CA2170766A1 - Substituted di- and tripeptide inhibitors of protein:farnesyl transferase - Google Patents
Substituted di- and tripeptide inhibitors of protein:farnesyl transferaseInfo
- Publication number
- CA2170766A1 CA2170766A1 CA002170766A CA2170766A CA2170766A1 CA 2170766 A1 CA2170766 A1 CA 2170766A1 CA 002170766 A CA002170766 A CA 002170766A CA 2170766 A CA2170766 A CA 2170766A CA 2170766 A1 CA2170766 A1 CA 2170766A1
- Authority
- CA
- Canada
- Prior art keywords
- obn
- tyr
- cbz
- ser
- co2me
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/06—Dipeptides
- C07K5/06139—Dipeptides with the first amino acid being heterocyclic
- C07K5/06147—Dipeptides with the first amino acid being heterocyclic and His-amino acid; Derivatives thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
Abstract
Novel inhibitors of protein:farnesyl transferase enzyme are described, as well as methods for the preparation and pharmaceutical compositions of the same, which are useful in controlling tissue proliferative diseases, including cancer and restenosis.
Description
~1 7 ~7 6 6 PCT~S94111553 W095/12612 L11 ~1 SUBS~ ul~ DI- AND TRIPEPTIDE INHIBITORS
OF PROTEIN:FARNESYL TRANSFERASE
FIELD OF THE INVENTION
The present invention pertains to a number of compounds which can be used in the medicinal field to treat, prophylactically or otherwise, uncontrolled or abnormal proliferation of ht1mAn tissues. More specifically, the present invention pertains to a number of compounds which act to inhibit the farnesyl transferase enzyme that has been determined to activate ras proteins which in turn activate cellular divi~ion and are implicated in cancer and restenosis.
BACKGROUND OF THE INVENTION
Ras protein (or p21) has been ~x~mi n~d extensively because mutant forms are found in 20~ of most types of human cancer and greater than 50~ of colon and pancreatic carc~nnm~s (Gibbs J.B., Cell, 65:l (l99l), Cartwright T., et al., Chimica Og~i, 10:26 (1992)).
These mutant ras proteins are deficient in the capability for feedback regulation that is present in native ras and this deficiency is associated with their oncogenic action since the ability to stimulate normal cell division can not be controlled by the normal endogenous regulatory cofactors. The recent discovery that the transforming activity of mutant ras is critically dependent on post-translational modifications (Gibbs J., et al., Microbiol. Rev., ~3:171 (1989)) has unveiled an important aspect of ras function and identified novel prospects for cancer therapy.
WO95/l2612 PCT~S94/1l553
OF PROTEIN:FARNESYL TRANSFERASE
FIELD OF THE INVENTION
The present invention pertains to a number of compounds which can be used in the medicinal field to treat, prophylactically or otherwise, uncontrolled or abnormal proliferation of ht1mAn tissues. More specifically, the present invention pertains to a number of compounds which act to inhibit the farnesyl transferase enzyme that has been determined to activate ras proteins which in turn activate cellular divi~ion and are implicated in cancer and restenosis.
BACKGROUND OF THE INVENTION
Ras protein (or p21) has been ~x~mi n~d extensively because mutant forms are found in 20~ of most types of human cancer and greater than 50~ of colon and pancreatic carc~nnm~s (Gibbs J.B., Cell, 65:l (l99l), Cartwright T., et al., Chimica Og~i, 10:26 (1992)).
These mutant ras proteins are deficient in the capability for feedback regulation that is present in native ras and this deficiency is associated with their oncogenic action since the ability to stimulate normal cell division can not be controlled by the normal endogenous regulatory cofactors. The recent discovery that the transforming activity of mutant ras is critically dependent on post-translational modifications (Gibbs J., et al., Microbiol. Rev., ~3:171 (1989)) has unveiled an important aspect of ras function and identified novel prospects for cancer therapy.
WO95/l2612 PCT~S94/1l553
2-In addition to cancer, there are other conditions of uncontrolled cellular proliferation that may be related to excessive expression and/or function of native ras proteins. Post surgical vascular restenosis is such a condition. The use of various surgical revascularization techniques such as saphenous vein bypass grafting, endarterectomy and translllm; n~ 1 coronary angioplasty is often accompanied by complications due to uncontrolled growth of neointimal tissue, known as restenosis. The biochemical causes of restenosis are poorly understood and numerous growth factors and protooncogenes have been implicated (Naftilan A.J., et al., Hypertension, 13:706 (1989) and J. Clin. Invest., 83:1419; Gibbons G.H., et al., Hypertension, 14:358 (1989); Satoh T., et al., Mollec.
Cell. Biol., 13:3706 (1993)). The fact that ras proteins are known to be involved in cell division processes makes them a candidate for intervention in many situations where cells are dividing uncontrol-lably. In direct analogy to the inhibition of mutant ras related cancer, blockade of ras dependant processes has the potential to reduce or eliminate the inappropriate tissue proliferation associated with restenosis, particularly in those instances where normal ras expression and/or function is exaggerated by growth stimulatory factors.
Ras functioning is dependent upon the modification of the proteins in order to associate with the inner face of plasma membranes. Unlike other membrane-associated proteins, ras proteins lack conventionaltr~ncm~mhrane or hydrophobic sequences and are initially synthesized in a cytosol soluble form. Ras protein membrane association is triggered by a series of posttranslational processing steps that are signaled by a carboxyl term;n~l amino acid consensus sequence that is recognized by protein:farnesyl transferase.
WO 95/12612 2 1 7 ~ 7 6 ~ PCT/US9~/115~i3 .
This consensus sequence consists of a cysteine residue located four amino acids from the carboxyl term;
followed by two lipophilic amino acids and the C-t~rm;n~l residue. The sulfhydryl group of the cysteine residue is alkylated by farnesyl pyrophosphate in a reaction that is catalyzed by protein:farnesyl transferase. Following prenylation, the C-termln~1 three amino acids are cleaved by an endoproteaQe and the newly exposed alpha-carboxyl group of the prenylated cysteine is methylated by a methyl transferase. The enzymatic processing of ras proteins that begins with farnesylation enables the protein to associate with the cell membrane. Mutational analysis of oncogenic ras proteins indicate that these posttranslational modifications are essential for transforming activity. Replacement of the consensus sequence cysteine residue with other amino acids gives a ras protein that is no longer farnesylated, fails to migrate to the cell membrane and lacks the ability to stimulate cell proliferation (Hancock J.F., et al., Cell, 57:1617 (1989); Scha~er w.R., et al., Science, 245:379 (1989); Casey P.J., Proc. Natl. Acad. Sci. USA, 86:8323 (1989)).
Recently, protein:farnesyl transferases (PFTs, also referred to as farnesyl:protein transferaseQ) have been identified and a specific PFT from rat brain was purified to homogeneity (Reiss Y., et al., Bioch. Soc.
Trans., 20:487-88 (1992)). The enzyme was charac-terized as a heterodimer composed of one alpha-subunit (49 kDa) and one beta-subunit (46 kDa), both of which are required for catalytic activity. High level expression of m~mm~lian PFT in a baculovirus system and purification of the recombinant enzyme in active fonm has also been accomplished (Chen W.-J., et al., J. Biol. Chem., 268:9675 (1993)).
PCT~S9411lS53 In light of the foregoing, the discovery that the function of oncogenic ras pro.teins is critically dependent on their posttranslational processing provides a means of cancer chemotherapy through S inhibition of the processing enzymes. The identification and isolation of a protein:farnesyl transferase that catalyzes the addition of a farnesyl group to ras proteins provides a promising target for such intervention. Recently it has been determined that prototypical inhibitors of PFT can inhibit ras processing and reverse cancerous morphology in tumor cell models (Kohl N.E., et al., Science, 260:1934 (1993); James G.L., et al., Science, 260:1937 (1993);
Garcia A.M., et al., J. Biol. Chem., 268:18415 (1993)).
Thus, it is possible to prevent or delay the onset of cellular proliferation in cancers that exhibit mutant ras proteins by blocking PFT. By analogous logic, inhibition of PFT would provide a potential means for controlling cellular proliferation associated with restenosis, especially in those cases wherein the expression and/or function of native ras is overstimulated.
PCT Application WO91/16340 discloses cysteine cont~in;ng tetrapeptide inhibitors of PFT of the formula CAAX.
European Patent Application 0461869 discloses cysteine cont~;n;ng tetrapeptide inhibitors of PFT of the formula Cys-Aaa1-Aaa2-Xaa.
European Patent Application 0520823 discloses cysteine cont~;n;ng tetrapeptide inhibitors of PFT of the formula Cys-Xaal-dXaa2-Xaa3.
European Patent Application 0523873 discloses cysteine cont~;n;ng tetrapeptide inhibitors of PFT of the formula Cys-Xaa1-Xaa2-Xaa3.
WO95/12612 2 1 7 0 7 6 6 PCT~S94/11553 European Patent Application 0528486 discloses cysteine cont~; n; ng tetrapeptide amides inhibitors of PFT of the formula Cys-Xaa1-Xaa2-Xaa3-NRRl.
European Patent Application 0535730 discloses pseudotetrapeptide inhibitors of PFT of the following two formulas:
X R2 o )~_,ZH
J~NH ~H~J~H~OH
HS'~ Y R 0 X R~ o HS
European Patent Application 0535731 (US 5,238,922) discloses esters of pseudotetrapeptide inhibitors of PFT of the formula:
X R2 o ()~ZH
- H ~ -~ N oR5 ~ Y R O
HS
European Patent Application 0482539 discloses tachykinin antagonists of the formula:
R
l,Y ~ A` ~ N~R5 0 R~ O
wos5ll26l2 PCT~S9~/11553 European Patent Application 0457195 discloses endothelin antagonists of the formula:
~
US 4,022,759 discloses tripeptide antagonists of luteinizing honmone releasing factor of the formula A-Rl-Tyr(benzyl)-Ser(benzyl)-R2, wherein one of the definitions of Rl is His(benzyl).
Compounds disclosed in the above references do not disclose or suggest a novel combination of structural variations found in the present invention described hereinafter. All cited references are hereby incorporated by reference.
SUMMARY OF THE IN V~N'1'10N
Accordingly, the present invention is a substituted di- or tripeptide compound of Formula I:
R
/
~rN
N~
~( 'H2) n A--N R O
R O ~ ) n 2 1 7~766 PCT~Ss~/115~3 W095/12~1~
wherein:
n = l or 2;
A = CoR3, Co2R3, CoNHR3, CSR3, C(S)oR3, C(S)NHR3, CF3SO2, aryl-SO2, or alkyl-SO2, wherein R3 is - 5 alkyl, (CH2) m~ cycloalkyl, (CH2) m~ aryl, ( CH2 ) m~ heteroaryl, or (CH2) m ~ alkyl, and m = 0, l, 2, or 3;
R = independently H or Me;
Y - independently H or Me;
Z - independently H or Me;
Rl - H, CO-aryl, (CH2) m ~ aryl, O(CH2) m ~ cycloalkyl, O(CH2)m-aryl, or O(CH2)m-heteroaryl, wherein m is as defined above and Rl i8 located at either the meta or para position;
X = one to four substituents, including H, alkyl, CF3, F, Cl, Br, I, HO, MeO, NO2, NH2, N(Me) 2' OPO3H2, or CH2PO3H2;
R2 = NR(CH2)nCo2R3, NR(CH2)nCoNHR3, NR(CH2)nR3, NR(CH2)n+l0R , NR(CH2)n+lSR ~
NRCH(CoR5)(CH2) n ~ heteroaryl, NRCH(CoR5)(CH2)noR3, NRCH(CoR5)(CH2)nSR3, or N N- R3 wherein R, R3, and n are as defined above, R4 = H
or R3, and R5 = OH, NH2, oR3, or NHR3; an optical isomer, diastereomer, or a ph~rm~ceutically acceptable salt thereof.
The present invention is also directed to the use of a compound of Formula I, or a pharmaceutically acceptable salt there~rom, to inhibit the activity of a protein:farnesyl transferase enzyme as a method for trea~ing tissue proliferative diseases.
A further embodiment of the present invention is the use of a pharmaceutical composition including an e~ective amount of a compound of Formula I as a method for the treatment of cancer.
A still further embodiment of the present invention is the use of a pharmaceutical composition including an effective amount of a compound of Formula I as a method for the treatment of restenosis.
A still further embodiment of the present invention i8 a ph~r~ceutical compo~ition for ~mi n; stering an effective amount of a compound of Fonmula I in unit dosage form in the treatment methods mentioned above.
A final embodiment of the present invention pertains to methods for the preparation of compounds of Formula I by solid phase synthesis, solution phase synthesis, and simultaneous multiple ~yntheses using a multiple simultaneous synthesis apparatus.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMæNTS
In the compounds of Formula I, the term "alkyl"
means a straight or branched hydrocarbon radical having from 1 to 6 carbon atoms and includes, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, and the like.
The term "cycloalkyl" means a saturated hydrocarbon ring which contains from 3 to 10 carbon atoms, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, adamantyl, and the like, unsubstituted or substituted by an alkyl or aryl group.
The term "aryl" means an aromatic ring which is a phenyl, 5-fluorenyl, 1-naphthyl or 2-naphthyl group, unsubstituted or substituted by 1 to 3 substituents selected from alkyl, O-alkyl and S-alkyl, O-aryl, OH, SH, F, Cl, Br, I, CF3, NO2, NH2, NHCH3, N(CH3) 2 NHCO alkyl, (CH2)mC02H, (CH2)mCO2-alkyl, (CH2)mSO3H, (CH2)mPO3H2, (CH2)mPO3(alkYl) 2 ~ ( CH2 ) mS2NH2 ~ and wossll26l2 2 1 7 0 7 6 6 pcT~ss4lll553 (CH2)mSO2NH-alkyl wherein alkyl is defined a~ above and m = 0, l, 2, or 3.
The term Uheteroaryl~ means a heteroaromatic ring which is a 2- or 3-thienyl, 2- or 3-furanyl, 2- or
Cell. Biol., 13:3706 (1993)). The fact that ras proteins are known to be involved in cell division processes makes them a candidate for intervention in many situations where cells are dividing uncontrol-lably. In direct analogy to the inhibition of mutant ras related cancer, blockade of ras dependant processes has the potential to reduce or eliminate the inappropriate tissue proliferation associated with restenosis, particularly in those instances where normal ras expression and/or function is exaggerated by growth stimulatory factors.
Ras functioning is dependent upon the modification of the proteins in order to associate with the inner face of plasma membranes. Unlike other membrane-associated proteins, ras proteins lack conventionaltr~ncm~mhrane or hydrophobic sequences and are initially synthesized in a cytosol soluble form. Ras protein membrane association is triggered by a series of posttranslational processing steps that are signaled by a carboxyl term;n~l amino acid consensus sequence that is recognized by protein:farnesyl transferase.
WO 95/12612 2 1 7 ~ 7 6 ~ PCT/US9~/115~i3 .
This consensus sequence consists of a cysteine residue located four amino acids from the carboxyl term;
followed by two lipophilic amino acids and the C-t~rm;n~l residue. The sulfhydryl group of the cysteine residue is alkylated by farnesyl pyrophosphate in a reaction that is catalyzed by protein:farnesyl transferase. Following prenylation, the C-termln~1 three amino acids are cleaved by an endoproteaQe and the newly exposed alpha-carboxyl group of the prenylated cysteine is methylated by a methyl transferase. The enzymatic processing of ras proteins that begins with farnesylation enables the protein to associate with the cell membrane. Mutational analysis of oncogenic ras proteins indicate that these posttranslational modifications are essential for transforming activity. Replacement of the consensus sequence cysteine residue with other amino acids gives a ras protein that is no longer farnesylated, fails to migrate to the cell membrane and lacks the ability to stimulate cell proliferation (Hancock J.F., et al., Cell, 57:1617 (1989); Scha~er w.R., et al., Science, 245:379 (1989); Casey P.J., Proc. Natl. Acad. Sci. USA, 86:8323 (1989)).
Recently, protein:farnesyl transferases (PFTs, also referred to as farnesyl:protein transferaseQ) have been identified and a specific PFT from rat brain was purified to homogeneity (Reiss Y., et al., Bioch. Soc.
Trans., 20:487-88 (1992)). The enzyme was charac-terized as a heterodimer composed of one alpha-subunit (49 kDa) and one beta-subunit (46 kDa), both of which are required for catalytic activity. High level expression of m~mm~lian PFT in a baculovirus system and purification of the recombinant enzyme in active fonm has also been accomplished (Chen W.-J., et al., J. Biol. Chem., 268:9675 (1993)).
PCT~S9411lS53 In light of the foregoing, the discovery that the function of oncogenic ras pro.teins is critically dependent on their posttranslational processing provides a means of cancer chemotherapy through S inhibition of the processing enzymes. The identification and isolation of a protein:farnesyl transferase that catalyzes the addition of a farnesyl group to ras proteins provides a promising target for such intervention. Recently it has been determined that prototypical inhibitors of PFT can inhibit ras processing and reverse cancerous morphology in tumor cell models (Kohl N.E., et al., Science, 260:1934 (1993); James G.L., et al., Science, 260:1937 (1993);
Garcia A.M., et al., J. Biol. Chem., 268:18415 (1993)).
Thus, it is possible to prevent or delay the onset of cellular proliferation in cancers that exhibit mutant ras proteins by blocking PFT. By analogous logic, inhibition of PFT would provide a potential means for controlling cellular proliferation associated with restenosis, especially in those cases wherein the expression and/or function of native ras is overstimulated.
PCT Application WO91/16340 discloses cysteine cont~in;ng tetrapeptide inhibitors of PFT of the formula CAAX.
European Patent Application 0461869 discloses cysteine cont~;n;ng tetrapeptide inhibitors of PFT of the formula Cys-Aaa1-Aaa2-Xaa.
European Patent Application 0520823 discloses cysteine cont~;n;ng tetrapeptide inhibitors of PFT of the formula Cys-Xaal-dXaa2-Xaa3.
European Patent Application 0523873 discloses cysteine cont~;n;ng tetrapeptide inhibitors of PFT of the formula Cys-Xaa1-Xaa2-Xaa3.
WO95/12612 2 1 7 0 7 6 6 PCT~S94/11553 European Patent Application 0528486 discloses cysteine cont~; n; ng tetrapeptide amides inhibitors of PFT of the formula Cys-Xaa1-Xaa2-Xaa3-NRRl.
European Patent Application 0535730 discloses pseudotetrapeptide inhibitors of PFT of the following two formulas:
X R2 o )~_,ZH
J~NH ~H~J~H~OH
HS'~ Y R 0 X R~ o HS
European Patent Application 0535731 (US 5,238,922) discloses esters of pseudotetrapeptide inhibitors of PFT of the formula:
X R2 o ()~ZH
- H ~ -~ N oR5 ~ Y R O
HS
European Patent Application 0482539 discloses tachykinin antagonists of the formula:
R
l,Y ~ A` ~ N~R5 0 R~ O
wos5ll26l2 PCT~S9~/11553 European Patent Application 0457195 discloses endothelin antagonists of the formula:
~
US 4,022,759 discloses tripeptide antagonists of luteinizing honmone releasing factor of the formula A-Rl-Tyr(benzyl)-Ser(benzyl)-R2, wherein one of the definitions of Rl is His(benzyl).
Compounds disclosed in the above references do not disclose or suggest a novel combination of structural variations found in the present invention described hereinafter. All cited references are hereby incorporated by reference.
SUMMARY OF THE IN V~N'1'10N
Accordingly, the present invention is a substituted di- or tripeptide compound of Formula I:
R
/
~rN
N~
~( 'H2) n A--N R O
R O ~ ) n 2 1 7~766 PCT~Ss~/115~3 W095/12~1~
wherein:
n = l or 2;
A = CoR3, Co2R3, CoNHR3, CSR3, C(S)oR3, C(S)NHR3, CF3SO2, aryl-SO2, or alkyl-SO2, wherein R3 is - 5 alkyl, (CH2) m~ cycloalkyl, (CH2) m~ aryl, ( CH2 ) m~ heteroaryl, or (CH2) m ~ alkyl, and m = 0, l, 2, or 3;
R = independently H or Me;
Y - independently H or Me;
Z - independently H or Me;
Rl - H, CO-aryl, (CH2) m ~ aryl, O(CH2) m ~ cycloalkyl, O(CH2)m-aryl, or O(CH2)m-heteroaryl, wherein m is as defined above and Rl i8 located at either the meta or para position;
X = one to four substituents, including H, alkyl, CF3, F, Cl, Br, I, HO, MeO, NO2, NH2, N(Me) 2' OPO3H2, or CH2PO3H2;
R2 = NR(CH2)nCo2R3, NR(CH2)nCoNHR3, NR(CH2)nR3, NR(CH2)n+l0R , NR(CH2)n+lSR ~
NRCH(CoR5)(CH2) n ~ heteroaryl, NRCH(CoR5)(CH2)noR3, NRCH(CoR5)(CH2)nSR3, or N N- R3 wherein R, R3, and n are as defined above, R4 = H
or R3, and R5 = OH, NH2, oR3, or NHR3; an optical isomer, diastereomer, or a ph~rm~ceutically acceptable salt thereof.
The present invention is also directed to the use of a compound of Formula I, or a pharmaceutically acceptable salt there~rom, to inhibit the activity of a protein:farnesyl transferase enzyme as a method for trea~ing tissue proliferative diseases.
A further embodiment of the present invention is the use of a pharmaceutical composition including an e~ective amount of a compound of Formula I as a method for the treatment of cancer.
A still further embodiment of the present invention is the use of a pharmaceutical composition including an effective amount of a compound of Formula I as a method for the treatment of restenosis.
A still further embodiment of the present invention i8 a ph~r~ceutical compo~ition for ~mi n; stering an effective amount of a compound of Fonmula I in unit dosage form in the treatment methods mentioned above.
A final embodiment of the present invention pertains to methods for the preparation of compounds of Formula I by solid phase synthesis, solution phase synthesis, and simultaneous multiple ~yntheses using a multiple simultaneous synthesis apparatus.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMæNTS
In the compounds of Formula I, the term "alkyl"
means a straight or branched hydrocarbon radical having from 1 to 6 carbon atoms and includes, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, and the like.
The term "cycloalkyl" means a saturated hydrocarbon ring which contains from 3 to 10 carbon atoms, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, adamantyl, and the like, unsubstituted or substituted by an alkyl or aryl group.
The term "aryl" means an aromatic ring which is a phenyl, 5-fluorenyl, 1-naphthyl or 2-naphthyl group, unsubstituted or substituted by 1 to 3 substituents selected from alkyl, O-alkyl and S-alkyl, O-aryl, OH, SH, F, Cl, Br, I, CF3, NO2, NH2, NHCH3, N(CH3) 2 NHCO alkyl, (CH2)mC02H, (CH2)mCO2-alkyl, (CH2)mSO3H, (CH2)mPO3H2, (CH2)mPO3(alkYl) 2 ~ ( CH2 ) mS2NH2 ~ and wossll26l2 2 1 7 0 7 6 6 pcT~ss4lll553 (CH2)mSO2NH-alkyl wherein alkyl is defined a~ above and m = 0, l, 2, or 3.
