Note: Descriptions are shown in the official language in which they were submitted.
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PYRIMIDINYLIMIDAZOLES AS TGF-BETA INHIBITORS
[0001] This application claims priority to U.S. Serial No. 60/606,045, which
was filed on
August 31, 2004. The entire contents of the aforementioned application are
incorporated herein
by reference.
BACKGROUND OF THE INVENTION
[0002] TGFO (Transforming Growth Factor (3) is a member of a large family of
dimeric
polypeptide growth factors that includes, for example, activins, inhibins,
bone morphogenetic
proteins (BMPs), growth and differentiation factors (GDFs) and mullerian
inhibiting substance
(MIS). TGFO exists in three isoforms (TGFo1, TGF02, and TGF,(33) and is
present in most cells,
along with its receptors. Each isoform is expressed in both a tissue-specific
and
developmentally regulated fashion. Each TGFO isoform is synthesized as a
precursor protein
that is cleaved intracellularly into a C-terminal region (latency associated
peptide (LAP)) and an
N-terminal region known as mature or active TGF,6. LAP is typically non-
covalently associated
with mature TGFO prior to secretion from the cell. The LAP-TGF6 complex cannot
bind to the
TGFO receptors and is not biologically active. TGFO is generally released (and
activated) from
the complex by a variety of mechanisms including, for example, interaction
with
thrombospondin-1 or plasmin.
[0003] Following activation, TGFO binds at high affinity to the type II
receptor (TGFRRII), a
constitutively active serine/threonine kinase. The ligand-bound type II
receptor phosphorylates
the TGFO type I receptor (Alk 5) in a glycine/serine rich domain, which allows
the type I
receptor to recruit and phosphorylate downstream signaling molecules, Smad2 or
Smad3. See,
e.g., Huse, M. et al.,lllol. Cell. 8: 671-682 (2001). Phosphorylated Smad2 or
Smad3 can then
complex with Smad4, and the entire hetero-Smad complex translocates to the
nucleus and
regulates transcription of various TGF(3-responsive genes. See, e.g.,
Massague, J. Ann. Rev
.Biochein. Med. 67: 773 (1998).
[0004] Activins are also members of the TGFO superfamily, which are distinct
from TGFO in
that they are homo- or heterodimers of activin (3a or Ob. Activins signal in a
manner similar to
TGFO , that is, by binding to a constitutive serine-threonine receptor kinase,
activin type II
receptor (ActRIIB), and activating a type I serine-threonine receptor, Alk 4,
to phosphorylate
Smad2 or Smad3. The consequent formation of a hetero-Smad complex with Smad4
also results
in the activin-induced regulation of gene transcription.
[0005] Indeed, TGFO and related factors such as activin regulate a large array
of cellular
processes, e.g., cell cycle arrest in epithelial and hematopoietic cells,
control of mesenchymal
-1-
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WO 2006/026306 PCT/US2005/030133
cell proliferation and differentiation, inflammatory cell recruitment,
immunosuppression, wound
healing, and extracellular matrix production. See, e.g., Massague, J. Ann.
Rev. .Cell. Biol. 6:
594-641 (1990); Roberts, A. B. and Sporn M. B. Peptide Growtlz Factors and
Tlzeiz- Receptors,
95: 419-472 Berlin: Springer-Verlag (1990); Roberts, A. B. and Sporn M. B.
Growtlz Factors
8:1-9 (1993); and Alexandrow, M. G., Moses, H. L. Cancer Res. 55: 1452-1457
(1995).
Hyperactivity of TGFO signaling pathway underlies many human disorders (e.g.,
excess
deposition of extracellular matrix, an abnormally high level of inflammatory
responses, fibrotic
disorders, and progressive cancers). Similarly, activin signaling and
overexpression of activin is
linked to pathological disorders that involve extracellular matrix
accumulation and fibrosis (see,
e.g., Matsuse, T. et al., Am. J. Respir. Cell Mol. Biol. 13: 17-24 (1995);
Inoue, S. et al.,
Biochem. Bioplzys. Res. Coznzn. 205: 441-448 (1994); Matsuse, T. et al, Am. J.
Pathol. 148: 707-
713 (1996); De Bleser et al., Hepatology 26: 905-912 (1997); Pawlowski, J.E.,
et al., J. Clin.
Invest. 100: 639-648 (1997); Sugiyama, M. et al., Gastroenterology 114: 550-
558 (1998); Munz,
B. et al., EMBO J. 18: 5205-5215 (1999)), inflammatory responses (see, e.g.,
Rosendahl, A. et
al., Am. J Repir. Cell Mol. Biol. 25: 60-68 (2001)), cachexia or wasting (see
Matzuk, M. M. et
al., Proc. Nat. Acad. Sci. USA 91: 8817-8821 (1994); Coerver, K.A. et al, Mol.
Endocrinol. 10:
534-543 (1996); Cipriano, S.C. et al. Endocz izzology 141: 2319-27 (2000)),
diseases of or
pathological responses in the central nervous system (see Logan, A. et al.
Eur. J. Neurosci. 11:
2367-2374 (1999); Logan, A. et al. Exp. Neui~ol. 159: 504-510 (1999); Masliah,
E. et al.,
Neurochem. Int. 39: 393-400 (2001); De Groot, C. J. A. et al, J. Neuropathol.
Exp. Neurol. 58:
174-187 (1999), John, G. R. et al, Nat Med. 8: 1115-21 (2002)) and
hypertension (see Dahly, A.
J. et al., Azn. J. Physiol. Regul. Izztegz . Comp. Physiol. 283: R757-67
(2002)). Studies have
shown that TGFO and activin can act synergistically to induce extracellular
matrix production
(see, e.g., Sugiyama, M. et al., Gastroenterology 114: 550-558, (1998)). It is
therefore desirable
to develop modulators (e.g., antagonists) to members of the TGFO family to
prevent and/or treat
disorders involving this signaling pathway.
SUMMARY OF THE 1NVENTION
[0006] The invention is based on the discovery that compounds of formula (I)
are unexpectedly
potent antagonists of the TGFO family type I receptors, A1k5 and/or Alk 4.
Thus, compounds of
formula (I) can be employed in the prevention and/or treatment of diseases
such as fibrosis (e.g.,
renal fibrosis, pulmonary fibrosis, and hepatic fibrosis), progressive
cancers, or other diseases
for which reduction of TGFO family signaling activity is desirable.
[0007] In one aspect, a compound of the following formula:
-2-
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(Ra)m
N~ N
2
(:~--X-Y-R-
A'
R'
where Rl can be heteroaryl.
[0008] Each Ra, independently, can be alkyl, alkenyl, alkynyl, alkoxy, acyl,
halo, hydroxy,
amino, nitro, oxo, thioxo, cyano, guanadino, amidino, carboxy, sulfo,
mercapto, alkylsulfanyl,
alkylsulfinyl, alkylsulfonyl, aminocarbonyl, alkylcarbonylamino,
arylcarbonylamino,
heteroarylcarbonylamino, alkylsulfonylamino, arylsulfonylamino,
heteroarylsulfonylamino,
alkoxycarbonyl, alkylcarbonyloxy, urea, thiourea, sulfamoyl, sulfamide,
carbamoyl, cycloalkyl,
cycloalkyloxy, cycloalkylsulfanyl, cycloalkylcarbonyl, heterocycloalkyl,
heterocycloalkyloxy,
heterocycloalkylsulfanyl, heterocycloalkylcarbonyl, aryl, aryloxy,
arylsulfanyl, aroyl, heteroaryl,
heteroaryloxy, heteroarylsulfanyl, or heteroaroyl.
[0009] X can be cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or a bond.
[0010] Y can be a bond, -C(O)-, -C(O)-O-, -O-C(O)-, -S(O)p O-, -O-S(O)P-, -
C(O)-N(Rb)-,
-N(Rb)-C(O)-, -O-C(O)-N(Rb)-, -N(Rb)-C(O)-0-, -C(O)-N(Rb)-0-, -O-N(Rb)-C(O)-,
-O-S(O)P-N(Rb)-, -N(Rb)- S(O)P O-, -S(O)P N(Rb)-0-, -O-N(R)-S(O)p , -N(R)-C(O)-
N(R )-,
-N(Rt')-S(O)p N(R )-, -C(O)-N(Rb)-S(O)p , -S(O)p N(R)-C(O)-, -C(O)-N(R)-S(O)p-
N(R )-,
-C(O)-O-S(O)p N(Rb)-, -N(Rb)-S(O)P N(R )-C(O)-, -N(Rb)-S(O)p O-C(O)-, -S(O)p
N(Rb)-,
-N(Rb)-S(O)P ,-N(Rb)-, -S(O)P-, -0-, -S-, or -(C(Rb)(R ))q-. Each of Rb and
Rc, independently,
can be hydrogen, hydroxy, alkyl, alkoxy, amino, aryl, aralkyl,
heterocycloalkyl, heteroaryl, or
heteroaralkyl. p can be 1 or 2, and q can be 1-4.
[0011] R2 can be hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl,
(cycloalkyl)alkyl, cycloalkenyl,
(cycloalkenyl)alkyl, aryl, aralkyl, arylalkenyl, heterocycloalkyl,
(heterocycloalkyl)alkyl,
heterocycloalkenyl, (heterocycloalkenyl)alkyl, heteroaryl, heteroaralkyl, or
(heteroaryl)alkenyl.
[0012] Each of A' and A2, independently, can be N or NRb. It is to be
understood that when A'
is NRb, A2 is N, and vice versa. The variable, m, can be 0, 1, 2, or 3. In
other words, the
pyrimidinyl ring can be unsubstituted or substituted with 1-3 Ra groups. Note
that when mL-2,
two adjacent Ra groups can join together to form a 4- to 8-membered optionally
substituted
cyclic moiety. That is, the pyrimidinyl ring can fuse with a cyclic moiety to
form a moiety, that
can be optionally substituted with one or more substituents such as alkyl
(including
-3-
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carboxyalkyl, hydroxyalkyl, and haloalkyl such as trifluoromethyl), alkenyl,
alkynyl, cycloalkyl,
heterocycloalkyl, alkoxy, aryl, heteroaryl, aryloxy, heteroaryloxy, aroyl,
heteroaroyl, amino,
nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl,
alkylcarbonylamino, cyano,
halo, hydroxy, acyl, mercapto, alkylthio, sulfoxy, sulfamoyl, oxo, or
carbamoyl.
[0013] Note also that if X is a bond, then Y is a bond; Rz is hydrogen or
alkyl; m is 1, 2, or 3;
and at least one Ra is substituted at the 2-pyrimidinyl position (i.e., the
position of the
pyrimidinyl ring that is between the two nitrogen ring atoms).
[0014] In an embodiment, X can be aryl or heteroaryl. For example, X can be an
optionally
substituted phenyl (e.g., alkyl or cyano). Y can be a bond, -N(R)-C(O)-, -N(R)-
S(O)2-, -C(O)-,
-C(O)-0-, -O-C(O)-, -C(O)-N(R)-, -S(O)P , -0-, -S(O)Z-N(Rb)-, - N(Rb)-, -N(Rb)-
C(O)-0-,
-N(R)-C(O)-N(Rc)-, -C(O)-N(R)-S(O)p N(R )-, or -C(O)-O-S(O)p N(R)-. R2 can be
hydrogen, C1_6 alkyl, aryl, heteroaryl, aryl-Cl-4 alkyl, or heteroaryl-Cl.4
alkyl.
[0015] In an embodiment, X can be a 4- to 8-membered monocyclic cycloalkyl or
heterocycloalkyl, or X can be a 4- to 8-membered bicyclic cycloalkyl or
heterocycloalkyl. For
example, X can be piperidinyl, piperazinyl, pyrrolidinyl, tetrahydrofuran,
cyclohexyl,
cyclopentyl, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2. 1
]octane,
2-oxa-bicyclo[2.2.2]octane, 2-aza-bicyclo[2.2.2]octane, 3-aza-bicyclo[3.2. 1
]octane, or
1 -aza-bicyclo [2.2.2] octane.
[0016] In an embodiment, X can be piperidinyl, piperazinyl, or pyrrolidinyl.
The piperdinyl,
piperazinyl, or pyrrolidinyl can be bonded to Y via its nitrogen ring atom. Y
can be a bond,
-C(0)0-, -C(O)-N(Rb)-, -S(0)2-, or -S(O)2-N(R)-, wherein Rb is hydrogen or C1-
4 alkyl.
Alternatively, X can be cyclohexyl, cyclopentyl, or bicyclo[2.2.2]octane, and
Y can be
-N(Rb)-C(O)-, -N(R)-S(O)2-, -C(O)-, -C(O)-0-, -O-C(O)-, -C(O)-N(Rb)-, -S(O)p ,
-0-,
-S(O)Z-N(Rb)-, - N(Rb)-, -N(R)-C(O)-0-, -C(O)-N(Rb)-0-, or -N(R)-C(O)-N(R )-.
[0017] In an embodiment, Y can be -N(R)-C(O)-, -N(Rb)-S(O)2-, -C(O)-, -C(O)-0-
, -O-C(O)-,
-C(O)-N(R)-, -S(O)P , -0-, -S(O)2-N(R)-, - N(Rb)-, -N(R)-C(O)-0-, -C(O)-N(R)-0-
,
-N(R)-C(O)-N(R )-, -C(O)-N(R)-S(O)P N(R )-, or -C(O)-O-S(O)p-N(R)-.
[0018] In an embodiment, X and Y are each a bond; R2 can be hydrogen or C1_6
alkyl (e.g., C1-4
alkyl such as methyl or t-butyl); m can be 1 or 2 (e.g., m can be 1); at least
one Ra is substituted
at the 2-pyrimidinyl position and this Ra can be Cl-4 alkyl, C3_6 cycloalkyl,
or amino (e.g, -CH3, -
CF3, cyclopropyl, -NHZ, -NH-C1-4 alkyl, or -NH-cycloalkyl such as -NH-
cyclopropyl).
