Synthesis and biological evaluation of GPR40/FFAR1 agonists containing 3,5-dimethylisoxazole

Article abstract of DOI:10.1016/j.ejmech.2016.03.054

GPR40 is an attractive target due to its glucose-stimulated insulin secretion effect with low risk of causing hypoglycemia, which also can be seen from the clinical studies using TAK-875 (fasiglifam). In the present studies, we discovered a series of anal

Full text of DOI:10.1016/j.ejmech.2016.03.054

Accepted Manuscript
Synthesis and biological evaluation of GPR40/FFAR1 agonists containing 3,5-
dimethylisoxazole
Lingyun Yang, Jian Zhang, Lianghui Si, Li Han, Bo Zhang, Hui Ma, Junhao Xing,
Leilei Zhao, Jinpei Zhou, Huibin Zhang
PII:
S0223-5234(16)30236-7
10.1016/j.ejmech.2016.03.054
EJMECH 8479
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 21 October 2015
Revised Date: 17 March 2016
Accepted Date: 18 March 2016
Please cite this article as: L. Yang, J. Zhang, L. Si, L. Han, B. Zhang, H. Ma, J. Xing, L. Zhao, J.
Zhou, H. Zhang, Synthesis and biological evaluation of GPR40/FFAR1 agonists containing 3,5-
dimethylisoxazole, European Journal of Medicinal Chemistry (2016), doi: 10.1016/j.ejmech.2016.03.054.
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ACCEPTED MANUSCRIPT
Graphical abstract

ACCEPTED MANUSCRIPT
Synthesis and biological evaluation of GPR40/FFAR1 agonists containing
3,5-dimethylisoxazole
Lingyun Yang a, Jian Zhang a, Lianghui Si a, Li Han a, Bo Zhang a, Hui Ma a, Junhao Xing a, Leilei
Zhao a, Jinpei Zhou b, Huibin Zhang a, c, *
a Center of Drug Discovery, State Key Laboratory of Natural Medicines, China Pharmaceutical University, 24
Tongjiaxiang, Nanjing 210009, PR China
b Department of Medicinal Chemistry, China Pharmaceutical University, TongjiaXiang 24, 210009 Nanjing, PR
China
c
Jiangsu Key Laboratory of Drug Discovery for Metabolic Disease, China Pharmaceutical University, Nanjing
210009, PR China
Abstract
GPR40 is an attractive target due to its glucose-stimulated insulin secretion effect with low risk of
causing hypoglycemia, which also can be seen from the clinical studies using TAK-875
(fasiglifam). In the present studies, we discovered a series of analogues containing
3,5-dimethylisoxazole as potent GPR40 agonists, especially compound 11k with an EC50 value of
15.9 nM. Moreover, compound 11k reduced glucose excursion to 23.1% in ICR mice and 29.5%
in type 2 diabetic C57BL/6 mice at 30 mg/kg. It also exhibited satisfactory PK profile. Docking
studies were conducted to explain the interaction mode of this series. In summary, compound 11k
with robust efficacy in vitro and in vivo is a promising drug candidate for further investigation.
Keywords
Diabetes; GPR40 agonist; 3,5-dimethylisoxazole; oral glucose tolerance test
1. Introduction
Type 2 diabetes mellitus (T2DM) is mainly characterized by insulin deficiency and insulin
resistance, often accompanied by multiple complications such as cardiovascular disease, renal
failure and retinopathy. Currently, the most common oral antidiabetic agents include sulfonylureas,
metformin, glitazones, all of which are associated with limited efficacy and adverse side effects [1,
2]. There are only a few insulin secretagogues with low risk of hypoglycemia and weight gain,
such as DPP-IV inhibitors, SGLT2 inhibitors and GLP-1 agonists [3, 4]. The urgent need for novel
oral antidiabetic agents with sustained safety and efficacy stimulates efforts to develop new
mechanisms to improve glycemic control.
GPR40 (also known as FFAR1) has, since its deorphanization as a cell surface free fatty acids
receptor in 2003, become an attractive and potential target for T2DM therapy [5-7]. GPR40 is a
G-protein-coupled receptor, highly expressed in pancreatic β-cells and intestinal enteroendocrine
cells [8]. Activation of GPR40 by medium (C6–C12) to long (C14–C24) chain saturated and
unsaturated fatty acids enhances glucose-stimulated insulin secretion (GSIS). However,
mechanisms of the activation of GPR40 to GSIS are only partially understood and remain to be
fully explained. Several studies have pointed that GPR40 is coupled with heterotrimeric G protein
Gαq/11, which activates phospholipase C, leading to the formation of diacylglycerol (DAG) and
inositol-triphosphate (IP3). IP3 binding to its receptor on endoplasmatic reticulum activates an
intracellular Ca2+ mobilization. The generation of DAG stimulates protein kinase D1 and promotes
F-Actin polymerization. Subsequently, both Ca2+ mobilization and F-Actin polymerization
enhance insulin secretion [9, 10]. The modulation of insulin secretion via GPR40 is dependent on
* Corresponding author. Center of Drug Discovery, State Key Laboratory of Natural Medicines, China
Pharmaceutical University, 24 Tongjiaxiang, Nanjing 210009, China.
E-mail addresses: yanglingyun130@163.com (L. Yang), zhanghb80@163.com (H. Zhang).

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glucose concentration, indicating GPR40 agonists could have a low risk of hypoglycemia.
A variety of synthetic GPR40 agonists containing 3-phenylpropanoic acid and its
bioisostere have been reported (Fig. 1) [11-14]. Among these potent molecules, TAK-875,
developed from a modification of compound 1, was the most advanced in clinical phase III.
TAK-875 showed significant glycemic control and ideal pharmacokinetics profiles. The
dihydrobenzofuran ring of TAK-875 is highly resistant to β-oxidation, thus slowing down its rate
of metabolism [15-17]. Unfortunately, the development of TAK-875 was terminated by Takeda
due to liver toxicity in December 2013 [18]. The modification of TAK-875 from compound 1
inspired us to choose compound 1 as a starting structure to explore novel GPR40 agonists in a
different way.
It has been recognized that GPR40 is expressed in the brain. While the consequences of
activation of GPR40 in the brain have not been identified [19, 20]. A recent study found a GPR40
agonist (AM-3189) with high topological polar surface area (tPSA) showed minimal CNS
penetration. tPSA is defined as the surface area occupied by nitrogen and oxygen atoms, and polar
hydrogens bonded to these heteroatoms [21]. It is a novel parameter associated with incidence of
toxicity for highly lipophilic and low polarity compounds. To avoid potential effects in central
nervous system, we decided to improve tPSA by introducing functional groups with greater tPSA
to compound 1, which has a high lipophilicity by itself [22, 23]. Compared with compound 1, the
replacement of the 2,6-dimethylphenyl group with 3,5-dimethylisoxazole in compound 11a led to
a noticeable increase in the tPSA [24]. The well-known rule of five describes clog P should less
than 5. For GPR40 agonists, strict range of cLog P has not been proposed as far. However, a high
cLog P value is related with undesirable ADMET properties [25]. Most of GPR40 agonists present
cLog P values between 4 and 5. We considered this range was probably ideal and the value of
compound 1 (cLog P = 6.16) was too high. The introduction of 3,5-dimethylisoxazole also
decreased cLog P. Based on compound 11a, we designed and synthesized a series of compounds
containing 3,5-dimethylisoxazole. On the one hand, we introduced halogen, alkyl or alkoxyl on
the phenyl to obtain compound 11b–l and 22a–h. On the other hand, we replaced phenyl with
pyridyl to obtain compound 28a–d [26-28]. TUG-770 is a highly potent GPR40 agonist with
favorable physicochemical and pharmacokinetic properties [29]. The introduction of the ethynyl
connecting the phenyl rings is very successful. Comparing the size of compound 11a and
TAK-875, we thought 11a was shorter. So we decided to introduce the ethynyl into compound 36
to make the 3,5-dimethylisoxazole closer to the solvent region. Most of these compounds were
proved to be novel GPR40 agonist by biological evaluation in vitro and in vivo, along with
optimized tPSA and cLog P. Among them, compound 11k was found to be the most efficacious in
vitro and in vivo as a promising lead compound.
Figure 1.
2. Results and discussion
2.1. Chemistry
3,5-dimethylisoxazole derivatives 11a–l, 22a–h, 28a–d and 36 were synthesized as outlined
in Schemes 1–4. All compounds were purified by flash chromatography and checked for purity
using HPLC before being tested in biological evaluation (purity was > 97%). The structures of
1
3,5-dimethylisoxazole derivatives were confirmed by H NMR and 13C NMR spectrum, mass
spectrometry, IR spectrum.

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As shown in Scheme 1, the synthesis of (E)-3-(2-fluoro-4-hydroxyphenyl)acrylic acid (3)
was carried out by Knoevenagel condensation of 2-fluoro-4-hydroxybenzaldehyde (2) with
propanedioic
acid
in
pyridine.
The
key
intermediate
methyl
3-(2-fluoro-4-hydroxyphenyl)propanoate (5) was prepared by reduction of intermediate 3 in the
presence of Pd/C, followed by methyl esterification. (3,5-dimethylisoxazol-4-yl)benzaldehyde
(7a–l) were synthesized by Suzuki coupling reaction. The chloride intermediates (9a–l) were
prepared by the substitution of corresponding benzyl alcohols (8a–l) with phosphorus oxychloride,
which were derived from reduction of benzaldehyde (7a–l) with NaBH4. The target compounds
11a–l were obtained by condensation of intermediates 9a–l with intermediate 5, followed by
hydrolysis of the methyl esters 10a–l. Derivatives 28a–d were synthesized as outlined in Scheme
3. The synthetic route was similar to that in Scheme 1.
Scheme 1.
As shown in Scheme 2, 3-bromo-5-nitrobenzaldehyde (13) was prepared by bromination with
3-nitrobenzaldehyde (12). The key intermediate 17 was synthesized by Suzuki coupling reaction
with borate ester and 3-bromo-5-hydroxybenzaldehyde (16), which was obtained in several steps
by the introduction of 1,3-dioxolane as protecting group in intermediate 13, reduction of
intermediate 14 employing iron powder, subsequent Sandmeyer reaction and removal of the
protecting group. Intermediate 18a-h were prepared by condensation of intermediate 17 with the
corresponding bromoalkanes. The following procedures were depicted in Scheme 1 to give
compounds 22a–h.
Scheme 2.
Scheme 3.
As shown in Scheme 4, 3-((trimethylsilyl)ethynyl)benzaldehyde (30) was synthesized by
Sonogashira coupling of 3-iodobenzaldehyde with trimethylsilylacetylene (29). Removal of
trimethylsilyl by TBAF and second Sonogashira reaction with 4-iodo-3,5-dimethylisoxazole were
conducted to give intermediate 32. The desired compound 36 was prepared as the synthetic route
described previously.
Scheme 4.
2.2. GPR40 agonist activity and SAR study
The GPR40 agonist activities of synthesized compounds were evaluated by FLIPR calcium
assay in Chinese hamster ovary (CHO) cells stably expressing human GPR40. TAK-875 was used
as positive control. The agonist activities were presented in Table 1–4. We initially modified the
substituents on the phenyl connecting with 3,5-dimethylisoxazole. As shown in Table 1,
compounds 11a-l containing halogen, alkyl or alkoxyl maintained strong GPR40 agonist activities
with improved tPSA and lowered cLog P values compared with compound 1. It demonstrated
3,5-dimethylisoxazole was highly tolerable as a terminal group in GPR40 agonists. The
introduction of nitrogen and oxygen to the heterocycle was beneficial to tPSA and cLog P. It was
expected to reduce the risk of the potential effects in central nervous system. In this series,
compounds 11b, 11f, 11g and 11k showed good potency and were selected for in vivo profiling.
To understand the binding mode of this series, we performed docking studies with Glide docking
in Schrodinger 9.4 (Fig. 2). The first crystal structure of human GPR40 receptor (PDB ID: 4PHU)
binding to TAK-875 was reported by Takeda in 2014 [30]. A comparison of TAK-875 to energy
minimized conformation of compound 11a indicated a high percentage of overlap. Both
carboxylate moiety in two molecules interacted with arginine residues Arg 183 and Arg 258