The term Uheteroaryl~ means a heteroaromatic ring which is a 2- or 3-thienyl, 2- or 3-furanyl, 2- or
3-pyrrolyl, 2-, 3- or 4-pyridyl, 2-, 3-, 4-, 5-, 6-, or 7-indolyl group, unsubstituted or with l or 2 substituents from the group of substituents described above for aryl.
The following table provides a list of abbreviations and definitions thereof used in the present invention.
TABLE OF ABBREVIATIONS
Abbreviation Amino Acid Ala Alanine Arg Arginine Asn Asparagine Asp Aspartic acid Cys Cysteine Glu Glutamic acid Gln Glutamine Gly Glycine His Histidine Ile Isoleucine Leu Leucine Lys Lysine Met Methionine Phe Phenylalanine Pro Proline Ser Serine Thr Threonine Trp Tryptophan If the optical activity of the amino acid iB other than L(S), the amino acid or abbreviation i8 preceded by the appropriate configuration D(R) or D~(RS).
PCT~S94/11553 Abbreviation Amino Acid (continued) Tyr Tyrosine Val Valine Abbreviation* Modified and Unusual Amino ~çid Aaa-CO2R An amino acid ester, for examples:
Gly-CO2Bn is Glycine, benzyl ester;
Ser(OBn)-CO2Me is O-Benzyl-serine, methyl ester.
Aaa-CONHR An amino acid amide, for examples:
Gly-CONHBn is Glycine, N-benzyl amide; Ser(OBn)-CONHEt is O-Benzyl-serine, N-ethyl amide;
Tyr(OBn)-CONHCH2CH20Bn is O-Benzyl-tyrosine, N-(2-(phenylmethoxy)ethyl) amide.
3Hyp 3-Hydroxyproline 4Hyp 4-Hydroxyproline Hcy Homocysteine Nva Norvaline Nle Norleucine Orn Ornithine Bal Beta-alanine (or 3-aminopropionic acid) Abu 4-Aminobutyric acid Ahe 7-Aminoheptanoic acid Acp 6-Aminocaproic acid Aoc 8-Aminooctanoic acid Apn 5-Aminopentanoic acid Bpa (4-Benzoylphenyl)alanine Chx 3-Cyclohexylalanine (or Hexahydrophenylalanine) Cit Citrulline If the optical activity of the amino acid i8 other than L(S), the amino acid or abbreviation i8 preceded by the appropriate configuration D(R) or DL(RS).
PCT~S94111553 WQ 9s/l26l2 2 1 ~ 0 7 6 6 Abbrevi~tion Modified and Unusual Amino Acid (continued) His(1-Me) l-Methyl-hi~tidine ~or N(~)-Methyl-histidine) His(Tr) 1-Triphenylmethyl-histidine (or N(~-Trityl-histidine) homoPhe 2-Amino-4-phenylbutanoic acid ~or Homophenylalanine) homoTyr 2-Amino-4-(4-hydroxyphenyl)butanoic acid (or Homotyrosine) homoTyr(OBn) 2-Amino-4-[4-(phenylmethoxy)phenyl]-butanoic acid (or O-Benzyl-homotyrosine) 1-Nal 3-(1'-Naphthyl)alanine 2-Nal 3-(2'-Naphthyl)alanine Pen Penicillamine Phe(3-OBn) (3-Benzyloxyphenyl)alanine Phe(4-Ph) 3-(l,l'Biphen-4-yl)alanine (or
The following table provides a list of abbreviations and definitions thereof used in the present invention.
TABLE OF ABBREVIATIONS
Abbreviation Amino Acid Ala Alanine Arg Arginine Asn Asparagine Asp Aspartic acid Cys Cysteine Glu Glutamic acid Gln Glutamine Gly Glycine His Histidine Ile Isoleucine Leu Leucine Lys Lysine Met Methionine Phe Phenylalanine Pro Proline Ser Serine Thr Threonine Trp Tryptophan If the optical activity of the amino acid iB other than L(S), the amino acid or abbreviation i8 preceded by the appropriate configuration D(R) or D~(RS).
PCT~S94/11553 Abbreviation Amino Acid (continued) Tyr Tyrosine Val Valine Abbreviation* Modified and Unusual Amino ~çid Aaa-CO2R An amino acid ester, for examples:
Gly-CO2Bn is Glycine, benzyl ester;
Ser(OBn)-CO2Me is O-Benzyl-serine, methyl ester.
Aaa-CONHR An amino acid amide, for examples:
Gly-CONHBn is Glycine, N-benzyl amide; Ser(OBn)-CONHEt is O-Benzyl-serine, N-ethyl amide;
Tyr(OBn)-CONHCH2CH20Bn is O-Benzyl-tyrosine, N-(2-(phenylmethoxy)ethyl) amide.
3Hyp 3-Hydroxyproline 4Hyp 4-Hydroxyproline Hcy Homocysteine Nva Norvaline Nle Norleucine Orn Ornithine Bal Beta-alanine (or 3-aminopropionic acid) Abu 4-Aminobutyric acid Ahe 7-Aminoheptanoic acid Acp 6-Aminocaproic acid Aoc 8-Aminooctanoic acid Apn 5-Aminopentanoic acid Bpa (4-Benzoylphenyl)alanine Chx 3-Cyclohexylalanine (or Hexahydrophenylalanine) Cit Citrulline If the optical activity of the amino acid i8 other than L(S), the amino acid or abbreviation i8 preceded by the appropriate configuration D(R) or DL(RS).
PCT~S94111553 WQ 9s/l26l2 2 1 ~ 0 7 6 6 Abbrevi~tion Modified and Unusual Amino Acid (continued) His(1-Me) l-Methyl-hi~tidine ~or N(~)-Methyl-histidine) His(Tr) 1-Triphenylmethyl-histidine (or N(~-Trityl-histidine) homoPhe 2-Amino-4-phenylbutanoic acid ~or Homophenylalanine) homoTyr 2-Amino-4-(4-hydroxyphenyl)butanoic acid (or Homotyrosine) homoTyr(OBn) 2-Amino-4-[4-(phenylmethoxy)phenyl]-butanoic acid (or O-Benzyl-homotyrosine) 1-Nal 3-(1'-Naphthyl)alanine 2-Nal 3-(2'-Naphthyl)alanine Pen Penicillamine Phe(3-OBn) (3-Benzyloxyphenyl)alanine Phe(4-Ph) 3-(l,l'Biphen-4-yl)alanine (or
4-Phenyl-phenylalanine) Pgl Phenylglycine Pyr 2-Amino-3-(3-pyridyl)-propanoic acid (or 3-Pyridylalanine) Ser(OBn) O-Benzyl-serine Thr(OBn) O-Benzyl-threonine Tic 1,2,3,4-Tetrahydro-3-i~oquinoline-carboxylic acid Tyr(OMe) O-Methyl-tyrosine Tyr(OEt) O-Ethyl-tyro~ine Tyr(OBn) O-Benzyl-tyrosine (~-Me)Tyr(OBn) 2-Amino-3-(4-benzyloxyphenyl)-2-methyl-propionic acid (or ~-Methyl-O-benzyl-tyrosine) (N-Me)Tyr(OBn) N-Methyl-O-benzyl-tyrosine Trp(For) Nln-Formyltryptophan If the optical activity of the amino acid i~ other than L(S), the amino acid or abbreviation i8 preceded by the a~op-iate configuration D(R) or DL(RS).
PCT/IJS9~/1 1553 2~a~
Abbreviation Mercapto Acids Maa Mercaptoacetic acid Mba 4-Mercaptobutyric acid Mpa 3-Mercaptopropionic acid Abbreviation Protectinq Group Ac Acetyl Ada 1-A~m~ntyl acetic acid Adoc A~m~ n tyloxycarbonyl Bn Benzyl MeBn 4-Methylbenzyl Cbz Benzyloxycarbonyl 2-Br-Cbz ortho-Bromobenzyloxycarbonyl 2-Cl-Cbz ortho-Chlorobenzyloxycarbonyl Bom Benzyloxymethyl Boc tertiary Butyloxycarbonyl Dnp 2,4-Dinitrophenyl For Formyl Fmoc 9-Fluorenylmethyloxycarbonyl NO2 Nitro TMS Trimethylsilyl Tos 4-Toluenesulfonyl (to~yl) Tr Triphenylmethyl (trityl) Abbreviation Solvents and Rea~ents HOAc Acetic acid CF3SO2H Trifluoromethanesulfonic acid DCM Dichloromethane DCC N,N'-Dicyclohexylcarbodiimide DIC N,N'-Diisopropylcarbodiimide DIEA N,N-Diisopropylethylamine DMAP 4-Dimethylaminopyridine DMF N,N'-Dimethylfonmamide EDAC N-Ethyl-N'-Dimethylaminopropyl-carbodiimide EtOAc Ethyl acetate PCT/US94/1 lS53 ~ 21 7~766 Abbreviation Solvents and Reaqents Et20 Diethyl ether HC1 Hydrochloric acid HF Hydrofluoric acid HOBT 1-Hydroxybenzotriazole KOH Potassium hydroxide MeCN Acetonitrile MeOH Methanol NHOS N-Hydroxysuccinimide NMP N-Methylpyrrolidone iPrOH iso-Propanol TBAF Tetra n-Butyl~mmo~-um Fluoride TFA Trifluoroacetic acid Abbreviation . Solid Phase Peptide Synthesis Resins HMP ~esin 4-(Hydroxymethyl)-phenoxymethyl-polystyrene resin MBHA Resin Methylbenzhydrylamine resin PAM Resin 4-(Hydroxymethyl)-phenylacetamidomethyl-polystyrene resin 2-Cl-Tr Resin 2-Chlorotrityl-polystyrene resin NH2-Rink Resin 4-(amino-(2',4'-dimethoxy-phenyl)methyl)-phenoxymethyl-polystyrene resin Abbreviation Biological Reaqents FPP Farnesyl pyrophosphate PFT Protein:farnesyl transferase DTT Dithiothreitol BSA Bovine serum albumin PCT~S94/11553 ~ 14-Abbreviation Miscellaneous CoR3 1IR3 CoNHR3 1l C ( S ) oR3 s CoR3 C(S)NHR3 lSI 3 CNHR
CH(CoR5)(CH2)noR3 (CH2)nCNHR3 CH(CoR5)(CH2) n~ heteroaryl o= Ic5 IC-(CH2) n ~ heteroaryl CH(CoR5)(CH2)noR3 O=C
C- (CH2)noR3 CH(CoR5)(CH2)noR3 o=c5 C- (CH2)nSR3 1l ~
CO(4-Bn-piperazin-1-yl) C-N ~ N-CH2Ph 2 1 7 n PCT~S94/11553 W095/1:612 -15 - u ~ 6 6 Preferred compounds of Formula I consist of compounds of Formula II below:
N-R
~; ~
R O
A'-N ~ z ~ RZ II
R O ~ "~
~ l ~ R
wherein:
A' ~ Co2R3, CoNHR3, C(S)NHR3, or aryl-S02, wherein R3 is alkyl, (CH2)m-cycloalkyl, (CH2)m-aryl, (CH2) m~ heteroaryl, and m = O, l, 2, or 3;
R = independently H or Me;
Y - independently H or Me;
Z = independently H or Me;
Rl = (CH2)m-aryl, O(CH2)m-aryl, OPO3H2, or CH2PO3H2, wherein m is as defined above;
R2 = NR(CH2) 2 oR4 ~ NR(CH2) 2 SR4, NRCH(CoR5)cH2oR3 s 3 ~
NRCH(COR )CH2SR , or , - N ~ N - CH2Ph wherein R, R3, and n are as defined above, R4 = H
or R3, and R5 = OH, NH2, oR3, or NHR3; an optical isomer, diastereomer, or a ph~rm~ceutically acceptable salt thereof.
Other preferred compounds of the present inventio are those of Formula I as defined above wherein A is Co2R3 or CoNHR3; or as defined above in Formula I
wherein at least one of Y and Z is Me; or as defined above in Formula I wherein R2 is (CH2)20R4 or CH(CoR5)CH2oR3; or as defined above in Formula I
PCT~Ss~/ll553 ~7 a7 66 -16-wherein A is CoNHR3, R2 is (CH2)2oR4, and at least one of Y and Z i8 Me.
The most preferred compounds of Fonmula I include the following:
Cbz-His-Tyr(OBn)-Ser(OBn)-C02Me;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-Tyr(OBn)-Ser(OBn);
Cbz-His-Tyr(OBn)-D-Ser(OBn)-CO2Me;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-D-His-Tyr(OBn)-Ser(OBn);
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn);
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-C02Me;
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His~l-Me)-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn);
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn);
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn);
Cbz-D-His-homoTyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-Phe(4-Ph)-Ser(OBn)-CO2Me;
Cbz-D-His-Phe(4-Ph)-Ser(OBn)-C02Me;
Cbz-His-Tyr(OBn)-Pyr-C02Me;
Cbz-D-His-Tyr(OBn)-Pyr-CO2Me;
Cbz-His-Tyr(OBn)-CONHCH2CH20Bn;
Cbz-D-His-Tyr(OBn)-CONHCH2CH2OBn;
pcT~ss~llls53 ~ 2 1 ~0~66 Cbz-His-(N-Me)Tyr(OBn)-CONHCH2CH20Bn;
Cbz-D-His-(N-Me)Tyr(OBn)-CONHCH2CH20Bn;
Cbz-His-Tyr(OBn)-CONH(CH2)2Ph;
Cbz-D-His-Tyr(OBn)-CONH(CH2)2Ph;
PCT/IJS9~/1 1553 2~a~
Abbreviation Mercapto Acids Maa Mercaptoacetic acid Mba 4-Mercaptobutyric acid Mpa 3-Mercaptopropionic acid Abbreviation Protectinq Group Ac Acetyl Ada 1-A~m~ntyl acetic acid Adoc A~m~ n tyloxycarbonyl Bn Benzyl MeBn 4-Methylbenzyl Cbz Benzyloxycarbonyl 2-Br-Cbz ortho-Bromobenzyloxycarbonyl 2-Cl-Cbz ortho-Chlorobenzyloxycarbonyl Bom Benzyloxymethyl Boc tertiary Butyloxycarbonyl Dnp 2,4-Dinitrophenyl For Formyl Fmoc 9-Fluorenylmethyloxycarbonyl NO2 Nitro TMS Trimethylsilyl Tos 4-Toluenesulfonyl (to~yl) Tr Triphenylmethyl (trityl) Abbreviation Solvents and Rea~ents HOAc Acetic acid CF3SO2H Trifluoromethanesulfonic acid DCM Dichloromethane DCC N,N'-Dicyclohexylcarbodiimide DIC N,N'-Diisopropylcarbodiimide DIEA N,N-Diisopropylethylamine DMAP 4-Dimethylaminopyridine DMF N,N'-Dimethylfonmamide EDAC N-Ethyl-N'-Dimethylaminopropyl-carbodiimide EtOAc Ethyl acetate PCT/US94/1 lS53 ~ 21 7~766 Abbreviation Solvents and Reaqents Et20 Diethyl ether HC1 Hydrochloric acid HF Hydrofluoric acid HOBT 1-Hydroxybenzotriazole KOH Potassium hydroxide MeCN Acetonitrile MeOH Methanol NHOS N-Hydroxysuccinimide NMP N-Methylpyrrolidone iPrOH iso-Propanol TBAF Tetra n-Butyl~mmo~-um Fluoride TFA Trifluoroacetic acid Abbreviation . Solid Phase Peptide Synthesis Resins HMP ~esin 4-(Hydroxymethyl)-phenoxymethyl-polystyrene resin MBHA Resin Methylbenzhydrylamine resin PAM Resin 4-(Hydroxymethyl)-phenylacetamidomethyl-polystyrene resin 2-Cl-Tr Resin 2-Chlorotrityl-polystyrene resin NH2-Rink Resin 4-(amino-(2',4'-dimethoxy-phenyl)methyl)-phenoxymethyl-polystyrene resin Abbreviation Biological Reaqents FPP Farnesyl pyrophosphate PFT Protein:farnesyl transferase DTT Dithiothreitol BSA Bovine serum albumin PCT~S94/11553 ~ 14-Abbreviation Miscellaneous CoR3 1IR3 CoNHR3 1l C ( S ) oR3 s CoR3 C(S)NHR3 lSI 3 CNHR
CH(CoR5)(CH2)noR3 (CH2)nCNHR3 CH(CoR5)(CH2) n~ heteroaryl o= Ic5 IC-(CH2) n ~ heteroaryl CH(CoR5)(CH2)noR3 O=C
C- (CH2)noR3 CH(CoR5)(CH2)noR3 o=c5 C- (CH2)nSR3 1l ~
CO(4-Bn-piperazin-1-yl) C-N ~ N-CH2Ph 2 1 7 n PCT~S94/11553 W095/1:612 -15 - u ~ 6 6 Preferred compounds of Formula I consist of compounds of Formula II below:
N-R
~; ~
R O
A'-N ~ z ~ RZ II
R O ~ "~
~ l ~ R
wherein:
A' ~ Co2R3, CoNHR3, C(S)NHR3, or aryl-S02, wherein R3 is alkyl, (CH2)m-cycloalkyl, (CH2)m-aryl, (CH2) m~ heteroaryl, and m = O, l, 2, or 3;
R = independently H or Me;
Y - independently H or Me;
Z = independently H or Me;
Rl = (CH2)m-aryl, O(CH2)m-aryl, OPO3H2, or CH2PO3H2, wherein m is as defined above;
R2 = NR(CH2) 2 oR4 ~ NR(CH2) 2 SR4, NRCH(CoR5)cH2oR3 s 3 ~
NRCH(COR )CH2SR , or , - N ~ N - CH2Ph wherein R, R3, and n are as defined above, R4 = H
or R3, and R5 = OH, NH2, oR3, or NHR3; an optical isomer, diastereomer, or a ph~rm~ceutically acceptable salt thereof.
Other preferred compounds of the present inventio are those of Formula I as defined above wherein A is Co2R3 or CoNHR3; or as defined above in Formula I
wherein at least one of Y and Z is Me; or as defined above in Formula I wherein R2 is (CH2)20R4 or CH(CoR5)CH2oR3; or as defined above in Formula I
PCT~Ss~/ll553 ~7 a7 66 -16-wherein A is CoNHR3, R2 is (CH2)2oR4, and at least one of Y and Z i8 Me.
The most preferred compounds of Fonmula I include the following:
Cbz-His-Tyr(OBn)-Ser(OBn)-C02Me;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-Tyr(OBn)-Ser(OBn);
Cbz-His-Tyr(OBn)-D-Ser(OBn)-CO2Me;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-D-His-Tyr(OBn)-Ser(OBn);
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn);
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-C02Me;
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His~l-Me)-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn);
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-(~-Me)Tyr(OBn)-Ser(OBn);
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-D-(~-Me)Tyr(OBn)-Ser(OBn);
Cbz-D-His-homoTyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-Phe(4-Ph)-Ser(OBn)-CO2Me;
Cbz-D-His-Phe(4-Ph)-Ser(OBn)-C02Me;
Cbz-His-Tyr(OBn)-Pyr-C02Me;
Cbz-D-His-Tyr(OBn)-Pyr-CO2Me;
Cbz-His-Tyr(OBn)-CONHCH2CH20Bn;
Cbz-D-His-Tyr(OBn)-CONHCH2CH2OBn;
pcT~ss~llls53 ~ 2 1 ~0~66 Cbz-His-(N-Me)Tyr(OBn)-CONHCH2CH20Bn;
Cbz-D-His-(N-Me)Tyr(OBn)-CONHCH2CH20Bn;
Cbz-His-Tyr(OBn)-CONH(CH2)2Ph;
Cbz-D-His-Tyr(OBn)-CONH(CH2)2Ph;
5 .Cbz-His-Tyr(OBn)-Gly-C02Bn;
Cbz-D-His-Tyr(OBn)-Gly-C02Bn;
Cbz-His-Tyr(OBn)-Gly-CONHBn;
Cbz-D-His-Tyr(OBn)-Gly-CONHBn;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-C02Me;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-Hi~-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-His-Tyr(OBn)-Ser(OBn);
BnNHCO-His-Tyr(OBn)-CONHCH2CH20Bn;
BnNHCO-His-Tyr(OBn)-CONHCH2CH2CH20Ph;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CO2Me;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn);
BnNHCO-D-His-Tyr(OBn)-CONHCH2CH20Bn;
BnNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH20Ph;
Cbz-His-Tyr(OBn)-CON(Me)CH2CH20Bn;
(4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)30Ph;
PhCH2CO-D-His-Tyr(OBn)-CONH(CH2)3-(2-MeOPh);
(4-PhOPh)NHCO-D-His-Tyr(OBn)-COHN(CH2)2Ph; and (4-MePh)S02-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl).
~ENERAL METHODS FOR THE PREPARATION, EVALUATION
AND USE OF COMPOUNDS OF FORMUhA I
The compounds of Formula I may be prepared by solid phase peptide synthesis on a peptide synthesizer, for example, an Applied Biosystems 43lA peptide 3~ synthesizer using activated esters or anhydrides of Boc or Fmoc protected amino acids, acid chlorides, WO9Sl12612 PCT~S94/11553 isocyanates, isothiocyanates, etc., on PAM, MBHA, or NH2-Rink resins with solution phase modifications to the carboxyl terminus as appropriate. Methodology for the solid phase synthesis of peptides is widely known to those skilled in the art thereof (see, for example:
J.M. Stewart and J.D. Young in Solid Phase Peptide Synthesis; Pierce Chemical Co.; Rockford, IL (1984);
Fields G.B. and Noble R.L., Int. J. Peptide Protein ~ç~, 35:161-214 (1990)).
Additionally, the compounds of Formula I may also be prepared by conventional solution peptide synthesis, substituting ~m; n~s, acid chlorides, isocyanates, etc, for amino acid derivatives where appropriate. Methods for solution phase synthesis of peptides are widely known to those skilled in the art (see, for example, M. Bodanszky, Principles of Peptide Synthesia, Springer-Verlag (1984)).
Finally, the compounds of Formula I may be prepared by simultaneous multiple solid phase syntheses using an apparatus described by S. H. DeWitt, et al., Proc. Natl. Acad. Sci. USA, 90:6909 (1993), and referred to by the trademark, Diversomer~, both trA~m~rk and apparatus being owned in whole by the Warner-~ambert Company. The multiple solid phase synthesis apparatus is currently the subject of now ab;tn~on~ US Serial 07/958,383 filed October 8, 1992 and pending continuation-in-part US Serial 08/012,557 filed February 2, 1993.
