An Amide Analog of (?)-Epigallocatechin Gallate Shows Preferential Cytotoxicity toward Triple-Negative Breast Cancer Cells

Full text of DOI:10.1002/bkcs.12007

Communication
DOI: 10.1002/bkcs.12007
BULLETIN OF THE
T. G. Lee et al.
KOREAN CHEMICAL SOCIETY
An Amide Analog of (−)-Epigallocatechin Gallate Shows Preferential
Cytotoxicity toward Triple-Negative Breast Cancer Cells
†,
Tae Gum Lee,†,# Yulim Kim,†,# Woong Jung,‡ Mi Kyoung Kim,†, and Youhoon Chong
*
*
†Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University,
120 Neungdong-ro, Gwangjin-gu, Seoul 05029, South Korea.
*E-mail: mkkim@konkuk.ac.kr; chongy@konkuk.ac.kr
‡Department of Emergency Medicine, Kyung Hee University Hospital at Gangdong, Seoul 134-727,
South Korea
#These authors contributed equally to this work.
Received January 28, 2020, Accepted March 6, 2020
Keywords: (−)-Epigallocatechin gallate, Amide, Cytotoxicity, Triple-negative breast cancer
MDA-MB-231 is a highly aggressive, invasive and poorly
differentiated triple-negative breast cancer (TNBC) cell line
as it lacks estrogen receptor and progesterone receptor
expression, as well as HER2 (human epidermal growth fac-
tor receptor 2) amplification.1,2 Similar to other invasive
cancers, there are only limited treatment options for TNBC
and, as a result, there is an urgent need to discover cyto-
toxic agents against the MDA-MB-231 cells.
ketone 5. Upon reductive amination with benzylamine followed
by debenzylation, was converted into 3-amino
5
epigallocatechin (6) in 64% combined yields. The reductive ami-
nation proceeded in a stereospecific manner to give only a
single stereoisomer, which was unequivocally confirmed to
have (2R, 3R) configuration by comparison of its spectro-
scopic data with those of the known compound.14 Conden-
sation of 6 with methyl 3,4,5-tribenzyloxybenzoate in the
presence of 1-ethyl-3-(3-dimethylaminopropyl)carbo-
diimide and 4-dimethylaminopyridine provided the protec-
ted EGCG-amide, which was converted into 7 after removal of
the TBS protecting group upon treatment with HF-pyridine
(65% yield). Debenzylation of 7 provided the EGCG-amide
(3) in 90% yield.
(−)-Epigallocatechin gallate [EGCG (1), Figure 1], the most
abundant tea catechin with a broad-spectrum bioactivity,3 is also
known as a cytotoxic agent against MDA-MB-231 cells.4 More
interestingly, a prodrug of EGCG [AcEGCG (2), Figure 1]
showed significantly improved antiproliferative effect against
MDA-MB-231 cells compared with EGCG,5,6 which prompted
structure–activity relationship study of EGCG esters.7–9 Glyco-
sylation10,11 as well as bioisosteric replacement of the D-ring phe-
nolic hydroxyl groups with a fluorine atom have also been
attempted.12 Nevertheless, the ester functionality bridging C- and
D-ring (bold lines in 1, Figure 1) has rarely been targeted for
structural modification of the EGCG scaffold. From a synthetic
point of view, replacement of the bridging ester with other func-
tionalities is more challenging than substitution at the peripheral
phenolic hydroxyl groups (OR, Figure 1). In addition, substitu-
tion at the epimerization-susceptible 3-position provides addi-
tional consideration.13 In this study, we devised a synthetic route
to an amide analog of EGCG [EGCG-Amide (3), Figure 1],
which showed preferential cytotoxicity toward triple-negative
breast cancer cell.
Synthesis of EGCG-amide (3) was performed by using the ste-
reospecific reductive amination as the key step (Scheme 1).
Starting from EGCG (1), global protection of the phenolic
hydroxyls with tert-butyldimethylsilyl chloride followed by
removal of the 3-gallate moiety by LiAlH4 reduction provided
TBS-protected epigallocatechin 4 in 78% yield. The substrate for
the reductive amination, 3-keto epigallocatechin (5, Scheme 1),
was prepared only under mild oxidation conditions, and Dess-
Martin oxidation of 4 provided clean conversion to the desired
Antioxidative activity15 and cytotoxicity against tumor
cells,16 the representative bioactivity of EGCG (1), were evalu-
ated for EGCG-amide (3), but no significantly different activity
was observed compared with EGCG (1) (data not shown). Thus,
EGCG-amide (3) was equipotent to EGCG (1) in 1,1-diphenyl-
2-picryl hydrazyl (DPPH) radical scavenging activity and MTT
[(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)
cell proliferation assay in various cancer cells (data not shown).
As such, the replacement of the ester linkage with an amide does
not seem to affect the bioactivity of EGCG (1).
However, to our surprise, EGCG-amide (3) was significantly
more potent than EGCG (1) in reducing the viability of MDA-
MB-231 cells (Figure 2(b)). MDA-MB-231 was found to be less
susceptible than MCF-7 to the cytotoxic activity of EGCG (1)
(Figure 2), and 1 showed twofold increase in IC50 value against
MDA-MB-231 compared to that against MCF-7 cells
(EC50 = 22.7 μM and 45.6 μM for MCF-7 and MDA-MB-231,
respectively). A decrease in cytotoxic effect against MDA-MB-
231 cells is not unprecedented, but a general trend is observed in
anticancer drug discovery.17,18 However, EGCG-amide (3) was
more potent as an inhibitor of cell proliferation in MDA-MB-
231 cells than in MCF-7 cells (IC50 = 34.7 μM and 27.4 μM for
MCF-7 and MDA-MB-231, respectively).
