Design, synthesis, and structure-activity relationship (SAR) of N-[7-(4-hydroxyphenoxy)-6-methylindan-4-yl]malonamic acids as thyroid hormone receptor β (TRβ) selective agonists

Article abstract of DOI:10.1016/j.bmc.2012.12.002

Highly TRβ selective thyromimetics have several potential therapeutic applications. Based on the novel indane derivative KTA-439 with high receptor (TRβ) and organ (liver) selectivity, a series of thyroid hormone analogues were prepared, in which the isopropyl at the 3′-position was replaced with alkyl and aralkyl moieties of variable lengths and branches. Binding assays for these human TRs and reporter cell assays showed that 2-arylethyl derivatives had higher TRβ selectivity than KTA-439. KTA-574, a representative 2-arylethyl derivative, had TRβ specificity in a binding assay and exhibited full agonism in a reporter cell assay.

Full text of DOI:10.1016/j.bmc.2012.12.002

Bioorganic & Medicinal Chemistry 21 (2013) 592–607
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Design, synthesis, and structure–activity relationship (SAR) of
N-[7-(4-hydroxyphenoxy)-6-methylindan-4-yl]malonamic acids
as thyroid hormone receptor b (TRb) selective agonists
a
a
a
a
Hiroaki Shiohara a, , Tetsuya Nakamura , Norihiko Kikuchi , Tomonaga Ozawa , Akane Matsuzawa ,
⇑
Ryuichi Nagano a, Hideki Ohnota a, Takahide Miyamoto b, Kazuo Ichikawa c, Kiyoshi Hashizume d
a Central Research Laboratory, Kissei Pharmaceutical Co., Ltd, 4365-1, Kashiwabara, Hotaka, Azumino-city, Nagano 399-8304, Japan
b Miyamoto Clinic, 1-6, Ryojima, Matsumoto-city, Nagano 390-0848, Japan
c Ichikawa Clinic, 1548-2, Minamiminowa-Village, Kamiina-County, Nagano 399-4598, Japan
d Matsumoto Dental University Hospital, Shiojiri-city, Nagano 399-0781, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Highly TRb selective thyromimetics have several potential therapeutic applications. Based on the novel
indane derivative KTA-439 with high receptor (TRb) and organ (liver) selectivity, a series of thyroid hor-
mone analogues were prepared, in which the isopropyl at the 30-position was replaced with alkyl and
aralkyl moieties of variable lengths and branches. Binding assays for these human TRs and reporter cell
assays showed that 2-arylethyl derivatives had higher TRb selectivity than KTA-439. KTA-574, a repre-
sentative 2-arylethyl derivative, had TRb specificity in a binding assay and exhibited full agonism in a
reporter cell assay.
Received 4 November 2012
Revised 2 December 2012
Accepted 3 December 2012
Available online 11 December 2012
Keywords:
Thyromimetics
TRb selectivity
TRb specificity
Full agonism
Indane
Ó 2012 Elsevier Ltd. All rights reserved.
Malonamic acid
1. Introduction
activation of the TRa1 isoform, whereas most of the actions of
these hormones on the liver (e.g., lipid-lowering effects) and other
Thyroid hormones (THs) affect the growth, development, and
metabolism of most tissues.1–3 The active endogenous thyroid hor-
tissues are mediated through the activation of the TRb1 isoform.8
Thyromimetics that specifically target TRb have been shown to
reduce plasma cholesterol levels and prevent atherosclerosis by
promoting reverse cholesterol transport in an animal model. These
compounds may be useful as complements to statin therapy for
preventing cardiovascular disease.8–13 One report suggested that
TRb was a critical TR isoform for T3-induced proliferation of hepa-
tocytes and pancreatic acinar cells.14 Recently, a thyroid hormone
receptor b subtype-selective thyromimetic was found to be effica-
cious in both mouse and monkey hair growth models after topical
applications.15 These reports suggested that highly TRb selective
thyromimetics have potential therapeutic applications.
We previously found that novel indane derivatives 2a–2e
had higher organ (liver) selectivity than eprotirome (3). N-[7-(4-
Hydroxy-3-isopropylphenoxy)-6-methylindan-4-yl] malonamic
acid (KTA-439, 2a), a representative indane derivative, exhibited
higher liver selectivity than 3 in a cholesterol-fed rat model and
had the same high human TRb selectivity as 3 in a binding assay.16
We sought to improve the TRb selectivity of 2a. Thus, in the pres-
ent study, we investigated the SARs for N-[7-(4-hydroxyphenoxy)-
6-methylindan-4-yl] malonamic acids. We describe our discovery
process for a TRb-specific agonist.
mone 3,5,30-triiodo-
L-thyronine (T3; 1 in Fig. 1) was anticipated to
be a potent lipid-lowering agent, although it cannot be used ther-
apeutically for patients with dyslipidemias or those who are obese
or have metabolic syndrome because of its side effect of
tachycardia.4
There are two major subtypes of thyroid hormone receptors
(TRs),
genes.1 Differential processing of ribonucleic acid (RNA) results in
the formation of several isoforms of each gene. TR 1, TRb1, and
a (TRa) and b (TRb), which are encoded for by two different
a
TRb2 isoforms bind to THs and act as ligand-regulated transcription
factors.5 The TRb1 isoform is prevalent particularly in the liver and
to a lesser degree in the heart.6 The TRb2 isoform is expressed in
the hypothalamus, anterior pituitary gland, and the developing
brain.1,7 The TRa1 isoform is also widely distributed, although its
levels are generally lower than those of the TRb1 isoform. The liter-
ature suggests that most of the effects of THs on the heart (partic-
ularly on heart rate and rhythm) are mediated through the
⇑
Corresponding author. Tel.: +81 (0)263 82 8820; fax: +81 (0)263 82 8827.
E-mail address: hiroaki_shiohara@pharm.kissei.co.jp (H. Shiohara).
0968-0896/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2012.12.002

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
593
O
OH
5'
HO
R5
Y
6
5
4
4'
3'
HO
I
6' I
OH
1
2
O
NH2
A
1' O
3
2'
I
R5
A
Y
1
Me
Br
O
O
NHCOCH2 2a (KTA-439)
NHCOCH2 2b
Me CH2 NHCOCH2 2c
H
N
OH
HO
Br
O
Me
O
NHCO
2d
Me
O
CONHCH2 2e
O
O
Br
3
O
H
H
N
O
N
O
HO
O
HO
Cl
N
HO
O
N
O
O
O
NH
Cl
4
5
F
O
O
HO
O
HO
O
Br
OH
OH
NH2
O
Br
N
NH
6
7
Figure 1. Structures of T3 (1), previously reported indane derivatives (2a–2e), eprotirome (3), reported benzamide (4), axitirome (5), L-94901 (6), and GC-24 (7).
of 12 with the carboxylic acids 14a–d in the presence of Tf2O21
gave ketones 15a–d. Demethylation, reduction, and hydrolysis
gave the final target ligands 16a–d.
2. Strategy
It has been shown that substituent modifications on the outer
ring of the biphenyl ether skeleton can result in enhanced TRb
selectivity. Benzamide 417 with a bulky moiety at its 30-position
reportedly was 105-fold more TRb-selective compared with TRa,
3.2. In vitro effects on TRs of indane derivatives with an alkyl
substituent at their 30-positions
although its amide moiety appeared to be metabolically labile. Li-
ver-selective axitirome (5)18 and L-94901 (6)19 also have steric
bulk moieties at their 30-positions as does TRb-selective GC-24
(7),20 which has a benzyl moiety at its 30-position. We speculated
that these liver-selective compounds 5 and 6 might also be TRb
selective. Thus, we focused on designing indane derivatives based
on 2a, in which the isopropyl at the 30-position was replaced with
Table 1 summarizes the results of a radioligand binding assay
for hTR
transfected with hTR
downstream thyroid response element.
The natural ligand T3 bound to hTR
of 2.29 and 2.33 nM, respectively. The 30-H compound 13 was
weakly, but moderately selective for hTRb compared with hTR
a
and hTRb and a reporter cell assay using COS1 cells stably
a
or hTRb and a luciferase reporter gene
a
and hTRb with Ki values
either alkyl, aralkyl including
6-dihydropyridazin-3-yl]methyl.
a-OH-4-F-benzyl, or 3-[6-oxo-1,
a
in terms of binding. When the hydrogen atom at the 30-position
was replaced with an ethyl and an iPr group (16a, 2a), the bind-
ing affinities increased for both isoforms, which resulted in high
hTRb selectivity. The iPr compound 2a was more hTRb-selective
than ethyl compound 16a. Compounds 16b and 16c, which
had elongated iPr moieties with methylene and ethylene, exhib-
ited high hTRb selectivity. When a cyclohexyl group was added
at the 2-ethyl of 16a (16d), the binding affinities increased for
both isoforms, resulting in high hTRb selectivity. Compound 13
exhibited partial agonism, whereas the other compounds had full
agonism in a reporter cell assay, except for the most bulky 16d
that exhibited full agonism to TRLuc-b and partial agonism to
3. Results and discussion
3.1. Synthesis of indane derivatives with alkyl substituents at
their 30-positions
As outlined in Scheme 1, four indane derivatives with alkyl sub-
stituents at their 30-positions were prepared. The nitro compound
816 was coupled with salt 918 to give ether 10. Nitro group reduc-
tion with H2/Pd gave aniline 11, which when coupled with ethyl
malonyl chloride gave anilide 12. Demethylation and hydrolysis
with BBr3 gave 30-H ligand 13. Dehydrative Friedel–Crafts acylation
TRLuc-a.

594
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
O
NO2
NO2
O
O
a
b
I+
+
O
HO
O
-
BF4
8
9
10
O
O
CO2Et
CO2H
NH2
O
NH
HO
NH
c
d
O
O
O
11
12
13
R
RCO2H
14a-d
e
Me
iPr
14a
14b
O
O
14c
CO2Et
CO2H
O
R
NH
HO
R
NH
14d
f, g, h
O
O
15a-d
O
16a-d
R
R
Me
iPr
15a
15b
Me
iPr
16a
16b
15c
15d
16c
16d
Scheme 1. Synthetic route for preparing 16a–d. Reagents and conditions: (a) Cu, Et3N, CH2Cl2, 5 days; (b) H2/Pd-C, EtOAc; (c) ClCOCH2CO2Et, pyridine, CH2Cl2, 0 °C; (d) BBr3,
CH2Cl2; (e) Tf2O, CH2Cl2; (f) BCl3, CH2Cl2; (g) Et3SiH, TFA, CH2Cl2; and (h) NaOH, MeOH.
Table 1
3.3. Synthesis of indane derivatives with p-fluorobenzyl and 3-
Thyroid hormone receptor binding affinities (Ki) and reporter cell line potency
[6-oxo-1,6-dihydropyridazin-3-yl]methyl at their 30-positions
efficacies (% agonism) of compounds 1, 2a, 13, 16a–c, and 16d
H
We attempted to synthesize indane derivatives with a-OH-4-F-
HO
R1
N
OH
benzyl moiety in a manner similar to that described for 5. How-
ever, this moiety was so acid labile that the desired acid was
decomposed by itself (data not shown). Thus, we prepared a 4-F-
O
O
O
Compounds R1
Kia (nM)
hTR
a
/bb
% Agonismc
TRLuc-b TRLuc-a
benzyl compound rather than a
a-OH-4-F-benzyl compound. The
preparation of this compound and a 3-[6-oxo-1,6-dihydropyrida-
zin-3-yl] methyl compound is outlined in Scheme 2. Benzylation
of known 17 gave benzyl ether 18, which was treated with
(CF3CO2)3I prepared from I2 to give iodonium salt 19.14 Nitro com-
pound 8 was coupled with 19 to give ether 20. Nitro group reduc-
tion and debenzylation of 20 with H2/Pd gave aniline 21. Coupling
21 with ethyl malonyl chloride and alkaline hydrolysis gave target
ligand 22. Coupling 8 with known 23,22 hydrolysis, and demethyl-
ation gave oxopyridazine 24. Nitro group reduction of 24 gave ani-
line 25. Coupling 25 with ethyl malonyl chloride and alkaline
hydrolysis gave target ligand 26.
hTRb
a
1
2.29
1662
53.6
7.82
2.33
>10000
710
1
100
62
99
100
47
96
13
H
>6
13
22
16a
Et
iPr
2ad
172
113
110
16b
16c
39.9
1431
503
36
109
94
96
94
31.7
16
3.4. Design and synthesis of indane derivatives with an aralkyl
moiety at their 30-positions
16d
17.0
429
25
73
48
a
b
c
Values are means of two experiments. The variability was 25% on average.
Because indane derivatives with a bulky alkyl moiety at their
30-positions had high hTRb potency and selectivity, we designed
and synthesized other indane derivatives with aralkyl moieties to ob-
tain additional SAR information, as outlined in Scheme 3. Acylation
of 12 with carboxylic acids 27a–c gave ketones 28a–c. Demethyl-
ation of 28a gave phenol 29, which was hydrolyzed to give ketone
ligand 30. Reduction of 29 with NaBH(OAc)3 and hydrolysis gave
Selectivity (Ki hTRa)/(Ki hTRb).
Values at 10ꢀ5 M; T3 set at 100%.
d
Ref. 16.
the acid stable
hydrolysis of 28a–c gave the final target ligands 32a–c.
a-OH ligand 31. Demethylation, reduction, and

