Discovery of novel osthole derivatives as potential anti-breast cancer treatment

Article abstract of DOI:10.1016/j.bmcl.2010.10.027

Osthole, an ingredient of Traditional Chinese Medicine (TCM) from natural product Cnidium monnieri (L.) Cusson, was used as a lead compound for structural modification. A series of osthole derivatives bearing aryl substituents at 3-position of coumarin, h

Full text of DOI:10.1016/j.bmcl.2010.10.027

Bioorganic & Medicinal Chemistry Letters 20 (2010) 7426–7428
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of novel osthole derivatives as potential anti-breast cancer treatment
⇑
Lisha You , Rui An, Xinhong Wang, Yimin Li
School of Traditional Chinese Pharmacy, Shanghai University of Traditional Chinese Medicine, 1200 Cai Lun Road, Zhangjiang Hi-Tech Park, Shanghai 201203, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 27 July 2010
Revised 20 September 2010
Accepted 6 October 2010
Available online 2 November 2010
Osthole, an ingredient of Traditional Chinese Medicine (TCM) from natural product Cnidium monnieri (L.)
Cusson, was used as a lead compound for structural modification. A series of osthole derivatives bearing
aryl substituents at 3-position of coumarin, has been prepared and evaluated for their growth inhibitory
activity against human breast cancer cell lines MCF-7 and MDA-MB-231. Interestingly, some derivatives
exhibited good inhibition, among them compound 8e was found to be the most potent compound with
IC50 values of 0.24
more than 100-folds compared with its parent compound osthole.
lM, 0.31 lM against MCF-7 and MDA-MB-231, respectively, which was improved
Keywords:
Osthole
Antitumor
Ó 2010 Elsevier Ltd. All rights reserved.
Breast cancer
Natural product
Structure modification
The coumarin(benzopyran-2-one, or chromen-2-one) ring
system, present in natural products (such as the anticoagulant
Warfarin, 1), that display interesting pharmacological properties,1,2
has intrigued chemists and medicinal chemists for decades to
explore the natural coumarins or synthetic analogs for their appli-
cability as drugs (Fig. 1). Many interesting molecules based on the
coumarin ring system have been synthesized utilizing innovative
synthetic techniques. Some new derivatives bearing coumarin ring
including the furanocoumarins (e.g., Imperatorin, 2), pyra-
nocoumarins (e.g., Seselin, 3), and coumarin sulfamates (Cou-
mates) (4), have been found to be useful in photochemotherapy,
antitumor and anti-HIV therapy and others.3,4 Among the diverse
biological activities of coumarins, the most intriguing is the nota-
ble effect of, some of the coumarins against breast cancer, some
coumarins and their active metabolite 7-hydroxycoumarin analogs
have shown sulfatase and aromatase inhibitory activities.5,6
Coumarin based selective estrogen receptor modulators
(SERMs) and coumarin estrogen conjugates have also been
described as potential anti-breast cancer agents according some
recently publications.7 Currently, tamoxifen is most commonly
used adjuvant drug for estrogen receptor (ER)-positive breast can-
cer,8 which competes with estrogen and downregulates estrogenic
actions in breast cancer, however, it is less effective in ER-negative
breast cancer, and its safety is also controversial.9–11 Therefore,
there is a strong impetus to identify new anti-breast cancer agents
with improved activity and reduced side effects.
Osthole, 7-methoxy-8-(3-methyl-2-butenyl) coumarin (5)
(Fig. 1), is a coumarin derivative clinically ingested as an important
component of medicinal plants and herbs12,13 in Tradition Chinese
Medicine (TCM), and it exhibits many pharmacological and biolog-
ical activities,14,15 The original study of osthole could be dated back
to more than 100 years ago. It was first discovered from Peuceda-
num ostruthium. Now osthole is being found to be widely distrib-
uted in the plant kingdom. Cnidium monnieri (L.) Cusson, a kind of
plant containing high percentage of osthole, which has been used
in China since several hundred years before as an herbal medicine
to treat male sexual dysfunction. It has been proved that osthole
has some important therapeutic function and safe profile com-
pared with other natural product, which makes it a very promising
lead compound in drug discovery area. Many reviews have sum-
marized its diverse biological activities and therapeutic applica-
tion. In this paper, we utilized osthole as lead compound to
modify in order to improve its anti-cancer activity and drug-like-
ness properties.
The studies on growth-inhibitory cytostatic activity in human
cancer cell line: MCF-7 breast carcinoma cells revealed that osthole
demonstrated some estrogenic activity by preventing the synthesis
and action of estrogens (ER antagonists)16,17 which indicated that
osthole has the potential to become a breast cancer treatment re-
agents. However, osthole exhibited comparatively weak activity,
low water solubility and limited permeability, and these properties
may lower its absorption upon oral administration and bioavail-
ability, which need improve for better drug-likeness.3,12
According to literature analysis18, 3- and 4-position of osthole
might play an important role in proliferation activity, therefore,
we tried to introduce some substituents at these positions to im-
prove its anti-cancer activity. Bromination and Vilsmeier–Haack
⇑
Corresponding author. Tel.: +86 021 51322183; fax: +86 021 2623938.
E-mail address: youlisha@shutcm.com (L. You).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.10.027

