Aryl urea derivatives of spiropiperidines as NPY Y5 receptor antagonists

Article abstract of DOI:10.1016/j.bmcl.2009.05.013

Continuing medicinal chemistry studies to identify spiropiperidine-derived NPY Y5 receptor antagonists are described. Aryl urea derivatives of a variety of spiropiperidines were tested for their NPY Y5 receptor binding affinities. Of the spiropiperidines

Full text of DOI:10.1016/j.bmcl.2009.05.013

Bioorganic & Medicinal Chemistry Letters 19 (2009) 3511–3516
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Aryl urea derivatives of spiropiperidines as NPY Y5 receptor antagonists
Toshiyuki Takahashi, Yuji Haga, Toshihiro Sakamoto, Minoru Moriya, Osamu Okamoto,
Katsumasa Nonoshita, Takunobu Shibata, Takuya Suga, Hirobumi Takahashi, Tomoko Hirohashi,
Aya Sakuraba, Akira Gomori, Hisashi Iwaasa, Tomoyuki Ohe, Akane Ishihara, Yasuyuki Ishii,
*
Akio Kanatani, Takehiro Fukami
Tsukuba Research Institute, Banyu Pharmaceutical Co., Ltd, Okubo 3, Tsukuba 300-2611, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Continuing medicinal chemistry studies to identify spiropiperidine-derived NPY Y5 receptor antagonists
are described. Aryl urea derivatives of a variety of spiropiperidines were tested for their NPY Y5 receptor
binding affinities. Of the spiropiperidines so far examined, spiro[3-oxoisobenzofurane-1(3H),40-piperi-
dine] was a useful scaffold for producing orally active NPY Y5 receptor antagonists. Oral administration
of 5c significantly inhibited the Y5 agonist-induced food intake in rats with a minimum effective dose of
3 mg/kg. In addition, this compound was efficacious in decreasing body weight in diet-induced obese
mice.
Received 23 March 2009
Revised 1 May 2009
Accepted 2 May 2009
Available online 9 May 2009
Keywords:
Neuropeptide Y
NPY
Ó 2009 Elsevier Ltd. All rights reserved.
Antagonist
Obesity
Neuropeptide Y (NPY) is a highly conserved C-terminus ami-
dated peptide consisting of 36 amino acid residues which has
been shown to have potent, centrally-mediated orexigenic ef-
fects.1–4 NPY is abundant in the central nervous system through-
out the cerebral cortex, forebrain, hypothalamus, brain stem and
spinal cord. In the periphery, NPY is present in most sympathetic
nerve fibers, especially around blood vessels. Reports of NPY
activity demonstrate a wide range of potential effects at both cen-
tral and peripheral targets, acting either alone or in combination
with other neurotransmitters such as norepinephrine and gluta-
mate. The effects of the NPY family of peptides are mediated
via a family of GPCRs, providing opportunities for subtype selec-
tive therapeutics. At least six receptor subtypes of the NPY family
have been characterized based on cloning and/or their pharmaco-
logical properties.5–19 Various pharmacological studies, employing
receptor deficient mice and/or subtype-selective agonists and
antagonists, have suggested that the Y1 and Y5 receptors are in-
volved in body weight regulation.14,16,20–32 Thus, the antagonism
of the Y1 and/or Y5 receptors may have considerable therapeutic
benefits for the treatment of obesity.
in a Y5 agonist-induced food intake model in rats, 1b was poorly
brain penetrant. We were intrigued by these results as the spiro-
indoline-3,40-piperidine structure is a so-called ‘privileged struc-
ture’.34–38 It is known that certain ‘privileged structures’ are
capable of providing useful ligands for more than one receptor
and that judicious modification of such structures could be a viable
alternative in the search for new receptor agonists and antago-
nists.39 We assumed that replacement of the spiroindoline-3,40-
piperidine with other spiropiperidine-based privileged structures
may yield potent NPY Y5 receptor antagonists. In addition, some
of them could possess more desirable profiles than the spiroindo-
line-3,40-piperidine Y5 antagonists from the viewpoint of clinical
development candidates, such as improvements in pharmacokinet-
ics, brain penetration and safety. We herein report structure–activ-
ity relationships of spiropiperidine–urea derivatives NPY Y5
receptor antagonists and identification of potent spiro[3-
oxoisobenzofurane-1(3H),40-piperidine]–urea
derivatives
Y5
receptor antagonists.
The spiroindoline phenylpyrazine urea derivative 1c was as
potent and as brain penetrant as 1a (Fig. 1). We employed a
2-amino-5-phenylpyrazine structure as a surrogate of the biphe-
nylamine moiety in 1a for initial exploration of the structure–
activity studies at the spiropiperidine moiety due to safety con-
cerns.40 2-Amino-3-phenylpyrazine 2 was reacted with phenyl
chloroformate in pyridine to afford phenyl carbamate 3, which
was reacted with spiropiperidines 4a–h to give the correspond-
ing urea derivatives of spiropiperidines 5a–h. Two conditions
were employed for the urea formation reaction; (a) the piperi-
In our preceding Letter,33 we reported discovery of potent and
orally active NPY Y5 receptor antagonists based on a spiroindo-
line-3,40-piperidine scaffold, exemplified by 1a and 1b. Compound
1a was highly potent and well brain penetrant, but lacked oral bio-
availability. In contrast, while 1b was orally bioavailable and active
* Corresponding author. Tel.: +81 29 877 2361; fax: +81 29 877 2029.
E-mail address: takehiro_fukami@merck.com (T. Fukami).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.05.013

