U.S. patent application number 11/210717 was filed with the patent office on 2006-03-02 for antipicornaviral compounds and compositions, their pharmaceutical uses, and materials for their synthesis.
This patent application is currently assigned to Agouron Pharmaceuticals, Inc.. Invention is credited to Peter Scott Dragovich, Theodore O. JR. Johnson, Joseph Timothy Marakovits, Thomas Jay Prins, Stephen Evan Webber, Ru Zhou.
Application Number | 20060046966 11/210717 |
Document ID | / |
Family ID | 26769788 |
Filed Date | 2006-03-02 |
United States Patent
Application |
20060046966 |
Kind Code |
A1 |
Dragovich; Peter Scott ; et
al. |
March 2, 2006 |
Antipicornaviral compounds and compositions, their pharmaceutical
uses, and materials for their synthesis
Abstract
Peptido and peptidomimetic compounds of the formula: ##STR1##
wherein the formula variables are as defined in the disclosure,
advantageously inhibit or block the biological activity of the
picornaviral 3C protease. These compounds, as well as
pharmaceutical compositions containing these compounds, are useful
for treating patients or hosts infected with one or more
picornaviruses, such as RVP. Intermediates and synthetic methods
for preparing such compounds are also provided.
Inventors: |
Dragovich; Peter Scott;
(Encinitas, CA) ; Webber; Stephen Evan; (San
Diego, CA) ; Prins; Thomas Jay; (Cardiff, CA)
; Zhou; Ru; (Carlsbad, CA) ; Marakovits; Joseph
Timothy; (Encinitas, CA) ; Johnson; Theodore O.
JR.; (San Diego, CA) |
Correspondence
Address: |
AGOURON PHARMACEUTICALS, INC.
10777 SCIENCE CENTER DRIVE
SAN DIEGO
CA
92121
US
|
Assignee: |
Agouron Pharmaceuticals,
Inc.
|
Family ID: |
26769788 |
Appl. No.: |
11/210717 |
Filed: |
August 24, 2005 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10289982 |
Nov 6, 2002 |
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11210717 |
Aug 24, 2005 |
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09647712 |
Nov 28, 2001 |
6906198 |
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10289982 |
Nov 6, 2002 |
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09301977 |
Apr 29, 1999 |
6531452 |
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09647712 |
Nov 28, 2001 |
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60098358 |
Aug 28, 1998 |
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60083828 |
Apr 30, 1998 |
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Current U.S.
Class: |
548/128 ;
514/20.1; 514/3.7; 514/362; 514/364; 548/132 |
Current CPC
Class: |
A61P 31/16 20180101;
C07D 261/18 20130101; C07D 207/26 20130101; C07K 5/0205 20130101;
A61P 31/12 20180101; C07D 413/12 20130101; Y02A 50/30 20180101;
A61K 38/00 20130101; Y02A 50/465 20180101; Y02A 50/463
20180101 |
Class at
Publication: |
514/019 ;
514/362; 514/364; 548/128; 548/132 |
International
Class: |
A61K 38/04 20060101
A61K038/04; C07K 5/04 20060101 C07K005/04; A61K 31/433 20060101
A61K031/433; A61K 31/4245 20060101 A61K031/4245 |
Claims
1. A compound of the formula I: ##STR136## wherein: Y is
--N(R.sub.y)--, --C(R.sub.y)(R.sub.y)--, or --O--, where each
R.sub.y is independently H or lower alkyl; R.sub.1 is H, F, an
alkyl group, OH, SH, or an O-alkyl group; R.sub.2 and R.sub.3 are
each independently H; ##STR137## where n is an integer from 0 to 5,
A.sub.1 is CH or N, A.sub.2 and each A.sub.3 are independently
selected from C(R.sub.41)(R.sub.41), N(R.sub.41), S, S(O),
S(O).sub.2, and 0, and A.sub.4 is NH or NR.sub.41, where each
R.sub.41 is independently H or lower alkyl, provided that no more
than 2 heteroatoms occur consecutively in the ring formed by
A.sub.1, A.sub.2, (A.sub.3).sub.n, A.sub.4 and C.dbd.O; and
provided that at least one of R.sub.2 and R.sub.3 is ##STR138##
R.sub.5 and R.sub.6 are each independently H, F, an alkyl group, a
cycloalkyl group, a heterocycloalkyl group, an aryl group, or a
heteroaryl group; R.sub.7 and R.sub.8 are each independently H, an
alkyl group, a cycloalkyl group, a heterocycloalkyl group, an aryl
group, a heteroaryl group, --OR.sub.17, --SR.sub.17,
--NR.sub.17R.sub.18, NR.sub.19NR.sub.17R.sub.18, or
--NR.sub.17OR.sub.18, where R.sub.17, R.sub.18, and R.sub.19 are
each independently H, an alkyl group, a cycloalkyl group, a
heterocycloalkyl group, an aryl group, a heteroaryl group, or an
acyl group; R.sub.9 is a five-membered heterocycle having from one
to three heteroatoms selected from O, N, and S, or R.sub.9 is
##STR139## where R.sub.2 is ##STR140## and Z and Z.sub.1 are each
independently H, F, an alkyl group, a cycloalkyl group, a
heterocycloalkyl group, an aryl group, a heteroaryl group,
--C(O)R.sub.21, --CO.sub.2R.sub.21, --CN, --C(O)NR.sub.21,R.sub.22,
--C(O)NR.sub.21OR.sub.22, --C(S)R.sub.21, --C(S)NR.sub.21R.sub.22,
--NO.sub.2, --SOR.sub.21, --SO.sub.2R.sub.21,
--SO.sub.2NR.sub.21R.sub.22, --SONR.sub.21)(OR.sub.22),
--SONR.sub.21, --SO.sub.3R.sub.21, --PO(OR.sub.21).sub.2,
--PO(R.sub.21)(R.sub.22), --PO(NR.sub.21R.sub.22)(OR.sub.23),
PO(NR.sub.21R.sub.22)(NR.sub.23R.sub.24),
--C(O)NR.sub.21NR.sub.22R.sub.23, or
--C(S)NR.sub.21NR.sub.22R.sub.23, where R.sub.21, R.sub.22,
R.sub.23, and R.sub.24 are each independently H, an alkyl group, a
cycloalkyl group, a heterocycloalkyl group, an aryl group, a
heteroaryl group, an acyl group, or a thioacyl group, or any two of
R.sub.21, R.sub.22, R.sub.23, and R.sub.24, together with the
atom(s) to which they are bonded, form a heterocycloalkyl group,
provided that Z and Z, are not both H; or Z.sub.1 and R.sub.1,
together with the atoms to which they are bonded, form a cycloalkyl
or heterocycloalkyl group; or Z and Z.sub.1, together with the
atoms to which they are bonded, form a cycloalkyl or
heterocycloalkyl group; or a prodrug, pharmaceutically active
metabolite, pharmaceutically acceptable salt, or solvate thereof.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is related to U.S. Provisional Application
No. 60/098,358, filed Aug. 28, 1998, and U.S. Provisional
Application No. 60/083,828, filed Apr. 30, 1998.
FIELD AND INDUSTRIAL APPLICABILITY OF THE INVENTION
[0002] The invention pertains to peptide-like and peptidomimetic
compounds that advantageously inhibit the enzymatic activity of
picornaviral 3C proteases, especially rhinovirus 3C proteases
(RVPs), and that retard viral growth in cell culture. The invention
also relates to the use of such compounds in pharmaceutical
compositions and therapeutic treatments for rhinoviral infections.
The invention further relates to processes for synthesizing such
compounds and compounds useful in such syntheses.
BACKGROUND OF THE INVENTION
[0003] The picornaviruses are a family of tiny non-enveloped
positive-stranded RNA-containing viruses that infect humans and
other animals. These viruses include the human rhinoviruses, human
polioviruses, human coxsackieviruses, human echoviruses, human and
bovine enteroviruses, encephalomyocarditis viruses, meningitis
virus, foot and mouth viruses, hepatitis A virus, and others. The
human rhinoviruses are a major cause of the common cold. To date,
there are no effective therapies on the market that cure the common
cold, only treatments that relieve the symptoms.
[0004] Picornaviral infections may be treated by inhibiting the
proteolytic 3C enzymes. These enzymes are required for the natural
maturation of the picornaviruses. They are responsible for the
autocatalytic cleavage of the genomic, large polyprotein into the
essential viral proteins. Members of the 3C protease family are
cysteine proteases, where the sulfhydryl group most often cleaves
the glutamine-glycine amide bond. Inhibition of 3C proteases is
believed to block proteolytic cleavage of the polyprotein, which in
turn can retard the maturation and replication of the viruses by
interfering with viral particle production. Therefore, inhibiting
the processing of this cysteine protease with selective small
molecules that are specifically recognized should represent an
important and useful approach to treat and cure viral infections of
this nature and, in particular, the common cold.
[0005] Some small-molecule inhibitors of the enzymatic activity of
picornaviral 3C proteases (i.e., antipicornaviral compounds) have
been recently discovered. See, for example: U.S. patent application
Ser. No. 08/850,398, filed May 2, 1997, by Webber et al.; U.S.
patent application Ser. No. 08/991,282, filed Dec. 16, 1997, by
Dragovich et al.; and U.S. patent application Ser. No. 08/991,739,
filed Dec. 16, 1997, by Webber et al. These U.S. patent
applications, the disclosures of which are incorporated herein by
reference, describe certain antipicornaviral compounds. There is
still a desire to discover small-molecule compounds that are
especially potent antipicornaviral agents.
SUMMARY OF THE INVENTION
[0006] Thus, an object of this invention is to discover
small-molecule compounds that are especially potent
antipicornaviral agents. A further object of the invention is to
provide intermediates useful for the synthesis of said
protease-inhibiting compounds and synthetic methods useful for such
syntheses. A yet further object of the invention is to achieve
pharmaceutical compositions that are highly effective for treating
maladies mediated by inhibition of picornaviral 3C proteases, such
as the common cold.
[0007] Such objects have been attained through the discovery of
compounds of the invention, which are picornaviral 3C protease
inhibitors displaying particularly strong antiviral activity. It
has surprisingly been discovered that peptido and peptidomimetic
compounds containing a five-membered heterocyclic group have high
rhinoviral-protease-inhibiting activity. It has further been
surprisingly found that the rhinoviral-protease-inhibiting activity
of peptido and peptidomimetic compounds may be significantly
enhanced by replacing a glutamine-like moiety found in some known
rhinoviral-protease-inhibiting compounds with a side-chain
comprising a gamma- or delta-lactam.
[0008] The inhibitors of the present invention are of the following
general formula (1): ##STR2## wherein: [0009] Y is --N(R.sub.y)--,
--C(R.sub.y)(R.sub.y)--, or --O--, where each R.sub.y is
independently --H or lower alkyl; [0010] R.sub.1 is --H, --F,
-alkyl, --OH, --SH, or an O-alkyl group; [0011] R.sub.2 and R.sub.3
are each independently H; ##STR3## where n is an integer from 0 to
5, A.sub.1 is CH or N, A.sub.2 and each A.sub.3 are independently
selected from C(R.sub.41)(R.sub.41), N(R.sub.41), S, S(O),
S(O).sub.2, and 0, and A.sub.4 is NH or NR.sub.41, where each
R.sub.4, is independently H or lower alkyl, provided that no more
than two heteroatoms occur consecutively in the above-depicted ring
formed by A.sub.1, A.sub.2, (A.sub.3).sub.n, A.sub.4 and C.dbd.O,
and at least one of R.sub.2 and R.sub.3 is ##STR4## [0012] R.sub.5
and R.sub.6 are each independently H, F, an alkyl group, a
cycloalkyl group, a heterocycloalkyl group, an aryl group, or a
heteroaryl group; [0013] R.sub.7 and R.sub.8 are each independently
H, an alkyl group, a cycloalkyl group, a heterocycloalkyl group, an
aryl group, a heteroaryl group, --OR.sub.17, --SR.sub.17,
--NR.sub.17R.sub.18, --NR.sub.19NR.sub.17R.sub.18, or
--NR.sub.17OR.sub.18, where R.sub.17, R.sub.18, and R.sub.19 are
each independently H, an alkyl group, a cycloalkyl group, a
heterocycloalkyl group, an aryl group, a heteroaryl group, or an
acyl group, provided that at least one of R.sub.7 and R.sub.8 is an
alkyl group, a cycloalkyl group, a heterocycloalkyl group, an aryl
group, a heteroaryl group, --OR.sub.17, --SR.sub.17,
--NR.sub.17R.sub.18, --NR.sub.19NR.sub.17R.sub.18, or
--NR.sub.17OR.sub.18; [0014] R.sub.9 is a suitable organic moiety;
and [0015] Z and Z.sub.1 are each independently H, F, an alkyl
group, a cycloalkyl group, a heterocycloalkyl group, an aryl group,
a heteroaryl group, --C(O)R.sub.21, --CO.sub.2R.sub.21, --CN,
--C(O)NR.sub.21R.sub.22, --C(O)NR.sub.21OR.sub.22, --C(S)R.sub.21,
--C(S)NR.sub.21R.sub.22, --NO.sub.2, --SOR.sub.21,
--SO.sub.2R.sub.21, --SO.sub.2NR.sub.21R.sub.22,
--SO(NR.sub.21)(OR.sub.22), --SONR.sub.21, --SO.sub.3R.sub.21,
--PO(OR.sub.21).sub.2, --PO(R.sub.21)(R.sub.22),
--PO(NR.sub.21R.sub.22)(OR.sub.23),
--PO(NR.sub.21R.sub.22)(NR.sub.23R.sub.24),
--C(O)NR.sub.21NR.sub.22R.sub.23, or
--C(S)NR.sub.21NR.sub.22R.sub.23, where R.sub.21, R.sub.22,
R.sub.23, and R.sub.24 are each independently H, an alkyl group, a
cycloalkyl group, a heterocycloalkyl group, an aryl group, a
heteroaryl group, an acyl group, or a thioacyl group, or where any
two of R.sub.21, R.sub.22, R.sub.23, and R.sub.24, together with
the atom(s) to which they are bonded, form a heterocycloalkyl
group, provided that Z and Z.sub.1 are not both H; [0016] or
Z.sub.1 and R.sub.1, together with the atoms to which they are
bonded, form a cycloalkyl or heterocycloalkyl group, where Z.sub.1
and R.sub.1 are as defined above except for moieties that cannot
form the cycloalkyl or heterocycloalkyl group; [0017] or Z and
Z.sub.1, together with the atoms to which they are bonded, form a
cycloalkyl or heterocycloalkyl group, where Z and Z.sub.1 are as
defined above except for moieties that cannot form the cycloalkyl
or heterocycloalkyl group. The invention also pertains to prodrugs,
pharmaceutically acceptable salts, pharmaceutically active
metabolites, and pharmaceutically acceptable solvates of compounds
of the formula I.
[0018] In preferred embodiments of the compounds of the formula I,
R.sub.2 and R.sub.3 are each independently H; ##STR5## where n is
an integer from 0 to 5, each R.sub.41 is independently H or lower
alkyl, and the stereochemistry at the carbon denoted with an
asterisk may be R or S; provided that at least one of R.sub.2 and
R.sub.3 is ##STR6## Preferably, R.sub.9 is a five-membered
heterocycle having one to three heteroatoms selected from O, N, and
S. Alternatively, R.sub.9 is ##STR7##
[0019] In other preferred embodiments, the variables of formula I
are as follows. Z and Z.sub.1 are each independently selected from
H, F, lower alkyl, --CO.sub.2R.sub.21, and --C(O)NR.sub.21R.sub.22,
provided that Z and Z.sub.1 are not both H, where R.sub.21 and
R.sub.22 are each independently H, an alkyl group, a cycloalkyl
group, a heterocycloalkyl group, an aryl group, a heteroaryl group,
an acyl group, or a thioacyl group, or R.sub.2, and R.sub.22,
together with the atom(s) to which they are bonded, form a
heterocycloalkyl group. At least one of R.sub.2 or R.sub.3 is
##STR8## and the other is H. R.sub.5 and R.sub.6 are each
independently selected from H, F, an alkyl group, a cycloalkyl
group, a heterocycloalkyl group, an aryl group, and a heteroaryl
group, more preferably one of R.sub.5 and R.sub.6 is H and the
other is alkyl or aryl (e.g., unsubstituted or substituted
phenylmethyl). R.sub.7 and R.sub.8 are each independently H, an
alkyl group, a cycloalkyl group, a heterocycloalkyl group, an aryl
group, or a heteroaryl group; and more preferably one of R.sub.7
and R.sub.8 is H and the other is alkyl (e.g., 2-propyl,
2-methyl-2-propyl, or 2-methyl-1-propyl) or arylmethyl (e.g.,
unsubstituted or substituted phenylmethyl or naphthylmethyl).
R.sub.9 is a five-membered heterocycle having from one to three
heteroatoms selected from O, N, and S, more preferably a
five-membered heterocycle having at least one nitrogen heteroatom
and at least one oxygen heteroatom (e.g., unsubstituted or
substituted 1,2-oxazolyl (i.e., isoxazolyl), 1,3-oxazolyl (i.e.,
oxazolyl), or oxadiazolyl (1,2,3-oxadiazolyl, 1,2,4-oxadiazolyl, or
1,2,5-oxadiazolyl). When R.sub.9 is oxadiazolyl, unsubstituted and
monomethyl-substituted 1,2,4-oxadiazolyl are preferred. In
especially preferred embodiments, R.sub.9 is 3-isoxazolyl or
5-isoxazolyl, either unsubstituted or substituted with one or two
methyl groups and/or halogens, with chlorine and fluorine being
preferred halogen substituents.
[0020] In a preferred embodiment, the compounds, prodrugs,
pharmaceutically acceptable salts, pharmaceutically active
metabolites, and solvates have an antipicornaviral activity with an
EC.sub.50 less than or equal to 100 .mu.M in the H1-HeLa cell
culture assay, and more preferably an antirhinoviral activity with
an EC.sub.50 less than or equal to 10 .mu.M in the H1-HeLa cell
culture.
[0021] In another aspect, the invention is directed to
intermediates of formula II, preferably of the formula II', which
are useful in the synthesis of certain compounds: ##STR9## wherein:
[0022] p is an integer of from 0 to 5; [0023] A.sub.11 is CH or N,
A.sub.12 and each A.sub.13 are independently selected from
C(R.sub.61)(R.sub.61), N(R.sub.61), S, S(O), S(O).sub.2, and O, and
A.sub.14 is NH or NR.sub.61, where each R.sub.61 is independently
H, alkyl, acyl, or aryl, provided that no more than two heteroatoms
occur consecutively in the above-depicted ring in formula II formed
by A.sub.11, A.sub.12, (A.sub.13).sub.n, A.sub.14 and C.dbd.O;
[0024] each R.sub.141 is independently H or lower alkyl; [0025]
R.sub.51, is H, alkyl, acyl, or aryl; [0026] R.sub.52, R.sub.53,
and R.sub.54 are each independently selected from H, hydroxyl,
alkyl, acyl, and aryl; or any two of R.sub.52, R.sub.53, and
R.sub.54 together form .dbd.O or .dbd.C(R.sub.57)(R.sub.58), where
R.sub.57 and R.sub.58 are each independently selected from H,
alkyl, CO.sub.2(C.sub.1-C.sub.6)alkyl, C(O)N(C.sub.1-C.sub.6)alkyl,
and CO.sub.2(aryl); and [0027] R.sub.55 and R.sub.56 are each
independently H or a suitable protecting group for nitrogen. The
invention is also directed to pharmaceutically acceptable salts of
the compounds of formulae II and II'.
[0028] The invention also relates to pharmaceutical compositions
containing a therapeutically effective amount of at least one
compound of the formula I, or a prodrug, pharmaceutically
acceptable salt, pharmaceutically active metabolite, or solvate
thereof. Additionally, the invention relates to methods of
inhibiting picornaviral 3C protease by administering a
therapeutically effective amount of at least one compound of the
formula I, or a prodrug, pharmaceutically acceptable salt,
pharmaceutically active metabolite, or solvate thereof.
DETAILED DESCRIPTION AND PREFERRED EMBODIMENTS
[0029] The present invention relates to compounds of the formula I:
##STR10## wherein Y, R.sub.1, R.sub.2, R.sub.3, R.sub.5, R.sub.6,
R.sub.7, R.sub.8, R.sub.9, Z, and Z.sub.1 are as defined above, and
to pharmaceutically acceptable salts, prodrugs, active metabolites,
and solvates thereof. Preferably, such compounds, pharmaceutically
acceptable salts, prodrugs, active metabolites, and solvates have
antipicornaviral activity, more preferably antirhinoviral activity,
corresponding to an EC.sub.50 less than or equal to 100 .mu.M in
the H1-HeLa cell culture assay, more preferably corresponding to an
EC.sub.50 less than or equal to 10 .mu.M in the H1-HeLa cell
culture assay.
[0030] The present invention additionally relates to preferred
compounds of the formulas I-A, I-B, and I-C: ##STR11## wherein
R.sub.y (in formula I-A) is H or lower alkyl, and R.sub.1, R.sub.2,
R.sub.3, R.sub.5, R.sub.6, R.sub.7, R.sub.8, R.sub.9, Z, and
Z.sub.1 are as defined above, and to pharmaceutically acceptable
salts, prodrugs, active metabolites, and solvates thereof.
[0031] The inventive compounds of formulas I-A, which are referred
to herein as "peptide-like" compounds, I-B, which are referred to
herein as "ketomethylene-type" compounds, and I-C, which are
referred to herein as "depsipeptide" compounds, differ in their
backbones, which may affect the specific biodistribution or other
physical properties; nonetheless each possesses a strong
rhinoviral-protease-inhibiting activity.
[0032] In preferred embodiments of compounds of formulas I-A, I-B,
and I-C above: [0033] R.sub.1 is H, F, or an alkyl group; [0034]
R.sub.y (in formula I-A) is H or methyl; [0035] R.sub.3, R.sub.5,
and R.sub.8 are each H; [0036] R.sub.2 is selected from one of the
following moieties: ##STR12## [0037] R.sub.6 is an alkyl group,
which has as a preferred optional substituent an aryl group; [0038]
R.sub.7 is an alkyl group, a cycloalkyl group, a heterocycloalkyl
group, an aryl group, or a heteroaryl group; [0039] R.sub.9 is a
five-membered heterocycle having from one to three heteroatoms
selected from O, N, and S, preferably where at least one of the
heteroatoms is nitrogen, that is unsubstituted or substituted,
where the optional substituents are preferably halogen or lower
alkyl, and more preferably mono-chloro or -fluoro or a methyl
group; and [0040] Z and Z.sub.1 are each independently H, F, an
alkyl group, a cycloalkyl group, a heterocycloalkyl group, an aryl
group, a heteroaryl group, --C(O)R.sub.21, --CO.sub.2R.sub.21,
--CN, --C(O)NR.sub.21R.sub.22, --C(O)NR.sub.21OR.sub.22,
--C(S)R.sub.21, --C(S)NR.sub.21R.sub.22, --NO.sub.2, --SOR.sub.21,
--SO.sub.2R.sub.21, --SO.sub.2NR.sub.21R.sub.22,
--SO(NR.sub.21)(OR.sub.22), --SONR.sub.21, --SO.sub.3R.sub.21,
--PO(OR.sub.21).sub.2, --PO(R.sub.21)(R.sub.22),
--PO(NR.sub.21R.sub.22)(OR.sub.23),
--PO(NR.sub.21R.sub.22)(NR.sub.23R.sub.24),
--C(O)NR.sub.21NR.sub.22R.sub.23, or
--C(S)NR.sub.21NR.sub.22R.sub.23, where Z and Z.sub.1 are not both
H, and where R.sub.21, R.sub.22, R.sub.23, and R.sub.24 are each
independently H, an alkyl group, a cycloalkyl group, a
heterocycloalkyl group, an aryl group, a heteroaryl group, an acyl
group, or a thioacyl group, or where any two of R.sub.21, R.sub.22,
R.sub.23, and R.sub.24, together with the atom(s) to which they are
bonded, form a heterocycloalkyl group, or Z and Z.sub.1 (both as
defined above), together with the atoms to which they are attached,
form a heterocycloalkyl group.
[0041] In preferred embodiments, the compounds of the invention are
of the formulae I-A', I-B', and I-C': ##STR13## wherein: [0042]
R.sub.1, Z, and Z.sub.1 are as defined above; [0043] n is 1 or 2;
[0044] R.sub.y (in formula I-A') is H or lower alkyl; [0045]
R.sub.6 is alkyl, cycloalkyl, heterocycloalkyl, aryl, or
heteroaryl; [0046] R.sub.7 is alkyl, cycloalkyl, heterocycloalkyl,
aryl, heteroaryl, --OR.sub.17, --SR.sub.17, --NR.sub.17R.sub.18,
--NR.sub.19NR.sub.17R.sub.18, or --NR.sub.17OR.sub.18, where
R.sub.17, R.sub.18, and R.sub.19 are each independently H, alkyl,
cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or acyl; [0047]
R.sub.9 is a five-membered heterocycle having one to three
heteroatoms selected from O, N, and S, that is unsubstituted or
substituted, where the optional substituents are preferably one or
two lower alkyl groups and/or halogens; and [0048] R.sub.20 is
H.
[0049] The invention also relates to prodrugs, pharmaceutically
acceptable salts, pharmaceutically active metabolites, and solvate
of such compounds.
[0050] In preferred embodiments, the RVP-inhibiting agents of the
invention are compounds of any of the stereospecific formulas
I-A'', I-B'', and I-C'': ##STR14## wherein R.sub.y, R.sub.1,
R.sub.2, R.sub.6, R.sub.7, R.sub.9, Z, and Z, are as defined above,
and pharmaceutically acceptable salts, prodrugs, active
metabolites, and solvates thereof.
[0051] In preferred embodiments of compounds of the formula I-A'',
I-B'', or I-C'': [0052] R.sub.1 is H, F, or methyl; [0053] Ry (in
formula I-A') is H or methyl; [0054] R.sub.2 is selected from one
of the following moieties: ##STR15## [0055] R.sub.6 is arylmethyl
or arylthiomethyl, where aryl is preferably an optionally
substituted phenyl group; [0056] R.sub.7 is an alkyl group, more
preferably selected from 2-propyl, 2-methyl-2-propyl,
2-methyl-1-propyl, and arylmethyl, where the aryl group is
preferably phenyl or naphthyl; [0057] R.sub.9 is isoxazolyl,
oxazolyl, or oxadiazolyl, optionally substituted with one or two
lower alkyl groups and/or halogens; and [0058] Z is H, and Z.sub.1
is --CO.sub.2R.sub.21, --CN, or --C(O)NR.sub.21R.sub.22, where
R.sub.21 and R.sub.22 are as defined above, or Z and Z.sub.1
together form a cyclic ester or amide.
[0059] Even more preferably, the RVP-inhibiting agents of the
invention are compounds of any of the formulas I-A''', I-B''', and
I-C''': ##STR16## wherein n, R.sub.y, R.sub.1, R.sub.20, R.sub.6,
R.sub.7, R.sub.9, Z, and Z.sub.1 are as defined above, and
pharmaceutically acceptable salts, prodrugs, active metabolites,
and solvates thereof.
[0060] In preferred compounds of the formula (I-A'''), (I-B'''), or
(I-C'''): [0061] R.sub.1 is H, F, or methyl; [0062] R.sub.y (in
formula I-A') is H or methyl; [0063] R.sub.20 is hydrogen; [0064]
R.sub.6 is arylmethyl or arylthiomethyl, where aryl is preferably
phenyl unsubstituted or substituted with halogen, lower alkyl,
and/or lower alkoxy; [0065] R.sub.7 is an alkyl group, and more
preferably is selected from 2-propyl, 2-methyl-2-propyl,
2-methyl-1-propyl, and arylmethyl, where the aryl group is
preferably phenyl or naphthyl; [0066] R.sub.9 is isoxazolyl,
oxazolyl, or oxadiazolyl, each optionally substituted with one or
two lower alkyl groups and/or halogens; and [0067] Z is H, and Z,
is --CO.sub.2R.sub.21, --CN, or --C(O)NR.sub.21R.sub.22, where
R.sub.21 and R.sub.22 are as defined above, or Z and Z, together
form a cyclic ester or amide.
[0068] In especially preferred compounds of the invention of the
generic formula I (and subgeneric formulae), R.sub.1 is H or F.
[0069] In another aspect, the invention is directed to intermediate
compounds of the formulas II and II': ##STR17## wherein the
variables (p, A.sub.11, A.sub.12, A.sub.13, A.sub.14, R.sub.51,
R.sub.52, R.sub.53, R.sub.54, R.sub.55, R.sub.56, and R.sub.141)
are as defined above. These compounds are useful for synthesizing
pharmaceutically useful compounds of the formula I.
[0070] Preferred R.sub.55 and R.sub.56 groups are H and suitable
protecting groups for nitrogen, for example, BOC
(t-butyloxycarbonyl), CBZ (benzyloxycarbonyl), FMOC
(fluorene-9-methyloxycarbonyl), other alkyloxycarbonyls (e.g.
methyloxycarbonyl), and trityl (triphenylmethyl). Other suitable
nitrogen-protecting groups may be readily selected by artisans
(see, e.g., Greene and Wutz, Protecting Groups in Chemical
Synthesis (2.sup.nd ed.), John Wiley & Sons, NY (1991)).
Preferred groups for R.sub.52, R.sub.53, and R.sub.54 are H,
alkoxy, hydroxy, and carbonyl.
[0071] Preferred formula-II compounds include the following, where
P.sub.N is a suitable protecting group for nitrogen and q is 1 or
2: ##STR18## ##STR19##
[0072] Other preferred intermediates include the following
compounds, where BOC is t-butyloxycarbonyl: ##STR20##
[0073] Of these, the preferred stereoisomers are: ##STR21##
[0074] Especially preferred intermediates include the following
compounds: ##STR22## ##STR23##
[0075] In accordance with a convention used in the art, ##STR24##
is used in structural formulas herein to depict the bond that is
the point of attachment of the moiety or substituent to the core or
backbone structure.
[0076] Where chiral carbons are included in chemical structures,
unless a particular orientation is depicted, both stereoisomeric
forms are intended to be encompassed.
[0077] As used herein, the term "alkyl group" is intended to mean a
straight- or branched-chain monovalent radical of saturated and/or
unsaturated carbon atoms and hydrogen atoms, such as methyl (Me),
ethyl (Et), propyl, isopropyl, butyl, isobutyl, t-butyl, ethenyl,
pentenyl, butenyl, propenyl, ethynyl, butynyl, propynyl, pentynyl,
hexynyl, and the like, which may be unsubstituted (i.e., containing
only carbon and hydrogen) or substituted by one or more suitable
substituents as defined below (e.g., one or more halogens, such as
F, Cl, Br, or I, with F and Cl being preferred). A "lower alkyl
group" is intended to mean an alkyl group having from 1 to 4 carbon
atoms in its chain.
[0078] A "cycloalkyl group" is intended to mean a non-aromatic
monovalent monocyclic, bicyclic, or tricyclic radical containing 3,
4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14 carbon ring atoms, each of
which may be saturated or unsaturated, and which may be
unsubstituted or substituted by one or more suitable substituents
as defined below, and to which may be fused one or more
heterocycloalkyl groups, aryl groups, or heteroaryl groups, which
themselves may be unsubstituted or substituted by one or more
substituents. Illustrative examples of cycloalkyl groups include
the following moieties: ##STR25##
[0079] A "heterocycloalkyl group" is intended to mean a
non-aromatic monovalent monocyclic, bicyclic, or tricyclic radical,
which is saturated or unsaturated, containing 3, 4, 5, 6, 7, 8, 9,
10, 11, 12, 13, 14, 15, 16, 17, or 18 ring atoms, which includes 1,
2, 3, 4, or 5 heteroatoms selected from nitrogen, oxygen, and
sulfur, where the radical is unsubstituted or substituted by one or
more suitable substituents as defined below, and to which may be
fused one or more cycloalkyl groups, aryl groups, or heteroaryl
groups, which themselves may be unsubstituted or substituted by one
or more suitable substituents. Illustrative examples of
heterocycloalkyl groups include the following moieties:
##STR26##
[0080] An "aryl group" is intended to mean an aromatic monovalent
monocyclic, bicyclic, or tricyclic radical containing 6, 10, 14, or
18 carbon ring atoms, which may be unsubstituted or substituted by
one or more suitable substituents as defined below, and to which
may be fused one or more cycloalkyl groups, heterocycloalkyl
groups, or heteroaryl groups, which themselves may be unsubstituted
or substituted by one or more suitable substituents. Illustrative
examples of aryl groups include the following moieties:
##STR27##
[0081] A "heteroaryl group" is intended to mean an aromatic
monovalent monocyclic, bicyclic, or tricyclic radical containing 4,
5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 ring atoms,
including 1, 2, 3, 4, or 5 heteroatoms selected from nitrogen,
oxygen, and sulfur, which may be unsubstituted or substituted by
one or more suitable substituents as defined below, and to which
may be fused one or more cycloalkyl groups, heterocycloalkyl
groups, or aryl groups, which themselves may be unsubstituted or
substituted by one or more suitable substituents. Illustrative
examples of heteroaryl groups include the following moieties:
##STR28##
[0082] A "heterocycle" is intended to mean a heteroaryl or
heterocycloalkyl group (each of which, as defined above, are
optionally substituted).
[0083] An "acyl group" is intended to mean a --C(O)--R radical,
where R is a substituent as defined below.
[0084] A "thioacyl group" is intended to mean a --C(S)--R radical,
where R is a substituent as defined below.
[0085] A "sulfonyl group" is intended to mean a --SO.sub.2R
radical, where R is a substituent as defined below.
[0086] A "hydroxy group" is intended to mean the radical --OH.
[0087] An "amino group" is intended to mean the radical
--NH.sub.2.
[0088] An "alkylamino group" is intended to mean the radical
--NHR.sub.a, where R.sub.a is an alkyl group.
[0089] A "dialkylamino group" is intended to mean the radical
--NR.sub.aR.sub.b, where R.sub.a and R.sub.b are each independently
an alkyl group.
[0090] An "alkoxy group" is intended to mean the radical
--OR.sub.a, where R.sub.a is an alkyl group. Exemplary alkoxy
groups include methoxy, ethoxy, propoxy, and the like.
[0091] An "alkoxycarbonyl group" is intended to mean the radical
--C(O)OR.sub.a, where R.sub.a is an alkyl group.
[0092] An "alkylsulfonyl group" is intended to mean the radical
--SO.sub.2R.sub.a, where R.sub.a is an alkyl group.
[0093] An "alkylaminocarbonyl group" is intended to mean the
radical --C(O)NHR.sub.a, where R.sub.a is an alkyl group.
[0094] A "dialkylaminocarbonyl group" is intended to mean the
radical --C(O)NR.sub.aR.sub.b, where R.sub.a and R.sub.b are each
independently an alkyl group.
[0095] A "mercapto group" is intended to mean the radical --SH.
[0096] An "alkylthio group" is intended to mean the radical
--SR.sub.a, where R.sub.a is an alkyl group.
[0097] A "carboxy group" is intended to mean the radical
--C(O)OH.
[0098] A "carbamoyl group" is intended to mean the radical
--C(O)NH.sub.2.
[0099] An "aryloxy group" is intended to mean the radical
--OR.sub.c, where R.sub.c is an aryl group.
[0100] A "heteroaryloxy group" is intended to mean the radical
--OR.sub.d, where R.sub.d is a heteroaryl group.
[0101] An "arylthio group" is intended to mean the radical
--SR.sub.c, where R.sub.c is an aryl group.
[0102] A "heteroarylthio group" is intended to mean the radical
--SR.sub.d, where R.sub.d is a heteroaryl group.
[0103] The term "suitable organic moiety" is intended to mean any
organic moiety recognizable, such as by routine testing, to those
skilled in the art as not adversely affecting the inhibitory
activity of the inventive compounds. Illustrative examples of
suitable organic moieties include, but are not limited to, hydroxy
groups, alkyl groups, oxo groups, cycloalkyl groups,
heterocycloalkyl groups, aryl groups, heteroaryl groups, acyl
groups, sulfonyl groups, mercapto groups, alkylthio groups, alkoxy
groups, carboxy groups, amino groups, alkylamino groups,
dialkylamino groups, carbamoyl groups, arylthio groups,
heteroarylthio groups, and the like.
[0104] The term "substituent" or "suitable substituent" is intended
to mean any suitable substituent that may be recognized or
selected, such as through routine testing, by those skilled in the
art. Illustrative examples of suitable substituents include hydroxy
groups, halogens, oxo groups, alkyl groups, acyl groups, sulfonyl
groups, mercapto groups, alkylthio groups, alkoxy groups,
cycloalkyl groups, heterocycloalkyl groups, aryl groups, heteroaryl
groups, carboxy groups, amino groups, alkylamino groups,
dialkylamino groups, carbamoyl groups, aryloxy groups,
heteroaryloxy groups, arylthio groups, heteroarylthio groups, and
the like.
[0105] The term "optionally substituted" is intended to expressly
indicate that the specified group is unsubstituted or substituted
by one or more suitable substituents, unless the optional
substituents are expressly specified, in which case the term
indicates that the group is unsubstituted or substituted with the
specified substituents. As defined above, various groups may be
unsubstituted or substituted (i.e., they are optionally
substituted) unless indicated otherwise herein (e.g., by indicating
that the specified group is unsubstituted).
