Synthesis of combretastatin A-4 and erianin

Article abstract of DOI:10.3184/030823408X324751

A concise route to two anti-tubulin natural products combretastatin A-4 and erianin has been developed. Combretastatin A-4 was obtained by a Perkin reaction between 3-bromo-4-methoxyphenyl acetic acid and 3,4,5- trimethoxybenzaldehyde, hydroxyl transformation, decarboxylation with a high level of cis-selectivity (cis/trans = 95/5), and erianin was obtained by subsequent hydrogenation. The overall yields of combretastatin A-4 and erianin were 37.5 and 30.8%, respectively.

Full text of DOI:10.3184/030823408X324751

354 RESEARCH PAPER
JUNE, 354–356
JOURNAL OF CHEMICAL RESEARCH 2008
Synthesis of combretastatin A-4 and erianin
Yong Zou*a, Chun-Fen Xiaoa,b, Rong-Qing Zhonga, Wen Weia, Wen-Ming Huanga and Shu-Jie Hea
aGuangzhou Institute of Chemistry, Chinese Academy of Sciences, Guangzhou, 510650, P. R. China
bGraduate School of Chinese Academy of Sciences, Beijing, 100039, P. R. China
A concise route to two anti-tubulin natural products combretastatin A-4 and erianin has been developed.
Combretastatin A-4 was obtained by a Perkin reaction between 3-bromo-4-methoxyphenyl acetic acid and 3,4,5-
trimethoxybenzaldehyde, hydroxyl transformation, decarboxylation with a high level of cis-selectivity (cis/trans =
95/5), and erianin was obtained by subsequent hydrogenation. The overall yields of combretastatin A-4 and erianin
were 37.5 and 30.8%, respectively.
Keywords: Perkin reaction, combretastatin A-4, erianin
Combretastatin A-4 is a polyphenolic cis-stilbene, which was
first isolated from the South African tree Combretum caffrum
in 1989 by Pettit et al.1 Since then, a series of combretastatins
have been isolated from this plant (Fig.1) and many of them
were found to be anti-tubulin and anti-vascular lead compounds.
Combretastatin A-4, which binds to the same binding site
of tubulin as colchicine and exhibited excellent activity by
inhibiting mitosis and microtubule assembly,2 is one of the
most potent compound and has attracted considerable interest
in recent years.3,4 A prodrug of combretastatin A-4 (CA-
4P), the water soluble disodium phosphate, is regarded as the
flagship compound of the vascular disrupting agents (VDA),5
and is in phase III clinical trials in the USA. However, the
trans-isomer showed a dramatic decrease in its inhibitory
effects on cancer cell growth and tubulin polymerisation.
Structure–activity relationship studies6 strongly indicate that a
(Z)-stilbene configuration is essential for the cytotoxicity of the
combretastatins. Consequently, the stereo-selective synthesis of
combretastatinA-4 has received considerable attention in the last
few years, but a large-scale preparation has not been reported.
Erianin 2, 3-hydroxy-3',4',4,5'-tetramethoxylbibenzyl, was
first isolated from the Orchid Eria carinata in 19847 and was
subsequently isolated from a traditional Chinese herb, the
Dendrobium chrysotoxum Lindl, by Chinese chemists in the
1990s. Since then the biological activity of erianin has received
more attention.8-10 Structurally, erianin can be regarded as a
dihydro analogue of combretastatin A-4. Although they are
closely related to each other, they do not occur in the same
plants. Studies showed that erianin significantly inhibits the
growth of HL-60 cells and that the activity was stronger
than that of vincristine.11 Further studies showed that erianin
inhibited angiogenesis in vivo and in vitro and induced
endothelial cytoskeleton disorganisation.12 Recently, Kang et
al.'s studies13 demonstrated that erianin had no mutagenicity
in vitro and in vivo. These findings suggest that erianin is an
anti-vascular agent exhibiting therapeutic potential in vivo and
in vitro with low toxicity.
