Discovery of N-(1-(3-(4-phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethyl)acetamides as novel acetyl-CoA carboxylase 2 (ACC2) inhibitors with peroxisome proliferator-activated receptor α/δ (PPARα/δ) dual agonistic activity

Article abstract of DOI:10.1016/j.bmc.2016.08.045

Acetyl-CoA carboxylases (ACCs) catalyze a critical step in de novo lipogenesis, and are considered as promising targets for treatment of obesity, dyslipidemia and type 2 diabetes mellitus. On the other hand, peroxisome proliferator-activated receptors (PP

Full text of DOI:10.1016/j.bmc.2016.08.045

Bioorganic & Medicinal Chemistry xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Discovery of N-(1-(3-(4-phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethyl)
acetamides as novel acetyl-CoA carboxylase 2 (ACC2) inhibitors with
peroxisome proliferator-activated receptor
agonistic activity
a/d (PPARa/d) dual
Shogo Okazaki a, Tomomi Noguchi-Yachide a, Taki Sakai a, Minoru Ishikawa a, Makoto Makishima b,
Yuichi Hashimoto a, Takao Yamaguchi a,
⇑
a Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan
b Nihon University School of Medicine, 30-1 Oyaguchi-kamicho, Itabashi-ku, Tokyo 173-8610, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Acetyl-CoA carboxylases (ACCs) catalyze a critical step in de novo lipogenesis, and are considered as
promising targets for treatment of obesity, dyslipidemia and type 2 diabetes mellitus. On the other hand,
peroxisome proliferator-activated receptors (PPARs) are well-established therapeutic targets for these
metabolic syndrome-related diseases. Therefore, we considered that dual modulators of ACC and
PPARs would be promising candidates as therapeutic agents. Here, we designed a series of acetamides
based on the molecular similarity between ACC inhibitors and PPAR agonists. Screening of the synthe-
sized compounds identified N-(1-(3-(4-phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethyl)acetamides as novel
Received 22 July 2016
Revised 24 August 2016
Accepted 25 August 2016
Available online xxxx
Keywords:
Acetyl-CoA carboxylase
Peroxisome proliferator-activated receptor
Multi-target drug
ACC2 inhibitors with PPARa/PPARd dual agonistic activity. Structure–activity relationship studies and
further structural elaboration afforded compounds with distinct activity profiles. Our findings should
be helpful for the discovery of candidate agents with an appropriate balance of ACC-inhibitory and
PPAR-activating activities for therapeutic lipid control.
Ó 2016 Elsevier Ltd. All rights reserved.
1. Introduction
Malonyl-CoA is a substrate of fatty acid synthetase (FAS) in de novo
lipogenesis and a negative regulator of carnitine palmitoyltrans-
The term metabolic syndrome refers to a cluster of risk factors,
including central obesity, hypertension (high blood pressure), ele-
vated fasting plasma glucose, dyslipidemia (e.g., elevated triglyc-
erides and low high-density lipoprotein cholesterol levels), and
insulin resistance, that lead to type 2 diabetes mellitus (T2DM) as
well as cardiovascular diseases (CVD). It is a serious public health
problem worldwide.1 Although the molecular mechanisms of the
pathogenesis of metabolic syndrome are not fully understood, it
is established that disordered lipid metabolism is closely related
to the development of obesity and insulin resistance.2–5 For exam-
ple, ectopic lipid accumulation in non-adipose tissues such as liver
and skeletal muscle is associated with insulin resistance.2 There-
fore, rebalancing of lipid metabolism is expected to be beneficial
for a whole range of complications associated with metabolic syn-
drome.5 Inhibition of acetyl-CoA carboxylases (ACCs) has recently
received particular attention in this area.6 ACCs catalyze the
ATP-dependent carboxylation of acetyl-CoA to form malonyl-CoA.
ferase 1 (CPT1), a mitochondrial outer membrane protein that
transports free fatty acyl-CoAs into mitochondria for b-oxidation
(fatty acid catabolism step). In humans, there are two ACC isoforms;
cytosolic ACC1 is primarily expressed in lipid-rich tissues (liver and
adipose) and is involved in lipogenesis, whereas mitochondrial
ACC2 is primarily found in oxidative tissues (muscle) and mainly
controls fatty acid oxidation. Knockdown of ACC1, ACC2 or both sig-
nificantly increases fatty acid oxidation in rat hepatocytes7, and an
ACC inhibitor, soraphen A, improves insulin sensitivity in mice fed a
high-fat diet.8 These data indicate that ACC inhibition could be use-
ful in the treatment of T2DM and obesity.
The peroxisome proliferator-activated receptors (PPARs) have
also been recognized as molecular targets for such metabolic syn-
drome-related diseases.9 PPARs are ligand-dependent transcription
factors belonging to the nuclear hormone receptor superfamily.
They regulate transcription of a large number of different genes
involved in lipid metabolism, glucose homeostasis and adipogene-
sis, in response to the binding of endogenous lipophilic ligands (e.g.
prostaglandins, fatty acids and their metabolites). There are three
⇑
Corresponding author.
http://dx.doi.org/10.1016/j.bmc.2016.08.045
0968-0896/Ó 2016 Elsevier Ltd. All rights reserved.
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2
S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
subtypes of PPAR, commonly designated as PPAR
PPARd (also known as PPARb). PPAR is found in tissues with high
rates of fatty acid catabolism, such as brown adipose tissue and
liver. PPAR is predominantly expressed in adipose tissue, whereas
the third subtype, PPARd, is expressed in many tissues. PPAR ago-
a, PPARc and
2. Results and discussion
2.1. Molecular design
a
c
a
Compounds 1–3 have been reported as ACC1/2 dual inhibitors
with low to moderate ACC2 selectivity (Fig. 1).18,19 These com-
pounds have linear lipophilic structures with a pendant polar acet-
amide group. Similar molecular features are seen in known PPAR
agonists (Fig. 2), i.e., bezafibrate, pioglitazone and GW501516 have
linear lipophilic cores with an acidic head group (carboxylic acid or
thiazolidine-2,4-dione). This observation suggests that there may
be an overlapping pharmacophore for ACC inhibitors and PPAR
agonists. We thus speculated that derivatives of these compounds
might be dual modulators of ACC and PPAR. Although the binding
mode of 1–3 is unknown, the acetamide group and adjacent
methyl group, and terminal alkoxy/phenoxy group seem to be
important for the ACC-inhibitory activity according to previously
reported SAR studies.18 Taking into account the fact that PPARs
have large ligand-binding cavities,9,20 we designed and prepared
a series of acetamide-based compounds as candidate dual modula-
tors (Fig. 3, compounds 4–11). 1,2,4-Oxadiazoles were designed
because of the ease of preparation of acetamides from nitriles
and N-acetylamino acids (e.g., N-acetylalanine).
nists, represented by fibrates (e.g. bezafibrate, fenofibrate), have
been clinically used for the treatment of dyslipidemia and insulin
resistance. PPARc agonists, for example thiazolidine-2,4-diones
such as pioglitazone, which improve insulin sensitivity and glyce-
mic control, are currently being used to treat patients with T2DM.
The therapeutic potential of PPARd agonists is still under investiga-
tion, but a potent subtype-selective PPARd agonist GW501516 was
shown to improve insulin sensitivity in animal models of T2DM.10
Because both ACCs and PPARs are key targets for regulation of
lipid metabolism, compounds that simultaneously inhibit ACCs
and activate PPARs may offer synergistic benefits. Reviews of
multi-target drugs,11 the magic shotgun approach12 and polyphar-
macology13 have suggested that drugs engaging two or more tar-
gets can be therapeutically superior to single-target drugs. For
example, dual modulators that target PPAR and soluble epoxide
hydrolase (sEH)14,15 or angiotensin II type 1 receptor (AT1)16 have
recently been developed. However, to our knowledge, there is no
report on ACC/PPAR dual modulators. Currently available fibrates
have significant therapeutic benefits, but their disadvantages
include weak agonistic activities and dose-dependent side
effects;17 it is possible that these issues might be overcome by
multi-targeting. Driven largely by these concepts, we designed a
library of candidate dual modulators of ACC and PPAR on the basis
of the molecular similarity between known ACC inhibitors and
PPAR agonists. Screening of the synthesized compounds resulted
in identification of N-(1-(3-(4-phenoxyphenyl)-1,2,4-oxadiazol-5-
2.2. Synthesis and evaluation of the designed compounds
The designed compounds 4–7 were synthesized as outlined in
Scheme 1. 4-Isopropoxyphenol, prepared from hydroquinone,21
was treated with 4-fluorobenzonitrile in the presence of K2CO3 to
afford 12. Compound 13 was similarly obtained from commercially
available 4-benzyloxyphenol. Compounds 12 and 13 were con-
verted into amidoximes 14 and 15, which in turn were condensed
with N-acetylamino acids to afford the desired compounds 4 and 5,
as well as O-benzyl-protected 16. Removal of the benzyl group of
16 gave 6, which was alkylated with 1-iodopropane to afford 7.
Benzimidazoles 8–10 were prepared as shown in Scheme 2. First,
yl)ethyl)acetamides as novel ACC2 inhibitors with PPARa/d dual
agonistic activity. Further structural elaboration based on struc-
ture–activity relationship (SAR) studies led to (S)-32, which inhib-
ited ACC2 and activated PPAR
concentrations [(S)-32: IC50 (ACC2) = 0.22
= 0.15 M, EC50 (PPARd) = 0.29 M].
a
and PPARd at submicromolar
l
M, EC50 (PPAR
a)
l
l
2-chloro-5-cyanobenzimidazole,
synthesized
from
3,4-
Figure 1. Chemical structures of known ACC1/2 dual inhibitors 1–3.
Figure 2. Chemical structures of PPARa agonists (bezafibrate and GW7647), PPARc agonists (pioglitazone and rosiglitazone) and PPARd agonists (GW501516 and TIPP-401).
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S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
3
Figure 3. The designed acetamide-based compounds.
Scheme 1. Reagents and conditions: (a) 4-fluorobenzonitrile, K2CO3, DMF, 150 °C, 97% (for 12), 97% (for 13); (b) aq NH2OH, EtOH, reflux, 58% (for 14), 52% (for 15);
(c) N-acetyl-DL-alanine (for 4 and 16), N-acetylglycine (for 5), HATU, DIPEA, DMF, rt to 80–100 °C, 46% (for 4), 65% (for 5), 70% (for 16); (d) BBr3, CH2Cl2, ꢀ50 to ꢀ20 °C, 91% ;
(e) 1-iodopropane, Cs2CO3, DMF, 80 °C, 94%.
