Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • C07K14/745—Blood coagulation or fibrinolysis factors
  • C07K14/75—Fibrinogen
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides

Definitions

• This invention relates to processes for the synthesis of platelet glycoprotein Ilb/IIIa inhibitors, and to intermediate compounds useful in said processes.
• Activation of platelets and the resulting platelet aggregation and secretion of factors by the platelets have been associated with different pathophysiological conditions including cardiovascular and cerebrovascular thromboembolic disorders, for example, the thromboembolic disorders associated with unstable angina, myocardial infarction, transient ischemic attack, stroke, atherosclerosis and diabetes.
• cardiovascular and cerebrovascular thromboembolic disorders for example, the thromboembolic disorders associated with unstable angina, myocardial infarction, transient ischemic attack, stroke, atherosclerosis and diabetes.
• the contribution of platelets to these disease processes stems from their ability to form aggregates, or platelet thrombi, especially in the arterial wall following injury.
• Platelets are known to play an essential role in the maintenance of hemostasis and in the pathogenesis of arterial thrombosis. Platelet activation has been shown to be enhanced during coronary thrombolysis which can lead to delayed reperfusion and reocclusion. Clinical studies with aspirin, ticlopidine and a monoclonal antibody for platelet glycoprotein Ilb/IIIa provide biochemical evidence for platelet involvement in unstable angina, early stage of acute myocardial infarction, transient ischemic attack, cerebral ischemia, and stroke.
• Platelets are activated by a wide variety of agonists resulting in platelet shape change, secretion of granular contents and aggregation. Aggregation of platelets serves to further focus clot formation by concentrating activated clotting factors in one site.
• endogenous agonists including adenosine diphosphate (ADP) , serotonin, arachidonic acid, thrombin, and collagen, have been identified. Because of the involvement of several endogenous agonists in activating platelet function and aggregation, an inhibitor which acts against all agonists would represent a more efficacious antiplatelet agent than currently available antiplatelet drugs, which are agonist-specific.
• Current antiplatelet drugs are effective against only one type of agonist; these include aspirin, which acts against arachidonic acid; ticlopidine, which acts against ADP; thromboxane A2 synthetase inhibitors or receptor antagonists, which act against thromboxane A2; and hirudin, which acts against thrombin.
• GPIIb/IIIa platelet glycoprotein Ilb/IIIa complex
• GPIIb/IIIa membrane protein mediating platelet aggregation.
• a recent review of GPIIb/IIIa is provided by Phillips et al. (1991) Cell 65: 359-362.
• the development of a GPIIb/IIIa antagonist represents a promising new approach for antiplatelet therapy.
• Recent studies in man with a monoclonal antibody for GPIIb/IIIa indicate the antithrombotic benefit of a GPIIb/IIIa antagonist.
• GPIIb/IIIa-specific antiplatelet agent which inhibits the activation and aggregation of platelets in response to any agonist.
• Such an agent should represent a more efficacious antiplatelet therapy than the currently available agonist-specific platelet inhibitors.
• GPIIb/IIIa does not bind soluble proteins on unstimulated platelets, but GPIIb/IIIa in activated platelets is known to bind four soluble adhesive proteins, namely fibrinogen, von illebrand factor, fibronectin, and vitronectin.
• fibrinogen The binding of fibrinogen and von Willebrand factor to GPIIb/IIIa causes platelets to aggregate.
• the binding of fibrinogen is mediated in part by the Arg-Gly-Asp (RGD) recognition sequence which is common to the adhesive proteins that bind GPIIb/IIIa.
• This invention is directed to a processes for the preparation of compounds of formula:
• Z is a suitable carboxylic acid protecting group and Y is a suitable amine protecting group, with a carboxylic acid derivitive of formula:
• G is a suitable amine protecting group, to produce a protected linear peptide of formula:
• Step (f) reacting the product of Step (e) with a reagent capable of converting an amine to guanidine to produce a compound of formula (I) ,
• p and p' are 0 or 1;
• ⁇ - 9 is a C 6 -C ⁇ 4 saturated, partially saturated, or aromatic carbocyclic ring system or heterocyclic ring system composed of at least 1-3 heteroatoms selected from N, 0, S; all these ring may be optionally substituted with 0-2 R 7 ;
• R 17 and R 16 ' are independently selected from 5 the group:
• R--5 and R-*- 8 are independently selected from 15 the group:
• heterocylic ring system composed of 5-10 atoms including 1-3 nitrogen, 30 oxygen, or sulfur heteroatoms, optionally substituted with 0-2 R 13 ;
• R 15 and R 17 can alternatively join to form a 5-7 membered carbocyclic ring 35 substituted with 0-2 R 13 ;
• R 18 and R 16 can alternatively join to form a 5-7 membered carbocyclic ring substituted with 0-2 R 13 ;
• R 15 and R 14 can alternatively join to form a 5-8 membered carbocyclic ring substituted with 0-2 R 13 , when R 17 is H;
• R 7 is independently selected at each occurrence from the group:
• R 8 is independently selected at each occurrence from the group:
• R 20 is independently selected at each occurrence from the group:
• R 21 is independently selected at each occurrence from the group:
• R 11 is H or C ⁇ -C 8 alkyl
• R 12 is H or C1-C8 alkyl
• R 14 is H or Ci-C ⁇ alkyl
• R 3 is H or Ci-C ⁇ alkyl
• A is selected from the group:
• R 9 is H, C1-C8 alkyl
• R 5 is H, C ⁇ C8 alkyl.
• R-*-- ⁇ is selected from:
• R 15 and R 18 are independently selected from H, C1-C4 alkyl, phenyl, benzyl, phenyl- (C2-C4)alkyl, C 1 -C 4 alkoxy;
• R 17 and R 16 are independently H or C 1 -C 4 alkyl;
• R 7 is H, Ci-C ⁇ alkyl, phenyl, halogen, or C 1 -C 4 alkoxy;
• R 11 is H or C 1 -C 3 alkyl
• R 12 is H or CH3
• R 9 is H, C 1 -C 3 alkyl ;
• R 5 is H, C 1 -C 3 alkyl.
