Benzimidazole-2-carboxamides as novel NR2B selective NMDA receptor antagonists

Article abstract of DOI:10.1016/j.bmcl.2006.06.002

A novel series of benzimidazole-2-carboxamide derivatives was prepared and identified as NR2B selective NMDA receptor antagonists. The influence of some structural elements, like H-bond donor groups placed on the benzimidazole skeleton and the substitution pattern of the piperidine ring, on the biological activity was studied. Compound 6a showed excellent analgetic activity in the mouse formalin test following po administration.

Full text of DOI:10.1016/j.bmcl.2006.06.002

Bioorganic & Medicinal Chemistry Letters 16 (2006) 4638–4640
Benzimidazole-2-carboxamides as novel NR2B selective
NMDA receptor antagonists
a,
a
a
a
a
*
´
´
´
´
´
´
´
Istvan Borza, Sandor Kolok, Aniko Gere, Jozsef Nagy, Laszlo Fodor,
a
b
b
b
a
´
´
´
´
´
Kornel Galgoczy, Jozsef Fetter, Ferenc Bertha, Bela Agai, Csilla Horvath,
a
a
´
´
Sandor Farkas and Gyo¨rgy Domany
aGedeon Richter Ltd., Budapest, 10. POB. 27, H-1475, Hungary
bBudapest Univ. of Techn. and Econ., Budapest, POB. 91., H-1521, Hungary
Received 11 April 2006; revised 1 June 2006; accepted 1 June 2006
Available online 16 June 2006
Abstract—A novel series of benzimidazole-2-carboxamide derivatives was prepared and identified as NR2B selective NMDA recep-
tor antagonists. The influence of some structural elements, like H-bond donor groups placed on the benzimidazole skeleton and the
substitution pattern of the piperidine ring, on the biological activity was studied. Compound 6a showed excellent analgetic activity
in the mouse formalin test following po administration.
Ó 2006 Elsevier Ltd. All rights reserved.
The NR2B subunit containing NMDA receptors became
thoroughly studied targets in a wide range of CNS pathol-
ogies including chronic pain states.1 Numerous structur-
ally diverse antagonists of these receptors have been
reported and have recently been reviewed as well.2–4 An
important class of the NR2B subtype selective NMDA
receptor antagonists can be characterised as an H-bond
donor moiety containing benzene ring connected to the
nitrogen of a 4-benzylpiperidine via a three-atom long
spacer, such as Ro 25-69815 and CI-1041.6
basis of this lead, 2 and 3 were designed by adding the
5(6)-MsNH and 5(6)-OH hydrogen-bond-donating sub-
stituents, that showed excellent in vitro and in vivo
activity.7
N
N
Q
N
H
1 Q = H; 2 Q = MsNH; 3 Q = OH
Their publication prompted us to report on our own re-
sults achieved in the field of NR2B selective NMDA
antagonist benzimidazole-2-carboxamides.
HO
CH3
N
O
N
OH
Ro 25-6981
H
HO
N
O
4
N
O
S
F
O
N
N
H
HO
HO
CI-1041
O
Researchers at Merck have recently reported a series of
benzimidazole derivatives. The screening of their com-
pound library led to the identification of the benzimid-
azole derivative 1 with modest NR2B activity. On the
5
N
N
N
H
O
6
Keywords: NMDA; NR2B; Benzimidazole.
*
Cinnamide derivatives were the first NR2B subtype-se-
lective NMDA receptor antagonists which did not con-
Corresponding author. Fax: +36
i.borza@richter.hu
1
432 6002; e-mail:
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.06.002

I. Borza et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4638–4640
4639
tain basic N.8 Piperidine-based cinnamide 4 was selected
from this family as starting point of our research. In an
earlier publication we verified our hypothesis that poten-
cy may be enhanced by increasing the rigidity of cinna-
mide moiety. Incorporation of an NH group between
the a carbon atom and the benzene ring of the cinnamic
acid part led to the 6-hydroxy-indole-2-carboxamide (5),
with more than 6-fold higher potency.9 Encouraged by
this result, we decided to explore other heterobicyclic
systems. Replacing of the indole skeleton in compound
5 with the potentially bioequivalent benzimidazole
ring-system resulted in a more active compound (6a),
so we decided to prepare a series of benzimidazole-2-
carboxamides.
