The total synthesis of an aurone isolated from Uvaria hamiltonii: Aurones and flavones as anticancer agents

Article abstract of DOI:10.1016/j.bmcl.2003.07.003

The naturally occurring aurone 1, isolated from Uvaria hamiltonii, and a series of aurones analogues based structurally on known tubulin binding agents were prepared and evaluated for anticancer activity. Aurone 20 was the most active (IC₅₀ K562 50 nM) and caused significant G₂/M cell-cycle arrest.

Full text of DOI:10.1016/j.bmcl.2003.07.003

Bioorganic & Medicinal Chemistry Letters 13 (2003) 3759–3763
The Total Synthesis of an Aurone Isolated from Uvaria
hamiltonii: Aurones and Flavones as Anticancer Agents
Nicholas J. Lawrence,a,b,* David Rennison,b,d Alan T. McGownc,d
and John A. Hadfieldc,d
aDepartment of Chemistry, Cardiff University, PO Box 912, Cardiff CF10 3TB, UK
bDepartment of Chemistry, UMIST, PO Box 88, Manchester M60 1QD, UK
cCentre for Molecular Drug Design, Department of Chemistry, University of Salford, Manchester M5 4WT, UK
dCancer Research UKDepartment of Drug Development, Paterson Institute for Cancer Research, Christie Hospital NHS Trust,
Wilmslow Road, Manchester M20 4BX, UK
Received 19 June 2003; revised 31 July 2003; accepted 31 July 2003
Abstract—The naturally occurring aurone 1, isolated from Uvaria hamiltonii, and a series of aurones analogues based structurally
on known tubulin binding agents were prepared and evaluated for anticancer activity. Aurone 20 was the most active (IC50 K56250
nM) and caused significant G2/M cell-cycle arrest.
# 2003 Elsevier Ltd. All rights reserved.
Interest in flavonoids as pharmacological agents has
been considerable since these natural products have
been shown to constitute the active ingredient of many
folk medicines.1 Aurones have been reported to display
analgesic activity,2 while aurone derivatives from plant
extracts have been used in the treatment of thyroid dis-
eases.3 Flavones are known to possess anticancer activ-
ity,4 along with selective inhibition of both cyclin-
dependent kinases (CDK’s)5 and tyrosine kinases.6
Aurone 1 is one of several constituents isolated from the
extracts of Uvaria hamiltonii by Wani and co-workers.7
The constituents were isolated by way of a bioassay-
guided fractionation based on DNA strand-scission8
and 9KB assays. While aurone 1 appeared to be inactive
in the 9KB assay it offered the greatest potency, of the
compounds isolated, in the DNA strand-scission assay.
It should be noted that in comparison to a bleomycin
standard, compounds of this nature actually offer weak
DNA strand-scission activity.
The development of a flexible synthesis of 1 would also
provide access to analogues to support our programme
of research into tumour vascular targeting agents.12 The
spatial relationship between the two aromatic rings of
combretastatin A-4, colchicine and similar drugs is an
important structural feature that determines their ability
to bind to tubulin.13 The chalcone 2 is a potent inhibitor
of tubulin polymerisation,14,15 which we believe adopts
an s-trans conformation.16 The aurones and flavones
would effectively provide conformationally restricted
analogues of 2. We hoped that this would present an
insight into the importance of aryl ring orientation
about the rotatable bonds a and c in influencing cyto-
toxicity and tubulin-binding properties.
The resemblance of aurone 1 to the tumour vascular
targeting agent combretastatin A-4,9ꢀ11 made it an
appealing target for total synthesis.
*Corresponding author. Tel.: +44-29-2087-4000x77109; fax: +44-29-
2087-4030; e-mail: lawrencenj1@cardiff.ac.uk
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.07.003

