A new strategy for the synthesis of crucigasterin A, and cytotoxic activity of this compound and its related analogues

Article abstract of DOI:10.1016/j.bmcl.2013.07.007

Stereoselective total synthesis of bioactive marine natural product crucigasterin A has been accomplished from commercially available and inexpensive l-(-)-malic acid as a starting material. Julia olefination and chelation controlled Grignard additions are the key steps involved in the present synthesis. Cytotoxic properties of crucigasterin A and its related analogues crucigasterins B and D have been evaluated. Crucigasterin A showed promising activities against both the human cervical cancer cell line and human breast adenocarcinoma cell line.

Full text of DOI:10.1016/j.bmcl.2013.07.007

Bioorganic & Medicinal Chemistry Letters 23 (2013) 5192–5194
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
A new strategy for the synthesis of crucigasterin A, and cytotoxic
activity of this compound and its related analogues q
b
Jayprakash Narayan Kumar a, Parigi Raghavendar Reddy a, Biswanath Das a, , C. Ganesh Kumar ,
⇑
Pombala Sujitha b
a Natural Product Chemistry Division, CSIR-Indian Institute of Chemical Technology, Hyderabad 500607, India
b Chemical Biology Laboratory, Medicinal Chemistry and Pharmacology Division, CSIR-Indian Institute of Chemical Technology, Hyderabad 500607, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Stereoselective total synthesis of bioactive marine natural product crucigasterin A has been accomplished
Received 13 June 2013
Revised 1 July 2013
Accepted 3 July 2013
Available online 13 July 2013
from commercially available and inexpensive
L
-(À)-malic acid as a starting material. Julia olefination and
chelation controlled Grignard additions are the key steps involved in the present synthesis. Cytotoxic
properties of crucigasterin A and its related analogues crucigasterins B and D have been evaluated. Cru-
cigasterin A showed promising activities against both the human cervical cancer cell line and human
breast adenocarcinoma cell line.
Keywords:
Total synthesis
Ó 2013 Elsevier Ltd. All rights reserved.
Crucigasterin A
Julia olefination
Chelation controlled Grignard addition
Cytotoxicity
Marine organisms are found to be a rich source of biologically
active compounds.1 Some secondary metabolites containing syn-
(2-amino-3-hydroxy) functionality were isolated from Mediterra-
nean colonial ascidian Pseudodistoma crucigaster.2 Among the five
newly isolated aliphatic unsaturated amino-alcohols crucigasetrins
A–E, crucigasterins B and E have been tested for antifungal and
antibacterial activities. Crucigasterin B was found to possess both
The retrosynthetic analysis (Scheme 1) reveals that the com-
pound 1 can be accessed starting from inexpensive and easily
available
L
-(À)-malic acid (5).
The esterification of
L
-(À)-malic acid (5) was accomplished by
reaction with thionyl chloride in absolute ethanol to provide ma-
late 6 (Scheme 2). The ester function proximal to hydroxyl group
was then reduced regioselectively using borane dimethyl sulfide
complex and 10 mol % sodium borohydride as a catalyst to furnish
the diol5 which was protected by 2,2-DMP in the presence of p-
TsOH to form the acetonide 7.6 The alcohol formed after reduction
of the ester functionality of 7 with LiAlH4,7 was oxidized with IBX
and the corresponding aldehyde was subjected to C-2 Wittig olef-
antifungal and antibacterial properties at 50
richia coli and Candida albicans, respectively, whereas crucigasterin
E was active at 100
g/ml against E. coli.2 However, to the best of
lg/ml against Esche-
l
our knowledge, cytotoxic properties were not evaluated for any
of these crucigasterins.
In continuation of our work3 on the stereoselective construction
of naturally occurring bioactive metabolites, earlier we synthesised
crucigasterins A, B and D (1–3) (Fig. 1).4
ination using Ph3PCHCOOEt to afford the a
, b-unsaturated ester 8.8
The compound 8 was treated with DIBAL-H to produce an alcohol,
which was further oxidized with IBX9 and the respective aldehyde
underwent Julia olefination using benzothiazolyl sulfone 94 and
KHMDS to furnish the triene 10.10 The acetonide group of 10 was
deprotected using a methanolic solution of p-TsOH to obtain a diol,
which on subsequent treatment with TBS-OTf and 2,6-lutidine pro-
duced the di-TBS compound11 and its TBS group of the primary
OTBS was selectively deprotected in the presence of Olah’s
reagent12 to provide 11.
While presently we evaluated the cytotoxicity of these com-
pounds we have observed that they possess significant activity
and the activity of crucigasterin A is more impressive in compari-
son to its other two analogues. This observation motivated us to
explore an alternative and more convenient synthetic route of cru-
cigasterin A. Herein, we report the new synthesis of this compound
and the cytotoxic properties of 1–3.
The free hydroxyl functionality of the compound 11 was oxi-
dized under Swern oxidation conditions and the corresponding
aldehyde was treated with benzyl amine in presence of anhydrous
MgSO4 to form the imine 4. Stereoselective chelation controlled
addition of methyl Grignard on chiral aldimine4 furnished the
q
Part 69 in the series, ‘Synthetic studies on natural products’.
Corresponding author. Tel./fax: +91 40 7160512.
E-mail address: biswanathdas@yahoo.com (B. Das).
⇑
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.07.007