a functional assay where the inhibition of NMDA-
evoked increase of intracellular Ca2+ level was deter-
mined on rat cortical cell culture. Baseline and
NMDA-evoked changes of intracellular Ca2+ were mon-
itored by fluorimetric method using a Ca2+-selective
fluorescent dye (Fluo-4/AM) and a plate reader fluorim-
eter.13 The results of the functional assay for selected
compounds are summarized in Table 1. Selectivity to-
wards NR2A subunit containing NMDA receptors
was tested by the same functional assay using cells
expressing recombinant NR1/NR2A receptors and none
of the compounds exhibited significant activity up to
15 lM concentration. 6a was selected for further exam-
ination of selectivity towards other NR2 subunit con-
taining NMDA receptors. 6a up to 10 lM
concentration did not inhibit (inh. < 20%) NMDA
evoked current in NR1/NR2A and NR1/NR2D trans-
fected cells measured by standard patch clamp tech-
nique.14 In vivo analgesic activity was tested in the
mouse formalin test,15,16 a model of persistent pain.
Synthesis of targeted compounds 6a–9 is shown in
Scheme 1. Acylation of the corresponding o-nitro-ani-
lines 10–12 with ethyl chlorooxoacetate under basic con-
ditions (Et3N/CH2Cl2), followed by amidation with n-
butylamine, afforded the oxalic acid diamides 16–18.
Reduction of their nitro groups and debenzylation of
17 at the same time gave the phenylenediamines 19–21.
The benzimidazole ring was obtained by thermal con-
densation of 19–21. Hydrolysis of amides 22–24, simulta-
neous demethylation of 22 and nitration of 27 readily
produced the corresponding substituted benzimidazole-
2-carboxylic acids 25–28, which were coupled with vari-
ous piperidine derivatives to give the desired compounds
6a–7, 29. Nitrobenzimidazole-carboxamide 29 was trans-
formed via the amine derivative 8 to methanesulfona-
mide 9 with standard procedures. For the preparation
of 4-(substituted-benzyl)piperidines a recently published
procedure was used.10 The IR, 1H NMR, 13C NMR and
MS spectra for all intermediates and final compounds
were consistent with the assigned structures. Purity of
the samples was checked by HPLC and HRMS analysis.
Majority of 5(6)-OH-benzimidazole-2-carboxamide
derivatives showed excellent activities. Surprisingly
modifications of Y and Z did not influence significantly
the potency of compounds. The meta (6d–6f) and para
(6b, 6c) position of Z substituents resulted in nearly
equipotent compounds, even the ortho (6g) position
was well tolerated. A slightly reduced potency could
be observed in case of para Me (6c) and meta F (6d)
derivatives. The phenoxy-piperidines (6h–6j) were the
most effective compounds of the series. Replacement
of OH group by NH2 (8) led to loss of activity, whereas
the corresponding mesylamino derivative (9) showed
high activity. IC50 value of 9 was 6-fold lower than that
of 2 while having the same affinity towards NR2B recep-
tors. The 4(7)-OH isomer of 6a (7) was practically equi-
potent with it.
Biological activity of the prepared compounds was mea-
sured in a binding assay on rat forebrain membrane
using tritiated Ro 25-6981 as radioligand5,11,12 and in
Compound 6a had the best oral efficacy in formalin test
(ED50: 1.6 mg/kg po).
