Design and synthesis of novel pinacolylboronate containing combretastatin 'antimitotic agent' analogues

Article abstract of DOI:10.1016/j.tetlet.2009.04.003

We developed a procedure to synthesize pinacolyl boronate containing stilbene derivatives and used this procedure to synthesize boron-containing combretastatin analogues. The key step involves the Wittig reaction of the ylide 4-(4,4,5,5-tetramethyl-1,3,2-

Full text of DOI:10.1016/j.tetlet.2009.04.003

Tetrahedron Letters 50 (2009) 3031–3034
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Tetrahedron Letters
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Design and synthesis of novel pinacolylboronate containing combretastatin
‘antimitotic agent’ analogues
b
c,
Bhaskar C. Das a,b, , Sakkarapalayam M. Mahalingam , Todd Evans
*
*
a Department of Nuclear Medicine, Albert Einstein College of Medicine, Bronx, NY 10461, USA
b Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA
c Department of Surgery, Weill Cornell Medical College, Cornell University, New York, NY 10021, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
We developed a procedure to synthesize pinacolyl boronate containing stilbene derivatives and used this
procedure to synthesize boron-containing combretastatin analogues. The key step involves the Wittig
reaction of the ylide 4-(4,4,5,5-tetramethyl-1,3,2-dioxaboratophenyl)-methyl triphenylphosphonium
bromide 11 with 3,4,5-trimethoxy benzaldehyde in the presence of tBuONa in DMF, providing 88% yield.
We are now in a position to evaluate the biological activity of these derivatives as modulators of TGF-beta
signaling pathways.
Received 25 February 2009
Revised 31 March 2009
Accepted 1 April 2009
Available online 7 April 2009
Keywords:
Ó 2009 Elsevier Ltd. All rights reserved.
Antimitotic agent
Combretastatin
Stilbene
Pinacolatoborane
TGF-beta
For an ongoing chemical biology project involving the study of
the TGF-beta (Transforming Growth Factor-beta) pathways using
developing zebrafish embryos, we synthesized a small library of
2-substituted 2H-chromene derivatives, and after screening, we
identified a lead molecule BT7 capable of modulating a specific rel-
evant pathway, namely p-SAPK/JNK, that is known to be down-
stream of TGF-beta and can mediate Smad-independent
signaling.1,2 To increase the activity and potency of our lead mole-
cule, and for structure/activity relationship studies, we envisioned
developing a boron-based stilbene-containing small molecule li-
brary (Fig. 1) based on a hypothesis that (a) introducing chromene
isosteres like stilbene may increase potency and (b) introducing a
boron atom into a biologically active framework, might allow
interaction with a target protein not only through hydrogen bonds
but also through covalent bonds, and this interaction would pro-
duce potent biological activity, and (c) from a literature search
we also found that combretastatin A modulates TGF-beta signaling
pathway. Therefore, we considered if different CA4 analogues
could provide additional insight into TGF-beta signaling pathways
to identify new gene targets or co-factors. In this context we
undertook a project to develop synthetic methodology to synthe-
size boron-containing stilbene derivatives, and use those methods
to synthesize CA4 analogues.
O
B
Cl
O
Cl
O
Cl
OH
Cl
1
BT7
Figure 1. Structure of BT7.
Stilbene (1,2-diphenylethene) represents a prototype of a 1,2-
disubstituted olefin. Stilbene itself does not occur in nature, but
substituted stilbenes (Fig. 2) such as pinosilvin 2, resveratrol 3,3,4
diethylstilbestrol 4,5 4,4-diaminostilbene-2,2-disulfonic acid,6 ni-
tro-substituted stilbene boronate pinacol esters,7 arotinoids8, com-
bretastatin 5, 69 (Fig. 2) are among the many synthetic and natural
stilbene derivatives that are biologically active compounds.
The use of boron atoms in pharmaceutical drug design pos-
sesses a high potential for discovery of new biological activity.10–
13
Among various boron compounds synthesized, much attention
has been paid to boronic acid containing peptides such as Velcade
and DPP-IV inhibitors.12 In these boropeptides, a carboxylic acid
has been replaced by a boronic acid group.
Combretastatin A-4 (CA-4) has been found to be a potent inhib-
itor of tubulin polymerization through binding to the colchicine
binding site. It is also a cytotoxic agent toward a wide variety of
human cancer cell lines and also modulates TGF-beta signaling
* Corresponding authors. Tel.: +1 718 430 2422; fax: +1 718 430 8853 (B.C.S.).
E-mail address: bdas@aecom.yu.edu (B.C. Das).
0040-4039/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2009.04.003

