Synthesis and SAR study of imidazoquinolines as a novel structural class of microsomal prostaglandin E2 synthase-1 inhibitors

Article abstract of DOI:10.1016/j.bmcl.2011.11.015

The imidazoquinoline derivative 1 was found as a novel mPGES-1 inhibitor. Optimization of 1 led to the identification of the 2-chlorophenyl group at the C(2)-position and the quinolone structure at the C(4)-position. Compound 33, the most potent synthesized compound, showed excellent mPGES-1 inhibition (IC ₅₀ = 9.1 nM) with high selectivity (>1000-fold) over both COX-1 and COX-2.

Full text of DOI:10.1016/j.bmcl.2011.11.015

Bioorganic & Medicinal Chemistry Letters 22 (2012) 285–288
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and SAR study of imidazoquinolines as a novel structural class
of microsomal prostaglandin E2 synthase-1 inhibitors
Tomoya Shiro a, Hirotada Takahashi a, Keisuke Kakiguchi a, Yoshifumi Inoue b, Keiki Masuda a,
Hidetaka Nagata a, Masanori Tobe b,
⇑
a Drug Research Division, Dainippon Sumitomo Pharma. Co. Ltd, 3-1-98 Kasugade-naka, Konohana-ku, Osaka 554-0022, Japan
b Drug Research Division, Dainippon Sumitomo Pharma. Co. Ltd, 33-94 Enoki, Suita, Osaka 564-0053, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
The imidazoquinoline derivative 1 was found as a novel mPGES-1 inhibitor. Optimization of 1 led to
the identification of the 2-chlorophenyl group at the C(2)-position and the quinolone structure at the
C(4)-position. Compound 33, the most potent synthesized compound, showed excellent mPGES-1
inhibition (IC50 = 9.1 nM) with high selectivity (>1000-fold) over both COX-1 and COX-2.
Ó 2011 Elsevier Ltd. All rights reserved.
Received 4 October 2011
Revised 4 November 2011
Accepted 5 November 2011
Available online 11 November 2011
Keywords:
mPGES-1
PGE2
Imidazoquinoline
COX-1
COX-2
Prostaglandin E2 (PGE2) is well known as an important lipid
mediator with broad range of biological activities in various cells
and tissues, and is biosynthesized by sequential conversion of
arachidonic acid with various metabolic enzymes, such as cyclo-
oxygenase-1 (COX-1), -2 (COX-2), and PGE synthases (PGES).
Among these enzymes, PGES is a terminal enzyme in the biosyn-
thesis of PGE2 classified into three isoforms: microsomal PGES-1
(mPGES-1), -2 (mPGES-2), and cytosolic PGES (cPGES).1 mPGES-1,
which is functionally linked to COX-2, is inducible and responsi-
ble for the release of PGE2 in response to inflammatory stimuli,
plasma without affecting that of other prostaglandins, such as
TXB2 and PGF1a 6
Unlike COX-1/2 inhibitors, administration of
selective mPGES-1 inhibitors resulted in no undesirable effects
on mice gastrointestinal system.6 Based on the information de-
scribed above, it is believed that selective mPGES-1 inhibitors
could be ideal agents for the treatment of inflammation, pain,
and other PGE2-related disorders.7a–e
To date, several selective mPGES-1 inhibitors have been re-
ported. Among them, the phenanthreneimidazole derivatives
were reported by Merck Frosst researchers and represented here
by MF-63.8a,8b MF-63 inhibits mPGES-1 with an IC50 value in the
subnanomolar range. Pfizer researchers reported the dioxobenzo-
thiazinone derivative PF-9184,8c which inhibits mPGES-1 with an
IC50 value of 16 nM (Fig. 1). However, no compound has reached
clinical development. In our search for novel potent mPGES-1
inhibitors, we conducted a high-throughput screening (HTS) cam-
paign9a of our chemical library and found the imidazoquinoline
.
