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WO1995035311A1 - Derives de l'acide 3-amino-2-oxo-1-piperidineacetique - Google Patents

Derives de l'acide 3-amino-2-oxo-1-piperidineacetique Download PDF

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Publication number
WO1995035311A1
WO1995035311A1 PCT/US1995/007661 US9507661W WO9535311A1 WO 1995035311 A1 WO1995035311 A1 WO 1995035311A1 US 9507661 W US9507661 W US 9507661W WO 9535311 A1 WO9535311 A1 WO 9535311A1
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Prior art keywords
argininal
substituted
carbon atoms
compound according
gly
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PCT/US1995/007661
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English (en)
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WO1995035311A9 (fr
Inventor
Joseph Edward Semple
Robert J. Ardecky
Ruth F. Nutt
William Charles Ripka
David C. Rowley
Marguerita S. L. Lim-Wilby
Terrence K. Brunck
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Corvas International, Inc.
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Priority to BR9508048A priority Critical patent/BR9508048A/pt
Priority to MX9606566A priority patent/MX9606566A/es
Priority to JP8502499A priority patent/JPH10503176A/ja
Priority to EP95923922A priority patent/EP0802923A1/fr
Priority to AU28630/95A priority patent/AU700808B2/en
Priority to NZ288940A priority patent/NZ288940A/xx
Publication of WO1995035311A1 publication Critical patent/WO1995035311A1/fr
Publication of WO1995035311A9 publication Critical patent/WO1995035311A9/fr
Priority to NO965353A priority patent/NO965353L/no

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06139—Dipeptides with the first amino acid being heterocyclic
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/05—Dipeptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
    • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
    • C07K5/06—Dipeptides
    • C07K5/06191—Dipeptides containing heteroatoms different from O, S, or N

