SAR Studies of N-[2-(1 H-Tetrazol-5-yl)phenyl]benzamide Derivatives as Potent G Protein-Coupled Receptor-35 Agonists

Article abstract of DOI:10.1021/acsmedchemlett.7b00510

G protein-coupled receptor-35 (GPR35) has emerged as a potential target in the treatment of pain and inflammatory and metabolic diseases. We have discovered a series of potent GPR35 agonists based on a coumarin scaffold and found that the introduction of

Full text of DOI:10.1021/acsmedchemlett.7b00510

Subscriber access provided by Eastern Michigan University | Bruce T. Halle Library
SAR Studies of N-[2-(1H-tetrazol-5-yl)-phenyl]-benzamide
Derivatives as Potent G Protein-Coupled Receptor-35 Agonists
Lai Wei, Tao Hou, Chang Lu, Jixia Wang, Xiuli Zhang, Ye Fang, Yaopeng
Zhao, Jiatao Feng, Jiaqi Li, Lala Qu, Hai-long Piao, and Xinmiao Liang
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.7b00510 • Publication Date (Web): 09 Apr 2018
Downloaded from http://pubs.acs.org on April 9, 2018
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a service to the research community to expedite the dissemination
of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in
full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully
peer reviewed, but should not be considered the official version of record. They are citable by the
Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore,
the “Just Accepted” Web site may not include all articles that will be published in the journal. After
a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web
site and published as an ASAP article. Note that technical editing may introduce minor changes
to the manuscript text and/or graphics which could affect content, and all legal disclaimers and
ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or
consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W.,
Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 6
ACS Medicinal Chemistry Letters
1
2
3
4
5
6
7
8
9
SAR Studies of N-[2-(1H-tetrazol-5-yl)-phenyl]-benzamide Deriva-
tives as Potent G Protein-Coupled Receptor-35 Agonists
Lai Wei†,‡,ǁ, Tao Hou†,ǁ, Chang Lu†,‡, Jixia Wang†, Xiuli Zhang*,†,§, Ye Fangǁ, Yaopeng Zhao†, Jiatao
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Feng†, Jiaqi Li†,‡, Lala Qu†,‡, Hailong Piao† and Xinmiao Liang*,†,§
†Key Lab of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of
Sciences, Dalian 116023, China
‡University of Chinese Academy of Sciences, Beijing, 100049, China
Biochemical Technologies, Science and Technology Division, Corning, New York 14831, United States
§Co-innovation Center of Neuroregeneration, Nantong University, Nantong 226019, China
*Corresponding authors: Xiuli Zhang, Xinmiao Liang
KEYWORDS: GPR35, dynamic mass redistribution, Nꢀ[2ꢀ(1Hꢀtetrazolꢀ5ꢀyl)ꢀphenyl]ꢀbenzamide derivatives, Lipinski’s Rule
ABSTRACT: G proteinꢀcoupled receptorꢀ35 (GPR35) has emerged as a potential target in the treatment of pain, inflammatory and
metabolic diseases. We have discovered a series of potent GPR35 agonists based on a coumarin scaffold and found that the introꢀ
duction of a 1H-tetrazolꢀ5ꢀyl group significantly increased their potency. We designed and synthesized a new series of N-[2ꢀ(1H-
tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀbenzamide derivatives through a twoꢀstep synthetic approach, and characterized their agonistic activities
against GPR35 using a dynamic mass redistribution (DMR) assay. N-(5ꢀbromoꢀ2ꢀ(1H-tetrazolꢀ5ꢀyl)phenyl)ꢀ4ꢀmethoxybenzamide
(56) and N-(5ꢀbromoꢀ2ꢀ(1H-tetrazolꢀ5ꢀyl)phenyl)ꢀ2ꢀfluoroꢀ4ꢀmethoxybenzamide (63) displayed the highest agonistic potency agoꢀ
nist GPR35 with an EC50 of 0.059 ꢁM and 0.041 ꢁM, respectively. The physicochemical properties of selected compounds were
calculated to evaluate their druglikeness, suggesting that compounds 56 and 63 have good drugꢀlike properties. Together, Nꢀ[2ꢀ(1Hꢀ
tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀbenzamide derivatives are potentially great candidates for developing potent GPR35 agonists.