L. You et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7426–7428
7427
CH3
R1
CH3
O
O
R2
O
O
H3C
O
O
O
O
O
O
O
O
O
CH3
CH3
Osthole (5)
O
NH2SO2O
CH3
O
CH3
1
2
3
4
Figure 1. Structures of coumarin Warfarin (1), Imperatorin (2), Seselin (3), and Coumates (4).
Br
b
a
O
O
O
O
O
O
O
O
O
6
7
R1
R1
R1
c
R2
e
d
HO
O
O
O
O
O
O
O
O
8
9
10
Scheme 1. Reagents and conditions: (a) PtO2, H2, 24 h, 80%; (b) NBS, NaOAc, CH3CN, MW, 2 h, 60%; (c) Pd(PPh3)4, phenylboronic acid, K3PO4, dioxane, 60–90%; (d) BBr3, DCM,
À78 °C, 40–70%; (e) bromide, DMF, NaH, 60–85%.
reaction were attempted to introduce bromo or formaldehyde at 3-
or 4-position of osthole (Scheme 1). However due to the existing of
double bond of isopentenyl, these efforts were failed finally; nei-
ther 3-position nor 4-position substituted products were obtained
successfully. It was believed that the modification at 3- or 4-posi-
tion could be realized after the hydrogenation of isopentenyl. It
was proved that platinum oxide could catalyze the hydrogenation
to afford reduced osthole derivative 6 with a good yield (more than
80%), the hydrogenated product 6 could carry out bromination un-
der microwave irradiations selectively at 3-position to yield the
important intermediate 7, which could be employed to couple with
various aryl boric acid and prepare a series of novel osthole deriv-
atives 8. After demethylation and nucleophilic substitution, a new
class of osthole derivatives 9 and 10 were obtained too.
The anti-cancer activity tests in vitro of the above osthole ana-
logs were carried on human breast cancer MCF-7 and MDA-MB-
231 cell lines. The structure and activity relationship studies
showed the methoxy at 7-position is critical for maintaining the
antitumor activity, and the modification at 3-position and the
hydrogenation on the double bond of isoamylene could obviously
improve its antitumor activity (Fig. 2).
Preliminary experiments illustrated that the series of novel ost-
hole derivatives with modification at 3-position exhibited im-
proved inhibitory activity against breast cancer cell (Table 1). The
R2
a: R1= CH3 , R2= CH3O, R3=CH3O
b: R1=CH3, R2=CF3, R3=H
f: R1=CH3, R2= F, R3=F
g: R1=CH3, R2=CH3, R3=H
R3
R1
c: R1=H, R2=CF3, R3=H
d: R1=CH3, R2= F, R3=H
e: R1=CH3, R2= OCF3, R3=H
h: R1=CH3, R2= OH, R3=H
i: R1=CH3, R2= Cl, R3=H
j: R1=H, R2= OCF3, R3=H
O
O
O
8
F
F
F
N
O
O
O
O
O
O
HO
O
O
O
O
O
R1
O
O
O
O
O
O
11
12
R1=CH3
R1=H
13
14
15
Figure 2. The novel derivatives of osthole.

