Design, synthesis and biological evaluation of E-ring modified evodiamine derivatives as novel antitumor agents

Article abstract of DOI:10.1016/j.cclet.2014.03.043

A series of novel E-ring modified evodiamine derivatives were designed and synthesized as antitumor agents. Their capacity to interfere with the catalytic activity of topoisomerase I and II was evaluated by the relaxation assay. In vitro antitumor activity results revealed that compound 12 showed good antitumor activity with a broad spectrum. Its binding modes with topoisomerase I and II were clarified by molecular docking.

Full text of DOI:10.1016/j.cclet.2014.03.043

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Chinese Chemical Letters xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
Original article
Design, synthesis and biological evaluation of E-ring modified
evodiamine derivatives as novel antitumor agents
Kun Fang a,1, Guo-Qiang Dong a,1, Hai Gong b,1, Na Liu a, Zhen-Gang Li a, Shi-Ping Zhu a,
Zhen-Yuan Miao a, Jian-Zhong Yao a, Wan-Nian Zhang a, Chun-Quan Sheng a,
*
a School of Pharmacy, Second Military Medical University, Shanghai 200433, China
b Department of Radiation Oncology, Ji’nan Military General Hospital, Jinan 250031, China
A R T I C L E I N F O
A B S T R A C T
Article history:
A series of novel E-ring modified evodiamine derivatives were designed and synthesized as antitumor
agents. Their capacity to interfere with the catalytic activity of topoisomerase I and II was evaluated by
the relaxation assay. In vitro antitumor activity results revealed that compound 12 showed good
antitumor activity with a broad spectrum. Its binding modes with topoisomerase I and II were clarified
by molecular docking.
Received 4 December 2013
Received in revised form 6 March 2014
Accepted 7 March 2014
Available online xxx
ß 2014 Chun-Quan Sheng. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights
reserved.
Keywords:
Evodiamine derivatives
Topoisomerase I
Topoisomerase II
Antitumor activity
1. Introduction
identified as a Top1 inhibitor by structure-based virtual screening
[7]. Further structural optimization studies identified several
The use of effective antitumor agents derived from natural
products is a promising strategy for cancer chemotherapy [1]. It is
reported that more than 50% of antitumor drugs have been
developed from natural products or their synthetic derivatives
[2,3]. Antitumor natural products also provide novel probes for
chemical biology and target identification studies. For example,
camptothecin (target: DNA topoisomerase I, Top1) and taxol
(target: microtubule) represent two historic achievements in
antitumor drug discovery [4].
highly active evodiamine derivatives which showed potent in vitro
and in vivo antitumor activities [8]. In silico target identification
and biological assays revealed that the evodiamine derivatives
were dual inhibitors of Top1 and topoisomerase II (Top2) [8].
Inspired by these encouraging results, herein a series of E-ring
modified evodiamine derivatives were designed and synthesized.
Compound 12 showed good antitumor activity with a broad
spectrum. Its binding modes with topoisomerase I and II were
clarified by molecular docking.
Evodiamine is a quinazolinocarboline alkaloid with diverse
biological activities which was isolated from the fruits of the
traditional Chinese herb Evodiae fructus (Chinese name: Wu-Chu-
Yu) [5]. In particular, evodiamine showed anti-proliferative
activities against a wide variety of cancer cells by inducing
apoptosis [6]. It is a good antitumor lead compound because of its
broad-antitumor spectrum, low toxicity, and suitable physico-
chemical properties. In our previous studies, evodiamine was
2. Experimental
Nuclear magnetic resonance (NMR) spectra were recorded on
Bruker AVANCE300 and AVANCE500 spectrometers (Bruker
Company, Germany), with TMS as an internal standard and
DMSO-d6 as the solvent. Chemical shifts (d values) and coupling
constants (J values) are expressed in ppm and Hz, respectively. ESI
mass spectra were performed on an API-3000 LC–MS spectrome-
ter. TLC analysis was carried out on silica gel plates GF254 (Qindao
Haiyang Chemical, China). Silica gel column chromatography was
performed with Silica gel 60G (Qindao Haiyang Chemical, China).
