Asymmetric synthesis of phenanthroindolizidine alkaloids with hydroxyl group at the C14 position and evaluation of their antitumor activities

Article abstract of DOI:10.1016/j.bmcl.2010.11.008

The asymmetric total synthesis of the strongly cytotoxic phenanthroindolizidine alkaloid 3 was achieved. Using the same route, various derivatives were also synthesized. Cytotoxicity of those synthetic compounds was evaluated and compounds 19, 23, and 27

Full text of DOI:10.1016/j.bmcl.2010.11.008

Bioorganic & Medicinal Chemistry Letters 21 (2011) 342–345
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Asymmetric synthesis of phenanthroindolizidine alkaloids with hydroxyl group
at the C14 position and evaluation of their antitumor activities
⇑
Takashi Ikeda , Takashi Yaegashi, Takeshi Matsuzaki, Syusuke Hashimoto, Seigo Sawada
Yakult Central Institute for Microbiological Research, 1796 Yaho, Kunitachi-shi, Tokyo 186-8650, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
The asymmetric total synthesis of the strongly cytotoxic phenanthroindolizidine alkaloid 3 was achieved.
Using the same route, various derivatives were also synthesized. Cytotoxicity of those synthetic com-
pounds was evaluated and compounds 19, 23, and 27 demonstrated potent cytotoxicities similar to that
of 3. The in vivo antitumor efficacy of selected compounds was also evaluated and 23 demonstrated mod-
erate antitumor efficacy.
Received 27 August 2010
Revised 28 October 2010
Accepted 1 November 2010
Available online 5 November 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Asymmetric synthesis
Phenanthroindolizidine alkaloid
Antitumor
Phenanthroindolizidine alkaloids, isolated mainly from plants
of the Asclepiadaceae family, are known to possess various biolog-
ical activities including anti-arthritis,1 antifungal,2 anti-inflamma-
tory,3 and antitumor4 activities. In particular, their significant
antitumor activities make them attractive candidates for novel
antitumor agents. Tylophorine (1) and tylocrebrine (2) are repre-
sentative of this group of alkaloids (Fig. 1).5 Compound 3 was
recently isolated from a butterfly (Ideopsis similis) and has exhib-
ited remarkable cytotoxicity against various human cancer cell
lines.6 The structural features of 3 include the presence of a
hydroxyl group at the C14 position. This compound has attracted
attention due to its unique origin, strong cytotoxicity, and struc-
tural features. Another unique aspect of this compound is that
its related phenanthroindolizidine alkaloids are suggested to have
a different antitumor mechanism from those of other antitumor
agents that are currently being used clinically.4,7 In this study,
we report the asymmetric total synthesis of 38 and its derivatives
together with their in vitro cytotoxicities and in vivo antitumor
activities.
successfully attained the asymmetric total synthesis of 3. We also
synthesized various derivatives of 3 using the same route.
Synthesis of 3 commenced with the condensation of commer-
cially available 4-benzyloxybenzaldehyde (4) and 3,4-dimethoxy-
benzyl cyanide (5) to the stilbene (6) (Scheme 1). Buckley and
Rapoport demonstrated that 2,3,6,7-substituted phenanthrene
(so-called tylophorine-type) could be obtained by vanadium(V)
oxidation of the corresponding stilbene; however, 2,3,6-substi-
tuted phenanthrene could not be synthesized in the same manner.
Although Compound 3 is 3,6,7-substituted phenanthrene rather
than 2,3,6-substituted phenanthrene, there is a possibility that
vanadium(V) oxidation will not work because of the lack of tyloph-
orine-type substituents. Therefore, we adopted the photo-induced
electron cyclization reaction to construct the phenanthrene ring
because of its feasibility.12 Subsequently, we successfully obtained
the desired phenanthrene (7) by this route.
Conversion of the nitrile (7) to the aldehyde (8) followed by the
reductive amination with the glutamic acid unit produced the par-
tially racemized 12 contrary to the report (Scheme 2, path A).11
Thus, we changed the method to introduce the glutamic acid unit
from the reductive amination of the aldehyde to the amination of
the bromide (11) (path B). Fortunately, racemization was not
observed via path B.
