Synthesis and antiproliferative activity of unnatural enantiomers of 7-epi-goniofufurone and crassalactone C

Article abstract of DOI:10.1016/j.bmcl.2008.08.093

A facile synthesis of 7-epi-(-)-goniofufurone as well as the first synthesis of (-)-crassalactone C was achieved starting from d-xylose. A comparison of their in vitro antitumour activities with those observed for the corresponding naturally occurring enantiomers was provided.

Full text of DOI:10.1016/j.bmcl.2008.08.093

Bioorganic & Medicinal Chemistry Letters 18 (2008) 5178–5181
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and antiproliferative activity of unnatural enantiomers
of 7-epi-goniofufurone and crassalactone C
a
a
a
a
Velimir Popsavin a, , Goran Benedekovic , Bojana Sreco , Mirjana Popsavin , Jovana Francuz ,
*
´
´
b
b
´
´
Vesna Kojic , Gordana Bogdanovic
a Department of Chemistry, Faculty of Sciences, University of Novi Sad, Trg D. Obradovi´ca 3, 21000 Novi Sad, Serbia
b Institute of Oncology Sremska Kamenica, Institutski put 4, 21204 Sremska Kamenica, Serbia
a r t i c l e i n f o
a b s t r a c t
Article history:
A facile synthesis of 7-epi-(ꢀ)-goniofufurone as well as the first synthesis of (ꢀ)-crassalactone C was
Received 21 July 2008
Revised 24 August 2008
Accepted 26 August 2008
Available online 29 August 2008
achieved starting from D-xylose. A comparison of their in vitro antitumour activities with those observed
for the corresponding naturally occurring enantiomers was provided.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Styryl-lactones
Stereocontrolled syntheses
D-Xylose
7-Epigoniofufurone
Crassalactone C
Goniofufurone
Unnatural enantiomers
Antitumor agents
7-epi-(+)-Goniofufurone (1, Fig. 1) is a naturally occurring styryl
lactone that was isolated from the stem bark of the tropical plant
Goniothalamus giganteus (Annonaceae).1 Its structure was eluci-
dated by spectral methods and the relative configuration was
established by X-ray crystal data.1 The absolute stereochemistry
of 1 was established from the total synthesis of 7-epi-(ꢀ)-gon-
iofufurone (ent-1).2 The combination of the unique structures
and a promising biological activity3 has prompted a number of
syntheses of similar styryl lactones.4,5 A new styryl lactone, (+)-
crassalactone C (2), was recently isolated from the leaves and twigs
of the Asian three Polyalthia crassa.6 Its structure and the relative
stereochemistry were assigned based on spectral data, and its
absolute configuration was confirmed by semi-synthesis starting
from the isolated (+)-goniofufurone. Recently, we have completed
the total synthesis of the naturally occurring styryl lactones 1 and
2,7 and evaluated their antiproliferative activity against several tu-
mour cell lines. As a consequence of the in vitro antitumour activ-
ity of these natural products, we were especially keen to prepare
and evaluate the opposite enantiomers of 1 and 2, since it is well
known that enantiomers of certain bioactive compounds may exhi-
OH
OH
6
O
O
3
7
7
2
Ph
Ph
Ph
4
1
1
5
OH
HO
O
O
O
O
O
O
O
ent -1
1
O
O
3'
2'
1'
6
O
Ph
Ph
O
O
O
3
7
7
2
Ph
4
1
1O
5
HO
OH
ent-
2
2
Figure 1. Structures of 7-epi-(+)-goniofufurone (1) and (+)-crassalactone C (2), and
of the corresponding unnatural enantiomers ent-1 and ent-2
bit improved potencies8 or even different biological activities
altogether.9
Herein, we report the total synthesis of the unnatural enantio-
mers of 7-epi-goniofufurone (ent-1) and crassalactone C (ent-2)
starting from D-xylose, and provide a comparison of their antipro-
liferative activities with those observed for the natural products 1
* Corresponding author. Tel.: +381 21 385 27 68; fax: +381 21 454 065.