Commercial solvents were used without any pretreatment.
*
Corresponding author.
E-mail address: shengcq@hotmail.com (C.-Q. Sheng).
These authors contributed equally to this article.
1
http://dx.doi.org/10.1016/j.cclet.2014.03.043
1001-8417/ß 2014 Chun-Quan Sheng. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Please cite this article in press as: K. Fang, et al., Design, synthesis and biological evaluation of E-ring modified evodiamine derivatives as
novel antitumor agents, Chin. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.cclet.2014.03.043

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K. Fang et al. / Chinese Chemical Letters xxx (2014) xxx–xxx
Scheme 1. Synthetic routes of the target compounds. Reagents and conditions: (a) Ethyl formate, reflux, 6 h, yield 100%; (b) POCl3, CH2Cl2, 0 8C, 6 h, yield 85%; (c) Triphosgene,
THF, reflux, 4 h, yield 90%-95%; (d) CH3I, KOH, DMF, rt, 2 h, yield 60–87%; (e) CH2Cl2, 45 8C, 6 h, yield 50%-62%; (f) 10% Pd/C, H2, DMF, r.t., 12 h, yield 11–73%; (g) 2-bromoacetyl
bromide, Et3N, CH2Cl2, 0 8C, 1 h, yield 84%; (h) Na2SÁ9H2O, DMF, r.t., 12 h, yield 70%; (i) 2-(1-piperazinyl)pyridine-4-boronic acid pinacol ester, Pd(PPh3)4, Na2CO3, H2O, reflux,
3.5 h, yield 51%.
2.1. Chemistry
(m, 3H), 7.35 (d, 1H, J = 7.5 Hz), 7.43 (d, 1H, J = 7.5 Hz), 8.37 (s, 1H),
10.90 (s, 1H). ESI-MS: Calcd. for C19H14ClN4O3 [MÀH]À: m/z
381.79; found: 381.81.
The synthetic route of E-ring modified evodiamine derivatives is
describedinScheme1. 3,4-Dihydro-
b
-carboline(4)waspreparedby
3-Iodoevodiamine (8c): White solid (0.07 g, yield: 50%).
reacting tryptamine 2 with ethyl formate, followed by intra-
molecular ring closure in the presence of POCl3. Substituted 2-
aminobenzoic acid (5) was treated with triphosgene to provide
isatoic anhydride, which was methylated by CH3I to afford
1H NMR (500 MHz, DMSO-d6):
d 2.75–2.78 (m, 1H), 2.91–2.92
(m, 1H), 2.96 (s, 3H), 3.19–3.21 (m, 1H), 4.58–4.62 (m, 1H), 6.17
(s, 1H), 6.87 (d, 1H, J = 8.6 Hz), 7.01 (t, 1H, J = 7.6 Hz), 7.11 (t, 1H,
J = 7.6 Hz), 7.35 (d, 1H, J = 8.1 Hz), 7.46 (d, 1H, J = 8.1 Hz), 7.45
(dd, 1H, J = 8.6 Hz, 2.1 Hz), 7.98 (d, 1H, J = 2.1 Hz), 11.03 (s, 1H).
ESI-MS: Calcd. for C19H17IN3O [M+H]+: m/z 430.26; found: 430.39.
Synthesis of 3-aminoevodiamine (9a): To a stirring solution of
8a (0.12 g, 0.34 mmol) in DMF (10 mL), 10% Pd/C (0.01 g) was
added. The reaction mixture was stirred under hydrogen atmo-
sphere for 12 h at room temperature. Then the mixture was diluted
with water (50 mL) and extracted with EtOAc (3 Â 50 mL). The
combined organic layers were washed with saturated sodium
chloride solution (3 Â 50 mL), dried over anhydrous Na2SO4 and
concentrated under reduced pressure. The residue was purified by
silica gel column chromatography (CH2Cl2–MeOH, gradient 100:1,
100:2) to give compound 9a (0.08 g, 73%) as a yellow solid. 1H NMR
compound 7 [8]. Condensation of 3,4-dihydro-b-carboline (4) with
N-methyl isatoic anhydride 7 provided E-ring derivatives 8a–c.
