Synthesis and CYP24A1 inhibitory activity of N-(2-(1H-imidazol-1-yl)-2- phenylethyl)arylamides

Article abstract of DOI:10.1016/j.bmc.2010.06.011

A series of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides were prepared, using an efficient three- to five-step synthesis, and evaluated for their inhibitory activity against human cytochrome P450C24A1 (CYP24A1) hydroxylase. Inhibition ranged from IC₅₀ 0.3-72 μM compared with the standard ketoconazole IC₅₀ 0.52 μM, with the styryl derivative (11c) displaying enhanced activity (IC₅₀ = 0.3 μM) compared with the standard, providing a useful preliminary lead for drug development.

Full text of DOI:10.1016/j.bmc.2010.06.011

Bioorganic & Medicinal Chemistry 18 (2010) 4939–4946
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and CYP24A1 inhibitory activity
of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides
Ahmed S. Aboraia a, , Sook Wah Yee a, Mohamed Sayed Gomaa a, Nikhil Shah a, Anna C. Robotham b,
Bart Makowski b, David Prosser b, Andrea Brancale a, Glenville Jones b, Claire Simons a,
*
a Medicinal Chemistry, Welsh School of Pharmacy, Cardiff University, King Edward VII Avenue, Cardiff CF10 3NB, UK
b Department of Biochemistry, Queens University, Kingston, Ontario, Canada K7L 3N6
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides were prepared, using an efficient three- to
Received 31 March 2010
Revised 27 May 2010
Accepted 4 June 2010
Available online 9 June 2010
five-step synthesis, and evaluated for their inhibitory activity against human cytochrome P450C24A1
(CYP24A1) hydroxylase. Inhibition ranged from IC50 0.3–72
zole IC50 0.52 M, with the styryl derivative (11c) displaying enhanced activity (IC50 = 0.3
with the standard, providing a useful preliminary lead for drug development.
l
M compared with the standard ketocona-
l
l
M) compared
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
N-(2-(1H-Imidazol-1-yl)-2-
phenylethyl)arylamides
Benzofuran-2-carboxamides
CYP24A1
Enzyme inhibition
Molecular modelling
1. Introduction
molecule inhibitors of CYP24A1 have been described by our group6,7
and others,8–11 with the non-specific P450 inhibitor ketoconazole8
(Fig. 1) used as a standard for comparison. The most promising
CYP24A1 inhibitors described to date are VID-4009 (IC50 = 15.1 nM
compared with ketoconazole IC50 = 126 nM in human epidermal
Calcitriol (1a,25-(OH)2D3), the biologically active metabolite of
vitamin D binds to the vitamin D receptor (VDR), whichmediates tis-
sue-specific effects including enhancement of intestinal calcium
transport, maintenance of skeletal health and epidermal integrity
and immune cell regulation (e.g., macrophages).1 Replacement cal-
citriol therapy has been used in chronic kidney disease, hyperprolif-
erative skin diseases and osteoporosis, with clinical trials for various
forms of cancer and autoimmune conditions ongoing.2,3 Although
the potential of vitamin D as an antiproliferative drug has been real-
ised in psoriasis and parathyroid cell hyperplasia, an anticancer
treatment incorporating vitamin D remains a challenge owing to in-
creased drug resistance.3 Evidence is growing that this resistance is
caused by up-regulation of the cytochrome P450 enzyme, CYP24A1,
in tumours resulting in accelerated metabolism of calcitriol and
derivatives.4,5 Inhibitors of CYP24A1 are expected to extend the
half-life of calcitriol thereby enhancing endogenous levels of
calcitriol, or vitamin D analogues, which may result in stabilized
CYP24A1 and enhanced stability of vitamin D compounds. Small
keratinocytes) and the vitamin D derived sulfoximes10 (IC50
=
7.4 nM compared with ketoconazole IC50 = 312 nM in V79-CYP24A1
recombinant cell line expressing human CYP24A1), which are based
on the structure of the natural substrate, calcitriol.
Here we report the synthesis and inhibitory activity of a series
of N-(2-(1H-imidazol-1-yl)-2-phenylethyl)arylamides. The scaffold
of the benzofuran series (7) were based on VID-400, while the
phenylmethyl (11b) and styryl (11c) derivatives combined phar-
macophores of VID-400 and calcitriol.
2. Chemistry
The synthesis of N2-[2-phenyl-2-(1H-imidazoyl)ethyl]-ben-
zo[b]furan-2-carboxamide derivatives (7a–e), was achieved using
a five-step reaction (Scheme 1) and involved a modification of the
procedure described by Schuster and Egger12 and Moenius et al.13
The substituted ethyl benzo[b]furan-2-carboxylates (2) were
prepared by the reaction of the appropriate salicylaldehyde (1) with
ethyl bromoacetate in anhydrous DMF at 130 °C for 6 h according to
the methodology previously described.14 After purification by flash
chromatography or recrystallisation with petroleum ether, the
* Corresponding author. Tel.: +44 (0) 2920 876307; fax: +44 (0) 2920 874149.
E-mail address: SimonsC@Cardiff.ac.uk (C. Simons).
URL: http://www.cardiff.ac.uk/phrmy/staffinfo/CS/homepage/CS.html (Claire Si-
mons).
Present address: Medicinal Chemistry Department, Faculty of Pharmacy, Assiut
University, Assiut, Egypt.
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.06.011

4940
A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
N
N
O
O
S
O
O
N
N
O
NH
Cl
H
H
Cl
ketoconazole
H
Cl
N
HO
OH
N
H
N
24-sulfoximine vit D3 analogue
O
VID-400
Figure 1. Inhibitors of CYP24A1.
O
OH
NH2
5
5
H
R1
R1
R1
+
CO2Et
CO2H
6
6
O
O
R2
OH
4
1
2
3
N
N
OH
H
N
H
N
O
O
R2
Hx
R2
R2
O
O
N
O
R1
HA
HB
O
R1
R1
5
6
7
6
5
Scheme 1. Reagents and conditions: (i) ethyl bromoacetate, K2CO3, DMF, 130 °C, 6–7 h, 22–53%; (ii) 2 M NaOH (aq), CH3OH, rt, 20 min, 85–98%; (iii) (a) CDI, DMF, rt, 1 h;
(b) substituted 2-amino-1-phenyl-ethanol (4), rt, 12 h, 72–88%; (iv) (CH3SO2)2O, Et3N, 0 °C, 24 h, 16–67%; (v) imidazole, isopropyl acetate, 125 °C, 24 h, 28–72%.
products (2) were obtained in low to moderate yields (22–53%). Base
hydrolysis of the ethyl benzo[b]furan-2-carboxylates (2) gave the
substituted benzo[b]furan-2-carboxylic acid compounds (3) in good
yields (88–98%). The synthesis of the substituted N2-(2-hydroxy-2-
phenylethyl)-benzo[b]furan-2-carboxamides (5) was carried out
using 1,10-carbonyldiimidazole (CDI) with the corresponding substi-
tuted 2-amino-1-phenyl-ethanol (4) (Scheme 1). The products were
recrystallised from ethanol and ice water to give good purity and
yield (72–88%).
The substituted 2-benzo[b]furan-2-yl-5-phenyl-4,5-dihydro-
1,3-oxazoles (6) were prepared by the reaction of the amide-con-
taining compound (5) with methanesulfonic anhydride and a base
(triethylamine).15,16 The presence of the 4,5-dihydro-1,3-oxazole
ring structure was observed on the 1H NMR spectra as an ABX sys-
tem, which showed the signals of the three carbon-bound protons
of the 4,5-dihydro-1,3-oxazole system. The geminal pair, HA and
HB, are diastereotopic; they are coupled to each other with the
largest of the three coupling constants (ꢀ15 Hz) and they are each
coupled to HX, with different coupling constants, one large
(ꢀ10 Hz) and one small (ꢀ8 Hz). The HX signal (ꢀ5.8 ppm) was
downfield from the HA and HB signals (ꢀ4.5 and 4.0 ppm).