J. N. Kumar et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5192–5194
5193
Table 1
IC50 values (
NH2
NH2
l
M) of test compounds against different cancer cell lines
IC50 valuesa
HeLa MDA 231
5.91 0.2 8.03 0.4 >50
5
No. Sample code
(lM)
4
1
2
OH
OH
A549
MCF-7
NH2
1
2
3
4
Crucigasterin A 14.71 0.9
Crucigasterin B >50
Crucigasterin D
19.21 1.1 10.42 1.3 15.54 0.8
7.81 1.1 39.71 0.5 12.71 0.7 13.51 1.2
0.459 0.5 0.509 0.3 0.91 0.1 13.51 1.2
Doxorubicin
OH
3
a
All tests were performed in duplicate (n = 2) and values are represented as
mean standard deviations.
Figure 1. Crucigasterins A (1), B (3) and D (3).
NH2
4
1
OH
Bn
N
4
4
OTBS
HO
O
OH
OH
O
5
Scheme 1. Retrosynthetic route for crucigasterin A.
Figure 2. IC50 values of the test compounds.
O
O
O
c
b
OR
OEt
OEt
of a five-membered chelate with the metal ion, lone pairs of 2-
silyloxy group and imino nitrogen atom.15 The physical and spec-
tral properties of 1 were found to be identical to those of crucigas-
terin A synthesised earlier.4
The present synthesis of crucigasterin A is more convenient
compared to our earlier synthesis of the compound.4 The less
expensive starting material has been used here. Earlier the selec-
tivity of a key step, aminohydroxylation, was less and the produced
regioisomers were not separable. The overall yield of the present
synthesis (13.4%) is also somewhat higher than that of the previous
one (12.6%).
Crucigasterins A, B and D were examined for in vitro cytotoxic-
ity against four cancerous cell lines: A-549 (human alveolar adeno-
carcinoma), HeLa (human cervical cancer), MDA-MB-231 (human
breast adenocarcinoma) and MCF-7 (human breast adenocarci-
noma). Doxorubicin was used as the positive control. The cells
were treated for two days with the test compounds. The MTT assay
(according to the method of Mosmann16) was applied to evaluate
RO
OH
5 : R = H
O
O
O
O
O
8
7
a
6 : R = Et
OH
O
O
e
d
4
4
4
OTBS
11
10
Bn
N
NH2
g
f
4
4
1
OTBS
OH
O
S
O
N
9
S
the activity. The IC50 value (in
lM) for each cell line was deter-
mined as the average of two independent experiments (Table 1)
(Fig. 2).
Scheme 2. Reagents and conditions: (a) EtOH, SOCl2, 95%; (b) (i) BH3ÁSMe2, cat.
NaBH4, THF, 0 °C (ii) 2,2-DMP, CH2Cl2, p-TsOH (catalytic), 0 °C to rt, 30 min., 85%
overall yield; (c) (i) LiAlH4, THF, À40 to À30 °C (ii) IBX, DMSO, CH2Cl2 (iii) C2-Wittig,
benzene, reflux, 2 h, 80% overall yield; (d) (i) DIBAL-H (2.5 equiv), CH2Cl2, À78 °C,
2 h (ii) IBX, DMSO, CH2Cl2 (iii) 9 (1.2 equiv), aldehyde (1 equiv), THF, KHMDS
(1 equiv), 62% overall yield; (e) (i) p-TsOH (catalytic), MeOH (ii) TBS-OTf (2.1 equiv),
2,6-lutidine (6 equiv), CH2Cl2 (iii) HF-pyridine (0.3 equiv), THF, 67% overall yield; (f)
(i) (COCl)2, DMSO, Et3N, CH2Cl2, À78 °C (ii) BnNH2, MgSO4, CH2Cl2, 94% overall
yield; (g) (i) CH3MgBr, Et2O, 0 °C (ii) Na/NH3, THF, À78 °C (iii) TBAF, THF, 0 °C, 53%
overall yield.
The results indicate that crucigasterin A exhibits significant
cytotoxic activity against A-549, MDA-MB-231 and MCF – 7 cell
lines. The activities against the last two cell lines are highly prom-
ising. Crucigasterin B is active against HeLa, MDA-MB-231 and
MCF-7 cell lines while crucigasterin D against all the tested four
cell lines.
In conclusion, we have carried out an alternative synthesis of
marine-derived aminoalcohol crucigasterin A through a highly ste-
reospecific and more convenient route from commercially avail-
-(À)-malic acid.17 The
desired product with 84% syn selectivity,13 and finally the reduc-
tion of the product under Birch reduction condition14 followed
by removal of the TBS ether group afforded the compound 1. The
syn selectivity of the reaction can be explained on the basis of che-
lation controlled Crams model which involves the prior formation
able, inexpensive starting material,
L
synthesis involves the well established reactions like Julia olefin-
ation and chelation controlled Grignard addition to the chiral
aldimine as the key steps. The method can be convincingly em-
ployed to synthesize the other anlogues of crucigasterin A. In