The remarkable medical importance of combretastatin
A-4 and its analogues has prompted studies of their synthesis.
Most of the syntheses of combretastatin A-4 involved a Wittig
reaction which was developed by Pettit et al.14 They reported
a five-step route from a phosphonium bromide and 3,4,5-
trimethoxybenzaldehyde, using n-butyllithium as the base.
Combretastatin A-4 was obtained as a mixture of cis and trans-
isomer in a ratio of 1/1.5. A stereoselective method developed
by the Fürstner group,15 was based on the Lindlar-type semi-
hydrogenation of an alkyne precursor assembled by 9-MeO-
9-BBN mediated Suzuki-type reaction. This gave a mixture
of combretastatin A-4 and alkane in a ratio of 86/14. Both
methods were limited by the availability of starting materials
and the preparation of intermediates. The Perkin condensation
developed by Gaukroger et al.16 seems more convenient.
Starting from 3,4,5-trimethoxyphenylacetic acid and isovanillin,
and using
a Perkin condensation and decarboxylation,
combretastatin A-4 was obtained in an overall yield of 16%
and with a stereoselective ratio of 99.8/0.2 after crystallisation.
However, the starting material 3,4,5-trimethoxyphenylacetic
acid is not commercially available and the yield is not high.
Most recently, Robinson and Taylor17 reported a Ramberg–
Backlund route starting from 3,4,5-trimethylbenzyl thiol
and protected 3-hydroxy-4-methoxybenzyl bromide to give
combretastatin A-4 and its trans-isomer in a ratio of 53/47
in their optimal result, and the overall yield is 35.28%. The
above-mentioned methods have shortcomings and are not ideal
for large-scale applications. We report a convenient synthetic
route for combretastatin A-4 1 and erianin 2 via Perkin reaction
methodology with a high level of cis-selectivity (cis/trans =
95/5 after one crystallisation) and good yields (Scheme 1). It is
the most efficient synthetic route for combretastatin A-4 1 and
erianin 2 starting from commercial materials.
Results and discussions
3-Bromo-4-methoxyphenylacetic acid 4 was obtained by
bromination of 4-methoxyphenylacetic acid 3. The Perkin
R1O
R2
R1O
R2
MeO
OMe
OH
OMe
OR3
OR3
OMe
R1 R2
Me OH
R3
H
R1
H
Me
Me
Me
R2
R3
Me
H
Combretastatin A-1
Combretastatin A-3
Combretastatin A-4
Combretastatin A-5
OMe
OMe
H
Combretastatin B-2
Combretastatin B-3
Combretastatin B-4
Erianin
H
H
H
H
H
Me
H
H
Me
H
OMe
Me
Fig. 1 Selected compounds of combretastatins and erianin.
* Correspondent. E-mail: zou_jinan@163.com

JOURNAL OF CHEMICAL RESEARCH 2008 355
CHO
OMe
COOH
COOH
MeO
MeO
MeO
COOH
OMe
1) NaOH/CuSO4
Br2
OMe
5
2) H
Br
Ac2O/Et3N
Br
OMe
OMe
4
OMe
3
MeO
MeO
MeO
MeO
MeO
COOH
quinoline/Cu
Raney-Ni
H2
MeO
OMe
OMe
OMe
6
OH
OH
OH
OMe
OMe
Combretastatin A-4
OMe
1
2
Erianin
Scheme 1 Synthesis of combretastatin A-4 and erianin.
condensation between 4 and 3,4,5-trimethoxybenzaldehyde in
the presence of acetic anhydride and triethylamine at 110°C
gave the 1,2-diarylacrylic acids 5 in 75% yield, in which the
cis-stilbene isomer (the E isomer by Cahn-Ingold rules) was
the main product (cis/trans = 95/5 after one crystallisation).
