Synthesis and estrogen receptor binding affinities of 7-hydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-ones containing a basic side chain

Article abstract of DOI:10.1016/S0960-894X(03)00132-X

Two isoflavones containing a sulfur or oxygen hinge with an amine-bearing side chain have been designed and synthesized as potential selective estrogen receptor modulators. The target compounds exhibited low affinities for estrogen receptors (ERs), and binding affinity data indicate that oxygen hinge is more favorable than sulfur for binding. These compounds also displayed selectivity for ERα over ERβ.

Full text of DOI:10.1016/S0960-894X(03)00132-X

Bioorganic & Medicinal Chemistry Letters 13 (2003) 1475–1478
Synthesis and Estrogen Receptor Binding Affinities of 7-Hydroxy-
3-(4-hydroxyphenyl)-4H-1-benzopyran-4-ones Containing a
Basic Side Chain
Young-Woo Kim,a James A. Mobleyb and Robert W. Brueggemeiera,*
aDivision of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, 500 West 12th Avenue,
Columbus, OH 43210, USA
bDivision of Urology, Department of Surgery, University of Massachusetts Medical School, 55 Lake Avenue North,
Worcester, MA01655, USA
Received 4 October 2002; revised 31 January 2003; accepted 3 February 2003
Abstract—Two isoflavones containing a sulfur or oxygen hinge with an amine-bearing side chain have been designed and synthe-
sized as potential selective estrogen receptor modulators. The target compounds exhibited low affinities for estrogen receptors
(ERs), and binding affinity data indicate that oxygen hinge is more favorable than sulfur for binding. These compounds also dis-
played selectivity for ERa over ERb.
# 2003 Elsevier Science Ltd. All rights reserved.
Over the past few years, our group has been interested
in the 4H-1-benzopyran-4-one ring system (Fig. 1) as a
core of potential therapeutic agents for the treatment of
hormone-dependent breast cancer.1 This ring system
can be found in a number of natural products termed
flavonoids. Compounds in this class have demonstrated
numerous biological activities such as antiviral, anti-
inflamatory, antiallergic, antimutagenic and anti-
carcinogenic activities.2 In breast cancer cells, in
particular, numerous flavonoids have shown interesting
pharmacological activities including binding affinities for
estrogen receptors (ERs),3 antiproliferative activities,4 and
inhibitory activities against aromatase enzyme.5
various breast cancer cell lines including MCF-7, T47D,
MD-MBA-231, and SKBR3 by several mechanisms of
action.6,8
Based on these findings, we became interested in identi-
fying a new series of selective estrogen receptor mod-
ulators (SERMs) constructed on an isoflavone scaffold.
SERMs are a class of nonsteroidal ER ligands with
mixed estrogen agonistic/antagonistic activity and are
represented by raloxifene (Fig. 2).9,10 The amine-bearing
side chain of raloxifene is reported to play a key role for
SERM activity by preventing the proper positioning of
11
helix 12for agonistic activity.
For this reason, most
published SERMs contain a similar amine-bearing side
chain.10
One isoflavonoid, genistein (Fig. 1), has been extensively
studied. Genistein is a weak phytoestrogen and has
structural characteristics in common with the non-
steroidal estrogen pharmacophore, that is, two phenolic
groups separated by approximately 11–12A planar
core.6 It is therefore not surprising that it shows rea-
sonable binding affinities for ERs and, more impor-
tantly, is able to discriminate between two ER subtypes
(ERa and ERb).7 Genistein also inhibits proliferation of
We have initially designed several isoflavone analogues
containing the basic side chain of raloxifene (Fig. 2).
However, compound A has already been synthesized by
another research group.12 Therefore, we have focused
on compounds 1 that contain a sulfur or oxygen as an
isostere of the carbonyl group. We envisioned these
heteroatoms could serve as a hinge to direct the basic
side chain to the proper region in the binding pocket of
the ER for the SERM profile.11 Herein we wish to
describe the synthesis of compounds 1 and their binding
affinities for ERs as preliminary biological data.
*Corresponding author. Tel.:+1-614-292-5231; fax:+1-614-292-2435;
e-mail: brueggemeier.1@osu.edu
0960-894X/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0960-894X(03)00132-X

