Alkyl and carboxylalkyl esters of 4′-demethyl-4-deoxypodophyllotoxin: Synthesis, cytotoxic, and antitumor activity

Article abstract of DOI:10.1016/j.ejmech.2003.10.002

Esters of 4′-demethyl-4-deoxypodophyllotoxin (DDPT) with alkanoic acids and alkanedioic acids were prepared and tested for cytotoxic and antitumor activity. Among 19 esters, esters of propanoic acid, tetradecanedioic acid, 13-carboxyundecanoic acid, and h

Full text of DOI:10.1016/j.ejmech.2003.10.002

European Journal of Medicinal Chemistry 39 (2004) 189–193
www.elsevier.com/locate/ejmech
Short communication
Alkyl and carboxylalkyl esters of 4′-demethyl-4-deoxypodophyllotoxin:
synthesis, cytotoxic, and antitumor activity
Young-JaeYou,Yong Kim, Nguyen-Hai Nam, Seong-Cheol Bang, Byung-Zun Ahn *
College of Pharmacy, Chungnam National University, Taejon 305-764, South Korea
Received 20 May 2003; received in revised form 3 October 2003; accepted 3 October 2003
Abstract
Esters of 4′-demethyl-4-deoxypodophyllotoxin (DDPT) with alkanoic acids and alkanedioic acids were prepared and tested for cytotoxic
and antitumor activity. Among 19 esters, esters of propanoic acid, tetradecanedioic acid, 13-carboxyundecanoic acid, and hexadecanedioic
acid improved the antitumor activity compared with that of the starting compounds, DDPT.
© 2003 Elsevier SAS. All rights reserved.
Keywords: Deoxypodophyllotoxin; Esters; Cytotoxicity; Antitumor activity
1. Introduction
O
O
O
O
O
4-Deoxypodophyllotoxin (DPT) is a potent antimitotic
agent first isolated from Anthriscus sylvestris [1]. It was
O
O
O
shown to exhibit potent cytotoxicity against a wide variety of
tumor cell lines [2,3]. Recently, we reported the presence of
H3CO
OCH3
OCH3
H3CO
OCH3
DPT in Pulsatilla koreana, one indigenous plant in Korea,
and demonstrated that this antimitotic agent completely in-
hibited the tube-like formation of human umbilical venous
endothelial cells (HUVEC) at a concentration considerably
below the cytotoxic activity. Moreover, DPT also exhibited
significant antitumor activity in BDF1 mice bearing murine
Lewis lung carcinoma (3LL) cells with inhibition rate (IR) of
59% [4].
OH
DPT
DDPT
Fig. 1. Structures of DPT and of DDPT.
alkanoic acids and alkanedioic acids as alternative protecting
groups and tested their cytotoxic and antitumor activity.
Previously, it was reported that 4′-demethyl-4-
deoxypodophyllotoxin (DDPT) exerted a comparable in
vitro potency as DPT [5]. In vivo experiments revealed a
substantial loss of the antitumor activity of DDPT in the
BDF1/3LL model. We assumed that the free hydroxy group
at the 4′ position in DDPT was not favorable for antitumor
activity (Fig. 1). Indeed, in previous studies, we notified that
the esterification of the phenolic hydroxy group with car-
bamic, carbonic and amino acids increased the activity [6].
Now we would like to report the synthesis of esters of
2. Chemistry
The starting material, DPT, was isolated from A. sylvestris
(1.6 g DPT from 2 kg dried roots of A. sylvestris). 4′ Methoxy
group of DPT was selectively demethylated with trimethyl-
silyl iodide (yield, 72%), which afforded DDPT (Fig. 2).
Then, DDPT was esterified with various organic acids using
DCC and DMAP (yield, 70–91%). The products were con-
firmed by the triplet peaks of a-methylene group at around
2.5 ppm (–OCOCH2C–) on 1H NMR. Esters with dicarboxy-
lic acids were prepared by the similar methods (yield,
60–70%).
* Corresponding author. Present address: SBP Pharmaceutical
Company, Bonggok-Li 533-2, Banpo-Myun, Gongju-shi 314-923, South
Korea.
E-mail address: ahnbj@hanbat.chungnam.ac.kr (B.-Z. Ahn).