[0019] In an embodiment, RZ can be hydrogen, C1_6 alkyl, aryl, heteroaryl,
aryl-C1-4 alkyl, or
heteroaryl-C1-4 alkyl. In an embodiment, RZ can be hydrogen, C1.4 alkyl,
phenyl, pyridyl,
imidazolyl, furanyl, thienyl, triazolyl, tetrazolyl, benzyl, phenylethyl,
benzimidazolyl,
-4-
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benzothiazolyl, naphthylmethyl, naphthylethyl, or -CI_2 alkyl-pyridyl; each of
which,
independently, is optionally substituted with one or more substituents
selected from the group
consisting of fluoro, chloro, trifluoromethyl, methyl, ethyl, aminocarbonyl,
alkylcarbonylamino,
sulfamoyl, alkoxycarbonyl, and alkylcarbonyloxy.
[0020] In another embodiment, R'' can be hydrogen, methyl, ethyl, n-butyl, t-
butyl, benzyl or
pyridylmethyl. For example, R2 can be hydrogen, hydroxymethyl, or
trifluoromethyl.
[0021] In an embodiment, R' can be benzo[1,3]dioxolyl, benzo[b]thiophenyl,
benzo-oxadiazolyl, benzothiadiazolyl, benzoimidazolyl, benzooxazolyl,
benzothiazolyl,
2-oxo-benzooxazolyl, pyridyl, pyrimidinyl, 2,3-dihydro-benzo[1,4]dioxyl,
2,3-dihydro-benzofuryl, 2,3-dihydro-benzo[b]thiophenyl, 3,4-dihydro-
benzo[1,4]oxazinyl,
3-oxo-benzo[1,4]oxazinyl, 1,1-dioxo-2,3-dihydro- benzo[b]thiophenyl,
[1,2,4]triazolo[1,5-a]pyridyl, [1,2,4]triazolo[4,3-a]pyridyl, quinolinyl,
quinoxalinyl,
quinazolinyl, isoquinolinyl, or cinnolinyl.
[0022] In an embodiment, m can be 0-2.
[0023] In an embodiment, Ra can be substituted at the 2-pyrimidinyl position.
[0024] In another embodiment, Ra can be C1-4 alkyl, C1-4 alkoxy, C1-4
alkylthio, halo, amino,
aminocarbonyl, or alkoxycarbonyl.
[0025] In an embodiment, Al can be N and A2 is NRb, or Al is NRb and A2 is N;
wherein Rb is
hydrogen or C 1-4 alkyl.
[0026] In an embodiment, m can be 0-2; Rl can be heteroaryl; R2 can be
hydrogen, C1_6 alkyl,
aryl, heteroaryl, -C1-4 alkyl-aryl, or -C1-4 alkyl-heteroaryl; X can be a 4-
to 8-membered
monocyclic or bicyclic cycloalkyl or heterocycloalkyl; and Y can be -N(R)-C(O)-
,
-N(R)-S(O)Z-, -C(O)-, -C(O)-O-, -O-C(O)-, -C(O)-N(R)-, -S(O)P , -0-, -S(O)2-
N(R)-,
-N(R)-, -N(R)-C(O)-0-, -N(Rb)-C(O)-N(Rc)-, -C(O)-N(R)-S(O)p N(Rc)-, or
-C(O)-O-S(O)p N(Rb)-.
[0027] In an embodiment, m can be 0-2; Rl can be heteroaryl; RZ can be
hydrogen, C1_6 alkyl,
aryl, heteroaryl, -C1-4 alkyl-aryl, or -C1-4 alkyl-heteroaryl; X can be
piperidinyl, piperazinyl,
pyrrolidinyl, tetrahydrofuran, cyclohexyl, cyclopentyl, bicyclo[2.2. 1
]heptane,
bicyclo[2.2.2]octane, bicyclo[3.2.1]octane, 2-oxa-bicyclo[2.2.2]octane,
2-aza-bicyclo[2.2.2]octane, 3-aza-bicyclo[3.2.1]octane, or 1 -aza-
bicyclo[2.2.2] octane; and Y can
be -N(Rb)-C(O)-, -N(R)-S(O)2-, -C(O)-, -C(O)-0-, -O-C(O)-, -C(O)-N(R)-, -S(O)p
, -0-,
-S(O)2-N(R)-, - N(R)-, -N(R)-C(O)-0-, -N(R)-C(O)-N(W)-, -C(O)-N(R)-S(O)P N(R )-
, or
-C(O)-O-S(O)p N(R)-.
[0028] In an embodiment, m can be 0-2; Rl can be heteroaryl; R2 can be
hydrogen, C1.6 alkyl,
-5-
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aryl, heteroaryl, -C1-4 alkyl-aryl, or -C1-4 alkyl-heteroaryl; and -X-Y- can
be
0
~-o-
I I N
N-SOa N-C-O-
,
/ SOz
N 11 ZC N-SO2- N C 0-
, or
[0029] In an embodiment, Al can be N and A2 can be NH. Alternatively, Al can
be NH and A2
can be N. RZ can be hydrogen, C1-4 alkyl, benzyl, or pyridylmethyl; m can be 1
and Ra can be
substituted at the 2-pyrimidinyl position.
[0030] In an embodiment, m can be 0-2; Rl can be heteroaryl; R' can be
hydrogen, C1_6 alkyl,
aryl, heteroaryl, aryl-C1.4 alkyl, or heteroaryl-C1-4 alkyl; X can be
cyclohexyl, cyclopentyl, or
bicyclo[2.2.2]octane; and Y can be -N(Rb)-C(O)-, -N(R)-S(O)2-, -C(O)-, -C(O)-O-
, -O-C(O)-,
-C(O)-N(R)-, -S(O)P , -0-, -S(O)2-N(R)-, - N(Rb)-, -N(R)-C(O)-0-, -N(Rb)-C(O)-
N(R )-,
-C(O)-N(Rb)-S(O)p N(R )-, or -C(O)-O-S(O)p N(R)-. Each of Rb and R ,
independently, can be
hydrogen or C1-4 alkyl. Al can be N and A2 can be NH, or alternatively, A' can
be NH and A2
can be N. RZ can be hydrogen, CI-4 alkyl, benzyl, or pyridylmethyl; m can be 1
and Ra can be
substituted at the 2-pyrimindyl position.
[0031] In an embodiment, X and Y can each be a bond; RZ can be hydrogen or C1-
4 alkyl; m can
be 1; Ra can be -CH3, -CF3, cyclopropyl, -NH2, -NH-C1-4 alkyl, or -NH-
cycloalkyl; and Rl can
be benzo[1,3]dioxolyl, benzo[b]thiophenyl, benzooxadiazolyl,
benzothiadiazolyl,
benzoimidazolyl, benzooxazolyl, benzothiazolyl, 2-oxo-benzooxazolyl, pyridyl,
pyrimidinyl,
2,3-dihydro-benzo[1,4]dioxyl, 2,3-dihydro-benzofuryl, 2,3-dihydro-
benzo[b]thiophenyl,
3,4-dihydro-benzo[1,4]oxazinyl, 3-oxo-benzo[1,4]oxazinyl, 1,1-dioxo-2,3-
dihydro-
benzo[b]thiophenyl, [1,2,4]triazolo[1,5-a]pyridyl, [1,2,4]triazolo[4,3-
a]pyridyl, quinolinyl,
quinoxalinyl, quinazolinyl, isoquinolinyl, or cinnolinyl.
[0032] The compound of formula (I) can be:
[0033] 4-[4-benzo[1,3]dioxol-5-yl-5-(2-methylsulfanyl-pyrimidin-4-yl)-1H-
imidazol-2-yl]-
benzamide;
[0034] 4-[4-benzo[1,3]dioxol-5-yl-5-(2-methylsulfanyl-pyrimidin-4-yl)-1H-
imidazol-2-yl]-
benzonitrile;
[0035] 4-[5-(2-methanesulfonyl-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-yl]-
-6-
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bicyclo[2.2.2]octane-l-carboxylic acid methyl ester;
[0036] 4-[5-(2-methoxy-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-imidazol-2-
yl]-
bicyclo[2.2.2]octane-1-carboxylic acid methyl ester;
[0037] 4-[5-(2-hydroxy-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-imidazol-2-
yl]-
bicyclo[2.2.2]octane-1-carboxylic acid methyl ester;
[0038] 4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester;
[0039] 4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-yl]-
bicyclo [2.2.2] octane- 1 -carboxylic acid;
[0040] 4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid amide;
[0041] 4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-yl]-
bicyclo [2.2.2] octane- 1 -carboxylic acid hydroxyamide;
[0042] 4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-iH-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methoxy-amide;
[0043] 4-[5-(2-amino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-imidazol-2-
yl]-
bicyclo[2.2.2]octane-l-carboxylic acid;
[0044] {4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-
yl]-bicyclo[2.2.2]oct-1-yl}-carbamic acid benzyl ester;
[0045] N-{4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-
1H-imidazol-
2-yl]-bicyclo[2.2.2]oct-l-yl}-acetamide;
[0046] N- {4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-
1 H-imidazol-
2-yl] -bicyclo [2.2.2] oct-1-yl } -methane sulfonarnide;
[0047] N-{4-[5-(2-cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-
1H-imidazol-
2-yl]-bicyclo[2.2.2]oct-1-yl} -2,2,2-trifluoro-acetamide;
[0048] 4-[5-quinoxalin-6-yl-4-(2-trifluoromethyl-pyrimidin-4-yl)-1H-imidazol-2-
yl]-
bicyclo[2.2.2]octan-l-ol;
[0049] 4-[4-(2-cyclopropyl-pyrimidin-4-yl)-5-quinoxalin-6-yl-lH-imidazol-2-yl]-
bicyclo[2.2.2]octan-l-ol;
[0050] 6-[2-tert-butyl-5-(2-cyclopropyl-pyrimidin-4-yl)-3H-imidazol-4-yl]-
quinoxaline;
(15686)
[0051] 6-[5-(2-byclopropyl-pyrimidin-4-yl)-3H-imidazol-4-yl]-quinoxaline;
[0052] {4-[4-(2-cyclopropyl-pyrimidin-4-yl)-5-quinoxalin-6-yl-lH-imidazol-2-
yl]-
bicyclo [2.2.2] o ct-1-yl } -methanol;
-7-
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[0053] 6-[5-(2-trifluoromethyl-pyrimidin-4-yl)-3H-imidazol-4-yl]-quinoxaline;
[0054] 6-[2-tert-butyl-5-(2-trifluoromethyl-pyrimidin-4-yl)-3H-imidazol-4-yl]-
quinoxaline;
[0055] 4-[5-quinoxalin-6-y1-4-(2-trifluoromethyl-pyrimidin-4-yl)-1H-imidazol-2-
yl]-piperidine-
1-carboxylic acid benzyl ester;
[0056] 4-[4-(2-cyclopropyl-pyrimidin-4-yl)-5-quinoxalin-6-yl-lH-imidazol-2-yl]-
piperidine-l-
carboxylic acid benzyl ester;
[0057] 6-[5-(2-cyclopropyl-pyrimidin-4-yl)-2-(1-methanesulfonyl-piperidin-4-
yl)-3H-imidazol-
4-yl]-quinoxaline;
[0058] 4-[5-(2-methyl-pyrimidin-4-yl)-4-[1,2,4]triazolo[4,3-a]pyridin-6-yl-lH-
imidazol-2-yl]-
bicyclo[2.2.2]octan-1-ol;
[0059] 4-[4-(2-methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo[1,5-a]pyridin-6-yl-lH-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid amide;
[0060] 4-[4-(2-methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo[1,5-a]pyridin-6-yl-lH-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid;
[0061] 4-[4-(2-methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo[1,5-a]pyridin-6-yl-lH-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester;
[0062] 4-[4-(2-methyl-pyrimidin-4-yl)-5-quinoxalin-6-yl-lH-imidazol-2-yl]-
cyclohexanol; or
[0063] 4-[4-(2-methyl-pyrimidin-4-yl)-5 -quinoxalin-6-y1-1 H-imidazol-2-yl] -
bicyclo [2.2.2] octan-
1-ol.
[0064] In another aspect, a pharmaceutical composition includes a compound of
formula (I) and
a pharmaceutically acceptable carrier.
[0065] In another aspect, a method of inhibiting the TGF,6 signaling pathway
in a subject
includes administering to said subject an effective amount of a compound of
formula (I).
[0066] In another aspect, a method of inhibiting the TGF,6 type I receptor in
a cell, includes
contacting said cell with an effective amount of a compound of formula (I).
[0067] In another aspect, a method of reducing the accumulation of excess
extracellular matrix
induced by TGF,6 in a subject includes administering to said subject an
effective amount of a
compound of formula (I).
[0068] In another aspect, a method of treating or preventing fibrotic
condition in a subject
includes administering to said subject an effective amount of a compound of
formula (I). The
fibrotic condition can be, for example, scleroderma, lupus nephritis,
connective tissue disease,
wound healing, surgical scarring, spinal cord injury, CNS scarring, acute lung
injury, pulmonary
fibrosis (such as idiopathic pulmonary fibrosis), chronic obstructive
pulmonary disease, adult
respiratory distress syndrome, drug-induced lung injury, glomerulonephritis,
diabetic
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nephropathy, hypertension-induced nephropathy, alimentary track or
gastrointestinal fibrosis,
renal fibrosis, hepatic or biliary fibrosis (such as liver cirrhosis, primary
biliary cirrhosis, fatty
liver disease, primary sclerosing cholangitis), restenosis, cardiac fibrosis,
opthalmic scarring,
fibrosclerosis, fibrotic cancers, fibroids, fibroma, fibroadenomas,
fibrosarcomas, transplant
arteriopathy, or keloid. The fibrotic condition can be idiopathic in nature,
genetically linked, or
induced by radiation.
[0069] In another aspect, a method of inhibiting growth or metastasis of tumor
cells and/or
cancers in a subject includes administering to said subject an effective
amount of a compound of
formula (I).
