Estrogenic activity of bis(4-hydroxyphenyl)methanes with cyclic hydrophobic structure

Article abstract of DOI:10.1016/j.bmc.2015.09.046

Monoalkylated bis(4-hydroxyphenyl)methanes (e.g., 1) are reported to show weak binding affinity for estrogen receptor (ER). We hypothesized that introduction of appropriately located hydrophobic substituents in these compounds would increase the binding affinity. Indeed, we found that bis(4-hydroxyphenyl)methane bearing a 3,3-dimethylcyclohexyl group (7) shows potent ERα binding affinity, comparable to that of estradiol. Bulkier substituents could be introduced at the 3,3-position without decreasing the affinity. However, the position of the substituents was critical: the 4,4-dimethylcyclohexyl derivative (2) showed very weak binding affinity. The compounds with high ER-binding affinity showed predominantly agonistic activity, together with weak antagonistic activity at high concentration, in cell proliferation assay with human breast cancer cell line MCF-7. Further structure-function studies of these compounds and their derivatives might lead to the development of more selective and potent estrogen receptor modulators.

Full text of DOI:10.1016/j.bmc.2015.09.046

Bioorganic & Medicinal Chemistry 23 (2015) 6900–6911
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Estrogenic activity of bis(4-hydroxyphenyl)methanes with cyclic
hydrophobic structure
Tomohiro Kojima a, Takumi Ogawa a, Souichiro Kitao a, Manabu Sato a, Akifumi Oda b,c, Kiminori Ohta a,
Yasuyuki Endo a,
⇑
a Faculty of Pharmaceutical Sciences, Tohoku Pharmaceutical University, 4-4-1 Komatsushima, Aoba-ku, Sendai 981-8558, Japan
b Faculty of Pharmacy, Institute of Medical, Pharmaceutical and Health Sciences, Kanazawa University, Kakuma-machi, Kanazawa, Ishikawa 920-1192, Japan
c Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Monoalkylated bis(4-hydroxyphenyl)methanes (e.g., 1) are reported to show weak binding affinity for
estrogen receptor (ER). We hypothesized that introduction of appropriately located hydrophobic sub-
stituents in these compounds would increase the binding affinity. Indeed, we found that bis(4-hydrox-
Received 14 September 2015
Revised 26 September 2015
Accepted 29 September 2015
Available online 1 October 2015
yphenyl)methane bearing a 3,3-dimethylcyclohexyl group (7) shows potent ER
a binding affinity,
comparable to that of estradiol. Bulkier substituents could be introduced at the 3,3-position without
decreasing the affinity. However, the position of the substituents was critical: the 4,4-dimethylcyclohexyl
derivative (2) showed very weak binding affinity. The compounds with high ER-binding affinity showed
predominantly agonistic activity, together with weak antagonistic activity at high concentration, in cell
proliferation assay with human breast cancer cell line MCF-7. Further structure–function studies of these
compounds and their derivatives might lead to the development of more selective and potent estrogen
receptor modulators.
Keywords:
Estrogen receptor
Partial agonist
SERM
Carborane
Ó 2015 Elsevier Ltd. All rights reserved.
1. Introduction
selectivity is considered to be quantitative and qualitative differ-
ences of co-regulatory proteins in the ER-mediated transcriptional
Estrogens, such as 17b-estradiol, play an important role in the
female and male reproductive systems,1 as well as in bone mainte-
nance, in the central nervous system2 and in the cardiovascular
system.3 The first step in the appearance of these activities is bind-
systems of each target tissue.9 These co-regulatory proteins alter
the conformational state of the ER–ligand complex to influence
the transcriptional action. Similarly, the agonist/antagonist balance
of SERMs is determined by the conformational state of the ER–
ligand complex. These complex macromolecular systems can be
controlled by low-molecular-weight ligands. Typical SERMs such
as tamoxifen10 and raloxifene11 were found to be agonistic in bone
and antagonistic in breast, but showed varying activity in uterus.
The agonist/antagonist balances of the two SERMs are different:
tamoxifen is more antagonistic and raloxifene is more agonistic.
Therefore, elucidation of the structure–activity relationships of
partial agonist/antagonists is important for elucidating ER activa-
tion mechanisms and for developing useful clinical medicines.
Based on the structure of the complex formed by estradiol and
ing of the ligands to estrogen receptors
a and b (ERa
4 and b5). This
binding results in a conformational change of the receptor, induc-
ing dimerization. The dimer functions as a transcription factor,
which causes biological responses by binding to specific promoter
elements of DNA to initiate gene transcription. It has been reported
that ERa
and ERb have different tissue distributions6 and biological
roles, and they sometimes act in opposition to each other.
Numerous subtype-selective ligands have been reported and
ERb-selective ligands are of potential clinical interest.7 However,
the nature of the differences between the two ER subtypes has
not been fully established, possibly because ER
a
is predominant
the human ER-
a
LBD (hER
a
LBD),12 the phenolic hydroxyl group is
LBD and the 17b-
as a transcription factor, compared with ERb.8
hydrogen-bonded to Glu-353 and Arg394 of hER
a
From a clinical point of view, there is great interest in selective
estrogen receptor modulators (SERMs), which are tissue-selective
ER agonists and antagonists. The major factor determining tissue
hydroxyl group is hydrogen-bonded to the d-nitrogen of His-524.
The importance of these two hydrogen-bondings is very different:
the phenolic hydrogen-bonding is indispensable, whereas the 17-
hydroxyl hydrogen-bonding can be replaced by hydrophobic inter-
action. Hydrophobic interaction of the steroidal skeleton of the
ligand with the LBD also plays an important role for stabilization
⇑
Corresponding author.
E-mail address: yendo@tohoku-pharm.ac.jp (Y. Endo).
http://dx.doi.org/10.1016/j.bmc.2015.09.046
0968-0896/Ó 2015 Elsevier Ltd. All rights reserved.

T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
6901
Figure 1. Various estrogen receptor modulators.
and binding activity of the ER–ligand complex. Since the discovery
of the potent ER agonist diethylstilbestrol and SERM tamoxifen,
triphenylethylene and diphenylethylene skeletons have attracted
considerable attention as basic structures for ER ligands.13
Katzenellenbogen et al. have reported that 1,1-diarylethylene
derivatives with bridged bicyclic structures have high ER-binding
activity, and these compounds mainly show ER antagonistic activ-
ity.14 Jordan et al. reported that 1,1,2-triphenylethylenes with two
or three hydroxyl groups in various phenyl rings showed ER ago-
nistic activity.15 Another important compound is bisphenol A,
which has weak estrogenic activity but has been manufactured
on a considerable scale as an industrial material for use in plastic
products. Recently, Hashimoto et al. reported that bisphenol A
We have focused on the application of three-dimensional
hydrophobic structural units, such as icosahedral boron clusters
(carboranes17), that fit in the hydrophobic cavity of the ER LBDs.18
In our design and biological activity studies of bisphenols with a
hydrophobic core structure, we have found that the hydrophobic
core structure has the ability to regulate agonist/antagonist bal-
ance. For example, BE360, bis(4-hydroxylphenyl)-o-carborane,
showed partial antagonist activity towards ER.19 However,
BE1060, in which the carborane cage of BE360 is replaced with a
hydrocarbon core, bicyclo[2,2,2]octene, exhibited potent ER ago-
nistic activity, even though the two phenolic groups appear to be
similarly directed.20 Thus, differences in receptor–ligand complex
structures arising from the presence of different hydrophobic
structures in the ligand can influence biological activity. 1,1-Diary-
lethylene derivatives such as cyclofenil have potent binding affin-
ity for ER, and show mixed agonist/antagonist activities.21 On the
other hand, hydrogenated 1,1-diarylmethane derivatives22 have
not received much attention because of their lower binding affin-
ity.14 However, we hypothesized that addition of appropriate
hydrophobic substituents would improve the binding affinity and
derivatives with extended alkyl chains show selective ER
a antago-
nistic activity16 (see Fig. 1).
provide
a means to regulate the agonist/antagonist activity
balance. In this article, we describe the synthesis and biological
evaluation of simple bisphenols, bis(4-hydroxyphenyl)methanes,
bearing cyclic hydrophobic structures.
2. Results
2.1. Chemistry
We presumed that the first phenolic hydroxyl group would act
as an anchor at the hydrogen-bonding site of ER, while the three-
dimensional hydrophobic core would fill the hydrophobic cavity
of ER. The steric and electronic effects of substituents on the
hydrophobic core were expected to determine the nature of the
estrogenic action. Therefore, we designed bis(4-hydroxyphenyl)
methanes with a cyclohexyl group bearing various substituents
(1–12), as shown in Figure 2. We also designed bis(4-hydrox-
yphenyl)methanes with a cyclopentyl group (13) and a carboranyl
group (14–16).
Synthesis of unsubstituted and 4,4-disubstituted cyclohexyl
derivatives is summarized in Scheme 1. Cyclohexyl (1) and 4,4-
dimethylcyclohexyl (2) derivatives were prepared by McMurry
Figure 2. ER Ligands with cyclic hydrophobic structures. In the icosahedral cage
structure, d represent carbon atoms and other vertices represent BH units.
coupling
reaction
of
4,4-dimethoxybenzophenone
with