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(displayed as Arg 2258 in Schrodinger 9.4). The fluorine atom overlapped with the oxygen atom
on dihydrobenzofuran of TAK-875. The 3,5-dimethylisoxazole in compound 11a and the left
2,6-dimethylphenyl in TAK-875 was orthogonal to the middle phenyl ring. The similarities
between 3,5-dimethylisoxazole analogues and TAK-875 allowed explanation of the GPR40
agonist activities we evaluated.
Table 1.
Figure 2.
Next, we explored alkoxyl into the meta-position of the middle phenyl ring to obtain
compounds 18a-h (Table. 2). The EC50 values ranged from 53.1 nM to 143 nM, indicating that
meta-position of the phenyl could tolerate substituents with differing length with minimal loss of
potency. However, the replacement of the middle phenyl ring with a variety of the pyridine rings
reduced agonist activity significantly (Table. 3). Although the tPSA value of compound 22a-d
were improved, the pyridine ring was not suitable for the middle ring of GPR40 agonists. The
introduction of the ethynyl connecting the 3,5-dimethylisoxazole and the phenyl gave compound
36 an EC50 value of 214 nM (Table. 4).
Table 2.
Table 3.
Table 4.
2.3. Antihyperglycemic effect of 11b, 11f, 11g and 11k in ICR mice
According to the results of GPR40 agonist activity in vitro, compound 11b, 11f, 11g and 11k
were selected for in vivo profiling. Acute oral glucose tolerance test (OGTT) was conducted in
male ICR mice initially. Mice were dosed orally with single doses of vehicle, TAK-875 (30 mg/kg)
and test compounds (30 mg/kg), 30 min prior to oral glucose load (2 g/kg). Among these
compounds, compound 11f and 11k exhibited robust potency of glycemic control. Compound 11f,
11k and TAK-875 reduced area under the curve from 0 to 120 min (AUC0–120 min) by 19.1%, 23.1%
and 23.3% (18f, 785.9 ± 31.0; 18k, 746.4 ± 52.3; TAK-875, 744.9 ± 24.9), respectively (Fig. 3).
Figure 3.
2.4. Antihyperglycemic effect of 11f and 11k in type 2 diabetic C57BL/6 mice
Based on the data in ICR mice OGTT, compound 11f and 11k were tested in type 2 diabetic
C57BL/6 mice. Mice were dosed orally with single doses of vehicle, TAK-875 (30 mg/kg),
compound 11f and 11k (30 mg/kg), 30 min prior to oral glucose load (1 g/kg). Fig. 4 depicted that
compound 11k reduced notably AUC0–120 min by 29.5% and its antihyperglycemic effect was
similar to that of TAK-875 (11k, 1298.5 ± 47.5; TAK-875, 1216.6 ± 75.0).
Figure 4.
2.5 Pharmacokinetic profiles of 11k in SD rats
Compound 11k was selected to the further evaluation for oral pharmacokinetic profiles in SD
rats (Table 5.). It exhibited favorable pharmacokinetic profiles, with lasting half-life (2.87 ± 0.33
h), fast oral absorption (Tmax = 1.0 h), high maximum plasma concentration (Cmax = 0.84 ± 0.15
ꢀg/mL) and high plasma drug exposure (AUC0-8 h = 4.18 ± 0.71 ꢀg·h/mL).
Table 5.
3. Conclusion
In summary, we designed and discovered a series of novel GPR40 agonists containing
3,5-dimethylisoxazole with improved tPSA and ideal cLog P. Most of these compounds were
confirmed as excellent GPR40 agonists in vitro. Especially, compound 11k exhibited robust

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potency of glycemic control in both ICR mice and type 2 diabetic C57BL/6 mice compared with
TAK-875. The pharmacokinetic profiles was evaluated and presented desirable results. In addition,
docking studies were conducted to explain the potency of 3,5-dimethylisoxazole analogues. Those
qualities made compound 11k a promising lead compound for the treatment of T2DM and worth
for further investigation.
4. Experimental section
4.1. Chemistry
All chemical reagents were commercially available and treated with standard methods before
use. Solvents were dried and redistilled before use. Column chromatography (CC): silica gel 60
(200–300 mesh). Thin-layer chromatography (TLC): silica gel 60 F254 plates (250 nm; Qingdao
Ocean Chemical Company, Qingdao, China). M.p.: capillary tube, RY-1 capillary apparatus
(Tianjin Optical Instrument Company, Tianjin, China), uncorrected. %Purity of the target
compounds (> 97%) were determined by HPLC analysis (UV detector, wavelength: 272 nm). IR
spectra: Nicolet Impact 410 spectrophotometer with KBr disks; in cm-1. 1H and 13C NMR spectra:
Bruker ACF-300Q apparatus (300 MHz for 1H NMR and 75 MHz for 13C NMR), in DMSO-d6 or
CDCl3 unless otherwise indicated, δ in ppm rel. to Me4Si, J in Hz. Mass spectrometry (MS):
Hewlett-Packard 1100 LC/MSD spectrometer; elemental analyses: CHN-O-Rapid instrument.
4.1.1. Preparation of (E)-3-(2-fluoro-4-hydroxyphenyl)acrylic acid (3)
To a solution of 2 (1.0 g, 7.1 mmol) in pyridine (10 mL) was added propanedioic acid (1.1 g,
10 mmol) at room temperature. The reaction mixture was stirred at 80°C for 24 h. After cooling to
the room temperature, the mixture was acidified with HCl (1 N) and filtered. The filter cake was
washed with water and dried to give 3 (1.1 g, 84%) as white solid. 1H NMR (300 MHz, DMSO-d6)
δ: 12.29 (s, 1H), 10.47 (s, 1H), 7.64 (t, J = 8.8 Hz, 1H), 7.56 (d, J = 16.1 Hz, 1H), 6.72–6.56 (m,
2H), 6.36 (d, J = 16.1 Hz, 1H).
4.1.2. Preparation of 3-(2-fluoro-4-hydroxyphenyl)propanoic acid (4)
To a solution of 3 (0.80 g, 4.4 mmol) in THF (25 mL) was hydrogenated on 10% Pd/C under
hydrogen atmosphere (balloon pressure) at room temperature for 24 h. The mixture was filtered
through a celite pad, and the filtrate was concentrated to afford 4 (0.79 g, 99%) as a colorless solid
1
without further purification. H NMR (300 MHz, DMSO-d6) δ: 11.80 (br s, 1H), 9.93 (br s, 1H),
7.06 (t, J = 8.8 Hz, 1H), 6.60–6.38 (m, 2H), 2.72 (t, J = 7.4 Hz, 2H), 2.44 (t, J = 7.5 Hz, 2H).
4.1.3. Preparation of methyl 3-(2-fluoro-4-hydroxyphenyl)propanoate (5)
To a solution of 4 (0.70 g, 3.8 mmol) in methanol (20 mL) was added concentrated sulfuric
acid (0.50 mL) and then refluxed for 12 h. After cooling to the room temperature, the solvent was
concentrated in vacuo. The residue was quenched with saturated NaHCO3 solution and extracted
with EtOAc (3 × 20 mL). The combined organic layers were washed with water (30 mL) and brine
(30 mL), dried over Na2SO4, and evaporated in vacuo to afford 5 (0.69 g, 92%) as a brown solid.
1H NMR (300 MHz, DMSO-d6) δ: 9.65 (s, 1H), 7.04 (t, J = 8.9 Hz, 1H), 6.57–6.42 (m, 2H), 3.56
(s, 3H), 2.74 (t, J = 7.5 Hz, 2H), 2.53 (t, J = 7.5 Hz, 2H).
4.1.3. The following procedures described the synthesis of compound (11a)
These procedures can also be applied to the synthesis of compounds 11b–l and 28a–d.
4.1.3.1 Preparation of 3-(3,5-dimethylisoxazol-4-yl)benzaldehyde (7a)
To a mixture of 3,5-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)isoxazole (0.50
g, 2.2 mmol), 3-bromobenzaldehyde (0.50 g, 2.7 mmol), Pd(PPh3)4 (0.13 g, 0.12 mmol) and

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sodium carbonate (0.71 g, 6.7 mmol) in toluene/ethanol/H2O (10 mL, 3/1/3) was refluxed under
nitrogen atmosphere for 24 h. Then the mixture was diluted with saturated ammonium chloride
solution and EtOAc, and the insoluble material was filtered through Celite. The organic layer of
the filtrate was washed with water (30 mL) and brine (30 mL), dried over Na2SO4, and evaporated
in vacuo. The residue was purified by column chromatography (EtOAc/petroleum ether, 1/15 to
1
1/5) to give 7a (0.37 g, 77%) as a white solid. H NMR (300 MHz, DMSO-d6) δ: 10.08 (s, 1H),
7.99–7.85 (m, 2H), 7.81–7.63 (m, 2H), 2.43 (s, 3H), 2.25 (s, 3H).
4.1.3.2 Preparation of (3-(3,5-dimethylisoxazol-4-yl)phenyl)methanol (8a)
To a solution of 7a (0.37 g, 1.8 mmol) in THF/methanol (20 mL, 5/1) was added NaBH4
(0.14 g, 3.7 mmol) in small portions at 0°C. The reaction mixture was stirred at room temperature
for 15 min, and then quenched with HCl (1 N). The mixture was extracted with EtOAc (3 × 20 mL)
and the combined organic layers were washed with water (30 mL) and brine (30 mL), dried over
Na2SO4, and concentrated in vacuo to afford 8a (0.35 g, 94%) as a colorless solid without further
purification. 1H NMR (300 MHz, DMSO-d6) δ: 7.42 (t, J = 7.5 Hz, 1H), 7.37–7.28 (m, 2H), 7.28–
7.15 (m, 1H), 5.24 (t, J = 5.8 Hz, 1H), 4.54 (d, J = 5.8 Hz, 2H), 2.39 (s, 3H), 2.22 (s, 3H).
4.1.3.3 Preparation of 4-(3-(chloromethyl)phenyl)-3,5-dimethylisoxazole (9a)
To a solution of 8a (0.35 g, 1.7 mmol) in N-methyl-2-pyrrolidone (3.0 mL) was added
phosphorus oxychloride (0.43 mL, 5.2 mmol) dropwise at 0°C. The reaction mixture was stirred at
room temperature for 2 h, then quenched with saturated sodium bicarbonate solution. The product
1
was prepiticated, filtered and washed with water to afford 9a (0.34 g, 89%) as a white solid. H
NMR (300 MHz, DMSO-d6) δ: 7.57–7.43 (m, 3H), 7.36 (dt, J = 6.9, 1.8 Hz, 1H), 4.82 (s, 2H),
2.41 (s, 3H), 2.23 (s, 3H).
4.1.3.4 Preparation of methyl 3-(4-((3-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-fluorophenyl)
propanoate (10a)
To a solution of 9a (0.20 g, 0.89 mmol), 5 (0.16 g, 0.81 mmol) and potassium carbonate
(0.22 g, 1.6 mmol) in DMF (3.0 mL) was stirred at 60°C for 5 h. Then the mixture was quenched
with saturated ammonium chloride solution and extracted with EtOAc (3 × 20 mL). The combined
organic layers were washed with water (30 mL) and brine (30 mL), dried over Na2SO4, and
concentrated in vacuo. The residue was purified by column chromatography (EtOAc/petroleum
1
ether, 1/9 to 1/3) to give 10a (0.23 g, 71%) as a white solid. H NMR (300 MHz, DMSO-d6) δ:
7.53–7.40 (m, 3H), 7.35 (d, J = 7.2 Hz, 1H), 7.20 (t, J = 8.8 Hz, 1H), 6.89 (dd, J = 12.2, 2.4 Hz,
1H), 6.80 (dd, J = 8.5, 2.5 Hz, 1H), 5.15 (s, 2H), 3.57 (s, 3H), 2.79 (t, J = 7.5 Hz, 2H), 2.56 (t, J =
7.5 Hz, 2H), 2.38 (s, 3H), 2.21 (s, 3H).
4.1.3.5 Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-fluorophenyl)pro-
panoic acid (11a)
To a solution of 10a (0.23 g, 0.60 mmol) and LiOH (60 mg, 2.4 mmol) in methanol/H2O (10
mL, 5/1) was stirred at 40°C for 6 h. The solvent was evaporated in vacuo and the residue was
acidified with HCl (1 N). The mixture was extracted with EtOAc (3 × 10 mL). The combined
organic layers were washed with water (20 mL) and brine (20 mL), dried over Na2SO4, and
concentrated in vacuo. The residue was purified by column chromatography (EtOAc/petroleum
ether, 1/5 to 1/2) to give compound 11a (0.14 g, 63%) as a white solid. m.p.: 106–108°C; 1H NMR
(300 MHz, DMSO-d6) δ: 11.71 (br s, 1H), 7.63–7.27 (m, 4H), 7.27–7.07 (m, 1H), 6.98–6.69 (m,
2H), 5.16 (s, 2H), 2.75 (t, J = 7.1 Hz, 2H), 2.47 (t, J = 7.1 Hz, 2H), 2.39 (s, 3H), 2.21 (s, 3H); 13
C
NMR (75 MHz, DMSO-d6) δ: 173.63 (s), 165.22 (s), 160.96 (d, J = 224.2 Hz), 158.07 (d, J = 5.8