Reductionof the nitro groupof compounds 8a and 8b using10% Pd/C
and H2 afforded the amino derivatives 9a and 9b. Compound 9a was
further acylated to give compound 10, which was subsequently
treated with Na2S to provide compound 11. The Suzuki-Miyaura
reaction of compound 8c with 2-(1-piperazinyl)pyridine-4-boronic
acid pinacol ester afforded the target compound 12.
Synthesis of 3-nitroevodiamine (8a): A solution of 3,4-dihydro-
b
-carboline [8] (4, 0.17 g, 1 mmol) and 1-methyl-6-nitro-1H-
benzo[d][1,3]oxazine-2,4-dione [8] (7a, 0.2 g, 1 mmol) in CH2Cl2
(15 mL) were heated to 45 8C and stirred for 6 h. Then the reaction
mixture was concentrated to one-half of its original volume. The
precipitate was collected by filtration and washed with alcohol to
yield the product 8a (0.21 g, 62%) as yellow solid. 1H NMR
(500 MHz, DMSO-d6):
d 2.27 (s, 3H), 2.77–2.79 (m, 1H), 2.88–2.92
(m, 1H), 3.11–3.13 (m, 1H), 4.63–4.67 (m, 1H), 5.10 (s, 2H), 5.90
(s, 1H), 6.78 (dd, 1H, J = 8.5 Hz, 2.5 Hz), 6.94 (d, 1H, J = 8.5 Hz), 7.01
(t, 1H, J = 7.5 Hz), 7.10–7.14 (m, 2H), 7.36 (d, 1H, J = 8.0 Hz), 7.51
(d, 1H, J = 8.0 Hz), 11.26 (s, 1H). ESI-MS: Calcd. for C19H19N4O
[M+H]+: m/z 319.15; found: 319.93.
The synthetic method for target compounds 9b was similar to
that of compound 9a.
2-Chloro-3-aminoevodiamine (9b): Red solid (0.09 g, yield:
11%). 1H NMR (500 MHz, DMSO-d6):
d 2.27 (s, 3H), 2.73–2.77 (m,
(300 MHz, DMSO-d6): d 2.68–2.70 (m, 1 H), 2.95–3.01 (m, 2H), 3.45
(s, 3H), 4.60–4.64 (m, 1H), 6.50 (s, 1H), 6.96–7.11 (m, 4H), 7.32 (d,
1H, J = 7.8 Hz), 7.42 (d, 1H, J = 7.8 Hz), 8.24 (dd, 1H, J = 9.0 Hz,
2.7 Hz), 8.43 (d, 1H, J = 2.7 Hz), 10.89 (s, 1H). ESI-MS: Calcd. for
C
19H16N4O3 [MÀH]À: m/z 347.12; found: 347.76.
The synthetic method for target compounds 8b–c was similar to
that of compound 8a.
2-Chloro-3-nitroevodiamine (8b): Yellow solid (0.33 g, yield:
2H), 3.11–3.13 (m, 1H), 4.63–4.66 (m, 1H), 5.10 (s, 2H), 5.90 (s, 1H),
6.78 (dd, 1H, J = 8.5 Hz, 2.5 Hz), 6.94 (d, 1H, J = 8.5 Hz), 7.01 (t, 1H,
J = 7.5 Hz), 7.10–7.14 (m, 2H), 7.36 (d, 1H, J = 8.0 Hz), 11.26 (s, 1H).
50%). 1H NMR (300 MHz, DMSO-d6):
d
2.68–2.73 (m, 1H), 2.95–3.01
(m, 2H), 3.46 (s, 3H), 4.61–4.67 (m, 1H), 6.52 (s, 1H), 7.00–7.16
Please cite this article in press as: K. Fang, et al., Design, synthesis and biological evaluation of E-ring modified evodiamine derivatives as
novel antitumor agents, Chin. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.cclet.2014.03.043

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3
ESI-MS: Calcd. for C19H18ClN4O [M+H]+: m/z 353.83; found:
353.49.
