LV10111B - Cyclic peptides containing aminomethylbenzoic acid useful as inhibitors of platelet glycoprotein iib/iiia - Google Patents

Abstract

New cyclic peptides containing aminomethylbenzoic acid are useful as described platelet glycoprotein complex IIb / IIIa antagonists. There are pharmaceutical compositions containing these cyclic peptides, and methods of using these compounds to inhibit platelet aggregation.

Description

LV 10111 ΣΙ.71Σ

Cyclic Peptides Containing Aminomethylbenzoic Acid Useful 5 as Inhibitors of Platelet Glycoprotein Ilb/IIIa

FIELD JDF-IHE ..INVENTION 10 15

This invention relates to novel cyclic peptides containing aminomethylbenzoic acid useful as antagonists of the platelet glycoprotein Ilb/IIIa complex# to pharamaceutical compositions containing such cyclic peptides# and to methods of using these compounds for the inhibition of platelet aggregation.

BACKGROUND QF THE INVENTION

Activation of platelets and the resulting platelet aggregation and secretion of factors by the platelets have 20 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 25 diabetes. The contribution of platelets to these disease processes stems from their ability to form aggregates# or platelet thrombi# especially in the arterial vali following injury.

Platelets are known to play an essential role in the 30 maintenance of hemostasis and in the pathogenesis of arterial thrombosis. Platelet activation has been shovrn 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 35 glycoprotein Ilb/IIIa provide biochemical evidence for platelet involvement in unstable angina# early stage of acute 1 2 myocardial infarction, transient ischemic attack, cerebral ischemia, and stroke.

Platelets are activated by a vide variety of agonists resulting in platelet shape change, secretion of granular 5 contents and aggregation. Aggregation of platelets serves to further focus clot formation by concentrating activated clotting factors in one site. Several endogenous agonists includirig adenosine diphosphate (ADP), serotonin, arachidonic acid/ thrombin, and collagen, have been identified. Because 10 of the involvement of several endogenous agonists in activating platelet function and aggregation, an inhibitor which acts against ali agonists would represent a more efficacious antiplatelet aģent than currently available antiplatelet drugs, vhich are agonist-specific. 15 Current antiplatelet drugs are effective against only one type of agonist; these include aspirin, vhich acts against arachidonic acid; ticlopidine, vhich acts against ADP; thromboxane A2 synthetase inhibitors or receptor antagonists, vhich act against thromboxane A2; and hirudin, 20 vhich acts against thrombin.

Recently, a common pathvay for ali knovn agonists has been identified, namely platelet glycoprotein Ilb/IIIa complex (GPIIb/IIIa), vhich is the membrane protein mediating platelet aggregation. A recent reviev of GPIIb/IIIa is 25 provided by Phillips et al. (1991) Celi 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. 30 There is presently a need for a GPIIb/IIIa-specific antiplatelet aģent vhich inhibits the activation and aggregation of platelets in response to any agonist. Such an aģent should represent a more efficacious antiplatelet therapy than the currently--available.agonist-specific 35 platelet inhibitors. 2 5 3 LV 10111 GPIIb/IIIa does not bind soluble proteīns on unstimulated platelets, but GPIIb/IIIa in activated platelets is kr.cwn to bind four sclubie achesive proteīns, r.atr.ely fibrinogen, von Millebrand factor, fibronectin, and vitronectin. 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) rečognition sequence which is common to the adhesive proteīns that bind GPIIb/IIIa. 10 15 20 25

Several RGD-containing peptides and related compounds have been reported which block fibrinogen binding and prevent the formation of platelet thrombi. For example, see Cadroy et al. (1989) J. Clin. Invest. 84: 939-944/ Klein et al. U.S. Patent 4,952,562, issued 8/28/90; European Patent Application EP 0319506 A; European Patent Application EP 0422938 Al; European Patent Application EP 0422937 Al; European Patent Application EP 0341915 A2; PCT Patent Application WO 89/07609; PCT Patent Application WO 90/02751; PCT Patent Application WO 91/04247; and European Patent Application EP 0343085 Al.

In the present invention we use rigid, conformationally-constraining amino acids (isomers of aminomethylbenzoic acid) as templates for cyclizing peptides such that they have high affinity and selectivity for GPIIb/IIIa.

DETAILED . DESCRIPTION OF THE INVENT.IDN

This invention is directed to novel compounds of the formula (I): 30 3

4/ «j \ 0=C κ' Μ \ Νκ/ . CHR4 (ΐ) ,10 or a pharmaceutically acceptable salt or prodrug form thereof vherein: 5 r1 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(C2-C4)alkyl; r2 is H or Ci-Ca alkyl; 10 R1^ is H' cl"c8 phenyl, halogen, or C1-C4 alkoxy; j is β-Ala or an L-isomer or D-isomer amino acid of structure -N(R3)C<R4) (R5)C(=0)-, vherein: 15 20 r3 is H or CH3; R4 is H or C1-C3 alkyl; R5 is H, Ci~C8 alkylf C3-C6 cycloalkyl, C3-C6 cycloalkylmethyl, Cļ-Cg cycloalkylethyl, phenyl, phenylmethyl, CH2OH, CH2SH, CH2OCH3, CH2SCH3, CH2CH2SCH3, (CH2)SX vhere X is -NH2 or -NHC(=NH) (NH2) and s = 3-5/ or r3 and R$ can be taken together to form - (CH2) t“ (t * 2-4) or -CH2SC(CH3)2-;or R4 and r5 can be taken together to form -(CH2)u"/ vhere u - 2-5; K is an L-isomer amino acid of structure -N(R6)CH(R7)C(*0)-, vherein: 4 25 5 5LV 10111 r6 is H or Ci-Ca alkyl; R7 is -(CH2>pNHC(=NH> (NK2>» vhere p = 3-5; 10 15 20 25

”(CH2)rX/vhere r * 4-6;

ch2x - ch2 ^-ch2x -(CH2)mS(CH2)2X/ vhere m - 1/2; and X is -NH2 or . muo when -NHC(=NH)(NH2)/ provided that X cannot be - ^ r = 4; or (CH2)nX r6 and R7 are taken together to form “CH2CHCH2 , n = 0,1 and X is -NHC(=NH)(NH2)i • 1 is -Y(CH2)vC(»0)vherein: γ is NH, 0, or S; and v « 1,2; B\C M is an L-isomer amino acid of structure -NH-CH(R > wherein: r8 is -CH2CO2H,-CH2SO3H, or -CH(CH3)C02H;

Preferred compounds of the invention are 1,3-disubstituted phenyl compounds of the formula (II): 5

(IX)°"CV^V U CHR* v/herein: R1 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(Cļ- C4)alkyl; is H or methyl; J is β-Ala or an L-isomer or D-isomer amino acid of structure -N(R3)C(R4) (R3)C(=0)-, vherein: R3 is H or CH3; R4 is H or C1-C3 alkyl; R3 is Hf C1-C8 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkylmethyl, C1-C6 cycloalkylethyl, phenyl, phenylmethyl, CH2OH, CH2SH, CH2OCH3, CH2SCH3, CH2CH2SCH3, (CH2)8X vhere X is NH2 or NHC(=NH) (NH2) and s - 3-5; or R3 and R3 can be taken together to form -CH2CH2CH2”/ or R4 and R3 can be taken together to form -(CH2)u"' where u * 2-5; K is an L-isomer amino acid of structure -N(R3)CH(R7)C(=0)-, vherein: r6 is H or C1-C8 alkyl; R7 is -(CH2)pNHC(=NH)(NH2)/ where p - 3,4; 7 5 LV 10111

-(CH2)mS(CH2>2X» where m - 1,2; and X is -NH2 or -NHC(=NH)(NH2)/ provided that X cannot be -NH2 when r » 4; or 10

(CH2)nX

I R6 and R7 are taken together to form "CH2CHCH2-/ where n - 0,1 and X is -NHC (=NH)(NH2); L is -Y(CH2)vC(=0)-, wherein: 15 y is NH, 0, or S; and v - 1,2; M 20 is an L-isomer amino acid of structure -NH-CH(R®)C(eO)-, wherein: R8 is -CH2CO2H,“CH2SO3H, or -CH(CH3)CO2H;

Further preferred compounds of the invention are 1»3-disubstituted phenyl compounds of the formula (II): 25 7

8 / J

\ 0=C

(II) wherein: r! is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(C2“ C4)alkyl; r2 is H or methyl; J is β-Ala or an L-isomer or D-isomer amino acid of structure -N (R^) C (R4) (R^) C (=0)wherein: r3 is H or CH3; R4 is H; r5 is H, C1-C8 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkylmethyl, Ci-Cg cycloalkylethyl, phenyl, phenylmethyl, CH2OH, CH2SH, CH2OCH3, CH2SCH3, CH2CH2SCH3, (CH2)SX where X is NH2 or NHC(=NH) (NH2) and s « 3,4,5; or r3 and R^ can be taken together to form -CH2CH2CH2-; K is an L-isomer amino acid of structure -N(R6)CH(R7)C(=0)-, vherein: R6 is H or C3-C8 alkyl; R7 is -(CH2)pNHC(«NH)(NH2)/ where p - 3,4;

8 9 LV 10111 9

- (CH2)rx»wkere r = 4-6;

5 -(CH2)mS(CH2)2X/ where m » 1,2; and X is -NH2 or -NHC(=NH)(NH2), provided that X cannot be -NH2 vhen r « 4; or L is -YCH2C (=0)-/ vherein: 10 Y is NH or 0; M 15 is an L-isomer amino acid of structure -NH-CH(R8)C(=0)-, wherein: R8 is -CH2CO2H or -CH2SO3H.

More preferred compounds of the present invention are compounds of the further preferred scope above, wherein: 20 is H; is H; is β-Ala or an L-isomer or D-isomer amino acid of formula -N(R3)CH(R5)C(=0)-, v/herein: R3 is H and RS is H, CH3, CH2ČH3, CH{CH3)2/ CH(CH3)CH2CH3, CH2CH (CH3) 2, (CH2) 4NH2; or R3 is CH3 and R5 is H; or R3 and R3 can be taken together to form -CH2CH2CH2·-. R1 R2 J 25 9 10 K is an L-isomer amino acid of formula -N(CH3)CH(R7)C(=0)-, vherein: R7 is -(CH2)3NHC(=NH)(NH2); L is -NHCH2C(e0)and M ' is an L-isomer amino acid of formula -NHCH(CH2C00H)C(=O)-.

Specifically preferred are the following compounds:

The compound of formula (II) vherein R* and R2 are H; J is D-Val; K is NMeArg; L is Gly/ and M is Asp.

The compound of formula (II) vherein R^ and R2 are H; J is D-2-aminobutyric acid; K is NMeArg; L· is Gly; and M is Asp.

The compound of formula (II) vherein R^ and R2 are H; J is D-Leu; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R^· and R2 are H; J is o-Ala; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R* and R2 are H; J is Gly; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R^· and R2 are H; J is D-Pro; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R1 and R2 are H; J is o-Ser; K is Arg; L is Gly; and M is Asp.

The compound of formula (II) vherein R1 and R2 are H; J is D-Lys; K is NMeArg; L is Gly; and M is Asp. 10 5 11 LV 10111

The compound of formula (II) wherein R* and R^ are H; J is D-2-aminobutyric acid; K is Arg; L is Gly; and M is Asp.

The compound of formula (II) vherein R* and are H; J is D-Ala; K is Arg; L is Gly; and M is Asp.

The compound of formula (II) wherein R* and R^ are H; J 10 is D-Val; K is Arg; L is Gly; and M is Asp.

The compound of formula (II) wherein R^· and R^ are H; J is β-Ala; K is NMeArg; L is Gly; and M is Asp. 15 The compound of formula (II) vherein R^· and are H; J is D-Val; K is NMeArg; L is β-Ala; and M is Asp.

The compound of formula (II) vherein R* and R^ are H; J is Pro; K is NMeArg; L is Gly; and M is Asp. 20

The compound of formula (II) vherein R* and R^ are H; J is Phe; K is Arg; L is Gly; and M is Asp.

The compound of formula (II) vherein R* and R^ are H; J 25 is Gly; K is Arg; L is Gly; and M is Asp.

The compound of formula (II) vherein R* and are H; J is NMeGly; K is NMeArg; L is Gly; and M is Asp. 30 The compound of formula (II) vherein R* and R^ are H; J is D-Leu; K is Arg; L is Gly; and M is Asp.

The compound of formula (III) vherein R* and R^ are H; J is D-Val; K is NMeArg; L is Gly; and M is Asp. 11 35 12 /

Μ \ NR1'

(III)

More specifically preferred are the folloving compounds of formula (I), ali of which have IC50 values of less than 5 1.0 μΜ for inhibiting the aggregation of platelets (as described below).

The compound of formula (II) wherein R^· and R^ are H; J is D-Val; K is NMeArg; L is Gly; and M is Asp. 10

The compound of formula (II) vherein R* and R^ are H; J is D-2-aminobutyric acid; K is NMeArg; L is Gly; and M is Asp. 15 The compound of formula (II) wherein R^· and R^ are H; J is D-Leu; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R* and R^ are H; J is D-Ala; K is NMeArg; L is Gly; and M is Asp. 20

The compound of formula (II) vherein R^ and R^ are H; J is Gly; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R* and R^ are H; J 25 is D-Pro; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) vherein R1 and R^ are H; J is D-Lys; K is NMeArg; L is Gly; and M is Asp. 12 13 LV 10111

The compound of formula (II) wherein R^ and R^ are H; J is (^-Ala; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (II) wherein R^· and R^ are H; J is NMeGly; K is NMeArg; L is Gly; and M is Asp.