Definitions

  • the present invention relates compounds which are potent and specific inhibitors of thrombin.
  • the present invention relates to novel peptide aldehydes, their pharmaceutically acceptable salts, and pharmaceutically acceptable compositions thereof which are useful as potent and specific inhibitors of blood coagulation in vitro and in vivo in mammals.
  • the invention relates to methods of using these inhibitors as therapeutic agents for disease states in mammals characterized by abnormal thrombosis.
  • Normal hemostasis is the result of a complex balance between the processes of clot formation (blood coagulation) and clot dissolution (fibrinolysis) .
  • Blood coagulation is the culmination of a series of amplified reactions in which several specific zymogens of serine proteases in plasma are activated by limited proteolysis. Nemerson, Y. and Nossel, H.L., Ann. Rev. Med., 33 : 479 (1982) . This series of reactions results in the formation of an insoluble fibrin matrix which is required for the stabilization of the primary hemostatic plug. The interaction and propagation of the activation reactions occurs through the extrinsic and intrinsic pathways of coagulation.
  • Factor Xa catalyzes the penultimate step in the blood coagulation cascade which is the formation of the serine protease thrombin. This step occurs following the assembly of the prothrombinase complex which is composed of factor Xa, the non-enzymatic co-factor Va and the substrate prothrombin assembled on the surface of adhered, activated platelets or .systemically circulating membranous microparticles.
  • Proteolytic activation of zymogen factor X to its catalytically active form, factor Xa can occur by either the intrinsic or extrinsic coagulation pathways.
  • the intrinsic pathway is referred to as "intrinsic” because everything needed for clotting is in the blood. Saito, H. , “Normal Hemostatic Mechanisms", Disorders of Hemostasis, pp. 27-29, Grune & Stratton, Inc. (0. D. Ratnoff, M.D. and C. D. Forbes, M.D. edit. 1984) .
  • This pathway is comprised of the zymogen serine proteases, factors IX and XI, and the non-enzymatic co-factor, factor VIII.
  • Factor XIa catalyzes the activation of factor IX to factor IXa which in combination with the activated form of factor VIII on an appropriate phospholipid surface, results in the formation of the tenase complex. This complex also catalyzes the formation of the serine protease, factor Xa, from its zymogen, factor X which subsequently results in clot formation.
  • extrinsic pathway is referred to as "extrinsic" because the tissue factor which binds to and facilitates the activation of factor VII comes from outside the blood. s
  • the major components of this pathway are the zymogen serine protease, factor VII, and the membrane bound protein, tissue factor. The latter serves as the requisite non-enzymatic co-factor for this enzyme.
  • the initiation of this pathway is thought to be an autocatalytic event resulting from the activation of zymogen factor VII by trace levels of activated factor VII (factor Vila) , both of which are bound to newly exposed tissue factor on membrane surfaces at sites of vascular damage.
  • factor Vila/tissue factor complex directly catalyzes the formation of the serine protease, factor Xa, from its zymogen, factor X. Exposure of blood to injured tissue initiates blood clotting by the extrinsic pathway.
  • thrombin is catalyzed by factor Xa following the assembly of the catalytic prothrombinase complex as reviewed by Mann, K. G. et. al., "Surface- Dependent Reactions of the Vitamin K-Dependent Enzyme Complexes", Blood, 76.: 1-16 (1990).
  • This complex is composed of factor Xa, the non-enzymatic co-factor Va and the substrate prothrombin all assembled on an appropriate phospholipid surface.
  • the requirement of a macromolecular complex for efficient catalysis results in the protection of factor Xa from natural anticoagulant mechanisms such as heparin-antithrombin III mediated inhibition.
  • Thrombin is the primary mediator of thrombus formation. Thrombin acts directly to cause formation of insoluble fibrin from circulating fibrinogen. In addition, thrombin activates the zymogen factor XIII to the active transglutaminase factor Xllla which acts to covalently stabilize the growing thrombus by crosslinking the fibrin strands. Lorand, L. and Konishi, K. , Arch. Biochem. y
  • thrombin is the most potent agonist of platelet activation, and it has been demonstrated to be the primary pathophysiologic-mediator of platelet-dependent arterial thrombus formation. Edit, J.F. et , al., J. Clin. Invest., 84* 18 (1989) .
  • Thrombin-mediated platelet activation leads to ligand-induced inter-platelet aggregation principally due to the bivalent interactions between adhesive ligands such as fibrinogen and fibronectin with platelet integrin receptors such as glycoprotein
  • Thrombin-activated platelets can also support further thrombin production through the assembly of new prothrombinase and tenase (factor IXa, factor Villa and factor X) catalytic complexes on the membrane surface of intact activated platelets and platelet-derived microparticles, following thrombin-mediated activation of the non-enzymatic cofactors V and VIII, respectively. Tans, G.
  • thrombin In contrast to its prothrombotic effects, thrombin has been shown to influence other aspects of hemostasis. These include its effect as an important physiological anticoagulant.
  • the anticoagulant effect of thrombin is expressed following binding of thrombin to the endothelial cell membrane glycoprotein, thrombomodulin. This is thought to result in an alteration of the substrate s specificity of thrombin thereby allowing it to recognize and proteolytically activate circulating protein C to give activated protein C (aPC) .
  • aPC activated protein C
  • aPC is a serine protease which selectively inactivates the non-enzymatic co-factors Va and Villa resulting in a down-regulation of thrombin formation by the prothrombinase and tenase catalytic complexes, respectively. Esmon, C.T., Science, 235: 1348 (1987) . The activation of protein C by thrombin in the absence of thrombomodulin is poor.
  • Thrombin has also been shown to be a potent direct mitogen for a number of cell types, including cells of mesenchymal origin such as vascular smooth muscle cells. Chen, L.B. and Buchanan, J.M. , Proc. Natl. Acad. Sci. USA, 72 . : 131 (1975) .
  • the direct interaction of thrombin with vascular smooth muscle also results in vasoconstriction.
  • Thrombin acts as a direct secretagogue inducing the release of a number of bioactive substances from vascular endothelial cells including tissue plasminogen activator. Levin, E.G. et al., Thromb. Haemost., .56.: 115 (1986) .
  • the enzyme can indirectly elaborate potent mitogenic activity on vascular smooth muscle cells by the release of several potent growth factors (e.g. platelet-derived growth factor and epidermal growth factor) from platelet a- granules following thrombin-induced activation. Ross, R. , N. Engl. J. Med., 3_14: 408 (1986).
  • abnormal hemostasis With respect to the coronary arterial vasculature, abnormal thrombus formation due to the rupture of an established atherosclerotic plaque is the major cause of acute myocardial infarction and unstable angina. Moreover, treatment of an occlusive coronary thrombus by either thrombolytic therapy or percutaneous transluminal coronary angioplasty (PTCA) is often accompanied by an acute thrombotic reclosure of the affected vessel which requires immediate resolution.
  • PTCA percutaneous transluminal coronary angioplasty
  • venous ⁇ _> vasculature With respect to the venous ⁇ _> vasculature, a high percentage of patients undergoing major surgery in the lower extremities or the abdominal area suffer from thrombus formation in the venous vasculature which can result in reduced blood flow to the affected extremity and a predisposition to pulmonary embolism.
  • Disseminated intravascular coagulopathy commonly occurs within both vascular systems during septic shock, certain viral infections and cancer and is characterized by the rapid consumption of coagulation factors and systemic coagulation which results in the formation of life- threatening thrombi occurring throughout the vasculature leading to widespread organ failure.
  • arterial thrombosis is the result of a complex interaction between fibrin formation resulting from the blood coagulation cascade and cellular components, particularly platelets, which make up a large percentage of arterial thrombi.
  • Heparin the most widely used clinical anticoagulant administered i.v., has not been shown to be universally effective in the treatment or prevention of acute arterial thrombosis or rethrombosis. Prins, M.H. and Hirsh, J., J. Am. Coll. Cardiol., .67: 3A (1991) .
  • the need for safe and effective therapeutic anticoagulants has in one aspect focused on the role of the serine protease thrombin in blood coagulation.
  • thrombin cleavage site on the A ⁇ chain of fibrinogen which is the primary physiological substrate for thrombin, is reported to contain a glycine residue in this position while the cleavage site on the B ⁇ chain contains a serine, as shown below:
  • Peptidyl derivatives having an uncharged residue in the P3 position are said to bind to the active site of thrombin and thereby inhibit the conversion of fibrinogen to fibrin and cellular activation have been reported.
  • These derivatives have either an aldehyde, chloromethyl ketone or boronic acid functionality associated with the PI amino acid.
  • substrate-like peptidyl derivatives such as D-phenylalanyl-prolyl-argininal (D-Phe- Pro-Arg-al) , D-phenylalanyl-prolyl-arginine-chloromethyl ketone (P-PACK) and acetyl-D-phenylalanyl-prolyl- boroarginine (Ac- (D-Phe) -Pro-boroArg) have been reported to s inhibit thrombin by directly binding to the active site of the enzyme. Bajusz, S., Symposia Biologica Hungarica, 25 : 277 (1984), Bajusz, S. et al, J. Med. Chem., 33 .
  • Peptidyl compounds which are said to be active site inhibitors of thrombin but which differ in structure from those containing a uncharged amino acid in the P3 recognition subsite have been reported.
  • the compound, Argatroban also called 2R,4R-4-methyl- 1- [N-2- (3-methyl-l,2,3,4-tetrahydro-8-quinolinesulfonyl) -L- argininal] -2-piperidinecarboxylic acid
  • 2R,4R-4-methyl- 1- [N-2- (3-methyl-l,2,3,4-tetrahydro-8-quinolinesulfonyl) -L- argininal] -2-piperidinecarboxylic acid
  • Argatroban has been reported to be a potent antithrombotic agent in several experimental models of acute arterial thrombosis. Jang, I.K. et al. , in both Circulation, .81: 219 (1990) and Circ. Res., £7: 1552 (1990) .
  • Peptidyl compounds which are said to be inhibitors of thrombin and whose mode of action is thought to be by binding to both the active site and another site on the enzyme have been reported.
  • Hirudin and certain peptidyl derivatives of hirudin have been reported to inhibit both conversion of fibrinogen to fibrin and platelet activation by binding to either both the active site and exo site, or * the exo site only, of thrombin.
  • Hirudin is reported to be a 65 amino acid polypeptide originally isolated from leech salivary gland extracts. It is said to be one of the most potent inhibitors of thrombin known. Marki, W.E. and
  • hirudin has been reported to also effectively inhibit smooth muscle proliferation and the associated restenosis following mechanical damage to a atherosclerotic rabbit femoral artery. Sarembock, I.J. et al., Circulation, £4_* 232 (1991).
  • Hirugen has been reported to be a peptide derived from the anionic carboxy-terminus of hirudin. It is reported to bind only to the anion binding exo-site of thrombin and thereby inhibit the formation of fibrin but not the catalytic turnover of small synthetic substrates which have access to the unblocked active site of the enzyme. Maraganore, J.M. et al. , J. Biol. Chem., 264: 8692 (1989); Naski, M.C. et al. , J. Biol. Chem., 265: 13484 (1990) .
  • the region of hirudin represented by hirugen has been reported, as according to by x-ray crystallographic analysis, to bind directly to the exo site of thrombin. Skrzypczak-Jankun, E. et al., Thromb. Haemostas., J55_: 830 at abstract 507 (1991) . Moreover, the binding of hirugen has also been reported to enhance the catalytic turnover of certain small synthetic substrates by thrombin, indicating that a conformational change in the enzyme active site may accompany occupancy of the exo-site. Liu, L.W. et al. , J. Biol. Chem, 26_6:16977 (1991) . Hirugen also is reported to block thrombin- mediated platelet aggregation. Jakubowski, J.A. and Maraganore, J.M., Blood, 75: 399 (1990) . /o
  • the hirugen-like sequence is said to be linked to this peptide through the C-terminal end of the peptide.
  • hirulogs have been reported to be an effective antithrombotic agents in preventing both fibrin-rich and platelet-rich thrombosis. Maraganone, J.M. et al . , Thromb. Haemostas., 65 : 651 at abstract 17 (1991) .
  • the present invention is directed to novel compounds which are peptide argininals which incorporate lactam groups which are potent inhibitors of thrombin in vivo and in vitro.
  • the present invention is directed to compounds of the formula:
  • R' is hydrogen, alkyl of 1 to about 4 carbon atoms, aryl of about 6 to about 14 carbon atoms or aralkyl of about 6 to about 16 carbon atoms, and R" is NR', OR', R', or SR', with the proviso that R" is not NH, OH, H, or SH, and;
  • Ri is selected from the group consisting of: //
  • alkyl of 1 to about 12 carbon atoms
  • heterocycloalkyl of 4 to about 10 ring atoms with the ring atoms selected from carbon and heteroatoms, wherein the heteroatoms are selected from the group consisting of oxygen, nitrogen, and S(0)i, wherein i is 0, 1, or 2, optionally substituted in the ring carbons with hydroxyl, alkoxyl or alkyl each of which is 1 to about 3 carbons, amino, guanidino, or amidino,
  • aryl of about 6 to about 14 carbon atoms which is optionally mono-, di- or tri-substituted with Yi, Y2, and/or Y3, respectively,
  • heteroaryl of 5 to 14 atoms with the ring atoms selected from carbon and heteroatoms, wherein the heteroatoms are selected from oxygen, nitrogen, and S(0)i, wherein i is 0, 1, or 2, optionally mono-, di- or tri- substituted with Yi, Y2, and/or Y3 , respectively, (9) aralkyl of about 7 to about 15 carbon atoms which is optionally mono-, di-, or tri-substituted in the aryl ring with Yi, Y2 , and/or Y3 , respectively,
  • Z--_ and Z 2 are independently selected from the group consisting of alkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbon atoms, heteroaryl of about 5 to about 14 atoms having 1 to about 9 carbon atoms, aralkyl of about 7 to about 15 carbon atoms, and heteroaralkyl of about 6 to about 11 atoms having about 3 to about 9 carbon atoms, or
  • Yi and Y2 are selected together to be -OC(Z3) (Z4)0-, wherein Z3 and Z4 are independently selected from the group consisting of hydrogen, alkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbon atoms heteroaryl of about 5 to about 14 atoms having 1 to about 9 carbon atoms, aralkyl of about 7 to about 15 carbon atoms, and heteroaralkyl of about 6 to about 11 atoms having about 3 to about 9 carbon atoms,
  • Q is -(CH2)n _ * wherein n is an integer from 1 to 4, or -(CH2)qR4-- wherein q is 1 or 2, and R4 is -S(0) p -, -0-, -N(Rs) -, wherein p is 0, 1, or 2 and R5 is selected from the group consisting of hydrogen, alkyl of 1 to 4 carbon atoms, and acyl of 1 to 4 carbon atoms; IH
  • R2 is selected from the group consisting of hydrogen, alkyl of 1 to 4 carbon atoms or alkenyl of 2 to 4 carbon atoms;
  • Y is selected from the group of Ri substituents, N V with the proviso that Y is not __- / , and pharmaceutically acceptable salts thereof.
  • the present invention is based on our finding that the novel compounds of our invention are active as selective inhibitors of thrombin in vivo and in vi tro .
  • certain of the preferred compounds of the present invention exhibit advantageous selectivity in that they are very potent inhibitors of thrombin but are inactive or significantly less active, (several orders of magnitude less) in inhibiting plasmin and are significantly less active in inhibiting trypsin.
  • This selectivity for inhibition of thrombin gives these compounds a therapeutic advantage in treating or preventing thrombosis in a mammal suspected of having a condition characterized by abnormal thrombosis.
  • Peptidyl arginine aldehydes have been reported to exist in equilibrium structures in aqueous solutions.
  • the present invention is directed to pharmaceutical compositions comprising a therapeutically effective amount of a compound of the present invention and a pharmaceutically acceptable carrier.
  • the present invention is directed to methods of using the compounds and pharmaceutical compositions of the present invention for the prevention of thrombosis in a mammal suspected of having a condition characterized by abnormal thrombosis, comprising administering to said mammal a therapeutically effective amount of a compound the present invention or pharmaceutical composition comprising such a compound.
  • alkenyl refers to unsaturated aliphatic groups having at least one double bond.
  • alkyl refers to saturated aliphatic groups including straight-chain, branched-chain and cyclic groups.
  • alkoxy and alkoxyl refer to a group having the formula, R-0-, wherein R is an alkyl group.
  • alkoxycarbonyl refers to -C(0)0R wherein R is alkyl.
