2-Amino-[1,2,4]triazolo[1,5-a]pyridines as JAK2 inhibitors

Article abstract of DOI:10.1016/j.bmcl.2013.06.008

The advancement of a series of ligand efficient 2-amino-[1,2,4]triazolo[1, 5-a]pyridines, initially identified from high-throughput screening, to a JAK2 inhibitor with pharmacodynamic activity in a mouse xenograft model is disclosed.

Full text of DOI:10.1016/j.bmcl.2013.06.008

Bioorganic & Medicinal Chemistry Letters 23 (2013) 5014–5021
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
2-Amino-[1,2,4]triazolo[1,5-a]pyridines as JAK2 inhibitors
a
a
a
a
a
Michael Siu a, , Richard Pastor , Wendy Liu , Kathy Barrett , Megan Berry , Wade S. Blair ,
Christine Chang a, Jacob Z. Chen a, Charles Eigenbrot a, Nico Ghilardi a, Paul Gibbons a, Haiying He b,
Christopher A. Hurley c, Jane R. Kenny a, S. Cyrus Khojasteh a, Hoa Le a, Leslie Lee a, Joseph P. Lyssikatos a,
Steve Magnuson a, Rebecca Pulk a, Vickie Tsui a, Mark Ultsch a, Yisong Xiao b, Bing-yan Zhu a, Deepak
Sampath a
⇑
a Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, United States
b WuXi AppTec Co., Ltd., 288 Fute Zhong Road, Waigaoqiao Free Trade Zone, Shanghai 200131, PR China
c Argenta, 8/9 Spire Green Centre, Flex Meadow, Harlow, Essex CM19 5TR, United Kingdom
a r t i c l e i n f o
a b s t r a c t
Article history:
The advancement of a series of ligand efficient 2-amino-[1,2,4]triazolo[1,5-a]pyridines, initially identified
from high-throughput screening, to a JAK2 inhibitor with pharmacodynamic activity in a mouse xeno-
graft model is disclosed.
Received 8 March 2013
Revised 28 May 2013
Accepted 3 June 2013
Available online 15 June 2013
N
N
N
N
N
N
N
HN
O
HN
HN
Keywords:
JAK2
Janus kinase
Triazolopyridine
N
N
5
5
8
HO
N
Cl
O
OCH3
OCH3
N
H3C
1
9
29
JAK2 Ki = 0.0597 uM
LE = 0.45
JAK2 Ki = 0.0006 uM
LE = 0.41
JAK2 Ki = 0.0017 uM
LE = 0.45
Ó 2013 Elsevier Ltd. All rights reserved.
JAK2 (Janus kinase 2) is a member of a family of non-receptor
cytoplasmic tyrosine kinases which include JAK1, JAK3, and
TYK2.1 This family of kinases share similar structural motifs, and
all have a JH1 kinase domain at the C terminus with an adjacent
JH2 pseudokinase domain. The JAK/TYK kinases are activated upon
cytokine to cytokine receptor binding. Transduction of the cytokine
signal occurs by phosphorylation of STAT (signal transducers and
activators of transcription) proteins that proceed to upregulate
their respective target genes. Mutation of JAK2 at codon 617 from
valine to phenylalanine (V617F) in the pseudokinase domain is
prevalent in myeloproliferative diseases.2 This particular mutation
leads to kinase activity without cytokine activation in hematopoe-
itic cells. In chronic myeloproliferative diseases, the activating
JAK2 V617F mutation is found in >95% of patients with polycythe-
mia vera (characterized by overproduction of red blood cells), in
ꢀ50% of patients with essential thrombocythemia (characterized
by overproduction of platelets), and in ꢀ50% of patients with mye-
lofibrosis (characterized by fibrosis of the bone marrow).3 Accord-
ingly, the inhibition of JAK2 presents an opportunity for targeted
therapy for these myeloproliferative diseases.
A variety of JAK inhibitors have been reported in the literature4
and have entered clinical trials5 with the most advanced being
tofacitinib (JAK1/JAK3 inhibitor)6 and ruxolitinib (JAK1/JAK2 inhib-
itor).7 Herein, we disclose our initial efforts toward JAK2 inhibitors
from chemical matter discovered by high-throughput screening
with the goal of generating a compound with greater than 10-fold
biochemical selectivity for JAK2 over JAK1, JAK3, and TYK2.