G proteinꢀcoupled receptors (GPCRs) as the most successꢀ
ful druggable receptor family have an important position in
drug development.1 However, the physiological functions of
many GPCRs, in particular orphan receptors,2 are far from
clear. Discovering potent probe molecules for these orphan
receptors has great significance in understanding their physioꢀ
logical functions and for drug development. The orphan G
proteinꢀcoupled receptor 35 (GPR35) was first discovered in
1998, and later has been implicated in a variety of diseases,3
such as cardiovascular diseases,4 gastric cancer,3, 5 artery disꢀ
ease6 and type 2 diabetes.7
In the past decade, several endogenous ligands have been
discovered to activate GPR35; these ligands include kynurenic
acid,5 lysophosphatidic acid,8 multiple tyrosine metabolites9
and the mucosal chemokine CXCL17,10 but with variable poꢀ
tency. Several classes of synthetic agonists including the
phosphodiesterase 5 inhibitor zaprinast (1),11 the antiasthma
drugs doxantrazole (2) and pemirolast (3)12 have also been
reported to display GPR35 agonistic activity. Furthermore,
zaprinast is the most widely used reference agonist to eluciꢀ
date the biological functions of GPR35. Many more GPR35
Figure 1. Selected GPR35 agonists with potencies at GPR35
(1-4) and antagonist (5).
agonists
were
discovered
through
highꢀthroughput
14
screening,13,
and only few structureꢀactivity relationship
Our previous studies have shown that tetrazole 4 possessed
significant agonism at GPR3517. Several other studies have
shown that halogen atom substitution also increased the agoꢀ
nistic activity in certain positions.14, 15, 16 The introduction of
lipophilic residues and hydrogen bond accepting groups at
(SAR) studies have been reported in the literature so far.15, 16
ACS Paragon Plus Environment

ACS Medicinal Chemistry Letters
Page 2 of 6
specific positions in the molecule could probably enhance the
tained not only gave rise to a doseꢀdependent DMR in HTꢀ29,
but also desensitized the cells to subsequent stimulation with 1
µM zaprinast. Notably, the potency to trigger DMR for all
compounds was found to be almost equal to that to desensitize
the zaprinast response (Table 1, Table 2), suggesting that
these compounds are GPR35 agonists.
The DMR antagonist assay further showed that the known
GPR35 antagonist MLꢀ145 (5, Fig. 1) doseꢀdependently and
completely blocked the DMR arising from all N-[2ꢀ(1H-
tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀamido derivatives, each at its respective
EC80 to EC100 concentration.20 This suggests that the DMR of
these N-[2ꢀ(1H-tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀamido derivatives were
specific to the activation of GPR35.
activity of compunds.15, 16 Inspired by these findings, a variety
of N-[2ꢀ(1H-tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀamido derivatives with
different halogen atoms at the 4 or 5ꢀpositions were syntheꢀ
sized. The compounds were prepared starting from 6a-6c and
8.
1
2
3
4
5
6
7
8
Scheme 1. Synthesis of Compound 7a and 7ba
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Inspired by our previous findings, a tetrazolyl group was
firstly introduced into the simplest compounds 6a and 6b,
yielding compounds 7a and 7b. Compared to the inactive
compounds 6a and 6b, 7a and 7b both showed moderate poꢀ
tency with an EC50 of 29.99 ꢁM and 44.06 ꢁM, respectively.
This result suggests that the introduction of the tetrazolyl to
the molecule significantly improves the agonistic activity.
Next, an amide linker was introduced into the compound 7b,
given that lipophilic residues and hydrogen bondꢀaccepting
groups may also play an important role in activating GPR35.15
The propanamide (14, EC50 0.62ꢁM) showed more than 50ꢀ
fold increase in potency compared to 7b. The introduction of
the bulkier alkyl substituents further improved potency. For
instance, isopropyl (15, EC50 0.38 ꢁM) and cyclohexyl (16,
EC50 0.44 ꢁM) substituted compounds displayed more than
100ꢀfold increased potency compared to 7b. Compounds with
an aromatic group in this position also had relatively higher
potency, such as furyl (17, EC50 1.06 ꢁM), thienyl (18, EC50
0.52ꢁM) and phenyl (42, EC50 0.87 ꢁM).
aReagents and conditions: (a) NaN3, AlCl3, THF, 90oC, 5h,
yield 81ꢀ86%.