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Hz), 157.88 (s), 137.48 (s), 130.96 (d, J = 6.8 Hz), 130.02 (s), 129.06 (s), 128.38 (s), 128.19 (s),
126.88 (s), 119.36 (d, J = 16.3 Hz), 115.72 (s), 110.96 (d, J = 2.8 Hz), 102.34 (d, J = 26.0 Hz),
69.23 (s), 34.16 (s), 23.18 (s), 11.31 (s), 10.45 (s); IR (cm-1): 3427.73, 2926.25, 1731.75, 1628.47,
1585.31, 1509.12, 1456.27, 1433.66, 1412.74, 1364.38, 1286.93, 1189.05, 1143.95, 1107.57,
1036.02, 844.93, 805.09, 757.31, 710.14; MS (ESI), m/z: 368.2 (M–H)–. Anal. calcd. for
C21H20FNO4: C, 68.28; H, 5.46; N, 3.79. Found: C, 68.31; H, 5.47; N, 3.74.
4.1.4. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl) -5-methylbenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11b)
1
Compound 11b (0.12 g, 54%) as a white solid. m.p.: 120–122°C; H NMR (300 MHz,
DMSO-d6) δ: 12.15 (br s, 1H), 7.34–7.11 (m, 4H), 6.95–6.74 (m, 2H), 5.10 (s, 2H), 2.75 (t, J = 7.1
Hz, 2H), 2.46 (t, J = 7.1 Hz, 2H), 2.37 (s, 6H), 2.19 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ:
173.55 (s), 165.11 (s), 162.44 (s), 159.21 (s), 157.99 (d, J = 11.8 Hz), 138.44 (s), 137.35 (s),
130.94 (d, J = 6.7 Hz), 129.94 (s), 128.97 (s), 127.50 (s), 125.39 (s), 119.27 (d, J = 16.1 Hz),
115.74 (s), 110.94 (d, J = 2.8 Hz), 102.31 (d, J = 26.1 Hz), 69.26 (s), 34.08 (s), 23.13 (s), 20.94 (s),
11.29 (s), 10.44 (s); IR (cm-1): 3440.82, 2930.61, 1705.68, 1627.51, 1587.51, 1511.23, 1427.25,
1319.48, 1287.01, 1251.36, 1164.35, 1105.05, 859.94, 846.44; MS (ESI), m/z: 382.2 (M–H)–.
Anal. calcd. for C22H22FNO4: C, 68.92; H, 5.78; N, 3.65. Found: C, 68.88; H, 5.73; N, 3.64.
4.1.5. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-fluorobenzyl)oxy)-2-fluorophenyl)
propanoic acid (11c)
1
Compound 11c (0.15 g, 68%) as a white solid. m.p.: 160–161°C; H NMR (300 MHz,
DMSO-d6) δ: 12.20 (br s, 1H), 7.49 (d, J = 10.2 Hz, 1H), 7.32 (d, J = 7.4 Hz, 2H), 7.17 (t, J = 8.8
Hz, 1H), 6.74 (d, J = 12.1 Hz, 1H), 6.65 (d, J = 8.4 Hz, 1H), 4.79 (s, 2H), 2.74 (t, J = 7.5 Hz, 2H),
2.45 (t, J = 7.5 Hz, 2H), 2.19 (s, 3H), 2.01 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.54 (s),
165.80 (s), 162.37 (s), 162.17 (d, J = 243.0 Hz), 159.00 (d, J = 22.5 Hz), 157.63 (d, J = 11.1 Hz),
138.80 (d, J = 7.7 Hz), 133.44 (d, J = 8.5 Hz), 130.97 (d, J = 6.8 Hz), 125.44 (d, J = 3.1 Hz),
119.64 (d, J = 2.4 Hz), 119.44 (s), 116.26 (d, J = 22.2 Hz), 115.55 (d, J = 21.2 Hz), 110.62 (d, J =
3.0 Hz), 102.06 (d, J = 26.0 Hz), 67.32 (s), 34.02 (s), 23.11 (s), 11.01 (s), 9.99 (s); IR (cm-1):
3351.03, 2938.05, 1746.94, 1714.40, 1625.61, 1586.89, 1507.90, 1409.29, 1278.97, 1264.92,
1243.20, 1179.46, 1162.99, 1148.78, 1096.45, 1043.51, 871.73, 826.81; MS (ESI), m/z: 386.1 (M–
H)–. Anal. calcd. for C21H19F2NO4: C, 65.11; H, 4.94; N, 3.62. Found: C, 65.06; H, 4.94; N, 3.60.
4.1.6. Preparation of 3-(4-((3-chloro-5-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-fluorophen-
yl)propanoic acid (11d)
1
Compound 11d (0.16 g, 64%) as a white solid. m.p.: 116–118°C; H NMR (300 MHz,
DMSO-d6) δ: 12.14 (br s, 1H), 7.53 (s, 1H), 7.45 (s, 1H), 7.41 (s, 1H), 7.23 (t, J = 8.8 Hz, 1H),
6.90 (dd, J = 12.1, 2.2 Hz, 1H), 6.82 (dd, J = 8.5, 2.0 Hz, 1H), 5.18 (s, 2H), 2.78 (t, J = 7.5 Hz,
2H), 2.47 (t, J = 7.5 Hz, 2H), 2.40 (s, 3H), 2.22 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.54
(s), 165.85 (s), 160.82 (d, J = 243.2 Hz), 158.02 (s), 157.69 (d, J = 11.2 Hz), 139.86 (s), 133.64 (s),
132.17 (s), 131.00 (d, J = 6.8 Hz), 127.87 (s), 126.76 (s), 126.45 (s), 119.56 (d, J = 15.8 Hz),
114.68 (s), 110.92 (d, J = 2.6 Hz), 102.41 (d, J = 25.9 Hz), 68.36 (s), 34.07 (s), 23.15 (s), 11.31 (s),
10.33 (s); IR (cm-1): 3439.53, 2926.25, 1704.01, 1625.66, 1588.05, 1576.71, 1510.35, 1428.02,
1408.71, 1315.85, 1293.73, 1278.87, 1248.75, 1162.90, 1148.62, 1104.29, 858.36, 782.15; MS
(ESI), m/z: 402.1 (M–H)–. Anal. calcd. for C21H19ClFNO4: C, 62.46; H, 4.74; N, 3.47. Found: C,
62.44; H, 4.71; N, 3.53.
4.1.7. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-(trifluoromethyl)benzyl)oxy)-2-

ACCEPTED MANUSCRIPT
fluorophenyl)propanoic acid (11e)
1
Compound 11e (0.12 g, 48%) as a white solid. m.p.: 153–156°C; H NMR (300 MHz,
DMSO-d6) δ: 12.15 (br s, 1H), 7.82 (s, 1H), 7.75 (s, 1H), 7.71 (s, 1H), 7.22 (t, J = 8.8 Hz, 1H),
6.96–6.87 (m, 1H), 6.87–6.79 (m, 1H), 5.26 (s, 2H), 2.77 (t, J = 7.5 Hz, 2H), 2.46 (t, J = 7.5 Hz,
2H), 2.40 (s, 3H), 2.22 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.53 (s), 166.07 (s), 160.83 (d,
J = 243.2 Hz), 158.04 (s), 157.69 (d, J = 11.9 Hz), 139.14 (s), 132.03 (s), 131.42 (s), 131.02 (d, J
= 6.7 Hz), 129.76 (s), 125.14–124.35 (m), 123.72–122.67 (m), 122.15 (s), 119.63 (d, J = 16.4 Hz),
114.68 (s), 110.91 (d, J = 2.7 Hz), 102.42 (d, J = 25.8 Hz), 68.40 (s), 34.07 (s), 23.13 (s), 11.27 (s),
10.27 (s); IR (cm-1): 3056.05, 2932.15, 2678.47, 1712.24, 1628.25, 1587.19, 1508.86, 1452.92,
1438.66, 1419.48, 1348.83, 1275.62, 1238.36, 1165.87, 1119.43, 878.08, 698.54; MS (ESI), m/z:
436.1 (M–H)–. Anal. calcd. for C22H19F4NO4: C, 60.41; H, 4.38; N, 3.20. Found: C, 60.47; H, 4.46;
N, 3.17.
4.1.8. Preparation of 3-(4-((5-(3,5-dimethylisoxazol-4-yl)-2-fluorobenzyl)oxy)-2-fluorophenyl)
propanoic acid (11f)
1
Compound 11f (0.15 g, 60%) as a white solid. m.p.: 139–142°C; H NMR (300 MHz,
DMSO-d6) δ: 12.16 (br s, 1H), 7.55 (d, J = 7.0 Hz, 1H), 7.49–7.29 (m, 2H), 7.22 (t, J = 8.8 Hz,
1H), 6.92 (d, J = 12.2 Hz, 1H), 6.82 (d, J = 8.7 Hz, 1H), 5.18 (s, 2H), 2.76 (t, J = 7.6 Hz, 2H), 2.46
(t, J = 7.6 Hz, 2H), 2.36 (s, 3H), 2.19 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.63 (s),
165.28 (s), 161.48 (s), 160.83 (d, J = 243.5 Hz), 158.14 (d, J = 8.5 Hz), 157.74 (d, J = 10.9 Hz),
131.31 (d, J = 4.0 Hz), 130.99 (d, J = 7.3 Hz), 126.24 (d, J = 3.5 Hz), 124.08 (d, J = 15.2 Hz),
119.61 (d, J = 16.2 Hz), 116.19 (t, J = 3.2 Hz), 115.91 (s), 114.91 (s), 110.87 (d, J = 2.7 Hz),
102.35 (d, J = 26.0 Hz), 63.81 (s), 34.12 (s), 23.16 (s), 11.21 (s), 10.31 (s); IR (cm-1): 3439.53,
2943.95, 1709.60, 1627.59, 1586.03, 1508.26, 1424.83, 1306.05, 1281.20, 1258.41, 1249.56,
1220.61, 1149.83, 1108.46, 1098.59, 1013.71, 836.12, 645.40; MS (ESI), m/z: 386.1 (M–H)–.
Anal. calcd. for C21H19F2NO4: C, 65.11; H, 4.94; N, 3.62. Found: C, 65.20; H, 4.92; N, 3.61.
4.1.9. Preparation of 3-(4-((2-chloro-5-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11g)
1
Compound 11g (0.14 g, 63%) as a white solid. m.p.: 131–132°C; H NMR (300 MHz,
DMSO-d6) δ: 12.18 (br s, 1H), 7.64–7.46 (m, 2H), 7.45–7.30 (m, 1H), 7.20 (t, J = 8.2 Hz, 1H),
6.97–6.67 (m, 2H), 5.11 (s, 2H), 2.75 (t, J = 7.3 Hz, 2H), 2.47 (t, J = 7.3 Hz, 2H), 2.31 (s, 3H),
2.13 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.55 (s), 166.51 (s), 160.82 (d, J = 243.0 Hz),
160.60 (s), 158.19 (d, J = 60.9 Hz), 157.69 (d, J = 43.6 Hz), 133.49 (d, J = 3.5 Hz), 131.30 (d, J =
3.3 Hz), 130.95 (d, J = 6.9 Hz), 129.94 (d, J = 8.4 Hz), 119.42 (d, J = 16.2 Hz), 117.12 (d, J =
16.2 Hz), 116.17 (d, J = 22.4 Hz), 110.96 (d, J = 2.8 Hz), 110.17 (s), 102.37 (d, J = 25.9 Hz),
68.58 (s), 34.10 (s), 23.16 (s), 11.22 (s), 10.11 (s); IR (cm-1): 3433.63, 3050.15, 3002.95, 1726.28,
1627.56, 1508.85, 1464.03, 1365.70, 1296.13, 1152.40, 1130.43, 1110.93, 838.37; MS (ESI), m/z:
402.1 (M–H)–. Anal. calcd. for C21H19ClFNO4: C, 62.46; H, 4.74; N, 3.47. Found: C, 62.43; H,
4.78; N, 3.50.
4.1.9. Preparation of 3-(4-((5-(3,5-dimethylisoxazol-4-yl)-2-methoxybenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11h)
1
Compound 11h (0.13 g, 54%) as a white solid. m.p.: 125–127°C; H NMR (300 MHz,
DMSO-d6) δ: 7.43–7.27 (m, 2H), 7.25–7.09 (m, 2H), 6.92–6.73 (m, 2H), 5.08 (s, 2H), 3.85 (s, 3H),
2.75 (t, J = 7.2 Hz, 2H), 2.45 (t, J = 7.2 Hz, 2H), 2.31 (s, 3H), 2.14 (s, 3H); 13C NMR (75 MHz,
DMSO-d6) δ: 173.54 (s), 164.65 (s), 160.85 (d, J = 243.1 Hz), 158.15 (s), 158.03 (s), 156.46 (s),