E-mail address: popsavin@ih.ns.ac.yu (V. Popsavin).
and 2. Three syntheses of ent-1 were reported,2,10 but there was no
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.08.093

V. Popsavin et al. / Bioorg. Med. Chem. Lett. 18 (2008) 5178–5181
5179
O
O
OH
OMe
OMe
Ph
O
O
O
O
7
Ph
O
O
7
Ph
BnO
OH
O
OBn
O
O
O
O
OH
O
ent-
O
2
6
ent-
1
9
Scheme 1. Retrosynthetic analysis of target compounds ent-1 and ent-2.
record in the literature concerning the biological activity of this
molecule. Neither the synthesis nor biological activity of ent-2
was reported in the literature.
tion of a A newly developed two-step
literature method.12
sequence that included di-O-benzylation of 3, followed by a regio-
selective C-5 deprotection gave the primary alcohol 5. Oxidation of
5 under modified Moffat conditions14 gave 6 in 71% overall yield
(from three steps). The preceding preparation of 6 was accom-
plished in 62% overall yield over four linear steps.12 The Z-selective
Wittig olefination15 of 6 with methyl (triphenylphosphoranylid-
ene)acetate gave the Z-unsaturated ester 7 (83%) as a dominant
reaction product, accompanied with a minor amount (12%) of the
corresponding E-isomer (not shown in the reaction scheme). It
was expected that the key intermediate 9 could be obtained by
an acid-catalyzed methanolysis of 7. The procedure followed here
is analogous to that previously developed by Prakash and Rao11
for the conversion of the isopropylidene analogue of 7 into lactone
9. Accordingly, compound 7 was refluxed in dry methanol in the
presence of a catalytic amount of sulphuric acid (2%). Although a
complete conversion of starting compound was observed after
4.5 h, only traces of the desired lactone 9 were detected in the reac-
tion mixture. The unsaturated lactone 8 was formed as a major
reaction product under these reaction conditions. However, when
Retrosynthetic analysis of ent-1 and ent-2 is outlined in Scheme
1. Both targets ent-1 and ent-2 contain five contiguous stereocen-
ters but differ in stereochemistry and functionality at C-7. The
presence of the cinnamoyl moiety in ent-2 implies that this mole-
cule could be obtained from ent-1 through a regioselective Mitsun-
obu reaction with cinnamic acid. Further disconnection of ent-1
shows that it can be derived from the protected aldehyde 9 by
three successive transformations that involve a hydrolysis of the
acetal functionality, stereo-selective addition of phenyl magne-
sium bromide to the liberated aldehyde group, followed by re-
moval of benzyl protective group. The intermediate 9 should be
accessible from the aldehyde 6, through an adopted literature pro-
cedure.11 Dialdose 6 is readily available from
D-xylose in several
synthetic steps.12
Synthesis of both ent-1 and ent-2 is shown in Scheme 2. The
synthetic sequence commenced with the formation of the pro-
tected dialdose 6 from
D
-xylose derivative 313 by a slight modifica-
OR
OH
O
OH
O
O
O
O
Ref. 13
c
b
OH
O
O
O
RO
O
BnO
BnO
O
O
HO
OH
3 R = H
a
D-Xylose
6
5
4 R = Bn
d
OMe
OMe
CO2Me
O
OH
OMe
HO
f
O
e
O
Ph
OMe
OBn
+
O
OBn
BnO
OBn
O
O
O
O
O
O
O
7
9
8
10
+
O
O
NOE
Ph
OH
Ph
OH
Ph
H
H
O
g
h
O
O
O
Ph
Ph
OH
7
+
3
O
OBn
OH
O
O
O
O
O
O
O
O
11
12
ent-1
ent-2
i
Scheme 2. Reagents and conditions: (a) BnBr, NaH, DMF, 0 °C for 0.5 h, then rt for 1 h; (b) H2–Pd/C (0.05 equiv of Pd), EtOH, EtOAc, rt, 1.5 h, 80% from 3; (c) DCC, anhyd H3PO4,
Py, DMSO, rt, 3.5 h, 89%; (d) Ph3P:CHCO2Me, MeOH, 0 °C for 0.5 h, then rt for 1.5 h, 83%; (e) i—H2SO4, MeOH, reflux 4.5 h, ii—NaHCO3, 35 °C, 1 h, 74% of 9, 4% of 8; (f) i—9:1 TFA/
H2O, 0 °C for 0.5 h, then rt for 0.5 h, ii—PhMgBr/THF, Et2O, ꢀ7 °C, 3 h, 56% of 11 (from 9), 3% of 10 (from 9); (g) H2–Pd/C, MeOH, rt, 72 h, 64%; (h) Cinnamic acid, Ph3P, THF,
DEAD, 0 °C for 1 h, then rt for 2.5 h, 53% of ent-2, 16% of 12; (i) Ph3P, DEAD, PhMe, reflux, 2.5 h, 40%.