[0070] In another aspect, a method of treating a disease or disorder mediated
by an
overexpression of TGF,6 includes administering to a subject in need of such
treatment an
effective amount of a compound of formula (I). The disease or disorder can be,
for example,
demyelination of neurons in multiple sclerosis, Alzheimer's disease, cerebral
angiopathy,
squamous cell carcinomas, multiple myeloma, melanoma, glioma, glioblastomas,
leukemia,
sarcomas, leiomyomas, mesothelioma, or carcinomas of the lung, breast, ovary,
cervix, liver,
biliary tract, gastrointestinal tract, pancreas, prostate, and head and neck.
[0071] An N-oxide derivative or a pharmaceutically acceptable salt of each of
the compounds of
formula (I) is also within the scope of this invention. For example, a
nitrogen ring atom of the
imidazole core ring or a nitrogen-containing heterocyclyl substituent can form
an oxide in the
presence of a suitable oxidizing agent such as m-chloroperbenzoic acid or
H202.
[0072] A compound of formula (I) that is acidic in nature (e.g., having a
carboxyl or phenolic
hydroxyl group) can form a pharmaceutically acceptable salt such as a sodium,
potassium,
calcium, or gold salt. Also within the scope of the invention are salts formed
with
pharmaceutically acceptable amines such as ammonia, alkyl amines,
hydroxyalkylamines, and
N-methylglycamine. A compound of formula (I) can be treated with an acid to
form acid
addition salts. Examples of such acids include hydrochloric acid, hydrobromic
acid, hydroiodic
acid, sulfuric acid, methanesulfonic acid, phosphoric acid, p-bromophenyl-
sulfonic acid,
carbonic acid, succinic acid, citric acid, benzoic acid, oxalic acid, malonic
acid, salicylic acid,
malic acid, fumaric acid, ascorbic acid, maleic acid, acetic acid, and other
mineral and organic
acids well known to those skilled in the art. The acid addition salts can be
prepared by treating a
compound of formula (I) in its free base form with a sufficient amount of an
acid (e.g.,
hydrochloric acid) to produce an acid addition salt (e.g., a hydrochloride
salt). The acid addition
salt can be converted back to its free base fotm by treating the salt with a
suitable dilute aqueous
basic solution (e.g., sodium hydroxide, sodium bicarbonate, potassium
carbonate, or ammonia).
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Compounds of formula (1) can also be, e.g., in a form of achiral compounds,
racemic mixtures,
optically active compounds, pure diastereomers, or a mixture of diastereomers.
[0073] Compounds of formula (I) exhibit surprisingly high affinity to the
TGF,6 family type I
receptors, Alk 5 and/or Alk 4, e.g., with ICSo and K; values of less than 10
M under conditions
as described below in Examples 34 and 36, respectively. Some compounds of
formula (I)
exhibit IC50 and K; values of less than 1 [tM (such as below 50 nM).
[0074] Compounds of formula (I) can also be modified by appending appropriate
functionalities
to enhance selective biological properties. Such modifications are known in
the art and include
those that increase biological penetration into a given biological system
(e.g., blood, lymphatic
system, central nervous system), increase oral availability, increase
solubility to allow
administration by injection, alter metabolism, and/or alter rate of excretion.
Examples of these
modifications include, but are not limited to, esterification with
polyethylene glycols,
derivatization with pivolates or fatty acid substituents, conversion to
carbamates, hydroxylation
of aromatic rings, and heteroatom-substitution in aromatic rings.
[0075] The present invention also features a pharmaceutical composition
comprising a
compound of formula (I) (or a combination of two or more compounds of formula
(I)) and at
least one pharmaceutically acceptable carrier. Also included in the present
invention is a
medicament composition including any of the compounds of formula (I), alone or
in a
combination, together with a suitable excipient.
[0076] The invention also features a method of inhibiting the TGF(3 family
type I receptors, Alk
and/or Alk 4 (e.g., with an IC50 value of less than 10 M; such as, less than
1 M; and for
example, less than 5 nM) in a cell, including the step of contacting the cell
with an effective
amount of one or more compounds of formula (I). Also within the scope of the
invention is a
method of inihibiting the TGF,(3 and/or activin signaling pathway in a cell or
in a subject (e.g., a
mammal such as a human), including the step of contacting the cell with or
administering to the
subject an effective amount of one or more of the compounds of formula (I).
[0077] Also within the scope of the present invention is a method of treating
a subject or
preventing a subject from suffering a condition characterized by or resulted
from an elevated
level of TGFO and/or activin activity. The method includes the step of
administering to the
subject an effective amount of one or more of a compound of formula (I). The
conditions
include an accumulation of excess extracellular matrix; a fibrotic condition
(which can be
induced by drug or radiation), e.g., scleroderma, lupus nephritis, connective
tissue disease,
wound healing, surgical scarring, spinal cord injury, CNS scarring, acute lung
injury, pulmonary
fibrosis (such as idiopathic pulmonary fibrosis and radiation-induced
pulmonary fibrosis),
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chronic obstructive pulmonary disease, adult respiratory distress syndrome,
acute lung injury,
drug-induced lung injury, glomerulonephritis, diabetic nephropathy,
hypertension-induced
nephropathy, alimentary track or gastrointestinal fibrosis, renal fibrosis,
hepatic or biliary
fibrosis, liver cirrhosis, primary biliary cirrhosis, cirrhosis due to fatty
liver disease (alcoholic
and nonalcoholic steatosis), primary sclerosing cholangitis, restenosis,
cardiac fibrosis,
opthalmic scarring, fibrosclerosis, fibrotic cancers, fibroids, fibroma,
fibroadenomas,
fibrosarcomas, transplant arteriopathy, and keloid); TGF(.3-induced growth or
metastasis of
tumor/cancer cells; and carcinomas (e.g, squamous cell carcinomas, multiple
myeloma,
melanoma, glioma, glioblastomas, leukemia, sarcomas, leiomyomas, mesothelioma,
and
carcinomas of the lung, breast, ovary, cervix, liver, biliary tract,
gastrointestinal tract, pancreas,
prostate, and head and neck); and other conditions such as cachexia,
hypertension, ankylosing
spondylitis, demyelination in multiple sclerosis, cerebral angiopathy and
Alzheimer's disease.
[0078] As used herein, an "alkyl" group refers to a saturated aliphatic
hydrocarbon group
containing 1-8 (e.g., 1-6 or 1-4) carbon atoms. An alkyl group can be straight
or branched.
Examples of an alkyl group include, but are not limited to, methyl, ethyl,
propyl, isopropyl,
butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl, and 2-ethylhexyl.
An alkyl group can be
optionally substituted with one or more substituents such as alkoxy,
cycloalkyloxy,
heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy,
amino, nitro,
carboxy, cyano, halo, hydroxy, sulfo, mercapto, alkylsulfanyl, alkylsulfinyl,
alkylsulfonyl,
aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, cycloalkyl-
alkylcarbonylamino,
arylcarbonylamino, aralkylcarbonylamino, heterocycloalkyl-carbonylamino,
heterocycloalkyl-
alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, urea,
thiourea,
sulfamoyl, sulfamide, alkoxycarbonyl, or alkylcarbonyloxy.
[0079] As used herein, an "alkenyl" group refers to an aliphatic carbon group
that contains 2-8
(e.g., 2-6 or 2-4) carbon atoms and at least one double bond. Like an alkyl
group, an alkenyl
group can be straight or branched. Examples of an alkenyl group include, but
are not limited to,
allyl, isoprenyl, 2-butenyl, and 2-hexenyl. An alkenyl group can be optionally
substituted with
one or more substituents such as alkoxy, cycloalkyloxy, heterocycloalkyloxy,
aryloxy,
heteroaryloxy, aralkyloxy, heteroarylalkoxy, amino, nitro, carboxy, cyano,
halo, hydroxy, sulfo,
mercapto, alkylsulfanyl, alkylsulfinyl, alkylsulfonyl, aminocarbonyl,
alkylcarbonylamino,
cycloalkylcarbonylamino, cycloalkyl-alkylcarbonylamino, arylcarbonylamino,
aralkylcarbonylamino, heterocycloalkyl-carbonylamino, heterocycloalkyl-
alkylcarbonylamino,
heteroarylcarbonylamino, heteroaralkylcarbonylamino, urea, thiourea,
sulfamoyl, sulfamide,
alkoxycarbonyl, or alkylcarbonyloxy.
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[0080] As used herein, an "alkynyl" group refers to an aliphatic carbon group
that contains 2-8
(e.g., 2-6 or 2-4) carbon atoms and has at least one triple bond. An alkynyl
group can be straight
or branched. Examples of an alkynyl group include, but are not limited to,
propargyl and
butynyl. An alkynyl group can be optionally substituted with one or more
substituents such as
alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy,
aralkyloxy,
heteroarylalkoxy, amino, nitro, carboxy, cyano, halo, hydroxy, sulfo,
mercapto, alkylsulfanyl,
alkylsulfinyl, alkylsulfonyl, aminocarbonyl, alkylcarbonylamino,
cycloalkylcarbonylamino,
cycloalkyl-alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino,
heterocycloalkyl-
carbonylamino, heterocycloalkyl-alkylcarbonylamino, heteroarylcarbonylamino,
heteroaralkylcarbonylamino, urea, thiourea, sulfamoyl, sulfamide,
alkoxycarbonyl, or
alkylcarbonyloxy.
[0081] As used herein, an "amino" group refers to NRxRY wherein each of Rx and
RY is
independently hydrogen, alkyl, cycloalkyl, (cycloalkyl)alkyl, aryl, aralkyl,
heterocycloalkyl,
(heterocycloalkyl)alkyl, heteroaryl, or heteroaralkyl. When the term "amino"
is not the terminal
group (e.g., alkylcarbonylamino), it is represented by -NRx-. Rx has the same
meaning as
defined above.
[0082] As used herein, an "aryl" group refers to phenyl, naphthyl, or a
benzofused group having
2 to 3 rings. For example, a benzofused group includes phenyl fused with one
or two C4_8
carbocyclic moieties, e.g., 1, 2, 3, 4-tetrahydronaphthyl, indanyl, or
fluorenyl. An aryl is
optionally substituted with one or more substituents such as alkyl (including
carboxyalkyl,
hydroxyalkyl, and haloalkyl such as trifluoromethyl), alkenyl, alkynyl,
cycloalkyl,
(cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl, aryl,
heteroaryl, alkoxy,
cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,
heteroaralkyloxy, aroyl,
heteroaroyl, amino, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy,
aminocarbonyl,
alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino,
arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino,
(heterocycloalkyl)alkylcarbonylamino, heteroarylcarbonylamino,
heteroaralkylcarbonylamino,
cyano, halo, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,
sulfamoyl,
sulfamide, oxo, or carbamoyl.
[0083] As used herein, an "aralkyl" group refers to an alkyl group (e.g., a C1-
4 alkyl group) that
is substituted with an aryl group. Both "alkyl" and "aryl" have been defined
above. An
example of an aralkyl group is benzyl.
[0084] As used herein, a "cycloalkyl" group refers to an aliphatic carbocyclic
ring of 3-10 (e.g.,
4-8) carbon atoms. Examples of cycloalkyl groups include cyclopropyl,
cyclopentyl,
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cyclohexyl, cycloheptyl, adamantyl, norbornyl, cubyl, octahydro-indenyl,
decahydro-naphthyl,
bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[3.3.1]nonyl, and
bicyclo[3.2.3]nonyl. A
"cycloalkenyl" group, as used herein, refers to a non-aromatic carbocyclic
ring of 3-10 (e.g., 4-
8) carbon atoms having one or more double bond. Examples of cycloalkenyl
groups include
cyclopentenyl, 1,4-cyclohexa-di-enyl, cycloheptenyl, cyclooctenyl, hexahydro-
indenyl,
octahydro-naphthyl, bicyclo[2.2.2]octenyl, and bicyclo[3.3.1]nonenyl. A
cycloalkyl or
cycloalkenyl group can be optionally substituted with one or more substituents
such as alkyl
(including carboxyalkyl, hydroxyalkyl, and haloalkyl such as trifluoromethyl),
alkenyl, alkynyl,
cycloalkyl, (cycloalkyl)alkyl, heterocycloalkyl, (heterocycloalkyl)alkyl,
aryl, heteroaryl, alkoxy,
cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy,
heteroaralkyloxy, aroyl,
heteroaroyl, amino, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy,
aminocarbonyl,
alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino,
arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl)carbonylamino,
(heterocycloalkyl)alkylcarbonylamino, heteroarylcarbonylamino,
heteroaralkylcarbonylamino,
cyano, halo, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea,
sulfamoyl,
sulfamide, oxo, or carbamoyl.
[0085] As used herein, a "heterocycloalkyl" group refers to a 3- to 10-
membered (e.g., 4- to 8-
membered) saturated ring structure, in which one or more of the ring atoms is
a heteroatom, e.g.,
N, 0, or S. Examples of a heterocycloalkyl group include piperidinyl,
piperazinyl,
tetrahydropyranyl, tetrahydrofuryl, dioxolanyl, oxazolidinyl, isooxazolidinyl,
morpholinyl,
octahydro-benzofuryl, octahydro-chromenyl, octahydro-thiochromenyl, octahydro-
indolyl,
octahydro-pyrindinyl, decahydro-quinolinyl, octahydro-benzo[b]thiophenyl, 2-
oxa-
bicyclo[2.2.2]octyl, 1-aza-bicyclo[2.2.2]octyl, 3-aza-bicyclo[3.2.1]octyl,
anad 2,6-dioxa-
tricyclo[3.3.1.03'7]nonyl. A "heterocycloalkenyl" group, as used herein,
refers to a 3- to 10-
membered (e.g., 4- to 8-membered) non-aromatic ring structure having one or
more double
bonds, and wherein one or more of the ring atoms is a heteroatom, e.g., N, 0,
or S. A
heterocycloalkyl or heterocycloalkenyl group can be optionally substituted
with one or more
substituents such as alkyl (including carboxyalkyl, hydroxyalkyl, and
haloalkyl such as
trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl,
heterocycloalkyl,
(heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy,
heterocycloalkyloxy, aryloxy,
heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro,
carboxy,
alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino,
cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino, arylcarbonylamino,
aralkylcarbonylamino, (heterocycloalkyl)carbonylamino,
(heterocycloalkyl)alkylcarbonylamino,
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heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo, hydroxy,
acyl, mercapto,
alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or
carbamoyl.