The cis-stilbene configuration of 5 was established by the
1H NMR spectrum, in which the field-effect of carboxyl group
in the cis-stilbene resulted in a remarkable down-field shift of
the acrylic alkene proton, and similarly, a noticeable field-
effect of carboxyl group can also be found in 2'-H in the B-
ring of trans-stilbene. This can be confirmed by chemical shift
of the acrylic alkene proton and the 2'-H in the B-ring of the
isomers, for 5, δ Ha = 7.82 ppm, δ Hb = 7.50 ppm, respectively,
and for the trans-isomer of 5, δHa = 6.90 ppm, δ Hb = 7.68 ppm,
respectively (Fig.2). The hydroxyl transformation from 5 to
6 in the presence of NaOH and CuSO4 at 100°C gave 6 in
75% yield. Fortunately, although the reaction was carried out
in a strong base for a few days, geometrical isomerisation did
not occur and the configuration of main product remained
cis. The decarboxylation was carried out in the presence of
quinoline and copper powder at 220°C under nitrogen, to
give the combretastatin A-4 in 71% yield. The geometrical
configuration of combretastatin A-4 was also determined by
1H NMR, for which the J(CH=CH) = 12.4 Hz for cis-stilbene.
Hydrogenation of combretastatin A-4 catalysed by Raney Ni
then gave erianin 2 in 82% yield.
difficult. TLC analysis showed that compound combretastatin
A-4 was highly lipophilic, whereas the byproducts were
poorly lipophilic. Thus, we used petroleum ether to extract
compound 1 from the dark oily reaction mixtures and leaving
the byproducts behind. The product was thus readily purified
without using column chromatography. The crude products
were then recrystallised with ethyl acetate.
In conclusion, we have developed an effective process
for syntheses of combretastatin A-4 and erianin in good
yields. Starting from 3,4,5-trimethoxybenzaldehyde and
4-methoxyphenylacetic acid, combretastatin A-4 and erianin
were synthesised with overall yields of 37.5 and 30.8%
respectively, on a multigram scale.
Experimental
The melting points were determined on Thiele apparatus and were
uncorrected. IR spectra were recorded on an Analect RFX-65A IR
spectrometer. 1H NMR were obtained from a Bruker DRX-400 MHz
spectrometer with TMS as an internal standard and CDCl3 was used
as the solvent. EI–MS analysis was performed using a Shimadzu
GCMS-QP5050A mass spectrometer. Elemental analyses were
carried out by Elementar Vario EL element analyser. All reactions
were monitored by TLC on silica gel GF254. The ratio of Z/E was
determined by HPLC. Unless otherwise mentioned, reagents were
obtained commercially and used without further purification.
Petroleum ether had a boiling point range 60–90°C.
3-Bromo-4-methoxyphenylacetic acid (4): To a stirred solution of 4-
methoxyphenylacetic acid 3 (332.00 g, 2 mol) in acetic acid (800 ml),
Br2 (106 ml, 2 mol) was slowly added over 1 h. After stirring for 3 h
at room temperature, the mixture was carefully poured into ice water
to afford a substantial light yellow precipitate. This was filtered to
give 4 as a light yellow powder in 94% yield. After recrystallising, a
white crystal was obtained. M.p. 113–114°C (lit18 114–115°C). MS,
m/z (%): 244 (M+, 50), 246 (M+ + 2, 48), 199 (100), 201 (98), 121
(10), 105 (30), 77 (70), 51 (45).
In the Perkin condensation, the product is an acid, so we
used a basification/extraction/acidification procedure as an
easy way to isolate the solid product and remove any non-
acidic impurities from the crude product. The product was
obtained in yield of 75%. In the decarboxylation reaction,
dark oily byproducts in the mixture made the purification
(7.82ppm)Ha
MeO
O
OMe
(6.90ppm)
Ha
MeO
OH
(7.50ppm)
Hb
Br
MeO
Hb(7.68ppm)
MeO
HO
OMe
O
OMe
5
Br
OMe
trans-isomer of 5
Fig. 2 Chemical shift of acrylic alkene and aromatic proton.

