Novel quinazoline-quinoline alkaloids with cytotoxic and DNA topoisomerase II inhibitory activities

Article abstract of DOI:10.1016/j.bmcl.2003.12.048

Two new synthetic analogues of luotonins A and F, 7-acetylaminoluotonin A (6) and 3-[3H(quinazolino-4-one)]quinoline (7) were synthesized. The new analogues, along with four natural quinazoline-quinoline alkaloids, luotonins A (1), B (2), E (3), F (4) and a synthetic deoxoluotonin F (5), showed cytotoxic activity (IC₅₀ 1.8-40.0 μg/mL) and DNA topoisomerase II inhibition at a concentration of 25 μM.

Full text of DOI:10.1016/j.bmcl.2003.12.048

Bioorganic & Medicinal Chemistry Letters 14 (2004) 1193–1196
Novel quinazoline–quinoline alkaloids with cytotoxic and DNA
topoisomerase II inhibitory activities
Zhongze Ma,a,* Yoshio Hano,a Taro Nomuraa and Yingjie Chenb
aFaculty of Pharmaceutical Sciences, Toho University, 2-2-1 Miyama, Funabashi, Chiba, 274-8510, Japan
bShenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, Liaoning 110015, China
Received 5 November 2003;revised 8 December 2003;accepted 12 December 2003
Abstract—Two new synthetic analogues of luotonins A and F, 7-acetylaminoluotonin A (6) and 3-[3H(quinazolino-4-one)]quino-
line (7) were synthesized. The new analogues, along with four natural quinazoline–quinoline alkaloids, luotonins A (1), B (2), E (3),
F (4) and a synthetic deoxoluotonin F (5), showed cytotoxic activity (IC50 1.8–40.0 mg/mL) and DNA topoisomerase II inhibition at
a concentration of 25 mM.
# 2003 Elsevier Ltd. All rights reserved.
Luotonins A (1), B (2), E (3) and F (4) are novel qui-
nazoline–quinoline alkaloids that were isolated from the
aerial parts of Peganum nigellastrum Bunge.1,2 Luotonin
A (1) has unique pyrroloquinazolinoquinoline ring sys-
tem and showed cytotoxic activity against mouse leuke-
mia P-388 cells in vitro at a concentration of 1.8 mg/
mL.1 The structure of luotonin A (1) is strikingly remi-
niscent of the cytotoxic alkaloid camptothecin, whose
derivatives are clinically useful anti-cancer agents.3 This
similarity has stimulated much activity directed toward
the synthesis of this compound by several groups.4 The
syntheses of luotonins B4c,j (2), E2 (3) and F2,5 (4) have
also been achieved, but no biological data was reported.
In a recent paper, Hecht et al.6 demonstrated that luo-
tonin A (1) stabilized the human DNA topoisomerase
I–DNA covalent binary complex, affording the same
pattern of cleavage as the structurally related topoi-
somerase I inhibitor camptothecin. Further it was
shown that luotonin A (1) also mediated topoisomerase
I-dependent cytotoxicity toward Saccharyomyces cere-
visiae lacking yeast topoisomerase I, but harboring a
plasmid having the human topoisomerase I gene under
the control of a galactose promoter.6 The above findings
make the synthesis of luotonin A analogues an attrac-
tive target to evaluate their biological activities. In this
communication, we report the syntheses of 7-acetyl-
aminoluotonin A (6) and a luotonin F analogue, 3-
[3H(quinazolino-4-one)]quinoline (7), and compare
their cytotoxic and topoisomerase II inhibitory activities
with natural alkaloids, luotonins A (1), B (2), E (3), F
(4) and synthetic deoxoluotonin F2 (5).
* Corresponding author at present address: Bio-organic and Natural
Products Laboratory, McLean Hospital, Harvard Medical School,
115 Mill Street, Belmont, MA 02478, USA. Tel.: +1-617-855-2039;
fax: +1-617-855-2040;e-mail: zhongze_ma@hms.harvard.edu
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.12.048

