Discovery of selective indole-based prostaglandin D2 receptor antagonist

Article abstract of DOI:10.1016/j.bmc.2011.06.014

A series of N-benzoyl-2-methylindole-3-acetic acids were synthesized and biologically evaluated as prostaglandin (PG) D₂ receptor antagonists. Some of the selected compounds significantly inhibited OVA-induced vascular permeability in guinea pig conjunctiva after oral dosing. Structure-activity relationship study is presented.

Full text of DOI:10.1016/j.bmc.2011.06.014

Bioorganic & Medicinal Chemistry 19 (2011) 4574–4588
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Discovery of selective indole-based prostaglandin D2 receptor antagonist
Maki Iwahashi a, , Atsushi Shimabukuro a, Takahiro Onoda b, Yoko Matsunaga a, Yutaka Okada a,
⇑
Ryoji Matsumoto c, Fumio Nambu a, Hisao Nakai a, Masaaki Toda a
a Minase Research Institute, Ono Pharmaceutical Co., Ltd, 3-1-1 Sakurai, Shimamoto, Mishima, Osaka 618-8585, Japan
b Fukui Research Institute, Ono Pharmaceutical Co., Ltd, 1-5-2 Technoport, Yamagishi, Mikuni, Sakai, Fukui 913-8538, Japan
c Ono Pharmaceutical Co., Ltd, 2-1-5 Doshomachi, Chuoh, Osaka 541-8526, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of N-benzoyl-2-methylindole-3-acetic acids were synthesized and biologically evaluated as
prostaglandin (PG) D2 receptor antagonists. Some of the selected compounds significantly inhibited
OVA-induced vascular permeability in guinea pig conjunctiva after oral dosing. Structure–activity
relationship study is presented.
Received 10 May 2011
Revised 3 June 2011
Accepted 3 June 2011
Available online 16 June 2011
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Prostaglandin
DP receptor
Antagonist
Indole
1. Introduction
CO2H
Coleman et al.1 proposed the existence of specific receptors for
thromboxane (TX), prostaglandin (PG)I2, PGE2, PGF2 , and PGD2
a
Me
N
such as TP, IP, EP, FP, and DP receptors, respectively. Among them,
the DP receptor is the most recently cloned prostanoid receptor
and perhaps the least characterized.2 PGD2 is the major prostanoid
released from mast cells after challenge with IgE3 and it has also
been shown to affect the sleep cycle4 and body temperature.5
The discovery of a DP selective receptor antagonist6,7 seems to of-
fer significant advantages, for investigating the role of this receptor
in the various pathologies described above. However, the role of
the DP receptor remains unclear because of the lack of potent
and subtype-selective ligands.
Several patent applications claim the use of indolyl acetic acid
derivatives as orphan receptor CRTH2 antagonists.8 Blockade of
the above-described receptors is known to inhibit much of the pro-
inflammatory effects of PGD2.9
PGD2 receptor antagonists might have therapeutic potential for
allergic disorders because PGD2 is considered to play an important
role in various allergic diseases such as allergic rhinitis,10 atopic
asthma,3b allergic conjunctivitis,11 and atopic dermatitis.12
In our previous paper,7d we reported discovery of N-benzoyl-2-
methylindole-4-acetic acid analog 2 as an optimized human DP
(hDP) receptor antagonist starting with chemical modification of
indomethacin 1 (Fig. 1). Synthetic study of the indole-based hDP
O
O
MeO
O
CO2H
Me
N
Me
2
N
O
CO2H
Me
Cl
1 (Indomethacin)
N
O
Me
O
N
11
Et
Figure 1. Molecular design of indole-based DP antagonists.
antagonist 2 prompted us to carry out molecular design of more
cost-effective structure as a chemical lead of a drug candidate be-
cause of the synthetic difficulty of ‘N-benzoyl-2-methylindole-
4-acetic acid’ scaffold. Besides, low hIP/DP receptor selectivity of
2 (Table 1) was considered a risk factor that might cause serious
side effects such as platelet aggregation. To solve the problems
described above, further optimization of the indole-3-acetic
⇑
Corresponding author. Tel.: +81 75 961 1151; fax: +81 75 962 9314.
E-mail address: iwahashi@ono.co.jp (M. Iwahashi).
0968-0896/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2011.06.014

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4575
Table 1
Effect of the 5-substituent of the indole moiety on the activity profiles
X
6
5
4
CO2H
Me
N
7
O
O
O
N
Me
a
Compound
X
Binding Ki (nM)
IC50 (nM)
mEP1
mEP2
mEP3
mEP4
mFP
hTP
hIP
mDP
mDP
1
2
>10,000
6200
>10,000
5200
>10,000
>10,000
>10,000
>10,000
99
210
340
330
>10,000
1700
1700
2800
>10,000
1900
>10,000
>10,000
>10,000
3600
>10,000
>10,000
3000
NTb
NTb
260
>10,000
>10,000
NTb
NTb
37
200
1100
49
>10,000
16
7.7
28
29
NTb
2.4
NTb
4.3
3.1
1.8
NTb
3100
2100
1700
2200
1700
NTb
3a
3b
3c
3d
3e
H
Me
Cl
F
250
310
>10,000
320
1000
5.7
9.9
OMe
2500
NTb
a
IC50 (nM): mDP receptor antagonist activity.
NT: not tested.
b
Table 2
acid-based chemical lead was carried out. We here report the
discovery process of new indole-based mouse DP (mDP) antagonist
11 starting from chemical modification of indomethacin 1, which
could be an excellent chemical lead for developing an orally active
drug candidate.
Effect of substitution of the N-benzoyl moiety on the activity profiles
X
CO2H
Me
N
2
2. Chemistry
3
O
O
Synthesis of the test compounds listed in Tables 1–3 is outlined
in Schemes 1 and 2. Synthesis of 3a–e, 4a–i and 5a–g is described
in Scheme 1. O-Alkylation of appropriately substituted methyl
4-hydroxybenzoates 18a–j with a tosylate 13, prepared from
N-methyl benzomorpholine 12,7d afforded 14a–j, alkaline hydroly-
sis of which provided the corresponding carboxylic acids 15a–j.
The carboxylic acids 15a–j were converted to the corresponding
acid chlorides 16a–j, which were used for the next N-acylation
reaction of the benzyl 2-methylindole-3-acetic acetates 19a–e. N-
Acylation of 19a–e, which were prepared from the corresponding
para-substituted phenylhyrazine hydrochloride and levulinic acid
by Fisher’s indole synthesis, with an optional acid chloride selected
among 16a–j under the reported reaction condition7d afforded
17a–u. Deprotection of 17a–e, 17f–n, and 17o–u by catalytic
hydrogenation afforded 3a–e, 4a–i, and 5a–g, respectively. Synthe-
sis of 6–11 is described in Scheme 2a. N-Acylation of 19a with
4-acetoxy-2-methylbenzoyl chloride in the presence of 20 M
sodium hydroxide aq gave 20, which was transformed to the
corresponding carboxylic acid 21 by catalytic hydrogenation.
Treatment of carboxylic acid 21 with a resin in the presence of
diisopropylethylamine afforded resin-supported intermediate 22,
deacetylation of which with piperidine provided phenol 23. Mits-
unobu reaction13 of alcohols 24–297d,14 with the resin-supported
phenol 23 followed by acidic cleavage from the supporting resin15
resulted in the production of 6–11. Among the bicyclic alcohols,
25a–b were prepared as described in Scheme 2b. O-Alkylation of
30a–b with (S)-(+)-glycidyl 3-nitrobenzenesulfonate in the pres-
ence of cesium carbonate afforded epoxides 31a–b. Intramolecular
ring opening reaction of epoxides 31a–b by addition of n-butyllith-
ium resulted in dihydrofurans 25a–b. As shown in Scheme 2c,
racemic 28b and optically active 29 were prepared from commer-
cially available 32a and 32b, respectively. Concurrent N-ethylation
6
O
Y
5
N
Me
a
Compound
X
Y
Binding Ki (nM) IC50 (nM) Binding Ki (nM)
mDP
hIP
3a
4a
4b
4c
4d
4e
4f
4g
4h
4i
H
H
H
H
H
H
H
H
H
H
H
7.7
3.5
4.4
3.8
140
380
240
340
NTb
1.1
2.2
200
130
92
110
72
91
210
430
98
2-Me
2-Cl
2-F
3-Me
3-Cl
3-F
2.5
NTb
NTb
NTb
NTb
NTb
NTb
3-OMe
2,3-Me 380
2,5-Me 360
94
3d
5a
5b
5c
5d
5e
5f
F
F
F
F
F
F
F
F
H
5.7
2.7
5.2
3.8
59
1.8
3.2
1.6
0.7
42
68
32
2.8
320
130
200
320
160
1500
230
93
2-Me
2-Cl
2-F
3-Me
3-OMe
NTb
2,3-Me NTb
2,5-Me 14
5g
a
IC50 (nM): mDP receptor antagonist activity.
NT: not tested.
b
and O-ethylation of 32a with ethyl iodide in the presence of cesium
carbonate afforded 33a, reduction of which with lithium aluminum
hydride gave 28b. According to the same procedure as described
for the preparation of 28b from 32a, 29 was prepared from the cor-
responding 2R-enantiomer 32b.

4576
M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
Table 3
hIP receptor affinity relative to 2. Introduction of 5-methyl and
5-chloro residues into the 2-methyl indole-3-acetic acid scaffold
afforded 3b and 3c, respectively, with nearly equipotent binding
affinity for DP receptor and 30-fold less potent and equipotent
hIP receptor affinity, respectively. Introduction of 5-fluoro and 5-
methoxy residues into the 2-methyl indole-3-acetic acid scaffold
afforded 3d and 3e, respectively, with slightly increased and nearly
equipotent binding affinity.
Effect of chemical modification of N-methyl benzomorpholine moiety on the activity
profiles
CO2H
Me
N
O
Me
R
O
Among them, 3a, 3d and 3e tended to show slightly more DP
selectivity than 2.
a
Compound
R
Binding
IC50
Binding
Regarding hIP receptor affinity, compound 3c afforded nearly
the same hIP receptor affinity as 2. Compounds 3a–b and 3d–e
showed less potent hIP receptor affinity relative to 2. 4-Substituted
and 6-substituted analogs were also synthesized but they were not
evaluated as a single product because of the difficulty in their sep-
aration. 7-Substituted indole analogs were not synthesized be-
cause of predicted difficulty of their N-acylation. As a result,
most of the tested compounds 3b–c and 3d showed potent antag-
onist activity.
Ki (nM)
(nM)
Ki (nM)
mDP
hIP
O
S
R
4a
6
3.5
22
1.1
3.8
130
570
N
Me
O
Table 2 displays the effect of simultaneous substitution of the
indole moiety and phenyl moiety of the N-benzoyl residue on the
activity profiles. As illustrated by the biological results of 4a–c,
2-methyl, 2-chloro, and 2-fluoro substitutions resulted in retention
of potent DP receptor affinity relative to unsubstituted 3a while as
illustrated by 4d–i, 3-methyl, 3-chloro, 3-fluoro, 3-methoxy, 2,
3-dimethyl, and 2,5-dimethyl substitution resulted in a decrease
of DP receptor affinity. A series of 5-fluoro indole analogs were
synthesized and evaluated. As illustrated by 5a–c, introduction of
2-methyl, 2-chloro, and 2-fluoro residues was effective for reten-
tion of potent DP receptor affinity whereas introduction of
3-methyl, 3-methoxy, and 2,3-dimethylbenzoyl residues resulted
in reduced activities relative to 3d as illustrated by 5d–f. Com-
pounds 4a–c, 5a–c, and 5g showed very potent antagonist activity
as expected from their corresponding Ki values. Regarding hIP
receptor affinity, compounds 4b, 4d–e, 4h–i, and 5g tended to
show increased affinity relative to 3a while they still retained
selectivity (Ki values, hIP/mDP). As a result, the 2-substituted
N-benzoyl analogs tended to show increased activity versus
3-substituted N-benzoyl analogs whereas 4i and 5g showed unex-
pected results in their binding affinity.
O
X
5
a X = H
b X = F
43
72
2100
520
7
5.4
1.8
R
S
O
O
8
9
7.1
7.0
43
280
O
NTb
3000
O
a R = Me
b R = Et
NTb
22
6.2
3.1
>10,000
5100
10
11
N
N
R
R
4.8
2.2
2200
Et
a
IC50 (nM): mDP receptor antagonist activity.
NT: not tested.
b
3. Results and discussion
Table 3 displays the effect of chemical modification of N-methyl
benzomorpholine moiety of 4a, which is the partial structure of the
previously optimized 2-methylindole-4-acetic acid analog 2.7d
Transformation of the terminal N-methyl benzomorpholine moiety
to a benzodioxane moiety provided 6 with 6.3-fold reduction of DP
receptor affinity. Replacement of the benzomorpholine moiety of
4a with benzofuran-3-yl moiety provided 7a with 12-fold
reduction of DP receptor affinity, whereas introduction of 5-fluoro
residue into the benzofuran moiety afforded 7b with nearly
equipotent receptor affinity and 4.1-fold less potent hIP receptor
affinity relative to 4a. Replacement of the benzomorpholine moiety
of 4a with benzofuran-2-yl moiety afforded 8 with nearly retained
DP receptor affinity and nearly 2-fold less potent hIP receptor
affinity. Introduction of another oxygen into the benzofuran
moiety of 8 afforded 9 with a reduction of antagonist activity.
Replacement of the oxygen atom of the benzofuran moiety of 8
with N-methyl moiety afforded 10a with nearly equipotent antag-
onist activity. Replacement of the oxygen moiety of 8 with N-ethyl
moiety afforded 10b tended to increase DP receptor antagonist
activity. Based on the information mentioned above, an optically
active form 11 was prepared and evaluated for DP receptor affinity
and hIP receptor affinity. The compound 11 showed potent DP
receptor affinity and equipotent antagonist activity relative to 4a.
Besides, DP receptor affinity of 11 showed remarkable improve-
ment in selectivity for the hIP receptor (11, hIP/DP = 458; 2, hIP/
DP = 2.3).
The test compounds listed in Tables 1–3 were all tested for inhi-
bition of the specific binding of a radiolabeled ligand, [3H]PGD2, to
membrane fractions prepared from cells stably expressing mDP
receptor. They were also evaluated for their potency to antagonize
mDP receptors by measuring PGD2-stimulated changes in intracel-
lular second messenger cAMP as an indicator of receptor function.
The mDP antagonism was measured in the presence of 0.1% bovine
serum albumin (BSA) for the discovery of potent antagonists in
protein-rich in vivo animal models. Because of their close homol-
ogy to human receptors, all the DP receptor affinities and antago-
nist activities are assayed by mouse receptor unless otherwise
noted. Our purpose was to identify more DP-selective antagonist
compared with 2, which showed relatively higher IP receptor affin-
ity. As such, our focus was placed on the binding affinity (Ki) of the
newly synthesized compounds for mEP1–4, mFP, hTP, and hIP in
addition to their Ki values for the mDP receptor. Compounds
3a–e (Table 1), 4a–i (Table 2), and 5a–g (Table 2), in which para-
position of the N-benzoyl residue is substituted with the para-N-
methyl benzomorpholinomethoxy moiety, were synthesized and
evaluated. On the basis of the previously optimized structure 2,7d
a series of 2R-enantiomers 3a–e were synthesized and evaluated.
Table 1 displays the effect of the 5-substituent of the indole
moiety on the activity profiles. Compound 3a demonstrated
slightly more potent DP receptor affinity and 5.4-fold less potent

