Molecular and Cellular Endocrinology 337 (2011) 43–51
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Molecular and Cellular Endocrinology
journal homepage: www.elsevier.com/locate/mce
LC, a novel estrone–rhein hybrid compound, concurrently stimulates
osteoprotegerin and inhibits receptor activator of NF-B ligand (RANKL) and
interleukin-6 production by human osteoblastic cells
, Ling Zhi Li
, Yong Liang Zhang
, Ya Qin Zhu
, Jian Wu
, Wei Jia Sun
Department of Immunology, Medical College of Chinese People’s Armed Police Forces, Tianjin, People’s Republic of China
TianJin Key Laboratory for Biomarkers of Occupational and Environmental Hazard, Tianjin, People’s Republic of China
Department of Pharmaceutical Chemistry, Medical College of Chinese People’s Armed Police Forces, Tianjin, People’s Republic of China
Received 29 October 2010
Received in revised form
31 December 2010
Accepted 25 January 2011
Estrogen analogues are promising drugs for postmenopausal osteoporosis, but because of their possible
side effects such as increased risk of cancer, estrogens which exert their estrogenic effects selectively on
bone are desired. It has been shown that rhein inhibits osteoclast formation and bone resorption activity
and has an antitumor role in several types of cancers. Having found that rhein had high afﬁnity for the
bone mineral, we synthesized estrone–rhein hybrid compounds and conﬁrmed that one of these hybrid
compounds, LC, exhibited a selective proﬁle in the bone and prevented bone loss but had no effect on
endometrium growth in ovariectomized rats. However, the mechanisms underlying its actions on human
bone cells have not been well deﬁned. Here we show that LC concurrently stimulates osteoprotegerin
(OPG) and inhibits receptor activator of nuclear factor-B ligand (RANKL) and Interleukin-6 (IL-6) pro-
duction by human osteoblastic MG-63 cells containing two estrogen receptor (ER) isotypes. Treatment
with the ER antagonist ICI 182,780 abrogates the above actions of LC on osteoblast-derived cells. Using
small interfering double-stranded RNAs (siRNA) technology, we further demonstrate that the effects of LC
on IL-6 production are mediated by both ER␣ and ER␤ but those on OPG and RANKL expression primarily
by ER␣. Furthermore, we also demonstrate that LC functions at least partially through activation of the
classic estrogen response element (ERE) pathway as well as Ras/MEK/ERK and PI3K/Akt signaling. The
effect of LC on bone is due to not only its estrogenic activity but also action of its rhein moiety. Also, this
compound shows much weaker effect on breast epithelial cell growth than that of estrone. Therefore,
using rhein for conjugating compounds is a promising method of effectively targeting estrogens to the
© 2011 Elsevier Ireland Ltd. All rights reserved.
Estrogen replacement therapy has long been an important
therapeutic modality for the prevention and treatment of post-
menopausal osteoporosis (Eastell, 1998). However, long-term
compliance with estrogen therapy is limited, and there are increas-
ing concerns regarding its safety (Eastell, 1998; Writing Group for
the Women’s Health Initiative Investigators, 2002). At present, two
approaches can be taken to create such bone-selective estrogens.
One approach is to make estrogens that act speciﬁcally on bone
Corresponding author at: Department of Pharmaceutical Chemistry, Medical
College of Chinese People’s Armed Police Forces, Hedong District Chenlin Road No.
211, Tianjin 300162, People’s Republic of China. Tel.: +86 22 60578184.
Corresponding author at: Department of Immunology, Medical College of Chi-
nese People’s Armed Police Forces, Hedong District Chenlin Road No. 211, Tianjin
300162, People’s Republic of China. Tel.: +86 22 60578099.
E-mail addresses: email@example.com (Y. Wang), firstname.lastname@example.org (L.Z. Li).
irrespective of their distribution, with a very weak effect on repro-
ductive tissues (Gennari et al., 2007; Kulak Júnior et al., 2010).
Examples are the so-called selective estrogen-receptor modulators
(SERMs) such as tamoxifen, raloxifene, bazedoxifene, lasofoxifene
and ospemifene (Gennari et al., 2007; Kulak Júnior et al., 2010).
These compounds function as estrogen antagonists in reproduc-
tive organs, whereas they act as estrogen agonists on bone and
lipid metabolism. The second approach is to make estrogens that
distribute speciﬁcally in bone. Examples of these are hybrids
made with tetracycline (Orme and Labroo, 1994; Neale et al.,
2009), bisphosphonate (Bauss et al., 1996; Tsushima et al., 2000),
small heterocycles (Willson et al., 1996), and acidic oligopeptide
(Yokogawa et al., 2001; Yokogawa, 2006).
To obtain a highly bone-selective estrogen that distributes in the
bone effectively, we have synthesized estrone–rhein hybrid com-
pounds (Li et al., 2004). Rhein was used because it is known to have
high afﬁnity for hydroxyapatite in the bone (Cui et al., 2008; Zhang
et al., 2008) and was suggested to function as a good carrier of estro-
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