U.S. patent application number 10/259041 was filed with the patent office on 2003-07-17 for compounds useful for treating hepatitis c virus.
Invention is credited to Decicco, Carl P., Hudyma, Thomas W., Priestley, Eldon Scott, Zheng, Xiaofan.
Application Number | 20030134853 10/259041 |
Document ID | / |
Family ID | 23265472 |
Filed Date | 2003-07-17 |
United States Patent
Application |
20030134853 |
Kind Code |
A1 |
Priestley, Eldon Scott ; et
al. |
July 17, 2003 |
Compounds useful for treating hepatitis C virus
Abstract
A series of compounds of Formula I are disclosed which are
useful in treating viral hepatitus C. 1
Inventors: |
Priestley, Eldon Scott;
(Hockessin, DE) ; Decicco, Carl P.; (Kennett
Square, PA) ; Hudyma, Thomas W.; (Durham, CT)
; Zheng, Xiaofan; (Cheshire, CT) |
Correspondence
Address: |
STEPHEN B. DAVIS
BRISTOL-MYERS SQUIBB COMPANY
PATENT DEPARTMENT
P O BOX 4000
PRINCETON
NJ
08543-4000
US
|
Family ID: |
23265472 |
Appl. No.: |
10/259041 |
Filed: |
September 26, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60324874 |
Sep 26, 2001 |
|
|
|
Current U.S.
Class: |
514/233.5 ;
514/254.06; 514/256; 514/314; 514/338; 514/362; 514/364; 514/394;
544/139; 544/333; 544/370; 546/167; 546/273.4; 548/127; 548/131;
548/304.7 |
Current CPC
Class: |
C07D 419/04 20130101;
C07D 401/14 20130101; C07D 413/04 20130101; A61P 31/00 20180101;
C07D 409/14 20130101; C07D 403/04 20130101; C07D 413/14 20130101;
C07D 403/14 20130101; C07D 471/04 20130101; C07D 405/14 20130101;
C07D 235/18 20130101 |
Class at
Publication: |
514/233.5 ;
514/256; 514/314; 514/254.06; 514/338; 514/362; 514/364; 514/394;
544/139; 544/333; 544/370; 546/167; 546/273.4; 548/127; 548/131;
548/304.7 |
International
Class: |
A61K 031/5377; A61K
031/506; A61K 031/4709; A61K 031/496; A61K 031/4439; A61K 031/4184;
C07D 417/02; C07D 413/02; C07D 45/02; C07D 43/02 |
Claims
We claim:
1. A compound of Formula I 58wherein: Q is CH or N; R.sup.1 is
tetrazolyl, MeCONHSO.sub.2--, PhCONHSO.sub.2--,
R.sup.5O.sub.2C(CH.sub.2)- .sub.0-3CONHSO.sub.2--, R.sup.2 is 59
--CH.sub.2Ar.sup.1, --CHPh.sub.2 , --CH.sub.2CO(4-FPh),
--CH.sub.2CO(4-CF.sub.3Ph), or --CH.sub.2CONp where Np is naphthyl;
R.sup.3 is C.sub.5-7cycloalkyl; R.sup.4 is hydrogen, Ar.sup.2, or
Ar.sup.3; Ar.sup.1 is selected from the following group: phenyl,
halophenyl, 60 61Ar.sup.2 is phenyl, naphthyl, or biphenyl,
optionally substituted with 1-3 substituents selected from the
group comprising halogen, C.sub.1-6 alkyl, hydroxyC.sub.1-6alkyl,
C.sub.1-6alkoxy, C.sub.1-6sulfoxy, C.sub.1-2perfluoroalkyl,
hydroxy, formyl, C.sub.1-6alkylcarbonyl, cyano, nitro,
C.sub.1-6alkylamido, CO.sub.2R.sup.5, CONR.sup.5R.sup.5,
C.sub.1-6alkylsulfonamido, and dioxolane; Ar.sup.3 is thienyl,
furanyl, pyrrolyl, benzothiophenyl, benzofuranyl, indolyl,
quinolinyl, or pyrimidinyl optionally substituted with 1-2
substituents selected from the group comprising C.sub.1-6alkyl,
formyl, acetoxy, trifluoroacetoxy, and t-butoxycarbonyl; R.sup.5 is
hydrogen or C.sub.1-6alkyl; R.sup.6 is halogen, methoxy,
CO.sub.2R.sup.5 or CONR.sup.7R.sup.8; R.sup.7 and R.sup.8 are
independently hydrogen, C.sub.1-6alkyl, --CH(Me)CO.sub.2R.sup.5,
--(CH.sub.2).sub.1-3CO.sub.2R.su- p.5,
--(CH.sub.2).sub.1-3CONR.sup.5R.sup.5, --(CH.sub.2).sub.1-3OH, 62
or 63 or R.sup.7 and R.sup.8 taken together with the nitrogen to
which they are attached form pyrrolidine, morpholine, piperidine,
4-hydroxypiperidine, piperazine, or 4-methylpiperazine; or a
pharmaceutically acceptable salt, solvate, or prodrug thereof.
2. A compound of claim 1 wherein R.sup.3 is cyclohexyl.
3. A compound of claim 1 wherein R.sup.1 is tetrazolyl and R.sup.2
is 64
4. A compound of claim 3 wherein R.sup.4 is Ar.sup.2.
5. A compound of claim 4 wherein R.sup.3 is cyclohexyl.
6. A compound of claim 3 wherein R.sup.4 is Ar.sup.3.
7. A compound of claim 6 wherein R.sup.3 is cyclohexyl.
8. A compound of claim 3 wherein R.sup.4 is hydrogen.
9. A compound of claim 8 wherein R.sup.3 is cyclohexyl.
10. A compound of claim 1 wherein R.sup.2 is
--CH.sub.2Ar.sup.1.
11. A compound of claim 10 wherein R.sup.3 is cyclohexyl.
12. A composition useful for treating hepatitus C comprising a
therapeutic amount of a compound of claim 1 and a pharmaceutically
acceptable carrier.
13. A method for treating hepatitus C comprising administering a
therapeutically effective amount of a compound of claim 1 to a
patient.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims the benefit of U.S. provisional
application U.S. Ser. No. 60/324,874, filed Sep. 26, 2001.
FIELD OF THE INVENTION
[0002] The present invention is directed to compounds which inhibit
the RNA-dependent RNA polymerase (RdRp) encoded by Hepatitis C
virus (HCV). The compounds, or pharmaceutically acceptable salts or
prodrugs thereof, are of value in the treatment and/or prevention
of infection by HCV.
BACKGROUND OF THE INVENTION
[0003] 1. The Field of the Invention
[0004] HCV is a major human pathogen, infecting an estimated 170
million persons worldwide--roughly five times the number infected
by human immunodeficiency virus type 1. A substantial fraction of
these HCV infected individuals develop serious progressive liver
disease, including cirrhosis and hepatocellular carcinoma. (Lauer,
G. M.; Walker, B. D. N. Engl. J. Med. (2001), 345, 41-52).
[0005] Presently, the most effective HCV therapy employs a
combination of alpha-interferon and ribavirin, leading to sustained
efficacy in 40% of patients. (Poynard, T. et al. Lancet (1998),
352, 1426-1432). Recent clinical results demonstrate that pegylated
alpha-interferon is superior to unmodified alpha-interferon as
monotherapy (Zeuzem, S. et al. N. Engl. J. Med. (2000), 343,
1666-1672). However, even with experimental therapeutic regimens
involving combinations of pegylated alpha-interferon and ribavirin,
a substantial fraction of patients do not have a sustained
reduction in viral load. In addition, the prospects for development
of a prophylactic or therapeutic vaccine appear dim, in spite of
intensive research efforts. Thus, there is a clear and long-felt
need to develop effective therapeutics for treatment of HCV
infection.
[0006] HCV is a positive-stranded RNA virus. Based on comparison of
deduced amino acid sequence and the extensive similarity in the 5'
untranslated region, HCV has been classified as a separate genus in
the Flaviviridae family. All members of the Flaviviridae family
have enveloped virions that contain a positive stranded RNA genome
encoding all known virus-specific proteins via translation of a
single, uninterrupted, open reading frame.
[0007] Considerable heterogeneity is found within the nucleotide
and encoded amino acid sequence throughout the HCV genome. At least
six major genotypes have been characterized, and more than 50
subtypes have been described. The major genotypes of HCV differ in
their distribution worldwide, and the clinical significance of the
genetic heterogeneity of HCV remains elusive despite numerous
studies of the possible effect of genotypes on pathogenesis and
therapy.
[0008] The RNA genome is about 9.6 Kb in length, and encodes a
single polypeptide of about 3000 amino acids. The 5' untranslated
region contains an internal ribosome entry site (IRES), which
directs cellular ribosomes to the correct AUG for initiation of
translation. The translated product contains the following
proteins: core-E1-E2-p7-NS2-NS3-NS4A-NS4B-NS5A-NS5B. This precursor
protein is cotranslationally and posttranslationally processed into
at least 10 viral structural (core, E1, E2) and nonstructural
(NS2-NS5B) proteins by the action of host cell signal peptidase and
by two distinct viral proteinase activities (NS2/3 and NS3).
[0009] Although the functions of the NS proteins are not completely
defined, it is known that NS3 is a serine protease/RNA helicase,
NS4A is a protease cofactor, and NS5B is an RNA dependent RNA
polymerase involved in viral replication. It has recently been
demonstrated that functional NS5B is required for virus infectivity
in chimpanzees (Kolykhalov, A. A. et al. J. Virol. (2000), 74,
2046-2051). This result strongly suggests that inhibition of the
NS5B RdRp is a viable approach for the development of HCV
therapeutic agents.
[0010] 2. Description of Related Art
[0011] Efforts toward the development of HCV NS5B RdRp inhibitors
have resulted in the following disclosures:
[0012] Altamura et al. (Istituto Di Ricerche Di Biologia
Molecolare) describe diketoacid RdRp inhibitors (WO 00/06529 and WO
02/06246 A1). Altamura et al. suggest that the diketoacids and
dihydroxypyrimidine carboxylic acids inhibit HCV RdRp by
interfering with the binding of phosphoryl groups at the active
site of the enzyme.
[0013] A series of three disclosures from Viropharma Inc. (Bailey,
T. R. et al, WO 00/10573; Bailey, T. R. et al, WO 00/13708; Young,
D. C. et al, WO 00/18231) describe HCV RdRp inhibitors. WO 00/10573
covers a series of rhodanine derivatives, WO 00/13708 covers a
series of barbituric acid or thiobarbituric acid derivatives, and
WO 0018231 covers a series of dihydrobenzothiophene
derivatives.
[0014] R. Storer (Biochem Pharma, Inc.) has disclosed the use of a
series of dioxolane nucleosides for treatment of HCV (WO
01/32153).
[0015] EP 1162196 (Japan Tobacco Inc.) discloses a series of fused
ring heterocycles as inhibitors of HCV RdRp. These compounds may be
distinguished from the applicants' compounds in the nature of the
"A" substituent in applicants' Formula I compounds.
[0016] WO 02/04425 (Boehringer Ingelheim) discloses a series of HCV
NS5B polymerase inhibitors which also may be distinguished from the
applicants' compounds in the nature of the "A" substituent in
applicants.degree. Formula I compounds.
[0017] WO 01/85172 (Smithkline Beecham) discloses a series of
1-(alkyl)-3-(1,1-dioxo-2H-benzo-1,2,4-thiadiazin-3-yl)-4-hydroxy-2-quinol-
ones as HCV inhibitors.
SUMMARY OF THE INVENTION
[0018] The present invention is directed to compounds according to
Formula I: 2
[0019] wherein all represented groups are defined below.
[0020] The present invention is also directed to compounds of
Formula I, or pharmaceutically acceptable salts or prodrugs
thereof, which are useful as inhibitors of HCV NS5B RdRp. It is
another object of the present invention to provide pharmaceutical
compositions comprising a pharmaceutically acceptable carrier and a
therapeutically effective amount of a compound of Formula I, or
pharmaceutically acceptable salt or prodrug thereof. It is another
object of the present invention to provide a method for the
treatment or prevention of HCV comprising administering to a host
in need of such treatment a therapeutically effective amount of a
compound of Formula I, or a pharmaceutically acceptable salt or
prodrug thereof. These and other objects of the invention, which
will become apparent during the following detailed description,
have been achieved by the discovery that compounds of Formula I
inhibit the HCV NS5B RdRp.
DETAILED DESCRIPTION OF THE INVENTION
[0021] The present invention is directed to compounds according to
Formula I: 3
[0022] wherein: Q is CH or N; R.sup.1 is tetrazolyl,
MeCONHSO.sub.2--, PhCONHSO.sub.2--, 4
[0023] R.sup.5O.sub.2C(CH.sub.2).sub.0-3CONHSO.sub.2--, 5
[0024] --CH.sub.2Ar.sup.1, --CHPh.sub.2, --CH.sub.2CO(4-FPh),
--CH.sub.2CO(4-CF.sub.3Ph), or --CH.sub.2CONp where Np is naphthyl;
R.sup.3 is C.sub.5-7cycloalkyl; R.sup.4 is hydrogen, Ar.sup.2, or
Ar.sup.3; Ar.sup.1 is selected from the following 6
[0025] group: phenyl, halophenyl,
[0026] optionally substituted with 1-3 substituents selected from
the group comprising halogen, C.sub.1-6 alkyl, hydroxy
C.sub.1-6alkyl, C.sub.1-6alkoxy, C.sub.1-6sulfoxy,
C.sub.1-2perfluoroalkyl, hydroxy, formyl, C.sub.1-6alkylcarbonyl,
cyano, nitro, C.sub.1-6alkylamido, CO.sub.2R.sup.5,
CONR.sup.5R.sup.5, C.sub.1-6alkylsulfonamido, and dioxolane;
Ar.sup.3 is thienyl, furanyl, pyrrolyl, benzothiophenyl,
benzofuranyl, indolyl, quinolinyl, or pyrimidinyl optionally
substituted with 1-2 substituents selected from the group
comprising C.sub.1-6alkyl, formyl, acetoxy, trifluoroacetoxy, and
t-butoxycarbonyl; R.sup.5 is hydrogen or C.sub.1-6alkyl; R.sup.6 is
halogen, methoxy, CO.sub.2R.sup.5 or CONR.sup.7R.sup.8; R.sup.7 and
R.sup.8 are independently hydrogen, C.sub.1-6alkyl,
--CH(Me)CO.sub.2R.sup.5, --(CH.sub.2).sub.1-3CO.sub.2R.su- p.5,
--(CH.sub.2).sub.1-3CONR.sup.5R.sup.5, --(CH.sub.2).sub.1-3OH,
7
[0027] or R.sup.7 and R.sup.8 taken together with the nitrogen to
which they are attached form pyrrolidine, morpholine, piperidine,
4-hydroxypiperidine, piperazine, or 4-methylpiperazine; or a
pharmaceutically acceptable salt, solvate, or prodrug thereof.
[0028] As used herein, the following terms shall be understood to
have the meaning set forth in the following definitions.
[0029] The term "compounds of the invention", and equivalent
expressions, are meant to embrace compounds of formula I, and
include prodrugs, pharmaceutically acceptable salts, and solvates,
e.g. hydrates. Similarly, reference to intermediates, whether or
not they themselves are claimed, is meant to embrace their salts,
and solvates, where the context so permits.
[0030] The term "derivative" means a chemically modified compound
wherein the modification is considered routine by the ordinary
skilled chemist, such as an ester or an amide of an acid,
protecting groups, such as a benzyl group for an alcohol or thiol,
and tert-butoxycarbonyl group for an amine.
[0031] The term "analog" means a compound which comprises a
chemically modified form of a specific compound or class thereof,
and which maintains the pharmaceutical and/or pharmacological
activities characteristic of said compound or class.
[0032] The term "solvate" means a physical association of a
compound of this invention with one or more solvent molecules. This
physical association includes hydrogen bonding. In certain
instances the solvate will be capable of isolation, for example
when one or more solvent molecules are incorporated in the crystal
lattice of the crystalline solid. "Solvate" encompasses both
solution-phase and isolable solvates. Exemplary solvates include
hydrates, ethanolates, methanolates, and the like.
[0033] The term "effective amount" means an amount of a
compound/composition according to the present invention effective
in producing the desired therapeutic effect, i.e. inhibiting
HCV.
[0034] The term "patient" includes both human and other
mammals.
[0035] The term "pharmaceutical composition" means a composition
comprising a compound of formula I in combination with at least one
additional pharmaceutical adjuvant, excipient, vehicle or carrier
component, such as diluents, preserving agents, fillers, flow
regulating agents, disintegrating agents, wetting agents,
emulsifying agents, suspending agents, sweetening agents, flavoring
agents, perfuming agents, antibacterial agents, antifungal agents,
lubricating agents and dispensing agents, depending on the nature
of the mode of administration and dosage forms. Any ingredient
listed in Remington's Pharmaceutical Sciences, 18.sup.th ed., Mack
Publishing Company, may be used.
[0036] The term "alkyl" is intended to include both branched and
straight-chain saturated aliphatic hydrocarbon groups having the
specified number of carbon atoms. Thus, C.sub.1-6 alkyl refers to
an alkyl group having from one to six carbon atoms. Examples of
alkyl include, but are not limited to, methyl, ethyl, n-propyl,
i-propyl, n-butyl, s-butyl, and t-butyl, pentyl, hexyl, heptyl and
octyl.
[0037] The terms "linear and cyclic heteroalkyl" are defined in
accordance with the term "alkyl" with the suitable replacement of
carbon atoms with an atom such as oxygen, nitrogen or sulfur which
would render a chemically stable species.
[0038] The term "heterocyclic C.sub.1-6 alkyl group means a
C.sub.1-6 alkyl that is substituted by a heterocyclic group.
[0039] The terms "halo" or "halogen" as used herein refer to
fluoro, chloro, bromo and iodo.
[0040] Alkanoyl refers to a substituent group having a C.sub.1-6
alkyl component bonded to a carbonyl group which is then bonded to
the backbone to which the substituent group is connected.
[0041] The term "alkoxy" is intended to represent an alkyl group
with the indicated number of carbon atoms attached to an oxygen
atom. Examples of alkoxy include, but are not limited to, methoxy,
ethoxy, n-propoxy, i-propoxy, n-butoxy, s-butoxy, and t-butoxy.
