Design and synthesis of 6-fluoro-2-naphthyl derivatives as novel CCR3 antagonists with reduced CYP2D6 inhibition

Article abstract of DOI:10.1016/j.bmc.2008.08.006

In our previous study on discovering novel types of CCR3 antagonists, we found a fluoronaphthalene derivative (1) that exhibited potent CCR3 inhibitory activity with an IC₅₀ value of 20 nM. However, compound 1 also inhibited human cytochrome P450 2D6 (CYP2D6) with an IC₅₀ value of 400 nM. In order to reduce its CYP2D6 inhibitory activity, we performed further systematic structural modifications on 1. In particular, we focused on reducing the number of lipophilic moieties in the biphenyl part of 1, using C log D_7.4 values as the reference index of lipophilicity. This research led to the identification of N-{(3-exo)-8-[(6-fluoro-2-naphthyl)methyl]-8-azabicyclo[3.2.1]oct-3-yl}-3-(piperidin-1-ylcarbonyl)isonicotinamide 1-oxide (30) which showed comparable CCR3 inhibitory activity (IC₅₀ = 23 nM) with much reduced CYP2D6 inhibitory activity (IC₅₀ = 29,000 nM) compared with 1.

Full text of DOI:10.1016/j.bmc.2008.08.006

Bioorganic & Medicinal Chemistry 16 (2008) 8607–8618
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design and synthesis of 6-fluoro-2-naphthyl derivatives as novel CCR3
antagonists with reduced CYP2D6 inhibition
a
a
a
a
a
Ippei Sato a, , Koichiro Morihira , Hiroshi Inami , Hirokazu Kubota , Tatsuaki Morokata , Keiko Suzuki ,
Yosuke Iura b, Aiko Nitta b, Takayuki Imaoka b, Toshiya Takahashi b, Makoto Takeuchi a,
Mitsuaki Ohta a, Shin-ichi Tsukamoto a
*
a Drug Discovery Research, Astellas Pharma. Inc., 21 Miyukigaoka, Tsukuba-shi, Ibaraki 305-8585, Japan
b Pharmaceutical Research Laboratories, Toray Industries, Inc., 6-10-1 Tebiro, Kamakura, Kanagawa 248-0036, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
In our previous study on discovering novel types of CCR3 antagonists, we found a fluoronaphthalene
derivative (1) that exhibited potent CCR3 inhibitory activity with an IC50 value of 20 nM. However, com-
pound 1 also inhibited human cytochrome P450 2D6 (CYP2D6) with an IC50 value of 400 nM. In order to
reduce its CYP2D6 inhibitory activity, we performed further systematic structural modifications on 1. In
particular, we focused on reducing the number of lipophilic moieties in the biphenyl part of 1, using
ClogD7.4 values as the reference index of lipophilicity. This research led to the identification of N-{(3-
e x o )-8-[(6-fluoro-2-naphthyl)methyl]-8-azabicyclo[3.2.1]oct-3-yl}-3-(piperidin-1-ylcarbonyl)isonicoti-
namide 1-oxide (30) which showed comparable CCR3 inhibitory activity (IC50 = 23 nM) with much
reduced CYP2D6 inhibitory activity (IC50 = 29,000 nM) compared with 1.
Received 27 June 2008
Revised 1 August 2008
Accepted 2 August 2008
Available online 7 August 2008
Keywords:
Allergic diseases
CCR3 antagonists
CYP2D6 inhibition
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
Thus, novel type CCR3 antagonists may have potential as ther-
apeutic agents for eosinophil-related allergic diseases, such as
Infiltration of eosinophils into the airways and the mucosal tis-
sues is one of the pathological features of various inflammatory
diseases, such as allergic asthma, allergic rhinitis, and atopic der-
matitis. In particular, asthma is a chronic inflammatory disease
accompanied by airway obstruction and hyper-responsiveness
caused by cytotoxic proteins, chemical mediators, and inflamma-
tory mediators released from activated eosinophils.1a,1b Therefore,
eosinophils are thought to play a critical role in the induction and
progression of these allergic diseases.
Chemokines are a family of small (8–12 kDa), heparin-binding,
basic proteins that induce the migration and activation of leuko-
cytes, such as monocytes, lymphocytes, eosinophils, and basophils.
They are classified into four sub-families, CC chemokines, CXC che-
mokines, C chemokines, and CX3C chemokines, based on the posi-
tions of the first two of their conserved N-terminal cysteine
residues. CCR3, which is one of the CC chemokine receptor, is
prominently expressed on eosinophils, and is responsible for eosin-
ophil chemotaxis.2a–e The specific antibody against eotaxin-1, a
selective CCR3 ligand, reduces the lung eosinophilia and airway
hyper-responsiveness in mice that occurs in response to
ovalbumin.3
allergic asthma, allergic rhinitis, and atopic dermatitis.
In our previous research aimed at discovery of a novel type of
CCR3 antagonist, a 6-fluoro-2-naphthyl derivative, compound 1,
was found to have potent CCR3 inhibitory activity with an IC50 va-
lue of 20 nM. However, further biological studies revealed that 1
also had potent inhibitory activity against human cytochrome
P450 2D6 (CYP2D6), which is a polymorphic member of the P450
superfamily (IC50: 400 nM), but weak inhibitory activity against
other CYP enzymes, such as CYP1A2, 2C9, 2C19, and 3A4
(IC50 > 20,000 nM) (Fig. 1). Since CYP2D6 is important to the
metabolism of about 20% of clinically used drugs, its inhibition
can cause unfavorable drug–drug metabolizing interaction.4 There-
fore, a program to further reduce the CYP2D6 inhibitory activity of
1 was designed as described below.
Recently, we reported that both the 6-fluoro-2-naphthyl moiety
and the nortropane ring of 1 were essential for potent inhibitory
activity against CCR3.5 Therefore, we attempted to modify the
biphenyl moiety of compound 1 by dividing into Part A and Part
B (Fig. 1). This study focused on compound lipophilicity since some
groups have already reported that it is an important contributor to
CYP inhibition.6a–c Reduction of the molecular lipophilicity was
speculated to be an effective way to reduce inhibitory activity
against CYP2D6, despite the fact that it is preferable to have bulky
and/or lipophilic moieties on Part A of 1.5 Therefore, in order to
* Corresponding author. Tel.: +81 29 863 6732; fax: +81 29 852 5387.
E-mail address: ippei.sato@jp.astellas.com (I. Sato).
0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.08.006

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I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
(9a–c, 12a, 12b, 15, and 17) carboxylic acids afforded compounds
6a–i.
Scheme 2 summarizes the synthesis of the carboxylic acids 9a–
Part A
c, 12a, 12b, 15, and 17. Compounds 9a–c were obtained by Suzuki
coupling of 2-methylphenyl boronic acid (7) and appropriate
bromopyridines, followed by oxidation of the methyl moiety using
KMnO4.10 The carboxylic acids 12a–b were prepared from 2-form-
ylphenylboronic acid (10) and appropriate arylbromides by Suzuki
coupling, followed by oxidation of the formyl group. The Mitsun-
obu reaction was performed using methyl salicylate (13) and
cyclohexanol to yield the ester 14,11 and subsequent hydrolysis
of the ester group afforded the carboxylic acid 15. The carboxylic
acid 17 was obtained from 2-fluorobenzonitrile (16) via a substitu-
tion reaction with a piperidine, followed by hydrolysis of the nitrile
group.
O
N
N
H
F
Part B
1
inhibitory activity
CCR3:IC50 = 20 nM
CYP2D6: IC50 = 400 nM
CYP1A2: IC50 > 20,000 nM
CYP2C9: IC50 > 20,000 nM
CYP2C19: IC50 > 20,000 nM
CYP3A4: IC50 > 20,000 nM
Compounds 20a–g were synthesized by condensation of the
amine 5 with appropriate carboxylic acids (19a–g) which were
prepared from phthalic anhydride (18) and the corresponding
amines (Scheme 3).
Figure 1. The structure of compound 1.
Scheme 4 shows the syntheses of several carboxylic acids (23,
26, 29, 27, and 35) required for generation of the corresponding
compounds (24, 28, 30, 32, and 36). The carboxylic acid 23, which
corresponded to compound 24, was synthesized according to the
method reported by Zhenrong Guo et al., as described below.12 Di-
methyl pyridine-2,3-dicarboxylate (21) was regioselectively con-
verted to the amide (22) by treatment with piperidine in the
presence of MgCl2. The amide (22) was then hydrolyzed to yield
compound 23. The carboxylic acid 26 was prepared from pyri-
dine-3,4-dicarboxylic acid (25) in three steps. Compound 25 was
treated with acetic anhydride to yield pyridine-3,4-dicarboxylic
anhydride, and then a ring-opening reaction by piperidine afforded
the 1:1 mixture of carboxylic acids 26 and 27. Compound 27 could
be easily removed by recrystallization from EtOH–EtOAc to yield
the pure carboxylic acid 26. The carboxylic acid 29 was synthesized
by oxidation of the 1:1 mixture of above-obtained carboxylic acids
26 and 27, followed by purification via silica gel column chroma-
tography. Pure Compound 27 was obtained by the regioselective
ring-opening reaction of pyridine-3,4-dicarboxylic anhydride with
MeOH,13 followed by direct conversion of the ester group to the
amide group by piperidine under neat conditions. The condensa-
tion of 3314 with piperidine using WSC as the coupling reagent
yielded the amide 34, and subsequent hydrolysis of 34 yielded
the carboxylic acid 35. Condensation of the carboxylic acids 23,
26, 29, 27, and 35 with the amine 5 yielded compound 24, 28,
30, 32, and 36, respectively.
obtain less lipophilic compounds with potent CCR3 inhibitory
activity, attempts were made to replace the phenyl ring with other
heteroaromatic rings or non-aromatic heterocyclic rings and opti-
mize their linkages. The n-octanol/water distribution coefficient of
designed compounds calculated at pH 7.4 (ClogD7.4) was used as
an index of lipophilicity. The ClogD7.4 values were calculated using
ACD/logD software, and the lead compound 1 was determined to
have a ClogD7.4 value of 3.79.7 Thus, the attenuation of CYP2D6
inhibitory activity was assumed to be achieved by designing and
synthesizing derivatives with ClogD7.4 values lower than those
of 1.
In this paper, we describe the successful development of a novel
type of CCR3 antagonist with reduced CYP2D6 inhibitory activity.