© 2003 Elsevier SAS. All rights reserved.
doi:10.1016/j.ejmech.2003.10.002

190
Y.-J. You et al. / European Journal of Medicinal Chemistry 39 (2004) 189–193
O
O
a
Anthriscus sylvestris
O
O
H3CO
OCH3
OCH3
b
O
O
O
O
O
O
c
O
O
H3CO
OCH3
R
H3CO
OCH3
O
OH
O
Fig. 2. (a) MeOH, 5 h, 3 times then silica gel column, cyclohexanes/EA (5:1); (b) TMSI, CH2Cl2, 0 °C, 5 h then BaCO3, 30 min; (c) DCC, DMAP, CH2Cl2, 0 °C,
1 h.
3. Biological results and discussion
from alkyl esters, esters with dicarboxylic acids seemed to
have an optimal chain length at least on A549 cells: esters
with sebacic acid to tetradecanedioic acid (DODE3–
DODE5) having stronger activity (ED50, 0.014–0.046 µg/ml)
than shorter or longer dioic acids.
In animal tests, unlike cytotoxic activity, esters with
shorter alkanoic acid showed weak antitumor activity except
DOE2 (Table 1). On the other hand, medium chain length
acids (DOE5–DOE10) (IR, 36–72%) were better for antitu-
mor activity. In particular, antitumor activity of DOE5 (IR,
72%) was comparable with the positive control, etoposide
(VP-16) (IR, 68%) at single injection (Fig. 3). This implies
that DOE5 is pharmacokinetically beneficial because it re-
Biological activities are reported in Tables 1 and 2. For
A549 cells, cytotoxic activity was dependent on chain length
of the alkyl moiety; the shorter were the alkyl chains, the
stronger were the cytotoxicities. Esters with acetic to oc-
tanoic acid (DOE1–DOE6; ED50, 0.003–0.013 µg/ml) were
more active than DDPT (ED50, 0.023 µg/ml), while esters of
longer alkanoic acids than hexadecanoic acid (DOE10–
DOE13) showed weaker activity (ED50, >2.670 µg/ml). A
similar tendency was observed against SK-MEL-2 cells. In-
terestingly, among the 13 esters, nine showed higher cyto-
toxic activity on A549 than on SK-MEL-2 cells. Different
Table 1
Cytotoxicities and antitumor activities of 4′-O-alkanoyl-DDPTs (DOE1–DOE13)
Compounds
R
ED50 (µg/ml)
A-549
0.003
0.004
0.005
0.006
0.013
0.009
0.027
0.030
0.041
0.268
2.670
>5
IR a (%)
Injection schedule
SK-MEL-2
0.006
0.010
0.017
0.089
0.078
0.030
0.004
0.025
0.050
0.290
>5
DOE1
DOE2
DOE3
DOE4
DOE5
DOE6
DOE7
DOE8
DOE9
DOE10
DOE11
DOE12
DOE13
Acetyl
11
70
21
25
72
45
63
57
36
40
–2
11
–8
NT b
<10
68
d1
Propanoyl
d1, d8
d1, d9
d1
Butanoyl
3-Methylbutanoyl
Heptanoyl
Octanoyl
d1
d1
Decanoyl
d1
Dodecanoyl
Tetradecanoyl
Hexadecanoyl
Octadecanoyl
Eicosanoyl
Docosanoyl
DPT
d1
d1, d8
d1, d5
d1, d5, d9
d1, d5, d9
d1, d5, d9
>5
>5
>5
0.012
0.023
1.102
0.009
0.015
NT b
DDPT
Etoposide (VP-16) c
d1, d5, d9
d1, d5, d9
a Inhibition rate with a dose of 60 mg/kg except VP-16.
b Not tested.
c VP-16 was treated at 36 mg/kg/day.

Y.-J. You et al. / European Journal of Medicinal Chemistry 39 (2004) 189–193
191
Table 2
Cytotoxicities and antitumor activities of 4′-O-carboxyalkanoyl-DDPTs (DODE1–DODE6)
Compounds
R
ED50 (µg/ml)
A-549
0.258
IR a (%)
Injection schedule
SK-MEL-2
0.062
0.066
0.015
0.068
0.070
0.027
0.009
0.015
NT b
DODE1
DODE2
DODE3
DODE4
DODE5
DODE6
5-Carboxypentanoyl
7-Carboxyheptanoyl
9-Carboxynonanoyl
11-Carboxydodecanoyl
13-Carboxyundecanoyl
15-Carboxypentadecanoyl (hexadecanedioic)
DPT
50
d1, d5, d9
d1, d5, d9
d1, d5, d9
d1, d5, d9
d1, d5, d9
d1, d5, d9
0.175
30
0.014
36
0.046
25
0.019
86
0.106
72
0.012
NT b
<10
84
DDPT
Etoposide (VP-16) c
0.023
d1, d5, d9
d1, d5, d9
1.102
a Inhibition rate with a dose of 60 mg/kg except VP-16.
b Not tested.
c VP-16 was treated at 36 mg/kg/day.
mains in the body for a longer amount of time than VP-16.