Overall, the synthesis of the oxazole compounds (6) gave moder-
ate yields after purification by column chromatography or recrystal-
lisation from ethanol. The isolated yield of compound 6c was very
low (16%), which may be owing to the presence of the electron
withdrawing nitro group affecting the ring closure.
In the last step of the reaction scheme, heating the oxazole
compound (6) in the presence of imidazole opened the oxazole ring
by nucleophilic displacement (Scheme 1)17 A reasonable yield of
compound (7a–e) was produced after purification by column chro-
matography and/or recrystallisation from ethanol/water (2:1 v/v).
In a similar manner 4-bromo-N-(2-(1H-imidazol-1-yl)-2-phen-
ylethyl)benzamide (11a), 4-benzyl-N-(2-(1H-imidazol-1-yl)-2-
phenylethyl)benzamide (11b), and (E)-4-N-(2-(1H-imidazol-1-yl)-
2-phenylethyl)styrylbenzamide (11c) were prepared (Scheme 2),
commencing from either commercially available or readily pre-
pared carboxylic acids (8), which on coupling with 2-amino-1-phe-
nyl–ethanol (4) gave the amides (9). The oxazoles (10) and final
imidazole products (11) were prepared as previously described
for the benzofuran series with good yields obtained for the oxaz-
oles (61–82%) and low to moderate yields obtained for the imida-
zoles (16–41%).

A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
4941
N
OH
N
CO2H
H
N
H
N
O
O
4
O
N
R
R
11
9
10
R
R
8
Scheme 2. Reagents and conditions: (i) (a) CDI, DMF, rt, 1 h; (b) 2-amino-1-phenyl-ethanol (4), rt, 12 h, 39–86%; (ii) (CH3SO2)2O, Et3N, 0 °C, 24 h, 71–82%; (iii) imidazole,
isopropyl acetate, 125 °C, 24 h, 16–41%.
Table 1
3. CYP24A1 inhibitory activity
IC50 data for the novel N-(2-(1H-imidazol-1-yl)-2-phenylethyl)aryl amides 7 and 11
N
The imidazole derivatives 7a–e and 11a–c were evaluated for
their calcitriol metabolism (CYP24A1) inhibitory activity using a cell
N
H
based assay employing a recombinant cell line expressing human
O
N
CYP24A1 enzyme (V79-CYP24),18 using radiolabelled [3H-1b]-calci-
triol as the substrate and ketoconazole as the standard for compar-
ison. In the benzofuran series (7), the 4-fluorophenyl (7d) and
unsubstituted (7a) derivatives were the most potent (IC50 = 2.8
R1
R2
Compound
R1
R2
H
IC50 (lM)
a
and 4.0 lM, respectively), with introduction of the larger chloro
group (7e) resulting in a small reduction in activity (Table 1). Incor-
poration of substituents in the benzofuran moiety was less favour-
7a
4.0
O
able (IC50 = 6.4
However all the benzofuran derivatives (7) were less active than
the standard ketoconazole (IC50 = 0.52 M) (Table 1). Replacing
the benzofuran group with either 4-bromobenzene (11a) or methy-
lenedibenzene (11b) resulted in a substantial loss of activity, how-
ever the introduction of the styryl moiety (11c) resulted in a
lM (5-NO2, 7c) and 36.9 lM (6-OCH3, 7b)).
7b
H
36.9
O
H3CO
O2N
l
7c
H
F
6.4
2.8
O
potent inhibitor (IC50 = 0.3 lM).
7d
O
O
4. Docking
7e
Cl
H
10.6
23
To investigate the possible binding mode of this series of com-
pounds, we have performed a set of molecular docking simula-
tions, using our model of the CYP24A1.19 In the benzofuran
analogues series (Fig. 2) it is possible to observe how the imidazole
N3 complexes the Haem–Fe, while the phenyl ring is placed in a
small hydrophobic pocked created by Ile242, Val391, Thr330, and
Ala326. The benzofuran ring is placed in a hydrophobic channel
and it establishes a series of interactions with Ile149, Pro392,
and Trp134. From these results it also appears that the amide func-
tion does not have a specific role in the binding, but acts just as a
linker between the two aromatic parts of the compounds.
The styryl analogue 11c docks in a very similar pose as 7a
(Fig. 3), but in this case the styryl group extends further in the
hydrophobic channel, establishing a contact with Phe104. Finally,
it should be noted that compound 11b does not dock successfully
in the CYP24A1 model, probably because of the steric hindrance of
the side chain, which cannot be properly placed in the narrow
hydrophobic channel.
11a
Br
11b
H
72
11c
H
—
0.3
Ketoconazole
—
0.52
a
IC50 values are the average ( 5%) of two experiments.
phy was performed with silica gel 60 (230–400 mesh) (Merck) and
TLC was carried out on precoated silica plates (kiesel gel 60 F254
,
BDH). Melting points were determined on an electrothermal instru-
ment and are uncorrected. Compounds were visualised by illumina-
tion under UV light (254 nm) or by the use of vanillin stain followed
by charring on a hotplate. All solvents were dried prior to use as de-
scribed by the handbook Purification of Laboratory Chemicals20 and
stored over 4 Å molecular sieves, under nitrogen.
5. Experimental
5.1. Materials and methods: chemistry
The ethyl benzo[b]furan-2-carboxylic acid derivatives (3) were
prepared according to the general procedure described by Suzuki
et al,14 compounds 2 and 3 (R1 = H) were previously reported.14
The 2-amino-1-phenyl-ethanol derivatives (4) were either avail-
able commercially (R = H) or prepared (R = F, Cl) according to the
method described by Cho et al.21
1H and 13C NMR spectra were recorded with a Brucker Avance
DPX500 spectrometer operating at 500 and 125 MHz, with Me4Si
as internal standard. Mass spectra were determined by the EPSRC
mass spectrometry centre (Swansea, UK). Microanalyses were
determined by Medac Ltd (Surrey, UK). Flash column chromatogra-

4942
A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
J = 8.7 Hz, 1H, H-4), 7.57 (d, J = 0.9 Hz, 1H, H-3), 7.17 (d,
J = 1.8 Hz, 1H, H-7), 7.03 (dd, J = 2.2, 8.7 Hz, 1H, H-5), 4.53 (q,
J = 7.1 Hz, 2H, CH2), 3.97 (s, 3H, OCH3), 1.52 (t, J = 7.1 Hz, 3H,
CH3). 13C NMR: d 160.9 (C, C-7a), 160.0 (C-6), 157.5 (C, C@O),
145.4 (C, C-2), 123.4 (CH, CH-4), 120.7 (C, C-3a), 114.5 (CH, CH-
3), 114.4 (CH, CH-5), 96.2 (CH, CH-7), 61.7 (CH2), 56.1 (CH3, –
OCH3), 14.8 (CH3). Anal. Calcd for C12H12O4 (220.223): C, 65.45;
H, 5.49. Found: C, 65.32; H, 5.53.
5.1.1.2. Ethyl 5-nitrobenzo[b]furan-2-carboxylate (2, R = 5-
NO2).
