Recombination of diterpenoid structure units: Synthesis of antitumor amides bearing functionalized bicyclo[3.2.1]octane ring

Article abstract of DOI:10.1016/j.bmcl.2010.05.075

In this work, 23 new amides (14-36) bearing a representative diterpenoid structure unit, the functionalized bicyclo[3.2.1]octane ring, have been synthesized and its antitumor potential is studied. In vitro studies demonstrate that a number of amides with the bicyclo[3.2.1]oct-3-en-2-one subunit are active against HL-60, SMMC-7721, A-549, SK-BR-3, and PANC-1 tumor cell lines. The hybrid derivative, compound 20, was found to be the most potent compound (IC₅₀ = 1.05 μM against HL-60) and more active than cisplatin (DDP), the positive control. Additionally, compound 20 exhibited broad spectrum in vitro anticancer activity with IC₅₀ values of 1.1-4.3 μM against the five tested cancer cell lines.

Full text of DOI:10.1016/j.bmcl.2010.05.075

Bioorganic & Medicinal Chemistry Letters 20 (2010) 4116–4119
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Recombination of diterpenoid structure units: Synthesis of antitumor
amides bearing functionalized bicyclo[3.2.1]octane ring
Zewei Mao a, Yan Li b, Jingbo Chen a, Yuanyuan Wang b, Hongbin Zhang a,
*
a Key Laboratory of Medicinal Chemistry for Natural Resource (Yunnan University), Ministry of Education, School of Chemical Science and Technology, Yunnan University,
Kunming, Yunnan 650091, PR China
b State Key Laboratory for Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Science, Kunming, Yunnan 650204, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
In this work, 23 new amides (14–36) bearing a representative diterpenoid structure unit, the functional-
ized bicyclo[3.2.1]octane ring, have been synthesized and its antitumor potential is studied. In vitro stud-
ies demonstrate that a number of amides with the bicyclo[3.2.1]oct-3-en-2-one subunit are active against
HL-60, SMMC-7721, A-549, SK-BR-3, and PANC-1 tumor cell lines. The hybrid derivative, compound 20,
Received 18 March 2010
Revised 14 May 2010
Accepted 18 May 2010
Available online 8 June 2010
was found to be the most potent compound (IC50 = 1.05
(DDP), the positive control. Additionally, compound 20 exhibited broad spectrum in vitro anticancer
activity with IC50 values of 1.1–4.3 M against the five tested cancer cell lines.
Ó 2010 Elsevier Ltd. All rights reserved.
lM against HL-60) and more active than cisplatin
Keywords:
Amide
Diterpenoid structure unit
Bicyclo[3.2.1]octane ring system
Antitumor activity
l
Natural diterpenes constitute a large family of important bioac-
tive compounds. Among them, the tetracyclic ent-kaurenoids are
important source of antitumor agents and have attracted the atten-
tion of numerousresearch groups.1 A number of antitumor ent-kaur-
enoid diterpenes ( Fig. 1) have been reported in recent years,
The synthesis of amides containing the functionalized bicy-
cle[3.2.1]octane framework commenced with protection of 1,3-
cyclohexandione. Treatment of commercially available 1,3-cyclo-
hexandione with isopropanol in reflux benzene in the presence of
p-toluene sulfonic acid resulted in the 3-isoproxyl-2-cyclohexenone
(6). Enolizationofcompound6withlithiumdiisopropyl amide(LDA)
followed by carbonylation with methyl chloroformate provided
intermediate 7 (82%). Allylationof compound7 in thepresenceof so-
dium hydride afforded compound 8 (91%). After reduction with lith-
including eriocalyxin B (1),2 oridonin (2),3 and ludongnin J (3).4
A
representative structure unit presented in those diterpenoid com-
pounds is the functionalized bridged C/D bicyclo[3.2.1]octane ring
system. In the preceding Letter5 we reported the synthesis and bio-
logicalevaluation of a series of gibberellin derivativesand foundthat
gibberellins bearing two
a,b-unsaturated ketone units are potent
O
OH
antitumor agents. It is noteworthy that a functionalized bicy-
clo[3.2.1]octane carbon framework is also presented in our antitu-
mor gibberellins. As an especially interesting example, the highly
bioactive platensimycin (Fig. 1, compound 4), an amide derived from
bacterial oxidation of ent-kaurene,6 is also endowed with the bicy-
clo[3.2.1]octane skeleton (Fig. 1). Because the structure of platensi-
mycin can be viewed as a hybrid of diterpenoid structure unit with
an amine by an amide linkage, we were inspired to initiatea research
program towards the recombination of diterpenoid structure unit,
with the aim of generation of antitumor amides on the basis of the
functionalized bicyclo[3.2.1]octane ring system (Scheme 1). Herein
we report the synthesis of bicyclo[3.2.1]oct-3-en-2-one ring con-
taining amides and its in vitro cytotoxic properties against a number
of human tumor cell lines.