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B. C. Das et al. / Tetrahedron Letters 50 (2009) 3031–3034
solvent at room temperature produced the 2,4-dichloro-6-{2-[4-
OH
R
(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-vinyl}-
phenol 1 in 86% yield with a mixture of E and Z isomers
(E:Z = 42:58) (Scheme 2).
After successfully synthesizing our target compound 1, next we
examined the scope of the Wittig reaction for the synthesis of pin-
acolylboronate-substituted stilbenes using various aryl alde-
hydes.20 The results are summarized in Table 1. The reaction
proved tolerant of both electron-withdrawing (NO2; Table 1, entry
4) and electron-donating groups (B(OH)2, di-t-butyl; Table 1, en-
HO
HO
2 : R = H
3: R = OH
OH
4
A
R1
MeO
B
MeO
R
A
B
B
R2
7
O
MeO
OMe OMe
O
R = OH: Combretastatin A-4 (CA-4) (5)
R = NH2: AC-77394 (6)
a: R1 = R2 = OMe
b: R1 = R2 = H
Table 1
Wittig reaction of boronate ylide (11) with various aldehydesa
Entry Aldehyde
Product
Yieldb
(%)
Figure 2. Biologically important stilbene derivatives.
pathways.14 From a structure/activity relationship point of view,
CA-4 belongs to the class of natural compounds related to biphe-
nyls and contains, as a key structural feature, the cis-stilbene motif.
Due to the strong biological significance of stilbene-containing
compounds, a general applicable synthetic protocol to synthesize
these highly demanding molecules with boronic acid groups would
be useful. The protocol should tolerate a large number of functional
groups in different positions on the aromatic ring. The most famil-
iar and general strategy for synthesis of boron-containing stilbenes
8 is based on disconnection A (Fig. 3) and involves the Wittig reac-
tion of the various substituted benzyl phosphonium ylide 9 with
the pinacol ester of boronate aldehyde 10.15
Although Wittig and Horner–Wadsworth–Emmons reactions
have been carried out on aldehyde derivatives of boronate es-
ters16,17 the problem with this approach (A) for the synthesis of a
library of boron-containing stilbene derivatives is the need to uti-
lize various substituted benzyl phosphonium ylides and boron-
containing aldehydes. Boron-containing aldehydes are very prone
to self-dimerization and oxidation and are unstable in air, so this
method is limited in scope. To overcome this problem, we under-
took the disconnection B approach (Fig. 3), envisaging the use of
the pinacolylboronate phosphonium ylide 11, which has not previ-
ously been explored at all. However, organotrifluoroborato phos-
phonium ylides are used to synthesize alkene derivatives by
Wittig reactions.18
To synthesize our model compound 1, we first synthesized
compound 11. 4-(4,4,5,5-tetramethyl-1,3,2 dioxaboratophenyl)-
methyl triphenylphosphonium bromide 11 was synthesized from
2-[40-(bromomethyl)phenyl]-4,4,5,5-tetramethyl-1,3,2-dioxaboro-
lane 13 in the presence of 1.01 equiv of triphenylphosphine in ace-
tonitrile at reflux conditions. The minor excess PPh3 was removed
from the product by trituration with ether 2–3 times and the prod-
uct is stable under normal atmospheric conditions (Scheme 1).
Compound 13, was synthesized by refluxing 14, NBS (N-bromo-
succinimide) and AIBN (azobisisobutyronitrile) in carbon tetra-
chloride for six hours to provide the intermediate 13 (85% yield).19
In this study, we found that 4-(4,4,5,5-tetramethyl-1,3,2-diox-
aboratophenyl)-methyl triphenylphosphonium bromide 11, when
treated with 2-hydroxy-3,5-dichloro benzaldehyde as a model sub-
strate in the presence of 3 equiv of sodium tert-butoxide in DMF as
O
CHO
B
O
1
86
72
71
76
78
H
1b E:Z = 74: 26
CHO
O
HO
B
O
B
2
3
4
5
H
HO
B
1c E:Z = 64 : 36
HO
OH
O
O
CHO
OH
B
H
OH
1d E:Z = 75 : 25
O2N
O
B
O
O N
2
CHO
OH
H
OH
1e E:Z = 55 : 45
O
B
O
CHO
N
N
H
1f E:Z = 100 : 0
All reactions were performed using 1 equiv of aldehyde and 3 equiv of tBuONa
a
in DMF for 2–12 h.
b
Isolated yield (mixture of E and Z) refers to aldehyde and the E/Z ratios were
determined by 1H NMR.
R
R
R
O
B
PPh3
O
O
A
O
O
B
O
O
B
B
12
8
9
O
Ph3P
10
11
Figure 3. Retrosynthetic analysis.