such as IL-1b, TNF-a
, and LPS.2 Indeed, a previous study has
shown that PGE2 production by LPS is completely suppressed in
peritoneal macrophages derived from mPGES-1 knockout mice.3
This allowed the use of mPGES-1 knockout mice as models of
various diseases, such as collagen induced arthritis, pain hyper-
sensitivity, and neuropathic pain. The symptoms of such diseases
were significantly relieved by suppression of PGE2 production.4a–c
As mPGES-1 knockout mice show increased prostacyclin (PGI2)
level in plasma with normal thromboxanes levels,5 it is expected
that cardiovascular system related side effects, such as thrombo-
sis and myocardial infarction, observed with selective COX-2
inhibitors, would not manifest with mPGES-1 inhibitors. A recent
study in human mPGES-1 knock-in mice has shown that a selec-
tive mPGES-1 inhibitor specifically reduced PGE2 production in
Cl
NC
4
N
OH
O
Cl
N
2
N
N
H
N
H
N
H
NH
O
NC
S
Cl
Br
O
PF-9184
1
MF-63
⇑
Corresponding author. Tel.: +81 6 6337 5927; fax: +81 6 6337 6010.
E-mail address: masanori-tobe@ds-pharma.co.jp (M. Tobe).
Figure 1. Structures of reported mPGES-1 inhibitors and compound 1.
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.11.015

286
T. Shiro et al. / Bioorg. Med. Chem. Lett. 22 (2012) 285–288
derivative 1 as a hit compound. Compound 1, which has a simple
structure favoring various substitutions, showed moderate inhib-
itory activity for mPGES-1 (60% at 10 lM). To improve the
We initially investigated the effects of different substituents at
the C(2)-position of 1 on mPGES-1 inhibitory activity (Table 1).9b
Replacement of the phenyl group by several alkyl groups (5À9) re-
sulted in a decrease in the inhibitory activity. However, among the
synthesized compounds, those with cyclic alkyl group (8 and 9)
showed slightly better inhibitory activity than those with straight
or blanched alkyl group (5À7), suggesting that sterically large
groups are preferable substituents at the C(2)-position. Com-
pounds 10À12, having 2-, 3-, and 4-pyridyl groups, respectively,
showed decreased inhibitory activity (19%, 26%, and 22%, respec-
mPGES-1 inhibitory activity of 1, we initiated a structure–activity
relationship (SAR) study. Here, we report the synthesis and
biological evaluation of a novel series of imidazoquinolines, in
particular, chemical modification of the substituents at the
C(2)- and the C(4)-positions of 1.
Procedure for the preparation of compounds 5À26 having the
C(2)-substituted imidazoquinoline ring system is shown in Scheme
1. Reaction of 7-bromo-4-chloro-3-nitroquinoline (2)10 with 28%
ammonia provided 4-amino-7-bromo-3-nitroquinoline (3). Reduc-
tion of 3 with iron powder in the presence of ammonium chloride
provided 7-bromo-3,4-diaminoquinoline (4).
With the exception of 5 and 6, the C(2)-substituted imidazo-
quinolines 7À26 were prepared by cyclization of 4 with appropri-
ate aldehydes in the presence of sodium pyrosulfate, respectively.
The methyl and the n-butyl derivatives 5 and 6 were prepared by
heating of 4 with appropriate orthoformates in the presence of cat-
alytic amount of p-toluenesulfonic acid, respectively. The tautom-
ers of the synthesized imidazoquinoline derivatives in the
imidazole moiety were not identified by the structural analysis.
The synthetic pathway to the C(4)-substituted imidazoquinolines
is shown in Scheme 2. Oxidation of 13 with mCPBA followed by
chlorination of the N-oxide with phosphorous oxychloride afforded
27 in 63% yield. The C(4)-substituted imidazoquinolines 28À32
were prepared by reaction of 27 with appropriate alkoxides,
respectively. Reaction of 27 with hydrochloric acid provided the
corresponding the imidazoquinolone derivative 33. The quinolone
structure of 33 was established on the basis of the characteristic
nuclear Overhauser effects (NOE; in DMSO), which were observed
between the N-proton and the imidazoquinolone proton at the 6
position by NOESY experiments. We also observed carbonyl vibra-
tion at 1654 cmÀ1 by Fourier transform infrared spectrum, indicat-
ing the presence of the quinolone structure of 33 in the solid state.
tively, at 10 lM). These results indicate that a lipophilic aromatic
group would be suitable at the C(2)-position of 1.