  • alkenyl refers to an alkenyl group substituted with an aryl group.
  • aralkyl refers to an alkyl group substituted with an aryl group. Suitable aralkyl groups include benzyl, picolyl, and the like, all of which may be optionally substituted.
  • aryl refers to aromatic groups which have at least one ring having a conjugated pi electron system and includes carbocyclic aryl, heterocyclic aryl and biaryl groups, all of which may be optionally substituted.
  • aryloxy refers to a group having the formula, R-O-, wherein R is an aryl group.
  • amino acid refers to natural amino acids, unnatural amino acids, and amino acid analogs in their D and L stereoisomers if their structure allow such stereoisomeric forms. Natural amino acids include alanine (Ala) , arginine (Arg) , asparagine (Asn) , aspartic acid (Asp) , cysteine (Cys) , gluta ine (Gin) , glutamic acid
  • Glu glycine
  • Gly histidine
  • His histidine
  • isoleucine lie
  • leucine Leu
  • lysine Lys
  • methionine Met
  • phenylalanine Phe
  • proline Pro
  • Ser serine
  • Thr threonine
  • Trp tryptophan
  • Tyr tyrosine
  • valine Val
  • Unnatural amino acids include, but are not limited to azetidinecarboxylic acid, 2-aminoadipic acid, 3-aminoadipic acid, beta-alanine, aminopropionic acid, 2- aminobutyric acid, 4-aminobutyric acid, 6-aminocaproic acid, 2-aminoheptanoic acid, 2-aminoisobutyric acid, 3- aminoisobutyric acid, 2-aminopimelic acid, 2,4 diaminoisobutyric acid, desmosine, 2,2 ' -diaminopimelic acid, 2,3-diaminopropionic acid, N-ethylglycine, N- ethylasparagine, hydroxylysine, allo-hydroxylysine, 3- hydroxyproline, 4-hydroxyproline, isodesmosine, allo- isoleucine, N-methylglycine, N-methylisoleucine, N- methylvaline, norvaline, norleucine, orni
  • Amino acid analogs include the natural and unnatural amino acids which are chemically blocked, reversibly or irreversibly, or modified on their N- terminal amino group or their side-chain groups, as for example, methionine sulfoxide, methionine sulfone, S- (carboxymethyl) -cysteine, S- (carboxymethyl) -cysteine sulfoxide and S- (carboxymethyl) -cysteine sulfone.
  • amino acid analog refers to an amino acid wherein either the C-terminal carboxy group, the N- terminal amino group or side-chain functional group has been chemically modified to another functional group.
  • aspartic acid- (beta-methyl ester) is an amino acid analog of aspartic acid
  • N-ethylglycine is an amino acid analog of glycine
  • alanine carboxamide is an amino acid analog of alanine.
  • amino acid residue refers to radicals having the structure: (1) -C(0)-R-NH-, wherein R typically is -CH(R')-, wherein R' is H or a carbon containing
  • aryl refers to phenyl substituted by carbocyclic or heterocyclic aryl as defined herein, ortho, meta or para to the point of attachment of the phenyl ring.
  • Carbocyclic aryl refers to aromatic groups wherein the ring atoms on the aromatic ring are carbon atoms. Carbocyclic aryl groups include monocyclic carbocyclic aryl groups and naphthyl groups, all of which may be optionally substituted. Suitable carbocyclic aryl groups include phenyl and naphthyl. Suitable substituted carbocyclic aryl groups include indene and phenyl substituted by one to two substituents such being advantageously lower alkyl, hydroxy, lower alkoxy, lower alkoxycarbonyl, halogen, trifluoromethyl, nitro, and cyano.
  • Substituted naphthyl refers to 1- or 2-naphthyl substituted by lower alkyl, lower alkoxy, or halogen.
  • Cycloalkenyl refers to a cyclic alkenyl group. Suitable cycloalkenyl groups include, for example, cyclopentenyl and cyclohexenyl.
  • Cycloalkyl refers to a cyclic alkyl group. Suitable cycloalkyl groups include, for example, cyclohexyl, cyclopropyl, cyclopentyl, and cycloheptyl.
  • Cyclohexylmethyl refers to a cyclohexyl group attached to CH2.
  • halogen refers to fluorine, chlorine, bromine and iodine.
  • Heteroaralkenyl refers to an alkenyl group substitued with a heteroaryl, and includes those heterocyclic systems described in "Handbook of Chemistry and Physics", 49th edition, 1968, R.C. Weast, editor;The Chemical Rubber Co., Cleveland, OH. See particularly Section C, Rules for Naming Organic Compounds, B. Fundamental Heterocyclic Systems.
  • Heteroaralkyl refers to an alkyl group substituted with a heteroaryl, and includes those heterocyclic systems described in "Handbook of Chemistry and Physics", 49th edition, 1968, R.C. Weast, editor,*The Chemical Rubber Co., Cleveland, OH. See particularly Section C, Rules for Naming Organic Compounds, B. Fundamental Heterocyclic Systems.
  • Heteroaryl refers to aryl groups having from 1 to 9 carbon atoms and the remainder of the atoms are heteroatoms, and includes those heterocyclic systems described in "Handbook of Chemistry and Physics", 49th edition, 1968, R.C. Weast, editor,*The Chemical Rubber Co., Cleveland, OH.
  • Suitable heteroatoms include oxygen, nitrogen, S(0)i, wherein i is 0, 1 or 2, and suitable heterocyclic aryls include furanyl, thienyl, pyridyl, pyrrolyl, pyrimidyl, pyrazinyl, i idazolyl, and the like.
  • Heterocyclo refers to a reduced heterocyclic ring system comprised of carbon, nitrogen, oxygen and/or sulfur atoms, and includes those heterocyclic systems described in / ⁇ ?
  • Heterocycloalkyl refers to an alkyl group substituted with a heterocyclo group, and includes those heterocyclic systems described in "Handbook of Chemistry and Physics", 49th edition, 1968, R.C. Weast, editor;The Chemical Rubber Co., Cleveland, OH. See particularly Section C, Rules for Naming Organic Compounds, B. Fundamental Heterocyclic Systems.
  • lower referred to herein in connection with organic radicals or compounds defines such with up to and including 5, preferably up to and including 4 and advantageously one or two carbon atoms.
  • Such groups may be straight chain or branched chain.
  • Perfluoroalkyl refers to an alkyl group which has every hydrogen replaced with fluorine.
  • Perfluoroaryl refers to an aryl group which has every hydrogen replaced with fluorine.
  • Perfluoroaryl alkyl refers an aralkyl group in which every hydrogen on the aryl moiety is replaced with fluorine.
  • “Pharmaceutically acceptable salt” includes salts of the compounds of the present invention derived from the combination of such compounds and an organic or inorganic acid. In practice the use of the salt form amounts to use of the base form. The compounds of the present invention are useful in both free base and salt form, with both forms being considered as being within the scope of the present invention.
  • Arg-al refers to the residue of L-argininal which has the formula: — HN ⁇ CHO
  • Arg-ol refers to the residue of L-argininol which has the formula:
  • N-alpha-t-butoxycarbonyl-N9-nitro-L-arginine refers to the compound which has the formula:
  • Boc refers to t-butoxycarbonyl
  • BoclS02 refers to benzylsulfonyl.
  • DCC refers to N,N'-dicyclohexylcarbodiimide.
  • EDC refers to l-ethyl-3- (3-dimethylamino- propyl)carbodiimide hydrochloride salt.
  • HBTU refers to 2- (lH-benzotriazol-1-yl) -1, 1,3,3- tetramethyluronium hexafluorophosphate.
  • HCl refers to hydrochloric acid
  • HPLC refers to high pressure liquid chromatography.
  • HOBt refers to 1-hydroxybenzotriazole monohydrate.
  • i-Pr refers to isopropyl
  • LiAlH4 refers to lithium aluminum hydride.
  • LiAlH2 (OEt)2 refers to lithium aluminum dihydride diethoxide.
  • Me refers to methyl.
  • OEt refers to ethoxy.
  • THF refers to tetrahydrofuran.
  • TLC refers to thin layer chromatography
  • Figure 1 depicts the reaction scheme for preparation of an intermediate used for the synthesis of the compounds of the present invention.
  • "i” through “iv” are defined as: i) hydrogen gas, 10% palladium on carbon; ii) glyoxylic acid; iii) hydrogen gas, 10% palladium on carbon; and iv) 50-60°C, N,N- dimethylformamide.
  • Figure 2 depicts the reaction scheme for preparation of an intermediate used for the synthesis of the compounds of the present invention.
  • "i” through “iii” are defined as: i) HOBt, DCC and triethylamine; ii) methyl iodide; and iii) sodium hydride later followed by ethyl acetate and water.
  • Figure 3 depicts a preferred reaction scheme for a synthesis of certain compounds of the present invention.
  • "i” through “v” are defined as: i) HOBt, EDC, 4-methylmorpholine; ii) IN HCl in ethyl acetate; iii) potassium carbonate, R1-SO2CI wherein Ri is as defined herein; iv) hydrogen gas, 10% palladium on carbon; and v) dimethylsulfoxide, toluene, dichloroacetic acid and EDC.
  • Figure 4 depicts a preferred reaction scheme for a synthesis of certain compounds of the present invention.
  • i through “vi” are defined as: i) methanol saturated with HCl; ii) triethylamine, R-SO2CI wherein R is alkyl, aryl or aralkyl; iii) 1.0M lithium hydroxide; iv) HOBt, N,N-diisopropylethylamine, EDC; v) hydrogen gas, 10% pallidium on carbon; and vi) 3N HCl.
  • Figure 5 depicts the anticoagulant effect of BzlS02 ⁇ norVal (cyclo) -Gly-Arg-al measured in citrated rat (•) and human (m) plasma using the activated partial thromboplastin time (APTT) assay.
  • the control clotting times (0 inhibitor) for rat and human plasma were 20 sec and 28 sec, respectively.
  • the concentration of BzlS ⁇ 2 ⁇ norVal (cyclo) -Gly-Arg-al which caused a doubling of the control clotting time m rat and human plasma was 22.1 micromolar and 12.8 micromolar, respectively.
  • the data is the mean of two independent determinations.
  • Figure 6 depicts a preferred reaction scheme for the preparation of certain compounds of the present invention.
  • i)-x are defined as: i) di-t-butyl dicarbonate, sodium bicarbonate in tetrahydrofuran and water; ii) lithium bis (trimethylsilyl)amide and tetrahydrofuran; iii) benzyl bromoacetate,- iv) ammonium chloride; v) HCl and ethyl acetate; vi) triethylamine, acetonitrile and R1-SO2-CI, where Ri is as defined herein; vii) hydrogen, palladium on carbon and ethanol; viii) acetonitrile, N ⁇ -nitro-L-argininal ethyl cyclol, hydrochloride salt, 1-hydroxybenzotriazole monohydrate, 1- ethyl-3- (3-dimethylaminopropyl)carbodiimide hydrochloride salt and
  • Figure 7 depicts the anticoagulant effect of N- (BnS02-norLeu(cyclo)-Gly) -L-argininal measured in citrated human plasma, closed circles (•) , using the activated partial thromboplastin time (APTT) assay.
  • the control clotting times (0 inhibitor) for human plasma was 29 seconds.
  • the concentration of N- (BnS ⁇ 2-norLeu(cyclo)- Gly) -L-argininal which caused a doubling of the control clotting time in human plasma was 4.5 micromolar.
  • the data is the mean of two independent determinations.
  • Figure 8 depicts a preferred reaction scheme for the preparation of certain compounds of the present invention.
  • i)-v) are defined as: i) hydrogen gas and palladium on carbon; ii) N9-nitro-L-argininal ethyl cyclol hydrochloride salt, 1-hydroxybenzotriazole monohydrate, 1- ethyl-3- (3-dimethylaminopropyl)carbodiimide hydrochloride salt, and N,N-diisopropylamine; iii) hydrogen gas and palladium on carbon in ethanol, acetic acid and water; iv) 3N HCl; and v) HPLC purification with 0.1% trifluoroacetic acid in acetonitrile and water.
  • Figure 9 depicts a preferred reaction scheme for the J ⁇ H preparation of certain compounds of the present invention.
  • i)-vii) are defined as: i) sodium hydride; ii) methyl iodide; iii) hydrogen gas and palladium on carbon; iv) N9-nitro-L-argininal ethyl cyclol hydrochloride salt, 1-hydroxybenzotriazole monohydrate, 1- ethyl-3- (3-dimethylaminopropyl)carbodiimide hydrochloride salt, and N,N-diisopropylamine; v) hydrogen gas and palladium on carbon in ethanol, acetic acid and water; vi)
  • Figure 10 depicts a preferred reaction scheme for preparation of compound 30, described in Examples 46 through 55.
  • the steps are defined as follows:
  • (21) from (20) : 1. H2, Pd/C, 35 PSI, MeOH, H2O, HOAc, 2.5 hours; 2. OHCCO2H, room temperature, 20 hours; 3. H2, Pd/C, 40 PSi, 37 hours; 4. DMF, 50-60°C, 2.5 hours; 5. extractive workup, for a 67% overall yield.
  • (23) from (21) : 1. EtOH, HCl, 0°C to room temperature, quantitative yield of (22); 2. PhCHO, Et3N, MgS04, CH2CI2, 0°C to room temperature, 89% yield of (23) .
  • Figure 11 depicts a preferred reaction scheme for preparation of compound (39), described in Examples 56 through 64.
  • the steps are defined as follows:
  • (32) PhCHO, benzene, reflux, -H2O, quantitative yield of (31); 2.
  • (34) from (32) : 1. IN, HCl, room temperature, 5 hours to yield (33); 2. Cbz-OSu, NaHC03, THF, H2 ⁇ , 0°C to room temperature, 14 hours, 79% overall yield.
  • (35) In the synthesis of (34) : 1.
  • Figure 12 depicts a preferred reaction scheme for preparation of compound (47) , described in Examples 65 through 72.
  • the steps are defined as follows:
  • (41) from (21): 1. BnBr, K2CO3, DMF, CH3CN, reflux, 96% yield; 2. 5N HCl, EtOAc, room temperature, 3 hours, quantitative yield of (41) .
  • (42) from (41), PhCH CHO, Na(OAc)3BH, triethylamine, DCE, room temperature, 66% yield of (42) .
  • (44) from (42) 1. B0C2O, NaHC ⁇ 3, THF, H2O, room temperature, 1.5 hours, 70% yield of (43); 2.
  • Figure 13 depicts a preferred reaction scheme for preparation of compound (56) , described in Examples 73 through 80. In this figure, the steps are defined as follows: In the synthesis of (50) from (48) : 1.
  • R' is hydrogen, alkyl of 1 to about 4 carbon atoms, aryl of about 6 to about 14 carbon atoms or aralkyl of about 6 to about 16 carbon atoms, and R" is NR', OR', R', or SR', with the proviso that R" is not NH, OH, H, or SH, and;
  • Ri is selected from the group consisting of:
  • alkyl of 1 to about 12 carbon atoms
  • heterocycloalkyl of 4 to about 10 ring atoms with the ring atoms selected from carbon and heteroatoms, wherein the heteroatoms are selected from the group consisting of oxygen, nitrogen, and S(0)i, wherein i is 0, 1, or 2, optionally substituted in the ring carbons with hydroxyl, alkoxyl or alkyl each of which is 1 to about 3 carbons, amino, guanidino, or amidino,
  • alkenyl of about 3 to about 6 carbon atoms which is optionally substituted with cyclic alkyl of about 5 to about 8 carbon atoms, which optionally is substituted in the ring carbons with hydroxyl, amino, guanidino, amidino, or alkoxyl or alkyl each of which is 1 to about 3 carbons,
  • heteroaryl of 5 to 14 atoms with the ring atoms selected from carbon and heteroatoms, wherein the heteroatoms are selected from oxygen, nitrogen, and S(0)i, wherein i is 0, 1, or 2, optionally mono-, di- or tri- substituted with Yi, Y2, and/or Y3 , respectively,
  • Z3 and Z4 are independently selected from the group consisting of hydrogen, alkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbon atoms heteroaryl of about 5 to about 14 atoms having 1 to about 9 carbon atoms, aralkyl of about 7 to about 15 carbon atoms, and heteroaralkyl of about 6 to about 11 atoms having about 3 to about 9 carbon atoms,
  • Q is -(CH2)n _ - wherein n is an integer from 1 to 4, or -(CH2)qR4 ⁇ , wherein q is 1 or 2, and R4 is -S(0) p -, -0-, -N(R5)-, wherein p is 0, 1, or 2 and R5 is selected from the group consisting of hydrogen, alkyl of 1 to 4 carbon atoms, and acyl of 1 to 4 carbon atoms;
  • R2 is selected from the group consisting of hydrogen, alkyl of 1 to 4 carbon atoms or alkenyl of 2 to 4 carbon atoms;
  • Y is selected from the group of Ri substituents
  • Preferred X groups include a direct link, -SO2-,
  • X groups include a direct link and -SO2-.
  • Preferred Ri groups include alkyl, aralkyl and aryl groups . Suitable aryl groups include substituted or unsubstituted phenyl and naphthyl. Preferred aryl substituents include, -C(0)OH, -C(0)OZ ⁇ , -CH3 , -OCH3, and -CF3. Meta and/or ortho substitutions are preferred.
  • Especially preferred Ri groups include aralkyl groups. Particularly preferred Ri groups include substituted or unsubstituted benzyl groups. Cyclohexyl and cyclohexylmethyl are especially preferred Ri groups.
  • Preferred R2 groups include hydrogen.
  • Preferred Y groups are selected from the group consisting of
  • phenyl- (CH2)i- wherein i is an integer from 0 to 3 , and the phenyl is optionally mono-, di-, or tri-substituted with with Yi, Y2, and/or Y3, respectively, as defined above,
  • heteroaryl- (CH2)i- wherein i is an integer from 0 to 3 , and the heteroaryl is optionally mono-, di-, or tri-substituted with Yi, Y2, and/or Y3, respectively, as defined above,
  • heterocycloalkyl- (CH2)i- wherein i is an integer from 0 to 3, and the heterocycloalkyl is optionally substituted with hydroxyl, alkoxyl or alkyl each of which is 1 to about 3 carbons,
  • More preferred Y groups include aralkyl or cycloalkyl groups. Particularly preferred are substituted or unsubstituted benzyl and 1-naphthylmethyl groups.
  • Preferred aromatic ring substituents include -C(0)OH, -C(0)OZi, -CH3, -OCH3, and -CF3. Meta and/or ortho substituents are preferred.
  • Preferred cycloalkyl groups include those containing 5-8 ring carbons .
  • Preferred cycloalkyl substitutents include -C(0)OH, -C(0)OZ ⁇ , -CH3, -OCH3.
  • X is -S(0)2-- Rl is substituted or unsubstituted aralkyl
  • Q is -(CH2)2 _
  • R2 is hydrogen.
  • Ri is substituted or unsubstituted benzyl.
  • X is -S(0)2-- Rl is substituted or unsubstituted aralkyl
  • Q is -(CH2)3 ⁇
  • R2 is hydrogen.
  • Ri is substituted or unsubstituted benzyl.
  • Preferred compounds of this invention include those whose synthesis is described in Examples 8 (and 18) , 26, 34, 36(a) through 36 (y) , 40(b), 55, 64, 72(a) and 72 (b) , 80, and 82(a) through 82(f) .