In order to identify novel small molecule JAK2 inhibitors, our
internal collection of small molecules was screened against the
⇑
Corresponding author. Tel.: +1 650 467 7764; fax: +1 650 467 8922.
E-mail address: siu.michael@gene.com (M. Siu).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.06.008

M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
5015
kinase domain of JAK2. For the primary screen, percent inhibition
was measured at a concentration of 5 M. For a subset of com-
docked using Glide13 after preparation with LigPrep using the de-
fault settings (LigPrep, version 2.4, Schrödinger, LLC, New York,
NY, 2010.)
l
pounds which showed >50% inhibition, half-maximal inhibitory
concentrations (IC50) were determined. Based on these activity
measurements, the 5-substituted 2-amino-[1,2,4]triazolo[1,5-
a]pyridine 18 was one of the chemotypes selected for optimization
that had high ligand efficiency (Table 1, LE = 0.45).9 This chemo-
type was potent against JAK2 but not selective against JAK1 and
TYK2 (Table 1). For initial SAR development, both the cyclopropyl
and chlorophenyl positions were explored via parallel synthesis
(representative examples shown in Table 1). An approximate 7.5-
fold improvement in JAK2 biochemical potency was achieved by
introduction of appropriate lipophilicity on the aryl ring while
maintaining high ligand efficiency (iso-propylphenyl analog 4, Ta-
ble 1); however, close-in replacements of the cyclopropanecarbox-
amide (propionamide 7 and benzamide 8) did not maintain the
necessary binding affinity (Table 1). Moreover, sulfonamides such
as methylsulfonamide and cyclopropylsulfonamide at the R1 posi-
tion were not tolerated (data not shown).
To further improve on our initial analogs, structure-based de-
sign efforts were also undertaken. As attempts to obtain a crystal
structure of triazolopyridine 1 in the active site of JAK2 were
unsuccessful, a docking protocol using the JAK2 crystal structure
of TG10120910,11 was developed to generate a model of the JAK2-
1 complex. The protein was prepared using the default settings
in the Protein Preparation Workflow in Maestro12 and 1 was
This comparison of the triazolopyridine 1 to TG101209 (Fig. 1)
suggested that the cyclopropanecarboxamide could be replaced
with a phenylpiperazine moiety in order to improve van der Waals
interactions in the active site. We synthesized a series of phenyl-
piperazine containing analogs shown in Table 2. Methoxyphenyl
and methylsulfonylphenyl analogs were initially examined on the
scaffold to balance the additional lipophilicity of the phenylpiper-
azine group. In general, these substitutions led to significant inhib-
itory activity of JAK2 in the biochemical assay. From this initial
examination, meta-substitution or large para-substitution on the
5-aryl ring improved JAK2 selectivity over JAK1. Moreover, these
compounds were potent in our cellular assay—inhibition of phos-
phorylation of STAT5 in SET-2 cells with triazolopyridine 9 being
the most potent.
Triazolopyridine 9 is synthesized as shown in Scheme 1. Treat-
ment of 2-amino-6-bromopyridine with ethyl isothiocyanatofor-
mate provides thiourea 14 which cyclizes to the triazolopyridine
core 15 upon treatment with hydroxylamine hydrochloride. Palla-
dium catalyzed cross coupling of bromide 15 with 4-meth-
oxyphenylboronic acid provides amine 16. Diazotization of amine
16 with sodium nitrite and in situ trapping with iodide give iodide
17. Lastly, palladium catalyzed amination of iodide 17 with 4-(4-
methylpiperazin-1-yl)aniline affords triazolopyridine 9.
