Synthesis and biological evaluation of α-methyl-chalcone for anti-cervical cancer activity

Article abstract of DOI:10.1007/s00044-017-1891-0

A series of 31 chalcones were synthesized and evaluated for anti-proliferative activity against the human cervical cancer cell lines (HeLa and SiHa). Compound 19, (E)-1-(2,4-dihydroxyphenyl)-3-(4-(dimethylamino) phenyl)-2-methylprop-2-en-1-one was found to be an effective anti-proliferative agent in HeLa and SiHa cells (IC₅₀ = 0.035 μM). Compound 19 increased the percentage of apoptosis in a dose-dependent manner, as measured by Annexin V-FITC (fluorescein isothiocyanate)/(propidium iodide) PI staining. Molecular modeling studies of compound 19 showed that the most potent structure was docked at the yeast 20 S proteasome binding site (Protein Data Bank code-3E47) and was stabilized in the cavity by various hydrophobic and hydrogen bonding interactions.

Full text of DOI:10.1007/s00044-017-1891-0

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-017-1891-0
ORIGINAL RESEARCH
Synthesis and biological evaluation of α-methyl-chalcone for anti-
cervical cancer activity
Bing-zhao Ren1 Mourboul Ablise1 Xu-chao Yang1 Bo-er Liao1 Zheng Yang1
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Received: 29 November 2016 / Accepted: 4 April 2017
© Springer Science+Business Media New York 2017
Abstract A series of 31 chalcones were synthesized and
evaluated for anti-proliferative activity against the human
cervical cancer cell lines (HeLa and SiHa). Compound 19,
response, cell cycle regulation, metabolism, and protein
expression. It is the 26S proteasome that completes ATP-
dependent ubiquitin protein degradation (Tanaka 2009).
The 26S proteasome is composed of one 20S catalytic
subunit, which is known as the “proteolytic core” and two
19S regulatory subunits, which are termed “caps”
(Ciechanover et al. 1984; Ciechanover 1993). The 20S
proteasome contains fourteen α subunits and fourteen β
subunits. Each β subunit is composed of three catalytic
sites: β1 (caspase-like), β2 (trypsin-like), β5 (chymotrypsin-
like). The proteolytic activities of three subunits use the γ-
hydroxyl group of an N-terminal threonine residue within
each catalytic site for nucleophilic attack of the α-amine
proton donor/acceptor within the targeted protein (Bazzaro
et al. 2008).
Chalcones are naturally occurring compounds that are
widely distributed in a variety of plant species belonging to
the flavonoid family. They are known to exhibit several
biological activities (Zhao et al. 2005; Valla et al. 2006;
Lawrence et al. 2006; Hsu et al. 2006; Ahmad et al. 2006;
Dimmock et al. 1999; Ni et al. 2005), including antiplatelet
activity, inhibition of pro-inflammatory mediators, anti-
proliferative action in cancer cells, apoptosis induction,
antimalarial activity, and others. The method of synthesis of
chalcones is mainly the Claisen—Schmidt condensation
reaction, with a basic catalyst, such as NaOH (Romagnoli
et al. 2009; Robert and Jarry 2003; Ahmed Belkacem et al.
2006), KOH (Zamora et al. 1988), and Ba(OH)2 (Lee et al.
1994; Liu et al. 2008), acetophenones, and benzene for-
maldehyde dehydration condensation. Other basic catalysts
such as magnesium tert-butoxide (Lawrence and McGown
2005), K2CO3, Al2O3 (Seelig and Landwojtowicz 2000),
MgO (Ducki et al. 1998), calcined hydrotalcites (Bois et al.
1998, 1999), the natural phosphate/NaNO3 (Pati et al. 2005;
Xia et al. 2000), the natural phosphate/KF (Dimmock et al.
(E)-1-(2,4-dihydroxyphenyl)-3-(4-(dimethylamino)
nyl)-2-methylprop-2-en-1-one was found to be an effective
anti-proliferative agent in HeLa and SiHa cells (IC50
phe-
=
0.035 μM). Compound 19 increased the percentage of
apoptosis in a dose-dependent manner, as measured by
Annexin V-FITC (fluorescein isothiocyanate)/(propidium
iodide) PI staining. Molecular modeling studies of com-
pound 19 showed that the most potent structure was docked
at the yeast 20 S proteasome binding site (Protein Data
Bank code-3E47) and was stabilized in the cavity by var-
ious hydrophobic and hydrogen bonding interactions.
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Keywords Chalcone α,β-Unsaturated Carbonyl Anti-
●
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cervical cancer molecular modeling Anti-mitotic
Synthesis
Introduction
The ubiquitin-proteasome system is one of the major
pathways of intracellular protein degradation, playing a
prominent role in signal transduction, cell death, immune
Electronic supplementary material The online version of this article
(doi:10.1007/s00044-017-1891-0) contains supplementary material,
which is available to authorized users.
* Mourboul Ablise
mourboul@sohu.com
1
Institute of Pharmacy, Xinjiang Medical University, Urumchi,
China

