Antitumor agents. Part 214: Synthesis and evaluation of curcumin analogues as cytotoxic agents

Article abstract of DOI:10.1016/S0968-0896(02)00249-3

Fifty-eight curcumin analogues were prepared and evaluated for in vitro cytotoxicity against a panel of human tumor cell lines. Compound 50 was the most potent analogue against several cell lines, including HOS (bone cancer) and 1A9 (breast cancer), with ED₅₀ values of 0.97 and <0.63 μg/mL, respectively.

Full text of DOI:10.1016/S0968-0896(02)00249-3

Bioorganic & Medicinal Chemistry 10 (2002) 3481–3487
y
Antitumor Agents. Part 214: Synthesis and Evaluation of
Curcumin Analogues as Cytotoxic Agents
a
b
b
Junko Ishida, Hironori Ohtsu, Yoko Tachibana,
b
b
a
Yuka Nakanishi, Kenneth F. Bastow, Masahiro Nagai,
b
b
b,
Hui-Kang Wang, Hideji Itokawa and Kuo-Hsiung Lee *
aHoshi University, 2-4-41 Ebara, Shinagawa, Tokyo 158-0098, Japan
bNatural Products Laboratory, School of Pharmacy, University of North Carolina, Chapel Hill, NC 27599, USA
Received 28 November 2001; accepted 14 June 2002
Abstract—Fifty-eight curcumin analogues were prepared and evaluated for in vitro cytotoxicity against a panel of human tumor
cell lines. Compound 50 was the most potent analogue against several cell lines, including HOS (bone cancer) and 1A9 (breast
cancer), with ED50 values of 0.97 and <0.63 mg/mL, respectively.
#
2002 Elsevier Science Ltd. All rights reserved.
Introduction
exhibited potent antitumor promoting effects on
12-O-tetradecanoylphorbol-13-acetate (TPA)-induced
mouse skin carcinogenesis. Furthermore, 17 inhibited
papilloma formation initiated by peroxynitrite.1
The rhizome of Curcuma longa L. (turmeric) has been
widely used as a yellow coloring agent and spice in
many foods, and it has also been used in indigenous
medicine for the treatment of inflammatory and other
3
2
diseases. Curcumin (1), a well-known acyclic dia-
In a continuing search for potent and selective cytotoxic
antitumor agents, we prepared 32 curcumin analogues
(1–32) and evaluated their cytotoxic effects against a
panel of human tumor cell lines. We also evaluated 26
b-diketone compounds (33–58) that are structurally
related to curcumin (1). In addition, the effects of selec-
ted compounds on drug resistance were evaluated using
a drug resistance reversal assay.
rylheptanoid, has been identified as the major con-
stituent in turmeric. Recently, numerous studies have
demonstrated the remarkable cancer preventive proper-
2À13
ties of curcumin (1).
The chemopreventive effects of
curcumin have been attributed to various biological
properties, including neutralization of carcinogenic free
3
radicals and anti-angiogenesis action, which limits the
blood supply to rapidly growing malignant cells.4,5 In
addition, curcumin possesses other pharmacological
properties, including stimulation of Phase I and Phase
II detox systems, (e.g., inhibition of COX-1 and COX-2
Chemistry
6
,7
enzymes, and stimulation of glutathione S-transferase).
In addition, curcumin exhibited remarkable cytotoxic
As shown in Figure 1, compounds 1, 2, and 3 were
obtained by column chromatography (silica gel, CHCl3–
MeOH) of commercially available (Aldrich) curcumin,
which contained 2 and 3 as minor components. Com-
pounds 4–6 were prepared by heating 1–3 overnight with
histidine hydrazide, acetic acid, and p-TsOH. Acetylation
of 1 gave 7. Compound 8 was obtained by demethylation of
1 with AlCl and pyridine in CH Cl . Compound 9 was
8
À10
effects on various cancer cells
cell death in human promyelocytic leukemia HL-60 cells
and induced apoptotic
1
1
and human oral squamous carcinoma HSC-4 cells.