[0106] A "prodrug" is intended to mean a compound that is converted
under physiological conditions or by solvolysis or metabolically to
a specified compound that is pharmaceutically active.
[0107] A "pharmaceutically active metabolite" is intended to mean a
pharmacologically active product produced through metabolism in the
body of a specified compound.
[0108] A "solvate" is intended to mean a pharmaceutically
acceptable solvate form of a specified compound that retains the
biological effectiveness of such compound. Examples of solvates
include compounds of the invention in combination with water,
isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid,
or ethanolamine.
[0109] A "pharmaceutically acceptable salt" is intended to mean a
salt that retains the biological effectiveness of the free acids
and bases of the specified compound and that is not biologically or
otherwise undesirable. Examples of pharmaceutically acceptable
salts include sulfates, pyrosulfates, bisulfates, sulfites,
bisulfites, phosphates, monohydrogenphosphates,
dihydrogenphosphates, metaphosphates, pyrophosphates, chlorides,
bromides, iodides, acetates, propionates, decanoates, caprylates,
acrylates, formates, isobutyrates, caproates, heptanoates,
propiolates, oxalates, malonates, succinates, suberates, sebacates,
fumarates, maleates, butyne-1,4-dioates, hexyne-1,6-dioates,
benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates,
hydroxybenzoates, methoxybenzoates, phthalates, sulfonates,
xylenesulfonates, phenylacetates, phenylpropionates,
phenylbutyrates, citrates, lactates, .gamma.-hydroxybutyrates,
glycollates, tartrates, methane-sulfonates, propanesulfonates,
naphthalene-1-sulfonates, naphthalene-2-sulfonates, and
mandelates.
[0110] If an inventive compound is a base, a desired salt may be
prepared by any suitable method known to the art, including
treatment of the free base with an inorganic acid, such as
hydrochloric acid; hydrobromic acid; sulfuric acid; nitric acid;
phosphoric acid; and the like, or with an organic acid, such as
acetic acid; maleic acid; succinic acid; mandelic acid; fumaric
acid; malonic acid; pyruvic acid; oxalic acid; glycolic acid;
salicylic acid; pyranosidyl acid, such as glucuronic acid or
galacturonic acid; alpha-hydroxy acid, such as citric acid or
tartaric acid; amino acid, such as aspartic acid or glutamic acid;
aromatic acid, such as benzoic acid or cinnamic acid; sulfonic
acid, such as p-toluenesulfonic acid or ethanesulfonic acid; or the
like.
[0111] If an inventive compound is an acid, a desired salt may be
prepared by any suitable method known to the art, including
treatment of the free acid with an inorganic or organic base, such
as an amine (primary, secondary, or tertiary); an alkali metal or
alkaline earth metal hydroxide; or the like. Illustrative examples
of suitable salts include organic salts derived from amino acids
such as glycine and arginine; ammonia; primary, secondary, and
tertiary amines; and cyclic amines, such as piperidine, morpholine,
and piperazine; as well as inorganic salts derived from sodium,
calcium, potassium, magnesium, manganese, iron, copper, zinc,
aluminum, and lithium.
[0112] In the case of compounds, salts, or solvates that are
solids, it is understood by those skilled in the art that the
inventive compounds, salts, and solvates may exist in different
crystal forms, all of which are intended to be within the scope of
the present invention and specified formulas.
[0113] The inventive compounds may exist as single stereoisomers,
racemates, and/or mixtures of enantiomers and/or diastereomers. All
such single stereoisomers, racemates, and mixtures thereof are
intended to be within the broad scope of the present invention.
Preferably, however, the inventive compounds are used in optically
pure form.
[0114] As generally understood by those skilled in the art, an
optically pure compound is one that is enantiomerically pure. As
used herein, the term "optically pure" is intended to mean a
compound comprising at least a sufficient activity. Preferably, an
optically amount of a single enantiomer to yield a compound having
the desired pharmacological pure compound of the invention
comprises at least 90% of a single isomer (80% enantiomeric
excess), more preferably at least 95% (90% e.e.), even more
preferably at least 97.5% (95% e.e.), and most preferably at least
99% (98% e.e.).
[0115] Preferably in the compounds of the formula I (or of any of
the subgeneric formula), R.sub.1 is H or F.
[0116] In the compounds of formula I, preferably R.sub.9 is an
unsubstituted or substituted isoxazolyl group, where the optional
substituents are preferably one or two methyl groups and/or
halogens.
[0117] Especially preferred embodiments of the invention are
described below in reference to the following formula I-A'':
##STR29##
[0118] Preferred compounds of the present invention include peptido
(peptide-like) Compounds (A-1)-(A-8) of the formula I-A'' above,
wherein R.sub.1 is H, Z is H, R.sub.y is H, and R.sub.2, R.sub.6,
R.sub.7, Z.sub.1, and R.sub.9 are as respectively defined below:
[0119] (A-1) R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is
CH.sub.2Ph, R.sub.7 is CH.sub.2CH(CH.sub.3).sub.2, Z.sub.1 is
CO.sub.2CH.sub.2CH.sub.3, and R.sub.9 is ##STR30## [0120] (A-2)
R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is CH.sub.2Ph,
R.sub.7 is CH.sub.2CH(CH.sub.3).sub.2, Z.sub.1 is
CO.sub.2CH.sub.2CH.sub.3, and R.sub.9 is ##STR31## [0121] (A-3)
R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is ##STR32##
R.sub.7 is C(CH.sub.3).sub.3, Z.sub.1 is CO.sub.2CH.sub.2CH.sub.3,
and R.sub.9 is ##STR33## [0122] (A-4) R.sub.2 is
CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is ##STR34## R.sub.7 is
C(CH.sub.3).sub.3, Z.sub.1 is CO.sub.2CH.sub.2CH.sub.3, and R.sub.9
is ##STR35## [0123] (A-5) R.sub.2 is ##STR36## R.sub.6 is ##STR37##
R.sub.7 is CH(CH.sub.3).sub.2, Z.sub.1 is CO.sub.2CH.sub.2CH.sub.3,
and R.sub.9 is ##STR38## [0124] (A-6) R.sub.2 is
CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is ##STR39## R.sub.7 is
CH(CH.sub.3).sub.2, Z, is CO.sub.2CH.sub.2CH.sub.3, and R.sub.9 is
##STR40## [0125] (A-7) R.sub.2 is ##STR41## R.sub.6 is ##STR42##
R.sub.7 is C(CH.sub.3).sub.3, Z.sub.1 is CO.sub.2CH.sub.2CH.sub.3,
and R.sub.9 is ##STR43## [0126] (A-8) R.sub.2 is ##STR44## R.sub.6,
is ##STR45## R.sub.7 is CH(CH.sub.3).sub.2, Z.sub.1 is
CO.sub.2CH.sub.2CH.sub.3, and R.sub.9 is ##STR46##
[0127] Preferred peptide-like compounds of the formula I-A''''
further include Compounds (A-9)-(A-13) below, wherein R.sub.1 is H,
Z is H, Z, is CO.sub.2CH.sub.2CH.sub.3, R.sub.y is CH.sub.3, and
R.sub.2, R.sub.6, R.sub.7, and R.sub.9 are as respectively defined
below: [0128] (A-9) R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2,
R.sub.6 is ##STR47## R.sub.7 is ##STR48## and R.sub.9 is ##STR49##
[0129] (A-10) R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is
CH.sub.2Ph, R.sub.7 is CH.sub.2CH(CH.sub.3).sub.2, and R.sub.9 is
##STR50## [0130] (A-11) R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2,
R.sub.6 is ##STR51## R.sub.7 is ##STR52## and R.sub.9 is ##STR53##
[0131] (A-12) R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is
##STR54## R.sub.7 is CH.sub.2CH(CH.sub.3).sub.2, and R.sub.9 is
##STR55## [0132] (A-13) R.sub.2 is CH.sub.2CH.sub.2C(O)N NH.sub.2,
R.sub.6 is ##STR56## R.sub.7 is ##STR57## and R.sub.9 is
##STR58##
[0133] Other preferred peptide-like compounds include the
following: ##STR59## ##STR60## ##STR61## ##STR62##
[0134] Preferred ketomethylene-type Compounds (B-1)-(B-4) of the
invention are described below in reference to the following formula
I-B'': ##STR63## (B-1) R.sub.2 is CH.sub.2CH.sub.2C(O)NH.sub.2,
R.sub.6 is ##STR64## R.sub.7 is CH(CH.sub.3).sub.2, Z is H, Z, is
CO.sub.2CH.sub.2CH.sub.3, and R.sub.9 is ##STR65## [0135] (B-2)
R.sub.2 is ##STR66## R.sub.6 is ##STR67## R.sub.7 is
CH(CH.sub.3).sub.2, Z is H, Z, is CO.sub.2CH.sub.2CH.sub.3, and
R.sub.9 is ##STR68## [0136] (B-3) R.sub.2 is ##STR69## R.sub.6 is
##STR70## R.sub.7 is CH(CH.sub.3).sub.2, Z and Z.sub.1 together are
##STR71## where the carbonyl group is cis to the hydrogen
corresponding to R.sub.1 in formula I, and R.sub.9 is ##STR72##
[0137] (B-4) R.sub.2 is ##STR73## R.sub.6 is ##STR74## R.sub.7 is
CH(CH.sub.3).sub.2, Z is H, Z.sub.1 is CO.sub.2CH.sub.2CH.sub.3,
and R.sub.9 is ##STR75##
[0138] Preferred depsipeptide-type Compounds (C-1) and (C-2) of the
invention are described below in reference to the following formula
I-C'', where R.sub.1 is H: ##STR76## [0139] (C-1) Z is H, R.sub.2
is CH.sub.2CH.sub.2C(O)NH.sub.2, R.sub.6 is ##STR77## R.sub.7 is
CH(CH.sub.3).sub.2, Z.sub.1 is CO.sub.2CH.sub.2CH.sub.3, and
R.sub.9 is ##STR78## [0140] (C-2) Z is H, R.sub.2 is ##STR79##
R.sub.6 is ##STR80## R.sub.7 is CH(CH.sub.3).sub.2, Z.sub.1 is
CO.sub.2CH.sub.2CH.sub.3, and R.sub.9 is ##STR81##
[0141] Additional compounds may be prepared in reference to formula
I by selecting the variables from the following substituents:
[0142] R.sub.y.dbd.H or CH.sub.3; [0143] R.sub.1 .dbd.H or
CH.sub.3; ##STR82## or phenylmethyl (i.e., benzyl), where the aryl
group is optionally substituted with one, two, or three
substituents each independently selected from halogens, methoxy and
methyl; [0144] R.sub.7=2-methyl-1-propyl, 2-propyl,
2-methyl-2-propyl, benzyl, or ##STR83##
[0145] The present invention is also directed to a method of
inhibiting picornaviral 3C protease activity, comprising contacting
the protease with an effective amount of a compound of formula I,
or a pharmaceutically acceptable salt, prodrug, pharmaceutically
active metabolite, or solvate thereof. For example, picornaviral 3C
protease activity may be inhibited in mammalian tissue by
administering a compound of formula I or a pharmaceutically
acceptable salt, prodrug, pharmaceutically active metabolite, or
solvate thereof. More preferably, the present method is directed at
inhibiting rhinoviral protease activity.
[0146] "Treating" or "treatment" is intended to mean at least the
mitigation of a disease condition in a mammal, such as a human,
that is alleviated by the inhibition of the activity of one or more
picornaviral 3C proteases, such as human rhinoviruses, human
poliovirus, human coxsackieviruses, encephalomyocarditis viruses,
meningitis virus, and hepatitis A virus, and includes: (a)
prophylactic treatment in a mammal, particularly when the mammal is
found to be predisposed to having the disease condition but not yet
diagnosed as having it; (b) inhibiting the disease condition;
and/or (c) alleviating, in whole or in part, the disease
condition.
[0147] The activity of the inventive compounds as inhibitors of
picornaviral 3C protease activity may be measured by any of the
suitable methods known to those skilled in the art, including in
vivo and in vitro assays. An example of a suitable assay for
activity measurements is the antiviral H1-HeLa cell culture assay
described herein.
[0148] Administration of the compounds of the formula I and their
pharmaceutically acceptable prodrugs, salts, active metabolites,
and solvates may be performed according to any of the accepted
modes of administration available to those skilled in the art.
Illustrative examples of suitable modes of administration include
oral, nasal, parenteral, topical, transdermal, and rectal.
Intranasal delivery is especially preferred.
[0149] An inventive compound of formula I or a pharmaceutically
acceptable salt, prodrug, active metabolite, or solvate thereof may
be administered as a pharmaceutical composition in any
pharmaceutical form recognizable to the skilled artisan as being
suitable. Suitable pharmaceutical forms include solid, semisolid,
liquid, or lyophilized formulations, such as tablets, powders,
capsules, suppositories, suspensions, liposomes, and aerosols.
Pharmaceutical compositions of the invention may also include
suitable excipients, diluents, vehicles, and carriers, as well as
other pharmaceutically active agents, depending upon the intended
use. In preferred embodiments, the inventive pharmaceutical
compositions are delivered intranasally in the form of
suspensions.
[0150] Acceptable methods of preparing suitable pharmaceutical
forms of the pharmaceutical compositions are known or may be
routinely determined by those skilled in the art. For example,
pharmaceutical preparations may be prepared following conventional
techniques of the pharmaceutical chemist involving steps such as
mixing, granulating, and compressing when necessary for tablet
forms, or mixing, filling, and dissolving the ingredients as
appropriate, to give the desired products for oral, parenteral,
topical, intravaginal, intranasal, intrabronchial, intraocular,
intraaural, and/or rectal administration.
[0151] Solid or liquid pharmaceutically acceptable carriers,
diluents, vehicles, or excipients may be employed in the
pharmaceutical compositions. Illustrative solid carriers include
starch, lactose, calcium sulfate dihydrate, terra alba, sucrose,
talc, gelatin, pectin, acacia, magnesium stearate, and stearic
acid. Illustrative liquid carriers include syrup, peanut oil, olive
oil, saline solution, and water. The carrier or diluent may include
a suitable prolonged-release material, such as glyceryl
monostearate or glyceryl distearate, alone or with a wax. When a
liquid carrier is used, the preparation may be in the form of a
syrup, elixir, emulsion, soft gelatin capsule, sterile injectable
liquid (e.g., solution), or a nonaqueous or aqueous liquid
suspension.
[0152] A dose of the pharmaceutical composition contains at least a
therapeutically effective amount of the active compound (i.e., a
compound of formula I or a pharmaceutically acceptable salt,
prodrug, active metabolite, or solvate thereof), and preferably is
made up of one or more pharmaceutical dosage units. The selected
dose may be administered to a mammal, for example, a human patient,
in need of treatment mediated by inhibition of picornaviral 3C
protease activity, by any known or suitable method of administering
the dose, including topically, for example, as an ointment or
cream; orally; rectally, for example, as a suppository;
parenterally by injection; or continuously by intravaginal,
intranasal, intrabronchial, intraaural, or intraocular
infusion.
[0153] A "therapeutically effective amount" is intended to mean the
amount of an inventive compound that, when administered to a mammal
in need thereof, is sufficient to effect treatment for disease
conditions alleviated by the inhibition of the activity of one or
more picornaviral 3C proteases, such as human rhinoviruses, human
poliovirus, human coxsackieviruses, encephalomyocarditis viruses,
menigovirus, and hepatitis A virus. The amount of a given compound
of the invention that will be therapeutically effective will vary
depending upon factors such as the particular compound, the disease
condition and the severity thereof, the identity of the mammal in
need thereof, which amount may be routinely determined by
artisans.
[0154] By way of illustration, a formulation for nasal delivery of
the inventive compounds for treatment of rhinoviral infections can
be prepared as follows, where all percentages are weight/weight and
the suspension is prepared in purified water. A formula-I compound
is micronized to a reduced particle size such that D.sub.90<10
.mu.m. A suspension is prepared to contain a final concentration of
from about 0.01% to about 2% of the active compound, preferably
about from 0.2% to 2%. An appropriate preservative selected from
those known in the art may be included, for example, benzalkonium
chloride/EDTA, in appropriate final-concentration ranges, e.g.,
about 0.02%/0.01%. A suspending agent, such as mixture of
microcrystalline cellulose (final concentration of about 1%-1.5%,
preferably about 1.2%) and sodium carboxymethylcellulose cellulose
(final concentration of about 0.1%-0.2%, preferably about 0.13%)
may be included. A surfactant such as polysorbate 80 may be
included in a final concentration of about from 0.05% to 0.2%,
preferably about 0.1%. A tonicity modifier such as dextrose may be
included to give a final concentration of about from 4% to 6%,
preferably about 5%. The pH of the final solution is adjusted as
appropriate to a physiological range, e.g., 4-6, using non-toxic
acid and/or base, such as HCl and/or NaOH.
[0155] An exemplary formulation for nasal delivery of the inventive
compound of Example 17 has the following composition, where all
percentages are weight/weight and the suspension is prepared in
purified water: TABLE-US-00001 Active Compound (B-2) 0.2-2%
Preservative Benzalkonium 0.02%/0.01% chloride/EDTA Suspending
Agent Microcrystalline cellulose/Na- 1.2%/0.13%
carboxymethylcellulose Surfactant Polysorbate 80 0.1% Tonicity
Modifier Dextrose 5% pH Adjustment NaOH/HCl pH 4-6
General Syntheses
[0156] The inventive compounds of formula (I) may be advantageously
prepared by the methods of the present invention, including the
general methods described below. In each of these general methods,
R.sub.1, R.sub.2, R.sub.3, R.sub.5, R.sub.6, R.sub.7, R.sub.8,
R.sub.9, R.sub.y, Z, and Z.sub.1 are as defined above, and R.sub.4
is used (as a shorthand representation) to mean: ##STR84## where
R.sub.7, R.sub.8, and R.sub.9 are as defined above.
[0157] In General Method I, useful for synthesis of peptide-like
compounds of formula I-A, an amino acid A, where P.sub.1 is an
appropriate protecting group for nitrogen, is subjected to an
amide-forming reaction with amino alcohol (or salt thereof) B to
produce amide C. Compound C is then deprotected to give free amine
(or salt thereof) D. Amine D and amino acid E, which may
incorporate either an R.sub.4 group or a protecting group for
nitrogen (P.sub.2), are subjected to a bond-forming reaction
generating compound F. Compound F is oxidized to intermediate G,
which is then transformed into unsaturated product H. If protecting
groups have been used on amino acid E, or on any R groups
(R.sub.1-R.sub.9) and/or on R.sub.y and/or on Z and/or Z.sub.1,
product H is deprotected and/or further modified to yield
deprotected or modified H. ##STR85##
[0158] An alternative method to prepare intermediate F is described
as follows. Amino acid E and amino acid (or salt thereof) I, where
P.sub.3 is an appropriate protecting group for oxygen, are
subjected to a bond-forming reaction to produce intermediate J.
Molecule J is deprotected to yield free carboxylic acid K, which is
subsequently subjected to an amide-forming reaction with amino
alcohol (or salt thereof) B to generate intermediate F.
##STR86##
[0159] In General Method II, which is also useful for synthesizing
peptide-like compounds of formula I-A, an amino acid L, where
P.sub.1 is an appropriate protecting group for nitrogen, is
converted to a carbonyl derivative M, where "Lv" is a leaving
group. Compound M is subjected to a reaction where Lv is replaced
by R.sub.1 to give derivative N. Derivative N is then transformed
into unsaturated product O. Unsaturated compound O is deprotected
to give free amine (or salt thereof) P, or modified at Z or Z.sub.1
first to give O' and then deprotected to P. Intermediate P is
subsequently subjected to an amide-forming reaction with carboxylic
acid K to give final product H. If protecting groups have been used
on any R group (R.sub.1-R.sub.9) and/or on R.sub.y and/or on Z
and/or Z.sub.1, product H is deprotected and/or further modified to
yield deprotected or modified H. ##STR87##
[0160] An alternative method to prepare intermediate N is described
as follows. Compound M is subjected to a reaction where "Lv" (or
more particularly --C(O)-Lv), is reduced to protected amino alcohol
Q. Intermediate Q is subsequently oxidized to derivative N.
##STR88##
[0161] In General Method III, useful for synthesis of peptide-like
compounds of formula I-A, an amino acid L, where P.sub.1 is an
appropriate protecting group for nitrogen, is converted to a
carbonyl derivative M, where "Lv" is a leaving group. Compound M is
deprotected to give free amine (or salt thereof) R, which
subsequently is subjected to an amide-forming reaction with
carboxylic acid K to give intermediate S. Compound S is then either
directly converted to carbonyl intermediate G, or successively
reduced to alcohol F first, which is oxidized to G. Compound G is
subjected to a reaction to yield the unsaturated final product H.
If protecting groups have been used on any R groups
(R.sub.1-R.sub.9) and/or on R.sub.y and/or on Z and/or Z.sub.1,
product H is deprotected and/or further modified to yield
deprotected or modified H. ##STR89##
[0162] In General Method IV, useful for synthesis of peptide-like
compounds of formula I-A, free amine (or salt thereof) P, prepared
from intermediate 0 as described in General Method II, is converted
to amide T by reaction with amino acid A, where P.sub.1 is an
appropriate protecting group for nitrogen. Compound T is further
deprotected to free amine (or salt thereof) U, which is
subsequently converted to H with amino acid E. If protecting groups
have been used on any R groups (R.sub.1-R.sub.9) and/or on Ry
and/or on Z and/or Z.sub.1, product H is deprotected and/or further
modified to yield deprotected or modified H. ##STR90##
[0163] In General Method V, useful for synthesis of ketomethylene
compounds of formula I-B, optically active lactone AA, where
P.sub.4 is an appropriate protecting group for nitrogen, and
R.sub.5 and R.sub.8 are H (which may be prepared by the method
described below and by various literature methods, including: (a)
Herold et al., J. Org. Chem. 1989, 54, 1178; (b) Bradbury et al.,
Tetrahedron Lett. 1989, 30, 3845; (c) Bradbury et al., J. Med.
Chem. 1990, 33, 2335; (d) Wuts et al., J. Org. Chem. 1992, 57,
6696; (e) Jones et al., J. Org. Chem. 1993, 58, 2286; (f) Pegorier
et al., Tetrahedron Lett. 1995, 36, 2753; and (g) Dondoni et al.,
J. Org. Chem. 1995, 60, 7927) is transformed by a two-step
procedure (basic hydrolysis and subsequent oxidation) into
carboxylic acid BB. This material is not isolated, but is subjected
to an amide-forming reaction with amine (or salt thereof) P to
provide final product CC. The P.sub.4 protecting group, along with
any additional protecting groups that have been used on any R
groups (R.sub.1, R.sub.2, R.sub.3, R.sub.6, and/or R.sub.7) and/or
on Z and/or on Z.sub.1, is subsequently deprotected and/or further
modified to yield deprotected or modified CC. ##STR91##
[0164] Lactone AA may be prepared in optically active form by the
following General Method VI (see: Herold et al., J. Org. Chem.
1989, 54, 1178; Bradbury et al., Tetrahedron Lett. 1989, 30, 3845;
and Bradbury et al., J. Med. Chem. 1990, 33, 2335). A
.gamma.,.delta.-unsaturated carboxylic acid DD, which incorporates
R.sub.7, is transformed into the corresponding acid chloride (not
shown). This acid chloride is subjected to an amide-forming
reaction with a chiral amine or a chiral oxazolidone to provide
derivative EE (in which X.sub.1 is a chiral amine or a chiral
oxazolidone). Compound EE is subsequently deprotonated, and the
resulting enolate is diastereoselectively alkylated with an
electrophile corresponding to R.sub.6 to provide compound FF, where
R.sub.5 is H. This material is then subjected to a
halolactonization reaction to provide halo-lactone GG, in which
R.sub.5 and R.sub.8 are H and "hal" is Br or I. Halo-lactone GG is
subsequently transformed into azide HH, and this material is then
converted into lactone AA, where P.sub.4 is an appropriate
protecting group for nitrogen. ##STR92##
[0165] .gamma.,.delta.-Unsaturated carboxylic acid DD may be
prepared by the following General Method VII (see: Herold et al.,
J. Org. Chem. 1989, 54, 1178). An aldehyde II, which incorporates
R.sub.7, is coupled with vinylmagnesium bromide to give alcohol JJ.
Alcohol JJ is then transformed into .gamma.,.delta.-unsaturated
carboxylic acid DD by a three-step procedure as follows: (i)
treatment with diethyl malonate and catalytic Ti(OEt).sub.4 at
160.degree. C. for 1 hour, (ii) heating at 190.degree. C. for 4
hours, and (iii) hydrolysis with ethanolic KOH at reflux.
##STR93##
[0166] Carboxylic acid BB may also be prepared by General Method
VIII (see Hoffman et al., Tetrahedron, 1997, 53, 7119). An amino
acid KK, which incorporates R.sub.7 and where P.sub.4 is an
appropriate protecting group for nitrogen, is transformed into
.beta.-ketoester LL. Compound LL is deprotonated and the resulting
anion is condensed with triflate MM, which incorporates R.sub.6.
The coupling product thus obtained is treated with trifluoroacetic
acid to provide ketoester NN, and this material is subsequently
hydrolyzed to afford carboxylic acid BB. If basic hydrolysis
results in epimerization, ketoester NN can be transesterified
(allyl alcohol, Ti(Oi-Pr).sub.4) and subsequently deprotected under
neutral conditions (Pd(PPh.sub.3).sub.4, morpholine) to give
carboxylic acid BB. Triflate MM, in turn, may be prepared from the
corresponding alcohol by treatment with trifluoromethanesulfonic
anhydride and 2,6-lutidine. ##STR94##
[0167] Lactone AA may also be prepared by General Method IX (see:
Askin et al., J. Org. Chem. 1992, 57, 2771; and McWilliams et al.,
J. Am. Chem. Soc. 1996, 118, 11970). An amino acid KK, which
incorporates R.sub.7 and where P.sub.4 is an appropriate protecting
group for nitrogen, is transformed into epoxide OO (single
diastereomer) by the method described in Luly et al., J. Org. Chem.
1987, 52, 1487. Epoxide OO is condensed with the anion derived from
compound PP, which incorporates R.sub.6 and in which X.sub.2 is a
chiral auxiliary (including (1S,2R)-1-aminoindan-2-ol acetonide) to
afford coupling product QQ. This material is subsequently cyclized
under acidic conditions to provide lactone AA. Compound PP may be
prepared from the corresponding carboxylic acid (not shown) by the
method outlined in Askin et al., J. Org. Chem. 1992, 57, 2771.
##STR95##
[0168] General Method X, useful in preparation of depsipeptide
compounds of the formula I-C, illustrates a method to prepare
intermediate TT. Amino acid E and alcohol RR, where P.sub.5 is an
appropriate protecting group for oxygen, are subjected to an ester
bond-forming reaction to produce intermediate SS. Molecule SS is
deprotected to yield free carboxylic acid TT, which may be utilized
in lieu of carboxylic acid K in any of the general methods
described above. ##STR96## Specific Syntheses
[0169] The following specific methods may also be used to prepare
various compounds according to the invention.
[0170] Specific Method I describes the preparation of specific
intermediate O1, which may be utilized as intermediate O in the
general methods described above. Thus, ester A1 (prepared as
described in Chida et al., J. Chem. Soc., Chem. Commun. 1992, 1064)
is hydrolyzed to give acid B1, which, in turn, is transformed into
oxazolidinone C1. Compound C1 is subsequently deprotonated, and the
resulting enolate is diastereoselectively alkylated to give allyl
intermediate D1. This entity is oxidized via ozonolysis, and the
resulting aldehyde (not shown) is subjected to a reductive
amination reaction producing lactam E1. Acid-catalyzed methanolysis
of E1 then affords alcohol F1. This intermediate is oxidized via
the method of Swern (or other suitable oxidation conditions) to the
resulting aldehyde (not shown), which is subsequently subjected to
an olefin-forming reaction to provide specific intermediate O1.
##STR97##
[0171] Specific Method II describes the preparation of specific
intermediate O2, which may be utilized as intermediate O in the
general methods described above. Allyl intermediate D1 is subjected
to a hydroboration/oxidation sequence to afford a primary alcohol
(not shown). This entity is oxidized via the method of Swern (or
other suitable oxidation conditions), and the resulting aldehyde
(not shown) is subjected to a reductive-amination reaction,
producing lactam G1. Acid-catalyzed methanolysis of G1 then affords
alcohol H1. This intermediate is oxidized via the method of Swern
(or other suitable oxidation conditions) to the resulting aldehyde
(not shown), which is subsequently subjected to an olefin-forming
reaction to provide specific intermediate O2. ##STR98##
[0172] The following intermediates P1, P2, and P3 may be used in
the above general methods in place of intermediate O, to vary the
substituent group in the R.sub.2 position. ##STR99##
[0173] A synthesis of intermediate P1 is described below.
Intermediate C1 (described above) is deprotonated, and the
resulting enolate is trapped with an appropriate disulfide
(symmetrical or mixed) to give sulfide p1 (P is a suitable
protecting group for oxygen). The oxygen-protecting group is then
removed to give alcohol p2. This intermediate is oxidized via the
method of Swern (or using other suitable oxidation conditions), and
the resulting aldehyde (not shown) is subjected to a reductive
amination reaction to give lactam p3. Acid-catalyzed methanolysis
of p3 then affords alcohol p4. This intermediate is oxidized via
the method of Swern (or using other suitable oxidation conditions)
to the resulting aldehyde (also not shown), which is subsequently
subjected to an olefin-forming reaction to provide intermediate p5.
This compound may be utilized in place of intermediate O in the
above general synthetic methods; alternatively, the
lactam-protecting group may be removed to give intermediate P1.
##STR100##
[0174] To synthesize intermediate P2, intermediate C1 is
deprotonated, and the resulting enolate is trapped with an
appropriate source of electrophilic oxygen (e.g., an oxaziridine)
to give alcohol p6. This intermediate is alkylated with a suitably
functionalized alkyl halide or triflate to give ether p7 (P is an
appropriate protecting group for nitrogen). The nitrogen-protecting
group is then removed, and the resulting amine (not shown) is
subjected to cyclization conditions to give lactam p8.
Acid-catalyzed methanolysis of p8 then affords alcohol p9. This
intermediate is oxidized via the method of Swern (or using other
suitable oxidation conditions) to the resulting aldehyde (also not
shown), which is subsequently subjected to an olefin-forming
reaction to provide intermediate P2. ##STR101##
[0175] A synthesis of specific intermediate P3 is now described.
Intermediate D1 (described above) is ozonized, and the resulting
aldehyde (not shown) is reduced to the corresponding alcohol (also
not shown). This intermediate is then protected to afford compound
p10 (P.sub.1 is an appropriate protecting group for oxygen). The
imide functionality present on p10 is hydrolyzed to carboxylic acid
p11, and this intermediate is coupled with a suitably protected
hydroxylamine derivative to give amide p12 (P.sub.2 is an
appropriate protecting group for oxygen that is stable to
conditions which will remove P.sub.1). The P.sub.1 protecting group
is then removed, and the resulting alcohol (p13) is transformed
into an appropriate leaving group (halide or triflate, not shown).
The P2 protecting group is then removed, and the resulting
hydroxamic acid is cyclized to give intermediate p14.
Acid-catalyzed methanolysis of p14 then affords alcohol p15. This
intermediate is oxidized via the method of Swern (or using other
suitable oxidation conditions) to the resulting aldehyde (not
shown), which is subsequently subjected to an olefin-forming
reaction to provide intermediate P3. ##STR102##
[0176] Specific Method III describes the preparation of
intermediates Q1, Q2, and Q3, which may be utilized in the general
methods described above. The known compound I1 is transformed into
the literature molecule J1 by a modification of a published
procedure (Ikuta et al., J. Med. Chem. 1987, vol. 30, p. 1995).
Independently, the amino acid ester K1 is protected to afford silyl
ether L1. The ether is reduced with DIBAL (or using other suitable
reduction conditions), and the resulting aldehyde (not shown) is
subjected to an olefin-forming reaction with intermediate J1,
producing M1. Silyl deprotection of M1 then affords alcohol N1.
This intermediate is subjected to a variety of hydrogenation
conditions to provide intermediates Q1, Q2, and Q3. These
intermediates may be transformed into intermediates analogous to
intermediate O1 (see Specific Method I above) by oxidation and
subsequent olefination. ##STR103## ##STR104##
[0177] The artisan will recognize that various compounds of the
invention may be made by following the above-described general and
specific methods as well as teachings in the art, including the
references cited herein, the disclosures of which are hereby
incorporated by reference. Additionally, the artisan may prepare
various compounds of the invention according to the example
described below or through routine modifications to the syntheses
described herein.
EXAMPLES
[0178] Examples of various preferred compounds of formula I are set
forth below. The structures of the compounds of the following
examples were confirmed by one or more of the following: proton
magnetic resonance spectroscopy, infrared spectroscopy, elemental
microanalysis, mass spectrometry, thin layer chromatography,
melting-point determination, and boiling-point determination. Where
there is any discrepancy between the given structural formula shown
for a compound and its chemical name provided, the structural
formula applies.
[0179] Proton magnetic resonance (.sup.1H NMR) spectra were
determined using a Varian UNITY plus 300 spectrometer operating at
a field strength of 300 megahertz (MHz). Chemical shifts are
reported in parts per million (ppm, .delta.) downfield from an
internal tetramethylsilane standard. Alternatively, .sup.1H NMR
spectra were referenced to residual protic solvent signals as
follows: CHCl.sub.3=7.26 ppm; DMSO=2.49 ppm; C.sub.6HD.sub.5=7.15
ppm. Peak multiplicities are designated as follows: s=singlet;
d=doublet; dd=doublet of doublets; t=triplet; q=quartet; br=broad
resonance; and m=multiplet. Coupling constants are given in Hertz.
Infrared absorption (IR) spectra were obtained using a Perkin-Elmer
1600 series FTIR spectrometer. Elemental microanalyses were
performed by Atlantic Microlab Inc. (Norcross, Ga.) and gave
results for the elements stated within .+-.0.4% of the theoretical
values. Flash column chromatography was performed using Silica gel
60 (Merck Art 9385). Analytical thin layer chromatography (TLC) was
performed using precoated sheets of Silica 60 F.sub.254 (Merck Art
5719). Melting points (abbreviated as mp) were determined on a
Mel-Temp apparatus and are uncorrected. All reactions were
performed in septum-sealed flasks under a slight positive pressure
of argon, unless otherwise noted. All commercial reagents were used
as received from their respective suppliers with the following
exceptions: tetrahydrofuran (THF) was distilled from
sodium-benzophenone ketyl prior to use; dichloromethane
(CH.sub.2Cl.sub.2) was distilled from calcium hydride prior to use;
anhydrous lithium chloride was prepared by heating at 110.degree.
C. under vacuum overnight.
[0180] The following abbreviations are used herein: Et.sub.2O
refers to diethyl ether; DMF refers to N,N-dimethylformamide; DMSO
refers to dimethylsulfoxide; and MTBE refers to tert-butyl methyl
ether. Other abbreviations include: CH.sub.3OH (methanol), EtOH
(ethanol), EtOAc (ethyl acetate), DME (ethylene glycol dimethyl
ether), Ac (acetyl), Me (methyl), Ph (phenyl), Tr
(triphenylmethyl), Cbz (benzyloxycarbonyl), Boc
(tert-butoxycarbonyl), TBS (tert-butyldimethylsilyl), TFA
(trifluoroacetic acid), DIEA (N,N-diisopropylethylamine), DBU
(1,8-diazabicyclo[5.4.0]undec-7-ene), HOBt (1-hydroxybenzotriazole
hydrate), PyBOP (benzotriazole1-yl-oxy-tris-pyrrolidino-phosphonium
hexafluorophosphate), HATU
(O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate), EDC
(1-(3-dimethylaminopropyl)-3-ethylcarbarbodiimide hydrochloride),
DCC (dicyclohexylcarbodiimide), DDQ
(2,3-dichloro-5,6-dicyano-1,4-benzoquinone), DMAP
(4-dimethylaminopyridine), Gln (glutamine), Leu (leucine), Phe
(phenylalanine), Val (valine), His (histidine), 1-Naphth
(1-naphthlyalanine), 2-Naphth (2-naphthylalanine),
.alpha.-t-Butyl-Gly (tert-butyl glycine), (S)-Pyrrol-Ala
((2S,3'S)-2-amino-3-(2'-oxopyrrolidin-3'-yl)-propionic acid), and
(S)-Piper-Ala
((2S,3'S)-2-amino-3-(2'-oxo-piperidin-3'-yl)-propionic acid).
Additionally, "L" represents naturally occurring amino acids.
