Synthesis of a dendritic estrogen cluster: A potential tool for studies of nuclear versus extranuclear pathways of estrogen actions

Article abstract of DOI:10.1016/j.cclet.2012.11.009

A novel estrogen dendrimer has been synthesized through a combination of divergent and convergent approaches in 9 practical steps and in good yields. It was characterized and confirmed by elemental analysis, FT-IR, MS, ¹H NMR, ¹³C NMR. The dendrimer contains 16 estrone units and is potentially a useful tool for the studies of estrogen actions.

Full text of DOI:10.1016/j.cclet.2012.11.009

Available online at www.sciencedirect.com
Chinese Chemical Letters 23 (2012) 1319–1322
www.elsevier.com/locate/cclet
Synthesis of a dendritic estrogen cluster: A potential tool for studies
of nuclear versus extranuclear pathways of estrogen actions
Jian Chen a, Hu Zheng b, Yan Song a, Yu Feng Liang b, Qing Rong Qi b,
*
a Functional Food (Drug) Laboratory, Institute of Agro-Product Processing Science and Technology, Sichuan Academy of
Agricultural Science, Chengdu 610066, China
b Key Laboratory of Drug-Targeting and Drug-Delivery Systems of the Ministry of Education, Department of Medicinal Chemistry,
West China School of Pharmacy, Sichuan University, Chengdu 610041, China
Received 14 July 2012
Available online 1 December 2012
Abstract
A novel estrogen dendrimer has been synthesized through a combination of divergent and convergent approaches in 9 practical
steps and in good yields. It was characterized and confirmed by elemental analysis, FT-IR, MS, 1H NMR, 13C NMR. The dendrimer
contains 16 estrone units and is potentially a useful tool for the studies of estrogen actions.
# 2012 Qing Rong Qi. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: Estrogen dendrimer; Synthesis; Nuclear and extranuclear; Receptor
Estrogen plays important roles in human physiology and pathology. It acts through two pathways. One is via
classical nuclear estrogen receptors (ERs), namely ERa and ERb, altering target gene transcription [1]. The other is an
extranuclear pathway via the modulation of signaling through secondary messengers, more commonly associated with
growth factor and G protein-coupled receptors (GPR30) [2,3]. The extranuclear pathway has become an active topic as
a novel therapeutic target in estrogen-related diseases [4–6].
To carry out selective studies of nuclear and extranuclear pathways of estrogen actions, one important approach is
to create membrane-impermeable estrogen ligands, based on the assumption that they only stimulate receptors with
binding sites on the external face of the plasma membrane, without stimulation of nuclear ERs [4].
In this work we report a novel design and synthesis of a dendritic estrogen cluster 11 (DEC, Fig. 1). The DEC
compound might be a potential tool for the studies of estrogen activities that follow extranuclear pathways. Compared
with previously reported membrane impermeable estrogen macromolecular conjugates [7–10], the new dendritic
estrogen cluster overcomes many of their reported defects. For example, DEC contains exactly 16 estrone molecules
on the surface of the dendrimer, not ‘‘approximately’’. It is a structurally well-defined single unique molecule with a
molecular weight 6744.4 (C401H516O88), so there is no ‘‘free’’ estrogen in the cluster. The second generation
dendrimer is composed of neutral aliphatic extension chains with appropriate length. The chains have chemically
stable ether and ester bonds, not non-covalent or chemical unstable oxime linkages. There is no amino group in the
* Corresponding author.
E-mail address: qiqingronghh@yahoo.com.cn (Q.R. Qi).
1001-8417/$ – see front matter # 2012 Qing Rong Qi. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
http://dx.doi.org/10.1016/j.cclet.2012.11.009

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J. Chen et al. / Chinese Chemical Letters 23 (2012) 1319–1322
Fig. 1. Chemical structure of the dendrimer 11.
dendrimer, so it bears no charge at physiological pH values, so there is no disturbance of cell membrane permeability.
Furthermore, the aliphatic chain will not introduce physiological side effects, other than estrogen action, before or
after the DEC degradation.
The synthesis of DEC was achieved through a strategy combining divergent and convergent approaches, as shown
in Schemes 1–3.
The first generation eight directional multi-hydroxyl aliphatic dendrimer 3 was synthesized using a divergent
method, as shown in Scheme 1. The starting compound 1 [11] was reacted with 1,3-O-benzylidene glycerol [12] to
produce 2. 1,3-O-Benzylidene glycerol has cis- and trans-forms. The hydroxyl group in the trans-form has higher
reactivity because of less steric hindrance, so we used trans-1,3-O-benzylidene glycerol to react with 1 to obtain 2 with
72% yield. The acetal groups in 2 were very sensitive to acid conditions so it had to be handled carefully in the
preparation and testing process to avoid a complicated decomposition mixture. On the other hand, 2 was easily
hydrolyzed to expose eight hydroxyl groups in the presence of p-TsOH in methanol, resulting in the oily compound 3
in 90% yield.
The important branched side chain 4, which possesses two estrone molecules, was synthesized from the extension
chain diethylene glycol derivative 5 [11], as shown in Scheme 2. In a similar way to the preparation method of
OH
OH
HO
HO
Ph
O
O
Ph
TsO
TsO
O
O
O
O
O
O
OTs
O
O
a
b
O
O
O
O
O
O
O
O
O
O
O
O
OTs
O
O
O
O
OH
OH
HO
Ph
Ph
O
O
HO
1
2
3
Scheme 1. (a) trans-1,3-O-Benzylidene glycerol (4.4 equiv.)/THF/NaH/reflux 48 h, 72% and (b) p-TsOH/MeOH/rt, 6 h, 90%.