Med Chem Res
2003) and piperidine (Liu and Go 2006) are also often used
for the synthesis of chalcones. The reaction can also use an
acid catalyst such as HCl, BF3, B2O3, PTSA, SOCl2/EtOH,
BF3–Et2O (Petrov et al. 2008), TiCl4 (Sogawa et al. 1993),
Zeolite (Chimenti et al. 2005), and RuCl3 (Climent et al.
1990). However, these catalysts have varying degrees of
shortcomings, such as low yield and harsh reaction condi-
tions, or are harmful to the environment. Microwave reac-
tors have high yield, simple operation and post-processing
(Iranpoor and Kazemi 1998; Deka and Sarma 2001; Kalita
et al. 1999; Saikia et al. 2009).
Groll et al. 2006 reported a crystallographic analysis of
the yeast 20 s proteasome in complex with the inhibitor
homobelactosin C. Catalytic activities of these β-type sub-
units are associated with their N-terminal threonine residue,
which acts as the nucleophile in hydrolysis, thus classifying
proteasomes as members of the family of N-terminal
nucleophilic hydrolases (Löwe et al. 1995; Brannigan
et al. 1995). The 20 s proteasome subunits utilize the
hydroxyl group of an N-terminal theronine within each
catalytic site for nucleophilic attack of the carbonyl group of
homobelactosin C, which leads inactivation of the protea-
some. The α,β-unsaturated carbonyl system is the activity
group of chalcone-based derivatives that inhibits protea-
some activity in human papilloma virus-positive cervical
cancer cells (Bazzaro et al. 2011).
Compounds 2–9, having electron donating groups,
exhibited anti-proliferative activity. Notably, the target
compounds of isoliquiritigenin derivatives showed dose-
dependent and time-dependent growth inhibition in SiHa
cells and HeLa cells. The isoliquiritigenins with an –OCH3
group in the meta-positions on aromatic ring B were
selectively anti-proliferative in SiHa cells, due to their
increased lipid solubility, which caused greater penetration
into the cells. Hydroxyl group substitutions in chalcones
have been shown to play a key role in anti-cancer activities
of chalcones. The isoliquiritigenin with –OH groups in the
meta-positions and in the ortho-positions on the aromatic
ring B induced a lower degree of growth inhibition than
those without –OH groups. Thus, the –OCH3 and –OH
groups are important in influencing selectivity and potency
of chalcone derivatives.
Compounds 10–16, bearing –F on their A-rings and B-
rings were initially synthesized and evaluated in terms of
their activity. Those with an electron withdrawing group
exhibited greater cytotoxic activity, and the cytotoxic
activities of these compounds were related to the electro-
philic nature of their substituent. For example, the IC50
value of compound 11 was 3.35 μM.
Comparisons were made to determine the effect of hav-
ing a methyl group in the α position and another substituent
(–OH, –NO3, –F, –N(CH3)2, –OCH3) on rings A and B
(chalcones 17–31). Chalcones 17–31 showed the most
potent anti-proliferative effect against HeLa and SiHa cells,
indicating that the methyl group on α position plays an
important role.
Chalcone 19 exhibited the most potent anti-proliferative
effect against SiHa cells with lowest IC50 (0.035 μM).
Morphological changes (Fig. 1) show that chalcone 19
exhibited a greater growth inhibitory rate in SiHa cells at all
concentrations than in HeLa cells. The anti-proliferative
effects exhibited by chalcone 19 were also dose-dependent.
From the photos (Fig. 1), it looks like cisplatin is more
potent than compound 19 against cancer cells. In non-
cancerous cells, cisplatin killed more cells than compound
19. This could be interpreted as better selectivity of com-
pound 19 for cancer cells than cisplatin and that it would be
investigated further. Figure 2 shows that compound 19
displayed the highest anti-proliferative activity and there-
fore was chosen for further investigation. All growth inhi-
bitory effects induced by chalcones were significantly
different (p < 0.05) as compared with cells treated with 1%
DMSO.
Results and discussion
Chemistry
The chalcones were classified into 4 groups according to
their substitution patterns on ring A and ring B (i.e., their 1-
phenyl moiety). Group 1 chalcones (1–4) have no ring A
substitution and ring B is substituted with a hydroxyl group
at one position; Group 2 chalcones (5–9) are iso-
liquiritigenin derivatives; Group 3 chalcones (10–16) are
fluorine isoliquiritigenin derivatives, and Group 4 chalcones
(17–31) are α-methyl-chalcone derivatives. All synthetic
compounds (Table 1) had satisfactory analysis and spectral
data, which were completely described by their structure.
Biological activity
Anti-proliferation assay
The methyl thiazolyl tetrazolium (MTT) assay system was
used to identify compounds with anti-proliferative activity.
Human cervical cancer cells (HeLa and SiHa), normal
Chinese Hamster Ovary (CHO) were incubated for 24 h/48
h/72 h at 37 °C. Table 2 lists the IC50 values of the chal-
cones for each cell line.
Apoptosis assay
Apoptosis is an essential mechanism used to eliminate
activated cells during the shut down process of excess
immune responses and to maintain proper immune

Med Chem Res
Table 1 Chalcone derivatives
and their melting points
Chalcone ring A
Ring B
No.
R2
R3
R4
R5
R6
R
R′2
R′3
R′4
R′5
R′6
MP °C
1
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
OH
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
55–56
2
H
H
H
H
H
149–151
223–224
159–161
198–200
207–209
198–203
220–221
196–198
103–105
108–109
104–105
106–107
105–107
110–111
110–112
163–164
176–178
89–92
3
H
H
H
OH
H
H
4
H
H
H
OH
OH
OH
OCH3
OH
OH
F
5
OH
OH
H
OH
OH
OH
OH
OH
F
H
H
6
H
OCH3
OH
OH
H
7
H
8
OH
H
H
9
H
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
F
H
H
OH
H
OH
OH
H
H
H
F
H
H
F
OH
F
H
H
F
F
H
H
OH
OH
OCH3
NO2
F
F
F
H
OCH3
OH
H
F
F
H
OH
OH
OH
OH
OH
H
OH
OH
OH
OH
OH
H
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
H
H
N(CH3)2
H
H
113–115
103–105
103–105
100–102
85–87
OCH3
H
OH
NO2
F
H
H
H
H
H
H
N(CH3)2
H
H
H
H
115–117
95–97
H
H
OCH3
H
OH
NO2
F
H
OH
OH
OH
OH
OH
91–93
H
H
90–93
H
H
N(CH3)2
H
121-123
92–93
H
H
H
OCH3
OH
123–125
homeostasis, while deficient apoptosis of activated cells is
associated with a wide variety of immune disorders. As a
representative of the chalcone thiosemicarbazide deriva-
tives, compound 19 was further evaluated for its apoptotic
effect. We assessed the mechanism underlying the
cell growth inhibition by compound 19 by flow
V-FITC/PI staining, in
(Figs 3 and 4).
a
dose-dependent manner
Molecular docking simulation
In order to obtain a deeper insight into the molecular basis of
the enzyme-inhibitor recognition process, docking simulation
of the most active (compound 19) was performed to inves-
tigate its binding mode to the 20S proteasome that is
co-crystallized with the β5-inhibitor, homobelactosin
cytometry in SiHa and HeLa cells following
a
24 h treatment with 0.01, 0.1, 1, 6.25, 12.5 μg mL−1 of
compound 19. The compound increased the percentage of
apoptosis in both cell lines, as revealed by Annexin

