Synthesis of 1,5-diarylhaloimidazole analogs and their inhibitory activities against PGE2 production from LPS-treated RAW 264.7 cells

Article abstract of DOI:10.1016/j.bmc.2012.09.014

A number of 1,5-diarylimidazole analogs were synthesized and evaluated their inhibitory activities of cyclooxygenase-2 catalyzed prostaglandin E ₂ production. Reactions of 1,5-diarylimidazoles with halogenating reagents (NCS, NBS, NIS) afforded halogenated analogs. Among the analogs tested, compounds Ib, IIa, IIb and IIe exhibited significantly improved inhibitory activities against COX-2-mediated PGE₂ production from LPS-induced RAW 264.7 cells compared to those of the parent 1,5-diarylimidazoles. Especially, the analogs Ib (IC₅₀ = 0.55 μM) and IIa (IC ₅₀ = 0.58 μM) showed best results. Halogenation on the 1,5-diarylimidazole ring enhanced inhibitory activities against COX-2 catalyzed PGE₂ production, however, inhibitory activities were significantly varied by position(s) and species of the substituted halogen(s).

Full text of DOI:10.1016/j.bmc.2012.09.014

Bioorganic & Medicinal Chemistry 20 (2012) 6256–6259
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Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis of 1,5-diarylhaloimidazole analogs and their inhibitory activities
against PGE2 production from LPS-treated RAW 264.7 cells
Zunhua Yang a, Tuyen N. Truong a,c, Tuan-Anh N. Pham a, Jae-Won Lee b, Sung-Soo Kim b, Haeil Park a,
⇑
a College of Pharmacy, Chuncheon 200-701, Republic of Korea
b College of Medicine of Kangwon National University, Chuncheon 200-701, Republic of Korea
c College of Pharmacy, Ho Chi Minh City University of Medicine & Pharmacy, Ho Chi Minh City, Viet Nam
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 14 July 2012
Revised 6 September 2012
Accepted 7 September 2012
Available online 16 September 2012
A number of 1,5-diarylimidazole analogs were synthesized and evaluated their inhibitory activities of
cyclooxygenase-2 catalyzed prostaglandin E2 production. Reactions of 1,5-diarylimidazoles with haloge-
nating reagents (NCS, NBS, NIS) afforded halogenated analogs. Among the analogs tested, compounds Ib,
IIa, IIb and IIe exhibited significantly improved inhibitory activities against COX-2-mediated PGE2
production from LPS-induced RAW 264.7 cells compared to those of the parent 1,5-diarylimidazoles.
Especially, the analogs Ib (IC50 = 0.55 lM) and IIa (IC50 = 0.58 lM) showed best results. Halogenation
Keywords:
Halogenated 1,5-diarylimidazoles
COX-2
Inhibitory activity
PGE2 production
Anti-inflammation
on the 1,5-diarylimidazole ring enhanced inhibitory activities against COX-2 catalyzed PGE2 production,
however, inhibitory activities were significantly varied by position(s) and species of the substituted
halogen(s).
Ó 2012 Elsevier Ltd. All rights reserved.
1. Introduction
1,5-diarylimidazole made success and was pursued in further
development. Our previous results related with 1,5-diarylimidaz-
Several chemicals and biologicals such as eicosanoids and tissue
degradation enzymes are involved in inflammatory process.
Cyclooxygenase (COX), is the key proinflammatory enzyme which
catalyses the conversion of arachidonic acid to prostaglandins
(PGs). Cyclooxygenase exists in two isoforms. Cyclooxygenase-1
(COX-1) is a constitutive enzyme processing homeostasis function,
while cyclooxygenase-2 (COX-2) is an inducible one and known as
a major isoform found in inflammatory lesions.1
During the past two decades, efforts have been focused on the
development of selective COX-2 inhibitors. Modification of the
central heterocycle in tricyclic series has been a popular area of
research. As a result, diverse heterocycles have been explored.2 It
has been observed from previous SAR studies that COX-2 selective
inhibitors require a 4-methylsulfonylphenyl or a 4-sulfonamid-
ophenyl group attached to an unsaturated ring system in which
an additional vicinal lipophilic moiety is needed to possess good
activity and selectivity (Fig. 1).