Scheme 2. Reagents and conditions: (a) SEMCl, Et3N, THF, rt, 87%; (b) 4-isopropoxyphenol (for 18), phenol (for 19), Cs2CO3, DMF, rt, 99% (for 18), 99% (for 19); (c) aq NH2OH,
EtOH, reflux, 87% (for 20), 93% (for 21); (d) N-acetylglycine (for 22), N-acetyl-DL-alanine (for 23), HATU, DIPEA, DMF, rt to 90 °C; (e) TBAF, THF, reflux, 13% in 2 steps (for 8), 22%
in 2 steps (for 9); (f) H2, Raney Ni, aq NH3, EtOH, rt, 97%; (g) Ac2O, Et3N, CH2Cl2, rt, 85%; (h) TBAF, THF, reflux, 55%.
diaminobenzonitrile,22 was protected using 2-(trimethylsilyl)
ethoxymethyl chloride (SEMCl) to give 17 as a 1:1 regioisomeric
mixture (1-N-SEM and 3-N-SEM). Coupling of 17 with phenols pro-
vided the corresponding 2-phenoxybenzimidazoles 18 and 19.
Compounds 8 and 9 were synthesized from 18 and 19 similarly
to the above 1,2,4-oxadiazole synthesis. Compound 10 was pre-
pared by reduction of the cyano group of 18 followed by acetyla-
tion and SEM deprotection. Carbamate 11 was simply obtained
from 4-aminobenzonitrile (Scheme 3).
Section 4). Some of compounds exhibited partial inhibition. Thus,
the IC50 values were determined from the dose–response curves,
and the values of percent inhibition at 30 lM were calculated
relative to the maximum ACC2 inhibition of 1. For evaluation of
PPAR-agonistic activities, we adopted a cell-based reporter gene
assay using CMX-GAL4N-hPPAR LBD as a recombinant receptor
gene, TK-MH100x4-LUC as a reporter gene, and CMX b-galactosi-
dase gene for normalization, as previously reported.23 The EC50 val-
ues were determined from the dose–response curves, and the
maximum efficacies (Emax) were calculated relative to those of
The prepared compounds were evaluated for ACC-inhibitory
and PPAR-agonistic activities (Table 1). We used recombinant
human ACC2, and performed ATP consumption assay19 (see
known PPAR agonists (GW7647 for PPARa, rosiglitazone for PPARc,
and GW501516 for PPARd).
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S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
Scheme 3. Reagents and conditions: (a) phenyl chloroformate, pyridine, THF, 0 °C to rt, 98%; (b) aq NH2OH, EtOH, 50 °C to reflux, 29%; (c) N-acetyl-DL-alanine, HATU, DIPEA,
DMF, rt to 90 °C, 78%.
Table 1
ACC2-inhibitory and PPAR-agonistic activities of acetamide-based compounds
Compound
ACC2a
IC50
PPARa b
PPARcb
PPARdb
[
l
M] (inhibition [%])c,d
EC50
[l
M] (efficacy [%])c,e
EC50
[l
M] (efficacy [%])c,e
EC50 [l
M] (efficacy [%])c,e
4
5
6
7
8
9
10
11
0.10 (74)
1.1 (83)
(24)
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
—
ia.f
ia.f
ia.f
1.0 (13–33)
1.0 (2–9)
0.61 (88)
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
ia.f
1
0.011 (100)
ia.f
ia.f
GW7647
Rosiglitazone
GW501516
—
—
—
0.021 (100)
—
—
—
—
0.26 (100)
—
0.0059 (100)
a
b
c
Inhibitory activity towards recombinant human ACC2.
Agonistic activities towards PPAR(LBD)-GAL4 chimeric receptors in transiently transfected HEK-293 cells.
Evaluations were performed in triplicate, and the EC50 and IC50 values were determined from the dose–response curves.
d
e
Percent inhibition at 30
Maximum PPAR-agonistic activities from several independent evaluations are shown in parenthesis. The maximal efficacies of GW7647, rosiglitazone and GW501516
, PPAR and PPARd, respectively.
lM is shown in parenthesis. The maximum ACC2 inhibition by 1 was defined as 100%.
were defined as 100% for PPAR
a
c
f
ia. = inactive at <20 lM.
Among the designed compounds, 4, 5, 6 and 7 exhibited ACC2-
inhibitory activity, and 4, which has a similar structure to 1, was
the most potent. Neither 4 nor 1 showed activity against PPARs.
However, 6 showed both ACC2-inhibitory activity and partial
activity (Table 2). The order of ACC2-inhibitory activity was OH
(6) ꢁ OMe (28) < OEt (29) < H (32) ꢁ OnHex (30) < OiPr (4) < OPh
(31), which is consistent with the reported SAR of acetamide-based
ACC inhibitors (OEt ꢂ OiPr 6 OPh).18,24 Agonistic activities towards
PPARs were also evaluated, and we found that 28, 29 and 32 were
active at submicromolar concentrations. Compounds 29 and 32
were identified as more potent dual modulators of ACC2 and
PPAR
weak (24% inhibition at 30
seen at concentrations up to 20
a
/d dual agonistic activity, although the former activity was
M). PPAR agonistic activity was not
M.
l
c
l
PPARa/d. Further, their maximum efficacies towards PPARa and
2.3. Structure–activity investigations of the dual modulator 6
PPARd were similar to those of compound 6. It is interesting that
structurally similar compounds exhibited dramatically different
activities (Table 2). For example, compounds 6 and 28 are PPAR-
selective, whereas the phenoxy analogue (31) exhibits selective
ACC inhibition. These compounds might be useful tools to clarify
the roles of ACC and PPAR in lipid metabolism.
We next focused on the methylene bridge between the aceta-
mide and the oxadiazole. Compounds 38 and 39 were prepared
by coupling with N-acetylglycine (Scheme 5), and their activities
To improve the ACC2-inhibitory activity and the PPAR /d dual
a
agonistic activity of 6, we initially focused on exploring sub-
stituents at the 4-position of the phenoxyphenyl group because it
is known that terminal alkoxy/phenoxy group affects greatly to the
ACC-inhibitory activity.19,24 For this purpose, compounds 28–32
were prepared using a similar route to that in Scheme 1 (Scheme 4),
and the obtained compounds were evaluated for ACC2-inhibitory
Scheme 4. Reagents and conditions: (a) Me2SO4 (for 28), K2CO3 (for 28), EtI (for 29), Cs2CO3 (for 29), DMF, 90 °C, 23% (for 28), 83% (for 29); (b) 4-fluorobenzonitrile, K2CO3,
DMF, 150 °C, 99% (for 33), 81% (for 34); (c) aq NH2OH, EtOH, reflux, 73% (for 35), 79% (for 36), 96% (for 37); (d) N-acetyl-DL-alanine, HATU, DIPEA, DMF, rt to 90 °C, 47% (for 30),
92% (for 31), 87% (for 32).
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S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
5
Table 2
SAR investigations of substituents at the 4-position of the phenoxyphenyl group
Compound
R
ACC2a
PPARab
PPARcb
PPARdb
IC50
[l
M] (inhibition [%])c,d
EC50
[l
M]c
EC50
[l
M]c
EC50 [l
M]c
28
29
30
31
32
6
OMe
OEt
OnHex
OPh
H
(17)
0.21
0.33
ia.e
ia.e
ia.e
ia.e
n.t.e
ia.e
ia.e
0.39
0.37
ia.e
0.42 (35)
0.26 (93)
0.048 (76)
0.29 (45)
(24)
ia.e
ia.e
0.32
1.0
0.32
1.0
OH
a
Inhibitory activity towards recombinant human ACC2.
Agonistic activities towards PPAR(LBD)-GAL4 chimeric receptors in transiently transfected HEK-293 cells. All active compounds showed partial agonistic properties, as
b
did compound 6 (see Table 1).
c
Evaluations were performed in triplicate, and the EC50 and IC50 values were determined from the dose–response curves.
d
Percent inhibition at 30
l
M is shown in parenthesis. The maximum ACC2 inhibition by 1 was defined as 100%.
e
ia. = inactive at <20
lM. n.t. = not tested.
Scheme 5. Reagents and conditions: (a) N-acetylglycine, HATU, DIPEA, DMF, rt to 90 °C, 76%; (b) BBr3, CH2Cl2, ꢀ50 to ꢀ20 °C, 92% (for 38); (c) N-acetylglycine, HATU, DIPEA,
DMF, rt to 90 °C, 62% (for 39).
Table 3
SAR investigations on the methylene bridge between acetamide and oxadiazole
Compound
R1
R2
ACC2a
PPARa b
PPARcb
PPARdb
IC50
[l
M] (inhibition [%])c,d
EC50
[l
M]c
EC50
[l
M]c
EC50 [l
M]c
38
6
39
32
H
Me
H
OH
OH
H
(3)
ia.e
1.0
n.t.e
ia.e
n.t.e
1.0
(24)
ia.e
ia.e
0.32
n.t.e
ia.e
n.t.e
0.32
Me
H
0.29 (45)
a
Inhibitory activity towards recombinant human ACC2.
Agonistic activities towards PPAR(LBD)-GAL4 chimeric receptors in transiently transfected HEK-293 cells. All active compounds showed partial agonistic properties, as
b
did compound 6 (see Table 1).
c
Evaluations were performed in triplicate, and the EC50 and IC50 values were determined from the dose–response curves.
d
Percent inhibition at 30
lM is shown in parenthesis. The maximum ACC2 inhibition by 1 was defined as 100%.
e
ia. = inactive at <20
l
M (for PPAR) or <30 M (for ACC). n.t. = not tested.
l
were evaluated (Table 3). Conversion of the monomethyl methy-
lene bridge (–CHMe–) to methylene (–CH2–) considerably
decreased the PPARa-agonistic activity as well as the ACC2-inhibi-
inhibitors.18,24 However, it is noteworthy that (S)-32 exhibited
PPAR /d dual agonistic activities, whereas (R)-32 did not. Thus,
among the synthesized compounds, (S)-32 was the most potent
a
tory activity. The results indicated that the methyl group at the
methylene bridge is critical for both ACC-inhibitory activity and
PPAR-agonistic activity.
dual modulator of ACC2 and PPARa/d.
