Novel non-benzimidazole chk2 kinase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2005.12.096

In a recent paper, [Arienti, K. L.; Brunmark, A.; Axe, F. U.; McClure, K. M.; Lee, A.; Blevitt, J.; Neff, D. K.; Huang, L.; Crawford, S.; Chennagiri, R. P.; Karlsson, L.; Brietenbucher, J. G. J. Med. Chem. 2005, 48, 1873], we described the discovery of a class of benzimidazole chk2 kinase inhibitors, exemplified by compound 1, which had radio-protective effects in human T-cells subjected to ionizing radiation. Here, a series of non-benzimidazole analogs intended to define the scope of the SAR about this new series of inhibitor, and allow for refinement of the binding model of these compounds to the chk2 kinase is described.

Full text of DOI:10.1016/j.bmcl.2005.12.096

Bioorganic & Medicinal Chemistry Letters 16 (2006) 1924–1928
Novel non-benzimidazole chk2 kinase inhibitors
Kelly J. McClure,* Liming Huang, Kristen L. Arienti, Frank U. Axe,
Anders Brunmark, Jon Blevitt and J. Guy Breitenbucher
Johnson and Johnson Pharmaceutical Research and Development, LLC, 3210 Merryfield Row, San Diego, CA 92121, USA
Received 28 October 2005; revised 19 December 2005; accepted 22 December 2005
Available online 25 January 2006
Abstract—In a recent paper, [Arienti, K. L.; Brunmark, A.; Axe, F. U.; McClure, K. M.; Lee, A.; Blevitt, J.; Neff, D. K.; Huang, L.;
Crawford, S.; Chennagiri, R. P.; Karlsson, L.; Brietenbucher, J. G. J. Med. Chem. 2005, 48, 1873], we described the discovery of a
class of benzimidazole chk2 kinase inhibitors, exemplified by compound 1, which had radio-protective effects in human T-cells sub-
jected to ionizing radiation. Here, a series of non-benzimidazole analogs intended to define the scope of the SAR about this new
series of inhibitor, and allow for refinement of the binding model of these compounds to the chk2 kinase is described.
Ó 2006 Elsevier Ltd. All rights reserved.
Radiotherapy is still the most commonly used treatment
for cancer patients, with >50% of all cancer patients
receiving some sort of radiotherapy. Side effects from
radiation therapy represent a major clinical problem,
which seriously affects both quality-of-life and clinical
outcome. Despite major improvements in the use of fo-
cused and fractionated dosing of radiation side effects
are still generally dose limiting. Thus, compounds that
act to increase the therapeutic window of radiotherapy
would have great clinical utility in the treatment of can-
cer. Additionally, such an agent might find utility in pro-
tecting humans from occupational exposure to ionizing
radiation.
model, is described. To probe the SAR of the benzimid-
azole-containing analogues, we replaced the benzimid-
azole core with closely related 5,6-fused heterocycles.
Maintaining a constant geometric arrangement of pen-
dant groups allowed for the precise determination of
the critical binding elements of the benzimidazole group.
The routes utilized to arrive at these analogues as well as
their corresponding biological data are reported below
(Table 1).
O
N
H2N
O
N
H
Chk2 is a serine/threonine kinase that was first described
in 1998.2 Chk2 is involved in the cellular response to
DNA strand breaks that occur upon exposure to c-radi-
ation.3 Chk2 knockout mice demonstrate increased sur-
vival to radiation. These animals also show suppressed
apoptosis in thymus, hippocampus, and skin tissues
after exposure to c-radiation.4,5 As a result, inhibition
of chk2 would be expected to be radio protective and
potentially useful as an adjunct to radiotherapy.
1
Simple methyl substitution at either nitrogen of the imid-
azole, as represented in compounds 4 and 7 of Table 1,
was accessed utilizing the route shown in Scheme 1.