Total synthesis of phenanthroindolizidine alkaloids has been re-
ported by various groups;9 however, those with stereoselectively
constructed 14-hydroxyl groups are rare.10 Buckley and Rapoport
reported an asymmetric total synthesis of (S)-tylophorine (ent-1),
and during this process they synthesized 14-
a-hydroxyl (S)-tyl-
Hydrolysis of 12 produced the amido acid 13 with a good yield
(Scheme 3). An intramolecular Friedel–Crafts acylation was com-
pleted successfully but partial cleavage of the 3-benzylether to
the corresponding 3-hydroxyl compound was also observed.13
The constructed carbonyl moiety was reduced to alcohol diastere-
ophorine stereoselectively as an intermediate product.11 In this
study, we made few modifications to their synthetic route and
oselectively using L-selectride. Reduction of the lactam moiety fol-
lowed by hydrogenolysis of the benzylether afforded 3. Synthetic 3
was confirmed as a single enantiomer by chiral HPLC analysis and
⇑
Corresponding author. Tel.: +81 42 577 8957; fax: +81 42 577 3020.
E-mail address: takashi-ikeda@yakult.co.jp (T. Ikeda).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.11.008

T. Ikeda et al. / Bioorg. Med. Chem. Lett. 21 (2011) 342–345
343
Figure 1. Chemical structures of phenanthroindolizidine alkaroids.
Scheme 1. Reagents and conditions: (a) NaOEt, EtOH (93%); (b) I2, propylene oxide, hm, CH3CN (79%).
was spectroscopically identical to the compound in a previous re-
port.6 Subsequently, 13% overall yield of Compound 3 was obtained
starting from 4 with 12-steps. By changing the starting material
appropriately, we also obtained derivatives of 3 that have different
substituents on the phenanthrene ring.14
The compounds synthesized were evaluated for their cytotoxic-
ities against three human cancer cell lines, KB (nasopharyngeal),
A549 (lung), and HT-29 (colorectal) (Table 1).15 Compound 3
exhibited potent cytotoxicity against all cell lines as previously re-
ported.6 Compounds 17 and 18 are derivatives of 3 that differed
from 3 in the position of the hydroxyl group from R2 to R1 or R3.
Although 3 exhibited potent cytotoxicity against all cell lines
tested, these two compounds exhibited cytotoxicities that were
cell line-dependent. This result stimulated us to probe the efficacy
of the substituent at R2. Therefore, we changed the hydroxyl group
of 3 to either a methoxy group (19), hydrogen atom (20), fluorine
atom (22), ethyl group (21), or acetoxy group (23) and evaluated
their cytotoxicities. Compounds 19 and 23 exhibited comparable
activities to 3, while 20 and 22 were found to be less active. In
contrast, Compound 21 showed a remarkable decrease in activity.
Scheme 2. Reagents and conditions: (c) DIBAL, CH2Cl2 (86%); (d) (
L)-diisopropyl glutamate, NaCNBH3, CH2Cl2; (e) NaBH4, MeOH, 1,4-dioxane; (f) PBr3, Et3N, CHCl3; (g) (L)-
diisopropyl glutamate, K2CO3, benzene, DMF; (h) AcOH, MeOH (via Path A: 88%, two-steps, via Path B: 84%, four-steps).

344
T. Ikeda et al. / Bioorg. Med. Chem. Lett. 21 (2011) 342–345
Scheme 3. Reagents and conditions: (i) KOHaq, MeOH, 1,4-dioxane (99%); (j) (COCl)2, DMF, CH2Cl2, then SnCl4 (49%); (k) L-selectride, THF (82%); (l) BH3–THF, THF (93%); (m)
H2, Pd-C, MeOH (62%).