3. Conclusions
Next, we synthesized both enantiomers of the most potent
ACC2 inhibitor 31 and dual modulator 32 by using commercially
available chiral N-Boc-alanines (Scheme 6). The (S)-isomers of 31
and 32 were found to show higher ACC2-inhibitory activities than
the corresponding (R)-isomers (Table 4). This result is consistent
with previous reports on other acetamide-based ACC
We designed a series of candidate dual modulators of ACC and
PPAR based on the molecular similarity between known ACC inhi-
bitors and PPAR agonists. Among the synthesized compounds, 6
showed weak ACC2-inhibitory activity together with PPAR
a/d dual
agonistic activity. Structure-activity investigations and structural
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S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
Scheme 6. Reagents and conditions: (a) N-(tert-butoxycarbonyl)-D-alanine, HATU, DIPEA, DMF, rt to 90 °C, 65% (for (R)-41), 69% (for (R)-42); (b) TFA, CH2Cl2, 0 °C to rt, quant.
(for (R)-43), 90% (for (R)-44); (c) Ac2O, pyridine, CH2Cl2, 0 °C to rt, 73% (for (R)-31), 90% (for (R)-32); (d) N-(tert-butoxycarbonyl)-
L-alanine, HATU, DIPEA, DMF, rt to 90 °C, 69%
(for (S)-41), 61% (for (S)-42); (e) TFA, CH2Cl2, 0 °C to rt, quant. (for (S)-43), 98% (for (S)-44); (f) Ac2O, pyridine, CH2Cl2, 0 °C to rt, 74% (for (S)-31), 79% (for (S)-32).
Table 4
SAR investigations on the stereochemistry of the methyl group near acetamide
Compound
Me
R
ACC2a
IC50
PPARab
PPARcb
PPARdb
[
l
M] (inhibition [%])c,d
EC50
[l
M]c
EC50
[
l
M]c
EC50 [l
M]c
31
rac
R
S
rac
R
S
OPh
OPh
OPh
H
H
H
0.048 (76)
7.5 (67)
0.016 (86)
0.29 (45)
(37)
ia.e
ia.e
ia.e
(R)-31
(S)-31
32
(R)-32
(S)-32
ia.e
n.t.e
n.t.e
ia.e
ia.e
ia.e
ia.e
0.32
0.32
ia.e
n.t.e
n.t.e
ia.e
0.22 (65)
0.15
0.29
a
Inhibitory activity towards recombinant human ACC2.
Agonistic activities towards PPAR(LBD)-GAL4 chimeric receptors in transiently transfected HEK-293 cells. All active compounds showed partial agonistic properties, as
b
did compound 6 (see Table 1).
c
Evaluations were performed in triplicate, and the EC50 and IC50 values were determined from the dose–response curves.
d
Percent inhibition at 30
lM is shown in parenthesis. The maximum ACC2 inhibition of 1 was defined as 100%.
e
ia. = inactive at <20
lM. n.t. = not tested.
elaboration of 6 led us to the more potent dual modulator 32 and
the ACC2-selective inhibitor 31. The (S)-isomers of 32 and 31 were
Bands were visualized using UV light or by application of appro-
priate reagents followed by heating. Flash chromatography was
identified as the eutomers for ACC2 inhibition and PPAR
a
/d activa-
M, EC50
M]. Importantly,
carried out with silica gel (Silica gel 60N, 40–50 lm particle size)
tion [(S)-32: IC50 (ACC2) = 0.22
l
M, EC50 (PPAR
a
) = 0.15
l
purchased from Kanto Chemical. NMR spectra were recorded on a
JEOL JNM-ECA500 (500 MHz) spectrometer, operating at 500 MHz
for 1H NMR and at 125 MHz for 13C NMR. Proton and carbon
chemical shifts are expressed in d values (ppm) relative to inter-
nal tetramethylsilane (0.00 ppm), residual CHCl3 (7.26 ppm),
CHD2OD (3.31 ppm) or C2HD5SO (2.49 ppm) for 1H NMR, and
internal tetramethylsilane (0.00 ppm) or CDCl3 (77.16 ppm),
methanol-d4 (49.00 ppm) or dimethyl sulfoxide-d6 (39.50 ppm)
for 13C NMR. Data are reported as follows: chemical shift, multi-
plicity (s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet;
br, broad), coupling constants (Hz), integration. High-resolution
mass spectra were recorded using a Bruker micrOTOF II mass
spectrometer. Fast atom bombardment (FAB) mass spectra were
recorded on a JEOL JMA-HX110 mass spectrometer with m-
nitrobenzyl alcohol as a matrix.
(PPARd) = 0.29 M; (S)-31: IC50 (ACC2) = 0.016
l
l
some of our compounds show distinct activity profiles; for exam-
ple, compound 31 selectively inhibits ACC2, whereas compound
28 mainly exhibits PPARa/d agonistic activity. These compounds
are expected to be useful tools for investigating the appropriate
balance of ACC inhibition and PPAR activation to achieve therapeu-
tic lipid control.25
4. Experimental
4.1. Chemistry
4.1.1. General
All chemical reagents and solvents were purchased from
Sigma–Aldrich, Kanto Chemical, Tokyo Chemical Industry and
Wako Pure Chemical Industries, and used without further purifi-
cation. Moisture-sensitive reactions were performed under an
atmosphere of argon, unless otherwise noted, and monitored by
thin-layer chromatography (TLC, Merck silica gel 60 F254 plate).
4.1.2. General procedure A: coupling reaction of phenol and 4-
fluorobenzonitrile
To a solution of phenol (1.2 equiv) and 4-fluorobenzonitrile
(1.0 equiv) in anhydrous DMF was added K2CO3 (1.2 equiv). The
Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045

S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
7
mixture was stirred at 150 °C for 2–9 h, then poured into ice-cold
water, and extracted with CH2Cl2 or AcOEt. The combined organic
layer was washed with water, dried over Na2SO4 and concentrated
under reduced pressure. The residue was purified by silica gel col-
umn chromatography to afford the product.
(125 MHz, CDCl3) d 162.7, 155.4, 147.8, 134.2, 121.9, 119.1,
117.4, 117.3, 105.4, 70.7, 22.2; MS (FAB) m/z 254 (M+H)+.
4.1.10. 4-(4-Benzyloxyphenoxy)benzonitrile (13)
General procedure A: A white solid (97%); 1H NMR (500 MHz,
CDCl3) d 7.57 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 7.46–7.40 (m, 4H),
7.33–7.37 (m, 1H), 7.01 (s, 4H), 6.96 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H),
5.08 (s, 2H); 13C NMR (125 MHz, CDCl3) d 162.5, 156.3, 148.2,
136.8, 134.2, 128.8, 128.3, 127.6, 121.9, 119.1, 117.3, 116.4,
105.4, 70.6; MS (FAB) m/z 302 (M+H)+.
4.1.3. General procedure B: reaction of benzonitrile and
hydroxylamine
To a solution of benzonitrile (1.0 equiv) in EtOH was added 50%
aq hydroxylamine (4.0–5.0 equiv). The mixture was refluxed for 1–
3 h, and then concentrated under reduced pressure. The residue
was purified by silica gel column chromatography to afford the
product.
4.1.11. 4-(4-Isopropoxyphenoxy)benzamidoxime (14)
General procedure B: An yellow solid (58%); 1H NMR (500 MHz,
CDCl3) d 7.56 (ddd, J = 9.7, 2.6, 2.6 Hz, 2H), 6.98–6.93 (m, 4H), 6.88
(ddd, J = 9.0, 2.3, 2.3 Hz, 2H), 4.84 (s, 2H), 4.50 (sep, J = 5.7 Hz, 1H),
1.34 (d, J = 5.7 Hz, 6H); 13C NMR (125 MHz, CDCl3) d 160.3, 154.7,
152.5, 149.4, 127.5, 126.6, 121.3, 117.4, 117.3, 70.7, 22.2; MS
(FAB) m/z 287 (M+H)+.
4.1.4. General procedure C: condensation of amidoxime and
amino acid
A solution of amino acid (1.5 equiv), HATU (1.5 equiv) and DIEA
(1.5 equiv) in anhydrous DMF was stirred at room temperature for
15–60 min. Amidoxime (1.0 equiv) was then added, and the result-
ing mixture was further stirred at room temperature for 25–
120 min and then at 80–100 °C for 0.5–9 h. After the reaction,
the mixture was diluted with AcOEt and washed with aq NaHCO3.
The organic layer was dried over Na2SO4, and the solvent was
removed under reduced pressure. The residue was purified by sil-
ica gel column chromatography to afford the product.
4.1.12. 4-(4-Benzyloxyphenoxy)benzamidoxime (15)
General procedure B: A white solid (52%); 1H NMR (500 MHz,
DMSO-d6) d 9.54 (s, 1H), 7.63 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.47–
7.44 (m, 2H), 7.41–7.38 (m, 2H), 7.35–7.31 (m, 1H), 7.06–7.00
(m, 4H), 6.88 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H) 5.78 (s, 2H), 5.08 (s,
2H); 13C NMR (125 MHz, DMSO-d6) d 158.6, 154.9, 150.4, 149.2,
137.0, 128.5, 127.9, 127.7, 127.1, 120.9, 116.6, 116.1, 69.6; MS
(FAB) m/z 335 (M+H)+.
4.1.5. General procedure D: removal of Bn group
BBr3 (0.2–0.4 equiv, 1 M in CH2Cl2) was slowly added to a solu-
tion of benzyloxy compound (1.0 equiv) in anhydrous CH2Cl2 at
ꢀ50 °C, and the mixture was stirred at ꢀ20 °C for 1.5 h. The reac-
tion was then quenched by addition of water. The resulting mix-
ture was warmed to ambient temperature, diluted with CH2Cl2
and aq HCl, and extracted with CH2Cl2. The combined organic layer
was dried over Na2SO4 and concentrated under reduced pressure.
The residue was purified by silica gel column chromatography to
afford the product.
4.1.13. N-(1-(3-(4-(4-Isopropoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (4)
General procedure C: A white solid (46%); 1H NMR (500 MHz,
CDCl3) d 7.99 (ddd, J = 9.2, 2.5, 2.5 Hz, 2H), 7.00 (ddd, J = 9.4, 2.0,
2.0 Hz, 4H), 6.90 (ddd, J = 9.2, 2.9, 2.9 Hz, 2H), 6.23 (d, J = 6.9 Hz,
1H), 5.47 (dq, J = 7.4, 6.9 Hz, 1H), 4.51 (sep, J = 6.3 Hz, 1H), 2.09
(s, 3H), 1.63 (d, J = 7.4 Hz, 3H), 1.35 (d, J = 6.3 Hz, 6H); 13C NMR
(125 MHz, CDCl3) d 179.5, 169.6, 168.0, 161.5, 154.9, 148.9,
129.3, 121.6, 120.4, 117.4, 117.3, 70.7, 43.0, 23.3, 22.2, 20.1; ESI-
TOF-HRMS calcd for C21H24N3O4 (m/z) [M+Na]+ 404.1581, found
404.1583.
