Discovery of LY3325656: A GPR142 agonist suitable for clinical testing in human

Article abstract of DOI:10.1016/j.bmcl.2019.126857

The discovery and optimization of a novel series of GPR142 agonists are described. These led to the identification of compound 21 (LY3325656), which demonstrated anti-diabetic benefits in pre-clinical studies and ADME/PK properties suitable for human dosing. Compound 21 is the first GPR142 agonist molecule advancing to phase 1 clinic trials for the treatment of Type 2 diabetes.

Full text of DOI:10.1016/j.bmcl.2019.126857

Journal Pre-proofs
Discovery of LY3325656: a GPR142 agonist suitable for clinical testing in hu-
man
Lian Zhu Liu, Tianwei Ma, Jingye Zhou, Zhi Long Hu, Xue Jun Zhang, Hai
Zhen Zhang, Mi Zeng, Jia Liu, Lei Li, Yi Jiang, Zack Zou, Fan Wang, Lei Zhang,
Jianfeng Xu, Jingru Wang, Fei Xiao, Xiankang Fang, Haixia Zou, Alexander
M. Efanov, Melissa K Thomas, Hua V. Lin, Jiehao Chen
PII:
S0960-894X(19)30831-5
DOI:
https://doi.org/10.1016/j.bmcl.2019.126857
BMCL 126857
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
16 August 2019
20 November 2019
25 November 2019
Please cite this article as: Zhu Liu, L., Ma, T., Zhou, J., Long Hu, Z., Jun Zhang, X., Zhen Zhang, H., Zeng, M.,
Liu, J., Li, L., Jiang, Y., Zou, Z., Wang, F., Zhang, L., Xu, J., Wang, J., Xiao, F., Fang, X., Zou, H., Efanov, A.M.,
Thomas, M.K., Lin, H.V., Chen, J., Discovery of LY3325656: a GPR142 agonist suitable for clinical testing in
human, Bioorganic & Medicinal Chemistry Letters (2019), doi: https://doi.org/10.1016/j.bmcl.2019.126857
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Bioorganic & Medicinal Chemistry Letters
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Discovery of LY3325656: a GPR142 agonist suitable for clinical testing in human
Lian Zhu Liua,b, Tianwei Maa , Jingye Zhoua, Zhi Long Hua, Xue Jun Zhanga, Hai Zhen Zhanga, Mi Zenga,
Jia Liua, Lei Lia, Yi Jianga, Zack Zoua, Fan Wanga, Lei Zhanga, Jianfeng Xua, Jingru Wanga, Fei Xiaoa,
Xiankang Fanga, Haixia Zoua,b, Alexander M. Efanovc, Melissa K Thomasc, Hua V. Lina, Jiehao Chenc
a Lilly China Research and Development Center (LCRDC), Eli Lilly & Company, Building 8, 338 Jia Li Lue Road, Shanghai 201203, PR China
b Lilly China Innovation and Partnerships (LCIP), Eli Lilly & Company, 16F, Tower1 HKRI, Taikoo Hui 288 Shimenyi Road, Shanghai, 200041, PR China
c Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN 46285, United States
ARTICLE INFO
ABSTRACT
Article history:
Received
Revised
Accepted
Available online
The discovery and optimization of a novel series of GPR142 agonists are described. These led to
the identification of compound 21 (LY3325656), which demonstrated anti-diabetic benefits in
pre-clinical studies and ADME/PK properties suitable for human dosing. Compound 21 is the
first GPR142 agonist molecule advancing to phase 1 clinic trials for the treatment of Type 2
diabetes.
2019 Elsevier Ltd. All rights reserved.
Keywords:
Type 2 diabetes
Glucose-dependent insulin secretion
GPR142 agonist
Metabolism
———
 Corresponding author. Tel.: +1-317-433-1095; fax: +1-317-277-7287; e-mail: chen_jiehao@lilly.com

Type 2 diabetes mellitus (T2DM) is a prevalent disease derived
from a combination of insulin resistance and insufficient insulin
secretion from pancreatic β cells. T2DM is characterized by
hyperglycemia in the fasting state and/or after administration of
glucose bolus during an oral glucose tolerance test [1]. An
estimated 135 million people worldwide are affected by T2DM
while the number of Americans affected ranges between 11.6
million and 14 million [2]. Patients with T2DM are faced with
increased risk for macro- and microvascular complications such as
coronary heart disease, stroke, neuropathy, and retinopathy [3].
