Structure of a novel multidrug resistance modulator, irciniasulfonic acid, isolated from a marine sponge, Ircinia sp.

Article abstract of DOI:10.1016/S0040-4039(01)00426-9

Irciniasulfonic acid was obtained from a marine sponge of Ircinia sp. Spectroscopic and chemical analyses revealed its structure consists of three different kinds of acids, i.e. common fatty acids, a novel unsaturated branched C-10 fatty acid and an isethionic acid. Irciniasulfonic acid and deacyl irciniasulfonic acid reverse multidrug resistance in human carcinoma cells caused by overexpression of membrane glycoprotein.

Full text of DOI:10.1016/S0040-4039(01)00426-9

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 42 (2001) 3335–3337
Structure of a novel multidrug resistance modulator,
irciniasulfonic acid, isolated from a marine sponge, Ircinia sp.
a
a
a,
b
Ayako Kawakami, Tomofumi Miyamoto, Ryuichi Higuchi, * Takeshi Uchiumi,
b
c
Michihiko Kuwano and Rob W. M. Van Soest
a
Graduate School of Pharmaceutical Sciences, Kyushu University, Maidashi 3-1-1, Higashi-ku, Fukuoka 812-8582, Japan
b
Department of Medical Biochemistry, Graduate School of Medical Sciences, Kyushu University, Maidashi 3-1-1, Higashi-ku,
Fukuoka 812-8582, Japan
c
Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, PO Box 94766, 1090 GT Amsterdam, Netherlands
Received 8 February 2001; revised 8 March 2001; accepted 9 March 2001
Abstract—Irciniasulfonic acid was obtained from a marine sponge of Ircinia sp. Spectroscopic and chemical analyses revealed its
structure consists of three different kinds of acids, i.e. common fatty acids, a novel unsaturated branched C-10 fatty acid and an
isethionic acid. Irciniasulfonic acid and deacyl irciniasulfonic acid reverse multidrug resistance in human carcinoma cells caused
by overexpression of membrane glycoprotein. © 2001 Elsevier Science Ltd. All rights reserved.
Multidrug resistance (MDR) is one of the main causes
for failure in chemotherapy treatment for cancer
patients. Overexpression of a membrane glycoprotein
While searching for other MDR modulators in marine
invertebrates, we isolated a novel fatty acid analogue
named irciniasulfonic acid (ISA) as an active con-
stituent from the Japanese marine sponge, Ircinia sp. In
this paper, we will report on the isolation and structural
determination of ISA.
(
termed P-glycoprotein or P-gp ) confers resistance by
acting as an energy-dependent drug transporter or an
efflux pump, which results in the decreased cellular
accumulation of drugs. Since the discovery of a calcium
channel blocker verapamil was recognized as an agent
for reversing MDR, a variety of compounds, including
cyclosporin A, have been reported to reverse MDR.
Ircinia sp. (wet weight 2.8 kg) was collected by hand at
depths of 10–15 m off Tsutsumi Island, Fukuoka pre-
4
fecture, Japan, in 1999. The Et O soluble fraction (3.0
2
1
2
For example, agosterol A, patellamide D, and lamel-
larin I have recently been isolated from marine inverte-
g) of the EtOH extract obtained from Ircinia sp.
reversed MDR at 33 mg/mL against P-gp overexpress-
3
5
brates as resistance modulators.
ing MDR tumor cells (KB/VJ300) in the presence of
O
2'
-
3
10
9
Irciniasulfonic acid (ISA, 1)
O S
3
O
1
1'
O
2
11
OR
15"
22"
O
O
5"
R=
9"
24"
23"
Deacyl irciniasulfonic acid (2): R=H
Figure 1.
Keywords: multidrug resistance; irciniasulfonic acid; Ircinia sp.; sulfonic acids and derivatives.
*
Corresponding author. Fax: +(81) 92-642-6639; e-mail: rhiguchi@phar.kyushu-u.ac.jp
0
040-4039/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(01)00426-9