• R 5 , R 9 , R 16 , R 17 and R 18 are H;
• R 11 , R 12 , and R 14 are H or CH3;
• R 15 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl- (C2-C4) alkyl;
• R 3 is H or C 1 -C 3 alkyl.
• R 5 , R 9 , R 17 , R 15 ,R n , R 12 , R 14 are H;
• R 2 is C2H5;
• R 3 is CH3; and
• A is -(CH2)3".
• R 1 is wherein:
• p and p 1 are 0 or 1; 5
• R 19 is a C ⁇ -Ci 4 saturated, partially saturated, or aromatic carbocyclic ring system or heterocyclic ring system composed of at least 1-3 10 heteroatoms selected from N, 0, S; all these ring systems may be optionally substituted with 0-2 R 7 ;
• R 17 and R 16 are independently selected from 15 the group:
• C 1 -C 4 alkyl optionally substituted with halogen; 20 C1-C2 alkoxy; benzyl;
• R- • - • -*- and R 18 are independently selected from 25 the group:
• heterocylic ring system composed of 5-10 atoms including 1-3 nitrogen, oxygen, or sulfur heteroatoms, optionally substituted with 0-2 R 13 ;
• R 15 and R 17 can alternatively join to form a 5-7 membered carbocyclic ring substituted with 0-2 R 13 ;
• R 18 and R 16 can alternatively join to form a
• R 15 and R 14 can alternatively join to form a 20 5-8 membered carbocyclic ring substituted with 0-2 R 13 , when R 17 is
• R 7 is independently selected at each 25 occurrence from the group:
• R 8 is independently selected at each occurrence from the group:
• R* 1 - 3 is independently selected at each occurrence from the group:
• R 20 is independently selected at each occurrence from the group:
• R 2 -*- is independently selected at each occurrence from the group:
• R 11 is H or Ci-C ⁇ alkyl
• R 12 is H or C1-C8 alkyl
• R 14 is H or Ci-C ⁇ alkyl
• R 3 is H or Ci-C ⁇ alkyl
• R 3 and A may also be taken together to form
• n 0-1;
• R 9 is H, C1-C8 alkyl
• R 5 is H, C1-C8 alkyl.
• R-*- 9 is selected from:
• R 15 and R 18 are independently selected from H, C1-C4 alkyl, phenyl, benzyl, phenyl- (C2-C 4 )alkyl, C 1 -C 4 alkoxy;
• R 17 and R 16 are independently H or C 1 -C 4 alkyl
• R 7 is H, Ci-C ⁇ alkyl, phenyl, halogen, or C1-C4 alkoxy;
• R 11 is H or C 1 -C 3 alkyl
• R 12 is H or CH3
• A is C1-C7 alkyl.
• R 3 and A may be taken together to form
• n 0-1 and Y is two hydrogen atoms
• R 9 is H, C1-C3 alkyl
• R 5 is H, C1-C3 alkyl
• R 11 , R 12 , and R 14 are H or CH3;
• R 15 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(C2-C4) alkyl;
• R 3 is H or C 1 -C 3 alkyl.
• R l9 is ⁇ ;
• R5, R9, R-*- 7 , R15.R11, R1 2 , R14 are H;
• R 2 is C2H5;
• R 3 is CH3; and A is - ( CH2 ) 3-.
• This invention also provides intermediate compounds useful in the claimed processes for the preparation of compounds of formula (I) .
• Said intermediate compounds have formulae:
• Z is H, t-butyl, benzyl, alkyl, t-BOC;
• G is H, t-BOC, CBZ;
• p and p' are 0 or 1;
• R 19 is a C6-C ⁇ 4 saturated, partially saturated, or aromatic carbocyclic ring system or heterocyclic ring system composed of at least 1-3 heteroatoms selected from N, 0, S; all these ring systems may be optionally substituted with 0-2 R 7 ;
• R 17 and R 16 are independently selected from 5 the group:
• R* * - * *-* 1 and R**- 8 are independently selected from 15 the group:
• heterocylic ring system composed of 5-10 atoms including 1-3 nitrogen, 30 oxygen, or sulfur heteroatoms, optionally substituted with 0-2 R 13 ;
• R 15 and R 17 can alternatively join to form a 5-7 membered carbocyclic ring 35 substituted with 0-2 R 13 ;
• R 18 and R 16 can alternatively join to form a 5-7 membered carbocyclic ring substituted with 0-2 R 13 ;
• R 15 and R 14 can alternatively join to form a 5-8 membered carbocyclic ring substituted with 0-2 R 13 , when R 17 is H;
• R 7 is independently selected at each occurrence from the group:
• R 8 is independently selected at each occurrence from the group:
• R 20 is independently selected at each occurrence from the group:
• R 21 is independently selected at each occurrence from the group:
• R 11 is H or Ci-C ⁇ alkyl
• R 12 is H or Ci-C ⁇ alkyl
• R 14 is H or Ci-C ⁇ alkyl
• R 3 is H or Ci-C ⁇ alkyl
• A is selected from the group:
• R 3 and A may also be taken together to form
• R 9 is H, Ci-C ⁇ alkyl
• R 5 is H, C ⁇ -C8 alkyl
• R 2* -** 1 is H or benzyl.
• Preferred intermediate compounds of formulae III and IV are those wherein:
• Z is H, t-butyl
• G is H, t-BOC
• R**- 9 is selected from:
• R 15 and R 18 are independently selected from H, C1-C4 alkyl, phenyl, benzyl, phenyl- (C2-C4) alkyl, C 1 -C 4 alkoxy;
• R 17 and R 16 are independently H or C 1 -C 4 alkyl
• R 7 is H, Ci-C ⁇ alkyl, phenyl, halogen, or C 1 -C 4 alkoxy;
• R 11 is H or C 1 -C3 alkyl
• R 12 is H or CH3
• A is C1-C7 alkyl.