2. Chemistry
The synthetic routes of compounds 6a–i are shown in Scheme
1. The protection of amine 28 with the Boc group and subsequent
deprotection of the benzyl group yielded the amine 3. Alkylation
of 3 with 2-(bromomethyl)-6-fluoronaphthalene,9 followed by re-
moval of the Boc group by creating an acidic condition yielded
the key intermediate, amine 5. Condensation of 5 with a commer-
cially available (in the cases of 6f and 6g) or synthesized
a,b
O
c
d
O
H2N
N
O
N
NH
HCl
O
N
N
H
H
F
2
3
4
2HCl
H2N
e
R
O
N
N
N
H
F
F
5
6a-i
N
N
N
N
N
R =
N
S
N
O
*
N
*
N
*
N
*
*
*
*
*
*
6a
6b
6c
6d
6e
6f
6g
6h
6i
Scheme 1. Synthesis of compounds 6a–i. Reagents and conditions: (a) Boc2O, THF, rt, 11 h; (b) H2, 10 wt% Pd(OH)2/C, 4 M HCl/EtOAc, EtOH–MeOH, rt, 3 h, 96% for two steps;
(c) 2-(bromomethyl)-6-fluoronaphthalene,9 K2CO3, DMF, rt, 21 h, quant.; (e) ArCOOH, WSC, HOBt, Et3N, CH2Cl2, rt.

I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
8609
HO OH
B
N
*
Ar
Ar
a
b
N
COOH
N
Ar =
Ar =
*
*
c
7
8a-c
9a-c
a
b
B(OH)2
Ar
Ar
N
N
c
d
N
S
CHO
CHO
COOH
*
*
10
11a-b
12a-b
a
b
OH O
f
F
e
g, h
O
O
O
O
O
CN
N
O
O
OH
OH
16
17
13
14
15
Scheme 2. Synthesis of compounds 9a–c, 12a–b, 15, and 17. Reagents and conditions: (a) bromopyridine, Pd(PPh3)4, CsF, DME, reflux; (b) KMnO4, H2O, reflux; (c) ArBr,
Pd(PPh3)4, Na2CO3, DME–H2O, reflux; (d) NaClO2, 31% H2O2 aq, NaH2PO4, H2O–MeCN; (e) cyclohexanol, PPh3, DEAD, THF, rt, 4 d, 57%; (f) 1 M NaOH, MeOH, rt, 3.5 d, quant.; (g)
piperidine, K2CO3, DMF, 70 °C, over night; (h) KOH, 2-ethoxyethanol–H2O, reflux, 4 h, 58% for two steps.
O
R
O
R
O
O
O
b
O
a
O
N
N
OH
H
F
18
19a-g
20a-g
N
N
O
R =
N
*
N
*
N
N
N
N
*
*
*
*
*
a
b
c
d
e
f
g
Scheme 3. Synthesis of compounds 20a–g. Reagents and conditions: (a) amine, THF, rt; (b) 5, WSC, HOBt, Et3N, CH2Cl2, rt.
compound 6e also showed reduced CYP2D6 inhibitory activity, as
3. Results and discussion
indicated by the reduction in ClogD7.4 values.
Next, the effect induced by replacing Part A of 1 with other het-
ero non-aromatic rings connected by hetero-atoms-containing
linkages was investigated (Tables 2 and 3). When Part A was re-
placed with a cyclohexyl ether moiety (6h), a decrease in CCR3
inhibitory activity of about sixfold occurred, and the activity of
the cyclohexyl amide derivative (20a) was moderate
(IC50 = 44 nM). The same tendency was observed in the pair of
the piperidine derivative (6i) and the piperidinocarbonyl deriva-
tive (20b). These results indicated that the carbonyl group of Part
A could play a significant role in the interaction with the CCR3
receptor. Thus, some potent inhibitors with hetero non-aromatic
rings at Part A of 1 were found by introducing a carbonyl group
there. With respect to CYP2D6 inhibition, 6h, which had a similar
ClogD7.4 value to that of 1, showed comparable activity to 1.
And, 20a and 20b, which both had lower ClogD7.4 values than 1,
showed weaker activity (IC50 = 2900 and 6500 nM, respectively).
These results therefore prompted us to investigate the effects of
replacing the piperidine ring in 20b with other cyclic amines. The
piperazine and morpholine derivatives (20c and 20d), which had
lower ClogD7.4 values than that of 20b, showed significantly less
potent CCR3 inhibitory activity, although their CYP2D6 inhibitory
activity was similar to that of 20b. These results suggest that the
presence of hetero-atoms in this position would not be suitable
for CCR3 inhibitory activity, but they would be effective for the
attenuation of CYP2D6 inhibition. Among compounds 20e, 20f,
and 20g, in which the piperidine ring in 20b was converted to 5-,
The synthesized compounds were evaluated for their CCR3
inhibitory activity (IC50) on eotaxin-induced Ca2+ influx using
CCR3-expressing preB cells. The lead compound (1) displayed po-
tent CCR3 inhibitory activity with an IC50 value of 20 nM. The
CYP2D6 inhibitory activity (IC50) of the selected compounds was
also evaluated according to the reported procedure.15 The lead
compound (1) displayed potent CYP2D6 inhibitory activity with
an IC50 value of 400 nM. The ClogD7.4 values were calculated using
the ACD/logD software, and that for the lead compound (1) was
determined to be 3.79.7
First, the effects of replacing the Part A of 1 with other aromatic
rings were explored (Table 1). While the CCR3 inhibitory activities
of both the 4-pyridyl derivative (6a) and the 2-pyridyl derivative
(6c) were about 30-fold less and threefold less, respectively, than
that of 1, the activity of the 3-pyridyl derivative (6b) was similar
to that of 1 (IC50 = 29 nM). However, the pyrimidyl derivative
(6d) exhibited about fivefold less activity than that of 1. It seemed
that only one nitrogen atom may be tolerated at the 3-positon of
Part A of 1 since other substituted patterns were not. As we ex-
pected, compounds 6b, 6c, and 6d showed reduced inhibitory
activity against CYP2D6 (IC50 = 1600, 4000, and 2900 nM, respec-
tively) probably due to the fall in their ClogD7.4 values. The CCR3
inhibitory activity of all 5-membered aromatic ring derivatives
(6e, 6f, and 6g) was lower than that of 1. It seemed that, in terms
of CCR3 inhibitory activity, Part A of 1 may prefer a larger molecule
than 5-membered aromatic ring system. Similar to the above,

8610
I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
O
N
O
a
b
c
O
N
N
O
N
N
O
O
N
N
O
O
O
O
O
N
OH
N
H
F
21
22
23
24
HO
N
O
N
O
O
N
O
f
O
d, e
N
O
O
O
OH
OH
OH
OH
N
N
N
25
mixture (26: 27 = 1 : 1)
26
c
N
N
O
O
N
g
N
H
F
28
c
N
O
O
N
O
O
N
H
OH
N
N+
N+
-O
c
-O
O
F
29
30
d, h
i
HO
N
O
O
O
O
N
O
N
O
O
O
OH
OH
N
OH
N
H
N
N
N
N
F
25
31
27
32
O
N
HO
j
k
c
O
N
O
N
O
N
O
N
O
O
O
O
N
H
O
OH
N
N
F
33
34
35
36
Scheme 4. Synthesis of compounds 24, 28, 30, 32, and 36. Reagents and conditions: (a) piperidine, MgCl2, THF, rt, 1 d, 37%; (b) 1 M NaOH, EtOH, rt, over night, quant.; (c) 5,
WSC, HOBt, Et3N; (d) Ac2O, reflux; (e) piperidine, THF, rt, 17 h; (f) recrystallization from EtOAc–EtOH, 23% from 25; (g) mCPBA, CH2Cl2, rt, 21 h, 34% from 1:1 mixture of 26 and
27; (h) MeOH, reflux, 4.5 h, 45%; (i) piperidine, neat, 70 °C, 1 d, 58%; (j) piperidine, WSCꢁHCl, HOBt, CH2Cl2, rt, 3 h, 74%; (k) 1 M NaOH, MeOH, rt, 1 d, quant.
7-, and 8-membered rings, respectively, compound 20f had the
most potent CCR3 inhibitory activity (IC50 = 12 nM). These results
indicated that the molecular size of the 6 or 7 member ring sys-
tems in Part A would be favorable for potent inhibitory activity
against the CCR3 receptor. The larger ClogD7.4 values the com-
pounds 20e, 20b, 20f, and 20g had, the stronger the CYP2D6 inhib-
itory activity was observed.
Table 1
CCR3 and CYP2D6 inhibitory activities of 6-fluoronaphthalene derivatives (6a–6g)
R
O
N
N
H
F
Correlation analyses were performed based on the results ob-
tained above and using the IC50 values of CYP2D6 inhibitory activ-
ity and the ClogD7.4 values of the synthesized compounds. The
pIC50 (ꢀlogIC50) values were plotted against the ClogD7.4 values
in a scatter diagram. As shown in Figure 2, the coefficient of deter-
mination obtained from this diagram showed that the value of R2
was 0.8241, which would explain the positive correlation between
the pIC50 values of CYP2D6 inhibitory activity and the ClogD7.4 val-
ues. Since the results of this analysis supported our hypothesis, we
assumed that the further attenuation of CYP2D6 inhibitory activity
could be accomplished by further lowering the ClogD7.4 values. To
accomplish this, we planned to replace the phenyl ring of 20b, the
position of which corresponded to the Part B of 1, with pyridine
rings.
a
b
c
Compound
R
CCR3, IC50
(nM)
CYP2D6, IC50
(nM)
ClogD7.4
1
Phenyl
20 3.9
613 100
29 7.8
57 16
93 22
89 27
400
NTd
0
3.59
2.23
2.30
2.27
1.27
2.64
3.07
1.91
6a
6b
6c
6d
6e
6f
Pyridin-4-yl
Pyridin-3-yl
Pyridin-2-yl
Pyrimidin-5-yl
1,3-Thiazol-2-yl
1H-pyrrol-1-yl
1H-imidazol-1-yl
1600 130
4000 150
2900e
2400e
221 49
217 73
NTd
6g
NTd
a
The IC50 values are shown as means SEM for at least three determinations.
The IC50 values are shown as means SEM for at least four determinations.
b
c
See Ref. 7.
Not tested.
Mean of two experiments.
d
e

I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
8611
Table 4
Table 2
CCR3 and CYP2D6 inhibitory activities of 6-fluoronaphthalene derivatives (24, 28, 30,
32, and 36)
CCR3 and CYP2D6 inhibitory activities of 6-fluoronaphthalene derivatives (6h, 6i, 20a,
and 20b)
R
O
N
O
O
2
N
H
N
5
4
N
N
F
Ar
3
H
F
a
b
c
Compound
R
CCR3, IC50
(nM)
CYP2D6, IC50
(nM)
ClogD7.4
a
b
c
Compound
Ar
CCR3, IC50 (nM)
CYP2D6, IC50 (nM)
6500d
ClogD7.4
*
1
20 3.9
118 38
400
0
3.59
*
20b
23 5.0
0.84
*
*
6h
20a
6i
580e
3.98
1.89
2.7
O
*
*
*
24
28
32
36
98 8.1
13 3.4
85 12
18,300 1630
6800 510
26,700d
0.72
0.17
0.34
N
N
N
O
*
44 0.6
279 64
23 5.0
2900 120
*
NTd
*
N
*
*
N
*
6500e
0.84
N
O
20b
131 6.0
23 4.7
23,500d
29,000d
0.13
*
*
a
The IC50 values are shown as means SEM for at least three determinations.