5180
V. Popsavin et al. / Bioorg. Med. Chem. Lett. 18 (2008) 5178–5181
the reaction solution was alkalized to pH 9 (NaHCO3) and stirred
for 1 h at 35 °C, the bicyclic lactone 9 was predominantly formed
as a result of the stereospecific Michael ring closure in 8. The bicy-
clic lactone 9 was isolated in a 74% yield, along with a minor
amount of 8 (4%). These results are in accord with Prakash and
Rao’s assumption that lactonisation precedes Michael attack in this
process, but dismisses their hypothesis that the tetrahydrofuran
ring closure in 8 occurs under the acidic conditions.11
Hydrolytic removal of the dimethyl acetal protection in 9 affor-
ded the corresponding aldehyde, which was subsequently treated
with phenyl magnesium bromide to give two diastereomeric alco-
hols 10 and 11 in a 1:19 ratio and 59% combined yield (calculated
to 9). Major isomer 11 was converted to target ent-1 after removal
of the benzyl protecting group. The physical and spectral data16 of
thus obtained sample ent-1 were identical to those previously re-
ported.10a This new synthesis of ent-1 proceeds in seven steps with
16% overall yield calculated to starting compound 3. Along with
the Gracza and Jäger approach, it appears to be one of the most effi-
cient routes to this molecule yet disclosed.17
Compound ent-1 was converted to (ꢀ)-crassalactone C (ent-2)
upon treatment with cinnamic acid, under the standard Mitsunobu
conditions.18 The target ent-2 that was isolated in 53% yield, dis-
played physical and spectral properties19 in reasonable agreement
with those previously reported for the natural (+)-crassalactone
C.6,7 A minor amount of 5,7-anhydro derivative 12 (16%) was also
obtained from this reaction. The side-product 12 was presumably
formed by a competitive intramolecular nucleophilic displacement
process.18 In order to verify this assumption, the Mitsunobu reac-
tion was performed in absence of cinnamic acid (Ph3P, DEAD,
PhMe";, 2.5 h), whereupon the oxetane 1220 was obtained as a ma-
jor product. The stereochemistry at C-7 was resolved on the basis
of a NOE interaction between H-3 and H-7 that is consistent with
tone C (ent-2) showed a potent antiproliferative activity towards
HL-60 cells, while the corresponding natural enantiomer 2 was
completely inactive against this cell line. Moreover, compound
ent-2 demonstrated a 27- and 10-fold greater cytotoxicity in Raji
and HeLa cells, respectively, when compared to the natural product
2. At the same time, this molecule demonstrated a 5-fold greater
cytotoxicity than DOX towards the Raji cells. Remarkably, tricyclic
derivative 12 showed sub-micromolar antiproliferative activities
against HL-60, Raji and HeLa malignant cells. In fact, it was the
most active compound towards these cell lines. Finally, molecule
12 demonstrated a 37- and 7-fold stronger cytotoxicity in HL-60
and Raji cell lines when compared to doxorubicin, respectively.
It is notable that natural styryl lactones and analogues present
promising and diverse biological activities. Some important mech-
anisms of action of this class of bioactive compounds have been re-
cently reviewed.3 The remarkable activity of the lactone 12
towards the all malignant cell lines tested deserves some addi-
tional comments. It is possible that this analogue acts as a specific
alkylating agent, and that the observed cytotoxicities originated
from an irreversible covalent binding to cellular serine or thiol pro-
teases. However, further studies will be needed to verify this
hypothesis.