To improve the inhibitory activity of 1 for mPGES-1, we inves-
tigated the effects of different substituents on the phenyl group
(Table 2). Among the chlorine atom substituted compounds
13À15, the ortho-chloro derivative 13 showed the most potent
inhibitory activity for mPGES-1 compared with the meta-chloro
and the para-chloro derivatives 14 and 15. Compound 13 showed
approximately 40-fold higher mPGES-1 inhibitory activity over
the unsubstituted 1. The same tendency was observed when the
position of the electron-donating methyl group on the phenyl ring
was changed from an ortho- to a meta- or a para-position (16 vs 17
and 18). These findings suggest that the substituent at the ortho
position should be selected to cause steric repulsion between the
phenyl group and the imidazoquinoline core. This steric repulsion
can force the phenyl group to orientate at proper twist angle for
potent inhibitory activity.
On the basis of these data, we performed a ortho-substitution
study on the phenyl ring at the C(2)-position of the imidazoquino-
line ring. As shown in Table 2, the bromine derivative 19 displayed
a two-fold loss in the inhibitory activity compared with the
chlorine derivative 13. The fluorine derivative 20 showed great loss
of the inhibitory activity (IC50 = 6970 nM). Furthermore, as shown
by 21, introduction of a bulky phenyl group also showed a 18-fold
loss in the inhibitory activity over 13. These results indicate that
the ortho-substituents on the phenyl ring should be a moderate-
sized substituent (e.g., the chlorine atom), which affects the above
mentioned steric repulsion, to show a potent mPGES-1 inhibitory
activity. The inhibitory activities of compounds having an elec-
tron-withdrawing group in the ortho position on the phenyl group
(such as the trifluoromethyl, the nitro, and the cyano derivatives
22À24) were not improved toward mPGES-1 response. The
dimethylamino and the hydroxyl derivatives 25 and 26 showed
significantly reduced inhibitory activity compared with the chlo-
rine derivative 13. These results suggest that the placement of a
hydrophilic group on the phenyl group was not suitable to exert
its mPGES-1 inhibitory activity.
NO2
NH2
NH2
NH2
NO2
Cl
N
N
N
a
b
Br
Br
Br
2
3
4
c or d
5: R1 = Me
6: R1 = n-Bu
7: R1 = i-Pr
13: R1 = 2-Cl-Ph 20: R1 = 2-F-Ph
14: R1 = 3-Cl-Ph 21: R1 = 2-Ph-Ph
15: R1 = 4-Cl-Ph 22: R1 = 2-CF3-Ph
N
N
R1
: R1 = c-pentyl
R
1 = 2-Me-Ph
1 = 2-NO2-Ph
23:
R
N
8
16:
H
9: R1 = 4-pyranyl 17: R1 = 3-Me-Ph 24: R1 = 2-CN-Ph
10: R1 = 2-pyridyl 18: R1 = 4-Me-Ph 25: R1 = 2-NMe2-Ph
11: R1 = 3-pyridyl 19: R1 = 2-Br-Ph 26: R1 = 2-OH-Ph
12: R1 = 4-pyridyl
Br
To further improve the inhibitory activity for mPGES-1, we next
focused on the substitution studies at the C(4)-position on the imi-
dazoquinoline ring, keeping the 2-(2-chloro)phenyl group constant.
As shown in Table 3, the phenanthreneimidazole derivative MF-63,
used here as positive control, showed very potent inhibitory activity
Scheme 1. Reagents and conditions: (a) 28% NH3 aq, MeCN, 40 °C, 5 h, 87%; (b) Fe,
NH4Cl, EtOH/H2O, reflux, 4 h, 86%; (c) R1-C(OEt)3, p-toluenesulfonic acid, NMP,
120 °C, 5 h, 50À56%; (d) R1-CHO, Na2S2O7, DMF, 140 °C, 4À8 h, 19À96%.
