-
4 90 (m, IH), 5 80-6 00 (2d, IH), 6 70-7 32 (m, 3H), N-f3-phenylcarbonyldιιo-2-(2-thjenyl)methyl-propιonyll-4-(2-thιazolyl)-L-phenylalanme methyl ester (compound 15)
^-NMR (200 MHz, CDC13) δ (ppm) 2 68-3 37 (m, 7H), 3 60-3 61 (2s, 3H, COOCH3),
4 31-4 45 (m, IH, CH-COOCH3), 6 01-6 10 (2d, IH, NH), 6 80-8 00 (m, 14H),
Example 7
Preparation of N-r(2SV3-mercapto-2-phenylmethylpropionyll-4-(2-thiazolyl)-L-phenylalanme (compound 16)
N-[(2S)-3-phenylcarbonyldτιo-2-phenylmethylpropιonyl]-4-(2-thιazolyl)-L-phenylalanme ethyl ester (1.4 g; 2.57 mmoles), prepared as described in example 6, was suspended in ethanol (30 ml), degassed by nitrogen bubbling to eliminate the oxygen.
An aqueous degassed solution of sodium hydroxide IN (7.7 ml) and, at the end of the addi- tion, further degassed ethanol (20 ml) were added dropwise at 5°C to me suspension.
The reaction mixture was kept under stirring for 4 hours at room temperature, then cooled at
0°C and acidified with hydrochloric acid 5% (10 ml).
The reaction mixture was evaporated to dryness and the residue was collected witii acetonitrile and water and, subsequently, was filtered thus affording N-[(2S)-3-mercapto-2-phenylmemyl- propιonyl]-4-(2-thiazolyl)-L-phenylalanine (l g). m.p. 180-182°C
Mass (C.I.) (M+H)+= 427
^-N R (200 MHz, DMSO-d6): δ (ppm): 1.80-1.88 (m, IH, SH); 2.22-2.84 (m, 5H, CH2-
CH-CH2); 2.86-3.18 (m, 2H, CH2-CH-COO); 4.46-4.57 (m, IH, CH-COO); 7.10-7.25 (m, 5H, phenyl); 7.32-7.84 (m, 4H, phenylene); 7.74 (d, IH, N-CH=CH-S); 7.89 (d, IH, N-
CH=CH-S); 8.35 (d, IH, NH); 12.76 (s, IH, COOH);
By working m an analogous way the following compounds were prepared:
N-r(2R)-3-mercapto-2-phenylmethylpropionyll-4-(2-tiazolyl>L-phenylalanme (compound 17)
JH-NMR (200 MHz, DMSO-dg): δ (ppm): 2.15-2.23 (m, IH, SH); 2.31-2.74 (m, 5H, CH2- CH-CH2); 2.78-3.07 ( , 2H, CH2-CH-COO); 4.42-4.53 (m, IH, CH-COO); 7.02-7.80 (m,
9H, phenyl and phenylene); 7.75 (d, IH, N-CH=CH-S); 7.90 (d, IH, N-CH=CH-S); 8.36 (d,
1H, NH);
N-r(2SV3-mercapto-2-phenylmemylpropionyl -4-(2-furyl)-L-phenylalanme (compound 18) m.p. 153-155°C Mass (CI.) (M+H)+= 410
1 H-NMR (200 MHz, CDC1 ): δ (ppm): 1.40 (t, IH, SH); 2.45-3.25 (m, 7H, CH2-CH-CH2,
CH2-CH-COO); 4.80-4.90 (m, IH, CH-COO); 5.86 (d, IH, NH); 6.42-7.42 (m, 3H, furyl);
7.07-7.57 (m, 9H, phenyl, phenylene);
N-f(2S)-3-mercapto-2-phenylmethylpropionyl1-4-(5-pyrirrudinyl)-L-phenylalanine (compound 19) m.p. 193-195°C Mass (C I ) (M+H)+= 422
^-NMR (200 MHz, DMSO-dg) δ (ppm) 1 81 (bm, IH, SH), 2 21-3 20 (m, 7H, CH2-CH- CH2, CH2-C6H4-pyπmιdmyl), 4 46-4 57 (m, IH, CH-COO), 7 06-7 29 (m, 5H, phenyl), 7 36-7 72 (m, 4H, phenylene), 8 33 (d, IH, NHCO), 9 08 (s, 2H, N-CH-C-CH-N), 9 15 (s, IH, N-CH-N),
N-r(2SV3-mer∞pto-2-phenylmethylpropιonyll-4-(2-pyrazmyl)-L-phenylalanme (compound 20) m p 176-178°C Mass (C I ) (M+H)+= 422
^-NMR (200 MHz, DMSO-d6) δ (ppm) 1 84 (bt, IH, SH), 2 21-3 21 (m, 7H, CH2-CH- CH2, CH2-CH-COO), 4 48-4 59 (m, IH, CH-COO), 7 10-7 26 (m, 5H, phenyl), 7 37-8 05 (m, 4H, phenylene), 8 37 (d, IH, NHCO), 8 58 (d, IH, CH-N-CH-CH-N), 8 68-8 70 (m, IH, CH-N-CH-CH-N), 9 21 (d, IH, C-CH-N), 12 78 (b, IH, COOH), N-f(2S)-3-mercapto-2-phenylmethylpropιonyll-4-(3-pyndyl)-L-phenylalanme (compound 21) m p 146-148°C Mass (C I ) (M+H)+= 421
