Chemoenzymatic syntheses of tumor-associated carbohydrate antigen globo-H and stage-specific embryonic antigen 4

Article abstract of DOI:10.1002/adsc.200800129

Gangliosides have attracted much attention due to their important biological properties. Herein we report the first chemoenzymatic syntheses oftwo globo series of ganglioside oligosaccharides, Globo-H 1 and stage-specific embryonic antigen-4 (SSEA-4) 2. The common precursor SSEA-3 pentasaccharide for these two compounds was assembled rapidly using the pre-activation-based one-pot glycosylation method. The stereoselectivity in forming the 1,2-cis linkage in SSEA-3 was attributed to a steric buttressing effect of the donor rather than electronic properties ofthe glycosyl donors. SSEA-3 was then successfully fucosylated by the fucosyltransferase WbsJ and sialylated by sialyltransferases CST-I and PmST1 producing Globo-H and SSEA-4, respectively.

Full text of DOI:10.1002/adsc.200800129

DEDICATED CLUSTER
FULL PAPERS
DOI: 10.1002/adsc.200800129
Chemoenzymatic Syntheses of Tumor-Associated Carbohydrate
Antigen Globo-H and Stage-Specific Embryonic Antigen 4
Zhen Wang,a Michel Gilbert,b Hironobu Eguchi,c Hai Yu,d Jiansong Cheng,d
Saddam Muthana,d Luyuan Zhou,a Peng George Wang,c Xi Chen,d
and Xuefei Huanga,*
a
Department ofChemistry, The University ofToledo, 2801 W. Bancroft Street, MS 602, Toledo, Ohio 43606, USA
Fax : (+1)-419-530-4033; e-mail: Xuefei.huang@utoledo.edu
National Research Council Canada, Institute for Biological Sciences, Glycobiology Program, 100 Sussex Drive, Ottawa,
b
ON K1A 0R6, Canada
c
The Ohio State University, Departments ofBiochemistry and Chemistry, 484 West 12th Avenue, Columbus, OH 43210,
USA
d
Department ofChemistry, University ofCalifornia, One Shields Avenue, Davis, CA USA
Received: February 29, 2008; Published online: April 11, 2008
th
Dedicated to Prof. Chi-Huey Wong on the occasion of his 60 birthday for his ground breaking contributions
to chemistry.
Supporting information for this article is available on the WWW under http://asc.wiley-vch.de/home/.
Abstract: Gangliosides have attracted much atten- buttressing effect of the donor rather than electronic
tion due to their important biological properties. properties ofthe glycosyl donors. SSEA-3 was then
Herein we report the first chemoenzymatic syntheses successfully fucosylated by the fucosyltransferase
oftwo globo series ofganglioside oligosaccharides,
WbsJ and sialylated by sialyltransferases CST-I and
Globo-H 1 and stage-specific embryonic antigen-4 PmST1 producing Globo-H and SSEA-4, respective-
(SSEA-4) 2. The common precursor SSEA-3 penta- ly.
saccharide for these two compounds was assembled
rapidly using the pre-activation-based one-pot glyco- Keywords: carbohydrates; chemoenzymatic synthe-
sylation method. The stereoselectivity in forming the sis; natural products; oligosaccharides; stereoselec-
1,2-cis linkage in SSEA-3 was attributed to a steric tivity
Introduction
receptors for uropathogenic Escherichia coli[12–14] and
human parvovirus B19.[15] In addition, SSEA-4 is a
The globo series ofgangliosides, including Globo-H 1 marker for human cancer cells[16–19] and the expression
and stage-specific embryonic antigen-4 (SSEA-4) 2, level ofSSEA-4 shows clear correlation with metasta-
possess the b-GalNAc-(1!3)-a-Gal-(1!4)-Gal oligo- sis potential and malignancy ofrenal cell carcino-
saccharide moiety.[1,2] This family of glycolipids has at- ma.[20–23]
tracted much attention due to their roles as tumor-as-
Due to their biological importance and the difficul-
sociated antigens[2] and as receptors for bacteria, vi- ties in accessing these complex oligosaccharides from
ruses and toxins, which present excellent targets for natural sources, great efforts have been devoted to
novel therapeutics design.[3–5] For example, Globo-H chemical synthesis.[1,24] Globo-H hexasaccharide has
has been found to be over-expressed on a variety of been assembled by a variety ofmethods including the
human cancer cells, including breast cancer, prostate glycal strategy,[25] the trichloroacetimidate method,[26]
cancer, ovarian cancer, and lung carcinomas.[6,7] Im- two-directional glycosylation,[27] automated solid-
munotherapy using Globo-H hexasaccharide as a phase synthesis,[28] the reactivity-based one-pot
cancer vaccine[8] has received encouraging preliminary method[29] as well as the pre-activation-based one-pot
results against breast cancer, which is now undergoing method.[30] With the presence ofthe sialic acid unit at
phase II clinical trials.[9–11] SSEA-4 is believed to be its non-reducing end, SSEA-4 presents additional syn-
involved in bacterial and viral infections, serving as thetic challenges. Although much progress has been
Adv. Synth. Catal. 2008, 350, 1717 – 1728
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
1717

DEDICATED CLUSTER
FULL PAPERS
Zhen Wang et al.
made in chemical sialylation,[31,32] it still remains one ically. Herein, we report our studies on chemoenzy-
of the most difficult glycosidic linkages to synthesize. matic syntheses ofGlobo-H 1b and SSEA-4 2b, both
The carboxylic acid moiety in sialic acid also requires ofwhich contain aminopropyl side chains and can be
additional consideration for protective group compati- conjugated to protein carriers for future immunologi-
bility in synthetic design.[33] To date, SSEA-4 has only cal studies.
been assembled twice by the Hasagawa[34] and
Schmidt/Garegg groups.[35] The successful completion
ofthese highly challenging structures serves as high-
lights ofthe power ofmodern carbohydrate synthetic
Results and Discussion
methodologies. However, despite these accomplish- Retrosynthetically, we envision that both Globo-H
ments, there is a continual need to further improve and SSEA-4 can be accessed by enzymatic glycosyla-
synthetic efficiencies of these molecules.