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T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
Scheme 1. Synthesis of unsubstituted and 4,4-disubstituted derivatives. Reagents and conditions: (a) 4,40-dimethyoxybenzophenone, TiCl4, Zn/THF, reflux, 74–85%; (b) H2,
Pd–C/CH3COOC2H5, rt, 93–98%; (c) BBr3/CH2C12, ꢀ40 °C, 61–90%; (d) SiO2, H2SO4/CH2C12, rt, 80%; (e) (1) DAST/CH2C12, rt, (2) OsO4, NMO/acetone–H2O, rt, 50%; (f) benzyl
bromide, K2CO3/acetone, reflux, 86%; (g) (1) N2H4, MS4A/CH3OH–CH2C12, rt, (2) CuCl2, Et3N/CH3OH–CH2C12, rt, 32%; (h) BC13/CH2C12, rt, 78%; (i) 2,3-dihydroxynaphthalene,
BF3OEt2, TMSC1/CH2C12, rt, 75%; (j) 4,40-dihydroxybenzophenone, TiCl4, Zn/THF, reflux, 69%.
cyclohexanone or 4,4-dimethylcyclohexanone to give 17 or 18.
Catalytic hydrogenation followed by deprotection afforded the
5-tetramethylcyclohexanone to give 22 or 23. Palladium-catalyzed
hydrogenation of 22 or 23 did not proceed smoothly, probably for a
steric hindrance, in contrast to the case of the 4,4-dimethyl deriva-
tives. Reduction of 22 or 23 with hydrogen iodide afforded 24 or 25
in low yield. Deprotection of the methyl group using BBr3 gave the
desired products 7 and 12. 3,3-Dihalogenated cyclohexyl deriva-
tives were prepared through the ketone 26. McMurry coupling
reaction of unprotected 4,40-dihydroxybenzophenone with 1,3-
cyclohexanedione mono-ethylene ketal followed by acidic treat-
ment afforded the conjugated ketone in 33% yield. The conjugated
ketone was readily hydrogenated to give ketone 26. Fluorination of
26 with DAST afforded 3,3-difluorocylclohexyl derivative 8 in 46%
yield. Chlorination of ketone 26 by a multi-step procedure in a
similar manner to the preparation of 4 afforded 3,3-dichlorocylclo-
hexyl derivative 9. The ketone 26 was also converted to 3,3-
dithoketal 10 by reaction with 1,3-propanedithiol in 78% yield.
3,3-Pentamethylene derivative (11) was prepared from spiro[5,5]
undecan-3-one (27). Compound 27 was converted to carboxyalde-
hyde 28 and then Friedel–Crafts reaction with phenol gave the
desired compound 11.
desired compounds
1 and 2, respectively. 4,4-Dihalogenated
derivatives were prepared though the ketone 21 as an intermedi-
ate. McMurry coupling reaction of 4,40-dimethoxybenzophenone
with 1,4-cyclohexanedione mono-ethylene ketal, followed by
acidic treatment and hydrogenation, gave ketone 21 in 53% yield.
Deprotection followed by fluorination with (diethylamino)sulfur
trifluoride (DAST) and treatment with OsO4 for separation of olefin
by-product afforded 4,4-difluorocyclohexyl derivative 3 in 50%
yield. After conversion of the protected group of the ketone 21 into
a benzyl group, reaction with hydrazine, followed by chlorination
with copper(II) chloride and deprotection afforded 4,4-dichlorocy-
clohexyl derivative 4. The carbonyl group of 21 was protected with
2,3-dihydroxynaphthalene ketal and treated with boron tribro-
mide to afford 4,4-dibromocyclohexyl derivative 5. The 4,4-ethy-
lene ketal derivative (6) was prepared from 1,4-cyclohexanedione
mono-ethylene ketal and 4,40-dihydroxybenzophenone by means
of the McMurry coupling reaction and catalytic hydrogenation.
Synthesis of 3,3-disubstituted and 3,3,5,5-tetrasubstituted
cyclohexyl derivatives is summarized in Scheme 2. 3,3-Dimethyl-
cyclohexyl (7) and 3,3,5,5-tetramethylcyclohexyl (12) derivatives
were prepared by McMurry coupling reaction of 4,4-dimethoxy-
benzophenone with 3,3-dimethylcyclohexanone or 3,3,5,
Synthesis of the cyclopentyl derivative (13) was carried out
according to the literature.23 Friedel–Crafts reaction of cyclohexene
oxide with anisole catalyzed by AlCl3 afforded 29 via rearrange-
ment of the carbon skeleton. Deprotection of the methyl group

T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
6903
Scheme 2. Synthesis of 3,3-disubstituted and 3,3,5,5-tetrasubstituted derivatives. Reagents and conditions: (a) 4,40-dimethoxybenzophenone, TiCl4, Zn/THF, reflux, 83–90%;
(b) HI/CH2C12 reflux, 11%; (c) BBr3/CH2C12, rt, 96–98%; (d) 4,40-dihydroxybenzophenone, TiCl4, Zn/THF, reflux, 45%; (e) HCl/acetone rt, 74%; (f) H2, Pd–C/C2H5OH, 88%; (g)
DAST/CH2C12, rt, 46%; (h) benzyl bromide, K2CO3/acetone, reflux, 70%; (i) (1) N2H4, MS4A/CH3OH–CH2C12, rt, (2) CuCl2, Et3N/CH3OH–CH2C12, rt, 11%; (j) BC13/CH2C12, rt, 59%;
(k) 1,3-propanedithiol, BF3OEt3, TMSC1/CH2C12, rt, 78%; (l) DMF, POC13/CH2C12, rt, 67%; (m) H2, Pd–C, Et3N/C2H5OH, rt, 84%; (n) Phenol, HC1, 70 °C, 32%.
gave 13 (Scheme 3). Carborane-containing compounds were pre-
pared from TBDMS-protected 4,40-dihydroxybenzophenone by
reaction with lithiated p- or m-carborane, followed by reduction
with triethylsilane, and then deprotection to afford the desired
compounds 14 and 15 (Scheme 3). In the case of the o-carboranyl
derivative, adduct of the 4,40-dihydroxybenzophenone with
o-carborane was not isolated because of its instability. Therefore,
substitution of 4,40-dimethoxydiphenylmethyl bromide with
lithiated o-carborane was employed to give 33. Deprotection using
BBr3 afforded 16.
and polar groups including oxygen on the 4,4-position decreased
the binding affinity. In contrast, introduction of substituents at
the 3,3-position had a quite different effect; the binding affinity
of the 3,3-dimethylcyclohexyl compound (7) was significantly
increased. Its potency for binding to ER
a (RBA = 99) was almost
the same as that of estradiol. The bulky trimethylenethioketal
group (10) or pentamethylene group (11) slightly decreased the
affinity. Introduction of halogen at the 3,3-position led to a
decrease of the binding affinity. On the other hand, the 3,3,5,5-
tetramethyl compound (12) showed somewhat lower binding
affinity than 7. Introduction of a bulkier carborane group (14, 15,
16) did not significantly affect the binding affinity. 3,3-Disubsti-
tuted and 4,4-disubstituted cyclohexyl derivatives (2–12) were
2.2. Biological evaluation
The binding activities of the synthesized compounds were
not particularly selective for ER
0.2–1.3). The compounds bearing carboranes showed similar
selectivity (b/ values of 1.2–1.6).
a or ERb (ba in the range of
examined by measurement of the inhibition of [6,7-3H]17b-estra-
diol binding (Kd = 0.4 nM) to human recombinant ER
a
and b, using
a
the nitrocellulose filter binding assay method. Table 1 summarizes
the binding affinity data of the test compounds as relative binding
affinity (RBA) with respect to that of estradiol (estradiol = 100).
Simple unsubstituted cyclohexyl (1), cyclopentyl (13) and 4,
4-dimethylcyclohexyl (2) derivatives showed weak binding
affinity. Introduction of a 4,4-difluoro group (3) did not affect the
binding affinity, and introduction of larger halogens (4, 5, and 6)
The estrogenic activity of the synthesized compounds was eval-
uated by cell proliferation assay with human breast cancer cell line
MCF-7. The EC50 values and IC50 values of these compounds are
summarized in Table 2. Measurement of antagonistic activity
was performed in the presence of 1 ꢁ 10ꢀ11 M estradiol. Unsubsti-
tuted cyclohexyl (1) and 4,4-disubstituted cyclohexyl (2–6) deriva-
tives showed weak agonistic activity (EC50 < 10ꢀ7 M). The potency

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T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
Scheme 3. Synthesis of cyclopentyl derivative and carborane derivatives. Reagents and conditions: (a) anisole, BE3 OEt2, 50 °C, 22%; (b) BBr3/CH2C12, rt, 78–92%; (c) m- or p-
carborane, n-BuLi/(C2H5)2O, rt to ꢀ30 °C; (d) Et3SiH, BF3 OEt2, ꢀ30 °C, 51–57% (two steps); (e) o-carborane, n-BuLi/(C2H5)2O, rt, 28%.
Table 1
Table 2
Relative binding affinity (RBA) of the compound estrogen receptors
a
and b
Estrogenic activities of the compounds in cell proliferation assay using MCF-7 cells
Compound
RBA (E2 = 100)a
b/a
Compound
Agonistic activity
Antagonistic activity
a
b
a
ER
a
ERb
EC50 (nM)
Emax (%)
IC50 (nM)
1
2
3
4
5
6
7
8
6.71
3.64
2.71
0.662
0.957
0.465
98.8
5.78
15.3
50.6
21.9
28.4
4.61
4.08
8.06
11.1
15.5
3.02
3.93
0.158
0.180
0.219
30.7
4.29
15.7
8.43
29.3
35.2
13.7
5.86
6.80
18.4
2.3
1.2
1.3
0.2
0.2
0.5
0.3
0.7
1.0
0.2
1.3
1.2
3.0
1.4
1.2
1.6
1
2
3
4
5
6
7
8
54.1
68.4
22.5
304
211
—
2.40
16.6
4.60
0.39
0.80
—
79
72
80
78
64
22
74
58
70
80
74
32
69
73
86
78
>1000
>1000
>1000
>1000
>1000
>1000
64.9
19.6
9
9
>1000
>1000
>1000
63.1
>1000
>1000
>1000
>1000
10
11
12
13
14
15
16
10
11
12
13
14
15
16
48.8
22.1
2.13
3.42
a
a
Relative binding affinity (RBA) values were determined by radiometric binding
assay with [3H]estradiol and ER
and ERb. Binding assay were performed in
MCF-7 cells were treated with the test compounds (1 ꢁ 10ꢀ13–1 ꢁ 10ꢀ6 M)
a
alone or in the presence of 0.1 nM E2. Cell proliferation assay was performed in
triplicate (n = 3). EC50 and IC50 values were estimated from the sigmoidal dose–
response curves using GraphPad Prism 4 software.
duplicate (n = 2). The RBA of estradiol (E2) is defined as 100.
b
Emax values indicate the efficacy for cell proliferation, based on the value for E2
taken as 100.
was strongly correlated to the binding affinity for ERa. Among
these compounds, unsubstituted cyclohexyl compound 1 showed
very weak antagonistic activity at high concentration. 3,3-Disubsti-
tuted cyclohexyl derivatives (7–11) showed moderate to potent
agonistic activity (EC50 10ꢀ7–10ꢀ10 M). 3,3-Dimethyl (7) and 3,3-
trimethylenethioketal (10) derivatives showed potent ER agonistic
activity, with weak antagonistic activity at high concentration. The
potency of agonistic activity was also correlated to the binding
affinity for ERa. 3,3-Difluorocyclohexyl compound 8 also showed
very weak antagonistic activity at high concentration. The com-
pounds containing a carborane group (14, 15, 16) were moderate
agonists, reflecting their moderate ERa binding affinity. In contrast,
the 3,3,5,5-tetramethyl derivative (12) was a very weak partial