12
In our previous paper, we reported that 13-oxomyricanol
16) and myricanone (17), two cyclic diarylheptanoids,
(
3
2
2
obtained by treating 1 with methyl iodide. Hydrogenation
of 1, 9, and 50 with Pd/C gave 10–13 and 51. Preparation of
yFor Paper 213, See ref 1.
12
14–23 was described in a previous paper. Compounds 24–
2 were synthesized from the appropriate benzaldehyde
*
3
Corresponding author. Tel.: +1-919-962-0066; fax: +1-919-966-
893; e-mail: khlee@unc.edu
3
0
968-0896/02/$ - see front matter # 2002 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(02)00249-3

3482
J. Ishida et al. / Bioorg. Med. Chem. 10 (2002) 3481–3487
and 2,4-pentanedione by the method reported by Ped-
1
dihydroxy (8) and trimethyl (9) derivatives, respectively,
significantly increased cytotoxic activity against 1A9,
KB and HCT-8 cell lines. Thus, the presence of catechol
or 3,4-dimethoxyphenyl constituents enhanced the
cytotoxic properties. (c) Saturation of the olefinic bonds
of 1 and 9 to give 10–13 abolished activity. In addition,
all cyclic diarylheptanopids (14–23) were inactive.
Therefore, the presence of the conjugated b-diketone in
the acyclic carbon chain appears to play an important
role for cytotoxicity in this class. (d) Fluorination at the
ortho position of both benzene rings (26) increased
activity. Introducing a methoxy group at the 4- (31) or
6- (32) position of both fluorinated rings decreased
cytotoxicity. Additional fluorination (30) or fluorination
solely (24) at the para position abolished activity. Thus,
the position and nature of substituents on the benzene
rings seem to modulate antitumor activity.
4
ersen et al. Compounds 33–50 and 52–58 were
obtained from Aldrich Inc. (Milwaukee, WI, USA).
Results and Discussion
Cytotoxicity of diarylheptanoids
Thirty-two curcumin derivatives (1–32) were tested for
cytotoxicity against a panel of human tumor cell lines.
Selected results for active compounds are summarized
in Table 1. Curcumin hydrazide (4) showed a broad
cytotoxicity spectrum (ED50 values from 1.0 to 3.9 mg/
mL against KB, A549, CAKI-1, MCF-7, 1A9, HCT-8,
SK-MEl-2, U-87-MG, HOS, PC-3, KB-VIN, HepG2,
and LNCaP (clone FGC) cell lines. Demethylcurcumin
(8) was selectively active against KB cells with an ED50
value of 1.0 mg/mL, and trimethylcurcumin (9) was also
active against a narrow spectrum of cell lines (1A9,
U87-MG, and HOS). All hydrogenated (10–13) and
cyclic (14–23) diarylheptanoids were inactive. Com-
pound 26, which is o-fluorinated on each benzene ring,
had a broad cytotoxicity spectrum, but borderline
activity, while the remaining fluorinated diarylhepta-
noids (24, 25, and 27–32) were inactive.