Table 1
Cytotoxicity of synthetic compounds
Compound
R1
R2
R3
R4
R5
Configuration of (13a,14)
IC50 (nM)
A549
KB
HT-29
3
17
18
ent-3
19
ent-19
20
ent-20
21
ent-21
22
ent-22
23
H
OH
H
H
H
H
H
H
H
H
H
H
H
H
OH
H
H
OH
CH3O
CH3O
H
H
H
OH
H
H
H
H
H
H
H
H
H
H
H
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
H
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
CH3O
H
(S,S)
(S,S)
(S,S)
(R,R)
(S,S)
(R,R)
(S,S)
(R,R)
(S,S)
(R,R)
(S,S)
(R,R)
(S,S)
(S,S)
5.2
43.8
1.4
3.0
4.2
0.5
4.7
3.3
10.9
0.3
15.5
7.4
603.8
559.0
267.6
11.2
92.8
1.5
1.6
6.8
82.1
12.5
2.1
20.8
10.6
583.8
134.6
320.5
19.6
105.6
0.2
47.4
17.5
H
2031.9
180.1
1589.5
49.0
653.2
1.7
CH3CH2
CH3CH2
F
F
AcO
OH
24
1964.8
353.7
458.4
25
H
OH
H
(S,S)
25.5
54.4
134.5
26
27
H
H
OH
OH
H
H
(S,S)
(S,S)
741.8
10.9
927.8
0.7
487.5
1.2
CH3CH2O
CH3CH2O
Subsequently, we examined the enantiomers of 3, 19, 20, 21, and
22 to observe how the configuration at C-13a and C-14 influences
cytotoxicity. As a result, enantiomers were found to be less active
in almost all cases. We then probed the effect of substituents at R4
and R5. Changing the methoxy groups at R4 and R5 of 3 to hydrogen
atoms (24) led to a remarkable decrease in activity. This demon-
strated the importance of alkoxy groups at these positions. Thus,
we examined three compounds that have alkoxy group(s) at R4
and R5. Compounds 25 and 26 that have a cyclic alkoxy group be-
tween R4 and R5 exhibited lower activity than 3, while 27, having
ethoxy groups at R4 and R5, showed similar potent cytotoxicity to
that of 3.
Table 2
Antitumor efficacy in vivo
Compound
Inhibition rate (%)
Total dose (mg/kg)
Mortality
3
30.6
—
25
50
25
50
25
50
25
50
25
50
50
100
0/5
5/5
0/5
0/5
2/5
5/5
0/5
2/5
0/5
0/5
0/5
0/5
ent-3
19
7.2
12.5
4.3
—
23
54.9
39.3
ꢀ3.1
ꢀ10.6
46.2
78.3
27
Subsequently, we planned to explore the in vivo antitumor effi-
cacy of these compounds.16 Thus, we selected 3, ent-3, 19, 23, and
27 as testing probes because of their potent in vitro activities.
CPT-11

T. Ikeda et al. / Bioorg. Med. Chem. Lett. 21 (2011) 342–345
345
7.84 (1H, d, J = 7.8 Hz), 9.48 (1H, s), 10.45 (1H, br s); HRMS (ESI) calcd for
Contrary to the promising results in an in vitro screening, all
compounds showed low to moderate tumor growth inhibition
rates (Table 2). Only 23 showed moderate antitumor activity, but
due to inherent toxicity, higher dosing examination could not be
performed. The same toxicity was observed in the cases of 3 and
19, while ent-3 and 27 showed neither activity nor toxicity.
In summary, we completed asymmetric total synthesis of 3.
This synthetic route could be applicable for various substrates, thus
we could acquire various derivatives of 3 in view of the substitu-
ents on the phenanthrene ring and/or configurations at the C13a
and C14 stereocenters. Evaluation of in vitro cytotoxicities of these
synthetic compounds showed that the substituents on the phenan-
threne ring and configuration strongly affected the activity. In vivo
antitumor efficacy was explored using compounds that showed
potent cytotoxicity in vitro. In this examination, 23 showed mod-
erate antitumor efficacy, but almost all other compounds showed
inherent toxicity, thus limiting higher dosing examination. To
overcome this problem, new research is currently underway.
C
22H24NO4 [M+H]+, 366.1705. Found: 366.1713; ½a 2D8
ꢁ
+88.17 (c = 0.11, CHCl3 :
CH3OH = 1:1).
Compound 19: yellow powder, 1H NMR (400 MHz, CDCl3) d: 1.86–2.06 (3H, m),
2.30–2.56 (4H, m), 3.30–3.42 (2H, m), 3.94 (3H, s), 4.04 (3H, s), 4.11 (3H, s),
5.00–5.08 (1H, m), 6.67 (1H, s), 7.28 (1H, dd, J = 8.8, 2.4 Hz), 7.76 (1H, s), 7.83
(1H, d, J = 2.4 Hz), 8.38 (1H, d, J = 8.8 Hz); HRMS (ESI) calcd for C23H26NO4
[M+H]+, 380.1862. Found:380.1686; ½a 2D8
ꢁ
+106.09 (c = 0.11,CHCl3).