NO2
NO2
NH2
NO2
NH2
i
iii
iv
ii
X
X
X
X
NH
C
NH
C
NH
C
O
O
O
O
C
O
C
O
C
OEt
NH
NH
nBu
nBu
10 X = 4-MeO
11 X = 3-BnO
12 X = H
13 X = 4-MeO
14 X = 3-BnO
15 X = H
16 X = 4-MeO
17 X = 3-BnO
18 X = H
19 X = 4-MeO
20 X = 3-OH
21 X = H
Y
N
N
N
v
vii
Z
X
X
X
N
NHnBu
OH
N
N
H
N
H
H
O
O
O
22 X = 5(6)-MeO
23 X = 4(7)-OH
24 X = H
6 a-g
7
29
25 X = 5(6)-OH
26 X = 4(7)-OH
27 X = H
X = 5(6)-OH
X = 4(7)-OH
X = 5(6)-NO2
iii
vi
28 X = 5(6)-NO2
8
9
X = 5(6)-NH2
X = 5(6)-MesNH
viii
Scheme 1. Reagents and conditions: (i) ClCOCOOEt, Et3N, CH2Cl2, 0 °C–rt; (ii) n-BuNH2, toluene, rt, 10 h; (iii) H2, 10% Pd/C, MeOH, rt; (iv)
240 °C, 10 min; (v) 48% HBr, reflux, 48 h; (vi) H2SO4–HNO3; (vii) 4-substituted piperidine, HBTU, Et3N, DMF; (viii) MesCl, pyridine, CH2Cl2,
0 °C–rt.

4640
I. Borza et al. / Bioorg. Med. Chem. Lett. 16 (2006) 4638–4640
Table 1. Receptor binding and functional assay results for compounds 6a–9
Y
N
Z
X
N
N
H
O
NMDA-evoked D[Ca2+
]
IC50 (nM)
n [3H]Ro 25-6981
bindinga IC50
(nM)
n
a,b
Compound
X
Y
Z
Mp (°C)
i
6a
6b
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
5(6)-OH
4(7)-OH
5(6)-NH2
5(6)-MsNH
CH2
CH2
CH2
CH2
CH2
CH2
CH2
O
H
4-F
90–92
99–100
158–160
92–94
2.2 0.4
1.2
4
5
4
2
2
3
3
2
3
2
2
2
2
3
2
2
3
3
3
3
3
4
4.4 1.2
5.2 1.9
6c
6d
4-Me
3-F
8.7 1.0
6.0 1.1
2.4 0.5
2.5 0.6
12.0 2.8
1.1 0.2
1.7 0.4
2.9 0.6
3.0 0.4
40 0.4
1.3 0.2
8.6 1.7
191 10
5
3
6e
3-Me
3-MeO
2-Me
4-Me
4-Cl
4-F
95–96
86–87
74–76
4
5
3.2 1.1
6f
6g
3
10
4
8.4
3
6h
6i
115–117
194–195
147–151
165-166
74–75
O
6
14
6
6j
O
3
7
CH2
CH2
CH2
H
4
10
143
3
4
8
H
3
9
H
222–223
4
2
5
3.5 1.1
196 43
4
2
5
18
4
2
12
6
2
1
1
Ro 25-6981
CI-1041
159 29
6.6 1.2
3
3
4
a Values represent means SEM. The number of experiments (n) is indicated.
b NMDA-evoked changes of intracellular Ca2+
.
ward, R. M.; Keana, J. F. J. Med. Chem. 1999, 42, 3412–
3420.
In summary, a series of new benzimidazole-2-carboxam-
ide derivatives were prepared and found to be potent
and selective antagonists of the NR2B subtype of
NMDA receptors. Modifications of Y and Z (6a–6j),
transfer of the OH group from the 5(6) position to the
4(7) position and replacement of phenolic OH with
mesylamino group did not influence significantly the
activity of the compounds. Results of further SAR stud-
ies and detailed biological data will be published under
separate cover.