  • Especially preferred compounds of the present invention include N-benzylsulfonyl-norVal (cyclo) -Gly-L-argininal (Example 8, 18),
  • the present invention is directed to salts of the compounds of formula (I) .
  • Salt includes within its definition, salts of the compounds of the present invention derived from the combination of such compounds and an organic or inorganic acid.
  • the use of the salt form amounts to use of the base form.
  • the compounds of the present invention are useful in both free base and salt form, with both forms being considered S3 as being within the scope of the present invention.
  • These salts include acid addition salts, for example, salts of trifluoroacetic acid, hydrochloric acid, hydrobromic acid, acetic acid, benzene sulfonic acid and other suitable acid addition salts.
  • Figure 1 exemplifies a preferred reaction scheme for the preparation of one preferred intermediate 3 used in the preparation of the compounds of the present invention.
  • Example 1 provides the details of the preferred scheme.
  • N-alpha-Boc-N-delta- benzyloxycarbonyl-L-ornithine 1 is hydrogenated with hydrogen gas and palladium on carbon to give 2, which is then reacted with glyoxylic acid, hydrogenated with hydrogen gas and palladium on carbon, and heated at an elevated temperature to give 3.
  • Figure 2 exemplifies a preferred reaction scheme for the preparation of one preferred intermediate, 2-5, used in the preparation of the compounds of the present invention.
  • Examples 19 through 21 provide the details of the preferred scheme.
  • N-alpha-Boc-L-methionine 2-1 and glycine methyl ester hydrochloride 2-2 are coupled by use of a carbodiimide and HOBt to give 2-3.
  • the sulfur atom on the methionine side chain of 2-3 is alkylated with methyl iodide to give 2-4.
  • 2-4 is treated with 3i sodium hydride which results in cyclization to give 2-5.
  • the compounds of the present invention may be prepared by the preferred reaction schemes depicted in Figures 3 and 4. Examples 2 through 8 provide the details of the preferred scheme of Figure 3, while Examples 9 through 18 provide the same for the preferred scheme of Figure 4.
  • 3-3 is coupled to (S) -NS-nitroargininol hydrochloride 3-4 (prepared from (S) -N-alpha-Boc-N9-nitroarginine as disclosed in Examples 2 through 3) by carbodiimide coupling to give 3-5.
  • 3-5 is treated with hydrogen chloride to remove the Boc group to give the hydrochloride 3-6, which is then reacted with a sulfonyl chloride, depicted by R ⁇ -S(0)2-Cl, to give 3-7.
  • Rl is as defined herein.
  • 3-7 is hydrogenated with hydrogen gas and palladium on carbon to remove the N9- nitro group, then the hydroxy group is oxidized using dimethylsulfoxide, dichloroacetic acid, toluene and EDC to give 3-8.
  • the preferred means of chemically coupling include formation of a peptide bond by using conventional coupling reagents known in the art. See 3 ⁇ Y Bodanszky, N. , Peptide Chemistry, pp. 55-73, Springer-
  • the chemical coupling may be either by means of one-step or two-step coupling.
  • one-step coupling the two coupling partners are coupled directly.
  • Preferred coupling reagents for one-step coupling include DCC with HOBt, EDC with HOBt, HBTU or TBTU.
  • two-step coupling an activated ester or anhydride of the C-terminal carboxy group of one coupling partner is formed prior to its coupling to the other coupling partner.
  • the compounds of the present invention also may be prepared by the preferred reaction scheme depicted in Figure 6. Examples 12 through 15 and 27 through 34 provide the details of this preferred scheme.
  • the amine of D- (alpha)- or L- (-) -alpha-amino-epsilon-caprolactam 6-1 is protected by treatment with di-t-butyldicarbonate and sodium bicarbonate in THF and water to give 6-2, which is then reacted with lithium bis (trimethylsilyl)amide in THF, followed by benzyl bromoacetate. The reaction is quenched with a saturated ammonium chloride solution to give 6-3.
  • the Boc protecting group of 6-3 is then removed by treatment with 5 N HCl in ethyl acetate to provide the HCl salt 6-4, which is then reacted with triethylamine and R1SO2CI in acetonitrile to give the sulfonamide 6-5.
  • 6-5 is then hydrogenated with hydrogen gas and palladium on carbon in a Parr Shaker to give 6-6.
  • 6-6 is coupled to N ⁇ -nitro-L-argininal ethyl cyclol hydrochloride salt (prepared as described in Examples 12 through 15) by carbodiimide coupling to give 6-7.
  • (S) -3- [ (tert-butoxycarboxyl)amino] -2-oxo- hexahydro-1-azepine acetic acid, benzyl ester 6-3 is deprotected to give the acid 8-10 by treatment with hydrogen gas and palladium on carbon in a Parr shaker at 40 psi. 8-10 is then coupled to N ⁇ -nitro-L-argininal ethyl cyclol using 1-hydroxybenztriazole monohydrate, 1- ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride salt, and then N,N-diisopropylamine to give 8-11.
  • the N ⁇ - nitro group of 8-11 is removed by treatment with hydrogen gas and palladium on carbon in ethanol, water, and acetic acid at 50 psi. This reaction produces acetic acid salt 8-12.
  • the ethyl cyclol group of 8-12 is hydrolyzed by treatment with 3N HCl, followed by HPLC purification with 0.1% trifluoroacetic acid in acetonitrile and water to produce 8-13.
  • the scheme of Figure 9 differs in that (S)- 3- [ (tert-butoxycarbonyl)amino] -2-oxo-hexahydro-l-azepine acetic acid, benzyl ester is first reacted with sodium hydride followed by methyl iodide to prepare the alkylated Boc protected amine 14. 14 is deprotected to form 15, which is treated to the same reactions as acid 8-10 in Figure 8.
  • the method used in Figure 9 illustrates the reactions to make compounds in which X is a direct link to Rl, when Ri is other than hydrogen.
  • Figure 10 depicts the synthesis of a preferred species.
  • Intermediate 20 is catalytically hydrogenated, condensed with glyoxylic acid, catalytically rehydrogenated and the resultant intermediate is thermally dehydrated in warm dimethylformamide to afford the lactam 21.
  • Treatment of 21 with dry hydrogen chloride in ethanol simultaneously effects both deblocking of the Boc group as well as esterification of the carboxyl moiety and produces a inoester hydrochloride 22.
  • Condensation of 22 with benzaldehyde in the presence of a base such as triethylamine employing a dehydrating agent such as magnesium sulfate affords the imine derivative 23.
  • Figure 11 depicts the synthesis of a preferred species which is the 7-membered ring analog of compound 30.
  • ⁇ -Amino- ⁇ -caprolactam is condensed with benzaldehyde in azeotropically refluxing benzene solution to afford imine 31.
  • Deprotonation f 31 with a selective base such as potassium tert-butoxide, lithium N,N-diisopropylamide or lithium bis (trimethylsilyl)amide delivers the alkylated compound 32.
  • Hydrolysis of 32 with dilute hydrochloric acid at ambient temperature affords the amine hydrochloride salt 33.
  • N-Cbz-lactam intermediate 34 Protection of the amino group by reaction of 33 with N- (benzyloxycarbonyloxy) -succinimide and sodium bicarbonate in aqueous THF provides N-Cbz-lactam intermediate 34.
  • Cleavage of 3 the tert-butyl ester moiety of 35 is effected with trifluoroacetic acid in methylene chloride at about 0° to room temperature and affords the carboxylic acid derivative 36.
  • the amino group of 42 is protected by reaction with di-tert-butyl dicarbonate in the presence of saturated sodium bicarbonate in THF and provides 43.
  • Catalytic hydrogenation of 43 using palladium on charcoal leads to the carboxylic acid 44.
  • Standard peptide coupling of 44 and N ⁇ -nitro-argininal ethyl cyclol using EDC, HOBt and N,N-diisopropylethylamine in an inert solvent such as acetonitrile at ambient temperature gives 45.
  • Catalytic hydrogenation of 45 in ethanol, acetic acid and water using a palladium on charcoal catalyst affords the acetate salt precursor 46.
  • hydrolysis of 46 with dilute hydrochloric acid at ambient temperature followed by HPLC purification gives the novel compound 47.
  • Figure 13 depicts the synthesis of a preferred species which is the 7-membered ring homolog of target 47.
  • Compound 48 prepared as described above (equivalent to compound 6-2 from figure 6) , is treated with a hindered base such as lithium bis- (trimethylsilyl)amide and is alkylated with ethyl bromoacetate to afford 49.
  • Cleavage of the Boc group of 49 with hydrogen chloride in ethanol gives amine hydrochloride salt 50.
  • Reductive amination of salt 50 with phenylacetaldehyde using sodium triacetoxyborohydride and triethylamine in an inert solvent such as 1,2-dichloroethane delivers the amine derivative 51.
  • the amino group of 51 is protected by reaction with di-tert-butyl dicarbonate in the presence of saturated sodium bicarbonate in THF and provides 52.
  • Hydrolysis of 52 in an aqueous alcoholic solvent with a strong base like lithium, sodium or potassium hydroxide followed by protonation using a strongly acidic cation exchange resin such as Dowex® 50WX8-400 affords the N-boc- protected lactam carboxylic acid intermediate 53.
  • Standard peptide coupling of 53 and N ⁇ -nitro-argininal ethyl cyclol using EDC, HOBt and N,N-diisopropylethylamine in an inert solvent such as acetonitrile at ambient temperature gives 54.
  • the reagent is also commercially available. Generally, the N ⁇ -nitro compound is treated with boron tris (trifluoroacetate) in trifluoroacetic acid at 0°C. Fieser, M. and Fieser, L. F., Reagents for Organic Synthesis. p. 46, John Wiley & Sons, New York (1974); yo
  • titanium trichloride is another preferred reagent for selective nitro group cleavage.
  • This reagent is commercially available.
  • the N ⁇ nitro compound is treated with titanium trichloride in aqueous methanol containing an ammonium acetate buffer followed by exposure of the reaction mixture to air or dimethyl sulfoxide.
  • an even more preferred method is to use the di-N-t- butoxycarbonyl protecting group for the L-argininal moiety for groups incompatible with hydrogenation with palladium on carbon.
  • alpha-N-benzyloxycarbonyl-omega, omega'-di-N-t-butoxycarbonylarginine is dissolved in acetonitrile and treated with hydroxybenzotriazole and 1- ethyl-3-(3-dimethylamino-propyl)carbodiimide HCl salt to form alpha-N-benzyloxycarbonyl-omega, omega'-di-N-t- butoxycarbonyl-L-arginine lactam.
  • the lactam is reduced by treatment with LiAlH4 in THF at -70°C to provide alpha- N-benzyloxycarbonyl-omega, omega'-di-N-t-butoxycarbonyl-L- argininal.
  • This aldehyde is protected as the diethyl acetal by treatment with ethanol and HCl.
  • the N- benzyloxycarbonyl protecting group is removed by treatment with hydrogen gas and palladium on carbon to give omega, omega'-di-N-t-butoxycarbonyl-L-argininal diethyl acetal, HCl salt.
  • This protected L-argininal moiety can then be coupled to a desired carboxylic acid by treatment with N- hydroxybenzotriazole and l-ethyl-3-(3-dimethylamino- propyl)carbodiimide HCl salt.
  • the diethyl acetal and the di-BOC protecting groups are removed by treatment with hexafluorophosphoric acid in acetonitrile at 0°C.
  • the reaction is quenched with 2.5 M aqueous sodium acetate until pH 4 is reached.
  • the mixture is filtered through a 2 micron filter.
  • Preparative HPLC using 0.1% CF3COOH in 10-40% aqueous acetonitrile provides the trifluoroacetate salt of the desired substituted L-argininal compound. HI 3. Selection of Preferred Compounds
  • the compounds of the present invention are screened for their ability to inhibit thrombin, plasmin, recombinant tissue plasminogen activator (rt-PA) , activated protein C (aPC) , chymotrypsin, fXa and trypsin as set forth below.
  • Certain of the preferred compounds are distinguished by their ability to inhibit thrombin, while not substantially inhibiting plasmin, t-PA, aPC, chymotrypsin, and trypsin.
  • the term "not substantially inhibiting" means that the IC50 (or Ki) for plasmin, t-PA, aPC, chymotrypsin, and trypsin for a given compound is greater than or equal to its IC50 (or Ki, respectively) for thrombin.
  • the compounds of the present invention are dissolved in buffer to give solutions containing concentrations such that assay concentrations range from 0 to 100 micromolar.
  • a chromogenic synthetic substrate is added to a solution containing test compound and the enzyme of interest and the residual catalytic activity of that enzyme is determined spectrophometrically.
  • the IC50 of a compound of the present invention is determined from the rate of substrate turnover caused by the specific enzyme being measured. IC50 is that concentration of test compound giving 50% inhibition of the rate of substrate turnover.
  • the Ki of a compound of the present invention is determined from the rate of substrate turnover caused by the specific enzyme being measured at various enzyme concentrations. Ki is that concentration of test compound giving 50% inhibition of the rate of substrate turnover. Examples A and B provide an exemplar of the in vitro assays used to select the compounds of the present invention.
  • Certain of the preferred compounds of the present invention have a Ki of about 0.001 to about 200 nM in the thrombin assay. Especially preferred compounds have a Ki of about 0.001 to about 50 nM. The more especially preferred compounds have a Ki of about 0.001 to about. 10 J- nM.
  • Certain of the preferred compounds of the present invention have a IC50 for plasmin, t-PA, aPC, chymotrypsin, and trypsin which is at least 10 times greater than its IC50 for thrombin.
  • Especially preferred compounds have an IC50 for plasmin, rt-PA, aPC, chymotrypsin, and trypsin which is about 20 to about 100,000 times greater than its IC50 for thrombin. More especially preferred compounds have an IC50 for plasmin, rt-PA, aPC, chymotrypsin, and trypsin which is about 100 to about 1,000,000 times greater than its IC50 for thrombin.
  • a compound of the present invention has an IC50 with respect to plasmin, rt-PA, aPC, chymotrypsin, or trypsin which is greater than the highest concentration of compound tested, the IC50 is taken to be that highest concentration of compound.
  • the present invention encompasses pharmaceutical compositions prepared for storage or administration which comprise a therapeutically effective amount of a compound of the present invention in a pharmaceutically acceptable carrier.
  • the "therapeutically effective amount" of a compound of the present invention will depend on the route of administration, the type of mammal being treated, and the physical characteristics of the specific mammal under consideration. These factors and their relationship to determining this amount are well known to skilled practitioners in the medical arts. This amount and the method of administration can be tailored to achieve optimal efficacy but will depend on such factors as weight, diet, concurrent medication and other factors which as noted those skilled in the medical arts will recognize.
  • the “therapeutically effective amount” of the compound of the present invention can range broadly depending upon the desired affects and the therapeutic indication. Typically, dosages will be between about 0.01 mg/kg and 100 H3 mg/kg body weight, preferably between about 0.01 and 10 mg/kg, body weight.
  • “Pharmaceutically acceptable carriers” for therapeutic use are well known in the pharmaceutical art, and are described, for example, in Remington' s Pharmaceutical Sciences, Mack Publishing Co. (A.R. Gennaro edit. 1985) .
  • sterile saline and phosphate-buffered saline at physiological pH may be used.
  • Preservatives, stabilizers, dyes and even flavoring agents may be provided in the pharmaceutical composition.
  • sodium benzoate, sorbic acid and esters of p-hydroxybenzoic acid may be added as preservatives. Id. at 1449.
  • antioxidants and suspending agents may be used. Id.
  • compositions of the present invention may be formulated and used as tablets, capsules or elixers for oral administration; suppositories for rectal administration; sterile solutions and suspensions for injectable administration; and the like.
  • the dose and method of administration can be tailored to achieve optimal efficacy but will depend on such factors as weight, diet, concurrent medication and other factors which those skilled in the medical arts will recognize.
  • injectable pharmaceutcial compositions can be prepared in conventional forms, either as liquid solutions or suspensions, solid forms suitable for solution or suspension in liquid prior to injection, or as emulsions.
  • Suitable excipients are, for example, water, saline, dextrose, mannitol, lactose, lecithin, albumin, sodium glutamate, cysteine hydrochloride, or the like.
  • the injectable pharmaceutical compositions may contain minor amounts of nontoxic auxilliary substances, such as wetting agents, pH buffering agents, and the like.
  • absorption enhancing preparations e.g., liposomes may be utilized.
  • Compounds of the present invention when made and HH selected as disclosed are useful as potent inhibitors of thrombin in vitro and in vivo. As such, these compounds are useful as in vitro diagnostic reagents to prevent the clotting of blood and as in vivo pharmaceutical agents to prevent thrombosis in mammals suspected of having a condition characterized by abnormal thrombosis.
  • the compounds of the present invention are useful as in vitro diagnostic reagents for inhibiting clotting in blood drawing tubes.
  • stoppered test tubes having a vaccum therein as a means to draw blood obtained by venipuncture into the tube is well known in the medical arts. Kasten, B.L. , "Specimen Collection", Laboratory Test Handbook, 2nd Edition, Lexi-Comp Inc., Cleveland pp. 16-17 (Edits. Jacobs, D.S. et al. 1990) .