The 2ꢀ(1H-tetrazolꢀ5ꢀyl)aniline derivatives 7a and 7b were
obtained from 6a and 6b via the [3+2] cycloaddition of nitriles
and sodium azide catalyzed by aluminum chloride at 90oC
with excellent yields (84% and 91%, respectively) (Scheme
1).
Scheme 2. Synthesis of Compound 14 and its Derivativesa
Table 1. Potency of 2-Substituted 2-(1H-tetrazol-5-
yl)aniline Derivatives in DMR Assays
aReagents and conditions: (a) R3COCl, pyridine, overnight,
yield 68ꢀ87%. (b) NaN3, AlCl3, THF, 90oC, 5h, yield 69ꢀ91%
The amines 6a-6c, 8 were reacted with various acid chloꢀ
rides in pyridine as the base and solvent at room temperature,
yielding a series of N-(2ꢀcyanophenyl)amido derivatives (19-
41) (Scheme 2, Scheme 3). The N-[2ꢀ(1H-tetrazolꢀ5ꢀyl)ꢀ
phenyl]ꢀbenzamide derivatives (42-64) were synthesized from
N-(2ꢀcyanophenyl)benzamide derivatives (19-41) via a [3+2]
cycloaddition, also with good yields (69% ~ 91%) (Scheme 2,
Scheme 3).
desensitization
IC50b (ꢁM)
antagonist
compd
R1
R2
R3
EC50a (ꢁM)
IC50c (ꢁM)
0.67±0.19
0.67±0.25
0.47±0.11
0.58±0.06
0.43±0.08
0.29±0.08
0.50±0.05
7a
7b
14
15
16
17
18
29.99±2.80
44.06±4.06
0.62±0.06
0.38±0.02
0.44±0.03
1.06±0.13
0.51±0.02
35.56±2.54
75.34±9.31
0.70±0.07
0.28±0.02
0.20±0.01
0.44±0.03
0.26±0.03
Scheme 3. Synthesis of Compound 41 and 64a
Br
Br
Br
Br
Br
H
H
H
H
H
ethyl
isopropyl
cyclohexyl
furyl
aReagents and conditions: (a) 4ꢀMethoxybenzoyl chloride,
pyridine, overnight. (b) NaN3, AlCl3, THF, 90oC, 5h.
thienyl
aEC50 to trigger DMR.bIC50 to desensitize upon cells repeated
All N-[2ꢀ(1H-tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀamido compounds obꢀ
tained were tested using a dynamic mass redistribution (DMR)
assay18 using HTꢀ29, a cell line endogenously expressing
GPR35.19 The reference agonist zaprinast was also used to
perform a DMR desensitization assay.19 Results showed that
all N-[2ꢀ(1H-tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀamido compounds obꢀ
c
stimulation with 1 µM zaprinast. IC50 of known GPR35 anꢀ
tagonist 5 to block the agonism. The data respresent mean ±
sd from two independent measurements, each with four repliꢀ
cates (n = 8).
ACS Paragon Plus Environment

Page 3 of 6
ACS Medicinal Chemistry Letters
N
N
Since the benzoic acid derivatives are easy to obtain, we
chose the phenylꢀsubstituted derivative 42 (N-(5ꢀbromoꢀ2ꢀ
(1H-tetrazolꢀ5ꢀyl)phenyl)benzamide) for further modification.
N-(5ꢀbromoꢀ2ꢀ(1H-tetrazolꢀ5ꢀyl)phenyl)benzamide derivatives
with different substituents in the o-, m-, and p-position were
investigated (monoꢀ or diꢀsubstitutions).