tion ofpentasaccharide SSEA-3 3, which is also a
An attractive alternative to chemical synthesis is tumor-associated carbohydrate antigen.[42] The penta-
enzymatic synthesis.[36,37] With their high regio- and saccharide 3, in turn, can be chemically assembled
stereoselectivities, glycosyltransferases can facilitate from disaccharide 5, galactoside 6, and lactoside 7
the formation of glycosidic bonds under aqueous con- (Figure 1). Based on our previous studies,[30] the pres-
ditions without relying heavily on protective groups. ence ofelectron-withdrawing benzoyl moieties on the
However, enzymatic activities can be highly depen- lactoside acceptor drastically reduced the glycosyla-
dent upon structures ofboth the glycosyl donor and
the acceptor. An effective enzymatic synthesis may are selected as protective groups masking hydroxy
require the preparation and screening ofan array of groups not undergoing glycosylation in order to en-
enzymes, which can be time-consuming. Thus, chemo- hance the nucleophilicity ofthe acceptors as well as
tion yield. Thus, the electron-donating benzyl groups
enzymatic synthesis, through the combination ofthe
power ofenzymatic synthesis with the lfexibility of
to simplify deprotection procedures. A side effect of
using benzyl groups is that building block galactoside
chemical synthesis, presents a potent approach to 6 possesses high anomeric reactivity (armed)[43] as a
access complex oligosaccharides. Several gangliosides glycosyl donor with its multiple electron-donating
such as GM3,[38] GM2,[39] GM1,[39] GD1a,[39] Gb3[40] protective groups.[36] Therefore, in chemoselective gly-
and SSEA-3[41] have been synthesized chemoenzymat- cosylation ofthiogalactoside 6 by the thioglycoyl
1718
asc.wiley-vch.de
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2008, 350, 1717 – 1728

DEDICATED CLUSTER
FULL PAPERS
Chemoenzymatic Syntheses ofTumor-Associated Carbohydrate Antigen Globo-H
Figure 1. Retrosynthetic analysis.
donor 5, caution must be taken to prevent the unde- dride (Tf2O),[55] S-(4-methoxyphenyl) benzenethiosul-
sired premature activation ofgalactoside 6. This can finate/Tf2O,[56] benzenesulfinylmorpholine/Tf2O,[57]
be achieved by pre-activating[44–48] disaccharide 5 first and diphenyl sulfoxide/Tf2O[58] have also been used as
with a stoichiometric promoter, generating a reactive thioglycoside promoters for pre-activation reactions.
intermediate. Upon complete donor activation, addi- However, electrophilic side products are typically
tion ofthe biufnctional galactoside 6 building block generated with these promoters, which often require
to the reaction mixture will produce the desired tri- the addition ofa quencher such as triethyl phosphite
saccharide. As donor activation and acceptor addition at the end ofthe reaction to prevent undesired ac-
occur as two distinct steps, even though the acceptor ceptor activation.[45,59] This renders it difficult to carry
6 has high anomeric reactivity, it will not be activated out multiple sequential glycosylations in the same re-
by the promoter.
action flask without intermediate purification.
For the pre-activation-based glycosylation ap-
Disaccharide donor 5 was prepared from disacchar-
proach, we have extensively used the p-TolSCl/ ide 8[30] starting with treatment ofsodium hydride and
[30,44,49–52]
AgOTfpromoter system,
which generates p- benzyl bromide. Reduction ofthe azide moiety and
TolSOTf in situ. p-TolSOTfis a powerufl thiophilic
subsequent Troc protection produced the disaccharide
agent, capable ofactivating thioglycosides even with
donor 5 in 81% overall yield for the three steps
very low anomeric reactivities.[48,53,54] Moreover, the (Scheme 1). Galactoside 6 and lactoside 7 were pre-
side product p-tolyl disulfide produced from pre-acti- pared according to literature methods.[30,60]
vation is not electrophilic, thus will not activate the
With all necessary building blocks in hand, we per-
thioglycoside acceptor. Alternatively, reagent combi- formed the assembly of SSEA-3 using the pre-activa-
nations such as benzenesulfinylpiperidine/triflic anhy- tion-based one-pot protocol.[30,44] With future automa-
Scheme 1.
Adv. Synth. Catal. 2008, 350, 1717 – 1728
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
1719

DEDICATED CLUSTER
FULL PAPERS
Zhen Wang et al.
Scheme 2.
tion in mind, we decided to carry out the one-pot syn- often obtained.[28,60,64] We have reported that glycosy-
thesis under the reaction conditions established previ- lation ofthe lactoside acceptor 7 by the galactoside
ously without optimization. Pre-activation ofthe dis-
9[30] gave the Gb3 trisaccharide 10 in a combined 85%
[30]
accharide donor 5 at ꢀ788C with p-TolSCl/AgOTf yield with an a:b ratio of27:1 (Table 1, entry 1),
was followed by the addition of the bifunctional which compared favorably with literature results on
building block 6 (Scheme 2, reaction a). A sterically Gb3 synthesis.[28,60,64] The large difference in stereo-
hindered base, 2,4,6-tri-tert-butylpyrimidine (TTBP)[61] chemical selectivity between Gb3 trisaccharide 10 and
was added with 6 to neutralize triflic acid generated
from glycosylation. The reaction temperature was
Table 1. Evaluation ofthe ofrmation ofthe
Gal linkage.
a-Gal-(1!4)-
raised to ꢀ208C to expedite glycosylation, and the ac-
ceptor 6 was completely consumed as judged by TLC
analysis. The reaction temperature was cooled back
down to ꢀ788C, followed by addition of the lactoside
acceptor 7 and p-TolSCl/AgOTf. The fully protected
SSEA-3 pentasaccharide 4a was obtained in 37%
yield from this three-component one-pot reaction se-
quence within six hours, which was fully characterized
1
by H NMR, 13C NMR, gCOSY, gHMQC and HR-
MS. In addition, the anomer 4b was also separated in
13% yield. The presence or absence ofthe base
TTBP did not significantly affect either the yield or
[62]
the stereochemical selectivity ofthis reaction.
Deprotection ofthe pentasaccharide 4a was per-
formed by first removing the Troc group with 1M
NaOH in THF followed by acetylation. Staudinger re-
duction ofthe azide group and subsequent catalytic
hydrogenation over Pearlmanꢁs catalyst[30,50,51,63] gave
the fully deprotected SSEA-3 3 in 55% overall yield
for all the deprotection steps (Scheme 2, reaction b).