[0086] A "heteroaryl" group, as used herein, refers to a monocyclic, bicyclic,
or tricyclic ring
structure having 5 to 15 ring atoms wherein one or more of the ring atoms is a
heteroatom, e.g.,
N, 0, or S and wherein one ore more rings of the bicyclic or tricyclic ring
structure is aromatic.
Some examples of heteroaryl are pyridyl, furyl, pyrrolyl, thienyl, thiazolyl,
oxazolyl, imidazolyl,
indolyl, tetrazolyl, benzofuryl, benzthiazolyl, xanthene, thioxanthene,
phenothiazine,
dihydroindole, and benzo[1,3]dioxole. A heteroaryl is optionally substituted
with one or more
substituents such as alkyl (including carboxyalkyl, hydroxyalkyl, and
haloalkyl such as
trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl)alkyl,
heterocycloalkyl,
(heterocycloalkyl)alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy,
heterocycloalkyloxy, aryloxy,
heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro,
carboxy,
alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino,
cycloalkylcarbonylamino, (cycloalkyl)alkylcarbonylamino, arylcarbonylamino,
aralkylcarbonylamino, (heterocycloalkyl)carbonylamino,
(heterocycloalkyl)alkylcarbonylamino,
heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halo, hydroxy,
acyl, mercapto,
alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or
carbamoyl. A
"heteroaralkyl" group, as used herein, refers to an alkyl group (e.g., a C1.4
alkyl group) that is
substituted with a heteroaryl group. Both "alkyl" and "heteroaryl" have been
defined above.
[0087] As used herein, "cyclic moiety" includes cycloalkyl, heterocycloalkyl,
cycloalkenyl,
heterocycloalkenyl, aryl, or heteroaryl, each of which has been defined
previously.
[0088] As used herein, an "acyl" group refers to a formyl group or alkyl-C(=0)-
where "alkyl"
has been defined previously. Acetyl and pivaloyl are examples of acyl groups.
[0089] As used herein, a "carbamoyl" group refers to a group having the
structure -O-CO-
NR"RY or -NR"-CO-O-RZ wherein R" and R'' have been defined above and W can be
alkyl, aryl,
aralkyl, heterocycloalkyl, heteroaryl, or heteroaralkyl.
[0090] As used herein, a "carboxy" and a "sulfo" group refer to -COOH and -
SO3H,
respectively.
[0091] As used herein, an "alkoxy" group refers to an alkyl-0- group where
"alkyl" has been
defined previously.
[0092] As used herein, a "sulfoxy" group refers to -O-SO-Rx or -SO-O-Rx, where
Rx has been
defined above.
[0093] As used herein, a"halogen" or "halo" group refers to fluorine,
chlorine, bromine or
iodine.
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[0094] As used herein, a"sulfamoyP' group refers to the structure -S(O)2-NR"Ry
or -NR" -
S(O)2-RZ wherein R", R}", and RZ have been defined above.
[0095] As used herein, a "sulfamide" group refers to the structure -NRx -S(O)Z-
NRYRZ wherein
Rx, RY, and RZ have been defined above.
[0096] As used herein, a "urea" group refers to the structure -NRx-CO-NRYRZ
and a "thiourea"
group refers to the structure -NRx-CS-NRYRz. Rx, RY, and RZ have been defined
above.
[0097] As used herein, an effective amount is defined as the amount required
to confer a
therapeutic effect on the treated patient, and is typically determined based
on age, surface area,
weight, and condition of the patient. The interrelationship of dosages for
animals and humans
(based on milligrams per meter squared of body surface) is described by
Freireich et al., Cancer
Chemot.laer. Rep., 50: 219 (1966). Body surface area may be approximately
determined from
height and weight of the patient. See, e.g., Scientific Tables, Geigy
Pharmaceuticals, Ardsley,
New York, 537 (1970). As used herein, "patient" refers to a mammal, including
a human.
[0098] An antagonist, as used herein, is a molecule that binds to the receptor
without activating
the receptor. It competes with the endogenous ligand(s) or substrate(s) for
binding site(s) on the
receptor and, thus inhibits the ability of the receptor to transduce an
intracellular signal in
response to endogenous ligand binding.
[0099] As compounds of formula (I) are antagonists of TGFO receptor type
I(AlkS) and/or
acfivin receptor type I(Alk4), these compounds are useful in inhibiting the
consequences of
TGFO and/or activin signal transduction such as the production of
extracellular matrix (e.g.,
collagen and fibronectin), the differentiation of stromal cells to
myofibroblasts, and the
stimulation of and migration of inflammatory cells. Thus, compounds of formula
(I) inhibit
pathological inflammatory and fibrotic responses and possess the therapeutic
utility of treating
and/or preventing disorders or diseases for which reduction of TGFO and/or
activin activity is
desirable (e.g., various types of fibrosis or progressive cancers). In
addition, the compounds of
formula (I) are useful for studying and researching the role of TGFO receptor
type I(AlkS)
and/or activin receptor type I(Alk4), such as their role in cellular
processes, for example, signal
transduction, production of extracellular matrix, the differentiation of
stromal cells to
myofibroblasts, and the stimulation of and migration of inflammatory cells.
[0100] Unless otherwise defined, all technical and scientific terms used
herein have the same
meaning as commonly understood by one of ordinary skill in the art to which
this invention
belongs. All publications, patent applications, patents, and other references
mentioned herein
are incorporated by reference in their entirety. In addition, the materials,
methods, and examples
are illustrative only and not intended to be limiting.
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[0101] Other features and advantages of the invention will be apparent from
the following
detailed description, and from the claims.
DETAILED DESCRIPTION OF THE INVENTION
[0102] In general, the invention features compounds of formula (I), which
exhibit surprisingly
high affinitiy for the TGFO family type I receptors, Alk 5 and/or Alk 4.
Synthesis of the Compounds of formula (I)
[0103] Compounds of formula (I) may be prepared by a number of known methods
from
commercially available or known starting materials. In one method, compounds
of formula (I)
are prepared according to Schemes 1a, lb, or 1c below. Specifically, in Scheme
la, optionally
substituted 2-methylpyrimidine (II) is deprotonated by LDA before reacting
with Rl-substituted
carboxylic acid methoxy-methyl-amide (V) to form an Rl-(6-methylpyrimidinyl)-
ketone (III).
R' has been defined above. The methoxy-methyl-amide can be prepared by
reacting a
corresponding acid chloride (i.e., R'-CO-Cl) with N, O-dimethylhydroxylamine
hydrochloride.
The Rl-(6-methylpyrimidinyl)-ketone (III) can then be treated with sodium
nitrite in acetic acid
to afford an a-keto-oxime (IV), which can undergo further reaction with an
appropriate
substituted (and optionally protected) aldehyde (VI) in the presence of
ammonium acetate to
yield a compound of formula (I).
Scheme la
(Ra
r Ra~
1. LDA R1~- m Na
NN CH3
II II 1
2. Rl ~N, OCHg 0 NN HOAc
(II) O
(V) (III)
1. NHqOAc
OII
NOH a HxX-Y-R2 R1
I ~R 1 ( al N
R I m (VI) \R /m ~ ~X-Y-R2
y 0 NN 2. TiCl3, MeOH or H
P(OMe)3 or P(OEt)3 NN
(IV) (I)
[0104] In another method, the above-described compounds of formula (I) can be
prepared
according to Scheme lb below. Specifically, 1,1-dimethoxy-propan-2-one can
first react with
dimethoxymethyl-dimethyl-amine at an elevated temperature to produce the
intermediate 4-
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dimethylamino-1,1-dimethoxy-but-3-en-2-one, which can then react with an Ra-
substituted
amidine to form an Ra-substituted pyrimidine-2-carbaldehyde (IIa). This
carbaldehyde (Ila) can
then reacted with aniline and diphenyl phosphite to form a resulting N,P-
acetal, which can
further couple with an Rl-substituted aldehyde to produced an (Rl-methyl)-
pyrimidinyl-ketone
(IIIa). See, e.g., Journet et al., Tetrahedron Lett. 39:1717-1720 (1998).
Treatment of the (Rl-
methyl)-pyrimidinyl-ketone (Illa) with sodium nitrite in acetic acid produces
an a-keto-oxime
(IVa), which can undergo reaction with an appropriate substituted (and
optionally protected)
aldehyde (VI) to yield a compound of formula (I) as described in Scheme la
above.
Scheme lb
NH
O NYO neat, 80 C RaK NH HBr/H2O
+ i RaN ~
~O o/n IOI O~
O1- ~OPh R'
P~OPh 1. Rl-CHO
CHO PhNH2 I~ NH Cs2CO3 O
N N N N b (Ph0)2P(O)H 2. HCI
Ra Ra Ra
(Ila) (Illa)
1. NH4OAc
1
R N, 0
/
OH H X-Y-R2 R' N
NaNOZ
I ~ O (VI) ~ }--X-Y-R2
HOAc/THF/HaO NYN H
2. TiCl3, MeOH or N N
Ra P(OMe)3 or P(OEt)3
I (I)
(IVa) Ra
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[0 . 105] In another method, the above-described compounds of formula (I) can
be prepared
according to Scheme 1c below. Specifically, an (Rl-methyl)-pyrimidinyl-ketone
(IIIa)
(described above) can be oxidized to form a pyrimidinyl-diketone (IVb), which
can undergo
reaction with an appropriate substituted (and optionally protected) aldehyde
(VI) to yield a
compound of formula (I) as described above.
Scheme lc
N ) N 0
(Ra)m ~'N Rt NBS/DMSO (Ra)m ' t
N R
O O
(Illa) (IVb)
1. NHqOAc
OII
HxX-Y-R ~
z (Ra)n~~ N N
(VI)
~}--X-Y-Rz
2. TiCl3, MeOH or R1 N
P(OMe)3 or P(OEt)3 H
(I)
[0106] If compound (VI) is in its protected form, appropriate deprotecting
agents can be applied
to the resulting compound after the coupling reaction of compound (IV) or
(IVa) and compound
(VI) to yield a compound of formula (I). See, e.g., T. W. Greene, Protective
Groups in Organic
Synthesis, John Wiley & Sons, Inc., New York (198 1), for suitable protecting
groups.
[0107] Alternatively, a compound of formula (I) can be prepared by reacting
intermediate (IV)
or (IVa) with an aldehyde (VII) to yield a further intermediate (VIII), which
can then react with
compound (IX) to yield a compound of formula (I). Note that moieties Y' and Y"
are precursors
of moiety Y. See Scheme 2 below. In addition, desired substitutions at Ra can
be obtained by
selecting, for example, the appropriate compound (IIa) intermediate.
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Scheme 2
N~N 0
(IV) (Ra)m
R1
NOH 1. NH4OAc
or 0
R1
x N
H X-Y' (Ra ~~--~ Y'
(IVa) (Ra)m \ N NOH (VII) 14 H
1
R 2. TiCl3, MeOH N,,,~,,N
O
or (VIII)
N
(IVb) (Ra)m ~
0
R1 Y"~ R2
1
0 (IX) (Ra m R I N~X-Y-R2
, \ H
N,,~,,, N
(I)
[0108] In some embodiments, moiety X in compound (VII) is a nitrogen-
containing
heterocycloalkyl (e.g., piperidine). The nitrogen ring atom can be protected
by a nitrogen
protecting group (e.g., Cbz, Boc, or FMOC) before coupling to compound (IV) or
(IVa) and
deprotected afterwards (see first step of Scheme 3) to yield compound (VIIIa).
This compound
can further react with various compounds (IX) to produce a compound of formula
(I). See
second steps of Scheme 3 below. It should be noted that compound (VIII) or
compound (VIIIa)
can be a compound of formula (I) as well.
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Scheme 3
N~N 0
(IV) (Ra)m \ I ' 1. NH40Ac
R
NOH 0
or -~
1 H N-Cbz
)i_3 a Ri
\\~~ N o_i
N ~ NOH (yll) ( \ Deprotection (using
R \m Y ~~-( N-Cbz
~-(h ~ s agents,e.g.,H2, Pd/C
(IVa) (Ra)m ~ \Y N
R' 2. TiCl3, MeOH NvN H ortiBr/HOAc)
O
or (VIII)
\
(lVb) (Ra)m N O R' O R2SO2CI (IX) (Ra \ R~~~ N) o-~
I DIEA /\ ~ N-SO~RZ
( \ H 1-3
N,,~,, N
(I)
RaOCOCI (IX) (R R' N ) o-~ O
aJ,~ Y -l(
NaHCO3 \~ ~ \ N O-Rz
r' \YI H )i-3
N,,,N
(I)
R' N
(Ra 1
\~N
1-3 H NH R2NC0 (IX) ~R R1 N )o-t ,,O'R
1~
N \ 2
N~N DIEA H t 3 H z
(Villa) N,,~,,N
(I)
R1
R2COCI (IX) or (R
RZ al N ) o-~ 0
' \ N-~
COH (IX), DIEA or I 1-3 R2
R2COOH (IX), N~ N H
coupling agent, e.g., ~
HATU (I)
R'
RZCHO (IX) (Ra 1 N ) o-i
NaB(OAc)3H /\ I ~N~ z
~H/13 R
N,,,, N
(I)
[0109] Similarly, when moiety X in compound (VII) is a cycloalkyl (e.g.,
cyclopentyl,
cyclohexyl, or bicyclo[2.2.2]octane), it can be further functionalized to form
a compound of
formula (I) as depicted in Schemes 4, 5a, 5b, and 5c below.