Cytotoxicity of ꢀ-diketones (1,3-diaryl-1,3-diketo-propane)
Twenty-six 1,3-diaryl-1,3-diketo-propane and related
analogues (33–58) were also tested for cytotoxicity
against the human tumor cell line panel. The ED50
values of the active compounds are summarized in
Table 1. Compound 38 (b-diketone) displayed moderate
cytotoxicity, whereas 34 (b-triketone) and 36 (b-tetra-
ketone) were inactive. Compound 44 (4-chlorophenyl)
was also more active than 35, the corresponding trike-
tone. Thus, it would appear that the b-diketone moiety
enhances the cytotoxic properties. Compound 40 (4-
tert-butylated phenyl) was more potent than 38 (unsub-
stituted) and 39 (3,5-dimethylphenyl) against 1A9 cells
selectively. Compound 41 (2,4-dimethoxyphenyl)
showed no activity. Thus, introducing the tert-butyl
group (an electron-donating substituent) on the phenyl
ring led to increased cytotoxicity against the 1A9 cell
By comparing the cytotoxicity results in Table 1, the
following structure–activity relationships (SARs) were
drawn. (a) Converting the keto-enol moiety (1–3) to the
corresponding pyrazole (4–6) led to increased cytotoxi-
city against various cell lines. Compound 4 was active in
all cell lines, whereas, compounds 5 and 6 exhibited
selective activity against the 1A9 cell line. Thus, the ring
substituents affected the activity in the pyrazole deriva-
tives. (b) Demethylation or methylation of 1 to form the
Table 1. Selected in vitro cytotoxicity of curcumin analogues
a
Compd
ED50 (mg/mL)
KBb A549 CAKI-1 MCF-7
1A9
>20
HCT-8 SK-MEL-2 U-87-MG HOS
PC-3 KB-VIN HepG2 LNCaP clone FGC
1
4
5
6
8
9
2
3
3
4
4
4
4
4
4
5
5
5
>20
1.0
9.0 >20
10.7 20
>20
>20
2.4
ND
ND
>20
3.9
8.8
13.5
9.5
>20
1.9
7.0
10.0
ND
2.2
6.3
7.5
7.5
10.0
12.0
5.8
8.0
9.7
>20
2.4
13.6
14.1
>20
2.4
14.0
>20
ND
3.0
6.1
>20
>20
>20
16.3
>20
>20
>20
>20
ND
10.2
10.5
ND
>20
2.8
>20
ND
ND
2.5
ND
2.0
1.7
1.8
4.2
1.2
3.8
1.1
3.9
6.0
4.8
3.9
4.0
13.0 >20
15.0 9.5
ND ND
6.7
6.1
>20
ND
ND
ND
5.4
6.9
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
1.0
7.9
4.3
8.0 >10
7.5
6.0
>10
ND
7.6
6.5
4.7
4.6
7.4
>20
>20
>20
16.0
>20
>20
>20
4.0
4.9
5.2
5.8
>20
13.0
6
8
9
0
2
3
4
5
6
0
1
7
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
>20
>20
>20
16.5 >20
20
19.5 >20
19.7
15.5
3.8
14.4
4.5
20
20
>20
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
ND
20
14.5
>20
15
13.5 <10
>20 5.0
17.5
15.0
7.0
16.7
4.0
<10
<10
16.0 >20
8.0 16.0
>10 >20
>20
>20
>20
16.5
>20
19.5
>20
15.0
4.5
>20
7.4
16.5
9.0 >20
10.0 >20
17.5
18
4.4
18.8
8.9
<2.5
3.9
<0.63
13.6
2.1
6.0
1.8
18.3
3.0
7.0
2.0
>20
6.0
15.0
12.0
0.97
18.0 >20
<2.5 8.0
14
5.0
3.8
18.8
7.7
ND, not determined.
a
Cytotoxicity as ED50 for each cell line, the concentration of compound that causes 50% reduction in absorbance at 562 nm relative to untreated
cells using the SRB assay.
b
The human cell line panel consisted of epidermoid carcinoma of the nasopharynx (KB), lung carcinoma (A549), renal cancer (CAKI-1), breast
cancer (MCF-7), ovarian cancer (1A9), ileocecal carcinoma (HCT-8), melanoma (SK-MEL-2), glioblastoma (U-87-MG), bone cancer (HOS), liver
cancer (HepG2), and prostate cancer (PC-3, LNCaP clone FGC). The human KB drug-resistant sub-line panel included KB-VIN (expressing P-
glycoprotein).