28: R2 = OPh
R2
Cl
: R2 = O(c-hexyl)
29
N
N
c
30: R2 = OBn
31: R2 = OMe
N
H
Cl
Cl
Cl
Cl
: R2 = OEt
Br
32
-O
a
+
b
N
N
N
N
N
N
N
N
N
H
H
O
H
3
Cl
NOE
5
4
Br
Br
d
H
Br
N
13
27
2
N
6
H
N
1
H
7
Br
33
Scheme 2. Reagents and conditions: (a) mCPBA, CH2Cl2, rt, 2 h; (b) POCl3, reflux, 2 h, 63% in 2 steps; (c) R2-Na or R2-K, NMP or DMSO, 90À150 °C, 0.5À1 h, 33À97%; (d) cHCl,
EtOH/H2O, reflux, 8 h, 89%.

T. Shiro et al. / Bioorg. Med. Chem. Lett. 22 (2012) 285–288
287
Table 1
cyclohexyloxy, and the benzyloxy derivatives 28À30, which had
IC50 values comparable to that of the unsubstituted 13. These find-
ings indicated that compounds having the imidazoquinoline scaf-
fold would have different SAR from MF-63.
Effects of substituents at the C(2)-position on mPGES-1 inhibitory activity
N
2
N
R1
N
Next, we introduced several small substituents, such as a lower
aliphatic alkoxy group and a hydroxyl group. The methoxy and the
ethoxy derivatives 31 and 32 exhibited improved inhibitory activ-
ity compared with 13. In particular, compound 31 led to a four-fold
higher inhibitory activity for mPGES-1 over 13. These findings indi-
cate that a small alkoxy group is preferable for better inhibitory
activity. Furthermore, we evaluated the inhibitory activity of the
imidazoquinolone derivative 33. Surprisingly, compound 33
showed significantly improved inhibitory activity, reaching 27-fold
that of 13 and was almost equipotent to that of MF-63. Compound
33,11 the most potent synthesized compound in this study, was
next evaluated for its mPGES-1 selectivity over COX-1 and COX-
H
Br
Compound
R1
mPGES-1 % inhibitiona at 10
lM
1
5
6
7
8
9
10
11
12
Ph
Me
n-Bu
i-Pr
60
27
22
13
39
46
19
26
22
c-Pentyl
4-Pyranyl
2-Pyridyl
3-Pyridyl
4-Pyridyl
a
See Ref. 9b for details.
2, and exhibited less than 10% inhibition at 10 l
M.12 These findings
indicate that compound 33 has over 1000-fold mPGES-1 selectivity
over COX-1 and COX-2.
Table 2
Effects of substituents on the C(2)-phenyl group on mPGES-1 inhibitory activity
In summary, HTS screening of our chemical library resulted in
the discovery of the imidazoquinoline derivative 1 as a novel
mPGES-1 inhibitor. Optimization of 1 led to the identification of
the 2-chlorophenyl group at the C(2)-position and the quinolone
structure at the C(4)-position. Compound 33, the most potent syn-
thesized compound, showed excellent mPGES-1 inhibition
(IC50 = 9.1 nM) with high selectivity (>1000-fold) over both COX-
1 and COX-2. At present, we are now investigating the SAR studies
of 7-substituted-imidazoquinolones. These studies will be reported
in due course.
N
2
N
R1
N
H
Br
a
Compound
R1
mPGES-1 IC50 (nM)
1
Ph
9500
251
859
>10,000
395
1490
>10,000
506
6970
4620
1000
669
687
6264
4116
13
14
15
16
17
18
19
20
21
22
23
24
25
26
2-Cl-Ph
3-Cl-Ph
4-Cl-Ph
2-Me-Ph
3-Me-Ph
4-Me-Ph
2-Br-Ph
2-F-Ph
2-Ph-Ph
2-CF3-Ph
2-NO2-Ph
2-CN-Ph
2-NMe2-Ph
2-OH-Ph
Acknowledgments
We are deeply grateful to Dr. Yoshiaki Isobe and Dr. Katsumi
Kubota for their support and encouragement.