The IC50 values are shown as means SEM for at least four determinations.
N
b
c
*
See Ref. 7.
Not tested.
d
e
*
30
ꢀ2.36
N+
Mean of two experiments.
O
a
The IC50 values are shown as means SEM for at least three determinations.
The IC50 values are shown as means SEM for at least four determinations.
See Ref. 7.
b
c
Table 3
CCR3 and CYP2D6 inhibitory activities of 6-fluoronaphthalene derivatives (20c–20g)
d
Mean of two experiments.
R
O
O
N
6.5
6
N
H
F
a
c
Compound
R
CCR3, IC50
(nM)
CYP2D6,
ClogD7.4
IC50b(nM)
20b
20c
20d
20e
20f
Piperidin-1-yl
23 5.0
6500d
0.84
ꢀ0.35
ꢀ0.7
0.29
1.41
1.97
4-Methylpiperazin-1-yl
Morpholin-4-yl
Pyrrolidin-1-yl
Azepan-1-yl
482 201
679 129
225 82
12 5.0
7100d
5.5
8200 570
7300d
3000 200
2700d
20g
Azocan-1-yl
62 7.9
5
a
R2 = 0.8241
The IC50 values are shown as means SEM for at least three determinations.
The IC50 values are shown as means SEM for at least four determinations.
See Ref. 7.
b
c
d
4.5
Mean of two experiments.
-2
0
2
4
6
ClogD7.4
Figure 2. Scatter plot of pIC50 of CYP2D6 against ClogD7.4 values.
The results are shown in Table 4. As for CCR3 inhibitory activity,
the potency of 4-pyridyl derivative 28 was twofold compared to
that of 1 (IC50 = 13 nM), whereas other substituted patterns (24,
32, and 36) led to four- to sixfold decreases in activity. This obser-
vation suggests that, for CCR3 inhibition, the nitrogen atom is tol-
erated only at the 4-position of Part B.
Unfortunately, the CYP2D6 inhibition of compound 28, which
was the most potent inhibitor in this study, showed an activity
equal to that of 20b. In contrast, activity of 24, 32, and 36 was
significantly improved because their ClogD7.4 values were lower
than that of 20b.
In order to reduce the CYP2D6 inhibitory activity of 28, we de-
signed and synthesized the pyridine N-oxide derivative 30, which
had much lower ClogD7.4 value (ꢀ2.36) than 28. Predictably, com-
pound 30’s inhibitory activity against CCR3 was equal to that of 1,
with reduced activity against CYP2D6 (IC50 = 29,000 nM). This

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I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
value was the least potent among this series. Thus, compound 30
was identified as having satisfactory selectivity for CCR3 inhibition
over CYP2D6 inhibition. In addition, compound 30 showed no
inhibitory effects against other CC chemokines (CCR1, CCR2, and
CCR5) at the concentration of 10 lM, and had weak inhibitory
activity against other CYP enzymes (CYP1A2, 2C9, 2C19, and 3A4)
3 h. The catalyst was removed by filtration on Celite and the filtrate
was concentrated in vacuo to yield 3 (29.12 g, 96%) as a colorless
solid. 1H NMR (400 MHz, DMSO-d6) d: 1.38 (s, 9H), 1.61 (dd,
J = 12.0, 11.6 Hz, 2H), 1.77–1.86 (m, 4H), 1.89–1.94 (m, 2H), 3.58–
3.70 (m, 1H), 3.82–3.88 (m, 2H), 6.94–6.99 (m, 1H); MS (FAB) m/
z = 227 [M+H]+.
with IC50 values of more than 40,000 nM.
Further biological, pharmacokinetic and pharmacotoxic
evaluations are now being carried out for 30.
5.1.2. tert-Butyl {(3-exo)-8-[(6-fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}carbamate (4)
To a solution of 3 (29.11 g, 111 mmol) in DMF (400 mL) were
added K2CO3 (45.93 g, 332 mmol) and 2-(bromomethyl)-6-fluoro-
naphthalene9 (29.13 g, 122 mmol), and the mixture was stirred
at room temperature for 21 h. The mixture was concentrated in va-
cuo. The residue was then partitioned between EtOAc and satd
NaHCO3 aq, and the organic layer was washed with brine, dried
over Na2SO4, filtered, and concentrated in vacuo. The residue was
purified by silica gel column chromatography (CHCl3/
MeOH = 100:0–98:2) to yield 4 (42.62 g, quant.) as a pale yellow
oil. 1H NMR (300 MHz, CDCl3) d: 1.44 (s, 9H), 1.45–1.56 (m, 2H),
1.69–1.77 (m, 2H), 1.79–1.86 (m, 2H), 2.03–2.08 (m, 2H),
3.21–3.26 (m, 1H), 3.67(s, 2H), 3.78–3.88 (m, 1H), 4.30–4.39 (m,
1H), 7.22 (dd, J = 8.6, 2.6 Hz, 1H), 7.43 (dd, J = 9.9, 2.4 Hz, 1H),
7.56 (d, J = 8.4 Hz, 1H), 7.70–7.81 (m, 3H). MS (FAB) m/z = 385
[M+H]+.
4. Conclusions
Over the course of our discovery research for a novel type of
CCR3 antagonist to attenuate the CYP2D6 inhibition of lead com-
pound 1, novel compounds with relatively low ClogD7.4 values
were synthesized and their inhibitory activities against CCR3 and
CYP2D6 were evaluated. These SAR studies revealed that the car-
bonyl group at Part A of 1 effectively inhibited CCR3 activity, and
it allowed the incorporation of hetero non-aromatic rings into Part
A. In addition, it could also lower the ClogD7.4 value accompanying
reduced CYP2D6 inhibition. Furthermore, replacements of the phe-
nyl ring at Part B with a pyridine ring lowered ClogD7.4 value, and
could cause further reduction of CYP2D6 inhibitory activity. In this
study, good correlation was observed in the ClogD7.4 values and
CYP2D6 inhibition, and we could confirm that the ClogD7.4 values
was an important contributor for CYP2D6 inhibition. These results
indicate the successful identification of 30 as a potent CCR3 inhib-
itor with reduced CYP2D6 activity. For our series of synthesized
compounds, the strategic synthesis of compounds that would
attenuate CYP2D6 inhibitory activity was accomplished by refer-
ring to the ClogD7.4 values.
5.1.3. (3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabicyclo-
[3.2.1]octan-3-amine dihydrochloride (5)
To a solution of 4 (42.62 g, 111 mmol) in EtOAc (300 mL) was
added 4 M HCl (g)/EtOAc (300 mL), and the mixture was stirred
at room temperature for 5.5 h. The mixture was concentrated in
vacuo to yield 5 (39.66 g, quant.) as a slightly brown foam. 1H
NMR (400 MHz, DMSO-d6) d: 1.89–2.09 (m, 7H), 2.18–2.27 (m,
2H), 2.36–2.42 (m, 1H), 3.78, 3.87(m, 2H), 4.32, 4.89 (m, 2H),
7.45–7.56 (m, 1H), 7.75–7.81 (m, 1H), 7.95–8.05 (m, 3H), 8.33–
8.39 (m, 2H), 8.64–8.70 (m, 1H), 11.21 (br, 1H); MS (FAB) m/
z = 285 [M+H]+.
5. Experimental
5.1. Chemistry
In general, reagents and solvents were used as purchased with-
out further purification. Melting points were determined with a
Yanaco MP-500D melting point apparatus and left uncorrected.
1H NMR spectra were recorded on a JEOL JNM-LA300 or a JEOL
JNM-EX400 spectrometer. Chemical shifts were expressed in d
(ppm) values with tetramethylsilane as an internal standard
(NMR descriptions; s = singlet, d = doublet, t = triplet, dt = double
triplet, m = multiplet and br = broad peak). Mass spectra were re-
corded on a JEOL JMS-LX2000 spectrometer. High resolution
(HR)-mass spectra were recorded using a Waters QTOF Premier
spectrometer. The elemental analyses were performed with a
Yanaco MT-5 microanalyzer (C, H, N) and Yokogawa IC-7000S ion
chromatographic analyzer (halogens) and were within 0.4% of
those theoretical values.
5.1.4. 4-(2-Methylphenyl)pyridine (8a)
To
a solution of 4-bromopyridine hydrochloride (1.94 g,
10.0 mmol) in DME (15 mL) were added 2-methylphenyl boronic
acid (7) (1.50 g, 11.0 mmol), CsF (3.34 g, 22.0 mmol), and Pd(PPh3)4
(0.35 g, 0.30 mmol), and the mixture was stirred at reflux for
14.5 h. The mixture was cooled to room temperature and the res-
idue was then partitioned between EtOAc and H2O, and the organic
layer was washed with brine, dried over Na2SO4, filtered, and con-
centrated in vacuo. The residue was purified by silica gel column
chromatography (hexane/EtOAc = 20:1–10:1) to yield 8a (0.918 g,
54%) as a colorless oil. 1H NMR (300 MHz, CDCl3) d: 2.28 (s, 3H),
7.18–7.36 (m, 6H), 8.65 (dd, J = 4.9, 1.7 Hz, 2H); MS (FAB) m/
z = 170 [M+H]+.