JH-NMR (200 MHz, DMSO-d6) δ (ppm) 1 69-1 89 (b, IH, SH), 2 22-3 18 (m, 7H, CH2- CH-CH2, CH2-phenylene), 4 45-4 56 (m, IH, CH-COO), 7 09-7 26 (m, 5H, phenyl), 7 42- 7 48 (m, IH, CH-N-CH-CH-CH), 7 62-7 33 (m, 4H, phenylene), 7 98-8 04 (m, IH, CH-N- CH-CH-CH), 8 30 (d, IH, CONH), 8 53 (dd, IH, CH-N-CH-CH-CH), 8 83 (d, IH, CH-N- CH-CH-CH),
N-r(2SV3-mercapto-2-phenylmemylpropιonvπ-4-(2-pyndyl'r-L-phenylalamne (compound 22) m p 157-159°C Mass (C I ) (M+H)+= 421
'H-NMR (200 MHz, DMSO-d6) δ (ppm) 1 87 (b, IH, SH), 2 23-3 19 (m, 7H, S-CH2-CH- CH2, CONH-CH-CH2), 4 44-4 45 (m, IH, CH-COO), 7 09-7 93 (m, 8H, N-CH-CH-CH- CH, phenyl), 7 30-7 99 (m, 4H, phenylene), 8 29 (d, IH, CONH), 8 61-8 65 (m, IH, C-N- CH), N-f(2S)-3-mercapto-2-phenylmethylpropιonvn-4-(2-thιenyl)-L-phenylalanme (compound 23) m p 152-154°C !H-NMR (200 MHz, CDC13) δ (ppm) 1 34-1 43 (m, IH, SH), 2 44-3 26 (m, 7H, S-CH2-
CH-CH2, CONH-CH-CH2), 4 80-4 89 (m, IH, NH-CH-COO), 5 81 (d, IH, NH), 7 02-7 52
(m, 12H, aryl), N-r(2S)-3-mercapto-2-phenylmethylpropιonyll-4-(3-thιenyl)-L-phenylalanme (compound 24) m p 169-171°C
Mass (C I ) (M+H)+= 426
TLC (ethyl acetate hexane acetic aαd=50 50 5), Rf = 0 44
!H-NMR (200 MHz, CDC13) δ (ppm) 1 84 (bs, IH, SH), 2 23-3 13 (m, 7H, S-CH2-CH- CH2, NH-CH-CH2), 4 42-4 53 (m, IH, NH-CH-COO), 7 12-7 80 (m, 12H, aryl), 8 30 (d,
IH, JHH=8 2 Hz, NH),
N-[(2SV3-mercapto-2-phenylmethylpropιonyl]-4-(3-furyl)-L-phenylalan ne (compound 25) m p 140-142°C
Mass (C I ) (M+H)+= 410 TLC (ethyl acetate hexane acetic aαd=50 50 5), Rf = 0 42
!H-NMR (200 MHz, CDC13) δ (ppm) 1 79-1 90 (m, IH, SH), 2 22-3 11 (m, 7H, S-CH2-
CH-CH2, NH-CH-CH2), 4 41-4 53 (m, IH, NH-CH-COO), 7 11-7 50 (m, 9H, phenyl, phenylene), 6 91-8 12 (m, 3H, furyl), 8 30 (d, IH, JHH=8 2 Hz, NH),
N-f3-mercapto-2-(3-pyπdylmethyl)propιonyll-4-(2-thιazolyl)-L-phenylalanme - stereoisomer A (compound 26) m p 193-196°C
Mass (C I ) (M+H)+= 428
TLC (methylene chlonde methanol acetic aαd=85 15 1 5), Rf = 0 53 H-NMR (200 MHz, DMSO-d6) δ (ppm) 2 37-3 05 (m, 7H, S-CH2-CH-CH2, NH-CH- CH2), 4 36-4 47 (m, IH, NH-CH-COO), 7 76-7 90 (m, 2H, thiazolyl), 7 10-8 33 (m, 9H,
NH, pyndyl, phenylene),
N-r3-mercapto-2-(3-pyπdylmethyl)propιonyll-4-(2-thιazolyl)-L-phenylalanme - stereoisomer
B (compound 27)
Mass (C I ) (M+H)+= 428 TLC (methylene chlonde methanol acetic aαd=85 15 1 5), Rf = 0 47
^-NMR (200 MHz, DMSO-d6) δ (ppm) 2 28-3 20 (m, 7H, S-CH2-CH-CH2, NH-CH- CH ), 4 20-4 35 (m, IH, NH-CH-COO), 7 70-7 90 (m, 2H, thiazolyl), 7 17-8 40 (m, 9H,
NH, pyndyl, phenylene),
N-(2-ιsobutyl-3-mercapto-propιonyl)-4-(2-thιazolyl)-L-phenylalanme (compound 28) ^-NMR (200 MHz, DMSO-d6) δ (ppm) 0 50-1 44 (m, 9H), 1 68-2 25 (m, 4H), 2 79-3 22
(m, 2H), 4 50-4 63 (m, IH), 7 34-7 85 (m, 4H), 7 73-7 90 (m, 2H), 8 27-8 39 (2d, IH),
Mass (C I ) (M+H)+= 393
N-r3-mer^pto-2-(3-memoxyphenyl)memyl-propιonylV4-(2-thιazolyl)-L-phenylalanme
(compound 29) Mass (C I ) (M+H)+=457
!H-NMR (200 MHz, DMSO-d6+D20) δ (ppm) 2 20-3 21 (m, 7H), 3 70 (d, 3H), 4 48 (m,
IH), 6 55-6 86 (m, 3H), 7 00-7 40 (m, 3H), 7 65-7 95 (m, 4H), 8 27-8 45 (bt, CONH),
N-f3-mercaptc-2-(2-fluorophenyl)methyl-propιonyll-4-(2-dιιazolyl)-L-phenylalanme
(compound 30) Mass (C I ) (M+H)+= 45
*H-NMR (200 MHz, DMSO-d6) δ (ppm) 2 20-3 18 (m, 8H), 4 35-4 55 (m, IH), 6 85-7 35
(m, 6H), 7 65-7 90 (m, 4H), 8 35 (d, IH),
N-r3-mercapto-2-(2-tmenyl)methyl-propιonyll-4-(2-thιazolyl)-L-phenylalarune (compound 31)
^-NMR (200 MHz, DMSO-d6) δ (ppm) 1 82-3 19 (m, 8H, CH2-CH2-CH2, NH-CH- CH2), 4 44-4 59 (m, IH, CH-COO), 6 62-7 91 ( , 9H, aryl), 8 42 (d, IH, NH),