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4577
R
O
b
RO
O
O
X
MeO2C
N
Me
N
Me
OH
X
18a-j
X = H
X = 2-Me
X = 2-Cl
X = 2-F
X = 3-Me
X = 3-Cl
X = 3-F
X = 3-OMe
X = 2,3-Me
X = 2,5-Me
12 : R = H
13 : R = Ts
a
14a-j : R = CO2Me
15a-j : R = CO2H
16a-j : R = COCl
c
a :
d
b :
c :
d :
e :
f :
g :
h :
i :
j :
Y
CO2Bn
Me
N
f
e
3a-e (Table 1),4a-i (Table 2),
5a-g (Table 2)
O
Y
O
CO2Bn
Me
O
X
N
Me
N
H
17a-u
19a-e
X
X
Y
H
Me
Cl
F
Y
a :
a :
b :
c :
d :
e :
H
H
H
H
H
Y = H
Y = Me
Y = Cl
Y = F
l : 3-OMe
m :2,3-Me
n : 2,5-Me
o : 2-Me
p : 2-Cl
H
H
H
F
F
F
F
F
F
F
b :
c :
d :
e :
OMe
H
Y = OMe
f : 2-Me
g : 2-Cl
h : 2-F
i : 3-Me
j : 3-Cl
k : 3-F
q : 2-F
H
r : 3-Me
s : 3-OMe
t : 2,3-Me
u : 2,5-Me
H
H
H
H
Scheme 1. Synthesis of 3a–e, 4a–i and 5a–g. Reagents: (a) TsCl, Et3N, DMAP, THF; (b) 18a–j, K2CO3, DMF; (c) NaOH, MeOH, THF; (d) (COCl)2, DMF, DME; (e) NaOH, TBACl,
CH2Cl2; or Et3N, DMAP, CH3CN, EtOAc; (f) H2, Pd(OH)2–C, EtOAc.
As shown in Table 4, representative compounds 4a, 4b, 5b, and
11 were evaluated for their potency to inhibit OVA-induced vascu-
lar permeability in guinea pig conjunctiva. Among the tested com-
pounds, 4b, 5b, and 11 showed significant inhibition after their oral
dosing (3, 3, and 1 mg/kg, respectively) whereas 4a (3 mg/kg) did
not show significant DP receptor antagonist activity because of
the presumed PK problem and/or species difference of DP receptor
affinity between mouse and guinea pig. Weaker in vivo potency of
4a relative to 4b, 5b and 11 was considered to be due to its less po-
tent antagonist activity and/or its lower blood level in guinea pig.
Table 5 displays activity profiles of 4a, 4b, 5b, and 11 including
other mEP receptors, mFP receptor, hTP receptor, hIP receptor,
mDP receptor, and mDP antagonist activity. As shown in Table 5,
4a, 4b, 5b, and 11 demonstrated much better IP/DP selectivity rel-
ative to the previously reported compound 2 (Table 1). Especially,
compound 11 showed better selectivity compared with 2 because
of its markedly reduced hIP receptor affinity and retained mDP
receptor affinity. Compounds 4a, 4b, and 5b also showed a similar
tendency as 11 regarding hIP/DP selectivity although reduction of
their hIP receptor affinity was not so much as that of 11. Based
on these results, more receptor-selective indole-based DP antago-
nist relative to 2 was discovered starting with the chemical
modification of indomethacin 1.
guinea pig conjunctiva. The newly identified DP antagonist 11
showed remarkable improvement of hIP/mDP selectivity in its
receptor affinity compared with 2.
4. Experimental
4.1. General directions
Analytical samples were homogeneous as confirmed by TLC,
and afforded spectroscopic results consistent with the assigned
structures. Proton nuclear magnetic resonance spectra (1H NMR)
were taken on a Varian Mercury 300 spectrometer or Varian
GEMINI-200 or VXR-200s spectrometer using deuterated chloro-
form (CDCl3) or deuterated methanol (CD3OD) or deuterated
dimethylsulfoxide (DMSO-d6) as the solvent. Fast atom bombard-
ment mass spectra (FAB-MS) and electron ionization (EI) were
obtained on a JEOL JMS-DX303HF spectrometer. The matrix as-
sisted laser desorption ionization-time of flight high-resolution
mass spectra (MALDI-TOF, HRMS) were obtained on a PerSeptive
Voyager Elite spectrometer. Atmospheric pressure chemical ioniza-
tion (APCI) was determined on a HITACHI M1200H spectrometer.
Infrared spectra (IR) were measured on a Perkin-Elmer FT-IR
1760X spectrometer. Melting points and results of elemental anal-
yses were uncorrected. Optical rotations were measured in a JASCO
DIP-1000 digital polarimeter. Column chromatography was carried
out on silica gel [Merck Silica Gel 60 (0.063–0.200 mm), Wako gel
C200 or Fuji Silysia BW235]. Thin layer chromatography was
performed on silica gel (Merck TLC or HPTLC plates, Silica Gel 60
In summary, we have discovered two series of indole-based DP
antagonists possessing 2-methylindole-4-acetic acid7 and 2-
methylindole-3-acetic acid as scaffolds starting with chemical
modification of indomethacin 1. An optimized compound 11
demonstrated efficacy in OVA-induced vascular permeability in

4578
M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
CO2R
Me
c
N
a
19a
O
Me
b
OAc
20 : R = Bn
21 : R = H
O
O
resin
R
1) e 24-29
Cl
Me
N
6-11 (Table 3)
Cl
polymer
resin-Cl :
2) f
O
Me
O
22 : R = Ac
23 : R = H
d
HO
O
O
HO
HO
X
O
O
2615(b)
25a-b
2415(a)
a : X = H
b : X = F
O
HO
HO
HO
O
N
R
N
Et
28a-b
29
277(d)
a
b
7(d) : R = Me
: R = Et
Scheme 2a. Synthesis of 6–11. Reagents: (a) 4-acetoxy-2-methylbenzoyl chloride, NaOH, TBACl, CH2Cl2; (b) H2, Pd(OH)2–C, EtOAc; (c) resin-Cl, diisopropylethylamine,
CH2Cl2; (d) piperidine, CH2Cl2; (e) DEAD, PPh3, CH2Cl2; (f) AcOH, CF3CH2OH, CH2Cl2.
F254). The following abbreviations for solvents and reagents are
used; tetrahydrofuran (THF), ethyl acetate (EtOAc), dimethylform-
amide (DMF), dichloromethane (CH2Cl2), chloroform (CHCl3),
methanol (MeOH), acetic acid (AcOH), triethylamine (TEA),
diisopropylethylamine (DIPEA), 4-dimethylaminopyridine (DMAP),
tert-butyl methyl ether (MTBE), 2-propanol (IPA), 1,2-dime-
thoxyethane (DME), tetrabutylammonium chloride (TBACl),
divinylbenzene (DVB), p-toluenesulfonyl chloride (TsCl), and
diethylazodi-carboxylate (DEAD).
DMAP (0.29 g, 2.38 mmol) in THF (1.5 mL) and a solution of TsCl
(5.45 g, 28.6 mmol) in THF (18 mL) at 0 °C. After stirring for 4 h
at room temperature, the reaction mixture was quenched with
water and extracted with MTBE (ꢀ2). The combined organic layers
were washed with 1 M HCl aq, water, NaHCO3 aq, brine, dried over
MgSO4 and concentrated in vacuo. The resulting residue was puri-
fied by recrystalization from MTBE (7.9 mL) and IPA (39 mL) to
yield 13 (5.53 g, 70%); TLC Rf = 0.45 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3)
d 7.81 (d, J = 8.4 Hz, 2H), 7.35 (d,
J = 8.4 Hz, 2H), 6.87–6.81 (m, 1H), 6.70–6.61 (m, 3H), 4.50–4.42
(m, 1H), 4.23–4.12 (m, 2H), 3.24 (dd, J = 11.7, 2.7 Hz, 1H), 3.09
(dd, J = 11.7, 6.3 Hz, 1H), 2.83 (s, 3H), 2.45 (s, 3H).
4.2. [(2S)-4-Methyl-3,4-dihydro-2H-1,4-benzoxazin-2-yl]methyl
4-methylbenzenesulfonate (13)
To a stirred solution of 12 (4.27 g, 23.8 mmol) in THF (10.5 mL)
were successively added a solution of TEA (8.0 mL, 57.2 mmol) and
4.3. General procedure for the preparation of 16a–j
4.3.1. Methyl 4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14a)
Br
X
To a stirred solution of methyl-4-hydroxybenzoic acid (2.43 g,
16.0 mmol) and 13 (5.43 g, 16.3 mmol) in DMF (10.8 mL) was
added K2CO3 (4.94 g, 35.2 mmol). After stirring for 10 h at 80 °C,
the reaction mixture was quenched with water and extracted with
MTBE (ꢀ2). The combined organic layers were washed with water,
brine, dried over Na2SO4 and evaporated to give 14a, which was
used for the next reaction without further purification; TLC
Rf = 0.46 (n-hexane/EtOAc, 7:3); 1H NMR (300 MHz, CDCl3) d 7.99
(d, J = 8.7 Hz, 2H), 6.96 (d, J = 8.7 Hz, 2H), 6.92–6.80 (m, 2H),
6.74–6.66 (m, 2H), 4.70–4.61 (m, 1H), 4.28 (dd, J = 9.6, 4.8 Hz,
1H), 4.17 (dd, J = 9.6, 6.6 Hz, 1H), 3.89 (s, 3H), 3.39 (dd, J = 11.7,
2.7 Hz, 1H), 3.26 (dd, J = 11.7, 6.6 Hz, 1H), 2.90 (s, 3H).
Br
X
a
b
25a-b
O
HO
H
O
30a-b
31a-b
a : X = H
b : X = F
Scheme 2b. Synthesis of 25a–b. Reagents: (a) (S)-(+)-glycidyl 3-nitrobenzenesulf-
onate, Cs2CO3, DMF; (b) n-BuLi, THF.
2
2
HO2C
EtO2C
Et
a
b
28b and 29
N
H
N
4.3.2. 4-{[(2S)-4-Methyl-3,4-dihydro-2H-1,4-benzoxazin-2-yl]
methoxy}benzoic acid (15a)
To a stirred solution of 14a in MeOH (25 mL)–THF (25 mL) was
added 5 M NaOH aq (25 mL). After stirring for 3 h at 50 °C, the reac-
tion mixture was diluted with water and extracted with MTBE. The
33a (2RS
33b (2R)
)
32a (2RS
32b (2R)
)
Scheme 2c. Synthesis of 28b and 29. Reagents: (a) ethyl iodide, Cs2CO3, DMF; (b)
LiAlH4, THF.