Compound 20: white powder, 1H NMR (400 MHz, DMSO-d6) d: 1.76–1.90 (3H,
m), 2.14–2.24 (1H, m), 2.32–2.48 (2H, m), 3.28–3.36 (1H, m), 3.51 (1H, d,
J = 15.4 Hz), 3.94 (3H, s), 4.01 (3H, s), 4.61 (1H, d, J = 15.4 Hz), 4.66–4.67 (1H,
m), 4.96–5.01 (1H, m), 7.26 (1H, s), 7.55–7.60 (2H, m), 8.16 (1H, s), 8.26–8.32
(1H, m), 8.72–8.76 (1H, m); HRMS (ESI) calcd for C22H24NO3 [M+H]+, 350.1756.
Found: 350.1754; ½a D28
ꢁ
+115.11 (c = 0.10, CHCl3).
Compound 21: white powder, 1H NMR (400 MHz, DMSO-d6) d: 1.32 (3H, t,
J = 7.8 Hz), 1.80–1.88 (3H, br s), 2.10–2.46 (3H, m), 2.86 (2H, q, J = 7.8 Hz),
3.29–3.38 (1H, m), 3.49 (1H, d, J = 15.6 Hz), 3.93 (3H, s), 4.02 (3H, s), 4.58 (1H,
d, J = 15.6 Hz), 4.62 (1H, d, J = 10.2 Hz), 4.91–4.99 (1H, m), 7.23 (1H, s), 7.40–
7.48 (1H, m), 8.15 (1H, s), 8.32 (1H, d, J = 8.8 Hz), 8.52 (1H, s); HRMS (ESI) calcd
for
CHCl3).
C
24H28NO3 [M+H]+, 378.2069. Found: 378.2079; a 2D9
½ ꢁ +91.29 (c = 0.02,
Compound 22: yellow powder, 1H NMR (400 MHz, DMSO-d6) d: 1.84 (3H, br s),
2.10–2.52 (3H, m), 3.30–3.33 (1H, m), 3.50 (1H, d, J = 15.6 Hz), 3.94 (3H, s), 4.01
(3H, s), 4.61 (1H, d, J = 15.6 Hz), 4.75 (1H, d, J = 9.76 Hz), 4.93–4.99 (1H, m),
7.27 (1H, s), 7.41–7.48 (1H, m), 8.11 (1H, s), 8.30–8.37 (1H, m), 8.55–8.61 (1H,
m); HRMS (ESI) calcd for C22H23FNO3 [M+H]+, 368.1662. Found: 368.1621; ½a 2D7
ꢁ
Acknowledgments
+159.42 (c = 0.34, CHCl3).
Compound 23: white powder, 1H NMR (400 MHz, DMSO-d6) d: 1.78–1.92 (3H,
m), 2.15–2.25 (1H, m), 2.30–3.05 (2H, m), 2.36 (3H, s), 3.25–3.40 (1H, m), 3.48–
3.63 (1H, m), 3.94 (3H, m), 4.01 (3H, m), 4.59–5.05 (3H, m), 7.26 (1H, s), 7.35
(1H, dd, J = 2.2, 9.0 Hz), 8.08 (1H, s), 8.32 (1H, d, J = 9.0 Hz), 8.48 (1H, d,
The authors thank Miyuki Uchida and Fukiko Nishisaka for their
excellent technical assistance.
J = 2.2 Hz); HRMS (ESI) calcd for
C
24H26NO5 [M+H]+, 408.1811. Found:
408.1850; ½a 2D7
ꢁ
+102.72 (c = 0.016, CHCl3).
References and notes
Compound 24: white powder, 1H NMR (400 MHz, DMSO-d6) d: 1.76–1.90 (3H,
m), 2.12–2.25 (1H, m), 2.30–2.48 (2H, m), 3.25–3.47 (1H, m), 3.49 (1H, d,
J = 15.9 Hz), 4.60 (1H, d, J = 15.9 Hz), 4.64–4.69 (0.5H, m), 4.90–4.96 (1H, m),
7.16 (1H, dd, J = 2.4, 8.8 Hz), 7.57–7.66 (2H, m), 7.90–7.96 (1H, m), 8.01 (1H, d,
J = 2.4 Hz), 8.18 (1H, d, J = 8.8 Hz), 8.55–8.62 (1H, m); HRMS (ESI) calcd for
1. You, X.; Pan, M.; Gao, W.; Shiah, H. S.; Tao, J.; Zhang, D.; Koumpouras, F.; Wang,
S.; Zhao, H.; Madri, J. A.; Baker, D.; Cheng, Y. C.; Yin, Z. Arthritis Rheum. 2006, 54,
877.
2. Baumgartner, B.; Erdelmeier, C. A. J.; Wright, A. D.; Rali, T.; Sticher, O.
Phytochemistry 1990, 29, 3327.