4.1.6. General procedure E: removal of SEM group
To a solution of SEM-protected compound (1.0 equiv) in THF
was added TBAF (3.0 equiv, 1 M in THF). The mixture was refluxed
for 11–20 h, then cooled to room temperature, and concentrated
under reduced pressure. The residue was purified by silica gel col-
umn chromatography or PTLC to afford the product.
4.1.14. N-((3-(4-(4-Isopropoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)methyl)acetamide (5)
General procedure C: A white solid (65%); 1H NMR (500 MHz,
CDCl3) d 7.98 (dd, J = 8.9, 1.4 Hz, 2H), 7.00 (d, J = 9.2 Hz, 4H), 6.90
(ddd, J = 9.2, 2.9, 2.9 Hz, 2H), 6.25 (s, 1H), 4.74 (dd, J = 5.7, 1.1 Hz,
2H), 4.51 (sep, J = 6.3 Hz, 1H), 2.12 (d, J = 1.1 Hz, 3H), 1.35 (d,
J = 6.3 Hz, 6H); 13C NMR (125 MHz, CDCl3) d 175.9, 170.3, 168.1,
161.6, 154.9, 148.9, 129.3, 121.6, 120.3, 117.4, 117.3, 70.7, 36.0,
32.1, 22.2; ESI-TOF-HRMS calcd for C20H21N3O4 (m/z) [M+Na]+
390.1424, found 390.1423.
4.1.7. General procedure F: removal of Boc group
To a solution of Boc-protected compound (1.0 equiv) in CH2Cl2
was added TFA (10 equiv) at 0 °C. The mixture was stirred at room
temperature for 1–5 h, and then concentrated under reduced pres-
sure to afford the product.
4.1.8. General procedure G: acetylation of amine
To a solution of amine (1.0 equiv) in CH2Cl2 was added pyri-
dine (5.0 equiv) and acetic anhydride (1.2–2.0 equiv) at 0 °C.
The mixture was stirred at room temperature for 2–14 h, and
then diluted with CH2Cl2 and 0.5 N aq HCl. The organic layer
was washed with 0.5 N aq HCl and saturated aq NaHCO3, dried
over Na2SO4 and concentrated under reduced pressure. The resi-
due was purified by recrystallization (n-hexane/AcOEt) to afford
the product.
4.1.15. N-(1-(3-(4-(4-Benzyloxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (16)
General procedure C: A yellow solid (70%); 1H NMR (500 MHz,
CDCl3) d 7.99 (ddd, J = 8.6, 2.9, 2.9 Hz, 2H), 7.47–7.43 (m, 2H),
7.42–7.39 (m, 2H), 7.36–7.33 (m, 1H), 7.03–6.98 (m, 6H), 6.15 (d,
J = 8.0 Hz, 1H), 5.46 (dq, J = 7.4, 7.4 Hz, 1H), 5.07 (s, 2H), 2.09 (s,
3H), 1.63 (d, J = 7.4 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 179.3,
169.5, 167.8, 161.3, 155.6, 149.1, 136.8, 129.2, 128.6, 128.1, 127.5,
121.4, 120.3, 117.3, 116.0, 70.5, 42.8, 23.1, 20.0; ESI-TOF-HRMS
calcd for C25H23N3O4 (m/z) [M+Na]+ 452.1581, found 452.1605.
4.1.9. 4-(4-Isopropoxyphenoxy)benzonitrile (12)
General procedure A: A white solid (97%); 1H NMR (500 MHz,
CDCl3) d 7.57 (ddd, J = 9.2, 2.6, 2.6 Hz, 2H), 7.00–6.97 (m, 2H),
6.96 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 6.92 (ddd, J = 9.2, 3.1, 3.1 Hz,
2H), 4.52 (sep, J = 6.3 Hz, 1H), 1.35 (d, J = 6.3 Hz, 6H); 13C NMR
4.1.16. N-(1-(3-(4-(4-Hydroxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (6)
General procedure D: A white solid (91%); 1H NMR (500 MHz,
CD3OD) d 7.96 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 6.99 (ddd, J = 9.2, 2.9,
Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045

8
S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
2.9 Hz, 2H), 6.96 (ddd, J = 8.6, 2.3, 2.3 Hz, 2H), 6.85 (ddd, J = 8.6, 2.3,
2.3 Hz, 2H), 5.27 (q, J = 6.9 Hz, 1H), 2.01 (s, 3H), 1.60 (d, J = 6.9 Hz,
3H); 13C NMR (125 MHz, CD3OD) d 181.5, 173.0, 169.1 163.2, 155.7,
149.2, 130.1, 122.7, 121.5, 117.9, 117.4, 44.0, 22.3, 18.8; ESI-TOF-
J = 6.0 Hz, 6H), 0.99–0.93 (m, 2H), ꢀ0.01 to ꢀ0.04 (m, 9H); ESI-
TOF-HRMS calcd for C23H29N3O3Si (m/z) [M+Na]+ 446.1870, found
446.1870.
HRMS calcd for
362.1138.
C
18H17N3O4 (m/z) [M+Na]+ 362.1111, found
4.1.20. 5-Cyano-2-phenoxy-1-((2-(trimethylsilyl)ethoxy)methy-
l)benzimidazole and 6-cyano-2-phenoxy-1-((2-(trimethylsilyl)
ethoxy)methyl)benzimidazole (19)
4.1.17. N-(1-(3-(4-(4-Propoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (7)
To a solution of 6 (75 mg, 0.22 mmol) and Cs2CO3 (120 mg,
0.38 mmol) in anhydrous DMF (1.0 mL) was added 1-iodopropane
To a solution of 17 (0.5 g, 1.6 mmol) and phenol (230 mg,
2.4 mmol) in anhydrous DMF (5.4 mL) was added Cs2CO3 (1.6 g,
4.9 mmol), and the mixture was stirred at room temperature for
3 h. The reaction was then quenched by addition of water, and
the resulting mixture was extracted with AcOEt. The combined
organic layer was washed with brine, dried over Na2SO4 and con-
centrated under reduced pressure. The residue was purified by sil-
ica gel column chromatography (n-hexane/AcOEt = 3:1) to afford
the product 19 (0.59 g, 99%, 1:1 regioisomeric mixture) as a white
solid.
(27 lL, 0.28 mmol). The mixture was heated at 80 °C for 5 h, and
then cooled to room temperature. The reaction was quenched by
addition of water and the resulting mixture was extracted with
AcOEt. The combined organic layer was washed with brine, dried
over Na2SO4 and concentrated under reduced pressure. The residue
was purified by silica gel column chromatography (CHCl3/
AcOEt = 3:2) to afford the product 7 (79 mg, 94%) as a white solid.
1H NMR (500 MHz, CDCl3) d 7.99 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H),
7.02–6.91 (m, 4H), 6.90–6.94 (m, 2H), 6.16 (d, J = 7.4 Hz, 1H),
5.49–5.44 (m 1H), 3.93 (t, J = 6.6 Hz, 2H), 2.09 (s, 3H), 1.86–1.79
(m, 2H), 1.64 (d, J = 6.9 Hz, 3H), 1.06 (t, J = 7.4 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 179.3, 169.4, 167.8, 161.4, 156.1, 148.7,
129.2, 121.4, 120.2, 117.2, 115.6, 70.0, 42.8, 23.2, 22.6, 20.0, 10.5;
ESI-TOF-HRMS calcd for C21H23N3O4 (m/z) [MꢀH]ꢀ 380.1605,
found 380.1609.
1H NMR (500 MHz, CDCl3) d 7.84 (s, 1/2H), 7.70 (d, J = 1.1 Hz,
1/2H), 7.59 (d, J = 8.0 Hz, 1/2H), 7.50–7.46 (m, 7/2H), 7.37–7.31
(m, 3H), 5.58 (s, 1H), 5.58 (s, 1H), 3.69–3.64 (m, 2H), 0.99–0.93
(m, 2H), ꢀ0.02 (s, 9H); 13C NMR (125 MHz, CDCl3) d 157.9, 157.3,
152.9, 143.4, 139.8, 136.3, 133.0, 130.1, 129.6, 126.7, 126.4,
126.4, 126.0, 123.0, 120.3, 120.2, 119.9, 119.4, 115.3, 113.7,
110.2, 105.8, 104.8, 71.8, 71.8, 67.0, 17.7, ꢀ1.5; MS (FAB) m/z 366
(M+H)+.
4.1.21. 2-(4-Isopropoxyphenoxy)-1-((2-(trimethylsilyl)ethoxy)
methyl)benzimidazole-5-amidoxime and 2-(4-isopropoxyphe-
noxy)-1-((2-(trimethylsilyl)ethoxy)methyl)benzimidazole-6-
amidoxime (20)
4.1.18. 2-Chloro-5-cyano-1-((2-(trimethylsilyl)ethoxy)methyl)
benzimidazole and 2-chloro-6-cyano-1-((2-(trimethylsilyl)
ethoxy)methyl)benzimidazole (17)
To
a
solution of 2-chloro-5-cyanobenzimidazole (1.4 g,
General procedure B: A white solid (87%, 1:1 regioisomeric mix-
ture); 1H NMR (500 MHz, CDCl3) d 7.88 (dd, J = 5.4, 1.4 Hz, 1/2H),
7.70 (d, J = 1.7 Hz, 1/2H), 7.57–7.53 (m, 1H), 7.48 (dd, J = 8.3,
1.4 Hz, 1/2H), 7.37 (d, J = 8.0 Hz, 1/2H), 7.27–7.23 (m, 2H), 6.96–
6.91 (m, 2H), 5.54 (s, 1H), 5.54 (s, 1H), 4.93 (s, 1H), 4.86 (s, 1H),
4.56–4.49 (m, 1H), 3.68–3.64 (m, 2H), 1.35 (d, J = 6.3 Hz, 3H),
1.35 (d, J = 6.3 Hz, 3H), 0.95 (t, J = 6.9 Hz, 1H), 0.95 (t, J = 6.9 Hz,
8.1 mmol) and triethylamine (3.4 mL, 24 mmol) in anhydrous
DMF (30 mL) was added 2-(trimethylsilyl)ethoxymethyl chloride
(2.2 mL, 12 mmol), and the mixture was stirred at room tempera-
ture for 2.5 h. Triethylamine (1.7 mL, 16 mmol) was further added,
and stirring was continued overnight. The reaction was then
quenched by addition of water, and the resulting mixture was
extracted with AcOEt. The combined organic layer was washed
with 2 N aq HCl and brine, dried over Na2SO4, and concentrated
under reduced pressure. The residue was purified by silica gel col-
umn chromatography (n-hexane/AcOEt = 3:1) to afford the product
17 (2.2 g, 87%, 1:1 regioisomeric mixture) as a white solid.