• R 3 and A may be taken together to form
• R 9 is H, C 1 -C 3 alkyl
• R 5 is H, C 1 -C 3 alkyl.
• R 5 , R 9 , R 16 , R 17 and R 18 are H;
• R 11 , R 12 , and R 14 are H or CH3;
• R 15 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl- (C2-C4)alkyl;
• R 3 is H or C 1 -C 3 alkyl.
• Specifically preferred compounds of formulae II and IV are those wherein:
• R 5 , R 9 , R 17 , R 15 ,R n , R 12 , R 14 are H;
• R 2 is C2H5;
• R 3 is CH3; and
• A is - ( CH2 ) 3-.
• any variable for example, R 1 through R 8 , m, n, p, X, Y, etc.
• its definition on each occurrence is independent of its definition at every other occurrence.
• combinations of substituents and/or variables are permissible only if such combinations result in stable compounds.
• alkyl is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms; “alkoxy” represents an alkyl group of indicated number of carbon atoms attached through an oxygen bridge; “cycloalkyl” is intended to include saturated ring groups, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl; and “biycloalkyl” is intended to include saturated bicyclic ring groups such as [3.3.0]bicyclooctane, [4.3.0]bicyclononane, [4.4.0]bicyclodecane (decalin) , [2.2.2]bicyclooctane, and so forth.
• Alkenyl is intended to include hydrocarbon chains of either a straight or branched configuration and one or more unsaturated carbon- carbon bonds which may occur in any stable point along the chain, such as ethenyl, propenyl and the like; and "alkynyl” is intended to include hydrocarbon chains of either a straight or branched configuration and one or more triple carbon-carbon bonds which may occur in any stable point along the chain, such as ethynyl, propynyl and the like.
• Halo or “halogen” as used herein refers to fluoro, chloro, bromo and iodo; and "counterion” is used to represent a small, negatively charged species such as chloride, bromide, hydroxide, acetate, sulfate and the like.
• aryl is intended to mean phenyl or naphthyl;
• Carbocyclic is intended to mean any stable 5- to 7- membered monocyclic or bicyclic or 7- to 14-membered bicyclic or tricyclic carbon ring, any of which may be saturated, partially unsaturated, or aromatic.
• carbocyles examples include, but are not limited to cyclopentyl, cyclohexyl, phenyl, biphenyl, naphthyl, indanyl or tetrahydronaphthyl (tetralin) .
• heterocycle or “heterocyclic ring system” is intended to mean a stable 5- to 7- membered monocyclic or bicyclic or 7- to 10-membered bicyclic heterocyclic ring which may be saturated, partially unsaturated, or aromatic, and which consists of carbon atoms and from 1 to 3 heteroatoms selected from the group consisting of N, 0 and S and wherein the nitrogen and sulfur heteroatoms may optionally be oxidized, and the nitrogen may optionally be quaternized, and including any bicyclic group in which any of the above-defined heterocyclic rings is fused to a benzene ring.
• the heterocyclic ring may be attached to its pendant group at any heteroatom or carbon atom which results in a stable structure.
• the heterocyclic rings described herein may be substituted on carbon or on a nitrogen atom if the resulting compound is stable.
• Examples of such heterocycles include, but are not limited to, pyridyl, pyrimidinyl, furanyl, thienyl, pyrolyl, pyrazolyl, imidazolyl, tetrazolyl, benzofuranyl, benzothiophenyl, indolyl, indolenyl, quinolinyl, isoquinolinyl or benzimidazolyl, piperidinyl, 4-piperidonyl, pyrrolidinyl, 2-pyrrolidonyl, pyroli ' nyl, tetrahydrofuranyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, deca
• stable compound or “stable structure” is meant herein a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and formulation into an efficacious therapeutic agent .
• substituted means that one or more hydrogen on the designated atom is replaced with a selection from the indicated group, provided that the designated atom's normal valency is not exceeded, and that the substitution results in a stable compound.
• amine protecting group means any group known in the art of organic synthesis for the protection of amine groups. Such amine protecting groups include those listed in Greene, “Protective Groups in Organic Synthesis” John Wiley & Sons, New York (1981) and “The Peptides: Analysis, Sythesis, Biology, Vol. 3, Academic Press, New York (1981), the disclosure of which is hereby incorporated by reference. Any amine protecting group known in the art can be used.
• amine protecting groups include, but are not limited to, the following: 1) acyl types such as formyl, trifluoroacetyl, phthalyl, and p-toluenesulfonyl; 2) aromatic carbamate types such as benzyloxycarbonyl (Cbz) and substituted benzyloxycarbonyls, 1- (p- biphenyl) -1-methylethoxycarbonyl, and 9-fluorenylmethyloxycarbonyl (Fmoc) ; 3) aliphatic carbamate types such as tert-butyloxycarbonyl (Boc) , ethoxycarbonyl, diisopropylmethoxycarbonyl, and allyloxycarbonyl; 4) cyclic alkyl carbamate types such as cyclopentyloxycarbonyl and adamantyloxycarbonyl; 5) alkyl types such as triphenylmethyl and benzyl; 6) trialkylsi
• pharmaceutically acceptable salts and prodrugs refer to derivatives of the disclosed compounds that are modified by making acid or base salts, or by modifying functional groups present in the compounds in such a way that the modifications are cleaved, either in routine manipulation or in vivo, to the parent compounds.
• Examples include, but are not limited to: mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; esters of carboxylates; acetate, formate and benzoate derivatives of alcohols and amines; and the like.
• compositions of the compounds of the invention can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in Remington's Pharmaceutical Sciences. 17th ed.. Mack
• amino acid as used herein means an organic compound containing both a basic amino group and an acidic carboxyl group. Included within this term are modified and unusual amino acids,such as those disclosed in, for example, Roberts and Vellaccio (1983) The Peptides. 5: 342-429, the teaching of which is hereby incorporated by reference.
• amino acid residue means that portion of an amino acid (as defined herein) that is present in a peptide or pseudopeptide.
