Tetrahydroindazole derivatives as potent and peripherally selective cannabinoid-1 (CB1) receptor inverse agonists

Article abstract of DOI:10.1016/j.bmcl.2016.09.025

A series of potent and receptor-selective cannabinoid-1 (CB1) receptor inverse agonists has been discovered. Peripheral selectivity of the compounds was assessed by a mouse tissue distribution study, in which the concentrations of a test compound in both plasma and brain were measured. A number of peripherally selective compounds have been identified through this process. Compound 2p was further evaluated in a 3-week efficacy study in the diet-induced obesity (DIO) mouse model. Beneficial effects on plasma glucose were observed from the compound-treated mice.

Full text of DOI:10.1016/j.bmcl.2016.09.025

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Tetrahydroindazole derivatives as potent and peripherally selective
cannabinoid-1 (CB1) receptor inverse agonists
⇑
Jay M. Matthews, James J. McNally, Peter J. Connolly, Mingde Xia, Bin Zhu , Shawn Black, Cailin Chen,
Cuifen Hou, Yin Liang, Yuting Tang, Mark J. Macielag
Janssen Research & Development, L.L.C., Welsh & McKean Roads, Spring House, PA 19477, USAy
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of potent and receptor-selective cannabinoid-1 (CB1) receptor inverse agonists has been discov-
ered. Peripheral selectivity of the compounds was assessed by a mouse tissue distribution study, in which
the concentrations of a test compound in both plasma and brain were measured. A number of peripher-
ally selective compounds have been identified through this process. Compound 2p was further evaluated
in a 3-week efficacy study in the diet-induced obesity (DIO) mouse model. Beneficial effects on plasma
glucose were observed from the compound-treated mice.
Received 3 August 2016
Revised 7 September 2016
Accepted 8 September 2016
Available online xxxx
Keywords:
Cannabinoid receptor
CB1 receptor
Ó 2016 Elsevier Ltd. All rights reserved.
Inverse agonist
Peripheral selectivity
Peripherally restricted
The cannabinoid receptors are part of the endocannabinoid sys-
tem, which regulates many important biological processes. Two
such receptors have been identified to date as cannabinoid-1
(CB1) receptor1 and cannabinoid-2 (CB2) receptor.2 The CB2 recep-
tor is mainly located in the immune system and regulates inflam-
matory responses.3 The CB1 receptor is expressed abundantly in
the central nervous system (CNS) and in peripheral tissues such
as liver, skeletal muscle, adipose tissue, and pancreas.4 Antagonism
of the CB1 receptor leads to decreasing food intake and increasing
insulin sensitivity, and has emerged as an attractive approach to
treat obesity and related metabolic diseases such as type 2 dia-
betes. Unfortunately, undesirable psychiatric side effects led to
the withdrawal of the only marketed brain penetrating CB1 recep-
tor inverse agonist/antagonist, rimonabant 1, as well as the termi-
nation of the development of other clinical-stage agents in this
class. However, in recent years, an increasing amount of evidence
suggested that some of the metabolic benefits seen in treatment
with CB1 receptor inverse agonists/antagonists may result from
actions in the peripheral tissues. Therefore peripherally restricted
CB1 receptor inverse agonists/antagonists that do not cross
blood–brain barrier (BBB) may be useful therapeutics for metabolic
disease without causing CNS-mediated adverse effects.5–9
Herein, we report the discovery of a series of potent and periph-
erally selective CB1 receptor inverse agonists 2 that originated
from our earlier brain-penetrant CB1 receptor inverse agonist pro-
gram (Fig. 1).10 The synthesis of compound 2 is illustrated in
Scheme 1. Cyclohexanone (n = 1) or cycloheptanone (n = 2) 3 was
reacted with aryl aldehyde 4 in aqueous sodium hydroxide solu-
tion at elevated temperature to give the condensation product 5.