T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
6905
agonist with a maximum activity of 32% of that of estradiol. These
compounds showed antagonistic activity at concentrations of
10ꢀ8 M order.
hydrogen-bonded to Glu353 and Arg394, and the second hydroxyl
group is hydrogen-bonded to Thr347. The hydrophobic 3,
3-dimethylcyclohexyl group is located in a hydrophobic pocket
surrounded by hydrophobic amino acid residues Leu346, Ile424,
Leu428 and Leu391. In docking simulations of 7 using the coordi-
nates of bisphenol A with mutated ER (3UU7)24 and bisphenol C
with ER (3UUC),24 similar stable structures were obtained. Thus,
high binding affinity is associated with the presence of bulky 3,3
substituents. On the other hand, introduction of a carboranyl struc-
ture did not increase the hydrophobic interaction. The compounds
2.3. Discussion
The crystal structure of the ER
complex with estradiol.12 The phenolic hydroxyl group of estradiol
is hydrogen-bonded to Glu353 and Arg394 of ER LBD and the
a LBD has been elucidated in the
a
17-b-hydroxyl group is hydrogen-bonded to the d-nitrogen of
His524. Hydrophobic interaction with hydrophobic amino acid
residues along the body of the steroid skeleton also contributes
to the stability of the complex. Interestingly, the lack of the second
hydrogen-bonding with His524 can be compensated by hydropho-
bic interaction or by alternative hydrogen-bonding, especially with
Thr347. For example, bisphenol bearing bicyclo[2,2,2]octane
(BE1060),20 and many 1,1-diarylethylene derivatives14 have potent
binding affinity for ER. It was reported that reduction of 1,1-diary-
lethylene derivatives greatly decreased the binding affinity.14
Indeed, the binding affinity of compound 1, which corresponds to
hydrogenated cyclofenil, was much lower than that of cyclofenil.
Introduction of methyl at the 4,4-position (2) did not increase
the hydrophobic interaction. Introduction of a larger halogen
(4, and 5), or polar group including oxygen (6) at the 4,4-position
decreased the binding affinity. In contrast, the 3,3-dimethyl deriva-
tive (7) showed greatly increased binding affinity for ER. This is an
example where increased hydrophobic interaction due to the
introduction of a hydrophobic substituent at an appropriate loca-
tion has a dramatic effect on the binding affinity. We performed
in this series were not particularly selective for ER
difference of ER and ERb within the ligand-binding pockets is lim-
ited to two amino acids, that is, Leu384 and Met421 in ER corre-
spond to Met336 and Ile 373 in ERb. It seem not to be effective for
distinction between the differences.
The agonist/antagonist balance of ER ligands depends on the
helix 12 capping conformation of the ER–ligand complex and the
consequent relationship with the steroid receptor coactivator-1
(SRC-1) peptide.12 The characteristic activated conformations of
ER are ‘agonist conformation’ and ‘antagonist conformation’, but
the change of agonist/antagonist balance appears to be continuous,
not stepwise, and small changes of the ligand structure causes
changes of the biological activities. Consequently, there are many
partial agonist/antagonists with various characteristics as regards
potency and agonist/antagonist balance. On the whole, the com-
pounds with 3,3-disubstituted cyclohexyl groups have agonistic
activity together with weak antagonistic activity at high concen-
a or ERb. The
a
a
tration, and their potency corresponds well to their ER
a binding
affinity. Some of them possessed different agonist/antagonist
balances, such as 10 and 11, which were more agonistic. 3,3,5,5-
Tetramethyl derivative (12) was conversely more antagonistic.
Antagonistic activity of unsubstituted and 4,4-disubstituted com-
pounds did not appear, presumably because of their essentially
weak activity.
docking simulation of compounds 7, 11, 12 and 2 into ER
a
(1ERE). Compounds 7 and 11 are racemic, so both enantiomers
were examined. Among the compounds, the most stable complex
was obtained in the case of S-7. Figure 3 shows docking simulation
of S-7 with ER
a using 1ERE. The first hydroxyl group is
Figure 3. Docking simulation of 5–7 into ERa LBD (1ERE).

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T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
(eluent: n-hexane/Et2O, 100:1) gave 19 (93%). 1H NMR (CDCl3) d
3. Conclusion
0.83 (2H, qua, J = 11.4 Hz), 1.11–1.25 (3H, m), 1.57–1.66 (5H, m),
2.00 (1H, quat, J = 10.6, 3.4 Hz), 3.38 (1H, s, J = 10.6 Hz), 3.75 (6H,
s), 6.79 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.16 (4H, ddd, J = 8.7, 2.9,
1.9 Hz); HRMS Calcd for C21H26O2 310.1934. Found 310.1933.
In this paper, we investigated the influence of structural modi-
fication of bis(4-hydroxyphenyl)methanes with cyclic hydrophobic
structure on ER binding affinity and agonist/antagonist balance.
Among the synthetic compounds, 3,3-disubstituted cyclohexyl
4.2.3. 4,40-(Cyclohexylmethylene)bisphenol (1)
derivatives (7–11) possessed potent ERa binding affinity and ago-
To a solution of 19 (9/96.4 mg, 3.21 mmol) in CH2Cl2 (10 mL)
was added a 1.0 M solution of BBr3 in CH2Cl2 (3.90 mL, 3.90 mmol)
at 0 °C under Ar. The mixture was stirred at room temperature for
48 h, then BBr3 in CH2Cl2 (2.20 mL, 2.20 mmol) was added at 0 °C,
and stirring was continued at room temperature for 24 h. The reac-
tion mixture was poured into ice water, and extracted with AcOEt.
The organic layer was washed with brine, dried over MgSO4, and
concentrated. Purification of the residue by silica column chro-
matography (eluent: n-hexane/AcOEt, 3:1) gave 1 (62%). 1: white
fibrous solid (CH2Cl2); mp 228–229 °C; 1H NMR (DMSO-d6) d
0.74 (2H, br qua, J = 10.6 Hz), 1.00–1.21 (3H, br m), 1.46 (2H, br
d, J = 12.6 Hz), 1.51–1.64 (3H, br m), 1.97 (1H, br qua,
J = 10.6 Hz), 3.25 (1H, d, J = 10.6 Hz), 6.61 (4H, d, J = 7.7 Hz), 7.04
(4H, d, J = 7.7 Hz), 9.07 (2H, s); 13C NMR (DMSO-d6) d 25.7, 26.1,
29.8, 31.6, 56.8, 115.0, 128.5, 135.5, 155.1; HRMS Calcd for
nistic activity, together with weak antagonistic activity at high
concentration. This result confirms the importance of hydrophobic
structure for binding and modulation of ER activity. Partial agonists
with various agonist/antagonist activity balances should be candi-
dates for tissue-selective estrogen receptor modulators. Further
structure–function studies of these compounds and their deriva-
tives could lead to the development of more selective and potent
estrogen receptor modulators.
4. Experimental section
4.1. General remarks
Melting points were determined with a Yanaco micro melting
point apparatus and were not corrected. 1H NMR and 13C NMR
spectra were recorded with JEOL JNM-LA-400 spectrometers.
Chemical shifts for 1H NMR spectra were referenced to tetram-
ethylsilane (0.0 ppm) as an internal standard. Chemical shifts for
13C NMR spectra were referenced to residual 13C present in deuter-
ated solvents. The splitting patterns are designated as follows: s
(singlet), d (doublet), t (triplet), q (quartet) and m (multiplet). Mass
C19H22O2 282.1621. Found 282.1625; Anal. Calcd for C19H22O2: C,
80.82; H, 7.85. Found C, 80.76; H, 7.45.
4.2.4. 1,10-(4,4-Dimethylcyclohexylidenemethylene)bis
(4-methoxy)benzene (18)
McMurry coupling of 4,40-dimethoxybenzophenone and 4,4-
dimethylcyclohexanone was performed by the same method as
that used for preparation of 17. Purification by silica gel column
chromatography (eluent: n-hexane/AcOEt, 40:1) gave 18 (85%).
1H NMR (CDCl3) d 0.98 (6H, s), 1.37 (4H, t, J = 6.4 Hz), 2.27 (4H, t,
J = 6.4 Hz), 3.79 (6H, s), 6.81 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.03
(4H, ddd, J = 8.7, 2.9, 1.9 Hz); HRMS Calcd for C23H28O2 336.2090.
Found 336.2095.
spectra were recorded on
a JEOL JMS-DX-303 spectrometer.
Elemental analyses were performed with a Perkin Elmer 2400
CHN spectrometer. Column chromatography was carried out using
Merck silica gel 60 (0.063–0.200 lm) and TLC was performed on
Merck silica gel F254. Carboranes were purchased from Katchem
s.r.o. (Prague, Czech Republic). Other reagents were purchased
from Wako Pure Chemical Industries, Ltd, Sigma–Aldrich Co., and
Tokyo Chemical Industry, Ltd (TCI). All solvents were commercial
products of reagent quality, and were used without further
purification.
4.2.5. 1,10-(4,4-Dimethylcyclohexylmethylene)bis(4-methoxy)
benzene (20)
Compound 20 was prepared from 18 by the same method as
that used for preparation of 19. Purification by silica gel column
chromatography (eluent: n-hexane/Et2O, 100:1) gave 20 (quant).
1H NMR (CDCl3) d 0.85 (3H, s), 0.87 (3H, s), 1.03 (2H, quad,
J = 13.1, 2.8 Hz), 1.14 (2H, td, J = 13.1, 2.8 Hz), 1.32 (2H, d,
J = 13.2 Hz), 1.41 (2H, d, J = 13.2 Hz), 1.92 (1H, quat, J = 10.7,
3.4 Hz), 3.40 (1H, d, J = 10.7 Hz), 3.74 (6H, s), 6.79 (4H, ddd,
J = 8.8, 2.9, 2.0 Hz), 7.16 (4H, ddd, J = 8.8, 2.9, 2.0 Hz); HRMS Calcd
for C23H30O2 338.2247. Found 338.2247.
4.2. Synthesis
4.2.1. 1,10-(Cyclohexylidenemethylene)bis(4-methoxy)benzene
(17)
To a suspension of zinc powder (6.30 g, 97.0 mmol) in THF
(65 mL) was added dropwise titanium (IV) chloride (5.20 mL,
47.3 mmol) at ꢀ10 °C under Ar. The mixture was refluxed with
stirring for 18 h. To the reaction mixture was added dropwise a
solution of 4,40-dimethoxybenzophenone (3.10 g, 12.8 mmol) and
cyclohexanone (1.30 g, 13.3 mmol) in THF (60 mL). The mixture
was refluxed with stirring for 4.5 h, then cooled to room tempera-
ture, and poured slowly into saturated aqueous NaHCO3. Et2O was
added to the aqueous solution, and the heterogeneous solution was
filtered through Celite. The filtrate was extracted with Et2O,
washed with brine, dried over MgSO4, and concentrated.
Purification of the residue by silica gel column chromatography
(eluent: n-hexane/AcOEt, 20:1) gave 17 (74%). 1H NMR (CDCl3) d
1.55–1.65 (6H, m), 2.20–2.28 (4H, m), 3.78 (6H, s), 6.80 (4H, ddd,
J = 8.8, 2.9, 2.4 Hz), 7.02 (4H, ddd, J = 8.8, 2.9, 2.4 Hz); HRMS Calcd
for C21H24O2 308.1777. Found 308.1785.
4.2.6. 4,40-[(4,4-Dimethylcyclohexyl)methylene]bisphenol (2)
Deprotection of 18 was performed by the same method as that
used for preparation of 1. Purification by silica gel column chro-
matography (eluent: n-hexane/AcOEt, 2:1) gave 2 (90%). 2: white
needles (Et2O–n-hexane); mp 221 °C; 1H NMR (CDCl3) d 0.85 (3H,
s), 0.87 (3H, s), 1.02 (2H, quad, J = 13.0, 2.9 Hz), 1.14 (2H, td,
J = 13.0, 2.9 Hz), 1.32 (2H, br d, J = 13.0 Hz), 1.40 (2H, br d,
J = 13.0 Hz), 1.89 (1H, quat, J = 11.1, 3.4 Hz), 3.38 (1H, d,
J = 11.1 Hz), 4.53 (2H, s), 6.72 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.10
(4H, ddd, J = 8.7, 2.9, 1.9 Hz); 13C NMR (CDCl3) d 24.4, 27.8, 39.9,
32.7, 39.2, 41.6, 57.5, 115.2, 129.0, 137.3, 153.5; HRMS Calcd for
C
21H26O2 310.1934. Found 310.1939; Anal. Calcd for C21H26O2: C,
4.2.2. 1,10-(Cyclohexylmethylene)bis(4-methoxy)benzene (19)
A suspension of 17 (92.4 mg, 0.300 mmol) and 10% palladium
on carbon (15.9 mg, 0.0150 mmol) in AcOEt (3 mL) was stirring
at room temperature under hydrogen gas for 24 h. The mixture
was filtered through Celite, and the filtrate was concentrated.
Purification of the residue by silica gel column chromatography
81.25; H, 8.44. Found C, 81.28; H, 8.59.
4.2.7. 4-[Bis(4-dimethoxyphenyl)methylene]cyclohexanone
(21)
McMurry coupling of 4,40-dimethoxybenzophenone and 1,4-
cyclohexanedione mono-ethylene ketal was performed by the