1H), ꢀ0.03 (s, 9/2H), ꢀ0.03 (s, 9/2H); ESI-TOF-HRMS calcd for C23
-
H
32N4O4Si (m/z) [M+Na]+ 457.2266, found 457.2291.
4.1.22. 2-Phenoxy-1-((2-(trimethylsilyl)ethoxy)methyl)benzim-
idazole-5-amidoxime and 2-phenoxy-1-((2-(trimethylsilyl)ethoxy)
methyl)benzimidazole-6-amidoxime (21)
1H NMR (500 MHz, CDCl3) d 8.03 (s, 1/2H), 7.81 (d, J = 1.1 Hz,
1/2H), 7.77 (d, J = 8.6 Hz, 1/2H), 7.61–7.55 (m, 3/2H), 5.59 (s, 2H),
3.60–3.55 (m, 2H), 0.95–0.89 (m, 2H), ꢀ0.03 (s, 9H); 13C NMR
(125 MHz, CDCl3) d 144.6, 144.1, 143.3, 141.3, 137.6, 134.5,
127.2, 126.9, 124.3, 120.5, 119.2, 115.0, 111.3, 73.6, 73.5, 67.2,
17.7, ꢀ1.5; MS (FAB) m/z 308 (M+H)+.
General procedure B: A white solid (93%, 1:1 regioisomeric mix-
ture); 1H NMR (500 MHz, CDCl3) d 7.93 (d, J = 1.1 Hz, 1/2H), 7.75 (d,
J = 1.1 Hz, 1/2H), 7.60 (dd, J = 8.6, 2.6 Hz, 1H), 7.54–7.48 (m, 5/2H),
7.44–7.40 (m, 5/2H), 7.33–7.30 (m, 1H), 5.60 (s, 1H), 5.60 (s, 1H),
4.98 (s, 1H), 4.91 (s, 1H), 3.72–3.68 (m, 2H), 1.01–0.97 (m, 2H),
0.00 (s, 9H); 13C NMR (125 MHz, CDCl3) d 156.6, 156.3, 153.2,
141.3, 139.7, 134.4, 133.2, 129.9, 127.3, 126.6, 125.9, 120.4,
120.2, 120.2, 118.6, 116.4, 109.4, 107.3, 71.6, 71.5, 66.7, 66.6,
17.7, 17.7, ꢀ1.4; MS (FAB) m/z 399 (M+H)+.
4.1.19. 5-Cyano-2-(4-isopropoxyphenoxy)-1-((2-(trimethylsilyl)
ethoxy)methyl)benzimidazole and 6-cyano-2-(4-isopropoxyph-
enoxy)-1-((2-(trimethylsilyl)ethoxy)methyl)benzimidazole (18)
To a solution of 17 (0.60 g, 2.0 mmol) and 4-isopropoxyphenol
(0.46 g, 3.0 mmol) in DMF (10 mL) was added Cs2CO3 (1.9 g,
5.9 mmol), and the mixture was stirred at room temperature
overnight. The reaction was quenched by addition of water, and
the resulting mixture was extracted with AcOEt. The combined
organic layer was washed with brine, dried over Na2SO4 and con-
centrated under reduced pressure. The residue was purified by
silica gel column chromatography (n-hexane/AcOEt = 3:1) to
afford the product 18 (0.83 g, 99%, 1:1 regioisomeric mixture) as
a pink oil.
4.1.23. N-((3-(2-(4-Isopropoxyphenoxy)-1-((2-(trimethylsilyl)
ethoxy)methyl)-1H-benzo[d]imidazol-5-yl)-1,2,4-oxadiazol-5-
yl)methyl)acetamide and N-((3-(2-(4-isopropoxyphenoxy)-1-
((2-(trimethylsilyl)ethoxy)methyl)-1H-benzo[d]imidazol-6-yl)-
1,2,4-oxadiazol-5-yl)methyl)acetamide (22)
General procedure C: A colorless oil (quant., 1:1 regioisomeric
mixture); 1H NMR (500 MHz, CD3OD) d 8.18 (d, J = 1.7 Hz, 1/2H),
8.10 (d, J = 1.1 Hz, 1/2H), 7.97–7.92 (m, 1H), 7.61 (d, J = 8.6 Hz,
1H), 7.50 (d, J = 8.6 Hz, 1H), 7.29–7.25 (m, 2H), 7.02 (d, J = 9.0 Hz,
2H), 5.67 (s, 1H), 5.65 (s, 1H), 4.69 (s, 1H), 4.67 (s, 1H), 4.65–4.58
(m, 1H), 3.76–3.72 (m, 2H), 2.07 (s, 3/2H), 2.05 (s, 3/2H), 1.34 (d,
J = 6.3 Hz, 6H), 0.96 (t, J = 7.9, 1H), 0.94 (t, J = 7.9 Hz, 1H), ꢀ0.05
(s, 9H).
1H NMR (500 MHz, CDCl3) d 7.83 (s, 1/2H), 7.68 (d, J = 1.1 Hz,
1/2H), 7.59 (d, J = 8.0 Hz, 1/2H), 7.51–7.47 (m, 1H), 7.44 (d,
J = 8.0 Hz, 1/2H), 7.26–7.23 (m, 2H), 6.96–6.93 m, 2H), 5.56 (s,
1H), 5.56 (s, 1H), 4.56–4.51 (m, 1H), 3.68–3.64 (m, 2H), 1.36 (d,
Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045

S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
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4.1.24. N-(1-(3-(2-Phenoxy-1-((2-(trimethylsilyl)ethoxy)
methyl)-1H-benzo[d]imidazol-5-yl)-1,2,4-oxadiazol-5-yl)ethyl)
acetamide and N-(1-(3-(2-phenoxy-1-((2-(trimethylsilyl)
ethoxy)methyl)-1H-benzo[d]imidazol-6-yl)-1,2,4-oxadiazol-5-
yl)ethyl)acetamide (23)
General procedure C: A yellow oil (quant. 1:1 regioisomeric mix-
ture); 1H NMR (500 MHz, CDCl3) d 8.32 (s, 1/2H), 8.16 (s, 1/2H),
7.99 (dd, J = 8.3, 1.4 Hz, 1H), 7.66 (d, J = 8.6 Hz, 1/2H), 7.51–7.49
(m, 5/2H), 7.44–7.41 (m, 2H), 7.35 (dd, J = 7.4, 6.9 Hz, 1H), 6.34
(d, J = 5.2 Hz, 1H), 5.65 (s, 1H), 5.62 (s, 1H), 5.55–5.47 (m, 1H),
3.75–3.70 (m, 2H), 2.14 (s, 3/2H), 2.12 (s, 3/2H), 1.69 (d,
J = 6.9 Hz, 3/2H), 1.67 (d, J = 6.9 Hz, 3/2H), 1.03–0.97 (m, 1H),
0.02 (s, 9H); MS (FAB) m/z 494 (M+H)+.
1H NMR (500 MHz, CDCl3) d 7.50 (d, J = 8.0 Hz, 1/2H), 7.45 (d,
J = 1.1 Hz, 1/2H), 7.33 (d, J = 8.0 Hz, 1/2H), 7.31 (d, J = 1.1 Hz,
1/2H), 7.25–7.21 (m, 2H), 7.15 (dd, J = 8.0, 2.0 Hz, 1/2H), 7.11 (dd,
J = 8.0, 1.5 Hz, 1/2H), 6.95–6.91 (m, 2H), 5.76 (br s, 1/2H), 5.72
(br s, 1/2H), 5.52 (s, 1H), 5.52 (s, 1H), 4.54–4.50 (m, 2H), 4.50–
4.47 (m, 1H), 3.66 (t, J = 8.3 Hz, 1H), 3.64 (t, J = 8.3 Hz, 1H), 2.03
(s, 3/2H), 2.00 (s, 3/2H), 1.35 (d, J = 6.6 Hz, 6H), 0.95 (t, J = 8.8 Hz,
1H), 0.94 (t, J = 8.5 Hz, 1H), ꢀ0.03 (s, 9H); ESI-TOF-HRMS calcd
for C25H35N3O4Si (m/z) [M+Na]+ 492.2289, found 492.2265.
4.1.29. N-((2-(4-Isopropoxyphenoxy)-1H-benzo[d]imidazol-5-
yl)methyl)acetamide (10)
General procedure E: A white solid (55%); 1H NMR (500 MHz,
CD3OD) d 7.30–7.27 (m, 2H), 7.19 (d, J = 6.4 Hz, 2H), 7.08 (d,
J = 8.0 Hz, 1H), 6.97 (d, J = 6.4 Hz, 2H), 4.61–4.56 (m, 1H), 4.40 (s,
2H), 1.97 (s, 3H), 1.32 (d, J = 5.7 Hz, 6H); ESI-TOF-HRMS calcd for
4.1.25. N-(1-(3-(2-(4-Isopropoxyphenoxy)-1H-benzo[d]
imidazol-5-yl)-1,2,4-oxadiazol-5-yl)ethyl)acetamide (8)
General procedure E: A white solid (13%); 1H NMR (500 MHz,
CD3OD) d 8.05 (s, 1H), 7.87 (dd, J = 8.0, 1.1 Hz, 1H), 7.45 (d,
J = 8.0 Hz, 1H), 7.26–7.22 (m, 2H), 7.02–6.98 (m, 2H), 4.67 (s,
2H), 4.65–4.58 (m, 1H), 2.06 (s, 3H), 1.34 (d, J = 6.3 Hz, 6H); ESI-
C
19H21N3O3 (m/z) [M+Na]+ 362.1475, found 362.1481.
