Discovery of a novel selective PPARγ modulator from (-)-Cercosporamide derivatives

Article abstract of DOI:10.1016/j.bmcl.2010.02.073

In an investigation of (-)-Cercosporamide derivatives with a plasma glucose-lowering effect, we found that N-benzylcarboxamide derivative 4 was a partial agonist of PPARγ. A SAR study of the substituents on carboxamide nitrogen afforded the N-(1-naphthyl)methylcarboxamide derivative 23 as the most potent selective PPARγ modulator. An X-ray crystallography study revealed that compound 23 bounded to the PPARγ ligand binding domain in a unique way without any interaction with helix12. Compound 23 displayed a potent plasma glucose-lowering effect in db/db mice without the undesirable increase in body fluid and heart weight that is typically observed when PPARγ full agonists are administrated.

Full text of DOI:10.1016/j.bmcl.2010.02.073

Bioorganic & Medicinal Chemistry Letters 20 (2010) 2095–2098
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of a novel selective PPAR
c
modulator
from (À)-Cercosporamide derivatives
a
b
c
a
Akihiro Furukawa a, , Tsuyoshi Arita , Susumu Satoh , Kenji Wakabayashi , Shinko Hayashi ,
*
Yumi Matsui a, Kazushi Araki a, Masanori Kuroha a, Jun Ohsumi a
a Shinagawa R&D Center, Daiichi Sankyo Co., Ltd, 1-2-58, Hiromachi, Shinagawa-ku, Tokyo, Japan
b Daiichi Sankyo RD Associe Co., Ltd, 3-10-2, Kitashinagawa, Shinagawa-ku, Tokyo, Japan
c Kasai R&D Center, Daiichi Sankyo Co., Ltd, 1-16-13, Kitakasai, Edogawa-ku, Tokyo, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
In an investigation of (À)-Cercosporamide derivatives with a plasma glucose-lowering effect, we found
Received 4 December 2009
Revised 15 February 2010
Accepted 17 February 2010
Available online 20 February 2010
that N-benzylcarboxamide derivative 4 was a partial agonist of PPARc. A SAR study of the substituents
on carboxamide nitrogen afforded the N-(1-naphthyl)methylcarboxamide derivative 23 as the most
potent selective PPAR modulator. An X-ray crystallography study revealed that compound 23 bounded
to the PPAR ligand binding domain in a unique way without any interaction with helix12. Compound 23
displayed a potent plasma glucose-lowering effect in db/db mice without the undesirable increase in
body fluid and heart weight that is typically observed when PPAR full agonists are administrated.
c
c
Keywords:
Cercosporamide
Diabetes
c
Ó 2010 Elsevier Ltd. All rights reserved.
PPAR
Modulator
Type 2 diabetes is one of the most serious health problems in
the world. The World Health Organization (WHO) estimates that
more than 180 million people worldwide have diabetes. This num-
ber is likely to more than double by 2030.1 Although many thera-
peutic agents have already been used in clinical situations, it is still
difficult to tightly control plasma glucose and prevent diabetic
complications.2,3 So there is great need for novel pharmacotherapy
which is able to achieve tight glycemic control singly or to be used
with existing agents.
We have previously reported that (À)-Cercosporamide had a
plasma glucose-lowering effect in KK/Ta mice, but it was accompa-
nied by a severe decrease in food intake and a loss of body weight.4
In that report it was also shown that N,O-acetal type derivatives 1
and 2 (Fig. 1) lowered the plasma glucose in KK/Ta mice without
notably affecting the food consumption or body weight. Although
they showed an obvious and promising effect on hyperglycemia,
the mechanism of action had not yet been made clear.
We designed benzylated compounds 3 and 4 to enhance the
desirable effect because the fact that compound 2 was superior
to 1 indicated that the phenyl ring near the carbamoyl group was
important for the plasma glucose-lowering effect. The preparation
of these compounds was accomplished in the following manner
(Scheme 1). For the preparation of compound 3, the carbamoyl
nitrogen and 3-hydroxyl group of (À)-Cercosporamide were re-
acted with 2,2-dimethoxypropane to form a acetonide. The carba-
myl nitrogen and 1-hydroxyl group were benzylated by
benzylbromide using sodium hydroxide as a base. The benzyl
group on the 1-hydroxy group was selectively deprotected by Pd/
C catalyzed hydrogenolysis to give compound 3. For the prepara-
tion of compound 4, the 3-hydroxyl group of (À)-Cercosporamide
was selectively methylated by iodomethane in the presence of
potassium carbonate. The carbamoyl nitrogen was benzylated to
compound 4 by reductive amination with benzaldehyde, triethylsi-
lane and trifluoroacetic acid in toluene.5
The plasma glucose-lowering effects of these compounds were
tested in hyperglycemic KK/Ta mice. The test compounds were
mixed with their diet in a ratio of 0.1% (about 100 mg/kg/day if
the food intake did not change). This mixture was fed to the mice
O
O
O
O
OH
OH
HO
HO
O
O
O
OH
NH2
(-)-Cercosporamide
R
O
N
O
H
1 : R=Me
2 : R=Ph
* Corresponding author. Tel.: +81 3 3492 3131; fax: +81 3 5436 8563.
E-mail address: furukawa.akihiro.zy@daiichisankyo.co.jp (A. Furukawa).
Figure 1. Previously reported (À)-Cercosporamide derivatives.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.02.073