The compound of formula (III) wherein R* and R^ are H; J is D-Val; K is NMeArg; L is Gly; and M is Asp.

(III)

In the present invention it has been discovered that the compounds above are useful as inhibitors of glycoprotein Ilb/IIIa (GPIIb/IIIa). As discussed above, GPIIb/IIIa mediates the process of platelet activation and aggregation.

The compounds of the present invention inhibit the activation and aggregation of platelets induced by ali ķnovm endogenous platelet agonists.

The present invention also provides methods for the treatment of conditions involving platelet activation and aggregation, including cardiovascular and cerebrovascular thromboembolic disorders, for example, thromboembolic disorders associated with unstable angina, myocardial infarction, transient ischemic attacJc, stroke, atherosclerosis, and diabetes, by administering to a host in need of such treatment a pharmaceutically effective amount of the compounds of formula (I) above. 13 14 GPIIb/IIIa is known to be overexpressed in metastatic tumor celis. The compounds of the present invention may also be useful for the treatmer.t cf metastatic cancer.

The compounds herein described may have asymmetric centers. Unless otherwise indicated, ali chiral, diastereomeric and racemic forms are included in the present invention. Two distinct isomers (cis and trans) of the peptide bond are known to occur; both can also be present in the compounds described herein, and ali such stable isomers are contemplated in the present invention. Unless othervise specifically noted, the L-isomer of the amino acid is used. The D and L-isomers of a particular amino acid are designated herein using the conventional 3-letter abbreviation of the amino acid, as indicated by the following examples: D-Leu, D-Leu, L-Leu, or L-Leu.

When any variable (for example, R1 through R®, m, n, p, Q, W, X, Y, Z, etc.) occurs more than one time in any constituent or in any formula, its definition on each occurrence is independent of its definition at every other occurrence. Also, combinations of substituents and/or variables are permissible only if such combinations resuit ir. stable compounds.

As used herein, ”alkyl" is intended to include both branched and straight-chain saturated aliphatic hydrocarbo.n 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. "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 14 15 LV 10111 and the like; and "alkynyl" is intended to include hydrocarbon chains of either a straight or branched cc-nf iguration and c-r.e c: m.ore tripie carbon-carbon bor.ds wnich may occur in any stable point along the chain, such as' 5 ethynyl, propynyl and the like. "Halo" as used herein refers to fluoro, chloro, bromo and iodo; and "counterion" is used to represent a small, negati.vely charged species such as chloride, bromide, hydroxide, acetate, sulfate and the like.

By "stable compound" or "stable structure" is meant 10 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 aģent.

As used herein, "pharmaceutically acceptable salts and 15 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 20 include, but are not limited to: mineral or organic acid salts of basie 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. 25 Pharmaceutically acceptable salts 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 30 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 Publishing Company, Easton, PA, 1985, p. 1418, the disclosure of which is hereby incorporated by reference. 35 The term "amino acid" as used herein means an organic compound containing both a basie amino group and an acidic 15 16 carboxyl group. Included vithin this term are modified and unusual amino acids.

The term "amino acid residue" as used herein means t'nat portion of an amino acid (as defined herein) that is present 5 in a peptide or pseudopeptide. The term "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. 10 The compounds of the present invention can be prepared in a number of ways well known to one skilled in the art of organic synthesis. Preferred methods include but are not limited to those methods described below. 15 The following abbreviations are used herein: D-Abu β-Ala or o-2-aminobutyric acid bAla 3-aminopropionic acid 20 Boc t-butyloxycarbonyl Boc-Mamb t-butyloxycarbonyl-3-aminomethylbenzoic acid Boc-ON 2- (tert-butyloxycarbonyloxylimino)-2-phenylacetonitrile DCC dicyclohexylcarbodiimide 25 DIEA diisopropylethylamine DMA? 4-dimethylaminopyridine HBTU NMeArg or 2-(lH-Benzotria2ol-l-yl)-l,l,3,3- tetramethyluronium hexafluorophosphate 30 MeArg NTMeGly or a-N-methyl arginine MeGly N-methyl glycine NMM N-methylmorpholine OcHex 0-cyclohexyl 35 OBzl 0-benzyl ' Tos tosyl 16 17 LV 10111

The folloving conventional three-letter amino acid abbreviat icr.s are used herein; the conventional one-letter amino acid abbreviations are HClL used herein:

Ala s alanine Arg 2t arginine Asn St asparagine Asp 3 aspartic acid Cys 3 cysteine Gln * glutamine Glu * glutamic acid Gly at glycine His 3 histidine Ile - isoleucine Leu 3 leucine Lys 3 lysine Met S* methionine Phe = phenylalanine Pro s proline Ser = serine Thr 3 threonine Trp = tryptophan Tyr 3 tyrosine Vai 3 valine

Peotide Svnthesis

The compounds of the present invention can be synthesized using Standard synthetic methods known to those skilled in the art. Generally, peptides are elongated by deprotecting the Ct-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 seguence is obtained. This coupling can be performed with the constituent amino acids in a stepvvise fashion, or 17 18 condensation of fragments (tvo to several amino acids), or combination of both processes, or by solid phase peptide synthesis according to the method originally described by Merrifield, J. Am. Chem. Soc., 85, 2149-2154 (1963), the disclosure of which is hereby incorporated by reference.

The compounds of the invention may also be synthesized using automated peptide synthesizing equipment. In addition to the foregoing, procedures for peptide synthesis are described in Stewart and Young, "Solid Phase Peptide Syrithesis", 2nd ed, Pierce Chemical Co., Rockford, IL (1984); Gross, Meienhofer, Udenfriend, Eds., "The Peptides: Analysis, Synthesis, Biology, Vol. 1, 2, 3, 5, and 9, Academic Press, New York, (1980-1987)/ Bodanszky, "Peptide Chemistry: A Practical Textbook", Springer-Verlag, New York (1988); and Bodanszky et al. "The Practice of Peptide Sythesis" Springer-Verlag, New York (1984), the disclosures of which are hereby incorporated by reference.

The coupling betveen two amino acid derivatives, an amino acid and a peptide, tvo 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 vater-soluble carbodiimides) method, active ester (p-nitrophenyl ester, N-hydroxysuccinic imido ester) method, Woodward reaģent K method, carbonyldiimidazole method, phosphorus reaģents such as BOP-C1, or oxidation-reduction method. Some of these methods (especially the carbodiimide) can be enhanced by the addition of l-hydroxybenzotriazole. These coupling reactions may be performed in either solution (liquid phase) or solid phase.

The functional groups of the constituent amino acids must be protected during the coupling reactions to avoid undesired bonds being formed. The protecting groups that can be used are listed in Greene, "Protective Groups in Organic Synthesis" John Wiley 4 Sons, Nev York (1981) and "The 18 19 LV 10111

Peptides: Analysis, Sythesis, Biology, Vol. 3, Academic Press, New York (1981), the disclosure o£ which is herebv ir.corporated by reference.

The ct-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. In the solid phase case, the C-terminal amino acid is attached to an insoluble carrier (usually polystyrene). These insoluble carriers contain a group vhich will react with the carboxyl group to form a bond vhich is stable to the elongation conditions but readily cleaved later. Examples of vhich are: oxime resin (DeGrado and Kaiser (1980) J. Org. Chem. 45, 1295-1300) chloro or bromomethyl resin, hydroxymethyl resin, and aminomethyl resin. Many of these resins are commercially available vith the desired C-terminal amino acid already incorporated.

The α-amino group of each amino acid must be protected. Any protecting group knovn in the art can be used. Examples of these are: 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)-l-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 triphenylmethy1 and benzyl; 6) trialkylsilane such as trimethylsilane; and 7) thiol containing types such as phenylthiocarbonyl and dithiasuccinoyl. The preferred (X- amino protecting group is either Boc or Fmoc. Many amino acid 19 derivatives suitably protected for peptide synthesis are ccmmercialiy available,

The α-amino protecting group is cleaved prior to the coupling of the next amino acid. When 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 basie Solutions such as aqueous buffers, or tertiary amines in dichloromethane or dimethylformamide.

When the Fmoc group is used, the reaģents of choice are piperidine or substituted piperidines in dimethylformamide, but any secondary amine or aqueous basie Solutions can be used. The deprotection is carried out at a temperature betv/een 0 °C and room temperature.

Any of the amino a.cids 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 a-amino group.

For example, when Boc is ehosen for the a-amine protection the foliowing 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. 20 21 LV 10111

When Fmoc is chosen fcr the α-amine protection usually tert-butyl based protecting groups are acceptable. For instance, Boc can be used fcr lysine, tert-butyi ether fcr serine, threonine and tyrosir.e, and tert-butyl ester for 5 glutamic and aspartic acids.

Once the elongation and cyclization of the peptide is completed ali of the protecting groups are removed. For the liquid phase synthesis the protecting groups are removed in whatever manner as dictated by the choice of protecting 10 groups. These procedures are well known to those skilled in the art.

When a solid phase synthesis is used, the peptide should be removed from the resin vithout simultaneously removing protecting groups from functional groups that might interfere 15 with the cyclization process. Thus, if the peptide is to be cyclized in solution, the cleavage conditions need to be chosen such that a free a-carboxylate and a free a-amino group are generated vithout simultaneously removing other protecting groups. Alternatively, the peptide may be removed 20 from the resin by hydrazinolysis, and then coupled by the azide method. Another very convenient method involves the synthesis of peptides on an oxime resin, folloved by intramolecular nucleophilic displacement from the resin, which generates a cyclic peptide (Osapay, Profit, and Taylor 25 (1990) Tetrahedron Letters 43, 6121-6124). When the oxime resin is employed, the Boc protection scheme is generally chosen. Then, the preferred method for removing side chain protecting groups generally involves treatment with anhydrous KF containing additives such as dimethyl sulfide, anisole, 30 thioanisole, or p-cresol at 0 ®C. The cleavage of the peptide can also be accomplished by other acid reaģents such as trifluoromethanesulfonic acid/trifluoroacetic acid mixtures.

Unusual amino acids used in this invention can be 35 synthesized by Standard methods familiar to those skilled in the art ("The Peptides: Analysis, Sythesis, Biology, Vol. 5, 21 22 ρρ. 342-449, Academic Press, N’ew York (1981)). N-Alkyl amino acids can be prepared usir.g proceedures described in previously (Cheung et ai., (1977) Can. J. Chem. 55, 906; Ereidinger et ali, (1982) J. Org, Chem. 48, 77 (1982)), which are incorporated here by reference.

The compounds of the present invention may be prepared using the procedures further detailed below.

Representative materiāls and methods that may be used in preparing the compounds of the invention are described further below.

Manual solid phase peptide synthesis was performed in 25 mL polypropylene filtration tubes purchased from BioRad Inc. Oxime resin (substitution Ievel =0.96 mmol/g) was prepared according to published procedures (DeGrado and Kaiser (1980) J. Org. Chem. 45, 1295). Ali Chemicals and solvents (reaģent grade) were used as supplied from the vendors cited without further purification. 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-(ĪH-Benzotriazol- l-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HBTU) was purchased from Advanced ChemTech. N-methylmorpholine (NMM), m-cresol, D-2-aminobutyric acid (Abu), trimethylacetylchloride, diisopropylethylamine (DIEA), 3-cyanobenzoic acid and [2-(tert-butyloxycarbonyloxyiimino)-phenylacetonitrile) (Boc-ON) were purchased from Aldrich Chemical Company. Dimethylfcrmamide (DMF), ethyl acetate, chloroform (CHCI3), methanol (MeOH), pyridine and hydrochloric acid (HC1) 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 Fluka Chemical Company. Absolute ethanol was obtained 22 23 LV 10111

from Quantum Chemical Corporation. Thin layer chromatography (TLC) was performed on Silica Gel 60 F254 TLC - - ·> —, plates (iayer thickr.es were purcr.asea frcn ΞΜ

Separations. TLC visualization was accomplished using UV light and/or ninhydrin spray. Melting points were determined using a Thomas Hoover melting point apparatus and are uncorrected. Ali HPLC analyses were performed on either a Hevlett Packard 1090, Rainin, or DuPont 8800 system. NMR spectra were recorded on a 300 MHz General Electric QE-300 spectrometer. Fast atom bombardment mass spectrometry (FAB-MS) was performed on a VG Zab-E double-focusing mass spectrometer using a Xenon FAB gun as the ion source.

Synthesis of 2. .3. and 4-substituted. Boc-aminometh/lbenzoic

Acid Derivatives 2, 3, and 4-substituted Boc-aminomethylbenzoic acid derivatives useful as intermediates in the synthesis of the compounds of the invention are prepared using Standard procedures, for example, as described in Tett. Lett., 4393 (1975); Modēm Synthetic Reactions, H.O. House (1972); or Harting et al. J. Am. Chem. Soc., 50: 3370 (1928), and as shown schematically below.