Top2a-mediated supercoiled pBR322 relaxation assay: Relaxa-
tion assays were performed using Topoisomerase II Drug Screening
Kits (TopoGEN, Inc.) [8]. The reaction buffer contained 50 mmol/L
Tris–HCl (pH 8.0), 150 mmol/L NaCl, 10 mmol/L MgCl2, 5 mmol/L
Synthesis of 2-bromoacetamidoevodiamine (10): To a stirring
solution of 9a (0.1 g, 0.3 mmol) and Et3N (0.04 mL, 0.3 mmol) in
CH2Cl2 (20 mL), 2-bromoacetyl bromide (0.04 mL, 0.47 mmol) was
added and stirred for 1 h at 0 8C. The solvent was removed under
reduced pressure. The residue was purified by silica gel column
chromatography (CH2Cl2:MeOH = 100:1, v/v) to afford 10 (0.11 g,
dithiothreitol, 30
ATP, 0.25 g pBR322 plasmid DNA, and 0.75 unit of Top2
(TopoGEN, Inc.) in a total volume of 20 L. After incubation at 37 8C
for 30 min, the reaction was terminated by the addition of 2
mg/mL bovine serum albumin (BSA), 2 mmol/L
m
a
m
m
L
yield 84%) as yellow solid. 1H NMR (300 MHz, DMSO-d6):
d
2.69 (s,
10Â gel loading buffer (0.25% bromophenol blue, 50% glycerol).
The reaction products were analyzed on 1% agarose gel at 8 V/cm
for 1 h with TAE (Tris–acetate–EDTA) as the running buffer. Gels
3H), 2.85 (m, 2H), 3.15 (m, 1H), 4.01 (s, 2H), 4.62 (m, 1H), 6.07 (s,
1H), 7.00 (t, 1H, J = 7.8 Hz), 7.08–7.13 (m, 2H), 7.35 (d, 1H,
J = 7.8 Hz), 7.48 (d, 1H, J = 7.8 Hz), 7.73 (dd, 1H, J = 9.0 Hz, 2.4 Hz),
8.03 (d, 1H, J = 2.7 Hz), 10.41 (s, 1H), 11.16 (s, 1H). ESI-MS: Calcd.
for C21H19BrN4O2 [M+H]+: m/z 439.07; found: 439.75.
were stained with ethidium bromide (0.5
mg/mL) for 60 min. The
DNA band was visualized over UV light and photographed with Gel
Doc Ez imager software (Bio-Rad Laboratories Ltd.).
Synthesis of 2-mercaptoacetamidoevodiamine (11): A solu-
tion of 10 (0.05 g, 0.11 mmol) and Na2SÁ9H2O (0.27 g, 1.1 mmol) in
DMF (5 mL) was stirred at room temperature for 12 h. The solvent
was removed in vacuo and the residue was diluted with water and
filtered from insolubles. The filtrate was acidified to pH 1.2 with
3 mol/L HCl. The precipitate was collected and purified by silica
gel column chromatography (CH2Cl2–MeOH, gradient 100:1–
100:3) to give compound 11 (0.03 g, yield 70%) as a yellow solid.
In vitro cytotoxicity assay: Cells were seeded in 96-well
microtiter plates at a density of 5 Â 103/well and treated in
triplicate with gradient concentrations of compounds in
a
humidified atmosphere with 5% CO2 at 37 8C for 72 h. 20
mL of
MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bro-
mide) solution (5 mg/mL) was added to each well and the plate
was incubated for an additional 4 h. The formazan was dissolved in
100
mL of DMSO. The absorbance (OD) was read on a WellscanMK-
1H NMR (300 MHz, DMSO-d6):
d
1.30 (s, 1 H), 2.66 (s, 3H), 2.86
2 microplate reader (Labsystems) at 570 nm. The concentration
causing 50% inhibition of cell growth (IC50) was determined by the
Logit method [9,10]. All experiments were performed three times.