5.1.5. 2-Pyridin-4-ylbenzoic acid (9a)
5.1.1. tert-Butyl (3-exo)-8-azabicyclo[3.2.1]oct-3-ylcarbamate
To a solution of 8a (918 mg, 5.42 mmol) in H2O (10 mL) was
added KMnO4 (3.24 g, 16.3 mmol) and the mixture was stirred at
reflux for 2 h. To the mixture was then added EtOH (4 mL) and
the mixture was stirred at reflux for 1 h. The mixture was cooled
to room temperature and filtered through a pad of Celite, and the
filtrate was concentrated in vacuo. The residue was dissolved in
H2O, and the mixture was acidified with 1 M HCl (pH 6). The pre-
cipitate was collected by filtration, washed with H2O, and dried in
vacuo to yield 9a (374 mg, 35%) as a colorless solid. 1H NMR
(300 MHz, DMSO-d6) d: 7.34 (dd, J = 4.5, 1.7 Hz, 2H), 7.41 (dd,
J = 7.6, 1.1 Hz, 2H), 7.55 (ddd, J = 7.6, 7.6, 1.1 Hz, 2H), 7.55 (ddd,
J = 7.6, 7.6, 1.4 Hz, 2H), 7.85 (dd, J = 7.6, 1.4 Hz, 2H), 8.59 (dd, J =
4.5, 1.7 Hz, 2H); MS (FAB) m/z = 200 [M+H]+.
hydrochloride (3)
To
a solution of 3-exo-8-benzyl-8-azabicyclo[3.2.1]octan-3-
amine (2)8 (31.32 g, 145 mmol) in THF (600 mL) was added
di-tert-butyl carbonate (34.76 g, 159 mmol), and the mixture was
stirred at room temperature for 11 h. The mixture was concen-
trated in vacuo. The crude solid was washed with Et2O, and the
precipitate was collected by filtration, washed with Et2O, and dried
in vacuo to yield tert-butyl (3-exo-8-benzyl-8-azabicyclo[3.2.1]oct-
3-yl)carbamate (36.54 g, 79%) as a colorless powder. To an ice-
cooled solution of above-obtained compound (36.54 g, 115 mmol)
in EtOH (500 mL) and MeOH (250 mL) were added 4 M HCl (g)/
EtOAc (27 mL), and Pd(OH)2 (20 wt%, 11.5 g), and the mixture
was stirred in a hydrogen atmosphere at room temperature for

I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
8613
5.1.6. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabicyclo
5.1.12. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
[3.2.1]oct-3-yl}-2-pyridin-4-ylbenzamide (6a)
azabicyclo[3.2.1]oct-3-yl}-2-pyridin-2-ylbenzamide (6c)
To a solution of 5 (324 mg, 0.75 mmol) in CH2Cl2 (3 mL)
were added 9a (166 mg, 0.83 mmol), HOBt (123 mg, 0.91 mmol),
WSC (175 mg, 1.13 mmol), and Et3N (0.33 mL, 2.37 mmol), and
this mixture was stirred at room temperature over night. The
mixture was then partitioned between CHCl3 and satd NaHCO3
aq, and the organic layer was dried over MgSO4, filtered, and
concentrated in vacuo. The residue was purified by silica gel
column chromatography (CHCl3/MeOH = 99:1–98:2). The crude
solid was recrystallized from MeCN to yield 6a (172 mg, 49%)
Compound 6c was prepared from 5 and 9c in a manner similar
to that described for compound 6a, with a yield of 77% as a color-
less crystal. Mp: 187–189 °C (EtOAc–EtOH). 1H NMR (400 MHz,
CDCl3) d: 1.24–1.33 (m, 2H), 1.63–1.68 (m, 2H), 1.69–1.74 (m,
2H), 2.02–2.07 (m, 2H), 3.14–3.20 (m, 2H), 3.62 (s, 2H), 4.11–4.23
(m, 1H), 5.78 (d, J = 8.8 Hz, 1H), 7.21–7.30 (m, 2H), 7.40–7.55 (m,
6H), 7.62–7.65 (m, 1H), 7.68–7.79 (m, 4H), 8.64–8.68 (m, 1H);
MS (FAB) m/z = 466 [M+H]+. Anal. Calcd for C30H28FN3O: C, 77.39;
H, 6.06; N, 9.03; F, 4.08. Found: C, 77.42; H, 6.05; N, 9.11; F, 4.07.
as
a
colorless crystal. Mp: 103–107 °C (MeCN). 1H NMR
(400 MHz, CDCl3) d: 1.17–1.26 (m, 2H), 1.59–1.66 (m, 2H),
1.68–1.76 (m, 2H), 2.02–2.08 (m, 2H), 3.14–3.20 (m, 2H), 3.58
(s, 2H), 4.10–4.23 (m, 1H), 5.10 (d, J = 8.3 Hz, 1H), 7.21–7.27
(m, 1H), 7.34–7.38 (m, 3H), 7.39–7.43 (m, 1H), 7.44–7.54 (m,
4H), 7.62 (dd, J = 7.3, 1.5 Hz, 1H), 7.67–7.68 (m, 1H), 7.72 (d,
J = 8.3 Hz, 1H), 7.78 (dd, J = 8.8, 5.9 Hz, 1H), 8.63 (dd, J = 4.7,
1.8 Hz, 1H); MS (FAB) m/z = 466 [M+H]+. Anal. Calcd for
5.1.13. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-pyrimidin-5-ylbenzamide
fumarate (6d)
To a solution of 5-bromopyrimidine (1.79 g, 11.0 mmol) in DME
(60 mL) were added Pd(PPh3)4 (347 mg, 0.30 mmol), Na2CO3
(1.07 g, 10.0 mmol) in H2O (15 mL), and o-formylphenylboronic acid
(10) (1.55 g, 10.0 mmol), and the mixture was stirred at reflux over
night. The mixture was then partitioned between EtOAc and H2O,
and the organic layer was washed with brine, dried over MgSO4, fil-
tered, and concentrated in vacuo. The residue was purified by silica
gel column chromatography (CHCl3/MeOH = 100:0–98:2) to yield
11a (1.84 g, quant.) as a pale yellow solid. To a solution of above-ob-
tained residual compound (920 mg, 5.00 mmol) in CH3CN (20 mL)
and H2O (8 mL) were added NaH2PO4 (0.16 g, 1.33 mmol), and 35%
H2O2 aq (0.5 mL, 5.15 mmol), was added NaClO2 (792 mg,
8.76 mmol) in H2O (7 mL) dropwise over 2 h and the mixture was
stirred under 15 °C for 4 h. The resulting precipitate was collected
by filtration, and washed with cold-H2O, and dried in vacuo to yield
12a (420 mg, 40%, 90% purity) as a cream-colored solid. To a solution
of 5 (324 mg, 0.75 mmol) in CH2Cl2 (3 mL) were added 12a (164 mg,
0.75 mmol, 90% purity), HOBt (123 mg, 0.91 mmol), WSC (175 mg,
1.13 mmol), and Et3N (0.33 mL, 2.37 mmol), and this mixture was
stirred at room temperature over night. The mixture was then parti-
tioned between CHCl3 and satd NaHCO3 aq, and the organic layer
was dried over MgSO4, filtered, and concentrated in vacuo. The res-
idue was purified by silica gel column chromatography (CHCl3/
MeOH = 99:1–96:4). The crude amorphous solid was converted to
its fumaric acid salt by treating it with fumaric acid (70 mg,
0.60 mmol). The crude solid was recrystallized from EtOH–EtOAc
to yield 6d (270 mg, 62%) as a colorless crystal. Mp: 193–196 °C
(EtOAc–EtOH). 1H NMR (400 MHz, DMSO-d6) d: 1.51–1.57 (m, 4H),
1.63–1.70 (m, 2H), 2.05–2.12 (m, 2H), 3.25–3.33 (m, 2H), 3.79 (s,
2H), 3.98–4.09 (m, 1H), 6.61 (s, 2H), 7.40–7.46 (m, 1H), 7.50–7.63
(m, 5H), 7.70 (dd, J = 10.2, 2.4 Hz, 1H), 7.82–7.92 (m, 2H), 7.97 (dd,
J = 9.3, 5.8 Hz, 1H) , 8.24 (d, J = 8.3 Hz, 1H), 8.76 (s, 2H) , 9.16 (s,
1H); MS (FAB) m/z = 467 [M+H]+. Anal. Calcd for C29H27FN4Oꢁ
C4H4O4: C, 68.03; H, 5.36; N, 9.62; F, 3.26. Found: C, 67.98; H, 5.43;
N, 9.62; F, 3.28.
C30H28FN3O: C, 77.39; H, 6.06; N, 9.03; F, 4.08. Found: C,
77.21; H, 6.04; N, 9.02; F, 3.98.
5.1.7. 3-(2-Methylphenyl)pyridine (8b)
Compound 8b was prepared from 7 and 3-bromopyridine in a
manner similar to that described for compound 8a, with a yield
of 97% as a pale yellow oil. 1H NMR (300 MHz, CDCl3) d: 2.28 (s,
3H), 7.19–7.38 (m, 5H), 7.63–7.68 (m, 1H), 8.57–8.62 (m, 2H);
MS (FAB) m/z = 170 [M+H]+.
5.1.8. 2-Pyridin-3-ylbenzoic acid (9b)
Compound 9b was prepared from 8b in a manner similar to that
described for compound 9a, with a yield of 28% as a colorless solid.
1H NMR (300 MHz, DMSO-d6) d: 7.40–7.47 (m, 2H), 7.53 (ddd,
J = 7.5, 7.5, 1.4 Hz, 1H), 7.64 (ddd, J = 7.5, 7.5, 1.7 Hz, 1H), 7.75
(ddd, J = 7.7, 2.2, 1.7 Hz, 2H), 7.84–7.88 (m, 1H), 8.51 (dd, J = 2.4,
0.7 Hz, 1H), 8.55 (dd, J = 4.7, 1.7 Hz, 1H); MS (FAB) m/z = 200
[M+H]+.
5.1.9. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabicyclo
[3.2.1]oct-3-yl}-2-pyridin-3-ylbenzamide dihydrochloride (6b)
Compound 6b was prepared from 5 and 9b in a manner sim-
ilar to that described for compound 6a, with a yield of 55% as a
colorless solid. Mp: 174–177 °C (MeCN). 1H NMR (400 MHz,
DMSO-d6) d: 1.64–2.00 (m, 6H), 2.15–2.45 (m, 2H), 4.02–4.25
(m, 1H), 4.25–4.32 (m, 2H), 4.74 (d, J = 5.8 Hz, 1H), 7.48–7.74
(m, 5H), 7.78 (dd, J = 10.3, 2.5 Hz, 1H), 7.81–7.95 (m, 1H), 7.96–
8.05 (m, 3H), 8.23–8.30 (m, 2H), 8.56 (d, J = 7.8 Hz, 1H), 8.77–
8.85 (m, 3H), 10.75–11.00 (m, 1H); MS (FAB) m/z = 466 [M+H]+.
Anal. Calcd for C30H28FN3Oꢁ2HClꢁ2H2O: C, 61.75; H, 6.05; N,
7.20; Cl, 12.15; F, 3.76. Found: C, 61.78; H, 6.03; N, 7.38; Cl,
11.99; F, 3.05.
5.1.14. 2-(1,3-Thiazol-2-yl)benzoic acid (12b)
5.1.10. 2-(2-Methylphenyl)pyridine (8c)
Compound 12b was prepared from 10 and 2-bromothiazole in a
manner similar to that described for compound 12a, with a yield of
39% from 10 as a cream-colored solid. 1H NMR (400 MHz, DMSO-
d6) d: 7.55–7.64 (m, 2H), 7.68–7.73 (m, 2H), 7.84 (d, J = 3.2 Hz,
1H), 7.92 (d, J = 3.2 Hz, 1H), 12.96 (s, H); MS (ESI+) m/z = 206.1
[M+H]+.