A yellow solid was obtained. Yield: 22%, Rf 0.60 (petro-
leum ether/EtOAc 4:1 v/v); mp 139–141 °C; 1H NMR: d 8.69 (d,
J = 2.3 Hz, 1H, H-4), 8.42 (dd, J = 2.3, 9.1 Hz, 1H, H-6), 7.75 (d,
J = 9.1 Hz, 1H, H-7), 7.70 (s, 1H, H-3), 4.53 (q, 2H, J = 7.1 Hz, CH2),
1.50 (t, 3H, J = 7.1 Hz, CH3). 13C NMR: d 159.0 (C, C-7a), 158.4
(C@O), 149.1 (C, C-2), 145.2 (C, C-5), 127.7 (C, C-3a), 123.3 (CH,
CH-6), 119.9 (CH, CH-4), 114.3 (CH, CH-3), 113.4 (CH, CH-7), 62.6
(CH2), 14.7 (CH3). Anal. Calcd for C11H9NO5 (235.194): C, 56.17;
H, 3.86; N, 5.95. Found: C, 56.38; H, 3.90; N, 5.88.
5.1.2. General procedure for the synthesis of ethyl
benzo[b]furan-2-carboxylic acid derivatives (3)
Figure 2. Proposed binding mode of compound 7a.
A solution of ethyl benzo[b]furan-2-carboxylate (2) (32 mmol)
in methanol (100 mL) was treated with 2 M aqueous sodium
hydroxide (50 mL) and warmed gently on a steam bath. After
hydrolysis was complete (ca. 30 min.), the reaction solution was
cooled and acidified to pH 1 by the dropwise addition of concen-
trated HCl. The solution was then extracted with diethyl ether
(3 ꢁ 150 mL) and the organic layer dried (MgSO4), filtered, and re-
duced in vacuo to give a crude yellow solid. The crude yellow solid
was washed with water, collected and dried under vacuum with
phosphorus pentoxide
5.1.2.1. 6-Methoxybenzo[b]furan-2-carboxylic acid (3, R = 6-
OMe).
Yield: 88%, Rf 0.0 (petroleum ether/EtOAc 1:1 v/v); mp
194–196 °C; 1H NMR (DMSO-d6): d 13.36 (s, 1H, COOH), 7.67 (d,
J = 8.7 Hz, 1H, H-4), 7.61 (s, 1H, H-3), 7.31 (s, 1H, H-7), 6.99 (split
d, J = 2.1, 8.6 Hz, H-5), 3.85 (s, 3H, OCH3). 13C NMR (DMSO-d6): d
160.4 (C@O and C, C-7a), 156.8 (C, C-6), 145.6 (C, C-2), 123.7 (CH,
CH-4), 120.3 (C, C3a), 114.2 (CH, CH-3), 114.1 (CH, CH-5), 96.2
(CH, CH-7), 56.1 (CH3, –OCH3). Anal. Calcd for C10H8O4 (192.169):
C, 62.50; H, 4.20. Found: C, 62.49; H, 4.19.
5.1.2.2. 5-Nitrobenzo[b]furan-2-carboxylic acid
(3, R = 5-
NO2). Yield: 95%, Rf 0.0 (petroleum ether/EtOAc 1:1 v/v); mp
268–270 °C; 1H NMR (DMSO-d6): d 14.00 (s, 1H, COOH), 8.75 (d,
J = 2.2 Hz, 1H, H-4), 8.36 (dd, J = 2.2, 9.2 Hz, 1H, H-4), 7.97 (d,
J = 9.2 Hz, 1H, H-7), 7.85 (s, 1H, H-3). 13C NMR (DMSO-d6): d
159.8 (C, C-7a), 157.9 (C@O), 149.4 (C, C-5), 144.5 (C, C-2), 127.8
(C, C-3a), 123.1 (CH, CH-6), 120.2 (CH, CH-4), 114.5 (CH, CH-3),
113.6 (CH, CH-7). Anal. Calcd for C9H5NO5 (207.140): C, 52.19; H,
2.43; N, 6.76. Found: C, 52.30; H, 2.43; N, 6.68.
Figure 3. Proposed binding mode of compound 11c.
5.1.1. General procedure for the synthesis of ethyl benzo[b]
furan-2-carboxylate derivatives (2)
Activated potassium carbonate (16.55 g, 120 mmol) was added
to a stirred solution of salicylaldehyde (1) (60 mmol) in anhydrous
DMF (15 mL). Ethyl bromoacetate (10.0 g, 60 mmol) in anhydrous
DMF (20 mL) was then added to the above mixture and the reac-
tion was stirred at 130 °C for 6 h. The solvent was then evaporated
to about 1/3 its volume to give a brown syrup. The crude product
was extracted with CH2Cl2 (200 mL) and water (3 ꢁ 100 mL). The
organic layer was dried (MgSO4), filtered and reduced in vacuo
and the resulting residue purified by flash column chromatography
(petroleum ether/ethyl acetate 100:0 v/v increasing to 95:5 v/v)
5.1.3. General procedure for the synthesis of N2-(2-hydroxy-2-
phenylethyl)aryl-2-carboxamide derivatives (5) and (9)
To a suspension of carboxylic acid (3 or 8) (28 mmol) in anhy-
drous DMF (30 mL) was added 1,10-carbonyldiimidazole (28 mmol)
and the reaction stirred at room temperature for 1 h. The reaction
was cooled to 0 °C and subsequently combined with a solution of
2-amino-1-phenyl-ethanol (4) (28 mmol) in DMF (10 mL). The
mixture was stirred at room temperature for 8 h. After the reaction
was complete, an aliquot amount of ice was poured into the flask
to precipitate out the product, which was then filtered and washed
with 10 mL of ethanol and dried under vacuum with phosphorus
pentoxide.
5.1.1.1. Ethyl 6-methoxybenzo[b]furan-2-carboxylate (2, R = 6-
OMe).
A brown solid was obtained. Yield: 53%, Rf 0.55 (petro-
leum ether/EtOAc 3:1 v/v); mp 66–68 °C; 1H NMR: d 7.63 (d,

A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
4943
5.1.3.1. N2-(2-Hydroxy-2-phenylethyl)benzo[b]furan-2-carbox-
amide (5a). A white solid was obtained. Yield: 72%, Rf 0.45
aqueous ethanol. Yield: 86%, Rf 0.8 (petroleum ether/EtOAc 3:2 v/
v); mp 188–190 °C; 1H NMR (DMSO-d6): d 8.61 (t, J = 5.2 Hz, 1H,
NH), 7.79 (dd, J = 2.0, 8.5 Hz, 2H, Ar), 7.36 (m, 2H, Ar), 7.26 (m,
5H, Ar), 5.51 (d, J = 4.5 Hz, 1H, OH), 4.78 (ddd, J = 4.6, 6.6, 1H, CH-
OH), 3.48 (ddd partially obscured, J = 5.8, 10.9 Hz, 2H, CH2). 13C
NMR (DMSO-d6): d 165.4 (C@O), 143.7 (C), 133.7 (C), 131.2,
131.2, 129.3, 129.3, 128.0, 128.0, 127.0, 125.49, 125.9 (9 ꢁ CH,
Ar), 124.8 (C), 71.0 (CH–OH), 47.7 (CH2). Anal. Calcd for
(petroleum ether/EtOAc 3:2 v/v); mp 140–142 °C; 1H NMR
(DMSO-d6): d 8.67 (t, J = 5.6 Hz, 1H, NH), 7.78 (d, J = 7.7 Hz, 1H,
Ar), 7.67 (d, J = 8.3 Hz, 1H, Ar), 7.56 (s, 1H, H-3), 7.47 (m, 1H, Ar),
7.41–7.24 (m, 5H, Ar), 5.60 (d, J = 4.4 Hz, 1H, OH), 4.83 (m, 1H, H-
1), 3.58–3.34 (m, 2H, CH2). 13C NMR (DMSO-d6): d 158.5 (C@O),
154.5 (C), 149.5 (C), 143.9 (C), 128.4, 128.4 (2 ꢁ CH), 127.5 (C),
127.4, 127.1, 126.4, 126.4, 124.0, 123.1, 112.1, 109.7 (8 ꢁ CH),
71.3 (CH–OH), 47.4 (CH2). Anal. Calcd for C17H15NO3 (281.309): C,
72.58; H, 5.37; N, 4.98. Found: C, 72.44; H, 5.32; N, 4.88.
