Synthesis of macrocyclic bisbibenzyl derivatives and their anticancer effects as anti-tubulin agents

Article abstract of DOI:10.1016/j.bmc.2012.02.004

Based on the core skeleton of the total synthesized bisbibenzyl marchantin C, riccardin D and plagiochin E, a series of brominated and aminomethylated derivatives of above three bisbibenzyls have been synthesized and their cytotoxic activity against KB, M

Full text of DOI:10.1016/j.bmc.2012.02.004

Bioorganic & Medicinal Chemistry 20 (2012) 2382–2391
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis of macrocyclic bisbibenzyl derivatives and their anticancer effects
as anti-tubulin agents
Juan Jiang a, , Bin Sun a,b, , Yan-yan Wang a, Min Cui c, Li Zhang a, Chang-zhi Cui a, Yan-feng Wang a,
Xi-gong Liu a, Hong-xiang Lou a,
⇑
a School of Pharmaceutical Sciences, Shandong University, Jinan 250012, PR China
b National Glycoengineering Research Center, Shandong University, Jinan 250012, PR China
c School of Oceanology, Shandong University, Weihai 264209, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Based on the core skeleton of the total synthesized bisbibenzyl marchantin C, riccardin D and plagiochin
E, a series of brominated and aminomethylated derivatives of above three bisbibenzyls have been synthe-
sized and their cytotoxic activity against KB, MCF-7 and PC3 cell lines has been preliminary evaluated.
The bio-test results revealed that the brominated derivatives 21, 22, 24, 25 and 28 exhibited excellent
antiproliferative activity, with IC50 value lower than their parent compounds. As a most potent microtu-
bule depolymerization agent, compound 28 was found to arrest cells at the G2/M phase of the cell cycle as
determined by the flow cytometry assay in PC3 cell line. The remarkable biological profile and novel
structure of these bisbibenzyl derivatives make them possible as promising candidates for clinical devel-
opment as chemotherapeutic agents.
Received 22 November 2011
Revised 30 January 2012
Accepted 1 February 2012
Available online 9 February 2012
Keywords:
Bisbibenzyls
Derivatives
Microtubule inhibitor
Anticancer
Ó 2012 Elsevier Ltd. All rights reserved.
1. Introduction
can block mitosis progress and induce apoptosis of cancer cell
in vivo and in vitro by interfering the microtubule polymeriza-
Microtubules are cytoskeletal protein polymers formed mainly
by dynamic assemblies of tubulin heterodimers.1 They play crucial
roles in mitosis and cell division and are recognized as important
targets for anticancer therapy. The anticancer agents targeting on
microtubules can be divided into two major groups: microtubule
stabilizers like taxanes and microtubule destabilizers such as com-
bretastatin A-4, colchicine, and vinca alkaloids.2 Some of them
(taxanes and vinca alkaloids) have been widely used in the clinical
treatment of diverse human cancers for decades. However, these
potent drugs still exhibit substantial limitations, such as low bio-
availability, systemic toxicity, drug resistance, complex syntheses,
and isolation procedures,3 encouraging scientists to develop novel
antimitotic agents for cancer therapy.
Bisbibenzyls are a series of phenolic natural products that are
found exclusively in bryophytes. These natural products exhibit
versatile biological activities,4–13 including 5-lipoxygenase, cyclo-
oxygenase and calmodulin inhibitory effects, and antifungal, anti-
microbial, antioxidative, muscle-relaxing, and cytotoxic activities.
Recently, we found that the natural bisbibenzyl compound
marchantin C (Fig. 1) was a novel microtubule inhibitor, which
tion.14 Another bisbibenzyl riccardin D (Fig. 1), previously isolated
in our group, was also proven to exhibit excellent anti-cancer
activity,15 and the structure similarity of riccardin D and marchantin
C on bisbibenzyl skeleton implies that riccardin D might also target
on microtubules.
The attractive biological results motivated us to synthesize the
bisbibenzyls and their derivatives in order to improve their bioac-
tivities and to discover more potent cytotoxic agents. It has been
reported that the antiproliferative effect of some natural com-
pounds can be enhanced obviously by halogenation and aminome-
thylation.16–21 In light of this point, we were interested in the effect
of the bromine and aminomethyl group on the macrocyclic bisbib-
enzyl system. Accordingly, some brominated and aminomethylat-
ed derivatives were prepared based on the total synthesized
marchantin C, riccardin D as well as the structure similarity com-
pound plagiochin E.22
Marchantin C has been previously synthesized,23 and in present
study, we report the synthetic details of riccardin D and plagiochin
E, which were slightly modified according to previous report, and
the preparation of brominated and aminomethylated derivatives
of above three bisbibenzyls, as well as their cytotoxic activities
against the KB, MCF-7 and PC3 cell lines. Molecular docking anal-
yses were also used to elucidate the potential binding modes of
the derivatives to tubulin.24,25
⇑
Corresponding author. Tel.: +86 531 88364778; fax: +86 531 88382019.
E-mail address: louhongxiang@sdu.edu.cn (H. Lou).
These authors contributed equally to this work.
0968-0896/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2012.02.004

J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
2383
commercially available 4-iodo-benzaldehyde, resulting in the for-
mation of the diphenyl ether 2.26 Compound 2 was then reduced
with sodium borohydride to give the benzyl alcohol, followed by
bromination and reaction with triphenylphosphine, affording
phosphonium salt 3 in three steps. The biphenyl building block 9
was synthesized from compound 6 and 8 by standard Suzuki reac-
tion, following the approach reported by Speicher et al.27 The
building blocks 3 and 9 were coupled by Wittig reaction in the
presence of potassium carbonate and 18-crown-6.28 The stilbene
10 (obtained as E/Z mixture) was then hydrogenated over Pd/C,
and the carboxylic ester group was reduced with lithium alumin-
ium hydride, followed by acidic hydrolysis. After bromination with
carbon tetrabromide and subsequent reaction with triphenylphos-
phine, compound 11 was obtained. Cyclization of 11 by means of
an intramolecular Wittig reaction was achieved with sodium
methoxide, leading to intermediate 12. Riccardin D was finally ob-
tained after the hydrogenation and subsequent methyl ether cleav-
age. The synthesis of plagiochin E was achieved in 15 steps as
shown in Scheme 2. The biphenyl moiety 15 was prepared from
2-bromo-5-methoxylbenzaldehyde 13 by the acetal protection,
halogen/lithium exchange and subsequent scavenging with tri-
methyl borate, followed by treatment with pinacol. The building
blocks 3 and 15 were connected by intermolecular Wittig reaction
and the macrocyclic stilbene 18 was synthesized by intramolecular
McMurry reaction, following the procedure depicted by Speicher
et al.22 Plagiochin E was finally obtained after hydrogenation and
methyl ether cleavage.29
Figure 1. The chemical structures of marchantin C, riccardin D and plagiochin E.