Among esters with alkanedioic acids, DODE5 (IR, 86%)
showed the most potent antitumor activity which is compa-
rable to VP-16 (IR, 84%) in a repeated animal test (Table 2).
More interesting is the pattern of tumor growth treated with
DODE5. As shown in Fig. 4, the antitumor activity of
DODE5 showed a plateau from the day 7 at around 0.3 cm–3
of tumor volume crossing the tumor volume treated with
VP-16 at the day 11. Considering the dosing frequency of
DOE5 and DODE5, it was assumed that the activities of two
compounds come from a different mechanism or pharmaco-
kinetic profile.
four esters of propanoic acid, tetradecanedioic acid, 13-
carboxyundecanoic acid, and hexadecanedioic acid showed a
promising antitumor activity.
4. Experimental protocol
1
The H NMR spectra were recorded on a JEOL EX-90
(90 MHz) NMR spectrometer, and proton chemical shifts are
relative to tetramethylsilane as an internal standard in CDCl3.
Mass spectra (MS) were collected on a Finnigan LCQ Ad-
vantage mass spectrometer using ion-trap MS with electro-
spray ionisation. All fractions from column chromatography
(EM Kiesegel 60, 230–400 mesh) were monitored by thin
layer chromatography (silica gel 60 GF-254, Merck, 230–
400 mesh ASTM). All reagents, commercially available,
were used without further purification unless otherwise
stated.
In conclusion, introducing ester group, we successfully
increased the in vivo antitumor activity of DDPT. Especially,
3.0
control
2.5
VP-16
DOE5
2.0
1.5
1.0
0.5
0.0
4.1. Isolation of deoxypodophyllotoxin
Air-dried A. sylvestris (roots, 2 kg) was extracted with 5 l
d6
d7
d8
d9
d10
d11
d12
d13
of ethyl acetate three times. The ethyl acetate extract was
Days after tumor transplantation
chromatographed over
a
silica gel column using
cyclohexane/ethyl acetate (5:1) as an eluent to afford the
crude DPT fraction. Recrystallization of the resulting crude
fraction from MeOH afforded DPT (1, 4.2 g), which was
confirmed by direct comparison with an authentic sample
isolated from P. koreana and spectral data reported previ-
ously [4]. DPT obtained as such was used as a starting
material for subsequent syntheses of the prodrugs.
Fig. 3. Inhibition of tumor growth in BDF1 mice by 4′-O-heptanoyl-DDPT
(DOE5).
3.0
control
VP-16
2.5
2.0
1.5
1.0
0.5
0.0
DODE5
4.2. Synthesis
4.2.1. Synthesis of 4′-demethyl-4-deoxypodophyllotoxin
(DDPT)
d5
d6
d7
d8
d9
d10
d11
d12
Days after tumor transplantation
The demethylation of DPT was performed under presence
of trimethylsilyl iodide according to the method in a refer-
ence [7].
Fig. 4. Inhibition of tumor growth in BDF1 mice by 4′-O-(13-
carboxyundecanoyl)-DDPT (DODE5).

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Y.-J. You et al. / European Journal of Medicinal Chemistry 39 (2004) 189–193
4.2.2. General procedures for the synthesis of esters
of DDPT
(1H, m), 3.66 (6H, s), 3.22–2.54 (4H, m), 2.56 (2H, t,
J = 7.4 Hz), 1.56–1.73 (2H, m), 1.30 (m, 8H), 0.87 (3H, t,
J = 3.4 Hz); MS: [M + H]+ 511.2 (calc. MS 510.2).