Med Chem Res
Table 2 The anti-proliferative
effects of chalcones for cancer
cells and CHO cells
Chalcone no. HeLa IC50 μM
SiHa IC50 μM
CHO IC50 μM
24 h
48 h
72 h
24 h
48 h
72 h
24 h
48 h
72 h
1
>100
>100
>100
>100
>100
63.01
>100
>100
98.21 89.27
2
67.21
70.45
>100
60.74
45.05
61.87
63.72 >100
66.82
52.2
97.66 80.33 75.23
40.32 35.56 23.71
51.23 45.65 32.71
55.12 43.57 27.42
60.87 53.45 38.27
59.36 47.51 40.03
56.15 48.35 38.76
48.69 45.68 32.11
49.36 47.23 35.12
48.71 44.36 31.56
45.63 38.52 33.28
47.64 40.66 34.75
40.36 35.36 28.57
43.81 37.83 29.49
41.86 37.92 30.01
47.56 40.42 31.2
50.94 45.36 34.75
54.68 47.55 38.49
55.11 48.21 37.14
56.47 46.38 37.08
59.46 45.55 38.33
63.45 59.46 40.59
57.31 48.54 40.91
58.64 49.59 34.31
48.31 36.89 32.66
57.13 52.81 45.76
41.36 39.46 37.21
39.45 38.23 36.32
37.64 23.16 20.36
49.15 45.63 42.34
3
65.34
76.81
23.92
34.6
74.23
68.67
80.53
23.61
25.34
44.25
1.74
62.3
75.6
18.41
21.63
33.6
0.182
6.89
10.23
3.35
12.76
5.02
8.1
4
5
31.01
46.34
48.03
62.4
19.75
23.38
16.78
6.87
29.21
6
38.14
46.1
7
31.57
29.43
14.56
23.66
32.67
26.57
27.43
19.44
30.23
16.94
6.5
8
75.21
34.97
23.32
15.54
25.21
17.67
16.97
22.54
22
9
17.72
42.54
40.34
39.34
34.44
27.25
38.45
39.65
10.43
13.35
10.45
17.04
13.32
20.15
23.65
19.84
25.98
17.56
25.61
19.58
18.66
25.98
20.36
13.13
19.75
10.25
15.65
16.57
12.43
20.56
14.56
0.231
2.34
14.81
16.72
10.43
16.54
12.87
12.48
15.67
15.76
2.12
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
10.79
5.87
0.187
0.673
0.035
0.178
0.105
1.36
2.03
0.94
3.12
2.41
1.34
2.13
5.79
6.85
4.31
5.11
6.22
5.35
8.48
4.45
0.082
0.56
4.35
1.01
5.56
4.24
3.46
8.76
2.53
6.32
1.34
10.11
15.23
11.58
16.42
17.22
11.03
13.23
9.55
2.54
10.53
16.54
14.13
18.36
23.54
23.12
17.64
19.71
18.54
17.64
2.65
1.65
4.31
1.11
1.85
1.84
6.31
0.912
0.847
1.035
2.156
1.65
5.24
6.17
8.23
5.46
6.89
5.24
14.63
2.360
3.85
C. Structures were retrieved from the Protein Data Bank,
PDB (Rocco et al. 2006). Compound 19 was docked at the
homobelactosin C binding site of the yeast 20S proteasome
(a, b in Fig. 5a) and the best fit conformation was selected on
the basis of the Gold score and visual inspection.
The docking study showed that compound 19 is stabilized
in the 20S proteasome by hydrogen bonding and hydrophobic
interactions. Many of the amino acid residues, which are in the
formation of the ligand-binding pocket, i.e., THR1, THR21,
GLY47, and SER129, formed H-bonds with hydroxyl and
carbonyl groups of compound 19. The oxygen atom of the
para-hydroxyl group of ring A acts as an H-bond acceptor and
is involved with chain NH of THR-21 (d = 2.9 Å). The car-
bonyl group of α,β-unsaturated carbonyl system was located
2.7 Å away from the nucleophilic serine residue (SER-129), a
distance that is typical of a hydrogen bonding interaction in
this type of emulation with a rigid protein frame. Figure 5
shows that additional but weak H-bonds were also observed
between compound 19 and other amino acid residues (d =
2.7–3.2 Å). The dimethylamino group of compound 19 is
extended toward the β subunits of the ligand-binding pocket
positioned in a hydrophobic cavity formed by Gly47 and
Gly98. The molecular docking studies suggest a nucleophilic
attack from the N-terminal threonine residue of the β-subunits
to the carbonyl group of compound 19 as a mechanism for
proteasomal inhibition.
Conclusion
A series of chalcone derivatives containing electron-
withdrawing and electron-donating substituents have been

Med Chem Res
Fig. 1 The morphological
changes of human cervical
cancer and CHO cells with
compound 19
Fig. 2 Inhibition rate of
compound 19 toward SiHa
a and HeLa b cells after 24,
48 and 72 h
designed and synthesized and their biological activities
were also evaluated as potent and selective anti-proliferative
activity against human cervical cancer cells (SiHa and
HeLa). Chalcone 19 showed the most potent growth inhi-
bitory activity toward SiHa and HeLa cells with IC50 values
of 0.035 and 0.082 μM, which was slightly lower than the
positive control (cisplatin), and ideal quality of a potential
cancer drug—low toxicity to normal tissue with IC50 value
of 38.49 μM toward CHO cells, which was obviously
higher than cisplatin.
X-4 melting point apparatus and are uncorrected. Thin layer
chromatography (TLC) analysis was done using silica gel
60 F254 analytical plates (Merck, Billerica, MA, USA). The
reaction products were purified by crystallization or flash
column chromatography using a mixture of petroleum ether/
ethyl acetate and hexane/ethyl acetate as the eluent. FT-IR
spectra were carried out on Bruker RFS100/S spectro-
photometer (Bio-Rad, Cambridge, MA, USA) in the
400–4000 cm−1 range. 1H-NMR spectra were recorded on
Varian Inova-600 MHz instruments (Varian, Palo Alto, CA,
USA). 13C-NMR spectra were recorded on a 600 MHz
Bruker ARX-600 spectrometer (Bruker Bioscience, Bill-
erica, MA, USA). The chemical shifts (δ) are reported in
parts per million downfield relative to tetramethylsilane. All
tested compounds were >95% purity as determined by
preparative TLC, which was made using silica gel GF254
on glass plates. The chalcones were prepared according to
methods previously described in the literature (Leong et al.
2007, 2005; Rocco et al. 2006).
Experimental
Chemistry
All of the solvents and reagents used in the current study
were purchased from commercial suppliers and used with-
out further purification. Melting points were recorded on an

Med Chem Res
Fig. 3 The apoptotic effect of compound 19 in SiHa cells
Fig. 4 The apoptotic effect of compound 19 in HeLa cells

Med Chem Res
Fig.5 Docking simulation of lead compound 19 to the 20S protea-
some. a Compound 19 (red) and the ligand of proteasome (green) are
binding at the A, B, and C active sites. b and c Schematic view of lead
compound 19 (green) docked to the crystal structure of the 20S pro-
teasome (color figure online)
General methods for chalcones 2–9
prepared according to this method were formed pre-
dominantly with the (E)-configuration.
The chalcones (2–9) evaluated in the current study were
synthesized by the base catalyzed Claisen–Schmidt con-
densation reaction of the appropriately substituted acet-
ophenones and aldehydes (Scheme 1). The chalcones
Acetophenones (0.01 mol), benzaldehydes (0.012 mol),
8 g alumina, and 4 g potassium hydroxide were homo-
genized in a mortar. Then, the mixture was irradiated in a
reaction vessel of a CW-2000 focused microwave reactor