The derivatives 19, 23 and 26 were then prepared from marchan-
tin C, riccardin D and plagiochin E by Mannich reaction, and the
derivatives 20–22, 24, 25, 27 and 28 were prepared by bromination
with N-bromosuccinimide (NBS) and were purified by HPLC
Scheme 1. Synthesis of riccardin D. Reagents and conditions: (a) (i) 1,3-propane-
diol, DMS–DMF, DCM, rt, 24 h; (ii) 4-iodo-benzaldehyde, Cs2CO3, CuBr, TMHD, NMP,
57%; (b) (i) NaBH4, EtOH, rt, 2–3 h; (ii) CBr4, PPh3, DCM; (iii) PPh3, toluene, reflux,
81%; (c) (i) LiAlH4, THF, ꢀ40 °C, 0.5–1 h; (ii) 2,3-Dihydropyran, p-toluenesulfonic
acid, DCM, 0–25 °C, 0.5 h, 77%; (d) n-BuLi, B(OMe)3, KH2PO4, THF, ꢀ40 °C, 0.5 h, rt,
2 h, 78%; (e) Tf2O, pyridine, DCM, rt, 1–2 h, 95%;(f) Pd(PPh3)4, toluene, EtOH, 2 mol/L
Na2CO3, reflux, 10 h, 81%; (g) K2CO3, 18-crown-6, DCM, reflux, 24 h, 91%; (h) (i) Pd/C
(5%), 3 bar H2, Et3N, EtOAc, rt, 24 h; (ii) 2 M HCl/THF (1:1), rt, 12 h; (iii) CBr4, PPh3,
DCM; (iv) PPh3, toluene, reflux,67%; (i) NaOMe, DCM, rt, 24 h, 67%; (j) (i) Pd/C (5%),
3 bar H2, EtOAc, rt, 24 h; (ii) BBr3, DCM, –40 °C to rt, 12 h, 81%.
2. Results and discussion
2.1. Chemistry
Scheme 2. Synthesis of plagiochin E. Reagents and conditions: (a) (i) 1,3-propane-
diol, p-toluenesul- fonic acid, toluene, reflux, 24 h; (ii) n-BuLi, B(OMe)3, KH2PO4,
THF, ꢀ40 °C, 0.5 h, rt, 2 h; (iii) pinacol, MgSO4, toluene, rt, 12 h, 69%; (b) compound
8, Pd(PPh3)4, toluene, EtOH, 2 mol/L Na2CO3, reflux, 10 h, 80%; (c) K2CO3, 18-crown-
6, CH2Cl2, reflux, 24 h, 91% (d) (i) Pd/C (5%), 3 bar H2, Et3N, EtOAc, rt, 24 h; (ii) 2 M
HCl/THF (1:1), rt, 12 h, 89%; (e) (i) Zn, TiCl4, THF, reflux, 24 h, 40%; (ii) Pd/C (5%),
3 bar H2, EtOAc, rt, 24 h; (f) BBr3, DCM, –40 °C to rt, 12 h, 70%.
The synthesis of riccardin D was achieved in 18 steps as shown
in Scheme 1. The synthetic route began with the Ullmann coupling
of the protected 3-hydroxy-4-methoxyl-benzaldehyde with

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J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
Scheme 3. Synthesis of marchantin C’s derivatives. Reagents and conditions: (a) Me2NH, HCHO, MeOH, reflux, 82%; (b) NBS, CH3CN, 0 °C, 89% in all.
2.2. Biological evaluation of bisbibenzyl derivatives
2.2.1. Bisbibenzyl derivatives inhibit cancer cell proliferation
in vitro
To evaluate the anticancer effects of bisbibenzyl derivatives
in vitro (19–28, 32), MTT assays were performed as described in
the Section 4 to examine their proliferative inhibitory activity
against three cancer cell lines: KB (oral cancer cell line), MCF-7
(human breast adenocarcinoma cell line) and PC3 (prostate cancer
Scheme 4. Synthesis of plagiochin E’s derivatives. Reagents and conditions: (a)
Me2NH, HCHO, MeOH, reflux, 79%; (b) NBS, CH3CN, 0 °C, 86% in all.
cell line). The inhibition percentage at a concentration of 20 lM
and the IC50 values of each compounds are reported in Table 1.
The data demonstrate that most derivatives exhibit obvious inhib-
itory activity against three human cancer cell lines, with IC50 value
ranging from 5.4 to 33.8 lM. Within the marchantin C derivatives,
the bromination at either the para- or ortho-position of hydroxyl
group on ring B improves the activity obviously (compounds 21
and 22); however, bromination at the meta-position and aminome-
thylation at the ortho-position of hydroxyl group on ring B im-
pacted the activity slightly, and the dimer compound 32 was less
potent than marchantin C. The cytotoxicity of brominated plagio-
chin E 24 and 25 was greatly improved comparing to the parent
compound and the aminomethylated product 23 also exhibited
good activity against KB cell line. The riccardin D derivative 28,
with a bromide at the para-position of hydroxyl group on ring B,
was the most cytotoxic compound against KB, MCF-7 and PC3 cell
Scheme 5. Synthesis of riccardin D’s derivatives. Reagents and conditions: (a)
Me2NH, HCHO, MeOH, reflux, 80%; (b) NBS, CH3CN, 0 °C, 76% in all.
lines, with IC50 values of 5.9, 5.4 and 5.6 lM, respectively. The sub-
stitution of aminomethyl group on ring B of riccardin D led to re-
duced activity (compound 26), and the bromination at the ortho-
position of hydroxyl group on ring B made the resulting compound
inactive (compound 27, IC50 >50 lM). Overall, the activities of most
brominated derivatives (compounds 21, 22, 24, 25, and 28) were
more potent than their parent compounds and aminomethylated
derivatives (compounds 19, 23 and 26) hardly exhibited the im-
proved activity.
2.2.2. Bisbibenzyl derivative induces cell cycle arrest at G2/M
phase
After the activity evaluation, we extend our work to the mech-
anism investigation and the flow cytometry was then used to ana-
lyze the effects of bisbibenzyl derivative on the cell growth and
division. The most potent antitubulin agent 28 was selected for cell
cycle studies in PC3, MCF-7 and KB cell lines by measuring the DNA
content. PC3 cells were cultured with 2.5, 5 or 10 lM 28, respec-
tively, for 24 h and then collected for flow cytometry assay. As
shown in Figure 3, in the absence of 28, there were 7.02% of cells
in G2/M phase. In contrast, significant accumulation of PC3 cells
in G2/M phase were observed after 28 treatment at the concentra-
Scheme 6. Synthesis of marchantin C’s dimmer. Reagents and conditions: (a)
NaOMe, DCM, 76% in all; (b) (i) H2, Pd/C, EtOAc; (ii) BBr3, DCM, 89%.
tion of 2.5 (6.15%), 5 (17.05%) and 10 lM (30.10%). Such cells ap-
pear to have the capacity to replicate their DNA but are not able
to proceed through the cell cycle to cell division. The effects of
28 on cell growth of MCF-7 and KB cell lines have also been eval-
uated by the same way as used for PC3 cells. As shown in Figure
4 and 5, the cell cycle of both cell lines were arrested in G2/M phase
(Schemes 3–5). In addition, the macrocyclic compound 31 was
obtained as byproduct when the synthesis of marchantin C was
scaled up, and the dimer of marchantin C 32 was prepared after
the hydrogenation and methyl ether cleavage, accordingly (Scheme
6).
