Synthesis and biological evaluation of 1-substituted-3-(6-methylpyridin-2- yl)-4-([1,2,4triazolo[1,5-apyridin-6-yl)pyrazoles as transforming growth factor-β type 1 receptor kinase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2011.08.064

A series of 1-substituted-3-(6-methylpyridin-2-yl)-4-([1,2,4triazolo[1,5- apyridin-6-yl)pyrazoles 14a-ae, 16a, 16b, and 21a-c has been prepared and evaluated for their ALK5 inhibitory activity in an enzyme assay and in a cell-based luciferase reporter ass

Full text of DOI:10.1016/j.bmcl.2011.08.064

Bioorganic & Medicinal Chemistry Letters 21 (2011) 6049–6053
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of 1-substituted-3-(6-methylpyridin-2-yl)-4-
([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles as transforming growth factor-b
type 1 receptor kinase inhibitors
Cheng Hua Jin, Maddeboina Krishnaiah, Domalapally Sreenu, Vura Bala Subrahmanyam,
Kota Sudhakar Rao, Annaji Venkata Nagendra Mohan, Chul-Yong Park, Jee-Yeon Son, Yhun Yhong Sheen,
⇑
Dae-Kee Kim
College of Pharmacy, Ewha Womans University, 11-1 Daehyun-dong, Seodaemun-gu, Seoul 120-750, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 25 May 2011
Revised 29 July 2011
Accepted 13 August 2011
Available online 22 August 2011
A series of 1-substituted-3-(6-methylpyridin-2-yl)-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles 14a–
ae, 16a, 16b, and 21a–c has been prepared and evaluated for their ALK5 inhibitory activity in an enzyme
assay and in
a cell-based luciferase reporter assay. The 4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)-
N-(4-methoxyphenyl)-3-(6-methylpyridin-2-yl)-1H-pyrazole-1-carbothioamide (14n) inhibited ALK5
phosphorylation with IC50 value of 0.57 nM and showed 94% inhibition at 100 nM in a luciferase reporter
assay using HaCaT cells permanently transfected with p3TP-luc reporter construct.
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
TGF-b
ALK5 inhibitor
Fibrosis
Cancer
Kinase assay
Cell-based luciferase reporter assay
The transforming growth factor-b (TGF-b) is the most potent and
ubiquitous profibrogenic cytokine. TGF-b transduces signals
through two distinct serine/threonine kinase receptors, termed type
I and type II.1–3 Following the binding of ligand to the constitutively
active type II receptor, the type I receptor, also called as activin
receptor-like kinase 5 (ALK5), is phosphorylated, which further
phosphorylates Smad2 or Smad3. Phosphorylated Smad2 and
Smad3 form a heteromeric complex with Smad4, which translocates
to the nucleus and binds to the promoters of TGF-b target genes in-
volved in cell differentiation, proliferation, apoptosis, migration, and
extracellular matrix production.4 TGF-b plays a pivotal role in the
development of fibrosis in various organ systems such as kidney,5
heart,6 lung,7 and liver.8 Deregulation of TGF-b signaling has been
also implicated in various human diseases including cancer,9 pan-
creatic diseases,10 and hematological malignancies.11 Recent studies
have shown that blocking the TGF-b signaling pathway with several
small molecule ATP-competitive ALK5 inhibitors (Fig. 1) such as 1
(SB-505154),12 2 (SB-525334),13 3 (GW6604),14 4 (SD-208),15,16
and 5 (LY-2157299)17 inhibited autophosphorylation of ALK5 and
TGF-b induced transcription of matrix genes in receptor assays at
sub-micromolar concentrations. Among them, 2, 3, and 4 effectively
retarded progressive fibrosis in kidney, liver and lung, respectively,
and 4 and 5 also strongly inhibited growth and invasiveness of can-
cer cells in animal models. We have prepared a number of the 2-pyr-
idyl-substituted five-membered heterocycles as ALK5 inhibitors
and found that insertion of a methylene, a methyleneamino, or an
aminomethylene linkage between a central five-membered hetero-
cyclic ring and a phenyl ring significantly increased ALK5 inhibitory
activity and selectivity.18–26 One of our preclinical candidates, IN-
1130 (6), demonstrated its remarkable activity as a suppressor of
fibrogenic process of unilateral ureteral obstruction in rats,27 ame-
liorated experimental autoimmune encephalomyelitis, a mouse
model for multiple sclerosis,28 and reduced tunical fibrosis in rats.29
It also inhibited cancer metastasis in MMTV/c-Neu breast cancer
mice and enhanced CTL response in cancer mice.30 Another preclin-
ical candidate, IN-1233 (7), effectively prevented development and
progression of pulmonary arterial hypertension in monocrotaline
rat model through inhibition of TGF-b signaling31 and reduced gran-
ulation tissue formation after bare metallic stent placement in a rat
urethral model.32
Recently, Tojo et al.33 and we23 have reported a novel class of
ALK5 inhibitors possessing a thioamide linkage between a pyrazole
ring and a phenyl ring, such as A-83-01 (8) and 3-(3-(6-methylpyri-
din-2-yl)-4-(quinolin-6-yl)-1H-pyrazol-1-carbothioamido )benzm-
ide (9).