  • Such vacuum tubes may be free of clot-inhibiting additives, in which case, they are useful for the isolation of mammalian serum from the blood.
  • clot-inhibiting additives such as heparin salts, EDTA salts, citrate salts or oxalate salts
  • clot-inhibiting additives such as heparin salts, EDTA salts, citrate salts or oxalate salts
  • the compounds of the present invention are potent inhibitors of factor Xa or thrombin, and as such, can be incorporated into blood collection tubes to prevent clotting of the mammalian blood drawn into them.
  • the compounds of the present invention are used alone, in combination of other compounds of the present invention, or in combination with other known inhibitors of clotting, in the blood collection tubes.
  • the amount to be added to such tubes is that amount sufficient to inhibit the formation of a clot when mammalian blood is drawn into the tube.
  • the addition of the compounds to such tubes may be accomplished by methods well known in the art, such as by introduction of a liquid composition thereof, as a solid composition thereof, or liquid composition which is lyophilized to a solid.
  • the compounds of the present invention are added to blood collection tubes in such amounts that, when combined with 2 to 10 mL of mammalian blood, the concentration of such compounds will be sufficient to inhibit clot formation. Typically, the required concentration will be about 1 to 10,000 nM, with 10 to 1000 nM being preferred.
  • the compounds of the present invention are useful as a pharmaceutical agent for preventing thrombosis in a mammal suspected of having a condition characterized by abnormal thrombosis.
  • abnormal thrombosis Conditions characterized by abnormal thrombosis are well known in the medical arts and include those involving the arterial and venous vasculature of mammals. With respect to the coronary arterial vasculature, abnormal thrombosis (thrombus formation) characterizes the rupture of an established atherosclerotic plaque which is the major cause of acute myocardial infarction and unstable angina, as well as also characterizing the occlusive coronary thrombus formation resulting from either thrombolytic therapy or percutaneous transluminal coronary angioplasty (PTCA) .
  • PTCA percutaneous transluminal coronary angioplasty
  • abnormal thrombosis characterizes the condition observed in patients undergoing major surgery in the lower extremities or the abdominal area who often suffer from thrombus formation in the venous vasculature resulting in reduced blood flow to the affected extremity and a predisposition to pulmonary embolism.
  • Abnormal thrombosis further characterizes disseminated intravascular coagulopathy which commonly occurs within both vascular systems during septic shock, certain viral infections and cancer, a condition wherein there is rapid consumption of coagulation factors and systemic coagulation which results in the formation of life-threatening thrombi occurring throughout the microvasculature leading to widespread organ failure.
  • the present invention includes methods for preventing a condition in a mammal suspected of having a condition characterized by abnormal thrombosis, comprising administering to said mammal a therapeutically effective amount of a compound or a pharmaceutical composition of the present invention.
  • the compounds or pharmaceutical compositions of the H ⁇ present invention are administered in vivo, ordinarily in a mammal, preferably in a human.
  • the compounds or pharmaceutical compositions can be administered to a mammal in a variety of ways, including orally, parenterally, intravenously, subcutaneously, intramuscularly, colonically, rectally, nasally or intraperitoneally, employing a variety of dosage forms.
  • Administration is preferably parenteral, such as intravenous on a daily basis.
  • administration is preferably oral, such as by tablets capsules or elixers taken on a daily basis.
  • the compounds or pharmaceutical compositions of the present invention are administered alone or in combination with one another, or in combination with other therapeutic or in vivo diagnostic agents.
  • a therapeutically effective amount of the compounds or pharmaceutical compositions of the present invention will vary depending upon the age, weight and mammalian species treated, the particular compounds employed, the particular mode of administration and the desired affects and the therapeutic indication. Because these factors and their relationship to determining this amount are well known in the medical arts, the determination of therapeutically effective dosage levels, the amount necessary to achieve the desired result of preventing thrombosis, will be within the ambit of one skilled in these arts. Typically, administration of the compounds or pharmaceutical composition of the present invention is commenced at lower dosage levels, with dosage levels being increased until the desired effect of preventing in vivo thrombosis is achieved which would define a therapeutically effective amount. For the compounds of the present invention, alone or as part of a pharmaceutical composition, such doses are between about 0.01 mg/kg and 100 mg/kg body weight, preferably between about 0.01 and 10 mg/kg, body weight.
  • N-alpha-Boc-N-delta-benzyloxycarbonyl-L-ornithine (100.3 g, 0.27 mole) was dissolved in a solution of methanol (450 mL) , water (320 mL) and acetic acid (46.5 mL) .
  • 10% palladium on carbon catalyst (10.0 g) was added and the mixture was hydrogenated on a Parr apparatus at 35 psi for 2.5 hours.
  • Thin-layer chromatography sica gel; 20:10:3 dichloromethane/methanol/acetic acid; ninhydrin
  • the solvent was removed under vacuum at 80°C.
  • the resultant oil was dissolved in 500 mL of dichloromethane and extracted with 500 mL of 1M sodium hydroxide solution.
  • the aqueous solution was extracted with 500 mL of dichloromethane, acidified with cooling with 550 mL of 1M HCl, re-extracted with 5x500 mL dichloromethane and 2x500 mL 9:1 dichloromethane/isopropanol.
  • the aqueous solution was briefly placed on the rotary evaporator to remove any volatiles.
  • the mixture was purified by reverse-phase HPLC using a standard C18 50x300 mm cartridge employing a 10-30% gradient system of acetonitrile-water (containing 0.1% trifluoroacetic acid) over 1 hour. Pooled fractions of interest were lyophilized and afforded 151 mg (63% yield) of the title compound as a colorless, amorphous solid.
  • HPLC (on C18 reverse phase) analysis revealed 3 product forms. Fast atom bombardment mass spectrometry confirmed the theoretical molecular weight of 466.
  • Example 8 The compound of Example 8 (19.1 g, 85.8 mmole) was slurried in 850 mL of dry acetonitrile and was treated with benzylsulfonyl chloride (32.7 g, 0.172 mole) . The solution was cooled to 0°C and treated dropwise with triethylamine (60.0 mL, 0.428 mole) . After 2 hours, an additional portion of benzylsulfonyl chloride (16.4 g, 85.8 mmole) was added. The solution gradually warmed to ambient temperature and was stirred for 16 hours. The solids were filtered and the filtrate was concentrated under vacuum to give an oil.
  • Example 9 The compound of Example 9 (17.2 g, 52.7 mmole) was dissolved in 350 mL of methanol, cooled to 0°C, and treated with 1.0M lithium hydroxide in water (116 mL) dropwise. After 1 hour, the reaction mixture was allowed to warm to ambient temperature and was stirred for 18 hours.
  • Dowex 50X8-400 ion-exchange resin H + form, 49 g was added to the slurry to adjust the pH to 3. After stirring for 30 minutes, the slurry was filtered and the resin was washed with several portions of water/methanol. The filtrate was concentrated under vacuum. The resulting residue was taken up in acetonitrile and concentrated under vacuum.
  • Alpha-N-t-butoxycarbonyl-N ⁇ -nitroarginine (2.00 g, 6.3 mmole) was dissolved in tetrahydrofuran (100 mL) by heating the solution to 50°C. The solution was allowed to cool to room temperature. N-methyl piperidine (0.84 mL, 6.9 mmole) was added, and the solution was cooled in an ice bath.
  • Rf 0.44 (silica gel; 5% isopropanol in dichloromethane) . Isobutanol was present as an impurity. This compound may be further purified by recrystallization from dichloromethane/hexanes or ethanol/water.
  • the title compound was made by the procedures which follows.
  • a 12 liter four-necked round bottom flask equipped with an overhead stirring apparatus was flame dried under a strong stream of nitrogen. After the flask had cooled, 120.0 g of alpha-N-t-butoxycarbonyl-N ⁇ -nitro-L-arginine (376 mmole, 1 equivalent) was added under a blanket of nitrogen followed by the addition of 6 liters of anhydrous tetrahydrofuran (Aldrich sure-seal) via canula. The flask was then fitted with a thermometer and the resulting suspension was warmed to 50°C with a heat gun while stirring. The reaction mixture was cooled to 5°C with an ice bath and further cooled to -5°C with an ice/acetone bath.
  • the crude intermediate was taken up in 300 mL of dichloromethane and applied to a column of silica gel (70 - 230 mesh, 7 x 50 cm) .
  • the column was first eluted with 2 liters of dichloromethane followed by 2 liters of 2% methanol in dichloromethane. This was followed by elution with 5% methanol in dichloromethane until all of the product had been eluted (the eluant was checked for UV activity and five one-liter fractions were collected once this UV activity was apparent) .
  • reaction When the reaction was judged to be complete, it was cooled to -70°C and 503 mL of 2M potassium bisulfate was added via dropping funnel at a slow enough rate to keep the reaction temperature below -30°C. The cooling bath was removed and the reaction mixture was allowed to come to 0°C over the course of 2 hours at which time a white precipitate was filtered off. The solids were washed with 500 mL of cold tetrahydrofuran. The filtrate was concentrated under vacuum on a rotary evaporator until most of the tetrahydrofuran was removed and the remaining white sludge was mostly aqueous.
  • the crude product was taken up in 1.5 liters of ethyl acetate and washed with 0.2 M HCl (2 x 200 mL) .
  • the HCl extracts were back-extracted with 400 mL of ethyl acetate and the organics were combined and extracted with saturated sodium bicarbonate (2 x 200 mL) .
  • the bicarbonate extracts were also back-extracted with 400 ml of ethyl acetate.
  • the organics were then combined and washed with brine (200 mL) followed by drying over anhydrous sodium sulfate.
  • the solution was filtered, concentrated under vacuum on a rotary evaporator and pumped on overnight to yield a white solid (89.0 g) of crude title compound.
  • Example 11 The compound of Example 11 (17.2 g, 52.7 mmole) was u dissolved in 215 mL of acetonitrile and was treated with the compound of Example 15 (14.2 g, 53.0 mmole) , EDC (15.2 g, 79.1mmole), and HOBt (7.12 g, 52.7 mmole) . The solution was stirred for 15 minutes at ambient temperature, cooled to 0°, and was then treated with N,N- diisopropylethyla ine (45.9 mL, 0.264 mole) . The reaction was warmed to ambient temperature and was stirred for 48 hours.
  • Example 17 The compound of Example 17 (4.5 g, 8.1 mmole) was dissolved in IN HCl (150 mL) and stirred at ambient temperature for 15 hours. 12N HCl (25.5 mL) was added to the solution and the reaction continued another 3.5 hours, Purification by reverse phase HPLC on a 50x300 mm C18 column using a gradient ranging from 10-30% of acetonitrile-water (containing 0.1% trifluoroacetic acid) over 1 hour yielded 3.8 g (81 % yield) of the title compound as a white solid. HPLC (on C18 reverse phase) analysis showed three peaks for the product. Fast atom bombardment mass spectrometry confirmed the theoretical molecular weight of 466.
  • N-Alpha-Boc-methionine (24.9 g, 0.1 mole) and glycine methyl ester hydrochloride (12.6 g, 0.1 mole) were mixed in degassed dimethylformamide (150 mL) .
  • Triethylamine (13.9 mL, 0.1 mole) and HOBt (15.3g, 0.1 mole) were dissolved in the mixture, and DCC (20.6 g, 0.1 mole) was added. The mixture was stirred at room temperature overnight and filtered. The filtrate was concentrated under vacuum, and the residue was redissolved in dichloromethane (150 mL) .
  • Example 19 The compound of Example 19 (960 mg, 3 mmole) was dissolved with stirring in 6 mL of methyl iodide at room temperature. The reaction mixture was stirred for 6.5 hours over which time a gummy solid had separated. The supernatant was drawn off and the residue was dried under vacuum to give 1.41 g (100%) of the title compound as a hygroscopic foam.
  • Example 20 The compound of Example 20 (7.3 g, 15.6 mmole) was dissolved in 312 mL of 1:1 dimethylformamide/ dichloromethane under nitrogen and cooled to 0°C. Sodium hydride (1.5 g of a 50% mineral oil suspension, 31.5 mmole) was added all at once, and the mixture was stirred at 0°C for 2.5 hours. Ethyl acetate (104 mL) followed by water (24 mL) was added, and the resultant solution was left overnight at room temperature. The solution was concentrated under vacuum to a small volume and partitioned between water (50 mL) and dichloromethane (50 mL) . The phases were separated, and the aqueous phase at pH 8 was acidified to pH 4 with 0.5M citric acid.
  • Sodium hydride 1.5 g of a 50% mineral oil suspension, 31.5 mmole
  • Example 21 The compound of Example 21 (5.16 g, 0.02 mole), the compound of Example 3 (5.83 g, 0.022 mole), HBTU (8.34 g, 0.022 mole), HOBt (2.97, 0.02 mole) was dissolved in 500 mL of acetonitrile. To this solution was added N- methylmorpholine (10 mL, 0.09 mole) slowly. The reaction was stirred for 12 hours at 25°C and. The solvent was removed under vacuum to yield a residue.
  • Example 22 The compound of Example 22 (3.3 g, 74 mmole) was dissolved in 200 mL of ethyl acetate and 50 mL of methanol. To this solution at 0°C was added 50 mL of ethyl acetate saturated with HCl gas. This mixtured was stirred for 30 minutes. The solvents were removed under vacuum to yield 3.1 g (100%) of the title compound as a white solid.
  • Example 25 The compound of Example 25 (200 mg, 0.3 mmole), EDC (0.76 g, 3.0 mmole) and 4 mL of dimethylsulfoxide were stirred at 25°C. To this solution was added dichloroacetic acid (170 microliters, 1.5 mmole) . The reaction mixture was stirred for 30 minutes. The reaction was poured into 80 mL of water, filtered and processed on an HPLC C18 50x300 mm cartridge employing a 10-40% gradient system of acetonitrile-water (containing 0.1% trifluoroacetic acid) over 20 minutes. The pure fractions were combined and lyopholized to give 130 mg of the title compound. Fast atom bombardment mass spectrometry confirmed the theoretical molecular weight of 452.
  • Example 32 To a solution of the compound of Example 30 (11.06 g, 0.0257 mole) in 200 mL ethanol, was added 10% Pd/C ( 1.11 g) ; the mixture was hydrogenated at 40 psi on the Parr Shaker for 5 hours. The catalyst was filtered off and the solvents were removed to afford 8.81 g (-quantitative yield) of product as a colorless foam, judged pure by TLC (silica gel
  • Example 31 The compound of Example 31 (6.81 g, 0.020 mole) was dissolved in 80 mL of acetonitrile. To that mixture was added the compound of Example 15 (N ⁇ -nitro-L-argininal ethyl cyclol, hydrochloride salt) (5.89 g, 0.022 mole), 1- hydroxybenzo-triazole monohydrate (3.06 g, 0.020 mole) and l-ethyl-3- (3-dimethylamino-propyl)carbodiimide hydrochloride salt (5.75 g, 0.030 mole) .
  • Example 15 N ⁇ -nitro-L-argininal ethyl cyclol, hydrochloride salt
  • N,N-diisopropylethylamine (12.83 g, 0.10 mole, 17.3 mL) .
  • the reaction mixture was stirred for 20 hours, diluted with 700 mL ethyl acetate and washed with 2x50 mL each of IN HCl, saturated sodium bicarbonate, and brine.
  • Example 32 The compound of Example 32 ( 7.54 g, 0.0136 mole) was dissolved in 200 mL of 4:1:1: ethanol/water/acetic acid and 10% Pd/C catalyst (3.8 g) was added. This mixture was hydrogenated on a Parr shaker at 50 psi for 19 hours. The catalyst was filtered off and the filtrate was evaporated. The residue was dissolved in a mixture of 1:1 ethanol/acetonitrile, reevaporated, and pumped at ⁇ 1 mm Hg on a vacuum pump for 3 days to afford 8.20 g
  • Example 33 The compound of Example 33 (2.84 g, 5.0 mmole) was dissolved in 3N HCL (80 mL) and stirred at ambient temperature for 2.5 hours. Purification via reverse phase HPLC on a 50 X 300 mm C18 column using a 10-30% gradient of acetonitrile/water (containing 0.1% trifluoroacetic acid) over 1 hour yielded 2.18 g (74% yield) of product as a colorless, amorphous solid. RP/HPLC analysis showed 13 three peaks for the product. Fast atom bombardment mass spectrometry confirmed the theoretical molecular weight of
  • Example 29 To a solution of the compound of Example 29 (9.38 g, 0.030 mole) in 300 mL dry acetonitrile is added the appropriate sulfonyl or sulfamoyl chloride listed below (0.033 mole) and the solution is cooled to 0°C under N2.
  • Example 35 the intermediates of Example 35 are used to synthesize the following compounds of the present invention (as their trifluoroacetic acid salts) :
  • Example 37 The compound of Example 37 (5.73 g, 0.020 mole) is dissolved in 80 mL of CH3CN and treated with the compound of Example 15 (N°-nitro-L-argininal ethyl cyclol, hydrochloride salt) (5.89 g, 0.022 mole), 1- Hydroxybenzotriazole monohydrate (3.06 g, 0.020 mole) and l-ethyl-3- (3-dimethylamino-propyl)carbodiimide hydrochloride salt (5.75 g, 0.030 mole) .