Insulin secretagogues such as sulfonylureas and meglitinides are
widely used for the treatment T2DM. However, their action is
independent of blood glucose concentration and, thus, leads to
increased risk of hypoglycemia [4]. Over the past two decades,
significant efforts have been made to discover novel anti-diabetic
agents, from which DPP-4 inhibitors, GLP-1 receptor agonists and
SGLT2 inhibitors have emerged as safe and effective treatment
options [5-7]. Despite the progress, nearly half of T2DM patients
still fail to meet glycemic goals. Therefore, it is highly desirable to
discover novel oral anti-diabetic agents that can offer other
benefits in addition to safe glycemic control [8].
Our effort to identify novel GPR142 agonists started with
screening of approximately 200K compounds in the Lilly’s
proprietary library, leading to only one hit with micro molar level
GPR142 agonistic activity. Early assessment of the screening hit
was focused on understanding key pharmacophore requirement,
improving metabolic stability and mitigating inhibition on
Scheme 1. Synthesis of compound 6 a. (Boc)2O, DIEA, H2O,
Dioxane, 25-30 oC, 98%; b. NaH, DMF, MeI, 0-25 oC, 16 h, 100%;
c. HCl, EtOAc; d. 3-(trifluoromethyl)benzoyl chloride, Et3N,
DCM; e. chiral SFC; f. LiOH, 96%; g. EDC-HCl, pyridine, 56%.
Insulin Secretion in IPGTT
(T=15min)
10
8
a
#
b
25000
20000
***
%
6
-9
-11%
-12%
Table 1
6 16
-
In vitro charaterization of
***
-27% ***
4
-34% **
***
15000
10000
5000
*
R-247%
2
O
or1 or1
0
N
N
e
n
i
l
N
pk
m
c
t
p
mpk
5
hi
i
5
1
gl
a
t
n
i
Ve
6
t
i
O
p
N
nd
ou
gli
+ S
6
0
a
t
0.01 0.03 0.1
0.3
1
10
Si
Veh
nd
omp
C
ou
R1
Compound 6 (mpk)
omp
C
d
EC50 (Emax)a
b
Compound c
2
Met. %
600
R1
R2
G protein-coupled receptors (GPCRs) are important cell surface
mediators of signal transduction and a class of druggable targets
that have been extensively studied for therapeutic intervention.
GPR142 is a Gq-coupled, type A GPCR with highly enriched
expression in pancreatic islets [9-10]. It is selectively activated by
aromatic amino acids with L-tryptophan being the most potent and
efficacious. Several groups including ours have reported that
activation of GPR142 results in glucose dependent insulin
secretion (GDIS) with low risk of hypoglycemia [11]. We have
also reported that GPR142 agonism is beneficial to β-cell health
[12]. In addition to GDIS, we and others have found that
tryptophan and synthetic GPR142 ligands impact the levels of
incretin hormones and glucagon in rodents, which suggests this
mechanism has the potential to become oral incretin secretagogues
[13].
(nM)
O
(m/r/d/h)
500
**
***
O
a
b
O
***
1.4
###
40N0 H
CF32
300
N
O
1
***
2.5
6
Methyl
***
***
30
H
(104%)
89/41/28/32
***
OH
OH
***
6b
***
6a
7
**
*
CF3
isopropyl
1
31 (115%)
76/52/33/48
***
200
cis racemic
cis racemic
***
0
Vephicle
m
r
p
o
ou
52 (118%)
Sitagliptin 5mpk
Compound 6 15mpk + Sitagliptin 5mpk
p
n
y
d
l
6
m
15
e
m
t
p
h
k
yl
97/88/66/72
*
cycl
i.p.
Cpd Glu
8
9
10
C
0
F3
C
o
o
2.8 mM G
11.1 mM G
+
p.o.
+
+
+
0
10uM Compound 6
-60
0
20
40
60
80 100 120
CF
2,2-difluoroethyl
63 (105%)
64/59/38/58
O
3
Time (min)
c
d
O
O
HN
10
N
CF3
O
2,2,2-trifluoroethyl
81 (114%)
67/59/38/62
O
Figure 2. Pharmacology studies of compound 6 (a) IPGTT study
O
6d
N
6c
in CmFale C57BL/6 mice, 6 was dosed 30 minutes prior to glucose
11
93 (96%)
cis racemic
56/21/6/26
3
cis crahceamllicenge; (b) GDIS study in mouse islet; IPGTT study in male
DIO mice (c) effNect on glOucose; (d) effect on plasma insulin
12
13
CF3
147 (103%)
38/29/24/20
O
level. Data are mean ± SEM.NN=5 per group. *,**,***: p<0.05,
O
O
0.01, 0.001 vs vehicle.
