Design and synthesis of harzialactone analogues: Promising anticancer agents

Article abstract of DOI:10.1016/j.bmcl.2010.10.100

New homologues of harzialactone were synthesized using D-glucose as chiral template. Wittig reaction to introduce aromatic moiety in 10 and chemoselective anomeric oxidation of 13 were used as key reactions in our synthesis. Anticancer activity of these target molecules was assessed against five cancer cell lines, P388D1, HL60, COLO-205, Zr-75-1 and HeLa. Both compound 5 and 6, showed significant activity against colon cancer (COLO-205) and cervical cancer (HeLa) and moderate with others. To the best of our knowledge, this is the first report of harzialactone analogues as potent inhibitors of human colon and cervical cancer.

Full text of DOI:10.1016/j.bmcl.2010.10.100

Bioorganic & Medicinal Chemistry Letters 20 (2010) 7243–7245
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of harzialactone analogues: Promising anticancer agents
Vishwas U. Pawar a, Sougata Ghosh b, Balu A. Chopade b, Vaishali S. Shinde a,
⇑
a Garware Research Centre, Department of Chemistry, University of Pune, Pune 411 007, India
b Institute of Bioinformatics and Biotechnology, University of Pune, Pune 411 007, India
a r t i c l e i n f o
a b s t r a c t
Article history:
New homologues of harzialactone were synthesized using D-glucose as chiral template. Wittig reaction
to introduce aromatic moiety in 10 and chemoselective anomeric oxidation of 13 were used as key reac-
tions in our synthesis. Anticancer activity of these target molecules was assessed against five cancer cell
lines, P388D1, HL60, COLO-205, Zr-75-1 and HeLa. Both compound 5 and 6, showed significant activity
against colon cancer (COLO-205) and cervical cancer (HeLa) and moderate with others. To the best of
our knowledge, this is the first report of harzialactone analogues as potent inhibitors of human colon
and cervical cancer.
Received 21 July 2010
Revised 1 October 2010
Accepted 20 October 2010
Available online 26 October 2010
Keywords:
Harzialactone
Ó 2010 Elsevier Ltd. All rights reserved.
D-Glucose
Chemoselective oxidation
Anticancer activity
Various biologically active metabolites have been obtained from
marine microorganisms. Numata and co-workers1 isolated and
characterised various secondary metabolites from a fungal strain
of Trichoderma harzianum OUPS-N115 originally separated from
the sponge Halichondria okadai. Among these, (+)-harzialactone A
1 (Fig. 1), exhibited cytotoxic activity against the P388 lymphocytic
leukaemia test system in cell cultures. Souza et al.2 isolated and
characterised 2, diastereomer of harzialactone A from the Eutypa-
like strain FED-3. Due to the potent bioactivity and unique struc-
tural feature of harzialactone A, synthesis of it remains a centre
of interest of chemists.3–9 Mereyala et al.3 developed chiron
approach for the synthesis of harzialactone A 1 starting from
O
O
O
1
5
O
O
O
O
O
3
OH
OH
OH
cis-(−), 2
trans-(−), 3
trans-(+), 1
Harzialactone A
Br
O
O
O
O
OH
OH
cis-(+), 4
OH
cis-(−), 5
cis-(−), 6
Figure 1. Harzialactone A and its analogues.
D
-glucose for the first time and assigned correct stereochemistry
as (3R,5R). Further, they also synthesized all its stereoisomers 2,
3, 4 from
-glucose.4 Recently, Chen et al.8 developed a chemo-
of 5 which is homologue of 4 and to evaluate its anticancer activity.
Though the racemic compound 5 is known,11 its bioevaluation was
not carried out. During the synthetic course towards 5, we also ob-
tained bromine substituted compound 6, whose anticancer activity
was also evaluated.
D
enzymatic route to harzialactone A and all its stereoisomers using
recombinant F. proliferatum lactonase (reFPL) starting from phenyl
acetone. Sabitha et al.9 reported the synthesis of harzialactone A
and its (3R,5S)-isomer 4 utilising the chiral-epoxide opening with
thioacetal and diastereoselective reductions as key reactions.
Though, the syntheses of stereoisomers of 1 have been reported,
their potential as anticancer agents has not yet been assessed. In
spite of the significant anticancer activity of harzialactone A, the
field of bioevaluation of its other stereoisomers and also the
synthesis of homologues is unattended. This need prompted us
to synthesize homologues and to estimate their anticancer
property. As a part of our continuing interest in the synthesis of
Retrosynthetic analysis of homologues
5 and 6 indicates
(Scheme 1) that it could be derived from anomers 13 by chemose-
lective oxidation of the anomeric hydroxyl group of the lactol and
this lactol could be obtained by acetonide deprotection of the
3-deoxy-gulose derivative 12. Compound 12 can be derived by
double bond reduction of styrene derivative 11 which in turn could
be obtained by Wittig olefination of in situ generated ylide from
benzyl triphenylphosphonium bromide with aldehyde 10. This
aldehyde can be synthesized from
procedures with slight modifications.4,12
Our synthesis started with -glucose (Scheme 2) which was
converted into 3-deoxy- -gulose derivative 8 according to known
procedures.12 Cleavage of 5,6-acetonide protection was selectively
D-glucose according to known
bioactive molecules from D
-glucose,10 we planned for the synthesis
D
D
⇑
Corresponding author. Tel.: +91 20 25601395; fax: +91 20 25691728.
E-mail address: vsshinde@chem.unipune.ac.in (V.S. Shinde).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.10.100