In the first step, the fluoro group of 4-fluoro-3-nitroben-
zoic acid (2) was displaced with methylamine followed
by conversion of the acid to a primary amide, and nitro
reduction giving the diamine intermediate 3. The dia-
mine was then oxidatively condensed under standard
conditions with 4-aryloxybenzaldehyde giving com-
pound 4 in good yield. Compound 7 was prepared in a
similar fashion starting from 3-fluoro-4-nitrobenzoic
acid.
(4-Aryloxy-phenyl)benzimidazoles such as compound 1
(also compounds 1a, 1b and 1c in Table 1) were discov-
ered to be potent inhibitors of the chk2 kinase.1 In this
paper, a series of benzimidazole replacement analogues,
which were designed to further define the SAR of this
series of inhibitors as well as refine the existing binding
The benzoxazoles 9 and 11 were prepared via the route
shown in Scheme 2.
*
Corresponding author. E-mail: kmcclure@prdus.jnj.com
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.12.096

K. J. McClure et al. / Bioorg. Med. Chem. Lett. 16 (2006) 1924–1928
1925
Table 1. Imidazole ring replacements
HO2C
NH2
OH
CHO
O
O
V
Z
U
X
O
8
Y
R
W
a, b, c
65%
+
or
HO2C
OH
Compound
R
U
V
W
X
Y
Z
IC50
(nM)
NH2
1
H
H
NH2 CH CH
OH CH CH
C
C
C
C
C
C
C
C
C
N
C
C
NH
NH
NH
NH
NMe
N
N
N
N
N
N
55
640
10
1a
1b
1c
4
4-Cl OH CH CH
4-Cl NH2 CH CH
133
16
O
X
Y
H2N
H
H
H
H
NH2 CH CH
NH2 CH CH
NH2 CH CH
NH2 CH CH
>10,000
O
7
NMe 1540
N
9
O
>10,000
>10,000
5800
11
16
23
27
32
N
O
9: X = N, Y = O
11: X = O, Y = NH
4-Cl OH CH CH
N
CH
C
NH
N
H NH2 CH
4-Cl NH2 CH
>10,000
2000
77
Scheme 2. Reagents and condition: (a) Neat, 150 °C; (b) Pb(OAc)4,
AcOH; (c) SOCl2, NH3.
N
CH
NH
NH
N
4-Cl NH2
N
N
CHO
EtO2C
NO2
O
O
NO2
F
a, b, c
90%
HO
NH2
NHMe
EtO2C
NO2
H2N
a
Cl
+
HO
O
60%
O
3
2
Cl
13
12
14
H
N
O
EtO2C
d
b,c
N
N
d
O
H2N
80%
O
43%
65%
15
4
Cl
H
N
HO2C
O
O
O
F
NO2
5
NHMe
NH2
a, b, c
70%
HO
H2N
Cl
16
Scheme 3. Reagents and conditions: (a) NaOEt, EtOH, DMSO; (b)
Fe, HCl/H2O; (c) RuCl2(PPh3)4, toluene, 100 °C; (d) 7 N KOH, THF,
80 °C.
6
O
N
N
d
condensed with the corresponding 4-chloro-phenoxy
benzaldehyde giving compound 14 in good yield. Reduc-
tion of the nitro group with iron followed by ruthenium-
catalyzed cyclization led to the indole intermediate 15.7
Finally, hydrolysis of the ester group gave the target
compound 16.
H2N
O
60%
7
Scheme 1. Reagents and condition: (a) MeNH2, DIPEA; (b) SOCl2,
NH3; (c) H2, Pd/C, EtOH; (d) Na2S2O5, 4-phenoxybenzaldehyde,
DMF, 100 °C.
Heterocycle 23 was synthesized starting from commer-
cially available 4-phenoxybenzoic acid (17) in a five-step
sequence as outlined in Scheme 4.