Formation of the Challenging a-Gal-(1!4)-Gal
Linkage, Stereochemical Dependence on Donor
We examined next the stereochemical outcome in for-
mation ofthe a-Gal-(1!4)-Gal linkage, which is a
major challenge in the syntheses ofall globo series of
gangliosides, with anomeric mixtures ofproducts
1720
asc.wiley-vch.de
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2008, 350, 1717 – 1728

DEDICATED CLUSTER
FULL PAPERS
Chemoenzymatic Syntheses ofTumor-Associated Carbohydrate Antigen Globo-H
Scheme 3.
SSEA-3 pentasaccharide 4 prompted us to prepare that as the glycosyl donor became larger, the a selec-
the disaccharide donor 11, the trisaccharide donor 12 tivity became lower although the overall yields were
and the tetrasaccharide donor 13 (Scheme 3). Pre-ac- similar. The stereoselectivity change must be reflec-
tivation ofdonor 14[30] by p-TolSCl/AgOTfofllowed
tive of the rate differential of axial versus equatorial
by addition ofthe galactoside 6 generated the disac- approach ofthe nucleophile onto the reactive inter-
charide 11 in 70% yield and the glycosylation of 6 by mediate(s),[51,65] because no anomeric bond isomeriza-
5 produced the trisaccharide 12 (Scheme 3, reactions tions were observed when the O-glycoside products
a and b). The successful glycosylation of armed galac- were re-submitted to the glycosylation reaction condi-
toside 6 by the disarmed donor 14 is a salient feature tions.
ofthe pre-activation method. One-pot sequential gly-
cosylation ofufcoside
15,[30] disaccharide 16[30] and glycosylating the acceptor 7 may be attributed to
galactoside 6 led to the tetrasaccharide donor 13 in changes in electronic properties and/or steric encum-
60% overall yield (Scheme 3, reaction c). brance ofthe glycosyl donors. The introduction ofa
It is possible that the stereochemical divergence in
The glycosylations oflactoside 7 by donors 11, 12 single electron-withdrawing group on glycosyl donors
and 13 were then performed. The reaction of disac- can greatly influence the stereoselectivity.[66,67] Induc-
charide donor 11 with 7 gave tetrasaccharides 17a tively, the glycosyl units on 3-O ofthe reducing end
(71%) and 17b (19%) (17a:17b=3.7:1, Table 1, galactose in donors 11–13 can be viewed as electron-
entry 2). Trisaccharide 12 glycosylated the lactoside 7 withdrawing groups.[51,68] Therefore we prepared
with an a:b ratio of2.6 (Table 1, entry 3), which was
donor 19 bearing a highly electron-withdrawing p-ni-
similar to that obtained in the one-pot synthesis of trobenzoyl (p-NO2Bz) moiety on its 3-O position
SSEA-3 (Scheme 2) indicating that the one-pot proce- through nitrobenzoylation ofgalactoside 6. Glycosyla-
dure did not significantly affect the stereoselectivity. tion ofthe lactoside 7 by the donor 19 produced the
A further reduction in a:b selectivity (1.9:1) was ob- trisaccharide 20a in 78% yield with no corresponding
served in glycosylation of 7 by the tetrasaccharide b anomer isolated (Table 1, entry 5). This suggests
donor 13 (Table 1, entry 4). These results suggested that electronic properties ofthe donors do not play a
Adv. Synth. Catal. 2008, 350, 1717 – 1728
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
1721

DEDICATED CLUSTER
FULL PAPERS
Zhen Wang et al.
major role in determining stereoselectivity in this Enzymatic Synthesis of Globo-H 1a and MSGG 2a
case.
Crich and co-workers have reported that the pres- Fucosylation ofthe SSEA-3 pentasaccharide 3 was
ence ofa tert-butyldimethylsilyl (TBDMS) group on carried out using WbsJ[71] with GDP-fucose. Despite
3-O ofthe mannosyl donor 21 led to a significant de- the presence ofthree galactosyl units in SSEA-3, the
crease in b-mannoside formation compared to the 2-OH ofthe non-reducing terminal galactose was se-
corresponding reaction with the benzyl-bearing donor lectively fucosylated leading to Globo-H 1a in 71%
22.[69] This was rationalized by a steric buttressing yield (Scheme 4, reaction a), the identity ofwhich was
confirmed by NMR comparison with the Globo-H 1a
assembled via chemical synthesis.[30]
Sialylation ofpentasaccharide 3 was performed
with the Campylobacter jejuni Cst-I sialyltransferase
(construct CST-06), which cleanly transformed SSEA-
3 into SSEA-4 2b with 1.5 equivalents ofCMP-
Neu5Ac (Scheme 4, reaction a). The conversion was
quantitative based on TLC but the isolated yield was
effect as the bulky 3-O-TBDMS moiety pushes the 34%. Presumably, some product was lost during the
axial 2-O-benzyl group towards the b face of the work-up and purification processes. SSEA-3 can also
anomeric center, 1,2-cis-(b-mannosyl) glycosylation is be sialylated by a multifunctional Pasteurella multoci-
hindered. Similar phenomena have been observed in da sialyltransferase PmST1[72] to form SSEA-4 2b in
modular synthesis ofalternating b-(1!3)-b-(1!4)- over 90% yield based on TLC analysis. In addition,
mannans,[70] where increased steric size on the 3-O po- we found that SSEA-3 is a substrate for a recombi-
sition ofthe donor in the ofrm ofanother glycosyl
nant Photobacterium damsela a-2,6-sialyltransferase
unit reduced the amount of b-mannoside product Pd2,6ST[73] with a yield of40% by TLC. These results
drastically. It is possible in our studies that the glyco- indicate that the large size ofthe pentasaccharide ac-
syl units on the 3-O ofreducing end galactose of
ceptor 3 does not present an obstacle to these enzy-
donors 11, 12 and 13 present a steric buttressing matic glycosylation reactions.
effect, pushing the equatorial 2-O-benzyl towards the
As we have obtained the tetrasaccharide 17a, we
anomeric center, thus reducing the amount of1,2- cis- investigated whether its fully deprotected form can
(a-galactosyl) products. As the size ofthe donor in-
also serve as an enzyme substrate. Compound 17a
creases from monosaccharide 9 to tetrasaccharide 13, was deprotected in the same manner as described for
the effect of the steric buttressing effect becomes the pentasaccharide 3, producing the tetrasaccharide
greater resulting in lower a selectivity.