In summary, the unnatural styryl lactones (ꢀ)-7-epi-goniofufu-
rone (ent-1), (ꢀ)-crassalactone C (ent-2), as well as a new conform-
ationally constrained analogue of (ꢀ)-goniofufurone (12) have
been synthesized and evaluated for their in vitro antitumour activ-
ities against a number of human neoplastic cell lines. Compound
ent-1 demonstrated a sub-micromolar antiproliferative activity
against Jurkat cells, comparable to that recorded for the reference
compound (DOX). The highest cytotoxicity of ent-2 was observed
in Raji cell line, with approximately 5-fold higher potency then
doxorubicin. Compound 12 showed the most potent antiprolifera-
tive activity towards HL-60 cells being ca. 37-fold more active than
the reference compound (DOX). Based upon the potent antitumour
activities of ent-1, ent-2 and 12, as well as upon their non-toxicity
against normal MRC-5 cells, we believe that these compounds may
serve as convenient leads in the synthesis of more potent and
selective antitumour agents. Finally, from a synthetic perspective,
the fact that both enantiomers efficiently inhibit tumour cells
growth also means that both are equally valuable synthetic targets.
In this sense, the synthesis of ent-1 and ent-2, along with our pre-
ceding approach to 1 and 2,7 represents a new enantiodivergent
route that provided the opportunity to access the cytotoxic proper-
ties of not only the natural products, but also their unnatural
enantiomers.
L
-glycero-
D
-ido stereochemistry of 12.
A side-by-side comparison of natural 1 and ent-1, as well as of 2
and ent-2 in a range of cytotoxic assays21 is presented in Table 1.
Antitumour activity of 12 was also preliminary evaluated, since
this compound formally represents a structurally constrained ana-
logue of (ꢀ)-goniofufurone. The doxorubicin (DOX) was used as a
reference compound.
In general, the unnatural enantiomer of crassalactone C (ent-2)
exhibited antiproliferative activities towards the all tested cell
lines while compound ent-1 was active against K562, Jurkat and
HeLa cell lines but was completely inactive against Raji cells.
Remarkably, all styryl lactones were devoid of any significant
toxicity against the normal foetal lung fibroblasts (MRC-5). A com-
parison of IC50 values revealed not only that the unnatural 7-epi-
(ꢀ)-goniofufurone (ent-1) was slightly more potent than the
corresponding natural enantiomer 1 against HL-60 cells, but also
that this molecule was approximately 345-fold more potent than
the natural enantiomer 1 against Jurkat cell line. On the other
hand, ent-1 showed a similar potency as the reference compound
(DOX) in the same cell line. The unnatural enantiomer of crassalac-
Acknowledgment
Financial support from the Ministry of Science of the Republic
of Serbia (Project No. 142005) is gratefully acknowledged.
References and notes
1. Fang, X. P.; Anderson, J. E.; Chang, C. J.; McLaughlin, J. L.; Fanwick, P. E. J. Nat.
Prod. 1991, 54, 1034.
2. (a) Shing, T. K. M.; Tsui, H.-C.; Zhou, Z-H. Tetrahedron 1992, 48, 8659; (b) Shing,
T. K. M.; Tsui, H.-C. J. Chem. Soc. Chem. Commun. 1992, 432.
Table 1
In vitro cytotoxicity of 1, ent-1, 2, ent-2 and 12
3. For reviews on biological activity of natural styryl lactones and analogues, see:
(a) de Fatima, A.; Modolo, L. V.; Conegero, L. S.; Pilli, R. A.; Ferreira, C. V.; Kohn,
L. K.; de Carvalho, J. E. Curr. Med. Chem. 2006, 13, 3371; (b) Mereyala, H. B.; Joe,
M. Curr. Med. Chem.. Anti-Cancer Agents 2001, 1, 293; (c) Blazquez, M. A.;
Bermejo, A.; Zafra-Polo, M. C.; Cortes, D. Phytochem. Anal. 1999, 10, 161.