NH—BOC 3-Cyanobenzoic acid (10.0 g, 68 mmol) was dissolved in 200 ml ethanol by heating in a 35-50°C water bath. Concentrated HC1 (6.12 ml, 201 mmol) was added and the solution was transferred to a 500 ml nitrogen-flushed round bottom flask 23 24 containing palladium on carbon catalyst (1.05 g, 10% Pd/C). The suspension vas stirred under an atmosphere cf hydrogen fcr 36 hours, filtered throuah a scintered glass funnel, ar.d vashed thoroughly vith H2O. The ethanol was removed under 5 reduced pressure and the remaining aqueous layer, which contained a white solid, was diluted to 250 ml with additional H2O. Ethyl ether (250 ml) vas added and the suspension vas transferred to a separatory funnel. Upon vigorous shaking, ali solids dissolved and the aqueous layer 10 was then vashed tvo times vith ether, evaporated under reduced pressure to a volume of 150 ml, and lyophiiized to give the title compound (3-aminomethylbenzoic acid*HCl) (8.10 g, 64%) as a beige solid. NMR (D2O) 4.27 (s, 2H), 7.60 (t, 1H), 7.72 (d,1H), 8.06Z (d, 2H) . 15 t-Butyloxycarbony1-3-aminomethylbenzoic Acid (Boc-Mambl

The title compound vas prepared according to a modification of Standard procedures previously reported in 20 the literature (Itoh, Hagivara, and Kamiya (1975) Tett.

Lett., 4393). 3-Aminomethylbenzoic acid (hydrochloride salt) (3.0 g, 16.0 mmol) vas dissolved in 60 ml H2O. To thiš vas added a solution of Boc-ON (4.33 g, 17.6 mmol) in 60 ml acetone folloved by triethylamine (5.56 ml, 39.9 mmol). The 25 solution turned yellov and the pH vas adjusted to 9 (wet pH paper) by adding an additional 1.0 ml (7,2 mmol) triethylamine. The solution vas stirred overnight at room temperature at vhich time the acetone vas removed under reduced pressure and the remaining'aqueous layer vas vashed 30 three times vith ether, The agueous layer vas then acidified to pH 2 vith 2N HC1 and then extracted three times vith ethyl acetate. The combined organic layers vere vashed three times vith H2O, dried over anhydrous magnesium sulfate, and evaporated to dryness under reduced pressure. The material 35 vas recrystallized from ethyl acetate/ hexane to give tvo crops of (2) (2,58 g, 64%) as an off-vhite solid. mp 123- 24 25 LV 10111 125°C /½ NMR (CDCI3) 1.47 (s, 9 H), 4.38 <br s, 2 H) , 4.95 (br s, 1H), 7.45 (t, 1H), 7.55 (d, 1H) , 8.02 <d, 2H) . tr5utV-lQ£V£fl.rbonvl-M-methyl-3-amlnQir.ethylbenzoic d n^nr-5 Mamb),

The title compound can be prepared according to Standard procedures, for examples, as disclosed in Olsen, J. Org.

Chem. (1970) 35: 1912), and as shown schematically below. 10

Synthesis of Aminomethvlbeiiroic Ācid Analogs

1) NaOH/EtOH 2) H* 15 Intermediates of the formula below may be prepared using

Standard synthetic procedures, for example, as shown in the indicated reaction schemes shown below.

For R =» CH3, CH2CH3, CH2CH2CH3, CH2CH2CH2CH3, CH(CH3)2, C(CH3)3, CH(CH3)CH2CH3, benzyl, cyclopentyl, cyclohexyl; see Scheme 1.

For R - CH3, CH2CH2CH2CH3, phenyl; see Scheme 2. 25

For R Γ , pher.yl; see Sc'r.er.e 3 ar.d 4.

10% Pd-C/H2/ HCI.EtOH

Ph

Ph

EtjN/Tol./BFj.EtjO PhjCO Reflu*

MeO

υ»

8'/RX

(i) Deprotection ’(iī) BOC-OH

26 27 ££iL£JH£_2:

N 0 0 o

HjS 04»q/Δ OR KOH/nPrOH/Δ ;N*N(SIM«3)2 »

LV 10111 27 28 Schem.e 3: Ο Ο

(i) NHj/CHjOH/MOL. SIEVES

(ii) NaBH4/HCI

OR NH4OAc/Na(CN)BHj/CH3OH/M.S. 5

HO

NH-BOC 28 29 Scheme 4: LV 10111 0 0

:NH2OH/HCI/C2H5OH/Py i

: (i) Reductlon i (II) BOC-ON t

0 NH-BOC

The title compound was prepared by a modification of procedures previously reported in the literature (Itoh, Hagivara, and Kamiya (1975) Tett. Lett., 4393), as shown in 10 the scheme below. 29 30 ΝΗ2

0

BOC-ON

pH = 9, EtjN

OH O D-2*amlnobutyrlc ecld D-2-aminobutyric acid (1.0 g, 9.70 mmol) was dissolved in 20 ml H2O and a solution of Boc-ON (2.62 g, 10.6 mmol) in 20 ml acetone was added. A white precipitate formed which dissolved upon addition of triethylamine (3.37 ml, 24.2 mmol) to give a pale yellow solution (pH « 9, wet pH paper). The solution was stirred at room temperature overnight at which time the acetone was removed under reduced pressure. The remaining aqueous layer was extracted with ether three times, acidified to pH 2 with concentrated HC1, and then extracted with ethyl acetate three times. The combined organic layers were dried over anhydrous magnesium sulfate and evaporated under reduced pressure to give t-butyloxycarbonyl-D-2-aminobutyric acid as an oil (2.05 g,greater than quantitative yield, contains solvent), which was used without further purification. NMR (CDCI3) 0.98 (t, 3H), 1.45 (s, 9H), 1.73 (m, 1H), 1.90 (m, 1H), 4.29 (m, 1H), 5.05 (m, 1H) .

Svnthesis of Cvclic Peotides t-Butyloxycarbonyl-3-aminomethylbenzoic acid (Boc-Mamb) is coupled to oxime resin by a modification of the method described by DeGrado and Kaiser (1980) J. Org. Chem. 45, 1295 using 1 equivalent of the 3-aminomethylbenzoic acid (with respect to the substitution Ievel of the resin), 1 equivalent of HBTU, and 3 equivalent of NMM. Alternatively, Boc-Mamb (1 equivalent) may be coupled to the oxime resin using 1 equivalent each of DCC and DMAP in methylene chloride, Coupling times range from 15 to 96 hours. The substitution Ievel is then determined using either the picric acid tēst (Sarin, Kent, Tam, and Merrifield, (1981) Anal. Biochem. 117, 30 31 LV 10111 145-157) or the quantitative ninhydrin assay (Gisin (1972) Anal. Chim. Acta 58, 248-249). Unreacted oxime groups are fclecked using 0.5 M crimec r.v lacety ichlor ide / 0.5 M diisopropy lethylamine in DMF for 2 hours. Deprotection of the Boc protecting group is accomplished using 25% TFA in DCM for 30 minūtes. The remaining amino acids or amino acid derivatives are coupled using between a two and ten fold excess (based on the loading of the first amino aciu or amino acid derivative) of the appropriate amino acid or amino acid derivatives and HBTU in approximately 8 ml of DMF. The resin is then neutralized in situ using 3 eq. of NMM (based on the amount of amino acid used) and the coupling times range from 1 hour to overnight. The completeness of coupling is monitored by qualitative ninhydrin assay, or picric acid assay in cases where the amino acid was coupled to a secondary amine. Amino acids are recoupled if necessary based on these results.

After the linear peptide had been assembled, the N-terminal Boc group is removed by treatment with 25% TFA in DCM for 30 minūtes. The resin is then neutralized by treatment with 10% DIEA in DCM. Cyclization with concomitant cleavage of the peptide is accomplished using the method of Osapay and Taylor ((1990) J. Am. Chem. Soc., 112, 6046) by suspending the resin in approximately 10 ml/g of DMF, adding one equivalent of HOAc (based on the loading of

O the first amino acid), and stirring at SOC for 60 to 72 hours. Folloving filtration through a scintered glass funnel, the DMF filtrate is evaporated, redissolved in HOAc or 1:1 acetonitrile: H2O, and lyophilized to obtain protected, cyclized material. This is then treated using Standard procedures with anhydrous hydrogen fluoride (Stevart and Young (1984) "Solid Phase Peptide Synthesis", 2nd. edition, Pierce Chemical Co., 85) containing 1 ml/g m-cresol 0 as scavenger at 0 C for 20 to 60 minūtes to remove side cham protecting groups. The crude product may be purified by reversed-phase HPLC using a 2.5 cm preparative Vydac C18 31 32 column with a linear acetonitrile gradient containing 0.1% TFA to producē pure cyclized tr.aterial. The following N-a-

Bcc-prctected amino acids m.ay be used for the syntheses: 5oc-Arg(Tos), Boc-N-a-MeArg(Tos), 3oc-Gly, Boc-Asp(OcHex), Boc-D-

Leu, Boc-D-Val, Boc-D-2-aminobutyric acid (Boc-D-Abu), Boc-Phe, Boc-D-Ser(Bzl) , Boc-D-Ala, Boc-3-aminomethylben2oic acid (Boc-Mamb), Boc-D-Lys(2-C1Z), Βοο-β-Ala, Boc-D-Pro, or Boc- NMeGly.

The synthesis of the compounds of the invention is further exemplified below. Tables 1-6 below set forth representative compounds of the present invention.

Example 1 cyclo-(Gly-NMeArg-Gly-Asp-Mamb); J =* Gly, K * NMeArg, L = Gly, M = Asp, R1 - R2 » H (SEQ ID NO:1)

The title compound was prepared using the general procedure described below for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb) . The peptide was prepared on a 0.336 minol scale to give the protected cyclic peptide (218 mg, 84%) . The peptide (200 mg) and 200 pL of m-cresol were treated with anhydrous hydrogen fluoride at 0°C for 1 hour. The crude material vas precipitated with ether, redissolved in aqueous HOAc, and lvophilized to generate the title compound as a pale yellow solid (158 mg, greater than quantitative yield; calculated as the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 2 to 11% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy white solid (21% recovery, overall yield 16.3%),

Mass spectrum: M+H » 533.26. 32 33 LV 10111

Example 2 cyclo- (D-Ala-NMeArg-Gly-Asp-Mamb); J » D-Ala, K « NMeArg,

L - Gly, M - Asp, R1 - R2 - H

The title compound was prepared using the general procedure described below for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb) . Recoupling of the Boc-N-MeArg(Tos) residue was found to be necessary. The peptide was prepared on a 0.24 4 mmol scale to give the protected cyclic peptide (117 mg, 61%).

The peptide (110 mg) and 110 μι. of m-cresol were treated

O with anhydrous hydrogen fluoride at 0 C for 1 hour. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yellow solid. Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.25%/ min. gradient of 2 to 11% acetonitrile containing 0.1% ΤΓΑ and then lyophilized to give the TFA salt of the title compound as a fluffy white solid.

Mass spectrum: M+H - 547.23.

Example 3 cyclo- (D-Abu-NMeArg-Gly-Asp-Mamb); J * D-Abu, K » NMeArg,

L - Gly, M - Asp, R^ R2 H

The title compound was prepared using the general procedure described below for Example 4. The peptide was prepared on a 0.101 mmol scale to give the protected cyclic peptide (51 mg, 63%) . The peptide (43 mg) and 50 μΐ, of m-

O cresol were treated with anhydrous hydrogen fluoride at 0 C for 30 minūtes The crude material was precipitated vith ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yellow solid (23 mg, 68.7%; calculated a3 the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/min. gradient of 7 to 14% 33 34 acetonitrile containing 0.1¾ tri lyophilized to give the ΤΓΑ salt fluffy vhite sclid (31* rectv^rv Mass spectrum: M+H = 561.45. fluoroacetic acid and t of the title compound ; c ve r a 11 y i e1d 12.4*). hen as a . Example 4 cyclo-(D-Val-NMeArg~Gly-Asp-Mamb) ; J = D-Val, K = NMeArg, l =

Gly, M » Asp, R1 « R2 = H

To a 25 ml polypropylene tube fitted with a frit was added Boc-Mamb (0.126 g, 0.5 mmol) and 6 ml of DMF. To this was added HBTU (0.194 g, 0.5 mmol), oxime resin (0.52 g, substitution Ievel = 0.96 mmol/g), and N-methylmorpholine (0.165 ml, 1.50 mmol). The suspension was mixed at room temperature for 24 hours. The resin was then vashed thoroughly (10-12 ml volumes) with DMF (3x), MeOH (lx), DCM (3x), MeOH (2x) and DCM (3x). The substitution Ievel was determined to be 0.389 mmol/g by quantitative ninhydrin assay. Unreacted oxime groups were blocked by treatment vith 0.5 M trimethylacetylchloride/ 0.5M DIEA in DMF for 2 hours.