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N. J. Lawrence et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3759–3763
Not widely distributed in nature, aurones are one of the
least common types of flavonoid, and so have received
little attention in comparison to the structurally related
and widely investigated flavones and isoflavones.
Therefore, not surprisingly, general methods for their
synthesis are not so common. The most general route to
aurones involves the acid or base catalyzed condensa-
tion of substituted 3(2H)-benzofuranones with aromatic
aldehydes. We therefore needed to develop a route to
the appropriate benzofuranones.
deemed to be identical to the natural product originally
disclosed by Wani and co-workers by 1H and 13C
NMR.7 The total synthesis of 1 was therefore achieved
in four steps in 37% overall yield.
With the benzofuranones 5 and 9 in hand, a series of
substituted aurones was prepared (Scheme 4). The
yields were generally lower when a deactivated (elec-
tron-rich) benzaldehyde was used (Table 1). Fluorinated
aurones are included in the series since we wish to
investigate the ability of combretastatin derivatives to
Benzofuranone 5 was prepared in two steps from 3,4,5-
trimethoxyphenol (3) in 51% overall yield, via the reac-
tion with chloroacetic acid and excess sodium hydride,
followed by polyphosphoric acid (PPA) mediated cycli-
zation of the first-formed phenoxyacetic acid
(Scheme 1).
4
Benzofuranone 9 was prepared in the same way as 5 by
PPA promoted intramolecular Friedel–Crafts reaction
of the phenoxyacetic acid 8. Phenol 7 was prepared by
sequential Baeyer–Villiger oxidation of the benzalde-
hyde 6, using m-chloroperbenzoic acid, and base-cata-
lysed methanolysis of the resulting formate (Scheme 2).
The overall yield of benzofuranone 9 from 6 was 38%.
The synthesis of aurone 1 and its regioisomer 14
required the use of the TBDMS-protected benzaldehyde
11, prepared in high yield from 3,4-dihydroxybenz-
aldehyde (10) using tert-butyldimethylsilyl chloride in
the presence of imidazole.17 On reaction with the
benzofuranones 5 and 9 in the presence of neutral alu-
mina—a protocol developed by Varma and Varma18—
the TBDMS-protected aurones 12 and 13 were obtained
in good yield. Subsequent treatment with tetra-
butylammonium fluoride afforded aurones 1 and 14
(Scheme 3). The aurone 1 was fully characterised and
Scheme 3. Reagents and conditions: (i) TBDMSCl, imidazole, DCM,
rt, overnight, 92%; (ii) Al2O3, DCM, rt, overnight, 74% (12) and 97%
(13); (iii) TBAF, DCM, rt, 30 min, 98% (12 to 1) or 78% (13 to 14).
Scheme 1. Reagents and conditions: (i) ClCH2CO2H, NaH, DMF, rt,
overnight, 85%; (ii) PPA, 80 ꢁC, 8 h, 60%.
Scheme 4. Reagents and conditions: (i) Al2O3, DCM, rt, 1–3 days,
yields (see Table 1).
Table 1. Synthesis of substituted aurones
0
0
0
R4
R7
R3
R4
R5
Yield (%)
15
16
17
18
OMe
OMe
OMe
OMe
H
H
H
H
H
OH
F
OMe
OMe
OMe
OMe
H
H
H
F
71
31
62
71
F
19
20
21
22
H
H
H
H
OMe
OMe
OMe
OMe
H
OH
F
OMe
OMe
OMe
OMe
H
H
H
F
78
48
80
82
Scheme 2. Reagents and conditions: (i) m-CPBA, DCM, rt, 18 h,
93%; (ii) NaOMe, MeOH, rt, 4 h, 88%; (iii) ClCH2CO2H, NaH,
DMF, rt, overnight, 81%; (iv) PPA, 80 ꢁC, 8 h, 58%.
F