[0042] The term "aryl" is intended to mean an aromatic moiety
containing the specified number of carbon atoms, such as, but not
limited to phenyl, indanyl or naphthyl. So C.sub.6-14 aryl refers
to an aromatic moiety having from six to fourteen carbon atoms
which may be in the form of a single, bicyclic or tricyclic
structure. The term "haloaryl" as used herein refers to an aryl
mono, di or tri substituted with halogen atoms.
[0043] As used herein, the term "cycloalkyl" refers to a saturated
monocyclic alkyl group. Examples of such carbocycles include, but
are not limited to, cyclopropyl, cyclobutyl, cyclopentyl,
cyclohexyl, cycloheptyl and cyclooctyl.
[0044] As used herein, the term "heterocycle" or "heterocyclic
system" is intended to mean a stable 5- to 7-membered monocyclic or
bicyclic or 7- to 14-membered bicyclic heterocyclic ring which is
saturated, partially unsaturated or unsaturated (aromatic), and
which consists of carbon atoms and from 1 to 4 heteroatoms
independently selected from the group consisting of N, O and S and
including any bicyclic group in which any of the above-defined
heterocyclic rings is fused to a benzene ring. The nitrogen and
sulfur heteroatoms may optionally be oxidized. The heterocyclic
ring may be attached to its pendant group at any heteroatom or
carbon atom which results in a stable structure. The heterocyclic
rings described herein may be substituted on carbon or on a
nitrogen atom if the resulting compound is stable. If specifically
noted, a nitrogen in the heterocycle may optionally be quaternized.
It is preferred that when the total number of S and o atoms in the
heterocycle exceeds 1, then these heteroatoms are not adjacent to
one another. It is preferred that the total number of S and O atoms
in the heterocycle is not more than 2. Also included are fused ring
and spiro compounds containing, for example, the above
heterocycles.
[0045] The term "aromatic heterocyclic system" or "heteroaryl"
means a stable 5- to 7-membered monocyclic or bicyclic or 7- to
10-membered bicyclic heterocyclic aromatic ring which consists of
carbon atoms and from 1 to 4 heterotams independently selected from
the group consisting of N, O and S. The term includes any bicyclic
group in which any of the above-defined heterocyclic rings is fused
to a benzene ring. It is preferred that the total number of S and O
atoms in the aromatic heterocycle is not more than 2.
[0046] Examples of heterocycles include, but are not limited to,
piperidinyl, morpholinyl, piperazinyl, 1H-indazole, 2-pyrrolidonyl,
2H,6H-1,5,2-dithiazinyl, 2H-pyrrolyl, 3H-indolyl, 4-piperidonyl,
4aH-carbazole, 4H-quinolizinyl, 6H-1,2,5-thiadiazinyl, acridinyl,
azocinyl, benzimidazolyl, benzofuranyl, benzothiofuranyl,
benzothiophenyl, benzoxazolyl, benzthiazolyl, benztriazolyl,
benztetrazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazalonyl,
carbazolyl, 4aH-carbazolyl, b-carbolinyl, chromanyl, chromenyl,
cinnolinyl, decahydroquinolinyl, 2H,6H-1,5,2-dithiazinyl,
dihydrofuro[2,3-b]tetrahydrofuran, furanyl, furazanyl,
imidazolidinyl, imidazolinyl, imidazolyl, 1H-indazolyl, indolenyl,
indolinyl, indolizinyl, indolyl, isobenzofuranyl, isochromanyl,
isoindazolyl, isoindolinyl, isoindolyl, isoquinolinyl,
isothiazolyl, isoxazolyl, morpholinyl, naphthyridinyl,
octahydroisoquinolinyl, oxadiazolyl, 1,2,3-oxadiazolyl,
1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl,
oxazolidinyl., oxazolyl, oxazolidinylperimidinyl, phenanthridinyl,
phenanthrolinyl, phenarsazinyl, phenazinyl, phenothiazinyl,
phenoxathiinyl, phenoxazinyl, phthalazinyl, piperazinyl,
piperidinyl, pteridinyl, piperidonyl, 4-piperidonyl, pteridinyl,
purinyl, pyranyl, pyrazinyl, pyrazolidinyl, pyrazolinyl, pyrazolyl,
pyridazinyl, pyridooxazole, pyridoimidazole, pyridothiazole,
pyridinyl, pyridyl, pyrimidinyl, pyrrolidinyl, pyrrolinyl,
pyrrolyl, quinazolinyl, quinolinyl, 4H-quinolizinyl, quinoxalinyl,
quinuclidinyl, carbolinyl, tetrahydrofuranyl,
tetrahydroisoquinolinyl, tetrahydroquinolinyl,
6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1,2,4-thiadiazolyl,
1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, thianthrenyl, thiazolyl,
thienyl, thienothiazolyl, thienooxazolyl, thienoimidazolyl,
thiophenyl, triazinyl, 1,2,3-triazolyl, 1,2,4-triazolyl,
1,2,5-triazolyl, 1,3,4-triazolyl, xanthenyl. Preferred heterocycles
include, but are not limited to, pyridinyl, furanyl, thienyl,
pyrrolyl, pyrazolyl, imidazolyl, indolyl, benzimidazolyl,
1H-indazolyl, oxazolidinyl, benzotriazolyl, benzisoxazolyl,
oxindolyl, benzoxazolinyl, or isatinoyl.
[0047] As used herein, "pharmaceutically acceptable salts" refer to
derivatives of the disclosed compounds wherein the parent compound
is modified by making acid or base salts thereof. Examples of
pharmaceutically acceptable salts include, but are not limited to,
mineral or organic acid salts of basic residues such as amines;
alkali or organic salts of acidic residues such as carboxylic
acids; and the like. The pharmaceutically acceptable salts include
the conventional non-toxic salts or the quaternary ammonium salts
of the parent compound formed, for example, from non-toxic
inorganic or organic acids. For example, such conventional
non-toxic salts include those derived from inorganic acids such as
hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric
and the like; and the salts prepared from organic acids such as
acetic, propionic, succinic, glycolic, stearic, lactic, malic,
tartaric, citric, ascorbic, palmitic, maleic, hydroxymaleic,
phenylacetic, glutamic, benzoic, salicylic, sulfanilic,
2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic, ethane
disulfonic, oxalic, isethionic, and the like.
[0048] The pharmaceutically acceptable salts of the present
invention can be synthesized from the parent compound which
contains a basic or acidic moiety by conventional chemical methods.
Generally, such salts can be prepared by reacting the free acid or
base forms of these compounds with a stoichiometric amount of the
appropriate base or acid in water or in an organic solvent, or in a
mixture of the two; generally, nonaqueous media like ether, ethyl
acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists
of suitable salts are found in Remington's Pharmaceutical Sciences,
18th ed., Mack Publishing Company, Easton, Pa., 1990, p. 1445, the
disclosure of which is hereby incorporated by reference.
[0049] The compounds of the present invention are useful in the
form of the free base or acid or in the form of a pharmaceutically
acceptable salt thereof. All forms are within the scope of the
invention.
[0050] The phrase "pharmaceutically acceptable" is employed herein
to refer to those compounds, materials, compositions, and/or dosage
forms which are, within the scope of sound medical judgment,
suitable for use in contact with the tissues of human beings and
animals without excessive toxicity, irritation, allergic response,
or other problem or complication commensurate with a reasonable
risk/benefit ratio.
[0051] The term "pharmaceutically acceptable prodrugs" as used
herein means those prodrugs of the compounds useful according to
the present invention which are, within the scope of sound medical
judgment, suitable for use in contact with the tissues of humans
and lower animals without undue toxicity, irritation, allergic
response, and the like, commensurate with a reasonable risk/benefit
ratio, and effective for their intended use, as well as
zwitterionic forms, where possible, of the compounds of the
invention.
[0052] The term "prodrugs", as the term is used herein, are
intended to include any covalently bonded carriers which release an
active parent drug of the present invention in vivo when such
prodrug is administered to a mammalian subject. Since prodrugs are
known to enhance numerous desirable qualities of pharmaceuticals
(i.e., solubility, bioavailability, manufacturing, etc.) the
compounds of the present invention may be delivered in prodrug
form. Thus, the skilled artisan will appreciate that the present
invention encompasses prodrugs of the presently claimed compounds,
methods of delivering the same, and compositions containing the
same. Prodrugs of the present invention are prepared by modifying
functional groups present in the compound in such a way that the
modifications are cleaved, either in routine manipulation or in
vivo, to form the parent compound. The transformation in vivo may
be, for example, as the result of some metabolic process, such as
chemical or enzymatic hydrolysis of a carboxylic, phosphoric or
sulphate ester, or reduction or oxidation of a susceptible
functionality. Prodrugs include compounds of the present invention
wherein a hydroxy, amino, or sulfhydryl group is bonded to any
group that, when the prodrug of the present invention is
administered to a mammalian subject, it cleaves to form a free
hydroxyl, free amino, or free sulfydryl group, respectively.
Functional groups which may be rapidly transformed, by metabolic
cleavage, in vivo form a class of groups reactive with the carboxyl
group of the compounds of this invention. They include, but are not
limited to such groups as alkanoyl (such as acetyl, propionyl,
butyryl, and the like), unsubstituted and substituted aroyl (such
as benzoyl and substituted benzoyl), alkoxycarbonyl (such as
ethoxycarbonyl), trialkylsilyl (such as trimethyl- and
triethysilyl), monoesters formed with dicarboxylic acids (such as
succinyl), and the like. Because of the ease with which the
metabolically cleavable groups of the compounds useful according to
this invention are cleaved in vivo, the compounds bearing such
groups can act as pro-drugs. The compounds bearing the
metabolically cleavable groups have the advantage that they may
exhibit improved bioavailability as a result of enhanced solubility
and/or rate of absorption conferred upon the parent compound by
virtue of the presence of the metabolically cleavable group. A
thorough discussion of prodrugs is provided in the following:
Design of Prodrugs, H. Bundgaard, ed., Elsevier, 1985; Methods in
Enzymology, K. Widder et al, Ed., Academic Press, 42, p.309-396,
1985; A Textbook of Drug Design and Development, Krogsgaard-Larsen
and H. Bundgaard, ed., Chapter 5; "Design and Applications of
Prodrugs" p.113-191, 1991; Advanced Drug Delivery Reviews, H.
Bundgard, 8, p.1-38, 1992; Journal of Pharmaceutical Sciences, 77,
p. 285, 1988; Chem. Pharm. Bull., N. Nakeya et al, 32, p. 692,
1984; Pro-drugs as Novel Delivery Systems, T. Higuchi and V.
Stella, Vol. 14 of the A.C.S. Symposium Series, and Bioreversible
Carriers in Drug Design, Edward B. Roche, ed., American
Pharmaceutical Association and Pergamon Press, 1987, each of which
is herein incorporated by reference in their entirety as though set
forth in full.
[0053] The term "treating" refers to: (i) preventing a disease,
disorder or condition from occurring in an animal which may be
predisposed to the disease, disorder and/or condition but has not
yet been diagnosed as having it; (ii) inhibiting the disease,
disorder or condition, i.e., arresting its development; and (iii)
relieving the disease, disorder or condition, i.e., causing
regression of the disease, disorder and/or condition.
PREPARATION OF COMPOUNDS OF THE INVENTION
[0054] Certain compounds of Formula I and intermediates used in
making these compounds may exhibit tautomerism and in some cases
one tautomer has been schematically drawn to represent all forms.
Certain compounds of formula I can exhibit isomerism, for example
geometrical isomerism, e.g., E or Z isomerism, and optical
isomerism, e.g., R or S configurations. Geometrical isomers include
the cis and trans forms of compounds of the invention having
alkenyl moieties. It is well known in the art how to prepare
optically active forms, such as by resolution of racemic forms or
by synthesis from optically active starting materials. All chiral,
diastereomeric, racemic forms and all geometric isomeric forms of a
structure are intended, unless the specific stereochemistry or
isomer form is specifically indicated.
[0055] Such isomers can be separated from their mixtures, by the
application or adaptation of known methods, for example
chromatographic techniques and recrystallization techniques, or
they are separately prepared from the appropriate isomers of their
intermediates, for example by the application or adaptation of
methods described herein.
[0056] Where the compound of the present invention is substituted
with a basic moiety, acid addition salts are formed and are simply
a more convenient form for use; and in practice, use of the salt
form inherently amounts to use of the free base form. The acids
which can be used to prepare the acid addition salts include
preferably those which produce, when combined with the free base,
pharmaceutically acceptable salts, that is, salts whose anions are
non-toxic to the patient in pharmaceutical doses of the salts, so
that the beneficial inhibitory effects on HCV RdRp inherent in the
free base are not vitiated by side effects ascribable to the
anions. Although pharmaceutically acceptable salts of said basic
compounds are preferred, all acid addition salts are useful as
sources of the free base form even if the particular salt, per se,
is desired only as an intermediate product as, for example, when
the salt is formed only for purposes of purification, and
identification, or when it is used as an intermediate in preparing
a pharmaceutically acceptable salt by ion exchange procedures.
[0057] According to a further feature of the invention, acid
addition salts of the compounds of this invention are prepared by
reaction of the free base with the appropriate acid, by the
application or adaptation of known methods. For example, the acid
addition salts of the compounds of this invention are prepared
either by dissolving the free base in aqueous or aqueous-alcohol
solution or other suitable solvents containing the appropriate acid
and isolating the salt by evaporating the solution, or by reacting
the free base and acid in an organic solvent, in which case the
salt separates directly or can be obtained by concentration of the
solution.
[0058] The acid addition salts of the compounds of this invention
can be regenerated from the salts by the application or adaptation
of known methods. For example, parent compounds of the invention
can be regenerated from their acid addition salts by treatment with
an alkali, e.g. aqueous sodium bicarbonate solution or aqueous
ammonia solution.
[0059] Where the compound of the invention is substituted with an
acidic moiety, base addition salts may be formed and can be simply
a more convenient form for use; and in practice, use of the salt
form inherently amounts to use of the free acid form. The bases
which can be used to prepare the base addition salts include those
which produce, when combined with the free acid, pharmaceutically
acceptable salts, that is, salts whose cations are non-toxic to the
animal organism in pharmaceutical doses of the salts, so that the
beneficial inhibitory effects on HCV RdRp inherent in the free acid
are not vitiated by side effects ascribable to the cations.
Pharmaceutically acceptable salts, including for example alkali and
alkaline earth metal salts, within the scope of the invention are
those derived from the following bases: sodium hydride, sodium
hydroxide, potassium hydroxide, calcium hydroxide, aluminum
hydroxide, lithium hydroxide, magnesium hydroxide, zinc hydroxide,
ammonia, ethylenediamine, N-methyl-glucamine, lysine, arginine,
ornithine, choline, N,N'-dibenzylethylenediamine, chloroprocaine,
diethanolamine, procaine, N-benzylphenethylamine, diethylamine,
piperazine, tris(hydroxymethyl)-aminomethane, tetramethylammonium
hydroxide, and the like.
[0060] Metal salts of compounds of the present invention may be
obtained by contacting a hydride, hydroxide, carbonate or similar
reactive compound of the chosen metal in an aqueous or organic
solvent with the free acid form of the compound. The aqueous
solvent employed may be water or it may be a mixture of water with
an organic solvent, preferably an alcohol such as methanol or
ethanol, a ketone such as acetone, an aliphatic ether such as
tetrahydrofuran, or an ester such as ethyl acetate. Such reactions
are normally conducted at ambient temperature but they may, if
desired, be conducted with heating.
[0061] Amine salts of compounds of the present invention may be
obtained by contacting an amine in an aqueous or organic solvent
with the free acid form of the compound. Suitable aqueous solvents
include water and mixtures of water with alcohols such as methanol
or ethanol, ethers such as tetrahydrofuran, nitriles such as
acetonitrile, or ketones such as acetone. Amino acid salts may be
similarly prepared.
[0062] The base addition salts of the compounds of this invention
can be regenerated from the salts by the application or adaptation
of known methods. For example, parent compounds of the invention
can be regenerated from their base addition salts by treatment with
an acid, e.g. hydrochloric acid.
[0063] Pharmaceutically acceptable salts also include quaternary
lower alkyl ammonium salts. The quaternary salts are prepared by
the exhaustive alkylation of basic nitrogen atoms in compounds,
including nonaromatic and aromatic basic nitrogen atoms, according
to the invention, i.e., alkylating the non-bonded pair of electrons
of the nitrogen moieties with an alkylating agent such as
methylhalide, particularly methyl iodide, or dimethyl sulfate.
Quaternarization results in the nitrogen moiety becoming positively
charged and having a negative counter ion associated therewith.
[0064] As will be self-evident to those skilled in the art, some of
the compounds of this invention do not form stable salts. However,
acid addition salts are more likely to be formed by compounds of
this invention having a nitrogen-containing heteroaryl group and/or
wherein the compounds contain an amino group as a substituent.
Preferable acid addition salts of the compounds of the invention
are those wherein there is not an acid labile group.
[0065] As well as being useful in themselves as active compounds,
salts of compounds of the invention are useful for the purposes of
purification of the compounds, for example by exploitation of the
solubility differences between the salts and the parent compounds,
side products and/or starting materials, by techniques well known
to those skilled in the art.
[0066] Compounds according to the invention, for example, starting
materials, intermediates or products, are prepared as described
herein or by the application or adaptation of known methods, by
which is meant methods used heretofore or described in the
literature, for example those described by R. C. Larock in
Comprehensive Organic Transformations, VCH publishers, 1989.
[0067] In the reactions described hereinafter it may be necessary
to protect reactive functional groups, for example hydroxy, amino,
imino, thio or carboxy groups, where these are desired in the final
product, to avoid their unwanted participation in the reactions.
Conventional protecting groups may be used in accordance with
standard practice, for examples see T. W. Green and P. G. M. Wuts
in "Protective Groups in Organic Chemistry" John Wiley and Sons,
1991; J. F. W. McOmie in "Protective Groups in Organic Chemistry"
Plenum Press, 1973.
[0068] The compounds useful according to the invention optionally
are supplied as salts. Those salts which are pharmaceutically
acceptable are of particular interest since they are useful in
administering the foregoing compounds for medical purposes. Salts
which are not pharmaceutically acceptable are useful in
manufacturing processes, for isolation and purification purposes,
and in some instances, for use in separating stereoisomeric forms
of the compounds of this invention. The latter is particularly true
of amine salts prepared from optically active amines.
[0069] Where the compound useful according to the invention
contains a carboxy group, or a sufficiently acidic bioisostere,
base addition salts may be formed and are simply a more convenient
form for use; and in practice, use of the salt form inherently
amounts to use of the free acid form.