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
BnO
595
OBn
HO
BnO
I+
a
b
c
F
B
F
F
F
F
F
F
F
17
18
19
O
CO2H
BnO
NO2
HO
NH2
HO
NH
d
e, f
O
O
O
20
21
22
F
F
F
O
HO
NO2
O
Cl
c, g
I+
O
N
N
O
O
O
N
N
N
NH
Cl
O
24
23
Cl
O
O
CO2H
NH
NH2
HO
HO
O
O
d
e, f
N
N
NH
NH
25
26
O
O
Scheme 2. Synthetic route for preparing 22 and 26. Reagents and conditions: (a) BnBr, Cs2CO3, DMF; (b) (i) (CF3CO2)3I, CH2Cl2; (ii) NaBF4; MeOH; (c) 8, Cu, Et3N, CH2Cl2,
5 days; (d) H2/Pd-C, EtOAc; (e) ClCOCH2CO2Et, pyridine, CH2Cl2, 0 °C; (f) NaOH, MeOH; (g) (i) NaOAc; and (ii) HBr, AcOH.
3.5. In vitro effects of ligands on TRs
3.6. Design and synthesis of indane derivatives with a 2-
arylethyl moiety at their 30-positions
The in vitro results for indane derivatives with an aralkyl moi-
ety at their 30-positions are summarized in Table 2. The 4-F-benzyl
ligand 22 had high affinity for hTRb and moderate hTRb selectivity,
which was less than that of 5. Interestingly, although amino acid 6
with a 3-[6-oxo-1,6-dihydropyridazin-3-yl] methyl moiety had
Finally, we designed and synthesized indane derivatives with a
2-arylethyl moiety to obtain additional SAR information as out-
lined in Scheme 4. Acylation of 12 with carboxylic acids 33a–e
and reduction gave ethers 34a–e. Coupling 34a–e with ethyl mal-
onyl chloride and alkaline hydrolysis gave target ligands 35a–e.
Dehydrative Friedel–Crafts acylations of 12 with methoxyphen-
ylacetic acids were not successful because these acids reacted with
themselves. Thus, we synthesized 40a–c as follows. Phenyl ether
10 was treated with Cl2CHOCH3/TiCl4 to give aldehyde 36. A Wittig
reaction with 37a–c and reduction of the nitro group and olefin
with H2/Pd gave anilines 38a–c. Coupling 38a–c with ethyl malo-
nyl chloride gave esters 39a–c, which were demethylated and
hydrolyzed to give target ligands 40a–c.
high hTRa affinity and moderate hTRa selectivity, ligand 26 with
the same moiety had high hTRb affinity and moderate hTRb selec-
tivity. This suggested that the indane skeleton strongly preferred
hTRb over hTR
When a phenyl group was added at the 2-ethyl of 16a (32a), the
binding affinity for hTRb increased slightly and decreased for hTR
a.
a,
which resulted in high hTRb selectivity. Compound 32b that had an
elongated 4-F-benzyl moiety with the methylene of 22 also im-
proved hTRb selectivity. It is noteworthy that this is the first report
on hTRb-selective thyromimetics with phenethyl moieties. Substi-
tuting the OH group at the
a
-position of 32a as in 31 negatively
3.7. In vitro effects on TRs of the indane derivatives with a 2-
arylethyl moiety at their 3’-positions
affected both subtypes’ affinities. Ketone 30 and the phenylpropyl
compound 32c both exhibited decreased subtype affinity and effi-
cacy. Based on these results, we focused on the 2-arylethyl group
at the 30-position to improve hTRb selectivity.
The in vitro results for indane derivatives with a 2-arylethyl
moiety at their 30-positions are summarized in Table 3. Introducing

596
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
O
CO2H
HO
O
NH
O
Ar
n
C6H5
1
1
2
27a
27b
27c
30
4-F-C6H4
C6H5
c
CO2H
O
O
( )n = 1,2
CO2Et
CO2Et
Ar
27a-c
O
NH
HO
NH
b
12
O
O
O
O
a
(Ar = C6H5,
n = 1)
( )n = 1,2
Ar
28a-c
29
Ar
n
C6H5
1
28a
28b
28c
e, c
b, e, c
d, c
1
2
4-F-C H4
6
C6H5
O
O
CO2H
CO2H
HO
HO
NH
HO
NH
O
O
( )n = 1,2
Ar
32a-c
31
Ar
n
C6H5
1
1
2
32a
32b
32c
4-F-C6H4
C6H5
Scheme 3. Synthetic route for preparing 30, 31, and 32a–c. Reagents and conditions: (a) Tf2O, CH2Cl2; (b) BCl3, CH2Cl2; (c) NaOH, MeOH; (d) NaBH(OAc)3, THF; and (e) Et3SiH,
TFA, CH2Cl2.
an additional fluorine into the 4-F-phenyl ring at the 2- or 3-posi-
tion (35a, 35b) was tolerated for both subtypes as compared with
32b. Introducing chlorine (35c, 35d, 35e) into the phenyl ring neg-
atively affected the efficacies. These chlorine compounds had hTRb
affinity and hTRb selectivity similar to 32a, but exhibited partial
agonism. Interestingly, introducing OH into the phenyl ring at
the 2-position greatly improved hTRb selectivity sufficiently to re-
gard 40a (KTA-574) as a specific hTRb agonist. Introducing OH at
the 3- or 4-position also improved hTRb selectivity, but reduced
the efficacies (40b, 40c).
Table 2
Thyroid hormone receptor binding affinities (Ki) and reporter cell line potency
efficacies (% agonism) of compounds 5, 6, 22, 26, 30, 31, 32a, 32b, and 32c
H
HO
R1
N
OH
O
O
O
Compounds R1
Kia (nM)
a
/bb
% Agonismc
TRLuc-b TRLuc-a
hTRb
2.17
238
3.70
hTR
a
5
6
22
40.0 18
94
104
163
115
8.72 0.04 121
4. Modeling
4-F-C6C4CH2
30.5
91.8
8
7
92
79
H
Molecular modeling studies were performed to investigate the
cause of the high affinity and selectivity of 40a. Docking models
for 40a were constructed from the crystal structures of hTR–Triac
(41, Fig. 2) complexes23 and modified from the crystal structure
of an hTR–7 complex.20 Figure 3 shows the docking model for hTRb
with 40a. This model suggested that interactions occurred between
COOH of 40a and Arg-316/Arg-320 of hTRb and between CONH of
40a and Asn-331 of hTRb. Other hydrogen bonds were observed
between 40-OH of 40a and His-435 of hTRb, which was thought
to play an important role in agonist activity. These interactions
O
N
N
26
14.1
73
32a
32b
31
30
32c
PhCH2CH2
40.2
8.20
124
441
223
1638
228
6751
>10000
4866
41
28
54
86
90
68
52
67
61
6
4-F-C6C4CH2CH2
PhCH2CH(OH)
PhCH2CO
>23 23
22 15
PhCH2CH2CH2
2
a
b
c
Values are means of two experiments. The variability was 25% on average.
Selectivity (Ki hTR )/(Ki hTRb).
Values at 10ꢀ5 M; T3 set at 100%.
a

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
597
CO2H
b
O
O
CO2Et
CO2H
R"
O
NH
HO
NH
c, d
33a-e
12
a
O
O
35a-e
34a-e
R"
R"
R"
R"
R"
2,4-F
3,4-F
2-Cl
3-Cl
4-Cl
33a
33b
33c
33d
33e
2,4-F
3,4-F
2-Cl
3-Cl
4-Cl
34a
34b
34c
34d
34e
2,4-F
3,4-F
2-Cl
3-Cl
4-Cl
35a
35b
35c
35d
35e
+
PPh3
O
NH2
O
NO2
Br
37a-c
R'
e
g
O
O
10
O
f
36
38a-c
R'
R'
R'
2-OMe
3-OMe
4-OMe
37a
37b
37c
2-OMe
3-OMe
4-OMe
38a
38b
38c
O
O
CO2Et
CO2H
O
NH
HO
NH
h
c, d
O
O
39a-c
40a-c
R'
R"
R'
R"
2-OMe
3-OMe
4-OMe
39a
39b
39c
2-OH
3-OH
4-OH
40a
40b
40c
Scheme 4. Synthetic route for preparing 35a–e and 40a–c. Reagents and conditions: (a) Tf2O, CH2Cl2; (b) Et3SiH, TFA, CH2Cl2; (c) BBr3, CH2Cl2; (d) NaOH, MeOH; (e)
MeOCHCl2, TiCl4, CH2Cl2; (f) tBuOK, THF ; (g) H2/Pd-C, EtOAc; and (h) ClCOCH2CO2Et, pyridine, CH2Cl2, 0 °C.
were previously suggested between thyromimetics with malo-
namic acid and hTRb. Another hydrogen bond was observed be-
tween 2-OH of the phenethyl of 40a and Gly-344 of hTRb (Fig
3a). This interaction may have affected the high affinity with ago-
nist activity, although the same interaction was observed between
through hydrogen bonds and CH/p interactions. However, no dif-
ferences in interactions were observed in the phenyl ethyl part.
The cause of the high selectivity of 40a remained unclear.
In an X-ray study, Borngraeber et al. showed that the benzyl
group at the 30-position of the distal ring of the hTRb-selective
thyromimetic 7 bound to hTRb and bent helices 3 and 11.20 They
concluded that 4 of the 14 side chains responsible for proper reg-
istration of helix 12 moved by 1.6–4.0 Å without affecting its posi-
tion in the protein. It is known that substitutions on hormones can
reach toward and alter the position of helix 12, which affects
receptor activity. Ligand 7 was accepted because of the flexibility
of helix 11 in hTRb. They speculated that helix 11 was better
40a and hTRa (data not shown). Because the aromatic rings of an
indane phenyl ether are closed to aliphatic side chains of non-polar
amino residues, we considered the contributions of CH/
interactions.24,25
p
The CHPAI program was used to search for CH/
Several CH/ interactions were observed between an indane phe-
nyl ether and TRs. Figure 3b shows CH/ interactions between
p
interactions.26
p
p
40a and Leu-330, Ile-276, Leu-346, Met-442, and Phe-451 of hTRb.
packed in hTRa and that changes in the spatial volume near the li-
Thus, compound 40a interacted with thyroid hormone receptors
gand substitution were probably less tolerated in hTR
a.

598
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
Table 3
When 35c–e, 40b, or 40c binds to hTRb, helix 11 is moved so as
to change the position of helix 12 slightly with the interaction be-
tween His-435 and OH of the distal ring of ligands; thus, they ex-
hibit partial agonism.
Thyroid hormone receptor binding affinities (Ki) and reporter cell line potency
efficacies (% agonism) of compounds 32a, 32b, 35a–e, and 40a–c
H
We are confident that the hTRb-specificity of 40a is explained
HO
N
OH
not only by the type and size of its 30-position, but also because
O
O
our indane skeleton strongly prefers hTRb over hTRa, as 6 and 26
exhibited opposite selectivity.
O
R"
5. Conclusion
R00
Kia (nM)
a
/bb
% Agonismc
TRLuc-b TRLuc-a
We designed novel thyromimetics with high receptor (hTRb)
selectivity based on the novel indane derivative 2a in which
the isopropyl at the 30-position was replaced with alkyl and aral-
kyl moieties of variable lengths and branches. The results of a
binding assay for hTRs and a reporter cell assay revealed that
a previously unreported 2-arylethyl moiety affected higher hTRb
selectivity. KTA-574 (40a) with the 2-OH-C6H4CH2CH2 moiety at
the 30-position showed hTRb specificity in a binding assay and
exhibited full agonism in a reporter cell assay. Both the nature
of the hTRb-selective indane skeleton and the size/shape of the
2-OH-C6H4CH2CH2 moiety at the 30-position contributed to the
hTRb specificity of 40a.
Compounds
hTRb
40.2
8.20
12.0
6.00
hTR
a
32a
32b
35a
35b
35c
35d
35e
40a(KTA-574)
H
4-F
1638
228
201
40
28
17
31
27
33
41
>187
>123
>174
86
90
73
101
35
41
40
81
50
44
52
67
49
45
11
13
11
66d
53d
25
2,4-F
3,4-F
2-Cl
3-Cl
4-Cl
2-OH
3-OH
4-OH
186
30.0
41.0
40.0
53.5
81.3
57.6
814
1366
1645
>10000
>10000
>10000
40b
40c
a
b
c
Values are means of two experiments. The variability was 25% on average.
The results of this study should provide useful SAR information
for the further design of potent, hTRb-specific, thyromimetics for
therapeutic uses and deepen our understanding of thyroid hor-
mone actions.31
Selectivity (Ki hTRa)/(Ki hTRb).
Values at 10ꢀ5 M; T3 set at 100%.
Values at 10ꢀ4 M; T3 set at 100%.
d
6. Experimental
HO
I
I
OH
6.1. Chemistry: general
O
O
I
Uncorrected melting points were obtained with a Yanako MP-
3S Micro melting point apparatus. 1H and 13C NMR spectra were
recorded using a Bruker Avance II+ 400 or Avance III 600, and
chemical shifts were reported in parts per million (d) downfield
from tetramethylsilane used as the internal standard. Peak pat-
terns are shown using the following abbreviations: br, broad; s,
singlet; d, doublet; t, triplet; q, quartet; and m, multiplet. Mass
spectra (HRMS) were obtained with an Agilent Technologies
6520 Accurate-Mass Q-TOF apparatus. Silica gel 60F254-precoated
glass plates from Merck KgaA or aminopropyl silica gel (APS)-
precoated NH plates from Fuji Silysia Chemical Ltd were used for
thin layer chromatography (TLC). Flash or medium-pressure liquid
chromatography (MPLC) was performed using silica gel BW-350
from Fuji Silysia Chemical Ltd or APS Daisogel IR-60 (particle size:
41
Figure 2. Structure of Triac (41).
Polikarpov et al. performed Molecular Dynamics (MD) simula-
tions of thyroid hormones and suggested the contributions of
waters on selectivity, which is the basis of ligand dissociation,
and the stabilization of helix 12 in agonist conformation.23,27–29
This group showed that the TRb ligand-binding cavity (LBC) was
more extensive relative to TRa. Introducing a large group was al-
lowed in TRb, while it was not allowed in TR
a that had a smaller
LBC compared with that of TRb.30
25–40 lM) from Daiso Co., Ltd. All reagents and solvents were
We also performed MD simulations for TRs and 40a to explain
receptor selectivity, but obtained no positive results. However,
commercially available, unless otherwise indicated. Purchased re-
agents and solvents were used without further purification, unless
otherwise noted.
the hypotheses that helix 11 was better packed in hTR
TRb LBC was more extensive relative to TR are supported by
our results that the most bulky 16d in Table 1 exhibited full agon-
ism to TRLuc-b and partial agonism to TRLuc-
Based on our results, modeling, and these hypotheses, one rea-
son for the hTRb specificity of 40a is the following.
When 40a binds to hTRb, helix 11 is slightly moved. However,
this movement is not enough to break the interaction between
His-435 and OH of the distal ring of 40a and is not enough to
change the position of helix 12; thus, it exhibits full agonism.
a and that
a
6.1.1. 4-(4-Methoxyphenoxy)-5-methyl-7-nitroindane (10)
To a solution of 8 (6.00 g, 31.1 mmol) and 9 (17.0 g, 39.7 mmol)
in CH2Cl2 (150 mL), copper bronze (2.00 g, 31.5 mmol) and Et3N
(5.6 mL, 40.2 mmol) were added at room temperature. The
mixture was stirred at room temperature for 5 days. Insoluble
materials were removed by filtration. The filtrate was evaporated
to dryness under reduced pressure. The residue was purified by
column chromatography on silica gel (eluent: hexane/EtOAc
1/0–3/1) to give 10 (7.30 g, 79%) as a beige solid. Beige solid;
mp 88–89 °C (EtOAc–hexane); 1H NMR (400 MHz, CDCl3)
d: 2.00–2.15 (2H, m), 2.24 (3H, s), 2.60–2.70 (2H, m), 3.30–3.45
(2H, m), 3.77 (3H, s), 6.73 (2H, d, J = 9.1 Hz), 6.81 (2H, d,
J = 9.1 Hz), 7.96 (1H, s); 13C NMR (100 MHz, CDCl3) d: 16.1, 24.8,
30.3, 34.3, 55.7, 114.9, 116.5, 125.7, 130.8, 139.5, 141.8, 141.8,
a.
The hydrogen bond between Gly-344 and 40a and the CH/p inter-
action between Met-442/Phe-451 and 40a may have affected the
high affinity with agonist activity. In contrast, when 40a attempts
to bind to hTR
tion between His-435 and OH of the distal ring of 40a because
of the better packing of helix 11 in hTR ; thus, 40a cannot bind
to hTR
a, helix 11 is moved enough to break the interac-
a
a.