4.1.30. Phenyl (4-cyanophenyl)carbamate (26)
To a solution of 4-aminobenzonitrile (440 mg, 3.7 mmol) and
pyridine (0.60 mL, 7.5 mmol) in THF (4.4 mL) was added
phenylchlorocarbonate (0.52 mL, 4.2 mmol) at 0 °C. The mixture
was stirred at room temperature for 2 h, then diluted with water,
and extracted with CH2Cl2. The combined organic layer was
washed with 1 N aq HCl and dried over Na2SO4. The solvent was
removed under reduced pressure to afford the product 26
(880 mg, 98%).
TOF-HRMS calcd for
407.1563.
C21H21N5O4 (m/z) [M] 407.1588, found
4.1.26. N-(1-(3-(2-Phenoxy-1H-benzo[d]imidazol-5-yl)-1,2,4-
oxadiazol-5-yl)ethyl)acetamide (9)
General procedure E: A white solid (22%); 1H NMR (500 MHz,
DMSO-d6)
d 8.73 (d, J = 7.4 Hz, 1H), 8.31 (s, 1H), 7.93 (d,
J = 1.1 Hz, 1H), 7.77 (dd, J = 8.0, 1.7 Hz, 1H), 7.50–7.47 (m, 3H),
7.42–7.40 (m, 2H), 7.32–7.29 (m, 1H), 5.21–5.15 (m, 1H), 1.89 (s,
3H), 1.52 (d, J = 7.4 Hz, 3H); 13C NMR (125 MHz, DMSO-d6) d
180.0, 168.8, 168.0 157.4, 153.3, 129.4, 124.8, 120.2, 119.7, 119.0,
114.5, 112.3, 41.8, 21.9, 21.8, 18.1; ESI-TOF-HRMS calcd for
1H NMR (500 MHz, CD3OD) d 7.71–7.66 (m, 4H), 7.41 (dd,
J = 8.0, 8.0 Hz, 2H), 7.26 (t, J = 7.0 Hz, 1H), 7.20 (d, J = 8.0 Hz, 2H);
13C NMR (125 MHz, CD3OD) d 151.3, 150.3, 141.7, 133.6, 129.7,
126.3, 121.6, 118.9, 118.7, 107.0; MS (FAB) m/z 239 (M+H)+.
C
19H17N5O3 (m/z) [M+Na]+ 386.1224, found 386.1242.
4.1.31. Phenyl (4-(N0-hydroxycarbamimidoyl)phenyl)carbamate
(27)
4.1.27. 5-Aminomethyl-2-(4-isopropoxyphenoxy)-1-((2-(trime-
thylsilyl)ethoxy)methyl)benzimidazole and 6-aminomethyl-2-
(4-isopropoxyphenoxy)-1-((2-(trimethylsilyl)ethoxy)methyl)
benzimidazole (24)
General procedure B: A white solid (29%); 1H NMR (500 MHz,
CD3OD) d 7.60 (d, J = 8.6 Hz, 2H), 7.54 (d, J = 8.6 Hz, 2H), 7.40 (dd,
J = 8.0, 8.0 Hz, 2H), 7.24 (t, J = 7.4 Hz, 1H), 7.19 (d, J = 8.0 Hz, 2H);
13C NMR (125 MHz, CD3OD) d 155.2, 154.0, 152.3, 141.3, 130.4,
129.0, 128.0, 126.7, 122.9, 119.4; MS (FAB) m/z 272 (M+H)+.
To a solution of 18 (0.21 g, 0.50 mmol) in 25% aq NH3 (1.6 mL)
and EtOH (16 mL) was added Raney Ni (1.6 mL, in water). The mix-
ture was stirred for 4 h under a hydrogen atmosphere, then filtered
through a Celite pad, and the filtrate was concentrated under
reduced pressure. The residue was purified by silica gel column
chromatography (CHCl3/MeOH = 8:1) to afford the product 24
(0.20 g, 97%, 1:1 regioisomeric mixture) as a yellow oil.
1H NMR (500 MHz, CDCl3) d 7.51 (s, 1/2H), 7.48 (d, J = 8.6 Hz,
1/2H), 7.43 (s, 1/2H), 7.31 (d, J = 8.0 Hz, 1/2H), 7.24–7.20 (m, 2H),
7.19–7.14 (m, 1H), 6.95–6.90 (m, 2H), 5.50 (s, 1H), 5.46 (s, 1H),
4.55–4.48 (m, 1H), 3.97 (s, 1H), 3.92 (s, 1H), 3.64 (t, J = 8.3 Hz,
1H), 3.63 (t, J = 8.0 Hz, 1H), 1.34 (d, J = 6.0 Hz, 3H), 1.34 (d,
J = 6.0 Hz, 3H), 0.93 (t, J = 8.0 Hz, 2H), ꢀ0.04 (s, 9/2H), ꢀ0.05 (s,
9/2H).
4.1.32. Phenyl (4-(5-(1-acetamidoethyl)-1,2,4-oxadiazol-3-yl)
phenyl)carbamate (11)
General procedure C: A white solid (78%); 1H NMR (500 MHz,
DMSO-d6)
d 10.59 (s, 1H), 8.72 (d, J = 6.9 Hz, 1H), 7.95 (d,
J = 8.6 Hz, 2H), 7.69 (d, J = 8.6 Hz, 2H), 7.43 (ddd, J = 8.6, 8.6,
2.3 Hz, 2H), 7.29–7.23 (m, 3H), 5.17 (dq, J = 7.4, 6.9 Hz, 1H), 1.88
(s, 3H), 1.51 (d, J = 7.4 Hz, 3H); 13C NMR (125 MHz, DMSO-d6) d
180.7, 169.2, 167.2, 151.6, 150.4, 141.7, 129.5, 128.0, 125.6,
122.0, 120.4, 118.6, 42.0, 22.3, 18.4; ESI-TOF-HRMS calcd for
C
19H18N4O4 (m/z) [M+Na]+ 389.1220, found 389.1192.
4.1.33. N-(1-(3-(4-(4-Methoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (28)
4.1.28. N-((2-(4-Isopropoxyphenoxy)-1-((2-(trimethylsilyl)
ethoxy)methyl)-1H-benzo[d]imidazol-5-yl)methyl)acetamide
and N-((2-(4-isopropoxyphenoxy)-1-((2-(trimethylsilyl)ethoxy)
methyl)-1H-benzo[d]imidazol-6-yl)methyl)acetamide (25)
To a solution of 24 (0.2 g, 0.46 mmol) in CH2Cl2 (4.0 mL) was
added triethylamine (0.13 mL, 0.92 mmol) and acetic anhydride
To a solution of 6 (41 mg, 0.12 mmol) and Me2SO4 (30 lL
0.31 mmol) in anhydrous DMF (0.80 mL) was added K2CO3 (36 mg,
0.26 mmol). The mixture was stirred at 90 °C for 4.5 h and then
cooled to room temperature. The reaction was quenched by addition
of water and the resulting mixture was extracted with AcOEt. The
combined organic layer was washed with brine, dried over Na2SO4
and concentrated under reduced pressure. The residue was purified
by silica gel column chromatography (CHCl3/MeOH = 10:1) to afford
the product 28 (10 mg, 23%) as a white solid.
(87 lL, 0.92 mmol). The mixture was stirred at room temperature
for 1 h, then diluted with CH2Cl2 and aq NaHCO3, and the organic
layer was washed with aq NaHCO3. After drying over Na2SO4, the
solvent was removed under reduced pressure. The residue was
purified by silica gel column chromatography (CHCl3/
MeOH = 10:1) to afford product 25 (0.18 g, 85%, 1:1 regioisomeric
mixture) as a yellow oil.
1H NMR (500 MHz, CDCl3) d 7.99 (d, J = 5.7 Hz, 2H), 7.05–6.92
(m, 6H), 6.13 (br s, 1H), 5.50–5.44 (m, 1H), 3.83 (s, 3H), 2.09 (s,
3H), 1.64 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 179.3,
169.4, 167.8, 161.4, 156.5, 148.9, 129.2, 121.5, 120.3, 117.2,
Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045

10
S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
115.0, 55.7, 42.8, 23.2, 20.1; ESI-TOF-HRMS calcd for C19H19N3O4
127.4, 127.1, 123.9, 119.4, 118.4; ESI-TOF-HRMS calcd for
C
13H12N2O2 (m/z) [M+H]+ 229.0972, found 229.0999.
(m/z) [M+Na]+ 376.1268, found 376.1286.
4.1.34. N-(1-(3-(4-(4-Ethoxyphenoxy)phenyl)-1,2,4-oxadiazol-5-
yl)ethyl)acetamide (29)
To a solution of 6 (83 mg, 0.24 mmol) and Cs2CO3 (140 mg,
0.43 mmol) in DMF (1.3 mL) was added iodoethane (25 lL,
4.1.40. N-(1-(3-(4-(4-Hexyloxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (30)
General procedure C: A white solid (47%); 1H NMR (500 MHz,
CDCl3) d 7.98 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.03–6.98 (m, 4H), 6.91
(ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 6.14 (d, J = 7.4 Hz, 1H), 5.47 (dq,
J = 7.4, 6.9 Hz, 1H), 3.96 (t, J = 6.6 Hz, 2H), 2.09 (s, 3H), 1.82–1.76
(m, 2H), 1.64 (d, J = 6.9 Hz, 3H), 1.49–1.46 (m, 2H), 1.38–1.34 (m,
4H), 0.93–0.90 (m, 3H); 13C NMR (125 MHz, CDCl3) d 179.5, 169.6,
168.0, 161.6, 156.2, 148.9, 129.3, 121.6, 120.4, 117.3, 115.8, 68.6,
43.0, 31.7, 29.4, 25.9, 23.3, 22.8, 20,1, 14.2; ESI-TOF-HRMS calcd
for C24H29N3O4 (m/z) [M+Na]+ 446.2050, found 446.2066.
0.31 mmol). The mixture was stirred at 80 °C for 5 h, and then
cooled to room temperature. The reaction was quenched by addi-
tion of water, and the resulting mixture was extracted with AcOEt.
The combined organic layer was washed with brine, dried over
Na2SO4 and concentrated under reduced pressure. The residue
was purified by silica gel column chromatography (CHCl3/
AcOEt = 3:2) to afford the product 29 (74 mg, 83%) as a white solid.