The following steps were then performed: (Stepi) The resin was washed with DMF(3x), MeOH (lx), DCM (3x), MeOH (2x), and DCM (3x). (Step 2) The t-Boc group was deprotected using 25% TFA in DCM for 30 minūtes. (Step 3) The resin was vashed vith DCM (3x), MeOH (lx), DCM (2x), MeOH (3x) and DMF (3x) (Step 4) Boc-Asp(OcHex) (0.613 g, 1.94 mmol), HBTU (0.753 g, 1.99 mmol), 8 ml of DMF, and N-methylmorpholine (0.642 ml, 5.84 mmol) were added to the resin and the reaction alloved to proceed for 2.5 hours. (Step 5) The coupling reaction was found to be complete as assessed by the qualitative ninhydrin assay. Steps 1-5 were repeated until the desired sequence had been attained. The coupling of Boc-D-Val to NMeArg was monitored by the picric acid tēst,

After the linear peptide was assembled, the N-terminal t-Boc group was removed by treatment with 25% TFA in DCM (30 min.) The resin was vashed thoroughly vith DCM (3x), MeOH 34 35 LV 10111 (2x) and DCM (3x), and ther. neutralized with 10¾ DIEA ir. DCM (2x1 min.) Tne resin was washed thoroughly with DCM (3x) ar.d MeCH (3x) and ther. dried. Half of the resin (0.101 mmol) was cyclized by treating with 6 ml of DMF containing HOAc (5.8 μ!,, 0.101 mmol) and heating at 50eC for 72 hours. The resin was then filtered through a scintered glass funnei and washed thoroughly with DMF. The DMF filtrate was evaporated to an oil, redissolved in 1:1 acetonitrile: H20, anu lyophilized to give the protected cyclic peptide (49 mg, 60%). The peptide (42 mg) was treated with anhydrous hydrogen fluoride at 0°C, in the presence of 50 μΐ> of m- cresol as scavenger, for 30 minūtes to remove side chain protecting groups . The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yellow solid (23 mg, 10%; calculated as the acetate salt) . Purification was accomplished using reversed-phase HPLC with a preparative Vydac C18 column (2.5 cm) and a 0.23%/ minūte gradient of 7 to 18% acetonitrile containing 0.1% trifluoroacetic acid to give the TFA salt of the title compound as a fluffy white solid (24% recovery; overall yield 9.4%); FAB-MS: [M+H] =* 575.45. E&ample -5 cyclo- (D-Leu-NMeArg-Gly-Asp-Mamb); J » D-Leu, K » NMeArg,

L - Gly, M = Asp, R1 =» R2 = H

The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb) . The peptide was prepared on a 0.115 mmol scale to give the protected cyclic peptide (92.4 mg, 98%). The peptide (92.4 mg) and 93 μΙ, of m-cresol were treated with

O anhydrous hydrogen fluoride at 0 C for 20 minūtes. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yellow solid (45.7 mg, 63%; calculated as the acetate 35 36 salt) . Purification vas accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 7 to 21% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title 5 compound as a fluffy vhite solid (29% recovery, overall yield 16.5%);FAB-MS: (M+H) - - 589.48. £aample 13i cyclo-(D-Lys-NMeArg-Gly-Asp-Mamb); J - D-Lys, K - NMeArg,

10 L - Gly, M - Asp, R1 - R2 - H

The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-

Mamb) . The DCC/DMAP rr.ethod vas used for attachment of Boc- 15 Mamb to the oxime resin. The peptide vas prepared on a 0.586 mmol scale to give the protected cyclic peptide (349 mg, 58.9%). The peptide (334 mg) and 334 μΐ. of anisole vere 0 treated vith anhydrous hydrogen fluoride at 0 C for 30 minūtes. The crude material vas precipitated vith ether, 20 redissolved in aqueous acetonitrile, and lyophilized to generate the title compound as a pale yellow solid (168 mg, 79.1%; calculated as the difluoride salt). Purification vas accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 5.4 to 25 14.4% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy vhite solid (33.6% recovery, overall yield 12.1%); FAB-MS: (M+H) * 604.32 30 Example 18 cyclo-(NMeGly-NMeArg-Gly-Asp-Mamb); J - NMeGly, K - NMeArg, L - Gly, M - Asp, r! « R2 - H (SEQ ID NO:6) 35 The title compound vas prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp- 36 LV 10111

Mamb). The DCC/DMAP method was used for attachment of Eoc-Mamb to the cxirr.e resin. The peptide was prepared on a 0.4 3 mmcl scale tc grve the prctected cyclic peptide (205 mg, £0%). The peptide (200 rr.g) and 200 pL of m-cresol were

O treated with anhydrous hydrogen fluoride at 0 C for 30 minūtes. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate (18) as a pale yellow solid (148 mg, 97%; calculated as une acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 7 to 22% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of (18) as a fluffy white solid (14.7% recovery, overall yield 7.9%); FAB-MS: (M+H) « 547.34.

Example. 21 cyclo-(Pro-NMeArg-Gly-Asp-Mamb) ; J = Pro, K = NMeArg, L = Gly, M = Asp, R1 » R2 - H (SEQ ID NO:3)

The tītie compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-

Mamb). The DCC/DMAP method was used for attachment of Boc-

Mamb to the oxime resin. The peptide was prepared on a 0.43 mmol scale to give the protected cyclic peptide (170 mg, 48,8%). The peptide (164 mg) and 164 μΐ, of m-cresol were 0 treated with anhydrous hydrogen fluoride at 0 C for 30 minūtes. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate (24) as a pale yellow solid (101 mg, 79% ; calculated as the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 7 to 22% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of (24) as a fluffy white solid (5.8% recovery, overall yield 2.1%),‘FAB-MS: (M+H) » 573.46. 37 5 23

Ex appīs.. 2JS cycio-(C-?rc-NXeArg-3iy-Asp-Marx>) ; J = 1 = Gly, M - Asp, R1 = R2

H V * *

NMeA t 10 15 20

The title compound was prepared using.the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb) . The DCC/DMAP method was used for attachment of Boc-Mamb to the oxime resin. The peptide was prepared on a 0.43 mmol scale to give the protected cyclic peptide (211mg, 60.8%). The peptide (200 mg) and 200 μΐ, of m-cresol were

O treated with anhydrous hydrogen fluoride at 0 C for 30 minūtes. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate (25) as a pale yellow solid (145 mg, 93.3%; calculated as the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 7 to 22% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of (25) as a fluffy white solid (6.4% recovery, overall yield 3.3%); FAB-MS: [M+H] - = 573.35.

Exa:nple-,2B-C

cyclo-((J-Ala-NMeArg-Gly-Asp-Mamb) ; J = β-Ala, K ** NMeArg, 25 L * Gly, M = Asp, R1 = R2 = H (SEQ ID NO:2)

The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-30 Mamb) . The DCC/DMAP method was used for attachment of Boc-Mamb to the oxime resin. The peptide was prepared on a 0.536 mmol scale to give the protected cyclic peptide (264 mg, 57.5%). The peptide (258 mg) and 258 μΐ of anisole were treated with anhydrous hydrogen fluoride at 0 C for 30 35 minūtes. The crude material was precipitated with ether, redissolved in agueous acetonitrile, and lyophilized to 38 LV 10111 generate the title compound as a pale yellow solid (231 mg, greater than quantitative yield; calculated as the fluoride salt) . Furificaticr. was acccrrplished ty reversed-phase HP1C on a preparative Vydac C18 column (2.5 cir) using a 0.23%/ min. gradient of 5.4 to 14.4% acetonitrile containing 0.1% TFA and then lyophilizedto give the TFA salt of the title compound as a fluffy white solid (53.2% recovery, overall yield 32.5%); FAB-MS: [M+H] * 547.28

Example 29 cyclo-(Gly-Arg-Gly-Asp-Mamb); J = Gly, K = Arg, L * Gly, M » Asp, r! ** R2 - H ($EQ ID NO:5)

The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb). The peptide was prepared on a 0.283 mmol scale and half was cyclized to give the protected cyclic peptide (62 mg, 58%) . The peptide (60 mg) and 60 μΐ< of m-cresol were

O treated with anhydrous hydrogen fluoride at 0 C for 1 hour. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophili2ed to generate the title compound as a pale yellow solid (48 mg, > quantitative yield; calculated as the acetate salt) . Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.30%/ min. gradient of 0 to 9% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy white solid (36% recovery, overall yield 19.9%); FAB-MS: [M+H] - 519.26. £xa.T.ple.. cyclo-(D-Ala-Arg-Gly-Asp-Mamb); J = D-Ala, K = Arg,

L - Gly, M = Asp, R1 = R2 = H

The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp- 39 40

Mamb). The peptide was prepared on a 0.189 mmol scale to give the protected cyclic peptide (211 mg, >quantitative yield) . The peptide (135 mg) and 195 μΐ cf m-crescl vere 0 treated with anhydrous hydrogen fluoride at 0 C for 1 hour. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yellow solid (125 mg, 83%; calculated as the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 2 to 11% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy white solid (12.5% recovery, overall yield 13.8%); FAB-MS: [M+H] * 533.26.

Eaample.31 cyclo-(Ala-Arg-Gly-Asp-Mamb); J * Ala, K = Arg,

L = Gly, M =* Asp, R1 = R2 » H

The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb). The peptide was prepared on a 0.324 mmol scale to give the protected cyclic peptide (191 mg, 76.4%). The peptide (100 mg) and 100 mL of m-cresol were treated with

O anhydrous hydrogen fluoride at 0 C for 1 hour. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yellow solid (75 mg, 97.4%; calculated as the acetate salt) . Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 2 to 11% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy white solid (15.5% recovery, overall yield 10.5%); FAB-MS: (M+H] * 533.25. 40 1 * LV 10111 "y^rpl^ ?? Z jf C - 0 " ( v 5 - * ·“ Γ 3 ~ “ ."ī w " /.-3 ΓΤ-£·},' ^ = C"»a./ Λ = “·* ^ i L = Gly, M * Asp, R1 = R2 » H 5

The title compound vas prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Glv-Asp-

Mamb). The peptide vas prepared on a 0.193 mmol scale to give the protected cvciic ceptide (193 mg, > ouantitative 10 yield) . The peptide (193 mg) and 193 μΐ of m-cresol were 0 treated with anhydrous hvdrcgen fluoride .at 0 C for 1 hour. The crude ir.aterial vas precipitated with ether, redissolved in aqueous HCAc, and lyophili:ed to ger.erate the title compound as a pale yeilow solid (130 mg, 86%; calculated as 15 the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 2 to 13% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy white solid (57% 20 recovery, overall yield 58.1%); FAB-MS: [M+H] * 561.22.

Fvample 33

cyclc-(D-Leu-Arg-Gly-Asp-Mamb); J = C-Leu, K * Arg, L = Gly, M - Asp, R1 = R2 = H 25

The title compound vas prepared using the general procedure described abcve for cvclo-(C-Vai-NMeArg-Gly-Asp·

Mamb). The peptide vas prepared on a C.2C2 mmol scale to give the protected cvciic peptide (152 mg, 93%). The peptide (150 mg) and 150 μΐ of m-cresol were treated vith ar.hydrous hydrogen fluoride at c'c for 1 hour. The crude material was precipitated vith ether, redissolved in aqueous KOAc, and lvophilized to generate the title compound as a pale yellow solid (78 mg, 66%; calculated as the acetate salt) . Purification.vas accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. 41 35

gradient of 5 tc 18». acetcr.itrile containing 0.1% trif 1 ucrcacetic acid ar.d then lycp'r.ili:ed to give the TFA recovery, overall yield 14.8%); FA5-MS: [M+K] = 575.45.

cyclo- (D-Abu-Arg-Gly-Asp-Mamb) ; J = D-Abu, K * Arg, L = Giy, M = Asp, R1 = R2 - H

The title ccmpcund was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp-Mamb). The peptice was prepared on a 0.193 mmol scale to give the protected cyclic peptice (210 mg, > quantitative yield) . The peptide (206 mg) and 206 μΐ, of m-cresol were

O treated with ar.hydrous hydrogen fluoride at 0 C for 1 hour. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a paie yeliow solid (158 mg, 99%; calculated as the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.22%/ min. gradient of 2 to 11% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA sa of the title ccmpcund as a fluffv white solid (57% reccverv overall yield 72.2%); FA2-MS: [M+H] * 547.21.

cyclo-C

The title ccr.pour.d vas prepared using the ger.eral procedure described above for cycic-(T-Val-N’XeArg-Gly-Asp-Mamb) . The peptide vas prepared on a 0.193 mmol scale to give the protected cyclic peptide (224 mg, > quantitative yield) . The peptide (210 mg) and 210 μΐ of m-cresol were

O treated with anhydrous hydrogen fluoride at 0 C for 1 hour. The crude material vas precipitated with ether, redissolved 42 LV 10111 in agueous HCAc, and 1 yc c h: 1 i: e c ts ger.erate the title ccmpound as a pale yellcw solid (145 mg, 834; calculated 'Γ.τ iCecetr i 2 -1; . i - - - Γ . Γ ž ' 1 ^ w.** a t Otrt 1 1« i z 7 reversed-phase HPLC cr. a preparative Vydac C18 cclumr. (2.š cm) using a 0.23i/ min. gradient of 2 to 13i acetcnitrile cor.taining 0.1% TFA and then iycphilized to give the TFA sal-of the title compound as a fluffy white solid (22% recovery overall yield 27%); FA3-MS: [M+H] = 549.31. 10 Er.arsle 3 5 “ Arg, L * Gly,

cvclo-(D-?he-Arg-Gly-Asp-Mamb) ; J D-Phe, K

M * Asp, R1 = R2 = H

The title compound was prepared using the general 15 procedure described above for cyclo-(D-Val-N’MeArg-Gly-Asp-Mamb). The peptide was prepared on a 0.266 mmol scale to give the protected cyclic peptide (202 mg, 90%). The peptide (157 mg) and 157 mL of m-cresol vere treated with anhvdrous