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A. Furukawa et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2095–2098
Accordingly, we embarked on a SAR study to acquire a more po-
tent PPAR modulator than compound 4. First, we incorporated
various substituents on the phenyl ring. Next, we changed the phe-
nyl ring to a heteroaryl ring or a benzo-fused aryl ring. All these
compounds were synthesized in the same way as compound 4
with the corresponding aromatic aldehyde in toluene or acetoni-
O
O
c
OH
O
O
OH
OH
a, b, c
HO
O
O
O
OH
NH2
(-)-Cercosporamide
O
N
trile. We tested the PPAR
pounds. Then, for any compounds which showed potent
transcriptional activities (EC50 <1 M), we tested the potency of
c transactivation activities of these com-
O
l
3
their effect as partial antagonists. Their structures and transcrip-
tional activities are summarized in Table 2. In the various substit-
uents on the phenyl ring, only chloride substitution on the ortho-
or para-position (compounds 11 and 13) sustained transactivation
activity. In the heteroaryl or benzo-fused compounds, 2-furyl com-
d, e
O
O
OH
HO
pound 16 and 1-naphtyl compound 23 had the potent PPAR
c
transactivation activity equal to that of compound 4. As well com-
pound 23 acted as a potent partial antagonist in comparison to
compounds 4 and 16. Then we tested the binding affinities to
O
O
NH
O
4
Table 2
PPARc transactivation
Scheme 1. Reagents and conditions: (a) 2,2-dimetoxypropane, TsOH, acetone,
reflux, 8 h, 80%; (b) BnBr, NaH, DMF 0–25 °C, 2 h, 79%; (c) Pd/C, EtOAc/EtOH, 6 h,
41%; (d) MeI, K2CO3, DMF, 25 °C, 24 h, 66%; (e) benzaldehyde, Et3SiH, TFA, toluene,
reflux, 8 h, 74%.
O
O
OH
HO
O
O
Table 1
O
NH
Plasma glucose-lowering effect in KK/Ta mice
Ar
Compound
number
Plasma glucose
correctiona (%)
Body Weight on
7th dayb (%)
Food intake on
7th dayb (%)
Compound
number
Ar
Transactivation
1
3
4
56
55
69
102
103
101
78
74
100
a
b
c
d
EC50
M)
Emax
IC50
M)
Imax
(%)
(
l
(%)
(l
a
The values are the % change in the plasma glucose concentration of the drug-
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Phenyl
0.22
NDe
3.8
1.7
1.6
2.4
6.0
0.28
6.7
0.77
Negative
11
0.56
3.2
1.2
4.4
44
NDe
treated mice relative to the vehicle controls. All the values are the means of 4 or 6
mice.
o-MeO-phenyl
m-MeO-phenyl
p-MeO-phenyl
o-NO2-phenyl
m-NO2-phenyl
p-NO2-phenyl
o-Cl-phenyl
m-Cl-phenyl
p-Cl-phenyl
2-Pyridinyl
3-Pyridinyl
2-Furyl
36
43
42
34
46
19
56
50
b
The values are the % degree of the drug-treated mice relative to the vehicle
controls.
NDe
for a week. The plasma glucose, body weight, and food intake data
are summarized in Table 1. Compound 4 showed the most potent
glucose-lowering effect and did not affect the body weight or food
intake at all. On the other hand, compounds 1 and 3 slightly in-
duced the decrease in food intake.
We were strongly interested in the mechanism of action of
promising compound 4. We thoroughly investigated the interac-
tion between compound 4 and various targets which were related
to diabetes. Finally, we found that compound 4 partially activated a
peroxisome proliferator-activated receptor gamma (PPAR
0.22 M, Emax: 44%). PPAR agonists promote adipocyte differenti-
ation and improve insulin resistance.6 The PPAR
full agonists
Pioglitazone and Rosiglitazone have been clinically used and show
beneficial effects in patients with type 2 diabetes. However, the use
of these drugs has been limited because of their adverse effects,
such as weight gain, edema, and anemia.7 Recently, it was pro-
Negative
51
34
54
40
45
93
77
13
70
3-Furyl
2-Thiophenyl
3-Thiophenyl
2-Benzofuryl
2-
Benzothiophenyl
3-
Benzothiophenyl
1-Naphthyl
2-Naphthyl
2.3
3.8
c (EC50:
22
NDe
l
c
23
24
0.18
47
0.62
45
c
NDe
Pioglitazone
Rosiglitazone
0.088
0.011
118
106
Negative
Negative
a
EC50 was defined as the compound concentration at which 50% of a given
compound’s intrinsic maximal response had been reached.
posed that modulating PPAR
c activity, rather than activating it
b
The transcriptional activity in the presence of an in-house potent PPAR
c full
fully, was effective in improving insulin resistance without unde-
agonist (3.3 nM) was defined as 100%, while that in the vehicle alone was defined as
zero. The maximum transcriptional activity in the presence of the test compound
was defined as Emax (%).
IC50 was defined as the compound concentration at which 50% of a given
compound’s intrinsic maximal inhibition had been reached.
sirable adverse effects.8 A number of compounds, such as Fmoc-
L
-Leucine,9 FK614,10 5-substituted 2-benzoylaminobenzoic acids,11
c
indole and azaindole derivatives,12 T2384,13 INT-131,14 and MBX-
102,15 are reported to modulate PPAR
c
transactivation and im-
d
The maximum inhibition of the test compound against the transcriptional
activity in the presence of an in-house potent PPAR
prove insulin resistance in hyperglycemic rodent models. In some
c full agonist (3.3 nM) was
cases, it was observed that the adverse effects which accompany
PPARc activation were attenuated.
defined as Imax (%).
e
ND means ‘not determined’ because the Emax or Imax was too low.