1H NMR (500 MHz, CDCl3) d 7.99 (dd, J = 9.4, 2.4 Hz, 2H), 7.03–
6.98 (m, 4H), 6.93–6.90 (m, 2H), 6.14 (d, J = 7.4 Hz, 1H), 5.49–5.44
(m, 1H), 4.04 (q, J = 6.9 Hz, 2H), 2.09 (s, 3H), 1.64 (d, J = 6.9 Hz, 3H),
1.43 (t, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 179.3, 169.4,
167.8, 161.4, 155.8, 148.8, 129.2, 121.4, 120.2, 117.2, 115.6, 63.9,
42.8, 23.2, 20.0, 14.9; ESI-TOF-HRMS calcd for C20H21N3O4 (m/z)
[MꢀH]ꢀ 366.1448 found 366.1432.
4.1.41. N-(1-(3-(4-(4-Phenoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethyl)acetamide (31)
General procedure C: A white solid (92%); 1H NMR (500 MHz,
CDCl3) d 8.02 (ddd, J = 8.6, 2.3, 2.3 Hz, 2H), 7.38–7.33 (m, 2H),
7.11 (t, J = 7.4 Hz, 1H), 7.07–7.02 (m, 8H), 6.13 (d, J = 8.0 Hz, 1H),
5.50–5.44 (m, 1H), 2.09 (s, 3H), 1.64 (d, J = 7.4 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 179.4, 169.4, 167.8, 160.7, 157.4, 153.6,
151.3, 129.8, 129.3, 123.3, 121.3, 120.8, 120.4, 118.6, 117.8, 42.9,
23.2, 20.0; ESI-TOF-HRMS calcd for C24H21N3O4 (m/z) [M+Na]+
438.1424, found 438.1450.
4.1.35. 4-(4-Hexyloxyphenoxy)benzonitrile (33)
General procedure A: A white solid (99%); 1H NMR (500 MHz,
CDCl3) d 7.57 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 6.99 (ddd, J = 9.2, 2.3,
2.3 Hz, 2H), 6.95 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 6.92 (ddd, J = 9.2,
2.3, 2.3 Hz, 2H), 3.96 (t, J = 6.6 Hz, 2H), 1.82–1.76 (m, 2H), 1.49–
1.46 (m, 2H), 1.40–1.32 (m, 4H), 0.93–0,90 (m, 3H); 13C NMR
(125 MHz, CDCl3) d 162.7, 156.7, 147.8, 134.2, 121.9, 119.1,
117.2, 115.9, 105.3, 68.6, 31.7, 29.4, 25.9, 22.7, 14.2; MS (FAB) m/
z 296 (M+H)+.
4.1.42. N-(1-(3-(4-Phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethyl)
acetamide (32)
General procedure C: A white solid (87%); 1H NMR (500 MHz,
CDCl3) d 8.02 (d, J = 9.2 Hz, 2H), 7.39 (dd, J = 7.4, 7.4 Hz, 2H), 7.18
(t, J = 7.4 Hz, 1H), 7.08–7.05 (m, 4H), 6.17 (d, J = 7.4 Hz, 1H),
5.50–5.44 (m, 1H), 2.09 (s, 3H), 1.64 (d, J = 6.9 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 179.4, 169.5, 167.8, 160.3, 155.9, 130.0,
129.2, 124.2, 120.9, 119.8, 118.3, 42.8, 23.1, 19.9; ESI-TOF-HRMS
calcd for C18H17N3O3 (m/z) [M+Na]+ 346.1162, found 346.1179.
4.1.36. 4-(4-Phenoxyphenoxy)benzonitrile (34)
General procedure A: A light yellow solid (81%); 1H NMR
(500 MHz, CDCl3) d 7.83 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 7.42–7.38
(m, 2H), 7.17 (dd, J = 6.9, 2.3 Hz, 2H), 7.14 (t, J = 7.4 Hz, 1H),
7.09 (ddd, J = 8.6, 2.3, 2.3 Hz, 4H), 7.04 (dd, J = 8.6, 1.1 Hz, 2H);
13C NMR (125 MHz, CDCl3) d 161.5, 156.8, 153.6, 149.8, 134.6,
130.1, 123.5, 122.1, 120.5, 118.8, 118.5, 117.6, 104.9; ESI-TOF-
4.1.43. N-((3-(4-(4-Benzyloxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)methyl)acetamide (40)
General procedure C: A white solid (76%); 1H NMR (500 MHz,
CDCl3) d 7.99 (ddd, J = 9.2, 2.4, 2.4 Hz, 2H), 7.47–7.43 (m, 2H),
7.42–7.39 (m, 2H), 7.36–7.33 (m, 1H), 7.03–6.98 (m, 6H), 6.20 (br
s, 1H), 5.07 (s, 2H), 4.75 (d, J = 5.7 Hz, 2H), 2.13 (s, 3H); 13C NMR
(125 MHz, CDCl3) d 175.7, 170.2, 167.9, 161.3, 155.7, 149.1,
136.8, 129.2, 128.6, 128.1, 127.5, 121.4, 120.2, 117.3, 116.0, 70.5,
35.9, 23.0; ESI-TOF-HRMS calcd for C24H21N3O4 (m/z) [M+Na]+
438.1424, found 438.1427.
HRMS calcd for
310.0838.
C
19H13NO2 (m/z) [M+Na]+ 310.0838, found
4.1.37. 4-(4-Hexyloxyphenoxy)benzamidoxime (35)
General procedure B: A white solid (73%); 1H NMR (500 MHz,
CDCl3) d 8.31 (br s, 1H), 7.55 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 6.98
(ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 6.93 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H),
6.89 (ddd, J = 9.2, 2.3, 2.3 Hz, 2H), 4.84 (s, 2H), 3.94 (t, J = 6.6 Hz,
2H), 1.81–1.76 (m, 2H), 1.49–1.44 (m, 2H), 1.38–1.31 (m, 4H),
0.93–0.90 (m, 3H); 13C NMR (125 MHz, CDCl3) d 160.4, 156.0,
152.5, 149.3, 127.5, 126.5, 121.3, 117.3, 115.7, 68.6, 31.7, 29.4,
25.9, 22.8, 14.2; MS (FAB) m/z 329 (M+H)+.
4.1.44. N-((3-(4-(4-Hydroxyphenoxy)phenyl)-1,2,4-oxadiazol-5-
yl)methyl)acetamide (38)
General procedure D: A white solid (92%); 1H NMR (500 MHz,
CDCl3) d 7.99 (d, J = 9.0 Hz, 2H), 7.00 (d, J = 8.5 Hz, 2H), 6.97 (d,
J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 2H), 6.17 (br s, 1H), 5.04 (br s,
1H), 4.76–4.74 (m, 2H), 2.13 (s, 3H); 13C NMR (125 MHz, CDCl3) d
175.7, 170.5, 167.9, 161.5, 152.8, 148.7, 129.2, 121.7, 120.0,
117.1, 116.5, 35.9, 23.0; ESI-TOF-HRMS calcd for C17H15N3O4 (m/
z) [M+Na]+ 348.0955, found 348.0947.
4.1.38. 4-(4-Phenoxyphenoxy)benzamidoxime (36)
General procedure B: A white solid (79%); 1H NMR (500 MHz,
CDCl3) d 8.08 (br s, 1H), 7.59 (ddd, J = 9.2, 2.3, 2.3, 2H), 7.36–7.32
(m, 2H), 7.10 (t, J = 7.4 Hz, 1H), 7.02–6.98 (m, 8H), 4.85 (s, 2H);
13C NMR (125 MHz, CDCl3) d 159.5, 157.5, 153.3, 152.2, 151.7,
129.8, 127.4, 127.0, 123.2, 121.0, 120.4, 118.5, 117.8; ESI-TOF-
4.1.45. N-((3-(4-Phenoxyphenyl)-1,2,4-oxadiazol-5-yl)methyl)
acetamide (39)
HRMS calcd for
321.1264.
C
19H16N2O3 (m/z) [M+H]+ 321.1234, found
General procedure C: A white solid (62%); 1H NMR (500 MHz,
CDCl3) d 8.02 (ddd, J = 8.0, 2.5, 2.5 Hz, 2H), 7.39 (dd, J = 8.0,
8.0 Hz, 2H), 7.19 (t, J = 8.0 Hz, 1H), 7.10–7.05 (m, 4H), 6.15 (br s,
1H), 4.75 (d, J = 6.0 Hz, 2H), 2.13 (s, 3H); 13C NMR (125 MHz,
DMSO-d6) d 177.7, 169.9, 167.1, 159.8, 155.3, 130.3, 129.1, 124.6,
120.6, 119.8, 118.2, 35.2, 22.2; ESI-TOF-HRMS calcd for
4.1.39. 4-Phenoxybenzamidoxime (37)
General procedure B: A white solid (96%); 1H NMR (500 MHz,
CDCl3) d 8.32 (br s, 1H), 7.59 (ddd, J = 8.6, 2.9, 1.7, 2H), 7.36 (dd,
J = 9.6, 9.6 Hz, 2H), 7.15 (t, J = 7.4 Hz, 1H), 7.05–6.99 (m, 4H), 4.86
(s, 2H); 13C NMR (125 MHz, CDCl3) d 159.0, 156.4, 152.2, 129.9,
C
17H15N3O3 (m/z) [M+Na]+ 332.1006, found 332.1029.
Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045

S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
11
4.1.46. tert-Butyl (R)-(1-(3-(4-(4-phenoxyphenoxy)phenyl)-
1,2,4-oxadiazol-5-yl)ethyl)carbamate ((R)-41)
7.13–7.09 (m, 1H), 7.06–7.02 (m, 8H), 5.18 (br s, 2H), 1.62 (d,
J = 6.9 Hz, 3H), 1.46 (s, 9H); 13C NMR (125 MHz, CDCl3) d 179.9,
167.8, 160.7, 157.4, 154.8, 153.6, 151.3, 129.8, 129.3, 123.3,
121.3, 121.0, 120.4, 118.6, 117.8, 80.5, 44.3, 28.3, 20.1; ESI-TOF-
General procedure C: A white solid (65%); 1H NMR (500 MHz,
CDCl3) d 8.03 (ddd, J = 9.4, 2.4, 2.4 Hz, 2H), 7.37–7.33 (m, 2H),
7.13–7.10 (m, 1H), 7.06–7.02 (m, 8H), 5.17 (br s, 2H), 1.62 (d,
J = 6.9 Hz, 3H), 1.46 (s, 9H); 13C NMR (125 MHz, CDCl3) d 179.9,
167.8, 160.7, 157.4, 154.8, 153.6, 151.3, 129.8, 129.3, 123.3,
121.3, 121.0, 120.4, 118.6, 117.8, 80.5, 44.3, 28.3, 20.1; ESI-TOF-
HRMS calcd for
472.1892.