The
mixture
of
acid
(1.2
mmole),
1,3-
dicyclohexylcarbodiimide (1.2 mmole), 4-dimethylamino-
pyridine (0.4 mmole), and DDPT (1 mmole) in dried dichlo-
romethane (20 ml) were stirred for 2 h at 0 °C. The resulting
suspension was filtered, and distilled water (100 ml) was
added. After extracting with dichloromethane (100 ml) three
times, the organic layer was dried and concentrated to give
the crude product, which then was purified by silica gel
column with a mixture of cyclohexane and ethyl acetate
(4:1).
4.2.2.7.
4′-Demethyl-4′-O-decanoyl-4-deoxypodophyllo-
toxin (DOE7). Amorphous powder (87%); 1H NMR
(90 MHz, CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.66 (6H, s), 3.22–2.54 (4H, m), 2.56 (2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.23 (12H, m), 0.92 (3H, t,
J = 5.4 Hz); MS: [M + H]+ 539.3 (calc. MS 538.3).
4.2.2.8. 4′-Demethyl-4′-O-dodecanoyl-4-deoxypodophyllo-
toxin (DOE8). Amorphous powder (91%); 1H NMR
(90 MHz, CDCl3): d 6.65 (1H, s), 6.51 (1H, s), 6.37 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.66 (6H, s), 3.22–2.54 (4H, m), 2.56 (2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.23 (16H, m), 0.88 (3H, t,
J = 5.4 Hz); MS: [M + H]+ 567.3 (calc. MS 566.3).
4.2.2.1. 4′-Demethyl-4′-O-acetyl-4-deoxypodophyllotoxin
(DOE1). Amorphous powder (85%); H NMR (90 MHz,
CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37 (2H, s), 5.91(2H, br
s), 4.61(1H, br s), 4.56–4.32 (1H, m), 3.72–4.04 (1H, m),
3.66 (6H, s), 3.22–2.54 (4H, m), 2.29(3H, s); MS: [M + H]+
427.2 (calc. MS 426.1).
1
4.2.2.9. 4′-Demethyl-4′-O-tetradecanoyl-4-deoxypodophyl-
lotoxin (DOE9). Amorphous powder (90%); 1H NMR
(90 MHz, CDCl3): d 6.66 (1H, s), 6.53 (1H, s), 6.38 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.66 (6H, s), 3.22–2.54 (4H, m), 2.56 (2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.25 (20H, m), 0.88 (3H, t,
J = 5.4 Hz); MS: [M + H]+ 595.3 (calc. MS 594.3).
4.2.2.2. 4′-Demethyl-4′-O-propanoyl-4-deoxypodophyllotoxin
(DOE2). Amorphous powder (83%); H NMR (90 MHz,
1
CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37 (2H, s), 5.91(2H, br
s), 4.61(1H, br s), 4.56–4.32 (1H, m), 3.72–4.04 (1H, m),
3.66 (6H, s), 3.22–2.54 (4H, m), 2.27 (2H, q, J = 7.7 Hz),
1.24 (3H, t, J = 7.7 Hz); MS: [M + H]+ 440.1 (calc. MS
441.2).
4.2.2.10.
4′-Demethyl-4′-O-hexadecanoyl-4-deoxypodo-
4.2.2.3. 4′-Demethyl-4′-O-butanoyl-4-deoxypodophyllotoxin
(DOE3). Amorphous powder (90%); H NMR (90 MHz,
phyllotoxin (DOE10). Amorphous powder (85%); 1H NMR
(90 MHz, CDCl3): d 6.63 (1H, s), 6.50 (1H, s), 6.37 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.67 (6H, s), 3.22–2.54 (4H, m), 2.56 (2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.25 (24H, m), 0.87 (3H, t,
J = 5.4 Hz); MS: [M + H]+ 623.3 (calc. MS 622.4).
1
CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37 (2H, s), 5.91(2H, br
s), 4.61(1H, br s), 4.56–4.32 (1H, m), 3.72–4.04 (1H, m),
3.66 (6H, s), 3.22–2.54 (4H, m), 2.55(2H, t, J = 7.4 Hz), 1.77
(2H, m), 1.02 (3H, t, J = 7.4 Hz); MS: [M + H]+ 454.2 (calc.
MS 455.3).
4.2.2.11. 4′-Demethyl-4′-O-octadecanoyl-4-deoxypodophyl-
lotoxin (DOE11). Amorphous powder (78%); 1H NMR
(90 MHz, CDCl3): d 6.66 (1H, s), 6.52 (1H, s), 6.37 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.68 (6H, s), 3.22–2.54 (4H, m), 2.57(2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.25 (28H, m), 0.87 (3H, t,
J = 5.4 Hz); MS: [M + H]+ 651.3 (calc. MS 650.4).