[0181] A simplified naming system employing amino acid
abbreviations is used to identify some intermediates and final
products. When naming compounds, italicized amino acid
abbreviations represent modifications at the C-terminus of that
residue where the following apply: (1) acrylic acid esters are
reported as "E" (trans) propenoates; (2) substituted
3-methylene-dihydrofuran-2-ones are reported as "E" (trans)
2-(.alpha.-vinyl-.gamma.-butyrolactones); and (3) 5-vinylisoxazoles
are reported as "E" (trans) propenisoxazoles. In addition, the
terminology "AA.sub.1.PSI.[COCH.sub.2]-AA.sub.2" indicates that,
for any peptide sequence, two amino acids (AA.sub.1 and AA.sub.2)
usually linked by an amide bond are replaced by a ketomethlyene
dipeptide isostere moiety. The terminology
"AA.sub.1-NCH.sub.3-AA.sub.2" indicates that, for any peptide
sequence, the amide bond that usually connects the two amino acids
(AA.sub.1 and AA.sub.2) is replaced by an N-methyl amide linkage.
The terminology "AA.sub.1-O-AA.sub.2" indicates that, for any
peptide sequence, the amide bond that usually connects the two
amino acids (AA.sub.1 and AA.sub.2) is replaced by an ester
linkage.
[0182] Examples of embodiments in accordance with the invention are
described below.
Example 1
Preparation of Comparison Compound #1:
5-(3'-(Cbz-L-Leu-L-Phe-L-Gln)-E-Propene)-isoxazole
[0183] ##STR105##
Preparation of Intermediate Cbz-L-(Tr-Gln)-OMe
[0184] Cbz-L-(Tr-Gln) (0.26 g, 0.50 mmol, 1 equiv.) was added to a
solution of acetyl chloride (0.25 mL, 3.52 mmol, 7.0 equiv.) in
CH.sub.3OH (5 mL), and stirring was continued at 23.degree. C. for
1 h (hour). The solvent was removed under reduced pressure, and the
residue was dissolved in CH.sub.2Cl.sub.2 (100 mL) and washed with
water (100 mL), saturated NaHCO.sub.3 (100 mL), and brine (100 mL).
The organic layer was dried over Na.sub.2SO.sub.4 and was
concentrated. The residue was purified by flash column
chromatography (20% EtOAc in hexanes) to afford Cbz-L-(Tr-Gln)-OMe
(0.23 g, 84% yield) as a white solid: mp=139-140.degree. C.; IR
(cm.sup.-1) 1742, 1207; .sup.1H NMR (DMSO-d.sub.6) .delta. 1.16 (t,
1H, J=7.0), 1.77 (m, 1H), 1.97 (m, 1H), 3.61 (s, 3H), 4.99 (m, 1H),
5.03 (s, 2H), 7.02-7.55 (m, 20H), 7.69 (d, 1H, J=7.7), 8.59 (s,
1H); Anal. (C.sub.33H.sub.32N.sub.2O.sub.5) C, H, N.
Preparation of Intermediate Cbz-L-(Tr-Glutaminol)
[0185] Lithium chloride (0.24 g, 5.66 mmol, 2.0 equiv.) was added
to a solution of Cbz-L-(Tr-Gln)-OMe (1.50 g, 2.79 mmol, 1 equiv.)
in 2:1 THF:EtOH (30 mL), and the mixture was stirred at 23.degree.
C. until all solids had dissolved (10 minutes). Sodium borohydride
(0.21 g, 5.55 mmol, 2.0 equiv.) was added, and the reaction mixture
was stirred overnight at 23.degree. C. The solvents were removed
under reduced pressure, the residue was taken up in water (50 mL),
and the pH was adjusted to 2-3 with 10% HCl. The product was
extracted with EtOAc (50 mL), and the organic layer was washed with
water (50 mL) and brine (50 mL) before drying over MgSO.sub.4. The
organic layer was concentrated, and the residue was purified by
flash column chromatography (gradient elution, 20-50% EtOAc in
benzene) to give Cbz-L-(Tr-glutaminol) (1.02 g, 72% yield) as a
white glassy solid: mp=66-70.degree. C.; IR (cm.sup.-1) 3318, 1699,
1510, 1240; .sup.1H NMR (DMSO-d.sub.6) .delta. 1.40 (m, 1H), 1.72
(m, 1H), 2.26 (m, 2H), 3.17-3.50 (m, 3H), 4.64 (t, 1H, J=5.0), 5.00
(s, 2H), 7.00-7.40 (m, 20H), 6.96 (d, 1H, J=8.5), 8.54 (s, 1H);
Anal. (C.sub.32H.sub.32N.sub.2O.sub.4) C, H, N.
Preparation of Intermediaite L-(Tr-Glutaminol)
[0186] A suspension of Cbz-L-(Tr-glutaminol) (1.93 g, 3.79 mmol) in
CH.sub.3OH (25 mL) and Pd/C (10%, 0.19 g) was stirred under a
hydrogen atmosphere (balloon) for 4 hours, then was filtered
through a layer of Celite. The filtrate was concentrated under
reduced pressure to give L-(Tr-glutaminol) as a white amorphous
solid (1.38 g, 98% yield): mp=191-193.degree. C.; IR (cm.sup.-1)
3255 (br), 1642, 1527; .sup.1H NMR (DMSO-d.sub.6) .delta. 1.29 (m,
1H), 1.53 (m, 1H), 2.29 (m, 2H), 3.08 (m, 1H), 3.18 (m, 2H), 3.38
(s, br, 2H), 4.43 (s, br, 1H), 7.14-7.28 (m, 15H), 8.62 (s,
1H).
Preparation of Intermediate Cbz-L-Leu-L-Phe-L-(Tr-Glutaminol)
[0187] Carbonyldiimidazole (0.17 g, 1.05 mmol, 1.0 equiv.) was
added to a solution of Cbz-L-Leu-L-Phe-OH (0.41 g, 1.0 mmol, 0.95
equiv.) in THF (10 mL), and the reaction mixture was stirred at
23.degree. C. for 1 hour. L-(Tr-Glutaminol) (0.39 g, 1.05 mmol, 1
equiv.) was then added, and the resulting solution was stirred
overnight. The volatiles were removed under reduced pressure, and
the residue was purified by flash column chromatography (gradient
elution, 2-4% CH.sub.3OH in CHCl.sub.3) to give
Cbz-L-Leu-L-Phe-L-(Tr-glutaminol) (0.47 g, 62% yield) as a white
amorphous solid: mp=92-95.degree. C.; IR (cm.sup.-1) 3302, 1657,
1520, 1238; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.79 (t, 6H, J=7.0),
1.30 (m, 2H), 1.44 (m, 2H), 1.75 (m, 1H), 2.22 (m, 2H), 2.82 (m,
1H), 2.97 (m, 1H), 3.14 (m, 1H), 3.25 (m, 1H), 3.63 (m, 1H), 3.95
(m, 1H), 4.48 (m, 1H), 4.65 (t, 1H, J=5.0), 4.96 (d, 1H, J=13.0),
5.02 (d, 1H, J=13.0), 7.07-7.33 (m, 25H), 7.42 (d, 1H, J=8.0), 7.66
(d, 1H, J=8.5), 7.86 (d, 1H, J=8.0), 8.52 (s, 1H); Anal.
(C.sub.47H.sub.52N.sub.4O.sub.6.0.5H.sub.2O)C, H, N.
Preparation of Intermediate Cbz-L-Leu-L-Phe-L-(Tr-Gln)-H
[0188] o-Iodoxybenzoic acid (0.63 g, 2.25 mmol, 3.0 equiv.) was
added to a solution of Cbz-L-Leu-L-Phe-L-(Tr-glutaminol) (0.58 g,
0.75 mmol, 1 equiv.) in DMSO (7.5 mL) at 23.degree. C. After
stirring for 2 hours, the DMSO was removed under reduced pressure.
The residue was twice diluted with CH.sub.2Cl.sub.2, and the
solvent evaporated to remove any remaining DMSO. The residue was
diluted with EtOAc (30 mL) and filtered, and the filtrate was
washed with 5% Na.sub.2S.sub.2O.sub.3/5% NaHCO.sub.3 solution (30
mL), water (30 mL), and brine (30 mL), and then dried over
Na.sub.2SO.sub.4. The solvent was removed under reduced pressure to
give Cbz-L-Leu-L-Phe-L-(Tr-Gln)-H (0.53 g, 92% yield) as a white
glassy solid, which was used immediately without further
purification: .sup.1H NMR (DMSO-d.sub.6) .delta. 0.79 (m, 6H),
1.00-1.98 (m, 5H), 2.27 (m, 2H), 2.84 (m, 1H), 3.02 (m, 1H), 3.98
(m, 2H), 4.58 (m, 1H), 4.99 (s, 2H), 7.14-7.32 (m, 25H), 7.39 (d,
1H, J=8.1), 7.97 (d, 1H, J=8.5), 8.38 (d, 1H, J=8.0), 8.60 (s, 1H),
9.20 (s, 1H).
Preparation of Intermediate
5-{3'-Cbz-L-Leu-L-Phe-L-(Tr-Gln)}-E-Propene-Isoxsazole
[0189] A solution of KN((CH.sub.3).sub.3Si).sub.2 (0.95 mL of a 0.5
M solution in THF, 0.477 mmol, 1.0 equiv.) was added to a
suspension of isoxazol-5-ylmethyl-triphenylphosphonium bromide
(0.222 g, 0.525 mmol, 1.1 equiv) in THF (20 mL) at 0.degree. C.,
and the reaction was stirred at 0.degree. C. for 30 minutes. A
solution of Cbz-L-Leu-L-Phe-L-(Tr-Gln)-H (0.366 g, 0.477 mmol, 1
equiv.) in THF (10 mL) was added, and the reaction mixture was then
stirred overnight at 23.degree. C. The solvent was removed in
vacuo, and the residue was diluted with EtOAc (30 mL), washed with
water (30 mL), and then dried over MgSO.sub.4. The solvent was
removed under reduced pressure and the residue purified by flash
silica gel chromatography (gradient elution, 0.fwdarw.1% CH.sub.3OH
in CHCl.sub.3) to give
5-{3'-(Cbz-L-Leu-L-Phe-L-(Tr-Gln))-E-propene}-isoxazole (0.307 g,
70% yield) as an amorphous solid: IR (cm.sup.-1) 3423, 1678, 1568,
1265, 1043, 711; .sup.1H NMR (DMSO-d.sub.6) S 0.77-0.81 (m, 6H),
1.21-1.36 (m, 2H), 1.40-1.55 (m, 1H), 1.60-1.80 (m, 2H), 2.34-2.45
(m, 2H), 2.82-2.87 (m, 1H), 2.91-3.04 (m, 1H), 3.95-4.00 (m, 1H),
4.41-4.50 (m, 1H), 4.53-4.60 (m, 1H), 4.99 (q, 2H, J=6.0), 6.19 (d,
1H, J=15.0), 6.36 (dd, 1H, J=15.0, 6.0), 6.46 (s, 1H), 7.15-7.33
(m, 20H), 7.42 (d, 1H, J=9.0), 7.56-7.63 (m, 5H), 7.96 (d, 1H,
J=9.0), 8.08 (d, 1H, J=9.0), 8.51 (s, 1H), 8.58 (s, 1H); HRMS
calcd. for C.sub.51H.sub.53N.sub.5O.sub.6+Cs 964.3050 (M+Cs), found
964.3018.
Preparation of Product:
5-(3'-(Cbz-L-Leu-L-Phe-L-Gln)-E-Propene)-Isoxazole
[0190] Trifluoroacetic acid (1 mL) was added to a solution of
5-{3'-(Cbz-L-Leu-L-Phe-L-(Tr-Gln))-E-propene}-isoxazole (0.214 g,
0.257 mmol) in CH.sub.2Cl.sub.2 (10 mL), and the reaction mixture
was stirred at 23.degree. C. overnight. The solvent was removed in
vacuo and the residue purified by flash silica gel chromatography
(gradient elution, 0.fwdarw.1% CH.sub.3OH in CHCl.sub.3) to give
5-(3'-(Cbz-L-Leu-L-Phe-L-Gln)-E-propene)-isoxazole as a white solid
(0.054 g, 36% yield): .sup.1H NMR (DMSO-d.sub.6) .delta. 0.77-0.83
(m, 6H), 1.26-1.46 (m, 2H), 1.47-1.62 (m, 1H), 1.69-1.79 (m, 2H),
2.04-2.29 (m, 2H), 2.83-2.88 (m, 1H), 2.97-3.10 (m, 1H), 3.99-4.12
(m, 1H), 4.37-4.43 (m, 1H), 4.48-4.57 (m, 1H), 5.01 (q, 2H, J=6.0),
6.20 (d, 1H, J=15.0), 6.36 (dd, 1H, J=15.0, 6.0), 6.45 (d, 1H,
J=3.0), 6.75 (s, 1H), 7.14-7.29 (m, 6H), 7.31-7.40 (m, 5H), 7.45
(d, 1H, J=9.0), 8.04 (d, 1H, J=9.0), 8.07 (d, 1H, J=9.0), 8.51 (d,
1H, J=3.0); Anal. (C.sub.32H.sub.39N.sub.5O.sub.6) C, H, N.
Example 2
Preparation of Compound A-1:
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gin)-E-Propenoate
[0191] ##STR106##
Preparation of Intermediate Boc-L-(Tr-Gln)-N(OMe)Me
[0192] Isobutyl chloroformate (4.77 mL, 36.8 mmol, 1.0 equiv.) was
added to a solution of Boc-L-(Tr-Gln)-OH (18.7 g, 36.7 mmol, 1
equiv.) and 4-methylmorpholine (8.08 mL, 73.5 mmol, 2.0 equiv.) in
CH.sub.2Cl.sub.2 (250 mL) at 0.degree. C. The reaction mixture was
stirred at 0.degree. C. for 20 min. (minutes), then
N,O-dimethylhydroxylamine hydrochloride (3.60 g, 36.7 mmol, 1.0
equiv.) was added. The resulting solution was stirred at 0.degree.
C. for 20 min. and at 23.degree. C. for 2 hours (h), and then was
partitioned between water (150 mL) and CH.sub.2Cl.sub.2
(2.times.150 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4, and were concentrated. Purification of the
residue by flash column chromatography (gradient elution, 40-20%
hexanes in EtOAc) provided Boc-L-(Tr-Gln)-N(OMe)Me (16.1 g, 82%
yield) as a white foam: IR (cm.sup.-1) 3411, 3329, 3062, 1701,
1659; .sup.1H NMR (CDCl.sub.3) .delta. 1.42 (s, 9H), 1.63-1.77 (m,
1H), 2.06-2.17 (m, 1H), 2.29-2.43 (m, 2H), 3.17 (s, 3H), 3.64 (s,
3H), 4.73 (s, br, 1H), 5.38-5.41 (m, 1H), 7.20-7.31 (m, 15H); Anal.
(C.sub.31H.sub.37N.sub.3O.sub.5) C, H, N.
Preparation of Intermediate BOC-L-(Tr-Gln)-H
[0193] Diisobutylaluminum hydride (50.5 mL of a 1.5 M solution in
toluene, 75.8 mmol, 2.5 equiv.) was added to a solution of
Boc-L-(Tr-Gln)-N(OMe)Me (16.1 g, 30.3 mmol, 1 equiv.) in THF at
-78.degree. C., and the reaction mixture was stirred at -78.degree.
C. for 4 hours. Methanol (4 mL) and 1.0 M HCl (10 mL) were added
sequentially, and the mixture was warmed to 23.degree. C. The
resulting suspension was diluted with Et.sub.2O (150 mL), and was
washed with 1.0 M HCl (3.times.100 mL), half-saturated NaHCO.sub.3
(100 mL), and water (100 mL). The organic layer was dried over
MgSO.sub.4, filtered, and concentrated to give crude
Boc-L-(Tr-Gln)-H (13.8 g, 97% yield) as a white solid:
mp=114-116.degree. C.; IR (cm.sup.-1) 3313, 1697, 1494; .sup.1H NMR
(CDCl.sub.3) .delta. 1.44 (s, 9H), 1.65-1.75 (m, 1H), 2.17-2.23 (m,
1H), 2.31-2.54 (m, 2H), 4.11 (s, br, 1H), 5.38-5.40 (m, 1H), 7.11
(s, 1H), 7.16-7.36 (m, 15H), 9.45 (s, 1H).
Preparation of Intermediate
Ethyl-3-(BOC-L-(Tr-Gln))-E-Propenoate
[0194] Sodium bis(trimethylsilyl)amide (22.9 mL of a 1.0 M solution
in THF, 22.9 mmol, 1.0 equiv.) was added to a solution of triethyl
phosphonoacetate (5.59 g, 22.9 mmol, 1.0 equiv.) in THF (200 mL) at
-78.degree. C., and the resulting solution was stirred for 20
minutes at that temperature. Crude Boc-L-(Tr-Gln)-H (10.8 g, 22.9
mmol, 1 equiv.) in THF (50 mL) was added via cannula, and the
reaction mixture was stirred for 2 hours at -78.degree. C., warmed
to 0.degree. C. for 10 minutes, and then partitioned between 0.5 M
HCl (150 mL) and a 1:1 mixture of EtOAc and hexanes (2.times.150
mL). The combined organic layers were dried over Na.sub.2SO.sub.4
and were concentrated. Purification of the residue by flash column
chromatography (40% EtOAc in hexanes) provided
ethyl-3-[Boc-L-(Tr-Gln)]-E-propenoate (10.9 g, 88% yield) as a
white foam: IR (cm.sup.-1) 3321, 1710; .sup.1H NMR (CDCl.sub.3)
.delta. 1.27 (t, 3H, J=7.2), 1.42 (s, 9H), 1.70-1.78 (m, 1H),
1.80-1.96 (m, 1H), 2.35 (t, 2H, J=7.0), 4.18 (q, 2H, J=7.2), 4.29
(s, br, 1H), 4.82-4.84 (m, 1H), 5.88 (dd, 1H, J=15.7, 1.6), 6.79
(dd, 1H, J=15.7, 5.3), 6.92 (s, 1H), 7.19-7.34 (m, 15H); Anal.
(C.sub.33H.sub.38N.sub.2O.sub.5) C, H, N.
Preparation of Intermediate
Ethyl-3-(Boc-L-Phe-L-(Tr-Gln))-E-Propenoate
[0195] A solution of HCl in 1,4-dioxane (4.0 M, 15 mL), was added
to a solution of ethyl-3-(Boc-L-(Tr-Gln))-E-propenoate (3.26 g,
6.01 mmol, 1 equiv.) in the same solvent (15 mL) at 23.degree. C.
After 2 h, the volatiles were removed under reduced pressure to
afford ethyl-3-(H.sub.2N-L-(Tr-Gln))-E-propenoate-HCl. This
material was dissolved in CH.sub.2Cl.sub.2 (60 mL) and Boc-L-Phe-OH
(1.59 g, 6.01 mmol, 1.0 equiv.), HOBt (1.22 g, 9.02 mmol, 1.5
equiv.), 4-methylmorpholine (1.98 mL, 18.03 mmol, 3 equiv.), and
EDC (1.73 g, 9.02 mmol, 1.5 equiv.) were added sequentially. The
reaction mixture was stirred at 23.degree. C. overnight, and then
was partitioned between water (100 mL) and CH.sub.2Cl.sub.2
(2.times.100 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4, concentrated, and the residue was purified by
flash column chromatography (40% EtOAc in hexane) to afford
ethyl-3-(Boc-L-Phe-L-(Tr-Gln))-E-propenoate (3.55 g, 85%) as white
foam: IR (cm.sup.-1) 3306, 1706, 1661; .sup.1H NMR (CDCl.sub.3)
.delta. 1.29 (t, 3H, J=7.2), 1.38 (s, 9H), 1.65-1.76 (m, 1H),
1.87-1.99 (m, 1H), 2.25-2.27 (m, 2H), 2.94-3.01 (m, 2H), 4.14-4.26
(m, 3H), 4.48-4.53 (m, 1H), 4.95 (s, br, 1H), 5.64 (d, 1H, J=15.8),
6.29 (d, 1H, J=8.1), 6.64 (dd, 1H, J=15.8, 5.4), 6.80 (s, br, 1H),
7.14-7.32 (m, 20H); Anal. (C.sub.42H.sub.47N.sub.3O.sub.6) C, H,
N.
Preparation of Intermediate
Ethyl-3-(BOC-L-Leu-L-Phe-L-(Tr-Gln))-E-Propenoate
[0196] A solution of HCl in 1,4-dioxane (4.0 M, 15 mL) was added to
a solution of ethyl-3-(Boc-L-Phe-L-(Tr-Gln))-E-propenoate (6.40 g,
9.28 mmol, 1 equiv.) in the same solvent (15 mL) at 23.degree. C.
After 2 hours, the volatiles were removed under reduced pressure.
The residue was dissolved in CH.sub.2Cl.sub.2 (100 mL), and
Boc-L-Leu-OH (2.58 g, 11.1 mmol, 1.2 equiv.), HOBt (1.88 g, 13.9
mmol, 1.5 equiv.), 4-methylmorpholine (3.06 mL, 27.8 mmol, 3
equiv.), and EDC (2.67 g, 13.92 mmol, 1.5 equiv.) were added
sequentially. The reaction mixture was stirred at 23.degree. C.
overnight, and then was partitioned between water (100 mL) and
CH.sub.2Cl.sub.2 (2.times.100 mL). The combined organic layers were
dried over Na.sub.2SO.sub.4 and concentrated, and the residue was
purified by flash column chromatography (2% CH.sub.3OH in
CH.sub.2Cl.sub.2) to afford
ethyl-3-(Boc-L-Leu-L-Phe-L-(Tr-Gln))-E-propenoate (6.46 g, 87%
yield) as white foam: IR (cm.sup.-1) 3284, 1651, 1515; .sup.1H NMR
(CDCl.sub.3) .delta. 0.86 (d, 3H, J=6.0), 0.89 (d, 3H, J=6.0), 1.29
(t, 3H, J=7.2), 1.34 (s, 9H), 1.38-1.60 (m, 3H), 1.62-1.89 (m, 1H),
1.95-1.97 (m, 1H), 2.28-2.30 (m, 2H), 3.06-3.08 (m, 2H), 3.92-3.94
(m, 1H), 4.17 (q, 2H, J=7.2), 4.48-4.51 (m, 2H), 4.67 (m, 1H), 5.66
(d, 1H, J=15.9), 6.51-6.57 (m, 2H), 6.69 (dd, 1H, J=15.6, 5.1),
7.10-7.33 (m, 21H); Anal.
(C.sub.48H.sub.58N.sub.4O.sub.7.0.33H.sub.2O)C, H, N.
Preparation of Intermediate
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln))-E-Prope-
noate
[0197] A solution of HCl in 1,4-dioxane (4.0 M, 3 mL), was added to
a solution of ethyl-3-(Boc-L-Leu-L-Phe-L-(Tr-Gln))-E-propenoate
(0.216 g, 0.27 mmol, 1 equiv.) in the same solvent (3 mL) at
23.degree. C. After 2 hours, the volatiles were removed under
reduced pressure. The residue was dissolved in CH.sub.2Cl.sub.2 (15
mL), cooled to 0.degree. C., and triethylamine (0.112 mL, 0.81
mmol, 3.0 equiv.) and 5-methylisoxazole-3-carbonyl chloride (0.058
g, 0.40 mmol, 1.5 equiv.) were added sequentially. The reaction
mixture was stirred at 0.degree. C. for 30 minutes, and then was
partitioned between water (50 mL) and CH.sub.2Cl.sub.2 (2.times.50
mL). The combined organic layers were dried over Na.sub.2SO.sub.4
and concentrated, and the residue was purified by flash column
chromatography (2% CH.sub.3OH in CH.sub.2Cl.sub.2) to afford
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln))-E-prope-
noate (0.199 g, 91% yield) as a white foam: IR (cm.sup.-1) 3286,
1650, 1541; .sup.1H NMR (CDCl.sub.3) .delta. 0.86 (d, 3H, J=5.4),
0.89 (d, 3H, J=5.7), 1.28 (t, 3H, J=7.2), 1.43-1.59 (m, 2H),
1.67-1.75 (m, 1H), 1.95-1.99 (m, 2H), 2.28 (t, 2H, J=7.2), 2.41 (s,
3H), 2.97-3.04 (m, 1H), 3.06-3.13 (m, 1H), 4.17 (q, 2H, J=7.2),
4.31-4.33 (m, 1H), 4.48-4.52 (m, 2H), 5.72 (d, 1H, J=15.9), 6.19
(s, 1H), 6.41 (d, 1H, J=7.5), 6.59 (d, 1H, J=8.1), 6.71 (dd, 1H,
J=15.3, 6.0), 6.95 (d, 1H, J=6.6), 7.09-7.21 (m, 21H); Anal.
(C.sub.48H.sub.53N.sub.5O.sub.7.H.sub.2O)C, H, N.
Preparation of Product
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln)-E-Propenoate
[0198] Triisopropylsilane (0.077 mL, 0.376 mmol, 1.8 equiv.) and
trifluoroacetic acid (3 mL) were added sequentially to a solution
of
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gin))-E-prope-
noate (0.185 g, 0.21 mmol, 1 equiv.) in CH.sub.2Cl.sub.2 (3 mL) at
23.degree. C., producing a bright yellow solution. The reaction
mixture was stirred for 30 minutes at 23.degree. C., during which
time it became colorless. The volatiles were removed under reduced
pressure, and the resulting white solid was triturated with
Et.sub.2O (10 mL), filtered, and air-dried to give
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln)-E-propenoate
(0.87 g, 81% yield) as white solid: mp=223-225.degree. C.; IR
(cm.sup.-1) 3298, 1662, 1544, 1457, 1278; .sup.1H NMR
(DMSO-d.sub.6) .delta. 0.81 (d, 3H, J=6.0), 0.85 (d, 3H, J=6.3),
1.23 (t, 3H, J=6.9), 1.38-1.42 (m, 1H), 1.48-1.77 (m, 4H), 2.04 (t,
2H, J=7.2), 2.46 (s, 3H), 2.78-2.86 (m, 1H), 2.93-3.00 (m, 1H),
4.11 (q, 2H, J=7.2), 4.36-4.54 (m, 3H), 5.63 (d, 1H, J=15.6), 6.56
(s, 1H), 6.68 (dd, 1H, J=15.9, 5.4), 6.76 (s, br, 1H), 7.19 (m,
6H), 8.09 (d, 1H, J=8.1), 8.14 (d, 1H, J=7.8), 8.58 (d, 1H, J=7.5);
Anal. (C.sub.29H.sub.39N.sub.5O.sub.7) C, H, N.
Example 3
Preparation of Compound A-2:
Ethyl-3-((Isoxazole-5'-carbonyl)-L-Leu-L-Phe-L-Gln)-E-Propenoate
[0199] ##STR107##
Preparation of Intermediate
Ethyl-3-((Isoxazole-5'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln))-E-Propenoate
[0200] This compound was prepared from
ethyl-3-(Boc-L-Leu-L-Phe-L-(Tr-Gln))-E-propenoate and
isoxazole-5-carbonyl chloride using the procedure described above
(Example 2) for the preparation of
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln))-E-prope-
noate: IR (cm.sup.-1) 3282, 1643, 1530; .sup.1H NMR (CDCl.sub.3)
.delta. 0.87 (t, 6H, J=6.6), 1.29 (t, 3H, J=7.2), 1.49-1.64 (m,
3H), 1.69-1.80 (m, 1H), 1.90-1.96 (m, 1H), 2.30 (t, 2H, J=7.2),
2.92-2.96 (m, 1H), 3.02-3.09 (m, 1H), 4.17 (q, 2H, J=7.2),
4.42-4.48 (m, 3H), 5.69 (d, 1H, J=15.3), 6.65 (s, br, 1H), 6.66
(dd, 1H, J=15.9, 5.4), 6.76-6.79 (m, 2H), 7.00-7.31 (m, 22H), 8.24
(s, 1H); Anal. (C.sub.47H.sub.51N.sub.5O.sub.7.0.75H.sub.2O)C, H,
N.
Preparation of
Ethyl-3-((Isoxazole-5'-carbonyl)-L-Leu-L-Phe-L-Gln)-E-Propenoate
[0201] The title compound was prepared from
ethyl-3-((isoxazole-5'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln))-E-propenoate
using a procedure analogous to that described above (Example 2) for
the preparation of
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln)-E-propenoate-
: mp=217-220.degree. C.; IR (cm.sup.-1) 3302, 1655, 1541; .sup.1H
NMR (DMSO-d.sub.6) .delta. 0.81 (d, 3H, J=6.0), 0.86 (d, 3H,
J=6.0), 1.21 (t, 3H, J=7.2), 1.42-1.75 (m, 5H), 2.04 (t, 2H,
J=7.2), 2.78-2.87 (m, 1H), 2.94-3.01 (m, 1H), 4.11 (q, 2H, J=7.2),
4.37 (m, 1H), 4.41-4.52 (m, 2H), 5.64 (d, 1H, J=15.6), 6.68 (dd,
1H, J=15.9, 5.4), 6.76 (s, br, 1H), 7.12-7.19 (m, 7H), 8.02 (d, 1H,
J=8.1), 8.20 (d, 1H, J=8.1), 8.74 (d, 1H, J=1.8), 8.94 (d, 1H,
J=7.8); Anal. (C.sub.28H.sub.37N.sub.5O.sub.7) C, H, N.
Example 4
Preparation of Compound A-3:
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-Gln)-E-Propenoate
[0202] ##STR108##
Preparation of Intermediate
Ethyl-3-(Boc-L-(4-Me-Phe)-L-(Tr-Gln))-E-Propenoate
[0203] Ethyl-3-(H.sub.2N-L-(Tr-Gln))-E-propenoate-HCl (prepared as
described in Example 2 above, 1.37 g, 3.10 mmol) was dissolved in
DMF (10 mL) at 23.degree. C. Diisopropylethylamine (1.08 mL, 6.20
mmol) was added, followed by Boc-L-(4-Me-Phe)-OH (0.87 g 3.10
mmol). The reaction was cooled to 0.degree. C. HATU (1.18 g, 3.10
mmol) was added, and the reaction allowed to warm to room
temperature. The DMF was removed in vacuo. The residue was
dissolved in EtOAc (30 mL), and the organic phase was washed
sequentially with 10% HCl solution (25 mL), saturated NaHCO.sub.3
solution (25 mL), H.sub.2O (25 mL), and brine (25 mL). The solvent
was dried (MgSO.sub.4) and filtered, and the residue purified by
flash column chromatography (gradient elution, 0.fwdarw.0.75%
CH.sub.3OH in CHCl.sub.3) to give
ethyl-3-(Boc-L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoate (1.48 g, 68%
yield) as a white amorphous solid: IR (cm.sup.-1) 1713, 1655, 1491,
1175; .sup.1H NMR (DMSO-d.sub.6) .delta. 1.20 (t, 3H, J=7.0), 1.30
(s, 9H), 1.62-1.66 (m, 2H), 2.23 (s, 3H), 2.32 (m, 2H), 2.72 (m,
1H), 2.84 (m, 1H), 4.07-4.09 (m, 1H), 4.10 (q, 2H, J=7.0), 4.38 (m,
1H), 5.64 (d, 1H, J=15.5), 6.72 (dd, 1H, J=15.5, 5.5), 6.88 (d, 1H,
J=8.0), 7.04 (d, 2H, J=7.7), 7.10 (d, 2H, J=7.7), 7.14-7.28 (m,
15H), 8.02 (d, 1H, J=8.0), 8.53 (s, 1H); Anal.
(C.sub.43H.sub.49N.sub.3O.sub.6) C, H, N.
Preparation of Intermediate
Ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-Phe)-L-(Tr-Gln))-E-Propenoat-
e
[0204] Ethyl-3-(Boc-L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoate (1.45 g,
2.06 mmol) was dissolved in 1,4-dioxane (27 mL), and a solution of
HCl in 1,4-dioxane (4.0 M, 14 mL) was added. The reaction was
stirred at room temperature for 4 hours. The solvent was removed by
evaporation, and the residue taken up in EtOAc (50 mL). The organic
phase was washed with saturated NaHCO.sub.3 solution (50 mL) and
then brine (50 mL), dried (MgSO.sub.4), and the solvent removed
under reduced pressure to give 1.23 g of an off-white amorphous
solid. This material was coupled with
Boc-L-.alpha.-(t-Butyl-Gly)-OH using the procedure described for
the synthesis of ethyl-3-(Boc-L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoate
above to afford
ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-Phe)-L-(Tr-Gln))-E-pr-
openoate (49% yield) as a white amorphous solid: IR (cm.sup.-1)
1655, 1507, 1248, 1171; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.81 (s,
9H), 1.21 (t, 3H, J=7.0), 1.37 (s, 9H), 1.52-1.70 (m, 2H), 2.22 (s,
3H), 2.26-2.28 (m, 2H), 2.73-2.91 (m, 2H), 3.86 (d, 1H, J=9.6),
4.05-4.14 (m, 2H), 4.31-4.36 (m, 1H), 4.47-4.55 (m, 1H), 5.54 (d,
1H, J=15.4), 6.37 (d, 1H, J=9.6), 6.65 (dd, 1H, J=15.8, 5.5), 7.01
(d, 2H, J=8.1), 7.07 (d, 2H, J=7.7), 7.11-7.32 (m, 15H), 8.03 (d,
1H, J=8.1), 8.10 (d, 1H, J=7.7), 8.49 (s, 1H); Anal.
(C.sub.49H.sub.60N.sub.4O.sub.7.0.4H.sub.2O)C, H, N.
Preparation of Intermediate
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-(Tr-Gln))-E-Propenoate
[0205]
Ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-Phe)-L-(Tr-Gln))-E-pr-
openoate was deprotected using the procedure described for the
deprotection of ethyl-3-(Boc-L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoate
above, and the resulting amine (0.22 g, 0.30 mmol) was dissolved in
CH.sub.2Cl.sub.2 (3 mL). Pyridine (0.025 mL, 0.32 mmol) was added,
and the reaction was cooled to 0.degree. C.
5-Methylisoxazole-3-carbonyl chloride (0.046 g, 0.32 mmol) was
added. The reaction was allowed to warm to room temperature, and
was stirred for one hour. The solvent was removed in vacuo, and the
residue subjected to flash column chromatography (gradient elution,
0.fwdarw.1% CH.sub.3OH in CH.sub.2Cl.sub.2) to afford
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-(Tr-Gln))-E-propenoate (0.19 g, 77% yield) as a white
amorphous solid: IR (cm.sup.-1) 1651, 1518; .sup.1H NMR
(DMSO-d.sub.6) .delta. 0.88 (s, 9H), 1.20 (t, 3H, J=7.0), 1.55-1.67
(m, 2H), 2.14 (s, 3H), 2.18-2.28 (m, 2H), 2.45 (s, 3H), 2.70-2.77
(m, 1H), 2.86-2.93 (m, 1H), 4.07-4.14 (m, 2H), 4.46 (d, 1H, J=9.6),
4.50-4.55 (m, 1H), 5.54 (d, 1H, J=15.8), 6.59 (s, 1H), 6.65 (dd,
1H, J=15.8, 5.5, 15.8), 6.95 (d, 2H, J=8.1), 7.05 (d, 2H, J=8.1),
7.13-7.28 (m, 15H), 7.60 (d, 1H, J=9.6), 8.13 (d, 1H, J=8.1), 8.41
(d, 1H, J=8.1), 8.51 (s, 1H); Anal.
(C.sub.49H.sub.55N.sub.5O.sub.7) C, H, N.
Preparation of Product
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-Gln)-E-Propenoate
[0206]
Ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)--
L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoate (0.17 g, 0.20 mmol) was
dissolved in CH.sub.2Cl.sub.2 (4 mL) at 23.degree. C.
Trifluoroacetic acid (0.4 mL) was added, and the reaction was
stirred at room temperature for six hours. The solvents were
removed in vacuo, and the residue was subjected to flash column
chromatography (gradient elution, 0.fwdarw.2% CH.sub.3OH in
CH.sub.2Cl.sub.2) to afford
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-Gln)-E-propenoate (0.085 g, 73% yield) as a white amorphous
solid: IR (cm.sup.-1) 1661, 1541, 1206; .sup.1H NMR (DMSO-d.sub.6)
.delta. 0.88 (s, 9H), 1.21 (t, 3H, J=7.0), 1.60-1.73 (m, 2H),
2.01-2.06 (m, 2H), 2.14 (s, 3H), 2.50 (s, 3H), 2.70-2.77 (m, 1H),
2.86-2.93 (m, 1H), 4.07-4.14 (m, 2H), 4.34-4.37 (m, 1H), 4.45 (d,
1H, J=9.6), 4.50-4.55 (m, 1H), 5.57 (d, 1H, J=15.8), 6.60 (s, 1H),
6.66 (dd, 1H, J=15.8, 5.5), 6.75 (s, br, 1H), 6.96 (d, 2H, J=8.1),
7.06 (d, 2H, J=7.7), 7.17 (s, br, 1H), 7.65 (d, 1H, J=9.6), 8.14
(d, 1H, J=8.1), 8.40 (d, 1H, J=7.7); Anal.
(C.sub.30H.sub.41N.sub.5O.sub.7.0.5TFA.0.5H.sub.2O)C, H, N.