J. Chen et al. / Chinese Chemical Letters 23 (2012) 1319–1322
1321
a
b
OH
OH
O
PhCH O
2
O
PhCH O
2
O
PhCH O
OTs
2
O
Ph
O
O
O
7
6
5
c
O
e
d
OTs
OTs
OTs
PhCH O
2
O
HO
O
O
O
O
O
O
HO
O
O
OTs
9
8
O
10
f
O
O
O
O
O
O
O
HO
O
4
Scheme 2. (a, b) cis,trans-1,3-O-Benzylidene glycerol (1 equiv.)/THF/NaH/reflux 3 h; p-TsOH/MeOH/rt, 4 h, 70%; (c) p-TsCl (2.15 equiv.)/THF/
5 mol/L NaOH, 5 8C, 5 h/rt 1 h/99.5%; (d) 10% Pd/C, 1 atm H2/acetone–EtOH/rt, 10 h, 100%; (e) estrone (2.01 equiv.)/THF/KOH/reflux 15 h, 65%;
and (f) succinic anhydride (5 equiv.)/pyridine/reflux 12 h, 78%.
HO
O
OH
OH
HO
O
O
a
O
O
O
O
O
DEC
O
O
O
O
HO
O
O
O
O
O
OH
HO
HO
OH
3
4
11
Scheme 3. (a) 3 and 4 (10 equiv.) DMAP/CH2Cl2/rt 24 h, 72%.
compound 2, trans-1,3-O-benzylidene glycerol was first used to react with 5 to synthesize 6. In acidic conditions, the
acetal of 6 was decomposed leading to the exposure of two hydroxyl groups to form the important compound 7.
Because trans-1,3-O-benzylidene glycerol is more expensive than the cis,trans-1,3-O-benzylidene glycerol mixture,
we later successfully used the mixture to react with 5 to efficiently get 7 in total 70% yield as described in supporting
information. Compound 7 was tosylated to produce the oily diester 8 in 99.5% yield. The diester 8 was then
hydrogenized to afford the oily diester 9 at almost 100% yield. The diester 9 reacted with estrone to give the di-
estrogen compound 10 as a semi-solid. Although there is a hydroxyl group in compound 9, with our successful
selective reaction conditions, namely refluxing in THF with a KOH catalyst, only the phenolic hydroxyl group of
estrone was active enough to react with the tosyl group of 9 to afford 10 (mp 108–110 8C). Thus, the side chain 4 was
finally obtained through the reaction of 10 with succinic anhydride in pyridine to form a waxy solid in 78% yield.
The target compound DEC 11 was successfully synthesized through the convergent reaction of 3 and 4, as shown in
Scheme 3. The most difficult challenges in this final assembly were obtaining a suitable solvent and making all the 8-
hydroxyl groups in 3 react completely to avoid any ‘‘residue’’ hydroxyl groups. The difficulties came from the
solubility of 3 and 4, because 3 was more soluble in polar solvents and 4 was soluble in non-polar solvents. According
to our research, CH2Cl2 provided the perfect solvent for this reaction, because 3 was very slightly soluble in CH2Cl2
and gradually disappeared from the reaction mixture as the reaction proceeded. Compound 11 was obtained as a pure
white solid (mp 65–67 8C) at 72% yield. The overall yields of the 9 steps required to synthesize DEC 11 is 16.5%.
Compounds 2, 3, 4, 6, 7, 8, 9, 10 and 11 have not been reported previously. Their preparation processes and structural
characterizations were described in supporting information.

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J. Chen et al. / Chinese Chemical Letters 23 (2012) 1319–1322
Fig. 2. Three-dimensional model of the dendrimer 11.
The three-dimensional molecular model of the novel estrogen dendrimer 11 was simulated using ChemBio3D
software (Fig. 2). This picture shows the flexibility of DEC and the full exposure of the estrogen, suggesting its
advantages and potential use for studies of nuclear and extranuclear pathways of estrogen activity. Further studies of its
biological activities will be reported in due course.
Acknowledgments
This work was supported by the National Key Basic Research Program of China (973 Program) Foundation
(2010CB529905). We also appreciate support from the Sichuan Academy of Agricultural Science, China.
Appendix A. Supplementary data
Supplementary material related to this article found, in the online version, at http://dx.doi.org/10.1016/
j.cclet.2012.11.009.
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