Treatment of compound 5 with lithium bis(trimethylsilyl)amide
at -78 °C followed by diethyl oxalate led to compound 6. Conden-
sation of 6 with aryl hydrazine 7 was achieved in the presence of
trifluoroacetic acid (TFA) in dioxane at elevated temperature to
give pyrazole 8. Hydrolysis of the ethyl ester of compound 8 under
basic conditions yielded carboxylic acid 9, which was then coupled
O
N
N
O
N
R
NH
Me
NH
n
N
N
Cl
Cl
Ar2
n = 1, 2
2
Ar1
Cl
⇑
Corresponding author.
1 (rimonabant)
y
All authors were employed by Janssen Research & Development, L.L.C. at the time
the work reported herein was conducted.
Figure 1.
http://dx.doi.org/10.1016/j.bmcl.2016.09.025
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Matthews, J. M.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.025

2
J. M. Matthews et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
O
with amine 10 under standard amide formation conditions (HATU,
Et3N) to give final product 2.
O
O
O
OLi
Ar1
H
OEt
b
Ar1
Ar1
4
a
The in vitro CB1 and CB2 receptor inverse agonist activities of
compounds 2 were assessed in cell-based functional assays mea-
suring cyclic adenosine monophosphate (cAMP) production.11
Compounds with acceptable CB1 receptor potency and CB1/CB2
receptor selectivity were then evaluated in a mouse tissue distri-
bution study.12 In this study, male C57bl/6j mice were dosed orally
with a test compound at 20 mg/kg, and the concentrations of the
test compound in both plasma and brain were measured at either
a 2 h or 4 h time point. The in vitro CB1 and CB2 receptor inverse
agonist activity and in vivo tissue distribution data of cyclohex-
anone-derived compounds 2a–k (6-membered ring series, n = 1)
are shown in Table 1, and those of cycloheptanone-derived com-
pounds 2l–w (7-membered ring series, n = 2) are shown in Table 2.
The calculated topological polar surface area (TPSA) and cLogP
data are also included in the tables.
O
n = 1, 2
H
n
n
6
5
3
O
O
OEt
N
OH
Ar2
NH2
d
7
c
n
N
n
N
N
N
Ar2
Ar2
Ar1
Ar1
8
9
O
R
NH
R
NH2
n
10
N
N
e
Ar2
Ar1
2
As demonstrated by the data in Table 1, both aryl (2a–f) and
heteroaryl (2g–k) Ar1 groups led to good CB1 receptor inverse ago-
nist activity, while substituted aryls are the optimal Ar2 groups in
terms of CB1 potency. Additionally, the R group plays an important
role. Compounds with 1-(pyridin-2-yl)piperidin-4-yl as the R
Scheme 1. Reagents and conditions: (a) NaOH, H2O, 65 °C; (b) (i) LiHMDS, Et2O,
ꢀ78 °C; (ii) diethyl oxalate, ꢀ78 °C to rt; (c) CF3CO2H, 1,4-dioxane; (d) aq NaOH,
EtOH; (e) HATU, DIPEA, DMF.
Table 1
In vitro pharmacology and in vivo tissue distribution of compounds 2a–k
O
N
R
NH
N
Ar2
r1
2a - 2k
A
Compd.
Ar1
Ar2
R
CB1 EC50
(l
M)
CB2 EC50
(lM)
Plasma
Brain Conc.