as well, and increasing concentration of 28 to 10 lM led to essen-

J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
2385
Table 1
In vitro cytotoxicity of three bisbibenzyls and their derivatives in three cancer cell lines
Compound
KB
MCF-7
PC3
b
b
b
% Inhiba
IC50
(
lM)
% Inhiba
IC50
(
lM)
% Inhiba
IC50
(lM)
19
20
21
22
23
24
25
26
83.75%
81.34%
68.54%
76.76%
2.62%
84.32%
90.71%
91.59%
1.63%
16.8 1.20
11.5 0.15
11.0 0.70
11.3 0.22
15.1 0.28
8.2 0.72
9.7 0.48
10.7 0.07
>50
85.46%
62.55%
58.45%
83.01%
0.75%
62.41%
81.37%
92.83%
6.76%
17.5 0.27
15.3 0.41
6.3 0.82
12.1 0.14
33.8 2.37
6.3 0.69
8.4 0.09
13.3 0.21
>50
69.26%
48.66%
56.80%
64.88%
39.58%
71.74%
91.82%
98.03%
2.23%
12.6 0.34
15.7 0.42
15.6 0.23
8.5 0.02
25.1 0.54
9.3 0.05
9.2 0.03
9.7 0.02
>50
27
28
32
95.26%
87.91%
22.30%
98.12%
10.27%
5.9 0.30
13.8 0.54
15.3 0.32
7.1 0.11
40.1 1.52
95.53%
56.85%
13.03%
59.69%
6.90%
5.4 0.07
16.4 0.24
12.8 0.22
6.6 0.70
34.9 1.32
97.45%
20.66%
17.39%
94.87%
8.99%
5.6 0.23
27.2 0.13
15.8 0.28
10.1 0.09
28.0 1.86
Marchantin C
Riccardin D
Plagiochin E
a
The inhibition percentage was tested at a concentration of 20
The IC50 values (lM) are the concentrations corresponding to 50% inhibition of each cell line.
l
M of each compound.
b
tially the same result in MCF-7 and KB cells (20.24% and 39.56% in
G2/M phase, respectively). Overall, these results demonstrated that
bisbibenzyl derivative 28 induced cell cycle arrest at G2/M phase in
a dose-dependent manner, which is consistent with those obtained
for classical tubulin-targeting drugs.30
2.2.3. Immunofluorescence microscopy observation
The significant cell growth inhibitory properties of bisbibenzyl
derivatives supported by their obvious G2/M blocking properties
promoted us to investigate the further biological mechanism. Since
the bisbibenzyl compound marchantin C has been reported to
Figure 2. The chemical structures of combretastatin A-4 and riccardin D. The blue
bonds emphasize the structure similarity of these two compounds.
Figure 3. Compound 28 induced PC3 cell cycle arrest at G2/M phase. PC3 cells were treated with 2.5, 5, 10
lM of 28 for 24 h and then trypsinized, fixed and stained with PI to
measure cell cycle profile by flow cytometry. Control cells were treated with DMSO alone. Results are representatives of three independent experiments.

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J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
Figure 4. Compound 28 induced MCF-7 cell cycle arrest at G2/M phase. MCF-7 cells were treated with 2.5, 5, 10
lM of 28 for 24 h and then trypsinized, fixed and stained with
PI to measure cell cycle profile by flow cytometry. Control cells were treated with DMSO alone. Results are representatives of three independent experiments.
exhibit the microtubule depolymerization activity, we next exam-
ined the effect of selected potent compounds 22, 24, 26 and 28 on
microtubule by immunofluorescent staining techniques. First, PC3
cells were cultured for 24 h in the presence of compounds 22, 24,
similarity between CA-4 and bisbibenzyls implies that bisbibenzyls
might also bind to the colchicine binding site. Based on this
judgment, the most potent compound 28 was docked into the
colchicine binding site of tubulin (PDB code 1SA0) using the GOLD
(Genetic Optimization for Ligand Docking) program,39 and the en-
ergy minimized. In the resulting hypothetical structure (Fig. 7 and
Fig. 8), the hydroxyl group on ring C may forms hydrogen bonds
with carbonyl group of Ser178 and hydroxyl group of Try224.
The hydroxyl group on ring B is involved in a potential hydrogen
bond with the carbonyl group of Lys254. It is also possible that ring
26 and 28 at the concentration of 8, 10, 6 and 6
and then fixed with cold methanol/acetone, followed by immuno-
staining for -tubulin and b-actin. The cellular microtubule net-
lM, respectively,
a
works and actins were then visualized by fluorescence
microscopy. As shown in Figure 6, intact microtubules arrays could
be observed in untreated cells. However, after treatment with com-
pounds 22, 24, 26 or 28, microtubules networks were decreased
and short microtubules fragments were observed in the cytoplasm.
Such morphological changes are indicative of depolymerized and
dispersed tubulin dimmers, which are very similar to the reported
cellular changes caused by treatment with marchantin C and
colchicine.14,31
B forms a CH-
were able to observe that the bromine atom on ring B could fit into
the pocket formed by Ala316 and Lys352. These combined CH–
p interaction with the methyl group of Leu255. We
p
interaction and hydrogen bonds could play a crucial role in tubulin
inhibitory activity of the bisbibenzyl compounds. In addition, the
introduction of bromine atom to riccardin D could change the elec-
tron distribution on ring B, which may create stronger hydrogen
2.2.4. Molecular modeling
bond with Lys254 and CH–p interaction with Leu255, and this
The excellent bioactivity of derivatives encouraged us to inves-
tigate the possible mechanism of action at the molecular level, spe-
cifically, the binding mode of bisbibenzyls to tubulin.32,33 It has
been reported that there are three ligand binding sites in tubulins:
the colchicine, vinca alkaloid, and taxane.34 The antimitotic agents
binding to taxane binding site inhibit the depolymerization of
microtubules, while the compounds binding to vinca alkaloid bind-
ing site and colchicine binding site inhibit the polymerization of
microtubules. Combretastatin A-4 (CA-4) (Fig. 2) is a potent antimi-
totic agent by inhibiting the polymerization of microtubules which
is similar to the mechanism of bisbibenzyl. CA-4 has been proved
to binds to colchicine binding site.14,35–38 The structure and effect
might be the reason for the improved potency of compound 28
than riccardin D.
3. Conclusions
As shown in Schemes 1–6, and Table 1, we have achieved the
synthesis of riccardin D and plagiochin E, prepared a series of bis-
bibenzyl derivatives (19–28 and 32) with the bromine and amino-
methyl groups, and tested their cytotoxic activity against PC3,
MCF-7 and KB cell lines. Among the tested compounds, 28 showed
the most potent antiproliferative activity toward three cell lines,
and compounds 21, 24, 25 and 26 also exhibited excellent bioactiv-

J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
2387
Figure 5. Compound 28 induced KB cell cycle arrest at G2/M phase. KB cells were treated with 2.5, 5, 10
lM of 28 for 24 h and then trypsinized, fixed and stained with PI to
measure cell cycle profile by flow cytometry. Control cells were treated with DMSO alone. Results are representatives of three independent experiments.
ity. In addition, the further mechanism study revealed that bisbib-
enzyl derivatives could arrest cancer cells at G2/M phase by
destroying the microtubules network. Finally, molecular modeling
was also used to elucidate the binding models of the derivatives to
tubulin. From the study of a preliminary structure-activity rela-
tionship, it was considered that the bisbibenzyl skeleton and phe-
nolic hydroxyl groups played essential roles in the tubulin
inhibitory activity. The introduction of bromine into the structure
could change the electron distribution on benzene ring, which
were obtained on a LTQ Orbitrap mass spectrometer. Reagents
were used as purchased without further purification. Solvents
(THF, DCM, pyridine and toluene) were dried and freshly distilled
before use according to procedures reported in the literature.