T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
6907
same method as that used for preparation of 17. Purification by sil-
ica gel column chromatography (eluent: n-hexane/Et2O, 5:1) gave
a mixture of ethylene ketal product and deprotected product
(98:2, 8.50 g). To a suspension of the mixture (8.50 g) and silica
gel (104 g) in CH2Cl2 (229 mL) was added 15% aqueous H2SO4
(14.8 mL) at room temperature. The mixture was stirred at room
temperature for 44 h, then filtered, and the filtrate was extracted
with AcOEt. The organic layer was washed with brine, dried over
Na2SO4, and concentrated. The residue was used for the next reac-
tion. A suspension of the residue (7.00 g) and 10% palladium on
carbon (4.60 g, 4.34 mmol) in AcOEt (70 mL) was stirred at room
temperature under hydrogen gas for 48 h. The mixture was filtered
through Celite, and concentrated. Purification of the residue by sil-
ica gel column chromatography (eluent: n-hexane/Et2O, 3:1) gave
21 (53%, 3 steps). 1H NMR (CDCl3) d 1.33–1.43 (2H, m), 1.89–1.99
(2H, m), 2.29–2.37 (4H, m), 2.50 (1H, quat, J = 10.7, 3.4 Hz), 3.49
(1H, d, J = 10.7 Hz), 3.76 (6H, s), 6.82 (4H, ddd, J = 8.8, 2.9,
2.0 Hz), 7.20 (4H, ddd, J = 8.8, 2.9, 2.0 Hz). HRMS Calcd for
for 31 h under Ar, then cooled to room temperature, and filtered.
The filtrate was concentrated. The residue was dissolved in CH2Cl2,
and this solution was washed with H2O and brine, dried over
Na2SO4, and concentrated. Purification by trituration (n-hexane)
gave the benzyl-protected product (86%). 1H NMR (CDCl3) d
1.33–1.43 (2H, m), 1.94 (2H, br d, J = 12.2 Hz), 2.27–2.35 (4H, m),
2.49 (1H, quat, J = 10.7, 2.5 Hz), 3.49 (1H, d, J = 10.7 Hz), 5.01 (4H,
s), 6.90 (4H, ddd, J = 8.8, 2.9, 2.0 Hz), 7.20 (4H, ddd, J = 8.8, 2.9,
2.0 Hz), 7.29–7.42 (10H, m); HRMS Calcd for C33H32O3 476.2353.
Found 476.2348. To a suspension of active MS4A (2.80 g) in MeOH
(14 mL) was added hydrazine monohydrate (2.80 mL, 57.7 mmol)
at room temperature under argon gas. After 1 h, a solution of the
benzyl-protected compound (1.30 g, 2.75 mmol) in CH2Cl2
(10 mL) was added dropwise. The mixture was stirred at room
temperature for 2 h, then filtered, and concentrated. Excess
hydrazine was further removed from the residue under vacuum
(1–2 Torr) with heating at 30 °C for 4 h. The residue was used for
the next reaction. To a solution of CuCl2 (2.20 g, 16.5 mmol) in
MeOH (17 mL) was added Et3N (1.20 mL, 8.26 mmol) with stirring
at room temperature. After 30 min, a solution of the residue in
CH2Cl2 (10 mL) was added dropwise at 0 °C. The reaction mixture
was stirred at room temperature for 4 h, then 28% aqueous NH3
was added. The whole was extracted with CH2Cl2, washed with
brine, dried over Na2SO4, and concentrated. Purification of the resi-
due by silica gel column chromatography (eluent: n-hexane/AcOEt,
300:1) gave the dichloro product (32%, 2 steps). 1H NMR (CDCl3) d
1.40 (2H, br quad, J = 12.7, 3.4 Hz) 1.60 (2H, br d, J = 11.6 Hz), 2.02–
2.18 (3H, br m), 2.47 (2H, br d, J = 13.5 Hz), 3.43 (1H, d, J = 11.1 Hz),
5.00 (4H, s), 6.88 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.16 (4H, ddd, J = 8.7,
2.9, 1.9 Hz), 7.32–7.39 (10H, m); HRMS Calcd for C33H3235Cl2O2
C21H24O3 324.1726. Found 324.1734.
4.2.8. 4,40-[(4,4-Difluorocyclohexyl)methylene]bisphenol (3)
To a solution of 21 (0.970 g, 2.99 mmol) in CH2Cl2 (30 mL) was
added dropwise a 1.0 M solution of BBr3 in CH2Cl2 (12.0 mL,
12.0 mmol) at ꢀ60 °C under argon gas. The mixture was stirred
at ꢀ40 °C for 21 h, then poured into ice water, and extracted with
AcOEt. The organic layer was washed with brine, dried over
Na2SO4, and concentrated. Purification of the residue by silica gel
column chromatography (eluent: n-hexane/AcOEt = 2:1) gave the
phenol (61%). 1H NMR (acetone-d6) d 1.29 (2H, br quad, J = 12.1,
4.3 Hz), 1.79–1.89 (2H, m), 2.16 (2H, br t, J = 14.5 Hz), 2.30 (2H,
br td, J = 13.6, 5.5 Hz), 2.59 (1H, quat, J = 10.6, 3.4 Hz), 3.50 (1H,
d, J = 11.1 Hz), 6.71 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.17 (4H, ddd,
530.1781. Found 530.1770. To
a solution of the compound
(210 mg, 0.395 mmol) in CH2Cl2 (4 mL) was added dropwise a
1.0 M solution of BCl3 in CH2Cl2 (0.800 mL, 0.800 mmol) at 0 °C.
The mixture was stirred at room temperature for 19 h, then poured
into ice water, and the whole was extracted with CH2Cl2. The
organic layer was washed with brine, dried over MgSO4, and
concentrated. Purification of the residue by silica gel column chro-
matography (eluent: CH2Cl2/AcOEt, 25:1) gave 4 (78%). 4: white
plates (toluene); mp 77–78 °C; 1H NMR (DMSO-d6) d 1.21 (2H, br
qua, J = 12.2 Hz), 1.47 (2H, br d, J = 11.6 Hz), 2.19 (3H, br t,
J = 11.3 Hz), 2.38 (2H, br d, J = 13.5 Hz), 3.34 (1H, d, J = 7.7 Hz),
6.62 (4H, d, J = 8.7 Hz), 7.07 (4H, d, J = 8.7 Hz), 9.11 (2H, s); 13C
NMR (CDCl3) d 29.1, 39.8, 45.9, 56.6, 91.6, 115.4, 128.8, 136.3,
153.9; HRMS Calcd for C19H2035Cl2O2 350.0842. Found 350.0838;
Anal. Calcd for C19H20Cl2O2ꢂ1/2C7H8: C, 68.01; H, 6.09. Found: C,
67.66; H, 6.23.
J = 8.7, 2.9, 1.9 Hz), 8.08 (2H, s). HRMS Calcd for
C19H20O3
296.1413. Found 296.1412. To a solution of the crude compound
(210 mg, 0.709 mmol) in CH2Cl2 (10 mL) was added DAST
(0.560 mL, 4.17 mmol) at room temperature under Ar. The mixture
was stirred at room temperature for 28 h. To the reaction mixture
was added MeOH/H2O (1:1), and the resulting mixture was
extracted with CH2Cl2. The organic layer was washed with H2O,
saturated aqueous NaHCO3, and brine, dried over MgSO4, and con-
centrated. The residue was used directly for the next reaction. To a
solution of the residue and 97% NMO (337 mg, 2.80 mmol) in ace-
tone/water (4:1, 35 mL) was added 0.1 M OsO4 in THF (0.350 mL,
0.0350 mmol) at 0 °C. The mixture was stirred at room tempera-
ture for 63 h, then NaHSO3 (35.0 mg) was added, and the whole
was concentrated. The residue was diluted with Et2O, and washed
with H2O. The organic layer was extracted with 10% aqueous
NaOH. The water layer was acidified with 10% aqueous HCl to pH
1–2, and extracted with Et2O. The organic layer was washed with
saturated aqueous NaHCO3 and brine, dried over MgSO4, and con-
centrated. Purification of the residue by silica gel column chro-
matography (eluent: n-hexane/AcOEt, 3:1) gave 3 (50%). 3: White
needles (Et2O–n-hexane); mp 194–195 °C; 1H NMR (DMSO-d6) d
1.00–1.09 (2H, m), 1.51 (2H, br d, J = 13.5 Hz), 1.72 (2H, br dt,
J = 32.8, 13.4 Hz), 1.90–1.95 (2H, br m), 2.18 (1H, br qua,
J = 9.7 Hz), 3.36 (1H, d, J = 11.1 Hz), 6.62 (4H, d, J = 8.7 Hz), 7.08
(4H, d, J = 8.7 Hz), 9.12 (2H, s); 13C NMR (CDCl3) d 27.9, 33.5,
39.8, 56.3, 115.4, 123.7, 128.8, 136.4, 153.9; HRMS Calcd for
4.2.10. 4,40-[(4,4-Dibromocyclohexyl)methylene]bisphenol (5)
To a solution of 21 (1.30 g, 4.01 mmol) and 2,3-dihydroxynaph-
thalene (1.90 g, 12.0 mmol) in CH2Cl2 (40 mL) was added 47%
BF3ꢂOEt2 in Et2O (0.400 mL, 1.30 mmol) and TMSCl (1.20 mL,
9.47 mmol) at 0 °C. The mixture was stirred at room temperature
for 9 h, then MeOH (2 mL) was added at 0 °C. The whole was
washed with 10% aqueous NaOH, H2O, and brine, dried over
Na2SO4, and concentrated. Purification of the residue by silica gel
column chromatography (eluent: n-hexane/AcOEt, 10:1) gave the
2,3-dihydroxynaphthalene ketal product (75%). 1H NMR (CDCl3) d
1.42 (2H, br qua, J = 13.2 Hz), 1.73 (2H, br d, J = 13.7 Hz), 1.82
(2H, td, J = 13.2, 3.9 Hz), 2.09 (2H, br d, J = 12.2 Hz), 2.16 (1H, quat,
J = 10.7, 3.4 Hz) 3.52 (1H, d, J = 10.7 Hz), 3.76 (6H, s), 6.82 (4H, d,
J = 8.8 Hz), 7.01 (2H, d, J = 7.3 Hz), 7.21 (4H, d, J = 8.8 Hz),
7.27–7.29 (2H, m), 7.62 (2H, dt, J = 6.3, 2.9 Hz); HRMS Calcd for
C19H20F2O2 318.1432. Found 318.1428; Anal. Calcd for
C19H20F2O2: C, 71.68; H, 6.33. Found C, 71.49; H, 6.34.
4.2.9. 4,40-[(4,4-Dichlorocyclohexyl)methylene]bisphenol (4)
Compound 21 was deprotected by the same method as that
used for preparation of 3. A suspension of the phenol (0.940 g,
3.17 mmol), benzyl bromide (0.830 mL, 6.80 mmol) and K2CO3
(1.30 g, 9.42 mmol) in acetone (64 mL) was refluxed with stirring
C31H30O4 466.2145. Found 466.2138. To a solution of this com-
pound (34.0 mg, 0.0730 mmol) in CH2Cl2 (1 mL) was added a
1.0 M solution of BBr3 in CH2Cl2 (0.560 mL, 0.560 mmol) at
ꢀ60 °C. The mixture was stirred at ꢀ40 °C for 7 days, then poured