For example (Scheme I below), Fmoc-D-His-Tyr(OBn)-CO2-CH2CH2Si(CH3)3 is linked to 2-Cl-Tr resin using a sterically hindered amine such as DIEA as an HCl scavenger, the Fmoc protecting group is removed with piperidine, the resulting free amino term;ntl~ is acylated with a series of isocyanates, isothiocyanates, activated esters, acid chlorides and the like, the TMS-ethyl ester is cleaved with TBAF, the resulting ~ ~ PCT~Ss4/11553 WO95112612 ~ ~ 70~66 .
free carboxy term;nl~ is activated with a carbodiimide reagent such as EDAC, DCC, or DIC, the activated carboxyl group is reacted with alcohols such as HOBT, NHOS, or pentachlorophenol to give an activated eQter, 5 the activated ester is reacted with a series of amines and the reculting array of compounds of Formula I i8 cleaved from the resin by with hot HOAc or by treatment with TFA at room temperature.
For all three synthetic methods described above appropriate consideration is given to protection and deprotection of reactive functional groups and to the sequence of synthetic steps. Knowledge of the use of common protecting groups and strategy for the assembly of complex organic molecules are within the usual realm of expertise of a practitioner of the art of organic chemistry (see, for example: T.W. Greene and P.G.M. Wuts, Protective Groups in Organic Chçmistry, John Wiley and Sons (l99l); E.J. Corey and X.-M. Cheng, The Logic of Chemical Synthesis, John Wiley and Sons (l989)).
The homogeneity and composition of the resulting compounds is verified by reverse-phase-high pressure liquid chromatography (RP-HPLC), capillary electrophoresis, thin layer chromatography (T~C), proton nuclear magnetic resonance spectrometry (NMR), amino acid analysis, fast atom bombardment mass spectrometry (FAB-MS) and electrospray mass spectrometry (ES-MS).
PCT~S9~/11553 ~7~ 20-SCHEME I: Multiple Simultaneous Synthesi~ Method Resin ~ Resin 1. Piperidine ~N~ Cl ~ N Cl \\ // 2. Eight acylating N ~ agents N--~
FmocNH ~ ~ OCH2CH2TMS ~ ~ OCH2CH2TMS
O / O
~ ~
BnO BnO
1. TBAF
2. EDAC, HOBT
3. Five amines, Resin =
N ~N Cl H ~ ~ TFA A~ ~ ~ ~N J ~R2 BnO
40 Compounds o~ Formula I /~~~
EnO
pcT~ss~llls53 Wo 95112612 2 ~ 6 6 The compounds of Formula I are capable of further forming both pharmaceutically acceptable acid addition and/or base salts. A11 of these forms are within the scope of the present invention.
Pharmaceutically acceptable acid addition salts of the compounds of Formula I include ~alts derived from nontoxic inorganic acids such as hydrochloric, nitric, pho~phoric, sulfuric, hydrobromic, hydroiodic, hyd~ofluoric, phosphorous, and the like, as well as the salts derived from nontoxic organic acids, such a~
aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, ~k~nedioic acids, aromatic acids, aliphatic and aromatic sulfonic acids, etc. Such salts thus include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, trifluoroacetate, propionate, caprylate, isobutyrate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, mandelate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, phthalate, benzenesulfonate, toluenesulfonate, phenylacetate, citrate, lactate, maleate, tartrate, methanesulfonate, and the like.
Also contemplated are salts of amino acids such as arginate and the like and gluconate, galacturonate, n-methyl glucamine (see, for example, Berge S.M., et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science, 66:1-19 (1977)).
The acid addition salts of said basic compounds are prepared by contacting the free base form with a sufficient amount of the desired acid to produce the salt in the conventional manner. Preferably a compound o~ Formula I can be converted to an acidic salt by treating with an aqueous solution of the desired acid, such that the resulting pH is less than 4. The b~ -22-solution can be passed through a C18 cartridge to absorb the compound, washed with copious amounts of water, the compound eluted with a polar organic solvent such as, for example, methanol, acetonitrile, and the like, and isolated by concentrating under reduced pressure followed by lyophilization. The free base form may be regenerated by contacting the salt form with a base and isolating the free base in the conventional manner or as above. The free base forms differ from their respective salt forms somewhat in certain physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free base for purposes of the present invention.
Pharmaceutically acceptable base addition salts are formed with metals or amines, such as alkali and alkaline earth metals or organic amines. Examples of metals used as cations are sodium, potassium, magnesium, calcium, and the like. Examples of suitable amines are N,N'-dibenzylethylene~'~m;n~, chloroprocaine, choline, diethanolamine, dicyclohexylamine, ethylenediamine, N-methylgluc~m;ne, and procaine (see, for example, Berge S.M., et al., "Pharmaceutical Salts", Journal of Pharmaceutical Science, 66:1-19 (1977)).
The base addition salts of said acidic compounds are prepared by contacting the free acid form with a sufficient amount of the desired base to produce the salt in the conventional manner. Preferably, a compound of Formula I can be converted to a base salt by treating with an aqueous solution of the desired base, such that the resulting pH is greater than 9.
The solution can be passed through a C18 cartridge to absorb the compound, washed with copious amounts of water, the compound eluted with a polar organic solvent such as, for example, methanol, acetonitrile and the PCT~S94/11553 Wo95112612 2 1 7 0 7 6 6 like, and i~olated by concentrating under reduced pressure followed by lyophilization. The free acid ~ form may be regenerated by contacting the salt form with an acid and isolating the free acid in the con~entional manner or as above. The free acid forms differ from their respective salt forms somewhat in certain physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free acid for purposes of the present invention.
Certain of the compounds of the present invention can exist in unsolvated forms as well as solvated ~orms, including hydrated forms. In general, the solvated forms, including hydrated forms, are equivalent to unsolvated forms and are intended to be encompassed within the scope of the present invention.
Certain of the compounds of the present invention possess one or more chiral centers and each center may exist in the R(D) or S(L) configuration. The present invention includes all enantiomeric and epimeric forms as well as the appropriate mixtures thereo~.
The PFT inhibitory activity of compounds o~
Formula I was assayed in 30 mM potassium phosphate buffer, pH 7.4, containing 7 mM DTT, 1.2 mM MgC12, 0.1 mM leupeptin, 0.1 mM pepstatin and 0.2 mM
phenylmethylsulfonyl fluoride. Assays were performed in 96 well plates (Wallec) and employed solutions composed of varying concentrations of a compound of Formula I in lO0~ DMS0. Upon addition of both substrates, radiolabeled ~arnesyl pyrophosphate (rl-3~], specific activity 15-30 Ci/mmol, final concentration 0.12 ~M) and (biotinyl)-Ahe-Tyr-Lys-Cys-Val-Ile-Met peptide (final concentration O.l ~M), the enzyme reaction was started by addition of 40-fold purified rat brain farnesyl protein transferase. After incubation at 37C for 30 minutes, the reaction was ~ 7 ~1 6~
t~rm;n~ted by diluting the reaction 2.5-fold with a stop buffer cont~;n;ng 1.5 M magnesium acetate, 0.2 M
H3P04, 0.5~ BSA, and strepavidin beads (Amersham) at a concentration of 1.3 mg/m~. After allowing the plate to settle for 30 minutes at room temperature, radioactivity was quantitated on a microBeta counter (model 1450, Wallec). Compounds of Formula I show IC50 values of 0.5 nM to 80 ~M (see data table) in this assay and are thus valuable inhibitors of protein:
farnesyl transferase enzyme which may be used in the medical treatment of tissue proliferative diseases, including cancer and restenosis.
IC50 Values for Selected Compounds of Formula I Against PFT
Example Number IC50 (~M) 1 4.4 4 1.0 5 (3) 2.1 5 (4) 7.3 5 (23) 0.64 5 (27) 30 5 (28) 0.73 5 (30) 73 5 (31) 0.76 5 (35) 66 5 (36) 1.9 5 (46) 1.0 5 (49) 2.9 5 (40) 0.75 5 (52) 1.6 5 (56) 1.1 5 (59) 20 5 (60) 1.4 PCT~S94/11553 ~ 2 1 7~766 IC50 Values for Selected Compounds of Formula I Against PFT
Example Number IC50 (~M) 5 (61) 7.2 5 (62) 1.5 5 (63) 1.0 5 (64) 1.7 5 (69) 0.48 5 (79) 3.0 5 (80) 1.6
Cbz-D-His-Tyr(OBn)-Gly-C02Bn;
Cbz-His-Tyr(OBn)-Gly-CONHBn;
Cbz-D-His-Tyr(OBn)-Gly-CONHBn;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-C02Me;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-Hi~-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-His-Tyr(OBn)-Ser(OBn);
BnNHCO-His-Tyr(OBn)-CONHCH2CH20Bn;
BnNHCO-His-Tyr(OBn)-CONHCH2CH2CH20Ph;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CO2Me;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn);
BnNHCO-D-His-Tyr(OBn)-CONHCH2CH20Bn;
BnNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH20Ph;
Cbz-His-Tyr(OBn)-CON(Me)CH2CH20Bn;
(4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)30Ph;
PhCH2CO-D-His-Tyr(OBn)-CONH(CH2)3-(2-MeOPh);
(4-PhOPh)NHCO-D-His-Tyr(OBn)-COHN(CH2)2Ph; and (4-MePh)S02-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl).
~ENERAL METHODS FOR THE PREPARATION, EVALUATION
AND USE OF COMPOUNDS OF FORMUhA I
The compounds of Formula I may be prepared by solid phase peptide synthesis on a peptide synthesizer, for example, an Applied Biosystems 43lA peptide 3~ synthesizer using activated esters or anhydrides of Boc or Fmoc protected amino acids, acid chlorides, WO9Sl12612 PCT~S94/11553 isocyanates, isothiocyanates, etc., on PAM, MBHA, or NH2-Rink resins with solution phase modifications to the carboxyl terminus as appropriate. Methodology for the solid phase synthesis of peptides is widely known to those skilled in the art thereof (see, for example:
J.M. Stewart and J.D. Young in Solid Phase Peptide Synthesis; Pierce Chemical Co.; Rockford, IL (1984);
Fields G.B. and Noble R.L., Int. J. Peptide Protein ~ç~, 35:161-214 (1990)).
Additionally, the compounds of Formula I may also be prepared by conventional solution peptide synthesis, substituting ~m; n~s, acid chlorides, isocyanates, etc, for amino acid derivatives where appropriate. Methods for solution phase synthesis of peptides are widely known to those skilled in the art (see, for example, M. Bodanszky, Principles of Peptide Synthesia, Springer-Verlag (1984)).
Finally, the compounds of Formula I may be prepared by simultaneous multiple solid phase syntheses using an apparatus described by S. H. DeWitt, et al., Proc. Natl. Acad. Sci. USA, 90:6909 (1993), and referred to by the trademark, Diversomer~, both trA~m~rk and apparatus being owned in whole by the Warner-~ambert Company. The multiple solid phase synthesis apparatus is currently the subject of now ab;tn~on~ US Serial 07/958,383 filed October 8, 1992 and pending continuation-in-part US Serial 08/012,557 filed February 2, 1993.
For example (Scheme I below), Fmoc-D-His-Tyr(OBn)-CO2-CH2CH2Si(CH3)3 is linked to 2-Cl-Tr resin using a sterically hindered amine such as DIEA as an HCl scavenger, the Fmoc protecting group is removed with piperidine, the resulting free amino term;ntl~ is acylated with a series of isocyanates, isothiocyanates, activated esters, acid chlorides and the like, the TMS-ethyl ester is cleaved with TBAF, the resulting ~ ~ PCT~Ss4/11553 WO95112612 ~ ~ 70~66 .
free carboxy term;nl~ is activated with a carbodiimide reagent such as EDAC, DCC, or DIC, the activated carboxyl group is reacted with alcohols such as HOBT, NHOS, or pentachlorophenol to give an activated eQter, 5 the activated ester is reacted with a series of amines and the reculting array of compounds of Formula I i8 cleaved from the resin by with hot HOAc or by treatment with TFA at room temperature.
For all three synthetic methods described above appropriate consideration is given to protection and deprotection of reactive functional groups and to the sequence of synthetic steps. Knowledge of the use of common protecting groups and strategy for the assembly of complex organic molecules are within the usual realm of expertise of a practitioner of the art of organic chemistry (see, for example: T.W. Greene and P.G.M. Wuts, Protective Groups in Organic Chçmistry, John Wiley and Sons (l99l); E.J. Corey and X.-M. Cheng, The Logic of Chemical Synthesis, John Wiley and Sons (l989)).
The homogeneity and composition of the resulting compounds is verified by reverse-phase-high pressure liquid chromatography (RP-HPLC), capillary electrophoresis, thin layer chromatography (T~C), proton nuclear magnetic resonance spectrometry (NMR), amino acid analysis, fast atom bombardment mass spectrometry (FAB-MS) and electrospray mass spectrometry (ES-MS).
PCT~S9~/11553 ~7~ 20-SCHEME I: Multiple Simultaneous Synthesi~ Method Resin ~ Resin 1. Piperidine ~N~ Cl ~ N Cl \\ // 2. Eight acylating N ~ agents N--~
FmocNH ~ ~ OCH2CH2TMS ~ ~ OCH2CH2TMS
O / O
~ ~
BnO BnO
1. TBAF
2. EDAC, HOBT
3. Five amines, Resin =
N ~N Cl H ~ ~ TFA A~ ~ ~ ~N J ~R2 BnO
40 Compounds o~ Formula I /~~~
EnO
pcT~ss~llls53 Wo 95112612 2 ~ 6 6 The compounds of Formula I are capable of further forming both pharmaceutically acceptable acid addition and/or base salts. A11 of these forms are within the scope of the present invention.
Pharmaceutically acceptable acid addition salts of the compounds of Formula I include ~alts derived from nontoxic inorganic acids such as hydrochloric, nitric, pho~phoric, sulfuric, hydrobromic, hydroiodic, hyd~ofluoric, phosphorous, and the like, as well as the salts derived from nontoxic organic acids, such a~
aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, ~k~nedioic acids, aromatic acids, aliphatic and aromatic sulfonic acids, etc. Such salts thus include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, trifluoroacetate, propionate, caprylate, isobutyrate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, mandelate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, phthalate, benzenesulfonate, toluenesulfonate, phenylacetate, citrate, lactate, maleate, tartrate, methanesulfonate, and the like.
Also contemplated are salts of amino acids such as arginate and the like and gluconate, galacturonate, n-methyl glucamine (see, for example, Berge S.M., et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science, 66:1-19 (1977)).
The acid addition salts of said basic compounds are prepared by contacting the free base form with a sufficient amount of the desired acid to produce the salt in the conventional manner. Preferably a compound o~ Formula I can be converted to an acidic salt by treating with an aqueous solution of the desired acid, such that the resulting pH is less than 4. The b~ -22-solution can be passed through a C18 cartridge to absorb the compound, washed with copious amounts of water, the compound eluted with a polar organic solvent such as, for example, methanol, acetonitrile, and the like, and isolated by concentrating under reduced pressure followed by lyophilization. The free base form may be regenerated by contacting the salt form with a base and isolating the free base in the conventional manner or as above. The free base forms differ from their respective salt forms somewhat in certain physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free base for purposes of the present invention.
Pharmaceutically acceptable base addition salts are formed with metals or amines, such as alkali and alkaline earth metals or organic amines. Examples of metals used as cations are sodium, potassium, magnesium, calcium, and the like. Examples of suitable amines are N,N'-dibenzylethylene~'~m;n~, chloroprocaine, choline, diethanolamine, dicyclohexylamine, ethylenediamine, N-methylgluc~m;ne, and procaine (see, for example, Berge S.M., et al., "Pharmaceutical Salts", Journal of Pharmaceutical Science, 66:1-19 (1977)).
The base addition salts of said acidic compounds are prepared by contacting the free acid form with a sufficient amount of the desired base to produce the salt in the conventional manner. Preferably, a compound of Formula I can be converted to a base salt by treating with an aqueous solution of the desired base, such that the resulting pH is greater than 9.
The solution can be passed through a C18 cartridge to absorb the compound, washed with copious amounts of water, the compound eluted with a polar organic solvent such as, for example, methanol, acetonitrile and the PCT~S94/11553 Wo95112612 2 1 7 0 7 6 6 like, and i~olated by concentrating under reduced pressure followed by lyophilization. The free acid ~ form may be regenerated by contacting the salt form with an acid and isolating the free acid in the con~entional manner or as above. The free acid forms differ from their respective salt forms somewhat in certain physical properties such as solubility in polar solvents, but otherwise the salts are equivalent to their respective free acid for purposes of the present invention.
Certain of the compounds of the present invention can exist in unsolvated forms as well as solvated ~orms, including hydrated forms. In general, the solvated forms, including hydrated forms, are equivalent to unsolvated forms and are intended to be encompassed within the scope of the present invention.
Certain of the compounds of the present invention possess one or more chiral centers and each center may exist in the R(D) or S(L) configuration. The present invention includes all enantiomeric and epimeric forms as well as the appropriate mixtures thereo~.
The PFT inhibitory activity of compounds o~
Formula I was assayed in 30 mM potassium phosphate buffer, pH 7.4, containing 7 mM DTT, 1.2 mM MgC12, 0.1 mM leupeptin, 0.1 mM pepstatin and 0.2 mM
phenylmethylsulfonyl fluoride. Assays were performed in 96 well plates (Wallec) and employed solutions composed of varying concentrations of a compound of Formula I in lO0~ DMS0. Upon addition of both substrates, radiolabeled ~arnesyl pyrophosphate (rl-3~], specific activity 15-30 Ci/mmol, final concentration 0.12 ~M) and (biotinyl)-Ahe-Tyr-Lys-Cys-Val-Ile-Met peptide (final concentration O.l ~M), the enzyme reaction was started by addition of 40-fold purified rat brain farnesyl protein transferase. After incubation at 37C for 30 minutes, the reaction was ~ 7 ~1 6~
t~rm;n~ted by diluting the reaction 2.5-fold with a stop buffer cont~;n;ng 1.5 M magnesium acetate, 0.2 M
H3P04, 0.5~ BSA, and strepavidin beads (Amersham) at a concentration of 1.3 mg/m~. After allowing the plate to settle for 30 minutes at room temperature, radioactivity was quantitated on a microBeta counter (model 1450, Wallec). Compounds of Formula I show IC50 values of 0.5 nM to 80 ~M (see data table) in this assay and are thus valuable inhibitors of protein:
farnesyl transferase enzyme which may be used in the medical treatment of tissue proliferative diseases, including cancer and restenosis.
IC50 Values for Selected Compounds of Formula I Against PFT
Example Number IC50 (~M) 1 4.4 4 1.0 5 (3) 2.1 5 (4) 7.3 5 (23) 0.64 5 (27) 30 5 (28) 0.73 5 (30) 73 5 (31) 0.76 5 (35) 66 5 (36) 1.9 5 (46) 1.0 5 (49) 2.9 5 (40) 0.75 5 (52) 1.6 5 (56) 1.1 5 (59) 20 5 (60) 1.4 PCT~S94/11553 ~ 2 1 7~766 IC50 Values for Selected Compounds of Formula I Against PFT
Example Number IC50 (~M) 5 (61) 7.2 5 (62) 1.5 5 (63) 1.0 5 (64) 1.7 5 (69) 0.48 5 (79) 3.0 5 (80) 1.6
6 0.42
7 0.26
8 0.074
9 0.27 0.10 11 0.17 12 0.028 13 0.083 16 0.60 17 0.039 18 0.82 19 0.31 21 0.31 22 0.37 23 1.9 IC50 Values for Selected Compounds of Formula I Against PFT (cont'd) Example Number IC50 (~M) 24 1.0 3.7 29 3.0 The compounds of the present invention can be prepared and ~min;stered in a wide variety of oral, rectal and parenteral dosage forms. Thus, the compounds of the present invention can be A~mi n; gtered by injection, that is, intravenously, intramuscularly, intracutaneously, subcutaneously, intraduodenally, or intraperitoneally. Also, the compounds of the present invention can be A~ml n; stered by inhalation, for example, intranasally. Additionally, the compounds of the present invention can be administered transdermally. It will be obvious to those skilled in the art that the following dosage forms may comprise as the active component, either a compound of Formula I or a corresponding phArm~ceutically acceptable salt of a compound of Formula I.
For preparing pharmaceutical compositions from the compounds of the present invention, pharmaceutically acceptable carriers can be either solid or liquid.
Solid form preparations include powders, tablets, pills, capsules, cachets, suppositories, and dispersible granules. A solid carrier can be one or more substances which may also act as diluents, flavoring agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material.
pcT~ss~ 553 ~ 2 1 70766 In powders, the carrier is a finely divided solid which is in a mixture with ~he finely di~ided active component.
In tablets, the active component is mixed with the carrier having the necessary binding properties in suitable proportions and compacted in the shape and size desired.
The powders and tablets preferably contain from 5 or lO to about 70 percent of the active compound.
Sui~able carriers are magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter, and the like. The term "preparation" is intended to include the formulation of the active compound with encapsulating material as a carrier providing a capsule in which the active component with or without other carriers, is surrounded by a carrier, which is thus in association with it. Similarly, cachets and lozenges are included. Tablet~, powders, capsules, pills, cachets, and lozenges can be used as solid dosage forms suitable for oral ~m; n; gtration.
For preparing suppositories, a low melting wax, such as a mixture of fatty acid glycerides or cocoa butter, is first melted and the active component is dispersed homogeneously therein, as by stirring. The molten homogenous mixture i5 then poured into convenient sized molds, allowed to cool, and thereby to ~olidify.
Liquid form preparations include solutions, suspensions, and emulsions, for example, water or water propylene glycol solutions. For parenteral injection liquid preparations can be formulated in solution in aqueous polyethylene glycol solution.
Aqueous solutions suitable for oral use can be prepared by dissolving the active component in water WO9S112612 PCT~S9~111553 .
1 ~1 6 ~ -28-and adding suitable colorants, flavors, stabilizing and thickening agents as desired.
Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, and other well-known suspending agents .
Also included are solid form preparations which are intended to be converted, shortly before use, to liquid form preparations for oral A~m; n; stration. Such liquid forms include solutions, suspensions, and emulsions. These preparations may contain, in addition to the active component, colorants, flavors, stabilizers, buffers, artificial and natural sweeteners, dispersants, thickeners, solubilizing agents, and the like.
The phAr~Aceutical preparation is preferably in unit dosage form. In such ~orm the preparation is subdivided into unit doses contA;n;ng appropriate quantities of the active component. The unit dosage form can be a packaged preparation, the package contA i nl ng discrete quantities of preparation, such as packeted tablets, capsules, and powders in vials or ampoules. Also, the unit dosage form can be a capsules, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form.
The quantity of active component in a unit dose preparation may be varied or adjusted from O.l mg to lO0 mg preferably 0.5 mg to lO0 mg according to the particular application and the potency of the active component. The composition can, if desired, also contain other compatible therapeutic agents.
In therapeutic use as inhibitors of PFT, the compounds utilized in the phanmaceutical methods of WO 95/1~612 _, ~ 6 6 this invention are administered at the initial dosage of about 0.01 mg/kg to about 20 mg/kg daily. A daily dose range of about 0.01 mg/kg to about 10 mg/kg is preferred. The dosages, however, may be varied depending upon the requirements of the patient, the severity of the condition being treated, and the compound being employed. Determination of the proper dosage for a particular situation is within the skill of the art. Generally, treatment is initiated with smaller dosages which are less than the optimum dose of the compound. Thereafter, the dosage is increased by small increments until the optimum effect under the circumstances is reached. For convenience, the total daily dosage may be divided and ~m~ n; stered in port:ions during the day, if desired.