O hydrogen fluoride at 0 C for 1 hour. The crude material was 20 precipitated with ether, redissolved in aqueous HOAc, and lyophilized to generate the title compound as a pale yelicw solid (125 mg, > guantitative yield; calculated. as the acetate salt). Purification was accompiished by reversed-phase HPLC on a preparative Vvcac C18 coiumn (2.5 cm) using a 25 0.23%/ min. gradient of 7 to 23% acetonitrile containir.g 0.1% TFA and then Ivophilized to give the TFA salt of the title compound as a fluffv wr.ite solid (35% reccvery, overall yie 1 d 2 9.3%); FA3-MS: [M^H] = 609.25 30 Zzzzzlz ,32 cyclo-(?he-Arg-Gly-Asp-Xamii) ; J = Phe, K * Arg, L = Gly, M = Asp, R- = R2 = H (SEQ ID NC:4) 35 The title compound was prepared using the general procedure described above for cyclo-(D-Val-NMeArg-Gly-Asp- 43 ‘Ι *1

Mamb) . The peptide was prepared on a 0.335 mmoi scale to cive the prctected cvclic peptide {306 mg, > auar.titattve 10 15 treated with. ar.hydrous hvdrcgen iluoride at 0 C for 1 hcur. The crude material was precipitated with ether, redissolved in aqueous HOAc, and lyophiiized to ger.erate the title compound as a pale yellow solid (214 mg, 98%; calculated as the acetate salt). Purification was accomplished reversed-phase HPLC on a preparative Vvdac C1S column {2.5 cm) using a 0.23%/ min. gradient of S to 23% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of the title compound as a fluffy white solid (32% recovery, overall yield 31.5%); FAB-MS: [M+H] = 609.26

Example 48a

The title compound may be synthesized using procedures described in Mosher et al. Tett. Lett. 29: 3183-3186, and as shown schematically below. This same procedure is a 20 generally useful method for convertir.g a primary amine ir.to a guanidine functionality.

44 LV 10111 42-45 he synthesis cf E.\arples <1 2-<35 is shcw.n scherr.aticaliy 5 below, .SH (CHzJn

N H ΖΛ baso ,S(CHj)2NH2 (CHJn

se« jynthesij of compound 41

,S(CH2)2NH

HNH f NH2 Θ • H S 03® 10

Exa.?.pies 46 and 47 are prepared according to Standard prccedures, for exan'.pie, as described in Garigipati, Tetz. Lett, (1990) 31: 1969-1972 and in Canadian Patent 2008311, as is shown scheir.atically below. The aspartic acid group may be 15 protected (e.g., with a phenacyl protection group) to avoid side reactions. 45 ΝΗ

ļ ΝΗ

ι

{CH2)„

t) MeAI(CI)NR'R" Toluene, 80°C R' = R" = H 2) HjO η = 0, 1 OR 1} KjS/Pyr/EtjN 2) Mel/aceton# 3) Amm. acetat·

Eaarcple 5i cyclo-(D-Val-N’MeArg-P-Ala-Asp-Mamb) ; J * D-Val, K = NMeArg,

L = β-Ala, M = Asp, R1 = R2 - H

The title compound was prepared using the general procedure described above for cyclo- (D-*Val-NMeArg-Gly-Asp-

Mamb) . The DCC/DMA? method was used for ar.tachr.ent of Boc-

Mamb to the oxime resin. The peptide was prepared on a 0.555 mmol scale to give the protected cyclic peptide (227 mg, 46.9¾). The peptide (219 mg) and 219 μι cf anisole were 0 treated wiih ar.hvcrcus hvdrocer. fluoride at C C ror 30 minūtes. The crude rr.aterial was crecipitated with erher, redissolved in agueous acetonitrile, and iyophiiized to ger.erate (54) as a pale ye!lcw solid (150 mg, 93.2%/ calcuiated as the fluoride salt) . Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of T.2 to 16.2% acetonitrile containing 0.1% TFA and then lyophilized to give the TFA salt of (54) as a fluffy white solid (43.6% recovery, overall yield 16.5%); FAB-MS: [M+H] 589.32. 46

The syr.tr.esis o: -y.ar.c.‘,

C V? 5 3 LV 10111 y 5 belcw.

1) 25% TFA in OCM

2) 10% DIEA in OCM BOC'Asp*Mamb*oxime -—

3) Br(CH2)nCOOH n s 1,2 DCC 0

Mamb-oxime

Br(CH2)n

TOS-NH 10

y=NH

•NH

BOC—Ņ^ļj^ N'(CH2)i^Mamb-oxime Me o contlnue a$ in ganeral sch«m· 55, 56, 57, and 58 47 ► V a ~ ** ***» > A 0? Ci - Mfc , t

The title compound vas prepared using the general procedure described above for cyclo- (D-Val-NMeArg-Gly-Asp-Mamb) . The DCC/DMAP method was used for attachment of Bcc-4-aminomethylbenzoic acid to the oxime resin. The peptide was prepared on a 0.43 mmol scale to give the protected cyclic peptide (2l2mg, 60.8%). The peptide (200 mg) and 200 μΐ of m-cresol were treated with anhydrous hydrogen fluoride at C°C for 30 minūtes. The crude material was precipitated with ether, redissolved ir. agueous HOAc, and lyophilized to generate the crude peptide as a pale yellow soiid (152 mg, 97% ; calculated as the acetate salt). Purification was accomplished by reversed-phase HPLC on a preparative Vydac C18 column (2.5 cm) using a 0.23%/ min. gradient of 7 to 22% acetonitrile containing 0.1% TFA. Two peaks were isolated to give isomer #1 (87) (17.1% recovery, overall yield 9.3%) and isomer #2 (88) (13.4% recovery, overall yield 7.3%); FAB-MS: [M+H] = 575.41 (isomer #1; 87); 575.44 (isomer #2; 88) telet , as i r 1 may

The compounds of this ir.ver.tion possess artiplatele efficacy, as evider.ced by the ir activicy ir. Standard pie aggregation assays or platelet fibrir.cgen binding assays described below. A compound is ccnsidered to te active trese assays if it has ar. 2C*q value c-f less than abcut Platelet aggregation and fibrinogen bir.di.ng assāys which used to demonstrāte the antiplatelet activity of the compounds of the invention are described below.

Platelet Aggreoation Assay: Venous blood was obtained from the arm of a healthy human donor who was drug-free and 48 LV 10111 aspirin-free for at least :wo veeks prior to blood collection. Blood vas ccllected ir.to 10 ml citratea Vscutair.er tūces. The tlccd vas centrifūgai for 1: -mutes at 150 x g at rcom temperature, ar.d platelet-rich piasm.a (PRP) vas removed, The remaining blood was cer.trifuged for 15 minūtes at 1500 x g at-room temperature, and platelet-poor plasma (PPP) was removed. Samples were assayed on a aggregometer (PAP-4 Platelet Aggregation Profiler), using PPP as the blank (100* transmittance) . 200 μΐ of PRP vas added 10 to each micro tēst tube, and transmittance was set to 0¾. 20 15 μΐ of various agonists (ADP, collagen, arachidonate, epinephrine, thrombin) were added to each tube, and the aggregation profilēs were plotted (% transmittance versus time) . The results were expressed as % inhibition of agonist-induced platelet aggregation. For the IC50 evaluation, the tēst compounds were added at various concentrations prior to the activation of the platelets. 20 25

Platelet-Γibrinocen. Bindina Assav: Binding of 125I-fibrinogen to platelets was performed as described by Bennett et al. (1983) Proc. Nati. Acad. Sci'. USA 80: 2417-2422, vith some modifications as described below. Human PRP (h-PR?) was applied to a Sepharose column for the purification of platelet fractions. Aliguots of platelets (5 X 108 celis) aiong with 1 mM caicium chloride vere added to removable 96 well plates pricr to the activaticr. of the human gel purified platelets (h-GP?). Activation of the human gel purified platelets was achieved using ATP, collagen, arachidonate, epinephrine, and/cr thrombin in the presence of the ligar.d, 125χ_£ibrinogen. The ^”l-fibrir.cgen bound to the activated, platelets vas separated from the free form by centrifugation and then ccunted on a gamma counter. For an IC50 evaluation, the tēst compounds were added at various concentrations prior to the activation of the platelets. 49 35 50

Cosaqe....and rormulat ion

The ccrr.pour.ds of this rnvention can be administered by any means that producēs contact of the active aģent with the agent’s site of action, glycoprotein Ilb/IIIa (GPIIb/IIIa), in the body of a mammai. They can be administered by any conventional means available for use in conjunction with pharmaceuticals, either as individual therapeutic aģents or in a combination of therapeutic aģents, such as a second antiplatelet aģent such as aspirin or ticlopidine vhich are agonist-specific. They can be administered alone, but generally administered with a pharmaceutical carrier selected on the basis of the chosen route of administration and Standard pharmaceutical practice.

The dosage administered will, of course, vary depending upon knovn factors, such as the pharmacodynamic characteristics of the particular aģent and its mode and route of administration; the age, health and veight of the recipient; the nature and extent of the symptoms; the kind of concurrent treatment; the frequency of treatment; and the effect desired. A daily dosage of active ingredient can be expected to be about 0.01 to 10 milligrams per kilogram of body veight.

Dosage forms (compositions suitable for administration contain from about 1 milligram to about 100 milligrams of active ingredient per unit. In these pharmaceutical compositions the active ingredient will ordinarily be present in an amount of about 0.5-95% by veight based on the total veight of the composition.

The active ingredient can be administered orally in solid dosage forms, such as capsules, tablets, and povders, or in liquid dosage forms, such as elixirs, syrups, and suspensions. It can also be administered parenterally, in sterile liquid dosage forms.

Gelatin capsules contain the active ingredient and povdered carriers, such as lactose, starch, cellulose 50 51 LV 10111 derivatives, magnesium stearate, stearic acid, and the like. Similar diluents can be used to make compressed tablets.

Both tablets and capsules ca.n be manufactured as sustai.ned release products to provide for continuous release of medication over a period of hours. Compressed tablets can be sugar coated or film coated to mask any unpleasant taste and protect the tablet from the atmosphere, or enteric coated for selective disintegration in the gastrointestinal tract.

Liquid dosage forms for oral administration can contain coloring and flavoring to increase patient acceptance.

In general, water, a suitable oil, saline, aqueous dextrose (glucose), and related sugar Solutions and glycols such as propylene glycol or polyethylene glycols are suitable carriers for parenteral Solutions. Solutions for parenteral administration preferably contain a water soluble salt of the active ingredient, suitable stabilizing aģents, and if necessary, buffer substances. Antioxidizing aģents such as sodium bisulfite, sodium sulfite, or ascorbic acid, either alone or combined, are suitable stabilizing aģents. Also used are citric acid and its salts and sodium EDTA. In addition, parenteral Solutions can contain preservatives, such as benzalkonium chloride, methyl- or propyl-paraben, and chlorobutanol.

Suitable pharmaceutical carriers are described in Remington’s Pharmaceutical Sciences. Mack Publishing Company, a Standard reference text in this field.

Useful pharmaceutical dosage-forms for administration of the compounds of this invention can be illustrated as follovs:

Capaalea A large number of unit capsules are prepared by filling Standard two-piece hard gelatin capsules each with 100 milligrams of powdered active ingredient, 150 milligrams of lactose, 50 milligrams of cellulose, and 6 milligrams magnesium stearate. 51 52

Soft.Gelatin Capsnles A rr.ixture o: active ir.credier.t ir. a digestafcle cil suc!r. as soybean oil, cottonseed oil or olive oil is prepared and injected by means of a positive displacement pump into gelatin to form soft gelatin capsules containing 100 milligrams of the active ingredient. The capsules are vashed and dried. 10 lablets A large number of tablets are prepared by conventional procedures so that the dosage unit was 100 milligrams of active ingredient, 0.2 milligrams of colloidal Silicon dioxide, 5 milligrams of magnesium stearate, 275 milligrams 15 of microcrystalline cellulose, 11 milligrams of starch and 98.8 milligrams of lactose. Appropriate coatings may be applied to increase palatabilļty or delay absorption.