⇑
Corresponding author. Tel.: +82 2 3277 3025; fax: +82 2 3277 2467.
E-mail address: dkkim@ewha.ac.kr (D.-K. Kim).
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.08.064

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C. H. Jin et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6049–6053
O
N
F
NH2
N
N
O
HN
N
N
N
O
N
N
N
N
N
N
N
NH
N
H
N
H
N
N
N
N
N
Cl
5 (LY-2157299)
3 (GW6604)
1 (SB-505124)
2 (SB-525234)
4 (SD-208)
N
N
N
N
O
O
S
S
N
NH2
NH2
N
N
N
N
N
HN
N
HN
N
H
N
H
N
N
N
N
NH2
O
8 (A-83-01)
9
6 (IN-1130)
7 (IN-1233)
Figure 1. ALK5 inhibitors under development.
Insertion of a thioamide linkage between a pyrazole ring and a
phenyl ring markedly increased ALK5 inhibitory activity, thus, 8
exhibited significant inhibition of the transcriptional activity in-
duced by ALK5,33 and 9 showed more than 90% inhibition at
100 nM in luciferase reporter assays using HaCaT cells transiently
transfected with p3TP-luc reporter construct and ARE-luc reporter
construct.23 As part of our continuing efforts to develop more potent
and selective ALK5 inhibitors, in this Letter, we have prepared a
series of 1-substituted-3-(6-methylpyridin-2-yl)-4-([1,2,4]triazol-
o[1,5-a]pyridin-6-yl)pyrazoles 14a–ae, 16a, 16b, and 21a–c pos-
sessing a thioamide linkage between a pyrazole ring and a phenyl
ring or a piperidinyl ring. The target molecules 14a–ae have a
[1,2,4]triazolo[1,5-a]pyridin-6-yl moiety rather than a quinoxalin-
6-yl moiety of 2 and 6 or a quinolin-6-yl moiety of 7 and 9 at the
4-position of the pyrazole and have various substituents such as F,
Cl, Me, CF3, OMe, OCF3, CN, and CONH2 in the phenyl ring. In
N
N
N
N
N
N
N
N
b
a
NH
N
N
O
CHO
N
N
10
11 (90%)
12 (80%)
SCN
N
R
N
N
H
N
N
13a-ae
N
R
c
S
N
14a-ae
a:
b:
c:
p: R = p-CN (75%)
q:
R = H
R = m-F
R = m-Cl
p: R = p-CN
q:
r: R = o-F
a:
R = H (55%)
R = p-CONH2 (46%)
r: R = o-F (24%)
s:
R = p-CONH2
b: R = m-F (50%)
c: R = m-Cl (40%)
d: R = m-Me (64%)
e: R = m-CF3 (48%)
f: R = m-OMe (50%)
R = o-Cl (53%)
s: R = o-Cl
d: R = m-Me
e: R = m-CF3
f: R = m-OMe
g: R = m-OCF3
t: R = o-Me (47%)
u: R = o-OMe (69%)
v: R = o-OCF3 (31%)
t:
R = o-Me
u: R = o-OMe
v:
R = o-OCF3
g:
R = m-OCF3 (4%)
w:
R = 3,4-F2 (38%)
x: R = 3,5-F2 (38%)
y:
w: R = 3,4-F2
x: R = 3,5-F2
y: R = 3,4-Cl2
z: R = 3,5-Cl2
aa: R = 3,4-Me2
h:
i:
j:
R = m-CN
R = m-CONH2
R = p-F
h: R = m-CN (34%)
i: R = m-CONH2 (52%)
j: R = p-F (45%)
k: R = p-Cl (52%)
l: R = p-Me (70%)
m: R = p-CF3 (46%)
R = 3,4-Cl2 (45%)
z: R = 3,5-Cl2 (22%)
k:
R = p-Cl
aa: R = 3,4-Me2 (29%)
l: R = p-Me
ab:
ab:
R = 3,5-Me2 (47%)
ac: R = 3,5-(CF3)2 (41%)
ad:
R = 3,5-Me2
ac: R = 3,5-(CF3)2
ad:
m: R = p-CF3
n: R = p-OMe
o: R = p-OCF3
n:
o:
R = 3,4-(OMe)2 (51%)
ae: R = 3,5-(OMe)2 (26%)
R = 3,4-(OMe)2
ae: R = 3,5-(OMe)2
R = p-OMe (59%)
R = p-OCF3 (39%)
Scheme 1. Reagents and conditions: (a) (i) diphenyl (6-methylpyridin-2-yl)(phenylamino)methylphosphonate, Cs2CO3, THF/i-PrOH (4:1), rt, 16 h; (ii) 3 N HCl, rt, 1 h; (b) (i)
DMFÁDMA, 90 °C, 4 h; (ii) N2H4ÁH2O, EtOH, reflux, 3 h; (c) NaH, anhydrous THF, 0 °C to rt, 1 h.