Compound 8c was prepared from 7 and 2-bromopyridine in a
manner similar to that described for compound 8a, with a yield
of 81% as a pale yellow oil. 1H NMR (300 MHz, CDCl3) d: 2.37 (s,
3H), 7.21–7.32 (m, 4H), 7.37–7.42 (m, 2H), 7.71–7.78 (m, 1H),
8.68–8.74 (m, 1H); MS (FAB) m/z = 170 [M+H]+.
5.1.11. 2-Pyridin-2-ylbenzoic acid (9c)
5.1.15. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
Compound 9c was prepared from 8c in a manner similar to that
described for compound 9a, with a yield of 55% as a colorless solid.
1H NMR (300 MHz, DMSO-d6) d: 7.35 (ddd, J = 7.5, 4.8, 1.0 Hz, 2H),
7.48–7.62 (m, 4H), 7.69–7.73 (m, 1H), 7.86 (ddd, J = 7.7, 7.7, 1.8 Hz,
2H); MS (FAB) m/z = 200 [M+H]+.
azabicyclo[3.2.1]oct-3-yl}-2-(1,3-thiazol-2-yl)benzamide (6e)
Compound 6e was prepared from 5 and 12b in a manner similar
to that described for compound 6a, with a yield of 73% as a color-
less crystal. Mp: 163–165 °C (MeCN). 1H NMR (400 MHz, CDCl3) d:
1.40–1.48 (m, 2H), 1.74–1.82 (m, 4H), 2.04–2.09 (m, 2H), 3.20–3.35

8614
I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
(m, 2H), 3.65 (s, 2H), 4.20–4.34 (m, 1H), 6.14 (d, J = 8.3 Hz, 1H),
7.21–7.27 (m, 1H), 7.41–7.50 (m, 4H), 7.56 (d, J = 8.8 Hz, 1H),
7.61–7.67 (m, 2H), 7.73–7.74 (m, 2H) , 7.78 (dd, J = 8.8, 5.9 Hz,
1H), 7.88 (d, J = 3.0 Hz, 1H); MS (FAB) m/z = 472 [M+H]+. Anal. Calcd
for C28H26FN3O2S: C, 71.31; H, 5.56; N, 8.91; S, 6.80; F, 4.03. Found:
C, 71.29; H, 5.53; N, 8.92; S, 6.79; F, 4.04.
5.1.20. 2-(Cyclohexyloxy)-N-{(3-exo)-8-[(6-fluoro-2-
naphthyl)methyl]-8-azabicyclo[3.2.1]oct-3-yl}benzamide
fumarate (6h)
Compound 6h was prepared from 5 and 15 in a manner similar
to that described for compound 6a, with a yield of 77% as a color-
less crystal. Mp: 211–214 °C (EtOAc–EtOH). 1H NMR (400 MHz,
DMSO-d6) d: 1.26–1.59 (m, 6H), 1.67–1.87 (m, 8H), 1.92–1.99 (m,
2H), 2.10–2.18 (m, 2H), 3.32–3.39 (m, 2H), 3.85 (s, 2H), 4.16–
4.29 (m, 1H), 4.50–4.58 (m, 1H), 6.61 (s, 2H), 7.00 (dd, J = 7.9,
7.3 Hz, 1H), 7.16 (d, J = 8.3 Hz, 1H), 7.39–7.46 (m, 2H), 7.64 (d,
J = 8.3 Hz, 1H) , 7.70 (dd, J = 10.8, 2.7 Hz, 1H), 7.73 (dd, J = 7.8,
2.0 Hz, 1H), 7.90 (d, J = 8.8 Hz, 1H) , 7.92–7.98 (m, 2H), 8.01 (d,
J = 7.9 Hz, 1H); MS (FAB) m/z = 487 [M+H]+. Anal. Calcd for
C31H35FN2O2ꢁC4H4O4: C, 69.75; H, 6.52; N, 4.65; F, 3.15. Found: C,
69.69; H, 6.59; N, 4.67; F, 3.18.
5.1.16. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-(1H-pyrrol-1-yl)benzamide (6f)
Compound 6f was prepared from 5 and 2-(1H-pyrrol-1-yl)ben-
zoic acid in a manner similar to that described for compound 6a,
with a yield of 55% as a colorless crystal. Mp: 144–146 °C (iPr2O–
MeOH); 1H NMR (400 MHz, CDCl3) d: 1.22–1.30 (m, 2H), 1.63–
1.75 (m, 4H), 2.02–2.07 (m, 2H), 3.15–3.20 (m, 2H), 3.64 (s, 2H),
4.08–4.20 (m, 1H), 5.03 (d, J = 8.3 Hz, 1H), 6.38 (dd, J = 2.3, 1.9 Hz,
1H), 6.84 (dd, J = 2.3, 1.9 Hz, 1H), 7.21–7.28 (m, 1H), 7.34 (dd,
J = 8.8, 1.5 Hz, 1H), 7.41–7.46 (m, 2H), 7.49 (ddd, J = 7.8, 7.3,
1.8 Hz, 1H), 7.54 (d, J = 8.8 Hz, 1H) , 7.71–7.75 (m, 2H), 7.78 (dd,
J = 8.8, 5.9 Hz, 1H), 7.84 (dd, J = 7.8, 2.2 Hz, 1H); MS (FAB) m/
z = 454 [M+H]+. Anal. Calcd for C29H28FN3O: C, 76.80; H, 6.22; N,
9.26; F, 4.19. Found: C, 76.71; H, 5.95; N, 9.33; F, 4.10.
5.1.21. 2-Piperidin-1-ylbenzoic acid (17)
To a solution of 2-fluorobenzonitrile (16) (2.47 g, 20.0 mmol)
in DMF (20 mL) were added piperidine (3.61 g, 42.0 mmol) and
K2CO3 (2.77 g, 20.0 mmol), and the mixture was stirred at
70 °C over night. The mixture was then partitioned between
EtOAc and H2O, and the organic layer was washed with brine,
dried over MgSO4, filtered, and concentrated in vacuo to yield
2-piperidin-1-ylbenzonitrile (3.18 g, 85%) as a brown oil. To a
solution of above-obtained residual compound (1.00 g,
5.37 mmol) in 2-ethoxyethanol (4 mL) and H2O (0.5 mL) was
added KOH (2.23 g, 39.8 mmol), and the mixture was stirred
at reflux for 4 h. The mixture was then cooled to 0 °C, and
neutralized with concd HCl (2.8 mL), and concentrated in va-
cuo. The residue was purified by silica gel column chromatog-
5.1.17. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-(1H-imidazol-1-yl)benzamide
difumarate (6g)
Compound 6g was prepared from 5 and 2-(1H-imidazol-1-
yl)benzoic acid in a manner similar to that described for compound
6a, with a yield of 16% as a colorless crystal. Mp: 202–204 °C
(MeCN–MeOH). 1H NMR (400 MHz, DMSO-d6) d: 1.48–1.70 (m,
6H), 2.05–2.15 (m, 2H), 3.23–3.31 (m, 2H), 3.77 (s, 2H), 3.96–4.08
(m, 1H), 6.61 (d, 4H), 7.01 (s, 1H), 7.28 (s, 1H), 7.38–7.53 (m,
4H), 7.54–7.63 (m, 2H), 7.69 (dd, J = 10.3, 2.4 Hz, 1H), 7.74 (s,
1H), 7.86–7.92 (m, 2H) , 7.96 (dd, J = 8.8, 5.8 Hz, 1H), 8.20 (d,
J = 7.9 Hz, 1H); MS (FAB) m/z = 455 [M+H]+. Anal. calcd for
raphy (CHCl3/MeOH = 98:2) to yield 17 (0.64 g, 58%) as
a
brown solid. 1H NMR (300 MHz, DMSO-d6) d: 1.59–1.68 (m,
2H), 1.70–1.81 (m, 4H), 3.01–3.08 (m, 4H), 3.30–3.34 (m,
2H), 7.40–7.47 (m, 1H), 7.67 (dt, J = 8.1, 1.7 Hz, 1H), 7.70–
7.75 (m, 1H), 8.05 (dd, J = 7.8, 1.6 Hz, 1H); MS (FAB) m/
z = 206 [M+H]+.
C
28H27FN4Oꢁ2C4H4O4: C, 62.97; H, 5.14; N, 8.16; F, 2.77. Found: C,
63.10; H, 5.19; N, 8.46; F, 2.70.
5.1.18. Methyl 2-(cyclohexyloxy)benzoate (14)
5.1.22. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-piperidin-1-ylbenzamide (6i)
Compound 6i was prepared from 5 and 17 in a manner similar
to that described for compound 6a, with a yield of 25% as a pale
yellow crystal. Mp: 157–159 °C (MeOH); 1H NMR (400 MHz, CDCl3)
d: 1.63–1.75 (m, 4H), 1.78–1.86 (m, 6H), 1.96–2.03 (m, 2H), 2.08–
2.15 (m, 2H), 2.89–2.96 (m, 4H), 3.27–3.33 (m, 2H), 3.73 (s, 2H),
4.32–4.44 (m, 1H), 7.18–7.27 (m, 3H), 7.38–7.46 (m, 2H), 7.61 (d,
J = 8.3 Hz, 1H), 7.74–7.80 (m, 3H), 8.19 (dd, J = 8.0, 1.5 Hz, 1H) ,
10.13 (d, J = 8.3 Hz, 1H); MS (FAB) m/z = 472 [M+H]+. Anal. Calcd
for C30H34FN3O: C, 76.40; H, 7.27; N, 8.91; F, 4.03. Found: C,
76.31; H, 7.40; N, 8.85; F, 3.87.
To a solution of methyl salicylate (13) (3.38 g, 22.0 mmol) in
THF (4 mL) was added cyclohexanol (2.02 g, 20.0 mmol), PPh3
(5.77 g, 22.0 mmol), and DEAD (3.31 mL, 21.0 mmol), and the mix-
ture was stirred at room temperature for 4 d. The mixture was con-
centrated in vacuo, and the solid was then diluted with Et2O and
insoluble matter was removed by filtration, and the filtrate was
concentrated in vacuo The residue was purified by silica gel col-
umn chromatography (hexane/Et2O = 95:5–90:10) to yield 14
(2.67 g, 57%) as a pale yellow oil. 1H NMR (400 MHz, CDCl3) d:
1.32–1.45 (m, 3H), 1.47–1.58 (m, 1H), 1.62–1.72 (m, 2H), 1.76–
1.87 (m, 2H), 1.88–1.97 (m, 2H), 3.88 (s, 3H), 4.32–4.39 (m, 1H),
6.78 (dd, J = 8.3, 7.3 Hz, 1H), 6.99 (d, J = 8.3 Hz, 1H), 7.40 (ddd,
J = 7.8, 7.3, 2.0 Hz, 1H), 7.75 (dd, J = 7.8, 2.0 Hz, 1H); MS (FAB)
m/z = 235 [M+H]+.