3. Yang, C.-W.; Chen, W.-L.; Wu, P.-L.; Tseng, H.-Y.; Lee, S.-J. Mol. Pharmacol. 2006,
69, 749.
4. Gao, W.; Lam, W.; Zhong, S.; Kaczmarek, C.; Baker, D. C.; Cheng, Y. C. Cancer Res.
2004, 64, 678.
C
20H20NO2 [M+H]+, 306.1494. Found: 306.1462; ½a 2D8
ꢁ
+97.1 (c = 0.04, CH3OH :
CHCl3 = 1 : 1).
Compound 25: white powder, 1H NMR (400 MHz, DMSO-d6) d: 1.75–1.87 (3H,
m), 2.08–2.24 (1H, m), 2.30–2.41 (2H, m), 3.27–3.32 (1H, m), 3.39 (1H, d,
J = 15.4 Hz), 4.47 (1H, d, J = 15.4 Hz), 4.58 (1H, d, J = 10.0 Hz), 4.89 (1H, dd,
J = 1.9, 10.0 Hz), 6.17 (2H, d, J = 4.0 Hz), 7.10 (1H, dd, J = 2.4, 8.8 Hz), 7.33 (1H,
s), 7.82 (1H, d, J = 2.4 Hz), 8.01 (1H, s), 8.12 (1H, d, J = 8.8 Hz), 9.73 (1H, br s);
HRMS (ESI) calcd for C21H20NO4 [M+H]+, 350.1392. Found: 350.1391.
Compound 26: yellow powder, 1H NMR (400 MHz, DMSO-d6) d: 1.71 (3H, s),
1.74 (3H, s), 1.76–1.86 (3H, m), 2.09–2.21 (1H, m), 2.30–2.40 (2H, m), 3.25–
3.31 (1H, m), 3.39 (1H, d, J = 15.1 Hz), 4.46 (1H, d, J = 15.1 Hz), 4.54 (1H, d,
J = 9.8 Hz), 4.89 (1H, dd, J = 2.2, 9.8 Hz), 7.08 (1H, dd, J = 2.4, 9.0 Hz), 7.25 (1H,
s), 7.80 (1H, d, J = 2.4 Hz), 7.91 (1H, s), 8.11 (1H, d, J = 9.0 Hz), 9.67 (1H, br s);
HRMS (ESI) calcd for C23H24NO4 [M+H]+, 378.1705. Found: 378.1700.
Compound 27: white powder, 1H NMR (400 MHz, DMSO-d6) d: 1.41 (3H, t,
J = 7.0 Hz), 1.43 (3H, t, J = 7.0 Hz), 1.72–1.87 (3H, m), 2.08–2.23 (1H, m), 2.28–
2.42 (2H, m), 3.21–3.34 (1H, m), 3.38 (1H, d, J = 15.5 Hz), 4.16 (2H, dq, J = 7.0,
11.7 Hz), 4.27 (2H, q, J = 7.0 Hz), 4.43 (1H, d, J = 15.5 Hz), 4.60 (1H, d, J = 8.9 Hz),
4.88 (1H, d, J = 8.9 Hz), 7.08 (1H, dd, J = 2.2, 8.8 Hz), 7.13 (1H, s), 7.88 (1H, d,
J = 2.2 Hz), 7.89 (1H, s), 8.11 (1H, d, J = 8.8 Hz), 9.60 (1H, s);.
5. Gellert, E. Alkaloids: Chemical and Biological Perspectives In Pelletier, S. W.,
Ed.; Academic Press: New York, 1987; pp 55–132.
6. Komatsu, H.; Watanabe, M.; Ohyama, M.; Enya, T.; Koyama, K.; Kanazawa, T.;
Kawahara, N.; Sugimura, T.; Wakabayashi, K. J. Med. Chem. 2001, 44, 1833.
7. Gao, W.; Chen, A. P.-C.; Leung, C.-H.; Gullen, E. A.; Furstner, A.; Shi, Q.; Wei, L.;
Lee, K.-H.; Cheng, Y.-C. Bioorg. Med. Chem. Lett. 2008, 18, 704.
8. Wang, Q. M.; Wang, K.; Huang, Z.; Liu, Y.; Li, H.; Hu, T. S.; Jin, Z.; Fan, Z.; Huang,
R. Q. CN Patent 2006101295556, 2006.
9. Li, Z.; Zhong, J.; Huang, R. Synthesis 2001, 2365.
10. Gao, W.; Bussom, S.; Grill, P. S.; Gullen, A. E.; Hu, Y.-C.; Huang, X.; Zhong, S.;
Kaczmarek, C.; Gutierrez, J.; Francis, S.; Baker, C. D.; Yu, S.; Cheng, Y.-C. Bioorg.