C15H14BrNO2 (320.18): C, 56.27; H, 4.41; N, 4.37. Found: C, 56.26;
H, 4.38; N, 4.31.
5.1.3.7.
(9b).
4-Benzyl-N-(2-hydroxy-2-phenylethyl)benzamide
A white solid was obtained after recrystallisation from
5.1.3.2.
furan-2-carboxamide (5b).
N2-(2-Hydroxy-2-phenylethyl)-6-methoxybenzo[b]
A light brown powder was ob-
aqueous ethanol. Yield: 39%, Rf 0.8 (petroleum ether/EtOAc 1:1 v/
v); mp 98–99 °C; 1H NMR (DMSO-d6): d 8.28 (t, J = 5.5 Hz, 1H,
NH), 7.15–7.39 (m, 14H, Ar), 5.48 (d, J = 4.5 Hz, 1H, OH), 4.76 (dd,
J = 5.2, 11.9 Hz, 1H, CH-OH), 4.03 (d, J = 2.2 Hz, 2H, Ar-CH2-Ar),
4.38 (m, 1H, CH2CH(OH)). 13C NMR (DMSO-d6): d 169.3 (C@O),
143.5, 141.0, 138.9, 136.8 (4 ꢁ C), 130.1, 129.4, 128.9, 128.9,
128.4, 128.4, 128.2, 128.2, 128.0, 127.2, 127.1, 126.1, 125.8, 125.8
(14 ꢁ CH, Ar), 71.25 (CH, CH-OH), 47.05 (CH2), 37.7 (CH2). Anal.
Calcd for C21H19NO2 (317.38): C, 79.73; H, 6.39; N, 4.22. Found:
C, 79.67; H, 6.41; N, 4.28.
tained. Yield: 88%, Rf 0.40 (petroleum ether/EtOAc 3:2 v/v); mp
147–149 °C; 1H NMR (DMSO-d6): d 8.45 (t, J = 5.7 Hz, 1H, NH),
7.62 (d, J = 8.7 Hz, 1H, Ar), 7.46 (d, J = 0.8 Hz, 1H, H-3), 7.39–7.30
(m, 4H, Ar), 7.27–7.20 (m, 2H, Ar), 6.95 (dd, J = 2.2, 8.6 Hz, 1H,
Ar), 5.58 (d, J = 4.4 Hz, 1H, OH), 4.79 (m, 1H, H-1), 3.82 (s, 3H,
OCH3), 3.55–3.30 (m, 2H, CH2). 13C NMR (DMSO-d6): d 159.8 (C),
158.6 (C@O), 155.9 (C), 148.7 (C), 143.9 (C), 128.4, 128.4, 127.5,
126.4, 126.4, 123.3 (6 ꢁ CH), 120.7 (C), 113.5 (CH), 109.8 (CH),
96.3 (CH), 71.5 (CH–OH), 56.1 (OCH3), 47.3 (CH2). Anal. Calcd for
C18H17NO4 (311.335): C, 69.44; H, 5.50; N, 4.50. Found: C, 69.47;
H, 5.46; N, 4.37.
5.1.3.8.
(9c).
N
-(2-Hydroxy-2-phenylethyl)-4-styrylbenzamide
A white solid was obtained after recrystallisation from
5.1.3.3. N2-(2-Hydroxy-2-phenylethyl)-5-nitrobenzo[b]furan-2-
ethanol. Yield: 71%, Rf 0.31 (petroleum ether/EtOAc 1:1 v/v); mp
218–219 °C; 1H NMR (DMSO-d6): d 8.52 (s, 1H, NH), 7.88 (d,
J = 8.0 Hz, 2H, Ar), 7.69 (d, J = 8.0 Hz, 2H, Ar), 7.64 (d, J = 7.5 Hz,
2H, Ar, 7.42–7.25 (m, 10H, Ar), 5.53 (d, J = 4.2 Hz, 1H, OH), 4.82
(m, 1H, CH-OH), 3.52 (m, 1H, CH2), 3.37 (m, 1H, CH2). 13C NMR
(DMSO-d6): d 166.0 (C@O), 143.8, 139.7, 136.8, 133.3 (4 ꢁ C),
130.1, 128.7, 128.7, 128.0, 128.0, 127.7, 127.7, 127.5, 127.5,
127.0, 126.7, 126.7, 126.2, 126.2, 126.0, 126.0 (16 ꢁ CH, Ar), 71.2
(CH, CH-OH), 47.7 (CH2). Anal. Calcd for C23H21NO2 (343.42): C,
80.44; H, 6.16; N, 4.08. Found: C, 80.27; H, 6.21; N, 3.88.
carboxamide (5c).
A light yellow solid was obtained. Yield:
88%, Rf 0.59 (petroleum ether/EtOAc 3:2 v/v); mp 174–176 °C; 1H
NMR (DMSO-d6): d 8.92 (t, J = 5.6 Hz, 1H, NH), 8.80 (d, J = 2.1 Hz,
1H, H-4), 8.35 (dd, J = 2.0, 9.1 Hz, 1H, H-6), 7.93 (d, J = 9.1 Hz, 1H,
H-7), 7.79 (s, 1H, H-3), 7.44–7.26 (m, 5H, Ar), 5.62 (d, J = 2.9 Hz,
1H, OH), 4.85 (m, 1H, H-1), 3.59–3.37 (m, 2H, CH2). 13C NMR
(DMSO-d6): d 157.8 (C, C@O), 157.2 (C), 152.3 (C), 144.5 (C),
143.8 (C), 128.5 (2 ꢁ CH), 128.1 (C), 127.5, 126.4, 126.4, 122.4,
119.9, 113.2, 110.4 (7 ꢁ CH), 71.3 (CH–OH), 47.5 (CH2). Anal. Calcd
for C17H14N2O5 (326.306): C, 62.57; H, 4.32; N, 8.58. Found: C,
62.42; H, 4.21; N, 8.59.
5.1.4. General procedure for the synthesis of 4,5-dihydro-1,3-
oxazole derivatives (6) and (10)
5.1.3.4. N2-[2-(4-Fluorophenyl)-2-hydroxyethyl]benzo[b]furan-
To a solution of N-(2-hydroxy-2-phenylethyl)benzo[b]furan-2-
carboxamide (5) (7.11 mmol) in anhydrous THF (15 mL) was added
methanesulfonic anhydride (10.66 mmol) dissolved in anhydrous
THF (5 mL). The homogenous mixture was stirred at 0 °C for
15 min, then triethylamine (3.0 mL, 21.33 mmol) was added drop-
wise to the above mixture. After keeping the homogenous mixture
in the fridge at 0 °C for 24 h the reaction went to completion. The
mixture was quenched by the addition of aqueous ammonia solu-
tion (28%, 1 mL). After stirring at room temperature for 15 min the
mixture was concentrated in vacuo and finally distributed between
ethyl acetate (150 mL) and aqueous NaHCO3 solution (2 ꢁ 100 mL).
After repeated extraction the organic phases were dried and evap-
orated in vacuo.
2-carboxamide (5d).