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Z. Ma et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1193–1196
Table 1. NMR data of 2, 6 and 8 at C-7 position
(MW: 343) which suggests the presence of an additional
nitrogen in compound 6. From the above evidence, the
structure of 6 was determined as 7-acetylaminoluotonin
A. The formation of 6 could be explained by Ritter
reaction9 as shown in Scheme 2.
Compd
1H (d)
13C (d)
2
6
8
7.14 s
7.16 s
8.06 s
80.9
63.8
78.3
As shown in Scheme 3, hydrolysis of 3-quinolinenitrile
(9) with concentrated sulfuric acid yielded 3-quinoline-
carboxamide (10) in 95% yield. Subsequent reaction of
the amide (10) with isatoic anhydride10 at 200–210 ꢀC
for 2 h afforded 3-[3H(quinazolino-4-one)]quinoline11
(7) in 39% yield. Compounds 7 and 4 have similar
structural features and contain quinoline and quinazo-
linone moieties. In compound 7, the C-3 of quinoline
moiety is directly linked to the C-2 of quinazolinone
moiety whereas in 4 the two moieties are linked through
a junction of the carbonyl group.
Luotonin B (2) was synthesized4c by CAN oxidation of
luotonin A (1) in acetonitrile under reflux conditions
(Scheme 1). After 6 h reaction, the desired luotonin B
(2) was isolated in 15% yield along with compound 6 in
25% yield. The structure of 6 was elucidated by com-
1
parison of H and 13C NMR data (see Table 1) of luo-
tonin B (2) and 7-acetoxyluotonin A7 (8). The chemical
shift value of the methine carbon (d 63.8) in 6 is at
higher-field than those of 2 (d 80.9) and 8 (d 78.3), while
the chemical shift value of the methine proton (d 8.06)
in 8 is at lower-field than those of 2 (d 7.14) and 6 (d
7.16). The NMR data suggested that nitrogen atom
should be adjacent to C-7 position. The structure of
compound 6 was further confirmed by its molecular
weight obtained by FAB-MS spectrum.8 The molecular
weight of 6 (MW: 342) is one unit less than that of 8
Luotonins A (1), B (2), E (3) and F (4) are less polar
alkaloids isolated from the hexane, benzene and
chloroform extracts of Peganum nigellastrum.1,2 It has
been reported12 that the lipophilic alkaloid fraction of
P. nigellastrum showed anti-tumor effect on mice
implanted with ascetic hepatoma cells, and inhibited
Scheme 1.
Scheme 2.