N
CF3
51 (97%)
Yield 44%
67/63/19/38
CF3
O
6f
(absolute)
O
Several synthetic GPR142 agonists (1-5 as listed in Figure 1)
have been reported with characterization of their ex-vivo
pharmacology in rodent and human islets as well as in vivo
pharmacology in rodents and non-human primates [14-18].
However, to the best of our knowledge, none of these molecules
has been progressed to human trials so far. Herein, we report the
identification of compound 21 (LY3325656), a novel GPR142
agonist with an attractive combination of potency and drug-like
properties, which becomes the first compound to test GPR142
agonism for the treatment of type 2 diabetes in humans [19].
inactive
14
OCF3
Methyl
71 (104%)
90/37/40/42
e
N
Figure 1
Examples of GPR142 Agonists
O
+
O
15
CF3
OCF3 2,2,2-trifluoroethyl
110 (114%)
O
86/56/51/76
O
6e
N
N
OH
O
cis racemic
f
N
16
Cl
2,2-difluoroethyl
N
104 (99%)
54/79/19/51
N
O
N
N
NH
N
N
NH
O
H
NH
a. ref [22] for assay protocol
Yield 45%
CF3
b.% Turnover of parent compound after 30 min of incubation with m/r/d/h
6g
(absolute)
1
2
microsomes. See [23] for assay protocol
active
EC50 54 nM
(Amgen)
EC50 52 nM
(Amgen)
H2N
HCl
6i
N
O
H
O
O
N
N
N
N
N
N
N
OH
N
H
N
O
N
N
g
N
NH
N
O
N
N
6
N
N
O
CF3
O
N
6h
CF3
OCF3
4
3
EC50 63 nM
(Merck)
EC50 22 nM
(Amgen)
O
N
N
Cl
N
N
H
Cl
N
N
5
EC50 27 nM
(Merck)
Cytochrome P450 (Cyp) enzymes, which led to the identification
of compound 6. This compound showed an EC50 of 30 nM in a
cell-based GPR142 mediated inositol phosphate (IP) accumulation
assay. Compound 6 is a very weak inhibitor of the major types of
cytochrome P450 enzymes at 10 uM. However, additional
structure-activity-relationship (SAR) efforts on the imidazole R2
substitutions did not result in desired improvement for potency and
cross species metabolic stability (Table 1).

essentially a conformational space holder of the pharmacophore,
we envisioned that by incorporating a hetero atom into the ring,
we should mitigate oxidative metabolism and improve the overall
physical properties while retaining GPR142 activity.
Compound 6 was synthesized through the route outlined in
Scheme 1. Commercially available compound 6a was first
protected with tert-butyloxycarbonyl (Boc) and then esterified to
give methyl ester 6c with excellent yield. Removal of the Boc
group followed by acylation with 3-(trifluoromethyl)benzoyl
chloride led to 6e, which was then separated through chiral
supercritical fluid chromatography (SFC) to give two enantiomers
6f and 6g. The desired isomer 6g was hydrolyzed to acid 6h.
Condensation between 6h and 1,4-dimethyl-1H-imidazol-5-amine
6i generated compound 6 with moderate yield. Synthesis of
compounds 7-16 followed similar routes as compound 6.
To test this hypothesis, we prepared a number of analogues of
compound 6, with 2-pyran to replace the cyclohexyl ring.
Consistent with our hypothesis, we saw significant improvement
of in vitro metabolic stability and retention of in vitro potency.
However, it came to our attention that the right-hand amide was
cleaved quite extensively soon after the parent molecule was dosed
in animals (mouse, rat, dog, and monkey) [20]. To solve this
problem, we explored a number of amide surrogates, from which
the un-substituted 1,3,4-triazole emerged as the best option
considering the combination of good in vitro potency and in vivo
pharmacokinetic properties, as shown by compound 21 (Table 2
and 3). Meanwhile, compound 21 exhibited reduced renal
clearance, presumably due to the lowered pKa (5.9) as compared
to compound 6 (pKa 6.9). Application of the 1,3,4-triazole amide
surrogate back to the cyclo-hexyl core led to compound 18, which
showed improved cross-species pharmacokinetic properties, as
compared to compound 6, but not as dramatic as compound 21
(Table 3). The 3-fold improvement in potency for compound 18 is
likely due to the better conformation offered by the cyclo-hexyl
core.