23 in 50% yield (Scheme 4, reaction b). Tetrasacchar-
Scheme 4.
1722
asc.wiley-vch.de
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2008, 350, 1717 – 1728

DEDICATED CLUSTER
FULL PAPERS
Chemoenzymatic Syntheses ofTumor-Associated Carbohydrate Antigen Globo-H
All CH2Cl2 solutions were combined and washed twice with
ide 23 can serve as a substrate to galactosyl transfer-
ase LgtD,[41,74] providing an alternative access to
SSEA-3.
a saturated aqueous solution ofNaHCO (20 mL) and twice
3
with water (10 mL). The organic layer was collected and
dried over Na2SO4. After removal of the solvent, the desired
disaccharide was purified from the reaction mixture via
silica gel flash chromatography.
Conclusions
p-Tolyl 2,3,4,6-Tetra-O-benzyl-b-d-galactopyranosyl-
(1!3)-4,6-benzylidene-2-(2,2,2-trichloroethoxycar-
bonylamino)-2-deoxy-1-thio-b-d-galactopyranoside
(5)
In conclusion, we report the first chemoenzymatic
syntheses oftwo tumor-associated carbohydrate anti-
gens, Globo-H hexasaccharide 1b and SSEA-4 hexa-
saccharide 2b. The common precursor SSEA-3 penta-
saccharide for these two compounds was assembled
rapidly using the pre-activation-based one-pot
method. Enzymatic glycosylations ofSSEA-3 by fuco-
syltransferase WbsJ and sialyltransferases CST-I or
PmST1 led to Globo-H and SSEA-4, respectively. In-
terestingly, we observed that the stereoselectivities in
formation of the challenging a-Gal-(1!4)-Gal link-
age decreased when larger oligosaccharide donors
were used. This was rationalized by a steric buttress-
ing effect due to the glycosyl unit(s) linked to 3-O of
the reducing end galactose in the donor.
Galactoside 8[30] (0.4 g, 0.48 mmol) was dissolved in DMF
(10 mL) and the solution was cooled to 08C. NaH (0.029 g,
60% NaH in mineral oil, 0.72 mmol) was added in portions,
followed by addition of benzyl bromide (0.086 mL,
0.72 mmol). The mixture was stirred at room temperature
under N2 for 2 h and then diluted with EtOAc (50 mL). The
mixture was washed with saturated NaHCO3, water and
then dried over Na2SO4, filtered and concentrated. Silica gel
column chromatography (2:1 hexanes-EtOAc) afforded p-
tolyl 2,3,4,6-tetra-O-benzyl-b-d-galactopyranosyl-(1!3)-2-
azido-4,6-benzylidene-2-deoxy-1-thio-b-d-galactopyranoside
S1 as white solid; yield: 0.44 g (quantitative).
The above prepared S1 (0.44 g, 0.48 mmol), 1,3-propane-
dithiol (0.48 mL, 4.8 mmol) and Et3N (0.67 mL, 4.8 mmol)
were dissolved in a mixture ofCH Cl2/MeOH (5 mL each).
Experimental Section
2
The mixture was heated at reflux overnight under N2 and
then concentrated. The resulting residue was diluted with
CH2Cl2 (60 mL) and then washed with a saturated aqueous
Characterization of Anomeric Stereochemistry
The stereochemistries ofthe newly ofrmed glycosidic link-
ages in oligosaccharides are determined by JH1,H2 through
solution ofNaHCO and water, dried over Na2SO4, filtered
3
3
and concentrated. The resulting residue was purified by
quickly passing it through a short silica gel column (20:1,
CH2Cl2-MeOH) and the obtained solid and solid NaHCO3
(0.078 g, 0.93 mmol) were put into THF (5 mL) and then
TrocCl (0.075 mL, 0.56 mmol) was added. The mixture was
stirred at room temperature under N2 for 4 h and filtered.
The filtrate was concentrated and then diluted with CH2Cl2
(50 mL). The mixture was washed with water and brine,
dried over Na2SO4, filtered and concentrated. Silica gel
column chromatography (2:1 Hexanes-EtOAc) afforded
compound 5 as white solid; yield: 0.42 g (81% for two
steps).
3
(around 3 Hz) indicate 1,2-cis a linkages and larger coupling
3
constants JH1,H2 (7.2 Hz or larger) indicate 1,2-trans b link-
1
ages. This can be further confirmed by JC1,H1 (~170 Hz) for
1
a linkages and JC1,H1 (~160 Hz) for b linkages.[75]
General Procedure for Single Step Pre-Activation-
Based Glycosylation
A solution ofdonor (0.060 mmol) and rfeshly activated mo-
lecular sieve MS 4 ꢂ (200 mg) in CH2Cl2 (2 mL) was stirred
at room temperature for 30 min, and cooled to ꢀ788C,
which was followed by addition of AgOTf (47 mg,
0.18 mmol) dissolved in Et2O (1 mL) without touching the
wall of the flask. After 5 min, orange colored p-TolSCl
(9.5 mL, 0.060 mmol) was added through a microsyringe.
Since the reaction temperature was lower than the freezing
point of p-TolSCl, p-TolSCl was added directly into the reac-
tion mixture to prevent it from freezing on the flask wall.
The characteristic yellow color of p-TolSCl in the reaction
solution dissipated rapidly within a few seconds indicating
depletion of p-TolSCl. After the donor was completely con-
sumed according to TLC analysis (about 5 min at ꢀ788C), a
solution ofacceptor (0.060 mmol) in CH 2Cl2 (0.2 mL) was
slowly added dropwise via a syringe. The reaction mixture
was warmed to ꢀ208C under stirring in 2 h. Then the mix-
ture was diluted with CH2Cl2 (20 mL) and filtered over
Celite. The Celite was further washed with CH2Cl2 until no
organic compounds were observed in the filtrate by TLC.