Example 5
Preparation of Compound A-4:
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-Gln)-E-Provenoate
[0207] ##STR109##
Preparation of Intermediate
Ethyl-3-(Boc-L-(4-F-Phe)-L-(Tr-Gin))-E-Propenoate
[0208] Boc-L-(4-F-Phe)-OH (1.41 g, 5.0 mmol) was dissolved in THF
(50 mL). Ethyl-3-(H.sub.2N-L-(Tr-Gln))-E-propenoate-HCl (prepared
as described in Example 2 above, 1.0 g, 5.0 mmol) was added,
followed by Et.sub.3N (0.70 mL, 5.0 mmol). Carbonyldiimidazole
(0.81 g, 5.0 mmol) was added, and the reaction was stirred at room
temperature for 20 hours. The solvent was removed in vacuo, and the
residue subjected to flash column chromatography (gradient elution,
0.fwdarw.1% CH.sub.3OH in CH.sub.2Cl.sub.2) to afford
ethyl-3-(Boc-L-(4-F-Phe)-L-(Tr-Gln))-E-propenoate (1.13 g, 32%
yield) as a white amorphous solid: IR (cm.sup.-1) 1712, 1666, 1510,
1169; .sup.1H NMR (DMSO-d.sub.6) .delta. 1.20 (t, 3H, J=7.0), 1.29
(s, 9H), 1.61-1.70 (m, 2H), 2.27-2.34 (m, 2H), 2.74-2.78 (m, 1H),
2.86-2.90 (m, 1H), 4.06-4.13 (m, 3H), 4.36-4.40 (m, 1H), 5.58 (d,
1H, J=15.6), 6.71 (dd, 1H, J=15.6, 5.5), 6.98 (d, 1H, J=8.1),
7.03-7.09 (m, 2H), 7.14-7.28 (m, 17H), 8.06 (d, 1H, J=8.1), 8.53
(s, 1H); LRMS (M+Na) 730.
Preparation of Intermediate
Ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-F-Phe)-L-(Tr-Gln))-E-Propenoate
[0209] Ethyl-3-(Boc-L-(4-F-Phe)-L-(Tr-Gln))-E-propenoate was
deprotected and coupled with Boc-L-.alpha.-(t-Butyl-Gly)-OH using
the procedures described above to prepare
ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoat-
e, to provide
ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-F-Phe)-L-(Tr-Gln))-E-propenoate
(54% yield) as a white amorphous solid: IR (cm.sup.-1) 1720, 1651,
1506, 1168; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.80 (s, 9H), 1.20
(t, 3H, J=7.0), 1.36 (s, 9H), 1.53-1.67 (m, 2H), 2.23-2.28 (m, 2H),
2.79-2.94 (m, 2H), 3.85 (d, 1H, J=9.9), 4.09 (q, 2H, J=7.0),
4.31-4.35 (m, 1H), 4.53-4.55 (m, 1H), 5.46 (d, 1H, J=15.8), 6.36
(d, 1H, J=9.2), 6.64 (dd, 1H, J=15.8, 5.5), 6.97-7.03 (m, 2H),
7.13-7.28 (m, 17H), 8.08 (d, 1H, J=8.1), 8.14 (d, 1H, J=8.1), 8.49
(s, 1H); Anal. (C.sub.48H.sub.57N.sub.4O.sub.7F) C, H, N.
Preparation of Intermediate
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-(Tr-Gln))-E-Propenoate
[0210]
Ethyl-3-(Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-F-Phe)-L-(Tr-Gln))-E-pro-
penoate was deprotected and coupled with
5-methylisoxazole-3-carbonyl chloride using the procedures
described above to prepare
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-(Tr-Gln))-E-propenoate to afford
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-(Tr-Gln))-E-propenoate (74% yield) as a white amorphous
solid: IR (cm.sup.-1) 1659, 1535, 1510; .sup.1H NMR (DMSO-d.sub.6)
.delta. 0.88 (s, 9H), 1.20 (t, 3H, J=7.4), 1.52-1.67 (m, 2H),
2.23-2.28 (m, 2H), 2.45 (s, 3H), 2.75-2.82 (m, 1H), 2.89-2.96 (m,
1H), 4.09 (q, 2H, J=7.0), 4.32-4.36 (m, 1H), 4.45 (d, 1H, J=9.6),
4.50-4.55 (m, 1H), 5.44 (d, 1H, J=15.6), 6.58 (s, 1H), 6.63 (dd,
1H, J=15.6, 5.5, 15.6), 6.93-6.99 (m, 2H), 7.13-7.28 (m, 17H), 7.64
(d, 1H, J=9.6), 8.16 (d, 1H, J=8.5), 8.46 (d, 1H, J=8.1), 8.51 (s,
1H); Anal. (C.sub.48H.sub.52N.sub.5O.sub.7F) C, H, N.
Preparation of
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-Gln)-E-Propenoate
[0211]
Ethyl-3-[(5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)--
L-(4-F-Phe)-L-(Tr-Gln)]-E-propenoate was deprotected using a
procedure analogous to that described above for the preparation of
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-Me-
-Phe)-L-Gln)-E-propenoate to provide
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-Gln)-E-propenoate (80% yield) as a white amorphous solid: IR
(cm.sup.-1) 1653, 1543, 1223; .sup.1H NMR (DMSO-d.sub.6) .delta.
0.88 (s, 9H), 1.21 (t, 3H, J=7.0), 1.59-1.75 (m, 2H), 2.01-2.06 (m,
2H), 2.46 (s, 3H), 2.75-2.82 (m, 1H), 2.89-2.96 (m, 1H), 4.09 (q,
2H, J=7.0), 4.33-4.36 (m, 1H), 4.44 (d, 1H, J=9.6), 4.50-4.58 (m,
1H), 5.47 (d, 1H, J=15.8), 6.59 (s, 1H), 6.64 (dd, 1H, J=15.8,
5.5), 6.75 (s, br, 1H), 6.94-7.00 (m, 2H), 7.16 (s, br, 1H),
7.18-7.23 (m, 2H), 7.69 (d, 1H, J=9.6), 8.16 (d, 1H, J=8.1), 8.44
(d, 1H, J=8.1); Anal. (C.sub.29H.sub.38N.sub.5O.sub.7F) C, H,
N.
Example 6
Preparation of Compound A-5:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Pyrrol-
-Ala)}-E-Propenoate
[0212] ##STR110##
Preparation of Intermediate
(4S)-4-(2'-Carboxyethyl)-2,2-dimethyloxazolidine-3-carboxylic Acid
tert-Butyl Ester
[0213] Sodium hydroxide (27 mL of a 4.0 M solution in H.sub.2O, 108
mmol, 3.0 equiv.) was added to a solution of
(4S)-4-(2-methoxycarbonylethyl)-2,2-dimethyl-oxazolidine-3-carboxylic
acid tert-butyl ester (prepared as described in Chida et al., J.
Chem. Soc., Chem. Commun. 1992, 1064) (10.5 g, 36.5 mmol, 1 equiv.)
in CH.sub.3OH (150 mL), and the resulting cloudy reaction mixture
was stirred at 23.degree. C. for 3.5 h. The mixture was
concentrated under reduced pressure to .about.30 mL volume, and
then was partitioned between 0.5 M HCl (150 mL) and EtOAc
(2.times.150 mL). The combined organic layers were dried over
MgSO.sub.4 and were gravity filtered. The filtrate was concentrated
under reduced pressure and the residue dried under vacuum, to
afford
(4S)-4-(2'-carboxyethyl)-2,2-dimethyloxazolidine-3-carboxylic acid
tert-butyl ester (10.0 g, 100% crude yield). This material was used
without further purification: .sup.1H NMR (CDCl.sub.3, mixture of
rotamers) .delta. 1.49 (s), 1.57 (s), 1.60 (s), 1.84-2.05 (m),
2.39-2.41 (m), 3.71-3.74 (m), 3.91-4.05 (m).
Preparation of Intermediate
(4S,4''S)-4-{3'-(4''-Benzyl-2''-oxo-oxazolidin-3''-yl)-3'-oxopropyl}-2,2--
dimethyloxazolidine-3-carboxylic Acid tert-Butyl Ester
[0214] Triethylamine (8.87 mL, 63.6 mmol, 3.0 equiv.) and pivaloyl
chloride (2.61 mL, 21.2 mmol, 1.0 equiv.) were added sequentially
to a solution of
(4S)-4-(2'-carboxyethyl)-2,2-dimethyloxazolidine-3-carboxylic acid
tert-butyl ester (5.80 g, 21.2 mmol, 1 equiv.) in THF (450 mL) at
0.degree. C. The cloudy reaction mixture was stirred at 0.degree.
C. for 3.5 h, then lithium chloride (0.988 g, 23.3 mmol, 1.1
equiv.) and (S)-(-)-4-benzyl-2-oxazolidinone (3.57 g, 20.1 mmol,
0.95 equiv.) were added sequentially. After warming to 23.degree.
C. and stirring for 19 h, the reaction mixture was partitioned
between 0.5 M HCl (150 mL) and EtOAc (2.times.150 mL). The combined
organic layers were washed with half-saturated Na.sub.2CO.sub.3
(150 mL), dried over MgSO.sub.4, and gravity filtered. The filtrate
was concentrated under reduced pressure and the residue was
purified by flash column chromatography (30% EtOAc in hexanes) to
give
(4S,4"S)-4-{3'-(4''-benzyl-2''-oxo-oxazolidin-3''-yl)-3'-oxopropyl}-2,2-d-
imethyloxazolidine-3-carboxylic acid tert-butyl ester (7.17 g, 83%)
as a colorless oil: IR (cm.sup.-1) 2978, 1783, 1694; .sup.1H NMR
(CDCl.sub.3, mixture of rotamers) .delta. 1.49 (s), 1.59 (s), 1.63
(s), 2.01-2.10 (m), 2.76 (dd, J=13.5, 9.8), 2.82-3.13 (m),
3.30-3.41 (m), 3.76-3.82 (m), 3.90 (s, br), 3.97 (dd, J=9.0, 5.6),
4.10-4.19 (m), 4.63-4.71 (m), 7.22-7.36 (m); Anal.
(C.sub.23H.sub.32N.sub.2O.sub.6) C, H, N. Preparation of
Intermediate
(2'S,4S,4''S)-4-{2'-(4''-Benzyl-2''-oxo-oxazolidine-3''-carbonyl)-pent-4'-
-enyl}-2,2-dimethyloxazolidine-3-carboxylic Acid tert-Butyl
Ester
[0215] A solution of
(4S,4''S)-4-{3'-(4''-benzyl-2''-oxo-oxazolidin-3''-yl)-3'-oxopropyl}-2,2--
dimethyloxazolidine-3-carboxylic acid tert-butyl ester (7.17 g,
16.6 mmol, 1 equiv.) in THF (50 mL) was added to a solution of
sodium bis(trimethylsilyl)amide (16.6 mL of a 1.0 M solution in
THF, 16.6 mmol, 1.0 equiv.) in the same solvent (150 mL) at
-78.degree. C. The reaction mixture was stirred for 20 min. at
-78.degree. C., and then allyl iodide (4.55 mL, 49.8 mmol, 3.0
equiv.) was added. After stirring an additional 3 h at -78.degree.
C., the reaction mixture was maintained at -45.degree. C. for 2 h,
and then was partitioned between a 2:1 mixture of half-saturated
NH.sub.4Cl and 5% Na.sub.2S.sub.2O.sub.3 (300 mL) and a 1:1 mixture
of EtOAc and hexanes (2.times.200 mL). The combined organic layers
were washed with H.sub.2O (200 mL), dried over MgSO.sub.4, and
gravity filtered. The filtrate was concentrated under reduced
pressure and the residue was purified by flash column
chromatography (15% EtOAc in hexanes) to provide
(2'S,4S,4''S)-4-{2'-(4''-benzyl-2''
''-oxo-oxazolidine-3''-carbonyl)-pent-4'-enyl}-2,2-dimethyloxazolidine-3--
carboxylic acid tert-butyl ester (4.29 g, 55%) as a colorless oil:
IR (cm.sup.-1) 2978, 1780, 1695; .sup.1H NMR (CDCl.sub.3, mixture
of rotamers) .delta. 1.45 (s), 1.49 (s), 1.68-1.80 (m), 2.13-2.47
(m), 2.49-2.67 (m), 3.32 (dd, J=13.4, 3.1), 3.69-3.97 (m),
4.11-4.21 (m), 4.66-4.74 (m), 5.06-5.13 (m), 5.74-5.88 (m),
7.20-7.36 (m); Anal. (C.sub.26H.sub.36N.sub.2O.sub.6) C, H, N.
Preparation of Intermediate
(1S,3S)-{3-(1'-(2'',4''-Dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydro-
xymethylpropyl}-carbamic Acid tert-Butyl Ester
[0216] Ozone was bubbled through a solution of
(2'S,4S,4''S)-4-(2'-(4''-benzyl-2''-oxo-oxazolidine-3''-carbonyl)-pent-4'-
-enyl)-2,2-dimethyloxazolidine-3-carboxylic acid tert-butyl ester
(4.29 g, 9.08 mmol, 1 equiv.) in CH.sub.2Cl.sub.2 (200 .mu.L) and
CH.sub.3OH (0.735 mL, 18.1 mmol, 2.0 equiv.) at -78.degree. C.
until a blue color persisted. The reaction mixture was then purged
with argon until it became colorless. Methyl sulfide (6.67 mL, 90.8
mmol, 10 equiv.) was added, the mixture was stirred at -78.degree.
C. for 3.5 h, and was maintained at 0.degree. C. for an additional
1 h. After partitioning the reaction mixture between H.sub.2O (200
mL) and a 1:1 mixture of EtOAc and hexanes (2.times.200 mL), the
combined organic layers were dried over MgSO.sub.4 and gravity
filtered. The filtrate was concentrated under reduced pressure and
the residue was immediately utilized without further
purification.
[0217] The above residue was dissolved in a 2:1 mixture of THF and
EtOH (240 mL) at 23.degree. C., and 2,4-dimethoxybenzylamine
hydrochloride (7.40 g, 36.3 mmol, 4.0 equiv.), sodium acetate (2.98
g, 36.2 mmol, 4.0 equiv.), and sodium cyanoborohydride (1.14 g,
18.1 mmol, 2.0 equiv.) were added sequentially. The resulting
suspension was stirred for 18 h at 23.degree. C., and then was
partitioned between 0.5 M HCl (400 mL) and EtOAc (2.times.200 mL).
The combined organic layers were washed with half-saturated
NaRCO.sub.3 (300 mL), dried over Na.sub.2SO.sub.4, and concentrated
under reduced pressure. The residue was passed through a short
silica gel column (eluting with 50% EtOAc in hexanes) to give
(3'S,4S)-4-{1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-ylmethyl}-2-
,2-dimethyl-oxazolidine-3-carboxylic acid tert-butyl ester
contaminated with (S)-(-)-4-benzyl-2-oxazolidinone.
[0218] This material was dissolved in CH.sub.3OH (100 mL), and
TsOH--H.sub.2O (0.345 g, 1.81 mmol, 0.20 equiv.) was added. The
reaction mixture was heated to 50.degree. C., and was maintained at
that temperature for 2.5 h. After cooling to 23.degree. C., the
reaction mixture was concentrated under reduced pressure to
.about.20 mL volume and was partitioned between half-saturated
NaHCO.sub.3 (150 mL) and a 9:1 mixture of CH.sub.2Cl.sub.2 and
CH.sub.3OH (2.times.150 mL). The combined organic layers were dried
over Na.sub.2SO.sub.4 and concentrated under reduced pressure.
Purification of the residue by flash column chromatography (3%
CH.sub.3OH in CH.sub.2Cl.sub.2) afforded
(1S,3'S)-{2-(1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydr-
oxymethylethyl}-carbamic acid tert-butyl ester (1.62 g, 44%) as a
foam: IR (cm.sup.-1) 3328, 1669; .sup.1H NMR (CDCl.sub.3) .delta.
1.44 (s, 9H), 1.50-1.75 (m, 2H), 1.90-2.00 (m, 1H), 2.17-2.27 (m,
1H), 2.52-2.62 (m, 1H), 3.14-3.24 (m, 2H), 3.51-3.65 (m, 3H),
3.70-3.78 (m, 1H), 3.80 (s, 6H), 4.35 (d, 1H, J=14.3), 4.48 (d, 1H,
J=14.3), 5.51-5.54 (m, 1H), 6.42-6.46 (m, 2H), 7.09-7.12 (m, 1H);
Anal. (C.sub.21H.sub.32N.sub.2O.sub.6) C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-((N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala])-E-Propenoate
[0219] DMSO (0.270 mL, 3.80 mmol, 3 equiv. was added dropwise to a
-78.degree. C. solution of oxalyl chloride (0.166 mL, 1.90 mmol,
1.5 equiv.) in CH.sub.2Cl.sub.2 (14 mL). The reaction mixture was
stirred 20 min., then a solution of
(1S,3'S)-{2-(1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydr-
oxymethylethyl}-carbamic acid tert-butyl ester (0.518 g, 1.27 mmol,
1 equiv.) in CH.sub.2Cl.sub.2 (13 mL) was added via cannula along
the side of the reaction vessel. After stirring 20 min.,
triethylamine (1.06 mL, 7.60 mmol, 6 equiv.) was added dropwise,
and the reaction mixture was stirred for 1.5 h. Acetic acid (0.479
mL, 8.37 mmol, 6.6 equiv.) was added, and the reaction mixture was
warmed to 0.degree. C. for 5 min., then diluted with MTBE (200 mL)
and washed with water, saturated NaHCO.sub.3, and brine (25 mL
each). The organic phase was dried over Na.sub.2SO.sub.4 and
concentrated to provide the crude aldehyde as a foam (0.516 g,
quant.), which was used without further purification.
[0220] Sodium bis(trimethylsilyl)amide (1.23 mL of a 1.0 M solution
in THF, 1.23 mmol, 1 equiv.) was added to a solution of triethyl
phosphonoacetate (0.244 mL, 1.23 mmol, 1 equiv.) in THF (15 mL) at
-78.degree. C., and the resulting solution was stirred for 20 min.
at that temperature. The crude aldehyde (prepared above, 0.500 g,
1.23 mmol, 1 equiv.) in THF (13 mL) was added via cannula along the
side of the reaction vessel, and the reaction mixture was stirred
for 45 min. at -78.degree. C., warmed to 0.degree. C. for 7 min.,
and partitioned between 0.5 M HCl (20 mL) and MTBE (2.times.50 mL).
The combined organic layers were dried over MgSO.sub.4 and were
concentrated. Purification of the residue by flash column
chromatography (60% EtOAc in hexanes) provided
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-pyrrol-Ala)}-E-prope-
noate (0.356 g, 61%) as a white foam: R.sub.f=0.43 (60% EtOAc in
hexanes); IR (cm.sup.-1) 3307, 1708, 1678; .sup.1H NMR (CDCl.sub.3)
.delta. 1.28 (t, 3H, J=7.2), 1.43 (s, 9H), 1.52-1.70 (m, 2H),
1.98-2.09 (m, 1H), 2.21-2.34 (m, 1H), 2.48-2.59 (m, 1H), 3.16-3.24
(m, 2H), 3.80 (s, 6H), 4.18 (q, 2H, J=7.2), 4.27-4.40 (m, 1H), 4.41
(s, 2H), 5.40 (d, 1H, J=8.1), 5.95 (dd, 1H, J=15.6, 1.6), 6.41-6.48
(m, 2H), 6.86 (dd, 1H, J=15.6, 5.3), 7.08-7.13 (m, 1H); Anal.
(C.sub.25H.sub.36N.sub.207-0.25H.sub.2O)C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-(4-F-Phe)-L-((N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-Pr-
openoate
[0221] This material was prepared from
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-pyrrol-Ala)}-E-propenoate
and Boc-L-(4-F-Phe)-OH using a procedure similar to that described
for the preparation of
ethyl-3-(Boc-L-(4-Me-Phe)-L-(Tr-Gln))-E-propenoate (Example 4)
above: R.sub.f=0.34 (60% EtOAc in hexanes); IR (cm.sup.-1) 3258,
1705, 1666; .sup.1H NMR (CDCl.sub.3) .delta. 1.28 (t, 3H, J=7.2),
1.45 (s, 9H), 1.51-1.66 (m, 2H), 1.78-1.90 (m, 1H), 2.06-2.23 (m,
2H), 2.99 (dd, 1H, J=13.7, 6.2), 3.11 (dd, 1H, J=13.7, 5.3),
3.17-3.23 (m, 2H), 3.80 (s, 3H), 3.81 (s, 3H), 4.18 (q, 2H, J=7.2),
4.35 (s, 2H), 4.38-4.51 (m, 2H), 5.29-5.37 (m, 1H), 5.76 (d, 1H,
J=15.8), 6.43-6.47 (m, 2H), 6.72 (dd, 1H, J=15.8, 5.3), 6.83-6.91
(m, 2H), 7.09-7.17 (m, 3H), 7.92 (br, 1H); Anal.
(C.sub.34H.sub.44FN.sub.3O.sub.8) C, H, N.
Preparation of Intermediate
Ethyl-3-{BOC-L-Val-L-(4-F-Phe)-L-[(N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala]-
}-E-Propenoate
[0222] This compound was prepared from
ethyl-3-{Boc-L-(4-F-Phe)-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-pr-
openoate and Boc-L-Val-OH using a similar procedure to that
described above for the preparation of
ethyl-3-(Boc-L-Phe-L-(Tr-Gln))-E-propenoate (Example 2):
R.sub.f=0.24 (60% EtOAc in hexanes); IR (cm.sup.-1) 3284, 1713,
1678 br, 1643; .sup.1H NMR (CDCl.sub.3) .delta. 0.91 (d, 3H,
J=6.8), 0.97 (d, 3H, J=6.8), 1.28 (t, 3H, J=7.2), 1.45 (s, 9H),
1.50-1.62 (m, 2H), 1.66-1.82 (m, 1H), 1.90-2.02m, 1H), 2.08-2.21
(m, 2H), 2.94 (dd, 1H, J=13.5, 5.8), 3.17-3.27 (m, 3H), 3.80 (s,
3H), 3.82 (s, 3H), 3.97-4.05 (m, 1H), 4.17 (q, 2H, J=7.2), 4.27 (d,
1H, J=14.3), 4.29-4.38 (m, 1H), 4.40 (d, 1H, J=14.3), 4.86-4.93 (m,
1H), 5.10 (d, 1H, J=8.7), 5.76 (dd, 1H, J=15.6, 1.2), 6.45-6.52 (m,
2H), 6.70 (dd, 1H, J=15.6, 5.4), 6.79-6.88 (m, 3H), 7.12-7.22 (m,
3H), 8.30 (d, 1H, J=5.9); Anal. (C.sub.39H.sub.53FN.sub.4O.sub.9)
C, H, N.
Preparation of Intermediate
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((N-2,4-Dim-
ethoxybenzyl)-(S)-Pyrrol-Ala)}-E-Propenoate
[0223] This compound was prepared from
ethyl-3-{Boc-L-Val-L-(4-F-Phe)-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)-
}-E-propenoate and isoxazole-5-carbonyl chloride using the
procedure described above (Example 2) for the preparation of
ethyl-3-((5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln))-E-prope-
noate: R.sub.f=0.36 (5% CH.sub.3OH in CH.sub.2Cl.sub.2); IR
(cm.sup.-1) 3284, 1717, 1650; .sup.1H NMR (CDCl.sub.3) .delta. 0.97
(d, 3H, J=6.8), 1.01 (d, 3H, J=6.8), 1.28 (t, 3H, J=7.2), 1.51-1.64
(m, 2H), 1.72-1.84 (m, 1H), 1.95-2.05 (m, 1H), 2.11-2.33 (m, 2H),
2.48 (s, 3H), 2.98 (dd, 1H, J=13.7, 5.6), 3.16-3.24 (m, 3H), 3.80
(s, 3H), 3.81 (s, 3H), 4.17 (q, 2H, J=7.2), 4.23 (d, 1H, J=14.3),
4.31-4.42 (m, 1H), 4.40 (d, 1H, J=14.3), 4.44-4.50 (m, 1H),
4.88-4.96 (m, 1H), 5.79 (dd, 1H, J=15.6, 1.4), 6.43-6.49 (m, 3H),
6.71 (dd, 1H, J=15.6, 5.3), 6.80-6.88 (m, 2H), 6.94 (d, 1H, J=9.3),
7.11-7.17 (m, 3H), 7.29 (d, 1H, J=8.7), 8.33 (d, 1H, J=6.2); Anal.
(C.sub.39H.sub.48FN.sub.5O.sub.9.0.5H.sub.2O)C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Pyrrol-
-Ala)}-E-Propenoate
[0224] A suspension of
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((N-2,4-dim-
ethoxybenzyl)-(S)-pyrrol-Ala)}-E-propenoate (0.263 g, 0.351 mmol, 1
equiv.), water (2 drops), and DDQ (0.104 g, 0.458 mmol, 1.3 equiv.)
was refluxed for 9 h and then allowed to cool to room temperature
over 8 h. The reaction mixture was diluted with CH.sub.2Cl.sub.2
(200 mL) and washed with a 2:1 mixture of saturated NaHCO.sub.3 and
1 N NaOH (20 mL). The organic phase was dried over MgSO.sub.4 and
evaporated. Purification of the residue by flash column
chromatography (gradient elution 2-3% CH.sub.3OH in
CH.sub.2Cl.sub.2) gave
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-pyrrol-
-Ala)}-E-propenoate (0.117 g, 56%) as a white solid:
mp=219-220.degree. C.; R.sub.f=0.23 (5% CH.sub.3OH in
CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3401 br, 3295, 1655 br; I H NMR
(CDCl.sub.3) .delta. 0.94 (d, 3H, J=6.8), 0.97 (d, 3H, J=6.5), 1.29
(t, 3H, J=7.2), 1.54-1.65 (m, 1H), 1.72-1.91 (m, 2H), 2.07-2.26 (m,
2H), 2.28-2.39 (m, 1H), 2.49 (d, 3H, J=0.9), 3.01 (dd, 1H, J=13.8,
6.1), 3.12 (dd, 1H, J=13.8, 6.4), 3.26-3.38 (m, 2H), 4.18 (q, 2H,
J=7.2), 4.34 (dd, 1H, J=8.7, 7.2), 4.43-4.54 (m, 1H), 4.90 (dt, 1H,
J=9.0, 6.2), 5.76 (dd, 1H, J=15.6, 1.6), 6.00 (s, 1H), 6.42 (q, 1H,
J=0.9), 6.72 (dd, 1H, J=15.6, 5.4), 6.86-6.94 (m, 2H), 7.01 (d, 1H,
J=9.0), 7.11-7.18 (m, 2H), 7.21 (d, 1H, J=8.7), 7.76 (d, 1H,
J=7.2); Anal. (C.sub.30H.sub.38FN.sub.5O.sub.7) C, H, N.
Example 7
Preparation of Compound A-6:
Ethyl-3-[(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-Gln]-E-Prop-
enoate
[0225] ##STR111##
Preparation of Intermediate
Ethyl-3-{Boc-L-(4-F)-Phe-L-(Tr-Gln)}-E-Propenoate
[0226] This compound was prepared from
ethyl-3-(Boc-L-(Tr-Gln))-E-propenoate and BOC-L-(4-F)-Phe-OH using
a procedure like that described above (Example 2) for the
preparation of ethyl-3-(Boc-L-Phe-L-(Tr-Gln))-E-propenoate: IR
(cm.sup.-1) 3328, 1707, 1506, 1168; .sup.1H NMR (CDCl.sub.3)
.delta. 1.29 (t, 3H, J=7.2), 1.37 (s, 9H), 1.66-1.78 (m, 1H),
1.88-1.98 (m, 1H), 2.32 (t, 2H, J=6.6), 2.85-2.92 (m, 1H),
2.97-3.04 (m, 1H), 4.18 (q, 2H, J=7.2), 4.52 (m, 1H), 4.96 (m, 1H),
5.60 (d, 1H, J=15.6), 6.56 (d, 1H, J=8.1), 6.66 (dd, 1H, J=15.6,
5.1), 6.80 (s, br, 1H), 6.92-6.98 (m, 2H), 7.08-7.12 (m, 2H),
7.17-7.23 (m, 6H), 7.24-7.33 (m, 10H); Anal.
(C.sub.42H.sub.46FN.sub.3O.sub.6) C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-Val-L-(4-F)-Phe-L-(Tr-Gln)}-E-Propenoate
[0227] This compound was prepared from
ethyl-3-{Boc-L-(4-F)-Phe-L-(Tr-Gln)}-E-propenoate and Boc-L-Val-OH
in the manner described above (Example 2) for the preparation of
ethyl-3-{Boc-L-Leu-L-Phe-L-(Tr-Gln)}-E-propenoate: IR (cm.sup.-1)
3319, 1657, 1511, 1172; .sup.1H NMR (CDCl.sub.3) .delta. 0.78 (d,
3H, J=6.9), 0.87 (d, 3H, J=6.9), 1.29 (t, 3H, J=7.2), 1.37 (s, 9H),
1.69-1.79 (m, 1H), 1.93-2.06 (m, 2H), 2.33 (t, 2H, J=7.2),
2.97-3.04 (m, 1H), 3.73-3.77 (m, 1H), 4.18 (q, 2H, J=7.2),
4.42-4.54 (m, 2H), 4.80 (d, 1H, J=6.9), 5.61 (dd, 1H, J=15.6, 1.5),
6.44 (d, 1H, J=7.8), 6.69 (dd, 1H, J=15.9, 5.4), 6.71 (s, br, 1H),
6.92-6.98 (m, 2H), 7.07-7.13 (m, 2H), 7.18-7.31 (m, 16H), 8.02 (s,
1H); Anal. (C.sub.47R.sub.55FN.sub.4O.sub.7) C, H, N.
Preparation of Intermediate
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F)-Phe-L-(Tr-Gln)}-E-
-Propenoate
[0228] This compound was prepared from
ethyl-3-{Boc-L-Val-L-(4-F)-Phe-L-(Tr-Gln)}-E-propenoate and
5-methylisoxazole-3-carbonyl chloride in a manner like that
described above (Example 2) for the preparation of
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln)}-E-prope-
noate: IR (cm.sup.-1) 3319, 1657, 1511, 1172; .sup.1H NMR
(CDCl.sub.3) .delta. 0.84 (d, 3H, J=6.9), 0.89 (d, 3H, J=6.9), 1.30
(t, 3H, J=7.2), 1.68-1.80 (m, 1H), 1.95-2.06 (m, 1H), 2.08-2.17 (m,
1H), 2.34 (t, 2H, J=7.2), 2.44 (s, 3H), 2.87-2.94 (m, 1H),
3.01-3.08 (m, 1H), 4.18 (q, 2H, J=7.2), 4.48-4.56 (m, 2H), 5.68
(dd, 1H, J=15.6, 1.8), 6.23 (s, 1H), 6.39 (d, 1H, J=7.8), 6.70 (dd,
1H, J=15.9, 5.4), 6.84-6.90 (m, 3H), 7.04-7.08 (m, 4H), 7.17-7.30
(m, 16H); Anal. (C.sub.47H.sub.50N.sub.5O.sub.7) C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F)-Phe-L-Gln}-E-Prop-
enoate
[0229]
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F)-Phe-L-(Tr--
Gln)}-E-propenoate was deprotected using the procedure described
above (Example 2) for the preparation of
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln}-E-propenoate-
: IR (cm.sup.-1) 3284, 1652, 1542; .sup.1H NMR (DMSO-d.sub.6)
.delta. 0.76 (d, 3H, J=6.9), 0.79 (d, 3H, J=6.9), 1.20 (t, 3H,
J=7.2), 1.57-1.76 (m, 2H), 1.96-2.06 (3H), 2.46 (s, 3H), 2.75-2.83
(m, 1H), 2.89-2.96 (m, 1H), 4.09 (q, 2H, J=7.2), 4.13-4.25 (m, 1H),
4.35 (m, 1H), 4.49-4.56 (m, 1H), 5.53 (d, 1H, J=15.6), 6.57 (s,
1H), 6.66 (dd, 1H, J=15.6, 5.4), 6.75 (s, br, 1H), 6.97-7.03 (m,
2H), 7.17-7.24 (m, 3H), 8.15 (d, 1H, J=7.8), 8.24 (d, 1H, J=8.7),
8.32 (d, 1H, J=8.1); Anal. (C.sub.28H.sub.36N.sub.5O.sub.7) C, H,
N.
Example 8
Preparation of Compound A-7:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-((S)-Pyrrol-Ala)}-E-Provenoate
[0230] ##STR112##
Preparation of Intermediate BOC-L-(4-F-Phe)-Obn
[0231] DCC (0.765 g, 3.71 mmol, 1.05 equiv.), benzyl alcohol (0.347
mL, 3.35 mmol, 0.95 equiv.) and DMAP (0.022 g, 0.18 mmol, 0.05
equiv.) were added sequentially to a solution of Boc-L-(4-F-Phe)-OH
(1.0 g, 3.53 mmol, 1 equiv.) in CH.sub.2Cl.sub.2 (15 mL). After
stirring 18 h, the precipitate was removed by filtration, and the
filtrate was diluted with MTBE (75 mL), washed with 10% KRSO.sub.4
and brine (10 mL each), dried over Na.sub.2SO.sub.4 and evaporated.
Purification of the residue by flash column chromatography (12%
EtOAc in hexanes) gave Boc-L-(4-F-Phe)-OBn (0.992 g, 79%) as a
white solid. .sup.1H NMR spectral data matches literature (see
Jackson et al., J. Org. Chem. 1992, vol. 57, 3397).
Preparation of Intermediate
BOC-L-.alpha.-(t-Butyl-Gly)-L-(4-F-Phe)-OBn
[0232] Boc-L-(4-F-Phe)-OBn (2.0 g, 5.36 mmol, 1 equiv.) was stirred
for 1 h in a mixture of CH.sub.2Cl.sub.2 (20 mL) and TFA (10 mL),
then more TFA (10 mL) was added and the reaction solution was
stirred an additional hour. The volatiles were evaporated and the
residue was dissolved in DMF (30 mL).
Boc-L-.alpha.-(t-butyl-Gly)-OH (1.24 g, 5.36 mmol, 1 equiv.) was
added, and the solution was cooled to 0.degree. C.
N,N-diisopropylethylamine (2.80 mL, 16.1 mmol, 3 equiv.) and HATU
(2.04 g, 5.37 mmol, 1 equiv.) were added sequentially. After
stirring 20 min., the reaction mixture was allowed to warm to room
temperature over 1 h, then diluted with MTBE (500 mL) and washed
with 5% KHSO.sub.4 (100 mL), saturated NaHCO.sub.3 (50 mL), and
brine (50 mL). The organic phase was dried and evaporated.
Purification of the residue by flash column chromatography (20%
EtOAc in hexanes) gave Boc-L-.alpha.-(t-butyl-Gly)-L-(4-F-Phe)-OBn
(2.04 g, 78%) as a white foam: R.sub.f=0.49 (25% EtOAc in hexanes);
IR (cm.sup.-1) 3307, 1737, 1655 br; .sup.1H NMR (CDCl.sub.3)
.delta. 0.94 (s, 9H), 1.45 (s, 9H), 3.04 (dd, 1H, J=14.2, 5.8),
3.11 (dd, 1H, J 14.2, 6.1), 3.79 (d, 1H, J=9.3), 4.88 (dt, 1H,
J=7.8, 5.8), 5.08 (d, 1H, J=12.0), 5.16-5.23 (m, 1H), 5.19 (d, 1H,
J=12.0), 6.08 (d, 1H, J=7.8), 6.83-6.97 (m, 4H), 7.28-7.40 (m, 5H);
Anal. (C.sub.27H.sub.35FN.sub.2O.sub.5) C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-.alpha.-(t-Butyl-Gly)-L-(4-F-Phe)-L-((N-2,4-Dimethoxybenzy-
l)-(S)-Pyrrol-Ala)}-E-Propenoate
[0233] A solution of HCl in 1,4-dioxane (4.0 M, 8 mL) was added to
a solution of
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-pyrrol-Ala)}-E-propenoate
(0.309 g, 0.648 mmol, 1 equiv.) in 1,4-dioxane (8 mL). After
stirring for 1.5 h, the volatiles were evaporated to give the crude
amine salt as a foam.
[0234] Palladium on carbon (10%, 200 mg) was added to a solution of
Boc-L-.alpha.-(t-butyl-Gly)-L-(4-F-Phe)-OBn (2.04 g, 4.19 mmol) in
EtOAc (200 mL). The atmosphere was replaced with hydrogen via
balloon. After stirring 3 h, the atmosphere was replaced with
argon, and the reaction mixture was filtered through #3 and #5
Whatman filter papers. The filtrate was evaporated to give a white
foam.
[0235] This foam was combined with the crude amine salt (prepared
above) in DMF (5 mL) and cooled to 0.degree. C.
N,N-diisopropylethylamine (0.339 mL, 1.95 mmol, 3 equiv.) and HATU
(0.247 g, 0.650 mmol, 1 equiv.) were added sequentially. After
stirring 20 min., the reaction mixture was allowed to warm to room
temperature over 1 h, then diluted with MTBE (100 mL) and washed
with 5% KHSO.sub.4, saturated NaHCO.sub.3 and brine (15 mL each).