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Y.-W. Kim et al. / Bioorg. Med. Chem. Lett. 13 (2003) 1475–1478
tively protected with benzyl group under Mitsunobu
reaction conditions to give monobenzyl ether 3. To
construct the 4H-1-benzopyran-4-one scaffold, we pre-
viously developed a convenient one-pot synthesis using
a phase transfer catalyst.1c Since this method is not
readily amenable to solid-phase synthesis, we modified
the reaction conditions in which sodium hydride was
utilized as a base. The resulting 2-(methylthio)isoflavone
4 was oxidized by mCPBA to 5, whose methylsulfonyl
group was then successfully displaced with sodium salt
of 4-mercaptophenol or hydroquinone to afford 2-(4-
hydroxythiophenoxy)-isoflavone 6a and 2-(4-hydroxy-
phenoxy)isoflavone 6b, respectively. The hydroxyl
group of each compound 6 was alkylated with 1-(2-
chloroethyl)piperidine to give compounds 7. Initial
attempts to generate the target molecules 1a and 1b
using typical reaction conditions for demethylation,
Figure 1. Structures of genistein and its core ring.
Figure 2. Structures of raloxifene and target molecules.
Many synthetic methods have been reported for the
synthesis of flavonoids.13 These previously reported
methods are not ideally suited for the synthesis of our
target isoflavones. Our initial goal was therefore to
develop an efficient synthetic route that avoids the harsh
conditions previously employed in flavonoid chemistry.
One of the authors has developed a convenient method
for the synthesis of 6-substituted 1,5-dialkyluracils, in
which a-oxoketene dithioacetals were employed as key
intermediates for the construction of pyrimidine ring.14
We anticipated this strategy would be suitable for the
synthesis of our target molecules. One of the advantages
of this synthetic route is that, after construction of the
ring system, the resulting methylthio substituent can be
converted into the methylsulfonyl group, which serves
as a good leaving group for the introduction of other
substituents. Thus, we investigated the feasibility of this
strategy for the synthesis of our target molecules. The
retrosynthesis is outlined in Scheme 1.
As shown in Scheme 2, our synthesis starts with deoxy-
benzoin 2, which can be prepared by the published
procedure from resorcinol and 4-methoxyphenylacetic
acid.15 Since we wished to eventually apply this method
to a solid-phase synthesis, we decided to use benzyl
group as a surrogate for hydroxymethyl polystyrene
resin. The 4-hydroxyl group of 2 was therefore selec-
Scheme 2. Synthesis. Reagents and conditions: (a) BnOH, Ph3P,
DIAD, THF, 0 ꢀC, 0.5 h; (b) NaH, CS2, DMF, 0 ꢀC, 5 min; MeI,
0 ꢀC!rt, 1 h; (c) mCPBA, CH2Cl2, reflux, 1 h; (d) 4-mercaptophenol
(for 6a) or hydroquinone (for 6b), NaH, DMF, 0 ꢀC, 1 h; (e) 1-(2-
chloroethyl)piperidine monohydrochloride, Cs2CO3, DMF, rt, over-
night; (f) BBr3, CH2Cl2, rt, overnight; (g) HCl, EtOAc.
Scheme 1. Retrosynthesis.