3-Azidopropyl 2,3,4,6-Tetra-O-benzyl-b-d-galactopy-
ranosyl-(1!3)-4,6-benzylidene-2-(2,2,2-trichloroeth-
oxycarbonylamino)-2-deoxy-b-d-galactopyranosyl-
(1!3)-2,4,6-tri-O-benzyl-a-d-galactopyranosyl-(1!4)-
2,3,6-tri-O-benzyl-b-d-galactopyranosyl-(1!4)-2,3,6-
tri-O-benzyl-b-d-glucopyranoside (4a) and 3-
Azidopropyl 2,3,4,6-Tetra-O-benzyl-b-d-galacto-
pyranosyl-(1!3)-4,6-benzylidene-2-(2,2,2-trichloro-
ethoxycarbonylamino)-2-deoxy-b-d-galacto-pyranosyl-
(1!3)-2,4,6-tri-O-benzyl-b-d-galacto-pyranosyl-(1!4)-
2,3,6-tri-O-benzyl-b-d-galacto-pyranosyl-(1!4)-2,3,6-
tri-O-benzyl-b-d-glucopyranoside (4b)
After donor 5 (50 mg, 46.7 mmol) and activated molecular
sieve MS-4 ꢂ (500 mg) had been stirred for 30 min at room
temperature in CH2Cl2 (3 mL), the solution was cooled to
ꢀ788C, followed by addition of AgOTf (36 mg, 140 mmol) in
Adv. Synth. Catal. 2008, 350, 1717 – 1728
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
1723

DEDICATED CLUSTER
FULL PAPERS
Zhen Wang et al.
Et2O (1.5 mL). The mixture was stirred for 5 min at ꢀ788C
and then p-TolSCl (7.4 mL, 46.7 mmol) was added into the
solution. (See the general procedure for single step pre-acti-
vation based glycosylation for precautions) The mixture was
vigorously stirred for 10 min, followed by addition of a solu-
tion ofacceptor 6 (22.1 mg, 39.7 mmol) and TTBP (6.9 mg,
46.7 mmol) in CH2Cl2 (1 mL). The reaction mixture was
stirred for 2 h from ꢀ78 to ꢀ208C and then the mixture was
cooled down to ꢀ788C, followed by sequential additions of
AgOTf(12 mg, 46.7 mmol) in Et2O (1 mL), acceptor 7
(22.5 mg, 23.3 mmol) and TTBP (6.9 mg, 46.7 mmol) in
CH2Cl2 (1 mL). The mixture was stirred for 5 min at ꢀ788C
and then p-TolSCl (6.3 mL, 39.7 mmol) was added into the
solution. The reaction mixture was stirred for 3 h from ꢀ78
to 108C and then was quenched with Et3N (40 mL), concen-
trated under vacuum to dryness. The resulting residue was
diluted with CH2Cl2 (30 mL), followed by filtration. The or-
ganic phase was washed with saturated aqueous NaHCO3,
H2O and then dried over Na2SO4, filtered and concentrated.
Silica gel column chromatography (2:1 hexanes-EtOAc) af-
forded 4a (yield: 20 mg, 37%) and 4b (yield: 7.2 mg, 13%),
respectively, as colorless gel.
3) and then the aqueous phase was dried under vacuum to
afford compound 3 (acetate salt) as a white solid; yield:
13 mg (55% for four steps).
p-Tolyl 3,4,6-tri-O-Acetyl-2-(2,2,2-trichloroethoxycar-
bonylamino)-2-deoxy-b-d-galactopyranosyl-(1!3)-
2,4,6-tri-O-benzyl-1-thio-b-d-galactopyranoside (11)
Compound 11 was synthesized from donor 14 and acceptor
6 in 70% yield following the general procedure of glycosyla-
tion.
p-Tolyl 2,3,4,6-Tetra-O-benzyl-b-d-galactopyranosyl-
(1!3)-4,6-benzylidene-2-(2,2,2-trichloroethoxy-
carbonylamino)-2-deoxy-b-d-galactopyranosyl-(1!3)-
2,4,6-tri-O-benzyl-1-thio-b-d-galactopyranoside (12)
Compound 12 was synthesized from donor 5 and acceptor 6
in 71% yield following the general procedure of single step
glycosylation.
3-Azidopropyl 2,3,4-Tri-O-benzyl-a-l-fucopyranosyl-
(1!2)-3,4,6-tri-O-benzyl-b-d-galactopyranosyl-(1!3)-
4,6-benzylidene-2-(2,2,2-trichloroethoxycarbonyl-
amino)-2-deoxy-b-d-galactopyranosyl-(1!3)-2,4,6-tri-
O-benzyl-1-thio-b-d-galactopyranoside (13)
Compounds 4a and 4b were also synthesized from donor
12 and acceptor 7 in 60% and 23% yield, respectively, as
colorless gel following the general procedure of single step
glycosylation.
Compound 13 was synthesized by a three-component one-
pot synthesis procedure. After the donor p-tolyl 2,3,4-tri-O-
benzyl-1-thio-b-l-fucopyranoside 15[30] (50 mg, 92.47 mmol)
and activated molecular sieve MS-4 ꢂ (500 mg) had been
stirred for 30 min at room temperature in Et2O (4 mL), the
solution was cooled to ꢀ788C, followed by addition of
AgOTf(72 mg, 277.4 mmol) in Et2O (1.5 mL). The mixture
was stirred for 5 min at ꢀ788C and then p-TolSCl (14.7 mL,
92.47 mmol) was added into the solution. (See the general
procedure for single step pre-activation based glycosylation
for precautions) The mixture was vigorously stirred for
10 min, followed by addition of a solution of acceptor p-
tolyl 3,4,6-tri-O-benzyl-b-d-galactopyranosyl-(1!3)-4,6-ben-
zylidene-2-(2,2,2-trichloroethoxycarbonylamino)-2-deoxy-1-
thio-b-d-galactopyranoside 16[30] (77.1 mg, 78.60 mmol) and
TTBP (23 mg, 92.47 mmol) in CH2Cl2 (1 mL). The reaction
mixture was stirred for 2 h from ꢀ78 to ꢀ208C and then the
mixture was cooled down to ꢀ788C, followed by addition of
AgOTf(24 mg, 92.47 mmol) in Et2O (1 mL). The mixture
was stirred for 10 min at ꢀ788C and then p-TolSCl (12.5 mL,
78.60 mmol) was added into the solution. After stirred for
5 min, a solution ofacceptor 6 (30.9 mg, 55.48 mmol) and
TTBP (23 mg, 92.47 mmol) in CH2Cl2 (1 mL) was added
slowly along the flask wall into the mixture and the reaction
mixture was stirred for 2 h from ꢀ78 to ꢀ208C, then reac-
tion was quenched with Et3N (40 mL) and concentrated
under vacuum to dryness. The resulting residue was diluted
with CH2Cl2 (30 mL), followed by filtration. The organic
phase was washed with saturated aqueous NaHCO3, H2O
and then dried over Na2SO4, filtered and concentrated.