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4579
Table 4
4.17 (dd, J = 9.6, 6.3 Hz, 1H), 3.39 (dd, J = 11.7, 3.0 Hz, 1H), 3.25 (dd,
J = 11.7, 6.3 Hz, 1H), 2.91 (s, 3H), 2.63 (s, 3H).
Inhibitory effects of 4a, 4b, 5b and 11 on OVA-induced vascular permeability in
guinea pig conjunctiva (n = 8)
Compound
Dose (mg/kg, po)
% Inhibition
4.3.6. 2-Methyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16b)
2
1
3
3
3
3
1
23
41a
29
1H NMR (300 MHz, CDCl3) d 8.26 (d, J = 8.7 Hz, 1H), 7.10–6.80
(m, 6H), 4.86–4.78 (m, 1H), 4.40–4.28 (m, 2H), 3.58 (dd, J = 11.7,
2.7 Hz, 1H), 3.45 (dd, J = 11.7, 2.7 Hz, 1H), 3.08 (s, 3H), 2.57 (s, 3H).
4a
4b
5b
11
48a
52a
42a
68
4.3.7. Methyl 2-chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14c)
Pyrilamine
1 (mg/kg, iv)
Inhibition of increase in conjunctival vascular permeability caused by topical
application of OVA (1%, 20 L/eye) in actively sensitized guinea pigs. All antagonists
were orally administered 1 h before the antigen challenge.
TLC Rf = 0.58 (n-hexane/EtOAc, 7:3); 1H NMR (300 MHz, CDCl3)
d 7.88 (d, J = 8.7 Hz, 1H), 7.02 (d, J = 2.7 Hz, 1H), 6.92–6.80 (m, 3H),
6.73–6.65 (m, 2H), 4.69–4.60 (m, 1H), 4.25 (dd, J = 9.6, 5.1 Hz, 1H),
4.16 (dd, J = 9.6, 6.3 Hz, 1H), 3.90 (s, 3H), 3.37 (dd, J = 11.7, 3.0 Hz,
1H), 3.24 (dd, J = 11.7, 6.3 Hz, 1H), 2.91 (s, 3H).
l
a
0.05 versus control with Student’s t-test.
aqueous layer was acidified with 2 M HCl aq and extracted with
EtOAc (ꢀ2). The combined organic layers were washed with water,
brine, dried over Na2SO4 and evaporated to yield 15a (4.26 g, 87%
in two steps); TLC Rf = 0.47 (CHCl3/MeOH, 9:1); 1H NMR
(300 MHz, CDCl3) d 8.06 (d, J = 8.7 Hz, 2H), 6.99 (d, J = 8.7 Hz, 2H),
6.92–6.80 (m, 2H), 6.74–6.66 (m, 2H), 4.72–4.62 (m, 1H), 4.30
(dd, J = 9.6, 5.1 Hz, 1H), 4.19 (dd, J = 9.6, 6.6 Hz, 1H), 3.40 (dd,
J = 11.4, 3.0 Hz, 1H), 3.27 (dd, J = 11.4, 6.9 Hz, 1H), 2.91 (s, 3H).
4.3.8. 2-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15c)
68% yield in two steps; TLC Rf = 0.43 (CHCl3/CH3OH, 9:1); 1H
NMR (300 MHz, CDCl3)
d 8.04 (d, J = 9.0 Hz, 1H), 7.05 (d,
J = 2.4 Hz, 1H), 6.94–6.80 (m, 3H), 6.75–6.67 (m, 2H), 4.70–4.61
(m, 1H), 4.27 (dd, J = 9.9, 5.4 Hz, 1H), 4.18 (dd, J = 9.9, 6.3 Hz, 1H),
3.35 (dd, J = 11.7, 3.0 Hz, 1H), 3.25 (dd, J = 11.7, 6.3 Hz, 1H), 2.91
(s, 3H).
4.3.3. 4-{[(2S)-4-Methyl-3,4-dihydro-2H-1,4-benzoxazin-2-yl]
methoxy}benzoyl chloride (16a)
To a stirred solution of 15a (120 mg, 0.4 mmol) in DME (5 mL)
were added (COCl)2 (0.09 mL, mmol) and DMF (0.3 lL,
0.004 mmol). After stirring for 1 h at 40 °C, the reaction mixture
was concentrated in vacuo to give 16a, which was used for the next
reaction without further purification; 1H NMR (300 MHz, CDCl3) d
8.10 (d, J = 9.0 Hz, 2H), 7.01 (d, J = 9.0 Hz, 2H), 6.98–6.80 (m, 4H),
4.80–4.70 (m, 1H), 4.34 (dd, J = 9.9, 4.5 Hz, 1H), 4.26 (dd, J = 9.9,
5.7 Hz, 1H), 3.47 (dd, J = 12.0, 2.7 Hz, 1H), 3.34 (dd, J = 12.0,
7.2 Hz, 1H), 2.98 (s, 3H).
4.3.9. 2-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16c)
1H NMR (300 MHz, CDCl3) d 8.21 (d, J = 9.3 Hz, 1H), 7.10–6.90
(m, 6H), 4.88–4.78 (m, 1H), 4.40–4.30 (m, 2H), 3.59 (dd, J = 12.3,
2.4 Hz, 1H), 3.43 (dd, J = 12.3, 8.4 Hz, 1H), 3.08 (s, 3H).
4.3.10. Methyl 2-fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14d)
TLC Rf = 0.36 (n-hexane/EtOAc, 3:1); 1H NMR (300 MHz, CDCl3)
d 7.90 (t, J = 8.4 Hz, 1H), 6.90–6.79 (m, 2H), 6.78–6.64 (m, 4H),
4.68–4.60 (m, 1H), 4.24 (dd, J = 9.6, 5.1 Hz, 1H), 4.15 (dd, J = 9.6,
6.3 Hz, 1H), 3.90 (s, 3H), 3.37 (dd, J = 11.7, 2.7 Hz, 1H), 3.24 (dd,
J = 11.7, 6.3 Hz, 1H), 2.90 (s, 3H).
According to the same procedure as described above for the
conversion of 13 to16a, 13 were converted to 16b–j.
4.3.4. Methyl 2-methyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14b)
4.3.11. 2-Fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15d)
TLC Rf = 0.61 (n-hexane/EtOAc, 7:3); 1H NMR (300 MHz, CDCl3)
d 7.93 (d, J = 9.3 Hz, 1H), 6.91–6.75 (m, 4H), 6.75–6.61 (m, 2H),
4.68–4.60 (m, 1H), 4.26 (dd, J = 9.6, 5.1 Hz, 1H), 4.15 (dd, J = 9.6,
6.6 Hz, 1H), 3.86 (s, 3H), 3.38 (dd, J = 11.4, 2.7 Hz, 1H), 3.24 (dd,
J = 11.4, 6.6 Hz, 1H), 2.90 (s, 3H), 2.60 (s, 3H).
96% yield in two steps; TLC Rf = 0.45 (CHCl3/CH3OH, 9:1); 1H
NMR (300 MHz, CDCl3) d 7.96 (t, J = 8.4 Hz, 1H), 6.92–6.75 (m,
3H), 6.75–6.65 (m, 3H), 4.70–4.61 (m, 1H), 4.26 (dd, J = 9.9,
5.1 Hz, 1H), 4.17 (dd, J = 9.9, 6.3 Hz, 1H), 3.37 (dd, J = 11.7, 2.7 Hz,
1H), 3.24 (dd, J = 11.7, 6.3 Hz, 1H), 2.90 (s, 3H).
4.3.5. 2-Methyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15b)
4.3.12. 2-Fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16d)
56% yield in two steps; TLC Rf = 0.27 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 8.05 (d, J = 9.6 Hz, 1H), 6.92–6.78 (m, 4H),
6.75–6.66 (m, 2H), 4.70–4.60 (m, 1H), 4.28 (dd, J = 9.6, 5.1 Hz, 1H),
1H NMR (300 MHz, CDCl3) d 8.13 (t, J = 8.4 Hz, 1H), 7.42–7.33
(m, 1H), 7.27–7.20 (m, 1H), 7.12–6.98 (m, 2H), 6.85 (dd, J = 8.7,
Table 5
Activity profiles of 4a, 4b, 5b and 11 on all the tested receptors
a
Compound
Binding Ki (nM)
IC50 (nM)
mDP
mEP1
mEP2
mEP3
mEP4
mFP
hTP
hIP
130
92
200
mDP
4a
4b
5b
11
2900
500
600
370
3600
>10,000
3200
>10,000
>10,000
4100
2600
NTb
1200
1400
>10,000
NTb
>10,000
7600
3.5
4.4
5.2
4.8
1.1
2.2
1.6
2.2
>10,000
>10,000
>10,000
2700
2800
1600
2200
a
IC50 (nM): mDP receptor antagonist activity.
NT: not tested.
b

4580
M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
2.4 Hz, 1H), 6.75 (dd, J = 12.3, 2.4 Hz, 1H), 5.02–4.92 (m, 1H),
4.48–4.38 (m, 2H), 3.84–3.54 (m, 2H), 3.23 (s, 3H).
2H), 6.73–6.65 (m, 2H), 4.74–4.65 (m, 1H), 4.35 (dd, J = 9.9,
5.1 Hz, 1H), 4.25 (dd, J = 9.9, 6.6 Hz, 1H), 3.41 (dd, J = 11.4, 2.7 Hz,
1H), 3.28 (dd, J = 11.4, 6.3 Hz, 1H), 2.90 (s, 3H).
4.3.13. Methyl 3-methyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14e)
4.3.21. 3-Fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16g)
1H NMR (300 MHz, CDCl3) d7.99–7.93 (m, 1H), 7.86 (dd, J = 11.4,
2.4 Hz, 1H), 7.30–7.24 (m, 1H), 7.20–6.96 (m, 4H), 5.02–4.93 (m,
1H), 4.52–4.46 (m, 2H), 3.74 (dd, J = 12.6, 2.4 Hz, 1H), 3.59 (dd,
J = 12.6, 9.3 Hz, 1H), 3.19 (s, 3H).
TLC Rf = 0.69 (n-hexane/EtOAc, 2:1); 1H NMR (300 MHz, CDCl3)
d
7.90–7.81 (m, 2H), 6.91–6.80 (m, 3H), 6.75–6.65 (m, 2H),
4.72–4.63 (m, 1H), 4.28 (dd, J = 9.6, 4.8 Hz, 1H), 4.18 (dd, J = 9.6,
6.6 Hz, 1H), 3.88 (s, 3H), 3.40 (dd, J = 11.4, 2.7 Hz, 1H), 3.28 (dd,
J = 11.4, 6.9 Hz, 1H), 2.91 (s, 3H), 2.26 (s, 3H).
4.3.14. 3-Methyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15e)
4.3.22. Methyl 3-methoxy-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoate (14h)
90% yield in two steps; TLC Rf = 0.19 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.94 (dd, J = 8.7, 2.4 Hz, 1H), 7.91–7.87
(m, 1H), 6.91–6.80 (m, 3H), 6.73–6.66 (m, 2H), 4.72–4.64 (m,
1H), 4.30 (dd, J = 9.9, 5.1 Hz, 1H), 4.20 (dd, J = 9.9, 6.9 Hz, 1H),
3.41 (dd, J = 11.4, 2.7 Hz, 1H), 3.29 (dd, J = 11.4, 6.3 Hz, 1H), 2.91
(s, 3H), 2.28 (s, 3H).
TLC Rf = 0.24 (n-hexane/EtOAc, 4:1); 1H NMR (300 MHz, CDCl3)
d 7.64 (dd, J = 8.1, 1.8 Hz, 1H), 7.55 (d, J = 1.8 Hz, 1H), 6.94 (d,
J = 8.1 Hz, 1H), 6.90–6.79 (m, 2H), 6.71–6.64 (m, 2H), 4.74–4.66
(m, 1H), 4.32 (dd, J = 9.9, 4.8 Hz, 1H), 4.21 (dd, J = 9.9, 6.9 Hz, 1H),
3.91 (s, 3H), 3.89 (s, 3H), 3.41 (dd, J = 11.7, 2.7 Hz, 1H), 3.26 (dd,
J = 11.7, 6.6 Hz, 1H), 2.89 (s, 3H).
4.3.15. 3-Methyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16e)
4.3.23. 3-Methoxy-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15h)
1H NMR (300 MHz, CDCl3) d 7.99 (dd, J = 8.4, 2.4 Hz, 1H), 7.92 (s,
1H), 6.97–6.78 (m, 5H), 4.80–4.72 (m, 1H), 4.35 (dd, J = 9.9, 4.8 Hz,
1H), 4.27 (dd, J = 9.9, 6.0 Hz, 1H), 3.48 (dd, J = 11.7, 2.4 Hz, 1H), 3.35
(dd, J = 11.7, 7.2 Hz, 1H), 2.98 (s, 3H), 2.28 (s, 3H).
85% yield in two steps; TLC Rf = 0.57 (CHCl3/CH3OH/CH3CO2H,
9:1:0.1); 1H NMR (300 MHz, CDCl3) d 7.73 (dd, J = 8.4, 2.1 Hz,
1H), 7.60 (d, J = 2.1 Hz, 1H), 6.98 (d, J = 8.4 Hz, 1H), 6.91–6.81 (m,
2H), 6.73–6.66 (m, 2H), 4.76–4.67 (m, 1H), 4.35 (dd, J = 9.9,
4.8 Hz, 1H), 4.23 (dd, J = 9.9, 6.9 Hz, 1H), 3.93 (s, 3H), 3.42 (dd,
J = 11.1, 2.4 Hz, 1H), 3.28 (dd, J = 11.1, 6.3 Hz, 1H), 2.90 (s, 3H).
4.3.16. Methyl 3-chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14f)
TLC Rf = 0.30 (n-hexane/EtOAc, 4:1); 1H NMR (300 MHz, CDCl3)
d 8.05 (d, J = 2.1 Hz, 1H), 7.91 (dd, J = 8.7, 2.1 Hz, 1H), 6.97 (d,
J = 8.7 Hz, 1H), 6.91–6.79 (m, 2H), 6.72–6.64 (m, 2H), 4.75–4.67
(m, 1H), 4.34 (dd, J = 9.6, 4.8 Hz, 1H), 4.22 (dd, J = 9.6, 7.2 Hz, 1H),
3.89 (s, 3H), 3.43 (dd, J = 11.7, 3.0 Hz, 1H), 3.33 (dd, J = 11.7,
6.3 Hz, 1H), 2.91 (s, 3H).
4.3.24. 3-Methoxy-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16h)
1H NMR (300 MHz, CDCl3) d 7.84 (dd, J = 8.4, 2.1 Hz, 1H), 7.56 (d,
J = 2.1 Hz, 1H), 7.44 (d, J = 8.4 Hz, 1H), 7.32–7.25 (m, 1H), 7.13–6.98
(m, 3H), 5.04–4.95 (m, 1H), 4.49–4.44 (m, 2H), 3.91 (s, 3H), 3.84–
3.76 (m, 1H), 3.74–3.64 (m, 1H), 3.26 (s, 3H).
4.3.17. 3-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15f)
4.3.25. Methyl 2,3-dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoate (14i)
45% yield in two steps; TLC Rf = 0.38 (CHCl3/CH3OH, 9:1); 1H
NMR (300 MHz, CDCl3) d 8.12 (s, 1H), 7.98 (d, J = 9.0 Hz, 1H), 7.01
(d, J = 9.0 Hz, 1H), 6.93–6.80 (m, 2H), 6.75–6.65 (m, 2H),
4.78–4.68 (m, 1H), 4.36 (dd, J = 9.9, 5.4 Hz, 1H), 4.24 (dd, J = 9.9,
6.3 Hz, 1H), 3.44 (dd, J = 11.7, 3.0 Hz, 1H), 3.34 (dd, J = 11.7,
6.3 Hz, 1H), 2.91 (s, 3H).
TLC Rf = 0.66 (n-hexane/EtOAc, 2:1); 1H NMR (300 MHz, CDCl3)
d 7.72 (d, J = 8.7 Hz, 1H), 6.91–6.80 (m, 2H), 6.77–6.66 (m, 3H),
4.72–4.63 (m, 1H), 4.26 (dd, J = 9.9, 4.8 Hz, 1H), 4.15 (dd, J = 9.9,
6.3 Hz, 1H), 3.86 (s, 3H), 3.40 (dd, J = 11.4, 2.7 Hz, 1H), 3.29 (dd,
J = 11.4, 6.9 Hz, 1H), 2.91 (s, 3H), 2.51 (s, 3H), 2.21 (s, 3H).
4.3.26. 2,3-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15i)
4.3.18. 3-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16f)
1H NMR (300 MHz, CDCl3) d 8.16 (d, J = 2.4 Hz, 1H), 8.08 (dd,
J = 9.0, 2.4 Hz, 1H), 7.46 (dd, J = 8.1, 1.5 Hz, 1H), 7.32 (dt, J = 8.1,
1.5 Hz, 1H), 7.15–7.03 (m, 3H), 5.13–5.03 (m, 1H), 4.57–4.50 (m,
2H), 3.88 (dd, J = 12.9, 2.1 Hz, 1H), 3.71 (dd, J = 12.9, 10.2 Hz, 1H),
3.29 (s, 3H).
88% yield in two steps; TLC Rf = 0.26 (n-hexane/EtOAc, 1:1); 1H
NMR (300 MHz, CDCl3) d 7.88 (d, J = 9.0 Hz, 1H), 6.91–6.65 (m, 5H),
4.72–4.63 (m, 1H), 4.27 (dd, J = 9.6, 4.8 Hz, 1H), 4.17 (dd, J = 9.6,
6.6 Hz, 1H), 3.40 (dd, J = 11.4, 2.7 Hz, 1H), 3.29 (dd, J = 11.4,
6.6 Hz, 1H), 2.91 (s, 3H), 2.57 (s, 3H), 2.22 (s, 3H).
4.3.27. 2,3-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16i)
4.3.19. Methyl 3-fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoate (14g)
1H NMR (300 MHz, CDCl3) d 8.12 (d, J = 9.3 Hz, 1H), 7.49 (d,
J = 7.8 Hz, 1H), 7.33 (t, J = 7.8 Hz, 1H), 7.15–7.02 (m, 2H), 6.82 (d,
J = 9.3 Hz, 1H), 5.07–4.96 (m, 1H), 4.47–4.40 (m, 2H), 3.89–3.81
(m, 1H), 3.72–3.58 (m, 1H), 3.29 (s, 3H), 2.48 (s, 3H), 2.19 (s, 3H).
TLC Rf = 0.30 (n-hexane/EtOAc, 4:1); 1H NMR (300 MHz, CDCl3)
d 7.82–7.71 (m, 2H), 7.01 (t, J = 8.1 Hz, 1H), 6.90–6.79 (m, 2H),
6.72–6.65 (m, 2H), 4.73–4.64 (m, 1H), 4.33 (dd, J = 9.9, 5.1 Hz,
1H), 4.23 (dd, J = 9.9, 6.6 Hz, 1H), 3.89 (s, 3H), 3.40 (dd, J = 11.4,
2.7 Hz, 1H), 3.28 (dd, J = 11.4, 6.3 Hz, 1H), 2.90 (s, 3H).
4.3.28. Methyl 2,5-dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoate (14j)
4.3.20. 3-Fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoic acid (15g)
69% yield in two steps; TLC Rf = 0.57 (CHCl3/CH3OH/CH3CO2H,
9:1:0.1); 1H NMR (300 MHz, CDCl3) d 7.89–7.83 (m, 1H), 7.80
(dd, J = 11.4, 1.8 Hz, 1H), 7.04 (t, J = 8.4 Hz, 1H), 6.91–6.79 (m,
TLC Rf = 0.57 (toluene/EtOAc, 9:1); 1H NMR (300 MHz, CDCl3) d
7.76 (s, 1H), 6.92–6.80 (m, 2H), 6.74–6.65 (m, 3H), 4.71–4.63 (m,
1H), 4.27 (dd, J = 9.6, 4.8 Hz, 1H), 4.15 (dd, J = 9.6, 6.6 Hz, 1H),
3.85 (s, 3H), 3.40 (dd, J = 11.7, 3.0 Hz, 1H), 3.28 (dd, J = 11.7,
6.6 Hz, 1H), 2.91 (s, 3H), 2.57 (s, 3H), 2.21 (s, 3H).