6908
T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
into ice water, and extracted with AcOEt. The organic layer was
washed with brine, dried over MgSO4, and concentrated. Purifica-
tion by silica gel column chromatography (eluent: n-hexane/Et2O,
1:1) gave 5 (81%). 5: colorless needles (toluene); mp 93–94 °C;
1H NMR (CDCl3) d 1.38–1.49 (4H, m), 2.09 (1H, quat, J = 10.6,
4.3 Hz), 2.33 (2H, td, J = 15.0, 3.9 Hz), 2.65 (2H, br d, J = 13.0 Hz),
3.42 (1H, d, J = 11.1 Hz), 4.61 (1H, s), 4.62 (1H, s), 6.73 (4H, ddd,
J = 8.7, 2.9, 1.9 Hz), 7.10 (4H, ddd, J = 8.7, 2.9, 1.9 Hz); 13C NMR
(CDCl3) d 30.2, 39.8, 49.0, 56.7, 71.5, 115.4, 128.8, 136.2, 153.8;
HRMS Calcd for C19H2078Br2O2 437.9831. Found: 437.9817; Anal.
Calcd for C19H20Br2O2ꢂ1.0C7H8: C, 58.67; H, 5.30. Found: C, 58.49;
H, 5.31.
1.25–1.41 (5H, m), 2.19 (1H, qua, J = 11.1 Hz), 3.30 (1H, d,
J = 10.6 Hz), 3.75 (3H, s), 3.76 (3H, s), 6.78 (2H, d, J = 8.7 Hz), 6.80
(2H, d, J = 8.7 Hz), 7.14 (4H, d, J = 8.7 Hz); HRMS Calcd for
C23H30O2 338.2247. Found 338.2234.
4.2.14. 4,40-[(3,3-Dimethylcyclohexyl)methylene]bisphenol (7)
To a solution of 24 (200 mg, 0.592 mmol) in CH2Cl2 (5 mL) was
added dropwise a 1.0 M solution of BBr3 in CH2Cl2 (1.20 mL,
1.20 mmol) at 0 °C under argon gas. The mixture was stirred at
room temperature for 9 h, then poured into ice water, and the
whole was extracted with AcOEt. The organic layer was washed
with brine, dried over MgSO4, and concentrated. Purification of
the residue by silica gel column chromatography (eluent: n-hex-
ane/Et2O, 1:1) gave 7 (quant). 7: white leaflets (Et2O–n-hexane);
mp 192–193 °C; 1H NMR (acetone-d6) d 0.61–0.73 (2H, m), 0.79
(3H, s), 0.90 (3H, s), 1.06 (1H, td, J = 12.9, 5.0 Hz), 1.25–1.60 (5H,
m), 2.26 (1H, quat, J = 11.4, 2.9 Hz), 3.26 (1H, d, J = 10.6 Hz),
6.69–6.71 (4H, m), 7.10–7.12 (4H, m), 8.01, (1H, s), 8.03 (1H, s);
13C NMR (acetone-d6) d 23.1, 25.0, 31.5, 32.8, 33.9, 38.0, 40.0,
46.0, 58.9, 115.8, 115.9, 129.6, 136.9, 137.2, 156.2; HRMS Calcd
4.2.11. 4-[Bis(4-hydroxyphenyl)methylene]cyclohexanone
ethylene ketal (6)
McMurry coupling of 4,40-dihydroxybenzophenone and 1,4-
cyclohexanedione mono-ethylene ketal was performed by the
same method as that used for preparation of 17. Purification by sil-
ica gel column chromatography (eluent: CHCl3/AcOEt, 2:1) gave
the product (69%). 1H NMR (CDCl3) d 1.71 (4H, br t, J = 6.3 Hz),
2.39 (4H, br t, J = 6.3 Hz), 3.97 (4H, s), 4.69 (2H, s), 6.74 (4H, ddd,
J = 8.3, 2.9, 2.0 Hz), 6.97 (4H, ddd, J = 8.3, 2.9, 2.0 Hz); HRMS Calcd
for C21H22O4 338.1519. Found 338.1517. This compound was
reduced by the same method as that used for preparation of 19
to give the hydrogenated product. Purification by silica gel column
chromatography (eluent: n-hexane/Et2O, 3:1) gave 6 (quant). 6:
white columns (Et2O–n-hexane); mp 230–231 °C; 1H NMR (ace-
tone-d6) d 0.97–1.08 (2H, m), 1.32 (2H, td, J = 12.8, 3.9 Hz), 1.42
(2H, br d, J = 13.5 Hz), 1.51 (2H, br d, J = 12.6 Hz), 2.00 (1H, quat,
J = 11.1, 3.4 Hz), 3.26 (1H, d, J = 10.6 Hz), 3.72 (4H, s), 6.59 (4H,
ddd, J = 8.7, 2.9, 1.9 Hz), 7.02 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.90
(2H, s); 13C NMR (DMSO-d6) d 28.7, 30.7, 33.8, 55.8, 63.5, 108.2,
115.0, 128.5, 135.5, 155.2; HRMS Calcd for C21H24O4 340.1675.
Found 340.1675; Anal. Calcd for C21H24O4: C, 74.09; H, 7.11. Found
C, 74.41; H, 7.13.
for
C21H26O2 310.1934. Found 310.1939; Anal. Calcd for
C21H26O2: C, 81.25; H, 8.44. Found C, 81.19; H, 8.36.
4.2.15. 1,10-(3,3,5,5-Tetramethylcyclohexylidenemethylene)bis
(4-methoxy)benzene (23)
McMurry coupling of 4,40-dimethoxybenzophenone and 3,3,5,5-
tetramethylcyclohexanone was performed by the same method as
that used for preparation of 22. Purification by silica gel column
chromatography (eluent: n-hexane/AcOEt, 40:1) gave 23 (83%).
1H NMR (CDCl3) d 0.93 (12H, s), 1.28 (2H, s), 1.98 (4H, s), 3.78
(3H, s), 3.79 (3H, s), 6.80 (4H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.07 (4H,
ddd, J = 8.7, 2.9, 1.9 Hz); HRMS Calcd for C25H32O2 364.2404. Found
364.2397.
4.2.16. 1,10-(3,3,5,5-Tetramethylcyclohexylmethylene)bis(4-
methoxy)benzene (25)
4.2.12. 1,10-(3,3-Dimethylcyclohexylidenemethylene)bis(4-
methoxy)benzene (22)
Compound 25 was prepared from 200 mg of 23 by the same
method as that used for preparation of 24. Purification by silica
gel column chromatography (eluent: n-hexane/Et2O, 80:1) gave
25 (48%). 1H NMR (CDCl3) d 0.63 (1H, d, J = 12.6 Hz), 0.66 (1H, d,
J = 12.6 Hz), 0.80 (6H, s), 0.99 (6H, s), 1.00 (1H, d, J = 13.0 Hz),
1.23 (1H, d, J = 14.0 Hz), 1.33 (2H, d, J = 13.0 Hz), 2.40 (1H, quat,
J = 10.6, 2.9 Hz), 3.33 (1H, d, J = 10.6 Hz), 3.79 (6H, s), 6.80 (4H,
ddd, J = 8.7, 2.9, 1.9 Hz), 7.14 (4H, ddd, J = 8.7, 2.9, 1.9 Hz); HRMS
Calcd for C25H34O2 366.2560. Found 366.2563.
To a suspension of zinc powder (1.50 g, 23.1 mmol) in THF
(15 mL) was added dropwise titanium(IV) chloride (1.20 mL,
10.9 mmol) at ꢀ10 °C under Ar. The mixture was refluxed with
stirring for 3 h, and a solution of 4,40-dimethoxybenzophenone
(726 mg, 3.00 mmol) and 3,3-dimethylcyclohexanone (0.420 mL,
3.03 mmol) in THF (13 mL) was added dropwise. The mixture
was refluxed with stirring for 2 h. The reaction mixture was cooled
to room temperature, and poured slowly into saturated aqueous
NaHCO3. Et2O was added to the aqueous solution, and the hetero-
geneous mixture was filtered through Celite. The filtrate was
extracted with Et2O, washed with brine, dried over MgSO4, and
concentrated. Purification of the residue by silica gel column chro-
matography (eluent: n-hexane/AcOEt, 40:1) gave 22 (90%). 1H NMR
(CDCl3) d 0.86 (6H, s), 1.39 (2H, t, J = 6.1 Hz), 1.58–1.64 (2H, m),
2.00 (2H, s), 2.18 (2H, t, J = 6.3 Hz), 3.776 (3H,s), 3.781 (3H, s),
6.77–6.83 (4H, m), 7.00 (2H, d, J = 8.8 Hz), 7.04 (2H, d, J = 8.8 Hz);
HRMS Calcd for C23H28O2 336.2090. Found 336.2083.
4.2.17. 4,40-[(3,3,5,5-Tetramethylcyclohexyl)methylene]
bisphenol (12)
Compound 12 was prepared from 200 mg of 25 by the same
method as that used for preparation of 7. Purification by silica
gel column chromatography (eluent: n-hexane/AcOEt, 2:1) gave
12 (96%). 12: white lamellar solid (Et2O–n-hexane); mp 203 °C;
1H NMR (CDCl3)
d 0.62 (1H, d, J = 12.6 Hz), 0.65 (1H, d,
J = 12.6 Hz), 0.80 (6H, s), 0.98 (6H, s), 1.00 (1H, d, J = 12.6 Hz),
1.23 (1H, d, J = 13.5 Hz), 1.31 (2H, d, J = 12.6 Hz), 2.36 (1H, quat,
J = 10.6, 2.4 Hz), 3.30 (1H, d, J = 10.6 Hz), 4.53 (2H, s), 6.72 (4H,
ddd, J = 8.2, 2.9, 1.9 Hz), 7.09 (4H, ddd, J = 8.2, 2.9, 1.9 Hz); 13C
NMR (CDCl3) d 27.4, 31.7, 34.3, 35.4, 45.2, 52.4, 57.5, 115.3,
128.9, 137.4, 153.5; HRMS Calcd for C24H30O2 338.2247. Found
338.2234; Anal. Calcd for C23H30O2: C, 81.62; H, 8.93. Found C,
81.60; H, 9.05.
4.2.13. 1,10-(3,3-Dimethylcyclohexylmethylene)bis(4-methoxy)
benzene (24)
To a solution of the above compound (67.0 mg, 0.183 mmol) in
CH2Cl2 (1.6 mL) was added 57% aqueous hydriodic acid (1.60 mL,
7.13 mmol). The mixture was refluxed with stirring for 24 h, then
cooled to room temperature, washed with aqueous Na2S2O3, and
brine, dried over Na2SO4, and concentrated. Purification of the resi-
due by silica gel column chromatography (eluent: n-hexane/Et2O,
100:1) gave 24 (11%). 1H NMR (CDCl3) d 0.70 (2H, td, J = 12.4,
3.7 Hz), 0.81 (3H, s), 0.9 (s, 3H), 1.05 (1H, td, J = 13.2, 4.5 Hz),
4.2.18. 3-[Bis(4-hydroxyphenyl)methylene]cyclohexanone (26)
McMurry coupling of 4,40-dihydroxybenzophenone and 1,3-
cyclohexanedione mono-ethylene ketal was performed by the
same method as that used for preparation of 22. Purification by