Tables 1-7 below set forth representative compounds of 20 the present invention. 52 LV 10111

Table 1 s Gly s, N’MeArg "S^ / \

J NH \ °=c J R4 0 1 H -N-C-C- 1 1 J= R3 R5 The optical isomer of J is indicated. E&ample β3 No. E4 E5 fAEL-JMS IM±ŪI Ogljgal laumai 1 H H H 533.26 2 H H CH3 547.23 (D) 2a H H CH3 (L) 3 H H CH2CH3 561.46 (D) 3a H H CH2CH3 (L) 4 H H CH (CH3) 2 575.45 (D) 4a H H CH(CH 3)2 (L) 5 H H CH2CH(CH3)2 589.48 (D) 5a H H CH2CH(CH3)2 (L) 6 H H CH2CH2CH3 (D) 6a H H CH2CH2CH3 (L) 7 H H CH2CH2CH2CH3 (D) 7a H H CH2CH2CH2CH3 (L) 8 H H (CH2)5CH3 (D) 8a H H (CH2)5CH3 (L) 9 H H (CH2)7CH3 (D) 9a H H (CH2)7CH3 (L) 53 54

Table 1 (continuedl

Example £3 E4 E5 EAEzilE Optical 10 H H C(CH 3)3 -ΙΜ·*·Η)_ Isopgr <D) 10a H H C(CH3)3 (L) 11 H H phenyl (D) lla H H phenyl (L) 12 H H phenylmethyl (D) 12a H H phenylmethyl (L) 13 H H CH20H (D) 13a H H CH2OH (L) 13b H H (CH2)3NH2 (D) 13c H H (CH2)3NH2 (L) 13d H H (CH2)3NHC{=NH)NH2 (D) 13e H H (CH2)3NHC(=NH)NH2 (L) 13f H H (CH2)4NH2 604.32 (D) I3g H H (CH2)4NH2 (L) 13h H H (CH2)4NHC(-NH)NH2 (D) 13i H H (CH2)4NHC(=NH)NH2 <L) 13 j H H (CH2)5NH2 (D) I3k H H (CH2)5NH2 (L) 131 H H (CH2)5NHC(=NH)NH2 (D) 13m H H (CH2)5NHC(»NH)NH2 (L) 13n H H (CH2)4CH3 (D) 13o H H (CH2)4CH3 (L) 13p H H (CH2)6CH3 (D) 54 55LV 10111

Table 1 (rnnf i m:pd) F v a *r. * <=> - 7 = 4 *5 rīc.vc ^ r*v *· No. 13q L· JU £k (M+H) īso H H <CH2)6CH3 (L) 13r H H CH (CH3)CH2CH3 (D) 13s H H CH(CH3)CH2CH3 (L) 14 H H CH2SH (D) 14a H H CH2SH (L) 15 H H CH2OCH3 (D) 15a H H CH2OCH3 (L) 16 H H CH2SCH3 (D) 16a H H CH2SCH3 <L) 17 H H CH2CH2SCH3 (D) 17a H H CH2CH2SCH3 (L) 18 CH3 H H 547.34 19 H CH3 CH3 20 H CH2CH3 CH2CH3 21 H H cyclopentyl (D) 21a H . H cyclopentyl (L) 22 H H cyclohexyl (D) 22a H H cyclohexyl (L) 23 H H cyclohexylmethyl (D) 55 56 Table 1 (continuedl . Gly.N NMeArg As^

J NH

Example jIKa. 24 o FAB-MS (M+S). ORticfll ISPPigC 573.46 (L) 573.35 (D) 25 26

-NH _ A 27 0 28

S (D) 28a 56 57LV 10111

Xable-l -fcor.t inued) Es.ample J FA3-MS fM+S) Odcica1 Ha. -Lscrce E " 28b 1 ^sv . (L) 28c -NHCH2CH2C(=0)- 547.28 28d 0 (D) 28e 0 (L) 57 58

TaJble 2 58

R4 0

I II

-N-C-C-I I H RS

The optical isomer of J is indicated. E^ampla £4 NO. fi5 FAB-MS (M+H) Optical Isomaj: 29 H H 519.26 30 H CH3 533.26 (D) 31 H CH3 533.25 (L) 32 H CH(CH3) 2 561.22 (D) 32a H CH(CH3)2 (L) 33 H CH2CH(CH3)2 575.45 (D) 33a H CH2CH(CH3)2 (L) 34 H CH2CH3 547.21 (D) 34a H CH2CH3 (L) 35 H CH20H 549.31 (D) 35a H CH2OH <L) 36 H phenylmethyl 609.25 (D) 37 H phenylmethyl 609.26 (L) 58 59 LV 10111 y Giy s ķ ' Asp / \

Table 3 D-Val \ NH 0=0 K = -N(R6)CH(R7)C(=0)- The optical isomer of K is indicated. dampis £6 Να. £7 FAB-MS iM+.H), Optical Isomer 32 H -(CH2)3NHC(=NH)(NH2) 561.22 (L) 4 ch3 -(CH2)3NHC(=NH)(NH2) 575.45 (L) 38 CH3 -(CH2)4NHC(=NH)(ŅH2) (L) 39 H - ch2 ch2nh2 (L) 40 CH3 - ch2 ch2nh2 (L) 41 CH3 "ch,*^3-ch,nh_<“h* (L) 42 CH3 -CH2SCH2CH2NH2 (L) 43 CH3 -CH2SCH2CH2NHC(=NH)(NH2) (L) 44 CH3 -CH2CH2SCH2CH2NH2 (L) 45 CH3 -CH2CH2SCH2CH2NHC(=NH)(NH2) (L) 46 CH3 NH (L) 47 CH3 -“'-OC (L) (L) 59 60

60 48 CH3 — CH2 ^-CHjNH; Table 3 (continued) Lsample Na. R7 FAB-MS JM+H) 48a CH3 -C_Q«.c£ 4 9 CH3 Example Number R £ΑΒζΜ2_ΙΜ±ϋ1. 52 -NK s 0 53 , .NH (L)

Qpt ica1 Isomer (L) (L) Ο 60 61LV 10111

Table 4

/L N

NMeArg / \ D-Val NH

£xample L FAB- Ua. 4 -NHCH2C(=0)- 575 54 -NHCH2CH2C(»0)- 589 55 -OCH2C(=0)- 56 -OCH2CH2C(=0)- 57 -SCH2C (=0)- 58 -SCH2CH2C(=0)- MŽ 1M+H1 45 32 61 62labie ž / Giy v.

NMeArg M / \

D-Val \ NH M Example Numbvr B8 4 -CH2C00H 63 -CH2C00H 64 -CH(CH3)COOH 66 -CH2SO3H NHC(R8) (R9)C(«0) - £9 FAB-MS m-t-Hl· H 575.45

CH3 H H 62 63 LV 10111

D-Val N· -R2 R1 The optical isomer of -CH (R^) N (R^) - is indicated. Exampls E1 E2 FAB- Optical Να. MS (M+H-L 4 H H 575.45 68 CH3 H R 68a CH3 H S 69 CH2CH3 H R 69a CH2CH3 H S 70 CH(CH3)2 H R 70a CH(CH3)2 H S 71 CH2 CH2 CH3 H R 71a CH2 C.H2 CH3 H S 72 CH2CH2CH2CH3 H R 72a CH2CH2CH2CH3 H S 73 C(CH3)3 H R 73a C(CH3)3 H S 74 CH(CH3)CH2CH3 H R 74a CH(CH3)CH2CH3 H S 75 benzyl H R 75a benzyl H S

Table-fi yGiyv NMeArg Sp / \ 63 64

Tahle.6 /ronr. inued) T v a rr-,τ*) ia B1 £2 BAB- Qc:iica,l MS (M+H). Jsoīner. 76 phenyl • H R 7 6a phenyl H S 77 cycļopentyl H R 77a cyclopentyl H S 78 cyclohexyl H R 78a cyclohexyl H 79 H CH3 80 H CH2CH3 81 H CH2CH2CH3 82 H CH(CH3)2 83 H CH2CH2CH2CH3 84 H C(CH3)3 · 85 H CH(CH3)CH2CH3 86 H benzyl 64 65 LV 10111 labie .1

Eaarcplg Structura

EĀB-MS 88 89

575.41 isomer 1 isomer 2

575.44 65 66 SEOUENCE USTINfi (1) GENERAL INFORMATION: (i) APPLICANT: William Frank DeGrado, Sharon Ar.ne

Jackson, Shaker Ahmed Mousa (ii) TITLE OF INVENTION: Cyclic Peptides Containing

Aminomethylbenzoic Acid Useful as Inhibitors of Platelet Glycoprotein Ilb/IIIa NUMBER OF SEQUENCES: 6 CORRESPONDENCE ADDRESS: (A) ADDRESSEE: The Du Pont Merck Pharmaceutical Company (B) STREET: 1007 Marķēt Street, Legal Department (C) CITY: Wilmington (D) STATE: DE (E) C0UNTRY: U.S. (F) ZIP: 19898 (iii) (v) COMPUTER READABLE FORM: (A) MEDIUM ΤΥΡΕ: 3.50 inch disk (B) COMPUTER: Apple Macintosh (C) OPERATING SīSTEM: Apple Macintosh (D) SOFTWARE: Microsoft Word (vi) CURRENT APPLICATION DATA: (A) APPLICATION NUMBER: unknovm (B) FILING DATĒ: unknown (C) CLASSIFICATION: unknown (vii) PRIOR APPLICATION DATA: none (viii) ATTORNEī/AĢENT INFORMATION: (A) NAME: Ferguson, Blair, Q. (B) REGISTRATION NUMBER: 34,329 (C) REFERENCE/DOCKET NUMBER: BP-6543 (ix) TELECOMMUNICATION INFORMATION: (A) TELEPHONE: 302-892-1676 (B) TELEFAX: 302-892-7283 66 67 LV 10111 (2) INFORMATION FOR SEQ ID NO:1: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 4 ar.ino acids (B) ΤΥΡΕ: amino acid (C) TOPOLOGĪ: circular (ii) MOLECULAR ΤΥΡΕ: peptide (vi) ORIGINAL SOURCE: synthetic (ix) FEATURE: (D) OTHER INFORMATION: Example Number 1; GPIIb/IIIa inhibitor (xi) SEQUENCE DESCRIPTION: SEQ ID NO:l

Gly MeArg GIy Asp Xaa 1 67 68 (2) INFORMATION FOR SEQ ID NO:2: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 4 amino acids (B) ΤΥΡΕ: amino acid (C) TOPOLOGī: circular (ii) MOLECULAR ΤΥΡΕ: peptide (vi) ORIGINAL SOURCE: synthetic (ix) FEATURE: (D) OTHER INFORMATION: Example Number 28c; GPIIb/IIIa inhibitor (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2 bAla MeArg Gly Asp Xaa 1 68 69 LV 10111 (2) INFORMATION FOR SEQ ID NO:3: (i) SEQUENCE CHARACTERISTICS: IA) LENGTH: 4 amir.o acids (B) ΤΥΡΕ: amino acid (C) TOPOLOGY: circular MOLECULAR ΤΥΡΕ: peptide (vi) ORIGINAL SOURCE: synthetic (ix) FEATURE: (D) OTHER INFORMATION: Example Number 24/ GPIIb/IIIa inhibitor (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3

Pro MeArg Gly Asp Xaa 1 69 70 (2) INFORMATION FOR SEQ ID NO:4: (i) SEQUENCE CHARACTERISTICS: (A) LENGTH: 4 a.T.ir.o acids (B) ΤΥΡΕ: amino acid (C) TOPOLOGY: circular MOLECULAR ΤΥΡΕ: peptide (vi) ORIGINAL SOURCE: synthetic (ix) FEATURE: (D) OTHER INFORMATION: Example Number 37; GPIIb/IIIa inhibitor (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4

Phe Arg Gly Asp Xaa 1 70 71 LV 10111 (2) INFORMATION FOR SEQ ID NO: 5: (i) SEQUENCE CHARACTERISTICS : (A) LEN'GTH: 4 amino acibs (B) ΤΥΡΕ: amino acid (C) TOPOLOGY: circular (ii) MOLECULAR ΤΥΡΕ: peptide (vi) ORIGINAL SOURCE: synthetic (ix) FEATURE: (D) OTHER INFORMATION: Example Number 29; GPIIb/IIIa inhibitor (xi) SEQUENCE DESCRIPTION: SEQ ID NO:5

Gly Arg Gly Asp Xaa 1 71 72 (2) INFORMATION FOR SEQ ID NO:6: (i) SEQUENCE CHAP.ACTERISTICS: (A) LENGTH: 4 arr.ino aeids (B) ΤΥΡΕ: amino acid (C) TOPOLOGī: circular (ii) MOLECULAR ΤΥΡΕ: peptide (vi) ORIGINAL SOURCE: synthetic (ix) FEATURE: (D) OTHER INFORMATION: Example Number 18; GPIIb/IIIa inhibitor (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6

MeGly MeArg Gly Asp Xaa 1 72

5 73 CLAIMS LV 10111 WHAT IS CLAIMED IS: 1. A compound of the formula:

or a pharmaceutically acceptable salt or prodrug form thereof 10 wherein: R1 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(C2“ C4)alkyl; R2 is H or Ci-Ce alkyl; 15 R3^ is H, Ci-Ce alkyl, phenyl, halogen, or C1-C4 alkoxy;

J 20 is β-Ala·or an L-isomer or D-isomer amino acid of structure -N(R3)C(R4) <r5)c(=0)wherein: R3 is H or CH3; R4 is H or C1-C3 alkyl; R^ is H, Ci*C8 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkylmethyl, C1-C6 cycloalkylethyl, phenyl, phenylmethyl, CH2OH, CH2SH, CH2OCH3, CH2SCH3, CH2CH2SCH3, (CH2)3X where X is -NH2 or -NHC(-NH)(NH2) and s - 3-5; or R3 and R3 can be taken together to form -(CH2)t“ (t * 73 25 7 4 2-4) or -CH2SC{CH3)2~;or and r5 can be taken together to form -(CH2)u-/ where u = 2-5; is an L-isomer amino acid of structure -N(R6)CH(R7)C{=0)-, wherein: R8 is H or Ci-Ce alkyl; R7 is*-(CH2)pNHC(=NH) (NH2) , where p * 3-5;