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4581
4.3.29. 2,5-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
4.4.4. Benzyl [5-fluoro-2-methyl-1-(4-{[(2S)-4-methyl-3,4-
benzoxazin-2-yl]methoxy}benzoic acid (15j)
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetate (17d)
97% yield in two steps; TLC Rf = 0.50 (CHCl3/CH3OH, 9:1); 1H
NMR (300 MHz, CDCl3)
d
7.87 (s, 1H), 6.91–6.80 (m, 2H),
Method B: 85% yield; TLC Rf = 0.46 (n-hexane/EtOAc, 1:1); 1H
NMR (300 MHz, CDCl3) d 7.68 (dd, J = 14.4 Hz, 2H), 7.33–7.25 (m,
5H), 7.20–7.10 (m, 2H), 7.02–6.65 (m, 7H), 5.15 (m, 2H), 4.70 (m,
1H), 4.31 (dd, J = 9.6, 2.1 Hz, 1H), 4.21 (dd, J = 9.6, 6.3 Hz, 1H),
3.71 (s, 2H), 3.40 (dd, J = 11.4, 2.1 Hz, 1H), 3.28 (dd, J = 11.4,
6.3 Hz, 1H), 2.92 (s, 3H), 2.37 (s, 3H).
6.74–6.65 (m, 3H), 4.72–4.63 (m, 1H), 4.28 (dd, J = 9.6, 4.8 Hz,
1H), 4.17 (dd, J = 9.6, 6.9 Hz, 1H), 3.40 (dd, J = 11.7, 2.7 Hz, 1H),
3.28 (dd, J = 11.7, 6.3 Hz, 1H), 2.91 (s, 3H), 2.61 (s, 3H), 2.22
(s, 3H).
4.3.30. 2,5-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl chloride (16j)
4.4.5. Benzyl [5-methoxy-2-methyl-1-(4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetate (17e)
1H NMR (300 MHz, CDCl3) d 8.06 (s, 1H), 7.44 (d, J = 8.1 Hz, 1H),
7.33–7.23 (m, 1H), 7.14–7.00 (m, 2H), 6.71 (s, 1H), 5.05–4.95 (m,
1H), 4.47–4.38 (m, 2H), 3.85–3.76 (m, 1H), 3.72–3.56 (m, 1H),
3.26 (s, 3H), 2.57 (s, 3H), 2.24 (s, 3H).
Method A: 26% yield; TLC Rf = 0.53 (toluene/EtOAc, 9:1); 1H NMR
(300 MHz, CDCl3) d 7.69 (d, J = 9.0 Hz, 2H), 7.38–7.28 (m, 5H), 7.04–
6.62 (m, 9H), 5.14 (s, 2H), 4.74–4.62 (m, 1H), 4.31 (dd, J = 9.9,
4.8 Hz, 1H), 4.21 (dd, J = 9.9, 6.3 Hz, 1H), 3.76 (s, 3H), 3.72 (s, 2H),
3.41 (dd, J = 11.4, 2.7 Hz, 1H), 3.28 (dd, J = 11.4, 6.3 Hz, 1H), 2.92
(s, 3H), 2.39 (s, 3H).
4.4. General procedure for the preparation of 17a–u
4.4.1. Benzyl [2-methyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetate
(17a)
4.4.6. Benzyl [2-methyl-1-(2-methyl-4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetate (17f)
Method A: To a stirred solution of 19a (1 g, 3.58 mmol) and 16a
(1.52 g, 5.37 mmol) in CH2Cl2 (10 mL) were added 20 M NaOH aq
(0.9 mL, 17.9 mmol) and TBACl (99 mg, 0.358 mmol). After stirring
for 1 h at room temperature, the reaction mixture was quenched
with water and extracted with EtOAc (ꢀ2). The combined organic
layers were washed with water, brine, dried over Na2SO4 and con-
centrated in vacuo. The resulting residue was purified by column
chromatography on silica gel to yield 17a (462 mg, 23%); TLC
Method A: 20% yield; TLC Rf = 0.48 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.47 (d, J = 7.5 Hz, 1H), 7.40–7.25 (m,
7H), 7.20–7.00 (m, 3H), 6.93–6.76 (m, 3H), 6.75–6.67 (m, 2H),
5.13 (s, 2H), 4.73–4.63 (m, 1H), 4.29 (dd, J = 9.6, 4.8 Hz, 1H), 4.18
(dd, J = 9.6, 6.3 Hz, 1H), 3.74 (s, 2H), 3.41 (dd, J = 11.4, 2.7 Hz,
1H), 3.28 (dd, J = 11.4, 6.6 Hz, 1H), 2.92 (s, 3H), 2.31 (s, 3H), 2.29
(s, 3H).
Rf = 0.63 (toluene/EtOAc, 9:1);1H NMR (300 MHz, CDCl3)
d
7.76–6.67 (m, 17H), 5.13 (s, 2H), 4.74–4.62 (m, 1H), 4.32 (dd,
J = 9.9, 5.4 Hz, 1H), 4.21 (dd, J = 9.9, 6.3 Hz, 1H), 3.76 (s, 2H), 3.41
(dd, J = 11.7, 2.7 Hz, 1H), 3.28 (dd, J = 11.4, 6.6 Hz, 1H), 2.92 (s,
3H), 2.40 (s, 3H).
4.4.7. Benzyl [1-(2-chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetate (17g)
Method A: 60% yield; TLC Rf = 0.42 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.54–7.40 (m, 2H), 7.40–7.03 (m, 9H),
6.97–6.82 (m, 3H), 6.75–6.67 (m, 2H), 5.12 (s, 2H), 4.72–4.64 (m,
1H), 4.29 (dd, J = 9.9, 4.8 Hz, 1H), 4.20 (dd, J = 9.9, 6.0 Hz, 1H),
3.73 (s, 2H), 3.40 (dd, J = 12.0, 2.7 Hz, 1H), 3.27 (dd, J = 12.0,
6.0 Hz, 1H), 2.92 (s, 3H), 2.30 (s, 3H).
4.4.2. Benzyl [2,5-dimethyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-
yl]acetate (17b)
Method B: To a stirred solution of 19b (410 mg, 1.40 mmol) in
CH3CN (4 mL) and EtOAc (4 mL) were added TEA (1.17 mL,
8.40 mmol), DMAP (51.3 mg, 0.42 mmol) and then 16a (593 mg,
2.10 mmol). After stirring for 14 h at 40 °C, the reaction mixture
was quenched with water and extracted with MTBE (ꢀ2). The com-
bined organic layers were washed with NaHCO3 aq, brine, dried
over MgSO4 and concentrated in vacuo. The resulting residue was
purified by column chromatography on silica gel to yield 17b
(370 mg, 46%); TLC Rf = 0.46 (n-hexane/EtOAc, 1:1); 1H NMR
(300 MHz, CDCl3) d 7.70 (dd, J = 6.9, 2.1 Hz, 2H), 7.34–7.27 (m,
5H), 7.01 (dd, J = 6.9, 2.1 Hz, 2H), 6.92–6.83 (m, 5H), 6.73–6.67
(m, 2H), 6.67 (s, 2H), 4.68 (m, 1H), 4.31 (dd, J = 9.6, 4.0 Hz, 1H),
4.20 (dd, J = 9.6, 6.6 Hz, 1H), 3.73 (s, 2H), 3.43 (dd, J = 12.0,
4.0 Hz, 1H), 3.28 (dd, J = 12.0, 6.6 Hz, 1H), 2.92 (s, 3H), 2.39 (s,
3H), 2.38 (s, 3H).
4.4.8. Benzyl [1-(2-fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetate (17h)
Method A: 41% yield; TLC Rf = 0.53 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.56–7.47 (m, 3H), 7.34–7.26 (m, 4H),
7.21–7.05 (m, 3H), 6.92–6.82 (m, 3H), 6.74–6.68 (m, 3H), 5.13 (s,
2H), 4.72–4.64 (m, 1H), 4.32–4.09 (m, 2H), 3.74 (s, 2H),
3.42–3.25 (m, 2H), 2.92 (s, 3H), 2.38 (s, 3H).
4.4.9. Benzyl [2-methyl-1-(3-methyl-4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetate (17i)
Method A: 70% yield; TLC Rf = 0.51 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.61 (d, J = 2.1 Hz, 1H), 7.54 (dd, J = 8.1,
2.1 Hz, 1H), 7.50 (d, J = 7.8 Hz, 1H), 7.34–7.29 (m, 5H), 7.18–6.98
(m, 3H), 6.92–6.80 (m, 3H), 6.75–6.65 (m, 2H), 5.14 (s, 2H),
4.75–4.65 (m, 1H), 4.34–4.17 (m, 2H), 3.76 (s, 2H), 3.45–3.28 (m,
2H), 2.92 (s, 3H), 2.40 (s, 3H), 2.27 (s, 3H).
4.4.3. Benzyl [5-chloro-2-methyl-1-(4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetate (17c)
Method B: 42% yield; TLC Rf = 0.81 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3)
d 7.68 (d, J = 9.0 Hz, 2H), 7.47 (d,
4.4.10. Benzyl [1-(3-chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetate (17j)
J = 2.1 Hz, 1H), 7.40–7.27 (m, 5H), 7.02–6.96 (m, 3H), 6.93–6.82
(m, 3H), 6.74–6.66 (m, 2H), 5.15 (s, 2H), 4.73–4.63 (m, 1H), 4.31
(dd, J = 9.6, 4.8 Hz, 1H), 4.21 (dd, J = 9.6, 6.0 Hz, 1H), 3.71 (s, 2H),
3.41 (dd, J = 11.4, 2.7 Hz, 1H), 3.28 (dd, J = 11.4, 6.6 Hz, 1H), 2.92
(s, 3H), 2.38 (s, 3H).
Method A: 75% yield; TLC Rf = 0.42 (toluene/EtOAc, 9:1); 1H NMR
(300 MHz, CDCl3) d 7.84 (d, J = 1.8 Hz, 1H), 7.60 (dd, J = 8.4, 2.1 Hz,
1H), 7.50 (d, J = 7.2 Hz, 1H), 7.34–7.29 (m, 5H), 7.20–6.98 (m, 4H),