C. H. Jin et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6049–6053
6051
SCN
The 4-([1,2,4]tria zolo[1,5-a]pyridin-6-yl)-N-(1-methylpiperidin-
4-yl)-3-(6-methylpyridin-2-yl)-1H-pyrazole-1-carbothioamide
(16a) and 4-([1,2,4] triazolo[1,5-a]pyridin-6-yl)-N-(1-benzylpi-
N
N
N
R
N
H
N
15
N
peridin-4-yl)-3-(6-met
hylpyridin-2-yl)-1H-pyrazole-1-carbo-
12
N
a
thioamide (16b) were prepared as shown in Scheme 2. The
pyrazole 12 was alkylated with 4-isothiocyanato-1-methylpiperi-
dine (15a)38 or 1-benzyl-4-isothiocyanatopiperidine (15b)39 in the
same reaction condition as described in Scheme 1 to afford the
16a or 16b in 25% or 41% yields, respectively.
S
N
R
N
16
a: R = Me (25%)
b: R = Benzyl (41%)
a: R = Me
b: R = Benzyl
The 3-(6-methylpyridin-2-yl)-4-(2-methyl-[1,2,4]triazolo[1,5-
a]pyridin-6-yl)-1-(phenylcarbothioamido)pyrazoles 21a–c were
prepared as shown in Scheme 3. Formylation of 6-iodo-3-methyl-
[1,2,4]triazolo[4,3-a]pyridine (17)40 with i-PrMgCl and N,N-dimeth-
ylformamide in anhydrous THF followed by quenching with 1 N HCl
gave the Dimroth-like rearranged aldehyde 18 in 36% yield. The
aldehyde 18 was coupled with diphenyl (6-methylpyridin-2-
yl)(phenylamino)methylphosphonate to afford the monoketone 19
in 88% yield. The monoketone 19 was converted to the pyrazole 20
in 45% yield in the same reaction condition as described in Scheme
1. The pyrazole 20 was further alkylated with phenyl isothiocya-
nates (13a, 13h, and 13i) in the presence of NaH in anhydrous THF
to afford the target compounds 21a–c in 53–88% yields.
To investigate whether these compounds 14a–ae, 16a, 16b, and
21a–c could inhibit ALK5, a kinase assay was performed using the
purified human ALK5 kinase domain produced in Sf9 insect cells
(Table 1). The ALK5 inhibitory activity of the pyrazoles 14b–14ae
having various substituents such as F, Cl, Me, CF3, OMe, OCF3, CN,
and CONH2 in the phenyl ring was compared with that of the
unsubstituted pyrazole 14a. All the pyrazoles 14a–ae inhibited
ALK5 at concentrations (IC50 = 0.57–15.60 nM) as shown in Table
1. Among the meta-substituted compounds 14b–i, the m-Cl substi-
tuted compound 14c (IC50 = 2.37 nM) and m-OMe substituted
compound 14f (IC50 = 2.26 nM) displayed slightly more potent
ALK5 inhibition than the unsubstituted compound 14a
(IC50 = 2.82 nM). Among the para-substituted compounds 14j–q,
the p-OMe substituted compound 14n (IC50 = 0.57 nM) was highly
potent and was five-fold more potent than the compound 14a. The
p-CN substituted compound 14p (IC50 = 2.95 nM) showed the sim-
ilar level of potency to that of the compound 14a. Introduction of a
substituent at the ortho-positon was proved to be not beneficial,
thus, the o-Cl substituted compound 14s (IC50 = 2.83 nM) only dis-
played the comparable ALK5 inhibitory activity to that of the 14a.