H
H
OH
O
O
O
O
OH
OH
H
H
OH
OH
Eriocalycin B (1)
Oridonin (2)
OH
O
O
H
H
HOOC
N
H
OH
O
H
O
O
O
OMe
H
O
Ludongnin J (3)
Platensimycin (4)
* Corresponding author. Tel.: +86 871 5031119; fax: +86 871 5035538.
E-mail addresses: zhanghb@ynu.edu.cn, zhanghbyd@gmail.com (H. Zhang).
Figure 1. Representative molecules of tetracyclic diterpenes (1–3) and platensi-
mycin (4).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.05.075

Z. Mao et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4116–4119
4117
OH
O
O
R1
N
R1
N
R1
N
R2
R2
R2
OH
O
O
O
O
Bicyclo[3.2.1]octane ring system
O
O
OiPr
OiPr
O
O
O
O
O
Scheme 1. Synthetic plan towards amides bearing a functionalized bicyclo[3.2.1]octane ring.
O
Oi-Pr
O Pr
i-
O
TsOH, PhH
isopropanol
reflux
1, LDA, THF
-78 oC
2, ClCOOMe
Oi-Pr
NaH, THF
oC, then
Allyl bromide
LiAlH4, THF
0 oC, then
10% HClO4
0
88%
82%
O
O
MeO
O
91%
O
55%
MeO
O
HO
O
5
9
6
7
8
MOMCl, DIPEA
CH2Cl2, rt
86%
O
O
O
O
1, LDA, THF, -78 oC, then
HMPA and TBDMSCl, rt
6N HCl,
MeOH, rt
1, IBX, EtOAc
reflux
2, Jones reagent
acetone
91%
2, 5% Pd(OAc)2, O2
DMSO, 50 o
64%
C
63%
OMOM
O
OH
OH
OMOM
12
13
11
10
Scheme 2. Synthesis of 6-methylene-4-oxobicyclo[3.2.1]oct-2-ene-1-carboxylic acid 13.
Table 1
Synthesis of primary amides bearing functionalized bicycle[3.2.1]octane ringa,b
O
O
a: DCC, DMAP, CH2Cl2, rt
H
N
RNH2
+
HO
or b: SOCl2, Et3N, CH2Cl2,
rt
R
O
O
Amide
R
Yield (%)
14
15
16
17
18
19
20
21
22
23
24
25
26
2-(4-Methoxy-phenyl)-ethyl
Phenyl
79a
72a
81a
82a
71a
69a
67b
73b
77b
81b
78b
62b
79b
4-Methoxy-phenyl
4-Bromo-phenyl
2-Bromo-4-methyl-phenyl
4-Bromo-2-fluoro-phenyl
3,4-Dichloro-phenyl
2-Chloro-phenyl
3-Methoxy-phenyl
2,4,6-Trimethyl-phenyl
2,6-Dimethyl-4-bromo-phenyl
4-Acetyl-phenyl
t-Butyl
a
Method a: Treatment of acid with DCC in the presence of DMAP in dichloromethane for 30 min (rt) then addition of amine at room temperature; Method b: Treatment of
acid with SOCl2 in dichloromethane for 30 min then addition of amine and Et3N at room temperature.
b
Yields represent isolated yields.