N-f3-mercapto-2-(2-fυryl)methyl-propιonyll-4-(2-thιazolyl)-L-phenylalanme (compound 32) H-NMR (200 MHz, DMSO-dg) δ (ppm) 1 79-3 18 (m, 8H, S-CH2-CH2-CH , NH-CH-
CH2), 4 43-4 58 (m, 4H, CH-NH), 5 79-7 47 (m, 3H, furyl), 7 30-7 85 (m, 4H, phenylene),
7 73-7 91 (m, 2H, thiazolyl), 8 40-8 46 (2d, IH, NH), N-r3-mercapto-2-(3-memyl-5-ιsoxazolyl)memyl-prc>pιonyll-4-(2-thιazolyl)-L-phenylalanme
(compound 33)
!H-NMR (200 MHz, DMSO-dό) δ (ppm) 2 00-2 12 (2S, 3H, CH3-ιsoxazolyl), 2 28-3 19
(m, 8H, S-CH2-CH2-CH2, CH2-ρhenylene), 4 43-4 59 (m, IH, CH-NH), 5 75-6 03 (2s, IH, isoxazolyl), 7 29-7 86 (m, 4H, phenylene), 7 74-7 90 (m, 2H, thiazolyl), 8 46-8 53 (2d, IH, NH),
Example 8 In vitro evaluation ofthe pharmacoloac activity a) NEP-inhibitorv activity
The NEP-inhibitory activity was evaluated m rat kidney cortex membranes prepared ac- cording to the procedure descnbed by T Maeda et al in Biochim Biophys Acta 1983,
731(1), 115-120
Kidneys were removed from male Sprague-Dawley rats weighing approximately 300 g and kept at 4CC
Kidney cortex was carefully dissected, finely minced and suspended m a homogenization buffer (10 mM sodium phosphate pH 7 4 containmg 1 mM MgCl2, 30 mM NaCl, 002%
NaN3) 1 15 weight/volume
The tissue was then cold homogenized for 30 seconds usmg an Ultra-Turrax homogenizer
Approximately 10 ml of homogenate were layered over 10 ml of sucrose (41% weight volume) and centπfuged at 31200 φm for 30 mmutes at 4°C in a fixed angle rotor The membranes were collected from die buffer/sucrose interface, washed twice with 50 mM TRIS/HC1 buffer (pH 7 4) and resuspended into the same buffer for storage in small aliquots at -80°C until use
The NEP-inhibitory activity was evaluated by usmg the method descπbed by C Llorens et al , Eur J Pharmacol , 69, (1981), 113-116, as hennafter reported Aliquots of die membrane suspension prepared as above descnbed (concentration 5 μg/ml of proteins) were preincubated m the presence of an aminopeptidase inhibitor (Bestatin - 1 mM) for 10 minutes at 30°C
[3H][Leu5]-enkephalme (15 nM) and buffer TRIS/HCl pH 7 4 (50 mM) were added in order to obtain a final volume of 100 μl Incubation (20 minutes at 30°C) was stopped by adding HCl 0 IM (100 μl)
The formation of the metabolite [- H]Tyr-Gly-Gly was quantified by chromatography on polystyrene columns (Porapak Q)
The percentage of inhibition of the metabolite formation m the membrane preparations treated with the compounds of formula I and wi the comparative compounds with re- spect to the untreated membrane preparations was expressed as IC50 (nM) value b) ACE-inhibitorv activity The ACE-inhibitory activity was evaluated according to die method reported in die litera¬ ture by B Holmquist et al., m Analytical Biochemistry 95, 540-548 (1979) 50 μM of ACE (250 mU/ml punfied by lung rabbit, EC 3 4 15 1 SIGMA) were premcu- bated with 50 μl of the compound of formula I or of die reference compound or related vehicle in mermostated cuvettes at 37°C
The reaction was started by add g furylacryloylphenylalanylglycylglycine 0 8 mM (FAPGG-SIGMA) Contemporaneously, by usmg a Beckman DU-50 spectrophotometer provided with a pro- gram for calculating delta A/minutes and regression coeffiαents of the enzyme kinetics curves, the absorbance at 340 nm was recorded in continuo for 5 minutes The percentage ofthe enzyme inhibition in the preparations treated with the compounds of formula I or with the comparative compounds with respect to the untreated preparations was expressed as IC,- (nM) value The IC50 (nM) values related to die ACE-inhibitory activity and NEP-inhibitory activity of the compounds 16, 18-25, 27-33 and of the compaπson compounds Thioφhan and Captopπl are reported m the following table 1