4582
M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
6.92–6.82 (m, 2H), 6.73–6.68 (m, 2H), 5.14 (s, 2H), 4.76–4.72 (m,
1H), 4.41–4.24 (m, 2H), 3.76 (s, 2H), 3.46 (dd, J = 11.4, 2.7 Hz,
1H), 3.36 (dd, J = 11.4, 6.0 Hz, 1H), 2.93 (s, 3H), 2.40 (s, 3H).
4.4.17. Benzyl [5-fluoro-1-(2-fluoro-4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-
1H-indol-3-yl]acetate (17q)
Method A: 82% yield; TLC Rf = 0.55 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.53 (t, J = 8.4 Hz, 1H), 7.38–7.08 (m,
7H), 6.93–6.76 (m, 4H), 6.76–6.66 (m, 3H), 5.13 (s, 2H),
4.73–4.63 (m, 1H), 4.29 (dd, J = 9.6, 5.1 Hz, 1H), 4.20 (dd, J = 9.6,
6.0 Hz, 1H), 3.68 (s, 2H), 3.40 (dd, J = 11.4, 2.7 Hz, 1H), 3.27 (dd,
J = 11.4, 6.3 Hz, 1H), 2.92 (s, 3H), 2.33 (s, 3H).
4.4.11. Benzyl [1-(3-fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetate (17k)
Method A: 38% yield; TLC Rf = 0.30 (n-hexane/EtOAc, 3:1); 1H
NMR (300 MHz, CDCl3) d 7.56–6.68 (m, 16H), 5.14 (s, 2H),
4.76–4.65 (m, 1H), 4.40–4.25 (m, 2H), 3.75 (s, 2H), 3.45–3.28 (m,
2H), 2.92 (s, 3H), 2.40 (s, 3H).
4.4.18. Benzyl [5-fluoro-2-methyl-1-(3-methyl-4-{[(2S)-4-
methyl-3,4-dihydro-2H-1,4-benzoxazin-2-
yl]methoxy}benzoyl)-1H-indol-3-yl]acetate (17r)
4.4.12. Benzyl [1-(3-methoxy-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-
3-yl]acetate (17l)
Method A: 84% yield; TLC Rf = 0.55 (toluene/EtOAc, 9:1); 1H NMR
(300 MHz, CDCl3) d 7.60–7.56 (m, 1H), 7.51 (dd, J = 8.7, 2.7 Hz, 1H),
7.38–7.27 (m, 5H), 7.14 (dd, J = 9.0, 2.7 Hz, 1H), 6.95 (dd, J = 9.0,
4.5 Hz, 1H), 6.93–6.67 (m, 6H), 5.15 (s, 2H), 4.74–4.65 (m, 1H),
4.31 (dd, J = 9.9, 5.1 Hz, 1H), 4.22 (dd, J = 9.9, 6.3 Hz, 1H), 3.71 (s,
2H), 3.42 (dd, J = 11.4, 2.7 Hz, 1H), 3.31 (dd, J = 11.4, 6.6 Hz, 1H),
2.92 (s, 3H), 2.37 (s, 3H), 2.27 (s, 3H).
Method A: 64% yield; TLC Rf = 0.38 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3)
d 7.50 (d, J = 8.1 Hz, 1H), 7.36 (d,
J = 2.4 Hz, 1H), 7.33–7.26 (m, 6H), 7.18–7.03 (m, 3H), 6.94–6.82
(m, 3H), 6.72–6.67 (m, 2H), 5.14 (s, 2H), 4.78–4.70 (m, 1H),
4.38–4.22 (m, 2H), 3.87 (s, 3H), 3.76 (s, 2H), 3.43 (dd, J = 11.4,
2.7 Hz, 1H), 3.30 (dd, J = 11.4, 6.6 Hz, 1H), 2.91 (s, 3H), 2.41
(s, 3H).
4.4.19. Benzyl [5-fluoro-1-(3-methoxy-4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-
1H-indol-3-yl]acetate (17s)
4.4.13. Benzyl [1-(2,3-dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-
3-yl]acetate (17m)
Method A: 58% yield; TLC Rf = 0.55 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.38–7.20 (m, 7H), 7.14 (dd, J = 6.0,
2.7 Hz, 1H), 7.02–6.65 (m, 7H), 5.15 (s, 2H), 4.78–4.68 (m, 1H),
4.35 (dd, J = 9.9, 4.8 Hz, 1H), 4.25 (dd, J = 9.9, 6.6 Hz, 1H), 3.87 (s,
3H), 3.71 (s, 2H), 3.42 (dd, J = 11.7, 3.0 Hz, 1H), 3.30 (dd, J = 11.7,
6.3 Hz, 1H), 2.91 (s, 3H), 2.38 (s, 3H).
Method A: 68% yield; TLC Rf = 0.57 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.47 (d, J = 7.5 Hz, 1H), 7.35–7.27 (m,
5H), 7.19–7.14 (m, 2H), 7.10–7.03 (m, 2H), 6.92–6.83 (m, 2H),
6.77–6.67 (m, 3H), 5.13 (s, 2H), 4.72–4.67 (m, 1H), 4.30–4.08 (m,
2H), 3.74 (s, 2H), 3.45–3.29 (m, 2H), 2.93 (s, 3H), 2.29 (s, 3H),
2.23 (s, 3H), 2.22 (s, 3H).
4.4.20. Benzyl [1-(2,3-dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-5-fluoro-2-methyl-
1H-indol-3-yl]acetate (17t)
Method A: 65% yield; TLC Rf = 0.54 (n-hexane/EtOAc, 2:1); 1H
NMR (300 MHz, CDCl3) d 7.38–7.23 (m, 5H), 7.17–7.03 (m, 3H),
6.92–6.66 (m, 6H), 5.13 (s, 2H), 4.74–4.65 (m, 1H), 4.27 (dd,
J = 9.6, 4.8 Hz, 1H), 4.18 (dd, J = 9.6, 6.0 Hz, 1H), 3.67 (s, 2H), 3.43
(dd, J = 11.4, 2.7 Hz, 1H), 3.32 (dd, J = 11.4, 6.9 Hz, 1H), 2.93 (s,
3H), 2.24 (s, 3H), 2.23 (s, 3H), 2.21 (s, 3H).
4.4.14. Benzyl [1-(2,5-dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-
3-yl]acetate (17n)
Method A: 47% yield; TLC Rf = 0.60 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.46 (d, J = 7.8 Hz, 1H), 7.34–7.22 (m,
5H), 7.20–7.12 (m, 2H), 7.06–7.00 (m, 2H), 6.92–6.81 (m, 2H),
6.74–6.66 (m, 3H), 5.12 (s, 2H), 4.73–4.64 (m, 1H), 4.29 (dd,
J = 9.9, 4.8 Hz, 1H), 4.19 (dd, J = 9.9, 6.6 Hz, 1H), 3.73 (s, 2H), 3.42
(dd, J = 11.7, 2.7 Hz, 1H), 3.31 (dd, J = 11.7, 6.3 Hz, 1H), 2.92 (s,
3H), 2.31 (s, 3H), 2.22 (s, 3H), 2.17 (s, 3H).
4.4.21. Benzyl [1-(2,5-dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-5-fluoro-2-methyl-
1H-indol-3-yl]acetate (17u)
Method A: 73% yield; TLC Rf = 0.52 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.40–7.23 (m, 5H), 7.20–7.08 (m, 2H),
7.07–6.97 (m, 1H), 6.93–6.66 (m, 6H), 5.13 (s, 2H), 4.74–4.64 (m,
1H), 4.30 (dd, J = 9.6, 4.5 Hz, 1H), 4.20 (dd, J = 9.6, 6.6 Hz, 1H),
3.68 (s, 2H), 3.42 (dd, J = 11.4, 3.0 Hz, 1H), 3.31 (dd, J = 11.4,
6.0 Hz, 1H), 2.93 (s, 3H), 2.27 (s, 3H), 2.21 (s, 3H), 2.18 (s, 3H).
4.4.15. Benzyl [5-fluoro-2-methyl-1-(2-methyl-4-{[(2S)-4-
methyl-3,4-dihydro-2H-1,4-benzoxazin-2-
yl]methoxy}benzoyl)-1H-indol-3-yl]acetate (17o)
Method A: 24% yield; TLC Rf = 0.48 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.38–7.24 (m, 6H), 7.12 (dd, J = 9.0,
2.4 Hz, 1H), 7.03 (dd, J = 9.0, 4.2 Hz, 1H), 6.93–6.67 (m, 7H), 5.13
(s, 2H), 4.72–4.62 (m, 1H), 4.29 (dd, J = 9.9, 5.1 Hz, 1H), 4.17 (dd,
J = 9.9, 6.3 Hz, 1H), 3.68 (s, 2H), 3.41 (dd, J = 11.4, 2.7 Hz, 1H),
3.28 (dd, J = 11.4, 6.6 Hz, 1H), 2.92 (s, 3H), 2.28 (s, 3H), 2.27
(s, 3H).
4.5. General procedure for the preparation of 3a–e, 4a–i, 5a–g
4.5.1. [2-Methyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic acid
(3a)
4.4.16. Benzyl [1-(2-chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-5-fluoro-2-methyl-1H-
indol-3-yl]acetate (17p)
To a stirred solution of 17a (462 mg, 0.82 mmol) in EtOAc
(20 mL) was added 20% Pd(OH)2/C (100 mg) at room temperature.
The resulting suspension was vigorously stirred for 80 min at room
temperature under hydrogen atmosphere. The reaction mixture
was filtered through a pad of Celite, and the filtrate was concen-
trated in vacuo. The resulting residue was purified by column chro-
matography on silica gel to yield 3a (380 mg, quant); TLC Rf = 0.42
(CHCl3/MeOH, 9:1); 1H NMR (300 MHz, CDCl3) d 7.73 (d, J = 8.7 Hz,
2H), 7.51 (d, J = 8.1 Hz, 1H), 7.17 (dt, J = 1.5, 8.1 Hz, 1H), 7.09–6.94
(m, 4H), 6.93–6.81 (m, 2H), 6.75–6.66 (m, 2H), 4.74–4.64 (m, 1H),
Method A: 40% yield; TLC Rf = 0.40 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.42 (d, J = 8.4 Hz, 1H), 7.40–7.10 (m,
7H), 7.04 (d, J = 2.7 Hz, 1H), 6.98–6.80 (m, 4H), 6.76–6.68 (m,
2H), 5.13 (s, 2H), 4.73–4.63 (m, 1H), 4.29 (dd, J = 9.6, 5.4 Hz, 1H),
4.20 (dd, J = 9.6, 5.7 Hz, 1H), 3.67 (s, 2H), 3.40 (dd, J = 11.1,
2.4 Hz, 1H), 3.27 (dd, J = 11.1, 6.3 Hz, 1H), 2.92 (s, 3H), 2.24
(s, 3H).

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4583
4.32 (dd, J = 9.9, 5.1 Hz, 1H), 4.21 (dd, J = 9.9, 6.3 Hz, 1H), 3.75 (s,
2H), 3.41 (dd, J = 11.7, 2.4 Hz, 1H), 3.28 (dd, J = 11.7, 6.6 Hz, 1H),
2.93 (s, 3H), 2.43 (s, 3H); MS (APCI, Neg, 20 V) m/z 469 (MꢁH)ꢁ;
IR (KBr) 3046, 2928, 1735, 1709, 1682, 1604, 1577, 1505, 1474,
1457, 1420, 1396, 1351, 1316, 1246, 1224, 1175, 1065, 1041,
924, 874, 840, 743, 662, 624, 525, 465 cmꢁ1; Optical rotation
1H), 7.11–6.95 (m, 2H), 6.94–6.76 (m, 3H), 6.76–6.65 (m, 2H),
4.72–4.62 (m, 1H), 4.29 (dd, J = 9.9, 5.1 Hz, 1H), 4.19 (dd, J = 9.9,
6.3 Hz, 1H), 3.72 (s, 2H), 3.40 (dd, J = 11.4, 1.8 Hz, 1H), 3.27 (dd,
J = 11.4, 6.6 Hz, 1H), 2.92 (s, 3H), 2.34 (s, 3H), 2.32 (s, 3H); MS
(APCI, Neg, 20 V) m/z 483 (MꢁH)ꢁ; IR (KBr) 3065, 2926, 2871,
2249, 1710, 1684, 1605, 1572, 1503, 1474, 1457, 1383, 1351,
1316, 1241, 1225, 1176, 1131, 1115, 1046, 1026, 909, 856, 822,
½
a 2D3
+10.88 (c 0.75, DMSO).
ꢂ
742, 647, 619, 571 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+9.03 (c 0.75,
4.5.2. [2,5-Dimethyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic acid
(3b)
DMSO).
4.5.8. [1-(2-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4b)
37% yield; TLC Rf = 0.39 (CHCl3/MeOH, 10:1); 1H NMR
(300 MHz, CDCl3) d 7.71 (d, J = 9.6 Hz, 2H), 7.28 (s, 1H), 6.99 (d,
J = 9.6 Hz, 2H), 6.90–6.80 (m, 4H), 6.73–6.70 (m, 2H), 4.67 (m,
1H), 4.30 (dd, J = 9.9, 4.8 Hz, 1H), 4.21 (dd, J = 9.9, 6.3 Hz, 1H),
3.71 (s, 2H), 3.40 (dd, J = 11.7, 3.0 Hz, 1H), 3.28 (dd, J = 11.7,
6.6 Hz, 1H), 2.92 (s, 3H), 2.41 (s, 3H), 2.40 (s, 3H); MS (APCI, Neg,
20 V) m/z 483 (MꢁH)ꢁ; IR (KBr) 2925, 1710, 1681, 1604, 1578,
1505, 1463, 1350, 1307, 1225, 1174, 1138, 1068, 1041, 926, 842,
75% yield; TLC Rf = 0.53 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.52–7.42 (m, 2H), 7.33–7.04 (m, 4H), 6.98–6.82 (m, 3H),
6.76–6.68 (m, 2H), 4.72–4.64 (m, 1H), 4.29 (dd, J = 9.9, 4.8 Hz, 1H),
4.20 (dd, J = 9.9, 6.0 Hz, 1H), 3.72 (s, 2H), 3.40 (dd, J = 12.0, 2.7 Hz,
1H), 3.27 (dd, J = 12.0, 6.0 Hz, 1H), 2.92 (s, 3H), 2.32 (s, 3H); MS
(APCI, Neg, 20 V) m/z 503 (MꢁH)ꢁ; IR (KBr) 3042, 2928, 2871,
2821, 1733, 1709, 1685, 1600, 1561, 1504, 1474, 1458, 1355,
1319, 1299, 1221, 1136, 1075, 1044, 983, 964, 931, 914, 854, 821,
805, 762, 744 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+9.70 (c 0.75, DMSO).
4.5.3. [5-Chloro-2-methyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic
acid (3c)
743, 607, 571, 487 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+8.80 (c 0.75, DMSO).
4.5.9. [1-(2-Fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4c)
50% yield; TLC Rf = 0.40 (CHCl3/MeOH, 10:1); 1H NMR
(300 MHz, CDCl3) d 7.71 (d, J = 9.0 Hz, 2H), 7.47 (d, J = 1.8 Hz, 1H),
7.05–6.95 (m, 3H), 6.92–6.80 (m, 3H), 6.73–6.70 (m, 2H), 4.69
(m, 1H), 4.31 (dd, J = 9.9, 5.1 Hz, 1H), 4.22 (dd, J = 9.9, 6.0 Hz, 1H),
3.71 (s, 2H), 3.40 (dd, J = 11.7, 2.7 Hz, 1H), 3.28 (dd, J = 11.7,
6.3 Hz, 1H), 2.92 (s, 3H), 2.41 (s, 3H); MS (APCI, Neg, 20 V) m/z
503 (MꢁH)ꢁ; IR (KBr) 2928, 1686, 1604, 1505, 1458, 1347, 1308,
1224, 1175, 1137, 1061, 1042, 842, 745 cmꢁ1; Optical rotation
63% yield; TLC Rf = 0.63 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.55 (t, J = 8.4 Hz, 1H), 7.49 (d, J = 7.5 Hz, 1H), 7.20 (dt,
J = 6.6, 1.5 Hz, 1H), 7.15–7.06 (m, 2H), 6.92–6.83 (m, 3H), 6.75–
6.68 (m, 3H), 4.72–4.65 (m, 1H), 4.32–4.17 (m, 2H), 3.72 (s, 2H),
3.40 (dd, J = 11.4, 2.7 Hz, 1H), 3.27 (dd, J = 11.4, 6.3 Hz, 1H), 2.92
(s, 3H), 2.42 (s, 3H); MS (APCI, Neg, 20 V) m/z 487 (MꢁH)ꢁ; IR
(KBr) 3068, 2929, 2871, 2822, 2616, 1709, 1685, 1619, 1578,
1504, 1474, 1458, 1397, 1355, 1320, 1297, 1225, 1168, 1131,
1119, 1066, 1040, 984, 918, 859, 745, 615, 528 cmꢁ1; Optical rota-
½
a 2D3
+10.19 (c 0.75, DMSO).
ꢂ
4.5.4. [5-Fluoro-2-methyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic
acid (3d)
tion ½a 2D3
ꢂ
+8.41 (c 0.75, DMSO).
12% yield; TLC Rf = 0.42 (CHCl3/MeOH, 10:1); 1H NMR
(300 MHz, CDCl3) d 7.70 (d, J = 9.0 Hz, 2H), 7.15 (m, 1H), 7.00 (d,
J = 9.0 Hz, 2H), 6.95–6.65 (m, 6H), 4.68 (m, 1H), 4.30 (dd, J = 9.6,
4.8 Hz, 1H), 4.21 (dd, J = 9.6, 6.0 Hz, 1H), 3.68 (s, 2H), 3.39 (dd,
J = 11.7, 2.7 Hz, 1H), 3.28 (dd, J = 11.7, 6.6 Hz, 1H), 2.91 (s, 3H),
2.39 (s, 3H); MS (APCI, Neg, 20 V) m/z 487 (MꢁH)ꢁ; IR (KBr)
3069, 2930, 1711, 1684, 1604, 1472, 1459, 1373, 1355, 1314,
1248, 1224, 1173, 1136, 1065, 1042, 930, 844, 806, 762,
4.5.10. [2-Methyl-1-(3-methyl-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic
acid (4d)
32% yield; TLC Rf = 0.52 (CHCl3/MeOH/AcOH, 9:1:0.1); 1H NMR
(300 MHz, CDCl3) d 7.61 (d, J = 1.2 Hz, 1H), 7.56 (dd, J = 8.7,
2.4 Hz, 1H), 7.50 (d, J = 7.8 Hz, 1H), 7.16 (dt, J = 6.9, 1.2 Hz, 1H),
7.04 (dt, J = 8.4, 1.2 Hz, 1H), 6.99 (d, J = 7.8 Hz, 1H), 6.92–6.82 (m,
3H), 6.73–6.68 (m, 2H), 4.74–4.66 (m, 1H), 4.34–4.19 (m, 2H),
3.74 (s, 2H), 3.42 (dd, J = 11.4, 2.7 Hz, 1H), 3.31 (dd, J = 11.4,
6.6 Hz, 1H), 2.92 (s, 3H), 2.41 (s, 3H), 2.26 (s, 3H); MS (APCI, Neg,
20 V) m/z 483 (MꢁH)ꢁ; IR (KBr) 3048, 2927, 2821, 1682, 1604,
1580, 1504, 1474, 1457, 1415, 1397, 1351, 1317, 1265, 1225,
1191, 1133, 1122, 1071, 1041, 962, 913, 822, 744, 607, 598, 573,
745 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+14.68 (c 0.60, DMSO).
4.5.6. [5-Methoxy-2-methyl-1-(4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic
acid (3e)
99% yield; TLC Rf = 0.42 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.71 (d, J = 8.7 Hz, 2H), 7.04–6.94 (m, 3H), 6.94–6.80 (m,
3H), 6.76–6.62 (m, 3H), 4.74–4.64 (m, 1H), 4.31 (dd, J = 9.6,
5.1 Hz, 1H), 4.21 (dd, J = 9.6, 6.0 Hz, 1H), 3.83 (s, 3H), 3.71 (s, 2H),
3.41 (dd, J = 11.7, 2.4 Hz, 1H), 3.28 (dd, J = 11.7, 6.3 Hz, 1H), 2.91
(s, 3H), 2.42 (s, 3H); MS (APCI, Neg, 20 V) m/z 499 (MꢁH)ꢁ; IR
(KBr) 2929, 2831, 1735, 1709, 1679, 1604, 1578, 1505, 1477,
1458, 1357, 1314, 1224, 1173, 1143, 1067, 1037, 992, 926, 840,
545, 527, 499, 488, 479, 464, 455 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+11.52 (c 0.75, DMSO).
4.5.11. [1-(3-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4e)
46% yield; TLC Rf = 0.52 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.84 (d, J = 2.1 Hz, 1H), 7.62 (dd, J = 8.4, 2.1 Hz, 1H), 7.51
(d, J = 7.8 Hz, 1H), 7.19 (dt, J = 6.9, 1.2 Hz, 1H), 7.09–6.96 (m, 3H),
6.92–6.82 (m, 2H), 6.73–6.67 (m, 2H), 4.78–4.71 (m, 1H), 4.41–
4.24 (m, 2H), 3.74 (s, 2H), 3.45 (dd, J = 11.4, 2.7 Hz, 1H), 3.36 (dd,
J = 11.4, 6.0 Hz, 1H), 2.93 (s, 3H), 2.42 (s, 3H); MS (APCI, Neg,
20 V) m/z 503 (MꢁH)ꢁ; IR (KBr) 3067, 2928, 2870, 2822, 1710,
1684, 1596, 1503, 1474, 1458, 1405, 1351, 1317, 1277, 1271,
1244, 1224, 1159, 1137, 1065, 1027, 944, 915, 822, 745, 610,
808, 762, 744, 676, 608, 461 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+9.69 (c
0.75, DMSO).
4.5.7. [2-Methyl-1-(2-methyl-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-3-yl]acetic
acid (4a)
50% yield; TLC Rf = 0.52 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.48 (d, J = 7.8 Hz, 1H), 7.40–7.28 (m, 2H), 7.23–7.13 (m,
571, 464, 454 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+17.78 (c 0.75, DMSO).