(m, 2H), 3.20 (m, 1H), 3.48 (s, 2H), 4.64 (m, 1H), 6.06 (s, 1H), 7.01
(t, 1H, J = 7.2 Hz), 7.10–7.15 (m, 2H), 7.36 (d, 1H, J = 8.1 Hz), 7.50
(d, 1H, J = 8.1 Hz), 7.73 (d, 1H, J = 9.3 Hz), 8.07 (s, 1H), 10.19 (s, 1H),
11.19 (s, 1H). ESI-MS: Calcd. for C21H20N4O2S [M + H]+: m/z
393.14; found: 393.84.
2.3. Molecular docking
Synthesis of 3-(2-(piperazin-1-yl)pyridin-4-yl)evodiamine
(12): Under an nitrogen atomosphere, 8c (0.1 g, 0.23 mmol), 2-
(1-piperazinyl)pyridine-4-boronic acid pinacol ester (0.1 g,
0.35 mmol) and tetrakistriphenylphosphine palladium (40 mg)
were dissolved in DMF (5 mL). An aqueous sodium carbonate
solution (2 mol/L, 1 mL) was added and the mixture was heated
under reflux for 3.5 h. After cooled to room temperature, the
reaction mixture was diluted with water (50 mL) and then
extracted with EtOAc (3 Â 50 mL). The combined organic layers
were washed with saturated sodium chloride solution (3 Â 50 mL),
dried over anhydrous Na2SO4 and concentrated under reduced
pressure. The residue was purified by silica gel column chroma-
tography (CH2Cl2:MeOH = 100:1, v/v) to givecompound 12 (0.056 g,
In silico docking was performed using GOLD 5.1 [11] on a Linux
PC as described before [8]. The X-ray crystallographic structures of
CPT-DNA-Top1 ternary complex (PDB code: 1T8I [12]) and ATPase
domain of humanTop2a(PDBcode:1ZXM[13]) were obtained from
the Protein Data Bank. They were prepared for molecular docking
analysisinDiscoveryStudio3.0[14]byremovingtheligandfromthe
binding pocket, merging nonpolar hydrogens, adding polar hydro-
gens, and rendering Gasteiger charges to each atom.
3. Results and discussion
3.1. In vitro antitumor activity
yield 51%) as a yellow solid. 1H NMR (300 MHz, DMSO-d6):
d 2.15
The growth inhibitory activities of the target compounds toward
human cancer cell lines A549 (lung cancer), MDA-MB-435 (breast
cancer), and HCT116 (colon cancer) were determined using MTT
assay. Evodiamine was used as the reference drug. When the 2-
(s, 3H), 2.82 (m, 1H), 3.01 (m, 1H), 3.16 (m, 1H), 3.74 (s, 3H), 3.81
(s, 3H), 4.64 (m, 1H), 6.25 (s, 1H), 6.85 (d, 1H, J = 9.3 Hz), 6.92 (s, 1H),
6.99 (t, 1H, J = 7.8 Hz), 7.05–7.11 (m, 2H), 7.34 (d, 1H, J = 7.8 Hz),
7.44 (d, 1H, J = 7.8 Hz), 7.85 (dd, 1H, J = 8.7 Hz, 2.1 Hz), 8.04 (d, 1H,
J = 2.1 Hz), 8.09 (d, 1H, J = 5.1 Hz), 10.99 (s, 1H). ESI-MS: Calcd.
for C28H28N6O [M+H]+: m/z 465.24; found: 465.84.