oles also revealed very similar results.6,8 Other important SAR
results were observed from a precedent literature,4 the analogs
with halogen substitution at the 2- or 4-position of 1,5-diarylimi-
dazoles exhibited significantly improved biological activity. These
results led us to conduct SAR study of 1,5-diarylimidazoles haloge-
nated at 2- or/and 4-position(s). To decipher halogen effects on
COX-2 catalyzed PGE2 production from LPS-induced RAW 264.7
cell, two analogs (2,4-difluorophenyl or 3,4-difluorophenyl at N1,
4-methylsulfonylphenyl at C5; Fig. 2) were selected for further
investigation based on our previous results.8 We planned to
observe the result whether the halogen substitutions enhance
the moderate inhibitory activity of the parent compounds (84%
and 55% at 10 lM). In this study, several halogenated 1,5-diarylim-
idazoles at 2- or/and 4-position(s) were synthesized and evaluated
their inhibitory activities against COX-2 catalyzed PGE2 production
from LPS-induced RAW 264.7 cells.
Also it was well acknowledged that the nature of central scaf-
fold is very important for activity. Therefore, choice of the central
scaffold is crucial for activity as well as selectivity. As central scaf-
fold, diverse regioisomeric imidazoles such as 1,2-,3 1,5-,4–8 4,5-9,10
and 2,4- diarylimidazoles11 have been explored. However, only
2. Chemistry & pharmacology
2.1. Synthesis of 1,5-diarylimidazole analogs
Halogenated 1,5-diarylimiazole analogs were prepared following
the procedures and conditions as shown in Scheme 1. 4-Halo- and
2,4-dihalo-1,5-diarylimidazoles (Ib, Ic, IIc, IId, IIe) were prepared
by reacting the 1,5-diarylimidazoles I and II with halogenating
⇑
Corresponding author. Tel.: +82 33 250 6920; fax: +82 33 255 7865.
E-mail address: haeilp@kangwon.ac.kr (H. Park).
0968-0896/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2012.09.014

Z. Yang et al. / Bioorg. Med. Chem. 20 (2012) 6256–6259
Cl
6257
CHF2
CF3
N
Cl
O
CH3
N
N
CH3
N
N
N
N
N
F
N
H3CO
SO2NH2
SO2NH2
H3C
SO2CH3
SO2NH2
H3C
SO2NH2
Valdecoxib
Cimicoxib
Lumiracoxib
Etoricoxib
Celecoxib
Figure 1. Chemical structures of known COX-2 selective inhibitors.
kit for PGE2 (Cayman Chem. Co.) according to the manufacturer’s
recommendation. Cell viability was assessed with MTT assay. All
tested compounds showed no or less than 10% reduction of MTT
assay at the tested concentrations, indicating that they were not
significantly cytotoxic to RAW 264.7 cells in the presence or ab-
sence of LPS. Therefore, the inhibition of PGE2 production by 1,5-
diarylimidazoles might be not associated with their cytotoxicity.
F
F
F
R1
F
R1
N
N
N
N
SO2CH3
SO2CH3
R2
R2
II ~ IIe
I ~ Ic
3. Results and discussion
Figure 2. 1,5-Diarylimidazoles (I and II) and their halogenated derivatives.
Halogenated 1,5-diarylimidazole analogs on the imidazole ring
were prepared via two different synthetic pathways. Mono-substi-
tution (Br, I) at 2-position increased the inhibitory activity of PGE2
production (IIa and IIb), whereas 2-chlorination decreased inhibi-
tory activity of PGE2 production (Ia) compared to those of the par-
ent compounds. Di-bromination at 2- and 4-positions decreased
the inhibitory activity of PGE2 production (Ic), but di-chlorination
increased the inhibitory activity of PGE2 production (Ib and IIe)
compared to those of the parent compounds. To monitor the rela-
tionship between physicochemical properties (electron density/
lipophilicity) and biological activity of 1,5-diarylhaloimidazole
analogs, pKa and logP values of each compound were obtained
and compared with bioactivity data. The active compounds Ib(p-
Ka = 1.07) and IIa(pKa = 4.05) were observed to possess different
electronic density. To observe the effects of lipophilicity, logP val-
ues of active compounds (Ib = 3.64; IIa = 3.70; IIb = 3.88; IIe = 4.44)
and inactive compounds (Ic = 4.64; IId = 3.72) were compared.