7244
V. U. Pawar et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7243–7245
R
O
O
O
O
OH
O
O
5, R = H
6, R = Br
OH
13
12
11
O
OH
O
O
O
O
6
H
4
O
O
5
1
D
-glucose
O
3
O
2
O
10
O
8
O
Scheme 1. Retrosynthetic analysis.
OH
O
O
O
HO
O
O
O
O
b)
a)
O
D-glucose
c)
O
O
O
O
9
O
8
7
O
O
O
e)
O
d)
O
H
O
O
O
O
12
O
Z:E = 85:15
O
10
11
f)
g)
O
(S)
O
OH
O
13
5
(R) OH
OH
h)
Br
i)
(S)
O
(R) OH
6
Scheme 2. Synthesis of (À)-5 and (À)-6. Reagents and conditions: (a) H2, Pd/C, AcOEt, rt, 2 h, 94%; (b) 30% HClO4, THF, 0 °C, <5 min, 88%; (c) NaIO4, satd NaHCO3, CH2Cl2, 0 °C
to rt, 2.5 h, 87%; (d) Ph3P+CH2PhBrÀ, n-BuLi, THF, À5 °C to rt, 12 h, (Z:E = 85:15), 71%; (e) H2, Pd/C, AcOEt, rt, 2 h, 96%; (f) cat. H2SO4, THF/H2O (4:1), 65 °C, 2.5 h, 85%; (g)
Ag2CO3–Celite, toluene, reflux, 3 h, 79%; (h) Br2, BaCO3, dioxane/H2O (2:1), (in dark), rt, 1 h, 94%; (i) H2, Pd/C, AcOEt, 260 psi, rt, 24 h, 96%.
carried out using 30% HClO4 to afford diol 9 in 88% yield with
recovery of some starting material. This diol on oxidative cleavage
using sodium meta-periodate afforded aldehyde 10 which was
found to be relatively unstable and used as such for further reac-
tion. Aldehyde 10 was subjected to the Wittig olefination with
in situ generated benzylidenetriphenylphosphonium ylide from
benzyl triphenylphosphonium bromide to afford styrene derivative
11 as a diastereomeric mixture13 of Z:E in the ratio of 85:15 as evi-
dent from 1H NMR spectrum in 71% yield. This olefinic mixture was
then hydrogenated by using 10% Pd/C in ethyl acetate which affor-
ded compound 12 in 96% yield. Cleavage of 1,2-isopropylidene
functionality at 65 °C using cat. H2SO4 afforded a mixture of hemi-
acetal 13 in good yield. In the final step, selective anomeric oxida-
tion was carried out by using Ag2CO3/Celite14 which afforded
target molecule 5 as a white crystalline solid in 79% yield. Appear-
ance of carbonyl frequency at 1773 cmÀ1 in IR along with peak at
3436 for hydroxyl group and signal at 177.7 ppm in 13C NMR con-
pound 5 from 6, ring de-bromination of 6 was achieved under
hydrogenation condition using 10% Pd/C in ethyl acetate at
260 psi afforded compound 5 in 96% yield and with overall 90%
yield from 13.
The cytotoxicity of compounds 5 and 6 was tested in vitro
against five cancer cell lines, P388D1 (murine lymphocytic leukae-
mia), HL60 (human promyelocytic leukaemia),16 COLO-205
(human colon cancer), Zr-75-1 (human epithelial breast cancer),17
HeLa (human epithelial cervical cancer)18 by MTT colorimetric
assay using Mitomycin C as a standard.19 The assay results were
then used to evaluate the corresponding percent inhibition. These
synthetic compounds exhibited interesting in vitro anticancer
activity. As shown in Table 1, compound 5 (at 100 lg/mL) was
found to be very potent inhibitor of COLO-205 and HeLa. It showed
moderate inhibition against Zr-75-1 as well as HL-60. Compound 5
exhibited reduced inhibitory activity against P388D1 cell lines as
compared to other cell lines. Compound 6 (at 100 lg/mL) showed
firmed the formation of
a
-hydroxy-
c
-lactone. In order to improve
very high activity against COLO-205 (Table 2). It also inhibited
HeLa to a significant level even at lower concentration which
was comparable with Mitomycin C. P388D1 and HL60 were inhib-
ited by 6 to a moderate level while it exhibited poor inhibition
against Zr-75-1. Thus, substitution of bromine at para position in
phenyl ring as in compound 6 increased its anticancer activity
the yield of target compound, we tried another method15 for
anomeric oxidation with Br2/BaCO3. This bromine mediated oxida-
tion worked smoothly with 2.5 equiv of bromine and 1.1 equiv of
BaCO3 within 1 h to give the required lactone, but in this reaction
we also observed electrophilic bromine substitution in phenyl ring
exclusively at para position to give 6 in 94% yield. Targeting com-
against four of the cancer cell lines at 100 lg/mL and also showed