Med Chem Res
Scheme 1 Schematic
representation of the synthesis
and chemical structures of
chalcones 2–9
Scheme 2 Schematic
representation of the synthesis
and chemical structures of
chalcones 1, 10–16
Scheme 3 Schematic
representation of the synthesis
and chemical structures of
chalcones 17–31
for 100 s after setting reaction power to 150 W. After
cooling to room temperature, ice water (250 mL) was added
to the reaction mixture and filtered through a general
laboratory filter paper to separate the solid catalyst.
Hydrochloric acid (2 mol L−1) was added to the filtrate until
the pH was 1–2 and the mixture was refrigerated at 4 °C for
20 min. The crude product was filtered and the residue was
chromatographed on a flash silica-gel column using
25–50% EtOAc in hexane as the eluent.
flash silica-gel column using 25% EtOAc in hexane as
eluant.
Characterization data
(E)-1,3-diphenyl-2-propen-1-one (compound 1) Yellow
1
solid (63.31%), m.p. 55.1–56.8 °C; H NMR (CDCl3, 600
MHz): δ 8.01–8.03 (m, 2H), 7.81 (d, J = 15.98 Hz, 1H),
7.63–7.65 (m, 2H), 7.55–7.60 (m, 2H), 7.48–7.52 (m, 2H),
7.39–7.42 (m, 3H); 13C NMR (CDCl3, 150 MHz): δ 190.52,
144.81, 138.16, 134.83, 132.76, 130.52, 128.93, 128.59,
128.47, 128.41, 122.02. MS: m/e 209 (M + H+). Anal:
C15H12O/208.
General methods for chalcones 1, 10–16
The chalcones (1, 10–16) were synthesized by the base
catalyzed Claisen–Schmidt condensation reaction of the
appropriately substituted acetophenones and aldehydes
(Scheme 2).
(E)-3-(2-hydroxyphenyl)-1-phenylprop-2-en-1-one (com-
pound 2) Light green solid (65.41%); m.p. 148.8–151.3 °
1
C; H NMR (CDCl3, 600 MHz): δ 8.09 (d, J = 16.05 Hz,
Acetophenones (0.01 mol), benzaldehydes (0.012 mol)
and absolute ethyl alcohol (32 mL) were stirred in a 250 mL
three-necked flask at 0 °C for 1 h. A solution of potassium
hydroxide (20 mmol) in a 4:1 (v/v) mixture of ethanol/H2O
(50 mL) was added into a flask containing N2 and the
resulting mixture was stirred at room temperature. The
reaction was then monitored by TLC using ethyl acetate/
petroleum ether (1:4 or 1:2 v/v) as the solvent system. The
crude product was filtered and the residue was chromato-
graphed on a flash silica-gel column using 25–50% EtOAc
in hexane as eluent.
1H), 7.97 (d, J = 6.93 Hz, 2H), 7.65 (d, J = 16.05 Hz, 1H),
7.51 (t, J = 8.75 Hz, 2H), 7.43 (t, J = 8.39 Hz, 2H), 7.23 (d,
J = 8.02 Hz, 1H), 6.89 (t, J = 8.02 Hz, 1H), 6.84 (d, J =
8.39 Hz, 1H), 6.31 (bs, OH); 13C NMR (CDCl3, 150 MHz):
δ 191.07, 155.62, 140.63, 138.36, 132.48, 131.81, 129.68,
128.68, 123.00, 122.34, 121.11, 116.63. MS: m/e 225 (M
+ H+). Anal: C15H12O2/224.
(E)-3-(3-hydroxyphenyl)-1-phenylprop-2-en-1-one (com-
pound 3) Yellow solid (70.45%); m.p. 223.4–224.6 °C;
1H NMR (CDCl3, 600 MHz): δ 8.01 (d, J = 7.80 Hz, 2H),
7.76 (d, J = 15.61 Hz, 1H), 7.58–7.61 (m, 1H), 7.51 (t, J =
7.80 Hz, 3H), 7.30 (t, J = 7.80 Hz, 1H), 7.22 (d, J = 8.30
Hz, 1H), 7.14 (s, 1H), 6.90–6.92 (m, 1H), 5.27 (bs, OH);
13C NMR (CDCl3, 150 MHz): δ 190.86, 155.98, 144.71,
138.08, 136.53, 133.02, 130.28, 128.76, 128.63, 122.49,
121.27, 117.82, 114.97. MS: m/e 225 (M + H+). Anal:
C15H12O2/224.
General methods for chalcones 17–31
The chalcones (17–31) were synthesized by the base cata-
lyzed Claisen–Schmidt condensation reaction of the
appropriately substituted acetophenones and aldehydes
(Scheme 3).
A solution of 2,5-dimethoxypropiophenone (14.8 g,
0.076 mol), 4-(dimethy1amino) benzaldehyde (11.6 g,
0.078 mol), piperidine (15 mL), and acetic acid (7.5 mL) in
ethanol (80 mL) was heated at reflux. After 18 h the solvent
was removed and the residue was chromatographed on a
(E)-3-(4-hydroxyphenyl)-1-phenylprop-2-en-1-one (com-
pound 4) Yellow solid (55.25%); m.p. 158.7–161.2 °C;
1H NMR (CDCl3, 600 MHz): δ 8.00 (d, J = 7.08 Hz,
2H),7.78 (d, J = 15.45 Hz, 1H), 7.55–7.59 (m, 3H), 7.50