Either of two aminohydroxybenzoic acids (8 or 10) were
condensed with 4-phenoxybenzaldehyde giving the
benzoxazolidine. This was then oxidized to the corre-
sponding benzoxazole by treatment with lead tetraace-
tate.6 The acid was converted to the acid chloride
followed by displacement with ammonia yielding the
primary amides 9 or 11.
Benzoic acid 17 was first converted to b-ketoester 18 by
treatment with methyl cyanoacetate. This intermediate
was then decarboxylated giving the a-cyanoketone 19.
Formation of the amino pyrazole substrate 20 was
achieved by addition of hydrazine. The key step in the
synthesis involved the formation of the pyrazolo pyrim-
idine ring system, which was accomplished by treating
the amino pyrazole with 3-dimethylamino-2-formyl-
acrylonitrile8 21. The nitrile was then converted to the
Indole 16 was prepared by the method outlined in
Scheme 3. The anion of 4-methyl-3-nitroethyl benzoate
was generated with sodium ethoxide and then

1926
K. J. McClure et al. / Bioorg. Med. Chem. Lett. 16 (2006) 1924–1928
CO2H
to the acid, followed by activation with thionyl chloride
and displacement with ammonia giving the target com-
pound 27.
O
O
b
OMe
a
CN
+
92%
NC
CO2Me
>99%
O
O
O
The regioisomeric imidazopyridine 32 was prepared as
illustrated in Scheme 6.
18
17
O
Functional group interconversions of the commercially
available pyridine 28 as described in preceding schemes
led to the diaminopyridine intermediate 30 in good
yield over five steps. The diaminopyridine 30 was then
oxidatively condensed with the appropriate benzalde-
hyde. Finally, the nitrile was hydrolyzed to the primary
amide by treatment with BF3Æ2AcOH affording the
compound 32.
O
O
CN
H
N
H
N
N
+
c
CN
21
>99%
H2N
20
19
NC
N
N
e
d
O
N
45%
35%
In a previous paper1 on benzimidazole inhibitors of
chk2, we described a binding model for this chemical
series based on docking of these inhibitors into a homol-
ogy model of the chk2 kinase as well as experimental
evidence, suggesting that these compounds are ATP
competitive inhibitors. The proposed binding model is
pictured in Figure 1. It shows that the 5-amido group
is involved in a key set of hydrogen bonding interactions
with the backbone carbonyl and amide nitrogen of Met
90 in the ATP pocket and that the benzimidazole and
the biaryl-ether groups lie in the hydrophobic region
of the ATP pocket. SAR in this study focused on the
biaryl-ether and 5-amido portions of the inhibitors.
22
O
N
H2N
N
O
N
23
Scheme 4. Reagents and conditions: (a) DEPC, DIEA, DMF; (b) wet
DMSO/NaCl; (c) N2H4, H2O, EtOH, reflux; (d) EtOH, reflux; (e)
BF3Æ2AcOH, H2O, 120 °C.
primary amide utilizing BF3Æ2AcOH/H2O yielding the
target compound 23.9
In the present study, changes were made to the benz-
imidazole, which maintained the topological [5,6]-ring
arrangement, but repositioned the heteroatoms to vari-
ous positions about the 5,6-ring system. The activities
of these analogs are shown in Table 1.