5194
J. N. Kumar et al. / Bioorg. Med. Chem. Lett. 23 (2013) 5192–5194
11. (a) Corey, E. J.; Cho, H.; Rucker, C.; Hua, D. H. Tetrahedron Lett. 1981, 22, 3455;
(b) Evans, D. A.; Ng, H. P.; Rieger, D. L. J. Am. Chem. Soc. 1993, 115, 11446.
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Y.; Lee, E. Bioorg. Med. Chem. Lett. 1905, 2004, 14.
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ties of three members of crucigasterin family, crucigasterins A, B
and D. The cytotoxic activity of crucigasterin A was found to be
quite impressive.
13. (a) Badorrey, R.; Cativiela, C.; Diaz-de-villegas Marie, D.; Galvez Jose, A.
Tetrahedron 1997, 53, 1411; (b) Cativiela, C.; Diaz-de-villegas, M. D.; Galvez, J.
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Veith, U.; Jäger, V.; Peters, E. M.; Peters, K.; Von Schnering, H. G. Angew. Chem.,
Int. Ed. Engl. 1994, 33, 1298.
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16. Mosmann, T. J. Immunol. Methods 1983, 65, 55.
17. Spectroscopic data for selected compounds are given below.
Acknowledgement
The authors thank CSIR and UGC, New Delhi for financial
assistance.
References and notes
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2012, 95, 1666; (c) Reddy, G. C.; Balasubhramanyu, P.; Salvanna, N.; Reddy, T.
S.; Das, B. Bioorg. Med. Chem. Lett. 2012, 22, 2415; (d) Reddy, P. R.; Sudhakar,
Ch.; Kumar, J. N.; Das, B. Helv. Chim. Acta 2013, 96, 289.
4. Kumar, J. P. N.; Das, B. Tetrahedron Lett. 2013, 54, 3865.
5. (a) Saito, S.; Hasegawa, T.; Inaba, M.; Nishida, R.; Fujii, T.; Nomizu, S.;
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3104.
Compound 10: [
a
]
= À9.5 (c = 0.25, CHCl3/MeOH 5:1); 1H NMR (200 MHz,
25
D
CDCl3): d 6.08-5.86 (2H, m), 5.76 (1H, m), 5.64-5.39 (2H, m), 5.01-4.90 (2H, m),
4.07 (1H, m), 3.97 (1H, m), 3.53 (1H, m), 2.21-2.52 (2H, m), 2.19-1.96 (4H, m),
1.42-1.38 (4H, m), 1.29 (6H, s); 13C NMR (50 MHz, CDCl3): d 138.9, 133.6, 131.1,
130.0, 128.4, 114.3, 108.9, 75.5, 68.9, 37.4, 33.6, 32.4, 30.0, 28.8, 26.8,; ESIMS:
m/z 251 [M + H]+, HRESIMS: cald. m/z 251.1705, found m/z 251.1701.
25
Compound 11: [
a
]
= À7.4 (c = 0.25, CHCl3/MeOH 5:1); 1H NMR (200 MHz,
D
CDCl3): d 6.10-5.89 (2H, m), 5.81 (1H, m), 5.63-5.43 (2H, m), 5.02-4.90 (2H, m),
3.76 (1H, m), 3.56-3.46 (2H, m), 2.37-2.23 (2H, m), 2.20-2.03 (4H, m), 1.91 (1H,
brs), 1.45-1.34 (4H, m), 0.89 (9H, s), 0.08 (6H, s); 13C NMR (50 MHz, CDCl3): d
138.9, 133.2, 130.8, 130.09, 128.4, 114.3, 72.8, 66.0, 37.4, 33.6, 32.4, 30.0, 28.8,
28.4, 25.8, À4.5; ESIMS: m/z 325 [M + H]+, HRESIMS: cald. m/z 325.1809, found
m/z 325.1813.
Compound 1: [
a
]
= À4.3 (c = 0.25, CHCl3/MeOH 5:1); 1H NMR (200 MHz,
25
D
CDCl3): d 6.01 (1H, m), 5.89-5.70 (2H, m), 5.61 (1H, m), 5.36 (1H, m), 5.05-4.88
(2H, m), 4.12 (1H, m), 3.66 (1H, m), 2.38-2.27 (2H, m), 2.11-1.96 (6H, m), 1.70-
1.52 (2H, m), 1.48-1.37 (2H, m), 0.82 (3H, d, J = 7.0 Hz); 13C NMR (50 MHz,
CDCl3): d 138.9, 133.5, 131.0, 130.0, 128.3, 114.3, 81.0, 51.5, 36.7, 33.8, 33.6,
32.6, 32.4, 14.1; ESIMS: m/z 224 [M + H]+, HRESIMS: cald. m/z 224.20114, found
m/z 224.20089.