References and notes
1. (a) Watanabe, K.; Kurihara, K.; Suzuki, T. Biochem. Biophys. Acta 1999, 1438,
406; (b) Tanioka, T.; Nakatani, Y.; Semmyo, N.; Murakami, M.; Kudo, I. J. Biol.
Chem. 2000, 275, 32775.
a
2. Murakami, M.; Naraba, H.; Tanioka, T.; Semmyo, N.; Nakatani, Y.; Kojima, F.;
Ikeda, T.; Fueki, M.; Ueno, A.; Oh-ishi, S.; Kudo, I. J. Biol. Chem. 2000, 275, 32783.
3. Uematsu, S.; Matsumoto, M.; Takeda, K.; Akira, S. J. Immunol. 2002, 168, 5811.
4. (a) Trebino, C. E.; Stock, J.; Gibbons, C. P.; Naiman, B. M.; Wachtmann, T. S.;
Umland, J. P.; Pandher, K.; Lapointe, J. M.; Saha, S.; Roach, M. L.; Carter, D.;
Thomas, N. A.; Durtschi, B. A.; McNeish, J. D.; Hambor, J. E.; Jakobsson, P. J.;
Carty, T. J.; Perez, J. R.; Audoly, L. P. Proc. Natl. Acad. Sci. U.S.A. 2003, 100, 9044;
(b) Kamai, D.; Yamakawa, K.; Takegoshi, Y.; Mikami-Nakanishi, M.; Nakatani,
Y.; Oh-Ishi, S.; Yasui, H.; Azuma, Y.; Hirasawa, N.; Ohuchi, K.; Kawaguchi, H.;
Ishikawa, Y.; Ishii, T.; Uematsu, S.; Akira, S.; Murakami, M.; Kudo, I. J. Biol. Chem.
2004, 279, 33684; (c) Mabuchi, T.; Kojima, H.; Abe, T.; Takagi, K.; Sakurai, M.;
Ohmiya, Y.; Uematsu, S.; Akira, S.; Watanabe, K.; Ito, S. Neuroreport 2004, 15,
1395.
See Ref. 9b for details.
Table 3
Effects of substituents at the C(4)-position on mPGES-1 inhibitory activity
R2
O
Cl
Cl
4
4
N
N
2
2
N
HN
7
N
N
7
H
H
Br
Br
33
13, 28−32
5. Cheng, Y.; Wang, M.; Yu, Y.; Lawson, J.; Funk, C. D.; FitzGerald, G. A. J. Clin.
Invest. 2006, 116, 1391.
6. Xu, D.; Rowland, S. E.; Clark, P.; Giroux, A.; Côté, B.; Guiral, S.; Salem, M.;
Ducharme, Y.; Friesen, R. W.; Méthot, N.; Mancini, J.; Audoly, L.; Riendeau, D. J.
Pharm. Exp. Ther. 2008, 326, 754.
R2
mPGES-1 IC50 (nM)
a
Compound
MF-63
13
28
29
30
31
32
33
—
H
4.3
251
330
306
172
62
OPh
O(c-hexyl)
OBn
OMe
OEt
—
7. (a) Claveau, D.; Sirinyan, M.; Guay, J.; Gordon, R.; Chan, C. C.; Bureau, Y.;
Riendeau, D.; Mancini, J. A. J. Immunol. 2003, 170, 4738; (b) Cohen, E. G.;
Almahmeed, T.; Du, B.; Golijanin, D.; Boyle, J. O.; Soslow, R. A.; Subbaramaiah,
K.; Dannenberg, A. J. Clin. Cancer Res. 2003, 9, 3425; (c) Ikeda–Matsuo, Y.; Ota,
A.; Fukada, T.; Uematsu, S.; Akira, S.; Sasaki, Y. Proc. Natl. Acad. Sci. U.S.A. 2006,
103, 11790; (d) Chaudhry, U. A.; Zhuang, H.; Crain, B. J.; Doré, S. Alzheimers
Dement. 2008, 4, 6; (e) Kihara, Y.; Matsushita, T.; Kita, Y.; Uematsu, S.; Akira, S.;
Kira, J.; Ishii, S.; Shimizu, T. Proc. Natl. Acad. Sci. U.S.A. 2009, 106, 21807.