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Scheme 4
N'- N 0
(IV) (Ra)m 1
R 1. NH4OAc
NOH
t N o-, NHCbz
0 NHCbz r ~ RI
or ~ )o-t (R'~m
)1-3
H Deprotection (using
N__'_N NOH H (VII) N ),.,
(IVa) (Ra)m-\ I NvN agents,e.g.,H2,Pd/C
RI 2. TiC13, MeOH (VIII) orHBr/HOAc)
0
or /Ra Rt N o-~ NHSO2R2
N\ 0 R~SO2CI (IX) ai
(IVb) (~''a)m ' / t DIEA H )~'3
- II R N N
O
(I)
0
R2OCOCI (IX) ~ Ra m R1 N o-~ oN--~ 2
O-R
NaHCO3 N )1-a
~
N,,:,, N
(I)
R'
Ra N a-, NH2 RI I..I~O
m
~\ I N ),a R2NCO (IX) (Ra lm I N a~ N NR2
H DIEA /\ \ H~ H
NI~/ N (I
(Villa) N,,'~,, N
(I)
R2COCI (IX) or Rt N 0-1 N40
R2COH (IX), DIEA or ~ R Rz
R COOH (IX), coupling ~iV H )1-3
agent, e.g., HATU N,,Z~,- IN
(I)
R2CHO (IX) (Ra Rt N o-~ N-\NaB(OAc)3H ~ 0-1 R~
II~H )7-3
NvN
(I)
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Scheme 5a
N~N 0
(IV) (Ra)m \ YR1
NOH 1. NH4OAc
or 0
H~COOMe R1 N
N~N NOH (VII) ( lRa ~ 'Z~~COOMe
(A) (IVa) (Ra)m -\ ~ \ \ H
R1 2. TiCl3, MeOH N~N
0
or (I)
\
(IVb) (RB)m N O R1
O rRa,1 R1 N j~COOH
\ m\ ~~(' \~-
Deprotection \ ~/
(using reagents, N ~ N H
e.g., LiOH)
(I)
(can be further modified
according to Scheme 5b
below)
N~ R
(IV) (Ra)m i 1
NOH 1. NH4OAc
or 0
H-1-&OH
Ra R1
(VII) ~ J N
N N NOH m\ ~OH
(B) (IVa) (Ra)m 1 \ N
R 2. TiCi3, MeOH NI~N H
0
or (I)
N \ 0
(IVb) (Ra)m ' N~ ~Rl
0 (RaR1 N
NH(Rb)SOzCI \\ N~~OSOaNH(Rb)
II
N~N H
(I)
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Scheme 5b
~ Ra) Rt N~ /~\ (Ra ~ Rt N 0
\ ?-E'\~-COOH RZOH m
m
~
(~) \ H \\ H OR2
N:~, N
(I) (~)
( R a ~ R ~ t N ~ (Ra ~ Rt N 0
)---~~-COOH RZNHa m
(2) \ \ H ~~// fl \\ H~ NHRz
N~N NvN
(~) (I)
a Rt /~ / a R
3 ~R\ll\>__--CooH m RzSOaNH(Rb) lR ~ t N
~// N NHSOZR2
N~N EDC,DMAP \ ~H
N~N
(I) (i)
(Ra
\ Rt N
~'COOH 1. HATU, NH3 (Ra ) Rt N' ~--~
H 2. 2reducing agents
~I I ~%~-(~~~/ -~.
(4) NvN (e.g., BH3=THF) I \ H NHz
N,,N
(i)
(~)
(can be further modified
according to Scheme 5c
below)
(Ra / Ri N/~ Rt
\m\~ ~~-f' \~-COOH diohenvlohosohorvlazide (Ra ~ N~
14 ~/ DIEA, benzyl alcohol, toluene \\ ~ H NH O
(5) N~ vN H
N~N O~-
(I)
(~)
H2, Pd/C or (Ra ) Rt N~ /~\
l \rIm }-(~\k NHa
HBr/HOAc \ H ~-/
NvN
(I)
(can be further modified
according to Scheme 5c
below)
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(Ra ) R' NBH3=THF (Ra ~ R' N OH 2
COOH R SO,CI
DIEA, THF
(s \\ ) H THF \ H
NvN NvN
(1) (1)
(Ra ~ R~ (R ) ~ N
\ N NaCN a R
m I ~
( ~O-~oR2 DMF \ N CN
N~N H O ~ N,,,, N H
aN3
LiCI / NHaCI HCI / H2O
(Ra ) R~ N
\~ \ r 1 (Ra ~ R1 N
H
'N \NCOOH
NN HN-N (I)
/Ra ' R~ N (Ra ) R~' N /~ Trifluoroactic
' I\ ~~--( \~-COOH HATU, NH3 \ \ ~)- (~~CONH2 anhydride
(7) N ~/ DMF ~\ H"N ~// Pyridine, THF
N,,,, N H N,,:~,, N
(1) (1)
a R~
( R m\ ~ N C N NaN3 (Ra ~m R1 I N N-N
\H LiCl / NH4CI \~ NN N
N~iN N ~ N H H
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Scheme 5c
/~ ooNHSO2R2
RZSOZCI (IX) (Ra ) R~ N
DIEA ~~--f\i}-F~
H
N,,,,N
(I)
O
R2OCOCI (IX) (Ra ) Ri N' N4 a
m O
NaHCO3
~ H
NN
(1)
(Ra ) R' N NHa O
I N~ ~o R2NCO (IX) (Ra ) F''1 N N~ 2
r~ H DIEA \\ H-R
N~N r % H
(~) N,,Z~,, N
(~)
R2COCI (IX) or Rt HO
R2COH (IX), DIEA or (Ra ~m I N~a R2
R2COOH (IX), coup ing N
agent, e.g., HATU H
NN (')
R1 H
R2CHO (IX) (Ra ) NN- \ z
m R
NaB(OAc)3H N
~ H
N,,::~, N
(~)
[01101 As is well known to a skilled person in chemistry, desired
substitutions can be placed on
the pyrimidinyl ring in the last steps of the synthesis. See Scheme 6 below,
for example.
Scheme 6
sl~ O, S%O HN14~
N)IIN N'I" N NJ\\N H
N H202, NaWO4HZO / HZN-a / N
~} -X-Y-R2 ~ --X-Y-Rz , X-Y-R Z
R~ N HZSO4, MeOH R' N Reflux R' N
(I) (I) (I)
[0111] Compounds of formula (I) wherein Rb is not hydrogen can be prepared by
known
methods. For example, compounds of formula (I) wherein Al is N and A2 is NH
(or vice versa)
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WO 2006/026306 PCT/US2005/030133
can be treated with R1(e.g., alkyl iodide) and CsCO3 to produce a compound of
formula (I)
wherein Rb is alkyl. See, e.g., Liverton, et al., J. Med. Chem., 42: 2180-2190
(1999).
[0112] As will be obvious to a skilled person in the art, some starting
materials and
intermediates may need to be protected before undergoing synthetic steps as
described above.
For suitable protecting groups, see, e.g., T. W. Greene, Protective Groups in
Organic Synthesis,
John Wiley & Sons, Inc., New York (1981).
Uses of Compounds of formula (I)
[0113] As discussed above, hyperactivity of the TGFO family signaling pathways
can result in
excess deposition of extracellular matrix and increased inflammatory
responses, which can then
lead to fibrosis in tissues and organs (e.g., lung, kidney, and liver) and
ultimately result in organ
failure. See, e.g., Border, W.A. and Ruoslahti E. J. Clin. Invest. 90: 1-7
(1992) and Border,
W.A. and Noble, N.A. N. Engl. J. Med. 331: 1286-1292 (1994). Studies have been
shown that
the expression of TGFO and/or activin mRNA and the level of TGFO and/or
activin are increased
in patients suffering from various fibrotic disorders, e.g., fibrotic kidney
diseases, alcohol-
induced and autoimmune hepatic fibrosis, myelofibrosis, bleomycin-induced
pulmonary fibrosis,
and idiopathic pulmonary fibrosis. Elevated TGF(3 and/or activin is has also
been demonstrated
in cachexia, demyelination of neurons in multiple sclerosis, Alzheimer's
disease, cerebral
angiopathy and hypertension.
[0114] Compounds of formula (I), which are antagonists of the TGFO family type
I receptors
Alk 5 and/or Alk 4, and inhibit TGFO and/or activin signaling pathway, are
therefore useful for
treating and/or preventing fibrotic disorders or diseases mediated by an
increased level of TGFO
and/or activin activity. As used herein, a compound inhibits the TGFO family
signaling pathway
when it binds (e.g., with an IC50 value of less than 10 M; such as, less than
1 M; and for
example, less than 5 nM) to a receptor of the pathway (e.g., Alk 5 and/or Alk
4), thereby
competing with the endogenous ligand(s) or substrate(s) for binding site(s) on
the receptor and
reducing the ability of the receptor to transduce an intracellular signal in
response to the
endogenous ligand or substrate binding. The aforementioned disorders or
diseases include any
condition (a) marked by the presence of an abnormally high level of TGFO
and/or activin; and/or
(b) an excess accumulation of extracellular matrix; and/or (c) an increased
number and synthetic
activity of myofibroblasts. These disorders or diseases include, but are not
limited to, fibrotic
conditions such as scleroderma, glomerulonephritis, diabetic nephropathy,
lupus nephritis,
hypertension-induced nephropathy, ocular or comeal scarring, alimentary track
or
gastrointestinal fibrosis, renal fibrosis, hepatic or biliary fibrosis, acute
lung injury, pulmonary
fibrosis (such as idiopathic pulmonary fibrosis and radiation-induced
pulmonary fibrosis), post-
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infarction cardiac fibrosis, fibrosclerosis, fibrotic cancers, fibroids,
fibroma, fibroadenomas, and
fibrosarcomas. Other fibrotic conditions for which preventive treatment with
compounds of
formula (I) can have therapeutic utility include radiation-induced fibrosis,
chemotherapy-
induced fibrosis, and surgically-induced scarring including surgical
adhesions, laminectomy,
and coronary restenosis.
[0115] Increased TGFO activity is also found to manifest in patients with
progressive cancers.
Studies have shown that in many cancers, the tumor cells, stromal cells,
and/or other cells within
a tumor generally overexpress TGF,6. This leads to stimulation of angiogenesis
and cell
motility, suppression of the immune system, and/or increased interaction of
tumor cells with the
extracellular matrix. See, e.g., Hojo, M. et al., Nature 397: 530-534 (1999)
and Lammerts E. et
al., Int. J. Cancer 102: 453-462 (2002). As a result, the tumors grow more
readily, become
more invasive and metastasize to distant organs. See, e.g., Maehara, Y. et
al., J. Clin. Oncol.
17: 607-614 (1999) and Picon, A. et al., Cancet Epidemiol. Bi markers Prev.
7: 497-504 (1998).
Thus, compounds of formula (I), which are antagonists of the TGFO type I
receptor and inhibit
TGFO signaling pathways, are also useful for treating and/or preventing
various cancers which
overexpress TGFO or benefit from TGFO's above-mentioned pro-tumor activities.
Such cancers
include carcinomas of the lung, breast, liver, biliary tract, gastrointestinal
tract, head and neck,
pancreas, prostate, cervix as well as multiple myeloma, melanoma, glioma, and
glioblastomas.
[0116] Importantly, it should be pointed out that because of the chronic, and
in some cases
localized, nature of disorders or diseases mediated by overexpression of TGFO
and/or activin
(e.g., fibrosis or cancers), small molecule treatments (such as treatment
disclosed in the present
invention) are favored for long-term treatment.
[0117] Not only are compounds of formula (I) useful in treating disorders or
diseases mediated
by high levels of TGFO and/or activin activity, these compounds can also be
used to prevent the
same disorders or diseases. It is known that polymorphisms leading to
increased TGFO and/or
activin production have been associated with fibrosis and hypertension.
Indeed, high serum
TGFO levels are correlated with the development of fibrosis in patients with
breast cancer who
have received radiation therapy, chronic graft-versus-host-disease, idiopathic
interstitial
pneumonitis, veno-occlusive disease in transplant recipients, and peritoneal
fibrosis in patients
undergoing continuous ambulatory peritoneal dialysis. Thus, the levels of TGFO
and/or activin
in serum and of TGFO and/or activin mRNA in tissue can be measured and used as
diagnostic or
prognostic markers for disorders or diseases mediated by overexpression of
TGFO and/or
activin, and polymorphisms in the gene for TGFO that determine the production
of TGFO and/or
activin can also be used in predicting susceptibility to disorders or
diseases. See, e.g., Blobe,
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G.C. et al., N. Engl. J. Med. 342(18): 1350-1358 (2000); Matsuse, T. et al.,
Am. J. Respir. Cell
Mol. Biol. 13: 17-24 (1995); Inoue, S. et al., Biochem. Biophys. Res. Comm.
205: 441-448
(1994); Matsuse, T. et al, Am. J Pathol. 148: 707-713 (1996); De Bleser et
al., Hepatology 26:
905-912 (1997); Pawlowski, J.E., et al., J Clin. Invest. 100: 639-648 (1997);
and Sugiyama, M.
et al., Gastroenterology 114: 550-558 (1998).
ADMINISTRATION OF COMPOUNDS OF FORMULA (I)
[0118] As defined above, an effective amount is the amount required to confer
a therapeutic
effect on the treated patient. For a compound of formula (I), an effective
amount can range, for
example, from about 1 mg/kg to about 150 mg/kg (e.g., from about 1 mg/kg to
about 100
mg/kg). Effective doses will also vary, as recognized by those skilled in the
art, dependant on
route of administration, excipient usage, and the possibility of co-usage with
other therapeutic
treatments including use of other therapeutic agents and/or radiation therapy.
[0119] Compounds of formula (I) can be administered in any manner suitable for
the
administration of pharmaceutical compounds, including, but not limited to,
pills, tablets,
capsules, aerosols, suppositories, liquid formulations for ingestion or
injection or for use as eye
or ear drops, dietary supplements, and topical preparations. The
pharmaceutically acceptable
compositions include aqueous solutions of the active agent, in an isotonic
saline, 5% glucose or
other well-known pharmaceutically acceptable excipient. Solubilizing agents
such as
cyclodextrins, or other solubilizing agents well-known to those familiar with
the art, can be
utilized as pharmaceutical excipients for delivery of the therapeutic
compounds. As to route of
administration, the compositions can be administered orally, intranasally,
transdermally,
intradermally, vaginally, intraaurally, intraocularly, buccally, rectally,
transmucosally, or via
inhalation, implantation (e.g., surgically), or intravenous administration.