The organic phase was dried and evaporated. Purification of the
residue by flash column chromatography (60% EtOAc in hexanes) gave
ethyl-3-{Boc-L-.alpha.-(t-butyl-Gly)-L-(4-F-Phe)-L-((N-2,4-dimethoxybenzy-
l)-(S)-pyrrol-Ala)}-E-propenoate (0.364 g, 74%) as a white foam:
R.sub.f=0.34 (60% EtOAc in hexanes); IR (cm.sup.-1) 3284, 1713,
1655; .sup.1H NMR (CDCl.sub.3) .delta. 1.01 (s, 9H), 1.28 (t, 3H,
J=7.2), 1.46 (s, 9H), 1.50-1.63 (m, 2H), 1.68-1.81 (m, 1H),
1.85-1.99 (m, 1H), 2.09-2.20 (m, 1H), 2.94 (dd, 1H, J=13.5, 5.4),
3.15-3.26 (m, 3H), 3.80 (s, 3H), 3.82 (s, 3H), 3.97 (d, 1H, J=9.3),
4.17 (q, 2H, J=7.2), 4.27-4.38 (m, 1H), 4.29 (d, 1H, J=14.3), 4.42
(d, 1H, J=14.3), 4.84-4.92 (m, 1H), 5.22 (d, 1H, J=9.6), 5.74 (dd,
1H, J=15.6, 1.6), 6.45-6.53 (m, 2H), 6.69 (dd, 1H, J=15.6, 5.4),
6.76-6.87 (m, 3H), 7.10-7.28 (m, 3H), 8.26 (d, 1H, J=5.9); Anal.
(C.sub.40H.sub.55FN.sub.4O.sub.9) C, H, N.
Preparation of Intermediate
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-((N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-Propenoate
[0236] This compound was prepared from
ethyl-3-{Boc-L-.alpha.-(t-butyl-Gly)-L-(4-F-Phe)-L-((N-2,4-dimethoxybenzy-
l)-(S)-pyrrol-Ala)}-E-propenoate and isoxazole-5-carbonyl chloride
using the procedure described above (Example 2) for the preparation
of
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-(Tr-Gln)}-E-prope-
noate: R.sub.f=0.60 (10% CH.sub.3OH in CHCl.sub.3); IR (cm.sup.-1)
3295, 1713, 1666, 1643; .sup.1H NMR (CDCl.sub.3) .delta. 1.07 (s,
9H), 1.29 (t, 3H, J=7.2), 1.51-1.64 (m, 2H), 1.71-1.83 (m, 1H),
1.96-2.07 (m, 1H), 2.11-2.21 (m, 1H), 2.49 (s, 3H), 2.99 (dd, 1H,
J=13.7, 5.9), 3.13-3.26 (m, 3H), 3.80 (s, 3H), 3.81 (s, 3H), 4.18
(q, 2H, J=7.2), 4.23-4.48 (m, 4H), 4.85-4.93 (m, 1H), 5.76 (dd, 1H,
J=15.6, 1.4), 6.40-6.52 (m, 3H), 6.71 (dd, 1H, J=15.6, 5.3),
6.79-6.88 (m, 2H), 6.92 (d, 1H, J=9.0), 7.09-7.22 (m, 3H), 7.37 (d,
1H, J=9.0), 8.27 (d, 1H, J=6.2); Anal.
(C.sub.40H.sub.50FN.sub.5O.sub.9.0.25H.sub.2O)C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0237] This compound was prepared from
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-.alpha.-(t-Butyl-Gly)-L-(4-F--
Phe)-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate using
a procedure as described above (Example 6) for the preparation of
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Pyrrol-
-Ala)}-E-propenoate: R.sub.f=0.30 (5% CH.sub.3OH in
CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3307 br, 1684, 1660; .sup.1H NMR
(CDCl.sub.3) .delta. 1.02 (s, 9H), 1.29 (t, 3H, J=7.2), 1.51-1.61
(m, 1H), 1.75-1.97 (m, 2H), 2.14-2.25 (m, 1H), 2.28-2.40 (m, 1H),
2.50 (s, 3H), 3.03 (d, 2H, J=6.5), 3.27-3.42 (m, 2H), 4.18 (q, 2H,
J=7.2), 4.36 (d, 1H, J=9.8), 4.52-4.63 (m, 1H), 4.86-4.95 (m, 1H),
5.71 (dd, 1H, J=15.6, 1.4), 6.44 (s, 1H), 6.57-6.64 (m, 1H), 6.73
(dd, 1H, J=15.6, 5.3), 6.83-6.91 (m, 2H), 7.09-7.15 (m, 2H),
7.28-7.36 (m, 2H), 7.59 (d, 1H, J=8.1); Anal.
(C.sub.31H.sub.10FN.sub.5O.sub.7) C, H, N.
Example 9
Preparation of Compound A-8:
Ethyl-3-[(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Piper--
Ala)}-E-Propenoate
[0238] ##STR113##
Preparation of Intermediate
(2'S,4S,4''S)-4-{2'-(4-''-Benzyl-2''-oxo-oxazoldine-3''-carbonyl)-5'-hydr-
oxypentyl}-2,2-dimethyloxazolidine-3-carboxylic Acid tert-Butyl
Ester
[0239] A solution of borane-tetrahydrofuran complex (0.96 mL of a
1.0 M solution in THF, 0.96 mmol, 1 equiv.) was added to a
0.degree. C. solution of
(2'S,4S,4''S)-4-{2'-(4''-benzyl-2''-oxo-oxazolidine-3''-carbonyl)-pent-4'-
-enyl}-2,2-dimethyloxazolidine-3-carboxylic acid tert-butyl ester
(prepared as described in Example 6, 0.455 g, 0.963 mmol, 1 equiv.)
in THF (3 mL). After stirring 30 min., water (3 mL) and sodium
perborate tetrahydrate (0.148 g, 0.962 mmol, 1 equiv.) were added,
and the ice bath was removed. After an additional hour, the
reaction mixture was diluted with MTBE (125 mL), washed with water
(15 mL) and brine (2.times.15 mL), dried over Na.sub.2SO.sub.4, and
concentrated. Purification of the residue by flash column
chromatography (50% EtOAc in hexanes) provided
(2'S,4S,4''S)-4-{2'-(4''-benzyl-2''-oxo-oxazolidine-3''-carbonyl)-5'-hydr-
oxypentyl}-2,2-dimethyloxazolidine-3-carboxylic acid tert-butyl
ester (0.339 g, 72%) as a colorless glass: R.sub.f=0.41 (50% EtOAc
in hexanes); IR (cm.sup.-1) 3486, 1780, 1693; .sup.1H NMR
(CDCl.sub.3) .delta. 1.42-1.85 (m, 21H), 2.13-2.24 (m, 1H), 2.70
(dd, 1H, J=13.1, 10.0), 3.29-3.38 (m, 1H), 3.61-4.22 (m, 8H),
4.63-4.76 (m, 1H), 7.19-7.38 (m, 5H); Anal.
(C.sub.26H.sub.38N.sub.2O.sub.7.0.5H.sub.2O)C, H, N.
Preparation of Intermediate
(1S,3'S)-{2-(1'-(2'',4''-Dimethoxybenzyl)-2'-oxo-piperidin-3'-yl)-1-hydro-
xymethylethyl}-carbamic Acid tert-Butyl Ester
[0240]
(2'S,4S,4''S)-4-{2'-(4''-Benzyl-2''-oxo-oxazolidine-3''-carbonyl)--
5'-hydroxypentyl}-2,2-dimethyloxazolidine-3-carboxylic acid
tert-butyl ester (3.88 g, 7.92 mmol, 1 equiv.) was dissolved in
Et.sub.3N (3.97 mL, 28.51 mmol, 3.6 equiv.). The mixture was cooled
to -12.degree. C., and a solution of sulfur trioxide-pyridine
complex (5.04 g, 31.67 mmol, 4 equiv.) in DMSO (150 mL) was added
at a rate to maintain the temperature between 8-17.degree. C. The
solution was stirred at 23.degree. C. for 3 h. The reaction mixture
was cooled in an ice water bath and quenched by the addition of
H.sub.2O (150 mL). The resulting solution was extracted with EtOAc
(2.times.150 mL). The combined organic layers were washed with 5%
citric acid (100 mL), brine (100 mL), dried over Na.sub.2SO.sub.4
and filtered. The solvent was removed under reduced pressure and
the residue was dried under vacuum to give a white foam (3.53
g).
[0241] To a solution of this material (3.53 g, 7.22 mmol, 1 equiv.)
in a 2:1 mixture of THF and EtOH (120 mL) was added
2,4-dimethoxybenzylamine hydrochloride (5.88 g, 28.89 mmol, 4
equiv.), NaOAc (2.37 g, 28.89 mmol, 4 equiv.), and NaBH.sub.3CN
(0.908 g, 14.45 mmol, 2 equiv.). The reaction mixture was stirred
overnight (20 h) and then diluted with MTBE (200 mL). The organic
layer was washed with 10% KHSO.sub.4 (100 mL), saturated
NaHCO.sub.3 (100 mL), and brine (100 mL), and dried over
Na.sub.2SO.sub.4, and concentrated to give a pale yellow foam.
[0242] To a solution of this foam (3.34 g, 7.22 mmol, 1 equiv.) in
CH.sub.3OH (50 mL) was added p-toluenesulfonic acid (0.275 g, 1.44
mmol, 0.2 equiv.). The reaction mixture was stirred at 50.degree.
C. for 2.5 h and then was diluted with CH.sub.2Cl.sub.2 (100 mL).
The organic layer was washed with saturated NaHCO.sub.3 (100 mL),
dried over Na.sub.2SO.sub.4, and concentrated. The residue was
purified by flash column chromatography (3% CH.sub.3OH in
CH.sub.2Cl.sub.2) to give
(1S,3'S)-{2-(1'-(2'',4''-dimethoxybenzyl)-2'-oxo-piperidin-3'-yl)-1-hydro-
xymethyl-ethyl}-carbamic acid tert-butyl ester as a white foam
(1.33 g, 44% over three steps): .sup.1H NMR (CDCl.sub.3): .delta.
1.44 (s, 9H), 1.71-1.85 (m, 2H), 1.92-1.98 (m, 2H), 2.40-2.48 (m,
1H), 2.71-2.78 (m, 1H), 3.19-3.32 (m, 2H), 3.45-3.69 (m, 4H),
4.11-4.20 (m, 2H), 4.68 (m, 1H), 5.47 (m, 1H), 6.44 (s, 1H),
7.20-7.33 (m, 2H).
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Piper--
Ala)})-E-Propenoate
[0243]
(1S,3'S)-{2-(1'-(2'',4''-Dimethoxybenyl)-2'-oxo-piperidin-3'-yl)-1-
-hydroxy-methylethyl}-carbamic acid tert-butyl ester was converted
to the product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S-
)-Piper-Ala)}-E-propenoate in a manner analogous to the conversion
of
(1S,3'S)-{2-(1'-(2'',4''-dimethoxybenzyl)-2'-oxo-piperidin-3'-yl]-1-hydro-
xymethylethyl)-carbamic acid tert-butyl ester to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Pyrrol-
-Ala)}-E-propenoate described in Example 6 above: R.sub.f=0.24 (5%
CH.sub.3OH in CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3284 br, 1713,
1655, 1637 br; .sup.1H NMR (CDCl.sub.3) .delta. 0.94 (d, 3H,
J=6.8), 0.98 (d, 3H, J=6.8), 1.29 (t, 3H, J=7.2), 1.43-1.56 (m,
2H), 1.66-1.78 (m, 1H), 1.83-2.05 (m, 4H), 2.16-2.28 (m, 1H), 2.49
(s, 3H), 3.00 (dd, 1H, J=13.7, 6.2), 3.13 (dd, 1H, J=13.7, 5.9),
3.21-3.37 (m, 2H), 4.18 (q, 2H, J=7.2), 4.36-4.45 (m, 2H),
4.80-4.88 (m, 1H), 5.76 (dd, 1H, J=15.6, 1.6), 5.96 (s, 1H), 6.43
(s, 1H), 6.70 (dd, 1H, J=15.6, 5.3), 6.81 (d, 1H, J=8.7), 6.86-6.98
(m, 2H), 7.09-7.19 (m, 2H), 7.22-7.29 (m, 1H), 8.07 (d, 1H,
J=6.5).
Example 10
Preparation of Compound B-1:
Ethyl-3-[(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-Gln}-E-Propenoate
[0244] ##STR114##
Preparation of Intermediate trans-6-Methyl-hept-4-enoic Acid
[0245] A solution of isobutyraldehyde (9.59 g, 133 mmol, 1 equiv.)
in THF (50 mL) was added dropwise via addition funnel to a solution
of vinylmagnesium bromide (133 mL of a 1.0 M solution in THF, 133
mmol, 1.0 equiv.) in THF (300 mL) at 0.degree. C. Upon completion
of the addition, the reaction mixture was stirred for 30 min. at
0.degree. C., and then ethyl malonyl chloride (17.0 mL, 133 mmol,
1.0 equiv.) was added. After stirring for 1 h at 0.degree. C., the
reaction mixture was partitioned between saturated NH.sub.4Cl (150
mL) and a 1:1 mixture of EtOAc and hexanes (2.times.200 mL). The
combined organic layers were dried over Na.sub.2SO.sub.4 and were
concentrated. Purification of the residue by filtration through
silica gel (eluting with 5% EtOAc in hexanes) afforded the
intermediate malonate ester (11.5 g, 40% yield). This material was
not characterized, but was combined (neat) with Ti(OEt).sub.4 (1.13
mL, 5.39 mmol, 0.10 equiv.) and was heated to 190.degree. C. for 4
h, and then was cooled to 60.degree. C. EtOH (50 mL) and 6.0 M KOH
(50 mL) were added sequentially, and the brown reaction mixture was
refluxed for 4 h. After cooling to 23.degree. C., the reaction
mixture was filtered through a medium frit, and the filtrate was
partitioned between water (150 mL) and Et.sub.2O (2.times.150 mL).
The aqueous layer was then acidified to pH=2 (as indicated by pH
paper) with concentrated HCl and was extracted with a 1:1 mixture
of EtOAc and hexanes (2.times.150 mL). The combined organic layers
were dried over Na.sub.2SO.sub.4, concentrated, and the residue was
distilled at reduced pressure to afford trans-6-methyl-hept-4-enoic
acid (3.58 g, 47%) as a colorless liquid: bp 107-112.degree. C. (1
Torr); IR (cm.sup.-1) 2960, 1711; .sup.1H NMR (CDCl.sub.3) .delta.
0.96 (d, 6H, J=6.5), 2.18-2.45 (m, 5H), 5.31-5.50 (m, 2H); Anal.
(C.sub.8H.sub.14O.sub.2) C, H.
Preparation of Intermediate trans-6-Methyl-hept-4-enoic Acid
(2R-hydroxy-1R-methyl-2-phenyl-ethyl)-methyl Amide
[0246] Oxalyl chloride (2.25 mL, 25.8 mmol, 1.05 equiv.) was added
to a solution of trans-6-methyl-hept-4-enoic acid (3.50 g, 24.6
mmol, 1 equiv.) and N,N-dimethylformamide (0.03 mL, 0.39 mmol,
0.016 equiv.) in benzene (60 mL) at 23.degree. C. The reaction
mixture was stirred at 23.degree. C. for 2 h, and then was
concentrated under reduced pressure. The resulting oil was
dissolved in THF (20 mL) and was added via cannula to a solution of
(1R,2R)-(-)-pseudoephedrine (3.87 g, 23.4 mmol, 1 equiv.) and
triethylamine (3.92 mL, 28.1 mmol, 1.2 equiv.) in THF (150 mL) at
0.degree. C. The reaction mixture was stirred at 0.degree. C. for
30 min., then was partitioned between half-saturated NH.sub.4Cl
(150 mL) and EtOAc (2.times.150 mL). The combined organic layers
were dried over Na.sub.2SO.sub.4, concentrated, and the residue
purified by flash column chromatography (gradient elution 40-50%
EtOAc in hexanes) to afford trans-6-methyl-hept-4-enoic acid
(2R-hydroxy-1R-methyl-2-phenyl-ethyl)-methyl amide (6.31 g, 93%) as
a viscous oil: R.sub.f=0.35 (50% EtOAc in hexanes); IR (cm.sup.-1)
3382, 1622; .sup.1H NMR (CDCl.sub.3, mixture of rotamers) .delta.
0.96 (d, J=6.8), 0.97 (d, J=6.5), 1.11 (d, J=6.9), 2.18-2.59 (m),
2.82 (s), 2.92 (s), 3.99-4.04 (m), 4.32-4.42 (m), 4.44-4.49 (m),
4.55-4.62 (m), 5.32-5.49 (m), 7.24-7.42 (m); Anal.
(C.sub.18H.sub.27NO.sub.2) C, H, N.
Preparation of Intermediate
trans-6-Methyl-2S-(4-fluorobenzyl)-hept-4-enoic Acid
(2R-Hydroxy-1R-methyl-2-phenylethyl)methyl Amide
[0247] n-Butyllithium (32.5 mL of a 1.6 M solution in hexanes, 52.0
mmol, 3.1 equiv.) was added to a suspension of anhydrous lithium
chloride (7.18 g, 169 mmol, 10 equiv.) and diisopropylamine (7.80
mL, 55.7 mmol, 3.3 equiv.) in THF (250 mL) at -78.degree. C. The
reaction mixture was stirred for 30 min. at -78.degree. C., was
maintained at 0.degree. C. for 5 min., and subsequently cooled
again to -78.degree. C. trans-6-Methyl-hept-4-enoic acid
(2R-hydroxy-1R-methyl-2-phenyl-ethyl)-methyl amide (4.91 g, 17.0
mmol, 1 equiv) in THF (50 mL) was added via cannula, and the
resulting solution was stirred at -78.degree. C. for 1.75 h,
maintained at 0.degree. C. for 20 min., stirred at 23.degree. C.
for 5 min., and then was cooled again to 0.degree. C. A solution of
4-fluorobenzyl bromide (6.34 mL, 50.9 mmol, 3 equiv.) in THF (15
mL) was added and the reaction mixture was stirred at 0.degree. C.
for 30 min., which was then partitioned between half-saturated
NH.sub.4Cl (230 mL) and a 1:1 mixture of EtOAc and hexanes (200 mL,
2.times.150 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and were concentrated. Purification of the residue
by flash column chromatography (gradient elution 20-40% EtOAc in
hexanes) provided trans-6-methyl-2S-(4-fluorobenzyl)-hept-4-enoic
acid (2R-hydroxy-1R-methyl-2-phenylethyl)methyl amide (6.33 g, 94%)
as a viscous oil: R.sub.f=0.38 (40% EtOAc in hexanes); IR
(cm.sup.-1) 3378, 1614; .sup.1H NMR (CDCl.sub.3, mixture of
rotamers) .delta. 0.85-0.95 (m), 0.96 (d, J=6.8), 2.10-2.32 (m),
2.34-2.46 (m), 2.58 (s), 2.67-2.79 (m), 2.82-2.94 (m), 3.00-3.18
(m), 3.94 (br), 4.37-4.52 (m), 5.24-5.42 (m), 5.44-5.56 (m),
6.89-7.01 (m), 7.08-7.14 (m), 7.19-7.38 (m); Anal.
(C.sub.25H.sub.32FNO.sub.2) C, H, N.
Preparation of Intermediate
5S-(1R-Bromo-2-methylpropyl)-3R-(4-fluorobenzyl)dihydrofuran-2-one
[0248] N-Bromosuccinimide (2.93 g, 16.5 mmol, 1.05 equiv.) was
added in small portions over 10 minutes to a solution of
trans-6-methyl-2S-(4-fluorobenzyl)-hept-4-enoic acid
(2R-hydroxy-1R-methyl-2-phenylethyl)methyl amide (6.24 g, 15.7
mmol, 1 equiv.) and glacial acetic acid (4.49 mL, 78.4 mmol, 5
equiv.) in a 4:1 mixture of THF and H.sub.2O (165 mL) at 0.degree.
C. The resulting yellow solution was stirred for 15 min. at
0.degree. C., and then was warmed to 23.degree. C. and subsequently
refluxed for 45 min. After cooling to 23.degree. C., the reaction
mixture was partitioned between half-saturated NaHCO.sub.3 (200 mL)
and a 1:1 mixture of EtOAc and hexanes (2.times.200 mL, 100 mL).
The combined organic layers were dried over Na.sub.2SO.sub.4 and
were concentrated. Flash chromatographic purification of the
residue (gradient elution 4.fwdarw.10% EtOAc in hexanes) gave
5S-(1R-bromo-2-methylpropyl)-3R-(4-fluorobenzyl)dihydrofuran-2-one
(4.14 g, 80%) as a pale yellow oil (containing approximately 5-10%
unidentified impurities by .sup.1H NMR): R.sub.f=0.56 (25% EtOAc in
hexanes); IR (cm.sup.-1) 1772; .sup.1H NMR (CDCl.sub.3, major
isomer) .delta. 0.94 (d, 3H, J=6.5), 1.00 (d, 3H, J=6.8), 2.05-2.35
(m, 3H), 2.83 (dd, 1H, J=13.6, 8.4), 2.92-3.03 (m, 1H), 3.11 (dd,
1H, J=13.6, 4.7), 3.90 (dd, 1H, J=9.0, 3.7), 4.33-4.40 (m, 1H),
6.98-7.06 (m, 2H), 7.14-7.20 (m, 2H); Anal.
(C.sub.15H.sub.18BrFO.sub.2) C, H.
Preparation of Intermediate
5S-(1S-Azido-2-methylpropyl)-3R-(4-fluorobenzyl)dihydrofuran-2-one
[0249] A suspension of sodium azide (1.90 g, 29.2 mmol, 2.5 equiv.)
and
5S-(1R-bromo-2-methylpropyl)-3R-(4-fluorobenzyl)dihydrofuran-2-one
(3.85 g, 11.7 mmol, 1 equiv.) in N,N-dimethylformamide (40 mL) was
heated at 50.degree. C. for 67 hours. The reaction mixture was
cooled to 23.degree. C. and was partitioned between half-saturated
NaCl (200 mL) and a 1:1:1 mixture of EtOAc, hexanes and acetone
(2.times.200 mL, 100 mL). The combined organic layers were dried
over Na.sub.2SO.sub.4, concentrated, and the residue purified by
flash column chromatography (gradient elution 9-17% EtOAc in
hexanes) to give
5S-(1S-azido-2-methylpropyl)-3R-(4-fluorobenzyl)dihydrofuran-2-one
(2.10 g, 62%) as a white solid (containing approximately 5-10%
unidentified impurities by .sup.1H NMR): mp 91-96.degree. C.;
R.sub.f=0.44 (25% EtOAc in hexanes); IR (cm.sup.-1) 2097, 1772;
.sup.1H NMR (CDCl.sub.3, major isomer) .delta. 0.99 (d, 3H, J=6.5),
1.02 (d, 3H, J=6.8), 1.95-2.20 (m, 3H), 2.78-2.88 (m, 1H), 2.94
(dd, 1H, J=7.0, 4.2), 3.03-3.17 (m, 2H), 4.37-4.43 (m, 1H),
6.97-7.09 (m, 2H), 7.14-7.21 (m, 2H).
Preparation of Intermediate
{2-Methyl-1S-(4R-(4-fluorobenzyl)-5-oxotetrahydrofuran-2S-yl)propyl}-carb-
amic Acid tert-Butyl Ester
[0250] A suspension of
5S-(1S-azido-2-methylpropyl)-3R-(4-fluorobenzyl)dihydrofuran-2-one
(2.02 g, 6.93 mmol, 1 equiv.), di-tert-butyl dicarbonate (2.12 g,
9.71 mmol, 1.4 equiv.) and Pd/C (10%, 0.20 g) in CH.sub.3OH (100
mL) was stirred under a hydrogen atmosphere (balloon) for 16 hours.
The reaction mixture was vacuum filtered through Whatman #3 paper
and concentrated. Purification of the residue by flash column
chromatography (15% EtOAc in hexanes) provided
{2-methyl-1S-(4R-(4-fluorobenzyl)-5-oxotetrahydrofuran-2S-yl)propyl}-carb-
amic acid tert-butyl ester (1.58 g, 62%) as a white foam:
R.sub.f=0.80 (5% MeOH in CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3331,
1766, 1702; .sup.1H NMR (CDCl.sub.3) .delta. 0.93 (d, 3H, J=6.8),
0.95 (d, 3H, J=6.5), 1.41 (s, 9H), 1.71-1.83 (m, 1H), 1.95-2.06 (m,
1H), 2.16-2.27 (m, 1H), 2.80 (dd, 1H, J=13.5, 8.6), 2.88-2.99 (m,
1H), 3.09 (dd, 1H, J=13.5, 4.4), 3.32-3.40 (m, 1H), 4.42-4.48 (m,
2H), 6.95-7.03 (m, 2H), 7.11-7.18 (m, 2H); Anal.
(C.sub.20H.sub.28FNO.sub.4) C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L-(Tr-Gln)}-E-Propenoate
[0251] Lithium hydroxide (9.62 mL of a 1 M aqueous solution, 9.62
mmol, 5 equiv.) was added to a solution of
{2-methyl-1S-(4R-(4-fluorobenzyl)-5-oxotetrahydrofuran-2S-yl)propyl}-carb-
amic acid tert-butyl ester (0.703 g, 1.92 mmol, 1 equiv.) in DMB
(25 mL) at 23.degree. C. The resulting suspension was stirred at
23.degree. C. for 30 min., and then was partitioned between 10%
KRSO.sub.4 (50 mL) and CH.sub.2Cl.sub.2 (3.times.100 mL). The
combined organic layers were dried over Na.sub.2SO.sub.4,
concentrated, and the residue dissolved in CH.sub.2Cl.sub.2 (30
nL). Powdered 4A molecular sieves (0.70 g), 4-methylmorpholine
N-oxide (0.451 g, 3.85 mmol, 2 equiv.), and tetrapropylammonium
perruthenate (0.068 g, 0.19 mmol, 0.10 equiv.) were added
sequentially. The resulting dark reaction mixture was stirred for
1.33 hours at 23.degree. C., then was vacuum filtered through
Whatman #3 paper and then through Whatman #5 paper. The filtrate
was concentrated under reduced pressure to provide a dark residue,
which was dissolved in CH.sub.2Cl.sub.2 (30 mL). Crude
ethyl-3-(H.sub.2N-L-(Tr-Gln))-E-propenoate-HCl (2.30 mmol, 1.2
equiv., prepared as described in Example 2 for the preparation of
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln}-E-propenoate-
), 4-methylmorpholine (0.846 mL, 7.69 mmol, 4 equiv.), HOBt (0.390
g, 2.89 mmol, 1.5 equiv.), and EDC (0.553 g, 2.88 mmol, 1.5 equiv.)
were added sequentially, and the reaction mixture was stirred for
19 hours at 23.degree. C. and then was partitioned between brine
(100 mL) and CH.sub.2Cl.sub.2 (3.times.100 mL). The combined
organic layers were dried over Na.sub.2SO.sub.4 and were
concentrated. Purification of the residue by flash column
chromatography (gradient elution 35-40% EtOAc in hexanes) provided
ethyl-3-{Boc-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L-(Tr-Gln)}-E-pr-
openoate (0.820 g, 53%) as a tan foam: R.sub.f=0.50 (50% EtOAc in
hexanes); IR (cm.sup.-1) 3307, 1708, 1666; .sup.1H NMR (CDCl.sub.3)
.delta. 0.67 (d, 3H, J=6.8), 0.92 (d, 3H, J=6.8), 1.28 (t, 3H,
J=7.2), 1.40 (s, 9H), 1.53-1.67 (m, 1H), 1.91-2.04 (m, 2H),
2.32-2.41 (m, 2H), 2.46-2.55 (m, 1H), 2.63 (dd, 1H, J=12.1, 5.9),
2.69-2.80 (m, 1H), 2.83 (dd, 1H, J=12.1, 8.2), 3.03 (dd, 1H,
J=17.7, 10.0), 4.05-4.11 (m, 1H), 4.17 (q, 2H, J=7.2), 4.40-4.50
(m, 1H), 4.84 (d, 1H, J=8.4), 5.38 (d, 1H, J=15.7), 6.01 (d, 1H,
J=8.4), 6.60 (dd, 1H, J=15.7, 5.0), 6.92-6.99 (m, 2H), 7.03-7.12
(m, 3H), 7.17-7.30 (m, 15H); Anal.
(C.sub.48H.sub.56FN.sub.3O.sub.7) C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-Gln}-E-Propenoate
[0252]
Ethyl-3-{Boc-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L-(Tr-Gin)}-E-prop-
enoate was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-Gln}-E-propenoate in a manner analogous to that described in
Example 2 above for the conversion of
ethyl-3-{Boc-L-Leu-L-Phe-L-(Tr-Gin)}-E-propenoate to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln}-E-propenoate-
: mp=220.degree. C. (dec); R.sub.f=0.35 (10% CH.sub.3OH in
CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3277, 1715, 1643; .sup.1H NMR
(DMSO-d.sub.6) .delta. 0.81 (d, 3H, J=6.2), 0.87 (d, 3H, J=6.9),
1.21 (t, 3H, J=6.5), 1.59-1.67 (m, 2H), 2.03 (s, br, 2H), 2.21-2.24
(m, 1H), 2.46 (s, 3H), 2.57-2.68 (m, 3H), 2.80-2.95 (m, 2H), 4.09
(q, 2H, J=6.5), 4.30-4.34 (m, 2H), 5.41 (d, 1H, J=15.5), 6.55 (s,
1H), 6.61 (dd, 1H, J=15.5, 5.5), 6.73 (s, 1H), 6.99-7.16 (m, 5H),
8.01 (d, 1H, J=7.8), 8.69 (d, 1H, J=8.7); Anal.
(C.sub.29H.sub.37FN.sub.4O.sub.7) C, H, N.
Example 11
Preparation of Compound A-10:
Ethyl-3-((5'-Methylisoxazole-3'-carbonyl)-L-Leu-NCH.sub.3-L-Phe-L-Gln)-E--
Provenoate
[0253] ##STR115##
Preparation of Intermediate Boc-L-Leu-NCH.sub.3-L-Phe-OCH.sub.3
[0254] NCH.sub.3-L-Phe-OCH.sub.3--HCl (1.4 g) was dissolved in
CH.sub.2Cl.sub.2 (50 mL) and poured into a combination of aqueous
(aq) 1 N NaOH (7 mL) and saturated aqueous NaHCO.sub.3 (25 mL).
After mixing, the organic phase was separated and the aqueous phase
was washed with CH.sub.2Cl.sub.2 (3.times.50 mL). The combined
organic phases were dried over Na.sub.2SO.sub.4 and evaporated to
give the free amine as a clear colorless oil (1.14 g, 5.90 mmol). A
solution of this amine and (iPr).sub.2NEt (1.13 mL, 6.49 mmol) in
DMF (10 mL) was added dropwise to a 0.degree. C. solution of
Boc-L-Leu-OH (1.50 g, 6.49 mmol) and HOBt (0.877 g, 6.49 mmol) in
DMF (10 mL). DCC (1.47 g, 7.12 mmol) was added. The reaction
mixture was stirred at 0.degree. C. for 1 h, and was then stirred
at 23.degree. C. for 48 h. The mixture was filtered to remove the
precipitate (ppt) and the filtrate was evaporated. The residue was
dissolved in CH.sub.2Cl.sub.2 (200 mL), washed with saturated
aqueous NaHCO.sub.3 (40 mL), dried over Na.sub.2SO.sub.4 and
evaporated. The residue was purified by flash column chromatography
(25% EtOAc in hexanes) to give Boc-L-Leu-NCH.sub.3-L-Phe-OCH.sub.3
as a white solid (2.04 g, 85%): mp=126-127.degree. C.; IR
(cm.sup.-1) 3401, 3319, 1743, 1708, 1649; .sup.1H NMR (CDCl.sub.3)
(major isomer) .delta. 0.92 (d, 3H, J=6.8), 0.95 (d, 3H, J=6.5),
1.32-1.48 (m, 2H), 1.41 (s, 9H), 1.61-1.77 (m, 1H), 2.90 (s, 3H),
3.04 (dd, 1H, J=14.5, 10.5), 3.37 (dd, 1H, J=14.5, 5.5), 3.72 (s,
3H), 4.48-4.57 (m, 1H), 4.98-5.04 (m, 1H), 5.20 (dd, 1H, J=10.5,
5.5), 7.16-7.32 (m 5H); Anal. (C.sub.22H.sub.34N.sub.2O.sub.5) C,
H, N.
Preparation of Intermediate Boc-L-Leu-NCH.sub.3-L-Phe-OH
[0255] Boc-L-Leu-NCH.sub.3-L-Phe-OCH.sub.3 (0.625 g, 1.54 mmol) was
dissolved in CH.sub.3OH (20 mL) and cooled to 0.degree. C. Aqueous
NaOH (6.15 mL of a 2N solution, 12.3 mmol) was added dropwise. The
reaction mixture was stirred for 3 h at 23.degree. C., and then
poured into 10% aqueous KHSO.sub.4 (150 mL). This mixture was
extracted with CH.sub.2Cl.sub.2 (3.times.100 mL), and the combined
organic phases were dried over Na.sub.2SO.sub.4 and evaporated to
give BOC-L-Leu-NCH.sub.3-L-Phe-OH as a white foam (0.617 g,
quant.), which was used without purification.
Preparation of Intermediate
Ethyl-3-{BOC-L-Leu-NCH.sub.3-L-Phe-L-(Tr-Gln)}-E-Propenoate
[0256] This intermediate was prepared from
Boc-L-Leu-NCH.sub.3-L-Phe-OH and
ethyl-3-{H.sub.2N-L-(Tr-Gln)}-E-propenoate.HCl (prepared as
described in Example 2) in a manner analogous to that described for
the preparation of
ethyl-3-{Boc-L-Leu-L-Phe-L-(Tr-Gln)}-E-propenoate in Example 2
above: IR (cm.sup.-1) 3295, 1713, 1672, 1649; .sup.1H NMR
(CDCl.sub.3) (mixture of isomers) .delta. 0.65 (d, J=6.2), 0.66 (d,
J=6.5), 0.84 (d, J=6.5), 0.88 (d, J=6.5), 1.02-1.22 (m), 1.23-1.38
(m), 1.33 (s), 1.41 (s), 1.55-1.82 (m), 1.89-2.07 (m), 2.23-2.30
(m), 2.90 (s), 2.94 (s), 3.01 (dd, J=14.6, 10.9), 3.03-3.13 (m),
3.26-3.37 (m), 3.27 (dd, J=14.6, 3.4), 3.42-3.54 (m), 4.00-4.22
(m), 4.37-4.73 (m), 4.82-4.89 (m), 5.63-5.70 (m), 5.95 (dd, J=15.9,
1.2), 6.23-6.28 (m), 6.66-6.75 (m), 6.79-6.89 (m), 7.09-7.34 (m),
8.14 (d, J=8.7); Anal. (C.sub.49H.sub.60N.sub.4O.sub.7) C, H,
N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Leu-NCH.sub.3-L-Phe-L-Gln}-E--
Propenoate
[0257] Ethyl-3-{Boc-L-Leu-NCH.sub.3-L-Phe-L-(Tr-Gln)}-E-propenoate
was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-NCH.sub.3-L-Phe-L-Gln}-E--
propenoate in a manner analogous to that described in Example 2
above for the conversion of
ethyl-3-{Boc-L-Leu-L-Phe-L-(Tr-Gln)}-E-propenoate to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln}-E-pr-
openoate: R.sub.f=0.23 (5% CH.sub.3OH in CH.sub.2Cl.sub.2); IR
(cm.sup.-1) 3295, 1713, 1666, 1637; .sup.1H NMR (CDCl.sub.3)
(mixture of isomers) .delta. 0.65 (d, J=6.5), 0.71 (d, J=6.5), 0.93
(d, J=6.5), 0.94 (d, J=6.5), 1.30 (t, J=7.2), 1.24-1.73 (m),
1.81-2.22 (m), 2.45 (s), 2.48 (s), 2.86-2.93 (m), 2.96 (s), 2.97
(s), 3.03-3.14 (m), 3.21-3.31 (m), 3.48 (dd, J=14.0, 5.9), 4.19 (q,
J=7.2), 4.20 (q, J=7.2), 4.38-4.45 (m), 4.52-4.70 (m), 4.74-4.81
(m), 5.62-5.67 (m), 5.73-5.79 (m), 5.81 (dd, J=15.6, 1.6), 5.99
(dd, J=15.6, 1.6), 6.03-6.09 (m), 6.35 (s), 6.39 (s), 6.40-6.45
(m), 6.77-6.94 (m), 7.42 (d, J=7.2), 8.13 (d, J=7.8).
Example 12
Preparation of Compound C-1:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-Gln}-E-Pr-
openoate
[0258] ##STR116##
Preparation of Intermediate Allyl
(S)-1-Hydroxy-3-(4-fluorophenyl)propionate
[0259] In a flask fitted with a thermometer and a reflux condenser
was dissolved methyl (S)-1-hydroxy-3-(4-fluorophenyl)propionate
(prepared from L-H.sub.2N-(4-F-Phe)-OCH.sub.3 by the method
described in Hoffman et al., Tetrahedron 1992, vol. 48, 3007) (0.99
g, 5.0 mmol) in allyl alcohol (50 m]L). Titanium tetraisopropoxide
(1.53 mL, 5.0 mmol) was added, and the reaction brought to
90.degree. C. for 3.5 h. The reaction was cooled to room
temperature and poured into 250 mL of 1:1 EtOAc/saturated
NH.sub.4Cl solution. The organic phase was separated and washed
with water (100 mL), brine (100 mL), dried (MgSO.sub.4), and the
solvent removed. The residue was subjected to flash column
chromatography eluting with a gradient of 5-10% EtOAc/hexanes to
afford 0.77 g (68%) of allyl
(S)-1-hydroxy-3-(4-fluorophenyl)propionate as a clear liquid:
R.sub.f 0.21 (15% EtOAc/hexanes); IR (neat) 3470 (broad), 1734,
1510, 1221 cm.sup.-1; .sup.1H NMR (DMSO-d.sub.6) .delta. 1.24-1.27
(m, 1H), 2.92-2.99 (m, 1H), 3.09-3.15 (m, 1H), 4.43-4.47 (m, 1H),
4.65 (d, 2H, J=5.9),5.28-5.37 (m, 2H), 5.86-5.95 (m, 1H), 6.95-7.01
(m, 2H), 7.16-7.21 (m, 2H).