bromoacetamide group was attached on the position
evodiamine E-ring, compound 10 showed moderate in vitro
antitumor activity against MDA-MB-435 (IC50 = 22.69 mol/L),
A549 (IC50 = 153.97 mol/L) and HCT 116 (IC50 = 28 mol/L) cell
3 of
m
m
m
2.2. Biological assay
lines (Table 1). However, the loss of the antitumor activity was
observed for the corresponding thio-derivative 11. 2-Chloro-3-nitro
evodiamine (8b) also showed moderate antitumor activity against
all three of the tested cancer cell lines. When its nitro group was
reduced to the amino group, compound 9b showed significantly
Top1-mediated supercoiled pBR322 relaxation assay: Relaxa-
tion assays were carried out as previously described [7,8]. The
reaction buffer contained 35 mmol/L Tris–HCl (pH 8.0), 72 mmol/L
KCl, 5 mmol/L MgCl2, 5 mmol/L dithiothreitol, 5 mmol/L spermi-
dine, 0.1% bovine serum albumin (BSA), 0.25
DNA, and 1 unit of Top1 (TaKaRa Biotechnology Co., Ltd., Dalian) in
a total volume of 20 L. After incubation at 37 8C for 15 min, the
reaction was stopped by the addition of 2
(0.9% sodium dodecyl sulfate (SDS), 0.05% bromophenol blue, and
50% glycerol). Then, the DNA samples were subjected to
electrophoresis in 0.8% agarose gel in TAE (Tris–acetate–EDTA)
at 8 V/cm for 1 h. Gels were stained with ethidium bromide
mg pBR322 plasmid
Table 1
In vitro antitumor activity of evodiamine derivatives (IC50, mmol/L).
m
Compounds
Structure
IC50 (mmol/L)
m
L of 10Â loading buffer
MDA-MB-435
A549
204.15
HCT116
8b
31.37
1.35
35.97
75.19
28.00
>250
9b
>250
153.97
>250
6.80
100.00
10
22.69
>250
2.78
11
(0.5
mg/mL) for 60 min. The DNA band was visualized over UV light
12
5.94
and photographed with Gel Doc Ez imager software (Bio-Rad
Laboratories Ltd.).
Evodiamine
20.20
100.00
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K. Fang et al. / Chinese Chemical Letters xxx (2014) xxx–xxx
improved activity against the MDA-MB-435 cell line
(IC50 = 1.35 mol/L). Interestingly, the compound with a 2-(piper-
m
azin-1-yl)pyridyl group at the position 3 of evodiamine E-ring
(compound 12) showed good antitumor activity with a broad
spectrum. The IC50 values of compound 12 against MDA-MB-435,
A549 and HCT 116 cell lines were 2.78
mmol/L, 6.80 mmol/L and
5.94 mol/L, respectively. As compared with evodiamine, most of
m
the target compounds showed better antitumor activity and broader
spectra.
3.2. Top1/Top2 inhibitory activity
Human Top2 can be classified into two subgroups, Top2
Top2 . Top2 increases its expression during the cell cycle and
mitosis, and its concentration is much higher in tumor cells. Top2
expression remains relatively constant and at low level throughout
the cellcycle [15,16]. Thus, Top2 isa good target forantitumor drug
discovery. Previously, evodiamine and its derivatives were identi-
fied as dual Top1/Top2 inhibitors [8]. Herein, Top1- and Top2
mediated pBR322 DNA relaxation assays with purified Top1 and
Top2 were used to investigate the inhibitory activity of compound
12 with its concentrations from 10 mol/L to 100 mol/L.
a and
b
a
b
a
a-
Fig. 1. Top1 and Top2 inhibitory activity of compound 12. (A) Inhibition of Top1
relaxation activity ranging from 10 mol/L to 100 mol/L. Lane 1, supercoiled
plasmid DNA; Lane 2, DNA + Top1; Lanes 3–5, DNA + Top1 + CPT (100, 50 and
10 mol/L); Lanes 6–8, DNA + Top1 + 12 (100, 50 and 10 mol/L). (B) Inhibition of
Top2 relaxation activity ranging from 10 mol/L to 100 mol/L. Lane 1, supercoiled
plasmid DNA; Lane 2, DNA + Top2; Lanes 3–5, DNA + Top1 + Eto (100, 50 and
10 mol/L); Lanes 6–8, DNA + Top2 + 12 (100, 50 and 10 mol/L). Lanes order from
left to right.