[0070] Also, where the compound useful according to the invention
contains a basic group, or a sufficiently basic bioisostere, acid
addition salts may be formed and are simply a more convenient form
for use; and in practice, use of the salt form inherently amounts
to use of the free base form.
[0071] The foregoing compounds useful according to the invention
may also be combined with another therapeutic compound to form
pharmaceutical compositions (with or without diluent or carrier)
which, when administered, provide simultaneous administration of
two or more active ingredients resulting in the combination therapy
of the invention.
[0072] While it is possible for compounds useful according to the
invention to be administered alone it is preferable to present them
as pharmaceutical compositions. The pharmaceutical compositions,
both for veterinary and for human use, useful according to the
present invention comprise at lease one compound of the invention,
as above defined, together with one or more acceptable carriers
therefor and optionally other therapeutic ingredients. The skilled
artisan will appreciate the abundance of publications setting forth
the state of the art for pharmaceutical administration.
[0073] Examples of suspending agents include ethoxylated isostearyl
alcohols, polyoxyethylene sorbitol and sorbitan esters,
microcrystalline cellulose, aluminum metahydroxide, bentonite,
agar-agar and tragacanth, or mixtures of these substances.
Prevention of the action of microorganisms can be ensured by
various antibacterial and antifungal agents, for example, parabens,
chlorobutanol, phenol, sorbic acid, and the like. It may also be
desirable to include isotonic agents, for example sugars, sodium
chloride and the like. Prolonged absorption of the injectable
pharmaceutical form can be brought about by the use of agents
delaying absorption, for example, aluminum monosterate and gelatin.
Examples of suitable carriers, diluents, solvents or vehicles
include water, ethanol, polyols, suitable mixtures thereof,
vegetable oils (such as olive oil) and injectable organic esters
such as ethyl oleate. Examples of excipients include lactose, milk
sugar, sodium citrate, calcium carbonate, dicalcium phosphate
phosphate. Examples of disintegrating agents include starch,
alginic acids and certain complex silicates. Examples of lubricants
include magnesium stearate, sodium lauryl sulphate, talc, as well
as high molecular weight polyethylene glycols.
[0074] In certain preferred embodiments, active ingredients
necessary in combination therapy may be combined in a single
pharmaceutical composition for simultaneous administration.
[0075] The choice of vehicle and the content of active substance in
the vehicle are generally determined in accordance with the
solubility and chemical properties of the active compound, the
particular mode of administration and the provisions to be observed
in pharmaceutical practice. For example, excipients such as
lactose, sodium citrate, calcium carbonate, dicalcium phosphate and
disintegrating agents such as starch, alginic acids and certain
complex silicates combined with lubricants such as magnesium
stearate, sodium lauryl sulphate and talc may be used for preparing
tablets. To prepare a capsule, it is advantageous to use lactose
and high molecular weight polyethylene glycols. When aqueous
suspensions are used they can contain emulsifying agents or agents
which facilitate suspension. Diluents such as sucrose, ethanol,
polyethylene glycol, propylene glycol, glycerol and chloroform or
mixtures thereof may also be used.
[0076] The oily phase of the emulsions of this invention may be
constituted from known ingredients in a known manner. While the
oily phase may comprise merely an emulsifier (otherwise known as an
emulgent), it desirably comprises a mixture of at least one
emulsifier with a fat or an oil or with both a fat and an oil.
Preferably, a hydrophilic emulsifier is included together with a
lipophilic emulsifier which acts as a stabilizer. It is also
preferred to include both an oil and a fat. Together, the
emulsifier(s) with or without stabilizer(s) make up the emulsifying
wax, and the wax together with the oil and fat make up the
emulsifying ointment base which forms the oily dispersed phase of a
cream formulation. Emulgents and emulsion stabilizers suitable for
use in the formulation of the present invention include Tween.RTM.
60, Span.RTM. 80, cetostearyl alcohol, benzyl alcohol, myristyl
alcohol, glyceryl mono-stearate and sodium lauryl sulfate.
[0077] If desired, the aqueous phase of the cream base may include,
for example, a least 30% w/w of a polyhydric alcohol, i.e. an
alcohol having two or more hydroxyl groups such as propylene
glycol, butane 1,3-diol, mannitol, sorbitol, glycerol and
polyethylene glycol (including PEG 400) and mixtures thereof. The
topical formulations may desirably include a compound which
enhances absorption or penetration of the active ingredient through
the skin or other affected areas. Examples of such dermal
penetration enhancers include dimethyl sulphoxide and related
analogues.
[0078] The choice of suitable oils or fats for the formulation is
based on achieving the desired cosmetic properties. Thus the cream
should preferably be a non-greasy, non-staining and washable
product with suitable consistency to avoid leakage from tubes or
other containers. Straight or branched chain, mono- or dibasic
alkyl esters such as di-isopropyl myristate, decyl oleate,
isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or a
blend of branched chain esters known as Crodamol CAP may be used,
the last three being preferred esters. These may be used alone or
in combination depending on the properties required. Alternatively,
high melting point lipids such as white soft paraffin and/or liquid
paraffin or other mineral oils can be used. Solid compositions may
also be employed as fillers in soft and hard-filled gelatin
capsules using such excipients as lactose or milk sugar as well as
high molecular weight polyethylene glycols, and the like.
[0079] The pharmaceutical compositions can be administered in a
suitable formulation to humans and animals by topical or systemic
administration, including oral, inhalational, rectal, nasal,
buccal, sublingual, vaginal, parenteral (including subcutaneous,
intramuscular, intravenous, intradermal, intrathecal and epidural),
intracisternal and intraperitoneal. It will be appreciated that the
preferred route may vary with for example the condition of the
recipient.
[0080] The formulations can be prepared in unit dosage form by any
of the methods well known in the art of pharmacy. Such methods
include the step of bringing into association the active ingredient
with the carrier which constitutes one or more accessory
ingredients. In general the formulations are prepared by uniformLy
and intimately bringing into association the active ingredient with
liquid carriers or finely divided solid carriers or both, and then,
if necessary, shaping the product.
[0081] A tablet may be made by compression or moulding, optionally
with one or more accessory ingredients. Compressed tables may be
prepared by compressing in a suitable machine the active ingredient
in a free-flowing form such as a powder or granules, optionally
mixed with a binder, lubricant, inert diluent, preservative,
surface active or dispersing agent. Moulded tablets may be made by
moulding in a suitable machine a mixture of the powdered compounds
moistened with an inert liquid diluent. The tablets may optionally
be coated or scored and may be formulated so as to provide slow or
controlled release of the active ingredient therein.
[0082] Solid compositions for rectal administration include
suppositories formulated in accordance with known methods and
containing at least one compound of the invention.
[0083] If desired, and for more effective distribution, the
compounds can be microencapsulated in, or attached to, a slow
release or targeted delivery systems such as a biocompatible,
biodegradable polymer matrices (e.g. poly(d,1-lactide
co-glycolide)), liposomes, and microspheres and subcutaneously or
intramuscularly injected by a technique called subcutaneous or
intramuscular depot to provide continuous slow release of the
compound(s) for a period of 2 weeks or longer. The compounds may be
sterilized, for example, by filtration through a bacteria retaining
filter, or by incorporating sterilizing agents in the form of
sterile solid compositions which can be dissolved in sterile water,
or some other sterile injectable medium immediately before use.
[0084] Actual dosage levels of active ingredient in the
compositions of the invention may be varied so as to obtain an
amount of active ingredient that is effective to obtain a desired
therapeutic response for a particular composition and method of
administration. The selected dosage level therefore depends upon
the desired therapeutic effect, on the route of administration, on
the desired duration of treatment and other factors.
[0085] Total daily dose of the compounds useful according to this
invention administered to a host in single or divided doses may be
in amounts, for example, of from about 0.0001 to about 100 mg/kg
body weight daily and preferably 0.01 to 10 mg/kg/day. Dosage unit
compositions may contain such amounts of such submultiples thereof
as may be used to make up the daily dose. The skilled artisan will
appreciate that the specific dose level for any particular patient
will depend upon a variety of factors including the patient's body
weight, general health, sex, diet, time and route of
administration, rates of absorption and excretion, combination with
other drugs and the severity of the particular disease being
treated.
[0086] The amount of each component administered is determined by
the attending clinicians taking into consideration the etiology and
severity of the disease, the patient's condition and age, the
potency of each component and other factors.
[0087] The formulations may be presented in unit-dose or multi-dose
containers, for example sealed ampoules and vials with elastomeric
stoppers, and may be stored in a freeze-dried (lyophilized)
condition requiring only the addition of the sterile liquid
carrier, for example water for injections, immediately prior to
use. Extemporaneous injection solutions and suspensions may be
prepared from sterile powders, granules and tablets of the kind
previously described.
[0088] Administration of a compound of the present invention in
combination with additional therapeutic agents, may afford an
efficacy advantage over the compounds and agents alone, and may do
so while permitting the use of lower doses of each. A lower dosage
minimizes the potential of side effects, thereby providing an
increased margin of safety. The combination of a compound of the
present invention with such additional therapeutic agents is
preferably a synergistic combination. Synergy, as described for
example by Chou and Talalay, Adv. Enzyme Regul. 22:27-55 (1984),
occurs when the therapeutic effect of the compound and agent when
administered in combination is greater than the additive effect of
either the compound or agent when administered alone. In general, a
synergistic effect is most clearly demonstrated at levels that are
(therapeutically) sub-optimal for either the compound of the
present invention or a known antiviral agent alone, but which are
highly efficacious in combination. Synergy can be in terms of
improved inhibitory response without substantial increases in
toxicity over individual treatments alone, or some other beneficial
effect of the combination compared with the individual
components.
[0089] Procedures for evaluating the biological activity of
compounds or compositions according to the invention are carried
out as described herein or by the application or adaptation of
procedures well known in the art as described in the literature.
The compounds of the present invention, their methods or
preparation and their biological activity will appear more clearly
from the examination of the following examples which are presented
as an illustration only and are not to be considered as limiting
the invention in its scope. The following examples are but
preferred methods of synthesizing the compounds of the invention,
which may be prepared according to any method known to the organic
chemist of ordinary skill. Other features of the invention will
become apparent during the following descriptions of exemplary
embodiments which are given for illustration of the invention and
are not intended to be limiting thereof. Each of the patents,
patent applications, and other cited references, are hereby
incorporated herein by reference in their entirity as though set
forth in full.
Synthesis
[0090] The compounds of the present invention can be prepared in a
number of ways well known to one skilled in the art of organic
synthesis. The compounds of the present invention can be
synthesized using the methods described below, together with
methods known in the art of synthetic organic chemistry, or
variations thereon as appreciated by those skilled in the art. A
particularly useful compendium of synthetic methods which may be
applicable to the preparation of compounds of the present invention
may be found in Larock, R. C. Comprehensive Organic
Transformations, VCH: New York, 1989. Preferred methods include,
but are not limited to, those described below. All references cited
herein are hereby incorporated in their entirety herein by
reference.
[0091] The novel compounds of this invention may be prepared using
the reactions and techniques described in this section. The
reactions are performed in solvents appropriate to the reagents and
materials employed and are suitable for the transformations being
effected. Also, in the description of the synthetic methods,
described below, it is to be understood that all proposed reaction
conditions, including choice of solvent, reaction atmosphere,
reaction temperature, duration of the experiment and workup
procedures, are chosen to be the conditions standard for that
reaction, which should be readily recognized by one skilled in the
art. It is understood by one skilled in the art of organic
synthesis that the functionality present on various portions of the
molecule must be compatible with the reagents and reactions
proposed. Such restrictions to the substituents which are
compatible with the reaction conditions will be readily apparent to
one skilled in the art and alternate methods must then be used. In
addition, it may be necessary to introduce or remove protecting
groups in order to carry certain substituents through the indicated
reaction conditions. A compendium of protecting groups which may be
useful, together with reaction conditions for introduction and
removal may be found in Greene, T. W.; Wuts, P. G. M. Protective
Groups in Organic Synthesis, Second Edition; Wiley: New York,
1991.
[0092] The starting materials and all reagents and intermediates
for these synthetic routes are either commercially available or may
be prepared by methods known to one skilled in the art of organic
synthesis from commercially available materials.
[0093] The following methods describe different preparations of the
compounds of the present invention. The methods are often general
in nature and may be used to make variations of the inventive
embodiments. Other variations would be apparent to those skilled in
the art.
[0094] Method A describes a general method of preparing compounds
of Formula I (Scheme 1). Amine 3 is produced by nucleophilic
displacement of aryl chloride 1 by cyclohexylamine (2). Reduction
of the nitro group of 3 produces amine 4 which is condensed with
imidate 5 to provide benzimidazole 6. The phenol of 6 can be
alkylated with a variety of agents to form Formula I compounds.
These agents include diphenylmethylbromide (7) which generates
nitrile 8. Transformation of the nitrile moiety of 8 to a tetrazole
yields 9, an example of a Formula I compound.
[0095] Additionally, the nitrile of 8 may be converted to a
5-oxo-1,2,4-oxadiazole by reaction with hydroxylamine followed by
reaction with methylchloroformate or carbonyldiimidazole. The
nitrile may also be converted to a 2-oxo-1,2,3,5-oxathiadiazole by
reaction with hydroxylamine followed by reaction with thionyl
chloride and pyridine. 8
[0096] Method B provides an alternative method of preparing Formula
I compounds (Scheme 2). Amine 4 can be condensed with a range of
acid chlorides including 10 to form amides such as 11 which are
cyclized to form benzimidazoles of which 12 is a representative
example. Conversion of the nitrile moiety of 12 into a tetrazole
provides compound 13, which is an example of a Formula I compound.
9
[0097] Method C describes an additional alkylative preparation of a
Formula I compound. The alkylating agent 14 is prepared by coupling
4-chlorophenylboronic acid with 2-bromo-5-methoxytoluene which is
in turn brominated on the tolyl methyl group. Many of the
benzyl-type alkylating agents can be prepared by this route. 10
[0098] Method D provides an alternative route to Formula I
compounds and describes the preparation of compounds where R.sup.2
is --CH.sub.2C.sub.6H.sub.3R.sup.4R.sup.6 and R.sup.4 is Ar.sup.2
or Ar.sup.3 (Scheme 4). Phenol 6 is protected as the
tert-butyldimethylsilyl (TBDMS) ether 20. The nitrile moiety of 20
is then converted to tetrazole 21 and the tetrazole protected with
a trityl group. Subsequent removal of the TBDMS protecting group
with tetrabutylammonium fluoride (TBAF) affords intermediate 22
which is alkylated with a 3-bromomethyl-4-bromo benzoate ester
(with, for example, R=methyl or tert-butyl). Reasonable variations
of this agent (for example with a triflate replacing the aryl
bromide or substituting a close analog of the alkyl ester) would be
known to those skilled in the art. The resulting intermediate 23
may be used as a coupling partner with various organometallic
compounds to afford compounds similar to 25. Trityl deprotection
affords ester 26. Hydrolysis of this ester provides acids also of
Formula 26 (R=H). The esters and acids are examples of Formula I
compounds. 1112
[0099] Method E describes the use of solid phase technology for the
preparation of compounds of Formula I (Scheme 5). Phenol 27 is
attached to a polymeric support, such as the Merrifield resin, and
converted into an appropriate linker such as 30. Intermediate
tetrazole 21 is then tethered to this linker. Deprotection and
alkylation as described previously provide compounds like 34a.
Cleavage from the resin affords compound 34, which is an example of
a Formula I compounds. 13
[0100] Method F provides an additional solid phase approach which
utilizes chloro trityl resin 35 (scheme 6). 14
[0101] Method G provides for a preparation of Formula I compounds
where R6 is CONR.sup.7R.sup.8 (scheme 7). Tetrazole 23 can be
coupled with a variety of organometallic compounds to afford
compounds like 39. Hydrolysis of the methyl ester of 39 gives
carboxylic acid 40. The carboxylic acid moiety is transformed to an
amide using amines of Formula HNR.sup.7R.sup.8. In this scheme the
amine is an alkyl ester of glycine, which is then hydrolyzed. The
protecting group of 42 is then removed to provide compound 43. The
esters, acids, and amides are examples of Formula I compounds.
15
[0102] Method H provides a preparation of various R.sup.1
substituents (scheme 8). Compounds of Formula 48 may be made in an
analogous fashion to those of Formula 8 or 12. By judicious choice
of the ester moiety of 48, ester hydrolysis may be carried out
under basic (R is methyl), acidic (R is t-butyl), or neutral (R is
benzyl) conditions to provide compounds like 49. Acid 49 may be
coupled with a sulfonamide via the acid chloride to provide
compounds like 55. In a similar manner, other acylsulfonamides
provide additional examples of Formula I compounds. 16
[0103] Method I provides methods for making compounds where Q is N
(scheme 9). Where appropriate, these methods may be used in the
previous procedures to prepare other examples of formula I
compounds. 17
[0104] Method J provides a method for preparing Formula I compounds
where R.sup.1 is a RCONHSO.sub.2-moiety (Scheme 10).
[0105] The preparation of the primary sulfonamide 69 follows the
previous methods. Acylation of the sulfonamide can be accomplished
by treating the anion of the sulfonamide with an appropriate
acylating agent. 18
DESCRIPTION OF THE SPECIFIC EMBODIMENTS
[0106] Abbreviations used in the examples are defined as follows:
"1.times." for once, "2.times." for twice, "3.times." for thrice,
"rt" for room temperature, "eq" for equivalent or equivalents, "g"
for gram or grams, "mg" for milligram or milligrams, "mL" for
milliliter or milliliters, "M" for molar, N for normal, "mmol" for
millimole or millimoles, "min" for minute or minutes, "h" for hour
or hours, "MS" for mass spectrometry, "ESI" for electrospray
ionization, "NMR" for nuclear magnetic resonance spectroscopy,
".sup.1H" for proton, ".delta." for delta, "s" for singlet, "d" for
doublet, "t" for triplet, "q" for quartet, "m" for multiplet, "br"
for broad, "Hz" for hertz, "HPLC" for high pressure liquid
chromatography, "ELSD" for evaporative light scattering detector,
"tlc" for thin layer chromatography, "v/v" for volume to volume
ratio, "atm" for atmosphere, "psi" for pounds per square inch,
".alpha.", ".beta.", "R", and "S" are stereochemical designations
familiar to one skilled in the art. DMF is N,N-dimethylformamide,
THF is tetrahydrofuran. Temperatures are expressed in degrees
Celsius.
