Synthesis and in vitro antiproliferative activities of lupanol derivatives towards human esophageal squamous carcinoma cells

Article abstract of DOI:10.1080/14786419.2020.1844687

A series of lupanol derivatives were synthesized and evaluated in vitro for their inhibitory activities against three human esophageal squamous carcinoma cells lines, Eca-109, TE-1 and EC-9706. Among lupanol derivatives, seven were new compounds, and lupanol cinnamate analogues 5 and 6, hydrazone analogues 9 and 10 presented high activities towards all the tested tumour cells, even higher activities than those of doxorubicine.

Full text of DOI:10.1080/14786419.2020.1844687

Natural Product Research
Formerly Natural Product Letters
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Synthesis and in vitro antiproliferative activities of
lupanol derivatives towards human esophageal
squamous carcinoma cells
Weijie Li & Yeyu Xiao
To cite this article: Weijie Li & Yeyu Xiao (2020): Synthesis and inꢀvitro antiproliferative activities
of lupanol derivatives towards human esophageal squamous carcinoma cells, Natural Product
Research, DOI: 10.1080/14786419.2020.1844687
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NATURAL PRODUCT RESEARCH
https://doi.org/10.1080/14786419.2020.1844687
Synthesis and in vitro antiproliferative activities of
lupanol derivatives towards human esophageal
squamous carcinoma cells
Weijie Lia and Yeyu Xiaob
aDepartment of Applied Chemistry, School of Chemical and Environmental Engineering, Hanshan
b
Normal University, Chaozhou, P.R. China; Guangzhou Hospital of Integrated Traditional and West
Medicine, Guangzhou, P.R. China
ABSTRACT
ARTICLE HISTORY
Received 16 July 2020
Accepted 27 October 2020
A series of lupanol derivatives were synthesized and evaluated
in vitro for their inhibitory activities against three human esopha-
geal squamous carcinoma cells lines, Eca-109, TE-1 and EC-9706.
Among lupanol derivatives, seven were new compounds, and
lupanol cinnamate analogues 5 and 6, hydrazone analogues 9
and 10 presented high activities towards all the tested tumour
cells, even higher activities than those of doxorubicine.
KEYWORDS
Lupanol; lupanol derivative;
synthesis; anticancer activity
1. Introduction
Lupanol (1) (Figure 1) is a natural pentacyclic triterpenoid with biological and pharma-
cological activities (Salem et al. 2016; Wang et al. 2020), its natural contents are rela-
tively small and there are fewer reports in the literatures. Recent studies have shown
that lupanol dicarboxylic acid monoester derivatives had good anti-HIV activities
because they were capable of inhibiting HIV-1 replication in lymphocytes (Yu et al.
2006) and had good anticancer activities against lung and liver cancers (Li et al. 2013).
Therefore, the study of lupanol derivatives has been paid more attention.
Except for structural determination of lupanone (7) and lupanone oxime (8), their
biological properties had not yet been described in the literatures (Wintersteiner et al.
CONTACT Y. Xiao
xyyu73@163.com
Supplemental data for this article can be accessed at https://doi.org/10.1080/14786419.2020.1844687.
ß 2020 Informa UK Limited, trading as Taylor & Francis Group

2
W. LI AND Y. XIAO
R2
R1O
O
H CO
5
6
. R1= 3
8. R =O
7
1
2. R =H (lupanol)
2
1
9.. RR2==HHO2NNN
3. R =ClCH CO
H3CO
H3CO
2
1
2
O
H
N
4.. RR1==((CCH2H5))N2NCCHH2CCOO
1
3 2
2
. R1=
N
10. R2=
H3CO
O2N
NO2
OCH3
Figure 1. Chemical Structures of lupanol derivatives.
1965; Kumar et al. 2008; Kuriata-Adamusiak et al. 2012; Khan et al. 2014). Here, to sys-
tematically investigate the anticancer effect of the 3b-hydroxy group of 1, nine lupanol
derivatives (2-10, Figure 1) were synthesized. Among lupanol derivatives, seven (2-6, 9
and 10) were new compounds. All the obtained compounds were evaluated in vitro
antiproliferative activities against three human esophageal squamous carcinoma cells
lines, Eca-109, TE-1 and EC-9706.
2. Results and discussion
2.1. Chemistry
Lupanol (1) reacted with excess chloroacetyl chloride in anhydrous dichloromethane
to obtain new compound 2 in high yield at room temperature (Figure 2), and the lat-
ter reacted with dimethylamine in 33% aqueous solution or diethylamine in dichloro-
methane to afford new compound 3 or 4, respectively with mild yields at ambient
temperature (Figure 2).
Esterification of various acid anhydrides or acids with various alcohols was per-
formed in good yields using N, N’-dicyclohexylcarbodiimide (DCC) as condensing
agent and 4-dimethylaminopyridine (DMAP) as catalyst under mild conditions (Yu
et al. 2006; Li et al. 2013). Here, 1 reacted with 3, 4-dimethoxycinnamic acid in anhyd-
rous dichloromethane to obtain new compound 5 with mild yield at room tempera-
ture in the presence of DCC as condensing agent and DMAP as catalyst and the
above reaction required strictly anhydrous condition (Figure 3).
3, 4, 5-Trimethoxycinnamic acid was refluxed with excess thionyl chloride to lead to
the formation of the corresponding acyl chloride, and the latter reacted with 1 in
anhydrous dichloromethane to give new compound 6 in good yield at room tempera-
ture (Figure 3).
Compound 7 is a known natural product, and its oxime (8) had been reported in
the literatures ( Kumar et al. 2008; Kuriata-Adamusiak et al. 2012; Banerjee et al. 2014
). Here, 7 and 8 were prepared according to the literature’s method (Kumar et al.
2008). Compound 7 was refluxed with hydrazine hydrate in 80% aqueous solution or