A white solid was obtained. Yield: 85%, Rf
0.41 (petroleum ether/EtOAc 3:2 v/v); mp 172–174 °C; 1H NMR
(DMSO-d6): d 8.69 (t, J = 5.7 Hz, 1H, NH), 7.79 (d, J = 7.7 Hz, 1H,
Ar), 7.68 (d, J = 8.3 Hz, 1H, Ar), 7.58 (s, 1H, Ar), 7.51–7.27 (m, 4H,
Ar), 7.18 (t, J = 8.9 Hz, 2H, Ar), 5.68 (d, J = 4.5 Hz, 1H, OH), 4.85
(m, 1H, CH-OH), 3.57–3.37 (m, 2H, CH2). 13C NMR (DMSO-d6): d
163.3 (C@O), 160.1 (C), 158.6 (C–F), 154.6 (C), 149.5 (C), 140.1
(C), 128.4 (CH), 128.3 (CH), 127.5 (C), 127.1, 124.05, 123.1, 115.3,
115.0, 112.2, 109.7 (7 ꢁ CH), 70.8 (CH–OH), 47.2 (CH2). Anal. Calcd
for C17H14FNO3 (299.300): C, 68.22; H, 4.71; N, 4.68. Found: C,
68.27; H, 4.67; N, 4.66.
5.1.3.5. N2-[2-(4-Chlorophenyl)-2-hydroxyethyl]benzo[b]furan-
2-carboxamide (5e).
A white solid was obtained. Yield: 84%, Rf
0.47 (petroleum ether/EtOAc 3:2 v/v); mp 189–191 °C; 1H NMR
(DMSO-d6): d 8.69 (t, J = 5.7 Hz, 1H, NH), 7.79 (d, J = 7.8 Hz, 1H, Ar),
7.68 (d, J = 8.3 Hz, 1H, Ar), 7.57 (s, 1H, Ar), 7.51–7.33 (m, 6H, Ar),
5.71 (d, J = 4.5 Hz, 1H, OH), 4.84 (ddd, J = 4.9, 7.2, 12.1 Hz, 1H, CH-
OH), 3.56–3.37 (m, 2H, CH2). 13C NMR (DMSO-d6): d 158.6 (C@O),
154.6 (C), 149.5 (C), 142.9 (C), 128.4, 128.4, 128.3, 128.3 (4 ꢁ CH),
127.5 (C), 127.2, 124.1, 123.1, 112.2, 109.8 (5 ꢁ CH), 70.8 (CH–OH),
47.1 (CH2). Anal. Calcd for C17H14ClNO3 (315.754): C, 64.67; H,
4.47; N, 4.43. Found: C, 64.58; H, 4.51; N, 4.44.
5.1.4.1. 2-Benzo[b]furan-2-yl-5-phenyl-4,5-dihydro-1,3-oxazole
(6a).
ethanol. Yield: 54%, Rf 0.57 (petroleum ether/EtOAc 1:1 v/v); mp
79–81 °C; 1H NMR:
7.71 (d, J = 7.8 Hz, 1H, Ar), 7.45 (d,
A yellow solid was obtained after recrystallisation from
d
J = 8.4 Hz, 1H, Ar), 7.49–7.32 (m, 8H, Ar), 5.75 (dd, Jx,a = 8.1 Hz,
Jx,b = 10.0 Hz, 1H, CHx), 4.60 (dd, Jb,x = 10.1 Hz, Jb,a = 15.1 Hz, 1H,
CHb), 4.14 (dd, Ja,x = 8.0 Hz, Ja,b = 15.1 Hz, 1H, CHa). 13C NMR: d
157.3, 156.2, 144.6, 140.7 (4 ꢁ C), 129.3 (CH), 129.3 (CH), 129.0
(CH), 127.8 (C), 127.2, 126.3, 126.3, 124.0, 122.7, 112.5, 111.2,
82.1 (8 ꢁ CH), 63.60 (CH2). LRMS (ES+) m/z: 286.1 (M+Na)+. Anal.
Calcd for C17H13NO2ꢂ0.1H2O (265.097): C, 77.02; H, 5.02; N, 5.28.
Found: C, 77.05; H, 4.95; N, 5.25.
5.1.3.6.
(9a).
4-Bromo-N-(2-hydroxy-2-phenylethyl)benzamide
A white solid was obtained after recrystallisation from

4944
A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
5.1.4.2.
dro-1,3-oxazole (6b).
2-(6-Methoxybenzo[b]furan-2-yl)-5-phenyl-4,5-dihy-
A light yellow solid was obtained after
(C), 125.6 (CH), 125.6 (CH), 125.3 (C), 80.4 (CH), 62.5 (CH2). Anal.
Calcd for C18H16BrN3Oꢂ0.3H2O (375.65): C, 57.55; H, 4.45; N,
purification by flash column chromatography (petroleum ether/
ethyl acetate 90:10 v/v increasing to 50:50 v/v). Yield: 67%, Rf
0.30 (petroleum ether/EtOAc 3:2 v/v); mp116–118 °C; 1H NMR: d
7.65 (d, J = 8.7 Hz, 1H, Ar), 7.50 (d, J = 0.5 Hz, 1H, Ar), 7.47–7.33
(m, 6H, Ar), 7.00 (d, J = 2.2, 8.7 Hz, 1H, Ar), 5.82 (dd, Jx,a = 7.5 Hz,
Jx,b = 10.0 Hz, 1H, CHx), 4.49 (dd, Jb,x = 10.0 Hz, Jb,a = 15.1 Hz, 1H,
CHb), 3.89 (dd, Ja,x = 7.5 Hz, Ja,b = 15.1 Hz, 1H, CHa), 3.86 (s, 3H,
OCH3). 13C NMR: d 159.9, 156.7, 155.7, 143.4, 141.1 (5 ꢁ C),
129.2, 129.2, 128.7, 126.2, 126.2, 123.2 (6 ꢁ CH), 120.5 (C), 113.8
(CH), 111.4 (CH), 96.3 (CH), 80.6 (CH), 62.9 (CH2), 56.1 (CH3). Anal.
Calcd for C18H15NO3 (293.320): C, 73.71; H, 5.15; N, 4.77. Found: C,
73.52; H, 5.03; N, 4.72.
11.19. Found: C, 57.38; H, 4.42; N, 11.22.
5.1.4.7. 2-(4-Benzylphenyl)-5-phenyl-4,5-dihydro-1,3-oxazole
(10b).
A yellow syrup was obtained. Yield: 80%, Rf 0.8 (petro-
7.83 (d,
leum ether/EtOAc 1:1 v/v); 1H NMR (DMSO-d6):
d
J = 7.9 Hz, 1H, Ar), 7.48 (dd, J = 1.4, 7.6 Hz, 1H, Ar), 7.36–7.19 (m,
15H, Ar), 5.68 (dd, Jx,a = 7.8 Hz, Jx,b = 10.2 Hz, 1H, CHx), 4.50 (d,
J = 14.5 Hz, 2H, Ar-CH2-Ar), 4.47 (dd, Jb,x = 10.2 Hz, Jb,a = 14.9 Hz,
1H, CHb), 3.85 (dd, Ja,x = 7.8 Hz, Ja,b = 14.9 Hz, 1H, CHa). 13C NMR
(DMSO-d6): d 162.5 (C), 141.2 (C), 141.2 (C), 141.1 (C), 131.2,
130.8, 129.7, 128.9, 128.7, 128.7, 128.5, 128.2, 128.1, 128.0
(10 ꢁ CH), 126.8 (C), 126.4, 126.3, 125.8, 125.7, 79.2 (5 ꢁ CH),
63.1 (CH2), 38.5 (CH2). Anal. Calcd for C25H23N3Oꢂ0.1H2O
(383.28): C, 78.34; H, 6.10; N, 10.96. Found: C, 78.39; H, 6.12; N,
11.10.
5.1.4.3.
1,3-oxazole (6c).