ACCEPTED MANUSCRIPT
130.94 (d, J = 6.8 Hz), 129.92 (d, J = 10.3 Hz), 124.78 (s), 121.71 (s), 119.22 (d, J = 15.9 Hz),
115.46 (s), 111.47 (s), 110.75 (d, J = 2.6 Hz), 102.24 (d, J = 25.7 Hz), 64.80 (s), 55.65 (s), 34.12
(s), 23.15 (s), 11.16 (s), 10.35 (s); IR (cm-1): 3091.45, 2979.35, 2843.66, 1740.33, 1628.35,
1609.89, 1585.33, 1509.33, 1491.55, 1464.35, 1405.91, 1292.77, 1252.61, 1155.33, 1103.87,
1037.17, 827.05; MS (ESI), m/z: 398.2 (M–H)–. Anal. calcd. for C22H22FNO5: C, 66.16; H, 5.55;
N, 3.51. Found: C, 66.13; H, 5.62; N, 3.47.
4.1.10. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-2-fluorobenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11i)
1
Compound 11i (0.12 g, 52%) as a white solid. m.p.: 132–134°C; H NMR (300 MHz,
DMSO-d6) δ: 12.20 (br s, 1H), 7.65–7.56 (m, 1H), 7.44 (td, J = 7.5, 1.9 Hz, 1H), 7.34 (t, J = 7.6
Hz, 1H), 7.22 (t, J = 8.8 Hz, 1H), 6.92 (dd, J = 12.1, 2.5 Hz, 1H), 6.82 (dd, J = 8.5, 2.4 Hz, 1H),
5.18 (s, 2H), 2.77 (t, J = 7.5 Hz, 2H), 2.46 (t, J = 7.5 Hz, 2H), 2.33 (s, 3H), 2.15 (s, 3H); 13C NMR
(75 MHz, DMSO-d6) δ: 173.62 (s), 166.55 (s), 160.86 (d, J = 243.3 Hz), 158.67 (s), 157.86 (d, J =
11.1 Hz), 155.93 (s), 131.89 (d, J = 2.6 Hz), 131.03 (d, J = 6.7 Hz), 130.67 (d, J = 4.0 Hz), 124.78
(d, J = 4.3 Hz), 124.41 (d, J = 15.1 Hz), 119.64 (d, J = 15.8 Hz), 117.47 (s), 110.86 (d, J = 2.7 Hz),
102.32 (d, J = 25.8 Hz), 63.96 (s), 34.12 (s), 23.17 (s), 11.32 (s), 10.20 (s); IR (cm-1): 2991.15,
1706.73, 1627.14, 1581.58, 1508.62, 1457.43, 1420.08, 1290.99, 1255.86, 1148.86, 1096.61,
1007.33, 966.47; MS (ESI), m/z: 386.1 (M–H)–. Anal. calcd. for C21H19F2NO4: C, 65.11; H, 4.94;
N, 3.62. Found: C, 65.12; H, 4.96; N, 3.65.
4.1.11. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-4-fluorobenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11j)
1
Compound 11j (0.15 g, 60%) as a white solid. m.p.: 111–113°C; H NMR (300 MHz,
DMSO-d6) δ: 12.15 (br s, 1H), 7.66– .55 (m, 2H), 7.42 (dd, J = 8.2, 2.2 Hz, 1H), 7.22 (t, J = 8.8
Hz, 1H), 6.93 (dd, J = 12.1, 2.4 Hz, 1H), 6.82 (dd, J = 8.4, 2.3 Hz, 1H), 5.19 (s, 2H), 2.76 (t, J =
7.5 Hz, 2H), 2.46 (t, J = 7.5 Hz, 2H), 2.35 (s, 3H), 2.18 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ:
173.57 (s), 165.58 (s), 160.86 (d, J = 243.3 Hz), 158.05 (s), 157.75 (d, J = 11.0 Hz), 134.55 (s),
132.07 (s), 131.06 (d, J = 6.9 Hz), 130.69 (s), 130.28 (s), 129.97 (s), 129.10 (s), 119.66 (d, J =
15.8 Hz), 114.80 (d, J = 1.7 Hz), 110.83 (s), 102.42 (d, J = 26.0 Hz), 67.06 (s), 34.09 (s), 23.15 (s),
11.27 (s), 10.35 (s); IR (cm-1): 2985.25, 1709.06, 1627.47, 1588.01, 1509.58, 1479.08, 1450.46,
1423.33, 1288.56, 1241.81, 1194.47, 1161.44, 1153.56, 1109.62, 1096.28, 1062.48, 1031.96,
968.22, 836.24; MS (ESI), m/z: 386.1 (M–H)–. Anal. calcd. for C21H19F2NO4: C, 65.11; H, 4.94; N,
3.62. Found: C, 65.15; H, 4.88; N, 3.57.
4.1.12. Preparation of 3-(4-((4-chloro-3-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11k)
1
Compound 11k (0.13 g, 54%) as a white solid. m.p.: 143–145°C; H NMR (300 MHz,
DMSO-d6) δ: 12.14 (br s, 1H), 7.64 (d, J = 8.3 Hz, 1H), 7.52 (dd, J = 8.2, 2.0 Hz, 1H), 7.46 (d, J =
1.9 Hz, 1H), 7.21 (t, J = 8.8 Hz, 1H), 6.87 (dd, J = 12.2, 2.4 Hz, 1H), 6.79 (dd, J = 8.4, 2.4 Hz,
1H), 5.14 (s, 2H), 2.76 (t, J = 7.5 Hz, 2H), 2.46 (t, J = 7.5 Hz, 2H), 2.25 (s, 3H), 2.07 (s, 3H); 13
C
NMR (75 MHz, DMSO-d6) δ: 173.56 (s), 166.23 (s), 160.83 (d, J = 243.2 Hz), 158.61 (s), 157.77
(d, J = 11.6 Hz), 136.47 (s), 132.77 (s), 131.64 (s), 130.98 (d, J = 6.7 Hz), 129.69 (d, J = 32.4 Hz),
128.44 (s), 119.49 (d, J = 15.6 Hz), 113.79 (d, J = 19.9 Hz), 110.99 (d, J = 3.0 Hz), 102.42 (d, J =
25.8 Hz), 68.44 (s), 34.07 (s), 23.13 (s), 11.34 (s), 10.18 (s); IR (cm-1): 3398.23, 2973.45, 1736.09,
1628.70, 1588.09, 1509.08, 1460.68, 1357.83, 1295.35, 1189.95, 1160.55, 1149.09, 1114.39,
1097.53, 1085.66, 818.17; MS (ESI), m/z: 402.1 (M–H)–. Anal. calcd. for C21H19ClFNO4: C,

ACCEPTED MANUSCRIPT
62.46; H, 4.74; N, 3.47. Found: C, 62.52; H, 4.69; N, 3.48.
4.1.13. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-4-methoxybenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (11l)
1
Compound 11l (0.15 g, 58%) as a white solid. m.p.: 140–142°C; H NMR (300 MHz,
DMSO-d6) δ: 12.16 (s, 1H), 7.46 (dd, J = 8.5, 2.2 Hz, 1H), 7.28 (d, J = 2.1 Hz, 1H), 7.19 (t, J =
8.8 Hz, 1H), 7.13 (d, J = 8.6 Hz, 1H), 6.86 (dd, J = 12.2, 2.5 Hz, 1H), 6.78 (dd, J = 8.4, 2.4 Hz,
1H), 5.05 (s, 2H), 3.78 (s, 3H), 2.75 (t, J = 7.4 Hz, 2H), 2.45 (t, J = 7.4 Hz, 2H), 2.23 (s, 3H), 2.06
(s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.57 (s), 165.58 (s), 160.83 (d, J = 243.2 Hz), 159.04
(s), 158.04 (d, J = 11.2 Hz), 156.59 (s), 131.22 (s), 130.90 (d, J = 6.7 Hz), 129.64 (s), 128.75 (s),
119.18 (d, J = 15.8 Hz), 118.01 (s), 111.56 (s), 111.01 (d, J = 2.7 Hz), 102.34 (d, J = 25.8 Hz),
69.06 (s), 55.44 (s), 34.10 (s), 23.15 (s), 11.30 (s), 10.32 (s); IR (cm-1): 3445.43, 2932.15, 1732.65,
1625.59, 1508.45, 1458.87, 1441.34, 1363.13, 1297.39, 1253.68, 1191.98, 1146.87, 1098.23,
1027.57, 813.30; MS (ESI), m/z: 398.2 (M–H)–. Anal. calcd. for C22H22FNO5: C, 66.16; H, 5.55;
N, 3.51. Found: C, 66.21; H, 5.56; N, 3.43.
4.1.14. Preparation of 3-bromo-5-nitrobenzaldehyde (13)
To a solution of 3-nitrobenzaldehyde (1.0 g, 6.6 mmol) in concentrated sulfuric acid (4.0 mL)
was added N-bromosuccinimide (1.4 g, 7.9 mmol) in small portions at room temperature and then
heated to 65°C for 1 h. After cooling to the room temperature, the solution was poured into ice
water and the precipitate was filtered. The crude product was dried over Na2SO4 and recrystallized
1
from EtOAc/petroleum ether (1/10) to obtain 13 (1.3 g, 82%) as white crystals. H NMR (300
MHz, DMSO-d6) δ: 10.09 (s, 1H), 8.79–8.55 (m, 2H), 8.51 (s, 1H).
4.1.15. Preparation of 2-(3-bromo-5-nitrophenyl)-1,3-dioxolane (14)
To a solution of 3-bromo-5-nitrobenzaldehyde 13 (0.90 g, 3.9 mmol), ethylene glycol
(2.2 mL, 39 mmol ) and p-TsOH (70 mg, 0.39 mmol) in toluene (25 mL) was refluxed for 5 h.
After cooling to the room temperature, the solution was diluted with EtOAc (25 mL) and washed
with brine (20 mL). The organic layers were dried over Na2SO4 and concentrated to afford a crude
product 14 (0.99 g, 92%) as a pale yellow solid without further purification.
4.1.16. Preparation of 3-bromo-5-(1,3-dioxolan-2-yl)aniline (15)
To a solution of 14 (0.99 g, 3.6 mmol) and ammonium chloride (0.29 g, 5.4 mmol) in
ethanol/H2O (30 mL, 5/2) was added iron powders (1.0 g, 18 mmol) at room temperature. The
reaction mixture was refluxed for 4 h and then filtered. The filtrate was diluted with EtOAc (30
mL) and washed with brine (20 mL). The organic layers were dried over Na2SO4 and concentrated
to afford 15 (0.72 g, 82%) as a yellow oily product. 1H NMR (300 MHz, DMSO-d6) δ: 6.71 (t, J =
1.8 Hz, 1H), 6.65 (s, 1H), 6.60 (s, 1H), 5.56 (s, 1H), 5.46 (s, 2H), 4.03–3.88 (m, 4H).
4.1.17. Preparation of 3-bromo-5-hydroxybenzaldehyde (16)
To a solution of 15 (0.71 g, 6.0 mmol) in AcOH/H2SO4/H2O (5.0 mL, 8/1/1) was added
sodium nitrite aqueous solution (0.24 g, 3.5 mmol, 1 mL) dropwise and stirred for 1 h at 0°C. The
reaction was added 10% sulphuric acid (20 mL) and refluxed for 2 h. After cooling to the room
temperature, the reaction mixture was extracted with EtOAc (3 × 30 mL), washed with saturated
sodium bicarbonate solution (3 × 30 mL) and brine (30 mL). The combined organic layers were
dried over Na2SO4 and concentrated. The residue was purified by column chromatography
(EtOAc/petroleum ether, 1/8) to give 16 (0.47 g, 80%) as a pale yellow solid. 1H NMR (300 MHz,
DMSO-d6) δ: 10.43 (s, 1H), 9.88 (s, 1H), 7.51 (s, 1H), 7.31–7.18 (m, 2H).
4.1.18. Preparation of 3-(3,5-dimethylisoxazol-4-yl)-5-hydroxybenzaldehyde (17)