Table 1
ACE-inhibitory and NEP-inhibitory activity, expressed as IC,_ (nM) value, of compounds
16, 18-25, 27-33 and of Thioφhan and Captopril.
Compound ACE-inhibitory activity IC50 (nM) NEP-inhibitory activity IC50 (nM) I
16 3.2 1.8
18 1.8 1.8
19 1.9 1.8
20 1.5 2.5
21 1.7 2.6
22 1.8 2.0
23 1.6 0.6
24 2.5 1.3
25 2.4 1
27 9.1 17.3
28 5.8 1.8
29 4.6 9.0
30 10.7 11.2
31 8.6 1.2
32 8.6 4.0
33 7.9 4.5
Thioφhan 99 18
Captopril 4.6 not active The data reported in table 1 show that d e compounds of formula I, object of the present in¬ vention, are endowed wrth a significant mixed ACE/NEP inhibitory activity Said activity is comparable to that of Captopπl, to what it concerns the ACE inhibitory activ- ity, and greater dian that of Thioφhan, to what it concerns the NEP inhibitory activity
Example 9 "Ex vivo" evaluation ofthe pharmacoloac activity a) NEP-mhibitorv activity
The ex vivo NEP-inhibitory activity was evaluated accordmg to the procedure reported m the literature by M Orlowsky et al , m Biochemistry 1981, 20, 4942-4950
The inhibitory activity of the compounds of formula I was evaluated kidneys of sponta¬ neously hypertensive rats (SHR), 5 mmutes after 1 v injection (0 6 and 21 μmoles/kg) and 30 mmutes, 60 mmutes and 4 hours after oral administration (30 μmoles/kg) of the tested compounds After die removal of the kidneys from SHR, the renal tissue was homogenized and incu¬ bated for 15 mmutes at 37°C m the presence of Glutaryl-Ala-Ala-Phe-2-naphthylamιde (GAAP), as a substrate, and arrunopeptidase M at pH 7 6
The reaction was stopped by addmg an aqueous solution at 10% of tnchloroacetic aαd The released 2-naphthylamme was determmed by addmg fast garnet dye (2 ml) Enzyme reaction rates were determmed by measuring the mcrease m the optical density at
524 nm (OD524) with respect to a standard obtamed with 2-naphthylamιne complexed with fast garnet
The NEP-inhibitory activity of the tested compounds was expressed as percentage of NEP-inhibition m SHR kidneys b) ACE-inhibitorv activity
The ex vivo ACE-inhibitory activity was evaluated by usmg a radiometπc assay method, as reported m the literature by J W Ryan et al in Biochem J (1977), 167, 501-504 The inhibitory activity ofthe compounds of formula I was evaluated lungs of spontane¬ ously hypertensive rats (SHR), 5 mmutes after l v injection (0 6 and 21 μmoles/kg) and 30 mmutes, 60 minutes and 4 hours after oral administration (30 μmoles/kg) of die tested compounds After the removal ofthe lungs from SHR, the lung tissue was homogenized and incubated for 2 hours at 37°C m the presence of [ H]Hyp-Gly-Gly, as a substrate
The reaction was stopped by addmg hydrochlonc aαd
The released radiolabeUed hyppuπc aαd was extracted with ethyl acetate and counted by liquid scintillation spectrometry, accordmg to conventional methods
The ACE-inhibttory activity of the tested compounds was expressed as percentage of
ACE-inhibition m SHR lungs
As an example, the percentages of basal enzymatic activity obtamed m die ex vivo tests after intravenous or oral administration of compound 16 and of compaπson compounds