2-(5-Nitrobenzo[b]furan-2-yl)-5-phenyl-4,5-dihydro-
A white solid was obtained after purification
by flash column chromatography (petroleum ether/ethyl acetate
90:10 v/v increasing to 60:40 v/v). Yield: 16%, Rf 0.30 (petroleum
ether/EtOAc 3:2 v/v); mp159–161 °C; 1H NMR (DMSO-d6): d 8.75
(d, J = 2.3 Hz, H-4), 8.36 (dd, J = 2.4, 9.1 Hz, H-6), 7.99 (d,
J = 9.1 Hz, 1H, H-7), 7.79 (s, 1H, H-3), 7.48–7.36 (m, 5H, Ar), 5.89
(dd, Jx,a = 7.8 Hz, Jx,b = 10.0 Hz, 1H, CHx), 4.55 (dd, Jb,x = 10.0 Hz,
Jb,a = 15.5 Hz, 1H, CHb), 3.96 (dd, Ja,x = 7.8 Hz, Ja,b = 15.5 Hz, 1H,
CHa). 13C NMR (DMSO-d6): d 157.9, 155.0, 147.1, 144.6, 140.6
(5 ꢁ C), 129.2 (CH), 129.2 (CH), 128.8 (CH), 128.0 (C), 126.3,
126.3, 122.6, 119.6, 113.3, 112.0, 81.2 (7 ꢁ CH), 62.9 (CH2). Anal.
Calcd for C17H12N2O4 (308.91): C, 66.23; H, 3.92; N, 9.08. Found:
C, 66.09; H, 3.89; N, 9.06.
5.1.4.8. (E)-5-Phenyl-2-(4-styrylphenyl)-4,5-dihydro-1,3-oxazole
(10c).
A white solid was obtained by recrystallisation from eth-
anol. Yield: 61%, Rf 0.57 (petroleum ether/EtOAc 1:1 v/v); mp 126–
127 °C; 1H NMR (DMSO-d6): d 7.97 (d, J = 8.3 Hz, 2H, Ar), 7.6 (d,
J = 8.3 Hz, 2H, Ar), 7.67 (d, J = 7.5 Hz, 2H, Ar), 7.46–7.33 (m, 10H,
Ar), 5.80 (dd, Jx,a = 7.7 Hz, Jx,b = 9.9 Hz, 1H, CHx), 4.48 (dd,
Jb,x = 10.1 Hz, Jb,a = 14.9 Hz, 1H, CHb), 3.85 (dd, Ja,x = 7.5 Hz,
Ja,b = 14.9 Hz, 1H, CHa). 13C NMR (DMSO-d6): d 162.2 (C), 141.3
(C), 140.1 (C), 136.7 (C), 130.4, 128.7, 128.7, 128.7, 128.2, 128.2,
128.1, 128.05, 127.5, 127.5, 126.7, 126.7, 126.6, 126.6, 125.6,
125.6 (16 ꢁ CH, Ar), 126.2 (C), 80.1 (CH), 62.6 (CH2). Anal. Calcd
for C23H19NO (325.40): C, 84.89; H, 5.88; N, 4.30. Found: C,
84.70; H, 5.94; N, 4.11.
5.1.4.4. 2-Benzo[b]furan-2-yl-5-(4-fluorophenyl)-4,5-dihydro-
1,3-oxazole (6d).
A yellow solid was obtained after recrystalli-
sation from ethanol. Yield: 66%, Rf 0.74 (petroleum ether/EtOAc 3:2
v/v); mp 88–90 °C; 1H NMR (DMSO-d6): d 7.78 (d, J = 7.8 Hz, 1H,
Ar), 7.72 (d, J = 8.3 Hz, 1H, Ar), 7.59 (s, 1H, Ar), 7.52–7.45 (m, 3H,
Ar), 7.39–7.33 (m, 1H, Ar), 7.27 (t, J = 8.9 Hz, 2H), 5.86 (dd,
Jx,a = 7.8 Hz, Jx,b = 9.9 Hz, 1H, CHx), 4.50 (dd, Jb,x = 9.9 Hz,
Jb,a = 15.3 Hz, 1H, CHb), 3.92 (dd, Ja,x = 7.8 Hz, Ja,b = 15.3 Hz, 1H,
CHa). 13C NMR (DMSO-d6): d 164.0, 160.7, 155.6, 155.3, 144.2,
137.2, 137.1 (7 ꢁ C), 128.6 (CH), 128.6 (CH), 128.5 (CH), 128.5
(CH), 127.4 (C), 127.3, 124.2, 122.9, 116.2, 116.2, 115.9, 115.9,
112.1, 111.3, 80.2 (10 ꢁ CH), 62.9 (CH2). Anal. Calcd for C17H12FNO2
(281.285): C, 72.59; H, 4.30; N, 4.98. Found: C, 72.40; H, 4.28; N,
4.90.
5.1.5. General procedure for the synthesis of N2-(2-phenyl-2-
(1H-1-imidazoyl)ethyl)-arylamide derivatives (7) and (11)
A mixture of oxazole (6) or (10) (1.25 mmol) and imidazole
(25 mmol) dissolved in isopropyl acetate (3 mL) was heated at
125 °C for 24 h. After completion of the reaction the mixture was
partitioned between water (100 mL) and ethyl acetate (150 mL).
The organic layer was washed three times with water
(3 ꢁ 100 mL), dried over MgSO4 and concentrated in vacuo.
5.1.5.1. N2-(2-Phenyl-2-(1H-1-imidazoyl)ethyl)-benzo[b]furan-
2-carboxamide (7a).
A white solid was obtained after recrys-
tallisation from ethanol. Yield: 28%, Rf 0.74 (CH2Cl2/MeOH 9:1 v/
v); mp 180–182 °C; 1H NMR (DMSO-d6): d 9.04 (t, J = 5.4 Hz, 1H,
NH), 7.90 (s, 1H, imid), 7.79 (d, J = 7.7 Hz, 1H, Ar), 7.66 (d,
J = 8.3 Hz, 1H, Ar), 7.56 (s, 1H, H-3), 7.52–7.33 (m, 8H, Ar and imid),
6.95 (s, 1H, H-imidazole), 5.75 (dd, J = 5.6, 9.3 Hz, 1H, H-1), 4.23–
3.98 (m, 2H, CH2). 13C NMR (DMSO-d6): d 158.8 (C@O), 154.5 (C),
149.0 (C), 139.5 (CH), 137.1 (C), 129.1, 129.1, 128.9, 128.5
(4 ꢁ CH), 127.4 (C), 127.3, 127.2, 127.2, 124.1, 123.2, 118.7,
112.1, 110.2, 59.8 (9 ꢁ CH), 43.2 (CH2). LRMS (ES+) m/z: 332.13
(M+H)+. Anal. Calcd for C20H17N3O2ꢂ0.4H2O (338.579): C, 70.95;
H, 5.30; N, 12.41. Found: C, 71.08; H, 5.40; N, 12.21.
5.1.4.5. 2-Benzo[b]furan-2-yl-5-(4-chlorophenyl)-4,5-dihydro-
1,3-oxazole (6e).
A light yellow solid was obtained by recrys-
tallisation from ethanol. Yield: 65%, Rf 0.62 (petroleum ether/EtOAc
3:2 v/v); mp 103–105 °C; 1H NMR (DMSO-d6): d 7.70 (d, J = 7.7 Hz,
1H, Ar), 7.74 (d, J = 8.3 Hz, 1H, Ar), 7.62 (s, 1H, Ar), 7.54–7.36 (m,
6H, Ar), 5.88 (dd, Jx,a = 7.5 Hz, Jx,b = 10.0 Hz, 1H, CHx), 4.53 (dd,
Jb,x = 10.0 Hz, Jb,a = 15.3 Hz, 1H, CHb), 3.92 (dd, Ja,x = 7.5 Hz,
Ja,b = 15.3 Hz, 1H, CHa). 13C NMR (DMSO-d6): d 155.6, 155.3,
144.1, 140.0, 133.3 (5 ꢁ C), 129.2 (CH), 129.2 (CH), 128.1 (CH),
128.1 (CH), 127.4 (C), 127.3, 124.2, 123.0, 112.1, 111.3, 80.0
(6 ꢁ CH), 62.9 (CH2). Anal. Calcd for C17H12ClNO2 (297.740): C,
68.58; H, 4.06; N, 4.70. Found: C, 68.56; H, 3.97; N, 4.72.