4118
Z. Mao et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4116–4119
O
O
O
1, NaH, THF
2, Methyl iodide or Allyl bromide
H
N
N
N
16: R = 4-methoxy-phenyl
17: R = 4-bromo-phenyl
R
R
R
O
O
19: R = 4-Bromo-2-Fluoro-phenyl
O
27 (77%)
28 (78%)
29 (74%)
30 (79%)
31 (88%)
32 (72%)
Ce Cl3-7H2O
Na BH4, MeOH
16: R = 4-methoxy-phenyl
19: R = 4-Bromo-2-Fluoro-phenyl
OH
OH
O
SeO2, t-BuOOH
CH2Cl2
IBX, EtOAc
reflux
H
N
H
N
H
N
R
R
R
OH
O
O
O
O
33 (89%)
34 (85%)
35 (61%)
36 (59%)
Scheme 3. Synthesis of amides bearing functionalized bicycle[3.2.1]octane ring.
Table 2
In vitro cytotoxic activities of amides 17–41 (IC50, l
M)a,b
Amides
Cell lines
A-549
HL-60
SMMC-7721
SK-BR-3
PANC-1
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
DDP
1
>40
>40
>40
>40
3.78 0.60
5.74 0.91
5.41 1.52
1.19 0.19
>40
>40
>40
6.08 1.03
>40
>40
>40
>40
>40
>40
>40
22.10 1.35
>40
>40
>40
>40
14.31 1.30
0.43 0.27
>40
>40
>40
4.62 1.79
15.49 1.06
11.87 5.10
3.16 0.68
>40
>40
>40
16.64 0.57
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
15.23 0.35
1.40 0.81
>40
>40
>40
10.03 1.40
15.51 0.24
11.27 0.25
4.30 0.26
>40
>40
>40
14.42 3.79
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
14.77 0.54
0.73 0.21
>40
>40
>40
16.59 0.43
13.09 0.45
2.80 0.42
4.40 2.20
3.45 0.10
1.05 0.65
6.85 0.35
>40
14.11 0.35
8.44 3.18
13.6 0.25
>40
>40
>40
>40
20.51 0.45
>40
7.66 0.05
15.14 0.46
7.10 2.12
3.14 0.18
19.80 0.35
>40
15.22 1.25
16.63 0.22
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
>40
21.87 6.62
0.74 0.04
15.20 0.15
>40
>40
>40
>40
1.42 0.35
0.66 0.14
a
Cytotoxicity as IC50 values for each cell line, the concentration of compound that caused 50% reduction in absorbance at 570 nm relative to untreated cells using the MTT
assay.
b
Human promyelocytic leukemia (HL-60), human hepatocellular carcinoma (SMMC-7721), human lung adenocarcinoma (A-549), human breast adenocarcinoma (SK-BR-
3), human pancreatic carcinoma (PANC-1).
ium aluminium hydride and subsequent acidification with per-
chloric acid, cyclohexenone 9 was obtained in 55% yield. Following
the well established Ihara’s protocol,7 the key intermediate (11)
was prepared in 64% yield over two steps (Scheme 2). Acid catalyzed
hydrolysis of 11 resulted in alcohol 12 (91%), which provided acid 13
(63% in two steps) when exposed to oxidation first with IBX (o-
iodoxybenzoic acid) then with John’s reagent.8
available amines.9 The synthesized amides are summarized in
Table 1.
To get further insight toward the structure and activity relation-
ship, a few tertiary amides (27–32) were synthesized (Scheme 3).
Two amides (33, 34), without the a,b-unsaturated ketone system,
were also prepared by reduction of compound 16 and 19, respec-
tively under Luche reduction condition.10 Two amides (35 and
With the desired acid (13) in hand, we began the synthesis of
amides by treatment of acid 13 with a number of commercially
36) containing two
pared (Scheme 3) from the corresponding amide 33 and 34 by
a,b-unsaturated ketone moiety were also pre-

Z. Mao et al. / Bioorg. Med. Chem. Lett. 20 (2010) 4116–4119
4119
allylic hydroxylation with selenium dioxide (SeO2, CH2Cl2, t-
References and notes
BuOOH) and finally oxidation with IBX (IBX, EtOAc).
1. (a) Garcia, P. A.; de Oliveira, A. B.; Batista, R. Molecules 2007, 12, 455; (b) Sun,
H.-D.; Huang, S.-X.; Han, Q.-B. Nat. Prod. Rep. 2006, 23, 673.
2. Wang, L.; Zhao, W.-L.; Yan, J.-S.; Liu, P.; Sun, H.-P.; Zhou, G.-B.; Weng, Z.-Y.; Wu,
W.-L.; Weng, X.-Q.; Sun, X.-J.; Chen, J.; Sun, H.-D.; Chen, S.-J. Cell Death Differ.