Table 1
N
HN
R1
N
N
R2
Ex
1
R1
R2
JAK2 Ki (
lM)
JAK1 Sel Indxa
JAK3 Sel Indxb
TYK2 Sel Indxc
ClogPd
LE
O
O
O
O
O
3-Cl
0.0597
0.7ꢁ
15ꢁ
1.1ꢁ
3.4
0.45
2
3
4
5
4-Cl
0.059
0.3ꢁ
0.44ꢁ
3.8ꢁ
1ꢁ
8.4ꢁ
18.8ꢁ
21ꢁ
1.6ꢁ
1.5ꢁ
6.2ꢁ
2.4ꢁ
3.4
3.6
4
0.45
0.39
0.46
0.40
3-CF3
3-iPr
0.0826
0.0080
0.207
3-OCH3
7.6ꢁ
2.7
O
6
7
3-iPr
3-iPr
0.0177
2.2
7.5ꢁ
32.1ꢁ
12ꢁ
4
0.46
0.30
H3CO
H3C
O
—
—
—
4.5
O
8
3-iPr
2.2
—
—
—
4.8
0.30
F
a
b
c
(JAK1 Ki/JAK2 Ki).
(JAK3 Ki/JAK2 Ki).
(TYK2 Ki/JAK2 Ki).
d
ClogP values were calculated using MoKa v1.1.0, available from Molecular Discovery Ltd.

5016
M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
H3C
CH3
CH3
Cl
CH3
N
HN
S
H
N
O
O
N
N
N
N
HN
HN
O
N
N
CH3
1
TG101209
(X-ray structure)
(model)
Figure 1. Left: triazolopyridine 1 modeled into X-ray structure of JAK2. Right: X-ray structure of TG101209–JAK2 complex.
Table 2
N
HN
N
N
R
N
N
H3C
Ex
R
JAK2 Ki (lM)
JAK1 Sel Indxa
JAK3 Sel Indxb
TYK2 Sel Indxc
pSTAT5 SET2 IC50
(lM)
ClogPd
LE
9
10
11
12
4-OCH3
3-SO2-CH3
3-OCH3
0.0006
0.0020
0.0009
0.0003
5.4ꢁ
12.4ꢁ
22.3ꢁ
13.5ꢁ
81ꢁ
12.3ꢁ
11ꢁ
0.0219
0.0731
0.0536
0.0329
5.1
3.9
5.2
3.8
0.41
0.36
0.40
0.40
11ꢁ
12.6ꢁ
12.7ꢁ
14.2ꢁ
17.4ꢁ
4-SO2-CH3
a
(JAK1 Ki/JAK2 Ki).
(JAK3 Ki/JAK2 Ki).
(TYK2 Ki/JAK2 Ki).
b
c
d
ClogP values were calculated using MoKa v1.1.0, available from Molecular Discovery Ltd.
Triazolopyridine 9 has desirable potency and appropriate phar-
With in vivo demonstration of JAK2 inhibition by triazolopyri-
dine 9 in hand, we targeted further improvement of the scaffold
for better JAK1 selectivity and pharmacokinetic properties. The
phenylpiperazine fragment of triazolopyridine 9 introduced signif-
icant binding affinity to JAK2, albeit with an electron-rich aniline.
In an attempt to address JAK1 selectivity, a cocrystal structure of
triazolopyridine 9 in the JAK2 kinase domain was obtained.15 In
Figure 3, the amino acid residue differences between JAK2 and
JAK1 are highlighted. Modification of the phenylpiperazine is ex-
pected to impact selectivity since this functional group binds in a
macokinetic properties in mice (CLp = 40 mL/min/kg, Vdss = 2 L/kg,
F = 13%) to examine in our pharmacokinetic/pharmacodynamic
model, the SET-2 human leukemic cell line derived from a patient
with essential thrombocythemia.14 Mice harboring SET-2 xeno-
graft tumors were treated with a single dose of triazolopyridine
9 at 100 mg/kg. After 1 and 4 h, pSTAT5 and total STAT5 in ex-
tracted tumors were measured by MSD electrochemiluminescence
detection (Fig. 2). At 1 and 4 h, there was a 75% and 52% reduction
in pSTAT5 levels relative to total STAT5, respectively.