4.1.1. General procedures
4.1.1.1. General procedure 1 (GP 1) for the bromination of bis-
bibenzyls.
A mixture of bisbibenzyl (riccardin D, plagiochin E
or marchantin C, 20 mg, 0.047 mmol) and a catalytic amount of
manganese dioxide in anhydrous CH2Cl2 (1 mL) was stirred at
might improve the strength of hydrogen bond and CH–
p interac-
tion between the protein and ligand. This model could be utilized
in the further modification of bisbibenzyls for the discovery of no-
vel anticancer agents.
room temperature. The bromine (2.4 lL, 0.047 mmol)in anhydrous
CH2Cl2 (0.5 mL) was added dropwise to the solution and the reac-
tion system was stirred for 24 h. The solvent was evaporated and
the residue was purified by HPLC (80% methanol in water).
4. Experimental section
4.1. Chemistry
4.1.1.2. General procedure 2 (GP 2) for the bromination of bis-
bibenzyls.
To a solution of bisbibenzyl (riccardin D, plagio-
chin or marchantin C, 20 mg, 0.047 mmol) in acetonitrile
E
(1.3 mL) was added NBS (4.2 mg, 0.0235 mmol). The mixture was
stirred at 0 °C for 12 h. After rise to room temperature, the solid
was filtered off and the solvent was removed in vacuo. The residue
was purified by HPLC (78% methanol in water).
Column chromatography was carried out on silica gel or alu-
mina (200–300 mesh). Reactions were monitored by thin-layer
chromatography, using Merck plates with fluorescent indicator.
Melting points were determined on an X-6 melting-point appara-
tus and are uncorrected. The NMR spectra were recorded on a Bru-
ker Spectospin spectrometer at 600 MHz (1H) and 150 MHz (13C),
using TMS as an internal standard. The chemical shifts are reported
in parts per million (ppm d) referenced to the residual 1H reso-
nance of the solvent (CDCl3, 7.28 ppm). Abbreviations used in the
splitting pattern were as follows: s = singlet, d = doublet, t = triplet,
quin = quintet, m = multiplet, and br=broad. All HRMS spectra (ESI)
4.1.1.3. General procedure 3 (GP 3) for the aminomethylation of
bisbibenzyls.
Formaldehyde aqueous solution (37%, 5 lL,
0.061 mmol) was added to a solution of bisbibenzyl (riccardin D,
plagiochin E or marchantin C, 20 mg, 0.047 mmol) in methanol
(1 mL). Dimethylamine was then added to the reaction mixture
at 65 °C. The mixture was refluxed for 12 h and after cooling to

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J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
Figure 7. Ribbon diagram illustrating the hypothetical complex structure of tubulin
with compound 28 (orange) bound to the colchicine binding site.
Figure 6. Effects of selective derivatives 22, 24, 26 and 28 on the microtubule
distribution in PC3 cells. PC3 cells were incubated with 22, 24, 26 and 28 at the
concentration of 8, 10, 6 and 6 lM, respectively, for 24 h and then fixed and
immunostained with monoclonal antibody. DMSO was used as control.
Figure 8. Potential interactions between compound 28 and Ala316, Lys352, Ser178,
Tyr224, Leu255 and Lys254 in tubulin colchicine binding site. The diagram is
programmed for wall-eyed (relaxed) viewing. The hydrogen bonds are labeled as
yellow lines.
room temperature the solvent was evaporated. The crude product
was purified by column chromatography on silica gel eluted with
acetone: petroleum ether (3:8).
4.2. Synthesis
3H, OCH3), 3.61 (m, 1H, –CH2–), 1.93 (m, 1H, –CH2–), 1.82 (m,
1H, –CH2–), 1.70 (m, 3H, –CH2–), 1.27 (m, 1H, –CH2–); HRMS
(ESI) calcd for C21H24O5 356.4127 found 356.4223; MS (ESI) 357
(M+H)+.
4.2.1. The data of three parent compounds and their
intermediates
4.2.1.1. The dimer of marchantin C (32).
Compound 32 was
prepared by following the procedure reported in the literature.23
White solid. Mp 151–152 °C. 1H NMR (CDCl3): d = 7.28 (s, 12H,
Ar-H), 7.19 (t, J = 7.8 Hz, 1H, Ar-H), 7.10 (t, J = 7.8 Hz, 1H, Ar-H),
6.94 (d, J = 8.4 Hz, 2H, Ar-H), 6.92 (d, 2H, Ar-H), 6.81 (m, 5H, Ar-
H), 6.72 (d, J = 7.8 Hz, 1H, Ar-H), 6.68 (s, 1H, Ar-H), 6.56 (s, 1H,
Ar-H), 5.44 (s, 1H, Ar-H), 5.33 (s, 1H, Ar-H), 2.72 (m, 9H, –CH2–),
1.27 (m, 7H, –CH2–); HRMS (ESI) calcd for C56H48O8 848.9732
found 848.9756; MS (ESI) 849 (M+H)+.
4.2.1.3. The Bibenzyl (11).
Compound 11 was prepared by
following the procedure reported in the literature.23 Yellow oil.
1H NMR (CDCl3): d = 7.29 (d, J = 3 Hz, 1H, Ar-H), 7.26 (dd,
J1 = 7.8 Hz, J2 = 1.8 Hz, 1H, Ar-H), 7.08 (d, J = 7.2 Hz, 2H, Ar-H),
7.05 (d, J = 5.4 Hz, 2H, Ar-H), 6.99 (d, J = 8.4 Hz, 1H, Ar-H), 6.92
(d, J = 7.2 Hz, 1H, Ar-H), 6.86 (d, J = 8.4 Hz, 1H, Ar-H), 6.83 (d,
J = 7.8 Hz, 2H, Ar-H), 6.79 (d, J = 8.4 Hz, 2H, Ar-H), 5.41 (s, 1H,
Ph–CH–O), 4.89 (dd, J1 = 12.44 Hz, J2 = 2.4 Hz, 1H, Ph–CH2–O),
4.80 (m, 1H, O–CH–O), 4.60 (d, J = 12.44 Hz, 1H, Ph–CH2–O), 4.23
(dd, J1 = 11.44 Hz, J2 = 4.8 Hz, 2H, –CH2–),3.97 (m, 3H, –CH2–),
3.85(s, 3H, OCH3), 3.78 (s, 3H, OCH3), 3.73 (s, 3H, OCH3) 3.60 (m,
1H, –CH2–), 2.68 (m, 3H, –CH2–), 2.57 (m, 1H, –CH2–), 2.20 (m,
1H, –CH2–), 2.08 (m, 1H, –CH2–), 1.92 (m, 1H, –CH2–), 1.79 (m,
1H, –CH2–), 1.72 (m, 1H, –CH2–), 1.64 (m, 3H, –CH2–); HRMS
(ESI) calcd for C39H44O8 640.7620 found 640.7596; MS (ESI) 641
(M+H)+.
4.2.1.2.
yloxy)methyl]biphenyl-2-carbaldehyde (9).