• peptide as used herein means a linear compound that consists of two or more amino acids (as defined herein) that are linked by means of peptide or pseudopeptide bonds.
• Phg phenylglycine
• Trp tryptophan
• Val valine
• the present invention provides a process for the synthesis of compounds of formula (I) .
• the provided process is accomplished using inexpensive, simple starting materials.
• the overall process is novel: it utilizes novel reaction steps, novel reaction sequences, and novel reaction intermediates.
• knowledge of a number of standard techniques known to those in the art is required. The following discussion and references are offered to provide such knowledge.
• peptides are elongated by deprotecting the ⁇ -amine of the C-terminal residue and coupling the next suitably protected amino acid through a peptide linkage using the methods described. This deprotection and coupling procedure is repeated until the desired sequence is obtained.
• This coupling can be performed with the constituent amino acids in a stepwise fashion, or condensation of fragments (two to several amino acids) , or combination of both processes, according to the method originally described by Merrifield, J. Am. Chem. Soc, 85, 2149- 2154 (1963), "The Peptides: Analysis, Synthesis, Biology, Vol. 1, 2, 3, 5, and 9, Academic Press, New York, (1980-1987); Bodanszky, "Peptide Chemistry: A
• the coupling of two amino acid derivatives, an amino acid and a peptide, two peptide fragments, or the cyclization of a peptide can be carried out using standard coupling procedures such as the azide method, mixed carbonic acid anhydride (isobutyl chloroformate) method, carbodiimide (dicyclohexylcarbodiimide, diisopropylcarbodiimide, or water-soluble carbodiimides) method, active ester (p-nitrophenyl ester, N-hydroxysuccinic imido ester) method.
• the functional groups of the constituent amino acids must be protected during the coupling reactions to avoid undesired bond formation.
• the protecting groups that can be used, methods of using them to protect amino acids, and methods to remove them are listed in Greene, "Protective Groups in Organic Synthesis” John Wiley & Sons, New York (1981) and "The Peptides: Analysis, Sythesis, Biology, Vol. 3, Academic Press, New York (1981), the disclosure of which is hereby incorporated by reference.
• the ⁇ -carboxyl group of the C-terminal residue is usually protected by an ester that can be cleaved to give the carboxylic acid.
• These protecting groups include: 1) alkyl esters such as methyl and t-butyl, 2) aryl esters such as benzyl and substituted benzyl, or 3) esters which can be cleaved by mild base treatment or mild reductive means such as trichloroethyl and phenacyl esters.
• the C-terminal amino acid is attached to an insoluble carrier (usually polystyrene) .
• insoluble carriers contain a group which will react with the carboxyl group to form a bond which is stable to the elongation conditions but readily cleaved later.
• examples of which are: oxime resin (DeGrado and Kaiser (1980) J. Org. Chem . 45, 1295-1300) chloro or bromomethyl resin, hydroxymethyl resin, and aminomethyl resin.
• oxime resin DeGrado and Kaiser (1980) J. Org. Chem . 45, 1295-1300
• chloro or bromomethyl resin hydroxymethyl resin
• aminomethyl resin aminomethyl resin.
• Many of these resins are commercially available with the desired C-terminal amino acid already incorporated.
• acyl types such as formyl, trifluoroacetyl, phthalyl, and p- toluenesulfonyl
• aromatic carbamate types such as benzyloxycarbonyl (Cbz) and substituted benzyloxycarbonyls, 1- (p-biphenyl)-1- methylethoxycarbonyl, and 9-fluorenylmethyloxycarbonyl (Fmoc)
• 3) aliphatic carbamate types such as tert- butyloxycarbonyl (Boc) , ethoxycarbonyl, diisopropylmethoxycarbonyl, and allyloxycarbonyl
• cyclic alkyl carbamate types such as cyclopentyloxycarbonyl and adamantyloxycarbonyl
• 5) alkyl types such as triphen
• the ⁇ -amino protecting group is cleaved prior to the coupling of the next amino acid.
• the reagents of choice are hydrogenation conditions using hydrogen at atmospheric pressure or in a Parr apparatus at elevated hydrogen pressure, or cyclohexene or ammonium formate over palladium, palladium hydroxide on charcoal or platinum oxide in methanol, ethanol or tetrahydrofuran, or combination of these solvents (P. N. Rylander, Hydrogenation Methods, Acedemic Press, 1985) .
• the Boc group is used, the methods of choice are trifluoroacetic acid, neat or in dichloromethane, or HC1 in dioxane.
• the resulting ammonium salt is then neutralized either prior to the coupling or in situ with basic solutions such as aqueous buffers, or tertiary amines in dichloromethane or dimethylformamide.
• the reagents of choice are piperidine or substituted piperidines in dimethylformamide, but any secondary amine or aqueous basic solutions can be used.
• the deprotection is carried out at a temperature between 0°C and room temperature.
• any of the amino acids bearing side chain functionalities must be protected during the preparation of the peptide using any of the above- identified groups.
• Those skilled in the art will appreciate that the selection and use of appropriate protecting groups for these side chain functionalities will depend upon the amino acid and presence of other protecting groups in the peptide. The selection of such a protecting group is important in that it must not be removed during the deprotection and coupling of the ⁇ -amino group.
• the following protecting groups are acceptable: p-toluenesulfonyl (tosyl) moieties for arginine; t-butyloxycarbonyl, phthalyl, or tosyl for lysine or ornithine; alkyl esters such as cyclopentyl for glutamic and aspartic acids; alkyl ethers for serine and threonine; and the indole of tryptophan can either be left unprotected or protected with a formyl group.