J. Ishida et al. / Bioorg. Med. Chem. 10 (2002) 3481–3487
3483
line. Replacing the hydrogen atom with fluorine (an
Conclusions
electron-withdrawing substituent) at the para position
on the benzene rings (45) increased activity against 1A9
cells compared with unsubstituted 38 and 4-chlor-
ophenyl compounds (43 and 44). b-Bromination
between the keto groups (42, 46, and 50) led to
enhanced activity compared with the unsubstituted
compounds (39, 38, and 49) against 1A9 cells. However,
nitroso (47), benzoyl methyl (53), and furan (56) sub-
stitution at this position abolished activity.
In summary, modification of curcumin produced com-
pounds with potential for further development as anti-
cancer agents. Based on these preliminary screening
results, compounds 45, 50, and 57 showed significant
activity in certain cancer cell lines and have been tar-
geted for further studies. Additional research, including
mode of action studies, is planned to accurately estab-
lish relative activity for SAR and rational design.
Compound 50, which has an a-bromo substituent and
Recently, the cancer chemopreventive effects of curcu-
min have been intensively investigated. Curcumin
exhibited pronounced antitumor activity by triggering
apoptosis in human tumor cells. Studies are underway
to investigate the apoptosis-inducing activity of com-
pounds found to be cytotoxic in this study.
4
-nitro and 4-methoxy groups on separate benzene
rings, demonstrated the strongest cytotoxic effects
against HOS and 1A9 cells with ED50 values of 0.97 and
<0.63 mg/mL, respectively. However, compound 51,
which has a 4-amino rather than the 4-nitro group,
1
3
showed decreased activity, and compound 48, which has
4
-nitro groups on both benzene rings, was inactive.
Thus, asymmetrical substitution led to enhanced activ-
ity and different electronegative aryl substituents
Experimental
General
(
4-nitro and 4-methoxy) led to increased activity against
A9 and HOS cells selectively. Compound 50 is highly
1
electrophilic and would be expected to be a DNA-
interactive agent.
Melting points were determined on a Fisher–Johns
1
melting apparatus and are uncorrected. H NMR spec-
tra were recorded on a Varian Gemini-300 spectro-
meter. The chemical shifts are presented in terms of
ppm with TMS as the internal reference. MS spectra
were recorded on an HP5989A instrument. Elemental
analyses were performed by Atlantic Microlab Inc.,
Norcross, GA, USA
In addition, replacing the phenyl groups in 46 with
thiophenyl groups (57) increased cytotoxicity against
HOS and 1A9 cell lines (ED50 values of<2.5 and 2.1
mg/mL, respectively).
Inhibitory activity of curcumin analogues on
P-glycoprotein mediated drug resistance
ꢀ
Curcumin (1). Yellow needles, mp 181–183 C (EtOH)
ꢀ
1
5
(
lit. 184–185 C).
Six compounds (1, 2, 3, 4, 17, and 21) were also assayed
for vincristine toxicity potentiation in tumor cells
Demethoxycurcumin (2). Orange crystalline powder, mp
ꢀ
ꢀ
16
(
KB-VIN) possessing P-glycoprotein mediated drug
173–175 C (CHCl ) (lit. 175–177 C).
3
resistance. Interestingly, the cyclic ketal 21 inhibited
P-glycoprotein in a dose-dependent manner in the pre-
sence of vincristine; the corresponding ketone (17) was
less active. Although 21 was less active than verapamil
as judged by potentiation of vincristine toxicity (Table
Bisdemethoxycurcumin (3). Orange solid, mp 216–
218 C (lit. 209–211 C). The structures of 1–3 were
ꢀ
confirmed by comparing their physical and spectral data
17
ꢀ
1
5À17
with those reported in the literature.
2
), it was also less toxic. Additional studies are planned
with this initial novel lead for potential modulation of
multidrug resistance.
General procedure for the preparation of pyrazole deriva-
tives (4–6). To a solution of starting material in butanol
and EtOH were added histidine hydrazide, HOAc, and
TsOH. The solution was refluxed for 24 h, then the sol-
vent was removed in vacuo. The residue was purified by
silica gel column chromatography and plc.