Compound 6 was tested in the intraperitoneal glucose tolerance
test (IPGTT) in lean mice for initial pharmacological proof-of-
concept in (Figure 2). The compound demonstrated a robust, dose-
dependent glucose lowering effect. In isolated mouse pancreatic
islets, it also stimulated glucose dependent insulin secretion.
Compound 6 also lowered glucose and increased insulin in IPGTT
in diet-induced obese (DIO) mice. When dosed together with a
DPP-4 inhibitor sitagliptin in DIO mice, compound
6
demonstrated an additive effect on both glucose reduction and
insulin secretion.
Further in vivo pharmacokinetic studies with compound 6
showed significant differences in cross species clearances (Table
3). This could be the result of differences in cross-species
metabolic stability and/or clearance pathways, e.g. renal or biliary
clearance. Metabolite identification studies suggested significant
metabolism occurring on the cyclo-hexyl ring, in addition to minor
right-hand amide bond cleavage. Since the cyclohexyl ring is
Synthesis of 21 followed the route shown in Scheme 2.
Commercially available carboxylic acid 21a was transferred to
methyl ester 21b, which was then hydrogenated to give 21c as a
pair of diastereo isomers. Compound 21c was oxidized with
Pyridinium Chlorochromate (PCC) to generate 21d at an overall
yield of 72% for 3 steps. Reductive amination with methylamine
Table 3
6 18 21
Parmacokinetic profiles of
,
,
Table 2
18 31
-
In vitro characteriaztion of
Dog PKb
Vss %F
Rat PKa
X
O
Cyp inhibition%
at 10 uM
3A4/2C9/2D6
Compound
H
N
Cl
Vss
%F
28
Cl
N
R2
(min/ml/kg) (L/kg)
(min/ml/kg) (L/kg)
N
N
R1
6
49.6
1.3
17.6c
1.2 66
6.1/14/0
EC50 (IP)a
(nM)
Met. %b
(m/r/d/h)
Compound
R1
R2
X
18
21
40.1
22.1
1.0
1.1
29
63
8.8
0.9 86
1.3 78
3.3/0/0
R3
8.4d
22/11/16
N
N
a. Compounds were dosed 1 mpk for IV study, 10 mpk for oral study
b. Compounds were dosed 1 mpk for IV study, 5 mpk for oral study
c. Fraction of renal clearance is 20%
R3= Me
17
18
19
20
21
22
23
CH2
CF3
11 (117%)
64/43/46/67
R3= Me
R3= H
CH2
CH2
Cl
14 (111%)
76 (101%)
39/55/33/26
36/36/9/13
d. Fraction of renal clearance <5%
CF3
was followed by acylation with 3-(trifluoromethyl)benzoyl
chloride and chiral supercritical fluid chromatography (SFC) to
furnish 21e as a single cis enantiomer. Compound 21e was then
hydrolyzed to acid 21f under mild condition. Imidazole 21g was
first methylated under Mitsunobu condition and then converted to
hydrazide 21i with hydrazine hydrate. Condensation between 21i
and 21f was followed by dehydration to give oxadiazole 21j at
96% yield. In our initial effort, compound 21j was transformed to
21 directly by heating together with ammonium acetate in acetic
acid under microwave. However, the yield was low and
purification was difficult. The use of microwave also limited the
scale of the reaction. A stepwise approach was then developed to
enable large scale synthesis. By heating together with benzyl
amine in xylene in the presence of catalytic amount of p-
Toluenesulfonic acid (TsOH), 21j was transferred to benzyl
protected triazole 21k at 63% yield. Finally, benzyl group was
removed by hydrogenation to complete the synthesis of 21 at 82%
yield.