4.2.2.4.
4′-Demethyl-4′-O-(3-methylbutanoyl)-4-deoxy-
1
podophyllotoxin (DOE4). Amorphous powder (88%); H
NMR (90 MHz, CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37
(2H, s), 5.91(2H, br s), 4.61(1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.66 (6H, s), 3.22–2.54 (4H, m), 2.24
(2H, d, J = 5.7 Hz), 2.10–2.22 (1H, m), 1.06 (3H, s), 0.99
(3H, s); MS: [M + H]+ 469.2 (calc. MS 468.2).
4.2.2.5. 4′-Demethyl-4′-O-heptanoyl-4-deoxypodophyllo-
toxin (DOE5). Amorphous powder (92%); 1H NMR
(90 MHz, CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.66 (6H, s), 3.22–2.54 (4H, m), 2.55 (2H, t,
J = 7.1 Hz), 1.55–1.72 (2H, m), 1.30 (4H, m), 0.88 (3H, t,
J = 6.0 Hz); MS: [M + H]+ 497.3 (calc. MS 496.2).
4.2.2.12. 4′-Demethyl-4′-O-eicosanoyl-4-deoxypodophyllo-
toxin (DOE12). Amorphous powder (79%); 1H NMR
(90 MHz, CDCl3): d 6.64 (1H, s), 6.50 (1H, s), 6.38 (2H, s),
5.90 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
(1H, m), 3.67 (6H, s), 3.22–2.54 (4H, m), 2.56 (2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.26 (32H, m), 0.87 (3H, t,
J = 5.4 Hz); MS: [M + H]+ 679.3 (calc. MS 678.4).
4.2.2.6.
4′-Demethyl-4′-O-octanoyl-4-deoxypodophyllo-
4.2.2.13. 4′-Demethyl-4′-O-docosanoyl-4-deoxypodophyllo-
toxin (DOE13). Amorphous powder (79%); 1H NMR
(90 MHz, CDCl3): d 6.66 (1H, s), 6.52 (1H, s), 6.38 (2H, s),
5.93 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04
toxin (DOE6). Amorphous powder (85%); 1H NMR
(90 MHz, CDCl3): d 6.65 (1H, s), 6.49 (1H, s), 6.37 (2H, s),
5.91 (2H, br s), 4.61 (1H, br s), 4.56–4.32 (1H, m), 3.72–4.04

Y.-J. You et al. / European Journal of Medicinal Chemistry 39 (2004) 189–193
193
(1H, m), 3.68 (6H, s), 3.22–2.54 (4H, m), 2.57 (2H, t,
J = 7.3 Hz), 1.55–1.72 (2H, m), 1.00–1.4 (36H, m), 0.88 (3H,
t, J = 5.4 Hz); MS: [M + H]+ 513.3 (calc. MS 512.2).
in culture medium) were seeded in each well of 96-well
plates. The plates were incubated in a 5% CO2 incubator at
37 °C for 24 h and then samples in a 20 µl culture medium
were added at various concentrations. The plates were incu-
bated for another 48 h. Cytotoxicity was measured by SRB
method as described in literature [8] with slight modifica-
tions [4,9]. The values shown for these compounds are aver-
ages of three determinations.
4.2.2.14.
4′-Demethyl-4′-O-(5-carboxypentanoyl)deoxy-
podophyllotoxin (DODE1). Amorphous powder (70%);
1H NMR (90 MHz, CDCl3): d 6.65 (1H, s), 6.51 (1H, s), 6.36
(2H, s), 5.93 (2H, br s), 4.63 (1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.67 (6H, s), 3.02–2.5 (6H, m), 2.30–2.50
(2H, m), 1.70–1.90 (4H, m); MS: [M + H]+ 541.2 (calc. MS
540.2).