Table 2. Effects of co-treatment with verapamil or 21 on vincristine
toxicity
3
,5-Bis[ꢀ-(4-hydroxy-3-methoxyphenyl)-ethynyl]pyrazole
Treatment
ED50
Potentiation
b
ꢀ
lit. 211–214 C). The structure of 4 was confirmed by
(
(
4). Yellow amorphous solid (yield 22%), mp 215 C
a
(
mM)
index
1
8
ꢀ
comparing experimental and literature physical and
Verapamil
30.5
NAc
NA
13.9
>2
21
>40.0
2.2
20
18
spectral data.
d
Verapamil + 0.125 mM Vincristine
1 + 0.125 mM Vincristine
2
Compound 5. Yellow amorphous solid (yield 10%), mp
184–185 C; H NMR (500 MHz, acetone-d ) d 3.78
ꢀ
(3H, s, OCH ) 6.53 (1H, s, pyrazole 4-H), 6.70 (1H, d,
1
aConcentration for 50% inhibition of KB-VIN cell growth in presence
or absence of a non-growth inhibitory concentration of vincristine.
6
3
,
b
The ratio of ED50 without vincristine treatment to the ED50 with
vincristine.
0
0
0
J=8.1 Hz, aryl-6 ), 6.72 (2H, d, J=8.6 Hz, aryl-3 ,5 ),
6.83 (1H, d, J=16.4 Hz, CH=CH–aryl), 6.86 (1H, d,
c
Not applicable. The isolation and characterization of KB-VIN
0
(1H, d, J=16.4 Hz, CH=CH–aryl), 7.00 (1H, d,
J=16.4 Hz, CH=CH–aryl), 6.86 (1H, bd, aryl 5 ), 6.99
(
expressing P-glycoprotein) are described in ref 16.
d
0.125 mM Vincristine is not toxic to this cell line.

3484
J. Ishida et al. / Bioorg. Med. Chem. 10 (2002) 3481–3487
Figure 1.
ꢀ
by comparing experimental and literature physical and
15
spectral data.
J=16.4 Hz, CH=CH–aryl), 7.07 (1H, d, J=1.9 Hz,
aryl 2 ), 6.99 (2H, d, J=8.8 Hz, aryl-2 ,6 ); EI–MS m/z:
(55%). Mp 142–145 C. The structure of 9 was confirmed
0
34 (M ), HR FABMS m/z 334.1316 (M ) (calcd for
0
0
+
+
3
C H N O : 334.1317).
2
0
18
2
3
Hydrogenation of curcumin (1). A solution of 1 (55.3
mg) in MeOH (2 mL) was hydrogenated for 4 h at room
temperature and 1 atm over 10% Pd/C catalyst. The
filtered solution was concentrated and separated by
silica gel column chromatography [benzene/ AcOEt
Compound 6. White amorphous solid (yield 7%), mp
ꢀ
1
2
72–273 C; H NMR (500 MHz, acetone-d ) d 6.53
6
0
(
1H, s, 1-H), 6.72 (4H, d, J=8.5 Hz, aryl-3bm5,5 ), 6.83
2H, d, J=16.5 Hz, CH=CH–aryl), 6.99 (2H, d,
(
J=16.5 Hz, CH=CH–aryl), 7.28 (4H, d, J=8.5 Hz,
(8:1)!(1:1)] and plc [CHCl /MeOH (6:1)] to yield 10,
3
0
04.1203 (M ) (calcd for C H N O : 304.1212).
0
+
aryl 2 ,6 ); EI–MS m/z: 304 (M ); HR FABMS m/z
+
11, and 12. The structures of 10–12 were confirmed by
comparing their physical and spectral data with those
3
1
9
16
2
2
1
5
reported in the literature.
Diacetylcurcumin (7). A solution of curcumin in anhy-
drous pyridine was treated with anhydrous Ac O and
Tetrahydrocurcumin (10). White amorphous solid (yield
ꢀ
2
21
ꢀ
stirred overnight. After a general workup procedure, a
yellow amorphous solid was obtained. Compound 7
14%), mp 92–93 C (lit. 95–96 C).