Z. Ma et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1193–1196
1195
Scheme 3.
Table 2. Cytotoxic activity against mouse leukemia P-388 cells
References and notes
Compd
1
2
3
4
5
6
7
1. Ma, Z. Z.;Hano, Y.;Nomura, T.;Chen, Y. J.
cycles 1997, 46, 541.
2. Ma, Z. Z.;Hano, Y.;Nomura, T.;Chen, Y. J.
cycles 1999, 51, 1883.
Hetero-
Hetero-
IC50 (mg/mL)
1.8
5.0
9.0
20.0
2.3
33.0
40.0
3. Slichenmyer, W. J.;Rowinsky, E. K.;Donehower, R. C.;
Kaufmann, S. H. J. Nat. Cancer Inst. 1993, 85, 271.
4. (a) Wang, H.;Ganesan, A. Tetrahedron Lett. 1998, 39,
9097. (b) Kelly, T. R.;Chamberland, S.;Silva, R. A. Tet-
rahedron Lett. 1999, 40, 2728. (c) Ma, Z. Z.;Hano, Y.;
Nomura, T.;Chen, Y. J. Heterocycles 1999, 51, 1593. (d)
Molina, P.;Terraga, A.;Gonzalez-Tejero, A. Synthesis
2000, 1523. (e) Toyota, M.;Komori, C.;Ihara, M. Het-
erocycles 2002, 56, 101. (f) Dallavalle, S.;Merlini, L. Tet-
rahedron Lett. 2002, 43, 1835. (g) Yadav, J. S.;Reddy,
B. V. S. Tetrahedron Lett. 2002, 43, 1905. (h) Osborne, D.;
Stevenson, P. J. Tetrahedron Lett. 2002, 43, 5469. (i) Lee,
E. S.;Park, J. G.;Jahng, Y. Tetrahedron Lett. 2003, 44,
1883. (j) Harayama, T.;Morikami, Y.;Shigeta, Y.;Abe,
H.;Takeuchi, Y. Synlett 2003, 847.
DNA and protein syntheses in the hepatoma cells. In
order to evaluate the anti-tumor activity of these novel
alkaloids (1–4) and their synthetic derivatives (5–7), we
carried out biological assays for the cytotoxic activity
against P 388 mouse leukemia cells13 and the inhibitory
activity against human topoisomerase II.14 The cyto-
toxic effects of compounds 1–7 were tested and the
results were summarized in Table 2. Among them, luo-
tonin A (1) and deoxoluotonin F (5) showed cytotoxic
activity at lower concentrations, IC50 1.8 mg/mL and
IC50 2.3 mg/mL respectively. Comparison of the activ-
ities of 1, 2, 3 and 6, having the same pyrroloquinazoli-
noquinoline skeleton, revealed that the substituents
containing oxygen or nitrogen atom at C-7 position
have lower cytotoxicity. These data suggested that the
methylene group is very important for the activity. The
same tendency was also observed for the analogues 4, 5
and 7. On the other hand, DNA unknotting assays with
human topoisomerase II on knotted P4 phage DNA
was examined to detect the topoisomerase II inhibitory
activity. All seven compounds (1–7) had potent inhibi-
tory activity against human topoisomerase II at the
concentration of 25 mM, and all of them showed the
similar inhibitory intensity. In another experiment, luo-
tonin A exhibited IC50 value of 28.5 mM, which was
comparable to ICRF-193,15 a DNA topoisomerase II
inhibitor (IC50 13.9 mM).
5. Mhaske, S. B.;Argade, N. P. Synthesis 2002, 323.
6. Cagir, A.;Jones, S. H.;Gao, R.;Eisenhauer, B. M.;
Hecht, S. M. J. Am. Chem. Soc. 2003, 125, 13628.
7. Compound 8: mp 282–285 ꢀC. UV lmax (MeOH) nm (log
e): 213 (4.70), 250 (4.75), 300 (sh, 4.18), 322 (sh, 4.31), 336
1
(4.38), 352 (4.28). H NMR (CDCl3, 400 MHz) d: 2.22 (s,