  • Example 15 N°-nitro-L-argininal ethyl cyclol, hydrochloride salt
  • 1- Hydroxybenzotriazole monohydrate (3.06 g, 0.020 mole)
  • l-ethyl-3- (3-dimethylamino-propyl)carbodiimide hydrochloride salt 5.75 g, 0.030 mole
  • Example 38 The compound of Example 38 (6.79 g, 0.0136 mole) is dissolved in 200 mL of a mixture of EtOH, H2O, HOAc: 4, 1,
  • Example 39 The compound of Example 39 (2.57 g, 5.0 mmole) is dissolved in 3N HCL (80 mL) and stirred at ambient temperature for 2.5 hours. Purification via reverse phase HPLC on a 50 X 300 mm C18 column using a 0-20% gradient of acetonitrile/water/0.1% TFA over 1 hour yields the title compound. RP/HPLC analysis shows three peaks for the product.
  • Example 42 The compound of Example 42 (6.01 g, 0.020 mole) is dissolved in 80 mL of CH3CN and treated with the compound of Example 15 (N°-nitro-L-argininal ethyl cyclol, hydrochloride salt) (5.89 g, 0.022 mole), 1- Hydroxybenzotriazole monohydrate (3.06 g, 0.020 mole) and l-ethyl-3- (3-dimethylamino-propyl)carbodiimide hydrochloride salt (5.75 g, 0.030 mole) .
  • Example 15 N°-nitro-L-argininal ethyl cyclol, hydrochloride salt
  • 1- Hydroxybenzotriazole monohydrate (3.06 g, 0.020 mole)
  • l-ethyl-3- (3-dimethylamino-propyl)carbodiimide hydrochloride salt 5.75 g, 0.030 mole
  • Example 43 The compound of Example 43 (6.99 g, 0.0136 mole) is dissolved in 200 mL of EtOH, H2O, HOAc: 4, 1, 1. Pd/C catalyst (3.5 g of 10%) is added and the mixture is hydrogenated on a Parr shaker at 50 PSI for 26 hours. The catalyst is filtered off and the filtrate evaporated. The residue is dissolved in a mixture of EtOH, CH3CN: 1,1; reevaporated, and pumped at ⁇ 1 mm Hg on a vacuum pump for 3 days to afford -Quantitative crude yield of product which is judged pure by TLC (silica gel; CH2CI2, MeOH, NH4OH: 20, 10, 2) .
  • Example 44 The compound of Example 44 (2.64 g, 5.0 mmole) is dissolved in 3N HCL (80 mL) and stirred at ambient temperature for 2.5 hours . Purification via reverse phase
  • Compound 21 was made according to the protocol of Example 1.
  • N,N-diisopropylethylamine (727.4 mg, 5.64 mmole, 0.98 mL) was added and the resultant clear light brown solution was stirred at room temperature for 3 days.
  • the reaction was diluted with 100 mL of ethyl acetate, extracted with 25 mL portions of 0.1 N HCl (2x) , saturated NaHC ⁇ 3 (2x) , water, and brine; then, it was dried over MgS04, filtered and evaporated.
  • N- (Benzyloxycarbonyloxy) -succinimide (12.46 g, 0.050 mole) rapidly over 2 minutes. The solution was stirred, allowing the temperature to increase from 0°C to ambient temperature over 14 hours and the THF was evaporated. Solid NaCl was added to the remaining aqueous layer and it was extracted with lx 200 mL and 2 x 100 mL of ethyl acetate. The combined organic layers were extracted with 50 mL portions of water (2x) and brine, then dried over MgS04, filtered, and evaporated.
  • N ⁇ -nitro-L-argininal ethyl cyclol (683.0 mg, 2.55 mmole), 1-hydroxybenzotriazole hydrate (325.6 mg, 2.126 mmole) and l-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (611.0 mg, 3.19 mmole) .
  • N,N-diisopropylethylamine (1.099 g, 8.50 mmole, 1.48 mL) was added and the resultant clear light brown solution was stirred at room temperature for 18 hours.
  • the reaction was diluted with 200 mL of ethyl acetate, extracted with 25 mL portions of 0.1N HCl (2x) , saturated NaHC ⁇ 3 (2x) , water and brine, then dried over MgS04, filtered and evaporated.
  • Examples 65 through 72 (a) describe synthesis of N- [ (S) -3-N-phenylethylamino-2-oxo-l-piperidineacetyl] -L- argininal, bis-trifluoroacetate salt (47) .
  • the synthesis 9 ⁇ ° scheme is shown in Figure 12.
  • the reaction mixture was stirred at room temperature for 15 hours and quenched with 30 mL of water.
  • the mixture was diluted with 50 mL of CH2CI2 and the organic phase was separated, washed with 30 mL portions of saturated NaHC ⁇ 3, water and brine; it was dried over MgS ⁇ 4, filtered, and evaporated.
  • the residue was purified by flash chromatography on silica gel eluting with CH2CI2, ethanol:
  • N ⁇ -nitro-L-argininal ethyl cyclol 712.0 mg, 2.66 mmole
  • 1-hydroxybenzotriazole hydrate 340.0 mg, 2.22 mmole
  • l-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride 638.0 mg, 3.33 mmole
  • N,N-diisopropylethylamine (1.15 g, 8.88 mmole, 1.55 mL) was added and the resultant clear light brown solution was stirred at room temperature for 18 hours.
  • the solvent was evaporated and the residue was dissolved in 150 mL of ethyl acetate, extracted with 25 mL portions of IN HCl (2x) , saturated NaHC ⁇ 3 (2x) , water, and brine; it was dried over MgS04, filtered and evaporated.
  • Examples 73 through 80 describe synthesis of N-[(S)- 3-N-Phenylethylamino-hexahydro-2-oxo-azepine-l-acetyl] -L- argininal, bis-trifluoroacetate salt (56) .
  • the synthesis scheme is shown in Figure 13.
  • N, N-diisopropylethylamine (1.99 g, 15.4 mmole, 2.68 mL) was added and the resultant clear light brown solution was stirred at room temperature for 22 hours.
  • the solvent was evaporated and the residue was dissolved in 300 mL of ethyl acetate, extracted with 25 mL portions of IN HCl (2x) , saturated NaHC ⁇ 3 (2x) , and brine, dried over NaS ⁇ 4, filtered and evaporated.
  • Example 35 (0.033 mole) and the solution is cooled to 0°C under 2.
  • a solution of triethylamine (6.68g, 0.066 mole,
  • Example 35 Using this method and the starting materials listed in Example 35, intermediates having the formula given below, with R defined as in Example 35, are made:
  • Example 81 the intermediates of Example 81 were used to synthesize the following compounds of the present invention (as their trifluoroacetic acid salts) :
  • Example 83 the intermediates of Example 83 are used to synthesize the following compounds of the present invention (as their trifluoroacetate salts) :
  • Purified human alpha- thrombin (3000 U/mg specific activity) was obtained from Enzyme Research Laboratories, Inc.
  • the buffer used for all assays was HBSA (10 mM HEPES, pH 7.5, 150 mM sodium chloride, 0.1% bovine serum albumin) .
  • the assay for Ki determinations was conducted by combining in appropriate wells of a Corning microtiter plate, 50 microliters of HBSA, 50 microliters of the test compound at a specified concentration diluted in HBSA (or HBSA alone for V 0 (uninhibited velocity) measurement), and 50 microliters of the chromogenic substrate (250 mM, 5X Km) At time zero, 50 microliters of alpha-thrombin diluted in HBSA, was added to the wells yielding a final concentration of 0.5 nM in a total volume of 200 microliters.
  • Velocities of chromogenic substrate hydrolysis which occurred over 40 minutes was measured by the change in absorbance at 405 nm using a Thermo Max® Kinetic Microplate Reader. Ki values were determined for test compounds using the relationships developed by Williams and Morrison, Methods in Enzymology, j53_: 437 (1979) using steady state velocities (Vs) measured over 40 minutes. The extent of substrate hydrolysis was less than 5% over the course of this assay.
  • Table 1 gives the Ki values for BzlS ⁇ 2 ⁇ norVal (cyclo) -Gly-Arg-al and N- (BnS02-norLeu(cyclo) -Gly) - L-argininal. The data shows the utility of these compounds as potent in vitro inhibitors of human alpha- thrombin.
  • thrombin catalytic activity was assessed by determining the concentration of test compound which inhibited the activity of this enzyme by 50%, (IC50) - and comparing this value to that determined for some or all of the following related serine proteases: recombinant tissue plasminogen activator (rt-PA) , plasmin, activated protein C (aPC) , chymotrypsin and trypsin.
  • rt-PA tissue plasminogen activator
  • aPC activated protein C
  • chymotrypsin chymotrypsin
  • trypsin trypsin.
  • the buffer used for all assays was HBSA (10 mM HEPES, pH 7.5, 150 mM sodium chloride, 0.1% bovine serum albumin) .
  • the assay for IC50 determinations was conducted by combining in appropriate wells of a Corning microtiter plate, 50 microliters of HBSA, 50 microliters of the test compound at a specified concentration (covering a broad concentration range) diluted in HBSA (or HBSA alone for V 0
  • Enzyme activity was determined using the chromogenic substrate, Pefachrome t-PA (CH3S ⁇ 2-D-hexahydrotyrosine- /// glycyl-L-Arginine-p-nitroaniline, obtained from Pentapharm
  • the substrate was reconstituted in deionized water prior to use.
  • Purified human ⁇ -thrombin was obtained from
  • HBSA HBSA (10 mM HEPES, pH 7.5, 150 mM sodium chloride, 0.1% bovine serum albumin) .
  • IC50 determinations were conducted where HBSA (50 ⁇ L) , ⁇ -thrombin (50 ⁇ l) and inhibitor (50 ⁇ l) (covering a broad concentration range) , were combined in appropriate wells and incubated for 30 minutes at room temperature prior to the addition of substrate Pefachrome-t-PA (50 ⁇ l) .
  • the initial velocity of Pefachrome t-PA hydrolysis was measured by the change in absorbance at 405nm using a Thermo Max® Kinetic Microplate Reader over a 5 minute period in which less than 5% of the added substrate was utilized.
  • the concentration of added inhibitor which caused a 50% decrease in the initial rate of hydrolysis was defined as the IC50 value.
  • rt-PA tissue plasminogen activator
  • rt-PA catalytic activity was determined using the substrate, Pefachrome t-PA (CH3SO2-D-hexahydrotyrosine- glycyl-L-arginine-p-nitroanilide, obtained from Pentapharm Ltd.) .
  • the substrate was made up in deionized water followed by dilution in HBSA prior to the assay in which the final concentration was 500 micromolar (about 3-times Km) .
  • Human rt-PA (Activase®) was obtained from Genentech Inc.
  • the enzyme was reconstituted in deionized water and diluted into HBSA prior to the assay in which the final concentration was 1.0 nM.
  • Plasmin catalytic activity was determined using the chromogenic substrate, S-2251 (D-valyl-L-leucyl-L-lysine- p- nitroanilide) , which was obtained from Kabi
  • the substrate was made up in deionized water followed by dilution in HBSA prior to the assay in which the final concentration was 300 micromolar (about 2.5- II J. times Km) .
  • Purified human plasmin was obtained from
  • the enzyme was diluted into HBSA prior to assay in which the final concentration was 1.0 nM.
  • Activated Protein C aPC catalytic activity was determined using the chromogenic substrate, Pefachrome PC (delta-carbobenzloxy- D-lysine-L-prolyl-L-arginine-p-nitroanilide) , obtained from Pentapharm Ltd.) .
  • the substrate was made up in deionized water followed by dilution in HBSA prior to the assay in which the final concentration was 250 micromolar (about 3-times Km) .
  • Purified human aPC was obtained from Hematologic Technologies, Inc. The enzyme was diluted into HBSA prior to assay in which the final concentration was 1.0 nM.
  • Chymotrypsin catalytic activity was determined using the chromogenic substrate, S-2586 (methoxy-succinyl-L- arginine-L-prolyl-L-tyrosyl-p-nitroanilide) , which was obtained from Kabi Diagnostica.
  • the substrate was made up in deionized water followed by dilution in HBSA prior to the assay in which the final concentration was 100 micromolar (about 9-times Km) .
  • Purified (3X- crystallized,*CDI) bovine pancreatic a-chymotrypsin was obtained from Worthington Biochemical Corp. The enzyme was reconstituted in deionized water and diluted into HBSA prior to assay in which the final concentration was 1.0 nM.
  • Trypsin catalytic activity was determined using the chromogenic substrate, S-2222 (benzoyl-L-isoleucine-L- glutamic acid- (gamma-methyl ester) -L-arginine-p- nitroanilide) , which was obtained from Kabi Diagnostica.
  • the substrate was made up in deionized water followed by dilution in HBSA prior to the assay in which the final ⁇ 3 concentration was 250 micromolar (about 4-times Km) .
  • Purified (3X-crystallized; TRL3) bovine pancreatic trypsin was obtained from Worthington Biochemical Corp. The enzyme was reconstituted in deionized water and diluted into HBSA prior to assay in which the final concentration was 0.5 nM.
  • Factor Xa catalytic activity was determined using the chromogenic substrate S-2765 (N-benzyloxycarbonyl-D- arginine-L-glycine-L-arginine-p-nitroaniline) , obtained from Kabi Pharmacia Hepar, Inc. (Franklin, OH) . All substrates were reconstituted in deionized water prior to use. The final concentration of S-2765 was 250 ⁇ M (about 5-times Km) . Purified human Factor X was obtained from Enzyme Research Laboratories, Inc.
  • Table 2 lists the determined IC50 values for the enzymes listed above and demonstrates the high degree of specificity for the inhibition of alpha-thrombin compared to these related serine proteases.
  • Table 3 displays the specificity of compounds of this invention, having variable Ri and X groups and ring size, with respect to thrombin, fXa and trypsin. The data demonstrate the thrombin specificity of the compounds.
  • Fresh frozen citrated pooled normal human plasma was obtained from George King Biomedical, Overland Park, KA. Pooled normal rat plasma was prepared from citrated whole blood collected from anesthetized rats using standard procedures. The plasma was flash frozen and stored at - 80°C until use. Measurements APTT was made using the Coag-A-Mate RA4 automated coagulometer (General Diagnostics, Organon Technica, Oklahoma City, OK) using the Automated APTT reagent (Organon Technica, Durham, NC) as the initiator of clotting according to the manufacturers instructions. The assay was conducted by making a series of dilutions of the test compounds in rapidly thawed plasma followed by adding 200 microliters to the wells of the assay carousel.
  • Example 34 were evaluated using the following established experimental model of acute vascular thrombosis.
  • mice Male Harlan Sprague Dawley rats (420-450 g) were acclimated at least 72 hours prior to use and fasted for 12 hours prior to surgery with free access to water. The animals were prepared, anesthetized with Nembutal followed by the insertion of catheters for blood pressure monitoring, drug and anesthesia delivery.
  • the left carotid artery was isolated by making a midline cervical incision followed by blunt dissection and spreading techniques to separate a 2 cm segment of the vessel from If? the carotid sheath. A silk suture is inserted under the proximal and distal ends of the isolated vessel to provide clearance for the placement of a ultrasonic flow probe
  • test compound BzlS ⁇ 2 -norVal(cyclo)-Gly-Arg-al or N-(BnS ⁇ 2 ⁇ norLeu(cyclo)-Gly)-L-argininal
  • the test compound was administered as a single intravenous bolus at the doses outlined in Table 3 after placement of the flow probe and 5 minutes prior to the thrombogenic stimulus.
  • the effective dose which prevents 50% of thrombotic occlusions in this model can be determined from the above data by plotting the incidence of occlusion versus the dose administered. This allows a direct comparison of the antithrombotic efficacy of BzlS ⁇ 2-norVal (cyclo) -Gly- Arg-al and N- (BnS ⁇ 2-norLeu(cyclo) -Gly)-L-argininal with other antithrombotic agents which have also been evaluated in this model as described above.
  • Table 6 lists the ED50 values for several well known anticoagulant agents in this 1/9 model compared to the present compounds
  • a ED50 is defined as the dose that prevents the incidence of complete thrombotic occlusion in 50% of animals tested.
  • the pharmacodynamic and pharmacokinetic parameters of the compound of Example 34 were evaluated after oral administration in dogs.
  • Adult beagle dogs (9-12 kg) were housed individually in standard cages, and were fed a standard certified commercial diet and tap water ad libi tum . Dogs were given an oral dose of 20 mg/kg test compound, in 55 ml deionized water, via a gastric tube, followed by a water rinse.
  • 5 mg/kg of test compound, in 5 ml 0.9% sodium chloride were administered by fast bolus i.v. injection through an indwelling saphenous vein catheter.
  • the same dogs were used for both routes of administration, with a one week wash out period between use of the dogs for the different protocols.
  • Blood samples were collected during the experimental period from the cephalic vein using sodium citrate (0.38% final concentration) as anticoagulant (nine volumes blood to one volume sodium citrate) . After mixing with the anticoagulant, each blood sample was immediately chilled by immersion in a slurry of ice and water. The plasma was separated by centrifugation (4°C, 2400 rpm, 10 minutes) within 10 minutes of collection, and then transferred to a cryotube and stored at -70°C until analysis. The blood levels of test compound were measured by extracting the compound from plasma, separation by HPLC, and post-column derivatization with a fluorogenic functional group.
  • the clotting time of blood was measured using an APTT assay, which measures the prolongation of the activated partial thromboplastin time (APTT) by the test compound.
  • Fresh frozen pooled normal citrated human plasma was obtained from George King Biomedical, Overland Park, KS. Measurements of APTT were made using the Coag- A-Mate RA4 automated coagulometer (General Diagnostics, Organon Technica, Oklahoma City, OK) using the Automated aPTT Platelin® L reagent (Organon Technica, Durham, NC) as an initiator of clotting according to the manufacturers instructions .
  • the assay was conducted by making a series of dilutions of each blood sample in rapidly thawed plasma followed by adding 200 microliters APTT reagent to the wells of the assay carousel.