1
was identified on the basis of H NMR data comparison
1
Hexahydrocurcumin (11). White amorphous solid (yield
ꢀ
5
21
ꢀ
with literature values.
9%), mp 87–88 C (lit. 78–80 C).
1
(
,7-Bis(3,4-dihydroxyphenyl)-11,6-heptadiene-3,5-dione
8). Yellow powder, compound 8 was prepared by
heating 1 with AlCl , pyridine in CHCl by the method
Octahydrocurcumin (12). Colorless oil (yield 5%).
Compound 13. A solution of 9 (546 mg) in EtOAc (100
mL) was hydrogenated overnight at room temperature
and 45 psi over 10% Pd/C catalyst. The filtered solution
was concentrated and separated by silica gel column
chromatography [benzene/ EtOAc (10:1)] and plc
3
2
1
in ref. 19. Compound 8 was identified on the basis of H
1
9
NMR data comparison with literature values.
Trimethylcurcumin (9). Curcumin in dry acetone with
methyl iodide was refluxed over anhydrous K CO for
4
[CHCl / MeOH (6:1)] to yield 13. Colorless oil (yield
3
2
3
1
8 h with stirring. After filtration, the solvent was eva-
7%); H NMR (300 MHz, CDCl ) d 0.95 (3H, d,
3
porated and the residue purified by silica gel column
chromatography to yield 9 as a yellow amorphous solid
CHCH ), 1.52 (1H, m, CHCH ), 1.84 (2H, m, CHaHb-
CHOH), 2.67 (6H, m, CHaHbCH ), 3.83 (14H, bs,
3
3
2

J. Ishida et al. / Bioorg. Med. Chem. 10 (2002) 3481–3487
3485
Figure 2.
OCH , CHOH), 6.78 (6H, m, aryl H). EI–MS m/z: 418
3
added dropwise over 15 min. Stirring was continued for
18 h at 40 C. The mixture was hydrolyzed by adding 10
+
+
ꢀ
mL of 1 N HCl and heating at 60 C for 1 h. Two layers
(M ); HR FABMS m/z: 418.236618 (M ) (calcd for
C H O : 418.2355392).
ꢀ
2
4
34
6
were separated, and the aqueous layer was extracted
three times with EtOAc. The combined organic layers
were washed until neutral, dried (Na SO ), and the sol-
vent was removed in vacuo. The crude products were
purified by silica gel column chromatography using an
n-hexane–EtOAc solvent system.
Compound 51. A solution of 50 (70 mg) in EtOAc (50
mL) was hydrogenated overnight at room temperature
and 45 psi over 10% Pd/C catalyst. The filtered solution
was concentrated and separated by silica gel column
chromatography [benzene/EtOAc (100:1)!(10:1)] and
2
4
plc [benzene/EtOAc (10:1)] to yield 51. White powder
1
ꢀ
(
CHBr), 4.02 (3H, s, OCH ), 6.83 (2H, d, J=9 Hz, aryl
yield 31%); H NMR (300 MHz, CDCl ) d 4.00 (1H, s,
Compound 24. Yield 5%, yellow needles, mp 167–169 C
1
3
(n-hexane–EtOAc); H NMR (CDCl ) d 5.81 (1H, enol
3
3
0
Hz, aryl 2 ,6 ), 8.09 (2H, d, J=9 Hz, aryl 2 ,6 ); EI–MS
0
0
0
3
,5 ), 7.11 (2H, d, J =9 Hz, aryl 3 ,5 ), 7.98 (2H, d, J=9
CH), 6.54 (2H, d, J=16.0 Hz, CH=CH–aryl), 7.09
(4H, m, aryl 3 ,5 ), 7.54 (4H, m, aryl 2 ,6 ), 7.63 (2H, d,
+
0
0
0
0
m/z: 348 (M ), HR FABMS m/z: 348.0299 (calcd for
0
0
0
J=16.0 Hz, CH=CH–aryl). EI–MS m/z 312 (M ).