NATURAL PRODUCT RESEARCH
3
a
b
R2NCH2CO2
HO
ClCH2CO2
3
4
R=CH3
R=C2H5
2
1
(a). ClCH2COCl, CH2Cl2, r.t., 29 h, 93.5%; (b). dimethyl-amine, 33% w/w aq. soln. or
diethylamine, CH2Cl2, r.t., 24-68 h, 61.8%-65.3%.
Figure 2. Synthesis of compound 2–4.
a
O
H3CO
H3CO
O
HO
1
5
O
H3CO
H3CO
OH
O
b
c
H3CO
H3CO
O
OCH3
6
OCH3
(a). 3, 4-dimethoxycinnamic acid, DCC, DMAP, CH2Cl2, r.t., 72 h, 50.6%; (b). SOCl2,
reflux, 4 h; (c). lupanol, CH2Cl2, r.t., 60 h, 81.6%.
Figure 3. Synthesis of compounds 5 and 6.
a
R
O
H
9
R=H2NN
N
7
N
10 R=
O2N
NO2
(a). hydrazine hydrate in 80% aqueous solution, reflux, 10 h, 58.5%; or 2, 4-dinitro-
phenylhydrazine, ethyl alcohol, reflux, 10 h, 53.2%.
Figure 4. Synthesis of compounds 9 and 10.

4
W. LI AND Y. XIAO
2, 4-dinitrophenylhydrazine in ethanol to afford new compound 9 or 10, respectively
with mild yields (Figure 4).
2.2. In vitro inhibitory activities on proliferation of human esophageal
squamoous carcinoma cells
Using MTT assay, the in vitro anticancer activities of all the synthesized lupanol deriva-
tives were evaluated against three human esophageal squamous carcinoma cells lines,
Eca-109, TE-1 and EC-9706. Here, doxorubicine was used as a positive control. It had
been proven one of the most effective single anticancer agents with an impressively
broad spectrum of activity encompassing many of the solid tumors. It was widely
used as a positive control for in vitro screening of anticancer compounds in the litera-
tures ( Cortes-Funes et al. 2007; Hao et al. 2011; Li et al. 2013 ). According to evalu-
ation criteria of in vitro anticancer activity, the anticancer activity of each compound
was expressed as IC50 value to cancer cells. The IC50 value represented the concentra-
tion of a compound that was required for 50% inhibition towards cancer cells growth
in vitro. Here, all data were presented as mean SD (standard deviation) of three dif-
ferent experiments. Statistical analysis was performed using SPSS 12.0 software. The
results were recorded in Table S1.
If IC50 value was less than 50 mM, the sample was evaluated to have anticancer
activity. The smaller IC50 value was, the better the anticancer activity of the tested
sample was. According to the IC50 values, doxorubicine had good inhibitory abilities
against all the tested human esophageal squamous carcinoma cells growth. Lupanol
(1), its chloroacetate (2) and glycine ester derivatives 3 and 4 had no anticancer activ-
ities against all the tested cancer cells. However, the IC50 values of lupanol cinnamate
analogues 5 and 6 were less than 8 mM towards Eca-109 cells and TE-1 cells, and less
than 33 mM against EC-9706 cells. Their anticancer activities surpassed those of doxor-
ubicine. The 3b-hydroxyl group of 1 was oxidized to form carbonyl group derivative 7,
the latter reacted with hydroxylamine to give lupanone oxime 8, and they did not
possess anticancer activities. However, hydrazone analogues 9 and 10 presented sig-
nificant inhibitory effects against all the tested three human esophageal squamous
carcinoma cells lines, even higher activities than those of doxorubicine.
3. Conclusions
Nine lupanol derivatives were synthesized and evaluated in vitro for their antiprolifera-
tive activities against Eca-109, TE-1 and EC-9706 cells. According to the above study
for the anticancer effect of 3b-hydroxy group of lupanol, the following conclusions
can be obtained: (1) The 3b-hydroxyl group of lupanol is esterified with 3, 4-dimethox-
ycinnamoyl or 3, 4, 5-trimethoxycinnamoyl group, the anticancer activities of the deriv-
atives are promoted. (2) The 3b-hydroxyl group is converted to carbonyl group, the
anticancer activities of the derivatives disappear. (3) After the imidization of carbonyl
group, only hydrazone analogues 9 and 10 exhibit good inhibitory abilities against
Eca-109, TE-1 and EC-9706 cells proliferation. (4) Among lupanol derivatives, the most
promising compounds 5, 6, 9 and 10 exhibit significant antiproliferative activities

NATURAL PRODUCT RESEARCH
5
against all the tested human esophageal squamous carcinoma cells, even higher activ-
ities than those of doxorubicine.
Disclosure statement
No potential conflict of interest was reported by the authors.
Funding
The work was financially supported by Scientific Research Project of Traditional Chinese
Medicine Bureau of Guangdong Province (20202141); Natural Science Foundation of Guangdong
Province, China (2018A0303070002).
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