R-l, R-2, R-3 and R-4 are reported in die following table 2
Table 2
Percentages of ex vivo ACE-inhibition and NEP-mhibition of compound 16 and of compounds
R-l, R-2, R-3 and R-4
Compound Treatment ACE-inhibition (lungs) NEP-inhibrtion (kidneys)
5 mmutes 60 minutes 5 minutes 60 minutes
16 i v ( 0 6 μmoles/kg) 15% 34%
16 i v (21 μmoles/kg) 72% 49%
16 oral (30 μmoles/kg) 36% 39%
30 mmutes 4 hours 30 mmutes 4 hours ]
16 oral (30 μmoles/kg) 60% 45% 55% 40%
R-l oral (30 μmoles/kg) 25% 20% 5% not active
R-2 oral (30 μmoles/kg) 30% 25% 30% not active
R-3 oral (30 μmoles/kg) 25% 20% 10% 5%
R-4 oral (30 μmoles/kg) 25% 10% not active not active
The data reported m table 2 confirm at the compounds of formula I, object of die present in¬ vention, are endowed with a significant ACE/NEP-inhibitory activity after intravenous ad¬ ministration as well as after oral administration
The ex-vivo ACE NEP-inhibitory activity ofthe compounds of formula L, moreover, results to be significantly higher than that ofthe compaπson compounds

Claims

Claims
1) A compound of formula
Ri
R-CH2-CH-CONH-CH-COOR2 (I)
I
CH2-R3 wherem R is a mercapto group or a R4COS group convertible in the organism to mercapto group, Rj IS a straight or branched C -C4 alkyl group or an aryl or arylalkyl group havmg from 1 to 6 carbon atoms the alkyl moiety wherein the aryl is a phenyl or a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consising of nitrogen, oxygen and sulphur, optionally substituted with one or more substituents, the same or different, selected from the group consisting of halogen atoms, hydroxy groups, alkoxy, alkyl, alkylthio, alkylsulphonyl or alkoxycarbonyl groups havmg from 1 to 6 car¬ bon atoms m the alkyl moiety, Cι-C3 alkyl groups containing one or more fluorine atoms, carboxy groups, nitro groups, ammo or aminocarbonyl groups, acylammo groups, arnino- sulphonyl groups, mono- or di-alkylanuno or mono- or di-alkylaminocarbonyl groups havmg from 1 to 6 carbon atoms m the alkyl moiety,
R2 is a hydrogen atom, a straight or branched C J-C4 alkyl or a benzyl group, R3 is a phenyl group substituted by a 5 or 6 membered aromatic heterocycle with one or two heteroatoms selected from the group consisting of nitrogen, oxygen and sulphur, bemg die phenyl and the heterocyclic groups optionally substituted with one or more substituents, me same or different, as indicated for R1 ,
R4 is a straight or branched C1-C4 alkyl group or a phenyl group, the carbon atoms marked witii an astensk are stereogenic centers, and pharmaceutically acceptable salts thereof
2) A compound of formula I according to claim 1 wherein R3 is a phenyl group substituted in position 4 with a heterocyclic group
3) A compound of formula I accordmg to claim 2 wherem R\ represents an arylalkyl group optionally substituted with one or more substituents, the same or different, selected from the groups consisting of halogen atoms, hydroxy, alkyl or alkoxy groups 4) A compound of formula I accordmg to claim 3 wherem Rj represents a phenylaikyl group optionally substituted with one or more substituents, me same or different, selected among halogen atoms, hydroxy, alkyl or alkoxy groups
5) A compound of formula I accordmg to claim 1 m the form of a salt with an alkali metal se¬ lected from the group consisting of sodium, lithium and potassium