0
+
C H O NBr: 348.2035).
1
Anal. calcd for C H F O : C 73.07, H 4.52, found: C
19 14
6
15
3
2
2
72.81, H 4.52.
Preparation of fluorinated diarylheptanoids (24–32). The
1
4
ꢀ
EtOAc); H NMR (CDCl ) 5.84 (1H, s, enol CH), 6.60
(2H, d, J=15.0 Hz, CH=CH–aryl), 7.06 (2H, m, aryl
procedure reported by Pedersen et al. was applied with
compounds 24–32. In general, 2,4-pentanedione (0.5
equiv) and boric anhydride (0.35 equiv), dissolved in 10
Compound 25. Yield 6%, mp 110–111 C (n-hexane–
1
3
ꢀ
0
0
Hz, CH=CH–aryl). FABMS m/z 313 (MH ). Anal.
0
0
mL of EtOAc, were stirred for 30 min at 40 C. Benzal-
2 ), 7.22–7.39 (6H, m, aryl 4 ,5 ,6 ), 7.60 (2H, d, J=15.0
+
dehyde (1 equiv) and tributylborate (1 equiv) were
added, and the mixture was stirred for 30 min. Butyla-
mine (0.75 equiv), dissolved in 10 mL of EtOAc, was
calcd for C H F O : C 73.07, H 4.52, found: C 72.87,
2
1
9
14
2
H 4.50.

3486
J. Ishida et al. / Bioorg. Med. Chem. 10 (2002) 3481–3487
Figure 3.
ꢀ
0
aryl), 7.72 (2H, m, aryl 6 ), 7.79 (2H, d, J=6.6 Hz, aryl
2 ). FABMS m/z 449 (MH ). Anal. calcd for
Compound 26. Yield 5%, mp 110–113 C (n-hexane–
1
0
C H F O : C 56.26, H 2.70, found: C 56.06, H 2.72.
+
EtOAc); H NMR (CDCl ) d 5.89 (1H, s, enol CH),
6.75 (2H, d, J=16.0 Hz, CH=CH–aryl), 7.08–7.21 (4H,
m, aryl CH), 7.35 (2H, m, aryl CH), 7.57 (2H, m, aryl
3
2
1
12
8
2
ꢀ
CH), 7.78 (2H, d, J=16.0 Hz, CH=CH–aryl). FABMS
+
Compound 29. Yield 10%, mp 124–126 C (n-hexane–
1
m/z 313 (MH ). Anal. calcd for C H F O : C 73.07,
1
EtOAc); H NMR (CDCl ) d 5.58 (1H, s, enol CH), 6.60
9
14
2
2
3
H 4.52, found: C 72.89, H 4.45.
(2H, d, J=16.0 Hz, CH=CH–aryl), 7.24 (4H, m, aryl
2
0
CH=CH–aryl). FABMS m/z 445 (MH ). Anal. calcd
0
0
0
6 ), 7.58 (4H, m, aryl 3 ,5 ), 7.65 (2H, d, J=16.0 Hz,
+
ꢀ
Compound 27. Yield 31%, mp 174–175 C (n-hexane–
1
EtOAc); H NMR (acetone-d ) d 3.92 (6H, s, OCH ),
for C H F O : C 56.77, H 3.18, found: C 56.87, H 3.14.
6
6
3
21 14
4
6
CH=CH-aryl), 7.18 (2H, t, J=8.4 Hz, aryl 5 ), 7.43–
7
.02 (1H, s, enol CH), 6.76 (2H, d, J=16.0 Hz,
0
.55 (4H, m, aryl 2 ,6 ), 7.58 (2H, d, J=16.0 Hz,
+
ꢀ
Compound 30. Yield 3%, mp 155–157 C (n-hexane–
1
0
CH=CH–aryl). FABMS m/z 373 (MH ). Anal. calcd
0
EtOAc); H NMR (CDCl ) d 5.86 (1H, s, enol CH),
3
6.68 (2H, d, J=16.0 Hz, CH=CH-aryl), 6.84–6.96 (4H,
m, aryl 5 ,6 ), 7.56 (2H, m, aryl 3 ), 7.71 (2H, d, J=16.0
+
0
Hz, CH=CH–aryl). FABMS m/z 349 (MH ). Anal.