Silica gel column chromatography (2:1 hexanes-EtOAc) af-
forded 13 as a colorless gel; yield: 60.6 mg (60%).
3-Aminopropyl b-d-Galactopyranosyl-(1!3)-2-acet-
amido-2-deoxy-b-d-galactopyranosyl-(1!3)-a-d-ga-
lactopyranosyl-(1!4)-b-d-galactopyranosyl-(1!4)-b-
d-glucopyranoside (3)
The mixture ofcompound 4a (60 mg, 25.5 mmol), 1M
NaOH (4 mL, 0.4 mmol) and THF (4 mL) was stirred at
508C overnight and then concentrated to dryness. The re-
sulting residue was diluted with CH2Cl2 (30 mL) and the or-
ganic phase was washed by H2O and then dried over
Na2SO4, filtered and concentrated to dryness. The resulting
residue was dissolved in methanol (2 mL) and acetic anhy-
dride (24 mL, 0.25 mmol) was added dropwise and the mix-
ture was stirred at room temperature under N2 overnight.
The reaction was quenched by adding a few drops of H2O
and then diluted with CH2Cl2 (30 mL). The organic phase
was washed with a saturated aqueous solution ofNaHCO ,
3
H2O and then dried over Na2SO4, filtered and concentrated
to dryness. Silica gel column chromatography (2:1 hexanes-
EtOAc) afforded the N-acetylation product as white solid.
The mixture ofthe N-acetylation product, 1M PMe3 in
THF (0.5 mL, 0.5 mmol), 0.1M NaOH (0.5 mL, 0.05 mmol)
and THF (4 mL) was stirred at 608C under N2 overnight.
The mixture was concentrated and the resulting residue was
diluted with CH2Cl2 (30 mL). The organic phase was washed
with H2O and then dried over Na2SO4, filtered and concen-
trated to dryness. The resulting residue was purified by
quickly passing through a short silica gel column (10:1,
CH2Cl2-MeOH). The mixture ofthe obtained solid and
Pd(OH)2 in MeOH/H2O/HOAc (3 mL/1 mL/1 mL) was
stirred under H2 at room temperature overnight and then
filtered. The filtrate was concentrated to dryness under
vacuum and then was co-evaporated with H2O (10 mL)
three times to remove the HOAc. The aqueous phase was
further washed with CH2Cl2 (5 mLꢃ3) and EtOAc (5 mLꢃ
1724
asc.wiley-vch.de
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2008, 350, 1717 – 1728

DEDICATED CLUSTER
FULL PAPERS
Chemoenzymatic Syntheses ofTumor-Associated Carbohydrate Antigen Globo-H
0.127 mmol) in Et2O (1 mL). The mixture was vigorously
stirred for 10 min and then p-TolSCl (6.74 mL, 42 mmol) was
added and the reaction mixture was stirred for 2 h from ꢀ78
to ꢀ208C. The reaction was quenched by Et3N and then
concentrated under vacuum to dryness. The resulting resi-
due was diluted with CH2Cl2 (10 mL), followed by filtration.
The organic phase was washed with saturated aqueous
NaHCO3 and H2O and then dried over Na2SO4, filtered and
concentrated. Silica gel column chromatography (3:1 hex-
anes-EtOAc) afforded 20a as gel-like solid; yield: 46.2 mg
(78%).
3-Azidopropyl 3,4,6-Tri-O-acetyl-2-(2,2,2-trichloro-
ethoxycarbonylamino)-2-deoxy-b-d-galactopyranosyl-
(1!3)-2,4,6-tri-O-benzyl-a-d-galactopyranosyl-(1!4)-
2,3,6-tri-O-benzyl-b-d-galactopyranosyl-(1!3)-2,4,6-
tri-O-benzyl-b-d-glucopyranoside (17a) and 3-
Azidopropyl 3,4,6-Tri-O-acetyl-2-(2,2,2-tri-
chloroethoxycarbonylamino)-2-deoxy-b-d-
galactopyranosyl-(1!3)-2,4,6-tri-O-benzyl-b-d-
galactopyranosyl-(1!4)-2,3,6-tri-O-benzyl-b-d-
galactopyranosyl-(1!3)-2,4,6-tri-O-benzyl-b-d-
glucopyranoside (17b)
3-Aminopropyl a-l-Fucopyranosyl-(1!2)-b-d-
galactopyranosyl-(1!3)-2-acetamido-2-deoxy-b-d-
galac-topyranosyl-(1!3)-a-d-galactopyranosyl-(1!4)-
b-d-galactopyranosyl-(1!4)-b-d-glucopyranoside (1b)
Compound 17a and 17b were synthesized from donor 11
and acceptor 7 in 71% and 19% yield, respectively, as color-
less gel following the general procedure of single-step glyco-
sylation.
To a mixture of10 mM SSEA-3 pentasaccharide 3, 15 mM
GDP-fucose in 20 mM Tris-HCl buffer (pH 7.5) was added
a-1,2-fucosyltransferase WbsJ (7 mU). The mixture was in-
cubated at room temperature for 2 days. The Globo-H hexa-
saccharide 1b was purified on a BioGel P-2 gel filteration
column (BioRad) followed by Dowex 1ꢃ8–400 ion ex-
change resin (chloride form) with water as the mobile
phase. The fractions containing the desired product 1b
(71%) were collected and lyophilized. NMR data of 1b were
collected, which were identical to those obtained from
Globo-H 1b synthesized chemically.[30]
3-Azidopropyl 2,3,4-Tri-O-benzyl-a-l-fucopyranosyl-
(1!2)-3,4,6-tri-O-benzyl-b-d-galactopyranosyl-(1!3)-
4,6-benzylidene-2-(2,2,2-trichloroethoxycarbonyl-
amino)-2-deoxy-b-d-galactopyranosyl-(1!3)-2,4,6-tri-
O-benzyl-a-d-galactopyranosyl-(1!4)-2,3,6-tri-O-
benzyl-b-d-galactopyranosyl-(1!4)-2,3,6-tri-O-
benzyl-b-d-glucopyranoside (18a) and 3-Azidopropyl
2,3,4-Tri-O-benzyl-a-l-fucopyranosyl-(1!2)-3,4,6-tri-
O-benzyl-b-d-galactopyranosyl-(1!3)-4,6-
benzylidene-2-(2,2,2-tri-chloroethoxycarbonylamino)-
2-deoxy-b-d-galactopyranosyl-(1!3)-2,4,6-tri-O-
benzyl-b-d-galactopyranosyl-(1!4)-2,3,6-tri-O-
benzyl-b-d-galactopyranosyl-(1!4)-2,3,6-tri-O-
benzyl-b-d-glucopyranoside (18b)
3-Aminopropyl 5-Acetamido-3,5-dideoxy-d-glycero-a-
d-galacto-non-2-ulopyranosylonic acid-(2!3)-b-d-
galactopyranosyl-(1!3)-2-acetamido-2-deoxy-b-d-
galactopyranosyl-(1!3)-a-d-galactopyranosyl-(1!4)-
b-d-galactopyranosyl-(1!4)-b-d-glucopyranoside (2b)
The Campylobacter jejuni CST-I a-2,3-sialyltransferase (con-
struct CST-06) was expressed as a fusion protein with the E.