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
599
150.9, 154.7, 155.0; HRMS calcd for C17H18NO4 (M+H)+ 300.1230,
found 300.1231.
due was purified by column chromatography on silica gel (eluent:
hexane/EtOAc = 19/1–0/1) to give 15a (29 mg, 14%) as a beige solid.
Beige solid; mp 120–122 °C (EtOAc–hexane); 1H NMR (400 MHz,
CDCl3) d: 1.34 (3H, t, J = 7.1 Hz), 2.00–2.10 (2H, m), 2.14 (3H, s),
2.58 (3H, s), 2.60–2.70 (2H, m), 2.85–2.95 (2H, m), 3.49 (2H, s),
3.87 (3H, s), 4.27 (2H, q, J = 7.1 Hz), 6.80–6.90 (2H, m), 7.17 (1H,
d, J = 3.0 Hz), 7.79 (1H, s), 9.18 (1H, s); 13C NMR (100 MHz, CDCl3)
d: 14.1, 16.1, 24.9, 30.2, 30.4, 31.8, 41.2, 56.0, 62.0, 112.9, 116.4,
119.8, 121.9, 129.0, 129.9, 130.4, 134.4, 137.3, 146.1, 151.7,
153.7, 162.6, 170.4, 199.4; HRMS calcd for C24H28NO6 (M+H)+
426.1911, found 426.1915.
6.1.2. [7-(4-Methoxyphenoxy)-6-methylindan-4-yl]amine (11)
To a solution of 10 (1.68 g, 5.61 mmol) in EtOAc (50 mL), 10%
Pd/C (56% wet weight with water, 1.37 g, 0.566 mmol) was
added. The mixture was stirred overnight under a hydrogen
atmosphere at room temperature. Insoluble materials were re-
moved by filtration and washed with EtOAc. The filtrate was
evaporated to dryness under reduced pressure to give 11
(1.51 g, 100%) as a beige solid. Beige solid; mp 102–103 °C
(EtOAc–hexane); 1H NMR (400 MHz, CDCl3) d: 1.95–2.15 (5H,
m), 2.60–2.80 (4H, m), 3.44 (2H, br s), 3.75 (3H, s), 6.41 (1H,
s), 6.65–6.85 (4H, m); 13C NMR (100 MHz, CDCl3) d: 15.8, 24.9,
29.7, 30.3, 55.7, 114.6, 115.5, 115.6, 128.4, 129.9, 137.7, 139.0,
6.1.6. Ethyl N-[7-(3-isobutyryl-4-methoxyphenoxy)-6-
methylindan-4-yl]malonamate (15b)
The title compound was prepared from 12 and isobutyric acid
(14b) in a manner similar to that described for 15a as a white solid
(52%). White solid; mp 147–148 °C (EtOAc–hexane); 1H NMR
(400 MHz, CDCl3) d: 1.12 (6H, d, J = 7.0 Hz), 1.34 (3H, t,
J = 7.2 Hz), 2.00–2.10 (2H, m), 2.14 (3H, s), 2.65–2.70 (2H, m),
2.85–2.95 (2H, m), 3.43 (1H, heptet, J = 7.0 Hz), 3.49 (2H, s), 3.83
(3H, s), 4.27 (2H, q, J = 7.2 Hz), 6.80–6.85 (2H, m), 6.95–7.00 (1H,
m), 7.78 (1H, s), 9.19 (1H, br s); 13C NMR (100 MHz, CDCl3) d:
14.1, 16.1, 18.5, 24.9, 30.2, 30.4, 40.1, 41.2, 56.1, 61.0, 62.0, 112.6,
116.2, 118.5, 121.9, 129.8, 129.9, 130.4, 134.3, 137.3, 146.1,
151.8, 152.3, 162.6, 170.4, 207.5; HRMS calcd for C26H32NO6
(M+H)+ 454.2224, found 454.2223.
142.2, 152.7, 153.9; HRMS calcd for
270.1489, found 270.1485.
C
17H20NO2 (M+H)+
6.1.3. Ethyl N-[7-(4-methoxyphenoxy)-6-methylindan-4-
yl]malonamate (12)
To a solution of 11 (1.30 g, 4.83 mmol) and pyridine (0.585 mL,
7.23 mmol) in CH2Cl2 (50 mL), ethyl malonyl chloride (0.680 mL,
5.33 mmol) was added dropwise at 0 °C. The mixture was stirred
for 3 h at room temperature. After adding water (20 mL), the mix-
ture was partitioned between water and CH2Cl2. The organic layer
was washed with a saturated aqueous solution of NaHCO3 and
brine, dried over anhydrous MgSO4, and concentrated in vacuo.
The residue was purified by column chromatography on silica gel
(eluent: hexane/EtOAc = 19/1–0/1) to give 12 (1.21 g, 65%) as a
beige solid. Beige solid; mp 113–115 °C (EtOAc–hexane); 1H NMR
(400 MHz, CDCl3) d: 1.34 (3H, t, J = 7.1 Hz), 2.00–2.15 (2H, m),
2.16 (3H, s), 2.60–2.75 (2H, m), 2.80–2.95 (2H, m), 3.49 (2H, s),
3.76 (3H, s), 4.27 (2H, q, J = 7.1 Hz), 6.65–6.85 (4H, m), 7.77 (1H,
s), 9.20 (1H, s); 13C NMR (100 MHz, CDCl3) d: 14.1, 16.1, 24.9,
30.2, 30.4, 41.2, 55.7, 61.9, 114.6, 115.9, 121.9, 130.0, 130.1,
134.4, 137.4, 146.6, 152.1, 154.2, 162.6, 170.4; HRMS calcd for
6.1.7. Ethyl N-{7-[4-methoxy-3-(3-methylbutyryl)phenoxy]-6-
methylindan-4-yl}malonamate (15c)
The title compound was prepared from 12 and 3-methylbutyric
acid (14c) in a manner similar to that described for 15a as a pale
yellow solid (55%). Pale yellow solid; mp 151–153 °C (EtOAc–
hexane); 1H NMR (400 MHz, CDCl3) d: 0.90–1.00 (6H, m), 1.34
(3H, t, J = 7.1 Hz), 2.00–2.10 (2H, m), 2.14 (3H, s), 2.15–2.25 (1H,
m), 2.60–2.70 (2H, m), 2.75–2.95 (4H, m), 3.49 (2H, s), 3.84 (3H,
s), 4.27 (2H, q, J = 7.1 Hz), 6.80–6.85 (2H, m), 7.05–7.10 (1H, m),
7.78 (1H, s), 9.19 (1H, br s); 13C NMR (100 MHz, CDCl3) d: 14.1,
16.1, 22.7, 24.9, 25.0, 30.2, 30.4, 41.2, 52.6, 56.0, 62.0, 112.8,
116.2, 119.0, 121.9, 129.9, 130.0, 130.4, 134.3, 137.3, 146.1,
151.8, 152.9, 162.6, 170.4, 202.6; HRMS calcd for C27H34NO6
(M+H)+ 468.2381, found 468.2380.
C
22H26NO5 (M+H)+ 384.1805, found 384.1800.
6.1.4. N-[7-(4-Hydroxyphenoxy)-6-methylindan-4-
yl]malonamic acid (13)
To a solution of 12 (32 mg, 83.5 lmol) in CH2Cl2 (1 mL), a 1 M
solution of BBr3 in CH2Cl2 (0.500 mL, 0.500 mmol) was added drop-
wise at ꢀ78 °C. The mixture was stirred overnight at room temper-
ature. After adding water, the mixture was partitioned between
water and CH2Cl2. The organic layer was extracted with a saturated
aqueous solution of NaHCO3. The alkaline water layer was washed
with EtOAc, acidified with 2 M HCl, and extracted with EtOAc. The
organic layer was washed with water and brine and dried over
anhydrous MgSO4. The solvent was removed under reduced pres-
sure to give 13 (6 mg, 21%) as a white solid. White solid; mp
200–204 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, CDCl3/
CD3OD = 7/1) d: 1.95–2.10 (2H, m), 2.15 (3H, s), 2.60–2.70 (2H,
m), 2.80–2.90 (2H, m), 3.47 (2H, s), 6.60–6.75 (4H, m), 7.63 (1H,
s); 13C NMR (100 MHz, CDCl3/CD3OD = 7/1) d: 16.2, 25.1, 30.5,
30.5, 40.6, 116.1, 116.2, 122.7, 129.7, 130.1, 135.4, 137.7, 147.2,
151.1, 151.6, 164.4, 171.7; HRMS calcd for C19H20NO5 (M+H)+
342.1336, found 342.1341.
6.1.8. Ethyl N-{7-[3-(2-cyclohexylacetyl)-4-methoxyphenoxy]-6-
methylindan-4-yl}malonamate (15d)
The title compound was prepared from 12 and cyclohexylacetic
acid (14d) in a manner similar to that described for 15a as a pale
yellow solid (72%). Pale yellow solid; mp 154–155 °C (EtOAc–
hexane); 1H NMR (400 MHz, CDCl3) d: 0.90–1.30 (5H, m), 1.34
(3H, t, J = 7.1 Hz), 1.60–1.75 (5H, m), 1.80–1.95 (1H, m), 2.00–
2.10 (2H, m), 2.14 (3H, s), 2.60–2.70 (2H, m), 2.75–2.95 (4H, m),
3.49 (2H, s), 3.83 (3H, s), 4.27 (2H, q, J = 7.1 Hz), 6.80–6.90 (2H,
m), 7.00–7.10 (1H, m), 7.78 (1H, s), 9.19 (1H, s); 13C NMR
(100 MHz, CDCl3) d: 14.1, 16.1, 24.9, 26.2, 26.3, 30.2, 30.4, 33.4,
34.3, 41.2, 51.3, 56.1, 61.9, 112.8, 116.2, 119.0, 121.9, 129.9,
130.1, 130.4, 134.3, 137.3, 146.1, 151.8, 152.9, 162.6, 170.4,
202.6; HRMS calcd for
508.2687.
C
30H38NO6 (M+H)+ 508.2694, found
6.1.5. Ethyl N-[7-(3-acetyl-4-methoxyphenoxy)-6-methylindan-
4-yl]malonamate (15a)
6.1.9. N-[7-(3-Ethyl-4-hydroxyphenoxy)-6-methylindan-4-
yl]malonamic acid (16a)
To a solution of 12 (192 mg, 0.501 mmol) and acetic acid (14a)
(0.043 mL, 0.751 mmol) in CH2Cl2 (0.5 mL), trifluoromethanesulfo-
nic anhydride (0.125 mL, 0.762 mmol) was added. The mixture was
stirred for 15 h at room temperature. After adding water, the reac-
tion mixture was extracted with CH2Cl2. The organic layer was
dried over anhydrous MgSO4 and concentrated in vacuo. The resi-
To a solution of 15a (27 mg, 63.5 lmol) in CH2Cl2 (2 mL), a 1 M
solution of BCl3 in CH2Cl2 (0.20 mL, 0.200 mmol) was added drop-
wise at 0 °C. The mixture was stirred for 3 days at room tempera-
ture. After adding EtOH, the mixture was concentrated in vacuo.
The residue was purified by column chromatography on silica gel

600
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
Figure 3. (a, b) Docking model for hTRb with 40a constructed from the crystal structure of hTRb–41 complex (PDB code 3JZC23), which was modified from the crystal
structure of hTRb–7 complex (PDB code 1Q4X20). (a) Hydrogen bonds or polar interactions between hTRb and 40a. (b) CH/
interactions between hTRb and 40a.
p
(eluent: hexane/EtOAc = 19/1–0/1) to give ethyl N-[7-(3-acetyl-4-
hydroxyphenoxy)-6-methylindan-4-yl]malonamate (25 mg, 96%).
To a solution of ethyl N-[7-(3-acetyl-4-hydroxyphenoxy)-6-
6.1.10. N-[7-(4-Hydroxy-3-isobutylphenoxy)-6-methylindan-4-
yl]malonamic acid (16b)
The title compound was prepared from 15b in a manner similar
to that described for 16a as a white solid (54%, 3 steps). White so-
lid; mp 152–154 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz,
DMSO-d6) d: 0.75–0.90 (6H, m), 1.75–2.00 (3H, m), 2.06 (3H, s),
2.25–2.35 (2H, m), 2.50–2.60 (2H, m), 2.75–2.85 (2H, m), 3.38
(2H, s), 6.35–6.50 (2H, m), 6.60–6.70 (1H, m), 7.38 (1H, s), 8.83
(1H, s), 9.50 (1H, br s), 12.60 (1H, br s); 13C NMR (100 MHz,
DMSO-d6) d: 15.7, 22.2, 24.5, 27.9, 28.3, 30.0, 30.5, 43.2, 113.0,
115.5, 117.1, 123.2, 128.2, 128.4, 130.3, 136.0, 136.6, 146.3,
149.8, 149.8, 164.3, 169.6; HRMS calcd for C23H28NO5 (M+H)+
398.1962, found 398.1967.
methylindan-4-yl] malonamate (25 mg, 60.8 lmol) in CH2Cl2
(0.3 mL), triethylsilane (0.058 mL, 0.366 mmol) and TFA (0.3 mL)
were added. The mixture was stirred overnight at room tempera-
ture. After adding an aqueous solution of NaHCO3, the reaction
mixture was extracted with CH2Cl2. The organic layer was dried
over anhydrous MgSO4 and concentrated in vacuo. The residue
was purified by column chromatography on silica gel (eluent: hex-
ane/EtOAc = 19/1–0/1) to give ethyl N-[7-(3-ethyl-4-hydroxyphen-
oxy)-6-methylindan-4-yl]malonamate (10 mg, 41%).
To a solution of ethyl N-[7-(3-ethyl-4-hydroxyphenoxy)-6-
methylindan-4-yl]malonamate (10 mg, 25.2 lmol) in MeOH
(1 mL), an aqueous solution of 1 M NaOH (1 mL) was added. The
mixture was stirred for 30 min under an argon atmosphere at
60 °C. After adding 2 M HCl (0.5 mL) at 0 °C, the precipitate was
collected to give 16a (5 mg, 54%) as a white solid. White solid;
mp 118–120 °C (dec) (EtOH–H2O); 1H NMR (400 MHz, DMSO-d6)
d: 1.07 (3H, t, J = 7.4 Hz), 1.85–2.00 (2H, m), 2.05 (3H, s), 2.46
(2H, q, J = 7.4 Hz), 2.50–2.60 (2H, m), 2.70–2.85 (2H, m), 3.37
(2H, s), 6.30–6.40 (1H, m), 6.50–6.70 (2H, m), 7.38 (1H, s), 8.89
(1H, s), 9.56 (1H, br s), 12.62 (1H, br s); 13C NMR (100 MHz,
DMSO-d6) d: 14.1, 15.7, 22.8, 24.5, 30.0, 30.5, 43.2, 112.3, 115.3,
115.6, 123.2, 128.2, 130.3, 131.1, 136.0, 136.6, 146.2, 149.4,
150.1, 164.3, 169.5; HRMS calcd for C21H24NO5 (M+H)+ 370.1649,
found 370.1650.
6.1.11. N-{7-[4-Hydroxy-3-(3-methylbutyl)phenoxy]-6-
methylindan-4-yl}malonamic acid (16c)
The title compound was prepared from 15c in a manner similar
to that described for 16a as a white solid (43%, 3 steps). White so-
lid; mp 132–135 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz,
CDCl3/CD3OD = 7/1) d: 0.90–0.95 (6H, m), 1.40–1.65 (3H, m),
1.95–2.10 (2H, m), 2.15 (3H, s), 2.50–2.70 (4H, m), 2.80–2.90
(2H, m), 3.46 (2H, s), 6.35–6.45 (1H, m), 6.55–6.65 (2H, m),
7.63 (1H, s); 13C NMR (100 MHz, CDCl3/CD3OD = 7/1) d: 16.1,
22.6, 25.0, 28.0, 28.1, 30.3, 30.4, 38.9, 40.3, 112.7, 115.5, 116.8,
122.5, 129.5, 130.0, 130.6, 135.2, 137.6, 147.2, 148.7, 151.4,
164.3, 171.9; HRMS calcd for C24H30NO5 (M+H)+ 412.2118, found
412.2124.