5.1.5.2.
N2-[2-Phenyl-2-(1H-1-imidazoyl)ethyl]-6-meth-
A brown solid was
oxybenzo[b]furan-2-carboxamide (7b).
obtained after recrystallisation from ethanol/water (2:1 v/v). Yield:
66%, Rf 0.63 (CH2Cl2/MeOH 9:1 v/v); mp 110–112 °C; 1H NMR
(DMSO-d6): d 8.83 (t, J = 5.5 Hz, 1H, NH), 7.87 (s, 1H, imid), 7.62
(d, J = 8.6 Hz, 1H, Ar), 7.45 (s, 1H, H-3), 7.38–7.29 (m, 6H, Ar and
imid), 7.16 (d, J = 1.8 Hz, 1H), 6.96 (dd, J = 1.8, 8.7 Hz, 1H, Ar),
6.92 (s, 1H, imid), 5.70 (dd, J = 5.7, 9.4 Hz, 1H, H-1), 4.17–3.94
(m, 2H, CH2), 3.82 (s, 3H, CH3). 13C NMR (DMSO-d6): d 158.8
(C@O), 154.5 (C), 149.0 (C), 139.5 (CH), 137.14 (C), 129.1, 129.1,
128.9, 128.5 (4 ꢁ CH), 127.4 (C), 127.3, 127.2, 127.2, 124.1, 123.2,
5.1.4.6. 2-(4-Bromophenyl)-5-phenyl-4,5-dihydro-1,3-oxazole
(10a).
A yellow syrup was obtained. Yield: 82%, Rf 0.8 (petro-
7.91 (d,
leum ether/EtOAc 1:1 v/v); 1H NMR (DMSO-d6):
d
J = 8.5 Hz, 2H, Ar), 7.80 (d, J = 6.8 Hz, 2H, Ar), 7.43 (m, 5H, Ar),
5.83 (dd, Jx,a = 7.5 Hz, Jx,b = 10.1 Hz, 1H, CHx), 4.50 (dd,
Jb,x = 10.1 Hz, Jb,a = 15.1 Hz, 1H, CHb), 3.88 (dd, Ja,x = 7.5 Hz,
Ja,b = 15.1 Hz, 1H, CHa). 13C NMR (DMSO-d6): d 161.7 (C), 141.0
(C), 131.8, 131.8, 129.8, 129.8, 128.8, 128.8, 128.2 (7 ꢁ CH), 126.5

A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
4945
118.7, 112.1, 110.2, 59.8 (9 ꢁ CH), 43.24 (CH2). Anal. Calcd for
mp 40–42 °C; 1H NMR (DMSO-d6): d 8.67 (t, J = 5.5 Hz, 1H, NH),
7.88 (s, 1H, imid), 7.06–7.88 (m, 15H, Ar), 6.95 (s, 1H, Ar), 5.67
(dd, Jx,b = 9.3 Hz, Jx,a = 6.0 Hz, 1H, CHx), 4.09 (m, 1H, CHb), 4.01
(m, 3H, Ar-CH2-Ar and CHa). 13C NMR (DMSO-d6): d 169.46
(C@O), 140.90 (C), 139.15 (C), 138.97 (C), 136.40 (C), 136.75,
130.05, 129.50, 128.94, 128.87, 128.67, 128.52, 128.19, 128.17,
128.06, 127.19, 126.9, 126.9, 125.8, 125.7, 125.3, 118.3 (17 ꢁ CH,
Ar), 59.39 (CH), 43.02 (CH2), 30.6 (CH2). Anal. Calcd for
C21H19N3O3ꢂ0.1H2O (363.201): C, 69.45; H, 5.33; N, 11.60. Found:
C, 69.43; H, 5.41; N, 11.54.
5.1.5.3. N2-[2-Phenyl-2-(1H-1-imidazoyl)ethyl)-5-nitrobenzo[b]
furan-2-carboxamide (7c).
A brown solid was obtained after
recrystallisation from ethanol/water (2:1 v/v). Further purification
by flash column chromatography (CH2Cl2/MeOH 100:0 v/v increas-
ing to 95:5 v/v) gave 7c as a white solid. Yield: 33%, Rf 0.23 (CH2Cl2/
MeOH 9:1 v/v); mp 204–206 °C; 1H NMR (DMSO-d6): d 9.23 (t,
J = 5.6 Hz, 1H, NH), 8.79 (d, J = 2.4 Hz, 1H, Ar), 8.33 (dd, J = 2.4,
9.1 Hz, 1H, Ar), 7.90 (d, J = 8.6 Hz, 2H, Ar), 7.75 (s, 1H, imid),
7.42–7.33 (m, 6H, Ar and imid), 6.93 (s, 1H, imid), 5.71 (dd,
J = 5.6, 9.3 Hz, 1H, Ar), 4.22–3.98 (m, 2H, CH2). 13C NMR (DMSO-
d6): d 158.1 (C@O), 157.2, 151.7, 144.6, 139.4 (4 ꢁ C), 137.2,
129.1, 129.1, 129.0, 129.0, 128.5 (6 ꢁ CH), 128.0 (C), 127.1, 122.6,
120.0, 118.7, 113.3, 111.0, 59.8 (7 ꢁ CH), 43.35 (CH2). Anal. Calcd
for C20H16N4O4ꢂ0.3H2O (381.775): C, 62.92; H, 4.38; N, 14.68.
Found: C, 62.72; H, 4.13; N, 14.50.
C25H23N3O (381.48): C, 78.71; H, 6.08; N, 11.02. Found: C, 78.91;
H, 6.13; N, 11.20.
5.1.5.8. (E)-4-N-(2-(1H-imidazol-1-yl)-2-phenylethyl)-4-styrylb-
enzamide (11c).
Purification by flash column chromatography
(CH2Cl2/MeOH 95:5 v/v) gave a white solid. Yield: 16%, Rf 0.40
(CH2Cl2/MeOH 9:1 v/v); mp 190–192 °C; 1H NMR (DMSO-d6): d
8.77 (s, 1H, NH), 7.89 (s, 1H, imid), 7.81 (d, J = 8.0 Hz, 2H, Ar), 7.71
(d, J = 8.0 Hz, 2H, Ar), 7.66 (d, J = 7.4 Hz, 2H, Ar), 7.43–7.32 (m,
11H, Ar), 6.96 (s, 1H, imid), 5.67 (dd, Jx,b = 8.7 Hz, Jx,a = 5.8 Hz, 1H,
CHx), 4.12 (m, 1H, CHb), 4.03 (m, 1H, CHa). 13C NMR (DMSO-d6): d
166.3 (C@O), 140.0, 139.3, 136.7, 132.8 (4 ꢁ C), 130.3, 128.7, 128.7,
128.6, 128.5, 128.5, 128.1, 128.0, 128.0, 127.6, 127.6, 127.4, 127.4,
126.8, 126.8, 126.7, 126.7, 126.2, 126.2 (19 ꢁ CH, Ar), 59.4 (CH),
43.5 (CH2). Anal. Calcd for C26H23N3O (393.48): C, 79.36; H, 5.89;
N, 10.67. Found: C, 79.10; H, 5.80; N, 10.43.
5.1.5.4. N2-[2-(4-Fluorophenyl)-2-(1H-1-imidazoyl)ethyl]-ben-
zo[b]furan-2-carboxamide (7d).