The imidazopyridine 27 in Table 1 was prepared as out-
lined in Scheme 5. The bromine on the commercially
available pyridine 24 was first displaced with copper
cyanide followed by reduction of the nitro group under
standard hydrogenation conditions. Formation of the
imidazopyridine ring 26 was accomplished using stan-
dard oxidative condensation conditions. Finally, the ni-
trile was converted to the primary amide by hydrolysis
NO2
Cl
NO2
NH2
d,e
a,b,c
30%
73%
MeO
N
Cl
N
29
28
CHO
O
CHO
NH2
f
+
NC
NH2
NH2
Br
NO2
NH2
c
a,b
80%
NC
NC
N
NH2
+
65%
40%
N
N
30
O
Cl
25
24
H
N
N
g
NC
N
O
d,e
37%
O
N
95%
N
H
N
31
Cl
26
Cl
O
H
N
N
O
H2N
O
N
H2N
N
O
N
H
N
32
Cl
27
Cl
Scheme 6. Reagents and conditions: (a) NH4OH, 3 days; (b) HBr,
AcOH, 60 °C; (c) POCl3, 100 °C to rt; (d) Zn(CN)2, Pd2dba3, DPPF,
DMF, H2O, 100 °C; (e) H2, Pd/C, EtOH; (f) Na2S2O5, DMF, 100 °C;
(g) BF3Æ2AcOH, H2O, 120 °C, 30 min.
Scheme 5. Reagents and conditions: (a) CuCN, NMP, 80 °C; (b) H2,
Pd/C, EtOH; (c) Na2S2O5, DMF, 100 °C; (d) HCO2H/HCl; (e) SO2Cl,
NH3.

K. J. McClure et al. / Bioorg. Med. Chem. Lett. 16 (2006) 1924–1928
1927
12
10
8
6
4
2
0
0
60
120
180
240
300
360
angle (degrees)
Figure 3. B3LYP/6-31G calculated potential energy surface for
*
picolinamide suggests an energy minimum favoring the amide in
preferred orientation for chk2 binding.
Figure 1. Binding model of compound 1 in ATP pocked of chk2
homology model.
The substitution of a nitrogen atom at the W position in
27 reduces activity considerably relative to 1. This result
is consistent with our proposed binding model since the
polar nitrogen atom is being buried in the hydrophobic
pocket, which is energetically unfavorable. On the con-
trary, substituting a nitrogen atom at the V position
(32) leads to a compound that is comparable in activity
to 1. This is consistent with our model for two reasons.
First, a nitrogen at the V position is more solvent ex-
posed and second, a nitrogen at the V position can form
an intra-molecular hydrogen bond with the additional
hydrogen on the 5-amido group, which results in a con-
formationally restricted orientation of the 5-CONH2
group. This latter notion that the 4-pyridyl-analog is
conformationally constrained is corroborated by the
X-ray crystal structure of picolinamide,11 which has
the amide group nitrogen oriented toward the pyridine
nitrogen and appears to form an intra-molecular hydro-
gen bond. This idea is also supported by both ab initio
DFT calculations that we performed as well.12 In fact,
one of the motivations for making this compound was
to constrain the amido group Figure 3.
It can be seen from analogue 4 that simple methyla-
tion of the Y-imidazole nitrogen altogether eliminates
the binding affinity for the kinase while methylation of
the Z-imidazole nitrogen reduces the binding affinity
by 30-fold relative to compound 1. In addition,
replacement of the Y nitrogen with a CH group, as
in compound 16, leads to a compound that is much
less active than compound 1 having an IC50 of
5.8 lM. These data strongly suggest that the unsubsti-
tuted imidazole nitrogen at the Y position is crucial
for activity as well as the NH group at the Z position
to a lesser extent. We can interpret these results in
terms of our binding model if there is a hydrogen
bond donor in proximity of the Y nitrogen and a
hydrogen bond acceptor in proximity of the Z nitro-
gen. There are a number of examples of kinase inhib-
itors in which the X-ray crystal structure with the
compound bound reveals that the conserved lysine at
the bottom of the ATP pocket often forms a hydrogen
bond with an acceptor group on the inhibitor.10 Crys-
tal structures also reveal that amino acid side chains
on the edge of the ATP pocket can serve as hydrogen
bond acceptors.10 Both of these donor and acceptor
sites occur in close proximity to the imidazole nitro-
gens in our models as depicted in Figure 2.