B. C. Das et al. / Tetrahedron Letters 50 (2009) 3031–3034
3033
tries 2 and 3) on the phenyl ring of the aldehyde. For the Wittig
O
B
t
reaction of aryl aldehydes containing a heteroatom such as nitro-
gen (Table 1, entry 5) gave the product 1f with good yield with
high E selectivity.
PPh , CH CN
12a, BuONa
3
3
13
O
7a
o
r.t., 4 h, 74%
90 C, 6 h
Next we focussed on target compound CA4 analogues 7a and
7b, bearing a boronic acid system that replaces the OMe group of
the natural CA-4 in ring B (Fig. 2). To synthesize 7a and 7b, the
ylide 11 was reacted with aldehydes 12a and 12b. Wittig reaction
between the aldehyde 12a and ylide 11 in the presence of tBuONa,
afforded the corresponding products 7a and 7b with 88% and 82%
yields, respectively. The products were a mixture of E and Z iso-
mers (Scheme 3). As Rf value is very close to each other, it was very
difficult to separate them, but from TLC we found Rf 0.69 is consid-
erably more fluorogenic than Rf 0.76 (1:1 ethyl acetate/hexane;
solvent system).
PPh3.Br
11
Scheme 4. One-pot synthesis of 7a.
1. NaBH4,
CHO
PPh3Br
MeO
MeO
EtOH, rt, 2 h
MeO
2. PPh3Br,
CH3CN, 90 oC, 6 h
MeO
OMe
16
OMe 15
O
Moreover, we have also developed a one-pot Wittig reaction for
synthesizing the compound 7a. In this case, 4-(4,4,5,5-tetramethyl-
1,3,2-dioxaboratophenyl)methyltriphenylphosphonium bromide
11, the intermediate generated by reaction of 13 with triphenyl-
phosphine, was reacted directly with 3,4,5-trimethoxy benzalde-
OHC
B
MeO
MeO
MeO
O
O
B
17
O
tBuONa, rt,
4 h, 80%
7a E : Z = 50 : 50
t
Scheme 5. Synthesis of 7a using approach A.
hyde 12a in the presence of BuONa as a base, and the desired
boron-containing combretastatin analogue 7a was isolated in
74% overall yield as a one-pot Wittig reaction (Scheme 4).
To compare our approach B with the approach A in (Fig. 3), we
synthesized the 7a starting from 16 using aldehyde 17 via the Wit-
tig ylide 15 (Scheme 5), and found our approach gives higher yields
with high stereoselectivity.
ments are currently underway to separate the E and Z isomers
and test the biological activity of these derivatives and to deter-
mine their utility as modulators of TGF-beta signaling pathways.
In summary, we developed a procedure to synthesize pinacolyl-
boronate containing stilbene derivatives and used this procedure
to synthesize boron-containing combretastatin analogues. Experi-
Acknowledgments
The author B.C.D. is thankful to AECOM for start up funding. T.E.
is supported by grants from the NIH (HL56182 and HL64282).
S.M.M. is thankful to K. Hema for initiating this project. The instru-
mentation in the AECOM Structural NMR Resource is supported by
the Albert Einstein College of Medicine and in part by Grants from
the NSF (DBI9601607 and DBI0331934), the NIH (RR017998) and
the HHMI Research Resources for Biomedical Sciences.
O
B
O
B
O
B
NBS, AIBN,
CCl
PPh ,
3
CH CN
3
O
O
4
O
reflux,
12 h, 85%
o
90 C, 6 h,
11
Br
92%
13
14
PPh3.Br
Supplementary data
Scheme 1. Synthesis of boronate ylide 11.
Supplementary data (experimental procedures and copies of 1H,
13C NMR) associated with this article can be found, in the online
version, at doi:10.1016/j.tetlet.2009.04.003.
O
B
References and notes
t
BuONa, DMF
O
11
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7
MeO
CHO
a: R1 = R2 = OMe; E : Z = 67 : 33
: R1 = R2 = H; E : Z = 97 : 3
R2
: R1 = R2 = OMe
: R1 = R2 = H
12
b
a
b
Scheme 3. Synthesis of boronate containing combretastatin analogues 7a–b.

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B. C. Das et al. / Tetrahedron Letters 50 (2009) 3031–3034
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with a magnetic stirring bar, a septum inlet, charged with 4-(4,4,5,5-
tetramethyl-1,3,2-dioxaboratophenyl)methyltriphenylphosphonium bromide
(11) (1 mmol), dry DMF (5 mL), and tBuONa (3 mmol) under nitrogen. The
mixture was stirred at room temperature for 5–10 min. To this solution was
added aldehyde (1 mmol) and the resulting mixture was then stirred at room
temperature for 4–6 h. The reaction mixture was treated with water (20 mL)
and was neutralized with 1 M HCl and the product was extracted with ethyl
acetate (3 ꢀ 10 mL) washed with brine and dried over Na2SO4. The product was
isolated by chromatography over silica gel.