The following nonlimiting examples illustrate the inventors' preferred methods for preparing the compounds of the invention. For added clarity, complex chemical names describing compounds of Formula I are followed by structural abbreviations, which are shown in braces, wherein the structural elements are as defi~ed in the Table of Abbreviations above.
~-rN-rN-[(Phenylmethoxy)carbonyl]-L-histidyll-O-(phenylmethyl)-L-tyrosyl]-O-(phenylmethyl)-L-serinamide ~Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2}
Using an ABI model 431A solid phase peptide synthesizer, Fmoc-NH2-Rink resin (0.25 mmol scale) was treated with 20~ piperidine in NMP to afford NH2-Rink resin. Sequential coupling of Fmoc-protected Ser(OBn) and Tyr(OBn) (DCC and HOBT in NMP) and Fmoc deprotection (20~ piperidine in NMP) reactions were run using a 4-fold excess of reagents in the coupling steps and traditional resin washing cycles to afford Tyr(OBn)-Ser(OBn)-CONH2-Rink resin. This dipeptide PCT~S94/11553 ~lQ16~ ' --resin was transferred to an uninstrumented reaction vessel and treated with a 4-fold excess of Cbz-His, DCC, and HOBT in DMF, shaking overnight at room temperature. After removal of excess reagents, the resulting substituted tripeptide was cleaved from the resin by treatment with 50~ TFA in DCM at room temperature for 2.5 hours. Evaporation of solvents and purification by reversed phase chromatography (Cl8-column, eluted with a 20-70~ gradient of MeCN in water (both solvents acidi~ied with 0.l~ TFA) afforded Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2 as its TFA salt upon lyophilization. ES-MS: 719 (m+l).
Using analogous methods the following most preferred compounds of Formula I with carboxamides at the C-terminus may be prepared:
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-DL-(~-Me)Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONH2; and BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONH2.
N-rN-rN-[(Phenylmethoxy)carbonyll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-serine {Cbz-His-Tyr(OBn)-Ser(OBn)}
Beginning with PAM resin or HMP resin, Fmoc-Ser(OBn), Fmoc-Tyr(OBn), and Cbz-His are sequentially coupled using the deprotection and coupling conditions described in Example l. Cleavage from the resin is accomplished by treatment with CF3SO2H for the PAM supported tripeptide or with 50 TFA in DCM for the HMP supported tripeptide.
Chromatography as in Example l provides Cbz-His-Tyr(OBn)-Ser(OBn) as its TFA salt upon PCT~S94/11553 W095/12612 2 1 ~ ~66 .
lyophilization. See also Example 7 for a solution phase method.
Using analogous methods the following most preferred compounds of Formula I with a free carboxyl term; m~ may be prepared:
Cbz-D-His-Tyr(OBn)-Ser(OBn);
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn);
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(O~n);
Cbz-Hi~-D~-(~-Me)Tyr(OBn)-Ser(OBn3;
BnNHCO-His-Tyr(OBn)-Ser(OBn); and BnNHCO-D-His-Tyr(OBn)-Ser(OBn).
Solid phase supported N-~N-~(9H-Fluoren-9-ylmethoxy)car~onYl]-L-histidyll-O-(phenylmethyl)-L-tyrosine, 2-trimethylsilylethyl ester {Fmoc-His(2-Cl-Tr Resin)-Tyr(OBn)-CO2CH2CH2TMS}
Step 1: Boc-Tyr(OBn)-C02CH2CH2TMS
2-Trimethylsilyl ethanol (2.6 g, 22.6 mmol) was added to a pr~m~ solution of EDAC (4.3 g, 22.6 mmol), DMAP (0.5 g), and Boc-Tyr(OBn)-OH (7.0 g, 18.8 mmol) in dry THF (25 mL). The resulting mixture was stirred for 18 hours at room temperature. The solu~ion was diluted with 1:1 EtOAc:Et20 (40 mL), 2~ washed with saturated aqueous NaHC03 (2 x 10 m~) and with saturated aqueous NaCl (2 x 10 mL), dried (MgSO4), filtered and concentrated in vacuo to provide an oil which was further purified by flash chromatography (SiO2, EtOAc:hexane eluent) to give the pure TM~-ethyl ester as an oil;
H NMR (HCDCl3): ~ 0.04 (s, 9H), 1.43 (8, 9H), 3.03 (m, 2H), 4.22 (m, 2H), 4.51 (m, lH), 4.95 (m, lH), 5.05 (s, 2H), 6.85-7.48 (m, 9H).
PCT~S94111553 ~7`~76~ --Step 2: Tyr(OBn)-CO2CH2CH2TMS
Eighty percent TFA in CH2C12 (20 mL, v/v) was added to an ice-cooled solution of Boc-Tyr(OBn)-CO2CH2CH2TMS (14.8 g, 31.4 mmol) in CH2Cl2 (40 mL). The reQulting mixture was stirred for 1.0 minute before concentrating in vacuo. The procedure was repeated once more, and the resulting residue was diluted with CH2Cl2 and saturated aqueous NaHCO3. The resulting mixture was filtered through celite. The organic layer was then separated, washed with saturated aqueous NaCl, and dried (MgSO4). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:MeOH eluent) to give the de~ired product;
1H NMR (HCDC13): ~ 0.06 (s, 9H), 1.68 (br ~, 2H), 2.85-3.03 (m, 2H), 3.67 (m, lH), 4.22 (m, 2H), 5.05 (s, 2H), 6.91-7.45 (m, 9H).
Step 3: Fmoc-His(Tr)-Tyr(OBn)-CO2CH2CH2TMS
To a solution of HOBT (2.6 g, 19.3 mmol) in DMF
(10 mL) was added Fmoc-His(Tr) ( 10.0 g, 16.1 mmol) followed by EDAC (3.7 g, 19.3 mmol). The mixture was stirred at room temperature for 20 minutes before adding a solution of Tyr(OBn?-CO2CH2CH2TMS (from Step 2 above, 5.8 g, 16.1 mmol) in DMF (10 mL). The mixture was stirred overnight at room temperature before partitioning between a mixture of water and 1:1 Et2O:EtOAc (50 mL). The layers were separated, and the organic phase was washed with saturated aqueous NaCl (4 x 20 mL) and dried (MgSO4). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:MeOH eluent) to give the protected His-Tyr dipeptide; FAB-MS 974 (m+1).
W095/12612 PCT~S94/11553 2 1 7~66 Step 4: Fmoc-His-Tyr(OBn)-CO2CH2CH2TMS
Fmoc-His(Tr)-Tyr(OBn)-CO2CH2CH2TMS (from Step 3 above, 5.0 g, 5.1 mmol) was treated with pyridineoHCl (1.0 g) in MeOH (20 mL). The mixture was allowed to stir 8 hours at 65 C. The solution was concentrated in vacuo, and the residue was dissolved in CH2Cl2, washed with H20 (lx), saturated aqueous NaHC03 (2x), and dried (MgS04). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:MeOH eluent) to give Fmoc-His-Tyr(OBn)-CO2CH2CH2STMS as a white solid;
FAB-MS 731 (m+1).
Step 5: Fmoc-His(2-Cl-Tr Resin)-Tyr(OBn)-CO2CH2CH2TMS
To a suspension of Fmoc-His-Tyr(OBn)- C02CH2CH2TM~
(from Step 4 above, 5.3 g, 7.3 mmol) in CHC13 (20 mL) was added 2-chloroltrityl chloride resin (Novabiochem) (7.1 g) followed by DIEA (0.96 g, 7.4 mmol). The resulting mixture was subjected to brief sonication to disperse the resin and then agitated on a shaker for 2.0 hours. The modified resin was collected by filtration, washed with DMF (2x), MeOH (2x), CHCl2 (2x), and dried in vacuo for 18 hours to yield 10.5 g (loading corresponds to approximately 1 mmol/g resin).
N-r3-PhenoxYpropyll-O-(phenylmethyl)-N~-rN-r r (phenylmethyl)aminolcarbonyll-L-histidyl]-L-tyrosinamide {BnNHCO-His-Tyr(OBn)-CONH(CH2)30Ph}
Fmoc-His(2-Cl-Tr-Resin)-Tyr(OBn)-C02CH2CH2TMS
(from Example 3 above, 2.0 g) was suspended in 20 piperidine in DMF. The resulting suspension was subjected to sonication for 10 minutes and then agitated by shaking for 30 minutes. The resin was filtered and washed with DMF (3x). The resin was again subjected to the same reaction conditions for an PCT~S94111553 ?~1~766 additional 20 minutes. The resin was filtered and washed with DMF (4x) and CHCl3 (3x) to provide His(2-Cl-Tr-Resin)-Tyr(OBn)-C02CH2CH2TMS which was suQpended in DCM (lO mL), agitated by shaking for 30 minutes, treated with benzyl isocyanate (l.l g, 8.0 mmol), and agitated for an additional 30 minutes.
The resin was filtered, washed with DCM (3x), resuspended in DCM, and the benzyl isocyanate treatment was repeated. The resin was filtered and washed with DMF (2x) and CHCl3 (5x) to give BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn)-C02CH2CH2TMS which was next suspended in a mixture of 4:3 dioxane:MeOH (14 mL) and treated with l.O M TBAF in THF (2.0 mL, 2.0 mmol). The suspension was agitated by shaking for 18 hours, filtered, washed sequentially with a 2:l mixture of dioxane and lO~ citric acid (3x lO mL), dioxane:MeOH
(3x lO mL), dioxane (3x lO mL), and CHCl3 (3x lO mL) to provide BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn). The BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn) was suspended in DMF
(lO mL) and treated with a carbodiimide coupling reagent such as DIC (0.2 g, l.6 mmol) and HOBT (0.22 g, l.6 mmol). The resulting mixtuEe was stirred 30 minutes and 3-phenoxypropylamine (0.24 g, 1.6 mmol) was added. The resulting mixture was shaken 18 hours before filtering the resin and washing with DMF (3x) and CHCl3 (3x). The resin was suspended in DMF ~lO mL) and the carbodiimide/HOBT/3-phenoxypropylamine coupling reaction was repeated. After 18 hours, the resin was filtered and washed with lO mL each of MeOH (2x), DCM
(3x), DMF (2x), MeOH (2x), and CHCl3 (2x) to give BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn)-CONH(C~2)30Ph. The highly substiuted dipeptide was cleaved from the resin by treatment with 40~ TFA in DCM, shaking for l hour at room temperature. The supernate, cont~in;ng the free dipeptide, was filtered away from the resin and the resin was washed with DCM (6x). The combined supernate PCT~S94/11553 ~ ~ l 7~66 and washings were concentrated in vacuo to pro~ide BnNHCO-His-Tyr(OBn)-CONH(CH2)30Ph TFA. The product was partitioned between water and DCM, and both layers were treated dropwise with saturated aqueous NaHCO3 until the aqueous layer r~m~;ned basic. The layers were separated, and the organic phase was washed with saturated aqueous NaCl and dried (MgSO4). Filtration and concentration yielded BnNHCO-His-Tyr(OBn)-CONH(CH2)30Ph; ES-M~ 675 (m+1).
Mul~iple, Simultaneous Solid Phase Synthe~is The method described in Example 4 may be employed in simultaneous multiple syntheses using the Diversomer apparatus described by S.H. DeWitt, et al., Proc. Natl.
Acad. Sci. USA, 90:6909 (1993). Fmoc-D-His(2-Cl-Tr Resin)-Tyr(OBn)-CO2iPr, prepared according to from Example 3 by substituting Fmoc-D-His(Tr) for Fmoc-His(Tr) in Step 3, (100-200 mg) is placed in each of 40 ~as dispersion tubes, and the tubes are placed in the Diversomer apparatus. The sequential deprotection and coupling reactions described in Example 4 are followed, employing the following acylating agents and ~m; neS in various combinations:
Acylatinq aqents Amines 1) benzyl isocyanate 1) 3-pheno~y~lu~ylamine 2) p-toluenesulfonyl chloride 2) 2-(phenylmethoxy)ethyl amine 3) cyclohexyl isocyanate 3) 2-[(phenylmethyl)-thio]-ethylamine 4) phenyl isocyanate 4) 4-phenylbutylamine 5) i-propyl isocyanate 5) 3-(2-methoxyphenyl)-propylamine 6) n-butyl isocyanate 6) 1-benzyl piperazine 7) 4-chlorophenyl isocyanate 7) o-benzyl-hy~o~ylamine 8) l-napthyl isocyanate 8) methionine methyl ester 9) 3-methoxypropyl isocyanate 9) benzylamine
For preparing pharmaceutical compositions from the compounds of the present invention, pharmaceutically acceptable carriers can be either solid or liquid.
Solid form preparations include powders, tablets, pills, capsules, cachets, suppositories, and dispersible granules. A solid carrier can be one or more substances which may also act as diluents, flavoring agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material.
pcT~ss~ 553 ~ 2 1 70766 In powders, the carrier is a finely divided solid which is in a mixture with ~he finely di~ided active component.
In tablets, the active component is mixed with the carrier having the necessary binding properties in suitable proportions and compacted in the shape and size desired.
The powders and tablets preferably contain from 5 or lO to about 70 percent of the active compound.
Sui~able carriers are magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter, and the like. The term "preparation" is intended to include the formulation of the active compound with encapsulating material as a carrier providing a capsule in which the active component with or without other carriers, is surrounded by a carrier, which is thus in association with it. Similarly, cachets and lozenges are included. Tablet~, powders, capsules, pills, cachets, and lozenges can be used as solid dosage forms suitable for oral ~m; n; gtration.
For preparing suppositories, a low melting wax, such as a mixture of fatty acid glycerides or cocoa butter, is first melted and the active component is dispersed homogeneously therein, as by stirring. The molten homogenous mixture i5 then poured into convenient sized molds, allowed to cool, and thereby to ~olidify.
Liquid form preparations include solutions, suspensions, and emulsions, for example, water or water propylene glycol solutions. For parenteral injection liquid preparations can be formulated in solution in aqueous polyethylene glycol solution.
Aqueous solutions suitable for oral use can be prepared by dissolving the active component in water WO9S112612 PCT~S9~111553 .
1 ~1 6 ~ -28-and adding suitable colorants, flavors, stabilizing and thickening agents as desired.
Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, and other well-known suspending agents .
Also included are solid form preparations which are intended to be converted, shortly before use, to liquid form preparations for oral A~m; n; stration. Such liquid forms include solutions, suspensions, and emulsions. These preparations may contain, in addition to the active component, colorants, flavors, stabilizers, buffers, artificial and natural sweeteners, dispersants, thickeners, solubilizing agents, and the like.
The phAr~Aceutical preparation is preferably in unit dosage form. In such ~orm the preparation is subdivided into unit doses contA;n;ng appropriate quantities of the active component. The unit dosage form can be a packaged preparation, the package contA i nl ng discrete quantities of preparation, such as packeted tablets, capsules, and powders in vials or ampoules. Also, the unit dosage form can be a capsules, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form.
The quantity of active component in a unit dose preparation may be varied or adjusted from O.l mg to lO0 mg preferably 0.5 mg to lO0 mg according to the particular application and the potency of the active component. The composition can, if desired, also contain other compatible therapeutic agents.
In therapeutic use as inhibitors of PFT, the compounds utilized in the phanmaceutical methods of WO 95/1~612 _, ~ 6 6 this invention are administered at the initial dosage of about 0.01 mg/kg to about 20 mg/kg daily. A daily dose range of about 0.01 mg/kg to about 10 mg/kg is preferred. The dosages, however, may be varied depending upon the requirements of the patient, the severity of the condition being treated, and the compound being employed. Determination of the proper dosage for a particular situation is within the skill of the art. Generally, treatment is initiated with smaller dosages which are less than the optimum dose of the compound. Thereafter, the dosage is increased by small increments until the optimum effect under the circumstances is reached. For convenience, the total daily dosage may be divided and ~m~ n; stered in port:ions during the day, if desired.
The following nonlimiting examples illustrate the inventors' preferred methods for preparing the compounds of the invention. For added clarity, complex chemical names describing compounds of Formula I are followed by structural abbreviations, which are shown in braces, wherein the structural elements are as defi~ed in the Table of Abbreviations above.
~-rN-rN-[(Phenylmethoxy)carbonyl]-L-histidyll-O-(phenylmethyl)-L-tyrosyl]-O-(phenylmethyl)-L-serinamide ~Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2}
Using an ABI model 431A solid phase peptide synthesizer, Fmoc-NH2-Rink resin (0.25 mmol scale) was treated with 20~ piperidine in NMP to afford NH2-Rink resin. Sequential coupling of Fmoc-protected Ser(OBn) and Tyr(OBn) (DCC and HOBT in NMP) and Fmoc deprotection (20~ piperidine in NMP) reactions were run using a 4-fold excess of reagents in the coupling steps and traditional resin washing cycles to afford Tyr(OBn)-Ser(OBn)-CONH2-Rink resin. This dipeptide PCT~S94/11553 ~lQ16~ ' --resin was transferred to an uninstrumented reaction vessel and treated with a 4-fold excess of Cbz-His, DCC, and HOBT in DMF, shaking overnight at room temperature. After removal of excess reagents, the resulting substituted tripeptide was cleaved from the resin by treatment with 50~ TFA in DCM at room temperature for 2.5 hours. Evaporation of solvents and purification by reversed phase chromatography (Cl8-column, eluted with a 20-70~ gradient of MeCN in water (both solvents acidi~ied with 0.l~ TFA) afforded Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2 as its TFA salt upon lyophilization. ES-MS: 719 (m+l).
Using analogous methods the following most preferred compounds of Formula I with carboxamides at the C-terminus may be prepared:
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-DL-(~-Me)Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONH2; and BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONH2.
N-rN-rN-[(Phenylmethoxy)carbonyll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-serine {Cbz-His-Tyr(OBn)-Ser(OBn)}
Beginning with PAM resin or HMP resin, Fmoc-Ser(OBn), Fmoc-Tyr(OBn), and Cbz-His are sequentially coupled using the deprotection and coupling conditions described in Example l. Cleavage from the resin is accomplished by treatment with CF3SO2H for the PAM supported tripeptide or with 50 TFA in DCM for the HMP supported tripeptide.
Chromatography as in Example l provides Cbz-His-Tyr(OBn)-Ser(OBn) as its TFA salt upon PCT~S94/11553 W095/12612 2 1 ~ ~66 .
lyophilization. See also Example 7 for a solution phase method.
Using analogous methods the following most preferred compounds of Formula I with a free carboxyl term; m~ may be prepared:
Cbz-D-His-Tyr(OBn)-Ser(OBn);
Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn);
Cbz-D-His(l-Me)-Tyr(OBn)-Ser(O~n);
Cbz-Hi~-D~-(~-Me)Tyr(OBn)-Ser(OBn3;
BnNHCO-His-Tyr(OBn)-Ser(OBn); and BnNHCO-D-His-Tyr(OBn)-Ser(OBn).
Solid phase supported N-~N-~(9H-Fluoren-9-ylmethoxy)car~onYl]-L-histidyll-O-(phenylmethyl)-L-tyrosine, 2-trimethylsilylethyl ester {Fmoc-His(2-Cl-Tr Resin)-Tyr(OBn)-CO2CH2CH2TMS}
Step 1: Boc-Tyr(OBn)-C02CH2CH2TMS
2-Trimethylsilyl ethanol (2.6 g, 22.6 mmol) was added to a pr~m~ solution of EDAC (4.3 g, 22.6 mmol), DMAP (0.5 g), and Boc-Tyr(OBn)-OH (7.0 g, 18.8 mmol) in dry THF (25 mL). The resulting mixture was stirred for 18 hours at room temperature. The solu~ion was diluted with 1:1 EtOAc:Et20 (40 mL), 2~ washed with saturated aqueous NaHC03 (2 x 10 m~) and with saturated aqueous NaCl (2 x 10 mL), dried (MgSO4), filtered and concentrated in vacuo to provide an oil which was further purified by flash chromatography (SiO2, EtOAc:hexane eluent) to give the pure TM~-ethyl ester as an oil;
H NMR (HCDCl3): ~ 0.04 (s, 9H), 1.43 (8, 9H), 3.03 (m, 2H), 4.22 (m, 2H), 4.51 (m, lH), 4.95 (m, lH), 5.05 (s, 2H), 6.85-7.48 (m, 9H).
PCT~S94111553 ~7`~76~ --Step 2: Tyr(OBn)-CO2CH2CH2TMS
Eighty percent TFA in CH2C12 (20 mL, v/v) was added to an ice-cooled solution of Boc-Tyr(OBn)-CO2CH2CH2TMS (14.8 g, 31.4 mmol) in CH2Cl2 (40 mL). The reQulting mixture was stirred for 1.0 minute before concentrating in vacuo. The procedure was repeated once more, and the resulting residue was diluted with CH2Cl2 and saturated aqueous NaHCO3. The resulting mixture was filtered through celite. The organic layer was then separated, washed with saturated aqueous NaCl, and dried (MgSO4). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:MeOH eluent) to give the de~ired product;
1H NMR (HCDC13): ~ 0.06 (s, 9H), 1.68 (br ~, 2H), 2.85-3.03 (m, 2H), 3.67 (m, lH), 4.22 (m, 2H), 5.05 (s, 2H), 6.91-7.45 (m, 9H).
Step 3: Fmoc-His(Tr)-Tyr(OBn)-CO2CH2CH2TMS
To a solution of HOBT (2.6 g, 19.3 mmol) in DMF
(10 mL) was added Fmoc-His(Tr) ( 10.0 g, 16.1 mmol) followed by EDAC (3.7 g, 19.3 mmol). The mixture was stirred at room temperature for 20 minutes before adding a solution of Tyr(OBn?-CO2CH2CH2TMS (from Step 2 above, 5.8 g, 16.1 mmol) in DMF (10 mL). The mixture was stirred overnight at room temperature before partitioning between a mixture of water and 1:1 Et2O:EtOAc (50 mL). The layers were separated, and the organic phase was washed with saturated aqueous NaCl (4 x 20 mL) and dried (MgSO4). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:MeOH eluent) to give the protected His-Tyr dipeptide; FAB-MS 974 (m+1).
W095/12612 PCT~S94/11553 2 1 7~66 Step 4: Fmoc-His-Tyr(OBn)-CO2CH2CH2TMS
Fmoc-His(Tr)-Tyr(OBn)-CO2CH2CH2TMS (from Step 3 above, 5.0 g, 5.1 mmol) was treated with pyridineoHCl (1.0 g) in MeOH (20 mL). The mixture was allowed to stir 8 hours at 65 C. The solution was concentrated in vacuo, and the residue was dissolved in CH2Cl2, washed with H20 (lx), saturated aqueous NaHC03 (2x), and dried (MgS04). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:MeOH eluent) to give Fmoc-His-Tyr(OBn)-CO2CH2CH2STMS as a white solid;
FAB-MS 731 (m+1).