R3= 2,2-difluoroethyl
R3= Me
CH2
O
CF3
CF3
41 (109%)
43 (109%)
30 (110%)
27 (112%)
16 (121%)
14 (123%)
82/67/83/90
22/14/13/14
39/14/16/10
17/15/16/7
46/13/48/82
83/64/63/69
R3= Me
O
OCF3
Cl
R3= Me
O
R3= c-butyl
O
CF3
CF3
24
25
R3= cyclopropylmethyl
O
N
26
27
28
CH2
CH2
O
CF3
CF3
CF3
46 (98%)
32 (102%)
53 (107%)
53/41/22/69
70/80/52/84
39/16/7/18
N
O
N
N
N
N
O
29
30
O
O
CF3
Cl
N
N
102 (108%)
35 (102%)
27/28/17/12
66/23/46/29
In preclinical studies, compound 21 lowered blood glucose
level through stimulation of GDIS and incretin hormones GLP-1
N
N
a. ref [21] for assay protocol
b.% Turnover of parent compound after 30 min of incubation with m/r/d/h
microsomes. See ref [22] for assay protocol

O
O
O
O
a
b
c
O
O
OH
O
O
O
O
HO
21d
O
O
O
O
21b
21a
21c
d,e,f
O
O
O
O
abasbs
abasbs
O
g
OH
N
N
O
O
21e
O
O
absabs
21f
CF3
CF3
N
O
j, k
N
N
N
N
N
N
N
21j
CF3
O
O
N
i
h
O
N
N
O
O
N
l
N
n
21g
21i
21h
Ph
O
O
O
O
abasbs
N
abasbs
N
N
m
N
N
N
N
N
N
N
N
N
21
21k
CF3
CF3
Scheme 2. Synthesis of compound 21 a. MeOH/H2SO4, 67 oC, 8 h, 73%; b. Pd/C, EtOAc, 35 oC, H2, 16 h; c. PCC,DCM, r.t, 16 h, 72% for
2 steps; d. CH3NH2-HCl, NaBH(OAc)3; e. m-CF3-benzoyl chloride/Et3N/DCM; f. chiral SFC, 25% from 21d; g. LiOH/MeOH/THF, 99%;
h. MeOH, PPh3, DEAD; i. Hydrazine hydrate, EtOH, 38% for 2 steps; j. HATU, DIPEA, THF, 15 oC; k. Burgess, THF,15 oC, 96% isolated
yield; l. Benzylamine, TsOH, xylene, 120 oC, 63%; m. Pd(OH)2 /C, MeOH, H2 40-45 psi, 45 oC, 82%; n. NH4OAc, AcOH, MicroWave,
150 oC, 26%
and GIP [23]. In addition to its attractive in vivo activities in
preclinical models, compound 21 also demonstrated good
properties for further clinical development, including high
solubility (>2.0 mg/mL for pH 2, 6 and 7.5) and permeability
(13.46 x 10-6 cm/s), with simulated maximum absorbable human
dose ≥2000 mg at standard gastric pH. It is neither an inhibitor nor
inducer of cytochrome P450 enzymes. The compound also showed
acceptable safety profiles, with no-observed-adverse-effect level
(NOAEL) as 200 mg/kg and 180 mg/kg in rat and dog respectively
in the 1-month repeat-dose tox studies, providing enough safety
window to test this first-in-class mechanism for type 2 diabetes in
human studies.
All authors are current or former employees of Eli Lilly and
Company and may hold stocks of Eli Lilly and Company.
Declare of ethics in animal study
All animal experiments followed the National Institutes of
Health guide for the care and use of Laboratory animals (NIH
Publications No. 8023, revised 1978).
Funding source
Eli Lilly and Company provided support in the form of salaries
for all authors (during employed periods) and funding for research
materials, but did not have any additional role in the study design,
data collection and analysis, decision to publish, or preparation of
the manuscript.
In summary, a novel class of GPR142 agonists have been
identified through our lead generation and optimization effort.
Compound 21, LY3325656, demonstrated very reasonable
pharmacokinetic properties, good permeability and solubility,
acceptable safety profiles, along with strong effect on glucose,
insulin and incretins, making it a suitable clinical candidate to test
the hypothesis of GPR142 agonism in human. To the best of our
knowledge, this is the first GPR142 agonist progressed to clinic
test. Further results of pharmacological studies and clinical studies
will be reported in separate manuscripts.
References and notes
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(IP-1 coupled to an organic HTRF acceptor) followed by cryptate
solution. The plates are incubated at 25 °C for 1 hr. Fluorescence is
read in an Envision Instrument at 665 nm and 620 nm wavelength.
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levels using an IP-1 standard curve. The data is fitted to a 4
parameter-fit logistics to determine EC50 values.
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6218−6223.
22. Microsome stability was measured in a high throughput format by
incubating compounds at 20 uM concentration with mouse(m),
rat(r), dog(d) or human (h) microsomes at 37 °C and the percentage
of the parent compounds remaining were measured after 30 min. of
incubation by liquid chromatography/mass spectrometry analysis.