V. U. Pawar et al. / Bioorg. Med. Chem. Lett. 20 (2010) 7243–7245
7245
Table 1
In vitro anticancer activity of compound 5 on P388D1, COLO-205, Zr-75-1, HL60 and HeLa
Sample (
lg/ml) 5
% Inhibition on various cell lines
Zr-75-1
P388D1
COLO-205
HL60
HeLa
100
80
60
40
20
10
5
45.06 1.06
40.00 0.64
38.27 3.21
37.65 3.56
35.18 1.06
31.48 1.42
27.78 0.71
55.39 0.06
69.29 0.60a
50.66 0.76
43.61 0.25
37.88 0.23
37.44 1.27
36.12 0.76
35.68 1.53
29.51 0.18
51.26 0.39
49.03 0.74
43.22 2.23
40.00 0.37
38.70 0.02
36.77 1.12
30.96 0.37
28.38 0.74
53.21 0.46
62.42 0.46a
40.97 0.20
38.98 0.26
33.97 0.32
31.31 0.27
31.27 0.29
23.44 0.32
56.48 0.12
a
65.35 0.45
35.95 0.47
25.45 0.76
22.83 0.30
16.79 0.31
09.18 0.90
56.27 0.71
Mitomycinb
The data is indicated as the mean SEM (n = 3).
a
Denoting more significant value (P < 0.05).
Concentration of mitomycin used was 5 lg/ml.
b
Table 2
In vitro anticancer activity of compound 6 on P388D1, COLO-205, Zr-75-1, HL60 and HeLa
Sample (
lg/ml) 6
% Inhibition on various cell lines
Zr-75-1
P388D1
COLO-205
HL60
HeLa
100
80
60
40
20
10
5
52.46 0.71
40.74 0.35
40.12 0.73
34.56 1.08
31.48 2.14
25.92 1.78
10.49 0.61
55.39 0.06
75.85 0.75a
42.29 1.02
38.32 0.50
37.00 0.53
36.56 0.56
35.24 0.00
33.03 0.25
20.95 0.27
51.26 0.39
52.9 0.74
47.74 0.37
32.58 0.00
39.35 0.36
24.51 0.74
22.58 0.00
20.64 0.73
53.21 0.46
69.14 0.06a
62.51 0.08
60.58 0.12
57.48 0.13
54.40 0.06
52.51 0.23
52.34 0.19
56.48 0.12
a
72.17 0.00
43.30 0.45
40.94 0.30
29.39 0.32
26.24 1.06
23.09 0.31
56.27 0.71
Mitomycinb
The data is indicated as the mean SEM (n = 3).
a
Denoting more significant value (P < 0.05).
Concentration of mitomycin used was 5 lg/ml.
b
significant activity against COLO-205 and HeLa even at lower
concentration (5 g/mL) when compared to parent compound 5.
By contrast, compound 5 was found to be more potent against
Zr-75-1 as compared to 6 at both 5 and 100 g/mL. At lower con-
centration (5 g/mL), compound 5 was found to be more active
of new compounds) associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2010.10.100.
l
l
l
References and notes
against P388D1 and HL60 than compound 6.
In conclusion, we have synthesized two new homologues of
harzialactone by chiron approach starting from D-glucose in 10 steps
with overall yields of 20% and 23% for compounds 5 and 6 respec-
tively. Br2/BaCO3 mediated reaction apart from being chemoselec-
tive also resulted in bromination of phenyl ring exclusively at para
position. Thus compound 5 and 6, were studied for their anticancer
activity against five cancer cell lines which indicated their specificity
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Acknowledgement
We gratefully acknowledge University of Pune, Pune (BCUD/
OSD/184-2009) for financial support. We are thankful to Professor
M. S. Wadia for insightful discussions. S.G. is thankful to University
of Pune for research stipend. VUP is thankful to UGC (New Delhi,
INDIA) for fellowship in the form of SRF.
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