Med Chem Res
(t, J = 7.72 Hz, 2H), 7.41 (d, J = 15.45 Hz, 1H), 6.89 (d, J
= 8.36 Hz, 2H), 5.62 (bs, OH); 13C NMR (CDCl3, 150
MHz): δ 191.06, 158.20, 145.04, 138.99, 132.78, 130.66,
128.70, 128.55, 127,74, 119.83, 116.10. MS: m/e 225 (M
+ H+). Anal: C15H12O2/224.
(E)-1,3-bis(4-hydroxyphenyl)prop-2-en-1-one (compound
9) Yellow solid (38.75%), mp.196.2–198.4 °C; H-NMR
1
(DMSO, 600 MHz) δ: 6.82 (2H, d, J = 8.4 Hz, H-3, 5), 6.87
(3H, d, J = 9 Hz, H-3′, 5′), 7.61 (1H, d, J = 15.6 Hz, H-α),
7.68 (1H, d, J = 15.6 Hz, H-β), 7.70 (2H, d, J = 8.4 Hz, H-
2, 6), 8.03 (3H, d, J = 9 Hz, H-2′, 6′), 10.03 (1H, s, OH),
10.34 (1H, s, OH); 13C-NMR (DMSO, 150 MHz) δ: 188.9,
163.8, 161.7, 145.0, 132.8, 132.6, 127.9, 120.4, 117.6,
117.2. MS: m/e 241 (M + H+). Anal: C15H12O3/240.
(E)-1-(2,4-dihydroxyphenyl)-3-(4-hydroxyphenyl)prop-2-
en-1-one (compound 5) Yellow solid (58.90%), m.p.
1
198.1–199.7 °C; H-NMR (DMSO, 600 MHz) δ: 6.26 (1H,
d, J = 1.8 Hz, H-3′), 6.4 (1H, dd, J = 1.8 Hz, J1 = 8.4 Hz,
H-5′), 6.83(2H, d, J = 9 Hz, H-3, 5), 7.74 (1H, d, J = 8.4
Hz, H-6′), 7.78 (1H, d, J = 15.2 Hz, α-H), 7.85 (1H, d, J =
15.2 Hz, β-H), 8.15 (2H, d, J = 9 Hz, H-2, 6), 13.60 (1H, s,
OH); 13C-NMR (DMSO,150 MHz) δ: 193.4, 167.7, 166.7,
162.1, 146.1, 134.7, 133.1, 127.6, 119.3, 117.7, 114.8,
110.1, 104.5. MS: m/e 257 (M + H+). Anal: C15H12O4/256.
(E)-1-(2,4-difluorophenyl)-3-(4-fluorophenyl)prop-2-en-1-
one (compound 10) Red solid (43.2%), m.p. 103.2–105.6 °C;
1H-NMR (DMSO, 600 MHz) δ: 8.27 (2H, d, J = 7.8 Hz, H-
2,6), 7.57 (2H, d, J = 8.4 Hz, H-3, 5), 7.48 (1H, d, J = 15 Hz,
α-H), 7.75 (1H, d, J = 15 Hz, β-H), 6.24 ~ 7.25 (3H, m, ArH).
13C-NMR (DMSO, 150 MHz) δ: 191.2, 164.2, 146.5, 144.1,
143.2, 140.3, 137.4, 133.1, 132.5, 129.0, 121.6, 112.5, 102.3.
MS: m/e 263 (M + H+). Anal: C15H9F3O/262.
(E)-1-(2,4-dihydroxyphenyl)-3-(4-hydroxy-3-methoxyphe-
nyl)prop-2-en-1-one
(compound
6) Yellow
solid
(57.60%), m.p. 206.7–209.6 °C; 1H-NMR (DMSO, 600
MHz) δ: 3.87 (3H, s, OCH3), 6.28 (1H, d, J = 1.8 Hz, H-2),
6.42 (1H, dd, J1 = 2.4 Hz, J2 = 9.0 Hz, H-5′), 6.78 (1H, d, J
= 8.4 Hz, H-5), 7.28 (1H, dd, J1 = 1.8 Hz, J2 = 8.4 Hz, H-
6), 7.54 (1H, d, J = 2.4 Hz, H-3′), 7.73 (1H, d, J = 15 Hz,
H-α), 7.79 (1H, d, J = 15 Hz, H-β), 8.2 (1H, d, J = 9.0 Hz,
H-6′), 13.65 (1H, s, OH); 13C-NMR (DMSO, 150 MHz) δ:
193.4, 167.7, 166.8, 151.8, 149.9, 146.6, 134.7, 128.1,
57.5. MS: m/e 287 (M + H+). Anal: C16H14O5/286.
(E)-1-(2,4-dihydroxyphenyl)-3-(4-fluorophenyl)prop-2-en-
1-one (compound 11) Yellow solid (32.1%), m.p.
108.3–109.0 °C; H-NMR (DMSO, 600 MHz) δ: 8.35 (2H,
d, J = 8.6 Hz, H-2,6), 7.71 (2H, d, J = 8.4 Hz, H-3, 5), 7.51