ACCEPTED MANUSCRIPT
To a mixture of 3,5-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)isoxazole (0.50
g, 2.0 mmol), 16 (0.41 g, 2.0 mmol), Pd(PPh3)4 (0.12 g, 0.10 mmol) and sodium carbonate (0.43 g,
4.0 mmol) in toluene/ethanol/H2O (15 mL, 3/1/3) was refluxed under nitrogen atmosphere for 24 h.
Then the mixture was diluted with saturated ammonium chloride solution and EtOAc, and the
insoluble material was filtered through Celite. The organic layer of the filtrate was washed with
water (30 mL) and brine (30 mL), dried over Na2SO4, and evaporated in vacuo. The residue was
purified by column chromatography (EtOAc/petroleum ether, 1/5 to 1/3) to give 17 (0.30 g, 69%)
as a pale yellow solid. 1H NMR (300 MHz, DMSO-d6) δ: 10.15 (s, 1H), 9.96 (s, 1H), 7.37 (s, 1H),
7.25 (s, 1H), 7.08 (s, 1H), 2.42 (s, 3H), 2.24 (s, 3H).
4.1.19. Preparation of 3-(3,5-dimethylisoxazol-4-yl)-5-methoxybenzaldehyde (18a)
To a solution of 17 (0.14 g, 0.64 mmol), iodomethane (0.11 g, 0.77 mmol) and potassium
carbonate (0.18 g, 1.3 mmol) in DMF (3.0 mL) was stirred at 60°C for 5 h. Then the mixture was
quenched with saturated ammonium chloride solution and extracted with EtOAc (3 × 20 mL). The
combined organic layers were washed with water (30 mL) and brine (30 mL), dried over Na2SO4,
and concentrated in vacuo. The residue was purified by column chromatography
1
(EtOAc/petroleum ether, 1/9 to 1/5) to give 10a (0.12 g, 81%) as a white solid. H NMR (300
MHz, DMSO-d6) δ: 10.03 (s, 1H), 7.52 (s, 1H), 7.43 (s, 1H), 7.29 (s, 1H), 3.88 (s, 3H), 2.43 (s,
3H), 2.25 (s, 3H).
4.1.20. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-methoxybenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (22a)
Compound 22a was prepared from 18a according to the general procedures of compound 11a.
These procedures can also be applied to the synthesis of compounds 22b-h. Compound 22a (80
1
mg, 64%) as a white solid. m.p.: 142–144°C; H NMR (300 MHz, DMSO-d6) δ: 7.19 (t, J = 8.7
Hz, 1H), 7.01 (s, 1H), 6.99 (s, 1H), 6.93–6.72 (m, 3H), 5.11 (s, 2H), 3.79 (s, 3H), 2.72 (t, J = 7.6
Hz, 2H), 2.44–2.30 (m, 5H), 2.20 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 174.36 (s), 165.28 (s),
160.41 (d, J = 204.8 Hz), 159.66 (s), 157.94 (d, J = 22.7 Hz), 138.97 (s), 131.26 (s), 130.90 (d, J =
7.2 Hz), 120.36 (s), 115.71 (s), 113.88 (s), 112.30 (s), 110.87 (d, J = 2.5 Hz), 102.29 (d, J = 25.8
Hz), 69.13 (s), 55.29 (s), 35.47 (s), 23.67 (s), 11.31 (s), 10.43 (s); IR (cm-1): 3457.23, 2938.05,
1711.15, 1628.62, 1593.95, 1508.72, 1457.13, 1417.74, 1334.09, 1287.19, 1253.87, 1156.05,
1114.12, 859.39, 830.45; MS (ESI), m/z: 398.2 (M–H)–. Anal. calcd. for C22H22FNO5: C, 66.16; H,
5.55; N, 3.51. Found: C, 66.21; H, 5.54; N, 3.53.
4.1.21. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-ethoxybenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (22b)
1
Compound 22b (0.10 g, 66%) as a white solid. m.p.: 166–168°C; H NMR (300 MHz,
DMSO-d6) δ: 12.17 (s, 1H), 7.20 (t, J = 8.8 Hz, 1H), 6.98 (d, J = 6.3 Hz, 2H), 6.91–6.76 (m, 3H),
5.11 (s, 2H), 4.07 (q, J = 6.9 Hz, 2H), 2.75 (t, J = 7.3 Hz, 2H), 2.47 (t, J = 7.4 Hz, 2H), 2.37 (s,
3H), 2.20 (s, 3H), 1.33 (t, J = 6.9 Hz, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.53 (s), 165.24
(s), 160.82 (d, J = 243.0 Hz), 158.92 (s), 158.08 (s), 157.91 (d, J = 11.0 Hz), 138.91 (s), 131.26 (s),
130.92 (d, J = 6.9 Hz), 120.21 (s), 119.32 (d, J = 16.0 Hz), 115.72 (s), 114.25 (s), 112.74 (s),
110.95 (d, J = 2.6 Hz), 102.35 (d, J = 25.8 Hz), 69.12 (s), 63.23 (s), 34.07 (s), 23.15 (s), 14.59 (s),
11.30 (s), 10.43 (s); IR (cm-1): 3445.43, 2991.15, 1709.11, 1629.33, 1589.76, 1511.65, 1284.11,
1267.68, 1252.88, 1155.03, 1116.18, 1045.04, 860.63; MS (ESI), m/z: 412.2 (M–H)–. Anal. calcd.
for C23H24FNO5: C, 66.82; H, 5.85; N, 3.39. Found: C, 66.86; H, 5.81; N, 3.41.
4.1.22. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-propoxybenzyl)oxy)-2-fluoro-

ACCEPTED MANUSCRIPT
phenyl)propanoic acid (22c)
Compound 22c (80 mg, 57%) as a pale yellow solid. m.p.: 126–128°C; 1H NMR (300 MHz,
DMSO-d6) δ: 12.16 (br s, 1H), 7.20 (t, J = 8.8 Hz, 1H), 7.00 (s, 1H), 6.97 (s, 1H), 6.91–6.83 (m,
2H), 6.83–6.76 (m, 1H), 5.11 (s, 2H), 3.97 (t, J = 6.5 Hz, 2H), 2.75 (t, J = 7.6 Hz, 2H), 2.45 (d, J =
7.6 Hz, 2H), 2.37 (s, 3H), 2.20 (s, 3H), 1.80–1.65 (m, 2H), 0.98 (t, J = 7.4 Hz, 3H); 13C NMR (75
MHz, DMSO-d6) δ: 173.51 (s), 165.23 (s), 160.80 (d, J = 243.3 Hz), 159.07 (s), 158.07 (s), 157.90
(d, J = 11.0 Hz), 138.90 (s), 131.25 (s), 130.92 (d, J = 6.8 Hz), 120.22 (s), 119.31 (d, J = 16.1 Hz),
115.72 (s), 114.27 (s), 112.82 (s), 110.95 (d, J = 2.8 Hz), 102.35 (d, J = 25.8 Hz), 69.11 (s), 69.03
(s), 34.07 (s), 23.12 (s), 22.00 (s), 11.30 (s), 10.42 (s), 10.39 (s); IR (cm-1): 2938.05, 1713.15,
1629.71, 1589.66, 1509.86, 1420.92, 1337.64, 1282.99, 1264.25, 1251.46, 1154.94, 1116.18,
859.26; MS (ESI), m/z: 426.2 (M–H)–. Anal. calcd. for C24H26FNO5: C, 67.43; H, 6.13; N, 3.28.
Found: C, 67.41; H, 6.10; N, 3.35.
4.1.23. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-isopropoxybenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (22d)
Compound 22d (80 mg, 56%) as a clear oily product. 1H NMR (300 MHz, DMSO-d6) δ: 7.21
(t, J = 9.0 Hz, 1H), 7.01–6.71 (m, 5H), 5.11 (s, 2H), 4.74–4.60 (m, 1H), 2.76 (t, J = 7.0 Hz, 2H),
2.46 (t, J = 7.0 Hz, 2H), 2.38 (s, 3H), 2.20 (s, 3H), 1.34–1.13 (m, 6H); 13C NMR (75 MHz,
DMSO-d6) δ: 173.57 (s), 165.24 (s), 160.82 (d, J = 243.1 Hz), 158.11 (s), 157.99 (s), 157.85 (d, J
= 5.1 Hz), 138.97 (s), 131.29 (s), 130.95 (d, J = 6.9 Hz), 120.10 (s), 119.33 (d, J = 15.8 Hz),
115.74 (s), 115.18 (s), 113.90 (s), 110.97 (d, J = 3.0 Hz), 102.36 (d, J = 25.7 Hz), 69.29 (s), 69.11
(s), 34.11 (s), 23.15 (s), 21.80 (s), 11.36 (s), 10.48 (s); IR (cm-1): 3439.53, 2979.35, 1710.75,
1627.51, 1589.22, 1508.42, 1421.24, 1323.76, 1285.66, 1258.54, 1207.13, 1152.89, 1110.11,
1015.84, 847.88; MS (ESI), m/z: 426.2 (M–H)–. Anal. calcd. for C24H26FNO5: C, 67.43; H, 6.13;
N, 3.28. Found: C, 67.28; H, 6.15; N, 3.29.
4.1.24. Preparation of 3-(4-((3-butoxy-5-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-fluoro-
phenyl)propanoic acid (22e)
1
Compound 22e (80 mg, 52%) as a clear oily product. H NMR (300 MHz, DMSO-d6) δ:
12.21 (br s, 1H), 7.20 (t, J = 8.8 Hz, 1H), 7.00 (s, 1H), 6.97 (s, 1H), 6.91–6.83 (m, 2H), 6.83–6.74
(m, 1H), 5.11 (s, 2H), 4.01 (t, J = 6.5 Hz, 2H), 2.75 (t, J = 7.5 Hz, 2H), 2.45 (t, J = 7.5 Hz, 2H),
2.37 (s, 3H), 2.20 (s, 3H), 1.76–1.64 (m, 2H), 1.50–1.36 (m, 2H), 0.93 (t, J = 7.4 Hz, 3H); 13C
NMR (75 MHz, DMSO-d6) δ: 173.55 (s), 165.26 (s), 160.81 (d, J = 242.8 Hz), 159.09 (s), 158.09
(s), 157.91 (d, J = 11.7 Hz), 138.90 (s), 131.25 (s), 130.94 (d, J = 6.9 Hz), 120.23 (s), 119.99 (d, J
= 66.0 Hz), 115.73 (s), 114.27 (s), 112.82 (s), 110.96 (d, J = 3.0 Hz), 102.35 (d, J = 25.9 Hz),
69.12 (s), 67.33 (s), 34.09 (s), 30.73 (s), 23.13 (s), 18.75 (s), 13.72 (s), 11.33 (s), 10.45 (s); IR
(cm-1): 2967.55, 1710.48, 1627.67, 1592.05, 1508.38, 1421.66, 1328.41, 1304.07, 1285.79,
1259.00, 1207.44, 1152.19, 1109.42, 1098.47, 1024.68, 851.01; MS (ESI), m/z: 440.2 (M–H)–.
Anal. calcd. for C25H28FNO5: C, 68.01; H, 6.39; N, 3.17. Found: C, 67.99; H, 6.35; N, 3.21.
4.1.25. Preparation of 3-(4-((3-(3,5-dimethylisoxazol-4-yl)-5-isobutoxybenzyl)oxy)-2-fluoro-
phenyl)propanoic acid (22f)
1
Compound 22f (90 mg, 62%) as a pale yellow solid. m.p.: 122–124°C. H NMR (300 MHz,
DMSO-d6) δ: 12.10 (br s, 1H), 7.20 (t, J = 8.8 Hz, 1H), 7.01 (s, 1H), 6.97 (s, 1H), 6.90–6.76 (m,
3H), 5.11 (s, 2H), 3.79 (d, J = 6.5 Hz, 2H), 2.76 (t, J = 7.4 Hz, 2H), 2.45 (t, J = 7.3 Hz, 2H), 2.37
(s, 3H), 2.19 (s, 3H), 2.07–1.96 (m, 1H), 0.99 (s, 3H), 0.97 (s, 3H); 13C NMR (75 MHz, DMSO-d6)
δ: 173.57 (s), 165.26 (s), 160.82 (d, J = 243.7 Hz), 159.04 (s), 158.11 (s), 157.90 (d, J = 11.2 Hz),