• p-toluenesulfonyl (tosyl) moieties for arginine t-butyloxycarbonyl, phthalyl, or tosyl for lysine or ornithine
• alkyl esters such as cyclopentyl for glutamic and aspartic acids
• alkyl ethers for serine and threonine
• the indole of tryptophan can either
• Boc is chosen for the ⁇ -amine protection the following protecting groups are acceptable: p- toluenesulfonyl (tosyl) moieties and nitro for arginine; benzyloxycarbonyl, substituted benzyloxycarbonyls, or tosyl for lysine; benzyl or alkyl esters such as cyclopentyl for glutamic and aspartic acids; benzyl ethers for serine and threonine; benzyl ethers, substituted benzyl ethers or 2-bromobenzyloxycarbonyl for tyrosine; p-methylbenzyl, p-methoxybenzyl, acetamidomethyl, benzyl, or t- butylsulfonyl for cysteine; and the indole of tryptophan can either be left unprotected or protected with a formyl group.
• tert-butyl based protecting groups are acceptable.
• Boc can be used for lysine, tert-butyl ether for serine, threonine and tyrosine, and tert-butyl ester for glutamic and aspartic acids.
• Unusual amino acids used in this invention can be synthesized by standard methods familiar to those skilled in the art ("The Peptides: Analysis, Sythesis, Biology, Vol. 5, pp. 342-449, Academic Press, New York (1981)) .
• N-Alkyl amino acids can be- prepared using proceedures described in previously (Cheung et al., (1977) Can . J. Chem . 55, 906; Freidinger et all, (1982) J. Org. Chem . 48, 77 (1982)), which are incorporated here by reference.
• Step 1 of the process a compound of formula (1) is reacted with an appropriate protecting group to give protected amine (2) .
• the compound of formula (2) wherein Y is an amine protecting group, such as t-Boc, acyl, phthalyl, or other suitable group previously described, can be prepared from an appropriately substituted ⁇ -amino acid by complexing the ⁇ -amine and carboxylic acid with a copper salt, in water, an alcohol, or dioxane, or combination of these solvents, and protecting the remote amine with a protecting group such as t-butyloxycarbonyl, acyl, phthalyl or another previously described protecting group.
• the protected compound (3) is prepared by reacting amine (2) with any of the previously described amine protecting groups precursors.
• the preferred protecting group is Cbz.
• reaction of amine compound (2) and benzyl ch oroformate in 1,4-dioxane and water with sodium hydroxide as the acid scavenger at ambient temperature affords compound (3) wherein X is Cbz. This is shown in Scheme 1.
• the oxazolidinone compound (4) is prepared, as shown in Scheme 1, by the condensation of an appropriately substituted aldehyde, such as formaldehyde, acetaldehyde, benzaldehyde, C3- C8 alkyl and branched alkyl aldehydes or aldehyde surrogates such as trioxane, dimethoxymethane or higher alkyl acetals, in a solvent like benzene, toluene, N,N-dimethylformamide, or dioxane with an acid catalyst such as p-toluenesulfonic acid, hydrochloric acid, or trifluoroacetic acid at a temperature between 50°-150°C with a dean stark trap, molecular sieves, magnesium sulfate or other drying agent.
• an appropriately substituted aldehyde such as formaldehyde, acetaldehyde, benzaldehyde, C3- C8 al
• N- ⁇ -alkyl compound (5) is prepared by reduction of oxazolidinone (4) with triethylsilane and an acid such as trifluoroacetic acid in solvents such as methylene chloride or chloroform between -25° and 60°C.
• the preferred procedure for the preparation of compound (5) wherein X is Cbz, R 22 is H and Y is phthalyl, from the corresponding compound (4) utilizes triethylsilane with trifluoroacetic acid in chloroform at ambient temperature to reflux temperature of the solvent.
• Step 5 the dipeptide (6) is prepared by coupling amino acid (5) with an appropriately substituted carboxy protected amino acid, (AA) .
• Step 5 utilizes any of the several amide bond forming reactions described previously.
• Step 5 for the preparation of the compound of formula (6) wherein Z is t-butyl alkyl and X is Cbz is via reaction of the corresponding carboxylic acid (5) with glycine t-butyl ester, in the presence of the water soluble carbodiimde, l-(3- dimthethylaminopropyl) -3-ethylcarbodiimide hydrocloride, in the solvent methylene chloride, with N-methylmorpholine as the acid scavenger, at ambient temperature; or via reaction with isobutylchloroformate and N-methylmorpholine in tetrahydrofuran at -30° to 0° temperature.
• Step 6 the N- ⁇ -alkyl dipeptide (7) is prepared by deprotection of the corresponding compound of formula (6) using the appropriate conditions for removal of the selected protecting group, as shown in Scheme 2.
• X is Cbz
• the preferred method for the preparation of compound (7) wherein Z is t-butyl alkyl involves hydrogenation of compound (6) wherein X is Cbz with 10% palladium on charcoal and ammonium formate, in an alcohol solvent, at a temperature between ambient temperature and 70°. Alternatively, the reaction may be carried out with 10% palladium on charcoal at elevated hydrogen pressure, in an alcohol solvent.
• Step 7 of the process involves preparation of tripeptide (10) by coupling of an appropriately substituted N- ⁇ protected amino acid compound of formula (22) with the N- ⁇ -alkyl dipeptide compound (7) .
• Compounds of formula (22) are commercially available (Sigma, BACHEM) . Step 7 may be accomplished using any of the previously described methods for forming amide bonds.
• Step 7 the preferred method to prepare compound (10) wherein X is Cbz, R 3 is alkyl, and Z is t-butyl alkyl, is to activate the N- ⁇ -Cbz protected amino acid of the corresponding compound of formula (22) with O-benzotriazol-1-yl- N,N,N' ,N'-tetramethylurion hexafluorophosphate, or a carbodiimide and 1-hydroxybenzotriazole in the presence of a compound of formula (7) wherein R 3 is alkyl Z is t-butyl alkyl, and Y is a protecting group, in methylene chloride, at ambient temperature.
• compound (11) is prepared by hydrogenation of the N- ⁇ -Cbz-protected tripeptide,
• the preferred reduction methods for preparation of a compound of formula (11) wherein Z is t-butyl alkyl, from compound (10) wherein X is Cbz include: 10% palladium on charcoal and ammonium formate or cyclohexene, in an alcohol solvent, over a temperature range between ambient temperature and 70°C, or 10% palladium on charcoal at elvated hydrogen pressure, in an alcohol solvent.