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  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Peptides Or Proteins (AREA)

Abstract

La présente invention concerne des aldéhydes de peptides qui sont des inhibiteurs puissants et spécifiques de la thrombine, leurs sels pharmaceutiquement acceptables, et des compositions pharmaceutiquement acceptables de ces derniers. L'invention a également pour objet des procédés pour les utiliser comme agents thérapeutiques pour soigner des maladies chez le mammifère, caractérisées par une thrombose anormale.
PCT/US1995/007661 1994-06-17 1995-06-19 Derives de l'acide 3-amino-2-oxo-1-piperidineacetique WO1995035311A1 (fr)

Priority Applications (7)

Application Number Priority Date Filing Date Title
BR9508048A BR9508048A (pt) 1994-06-17 1995-06-19 Derivados 3-amino 2-oxo 1-piperidina acéticos como inibidores de enzimas
MX9606566A MX9606566A (es) 1994-06-17 1995-06-19 Derivados 3-amina-2-0xo-1-piperidin aceticos como inhibidores de enzima.
JP8502499A JPH10503176A (ja) 1994-06-17 1995-06-19 酵素インヒビターとしての３−アミノ−２−オキソ−１−ピペリジン酢酸誘導体
EP95923922A EP0802923A1 (fr) 1994-06-17 1995-06-19 Derives de l'acide 3-amino-2-oxo-1-piperidineacetique
AU28630/95A AU700808B2 (en) 1994-06-17 1995-06-19 3-amino-2-oxo-1-piperidineacetic derivatives as enzyme inhibitors
NZ288940A NZ288940A (en) 1994-06-17 1995-06-19 4-amino-2-oxo-1-piperidineacetic acid tripeptide derivatives as thrombin inhibitors
NO965353A NO965353L (no) 1994-06-17 1996-12-13 3-amino-2-okso-1-piperidineddiksyrederivater som enzyminhibitorer

Applications Claiming Priority (5)

Application Number Priority Date Filing Date Title
US26137894A 1994-06-17 1994-06-17
US08/261,378 1994-06-17
US35683194A 1994-12-13 1994-12-13
US08/356,831 1994-12-13
US08/487,007 1995-06-07

Publications (2)

Publication Number Publication Date
WO1995035311A1 true WO1995035311A1 (fr) 1995-12-28
WO1995035311A9 WO1995035311A9 (fr) 1996-02-01

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JP (1) JPH10503176A (fr)
WO (1) WO1995035311A1 (fr)