5.1.23. 2-[Cyclohexyl(methyl)carbamoyl]benzoic acid (19a)
To a solution of cyclohexylmethylamine (1.55 g, 13.7 mmol)
in THF (30 mL) was added phthalic anhydride (18) (2.04 g,
13.7 mmol), and the mixture was stirred at room temperature
for 15.5 h. The mixture was then concentrated in vacuo. The res-
idue was purified by silica gel column chromatography (CHCl3/
MeOH = 98:2–90:10) to yield 19a (2.72 g, 76%) as a colorless
amorphous solid. 1H NMR (400 MHz, DMSO-d6) d: 0.85–1.02
(m, 2H), 1.25–1.84 (m, 8H), 1.94–2.12 (m, 6H), 2.32–2.50 (m,
2H), 2.56–2.59, 2.78–2.84 (each m, 3H), 3.05–3.14 (m, 1H),
3.76–3.92, 4.31–4.36 (each m, 2H), 4.10–4.30 (m, 1H), 4.79–
4.84 (m, 1H), 7.18–7.29 (m, 2H), 7.39–7.56 (m, 4H), 7.76–7.82
(m, 2H), 7.90–8.08 (m, 2H), 8.23–8.27 (m, 1H), 8.29–8.33,
8.39–8.43 (each m, 1H), 10.40–10.66 (m,1H); MS (FAB)
m/z = 260 [M+H]+.
5.1.19. 2-(Cyclohexyloxy)benzoic acid (15)
To a solution of 14 (1.17 g, 5.00 mmol) in MeOH (10 mL) was
added 1 M NaOH aq (6.5 mL), and the mixture was stirred at room
temperature for 3.5 d. The mixture was concentrated in vacuo, and
the residue was then partitioned between CHCl3 and 1 M HCl aq,
and the organic layer was washed with brine, dried over MgSO4,
filtered, and concentrated in vacuo. to yield 15 (1.10 g, quant.) as
a pale yellow oil. 1H NMR (400 MHz, DMSO-d6) d: 1.28–1.41 (m,
3H), 1.42–1.56 (m, 3H), 1.66–1.78 (m, 2H), 1.80–1.88 (m, 2H),
4.41–4.49 (m, 1H), 6.96 (dd, J = 7.8, 7.3 Hz, 1H), 7.13 (d,
J = 8.3 Hz, 1H), 7.44 (ddd, J = 8.3, 7.3, 2.0 Hz, 1H), 7.59 (dd, J = 7.8,
2.0 Hz, 1H 1H), 12.48 (s, 1H); MS (FAB) m/z = 219 [MꢀH]+.

I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
8615
5.1.24. N-Cyclohexyl-N0-{(3-exo)-8-[(6-fluoro-2-naphthyl)-
methyl]-8-azabicyclo[3.2.1]oct-3-yl}-N-methylphthalamide
hydrochloride (20a)
and the mixture was stirred at reflux for 4 h. The mixture was
cooled to room temperature and then concentrated in vacuo. The
residue was purified by silica gel column chromatography
(CHCl3/MeOH = 98:2–90:10) to yield crude 19d (1.59 g) as a pale
yellow solid. To a solution of 5 (501 mg, 1.40 mmol) in 1,2-dichlo-
roethane (14 mL) were added crude 19d (330 mg, 1.40 mmol),
HOBt (189 mg, 1.14 mmol), WSC (219 mg, 1.41 mmol), and Et3N
(0.42 mL, 3.05 mmol), and this mixture was stirred at room tem-
perature over night. The mixture was then partitioned between
CHCl3 and satd NaHCO3 aq, and the organic layer was dried over
MgSO4, filtered, and concentrated in vacuo. The residue was puri-
fied by silica gel column chromatography (CHCl3/MeOH = 99:1–
98:2) to yield 20d (343 mg, 48%) as a pale yellow amorphous solid.
1H NMR (400 MHz, CDCl3) d: 1.60–1.70 (m, 2H), 1.75–1.92 (m, 4H),
2.06–2.18 (m, 2H), 3.16–3.21 (m, 2H), 3.25–3.30 (m, 2H), 3.53–3.60
(m, 2H), 3.73–3.82 (m, 4H), 4.25–4.38 (m, 1H), 6.43 (d, J = 8.3 Hz,
1H), 7.22–7.26 (m, 1H), 7.41–7.51 (m, 3H), 7.59 (d, J = 8.3 Hz,
1H), 7.68–7.72 (m, 1H), 7.73–7.82 (m, 4H); MS (FAB) m/z = 501
[M+H]+. Anal. Calcd for C30H32FN3O3ꢁ0.5H2O: C, 70.57; H, 6.51; N,
8.23; F, 3.72. Found: C, 70.80; H, 6.50; N, 8.37; F, 3.78.
Compound 20a was prepared from 5 and 19a in a manner sim-
ilar to that described for compound 6a, with a yield of 59% as a col-
orless crystal. Mp: 270–272 °C (MeCN–Et2O). 1H NMR (400 MHz,
DMSO-d6) d: 0.85–1.02 (m, 2H), 1.25–1.84 (m, 8H), 1.94–2.12 (m,
6H), 2.32–2.50 (m, 2H), 2.56–2.59, 2.78–2.84 (each m, 3H), 3.05–
3.14 (m, 1H), 3.76–3.92, 4.31–4.36 (each m, 2H), 4.10–4.30 (m,
1H), 4.79–4.84 (m, 1H), 7.18–7.29 (m, 2H), 7.39–7.56 (m, 4H),
7.76–7.82 (m, 2H), 7.90–8.08 (m, 2H), 8.23–8.27 (m, 1H), 8.29–
8.33, 8.39–8.43 (each m, 1H), 10.40–10.66 (m,1H); MS (FAB) m/
z = 528 [M+H]+. Anal. Calcd for C33H38FN3O2ꢁHCl: C, 70.26; H,
6.97; N, 7.45; Cl, 6.28; F, 3.37. Found: C, 69.93; H, 7.14; N, 7.38;
Cl, 6.21; F, 3.42.
5.1.25. 2-(Piperidin-1-ylcarbonyl)benzoic acid (19b)
Compound 19b was prepared from piperidine and phthalic
anhydride (18) in a manner similar to that described for compound
19a, with a yield of 73% as a pale brown amorphous solid. 1H NMR
(400 MHz, CDCl3) d: 1.78–1.84 (m, 2H), 3.08–3.20 (m, 8H), 7.23–
7.28 (m, 1H), 7.41–7.47 (m, 1H), 7.54–7.58 (m, 1H),
8.08(J = 7.9 Hz, 1H), 9.98 (br, 1H); MS (FAB) m/z = 234 [M+H]+.
5.1.30. 2-(Pyrrolidin-1-ylcarbonyl)benzoic acid (19e)
Compound 19e was prepared from pyrrolidine and phthalic
anhydride (18) in a manner similar to that described for compound
19a, with a yield of 75% as a pale brown amorphous solid. 1H NMR
(400 MHz, CDCl3) d: 1.82–1.99 (m, 4H), 3.10–3.17 (m, 2H), 3.62–
3.89 (m, 2H), 7.32 (d, J = 7.4 Hz, 1H), 7.45 (dd, J = 7.8, 7.8 Hz, 1H),
7.55 (dd, J = 7.8, 7.4 Hz, 1H), 8.08 (d, J = 7.8 Hz, 1H); MS (FAB) m/
z = 220 [M+H]+.
5.1.26. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-(piperidin-1-ylcarbonyl)benzamide
(20b)
Compound 20b was prepared from 5 and 19b in a manner
similar to that described for compound 6a, with a yield of 45%
as a pale yellow amorphous solid. 1H NMR (400 MHz, CDCl3) d:
1.37–1.46 (m, 2H), 1.60–1.83 (m, 10H), 2.08–2.12 (m, 2H), 3.10
(t, J = 5.4 Hz, 2H), 3.24–3.29 (m, 2H), 3.68–3.72 (m, 2H), 4.24–
4.36 (m, 1H), 6.73 (d, J = 8.3 Hz, 1H), 7.18–7.27 (m, 2H), 7.41–
7.49 (m, 3H), 7.60 (d, J = 8.8 Hz, 1H), 7.73–7.82 (m, 4H); MS
(FAB) m/z = 500 [M+H]+. Anal. Calcd for C31H34FN3O2ꢁ0.5H2O: C,
73.20; H, 6.94; N, 8.26; F, 3.74. Found: C, 73.08; H, 7.01; N,
8.57; F, 3.80.
5.1.31. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-(pyrrolidin-1-
ylcarbonyl)benzamide (20e)
Compound 20e was prepared from 5 and 19e in a manner sim-
ilar to that described for compound 6a, with a yield of 33% as a
cream-colored amorphous solid. 1H NMR (400 MHz, CDCl3) d:
1.74–1.89 (m, 6H), 1.89–1.98 (m, 2H), 2.10–2.18 (m, 2H), 3.12 (t,
J = 6.8 Hz, 2H), 3.30–3.36 (m, 2H), 3.63 (t, J = 6.8 Hz, 1H), 3.78 (s,
2H), 4.26–4.39 (m, 1H), 6.80 (d, J = 7.3 Hz, 1H), 7.23–7.29 (m,
2H), 7.41–7.50 (m, 3H), 7.66 (d, J = 8.3 Hz, 1H), 7.73–7.84 (m,
4H); MS (FAB) m/z = 486 [M+H]+. HR-MS calcd for C30H32FN3O2
m/z = 486.2557 [M+H]+. Found: 486.2539.
5.1.27. 2-[(4-Methylpiperazin-1-yl)carbonyl]benzoic acid (19c)
Compound 19c was prepared from 1-methylpiperazine and
phthalic anhydride (18) in a manner similar to that described for
compound 19a, with a yield of 96% as a colorless powder. 1H
NMR (400 MHz, D2O) d: 2.94 (s, 3H), 3.05–3.80 (m, 8H), 7.28–
7.35 (m, 1H), 7.55–7.62 (m, 2H), 7.82–7.89 (m, 1H); MS (FAB) m/
z = 249 [M+H]+.
5.1.32. 2-(Azepan-1-ylcarbonyl)benzoic acid (19f)
Compound 19f was prepared from azepane and phthalic
anhydride (18) in a manner similar to that described for com-
pound 19a, with a quantitative yield as a pale brown amorphous
solid. 1H NMR (400 MHz, CDCl3) d: 1.50–1.68 (m, 6H), 1.75–1.86
(m, 2H), 3.13–3.22 (m, 2H), 3.62–3.77 (m, 2H), 7.27 (d, J = 7.4 Hz,
1H), 7.42 (dd, J = 7.4, 7.4 Hz, 1H), 7.55 (dd, J = 7.4, 7.4 Hz, 1H),
8.06 (d, J = 7.4 Hz, 1H), 10.16 (br, 1H); MS (FAB) m/z = 248
[M+H]+.