0
0
for C H F O : C 67.74, H 4.87, found: C 67.76, H
4
2
1
18
2
4
.87.
calcd for C H F O : C 65.52, H 3.47, found: C 65.61,
1
H 3.5.
9
12
4
2
ꢀ
Compound 28. Yield 6%, mp 172–174 C (n-hexane–
1
EtOAc); H NMR (CDCl ) d 5.86 (1H, s, enol CH),
6.60 (2H, d, J=16.0 Hz, CH=CH–aryl), 7.24 (2H, t,
0
J=9.0 Hz, aryl 5 ), 7.63 (2H, d, J=16.0 Hz, CH=CH–
3
ꢀ
Compound 31. Yield, 10%, mp 162–164 C (n-hexane–
1
EtOAc); H NMR (CDCl ) d 3.84 (6H, s, OCH ), 5.82
3
3

J. Ishida et al. / Bioorg. Med. Chem. 10 (2002) 3481–3487
3487
(
6
3
1H, s, enol CH), 6.62 (2H, d, J=16.0 Hz, CH=CH–aryl),
0
References and Notes
0
.62–6.75 (4H, m, aryl 5 ,6 ), 7.48 (2H, t, J=8.4 Hz, aryl
), 7.71 (2H, d, J=16.0 Hz, CH=CH–aryl). FABMS m/z
0
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373 (MH ). Anal. calcd for C H F O : C 67.74, H 4.87,
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2
1
18 2 4
ꢀ
Compound 32. Yield 9%, mp 144–146 C (n-hexane–
1
EtOAc); H NMR (CDCl ) d 3.82 (6H, s, OCH ), 5.90
3
3
(
aryl), 6.88 (2H, m, aryl 4 ), 7.00–7.06 (4H, m, aryl 3 ,5 ),
7
3
4
1H, s, enol CH), 6.72 (2H, d, J=16.0 Hz, CH=CH–
0
9
5
0
0
.74 (2H, d, J=16.0 Hz, CH=CH–aryl). FABMS m/z
+
73 (MH ). Anal. calcd for C H F O : C 67.74, H
2
1
18
2
4
.87, found: C 67.60, H 4.96.
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2
0
the procedures described in Rubinstein et al. Drug
stock solutions were prepared in DMSO, and the final
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(
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1
The human tumor cell line panel constituted of epi-
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press.
(
(
MCF-7), ovarian cancer (1A9), ileocecal carcinoma
HCT-8), melanoma cancer (SKMEl-2) and glio-
blastoma (U-87-MG), bone cancer (HOS), liver cancer
HepG2), and prostate cancer (PC-3 and LNCaP clone
(
1
4. Pedersen, U.; Rasmussen, P. B.; Lawesson, S. O. Lebigs
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1
For drug-resistance reversal, KB-VIN cells cultured in
the presence or absence of 125 nM vincristine (a con-
centration without effect on cell growth) were used.
Cells were exposed to doses of verapamil or compound
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1
K.; Itokawa, H. Chem. Pharm. Bull. 1987, 3298.
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index (Table 2) was calculated.
1
Pertz, H.; Eich, E.; Pommier, Y. J. Med. Chem. 1997, 40,
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Acknowledgements
3
2
057.
0. Rubinstein, L. V.; Shoemaker, R. H.; Paull, K. D.; Simo,
This research was supported by a grant from the National
Cancer Institute (CA 17625) awarded to K. H. Lee.
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Boyd, M. R. J. Natl. Cancer Inst. 1990, 82, 1113.