[0107] The majority of the final compounds were purified by reverse
phase chromatography using a preparative C-18 column employing
gradients of methanol--water containing 0.1% of trifluoroacetic
acid (TFA), and using a Shimadzu High Perfomance Liquid Preparative
Chromatographic System employing an XTERRA 3.0.times.50 mm S7
column at 5 mL/min flow rate with a 2 min gradient. The final
compounds were usually isolated and submitted for biological
evaluations as their acid addition salts with trifluoroacetic acid.
Molecular weights and purities were usually determined using a
Shimadzu LCMS. NMR spectra were usually obtained on either a Bruker
500 or 300 MHz instrument.
Intermediate 3
4-(Cyclohexylamino)-3-nitrobenzonitrile (3)
[0108] A solution of 4-chloro-3-nitrobenzonitrile (5.28 g, 28.9
mmol), cyclohexylamine (5.0 mL, 43.7 mmol), and triethylamine (6.0
mL, 43.0 mL) in acetonitrile (75 mL) was stirred at 50.degree. C.
for 15 h. The reaction mixture was cooled to rt and then poured
into 100 mL ice water. The solid precipitate was filtered, washed
with water, and dried to afford compound 3 as a yellow solid (6.42
g, 90%). ESI-MS m/e 246.3 (M+1).
Intermediate 4
3-Amino-4-(cyclohexylamino)benzonitrile (4)
[0109] A solution of compound 3 (1.23 g, 5.01 mmol) in ethyl
acetate (30 mL) and methanol (10 mL) was hydrogenated over 10%
palladium on carbon (0.11 g) at 20 psi for 1 h. The reaction
mixture was filtered and concentrated on a rotary evaporator to
give compound 4 (1.07 g, 99%) as a red-brown solid. ESI-MS m/e
216.3 (M+1).
Intermediate 6
2-[4-Hydroxyphenyl]-1-cyclohexyl-1H-benzimidazole-5-carbonitrile
(6)
[0110] A solution of ethyl 4-hydroxybenzimidate hydrochloride (5,
1.26 g, 6.7mmol) and compound 4 (1.5 g, 5.0 mmol) in methanol (10
mL) was refluxed overnight under a nitrogen atmosphere. The
reaction mixture was cooled to rt, filtered, and washed with
methanol to afford 6 as a pinkish brown solid (1.26 g, 68%). ESI-MS
m/e 372.1 (M+1).
Intermediate 8
2-[4-(Diphenylmethoxy)phenyl]-1-cyclohexyl-1H-benzimidazole-5-carbonitrile
(8)
[0111] A 60% dispersion of sodium hydride in mineral oil (50 mg,
1.25 mmol) was added in a single portion to a stirred mixture of
compound 6 in DMF (3.5 mL). When the rapid evolution of hydrogen
had ceased, the mixture was cooled in an ice-water bath.
Diphenylmethyl bromide (297 mg, 1.2 mmol) was added. The mixture
was stirred at an oil bath temperature of 55.degree. C. for 30 min.
The mixture was cooled and poured into cold water to precipitate a
solid. The solid was dried and crystallized from ethyl
acetate-hexanes to afford 8 as off white needles. ESI-MS m/e 484
(M+1).
EXAMPLE 1
2-[4-(Diphenylmethoxy)phenyl]-1-cyclohexyl-5-(1H-tetrazol-5-yl)-1H-benzimi-
dazole (9)
[0112] Aluminum trichloride (33 mg, 0.247 mmol) was added to ice
cold THF (4 mL). Sodium azide (48 mg, 0.744 mmol) was added and the
resulting mixture was stirred under reflux for 30 min to complete
the formation of the aluminum azide. Compound 8 (100 mg, 0.206
mmol) was added and the mixture stirred under reflux for 18 h. The
reaction mixture was poured into ice cold dilute HCl to precipitate
a solid. The solid was dissolved in DMF (2 mL) and the solution
applied to a Shimadzu Prep HPLC. The product containing fractions
were combined. Removal of the methanol resulted in the
precipitation of 9 as a colorless solid (22.3 mg, 20.6% yield).
ESI-MS m/e 527 (M=1). .sup.1H NMR (DMSO) .delta.8.33 (s, 1H), 8.20
(d, 1H, J=8.6 Hz), 8.0 (1H, d, J=7.41 Hz), 7.63 (d, 2H, J=8.7 Hz),
7.55 (d, 4H, J=7.2 Hz), 7.41-7.37 (m, 4H), 7.34-7.26 (m, 4H). 6.72
(s, 1H), 4.34-4.27 (m, 1H),2.36-2.23 (m, 2H), 1.97-1.83 (m, 4H),
1.66-1.62 (m, 1H), 1.43-1.21 (m, 3H).
Intermediate 10
4-(Benzyloxy)benzoyl Chloride (10)
[0113] A solution of 4-benzyloxybenzoic acid (2.50 g, 1.0 mmol) and
thionyl chloride (1.2 mL, 16 mmol) in benzene (35 mL) was refluxed
for 3 h under an argon atmosphere. The reaction mixture was cooled
to rt and concentrated on a rotary evaporator. The residue was
re-evaporated with benzene (2.times.) to afford the acid chloride
as a white solid.
Intermediate 11
4-(Benzyloxy)-N-[5-cyano-2-(cyclohexylamino)phenyl]benzamide
(11)
[0114] A solution of the acid chloride of 4-benzyloxybenzoic acid
(2.56 g, 10.4 mmol) in dichloromethane (30 mL) and
dimethylformamide (5 mL) was added dropwise to a solution of
compound 4 (2.23 g, 10.4 mmol) and triethylamine (2.4 mL, 17.2
mmol) in dichloromethane (15 mL). After 1.5 h,
4-dimethylaminopyridine (0.12 g) was added. The reaction was
stirred at rt for 18 h and then heated to 45.degree. C. for 4 h.
The reaction was cooled to rt and then extracted with 1 N
KHSO.sub.4 (1.times.) and brine (1.times.). The organic layer was
dried (MgSO.sub.4) and concentrated on a rotary evaporator. The
residue was purified by column chromatography (silica gel, 10-20%
ethyl acetate/hexanes (v/v)) to afford compound 11 as a slightly
purple solid (2.08 g, 47%). ESI-MS m/e 426.3 (M+1).
Intermediate 12
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazole-5-carbonitrile
(12)
[0115] A solution of compound 11 (2.08 g, 4.89 mmol) in acetic acid
(15 mL) was refluxed for 4.5 h. The reaction mixture was cooled to
rt and concentrated on a rotary evaporator. The residue was
dissolved in ethyl acetate and washed with saturated NaHCO.sub.3
(2.times.) and water (1.times.). The organic layer was dried
(MgSO.sub.4) and concentrated on a rotary evaporator. The residue
was recrystallized from hexane/ethyl acetate to give compound 12 as
a tan solid (1.8 g, 90%). ESI-MS m/e 408.3 (M+1).
EXAMPLE 2
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-5-(1H-tetrazol-5-yl)-1H-benzimidazole
(13)
[0116] A suspension of compound 12 (0.041 g, 0.10 mmol) and
trimethyltin azide (0.023 g, 0.11 mmol) in toluene (2 mL) was
refluxed for 15 h. The reaction mixture was allowed to stand at rt
for 48 h. The resulting suspension was filtered and washed with
toluene (2.times.). The solid was treated with hydrogen chloride (4
M in dioxane, several mL) for 4 h at rt. The reaction mixture was
concentrated on a rotary evaporator. The residue was purified by
reverse phase HPLC (C18 column, water/acetonitrile gradient
containing 0.05% trifluoroacetic acid) to afford 13 as a white
solid (0.009 g, 20%). .sup.1H NMR (CDCl.sub.3) .delta.8.22 (br s,
1H), 8.05 (br m, 1H), 7.82 (br m, 3H), 7.52-7.38 (m, 5H), 7.30 (br
m, 2H), 5.23 (s, 2H), 4.53 (br m, 1H), 2.27 (br m, 4H), 2.06 (br m,
2H), 1.84 (br m, 1H), 1.40 (br m, 3H); ESI-MS m/e 451.3 (M+1); HPLC
purity (ELSD) >95%.
Intermediate 14
4'-Chloro-2-methyl-1,1'-biphenyl-4-yl)methyl ether
(2 Steps)
[0117] (Step 1) A solution of 4-bromo-3-methylanisole (6.48 g, 32.2
mmol), 4-chlorophenyl-boronic acid (6.02 g, 38.5 mmol), tri-o-tolyl
phosphine (0.98 g, 3.2 mmol), and NaHCO.sub.3 (11.0 g, 130.9 mmol)
in ethylene glycol dimethyl ether (100 mL) and water (33 mL) was
degassed with a stream of nitrogen for 20 min. Palladium (II)
acetate (0.361 g, 1.6 mmol) was added under a nitrogen atmosphere.
The reaction mixture was heated at 90.degree. C. for 2 h. The
mixture was cooled to rt, and the organic layer was washed with
water (1.times.), brine (1.times.), dried (MgSO.sub.4) and
concentrated on a rotary evaporator. The residue was suspended in
hexane and filtered. The filtrate was concentrated and purified by
column chromatography (silica gel, eluting with hexanes, followed
by 10% ethyl acetate/hexane (v/v) to give the product as a clear
oil (5.2 g, 70%). ESI-MS m/e 233.1 (M+1).
2-(Bromomethyl)-4'-chloro-4-methoxy-1,1'-biphenyl (14)
[0118] (Step 2) A solution of the previous product(5.2 g, 22.3
mmol), N-bromosuccinimide (4.12 g, 23.1 mmol), AIBN (0.183 g, 1.11
mmol) in carbon tetrachloride (100 mL) was refluxed for 1 h. The
mixture was cooled to rt, filtered through a silica plug and
concentrated on a rotary evaporator. The residue was dissolved in
hexane and cooled in an ice bath. The white crystals were filtered
to give 14 (3.6 g, 52%). ESI-MS m/e 311.1(M+1).
Intermediate 15
2-{4-[(4'-Chloro-4-methoxy-1,1'-biphenyl-2-yl)methoxy]phenyl}-1-cyclohexyl-
-1H-benzimidazole-5-carbonitrile (15)
[0119] A suspension of compound 6 (0.330 g, 1.04 mmol) in DMF (2.4
mL) and cesium carbonate (339 mg, 1.04 mmol) was heated at
40.degree. C. until almost clear. Compound 14 (356 mg, 1.14 mmol)
was added in portions. The reaction mixture was stirred at rt for
18 h. The mixture was concentrated on a rotary evaporator. The
residue was washed with water and collected by filtration. This
material was purified by column chromatography (silica gel, 50% to
100% hexane/ethyl acetate (v/v) to give compound 15 (0.55 g, 100%
yield). ESI-MS m/e 548 (M=1).
EXAMPLE 3
2-{4-[(4'-Chloro-4-methoxy-1,1'-biphenyl-2-yl)methoxy]phenyl}-1-cyclohexyl-
-5-(1H-tetrazol-5-yl)-1H-benzimidazole (16)
[0120] Compound 15 (0.110 g, 0.20 mmol) was converted to compound
16 (0.012 g, 10%) by the same method used in example 2. .sup.1H NMR
(CDCl.sub.3) .delta.8.18 (br s, 1H), 7.98 (d, 2H, J=8.7 Hz ), 7.67
(d, 2H, J=8.8 Hz ), 7.36-7.18 (m, 7H), 7.11 (d, 1H, J=2.6 Hz ),
6.92 (dd, 1H, J=2.9, 8.5 Hz ), 6.81 (d, 2H, J=8.4 Hz), 4.85 (s,
2H), 4.25 (br m, 1H), 3.38 (s, 3H), 2.28 (br m, 4H), 2.05 (br m,
2H), 1.80 (br m, 1H), 1.35 (br m, 3H); ESI-MS m/e 591.2 (M+1); HPLC
purity (UV) >95%.
Intermediate 20
2-[4-(tert-Butyldimethylsilyoxy)phenyl]-1-cyclohexyl-1H-benzimidazole-5-ca-
rbonitrile (20)
[0121] Imidazole (2.72 g, 0.040 mol) and tert-butyldimethylchloro
silane (2.89 g, 192 mmol) were added to a stirred solution of
compound 6 (5.1 g, 16 mmol) in DMF (10 mL). Stirring was continued
for 15 h at 100.degree. C. Upon cooling 20 crystallized. The
crystals were collected and washed with ethyl acetate followed by
water. The yield of 20 (after drying) was 4.70 g (62%).
Intermediate (21)
2-[4-(tert-Butyldimethylsilyoxy)phenyl]-1-cyclohexyl-5-(1H-tetrazol-5-yl)--
1H-benzimidazole (21)
[0122] Trimethylaluminum (20 mL of a 2.0 M solution in toluene,
0.040 mol) was added to a dry flask. Azidotrimethylsilane (5.3 mL,
0.040 mol) and compound 20 were added. The solution was stirred for
18 h at an oil bath temperature of 95.degree. C. The solution was
cooled and poured into a mixture of ethyl acetate and ice. The
mixture was carefully acidified with dilute hydrochloric acid
(caution-done in hood, hydrazoic acid liberated) to precipitate
compound 21. The compound was crystallized from ethyl
acetate--hexanes to afford 21 as a pinkish solid (4.13 g, 87%).
Intermediate 22
2-[4-Hydroxyphenyl]-1-cyclohexyl-5-[2-(trityl)-2H-tetrazol-5-yl]-1H-benzim-
idazole (22)
[0123] Trityl chloride (4.57 g, 0.0164 mol) and
N,N-diisopropylethylamine (2.42 g, 0.0187 mol) were added to a
stirred mixture of compound 21 (7.4 g, 0.0156 mol) in DMF (36 mL).
The mixture was stirred for 18 h at an oil bath temperature of
70.degree. C. The mixture was cooled and poured into a mixture of
ethyl acetate and water to precipitate the fully protected
benzimidazole,
2-[4-(tert-butyldimethylsilyoxy)phenyl]-1-cyclohexyl-5-[2--
(trityl)-2H-tetrazol-5-yl]-1H-benzimidazole. The solid was
collected, washed with cold water and dried to afford the protected
benzimidazole as a colorless solid (9.1 g, 81% yield). ESI-MS m/e
717 (M+1). HPLC purity (uv) 94%.
[0124] An additional 770 mg was obtained from the ethyl acetate
layer after concentration on a rotary evaporator.
[0125] A 1M solution of tetrabutylammonium fluoride in THF (34 mL,
0.034 mol) was added in one portion to stirred mixture of the fully
protected benzimdazole (9.8 g, 0.0137 mol) in THF (100 mL). The
mixture was stirred at rt for 40 min when acetic acid (2.0 mL) was
added. Stirring was continued for an additional 5 min. The mixture
was poured into ethyl acetate-water. The organic layer was washed
(water, dilute sodium bicarbonate, and brine) and partially
concentrated on a rotary evaporator. Diethyl ether was added to the
resulting slurry to fully precipitate 22 as a pinkish solid (8.2 g,
100% yield). ESI-MS M/e 603 (M+1).
Methyl 4-bromo-3-(bromomethyl)benzoate
[0126] A stirred mixture of methyl 4-bromo-3-methylbenzoate (25.0
g, 0.109 mol) in carbon tetrachloride (250 mL) was warmed to near
boiling when N-bromosuccinimide 21.4 g, 0.12 mol) and
2,2'-azobisisobutyronitrile (250 mg) were added. The mixture was
stirred under reflux for 18 h, during which time it was irradiated
with a 200 W light bulb. The mixture was cooled and filterd to
remove the succinimde. Partial concentration on a rotating
evaporator resulted in crystallization of the methyl ester (18.4 g,
73.7% yield). .sup.1H NMR (CDCl.sub.3) .delta.8.10 (s, 1H), 7.81(d,
1H, J=8 Hz), 7.64 (d, 1H,J=8 Hz), 4.61 (s, 2H),3.95 (s, 3H).
tert-Butyl 4-bromo-3-(bromomethyl)benzoate
[0127] A suspension of 4-bromo-3-methylbenzoic acid (10 g, 46.5
mmol) was heated to 80.degree. C. and N,N-dimethylformamide
di-tert-butyl acetal (44.6 mL, 186.0 mmol) was added dropwise over
30 minutes under a nitrogen atmosphere; stirring was continued at
this temperature for 1.5 h. The reaction mixture was allowed to
cool and then washed with water (1.times.), saturated NaHCO.sub.3
(1.times.), and brine (1.times.). The organic layer was dried
(Na.sub.2SO.sub.4) and concentrated to afford tert-Butyl
4-bromo-3-methylbenzoate (as an off-white oil (8.8 g, 70%). ESI-MS
m/e 311.9 (M+CH.sub.3CN+1).
[0128] A stirred mixture of tert-butyl 4-bromo-3-methylbenzoate
(25.0 g, 0.116 mol) in carbon tetrachloride (250 mL) was heated to
near boiling when N-bromosuccinimide (18.34 g, 0.103 mol) and
2,2'-azobisisobutyronitr- ile (358 mg) were added. The mixture was
stirred under reflux for 18 h. The succinimide was removed by
filtration and the filtrate concentrated on a rotary evaporator.
The residue was chromatographed on silicic acid (300 g), using the
flash technique and eluting with hexanes to afford the product as a
pale yellow oil (25.4 g, 77% yield) which eventually crystallized.
.sup.1H NMR (CDCl.sub.3) .delta.8.03 (s, 1H), 7.73 (d,1H, J=8 Hz),
7.61 (d, 1H, J=8 Hz), 7.25 (s, 1H), 4.61 (s, 2H), 1.599 (s,9H).
Intermediate 23
(R=methyl)
Methyl
4-Bromo-3-[[4-[1-cyclohexyl-5-(triphenylmethyl)-2H-tetrazol-5-yl]1--
H-benzimidazol-2-yl]phenoxy]methyl]benzoate
[0129] A mixture of compound 22 (400 mg, 0.68 mmol) and potassium
carbonate (282 mg, 2.04 mmol) in DMF (10 mL) was stirred at rt for
10 min. Methyl 4-bromo-3-(bromomethyl)benzoate (230 mg, 0.75 mmol)
was added and stirring was continued for 18 h at 80.degree. C. The
mixture was poured into ethyl acetate-water. The aqueous layer was
re-extracted with ethyl acetate. The organic layers were combined
and washed (water, brine) and dried over sodium sulfate. The
solution was concentrated on a rotary evaporator. The residue was
purified by flash chromatography on silicic acid and eluting with
hexanes-ethyl acetate to afford 23 (R=methyl) as a colorless oil
(400 mg, 71% yield) which eventually crystallized. ESI-MS m/e 83
(M+1) .sup.1H NMR (CDCl.sub.3) .delta.8.55 (s, 1H), 8.25 (s, 1H),
8.05 (d, 1H, J=8 Hz), 7.86 (d, 1H, J=8 Hz), 7.68 (d, 2H, J=8.5 Hz),
7.61 (d, 2H, J=8.5 Hz), 7.33 (m, 10H), 7.18 (m, 6H), 7.15 (s, 1H),
5.22 (s, 2H), 4.38 (m, 1H), 3.91 (s, 3H), 2.30 (m, 2H), 1.95 (m,
4H), 1.75 (m, 2H), 1.34 (m, 3H).