Step 5: Fmoc-His(2-Cl-Tr Resin)-Tyr(OBn)-CO2CH2CH2TMS
To a suspension of Fmoc-His-Tyr(OBn)- C02CH2CH2TM~
(from Step 4 above, 5.3 g, 7.3 mmol) in CHC13 (20 mL) was added 2-chloroltrityl chloride resin (Novabiochem) (7.1 g) followed by DIEA (0.96 g, 7.4 mmol). The resulting mixture was subjected to brief sonication to disperse the resin and then agitated on a shaker for 2.0 hours. The modified resin was collected by filtration, washed with DMF (2x), MeOH (2x), CHCl2 (2x), and dried in vacuo for 18 hours to yield 10.5 g (loading corresponds to approximately 1 mmol/g resin).
N-r3-PhenoxYpropyll-O-(phenylmethyl)-N~-rN-r r (phenylmethyl)aminolcarbonyll-L-histidyl]-L-tyrosinamide {BnNHCO-His-Tyr(OBn)-CONH(CH2)30Ph}
Fmoc-His(2-Cl-Tr-Resin)-Tyr(OBn)-C02CH2CH2TMS
(from Example 3 above, 2.0 g) was suspended in 20 piperidine in DMF. The resulting suspension was subjected to sonication for 10 minutes and then agitated by shaking for 30 minutes. The resin was filtered and washed with DMF (3x). The resin was again subjected to the same reaction conditions for an PCT~S94111553 ?~1~766 additional 20 minutes. The resin was filtered and washed with DMF (4x) and CHCl3 (3x) to provide His(2-Cl-Tr-Resin)-Tyr(OBn)-C02CH2CH2TMS which was suQpended in DCM (lO mL), agitated by shaking for 30 minutes, treated with benzyl isocyanate (l.l g, 8.0 mmol), and agitated for an additional 30 minutes.
The resin was filtered, washed with DCM (3x), resuspended in DCM, and the benzyl isocyanate treatment was repeated. The resin was filtered and washed with DMF (2x) and CHCl3 (5x) to give BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn)-C02CH2CH2TMS which was next suspended in a mixture of 4:3 dioxane:MeOH (14 mL) and treated with l.O M TBAF in THF (2.0 mL, 2.0 mmol). The suspension was agitated by shaking for 18 hours, filtered, washed sequentially with a 2:l mixture of dioxane and lO~ citric acid (3x lO mL), dioxane:MeOH
(3x lO mL), dioxane (3x lO mL), and CHCl3 (3x lO mL) to provide BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn). The BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn) was suspended in DMF
(lO mL) and treated with a carbodiimide coupling reagent such as DIC (0.2 g, l.6 mmol) and HOBT (0.22 g, l.6 mmol). The resulting mixtuEe was stirred 30 minutes and 3-phenoxypropylamine (0.24 g, 1.6 mmol) was added. The resulting mixture was shaken 18 hours before filtering the resin and washing with DMF (3x) and CHCl3 (3x). The resin was suspended in DMF ~lO mL) and the carbodiimide/HOBT/3-phenoxypropylamine coupling reaction was repeated. After 18 hours, the resin was filtered and washed with lO mL each of MeOH (2x), DCM
(3x), DMF (2x), MeOH (2x), and CHCl3 (2x) to give BnNHCO-His(2-Cl-Tr-Resin)-Tyr(OBn)-CONH(C~2)30Ph. The highly substiuted dipeptide was cleaved from the resin by treatment with 40~ TFA in DCM, shaking for l hour at room temperature. The supernate, cont~in;ng the free dipeptide, was filtered away from the resin and the resin was washed with DCM (6x). The combined supernate PCT~S94/11553 ~ ~ l 7~66 and washings were concentrated in vacuo to pro~ide BnNHCO-His-Tyr(OBn)-CONH(CH2)30Ph TFA. The product was partitioned between water and DCM, and both layers were treated dropwise with saturated aqueous NaHCO3 until the aqueous layer r~m~;ned basic. The layers were separated, and the organic phase was washed with saturated aqueous NaCl and dried (MgSO4). Filtration and concentration yielded BnNHCO-His-Tyr(OBn)-CONH(CH2)30Ph; ES-M~ 675 (m+1).
Mul~iple, Simultaneous Solid Phase Synthe~is The method described in Example 4 may be employed in simultaneous multiple syntheses using the Diversomer apparatus described by S.H. DeWitt, et al., Proc. Natl.
Acad. Sci. USA, 90:6909 (1993). Fmoc-D-His(2-Cl-Tr Resin)-Tyr(OBn)-CO2iPr, prepared according to from Example 3 by substituting Fmoc-D-His(Tr) for Fmoc-His(Tr) in Step 3, (100-200 mg) is placed in each of 40 ~as dispersion tubes, and the tubes are placed in the Diversomer apparatus. The sequential deprotection and coupling reactions described in Example 4 are followed, employing the following acylating agents and ~m; neS in various combinations:
Acylatinq aqents Amines 1) benzyl isocyanate 1) 3-pheno~y~lu~ylamine 2) p-toluenesulfonyl chloride 2) 2-(phenylmethoxy)ethyl amine 3) cyclohexyl isocyanate 3) 2-[(phenylmethyl)-thio]-ethylamine 4) phenyl isocyanate 4) 4-phenylbutylamine 5) i-propyl isocyanate 5) 3-(2-methoxyphenyl)-propylamine 6) n-butyl isocyanate 6) 1-benzyl piperazine 7) 4-chlorophenyl isocyanate 7) o-benzyl-hy~o~ylamine 8) l-napthyl isocyanate 8) methionine methyl ester 9) 3-methoxypropyl isocyanate 9) benzylamine
10) 4 ethoxyphenyl isocyanate 10) 2-phenylethylamine
11) 2~phenethyl isocyante PCTrUS9~/11553 ~7~ 36-
12) 3-phenylpropionyl chloride 14) phenylacetyl chloride 15) 4-phenoxyphenyl isocyanate 16) benzyl chloroformate 17) (trans)-2-phenylcyclopropyl isocyanate 18) l-adamantyl chloroformate Array 1. Following cleavage from the resin and work-up as described in Example 4, the following substituted dipeptides (1-40) o~ Formula I are prepared:
1. PhNHCO-D-His-Tyr(OBn)-CONHCH2CH20Bn 2. PhNHCO-D-His-Tyr(OBn)-CONHCH2CH2SBn BS-~S 677 (m+l) 3. PhNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH20Ph BS-MS 661 (m+1) 4. BuNHCO-D-His-Tyr(OBn)-CONH(CH2)4Ph ES-NS 639 (m+1) 5. BuNHCO-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl) BS-MS 665 (m) 6. (4-MePh)SO2-D His-Tyr(OBn)-CONHCH2CH2Ph BS-MS 666 (m+l) 7. (4-MePh)S02-D-His-Tyr(OBn)-CONHCH2CH2SBn BS-MS 712 (m+l) 8. CF3S02-D-His-Tyr(OBn)-CONH-Met-C02Me 9 CF3S02-D-His-Tyr(OBn)-CONHCH2CH2CH2-(2-MeO-Ph) 10. CF3S02-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) 11 BnNHCO-D-His-Tyr(OBn)CONHCH2cH2Ph 12. MeO(CH2~3NHCO-D-His-Tyr(OBn)CON~ORn
1. PhNHCO-D-His-Tyr(OBn)-CONHCH2CH20Bn 2. PhNHCO-D-His-Tyr(OBn)-CONHCH2CH2SBn BS-~S 677 (m+l) 3. PhNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH20Ph BS-MS 661 (m+1) 4. BuNHCO-D-His-Tyr(OBn)-CONH(CH2)4Ph ES-NS 639 (m+1) 5. BuNHCO-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl) BS-MS 665 (m) 6. (4-MePh)SO2-D His-Tyr(OBn)-CONHCH2CH2Ph BS-MS 666 (m+l) 7. (4-MePh)S02-D-His-Tyr(OBn)-CONHCH2CH2SBn BS-MS 712 (m+l) 8. CF3S02-D-His-Tyr(OBn)-CONH-Met-C02Me 9 CF3S02-D-His-Tyr(OBn)-CONHCH2CH2CH2-(2-MeO-Ph) 10. CF3S02-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) 11 BnNHCO-D-His-Tyr(OBn)CONHCH2cH2Ph 12. MeO(CH2~3NHCO-D-His-Tyr(OBn)CON~ORn
13. MeO(CH2)3NHCO-D-His-Tyr(OBn)CONHCH2CH2CH20Ph
14. MeO(CH2)3NHCO-D-His-Tyr(OBn)CONH(CH2)3-(2-MeO-Ph)
15. BnNHCO-D-His-Tyr(OBn)CONHCH2Ph BS-MS 631 (m+1)
16. (4-ClPh)NHCO-D-His-Tyr(OBn)CONHCH2CH20Bn BS-MS 695 (m+1)
17. l-Napthyl-NHCO-D-His-Tyr(OBn)C~N~ORn ES-~S 683 (m+1)
18. 1-Napthyl-NHCO-D-His-Tyr(OBn)CONH-Met-C02Me ES-MS 723 (m+l)
19. (4-ClPh)NHCO-D-His-Tyr(OBn)CONH(CH2)4Ph ES-MS 693 (m+l)
20. (4-ClPh)NHCO-D-His-Tyr(OBn)CONHCH2Ph ES-MS 651 (m+1)
21. BnOCO-D-His-Tyr(OBn)CONHCH2CH20Bn ES-MS 676 (m+l)
22. 1-adamantyl-OCO-D-His-Tyr(OBn)CONHCH2CH2SBn
23. BnOCO-D-His-Tyr(OBn)CONHCH2CH2CH20Ph ES-MS 676 (m+1)
24. 1-adamantyl-OCO-D-His-Tyr(OBn)CONHCH2CH2CH2CH2Ph
25. BnOCO-D-His-Tyr(OBn)CO(4-Bn-piperazin-1-yl) ES-MS 700 (m)
26. PhCH2CO-D-His-Tyr(OBn)CONHCH2CH2Ph ES-MS 630 (m+1)
27. PhCH2CH2NHCO-D-His-Tyr(OBn)CONHCH2CH2SBn BS-MS 705 (m+1) PCTrUS9~/11553
28. PhCH2CH2NHCO-D-His-Tyr(OBn)CONH-Met-CO2Me . ES-MS 701 (m+l)
29. PhCH2CH2NHCO-D-His-Tyr(OBn)CONH(CH2)3-(2-MeO-Ph) ES-MS 703 (m+1)
30. PhCH2CO-D-His-Tyr(OBn)CO(4-Bn-piperazin-1-yl) ES-MS 684 (m)
31. (t-2-Ph-c-propyl)-NHCO-D-His-Tyr(OBn)CONHCH2CH2Ph ES-MS 671 (m+l)
32. (t-2-Ph-c-propyl)-NHCO-D-His-Tyr(OBn)CONUoRn ES-MS 673 (m+l)
33. c-hexyl-NHCO-D-His-Tyr(OBn) CONH(CH2)3Oph ES-MS 666 (m+1)
34. c-hexyl-NHCO-D-His-Tyr(OBn)CONH(CH2)3-(2-MeO-Ph) ES-MS 681 (m+l)
35. c-hexyl-NHCO-D-His-Tyr(OBn)CONHCH2Ph ES-MS 623 (m+l)
36. PhCH2CH2CO-D-His-Tyr(OBn)CONHCH2CH2OBn ES-MS 674 (m+1)
37. PhCH2CH2CO-D-His-Tyr(OBn)C~N~ORn ES-MS 646 (m+l)
38. (CH3)2CHNHCO-D-His-Tyr(OBn)CONH-Met-CO2Me ES-MS 639 (m+1)
39. PhCH2CH2CO-D-His-Tyr(OBn)CoNH(cH2)3ph ES-MS 672 (m+l)
40. (CH3)2CHNHCO-D-His-Tyr(OBn)CONHCH2Ph ES-MS 583 (m+l) Array 2. Following cleavage from the resin and work-up as described in Example 4, the following substi~uted dipeptides (41-80) of Formula I are prepared:
41. n-BuNHCO-D-HiR-Tyr(OBn)-CONHCH2CH2OBn ES-MS 641 (m+1)
42. n-BuNHCO-D-HiR-Tyr(OBn)-CONHCH2CH2SBn ES-MS 657 (mll)
43. n-BuNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH2OPh ES-MS 641 (m+1)
44. Ph~HCO-D-His-Tyr(OBn)-CONHCH2CH2CH2CH2Ph ES-MS 659 (m+1)
45. PhNHCO-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl)ES-MS 685 (m)
46. (4-PhOPh)NHCO-D-His-Tyr(OBn)-CONHCH2CH2Ph ES-MS 723 (m+l)
47. (4-PhOPh)NHCO-D-His-Tyr(OBn)-CONHCH2CH2SBn ES-MS 769 (m+1)
48. (4-MePh)SO2-D-His-Tyr(OBn)-CONH-Met-CO2Me ES-MS 708 (m+1)
49. (4-MePh)SO2-D-His-Tyr(OBn)-CONHCH2CH2CH2-(2-MeO-Ph)
50. (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) ES-MS 720 (m)
51. MeO(CH2)3NHCO-D-His-Tyr(OBn)CONHCH2CH2Ph
52. BnNHCO-D-His-Tyr(oBn)cnN~oR~ ES-MS 647 (m+1)
53. BnNHCO-D-His-Tyr(OBn)CONHCH2CH2CH2OPh ES-~S 675 (m+l) PC~rUS94111553 6~ _
54. BnNHCO-D-His-Tyr(OBn)CONH(CH2)3-(2-MeO-Ph) ES-MS 689 (m+l)
55. MeO(CH2)3NHCO-D-His-Tyr(OBn)CONHCH2Ph
56. 1-napthyl-NHCO-D-His-Tyr(OBn)CONHCH2CH2OBn ES-MS 711 (m+l)
57. (4-ClPh)NHCO-D-His-Tyr(oBn)coNu~Rn
58. (4-ClPh)NHCO-D-His-Tyr(OBn)CONH-Met-CO2Me BS-MS 707 (m+l)
59. 1-napthyl-NHCO-D-His-Tyr(OBn)CONHCH2CH2CH2CH2Ph BS-MS 709 (m+1)
60. 1-napthyl-NHCO-D-His-Tyr(OBn)CONHCH2Ph
61. (4-EtOPh)NHCO-D-Hi 8 -Tyr(OBn)CONHCH2CH2OBn ES-MS 705 (m+l)
62. BnOCO-D-His-Tyr(OBn)CONHCH2CH2SBn
63. (4-BtOPh)NHCO-His-Tyr(OBn)CONHCH2CH2CH2OPh ES-MS 705 (m+l)
64. BnOCO-D-His-Tyr(OBn)CONHCH2CH2CH2CH2Ph
65. (4-EtOPh)NHCO-D-His-Tyr(OBn)CO(4-Bn-piperazin-1-yl) ES-MS 729 (m)
66. PhCH2CO-D-His-Tyr(OBn)CONHCH2CH2Ph ES-MS 659 (m+1)
67. PhCH2CO-D-His-Tyr(OBn)CONHCH2CH2SBn ES-MS 676 (m+l)
68. PhCH2CO-D-His-Tyr(OBn)CONH-Met-CO2Me
69. PhCH2CO-D-His-Tyr(OBn)CONH(CH2)3-(2-MeO-Ph) ES-MS 674 (m+l)
70. PhCH2CH2NHCO-D-His-Tyr(OBn)CO(4-Bn-piperazin-1-yl) ES-MS 713 (m)
71. c-hexyl-NHCO-D-His-Tyr(OBn)CONHCH2CH2Ph BS-NS 637 (m+l)
72. c-hexyl-NHCO-D-His-Tyr(oBn)CoN~nRn ES-NS 639 (m+l)
73. (t-2-Ph-c-propyl)-NHCO-D-His-Tyr(OBn)CONH(CH2)30Ph ES-MS 701 (m+l)
74. (t-2-Ph-c-propyl)-NHCO-D-His-Tyr(OBn)CONH(CH2)3-(2-MeO-Ph)
75. (t-2-Ph-c-propyl)-NHCO-D-His-Tyr(OBn)CONHCH2Ph ES-NS 657 (m+l)
76. (CH3)2CUNUCO-D-His-Tyr(OBn)CONHCH2CH2OBn ES-NS 627 (m+l)
77- (cH3)2cH~Hco-D-His-Tyr(oBn)c~N~oRn ES-MS 599 (m+l)
78. PhCH2CH2CO-D-His-Tyr(OBn)CONH-Met-CO2Me ES-MS 686 ~m+l)
79. (CH3)2CHNHCO-D-His-Tyr(OBn)CONHCH2CH2CH2CH2Ph ES-~S 625 (m+l)
80. PhCH2CH2CO-D-His-Tyr(OBn)CONHCH2Ph ES-~S 630 (m+l) PCT~S94111553 2 t 70766 N-rN-rN- r (Phenylmethoxy)carbonyll-L-histidyl]-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-qerine, methyl ester {Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me}
Step 1: Boc-Tyr(OBn)-Ser(OBn)-CO2Me To a solution of Boc-Tyr(OBn) (1.88 g, 6.50 mmol) in EtOAc ~30 mL) at 0C was added HOBT hydrate (1.19 g, 7.80 mmol) followed by DCC (1.61 g, 7.80 mmol). A
solution of Ser(OBn)-CO2Me TFA (2.1 g, 6.50 mmol) in EtOAc (20 mL) was added followed by Et3N (1.09 mL, 7.80 mmol). The mixture was allowed to warm to room temperature and stirred overnight. The mixture was filtered, diluted with EtOAc, and washed twice with saturated aqueous NaHC03, brine, dried over MgSO4, and concentrated. Flash chromatography (40~ EtOAc/hexane) gave 2.67 g (73~) of the title compound as a white solid, mp 81-84C.
Step 2: Tyr(OBn)-Ser(OBn)-CO2Me TFA
Boc-Tyr(OBn)-Ser~OBn)-C02Me (from Step 1 above, 2.64 g, 4.69 mmol) was dissolved in CH2C12 (15 mL), cooled to 0C and TFA (5 mL) was added. The ~olution was warmed to room temperature and stirred for 4 hours.
The solution was concentrated, taken up in CH2Cl2 and reconcentrated twice. The resulting oil was triturated with ether to provide 2.7 g of the title compound as a white solid.
Step 3: Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me To a solution of Cbz-His (1.00 g, 3.47 mmol) in DMF (15 mL) at 0C was added HOBT (0.64 g, 4.16 mmol) and DCC (0.86 g, 4.16 mmol). Tyr(OBn)-Ser(OBn)-CO2Me TFA (from Step 2 above, 2.0 g, 3.47 mmol) was added followed by Et3N (0.58 mL, 4.16 mmol). The mixture was allowed to warm to room temperature and stirred overnight. The mixture was filtered and the PCT~S94tllS53 %~ 40-filtrate was diluted with CHC13, washed twice with saturated aqueous NaHCO3, brine, dried over MgSO4, and concentrated. Flash chromatography (2-5~ MeOH/CHCl3) gave 2.14 g of the title compound as a white solid, mp 175-176C; FAB-MS 734 (m~1);
Anal. Calc. for C4lH43N58 C, 67.11; H, 5.91; N, 9.54;
Found: C, 66.96; H, 6.01; N, 9.41.
N-[N-rN-~(Phenylmethoxy)carbonyl]-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-serine {Cbz-His-Tyr(OBn)-Ser(OBn)}
To a solution of Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me (from Example 6 above, 2.02 g, 2.75 mmol) in THF
(50 m~) and MeOH (15 mL) at 0C was added 0.lN LiOH
(30.3 mL, 3.03 mmol). The solution was stirred for 6 hours at 0C, then concentrated. Water was added and the pH was adjusted to 4-5 with lN HCl. The mixture was filtered, and the solid was collected and dried to provide 1.55 g (78~) of the title compound as a white solid, mp 187-192C; ES-MS 720 (m+1);
Anal. Calc. for C40H4lN58 1 5H2O:
C, 64.33; H, 5.94; N, 9.38;
Found: C, 64.29; H, 5.73; N, 9.15.
EXAMP~E 8 N-rN-[N-[(Phenylmethoxy)carbonyl]-D-histidyll-O-(phenylmethyl)-L-tyrosyl]-O-(phenylmethyl)-L-serine, methyl ester ~Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me}
Step 1: Tyr(OBn)-Ser(OBn)-CO2Me HCl A solution of Boc-Tyr(OBn)-Ser(OBn)-CO2Me (from Bxample 6, Step 1 above, 9.90 g, 17.6 mmol) in EtOAc was cooled to 0C. Anhydrous HCl gas was bubbled through the cold solution for 5 minutes. The solution was allowed to warm to room temperature and stirred Wo 95/12612 ~ 21 70766 overnight. The solution was concentrated, taken up in EtOAc and reconcentrated to provide 8.75 g of the title compound as a foam; CI-MS 463 (m+1).
Step 2: Cbz-D-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me According to Example 6, Step 3, by substituting Cbz-D-His(Tr) for Cbz-His and Tyr(OBn)-Ser(OBn)-CO2Me HCl for Tyr(OBn)-Ser(OBn)-CO2Me TFA, the title compound was obtained as a white solid, mp 78-88C;
FAB-MS 976 (m+1).
Step 3: Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me A solution of Cbz-D-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me (from Step 2 above, 0.27 g, 0.28 mmol) in HOAc:H2O (4:1, 2 mL) was stirred at 80C for 5 minutes, then cooled to room temperature. The solution was partitioned between EtOAc and saturated aqueous NaHCO3.
The organic layer was washed with brine, dried (MgSO4), and concentrated. Flash chromatography (2-5~
MeOH:CHC13) yielded 0.10 g of the title compound as a foam; FAB-MS 734 (m+l).
N- rN- rN- r (Phenylmethoxy)carbonyll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-D-~erine, methyl ester {Cbz-His-Tyr(OBn)-D-Ser(OBn)-CO2Me}
According to Example 6, by substituting D-Ser(OBn)-CO2Me TFA for Ser(OBn)-CO2Me-TFA in Step 1, the title compound was obtained, mp 168-170C;
FAB-MS 734 (m+l).
PCT~S94/11553 z~3~ 42-N-r~-Methyl-N-rN- r (phenylmethoxy)carbonyll-L-hi tidyll-O-(phenylmethyl)-DL-tyrosyll-O-(phenylmethyl)-L-serine.
methyl ester {Cbz-His-DL-(~-Me)Tyr(OBn)-Ser(OBn)-CO
According to Example 6, by substituting Boc-DL-(~-Me)Tyr(OBn) for Boc-Tyr(OBn) in Step 1, the title compound was obtained; FAB-MS 748 (m+l).
N-Ethyl-N~-[N- rN- r (phenylmethoxy)carbonyl]-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-h-serinamide {Cbz-His-Tyr(OBn)-Ser(OBn)-CONHEt}
According to Example 6, by substituting Boc-Tyr(OBn)-Ser(OBn)-CONHEt for Boc-Tyr(OBn)-Ser(OBn)-CO2Me in Step 2, the title compound was obtained, mp 182-188C; FAB-MS 747 (m+l).