M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
5017
H
N
H
N
To improve in vivo metabolic stability and avoid potential reac-
tive metabolites we modified the electron-rich phenylpiperazine.16
Thus, a series of electron-deficient analogs were prepared that in-
cluded a variety of substituted pyridine and phenyl isomers
(Table 3). 2-Methyl pyridines 18 and 23, 2-hydroxymethyl pyridine
19 and benzoic acid 24 maintained JAK2 potency with good ligand
efficiency. These modifications improved biochemical selectivity
against JAK1. Pharmacokinetic evaluation of compound 18 in rats
showed high plasma clearance17 and low bioavailability (CLp =
42 mL/min/kg, Vdss = 1.6 L/kg, F = 3%).
a
b
c
H2N
EtO
N
N
H2N
N
N
O
S
N
Br
Br
Br
13
14
15
N
N
N
N
N
N
H2N
I
HN
N
d
N
e
N
In order to reduce in vivo CL, we subsequently targeted com-
pounds with lower ClogP by examining a variety of polar 3-iso-
propyl phenyl replacements as shown in Table 4. Polar groups such
as 3-methylsulfonylphenyl in addition to alkyl pyrazoles were tol-
erated in terms of inhibition of the JAK2 enzyme; however,
evaluation of rat pharmacokinetics revealed that 3-methylsulfo-
nylphenyl 25 (CLp = 138 mL/min/kg, Vdss = 7.2 L/kg, F = 6%) and
N-ethyl pyrazole 27 (CLp = 100 mL/min/kg, Vdss = 2.4 L/kg, F = 1%)
resulted in CLp that were greater than liver blood flow.18 The rea-
son for the extremely high plasma clearance was not determined,19
and similar pharmacokinetic properties for these molecules with a
wide lipophilicity range were typically observed. Therefore, further
efforts to replace the aminopyridine and to examine an isomeric
scaffold were attempted.
N
N
OCH3
OCH3
OCH3
H3C
16
17
9
Scheme 1. Synthesis of compound 9. Reagents and conditions: (a) ethyl
isothiocyanatoformate, CH2Cl2, 90%; (b) NH2OHꢂHCl, diisopropylethyl amine, 1:1
methanol/ethanol, 70 °C, 80%; (c) PdCl2(PPh3)2, Na2CO3, 4-methoxyphenylboronic
acid, CH3CN, 120 °C, microwave, 83%; (d) p-toluenesolfonic acid, KI, NaNO2, CH3CN,
71%; (e) 4-(4-methylpiperazin-1-yl)aniline, 2-dicyclohexylphosphino-20,40,60-tri-
isopropylbiphenyl, NaOt-Bu, Pd2(dba)3, (CH3)2NCHO, 100 °C, 30%.
An isomeric scaffold was made by repositioning the ring junc-
tion nitrogen (Tables 5 and 6). Analogs of the isomeric 8-substi-
tuted 2-amino-[1,2,4]triazolo[1,5-a]pyridine had similar potency
(24 cf. 29), and a variety of benzoic acids, benzamides, and pyra-
zoles20 were examined. Benzamides 30–31 and pyrazoles 32–33
maintained biochemical and cellular potency against JAK2. Addi-
tionally, the benzamides with low ClogP exhibited improved rat
in vivo CLp (Example 31: in vivo rat CLp = 10 mL/min/kg, Vdss
=
0.3 L/kg, F = 15%). Heteroaromatic groups (pyrazole and pyridine)
with lipophilic substituents at the C-8 position of the scaffold were
also potent inhibitor of JAK2 as shown in Table 6.
Of the examples in Tables 5 and 6, benzoic acid 29 (logD (pH
7.4) = 1.01) has the best balance of physical properties. Benzoic
acid 29 does not form reactive metabolites (no glutathione adducts
upon incubation with liver microsomes and no time dependent
inhibition of cytochrome P450s were observed) and exhibits good
pharmacokinetic properties in rodents (mouse: CLp = 15 mL/min/
kg, Vdss = 0.97 L/kg, F = 81%, PPB = 96.2%; rat: CLp = 15 mL/min/kg,
Figure 2. Reduction of pSTAT5a,b in SET2 tumors after dosing of Triazolopyridine 9
(100 mg/kg PO) at 1 and 4 h, student t-test ⁄p <0.05, ⁄⁄p <0.001.
region of JAK2 with amino acid residue differences to JAK1 (JAK2
residues 931 on the hinge, 853 above the active site, and 939 which
is hydrogen bonded to an observed water).