ACCEPTED MANUSCRIPT
138.89 (s), 131.25 (s), 130.94 (d, J = 6.8 Hz), 120.23 (s), 119.34 (d, J = 15.6 Hz), 115.73 (s),
114.38 (s), 112.79 (s), 110.97 (d, J = 2.7 Hz), 102.37 (d, J = 25.9 Hz), 72.23 (s), 69.12 (s), 34.11
(s), 23.17 (s), 11.33 (s), 10.46 (s), 10.13 (s), 3.15 (s); IR (cm-1): 3463.13, 2908.55, 1716.10,
1628.88, 1588.68, 1508.91, 1423.01, 1335.26, 1305.24, 1282.11, 1263.62, 1248.12, 1202.33,
1157.26, 1115.32, 1100.72, 1028.72, 965.75, 856.72, 830.20; MS (ESI), m/z: 440.2 (M–H)–. Anal.
calcd. for C25H28FNO5: C, 68.01; H, 6.39; N, 3.17. Found: C, 68.00; H, 6.37; N, 3.22.
4.1.26. Preparation of 3-(4-((3-(cyclopropylmethoxy)-5-(3,5-dimethylisoxazol-4-yl)benzyl)
oxy)-2-fluorophenyl)propanoic acid (22g)
1
Compound 22g (0.10 g, 69%) as a clear oily product. H NMR (300 MHz, DMSO-d6) δ:
12.07 (br s, 1H), 7.20 (t, J = 8.8 Hz, 1H), 7.05–6.93 (m, 2H), 6.91–6.74 (m, 3H), 5.11 (s, 2H), 3.86
(d, J = 7.0 Hz, 2H), 2.76 (t, J = 7.5 Hz, 2H), 2.45 (d, J = 7.5 Hz, 2H), 2.37 (s, 3H), 2.19 (s, 3H),
1.29–1.18 (m, 1H), 0.65 – 0.49 (m, 2H), 0.39–0.25 (m, 2H); 13C NMR (75 MHz, DMSO-d6) δ:
173.55 (s), 165.25 (s), 160.81 (d, J = 244.9 Hz), 158.09 (s), 157.97 (d, J = 2.5 Hz), 157.84 (s),
138.91 (s), 131.27 (s), 130.94 (d, J = 7.0 Hz), 120.28 (s), 119.33 (d, J = 16.4 Hz), 115.73 (s),
114.30 (s), 112.91 (s), 110.95 (d, J = 2.8 Hz), 102.36 (d, J = 25.7 Hz), 73.89 (s), 69.12 (s), 34.10
(s), 27.71 (s), 23.15 (s), 19.06 (s), 11.32 (s), 10.44 (s); IR (cm-1): 2960.05, 1711.11,
1627.59,1592.24, 1508.28, 1469.64, 1421.73, 1327.62, 1285.32, 1259.37, 1208.02, 1152.09,
1109.38, 1098.30, 1031.56, 846.61; MS (ESI), m/z: 438.2 (M–H)–. Anal. calcd. for C25H26FNO5:
C, 68.32; H, 5.96; N, 3.19. Found: C, 68.35; H, 6.01; N, 3.15.
4.1.27. Preparation of 3-(4-((3-(cyclopentyloxy)-5-(3,5-dimethylisoxazol-4-yl)benzyl)oxy)-2-
fluorophenyl)propanoic acid (22h)
1
Compound 22h (90 mg, 62%) as a clear oily product. H NMR (300 MHz, DMSO-d6) δ:
12.18 (br s, 1H), 7.20 (t, J = 8.8 Hz, 1H), 6.95 (s, 2H), 6.91–6.74 (m, 3H), 5.10 (s, 2H), 4.86 (t, J =
5.8 Hz, 1H), 2.75 (t, J = 7.5 Hz, 2H), 2.45 (t, J = 7.5 Hz, 2H), 2.38 (s, 3H), 2.20 (s, 3H), 2.00–1.84
(m, 2H), 1.77–1.51 (m, 6H); 13C NMR (75 MHz, DMSO-d6) δ: 173.56 (s), 165.24 (s), 160.83 (d, J
= 242.9 Hz), 159.71 (s), 158.05 (d, J = 6.2 Hz), 157.86 (s), 138.92 (s), 131.22 (s), 130.95 (d, J =
6.9 Hz), 119.97 (s), 119.33 (d, J = 15.8 Hz), 115.75 (s), 114.90 (s), 113.88 (s), 110.97 (d, J = 2.8
Hz), 102.36 (d, J = 25.7 Hz), 78.82 (s), 69.11 (s), 34.11 (s), 32.26 (s), 23.66 (s), 23.15 (s), 11.35
(s), 10.48 (s); IR (cm-1): 3463.13, 2960.75, 1710.08, 1627.41, 1589.73, 1508.18, 1421.09, 1327.49,
1302.92, 1285.50, 1258.64, 1206.58, 1152.32, 1109.13, 1098.16, 1016.96, 849.80; MS (ESI), m/z:
452.2 (M–H)–. Anal. calcd. for C26H28FNO5: C, 68.86; H, 6.22; N, 3.09. Found: C, 68.89; H, 6.17;
N, 3.11.
4.1.28. Preparation of 3-(4-((6-(3,5-dimethylisoxazol-4-yl)pyridin-2-yl)methoxy)-2-fluoro-
phenyl)propanoic acid (28a)
1
Compound 28a (0.11 g, 76%) as a white solid. m.p.: 97–100°C; H NMR (300 MHz,
DMSO-d6) δ: 12.18 (br s, 1H), 7.92 (t, J = 7.8 Hz, 1H), 7.56–7.40 (m, 2H), 7.21 (t, J = 8.9 Hz,
1H), 6.96–6.75 (m, 2H), 5.22 (s, 2H), 2.76 (t, J = 7.4 Hz, 2H), 2.56 (s, 3H), 2.46 (t, J = 7.4 Hz,
2H), 2.37 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.57 (s), 167.50 (s), 160.83 (d, J = 243.2
Hz), 158.38 (s), 157.87 (d, J = 11.3 Hz), 156.54 (s), 149.50 (s), 137.95 (s), 131.00 (d, J = 6.9 Hz),
121.93 (s), 119.83 (s), 119.49 (d, J = 16.0 Hz), 115.16 (s), 110.86 (d, J = 2.8 Hz), 102.36 (d, J =
25.9 Hz), 70.46 (s), 34.05 (s), 23.13 (s), 12.20 (s), 11.26 (s); IR (cm-1): 3415.93, 2926.25, 1739.94,
1624.57, 1584.38, 1571.98, 1510.54, 1474.32, 1449.04, 1439.21, 1296.20, 1185.13, 1166.36,
1102.90, 963.15; MS (ESI), m/z: 369.2 (M–H)–. Anal. calcd. for C20H19FN2O4: C, 64.86; H, 5.17;
N, 7.56. Found: C, 64.89; H, 5.15; N, 7.55.

ACCEPTED MANUSCRIPT
4.1.29. Preparation of 3-(4-((5-(3,5-dimethylisoxazol-4-yl)pyridin-3-yl)methoxy)-2-fluoro-
phenyl)propanoic acid (28b)
1
Compound 28b (0.12 g, 83%) as a white solid. m.p.: 162–164°C; H NMR (300 MHz,
DMSO-d6) δ: 12.13 (br s, 1H), 8.68 (s, 1H), 8.60 (s, 1H), 7.91 (s, 1H), 7.24 (t, J = 8.8 Hz, 1H),
6.92 (d, J = 12.1 Hz, 1H), 6.85 (d, J = 8.4 Hz, 1H), 5.22 (s, 2H), 2.78 (t, J = 7.5 Hz, 2H), 2.48 (d, J
= 7.5 Hz, 2H), 2.43 (s, 3H), 2.25 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.59 (s), 166.25 (s),
160.36 (d, J = 315.7 Hz), 158.05 (d, J = 32.9 Hz), 148.93 (s), 148.09 (s), 135.83 (s), 132.50 (s),
131.04 (d, J = 6.6 Hz), 125.84 (s), 119.63 (d, J = 16.1 Hz), 112.74 (s), 110.98 (d, J = 2.4 Hz),
102.46 (d, J = 25.9 Hz), 67.04 (s), 34.10 (s), 23.16 (s), 11.33 (s), 10.34 (s); IR (cm-1): 3439.53,
2938.05, 1711.52, 1628.40, 1586.42, 1510.45, 1433.59, 1387.10, 1284.10, 1233.01, 1148.92,
1111.69, 841.96, 714.98; MS (ESI), m/z: 369.1 (M–H)–. Anal. calcd. for C20H19FN2O4: C, 64.86;
H, 5.17; N, 7.56. Found: C, 64.83; H, 5.18; N, 7.45.
4.1.30. Preparation of 3-(4-((2-(3,5-dimethylisoxazol-4-yl)pyridin-4-yl)methoxy)-2-fluoro-
phenyl)propanoic acid (28c)
1
Compound 28c (0.11 g, 73%) as a white solid. m.p.: 162–164°C; H NMR (300 MHz,
DMSO-d6) δ: 12.15 (s, 1H), 8.64 (d, J = 5.6 Hz, 1H), 7.51 (s, 1H), 7.35 (d, J = 9.7 Hz, 1H), 7.20 (t,
J = 8.8 Hz, 1H), 6.94–6.71 (m, 2H), 5.23 (s, 2H), 2.74 (t, J = 7.6 Hz, 2H), 2.51 (s, 3H), 2.44 (d, J
= 7.4 Hz, 2H), 2.32 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.59 (s), 167.43 (s), 160.87 (d, J
= 243.3 Hz), 158.34 (s), 157.57 (d, J = 11.2 Hz), 150.14 (s), 149.88 (s), 146.91 (s), 131.09 (d, J =
6.8 Hz), 120.83 (s), 120.07 (s), 119.74 (d, J = 16.2 Hz), 115.36 (s), 110.90 (d, J = 2.9 Hz), 102.45
(d, J = 25.9 Hz), 67.74 (s), 34.06 (s), 23.15 (s), 12.18 (s), 11.27 (s); IR (cm-1): 3433.63, 2926.25,
1719.39, 1627.41, 1585.40, 1511.83, 1449.93, 1423.81, 1296.09, 1194.35, 1182.19, 1153.18,
965.96, 842.49, 824.80, 618.05; MS (ESI), m/z: 369.1 (M–H)–. Anal. calcd. for C20H19FN2O4: C,
64.86; H, 5.17; N, 7.56. Found: C, 64.92; H, 5.23; N, 7.51.
4.1.31. Preparation of 3-(4-((4-(3,5-dimethylisoxazol-4-yl)pyridin-2-yl)methoxy)-2-fluoro-
phenyl)propanoic acid (28d)
1
Compound 28d (0.12 g, 81%) as a white solid. m.p.: 176–179°C; H NMR (300 MHz,
DMSO-d6) δ: 8.72 (d, J = 5.2 Hz, 1H), 7.65 (s, 1H), 7.55 (d, J = 5.1 Hz, 1H), 7.23 (t, J = 8.8 Hz,
1H), 6.93 (d, J = 12.0 Hz, 1H), 6.84 (d, J = 8.5 Hz, 1H), 5.29 (s, 2H), 2.77 (t, J = 7.4 Hz, 2H),
2.49–2.39 (m, 5H), 2.27 (s, 3H); 13C NMR (75 MHz, DMSO-d6) δ: 173.57 (s), 168.11 (s), 160.85
(d, J = 243.3 Hz), 158.04 (s), 157.44 (d, J = 11.1 Hz), 154.64 (s), 146.87 (s), 142.35 (s), 131.14 (d,
J = 6.8 Hz), 123.61 (s), 122.64 (s), 120.02 (d, J = 15.9 Hz), 113.40 (s), 110.95 (d, J = 2.6 Hz),
102.63 (d, J = 25.9 Hz), 68.49 (s), 34.10 (s), 23.19 (s), 11.79 (s), 10.57 (s); IR (cm-1): 3362.83,
3079.65, 1730.02, 1612.11, 1506.15, 1424.41, 1403.28, 1285.17, 1261.58, 1174.88, 1157.83,
1103.49, 964.52, 843.18; MS (ESI), m/z: 369.1 (M–H)–. Anal. calcd. for C20H19FN2O4: C, 64.86;
H, 5.17; N, 7.56. Found: C, 64.86; H, 5.24; N, 7.53.
4.1.32. Preparation of 3-((trimethylsilyl)ethynyl)benzaldehyde (30)
To a mixture of 3-iodobenzaldehyde (0.50 g, 2.2 mmol), trimethylsilylacetylene (0.24 g, 2.4
mmol), Pd(PPh3)4 (70 mg, 0.060 mmol) and CuI (40 mg, 0.19 mmol) in triethylamine (15 mL)
was refluxed under nitrogen atmosphere for 24 h. Then the mixture was filtered through a celite
pad and the filtrate concentrated in vacuo. The residue was purified by column chromatography
(EtOAc/petroleum ether, 1/20 to 1/15) to give 30 (0.31 g, 71%) as a yellow oily product. 1H NMR
(300 MHz, DMSO-d6) δ: 10.00 (s, 1H), 7.98 (s, 1H), 7.91 (d, J = 7.7 Hz, 1H), 7.77 (d, J = 7.7 Hz,
1H), 7.61 (t, J = 7.7 Hz, 1H), 0.25 (s, 9H).