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
601
6.1.12. N-{7-[3-(2-Cyclohexylethyl)-4-hydroxyphenoxy]-6-
methylindan-4-yl}malonamic acid (16d)
(2H, m), 7.25–7.40 (5H, m), 7.94 (1H, s); 13C NMR (100 MHz, CDCl3)
d: 16.1, 24.8, 30.4, 34.3, 35.5, 70.6, 112.8, 113.6, 114.9, 115.1, 118.3,
125.7, 127.3, 127.9, 128.5, 130.2, 130.3, 130.7, 131.6, 136.1, 137.0,
139.4, 141.8, 150.8, 151.8, 154.6; HRMS calcd for C30H27FNO4
(M+H)+ 484.1919, found 484.1910.
The title compound was prepared from 15d in a manner similar
to that described for 16a as a white solid (56%, 3 steps). White so-
lid; mp 166–167 °C(dec) (EtOAc–hexane); 1H NMR (400 MHz,
CDCl3/CD3OD = 7/1) d: 0.85–1.00 (2H, m), 1.05–1.35 (4H, m),
1.40–1.50 (2H, m), 1.55–1.80 (5H, m), 2.16 (3H, s), 2.50–2.75 (4H,
m), 2.80–2.90 (2H, m), 3.46 (2H, s), 6.35–6.45 (1H, m), 6.55–6.65
(2H, m), 7.63 (1H, s); 13C NMR (100 MHz, CDCl3/CD3OD = 7/1) d:
16.0, 24.9, 26.3, 26.7, 27.4, 30.2, 30.3, 33.3, 37.2, 37.5, 40.4,
112.7, 115.3, 116.6, 122.4, 129.4, 129.9, 130.7, 135.1, 137.5,
147.1, 148.7, 151.2, 164.3, 171.5; HRMS calcd for C27H34NO5
(M+H)+ 452.2431, found 452.2428.
6.1.16. 4-(7-Amino-5-methylindan-4-yloxy)-2-(4-fluorobenzyl)
phenol (21)
The title compound was prepared from 20 in a manner similar
to that described for 11 as a beige solid (98%). Beige solid; mp 169–
170 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, DMSO-d6) d:
1.80–1.95 (5H, m), 2.40–2.50 (2H, m), 2.55–2.65 (2H, m), 3.77
(2H, s), 4.61 (2H, s), 6.28 (1H, s), 6.35 (1H, dd, J = 3.0, 8.7 Hz),
6.49 (1H, d, J = 3.0 Hz), 6.67 (1H, d, J = 8.7 Hz), 7.00–7.10 (2H, m),
7.15–7.25 (2H, m), 8.97 (1H, s); 13C NMR (100 MHz, DMSO-d6) d:
15.6, 24.3, 29.5, 29.9, 34.4, 112.7, 114.3, 114.6, 115.5, 116.5,
126.9, 128.1, 130.2, 136.2, 137.2, 140.0, 140.9, 148.9, 151.0,
159.3, 161.7; HRMS calcd for C23H23FNO2 (M+H)+ 364.1707, found
364.1721.
6.1.13. 1-Benzyloxy-2-(4-fluorobenzyl)benzene (18)
Benzyl bromide (30 mL, 252 mmol) and Cs2CO3 (100 g,
307 mmol) were added to a solution of 2-(4-fluorobenzyl)phenol
(51.3 g, 254 mmol) in DMF (200 mL), and the mixture was stir-
red overnight at 80 °C. After adding water, the mixture was ex-
tracted with EtOAc. The organic layer was washed with water
and brine, and dried over anhydrous MgSO4. The solvent was re-
moved under reduced pressure to give 18 (66.6 g, 90%) as a col-
orless oil. Colorless oil; 1H NMR (600 MHz, CDCl3) d: 3.98 (2H, s),
5.04 (2H, s), 6.85–7.00 (4H, m), 7.05–7.20 (4H, m), 7.25–7.50
(5H, m); 13C NMR (150 MHz, CDCl3) d: 35.6, 69.9, 111.7, 114.8,
115.0, 120.8, 127.3, 127.6, 127.8, 128.5, 130.3, 130.3, 130.4,
6.1.17. N-{7-[3-(4-Fluorobenzyl)-4-hydroxyphenoxy]-6-
methylindan-4-yl}malonamic acid (22)
Ethyl N-{7-[3-(4-fluorobenzyl)-4-hydroxyphenoxy]-6-methy-
lindan-4-yl}malonamate was prepared from 21 in a manner simi-
lar to that described for 12.
To a solution of ethyl N-{7-[3-(4-fluorobenzyl)-4-hydroxyphen-
oxy]-6-methylindan-4-yl}malonamate (2.16 g, 4.52 mmol) in EtOH
(30 mL), an aqueous solution of 1 M NaOH (20 mL) was added. The
mixture was stirred for 30 min under an argon atmosphere at
60 °C. After adding 1 M HCl (20 mL) at 0 °C, the mixture was ex-
tracted twice with EtOAc. The organic layer was washed with
brine, dried over anhydrous MgSO4, and evaporated under reduced
pressure. The residue was crystallized from a small amount of
EtOH and water to give 22 (40%, 2 steps) as a white solid. White
solid; mp 162–164 °C (dec) (EtOH–H2O); 1H NMR (400 MHz,
DMSO-d6) d: 1.85–1.95 (2H, m), 2.03 (3H, s), 2.70–2.80 (2H, m),
3.25–3.35 (2H, m), 3.38 (2H, s), 3.79 (2H, s), 6.39 (1H, dd, J = 3.0,
8.7 Hz), 6.54 (1H, d, J = 3.0 Hz), 6.69 (1H, d, J = 8.7 Hz), 7.00–7.10
(2H, m), 7.15–7.25 (2H, m), 7.36 (1H, s), 9.08 (1H, s), 9.48 (1H, s),
12.60 (1H, s); 13C NMR (100 MHz, DMSO-d6) d: 15.7, 24.5, 30.0,
30.5, 34.4, 43.2, 113.2, 114.9, 115.6, 117.0, 123.2, 128.1, 128.4,
130.3, 130.4, 136.0, 136.6, 137.1, 146.2, 149.4, 150.1, 159.7,
136.7, 137.1, 156.4; HRMS calcd for
293.1336, found 293.1336.
C
20H18FO (M+H)+
6.1.14. Bis[4-benzyloxy-3-(4-fluorobenzyl)phenyl]iodonium
tetrafluoroborate (19)
Fuming nitric acid (85 mL, 1980 mmol) was added to Ac2O
(226 mL, 2400 mmol) under ice cooling. Then, iodine (76.1 g,
300 mmol) was added to this reaction mixture. Later, TFA
(175 mL, 2250 mmol) was added dropwise. After stirring for
1 h at room temperature, the mixture was evaporated to dryness
under reduced pressure at <35 °C. Ac2O (500 mL) and 18 (391 g,
1337 mmol) were then added to the residue. Later, TFA (100 mL)
was added dropwise under ice cooling. After stirring at 4 °C for
4 days, the reaction mixture was evaporated to dryness under
reduced pressure at <35 °C. MeOH (1000 mL), an aqueous solu-
tion of K2S2O5 (100 g/500 mL), and 4 M aqueous NaBF4
(1250 mL) were added in succession to the residue. The mixture
was stirred for 2 h. After the precipitate was aggregated, the
supernatant was decanted. The residue was dissolved in CH2Cl2
(1000 mL), and the organic layer was washed with a 4.5 M aque-
ous solution of NaBF4 (500 mL) and dried over anhydrous MgSO4.
The solvent was removed under reduced pressure, and the resi-
due was triturated with diethyl ether. Insoluble material was
collected by filtration to give 19 (263 g, 68%) as a white solid.
White solid; mp 131–132 °C (dec); 1H NMR (400 MHz, CDCl3)
d: 3.89 (4H, s), 5.03 (4H, s), 6.80–6.95 (6H, m), 7.00–7.10 (4H,
m), 7.20–7.40 (10H, m), 7.60 (2H, d, J = 2.5 Hz), 7.80 (2H, dd,
J = 2.5, 8.9 Hz); 13C NMR (100 MHz, CDCl3) d: 35.4, 70.5, 101.3,
115.1, 115.2, 115.3, 127.5, 128.5, 128.7, 130.6, 134.5, 135.2,
135.4, 135.6, 136.4, 159.9; HRMS calcd for C40H32F2IO2 (M)+
709.1410, found 709.1398.
161.3, 164.3, 169.6; HRMS calcd for
450.1711, found 450.1712.
C
26H25FNO5 (M+H)+
6.1.18. 6-[2-Hydroxy-5-(5-methyl-7-nitroindan-4-yloxy)
benzyl]-2H-pyridazine-3-one (24)
3-Chloro-6-[2-methoxy-5-(5-methyl-7-nitroindan-4-yloxy)
benzyl]pyridazine was prepared from 23 in a manner similar to
that described for 10.
A
mixture of 3-chloro-6-[2-methoxy-5-(5-methyl-7-
nitroindan-4-yloxy)benzyl]pyridazine (760 mg, 1.78 mmol), so-
dium acetate (50 mg, 0.610 mmol), and acetic acid (10 mL) was
heated for 2 h at reflux temperature under Ar. The reaction mixture
was evaporated to dryness under reduced pressure. After adding
water, the residue was stirred for 30 min. The mixture was ex-
tracted with dichloromethane. The organic layer was washed with
a saturated aqueous solution of NaHCO3 and brine and dried over
anhydrous MgSO4. The solvent was removed under reduced pres-
sure to give 6-[2-methoxy-5-(5-methyl-7-nitroindan-4-yloxy)ben-
zyl]-2H-pyridazine-3-one (707 mg).
To a solution of 6-[2-methoxy-5-(5-methyl-7-nitroindan-4-
yloxy)benzyl]-2H-pyridazine-3-one (707 mg) in acetic acid
(10 mL) hydrobromic acid (48%, 10 mL) was added. The mixture
was heated overnight at reflux temperature under Ar. After add-
ing water, the reaction mixture was extracted with dichloro-
6.1.15. 4-[4-Benzyloxy-3-(4-fluorobenzyl)phenoxy]-5-methyl-7-
nitroindane (20)
The title compound was prepared from 19 in a manner similar
to that described for 10 as a white solid (54%). White solid; mp
106–107 °C (EtOAc–hexane); 1H NMR (400 MHz, CDCl3) d: 2.00–
2.10 (2H, m), 2.22 (3H, s), 2.55–2.65 (2H, m), 3.30–3.40 (2H, m),
3.92 (2H, s), 4.99 (2H, s), 6.52 (1H, dd, J = 3.0, 8.8 Hz), 6.63 (1H, d,
J = 3.0 Hz), 6.79 (1H, d, J = 8.8 Hz), 6.85–6.95 (2H, m), 7.05–7.15