20,6-Dimethoxy-40-[(tetrahydro-2H-pyran-2-
Compound
9
was by following the procedure reported in the literature.23 Color-
less oil. 1H NMR (CDCl3): d = 9.69 (s, 1H, CHO), 7.63 (d, J = 7.8 Hz,
1H, Ar-H), 7.47 (t, J = 7.8 Hz, 1H, Ar-H), 7.22 (t, J = 7.2 Hz, 2H, Ar-
H), 7.07 (m, 1H, Ar-H), 7.04 (d, J = 4.8 Hz, 1H, Ar-H), 4.89 (d,
J = 12.44 Hz, 1H, Ph –CH2–O), 4.79 (m, 1H, O–CH–O), 4.59 (dd,
1H, Ph –CH2–O), 3.98 (m, 1H, –CH2–), 3.80 (s, 3H, OCH3), 3.76 (s,

J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
2389
4.2.1.4. Riccardin D trimethyl ether (12).
Compound 12 was
J = 8.4 Hz, 1H, Ar-H), 6.73 (d, J = 7.8 Hz, 2H, Ar-H), 6.69 (d,
J = 7.8 Hz, 1H, Ar-H), 6.67 (s, 1H, Ar-H), 6.66 (dd, J1 = 7.8 Hz,
J2 = 2.4 Hz, 1H, Ar-H), 5.60 (s, 1H, Ar-H), 3.32 (m, 2H, –CH2–),
3.05 (m, 2H, –CH2–), 2.96 (m, 2H, –CH2–), 2.85 (m, 2H, –CH2–);
HRMS (ESI) calcd for C28H24O4 424.1713 found 424.1610; MS
(ESI) 425 (M+H)+.
prepared by following the procedure reported in the literature.22
White solid. Mp 116–117 °C. 1H NMR (CDCl3): d = 7.39 (t,
J = 7.8 Hz, 1 H, Ar-H), 7.12 (d, J = 7.8 Hz, 1H, Ar-H), 6.95 (d,
J = 8.4 Hz, 1H, Ar-H), 6.93 (d, J = 8.4 Hz, 1H, Ar-H), 6.91 (d,
J = 7.2 Hz, 1 H, Ar-H), 6.88 (d, J = 7.2 Hz, 1H, Ar-H), 6.87 (d,
J = 7.8 Hz, 1H, Ar-H), 6.83 (dd, J1 = 7.2 Hz, J2 = 1.8 Hz, 2H, Ar-H),
6.79 (d, J = 7.8 Hz, 1H, Ar-H), 6.45 (d, J = 7.8 Hz, 1H, Ar-H), 6.43 (s,
1H, Ar-H), 5.51 (s, 1H, Ar-H), 3.99 (s, 3H, OCH3), 3.72 (s, 3H,
OCH3), 3.70 (s, 3H, OCH3), 2.98 (m, 3H, –CH2–), 2.76 (m, 5H, –
CH2–); HRMS (ESI) calcd for C31H30O4 466.5715 found 466.5618;
MS (ESI) 467 (M+H)+.
4.2.2. The data of all the derivatives
4.2.2.1. Brominated derivative of marchantin C (19).
Com-
pound 19 was prepared by following the GP3 from marchantin C,
yield 80%, White powder, mp 79–80 °C. 1H NMR (CDCl3): d = 7.01
(d, J = 8.4 Hz, 2H, Ar-H), 6.94 (t, J = 7.8 Hz, 1H, Ar-H), 6.89 (d,
J = 4.2 Hz, 1H, Ar-H), 6.88 (d, J = 4.2 Hz, 1H, Ar-H), 6.88 (dd,
J1 = 8.4 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.74 (dd, J1 = 7.8 Hz, J2 = 1.8 Hz,
1H, Ar-H), 6.67 (s, 1H, Ar-H), 6.62 (d, J = 8.4 Hz, 2H, Ar-H), 6.49
(dd, J1 = 8.4 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.29 (s, J = 7.8 Hz, 1H, Ar-
H), 5.55 (d, J = 1.8 Hz, 1H, Ar-H), 5.32 (s, 1H, OH), 3.75 (d,
J = 7.2 Hz, 1H, Ph –CH2–N), 3.73 (d, J = 7.2 Hz, 1H, Ph –CH2–N),
3.07–3.01 (m, 4H, Ph-CH2CH2-Ph), 2.88–2.84 (m, 2H, Ph-CH2CH2-
Ph), 2.78–2.76 (m, 2H, Ph-CH2CH2-Ph), 2.31 (s, 6H, N-CH3); 13C
NMR (CDCl3): d = 157.86, 152.58, 150.42, 145.95, 143.33, 142.08,
140.02, 139.25, 135.73, 133.00, 129.72, 129.63, 127.94, 124.46,
122.51, 122.24, 121.28, 120.06, 115.57, 115.43, 114.73, 112.06,
58.47, 44.18, 36.40, 36.10, 34.71, 30.36; HRMS (ESI) calcd for
4.2.1.5. Riccardin D.
Riccardin D was prepared by following
the procedure reported in the literature.22 White solid. Mp 152–
153 °C. 1H NMR (CDCl3): d = 7.37 (t, J = 7.8 Hz, 1H, Ar-H), 7.09 (d,
J = 7.8 Hz, 1H, Ar-H), 6.95 (d, J = 7.8 Hz, 2H, Ar-H), 6.93 (d,
J = 8.4 Hz, 1H, Ar-H), 6.88 (d, J = 8.4 Hz, 1H, Ar-H), 6.85 (d,
J = 7.8 Hz, 2H, Ar-H), 6.80 (d, J = 8.4 Hz, 1H, Ar-H), 6.77 (d,
J = 7.8 Hz, 1H, Ar-H), 5.01 (s, 1H, -OH), 4.98 (s, 1H, -OH), 3.01 (m,
2H, –CH2–), 2.93 (t, 1H, –CH2–), 2.79 (m, 2H, –CH2–), 2.68 (m,
2H, –CH2–), 2.63 (t, 1H, –CH2–); HRMS (ESI) calcd for C28H24O4
424.4775 found 424.4735; MS (ESI) 465 (M+H)+.
4.2.1.6. 20-(1,3-Dioxan-2-yl)-40,6-dimethoxybiphenyl-2-carbal-
C
31H32O4N 482.2323 found 482.2326; MS (ESI) 482 (M+H)+.
dehyde (15).