3512
T. Takahashi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3511–3516
N
N
O
O
O
N
N
N
N
N
H
N
H
N
H
N
N
N
O
O
O
1a
hY5 IC50: 0.85 nM
rat brain/plasma ratioa: 1.30
1b
1c
hY5 IC50: 0.97 nM
rat brain plasma ratioa: 1.14
S
S
S
CH3
CH3
hY5 IC50: 4.1 nM
CH3
O
O
O
rat brain/plasma ratioa: 0.12
Figure 1. Spiroindoline-3,40-piperidine Y5 leads. At 10 min after 3 mg/kg iv. n = 2.
dine derivative and the phenyl carbamate were refluxed in
CHCl3 in the presence of Et3N or (b) the piperidine derivative
and the phenyl carbamate were reacted in DMSO in the pres-
ence of 10 N aqueous NaOH at room temperature, and the reac-
tion condition was chosen in accordance with solubility of the
substrates (Scheme 1). The general synthetic method was ap-
plied for replacement of the 4-phenylpyrazine group with vari-
ous groups.
Compounds 5a–h were evaluated for their Y5 binding affini-
ties.41 Compounds 5b, 5c, 5e and 5g showed potent Y5 receptor
binding affinities (although other analogs were very weak) sug-
gesting that privileged structures are applicable to produce potent
Y5 antagonists. The potent compounds 5b, 5c and 5e were tested
for their oral bioavailability in rats. Although 5g showed a potent
Y5 binding affinity, this compound exhibited unacceptable CYP3A
pre-incubation time dependent inhibition,42 and we did not fur-
ther investigate this scaffold. Of the three compounds tested for
their oral bioavailability, 5c showed the best oral bioavailability
(49%). Compound 5c also exhibited a high brain level as well as
a high cerebrospinal fluid (CSF) level at 2 h after oral administra-
tion in rat (Table 1). Compound 5c showed potent binding affin-
ities to rat and mouse Y5 receptors43 (IC50 values of 1.7 nM and
1.7 nM, respectively) while the compound was very weak at hu-
As the spiro[3-oxoisobenzofurane-1(3H),40-piperidine] was
identified as a particularly useful scaffold for potent and selec-
tive Y5 antagonists, we further explored further modifications
of this scaffold. Although compound 5c was in vivo active,
its solubility in aqueous media was limited (1.9
lg/mL and
1.2
l
g/mL in JP1 at pH 1.2 and JP2 at pH 6.8, respectively).46
More soluble compounds may be more preferable in terms of
oral absorption and developability, so the 4-phenylpyrazinyl
group in 5c was replaced with various biaryl groups to inves-
tigate structure-activity relationships at this moiety. While var-
ious biaryl groups were tolerable in terms of the Y5 binding
affinity although most compounds showed limited solubility,
only the 4-phenylpyrimidine analog 6a was particularly inter-
esting for its good solubility. Introduction of substituents, such
as Cl, CH3O, F and CF3, on the outer phenyl ring in 6a was
examined. In all cases, introduction of substituents at the
para-position decreased the Y5 binding affinity whereas those
at the ortho or meta-positions produced similar or modestly in-
creased Y5 binding affinity. The phenylisoxazole analog 9a was
another interesting compound in terms of good Y5 binding
affinity, and introduction of Cl on the outer phenyl ring was
examined. In this case, the p-Cl substituent was not detrimen-
tal, and o-Cl, m-Cl and p-Cl analogs 9b–d were as potent as 9a
unlike the prior case of 4-phenylpyrimidine analogs. (Table 2)
Of the 4-phenylisoxazole analogs 9a–d, 9d was orally available
in rats while compounds 9a–c were very poorly available pre-
sumably due to their insufficient metabolic stability (data not
shown).
man Y1, Y2 and Y4 receptors (IC50 > 10 lM). The antagonistic
activity of 5c was assessed by its ability to inhibit NPY-induced
[Ca2+]i increases in LMtk-cells expressing the recombinant human
Y5 receptor.43 In this Y5 functional assay, 5c inhibited the [Ca2+
]
i
increase with IC50 of 6.1 nM, demonstrating that 5c is a Y5-selec-
tive antagonist. Oral administration of 5c strongly inhibited the
Compounds 6a and 9d were tested for their inhibitory effects
Y5 agonist,
D
-Trp34NPY-induced food intake44 in rats with a min-
on D
-Trp34NPY-induced food intake in rats after 10 mg/kg po dos-
imum effective dose of 3 mg/kg. In contrast, 5c did not signifi-
ing; plasma, brain and CSF exposures were also determined. The
exposure data at 2 h after 10 mg/kg oral administration together
with rat Y5 receptor binding affinities are listed in Table 3. Com-
pounds 5c, 6a and 9d showed equally potent binding affinities to
the rat Y5 receptor as well as to the human Y5 receptor. In the
cantly inhibit NPY-induced food intake even at 10 mg/kg,
demonstrating that the inhibitory effect of 5c in D
-Trp34NPY-in-
duced food intake was due to the Y5 receptor-selective antago-
nism (Fig. 2).43,45
NH
X
4a-h
O
O
Y
H2N
NH
NH
Et3N/CHCl3
N
N
N
O
N
PhOCOCl/Pyridine
or 10N NaOH aq./DMSO
N
N
N
X
Y
2
3
5a-h
4a, 5a: X = N-CH3, Y = C(O)
4b, 5b: X = NH, Y = (CO)
4c, 5c: X = O, Y = C(O)
4d, 5d: X = O, Y = CH2
4e, 5e: X = NH, Y = CH2C(O)
4f, 5f: X = CH2NH, Y = (CO)
4g, 5g: X = O, Y = CH2C(O)
4h, 5h: X = O, Y = CH2CH2
Scheme 1. Synthesis of urea derivatives of spiropiperidines.