602
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
methane. The organic layer was washed with brine, dried over
anhydrous MgSO4, and concentrated in vacuo. The residue was
crystallized from a small amount of ethyl acetate to give 24
(270 mg, 36%, 3 steps) as a white solid. White solid; mp 246–
248 °C (EtOAc); 1H NMR (400 MHz, CDCl3) d: 2.00–2.10 (2H,
m), 2.22 (3H, s), 2.55–2.70 (2H, m), 3.30–3.40 (2H, m), 3.87
(2H, s), 6.51 (1H, dd, J = 3.0, 8.7 Hz), 6.66 (1H, d, J = 3.0 Hz),
6.73 (1H, d, J = 8.7 Hz), 6.86 (1H, d, J = 9.6 Hz), 7.30 (1H, d,
J = 9.6 Hz), 7.97 (1H, s); 13C NMR (100 MHz, CDCl3) d: 16.1,
24.8, 30.4, 34.3, 36.1, 115.6, 117.5, 117.7, 124.9, 125.7, 130.7,
131.2, 134.5, 139.4, 141.8, 141.9, 148.0, 149.7, 150.7, 154.5,
d, J = 9.0 Hz), 6.90 (1H, dd, J = 3.1, 9.0 Hz), 6.95–7.00 (2H, m), 7.06
(1H, d, J = 3.1 Hz), 7.10–7.20 (2H, m), 7.77 (1H, s), 9.18 (1H, s);
13C NMR (150 MHz, CDCl3) d: 14.1, 16.1, 24.9, 30.2, 30.3, 41.2,
49.2, 56.0, 62.0, 112.9, 115.1, 115.2, 116.5, 120.0, 121.9, 128.6,
129.8, 130.5, 130.8, 131.2, 134.3, 137.3, 146.0, 151.8, 153.1,
162.6, 170.4, 199.3; HRMS calcd for
520.2130, found 520.2124.
C
30H31FNO6 (M+H)+
6.1.23. Ethyl N-{7-[4-methoxy-3-(3-phenylpropionyl)phenoxy]-
6-methylindan-4-yl}malonamate (28c)
The title compound was prepared from 12 and 3-phenylpropi-
onic acid (27c) in a manner similar to that described for 15a as a
pale yellow solid (76%). Pale yellow solid; mp 107–109 °C
(EtOAc–hexane); 1H NMR (400 MHz, CDCl3) d: 1.34 (3H, t,
J = 7.1 Hz), 2.00–2.10 (2H, m), 2.14 (3H, s), 2.60–2.70 (2H, m),
2.85–2.95 (2H, m), 2.99 (2H, t, J = 7.7 Hz), 3.28 (2H, t, J = 7.7 Hz),
3.49 (2H, s), 3.83 (3H, s), 4.27 (2H, q, J = 7.1 Hz), 6.83 (1H, d,
J = 9.0 Hz), 6.87 (1H, dd, J = 3.0, 9.0 Hz), 7.13 (1H, d, J = 3.0 Hz),
7.15–7.30 (5H, m), 7.79 (1H, s), 9.20 (1H, br s); 13C NMR
(100 MHz, CDCl3) d: 14.1, 16.1, 24.9, 30.2, 30.4, 30.4, 41.2, 45.3,
56.0, 62.0, 112.8, 116.4, 119.6, 121.9, 125.9, 128.4, 128.5, 129.0,
129.9, 130.5, 134.3, 137.3, 141.7, 146.1, 151.8, 153.3, 162.6,
170.4, 201.2; HRMS calcd for C31H34NO6 (M+H)+ 516.2381, found
516.2381.
160.9; HRMS calcd for
394.1403.
C
21H20N3O5 (M+H)+ 394.1397, found
6.1.19. 6-[5-(7-Amino-5-methylindan-4-yloxy)-2-
hydroxybenzyl]-2H-pyridazine-3-one (25)
The title compound was prepared from 24 in a manner similar
to that described for 11 as a beige solid (100%). Beige solid; mp
141–144 °C (EtOAc–hexane); 1H NMR (400 MHz, CDCl3/CD3OD =
9/1) d: 1.95–2.10 (5H, m), 2.55–2.75 (4H, m), 3.85 (2H, s), 6.42
(1H, s), 6.52 (1H, dd, J = 2.9, 8.8 Hz), 6.57 (1H, d, J = 2.9 Hz), 6.69
(1H, d, J = 8.8 Hz), 6.85 (1H, d, J = 9.6 Hz), 7.27 (1H, d, J = 9.6 Hz);
13C NMR (100 MHz, CDCl3/CD3OD = 9/1) d: 15.7, 24.8, 29.5, 30.2,
35.4, 114.4, 115.8, 116.4, 116.8, 124.5, 128.8, 129.8, 130.0, 134.7,
137.5, 138.6, 142.3, 148.7, 152.0, 161.5; HRMS calcd for
C
21H22N3O3 (M+H)+ 364.1656, found 364.1652.
6.1.24. Ethyl N-[7-(4-hydroxy-3-phenylacetylphenoxy)-6-
methylindan-4-yl]malonamate (29)
6.1.20. N-{7-[4-Hydroxy-3-(6-oxo-1,6-dihydropyridazin-3-
ylmethyl)phenoxy]-6-methylindan-4-yl}malonamic acid (26)
The title compound was prepared from 25 in a manner sim-
ilar to that described for 22 as a white solid (9%, 2 steps). White
solid; mp 215–218 °C (dec) (EtOH); 1H NMR (400 MHz, DMSO-
d6) d: 1.85–1.95 (2H, m), 2.04 (3H, s), 2.70–2.80 (2H, m), 3.38
(2H, s), 3.74 (2H, s), 6.45 (1H, dd, J = 3.0, 8.6 Hz), 6.54 (1H, d,
J = 3.0 Hz), 6.72 (1H, d, J = 8.6 Hz), 6.79 (1H, d, J = 9.8 Hz), 7.22
(1H, d, J = 9.8 Hz), 7.38 (1H, s), 9.20 (1H, s), 9.52 (1H, br s),
12.73 (1H, br s); 13C NMR (100 MHz, DMSO-d6) d: 15.7, 24.5,
30.0, 30.5, 34.3, 43.2, 113.9, 115.8, 117.0, 123.2, 125.3, 128.1,
129.7, 130.5, 134.0, 136.0, 136.6, 146.1, 146.6, 149.6, 150.1,
To a solution of 28a (83 mg, 0.166 mmol) in CH2Cl2 (2 mL), a
1 M solution of BCl3 in CH2Cl2 (0.50 mL, 0.500 mmol) was added
dropwise at 0 °C. The mixture was stirred overnight at room tem-
perature. After adding EtOH, the mixture was concentrated in va-
cuo. The residue was purified by column chromatography on
silica gel (eluent: hexane/EtOAc = 3/1–1/1) to give 29 (49 mg,
61%) as a beige solid. Beige solid; mp 188–190 °C (EtOAc–hexane);
1H NMR (600 MHz, CDCl3) d: 1.34 (3H, t, J = 7.1 Hz), 2.00–2.10 (2H,
m), 2.09 (3H, s), 2.55–2.65 (2H, m), 2.85–2.95 (2H, m), 3.52 (2H, s),
4.09 (2H, s), 4.28 (2H, q, J = 7.1 Hz), 6.90 (1H, d, J = 9.1 Hz), 7.05
(1H, dd, J = 3.0, 9.1 Hz), 7.09 (1H, s), 7.20–7.35 (5H, m), 7.85 (1H,
s), 9.29 (1H, br s), 11.81 (1H, s); 13C NMR (150 MHz, CDCl3) d:
14.1, 16.0, 24.8, 30.2, 30.3, 41.1, 45.8, 62.0, 114.5, 118.4, 119.7,
122.0, 125.0, 127.1, 128.8, 129.1, 129.9, 130.6, 133.9, 134.3,
137.3, 145.8, 150.0, 157.7, 162.7, 170.5, 203.4; HRMS calcd for
160.2, 164.4, 169.6; HRMS calcd for
450.1660, found 450.1665.
C
24H24N3O6 (M+H)+
6.1.21. Ethyl N-[7-(4-methoxy-3-phenylacetylphenoxy)-6-
methylindan-4-yl]malonamate (28a)
C
29H30NO6 (M+H)+ 488.2068, found 488.2079.
The title compound was prepared from 12 and phenylacetic
acid (27a) in a manner similar to that described for 15a as a yel-
low solid (49%). Yellow solid; mp 123–124 °C (EtOAc–hexane);
1H NMR (600 MHz, CDCl3) d: 1.33 (3H, t, J = 7.1 Hz), 2.00–2.10
(2H, m), 2.11 (3H, s), 2.55–2.65 (2H, m), 2.80–2.90 (2H, m),
3.49 (2H, s), 3.86 (3H, s), 4.26 (2H, s), 4.27 (2H, q, J = 7.1 Hz),
6.84 (1H, d, J = 9.1 Hz), 6.89 (1H, dd, J = 3.0, 9.1 Hz), 7.06 (1H,
d, J = 3.0 Hz), 7.15–7.30 (5H, m), 7.77 (1H, s), 9.18 (1H, br s);
13C NMR (150 MHz, CDCl3) d: 14.1, 16.1, 24.9, 30.2, 30.3, 41.2,
50.1, 56.0, 61.9, 112.8, 116.5, 119.8, 121.9, 126.6, 128.3, 128.8,
129.7, 129.8, 130.4, 134.3, 135.1, 137.3, 146.0, 151.8, 153.1,
6.1.25. N-[7-(4-Hydroxy-3-phenylacetylphenoxy)-6-
methylindan-4-yl]malonamic acid (30)
To a solution of 29 (12 mg, 0.0246 mmol) in MeOH (1 mL), an
aqueous solution of 1 M NaOH (1 mL) was added. The mixture
was stirred for 30 min under Ar at 60 °C. After adding 2 M HCl
(0.55 mL) at 0 °C, the precipitate was collected to give 30 (10 mg,
89%) as a beige solid. Beige solid; mp 155–156 °C (dec) (EtOH–
H2O); 1H NMR (400 MHz, DMSO-d6) d: 1.85–2.00 (2H, m), 2.04
(3H, s), 2.45–2.55 (2H, m), 2.75–2.85 (2H, m), 3.41 (2H, s), 4.29
(2H, s), 6.92 (1H, d, J = 9.0 Hz), 7.01 (1H, dd, J = 3.0, 9.0 Hz), 7.10–
7.35 (6H, m), 7.44 (1H, s), 9.54 (1H, br s), 11.27 (1H, s), 12.62
(1H, br s); 13C NMR (100 MHz, DMSO-d6) d: 15.6, 24.5, 29.9, 30.5,
43.2, 46.3, 115.4, 119.0, 120.5, 123.4, 123.5, 126.5, 128.1, 128.4,
129.4, 130.8, 134.7, 136.2, 136.6, 145.5, 149.6, 155.2, 164.4,
169.5, 202.4; HRMS calcd for C27H26NO6 (M+H)+ 460.1755, found
460.1759.
162.6, 170.4, 199.6; HRMS calcd for
502.2224, found 502.2235.
C
30H32NO6 (M+H)+
6.1.22. Ethyl N-{7-[3-(4-fluorophenyl)acetyl-4-
methoxyphenoxy]-6-methylindan-4-yl}malonamate (28b)
The title compound was prepared from 12 and (4-fluoro-
phenyl)acetic acid (27b) in a manner similar to that described for
15a as a beige solid (55%). Beige solid; mp 134–135 °C (EtOAc–hex-
ane); 1H NMR (600 MHz, CDCl3) d: 1.34 (3H, t, J = 7.1 Hz), 2.00–2.10
(2H, m), 2.12 (3H, s), 2.60–2.65 (2H, m), 2.85–2.90 (2H, m), 3.49
(2H, s), 3.88 (3H, s), 4.23 (2H, s), 4.27 (2H, q, J = 7.1 Hz), 6.85 (1H,
6.1.26. N-{7-[4-Hydroxy-3-(1-hydroxy-2-phenylethyl)phenoxy]-
6-methylindan-4-yl}malonamic acid (31)
To a solution of 29 (19 mg, 0.039 mmol) in THF (10 mL), NaB-
H(OAc)3 (42 mg, 0.198 mmol) was added. The mixture was stir-
red overnight at room temperature. After adding water, the