´
9. Borza, I.; Kolok, S.; Gere, A.; Agai-Csongor, E.;
´
´
Agai, B.; Tarkanyi, G.; Horvath, Cs.; Barta-Szalai, G.;
´
Bozo, E.; Kiss, Cs.; Bielik, A.; Nagy, J.; Farkas, S.;
´
´
´
´
Domany, Gy. Bioorg. Med. Chem. Lett. 2003, 13,
´
3859–3861.
´
´
´
´
´
10. Agai, B.; Proszenyak, A.; Tarkanyi, G.; Vida, L.; Faigl, F.
Eur. J. Org. Chem. 2004, 2004, 3623–3632.
11. Mutel, V.; Buchy, D.; Klingelschmidt, A.; Messer, J.;
Bleuel, Z.; Kemp, J. A.; Richards, J. G. J. Neurochem.
1998, 70, 2147–2155.
12. Richards, J. G.; Messer, J.; Buchy, D.; Klingel-schmidt,
A.; Mutel, V. Br. J. Pharmacol 1997, 120, Proc. Suppl.,
39P.
References and notes
}
13. Nagy, J.; Boros, A.; Dezso, P.; Kolok, S.; Fodor, L.
Neurochem. Int. 2003, 43, 19–29.
1. Chazot, P. L. Curr. Med. Chem. 2004, 11, 389–396.
2. Chenard, B. L.; Menniti, F. S. Curr. Pharm. Des. 1999, 5,
381–404.
3. Nikam, S. S.; Meltzer, L. T. Curr. Pharm. Des. 2002, 8,
845–855.
}
14. Kurko, D.; Dezso, P.; Boros, A.; Kolok, S.; Fodor, L.;
Nagy, J.; Szombathelyi, Zs. Neurochem. Int. 2005, 46,
369–379.
15. Peak effect of pain-related behaviour (licking/biting of
paw) elicited by formalin injection in mice 20–25 min post-
formalin was measured. Inhibition of this late phase
response is considered as analgesic effect against chemi-
cally induced persistent pain.Fasted male NMRI mice
(20–25 g, Charles River Hungary) were po treated with
vehicle (5% Tween 80) or test drugs (in dose-range:
6a = 0.5–8 mg/kg, 6c = 2–32 mg/kg) 15 min pre-formalin.
Then 20 ml of 1% formalin was s.c. injected into the right
hindpaw. Animals were visually observed in an open glass
cylinder.Percent decrease in licking time was determined
for each dose in comparison with the daily vehicle-treated
groups. Statistical significance was tested by Mann–
Whitney U-test. ED50 values were calculated by Boltz-
mann’s sigmoidal fitting.
4. McCauley, J. A. Expert Opin. Ther. Patents 2005, 15, 389–
407.
5. Fischer, G.; Mutel, V.; Trube, G.; Malherbe, P.; Kew, J.
N.; Mohacsi, E.; Heitz, M. P.; Kemp, J. A. J. Pharmacol.
Exp. Ther. 1997, 283, 1285–1292.
6. Wright, J. L.; Kesten, S. R.; Upasani, R. B.; Lan, N. C.
PCT Int. Appl. WO2000000197-A1. 2000; Chem. Abstr.
2000, 132, 64255.
7. McCauley, J. A.; Theberge, C. R.; Romano, J. J.; Billings,
S. B.; Anderson, K. D.; Claremon, D. A.; Freidinger, R.
M.; Bednar, R. A.; Mosser, S. D.; Gaul, S. L.; Connolly,
T. M.; Condra, C. L.; Xia, M.; Cunningham, M. E.;
Bednar, B.; Stump, G. L.; Lynch, J. J.; Macaulay, A.;
Wafford, K. A.; Koblan, K. S.; Liverton, N. J. J. Med.
Chem. 2004, 47, 2089–2096.
16. Hunskaar, S.; Fasmer, O. B.; Hole, K. J. Neurosci. Meth.
1985, 14, 69–76.
8. Tamiz, A. P.; Cai, S. X.; Zhou, Z. L.; Yuen, P. W.;
Schelkun, R. M.; Whittemore, E. R.; Weber, E.; Wood-