N-Ethyl-N~- rN- rN- r (Phenylmethoxy)carbonyll-D-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-serinamide {Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONHEt}
According to Example 6, by substituting Boc-Tyr(OBn)-Ser(OBn)-CONHEt for Boc-Tyr(OBn)-Ser(OBn)-CO2Me in Step 2 and Cbz-D-His for Cbz-His in Step 3, the title compound was obtained, mp 193-196C;
ES-MS 747 (m+l).
N-rN-rl-Methyl-N- r (phenylmethoxy)carbonylll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-Serine~
methyl ester {Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CO2Me}
Step 1: Cbz-His(l-Me) Benzyl chloroformate (0.24 mL, 1.7 mmol) was added dropwise to a slurry of l-methyl-~-histidine (0.25 g, WO9S/12612 P~T~S94111553 ~ 21 70766 1.5 mmol) in THF (5 mL) and saturated aqueous NaHCO3 (5 mL) at 0C. The mixture was allowed to warm to room temperature and stirred overnight. The mixture was concentrated and diluted with H2O, washed with ether, and the pH adjusted to 6-7 with lN HCl. The mixture was concentrated, then diluted with CHCl3 (150 mL) and MeOH (15 mL), and stirred for 1 hour. The mixture was dried (MgSO4) and concentrated to pro~ide 0.48 g of the title compound which was used without further purification.
Step 2: Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CO2Me To a slurry of Cbz-His(1-Me) (from Step 1 abo~e, 0.36 g, 1.2 mmol), Tyr(OBn)-Ser(OBn)-CO2Me HCl (from Example 8, Step 1 above, 0.60 g, 1.2 mmol), DCC
(0.30 g, 1.4 mmol), and HOBT (0.19 g, 1.4 mmol) in CH2Cl2 was added Et3N (0.17 mL, 1.2 mmol) and the mixture was stirred overnight at room temperature. The mixture was diluted with CHCl3, washed with saturated aqueous NaHCO3, brine, dried (MgSO4), and concentrated.
Flash chromatography (1~ MeOH:CHCl3) provided the title compound as a white solid, mp 161.5-163.5C; FA~3-MS 748 (m+1).
N-~M-rl-Methyl-N- r (phenylmethoxy)carbonyll-D-histidyll-O-(phenylmethyl)-L-tyrosyl]-O-(~henylmethyl)-L-serine.
methyl ester {Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-CO2Me}
According to Example 13, by substituting 1-methyl-D-histidine for 1-methyl-L-histidine, the title compound was obtained; FAB-MS 748 (m+1).
WO9S/12612 PCT~S94/11553 .
N-[L-2-Amino-N - rN - r ( Phenylmethoxy)carbonyll-D-histidyll-4-r4-(phenylmethoxy)phenyllbutanoyll-0-(phenylmethyl-L-serine methyl ester {Cbz-D-His-homoTyr(OBn)-Ser(OBn)-CO2Me}
According to Example 6, by substituting Boc-homoTyr(OBn) for Boc-Tyr(OBn) in Step 1, and substituting Cbz-D-His for Cbz-His in Step 3, the title compound was obtained; ES-MS 748 (m+l).
N-r4-Phenyl-N-rN- r (phenylmethoxy)carbonyll-L-histidyll-L-phenylalanyll-O-(phenylmethyl)-L-serine methyl ester {Cbz-His-Phe(4-Ph)-Ser(OBn)-CO2Me}
According to Example 6, by substituting Boc-Phe(4-Ph) for Boc-Tyr(OBn) in Step 1, the title compound was obtained, mp 184-187C; FAB-MS 704 (m+l).
N- ro- (Phenylmethyl)-NrN- r r (Phenylmethyl)aminol-carbonyll-L-histidyll-L-tyrosyll-O-(phenylmethyl)-serine, methyl ester {BnNHCO-His-Tyr(OBn)-Ser(OBn)-C02Me }
Step 1: Fmoc-His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me According to Example 13, Step 2, by substituting Fmoc-His(Tr) for Cbz-His(l-Me), the title compound was obtained, mp 82-92C.
Step 2: His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me Piperidine (4.0 mL) was added to a slurry of Fmoc-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me (from Step 1 above, 1.85 g, 1.74 mmol) in CH2C12 (20 mL). The solution was stirred for 2 hours at room temperature, then concentrated. The residue was taken up in EtOAc (150 mL), washed with water (3 x 50 mL), dried (MgS04), and concentrated. The resulting oil was triturated W O 95/12612 2 PCTrUS94/11553 with Et2O/hexane. Flash chromatography of the residue (2~ MeOH/CHC13) gave 1.03 g o~ the title compound as a foam, mp 61.5-70C; ES-MS 843 (m+1).
Ste~ 3: BnNHCO-His(Tr)-Tyr(OBn~-Ser(OBn)-CO2Me Benzyl isocyanate (0.053 mL, 0.43 mmol) was added in one portion to a solution of His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me (~rom Step 2 above, 0.33 g, 0.39 mmol) in EtOAc (5 mL). The resulting slurry was stirred for 3 hours at room temperature, then concentrated to yield the title compound (0.4 g), which was used without fur~her purification.
Step 4: BnNHCO-His-Tyr(OBn)-Ser(OBn)-CO2Me According to Example 8, by substituting BnNHCO-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me for Cbz-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me, the title compound was obtained, mp 19 6.5- 199 C; ES-MS 733 (m+1).
N-rN-rN-(1-Oxo-3-phenylpropvl)-L-histidyll-O-(phenYlmethyl)-L-tyrosyl]-O-(phenylmethyl)-L-serine methyl ester {PhCH2CH2CO-His-Tyr(OBn)-Ser(OBn)-C02Me}
Step 1: PhCH2CH2CO-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me To a cooled (0C) solution of His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me (from Example 17, Step 2 above, 0.33 g, 0.39 mmol) in THF (5 mL) was added Et3N (O. 06 mL, 0.43 mmol) followed by phenylpropionyl chloride (0.064 mL, 0.43 mmol). The resulting slurry was brought to room temperature and stirred overnight. The mixture was partitioned between EtOAc and saturated aqueous NaHCO3. The organic layer was washed with brine, dried (MgSO4), and concentrated to yield the title compound as a solid which was u~ed without ~urther purification.
wossll26l2 pcT~s9~ ss3 .
~1 31 ~ -46-Step 2: PhCH2CH2CO-His-Tyr(OBn)-Ser(OBn)-CO2Me According to Example 8, Step 3, by substituting PhCH2CH2CO-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me for Cbz-D-His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me, the title compound was obtA;ne~, mp 193-196.5C; ES-MS 732 (m+l).
~-rN-~(Phenylmethoxy)carbonyll-L-histidyl]-O-(~henylmethyl)-N-r2-(phenylmethoxy)ethyll-~-tyrosinamide {Cbz-His-Tyr(OBn)-CONHCH2CH20Bn}
Step 1: Cbz-His-Tyr(OBn)-CO2Me According to Example 6, Step 3, by substituting Tyr(OBn)-CO2Me TFA for Tyr(OBn)-Ser(OBn)-CO2Me TFA, the title compound was obtained as a white powder, mp 145-148C; CI-MS 557 (m+l).
Step 2: Cbz-His-Tyr(OBn) According to Example 7, by substituting Cbz-His-Tyr(OBn)-CO2Me ~or Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me, the title compound was obtained as a white powder, mp 79-92C; CI-MS 543 (m+l).
Step 3: Cbz-His -Tyr(OBn)-CONHCH2CH20Bn To a solution of Cbz-His-Tyr(OBn) (from Step 2 above, 0.43 g, O.79 mmol) in DMF (4 mL) at 0C was added HOBT (0.15 g, 0.95 mmol) and DCC (0.20 g, 0.95 mmol). A solution of 2-(phenylmethoxy)ethylamine (0.12 g, 0.79 mmol) in DMF (1 mL) was then added. The mixture was allowed to warm to room temperature and stirred overnight. The mixture was filtered, diluted with CHCl3, washed twice with saturated aqueous NaHC03, washed with brine, dried over MgSO4, and concentrated.
Flash chromatography (3-5~ MeOH/CHC13) afforded 0.34 g (63~) of the title compound as a white solid, mp 136-150C; FAB-MS 676 (m+l);
WO 95/12612 2 1 7 0 l 6 6 PCTIUS94/11553 Anal. Calc. :Eor C39H41N506:
C, 69.32; H, 6.12; N, 10.36;
Found: C, 69.43; H, 6.24; N, 10.45.
~- rN- r (Phenylmethoxy) carbonyl]-D-histidyl]-N-r 2-(phenylmethoxy)ethyll-O-(phenylmethyl)-L-tyrosin~nide {cbz-D-His-Tyr(oBn)-cor~HcH2~H2c:)Bn}
According to Example 6, by substituting 2- (phenyl-methoxy)ethylamine for Ser(OBn) TFA and omitting Et3N
in Step 1 and by substituting Cbz-D-His for Cbz-His in Step 3, the title compound was prepared, mp 161-165C;
FAB-MS 676 (m+1).
~a~ rN-Methyl-Nr(phenylmethoxy)carbonyl]-D-histidyll-N-r2-~phenylmethoxy)ethyll-0-(phenylmethyl)-L-tyrosinamide {Cbz-D-His-(N-Me)Tyr(OBn)-CONHCH2CH20Bn}
According to Example 20, by substituting Boc-(N-Me)Tyr(OBn) for Boc-Tyr(OBn), the title compound was obtained, mp 64-78C; ES-MS 690 (m+l).
~a-rcY-Methyl-N-rN-~(~henylmethoxy)carbonyll-D-histidyll-N-r2-(phenylmethoxy)ethyll-O-(phenyl-methyl)-L-tyrosin~mide {Cbz-D-His-(cY-Me)Tyr(QBn)-CONHCH2CH2OBn}
According to Example 20, by substituting Boc-(cY-Me)Tyr(OBn) for Boc-Tyr(OBn), the title compound was obtained, mp 66-78C; ES-MS 690 (m+1).
W095/12612 PCT~S91111553 N-(2-Phenylethyl~-N~-[N- r (phenylmethoxy)carbonyll-T-histidyll-O-(phenylmethyl)-L-tyrosinamide {Cbz-His-Tyr(OBn)-CONHCH2CH2Ph}
According to Example 19, Step 3, by substituting 2-phenylethylamine for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid, mp 188-189.5C; FAB-MS 646 (m+1).
N-rN-rN-r(Phenylmethoxy)carbonyll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-3-(3-pyridinyl)-L-alanine, methyl ester {Cbz-His-Tyr(OBn)-Pyr-CO2Me}
According to Example 19, Step 3, by substituting Pyr-CO2Me for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid, mp 180-182.5C
(dec); FAB-MS 705 (m+1).
(S.R)-N- r 2-(4-Benzyloxy-phenyl)-1-(3-phenoxy-propylcarbamoyl)-ethyll-2-r3-(4-ethoxy-phenyl)-ureidol-3-(3H-imidazol-4-yl)-propionamide {(4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)30Ph}
Step 1. Boc-Tyr(OBn)-CONH(CH2)30Ph 2-(Phenylmethoxy)ethylamine (0.81 g, 5.4 mmol) was added to a premixed solution of EDAC (1.2 g, 6.5 mmol), HOBT (0.87 g, 6.5 mmol), and Boc-Tyr(OBn)-OH (2.0 g, 5.4 mmol) in dry DMF (15 mL). The resulting mixture was stirred for 18 hours at room temperature. The solution was diluted with 1:1 EtOAc:Et20 (40 mL), washed with saturated aqueous NaCl (4 x 10 mL), dried (MgSO4), filtered, and concentrated in vacuo to provide a solid which was further purified by trituration with h~n~ to give the pure product, mp 145-146-C.
WO95/12612 PCT~S94/11553 21~0766 Step 2. Tyr(OLn)-CONH(CH2)3OPh Dry HCl gas was bubbled into an ice cold ~olution of Boc-Tyr(OBn)-CONH(CH2)3OPh (from Step l above, 2.0 g, 3.9 mmol) in MeOH (15 mL) for 4 minutes. The resulting mixture was stirred for l hour at 0 C and then allowed to warm to room temperature and stir l hour. The solution was concentrated in vacuo to provide a solid which was triturated with ether to provide Tyr(OBn)-CONH(CH2)3OPh HCl; CI-MS 405 (m+l).
The title compound was suspended in CHCl3, cooled in an ice bath, and NH3 gas was bubbled through the mixture for 2 minutes. The NH4Cl was filtered off, and the supernate was concentrated in vacuo to yield the free base of the title compound which was used in the next step without further purification.
Step 3. Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)3OPh To a solution of HOBT (0.48 g, 3.5 mmol) in DMF
(10 m~) was added Fmoc-D-His(Tr)-CO2H (2.0 g, 3.2 mmol) followed by EDAC (0.67 g, 3.5 mmol). The mixture was stirred at room temperature for 20 minutes before adding a solution of Tyr(OBn)-CONH(CH2)3OPh (from Step 2 above, 1.4 g, 3.2 mmol) in DMF (l0 mL). The mixture was stirred overnight at room temperature before partitioniny between a mixture of water (20 mL) and l:l Et2O:EtOAc (50 mL). The layers were separated, and the organic phase was washed with saturated aqueous NaCl (4x 20 mL) and dried (MgSO4). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:
MeOH eluent) to give the protected His-Tyr dipeptide;
FAB-MS 1006 (m).
Step 4. D-His(Tr)-Tyr(OBn)-CONH(CH2)3OPh Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph (from Step 3 above, 1.O g, 0.99 mmol) in CH2C12 (5 mL) was treated PCT~S94/1l553 Wo9SI12612 so- ' with piperidine (0.18 g, 2.1 mmol). The resulting mixture was stirred 2 hours before concentrating in vacuo and purifying the resulting oil by flash chromatography (SiO2, CHC13:MeOH eluent) to give (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph;
ES-MS 784 (m+l).
Step 5. (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2L30Ph D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph (from Step 4 above, 0.55 g, 0.7 mmol) in CH2Cl2 (5 mL) was treated with 4-ethoxyphenyl isocyanate (0.1 g, 0.7 mmol). The resulting mixture was stirred 1 hour at room temperature. Concentrated in vacuo. The resulting oil was purified by flash chromatography (SiO2, CHC13:MeOH
eluent) to give (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph; ES-MS 947 (m+1).
Step 6. (4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2L30Ph (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph (from Step 5 above, 0.5 g, 0.52 mmol) in MeOH (5 mL) was treated with Pyridine HCl (catalytic). The resulting mixture was stirred at 65 C for 6 hours.
Concentrated in vacuo to obtain an oil which was purified by by flash chromatography (SiO2, CHCl3:MeOH
eluent) to give (4-EtO-Ph)NHCO-D-His-Tyr(OBn)-CONH(CH2)30Ph, mp 185-187 C; ES-MS 705 (m+1).
(S.R)-N-{2-(4-Benzyloxy-phenyl~-1-r3-(2-methoxy-phenyl)-propylcarbamoyll-ethyl}-3-(3H-imidazol-4-yl)-2-phenylacetylamino-propionamide {PhCH2CO-D-His-Tyr(OBn)- CONH(CH2L3(2-MeOPh)}
Step 1. Boc-Tyr(OBn)-CONH(CH2)3(2-MeOPh) According to Example 25, Step 1, by substituting 3-(2-methoxyphenyl) propyl amine for 2-(phenylmethoxy)-WO 95/12612 ~ 1 ~ O ~ ~ 6 ethylamine, the title compound was obtained as a white solid, mp 125-126.5-C.
.
Stel? 2. Tyr(OBn)-CONH(CH2)3(2-MeOPh) According to Example 25, Step 2, by substituting Boc-Tyr(OBn)-CONH(CH2)3(2-MeOPh) for Boc-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white solid; CI-MS 419 (m+1).
Step 3. Fmoc-D-His(Tr)-Tyr(OBn)-CQNH(CH2)3(2-MeOPh) According to Example 25, Step 3, by substituting Tyr~OBn)-CONH(CH2) 3 ( 2-MeOPh) for Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a foam; ES-MS 1020 (m).
Step 4. D-His(Tr)-Tyr(OBn)-CONH(CH2)3(2-MeOPh) According to Example 25, Step 4, by substituting Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)3(2-MeOPh) for Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obt~;ne~ as a white foam; ES-MS 798 (m+1).
Step 5. PhCH2CO-D-His(Tr)-Tyr(OBn)-CONH(CH2) 3 (2-MeOPh) To a solution of D-His(Tr)-Tyr(OBn)-CONH(CH2)3(2-MeOPh) (0.4 g, 0.5 mmol) in CH2Cl2 (5 mL) was added N-methyl morpholine (0.05 g, 0.5 mmol) followed by phenyl acetyl chloride (0.08 g, 0.5 mmol).
The resulting mixture was stirred 2 hours at room temperature. Diluted with DCM and washed with saturated aqueous NaHC03, saturated aqueous NaCl, and dried (MgSO4). Purified by flash chromatography (SiO2, CHCl3:MeOH eluent). The title compound was obtained as a foam; FAB-MS 916 (m).
Step 6. PhCH2CO-D-His-Tyr(OBn)-CONH(CH2) 3 (2-MeOPh) According to Example 25, Step 6, by substituting PhCH2CO-D-His(Tr)-Tyr(OBn)-CONH(CH2) 3 (2-MeOPh) for WO9S/12612 PCT~S94111S53 .
(4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white foam; ES-MS 674 (m+1).
(S,R)-N-r2-(4-Benzyloxy-phenyl)-1-phenethylcarbamoyl-ethyl]-3-(3H-imidazol-4-yl)-2-r3-(4-phenoxy-phenyl)-ureidol-propionamide {(4-PhOPh)NHCO-D-His-Tyr(OBn)-CoNH(cH2L2ph}
Step 1. Boc-Tyr(OBn)CONH(CH2)2Ph According to Example 25, Step 1, by substituting phenethylamine for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid; CI-MS 475 (m+1).
Step 2. Tyr(OBn)-cONH(cH2L2Ph According to Example 25, Step 2, by substituting BocTyr(OBn)-CONH(CH2)2Ph for BocTyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white solid;
CI-MS 375 (m+1).
Step 3. Fmoc-D-His(Tr)-Tyr(OBn?-CONH(CH2L2Ph According to Example 25, Step 3, by substituting Tyr(OBn)-CONH(CH2)2Ph for Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a foam; ES-MS 977 (m+1).
Step 4. Fmoc-D-His-Tyr(OBn)-CONH(CH2L2Ph Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)2Ph (1.2 g, 1.6 mmol) in MeOH (5.mL) was treated with Pyridine HCl (catalytic). The resulting mixture was stirred at 65 C
overnight and concentrated in vacuo to obtain an oil which was purified by by flash chromatography (SiO2, CHC13:MeOH eluent) to give a white solid; ES-MS 734 (m+1).
PCT~S94111553 ~t~ 66 Step 5. (4-phoph)NHco-D-His-Tyr(oBn)-coNH(cH2L2ph Fmoc-D-His-Tyr(OBn)-CONH(CH2)2Ph (0.6 g, O.8 mmol) in CH2C12 (5 mL) was treated with piperdine (0.14 g, 1.6 mmol). The resulting mixture was stirred 2 hours - 5 before concentrating in vacuo and purifying the resulting by flash chromatography (SiO2, CHC13:MeOH
eluent) to give D-His-Tyr(OBn)-CONH(CH2)2Ph. The foam was ~issolved in CH2Cl2 (5 mL) and treated with 4-phenoxyphenyl isocyanate (0.05 g, 0.23 mmol). The resulting mixture was stirred 1 hour at room temperature, concentrated in vacuo, and purified the re~u:Lting oil by flash chromatography (SiO2, CHCl3:MeOH
eluent) to obtain (4-PhOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)2Ph as a foam; ES-MS 723 (m+1).
(S,R)-N-rl-(4-Benzyloxy-benzyl)-2-(4-benzyl-piperazin-1-yl)-2-oxo-ethyl]-3-(3H-imidazol-4-yl)-2-(toluene-4-sulfonylamino)-propionamide {(4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) HCl}
Step 1. Boc-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) According to Example 25, Step 1, by substituting l-benzylpiperizine for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid;
CI-MS 530 (m+l).
Step 2. Tyr(OBn)-CO(4-Bn-piperazin-1-yl) According to Example 25, Step 2, by substituting Boc-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) for Boc-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white solid; CI-MS 430 (m+l).
WO95/12612 PCT~S9~111553 Step 3. Fmoc-D-His(Tr)-Tyr(OBn)-CO(4-Bn-pi~erazin-l-yl) According to Example 25, Step 3, by substituting Tyr(OBn)-CO(4-Bn-piperazin-1-yl) ~or Tyr(OBn)-CONH(CH2)30Ph, the title compound was obt~;ne~ as a foam; ES-MS 1032 (m+1).
Step 4. (4-MePh)SO2-D-His(Tr)-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) Fmoc-D-His-Tyr(OBn)-CONH(CH2)30Ph, (0.7 g, 0.69 mmol) in CH2C12 (5 mL) was treated with piperidine (0.14 g, 1.6 mmol). The resulting mixture was stirred 2 hours be~ore concentrating in vacuo and purifying the resulting oil by flash chromatography (SiO2, CHC13:MeOH
eluent) to give D-His-Tyr(OBn)-CONH(CH2)2Ph. The ~oam was dissolved in CH2C12 (5 mL) and treated with pyridine (0.05 g, 0.63 mmol), followed by 4-toluenesulfonyl chloride (0.12 g, 0.63 mmol). The resulting mixture was stirred 3 hours at room temperature, concentrated in vacuo, and purified the resulting oil by flash chromatography (SiO2, CHC13:MeOH
eluent) to obtain (4-MePh)SO2-D-His(Tr)-Tyr(OBn)-CO(4-Bn-piperazin-1-yl); ES-MS 963 (m).
5 Step 5. (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) HCl (4-MePh)SO2-D-His(Tr)-Tyr(OBn)-CO(4-Bn-piperazin-l-yl) (0.21 g, 0.22 mmol) was treated with 80~ aqueous HCl (3 mL) and heated to 80 C for 5 minutes. The mixture was cooled and diluted with water (5 mL). The solid was ~iltered of~, and the supernate was concentrated in vacuo to provide an oil. The oil was dissolved in water (15 mL), frozen, and lyophilized to provide (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl) HCl; ES-MS 720 (m).
W095/12612 2 ~ 7 0 7 6 6 PCT~S94/11553 .
N- r (Phenylmethoxy)carbonyll-L-histidyl]-O-(~henylmethyl)-N-methyl-N-[2-(phenylmethoxY)ethyll-L-tyrosinamide {Cbz-His-Tyr(OBn)-CON(Me)CH2CH20Bn}
According to Example 19, Step 3, by substituting N-methyl-N-[2-(phenylmethoxy)ethyl]amine for 2-(phenylmethoxy)ethylamine, the title compound was prepared; FAB-MS 690 (m+1).