2007, 14, 306.
3. Meade-Tollin, L. C.; Wijeratne, E. M. K.; Cooper, D.; Guild, M.; Jon, E.; Fritz, A.;
Zhou, G. X.; Whitesell, L.; Liang, J. Y.; Gunatilaka, A. A. L. J. Nat. Prod. 2004, 67, 2.
4. Han, Q.-B.; Zhao, A.-H.; Zhang, J.-X.; Lu, Y.; Zhang, L.-L.; Zheng, Q.-T.; Sun, H.-D.
J. Nat. Prod. 2003, 66, 1391.
5. Chen, J.; Sun, Z.; Zhang, Y.; Zeng, X.; Qing, C.; Liu, J.; Li, L.; Zhang, H. Bioorg. Med.
Chem. Lett. 2009, 19, 5496.
Next, the cytotoxic properties of all newly synthesized amides
were evaluated in vitro against five human tumor cell lines (includ-
ing HL-60, SMMC-7721, A-549, SK-BR-3, PANC-1) by MTT assay,11
cisplatin (DDP) and eriocalycin B (1) were used as reference drugs.
The results are summarized in Table 2 (IC50 value, defined as the con-
centrations corresponding to 50% growth inhibition). To our delight,
compound 17 (IC50 = 2.80 0.42
(IC50 = 3.45 0.10 M), 20 (IC50 = 1.05 0.65
0.35 M) and 24 (IC50 = 8.44 3.18 M) showed potential activities
lM), 18 (IC50 = 4.41 2.20
lM), 19
l
lM), 21 (IC50 = 6.85
l
l
6. For a review, see: Tiefenbacher, K.; Mulzer, J. Angew. Chem., Int. Ed. 2008, 47,
2548.
against myeloid leukaemia (HL-60); Compound 20 displayed potent
orsimilarcytotoxicactivityinvitrocomparedtoDDPanderiocalycin
B, and also possessed potent activities against SMMC-7721, A-549,
SK-BR-3 and PANC-1 cells. Although secondary amides (compound
15–25, except amide 22) derived from aniline derivatives were
shown to have certain level of activity against myeloid leukaemia
(HL-60), secondary amide 14 and 26, prepared from alkyl amines,
were both inactive in this assay. In comparison with secondary
amides, tertiary amides (27–32) were generally less potent than its
secondary amide counterparts (16, 17 and 19) against all tumor cell
lines investigated. The loss of potency in tertiary amides is possibly
due to the loss of a hydrogen bond donor (CONH bond). The fact that
compound 33 and 34 were both inactive suggests the requirement of
7. Recent examples: (a) Toyota, M.; Sasaki, M.; Ihara, M. Org. Lett. 2003, 5, 1193;
(b) Toyota, M.; Odashima, T.; Wada, T.; Ihara, M. J. Am. Chem. Soc. 2000, 122,
9036; A recent review: (c) Toyota, M.; Ihara, M. Synlett 2002, 1211.
8. Acid 13: colorless oil, 1H NMR (CDCl3, 300 MHz) d: 10.98 (s, 1H), 7.48–7.51 (dd,
J = 1.5 Hz, 1.5 Hz, 1H), 5.90–5.93 (dd, J = 1.5 Hz, 1.5 Hz, 1H), 5.34 (s, 1H), 5.13 (s,
1H), 3.58 (d, J = 1.6 Hz, 1H), 2.97–3.03 (m, 1H), 2.60 (d, J = 15.8 Hz, 1H), 2.31–
2.41 (m, 2H); 13C NMR (CDCl3, 75 MHz) d: 197.8, 178.6, 151.8, 142.7, 127.0,
113.7, 57.9, 51.6, 43.4, 41.3. EIMS m/z (%): 179 (M++1, 21%), 178 (M+, 68%), 160
(50), 150 (51), 133 (72), 132 (68), 105 (100), 91 (58), 79 (74), 77 (76). HRMS m/z
Found: 178.0626, Calcd for C10H10O3 (M)+: 178.0630.