N
N
HN
N
3,
O
O
HN
N
2,
1,
R3
CN
4,
H
1
2
H
N
H
N
H
N
1
N
6
H
2
3
O
O
O
N
R2
O
5
N
N
N
R1
42-63
4
3
37
64
4
5
6
7
8
R3
desensitization
IC50b (ꢁM)
antagonist
IC50c (ꢁM)
For the monoꢀsubstitutions, the same substituent in different
positions of the benzene ring could significantly change the
activity. The following rank order of potency among comꢀ
pounds with the same substituent in different positions (o-, m-,
and p-position) was observed: mꢀmethyl (44, EC50 0.86 ꢁM) ≈
pꢀmethyl (45, EC50 1.33 ꢁM) ＞ oꢀmethyl (43, EC50 2.05 ꢁM)
compd
R1
R2
EC50a (ꢁM)
ortho
meta
para
37
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
inactive
0.87±0.09
2.05±0.11
0.86±0.04
1.33±0.13
0.87±0.10
2.18±0.37
0.52±0.03
0.31±0.03
0.35±0.02
0.61±0.07
0.45±0.02
0.13±0.01
0.86±0.06
1.94±0.08
0.059±0.007
0.83±0.07
0.30±0.03
0.20±0.01
0.65±0.05
4.60±0.46
3.65±0.19
0.041±0.005
17.58±3.30
——
——
9
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
Br
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Br
H
H
H
H
H
H
CH3
H
H
H
H
H
0.30±0.03
0.81±0.05
0.36±0.02
0.51±0.03
0.41±0.03
0.48±0.03
0.21±0.01
0.11±0.01
0.17±0.01
0.21±0.02
0.16±0.01
0.06±0.01
0.40±0.03
0.81±0.07
0.026±0.003
0.26±0.02
0.11±0.01
0.082±0.005
0.28±0.02
1.28±0.08
1.36±0.16
0.014±0.002
8.36±0.99
0.47±0.06
0.19±0.02
0.12±0.03
0.13±0.04
0.77±0.08
0.42±0.11
0.52±0.05
1.11±0.42
0.52±0.07
0.57±0.07
0.18±0.06
0.22±0.03
0.28±0.05
0.56±0.08
0.20±0.09
1.00±0.16
1.38±0.15
0.34±0.08
1.00±0.22
0.84±0.06
0.25±0.05
1.69±0.16
0.63±0.19
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(Figure S1a); pꢀchloro (48, EC50 0.52 ꢁM) ＞ oꢀchloro (46,
CH3
H
H
EC50 0.87 ꢁM) ＞ mꢀchloro (47, EC50 2.18 ꢁM) (Figure S1b);
pꢀfluoro (52, EC50 0.45 ꢁM) ≈ oꢀfluoro (50, EC50 0.35ꢁM) ＞
mꢀfluoro (51, EC50 0.61 ꢁM) (Figure S1c); pꢀmethoxyl (56,
EC50 0.059 ꢁM) ＞ oꢀmethoxyl (54, EC50 0.86 ꢁM) ＞ mꢀ
methoxyl (55, EC50 1.94 ꢁM) (Figure S1d); pꢀtrifluoromethyl
(54, EC50 3.65 ꢁM) ＞ mꢀtrifluoromethyl (55, EC50 4.60ꢁM).
These results suggest that substituents in paraꢀposition led to a
considerable increase in potency except for the methylꢀ
derivatives with better tolerance in metaꢀposition. For N-(5ꢀ
bromoꢀ2ꢀ(1H-tetrazolꢀ5ꢀyl)phenyl)benzamide derivatives with
varying substituents in the paraꢀposition, the rank order of
potency was as follows: methoxyl (56, EC50 0.059 ꢁM) ＞
fluoro (52, EC50 0.45 ꢁM) ＞ chloro (48, EC50 0.52 ꢁM) ＞
H
CH3
H
Cl
H
H
Cl
H
H
H
Cl
Cl
F
H
Cl
H
H
H
F
H
H
H
F
F
H
F
methyl (45, EC50 1.33 ꢁM) ＞ trifluoromethyl (54, EC50 3.65
ꢁM), suggesting that methoxyl in the paraꢀposition was favorꢀ
able for the agonistic activity.
OCH3
H
H
H
OCH3
H
H
H
OCH3
OCH3
OCH3
OCH3
OCH3
H
H
OCH3
H
H
F
H
H
H
H
H
Figure 2. (a) Real time kinetic responses of 63 at different
doses in HTꢀ29 cells. (b) DMR amplitudes of compound 63 as
a function of doses compared with the doseꢀdependent desenꢀ
sitization of zaprinast DMR by 63, and the doseꢀdependent
inhibition of the DMR of 100 nM compound 63 by compound
5. The data represents mean ± sd from two independent measꢀ
urement, each with four replicates (n=8).