Compound 15 was prepared by following the
procedure reported in the literature.22 Yellow solid. Mp 105–
106 °C. 1H NMR (CDCl3): d = 9.57 (s, 1H, CHO), 7.64 (d, J = 7.8 Hz,
1H, Ar-H), 7.49 (t, J = 7.8 Hz, 1H, Ar-H), 7.35 (d, J = 1.2 Hz, 1H, Ar-
H), 7.20 (d, J = 7.8 Hz, 1H, Ar-H), 7.11 (d, J = 8.4 Hz, 1H, Ar-H),
6.98 (dd, J1 = 8.4 Hz, J2 = 1.8 Hz, 1H, Ar-H), 5.00 (s, 1H, -CH-Ph),
4.18 (m, 1H, –CH2–), 3.93 (m, 1H, –CH2–), 3.92 (s, 3H, OCH3),
3.78 (s, 3H, OCH3), 3.71 (t, 1H, –CH2–), 3.42 (t, 1H, –CH2–), 2.12
(m, 1H, –CH2–), 1.27 (m, 1H, –CH2–); HRMS (ESI) calcd for
4.2.2.2. Brominated derivative of marchantin C (20).
Com-
pound 20 was prepared by following the GP1 from marchantin C,
yield 70%, White powder, mp 110–111 °C. 1H NMR (CDCl3):
d = 6.99 (d, J = 8.4 Hz, 2H, Ar-H), 6.92 (t, J = 7.8 Hz, Ar-H), 6.89 (d,
J = 7.8 Hz, 1H, Ar-H), 6.87 (d, J = 8.4 Hz, 1H, Ar-H), 6.72 (dd,
J1 = 8.4 Hz, J2 = 1.8 Hz, 1H, Ar-H), 6.64 (s, 1H, Ar-H), 6.61 (d,
J = 7.8 Hz, 2H, Ar-H), 6.47 (dd, J1 = 8.4 Hz, J2 = 1.8 Hz, 1H, Ar-H),
6.28 (d, J = 7.2 Hz, 1H, Ar-H), 5.53 (d, J = 1.8 Hz, 1H, Ar-H), 3.06–
2.88 (m, 4H, Ph–CH2CH2–Ph), 2.84–2.74 (m, 4H, Ph–CH2CH2–Ph);
13C NMR (CDCl3): d = 157.74, 152.60, 145.94, 143.33, 142.19,
140.09, 139.22, 132.96, 129.72, 128.04, 122.63, 122.26, 121.30,
120.30, 115.54, 115.45, 114.73, 112.02, 36.36, 36.03, 34.67,
C
19H20O5 328.3535 found 328.2126; MS (ESI) 329 (M+H)+.
4.2.1.7. The bibenzyl dialdehyde (17). Compound 17 was
prepared by following the procedure reported in the literature.22
Yellow oil. 1H NMR (CDCl3): d = 9.82 (s, 1H, CHO), 9.60 (s, 1H,
CHO), 7.64 (dd, J1 = 8.4 Hz, J2 = 1.8 Hz, 1H, Ar-H), 7.54 (d,
J = 2.4 Hz, 1H, Ar-H), 7.41 (d, J = 1.8 Hz, 1H, Ar-H), 7.35 (t,
J = 7.8 Hz, 1H, Ar-H), 7.23 (dd, J1 = 8.4 Hz, J2 = 3.0 Hz, 1H, Ar-H),
7.11 (d, J = 8.4 Hz, 2H, Ar-H), 6.98 (d, J = 7.8 Hz, 1H, Ar-H), 6.84
(m, 5H, Ar-H), 3.94 (s, 3H, OCH3), 3.92 (s, 3H, OCH3), 3.68 (s, 3H,
OCH3) 2.68 (m, 4H, –CH2–); HRMS (ESI) calcd for C31H28O6
496.2323 found 496.2315; MS (ESI) 497 (M+H)+.
30.39; HRMS (ESI) calcd for
C28H27O4NBr 520.1118; found
520.1112 (M+NH4+); MS (ESI) 503, 501 (MꢀH)ꢀ.
4.2.2.3. Brominated derivative of marchantin C (21).
Com-
pound 21 was prepared by following the GP2 from marchantin C,
yield 40%, White powder, mp 116–117 °C. 1H NMR (CDCl3):
d = 7.41 (d, J = 9 Hz, 1H, Ar-H), 7.03 (d, J = 8.4 Hz, 2H, Ar-H), 6.97
(t, J = 7.8 Hz, 1H, Ar-H), 6.91 (d, J = 8.4 Hz, 1H, Ar-H), 6.81 (d,
J = 9 Hz, 1H, Ar-H), 6.78 (s, 1H, Ar-H), 6.76 (dd, J1 = 7.8 Hz,
J2 = 1.2 Hz, 1H, Ar-H), 6.70 (d, J = 7.8 Hz, 2H, Ar-H), 6.45 (d,
J = 7.2 Hz, 1H, Ar-H), 6.37 (dd, J1 = 8.4 Hz, J2 = 1.8 Hz, 1H, Ar-H),
5.60 (d, J = 1.2 Hz, 1H, Ar-H), 5.55 (s, 1H, OH), 4.66 (s, 1H, OH),
3.15–3.13 (m, 2H, Ph–CH2CH2–Ph), 3.06–3.05 (m, 2H, Ph–
CH2CH2–Ph), 2.88–2.86 (m, 2H, Ph–CH2CH2–Ph), 2.81–2.80 (m,
2H, Ph–CH2CH2–Ph); 13C NMR (CDCl3): d = 156.21, 152.94,
147.76, 146.27, 143.80, 143.61, 139.78, 136.67, 133.01, 130.04,
129.93, 129.06, 124.35, 122.42, 121.54, 116.26, 116.02, 115.80,
114.74, 114.98, 110.78, 110.02, 36.66, 34.93, 34.58, 31.82; HRMS
(ESI) calcd for C28H27O4NBr 520.1118; found 520.1112 (M+NH4+);
MS (ESI) 503, 501 (MꢀH)ꢀ.
4.2.1.8. Plagiochin E trimethyl ether (18).
Compound 18
was prepared by following the procedure reported in the litera-
ture.22 White solid. Mp 103–104 °C. 1H NMR (CDCl3): d = 7.38 (d,
J = 6.0 Hz, 1H, Ar-H), 7.37 (d, J = 7.8 Hz, 1H, Ar-H), 7.05 (d,
J = 8.4 Hz, 2H, Ar-H), 6.93 (dd, J1 = 9.6 Hz, J2 = 3.0 Hz, 1H, Ar-H),
6.91 (d, J = 7.8 Hz, 1 H, Ar-H), 6.78 (t, J = 7.8 Hz, 1H, Ar-H), 6.76
(d, J = 8.4 Hz, 2H, Ar-H), 6.74 (d, J = 7.8 Hz, 1H, Ar-H), 6.73 (s, 1H,
Ar-H), 6.71 (dd, J1 = 9.6 Hz, J2 = 3.0 Hz, 1H, Ar-H), 5.26 (s, 1H, Ar-
H), 3.95 (s, 3H, OCH3), 3.81 (s, 3H, OCH3), 3.66 (s, 3H, OCH3), 3.32
(t, 1H, –CH2–), 3.05 (m, 2H, –CH2–), 2.96 (m, 1H, –CH2–), 2.85
(m, 2H, –CH2–), 2.55 (t, 1H, –CH2–), 1.16 (m, 1H, –CH2–); HRMS
(ESI) calcd for C31H30O4 466.5670 found 466.5623; MS (ESI) 467
(M+H)+.
4.2.2.4. Aminomethylated derivative of marchantin
(22).