The % of parent compounds which was metabolized is reported as
% turnover.
23. E. Pratt; T., Oura, X. Ma; H. V. Lin, T. Ma; A. Efanov; M.
Thomas. 20-OR: GPR142 Agonists Increase Glucose-Dependent
Insulin Secretion and Differentially Regulate Hormone Secretion
in Preclinical and Phase 1 Studies. https://doi.org/10.2337/db19-
20-OR
12. Lin, H. V.; Wang, J.; Wang, J.;, Li W.; Wang, X.; Alston, J. T.;
Thomas M. K.; Briere, D. A.; Syed, S. K.; Efanov, A. M. GPR142
prompts glucagon-like Peptide-1 release from islets to improve β
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13. 1) Lin, H.V., Efanov, A.M., Fang, X., Beavers, L.S., Wang, X.,
Wang, J., et al., 2016. GPR142 controls tryptophan-induced insulin
and incretin hormone secretion to improve glucose metabolism.
PLoS One 11:e0157298. 2) Rudenko O, Shang J, Munk A, Ekberg
J P, Petersen N, Engelstoft M S, Egerod K L, Hjorth S A, Wu M,
Feng Y, Zhou Y P, Mokrosinski J, Thams P, Reimann F, Gribble F,
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The aromatic amino acid sensor GPR142 controls metabolism
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Fu, Z.; Liu, Q.; Zhang, Y.; Motani, A.; Oda, K.; Okuyama, R.; Nara,
F.; Murakoshi, M.; Fu, A.; Reagan, J. D.; Fan, P.; Xiong, Y.; Shen,
W.; Li, L.; Houze, J.; Medina, J. C. Phenylalanine derivatives as
GPR142 agonists for the treatment of Type II diabetes. Bioorg.
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Supplementary Material
Supplementary material including synthetic procedures and
assay protocols are available.
Declaration of Interest Statement
All authors are current or former employees of Eli Lilly and
Company and may hold stocks of Eli Lilly and Company.
15. Yu, M.; Lizarzaburu, M.; Motani, A.; Fu, Z.; Du, X.; Liu, J.; Jiao,
X.; Lai, S.; Fan, P.; Fu, A.; Liu, Q.; Murakoshi, M.; Nara, F.; Oda,
K.; Okuyama, R.; Reagan, J. D.; Watanabe, N.; Yamzaki, M.;
Xiong, Y.; Zhang, Y.; Zhuang, R.; Lin, D. C.; Houze, J. B.; Medina,
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Kim, Y.; Lizarzaburu, M.; Lively, S.; Lawlis, S.; Murakoshi, M.;
Nara, F.; Watanabe, N.; Reagan, J. D.; Tian, H.; Fu, A.; Motani, A.;
Liu, Q.; Lin, Y.-J.; Zhuang, R.; Xiong, Y.; Fan, P.; Medina, J.; Li,
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17. Wilson, J. E.; Kurukulasuriya, R.; Sinz, C.; Lombardo, M.; Bender,
K.; Parker, D.; Sherer, E. C.; Costa, M.; Dingley, K.; Li, X.;
Mitelman, S.; Tong, S.; Bugianesi, R.; Ehrhardt, A.; Priest, B.;
Ratliff, K.; Ujjainwalla, F.; Nargund, R.; Hagmann, W. K.;
Edmondson, S. Discovery and development of benzo-[1,2,4]-
triazolo-[1,4]-oxazepine GPR142 agonists for the treatment of
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C.; Buist, N.; Terebetski, J.; Kelly, T.; Bugianesi, R.; Priest, B. T.;
Dingley, K. H.; Li, X.; Mitelman, S.; Salituro, G.; Trujillo, M. E.;
Pachanski, M.; Kirkland, M.; Powles, M. A.; Eiermann, G. J.; Feng,
Y.; Shang, J.; Howard, A. D.; Ujjainwalla, F.; Sinz, C. J.;
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19. ClinicalTrials.gov Identifier: NCT03115099
20. Data of 2-pyran analogues compounds is not published.
21. HEK293 cells expressing human GPR142 are maintained in
DMEM supplemented with 10% FBS and 800 ug/mL G418
(Geneticin®) at 37 °C and 5% CO2. The cells are plated in 384 well
plates at 5000 cells per well and allowed 18 hours for attachment.
After addition of compounds at varying concentrations ranging
from 30 uM to 1 nM, cells are incubated for 1 hour. IP-1
measurements are performed using an IP-One HTRF® assay kit