The compositions can
be administered to an animal (e.g., a mammal such as a human, non-human
primate, horse, dog,
cow, pig, sheep, goat, cat, mouse, rat, guinea pig, rabbit, hamster, gerbil,
or ferret, or a bird, or a
reptile, such as a lizard).
[0120] Optionally, compounds of formula (I) can be administered in conjunction
with one or
more other agents that inhibit the TGF(3 signaling pathway or treat the
corresponding
pathological disorders (e.g., fibrosis or progressive cancers) by way of a
different mechanism of
action. Examples of these agents include angiotensin converting enzyme
inhibitors, nonsteroid
and steroid anti-inflammatory agents, immunotherapeutics, chemotherapeutics,
as well as agents
that antagonize ligand binding or activation of the TGFO receptors, e.g., anti-
TGFO, anti-TGF,(3
receptor antibodies, or antagonists of the TGFO type II receptors. Compounds
of formula (I) can
also be administered in conjunction with other treatments, e.g., radiation.
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The invention will be iurther described in the following examples, which do
not limit the
scope of the invention described in the claims.
Example 1
4-[1-Hydroxy-4-(2-methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo [1,5-a] pyridin-6-
yl-1H-
imidazol-2-yl]-bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
Synthesis of the title compound is described in parts (a)-(b) below.
(a) 1-(2-Methyl-pyrimidin-4-yl)-2-[1,2,4]triazolo[1,5-a]pyridin-6-yl-ethane-
1,2-dione 2-
oxime (IVa)
[0121] Sodium nitrite (1.03 g, 15 mmol) was added to a solution of 1-(2-methyl-
pyrimidin-4-
yl)-2-[1,2,4]triazolo[1,5-a]pyridin-6-yl-ethanone (prepared according to
Scheme lb above) (2.5
g, 10 mmol) in a nzixture of HOAc/THF/HZO (6:4:1, 55 mL). The mixture was
stirred at 0 C
for 1 hour and then room temperature for 1 hour. Solvent was removed under
reduced pressure.
Residue was dissolved in water and NaOH (3N) was added until the pH was
greater than 8. The
aqueous solution was extracted with ethyl acetate. The organic layer was
washed with brine,
dried over sodium sulfate, filtered, and concentrated to give 1.8 g (64%) of
the title compound as
a yellow foam.
(b) 4-[1-Hydroxy-4-(2-methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo[1,5-a]pyridin-6-
yl-1H-
imidazol-2-yl]-bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
[0122] 4-Formyl-bicyclo[2.2.2]octane-1-carboxylic acid methyl ester (0.170 g,
1.0 mmol) was
added to a solution of 1-(2-methyl-pyrimidin-4-yl)-2-[1,2,4]triazolo[1,5-
a]pyridin-6-yl-ethane-
1,2-dione 2-oxime (0.282 g, 1 mmol) and ammonium acetate (1.54 g, 20 mmol) in
acetic acid (5
mL). The mixture was refluxed for 3 hours. Solvent was removed under reduced
pressure. The
reaction mixture was then quenched with an ammonia/ice mixture. The aqueous
solution was
extracted with ethyl acetate. The organic layer was washed with brine, dried
over sodium
sulfate, filtered, and concentrated to give 0.400g (87%) of the title compound
as a yellow solid.
Example 2
4-[4-(2-Methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo [1,5-a] pyridin-6-yl-1H-
imidazol-2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
[0123] Triethylphosphite (0.343 uL, 2.0 mmol) was added to a solution of 4-[1-
hydroxy-4-(2-
methyl-pyrimidin-4-yl)-5-[ 1,2,4]triazolo[ 1,5-a]pyridin-6-yl-1 H-irnidazol-2-
yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester (0.40 g, 0.87 mmol; see
Example 1 above)
in DMF (10 mL). The mixture was heated at 110 C for 18 hours. The solvent was
removed.
The residue was portioned between ethyl acetate and brine. The organic layer
was dried over
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sodium sulfate, filtered, and concentrated to give a yellow oil. HPLC
purification gave 0.30 g
(77%) of the title compound as a yellow oil. 1H NMR (300 MHz, Methanol-d4) S
9.32 (s, 1H),
8.69 (d, 1H, J = 5.7 Hz), 8.58 (s, 1H), 7.93 (m, 2H), 7.74 (d, 1H, J = 5.7
Hz), 3.68 (s, 3H), 2.65
(s, 3H), 2.15 (m, 6H), 1.99 (m, 6H). MS (ES+) m/z (M+l) 444.24.
Example 3
4-[4-(2-Methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo [1,5-a]pyridin-6-yl-lH-
imidazol-2-yl]-
bicyclo [2.2.2] octane-l-carboxylic acid
[0124] Lithium hydroxide monohydrate (0.046 g, 1.12 mmol) was added to a
solution of 4-[4-
(2-methyl-pyrimidin-4-yl)-5-[ 1,2,4]triazolo[ 1,5-a]pyridin-6-yl-1 H-imidazol-
2-yl]-
bicyclo[2.2.2]octane-1-carboxylic acid methyl ester (0.25 g, 0.56 mmol) in a
mixture of
THF/MeOH/H20 (2/1/1, 4 mL). The mixture was stirred for 3 hours, and the
solvent was
removed. The residue was diluted with water (30 mL). Citric acid was added to
the solution to
make the pH lower than 7. The aqueous solution was extracted with ethyl
acetate. The organic
layer was washed with brine, dried over sodium sulfate, filtered and
concentrated to give 0.180 g
(75%) of the title compound as a yellow solid. 1H NMR (300 MHz, Methanol-d4) 6
9.29 (s,
1H), 8.65 (d, 1H, J= 5.4 Hz), 8.54 (s, 1H), 7.91 (m, 2H), 7.67 (d, 1H, J = 5.7
Hz), 2.64 (s, 3H),
2.14 (m, 6H), 2.00 (m, 6H). MS (ES+) m/z (M+1) 430.28.
Example 4
4-[4-(2-Methyl-pyrimidin-4-yl)-5-[1,2,4]triazolo [1,5-a]pyridin-6-yl-lH-
imidazol-2-yl]-
bicyclo [2.2.2] octane-l-carboxylic acid amide
[0125] HATU (0.265 g, 0.70 mmol) was added to a solution of 4-[4-(2-methyl-
pyrimidin-4-yl)-
5-[1,2,4]triazolo[1,5-a]pyridin-6-yl-lH-imidazol-2-yl]-bicyclo[2.2.2]octane-l-
carboxylic acid
(0.150 g, 0.35 mmol) and potassium carbonate (0.1242 g, 1.75 mmol) in DMF (5
mL). The
mixture was stirred for 10 minutes. NH3 was bubbled into the reaction mixture
for 10 minutes.
The mixture was stirre for an additional2 hours. The mixture was filtered, and
DMF was
removed under reduced pressure. The residue was dissolved in DMSO and the DMSO
solution
was filtered. HPLC purification of the DMSO solution gave 0.040 g (27%) of the
title
compound as a yellow solid. 1H NMR (300 MHz, Methanol-d4) S 9.30 (s, 1H), 8.67
(d, 1H, J
6.0 Hz), 8.56 (s, 1H), 7.92 (m, 2H), 7.69 (d, 1H, J = 6.0 Hz), 2.65 (s, 3H),
2.15 (m, 6H), 1.99 (m,
6H). MS (ES+) m/z (M+1) 429.25.
Example 5
4-[4-(2-Methyl-pyrimidin-4-y1)-5-quinoxalin-6-yl-lH-imidazol-2-yl]-bicyclo
[2.2.2] octan-l-
ol
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Synthesis of the title compound is described in parts (a)-(b) below.
(a) 1-(2-Methyl-pyrimidin-4-yl)-2-quinoxalin-6-yl-ethane-1,2-dione (IVb)
[0126] To a solution of 2-(2-methyl-pyrimidin-4-yl)-1-quinoxalin-6-yl-ethanone
(0.500 g, 1.9
mmol; prepared according to Scheme lb above)) in DMSO (5 mL) was added NBS
(0.337 g, 1.9
mmol) and then stirred at room temperature for 3 days. The mixture was
partitioned between
ether and water. Ether was washed with brine, dried over sodium sulfate,
filtered and
concentrated to give 0.200 g (38%) of 1-(2-methyl-pyrimidin-4-yl)-2-quinoxalin-
6-yl-ethane-
1,2-dione as a yellow solid.
(b) 4-[4-(2-Methyl-pyrimidin-4-y1)-5-quinoxalin-6-y1-1H-imidazol-2-y1]-
bicyclo[2.2.2]octan-
1-ol
[0127] 4-Hydroxy-bicyclo [2.2.2] octane- 1 -carbaldehyde (0.130 g, 0.86 mmol)
was added to a
solution of 1-(2-methyl-pyrimidin-4-yl)-2-quinoxalin-6-yl-ethane-l,2-dione
(0.200 g, 0.72
mmol) and ammonium acetate (0.554 g, 7.2 mmol) in acetic acid (10 mL). The
mixture was
reflux for 3 hours. Solvent was removed under reduced pressure. Reaction
mixture was then
quenched with ammonia/ice mixture. The aqueous solution was extracted with
ethyl acetate.
Ethyl acetate was washed with brine, dried over sodium sulfate, filtered, and
concentrated.
HPLC purification eluting with acetonitrile:water gave 0.06 g (20%) of the
title compound as a
yellow solid. 1H NMR (300 MHz, Methanol-d4) S 8.99 (m, 2H), 8.63 (d, 1H, J=
5.8 Hz), 8.43
(d, 1H, J = 1.8 Hz), 8.25 (d, 1H, J = 8.7 Hz), 8.03 (m, 1H), 7.47 (d, 1H, J =
5.7 Hz), 2.30 (m,
6H), 1.87 (m, 6H). MS (ES) m/z (M+l) 413.28.
Example 6
4-[1-Hydroxy-4-(6-methyl-pyridin-2-yl)-5-(2-methylsulfanyl-pyrimidin-4-yl)-1H-
imidazol-
2-yl]-bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
Synthesis of the title compound is described in parts (a)-(b) below.
(a) 1-(6-Methyl-pyridin-2-y1)-2-(4-methylsulfanyl-pyrimidin-2-y1)-ethane-1,2-
dione 2-
oxime (IV)
[0128] Sodium nitrite (0.479 g, 6.9 mmol) was added to a solution of 1-(6-
methyl-pyridin-2-yl)-
2-(4-methylsulfanyl-pyrimidin-2-yl)-ethanone (1.2 g, 4.6 mmol; prepared
according to Scheme
1 above) in a mixture of HOAc/THF/H20 (6:4:1, 35 mL). The mixture was stirred
at 0 C for 1
hour and then at room temperature for another hour. Solvent was removed under
reduced
pressure. Residue was dissolved in water, to which NaOH (3N) was added until
pH was larger
than 8. The aqueous solution was extracted with ethyl acetate. The organic
layer was washed
with brine, dried over sodium sulfate, filtered, and concentrated to give 1.3
g (98%) of the title
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oxime as a yellow foam.
(b) 4-[1-Hydroxy-4-(6-methyl-pyridin-2-yl)-5-(2-methylsulfanyl-pyrimidin-4-yl)-
1H-
imidazol-2-yl]-bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
[0129] 4-Formyl-bicyclo[2.2.2]octane-l-earboxylic acid methyl ester (0.72 g,
2.5 mmol) was
added to a solution of 1-(6-methyl-pyridin-2-yl)-2-(4-methylsulfanyl-pyrimidin-
2-yl)-ethane-
1,2-dione 2-oxime (0.6 g, 2.1 mmol) and ammonium acetate (3.1 g, 40 mmol) in
acetic acid (30
mL). The mixture was reflux for 2 hours. Solvent was removed under reduced
pressure.
Reaction mixture was then quenched with ammonia/ice mixture. The aqueous
solution was
extracted with ethyl acetate. Ethyl acetate was washed with brine, dried over
sodium sulfate,
filtered, and concentrated to 0.9 g (92%) of the title methyl ester as a
yellow solid.
Example 7
4-[4-(6-Methyl-pyridin-2-yl)-5-(2-methylsulfanyl-pyrimidin-4-yl)-1H-imidazol-2-
yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
[0130] Trimethylphosphite (1.0 mL, 9.7 mmol) was added to a solution of 4-[1-
hydroxy-4-(6-
methyl-pyridin-2-yl)-5-(2-methylsulfanyl-pyrimidin-4-yl)-1 H-imidazol-2-yl]-
bicyclo[2.2.2]octane-1-carboxylic acid methyl ester (0.9 g, 1.93 mmol; see
Example 6 above) in
DMF (10 mL). The mixture was stirred at 110 C for 18 hours. Solvent was
removed to give a
yellow oil. Column chromatography eluting with ethyl acetate:hexanes (50:50)
gave 0.8 g
(97%) of the title compound as a yellow solid. 1H NMR (300 MHz, DMSO-d6) S
12.25 (s, 1H),
8.51 (m, 1H), 7.71 (m, 2H), 7.59 (d, 1H, J= 6.0 Hz), 7.22 (t, 1H, J = 3.0 Hz),
3.61 (s, 3H), 3.29
(s, 3H), 2.11 (s, 3H), 1.98 (m, 6H), 1.83 (m, 6H). MS (ES+) m/z (M+l) 450.15.