853
JAK2 Q → JAK1 R
931
JAK2 Y → JAK1 F
H2O
939
JAK2 D → JAK1 E
Figure 3. X-ray structure of Triazolopyridine 9 complexed to JAK2 kinase domain. Active site amino acid differences between JAK2 and JAK1 highlighted in magenta.

5018
M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
Table 3
N
HN
N
R1
N
R2
Ex
18
R1
R2
JAK2 Ki (
l
M)
JAK1 Sel Indxa
JAK3 Sel Indxb
TYK2 Sel Indxc
pSTAT5 SET2 IC50
0.190
(lM)
ClogPd
LE
H3C
3-i-Pr
0.0011
26.3ꢁ
18.7ꢁ
13.2ꢁ
5.5
0.47
N
HO
19
20
3-i-Pr
3-i-Pr
0.0006
0.0406
20.8ꢁ
50ꢁ
27.1ꢁ
31ꢁ
11.3ꢁ
23.6ꢁ
0.117
1.4
4.2
6
0.47
0.39
N
H3C
N
H3C
H3C
H3C
N
21
22
23
3-i-Pr
0.0709
0.0091
0.0050
—
—
—
—
6
0.38
0.42
0.46
3-i-Pr
24.1ꢁ
5.3ꢁ
17.9ꢁ
18.3ꢁ
17.4ꢁ
7.9ꢁ
2
6.7
4.2
4-OCH3
0.174
N
O
24
4-OCH3
0.0016
2.3ꢁ
26.2ꢁ
5.4ꢁ
—
4.9
0.45
OH
a
(JAK1 Ki/JAK2 Ki).
(JAK3 Ki/JAK2 Ki).
(TYK2 Ki/JAK2 Ki).
b
c
d
ClogP values were calculated using MoKa v1.1.0, available from Molecular Discovery Ltd.
Table 4
Ex
25
R1
H
R2
JAK2 Ki (
lM)
JAK1 Sel Indxa
JAK3 Sel Indxb
TYK2 Sel Indxc
pSTAT5 SET2 IC50
0.446
(
lM)
ClogPd
LE
0.0193
9.6ꢁ
37ꢁ
9ꢁ
2.9
0.39
26
27
H
0.0048
0.003
8.4ꢁ
6ꢁ
38ꢁ
36ꢁ
7.9ꢁ
5ꢁ
–
3.2
1.9
0.48
0.47
OH
0.115
28
H
0.002
19.4ꢁ
50.9ꢁ
11.9ꢁ
0.165
4
0.46
a
(JAK1 Ki/JAK2 Ki).
(JAK3 Ki/JAK2 Ki).
(TYK2 Ki/JAK2 Ki).
b
c
d
CLogP values were calculated using MoKa v1.1.0, available from Molecular Discovery Ltd.

M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
5019
Table 5
N
N
HN
R1
N
R2
Ex
29
R1
R2
JAK2 Ki (
l
M)
JAK1 Sel Indxa
JAK3 Sel Indxb
TYK2 Sel Indxc
pSTAT5 SET2 IC50
0.212
(l
M)
ClogPd
LE
O
4-OCH3
0.0017
2.4ꢁ
35.2ꢁ
7.1ꢁ
4.9
0.45
0.41
0.38
OH
CH3
N
30
4-SO2-CH3
4-SO2-CH3
0.0006
0.0004
8.3ꢁ
86.5ꢁ
70ꢁ
16ꢁ
0.117
2.6
2.4
H3C
O
O
N
31
32
11.6ꢁ
17.3ꢁ
0.0534
O
H3C
HO
N
N
4-OCH3
4-OCH3
0.0014
0.0007
4.5ꢁ
45.9ꢁ
49.5ꢁ
11.4ꢁ
21.3ꢁ
0.089
3.2
4
0.45
0.43
O
N
N
33
15.2ꢁ
0.0869
a
(JAK1 Ki/JAK2 Ki).