3
336
A. Kawakami et al. / Tetrahedron Letters 42 (2001) 3335–3337
1
0 ng/mL of vincristine, but no cytotoxicity was seen
as shown in Fig. 2. The geometry of the tri-substituted
13
against parental KB cells at 100 mg/mL. Bioassay
guided the separation of the active fraction by Sep-
hadex LH-20 (CHCl :MeOH 1:1) and silica gel chro-
olefin was assigned as Z from the C-chemical shifts
10
value (l =23.8).
C
3
matography (EtOAc:MeOH 10:1) to give an active
compound, ISA (1, Fig. 1) (49.5 mg, 1.7×10 % yield).
The absolute configuration at C-9 was investigated by
11
−
3
application of modified Mosher’s method. Deacyl ISA
was treated with S-(−)- or R-(+)-2-methoxy-2-phenyl-2-
trifluoromethyl-acetyl chloride to furnish the 9-O-R-(+)
6
ISA (1) was obtained as a colorless oil and showed a
positive reaction to the qualitative analysis of sulfonic
(
2a) and 9-O-S-(−)-MTPA ester (2b). The absolute
7
acid. The IR spectrum revealed a strong absorption
configuration at C-9 was determined to be R on the
basis of Dl values, which were obtained from the
−
1
1
13
band due to ester carbonyl (1719 cm ). The H and
C
NMR data suggested the presence of a primary methyl,
a secondary methyl, a tri-substituted olefin, an oxy-
genated methine, esters and a long alkyl chain. The
negative-FABMS showed a series of quasi-molecular
12
proton signals of 2a and 2b as shown in Fig. 3.
Fatty acid methyl esters (FAMs) were purified by
reversed phase HPLC, and each FAM was analyzed by
GC–MS. The positions of double bonds were deter-
mined by the mass fragmentation of their DMDS
−
ion peaks at m/z 653, 643, 627 [M−H] , caused by the
diversity of a fatty acyl moiety, together with a base
−
peak at m/z 289 [M−fatty acids] , and a fragmentation
13
derivatives, and their geometry was elucidated by the
−
peak at m/z 80 [SO ] . Methanolysis with 5% HCl/
3
13
14
allylic C carbon signals. Thus, the three kinds of
8
MeOH gave deacyl ISA (2), a mixture of fatty acid
constituent fatty acid components were identified as
methyl ester, and a small amount of isethionic acid
1
5a
15b
9
tricosanoic acid,
(
(Z)-15-docosenoic acid,
and
(
HO SCH CH OH).
3 2 2
15c
5Z,9Z)-5,9-tetracosenoic acid. Accordingly, the final
structure of 1 was determined as shown in Fig. 1.
Deacyl ISA (2) showed the presence of sulfonic acid,
and the UV spectrum indicated the presence of a,b-
ISA reversed the multidrug resistance to vincristine in
KB/VJ300 cells at 25 mg/mL (ca. 38 mM), which corre-
sponds to 1 mM of verapamil. However, 2 was 13-fold
stronger than 1 (ca. 3 mM), but isethionic acid had no
activity. The details of the biological activities will be
reported elsewhere.
unsaturated carbonyl group [u
220 nm (m 6300)]. The
max
IR spectrum showed the absorption bands due to
−
1
−1
hydroxyl (3412 cm ) and ester carbonyl (1705 cm ).
The HR-API-TOFMS showed the molecular ion peak
at m/z 307.1218 (D+0.3 mmu) corresponding to the
1
13
molecular formula of C H O S. The H, C NMR
1
3
23
6
and HSQC spectral data suggested the presence of one
secondary methyl, one olefinic methyl, seven methyl-
enes, one oxygenated methine, one tri-substituted olefin
and one conjugated ester. The H– H COSY and
TOCSY spectra of 2 afforded partial structures A and
B and these two partial structures and the conjugated
ester were merged by the aid of the HMBC experiment
To our knowledge, irciniasulfonic acid is the first natu-
rally occurring product composed of normal fatty
acids, isethionic acid and a unique fatty acid, 9-
hydroxy-3-methyl-2-decenoic acid. Irciniasulfonic acid
and its analogues will be useful for human cancer
chemotherapy as MDR modulators in a clinical cure.
1
1
B
A
OH
67.1 22.0
O
O S
O-
4
9.9
59.0
166.1 115.1 161.6
32.8
27.8
29.3
25.2
38.7
C
H2
C
H2
4.32
O
C
O
C
C
C
H2
2.54
C
H2
1.39 1.19
(m)
C
H2
C
C
C CH3
H2
H2
1.05
H
.60
H
3.05
1.28
1.30
3.61 (3H, d, 6.4)
(m)
5
(t, 7.2) (t, 7.2)
CH3 (m)
.80
3H, d, 1.3)
(m)
(m)
(m)
(br.s)
1
23.8
(
HMBC COSY & TOCSY
1
1
Figure 2. H– H COSY, TOCSY and HMBC correlations of 2.
O
+
0.09
0.00
-0.06
-0.12
-0.06
-
O S
3
9R CH3
O
0.00
-0.09
-0.02
H
CH3
H
.00
OMTPA
δ=δS-δR (ppm)
-0.01
-0.01
0
∆
Figure 3. Application of modified Mosher’s method to 2.