9. Amide 17: White powder, mp 133.1–134.2 °C. 1H NMR (CDCl3, 300 MHz) d:
8.01 (s, 1H), 7.38–7.45 (m, 5H), 5.88 (dd, J = 1.3, 9.9 Hz, 1H), 5.29 (s, 1H), 5.12 (s,
1H), 3.52 (d, J = 4.5 Hz, 1H), 3.00 (dt, J = 2.4, 16.0 Hz, 1H), 2.64 (d, J = 16.0 Hz,
1H), 2.36 (dd, J = 2.1, 10.8 Hz, 1H), 2.25–2.33 (m, 1H). 13C NMR (CDCl3, 75 MHz)
d: 197.3, 170.7, 152.8, 142.6, 136.6, 132.0, 127.4, 122.2, 117.7, 113.8, 58.1, 53.8,
42.9, 40.4. EIMS m/z (%): 333 (M++2, 74%), 331 (M+, 74%), 305 (10), 303 (11),
288 (5), 280 (13), 278 (14), 250 (2), 224 (4), 199 (5), 198 (5), 172 (23), 170 (23),
160 (19), 145 (10), 133 (61), 119 (25), 105 (100), 91 (66), 79 (51), 77 (77).
HRMS m/z Found: 331.0175, Calcd for C16H14NO2Br (M)+: 331.0208. Amide 18:
White plates, mp 146.2–146.9 °C 1H NMR (CDCl3, 300 MHz) d: 8.15 (d,
J = 8.4 Hz, 1H), 7.82 (s, 1H), 7.46 (dd, J = 2.0, 9.9 Hz, 1H), 7.37 (d, J = 1.2 Hz,
1H), 7.14 (d, J = 8.4 Hz, 1H), 5.97 (dd, J = 1.3, 9.9 Hz, 1H), 5.36 (s, 1H), 5.17 (s,
1H), 3.61 (d, J = 4.8 Hz, 1H), 3.04 (dt, J = 2.4, 16.0 Hz, 1H), 2.71 (d, J = 16.0 Hz,
1H), 2.46 (dd, J = 2.1, 10.8 Hz, 1H), 2.30–2.40 (m, 1H), 2.31 (s, 3H). 13C NMR
(CDCl3, 75 MHz) d: 196.7, 170.4, 152.0, 142.6, 136.1, 132.6, 132.4, 129.2, 127.7,
122.1, 114.0, 113.8, 58.0, 54.0, 43.0, 40.6, 20.6. EIMS m/z (%): 347 (M++2, 84%),
345 (M+, 84%), 332 (2), 330 (2), 319 (27), 317 (27), 303 (27), 294 (37), 292 (38),
267 (22), 266 (80), 238 (34), 212 (17), 210 (15), 187 (51), 185 (57), 184 (36),
161 (32), 160 (40), 149 (12), 134 (59), 133 (85), 132 (50), 119 (48), 105 (100),
104 (57), 91 (53), 78 (68), 77 (85). HRMS m/z Found: 345.0370, Calcd for
an
system (see Table 2). To our surprise, however, presence of two
,b-unsaturated ketone moiety did not appear to improve potency
a,b-unsaturated ketone moiety in the bicyclo[3.2.1]octane ring
a
(Table 2, compound 35 and 36). Even though the results are preli-
minary,wefounda cleartrendinthestructure–activityrelationship:
thea,b-unsaturatedketonemoietyis necessaryforpotency,withthe
best result being coupled with halogen containing anilines. Among
the five tested cell lines, HL-60 exhibited more sensitivity to the rest
of tumor cell lines. Particularly, HL-60 cells and SMMC-7721 cells
were 3–4-fold more sensitive to amide 20 than the rest of tumor cell
lines. Compound 20 was shown to have promising antiproliferative
activities against a broad spectrum of human tumor cell lines from a
diverse set of target organs, including leukemia and solid tumors (li-
ver, lung, breast and pancreas cancer cell lines).
In conclusion, we have designed and synthesized a number of
amides bearing bicycle[3.2.1]octane ring system. We also evalu-
ated their antitumor activities by in vitro MTT assay. This study
has revealed that the bicyclo[3.2.1]oct-3-en-2-one moiety is a po-
tential cytotoxic pharmacophore, in particular, the amide 20 is a
promising lead due to the cytotoxic potencies displayed. It is also
of interests to note that amides bearing bicyclo[3.2.1]oct-3-en-2-
one moiety are more selective towards HL-60 cell line. Further bio-
logical investigation is currently underway in our laboratory and
the results will be reported in due course.