The aim of the work presented here was to determine the
importance of the benzimidazole group in binding to the
receptor and to gain a better understanding of the SAR
around this series of molecules. A number of heterocy-
clic benzimidazole replacements were made to answer
these questions. As can be seen from the decrease in
binding affinity with almost all of the prepared ana-
logues, the benzimidazole group seems to be preferred
for binding to the receptor. An extension of the binding
model to include key hydrogen bonding interactions be-
tween the benzimidazole nitrogens and Glu 94 and Lys
35 residues in the receptor would account for these find-
ings. The comparable activity to the parent benzimid-
azole found in compound 32 can readily be explained
by the solvent exposure to the pyridine nitrogen as well
as the conformational restriction of the amide group via
intramolecular hydrogen bonding to the pyridine
nitrogen.
LYS 35
N
GLU 88
H3N
O
HN
LEU 89
N
O
O
O
H
N
H
N
H
O
MET 90
N
H
S
O
O
Acknowledgments
GLU 94
The authors thank Dr. Jiejun Wu for his help in the
structure confirmation of the heterocyclic benzimidazole
Figure 2. Proposed hydrogen binding model explaining the impor-
tance of benzimidazole for chk2 binding.

1928
K. J. McClure et al. / Bioorg. Med. Chem. Lett. 16 (2006) 1924–1928
Suzki, H.; Nagashima, K.; Sawa, H.; Ikeda, K.; Motoy-
ama, N. EMBO J. 2002, 21, 5195.
replacements described in this paper. The authors also
thank Dr. Michael Hack for potential energy calcula-
tions of picolinamide.
5. Hirao, A.; Cheung, A.; Duncan, G.; Girard, P. M.; Elia,
A. J.; Wakeham, A.; Okada, H.; Sarkissian, T.; Wong, J.
A.; Sakai, T.; De Stanchina, E.; Bristow, R. G.; Suda, T.;
Lowe, S. W.; Jego, P. A.; Elledge, S. J.; Mak, T. W. Mol.
Cell. Biol. 2002, 22, 6521.
References and notes
6. Dunwell, D. W.; Evans, D.; Hicks, T. A. J. Med. Chem.
1975, 18, 1158.
7. Tsuji, Y.; Kotachi, S.; Huh, K. T.; Watanabe, Y. J. Org.
Chem. 1990, 55, 580.
8. Jachak, M.; Kriessmann, U.; Mittelbach, M.; Junek, H.
Monatsh. Chem. 1993, 124, 199.
9. Hauser, C. R.; Hoffenberg, D. S. J. Org. Chem. 1955, 20,
1448.
10. Huwe, A.; Mazitschek, R.; Giannis, A. Angew. Chem.
2003, 42, 2122.
11. Takano, T.; Sasada, Y.; Kakudo, M. Acta Cryst. 1966, 21,
514.
1. Arienti, K. L.; Brunmark, A.; Axe, F. U.; McClure, K.
M.; Lee, A.; Blevitt, J.; Neff, D. K.; Huang, L.; Crawford,
S.; Chennagiri, R. P.; Karlsson, L.; Brietenbucher, J. G. J.
Med. Chem. 2005, 48, 1873.
2. Matsuoka, S.; Huang, M.; Elledge, S. J. Science 1998, 282,
1893.
3. Chaturvedi, P.; Eng, W. K.; Zhu, Y.; Mattern, M. R.;
Mishra, R.; Hurle, M. R.; Zhang, X.; Annan, R. S.; Lu,
Q.; Faucette, L. F.; Scott, G. F.; Li, X.; Carr, S. A.;
Johnson, R. K.; Winkler, J. D.; Zhou, B. B. Oncogene
1999, 18, 4047.
4. Takai, H.; Naka, K.; Okada, Y.; Watanabe, M.; Harada,
N.; Saito, S.; Anderson, C. W.; Apella, E.; Nakanishi, M.;
12. Dewar, M. J. S.; Zoebisch, E. G.; Healy, E. F. J. Am.
Chem. Soc. 1985, 107, 3902.