4. For recent reviews on the synthesis of styryl lactones, see: (a) Mondon, M.;
Gesson, J.-P. Curr. Org. Synth. 2006, 3, 41; (b) Zhao, G.; Wu, B.; Wu, X. Y.; Zhang,
Y. Z. Mini-Rev. Org. Chem. 2005, 2, 333.
5. (a) Kapitán, P.; Gracza, T. Tetrahedron: Asymmetry 2008, 19, 38; (b) Prasad, K. R.;
Gholap, S. L. J. Org. Chem. 2008, 73, 2; (c) Yadav, K. V.; Agrawal, D. J. Chem. Soc.
Chem. Commun. 2007, 5232. and references therein.
6. Tuchinda, P.; Munyoo, B.; Pohmakotr, M.; Thinapong, P.; Sophasan, S.; Santisuk,
T.; Reutrakul, V. J. Nat. Prod. 2006, 69, 1728.
a
Compound
IC50
(
lM)
HL 60
Jurkat
Raji
HeLa
MRC-5
1
22.02
13.93
>100
1.87
0.025
0.92
18.64
0.05
25.45
15.67
28.45
0.03
1.25
>100
15.46
0.57
0.45
2.98
0.89
4.38
11.25
1.18
0.97
>100
>100
>100
>100
>100
0.10
ent-1
2
ent-2
12
DOX
0.07
a
IC50 is the concentration of compound required to inhibit the cell growth by 50%
compared to an untreated control.

V. Popsavin et al. / Bioorg. Med. Chem. Lett. 18 (2008) 5178–5181
5181
7. Popsavin, V.; Benedekovic´, G.; Srec´o, B.; Popsavin, M.; Francuz, J.; Kojic´, V.;
17. Gracza and Jäger10b synthesis: 16% yield over seven linear steps; Zhao et al.10a
synthesis: 11% yield over nine steps Shing et al.2 synthesis: 10 steps and 12%
overall yield.
´
Bogdanovic, G. Org. Lett. 2007, 9, 4235.
8. (a) de Fatima, A.; Kohn, L. K.; de Carvalho, J. E.; Pilli, R. A. Bioorg. Med. Chem.
2006, 14, 622; (b) Liu, S.; Kulp, S. K.; Sugimoto, Y.; Jiang, J.; Chang, H.-L.; Dowd,
M. K.; Wan, P.; Lin, Y. C. Anticancer Res. 2002, 22, 33; (c) Boger, D. L.; Hong, J. J.
Am. Chem. Soc. 2001, 123, 8515.
9. Nishimura, Y.; Umezawa, Y.; Adachi, H.; Kondo, S.; Takeuchi, T. J. Org. Chem.
1996, 61, 480.
10. (a) Su, Y.-L.; Yang, C.-S.; Teng, S.-J.; Zhao, G.; Ding, Y. Tetrahedron 2001, 57,
2147; (b) Gracza, T.; Jäger, V. Synthesis 1994, 1359; (c) Gracza, T.; Jäger, V.
Synlett 1992, 191.
11. Prakash, K. R. C.; Rao, S. P. Tetrahedron Lett. 1991, 32, 7473.
12. Popsavin, V.; Krstic´, I.; Popsavin, M.; Srec´o, B.; Benedekovic´, G.; Kojic´, V.;
18. Mitsunobu, O. Synthesis 1981, 1.
19. Selected data for ent-2: Mp 147–150 °C,
[a
]
ꢀ110.1 (c 0.5, EtOH). 1H
20
D
NMR (250 MHz, CDCl3): d 2.54 (d, 1H, J2a,2b = 18.6 Hz, H-2a), 2.69 (dd, 1H,
J2a,2b = 18.6, J2b,3 = 5.9 Hz, H-2b), 4.25 (br s, 1H, OH), 4.27 (dd, 1H,
J6,7 = 9.3, J5,6 = 2.4 Hz, H-6), 4.43 (br s, 1H, H-5), 5.01 (m, 2H, H-3 and H-
4), 6.00 (d, 1H, J6,7 = 9.3 Hz, H-7), 6.46 (d, 1H, J2 ,3 = 15.8 Hz, H-20), 7.35–
0
0
7.60 (m, 10H, 2ꢁ Ph), 7.78 (d, 1H, J2 ,3 = 15.8 Hz, H-30). 13C NMR
0
0
(62.9 MHz, CDCl3):
d 35.79 (C-2), 72.80 (C-7), 73.10 (C-5), 77.09 (C-3),
82.43 (C-6), 87.01 (C-4), 116.59 (C-20), 127.70, 128.35, 128.65, 128.95,
128.98, 130.95, 133.71 and 136.71 (2ꢁ Ph), 147.29 (C-30), 167.46 (C-10),
175.40 (C-1). HR MS (ESI): m/z 403.1133 (M++Na). Calcd for C22H20NaO6:
´
Bogdanovic, G. Tetrahedron 2006, 62, 11044.