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
603
reaction mixture was extracted with EtOAc. The organic layer
was washed with brine, dried over anhydrous MgSO4, and con-
centrated in vacuo. The residue was purified by column chroma-
tography on silica gel (eluent: hexane/EtOAc = 1/0–0/1) to give
ethyl N-{7-[4-hydroxy-3-(1-hydroxy-2-phenylethyl)phenoxy]-6-
methylindan-4-yl}malonamate (15 mg, 79%). The title compound
was prepared from ethyl N-{7-[4-hydroxy-3-(1-hydroxy-2-phen-
ylethyl)phenoxy]-6-methylindan-4-yl}malonamate in a manner
similar to that described for 30 as a white solid (99%). White so-
lid; mp 105–108 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz,
DMSO-d6) d: 1.85–2.00 (2H, m), 2.03 (3H, s), 2.60–2.95 (4H,
m), 4.90–5.10 (2H, m), 6.44 (1H, dd, J = 3.0, 8.7 Hz), 6.67 (1H,
d, J = 8.7 Hz), 6.71 (1H, d, J = 3.0 Hz), 7.10–7.25 (5H, m), 7.38
(1H, s), 9.08 (1H, br s), 9.59 (1H, br s); 13C NMR (100 MHz,
DMSO-d6) d: 15.8, 24.5, 30.0, 30.5, 43.3, 43.4, 68.3, 113.1,
113.4, 115.4, 123.1, 125.6, 127.7, 128.2, 129.4, 130.3, 133.1,
135.9, 136.7, 139.6, 146.2, 147.9, 150.1, 164.4, 169.6; HRMS
calcd for C27H28NO6 (M+H)+ 462.1911, found 462.1956.
(2,4-difluorophenyl)acetic acid (33a) in a manner similar to that
described for 15a (83%).
To a solution of ethyl N-{7-[3-(2,4-difluorophenyl)acetyl-4-
methoxyphenoxy]-6-methylindan-4-yl}malonamate
(100 mg,
0.186 mmol) in CH2Cl2 (0.37 mL), triethylsilane (0.104 mL,
0.651 mmol) and TFA (0.130 mL) were added. The mixture was
stirred overnight at room temperature. After adding water, the
reaction mixture was extracted with Et2O. The organic layer was
washed with water, a saturated aqueous solution of NaHCO3, and
brine. The organic layer was dried over anhydrous MgSO4 and con-
centrated in vacuo. The residue was purified by column chroma-
tography on silica gel (eluent: hexane/EtOAc = 5/1) to give 34a
(65 mg, 67%). White solid; mp 105–107 °C (EtOAc–hexane); 1H
NMR (400 MHz, CDCl3) d: 1.34 (3H, t, J = 7.2 Hz), 2.00–2.10 (2H,
m), 2.12 (3H, s), 2.55–2.65 (2H, m), 2.75–2.90 (6H, m), 3.49 (2H,
s), 3.76 (3H, s), 4.27 (2H, q, J = 7.2 Hz), 6.50 (1H, d, J = 2.9 Hz),
6.53 (1H, dd, J = 2.9, 8.9 Hz), 6.65–6.75 (3H, m), 6.95–7.05 (1H,
m), 7.75 (1H, s), 9.18 (1H, s); 13C NMR (100 MHz, CDCl3) d: 14.1,
16.1, 24.9, 28.5, 30.2, 30.3, 31.0, 41.2, 55.8, 61.9, 103.2, 103.4,
103.5, 110.5, 110.7, 111.0, 112.9, 117.2, 121.8, 130.0, 130.1,
130.7, 131.2, 134.3, 137.4, 146.6, 151.6, 152.2, 162.6, 170.5; HRMS
calcd for C30H32F2NO5 (M+H)+ 524.2243, found 524.2252.
6.1.27. N-[7-(4-Hydroxy-3-phenethylphenoxy)-6-methylindan-
4-yl]malonamic acid (32a)
The title compound was prepared from 28a in a manner similar
to that described for 16a as a white solid (35%, 3 steps). White so-
lid; mp 155–156 °C (dec) (EtOH–H2O); 1H NMR (400 MHz, DMSO-
d6) d: 1.85–2.00 (2H, m), 2.01 (3H, s), 2.40–2.50 (2H, m), 2.70–2.85
(6H, m), 3.38 (2H, s), 6.38 (1H, dd, J = 3.0, 8.7 Hz), 6.43 (1H, d,
J = 3.0 Hz), 6.68 (1H, d, J = 8.7 Hz), 7.10–7.30 (5H, m), 7.36 (1H, s),
8.99 (1H, br s), 9.49 (1H, br s), 12.62 (1H, br s); 13C NMR
(100 MHz, DMSO-d6) d: 15.7, 24.5, 29.9, 30.5, 31.7, 34.9, 43.2,
112.9, 115.4, 116.6, 123.2, 125.6, 128.0, 128.1, 128.3, 128.6,
130.3, 136.0, 136.6, 141.7, 146.2, 149.6, 149.9, 164.3, 169.6; HRMS
calcd for C27H28NO5 (M+H)+ 446.1962, found 446.1944.
6.1.31. Ethyl N-(7-{3-[2-(3,4-difluorophenyl)ethyl]-4-
methoxyphenoxy}-6-methylindan-4-yl)malonamate (34b)
The title compound was prepared from 12 and (3,4-difluoro-
phenyl)acetic acid (33b) in a manner similar to that described for
34a as a white solid (59%, 2 steps). White solid; mp 115–117 °C
(EtOAc–hexane); 1H NMR (600 MHz, CDCl3) d: 1.34 (3H, t,
J = 7.1 Hz), 2.00–2.10 (2H, m), 2.12 (3H, s), 2.55–2.65 (2H, m),
2.75–2.90 (6H, m), 3.50 (2H, s), 3.77 (3H, s), 4.27 (2H, q,
J = 7.1 Hz), 6.47 (1H, d, J = 3.0 Hz), 6.56 (1H, dd, J = 3.0, 8.8 Hz),
6.70 (1H, d, J = 8.8 Hz), 6.75–7.05 (3H, m), 7.75 (1H, s), 9.18 (1H,
br s); 13C NMR (150 MHz, CDCl3) d: 14.1, 16.0, 24.9, 30.2, 30.3,
32.3, 35.0, 41.2, 55.8, 61.9, 111.1, 113.0, 116.6, 116.8, 117.1,
117.2, 117.3, 121.9, 124.3, 130.0, 130.1, 130.5, 134.3, 137.4,
139.0, 146.5, 151.6, 152.1, 162.6, 170.4; HRMS calcd for
6.1.28. N-(7-{3-[2-(4-Fluorophenyl)ethyl]-4-hydroxyphenoxy}-
6-methylindan-4-yl)malonamic acid (32b)
The title compound was prepared from 28b in a manner similar
to that described for 16a as a beige solid (46%, 3 steps). Beige solid;
mp 82–85 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, CDCl3/
CD3OD = 7/1) d: 1.95–2.10 (2H, m), 2.12 (3H, s), 2.55–2.65 (2H,
m), 2.75–2.90 (6H, m), 3.48 (2H, s), 6.40–6.50 (2H, m), 6.63 (1H,
d, J = 8.6 Hz), 6.85–6.95 (2H, m), 7.05–7.15 (2H, m), 7.64 (1H, s);
13C NMR (100 MHz, CDCl3/CD3OD = 7/1) d: 16.1, 24.9, 30.3, 30.4,
32.5, 34.9, 40.2, 113.5, 114.8, 115.0, 115.7, 117.0, 122.4, 128.9,
129.84, 129.89, 135.0, 137.6, 147.0, 148.6, 131.4, 160.5, 162.1,
164.2, 171.5; HRMS calcd for C27H27FNO5 (M+H)+ 464.1868, found
464.1870.
C
30H32F2NO5 (M+H)+ 524.2243, found 524.2248.
6.1.32. Ethyl N-(7-{3-[2-(2-chlorophenyl)ethyl]-4-
methoxyphenoxy}-6-methylindan-4-yl)malonamate (34c)
The title compound was prepared from 12 and (2-chloro-
phenyl)acetic acid (33c) in a manner similar to that described for
34a as a pale yellow solid (74%, 2 steps). Pale yellow solid; mp
111–113 °C (EtOAc–hexane); 1H NMR (600 MHz, CDCl3) d: 1.34
(3H, t, J = 7.1 Hz), 2.00–2.10 (2H, m), 2.13 (3H, s), 2.60–2.70 (2H,
m), 2.80–2.90 (4H, m), 2.95–3.00 (2H, m), 3.49 (2H, s), 3.76 (3H,
s), 4.27 (2H, q, J = 7.1 Hz), 6.51 (1H, dd, J = 3.0, 8.8 Hz), 6.58 (1H,
d, J = 3.0 Hz), 6.68 (1H, d, J = 8.8 Hz), 7.05–7.15 (3H, m), 7.30–7.35
(3H, m), 7.76 (1H, s), 9.18 (1H, br s); 13C NMR (150 MHz, CDCl3)
d: 14.1, 16.1, 24.9, 30.2, 30.4, 30.5, 33.6, 41.2, 55.8, 62.0, 111.0,
112.7, 117.3, 121.8, 126.5, 127.2, 129.3, 130.0, 130.1, 130.6,
131.1, 134.0, 134.2, 137.5, 139.6, 146.6, 151.6, 152.3, 162.6,
170.5; HRMS calcd for C30H33ClNO5 (M+H)+ 522.2042, found
522.2049.
6.1.29. N-{7-[4-Hydroxy-3-(3-phenylpropyl)phenoxy]-6-
methylindan-4-yl}malonamic acid (32c)
The title compound was prepared from 28c in a manner similar
to that described for 16a as a beige solid (33%, 3 steps). Beige solid;
mp 111–115 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, DMSO-
d6) d: 1.70–2.00 (4H, m), 2.07 (3H, s), 2.40–2.60 (6H, m), 2.70–2.85
(2H, m), 3.40 (2H, s), 6.43 (1H, dd, J = 2.9, 8.7 Hz), 6.50 (1H, d,
J = 2.9 Hz), 6.69 (1H, d, J = 8.7 Hz), 7.05–7.30 (5H, m), 7.41 (1H, s),
8.92 (1H, br s), 9.51 (1H, s), 12.62 (1H, br s); 13C NMR (100 MHz,
DMSO-d6) d: 15.8, 24.5, 29.2, 30.0, 30.5, 30.8, 34.7, 43.2, 112.9,
115.5, 116.2, 123.2, 125.6, 128.2, 128.2, 128.3, 129.1, 130.4,
136.0, 136.6, 142.0, 146.3, 149.6, 150.1, 164.3, 169.6; HRMS calcd
for C28H30NO5 (M+H)+ 460.2118, found 460.2122.
6.1.33. Ethyl N-(7-{3-[2-(3-chlorophenyl)ethyl]-4-
methoxyphenoxy}-6-methylindan-4-yl)malonamate (34d)
The title compound was prepared from 12 and (3-chloro-
phenyl)acetic acid (33d) in a manner similar to that described for
34a as a white solid (54%, 2 steps). White solid; mp 135–136 °C
(EtOAc–hexane); 1H NMR (600 MHz, CDCl3) d: 1.34 (3H, t,
J = 7.2 Hz), 2.00–2.10 (2H, m), 2.13 (3H, s), 2.55–2.65 (2H, m),
2.80–2.90 (6H, m), 3.49 (2H, s), 3.77 (3H, s), 4.27 (2H, q,
J = 7.2 Hz), 6.51 (1H, d, J = 3.0 Hz), 6.55 (1H, dd, J = 3.0, 8.9 Hz),
6.70 (1H, d, J = 8.9 Hz), 6.95–7.05 (3H, m), 7.10–7.20 (3H, m),
6.1.30. Ethyl N-(7-{3-[2-(2,4-difluorophenyl)ethyl]-4-
methoxyphenoxy}-6-methylindan-4-yl)malonamate (34a)
Ethyl N-{7-[3-(2,4-difluorophenyl)acetyl-4-methoxyphenoxy]-
6-methylindan-4-yl}malonamate was prepared from 12 and

604
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
7.75 (1H, s), 9.18 (1H, br s); 13C NMR (150 MHz, CDCl3) d: 14.1,
16.1, 24.9, 30.2, 30.3, 32.2, 35.6, 41.2, 55.8, 62.0, 111.1, 112.9,
117.1, 121.9, 125.9, 126.8, 128.7, 129.4, 130.0, 130.1, 130.8,
133.9, 134.3, 137.5, 144.2, 146.5, 151.6, 152.2, 162.6, 170.5; HRMS
calcd for C30H33ClNO5 (M+H)+ 522.2042, found 522.2046.
dd, J = 2.4, 8.6 Hz), 6.54 (1H, d, J = 2.4 Hz), 6.63 (1H, d, J = 8.6 Hz),
7.05–7.20 (3H, m), 7.30–7.35 (1H, m), 7.63 (1H, s); 13C NMR
(100 MHz, CDCl3/CD3OD = 9/1) d: 16.1, 24.9, 30.3, 30.4, 30.5, 33.6,
40.2, 113.3, 115.7, 117.2, 122.4, 122.5, 126.7, 127.4, 128.7, 129.4,
130.0, 130.6, 133.9, 135.0, 137.6, 139.3, 147.0, 148.5, 151.5,
164.2, 171.3; HRMS calcd for C27H27ClNO5 (M+H)+ 480.1572, found
480.1577.
6.1.34. Ethyl N-(7-{3-[2-(4-chlorophenyl)ethyl]-4-
methoxyphenoxy}-6-methylindan-4-yl)malonamate (34e)
The title compound was prepared from 12 and (4-chloro-
phenyl)acetic acid (33e) in a manner similar to that described for
34a as a colorless amorphous solid (58%, 2 steps). Colorless amor-
phous solid; 1H NMR (600 MHz, CDCl3) d: 1.34 (3H, t, J = 7.2 Hz),
2.00–2.10 (2H, m), 2.12 (3H, s), 2.55–2.65 (2H, m), 2.80–2.90 (6H,
m), 3.50 (2H, s), 3.77 (3H, s), 4.27 (2H, q, J = 7.2 Hz), 6.48 (1H, d,
J = 2.9 Hz), 6.56 (1H, dd, J = 2.9, 8.7 Hz), 6.70 (1H, d, J = 8.7 Hz),
7.00–7.10 (2H, m), 7.15–7.25 (2H, m), 7.75 (1H, s), 9.18 (1H, br
s); 13C NMR (150 MHz, CDCl3) d: 14.1, 16.1, 24.9, 30.2, 30.3, 32.3,
35.2, 41.2, 55.8, 62.0, 111.1, 113.0, 117.1, 121.9, 128.2, 129.9,
130.0, 130.0, 130.8, 131.4, 134.3, 137.4, 140.5, 146.6, 151.6,
6.1.38. N-(7-{3-[2-(3-Chlorophenyl)ethyl]-4-hydroxyphenoxy}-
6-methylindan-4-yl)malonamic acid (35d)
The title compound was prepared from 34d in a manner similar
to that described for 35a as a beige solid (51%, 2 steps). Beige solid;
mp 154–155 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, CDCl3/
CD3OD = 9/1) d: 1.95–2.10 (2H, m), 2.12 (3H, s), 2.55–2.65 (2H,
m), 2.80–2.90 (6H, m), 3.46 (2H, s), 6.40–6.50 (2H, m), 6.60–6.70
(1H, m), 7.00–7.20 (4H, m), 7.63 (1H, s); 13C NMR (100 MHz,
CDCl3/CD3OD = 9/1) d: 16.0, 24.9, 30.2, 30.3, 32.2, 35.3, 40.2,
113.4, 115.5, 116.9, 122.4, 125.9, 126.8, 128.6, 128.7, 129.4,
129.4, 129.9, 133.8, 135.1, 137.5, 144.1, 147.0, 148.9, 151.1,
164.3, 171.5; HRMS calcd for C27H27ClNO5 (M+H)+ 480.1572, found
480.1573.
152.1, 162.6, 170.5; HRMS calcd for
522.2042, found 522.2045.
C
30H33ClNO5 (M+H)+
6.1.35. N-(7-{3-[2-(2,4-Difluorophenyl)ethyl]-4-
6.1.39. N-(7-{3-[2-(4-Chlorophenyl)ethyl]-4-hydroxyphenoxy}-
6-methylindan-4-yl)malonamic acid (35e)
hydroxyphenoxy}-6-methylindan-4-yl)malonamic acid (35a)
To a solution of 34a (69 mg, 0.132 mmol) in CH2Cl2 (2 mL), a
1 M solution of BBr3 in CH2Cl2 (0.527 mL, 0.527 mmol) was added
dropwise at ꢀ78 °C. The mixture was stirred overnight at room
temperature. After adding EtOH, the mixture was concentrated in
vacuo. The residue was purified by column chromatography on sil-
ica gel (eluent: hexane/EtOAc = 1/1) to give ethyl N-(7-{3-[2-(2,4-
difluorophenyl)ethyl]-4-hydroxyphenoxy}-6-methylindan-4-
yl)malonamate (63 mg, 94%).
The title compound was prepared from 34e in a manner similar
to that described for 35a as a beige solid (46%, 2 steps). Beige solid;
mp 125–128 °C (dec) (EtOAc–hexane); 1H NMR (600 MHz, DMSO-
d6) d: 1.85–1.95 (2H, m), 1.99 (3H, s), 2.40–2.50 (2H, m), 2.70–2.85
(6H, m), 3.38 (2H, s), 6.36 (1H, d, J = 3.0 Hz), 6.39 (1H, dd, J = 3.0,
8.6 Hz), 6.68 (1H, d, J = 8.6 Hz), 7.10–7.15 (2H, m), 7.25–7.30 (2H,
m), 7.36 (1H, s), 9.00 (1H, s), 9.49 (1H, s), 12.62 (1H, br s); 13C
NMR (150 MHz, DMSO-d6) d: 15.7, 24.5, 29.9, 30.5, 31.5, 34.0,
43.2, 113.1, 115.4, 116.6, 123.2, 127.9, 127.9, 128.0, 128.1, 128.2,
130.3, 136.0, 136.6, 140.6, 146.2, 149.6, 149.9, 164.3, 169.6; HRMS
calcd for C27H27ClNO5 (M+H)+ 480.1572, found 480.1540.
The title compound was prepared from ethyl N-(7-{3-[2-(2,4-
difluorophenyl)ethyl]-4-hydroxyphenoxy}-6-methylindan-4-yl)
malonamate in a manner similar to that described for 30 as a beige
solid (74%). Beige solid; mp 85–88 °C (dec) (EtOAc–hexane); 1H
NMR (400 MHz, CDCl3/CD3OD = 9/1) d: 1.95–2.10 (2H, m), 2.11
(3H, s), 2.55–2.65 (2H, m), 2.75–2.95 (6H, m), 3.45 (2H, s), 6.40–
6.50 (2H, m), 6.60–6.80 (3H, m), 7.00–7.10 (1H, m), 7.61 (1H, s);
13C NMR (100 MHz, CDCl3/CD3OD = 9/1) d: 15.9, 24.9, 28.2, 30.2,
30.9, 40.6, 103.3, 110.7, 113.4, 115.5, 117.0, 122.5, 124.5, 128.7,
129.4, 129.8, 131.3, 135.3, 137.5, 147.1, 149.0, 151.1, 160.2,
6.1.40. 2-Methoxy-5-(5-methyl-7-nitroindan-4-
yloxy)benzaldehyde (36)
To a solution of 10 (5.00 g, 16.7 mmol) and dichloromethyl
methyl ether (3.02 mL, 33.4 mmoL) in CH2Cl2 (50 mL), TiCl4
(42 mg, 0.198 mmol) was added dropwise at 0 °C. The mixture
was stirred overnight at room temperature. After adding ice water,
the reaction mixture was extracted with EtOAc. The organic layer
was washed with a saturated aqueous solution of NaHCO3 and
brine, and dried over anhydrous MgSO4. The solvent was removed
under reduced pressure. The residue was triturated with hexane
and diethyl ether, and the insoluble material was collected by fil-
tration to give 36 (4.37 g, 80%) as a beige solid. Beige solid; mp
175–176 °C (EtOAc–hexane); 1H NMR (400 MHz, CDCl3) d: 2.00–
2.15 (2H, m), 2.22 (3H, s), 2.64 (2H, t, J = 7.5 Hz), 3.39 (2H, t,
J = 7.5 Hz), 3.92 (3H, s), 6.97 (1H, d, J = 9.0 Hz), 7.11 (1H, dd,
J = 3.2, 9.0 Hz), 7.15 (1H, d, J = 3.2 Hz), 7.97 (1H, s), 10.41 (1H, s);
13C NMR (100 MHz, CDCl3) d: 16.0, 24.8, 30.3, 34.3, 56.2, 113.4,
113.8, 123.5, 125.4, 125.9, 130.7, 139.5, 142.0, 142.1, 150.8,
153.9, 157.5, 189.1; HRMS calcd for C18H18NO5 (M+H)+ 328.1179,
found 328.1187.
161.8, 164.6, 171.7; HRMS calcd for
482.1774, found 482.1779.
C
27H26F2NO5 (M+H)+
6.1.36. N-(7-{3-[2-(3,4-Difluorophenyl)ethyl]-4-
hydroxyphenoxy}-6-methylindan-4-yl)malonamic acid (35b)
The title compound was prepared from 34b in a manner similar
to that described for 35a as a beige solid (29%, 2 steps). Beige solid;
mp 153–154 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, DMSO-
d6) d: 1.85–2.00 (5H, m), 2.35–2.50 (2H, m), 2.70–2.95 (6H, m),
6.30–6.40 (2H, m), 6.65–6.70 (1H, m), 6.90–7.00 (1H, m), 7.10–
7.30 (2H, m), 7.79 (1H, s), 9.04 (1H, s), 12.45 (1H, br s); 13C NMR
(100 MHz, DMSO-d6) d: 15.6, 24.4, 29.9, 30.5, 31.4, 33.8, 43.2,
113.2, 115.5, 116.6, 116.7, 116.9, 117.0, 117.2, 123.2, 125.0,
128.0, 128.1, 130.3, 135.9, 136.6, 139.4, 146.2, 149.6, 149.9,
164.3, 169.6; HRMS calcd for C27H26F2NO5 (M+H)+ 482.1774, found
482.1780.
6.1.41. 7-{4-Methoxy-3-[2-(2-methoxyphenyl)ethyl]phenoxy}-
6-methylindan-4-ylamine (38a)
6.1.37. N-(7-{3-[2-(2-Chlorophenyl)ethyl]-4-hydroxyphenoxy}-
6-methylindan-4-yl)malonamic acid (35c)
To a solution of (2-methoxybenzyl)triphenylphosphonium bro-
mide (37a) (546 mg, 1.30 mmol) in THF (20 mL), KOtBu (135 mg,
1.20 mmol) was added under Ar at room temperature. After stir-
ring for 15 min, 36 (328 mg, 1.00 mmol) was added to give 4-{4-
methoxy-3-[2-(2-methoxyphenyl)vinyl]phenoxy}-5-methyl-7-
nitroindane (247 mg, 57%)
The title compound was prepared from 34c in a manner similar
to that described for 35a as a beige solid (30%, 2 steps). Beige solid;
mp 134–138 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, CDCl3/
CD3OD = 9/1) d: 1.95–2.10 (2H, m), 2.12 (3H, s), 2.55–2.65 (2H,
m), 2.80–2.90 (4H, m), 2.95–3.05 (2H, m), 3.50 (2H, s), 6.43 (1H,