a
m
m
m
m
Camptothecin (CPT) and etoposide (Eto) were employed as positive
controls. As shown in Fig. 1A, compound 12 inhibited Top1-
mediated relaxation of pBR322 DNA in a concentration-dependent
m
m
m
m
m
m
manner. It showed moderate inhibitory activity at 100
lower activity at 50 mol/L. The inhibitory activity of compound 12
was much higher than evodiamine, which only showed marginal
activity at 100 mol/L [7]. Compound 12 was also proven to be a
potent inhibitor of the catalytic activity of Top2 (Fig. 1B).
Particularly, it showed comparable inhibitory activity against
Top2-mediated DNA relaxation to that of Eto at 100 mol/L and
50 mol/L, respectively.
mmol/L and
m
m
a
observed to be well-fitted into the ATP-binding domain of Top2
a
m
(Fig. 2B). It is capable of making four hydrogen-bonds to enhance
the stabilization of the 12-ATPase complex. The N atom of pyridine
is shown to participate in hydrogen bonding interactions with
Ala167 and Lys168, respectively. The amino group on the
piperazine also can form two hydrogen bonds with backbone
amide of Arg162 and Asn163. ChemPLP score was also calculated
to evaluate the binding affinity. The most active compound
m
3.3. Binding mode with Top1/Top2
In an attempt to understand the binding mode of compound 12
to Top1-DNA complex and ATP-binding domain of Top2 , docking
a
simulation was performed. As indicated in the interaction model
12 showed the highest binding affinity to Top1 and Top2a with
between 12 and Top1-DNA complex (Fig. 2A), its E-ring and pyridyl
group intercalates at the site of DNA cleavage and forms p-stacking
ChemPLP sore 82.54 and 61.00, respectively. In contrast, removal
of the piperazine group led to the decrease of the binding affinity
interactions with both the À1 (upstream) and +1 (downstream)
base pairs. The N atom of pyridine forms a hydrogen bond with the
hydroxyl group of Thr718. AB-ring is faced into the major groove
and its indole NH undergoes a strong hydrogen bond interaction
with the oxygen atom of Guanine. Moreover, compound 12 is
(ChemPLP sore: 68.04 for Top1 and 54.48 for Top2
showed the lowest ChemPLP scores (Top1: 64.45, Top2
The computational results were consistent with the biological
activities and highlighted the importance of the pyridine and
a
). Evodiamine
a
: 53.34).
piperazine group for Top1 and Top2a binding.
Fig. 2. Binding mode with Top1/Top2. (A) Binding mode of 12 to Top1–DNA complex. (B) Binding mode of 12 to ATP-binding domain of Top2. The figure was generated using
PyMol (http://www.pymol.org/).
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5
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In summary, a series of novel E-ring modified evodiamine
derivatives were designed and synthesized as antitumor agents.
Top1/Top2 inhibitory assay and further molecular docking
confirmed that the derivatives acted by dual inhibition of Top1
and Top2. As compared with evodiamine, most of the compounds
showed better antitumor activity and broader spectra. Compound
12 showed good antitumor activity with an IC50 ranging from
2.78
mmol/L to 5.94 mmol/L. It can be used as a good starting point
for further optimization.
Acknowledgments
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This work was supported by National Natural Science Founda-
tion of China (Nos. 81222044, 81373278), Key Project of Science
and Technology of Shanghai (No. 11431920402), the National Basic
Research Program of China (No. 2014CB541800), Shanghai Rising-
Star Program (No. 12QH1402600), and Shanghai Municipal Health
Bureau (No. XYQ2011038).
[14] Accelrys Software Inc., Discovery Studio Modeling Environment, Release 3.0,
Accelrys Software Inc., San Diego, 2010.
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Please cite this article in press as: K. Fang, et al., Design, synthesis and biological evaluation of E-ring modified evodiamine derivatives as
novel antitumor agents, Chin. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.cclet.2014.03.043