N. J. Lawrence et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3759–3763
3761
induce tumour vasculature damage via 18F positron
emission tomography (PET).12
and its regioisomer 14 (IC50 9.7 mM) display similarly
poor cell-growth inhibitory properties. The aurone 20
(IC50 50 nM) bearing the greatest resemblance to com-
bretastatin A-4 displays the highest activity, along with
the monofluorinated aurone 21 (IC50 0.11 mM). Again,
as with the corresponding 5,6,7-trimethoxyaurones, it is
clear that the structurally related flavones bearing a
6,7,8-trimethoxyphenyl A-ring arrangement display the
greatest cytotoxicity, with flavone 30 (IC50 40 nM) being
equipotent to aurone 20 (IC50 50 nM).
In all cases a single geometric isomer (Z) was obtained,
this generally being more thermodynamically stable
than the (E)-isomer.19 Although an unambiguous
assignment of our aurone series was not made, the
values are consistent with those present in the literature
for known (Z)-aurones.20 In a single step—using the
protocol developed by Wheeler and co-workers21—the
aurones were transformed into their corresponding fla-
vones22 by simply heating in the presence of potassium
cyanide (Scheme 5 and Table 2). The reaction is
impeded by the presence of a phenol group. The re-
arrangement is also clearly complicated by the presence
of two fluorine atoms.
Preliminary results from modeling of the structures are
shown in Figure 1.24 Superposition of energy-minimized
structures of 20 and 30 onto the chalcone 2, indicates
that a reasonable degree of overlap can be achieved in
both cases. The positions of the A-ring and linker are
similar. The main difference in both cases seems to be
the tilt of the B-ring, which arises in 2 from the presence
of the a-methyl group.
The cell growth inhibitory properties of the substituted
aurones and flavones (summarised in Table 3) were
determined in the K562human chronic myelogenous
leukaemia cell line using the MTT assay as detailed by
Edmondson et al.23 The IC50 value represents the con-
centration which results in a 50% inhibition in cell
growth after 5 days incubation.
The effects of the most active compounds upon the cell
cycle were measured by flow cytometry. The results
show aurones 20 and 21 to cause significant arrest of the
cell cycle at the G2/M point, relative to the untreated
control, at this stage consistent with the behaviour of
tubulin binding agents. As expected (from the lack of
cytotoxicity), the natural product aurone 1 and its
regioisomer 14 fail to induce significant levels of G2/M
block, along with flavone 30 (Table 4).
It is clear that the 5,6,7-trimethoxyaurones (19–22) offer
greater cell growth inhibitory properties than their cor-
responding 4,5,6-trimethoxy isomers. It would appear
that either the presence of a methoxyl group at the 7-
position of the A-ring or the lack of a methoxyl group
at the 4-position is responsible for the differences in
cytotoxicity. Natural product aurone 1 (IC50 12 mM)
Table 3. Cell growth inhibitiona against the K562cell line
IC50 (mM)
IC50 (mM)
IC50 (mM)
1
12
>50
18
>50
>50
0.0002
14
19
20
21
22
9.7
24
25
26
29
30
31
42
22
>50
0.83
0.04
0.59
15
16
17
18
2
0.15
0.05
0.11
0.15
aAs measured by the MTT assay after 5 days incubation of the drug
with the cells cultured at 37 ꢁC.
Scheme 5. Reagents and conditions: (i) KCN, EtOH/DCM, reflux, 12
h, yields (see Table 2).
Table 2. Synthesis of substituted flavones
0
0
0
R5
R8
R3
R4
R5
Yield (%)
23
24
25
26
27
OMe
OMe
OMe
OMe
OMe
H
H
H
H
H
OH
H
OH
F
OH
H
H
H
H
F
0
73
20
57
0
OMe
OMe
OMe
OMe
F
28
29
30
31
32
H
H
H
H
H
OMe
OMe
OMe
OMe
OMe
OH
H
OH
F
OH
H
H
H
H
F
0
80
32
64
0
OMe
OMe
OMe
OMe
Figure 1. Superposition of the structures aurone 20 (A) and flavone 30
(B) onto chalcone 2. Structures 20 and 30 are shown in red, while 2 is
shown in blue.
F

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Table 4. Effects upon the cell cyclea
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G0–G1
S
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Microtubule
assemblya IC50 (mM)
Microtubule
assemblya IC50 (mM)
1
>50
>50
>50
>50
22
21
25
26
30
31
>50
>50
>50
25
16
17
14
20
2
>50
0.5
Colchicine displacementb IC50 (mM)
20
30
60
40
aMicrotubule assembly assay was carried out as previously descri-
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50% inhibition in tubulin assembly as measured by an increase in
turbidity at 350 nm.
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Acknowledgements
We thank the EPSRC for a studentship (to D.R.) and
for Research Grants (GR/L52246: NMR spectrometer;
GR/L84391: chromatographic equipment). We thank
Cancer Research UK and the Department of Chem-
istry, UMIST for additional support of this work and
Nigel Smith of the PICR Cell Culture Unit for valuable
assistance during the cell growth inhibition experiments.

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