C
Compound 22 was prepared by following the GP2 from
4.2.1.9. Plagiochin E.
Plagiochin E was prepared by following
marchantin C, yield 40%, White powder, mp 104–105 °C. 1H NMR
(CDCl3): d = 7.39 (d, J = 8.4 Hz, 1H, Ar-H), 6.99 (t, J = 7.8 Hz, 1H,
Ar-H), 6.97 (d, J = 8.4 Hz, 2H, Ar-H), 6.97 (dd, J1 = 8.4 Hz,
J2 = 2.4 Hz, 1H, Ar-H), 6.90 (d, J = 8.4 Hz, 1H, Ar-H), 6.76 (dd,
J1 = 7.8 Hz, J2 = 1.8 Hz, 1H, Ar-H), 6.63 (d, J = 8.4 Hz, 2H, Ar-H),
the procedure reported in the literature.22 White solid. Mp 197–
198 °C. 1H NMR (CDCl3): d = 7.29 (d, J = 7.8 Hz, 1H, Ar-H), 7.23 (d,
J = 7.8 Hz, 1H, Ar-H), 7.08 (d, J = 8.4 Hz, 2H, Ar-H), 6.85 (d,
J = 8.4 Hz, 1H, Ar-H), 6.82 (d, J = 7.8 Hz, 1 H, Ar-H), 6.74 (t,

2390
J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
6.52 (dd, J1 = 7.8 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.40 (d, J = 7.8 Hz, 1H,
Ar-H), 5.56 (d, J = 1.8 Hz, 1H, Ar-H), 5.50 (s, 1H, OH), 5.38 (s, 1H,
OH), 3.04–3.01 (m, 4H, Ph–CH2CH2–Ph), 2.89–2.87 (m, 2H, Ph–
CH2CH2–Ph), 2.81–2.79 (m, 2H, Ph–CH2CH2–Ph); 13C NMR (CDCl3):
d = 157.52, 150.23, 147.31, 145.50, 140.66, 140.09, 138.98, 136.09,
129.59, 127.00, 124.90, 122.87, 120.30, 119.95, 115.78, 115.15,
114.22, 113.31, 36.77, 36.07, 30.93, 29.70; HRMS (ESI) calcd for
4.2.2.8. Aminomethylated derivative
(26). Compound 26 was prepared by following the GP3 from
of riccardin
D
riccardin D, yield 80%, White powder, mp 110–111 °C. 1H NMR
(CDCl3): d = 7.03 (d, J = 7.2 Hz, 1H, Ar-H), 6.94 (d, J = 7.8 Hz, 1H,
Ar-H), 6.89 (d, J = 7.8 Hz, 2H, Ar-H), 6.87 (d, J = 7.8 Hz, 1H, Ar-H),
6.85 (d, J = 9.6 Hz, 1H, Ar-H), 6.79 (t, J = 9.6 Hz, 2H, Ar-H), 6.71 (d,
J = 7.8 Hz, 1H, Ar-H), 6.52 (d, J = 7.2 Hz, 1H, Ar-H), 6.25 (s, 1H, Ar-
H), 5.33 (s, 1H, Ar-H), 5.28 (s, 1H, OH), 3.83 (d, J = 7.8 Hz, 1H, Ph–
CH2–N), 3.61 (d, J = 7.8 Hz, 1H, Ph–CH2–N), 2.96–2.90 (m, 3H, Ph–
CH2CH2–Ph), 2.83–2.77 (m, 3H, Ph–CH2CH2–Ph), 2.61–2.55 (m,
1H, Ph–CH2CH2–Ph), 2.51–2.48 (m, 1H, Ph–CH2CH2–Ph), 2.32 (s,
6H, N–CH3); 13C NMR (CDCl3): d = 155.00, 153.19, 152.76, 146.70,
143.67, 143.33, 142.06, 140.06, 140.35, 133.29, 132.03, 129.69,
128.87, 128.29, 124.30, 122.49, 122.08, 122.03, 121.85, 121.51,
120.88, 119.17, 118.70, 116.19, 114.87, 58.46, 44.17, 38.10, 37.67,
36.81, 34.50; HRMS (ESI) calcd for C31H32O4N 482.2323 found
482.2326; MS (ESI) 482 (M+H)+.
C
28H27O4NBr 520.1118; found 520.1112 (M+NH4+); MS (ESI) 503,
501 (MꢀH)ꢀ.
4.2.2.5. Aminomethylated derivative of plagiochin
E
(23).
Compound 23 was prepared by following the GP3 from
plagiochin E, yield 80%, White powder, mp 94–95 °C. 1H NMR
(CDCl3): d = 7.17 (d, J = 7.8 Hz, 1H, Ar-H), 7.05 (dd, J1 = 8.4 Hz,
J2 = 2.4 Hz, 1H, Ar-H), 7.02 (d, J = 7.8 Hz, 1H, Ar-H), 6.78 (dd,
J1 = 8.4 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.76 (d, J = 8.4 Hz, 1H, Ar-H),
6.73 (d, J = 7.8 Hz, 1H, Ar-H), 6.66 (dd, J1 = 7.8 Hz, J2 = 1.8 Hz, 1H,
Ar-H), 6.65 (dd, J1 = 8.4 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.60 (dd,
J1 = 7.8 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.54 (d, J = 2.4 Hz, 1H, Ar-H),
5.29 (d, J = 1.8 Hz, 1H, Ar-H), 3.72 (d, J = 7.8 Hz, 1H, Ph –CH2–N),
3.54 (d, J = 7.8 Hz, 1H, Ph–CH2–N), 3.22–3.19 (m, 4H, Ph–
CH2CH2–Ph), 2.74–2.61 (m, 4H, Ph–CH2CH2–Ph), 2.26 (s, 6H, N–
CH3); 13C NMR (CDCl3): d = 156.53, 156.14, 155.78,155.40,
143.43, 142.21, 140.36, 138.47, 132.72, 131.94, 130.06, 129.96,
128.58, 127.58, 126.67, 123.81, 122.21, 121.09, 119.00, 118.64,
115.62, 115.53, 112.74, 110.71, 62.38, 43.40, 35.23, 33.36, 29.66,
4.2.2.9. Brominated derivative of riccardin D (27).
Com-
pound 27 was prepared by following the GP2 from riccardin D,
yield 40%, White powder, mp 99–100 °C. 1H NMR (CDCl3):
d = 7.58 (d, J = 7.8 Hz, 1H, Ar-H), 7.01 (d, J = 8.4 Hz, 1H, Ar-H),
6.95 (d, J = 8.4 Hz, 1H, Ar-H), 6.94 (dd, J1 = 7.8 Hz, J2 = 1.2 Hz, 1H,
Ar-H), 6.88 (dd, J1 = 7.8 Hz, J2 = 1.2 Hz, 1H, Ar-H), 6.85 (dd,
J1 = 8.4 Hz, J2 = 1.2 Hz, 1H, Ar-H), 6.84 (d, J = 7.8 Hz, 1H, Ar-H),
6.82 (dd, J1 = 8.4 Hz, J2 = 1.2 Hz, 1H, Ar-H), 6.77 (dd, J1 = 8.4 Hz,
J2 = 2.4 Hz, 1H, Ar-H), 6.47 (d, J = 1.8 Hz, 1H, Ar-H), 6.39 (dd,
J1 = 7.8 Hz, J2 = 1.2 Hz, 1H, Ar-H), 5.51 (s, 1H, OH), 5.43 (d,
J = 2.4 Hz, 1H, Ar-H), 5.61 (s, 1H, OH), 4.75 (s, 1H, OH), 2.97–2.90
(m, 4H, Ph-CH2CH2-Ph), 2.79–2.67 (m, 4H, Ph–CH2CH2–Ph); 13C
NMR (CDCl3): d = 156.83, 156.79, 152.65, 144.78, 143.58, 143.06,
140.59, 140.10, 139.06, 136.76, 133.09, 132.63, 132.56, 131.53,
129.27, 123.94, 122.28, 122.13, 117.45, 117.31, 116.02, 115.05,
37.70, 37.66, 36.64, 34.78; HRMS (ESI) calcd for C28H27O4NBr
520.1118; found 520.1112 (M+NH4+); MS (ESI) 503, 501 (MꢀH)ꢀ.