Cited By (42)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997030073A1 (fr) * 1996-02-13 1997-08-21 Akzo Nobel N.V. Inhibiteurs de la serine protease
WO1997031937A1 (fr) * 1996-03-01 1997-09-04 Akzo Nobel N.V. Inhibiteur de la serine-protease
WO1997031939A1 (fr) * 1996-03-01 1997-09-04 Akzo Nobel N.V. Inhibiteurs de la serine-protease
WO1998009987A1 (fr) * 1996-09-06 1998-03-12 Biochem Pharma, Inc. Inhibiteurs lactame de la thrombine
WO1998016547A1 (fr) * 1996-10-11 1998-04-23 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
WO1998016524A1 (fr) * 1996-10-11 1998-04-23 Cor Therapeutics, Inc. DERIVES HETEROCYCLIQUES UTILISES COMME INHIBITEURS DE FACTEUR Xa
WO1998016525A1 (fr) * 1996-10-11 1998-04-23 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
WO1999007732A1 (fr) * 1997-08-11 1999-02-18 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
WO1999007731A1 (fr) * 1997-08-11 1999-02-18 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
US6011047A (en) * 1997-11-26 2000-01-04 Corvas International Inc. Substituted 3-amino-2-hydroxyphenylacetamide derivatives as enzyme inhibitors
US6063794A (en) * 1996-10-11 2000-05-16 Cor Therapeutics Inc. Selective factor Xa inhibitors
US6133256A (en) * 1997-04-14 2000-10-17 Cor Therapeutics Inc Selective factor Xa inhibitors
US6194435B1 (en) 1996-10-11 2001-02-27 Cor Therapeutics, Inc. Lactams as selective factor Xa inhibitors
US6204384B1 (en) 1997-11-26 2001-03-20 Corvas International, Inc. Substituted 3-amino-2-hydroxyphenylacetamide derivatives as enzyme inhibitors (II)
US6204268B1 (en) 1997-04-14 2001-03-20 Cor Therapeutics, Inc Selective factor Xa inhibitors
WO2001019795A1 (fr) * 1999-09-13 2001-03-22 3-Dimensional Pharmaceuticals, Inc. Inhibiteurs de serine protease azacycloalcanone
US6211183B1 (en) 1997-04-14 2001-04-03 Cor Therapeutics, Inc. Selective factor Xa inhibitors
EP0944386A4 (fr) * 1996-12-13 2001-04-11 Aventis Pharm Prod Inc Composes a base d'acide sulfonique ou de sulfonylamino n-(heteroaralkyl)-azaheterocyclylamide
US6218382B1 (en) 1997-08-11 2001-04-17 Cor Therapeutics, Inc Selective factor Xa inhibitors
US6228854B1 (en) 1997-08-11 2001-05-08 Cor Therapeutics, Inc. Selective factor Xa inhibitors
US6262047B1 (en) 1996-10-11 2001-07-17 Cor Therapeutics, Inc. Selective factor Xa inhibitors
US6297233B1 (en) 1999-02-09 2001-10-02 Bristol-Myers Squibb Company Lactam inhibitors of FXa and method
WO2001079261A1 (fr) * 2000-04-14 2001-10-25 Corvas International, Inc. Derives de tetrahydro-azepinone servant d'inhibiteurs de la thrombine
US6333321B1 (en) 1997-08-11 2001-12-25 Cor Therapeutics, Inc. Selective factor Xa inhibitors
US6344450B1 (en) 1999-02-09 2002-02-05 Bristol-Myers Squibb Company Lactam compounds and their use as inhibitors of serine proteases and method
US6369080B2 (en) 1996-10-11 2002-04-09 Cor Therapeutics, Inc. Selective factor Xa inhibitors
US6369063B1 (en) 1997-04-14 2002-04-09 Cor Therapeutics, Inc. Selective factor Xa inhibitors
EP1175405A4 (fr) * 1999-02-09 2002-05-15 Bristol Myers Squibb Co INHIBITEURS LACTAME DU FACTEUR Xa ET METHODE ASSOCIEE
AU753842B2 (en) * 1997-08-11 2002-10-31 Millennium Pharmaceuticals, Inc. Selective factor Xa inhibitors
US6511973B2 (en) 2000-08-02 2003-01-28 Bristol-Myers Squibb Co. Lactam inhibitors of FXa and method
US6555542B1 (en) 2001-01-30 2003-04-29 Bristol-Myers Squibb Company Sulfonamide lactam inhibitors of FXa and method
US6777431B2 (en) 2001-07-13 2004-08-17 Corvas International, Inc. Non-convalent thrombin inhibitors
US7179835B2 (en) 2001-11-16 2007-02-20 Glaxo Group Limited 2-(3-sulfonylamino-2-oxopyrrolidin-1-yl)propanamides as factor xa inhibitors
US7279477B2 (en) 2001-01-26 2007-10-09 Chugai Seiyaku Kabushiki Kaisha Malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US7285562B2 (en) 2003-08-01 2007-10-23 Chugai Seiyaku Kabushiki Kaisha Cyanoamide compounds useful as malonyl-CoA decarboxylase inhibitors
US7385063B2 (en) 2001-01-26 2008-06-10 Chugai Seiyaku Kabushiki Kaisha Method for preparing imidazole derivatives
US7449482B2 (en) 2003-08-01 2008-11-11 Chugai Seiyaku Kabushiki Kaisha Piperidine compounds useful as malonyl-CoA decarboxylase inhibitors
US7524969B2 (en) 2001-01-26 2009-04-28 Chugai Seiyaku Kabushiki Kaisha Malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US7696365B2 (en) 2003-08-01 2010-04-13 Chugai Seiyaku Kabushiki Kaisha Heterocyclic compounds useful as malonyl-CoA decarboxylase inhibitors
US7709510B2 (en) 2001-02-20 2010-05-04 Chugai Seiyaku Kabushiki Kaisha Azoles as malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US7723366B2 (en) 2001-02-20 2010-05-25 Chugai Seiyaku Kabushiki Kaisha Azole compounds as malonyl-CoA decarboxylase inhibitors for treating metabolic diseases
US7786145B2 (en) 2003-08-01 2010-08-31 Chugai Seiyaku Kabushiki Kaisha Cyanoguanidine-based azole compounds useful as malonyl-CoA decarboxylase inhibitors

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
FR2490632A1 (fr) * 1980-09-19 1982-03-26 Nippon Kayaku Kk Derives de l-argininal et procede de preparation
EP0526877A2 (fr) * 1991-08-06 1993-02-10 Bristol-Myers Squibb Company Aldéhydes peptidiques ayant une activité anti-thrombotique

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
FR2490632A1 (fr) * 1980-09-19 1982-03-26 Nippon Kayaku Kk Derives de l-argininal et procede de preparation
EP0526877A2 (fr) * 1991-08-06 1993-02-10 Bristol-Myers Squibb Company Aldéhydes peptidiques ayant une activité anti-thrombotique

Cited By (53)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1997030073A1 (fr) * 1996-02-13 1997-08-21 Akzo Nobel N.V. Inhibiteurs de la serine protease
US6034067A (en) * 1996-02-13 2000-03-07 Akzo Nobel, N.V. Serine protease inhibitors
US6410684B1 (en) 1996-03-01 2002-06-25 Akzo Nobel N.V. Serine protease inhibitors
WO1997031937A1 (fr) * 1996-03-01 1997-09-04 Akzo Nobel N.V. Inhibiteur de la serine-protease
WO1997031939A1 (fr) * 1996-03-01 1997-09-04 Akzo Nobel N.V. Inhibiteurs de la serine-protease
US6218365B1 (en) * 1996-03-01 2001-04-17 Akzo Nobel N.V. Serine protease inhibitors
WO1998009987A1 (fr) * 1996-09-06 1998-03-12 Biochem Pharma, Inc. Inhibiteurs lactame de la thrombine
WO1998016524A1 (fr) * 1996-10-11 1998-04-23 Cor Therapeutics, Inc. DERIVES HETEROCYCLIQUES UTILISES COMME INHIBITEURS DE FACTEUR Xa
US6525076B1 (en) 1996-10-11 2003-02-25 Millennium Pharmaceuticals, Inc. Selective factor Xa inhibitors
WO1998016525A1 (fr) * 1996-10-11 1998-04-23 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
US6063794A (en) * 1996-10-11 2000-05-16 Cor Therapeutics Inc. Selective factor Xa inhibitors
US6194435B1 (en) 1996-10-11 2001-02-27 Cor Therapeutics, Inc. Lactams as selective factor Xa inhibitors
US6369080B2 (en) 1996-10-11 2002-04-09 Cor Therapeutics, Inc. Selective factor Xa inhibitors
WO1998016547A1 (fr) * 1996-10-11 1998-04-23 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
US6262047B1 (en) 1996-10-11 2001-07-17 Cor Therapeutics, Inc. Selective factor Xa inhibitors
EP0944386A4 (fr) * 1996-12-13 2001-04-11 Aventis Pharm Prod Inc Composes a base d'acide sulfonique ou de sulfonylamino n-(heteroaralkyl)-azaheterocyclylamide
US6369063B1 (en) 1997-04-14 2002-04-09 Cor Therapeutics, Inc. Selective factor Xa inhibitors
US6133256A (en) * 1997-04-14 2000-10-17 Cor Therapeutics Inc Selective factor Xa inhibitors
US6204268B1 (en) 1997-04-14 2001-03-20 Cor Therapeutics, Inc Selective factor Xa inhibitors
US6211183B1 (en) 1997-04-14 2001-04-03 Cor Therapeutics, Inc. Selective factor Xa inhibitors
US6333321B1 (en) 1997-08-11 2001-12-25 Cor Therapeutics, Inc. Selective factor Xa inhibitors
AU753842B2 (en) * 1997-08-11 2002-10-31 Millennium Pharmaceuticals, Inc. Selective factor Xa inhibitors
US6218382B1 (en) 1997-08-11 2001-04-17 Cor Therapeutics, Inc Selective factor Xa inhibitors
US6228854B1 (en) 1997-08-11 2001-05-08 Cor Therapeutics, Inc. Selective factor Xa inhibitors
WO1999007732A1 (fr) * 1997-08-11 1999-02-18 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
WO1999007731A1 (fr) * 1997-08-11 1999-02-18 Cor Therapeutics, Inc. INHIBITEURS SELECTIFS DU FACTEUR Xa
US6204384B1 (en) 1997-11-26 2001-03-20 Corvas International, Inc. Substituted 3-amino-2-hydroxyphenylacetamide derivatives as enzyme inhibitors (II)
US6011047A (en) * 1997-11-26 2000-01-04 Corvas International Inc. Substituted 3-amino-2-hydroxyphenylacetamide derivatives as enzyme inhibitors
US6297233B1 (en) 1999-02-09 2001-10-02 Bristol-Myers Squibb Company Lactam inhibitors of FXa and method
US6344450B1 (en) 1999-02-09 2002-02-05 Bristol-Myers Squibb Company Lactam compounds and their use as inhibitors of serine proteases and method
EP1175405A4 (fr) * 1999-02-09 2002-05-15 Bristol Myers Squibb Co INHIBITEURS LACTAME DU FACTEUR Xa ET METHODE ASSOCIEE
US6962942B2 (en) 1999-09-13 2005-11-08 3-Dimensional Pharmaceuticals, Inc. Azacycloalkanone serine protease inhibitors
WO2001019795A1 (fr) * 1999-09-13 2001-03-22 3-Dimensional Pharmaceuticals, Inc. Inhibiteurs de serine protease azacycloalcanone
US6469029B1 (en) 1999-09-13 2002-10-22 3-Dimensional Pharmaceuticals, Inc. Azacycloalkanone serine protease inhibitors
WO2001079261A1 (fr) * 2000-04-14 2001-10-25 Corvas International, Inc. Derives de tetrahydro-azepinone servant d'inhibiteurs de la thrombine
US6511973B2 (en) 2000-08-02 2003-01-28 Bristol-Myers Squibb Co. Lactam inhibitors of FXa and method
US7385063B2 (en) 2001-01-26 2008-06-10 Chugai Seiyaku Kabushiki Kaisha Method for preparing imidazole derivatives
US7279477B2 (en) 2001-01-26 2007-10-09 Chugai Seiyaku Kabushiki Kaisha Malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US8119819B2 (en) 2001-01-26 2012-02-21 Chugai Seiyaku Kabushiki Kaisha Malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US7524969B2 (en) 2001-01-26 2009-04-28 Chugai Seiyaku Kabushiki Kaisha Malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US7166586B2 (en) 2001-01-30 2007-01-23 Bristol Myers Squibb Co. Sulfonamide lactam inhibitors of FXa and method
US6555542B1 (en) 2001-01-30 2003-04-29 Bristol-Myers Squibb Company Sulfonamide lactam inhibitors of FXa and method
US7723366B2 (en) 2001-02-20 2010-05-25 Chugai Seiyaku Kabushiki Kaisha Azole compounds as malonyl-CoA decarboxylase inhibitors for treating metabolic diseases
US8110686B2 (en) 2001-02-20 2012-02-07 Chugai Seiyaki Kabushiki Kaisha Azoles as malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US7709510B2 (en) 2001-02-20 2010-05-04 Chugai Seiyaku Kabushiki Kaisha Azoles as malonyl-CoA decarboxylase inhibitors useful as metabolic modulators
US6777431B2 (en) 2001-07-13 2004-08-17 Corvas International, Inc. Non-convalent thrombin inhibitors
US7179835B2 (en) 2001-11-16 2007-02-20 Glaxo Group Limited 2-(3-sulfonylamino-2-oxopyrrolidin-1-yl)propanamides as factor xa inhibitors
US7696365B2 (en) 2003-08-01 2010-04-13 Chugai Seiyaku Kabushiki Kaisha Heterocyclic compounds useful as malonyl-CoA decarboxylase inhibitors
US7786145B2 (en) 2003-08-01 2010-08-31 Chugai Seiyaku Kabushiki Kaisha Cyanoguanidine-based azole compounds useful as malonyl-CoA decarboxylase inhibitors
US7897615B2 (en) 2003-08-01 2011-03-01 Chugai Sciyaku Kabushiki Kaisha Cyanoamide compounds useful as malonyl-CoA decarboxylase inhibitors
US8080665B2 (en) 2003-08-01 2011-12-20 Chugai Seiyaku Kabushiki Kaisha Piperidine compounds useful as malonyl-CoA decarboxylase inhibitors
US7449482B2 (en) 2003-08-01 2008-11-11 Chugai Seiyaku Kabushiki Kaisha Piperidine compounds useful as malonyl-CoA decarboxylase inhibitors
US7285562B2 (en) 2003-08-01 2007-10-23 Chugai Seiyaku Kabushiki Kaisha Cyanoamide compounds useful as malonyl-CoA decarboxylase inhibitors

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