356 JOURNAL OF CHEMICAL RESEARCH 2008
(E)-2-(3'-Bromo-4'-methoxyphenyl)-3-(3'',4'',5''-trimethoxyphenyl)
acrylic acid (5): To a solution of compound 4 (98.40 g, 0.4 mol) and
3,4,5-trimethoxybenzaldehyde (78.40 g, 0.4 mol) in acetic anhydride
(110 ml), triethylamine (110 ml) was added. The mixture was heated
at 130°C and stirred for 5 h. After cooling, it was acidified with
concentrated hydrochloric acid, then poured into ice-water, stirred
and stored for several hours. A yellow solid was obtained. This
was filtered and dissolved in 10% NaOH (400 ml), extracted and
decoloured with ethyl acetate and the organic layers were separated.
Hydrochloric acid was added to the aqueous phase to pH 2–3.
A yellow solid was precipitated, filtered and recrystallised from ethyl
acetate to afford 5 in 75% yield. M.p. 237–238°C. IR (KBr): 3318
(COOH), 3014 (CH= ), 2937, 2838 (CH3), 1708 (C=O), 1619 (C=C),
1577, 1504, 1457, 1419, 1396, 1284, 1249, 883, 742. 1H NMR
(CDCl3, 400 MHz) δ 7.82 (s, 1H, CH=), 7.50 (d, 1H, J = 2.0 Hz,
2'–ArH), 7.17 (dd, 1H, J1 = 8.4 Hz, J2 = 2.0 Hz, 6'–ArH), 6.93 (d, 1H,
J = 8.4 Hz, 5'–ArH), 6.36 (s, 2H, 2'',6''–ArH), 3.89 (s, 3H, 4'–OCH3),
3.81 (s, 3H, 4''–OCH3), 3.57 (s, 6H, 3'',5''–OCH3). For trans-isomer
of 5: 1H NMR (CDCl3, 400 MHz) δ 7.68 (d, 1H, J = 2.2 Hz, 2'–ArH),
7.38 (dd, 1H, J1 = 8.4 Hz, J2 = 2.2 Hz,, 6'–ArH), 6.90 (s, 1H, CH=),
6.89 (d, 1H, J = 8.4 Hz, 5'–ArH), 6.69 (s, 2H, 2'',6''–ArH), 3.90 (s,
3H, 4'–OCH3), 3.84 (s, 3H, 4''–OCH3), 3.59 (s, 6H, 3'',5''–OCH3).
MS, m/z (%): 422 (M+, 100), 424 (M+ + 2, 98), 407 (20), 409 (18),
156 (18). Anal. Calc. For C19H19BrO6: C 53.9; H 4.5%. Found: C
53.8; H 4.5%.
5.49 (s, 1H, OH), 3.89 (s, 3H, 4'–OCH3), 3.84 (s, 3H, 4''–OCH3),
3.68 (s, 6H, 3'',5''–OCH3). MS, m/z (%): 316 (M+, 100), 301 (75), 241
(8), 226 (6), 211 (5), 141 (12), 115 (8), 93 (5), 57 (8). Anal.Calc. For
C18H20O5: C 68.4, H 6.3%. Found: C 68.3, H 6.3%.
Erianin (2): A solution of combretastatin A-4 (20.00 g, 63.2 mmol)
in ethanol was hydrogenated at 25–30atm pressure in the presence of
Raney-Ni (6.00 g) at ambient temperature for 8 h. The catalyst was
removed by filtration. The filtrate was evaporated in vacuum and the
residue recrystallised from petroleum/chloroform to give erianin 2 as
a pale reddish crystal in 82% yield. M.p. 78.5–79°C (lit19 79.5–80°C).
IR (KBr) cm-1: 3426 (OH), 2937, 2838 (CH3), 1590, 1459, 1384,
1
1238, 1008; H NMR(CDCl3,400 MHz) δ 6.79 (d, 1H, J = 2.0 Hz,
2'–ArH), 6.75 (d, 1H, J = 8.0 Hz, 6'–ArH), 6.62 (dd, 1H, J1 = 8.0 Hz,
J2 = 2.0 Hz, 5'–ArH), 6.34 (s, 2H, 2'',6''–ArH), 5.55 (s, w, 1H, OH),
3.85 (s, 3H, 4''–OCH3), 3.79 (s, 9H, 4', 3'', 5''–OCH3), 2.80 (s, 4H,
CH2CH2). MS, m/z (%): 318 (M+, 16), 181 (100), 137 (20). Anal.