coli maltose-binding protein and purified.[39] To a mixture of
10 mM MgCl2, 5 mM SSEA-3 pentasaccharide 3, 7.5 mM
CMP-Neu5Ac in Hepes buffer (50 mM, pH 7.5) was added
CST-I a-2,3-sialyltransferase (0.32 U/mL). The reaction was
incubated at 378C for 2 h and TLC (BPH:AMW=1:1, BPH
Compound 18a and 18b were synthesized from donor 13
and acceptor 7 in 57% and 30% yield, respectively, as color-
less gel following the general procedure of single step glyco-
sylation. Their identities were confirmed by NMR compari-
son with literature data.[30]
p-Tolyl 2,4,6-Tri-O-benzyl-3-O-p-nitrobenzoyl-1-thio-
b-d-galactopyranoside (19)
1:2:1
n-butanol:propanol:0.1M
HCl;
AMW
1:1:1
Compound 6 (43.5 mg, 0.078 mmol) and N,N-dimethylami-
nopyridine (9.5 mg, 0.078 mmol) were dissolved in CH2Cl2
(5 mL), followed by the addition of p-nitrobenzoyl chloride
(10.9 mL, 0.09 mmol). The mixture was stirred overnight at
room temperature and then diluted with CH2Cl2 (20 mL),
washed with saturated aqueous NaHCO3 and H2O and then
dried over Na2SO4, filtered and concentrated. Silica gel
column chromatography (4:1 hexanes-EtOAc) afforded 19
as a gel-like solid; yield: 50.6 mg (92%).
CH3CN:methanol:water) showed complete consumption of
the pentasaccharide 3. Multiple reactions were set to con-
vert all ofthe pentasaccharide 3 (9 mg, 9.1 mmol). The reac-
tion volume was reduced using a centrifugal concentrator
and the material was loaded (in 3 runs) on a 10 mmꢃ30 cm
Superdex Peptide column (GE Healthcare). The column
was eluted with an ammonium acetate buffer (pH 7,
20 mM) at 0.5 mLminꢀ1. The product was eluted between 7
and 8 mL. The fractions were pooled and lyophilized three
times to remove the ammonium acetate. We recovered 4 mg
(3.1 mmol) ofpuriifed 2b with a yield of34%. As the condi-
tions for enzymatic synthesis were very mild and previously
we did not observe any product decomposition using this
enzyme,[39] the low isolated yield was probably due to multi-
ple small-scale reactions performed resulting in loss of prod-
uct during work-up and purification.
3-Azidopropyl 2,4,6-Tri-O-benzyl-3-O-p-nitrobenzoyl-
a-d-galactopyranosyl-(1!4)-2,3,6-tri-O-benzyl-b-d-
galactopyranosyl-(1!4)-2,3,6-tri-O-benzyl-b-d-
glucopyranoside (20a)
After donor 19 (30 mg, 42 mmol), acceptor 7 (37 mg, 38
mmol) and activated molecular sieve MS-4 ꢂ (200 mg) were
stirred for 30 min at room temperature in a mixture solvent
ofEt O (1 mL) and CH2Cl2 (2 mL), the mixture was cooled
2
to ꢀ788C, followed by addition of AgOTf (33 mg,
Adv. Synth. Catal. 2008, 350, 1717 – 1728
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
1725

DEDICATED CLUSTER
FULL PAPERS
Zhen Wang et al.
Enzymatic Sialylation of Pentasaccharide SSEA-3
using PmST1 and Pd2,6ST
Supporting Information
General experimental procedures as well as analytical and
spectroscopic characterization data for all compounds are
given in the Supporting information.
The enzymatic assays were carried out in a total volume of
10 mL in a Tris-HCl buffer (100 mm, pH 8.5) containing
CMP-Neu5Ac (20 mM), SSEA-3
3
(10 mM), MgCl2
(20 mM) and the corresponding sialyltransferases (PmST1
or Pd2,6ST). Reactions were allowed to proceed for 60 min
at 378C. The reactions were then monitored by TLC (BPH:
AMW=1:1) and the products formed were confirmed by
MS. ESI-MS (for sialylation product catalyzed by PmST1):
m/z=1262.29 calcd. for [M+Na]+ C46H78N3Na2O34: 1262.43;
ESI-MS (for sialylation product catalyzed by Pd2,6ST):
m/z=1262.28, calcd. for [M+Na]+ C46H78N3Na2O34: 1262.43.
Acknowledgements
We are grateful for financial supports from the National Insti-
tutes of Health (R01-GM-72667 to XH, R01-GM-76360 to
XC). We would like to thank Dr. Warren Wakarchuk (Na-
tional Research Council Canada) for helpful suggestions, Dr.
Jianjun Li (National Research Council Canada) for mass
spectrometry analysis of the sialylated product and Marie-
France Karwaski (National Research Council Canada) for
technical help.
