Concise synthesis of Bacillus anthracis exosporium tetrasaccharide via two-stage activation of allyl glycosyl donor strategy

Article abstract of DOI:10.1016/j.tetlet.2011.05.089

A highly efficient glycosylation protocol recently developed in our laboratory has been utilized in the short synthesis of the tetrasaccharide 1, an antigen important to the development of carbohydrate-based diagnostic tools and vaccines against anthrax.

Full text of DOI:10.1016/j.tetlet.2011.05.089

Tetrahedron Letters 52 (2011) 3912–3915
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Concise synthesis of Bacillus anthracis exosporium tetrasaccharide via
two-stage activation of allyl glycosyl donor strategy
⇑
Yun Wang, Xing Liang, Pengfei Wang
Department of Chemistry, University of Alabama at Birmingham, Birmingham, AL 35294, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A highly efficient glycosylation protocol recently developed in our laboratory has been utilized in the
short synthesis of the tetrasaccharide 1, an antigen important to the development of carbohydrate-based
diagnostic tools and vaccines against anthrax. The protocol employs allyl glycosides as building blocks
and improves the overall synthetic efficiency.
Received 2 May 2011
Revised 17 May 2011
Accepted 19 May 2011
Available online 27 May 2011
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Carbohydrate
Anthrax
Allyl glycoside building blocks
Step-economical synthesis
Anthrax is a fatal infectious disease in humans and other mam-
mals caused by spores of the bacterium Bacillus anthracis. The
durable spores enter the host, germinate, grow, and kill the host
within several days of infection. Due to their lethality, ready pro-
duction, resistance to harsh conditions, and ease of spread, B.
anthracis spores are considered as the foremost threat in biological
warfare. Unfortunately, there are few medical remedies for bioter-
rorism preparedness against B. anthracis spores. The known pre-
exposure prevention against anthrax is vaccination. Development
of vaccines targeting spores could inactivate spores prior to their
establishment of infectious loci and vegetative cell outgrowth,
and thus prevent development of anthrax.
tic tools and vaccines against spores of the known virulent strains
of B. anthracis.2f,l,3,4 The potential biomedical applications of the
tetrasaccharide highlights the need for rapid access to the material
in a sufficient amount. Herein we report a highly efficient route to
this tetrasaccharide from allyl glycoside building blocks.
We have been pursuing step economy-oriented carbohydrate
synthesis in solution phase for rapid access to oligosaccharides of
biochemical and biomedical significance. An efficient synthesis de-
mands (1) efficient and effective glycosylation methods with min-
imal anomeric position manipulation of glycosyl donors and (2)
tactically planned protecting group strategy to minimize selective
protecting/deprotecting steps. Along this direction, we have re-
cently demonstrated a simple and mild glycosylation reaction
employing only allyl glycosides as building blocks. Thus, a prop-
1-enyl glycosyl donor readily derived from an allyl glycoside can
undergo a highly effective glycosylation reaction with an allyl gly-
cosyl acceptor upon activation with NIS or NIS/TfOH at room tem-
perature.5,6 As prop-1-enyl glycosyl donors are directly isomerized
from allyl glycosides, time-consuming anomeric group replace-
ment and intermediate purification are avoided. The isomerization
of the anomeric allyl group to the corresponding prop-1-enyl is
typically of high efficiency with a variety of facile methods avail-
able in literature.7 Mindful of the ease of anomeric allyl group
placement and allyl to prop-1-enyl isomerization, and simple con-
ditions of glycosylation with prop-1-enyl donors, we anticipate
that the new protocol would be suitable for rapid oligosaccharide
synthesis with high overall synthetic efficiency.
The exosporium is the outermost layer of a spore, the first point
of contact of spores with host cells and the source of spore surface
antigens. It can be the target of diagnostic and preventative proce-
dures. In 2004, Turnbough and coworkers found multiple copies of
a
tetrasaccharide on BclA (Bacillus collagen-like protein of
anthracis) of the B. anthracis exosporium.1 This unique tetrasaccha-
ride, that is, 4,6-di-deoxy-4-(3-hydroxy-3-methylbutamido)-2-O-
methyl-b-
-l-rhamnopyranosyl-(1?2)-
the non-reducing end with a novel
D
-glucopyranosyl-(1?3)-
-rhamnopyranose (1), is capped at
-anthrose. Since its structure
a-L-rhamnopyranosyl-(1?3)-
a
L
D
was published in 2004, synthesis of the tetrasaccharide 1 and re-
lated anthrose-containing oligosaccharides has become a high pri-
ority.2 Immunological studies of the synthetic tetrasaccharides and
their conjugates demonstrate that the tetrasaccharide 1 is a prom-
ising antigen for the development of carbohydrate-based diagnos-
With the new glycosylation approach, the tetrasaccharide can
be synthesized from simple allyl glycoside building blocks 2–4 as
shown in the retrosynthesis (Scheme 1). The building blocks 2
⇑
Corresponding author. Tel.: +1 205 9965625; fax: +1 205 9342543.
E-mail address: wangp@uab.edu (P. Wang).
0040-4039/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2011.05.089