5.1.28. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-[(4-methylpiperazin-1-
yl)carbonyl]benzamide (20c)
Compound 20c was prepared from 5 and 19c in a manner sim-
ilar to that described for compound 6a, with a yield of 36% as a
slightly brown amorphous solid. 1H NMR (400 MHz, CDCl3) d:
1.56–1.68 (m, 2H), 1.78–1.88 (m, 4H), 2.06–2.14 (m, 2H), 2.23–
2.32 (m, 2H), 2.28 (s, 3H), 2.47 (t, J = 4.9 Hz, 2H), 3.17 (t,
J = 4.9 Hz, 2H), 3.24–3.30 (m, 2H), 3.70 (s, 2H), 3.77–3.86 (m, 2H),
4.24–4.37 (m, 1H), 6.57 (d, J = 8.8 Hz, 1H), 7.21–7.25 (m, 2H),
7.41–7.50 (m, 3H), 7.60 (d, J = 8.3 Hz, 1H), 7.73–7.82 (m, 4H); MS
(FAB) m/z = 515 [M+H]+. Anal. Calcd for C31H35FN4O2ꢁ0.25H2O: C,
71.72; H, 6.89; N, 10.79; F, 3.66. Found: C, 71.57; H, 6.96; N,
10.57; F, 3.75.
5.1.33. 2-(Azepan-1-ylcarbonyl)-N-{(3-exo)-8-[(6-fluoro-2-
naphthyl)methyl]-8-azabicyclo[3.2.1]oct-3-yl}benzamide (20f)
Compound 20f was prepared from 5 and 19f in a manner sim-
ilar to that described for compound 6a, with a yield of 48% as a pale
yellow amorphous solid. 1H NMR (300 MHz, CDCl3) d: 1.47–1.58
(m, 4H), 1.59–1.70 (m, 4H), 1.75–1.89 (m, 6H), 2.05–2.23 (m,
2H), 3.14–3.21 (m, 2H), 3.22–3.29 (m, 2H), 3.65–3.75 (m, 2H),
3.70 (s, 2H), 4.21–4.32 (m, 1H), 6.86 (d, J = 8.0 Hz, 1H), 7.19–7.25
(m, 2H), 7.42–7.49 (m, 3H), 7.59 (d, J = 8.4 Hz, 1H), 7.72–7.82 (m,
5.1.29. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-2-(morpholin-4-
ylcarbonyl)benzamide (20d)
To a solution of morpholine (880 mg, 10.0 mmol) in THF
(50 mL) was added phthalic anhydride (18) (1.48 g, 10.0 mmol),
4H);
32H36FN3O2ꢁ0.5H2O: C,73.54; H, 7.14; N, 8.04; F, 3.63. Found: C,
73.43; H, 7.08; N, 8.19; F, 3.65.
MS
(FAB)
m/z = 514
[M+H]+.
Anal.
Calcd for
C

8616
I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
5.1.34. 2-(Azocan-1-ylcarbonyl)-N-{(3-exo)-8-[(6-fluoro-2-
naphthyl)methyl]-8-azabicyclo[3.2.1]oct-3-yl}benzamide (20g)
Compound 20g was prepared from 5 and 19g in a manner sim-
ilar to that described for compound 20d, with a yield of 56% as a
colorless amorphous solid. 1H NMR (400 MHz, CDCl3) d: 1.40–
1.70 (m, 10H), 1.74–1.98 (m, 8H), 2.05–2.14 (m, 2H), 3.08–3.20
(m, 2H), 3.23–3.29 (m, 2H), 3.70 (s, 2H), 4.22–4.35 (m, 1H), 6.80
(d, J = 7.8 Hz, 1H), 7.20–7.27 (m, 2H), 7.41–7.50 (m, 3H), 7.60 (d,
J = 8.3 Hz, 1H), 7.73–7.82 (m, 4H); MS (FAB) m/z = 528 [M+H]+.
Anal. Calcd for C33H38FN3O2ꢁ0.25H2O: C, 74.48; H, 7.29; N, 7.90;
F, 3.57. Found: C, 74.63; H, 7.47; N, 8.11; F, 3.59.
pounds 26, 27, and piperidine hydrochloride (5.42 g). The
above-obtained residual compound (959 mg) was diluted with
CHCl3 and washed with NaCl–saturated 5% citric acid aq twice,
and the organic layer was dried over Na2SO4, filtered, and concen-
trated in vacuo to give the 1:1 mixture of 26 and 27. The residue
was recrystallized from EtOAc–EtOH to yield pure compound 26
(173 mg, 23% from 25) as
a
colorless powder. 1H NMR
(400 MHz, DMSO-d6) d: 1.38–1.47 (m, 2H), 1.52–1.65 (m, 4H),
3.05–3.10 (m, 2H), 3.55–3.61 (m, 2H), 7.78 (d, J = 4.8 Hz,1H),
8.57 (s, 1H), 8.75 (d, J = 4.8 Hz, 1H), 13.80 (s, 1H); MS (FAB) m/
z = 233 [M+H]+.
5.1.35. Methyl 2-(piperidin-1-ylcarbonyl)nicotinate (22)
5.1.38. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabi-
cyclo[3.2.1]oct-3-yl}-3-(piperidin-1-ylcarbonyl)isonicotinamide
(28)
To
a solution of dimethyl pyridine-2,3-dicarboxylate (21)
(1.02 g, 5.12 mmol) and MgCl2 (249 mg, 2.56 mmol) in THF
(20 mL) was added piperidine (1.33 g, 15.56 mmol) in THF (9 mL)
dropwise and the mixture was stirred at room temperature over
night. The mixture was then partitioned between EtOAc and 1 M
HCl aq, and the organic layer was washed with brine, dried over
MgSO4, filtered, and concentrated in vacuo. The residue was puri-
fied by silica gel column chromatography (CHCl3/MeOH = 99:1–
98:2) to yield 22 (444 mg, 37%) as a pale brown oil. 1H NMR
(400 MHz, CDCl3) d: 1.50–1.58(m, 2H), 1.66–1.77 (m, 4H), 3.14–
3.18 (m, 2H), 3.75–3.80 (m, 2H), 3.92 (s, 2H), 7.39 (dd, J = 8.3,
4.9 Hz, 1H), 8.31 (dd, J = 8.3, 1.4 Hz, 1H), 8.73 (dd, J = 4.9, 1.4 Hz,
1H); MS (FAB) m/z = 249 [M+H]+.
Compound 28 was prepared from 5 and 26 in a manner similar
to that described for compound 6a, with a yield of 51% as a slightly
brown amorphous solid. 1H NMR (400 MHz, CDCl3) d: 1.44–1.50
(m, 2H), 1.64–1.70 (m, 6H), 1.74–1.87 (m, 4H), 2.08–2.13 (m,
2H), 3.12–3.17 (m, 2H), 3.25–3.30 (m, 2H), 3.70 (s, 2H), 3.73–3.78
(m, 2H), 4.23–4.35 (m, 1H), 6.95 (d, J = 8.1 Hz, 1H), 7.21–7.28 (m,
1H), 7.44 (dd, J = 9.8, 2.5 Hz, 1H), 7.59 (d, J = 9.1 Hz, 1H), 7.64–
7.66 (m, 1H), 7.73–7.78 (m, 3H), 8.52 (s, 1H), 7.59 (d, J = 5.4 Hz,
1H);
MS
(FAB)
m/z = 501
[M+H]+.
Anal.
Calcd
for
C
30H33FN4O2ꢁ0.25H2O: C, 71.34; H, 6.68; N, 11.09; F, 3.76. Found:
C, 71.24; H, 6.69; N, 10.93; F, 3.63.
5.1.36. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabi-
cyclo[3.2.1]oct-3-yl}-2-(piperidin-1-ylcarbonyl)nicotinamide (24)
To a solution of 22 (464 mg, 1.87 mmol) in EtOH (4 mL) was
added 1 M NaOH aq (3.8 mL), and the mixture was stirred at room
temperature over night. This mixture was acidified with 1 M HCl
aq, and the precipitate was collected by filtration, washed with
H2O, and dried in vacuo to yield 23 (440 mg, quant.) as a pale
brown solid. To a solution of 5 (669 mg, 1.87 mmol) in DMF
(18 mL) were added 23 (440 mg, 1.87 mmol), HOBt (206 mg,
1.52 mmol), WSC (293 mg, 1.88 mmol), and Et3N (0.57 mL,
4.09 mmol), and this mixture was stirred at room temperature
over night. The mixture was then partitioned between EtOAc and
satd NaHCO3 aq, and the organic layer was washed with H2O and
brine, dried over MgSO4, filtered, and concentrated in vacuo. The
residue was purified by silica gel column chromatography
(CHCl3/MeOH = 99:1–98:2) to yield 24 (575 mg, 61%) as a colorless
amorphous soild. 1H NMR (400 MHz, CDCl3) d: 1.44–1.53 (m, 2H),
1.60–1.88 (m, 10H), 2.07–2.15 (m, 2H), 3.04–3.08 (m, 2H), 3.25–
3.29 (m, 2H), 3.61 (s, 2H), 3.75–3.79 (m, 2H), 4.23–4.32 (m, 1H),
7.23 (dd, J = 8.8, 2.4 Hz, 1H), 7.35–7.48 (m, 3H), 7.60 (d, J = 8.3 Hz,
1H), 7.73–7.82 (m, 3H), 8.18 (dd, J = 8.1, 1.5 Hz, 1H), 8.65 (dd,
J = 4.9, 1.5 Hz, 1H); MS (FAB) m/z = 501 [M+H]+. Anal. Calcd for
C30H33FN4O2ꢁ0.25H2O: C, 71.34; H, 6.68; N, 11.09; F, 3.76. Found:
C, 71.24; H, 6.59; N, 11.27; F, 3.78.
5.1.39. 3-(Piperidin-1-ylcarbonyl)isonicotinic acid 1-oxide (29)
To a solution of the 1:1 mixture of 26 and 27 (269 mg,
1.15 mmol) obtained by the same manner described for compound
26 in CH2Cl2 (3 mL) was added mCPBA (238 mg, 1.38 mmol), and
the mixture was stirred at room temperature for 21 h. The mixture
was directly purified by silica gel column chromatography (CHCl3/
MeOH/AcOH = 97:3:1–94:6:1) to yield 29 (98 mg, 34%) as a slightly
yellow amorphous solid. 1H NMR (400 MHz, DMSO-d6) d: 1.39–
1.50 (m, 2H), 1.51–1.63 (m, 4H), 3.05–3.20 (m, 2H), 3.45–3.62
(m, 2H), 7.82 (d, J = 6.9 Hz, 1H), 8.23–8.29 (m, 2H); MS (FAB) m/
z = 251 [M+H]+.