Among the di-substituted compounds 14w–ae, the 3,4-F2 substi-
Scheme 2. Reagents and conditions: (a) NaH, anhydrous THF, 0 °C to rt, 1 h.
pharmacokinetic studies, the bioavailability of orally administered 6
was 84.9%, 34.4%, 11.4%, and 8.95% in dogs, monkeys, rats, and mice,
respectively.25 The major metabolite of 6 was detected in the
systemic circulation of rat and mouse and was identified as 3-((4-
(2-hydroxyquinoxalin-6-yl)-5-(6-methylpyridin-2-yl)-1H-imidazo
l-2-yl)methyl)benzamide or 3-((4-(3-hydroxyquinoxalin-6-yl)-5-
(6-methylpyridin-2-yl)-1H-imidazol-2-yl)methyl)benzamide.25 To
overcome metabolic oxidation of 2- or 3-position of quinoxalin-6-
yl moiety of 6, we adopted a [1,2,4]triazolo[1,5-a]pyridin-6-yl moi-
ety since metabolic oxidation of 2-position of this moiety is assumed
to be difficult due to the presence of two adjacent nitrogen atoms.
The target molecules 16a and 16b have been prepared to com-
pare the role of a phenyl ring of 14a–ae with a piperidinyl ring. To
examine the influence of a methyl substituent at the 2-position of
the [1,2,4]triazolo[1,5-a]pyridin-6-yl moiety on ALK5 inhibition,
we have also prepared the target molecules 21a–c.
A
series of 3-(6-methylpyridin-2-yl)-1-(phenylcarbothioa-
mido)-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles 14a–ae was
prepared as shown in Scheme 1. The [1,2,4]triazolo[1,5-a]pyri-
dine-6-carbaldehyde (10)34 was coupled with diphenyl (6-methyl-
pyridin-2-yl)(phenylamino)methylphosphonate35 in a mixture of
THF and i-PrOH (4:1) in the presence of Cs2CO3 at room temperature
and subsequently treated with 3 N HCl to afford the monoketone
1136 in 90% yield. Treatment of 11 with N,N-dimethylformamide di-
methyl acetal at 90 °C and followed by reaction with hydrazine
monohydrate in absolute EtOH produced the pyrazole 1237 in 80%
yield. The pyrazole 12 was alkylated with appropriately substituted
phenyl isothiocyanates (13a–ae) in the presence of NaH in anhy-
drous THF to afford the target compounds 14a–ae in various yields
(4–75%). Compound 14g was obtained only in 4% yield due to the
low reactivity of 3-(trifluoromethoxy)phenyl isothiocyanate (13g).
N
N
N
N
N
N
a
b
N
N
O
N
I
CHO
N
18 (36%)
17
19 (88%)
SCN
N
N
N
N
R
N
N
H
N
c
13
NH
N
N
N
d
R
S
N
N
a: R = H
21
h: R = m-CN
20 (45%)
i: R = m-CONH2
a
: R = H (70%)
b: R = m-CN (88%)
c
: R = m-CONH2 (53%)
Scheme 3. Reagents and conditions: (a) i-PrMgCl, anhydrous THF, anhydrous DMF, 1 N HCl, 0 °C to rt, 4 h; (b) (i) diphenyl (6-methylpyridin-2-yl)(phenylamino)meth-
ylphosphonate, Cs2CO3, THF/i-PrOH (4:1), rt, 16 h; (ii) 3 N HCl, rt, 1 h; (c) (i) DMFÁDMA, 90 °C, 4 h; (ii) N2H4ÁH2O, EtOH, reflux, 3 h; (d) NaH, anhydrous THF, 0 °C to rt, 1 h.