6) A process for the preparation of a compound of formula I accordmg to claim 1 compπsmg the reaction between a compound of formula
Ri
I
R-CH2-CH-COOH (II)
wherem
R and Rj have the above reported meanings, and a phenylalanine den vati ve of formula
*
H2N-CH-COOR2 (TH)
I
CH2-R3 wherem
R2 and R have die above reported meanings
7) A pharmaceutical composition containing a therapeutically effective amount of a compound of formula I accordmg to claim 1 in admixture with a earner for pharmaceutical use
8) A pharmaceutical composition accordmg to claim 6 for me treatment of cardiovascular dis- eases
9) A method for die treatment of cardiovascular diseases compnsmg the administration of a merapeutically effective amount of a compound accordmg to claim 1
10) A compound selected from the group consisting of N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyl)-4-(2-thιazolyl)-L-phenylalanme methyl ester,
N-(3-phenylrarbonylthιo-2-phenylmethylpropιonyl)-4-(2-pyπdyl)-L-phenylaianme methyl ester,
N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyI)-4-(3-pyndyl)-L-phenylalanme benzyl ester, N-(3-phenylcarbonylt o-2-phenylmemylpropιonyl)-4-(2-furyl)-L-phenylalanme methyl ester, N-(3-phenylcarbonylmιo-2-phenylmemylpropιonyl)-4-(5-pynrmdmyl)-L-phenyialanme ethyl ester, N-(3-phenylcarbonylthιo-2-phenylmemylpropιonyl)-4-(2-pyrazmyl)-L-phenylalanme methyl ester,
N-(3-phenylcarbc«yltruo-2-phenylmemylpropionyl)-4-(2-t enyl)-L-phenylalanme methyl ester, N-(3-phenylcarbonylthιo-2-phenylmethylpropιonyl)-4-(3-thιenyl)-L-phenylalanme methyl ester,
N-(3-phenyicarbonylmιo-2-phenylmcrthylpropιonyl)-4-(3-furyl)-L-phenylalanme methyl ester, N-[3-phenylcarbonyltluo-2-(3-pyπdylmemyl)propιonyl]-4-(2-thιazolyl)-L-phenylalanme methyl ester, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-tiιιazoiyl)-L-phenylalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-pyndyl)-L-phenylalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(3-pyndyl)-L-phenylalanme, N-(3-mercapto-2-phenylmemylpropιonyI)-4-(2-furyl)-L-phenylalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(5-pyπmι nyl L-phenylalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-pyrazmyl)-L-phenvlalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(2-dιιenyl)-L-phenylalanme, N-(3-mer(*apto-2-phenylmethylpropιonyl)-4-(3-tiιιenyl)-L-phenylalanme, N-(3-mercapto-2-phenylmethylpropιonyl)-4-(3-furyl)-L-phenylalanme, N-[3-mercaptj 2-(3-pyndylmethyl)propιonyl]-4-(2-thιazolyl)-L-phenylalanme, N-[3-acetyltlυo-2-(3-methoxyphenyl)methyl-propιonyl]-4-(2-thιazolyl)-L-phenylalanme methyl ester,
N-[3-acetylmιo-2-(2-fluorophenyl)methyl-propιonyl]-4-(2-thιazolyl)-L-phenylalanme methyl ester,
N-[3-phenylcarbonylthιcr-2-(2-thιenyl)memyl-propιonyl]-4-(2-tbazolyl)-L-phenylalan e methyl ester, N-[2-(2-furyl)memyl-3-phenylcarbonylthiCr-propionyl]-4-(2-thiazolyl)-L-phenylaIanme methyl ester, N-[2-(3-memyl-5-isoxazolyl)memyl-3-phenylcarbonylthio-propionyl]-4-(2-thiazolyl)-L- phenylalanine methyl ester;
N-[3-mercapto-2-(3-memoxyphenyl)me yl-propionyl]-4-(2-thiazolyl)-L-phenylalanine; N-[3-mercapto-2-(2-tmenyi)memyl-propionyl]-4-(2-tlιiazolyl)-I^phenylalanine; N-[3-mercapto-2-(3-meώyI-5-isoxazolyl)methyl-propionyl]-4-(2-thiazolyl)-L-phenylalanine; N-[2-(2-fluorophenyl)medιyl-3-mercapto-propionyl]-4-(2-thiazolyl)-L-phenylalanine; N-[2-(2-furyl)memyl-3-mercapto-propionyl]-4-(2-thiazolyl)-L-phenylalanine; and die stereoisomers (2S) or (2R) thereof.