Y. Wang et al. / Tetrahedron Letters 52 (2011) 3912–3915
3913
O
O
O
c
O
c
HO
BnO
2
HO
BnO
O
OH
α
O
+
+
O
b
O
b
HO
BnO
O
AcO
OH
3
α
OBn
O
b
O
HO
O
O
N3
O
β
a
BnO
N
OH
H
a
1
O
HO
O
HO
4
Piv
OMe
Scheme 1. Retrosynthesis of tetrasaccharide 1.
and 3 were obtained from commercially available
L
-rhamnose in
rasaccharide 10 in 59% yield along with a trace amount of the b
anomer. It is unclear why the yields of producing 10 and 11 were
lower than those of 8 and 9, respectively.
six steps.5b
The building block 4a was prepared from commercially avail-
able -fucose in five steps (Scheme 2). Thus, installation of the ano-
D
We demonstrated that the tetrasaccharide skeleton 10 was con-
structed in 17 steps from commercially available free monosaccha-
rides, a synthesis shorter than previously reported work (typically
taking over 30 steps to reach similar complexity from commer-
cially available starting materials).2 The anomeric allyl group of
the tetrasaccharide 10 can be functionalized through ozonolysis,13
olefin metathesis,14 or thiol-ene click chemistry15 for future immo-
bilization or conjugation before or after2a,j the subsequent se-
quence of global deprotection, reduction of the azido group and
amide formation. In this work we explored a different approach.
Thus, from the tetrasaccharide 10, the anomeric allyl group was
converted to the (3-hydroxyl)propyl group under hydroboration
conditions to provide 13 in 87% yield (Scheme 5). The subsequent
one step global deprotection of 13 removed benzyl protecting
groups and reduced the azido group to the amino group through
palladium catalyzed hydrogenolysis. A standard amide formation
reaction with HATU and the butanoic acid2a,j achieved the final
product in 72% yield over two steps. The obtained tetrasaccharide
equipped with a functionalized anomeric group provided another
option for further elaboration.
meric allyl group followed by selective benzylation at 3-OH
provided the allyl fucoside 5 in 76% yield in two steps.8 Subsequent
placement of a O-2 pivalolyl group generated 6 in 89% yield. The
axial 4-OH of 6 was then converted to an equatorial azido group
via a triflate intermediate in 91% yield.9
With the building blocks in hand, we began to assemble the tet-
rasaccharide (Scheme 3). The allyl glycoside 3 was first isomerized
to the corresponding prop-1-enyl glycosyl donor with hydrogen-
activated [Ir(COD)(PMePh2)2]PF6.5,10 Upon activation of the vinyl
glycoside with NIS/TfOH in the presence of the acceptor 2, the de-
sired disaccharide 7 was obtained in 82% yield after alkali workup.
The b anomer of 7 was not produced in a detectable amount. How-
ever, when the same procedure was repeated with the donor 3 and
the acceptor 7, the trisaccharide 8 was obtained as a mixture of
two anomers (a/b = 7:1) in 84% yield after alkali workup. These
two anomers were separated by column chromatography. The ano-
meric stereochemistry of the newly constructed glycosidic bonds
1
in 7 and 8 was confirmed by the JC1–H1 coupling constants.11,12
The iterative process continued with the donor 4 and the acceptor
8
a
to provide the tetrasaccharide 10 after the O-2 pivalolyl group
In summary, a concise synthesis of the important tetrasaccha-
ride target was achieved by utilizing the effective and efficient gly-
cosylation method recently developed in our laboratory.5 This
method employed allyl glycosides as building blocks. The anomeric
allyl protecting group of glycosyl donors was first converted to the
more reactive prop-1-enyl leaving group. Without purification, the
prop-1-enyl intermediate was activated in the presence of an allyl
glycosyl acceptor for glycosidic bond construction. This procedure
avoided time-consuming anomeric group removal/installation and
intermediate purification. Moreover, iterative use of allyl
of 9 was replaced by a methyl group. With the neighboring direct-
ing effect of the O-2 pivalolyl group, stereoselective construction of
the new glycosidic bond was accomplished based on 1H NMR
analysis.
Alternatively, synthesis of the tetrasaccharide was attempted
via a [2+2] route (Scheme 4). Thus, the disaccharide 11 was pre-
pared in 66% yield from the allyl glycosyl donor 4 and the acceptor
12, derived from 3 by removing the O-3 acetyl group. The reducing
end expansion with the donor 11 and the acceptor 7 led to the tet-
OH
1) allyl alcohol, AcCl, 92%
O
PivCl, Py
89%
D-Fucose
BnO
2)Bu2SnO; BnBr, CsF, 84%
OH
O
5
OH
BnO
Tf2O, Py; NaN3.
91%
O
O
N3
BnO
PivO
POiv
O
O
6
4
α
Scheme 2. Synthesis of building block 4.