Intermediate 23
(R=tert-butyl)
4-Bromo-3-[[4-[1-cyclohexyl-5-(triphenylmethyl)-2H-tetrazol-5-yl]-1H-benzi-
midazol-2-yl]phenoxy]methyl]-benzoic acid 1,1-dimethylethyl ester
(23)
[0130] A mixture of compound 22 (2.04 g, 3.88 mmol) and cesium
carbonate (1.38 g, 4.23 mmol) in DMF (20 mL) was stirred at rt for
10 min. tert-Butyl 4-bromo-3-(bromomethyl)benzoate (1.24 g, 3.55
mmol) was added and stirring continued for 18 h at rt. The mixture
was poured into ethyl acetate-cold water. The organic layer was
washed (water, brine), dried (MgSO.sub.4), and concentrated on a
rotary evaporator. The residue was crystallized from ethyl
acetate-hexanes to afford 23 (R=tert-butyl) (2.3 g, 78% yield).
ESI-MS m/e 872 (M+1).
Intermediate 25
3-[[4-[1-Cyclohexyl-5-[2-(triphenylmethyl)-2H-tetrazol-5-yl]-1H-benzimidaz-
ol-2-yl]phenoxymethyl]-4-(5-pyrimidinyl)-benzoic acid
1,1-dimethylethyl ester (25)
[0131] A solution of compound 23 (215 mg, 0.25 mmol) and
pyrimidine-5-boronic acid (24, 37 mg, 0.29 mmol) in dimethoxyethane
(3.5 mL) was degassed at rt with a stream of nitrogen.
Tetrakis(triphenylphosp- hine)palladium(0) (29 mg, 0.025 mmol) was
added, followed by a degassed solution of potassium phosphate(315
mg, 1.48 mmol) in water (0.22 mL). The contents of the sealed vial
were shaken at 90.degree. C. for 18 h on an Innova platform shaker
mounted with a Thermolyne Dri-Bath heater. The vial was cooled and
the contents poured into a mixture of methylene chloride and water.
The organic layer was washed 2.times. with water followed by brine.
The organic layer was dried over sodium sulfate and crude 25
isolated as a froth after concentration on a rotating
evaporator.
EXAMPLE 4
3-[[4-[1-Cyclohexyl-5-(1H-tetrazol-5-yl)-1H-benzimidazol-2-yl]phenoxy]meth-
yl]-4-(5-pyrimidinyl)benzoic acid (26, R=H)
[0132] A solution of 4N hydrochloric acid in dioxane (1.5 mL) was
added to a vial containing crude 25 (40 mg). The mixture was
sonicated for 2 h. Dilution with diethyl ether precipitated crude
26 (30 mg) which was collected, washed with ether and dried. A
solution of 26 in DMF was purified by preparative HPLC to afford 26
as an acid addition salt with trifluoroacetic acid (1.9 mg, 6.6%
yield). ESI-MS m/e573 (M+1). Purity 98%(uv) by HPLC
Intermediate 29
Resin bound 4-Hydroxy-2-methoxybenzaldehyde (29)
[0133] A solution of 4-hydroxy-2-methoxybenzaldehyde (45.6 g, 0.30
mol) in DMF (70 mL) was added dropwise to a cooled ice/water bath
and stirred mixture of 95% sodium hydride (7.58 g, 0.03 mol) in DMF
(200 mL). After the vigorous evolution of hydrogen had subsided, a
catalytic amount (60 mg) of tetrabutylammonium iodide was added
followed by Merrifield resin (60 g of 100-200 mesh containing 1.25
mmol/g). The cooling bath was removed and stirring continued at an
oil bath temperature of 65.degree. C. for 15 min when the reaction
mixture set solid. It was diluted with additional DMF (250 mL) and
stirring continued for an additional 20 h. The mixture was cooled
and the solid collected and washed sequentially with DMF
(2.times.100 mL), water(3.times.150 mL), DMF:water (1:1,
2.times.100 mL), DMF (2.times.100 mL), methanol (3.times.100 mL),
methylene chloride (2.times.100 mL), and methanol (3.times.100 mL).
The bound aldehyde was dried over phosphorous pentoxide for 24 hr
at 0.1 mm to afford 68.5 g of 29 (100% yield) with a loading of
about 1.24 mmol/g.
Intermediate 30
Resin bound 4-Hydroxy-2-methoxybenzyl alcohol (30)
[0134] Sodium borohydride (10.0 g, 0.26 mol) was added to a mixture
of compound 29 (35 g, 0.044 mol) in 250 mL of a 1:1 mixture of THF
and 95% ethyl alcohol. The mixture was shaken for 18 h at rt. The
resin was washed sequentially with DMF (100 mL), water (100 mL),
DMF (100 mL), methanol (100 mL), and methylene chloride (100 mL).
This latter washing sequence was repeated (3.times.). The resin was
dried in vacuo to afford 30 as a colorless solid (33.9 g), with an
approximate loading of 1.25 mmol/g. An IR spectrum of 30 was
negative for an aldehyde carbonyl peak.
Intermediate 31
Resin bound
2-[4-(tert-Butyldimethylsilyoxy)phenyl]-1-cyclohexyl-5-(2H-tet-
razol-5-yl)-1H-benzimidazole (31)
[0135] Compound 21 (4.13 g, 8.7 mmol), diisopropylazodicarboxylate
(2.07 mL, 10.5 mmol) and triphenylphosphine (2.75 g, 10.5 mmol)
were added to a suspension of resin 30 (2.8 g, 3.5 mmol) in a 1:1
mixture of THF and methylene chloride (50 mL) at 0.degree. C. The
mixture was allowed to reach rt, and was shaken for 18 h and
filtered. The resin was washed successively with DMF (2.times.50
mL), 1:1 DMF-water (50 mL), DMF (2.times.50 mL), methanol
(3.times.50 mL), and methylene chloride (3.times.50 mL). The resin
was dried in vacuo to afford 2.65 g of 31.
Intermediate 32
Resin bound
2-[4-hydroxyphenyl]-1-cyclohexyl-5-(2H-tetrazol-5-yl)-1H-benzi-
midazole (32)
[0136] Compound 31 (2.65 g, 3.3 mmol) was added to
tetrabutylammonium fluoride (17.5 mL of a 1.0 N solution in THF,
17.5 mmol) at rt. The suspension was shaken for 66 h and was
filtered. The resin was washed and dried as described in the
preceding experiment to afford resin bound
2-[4-hydroxyphenyl]-1-cyclohexyl-5-(2H-tetrazol-5-yl)-1H-benzimidazole
(2.5 g).
EXAMPLE 5
1-Cyclohexyl-2-[4-[5-(2-methoxyphenyl)-1,2,4-oxadiazol-3-ylmethoxy]phenyl]-
-5-[1H-tetrazol-5-yl]-1H-benzimidazole (34)
[0137] Compound 32 (50 mg, 0.06 mmol)
3-chloromethyl-5-(2-methoxyphenyl)-1- ,2,4-oxadiazole (67 mg, 0.30
mmol), and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU, 46 mg, 0.3
mmol) were added to DMF (1.0 mL) in a fritted vial. The vial was
shaken at rt for 18 h in a Bodan MiniBlock.TM. II reactor. The
resin was filtered and washed successively with DMF (1 mL), water
(1 mL), DMF (2.times.1 mL), methyl alcohol (3.times.1 mL), and
methylene chloride (3.times.1 mL). The vial was recharged with 1 mL
of TFA:methylene chloride (95:5) and was shaken for 18 h at rt. The
contents of the vial were filtered and the filtrate concentrated to
dryness using a Speed Vac.RTM. Plus. The residue was dissolved in
DMF and the solution purified by preparative HPLC to afford 34 as a
salt with TFA (0.7 mg, 1.8% yield).
Intermediate 36
Resin bound
2-[4-(tert-Butyldimethylsilyoxy)phenyl]-1-cyclohexyl-5-(1H-tet-
razol-5-yl)-1H-benzimidazole (36)
[0138] The chlorotrityl resin (2.5 g of 200-400 mesh containing
1.14 mmol/g) was added to dry methylene chloride(25 mL) in a resin
flask and under argon. Diisopropylethylamine (1.47 g, 0.0115 mol)
and compound 21 (5.41 g, 0.0114 mol) were added. The mixture was
shaken for 4 h. The resin was collected and successively washed
with methylene chloride (3.times.50 mL), methanol (2.times.50 mL),
and methylene chloride (2.times.50 mL). The resin was collected and
dried to afford 36 (4.8 g).
Intermediate 37
Resin bound
1-Cyclohexyl-2-[4-[5-(2-methoxyphenyl)-1,2,4-oxadiazol-3-ylmet-
hoxy]phenyl]-5-[2H-tetrazol-5yl]-1H-benzimidazole (37)
[0139] The protected tetrazole 36 (4.8 g) was suspended in THF (70
mL). A solution of TBAF (28 mL of 1.0 N, ca 5 eq) was added and the
mixture shaken for 5 h at rt. The resin was collected and washed
successively with THF (70 mL), methylene chloride (2.times.70 mL),
methanol (2.times.70 mL), and methylene chloride (2.times.70 mL).
The resin was collected and dried to afford the desilylated
intermediate (3.8 g).
[0140] The desilylated resin (100 mg, 0.12 mmol),
3-chloromethyl-5-(2-meth- oxyphenyl)-1,2,4-oxadiazole (81 mg, 0.36
mmol), potassium carbonate (50 mg, 0.27 mmol), and sodium iodide
(18 mg, 0.12 mmol) were added to a vial containing DMF (1.0 mL).
The mixture was shaken at 75.degree. C. for 18 h. The resin was
collected and was washed successively with DMF (2.times.2 mL),
DMF-70% water (2.times.2 mL), DMF (2 mL), methanol (2 mL), and
methylene chloride (2.times.2 mL). The resin was dried to afford 37
which was directly cleaved to afford 34.
1-Cycohexyl-2-[4-[5-(2-methoxyphenyl)-1,2,4-oxadiazol-3-ylmethoxy]phenyl]--
5-[H-tetrazol-5yl]-1H-benzimidazole (34) (alternate route to
example 5)
[0141] Compound 37 from the preceding experiment was added to 2.0
mL of methylene chloride:TFA:triisopropylsilane (6:2:0.5). The
mixture was shaken at rt for 20 min and was filtered. The resin was
washed with methylene chloride (2.times.2 mL). The washings and
filtrate were combined and concentrated on a rotary evaporator to
afford 34 as a TFA salt (56 mg, 70% yield). ESI MS m/e 549 (M+1).
The product was identical to that described in Example 5.
Intermediate 39
3-[[4-[1-Cyclohexyl-5-[2-(triphenylmethyl)-2H-tetrazol-5-yl]-1H-benzimidaz-
ole-2-yl]phenoxymethyl]-4-(5-acetyl-2-thienyl)benzoic acid methyl
ester (39)
[0142] A mixture of compound 23 (500 mg, 0.60 mmol) and
2-acetyl-5-thiopheneboronic acid (112 mg, 0.66 mmol) in
dimethoxyethane (4 mL) was degassed for 15 min with a gentle stream
of nitrogen. Tetrakis(triphenylphosphine)palladium(0) (69 mg, 0.06
mmol) and a degassed solution of potassium phosphate (1 g, 4.7
mmol) in water (0.7 mL) were added. The mixture was stirred for 18
h at 80.degree. C. and concentrated. The residue was extracted with
ethyl acetate and filtered. The filtrate was reconcentrated and the
residue chromatographed on silicic acid (14 g) using the flash
technique and eluting with mixtures of ethyl acetate and hexanes
(1:4) to afford 39 (173 mg, 33% yield). ESI MS m/e 876 (M+1)
Intermediate 40
3-[[4-[1-Cyclohexyl-5-[2-(triphenylmethyl)-2H-tetrazol-5-yl]-1H-benzimidaz-
ole-2-yl]phenoxymethyl]-4-(5-acetyl-2-thienyl)benzoic acid (40)
[0143] A mixture of compound 39 (169 mg, 0.19 mmol) and lithium
hydroxide (23 mg, 0.97 mmol) in water (1 mL), ethanol (2 mL), and
THF (2 mL) was stirred under reflux for 3 h and cooled and
acidified with dilute hydrochloric acid to precipitate 40 which was
dried in vacuo to afford 147 mg (90% yield). ESI MS m/e 862
(M+1)
EXAMPLE 6
tert-Butyl
N-[4-(5-acetylthien-2-yl)-3-({4-[1-cyclohexyl-5-(1H-tetrazol-5--
yl]-1H-benzimidazol-2-yl]phenoxy}methyl)benzoyl]glycinate (42)
[0144] Compound 40 (23.2 mg, 0.027 mmol), tert-butyl glycinate (4.4
mg, 0.34 mmol), N,N-diisopropylethylamine (14 mg, 0.11 mmol), and
2-[1H-benzotriazol-1-yl]-1,1,3,3,-tetramethyluronium
tetrafluoroborate (TBTU, 11 mg, 0.034 mmol) were added to DMF (0.6
mL) in a fritted vial. The vial was sealed and shaken at rt for 2 h
in a Bodan MiniBl II reactor Block.TM.. The mixture was filtered
and the residual solid was washed with methanol (3.times.0.3 mL).
The combined filtrate and washings were concentrated to dryness
using a Speed Vac.RTM. Plus. The residue was dissolved in methanol
and the solution purified by preparative HPLC to afford 42 as a TFA
salt (6.0 mg, 26% yield). ESI MS m/e 732 (M+1).
EXAMPLE 7
N-[4-(5-Acetylthien-2-yl)-3-({4-[1-cyclohexyl-5-(1H-tetrazol-5-yl]-1H-benz-
imidazol-2-yl]phenoxy}methyl)benzoyl]glycine (43)
[0145] TFA:methylene chloride (0.6 mL of 1:1) was added to compound
42 (3.0 mg, 0.0035 mmol) in a vial contained in a Bodan MiniBl II
reactor Block.TM.The block was shaken for 30 min and the contents
of the vial filtered. The vial was washed with methylene chloride
(2.times.0.4 mL) and the combined filtrate and washings
concentrated to dryness using a Speed Vac.RTM. Plus. The residue
was dissolved in methanol and the solution purified by preparative
HPLC to afford 43 as a TFA salt (1.9 mg, 70% yield). ESI MS m/e 676
(M+1).
Intermediate 45
Methyl 4-(cyclohexylamino)-3-nitrobenzoate (45)
[0146] A solution of methyl 4-chloro-3-nitrobenzoate (5.00 g, 23.2
mmol), cylohexylamine (3.98 mL, 34.8 mmol), and triethylamine (4.8
mL, 35 mmol) in acetonitrile (75 mL) was reluxed for 22 h under an
argon atmosphere. The reaction mixture was concentrated to a volume
of .about.20 mL and the precipitated solid was filtered. The
filtrate was diluted with ethyl acetate and was washed with 1M
citric acid (1.times.) followed by saturated NaHCO.sub.3
(1.times.). The organic layer was dried (MgSO.sub.4) and
concentrated on a rotary evaporator. The residue was purified by
column chromatography (silica gel, 50-70% dichloromethane/hexanes
(v/v)) to give compound 45 as a yellow solid (6.32 g, 98%). ESI-MS
m/e 279.2 (M+1).
Intermediate 46
Methyl 3-amino-4-(cyclohexylamino)benzoate (46)
[0147] A solution of compound 45 (2.03 g, 7.29 mmol) in a mixture
of methanol (25 mL) and ethyl acetate (25 mL) was hydrogenated over
10% palladium on carbon (0.20 g) at 25 psi for 15 h. The reaction
mixture was filtered and concentrated to afford compound 46 as a
crude grey-brown oil (1.99 g, 100%). ESI-MS m/e 249.3 (M+1).
Intermediate 47
Methyl 3-{[4-(benzyloxy)benzoyl]amino}-4-(cyclohexylamino)benzoate
(47)
[0148] A solution of 4-benzyloxybenzoic acid (2.50 g, 11.0 mmol)
and thionyl chloride (1.2 mL, 16 mmol) in benzene (35 mL) was
refluxed for 3 h under an argon atmosphere. The reaction mixture
was cooled to rt and concentrated on a rotary evaporator. The
residue was coevaporated with benzene (2.times.) to afford the acid
chloride as a white solid. A solution of the acid chloride (0.333
g, 1.35 mmol) and compound 46 (0.275 g, 1.11 mmol) in
dichloromethane (3 mL) was treated with triethylamine (0.22 mL) at
rt. After 30 min, the reaction mixture was diluted with ethyl
acetate and washed with 1 M KHSO.sub.4 (1.times.), saturated
NaHCO.sub.3 (1.times.), and brine (1.times.). The organic layer was
dried (MgSO.sub.4) and concentrated on a rotary evaporator to
afford compound 47 as a burgundy foam (0.558 g, 100%). ESI-MS m/e
459.3 (M+1).
Intermediate 48
Methyl
2-[4-(benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazole-5-carboxylate
(48)
[0149] A solution of compound 47 (0.558 g, 1.11 mmol) in acetic
acid (3 mL) was refluxed for 1 h and then stirred at 100.degree. C.
for 15 h. The reaction mixture was cooled to rt and concentrated on
a rotary evaporator. The residue was coevaporated with toluene
(1.times.) and then purified by column chromatography (silica gel,
8% acetone/toluene (v/v)) to give compound 48 as a white solid
(0.234 g, 53%). ESI-MS m/e 441.3 (M+1).
Intermediate 49
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazole-5-carboxylic
acid (49)
[0150] A solution of compound 48 (0.514 g, 1.17 mmol) in
tetrahydrofuran (5 mL) and ethanol (5 mL) was treated with 4 M NaOH
(5 mL). The reaction mixture was refluxed for 1 h, cooled to rt,
and concentrated on a rotary evaporator. The residue was dissolved
in a small amount of water and made acidic with concentrated
hydrochloric acid. The solid precipitate was filtered, washed with
water, and dried to afford 49 as a white solid (0.52 g, 100%).