-(CH2)r*fwhere r = 4-6;

-(CH2)ms(CH2)2Xr where m = 1,2; and X is -NH2 or -NHC(=NH)(NH2), provided that X cannot be -NH2 when r 4; or

(CH2) nX

I R8 and R7 are taken together to form -CH2CHCH2-^ where n - 0,1 and X is -NHC(=NH)(NH2); is -Y(CH2)vC(=0)-, wherein: Y is NH, 0, or S; and v = 1,2; is an L-isomer amino acid of structure -NH-CH(R8)C(=0)-, vherein: R8 is -CH2CO2H,-CH2SO3H, or -CH(CH3)CO2H. 74 75 LV 10111 2. A compound of Claim 1 of the formula (II) : K / NR2 \ I ,0=cY^yCHR‘ 5 wherein: R1 10 r2 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(Ci- C4)alkyl; is H or methyl; J is β-Ala or an L-isomer or D-isomer amino acid of structure -N(R3)c(R4)(R3)c(=0)wherein: 15 20 κ R3 is H or CH3; R4 is H or C1-C3 alkyl; R3 is H, C1-C8 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkylmethyl, C1-C6 cycloalkylethyl, phenyl, phenylmethyl, CH2OH, CH2SH, CH2OCH3, CH2SCH3/ CH2CH2SCH3, (CH2)3X where X is NH2 or NHC(=NH)(NH2) and s * 3-5; or R3 and R3 can be taken together to form -CH2CH2CH2-; or R4 and R3 can be taken together to form -(CH2)u"/ vhere u - 2-5; is an L-isomer amino acid of structure -N(R6)CH(R7)C(»0)-, vherein; R3 is H or C1-C8 alkyl; 75 25 76 R7 is - (CH2)pNHC(=NH) (NH2), where p = 3,4; 5

-(CH2)r*/where r * 4-6;

10 15

L - (CH2) ms (CH2) 2^·! where m = 1,2; and X is -NH2 or -NHC(=NH)(NH2), provided that X cannot be -NH2 when r = 4; or (CH2) nx R8 and R7 are taken together to form where n * 0,1 and X is -NHC (=NH) (NH2); is -Y(CH2)VC(=0)-, vherein; Y is NH, 0, or S; and v - 1,2; 2 0 M is an L-isomer amino acid of structure -NH-CH (R8) C (=0)-, wherein: R8 is -CH2CO2H,-CH2SO3H, or -CH(CH3)C02H. 25 3. A compound of Claim 2 vherein;

Rl is H, Cļ-C4 alkyl, phenyl, benzyl, or phenyl-(C2-C4)alkyl; 3 0 is H or methyl; 76 77 LV 10111 J is β-Ala or an L-isomer or D-is.omer amino acid of structure -N (R^) c (R4) (R^) C (=0)-, wherein: is H or CH3; R4 is H; R5 is H, C1-C8 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkylmethyl, C1-C6 cycloalkylethyl, phenyl, phenylmethyl, CH2OH, CH2SH, CH2OCH3, CH2S^H3, CH2CH2SCH3, (CH2)sX where X is NH2 or NHC(=NH)(NH2) and s = 3,4,5; or r3 and r5 can be taken together to form -CH2CH2CH2-; K is an L-isomer amino acid of structure -N(R6)CH(R7)C(~0)-, vherein: R^ is H or CH3 alkyl; R7 is -(CH2)pNHC(=NH)(NH2), where p = 3,4;

- (CH2) rX/ where r =» 4-6;

-(CH2)mS(CH2)2Xf where m - 1,2; and X is -NH2 or -NHC(=NH)(NH2), provided that X cannot be -NH2 when r » 4; or L is -YCH2C(=0)-, wherein: Y is NH or O; 77 78 M is an L-isomer amino acid of structure -NH-CH(R8)C(*0) wherein: R8 is -CH2CO2H or -CH2SO3H. 5 4. A compound of Claim 3 wherein: R1 is H; 10 R2 is H; J is β-Ala or an L-isomer or D-isomer amino acid of formula -N<R3)CH(R5)C(=0)-, wherein: 15 R3 is H and R5 is H, CH3, CH2CH3, CH(CH3)2/ CH(CH3)CH2CH3, CH2CH {CH3)2, (CH2)4NH2; or R3 is CH3 and R5 is H; or 20 R3 and R3 can be taken together to form -CH2CH2CH2 K is an L-isomer amino acid of formula -N(CH3)CH(R7)C(=0)-, vherein: 25 R7 is ·-(CH2) 3NHC (=NH) (NH2) / L is -NHCH2C (=0)-; and M is an L-isomer amino acid of formula 30 -NHCH(CH2C00H)C(=0)-. 5. A compound of Claim 1 of the formula (III): 78 79 LV 10111

(III) vherein: 5 R1 is H, C1-C4 alkyl, phenyl, benzyl, or phenyl-(Ci-C4)alkyl; R2 is H or methyl; J is β-Ala or an L-isomer or D-isomer amino acid of 10 structure -N <R3)C (R4) (R5) C (=0)-, wherein: R3 is H or CH3; R4 is H or C1-C3 alkyl; r5 is H, C1-C8 alkyl, C3-C6 cycloalkyl, C3-C6 15 20 cycloalkylmethyl, C1-C6 cycloalkylethyl, phenyl, phenylmethyl, CH2OH/ CH2SH, CH20CH3, CH2SCH3, CH2CH2SCH3, (CH2)sX where X is NH2 or NHC(*NH)(NH2) and s * 3-5; or R3 and R5 can be taken together to form -CH2CH2CH2-; or R4 and R5 can be taken together to form -(CH2)u-> where u = 2-5; K is an L-isomer amino acid of structure -N(R6)CH<R7)C(=0)-, wherein: r6 is H or C1-C8 alkyl; R7 is' -(CH2)pNHC(-NH) (NH2)» where p - 3,4; 79 25 80

-(CH2)rXfVke*e r “ — ch2 -o- ch2x^ -ch2 -CyCH2x “(CH2)mS(CH2)2x» where m - 1,2; and X is -NH2 or -NHC(*NH)(NH2) t provided that X cannot be -NH2 when r - 4; or

(CH2) nX 1 R6 and R7 are taken together to form ~CH2CHCH2", where n - 0,1 and X is -NHC(=NH)(NH2>; L is -Y(CH2)vC(=0)-/ vherein: 15 Y is NH, 0, or S; and v - 1,2; M is an l-isomer amino acid of structure -NH-CH(R8)Č(*0)-, wherein: 20 R8 is -CH2CO2H,-CH2SO3H, or -CH(CH3)C02H. 6. A compound of Claim 5 vherein; 25 R1 is H; R2 is H; J is β-Ala or an L>isomer or D-isomer amino acid of 30 formula -N(r3)CH(r5)C(-0)-, vherein: 80 81 LV 10111 R1 2 is H and R5 is H, CH3, CH2CH3, CH(CH3)2, CH(CH3)CH2CH3/· CH2CH(CH3)2, <CH2)4NH2; or R1 is CH3 and R5 is H; or 5 R1 and R1 can be taken together to form -CH2CH2CH2-. K is an L-isomer amino acid of formula -N(CH3)CH(R7)C(=0)wherein : 10 R7 is -(CH2)3NHC(=NH)(NH2); L is -NHCH2C(=0)and 15 M is an L-isomer amino acid of formula -NHCH(CH2C00H)C(=0)-. 7. The compound of Claim 2 which is selected 2 0 from the group consisting of:

the compound wherein R^ and R1 are D_val/ K is NMeArg, L is Gly, and M is Asp; 2 5 the compound vherein R1 and R1 are H, J is d-2- aminobutyric acid, K is NMeArg, L Gly, and M is Asp;

the compound vherein r! and R1 are H, J is D-Leu, K 3 0 is NMeArg, L is Gly, and M is Asp; the compound vherein R* and R1 are H, J is D-Ala, K is NMeArg, L is Gly, and M is Asp; 81 1 5 the compound vherein r!· and R1 are H, J is Gly, K is NMeArg, 2 L is Gly, and M is Asp; 32 the compound wherein R1 and R2 are H, J is D-Fro, K is NMeArg, 1 is Gly, and M is Asp,· 5 the compound vherein R* and R2 are H, J is D-Ser, K is Arg, L is Gly, and M is Asp; the compound vherein R* and R2 are H, J is D-Lys, K is NMeArg, L is Gly, and M is Asp; 10 the compound wherein R^ and R2 are H, J is d-2-aminobutyric acid, K is Arg, L is Gly, and M is Asp;

1 5 the compound wherein R* and R2 are H, J is D-Ala, K is Arg, L is Gly, and M is Asp; the compound wherein R* and R2 are H, J is D-Val, K is Arg, L is Gly, and M is Asp; 20 the compound wherein R1 and R2 are H, J is β-Ala, K is NMeArg, L is Gly, and M is Asp; the compound vherein R1 and R2 are H, J is D-Val, K is 2 5 NMeArg, L is β-Ala, and M is Asp; the compound vherein R1 and R2 are H, J is Pro, K is NMeArg, L is Gly, and M is Asp; 30

the compound vherein R* is Arg, L is Gly, and M and R2 are H, is Asp;

J is Phe, K the compound vherein R1 and R2 are H, J is Gly, K is Arg, L is Gly, and M is Asp; 82 83 LV 10111 the compound wherein R^ and R2 are H, J is NMeGly, K is NMeArg, L is Gly, and M is Asp; the compound wherein R* and R2 are H, J is D-Leu, K is Arg, L is Gly, and M is Asp. 8. The compound of Claim 2, which is selected from the group consisting of: the compound wherein R^ and R2 are H, J is D-Val, K is NMeArg, L is Gly, and M is Asp; the compound wherein R^ and R2 are H, J is d-2-aminobutyric acid, K is NMeArg, L is Gly, and M is Asp; the compound wherein R1 and R2 are H, J is D-Leu, K is NMeArg, L is Gly, and M is Asp; the compound wherein R* and R2 are H, J is D-Ala, K is NMeArg, L is Gly, and M is Asp; the compound vherein R1 and R2 are H, J is Gly, K is NMeArg, L is Gly, and' M is Asp; the compound wherein R1 and R2 are H, J is D-Pro, K is NMeArg, L is Gly, and M is Asp; the compound wherein R1 and R2 are H, J is D-Lys, K is NMeArg, L is Gly, and M is Asp; the compound wherein R1 and R2 are H, J is β-Ala, K is NMeArg, L is Gly, and M is Asp; 83 84 the compound wherein R* and are H, J is NMeGly, K is NMeArg, L is Gly, and M is Asp. 5 9. The compound of Claim 5 wherein R^ and R^ are H, J is D-Val, K is NMeArg, L is Gly, and M is Asp. 10 10. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of a compound of Claim 1. 15 11. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of a compound of Claim 2. 2 0 12. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of.,.a compound of Claim 3. 2 5 13. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of a compound of Claim 4. 3 0 14. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of a compound of Claim 5. 35 15. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of 84 85 LV 10111 such treatment a pharmaceutically effective amount of a compound of Claim 6. 16. A method for the treatment of thrombcembolic disorders which comprises administering to a host in need of such treatment a phar’maceutically effective amount of a compound of Claim 7. 17. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of a compound of Claim 8. 18. A method for the treatment of thromboembolic disorders which comprises administering to a host in need of such treatment a pharmaceutically effective amount of a compound of Claim 9. 19. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Claim 1. 20. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Claim 2. 21, A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Claim 3. 22. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically 85 86 effective platelet-inhibiting amount of a corr.pound of Ciaim 4 . 23. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceuticaliy effective platelet-inhibiting amount of a compound of Ciaim 5. · 24. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Ciaim 6. 25. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Ciaim 7. 26. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Ciaim 8. 27. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective platelet-inhibiting amount of a compound of Ciaim 9. 86

Claims (27)