(1H, d, J = 15 Hz, α-H), 7.79 (1H, d, J = 15 Hz, β-H), 6.25
~ 7.67 (3H, m, Ar′H) 10.50(1H, s, OH-4′), 12.62(1H, s,
OH-2′). 13C-NMR (DMSO, 150 MHz) δ: 191.8, 169.2,
167.5, 164.9, 144.6, 132.5, 131.4, 120.6, 118.1, 116.1,
113.3, 108.1, 103.5. MS: m/e 259 (M + H+). Anal:
C15H11FO3/258.
1
(E)-3-(3-hydroxy-4-methoxyphenyl)-1-(4-hydroxyphenyl)
prop-2-en-1-one (compound 7) Yellow solid (52.90%), m.
p. 198.1–201.6 °C; H-NMR (DMSO, 600 MHz) δ: 3.83
(E)-3-(4-fluorophenyl)-1-(4-hydroxyphenyl)prop-2-en-1-
one (compound 12) Yellow solid (36.0%), m.p.
104.7–105.4 °C; H-NMR (DMSO, 600 MHz) δ: 8.31 (2H,
d, J = 8.6 Hz, H-2,6), 7.70(2H, d, J = 8.4 Hz, H-3, 5), 7.67
(1H, d, J = 15 Hz, α-H), 7.82 (1H, d, J = 15 Hz, β-H), 7.91
(2H, d, J = 8.7 Hz, H-3′, 5′), 8.40 (2H, d, J = 8.4 Hz, H-
2′,6′), 10.54 (1H, s, OH-4′). 13C-NMR (DMSO, 150 MHz)
δ: 193.5, 170.5, 167.3, 145.4, 141.2,123.1, 129.6, 120.5,
115.6, 114.6, 101.6. MS: m/e 243 (M + H+). Anal:
C15H11FO2/242.
1
1
(3H, s, OCH3), 6.82 (1H, d, J = 1.8 Hz, H-2), 6.88 (2H, d, J
= 9.0 Hz, H-3′,5′), 6.98 (1H, d, J = 9 Hz, H-5), 7.26 (1H,
dd, J1 = 1.8 Hz, J2 = 9 Hz, H-1,6), 7.55 (1H, d, J = 15.6
Hz, H-α), 7.65 (1H, d, J = 15.6 Hz, H-β), 8.03 (2H, d, J =
8.4 Hz, H-2′,6′), 9.13 (1H, s, OH-3), 10.36 (1H, s, OH-4′);
13C-NMR (DMSO, 150 MHz) δ: 193.4, 164.0, 151.9,
148.5, 145.1, 133.1, 132.9, 131.3, 129.7, 126.81, 123.62,
121.3, 117.2, 113.8, 57.5. MS: m/e 271 (M + H+). Anal:
C16H14O4/270.
(E)-3-(4-fluorophenyl)-1-(2-hydroxyphenyl)prop-2-en-1-
one (compound 13) Yellow solid (48.9%), m.p.
105.8–106.9 °C; 1H-NMR (DMSO, 600 MHz) δ: 8.35 (2 H,
d, J = 8.6 Hz, H-2,6), 7.71 (2H, d, J = 8.4 Hz, H-3, 5), 7.64
(1H, d, J = 15 Hz, α-H), 8.13 (1H, d, J = 15 Hz, β-H), 6.30
~ 7.65 (3H, m, Ar’H), 12.74 (1H, s, OH-2′). 13C-NMR
(DMSO, 150 MHz) δ: 194.1, 169.1, 165.7, 145.7, 131.7,
131.5, 129.6, 121.6, 120.6, 115.9, 112.5, 110.4, 107.5. MS:
m/e 243 (M + H+). Anal: C15H11FO2/242.
(E)-1-(2,4-dihydroxyphenyl)-3-(3,4-dihydroxyphenyl)prop-
2-en-1-one (compound 8) Yellow solid (48.89%), m.p.
220.3–221.6 °C; 1H-NMR (DMSO, 600 MHz) δ: 6.27 (1 H,
d, J = 2.4 Hz, H-3′), 6.40 (1H, dd, J1 = 2.4 Hz, J2 = 9 Hz,
H-5′), 6.81 (1H, d, J = 8.4 Hz, H-2), 7.20 (1H, dd, J1 = 1.8
Hz, J2 = 8.2 Hz, H-6), 7.37 (1H, d, J = 1.8 Hz, H-5), 7.65
(1H, d, J = 15.3 Hz, H-α), 7.71 (1H, d, J = 15.3 Hz, H-β),
8.14 (1H, d, J = 9 Hz, 6′), 13.60 (1H, s, OH); 13C-NMR
(DMSO, 150 MHz) δ: 193.3, 167.7, 166.8, 150.8, 147.5,
146.6, 134.7, 128.1, 124.3, 119.2, 117.7, 117.6, 114.9,
110.0, 104.5. MS: m/e 273 (M + H+). Anal: C15H12O5/272.
(E)-1-(2,4-difluorophenyl)-3-(4-hydroxyphenyl)prop-2-en-
1-one (compound 14) Yellow solid (35.4%), m.p.