29.59; HRMS (ESI) calcd for
C31H32O4N 482.2323 found
482.2326; MS (ESI) 482 (M+H)+.
4.2.2.6. Brominated derivative of plagiochin E (24).
Com-
pound 24 was prepared by following the GP2 from plagiochin E,
yield 50%, White powder, mp 126–127 °C. 1H NMR (CDCl3):
d = 7.52 (d, J = 8.4 Hz, 1H, Ar-H), 7.20 (d, J = 8.4 Hz, 1H, Ar-H),
7.04 (t, J = 8.4 Hz, 2H, Ar-H), 6.90 (dd, J1 = 8.4 Hz, J2 = 2.4 Hz, 1H,
Ar-H), 6.88 (d, J = 8.4 Hz, 1H, Ar-H), 6.82 (d, J = 7.8 Hz, 1H, Ar-H),
6.73 (dd, J1 = 9 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.71 (d, J = 9 Hz, 1H, Ar-
H), 6.69 (dd, J1 = 7.8 Hz, J2 = 1.8 Hz, 1H, Ar-H), 6.65 (d, J = 2.4 Hz,
1H, Ar-H), 5.46 (s, 1H, OH), 5.30 (s, 1H, OH), 5.21 (d, J = 2.4 Hz,
Ar-H), 4.77 (s, 1H, OH); 13C NMR (CDCl3): d = 157.24, 156.83,
154.32, 151.13, 142.66, 142.64, 140.01, 132.96, 132.52, 130.58,
130.36, 129.90, 124.26, 123.54, 122.77, 121.56, 121.38, 119.11,
118.56, 117.56, 115.42, 114.73, 113.88, 112.19, 35.37, 33.18,
30.63, 30.02; HRMS (ESI) calcd for C28H27O4NBr 520.1118; found
520.1112 (M+NH4+); MS (ESI) 503, 501 (MꢀH)ꢀ.
4.2.2.10. Brominated derivative of riccardin D (28).
Com-
pound 28 was prepared by following the GP2 from riccardin D,
yield 40%, White powder, mp 129–130 °C. 1H NMR (CDCl3):
d = 7.61 (d, J = 8.4 Hz, 1H, Ar-H), 7.01 (d, J = 8.4 Hz, 1H, Ar-H),
6.93 (s, 1H, Ar-H), 6.95 (dd, J1 = 7.8 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.85
(dd, J1 = 7.8 Hz, J2 = 1.8 Hz, 2H, Ar-H), 6.87 (t, J = 8.4 Hz, 1H, Ar-H),
6.82 (dd, J1 = 7.8 Hz, J2 = 1.8 Hz, 1H, Ar-H), 6.76 (dd, J1 = 7.8 Hz,
J2 = 1.2 Hz, Ar-H), 6.59 (s, 1H, Ar-H), 6.45 (d, J = 7.8 Hz, 1H, Ar-H),
5.64 (s, 1H, OH), 5.41 (d, J = 1.2 Hz, 1H, Ar-H), 4.85 (s, 1H, OH),
4.76 (s, 1H, OH), 3.05–2.83 (m, 4H, Ph–CH2CH2–Ph), 2.80–2.67
(m, 4H, Ph–CH2CH2–Ph); 13C NMR (CDCl3): d = 156.98, 155.06,
153.01, 152.93, 144.75, 142.74, 140.65, 135.46, 134.42, 133.31,
131.50, 129.68, 129.67, 122.68, 122.52, 122.36, 117.26, 117.19,
116.40, 115.80, 115.45, 113.61, 37.44, 36.08, 36.01, 35.52; HRMS
(ESI) calcd for C28H27O4NBr 520.1118; found 520.1112 (M+NH4+);
MS (ESI) 503, 501 (MꢀH)ꢀ.
4.2.2.7. Brominated derivative of plagiochin E (25).
Com-
pound 20 was prepared by following the GP2 from plagiochin E,
yield 30%, White powder, mp111–112 °C. 1H NMR (CDCl3):
d = 7.53 (d, J = 9 Hz, 1H, Ar-H), 7.22 (d, J = 8.4 Hz, 1H, Ar-H), 7.09
(s, 1H, Ar-H), 7.08 (dd, J1 = 7.8 Hz, J2 = 2.4 Hz, 1H, Ar-H), 7.01 (dd,
J1 = 8.4 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.94 (dd, J1 = 7.8 Hz, J2 = 2.4 Hz,
1H, Ar-H), 6.86 (s, 1H, Ar-H), 6.81 (d, J = 7.8 Hz, 1H, Ar-H), 6.72
(dd, J1 = 8.4 Hz, J2 = 2.4 Hz, 1H, Ar-H), 6.69 (dd, J1 = 7.8 Hz,
J2 = 1.8 Hz, 1H, Ar-H), 5.49 (s, 1H, OH), 5.47 (s, 1H,OH), 5.37 (s,
1H, OH), 5.20 (d, J = 1.8 Hz, 1H, Ar-H), 3.27–3.03 (m, 1H, Ph–
CH2CH2–Ph), 3.09–3.03 (m, 2H, Ph–CH2CH2–Ph), 3.00–2.94 (m,
1H, Ph–CH2CH2–Ph), 2.84–2.79 (m, 2H, Ph–CH2CH2–Ph), 2.50–
2.45 (m, 1H, Ph–CH2CH2–Ph), 1.91–1.86 (m, 1H, Ph–CH2CH2–Ph);
13C NMR (CDCl3): d = 155.93, 155.80, 153.04, 151.96, 142.78,
142.76, 142.75, 140.29, 139.17, 133.82, 133.80, 132.87, 130.76,
130.39, 129.89, 127.65, 124.41, 122.76, 121.63, 121.97, 121.42,
115.50, 114.76, 114.36, 35.44, 33.05, 30.85, 30.20; HRMS (ESI)
calcd for C28H26O4NBr2 600.2803; found 600.2805 (M+NH4+); MS
(ESI) 580, 582, 584 (MꢀH)ꢀ.
4.3. Biological evaluation
4.3.1. Cell culture and cytotoxicity assay
The human squamous cell carcinoma KB, human breast adeno-
carcinoma cell line MCF-7 and human human prostate cancer PC3
cells (The Cell Bank of Chinese Academy of Sciences, Shanghai)
were maintained in RPMI 1640 medium (Hyclone, Logan, USA)
supplemented with 10% fetal bovine serum (HyClone) and supple-
mented with 100 U/mL of penicillin and 100 lg/mL of streptomy-
cin. All cells were cultured in a humidified atmosphere of 5% CO2 at
37 °C. The cytotoxity of the candidate drug on tumor cells was
measured by MTT (3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-
phenytetra- zoliumromide) method as previously described.35

J. Jiang et al. / Bioorg. Med. Chem. 20 (2012) 2382–2391
2391
Briefly, after treatment of the candidate drug for 24 h, aborrbance
References and notes
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This work was supported by grant from the National Natural
Science Foundation (NNSF) of China (No.30925038) and Drug Devel-
opment Fund from Ministry of Science and Technology (ZX09102–
127).