Preparation of Intermediate
Boc-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.dbd.CH.sub.2
[0260] Allyl (S)-1-hydroxy-3-(4-fluorophenyl)propionate (0.070 g,
0.31 mmol) was dissolved in CH.sub.2Cl.sub.2 (20 mL). Boc-L-Val-OH
(0.068 g, 0.31 mmol) was added, followed by DMAP (0.004 g, 0.03
mmol) and DCC (0.067 g, 0.33 mmol). The reaction was stirred at
room temperature overnight, and the solvent was removed in vacuo.
The residue was subjected to flash column chromatography eluting
with a gradient of 3-5% EtOAc/hexanes. The
Boc-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.dbd.CH.sub.2 product was
obtained as 0.12 g (90%) of a clear oil: R.sub.f=0.18 (10%
EtOAc/hexanes); IR (neat) 1752, 1717 1510 cm.sup.-1; .sup.1H NMR
(DMSO-d.sub.6) .delta. 0.80-0.85 (m, 6H), 1.36 (s, 9H), 1.97-2.04
(m, 1H), 3.07-3.15 (m, 2H), 3.89-3.94 (m, 1H), 4.51-4.55 (m, 2H),
5.17-5.30 (m, 3H), 5.75-5.84 (m, 1H), 7.06-7.17 (m, 3H), 7.27-7.32
(m, 2H); Anal. (C.sub.22H.sub.30NO.sub.6F) C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-Val-O-L-(4-F-Phe)-L-(Tr-Gln)}-E-Propenoate
[0261] Boc-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.dbd.CH.sub.2 (0.65 g,
1.52 mmol) was dissolved in THF (15 mL).
Tetrakis(triphenylphosphine)palladium(0) (0.035 g, 0.03 mmol) was
added, and the reaction stirred 5 min. at 23.degree. C. Morpholine
(0.16 mL, 1.83 mmol) was added dropwise, and the reaction stirred
at room temperature for 2 h. The solvent was removed in vacuo, and
the residue taken up in 50 mL of 4:1 hexanes/Et.sub.2O. The product
was extracted into sat. NaHCO.sub.3 solution (50 mL), and the
organic phase discarded. The aqueous phase was acidified to pH=1-2
with solid KHSO.sub.4, and the product re-extracted into EtOAc (50
mL). The organic phase was washed with brine (50 mL), dried
(MgSO.sub.4), and concentrated to give 0.50 g (86%) of the free
acid as a clear oil. This material was dissolved in DMF (6 mL).
Diisopropylethylamine (0.43 mL, 2.50 mmol) was added, followed by
ethyl-3-{H.sub.2N-L-(Tr-Gln)}-E-propenoate.HCl (prepared as
described in Example 2 above, 0.55 g, 1.25 mmol). The reaction was
cooled to 0.degree. C. HATU (0.48 g, 1.25 mmol) was added, and the
reaction allowed to warm to room temperature. The DMF was removed
in vacuo. The residue was dissolved with EtOAc (30 mL), and the
organic phase washed consecutively with 10% HCl solution (25 mL),
saturated NaHCO.sub.3 solution (25 mL), H.sub.2O (25 mL), and brine
(25 mL). The organic layer was dried (MgSO.sub.4), filtered, and
concentrated, and the residue was purified by flash column
chromatography (0.fwdarw.1.0% MeOH/CH.sub.2Cl.sub.2) to give 0.40 g
(39%) of ethyl-3-{Boc-L-Val-O-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
as a white amorphous solid: R.sub.f=0.25 (3% MeOH/CHCl.sub.3);
IR(KBr) 1691, 1512, 1159 cm.sup.-1; .sup.1H NMR (DMSO-d.sub.6)
.delta. 0.78-0.83 (m, 6H), 1.20 (t, 3H, J=7.0), 1.35 (s, 9H),
1.58-1.66 (m, 2H), 1.96-2.02 (m, 1H), 2.19-2.33 (m, 2H), 2.99-3.02
(m, 2H), 3.82-3.87 (m, 1H), 4.09 (q, 2H, J=7.0), 4.33-4.37 (m, 1H),
5.03-5.08 (m, 1H), 5.60 (d, 1H, J=15.8), 6.67 (dd, 1H, J=15.8,
5.5), 7.02-7.08 (m, 2H), 7.14-7.28 (m, 18H), 8.12 (d, 1H, J=8.1),
8.59 (s, 1H); Anal. (C.sub.47H.sub.54N.sub.3O.sub.8F) C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-Gln}-E-Pr-
openoate
[0262] Ethyl-3-{Boc-L-Val-O-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-Gln}-E-pr-
openoate in a manner analogous to that described in Example 2 above
for the conversion of
ethyl-3-{Boc-L-Leu-L-Phe-L-(Tr-Gin)}-E-propenoate to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln}-E-pr-
openoate: R.sub.f=0.05 (3% MeOH/CHCl.sub.3); IR (KBr) 1746, 1719,
1661, 1549 cm.sup.-1; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.87 (d,
3H, J=6.6), 0.92 (d, 3H, J=6.6), 1.20 (t, 3H, J=7.0), 1.61-1.74 (m,
2H), 1.96-2.01 (m, 2H), 2.15-2.22 (m, 1H), 2.46 (s, 3H), 3.00-3.03
(m, 2H), 4.10 (q, 2H, J=7.0), 4.27-4.32 (m, 1H), 4.33-4.38 (m, 1H),
5.06-5.11 (m, 1H), 5.63 (d, 1H, J=15.6), 6.54 (s, 1H), 6.68 (dd,
1H, J=15.6, 5.5), 6.78 (s, br, 1H), 6.95-7.00 (m, 2H), 7.20-7.24
(m, 3H), 8.06 (d, 1H, J=8.1), 8.87 (d, 1H, J=7.7); Anal.
(C.sub.28H.sub.35N.sub.4O.sub.8F) C, H, N.
Example 13
Preparation of Compound A-11:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-(2-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-Propenoate
[0263] ##STR117##
Preparation of Intermediate
Ethyl-3-{Boc-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-Propenoate
[0264] Boc-(4-F-Phe)-OH (4.46 g, 15.75 mmol, 1 equiv.) and
CH.sub.3I (7.84 mL, 126 mmol, 8 equiv.) were dissolved in dry THF
(100 mL) and cooled to 0.degree. C. NaH (1.89 g, 47.25 mmol, 3
equiv.) was added to this solution with vigorous stirring. After
stirring at 23.degree. C. for 24 h, EtOAc (3 mL) and H.sub.2O (3
mL) were added carefully to the mixture, and the resulting
suspension was evaporated to dryness. After dissolving in H.sub.2O
(100 mL), the reaction mixture was washed with Et.sub.2O
(2.times.100 mL). The aqueous layer was acidified to pH=3 with 10%
citric acid solution, and then extracted with EtOAc (2.times.100
mL). The combined EtOAc extracts were washed successively with
half-saturated NaHCO.sub.3 (150 mL), 5% Na.sub.2S.sub.2O.sub.3 (150
mL), and H.sub.2O (150 mL), dried over Na.sub.2SO.sub.4, and
concentrated to give Boc-NCH.sub.3-(4-F-Phe)-OH as a pale yellow
foam (4.37 g, 80%), which was used without further purification:
.sup.1H NMR (CDCl.sub.3, mixture of isomers) .delta. 1.35 (s), 1.40
(s), 2.69 (s), 2.75 (s), 2.96-3.13 (m), 3.23-3.33 (m), 4.54-4.58
(m), 4.76-4.81 (m), 6.96-7.01 (m), 7.15-7.17 (m).
[0265] A solution of HCl in 1,4-dioxane (4.0 M, 15 mL) was added to
a solution of ethyl-3-(Boc-L-(Tr-Gln))-E-propenoate (prepared as
described in Example 2 above, 3.57 g, 6.73 mmol, 1 equiv.) in the
same solvent (15 mL) at 23.degree. C. After 2 h, the volatiles were
removed under reduced pressure. The residue was dissolved in
CH.sub.2Cl.sub.2 (50 mL), and Boc-NCH.sub.3-(4-F-Phe)-OH (prepared
as in the preceding paragraph, 2.0 g, 6.73 mmol, 1.0 equiv.), HOBt
(1.23 g, 9.09 mmol, 1.5 equiv), 4-methylmorpholine (2.0 mL, 18.19
mmol, 3 equiv.), and EDC (1.74 g, 9.09 mmol, 1.5 equiv.) were added
sequentially. The reaction mixture was stirred at 23.degree. C.
overnight, and then was partitioned between water (100 mL) and
CH.sub.2Cl.sub.2 (2.times.100 mL). The combined organic layers were
dried over Na.sub.2SO.sub.4, concentrated, and the residue was
purified by flash column chromatography (30% EtOAc in hexane) to
afford ethyl-3-{Boc-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
(4.07 g, 84%) as white foam: IR (cm.sup.-1) 1666, 1510, 1167;
.sup.1H NMR (CDCl.sub.3, mixture of isomers) .delta. 1.29 (t,
J=7.2), 1.37 (s), 1.65-1.75 (m), 1.95-2.06 (m), 2.29-2.33 (m), 2.66
(s), 2.91-2.99 (m), 3.22-3.29 (m), 4.18 (q, J=7.2), 4.52-4.58 (m),
5.68 (d, J=15.9), 6.45 (d, J=8.4), 6.74 (dd, J=15.6, 5.4),
6.91-6.99 (m), 7.11-7.33 (m); Anal.
(C.sub.43H.sub.48FN.sub.3O.sub.6) C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-(2-Naphth)-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-Propenoate
[0266] A solution of HCl in 1,4-dioxane (4.0 M, 3 mL), was added to
a solution of
ethyl-3-{Boc-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate (0.388
g, 0.54 mmol, 1 equiv.) in the same solvent (3 mL) at 23.degree. C.
After 2 h, the volatiles were removed under reduced pressure. The
residue was dissolved in DMF (10 mL), cooled at 0.degree. C., and
DIEA (0.188 mL, 1.08 mmol, 2 equiv.), Boc-L-(2-Naphth)-OH (0.170 g,
0.54 mmol, 1.0 equiv.) and HATU (0.205 g, 0.54 mmol, 1 equiv.) were
added sequentially. The reaction mixture was stirred at 23.degree.
C. for 1 h. The volatiles were removed under reduced pressure, and
the resulting residue was taken into EtOAc (50 mL), and washed with
0.5 N HCl (50 mL), saturated NaHCO.sub.3 (50 mL) and brine (50 mL).
The organic layer was dried over Na.sub.2SO.sub.4, concentrated,
and the residue was purified by flash column chromatography (40%
EtOAc in hexane) to afford
ethyl-3-{Boc-L-(2-Naphth)-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
(0.437 g, 88%) as white foam: IR (cm.sup.-1) 1656, 1509, 1162;
.sup.1H NMR (CDCl.sub.3, mixture of isomers) .delta. 0.88 (t,
J=7.2), 1.27 (s), 1.30 (s), 1.48-1.58 (m), 1.64-1.67 (m), 1.97-2.11
(m), 2.23-2.28 (m), 2.42-2.50 (m), 2.62-2.69 (m), 2.80 (s), 2.90
(s), 3.00-3.07 (m), 3.15-3.20 (m), 3.25-3.32 (m), 4.18 (q, J=7.2),
4.42-4.46 (m), 4.53-4.56 (m), 4.61-4.66 (m), 4.72-4.82 (m),
5.94-5.00 (m), 5.63 (d, J=15.6), 6.12 (d, J=15.6), 6.60 (dd,
J=15.6, 5.4), 6.75-6.89 (m), 6.70-7.08 (m), 7.19-7.30 (m),
7.41-7.50 (m), 7.70-7.82 (m), 8.80 (d, J=8.4); Anal.
(C.sub.56H.sub.59FN.sub.4O.sub.7) C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-(2-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-Propenoate
[0267]
Ethyl-3-{Boc-L-(2-Naphth)-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-prop-
enoate was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-(2-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-propenoate in a manner analogous to that described in
Example 2 above for the conversion of
ethyl-3-{Boc-L-Leu-L-Phe-L-(Tr-Gln)}-E-propenoate to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-L-Phe-L-Gln}-E-propenoate
IR (cm.sup.-1) 3296, 1654, 1510; .sup.1H NMR (DMSO-d.sub.6, mixture
of isomers) .delta. 1.17-1.24 (m), 1.62-1.78 (m), 2.04-2.15 (m),
2.38 (s), 2.42 (s), 2.71-2.79 (m), 2.84 (s), 2.87-2.92 (m), 3.03
(s), 3.15 (d, J=7.5), 3.98-4.06 (m), 4.08-4.12 (m), 4.38-4.42 (m),
4.94 (m), 5.03-5.07 (m), 5.09-5.18 (m), 5.66-5.82 (m), 6.42 (s),
6.43 (s), 6.66-6.81 (m), 6.88-6.94 (m), 7.01-7.06 (m), 7.13-7.17
(m), 7.24-7.34 (m), 7.43-7.46 (m), 7.56 (s), 7.76-7.84 (m), 8.08
(d, J=7.8), 8.60 (d, J=8.4), 9.01 (d, J=7.2); Anal.
(C.sub.37H.sub.40FN.sub.5O.sub.7.0.75H.sub.2O)C, H, N.
Example 14
Preparation of Compound A-9:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-His-NCH.sub.3-L-(4-F-Phe)-L-G-
ln}-E-Propenoate
[0268] ##STR118##
[0269] Ethyl-3-{Boc-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
(described in Example 13 above) was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-His-NCH.sub.3-L-(4-F-Phe)-L-G-
ln}-E-propenoate in a manner analogous to the preparation of
product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-(2-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-propenoate described in Example 13 above (utilizing
Boc-L-(Tr-His)-OH in lieu of Boc-L-(2-Naphth)-OH): IR (cm.sup.-1)
3302, 1665, 1202; .sup.1H NMR (DMSO-d.sub.6, mixture of isomers)
.delta. 1.21 (t, J=7.2), 1.70-1.78 (m), 2.05-2.09 (m), 2.41 (s),
2.44 (s), 2.69-3.26 (m), 4.11 (q, J=7.2), 4.38-4.53 (m), 5.07-5.19
(m)+6.51-5.84 (m), 6.44 (s), 6.48 (s), 6.63-6.86 (m), 6.89-7.01
(m), 7.09-7.19 (m), 7.23-7.42 (m), 8.02 (d, J=8.7), 8.15 (d,
J=8.1), 8.59 (d, J=8.7), 8.94 (s), 9.04 (s), 9.14 (d, J=6.9); Anal.
(C.sub.30H.sub.36FN.sub.7O.sub.7.TFA.H.sub.2O)C, H, N.
Example 15
Preparation of Compound A-12:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Leu-NCH.sub.3-L-(4-F-Phe)-L-G-
ln}-E-Propenoate
[0270] ##STR119##
[0271] Ethyl-3-{Boc-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
(described in Example 13 above) was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Leu-NCH.sub.3-L-(4-F-Phe)-L-G-
ln}-E-propenoate in a manner analogous to the preparation of
product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-(2-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-propenoate described in Example 13 above (utilizing
Boc-L-Leu-OH in lieu of Boc-L-(2-Naphth)-OH): IR (cm.sup.-1) 3325,
1663, 1171; .sup.1H NMR (DMSO-d.sub.6, mixture of isomers) .delta.
0.64-0.67 (m), 0.86-0.88 (m), 1.17-1.23 (m), 1.32-1.40 (m),
1.59-1.75 (m), 1.98-2.07 (m), 2.42 (s), 2.45 (s), 2.08 (s),
2.86-2.93 (m), 2.99 (s), 3.12-3.20 (m), 4.05-4.15 (m), 4.41-4.50
(m), 4.82-5.07 (m), 5.62 (d, J=15.9), 5.86 (d, J=15.9), 6.51 (s),
6.53 (s), 6.68-6.73 (m), 6.93-6.99 (m), 7.09-7.21 (m), 7.26-7.31
(m), 8.03 (d, J=7.8), 8.08 (d, J=7.8), 8.41 (d, J=8.1), 8.94 (d,
J=7.2); Anal.
(C.sub.30H.sub.40FN.sub.5O.sub.7.1.25CH.sub.2Cl.sub.2) C, H, N.
Example 16
Preparation of Compound A-13:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-(1-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-Propenoate
[0272] ##STR120##
[0273] Ethyl-3-{Boc-NCH.sub.3-L-(4-F-Phe)-L-(Tr-Gln)}-E-propenoate
(described in Example 13 above) was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-(1-Naphth)-NCH.sub.3-L-(4-F-P-
he)-L-Gln}-E-propenoate in a manner analogous to the preparation of
product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-(2-Naphth)-NCH.sub.3--
L-(4-F-Phe)-L-Gln}-E-propenoate described in Example 13 above, but
utilizing Boc-L-(1-Naphth)-OH in lieu of Boc-L-(2-Naphth)-OH: IR
(cm.sup.-1) 3308, 1659, 1169; .sup.1H NMR (DMSO-d.sub.6, mixture of
isomers) .delta. 1.16-1.23 (m), 1.61-1.78 (m), 1.98-2.02 (m),
2.07-2.12 (m), 2.41 (s), 2.43 (s), 2.77 (s), 2.78 (s), 2.84-2.87
(m), 2.93-3.03 (m), 3.08-3.14 (m), 3.31-3.38 (m), 4.00-4.15 (m),
4.27-4.32 (m), 4.40-4.46 (m), 4.58-4.64 (m), 5.07-5.17 (m),
5.57-5.73 (m), 6.45 (s), 6.57-6.61 (m), 6.71-6.88 (m), 6.89-6.91
(m), 7.11-7.19 (m), 7.31-7.38 (m), 7.50-7.58 (m), 7.73-7.78 (m),
7.83-7.94 (m), 8.08 (d, J=8.1), 8.13 (d, J=8.7), 8.62 (d, J=8.1),
9.14 (d, J=8.1); Anal. (C.sub.37H.sub.40FN.sub.5O.sub.7) C, H,
N.
Example 17
Preparation of Compound B-2:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0274] ##STR121##
Preparation of Intermediate
Ethyl-3-{Boc-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L-{(N-2,4-Dimethoxybenzyl-
)-(S)-Pyrrol-Ala}-E-Propenoate
[0275] This intermediate was prepared from
{2-methyl-1S-(4R-(4-fluorobenzyl)-5-oxotetrahydrofuran-2S-yl)propyl}-carb-
amic acid tert-butyl ester (described in Example 10) and
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-pyrrol-Ala)}-E-propenoate
(described in Example 6) in a manner analogous to the preparation
of ethyl-3-{Boc-L-(4-Me-Phe)-L-(Tr-Gln)}-E-propenoate (Example 4)
above: R.sub.f=0.24 (60% EtOAc in hexanes); IR (cm.sup.-1) 3293,
1717, 1668; .sup.1H NMR (CDCl.sub.3) .delta. 0.82 (d, 3H, J=6.8),
1.01 (d, 3H, J=6.8), 1.30 (t, 3H, J=7.2), 1.51-1.65 (m, 2H),
1.84-1.96 (m, 1H), 2.16-2.37 (m, 2H), 2.47 (s, 3H), 2.49-2.55 (m,
3H), 2.85-3.01 (m, 2H), 3.12-3.25 (m, 3H), 3.77 (s, 3H), 3.78 (s,
3H), 4.18 (q, 2H, J=7.2), 4.31-4.49 (m, 3H), 4.65-4.70 (m, 1H),
5.53 (dd, 1H, J=15.7, 1.4), 6.39-6.44 (m, 3H), 6.63 (dd, 1H,
J=15.7, 5.4), 6.93-7.01 (m, 2H), 7.05-7.10 (m, 1H), 7.12-7.18 (m,
2H), 7.24 (d, 1H, J=8.7), 7.47 (d, 1H, J=6.5); Anal.
(C.sub.40H.sub.49FN.sub.4O.sub.9.0.5H.sub.2O)C, H, N.
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)L-(4-F-Ph-
e)-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0276]
Ethyl-3-{Boc-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L-((N-2,4-dimethox-
ybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate was converted to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-((S)-Pyrrol-Ala)}-E-propenoate in a manner analogous to the
conversion of
ethyl-3-{Boc-L-Val-L-(4-F-Phe)-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)-
}-E-propenoate to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Pyrrol-
-Ala)}-E-propenoate described in Example 6 above:
mp=178-181.degree. C.; R.sub.f=0.49 (10% CH.sub.3OH in CHCl.sub.3);
IR (cm.sup.-1) 3295, 1678 br; .sup.1H NMR (CDCl.sub.3) .delta. 0.85
(d, 3H, J=6.8), 1.03 (d, 3H, J=6.5), 1.30 (t, 3H, J=7.2), 1.51-1.62
(m, 1H), 1.71-1.93 (m, 2H), 2.27-2.40 (m, 2H), 2.47 (s, 3H),
2.51-2.75 (m, 3H), 2.82-2.98 (m, 2H), 3.11-3.24 (m, 1H), 3.26-3.42
(m, 2H), 4.18 (q, 2H, J=7.2), 4.41-4.53 (m, 1H), 4.63-4.72 (m, 1H),
5.50 (d, 1H, J=15.4), 5.88 (s, 1H), 6.39 (s, 1H), 6.63 (dd, 1H,
J=15.4, 5.3), 6.92-7.03 (m, 2H), 7.08-7.31 (m, 4H); Anal.
(C.sub.31H.sub.39FN.sub.4O.sub.7) C, H, N.
Example 18
Preparation of Compound C-2:
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-((S)-Pyrr-
ol-Ala)}-E-Propenoate
[0277] ##STR122##
Preparation of Intermediate
(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.dbd.CH.s-
ub.2
[0278] Boc-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.dbd.CH.sub.2 (prepared
as described in Example 12 above, 0.91 g, 2.15 mmol) was dissolved
in 1,4-dioxane (28 mL), and a solution of HCl in 1,4-dioxane (4.0
M, 14 mL) was added. The reaction was stirred at room temperature
for 14 h. The solvent was removed by evaporation, and the residue
taken up in EtOAc (50 mL). The organic phase was washed with
saturated NaHCO.sub.3 solution (50 mL) and then brine (50 mL),
dried (MgSO.sub.4), and the solvent removed to give 0.66 g (quant.)
of a clear oil.
[0279] This material was dissolved in CH.sub.2Cl.sub.2 (20 mL).
Pyridine (0.17 mL, 2.08 mmol) was added, and the reaction was
cooled to 0.degree. C. 5-Methylisoxazole-3-carbonyl chloride (0.33
g, 2.27 mmol) dissolved in 2 mL of CH.sub.2Cl.sub.2 was added, and
the reaction warmed to room temperature over 1 h. The solvent was
removed in vacuo, and the residue purified by flash column
chromatography eluting with a gradient of 5-10% EtOAc/hexanes. The
(5'-methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.dbd.CH.s-
ub.2 product was obtained as 0.70 g (82%) of a white crystalline
solid: R.sub.f=0.20 (30% EtOAc/hexanes); IR (KBr) 1745, 1661, 1553,
1186 cm.sup.-1; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.85-0.92 (m,
6H), 2.14-2.21 (m, 1H), 2.46 (s, 3H), 3.05-3.19 (m, 2H), 4.32-4.37
(m, 1H), 4.53-4.60 (m, 2H), 5.16-5.28 (m, 3H), 5.74-5.85 (m, 1H),
6.56 (s, 1H), 7.01-7.07 (m, 2H), 7.26-7.29 (m, 2H), 8.76 (d, 1H,
J=8.1); Anal. (C.sub.22H.sub.25N.sub.2O.sub.6F) C, H, N.
Preparation of Intermediate
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-((N-2,4-D-
imethoxybenzyl)-(S)-Pyrrol-Ala)}-E-Propenoate
[0280]
(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-OCH.sub.2CH.d-
bd.CH.sub.2 (0.67 g, 1.55 mmol) was dissolved in THF (15 mL).
Tetrakis(triphenylphosphine)palladium(0) (0.036 g, 0.03 mmol) was
added, and the reaction mixture was stirred for 5 minutes.
Morpholine (0.16 mL, 1.86 mmol) was added dropwise, and the
reaction stirred at room temperature for 6 h. The solvent was
removed in vacuo, and the residue taken up in 50 mL of Et.sub.2O.
The product was extracted twice into saturated NaHCO.sub.3 solution
(50 mL), and the organic phase discarded. The aqueous phase was
acidified to pH=1-2 with 10% HCl, and the product extracted twice
with EtOAc (40 mL). The organic phase was washed with brine (50
mL), dried (MgSO.sub.4), and concentrated to give 0.57 g (95%) of
an oil which crystallized upon standing.
[0281] A portion of this material (0.19 g, 0.50 mmol) was dissolved
in DMF (3 mL). Diisopropylethylamine (0.34 mL, 1.0 mmol) was added,
followed by
ethyl-3-{H.sub.2N-L-((N-2,4-dimethoxybenzyl)-(S)-pyrrol-Ala)}-E-propen-
oate-HCl (prepared as described in Examples 4 and 6 above, 0.19 g,
0.50 mmol). The reaction was cooled to 0.degree. C. HATU (0.19 g,
0.50 mmol) was added, and the reaction was allowed to warm to room
temperature. The DMF was removed in vacuo. The residue was
dissolved with EtOAc (30 mL), and the organic phase washed
consecutively with 10% HCl solution (25 mL), saturated NaHCO.sub.3
solution (25 mL), H.sub.2O (25 mL), and brine (25 mL). The solvent
was dried (MgSO.sub.4) and filtered, and the residue purified by
flash column chromatography (0.fwdarw.1.0% MeOH/CH.sub.2Cl.sub.2)
to give
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-((N-2,4-d-
imethoxybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate (0.27 g, 72%) as a
white amorphous solid: R.sub.f=0.18 (3% MeOH/CHCl.sub.3); IR(KBr)
1671, 1547, 1510, 1209 cm.sup.-1; .sup.1H NMR (DMSO-d.sub.6)
.delta. 0.88 (d, 3H, J=7.0), 0.93 (d, 3H, J=7.0), 1.20 (t, 3H,
J=7.0), 1.41-1.58 (m, 2H), 1.79-1.98 (m, 2H), 2.07-2.24 (m, 2H),
2.44 (s, 3H), 3.01-3.13 (m, 4H), 3.73 (s, 3H), 3.76 (s, 3H), 4.09
(q, 2H, J=7.0), 4.24 (s, 2H), 4.30 (t, 1H, J=7.4), 4.45-4.48 (m,
1H), 5.12 (t, 1H, J=6.3), 5.59 (d, 1H, J=15.8), 6.45 (dd, 1H,
J=8.5, 2.2), 6.54-6.56 (m, 2H), 6.73 (dd, 1H, J=15.5, 4.8),
6.89-6.95 (m, 3H), 7.20 (d, 1H, J=8.5), 7.22 (d, 1H, J=8.5), 8.08
(d, 1H, J=8.8), 8.89 (d, 1H, J=7.4).
Preparation of Product
Ethyl-3-{(5'-Methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-((S)-Pyrr-
ol-Ala)}-E-Propenoate
[0282]
Ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-((-
N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate (0.25 g, 0.33
mmol) was dissolved in CHCl.sub.3 (6 mL). Two drops of water were
added, followed by DDQ (0.10 g, 0.43 mmol). The reaction was heated
to 50-55.degree. C. for 8 h. Upon cooling, the reaction mixture was
poured into EtOAc (30 mL). The organic phase was washed with 30 mL
of 2:1 NaHCO.sub.3/1N NaOH solution and then brine (30 mL), dried
(MgSO.sub.4), and concentrated. The residue was subjected to flash
column chromatography eluting with 0.fwdarw.2%
MeOH/CH.sub.2Cl.sub.2. The
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-O-L-(4-F-Phe)-L-((S)-Pyrr-
ol-Ala)}-E-propenoate product was obtained as 0.18 g (90%) of a
white amorphous solid: R.sub.f=0.09 (3% MeOH/CHCl.sub.3); IR(KBr)
1680, 1549 cm.sup.-1; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.91 (d,
3H, J=6.6), 0.95 (d, 3H, J=6.6), 1.20 (t, 3H, J=7.0), 1.36-1.44 (m,
1H), 1.56-1.61 (m, 1H), 1.74-1.82 (m, 1H), 1.94-1.99 (m, 2H),
2.18-2.25 (m, 1H), 2.45 (s, 3H), 2.98-3.18 (m, 4H), 4.09 (q, 2H,
J=7.0), 4.28-4.32 (m, 1H), 4.43-4.46 (m, 1H), 5.12 (m, 1H), 5.58
(d, 1H, J=15.8), 6.56(s, 1H), 6.72 (dd, 1H, J=15.8, 4.8), 6.91-6.97
(m, 2H), 7.21 (d, 1H, J=5.9), 7.23 (d, 1H, J=5.9), 7.59 (s, 1H),
8.06 (d, 1H, J=8.8), 8.92 (d, 1H, J=7.4); Anal.
(C.sub.30H.sub.37N.sub.4O.sub.8F) C, H, N.
Example 19
Preparation of Compound B-3:
2-{(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L--
((S)-Pyrrol-Ala)}-E-(.alpha.-Vinyl-.gamma.-Butyrolactone)
[0283] ##STR123##
Preparation of Intermediate
Boc-L-{(N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala}-E-(.alpha.-Vinyl-.gamma.-B-
utyrolactone)
[0284]
(1S,3'S)-{2-(1'-(2'',4''-Dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-
-1-hydroxymethylethyl}-carbamic acid tert-butyl ester (0.360 g,
0.881 mmol, 1 equiv.) was oxidized to the corresponding aldehyde in
the manner described in the preparation of
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate
(Example 6). This aldehyde was combined with
3-(triphenyl-.lamda..sup.5-phosphanylidene)-dihydrofuran-2-one
(prepared in a manner analogous to that described in Baldwin et
al., J. Org. Chem. 1971, vol. 36, 1441) (0.320 g, 0.924 mmol, 1.05
equiv.) in a mixture of ethylene glycol dimethyl ether (10 mL) and
DMF (2 mL). The reaction mixture was warmed in a 100.degree. C. oil
bath for 1.5 h, allowed to cool to 23.degree. C. overnight, and
then diluted with MTBE (200 mL), washed with water (20 mL) and
brine (20 mL), dried over Na.sub.2SO.sub.4 and evaporated. The
residue was purified by flash column chromatography (2.5%
CH.sub.3OH in CH.sub.2Cl.sub.2, then 67% EtOAc in CH.sub.2Cl.sub.2)
to give
Boc-L-{(N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala}-E-(.alpha.-vinyl-.-
gamma.-butyrolactone) as an oil (0.250 g, 60%): R.sub.f=0.50 (67%
EtOAc in CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3307, 1754, 1678;
.sup.1H NMR (CDCl.sub.3) .delta. 1.42 (s, 9H), 1.46-1.68 (m, 214),
2.02-2.13 (m, 1H), 2.18-2.30 (m, 1H), 2.44-2.56 (m, 1H), 2.91-3.04
(m, 1H), 3.15-3.27 (m, 3H), 3.80 (s, 6H), 4.34-4.43 (m, 5H),
5.63-5.69 (m, 1H), 6.42-6.47 (m, 2H), 6.48-6.53 (m, 1H), 7.09-7.13
(m, 1H).
Preparation of Product
2-{(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L--
((S)-Pyrrol-Ala)}-E-(.alpha.-Vinyl-.gamma.-Butyrolactone)
[0285]
Boc-L-{(N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala}-E-(.alpha.-vinyl-.g-
amma.-butyrolactone) was converted to product
2-{(5'-methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-Phe)-L--
((S)-Pyrrol-Ala)}-E-(.alpha.-vinyl-.gamma.-butyrolactone) in a
manner analogous to that described above for the preparation of
product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-((S)-Pyrrol-Ala)}-E-propenoate (Example 17): R.sub.f=0.28 (5%
CH.sub.3OH in CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3378 br, 1749, 1678
br; .sup.1H NMR (CDCl.sub.3) .delta. 0.84 (d, 3H, J=6.8), 1.03 (d,
3H, J=6.8), 1.45-1.55 (m, 1H), 1.75-2.00 (m, 2H), 2.25-2.43 (m,
2H), 2.46-2.59 (m, 2H), 2.48 (s, 3H), 2.63-2.72 (m, 1H), 2.77-2.90
(m, 3H), 3.06-3.26 (m, 2H), 3.29-3.44 (m, 2H), 4.32-4.46 (m, 3H),
4.65-4.71 (m, 1H), 5.72 (s, 1H), 6.14-6.20 (m, 1H), 6.40 (s, 1H),
6.94-7.02 (m, 2H), 7.03-7.11 (m, 2H), 7.24 (d, 1H, J=9.0), 7.60 (d,
1H, J=6.2); Anal. (C.sub.31H.sub.37FN.sub.4O.sub.7) C, H, N.
Example 20
Preparation of Compound B-4:
Ethyl-3-[(5'-Methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-((S)-Piper-Ala)}-E-Propenoate
[0286] ##STR124##
[0287] This product was prepared in analogy to product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val-L-(4-F-Phe)-L-((S)-Piper--
Ala)}-E-propenoate (Example 9) and product
ethyl-3-{(5'-methylisoxazole-3'-carbonyl)-L-Val.PSI.(COCH.sub.2)-L-(4-F-P-
he)-L-Gln}-E-propenoate (Example 10) described above:
mp=161-162.degree. C.; R.sub.f=0.30 (5% CH.sub.3OH in
CH.sub.2Cl.sub.2); IR (cm.sup.-1) 3295, 1713, 1649; .sup.1H NMR
(CDCl.sub.3) .delta. 0.84 (d, 3H, J=6.8), 1.03 (d, 3H, J=6.8), 1.30
(t, 3H, J=7.2), 1.43-1.55 (m, 2H), 1.77-1.90 (m, 2H), 1.95-2.12 (m,
2H), 2.27-2.38 (m, 1H), 2.44-2.58 (m, 2H), 2.48 (s, 3H), 2.66-2.76
(m, 1H), 2.80-2.93 (m, 2H), 3.12-3.42 (m, 3H), 4.18 (q, 2H, J=7.2),
4.39-4.49 (m, 1H), 4.65-4.72 (m, 1H), 5.50 (dd, 1H, J=15.9, 1.6),
5.80 (s, 1H), 6.38-6.41 (m, 1H), 6.62 (dd, 1H, J=15.9, 5.3),
6.94-7.02 (m, 2H), 7.08-7.28 (m, 4H).
Examples 21 through 30
[0288] For Examples 21-30, the following Compounds (A-14) through
(A-23), respectively, were prepared using synthetic methods
analogous to those described above for compounds of the formula
I-A: ##STR125## ##STR126## ##STR127##
Example 31
Preparation of Comparison Compound #2:
Ethyl-3-(Cbz-L-Leu-L-Phe-L-Gln)-E-Propenoate
[0289] ##STR128##
Preparation of Intermediate [Boc-L-(Tr-Gln)]-N(OMe)Me
[0290] Isobutyl chloroformate (4.77 mL, 36.8 mmol, 1.0 equiv.) was
added to a solution of N-.alpha.-Boc-.gamma.-trityl-L-glutamine
(18.7 g, 36.7 mmol, 1 equiv.) and 4-methylmorpholine (8.08 mL, 73.5
mmol, 2.0 equiv.) in CH.sub.2Cl.sub.2 (250 mL) at 0.degree. C. The
reaction mixture was stirred at 0.degree. C. for 20 minutes, and
then N,O-dimethylhydroxylamine hydrochloride (3.60 g, 36.7 mmol,
1.0 equiv.) was added. The resulting solution was stirred at
0.degree. C. for 20 minutes and at 23.degree. C. for 2 hours, and
then was partitioned between water (150 mL) and CH.sub.2Cl.sub.2
(2.times.150 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and were concentrated. Purification of the residue
by flash column chromatography (gradient elution 40.fwdarw.20%
hexanes in EtOAc) provided {Boc-L-(Tr-Gln)}-N(OMe)Me (16.1 g, 82%)
as a white foam: IR (cm.sup.-1) 3411, 3329, 3062, 1701, 1659;
.sup.1H NMR (CDCl.sub.3) .delta. 1.42 (s, 9H), 1.63-1.77 (m, 1H),
2.06-2.17 (m, 1H), 2.29-2.43 (m, 2H), 3.17 (s, 3H), 3.64 (s, 3H),
4.73 (s, br, 1H), 5.38-5.41 (m, 1H), 7.20-7.31 (m, 15H); Anal.