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
605
To
a
solution of 4-{4-methoxy-3-[2-(2-methoxyphenyl)
95 °C (EtOAc–hexane); 1H NMR (400 MHz, CDCl3) d: 1.34 (3H, t,
J = 7.1 Hz), 1.95–2.10 (2H, m), 2.13 (3H, s), 2.55–2.70 (2H, m),
2.75–2.95 (6H, m), 3.49 (2H, s), 3.77 (3H, s), 3.78 (3H, s), 4.27
(2H, q, J = 7.1 Hz), 6.45–6.60 (2H, m), 6.65–6.80 (4H, m), 7.10–
7.25 (1H, m), 7.75 (1H, s), 9.18 (1H, s); 13C NMR (100 MHz, CDCl3)
d: 14.1, 16.1, 24.9, 30.2, 30.3, 32.4, 36.0, 41.2, 55.1, 55.8, 61.9,
111.1, 111.2, 112.7, 114.1, 117.1, 121.0, 121.8, 129.1, 130.0,
130.0, 131.4, 134.3, 137.5, 143.8, 146.6, 151.6, 152.2, 159.5,
162.6, 170.4; HRMS calcd for C31H36NO6 (M+H)+ 518.2537, found
518.2533.
vinyl]phenoxy}-5-methyl-7-nitroindane (257 mg, 0.572 mmol) in
EtOAc (50 mL), 10% Pd/C (56% wet with water, 200 mg,
0.0827 mmol) was added. The mixture was stirred overnight under
a hydrogen atmosphere at room temperature. Insoluble materials
were removed by filtration and washed with EtOAc. The filtrate
was concentrated in vacuo. The residue was purified by column
chromatography on silica gel (eluent: hexane/EtOAc = 19/1–0/1)
to give 38a (192 mg, 83%) as a colorless amorphous solid. Colorless
amorphous solid; 1H NMR (400 MHz, CDCl3) d: 1.90–2.15 (5H, m),
2.55–2.92 (8H, m), 3.45 (2H, br s), 3.75 (3H, s), 3.80 (3H, s), 6.40
(1H, d, J = 4.5 Hz), 6.48 (1H, dd, J = 8.8 Hz, 3.0 Hz), 6.62 (1H, d,
J = 3.0 Hz), 6.67 (1H, d, J = 8.8 Hz), 6.75–6.95 (2H, m), 7.00–7.25
(2H, m); 13C NMR (100 MHz, CDCl3) d: 15.9, 24.9, 29.7, 30.3, 30.4,
30.4, 55.3, 55.9, 110.2, 111.0, 112.0, 115.5, 117.0, 120.3, 126.9,
128.3, 129.9, 130.0, 130.6, 132.1, 137.8, 138.8, 142.3, 152.0,
152.2, 157.5; HRMS calcd for C26H30NO3 (M+H)+ 404.2220, found
404.2219.
6.1.46. Ethyl N-(7-{4-methoxy-3-[2-(4-methoxyphenyl)
ethyl]phenoxy}-6-methylindan-4-yl)malonamate (39c)
The title compound was prepared from 38c in a manner similar
to that described for 12 as a colorless amorphous solid (77%). Col-
orless amorphous solid; 1H NMR (400 MHz, CDCl3) d: 1.34 (3H, t,
J = 7.1 Hz), 1.95–2.10 (2H, m), 2.12 (3H, s), 2.55–2.70 (2H, m),
2.75–2.95 (6H, m), 3.50 (2H, s), 3.77 (3H, s), 3.78 (3H, s), 4.27
(2H, q, J = 7.1 Hz), 6.50–6.60 (2H, m), 6.65–6.85 (3H, m), 7.00–
7.10 (2H, m), 7.74 (1H, s), 9.20 (1H, s); 13C NMR (100 MHz, CDCl3)
d: 14.1, 16.1, 24.9, 30.2, 30.4, 32.7, 35.0, 41.2, 55.2, 55.9, 61.9,
111.1, 112.7, 113.6, 117.1, 121.8, 129.4, 130.0, 130.0, 131.5,
134.3, 134.3, 137.5, 146.6, 151.6, 152.2, 157.7, 162.6, 170.4; HRMS
calcd for C31H36NO6 (M+H)+ 518.2537, found 518.2535.
6.1.42. 7-{4-Methoxy-3-[2-(3-methoxyphenyl)ethyl]phenoxy}-
6-methylindan-4-ylamine (38b)
The title compound was prepared from (3-methoxybenzyl)tri-
phenylphosphonium bromide (37b) in a manner similar to that de-
scribed for 38a as a colorless amorphous solid (70%, 2 steps).
Colorless amorphous solid; 1H NMR (400 MHz, CDCl3) d: 1.90–
2.15 (5H, m), 2.55–2.95 (8H, m), 3.45 (2H, br s), 3.70–3.85 (6H,
m), 6.35–6.90 (7H, m), 7.10–7.25 (1H, m); 13C NMR (100 MHz,
CDCl3) d: 15.8, 24.9, 29.7, 30.3, 32.4, 36.1, 55.1, 55.9, 111.1,
111.2, 112.4, 114.1, 115.5, 116.9, 121.0, 128.4, 129.1, 129.9,
131.3, 137.7, 138.8, 142.3, 143.9, 151.9, 152.2, 159.5; HRMS calcd
for C26H30NO3 (M+H)+ 404.2220, found 404.2215.
6.1.47. N-(7-{4-Hydroxy-3-[2-(2-hydroxyphenyl)
ethyl]phenoxy}-6-methylindan-4-yl}malonamic acid (40a)
The title compound was prepared from 39a in a manner similar
to that described for 35a as a beige solid (56%, 2 steps). Beige solid;
mp 102–106 °C (dec) (EtOAc–hexane); 1H NMR (400 MHz, CDCl3/
CD3OD = 7/1) d: 1.95–2.10 (2H, m), 2.13 (3H, s), 2.60–2.95 (8H,
m), 3.49 (2H, s), 6.50–6.60 (2H, m), 6.65–6.80 (3H, m), 7.05–7.15
(2H, m), 7.59 (1H, s); 13C NMR (100 MHz, CDCl3/CD3OD = 7/1) d:
16.1, 25.2, 30.5, 30.6, 30.9, 31.3, 41.0, 113.6, 115.3, 115.9, 116.7,
120.1, 123.2, 127.5, 128.2, 129.6, 130.0, 130.1, 130.2, 136.2,
137.9, 147.5, 149.2, 151.4, 154.8, 165.2, 171.6; HRMS calcd for
6.1.43. 7-{4-Methoxy-3-[2-(4-methoxyphenyl)ethyl]phenoxy}-
6-methylindan-4-ylamine (38c)
The title compound was prepared from (4-methoxybenzyl)tri-
phenylphosphonium bromide (37c) in a manner similar to that de-
scribed for 38a as a colorless amorphous solid (49%, 2 steps).
Colorless amorphous solid; 1H NMR (400 MHz, CDCl3) d: 1.95–
2.15 (5H, m), 2.55–2.95 (8H, m), 3.45 (2H, br s), 3.76 (3H, s), 3.78
(3H, s), 6.40 (1H, s), 6.45–6.55 (2H, m), 6.69 (1H, d, J = 8.7 Hz),
6.75–6.85 (2H, m), 7.00–7.10 (2H, m); 13C NMR (100 MHz, CDCl3)
d: 15.8, 24.9, 29.6, 30.3, 32.7, 35.1, 55.2, 55.9, 111.1, 112.3, 113.6,
115.5, 116.9, 128.4, 129.4, 129.9, 131.4, 134.4, 137.7, 138.8,
142.3, 151.9, 152.2, 157.7; HRMS calcd for C26H30NO3 (M+H)+
404.2220, found 404.2218.
C
27H28NO6 (M+H)+ 462.1911, found 462.1913.
6.1.48. N-(7-{4-Hydroxy-3-[2-(3-hydroxyphenyl)
ethyl]phenoxy}-6-methylindan-4-yl}malonamic acid (40b)
The title compound was prepared from 39b in a manner similar
to that described for 35a as a white solid (65%, 2 steps). White so-
lid; mp 150–152 °C (dec) (EtOAc–hexane); 1H NMR (600 MHz,
DMSO-d6) d: 1.85–1.95 (2H, m), 2.02 (3H, s), 2.45–2.50 (2H, m),
2.65–2.80 (6H, m), 3.39 (2H, s), 6.36 (1H, dd, J = 3.0, 8.6 Hz), 6.47
(1H, d, J = 3.0 Hz), 6.50–6.60 (3H, m), 6.67 (1H, d, J = 8.6 Hz),
6.95–7.05 (1H, m), 7.36 (1H, s), 8.97 (1H, s), 9.16 (1H, s), 12.60
(1H, br s); 13C NMR (150 MHz, DMSO-d6) d: 15.7, 24.5, 29.9, 30.5,
31.7, 35.1, 43.2, 112.6, 112.8, 115.2, 115.4, 116.6, 119.0, 123.2,
128.2, 128.8, 129.0, 130.3, 136.0, 136.7, 143.1, 146.2, 149.6,
149.9, 157.2, 164.3, 169.6; HRMS calcd for C27H28NO6 (M+H)+
462.1911, found 462.1923.
6.1.44. Ethyl N-(7-{4-methoxy-3-[2-(2-methoxyphenyl)
ethyl]phenoxy}-6-methylindan-4-yl)malonamate (39a)
The title compound was prepared from 38a in a manner similar
to that described for 12 as a white solid (80%). White solid; mp
101–102 °C (EtOAc–hexane); 1H NMR (400 MHz, CDCl3) d: 1.34
(3H, t, J = 7.1 Hz), 1.95–2.10 (2H, m), 2.13 (3H, s), 2.63 (2H, t,
J = 7.4 Hz), 2.75–2.95 (6H, m), 3.49 (2H, s), 3.76 (3H, s), 3.79 (3H,
s), 4.27 (2H, q, J = 7.1 Hz), 6.49 (1H, dd, J = 3.0, 8.8 Hz), 6.60 (1H,
d, J = 3.0 Hz), 6.68 (1H, d, J = 8.8 Hz), 6.75–6.90 (2H, m), 7.00–7.20
(2H, m), 7.75 (1H, s), 9.18 (1H, s); 13C NMR (100 MHz, CDCl3) d:
14.1, 16.1, 24.9, 30.2, 30.3, 30.4, 30.4, 41.2, 55.3, 55.9, 61.9, 110.2,
111.0, 112.3, 117.3, 120.2, 121.8, 127.0, 129.9, 130.0, 130.0,
130.5, 132.2, 134.2, 137.5, 146.7, 151.6, 152.3, 157.5, 162.6,
6.1.49. N-(7-{4-Hydroxy-3-[2-(4-hydroxyphenyl)
ethyl]phenoxy}-6-methylindan-4-yl}malonamic acid (40c)
The title compound was prepared from 39c in a manner similar
to that described for 35a as a colorless amorphous solid (56%, 2
steps). Colorless amorphous solid; 1H NMR (400 MHz, CDCl3/
CD3OD = 7/1) d: 1.95–2.10 (5H, m), 2.55–2.65 (2H, m), 2.75–2.90
(6H, m), 3.48 (2H, s), 6.34 (1H, d, J = 3.0 Hz), 6.49 (1H, dd, J = 3.0,
8.9 Hz), 6.60–6.70 (3H, m), 6.90–7.00 (2H, m), 7.55 (1H, s); 13C
NMR (100 MHz, CDCl3/CD3OD = 7/1) d: 15.9, 24.9, 29.6, 30.3, 32.2,
34.6, 40.3, 113.5, 115.0, 115.6, 116.8, 122.6, 122.7, 129.2, 129.4,
129.8, 133.0, 135.3, 137.6, 147.2, 148.8, 151.0, 154.4, 164.5,
170.4; HRMS calcd for
518.2532.
C
31H36NO6 (M+H)+ 518.2537, found
6.1.45. Ethyl N-(7-{4-methoxy-3-[2-(3-methoxyphenyl)
ethyl]phenoxy}-6-methylindan-4-yl)malonamate (39b)
The title compound was prepared from 38b in a manner similar
to that described for 12 as a white solid (78%). White solid; mp 94–
171.5; HRMS calcd for
462.1943.
C
27H28NO6 (M+H)+ 462.1911, found