(nmol/g)
B/P
ratio
cLogP
TPSA
Conc. (
l
M)
N
Cl
Cl
2a
2b
0.030
0.034
0.854
>10
1.60a
0.32a
0.04a
0.20
0.09
7.40
7.33
59.8
63.1
Cl
Cl
N
Cl
Cl
N
0.45a
N
N
N
Cl
Cl
F
Cl
Cl
Cl
2c
0.127
0.007
>10
3.72b
1.61a
0.97b
1.28a
0.26
0.82
6.68
6.72
59.8
59.8
2d
1.61
2e
0.004
>10
1.56b
1.39b
0.92
6.83
59.8
Cl
Cl
N
F
Cl
N
2f
0.012
0.022
0.035
>11
>10
3.82
1.52b
2.93b
2.96b
0.035b
1.91b
0.24b
0.02
0.65
0.08
7.71
5.27
7.32
50.2
72.7
88.1
N
N
Cl
Cl
Cl
Cl
2g
2h
N
S
S
S
S
Cl
N
N
N
Cl
Cl
Cl
Cl
2i
0.037
0.018
0.011
>10
>10
>10
3.18a
0.46a
4.63a
0.10a
0.051a
0.64a
0.03
0.11
0.14
7.20
7.56
7.59
88.1
88.1
88.1
Cl
F3C
2j
Cl
F3C
2k
Cl
a
Sample was collected at 4 h.
Sample was collected at 2 h. B/P ratio: brain/plasma concentration ratio.
b
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J. M. Matthews et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
Table 2
In vitro pharmacology and in vivo tissue distribution of compounds 2l–w
O
N
R
NH
N
Ar2
Ar1
2l - 2w
Compd.
Ar1
Ar2
R
CB1 EC50
(lM)
CB2 EC50
(
lM)
Plasma
Brain Conc.
(nmol/g)
B/P ratio
cLogP
TPSA
Conc. (lM)
N
N
Cl
Cl
2l
0.007
0.037
0.006
0.028
0.035
0.743
>10
0.83b
0.13b
0.05b
BLOQb
0.05b
0.07b
0.16
0.02
7.96
7.84
8.88
7.32
7.27
59.8
59.8
46.9
63.1
59.8
Cl
Cl
F
Cl
2m
2n
2o
2p
2.40b
1.29b
2.03b
1.62b
Cl
Cl
Cl
Cl
Cl
5.59
>10
Cl
Cl
N
F
N
0.03
0.05
N
N
F
2.00
Cl
Cl
F
2q
2r
2s
0.013
0.028
0.004
0.807
>10
2.41b
2.47b
0.38b
0.11b
1.02b
0.16b
0.05
0.41
0.48
7.30
6.67
6.54
59.8
59.8
72.7
N
N
F
Cl
Cl
Cl
0.56
N
Cl
N
N
N
N
F3C
F3C
Cl
Cl
Cl
0.64a
1.54a
3.63b
4.76b
0.17a
0.36a
0.56b
0.05b
0.28
0.24
0.15
0.01
8.12
8.15
7.88
8.01
88.1
88.1
88.1
59.8
S
S
S
2t
0.008
0.008
0.002
0.056
>10
>10
9.41
>10
Cl
2u
2v
2w
Cl
Cl
Cl
Cl
F3C
Cl
a
Sample was collected at 4 h.
b
Sample was collected at 2 h. B/P ratio: brain/plasma concentration ratio. BLOQ: Below limit of quantitation (LOQ: 5 ng/ml).
Table 3
Structurally similar analogs from both series showed comparable
CB1 potency and peripheral selectivity (i.e. 2a vs 2l, 2f vs 2o). How-
ever, in the case of the Ar1 group being 5-chloro-thiophen-2-yl, the
7-membered ring compounds (2t, 2u and 2v) are less peripherally
selective than their corresponding 6-membered ring analogs (2h,
2j and 2k). Interestingly, when the Ar2 group is 3,4-dichlorophenyl,
compounds from both series consistently exhibited good periph-
eral selectivity (2i, 2m, 2p, 2q and 2w). In addition, replacing the
pyridine of the R group with a phenyl led to compound 2n, which
possesses excellent peripheral selectivity with no detectable con-
centration of the compound in the brain. The correlation between
the compound’s cLogP and its brain/plasma ratio largely holds true
for this series. Compounds with calculated cLogP < 7 showed
higher degree of brain penetration (2r and 2s). On the other hand,
the cLogP of compound 2n is 8.88, which may explain its excep-
tional peripheral selectivity.