(JAK3 Ki/JAK2 Ki).
(TYK2 Ki/JAK2 Ki).
b
c
d
ClogP values were calculated using MoKa v1.1.0, available from Molecular Discovery Ltd.
may explain the reasonable plasma protein binding, volume of dis-
tribution, and absorption. Compound 29 has JAK1 selectivity of
2.4ꢁ in our enzyme assays and sufficient JAK2 potency and favor-
able pharmacokinetic properties to examine in our pharmacody-
namic model.
In mice bearing SET-2 xenograft tumors, 29 resulted in sig-
nificant tumor growth inhibition at 100 mg/kg (daily oral dose)
that corresponded to 38% inhibition over 21 days of dosing
(Fig. 4).
8-Substituted triazolopyridine 29 is synthesized as shown in
Scheme 2. The isomeric triazolopyridine 41 is prepared from 2-
amino-3-bromopyridine 39 using similar chemistry as the 5-
substituted triazolopyridine 15 (Scheme 1). Amine 41 is converted
to carbamate 42 to enhance solubility for the subsequent palla-
dium-mediated cross coupling with 4-methoxyphenyl boronic
acid to form biaryl 43 (following an acidic deprotection). Palla-
dium catalyzed amination of amine 43 and methyl 4-iodobenzoate
provides 44. Saponification of the methyl ester affords benzoic
acid 29.
In conclusion, a series of triazolopyridines identified by high-
throughput screening were advanced using structure-based design
to ligand efficient inhibitors of JAK2 that demonstrate pharmaco-
dynamic response in a human tumor mouse model. During the
course of the optimization, we observed poor pharmacokinetic
properties for lipophilic and polar compounds with this scaffold.
As a result, we introduced an acid moiety to achieve compounds
with reasonable pharmacokinetic properties and moderate JAK2
potency and selectivity.
Figure 4. Tumor growth inhibition (38%; p-value = 0.0028) in mice (n = 10/group)
harboring SET-2 acute leukemia xenograft tumors with PO dosing of triazolopyri-
dine 29 QD for 21 days. Percent tumor growth = 100ꢁ (mean volume of tumors in
animals administered vehicle—mean volume of tumors in animals administered
triazolopyridine 29)/(mean volume of tumors in animals administered vehicle). The
p-values were calculated using the Dunnett’s test using JMP 5.0.
Vdss = 0.65 L/kg, F = 75%, PPB = 98.3%).21 Although compound 29 is
an acid, the reduced acidity of the 4-aminobenzoic acid moiety

5020
M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
Table 6
N
N
HN
N
R
O
OH
Ex
34
R
JAK2 Ki (
lM)
JAK1 Sel Indxa
JAK3 Sel Indxb
TYK2 Sel Indxc
pSTAT5 SET2 IC50
0.156
(lM)
ClogPd
LE
0.0005
6.8ꢁ
21ꢁ
7.5ꢁ
4.7
0.46
H3C
N
N
H3C
35
36
0.0011
0.0006
16.4ꢁ
8.9ꢁ
30.6ꢁ
10ꢁ
13.1ꢁ
7.4ꢁ
0.379
5
0.44
0.44
H3C
N
CH3
0.0237
4.8
N
N
37
N
N
0.0005
0.0005
8.2ꢁ
7.8ꢁ
18.1ꢁ
19.7ꢁ
9.6ꢁ
9ꢁ
0.031
0.151
5.2
4.2
0.43
0.47
38
N
N
H3C
CH3
a
(JAK1 Ki/JAK2 Ki).
(JAK3 Ki/JAK2 Ki).
(TYK2 Ki/JAK2 Ki).
b
c
d
ClogP values were calculated using MoKa v1.1.0, available from Molecular Discovery Ltd.