The present invention may be embodied in other spec:ific forms without departing from its ~pirit or essential characteristics. The described embodiments are to be considered in all respects only as illustrative and not restrictive. The scope of the invention is, therefore, indicated by the appended claims rather than by the foregoing description. All changes which come within the me~nlng and range of equivalency of the claims are to be embraced within their scope.
,
Step 1: Boc-Tyr(OBn)-Ser(OBn)-CO2Me To a solution of Boc-Tyr(OBn) (1.88 g, 6.50 mmol) in EtOAc ~30 mL) at 0C was added HOBT hydrate (1.19 g, 7.80 mmol) followed by DCC (1.61 g, 7.80 mmol). A
solution of Ser(OBn)-CO2Me TFA (2.1 g, 6.50 mmol) in EtOAc (20 mL) was added followed by Et3N (1.09 mL, 7.80 mmol). The mixture was allowed to warm to room temperature and stirred overnight. The mixture was filtered, diluted with EtOAc, and washed twice with saturated aqueous NaHC03, brine, dried over MgSO4, and concentrated. Flash chromatography (40~ EtOAc/hexane) gave 2.67 g (73~) of the title compound as a white solid, mp 81-84C.
Step 2: Tyr(OBn)-Ser(OBn)-CO2Me TFA
Boc-Tyr(OBn)-Ser~OBn)-C02Me (from Step 1 above, 2.64 g, 4.69 mmol) was dissolved in CH2C12 (15 mL), cooled to 0C and TFA (5 mL) was added. The ~olution was warmed to room temperature and stirred for 4 hours.
The solution was concentrated, taken up in CH2Cl2 and reconcentrated twice. The resulting oil was triturated with ether to provide 2.7 g of the title compound as a white solid.
Step 3: Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me To a solution of Cbz-His (1.00 g, 3.47 mmol) in DMF (15 mL) at 0C was added HOBT (0.64 g, 4.16 mmol) and DCC (0.86 g, 4.16 mmol). Tyr(OBn)-Ser(OBn)-CO2Me TFA (from Step 2 above, 2.0 g, 3.47 mmol) was added followed by Et3N (0.58 mL, 4.16 mmol). The mixture was allowed to warm to room temperature and stirred overnight. The mixture was filtered and the PCT~S94tllS53 %~ 40-filtrate was diluted with CHC13, washed twice with saturated aqueous NaHCO3, brine, dried over MgSO4, and concentrated. Flash chromatography (2-5~ MeOH/CHCl3) gave 2.14 g of the title compound as a white solid, mp 175-176C; FAB-MS 734 (m~1);
Anal. Calc. for C4lH43N58 C, 67.11; H, 5.91; N, 9.54;
Found: C, 66.96; H, 6.01; N, 9.41.
N-[N-rN-~(Phenylmethoxy)carbonyl]-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-serine {Cbz-His-Tyr(OBn)-Ser(OBn)}
To a solution of Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me (from Example 6 above, 2.02 g, 2.75 mmol) in THF
(50 m~) and MeOH (15 mL) at 0C was added 0.lN LiOH
(30.3 mL, 3.03 mmol). The solution was stirred for 6 hours at 0C, then concentrated. Water was added and the pH was adjusted to 4-5 with lN HCl. The mixture was filtered, and the solid was collected and dried to provide 1.55 g (78~) of the title compound as a white solid, mp 187-192C; ES-MS 720 (m+1);
Anal. Calc. for C40H4lN58 1 5H2O:
C, 64.33; H, 5.94; N, 9.38;
Found: C, 64.29; H, 5.73; N, 9.15.
EXAMP~E 8 N-rN-[N-[(Phenylmethoxy)carbonyl]-D-histidyll-O-(phenylmethyl)-L-tyrosyl]-O-(phenylmethyl)-L-serine, methyl ester ~Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me}
Step 1: Tyr(OBn)-Ser(OBn)-CO2Me HCl A solution of Boc-Tyr(OBn)-Ser(OBn)-CO2Me (from Bxample 6, Step 1 above, 9.90 g, 17.6 mmol) in EtOAc was cooled to 0C. Anhydrous HCl gas was bubbled through the cold solution for 5 minutes. The solution was allowed to warm to room temperature and stirred Wo 95/12612 ~ 21 70766 overnight. The solution was concentrated, taken up in EtOAc and reconcentrated to provide 8.75 g of the title compound as a foam; CI-MS 463 (m+1).
Step 2: Cbz-D-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me According to Example 6, Step 3, by substituting Cbz-D-His(Tr) for Cbz-His and Tyr(OBn)-Ser(OBn)-CO2Me HCl for Tyr(OBn)-Ser(OBn)-CO2Me TFA, the title compound was obtained as a white solid, mp 78-88C;
FAB-MS 976 (m+1).
Step 3: Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me A solution of Cbz-D-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me (from Step 2 above, 0.27 g, 0.28 mmol) in HOAc:H2O (4:1, 2 mL) was stirred at 80C for 5 minutes, then cooled to room temperature. The solution was partitioned between EtOAc and saturated aqueous NaHCO3.
The organic layer was washed with brine, dried (MgSO4), and concentrated. Flash chromatography (2-5~
MeOH:CHC13) yielded 0.10 g of the title compound as a foam; FAB-MS 734 (m+l).
N- rN- rN- r (Phenylmethoxy)carbonyll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-D-~erine, methyl ester {Cbz-His-Tyr(OBn)-D-Ser(OBn)-CO2Me}
According to Example 6, by substituting D-Ser(OBn)-CO2Me TFA for Ser(OBn)-CO2Me-TFA in Step 1, the title compound was obtained, mp 168-170C;
FAB-MS 734 (m+l).
PCT~S94/11553 z~3~ 42-N-r~-Methyl-N-rN- r (phenylmethoxy)carbonyll-L-hi tidyll-O-(phenylmethyl)-DL-tyrosyll-O-(phenylmethyl)-L-serine.
methyl ester {Cbz-His-DL-(~-Me)Tyr(OBn)-Ser(OBn)-CO
According to Example 6, by substituting Boc-DL-(~-Me)Tyr(OBn) for Boc-Tyr(OBn) in Step 1, the title compound was obtained; FAB-MS 748 (m+l).
N-Ethyl-N~-[N- rN- r (phenylmethoxy)carbonyl]-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-h-serinamide {Cbz-His-Tyr(OBn)-Ser(OBn)-CONHEt}
According to Example 6, by substituting Boc-Tyr(OBn)-Ser(OBn)-CONHEt for Boc-Tyr(OBn)-Ser(OBn)-CO2Me in Step 2, the title compound was obtained, mp 182-188C; FAB-MS 747 (m+l).
N-Ethyl-N~- rN- rN- r (Phenylmethoxy)carbonyll-D-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-serinamide {Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONHEt}
According to Example 6, by substituting Boc-Tyr(OBn)-Ser(OBn)-CONHEt for Boc-Tyr(OBn)-Ser(OBn)-CO2Me in Step 2 and Cbz-D-His for Cbz-His in Step 3, the title compound was obtained, mp 193-196C;
ES-MS 747 (m+l).
N-rN-rl-Methyl-N- r (phenylmethoxy)carbonylll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-O-(phenylmethyl)-L-Serine~
methyl ester {Cbz-His(l-Me)-Tyr(OBn)-Ser(OBn)-CO2Me}
Step 1: Cbz-His(l-Me) Benzyl chloroformate (0.24 mL, 1.7 mmol) was added dropwise to a slurry of l-methyl-~-histidine (0.25 g, WO9S/12612 P~T~S94111553 ~ 21 70766 1.5 mmol) in THF (5 mL) and saturated aqueous NaHCO3 (5 mL) at 0C. The mixture was allowed to warm to room temperature and stirred overnight. The mixture was concentrated and diluted with H2O, washed with ether, and the pH adjusted to 6-7 with lN HCl. The mixture was concentrated, then diluted with CHCl3 (150 mL) and MeOH (15 mL), and stirred for 1 hour. The mixture was dried (MgSO4) and concentrated to pro~ide 0.48 g of the title compound which was used without further purification.
Step 2: Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CO2Me To a slurry of Cbz-His(1-Me) (from Step 1 abo~e, 0.36 g, 1.2 mmol), Tyr(OBn)-Ser(OBn)-CO2Me HCl (from Example 8, Step 1 above, 0.60 g, 1.2 mmol), DCC
(0.30 g, 1.4 mmol), and HOBT (0.19 g, 1.4 mmol) in CH2Cl2 was added Et3N (0.17 mL, 1.2 mmol) and the mixture was stirred overnight at room temperature. The mixture was diluted with CHCl3, washed with saturated aqueous NaHCO3, brine, dried (MgSO4), and concentrated.
Flash chromatography (1~ MeOH:CHCl3) provided the title compound as a white solid, mp 161.5-163.5C; FA~3-MS 748 (m+1).
N-~M-rl-Methyl-N- r (phenylmethoxy)carbonyll-D-histidyll-O-(phenylmethyl)-L-tyrosyl]-O-(~henylmethyl)-L-serine.
methyl ester {Cbz-D-His(l-Me)-Tyr(OBn)-Ser(OBn)-CO2Me}
According to Example 13, by substituting 1-methyl-D-histidine for 1-methyl-L-histidine, the title compound was obtained; FAB-MS 748 (m+1).
WO9S/12612 PCT~S94/11553 .
N-[L-2-Amino-N - rN - r ( Phenylmethoxy)carbonyll-D-histidyll-4-r4-(phenylmethoxy)phenyllbutanoyll-0-(phenylmethyl-L-serine methyl ester {Cbz-D-His-homoTyr(OBn)-Ser(OBn)-CO2Me}
According to Example 6, by substituting Boc-homoTyr(OBn) for Boc-Tyr(OBn) in Step 1, and substituting Cbz-D-His for Cbz-His in Step 3, the title compound was obtained; ES-MS 748 (m+l).
N-r4-Phenyl-N-rN- r (phenylmethoxy)carbonyll-L-histidyll-L-phenylalanyll-O-(phenylmethyl)-L-serine methyl ester {Cbz-His-Phe(4-Ph)-Ser(OBn)-CO2Me}
According to Example 6, by substituting Boc-Phe(4-Ph) for Boc-Tyr(OBn) in Step 1, the title compound was obtained, mp 184-187C; FAB-MS 704 (m+l).
N- ro- (Phenylmethyl)-NrN- r r (Phenylmethyl)aminol-carbonyll-L-histidyll-L-tyrosyll-O-(phenylmethyl)-serine, methyl ester {BnNHCO-His-Tyr(OBn)-Ser(OBn)-C02Me }
Step 1: Fmoc-His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me According to Example 13, Step 2, by substituting Fmoc-His(Tr) for Cbz-His(l-Me), the title compound was obtained, mp 82-92C.
Step 2: His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me Piperidine (4.0 mL) was added to a slurry of Fmoc-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me (from Step 1 above, 1.85 g, 1.74 mmol) in CH2C12 (20 mL). The solution was stirred for 2 hours at room temperature, then concentrated. The residue was taken up in EtOAc (150 mL), washed with water (3 x 50 mL), dried (MgS04), and concentrated. The resulting oil was triturated W O 95/12612 2 PCTrUS94/11553 with Et2O/hexane. Flash chromatography of the residue (2~ MeOH/CHC13) gave 1.03 g o~ the title compound as a foam, mp 61.5-70C; ES-MS 843 (m+1).
Ste~ 3: BnNHCO-His(Tr)-Tyr(OBn~-Ser(OBn)-CO2Me Benzyl isocyanate (0.053 mL, 0.43 mmol) was added in one portion to a solution of His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me (~rom Step 2 above, 0.33 g, 0.39 mmol) in EtOAc (5 mL). The resulting slurry was stirred for 3 hours at room temperature, then concentrated to yield the title compound (0.4 g), which was used without fur~her purification.
Step 4: BnNHCO-His-Tyr(OBn)-Ser(OBn)-CO2Me According to Example 8, by substituting BnNHCO-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me for Cbz-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me, the title compound was obtained, mp 19 6.5- 199 C; ES-MS 733 (m+1).
N-rN-rN-(1-Oxo-3-phenylpropvl)-L-histidyll-O-(phenYlmethyl)-L-tyrosyl]-O-(phenylmethyl)-L-serine methyl ester {PhCH2CH2CO-His-Tyr(OBn)-Ser(OBn)-C02Me}
Step 1: PhCH2CH2CO-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me To a cooled (0C) solution of His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me (from Example 17, Step 2 above, 0.33 g, 0.39 mmol) in THF (5 mL) was added Et3N (O. 06 mL, 0.43 mmol) followed by phenylpropionyl chloride (0.064 mL, 0.43 mmol). The resulting slurry was brought to room temperature and stirred overnight. The mixture was partitioned between EtOAc and saturated aqueous NaHCO3. The organic layer was washed with brine, dried (MgSO4), and concentrated to yield the title compound as a solid which was u~ed without ~urther purification.
wossll26l2 pcT~s9~ ss3 .
~1 31 ~ -46-Step 2: PhCH2CH2CO-His-Tyr(OBn)-Ser(OBn)-CO2Me According to Example 8, Step 3, by substituting PhCH2CH2CO-His(Tr)-Tyr(OBn)-Ser(OBn)-CO2Me for Cbz-D-His(Tr)-Tyr(OBn)-Ser(OBn)-C02Me, the title compound was obtA;ne~, mp 193-196.5C; ES-MS 732 (m+l).
~-rN-~(Phenylmethoxy)carbonyll-L-histidyl]-O-(~henylmethyl)-N-r2-(phenylmethoxy)ethyll-~-tyrosinamide {Cbz-His-Tyr(OBn)-CONHCH2CH20Bn}
Step 1: Cbz-His-Tyr(OBn)-CO2Me According to Example 6, Step 3, by substituting Tyr(OBn)-CO2Me TFA for Tyr(OBn)-Ser(OBn)-CO2Me TFA, the title compound was obtained as a white powder, mp 145-148C; CI-MS 557 (m+l).
Step 2: Cbz-His-Tyr(OBn) According to Example 7, by substituting Cbz-His-Tyr(OBn)-CO2Me ~or Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me, the title compound was obtained as a white powder, mp 79-92C; CI-MS 543 (m+l).
Step 3: Cbz-His -Tyr(OBn)-CONHCH2CH20Bn To a solution of Cbz-His-Tyr(OBn) (from Step 2 above, 0.43 g, O.79 mmol) in DMF (4 mL) at 0C was added HOBT (0.15 g, 0.95 mmol) and DCC (0.20 g, 0.95 mmol). A solution of 2-(phenylmethoxy)ethylamine (0.12 g, 0.79 mmol) in DMF (1 mL) was then added. The mixture was allowed to warm to room temperature and stirred overnight. The mixture was filtered, diluted with CHCl3, washed twice with saturated aqueous NaHC03, washed with brine, dried over MgSO4, and concentrated.
Flash chromatography (3-5~ MeOH/CHC13) afforded 0.34 g (63~) of the title compound as a white solid, mp 136-150C; FAB-MS 676 (m+l);
WO 95/12612 2 1 7 0 l 6 6 PCTIUS94/11553 Anal. Calc. :Eor C39H41N506:
C, 69.32; H, 6.12; N, 10.36;
Found: C, 69.43; H, 6.24; N, 10.45.
~- rN- r (Phenylmethoxy) carbonyl]-D-histidyl]-N-r 2-(phenylmethoxy)ethyll-O-(phenylmethyl)-L-tyrosin~nide {cbz-D-His-Tyr(oBn)-cor~HcH2~H2c:)Bn}
According to Example 6, by substituting 2- (phenyl-methoxy)ethylamine for Ser(OBn) TFA and omitting Et3N
in Step 1 and by substituting Cbz-D-His for Cbz-His in Step 3, the title compound was prepared, mp 161-165C;
FAB-MS 676 (m+1).
~a~ rN-Methyl-Nr(phenylmethoxy)carbonyl]-D-histidyll-N-r2-~phenylmethoxy)ethyll-0-(phenylmethyl)-L-tyrosinamide {Cbz-D-His-(N-Me)Tyr(OBn)-CONHCH2CH20Bn}
According to Example 20, by substituting Boc-(N-Me)Tyr(OBn) for Boc-Tyr(OBn), the title compound was obtained, mp 64-78C; ES-MS 690 (m+l).
~a-rcY-Methyl-N-rN-~(~henylmethoxy)carbonyll-D-histidyll-N-r2-(phenylmethoxy)ethyll-O-(phenyl-methyl)-L-tyrosin~mide {Cbz-D-His-(cY-Me)Tyr(QBn)-CONHCH2CH2OBn}
According to Example 20, by substituting Boc-(cY-Me)Tyr(OBn) for Boc-Tyr(OBn), the title compound was obtained, mp 66-78C; ES-MS 690 (m+1).
W095/12612 PCT~S91111553 N-(2-Phenylethyl~-N~-[N- r (phenylmethoxy)carbonyll-T-histidyll-O-(phenylmethyl)-L-tyrosinamide {Cbz-His-Tyr(OBn)-CONHCH2CH2Ph}
According to Example 19, Step 3, by substituting 2-phenylethylamine for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid, mp 188-189.5C; FAB-MS 646 (m+1).
N-rN-rN-r(Phenylmethoxy)carbonyll-L-histidyll-O-(phenylmethyl)-L-tyrosyll-3-(3-pyridinyl)-L-alanine, methyl ester {Cbz-His-Tyr(OBn)-Pyr-CO2Me}
According to Example 19, Step 3, by substituting Pyr-CO2Me for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid, mp 180-182.5C
(dec); FAB-MS 705 (m+1).
(S.R)-N- r 2-(4-Benzyloxy-phenyl)-1-(3-phenoxy-propylcarbamoyl)-ethyll-2-r3-(4-ethoxy-phenyl)-ureidol-3-(3H-imidazol-4-yl)-propionamide {(4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)30Ph}
Step 1. Boc-Tyr(OBn)-CONH(CH2)30Ph 2-(Phenylmethoxy)ethylamine (0.81 g, 5.4 mmol) was added to a premixed solution of EDAC (1.2 g, 6.5 mmol), HOBT (0.87 g, 6.5 mmol), and Boc-Tyr(OBn)-OH (2.0 g, 5.4 mmol) in dry DMF (15 mL). The resulting mixture was stirred for 18 hours at room temperature. The solution was diluted with 1:1 EtOAc:Et20 (40 mL), washed with saturated aqueous NaCl (4 x 10 mL), dried (MgSO4), filtered, and concentrated in vacuo to provide a solid which was further purified by trituration with h~n~ to give the pure product, mp 145-146-C.
WO95/12612 PCT~S94/11553 21~0766 Step 2. Tyr(OLn)-CONH(CH2)3OPh Dry HCl gas was bubbled into an ice cold ~olution of Boc-Tyr(OBn)-CONH(CH2)3OPh (from Step l above, 2.0 g, 3.9 mmol) in MeOH (15 mL) for 4 minutes. The resulting mixture was stirred for l hour at 0 C and then allowed to warm to room temperature and stir l hour. The solution was concentrated in vacuo to provide a solid which was triturated with ether to provide Tyr(OBn)-CONH(CH2)3OPh HCl; CI-MS 405 (m+l).
The title compound was suspended in CHCl3, cooled in an ice bath, and NH3 gas was bubbled through the mixture for 2 minutes. The NH4Cl was filtered off, and the supernate was concentrated in vacuo to yield the free base of the title compound which was used in the next step without further purification.
Step 3. Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)3OPh To a solution of HOBT (0.48 g, 3.5 mmol) in DMF
(10 m~) was added Fmoc-D-His(Tr)-CO2H (2.0 g, 3.2 mmol) followed by EDAC (0.67 g, 3.5 mmol). The mixture was stirred at room temperature for 20 minutes before adding a solution of Tyr(OBn)-CONH(CH2)3OPh (from Step 2 above, 1.4 g, 3.2 mmol) in DMF (l0 mL). The mixture was stirred overnight at room temperature before partitioniny between a mixture of water (20 mL) and l:l Et2O:EtOAc (50 mL). The layers were separated, and the organic phase was washed with saturated aqueous NaCl (4x 20 mL) and dried (MgSO4). Filtration and concentration in vacuo provided an oil which was further purified by flash chromatography (SiO2, CHCl3:
MeOH eluent) to give the protected His-Tyr dipeptide;
FAB-MS 1006 (m).
Step 4. D-His(Tr)-Tyr(OBn)-CONH(CH2)3OPh Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph (from Step 3 above, 1.O g, 0.99 mmol) in CH2C12 (5 mL) was treated PCT~S94/1l553 Wo9SI12612 so- ' with piperidine (0.18 g, 2.1 mmol). The resulting mixture was stirred 2 hours before concentrating in vacuo and purifying the resulting oil by flash chromatography (SiO2, CHC13:MeOH eluent) to give (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph;
ES-MS 784 (m+l).
Step 5. (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2L30Ph D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph (from Step 4 above, 0.55 g, 0.7 mmol) in CH2Cl2 (5 mL) was treated with 4-ethoxyphenyl isocyanate (0.1 g, 0.7 mmol). The resulting mixture was stirred 1 hour at room temperature. Concentrated in vacuo. The resulting oil was purified by flash chromatography (SiO2, CHC13:MeOH
eluent) to give (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph; ES-MS 947 (m+1).
Step 6. (4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2L30Ph (4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph (from Step 5 above, 0.5 g, 0.52 mmol) in MeOH (5 mL) was treated with Pyridine HCl (catalytic). The resulting mixture was stirred at 65 C for 6 hours.
Concentrated in vacuo to obtain an oil which was purified by by flash chromatography (SiO2, CHCl3:MeOH
eluent) to give (4-EtO-Ph)NHCO-D-His-Tyr(OBn)-CONH(CH2)30Ph, mp 185-187 C; ES-MS 705 (m+1).
(S.R)-N-{2-(4-Benzyloxy-phenyl~-1-r3-(2-methoxy-phenyl)-propylcarbamoyll-ethyl}-3-(3H-imidazol-4-yl)-2-phenylacetylamino-propionamide {PhCH2CO-D-His-Tyr(OBn)- CONH(CH2L3(2-MeOPh)}
Step 1. Boc-Tyr(OBn)-CONH(CH2)3(2-MeOPh) According to Example 25, Step 1, by substituting 3-(2-methoxyphenyl) propyl amine for 2-(phenylmethoxy)-WO 95/12612 ~ 1 ~ O ~ ~ 6 ethylamine, the title compound was obtained as a white solid, mp 125-126.5-C.
.
Stel? 2. Tyr(OBn)-CONH(CH2)3(2-MeOPh) According to Example 25, Step 2, by substituting Boc-Tyr(OBn)-CONH(CH2)3(2-MeOPh) for Boc-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white solid; CI-MS 419 (m+1).
Step 3. Fmoc-D-His(Tr)-Tyr(OBn)-CQNH(CH2)3(2-MeOPh) According to Example 25, Step 3, by substituting Tyr~OBn)-CONH(CH2) 3 ( 2-MeOPh) for Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a foam; ES-MS 1020 (m).