• Step 9 the fully elaborated protected linear peptide compound, (15) , is prepared by coupling the carboxylic acid compound, (14) , and the amino tripeptide compound, (11) .
• This step may be carried out using any of the previously described methods for forming amide bonds.
• the preferred coupling method for the preparation of the linear peptide of formula (15) wherein Z is t-butyl alkyl and G is t-Boc, from the amino compound (11) wherein Z is t-butyl alkyl and the carboxylic acid compound of formula (14) wherein G is t-Boc, utilizes l-(3- dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride or O-benzotriazol-1-yl-N,N,N' ,N'-tetra ethylurion hexafluorophosphate carbonyldiimiazole, hydroxysuccinate ester or isobutylchloroformate and 1- hydroxybentriazole with N-methylmorpholine in N,N- dimethylformamide or acetonitrile at ambient temperature.
• Step 10 the free amino acid peptide compound, (16) is prepared by the deprotection of compound (15) .
• deprotection of (15) wherein Z is t-butyl alkyl and G is t-Boc may be accomplished using any of the variety of methods well known in the literature for the deprotection of t- butyl esters and t-Boc groups. Such methods include: hydrogen chloride in dioxane or ethyl acetate; and trifluoroacetic acid neat or in methylene chloride, chloroform, ether, or toluene.
• the preferred method to prepare the free amino acid compound, (16) by deprotection of compound (15) wherein G is t-Boc and Y is t-butyl alkyl, utilizes trifluoroacetic acid in methylene chloride or hydrogen chloride in dioxane, at ambient temperature.
• the cyclic compound, (17) is prepared by cyclization of the linear pentapeptide compound, (16) .
• This Step may be accomplished using any of the variety of amide bond forming reactions well known in the literature, as previously described.
• the cyclization step may be carried out using macrocyclization techniques.
• the preferred cyclization methods for the preparation of compounds of formula (17) from the linear compound, (16) utilizes a carbodiimide such as l-(3- dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, dicyclohexylcarbodiimide, or 0- benzotriazol-1-yl-N,N, ', '-tetramethylurion hexafluorophosphate and 1-hydroxybentriazole with N- methylmorpholine, in a solvent such as N,N- dimethylformamide or acetonitrile, at ambient temperature.
• a carbodiimide such as l-(3- dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, dicyclohexylcarbodiimide, or 0- benzotriazol-1-yl-N,N, ', '-tetramethylurion hexafluorophosphate and 1-hydroxy
• Step 12 the ⁇ carboxylic acid
• the amino compound of formula (19) is prepared by removal of the protecting group, Y, of (18) via methods appropriate for removing the selected protecting group, Y.
• the preferred methods for preparation of the amino compound (19) from the protected compound (18) wherein Y is phthalyl utilize hydrazine, or. methyl amine or other lower alkyl " amines in solvents such as water, methanol, ethanol or combination of these solvents, at a temperature between ambient temperature and 80°.
• solvents such as water, methanol, ethanol or combination of these solvents
• the compound of formula (I) is prepared.
• the preparation is effected by reacting the amine compound, (19) , with any of the variety of reagents capable of converting an amine to a guanidine compound or a protected guanidine.
• Suitable reagents include: S-methyl and O-methyl isoureas, guanyl-3,5-dimethylpyrazole, cyanamide, formamidine sulfonic acid, bis-carbamate protected s- alkyl isoureas, "The Peptide” vol 2, 169-175, Garigipat et al. Tetrahedron Letters 31, 1969 (1990), Kim et al.
• the preferred method for the conversion of the amino compound (19) to the guanidine compound (I) utilizes formamidine sulfonic acid and N,N-dimethylaminopyridine, in solvents such as water, methanol, ethanol, dioxane or combination of these solvents at ambient temperature to reflux temperature of the solvent.
• n 0,1
• the preparation of intermediate compound (14) is shown in Scheme 5.
• the pseudodipeptide (14) is prepared by coupling the amino carboxylic acid compound of formula (13) or formula (13A) , with the activated carboxylic acid of an appropriately substituted N- ⁇ protected amino acid of formula (21) wherein G is a protecting group such as t-Boc, using any of the amide bond forming reactions previously described.
• the preferred method for preparing the carboxylic acid compound, (14), wherein R-*- is phenyl is by reaction of the free amino acid compound, (13) wherein R-*- is phenyl, with a ' carboxylic acid, (21) , activated with N,N'-carbonyldiimidazole, in the solvent N,N-dimethylformamide, at ambient temperature.
• the carboxylic acid can be activated as the N-hyroxysuccinate ester in a solvent such as methylene chloride or N,N-dimethylformamide.
• the amino carboxylic acid compound of formula (13) or formula (13A) can be purchased or can be prepared by reduction of the appropriately substituted cyano carboxylic acid compound (12) by methods well known in the literature for reducing cyano groups, as described in Tett . Lett . , 4393 (1975); Modern Synthetic Reactions, H.O. House (1972); or Harting et al. J. Am . Chem . Soc , 50: 3370 (1928) .
• the preferred method for preparing the amino acid (13) , wherein R-*- is phenyl from (12) involves reductive hydrogenation at elevated hydrogen pressure, with 10% palladium on charcoal in an alcohol solvent like ethanol between ambient temperature and 60°C. For example, reduction of 3- or 4-cyanobenzoic acid, which is a compound of formula (12) wherein R 1 is phenyl, under these conditions affords the corresponding benzyl amine of formula (13) .
• N-alkylated compound of formula (23) can be prepared according to standard procedures, for example, Olsen, J. Org. Chem . (1970) 35: 1912) . This compound may also be prepared as shown in Scheme 6. e
• Schemes 7-10 show a number of routes to intermediate compounds of formula (24) .
• Compound (24) falls within general formula (13) and is useful for the synthesis of compounds of formula (14) .