EN 10111 Claims of the Formula 1 Compound of Formula (I) or a pharmaceutically acceptable salt thereof, or a prodrug thereof, wherein: 1 R is hydrogen, 1-6 alkyl, phenyl, benzyl, phenyl-C 1-6 alkyl; 2 R is hydrogen, C-alkyl; 1 'o R' is hydrogen, C, alkyl, phenyl, halo, < 1 >4-alkoxy; 1 J is a residue of the β-Ala or amino acid L- or D-isomer of the formula -N (R 3) C (R 4) (R 5) C 0-, wherein: 3 R is hydrogen or methyl; 4 R is hydrogen or C 1-4 alkyl; R 5 is hydrogen, C 1-6 -alkyl, C 1-6 -cycloalkyl, C 1-6 -cycloalkylmethyl, C 1-6 -cycloalkylethyl, phenyl, phenylmethyl, CH 2 OH, CH 2 SH, CH 2 OCH 2, CH 2 SCH 3, CH 2 CH 2 SCH 3 > (CH2) gX, wherein X is -NH2 or a group -NHC (= NH) (NH2) and s = 3 to 5; or R 5 and R 5 together form - (CH) -, where t = 2-4, or a group -CH 2 S (CH 3) 2 -; or R 5 and R 5 together form - (CH 0) where u = 2 - 5: 2 u 6 7 K is a L-amino acid residue of the formula -N (R) CH (R) C 0 -, wherein: R 6 is hydrogen atom or C 1-6 alkyl; R7 and - (CH2) pNHC (= NH) (NH2), wherein p = 3 - 5; wherein q = 0, 1; - (CH 2) r X = wherein r = 4 - 6; 2 LV 10111 L and M and 2. - (CH) S (CH) -, wherein m = 1.2; X is -NH- or -NHC (= NH) (NH 0), with 2 m 2 2 2 2, provided that X cannot be -NH 2 > that r = 4; or 6 7 R and R together form a group -C 1-4 CHCH 2 -, where η = 0, 1 and (CH) X 2 n X are -NHC (= NH) (NH 2); -Y (CH 2) v CO-, wherein Y is NH, O or S; v = 1, 2; 8 L-amino acids remain with the formula -NHCH (R) C 0-, wherein: R 8 is -CH 2 CO H, -CH 2 SOH, or -CH (CH 3) C 100 H. Compound of formula (II) according to formula (II): k-LX / NJ, i.e., -CH 2 / CHR · NR (II) 3 wherein: R 1 is hydrogen, C 1-4 alkyl, phenyl, benzyl, phenyl-C 2 - 4 -alkyl; 2 R is hydrogen or methyl; J is a residue of the β-Ala or amino acid L- or D-isomer of formula -N (R 3) C (R 4) (R 5) C 0-, wherein: R 3 is hydrogen or methyl; 4 R is hydrogen or 3 * -alkyl; R 5 is hydrogen, C 0 -C 6 alkyl, C 1-6 cycloalkyl, C 0 -C 10 cycloalkylmethyl, C 1 -Cycloalkylethyl, J-O-6 phenyl, phenylmethyl, CH 2 OH, CH 2 SH, CH 2 OCH 2, CH 2 CH 3, CH 2 CH 2 SCH 3 , (CH 2) X, wherein X is -NH 2 or a group -NHC (= NH) (NH 2) and s = 3 to 5; or R 3 and R 5 together form - (CH 2) 3 -; or 4 5 R and R both sets up -, ^ Ur u = 2 - 5; 6 7 K is a residue of the L-amino acid of formula -N (R) CH (R) C 0-, wherein: g R is hydrogen or C 1-6 alkyl; R 7 is - (CH 2 NHC (= NH) (NH 0}, where p = 3 or 4; z p z.) wherein q = 0, 1; 4 LV 10111 - (CH) -X, where r = 4 - 6; Z r CH2 ch2x9 - (CH2) mS (CH2) 2X, where m = 1.2; X is -NH2 or -NHC (= NH) (N1 ^), provided that X cannot be -NH2 > that r = 4; or 6 7 R and R together form a group -CH 2 CHCH 2 -, wherein η = 0, 1 and (OL) X z n X is -NHC (= NH) (NH 2); L is -Y (CH 2) v CO-, wherein Y is NH, O or S; v = 1, 2; g M is the residue of the L-amino acid of formula -NHCH (R) C 0-, wherein: R 8 is -CH 2 COOH, -Cl 2 SOH, or -CH (CH 3) C 100 H. A compound according to claim 2 wherein: R * is hydrogen, C 1-4 alkyl, phenyl, benzyl, phenyl C 2-4 alkyl; 2 R is hydrogen or methyl; J is a residue of the B-Ala or amino acid L- or D-isomer of formula -N (R 3) C (R 1) (R 2) C 0-, wherein: R 3 is hydrogen or methyl; - 5 1 2 R is hydrogen; are hydrogen, C, alkyl, C 1 -C 6 cycloalkyl, C 1 -C 5 cycloalkylmethyl, C-cycloalkylethyl, J 'b J' b phenyl, phenylmethyl, CH 2 OH, CH 2 SH, CH 2 OCH 2, CH 2 SCH 3, CH 2 CH 2 SCH 3, (CH 2) s X wherein X is -NH 2 or a group -NHC (= NH) (NH 2) and s = 3 to 5; or 3 5 R and R both form - (CH 2) 3 ~; 6 7 K is a residue of the L-amino acid of formula -N (R) CH (R) CO-, wherein: R 1 is hydrogen or methyl; R7 and - (CH2) pNHC (= NH) (NH2) wherein p = 3 - 4; - (CH2) -NH-C4NH2 - (CH2) / where q = 0, 1; 6 LV 10111 - (CH2) r-X, where r = 4 - 6; -CH2 ch2 ch2x " · 9 - (CH-) S (CH) - X, wherein m '1,2; X is -NH0 or -NHC (= NH) (NH), with the proviso that X cannot be " NH ^ when r = 4; L is -YCH 2 CO-, where Y is NH or 0; g M is the residue of the L-amino acid of formula -NHCH (R) CO-, wherein: R 8 is -CH 2 CO 2 H, -CH 2 SO 3. 4. A compound according to claim 3 wherein: 1 R is hydrogen; 2 R is hydrogen; J is a residue of the β-Ala or amino acid L- or D-isomer of formula -N (R 3) CH (R 1 3 3) C 0-, wherein: R 3 is hydrogen; 7 1 R is hydrogen, methyl, ethyl, isopropyl, sec-butyl, isobutyl, 4-aminobutyl, or 2 3 5 3 5 R is methyl and R is hydrogen; or both R and R together form a group -CH 2 CH 2 CH 2 -; 3 K is a residue of L-amino acids with the formula -N (CH) CH (R) CO-, wherein: o R 7 is - (CH 2) 3 NHC (= NH) (NH 2); L is -NHCH 2 CO 2 -; M is the residue of the L-amino acid of formula -NHCH (CH 2 Cl 2 H) C 0 -. Section 5, S2ym5 After Clause 1, Formula (III) (III): 1 R is hydrogen, C 1-4 alkyl, phenyl, benzyl, phenyl-C 2-4 alkyl; 2 R is hydrogen or methyl; J is a residue of the β-Ala or amino acid L- or D-isomer of formula -N (R 3) C (R 4) (R 5) CO-, wherein: 8 LV 10111 R is hydrogen or methyl; 4 R is hydrogen or C 1-4 alkyl; R 'is hydrogen, C 1-6 -alkyl, C 3-6 -cycloalkyl, C 1-6 -cycloalkylmethyl, C 3-6 -cycloalkylethyl, phenyl, phenylmethyl, CH 2 OH, CH 2 SH, CH 2 OCHg, CH 2 CH 3, CH 2 CH 2 SCH 3 > (CH 2) s X wherein X is -NH 2 or a group -NHC (= NH) (NH 2) and s = 3 to 5; or 3 5 R and R both form - (CH 2) 3 ~; or R 4 and R 5 together form - (CH 2) -, wherein u = 2-5; 6 7 K is a residue of the L-amino acid of formula -N (R) CH (R) CO-, wherein: R 1 is hydrogen or C-alkyl; * i "O R7 and - (CH2) pNHC (= NH) (NH2), wherein p = 3 - 4; - (CH;) -NH- < nnh2 - (CH2) q t where q = 0, 1; 9 9 - (CH 2) r -X, where r = 4 - 6; - (CH 2) m S (CH 2) 2 X wherein m = 1.2; X is -NH 2 or -NHC (= NH) (NH 2), provided that X cannot be -NH 2 > that r = 4; or 6 7 R and R together form a group -CR 1 CHCH 2 -, where η = 0, 1 and (CRJ X 2 n X is -NHC (= NH) (NH 2); L is -Y (CH 2) v C 0 - wherein Y is NH, S or 0; v = 1, 2; g M is the residue of the L-amino acid of formula -NHCH (R) CO-, wherein: R 8 is -CH 2 CO 2 H, -CH 2 S 0H. The compound of claim 5, wherein: R 1 is hydrogen; 2 R is hydrogen; J is the remainder of the beta-alpha or amino acid L- or D-isomer of formula -N (R 3) CH (R 5) C 0-, wherein: 3 5 R is hydrogen and R is hydrogen, methyl, ethyl, isopropyl, sec. butyl, isobutyl, 4-aminobutyl or R 10 is methyl and R is hydrogen; or 3 5 R and R both together form - (CH 2) 2 -; 7 K is the remainder of the L-amino acids with the formula -NCH 2 CH 2 R 1 CO-, wherein: R 7 is - (CH 2) 3 NHC (= NH) (NH 2); L is -NHCH 2 CO 2 -; M is the L-amino acid residue with the formula -NHCH (CH9C00H) C0-. 7. The compound of claim 2, taken from the group: is D-Val, K is NMeArg, 1 2 is a compound having R, R is hydrogen, J is Gly, M is Asp; is a compound of the D-2-amino acid moiety 1 2 having R, R is hydrogen, J K is NMeArg, L is Gly, M is Asp; Compound 1 2 wherein R, R is hydrogen, J is D-Leu, K is NMeArg, L is Gly, M is Asp; A 1 '2 compound wherein R, R is hydrogen, J is D-Ala, K is NMeArg, L is Gly, M is Asp; _12 is a compound having R, R is hydrogen, J is Gly, K is NMeArg, L is Gly, M is Asp; Compound 1 2 having R, R is hydrogen, J is D-Pro, K is NMeArg, L is Gly, M is Asp; Compound 1 2 wherein R, R is hydrogen, J is D-Ser, K is Arg, L is Gly, M is Asp; Compound 1 2 wherein R, R is hydrogen, J is D-Lys, K is NMeArg, L is Gly, M is Asp; 11 compound, compound, compound, compound, compound, compound, compound, compound, compound, compound having R, R is hydrogen, J K is Arg, L is Gly, M is Asp; 12 having R, R is hydrogen, J is Gly, M is Asp; Wherein R, R are hydrogen, J L is Gly, M is Asp; Wherein R, R and hydrogen are hydrogen, J is NMeArg, L is Gly, M is Asp; Wherein R, R is hydrogen, J L is beta-Ala, M is Asp; 12 wherein R, R is hydrogen, J L is Gly, M is Asp; 12 wherein R, R is hydrogen, J L is Gly, M is Asp; Wherein R, R are hydrogen, J L is Gly, M is Asp; 1 2 having R, R and H, J K is NMeArg, L is Gly, M is Asp; 12 having R, R is hydrogen, J is Gly, M is Asp. and D-2-amino-terminal and D-Ala, K and Arg, and D-Val, K and Arg, and beta-ala, K and D-Val, K and NMeArg, and Pro, K and NMeArg, and Phe, K and Arg, and Gly, K and Arg, and N-MeGly, and D-Leu, K and Arg, The compound of claim 2, wherein R, R is hydrogen, L is Gly, M is Asp; Compound 1 2 wherein R, R is hydrogen, J J K is NMeArg, L is Gly, M is Asp; 1 2 compound of R, R is hydrogen, J of the group is D-Val, K is NMeArg, is D-2-aminobutyric acid is D-Leu, K is NMeArg, 12 LV 10111 compound, "compound, compound, compound compound, compound, compound, NMeArg, L is L is Gly, M is Asp; 1 2 of which R, R is hydrogen, L is Gly, M is Asp; 1 2 of which R, R is hydrogen, L is Gly, M is Asp; 1 2 of which R, R is hydrogen, L is Gly, M is Asp; 1 2 of which R, R is hydrogen, L is Gly, M is Asp; 12 having R, R is hydrogen, NMeArg, L is Gly, M is Asp; 1 2 where R, R and H, J and D-Ala, K and NMeArg, J and Gly, K and NMeArg, J and D-Pro, K and NMeArg, J and D-Lys, K are NMeArg, J is Beta-Ala, K and J and N-MeGly, K and Gly, M and Asp. 12 The compound of claim 5, wherein R and R are hydrogen, J and D-Val, K are NMeArg, L is Gly, M is Asp. 10. for the treatment of thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound according to claim 1. 11. A treatment for thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound of claim 2. A method of treating thromboembolic disorders, comprising administering to a subject a pharmaceutically effective amount of a compound of claim 3. A method of treating thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound according to claim 4. 14. A method for treating thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound according to claim 5. 15. for the treatment of thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound of claim 6. 16. A method for treating thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound according to claim 7. 17. for the treatment of thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound according to claim 8. 18. For the treatment of thromboembolic disorders, comprising administering to the subject a pharmaceutically effective amount of a compound according to claim 9. 14 EN 10111 A pharmaceutical composition for inhibiting erythrocyte adhesion comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of claim 1. 20. E1C5? ^ 2 Rhodium-inhibiting inhibition of erythrocyte cleavage comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of Claim 2. 21. inhibiting erythrocyte adhesion comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of Claim 3. A pharmaceutical composition for inhibiting erythrocyte adhesion comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of Claim 4. 23. £ 2EE222i: β2? 2? 2? 22? 2? 2 for inhibiting erythrocyte adhesion comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of claim 5. A pharmaceutical composition for inhibiting erythrocyte cleavage comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of Claim 6. A pharmaceutical composition for inhibiting erythrocyte adhesion comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of Claim 7. 15 A pharmaceutical composition for inhibiting erythrocyte cleavage comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of claim 8. 27. A pharmaceutical composition for inhibiting erythrocyte adhesion comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of claim 9. 16