T. Takahashi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3511–3516
3513
Table 1
Human Y5 receptor binding affinities, bioavailability, and plasma, brain and CSF exposure data in rats
Rat Fb
Brain, plasma and CSF levels at 2 h after 10 mpk po in ratsc
Plasma level ( M) Brain level, nmol/g tissue CSF level (lM)
a
Compounds
Structure
hY5 binding IC50 (nM)
l
O
NH
N
N
N
N
5a
>1000d
NT
NT
NT
NT
N
N
CH3
O
O
NH
N
5b
5c
5d
5e
5f
3.7
8%
0.36
2.98
NT
0.029
2.92
NT
ND
0.10
NT
NT
NT
NT
NT
N
NH
O
O
N
NH
2.4
49%
NT
3%
N
O
O
O
NH
N
N
N
>1000d
N
O
O
NH
N
1.7
NT
NT
N
NH
O
O
NH
N
N
N
>1000d
NT
NT
NT
NT
NT
N
NH
O
O
NH
N
5g
5.7
NT
NT
N
O
O
O
NH
N
N
5h
>1000d
NT
NT
N
O
a
In vitro data in nM are the average of at least two experiments.
3 mg/kg iv (n = 2) vs 10 mg/kg po (n = 2).
n = 3.
b
c
d
n = 1. NT, not tested. ND, not detected.