N
N
Boc
N
N
N
N
O
S
N
N
N
a
b
c
H2N
N
Boc
H2N
EtO
N
H
N
H
Br
Br
Br
Br
39
40
41
42
N
N
N
N
N
N
HN
H2N
HN
d
e
f
N
N
H3CO
HO
O
O
OCH3
OCH3
OCH3
43
44
29
Scheme 2. Synthesis of compound 29. Reagents and conditions: (a) ethyl isothiocyanatoformate, CH2Cl2; (b) NH2OHꢂHCl, diisopropylethyl amine, 1:1 methanol/ethanol,
70 °C, 73% over two-steps; (c) Boc anhydride, N,N-dimethyl-4-aminopyridine, pyridine, 70%; (d) (i) Pd(dppf)2Cl2, 4-methoxyphenylboronic acid, K3PO4, tetrahydrofuran,
100 °C, (ii) CF3CO2H, 79%; (e) Pd(OAc)2, Cs2CO3, Xantphos, methyl 4-iodobenzoate, dioxane, 80 °C, 73%; (f) 2 M aqueous NaOH, 85 °C, 75%.

M. Siu et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5014–5021
5021
Serdikoff, C.; Jan, M.; Angeles, T. S.; Albom, M. S.; Mason, J. L.; Aimone, L. D.;
Meyer, S. L.; Huang, Z.; WellsKnecht, K. J.; Ator, M. A.; Ruggeri, B. A.; Dorsey, B.
D. J. Med. Chem. 2012, 55, 5243.
References and notes
1. Yamaoka, K.; Saharinen, P.; Pesu, M.; Holt, V. E. T.; Silvennoinen, O.; O’Shea, J. J.
Genome Biol. 2004, 5, 253.
2. Levine, R. L.; Pardanani, A.; Tefferi, A.; Gilliland, D. G. Nat. Rev. Cancer 2007, 7,
673.
3. (a) Levine, R. L.; Belisle, C.; Wadleigh, M.; Zahrieh, D.; Lee, S.; Chagnon, P.;
Gilliand, D. G.; Busque, L. Blood 2006, 107, 4139; (b) Baxter, E. J.; Scott, L. M.;
Campbell, P. J.; East, C.; Fourouclas, N.; Swanton, S.; Vassiliou, G. S.; Bench, A. J.;
Boyd, E. M.; Curtin, N.; Scott, M. A.; Erber, W. N.; Green, A. R. Lancet 2005, 365,
1054; (c) Kralovics, R.; Passamonti, F.; Buser, A. S.; Teo, S. S.; Tiedt, R.; Passweq,
J. R.; Tichelli, A.; Cazzola, M.; Skoda, R. C. N. Engl. J. Med. 2005, 352, 1779.
9. Hopkins, A. L.; Groom, C. R.; Alex, A. Drug Discovery Today 2004, 9, 430.
10. PDB 4JI9.
11. Pardanani, A.; Hood, J.; Lasho, T.; Levine, R. L.; Martin, M. B.; Noronha, G.; Finke,
C.; Mak, C. C.; Mesa, R.; Zhu, H.; Soll, R.; Gilliland, D. G.; Tefferi, A. Leukemia
2007, 27, 1658.
12. Maestro, version 9.1, Schrodinger, LLC, New York, NY, 2010.
13. Glide, version 5.6, Schrödinger, LLC, New York, NY, 2010 (a) Friesner, R. A.;
Banks, J. L.; Murphy, R. B.; Halgren, T. A.; Klicic, J. J.; Mainz, D. T.; Repasky, M. P.;
Knoll, E. H.; Shaw, D. E.; Shelley, M.; Perry, J. K.; Francis, P.; Shenkin, P. S. J. Med.
Chem. 2004, 47, 1739; (b) Halgren, T. A.; Murphy, R. B.; Friesner, R. A.; Beard, H.
S.; Frye, L. L.; Pollard, W. T.; Banks, J. L. J. Med. Chem. 2004, 47, 1750; (c)
Friesner, R. A.; Murphy, R. B.; Repasky, M. P.; Frye, L. L.; Greenwood, J. R.;
Halgren, T. A.; Sanschagrin, P. C.; Mainz, D. T. J. Med. Chem. 2006, 49, 6177.
14. Uozumi, K.; Otsuka, M.; Ohno, N.; Moriyama, T.; Suzuki, S.; Shimotakahara, S.;
Matsumura, I.; Hanada, S.; Arima, T. Leukemia 2000, 14, 142.