A. Furukawa et al. / Bioorg. Med. Chem. Lett. 20 (2010) 2095–2098
2097
Table 3
PPARc. It was surprising that compound 23 showed a higher affin-
Anti-diabetic effect and adverse effects of compound 23 in db/db mice
ity (Ki: 14 nM) than Rosiglitazone (Ki: 82 nM), although the other
compounds, even compound 4, had relatively low affinities (data
not shown). Moreover, we tested the selectivity of compound 23
Compound The
ratio in
Plasma
glucose
Body
Red blood
Heart
weight
(mg)
weight gain cells (Â104
the diet (mg/dL)
for 7 days
(g)
cells/lL)
and it did not transactivate either PPARa or PPARd. We therefore
concluded that compound 23 was a potent selective PPAR
c
Vehicle
23
None
0.01%
0.1%
627 72
357 42
223 21
206 12
3.19 0.51
4.83 0.38
5.28 0.45
6.37 0.41
863 11
846 21
106.6 4.8
102.8 1.0
103.8 2.3
124.5 4.3
modulator.
In order to reveal how compound 23 interacts with PPAR
c, the
812
7
crystal structure of PPAR -LBD (ligand binding domain) with com-
c
Farglitazar 0.01%
746 19
pound 23 and a peptide derived form coactivator SRC-1 was deter-
The values are expressed as means SEM from four mice.
mined.16 The ternary complex structure showed that compound 23
bound to the PPAR
nists. However, compound 23 lacks direct interaction with helix12,
though most PPAR full agonists, including Rosiglitazone, interact
with Tyr473 in helix12 to stabilize PPAR in the active conforma-
c ligand binding site similar to other PPARc ago-
Finally, we tested whether the novel selective PPAR
c
modulator
c
23 showed an anti-diabetic effect and attenuated adverse effects in
c
comparison with the PPAR
c
full agonist Farglitazar (GI262570).18
tion (Fig. 2, A).17 The Cercosporamide scaffold of compound 23 is
located between helix3 and b-sheet, while the 1-naphthyl group
is embedded in a hydrophobic region which consisted of helix3,
helix5, b-strand2 and helix7. The Cercosporamide scaffold also
makes water-mediated hydrogen bonds with Leu340 and Ser342
(Fig. 2, B). Together with previous works11,13,14 which reported
The test compounds were mixed with the diet in a certain ratio.
The mixture was fed to hyperglycemic male db/db mice19 for a
week (Table 3). Compound 23 lowered the plasma glucose levels
in a dose-dependent manner and a high dose (0.1%) was as effec-
tive as Farglitazar (0.01%). Compound 23 also increased the body
weight in a dose-dependent manner, but the increases in body
weight tended to be less than the increase by Farglitazar. In the
group treated with Farglitazar, it was also observed that the num-
ber of red blood cells decreased and the heart weight increased.
The decrease in red blood cells indicated the fluid retention which
is associated with edema and anemia. As well, cardiac hypertrophy
was considered to be compensation for this fluid retention. On the
other hand, in the groups treated with compound 23, the tendency
of fluid retention was slightly observed and cardiac hypertrophy
was not recognized at all.
PPARc partial agonists bound with PPARc without direct interac-
tion to residues in helix12, the lack of direct interaction of com-
pound 23 with helix12 might also be responsible for its partial
agonist and partial antagonist activities.
In conclusion, we succeeded in acquiring a potent selective
PPAR
lography study revealed unique interaction between compound
23 and PPAR -LBD. Furthermore, compound 23 showed a potent
anti-diabetic effect in db/db mice. In comparison with the PPAR
c
modulator 23 from (À)-Cercosporamide. An X-ray crystal-
c
c
full agonist Farglitazar, the adverse effects such as body weight
gain and fluid retention were attenuated and no increase in heart
weight was observed at all. Further investigations to enhance these
desirable profiles are ongoing and the results will be reported
elsewhere.
Acknowledgments
We would like to thank the staff at the Photon Factory for their
excellent support in the use of the synchrotron beam lines. We
would also like to thank Mr. Takashi Suzuki and his co-workers
at Process Technology Research Laboratories, Daiichi Sankyo Co.
Ltd, for providing the purified (À)-Cercosporamide.
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