ESI-MS m/e 427.3 (M+1).
EXAMPLE 8
({2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazol-5-yl}carbonyl)benze-
nesulfonamide (55)
[0151] A suspension of compound 49 (0.060 g, 0.14 mmol) and thionyl
chloride (0.051 mL) in benzene (2 mL) was refluxed for 2 h under an
argon atmosphere. The reaction mixture was cooled to rt and
concentrated on a rotary evaporator. The residue was coevaporated
with benzene (2.times.) to afford the acid chloride as a white
solid (0.062 g, 100%). Sodium hydride (0.0101 g, 0.252 mmol, 60%
dispersion in oil was added to a solution of phenylsulfonamide
(0.0425 g, 0.270 mmol) in dimethylformamide (1 mL) at rt under an
argon atmosphere. After 1 h, dichloromethane was added (1 mL),
followed by the acid chloride of 49 at 0.degree. C. The reaction
was warmed to rt. After 1 h, the reaction was diluted with
dichloromethane and quenched with 1 N hydrochloric acid. The
aqueous layer was washed with dichloromethane (3.times.). The
combined organic layers were dried (MgSO.sub.4) and concentrated on
a rotary evaporator. The residue was purified by column
chromatography (silica gel, 25% acetone/toluene (v/v)). This
material was further purified by reverse phase HPLC (C18 column,
water/acetonitrile gradient containing 0.05% trifluoroacetic acid)
to afford 55 as a white solid (0.0219 g, 54%). .sup.1H NMR
(CDCl.sub.3) .delta.8.71 (d, 1H, J=1.1 Hz), 8.05 (m, 3H), 7.82 (d,
1H, J=8.8 Hz), 7.69 (d, 2H, J=9.1 Hz), 7.58 (m, 1H), 7.51-7.36 (m,
7H), 7.21 (d, 2H, 8.8 Hz), 5.16 (s, 2H), 4.52 (m, 1H), 2.32 (br m,
2H), 2.01 (br m, 4H), 1.80 (br m, 1H), 1.35 (br m, 3H); ESI-MS m/e
566.4 (M+1); HPLC purity (ELSD)>95%.
EXAMPLE 9
N-({2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazol-5-yl}carbonyl)met-
hanesulfonamide
[0152] Methanesulfonamide (0.0188 g, 0.198 mmol), sodium hydride,
and the acid chloride of 49 (0.0083 g, 0.019 mmol) were reacted in
dimethylformamide and dichloromethane according to the procedure
described for example 8. The crude material was purified by reverse
phase HPLC (C18 column, water/acetonitrile gradient containing
0.05% trifluoroacetic acid) to afford the product as a white solid
(0.0055 g, 52%). .sup.1H NMR (CDCl.sub.3) .delta.8.68 (s, 1H), 8.17
(d, 1H, J=8.8 Hz), 7.91 (d, 1H, J=8.8 Hz), 7.69 (d, 2H, J=8.5 Hz),
7.48-7.37 (m, 5H), 7.24 (d, 2H, J=8.7 Hz), 5.18 (s, 2H), 4.57 (m,
1H), 3.34 (s, 3H), 2.34 (br m, 2H), 2.06 (br m, 4H), 1.85 (br m,
1H), 1.39 (br m, 3H); ESI-MS m/e 504.3 (M+1); HPLC purity
(ELSD)>95%.
EXAMPLE 10
3-{2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazol-5-yl}-1,2,4-oxadia-
zol-5(4H)-one
(Two Steps)
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-N'-hydroxy-1H-benzimidazole-5-carboxi-
midamide
[0153] (Step 1) Triethylamine (0.040 mL, 2.9 mmol) was added to
hydroxylamine hydrochloride (0.212 g, 3.05 mmol) in dimethyl
sulfoxide (1.5 mL). The insoluble material was removed by
filtration and washed with tetrahydrofuran (5 mL). The filtrate was
concentrated on a rotary evaporator. To the resulting solution of
hydroxylamine in dimethylsulfoxide was added nitrile 12 (0.250 g,
0.61 mmol). The solution was heated to 75.degree. C. for 16 h. The
resulting suspension was diluted with water, extracted with ethyl
acetate, and washed with 1N hydrochloric acid. The aqueous layer
was made basic with 1N sodium hydroxide and extracted with ethyl
acetate (2.times.). This ethyl acetate solution was washed with
water (1.times.) and brine (1.times.), dried (MgSO.sub.4) and
concentrated on a rotary evaporator to afford the product as a
white solid (0.197 g, 96%). ESI-MS m/e 441.3 (M+1).
3-{2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazol-5-yl}-1,2,4-oxadia-
zol-5(4H)-one
[0154] (Step 2) Methylchloroformate (0.020 mL, 0.26 mmol) was added
dropwise to a suspension of the preceding product (0.088 g, 0.2
mmol) and pyridine (0.020 mL, 0.25 mmol) in dimethylformamide (1
mL) at 0.degree. C. The resulting clear solution was stirred
overnight at rt. Water was added to the mixture and a precipitate
was collected by filtration. The solid was refluxed in xylene for 3
hours, during which time the reaction became a clear solution, then
formed a new precipitate. This material was filtered and washed
with hexane, to afford the product as a white solid (0.038 g, 41%),
.sup.1H NMR (CD.sub.3OD) .delta.8.06 (s, 1H), 7.91 (d, 1H, J=8.8
Hz), 7.78 (d, 1H, J=8.4 Hz), 7.59 (d, 2H, J=8.8 Hz), 7.49 (d, 2H,
J=7.7 Hz) 7.44-7.32 (m, 3H), 7.22 (d, 2H, J=8.8 Hz), 5.21 (s, 2H),
4.41 (m, 1H), 2.45-2.25 (m, 2H), 1.98 (m, 4H), 1.78 (m, 1H), 1.39
(m, 3H); ESI-MS m/e 467.3 (M+1); HPLC purity (ELSD) 91%.
EXAMPLE 11
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-5-(2-oxido-3H-1,2,3,5-oxathiadiazol-4-
-yl)-1H-benzimidazole
[0155] Thionyl chloride (0.010 mL, 0.14 mmol) was added dropwise to
a suspension of
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-N'-hydroxy-1H-benzimid-
azole-5-carboximidamide (0.025 g, 0.057 mmol, see above for
preparation) and pyridine (0.020 mL, 0.25 mmol) in tetrahydrofuran
(2 mL) at 0.degree. C. The resulting solution was stirred for 3 h,
and then concentrated on a rotary evaporator. As starting material
remained, the residue was resubjected to the reaction conditions
(0.020 mL thionyl chloride and 0.040 mL pyridine). The residue was
purified by reverse phase HPLC (C18 column, water/acetonitrile
gradient containing 0.05% trifluoroacetic acid) to afford the
product as a yellow solid (0.011 g, 32%). .sup.1H NMR (CD.sub.3OD)
.delta.8.32 (d, 1H, J=8.8Hz), 8.25 (s, 1H), 8.04 (d, 1H, J=8.8 Hz),
7.77 (d, 2H, J=9.1 Hz), 7.49 (d, 2H, J=6.6 Hz) 7.43-7.35 (m, 5H),
5.27 (s, 2H), 4.55 (m, 1H), 2.45 (m, 2H), 2.15 (m, 2H), 2.00 (m,
2H), 1.80 (m, 1H), 1.44 (m, 3H); ESI-MS m/e 487.3 (M+1); HPLC
purity (ELSD)>95%.
Intermediate 56
6-Chloro-5-nitronicotinic acid (56)
[0156] A suspension of 6-hydroxy-5-nitronicotinic acid (8 g, 43.5
mmol) (prepared by literature nitration of 6-hydroxynicotinic acid)
in phosphorus oxychloride (24.3 mL, 261.0 mmol) was heated to
reflux (105.degree. C.) for 3 h under a nitrogen atmosphere. At the
end of this time period all of the solids had dissolved. The
reaction was cooled to rt and poured over 500 mL crushed ice, and
stirring was continued for 30 minutes until all of the ice had
melted. The aqueous mixture was extracted with a 1:2 mixture of THF
in diethyl ether (3.times.150 mL). The pooled organic extracts were
washed with brine (1.times.), dried (MgSO.sub.4), and concentrated
on a rotary evaporator to afford intermediate 56 as a light brown
solid (8.37 g, 95%). ESI-MS m/e 405.3 (2M-1).
Intermediate 57
6-(Cyclohexylamino)-5-nitronicotinic acid (57)
[0157] To a solution of intermediate 56 (10.1 g, 49.9 mmol) and
triethylamine (14.6 mL, 125.0 mmol) in acetonitrile (130 mL) and
DMSO (25 mL) was added cyclohexylamine (13.7 mL, 100 mmol). The
reaction mixture was heated to reflux (90.degree. C.) under a
nitrogen atmosphere for 20 h. The reaction mixture was cooled to rt
and concentrated on a rotary evaporator and the residue was
dissolved in 800 mL 1N sodium hydroxide solution. This solution was
washed with dichloromethane (3.times.75 mL) and then made acidic
(pH=5) with concentrated hydrochloric acid. The precipitated
product was collected by vacuum filtration and washed sparingly
with cold water to afford the HCl salt of intermediate 57 as a
yellow solid (9.8 g, 65%). ESI-MS m/e 266.2 (M+1).
Intermediate 58
Methyl 6-(cyclohexylamino)-5-nitronicotinate (58)
[0158] To a solution of intermediate 57 (9.7 g, 32.2 mmol) in
methanol (400 mL) was added chlorotrimethylsilane (3 mL) to achieve
pH=2. The reaction mixture was heated at strong reflux for 72 h,
during which time the pH was monitored and more
chlorotrimethylsilane added as necessary to maintain an acidic
solution. The reaction mixture was allowed to cool and was then
concentrated on a rotary evaporator. The residue was purified by
column chromatography (silica gel, 7:2:1
hexanes/dichloromethane/methanol- ) to afford the crude ester as a
yellow solid (9.0 g, 89%). ESI-MS m/e 280.1 (M+1).
[0159] A solution of the ester (9.0 g, 32.2 mmol) and 10% palladium
on carbon (450 mg) in 1.3:1 ethyl acetate/methanol (220 mL) was
placed on a PARR hydrogenation shaker under a hydrogen atmosphere
at 55 p.s.i. for 2.5 h. After hydrogen uptake had ceased, the
solution was filtered to remove the Pd/C and then concentrated on a
rotary evaporator. The oily residue was dissolved in acetonitrile
(10 mL) and a small amount of water (1-2 mL) was added to cause a
slight cloudiness. This solution was then frozen solid in a dry
ice/acetone bath and placed on the lyophilizer overnight to afford
intermediate 58 as a yellow solid (8.04 g, 100%). ESI-MS m/e 250.1
(M+1).
Intermediate 59
Methyl
3-cyclohexyl-2-(4-hydroxyphenyl)-3H-imidazo[4,5-b]pyridine-6-carbox-
ylate (59)
[0160] A solution of intermediate 58 (5.0 g, 20.0 mmol) and ethyl
4-hydroxybenzimidate hydrochloride (6.0 g, 30.0 mmol) in methanol
(67 mL) was heated at reflux for 22 h. The reaction mixture was
then cooled to rt and the precipitated solid was collected by
vacuum filtration and washed sparingly with cold methanol to afford
intermediate 59 as a white solid (6.13 g, 88%). ESI-MS m/e 352.2
(M+1).
Intermediate 60
3-Cyclohexyl-2-(4-hydroxy-phenyl)-3H-imidazo[4,5-b]pyridine-6-carboxylic
acid (2-cyano-ethyl)-amide (60)
[0161] A solution of Intermediate 59 (1.5 g, 4.3 mmol) in
tetrahydrofuran (5 mL) and ethanol (5 mL) was treated with 4 M NaOH
(5 mL). The reaction mixture was refluxed for 1 h, cooled to rt,
and concentrated on a rotary evaporator. The residue was dissolved
in a small amount of water and made acidic with concentrated
hydrochloric acid. The solid precipitate was filtered, washed, and
dried to afford the acid as a white solid (1.4 g, 97%). ESI-MS m/e
338.1 (M+1).
[0162] The crude acid (1.4 g, 4.15 mmol), 1-hydroxybenzotriazole
(1.27 g, 8.3 mmol), sodium bicarbonate (628 mg, 7.47 mmol), and
2-cyanoethylamine (332 .mu.L, 4.98 mmol) were dissolved in a 5:1
mixture of dimethylformamide/dichloromethane (12 mL). To this
solution was added ethyl-3-(3dimethylamino-propyl)carbodiimide (955
mg, 4.98 mmol) at rt, and the resulting reaction mixture was
stirred for 18 h at rt under a nitrogen atmosphere. The reaction
mixture was concentrated on a rotary evaporator to an approximate
volume of 4 mL and then ethyl acetate (75 mL) was added and the
solids removed by vacuum filtration. The filtrate was washed with
0.1 N HCl (1.times.), saturated NaHCO3 (1.times.), and brine
(1.times.). The organic phase was dried (Na.sub.2SO.sub.4) and
concentrated to afford the HCl salt of intermediate 60 as a white
solid (1.4 g, 80%). ESI-MS m/e 390.1 (M+1).
Intermediate 61
2-[4-(tert-Butyl-dimethyl-silanyloxy)-phenyl]-3-cyclohexyl-3H-imidazo[4,5--
b]pyridine-6-carboxylic acid (2-cyano-ethyl)-amide (61)
[0163] To a solution of intermediate 60 (1.5 g, 3.85 mmol) in
dimethylformamide (5 mL) was added imidazole (0.917 g, 13.5 mmol)
and tert-butyldimethylsilyl chloride (0.870 g, 5.77 mmol). The
reaction mixture was stirred at 35.degree. C. for 20 h, and then
ethyl acetate (100 mL) was added. The resulting organic phase was
washed with water (4.times.20 mL) and brine (1.times.) and then
dried (Na.sub.2SO.sub.4). After concentrating on a rotary
evaporator the resulting residue was purified by column
chromatography (silica gel, 1:1 hexanes/ethyl acetate-3:7
hexanes/ethyl acetate) to afford intermediate 61 as a white solid
(1.6 g, 83%). ESI-MS m/e 504.0 (M+1).
Intermediate 62
3-(5-{2-[4-(tert-Butyl-dimethyl-silanyloxy)-phenyl]-3-cyclohexyl-3H-imidaz-
o[4,5-b]pyridin-6-yl}-tetrazol-1-yl)-propionitrile (62)
[0164] A solution of intermediate 61 (250 mg, 0.5 mmol), sodium
azide (33 mg, 0.5 mmol), and 2,6-lutidine (116 .mu.L, 1.0 mmol) in
dichloromethane (3 mL) was cooled to 0.degree. C. and
trifluoromethanesulfonic anhydride (84 .mu.L, 0.5 mmol) from a
freshly opened ampule was added under a nitrogen atmosphere. The
reaction mixture was allowed to warm up slowly to rt over 2 h and
stirring continued for 20 h at which point dichloromethane was
added (25 mL). The resulting organic phase was washed with
saturated NaHCO.sub.3 (1.times.20 mL), dried (Na.sub.2SO.sub.4),
and concentrated on a rotary evaporator. The foamy residue was
purified by reverse phase HPLC (C18 column, water/acetonitrile
gradient containing 0.05% trifluoroacetic acid) to afford
intermediate 62 as a white solid (48 mg, 18%). ESI-MS m/e 529.1
(M+1).
Intermediate 63
3-{5-[3-Cyclohexyl-2-(4-hydroxy-phenyl)-3H-imidazo[4,5-b]pyridin-6-yl]-tet-
razol-1-yl}-propionitrile (63)
[0165] A solution of intermediate 62 (78 mg, 0.15 mmol) in glacial
acetic acid (3 mL), tetrahydrofuran (1 mL), and water (1 mL) was
heated to 100.degree. C. for 3 h under a nitrogen atmosphere. Ethyl
acetate (40 mL) was added and the organic phase washed with water
(2.times.20 mL), NaHCO.sub.3 (2.times.20 mL), and brine (1.times.).
The organic phase was dried (Na.sub.2SO.sub.4) and concentrated on
a rotary evaporator to afford intermediate 63 as a white solid (54
mg, 89%). ESI-MS m/e 415.1 (M+1).
Intermediate 63a
2-(Bromomethyl)-4'-cyano-N-methyl-1,1'-biphenyl-4-carboxamide
(3 Steps)
[0166] (Step 1) A solution of tert butyl 4-bromo-3-methylbenzoate
(3.3 g, 12.2 mmol) was reacted with 4-cyanophenylboronic acid (2.15
g, 14.6 mmol) according to the procedure described for compound 14.
The crude product was purified by column chromatography (silica
gel, 3:2 hexanes/methylene dichloride) to afford tert-butyl
4'-cyano-2-methyl-1,1'-biphenyl-4-carbox- ylate as a white solid
(3.0 g, 84% yield). ESI-MS m/e 294.1 (M+1).
[0167] (Step 2) To a solution of the preceding ester (3.0 g, 10.2
mmol) in methylene chloride (75 mL) at 0.degree. C. was added TFA
(75 mL) dropwise over 10 min. The ice bath was removed and stirring
continued for 1 h at rt. The mixture was concentrated on a rotary
evaporator and then diluted with toluene and reconcentrated to
afford 4'-cyano-2-methyl-1,1'-biphenyl- -4-carboxylic acid. ESI-MS
m/e 228.1 (M+1).
[0168] (Step 3) To a solution of the preceding acid (760 mg, 3.2
mmol), 2M methylamine in tetrahydrofuran (3.1 mL, 6.2 mmol), and
diisopropylethylamine (1.71 mL, 9.6 mmol) in dimethylformamide (15
mL) was added benzotriazol-1-yloxytris(dimethylamino)phosphonium
hexafluorophosphate (1.72 g, 3.89 mmol) under a nitrogen
atmosphere. The reaction mixture was stirred at rt for 1 h, at
which point the reaction mixture was diluted with dichloromethane
50 mL) and washed with water (2.times.15 mL), saturated NaHCO3
(2.times.15 mL), and brine (1.times.). The organic phase was then
dried (Na.sub.2SO4) and concentrated on a rotary evaporator. The
crude material was purified by column chromatography (silica gel,
1:1 dichloromethane/hexane to 100% dichloromethane) to obtain the
amide intermediate,
2-methyl-4'-cyano-N-methyl-1,1'-biphenyl-4-carboxamide, which was
dissolved in dichloromethane (15 mL). N-bromosuccinimide (1.14 g,
6.4 mmol) was added. The reaction mixture was irradiated with an
ultraviolet light for 4 h under reflux and under a nitrogen
atmosphere. The reaction mixture was concentrated with a rotary
evaporator and the residue purified by reverse phase HPLC (C18
column, water/acetonitrile gradient containing 0.05%
trifluoroacetic acid) to afford the titled compound as a white
solid (210 mg, 20% yield). ESI-MS m/e 329.1 (M+1).