Thus, no relationship between electron density/lipophilicity and
bioactivity was observed. However, in some cases, halogenation
on the 1,5-diarylimidazole ring enhanced inhibitory activities of
synthesized imidazole analogs against COX-2 catalyzed PGE2 pro-
duction. In detail, inhibitory activities were significantly varied
by halogenation position(s) and species of the substituted halo-
gen(s) in some cases as shown in Table 1. Two analogs exhibited
much stronger inhibitory activities of PGE2 production (Ib/
R
R
R1
N
NCS/NBS
reflux
N
N
N
SO2CH3
SO2CH3
R2
(R1, R2 = H, Cl, Br)
(R = 2,4-F2, 3,4-F2)
R
R
1. NCS/NBS/NIS
LiHMDS/-20 °C
R1
N
N
N
N
2. mCPBA
SO2CH3
SCH3
(R1 = Cl, Br, I)
(R = 2,4-F2, 3,4-F2)
Scheme 1. Synthetic pathway for halogenated 1,5-diarylimidzole derivatives.
reagents (NCS, NBS) in CHCl3 at reflux condition for 2–5 h gave
4-halo and 2,4-dihalo products.12,13 The halogenation reactions
with NIS in reflux conditions gave decomposed products.
2-Halo-1,5-diarylimidazoles (Ia, IIa, IIb) were prepared from
(methylthiophenyl)imidazoles in two steps. Reaction of (methylthi-
ophenyl)imidazoles, halogenating reagents (NCS, NBS, NIS) and lith-
ium hexamethyldisilazide (LiHMDS) in THF followed by oxidation
with m-chloroperbenzoic acid (mCPBA) provided 2-halogeno-1,5-
diarylimidazoles in crude form. Purification of the crude products
by column chromatography yielded pure 2-halogeno-1,5-diarylim-
idazole analogs.
Table 1
Yields of each compounds and % inhibitory activities of halogenated 1,5-diarylimi-
dazoles against COX-2 catalyzed PGE2 production from LPS-induced RAW 264.7 cellsa
Compounds
R1/R2
Yields (%)
% Inhibitionb,c (IC50
lM)
I
R1 = R2 = H
84 (6.52)
Ia
Ib
Ic
R1 = Cl,R2 = H
60
80
55
42
98 (0.55)
15
55
R
R
R
R
R
R
R
R
1 = R2 = Cl
1 = R2 = Br
1 = R2 = H
1 = Br, R2 = H
1 = I, R2 = H
1 = H, R2 = Cl
1 = H, R2 = Br
1 = R2 = Cl
2.2. RAW 264.7 cell culture and measurement of PGE2
concentrations
II
IIa
IIb
IIc
IId
IIe
40
85
55
50
74
99 (0.58)
90
56
7
RAW 264.7 cells obtained from American Type Culture Collec-
tion (ATCC, Rockville, MD) were cultured in DMEM supplemented
with 10% FBS and 1% antibiotics under 5% CO2 at 37 °C based on the
previously described procedures.14 Briefly, cells were plated in 96-
well plates (2 Â 105 cells/well). After pre-incubation with the test
81
a
All compounds were treated at 10
l
M. Treatment of LPS to RAW cells increased
M.
PGE2 production (10.0 M) from the basal level of 0.5
l
l
b
% Inhibition = 100 Â [1À(PGE2 of LPS with the flavones treated groupÀPGE2 of
the basal)/(PGE2 of LPS treated groupÀPGE2 of the basal)].
compounds for 1 h, LPS (1 lg/mL) were added and incubated for
c
All data are the arithmetic means (n = 3).
24 h. PGE2 concentration in the medium was measured using ELISA

6258
Z. Yang et al. / Bioorg. Med. Chem. 20 (2012) 6256–6259
IC50 = 0.55
l
M and IIa/0.58
l
M) than those of the parent com-
4.5. 1-(3,4-Difluorophenyl)-4-chloro-5-(4-methylsulfonyl
phenyl)imidazole (IIc)
pounds (I and II). Our present results imply that structural modifi-
cation at the imidazole ring system plays very important roles to
possess strong bioactivity. Further SAR study with large number
of halogenated imidazole analogs is under progress.