Step 4. D-His(Tr)-Tyr(OBn)-CONH(CH2)3(2-MeOPh) According to Example 25, Step 4, by substituting Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)3(2-MeOPh) for Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obt~;ne~ as a white foam; ES-MS 798 (m+1).
Step 5. PhCH2CO-D-His(Tr)-Tyr(OBn)-CONH(CH2) 3 (2-MeOPh) To a solution of D-His(Tr)-Tyr(OBn)-CONH(CH2)3(2-MeOPh) (0.4 g, 0.5 mmol) in CH2Cl2 (5 mL) was added N-methyl morpholine (0.05 g, 0.5 mmol) followed by phenyl acetyl chloride (0.08 g, 0.5 mmol).
The resulting mixture was stirred 2 hours at room temperature. Diluted with DCM and washed with saturated aqueous NaHC03, saturated aqueous NaCl, and dried (MgSO4). Purified by flash chromatography (SiO2, CHCl3:MeOH eluent). The title compound was obtained as a foam; FAB-MS 916 (m).
Step 6. PhCH2CO-D-His-Tyr(OBn)-CONH(CH2) 3 (2-MeOPh) According to Example 25, Step 6, by substituting PhCH2CO-D-His(Tr)-Tyr(OBn)-CONH(CH2) 3 (2-MeOPh) for WO9S/12612 PCT~S94111S53 .
(4-EtOPh)NHCO-D-His(Tr)-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white foam; ES-MS 674 (m+1).
(S,R)-N-r2-(4-Benzyloxy-phenyl)-1-phenethylcarbamoyl-ethyl]-3-(3H-imidazol-4-yl)-2-r3-(4-phenoxy-phenyl)-ureidol-propionamide {(4-PhOPh)NHCO-D-His-Tyr(OBn)-CoNH(cH2L2ph}
Step 1. Boc-Tyr(OBn)CONH(CH2)2Ph According to Example 25, Step 1, by substituting phenethylamine for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid; CI-MS 475 (m+1).
Step 2. Tyr(OBn)-cONH(cH2L2Ph According to Example 25, Step 2, by substituting BocTyr(OBn)-CONH(CH2)2Ph for BocTyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white solid;
CI-MS 375 (m+1).
Step 3. Fmoc-D-His(Tr)-Tyr(OBn?-CONH(CH2L2Ph According to Example 25, Step 3, by substituting Tyr(OBn)-CONH(CH2)2Ph for Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a foam; ES-MS 977 (m+1).
Step 4. Fmoc-D-His-Tyr(OBn)-CONH(CH2L2Ph Fmoc-D-His(Tr)-Tyr(OBn)-CONH(CH2)2Ph (1.2 g, 1.6 mmol) in MeOH (5.mL) was treated with Pyridine HCl (catalytic). The resulting mixture was stirred at 65 C
overnight and concentrated in vacuo to obtain an oil which was purified by by flash chromatography (SiO2, CHC13:MeOH eluent) to give a white solid; ES-MS 734 (m+1).
PCT~S94111553 ~t~ 66 Step 5. (4-phoph)NHco-D-His-Tyr(oBn)-coNH(cH2L2ph Fmoc-D-His-Tyr(OBn)-CONH(CH2)2Ph (0.6 g, O.8 mmol) in CH2C12 (5 mL) was treated with piperdine (0.14 g, 1.6 mmol). The resulting mixture was stirred 2 hours - 5 before concentrating in vacuo and purifying the resulting by flash chromatography (SiO2, CHC13:MeOH
eluent) to give D-His-Tyr(OBn)-CONH(CH2)2Ph. The foam was ~issolved in CH2Cl2 (5 mL) and treated with 4-phenoxyphenyl isocyanate (0.05 g, 0.23 mmol). The resulting mixture was stirred 1 hour at room temperature, concentrated in vacuo, and purified the re~u:Lting oil by flash chromatography (SiO2, CHCl3:MeOH
eluent) to obtain (4-PhOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)2Ph as a foam; ES-MS 723 (m+1).
(S,R)-N-rl-(4-Benzyloxy-benzyl)-2-(4-benzyl-piperazin-1-yl)-2-oxo-ethyl]-3-(3H-imidazol-4-yl)-2-(toluene-4-sulfonylamino)-propionamide {(4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) HCl}
Step 1. Boc-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) According to Example 25, Step 1, by substituting l-benzylpiperizine for 2-(phenylmethoxy)ethylamine, the title compound was obtained as a white solid;
CI-MS 530 (m+l).
Step 2. Tyr(OBn)-CO(4-Bn-piperazin-1-yl) According to Example 25, Step 2, by substituting Boc-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) for Boc-Tyr(OBn)-CONH(CH2)30Ph, the title compound was obtained as a white solid; CI-MS 430 (m+l).
WO95/12612 PCT~S9~111553 Step 3. Fmoc-D-His(Tr)-Tyr(OBn)-CO(4-Bn-pi~erazin-l-yl) According to Example 25, Step 3, by substituting Tyr(OBn)-CO(4-Bn-piperazin-1-yl) ~or Tyr(OBn)-CONH(CH2)30Ph, the title compound was obt~;ne~ as a foam; ES-MS 1032 (m+1).
Step 4. (4-MePh)SO2-D-His(Tr)-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) Fmoc-D-His-Tyr(OBn)-CONH(CH2)30Ph, (0.7 g, 0.69 mmol) in CH2C12 (5 mL) was treated with piperidine (0.14 g, 1.6 mmol). The resulting mixture was stirred 2 hours be~ore concentrating in vacuo and purifying the resulting oil by flash chromatography (SiO2, CHC13:MeOH
eluent) to give D-His-Tyr(OBn)-CONH(CH2)2Ph. The ~oam was dissolved in CH2C12 (5 mL) and treated with pyridine (0.05 g, 0.63 mmol), followed by 4-toluenesulfonyl chloride (0.12 g, 0.63 mmol). The resulting mixture was stirred 3 hours at room temperature, concentrated in vacuo, and purified the resulting oil by flash chromatography (SiO2, CHC13:MeOH
eluent) to obtain (4-MePh)SO2-D-His(Tr)-Tyr(OBn)-CO(4-Bn-piperazin-1-yl); ES-MS 963 (m).
5 Step 5. (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl) HCl (4-MePh)SO2-D-His(Tr)-Tyr(OBn)-CO(4-Bn-piperazin-l-yl) (0.21 g, 0.22 mmol) was treated with 80~ aqueous HCl (3 mL) and heated to 80 C for 5 minutes. The mixture was cooled and diluted with water (5 mL). The solid was ~iltered of~, and the supernate was concentrated in vacuo to provide an oil. The oil was dissolved in water (15 mL), frozen, and lyophilized to provide (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-l-yl) HCl; ES-MS 720 (m).
W095/12612 2 ~ 7 0 7 6 6 PCT~S94/11553 .
N- r (Phenylmethoxy)carbonyll-L-histidyl]-O-(~henylmethyl)-N-methyl-N-[2-(phenylmethoxY)ethyll-L-tyrosinamide {Cbz-His-Tyr(OBn)-CON(Me)CH2CH20Bn}
According to Example 19, Step 3, by substituting N-methyl-N-[2-(phenylmethoxy)ethyl]amine for 2-(phenylmethoxy)ethylamine, the title compound was prepared; FAB-MS 690 (m+1).
The present invention may be embodied in other spec:ific forms without departing from its ~pirit or essential characteristics. The described embodiments are to be considered in all respects only as illustrative and not restrictive. The scope of the invention is, therefore, indicated by the appended claims rather than by the foregoing description. All changes which come within the me~nlng and range of equivalency of the claims are to be embraced within their scope.
,
Claims (25)
1. A compound of the Formula I:
I
wherein:
n = 1 or 2;
A = COR3, CO2R3, CONHR3, CSR3, C(S)OR3, C(S)NHR3, CF3SO2, aryl-SO2, or alkyl-SO2, wherein R3 is alkyl, (CH2)m-cycloalkyl, (CH2)m-aryl, (CH2)m-heteroaryl, or (CH2)mO-alkyl, and m =
0, 1, 2, or 3;
R = independently H or Me;
Y = independently H or Me;
Z = independently H or Me;
Rl = H, CO-aryl, (CH2)m-aryl, O(CH2)m-cycloalkyl, O(CH2)m-aryl, or O(CH2)m-heteroaryl wherein m is as defined above and R1 is located at either the meta or para position;
X = one to four substituents, including H, alkyl, CF3, F, Cl, Br, I, HO, MeO, NO2, NH2, N(Me)2, OPO3H2, or CH2PO3H2; and R2 = NR(CH2)nCO2R3, NR(CH2)nCONHR3, NR(CH2)nR3, NR(CH2)n+1OR4, NR(CH2)n+1SR4, NRCH(COR5)(CH2)n-heteroaryl, NRCH(COR5)(CH2)nOR3, NRCH(COR5)(CH2)nSR3, or wherein R, R3, and n are as defined above, R4 = H or R3, and R5 - OH, NH2, OR3, or NHR3; an optical isomer, diastereomer, or a pharmaceutically acceptable salt thereof.
I
wherein:
n = 1 or 2;
A = COR3, CO2R3, CONHR3, CSR3, C(S)OR3, C(S)NHR3, CF3SO2, aryl-SO2, or alkyl-SO2, wherein R3 is alkyl, (CH2)m-cycloalkyl, (CH2)m-aryl, (CH2)m-heteroaryl, or (CH2)mO-alkyl, and m =
0, 1, 2, or 3;
R = independently H or Me;
Y = independently H or Me;
Z = independently H or Me;
Rl = H, CO-aryl, (CH2)m-aryl, O(CH2)m-cycloalkyl, O(CH2)m-aryl, or O(CH2)m-heteroaryl wherein m is as defined above and R1 is located at either the meta or para position;
X = one to four substituents, including H, alkyl, CF3, F, Cl, Br, I, HO, MeO, NO2, NH2, N(Me)2, OPO3H2, or CH2PO3H2; and R2 = NR(CH2)nCO2R3, NR(CH2)nCONHR3, NR(CH2)nR3, NR(CH2)n+1OR4, NR(CH2)n+1SR4, NRCH(COR5)(CH2)n-heteroaryl, NRCH(COR5)(CH2)nOR3, NRCH(COR5)(CH2)nSR3, or wherein R, R3, and n are as defined above, R4 = H or R3, and R5 - OH, NH2, OR3, or NHR3; an optical isomer, diastereomer, or a pharmaceutically acceptable salt thereof.
2. A compound according to Claim 1 which is a compound of Formula II:
II
wherein:
A' = CO2R3, CONHR3, C(S)NHR3, or aryl-SO2, wherein R3 is alkyl, (CH2)m-cycloalkyl, (CH2)m-aryl, (CH2)m-heteroaryl, and m = 0, 1, 2 or 3;
R = independently H or Me;
Y = independently H or Me;
Z = independently H or Me;
R = (CH2)m-aryl, O(CH2)m-aryl, OPO3H2, or CH2PO3H2, wherein m is as defined above;
R2 = NR(CH2)2OR4, NR(CH2)2SR4, NRCH(COR5)CH2OR3, or NRCH(COR5)CH2SR3, wherein R3 and n are as defined above, R4 = H or R3, and R5 = OH, NH2, OR3, or NHR3; an optical isomer, diastereomer, or a pharmaceutically acceptable salt thereof.
II
wherein:
A' = CO2R3, CONHR3, C(S)NHR3, or aryl-SO2, wherein R3 is alkyl, (CH2)m-cycloalkyl, (CH2)m-aryl, (CH2)m-heteroaryl, and m = 0, 1, 2 or 3;
R = independently H or Me;
Y = independently H or Me;
Z = independently H or Me;
R = (CH2)m-aryl, O(CH2)m-aryl, OPO3H2, or CH2PO3H2, wherein m is as defined above;
R2 = NR(CH2)2OR4, NR(CH2)2SR4, NRCH(COR5)CH2OR3, or NRCH(COR5)CH2SR3, wherein R3 and n are as defined above, R4 = H or R3, and R5 = OH, NH2, OR3, or NHR3; an optical isomer, diastereomer, or a pharmaceutically acceptable salt thereof.
3. A compound according to Claim 1 wherein A is CO2R3 or CONHR3.
4. A compound according to Claim 1 wherein at least one of Y and Z is Me.
5. A compound according to Claim 1 wherein R2 is NH(CH2)2OR4 or NHCH(COR5)CH2OR3.
6. A compound according to Claim 1 wherein A is NHCONHR3, R2 is (CH2)2OR4, and at least one of Y
and Z is Me.
and Z is Me.
7. A compound according to Claim 1 selected from the group consisting of:
Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONHEt; and Cbz-His-Tyr(OBn)-Ser(OBn).
Cbz-His-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-Tyr(OBn)-Ser(OBn)-CONHEt; and Cbz-His-Tyr(OBn)-Ser(OBn).
8. A compound according to Claim 1 selected from the group consisting of:
Cbz-His-Tyr(OBn)-D-Ser(OBn)-CO2Me;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me; and Cbz-D-His-Tyr(OBn)-Ser(OBn).
Cbz-His-Tyr(OBn)-D-Ser(OBn)-CO2Me;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-D-His-Tyr(OBn)-Ser(OBn)-CO2Me; and Cbz-D-His-Tyr(OBn)-Ser(OBn).
9. A compound according to claim 1 selected from the group consisting of:
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn);
Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONHEt; and Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn).
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His(1-Me)-Tyr(OBn)-Ser(OBn);
Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn)-CONHEt; and Cbz-D-His(1-Me)-Tyr(OBn)-Ser(OBn).
10. A compound according to claim 1 selected from the group consisting of:
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn);
Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONHEt; and Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn).
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONHEt;
Cbz-His-(.alpha.-Me)Tyr(OBn)-Ser(OBn);
Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONH2;
Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn)-CONHEt; and Cbz-His-D-(.alpha.-Me)Tyr(OBn)-Ser(OBn).
11. A compound according to Claim 1 selected from the group consisting of:
Cbz-D-His-homoTyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-Phe(4-Ph)-Ser(OBn)-CO2Me;
Cbz-D-His-Phe(4-Ph)-Ser(OBn)-CO2Me;
Cbz-His-Tyr(OBn)-Pyr-CO2Me; and Cbz-D-His-Tyr(OBn)-Pyr-CO2Me.
Cbz-D-His-homoTyr(OBn)-Ser(OBn)-CO2Me;
Cbz-His-Phe(4-Ph)-Ser(OBn)-CO2Me;
Cbz-D-His-Phe(4-Ph)-Ser(OBn)-CO2Me;
Cbz-His-Tyr(OBn)-Pyr-CO2Me; and Cbz-D-His-Tyr(OBn)-Pyr-CO2Me.
12. A compound according to Claim 1 selected from the group consisting of:
Cbz-His-Tyr(OBn)-CONHCH2CH2OBn;
Cbz-D-His-Tyr(OBn)-CONHCH2CH2OBn;
Cbz-His-(N-Me)Tyr(OBn)-CONHCH2CH2OBn;
Cbz-D-His-(N-Me)Tyr(OBn)-CONHCH2CH2OBn;
Cbz-His-Tyr(OBn)-CONH(CH2)2Ph; and Cbz-D-His-Tyr(OBn)-CONH(CH2)2Ph.
Cbz-His-Tyr(OBn)-CONHCH2CH2OBn;
Cbz-D-His-Tyr(OBn)-CONHCH2CH2OBn;
Cbz-His-(N-Me)Tyr(OBn)-CONHCH2CH2OBn;
Cbz-D-His-(N-Me)Tyr(OBn)-CONHCH2CH2OBn;
Cbz-His-Tyr(OBn)-CONH(CH2)2Ph; and Cbz-D-His-Tyr(OBn)-CONH(CH2)2Ph.
13. A compound according to Claim 1 selected from the group consisting of:
Cbz-His-Tyr(OBn)-Gly-CO2Bn;
Cbz-D-His-Tyr(OBn)-Gly-CO2Bn;
Cbz-His-Tyr(OBn)-Gly-CONHBn; and Cbz-D-His-Tyr(OBn)-Gly-CONHBn.
Cbz-His-Tyr(OBn)-Gly-CO2Bn;
Cbz-D-His-Tyr(OBn)-Gly-CO2Bn;
Cbz-His-Tyr(OBn)-Gly-CONHBn; and Cbz-D-His-Tyr(OBn)-Gly-CONHBn.
14. A compound according to Claim 1 selected from the group consisting of:
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CO2Me;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-His-Tyr(OBn)-Ser(OBn);
BnNHCO-His-Tyr(OBn)-CONHCH2CH2OBn; and BnNHCO-His-Tyr(OBn)-CONHCH2CH2CH2OPh.
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CO2Me;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-His-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-His-Tyr(OBn)-Ser(OBn);
BnNHCO-His-Tyr(OBn)-CONHCH2CH2OBn; and BnNHCO-His-Tyr(OBn)-CONHCH2CH2CH2OPh.
15. A compound according to Claim 1 selected from the group consisting of:
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CO2Me;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn);
BnNHCO-D-His-Tyr(OBn)-CONHCH2CH2OBn; and BnNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH2OPh.
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CO2Me;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONH2;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn)-CONHEt;
BnNHCO-D-His-Tyr(OBn)-Ser(OBn);
BnNHCO-D-His-Tyr(OBn)-CONHCH2CH2OBn; and BnNHCO-D-His-Tyr(OBn)-CONHCH2CH2CH2OPh.
16. A compound according to Claim 1 selected from the group consisting of:
Cbz-His-Tyr(OBn)-CON(Me)CH2CH2OBn;
(4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)3OPh;
PhCH2CO-D-His-Tyr(OBn)-CONH(CH2)3-(2-MeOPh);
(4-PhOPh)NHCO-D-His-Tyr(OBn)-COHN(CH2)2Ph;
and (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl).
Cbz-His-Tyr(OBn)-CON(Me)CH2CH2OBn;
(4-EtOPh)NHCO-D-His-Tyr(OBn)-CONH(CH2)3OPh;
PhCH2CO-D-His-Tyr(OBn)-CONH(CH2)3-(2-MeOPh);
(4-PhOPh)NHCO-D-His-Tyr(OBn)-COHN(CH2)2Ph;
and (4-MePh)SO2-D-His-Tyr(OBn)-CO(4-Bn-piperazin-1-yl).
17. A method of treating tissue proliferative diseases comprising administering to a host suffering therefrom a therapeutically effective amount of a compound according to Claim 1 in unit dosage form.
18. A pharmaceutical composition adapted for administration as an antiproliferative agent comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a pharmaceutically acceptable excipient, diluent or carrier.
19. A method of treating cancer comprising administering to a host suffering therefrom a therapeutically effective amount of a compound according to Claim 1 in unit dosage form.
20. A pharmaceutical composition adapted for administration as an anticancer agent comprising a therapeutically effective amount of a compound according to Claim 1 in admixture with a pharmaceutically acceptable excipient, diluent or carrier.
21. A method of treating restenosis comprising administering to a host suffering therefrom a therapeutically effective amount of a compound according to Claim 1 in unit dosage form.
22. A pharmaceutical composition adapted for administration as a restenosis inhibiting agent comprising a therapeutically effective amount of a compound according to Claim 1 in A admixture with a pharmaceutically acceptable excipient, diluent, or carrier.
23. A process for the preparation of organic compounds according to Claim 1, or a pharmaceutically acceptable salt thereof, comprising the steps of employing solid phase support technology and sequentially coupling building blocks utilizing a solid phase peptide synthesizer, cleaving the organic compound from the solid phase support and subsequently optionally modifying the C-terminus of the organic compound in solution phase to afford a compound or a pharmaceutically acceptable salt thereof of Formula I.
24. A process for the preparation of compounds according to Claim 1, or a pharmaceutically acceptable salt thereof, comprising the steps of employing solution phase technology and sequentially coupling building blocks to afford a compound or a pharmaceutically acceptable salt thereof of Formula I.
25. A process for the preparation of compounds according to Claim 1, or a pharmaceutically acceptable salt thereof, comprising simultaneous synthesis of compounds of Formula I in a multiple simultaneous synthesis apparatus, using a D- or L-histidine containing dipeptide derivative that is supported on 2-chlorotrityl resin with sequential deprotection and acylation of the N-terminus followed by sequential deprotection of the carboxy terminus, carboxyl activation and condensation with a series of amines, followed by cleavage from the solid support.
Applications Claiming Priority (4)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US14873593A | 1993-11-05 | 1993-11-05 | |
| US148,735 | 1993-11-05 | ||
| US30330194A | 1994-09-13 | 1994-09-13 | |
| US303,301 | 1994-09-13 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA2170766A1 true CA2170766A1 (en) | 1995-05-11 |
Family
ID=26846110
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002170766A Abandoned CA2170766A1 (en) | 1993-11-05 | 1994-10-12 | Substituted di- and tripeptide inhibitors of protein:farnesyl transferase |
Country Status (10)
| Country | Link |
|---|---|
| EP (1) | EP0730605A1 (en) |
| JP (1) | JP3597863B2 (en) |
| KR (1) | KR100362338B1 (en) |
| AU (1) | AU681454B2 (en) |
| CA (1) | CA2170766A1 (en) |
| FI (1) | FI961819A0 (en) |
| HU (1) | HUT75308A (en) |
| NO (1) | NO961814L (en) |
| NZ (1) | NZ275691A (en) |
| WO (1) | WO1995012612A1 (en) |
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-
1994
- 1994-10-12 AU AU79760/94A patent/AU681454B2/en not_active Ceased
- 1994-10-12 JP JP51322495A patent/JP3597863B2/en not_active Expired - Fee Related
- 1994-10-12 EP EP94930725A patent/EP0730605A1/en not_active Withdrawn
- 1994-10-12 NZ NZ275691A patent/NZ275691A/en unknown
- 1994-10-12 WO PCT/US1994/011553 patent/WO1995012612A1/en not_active Application Discontinuation
- 1994-10-12 KR KR1019960702309A patent/KR100362338B1/en not_active IP Right Cessation
- 1994-10-12 HU HU9601193A patent/HUT75308A/en unknown
- 1994-10-12 CA CA002170766A patent/CA2170766A1/en not_active Abandoned
-
1996
- 1996-04-29 FI FI961819A patent/FI961819A0/en unknown
- 1996-05-03 NO NO961814A patent/NO961814L/en not_active Application Discontinuation
Also Published As
| Publication number | Publication date |
|---|---|
| FI961819A (en) | 1996-04-29 |
| JPH09504547A (en) | 1997-05-06 |
| NO961814D0 (en) | 1996-05-03 |
| FI961819A0 (en) | 1996-04-29 |
| HUT75308A (en) | 1997-05-28 |
| HU9601193D0 (en) | 1996-07-29 |
| AU7976094A (en) | 1995-05-23 |
| EP0730605A1 (en) | 1996-09-11 |
| JP3597863B2 (en) | 2004-12-08 |
| KR100362338B1 (en) | 2003-04-08 |
| WO1995012612A1 (en) | 1995-05-11 |
| AU681454B2 (en) | 1997-08-28 |
| NZ275691A (en) | 1998-03-25 |
| NO961814L (en) | 1996-05-06 |
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