EXAMPLE 12
4'-Cyano-2-{4-[3-cyclohexyl-6-(1H-tetrazol-5-yl)-3H-imidazo[4,5-b]pyridin--
2-yl]-phenoxymethyl}-biphenyl-4-carboxylic acid methylamide
(64)
[0169] Intermediate 63 (50 mg, 0.12 mmol) in dimethylformamide was
reacted with
2-(bromomethyl)-4'-cyano-N-methyl-1,1'-biphenyl-4-carboxamide (63a)
(40 mg, 0.12 mmol), cesium carbonate (20 mg, 0.06 mmol) and the
resulting suspension stirred under a nitrogen atmosphere at rt for
3 h at which point ethyl acetate (30 mL) was added. The organic
phase was washed with water (2.times.25 mL) and brine (1.times.),
dried (Na.sub.2SO.sub.4), and concentrated on a rotary evaporator.
The resulting residue was dissolved in 10% methanolic sodium
hydroxide (5 mL) and water (2 mL) was added. The resulting solution
was stirred at rt under a nitrogen atmosphere for 30 minutes at
which point 10% hydrochloric acid was added until pH=2. This
solution was extracted with ethyl acetate, dried
(Na.sub.2SO.sub.4), and concentrated on a rotary evaporator and the
residue purified by reverse phase HPLC (C18 column,
water/acetonitrile gradient containing 0.05% trifluoroacetic acid)
to afford the product as a white solid (6 mg, 7%). .sup.1H NMR
(CD.sub.3OD) .delta.9.16 (s, 1H), 8.7 (br s, 1H), 8.15 (d, 1H,
J=1.46), 7.94 (dd, 1H, J=8.06, 1.83), 7.80 (d, 2H, J=8.42), 7.71
(br m, 2H), 7.65 (d, 2H, J=8.06), 7.49 (d, 1H, J=8.06), 7.17 (d,
2H, 6.59), 5.14 (s, 1H), 4.46 (br m, 1H), 2.96 (s, 3H), 2.83 (br m,
2H), 1.96 (br m, 4H), 1.75 (br m, 1H), 1.39 (br m, 3H); ESI-MS m/e
610.24 (M+H); HPLC purity (ELSD)>95%.
[0170] Using these methods examples 13 and 14 were prepared.
EXAMPLE 13
4'-Chloro-2-{4-[3-cyclohexyl-6-(1H-tetrazol-5-yl)-3H-imidazo[4,5-b]pyridin-
-2-yl]-phenoxymethyl}-biphenyl-4-carboxylic acid methylamide
[0171] White solid (6 mg, 15%). .sup.1H NMR (CD.sub.3OD)
.delta.9.17 (s, 1H), 8.67 (s, 1H), 8.13 (s, 1H), 7.90 (dd, 1H,
J=8.06, 1.46 Hz), 7.69 (d, 2H, J=8.06), 7.48 (s, 1H), 7.45 (m, 4H),
7.18 (d, 2H, J=8.06), 5.13 (s, 2H), 4.46 (m, 1H), 2.96 (s, 3H),
2.83 (br m, 2H), 1.98 (br m, 4H), 1.75 (br m, 1H), 1.4 (br m, 3H);
ESI-MS m/e 619.2 (M+1); HPLC purity (ELSD)>95%.
EXAMPLE 14
N-[(2-{4-[(4'-Chloro-4-methoxy-1,1'-biphenyl-2-yl)methoxy]phenyl}-3-cycloh-
exyl-3H-imidazo[4,5-b]pyridin-6-yl)carbonyl]methanesulfonamide
[0172] White solid (11 mg, 14%). .sup.1H NMR (CDCl.sub.3)
.delta.8.84 (d, 1H, J=1.8 Hz), 8.48 (d, 1H, J=1.9 Hz), 7.61 (d, 2H,
J=8.8 Hz), 7.42 (m, 4H), 7.34 (d, 1H, 9.2), 7.29 (d, 1H, J=4.4 Hz),
7.04 (m, 3H), 5.03 (s, 2H), 4.36 (br m, 1H), 3.84 (s, 3H), 3.29 (s,
3H), 2.69 (br m, 2H), 1.94 (br m, 4H), 1.70 (br m, 1H), 1.32 (br m,
3H); ESI-MS m/e 645.1 (M+1); HPLC purity (ELSD)>95%.
Intermediate 66
4-(cyclohexylamino)-3-nitrobenzenesulfonamide (66)
[0173] A solution of 4-chloro-3-nitro-benzene sulfonamide (5.49 g,
23.2 mmol), cyclohexylamine (4.00 mL, 35.0 mmol), and triethylamine
(5.0 mL, 35 mmol) in acetonitrile (75 mL) was refluxed overnight
under nitrogen atmosphere. The reaction mixture was concentrated
almost to dryness. The residue was diluted in ethyl acetate and
washed with 1N hydrochloric acid (1.times.) and saturated
NaHCO.sub.3 (1.times.). The organic layer was dried (MgSO.sub.4)
and concentrated on a rotary evaporator. The residue was filtered
and washed with hexane to give the product as a yellow solid (6.34
g, 91%). ESI-MS m/e 300.1 (M+1).
Intermediate 67
3-amino-4-(cyclohexylamino)benzenesulfonamide (67)
[0174] A solution of intermediate 66 (2.56 g, 8.55 mmol) in a
mixture of methanol (50 mL) and ethyl acetate (50 mL) was
hydrogenated over 10% palladium on carbon (0.14 g) at 50 psi for
2.5 h. The reaction mixture was filtered and concentrated to afford
the product as a crude purple-brown foam (2.45 g, 100%). ESI-MS m/e
270.1 (M+1).
Intermediate 68
1-cyclohexyl-2-(4-(hydroxy)phenyl-1H-benzimidazole-5-sulfonamide
(68)
[0175] A solution of ethyl-4 hydroxy benzimidate hydrochloride
(1.01 g, 5.0 mmol) and intermediate 67 (1.5 g, 5.0 mmol) in
methanol (10 mL) was refluxed overnight under a nitrogen
atmosphere. The reaction mixture was cooled to rt and t solid was
filtered and washed with methanol to afford the pinkish brown solid
(1.26 g, 68%). ESI-MS m/e 372.1 (M+1).
Intermediate 69
2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazole-5-sulfonamide
(69)
[0176] A suspension of intermediate 68 (0.556 g, 1.50 mmol) in
N,N-dimethylformamide (4 mL) and cesium carbonate (0.487 g, 1.49
mmol) was heated at 40.degree. C. until almost clear. Benzyl
bromide (0.201 ml, 1.65 mmol) was added drop wise. The reaction
mixture was stirred at rt overnight. The mixture was concentrated
on a rotary evaporator. The residue was washed with water and
filtered to give a white solid precipitate which was filtered, and
then purified by column chromatography (silica gel, 50% to 100%
hexane/ethyl acetate (v/v)) to give the product as a white solid
(0.300 g, 44%). ESI-MS m/e 462.1 (M+1).
EXAMPLE 15
N-acetyl-2-[4-(Benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazol-5-sulfonamid-
e
[0177] A solution of intermediate 69 (0.023 g, 0.05 mmol) in
N,N-dimethylformamide (0.200 mL) and lithium
bis(trimethylsilyl)amide (1.0 M solution in tetrahydrofuran) (0.050
ml, 0.05 mmol) was cooled to 0.degree. C. Acetyl chloride (0.005
ml, 0.06 mmol) was added. The reaction mixture was stirred at rt
for 1.5 h. At this time, more lithium bis(trimethylsilyl)amide
solution (0.050 ml, 0.05 mmol) and acetyl chloride (0.010 ml, 0.014
mmol) were added. The reaction was stirred at rt overnight and
concentrated. The crude material was purified by reverse phase HPLC
(C18 column, water/acetonitrile gradient containing 0.05%
trifluoroacetic acid) to afford the product as a white solid (0.006
g, 18%). .sup.1H NMR (CDCl.sub.3) .delta.8.52 (br s, 1H), 8.15 (br
s, 1H), 7.84 (br m, 1H), 7.61 (br m, 2H), 7.47-7.29 (br m, 5H),
7.22 (br m, 2H), 5.18 (s, 2H), 4.46 (m, 1H), 2.30 (br m, 4H), 2.01
(br m, 2H), 1.82 (br m, 1H), 1.37 (br m, 3H);. ESI-MS m/e 504.1
(M+1); HPLC purity (ELSD) 100%.
EXAMPLE 16
N-Benzoyl-2-[4-(benzyloxy)phenyl]-1-cyclohexyl-1H-benzimidazol-5-sulfonami-
de
[0178] Intermediate 69 (0.023 g, 0.05 mmol) and benzoyl chloride
(0.005 ml, 0.06 mmol) were used to prepare the titled compound
(0.006 g, 18%) by the same method as in example 15. .sup.1H NMR
(CDCl.sub.3) .delta.8.59 (s, 1H), 8.26 (d, 1H, J=8.8 Hz), 7.84 (m,
3H), 7.59 (d, 2H, J=8.8 Hz), 7.56-7.37 (m, 8H), 7.18 (d, 2H, J=8.4
Hz), 5.17 (s, 2H), 4.42 (m, 1H), 2.33 (br m, 4H), 1.99 (br m, 2H),
1.81 (br m, 1H), 1.36 (br m, 3H); ESI-MS m/e 566.1 (M+1); HPLC
purity (ELSD) 100%.
EXAMPLE 17
4-[2-(4-Benzyloxyphenyl)-1-cyclohexyl-1H-benzoimidazole-5-sulfonylaminocar-
bonyl]-benzoic acid methyl ester
[0179] A solution of intermediate 69 (0.046 g, 0.1 mmol) in
dichloromethane (0.500 mL) was cooled to 0.degree. C. and lithium
bis(trimethylsilyl)amide (1.0 M solution in tetrahydrofuran) (0.12
ml, 0.12 mmol) was added and stirred for 5 min. P-phthalic acid
mono methyl ester chloride (0.021 g, 0.11 mmol) was added. The
reaction mixture was stirred at rt for 4 hrs. The crude material
was concentrated and purified by reverse phase HPLC (C18 column,
water/acetonitrile gradient containing 0.05% trifluoroacetic acid)
to afford the titled compound (0.024 g, 38%). .sup.1H NMR
(CDCl.sub.3) .delta.8.45 (s, 1H), 8.17 (d, 1H, J=8.8 Hz), 8.07 (d,
2H, J=8.4 Hz), 7.91(d, 2H, J=8.8 Hz), 7.85 (d, 1H, J=8.8 Hz), 7.56
(d, 2H, J=8.8 Hz), 7.50-7.37 (m, 5H), 7.18 (d, 2H, J=8.8 Hz), 5.18
(s, 2H), 4.41 (m, 1H), 4.00 (bs, 3H), 2.31 (br m, 2H), 2.02 (br m,
4H), 1.80 (br m, 1H), 1.36 (br m, 3H); ESI-MS m/e 624.2(M+1); HPLC
purity (UV) 99%.
EXAMPLE 18
4-[2-(4-Benzyloxyphenyl)-1-cyclohexyl-1H-benzimidazole-5-sulfonylaminocarb-
onyl]-benzoic acid
[0180] A solution of the preceding ester (0.020 g, 0.03 mmol) in
ethanol (1.0 mL), tetrahydrofuran (1.0 mL) and 4N NaOH aq. solution
(0.5 mL), was refluxed for 30 min. The reaction mixture was
concentrated and treated with 4N HCl aq. Solution until acidic. The
solid was filtered and dried to afford the product (0.014 g, 72%).
.sup.1H NMR (CDCl.sub.3) .delta.8.46 (s, 1H), 8.2 (d, 1H, J=8.8
Hz), 8.08 (d, 2H, J=8.4 Hz), 7.90(d, 2H, J=8.4 Hz), 7.85 (d, 1H,
J=8.8 Hz), 7.56 (d, 2H, J=8.8 Hz), 7.47-7.41 (m, 5H), 7.17 (d, 2H,
J=8.8 Hz), 5.18 (s, 2H), 4.40 (m, 1H), 2.31 (br m, 2H), 1.99 (br m,
4H), 1.80 (br m, 1H), 1.36 (br m, 3H); ESI-MS m/e 610.1(M+1); HPLC
purity (UV) 94%.
EXAMPLE 19
3-[2-(4-Benzyloxyphenyl)-1-cyclohexyl-1H-benzimidazole-5-sulfonylamino]-3--
oxo-propionic acid methyl ester
[0181] A solution of intermediate 69 (0.046 g, 0.1 mmol) in
dichloromethane (0.700 mL) was cooled to 0.degree. C. Lithium
bis(trimethylsilyl)amide (1.0 M solution in tetrahydrofuran) (0.10
ml, 0.10 mmol) was added and the mixture was stirred for 1 hr.
Methyl-3-chloro-3-oxo-propionate (0.011 ml, 0.10 mmol was added.
The reaction mixture was stirred at rt overnight. The reaction
mixture was concentrated and the residue purified by reverse phase
HPLC (C18 column, water/acetonitrile gradient containing 0.05%
trifluoroacetic acid) to afford the product (0.005 g, 7%). .sup.1H
NMR (CDCl.sub.3) .delta.8.60 (s, 1H), 8.16 (d, 1H, J=8.8 Hz), 7.85
(d, 1H, J=8.8 Hz), 7.64(d, 2H, J=8.8 Hz), 7.50-7.38 (m, 5H), 7.21
(d, 2H, J=8.8 Hz), 5.18 (s, 2H), 4.49 (m, 1H), 3.70 (s, 3H), 3.35
(s, 2H), 2.30 (br m, 2H), 2.02 (br m, 4H), 1.82 (br m, 1H), 1.35
(br m, 3H);ESI-MS m/e 562.18(M+1); HPLC purity (UV) 99%.
EXAMPLE 20
3-[2-(4-Benzyloxy-phenyl)-1-cyclohexyl-1H-benzoimidazole-5-sulfonylamino]--
3-oxo-propionic acid
[0182] The preceding ester was hydrolyzed to yield the product
(0.006 g) by the same method as in example 18. .sup.1H NMR
(CDCl.sub.3) .delta.8.60 (s, 1H), 8.19 (d, 1H, J=8.5 Hz), 7.97 (d,
1H, J=8.8 Hz), 7.79(d, 2H, J=7.6 Hz), 7.48-7.37 (m, 5H), 7.26 (m,
2H), 5.18 (s, 2H), 4.57 (m, 1H), 3.35 (bs, 2H), 2.32 (br m, 2H),
2.10 (br m, 2H), 2.02 (br m, 2H), 1.80 (br m, 1H), 1.37 (br m,
3H);ESI-MS m/e 548.04(M+1); HPLC purity (UV)>93%.
EXAMPLE 21
3-[2-(4-Benzyloxy-phenyl)-1-cyclohexyl-1H-benzoimidazole-5-sulfonylamino]--
3-oxo-acetic acid methyl ester
[0183] Intermediate 69 (0.046 g, 0.1 mmol) and methyl-oxalyl
chloride (0.010 ml, 0.11 mmol) were used to prepare the titled
compound (0.020 g, 30%). .sup.1H NMR (CDCl.sub.3) .delta.8.71 (s,
1H), 8.24 (d, 1H, J=8.8 Hz), 7.91 (d, 1H, J=8.8 Hz), 7.64(d, 2H,
J=8.8 Hz), 7.48-7.38 (m, 5H), 7.22 (d, 2H, J=8.8 Hz), 5.18 (s, 2H),
4.53 (m, 1H), 3.85 (s, 3H), 2.31 (br m, 2H), 2.09 (br m, 4H), 1.84
(br m, 1H), 1.38 (br m, 3H); ESI-MS m/e 548.17(M+1); HPLC purity
(UV)>78%.
EXAMPLE 22
[2-(4-Benzyloxy-phenyl)-1-cyclohexyl-1H-benzoimidazole-5-sulfonylamino]-3--
oxo-acetic acid
[0184] The preceding ester (0.012 g, 0.018 mmol) was used to
prepare the titled compound (0.005 g, 52%) by the same method as in
the previous examples. .sup.1H NMR (CDCl.sub.3) .delta.8.67 (s,
1H), .delta.8.47 (bs, 1H), 8.13 (d, 1H, J=8.8 Hz), 7.91 (m, 1H),
7.71(d, 2H, J=8.8 Hz), 7.49-7.37 (m, 5H), 7.23 (d, 2H, J=8.4 Hz),
5.17 (s, 2H), 4.53 (m, 1H), 2.31 (br m, 2H), 2.06 (br m, 4H), 1.81
(br m, 1H), 1.37 (br m, 3H); ESI-MS m/e 534.1(M+1); HPLC purity
(UV) 100%.
[0185] Other compounds prepared by these methods are included in
the following table.