Med Chem Res
1
104.6–106.7 °C; H-NMR (DMSO, 600 MHz) δ: 8.31 (2H,
153.7, 131.9, 130.2, 127.1, 125.5, 120.3, 113.6, 109.8,
105.6, 105.3, 40.7, 27.3. MS: m/e 298 (M + H+). Anal:
C18H19NO3/297.
d, J = 8.6 Hz, H-2′,6′), 7.70 (2H, d, J = 8.4 Hz, H-3′, 5′),
7.67 (1H, d, J = 15 Hz, α-H), 7.82 (1H, d, J = 15 Hz, β-H),
6.60 ~ 7.65 (3H, m, ArH), 10.03 (1H, s, OH-4). 13C-NMR
(DMSO, 150 MHz) δ: 194.7, 171.6, 169.9, 169.1, 149.5,
146.3, 124.7, 123.6, 126.8, 119.5, 112.1, 110.5, 105.7. MS:
m/e 261 (M + H+). Anal: C15H10F2O2/260.
(E)-1-(2,4-dihydroxyphenyl)-2-methyl-3-phenylprop-2-en-
1-one (compound 20) Yellow solid (65.4%), m.p.
1
113.3–114.8 °C; H-NMR (DMSO, 600 MHz) δ: 12.5 (s,
OH), 7.5 (d, J = 8.7 Hz, H), 7.38–7.34 (m, 4H), 7.21 (s, H),
6.96 (d, J = 16.1 Hz), 6.85 (dd, J = 8.7 Hz, 2.6 Hz, H), 6.5
(d, J = 2.6 Hz), 1.83 (brs, 3H). 13C-NMR (DMSO, 150
MHz) δ: 189.5, 170.1, 163.2, 148.6, 129.8, 129.1, 128.8,
128.4, 126.9, 115.7, 114.7, 104.9, 103.8, 28.4. MS: m/e 255
(M + H+). Anal: C16H14O3/254.
(E)-1-(2,4-difluorophenyl)-3-(4-hydroxy-3-methoxyphenyl)
prop-2-en-1-one (compound 15) Yellow solid (26.4%), m.
p. 110.3–111.5 °C; H-NMR (DMSO, 600 MHz) δ: 3.37
(3H, s, OCH3), 6.43 ~ 6.65 (3H, m, ArH), 6.20 ~ 7.65 (3H,
m, Ar′H), 10.54 (1H, s, OH). 13C-NMR (DMSO, 150 MHz)
δ: 196.1, 163.5, 162.4, 161.7, 159.7, 149.5, 132.5, 132.1,
127.8, 124.5, 116.8, 116.3, 110.5, 103.7, 102.5. 55.9. MS:
m/e 291 (M + H+). Anal: C16H12F2O3/290.
1
(E)-1-(2,4-dihydroxyphenyl)-3-(4-hydroxyphenyl)-2-
methylprop-2-en-1-one (compound 21) Yellow solid
(67.4%), m.p. 103.3–104.9 °C; 1H-NMR (DMSO, 600
MHz) δ: 7.92 (d, J = 8.5 Hz, H), 7.83 (d, J = 7.0 Hz,2H),
7.21 (s, H), 6.89 (d, J = 8.32 Hz, 2H), 6.40 (dd, J = 8.5 Hz,
2.4 Hz, H), 6.32 (d, J = 2.4 Hz, H), 5.32 (brs, H), 2.65 (d, J
= Hz). 13C-NMR (DMSO, 150 MHz) δ: 191.5, 172.3,
170.5, 149.8, 130.5, 130.1, 128.6, 127.5, 120.4, 113.6,
112.7, 109.2, 105.9, 25.1. MS: m/e 271 (M + H+). Anal:
C16H14O4/270.
(E)-1-(2,4-difluorophenyl)-3-(3-hydroxy-4-methoxyphenyl)
prop-2-en-1-one (compound 16) Yellow solid (40.3%), m.
1
p. 110.7–112.0 °C; H-NMR (DMSO, 600 MHz) δ: 3.74
(3H, s, OCH3), 6.45 ~ 6.75 (3H, m, ArH), 6.61 ~ 7.47 (3H,
m, Ar’H), 10.09 (1H, s, OH). 13C-NMR (DMSO, 150 MHz)
δ: 197.5, 163.7, 163.4, 159.7, 157.7, 149.6, 132.6, 131.2,
127.7, 124.5, 116.8, 116.1, 110.5, 102.7, 101.5. 55.5. MS:
m/e 291 (M + H+). Anal: C16H12F2O3/290.
(E)-1-(2,4-dihydroxyphenyl)-2-methyl-3-(4-nitrophenyl)
prop-2-en-1-one (compound 17) Yellow solid (40.3%), m.
p. 162.7–164.1.0 °C; H-NMR (DMSO, 600 MHz) δ: 13.1
(E)-2-methyl-3-(4-nitrophenyl)-1-phenylprop-2-en-1-one
(compound 22) Yellow solid (77.4%), m.p. 103.3–104.9 °
1
1
C; H-NMR (DMSO, 600 MHz) δ: 8.00 (d, J = 7.08 Hz,
(s, OH), 8.10 (d, J = 8.8 Hz, H), 7.82(s, H), 7.73 (d, J = 8.8
Hz, 2H), 6.92 (d, J = 8.8 Hz, 2H), 6.46 (dd, J = 8.8 Hz, 2.0
Hz, 6.36 (d, J = 2.0 Hz, H), 2.25 (brs, 3H). 13C-NMR
(DMSO, 150 MHz) δ: 192.3, 167.6, 165.6, 145.7, 133.1,
132.1, 129.0, 128.6, 118.6, 114.5, 112.5, 108.7, 103.7,
22.5. MS: m/e 300 (M + H+). Anal: C16H13NO5/299.
2H), 7.78 (s, H), 7.55–7.59 (m, 3H), 7.50 (t, J = 7.72 Hz,
2H), 6.89 (d, J = 8.36 Hz, 2H). 2.67 (brs, 3H). 13C-NMR
(DMSO, 150 MHz) δ: 185.6, 158.2, 132.7, 131.8, 131.6,
130.5, 128.7, 127.8, 119.6, 116.2, 110.5, 29.6. MS: m/e 268
(M + H+). Anal: C16H13NO3/267.
(E)-3-(4-fluorophenyl)-2-methyl-1-phenylprop-2-en-1-one
(compound 23) Yellow solid (59.4%), m.p. 100.3–102.9 °
C; 1H-NMR (DMSO, 600 MHz) δ: 7.93(d, J = 7.1 Hz, 2H),
7.6 (s, H), 7.50–7.63 (m, 3H), 7.4 (t, J = 7.2, 2H), 6.75 (d,
J = 8.21 Hz, 2H), 2.81 (s, 6H), 2.32 (brs, 3H). 13C-NMR
(DMSO, 150 MHz) δ: 193.5, 149.8, 133.6, 132.1, 130.7,
129.5, 128.6, 126.4, 120.1, 116.3, 111.5, 21.6. MS: m/e 241
(M + H+). Anal: C16H13FO/240.
(E)-1-(2,4-dihydroxyphenyl)-3-(4-fluorophenyl)-2-methyl-
prop-2-en-1-one (compound 18) Yellow solid (55.3%), m.
p. 176.7–178.1.0 °C; H-NMR (DMSO, 600 MHz) δ: 13.6
(s, OH), 8.33 (d, J = 9.0 Hz, 2H), 7.83 (d, J = 9.0 Hz, H),
7.71 (d, J = 9.0 Hz, 2H), 7.58 (s, H), 5.97 (dd, J = 9.0 Hz,
2.0 Hz, H), 2.13 (brs, 3H). 13C-NMR (DMSO, 150 MHz) δ:
189.7, 173.8, 170.5, 136.9, 135.3, 133.6, 130.6, 127.5,
115.8, 111.9, 110.5, 105.8, 100.1, 25.6. MS: m/e 273 (M +
H+). Anal: C16H13FO3/272.
1
(E)-3-(4-(dimethylamino)phenyl)-2-methyl-1-phenylprop-
2-en-1-one (compound 24) Yellow solid (56.4%), m.p.
85.3–87.9 °C; 1H-NMR (DMSO, 600 MHz) δ: 8.01 (d, J =
6.9 Hz, 2H), 7.8 (s, H), 7.2–7.5 (m, 3H), 6.9 (t, J = 7.0 Hz,
2H), 6.52 (d, J = 7.9 Hz, 2H), 2.50 (brs, 3H). 13C-NMR
(DMSO, 150 MHz) δ: 182.1, 149.6, 134.8, 133.7, 131.1,
129.6, 126.3, 124.9, 120.7, 115.6, 108.9, 41.8, 28.6. MS:
m/e 266 (M + H+). Anal: C16H13NO/265.
(E)-1-(2,4-dihydroxyphenyl)-3-(4-(dimethylamino)phenyl)-
2-methylprop-2-en-1-one (compound 19) Yellow solid
(55.3%), m.p. 89.3–91.8 °C; H-NMR (DMSO, 600 MHz)
δ: 12.,15 (s, OH), 7.9 (s, H), 7.65 (d, J = 8.6 Hz, H), 6.93 (s,
H), 6.83 (d, J = 8.6 Hz, 2H), 6.50 (dd, J = 8.6 Hz, 2.2 Hz,
H), 6.21 (d, J = 2.2 Hz, H), 2.91 (s, 6H), 2.36 (brs, 3H).
13C-NMR (DMSO, 150 MHz) δ: 187.5, 172.3, 159.8,
1