A brown solid was obtained
after recrystallisation from ethanol/water (2:1 v/v). Yield: 73%,
Rf 0.38 (CH2Cl2/MeOH 9:1 v/v); mp 198–200 °C; 1H NMR (DMSO-
d6): d 9.00 (t, J = 5.5 Hz, 1H, NH), 7.88 (s, 1H, imid), 7.78 (d,
J = 7.7 Hz, 1H, Ar), 7.54 (s, 1H, Ar), 7.65 (d, J = 8.3 Hz, 1H, Ar), 7.48
(m, 3H, Ar), 7.35 (m, 2H, Ar), 7.23 (t, J = 8.8 Hz, 2H, Ar), 6.93 (s,
1H, imid), 5.73 (dd, J = 6.2, 8.9 Hz, 1H, H-1), 4.18–3.96 (m, 2H,
CH2). 13C NMR (DMSO-d6): d 163.8 (C@O), 160.5, 158.8, 154.6,
149.0 (4 ꢁ C), 137.1 (CH), 135.8 (C), 135.5 (C), 129.5 (CH), 129.4
(CH), 129.0 (CH), 127.4 (C), 127.3, 124.1, 123.2, 118.6, 116.1,
115.8, 112.1, 110.3, 59.1 (9 ꢁ CH), 43.20 (CH2). Anal. Calcd for
5.2. Materials and methods: CYP24A1 assay
A recombinant cell line expressing human CYP24A1 enzyme
(V79-CYP24) was used to measure the inhibitory properties of imi-
dazoles 7 and 11 on the enzyme activity as described before.18,22
Briefly, the inhibitors were dissolved in 100% ethanol or 100%
DMSO to form a 0.02 M stock solution. All stock solutions were di-
luted with 100% ethanol to obtain the range of inhibitor solutions.
Ketoconazole (Sigma) was weighed in a darkened room immedi-
ately prior to incubation and dissolved in 0.05 M HCl to form a
C20H16FN3O2 (349.363): C, 68.76; H, 4.62; N, 12.02. Found: C,
68.56; H, 4.62; N, 12.11.
5.1.5.5. N2-[2-(4-Chlorophenyl)-2-(1H-1-imidazoyl)ethyl]-ben-
1.9 mM solution, which was diluted to 190
Each millilitre of incubation media contained 1 or 2
phenyl-p-phenylenediamine (DPPD) (Sigma), substrate, 5
inhibitor or carrier or enough 190 M ketoconazole to give the cor-
rect final concentration, and the same medium used for cell culture
of that cell type but containing 1% BSA (Boehringer Mannheim,
Laval, PQ, Canada) instead of FCS. The substrate for CYP24A1 cells
l
M using 0.05 M HCl.
L N0,N-di-
L of
zo[b]furan-2-carboxamide (7e).
A brown residue was ob-
l
tained after recrystallisation from ethanol/water (2:1 v/v). Further
purification by flash column chromatography (dichloromethane/
methanol 100:0 v/v increasing to 96:4 v/v) gave a light yellow solid.
Yield: 72%, Rf 0.16(petroleumether/EtOAc 3:2 v/v); mp 76–78 °C; 1H
NMR (DMSO-d6): d 9.00 (t, J = 5.4 Hz, 1H, NH), 7.88 (s, 1H, imid), 7.76
(d, J = 7.7 Hz, 1H, Ar), 7.63 (d, J = 8.3 Hz, 1H, Ar), 7.53 (s, 1H, Ar), 7.49–
7.31 (m, 7H, Ar and imid), 6.91 (s, 1H, imid), 5.73 (dd, J = 6.2, 8.8 Hz,
1H, H-1), 4.16–3.95 (m, 2H, CH2). 13C NMR (DMSO-d6): d 158.8
(C@O), 154.6 (C), 149.0 (C), 138.5 (C), 137.2 (CH), 133.2 (C), 129.2,
129.2, 129.1, 129.1 (4 ꢁ CH), 127.4 (C), 127.3, 124.1, 123.2, 123.2,
118.7, 112.1, 110.3 (7 ꢁ CH, Ar), 59.1 (CH), 43.10 (CH2). Anal. Calcd
for C20H16ClN3O2ꢂ0.2H2O (369.421): C, 65.03; H, 4.47; N, 11.37.
Found: C, 64.93; H, 4.35; N, 11.36.
l
l
was 250,000 cpm of [1b-3H]1a,25-(OH)2D3 dissolved in ethanol
per mL incubation media. Dead cell controls were prepared by
microwaving plates for 3 min. A 1 mL volume of incubation media
was used for each well of a 6-well plate, whereas 2 mL of incuba-
tion media was used for each 60-mm plate. All conditions were
conducted in triplicate.
The reaction was terminated by the addition of 500 lL of metha-
nol and transferred to a glass tube. The aqueous phase was extracted
by standard Bligh–Dyer extraction in which we substituted dichlo-
romethane for chloroform.23 Samples were then spun at 4000 rpm
5.1.5.6. 4-Bromo-N-(2-(1H-imidazol-1-yl)-2-phenylethyl)benz-
amide (11a).
A pale yellow solid was obtained after recrystal-
for 5 min. Triplicate 100
lL aliquots of aqueous fraction containing
lisation from ethanol. Yield: 41%, Rf 0.5 (CH2Cl2/MeOH 9:1 v/v);
mp 180–181 °C; 1H NMR (DMSO-d6): d 8.86 (t, J = 5.4 Hz, 1H,
NH), 7.87 (s, 1H, imid), 7.74 (m, 4H, Ar), 7.38 (m, 6H, Ar), 6.95 (s,
1H, imid), 5.70 (dd, J = 9.3 Hz, 1H, CHx), 4.11 (m, 1H, CHb), 4.01
(m, 1H, CHa). 13C NMR (DMSO-d6): d 165.79 (C@O), 139.18 (C),
133.16 (C), 125.08 (C), 136.75, 131.31, 129.26, 128.69, 128.52,
128.06, 126.92, 119.60, 118.30 (9 ꢁ CH, Ar), 59.34 (CH), 43.48
(CH2). Anal. Calcd for C18H16BrN3O (370.24): C, 58.39; H, 4.36; N,
11.34. Found: C, 58.67; H, 4.36; N, 11.56.
water-soluble CYP24A1 products were mixed with 600
l
L of scintil-
lation fluid and the radioactivity was measured by use of a scintilla-
tion counter. All values were normalized for background.
The log of the inhibitor concentration was plotted versus the
quantification property using GraphPad Prism and the curve was
analysed using sigmoidal dose–response analysis to calculate an
IC50 value for the inhibitor.
5.3. Molecular docking
5.1.5.7. 4-Benzyl-N-(2-(1H-imidazol-1-yl)-2-phenylethyl)benz-
All molecular modelling studies were performed on a MacPro
dual 2.66 GHz Xeon running Ubuntu 8. Docking simulations were
performed using PLANTS24 and the ligands were built with Molec-
amide (11b).
A pale brown solid was obtained after recrystal-
lisation from ethanol. Yield: 40%, Rf 0.55 (CH2Cl2/MeOH 9:1 v/v);

4946
A. S. Aboraia et al. / Bioorg. Med. Chem. 18 (2010) 4939–4946
ular Operating Environment (MOE).25 The docking results were
visualized with MOE.
10. Kahraman, M.;Sinishtaj, S.; Dolan, P. M.;Kensler, T. W.;Peleg, S.; Saha, U.; Chuang,
S. S.; Bernstein, G.; Korczak, B.; Posner, G. H. J. Med. Chem. 2004, 47, 6854.
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241, 49.
12. Schuster, I.; Egger, H. Acylated aminoalkanimidazoles and -triazoles. Austria,
Patent No.: U.S. 5622,982, 1997.
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For A.S.A. and M.S.G. we acknowledge the Embassy of the Arab
Republic of Egypt for the award of a Ph.D. scholarship. For S.W.Y.
we would like to acknowledge the ORS Awards Scheme for a
United Kingdom Scholarship.
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