8. References of reported mPGES-1 inhibitor: (a) Côté, B.; Boulet, L.; Brideau, C.;
Claveau, D.; Ethier, D.; Frenette, R.; Gagnon, M.; Giroux, A.; Guay, J.; Guiral, S.;
Mancini, J.; Martins, E.; Massé, F.; Méthot, N.; Riendeau, D.; Rubin, J.; Xu, D.; Yu,
H.; Ducharme, Y.; Friesen, R. W. Bioorg. Med. Chem. Lett. 2007, 17, 6816; (b)
Giroux, A.; Boulet, L.; Brideau, C.; Chau, A.; Claveau, D.; Côté, B.; Ethier, D.;
Frenette, R.; Gagnon, M.; Guay, J.; Guiral, S.; Mancini, J.; Martins, E.; Massé, F.;
Méthot, N.; Riendeau, D.; Rubin, J.; Xu, D.; Yu, H.; Ducharme, Y.; Friesen, R. W.
Bioorg. Med. Chem. Lett. 2009, 19, 5837; (c) Wang, J.; Limburg, D.; Carter, J.;
Mbalaviele, G.; Gierse, J. G. M. Bioorg. Med. Chem. Lett. 2010, 20, 1604.
108
9.1
a
See Ref. 9b for details.
for mPGES-1 (IC50 = 4.3 nM). Considering the structural similarity
between MF-63 and 13, we speculated that introduction of a hydro-
phobic bulky group at the C(4)-position of 13 would lead to im-
proved inhibitory activity for mPGES-1. However, no improvement
in mPGES-1 inhibitory activity was seen with the phenoxy, the

288
T. Shiro et al. / Bioorg. Med. Chem. Lett. 22 (2012) 285–288
9. (a) Takahashi, H.; Nagata, H.; Shiro, T.; Kase, H.; Tobe, M.; Shimonishi, M.; Ido,
10. Bryon, M.; Chad, H.; Tushar, K.; Shri, N. WO2005/123080 A2, 2005.
11. Compound 33; mp 320–324 °C (dec); 1H NMR (DMSO-d6, 300 MHz) d 13.90
(1H, s), 11.75 (1H, s), 8.02 (1H, d, J = 8.4 Hz), 7.79 (1H, , dd, J = 7.2, 1.8 Hz), 7.63–
7.67 (2H, m), 7.49–7.59 (2H, m), 7.43 (1H, dd, J = 8.4, 1.8 Hz). 13C NMR (DMSO-
d6, 75 MHz) d 137.6, 132.1, 131.5, 130.2, 129.4, 127.4, 124.7, 123.6, 120.7,
M. in preparation.; (b) Biological assay method for mPGES-1 inhibition: The
test compound in DMSO (final 1%) was added to 15 mg/mL microsome in
100 mM Tris–HCl at pH 8.0, 5 mM EDTA, 500 mM phenol, 1 mM GSH, 10 mM
Hematin. The microsome was prepared from mPGES-1 and COX-1 co-
transfected HEK293 cells. Next, the mixture was pre-incubated at rt for
30 min. The reaction was started by the addition of 12.5 mM arachidonic acid
dissolved in 0.1 N KOH. The reaction mixture was incubated at rt for 60 min.
The reaction was stopped by addition of 1 N HCl. The reaction mixture was
neutralized by 1 N NaOH and PGE2 levels were measured by HTRF. The IC50
values were calculated by a logistic regression method.
118.2.; IR (ATR)
m
1654, 3392 cmÀ1.; MS (ESI) m/z calcd for C16H9BrClN3O 375;
found 376 (M+H); Anal. Calcd for C16H9BrClN3OÁ0.25H2O; C, 50.69; H, 2.53; N,
11.08. Found: C, 50.59; H, 2.53; N, 11.01.
12. The inhibition assay for the COX-1 and -2 was performed by Ricerca
Biosciences.