6052
C. H. Jin et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6049–6053
Table 1
Inhibitory activity of 1-substituted-3-(6-methylpyridin-2-yl)-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles 14a–ae, 16a, 16b, and 21a–c on ALK5
N
N
N
N
N
N
N
N
N
H
N
H
N
H
N
N
N
N
N
N
N
R
R
S
S
N
S
N
N
R
N
14a-ae
21a-c
16a, 16b
Compd
R
IC50 (nM)
ALK5a
Selectivity indexc
p3TP-luciferase activity (% control)d,e
p38ab
Mock
TGF-b
14a
14b
14c
14d
14e
14f
14g
14h
14i
14j
14k
14l
14m
14n
14o
14p
14q
14r
14s
14t
14u
14v
14w
14x
14y
—
—
H
m-F
m-Cl
m-Me
m-CF3
m-OMe
m-OCF3
m-CN
m-CONH2
p-F
p-Cl
—
—
—
—
—
—
6
1.9
100 2.6
10 1.4
2.82
4.43
2.37
3.98
3.01
2.26
10.00
4.50
3.45
4.09
3.87
6.61
3.04
0.57
3.30
2.95
15.60
5.39
2.83
4.00
5.69
3.98
2.12
3.20
3.78
5.08
3.54
3.97
4.12
4.37
2.49
39.47
95.70
102.00
85.30
66.80
54.40
17.20
2800
5680
5440
3680
1980
>10000
>10000
6430
4280
6670
5530
2960
4120
6360
5220
5230
4950
3000
3220
3680
7560
3830
1450
3810
1690
2850
5810
5390
5040
3840
2460
496
993
1282
2295
925
658
>4425
>1000
1429
1241
1631
1429
448
1355
11158
1582
1773
317
557
1138
920
1329
962
684
1191
447
561
1641
1358
1223
879
988
13
8
8
8
5
8
7
5
8
6
7
6
6
6
7
5
9
4
5
0.1
0.6
0.4
0.1
0.5
0.4
0.4
0.4
0.1
0.7
0.1
0.2
0.2
0.8
0.1
0.4
0.3
0.3
p-Me
p-CF3
p-OMe
p-OCF3
p-CN
p-CONH2
o-F
o-Cl
o-Me
21 0.7
11 1.3
o-OMe
o-OCF3
3,4-F2
3,5-F2
3,4-Cl2
3,5-Cl2
3,4-Me2
3,5-Me2
3,5-(CF3)2
3,4-(OMe)2
3,5-(OMe)2
Me
Benzyl
H
m-CN
m-CONH2
—
5
7
6
6
7
7
9
8
0.1
1.0
0.2
0.3
0.4
0.5
0.2
1.4
14z
14aa
14ab
14ac
14ad
14ae
16a
16b
21a
21b
21c
10 1.1
0.4
8
81 0.8
90 4.5
64 3.2
53 2.1
63 1.5
34 2.7
20 0.7
598
6
71
38
99
11
28
7260
3200
6580
594
1 (SB-505124)
6 (IN-1130)
—
480
a
ALK5 was expressed in Sf9 insect cells as human recombinant GST-fusion protein by means of the vaculovirus expression system. A proprietary radioisotopic protein
kinase assay (33PanQinaseÒ Activity Assay) was performed at ProQinase GmbH (Freiburg, Germany) using casein as a substrate.
b
p38a MAP kinase was expressed as untagged human recombinant protein in E. coli. The enzyme was purified by Ni-NTH-agarose (Qiagen). A proprietary radioisotopic
protein kinase assay (33PanQinaseÒ Activity Assay) was performed at ProQinase GmbH (Freiburg, Germany) using ATF2 as a substrate.
c
IC50 of p38a/IC50 of ALK5.
Luciferase activity was determined at a concentration of 100 nM of inhibitor.
Activity is given as the mean SD of three independent experiments run in triplicate relative to control incubations with DMSO vehicle.
d
e
tuted compound 14w (IC50 = 2.12 nM) and 3,5-(OMe)2 substituted
compound 14ae (IC50 = 2.49 nM) displayed slightly more potent
ALK5 inhibition than the 14a.
activity compared to the unsubstituted ones, thus, 21a (IC50 =
102.00 nM), 21b (IC50 = 85.30 nM), and 21c (IC50 = 66.80 nM) were
36-fold, 19-fold, and 19-fold less potent than the 14a, 14h
(IC50 = 4.50 nM), and 14i (IC50 = 3.45 nM), respectively. The pyra-
zole 14n showed the most potent ALK5 inhibitory activity in this
series of compounds, and was 95-fold and 30-fold more potent
than 1 (IC50 = 54.40 nM) and 6 (IC50 = 17.20 nM), respectively.