Calc. For C18H22O5: C 67.9, H 6.9%. Found: C 67.9, H 6.9%.
We thank the Science and Technology Program of Guangdong
Province and Guangzhou City, P. R. China (2003B31603,
2006B35604002, 2007J1-C0261) for financial support.
Received 5 May 2008; accepted 17 May 2008
Paper 08/5261
doi: 10.3184/030823408X324751
(E)-2-(3'-Hydroxyl-4'-methoxyphenyl)-3-(3'', 4'', 5''-trimethoxyphenyl)
acrylic acid (6): NaOH (50.00 g) was dissolved in 500 ml water,
then 5 (64.00 g 151.2 mmol) and CuSO4 (20.00 g) were added to
the clear solution, and the mixture was stirred at 110°C for 2.5 days.
Upon cooling, filtration and acidification with 2M HCl, the solution
was extracted with ethyl acetate. The organic layer was washed
with brine, dried and evaporated in vacuum to afford a yellow solid
which recrystallised with ethyl acetate and petroleum to afford 6 as
pale yellow crystals in 75% yield. M.p. 233–234°C. IR (KBr) cm-1:
3401 (OH), 2998 (CH3), 2944 (CH2), 2838–2518 (COOH), 1679
(C=O), 1581, 1508, 1459, 1378 (CH3), 1334, 1184, 998, 946 (ArH).
1H NMR (CDCl3, 400 MHz) δ 7.78 (s, 1H, CH= ), 6.89 (d, 1H,
J = 8.4 Hz, 5'–ArH), 6.85 (d, 1H, J = 2.0 Hz, 2'–ArH), 6.75 (dd, 1H,
J1 = 8.4 Hz, J2 = 2.0 Hz, 6'–ArH), 6.39 (s, 2H, 2'',6''–ArH), 5.76 (w,
2H, COOH, OH), 3.89 (s, 3H, 4'–OCH3), 3.81 (s, 3H, 4''–OCH3),
3.57 (s, 6H, 3'',5''–OCH3). MS, m/z (%): 360 (M+, 100), 345 (20, M–
OCH3), 327 (5), 285 (15). Anal. Calc. For C19H20O7: C 63.3, H 5.6%.
Found: C 63.3, H 5.6%.
Combretastatin A-4 (1): A mixture of compound 6 (40.00 g,
111.2 mmol) and copper powder (40.00 g, 62.5 mmol) in 200 ml
quinoline was stirred at 200°C for 3 h. After cooling, ethyl acetate
was added and the copper was filtered off. The filtrate was washed
with 2M hydrochloric acid, and the water layer was separated and
extracted with ethyl acetate. The organic layers were combined and
washed with brine, dried (MgSO4) and evaporated in vacuum to
afford a brown viscous solid, which was recrystallised from ethyl
acetate to afford combretastatin A-4 1 as colourless crystals in 71%
yield. M.p. 116–117°C (1it:15 115–116°C). IR (KBr): 3424 (OH),
3002 (CH= ), 2938, 2836, 1610, 1579, 1508, 1459, 1419, 1328, 1182,
1025, 1004, 944, 881, 854, 796. 1H NMR (CDCl3, 400 MHz) δ 6.91
(d, 1H, J = 2.0 Hz, 2'–ArH), 6.79 (dd, 1H, J1 = 8.0 Hz, J2 = 2.0 Hz,
6'–ArH), 6.71 (d, 1H, J = 8.0 Hz, 5'–ArH), 6.51 (s, 2H, 2'',6''–ArH),
6.45 (d, 1H, J = 12.4 Hz, CH=), 6.42 (d, 1H, J = 12.4 Hz, CH= ),
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