A. Kawakami et al. / Tetrahedron Letters 42 (2001) 3335–3337
3337
Acknowledgements
70.7 (C-9), 115.1 (C-2), 128.5, 128.7, 129.4, 129.9
(
(
-C
C-1%%).
6 Hꢀ C6 H-), 161.6 (C-3), 165.9 (C-1), 173.0, 173.6, 173.8
We would like to thank Professor T. Murae of the
Graduate School of Science, Kyushu University for
providing HR-API-TOFMS data. This work was sup-
ported in part by a Scientific Research Grant (No.
7. Momose, T.; Ohkura, Y.; Tanaka, H. Pharm. Bull. 1954,
2, 154–156.
−
1
8. Deacyl irciniasulfonic acid (2): IR (CHCl , cm ) 3412
3
(OH), 2931, 2857, 1705, 1645, 1170, UV (MeOH, umax)
1
13
1
2672055) from The Ministry of Education, Science,
Sports and Culture, Japan.
220 nm (m 6300), H, and C NMR (CD OD): summa-
3
rized in Fig. 1.
9. Isethionic acid was identical with the authentic sample
(
Wako Pure Chemical Industries Ltd.) in spectral data;
References
. Aoki, S.; Yoshioka, Y.; Miyamoto, Y.; Higuchi, K.;
however, the O-methyl-9-hydroxy-3-methyl-2-decenoate
could not be isolated.
10. De Haan, W. J.; Van de Ven, J. L. M. Org. Magn. Reson.
1
Setiawan, A.; Murakami, N.; Chen, Z.-S.; Sumizawa, T.;
1973, 5, 147–153.
Akiyama, S.; Kobayashi, M. Tetrahedron Lett. 1998, 39,
11. Ohtani, I.; Kusumi, T.; Kashman, Y.; Kakishima, H. J.
Am. Chem. Soc. 1991, 113, 4092–4096.
6
303–6306.
1
2
3
. Allen, B. W.; Robert, S. J. Cancer Lett. 1993, 71, 97–102.
. Quesada, A. R.; Garcia Gravalos, M. D.; Fernandez
Puentes, J. L. Br. J. Cancer 1996, 74, 677–682.
. Ircinia sp.; Class Demospongiae, order Dictyoceratida,
family Irciniidae. A voucher specimen was deposited in
the Zoological Museum in University of Amsterdam
12. 2a: H NMR (CD OD, 600 MHz, l): 1.15 (H -10), 1.27
3
3
(H -6), 1.37 (H -5), 1.51 (H -7), 1.57 (H -8), 1.79 (H -11),
2
2
2
2
3
2.52 (H -4), 3.03 (H -1%), 4.32 (H -2%), 5.03 (H-9), 5.60
2
2
2
1
4
(H-2), 2b: H NMR (CD OD, 600 MHz, l): 1.24 (H -10),
3
3
1.14 (H -6), 1.28 (H -5), 1.49 (H -7), 1.51 (H -8), 1.78
2
2
2
2
(H -11), 2.46 (H -4), 3.03 (H -1%), 4.32 (H -2%), 5.03 (H-9),
3
2
2
2
(ZMA Por 16383).
5.60 (H-2).
5
6
. Kohno, K.; Kikuchi, J.; Sato, S.; Takano, H.; Saburi, Y.;
Asoh, K.; Kuwano, M. Jpn. J. Cancer Res. (Gann) 1988,
13. Vincenti, M.; Guglielmetti, G.; Cassani, G.; Tonini, C.
Anal. Chem. 1987, 59, 694–699.
14. Fusetani, N.; Yasumuro, K.; Matsunaga, S.; Hirota, H.
Tetrahedron Lett. 1989, 30, 6891–6894.
7
9, 1238–1246.
−
1
. Irciniasulfonic acids (1): IR (CHCl , cm ) 2927, 2855,
3
1
645, 1719, UV (MeOH, umax) 220 nm (m 6100), negative-
15. (a) O-methyl tricosanoate (41.0% by GC); EIMS (m/z)
−
+
FABMS (m/z) 653, 643, 627 [M−H] , 363, 353, 337
368 (M ), 337, 325, 87, 74; (b) O-methyl (Z)-15-
−
−
−
1
+
13
[
R−COO] , 289 [M−fatty acid] , 80 [SO ] , H NMR
docosenoate (17.1%): EIMS (m/z) 352 (M ); C NMR
(CDCl , l): 26.9 (-CH -CHꢀCH-CH -); DMDS deriva-
3
(
CD OD, 600 MHz, l): 0.77, 0.80 (t, methyls), 1.11 (3H,
6
6
3
3
2
2
+
+
d, 6.6 Hz, H -10), 1.15–2.55 (methylenes), 1.39 (m, H -5),
tive: EIMS (m/z) 446 (M ), 301 (C H O S ), 145
3
2
17 33 2
+
1
.80 (s, H -11), 3.08 (t, 7.2 Hz, H -1%), 4.33 (t, 7.2 Hz,
(C H S ); (c) O-methyl (5Z,9Z)-5,9-tetracosenoate
8 17
(41.9%); EIMS (m/z) 378 (M ); C NMR (CDCl , l):
3
2
+
13
H -2%), 4.79 (m, H-9), 5.25, 5.27 (m, -CH
H-2), C NMR (CD OD, 600 MHz, l): 13.0 (terminal
methyl), 18.9 (C-10), 23.9 (C-11), 25.0–35.0 (methylenes),
6
ꢀCH
6
-), 5.60 (s,
2
3
13
24.8, 27.2 (-C6 H -CHꢀCH- C6 H -); DMDS derivative:
2 2
+
3
+
+
EIMS (m/z); 504 (M ), 343 (C H S ), 257 (C H S ),
20 39 2 16 33
+
+
2
7.7 (C-5), 32.7 (C-4), 34.1 (C-2%%), 49.9 (C-1%), 58.8 (C-2%),
247 (C H O S ), 161 (C H O S ).
11
19
2
2
7
13
2
.
.