To evaluate TGF-b-induced downstream transcriptional activa-
tion to ALK5 signaling, cell-based luciferase activity of 14a–ae,
16a, 16b, and 21a–c was measured using HaCaT cells permanently
transfected with p3TP-luciferase reporter construct at a concentra
tion of 100 nM (Table 1). The p3TP-luciferase reporter construct
Contrary to the pyrazoles 14a–ae possessing
a phen-
ycarbothioamido moiety, the pyrazoles 16a (IC50 = 39.47 nM) and
16b (IC50 = 95.70 nM) possessing a 4-piperidinylcarbothioamido
moiety were 14-fold and 34-fold less potent in ALK5 inhibition
than the 14a. This result may suggest us that the binding region
for the phenyl ring of 14a–ae or piperidinyl ring of 16a and 16b
in ATP binding site of ALK5 is rather planar. The compounds
21a–c having a methyl substituent at the 2-position of [1,2,4]
triazolo[1,5-a]pyridin-6-yl moiety displayed much lower ALK5

C. H. Jin et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6049–6053
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contains three Ap-1 binding elements and the plasminogen-activa-
tor inhibitor-1 (PAI-1) promoter.41 Similar to kinase assay, all the
pyrazoles 14a–ae significantly inhibited the luciferase activity,
thus, showing 89–96% inhibition except the o-Me substituted com-
pound 14t (79% inhibition). But, the pyrazoles 16a and 16b showed
no significant ALK5 inhibition (<20% inhibition), and 21a–c showed
only modest ALK5 inhibition (36–47% inhibition). The competitor
compounds 1 and 6 showed 66% and 80% inhibition in this assay,
respectively.
The kinase domain of p38a MAP kinase is known to be one of the
most homologous to that of ALK5,42 therefore, it was chosen to
examine the selectivity profile of this series of compounds. The pyr-
azoles 14a–ae and 21a–cpossessing a phenycarbothioamido moiety
did not inhibit p38a MAP kinase efficiently, showing IC50 values of
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>1450 nM, whereas the pyrazoles 16a (IC50 = 496 nM) and 16b
(IC50 = 598 nM) possessing a 4-piperidinylcarbothioamido moiety
were more inhibitory than the 14a–ae and 21a–c. The p-OMe substi-
tuted compound 14n was the most selective in this series of com-
pounds, showing a selectivity index of 11158 that is much higher
than those of 1 (11) and 6 (28). It is noteworthy that the compound
14n43 is the most potent and selective ALK5 inhibitor reported to
date.
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In this Letter, a series of 1-substituted-3-(6-methylpyridin-2-
yl)-4-([1,2,4]triazolo[1,5-a]pyridin-6-yl)pyrazoles having
a thi-
oamido linkage between a pyrazole ring and a phenyl ring or a
piperidinyl ring has been prepared and evaluated for their ALK5
inhibitory activity in an enzyme assay and in a cell-based luciferase
reporter assay. It has been proved that incorporation of the
[1,2,4]triazolo[1,5-a]pyridin-6-yl moiety and phenycarbothioami-
do moiety at the 4- and 1-positions of the pyrazole ring, respec-
tively, significantly increased both ALK5 inhibitory activity and
selectivity. The most potent compound 14n inhibited ALK5 phos-
phorylation with IC50 value of 0.57 nM and showed 94% inhibition
at 100 nM in a luciferase reporter assay using HaCaT cells perma-
nently transfected with p3TP-luc reporter construct. The selectivity
index of 14n against p38a MAP kinase is 11158 that is much high-
er than those of 1 (11) and 6 (28).
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Acknowledgment
38. Gerlach, M.; Seipelt, I.; Guenther, E.; Polymeropoulos, E.; Schuster, T.; Claus, E.
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This work was supported by RP-Grant 2010 of Ewha Womans
University.
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2.50 (s, 3H), 3.86 (s, 3H), 7.00 (m, 2H), 7.19 (dd, 1H, J = 7.2, 1.2 Hz), 7.63–7.69
(m, 5H), 7.73 (dd, 1H, J = 9.2, 0.8 Hz), 8.36 (s, 1H), 8.96 (dd, 1H, J = 1.6, 0.8 Hz),
9.00 (s, 1H), 10.64 (br s, 1H); HRMS-ESI: m/z [M+H]+ calcd for C23H20N7OS:
442.1445, found 442.1447.