Med Chem Res
(E)-2-methyl-1,3-diphenylprop-2-en-1-one(compound
25) Yellow solid (66.4%), m.p. 115.3–117.9 °C; 1H-NMR
(DMSO, 600 MHz) δ: 8.01–8.03 (m, 2H), 7.81 (s, 1H),
7.63–7.65 (m, 2H), 7.55–7.60 (m, 2H), 7.48–7.52 (m, 2H),
7.39–7.42 (m, 2H), 2.31 (brs, 3H). 13C NMR (DMSO, 150
MHz): δ 190.52, 144.81, 138.16, 134.83, 132.76,130.52,
128.93, 128.59, 128.47, 128.41, 122.02, 27.8. MS: m/e 223
(M + H+). Anal: C16H14O/222.
126.9, 121.9, 120.1, 27.8. MS: m/e 239 (M + H+). Anal:
C14H16O2/238.
(E)-1,3-bis(4-hydroxyphenyl)-2-methylprop-2-en-1-one
(compound 31) Yellow solid (43.3%), m.p. 123.1–124.9 °
C; 1H-NMR (DMSO, 600 MHz): δ 8.30 (d, J = 8.7 Hz, 2H),
8.21 (s, H), 8.02 (d, J = 7.3Hz, 2H), 6.73 (d, J = 8.0, 2H),
5.6 (d, J = 8.6 Hz, 2H), 2.76 (brs, 3H). 13C NMR (DMSO,
150 MHz) δ: 187.5, 169.2, 158.6, 135.6, 132.7, 131.6,
129.3, 127.3, 121.8, 121.5, 110.8, 26.3. MS: m/e 255 (M +
H+). Anal: C16H14O3/254.
(E)-3-(4-hydroxyphenyl)-2-methyl-1-phenylprop-2-en-1-
one(compound 26) Yellow solid (66.4%), m.p. 95.2–97.8 °C;
1H-NMR (DMSO, 600 MHz) δ: 8.02 (d, J = 7.3 Hz, 2H), 7.91
(t, J = 7.3 Hz, 2H), 7.42–7.86 (m, 3H), 7.01 (s, H), 6.73 (d, J
= 8.0 Hz, 2H), 2.42 (brs, 3H). 13C NMR (DMSO, 150 MHz):
δ 190.2, 149.8, 135.6, 134.2, 133.8, 130.6, 128.1, 127.9, 120.1,
115.8, 110.6, 22.3. MS: m/e 239 (M + H+). Anal: C16H14O2/
238.
Biological
Cell culture
HeLa, SiHa, and CHO cells were grown in Dulbecco’s
Modified Eagle Media. The media were supplemented with
100 U/mL penicillin, 100 mg/mL streptomycin and 10%
fetal bovine serum. Cells were incubated at 37 °C in a
humidified atmosphere containing 5% CO2.
(E)-1-(4-hydroxyphenyl)-2-methyl-3-(4-nitrophenyl)prop-
2-en-1-one (compound 27) Yellow solid (26.4%), m.p.
91.3–93.9 °C; 1H-NMR (DMSO, 600 MHz) δ: 8.04 (d, J =
8.8 Hz, 2H), 7.95 (s, H), 7.70 (d, J = 8.6 Hz, 2H), 7.56 (d, J
= 8.3 Hz, 2H), 7.01 (d, J = 8.6 Hz, 2H), 6.83 (d, J = 9.3 Hz,
2H), 6.02 (d, J = 9.0 Hz, 2H), 5.409 (s, OH), 2.56 (brs, 3H).
13C NMR (DMSO,150 MHz): δ 190.5, 161.6, 150.8, 132.8,
131.2, 129.0, 119.2, 115.3, 24.7. MS: m/e 284 (M + H+).
Anal: C16H13NO4/283.
In vitro anti-proliferative assay
Cells were seeded into 96-well plates (5000 cells/well) and
incubated at 37 °C in a humidified 5% CO2 atmosphere.
After 24 h, the cells were treated with different concentra-
tions of the compounds. Concentrated (5 mg) solutions of
chalcones were prepared in DMSO. Cell viability after 24,
48, and 72 h of drug exposure was determined using the
MTT cell viability assay, as described previously.
(E)-3-(4-fluorophenyl)-1-(4-hydroxyphenyl)-2-methylprop-
2-en-1-one (compound 28) Yellow solid (86.3%), m.p.
90.3–92.9 °C; 1H-NMR (DMSO, 600 MHz): δ 8.60 (d, J =
7.6 Hz, 2H), 8.24 (s, H), 7.92 (d, J = 9.1 Hz, 2H), 6.52 (d, J
= 8.2 Hz, 2H), 5.64 (d, J = 9.1 Hz, 2H), 2.35 (brs, 3H). 13
C
Apoptosis assay
NMR (DMSO,150 MHz):δ: 193.1, 165.8, 153.2, 132.7,
130.5, 129.6, 128.6, 123.9, 120.5, 117.8, 112.4, 26.5. MS:
m/e 257 (M + H+). Anal: C16H13FO2/256.
The SiHa and HeLa cells were treated with various con-
centrations (0, 1, 2, 4 μg mL−1) of compound 19 for 24 h
and then stained with both Annexin V-FITC and PI 50.
Then samples were analyzed with an EPICS XL/XL-MCL
flow cytometer (USA Beckmann).
(E)-3-(4-(dimethylamino)phenyl)-1-(4-hydroxyphenyl)-2-
methylprop-2-en-1-one (compound 29) Yellow solid
(62.3%), m.p. 121.3–122.9 °C; 1H-NMR (DMSO, 600
MHz): δ 8.50 (d, J = 7.9 Hz, 2H), 8.34 (s, H), 8.02 (d, J =
8.6 Hz, 2H), 6.53 (d, J = 8.2 Hz, 2H), 2.98 (brs, 3H), 2.54
(s, 6H). 13C NMR (DMSO,150 MHz) δ: 187.6, 165.9,
154.9, 133.1, 130.5, 128.7, 127.5, 124.6, 121.7, 118.2,
107.8, 45.6, 25.7. MS: m/e 282 (M + H+). Anal:
C18H19NO2/281.
Docking simulation
The molecule was docked to the crystal structure of the
yeast 20S proteasome in complex with the inhibitor
homobelactosin C covalently bound to the terminal threo-
nine of the β5 active site [PDB code: 3E47]. The structures
were retrieved from the PDB archive (www.pdb.org), and
the docking simulation has been performed using the MOE/
Dock procedure integrated in the AutoDock Tools
1.5.6 system of programs (Chemical Computing Group).
During the first step of the docking application, the ligand
was treated in a flexible manner by rotating routable bonds.
Several configurations were generated and scored in an
(E)-1-(4-hydroxyphenyl)-2-methyl-3-phenylprop-2-en-1-
one(compound 30) Yellow solid (62.3%), m.p. 92.1–92.9
1
°C; H-NMR (DMSO, 600 MHz): δ 8.51 (d, J = 8.3 Hz,
2H), 8.3 (s, H), 8.15 (d, J = 8.1 Hz, 2H), 6.49 (d, J = 8.2
Hz, 2H), 5.3 (d, J = 8.2 Hz, 2H), 2.65 (brs, 3H). 13C NMR
(DMSO,150 MHz) δ: 191.6, 167.2, 159.7, 133.8, 129.1,

Med Chem Res
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Acknowledgements This research work was financially supported
by Natural National Science Foundation of China (81260379). We
would like to thank Professor Mourboul for helping us with the
synthesis, biological evaluation and English language editing.
Compliance with ethical standards
Groll M, Oleg V, Huber R, Meijere A (2006) Inhibitor-binding mode
of homobelactosin C to proteasomes: new insights into class I
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proliferation of human breast cancer cell by blocking cell cycle
progression and inducing apoptosis. Food Chem Toxicol
44:704–713
Conflict of interest The authors declare that they have no competing
interests.
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