5.1.40. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabi-
cyclo[3.2.1]oct-3-yl}-3-(piperidin-1-ylcarbonyl)isonicotinamide
1-oxide (30)
Compound 30 was prepared from 5 and 29 in a manner simi-
lar to that described for compound 6a, with a yield of 58% as a
slightly yellow amorphous solid. 1H NMR (400 MHz, CDCl3) d:
1.45–1.55 (m, 2H), 1.57–1.72 (m, 6H), 1.74–1.87 (m, 4H), 2.08–
2.13 (m, 2H), 3.15–3.21 (m, 2H), 3.25–3.30 (m, 2H), 3.69 (s,
2H), 3.70–3.75 (m, 2H), 4.20–4.32 (m, 1H), 6.87 (d, J = 7.8 Hz,
1H), 7.21–7.29 (m, 1H), 7.44 (dd, J = 9.8, 2.4 Hz, 1H), 7.59 (d,
J = 8.3 Hz, 1H), 7.71 (d, J = 6.9 Hz, 1H), 7.73–7.82 (m, 3H), 8.02
(d, J = 1.4 Hz, 1H), 8.19 (dd, J = 6.9, 1.4 Hz, 1H); MS (FAB) m/
z = 517 [M+H]+. Anal. Calcd for C30H33FN4O3ꢁ0.2CHCl3ꢁ0.25H2O:
C, 66.56; H, 6.23; N, 10.28; F, 3.49; Cl, 3.90. Found: C, 66.40; H,
6.47; N, 9.99; F, 3.29; Cl, 3.53.
5.1.37. 3-(Piperidin-1-ylcarbonyl)isonicotinic acid (26)
A
solution of pyridine-3,4-dicarboxylic acid (25) (3.00 g,
18.0 mmol) in Ac2O (12 mL) was stirred at reflux for 1.5 h. The
mixture was filtered and the filtrate was concentrated in vacuo
to yield pyridine-3,4-dicarboxylic anhydride. To an ice-cooled
solution of above-obtained compound in THF (60 mL) was added
piperidine (3.9 mL, 39 mmol), and the mixture was stirred at
room temperature for 17 h. The mixture was concentrated in va-
cuo. The residue was then diluted with satd NaHCO3 aq and ex-
tracted with CHCl3 three times. The organic layer was acidified
by 6 M HCl aq and concentrated in vacuo, and the solid was di-
luted with hot EtOH, and insoluble matter was removed by filtra-
tion, and the filtrate was concentrated in vacuo. The residue was
crystallized from hexane–EtOAc to yield the mixture of com-
5.1.41. 4-(Piperidin-1-ylcarbonyl)nicotinic acid (27)
A
mixture of 3113 (500 mg, 2.76 mmol) and piperidine
(1.64 mL, 16.6 mmol) was stirred at 70 °C for 1 d. The mixture
was diluted with NaCl–saturated 5% citric acid aq, extracted
with CHCl3, and the organic layer was dried over Na2SO4, fil-
tered, and concentrated in vacuo to yield 27 (382 mg, 59%) as
a brown foam. 1H NMR (400 MHz, DMSO-d6) d: 1.35–1.47 (m,
2H), 1.50–1.63 (m, 4H), 2.97–3.04 (m, 4H), 7.29 (d, J = 5.1 Hz,
1H), 8.72 (d, J = 5.1 Hz, 1H), 9.02 (s, 1H); MS (FAB) m/z = 233
[M+H]+.

I. Sato et al. / Bioorg. Med. Chem. 16 (2008) 8607–8618
8617
5.1.42. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-azabi-
cyclo[3.2.1]oct-3-yl}-4-(piperidin-1-ylcarbonyl)nicotinamide (32)
Compound 32 was prepared from 5 and 27 in a manner similar
to that described for compound 6a, with a yield of 6% as a colorless
amorphous solid. 1H NMR (400 MHz, DMSO-d6) d: 1.38–1.75 (m,
12H), 2.01–2.09 (m, 2H), 3.00–3.09 (m, 2H), 3.19–3.24 (m, 2H),
3.45–6.59 (m, 2H), 3.71 (s, 2H), 4.05–4.15 (m, 1H), 7.31 (d,
J = 5.4 Hz, 1H), 7.41 (ddd, J = 9.3, 8.8, 2.6 Hz, 1H), 7.61 (d,
J = 8.3 Hz, 1H), 7.68 (dd, J = 10.3, 2.5 Hz, 1H), 7.85–7.90 (m, 2H),
7.96 (dd, J = 8.8, 5.9 Hz, 1H), 8.35 (d, J = 7.8 Hz, 1H), 8.66
(d, J = 5.9 Hz, 1H), 8.79 (s, 1H); HR-MS calcd for C30H33FN4O2
m/z = 501.2666 [M+H]+. Found: 501.2663.
1 mM MgCl2, 2.5 mM CaCl2, and 5.5 mM glucose. The cell suspen-
sion (490 L) was transferred into cuvettes and placed under con-
l
stant agitation. Changes in fluorescence were monitored at 25 °C
using a spectrophotometer at excitation wavelengths of 340 nM
and 380 nM and at an emission wavelength of 510 nM. Calculation
of Ca2+ concentration was performed using the Kd for the Ca2+
binding of 224 nM. The antagonist was dissolved in 100% DMSO
solution (1 lL) and added to the cuvette 1 min prior to the addition
of eotaxin (final concentration of 50 ng/mL). Linear regression
analysis using EXSAS-STAT was used to calculate the IC50 values.
Values are reported as means SEM of triplicate experiments.
5.2.2. CYP2D6 inhibition
5.1.43. Methyl 3-(piperidin-1-ylcarbonyl)pyridine-2-
carboxylate (34)
This test was carried out in accordance with the method of Cres-
pi et al.15 Using a 96-well plate, 3-[2-(N,N-diethyl-N-methyl-
To a solution of 3314 (414 mg, 2.29 mmol) in CH2Cl2 (5 mL) were
added piperidine (0.25 mL, 2.50 mmol), HOBt (340 mg,
2.51 mmol), and WSCꢁHCl (570 mg, 2.97 mmol), and the mixture
was stirred at room temperature for 3 h. The mixture was then par-
titioned between EtOAc and satd NaHCO3 aq, and the organic layer
was washed with brine, dried over Na2SO4, filtered, and concen-
trated in vacuo. The residue was purified by silica gel column chro-
matography (CHCl3/MeOH = 200:1) to yield 34 (423 mg, 74%) as a
slightly yellow oil. 1H NMR (400 MHz, CDCl3) d: 1.46–1.55 (m,
2H), 1.65–1.74 (m, 4H), 3.12–3.16 (m, 2H), 3.75–3.79 (m, 2H),
3.99 (s, 3H), 7.52 (dd, J = 8.0, 4.8 Hz, 1H), 7.68 (dd, J = 8.0, 1.6 Hz,
1H), 8.75 (dd, J = 4.8, 1.6 Hz, 1H); MS (FAB) m/z = 249 [M+H]+.
amino)ethyl]-7-methoxy-4-methylcoumarin (1.5
compound (from 0.031 to 50 M), and the enzyme (5 nmol) were
incubated at 37 °C for 20 min in 200 L in total volume of
100 mM phosphate buffer (pH 7.4) containing 8.2
M NADP+,
0.41 mM glucose-6-phosphate, 0.41 mM MgCl2 and 0.4 U/mL glu-
cose-6-phosphate dehydrogenase. Thereafter, the reaction was
stopped by adding 0.5 M 2-amino-2-hydroxymethyl-1,3-propane-
diol aqueous solution containing 80% acetonitrile, and the fluores-
cence intensity (excitation wavelength; 390 nM, fluorescence
wavelength; 460 nM) was measured using a fluorescence plate
reader. The inhibition ratio was calculated based on the following
formula, and concentration of each test compound by which the
inhibition ratio becomes 50% (IC50) was obtained.
lM), each test
l
l
l
5.1.44. 3-(Piperidin-1-ylcarbonyl)pyridine-2-carboxylic acid
(35)
The inhibition ratio ð%Þ ¼ 100 ꢀ ðC1 ꢀ B1Þ=ðC0 ꢀ B1Þ ꢂ 100
To a solution of 34 (396 mg, 1.59 mmol) in MeOH (4 mL) was
added 1 M NaOH aq (2.5 mL, 2.50 mmol), and the mixture was stir-
red at room temperature for 1 d. The mixture was then concen-
trated in vacuo, and the residue was partitioned between Et2O
and H2O, and the aqueous layer was acidified with 1 M HCl aq
(pH 5), and concentrated in vacuo. The solid was diluted with
EtOH, and insoluble matter was removed by filtration, and the fil-
trate was concentrated in vacuo to yield 35 (373 mg, quant.) as a
slightly brown amorphous solid. 1H NMR (400 MHz, DMSO-d6) d:
1.32–1.60 (m, 6H), 2.95–3.03 (m, 2H), 3.40–3.60 (m, 2H), 7.31–
7.39 (m, 1H), 7.51–7.56 (m, 1H), 8.46 (m, 1H); MS (FAB) m/
z = 233 [M+H]+.
C1: Fluorescence intensity in the presence of test compound
having known concentration, enzyme, and substrate.
C0: Fluorescence intensity in the absence of test compound and
in the presence of enzyme and substrate.
B1: Fluorescence intensity of blank well.
Acknowledgments
The authors wish to offer their deep thanks to the staff of the
Division of Analysis & Pharmacokinetics Research Laboratories
for their help with the evaluation of CYP2D6 inhibitory activity
as well as the elemental analysis and spectral measurements.
5.1.45. N-{(3-exo)-8-[(6-Fluoro-2-naphthyl)methyl]-8-
azabicyclo[3.2.1]oct-3-yl}-3-(piperidin-1-ylcarbonyl)pyridine-
2-carboxamide (36)
References and notes
Compound 36 was prepared from 5 and 35 in a manner similar
to that described for compound 6a, with a yield of 52% as a slightly
brown amorphous solid. 1H NMR (400 MHz, DMSO-d6) d: 1.25–
1.75 (m, 10H), 1.79–1.88 (m, 2H), 2.00–2.08 (m, 2H), 2.91–3.05
(m, 2H), 3.18–3.23 (m, 2H), 3.25–3.41 (m, 2H), 3.78 (s, 2H), 4.10–
4.21 (m, 1H), 7.40 (ddd, J = 8.8, 8.8, 2.5 Hz, 1H), 7.59–7.64 (m,
2H), 7.67 (dd, J = 10.2, 2.5 Hz, 1H), 7.75 (dd, J = 7.8, 1.5 Hz, 1H),
7.85–7.92 (m, 2H), 7.96 (dd, J = 8.3, 5.8 Hz, 1H), 8.50 (d, J = 8.8 Hz,
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Calcd for C30H33FN4O2ꢁ0.5H2O: C, 70.70; H, 6.72; N, 10.99; F, 3.73.
Found: C, 70.82; H, 6.71; N, 11.04; F, 3.70.
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