55%; White solid; mp 191–192 °C; Rf = 0.50 (hexane:ace-
tone = 1:1); 1H NMR (200 MHz, CDCl3) d 7.94–7.88 (m, 2H, Ar-H),
7.63 (s, 1H, H2-imidazole), 7.44–7.36 (m, 2H, Ar-H), 7.30–7.17
(m, 1H, Ar-H), 7.09–6.98 (m, 1H, Ar-H), 6.94–6.88 (m, 1H, Ar-H),
3.07 (s, 3H, SO2CH3); HRMS (EI) m/z Calcd for C16H11ClF2N2O2S
[M]+ 368.0198. Found 368.0155.
4. Experimental
4.1. Materials and methods
4.6. 1-(3,4-Difluorophenyl)-4-bromo-5-(4-methylsulfonyl
phenyl)imidazole (IId)
All chemicals were obtained from commercial suppliers, and
used without further purification. All solvents used for reaction
were freshly distilled from proper dehydrating agent under nitro-
gen gas. All solvents used for chromatography were purchased
and directly applied without further purification. 1H/13C NMR
spectra were recorded on a Varian Gemini 2000 instrument
(200 MHz) spectrometer. Chemical shifts were reported in parts
per million (ppm) downfield relative to tetramethylsilane as an
internal standard. Peak splitting patterns were abbreviated as m
(multiplet), s (singlet), br s (broad singlet), d (doublet), br d (broad
doublet), t (triplet) and dd (doublet of doublets). Mass spectra were
recorded on a Autospec M363. Melting points were recorded on a
Fisher-Johns microscopic scale melting point apparatus. Analytical
thin-layer chromatography (TLC) was performed using commercial
glass plate with silica gel 60F254 purchased from Merck. Chromato-
graphic purification was carried out by flash chromatography using
Kieselgel 60 (230–400 mesh, Merck).
50%; White solid; mp 195–196 °C; Rf = 0.40 (hexane:ace-
tone = 1:1); 1H NMR (200 MHz, CDCl3) d 7.90 (d, J = 8.2 Hz, 2H,
Ar-H), 7.67 (s, 1H, H2-imidazole), 7.42 (d, J = 8.2 Hz, 2H, Ar-H),
7.29–7.16 (m, 1H, Ar-H), 7.06–6.96 (m, 1H, Ar-H), 6.92–6.84 (m,
1H, Ar-H), 3.08 (s, 3H, SO2CH3); HRMS (EI) m/z Calcd for
C16H11BrF2N2O2S [M]+ 411.9693. Found 411.9659.
4.7. 1-(3,4-Difluorophenyl)-2,4-dichloro-5-(4-methylsulfonyl
phenyl)imidazole (IIe)
74%; White solid; mp 201 °C; Rf = 0.65 (hexane:acetone = 1:1);
1H NMR (200 MHz, CDCl3) d 7.89–7.85 (m, 2H, Ar-H), 7.38–7.33
(m, 2H, Ar-H), 7.28–7.20 (m, 1H, Ar-H), 7.13–7.04 (m, 1H, Ar-H),
6.99–6.93 (m, 1H, Ar-H), 3.05 (s, 3H, SO2CH3); HRMS (EI) m/z Calcd
for C16H10Cl2F2N2O2S [M]+ 401.9808. Found 401.9819.
4.2. General synthetic conditions for 4-halogeno and 2,4-
dihalogeno-1,5-diarylimidazoles
4.8. General synthetic conditions for 2-halogeno-1,5-
diarylimidazoles
4-Halogeno and 2,4-dihalogeno-1,5-diarylimiazole analogs
were prepared from 1,5-diarylimidazoles (I and II) following the
procedure as shown in Scheme 1. To the solution of I or II
(0.5 mmol) in CHCl3 (4 mL) was added NCS or NBS (0.75 mmol).
The mixture was refluxed for 5 h, extracted with DCM, washed
with aqueous NaHSO3 and brine and dried over anhydrous MgSO4
and concentrated. The residue was purified by silica gel column
chromatography to give 4-halogeno and 2,4-dihalogeno imida-
zoles. Reactions with excess halogenating reagents (3.0 equiv)
and extended reaction time (3–12 h) in chloroform or acetonitrile
exclusively provided 2,4-di-halogeno products.10,11 After the time
required in each case, the solvent was evaporated and ether was
added. The ethereal phase was washed with aqueous NaHSO3 solu-
tion, water, brine, dried with anhydrous MgSO4 and evaporated to
dryness. The residue was purified by column chromatography with
hexane:acetone as the mobile phase to get title products.