4.2.4. Antitumor experiments
Antitumor experiments were carried out as described in
our previous reports [4,9]. Briefly, 3LL cells were inoculated
subcutaneously into BDF1 mice on the day 0 (1 × 107
cells/mouse/0.2 ml PBS). Esters were dissolved in a medium
comprising of 5% DMSO and 20% Cremophor® to make a
concentration of 6 mg/ml. 0.2 ml of the solution was intrap-
eritoneally injected per mouse (20 g) corresponding to a dose
of 60 mg/kg/day, and the injection schedules were adjusted
by the change of body weight as in Tables 1 and 2. Body
weights of mice were tracked every day and tumor sizes were
measured with calipers from day 9. Tumor volumes were
calculated by the following equation: tumor volume
(mm3) = (length (mm) × width (mm)2)/2. The inhibition rate
was evaluated as (1–T/C) × 100% (where T is the mean tumor
volume of the treated group and C is the mean tumor volume
of the control group). Each group consisted of six mice.
Etoposide was administered at day 1, 5, and 9 at
36 mg/kg/day.
4.2.2.15. 4′-Demethyl-4′-O-(7-carboxyheptanoyl)-4-deoxy-
1
podophyllotoxin (DODE2). Amorphous powder (65%); H
NMR (90 MHz, CDCl3): d 6.65 (1H, s), 6.50 (1H, s), 6.36
(2H, s), 5.92 (2H, br s), 4.59 (1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.67 (6H, s), 3.02–2.50 (4H, m), 2.56
(2H, t, J = 7.1 Hz), 2.33 (2H, t, J = 7.1 Hz), 1.55–1.80 (2H,
m), 1.31 (8H, m); MS: [M + H]+ 569.2 (calc. MS 568.2).
4.2.2.16. 4′-Demethyl-4′-O-(9-carboxynonanoyl)-4-deoxy-
1
podophyllotoxin (DODE3). Amorphous powder (63%); H
NMR (90 MHz, CDCl3): d 6.65 (1H, s), 6.51 (1H, s), 6.36
(2H, s), 5.92 (2H, br s), 4.59 (1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.67 (6H, s), 3.02–2.50 (4H, m), 2.56
(2H, t, J = 7.1 Hz), 2.33 (2H, t, J = 7.1 Hz), 1.55–1.80 (2H,
m), 1.31 (12H, m); MS: [M + H]+ 597.3 (calc. MS 596.3).
4.2.2.17.4′-Demethyl-4′-O-(11-carboxyundecanoyl)-4-deoxy-
podophyllotoxin (DODE4). Amorphous powder (62%);
1H NMR (90 MHz, CDCl3): d 6.66 (1H, s), 6.52 (1H, s), 6.37
(2H, s), 5.92 (2H, br s), 4.60 (1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.66 (6H, s), 3.02–2.50 (4H, m), 2.57
(2H, t, J = 7.1 Hz), 2.34 (2H, t, J = 7.1 Hz), 1.55–1.80 (2H,
m), 1.31 (16H, m); MS: [M + H]+ 624.3 (calc. MS 625.3).
Acknowledgements
This work was supported by SBP Pharmaceutical Com-
pany.
4.2.2.18. 4′-Demethyl-4′-O-(13-carboxytridecanoyl)-4-deoxy-
podophyllotoxin (DODE5). Amorphous powder (60%);
1H NMR (90 MHz, CDCl3): d 6.66 (1H, s), 6.52 (1H, s), 6.38
(2H, s), 5.93 (2H, br s), 4.60 (1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.69 (6H, s), 3.02–2.50 (4H, m), 2.57
(2H, t, J =7.1 Hz), 2.34 (2H, t, J = 7.1 Hz), 1.55–1.80 (2H,
m), 1.31 (20H, m); MS: [M + H]+ 653.4 (calc. MS 652.3).
References
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4.2.2.19. 4′-Demethyl-4′-O-(15-carboxyheptadecanoyl)-4-
deoxypodophyllotoxin (DODE6). Amorphous powder (63%);
1H NMR (90 MHz, CDCl3): d 6.66 (1H, s), 6.52 (1H, s), 6.37
(2H, s), 5.93 (2H, br s), 4.63 (1H, br s), 4.56–4.32 (1H, m),
3.72–4.04 (1H, m), 3.68 (6H, s), 3.02–2.50 (4H, m), 2.57
(2H, t, J =7.1 Hz), 2.33 (2H, t, J =7.1 Hz), 1.55–1.80 (2H, m),
1.31 (24H, m); MS: [M + H]+ 681.3 (calc. MS 680.4).
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4.2.3. Cytotoxicity
Tumor cells were maintained in plastic dishes in a RPMI-
1640 medium supplemented with 10% fetal bovine serum.
On day 0, 180 µl of a tumor cell suspension (3 × 104 cells/ml