ACCEPTED MANUSCRIPT
4.1.33. Preparation of 3-ethynylbenzaldehyde (31)
To a solution of 30 (0.30 g, 1.5 mmol) in THF (20 mL) was added tetrabutylammonium
fluoride (0.78 g, 3.0 mmol) in small portions at room temperature. The reaction mixture was
stirred for 3 h at room temperature and concentrated in vacuo. The residue was purified by column
chromatography (EtOAc/petroleum ether, 1/20 to 1/10) to give 31 (0.17 g, 88%) as a white solid.
1H NMR (300 MHz, DMSO-d6) δ: 10.01 (s, 1H), 8.00 (s, 1H), 7.93 (d, J = 7.7 Hz, 1H), 7.81 (d, J
= 7.8 Hz, 1H), 7.63 (t, J = 7.7 Hz, 1H), 4.38 (s, 1H).
4.1.34. Preparation of 3-((3,5-dimethylisoxazol-4-yl)ethynyl)benzaldehyde (32)
To a mixture of 4-iodo-3,5-dimethylisoxazole (0.26 g, 1.2 mmol), 31 (0.15 g, 1.2 mmol),
Pd(PPh3)4 (40 g, 0.030 mmol) and CuI (20 mg, 0.10 mmol) in triethylamine (10 mL) was refluxed
under nitrogemn atmosphere for 24 h. Then the mixture was filtered through a celite pad and the
filtrate concentrated in vacuo. The residue was purified by column chromatography
1
(EtOAc/petroleum ether, 1/20) to give 32 (0.22 g, 85%) as a white solid. H NMR (300 MHz,
DMSO-d6) δ: 10.03 (s, 1H), 8.08 (s, 1H), 7.94 (d, J = 7.7 Hz, 1H), 7.88 (d, J = 7.8 Hz, 1H), 7.67 (t,
J = 7.7 Hz, 1H), 2.54 (s, 3H), 2.31 (s, 3H).
4.1.35. Preparation of 3-(4-((3-((3,5-dimethylisoxazol-4-yl)ethynyl)benzyl)oxy)-2-fluoro-
phenyl)propanoic acid (36)
Compound 36 was prepared from 32 according to the general procedures of compound 11a.
Compound 36 (90 mg, 92%) as a white solid. m.p.: 104–106°C; 1H NMR (300 MHz, DMSO-d6) δ:
12.18 (s, 1H), 7.63 (s, 1H), 7.56–7.42 (m, 3H), 7.21 (t, J = 8.8 Hz, 1H), 6.91–6.76 (m, 2H), 5.11 (s,
2H), 2.76 (t, J = 7.6 Hz, 2H), 2.51 (s, 3H), 2.46 (t, J = 7.4 Hz, 2H), 2.29 (s, 3H); 13C NMR (75
MHz, DMSO-d6) δ: 173.62 (s), 171.70 (s), 161.29 (d, J = 178.5 Hz), 159.25 (s), 157.92 (d, J =
11.3 Hz), 137.57 (s), 130.99 (d, J = 6.8 Hz), 130.83 (s), 130.42 (s), 129.02 (s), 128.24 (s), 122.13
(s), 119.45 (d, J = 16.1 Hz), 111.34 (s), 110.89 (d, J = 2.7 Hz), 102.33 (d, J = 25.8 Hz), 93.93 (s),
77.60 (s), 68.95 (s), 34.12 (s), 23.20 (s), 11.72 (s), 10.13 (s); IR (cm-1): 3421.83, 2949.85, 1717.33,
1629.28, 1583.29, 1508.97, 1309.06, 1293.68, 1263.90, 1228.85, 1163.39, 1098.05, 1018.39,
830.69; MS (ESI), m/z: 392.2 (M–H)–. Anal. calcd. for C23H20FNO4: C, 70.22; H, 5.12; N, 3.56.
Found: C, 70.26; H, 5.18; N, 3.49.
4.2. Biological evaluation
4.2.1. Calcium flux assay in the GPR40 FLIPER assay
The culture medium was removed carefully and CHO cells stably expressing human GPR40
were washed with PBS (5 mL). Trypsin (3 mL) was added and cells were incubated in 37°C for 2–
5 min. Then plate medium (10 mL) was added to suspend cells. The cells suspension (50 ꢀL) was
seeded into 96-well plates at a density of 1.5 × 104 cells/well, and the assay plates were placed in
37°C, 5% CO2 incubator for 16–24 hours. After incubation, the plate medium was removed and
dye solution (30 ꢀL) was added to each well. The assay plates were placed in 37°C, 5% CO2 to
incubator for 1 hour. Various concentrations of test compounds were added into the cells, and
calcium flux signal after addition was monitored by FlexStation3 Molecular Devices. EC50 of each
compound was calculated using GraphPad Prism V5.0 software.
4.2.2. Animals
Male ICR mice (20 ± 2 g), male C57BL/6 mice (20 ± 2 g) and male SD rats (200 ± 20 g)
were obtained from Comparative Medicine Centre of Yangzhou University and left to acclimatize
for 1 week before the experimental period. The animal were selected for the study and maintained
at a controlled temperature of 25 ± 2 °C and constant humidity (50–70%) under a 12-h light–dark

ACCEPTED MANUSCRIPT
cycle, with free access to diet and water. The animal study was performed accord the international
rules considering animal experiments and the internationaly accepted ethical principles for
laboratoryanimal use and care.
4.2.2.1. Oral glucose tolerance test in male ICR mice
Male ICR mice aged 10 weeks were fasted for 12 h, weighted, randomized into groups (n =
6). Mice were dosed orally with single doses of vehicle (0.5% methylcellulose aqueous solution),
TAK-875 (suspended in vehicle; 10 mL/kg; 30 mg/kg) or test compounds (suspended in vehicle;
10 mL/kg; 30 mg/kg), 30 min prior to oral glucose load (20% aqueous glucose solution, 2 g/kg).
The blood glucose levels were measured by blood glucose test strips (SanNuo GA-3 type,
ChangSha, China) before administration of vehicle, TAK-875 or test compounds (–30 min). After
glucose oral adminstration, blood glucose were measured at 0, 15, 30, 45, 60 and 120 min.
4.2.2.2. Oral glucose tolerance test in male type 2 diabetic C57BL/6 mice
Male C57BL/6 mice left to acclimatize for 1 week were fed with high-fat diet (MD 12032,
rodent diet with 45 kcal% fat, from Mediscience Ltd., Yangzhou, China) for 12 weeks to induce
insulin resistance. The mice with fasting blood glucose level 11.1 mmol/L or higher were
considered as type 2 diabetic model and selected for acute oral glucose tolerance test[31, 32]. Type
2 diabetic C57BL/6 mice were fasted for 12 h, weighted, randomized into groups (n = 6). Mice
were dosed orally with single doses of vehicle (0.5% methylcellulose aqueous solution), TAK-875
(suspended in vehicle; 10 mL/kg; 30 mg/kg) or test compounds (suspended in vehicle; 10 mL/kg;
30 mg/kg), 30 min prior to oral glucose load (10% aqueous glucose solution, 1 g/kg). The blood
glucose levels were measured by blood glucose test strips (SanNuo GA-3 type, ChangSha, China)
before administration of vehicle, TAK-875 or test compounds (–30 min). After glucose oral
adminstration, blood glucose were measured at 0, 15, 30, 45, 60 and 120 min.
4.2.2.3. Pharmacokinetic profiles of 11k in SD rats
Male SD rats were fasted for 12 h and were administered orally with compound 11k
(suspended in 0.5% methylcellulose aqueous solution; 1 mL/kg; 1 mg/kg). Blood samples were
collected at 0, 0.25, 0.5, 0.75, 1, 2, 4, 8, 12, 24 h after the dose via orbital sinus puncture into
microcentrifuge tubes containing heparin sodium. Then blood samples were centrifuged at 10000
rpm for 10 min to separate plasma. 300 ꢀL of acetonitrile and 5 ꢀL of internal standard (diazepam)
were added to100 ꢀL of plasma sample to precipitate plasma proteins by vortexing and
centrifugation at 14000 rpm for 10 min. The supernatant was concentrated and dissolved in 100
ꢀL of methanol again. The mixture was vortexed and centrifuged at 10000 rpm for 5 min. 20 ꢀL of
the supernatant was injected into the LC-MS/MS system for analysis. Pharmacokinetic profiles
were performed using DAS 2.0 statistical software program.
4.3. Docking studies
Docking studies were performed with Glide docking in Schrodinger 9.4. The crystal structure
of GPR40 with TAK-875 (PDB ID: 4PHU) was downloaded from the Protein Data Bank (PDB).
Before ligand docking, the protein was treated with removal of crystallized ligand, deletion of
water, automatical optimization with ProtAssign. Subsequently, OPLS-2005 force field was added
to constrain and the binding site was defined precisely by grid calculation. Glide docking was
carried out in standard precision mode to ontput one conformation for every molecule and then
these conformations were ranked with the scoring function.
Acknowledgments
This study was supported by the Natural Science Foundation of Jiangsu Province (No. BK

ACCEPTED MANUSCRIPT
20141349) and the China National Key HiTech Innovation Project for the R&D of Novel Drugs
(No. 2013ZX09301303-002).
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COOH
F3C
COOH
O
O
O
N
H
O
S
O
O
O
COOH
TAK-875
GW9508
AMG-837
CF3
O
COOH
Cl
N
N
N
N
COOH
NC
F3C
F
O
O
COOH
O
BMS-986118
DS-1558
TUG-770
COOH
O
F
1
Figure 1. Representative GPR40 agonists.
Figure 2. The interaction mode of compound 11a and TAK-875 within the binding site of hGPR40 and their
overlapped conformations. GPR40 is represented as blue ribbon. The carbons of Arg 183 and Arg 258 are shown in
green. The carbons of compound 11a are shown in yellow. The carbons of TAK-875 are shown in blue. Hydrogen
bonds are represented by red dashes.