T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
6909
silica gel column chromatography (eluent: n-hexane/AcOEt, from
3:1 to 2:1) gave the methylene product (45%). 1H NMR (CDCl3) d
1.62–1.68 (2H, m), 1.77 (2H, t, J = 5.8 Hz), 2.25 (2H, t, J = 5.8 Hz),
2.41 (2H, s), 3.82–3.94 (4H, m), 4.79 (2H, s), 6.73 (4H, d,
J = 8.7 Hz), 6.98 (2H, d, J = 8.7 Hz), 7.05 (2H, d, J = 8.7 Hz); HRMS
Calcd for C21H22O4 338.1519. Found 338.1522. To a solution of this
compound (1.92 g, 5.68 mmol) in acetone (30 mL) was added 1.0 M
aqueous HCl (0.290 mL, 0.290 mmol) at 0 °C. The mixture was stir-
red at room temperature for 48 h, then saturated aqueous NaHCO3
and H2O were added, and acetone was removed. The residue was
extracted with AcOEt, and the organic solution was washed with
brine, dried over MgSO4, and concentrated. Purification by silica
gel column chromatography (eluent: n-hexane/AcOEt, from 2:1
to 1:1) gave the unsaturated ketone product (74%). 1H NMR
(DMSO-d6) d 1.84 (2H, qua, J = 6.3 Hz), 2.21 (2H, t, J = 5.8 Hz),
2.26 (2H, t, J = 6.5 Hz), 4.79 (1H, s), 5.35 (1H, s), 6.69 (4H, d,
J = 8.2 Hz), 6.91 (4H, d, J = 8.2 Hz), 9.31 (2H, s); HRMS Calcd for
6.88 (2H, d, J = 8.7 Hz), 6.91 (2H, d, J = 8.7 Hz), 7.12 (2H, d,
J = 8.7 Hz), 7.16 (2H, d, J = 8.2 Hz), 7.29–7.43 (10H, m); HRMS Calcd
for C33H3235Cl2O2 530.1782 Found 530.1794. To a solution of the
dichloro product (49.8 mg, 0.0938 mmol) in CH2Cl2 (1 mL) was
added dropwise a 1.0 M solution of BCl3 in CH2Cl2 (0.200 mL,
0.200 mmol) at 0 °C. The mixture was stirred at room temperature
for 110 h, then poured into ice water, and the whole was extracted
with CH2Cl2. The organic layer was washed with brine, dried over
MgSO4, and concentrated. Purification by silica gel column chro-
matography (eluent: CH2Cl2/AcOEt, 3:1) gave 9 (59%). 9: white
leaflets (toluene); mp 77–78; 1H NMR (CDCl3) d 0.80–0.90 (1H,
m), 1.60–1.74 (4H, m), 2.00–2.08 (1H, m), 2.50–2.57 (3H, m),
3.39 (1H, d, J = 10.6 Hz), 4.61 (1H, s), 4.64 (1H, s), 6.73 (2H, ddd,
J = 8.7, 2.9, 1.9 Hz), 6.76 (2H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.09 (2H, d,
J = 8.2 Hz), 7.11 (2H, d, J = 8.2 Hz); 13C NMR (CDCl3) d 23.5, 30.1,
39.2, 46.2, 51.3, 56.5, 91.9, 115.4, 115.6, 125.3, 128.2, 128.87,
128.90, 129.0, 135.4, 136.1, 153.85, 153.92; HRMS Calcd for
C
19H18O3 294.1256. Found 294.1245. A suspension of this com-
pound (30 mg, 0.102 mmol) and 10% palladium on carbon
(5.02 mg, 4.74 mol) in EtOH (1 mL) was stirred at room tempera-
C
19H2035Cl2O2 350.0842. Found: 350.0833; Anal. Calcd for
C19H20Cl2O2ꢂC7H8: C, 70.43; H, 6.36. Found: C,70.63; H, 6.50.
l
ture under hydrogen gas for 48 h, then filtered through Celite, and
concentrated. Purification of the residue by silica gel column chro-
matography (eluent: n-hexane/AcOEt, 2:1) gave 26 (88%). 1H NMR
(acetone-d6) d 1.25–1.37 (1H, m), 1.57 (1H, quat, J = 12.2, 4.1 Hz),
1.72–1.82 (1H, m), 1.91–1.99 (2H, m), 2.11–2.23 (2H, m), 2.28
(1H, td, J = 13.2, 5.6 Hz), 2.57 (1H, quat, J = 10.8, 3.6 Hz), 3.54 (1H,
d, J = 10.6 Hz), 6.72 (2H, d, J = 8.2 Hz), 6.74 (2H, d, J = 8.2 Hz), 7.13
(2H, ddd, J = 8.2, 2.9, 1.9 Hz), 7.18 (2H, ddd, J = 8.2, 2.9, 1.9 Hz),
8.11 (1H, s), 8.12 (1H, s). HRMS Calcd for C29H20O3 296.1413. Found
296.1410.
4.2.21. 3-[Bis(4-hydroxyphenyl)methylene]cyclohexanone
trimethylenethioketal (10)
To a solution of 26 (116 mg, 0.392 mmol) and 1,3-propane-
dithiol (82.0
47% BF3ꢂOEt2 in Et2O (351
l
L, 0.820 mmol) in CH2Cl2 (4 mL) was added dropwise
L, 1.17 mmol) at 0 °C under argon gas.
l
The mixture was stirred at room temperature for 2.5 h, then
poured into saturated aqueous NaHCO3, and extracted with Et2O.
The organic solution was washed with brine, dried over Na2SO4,
and concentrated. Purification of the residue by silica gel column
chromatography (eluent: n-hexane/AcOEt, 2:1) gave 10 (78%). 10:
white fibrous solid (toluene); mp 86–87 °C; 1H NMR (CDCl3) d
0.82 (1H, quad, J = 12.1, 3.9 Hz), 1.17–1.24 (1H, m), 1.52–1.62
(3H, m), 1.72 (1H, quat, J = 13.0, 2.9 Hz), 1.89–1.94 (2H, m), 2.26
(1H, d, J = 12.1 Hz), 2.34 (1H, dd, J = 13.5, 1.9 Hz), 2.49–2.62 (2H,
m), 2.67–2.75 (3H, m), 3.35 (1H, d, J = 11.1 Hz), 4.60 (1H, s), 4.62
(1H, s), 6.72 (2H, d, J = 8.7 Hz), 6.75 (2H, d, J = 8.7 Hz), 7.10 (2H,
d, J = 8.7 Hz), 7.14 (2H, d, J = 8.7 Hz); 13C NMR (CDCl3) d 21.9,
25.8, 26.0, 26.1, 31.6, 37.2, 38.0, 42.8, 50.6, 57.0, 115.3, 115.5,
128.6, 128.9, 136.4, 136.6, 153.68, 153.71; HRMS Calcd for
4.2.19. 4,40-[(3,3-Difluorocyclohexyl)methylene]bisphenol (8)
To a solution of 26 (296 mg, 1.00 mmol) in CH2Cl2 was added
DAST (0.780 mL, 5.81 mmol) at 0 °C. The mixture was stirred at
room temperature for 22 h, then MeOH/H2O (1:1) was added at
0 °C, and the whole was extracted with AcOEt. The organic layer
was washed with saturated aqueous NaHCO3 and brine, dried over
MgSO4, and concentrated. Purification of the residue by silica gel
column chromatography (eluent: n-hexane/AcOEt, 3:1) and recrys-
tallization (Et2O–n-hexane) gave 8 (46%). 8: white lamellar solid
(Et2O–n-hexane); mp 194–195 °C; 1H NMR (actone-d6) d 0.93
(1H, qua, J = 12.9 Hz), 1.26–1.50 (2H, m), 1.55–1.79 (3H, m),
1.83–1.98 (2H, m), 2.38 (1H, qua, J = 11.1 Hz), 3.46 (1H, d,
J = 11.1 Hz), 6.73 (2H, d, J = 8.2 Hz), 6.74 (2H, d, J = 8.2 Hz), 7.14
(2H, d, J = 8.2 Hz), 7.16 (2H, d, J = 8.2 Hz), 8.07 (1H, s), 8.09 (1H,
s); 13C NMR (acetone-d6) d 22.7, 30.5, 34.3, 39.8, 40.0, 57.5,
116.0, 116.1, 125.2, 129.6, 135.8, 136.2, 156.5, 156.5; HRMS Calcd
C22H26O2S2 386.1376. Found 386.1377; Anal. Calcd for
C22H26O2S2ꢂ1/2H2O: C, 66.80; H, 6.75. Found: C, 66.88; H, 6.91.
4.2.22. Spiro[5,5]undecane-2-carboxaldehyde (28)
To a solution of DMF (0.800 mL, 10.4 mmol) in CH2Cl2 (9 mL)
was added POCl3 (0.860 mL, 8.99 mmol) at 0 °C under argon gas.
The mixture was stirred at room temperature for 1 h, then a solu-
tion of 27 (1.10 g, 6.63 mmol) in CH2Cl2 (9 mL) was added at 0 °C.
Stirring was continued at room temperature for 20 h, then satu-
rated aqueous NaHCO3 was added, and the whole was extracted
with AcOEt. The organic solution was washed with brine, dried
over Na2SO4, and concentrated. Purification by silica gel column
chromatography (eluent: n-hexane/AcOEt, 60:1) gave the formy-
late product (67%). 1H NMR (CDCl3) d 1.23–1.45 (10H, m), 1.60
(2H, t, J = 6.6 Hz), 2.14 (2H, t, J = 2.2 Hz), 2.57 (2H, tt, J = 6.6,
2.2 Hz), 10.2 (1H, s); HRMS Calcd for C12H1735ClO 212.0969
for
C19H20F2O2 318.1432. Found 318.1429; Anal. Calcd for
C19H20F2O2: C, 71.68; H, 6.33. Found C, 71.71; H, 6.22.
4.2.20. 4,40-[(3,3-Dichlorocyclohexyl)methylene]bisphenol (9)
Compound 26 was converted to the benzyl ether by the same
method as used for preparation of 4. Purification by trituration
with Et2O gave the benzyl-protected product (70%). 1H NMR
(CDCl3) d 1.29 (1H, br qua, J = 12.1 Hz), 1.61 (1H, quat, J = 12.1,
3.9 Hz), 1.82 (1H, br d, J = 13.0 Hz), 1.91–2.03 (2H, m), 2.21–2.37
(3H, m), 2.53 (1H, quat, J = 11.1, 3.1 Hz), 3.53 (1H, d, J = 10.6 Hz),
4.99 (2H, s), 5.00 (2H, s), 6.86 (2H, d, J = 8.7 Hz), 6.89 (2H, d,
J = 8.7 Hz), 7.12 (2H, d, J = 8.7 Hz), 7.16 (2H, d, J = 8.7 Hz), 7.29–
(100%),
C
12H1737ClO 214.0939 (33%). Found 212.0968 (33%),
214.0927 (11%). To a suspension of this compound (986 mg,
4.65 mmol) and 10% palladium on carbon (247 mg, 0.233 mmol)
in EtOH (18 mL) was added Et3N (1.4 mL, 10.1 mmol) at room tem-
perature. The mixture was stirred at room temperature under
hydrogen gas for 24 h, then filtered through Celite, and concen-
trated. The residue was dissolved in H2O and extracted with AcOEt.
The organic solution was washed with brine, dried over Na2SO4,
and concentrated. Purification of the residue by silica gel column
chromatography (eluent: n-hexane/AcOEt, 80:1) gave 28 (84%).
1H NMR (CDCl3) d 0.98 (1H, td, J = 13.0, 3.9 Hz), 1.01 (1H, t,
7.41 (10H, m); HRMS Calcd for
C33H32O3 476.2353. Found
476.2350. The benzyl ether was converted to the dichloro product
by the same method as that used for preparation of 4. Purification
by silica gel column chromatography (eluent: n-hexane/AcOEt,
20:1) gave the dichloro product (11%, 2 steps). 1H NMR (CDCl3)
0.81–0.91 (1H, m), 1.59–1.75 (4H, m), 2.00–2.08 (1H, m), 2.52–
2.58 (3H, m), 3.42 (1H, d, J = 10.6 Hz), 5.00 (2H, s), 5.02 (2H, s),