(C.sub.31H.sub.37N.sub.3O.sub.5) C, H, N.
Preparation of Intermediate {Boc-L-(Tr-Gln)}-H
[0291] Diisobutylaluminum hydride (50.5 mL of a 1.5 M solution in
toluene, 75.8 mmol, 2.5 equiv.) was added to a solution of
{Boc-L-(Tr-Gln)}-N(OMe)Me (16.1 g, 30.3 mmol, 1 equiv.) in THF at
-78.degree. C., and the reaction mixture was stirred at -78.degree.
C. for 4 h. Methanol (4 mL) and 1.0 M HCl (10 mL) were added
sequentially, and the mixture was warmed to 23.degree. C. The
resulting suspension was diluted with Et.sub.2O (150 mL) and was
washed with 1.0 M HCl (3.times.100 mL), half-saturated NaHCO.sub.3
(100 mL), and water (100 mL). The organic layer was dried over
MgSO.sub.4, filtered, and was concentrated to give crude
{Boc-L-(Tr-Gln)}-H (13.8 g, 97%) as a white solid:
mp=114-116.degree. C.; IR (cm.sup.-1) 3313, 1697, 1494; .sup.1H NMR
(CDCl.sub.3) .delta. 1.44 (s, 9H), 1.65-1.75 (m, 1H), 2.17-2.23 (m,
1H), 2.31-2.54 (m, 2H), 4.11 (s, br, 1H), 5.38-5.40 (m, 1H), 7.11
(s, 1H), 7.16-7.36 (m, 1SH), 9.45 (s, 1H).
Preparation of Intermediate
Ethyl-3-{Boc-L-(Tr-Gln)}-E-Propenoate
[0292] Sodium bis(trimethylsilyl)amide (22.9 mL of a 1.0 M solution
in THF, 22.9 mmol, 1.0 equiv.) was added to a solution of triethyl
phosphonoacetate (5.59 g, 22.9 mmol, 1.0 equiv.) in THF (200 mL) at
-78.degree. C., and the resulting solution was stirred for 20
minutes at that temperature. Crude {Boc-L-(Tr-Gln)}-H (10.8 g, 22.9
mmol, 1 equiv.) in THF (50 mL) was added via cannula, and the
reaction mixture was stirred for 2 hours at -78.degree. C., warmed
to 0.degree. C. for 10 minutes, and partitioned between 0.5 M HCl
(150 mL) and a 1:1 mixture of EtOAc and hexanes (2.times.150 mL).
The combined organic layers were dried over Na.sub.2SO.sub.4 and
were concentrated. Purification of the residue by flash column
chromatography (40% EtOAc in hexanes) provided
ethyl-3-{Boc-L-(Tr-Gln)}-E-propenoate (10.9 g, 88%) as a white
foam: IR (cm.sup.-1) 3321, 1710; .sup.1H NMR (CDCl.sub.3) .delta.
1.27 (t, 3H, J=7.2), 1.42 (s, 9H), 1.70-1.78 (m, 1H), 1.80-1.96 (m,
1H), 2.35 (t, 2H, J=7.0), 4.18 (q, 2H, J=7.2), 4.29 (s, br, 1H),
4.82-4.84 (m, 1H), 5.88 (dd, 1H, J=15.7, 1.6), 6.79 (dd, 1H,
J=15.7, 5.3), 6.92 (s, 1H), 7.19-7.34 (m, 15H); Anal.
(C.sub.33H.sub.38N.sub.2O.sub.5) C, H, N.
Preparation of Intermediate
Ethyl-3-{Cbz-L-Leu-L-Phe-L-(Tr-Gln)}-E-Propenoate
[0293] A solution of HCl in 1,4-dioxane (4.0 M, 20 mL), was added
to a solution of ethyl-3-{Boc-L-(Tr-Gln)}-E-propenoate (1.00 g,
1.84 mmol, 1 equiv.) in the same solvent (20 mL) at 23.degree. C.
After 3 hours, the volatiles were removed under reduced pressure.
The residue was dissolved in CH.sub.2Cl.sub.2 (50 mL) and
Cbz-L-Leu-L-Phe-OH (0.759 g, 1.84 mmol, 1.0 equiv.), HOBt (0.373 g,
2.76 mmol, 1.5 equiv.), 4-methylmorpholine (0.809 mL, 7.36 mmol,
4.0 equiv.), and EDC (0.529 g, 2.76 mmol, 1.5 equiv.) were added
sequentially. The reaction mixture was stirred at 23.degree. C. for
18 hours, and then was partitioned between water (150 mL) and EtOAc
(2.times.150 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and were concentrated. Flash chromatographic
purification of the residue (5% CH.sub.3OH in CH.sub.2Cl.sub.2)
afforded ethyl-3-{Cbz-L-Leu-L-Phe-L-(Tr-Gln)}-E-propenoate (1.25 g,
83%) as a white solid: mp=192-194.degree. C.; IR (cm.sup.-1) 3295,
1696, 1678, 1655, 1519; .sup.1H NMR (CDCl.sub.3) .delta. 0.84 (d,
3H, J=6.5), 0.86 (d, 3H, J=6.5), 1.24-1.32 (m, 1H), 1.28 (t, 3H,
J=7.2), 1.43-1.75 (m, 3H), 1.91-2.06 (m, 1H), 2.20-2.38 (m, 2H),
2.93-3.02 (m, 1H), 3.07-3.18 (m, 1H), 3.95-4.02 (m, 1H), 4.17 (q,
2H, J=7.2), 4.43-4.55 (m, 2H), 4.82-4.95 (m, 2H), 5.69 (d, 1H,
J=15.7), 6.46 (d, 1H, J=7.5), 6.60 (d, 1H, J=8.1), 6.69 (dd, 1H,
J=15.7, 5.1), 7.09-7.38 (m, 27H); Anal.
(C.sub.51H.sub.56N.sub.4O.sub.7) C, H, N.
Preparation of Product
Ethyl-3-(Cbz-L-Leu-L-Phe-L-Gln)-E-Propenoate
[0294] Trifluoroacetic acid (20 mL) was added to a solution of
ethyl-3-{Cbz-L-Leu-L-Phe-L-(Tr-Gln)}-E-propenoate (1.25 g, 1.49
mmol, 1 equiv.) and triisopropylsilane (1.53 mL, 7.47 mmol, 5.0
equiv.) in CH.sub.2Cl.sub.2 (20 mL) at 23.degree. C., producing a
bright yellow solution. The reaction mixture was stirred for 20
minutes at 23.degree. C., during which time it became colorless.
Carbon tetrachloride (20 mL) was added, and the volatiles were
removed under reduced pressure. The residue was triturated with
Et.sub.2O (20 mL), and the resulting white solid was collected by
vacuum filtration, washed with Et.sub.2O (3.times.50 mL), and
air-dried to afford ethyl-3-(Cbz-L-Leu-L-Phe-L-Gln)-E-propenoate
(0.717 g, 81%): mp=219-221.degree. C.; IR (cm.sup.-1) 3300, 1672,
1535; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.78 (d, 3H, J=6.8), 0.82
(d, 3H, J=6.5), 1.21 (t, 3H, J=7.0), 1.25-1.37 (m, 2H), 1.42-1.54
(m, 1H), 1.58-1.80 (m, 2H), 2.02-2.09 (m, 2H), 2.84 (dd, 1H,
J=13.2, 8.9), 2.97 (dd, 1H, J=13.2, 5.8), 3.93-4.01 (m, 1H), 4.11
(q, 2H, J=7.0), 4.33-4.52 (m, 2H), 4.97 (d, 1H, J=12.3), 5.04 (d,
1H, J=12.3), 5.64 (d, 1H, J=15.9), 6.69 (dd, 1H, J=15.9, 5.4), 6.76
(s, 1H), 7.13-7.37 (m, 1H), 7.43 (d, 1H, J=7.8), 7.99 (d, 1H,
J=8.1), 8.04 (d, 1H, J=8.1); Anal. (C.sub.32H.sub.2N.sub.4O.sub.7)
C, H, N.
Example 32
Preparation of Comparison Compound #3:
Ethyl-3-(Cbz-L-Val-L-Phe-L-Gln)-E-Propenoate
[0295] ##STR129##
Preparation of Intermediate
Ethyl-3-{Boc-L-Phe-L-(Tr-Gln)}-E-Propenoate
[0296] A solution of HCl in 1,4-dioxane (4.0 M, 10 mL) was added to
a solution of ethyl-3-{Boc-L-(Tr-Gln)}-E-propenoate (prepared as
described in Example 31, 3.05 g, 5.62 mmol, 1 equiv.) in the same
solvent (20 mL) at 23.degree. C. After 3 hours, the volatiles were
removed under reduced pressure. The residue was dissolved in
CH.sub.2Cl.sub.2 (50 mL), and Boc-L-Phe-OH (1.49 g, 5.62 mmol, 1.0
equiv.), HOBt (0.911 g, 6.74 mmol, 1.2 equiv.), 4-methylmorpholine
(1.85 mL, 16.8 mmol, 3.0 equiv.) and EDC (1.29 g, 6.73 mmol, 1.2
equiv.) were added sequentially. The reaction mixture was stirred
at 23.degree. C. for 18 hours, and was then partitioned between
water (150 mL) and EtOAc (2.times.150 mL). The combined organic
layers were dried over Na.sub.2SO.sub.4 and were concentrated.
Flash chromatographic purification of the residue (gradient
elution, 40-50% EtOAc in hexanes) afforded
ethyl-3-{Boc-L-Phe-L-(Tr-Gln)}-E-propenoate (2.77 g, 71%) as a
white foam: IR (cm.sup.-1) 3306, 1706, 1661; .sup.1H NMR
(CDCl.sub.3) .delta. 1.29 (t, 3H, J=7.2), 1.38 (s, 9H), 1.65-1.76
(m, 1H), 1.87-1.99 (m, 1H), 2.25-2.27 (m, 2H), 2.94-3.01 (m, 2H),
4.14-4.26 (m, 3H), 4.48-4.53 (m, 1H), 4.95 (s, br, 1H), 5.64 (d,
1H, J=15.8), 6.29 (d, 1H, J=8.1), 6.64 (dd, 1H, J=15.8, 5.4), 6.80
(s, br, 1H), 7.14-7.32 (m, 20H); Anal.
(C.sub.42H.sub.47N.sub.3O.sub.6) C, H, N.
Preparation of Intermediate
Ethyl-3-{Cbz-L-Val-L-Phe-L-(Tr-Gln)}-E-Propenoate
[0297] A solution of HCl in 1,4-dioxane (4.0 M, 8 mL) was added to
a solution of ethyl-3-{Boc-L-Phe-L-(Tr-Gln)}-E-propenoate (0.296 g,
0.429 mmol, 1 equiv.) in the same solvent (10 mL) at 23.degree. C.
After 3 hours, the volatiles were removed under reduced pressure.
The residue was dissolved in CH.sub.2Cl.sub.2 (10 mL), and then
Cbz-L-Val-OH (0.108 g, 0.430 mmol, 1.0 equiv.), HOBt (0.070 g,
0.518 mmol, 1.2 equiv.), 4-methylmorpholine (0.142 mL, 1.29 mmol,
3.0 equiv.) and EDC (0.099 g, 0.516 mmol, 1.2 equiv.) were added
sequentially. The reaction mixture was stirred at 23.degree. C. for
4 hours, and then was partitioned between water (100 mL) and EtOAc
(2.times.100 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and were concentrated. Flash chromatographic
purification of the residue (3% CH.sub.3OH in CH.sub.2Cl.sub.2)
afforded ethyl-3-{Cbz-L-Val-L-Phe-L-(Tr-Gln)}-E-propenoate (0.220
g, 62%) as a white solid: mp=195-198.degree. C.; IR (cm.sup.-1)
3284, 1689, 1646; .sup.1H NMR (CDCl.sub.3) .delta. 0.69 (d, 3H,
J=6.9), 0.82 (d, 3H, J=6.5), 1.28 (t, 3H, J=7.2), 1.63-1.74 (m,
1H), 1.96-2.02 (m, 2H), 2.22-2.35 (m, 2H), 2.93 (dd, 1H, J=14.0,
7.6), 3.10 (dd, 1H, J=14.0, 6.7), 3.81-3.85 (m, 1H), 4.17 (q, 2H,
J=7.2), 4.48-4.58 (m, 2H), 4.87 (d, 1H, J=12.0), 4.94 (d, 1H,
J=12.0), 5.06 (d, 1H, J=6.9), 5.67 (d, 1H, J=15.6), 6.43 (d, 1H,
J=7.5), 6.63-6.72 (m, 2H), 7.10-7.40 (m, 26H); Anal.
(C.sub.50H.sub.54N.sub.4O.sub.7) C, H, N.
Preparation of Product
Ethyl-3-(Cbz-L-Val-L-Phe-L-Gln)-E-Propenoate
[0298] Trifluoroacetic acid (5 mL) was added to a solution of
ethyl-3-{Cbz-L-Val-L-Phe-L-(Tr-Gln)}-E-propenoate (0.188 g, 0.229
mmol, 1 equiv.) and triisopropylsilane (0.300 mL, 1.46 mmol, 6.4
equiv.) in CH.sub.2Cl.sub.2 (10 mL) at 23.degree. C., producing a
bright yellow solution. The reaction mixture was stirred for 20
min. at 23.degree. C., during which time it became colorless.
Carbon tetrachloride (10 mL) was added, and the volatiles were
removed under reduced pressure. The residue was triturated with
Et.sub.2O (20 mL), and the resulting white solid was collected by
vacuum filtration, washed with Et.sub.2O (3.times.50 mL), and
air-dried to afford ethyl-3-(Cbz-L-Val-L-Phe-L-Gln)-E-propenoate
(0.094 g, 71%): mp=240.degree. C. (dec); IR (cm.sup.-1) 3263, 1686,
1640; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.73 (d, 6H, J=6.9), 1.21
(t, 3H, J=7.2), 1.60-1.75 (m, 2H), 1.83-1.90 (m, 1H), 2.03-2.08 (m,
2H), 2.83 (dd, 1H, J=13.6, 8.6), 2.96 (dd, 1H, J=13.6, 6.2),
3.79-3.84 (m, 1H), 4.10 (q, 2H, J=7.2), 4.37-4.49 (m, 1H),
4.51-4.56 (m, 1H), 4.99 (d, 1H, J=12.5), 5.06 (d, 1H, J=12.5), 5.61
(d, 1H, J=15.5), 6.67 (dd, 1H, J=15.5, 5.5), 6.76 (s, 1H),
7.13-7.36 (m, 12H), 8.06 (d, 2H, J=8.1); Anal.
(C.sub.31H.sub.40N.sub.4O.sub.7) C, H, N.
Example 33
Preparation of Compound (A-24):
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0299] ##STR130##
Preparation of Intermediate
Ethyl-3-{Cbz-L-Leu-L-Phe-L-(N-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-Pro-
penoate
[0300] A solution of HCl in 1,4-dioxane (4.0 M, 4 mL) was added to
a solution of
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-pyrrol-Ala)}-E-propenoate
(prepared as described in Example 6) (0.139 g, 0.292 mmol, 1
equiv.) in 1,4-dioxane (4 mL). After stirring 1.5 h, the volatiles
were evaporated to give the crude amine salt, which was used
without purification.
[0301] This amine salt was dissolved in CH.sub.2Cl.sub.2 (7 mL),
and then Cbz-L-Leu-L-Phe-OH (0.156 g, 0.378 mmol, 1.3 equiv.),
4-methylmorpholine (0.128 mL, 1.16 mmol, 4 equiv.), HOBt (0.067 g,
0.50 mmol, 1.7 equiv.) and EDC (0.095 g, 0.50 mmol, 1.7 equiv) were
added sequentially. After stirring 20 hours, the reaction mixture
was poured into brine (15 mL) and extracted with 10% CH.sub.3OH in
CH.sub.2Cl.sub.2 (3.times.25 mL). The combined organic phases were
dried over Na.sub.2SO.sub.4 and evaporated. Purification of the
residue by flash column chromatography (60% EtOAc in hexanes)
provided
ethyl-3-{Cbz-L-Leu-L-Phe-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-pr-
openoate (0.158 g, 70%) as a foam: .sup.1H NMR (CDCl.sub.3) .delta.
0.87-0.92 (m, 6H), 1.28 (t, 3H, J=7.2), 1.46-1.68 (m, 5H),
1.74-1.86 (m, 1H), 1.97-2.19 (m, 2H), 3.02 (dd, 1H, J=13.7, 5.6),
3.11-3.24 (m, 3H), 3.78 (s, 3H), 3.79 (s, 3H), 4.17 (q, 2H, J=7.2),
4.20-4.30 (m, 2H), 4.35-4.45 (m, 2H), 4.82-4.90 (m, 1H), 5.07 (d,
1H, J=12.3), 5.13 (d, 1H, J=12.3), 5.36 (d, 1H, J=7.8), 5.82 (dd,
1H, J=15.6, 1.2), 6.42-6.46 (m, 2H), 6.72 (dd, 1H, J=15.6, 5.3),
6.88 (d, 1H, J=8.7), 7.09 (d, 1H, J=9.0), 7.13-7.20 (m, 5H),
7.29-7.37 (m, 5H), 8.09 (d, 1H, J=6.5).
Preparation of Product
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0302]
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala-
)}-E-propenoate (0.156 g, 0.202 mmol, 1 equiv.) and ammonium
cerium(IV) nitrate (0.277 g, 0.505 mmol, 2,5 equiv.) were combined
in a mixture of THF/water 2:1 (3 mL) and stirred 2 hours. The
reaction mixture was poured into brine (15 mL) and extracted with
10% CH.sub.3OH in CH.sub.2Cl.sub.2 (3.times.25 mL). The combined
organic phases were dried over Na.sub.2SO.sub.4 and evaporated.
Purification of the residue by flash column chromatography
(gradient elution, 2-5% CH.sub.3OH in CH.sub.2Cl.sub.2) provided
ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Pyrrol-Ala)}-E-propenoate (0.059 g,
47%) as an off-white solid: .sup.1H NMR (CDCl.sub.3) .delta.
0.85-0.92 (m, 6H), 1.28 (t, 3H, J=7.2), 1.39-1.65 (m, 4H),
1.68-1.93 (m, 2H), 2.08-2.20 (m, 1H), 2.27-2.38 (m, 1H), 3.02-3.13
(m, 2H), 3.24-3.32 (m, 2H), 4.11-4.20 (m, 1H), 4.18 (q, 2H, J=7.2),
4.47-4.58 (m, 1H), 4.81-4.89 (m, 1H), 5.05 (d, 1H, J=12.1), 5.12
(d, 1H, J=12.1), 5.26 (d, 1H, J=8.1), 5.78 (dd, 1H, J=15.7, 1.2),
6.23 (s, 1H), 6.72 (dd, 1H, J=15.7, 5.3), 7.13-7.25 (m, 6H),
7.30-7.37 (m, 5H), 7.54 (d, 1H, J=7.2).
Example 34
Preparation of Compound (A-25):
Ethyl-3-{Cbz-L-Val-L-Phe-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0303] ##STR131##
Preparation of Intermediate
Ethyl-3-{Cbz-L-Val-L-Phe-L-((V-2,4-Dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-Pr-
openoate
[0304] In a manner analogous to that used for the conversion of
ethyl-3-{Boc-L-(Tr-Gln)}-E-propenoate to
ethyl-3-{Cbz-L-Leu-L-Phe-L-(Tr-Gin)}-E-propenoate described in
Example 31,
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate
(prepared as in Example 33) was coupled with Cbz-L-Val-L-Phe-OH to
afford
ethyl-3-{Cbz-L-Val-L-Phe-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-pr-
openoate: IR (cm.sup.1) 3288, 1699, 1652; .sup.1H NMR (CDCl.sub.3)
.delta. 0.87 (d, 3H, J=6.8), 0.95 (d, 3H, J=6.5), 1.28 (t, 3H,
J=7.2), 1.48-1.60 (m, 2H), 1.70-1.84 (m, 1H), 1.95-2.20 (m, 3H),
3.01 (dd, 1H, J=13.4, 5.6), 3.09-3.25 (m, 3H), 3.78 (s, 3H), 3.80
(s, 3H), 4.03-4.10 (m, 1H), 4.17 (q, 2H, J=7.2), 4.24 (d, 1H,
J=14.2), 4.33-4.44 (m, 1H), 4.38 (d, 1H, J=14.2), 4.85-4.94 (m,
1H), 5.08 (d, 1H, J=12.1), 5.14 (d, 1H, J=12.1), 5.39 (d, 1H,
J=8.1), 5.80 (dd, 1H, J=15.6, 1.2), 6.42-6.47 (m, 2H), 6.70 (dd,
1H, J=15.6, 5.3), 6.81 (d, 1H, J=9.0), 7.11-7.20 (m, 6H), 7.31-7.39
(m, 5H), 8.11 (d, 1H, J=6.2); Anal.
(C.sub.42H.sub.52N.sub.4O.sub.9) C, H, N.
Preparation of Product
Ethyl-3-{Cbz-L-Val-L-Phe-L-((S)-Pyrrol-Ala)}-E-Propenoate
[0305] A suspension of
ethyl-3-{Cbz-L-Val-L-Phe-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-pr-
openoate (0.215 g, 0.284 mmol, 1 equiv.), DDQ (0.071 g. 0.31 mmol,
1.1 equiv.) and water (3 drops) in CHCl.sub.3 (4 mL) was stirred 1
h at 23.degree. C., and was then warmed to reflux for 6 h. After
cooling overnight, more DDQ (0.019 g, 0.084 mmol, 0.3 equiv.) was
added, and the mixture was warmed to 67.degree. C. for 1 h and then
evaporated. Purification of the residue by flash column
chromatography (gradient elution, 2-5% CH.sub.3OH in
CH.sub.2Cl.sub.2) provided slightly impure material, which was
dissolved in CH.sub.2Cl.sub.2 (70 mL) and washed with saturated
NaHCO.sub.3 (2.times.30 mL) and brine (30 mL), and then dried over
Na.sub.2SO.sub.4 and evaporated. The residue was stirred in
Et.sub.2O (10 mL) for 20 minutes, and the solid was collected by
filtration and dried under vacuum to provide
ethyl-3-{Cbz-L-Val-L-Phe-L-((S)-Pyrrol-Ala)}-E-propenoate (0.060 g,
35%) as a an off-white solid: mp=215-217.degree. C.; IR (cm.sup.-1)
3413, 3295, 1696, 1649; .sup.1H NMR (CDCl.sub.3) .delta. 0.83 (d,
3H, J=6.5), 0.91 (d, 3H, J=6.8), 1.28 (t, 3H, J=7.2), 1.50-1.59 (m,
1H), 1.70-1.91 (m, 2H), 2.03-2.17 (m, 2H), 2.26-2.38 (m, 1H), 3.03
(dd, 1H, J=13.5, 6.4), 3.12 (dd, 1H, J=13.5, 6.4), 3.21-3.34 (m,
2H), 3.96 (dd, 1H, J=8.3, 6.4), 4.17 (q, 2H, J=7.2), 4.45-4.56 (m,
1H), 4.83-4.92 (m, 1H), 5.07 (d, 1H, J=12.1), 5.13 (d, 1H, J=12.1),
5.29 (d, 1H, J=8.3), 5.77 (dd, 1H, J=15.8, 1.2), 5.94 (s, 1H), 6.71
(dd, 1H, J=15.8, 5.3), 6.95 (d, 1H, J=9.0), 7.14-7.27 (m, 5H),
7.31-7.38 (m, 5H), 7.57 (d, 1H, J=7.2); Anal.
(C.sub.33H.sub.42N.sub.4O.sub.7) C, H, N.
Example 35
Preparation of Compound (A-26):
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Piper-Ala)}-E-Provenoate
[0306] ##STR132##
Preparation of Product
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Piper-Ala)}-E-Propenoate
[0307]
(1S,3'S)-{2-(1'-(2'',4''-Dimethoxybenzyl)-2'-oxo-piperidin-3'-yl)--
1-hydroxy-methylethyl}-carbamic acid tert-butyl ester (prepared as
described in Example 8) was converted to the product
ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Piper-Ala)}-E-propenoate in a
manner analogous to the conversion of
(1S,3'S)-{2-(1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydr-
oxymethyl-ethyl}-carbamic acid tert-butyl ester to product
ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Pyrrol-Ala)}-E-propenoate as
described in Example 34: IR (cm.sup.-1) 3422, 3307, 1713, 1649;
.sup.1H NMR (CDCl.sub.3) .delta. 0.86-0.92 (m, 6H), 1.28 (t, 3H,
J=7.2), 1.38-1.75 (m, 6H), 1.77-1.89 (m, 1H), 1.96-2.11 (m, 2H),
3.07 (d, 2H, J=6.2), 3.20-3.27 (m, 2H), 4.13-4.24 (m, 1H), 4.18 (q,
2H, J=7.2), 4.41-4.53 (m, 1H), 4.76-4.85 (m, 1H), 5.06 (d, 1H,
J=12.1), 5.12 (d, 1H, J=12.1), 5.34 (d, 1H, J=7.8), 5.78 (dd, 1H,
J=15.6, 5.4), 6.17 (s, 1H), 6.70 (dd, 1H, J=15.6, 5.4), 7.00 (d,
1H, J=8.4), 7.13-7.27 (m, 6H), 7.30-7.37 (m, 5H), 7.83 (d, 1H,
J=6.8); Anal. (C.sub.35H.sub.46N.sub.407-0.5H.sub.2O)C, H, N.
Example 36
Preparation of Compound (A-27):
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((R)-Pyrrol-Ala)}-E-Propenoate
[0308] ##STR133##
Preparation of Intermediate
(4S,4''R)-4-{3'-(4''-Benzyl-2''-oxo-oxazolidin-3''-yl)-3'-oxopropyl}-2,2--
dimethyloxazolidine-3-carboxylic Acid tert-Butyl Ester
[0309] Triethylamine (6.43 mL, 46.1 mmol, 3.0 equiv.) and pivaloyl
chloride (1.89 mL, 15.3 mmol, 1.0 equiv.) were added sequentially
to a solution of
(4S)-4-(2'-carboxyethyl)-2,2-dimethyloxazolidine-3-carboxylic acid
tert-butyl ester (4.20 g, 15.3 mmol, 1 equiv.) in THF (300 mL) at
0.degree. C. The cloudy reaction mixture was stirred at 0.degree.
C. for 3.5 h, and then lithium chloride (0.716 g, 16.9 mmol, 1.1
equiv.) and (R)-(+)-4-benzyl-2-oxazolidinone (2.59 g, 14.6 mmol,
0.95 equiv.) were added sequentially. After warming to 23.degree.
C. and stirring for 19 h, the reaction mixture was partitioned
between 0.5 M HCl (150 mL) and EtOAc (2.times.150 mL). The combined
organic layers were washed with half-saturated Na.sub.2CO.sub.3
(150 mL), dried over MgSO.sub.4, and gravity filtered. The filtrate
was concentrated under reduced pressure and the residue was
purified by flash column chromatography (30% EtOAc in hexanes) to
give
(4S,4''R)-4-{3'-(4''-benzyl-2''-oxo-oxazolidin-3''-yl)-3'-oxopropyl}-2,2--
dimethyloxazolidine-3-carboxylic acid tert-butyl ester (6.15 g,
97%) as a colorless oil: IR (cm.sup.-1) 2978, 1783, 1694; .sup.1H
NMR (CDCl.sub.3, mixture of isomers) .delta. 1.46 (s), 1.58 (s),
1.63 (s), 2.01-2.05 (m), 2.72-3.13 (m), 3.29-3.33 (m), 3.74-3.79
(m), 3.82-4.09 (m), 4.11-4.25 (m), 4.67-4.70 (m), 7.20-7.37 (m);
Anal. (C.sub.23H.sub.32N.sub.2O.sub.6) C, H, N.
Preparation of Intermediate
(2'R,4S,4''R)-4-{2'-(4''-Benzyl-2''-oxo-oxazolidine-3''-carbonyl)-pent-4'-
-enyl}-2,2-dimethyloxazolidine-3-carboxylic Acid tert-Butyl
Ester
[0310] A solution of
(4S,4''R)-4-{3'-(4''-benzyl-2''-oxo-oxazolidin-3''-yl)-3'-oxopropyl}-2,2--
dimethyloxazolidine-3-carboxylic acid tert-butyl ester (6.15 g,
14.2 mmol, 1 equiv.) in THF (25 mL) was added to a solution of
sodium bis(trimethylsilyl)amide (14.2 mL of a 1.0 M solution in
THF, 14.2 mmol, 1.0 equiv.).in the same solvent (50 m]L) at
-78.degree. C. The reaction mixture was stirred for 15 minutes at
-78.degree. C., and then allyl iodide (3.90 mL, 42.6 mmol, 3.0
equiv.) was added. After stirring an additional 2 h at -78.degree.
C., the reaction mixture was maintained at -45.degree. C. for 2 h,
and then was partitioned between a 2:1 mixture of half-saturated
NH.sub.4Cl and 5% Na.sub.2S.sub.2O.sub.3 (200 mL) and a 1:1 mixture
of EtOAc and hexanes (2.times.150 mL). The combined organic layers
were washed with H.sub.2O (100 mL), dried over MgSO.sub.4, and
gravity filtered. The filtrate was concentrated under reduced
pressure and the residue was purified by flash column
chromatography (15% EtOAc in hexanes) to provide
(2'R,4S,4''R)-4-{2'-(4''-benzyl-2''
"-oxo-oxazolidine-3''-carbonyl)-pent-4'-enyl}-2,2-dimethyloxazolidine-3-c-
arboxylic acid tert-butyl ester (3.12 g, 46%) as a viscous foam: IR
(cm.sup.-1) 2978, 1782, 1685; .sup.1H NMR (CDCl.sub.3, mixture of
isomers) .delta. 1.42 (s), 1.45 (s), 1.49 (s), 1.52 (s), 1.62-1.78
(m), 1.80-2.01 (m), 2.23-2.49 (m), 2.51-2.56 (m), 2.76 (dd, J=13.3,
9.7), 3.26 (dd, J=13.3, 3.6), 3.58-3.64 (m), 3.67 (d, J=8.7),
3.90-3.98 (m), 4.02-4.15 (m), 4.16-4.30 (m), 4.75-4.82 (m),
5.06-5.11 (m), 5.74-5.88 (m), 7.22-7.36 (m); Anal.
(C.sub.26H.sub.36N.sub.2O.sub.6) C, H, N.
Preparation of Intermediate
(1S,3'R)-{2-(1'-(2'',4''-Dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydr-
oxymethylethyl}-carbamic Acid tert-Butyl Ester
[0311] Ozone was bubbled through a solution of
(2'R,4S,4''R)-4-{2'-(4''-benzyl-2''-oxo-oxazolidine-3''-carbonyl)-pent-4'-
-enyl}-2,2-dimethyloxazolidine-3-carboxylic acid tert-butyl ester
(3.12 g, 6.60 mmol, 1 equiv.) in CH.sub.2Cl.sub.2 (200 mL) and
CH.sub.3OH (0.535 mL, 13.2 mmol, 2.0 equiv.) at -78.degree. C.
until a blue color persisted. The reaction mixture was then purged
with argon until it became colorless. Methyl sulfide (4.85 mL, 66.0
mmol, 10 equiv.) was added, and the mixture was stirred at
-78.degree. C. for 3.5 h and then was maintained at 0.degree. C.
for an additional 1 h. After partitioning the reaction mixture
between H.sub.2O 150 mL) and a 1:1 mixture of EtOAc and hexanes
(2.times.150 mL), the combined organic layers were dried over
MgSO.sub.4 and gravity filtered. The filtrate was concentrated
under reduced pressure, and the residue was immediately utilized
without further purification.
[0312] The above residue was dissolved in a 2:1 mixture of THF, and
then EtOH (180 mL) at 23.degree. C. and 2,4-dimethoxybenzylamine
hydrochloride (5.38 g, 26.4 mmol, 4.0 equiv.), sodium acetate (2.17
g, 26.4 mmol, 4.0 equiv.), and sodium cyanoborohydride (0.829 g,
13.2 mmol, 2.0 equiv.) were added sequentially. The resulting
suspension was stirred for 19 h at 23.degree. C., and then was
partitioned between 0.5 M HCl (150 mL) and EtOAc (2.times.100 mL).
The combined organic layers were washed with half-saturated
NaHCO.sub.3 (100 mL), dried over Na.sub.2SO.sub.4, and concentrated
under reduced pressure. The residue was passed through a short
silica gel column (eluting with 50% EtOAc in hexanes) to give
(3'R,4S)-4-{1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-ylmethyl}-2-
,2-dimethyl-oxazolidine-3-carboxylic acid tert-butyl ester
contaminated with (R)-(+)-4-benzyl-2-oxazolidinone.
[0313] This material was dissolved in CH.sub.3OH (80 mL), and
TsOH.H.sub.2O (0.251 g, 1.32 mmol, 0.20 equiv.) was added. The
reaction mixture was heated to 50.degree. C. and was maintained at
that temperature for 4 h. After cooling to 23.degree. C., the
reaction mixture was concentrated under reduced pressure to -20 mL
volume and was partitioned between half-saturated NaHCO.sub.3 (150
mL) and a 9:1 mixture of CH.sub.2Cl.sub.2 and CH.sub.3OH
(2.times.150 mL). The combined organic layers were dried over
Na.sub.2SO.sub.4 and concentrated under reduced pressure.
Purification of the residue by flash column chromatography (3%
CH.sub.3OH in CH.sub.2Cl.sub.2) afforded
(1S,3'R)-{2-(1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydr-
oxymethylethyl}-carbamic acid tert-butyl ester (0.92 g, 34%) as a
foam: IR (cm.sup.-1) 3347 (br), 2937, 1669; .sup.1H NMR
(CDCl.sub.3) .delta. 1.44 (s, 9H), 1.62-1.77 (m, 2H), 1.94-2.04 (m,
1H), 2.15-2.26 (m, 1H), 2.40-2.50 (m, 1H), 3.13-3.24 (m, 2H),
3.56-3.77 (m, 3H), 3.80 (s, 6H), 3.82-4.16 (m, 1H), 4.37 (d, 1H,
J=14.3), 4.45 (d, 1H, J=14.3), 5.49 (d, 1H, J=7.8), 6.42-6.45 (m,
2H), 7.08-7.11 (m, 1H); Anal.
(C.sub.21H.sub.32N.sub.2O.sub.6.0.25H.sub.2O)C, H, N.
Preparation of Intermediate
Ethyl-3-{Boc-L-((N-2,4-Dimethoxybenzyl)-(R)-Pyrrol-Ala)}-E-Propenoate
[0314] In a manner analogous to the conversion of
(1S,3'S){2-1'-(2'',4''dethoxybenzyl)-2'-oxo-pyrrolidin-3'-yl)-1-hydroxyme-
thylethyl}-carbamic acid tert-butyl ester to
ethyl-3-{Boc-L-(N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-propenoate
described in Example 33,
(1S,3'R)-{2-1'-(2'',4''-dimethoxybenzyl)-2'-oxo-pyrolidin-3'-yl)-1-hydrox-
ymethylethyl}-carbamic acid tert-butyl ester was transformed into
ethyl-3-{Boc-L-(N-2,4-dimethoxybenzyl)-(R)-Pyrrol-Ala)}-E-propenoate:
IR (cm.sup.-1) 3307, 1713, 1674; .sup.1H NMR (CDCl.sub.3) .delta.
1.28 (t, 3H, J=7.2), 1.45 (s, 9H), 1.57-1.82 (m, 2H), 2.02-2.19 (m,
2H), 2.42-2.55 (m, 1H), 3.11-3.25 (m, 2H), 3.79 (s, 3H), 3.80 (s,
3H), 4.19 (q, 2H, J=7.2), 4.32-4.50 (m, 3H), 5.97 (dd, 1H, J=15.7,
1.4), 6.38 (d, 1H, J=7.8), 6.42-6.47 (m, 2H), 6.86 (dd, 1H, J=15.7,
5.1), 7.08-7.13 (m, 1H); Anal. (C.sub.25H.sub.36N.sub.2O.sub.7) C,
H, N.
Preparation of Intermediate
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((N-2,4-Dimethoxybenzyl)-(R)-Pyrrol-Ala)}-E-Pr-
openoate
[0315] A solution of HCl in 1,4-dioxane (4.0 M, 6 mL) was added to
a solution of
ethyl-3-{Boc-L-((N-2,4-dimethoxybenzyl)-(R)-Pyrrol-Ala)}-E-propenoate
(0.233 g, 0.489 mmol, 1 equiv.) in 1,4-dioxane (6 mL). After
stirring 1.5 h, the volatiles were evaporated to give the crude
amine salt, which was used without purification.