PCT/EP1996/005663 1995-12-28 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity WO1997024342A1 (en)

Priority Applications (21)

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KR1019980704934A KR19990076809A (en) 1995-12-28 1996-12-17 Thiol derivatives showing metallopeptidase inhibitory activity
EE9800192A EE03766B1 (en) 1995-12-28 1996-12-17 Thiol derivatives having metallopeptidase inhibitory activity
RO98-01109A RO119882B1 (en) 1995-12-28 1996-12-17 Phenylalanine derivatives with metallopeptidase inhibitory activity
SK889-98A SK282977B6 (en) 1995-12-28 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity
HU9903636A HUP9903636A3 (en) 1996-12-17 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity, process for their preparation, pharmaceutical compositions containing such thiol derivatives and process for their use
JP09524001A JP2000502687A (en) 1995-12-28 1996-12-17 Thiol derivatives having metallopeptidase inhibitory activity
UA98074090A UA62923C2 (en) 1995-12-28 1996-12-17 Thiol derivatives having an inhibiting activity relative to metal-peptidases, a pharmaceutical composition and a method for the treatment of cardiovascular diseases
DE69634543T DE69634543D1 (en) 1995-12-28 1996-12-17 THIOL-DERIVATIVES WITH METAL-OPEPTIDASE-HEMDERING EFFECT
PL96327580A PL188151B1 (en) 1995-12-28 1996-12-17 Derivatives of thioles exhibiting metallopepsidase inhibiting activity
AU13018/97A AU713156B2 (en) 1995-12-28 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity
IL12491596A IL124915A (en) 1995-12-28 1996-12-17 Thiol derivatives, process for their preparation and pharmaceutical compositions containing them
EP96944583A EP0877740B1 (en) 1995-12-28 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity
APAP/P/1998/001264A AP1186A (en) 1995-12-28 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity.
BR9612373A BR9612373A (en) 1995-12-28 1996-12-17 Thiol derivatives with metallopeptidase inhibitory activity
AT96944583T ATE292123T1 (en) 1995-12-28 1996-12-17 THIOL DERIVATIVES WITH METALLOPEPTIDASE INHIBITING EFFECT
EA199800544A EA000991B1 (en) 1995-12-28 1996-12-17 Thiol derivativeswith metallopeptidase inhibitory activity
NZ325785A NZ325785A (en) 1995-12-28 1996-12-17 Thiol-phenylalanine derivatives and medicaments having metallopeptidase inhibitory activity
BG102553A BG63942B1 (en) 1995-12-28 1998-06-18 Thiol derivatives having metallopeptidase inhibitor activity
IS4780A IS1885B (en) 1995-12-28 1998-06-22 Thiol derivatives with metal peptidase depressant activity, their method of preparation, pharmaceutical preparations containing such compound, together with the use of the compounds for the manufacture of pharmaceuticals
NO19982977A NO310914B1 (en) 1995-12-28 1998-06-26 Thiol derivatives having metallopeptidase inhibitory activity, process for their preparation, pharmaceutical compositions containing such a compound, and use of the compound for the preparation of medicaments
HK99103110A HK1018008A1 (en) 1995-12-28 1999-07-20 Thiol derivatives with metallopeptidase inhibitory activity

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IT95MI002773A IT1277737B1 (en) 1995-12-28 1995-12-28 TIOLIC DERIVATIVES FOR METALLOPEPTIDASE INHIBITIVE ACTIVITY
ITMI95A002773 1995-12-28

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WO2003068725A2 (en) * 2002-02-15 2003-08-21 Glaxo Group Limited Process for the preparation of 4-hetero-substituted phenylalanine derivatives
WO2003104200A1 (en) * 2002-06-07 2003-12-18 Glaxo Group Limited N-mercaptoacyl phenyalanine derivatives, process for their preparation, and pharmaceutical compositions containing them