1MASTER TABLE 19 Retention Ex. Synthetic Time Purity MS No. Q
R.sup.3 R.sup.4 R.sup.6 method (min) (AP) (M + 1) 23 CH cHexyl
4-ClPh CO.sub.2H C 2.55 98 605 24 CH cHexyl 4-FPh CO.sub.2H D 1.82
95 589 25 CH cHexyl 4-MeOPh CO.sub.2H D 1.92 90 601 26 CH cHexyl
4-MeSPh CO.sub.2H D 2.00 90 617 27 CH cHexyl 3.5-di-ClPh CO.sub.2H
D 2.09 95 639 28 CH cHexyl 1-naphthyl CO.sub.2H D 2 03 95 621 29 CH
cHexyl 2-biphenyl CO.sub.2H D 1.93 95 647 30 CH cHexyl 3-Cl-4-FPh
CO.sub.2H D 1 87 90 623 31 CH cHexyl 4-CF.sub.3Ph CO.sub.2H D 1.87
98 639 32 CH cHexyl 3-AcNHPh CO.sub.2H D 1 61 98 628 33 CH cHexyl
3-(OH)Ph CO.sub.2H D 1.78 94 587 34 CH cHexyl 4-(CN)Ph CO.sub.2H D
1.78 92 596 35 CH cHexyl Ph CO.sub.2H D 1.92 95 571 36 CH cHexyl
2-(CHO)Ph CO.sub.2H D 1.79 90 599 37 CH cHexyl 4-(CHO)Ph CO.sub.2H
D 1.80 82 599 38 CH cHexyl 3-(CO.sub.2H)Ph CO.sub.2H D 1 80 90 615
39 CH cHexyl 4-(CO.sub.2H)Ph CO.sub.2H D 1.78 94 614 40 CH cHexyl
4-MeSO.sub.2Ph CO.sub.2H D 1.69 92 648 41 CH cHexyl 4-NO.sub.2Ph
CO.sub.2H D 1.73 98 616 42 CH cHexyl 4-(CH.sub.2OH)Ph CO.sub.2H D 1
50 83 601 43 CH cHexyl 3-NH.sub.2-4-MePh CO.sub.2H D 1.40 90 600 44
CH cHexyl 3,4-di-MeOPh CO.sub.2H D 1.60 95 631 45 CH cHexyl
4-MeCOPh CO.sub.2H D 1.60 95 613 46 CH cHexyl 4-(OH)Ph CO.sub.2H D
1.53 95 587 47 CH cHexyl 4-(CH.dbd.CHCO.sub.2H)Ph CO.sub.2H D 1.60
95 641 48 CH cHexyl 4-ClPh 20 G 1 68 90 688 49 CH cHexyl 4-CNPh
CONHMe G 99 609 50 CH cHexyl 4-ClPh CONHMe G 99 618 51 CH cHexyl H
21 G 1 89 90 770 52 CH cHexyl 3-(OH)Ph CONHCH(Me)CO.sub.2H G 1.52
70 658 53 CH cHexyl 3-(OH)Ph 22 G 1.54 90 750 54 CH cHexyl 3-(OH)Ph
23 G 1.93 96 773 55 CH cHexyl 3-(OH)Ph
CONHCH.sub.2CH.sub.2CONH.sub.2 G 1 45 97 657 56 CH cHexyl
3-Cl-4-FPh CONHCH(Me)CO.sub.2H G 1.63 100 694 57 CH cHexyl
3-Cl-4-FPh 24 G 1.74 98 786 58 CH cHexyl 3-Cl-4-FPh 25 G 1.98 99
809 59 CH cHexyl 3-Cl-4-FPh CONHCH.sub.2CH.sub.2CONH.sub.2 G 1 58
99 693 60 CH cHexyl 3,4-di-MeOPh CONHCH(Me)CO.sub.2H G 1.44 90 701
61 CH cHexyl 3,4-di-MeOPh 26 G 1 50 100 794 62 CH cHexyl
3,4-di-MeOPh 27 G 1 88 100 816 63 CH cHexyl 3,4-di-MeOPh
CONHCH.sub.2CH.sub.2CONH.sub.2 G 1.54 90 701 64 CH cHexyl 3-(OH)Ph
28 G 1.46 100 669 65 CH cHexyl 3-(OH)Ph 29 G 1 53 100 656 66 CH
cHexyl 3-(OH)Ph 30 G 1.61 100 640 67 CH cHexyl 3-(OH)Ph
CONHCH.sub.2CH.sub.2CH.sub.2OH G 1.47 100 644 68 CH cHexyl 3-(OH)Ph
CONHMe G 1.59 100 600 69 CH cHexyl 3-(OH)Ph 31 G 1 59 100 670 70 CH
cHexyl 3-(OH)Ph CON(CH.sub.2CH.sub.2OH).sub.2 G 1.52 100 674 71 CH
cHexyl 3-(OH)Ph CONHCH(Me)CO.sub.2tBu G 1 72 86 714 72 CH cHexyl
3-(OH)Ph CONHCH.sub.2CO.sub.2tBu G 1 79 98 700 73 CH cHexyl
3-(OH)Ph 32 G 1 93 98 829 74 CH cHexyl 3-(OH)Ph
CONHCH.sub.2CO.sub.2H G 1 59 100 644 75 CH cHexyl 3-Cl-4-FPh 33 G
1.70 100 704 76 CH cHexyl 3-Cl-4-FPh 34 G 1 86 100 692 77 CH cHexyl
3-Cl-4-FPh 35 G 1.84 86 676 78 CH cHexyl 3-Cl-4-FPh
CONHCH.sub.2CH.sub.2CH.sub.2OH G 1.74 91 680 79 CH cHexyl
3-Cl-4-FPh CONHMe G 1 88 100 636 80 CH cHexyl 3-Cl-4-FPh 36 G 1.75
94 706 81 CH cHexyl 3-Cl-4-FPh CON(CH.sub.2CH.sub.2OH).sub.2 G 1 79
100 710 82 CH cHexyl 3-Cl-4-FPh CONHCH(Me)CO.sub.2tBu G 1.91 90 750
83 CH cHexyl 3-Cl-4-FPh 37 G 1 92 90 842 84 CH cHexyl 3-Cl-4-FPh
CONHCH.sub.2CO.sub.2tBu G 1.98 98 736 85 CH cHexyl 3-Cl-4-FPh 38 G
2.00 85 865 86 CH cHexyl 3-Cl-4-FPh CONHCH.sub.2CO.sub.2H G 1 83
100 680 87 CH cHexyl 3,4-di-MeOPh 39 G 1.39 100 712 88 CH cHexyl
3,4-di-MeOPh 40 G 1 67 95 700 89 CH cHexyl 3,4-di-MeOPh 41 G 1.76
100 684 90 CH cHexyl 3,4-di-MeOPh CONHCH.sub.2CH.sub.2CH.sub.2OH G
1.53 84 688 91 CH cHexyl 3,4-di-MeOPh CONHMe G 1.57 96 644 92 CH
cHexyl 3,4-di-MeOPh 42 G 1.54 95 713 93 CH cHexyl 3,4-di-MeOPh
CON(CH.sub.2CH.sub.2OH).sub.2 G 1 56 100 718 94 CH cHexyl
3,4-di-MeOPh CONHCH(Me)CO.sub.2tBu G 1.78 85 758 95 CH cHexyl
3,4-di-MeOPh 43 G 1 79 89 850 96 CH cHexyl 3,4-di-MeOPh
CONHCH.sub.2CO.sub.2tBu G 1.71 93 744 97 CH cHexyl 3,4-di-MeOPh 44
G 1.88 95 873 98 CH cHexyl 3,4-di-MeOPh CONHCH.sub.2CO.sub.2H G 1
62 100 688 99 CH cHexyl 4-ClPh 45 G 1 74 95 688 100 CH cHexyl
4-MeSPh CONH.sub.2 G 1 06 90 616 101 CH cHexyl 4-MeSPh
CONHCH.sub.2CH.sub.2CONH.sub.2 G 1 05 93 681 102 CH cHexyl 4-MeSPh
46 G 1 05 95 700 103 CH cHexyl 4-MeSPh CONHCH.sub.2CO.sub.2tBu G
1.10 90 730 104 CH cHexyl 4-MeSPh CONMe.sub.2 G 1.07 96 644 105 CH
cHexyl 4-CNPh CONMe.sub.2 G 1 59 85 623 106 CH cHexyl 4-ClPh CONHMe
G 1.73 90 618 107 CH cHexyl 4-ClPh 47 G 1 71 90 688 108 CH cHexyl
4-ClPh CONHCH.sub.2CO.sub.2tBu G 1.85 75 718 109 CH cHexyl 4-ClPh
CONHCH.sub.2CO.sub.2H G 1 71 90 662 110 CH cHexyl 4-ClPh CONH.sub.2
G 1.71 90 604 111 CH cHexyl 4-ClPh CONHCH.sub.2CH.sub.2NH.sub.2 G 1
68 85 675 112 CH cHexyl 4-ClPh CONMe.sub.2 G 1 75 85 632 113 CH
cHexyl 4-MeSPh CONHMe G 1 06 90 630 114 CH cHexyl 2-thienyl
CO.sub.2H D 1 78 95 577 115 CH cHexyl 3-thienyl CO.sub.2H D 1.77 95
577 116 CH cHexyl 2-benzofuranyl CO.sub.2H D 1.91 90 611 117 CH
cHexyl 2-furanyl CO.sub.2H D 1 86 85 561 118 CH cHexyl
3,4-dioxolanePh CO.sub.2H D 1.77 98 615 119 CH cHexyl
2-benzothiophenyl CO.sub.2H D 2 07 95 627 120 CH cHexyl
5-Ac-2-thienyl CO.sub.2H D 1.91 98 619 121 CH cHexyl 5-indolyl
CO.sub.2H D 1 90 97 610 122 CH cHexyl 3-CHO-2-furanyl CO.sub.2H D
1.65 85 589 123 CH cHexyl 8-quinolinyl CO.sub.2H D 1 67 90 622 124
CH cHexyl 2-indolyl CO.sub.2H D 1.75 95 609 125 CH cHexyl
2,4-di-MeO-5-pyrimidinyl CO.sub.2H D 1 91 95 632 126 CH cHexyl
N-BOC-2-pyrrolyl CO.sub.2H D 1 81 92 659 127 CH cHexyl
N-Me-5-indolyl CO.sub.2H D 1.80 98 623 128 CH cHexyl 5-pyrimidinyl
CO.sub.2H D 1.60 98 572 129 CH cHexyl 3-thienyl 48 G 1.54 92 740
130 CH cHexyl 3-thienyl 49 G 1.91 100 763 131 CH cHexyl 3-thienyl
CONHCH.sub.2CH.sub.2CONH.sub.2 G 1.60 98 647 132 CH cHexyl
3-thienyl 50 G 1.56 100 659 133 CH cHexyl 3-thienyl 51 G 1.74 100
646 134 CH cHexyl 3-thienyl 52 G 1.72 100 630 135 CH cHexyl
3-thienyl CONHCH.sub.2CH.sub.2CH.sub.2OH G 1 71 92 634 136 CH
cHexyl 3-thienyl CONHMe G 1.73 100 590 137 CH cHexyl 3-thienyl 53 G
1 71 95 660 138 CH cHexyl 3-thienyl CON(CH.sub.2CH.sub.2OH).sub.2 G
1.63 100 664 139 CH cHexyl 3-thienyl CONH(Me)CO.sub.2tBu G 1.80 77
704 140 CH cHexyl 3-thienyl 54 G 1.91 92 796 141 CH cHexyl
3-thienyl CONHCH.sub.2CO.sub.2tBu G 1 86 94 690 142 CH cHexyl
3-thienyl 55 G 1 90 95 819 143 CH cHexyl 3-thienyl
CONHCH.sub.2CO.sub.2H G 1 58 98 634 144 CH cHexyl 5-Ac-2-thienyl
CONHCH.sub.2CO.sub.2tBu G 1.55 95 731 145 CH cHexyl 5-Ac-2-thienyl
CONHCH.sub.2CO.sub.2H G 1 48 90 676 146 CH cHexyl 5-Ac-2-thienyl
CONH.sub.2 G 1.58 94 618 147 CH cHexyl 5-Ac-2-thienyl
CONHCH.sub.2CH.sub.2CONH.sub.2 G 1 44 91 689 148 CH cHexyl
5-Ac-2-thienyl CONMe.sub.2 G 1.37 85 646 149 CH cHexyl 4-(CN)Ph 56
G 1.36 95 679 150 CH cHexyl 5-Ac-2-thienyl CONHMe G 1.51 92 632 151
CH cHexyl 5-Ac-2-thienyl 57 G 1 47 90 702 152 CH cHexyl
3-CF.sub.3CO-5-indolyl CONMe.sub.2 G 1 58 100 733 153 CH cHexyl
3-CF.sub.3CO-5-indolyl CONHCH.sub.2CO.sub.2H G 1 55 83 763 154 CH
cHexyl 3-CF.sub.3CO-5-indolyl CONH.sub.2 G 1 69 86 706 155 CH
cHexyl 3-CF.sub.3CO-5-indolyl CONHCH.sub.2CH.sub.2CONH.sub.2 G 1 52
80 776
UTILITY
[0186] The compounds of Formula I inhibit the activity of Hepatitis
C Virus NS5B RdRp as demonstrated using assays for NS5B RdRp
activity. Thus, the compounds of Formula I are potentially useful
in the cure and prevention of HCV infections.
[0187] HCV NS5B RdRp cloning, expression and purification. The cDNA
encoding the NS5B protein of HCV, genotype 1b, was cloned into the
pET21a expression vector. The protein was expressed with an 18
amino acid C-terminal truncation to enhance the solubility. The E.
coli competent cell line BL21(DE3) was used for expression of the
protein. Cultures were grown at 37.degree. C. for .about.4 hours
until the cultures reached an optical density of 2.0 at 600 nm. The
cultures were cooled to 20.degree. C. and induced with 1 mM IPTG.
Fresh ampicillin was added to a final concentration of 50 ug/ml and
the cells were grown overnight at 20.degree. C.
[0188] Cell pellets (3L) were lysed for purification to yield 15-24
mgs of purified NS5B. The lysis buffer consisted of 20 mM Tris-HCl,
pH 7.4, 500 mM NaCl, 0.5% triton X-100, 1 mM DTT, 1 mM EDTA, 20%
glycerol, 0.5 mg/ml lysozyme, 10 mM MgCl2, 15 ug/ml
deoxyribonuclease I, and Complete TM protease inhibitor tablets
(Roche). After addition of the lysis buffer, frozen cell pellets
were resuspended using a tissue homogenizer. To reduce the
viscosity of the sample, aliquots of the lysate were sonicated on
ice using a microtip attached to a Branson sonicator. The sonicated
lysate was centrifuged at 100,000.times.g for 1 hr at 4.degree. C.
and filtered through a 0.2 .mu.m filter unit (Corning).
[0189] The protein was purified using three sequential
chromatography steps: Heparin sepharose CL-6B, polyU sepharose 4B,
and Hitrap SP sepharose (Pharmacia). The chromatography buffers
were identical to the lysis buffer but contained no lysozyme,
deoxyribonuclease I, MgCl2 or protease inhibitor and the NaCl
concentration of the buffer was adjusted according to the
requirements for charging the protein onto the column. Each column
was eluted with a NaCl gradient which varied in length from 5-50
column volumes depending on the column type. After the final
chromatography step, the resulting purity of the enzyme is >90%
based on SDS-PAGE analysis. The enzyme was aliquoted and stored at
-80.degree. C.
[0190] HCV NS5B RdRp enzyme assay. HCV RdRp genotype 1b assays were
run in assay buffer composed of 20 mM Tris-HCl, pH 7.5, 2.5 mM KCl,
5 mM MgCl.sub.2, 1 mM DTT, 1.6 U RNAse inhibitor (Promega N2515),
in 96 well plates (Falcon 3918). All compounds were serially
diluted in DMSO and diluted further in assay buffer such that the
final concentration of DMSO in the assay was 2%. Compounds were
serially diluted (3-fold each time) for a 7 point inhibition
analysis. HCV RdRp genotype 1b enzyme was used at a final
concentration of 28 nM. A polyA template was used at 28 nM, and the
oligo-dTI.sub.12-18 primer was used at 840 nM final concentration.
Preannealed primer and template were obtained commercially
(Amersham 27-787802). .sup.3H-UTP was used at 0.125 .mu.Ci (1 .mu.M
total UTP). Reaction was initiated by the addition of enzyme.
Reactions were incubated at 30.degree. C. for 45 min, and stopped
by adding 30 ul of 20% ice cold TCA. Plates were chilled for 30
minutes and harvested onto Unifilter-96, GF/B plates(Packard,
6005177) using a Packard FilterMate Cell Harvester. The harvest
plates were prewashed 3 times, 200 ul/well, with 100 mM NaPPi.
Harvested filters were washed 30 times, 200 ul/well, with distilled
water followed by ethanol. Filter plates were dried, and 30 ul
/well microscint-20 was added. Plates were read on a Packard Top
Count NXT.
[0191] The IC.sub.50 values for compounds were determined using six
different [I], combined with 7nM enzyme, 800 ng of the
template-primer polyC/oligoG.sub.12 (1:5 molar ratio), and 0.7 uM
of .sup.3H GTP containing 1 uCi. The observed fractional activity
(fa=vi/vo) was used in the equation IC.sub.50=[I]/(1/fa-1) to
determine a single point IC.sub.50 value. Typically, the single
point IC.sub.50 values derived from [I] that produced fractional
activities in the range of 0.1 to 0.8 relative to the uninhibited
control were averaged to calculate the IC.sub.50 value for each
compound.
[0192] The HCV NS5B RdRp enzyme assay results are tabulated in the
following table.
2 Example IC50 No. (.mu.M) 1 0.63 3 0.14 4 0.63 5 0.36 10 0.85 17
5.21 18 1.52 20 8.20 22 8.09 23 0.02 24 0.05 25 0.04 26 0.19 27
1.01 28 0.62 29 0.25 30 0.04 31 0.06 32 0.04 33 0.04 34 0.04 35
0.07 36 0.83 37 0.07 38 0.00 39 0.01 40 0.08 41 0.05 42 0.05 43
0.24 44 0.08 45 0.08 46 0.06 47 0.05 48 0.32 49 0.08 50 0.23 51
0.93 52 0.47 53 0.57 54 0.68 55 0.20 56 0.19 57 0.18 58 0.38 59
0.92 60 0.28 61 0.09 62 0.11 63 0.15 64 1.20 65 1.70 66 1.40 67
0.79 68 0.54 69 0.66 70 0.50 71 0.52 72 0.93 73 1.30 74 0.15 75
0.35 76 1.40 77 1.20 78 0.69 79 0.55 80 0.94 81 1.10 82 1.30 83
0.56 84 1.20 85 2.30 86 0.32 87 0.43 88 0.47 89 0.46 90 0.63 91
0.26 92 0.30 93 0.69 94 2.10 95 1.40 96 1.60 97 2.20 98 0.21 99
0.86 106 0.66 114 0.07 115 0.04 116 0.17 117 0.29 118 0.07 119 0.03
120 0.02 121 0.03 122 0.67 123 0.44 124 0.12 125 1.28 126 0.46 127
0.09 128 1.04 129 0.29 130 0.58 131 0.23 132 0.83 133 1.10 134 0.37
135 0.70 136 0.22 137 1.30 138 3.00 139 2.60 140 2.15 141 2.80 142
1.20 143 0.08 149 0.22 152 0.42 153 0.26 154 0.33 155 0.27
* * * * *