For the diꢀsubstitution of the benzamide ring, the rank order
of potency of o,pꢀdichloro (49), o,pꢀdifluoro (53), m,pꢀ
dimethoxy (57) was as follows: o,pꢀdifluoro (53, EC50 0.13
ꢁM) ＞ o,pꢀdichloro (49, EC50 0.31 ꢁM) ＞ m,pꢀdimethoxy
(57, EC50 0.83ꢁM), suggesting that the o,p-positions led to a
visible increase in activity compared to the oꢀ or p-monoꢀ
substituted compounds. However, the diꢀmethoxy in m,pꢀ
positions (57) led to a large reduction in activity compared to
compound 56, suggesting that bulky m,pꢀsubstituted comꢀ
pounds are not well tolerated, but o,pꢀdiꢀsubstitutions are.
Compounds 50 and 56 showed the highest agonistic potency
among the derivatives. Therefore, we devised the synthesis of
compound 63, which was substituted with a fluoro in oꢀ
position and a methoxy in the pꢀposition. As expected, comꢀ
pound 63 displayed further improved potency (EC50 0.041ꢁM,
Figure 2).
Br
Br
Br
H
CF3
H
H
CF3
OCH3
OCH3
F
H
aEC50 to trigger DMR.bIC50 to desensitize cells upon cells
repeated stimulation with 1µM zaprinast. cIC50 of known
GPR35 antagonist 5 to block the agonistꢀinduced DMR. The
data represents mean ± sd from two independent measureꢀ
ments, each with four replicates (n = 8).
There is evidence that the halogen atom bromine is very
important for the retention of compound activity agonist
GPR35.14ꢀ17, 21 In order to study the effect of the change of the
bromine atom on the activity of the compound, we chose the
compound 56 for further study. Results showed that transꢀ
forming the substitution position of bromine atom (58, EC50
0.30ꢁM), replacing the bromine with a fluorine (59, EC50
0.20ꢁM), or removing the bromine (60, EC50 0.65ꢁM) all deꢀ
creased potency.
Compound 37, a synthetic precursor of 60 was also tested
and found to be devoid of activity, demonstrating the necessity
of the tetrazole. The activity was completely lost when the
Table
2.
Potency
of
N-(2-(1H-tetrazol-5-
yl)phenyl)benzamide Derivatives in DMR Assay
ACS Paragon Plus Environment

ACS Medicinal Chemistry Letters
Page 4 of 6
42
6.06
6.06
6.06
6.28
6.50
6.46
6.21
6.35
6.88
6.07
7.23
6.08
6.52
6.70
6.19
7.38
2.23
2.73
2.19
3.02
2.93
2.04
2.45
2.45
2.21
2.29
2.36
2.01
2.36
1.64
1.31
2.15
0.29
0.28
0.28
0.29
0.28
0.29
0.28
0.29
0.30
0.26
0.31
0.24
0.28
0.29
0.28
0.32
3.83
3.33
3.87
3.26
3.57
4.42
3.76
3.90
4.67
3.78
4.87
4.07
4.16
5.06
4.88
5.23
tetrazolyl was replaced by cyano. (Figure S1e). In addition,
changing the position of the tetrazoyl also significantly reꢀ
duced the activity of the compound (64, EC50 17.58 ꢁM).
1
2
3
4
5
6
7
8
44
46
48
49
50
51
52
53
54
56
57
58
59
60
63
We further examined the activity of compounds 53 and 56
through the ERK phosphorylation. Compound 53 and comꢀ
pound 56 increased ERK phosphorylation (Figure. 3a). As
control, the known GPR35 agonist zaprinast also triggered
ERK phosphorylation (Figure. 3b). Moreover, the GPR35
antagonist MLꢀ145 attenuated ERK phosphorylation induced
by these compounds (Figure. 3b). These results suggested that
compounds 53 and 56 induved the phosphorylation of ERK
via the activation of GPR35.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Figure 3. (a) Western blot of ERK1/2 and pꢀERK1/2 after
treatment with compounds 53 and 56 for 15 minutes at conꢀ
centrations of 100 nM, 1 ꢁM respectively. (b) Western blot of
ERK1/2 and pꢀERK1/2 after treatment with the known
GPR35 agonist Zaprinast (Zap) at 1 ꢁM and western blot of
ERK1/2 and pꢀERK1/2 after treatment with compounds for
10 minutes at the same concentrations as in (a) in the presence
of MLꢀ145 (25 ꢁM) for 5 minutes.
aCalculated by the Chembiodraw Ultra 11.0.