[0316] This amine salt was dissolved in DMF (4 mL) and cooled to
0.degree. C. Cbz-L-Leu-L-Phe-OH (0.202 g, 0.490 mmol, 1 equiv.),
DIEA (0.255 mL, 1.46 mmol, 3 equiv.) and HATU (0.186 g, 0.489 mmol,
1 equiv.) were added sequentially. After stirring 1.5 h, the
reaction mixture was diluted with MTBE (100 mL), washed with 5%
KHSO.sub.4, saturated NaHCO.sub.3 and brine (10 mL), dried over
MgSO.sub.4, and evaporated. Purification of the residue by flash
column chromatography (gradient elution, 60.fwdarw.75% EtOAc in
hexanes) provided
ethyl-3-{Cbz-L-Leu-L-Phe-L-((N-2,4-dimethoxybenzyl)-(R)-Pyrrol-Ala)}-E-pr-
openoate (0.208 g, 55%) as a white solid (after evaporation from
Et.sub.2O): mp=174-176.degree. C.; IR (cm.sup.-1) 3287, 1713, 1649;
.sup.1H NMR (CDCl.sub.3) .delta. 0.84-0.91 (m, 6H), 1.29 (t, 3H,
J=7.2), 1.42-1.66 (m, 4H), 1.67-1.77 (m, 1H), 1.84-1.95 (m, 1H),
2.20-2.12 (m, 1H), 2.33-2.45 (m, 1H), 3.04-3.23 (m, 4H), 3.78 (s,
3H), 3.79 (s, 3H), 4.15-4.29 (m, 1H), 4.17 (q, 2H, J=7.2), 4.31 (d,
1H, J=14.5), 4.40 (d, 1H, J=14.5), 4.67-4.84 (m, 2H), 5.05 (d, 1H,
J=12.1), 5.11 (d, 1H, J=12.1), 5.35 (d, 1H, J=8.1), 5.76 (dd, 1H,
J=15.6, 1.6), 6.42-6.46 (m, 2H), 6.74-6.81 (m, 1H), 6.75 (dd, 1H,
J=15.6, 5.0), 7.06-7.10 (m, 1H), 7.15-7.24 (m, 5H), 7.29-7.36 (m,
5H), 8.79 (d, 1H, J=5.9); Anal. (C.sub.43H.sub.54N.sub.4O.sub.9) C,
H, N.
Preparation of Product
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((R)-Pyrrol-Ala)}-E-Propenoate
[0317]
Ethyl-3-{Cbz-L-Leu-L-Phe-L-((N-2,4-dimethoxybenzyl)-(R)-Pyrrol-Ala-
)}-E-propenoate was converted to the product
ethyl-3-{Cbz-L-Leu-L-Phe-L-((R)-Pyrrol-Ala)}-E-propenoate in a
manner analogous to the conversion of
ethyl-3-{Cbz-L-Leu-L-Phe-L-((N-2,4-dimethoxybenzyl)-(S)-Pyrrol-Ala)}-E-pr-
openoate to product
ethyl-3-{Cbz-L-Leu-L-Phe-L-((S)-Pyrrol-Ala)}-E-propenoate described
in Example 34: IR (cm.sup.-1) 3290, 1702, 1642; .sup.1H NMR
(CDCl.sub.3) .delta. 0.85-0.92 (m, 6H), 1.30 (t, 3H, J=7.2),
1.35-1.49 (m, 1H), 1.52-1.71 (m, 3H), 1.73-1.94 (m, 2H), 2.15-2.26
(m, 1H), 2.32-2.43 (m, 1H), 3.02-3.18 (m, 2H), 3.19-3.29 (m, 2H),
4.15-4.27 (m, 1H), 4.18 (q, 2H, J=7.2), 4.65-4.74 (m, 1H),
4.76-4.85 (m, 1H), 5.07 (d, 1H, J=12.3), 5.12 (d, 1H, J=12.3),
5.18-5.25 (m, 1H), 5.76-5.84 (m, 2H), 6.64-6.78 (m, 2H), 7.15-7.40
(m, 10H), 7.91-7.98 (m, 1H); Anal. (C.sub.34H.sub.44N.sub.4O.sub.7)
C, H, N.
Example 37
Preparation of Compound (A-28):
Ethyl-3-{Cbz-L-Leu-L-Phe-L-1-(2-imidazolidone)Ala}-E-Propenoate
[0318] ##STR134##
Preparation of Intermediate
Ethyl-3-{Cbz-L-Leu-L-Phe-L-Boc-aminoAla}-E-Propenoate
[0319] (Carbethoxymethlene)triphenylphosphorane (1.20 g, 3.28 mmol,
1.2 equiv.) was added to a solution of
Cbz-L-Leu-L-Phe-L-N-Boc-aminoalaninal (prepared as described in
Webber et al., J. Med. Chem. 1998, vol. 41, 2786) (1.60 g, 2.73
mmol, 1 equiv.) in THF (55 mL), and the reaction was stirred at
room temperature overnight. The volatiles were then removed in
vacuo, and the residue was purified by flash column chromatography
eluting (gradient elution, 0.fwdarw.1.5% CH.sub.3OH in
CH.sub.2Cl.sub.2) to give
ethyl-3-{Cbz-L-Leu-L-Phe-L-N-Boc-aminoAla}-E-propenoate (0.968 g,
contaminated with triphenylphosphine oxide). This material was used
without further purification.
Preparation of Intermediate
Ethyl-3-{Cbz-L-Leu-L-Phe-L-(2-Boc-2-aminoethyl)aminoAla}-E-Propenoate
[0320] A solution of HCl in 1,4-dioxane (4.0 M, 10 mL) was added to
a solution of ethyl-3-{Cbz-L-Leu-L-Phe-L-Boc-aminoAla}-E-propenoate
(0.95 g, 1.46 mmol, 1 equiv.) in the same solvent (20 mL) at
23.degree. C. The reaction mixture was stirred at that temperature
for 1.5 h, and then additional HCl in 1,4-dioxane (4.0 M, 10 mL)
was added. After stirring overnight, the volatiles were removed in
vacuo and the residue was triturated with Et.sub.2O (20 mL). The
resulting solids were filtered and were washed with Et.sub.2O
(3.times.10 mL) to give the crude amine salt (0.63 g 73%, 1.05
mmol) as a white solid.
[0321] This material was dissolved in CH.sub.3OH (10 mL), and then
N-Boc-2-aminoethanal (prepared as described in Bischofberger et
al., J. Org. Chem. 1988, vol. 53, 3457) (0.19 g, 1.16 mmol, 1.1
equiv.) and NaBH.sub.3CN (0.069 g, 1.05 mmol, 1.0 equiv.) were
added sequentially. The reaction mixture was stirred at 23.degree.
C. overnight, and then the volatiles were removed under reduced
pressure. The residue was dissolved in EtOAc (25 mL) and the
organic layer was washed with water (25 mL) and brine (25 mL), and
then dried over MgSO.sub.4 and concentrated. The residue was
purified by flash column chromatography (gradient elution,
0.fwdarw.3% CH.sub.3OH in CH.sub.2Cl.sub.2) to provide
ethyl-3-{Cbz-L-Leu-L-Phe-L-(2-Boc-2-aminoethyl)aminoAla}-E-propenoate
(0.32 g, 44%) as a white amorphous solid: R.sub.f=0.20 (5%
CH.sub.3OH in CHCl.sub.3); IR (cm.sup.-1) 1712, 1649, 1537, 1252,
1175; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.79 (d, 3H, J=6.6),
0.82d, 3H, J=6.6), 1.21 (t, 3H, J=7.0), 1.26-1.37 (m, 13H),
1.46-1.54 (m, 1H), 2.56-2.60 (m, 2H), 2.82-2.97 (m, 4H), 3.98-4.04
(m, 1H), 4.10 (q, 2H, J=7.0), 4.42-4.49 (m, 2H), 4.98 (d, 1H,
J=12.5), 5.04 (d, 1H, J=12.9), 5.59 (d, 1H, J=15.8), 6.73-6.75 (m,
1H), 6.77 (dd, 1H, J=15.8, 4.8), 7.20-7.34 (m, 10H), 7.41 (d, 1H,
J=8.1), 7.97 (d, 1H, J=7.0), 8.07 (d, 1H, J=7.0); Anal.
(C.sub.37H.sub.53N.sub.5O.sub.8.0.50H.sub.2O)C, H, N.
Preparation of Product
Ethyl-3-{Cbz-L-Leu-L-Phe-L-1-(2-imidazolidone)Ala}-E-Propenoate
[0322] TFA (0.8 mL) was added to a solution of
ethyl-3-{Cbz-L-Leu-L-Phe-L-(2-Boc-2-aminoethyl)aminoAla}-E-propenoate
(0.29 g, 0.42 mmol, 1 equiv.) in CH.sub.2Cl.sub.2 (8 mL), and the
reaction mixture was stirred at 23.degree. C. for 2 h. The
volatiles were removed in vacuo, and the residue was dissolved in
EtOAc (25 mL) and washed with saturated NaHCO.sub.3 solution (25
mL), water (25 mL), and brine (25 mL). The organic layer was dried
over MgSO.sub.4 and was concentrated to give the crude diamine
(0.23 g, 92%, 0.39 mmol) as a tan amorphous solid.
[0323] This material was dissolved in THF (4 mL),
carbonyldiimidazole (0.06 g, 0.36 mmol, 0.92 equiv.) was added, and
the reaction mixture was stirred at 23.degree. C. for 3.5 h. The
solvent was removed in vacuo, and the residue was purified by flash
column chromatography (gradient elution, 0.fwdarw.2% CH.sub.3OH in
CH.sub.2Cl.sub.2) to give
ethyl-3-{Cbz-L-Leu-L-Phe-L-J-(2-imidazolidone)Ala}-E-propenoate
(0.12 g, 54%) as a white amorphous solid: mp=161-164.degree. C.;
R.sub.f=0.21 (5% CH.sub.3OH in CHCl.sub.3); IR (cm.sup.-1) 1701,
1647, 1535, 1277; .sup.1H NMR (DMSO-d.sub.6) .delta. 0.79 (d, 3H,
J=6.6), 0.82 (d, 3H, J=6.6), 1.21 (t, 3H, J=7.0), 1.27-1.35 (m,
2H), 1.48-1.52 (m, 1H), 2.79-2.86 (m, 1H), 2.92-3.05 (m, 3H),
3.14-3.19 (m, 2H), 3.25-3.30 (m, 2H), 3.98-4.03 (m, 1H), 4.10 (q,
2H, J=7.0), 4.47-4.49 (m, 1H), 4.59-4.63 (m, 1H), 4.97-5.02 (m,
2H), 5.72 (d, 1H, J=15.8), 6.37 (s, 1H), 6.71 (dd, 1H, J=15.8,
5.5), 7.15-7.39 (m, 10H), 7.42 (d, 1H, J=8.1), 8.00 (d, 1H, J=8.1),
8.18 (d, 1H, J=8.1); Anal. (C.sub.33H.sub.43N.sub.5O.sub.7) C, H,
N.
Example 38
Synthesis of Intermediates Q1, Q2, and Q3
[0324] ##STR135##
Preparation of Intermediate
1-Acetyl-3-(triphenylphosphanylidine)pyrrolidin-2-one
[0325] 2,4-Dibromobutyride (prepared according to Ikuta et al., J.
Med. Chem, 1987, vol. 30, 1995) (46.1 g, 188.2 mmol) in THF (1L)
was cooled to 0.degree. C. and treated with a solution of lithium
bis(trimethylsilylamide) (40.9 g, 244.6 mmol) in THF (200 mL). The
solution was held at 0.degree. C. for 2.5 h, and then poured into
brine (800 mL), extracted with ethyl acetate (2 L), and dried
(MgSO.sub.4). Evaporation yielded 25.5 g of
3-bromo-pyrrolidin-2-one as a brown oil. This material was treated
with Ac.sub.2O (76 mL) and refluxed 5 hours. Evaporation followed
by purification (silica gel filtration, EtOAc elutant) yielded 28 g
of 1-acetyl-3-bromo-pyrrolidin-2-one as a dark oil. THF (272 mL)
and triphenylphosphine (42.8 g, 163.3 mmol) were added and the
resulting solution was refluxed for 8 hours. Upon cooling to
23.degree. C., a precipitate of
1-acetyl-3-(triphenylphosphanyl)-pyrrolidin-2-one bromide formed
and was collected by filtration (27.1 g). Concentration of the
mother liquor, followed by cooling to 0.degree. C., yielded an
additional 6.6 g. The combined material in CH.sub.2Cl.sub.2 (1 L)
was washed with 1N NaOH (100 mL) and then brine (2.times.100 mL).
Evaporation of the organic layer yielded 26.9 g (37% overall) of
1-acetyl-3-(triphenylphosphanylidine)-pyrrolidin-2-one as a tan
oil. .sup.1H NMR (CDCl.sub.3) .delta. 7.76-7.32 (15H, m), 3.90-3.85
(2H, m), 2.50 (3H, s), 2.56-2.30 (2H, m).
Preparation of Intermediate
2-t-Butoxycarbonyl-3-(t-butyldimethylsilanoxy)-propionic Acid
Methyl Ester
[0326] Boc-D-serine methyl ester (20.0 g, 91.2 mmol) in DMF (300
mL) was treated with imidazole (18.6 g, 273.7 mmol) and then TBSCl
(13.0 g, 86.7 mmol). The solution was held at room temperature (rt)
for 8 h, and then washed with saturated aqueous ammonium chloride
(300 mL), and extracted with ethyl acetate (800 mL). The organic
layer was washed with brine (300 mL) and then dried (MgSO.sub.4),
to yield 30.2 g (100%) of
2-t-butoxycarbonyl-3-(t-butyldimethylsilanoxy)-propionic acid
methyl ester as a colorless oil. .sup.1H NMR (CDCl.sub.3) .delta.
5.32 (1H, d, J=8.3), 4.33 (1H, dt, J=8.8, 2.7), 4.02 (1H, dd,
J=10.1, 2.6), 3.80(1H, dd, J=9.8, 3.1), 3.72(3H, s), 1.43 (9H, s),
0.85 (9H, s), 0.0(6H, s).
Preparation of Intermediate
{1-(1-Acetyl-2-oxo-pyrrolidin-3-ylidenemethyl)-2-(t-butyldimethylsilanylo-
xy)-ethyl}-carbamic Acid t-Butyl Ester
[0327] 2-t-Butoxycarbonyl-3-(t-butyldimethylsilanoxy)-propionic
acid methyl ester (12.7 g, 38.0 mmol) in toluene (190 mL) was
cooled to -78.degree. C. and treated with a solution of
diisobutylaluminum hydride (15.6 mL, 87.4 mmol) in toluene (175
mL). The internal temperature was kept below -70.degree. C. The
solution was held at -78.degree. C. for an additional 90 min., and
then methanol (7.7 mL, 190 mmol) was added.
1-Acetyl-3-(triphenylphosphanylidine)-pyrrolidin-2-one (11.1 g,
28.6 mmol) in CH.sub.2Cl.sub.2 (50 mL) was added at -78.degree. C.,
and the resulting solution was allowed to warm to room temperature
and held for 30 minutes. A solution of sodium potassium tartrate
(150 g) in water (600 mL) was added, and stirred vigorously for 30
minutes. The mixture was extracted with ethyl acetate (4.times.250
mL), dried (MgSO.sub.4), and evaporated. Purification by silica gel
chromatography yielded 7.04 g (60%) of
{1-(1-acetyl-2-oxo-pyrrolidin-3-ylidenemethyl)-2-(t-butyldimethy-
lsilanyloxy)-ethyl}-carbamic acid t-butyl ester as a colorless oil.
.sup.1H NMR (CDCl.sub.3) .delta. 6.59 (1H, dt, J=8.7, 2.9), 4.98
(1H, d, J=6.8), 4.37-4.25 (1H, m), 3.77 (2H, t, J=7.3), 3.70-3.58
(2H, m), 2.90-2.80 (1H, m), 2.75-2.60 (1H, m), 5.53 (3H, s), 1.41
(9H, s), 0.87 (9H, s), 0.04 (6H, s).
Preparation of Intermediate
{2-Hydroxy-1-(2-oxo-pyrrolidin-3-ylidenemethyl)-ethyl}-carbamic
Acid t-Butyl Ester
[0328]
{1-(1-Acetyl-2-oxo-pyrrolidin-3-ylidenemethyl)-2-(t-butyldimethyls-
ilanyloxy)-ethyl}-carbamic acid t-butyl ester (9.18 g, 22.2 mmol)
in THF (150 mL) was treated with TBAF (22.2 mL of a 1M solution in
THF, 22.2 mmol) at 0.degree. C., and held for 1 hour. A solution of
saturated aqueous ammonium chloride was added and stirred for 10
min., and then the solution was extracted with ethyl acetate
(2.times.200 mL). The organic layer was dried (MgSO.sub.4) and then
evaporated. Purification by silica gel chromatography yielded 4.82
g (73%) of a colorless oil. This material was taken up in methanol
(160 mL), treated with potassium carbonate (223 mg, 1.62 mmol), and
held for 1 h at rt. The mixture was then treated with solid citric
acid (311 mg, 1.62 mmol), and ethyl acetate (800 mL) was added. The
solution was filtered through silica gel. Evaporation yielded 4.30
g (73% overall) of
{2-hydroxy-1-(2-oxo-pyrrolidin-3-ylidenemethyl)-ethyl}-carbamic
acid t-butyl ester as a colorless oil. .sup.1H NMR (CDCl.sub.3)
.delta. 7.03 (1H, br s), 6.35 (1H, dt, J=8.6, 2.6), 5.37 (1H, d,
J=6.5), 4.40-4.20 (1H, m), 3.66 (br s, 3H), 3.4 (2H, t, J=6.7),
3.10-2.80 (1H, m), 2.80-2.70 (1H, m), 1.41 (9H, s).
Preparation of Intermediate
{2-Hydroxy-1-(2-oxy-pyrrolidin-3-ylmethyl)-ethyl}-carbamic Acid
t-Butyl Ester (Mixture of Diastereomers) (Q1)
[0329]
{2-Hydroxy-1-(2-oxo-pyrrolidin-3-ylidenemethyl)-ethyl}-carbamic
acid t-butyl ester (4.30 g, 16.8 mmol) in ethyl acetate (168 mL)
was treated with 5% palladium on carbon (1.78 g), and hydrogenated
at ambient pressure for 1 h. The mixture was filtered and then
evaporated to yield 3.92 g (91%) of
{2-hydroxy-1-(2-oxy-pyrrolidin-3-ylmethyl)-ethyl}-carbamic acid
t-butyl ester as a colorless oil (1.5:1 mixture of diastereomers):
.sup.1H NMR (CDCl.sub.3) .delta. 6.99 (1H, s), 5.49 (1H, d, J=8.4),
3.70-3.50 (3H, m), 3.38-3.20 (2H, m), 2.60-2.20 (2H, m), 2.00-1.70
(2H, m), 1.65-1.45 (1H, m), 1.37 (9H, s).
Preparation of Intermediate
{2-Hydroxy-1-(R-2-oxy-pyrrolidin-3-ylmethyl)-ethyl}-carbamic Acid
t-Butyl Ester (Q2)
[0330]
{2-Hydroxy-1-(2-oxo-pyrrolidin-3-ylidenemethyl)-ethyl}-carbamic
acid t-butyl ester (98 mg, 0.39 mmol) in methanol (5 mL) was
treated with (R)-BINAP-RuCl (19 mg, 0.02 mmol), and then put under
a hydrogen atmosphere (1200 psi) at 50.degree. C. for 48 h. The
solution was evaporated and then filtered through silica gel (10%
MeOH-EtOAc elutant). Evaporation yielded 75 mg (75%) of
{2-hydroxy-1-(R-2-oxy-pyrrolidin-3-ylmethyl)-ethyl}-carbamic acid
t-butyl ester as a colorless oil (9:1 mixture of diastereomers):
.sup.1H NMR (CDCl.sub.3) .delta. 6.32 (1H, br s), 5.40(1H, d,
J=7.5), 3.82 (1H, br s), 3.71-3.63 (2H, m), 3.34-3.31 (2H, m),
2.45-2.30(2H, m), 2.09-1.86 (2H, m), 1.70-1.63 (1H, m), 1.42 (9H,
s).
Preparation of Intermediate
{2-Hydroxy-1-(S-2-oxy-pyrrolidin-3-ylmethyl)-ethyl}-carbamic Acid
t-Butyl Ester (Q3)
[0331]
{2-Hydroxy-1-(2-oxo-pyrrolidin-3-ylidenemethyl)-ethyl}-carbamic
acid t-butyl ester (0.10 g, 0.39 mmol) in methanol (5 mL) was
treated with (S)-BINAP-RuCl (19 mg, 0.02 mmol), and then put under
a hydrogen atmosphere (1200 psi) at 50.degree. C. for 48 h. The
solution was evaporated and then filtered through silica gel (10%
MeOH-EtOAc elutant). Evaporation yielded 74 mg (74%) of
{2-hydroxy-1-(S-2-oxy-pyrrolidin-3-ylmethyl)-ethyl}-carbamic acid
t-butyl ester as a colorless oil (9:1 mixture of diastereomers):
.sup.1H NMR (CDCl.sub.3) .delta. 6.66 (1H, br s), 5.51 (1H, d,
J=8.2), 3.72-3.57 (3H, m), 3.34-3.31 (2H, m), 2.52-2.33 (2H, m),
2.20-1.86 (1H, m), 1.86-1.70 (1H, m), 1.62-1.50 (1H, m), 1.40 (9H,
s).
[0332] Results of biochemical and biological tests conducted using
various compounds of the invention are described below.
Biochemical and Biological Evaluation
[0333] Inhibition of Rhinovirus 3C Protease
[0334] Stock solutions (50 mM, in DMSO) of various compounds were
prepared; dilutions were in the same solvent. Recombinant
rhinovirus 3C proteases (see Birch et al., "Purification of
recombinant human rhinovirus 14 3C protease expressed in
Escherichia coli," Protein Expr. Pur. 1995, 6(5), 609-618) from
serotypes 14, 16, and 2 were prepared by the following standard
chromatographic procedures: (1) ion exchange using Q Sepharose Fast
Flow from Pharmacia; (2) affinity chromatography using Affi-Gel
Blue from Biorad; and (3) sizing using Sephadex G-100 from
Pharmacia. Each assay sample contained 2% DMSO, 50 mM tris pH 7.6,
1 mM EDTA, a test compound at the indicated concentration,
approximately 1 .mu.M substrate, and 50-100 nM protease. The
k.sub.obs/I values were obtained from reactions initiated by
addition of enzyme rather than substrate. RVP activity was measured
in the fluorescence resonance energy transfer assay. The substrate
was (N-terminal)
DABCYL-(Gly-Arg-Ala-Val-Phe-Gln-Gly-Pro-Val-Gly)-EDANS. In the
uncleaved peptide, the EDANS fluorescence was quenched by the
proximal DABCYL moiety. When the peptide was cleaved, the quenching
was relieved, and activity was measured as an increase in
fluorescence signal. Data were analyzed using standard non-linear
fitting programs (Enzfit), and are shown in Tables 1 and 2 below.
In Table 1, all data are for rhinovirus 3C protease from HRV
serotype-14 (produced from the infectious cDNA clone constructed by
Dr. Robert Rueckert, Institute for Molecular Virology, University
of Wisconsin, Madison, Wis.). Table 2 shows protease-inhibiting
activity of compounds against proteases from several rhinovirus of
serotype other than RVP serotype-14. The data in the column
designated k.sub.obs/[I] were measured from progress curves in
enzyme start experiments.
Antirhinoviral H1-HeLa Cell Culture Assay
[0335] In this cell protection assay, the ability of compounds to
protect cells against HRV infection was measured by the XTT dye
reduction method, which is described in Weislow et al., J. Natl.
Cancer Inst. 1989, vol. 81, 577-586.
[0336] H1-HeLa cells were infected with HRV-14 at a multiplicity of
infection (m.o.i.) of 0.13 (virus particles/cell) or mock-infected
with medium only. Infected or mock-infected cells were resuspended
at 8.times.10.sup.5 cells per mL, and incubated with appropriate
concentrations of the compounds to be tested. Two days later,
XTT/PMS was added to test plates and the amount of formazan
produced was quantified spectrophotometrically at 450/650 nm. The
EC.sub.50 value was calculated as the concentration of compound
that increased the percentage of formazan production in
compound-treated, virus-infected cells to 50% of that produced by
compound-free, mock-infected cells. The 50% cytotoxic dose
(CC.sub.50) was calculated as the concentration of compound that
decreased the percentage of formazan produced in compound-treated,
mock-infected cells to 50% of that produced by compound-free,
mock-infected cells. The therapeutic index (TI) was calculated by
dividing the CC.sub.50 value by the EC.sub.50 value.
[0337] All strains of human rhinovirus (HRV) for use in this assay
were purchased from American Type Culture Collection (ATCC), except
for HRV serotype-14 (produced from the infectious cDNA clone
constructed by Dr. Robert Rueckert, Institute for Molecular
Virology, University of Wisconsin, Madison, Wis.). HRV stocks were
propagated and viral assays were performed in H1-HeLa cells (ATCC).
Cells were grown in minimal essential medium with 10% fetal bovine
serum, available from Life Technologies (Gaithersburg, Md.).
[0338] The compounds were tested against control compounds WIN
51711, WIN 52084, and WIN 54954 (obtained from Sterling-Winthrop
Pharmaceuticals), Pirodavir (obtained from Janssen
Pharmaceuticals), and Pleconaril (prepared according to the method
described in Diana et al., J. Med. Chem 1995, vol. 38, 1355).
Antiviral data obtained for the test compounds are shown in Tables
1 and 3, where all data are for HRV serotype-14 unless otherwise
noted in parentheses. The designation "ND" indicates that a value
was not determined for that compound. TABLE-US-00002 TABLE 1
Activity vs. HRV Serotype-14 Protease Inhibition Cell Protection
k.sub.obs/[I] EC.sub.50 Compound (M.sup.-1sec.sup.-1) (.mu.M)
Example 31 25,000 0.61 (Comparison Compound #2) Example 35 (A-26)
239,000 0.03 Example 33 (A-24) 257,000 0.10 Example 36 (A-27)
18,000 1.6 Example 32 62,500 0.38 (Comparison Compound #3) Example
34 (A-25) 500,000 0.03 Example 5 (A-4) 270,000 0.10 Example 8 (A-7)
980,000 0.004 Example 7 (A-6) 248,000 0.42 Example 9 (A-8) 900,000
ND Example 6 (A-5) 1,500,000 0.005 Example 12 (C-1) 68,400 0.10
Example 18 (C-2) 270,000 0.002 Example 10 (B-1) 240,000 0.10
Example 20 (B-4) 500,000 <0.03 Example 17 (B-2) 1,090,000 0.005
Example 1 573 >320 (Comparison Compound #1) Example 2 (A-1)
260,000 0.25 Example 3 (A-2) 46,900 1.6 Example 4 (A-3) 310,000
0.05 Example 11 (A-10) 108,000 0.14 Example 13 (A-11) 108,000 0.03
Example 14 (A-9) 66,000 1.8 Example 15 (A-12) 59,300 0.40 Example
16 (A-13) 95,800 0.20 Example 19 (B-3) 465,000 0.18 Example 21
(A-14) 54,500 0.48 Example 22 (A-15) 237,100 0.22 Example 23 (A-16)
172,800 0.45 Example 24 (A-17) 167,000 0.06 Example 25 (A-18)
292,000 1.5 Example 26 (A-19) 27,750 25.1 Example 27 (A-20) 1,020
12.6 Example 28 (A-21) 17,800 2.5 Example 29 (A-22) 2,400 nd
Example 30 (A-23) 26,000 nd
[0339] TABLE-US-00003 TABLE 2 Activity vs. Other HRV Serotypes
Compound Rhinovirus Serotype k.sub.obs/I (M.sup.-1sec.sup.-1)
Comparison Compound #2 (16) 6,500 '' (89) 3,400 '' (2) 2,000
Comparison Compound #3 (2) 8,000 '' (16) 16,900 (A-24) (2)
31,000
[0340] TABLE-US-00004 TABLE 3 Anti-Rhinovirus Activity # HRV
EC.sub.50 (.mu.M) CC.sub.50 (.mu.M) TI Comparison >320 >320
-- Compound #1 (A-1) 0.25 >100 >400 '' (2) 0.41 '' >243 ''
(1A) 1.0 '' >100 '' (89) 0.22 '' >450 (A-2) 1.6 >100
>63 (A-3) 0.05 >10 >200 (A-4) 0.10 >100 >1000 (A-5)
0.005 >10 >2000 '' (2) 0.01 '' >1000 '' (16) 0.02 ''
>500 '' (39) 0.02 '' >500 '' (89) 0.02 '' >500 '' (10)
0.05 '' >200 '' (1A) 0.03 '' >333 '' (3) 0.05 '' >200 ''
(9) 0.04 '' >250 '' (12) 0.06 '' >166 '' (13) 0.02 '' >500
'' (17) 0.02 '' >500 '' (25) 0.18 '' >55 '' (30) 0.06 ''
>166 '' (38) 0.13 '' >76 '' (87) 0.21 '' >47 (A-6) 0.42
>100 >237 (A-7) 0.004 >10 >2500 '' (2) 0.02 '' >500
'' (16) 0.04 '' >250 '' (39) 0.02 '' >500 '' (89) 0.02 ''
>500 '' (10) 0.06 '' >166 '' (1A) 0.03 '' >333 '' (3) 0.02
'' >500 '' (9) 0.04 '' >250 '' (12) 0.07 '' >143 '' (13)
0.03 '' >333 '' (17) 0.02 '' >500 '' (25) 0.20 '' >50 ''
(30) 0.09 '' >111 '' (38) 0.17 '' >58 '' (87) 0.59 '' >16
(A-8) ND ND ND (B-1) 0.10 >100 >1000 '' (1A) 0.30 '' >333
'' (10) 0.40 '' >250 (A-10) 0.14 >100 >714 (C-1) 0.10
>10 >100 (A-11) 0.03 50 1667 (A-9) 1.8 >100 >55 (A-12)
0.40 >100 >250 (A-13) 0.20 >10 >50 (B-2) 0.005 >100
>20000 '' (2) 0.02 '' >5000 '' (16) 0.01 '' >10000 '' (39)
0.05 '' >2000 '' (89) 0.009 '' >11111 '' (10) 0.02 ''
>5000 '' (1A) 0.02 '' >5000 '' (3) 0.02 '' >5000 '' (9)
0.006 '' >16666 '' (12) 0.05 '' >2000 '' (13) 0.01 ''
>10000 '' (17) 0.01 '' >10000 '' (25) 0.03 '' >3333 ''
(30) 0.04 '' >2500 '' (38) 0.07 '' >1428 '' (87) 0.06 ''
>1666 (C-2) 0.002 >10 >5000 '' (2) 0.004 '' >2500 ''
(16) 0.01 '' >1000 '' (39) 0.01 '' >1000 '' (89) 0.004 ''
>2500 '' (10) 0.02 '' >500 '' (1A) 0.01 '' >1000 '' (3)
0.02 '' >500 '' (9) 0.01 '' >1000 '' (12) 0.04 '' >250 ''
(13) 0.007 '' >1428 '' (17) 0.007 '' >1428 '' (25) 0.07 ''
>142 '' (30) 0.03 '' >333 '' (38) 0.05 '' >200 '' (87)
0.02 '' >500 (B-3) 0.18 >100 >543 (B-4) <0.03 >100
>3333 (A-14) 0.48 >100 >208 (A-15) 0.22 >100 >454 ''
(1A) 7.1 '' >14 '' (10) 2.7 '' >37 (A-16) 0.45 >100
>222 '' (1A) 4.8 '' >21 '' (10) 4.5 '' >22 (A-17) 0.06
>100 >1786 '' (1A) 1.8 '' >56 '' (10) 3.3 '' >30 (A-18)
1.5 >100 >67 (A-19) 25.1 >100 >4 (A-20) 12.6 >100
>8 (A-21) 2.5 >100 >40 (A-22) ND ND ND (A-23) ND ND ND
Comparison 0.61 >320 >524 Compound #2 Comparison (16) 2.3
>320 >139 Compound #2 Comparison (89) 6.3 >320 >50
Compound #2 Comparison (10) 0.60 >320 >533 Compound #2
Comparison 0.38 >320 >842 Compound #3 (A-24) 0.10 >100
>1000 (A-25) 0.030 >100 >3333 (A-26) 0.030 >100
>3333 (A-27) 1.6 >100 >62 WIN 51711 0.78 >60 >77 WIN
52084 0.07 >10 >143 WIN 54954 2.13 >63 >30 Pirodavir
0.03 >10 >300 Pleconaril 0.01 >10 >1000
Anticoxsackieviral Cell Culture Assay
[0341] Coxsackievirus types A-21 (CAV-21) and B3 (CVB-3) were
purchased from American Type Culture Collection (ATCC, Rockville,
Md.). Virus stocks were propagated and antiviral assays were
performed in H1-HeLa cells (ATCC). Cells were grown in minimal
essential medium with 10% fetal bovine serum (Life Technologies,
Gaithersburg, Md.).
[0342] The ability of compounds to protect cells against either
CAV-21 or CVB-3 infection was measured by the XTT dye reduction
method. This method is described in Weislow et al., J. Natl. Cancer
Inst. 1989, vol. 81, 577-586. H1-HeLa cells were infected with
CAV-21 or CVB-3 at a multiplicity of infection (m.o.i.) of 0.025 or
0.075, respectively, or mock-infected with medium only. H1-HeLa
cells were plated at 4.times.10.sup.4 cells per well in a 96-well
plate and incubated with appropriate concentrations of the test
compound. One day (CVB-3) or two days (CAV-21) later, XTT/PMS was
added to test plates and the amount of formazan produced was
quantified spectrophotometrically at 450/650 nm. The EC.sub.50 was
calculated as the concentration of compound that increased the
formazan production in compound-treated, virus-infected cells to
50% of that produced by compound-free, uninfected cells. The 50%
cytotoxic dose (CC.sub.50) was calculated as the concentration of
compound that decreased formazan production in compound-treated,
uninfected cells to 50% of that produced in compound-free,
uninfected cells. The therapeutic index TI) was calculated by
dividing the CC.sub.50 by the EC.sub.50.
[0343] The compounds were tested against control compounds WIN
54954 (obtained from Sterling-Winthrop Pharmaceuticals), Pirodavir
(obtained from Janssen Pharmaceuticals), and Pleconaril (prepared
according to Diana et al., J. Med. Chem. 1995, 38, 1355). Antiviral
data obtained for the test compounds against CAV-21 and CVB-3 are
shown in Table 4. TABLE-US-00005 TABLE 4 Anti-Coxsackievirus
Activity Compound Strain EC.sub.50 (.mu.M) CC.sub.50 (.mu.M) TI
(A-5) CAV-21 0.23 >10 >43 '' CVB-3 1.0 '' >10 (B-2) CAV-21
0.16 >100 >625 '' CVB-3 0.18 '' >555 WIN 54954 CAV-21
>100 >100 '' CVB-3 >100 '' Pirodavir CAV-21 >100
>100 '' CVB-3 >100 '' Pleconaril CAV-21 0.09 >10 >107
'' CVB-3 >10 ''
[0344] Anti-Echoviral and -Enteroviral Cell Culture Assays
[0345] Echovirus type 11 (EV 11) and enterovirus type 70 (EV 70)
were purchased from ATCC (Rockville, Md.). Virus stocks were
propagated and antiviral assays were performed in MRC-5 cells
(ATCC). Cells were grown in minimal essential medium with 10% fetal
bovine serum (Life Technologies, Gaithersburg, Md.).
[0346] The ability of compounds to protect cells against either EV
11 or EV 70 infection was measured by the XTT dye reduction method
(Weislow et al., J. Natl. Cancer Inst. 1989, vol. 81, 577-586).
MRC-5 cells were infected with EV 11 or EV 70 at an m.o.i. of 0.003
or 0.004, respectively, or mock-infected with medium only. Infected
or uninfected cells were added at 1.times.10.sup.4 cells per well
and incubated with appropriate concentrations of compound. Four
days later, XTT/PMS was added to test plates, and the amount of
formazan produced was quantified spectrophotometrically at 450/650
nm. The EC.sub.50 was calculated as the concentration of compound
that increased the formazan production in compound-treated,
virus-infected cells to 50% of that produced by compound-free,
uninfected cells. The 50% cytotoxic dose (CC.sub.50) was calculated
as the concentration of compound that decreased formazan production
in compound-treated, uninfected cells to 50% of that produced in
compound-free, uninfected cells. The therapeutic index (TI) was
calculated by dividing the CC.sub.50 by the EC.sub.50.
[0347] The compounds were tested against control compounds
Pirodavir (obtained from Janssen Pharmaceuticals) and Pleconaril
(prepared according to Diana et al., J. Med. Chem. 1995, vol. 38,
1355). Antiviral data obtained for the test compounds against
strain EV 11 and EV 70 are shown below in Table 5. TABLE-US-00006
TABLE 5 Anti-Echovirus and Anti-Enterovirus Activity Compound
Strain EC.sub.50 (.mu.M) CC.sub.50 (.mu.M) TI (A-5) EV-11 0.08
>10 >125 '' EV-70 0.04 '' >250 (B-2) EV-11 0.01 >100
>10000 '' EV-70 0.003 '' >33333 Pirodavir EV-11 3.7 >10
>3 '' EV-70 0.06 '' >167 Pleconaril EV-11 0.16 >10 >62
'' EV-70 ND ND ND
[0348] While the invention has been described in terms of preferred
embodiments and specific examples, those skilled in the art will
recognize that various changes and modifications can be made
without departing from the spirit and scope of the invention. Thus,
the invention should be understood as not being limited by the
foregoing detailed description, but as being defined by the
appended claims and their equivalents.
* * * * *