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Figure 3. Effect of compounds (30 mg/kg) on oral glucose tolerance test in male ICR mice. (A) show
time-dependent changes of blood glucose after oral administration of compounds, followed by 2 g/kg oral glucose
challenge. Date in (B) represent AUC0–120 min of blood glucose levels. Values are mean ± SEM (n = 6). ** P < 0.01
compared to vehicle-treated ICR mice by Student’s t test; *** P < 0.001 compared to vehicle-treated ICR mice by
Student’s t test.
Figure 4. Effect of compounds (30 mg/kg) on oral glucose tolerance test in type 2 diabetic C57BL/6 mice. (A)
show time-dependent changes of blood glucose after oral administration of compounds, followed by 1 g/kg oral
glucose challenge. Date in (B) represent AUC0–120 min of blood glucose levels. Values are mean ± SEM (n = 6). * P
< 0.05 compared to vehicle-treated type 2 diabetic C57BL/6 mice by Student’s t test; *** P < 0.001 compared to
vehicle-treated type 2 diabetic C57BL/6 mice by Student’s t test.
Scheme
CHO
F
COOMe
COOH
COOH
a
c
b
HO
HO
F
HO
O
F
HO
F
2
3
4
5
N
N
N
O
R3
R1
R3
R3
R3
O
Br
R4
CHO
R2
CHO
R2
OH
R2
Cl
d
e
f
R4
R4
R4
R2
R1
R1
8a-l
R1
6a-l
7a-l
9a-l
COOMe
COOH
N
N
O
R3
R1
R3
O
g
h
O
F
O
F
R4
R2
R4
R2
R1
11a-l
10a-l
Scheme 1. General synthesis of 11a-l. Reagents and conditions: (a) propanedioic acid, pyridine, 80°C; (b) Pd/C, H2,
THF, rt; (c) conc. H2SO4, methanol, reflux; (d) 3,5-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)
isoxazole, Pd(PPh3)4, Na2CO3, toluene, ethanol, H2O, 80°C; (e) NaBH4, methanol, THF, 0°C to rt; (f) POCl3,
N-methyl-2-pyrrolidone, 0°C to rt; (g) K2CO3, DMF, 60°C; (h) LiOH, methanol, H2O, 40°C.

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Scheme 2. General synthesis of 12a-h. Reagents and conditions: (a) NBS, H2SO4, 65°C; (b) ethylene glycol,
p-TsOH, toluene, reflux; (c) Fe, NH4Cl, ethanol, H2O, reflux; (d) NaNO2, 10% H2SO4, AcOH, conc. H2SO4, H2O,
0°C to reflux; (e) 3,5-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)isoxazole, Pd(PPh3)4, Na2CO3,
toluene, ethanol, H2O, 80°C; (f) CH3I or brominated alkanes, K2CO3, DMF, 60°C; (g) NaBH4, methanol, THF, 0°C
to rt; (h) POCl3, N-methyl-2-pyrrolidone, 0°C to rt; (i) K2CO3, DMF, 60°C; (j) LiOH, methanol, H2O, 40°C.
Scheme 3. General synthesis of 28a-d. Reagents and conditions: (a) 3,5-dimethyl-4-(4,4,5,5-tetramethyl-1,3,2-
dioxaborolan-2-yl)isoxazole, Pd(PPh3)4, Na2CO3, toluene, ethanol, H2O, 80°C; (b) NaBH4, methanol, THF, 0°C to
rt; (c) POCl3, N-methyl-2-pyrrolidone, 0°C to rt; (d) K2CO3, DMF, 60°C; (e) LiOH, methanol, H2O, 40°C.
Scheme 4. The synthesis of compound 36. Reagents and conditions: (a) trimethylsilylacetylene, Pd(PPh3)4, CuI,
triethylamine, reflux; (b) TBAF, THF, rt; (c) 4-iodo-3,5-dimethylisoxazole, Pd(PPh3)4, CuI, triethylamine, reflux;
(d) NaBH4, methanol, THF, 0°C to rt; (e) POCl3, N-methyl-2-pyrrolidone, 0°C to rt; (f) K2CO3, DMF, 60°C; (g)
LiOH, methanol, H2O, 40°C.
Table
Table 1
Structures and GPR40 agonistic activities of compounds 11a–l

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Compound R1
R2
R3 R4
hGPR40
EC50 (nM)a
27.5 ± 0.6
7.7e
Emax (%)b
tPSAc cLog Pd
-
-
-
-
100.0 ± 3.3 99.1
- 46.5
4.70
6.16
3.68
4.18
3.85
4.42
4.62
3.85
4.42
3.70
3.85
3.85
4.17
3.14
TAK-875
1
-
-
-
-
H
Me
F
H
H
H
H
H
F
H
H
H
H
H
H
H
H
F
H
H
H
H
H
H
H
H
H
F
24.8 ± 1.4
12.0 ± 3.2
33.2 ± 1.9
38.9 ± 0.4
114 ± 16
11.1 ± 0.8
9.3 ± 1.8
24.3 ± 4.1
25.9 ± 3.1
25.2 ± 7.3
15.9 ± 2.6
103.7 ± 0.3 68.1
107.0 ± 1.9 68.1
11a
11b
11c
11d
11e
11f
82.7 ± 1.8
98.5 ± 0.8
68.1
68.1
Cl
CF3
H
H
H
H
H
H
H
100.6 ± 0.5 68.1
102.4 ± 0.1 68.1
Cl
OMe
H
H
H
H
101.5 ± 0
98.7 ± 1.3
88.8 ± 1.4
68.1
77.4
68.1
11g
11h
11i
H
H
H
101.5 ± 1.2 68.1
105.9 ± 1.0 68.1
108.4 ± 1.3 77.4
11j
Cl
11k
OMe 22.9 ± 0.1
11l
a Calcium flux assay in GPR40-transfected CHO cells, values are mean ± SEM (n = 3).
b % Compared to reference TAK-875, values are mean ± SEM (n = 3).
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
e The EC50 value of compound 1 was presented in reference[17].
Table 2
Structures and GPR40 agonistic activities of compounds 22a–h
Compound
R
hGPR40
Emax (%)b
tPSAc cLog Pd
EC50 (nM)a
53.1 ± 5.3
69.1 ± 1.4
96.7 ± 13.9
79.2 ± 5.6
89.3 ± 5.7
143 ± 10
Me
98.7 ± 1.5
101.4 ± 1.6
102.0 ± 0.2
105.3 ± 0
77.4
77.4
77.4
77.4
77.4
77.4
77.4
3.70
4.23
4.76
4.54
5.29
5.16
4.68
22a
22b
22c
22d
22e
22f
Et
n-Pr
i-Pr
n-Bu
i-Bu
101.3 ± 0.8
102.0 ± 0
cyclopropyl 90.9 ± 0.7
methyl
104.3 ± 0.9
22g
cyclopentyl 102 ± 3
106.2 ± 0.1
77.4
5.18
22h
a Calcium flux assay in GPR40-transfected CHO cells, values are mean ± SEM (n = 3).
b % Compared to reference TAK-875, values are mean ± SEM (n = 3).

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c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
Table 3
Structures and GPR40 agonistic activities of compounds 28a–d
Compound
28a
R
hGPR40
EC50 (nM)a
506 ± 49
Emax (%)b
tPSAc cLog Pd
98.1 ± 0
80.5
80.5
80.5
80.5
2.57
2.36
2.57
2.36
4980 ± 1379 80.5 ± 5.1
28b
862 ± 27
107.6 ± 3.4
85.0 ± 1.0
28c
1930 ± 107
28d
a Calcium flux assay in GPR40-transfected CHO cells, values are mean ± SEM (n = 3).
b % Compared to reference TAK-875, values are mean ± SEM (n = 3).
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
Table 4
Structure and GPR40 agonistic activity of compound 36
Compound hGPR40
EC50 (nM)a
E
max (%)b tPSAc cLog Pd
214 ± 2
99.5 ± 1.0 68.1
5.03
36
a Calcium flux assay in GPR40-transfected CHO cells, values are mean ± SEM (n = 3).
b % Compared to reference TAK-875, values are mean ± SEM (n = 3).
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
Table 5.
Pharmacokinetic profiles of 11k in SD ratsa
Route Dose (mg/kg) T1/2 (h)
po 1.0 2.87 ± 0.33 1.0
a Data are expressed as mean ± SEM (n = 4).
Tmax (h) Cmax (µg/mL)
AUC0-8 h (µg·h/mL)
0.84 ± 0.15
4.18 ± 0.71

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Table 1
Structures and GPR40 agonistic activities of compounds 11a–l
Compound R1
R2
R3 R4
hGPR40
EC50 (nM)a
27.5
Emax (%)b tPSAc cLog Pd
-
-
-
-
100
-
99.1
46.5
68.1
68.1
68.1
68.1
68.1
68.1
68.1
77.4
68.1
68.1
68.1
77.4
4.70
6.16
3.68
4.18
3.85
4.42
4.62
3.85
4.42
3.70
3.85
3.85
4.17
3.14
TAK-875
1
-
-
-
-
7.7e
H
Me
F
H
H
H
H
H
F
H
H
H
H
H
H
H
H
F
H
H
H
H
H
H
H
H
H
F
24.8
104
99
83
98
101
98
102
91
89
95
93
99
11a
11b
11c
11d
11e
11f
12.0
33.2
Cl
CF3
H
H
H
H
H
H
H
38.9
114
11.1
Cl
OMe
H
H
H
H
9.3
11g
11h
11i
24.3
25.9
H
H
H
25.2
11j
Cl
15.9
11k
OMe 22.9
11l
a Calcium flux assay in GPR40-transfected CHO cells. Means of three experiments.
b % Compared to reference TAK-875.
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
e The EC50 value of compound 1 was presented in reference[17].
Table 2
Structures and GPR40 agonistic activities of compounds 22a–h
Compound
R
hGPR40
EC50 (nM)a
53.1
Emax (%)b tPSAc cLog Pd
Me
99
77.4
77.4
77.4
77.4
77.4
77.4
77.4
3.70
4.23
4.76
4.54
5.29
5.16
4.68
22a
22b
22c
22d
22e
22f
Et
69.1
101
94
n-Pr
i-Pr
n-Bu
i-Bu
96.7
79.2
99
89.3
98
143
102
92
cyclopropyl 90.9
methyl
22g
cyclopentyl 102
99
77.4
5.18
22h

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a Calcium flux assay in GPR40-transfected CHO cells. Means of three experiments.
b % Compared to reference TAK-875.
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
Table 3
Structures and GPR40 agonistic activities of compounds 28a–d
Compound
28a
R
hGPR40
EC50 (nM)a
506
Emax (%)b tPSAc
cLog Pd
2.57
98
80.5
80.5
80.5
80.5
4980
862
81
2.36
28b
108
85
2.57
28c
1930
2.36
28d
a Calcium flux assay in GPR40-transfected CHO cells. Means of three experiments.
b % Compared to reference TAK-875.
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
Table 4
Structure and GPR40 agonistic activity of compound 36
Compound hGPR40
EC50 (nM)a
Emax (%)b tPSAc cLog Pd
214
105
68.1
5.03
36
a Calcium flux assay in GPR40-transfected CHO cells. Means of three experiments.
b % Compared to reference TAK-875.
c Topological polar surface area values were calculated by ChemBioDraw Ultra 14.0.
d cLog P values were calculated by the BioByte’s algorithm as implemented in ChemBioDraw Ultra 14.0.
Table 5
Pharmacokinetic profiles of 11k in SD ratsa
Route Dose (mg/kg) T1/2 (h)
po 1.0 2.87 ± 0.33 1.0
a Data are expressed as mean ± SEM (n = 4).
Tmax (h) Cmax (µg/mL)
AUC0-8 h (µg·h/mL)
0.84 ± 0.15
4.18 ± 0.71

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