13. Popsavin, M.; Popsavin, V.; Vukojevic´, N.; Miljkovic´, D. Collect Czech. Chem.
403.1152.
ꢀ48.1 (c 0.5, CHCl3). 1
H
20
Commun. 1994, 59, 1884.
14. Jones, G. H.; Moffat, J. G. Methods Carbohydr. Chem. 1972, 6, 315.
15. Valverde, S.; Martin-Lomas, M.; Herradon, B.; Garcia-Ochoa, S. Tetrahedron
20. Selected data for 12: Mp 146°C (CH2Cl2/hexane), [
a]
D
NMR (250 MHz, CDCl3): d 2.83 (d, 1H, J2a,2b = 18 Hz, H-2a), 2.93 (dd, 1H,
J2a,2b = 18, J2b,3 = 4.1 Hz, H-2b), 2.84 (dd, 1H, J5,6 = 4.2, J6,7 = 2.6 Hz, H-6), 5.09 (d,
1H, J3,4 = 3.5 Hz, H-4), 5.37 (dd, J2b,3 = 4.1, J3,4=3.5 Hz, H-3), 5.50 (d, 1H,
J5,6 = 4.2 Hz, H-5), 5.52 (d, 1H, J6,7 = 2.6 Hz, H-7), 7.50–7.32 (m, 5H, Ph); NOE
contact: H-3 and H-7. 13C NMR (62.9 MHz, CDCl3): d 36.04 (C-2), 79.24 (C-3),
84.92 (C-4), 85.08 (C-6), 85.31 (C-5), 89.11 (C-7), 125.0, 128.51, 128.85 and
138.56 (Ph), 174.16 (C-1). HR MS (ESI): m/z 233.0805 (M++H). Calcd for
1987, 43, 1895.
16. Selected data for ent-1: Mp 205–206 °C, [
20
a]
D
ꢀ98.3 (c 0.47, Me2CO); lit.2 mp
20
208–209 °C, [a]
ꢀ92.5 (c 1.1, Me2CO). 1H NMR (250 MHz, DMSO-d6 + D2O): d
D
2.53 (d, 1H, J2a,2b = 18.7 Hz, H-2a), 2.85 (dd, 1H, J2a,2b = 18.7, J2b,3 = 6.5 Hz, H-2b),
3.60 (d, 1H, J5,6 = 2.8 Hz, H-5), 3.84 (dd, 1H, J5,6 = 2.8, J6,7 = 8.3 Hz, H-6), 4.73 (d,
1H, J6,7 = 8.3 Hz, H-7), 4.78 (d, 1H, J3,4 = 4.7 Hz, H-4), 4. 94 (dd, 1H, J3,4 = 4.7,
J2b,3 = 6.5 Hz, H-3), 7.21–7.37 (m, 5H, Ph). 13C NMR (62.9 MHz, DMSO-d6):d36.42
(C-2), 72.16 (C-7), 73.48 (C-5), 77.35 (C-3), 85.35 (C-6), 88.48 (C-4), 127.54,
128.35, 128.84 and 142.40 (Ph), 177.44 (C-1). LR-MS (ESI): m/z 289 (M++K), 273
(M++Na).
C
13H13O4: 233.0808.
21. Cytotoxic activities were evaluated by using standard MTT assay after
exposure of cells to the tested compounds for 72 h. Results are presented as
mean values of three independent experiments done in quadriplicates.
Coefficients of variation were <10%.