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T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
J = 13.0 Hz), 1.15–1.27 (2H, m), 1.34–1.51 (10H, m), 1.57–1.70 (2H,
m), 1.84 (1H, dt, J = 13.0, 1.4 Hz), 1.90 (1H, dqui, J = 13.0, 1.4 Hz),
2.38 (1H, ttd, J = 12.3, 3.9, 1.4 Hz), 9.59 (1H, d, J = 1.4 Hz); HRMS
Calcd for C12H20O 180.1515. Found 180.1508.
Purification of the residue by silica gel column chromatography
(eluent: n-hexane/Et2O, 40:1) gave 30 (58%). 1H NMR (CDCl3) d
0.18 (12H, s), 0.97 (18H, s), 2.65 (1H, s), 2.79 (1H, s), 1.36–3.02
(10H, br m), 6.74 (4H, d, J = 8.7 Hz), 7.46 (4H, d, J = 8.7 Hz); HRMS
Calcd for C27H5010B211B8O3Si2 586.4304. Found 586.4297.
4.2.23. 4,40-[(Spiro[5,5]undecan-2-yl)methylene]bisphenol (11)
To a solution of 28 (700 mg, 3.89 mmol) in phenol (2.90 g,
30.9 mmol) was added H2SO4 (0.100 mL, 1.88 mmol) at 70 °C.
The mixture was heated with stirring at the same temperature
for 19 h, and then saturated aqueous NaHCO3 was added. The
whole was extracted with AcOEt. The organic solution was washed
with brine, dried over MgSO4, and concentrated. Purification of the
residue by crystallization (CHCl3, ꢀ25 °C), trituration (CHCl3), and
silica gel column chromatography (eluent: n-hexane/AcOEt, from
10:1 to 4:1) gave 11 (32%). 11: white leaflets (Et2O); mp 228 °C;
1H NMR (CDCl3) d 0.54 (1H, t, J = 12.6 Hz), 0.71 (1H, quad,
J = 12.6, 4.3 Hz), 0.88 (1H, td, J = 13.0, 4.3 Hz), 1.04–1.16 (2H, m),
1.21–1.47 (10H, m), 1.52 (1H, br d, J = 11.6 Hz), 1.56 (1H, br d,
J = 13.0 Hz), 1.63 (1H, br d, J = 12.6 Hz), 2.16 (1H, quat, J = 11.1,
3.1 Hz), 3.26 (1H, d, J = 10.6 Hz), 4.70 (2H, s), 6.70 (2H, ddd,
J = 8.7, 2.9, 1.9 Hz), 6.73 (2H, ddd, J = 8.7, 2.9, 1.9 Hz), 7.06–7.10
(4H, m); 13C NMR (acetone-d6) d 22.2, 22.3, 33.2, 33.4, 33.9, 37.0,
37.8, 43.0, 43.6, 59.0, 115.8, 115.9, 129.5, 129.6, 137.1, 137.3,
156.2; HRMS Calcd for C24H30O2 350.2247. Found 350.2255; Anal.
Calcd for C24H30O2: C, 82.24; H, 8.63. Found C, 82.17; H, 8.73.
4.2.27. 4,40-[(1,12-Dicarba-closo-dodecaboran-1-yl)methylene]
bisphenol (14)
To a solution of 30 (169 mg, 0.288 mmol) and Et3SiH (0.160 mL,
1.01 mmol) in CH2Cl2 (3 mL) was added dropwise 47% BF3ꢂOEt2 in
Et2O (0.300 mL, 0.993 mmol) at 0 °C under argon gas. The reaction
mixture was stirred at room temperature for 23 h, and then 47%
BF3ꢂOEt2 in Et2O (0.300 mL, 0.993 mmol) was added dropwise at
0 °C. The mixture was stirred at room temperature for 24 h, then
poured into saturated aqueous NaHCO3, and extracted with AcOEt.
The organic layer was washed with brine, dried over Na2SO4, and
concentrated. Purification of the residue by silica gel column chro-
matography (eluent: n-hexane/AcOEt, 3:1) gave 14 (88%). 14:
white plates (Et2O); mp 259–260 °C; 1H NMR (CDCl3) d 1.42–3.67
(10H, br m), 2.67 (1H, s), 3.98 (1H, s), 4.86 (2H, s), 6.72 (4H, d,
J = 8.7 Hz), 7.13 (4H, d, J = 8.7 Hz). 13C NMR (CD3OD) d 60.0, 61.5,
92.0, 115.8, 131.2, 134.0, 157.4; HRMS Calcd for C15H2210B211B8-
O2:342.2624. Found: 342.2616; Anal. Calcd for C15H22B10O2: C,
52.61; H, 6.48. Found C, 52.52; H, 6.44.
4.2.28. [Bis(4-(tert-butyldimethyl)siloxyphenyl)](1,7-dicarba-
closo-dodecarboran-1-yl)methanol (31)
4.2.24. 4,40-(Cyclopentylmethylene)bis(4-methoxy)phenol (29)
To a solution of cyclohexene oxide (2.0 g, 20.4 mmol) and ani-
sole (5 mL) was added boron trifluoride ether complex (7.24 g,
50.9 mmol) at ꢀ78 °C. The mixture was stirred for 12 h at 50 °C,
then cooled, poured into water, and extracted with Et2O. The
organic layer was washed with brine, dried over Na2SO4, and con-
centrated. The residue was purified by silica gel column chro-
matography with 1:25 to 1:15 AcOEt/n-hexane to give 29 (22%)
as a yellow oil. 1H NMR (CDCl3) d 1.05–1.20 (m, 2H), 1.45–1.70
(m, 6H), 2.59 (m, 1H), 3.47 (d, J = 11.2 Hz, 1H), 3.73 (s, 6H), 6.78
(d, J = 8.7 Hz, 4H), 7.17 (d, J = 8.6 Hz, 4H); HRMS Calcd for
Compound 31 was prepared from 1,7-dicarba-closo-dodecabo-
rane by the same method as that used for preparation of 30.
Purification by silica gel column chromatography (eluent:
n-hexane/Et2O, 60:1) gave a mixture of 31 and bis(4-(tert-butyldi-
methyl)siloxyphenyl)ketone (98:2, 1.96 g). 1H NMR (CDCl3) d 0.19
(12H, s), 0.97 (18H, s), 2.76 (1H, br s), 2.85 (1H, s), 1.62–2.85 (10H,
br m), 6.78 (4H, ddd, J = 8.8, 3.4, 2.0 Hz), 7.57 (4H, ddd, J = 8.8, 3.4,
2.0 Hz); HRMS Calcd for C27H5010B211B8O3Si2 586.4304. Found
586.4297.
C20H24O2 296.1776. Found: 296.1757.
4.2.29. 4,40-[(1,7-Dicarba-closo-dodecaboran-1-yl)methylene]
bisphenol (15)
4.2.25. 4,40-(Cyclopentylmethylene)bisphenol (13)
Compound 15 was prepared from 31 by the same method as
that used for preparation of 14. Purification by trituration with
CHCl3 gave 15 (57%, 2 steps). 15: white lamellar solid (Et2O); mp
221–222 °C; 1H NMR (CD3OD) d 1.06–2.97 (10H, br m), 3.33 (1H,
br s), 4.35 (1H, s), 6.70 (4H, d, J = 8.7 Hz), 7.24 (4H, d, J = 8.7 Hz).
13C NMR (CD3OD) d 56.4, 58.4, 84.5, 116.0, 131.4, 134.2, 157.7;
HRMS Calcd for C15H2210B211B8O2 342.2624. Found Anal. Calcd for
To a solution of 25 (0.35 g, 1.18 mmol) in CH2Cl2 (3 mL) was
added a 1 M solution of BBr3 in CH2Cl2 (2.9 mL, 2.90 mmol) at
ꢀ78 °C. The mixture was stirred at room temperature for 1 h, then
poured onto ice, and extracted with AcOEt. The organic layer was
washed with brine, dried over Na2SO4, and concentrated. The resi-
due was purified by silica gel column chromatography with 1:3 to
1:1 AcOEt/n-hexane to give 13 (92%). Colorless needles (AcOEt–
hexane) mp 190.0–191.5 °C; 1H NMR (CDCl3) d 1.05–1.20 (m,
2H), 1.45–1.70 (m, 6H), 2.56 (m, 1H), 3.44 (d, J = 11.2 Hz, 1H),
4.61 (s, 2H), 6.71 (d, J = 8.6 Hz, 4H), 7.11 (d, J = 8.6 Hz, 4H); 1HRMS
Calcd for C18H20O2: 268.1463. Found: 268.1450: Anal. Calcd for
C15H22B10O2ꢂ1/2H2O: C, 51.26; H, 6.45. Found: C, 51.52; H, 6.51.
4.2.30. 1,10-[(1,2-Dicarba-closo-dodecaboran-1-yl)methylene]
bis(4-methoxy)benzene (32)
To a solution of 10,20-dicarba-closo-dodecarborane (72.0 mg,
0.500 mmol) in Et2O (1 mL) was added dropwise a 2.14 M solution
of n-BuLi in n-hexane (0.300 mL, 0.642 mmol) at 0 °C under Ar. The
mixture was stirred at room temperature for 30 min. To the reac-
tion mixture was added dropwise a solution of bis(4-methoxyphe-
nyl)bromomethane (200 mg, 0.649 mmol) in Et2O (1 mL) at 0 °C.
Stirring was continued at room temperature for 2 h, and then sat-
urated aqueous NH4Cl was added dropwise at 0 °C. The whole was
warmed to room temperature and extracted with AcOEt. The
organic layer was washed with brine, dried over MgSO4, and con-
centrated. Purification of the residue by silica gel column chro-
matography (eluent: n-hexane/CH2Cl2, 5:1) gave 32 (28%). 1H
NMR (CDCl3) d 1.38–2.92 (10H, br m), 3.25 (1H, br s), 3.79 (6H,
s), 4.73 (1H, s), 6.87 (4H, ddd, J = 8.8, 2.9, 2.4 Hz), 7.36 (4H, ddd,
J = 8.8, 2.9, 2.4 Hz); HRMS Calcd for C17H2610B211B8O2 370.2937.
Found 370.2941.
C18H20O2: C, 80.56; H, 7.51. Found C, 80.28; H, 7.48.
4.2.26. [Bis(4-(tert-butyldimethyl)siloxyphenyl)](1,12-dicarba-
closo-dodecarboran-1-yl)methanol (30)
To a solution of 1,12-dicarbora-closo-dodecaborane (72.0 mg,
0.500 mmol) in Et2O (1 mL) was added a 2.56 M solution of n-BuLi
in n-hexane (0.210 mL, 0.538 mmol) at 0 °C under argon gas. The
mixture was stirred at room temperature for 30 min. To the reac-
tion mixture was added dropwise a solution of 4,40-di(tert-butyl)
(dimethyl)siloxybenzophenone (243 mg, 0.548 mmol) in Et2O
(1.5 mL) at ꢀ30 °C for 30 min. Stirring was continued at the same
temperature for 30 min, and then saturated aqueous NH4Cl was
added dropwise at the same temperature. The resulting suspension
was stirred at 0 °C, and extracted with AcOEt. The organic layer
was washed with brine, dried over Na2SO4, and concentrated.