WO2003104189A2 (en) * 2002-06-07 2003-12-18 Glaxo Group Limited Compounds
US6855843B2 (en) 1998-01-20 2005-02-15 Tanabe Seiyaku Co., Ltd. Inhibitors of α4 mediated cell adhesion
WO2005054177A1 (en) 2003-12-01 2005-06-16 Novartis Ag Delta-amino-gamma-hydroxy-omega-aryl-alkanoic acid amides and use as renin inhibitors
US7045653B2 (en) 2002-12-23 2006-05-16 Pfizer, Inc. Pharmaceuticals
US7271280B2 (en) 2002-03-05 2007-09-18 Sumitomo Chemical Company, Limited Process for preparing a biaryl compound
EP1915993A1 (en) 2000-11-17 2008-04-30 Novartis AG Synergistic combinations comprising a renin inhibitor for cardiovascular diseases
EP1977741A2 (en) 2004-03-17 2008-10-08 Novartis AG Use of renin inhibitors in therapy
US7476758B2 (en) 2002-02-28 2009-01-13 Mitsubishi Tanbe Pharma Corporation Process for preparing a phenylalanine derivative and intermediates thereof
WO2011116115A1 (en) 2010-03-16 2011-09-22 Novartis Ag Aliskiren composition comprising a medium chain fatty acid, their process of manufacturing

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US9199914B2 (en) 2010-02-03 2015-12-01 Meh Associates, Inc. Multiple substituted fluoromethanes as selective and bioactive isosteres

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Cited By (17)

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Publication number Priority date Publication date Assignee Title
US6448408B1 (en) 1996-12-24 2002-09-10 Zambon Group S.P.A. Process for the preparation of heteroaryl-phenylalanines
WO1998028284A1 (en) * 1996-12-24 1998-07-02 Zambon Group S.P.A. Process for the preparation of heteroaryl-phenylalanines
US6855843B2 (en) 1998-01-20 2005-02-15 Tanabe Seiyaku Co., Ltd. Inhibitors of α4 mediated cell adhesion
EP1930000A1 (en) 2000-11-17 2008-06-11 Novartis AG Combination of organic compounds
EP1915993A1 (en) 2000-11-17 2008-04-30 Novartis AG Synergistic combinations comprising a renin inhibitor for cardiovascular diseases
WO2003068725A2 (en) * 2002-02-15 2003-08-21 Glaxo Group Limited Process for the preparation of 4-hetero-substituted phenylalanine derivatives
WO2003068725A3 (en) * 2002-02-15 2003-12-24 Glaxo Group Ltd Process for the preparation of 4-hetero-substituted phenylalanine derivatives
US7476758B2 (en) 2002-02-28 2009-01-13 Mitsubishi Tanbe Pharma Corporation Process for preparing a phenylalanine derivative and intermediates thereof
US7271280B2 (en) 2002-03-05 2007-09-18 Sumitomo Chemical Company, Limited Process for preparing a biaryl compound
US7714157B2 (en) 2002-03-05 2010-05-11 Sumitomo Chemical Company, Limited Process for preparing a biaryl compound
WO2003104189A2 (en) * 2002-06-07 2003-12-18 Glaxo Group Limited Compounds
WO2003104189A3 (en) * 2002-06-07 2005-09-01 Glaxo Group Ltd 4-pyrazol-phenylalanine derivatives as ace-nep inhibitors
WO2003104200A1 (en) * 2002-06-07 2003-12-18 Glaxo Group Limited N-mercaptoacyl phenyalanine derivatives, process for their preparation, and pharmaceutical compositions containing them
US7045653B2 (en) 2002-12-23 2006-05-16 Pfizer, Inc. Pharmaceuticals
WO2005054177A1 (en) 2003-12-01 2005-06-16 Novartis Ag Delta-amino-gamma-hydroxy-omega-aryl-alkanoic acid amides and use as renin inhibitors
EP1977741A2 (en) 2004-03-17 2008-10-08 Novartis AG Use of renin inhibitors in therapy
WO2011116115A1 (en) 2010-03-16 2011-09-22 Novartis Ag Aliskiren composition comprising a medium chain fatty acid, their process of manufacturing

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IS4780A (en) 1998-06-22

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