3H), 7.60 (dt, J=1.5 and 8.0 Hz, 1H), 7.73 (dt, J=1.5 and
8.0 Hz, 1H), 7.86 (dt, J=1.5 and 8.0 Hz, 1H), 7.90 (dt,
J=1.5 and 8.5 Hz, 1H), 7.99 (d, J=8.0 Hz, 1H), 8.06 (s,
1H), 8.10 (dd, J=1.5 and 8.0 Hz, 1H), 8.41 (dd, J=1.5
and 8.0 Hz, 1H), 8.48 (d, J=8.5 Hz, 1H), 8.60 (s, 1H). 13
C
NMR (CDCl3, 100 MHz) d: 20.4, 78.3, 121.7, 126.6,
127.8, 128.5, 128.6, 128.8, 129.4, 130.5, 130.6, 131.3,
133.8, 134.8, 148.6, 150.0, 150.1, 150.9, 160.4, 169.9. FAB-
MS: m/z 344 (M+H)+.
8. Compound 6: mp >300 ꢀC. UV lmax (MeOH) nm (log
e): 207 (4.79), 248 (4.67), 297 (4.20), 325 (4.23), 338 (4.25),
In conclusion, the present study demonstrated that luo-
tonin A (1) and its analogues (2–7) exhibited cytotoxic
activity through inhibition of DNA topoisomerase II.
Considering the new finding by Hecht’s group,6 luoto-
nin A was suggested as a potent inhibitor for both
topoisomerases I and II. Luotonin A (1) may have uti-
lity as an antineoplastic agent because of its novel
mechanism of cytotoxicity.
1
354 (4.14). H NMR (CDCl3–CD3OD: 3:1, 400 MHz) d:
1.95 (s, 3H), 7.16 (s, 1H), 7.41 (t, J=8.0 Hz, 1H), 7.55 (t,
J=8.0 Hz, 1H), 7.70 (t, J=8.0 Hz, 1H), 7.72 (t, J=8.0
Hz, 1H), 7.82 (d, J=8.0 Hz, 1H), 7.86 (d, J=8.0 Hz, 1H),
8.16 (d, J=8.0 Hz, 1H), 8.21 (d, J=8.0 Hz, 1H), 8.35 (s,
1H). 13C NMR (CDCl3–CD3OD: 3:1, 100 MHz) d: 22.7,
63.8, 121.7, 126.2, 127.6, 128.2, 128.3, 128.5, 128.9, 129.9,
130.9, 131.7, 132.0, 134.7, 148.5, 149.4, 149.7, 151.4,
160.1, 171.4. FAB-MS: m/z 343 (M+H)+.
9. Ritter, J. J.;Minieri, P. P. J. Am. Chem. Soc. 1948, 70,
4045.
10. Kametani, T.;Ohsawa, T.;Ihara, M.;Fukumoto, K.
Chem. Pharm. Bull. 1978, 26, 1922.
Acknowledgements
The authors thank Drs. H. Itokawa and K. Takeya,
Tokyo College of Pharmacy and Life Science, for con-
ducting cytotoxicity against mouse leukemia P-388 cells
and Drs. M. Sato and S. Terada, Zenyaku Kogyo Co.,
Ltd, for performing DNA topoisomerase II inhibitory
study.
11. Compound 7: mp >300 ꢀC. IR nmax (KBr) cmÀ1: 3403,
1669, 1620, 1572, 1498, 1470, 1307, 1149, 930, 772. 1H
NMR (DMSO-d6, 400 MHz), d: 7.57 (dt, J=1.2 and 8.1
Hz, 1H), 7.73 (t, J=8.0 Hz, 1H), 7.82 (d, J=8.1 Hz, 1H),
7.88 (dt, J=1.4 and 8.4 Hz, 1H), 7.90 (dt, J=1.2 and 8.2
Hz, 1H), 8.12 (d, J=8.3 Hz, 2H), 8.20 (dd, J=1.2 and 8.1

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Z. Ma et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1193–1196
13. Nagara, H.;Tobita, Y.;Nagae, T.;Itokawa, H.;Takeya,
Hz, 1H), 9.15 (d, J=2.2 Hz, 1H), 9.60 (d, J=2.2 Hz, 1H),
12.82 (brs, 1H). EI–MS: m/z 273 (M+, 78), 119 (100).
HR-EIMS: m/z 273.0904 (M+, C17H11N3O, requires
273.0902).
K.;Halim, A. F.;Abdel-Halim, O. B.
1997, 44, 1115.
Phytochemistry
14. Ma, Z. Z.;Hano, Y.;Nomura, T.;Chen, Y. J.
Prod. 2000, 63, 390.
J. Nat.
Cancer
12. Xiao, X. H.;Qou, G. L.;Wang, H. L.;Lui, L. S.;Zheng,
Y. L.;Jia, Z. J.;Deng, Z. B. Chinese J. Pharmacol. Tox-
icol 1988, 232.
15. Tanabe, K.;Ikegami, Y.;Ishida, R.;Andoh, T.
Res. 1991, 51, 4903.