3914
Y. Wang et al. / Tetrahedron Letters 52 (2011) 3912–3915
O
O
O
BnO
82%
BnO
O
O
BnO
84%
1. [Ir(COD)(PMePh2)2]PF6
(H2-activated); NIS/TfOH, 2
2. NaOMe
AcO
O
OBn
BnO
HO
3
OBn
7
1. [Ir(COD)(PMePh2)2]PF6
(H2-activated); NIS/TfOH, 7
2. NaOMe
O
(α/β = 7:1)
O
BnO
O
O
BnO
O
BnO
1. 4, [Ir(COD)(PMePh2)2]PF6
O
BnO
(H2-activated); 8, NIS/TfOH
BnO
O
2. NaOMe
O
OBn
O
O
BnO
85%
BnO
O
O
OBn
O
N3
BnO
OBn
O
BnO
OR
HO
9
: R = H
OBn
NaH, MeI
92%
8
10: R = Me
Scheme 3. Glycosidic bond construction.
66%
O
59%
4
10
O
1. [Ir(COD)(PMePh2)2]PF6
[Ir(COD)(PMePh2)2]PF6
(H2-activated); NIS/TfOH, 7
BnO
O
(H2-activated); 12
O
N3
OBn
NIS/TfOH
BnO
2. NaOMe; MeI
11
OMe
O
12 =
O
BnO
HO
OBn
Scheme 4. Synthesis of the tetrasaccharide via a [2+2] route.
O
OH
O
OH
O
O
BnO
HO
O
BnO
HO
O
O
1) H2, Pd/C
10
2) (CH3)2C(OH)CH2COOH
O
O
9-BBN; H2O2
NaOH
BnO
HO
HATU, DIPEA,DMF
O
O
OBn
OH
87%
72%
O
BnO
O
HO
O
O
O
O
N3
OBn
N
H
OH
BnO
HO
HO
OMe
OMe
1
13
Scheme 5. Synthesis of the tetrasaccharide.

Y. Wang et al. / Tetrahedron Letters 52 (2011) 3912–3915
3915
180; (c) Kubler-Kielb, J.; Vinogradov, E.; Hu, H.; Leppla, S. H.; Robbins, J. B.;
glycosides for oligosaccharide synthesis does not require any
armed/disarmed property tuning, significantly simplifying build-
ing block preparation, and improving the overall synthetic
efficiency.
ˇ
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We thank the University of Alabama at Birmingham for support.
Supplementary data
Supplementary data associated with this article can be found, in
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