Efficacy of compound 2p in 3-week DIO mouse model
Group
OGTT Plasma glucose AUC (mg/dL⁄min)
2p (n = 10)
vehicle (n = 10)
22000.50 647.03a
29890.50 2516.45a
a
On day 19 from 0 to 120 min after oral glucose (2 g/kg) dosing.
group generally exhibited good peripheral selectivity (2b and 2f).
However, when R is (R)-1-(pyridine-2-yl)ethyl, good peripheral
selectivity was only observed with compounds whose Ar1 group
is 5-chloro-thiophen-2-yl (2h–j). There is a correlation between
the compound’s cLogP and its brain/plasma ratio. Compounds with
cLogP < 7 showed higher brain/plasma ratio (2c–e, 2g).
The 7-membered ring series (2l–w) generally follows the same
trend as that of the 6-membered ring series (2a–k) in terms of
structure–activity relationships (SAR) and peripheral selectivity.
In vivo efficacy of compound 2p in a 3-week diet-induced obe-
sity (DIO) mice model is shown in Table 3.13 In this chronic efficacy
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4
J. M. Matthews et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
study, groups of DIO mice were dosed orally with compound 2p
(30 mg/kg, 5 mL/kg), or the vehicle (10% PEG400 + 10% solutol,
5 mL/kg) once a day for 3 weeks. Encouragingly, the DIO mice that
were treated with compound 2p showed statistically significant
lower plasma glucose levels in an oral glucose tolerance test
(OGTT) compared to the vehicle group.14 Furthermore, a PK (phar-
macokinetic) study of compound 2p was conducted concurrent
with the efficacy study. The PK results indicated that the high
peripheral selectivity of 2p was maintained after the 22-day
repeated dosing. Two hours after the last dose, the plasma and
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12. Male C57bl/6j mice (n = 3) were dosed orally with test compounds at 20 mg/
kg. Plasma was collected via retro-orbital bleeding at 1 h and 2 h or 1 h and 4 h
after dosing. Whole brain without cerebellum was collected at 2 h or 4 h after
dosing. Wet brain weight was recorded before freezing. Brains were
homogenized in saline and sent for analysis by LC/MS to determine the
concentration of the test compound.
13. Compound 2p was formulated in 10% PEG400 and 10% solutol. DIO mice
received vehicle or compound 2p at 30 mg/kg daily. On day 19, after an
overnight fast, an oral glucose tolerance test (2 g/kg glucose) was performed.
Blood glucose was measured at 0, 30, 60 and 120 min after glucose challenge.
Blood glucose was measured from the tail vein with a Lifescan glucometer. The
dosing was continued until day 22. On day 23, the mice were euthanized, and
blood and tissues were collected.
In summary, a novel series of CB1 receptor inverse agonists has
been investigated to search for peripherally restricted compounds
that will not cross the blood–brain barrier (BBB). A large number of
compounds that exhibited potent inverse agonist activity against
CB1 receptor and good selectivity versus CB2 receptor have been
identified. The peripheral selectivity of these compounds was then
assessed in a mouse tissue distribution study, which led to the dis-
covery of several highly peripherally selective compounds, such as
2f, 2i, 2m, 2n, 2o, 2p, 2q and 2w. Furthermore, compound 2p was
evaluated in a 3-week efficacy study in DIO mice and demon-
strated beneficial effects on plasma glucose levels. These analogs
can be useful tool compounds to further investigate the therapeu-
tic potential of peripherally restricted CB1 receptor inverse
agonists.
Acknowledgments
The authors would like to thank George Ho, Tom Kirchner, and
Fuyong Du for in vivo studies, Shuyuan Zhao and Jack Kauffman for
in vitro studies, and Jason Riggs for performing the formulation
study.
14. One-way ANOVA multiple comparisons.
15. Sprague Dawley rats were dosed orally with vehicle (10% PEG400 and 10%
solutol) or compound 2p (30 mg/kg). Automated recording of stereotypic
activity started 15 min post-dose and continued for a total of 2 h.
Please cite this article in press as: Matthews, J. M.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.09.025