• Scheme 7 details a method for the preparation of compounds of formula (24) wherein R 23 is CH3, CH2CH3, CH2CH2CH3, CH2CH2CH2CH3, CH(CH3)2, C(CH3)3, CH(CH 3 )CH2CH3, benzyl, cyclopentyl, or cyclohexyl.
• Formula (25) which is another compound of general formula (13) and is useful as an intermediate in the synthesis of the compounds of the invention is prepared using standard procedures, for example, as described in Collman and Groh (1982) J. Am . Chem . Soc , 104: 1391, and as shown below:
• the 4, 5, and 6-Substituted 3-aminomethylbenzoic acid'HCl of formula (26) can be prepared using standard procedures, for example, as described in Felder et al Helv. Chim . Acta, 48: 259 (1965); de Diesbach Helv. Chim . Acta, 23: 1232 (1949); Truitt and Creagn J. Org. Chem . , 27: 1066 (1962); or Sekiya et al Chem . Phar . Bull . , 11: 551 (1963), and as shown below. Such compounds fall within general formula (13) and are useful for the preparation of compounds of formula (14) as shown in Scheme 3.
• carbocylic residues for R 1 of the invention include aminoalkyl-naphthoic acid.
• Formula (29) and aminoalkyl-tetrahydronaphthoic acid.
• Formula (30) The synthesis of these intermediates is outlined below in Scheme 11:
• R-*- Formula (I) Some other possible analogues for R-*- Formula (I) can be prepared according to a modification of standard procedures previously reported in the literature (Earnest, I., Kalvoda, J., Rihs, G., and Mutter, M., Tett. Lett., Vol. 31, No. 28, pp 4011- 4014, 1990) .
• An alternative process for the preparation of compounds of formula (I) involves introduction of the guanidine residue at an earlier stage.
• a synthon bearing a guanidine residue such as compound (9)
• a synthon bearing a protected guanidine residue such as compound (9A)
• Scheme 12 shows methods for the preparation of compounds of formula (9) and (9A) .
• the amine compound Formula (8) can be prepared from the protected amine compound (5) wherein Y is phthalyl using methods previously discussed for the removal of a phthalyl group.
• the preferred method involves the use of hydrazine or methylamine, in a solvent such as ethanol, at a temperature between ambient temperature to reflux temperature of the solvent.
• the compound of formula (9) can be prepared from the amino compound (8) , using any of the variety of methods previously described for converting an amine to a guanidine.
• compound Formula (9A) can be prepared from the amino compound Formula (8),or compound Formula 9 using any of the variety of methods in the literature that describe preparation of protected guanidines.
• the preferred methods for the preparation of the protected quanidine compound of formula (9A) are di-t-Boc S-methyl isothiourea or di-CBZ S-methyl isothiourea (Delle Monche, et al, EPO 0330629A2) in methanol or ethanol at tempertatures between ambient and reflux temperature of the solvent.
• t-Butyloxycarbonyl (Boc) amino acids and other starting amino acids may be obtained commercially from Bachem Inc., Bachem Biosciences Inc. (Philadelphia, PA) , Advanced ChemTech (Louisville, KY) , Peninsula Laboratories (Belmont, CA) , or Sigma (St. Louis, MO) .
• 2-(lH-Benzotriazol-l-yl)-l,l,3,3- tetramethyluronium hexafluorophosphate (HBTU) and TBTU were purchased from Advanced ChemTech.
• N-methylmorpholine (NMM) , m-cresol, D-2-aminobutyric acid (Abu) , trimethylacetylchloride, diisopropylethylamine (DIEA) , 3-cyanobenzoic acid and [2-(tert-butyloxycarbonyloxylimino)- phenylacetonitrile] (Boc-ON) were purchased from Aldrich Chemical Company.
• Dimethylformamide (DMF) , ethyl acetate, chloroform (CHCI3) , methanol (MeOH) , pyridine and hydrochloric acid (HCI) were obtained from Baker.
• Acetonitrile, dichloromethane (DCM) , acetic acid (HOAc) , trifluoroacetic acid (TFA) , ethyl ether, triethylamine, acetone, and magnesium sulfate were purchased from EM Science. Palladium on carbon catalyst (10% Pd) was purchased from Aldrich Chemical Company or Fluka Chemical Company. Absolute ethanol was obtained from Quantum Chemical Corporation. Thin layer chromatography (TLC) was performed on Silica Gel 60 F254 TLC plates (layer thickness 0.2 mm) which were purchased from EM Separations. TLC visualization was accomplished using UV light, iodine, and/or ninhydrin spray.
• the amine hydrochloride (0.40 g, 2 mmol) was dissolved in 15 ml water.
• a solution of BOC-ON (0.52 g, 2.1 mmol) in 15 ml acetone was added, followed by the addition of triethylamine (0.8 ml, 6 mmol) .
• reaction was allowed to proceed for 20 h.
• the reaction mixture was concentrated and partitioned between ethyl acetate and water. Aqueous layer was acidified to pH 2 using 10% HCI solution. Product was extracted in ethyl acetate, which after the usual work up and recrystallization from ethyl acetate/hexane, gave the title compound as a white solid (0.30 g ; 57% yield) . m.p. 116-118° C.
• Part E A mixture of 4-carboxyphenylbutyric acid (10.40 g, 0.05 mol), aluminum chloride (33.34 g, 0.25 mol) and sodium chloride (2.90 g, 0.05 mol) was heated with continual stirring to 190°C over 30 minutes. As the mixture cooled to 60°C, cold hydrochloric acid (IN, 250 mL) was carefully added. The mixture was extracted with dichloromethane. The combined organic layers were backwashed with dilute hydrochloric acid and water, dried over anhydrous magnesium sulfate and evaporated to dryness under reduced pressure.