C
17H16NBrO2 (M)+: 345.0364. Amide 19: grey powder, mp 123.2 °C 1H NMR
(CDCl3, 300 MHz) d: 8.18 (t, J = 8.4 Hz, 1H), 7.58 (s, 1H), 7.43 (dd, J = 2.1, 9.9 Hz,
1H), 7.27–7.33 (m, 2H), 5.98 (dd, J = 1.5, 9.9 Hz, 1H), 5.37 (s, 1H), 5.18 (s, 1H),
3.61 (d, J = 4.8 Hz, 1H), 3.04 (dt, J = 2.4, 16.0 Hz, 1H), 2.71 (dd, J = 1.2, 16.0 Hz,
1H), 2.44 (dd, J = 2.1, 10.8 Hz, 1H), 2.30–2.40 (m, 1H). 13C NMR (CDCl3, 75 MHz)
d: 196.5, 170.6, 154.1 (150.8), 151.5, 142.3, 127.9, 125.1 (124.9), 123.3, 118.8,
118.5, 116.9 (116.8), 114.0, 57.9, 53.9, 42.9, 40.6. EIMS m/z (%): 351 (M++2,
80%), 349 (M+, 80%), 336 (10), 334 (11), 323 (32), 321 (32), 308 (17), 306 (18),
298 (36), 296 (37), 270 (12), 268 (9), 256 (2), 242 (8), 228 (3), 217 (11), 216
(11), 202 (7), 190 (43), 189 (46), 188 (45), 161 (93), 160 (50), 149 (27), 134 (51),
133 (85), 132 (51), 119 (51), 105 (100), 103 (56), 91 (54), 81 (39), 79 (72), 77
(80). HRMS m/z Found: 349.0109, Calcd for C16H13NBrFO2 (M)+: 349.0114.
Amide 20: pale pink plates, mp 135.4–136.8 °C 1H NMR (CDCl3, 300 MHz) d:
7.97 (s, 1H), 7.76 (d, J = 2.1 Hz, 1H), 7.34–7.45 (m, 3H), 5.91 (dd, J = 1.5, 9.9 Hz,
1H), 5.31 (s, 1H), 5.15 (s, 1H), 3.55 (d, J = 4.6 Hz, 1H), 3.03 (dt, J = 2.4, 16.0 Hz,
1H), 2.67 (d, J = 16.0 Hz, 1H), 2.28–2.42 (m, 2H). 13C NMR (CDCl3, 75 MHz) d:
197.2, 170.8, 152.4, 142.3, 136.9, 132.8, 130.5, 128.2, 127.5, 122.2, 119.7, 114.0,
58.0, 53.8, 42.9, 40.4. EIMS m/z (%): 323 (M++2, 78%), 322 (M++1, 42%), 321 (M+,
86%), 308 (6), 306 (9), 295 (29), 293 (40), 280 (13), 278 (22), 270 (30), 268 (40),
253 (5), 242 (8), 240 (11), 213 (11), 188 (10), 187 (9), 163 (30), 162 (32), 161
(91), 160 (79), 149 (23), 145 (21), 133 (100), 132 (55), 125 (18), 119 (50), 109
(17), 105 (94), 103 (49), 91 (52), 79 (67), 77 (75). HRMS m/z Found: 321.0329,
Calcd for C16H13NCl2O2 (M)+: 321.0323.
Acknowledgments
This work was supported by the Natural Science Foundation of
China(20832005, 20925205), the National Basic Research Program
of China (973 Program 2009CB522300) and the Natural Science
Foundation of Yunnan Province (2007B0006Z).
10. Gemal, A. L.; Luche, J. L. J. Am. Chem. Soc. 1981, 103, 5454.
11. The cytotoxicity assay was in five kinds of cell lines (HL-60, SMMC-7721, A-
549, SK-BR-3, PANC-1). Cells were cultured at 37 °C under
a humidified
atmosphere of 5% CO2 in RPMI 1640 medium supplemented with 10% fetal
serum and dispersed in replicate 96-well plates. Compounds were then added.
After 48-h exposure to the compounds, cells viability were determined by the
[3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide] (MTT)
cytotoxicity assay by measuring the absorbance at 570 nm with a microplate
spectrophotometer. Each test was performed in triplicate.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.05.075.