3514
T. Takahashi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3511–3516
Table 2 (continued)
4
3
2
1
0
6
Compounds Ar
hY5 binding
IC50 (nM)
Solubility (lg/mL)
5
a
JP1 at
JP2 at
4
3
pH 1.2
pH 6.8
CF3
*
*
N
2
1
0
6k
6l
1.3
0.85
20
0.8
4.4
NT
0.1
0.3
NT
N
N
CF3
vehicle
1
3
10
vehicle Comp. 5c
(10 mg/kg)
Compound 5c (mg/kg)
D-Trp34NPY 1 µg
N
N
hNPY 0.3 µg
6m
7
CF3
Figure 2. Inhibitory effect of compound 5c on D
-Trp34NPY—or NPY-induced food
N
N
intake in rats. *P < 0.05 versus vehicle treated group. n = 13–16.
N
N
N
2.2
0.6
NT
0.2
NT
Table 2
Human Y5 binding affinities and solubility data
8
6.1
O
H
N
N
O
9a
0.84
0.1
0.2
N
Cl
O
N
N
O
O
9b
9c
9d
0.83
1.0
NT
NT
O
Compounds Ar
hY5 binding
IC50 (nM)
Solubility (lg/mL)
Cl
a
JP1 at
JP2 at
NT
NT
pH 1.2
pH 6.8
N
N
N
N
O
5c
6a
6b
2.4
1.4
1.1
1.9
>100
2.6
1.2
85
0.69
<0.1
<0.1
N
Cl
HN
N
N
10
11
1.9
2.9
2.5
27
17
23
Cl
0.2
N
N
N
Cl
N
6c
6d
6e
6f
0.91
3.9
7.1
NT
3.1
NT
NT
5.9
43
0.7
NT
N
N
N
N
12
13
2.3
3.5
0.8
0.7
4.3
Cl
N
N
N
OCH3
N
>100
1.1
0.1
NT
N
N
OCH3
N
S
N
N
14
15
16
17
1.8
7.9
1.2
3.4
0.4
NT
<0.1
NT
1.8
S
N
N
N
6g
6h
6i
14
NT
OCH3
N
N
F
S
0.99
1.1
0.6
4.5
4.0
0.6
NT
0.5
NT
N
N
N
F
N
N
N
S
6j
2.4
48
F
NT, not tested.
a
N
In vitro data in nM are the average of at least two experiments.

T. Takahashi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3511–3516
3515
Table 3
Rat Y5 receptor binding affinities, inhibitory effect in D-Trp34NPY-induced food intake, and plasma, brain and CSF exposure data in rats
Minimum effective dose (mg/kg po)b
Brain, plasma and CSF levels at 2 h after 10 mpk po in ratsc
a
Compounds
rY5 agbinding IC50 (nM)
Plasma level (lM)
Brain level, nmol/g tissue
CSF level (lM)
5c
6a
9d
1.7
0.67
1.3
3
3
>30
2.98
3.86
1.42
2.92
0.26
1.09
0.10
0.13
<0.01
a
In vitro data in nM are the average of at least two experiments.
b
c
Inhibitory effect in D
-Trp34NPY-induced food intake.
n = 3.
Acknowledgements
We thank Dr. Arthur A. Patchett, Dr. Lex H. T. Van der Ploeg and
Dr. Douglas J. MacNeil for their suggestions and discussion on the
privileged structures and the NPY Y5 receptor antagonist program.
We are grateful to Dr. Peter T. Meinke, Merck Research Laborato-
ries, for his critical reading of the manuscript.
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In summary, we identified a series of spiro[3-oxoisobenzofura-
ne-1(3H),40-piperidine]–urea derivatives that are potent, selective
and orally active NPY Y5 receptor antagonists. In particular, 5c
significantly inhibited
with a minimum effective dose of 3 mg/kg. The CSF exposure
level was useful indicator to predict efficacy in the rat
-Trp34NPY-induced food intake model. Compound 5c showed
D
-Trp34NPY-induced food intake in rats
a
D
sustained brain Y5 receptor occupancy in mice, and oral once-dai-
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40. According to the Merck Index, 14th edition, potential symptoms of
overexposure are headache, dizziness; lethargy, dyspnea; ataxia, weakness;
methemoglobinemia; urinary burning; acute hemorrhagic cystitis. See NIOSH
Pocket Guide to Chemical Hazards (DHHS/NIOSH 97-140, 1997) p 14. This
substance is listed as a known human carcinogen: Report on Carcinogens,
Eleventh Edition (PB2005-104914, 2004) p III-13.
41. [
125I]PYY binding to human recombinant Y5 receptors in LMtk- cells; see Ref.
43.
42. Compound 5g at 10
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