• 4-carboxyphenylbutyric acid 10.40 g, 0.05 mol
• aluminum chloride 33.34 g, 0.25 mol
• sodium chloride 2.90 g, 0.05 mol
• Part F A solution of l-tetralon-7-carboxylic acid (1.0 g, 0.0053 mol) and sodium acetate (1.93 g, 0.024 mol) and hydroxylamine hydrochloride (1.11 g, 0.016 mol) in a mixture of methanol and water (1:1, 15 mL) was stirred at reflux over 4 hours. The mixture was cooled and then added was more water (50 mL) .
• Part G A mixture of l-tetralonoxime-7-carboxylic acid (0.75 g, 0.0037 mol) in methanol (25 mL) with concentrated hydrochloric acid (0.54 mL, 0.20 g, 0.0056 mol) and palladium on carbon catalyst (0.10 g, 5% Pd/C) was shaken for 20 hours at ambient temperature under an atmosphere of hydrogen (60 psi) . The reaction mixture was filtered over CeliteTM and washed with methanol.
• Part A A mixture of l-tetralon-7-carboxylic acid (7.0 g, 0.037 mol) in methanol (13.6 mL, 10.8 g, 0.30 mol) with a catalytic amount of hydrochloriic acid (0.07 mL, 0.12 g, 0.0012 mol) was stirred at reflux over 5 hours. The cooled reaction mixture was poured into ice water and extracted with ethyl acetate. The combined organic layers were backwashed with water and brine, dried over anhydrous magnesium sulfate and evaporated to dryness under reduced pressure. The resulting solid was purified by flash chromatography using hexane:ethyl acetate: :75:25.
• Part B A solution of l-tetralon-7-carboxylic acid methyl ester (3.50 g, 0.017 mol), trimethylsilylcyanide (1.98 g, 0.02 mol) and zinc iodide (0.10 g) in benzene (20 mL) was stirred at ambient temperature over 15 hours. Then added, sequentially and dropwise, was pyridine (20 mL) and phosphorous oxychloride (4.0 L, 6.55 g, 0.0425 mol) . The reaction mixture was stirred at reflux over 1 hour then evaporated to dryness under reduced pressure.
• Part C A mixture of methyl 8-cyano-5, 6-dihydro-2- naphthoate (0.80 g, 0.0038 mol) in methanol (25 mL) with concentrated hydrochloric acid (0.56 mL) and palladium on carbon catalyst (0.40 g, 5% Pd/C) was shaken for 20 hours at ambient temperature under an atmosphere of hydrogen (50 psi) . The reaction mixture was filtered over Celite and washed with methanol.
• Part D A solution of methyl 8-aminomethyl-5, 6,7, 8- tetrahydro-2-naphthoate (0.78 g, 0.0036 mol) and triethylamine (0.55 mL, 0.40 g, 0.004 mol) in aqueous tetrahydrofuran (50%, 75 mL) was added, portionwise as a solid, 2- (tert-butoxycarbonyloxyimino)-2- phenylacetonitrile (0.99 g, 0.004 mol). All was stirred at ambient temperature over 3 hours. The solution was concentrated to half volume and extracted with diethylether.
• the aqueous layer was then acidified to a pH of 1.0 using hydrochloric acid (IN) and then extraced with ethyl acetate.
• the combined organic layers were dried over anhydrous magnesium sulfate and evaporated to dryness under reduced pressure.
• Part B A mixture of methyl 8-cyano-2-naphthoate (1.0 g, 0.0047 mol) in methanol (35 mL) with concentrated hydrochloric acid (0.69 mL) and palladium on carbon catalyst (0.20 g, 5% Pd/C) was shaken for 6 hours at ambient temperature under an atmosphere of hydrogen (50 psi) . The reaction mixture was filtered over Celite 6 and washed with methanol. The filtrate was evaporated to dryness under reduced pressure and the residue was triturated with hexane to give methyl 8- aminomethyl-2-naphthoate (0.76 g, 0.0035 mol, 75%) as an oil.
• Part D A solution of 8-aminomethyl-2-naphthoic acid (0.50 g, 0.00025 mol) and triethylamine (0.038 mL, 0.028 g, 0.000275 mol) in aqueous tetrahydrofuran (50%, 5 mL) was added, portionwise as a solid, 2- (tert-butoxycarbonyloxyimino)-2-phenylacetonitrile (0.068 g, 0.000275 mol). All was stirred at ambient temperature over 5 hours. The solution was concentrated to half volume and extracted with diethylether.
• reaction mixture was stirred for 1.5 hrs to give a white precipitate.
• the solids were filtered off and washed with 6N hydrochloric acid until the filtrate remained clear and then air dried.
• the product was dissolved methanol (500 mL) then ethyl acetate (1000 mL) was added to crystallize 5- (1, 3-dihydro-l, 3- dioxo-2H-isoindol-2-yl) -L-norvaline hydrochloride as white needles mp >230° (131 gm, 0.438, 74%) .
• reaction mixture was extracted with ethyl acetate (2 X 500 mL) .
• the aqueous layer was made acidic with hydrochloric acid (cone) and extracted with ethyl acetate (4 X 500 mL) .
• the combined organic layer was washed with water, brine, dried over magnesium sulfate and concentrated in vacuo to give a colorless oil.
• Part D A suspension of 5- (1,3-dihydro-l,3-dioxo- 2H-isoindol-2-yl)-N-[ (phenylmethoxy)carbonyl]-L- norvaline (140 gm, 0.353 mol), p-toluenesulfonic acid (5.00 gm) and paraformaldehyde (82.8 gm, 2.76 mol) in toluene (1500 mL) in a flask fitted with a Dean Stark trap was heated to reflux for 1 hr.
• reaction mixture was stirred for 18 hrs at ambient temperature, poured into IN hydrochloric acid (600 mL) and was extracted with ethyl acetate (2 X 800 mL) . The combined organic layer was washed with IH hydrochloric acid, water, brine, dried over magnesium sulfate and concentrated in vacuo to give a viscous oil.
• reaction mixture was poured into IN hydrochloric acid (200 mL) and was extracted with ethyl acetate (2 X 300 mL) .
• the combined organic layer was washed with water, brine, dried over magnesium sulfate and concentrated in vacuo to give a viscous oil.

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