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After exploring the phenyl
moiety, we could conclude that
the best substitution pattern
was ortho-trifluoromethyl (com-
pound 7, IC50 =0.6 mm). There-
fore, derivatives of compounds
bearing altered eastern substitu-
tion were synthetized. Electron-
donating groups as in methoxy-
substituted 61 (IC50 =4.9 mm) ex-
hibited inhibitory activity; how-
ever, 52 (IC50 =1.9 mm), 55 (IC50 =
1.6 mm) and 58 (IC50 =2.7 mm)
Scheme 3. Synthesis of (E)-styryl-1H-benzo[d]imidazoles 73, 74 and 75 through introduction of aromatic residues
at position 3. Reagents and conditions: a) MeOH, H2SO4 (0.1 equiv), reflux, 16 h, 70%; b) R1ÀB(OH)2 (2.0 equiv),
Pd(PPh3)4 (0.1 equiv), 1m aq Na2CO3, toluene/EtOH (4:1), 858C, 16 h, 26–49%; c) KOH (2.85 equiv), MeOH, 258C,
3.5 h, 64–95%; d) EDC (1.5 equiv), DMAP (0.05 equiv), o-phenylenediamine (1.0 equiv), imidazole (0.05 equiv), RT,
4 h, 8–32%; e) 6m aq HCl, EtOH, 1208C, 10 min, MW, 34–63%.
bearing
electron-withdrawing
groups partially restored the ac-
tivity.
The structure–activity relation-
ship (SAR) of the investigated
styryl benzimidazoles on sEH are
using a synthetic epoxide as a substrate (3-phenyl-cyano(6-me-
thoxy-2-naphthalenyl)methyl ester-2-oxiraneacetic acid,
rather flat, which correlates with the possible binding mode
proposed by the molecular docking experiment of the most
potent compound 7 (Figure 1, see the Supporting Information
PHOME).[38] As shown in Table 1, we examined the different
substitutions at the phenyl ring of the styryl moiety (western
part) and the substitutions at the benzimidazole scaffold (east-
ern part). Regarding the western part of the molecule, several
compounds were synthesized bearing ortho, meta and para
substitutions. Ortho-substituted compounds (7, 10, 13, 16, 19)
yielded highest inhibitory activity, with the exception of
bromo-substituted 22. Electron-donating groups like methoxy
(compound 13) were less tolerated (IC50 =18.3 mm), and the
best inhibition was obtained with trifluoromethyl-substituted
compound 7 (IC50 =0.6 mm). Unsubstituted compound 3 result-
ed in a 10-fold loss of potency compared with the best com-
pound 7. With the exception of bromo-substituted 22
(IC50 =32.1 mm), halogen groups in the ortho position exhibited
moderate inhibition with IC50 values in the range of 1.7 to
2.6 mm .
Lower inhibitory activity was observed for 29 and 30, indi-
cating that meta substitution is less favored than ortho substi-
tution. However, we confirmed the tendency concerning the
electron-donating properties of the substituent, as compound
30 decreases the inhibitory activity (IC50 =5.8 mm). The intro-
duction of heterocyclic moieties, such as furane (73), pyridine
(74) or 2,3-dihydrobenzo[b][1,4]dioxine (75), increased the in-
hibitory potency (73: IC50 =1.2 mm, 74: IC50 =0.8 mm, 75: IC50 =
1.5 mm, Table 1). In contrast to the positive substituent effect
observed for ortho-substituted compounds, para-substitution
pattern resulted in a weaker inhibitory activity regardless of
the electron-donating nature of the substituent.
Figure 1. Docking pose of (E)-styryl-1H-benzo[d]imidazole 7 in the binding
pocket of the catalytic domain of sEH (PDB: 3KOO).
for details). Both nitrogen atoms of the benzimidazole core
seem to interact with the catalytic center of sEH (Tyr381,
Asp333, Tyr465). The largest part of the benzimidazole scaffold
is enclosed in the Trp334 niche (15 ꢃ) capable for aryl interac-
tions. The phenyl moiety showed p-stacking with Met419[39]
and His524, and the benzimidazole core showed p-stacking
with Trp334. The additional space around both aromatic cores
correlates with the high tolerability of different substituents, as
demonstrated by the SAR.
Additionally, double substitution at the phenyl moiety was
explored and led to a decrease in the inhibitory activity. 2,5-
Methoxy-substituted compound 43 (IC50 =17.3 mm) exhibited
the lowest inhibitory activity, which could be restored by a 3,4-
dimethoxy (46, IC50 =4.3 mm) or 3,5-dimethoxy substitution
pattern (2, IC50 =5.2 mm). The potency was not completely re-
stored by exchanging the methoxy groups to a 3,4-dichloro
pattern (49, IC50 =9.7 mm).
We evaluated the effect of compounds IPS (1; IC50 =
10.0 mm), 16 (IC50 =1.7 mm), 19 (IC50 =2.6 mm), 74 (IC50 =
1.2 mm) and 75 (IC50 =0.8 mm) on cell viability of several cancer
lines (Table 2). IPS inhibited the proliferation activity of all cell
lines except MCF-7. In the water-soluble tetrazolium salt (WST-
1) assay, EC50 values were in the range of 2.7 to 10.4 mm. Ortho-
substituted compounds 16 and 19 affected the proliferation
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ChemMedChem 2013, 8, 919 – 923 921