4584
M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4.5.12. [1-(3-Fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4f)
1355, 1317, 1279, 1241, 1224, 1198, 1174, 1118, 1064, 1049,
1024, 996, 911, 850, 807, 764, 737, 693, 673, 658, 620, 599, 589,
571, 523, 465, 454 cmꢁ1; Optical rotation ½a 2D4
ꢂ
+7.71 (c 0.75,
32% yield; TLC Rf = 0.55 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.57–7.48 (m, 3H), 7.19 (dt, J = 7.4, 1.2 Hz, 1H), 7.09–7.03
(m, 2H), 6.97 (d, J = 8.1 Hz, 1H), 6.92–6.81 (m, 2H), 6.73–6.67 (m,
2H), 4.75–4.68 (m, 1H), 4.40–4.25 (m, 2H), 3.74 (s, 2H),
3.45–3.28 (m, 2H), 2.92 (s, 3H), 2.42 (s, 3H), 1.90 (br s, 1H) m/z
487 (MꢁH)ꢁ; IR (KBr) 3069, 2931, 1689, 1604, 1578, 1506, 1459,
DMSO).
4.5.17. [1-(2-Chloro-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-5-fluoro-2-methyl-1H-
indol-3-yl]acetic acid (5b)
75% yield; TLC Rf = 0.48 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.44 (d, J = 8.7 Hz, 1H), 7.22 (dd, J = 8.7, 4.5 Hz, 1H), 7.15
(dd, J = 8.7, 2.1 Hz, 1H), 7.05 (d, J = 2.1 Hz, 1H), 7.00–6.80 (m, 4H),
6.75–6.68 (m, 2H), 4.73–4.63 (m, 1H), 4.29 (dd, J = 9.9, 5.4 Hz,
1H), 4.20 (dd, J = 9.9, 6.0 Hz, 1H), 3.67 (s, 2H), 3.40 (dd, J = 11.4,
2.7 Hz, 1H), 3.27 (dd, J = 11.4, 6.3 Hz, 1H), 2.92 (s, 3H), 2.27 (s,
3H); MS (APCI, Neg, 20 V) m/z 521 (MꢁH)ꢁ; IR (KBr) 3068, 2930,
2871, 2822, 1908, 1710, 1688, 1601, 1561, 1504, 1471, 1460,
1359, 1322, 1300, 1278, 1223, 1139, 1072, 1046, 996, 935, 926,
850, 811, 745, 621, 593, 585, 572, 515, 485, 475 cmꢁ1; Optical rota-
1318, 1173, 1117, 1056, 745 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+11.38
(c 0.75, DMSO).
4.5.13. [1-(3-Methoxy-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4g)
56% yield; TLC Rf = 0.65 (CHCl3/MeOH/AcOH, 9:1:0.1); 1H NMR
(300 MHz, CDCl3) d 7.50 (d, J = 7.5 Hz, 1H), 7.35 (d, J = 1.8 Hz, 1H),
7.30 (dd, J = 8.4, 2.1 Hz, 1H), 7.20–7.15 (m, 1H), 7.08–6.81 (m,
5H), 6.72–6.66 (m, 2H), 4.77–4.69 (m, 1H), 4.38–4.22 (m, 2H),
3.87 (s, 3H), 3.74 (s, 2H), 3.45–3.26 (m, 2H), 2.91 (s, 3H), 2.42 (s,
3H); MS (APCI, Neg, 20 V) m/z 499 (MꢁH)ꢁ; IR (KBr) 3067, 2932,
2871, 2827, 2617, 1710, 1682, 1598, 1506, 1458, 1417, 1350,
1317, 1273, 1244, 1215, 1183, 1137, 1072, 1028, 943, 871, 820,
tion ½a 2D4
ꢂ
+6.49 (c 0.75, DMSO).
4.5.18. [5-Fluoro-1-(2-fluoro-4-{[(2S)-4-methyl-3,4-dihydro-2H-
1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (5c)
787, 746, 611, 577, 462 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+13.37 (c
43% yield; TLC Rf = 0.40 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.53 (t, J = 8.1 Hz, 1H), 7.17–7.12 (m, 2H), 6.92–6.68 (m,
7H), 4.75–4.64 (m, 1H), 4.31–4.17 (m, 2H), 3.64 (s, 2H), 3.39 (dd,
J = 11.4, 3.0 Hz, 1H), 3.26 (dd, J = 11.4, 6.3 Hz, 1H), 2.92 (s, 3H),
2.34 (s, 3H); MS (APCI, Neg, 20 V) m/z 505 (MꢁH)ꢁ; IR (KBr)
3070, 2929, 2871, 2612, 1688, 1619, 1578, 1505, 1472, 1460,
1395, 1372, 1358, 1324, 1297, 1277, 1224, 1197, 1172, 1118,
1065, 1040, 997, 923, 850, 809, 745, 594 cmꢁ1; Optical rotation
0.75, DMSO).
4.5.14. [1-(2,3-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4h)
56% yield; TLC Rf = 0.63 (CHCl3/MeOH/AcOH, 9:1:0.1); 1H NMR
(300 MHz, CDCl3) d 7.47 (d, J = 7.8 Hz, 1H), 7.21–7.16 (m, 2H),
7.06–7.04 (m, 2H), 6.91–6.83 (m, 2H), 6.77–6.67 (m, 3H),
4.73–4.66 (m, 1H), 4.30–4.15 (m, 2H), 3.71 (s, 2H), 3.45–3.28 (m,
2H), 2.93 (s, 3H), 2.31 (s, 3H), 2.24 (s, 6H); MS (APCI, Neg, 20 V)
m/z 497 (MꢁH)ꢁ; IR (KBr) 3044, 2927, 2819, 2626, 2564, 1685,
1607, 1592, 1580, 1504, 1476, 1458, 1378, 1352, 1317, 1301,
1268, 1243, 1227, 1188, 1132, 1112, 1091, 1043, 957, 913, 866,
800, 764, 741, 687, 608, 567, 524, 514, 489, 469, 459 cmꢁ1; Optical
½
a 2D4
+8.67 (c 0.75, DMSO).
ꢂ
4.5.19. [5-Fluoro-2-methyl-1-(3-methyl-4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetic acid (5d)
76% yield; TLC Rf = 0.48 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.61–7.51 (m, 2H), 7.15 (dd, J = 8.7, 2.1 Hz, 1H), 6.98–6.67
(m, 7H), 4.74–4.66 (m, 1H), 4.32 (dd, J = 9.6, 5.1 Hz, 1H), 4.22 (dd,
J = 9.6, 6.3 Hz, 1H), 3.69 (s, 2H), 3.42 (dd, J = 11.4, 2.4 Hz, 1H),
3.31 (dd, J = 11.4, 6.6 Hz, 1H), 2.92 (s, 3H), 2.39 (s, 3H), 2.27 (s,
3H); MS (APCI, Neg, 20 V) m/z 501 (MꢁH)ꢁ; IR (KBr) 3067, 2927,
2822, 1711, 1685, 1604, 1561, 1504, 1472, 1459, 1415, 1395,
1373, 1354, 1317, 1265, 1225, 1204, 1122, 1069, 1041, 993, 910,
rotation ½a 2D3
ꢂ
+11.53 (c 0.75, DMSO).
4.5.15. [1-(2,5-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-3-
yl]acetic acid (4i)
12% yield; TLC Rf = 0.35 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.48 (t, J = 7.5 Hz, 1H), 7.21–7.15 (m, 2H), 7.09–6.99 (m,
2H), 6.92–6.82 (m, 2H), 6.75–6.68 (m, 3H), 4.73–4.66 (m, 1H),
4.33–4.18 (m, 2H), 3.72 (s, 2H), 3.42 (dd, J = 11.4, 2.7 Hz, 1H),
3.31 (dd, J = 11.4, 6.6 Hz, 1H), 2.93 (s, 3H), 2.34 (s, 3H), 2.26 (s,
3H), 2.17 (s, 3H); MS (APCI, Neg, 20 V) m/z 497 (MꢁH)ꢁ; IR (KBr)
2927, 2871, 1769, 1710, 1684, 1656, 1637, 1610, 1570, 1543,
1505, 1473, 1458, 1376, 1350, 1319, 1262, 1224, 1176, 1132,
1104, 1040, 986, 915, 859, 805, 744, 684, 671, 657, 638, 628,
618, 612, 603, 596, 578, 563, 552, 486, 478, 468, 462 cmꢁ1; Optical
808, 759, 744, 638, 600, 459 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+10.81
(c 0.75, DMSO).
4.5.20. [5-Fluoro-1-(3-methoxy-4-{[(2S)-4-methyl-3,4-dihydro-
2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-2-methyl-1H-indol-
3-yl]acetic acid (5e)
43% yield; TLC Rf = 0.58 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.33 (d, J = 1.8 Hz, 1H), 7.27 (dd, J = 8.7, 1.8 Hz, 1H),
7.19–7.14 (m, 1H), 7.00–6.66 (m, 7H), 4.77–4.69 (m, 1H), 4.38–
4.22 (m, 2H), 3.88 (s, 3H), 3.70 (s, 2H), 3.43 (dd, J = 11.4, 2.7 Hz,
1H), 3.30 (dd, J = 11.4, 6.3 Hz, 1H), 2.91 (s, 3H), 2.40 (s, 3H); MS
(APCI, Neg, 20 V) m/z 517 (MꢁH)ꢁ; IR (KBr) 3069, 2933, 2871,
2622, 2360, 1710, 1684, 1598, 1506, 1472, 1459, 1417, 1395,
1373, 1355, 1316, 1272, 1244, 1218, 1177, 1129, 1070, 1028,
911, 850, 808, 753, 669, 648, 582, 501, 471, 461 cmꢁ1; Optical rota-
rotation ½a 2D4
ꢂ
+11.08 (c 0.88, DMSO).
4.5.16. [5-Fluoro-2-methyl-1-(2-methyl-4-{[(2S)-4-methyl-3,4-
dihydro-2H-1,4-benzoxazin-2-yl]methoxy}benzoyl)-1H-indol-
3-yl]acetic acid (5a)
50% yield; TLC Rf = 0.43 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.31 (d, J = 8.7 Hz, 1H), 7.14 (dd, J = 8.7, 2.4 Hz, 1H), 7.01
(dd, J = 9.3, 4.5 Hz, 1H), 6.92–6.74 (m, 5H), 6.74–6.66 (m, 2H),
4.72–4.62 (m, 1H), 4.29 (dd, J = 9.9, 4.8 Hz, 1H), 4.18 (dd, J = 9.9,
6.3 Hz, 1H), 3.68 (s, 2H), 3.41 (dd, J = 11.4, 2.1 Hz, 1H), 3.27 (dd,
J = 11.4, 6.6 Hz, 1H), 2.92 (s, 3H), 2.31 (s, 3H), 2.30 (s, 3H); MS
(APCI, Neg, 20 V) m/z 501 (MꢁH)ꢁ; IR (KBr) 2927, 2611, 2360,
2249, 1865, 1686, 1604, 1573, 1504, 1471, 1459, 1384, 1372,
tion ½a 2D3
ꢂ
+9.73 (c 0.75, DMSO).
4.5.21. [1-(2,3-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-5-fluoro-2-methyl-1H-
indol-3-yl]acetic acid (5f)
41% yield; TLC Rf = 0.54 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.18–7.02 (m, 3H), 6.92–6.67 (m, 6H), 4.73–4.66 (m, 1H),