Example 8
4-[5-(2-Methanesulfonyl-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1 H-imidazol-
2-yl]-
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
[0131] Hydrogen peroxide (0.19 mL,6.68 mmol), 4N H2S04 (0.06 mL), and NaWO4HZO
(10
mg) were added to a solution of 4-[4-(6-methyl-pyridin-2-yl)-5-(2-
methylsulfanyl-pyrimidin-4-
yl)-1H-imidazol-2-yl]-bicyclo[2.2.2]octane-1-carboxylic acid methyl ester
(0.60 g, 1.34 mmol;
see Example 7 above) in methonal (20 mL). The mixture was stirred at 50 C for
10 hours. The
mixture was then quenched with water and stirred for 30 minutes. Saturated
Na2SZO3 aqueous
solution was added to neutralize the excess hydrogen peroxide. The aqueous
solution was
extracted with ethyl acetate. Ethyl acetate was washed with brine, dried over
sodium sulfate,
filtered, and concentrated. Column chromatography eluting with ethyl acetate
gave 0.5 g (91%)
of the title compound as a yellow solid. 1H NMR (300 MHz, Methanol-d4) 8 8.98
(m, 1H), 8.43
(m, 1H), 8.22 (m, 1H), 8.09 (m, 1H), 7.84 (m, 1H), 3.68 (s, 3H), 3.26 (s, 3H),
2.86 (s, 3H), 2.11
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(m, 6h), 1.99 (m, 6H). MS (ES+) m/z (M+1) 482.02.
Example 9
4-[5-(2-Methanesulfonyl-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-imidazol-
2-yl] -
bicyclo[2.2.2]octane-l-carboxylic acid methyl ester
[0132] 4-[5-(2-Methanesulfonyl-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-
imidazol-2-yl]-
bicyclo[2.2.2]octane-1-carboxylic acid methyl ester (0.30 g, 0.62 mmol; see
Example 8 above)
was dissolved in cyclopropylamine (10 mL) and the mixture was placed in a
sealed tube. The
mixture was heated at 80 C for 18 hours. Solvent was removed to give 0.285 g
(99%) of the
title compound as a yellow solid. 1H NMR (300 MHz, Acetone-d6) 6 8.56 (m, 1H),
8.49 (m,
1H), 8.24 (m, 1H), 7.65 (m, 2H), 3.63 (s, 3H), 2.87 (m, 1H), 2.61 (s, 3H),
2.05 (m, 6H), 1.91 (m,
6H), 0.75 (m, 2H), 0.64 (m, 2H). MS (ES+) m/z (M+1) 459.17.
Example 10
4-[5-(2-Cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-imidazol-
2-yl] -
bicyclo [2.2.2] octane-l-carboxylic acid
[0133] Lithium hydroxide monohydrate (0.10 g, 2.44 mmol) was added to 4-[5-(2-
Cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)- l H-imidazol-2-yl]-
bicyclo[2.2.2]octane-1-carboxylic acid methyl ester (0.28 g, 0.61 mmol) in a
mixture of
THF/MeOH/HZO (2/1/1, 5 mL). The mixture was stirred for 3 h. Solvent was
removed.
Residue was diluted with water (30 mL). Citric acid was added to the solution
to make the pH
lower than 7. The aqueous solution was extracted with ethyl acetate. Ethyl
acetate was washed
with brine, dried over sodium sulfate, filtered and concentrated to give 0.27
g (99%) of the title
compound as a yellow solid. 'H NMR (300 MHz, Methanol-d4) 8 8.50 (m, 1H), 8.29
(m, 2H),
7.66 (m, 1H), 7.30 (m, 1H), 2.75 (m, 1H), 2.65 (s, 3H), 2.07 (m, 6H), 1.97 (m,
6H), 0.86 (m,
2H), 0.62 (m, 2H). MS (ES+) m/z (M+l) 445.10.
Example 11
4-[5-(2-Cyclopropylamino-pyrimidin-4-yl)-4-(6-methyl-pyridin-2-yl)-1H-imidazol-
2-yl]-
bicyclo [2.2.2] octane-l-carboxylic acid amide
[0134] HATU (0.17 g, 0.45 mmol) was added to a solution of 4-[5-(2-
Cyclopropylamino-
pyrimidin-4-yl) -4-(6-methyl-pyridin-2-yl)-1 H-imidazol-2-yl] -bicyclo [2.2.2]
octanc-l-c arboxylic
acid (0.10 g, 0.225 mmol; see Example 10 above) and potassium carbonate (0.155
g, 1.13 mmol)
in anhydrous DMF (5 mL). The mixture was stirred for 30 minutes. Ammonia was
bubbled
through the mixture for 10 minutes. The mixture was continued to stir for 2
hours. The mixture
was then filtered and concentrated. HPLC purification eluting with
acetonitrile:water gave
0.070 g (70%) of the title compound as a yellow solid. 1H NMR (300 MHz,
Methanol-d4) S
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8.50 (d, 1H, J = 5.7 Hz), 8.30 (m, 2H), 7.67 (m, 1H), 7.32 (d, 1H, J= 5.7 Hz),
2.75 (m, 1H), 2.65
(s, 3H), 2.08 (m, 6H), 1.95 (m, 6H), 0.84 (m, 2H), 0.61 (m, 2H). MS (ES+) m/z
(M+l) 444.21.
[0135] The compounds listed in the following Table were prepared in an
analogous manner to
those described in the methods and examples above.
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Example Chemical Name 1H-NMR MS Method
(ES+)
m/z
(M+l )
Ex. 12 4-[4-(2-Methyl-pyrimidin-4-yl)- (300 MHz, Methanol-d4) d 9.00
5-quinoxalin-6-yl-lH-imidazol- (m, 2H), 8.62 (d, 1H, J = 5.4 Hz),
2-yl]-cyclohexanol 8.46 (d, 1H, J = 1.2 Hz), 8.28 (d,
1H, J = 8.7 Hz), 8.06 (m, 1H), 387.1 Ex. 1
7.37 (m, 1H), 3.68 (m, 1H), 3.31
(m, 1H), 2.66 (s, 3H), 2.22 (m,
2H), 1.92 (m, 4H), 1.51 (m, 2H)
Ex. 13 4-[5-Quinoxalin-6-yl-4-(2- (300 MHz, Methanol-d4) d 8.97
trifluoromethyl-pyrimidin-4-yl)- (m, 2H), 8.87 (d, 1H, J= 6.0 Hz),
1 H-imidazol-2-yl]- 8.37 (d, 1H, J = 3.0 Hz), 8.20 (d,
bicyclo[2.2.2]octan-l-ol 1H, J = 9.0 Hz), 8.06 (m, 1H), 467.35 Ex. 5
7.86 (d, 1H, J = 6.0 Hz), 2.26 (m,
6H), 1.85 (m, 6H)
Ex. 14 4-[4-(2-Cyclopropyl-pyrimidin- (300 MHz, Methanol-d4) d 8.99
4-yl)-5-quinoxalin-6-yl-1H- (m, 2H), 8.58 (d, 1H, J= 6.0 Hz),
imidazol-2-yl]- 8.34 (d, 1H, J = 3.0 Hz), 8.25 (d,
bicyclo[2.2.2]octan-l-ol 1H, J = 9.0 Hz), 7.98 (m, 1H), 439.41 Ex. 5
7.59 (d, 1H, J= 6.0 Hz), 2.25 (m,
6H), 2.14 (m, 1H), 1.85 (m, 6H),
0.94 (m, 2H), 0.76 (m, 2H)
Ex. 15 6-[2-tert-Butyl-5-(2- (300 MHz, Methanol-d4) d 9.00
cyclopropyl-pyrimidin-4-yl)- (m, 2H), 8.60 (d, 1H, J = 6.0 Hz),
3H-imidazol-4-yl]-quinoxaline 8.38 (d, 1H, J = 3.0 Hz), 8.26 (d,
1H, J = 9.0 Hz), 8.00 (m, 1H), 371.35 Ex. 5
7.59 (d, 1H, J = 6.0 Hz), 2.14 (m,
1H), 1.61 (s, 9H), 0.94 (m, 2H),
0.77 (m, 2H)
Ex. 16 6-[5-(2-Cyclopropyl-pyrimidin- (300 MHz, Methanol-d4) d 9.00
4-yl)-3H-imidazol-4-yl]- (m, 2H), 8.80 (s, 1H), 8.58 (d, 1H,
quinoxaline J = 6.0 Hz), 8.39 (d, 1H, J = 1.5
Hz), 8.28 (d, 1H, J = 9.0 Hz), 8.05 315.18 Ex. 5
(m, 1 H), 7.5 3 (d, 1 H, J= 6.0 Hz),
2.19 (m, 1H), 1.01 (m, 2H), 0.93
(m, 2H)
Ex. 17 {4-[4-(2-Cyclopropyl- (300 MHz, Methanol-d4) d 9.00
pyrimidin-4-yl)-5-quinoxalin-6- (m, 2H), 8.58 (d, 1H, J= 3.0 Hz),
yl-lH-imidazol-2-yl]- 8.35 (d, 1H, J = 3.0 Hz), 8.25 (d,
bicyclo[2.2.2]oct-1-yl}- 1H, J = 9.0 Hz), 7.99 (m, 1H), 453.4 Ex. 5
methanol 7.49 (d, 1H, J = 6.0 Hz), 3.59 (s,
2H), 2.13 (m, 6H), 1.69 (m, 1 H),
1.64 (m, 6H), 0.95 (m, 2H), 0.79
(m, 2H)
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Ex. 18 6-[5-(2-Trifluoromethyl- (300 MHz, Methanol-d4) d 8.97
pyrimidin-4-yl)-3H-imidazol-4- (m, 2H), 8.88 (d, 1H, J = 6.0 Hz),
yl]-quinoxaline 8.61 (s, 1H), 8.42 (d, 1H, J= 1.5 343.15 Ex. 5
Hz), 8.23 (d, 1 H, J = 9.0 Hz), 8.13
(m, 1 H), 7.88 (d, 1 H, J = 6.0Hz),
2.65 (s, 3H)
Ex. 19 6-[2-tert-Butyl-5-(2- (300 MHz, Methanol-d4) d 8.99
trifluoromethyl-pyrimidin-4-yl)- (m, 2H), 8.91 (d, 1H, J = 6.0 Hz),
3H-imidazol-4-yl]-quinoxaline 8.42 (d, 1H, J = 3.0 Hz), 8.24 (d, 399.3 Ex. 5
1H, J =9.0 Hz), 8.06 (m, 1H), 7.84
(d, 1H, J = 6.0 Hz), 1.58 (s, 9H)
Ex. 20 4-[5-Quinoxalin-6-yl-4-(2- (300 MHz, Methanol-d4) d 8.98
trifluoromethyl-pyrimidin-4-yl)- (m, 2H), 8.86 (d, 1H, J = 6.0 Hz),
1H-imidazol-2-yl]-piperidine-l- 8.44 (d, 1H, J = 1.5 Hz), 8.26 (d,
carboxylic acid benzyl ester 1H, J = 9.0 Hz), 8.06 (d, 1H, J = 560.26 Ex. 5
1.5 Hz), 7.84 (d, 1H, J = 6.0 Hz),
7.35 (m, 5H), 5.13 (s, 2H), 4.37
(m, 1 H), 4.05 (m, 1 H), 3.33 (m,
1H), 3.00 (m, 2H), 2.13 (m, 4H)
Ex. 21 4-[4-(2-Cyclopropyl-pyrimidin- (300 MHz, Methanol-d4) d 8.81
4-yl)-5-quinoxalin-6-yl-1H- (m, 2H), 8.45 (d, 1H, J = 6.0 Hz),
imidazol-2-yl]-piperidine-l- 8.25 (d, 1H, J = 1.5 Hz), 8.11 (d,
carboxylic acid benzyl ester 1H, J = 9.0 Hz), 7.83 (d, 1H, J =
9.0 Hz), 7.56 (d, 1H, J= 6.0 Hz), 532.38 Ex. 5
7.20 (m, 5H), 4.96 (s, 2H), 4.19
(m, 2H), 3.3 8(m, 1H), 2.82 (m,
2H), 1.91 (m, 5H), 0.92 (m, 2H),
0.79 (m, 2H)
Ex. 22 6-[5-(2-Cyclopropyl-pyrimidin- (300 MHz, Methanol-d4) d 8.99
4-yl)-2-(1-methanesulfonyl- (s, 2H), 8.57 (d, 1H, J = 5.7 Hz),
piperidin-4-yl)-3H-imidazol-4- 8.37 (d, 1H, J = 1.8 Hz), 8.26 (d,
yl]-quinoxaline 1H, J = 8.7 Hz), 8.03 (dd, 1H, J =
2.1 Hz, 8.7 Hz), 7.58 (d, 1H, J = 476.47 Ex. 5
5.7 Hz), 3.93 (m, 2H), 3.21 (m,
3H), 2.65 (s, 31-1), 2.10 (m, 5H),
0.98 (m, 2H), 0.86 (m, 2H)
Ex. 23 4-[5-(2-Methyl-pyrimidin-4-yl)- (300 MHz, Methanol-d4) d 9.28
4-[1,2,4]triazolo[4,3-a]pyridin- (d, 1H, J = 3.0 Hz), 8.65 (d, 1H, J
6-yl-lH-imidazol-2-yl]- = 6.0 Hz), 8.54 (s, 1H), 7.92 (m, 402.46 Ex. 5
bicyclo[2.2.2]octan-l-ol 2H), 7.65 (d, 1H, J = 6.0 Hz), 2.65
(s, 3H), 2.23 (m, 6H), 1.84 (m,
6H)
Ex. 24 4-[5-(2-Methoxy-pyrimidin-4- (300 MHz, Methanol-d4) 8 8.85
yl)-4-(6-methyl-pyridin-2-yl)- (d, 1H, J = 5.1 Hz), 8.61 (m, 111),
1H-imidazol-2-yl]- 8.44 (t, 1H, J = 8.1 Hz), 7.89 (m,
bicyclo[2.2.2]octane-l- 1H), 7.77 (d, 1H, J = 7.5 Hz), 4.18 434.05 Ex.
carboxylic acid methyl ester (s, 3H), 3.68 (s, 3H), 3.03 (s, 3H), 9&10
2.10 (m, 6H), 1.98 (m, 6H)
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Ex. 25 4-[5-(2-Hydroxy-pyrimidin-4- (300 MHz, Methanol-d4) 6 8.21
yl)-4-(6-methyl-pyridin-2-yl)- (m, 3H), 7.55 (m, 1H), 7.37 (m,
1H-imidazol-2-yl]- 1H), 3.58 (s, 3H), 2.90 (s, 3H), 420.16 Ex.
bicyclo[2.2.2]octane-l- 2.00 (m, 6H), 1.87 (m, 6H) 9&10
carboxylic acid methyl ester
Ex. 26 4-[5-(2-cyclopropylamino- (300 MHz, Acetone-d6) 5 8.56 (m,
pyrimidin-4-yl)-4-(6-methyl- 1H), 8.49 (m, 1H), 8.24 (m, 1H),
pyridin-2-yl)-1H-imidazol-2- 7.65 (m, 2H), 3.63 (s, 3H), 2.87
yl]-bicyclo[2.2.2]octane-l- (m, 1H), 2.61 (s, 3H), 2.05 (m, 459.17 Ex.
carboxylic acid methyl ester 6H), 1.91 (m, 6H), 0.75 (m, 2H), 9&10
0.64 (m, 2H).