Y.-W. Kim et al. / Bioorg. Med. Chem. Lett. 13 (2003) 1475–1478
1477
such as boron tribromide in dichloromethane, were
unsuccessful; the competitive cleavage of piperidinyl
ethoxy group led to compounds 8 as major products,
which can be obtained from compounds 6 using the
same reaction conditions. This partial cleavage of the
basic side chain has been previously noted by Katzen-
ellenbogen and co-workers with their pyrazole derivati-
ves.10b They have successfully overcome this problem
using the milder AlCl3-EtSH reagent. However, the
application of this reagent to our compounds which
contain a Michael acceptor was considered to be inap-
propriate because of the intrinsic nucleophilicity of
ethanethiol.16 The basic amine was considered to be
responsible for the susceptibility of the side chain, pre-
sumably by coordinating with the Lewis acid and
thereby allowing access of the Lewis acid to the suscep-
tible ethoxy group. Based on this explanation, we were
able to avoid the undesired cleavage of the side chain by
using hydrochloride salts of compounds 7 as substrates.
The positively charged ammonium species might pre-
vent the access of Lewis acid to the ethoxy group in the
side chain. Thus the desired targets 1a and 1b were syn-
thesized from deoxybenzoin 2 in seven steps in excellent
overall yields (60 and 62%, respectively).17
pound A has been reported to bind well to estrogen
receptors with Ki value of 3.9ꢁ0.3 nM.12 While further
investigation is needed, the result indicated that both
sulfur and oxygen atoms in this series may significantly
alter the orientation of the basic side chain, which may
not be well tolerated by the receptors. To our surprise,
both compound 8a and 8b, which lack the bulky side
chain, bind to ERa with even lower affinities. This is
interesting because most triphenolic compounds tend to
bind reasonably well to the estrogen receptors.20 It is
plausible that, in this series of compounds, the basic side
chain may provide an additional interaction with ER in the
binding pocket, which can compensate for its bulkiness.
In summary, we have designed new isoflavone deriva-
tives as potential SERMs and have developed an effi-
cient synthetic route for the preparation of these
molecules. Although these isoflavones display common
structural features that are found in many of published
SERMs, the ER binding data indicated they are poor
ligands for estrogen receptors. Further work is currently
underway to understand the consequences of this study.
Our medicinal chemistry efforts are now focused on
manipulating functional groups around the isoflavone
core to identify new lead compounds for the treatment
of hormone-dependent breast cancer.
Since SERMs display their activities through the estro-
gen receptors, we evaluated binding affinities of com-
pounds 1 for human ERa and ERb. We also included
the triphenolic compounds 8 in the assay. The binding
affinities were determined by fluorescence polarization
using a modified protocol of the previously reported
procedure.18 All the compounds showed no significant
binding to ERb below 5 mM (data not shown). Binding
results for ERa are listed in Table 1 as relative binding
affinity.
Acknowledgements
This work was financially supported in part by a
USAMRMC Breast Cancer Program grant DMAD 17-
00-1-0388.
References and Notes
As previously mentioned, no significant binding to ERb
was observed for both 1 and 8 while they showed bind-
ing affinities for ERa. This is opposite to our expecta-
tion because genistein is known to bind to ERb with
higher affinity than to the other subtype. Albeit with a
preference for ERa, binding affinities of 1 for the sub-
type are still low. However, oxygen linkage seems to be
a more favorable hinge than sulfur based on the binding
affinity of 1b, which is almost four times greater than
that of 1a. It has been reported that, in raloxifene ana-
logues, oxygen linker provides an enhanced affinity for
ER relative to carbonyl group of raloxifene.19 It is
interesting to note that their structurally related com-
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Table 1. Binding affinities for ERa of compounds 1 and 8
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EC50, mM
Log EC50 (ꢁSE)a
RBAb
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4-OHTc
0.0157
0.77
0.20
1.86
1.74
ꢂ7.81 (ꢁ0.12)
ꢂ6.11 (ꢁ0.07)
ꢂ6.70 (ꢁ0.11)
ꢂ5.73 (ꢁ0.55)
ꢂ5.76 (ꢁ0.13)
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1.95
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1
IR (KBr) 1610, 1558, 1500, 1385, 1270, 1243, 1194 cmꢂ1; H
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2.60 (bt, 2H), 2.46–2.2n8 (m, 4H), 1.55–1.28 (m, 6H); 13C NMR
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115.44, 107.70, 102.60, 66.86, 58.22, 55.26, 26.41, 24.77; HRMS
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