LLE (LEE = pEC50 ꢀ clogP), another useful parameter,
combines the potency and lipophilicity. Among these comꢀ
pounds, 14, 15, 56, 59, 60 and 63 showed a LLE value of 5,. A
suitable drug candidate should have a LE ＞ 0.3 and LLE ＞
5. In summary, the new compounds 56 and 63 with potent
potency showed a very good clogP, and suitable LE and LLE
values.
In conclusion, a series of N-[2ꢀ(1H-tetrazolꢀ5ꢀyl)ꢀphenyl]ꢀ
amido derivatives were synthesized as potent GPR35 agonists
through a twoꢀstep synthesis method. SAR analysis showed
that a bromine substituent in the 5ꢀposition and a pꢀmethoxyꢀ
benzamide in the 2ꢀposition would significantly improve the
activity (56, EC50 0.059 ꢁM). The o,pꢀdistribution such as o,pꢀ
dichloro (49) and o,pꢀdifluoro (53) could also improve the
potency when compared with the oꢀ or p-monosubstituted
analogs. Combining these findings, compound 63 was syntheꢀ
sized and found to display the highest potency (EC50 0.041
ꢁM) and good physicochemical properties. Together, this
study provides a new series of potent GPR35 agonists, such as
56 and 63, which may become useful leading ligands to furꢀ
ther elucidate the physiological roles of GPR35.
In order to evaluate the druglikeness of a compound, several
physiochemical properties such as partition coefficient
(clogP), the ligand efficiency (LE) and the ligandꢀlipophilicity
efficiency (LLE) were introduced.22ꢀ24 These parameters were
calculated for the compounds with relatively high potency,
including compounds 14-16, 18, 42, 44, 46, 48-54, 56-60 and
63 (Table 3). Lipinski’s Rule of Five considered that the
clogP value of a druglike compound should be lower than 5.25,
26
The clogP values of all these selected compounds were
found to be within this range. In fact, most of these comꢀ
pounds exhibited a clogP value between 2 and 3, which is
favorable for orally administered drugs.
The LE combines physiochemical with pharmacological
properties, and represents the binding force between each atꢀ
om and receptors and calculates as follows: LE = pEC50/N (N
= nonꢀhydrogen atoms). The LE of most selected compounds
were found to be about 0.3, except for compounds 54 and 57.
ASSOCIATED CONTENT
Supporting Information
Table 3. Physicochemical Properties of Selected Com-
pounds
The Supporting Information is available free of charge on the
ACS Publications website.
Experimental details and characterization data for the re-
ported compounds; NMR spectra; and biological assays.
compd
14
pEC50
6.21
6.42
6.36
6.29
clogPa
1.27
1.58
2.78
1.98
LE
LLE
4.94
4.84
3.58
4.31
0.37
0.36
0.30
0.31
AUTHOR INFORMATION
Corresponding Author
15
16
*For Xiuli Zhang: phone, +86 411 84379519; Eꢀmail, zhangꢀ
xiuli@dicp.ac.cn.
18
ACS Paragon Plus Environment

Page 5 of 6
ACS Medicinal Chemistry Letters
*For Xinmiao Liang: phone, +86 411 84379519; Eꢀmail,
liangxm@dicp.ac.cn,
(11) Taniguchi, Y.; TonaiꢀKachi, H.; Shinjo, K. Zaprinast, a
wellꢀknown cyclic guanosine monophosphateꢀspecific
phosphodiesterase inhibitor, is an agonist for GPR35. FEBS
Lett. 2006, 580, 5003ꢀ5008.
(12) MacKenzie, A. E.; Caltabiano, G.; Kent, T. C.; Jenkins,
L.; McCallum, J. E.; Hudson, B. D.; Nicklin, S. A.; Fawcett,
L.; Markwick, R.; Charlton, S. J.; Milligan, G. The antiallergic
mast cell stabilizers lodoxamide and bufrolin as the first high
and equipotent agonists of human and rat GPR35. Mol
Pharmacol 2014, 85, 91ꢀ104.