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4585
4.30–4.16 (m, 2H), 3.66 (s, 2H), 3.43 (dd, J = 11.4, 2.7 Hz, 1H), 3.32
(dd, J = 11.4, 6.9 Hz, 1H), 2.93 (s, 3H), 2.27 (s, 3H), 2.24 (s, 3H), 2.23
(s, 3H); MS (APCI, Neg, 20 V) m/z 515 (MꢁH)ꢁ; IR (KBr) 3041, 2929,
2871, 2372, 1710, 1686, 1649, 1606, 1593, 1561, 1543, 1505, 1472,
1459, 1372, 1355, 1317, 1302, 1269, 1243, 1225, 1203, 1149, 1110,
1086, 1042, 994, 915, 852, 803, 766, 743, 617, 603, 593, 567, 548,
(26 mL, 150 mmol). After agitating for 1 h at room temperature,
the reaction mixture was filtered, washed with CH2Cl2 (ꢀ5) to af-
ford 22 (8.55 g, quant); IR (KBr) 3568, 3058, 3024, 2920, 2849,
1950, 1807, 1767, 1742, 1686, 1601, 1568, 1505, 1492, 1446,
1415, 1347, 1314, 1265, 1197, 1154, 1125, 1056, 1039, 1013,
902, 827, 755, 698, 632, 551 cmꢁ1
.
527, 505, 487, 459 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+8.63 (c 0.75,
DMSO).
4.9. Polymer supported [1-(4-hydroxy-2-methylbenzoyl)-2-
methyl-1H-indol-3-yl]acetate (23)
4.5.22. [1-(2,5-Dimethyl-4-{[(2S)-4-methyl-3,4-dihydro-2H-1,4-
benzoxazin-2-yl]methoxy}benzoyl)-5-fluoro-2-methyl-1H-
indol-3-yl]acetic acid (5g)
To an agitated mixture of 22 (8.55 g, 7.5 mmol) in CH2Cl2
(95 mL) was added piperidine (5 mL, mmol) at room temperature.
After agitating for 30 min at room temperature, the reaction mix-
ture was filtered, washed with CH2Cl2 (ꢀ5), dried in vacuo to afford
23 (8.23 g, quant); IR (KBr) 3564, 3058, 3025, 2922, 2850, 1951,
1808, 1739, 1682, 1601, 1492, 1446, 1348, 1310, 1219, 1153,
39% yield; TLC Rf = 0.44 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.19–7.11 (m, 2H), 7.06–6.96 (m, 1H), 6.93–6.67 (m, 6H),
4.74–4.64 (m, 1H), 4.31 (dd, J = 9.6, 3.9 Hz, 1H), 4.20 (dd, J = 9.6,
6.6 Hz, 1H), 3.68 (s, 2H), 3.42 (dd, J = 11.7, 3.0 Hz, 1H), 3.31 (dd,
J = 11.7, 6.3 Hz, 1H), 2.93 (s, 3H), 2.31 (s, 3H), 2.25 (s, 3H), 2.18
(s, 3H); MS (APCI, Neg, 20 V) m/z 515 (MꢁH)ꢁ; IR (KBr) 3041,
2927, 2870, 1711, 1685, 1609, 1578, 1505, 1472, 1459, 1407,
1396, 1372, 1354, 1321, 1279, 1261, 1242, 1224, 1200, 1153,
1137, 1120, 1102, 1063, 1040, 981, 921, 849, 803, 766, 743, 655,
1055, 1013, 858, 825, 754, 697, 631, 553 cmꢁ1
.
4.10. General procedure for the preparation of N-(4-(alkyloxy)-
2-methylbenzoyl)-2-methylindol-3-acetic acids (6–11)
593, 572, 459 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+7.95 (c 0.75, DMSO).
4.10.1. (1-{4-[(2R)-2,3-Dihydro-1,4-benzodioxin-2-ylmethoxy]-
2-methylbenzoyl}-2-methyl-1H-indol-3-yl)acetic acid (6)
To an agitated mixture of 23 (300 mg, 0.3 mmol) in CH2Cl2
(4 mL) were added a solution of 24 (299 mg, 1.8 mmol) in CH2Cl2
(1 mL), Ph3P (472 mg, 1.8 mmol) and DEAD (40% in toluene,
0.82 mL, 1.8 mmol) at room temperature. After agitating for 2 h
at room temperature, the reaction mixture was filtered to collect
insoluble substance, which was washed with CH2Cl2 (ꢀ5), and
dried in vacuo.
4.6. Benzyl {1-[4-(acetyloxy)-2-methylbenzoyl]-2-methyl-1H-
indol-3-yl}acetate (20)
To a stirred solution of 19a (2.3 g, 8.3 mmol) in CH2Cl2 (15 mL)
were added 20 M NaOH aq (2.1 mL, 41.5 mmol), TBACl (231 mg,
0.83 mmol) and then a solution of 4-acetoxy-2-methylbenzoyl
chloride (2.3 g, 10.8 mmol) in CH2Cl2 (5 mL). After stirring for 1 h
at room temperature, the reaction mixture was quenched with
water and extracted with EtOAc (ꢀ2). The combined organic layers
were washed with water, brine, dried over Na2SO4 and concen-
trated in vacuo. The resulting residue was purified by column
chromatography on silica gel to yield 20 (3.5 g, 92%); TLC
Rf = 0.49 (n-hexane/EtOAc, 7:3); 1H NMR (300 MHz, CDCl3) d 7.47
(d, J = 7.5 Hz, 1H), 7.39 (d, J = 8.4 Hz, 1H), 7.36–7.25 (m, 6H),
7.22–7.15 (m, 1H), 7.13–7.06 (m, 2H), 7.02 (dd, J = 8.4, 2.4 Hz,
1H), 5.13 (s, 2H), 3.73 (s, 2H), 2.33 (s, 3H), 2.28 (s, 6H).
To a stirred suspension of the insoluble substance described
above in CH2Cl2 (2.1 mL) and CF3CH2OH (0.6 mL) was added acetic
acid (0.3 mL) at room temperature. After agitating for 2 h at room
temperature, insoluble substance was removed by filtration. The
filtrate was concentrated in vacuo to give a crude product, which
was purified by column chromatography on silica gel to yield 6
(50 mg, 35%); TLC Rf = 0.44 (CHCl3/MeOH, 9:1); 1H NMR
(300 MHz, CDCl3) d 7.48 (d, J = 7.8 Hz, 1H), 7.33 (d, J = 8.4 Hz, 1H),
7.18 (t, J = 7.8 Hz, 1H), 7.10–6.84 (m, 7H), 6.79 (dd, J = 8.4, 2.4 Hz,
1H), 4.64–4.56 (m, 1H), 4.42 (dd, J = 11.7, 2.4 Hz, 1H), 4.35–4.18
(m, 3H), 3.71 (s, 2H), 2.34 (s, 3H), 2.32 (s, 3H); MS (APCI, Neg,
20 V) m/z 470 (MꢁH)ꢁ; IR (KBr) 3049, 2927, 2612, 2360, 2251,
1892, 1709, 1684, 1638, 1604, 1572, 1542, 1494, 1475, 1457,
1440, 1351, 1316, 1266, 1243, 1176, 1113, 1047, 909, 849, 747,
4.7. {1-[4-(Acetyloxy)-2-methylbenzoyl]-2-methyl-1H-indol-3-
yl}acetic acid (21)
To a stirred solution of 20 (3.5 g, 8.0 mmol) in EtOAc (20 mL)
was added 20% Pd(OH)2/C (820 mg) at room temperature. The
resulting suspension was vigorously stirred for 30 min at room
temperature under hydrogen atmosphere. The reaction mixture
was filtered through a pad of Celite, and the filtrate was concen-
trated in vacuo to give 21, which was used for the next reaction
without further purification; TLC Rf = 0.56 (CHCl3/MeOH, 9:1); 1H
669, 647, 637, 542, 463 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+19.85 (c
0.75, DMSO).
4.10.2. (1-{4-[(3R)-2,3-Dihydro-1-benzofuran-3-ylmethoxy]-2-
methylbenzoyl}-2-methyl-1H-indol-3-yl)acetic acid (7a)
50% yield; TLC Rf = 0.49 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.51–7.45 (m, 1H), 7.35–7.28 (m, 2H), 7.25–6.80 (m, 7H),
6.79–6.73 (m, 1H), 4.73 (t, J = 8.4 Hz, 1H), 4.55 (dd, J = 9.3, 5.1 Hz,
1H), 4.21 (dd, J = 9.3, 5.7 Hz, 1H), 4.08 (t, J = 8.4 Hz, 1H), 4.02–
3.90 (m, 1H), 3.72 (s, 2H), 2.34 (s, 3H), 2.32 (s, 3H); MS (APCI,
Neg, 20 V) m/z 454 (MꢁH)ꢁ; IR (KBr) 3050, 2927, 2616, 2564,
2362, 1711, 1685, 1603, 1571, 1498, 1483, 1458, 1351, 1316,
1240, 1174, 1128, 1111, 1026, 968, 853, 815, 749, 636, 605,
NMR (300 MHz, CDCl3)
d 7.48 (d, J = 7.5 Hz, 1H), 7.40 (d,
J = 8.4 Hz, 1H), 7.24–7.17 (m, 1H), 7.15–7.00 (m, 4H), 3.72 (s, 2H),
2.33 (s, 3H), 2.31 (s, 6H).
4.8. Polymer supported {1-[4-(acetyloxy)-2-methylbenzoyl]-2-
methyl-1H-indol-3-yl}acetate (22)
To an agitated solution of 21 (2.63 g, 7.5 mmol) and DIPEA
(5.2 mL, 30 mmol) in CH2Cl2 (60 mL) was added a polymer sup-
ported 2-chloro triphenylmethyl chloride (1.22 mmol/g, 100–200
mesh, 1% DVB) (6.15 g, 7.5 mmol) at room temperature. After
agitating for 2.5 h at room temperature, the reaction mixture was
filtered to collect insoluble substance, which was washed with
CH2Cl2 (ꢀ5).
453 cmꢁ1; Optical rotation ½a 2D3
ꢂ
+19.15 (c 0.75, DMSO).
4.10.3. [1-(4-{[(3R)-5-Fluoro-2,3-dihydro-1-benzofuran-3-
yl]methoxy}-2-methylbenzoyl)-2-methyl-1H-indol-3-yl]acetic
acid (7b)
68% yield; TLC Rf = 0.53 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.48 (d, J = 7.8 Hz, 1H), 7.32 (d, J = 8.4 Hz, 1H), 7.22–6.97
(m, 4H), 6.84 (d, J = 2.4 Hz, 1H), 6.76 (dd, J = 8.4, 2.4 Hz,
1H), 6.67–6.57 (m, 2H), 4.80–4.64 (m, 2H), 4.44–4.36 (m, 1H),
To a suspension of the insoluble substance in CH2Cl2 (70 mL)
were added a solution of AcOH (4.3 mL, 75 mmol) and DIPEA