7428
L. You et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7426–7428
Table 1
replacement at 3-position of coumarin core, of which the in vitro
anti-breast cancer activities against MCF-7 and MDA-MB-231 cell
lines were investigated, and preliminary SAR around this scaffold
was established. The results indicated that 7-methoxy and 3-aryl
ring are critical for maintaining cytotoxicity potency. Compound
8e were the most potent analogs (IC50 values of 0.24 and
Cell growth inhibitory activity of osthole derivatives 8–15 in various cell lines with
MTT assay
Compound no.
MCF-7 Cell growth
MDA-MB-231
IC50 (lM)
inhibition IC50
(l
M)b
Tamoxifen citratea
Osthole (1)
6
8ª
8b
8c
8d
8e
8f
8g
8h
8i
10.3
25.8
47.0
51.6
24.5
42.9
52.6
0.24
27.3
50.2
3.23
1.27
5.70
NA
12.8
30.2
NAc
NA
38.1
NA
1.27 lM, respectively, against the MCF-7 and MDA-MB-231 cell
lines), and showed 100 higher antitumor activity compared with
osthol, and lacked cytotoxicity to normal HERK-293 cells. Mecha-
nism of action study is ongoing and the latest progress will be re-
ported in due course. In summary, 8e and 8i are the two novel and
promising lead compounds suitable for further development to-
ward a potential clinical trials candidate.
NA
0.31
33.6
NA
4.15
5.23
7.42
NA
NA
NA
NA
NA
Acknowledgments
8j
11
12
13
14
15
Financial assistance from Project of Shanghai Educational Com-
mission (08CZ08). ‘Xinlin’ scholars and outstanding team training
plan of Shanghai University of Traditional Chinese Medicine.
NA
NA
NA
45.9
a
b
c
Supplementary data
Positive control.
Mean IC50
(
lM), from three or more independent tests.
NA means not active and having IC50 >50
lM.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.10.027.
most potent analog investigated in this study appears to be triflu-
ormethyl phenyl derivative 8e, of which the cytotoxic activity
against MCF-7 and MDA-MB-231 cell lines was improved more
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than 100-fold to 0.24
its parent compound osthole.
For the analog 8i bearing p-chloro phenyl substituted at 3-posi-
tion, the antitumor activity was improved significantly (1.27 M vs
lM, 0.31 lM, respectively, compared with
l
25.8 against MCF-7), about 20-fold higher than its parent com-
pound. The inhibitory activity of compound 8h containing p-phe-
nol also had eight times increase (IC50 value is 3.23 lM).
Compounds 8a, 8b, 8d and 8f, which incorporate dimethoxyl phe-
nyl, trifluormethyl, fluoro, and difluoro phenyl at 3-position of the
coumarin ring, respectively, showed slightly increase (two, three-
fold) or comparable cytotoxic activity with osthole. In contrast,
methyl phenyl derivative 8g was found to weakly inhibit human
cancer cell growth in vitro with IC50 values around 50 lM.
The activity of demethylated compound 8j declined more than
20-folds, 8c is a 7-position analog prepared from 8b, exhibited re-
duced activity (IC50 = 42.9 lM), it revealed that the 7-methyloxy
group is a very sensitive substituent for maintaining the antitumor
activity. Same impacts were also observed in compound 11, which
is a hydrogenated and demethylated osthole derivative, of which
the activity was totally lost. The replacement of the phenyl ring
12 by a 3-pyridyl moiety also leads to the losing of cellular activity.
Compounds 13 and 14 did not show even lowest activity.
Since 8e and 8i showed better cell growth inhibition for breast
cancer cell lines than tamoxifen citrate and osthole, their cytotox-
icity was also tested in normal cells, Human Embryonic Kidney
(HEK)-293. Results show that 8e and 8i had no cell growth inhib-
itory effect in HEK-293 cells. The further MOA and in vivo study
for 8e is under going now.
In conclusion, we designed and prepared a series of novel ost-
hole derivatives by structural modification and bio-isosteric