(13) Deng, H. Y.; Hu, H. B.; Ling, S. Z.; Ferrie, A. M.; Fang,
Y. Discovery of Natural Phenols as G ProteinꢀCoupled
Receptorꢀ35 (GPR35) Agonists. ACS Med. Chem. Lett. 2012,
3, 165ꢀ169.
(14) Deng, H. Y.; Hu, H. B.; He, M. Q.; Hu, J. Y.; Niu, W. J.;
Ferrie, A. M.; Fang, Y. Discovery of 2ꢀ(4ꢀMethylfuranꢀ2(5H)ꢀ
ylidene)malononitrile and Thieno 3,2ꢀb thiopheneꢀ2ꢀ
carboxylic Acid Derivatives as G ProteinꢀCoupled Receptor
35 (GPR35) Agonists. J. Med. Chem. 2011, 54, 7385ꢀ7396.
(15) Funke, M.; Thimm, D.; Schiedel, A. C.; Muller, C. E. 8ꢀ
Benzamidochromenꢀ4ꢀoneꢀ2ꢀcarboxylic Acids: Potent and
1
2
3
4
5
6
7
8
Author Contributions
ǁ
These authors contributed equally to this work.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
9
This work is supported by Project of National Science Foundation
of China (81473436). We are also grateful for the support by the
State Key Program of National Natural Science of China (Grant
No.U1508221) and innovation program (DICP TMSR201601) of
science and research from DICP, CAS.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
ABBREVIATIONS USED
GPR35, G proteinꢀcoupled receptor 35; GPCR, G proteinꢀ
coupled receptor; SARs, structure activity relationships; DMR,
dynamic mass redistribution; THF, tetrahydrofuran; LE, ligand
efficiency; LLE, ligandꢀlipophilicity efficiency.
Selective Agonists for the Orphan
G ProteinꢀCoupled
Receptor GPR35. J. Med. Chem. 2013, 56, 5182ꢀ5197.
(16) Thimm, D.; Funke, M.; Meyer, A.; Muller, C. E. 6ꢀ
Bromoꢀ8ꢀ(4ꢀ[(3)H]methoxybenzamido)ꢀ4ꢀoxoꢀ4Hꢀchromeneꢀ
2ꢀcarboxylic Acid: a powerful tool for studying orphan G
proteinꢀcoupled receptor GPR35. J Med Chem 2013, 56, 7084ꢀ
99.
(17) Wei, L.; Wang, J. X.; Zhang, X. L.; Wang, P.; Zhao, Y. P.;
Li, J. Q.; Hou, T.; Qu, L. L.; Shi, L. Y.; Liang, X. M.; Fang, Y.
Discovery of 2HꢀChromenꢀ2ꢀone Derivatives as G Proteinꢀ
Coupled Receptorꢀ35 Agonists. J. Med. Chem. 2017, 60, 362ꢀ
372.
REFERENCES
(1) RaskꢀAndersen, M.; Almen, M. S.; Schioth, H. B. Trends
in the exploitation of novel drug targets. Nat. Rev. Drug
Discov. 2011, 10, 579ꢀ590.
(2) Chung, S.; Funakoshi, T.; Civelli, O. Orphan GPCR
research. Br. J. Pharmacol. 2008, 153, S339ꢀS346.
(3) O'Dowd, B. F.; Nguyen, T.; Marchese, A.; Cheng, R.;
Lynch, K. R.; Heng, H. H. Q.; Kolakowski, L. F.; George, S.
R. Discovery of three novel Gꢀproteinꢀcoupled receptor genes.
Genomics 1998, 47, 310ꢀ313.
(4) Min, K. D.; Asakura, M.; Liao, Y. L.; Nakamaru, K.;
Okazaki, H.; Takahashi, T.; Fujimoto, K.; Ito, S.; Takahashi,
A.; Asanuma, H.; Yamazaki, S.; Minamino, T.; Sanada, S.;
Seguchi, O.; Nakano, A.; Ando, Y.; Otsuka, T.; Furukawa, H.;
Isomura, T.; Takashima, S.; Mochizuki, N.; Kitakaze, M.