To the solution of (methylthiophenyl)imidazoles (0.4 mmol) in
THF (3 mL) was added, at À20 °C, LiHMDS (1 M in THF, 1.2 mL)
dropwise. The mixture was stirred for 0.5 h, then halo genating re-
agent (NCS, NBS or NIS) (1.6 mmol) in THF (3 mL) was added. The
mixture was stirred for 0.5 h at À20 °C and 6 h at room tempera-
ture. Aqueous NH4Cl was added to the mixture, and the reaction
mixture was extracted with EtOAc. The organic layer was washed
with aqueous NaHSO3 and brine, dried over anhydrous MgSO4
and concentrated. To the solution of crude intermediates (1 mmol)
in DCM (10 mL) was added, at 0 °C, m-chloroperbenzoic acid
(0.56 g, 2.5 mmol). The mixture was stirred for 2 h, added more
DCM, washed with aqueous Na2S2O3, NaHCO3 and brine and dried
over anhydrous MgSO4 and concentrated. The residue was purified
by silica gel column chromatography with hexane–EtOAc mixture
(8:1).
4.9. 1-(2,4-Difluorophenyl)-2-chloro-5-(4-methylsulfonyl
phenyl) imidazole (Ia)
4.3. 1-(2,4-Difluorophenyl)-2,4-dichloro-5-(4-methylsulfonyl
phenyl)imidazole (Ib)
60%; White solid; mp 183 °C; Rf = 0.55 (hexane:acetone = 1:1);
1H NMR (200 MHz, CDCl3) d 7.88 (d, J = 8.6 Hz, 2H, Ar-H), 7.59 (s,
1H, H4-imidazole), 7.40 (d, J = 8.6 Hz, 2H, Ar-H), 7.29–7.18 (m,
1H, Ar-H), 7.00–6.90 (m, 2H, Ar-H), 3.06 (s, 3H, SO2CH3); HRMS
(EI) m/z Calcd for C16H11ClF2N2O2S [M]+ 368.0198. Found 368.0184.
80%; White solid; mp 207 °C; Rf = 0.70 (hexane:acetone = 1:1);
1H NMR (200 MHz, CDCl3) d 7.87 (d, J = 8 Hz, 2H, Ar-H), 7.38 (d,
J = 8 Hz, 2H, Ar-H), 7.28–7.17 (m, 1H, Ar-H), 7.03–6.95 (m, 2H,
Ar-H), 3.05 (s, 3H, SO2CH3); HRMS (EI) m/z Calcd for
C
16H10Cl2F2N2O2S [M]+ 401.9808. Found 401.9882.
4.10. 1-(3,4-Difluorophenyl)-2-bromo-5-(4-methylsulfonyl
phenyl)imidazole (IIa)
4.4. 1-(2,4-Difluorophenyl)-2,4-dibromo-5-(4-methylsulfonyl
phenyl)imidazole (Ic)
40%; White solid; mp 169–170 °C; Rf = 0.45 (hexane:ace-
tone = 1:1); 1H NMR (200 MHz, CDCl3) d 7.83 (d, J = 8.4 Hz, 2H,
Ar-H), 7.36 (s, 1H, H4-imidazole), 7.33–7.22 (m, 1H, Ar-H), 7.24
(d, J = 8.4 Hz, 2H, Ar-H), 7.18–6.90 (m, 2H, Ar-H), 3.04 (s, 3H,
SO2CH3); HRMS (EI) m/z Calcd for C16H11BrF2N2O2S [M]+
411.9693. Found 411.9675.
55%; White solid; mp 239–240 °C; Rf = 0.55 (hexane:ace-
tone = 1:1); 1H NMR (200 MHz, CDCl3) d 7.87 (d, J = 8.4 Hz, 2H,
Ar-H), 7.40 (d, J = 8.4 Hz, 2H, Ar-H), 7.28–7.16 (m, 1H, Ar-H),
7.03–6.93 (m, 2H, Ar-H), 3.06 (s, 3H, SO2CH3); HRMS (EI) m/z Calcd
for C16H10Br2F2N2O2S [M]+ 489.8798. Found 489. 8725.

Z. Yang et al. / Bioorg. Med. Chem. 20 (2012) 6256–6259
6259
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