15. PDB 4JIA.
16. (a) Blagg, J. Annu. Rep. Med. Chem. 2006, 41, 353; (b) Hop, C. E. C. A.; Kalgutkar,
A. S.; Soglia, J. R. Annu. Rep. Med. Chem. 2006, 41, 369; (c) Park, B. K.; Boobis, A.;
Clarke, S.; Goldring, C. E.; Jones, D.; Kenna, J. G.; Lambert, C.; Laverty, H. G.;
Naisbitt, D. J.; Nelson, S.; Nicoll-Griffith, D. A.; Obach, R. S.; Routledge, P.; Smith,
D. A.; Tweedie, D. J.; Vermeulen, N.; Williams, D. P.; Wilson, I. D.; Baillie, T. A.
Nat. Rev. Drug Disc. 2011, 10, 292.
}
4. (a) Kiss, R.; Sayeski, P. P.; Keseru, G. M. Expert Opin. Ther. Patents 2010, 20, 471.
and references cited therein; (b) Wilson, L. J. Expert Opin. Ther. Patents 2010, 20,
609. and references cited therein; (c) Wrobleski, S. T.; Pitts, W. J. Annu. Rep.
Med. Chem. 2009, 44, 247; (d) Purandare, A. V.; Lorenzi, M. V.; Lombardo, L. J.
Annu. Rep. Med. Chem. 2010, 45, 211.
5. (a) Quintás-Cardama, A.; Verstovsek, S. Expert Opin. Investig. Drugs 2011, 20,
961. and references cited therein; (b) Lucia, E.; Recchia, A. G.; Gentile, M.;
Bossio, S.; Vigna, E.; Mazzone, C.; Madeo, A.; Morabito, L.; Gigliotti, V.; de
Stefano, L.; Caruso, N.; Servillo, P.; Franzese, S.; Bisconte, M. G.; Gentile, C.;
Morabito, F. Expert Opin. Investig. Drugs 2011, 20, 41. and references cited
therein.
6. Flanagan, M. E.; Blumenkopf, T. A.; Brissette, W. H.; Brown, M. F.; Casavant, J.
M.; Shang-Poa, C.; Doty, J. L.; Elliott, E. A.; Fisher, M. B.; Hines, M.; Kent, C.;
Kudlacz, E. M.; Lillie, B. M.; Magnuson, K. S.; McCurdy, S. P.; Munchhof, M. J.;
Perry, B. D.; Sawyer, P. S.; Strelevitz, T. J.; Subramanyam, C.; Sun, J.; Whipple, D.
A.; Changelian, P. S. J. Med. Chem. 2010, 53, 8468.
7. Verstovsek, S.; Kantarjian, H.; Mesa, R. A.; Pardanani, A. D.; Cortes-Franco, J.;
Thomas, D. A.; Estrov, Z.; Fridman, J. S.; Bradley, E. C.; Erickson-Viitanen, S.;
Vaddi, K.; Levy, R.; Tefferi, A. N. Engl. J. Med. 2010, 363, 1117.
8. Contemporaneous to this work, Dugan et al. reported 2-amino-
[1,2,4]triazolo[1,5-a]pyridines as JAK2 inhibitors. Dugan, B. J.; Gingrich, D. E.;
Mesaros, E. F.; Milkiewicz, K. L.; Curry, M. A.; Zulli, A. L.; Dobrzanski, P.;
17. Assuming rat liver blood flow = 55 mL/min/kg.
18. Rat liver microsome CLhep: compound 25 = 10 mL/min/kg; compound
27 = 15 mL/min/kg; rat hepatocyte CLhep: compound 25 = 19 mL/min/kg;
compound 27 = 37 mL/min/kg).
19. Blood/plasma partition ratios are ꢀ1 for this series.
20. 1-alkyl-4-aminopyrazole as aniline replacements: Zabierek, A. A.; Konrad, K.
M.; Haidle, A. M. Tetrahredron Lett. 2008, 49, 2996.
21. Human liver microsomes CLhep = 6 mL/min/kg; human hepatocyte
CLhep = 0.2 mL/min/kg.