Identification of genes related to heart failure using global
gene expression profiling of human failing myocardium.
Biochem. Biophys. Res. Commun. 2010, 393, 55ꢀ60.
(5) Wang, J. H.; Simonavicius, N.; Wu, X. S.; Swaminath, G.;
Reagan, J.; Tian, H.; Ling, L. Kynurenic acid as a ligand for
orphan G proteinꢀcoupled receptor GPR35. J. Biol. Chem.
2006, 281, 22021ꢀ22028.
(6) Sun, Y. V.; Bielak, L. E.; Peyser, P. A.; Turner, S. T.;
Sheedy, P. E.; Boerwinkle, E.; Kardia, S. L. R. Application of
machine learning algorithms to predict coronary artery
calcification with a sibshipꢀbased design. Genet. Epidemiol.
2008, 32, 350ꢀ360.
(7) Costa, V.; Federico, A.; Pollastro, C.; Ziviello, C.; Cataldi,
S.; Formisano, P.; Ciccodicola, A. Computational Analysis of
Single Nucleotide Polymorphisms Associated with Altered
Drug Responsiveness in Type 2 Diabetes. Int. J. Mol. Sci.
2016, 17, 14.
(8) Oka, S.; Ota, R.; Shima, M.; Yamashita, A.; Sugiura, T.
GPR35 is a novel lysophosphatidic acid receptor. Biochem.
Biophys. Res. Commun. 2010, 395, 232ꢀ237.
(18) Ferrie, A. M.; Wu, Q.; Fang, Y. Resonant waveguide
grating imager for live cell sensing. Appl. Phys. Lett. 2010, 97,
3.
(19) Deng, H. Y.; Hu, H. B.; Fang, Y. Tyrphostin analogs are
GPR35 agonists. FEBS Lett. 2011, 585, 1957ꢀ1962.
(20) Deng, H. Y.; Fang, Y. Discovery of nitrophenols as
GPR35 agonists. Medchemcomm 2012, 3, 1270ꢀ1274.
(21) Deng, H. Y.; Fang, Y. Synthesis and Agonistic Activity at
the GPR35 of 5,6ꢀDihydroxyindoleꢀ2ꢀcarboxylic Acid
Analogues. ACS Med. Chem. Lett. 2012, 3, 550ꢀ554.
(22) Edwards, M. P.; Price, D. A. Role of physicochemical
properties and ligand lipophilicity efficiency in addressing
drug safety risks. Annu. Rep. Med. Chem. 2010, 45, 380ꢀ391.
(23) Ertl, P.; Rohde, B.; Selzer, P. Fast calculation of molecular
polar surface area as a sum of fragmentꢀbased contributions
and its application to the prediction of drug transport
properties. J. Med. Chem. 2000, 43, 3714ꢀ3717.
(24) Kuntz, I. D.; Chen, K.; Sharp, K. A.; Kollman, P. A. The
maximal affinity of ligands. Proc. Natl. Acad. Sci. U. S. A.
1999, 96, 9997ꢀ10002.
(25) Leeson, P. D.; Springthorpe, B. The influence of drugꢀlike
concepts on decisionꢀmaking in medicinal chemistry. Nat. Rev.
Drug Discov. 2007, 6, 881ꢀ890.
(26) Lipinski, C. A.; Lombardo, F.; Dominy, B. W.; Feeney, P.
J. Experimental and computational approaches to estimate
solubility and permeability in drug discovery and development
settings. Adv. Drug Deliv. Rev. 2012, 64, 4ꢀ17.
(9) Deng, H. Y.; Hu, H. B.; Fang, Y. Multiple tyrosine
metabolites are GPR35 agonists. Sci. Rep. 2012, 2, 12.
(10) MaravillasꢀMontero, J. L.; Burkhardt, A. M.; Hevezi, P.
A.; Carnevale, C. D.; Smit, M. J.; Zlotnik, A. Cutting Edge:
GPR35/CXCR8 Is the Receptor of the Mucosal Chemokine
CXCL17. J. Immunol. 2015, 194, 29ꢀ33.
ACS Paragon Plus Environment

ACS Medicinal Chemistry Letters
Page 6 of 6
Insert Table of Contents artwork here
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
6
ACS Paragon Plus Environment