C
27H27N3O5 (m/z) [MꢀH]ꢀ 472.1867, found
4.1.53. tert-Butyl (S)-(1-(3-(4-phenoxyphenyl)-1,2,4-oxadiazol-
5-yl)ethyl)carbamate ((S)-42)
HRMS calcd for
472.1871.
C
27H27N3O5 (m/z) [MꢀH]ꢀ 472.1867, found
General procedure C: A white solid (61%); 1H NMR (500 MHz,
CDCl3) d 8.03 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.41–7.37 (m, 2H),
7.20–7.16 (s, 1H), 7.08–7.05 (m, 4H), 5.18 (br s, 2H), 1.63 (d,
J = 6.6 Hz, 3H), 1.46 (s, 9H); 13C NMR (125 MHz, CDCl3) d 179.9,
167.8, 160.2, 156.0, 154.8, 130.0, 129.3, 124.2, 121.2, 119.8,
118.3, 80.5, 44.3, 28.3, 20.1; ESI-TOF-HRMS calcd for C21H23N3O4
(m/z) [M+H]+ 404.1581, found 404.1580.
4.1.47. tert-Butyl (R)-(1-(3-(4-phenoxyphenyl)-1,2,4-oxadiazol-
5-yl)ethyl)carbamate ((R)-42)
General procedure C: A white solid (69%); 1H NMR (500 MHz,
CDCl3) d 8.03 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.41–7.37 (m, 2H),
7.20–7.16 (m, 1H), 7.08–7.05 (m, 4H) 5.18 (br s, 2H), 1.63 (d,
J = 6.6 Hz, 3H), 1.46 (s, 9H); 13C NMR (125 MHz, CDCl3) d 179.9,
167.8, 160.2, 156.0, 154.8, 130.0, 129.3, 124.2, 121.2, 119.8,
118.3, 80.5, 44.3, 28.3, 20.1; ESI-TOF-HRMS calcd for C21H23N3O4
(m/z) [M+H]+ 404.1581, found 404.1578.
4.1.54. (S)-1-(3-(4-(4-Phenoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethan-1-amine ((S)-43)
General procedure F: A white solid (quant.); 1H NMR (500 MHz,
CDCl3) d 7.92 (ddd, J = 9.4, 2.4, 2.4 Hz, 2H), 7.36–7.32 (m, 2H), 7.12–
7.09 (m, 1H), 7.05–6.99 (m, 8H), 4.85 (q, J = 7.1 Hz, 1H), 1.83 (d,
J = 6.9 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 174.9, 168.1, 161.3,
157.4, 153.8, 150.9, 129.8, 129.4, 123.3, 121.5, 120.4, 119.5,
118.6, 117.7, 44.8, 17.3; ESI-TOF-HRMS calcd for C22H19N3O3 (m/
z) [M+H]+ 374.1499, found 374.1470.
4.1.48. (R)-1-(3-(4-(4-Phenoxyphenoxy)phenyl)-1,2,4-oxadiazol-
5-yl)ethan-1-amine ((R)-43)
General procedure F: A white solid (quant.); 1H NMR (500 MHz,
CDCl3) d 7.92 (ddd, J = 9.4, 2.4, 2.4 Hz, 2H), 7.36–7.32 (m, 2H), 7.12–
7.09 (m, 1H), 7.05–6.99 (m, 8H), 4.85 (q, J = 7.1 Hz, 1H), 1.83 (d,
J = 7.4 Hz, 3H); 13C NMR (125 MHz, CDCl3) d 174.9, 168.1, 161.3,
157.4, 153.8, 150.9, 129.8, 129.4, 123.3, 121.5, 120.4, 119.5,
118.6, 117.7, 44.8, 17.3; ESI-TOF-HRMS calcd for C22H19N3O3 (m/
z) [M+H]+ 374.1499, found 374.1485.
4.1.55. (S)-1-(3-(4-Phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethan-
1-amine ((S)-44)
General procedure F: A white solid (98%); 1H NMR (500 MHz,
CD3OD) d 8.07 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.44–7.40 (m, 2H),
7.21 (t, J = 7.4 Hz, 1H), 7.10–7.06 (m, 4H), 4.95 (q, J = 6.9 Hz, 1H),
1.78 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CD3OD) d 177.5,
169.3, 162.3, 157.2, 131.2, 130.4, 125.6, 121.7, 121.0, 119.2, 45.2,
4.1.49. (R)-1-(3-(4-Phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethan-
1-amine ((R)-44)
General procedure F: A white solid (90%); 1H NMR (500 MHz,
CD3OD) d 8.07 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.44–7.40 (m, 2H),
7.21 (t, J = 7.4 Hz, 1H), 7.10–7.06 (m, 4H), 4.95 (q, J = 6.9 Hz, 1H),
1.78 (d, J = 6.9 Hz, 3H); 13C NMR (125 MHz, CD3OD) d 177.5,
169.3, 162.3, 157.2, 131.2, 130.4, 125.6, 121.7, 121.0, 119.2, 45.2,
17.4; ESI-TOF-HRMS calcd for
282.1237, found 282.1207.
C
16H15N3O2 (m/z) [M+H]+
4.1.56. (S)-N-(1-(3-(4-(4-Phenoxyphenoxy)phenyl)-1,2,4-
oxadiazol-5-yl)ethyl)acetamide ((S)-31)
17.4; ESI-TOF-HRMS calcd for
282.1237, found 282.1221.
C
16H15N3O2 (m/z) [M+H]+
General procedure G: A white solid (74%); 1H NMR (500 MHz,
CDCl3) d 8.02 (ddd, J = 8.6, 2.4, 2.4 Hz, 2H), 7.38–7.33 (m, 2H),
7.11 (t, J = 7.4 Hz, 1H), 7.07–7.02 (m, 8H), 6.15 (d, J = 7.4 Hz, 1H),
5.50–5.44 (m, 1H), 2.09 (s, 3H), 1.64 (d, J = 6.9 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 179.4, 169.4, 167.8, 160.7, 157.4, 153.6,
151.2, 129.8, 129.3, 123.3, 121.3, 120.8, 120.4, 118.6, 117.8, 42.8,
23.2, 20.0; ESI-TOF-HRMS calcd for C24H21N3O4 (m/z) [MꢀH]ꢀ
414.1448, found 414.1442.
4.1.50. (R)-N-(1-(3-(4-(4-Phenoxyphenoxy)phenyl)-1,2,4-oxadi-
azol-5-yl)ethyl)acetamide ((R)-31)
General procedure G: A white solid (73%); 1H NMR (500 MHz,
CDCl3) d 8.02 (ddd, J = 8.6, 2.4, 2.4 Hz, 2H), 7.38–7.33 (m, 2H),
7.11 (t, J = 7.4 Hz, 1H), 7.07–7.02 (m, 8H), 6.15 (d, J = 8.0 Hz, 1H),
5.50–5.44 (m, 1H), 2.09 (s, 3H), 1.64 (d, J = 6.9 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 179.4, 169.4, 167.8, 160.7, 157.4, 153.6,
151.2, 129.8, 129.3, 123.3, 121.3, 120.8, 120.4, 118.6, 117.8, 42.8,
23.2, 20.0; ESI-TOF-HRMS calcd for C24H21N3O4 (m/z) [MꢀH]ꢀ
414.1448, found 414.1443.
4.1.57. (S)-N-(1-(3-(4-Phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethyl)
acetamide ((S)-32)
General procedure G: A white solid (79%); 1H NMR (500 MHz,
CDCl3) d 8.02 (ddd, J = 9.4, 2.3, 2.3 Hz, 2H), 7.41–7.37 (m, 2H),
7.20–7.16 (m, 1H), 7.08–7.05 (m, 4H), 6.19 (d, J = 7.4 Hz, 1H),
5.50–5.44 (m, 1H), 2.09 (s, 3H), 1.64 (d, J = 6.9 Hz, 3H); 13C
4.1.51. (R)-N-(1-(3-(4-Phenoxyphenyl)-1,2,4-oxadiazol-5-yl)ethyl)
acetamide ((R)-32)
General procedure G: A white solid (90%); 1H NMR (500 MHz,
CDCl3) d 8.02 (ddd, J = 9.4, 2.4, 2.4 Hz, 2H), 7.41–7.37 (m, 2H),
7.20–7.16 (m, 1H), 7.08–7.05 (m, 4H), 6.20 (d, J = 7.4 Hz, 1H),
5.50–5.44 (m, 1H), 2.09 (s, 3H), 1.64 (d, J = 7.4 Hz, 3H); 13C NMR
(125 MHz, CDCl3) d 179.4, 169.5, 167.8, 160.3, 155.9, 130.0,
129.2, 124.2, 120.9, 119.8, 118.3, 42.8, 23.1, 19.9; ESI-TOF-HRMS
calcd for C18H17N3O3 (m/z) [M+H]+ 346.1162, found 346.1152.
NMR (125 MHz, CDCl3)
d 179.4, 169.5, 167.8, 160.3, 160.0,
130.0, 129.2, 124.2, 120.9, 119.8, 118.3, 42.8, 23.1, 19.9; ESI-
TOF-HRMS calcd for C18H17N3O3 (m/z) [M+H]+ 346.1162, found
346.1154.
4.2. Biology
4.2.1. Cell culture
4.1.52. tert-Butyl (S)-(1-(3-(4-(4-phenoxyphenoxy)phenyl)-1,2,
4-oxadiazol-5-yl)ethyl)carbamate ((S)-41)
Human embryonic kidney 293 (HEK293) cells were cultured in
DMEM supplemented with 10% FBS and penicillin/streptomycin
mixture at 37 °C in a humidified incubator (5% CO2).
General procedure C: A white solid (69%); 1H NMR (500 MHz,
CDCl3) d 8.03 (ddd, J = 9.4, 2.4, 2.4 Hz, 2H), 7.37–7.33 (m, 2H),
Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045

12
S. Okazaki et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
4.2.2. ACC2 inhibitory activity assay
References and notes
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The work described in this paper was partially supported by
Grants-in-Aid for Scientific Research (KAKENHI, Grant-in-Aid for
Young Scientists (B), No. 26810091 and 16K17930 to T.Y.) from
The Ministry of Education, Culture, Sports, Science and Technology
in Japan (MEXT), and the Japan Society for the Promotion of Science
(JSPS), and Platform for Drug Discovery, Informatics, and Structural
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A. Supplementary data
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Supplementary data associated with this article can be found, in
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Please cite this article in press as: Okazaki, S.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.08.045