4586
M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4.18–4.03 (m, 2H), 3.72 (s, 2H), 2.34 (s, 3H), 2.32 (s, 3H); MS (APCI,
Neg, 20 V) m/z 472 (MꢁH)ꢁ; IR (KBr) 3052, 2965, 2928, 1712, 1684,
1625, 1603, 1498, 1475, 1457, 1351, 1316, 1241, 1174, 1129, 1111,
1015, 956, 857, 816, 779, 764, 756, 746, 618, 478 cmꢁ1; Optical
558, 546, 537, 524, 514, 502, 489, 477, 470, 464 cmꢁ1; Optical
rotation ½a 2D4
ꢂ
-6.49 (c 0.75, DMSO).
4.11. General procedure for the preparation of 25a–b
rotation ½a 2D3
ꢂ
+27.43 (c 0.66, DMSO).
4.11.1. (2S)-2-[(2-Bromophenoxy)methyl]oxirane (31a)
To a stirred solution of 2-bromophenol 30a (3.5 g, 20 mmol) in
DMF (20 mL) were added Cs2CO3 (9.9 g, 30 mmol) and (S)-(+)-glyc-
idyl 3-nitrobenzenesulfonate (3.9 g, 20 mmol) at room tempera-
ture under argon atmosphere, and stirring was continued for 1 h.
The reaction mixture was quenched with water and extracted with
EtOAc (ꢀ2). The combined organic layers were washed with water,
brine, dried over Na2SO4 and concentrated in vacuo to give 31a
(3.95 g, 86%), which was used for the next reaction without further
purification; TLC Rf = 0.50 (n-hexane/EtOAc, 5:1).
4.10.4. (1-{4-[(2S)-2,3-Dihydro-1-benzofuran-2-ylmethoxy]-2-
methylbenzoyl}-2-methyl-1H-indol-3-yl)acetic acid (8)
34% yield; TLC Rf = 0.56 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.47 (d, J = 7.8 Hz, 1H), 7.32 (d, J = 8.7 Hz, 1H), 7.24–7.10
(m, 3H), 7.10–6.94 (m, 2H), 6.92–6.75 (m, 4H), 5.24–5.12 (m,
1H), 4.27 (dd, J = 10.5, 6.3 Hz, 1H), 4.17 (dd, J = 10.5, 4.2 Hz, 1H),
3.71 (s, 2H), 3.41 (dd, J = 15.6, 9.6 Hz, 1H), 3.16 (dd, J = 15.6,
7.2 Hz, 1H), 2.34 (s, 3H), 2.31 (s, 3H); MS (APCI, Neg, 20 V) m/z
454 (MꢁH)ꢁ; IR (KBr) 3050, 2927, 2562, 1734, 1710, 1685, 1603,
1571, 1499, 1481, 1458, 1351, 1316, 1228, 1173, 1129, 1111,
1047, 1026, 982, 905, 868, 747, 711, 607, 469, 458 cmꢁ1; Optical
4.11.2. (3R)-2,3-Dihydro-1-benzofuran-3-ylmethanol (25a)
To a stirred solution of the above-described crude product 31a
(3.92 g, 17 mmol) in THF (20 mL) was added dropwise a 1.6 M
solution of n-butyl lithium in hexane (11.3 mL, 17.9 mmol) at
ꢁ78 °C. After stirring for 1 h at ꢁ78 °C and 1 h while gradually
warmed up to room temperature, the reaction mixture was
quenched with 1 M HCl aq and extracted with EtOAc (ꢀ2). The
combined organic layers were washed with brine, dried over
Na2SO4 and concentrated in vacuo to give a crude product, which
was purified by column chromatography on silica gel to yield
25a (3.24 g, quant); TLC Rf = 0.70 (n-hexane/EtOAc, 1:1); 1H NMR
(300 MHz, CDCl3) d 7.24–7.12 (m, 2H), 6.90–6.78 (m, 2H), 4.64 (t,
J = 9.0 Hz, 1H), 4.48 (dd, J = 9.0, 5.1 Hz, 1H), 3.86–3.77 (m, 2H),
3.70–3.58 (m, 1H).
rotation ½a 2D3
ꢂ
+37.42 (c 0.59, CHCl3).
4.10.5. {1-[4-(1,3-Benzodioxol-2-ylmethoxy)-2-methylbenzoyl]-
2-methyl-1H-indol-3-yl}acetic acid (9)
47% yield; TLC Rf = 0.50 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.48 (d, J = 7.5 Hz, 1H), 7.34 (d, J = 8.7 Hz, 1H), 7.24–6.94
(m, 3H), 6.92–6.84 (m, 5H), 6.79 (dd, J = 8.7, 2.1 Hz, 1H), 6.48 (t,
J = 3.9 Hz, 2H), 4.33 (t, J = 3.9 Hz, 1H), 3.72 (s, 2H), 2.34 (s, 3H),
2.32 (s, 3H); MS (APCI, Neg, 20 V) m/z 456 (MꢁH)ꢁ; IR (KBr)
1711, 1684, 1604, 1484, 1457, 1352, 1316, 1230, 1176, 1108,
742 cmꢁ1
.
4.10.6. (2-Methyl-1-{2-methyl-4-[(1-methyl-2,3-dihydro-1H-
indol-2-yl)methoxy]benzoyl}-1H-indol-3-yl)acetic acid (10a)
32% yield; TLC Rf = 0.50 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d7.54–6.48 (m, 11H), 4.26 (dd, J = 9.6. 5.7 Hz, 1H), 4.21–4.04
(m, 2H), 3.90–3.80 (m, 1H), 3.72 (s, 2H), 3.27 (dd, J = 15.3, 8.4 Hz,
1H), 2.91 (s, 3H), 2.34 (s, 3H), 2.33 (s, 3H); MS (APCI, Neg, 20 V)
m/z 467 (MꢁH)ꢁ; IR (KBr) 3050, 2926, 2857, 1710, 1684, 1604,
1569, 1497, 1487, 1475, 1457, 1351, 1316, 1242, 1174, 1128,
1113, 1042, 984, 941, 854, 815, 746, 715, 611, 537, 526, 492,
According to the same procedure as described above for the
conversion of 30a to 25a, 30b was converted to 25b.
4.11.3. (2S)-2-[(2-Bromo-4-fluorophenoxy)methyl]oxirane
(31b)
93% yield; TLC Rf = 0.57 (n-hexane/EtOAc, 2:1).
4.11.4. [(3R)-5-Fluoro-2,3-dihydro-1-benzofuran-3-yl]methanol
(25b)
461 cmꢁ1
.
34% yield; TLC Rf = 0.35 (n-hexane/EtOAc, 2:1); 1H NMR
(300 MHz, CDCl3) d 7.10 (dd, J = 8.1, 6.0 Hz, 1H), 6.63–6.53 (m,
2H), 4.67 (t, J = 9.0 Hz, 1H), 4.59 (dd, J = 9.0, 5.1 Hz, 1H), 4.00–
3.74 (m, 3H).
4.10.7. (1-{4-[(1-Ethyl-2,3-dihydro-1H-indol-2-yl)methoxy]-2-
methylbenzoyl}-2-methyl-1H-indol-3-yl)acetic acid (10b)
45% yield; TLC Rf = 0.55 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.48 (d, J = 7.8 Hz, 1H), 7.33 (d, J = 8.4 Hz, 1H), 7.22–6.97
(m, 5H), 6.86 (d, J = 2.1 Hz, 1H), 6.77 (dd, J = 8.7, 2.1 Hz, 1H), 6.66
(t, J = 7.8 Hz, 1H), 6.47 (d, J = 7.8 Hz, 1H), 4.30–4.04 (m, 3H), 3.72
(s, 2H), 3.47–3.22 (m, 3H), 2.89 (dd, J = 16.2, 7.2 Hz, 1H), 2.34 (s,
3H), 2.33 (s, 3H), 1.17 (t, J = 7.5 Hz, 3H); MS (APCI, Neg, 20 V) m/z
481 (MꢁH)ꢁ; IR (KBr) 3048, 2966, 2927, 2871, 2558, 2353, 1732,
1683, 1604, 1540, 1486, 1457, 1416, 1395, 1374, 1351, 1316,
1241, 1174, 1128, 1109, 1024, 940, 856, 808, 744, 712, 682, 612,
4.12. General procedure for the preparation of 28b, 29
4.12.1. Ethyl 1-ethylindoline-2-carboxylate (33a)
To a stirred solution of 32a (150 mg, 0.9 mmol) in DMF (3 mL)
were added Cs2CO3 (1.2 g, 3.7 mmol) and ethyl iodide (0.28 mL,
3.5 mmol) at room temperature. After stirring for 3 h at 40 °C,
The reaction mixture was diluted with water and extracted with
EtOAc (ꢀ2). The combined organic layers were washed with
water, brine, dried over Na2SO4 and concentrated in vacuo to
give 33a, which was used for the next reaction without further
purification; TLC Rf = 0.78 (n-hexane/EtOAc, 7:3); 1H NMR
(300 MHz, CDCl3) d 7.11–7.00 (m, 2H), 6.66 (t, J = 7.5 Hz, 1H),
6.48 (d, J = 7.5 Hz, 1H), 4.30–4.18 (m, 3H), 3.41–3.18 (m, 3H),
3.12 (dd, J = 15.9, 8.7 Hz, 1H), 1.30 (t, J = 7.2 Hz, 3H), 1.14 (t,
J = 7.2 Hz, 3H).
580 cmꢁ1
.
4.10.8. [1-(4-{[(2R)-1-Ethyl-2,3-dihydro-1H-indol-2-yl]
methoxy}-2-methylbenzoyl)-2-methyl-1H-indol-3-yl]acetic
acid (11)
36% yield; TLC Rf = 0.55 (CHCl3/MeOH, 9:1); 1H NMR (300 MHz,
CDCl3) d 7.48 (d, J = 7.5 Hz, 1H), 7.33 (d, J = 8.4 Hz, 1H), 7.22–6.96
(m, 5H), 6.90–6.74 (m, 2H), 6.66 (t, J = 7.5 Hz, 1H), 6.47 (d,
J = 8.1 Hz, 1H), 4.26–4.04 (m, 3H), 3.72 (s, 2H), 3.57–3.16 (m, 3H),
2.89 (dd, J = 15.3, 7.2 Hz, 1H), 2.35 (s, 3H), 2.33 (s, 3H), 1.17 (t,
J = 6.9 Hz, 3H); MS (APCI, Neg, 20 V) m/z 481 (MꢁH)ꢁ; IR (KBr)
3049, 2970, 2928, 2869, 2563, 2345, 2233, 1892, 1709, 1685,
1604, 1570, 1486, 1458, 1351, 1316, 1242, 1175, 1129, 1111,
1024, 945, 863, 820, 764, 746, 725, 711, 696, 666, 625, 612, 599,
4.12.2. (1-Ethyl-2,3-dihydro-1H-indol-2-yl)methanol (28b)
To a stirred suspension of lithium aluminium hydride (42 mg,
1.1 mmol) in THF (3 mL) was added a solution of the above-
described crude product 33a (0.90 mmol) in THF (3 mL) at room
temperature. After stirring for 30 min at room temperature, the
reaction mixture was diluted with ether and quenched with brine

M. Iwahashi et al. / Bioorg. Med. Chem. 19 (2011) 4574–4588
4587
Ki ¼ IC50=ð1 þ ½Lꢂ=KdÞ
under cooling. The resulting mixture was stirred for additional
10 min at room temperature and dried over MgSO4. Insoluble sub-
stance was removed by filtration and the filtrate was evaporated to
give a crude product, which was purified by column chromatogra-
phy on silica gel to yield 28b (90 mg, 56% in two steps); TLC
Rf = 0.38 (n-hexane/EtOAc, 7:3); 1H NMR (300 MHz, CDCl3) d
7.11–7.04 (m, 2H), 6.68 (td, J = 7.8, 0.9 Hz, 1H), 6.51 (d, J = 7.8 Hz,
1H), 3.92–3.84 (m, 1H), 3.83–3.73 (m, 1H), 3.68–3.58 (m, 1H),
3.38–3.12 (m, 2H), 3.06 (d, J = 9.9 Hz, 2H), 1.86 (dd, J = 9.9, 2.1 Hz,
1H), 1.12 (t, J = 7.2 Hz, 3H).
½Lꢂ : Concentration of radioligand
Kd : Dissociation constant of radiolabeled ligand towards
the prostanoid receptors
5.2. Measurement of the mDP receptor antagonist activity
To confirm that test compounds antagonize the mDP receptor
and to estimate potencies of antagonism for the mDP receptor, a
functional assay was performed by measuring PGD2-stimulated
changes in intracellular second messenger cAMP as an indicator
of receptor function.
According to the same procedure as described above for the
conversion of 33a to 28b, 32b was converted to 29.
For the assessment of the antagonist activity of test compounds, a
suspension of CHO cells expressing mDP receptor was seeded at a
cell density of 1 ꢀ 105 cells per well and cultivated for 2 days. The
cells in each well were rinsed with minimum essential medium
4.12.3. Ethyl (2R)-1-ethylindoline-2-carboxylate (33b)
TLC Rf = 0.78 (n-hexane/EtOAc, 7:3); 1H NMR (300 MHz, CDCl3)
d 7.11–7.00 (m, 2H), 6.66 (t, J = 7.5 Hz, 1H), 6.48 (d, J = 7.5 Hz, 1H),
4.30–4.18 (m, 3H), 3.41–3.18 (m, 3H), 3.12 (dd, J = 15.9, 8.7 Hz, 1H),
1.30 (t, J = 7.2 Hz, 3H), 1.14 (t, J = 7.2 Hz, 3H).
(MEM), and MEM containing 2 lM of Diclofenac was added to each
well. The cells were incubated for approximately 10 min at 37 °C and
the culture medium was removed. The assay medium (MEM con-
taining 0.1% BSA, 1 mM IBMX and 2 lM Diclofenac) was added to
4.12.4. [(2R)-1-ethyl-2,3-dihydro-1H-indol-2-yl]methanol (29)
60% yield in two steps; TLC Rf = 0.38 (n-hexane/EtOAc, 7:3); 1H
NMR (300 MHz, CDCl3) d 7.11–7.04 (m, 2H), 6.68 (td, J = 7.8, 0.9 Hz,
1H), 6.51 (d, J = 7.8 Hz, 1H), 3.92–3.84 (m, 1H), 3.83–3.73 (m, 1H),
3.68–3.58 (m, 1H), 3.38–3.12 (m, 2H), 3.06 (d, J = 9.9 Hz, 2H), 1.86
(dd, J = 9.9, 2.1 Hz, 1H), 1.12 (t, J = 7.2 Hz, 3H).
each well and the cells were incubated for approximately 10 min
at 37 °C. The assay medium, assay medium containing 10 nM of
PGD2, or assay medium containing various concentrations of test
compounds and 10 nM of PGD2 was added to each well and the cells
were further incubated for 10 min at 37 °C. The reaction was termi-
nated by the addition of ice-cold trichloroacetic acid (TCA; 10 w/v%).
After centrifugation of the reaction mixture, TCA was extracted
by adding a mixture of tri-n-octylamine and chloroform (5:18 v/v)
to the resultant supernatant, mixing and re-centrifugation. The
cAMP level in the resultant aqueous layer (upper layer) was deter-
mined by enzymeimmunoassay using a cAMP assay kit (GE Health-
care UK Ltd). The relative responsiveness (%) of cAMP production
was calculated relative to the maximum increase in cAMP that oc-
curred in the absence of test compound (100%) to estimate of the
IC50 values.
5. Biological assay method
5.1. Prostanoid mEP1–4, mDP, hTP, mFP and hIP receptor
binding assay
Competitive binding studies were conducted using radiolabeled
ligands and membrane fractions prepared from Chinese hamster
ovary (CHO) cells stably expressing the respective prostanoid
receptors, mEP1, mEP2, mEP3a, mEP4, mDP, hTP mFP, and hIP.
Membranes from CHO cells expressing prostanoid receptors
were incubated with radioligands (2.5 nM of [3H]PGE2 for mEP1–
4; 2.5 nM of [3H]PGD2 for mDP; 5.0 nM [3H]-SQ29548 for hTP;
5.3. Inhibitory effects of 4a, 4b, 5b and 11 on ovalbumin-
induced vascular permeability in guinea pig conjunctiva
2.5 nM [3H]PGF2
a
for mFP; 5.0 nM of [3H]Iloprost for hIP) and
Male Hartley guinea pigs were sensitized by intraperitoneal
injection of a mixture of ovalbumin (OVA) (1 mg) and inactivated
Bordetella pertussis (5 ꢀ 109). Two weeks later, animals were chal-
the test compounds at various concentrations in assay buffer
(10 mM KH2PO4–KOH buffer containing 1 mM EDTA, 10 mM
MgCl2, and 100 mM NaCl, pH 6.0 for mEP1–4 and mFP; 25 mM
HEPES–NaOH buffer containing 1 mM EDTA, 5 mM MgCl2 and
10 mM MnCl2, pH 7.4 for mDP; 10 mM Tris–HCl buffer containing
100 mM NaCl, pH 7.5, for hTP; 50 mM Tris–HCl buffer containing
1 mM EDTA and 10 mM MgCl2, pH 7.5, for hIP). Incubation was car-
ried out at room temperature for 60 min except for mEP1, and mDP
(20 min), and hTP and hIP (30 min). The incubation was terminated
by filtration through Whatman GF/B filters. The filters were
washed with ice-cold buffer (10 mM KH2PO4–KOH buffer contain-
ing 100 mM NaCl, pH 6.0 for mEP1–4 and mFP; 10 mM Tris–HCl
buffer containing 100 mM NaCl and 0.01 w/v% BSA, pH 7.4 for
mDP; 10 mM Tris–HCl buffer containing 100 mM NaCl, pH 7.5,
for hTP and hIP), dried for 60 min at 60 °C and the radioactivity
on the filter was measured in 6 mL of liquid scintillation (ACSII)
mixture with a liquid scintillation counter. Nonspecific binding
was achieved by adding excess amounts of unlabeled PGE2 (for
mEP1–4), unlabeled PGD2 (for mDP), unlabeled SQ29548 (for
lenged by topical application of OVA (1%, 20 lL/eye) to the eye,
and then Evans blue dye (20 mg/kg, iv) was immediately injected
as a marker of plasma exudation. All antagonists were orally admin-
istered 1 h before the antigen challenge. After 30 min, the guinea
pigs were exsanguinated, and the eye tissue including conjunctiva
were extracted. Isolated conjunctiva was incubated in DMF (1 mL)
at 37 °C to extract the extravasated dye, and the incubation mixture
was centrifuged. The absorption of the supernatant at 620 nm was
determined, and the amount of Evans blue dye leaked into the tis-
sues was quantified by interpolation on the standard curve.
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