606
H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
6.2. Biology
Optimization was performed stepwise. Initially, structures were
minimized under conditions that constrained non-hydrogen
atoms. Next, the protein backbone atoms were constrained. At
the final step, all atoms were minimized with a harmonic atom
constraint. The force constants of the harmonic atom constraints
gradually decreased from 10.0 to 1.0 kcal/mol Å2.
6.2.1. Receptor binding assay
Recombinant hTR
Cell homogenates containing the respective receptors were mixed
with appropriate concentrations of test compounds.
-3,5,30-[125I]-
triiodothyronine ([125I]-T3, 0.95 nM, 160 Ci/mmol, [125I]-T3(NEN)
was diluted with
-3,5,30-triiodothyronine (Sigma) in a buffer con-
a
1 and hTRb1 were expressed in insect cells.32
L
CH/
p interactions were evaluated using the program CHPI
L
implemented in BioStation Viewer.26 This program determined
the distances and angles between CHs and interacting aromatic
taining 0.4 M KC1, 1 mM MgCl2, 10 mM Tris–HCl, and 1 mM dithi-
othreitol (pH 8.0). A 0.5-mL aliquot of the mixture was incubated
in a glass tube in an ice bath for 16–48 h. After incubation,
rings. This program searched for CH/
p interactions based on the
distances (<3.05 Å) and angles between CHs and interacting aro-
500 lL of an ion-exchange resin (Muromachi Kagaku, Dowex
matic rings.38
1-X8, 80 mg/mL, suspended in the buffer indicated above) was
added to each test tube and stirred. Stirring was repeated after
the resin sedimented at the bottom of the tube, followed by addi-
tional stirring. The tubes were centrifuged at 1000 rpm for 5 min at
1 °C using a centrifuge separator (KUBOTA, 8800). An aliquot of the
Acknowledgments
We thank Professor Dr. Atsushi Kittaka for helpful comments
while reviewing the manuscript, and Dr. Yoshinobu Nonaka and
Dr. Hideyuki Muranaka of Kissei Pharmaceutical Co., Ltd for NMR
and HRMS measurements of the compounds, respectively.
supernatant (500
lL) was transferred to another tube, and radioac-
tivity was measured with a
c
-ray detector. The detected radioac-
tivity was proportional to the amount of [125I]-T3 bound to the
soluble thyroid hormone receptor. The amount of recombinant
thyroid hormone receptor cell homogenate used in the experiment
was in a range for which the radioactivity associated with T3 bind-
ing showed a concentration-dependent increase in the amount of
the homogenate.
References and notes
1. Pramfalk, C.; Pedrelli, M.; Parini, P. Biochim. Biophys. Acta 1812, 2011, 929.
2. Moreno, M.; de Lange, P.; Lombardi, A.; Silvestri, E.; Lanni, A.; Goglia, F. Thyroid
2008, 18, 239.
3. Oetting, A.; Yen, P. M. Best Pract. Res. 2007, 21, 193.
The Kd value of T3 to a particular receptor subtype was deter-
mined from Scatchard analysis of the binding data obtained with
the different [125I]-T3 levels. Under these experimental conditions,
the Kd values of T3 binding for hTRa1 and hTRb1 were 0.268 and
0.304 nM, respectively.
4. Scanlan, T. S. Heart Fail. Rev. 2010, 15, 177.
5. Privalsky, M. L.; Lee, S.; Hahm, J. B.; Young, B. M.; Fong, R. N. G.; Chan, I. H. J.
Biol. Chem. 2009, 284, 19554.
6. Malm, J.; Grover, G. J.; Färnegårdh, M. Mini Rev. Med. Chem. 2007, 7, 79.
7. Grommen, S. V.; Arckens, L.; Theuwissen, T.; Darras, V. M.; De Groef, B. J.
Endocrinol. 2008, 196, 519.
8. Pedrelli, M.; Pramfalk, C.; Parini, P. World J. Gastroenterol. 2010, 16, 5958.
9. Erion, M. D.; Cable, E. E.; Ito, B. R.; Jiang, H.; Fujitaki, J. M.; Finn, P. D.; Zhang, B.
H.; Hou, J.; Boyer, S. H.; van Poelje, P. D.; Linemeyer, D. L. Proc. Natl. Acad. Sci.
U.S.A. 2007, 104, 15490.
10. Tancevski, I.; Wehinger, A.; Demetz, E.; Hoefer, J.; Eller, P.; Huber, E.; Stanzl, U.;
Duwensee, K.; Auer, K.; Schgoer, W.; Kuhn, V.; Fievet, C.; Stellaard, F.; Rudling,
M.; Foeger, B.; Patsch, J. R.; Ritsch, A. J. Lipid Res. 2009, 50, 938.
11. Tancevski, I.; Demetz, E.; Eller, P.; Duwensee, K.; Hoefer, J.; Heim, C.; Stanzl, U.;
Wehinger, A.; Auer, K.; Karer, R.; Huber, J.; Schgoer, W.; Van Eck, M.;
Vanhoutte, J.; Fievet, C.; Stellaard, F.; Rudling, M.; Patsch, J. R.; Ritsch, A. PLoS
ONE 2010, 5, e8722.
The Ki values of each compound were calculated using the
equation
Ki ðnMÞ ¼ ½IC50ꢁ=ð1 þ Kd=0:95Þ;
where IC50 indicated the concentration of the compound that inhib-
ited [125I]-T3 binding by 50%.
6.2.2. Luciferase reporter gene assay
hTRa133 and hTRb134 were cloned in a mammalian cell expression
vector (pCDM8, Promega) as described previously.35 The luciferase
reporter gene was constructed by inserting the thyroid hormone
responsive element (50-GATCCAGGTCATGACCTGGATCC-30) into a
commercial luciferase expression vector (pGL2-promoter, Prome-
ga). Each thyroid hormone receptor vector and the luciferase repor-
ter vector were co-transfected into trypsin-digested and suspended
COS1 cells using the calcium phosphate-mediated method.36 These
cells were seeded into 96-well multiwell plates and cultured over-
12. Joharapurkar, A. A.; Dhote, V. V.; Jain, M. R. J. Med. Chem. 2012, 55, 5649.
13. Joy, T. R. Pharmacol. Ther. 2012, 135, 31.
14. Kowalik, M. A.; Perra, A.; Pibiri, M.; Cocco, M. T.; Samarut, J.; Plateroti, M.;
Ledda-Columbano, G. M.; Columbano, A. J. Hepatol. 2010, 53, 686.
15. Li, J. J.; Mitchell, L. H.; Dow, R. L. Bioorg. Med. Chem. Lett. 2010, 20, 306.
16. Shiohara, H.; Nakamura, T.; Kikuchi, N.; Ozawa, T.; Nagano, R.; Matsuzawa, A.;
Ohnota, H.; Miyamoto, T.; Ichikawa, K.; Hashizume, K. Bioorg. Med. Chem. 2012,
20, 3622.
17. Dow, R. L.; Schneider, S. R.; Paight, E. S.; Hank, R. F.; Chiang, P.; Cornelius, P.;
Lee, E.; Newsome, W. P.; Swick, A. G.; Spitzer, J.; Hargrove, D. M.; Patterson, T.
A.; Pandit, J.; Chrunyk, B. A.; LeMotte, P. K.; Danley, D. E.; Rosner, M. H.;
Ammirati, M. J.; Simons, S. P.; Schulte, G. K.; Tate, B. F.; DaSilva-Jardine, P.
Bioorg. Med. Chem. Lett. 2003, 13, 379.
18. Yokoyama, N.; Walker, G. N.; Main, A. J.; Stanton, J. L.; Morrissey, M. M.;
Boehm, C.; Engle, A.; Neubert, A. D.; Wasvary, J. M.; Stephan, A. F.; Steele, R. E. J.
Med. Chem. 1995, 38, 695.
19. Leeson, P. D.; Emmett, J. C.; Shah, V. P.; Showell, G. A.; Novelli, R.; Prain, H. D.;
Benson, M. G.; Ellis, D.; Pearce, N. J.; Underwood, A. H. J. Med. Chem. 1989, 32,
320.
night. The following day, thyromimetics (at 10ꢀ9, 10ꢀ8, 10ꢀ7, 10ꢀ6
,
10ꢀ5, 10ꢀ4 M) were added and culture continued for 1 day. Lucifer-
ase activities were determined using a commercial kit (Luciferase
Assay System, Promega) and Top count (Packard) on the third day.
All thyromimetics showed dose-dependent responses.
20. Borngraeber, S.; Budny, M. J.; Chiellini, G.; Cunha-Lima, S. T.; Togashi, M.;
Webb, P.; Baxter, J. D.; Scanlan, T. S.; Fletterick, R. J. Proc. Natl. Acad. Sci. U.S.A.
2003, 100, 15358.
21. Similar reactions have been reported Khodaei, M. M.; Alizadeh, A.; Nazari, E.
Tetrahedron Lett. 2007, 48, 4199.
22. Hickey, D. M. B.; Leeson, P. D.; Novelli, R.; Shah, V. P.; Burpitt, B. E.; Crawford, L.
P.; Davies, B. J.; Mitchell, M. B.; Pancholi, K. D.; Tuddenham, D.; Lewis, N. J.;
O’Farrell, C. J. Chem. Soc., Perkin Trans. 1 1988, 3103.
23. Martínez, L.; Nascimento, A. S.; Nunes, F. M.; Phillips, K.; Aparicio, R.; Dias, S.
M.; Figueira, A. C.; Lin, J. H.; Nguyen, P.; Apriletti, J. W.; Neves, F. A.; Baxter, J.
D.; Webb, P.; Skaf, M. S.; Polikarpov, I. Proc. Natl. Acad. Sci. U.S.A. 2009, 106,
20717.
6.3. Molecular modeling
A docking model for 40a was constructed based on the crystal
structure of hTRb1 with 41 (PDB code 3JZC23). The malonic acid
part of 40a was modeled by keeping the hydrogen bonds that were
observed between TRb and 41. The phenethyl part of 40a was mod-
eled with reference to the complex between TRb and 7, which had
a large substituent next to the hydroxyl group of di-phenyl ether
(PDB code 1Q4X20).
24. Takahashi, O.; Kohono, Y.; Nishio, M. Chem. Rev. 2010, 110, 6049.
25. Ozawa, T.; Tsuji, E.; Ozawa, M.; Handa, C.; Mukaiyama, H.; Nishimura, T.;
Kobayashi, S.; Okazaki, K. Bioorg. Med. Chem. 2008, 16, 10311.
26. http://www.ciss.iis.u-tokyo.ac.jp/rss21/theme/life/synergy/synergy_softwareinfo.
html.
The constructed model was optimized using CHARMm force-
field implemented in Discovery Studio3.1 (Accelrys Inc., San Diego,
CA, USA).37 Protein structures were optimized by the steepest des-
cent (SD) method at a dielectric constant of
e = 4R (R: distance).

H. Shiohara et al. / Bioorg. Med. Chem. 21 (2013) 592–607
607
27. Martínez, L.; Sonoda, M. T.; Webb, P.; Baxter, J. D.; Skaf, M. S.; Polikarpov, I.
Biophys. J. 2005, 89, 2011.
34. Weinberger, C.; Thompson, C. C.; Ong, E. S.; Lebo, R.; Gruol, D. J.; Evans, R. M.
Nature 1986, 324, 641.
28. Martínez, L.; Webb, P.; Polikarpov, I.; Skaf, M. S. J. Med. Chem. 2006, 49, 23.
29. Martínez, L.; Polikarpov, I.; Skaf, M. S. J. Phys. Chem. B. 2008, 112, 10741.
30. DeAraujo, A. S.; Martínez, L.; DePaulaNicoluci, R.; Skaf, M. S.; Polikarpov, I. Eur.
Biophys. J. 2010, 39, 1523.
35. Nakai, A.; Sakurai, A.; Macchia, E.; Fang, V.; DeGroot, L. J. Mol. Cell. Endocrinol.
1990, 72, 143.
36. Sambrook, J.; Fritsch, E. F.; Maniatis, T. Molecular Cloning: A Laboratory Manual,
2nd ed.; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, New York,
1989. Chapter 16.33.
31. Mariash, C. N. Thyroid 2010, 20, 451.
32. Miyamoto, T.; Kaneko, A.; Kakizawa, T.; Yajima, H.; Kamijo, K.; Sekine, R.; Hiramatsu,
K.; Nishii, Y.; Hashimoto, T.; Hashizume, K. J. Biol. Chem. 1997, 272, 7752.
33. Nakai, A.; Sakurai, A.; Bell, G. I.; DeGroot, L. J. Mol. Endocrinol. 1988, 2, 1087.
37. http://accelrys.com/products/discovery-studio/.
38. Umezawa, Y.; Tsuboyama, S.; Honda, K.; Uzawa, J.; Nishio, M. Bull. Chem. Soc.
Jpn. 1998, 71, 1207.