T. Kojima et al. / Bioorg. Med. Chem. 23 (2015) 6900–6911
6911
4.2.31. 4,40-[(1,2-Dicarba-closo-dodecaboran-1-yl)methylene]
bisphenol (16)
Hermes.27 The center of the active site was defined as the center
of the ligand in 1ERE, 3UU7 or 3UUC, as appropriate, and the active
site radius was set to 10.0 Å. Structural optimizations of ligands
were carried out at the B3LYP/6-31G(d,p) level using Gaussian
09, Revision C.01.28
To a solution of 33 (171 mg, 0.445 mmol) in CH2Cl2 (5 mL) was
added a 1.0 M solution of BBr3 in CH2Cl2 (0.920 mL, 0.920 mmol) at
0 °C under Ar. The mixture was stirring at room temperature for
43 h, then poured into ice water, and extracted with AcOEt. The
organic layer was washed with brine, dried over MgSO4, and con-
centrated. Purification of the residue by silica gel column chro-
matography (eluent: n-hexane/Et2O, 1:1) gave 16 (78%). 16:
white fibrous solid (Et2O); mp 237–238 °C; 1H NMR (CD3OD) d
1.35–2.89 (10H, br m), 3.84 (1H, br s), 4.66 (1H, s), 6.72 (4H, ddd,
J = 8.7, 2.9, 1.9 Hz), 7.27 (4H, ddd, J = 8.7, 2.9, 1.9 Hz); 13C NMR
(CD3OD) d 58.7, 63.4, 82.4, 116.4, 131.5, 132.1, 158.3; HRMS Calcd
for C15H2210B211B8O2 342.2624. Found 342.2621; Anal. Calcd for
Acknowledgments
This research was supported by a Grant-in-Aid for the Strategic
Research Program for Private Universities (2010–2014, 2015–
2019) and
a Grant-in-Aid for Scientific Research (C) (Nos.
26460151 and 26460034) from JSPS – Japan.
References and notes
C15H22B10O2 C, 52.61; H, 6.48. Found C, 52.52; H, 6.42.
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with serial dilutions of compounds or DMSO (control) in the pres-
ence or absence of 0.1 nM estradiol. Cells were incubated in tripli-
cate microcultures for 4 days, and the medium with compounds or
DMSO (control) in the presence or absence of 0.1 nM estradiol was
replaced once after 2 days. At the end of the incubation period,
WST-8 (10 nM) was added to the microcultures, and cell prolifera-
tion was evaluated 2–4 h later by measuring the absorbance at
450 nm as a parameter of the number of living cells in the culture.
4.4. Docking simulation
The docking procedures were similar to those described in our
previous paper.25 The computational docking trials were per-
formed with GOLD 5.2 software26 using the default settings. The
3D structures of human ER
a were retrieved from the Protein
DataBank (PDB) (PDB IDs: 1ERE, 3UU7, and 3UUC). Missing hydro-
gen atoms in the PDB structures were computationally added by