Bull. Korean Chem. Soc. 2020
© 2020 Korean Chemical Society, Seoul & Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Wiley Online Library
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Communication
ISSN (Print) 0253-2964 | (Online) 1229-5949
BULLETIN OF THE
KOREAN CHEMICAL SOCIETY
(A)
(B)
120
120
MDA-MB-231 (EGCG, 1)
MDA-MB-231 (EGCG-Amide, 3)
MCF-7 (EGCG, 1)
MCF-7 (EGCG-Amide, 3)
100
80
60
40
20
0
100
80
60
40
20
0
Figure 1. Structures of the EGCG derivatives.
-20
0
20
40
60
80
100 120
0
20
40
60
80
100 120
Concentration (uM)
Concentration (uM)
Figure 2. Comparison of anticancer effects of EGCG (1) and
EGCG-amide (3) against (a) MCF-7 and (b) MDA-MB-231 cells.
Supporting Information. Additional supporting informa-
tion may be found online in the Supporting Information
section at the end of the article.
References
1. H. Liu, C. Zang, M. H. Fenner, K. Possinger, E. Elstner,
Breast Cancer Res. Treat. 2003, 79, 63.
2. K. J. Chavez, S. V. Garimella, S. Lipkowitz, Breast Dis.
2010, 32, 35.
3. H. J. Fujiki, Cancer Res. Clin. Oncol. 1999, 125, 589.
4. N. Khan, H. Mukhtar, Cancer Lett. 2008, 269, 269.
5. D. Kuhn, W. H. Lam, A. Kazi, K. G. Daniel, S. Song,
L. M. Chow, T. H. Chan, Q. P. Dou, Front. Biosci. 2005,
10, 1010.
6. K. R. Landis-Piwowar, C. Huo, D. Chen, V. Milacic, G. Shi,
T. H. Chan, Q. P. Dou, Cancer Res. 2007, 67, 4303.
7. Y. Zhong, Y. -S. Chiou, M. -H. Pan, C. -T. Ho, F. J. Shahidi,
Funct. Foods 2012, 4, 323.
SCHEME 1 Synthesis of EGCG-amide (3).
8. J. H. Jung, M. Yun, E.-J. Choo, S.-H. Kim, M.-S. Jeong, D.-
B. Jung, H. Lee, E.-O. Kim, N. Kato, B. Kim, S. K. Srivastava,
K. Kaihatsu, S.-H. Kim, Br. J. Pharmacol. 2015, 172, 3565.
9. Y. Zhong, F. J. Shahidi, Agric. Food Chem. 2011, 59, 6526.
10. S. Y. Han, E. Kim, K. Hwang, Z. A. Ratan, H. Hwang, E. -M. Kim,
D. Kim, J. Park, J. Y. Cho, Int. J. Mol. Sci. 2018, 19, 1466.
11. S. Y. Han, E. Kim, K. Hwang, Z. A. Ratan, H. Hwang, E.-M. Kim,
D. Kim, J. Park, J. Y. Cho, Cancer Cell Int. 2019, 19, 266.
12. H. Yang, D. K. Sun, D. Chen, Q. C. Cui, Y. Y. Gu, T. Jiang,
W. Chen, S. B. Wan, Q. P. Dou, Cancer Lett. 2010, 292, 48.
13. S. Bansal, S. Vyas, S. Bhattacharya, M. Sharma, Nat. Prod.
Rep. 2013, 30, 1438.
Considering the minimal structural difference between
EGCG (1) and EGCG-amide (3) resulting from the isosteric
replacement of an ester (1) with an amide (3) functionality,
the peculiar cytotoxicity profile of the EGCG-amide (3)
against MDA-MB-231 draws special attention. From a
structural point of view, the most notable difference
between an ester and an amide is the presence of a hydro-
gen bond donor on the amide functionality. Thus, one
might assume that the amide group on EGCG-amide (3) is
involved in a specific interaction with the characteristic tar-
get molecule in MDA-MB-231 cells. Identification of the
MDA-MB-231-specific target by using EGCG-amide (3) as
a probe molecule would provide valuable information for
development of anticancer agents for treatment of the
chemotherapy-resistant TNBC.
14. A. Kazi, Z. Wang, N. Kumar, S. C. Falsetti, T.-H. Chan,
Q. P. Dou, Anticancer Res. 2004, 24, 943.
15. S. C. Forester, J. D. Lambert, Mol. Nutr. Food Res. 2011, 55, 844.
16. K.-J. Min, T. K. Kwon, Integr. Med. Res. 2014, 3, 16.
17. C. Gest, U. Joimel, L. Huang, L.-L. Pritchard, A. Petit,
C. Dulong, C. Buquet, C.-Q. Hu, P. Mirshahi, M. Laurent,
F. Fauvel-Lafève, L. Cazin, J.-P. Vannier, H. Lu, J. Soria,
H. Li, R. Varin, C. Soria, BMC Cancer 2013, 13, 63.
18. Y. K. Park, Y. S. Ryu, H. Cho, H. S. Chung, S. H. Lee, Bull.
Kor. Chem. Soc. 2018, 39, 512.
Acknowledgments.
This research was supported by
Konkuk University in 2018.
Bull. Korean Chem. Soc. 2020
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