Ex. 27 4-[5-(2-cyclopropylamino- (300 MHz, Methanol-d4) 6 8.49
pyrimidin-4-yl)-4-(6-methyl- (m, 1H), 8.30 (m, 2H), 7.66 (m,
pyridin-2-yl)-1H-imidazol-2- 1H), 7.32 (m, 1H), 2.74 (m, 1H), 460.19 Ex.
yl]-bicyclo[2.2.2]octane-1- 2.65 (s, 3H), 2.07 (m, 6H), 1.93 9&10
carboxylic acid hydroxyamide (m, 6H), 0.85 (m, 2H), 0.61 (m,
2H).
Ex. 28 4-[5-(2-cyclopropylamino- (300 MHz, Methanol-d4) b 8.49
pyrimidin-4-yl)-4-(6-methyl- (m, 1H), 8.30 (m, 2H), 7.67 (m,
pyridin-2-yl)-1H-imidazol-2- 1H), 7.31 (m, 1H), 3.67 (s, 3H),
yl]-bicyclo[2.2.2]octane-l- 2.75 (m, 1H), 2.65 (s, 3H), 2.06 474.21 Ex.
carboxylic acid methoxy-amide (m, 6H), 1.92 (m, 6H), 0.84 (m, 9&10
2H), 0.62 (m, 2H).
Ex. 29 4-[5-(2-amino-pyrimidin-4-yl)- (300 MHz, Methanol-d4) 6 8.50
4-(6-methyl-pyridin-2-yl)-1H- (d, 1H, J = 5.4 Hz), 8.42 (d, 1H, J
imidazol-2-yl]- = 8.1 Hz), 8.34 (t, 1H, J= 7.8 Hz),
bicyclo[2.2.2]octane- 1 - 7.68 (d, 1H, J = 7.5 Hz), 7.30 (d, 405.11 Ex.
carboxylic acid 1H, J = 5.4 Hz), 2.90 (s, 3H), 2.08 9&10
(m, 6H), 1.98 (m, 6H).
Ex. 30 {4-[5-(2-Cyclopropylamino- (300 MHz, Methanol-d4) 6 8.14
pyrimidin-4-yl)-4-(6-methyl- (m, 1H), 7.73 (m, 2H), 7.31 (m,
pyridin-2-yl)-1H-imidazol-2- 5H), 7.06 (m, 1H), 6.70 (m, 1H),
yl]-bicyclo[2.2.2]oct-1-yl}- 5.02 (s, 2H), 2.60 (m, 1H), 2.54 (s, 550.15 Ex.
carbamic acid benzyl ester 3H), 2.10 (m, 6H), 2.00 (m, 6H), 9&10
0.79 (m, 2H), 0.45 (m, 2H)
Ex. 31 N-{4-[5-(2-cyclopropylamino- (300 MHz, Methanol-d4) S 8.47
pyrimidin-4-yl)-4-(6-methyl- (m, 1H), 8.29 (m, 2H), 7.69 (m,
pyridin-2-yl)-1H-imidazol-2- 1H), 7.35 (m, 1H), 2.80 (s, 3H), 458.24 Ex.
yl]-bicyclo[2.2.2]oct-1-yl}- 2.65 (s, 3H), 2.10 (m, 12H), 1.36 9&10
acetamide (m, 1H), 0.85 (m, 2H), 0.62 (m,
2H).
Ex. 32 N-{4-[5-(2-cyclopropylamino- (300 MHz, Methanol-d4) 8 8.50
pyrimidin-4-yl)-4-(6-methyl- (m, 1H), 8.30 (m, 1H), 7.65 (m,
pyridin-2-yl)-1H-imidazol-2- 1H), 7.49 (m, 1H), 7.29 (m, 1H),
yl]-bicyclo[2.2.2]oct-1-yl}- 2.80 (s, 3H), 2.65 (s, 3H), 2.26 (m, 494.20 Ex.
methanesulfonamide 1H), 2.10 (m, 12H), 0.84 (m, 2H), 9&10
0.61 (m, 2H).
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.. .... ..... ..
Ex. 33 N-{4-[5-(2-cyclopropylamino- (300 MHz, Methanol-d4) b 8.52
pyrimidin-4-yl)-4-(6-methyl- (d, 1H, J = 5.4 Hz), 8.31 (m, 1H),
pyridin-2-yl)-1H-imidazol-2- 7.66 (m, 111), 7.49 (m, 1H), 7.30
yl]-bicyclo[2.2.2]oct-l-yl}- (d, 111, J = 5.4 Hz), 2.65 (s, 3H), 512.08 Ex.
2,2,2-trifluoro-acetamide 2.26 (m, 1H), 2.15 (m, 12H), 0.84 9&10
(m, 2H), 0.60 (m, 2H).
[0136] The TGF,6 inhibitory activity of compounds of formula (I) can be
assessed by methods
described in the following examples.
Example 34
Cell-Free Assay for Evaluating Inhibition of Autophosphorylation of TGF# Type
I
Receptor
[0137] The serine-threonine kinase activity of TGF(.i type I receptor was
measured as the
autophosphorylation activity of the cytoplasmic domain of the receptor
containing an N-terminal
poly histidine, TEV cleavage site-tag, e.g., His-TGF,l3RI. The His-tagged
receptor cytoplasmic
kinase domains were purified from infected insect cell cultures using the
Gibco-BRL FastBac
HTb baculovirus expression system.
[0138] To a 96-well Nickel FlashPlate (NEN Life Science, Perkin Elmer) was
added 20 L of
1.25 Ci 33P-ATP/25 M ATP in assay buffer (50 mM Hepes, 60 mM NaCl, 1 mM
MgC12, 2
mM DTT, 5 mM MnC12, 2% glycerol, and 0.015% Brij 35). 10 L of each test
compound of
formula (I) prepared in 5% DMSO solution were added to the FlashPlate. The
assay was then
initiated with the addition of 20 L of assay buffer containing 12.5 pmol of
His-TGFORI to each
well. Plates were incubated for 30 minutes at room temperature and the
reactions were then
terminated by a single rinse with TBS. Radiation from each well of the plates
was read on a
TopCount (Packard). Total binding (no inhibition) was defined as counts
measured in the
presence of DMSO solution containing no test compound and non-specific binding
was defined
as counts measured in the presence of EDTA or no-kinase control.
[0139] Alternatively, the reaction performed using the above reagents and
incubation conditions
but in a microcentrifuge tube was analyzed by separation on a 4-20% SDS-PAGE
gel and the
incorporation of radiolabel into the 40 kDa His-TGF[3RI SDS-PAGE band was
quantitated on a
Storm Phosphoimager (Molecular Dynamics).
[0140] Compounds of formula (I) typically exhibited IC5o values of less than
10 M; some
exhibited IC50 values of less than 1 M; and some even exhibited IC50 values
of less than 50 nM.
Example 35
Cell-Free Assay for Evaluating Inhibition of Activin Type I Receptor Kinase
Activity
[0141] Inhibition of the Activin type I receptor (Alk 4) kinase
autophosphorylation activity by
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test compounds of formula (I) can be determined in a similar manner to that
described above in
Example 34 except that a similarly His-tagged form of Alk 4 (His-Alk 4) is
used in place of the
His-TGFRRI.
Example 36
TGF# Type I Receptor Ligand Displacement FlashPlate Assay
[0142] 50 nM of tritiated 4-(3-pyridin-2-yl-lH-pyrazol-4-yl)-quinoline (custom-
ordered from
PerkinElmer Life Science, Inc., Boston, MA) in assay buffer (50 mM Hepes, 60
mM NaC12, 1
mM MgC12, 5 mM MnC12, 2 mM 1,4-dithiothreitol (DTT), 2% Brij 35; pH 7.5) was
premixed
with a test compound of formula (I) in 1% DMSO solution in a v-bottom plate.
Control wells
containing either DMSO without any test compound or control compound in DMSO
were used.
To initiate the assay, His-TGFO Type I receptor in the same assay buffer
(Hepes, NaC12, MgC12,
MnC12, DTT, and 30% Brij added fresh) was added to a nickel coated FlashPlate
(PE, NEN
catalog number: SMP 107), while the control wells contained only buffer (i.e.,
no His-TGF,(i
Type I receptor). The premixed solution of tritiated 4-(3-pyridin-2-yl-lH-
pyrazol-4-yl)-
quinoline and test compound of formula (I) was then added to the wells. The
wells were
aspirated after an hour at room temperature and radioactivity in wells
(emitted from the tritiated
compound) was measured using TopCount (PerkinElmer Lifesciences, Inc., Boston
MA).
Compounds of formula (I) typically exhibited Ki values of less than 10 M;
some
exhibited Ki values of less than 1 M; and some even exhibited Ki values of
less than 50 nM.
Example 37
Assay for Evaluating Cellular Inhibition of TGF# Signaling and Cytotoxicity
[0143] Biological activity of the compounds of formula (I) was determined by
measuring their
ability to inhibit TGF(3-induced PAI-Luciferase reporter activity in HepG2
cells.
[0144] HepG2 cells were stably transfected with the PAI-luciferase reporter
grown in DMEM
medium containing 10% FBS, penicillin (100 U/mL), streptomycin (100 g/mL), L-
glutamine
(2 mM), sodium pyruvate (1 mM), and non-essential amino acids (lx). The
transfected cells
were then plated at a concentration of 2.5 x 104 cells/well in 96 well plates
and starved for 3-6
hours in media with 0.5% FBS at 37 C in a 5% COZ incubator. The cells were
then stimulated
with 2.5 ng/mL TGF(3ligand in the starvation media containing 1% DMSO either
in the
presence or absence of a test compound of formula (I) and incubated as
described above for 24
hours. The media was washed out the following day and the luciferase reporter
activity was
detected using the LucLite Luciferase Reporter Gene Assay kit (Packard, cat.
no. 6016911) as
recommended. The plates were read on a Wallac Microbeta plate reader, the
reading of which
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was used to determine the IC50 values of compounds of formula (I) for
inhibiting TGFO-induced
PAI-Luciferase reporter activity in HepG2 cells. Compounds of formula (I)
typically exhibited
IC50 values of less 10 M.
[0145] Cytotoxicity was determined using the same cell culture conditions as
described above.
Specifically, cell viability was determined after overnight incubation with
the CytoLite cell
viability kit (Packard, cat. no. 6016901). Compounds of formula (I) typically
exhibited LD25
values greater than 10 M.
Example 38
Assay for Evaluating Inhibition of TGFO Type I Receptor Kinase Activity in
Cells
[0146] The cellular inhibition of activin signaling activity by the test
compounds of formula (I)
is determined in a similar manner as described above in Example 37 except that
100 ng/mL of
activin is added to serum starved cells in place of the 2.5 ng/mL TGF(.3.
Example 39
Assay for TGFO-Induced Collagen Expression
Preparation of Iinmortalized Collagen Promotor-Green Fluorescent Protein Cells
[0147] Fibroblasts are derived from the skin of adult transgenic mice
expressing Green
Fluorescent Protein (GFP) under the control of the collagen 1A1 promoter (see
Krempen, K. et
al., Gene Exp. 8: 151-163 (1999)). Cells are immortalized with a temperature
sensitive large T
antigen that is in an active stage at 33 C. Cells are expanded at 33 C and
then transferred to 37
C at which temperature the large T antigen becomes inactive (see Xu, S. et
al., Exp. Cell Res.
220: 407-414 (1995)). Over the course of about 4 days and one split, the cells
cease
proliferating. Cells are then frozen in aliquots sufficient for a single 96
well plate.
Assay of TGFO-induced Collagen-GFP Expression
[0148] Cells are thawed, plated in complete DMEM (contains non-essential amino
acids, 1mM
sodium pyruvate and 2mM L-glutamine) with 10 % fetal calf serum, and then
incubated for
overnight at 37 C, 5% CO2. The cells are trypsinized in the following day and
transferred into
96 well format with 30,000 cells per well in 50 L complete DMEM containing 2
% fetal calf
serum, but without phenol red. The cells are incubated at 37 C for 3 to 4
hours to allow them to
adhere to the plate. Solutions containing a test compound of formula (I) are
then added to wells
with no TGFO (in triplicates), as well as wells with 1 ng/mL TGFO (in
triplicates). DMSO is
also added to all of the wells at a final concentration of 0.1%. GFP
fluorescence emission at 530
nm following excitation at 485 nm is measured at 48 hours after the addition
of solutions
containing a test compound on a CytoFluor microplate reader (PerSeptive
Biosystems). The
data are then expressed as the ratio of TGF(3-induced to non-induced for each
test sample.
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OTHER EMBODIMENTS
[0149] It is to be understood that while the invention has been described in
conjunction with the
detailed description thereof, the foregoing description is intended to
illustrate and not limit the
scope of the invention, which is defined by the scope of the appended claims.
Other aspects,
advantages, and modifications are within the scope of the following claims.
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