Med. Chem. Lett. 2007, 17, 4338.
11. Buckley, T. F., III; Rapoport, H. J. Org. Chem. 1983, 48, 4222.
12. Lir, L.; Yang, B.; Katz, T.; Poindexter, M. K. J. Org. Chem. 1991, 56, 3769.
13. Debenzyl derivative could also be utilized for synthesis of 3.
14. Compound 3: yellow powder, 1H NMR (400 MHz, DMSO-d6) d: 1.75–1.88 (3H,
m), 2.12–2.20 (1H, m), 2.30–2.42 (2H, m), 3.40–3.49 (2H, m), 3.92 (3H, m), 3.99
(3H, m), 4.53 (1H, d, J = 16.1 Hz), 4.58–4.61 (1H, m), 4.90–4.92 (1H, m), 7.09
(1H, d, J = 2.2, 9.0 Hz), 7.19 (1H, s), 7.91 (1H, s), 7.91 (1H, s), 8.12 (1H, d,
J = 9.0 Hz), 9.63 (1H, br s); HRMS (ESI) calcd for C22H24NO4 [M+H]+, 366.1705.
15. Cell viability was assayed in a 96-well plate using a TetraColor ONE (Seikagaku
Corp., Tokyo, Japan), according to the manufacturer’s protocol. Briefly,
exponentially growing KB, A549 or HT-29 cells were seeded into 96-well
plates at a density of 103 cells/well, respectively. The next day, serial diluted
compounds were added. After 96 h incubation, 2-(2-methoxy-4-nitrophenyl)-
3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium monosodium salt:
WST-8 reagent was added to each well and incubated at 37 °C for 1 h.
Absorbance at 450 nm was measured with SPECTRA Max PLUS384 (Molecular
Devices, Sunnyvale, CA). Cell viability IC50 was defined as the concentration of
compound that inhibited cell viability by 50% compared to solvent-treated
control cells.
Found: 366.1711; ½a D25
ꢁ
+102.30 (c = 0.12, CHCl3: CH3OH = 1:1).
Compound 17: yellow powder, 1H NMR (400 MHz, DMSO-d6) d: 1.78–1.91 (3H,
m), 2.10–2.26 (1H, m), 2.28–2.45 (2H, m), 3.20–3.35 (1H, m), 3.47 (1H, d,
J = 15.6 Hz), 3.90 (3H, s), 3.98 (3H, s), 4.52 (1H, d, J = 10.0 Hz), 4.57 (1H, d,
J = 15.6 Hz), 4.80 (1H, dd, J = 2.0, 10.0 Hz), 7.09 (1H, dd, J = 2.6, 8.9 Hz), 7.19
(1H, s), 7.62 (1H, d, J = 2.6 Hz), 8.0 (1H, s), 8.55 (1H, d, J = 8.9 Hz), 9.62 (1H, br
s); HRMS (ESI) calcd for C22H24NO4 [M+H]+, 366.1705. Found: 366.1700; ½a 2D8
ꢁ
16. Meth A cells (sarcoma, 2.5 ꢂ 105 cells/mouse) were inoculated sc into 7-wk old
male BALB/c mice (5/group), and samples were injected iv on days 1, 5, and 9.
Tumors were weighed on day 21 after tumor inoculation. The inhibition rate
(%) was calculated by the formula[1 ꢀ (mean tumor weight of tested mice)/
(mean tumor weight of control mice] ꢂ 100.
+137.83 (c = 0.11, CHCl3 : CH3OH = 1:1).
Compound 18: yellow powder, 1H NMR (400 MHz, DMSO-d6) d: 1.75–1.91 (3H,
m), 2.10–2.26 (1H, m), 2.28–2.45 (2H, m), 3.50 (1H, d, J = 15.9 Hz), 3.91 (3H, s),
3.93 (3H, s), 4.10–4.15 (1H, m), 4.52–4.58 (1H, m), 4.59 (1H, d, J = 15.9 Hz), 4.89
(1H, dd, J = 2.0, 10.0 Hz), 7.06 (1H, s), 7.36 (1H, t, J = 7.8 Hz), 7.64–7.75 (1H, m),