3-Aminopropyl 2-Acetamido-2-deoxy-b-d-galactop-
yranosyl-(1!3)-a-d-galactopyranosyl-(1!4)-b-d-
galactopyranosyl-(1!4)-b-d-glucopyranoside (23)
The mixture ofcompound 17a (0.050 g, 0.027 mmol), 1M
NaOMe (4.0 mL, 4.0 mmol) and THF (4 mL) was stirred at
508C overnight and then concentrated to dryness. The re-
sulting residue was diluted with CH2Cl2 (50 mL) and the or-
ganic phase was washed by H2O and then dried over
Na2SO4, filtered and concentrated to dryness. The resulting
residue was dissolved in methanol (3 mL). Acetic anhydride
(1.0 mL) was added dropwise and the mixture was stirred at
room temperature under N2 for 6 h. The reaction was
quenched by adding EtOH and then diluted with EtOAc
(20 mL). The organic phase was washed with saturated
aqueous solution ofNaHCO 3, H2O and then dried over
Na2SO4, filtered and concentrated to dryness. Silica gel
column chromatography (2:1 hexanes-EtOAc) afforded the
N-acetylation product as white solid.
References
[1] H. Ishida, M. Kiso, Trends Glycosci. Glycotechnol.
2001, 13, 57–64.
[2] S. Hakomori, Y. Zhang, Chem. Biol. 1997, 4, 97–104.
[3] A. Wedeking, G. van Echten-Deckert, Curr. Org.
Chem. 2007, 11, 579–598.
[4] M. Izumi, H. Uzawa, Trends Glycosci. Glycotechnol.
2005, 17, 107–119.
[5] S. Birkle, G. Zeng, L. Gao, R. K. Yu, J. Aubry, Bio-
chimie 2003, 85, 455–463.
[6] S. Zhang, H. S. Zhang, V. E. Reuter, S. F. Slovin, H. I.
Scher, P. O. Livingston, Clin. Cancer Res. 1998, 4, 295–
302.
The mixture ofthe N-acetylation product, 1M PMe3 in
THF (0.5 mL, 0.5 mmol) and THF (3 mL) was stirred at
508C under N2 overnight. The mixture was concentrated
and the resulting residue was diluted with CH2Cl2 (50 mL).
The organic phase was washed with H2O and then dried
over Na2SO4, filtered and concentrated to dryness. The re-
sulting residue was purified by quickly passing through a
short silica gel column (10:1, CH2Cl2-MeOH). The mixture
[7] E. G. Bremer, S. B. Levery, S. Sonnino, R. Ghidoni, S.
Canevari, R. Kannagi, S. Hakomori, J. Biol. Chem.
1984, 259, 14773–14777.
[8] S. J. Danishefsky, J. R. Allen, Angew. Chem. 2000, 112,
882–912; Angew. Chem. Int. Ed. 2000, 39, 836–863.
[9] T. Gilewske, G. Ragupathi, S. Bhuta, L. J. Williams, C.
Musselli, X.-F. Zhang, K. P. Bencsath, K. S. Panageas, J.
Chin, C. A. Hudis, L. Norton, A. N. Houghton, P. O.
Livingston, S. J. Danishefsky, Proc. Natl. Acad. Sci.
USA 2001, 98, 3270–3275.
[10] Z. G. Wang, L. J. Williams, X. F. Zhang, A. Zatorski, V.
Kudrya-shov, G. Ragupathi, M. Spassova, W. Born-
mann, S. F. Slovin, H. I. Scher, P. O. Livingston, K. O.
Lloyd, S. J. Danishefsky, Proc. Natl. Acad. Sci. USA
2000, 97, 2719.
[11] S. F. Slovin, G. Ragupathi, S. Adluri, G. Ungers, K.
Terry, S. Kim, M. Spassova, W. G. Bornmann, M. Faz-
zari, L. Dantis, K. Olkiewicz, K. O. Lloyd, P. O. Living-
ston, S. J. Danishefsky, H. I. Scher, Proc. Natl. Acad.
Sci. USA 1999, 96, 5710.
ofthe obtained solid and Pd(OH) in MeOH/H2O/HOAc
2
(5 mL, 3:1:1) was stirred under H2 at room temperature
overnight and then filtered. The filtrate was concentrated to
dryness under vacuum and then was co-evaporated with
H2O (10 mL) three times to remove the HOAc. The aque-
ous phase was further washed with CH2Cl2 (5 mL ꢃ 3) and
EtOAc (5 mL ꢃ 3) and then the aqueous phase was dried
under vacuum to afford compound 23 as white solid; yield:
10.4 mg (50% for three steps).
Enzymatic Galactosylation of Tetrasaccharide 23
To a mixture of10 mM tetrasaccharide 23, 20 mM UDP-gal-
actose, 1 mM MnCl2, 1 mM dithiothreitol and 1% BSA in
20 mM Tris-HCl buffer (pH 7.5) was added b-1,3-galactosyl-
transferase (LgtD) (2 mU). The mixture was incubated at
room temperature for 2 days. The SSEA-3 pentasaccharide
3 (70%) was purified on a BioGel P-2 gel filteration column
(BioRad) followed by Dowex 1ꢃ8–400 ion exchange resin
(chloride form) with water as the mobile phase.
[12] A. E. Stapleton, M. R. Stroud, S. I. Hakomori, W. E.
Stamm, Infect. Immun. 1998, 66, 3856–3861.
[13] M. R. Stroud, A. E. Stapleton, S. B. Levery, Biochem-
istry 1998, 37, 17420–17428.
[14] A. Stapleton, E. Nudelman, H. Clausen, S. Hakomori,
W. E. Stamm, J. Clin. Invest. 1992, 90, 965–972.
[15] L. L. W. Cooling, T. A. W. Koerner, S. J. Naides, J.
Infect. Dis. 1995, 172, 1198–1205.
1726
asc.wiley-vch.de
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2008, 350, 1717 – 1728

DEDICATED CLUSTER
FULL PAPERS
Chemoenzymatic Syntheses ofTumor-Associated Carbohydrate Antigen Globo-H
[16] Y. U. Katagiri, K. Ohmi, C. Katagiri, T. Sekino, H. Na-
kajima, T. Ebata, N. Kiyokawa, J. Fujimoto, Glycocon-
jugate J. 2001, 18, 347–353.
[42] R. Kannagi, N. A. Cochran, F. Ishigami, S. Hakomori,
P. W. Andrews, B. B. Knowles, D. Solter, EMBO J.
1983, 2, 2355–2361.
[17] J. Wenk, P. W. Andrews, J. Casper, J. Hata, M. F. Pera,
A. von Keitz, I. Damjanov, B. A. Fenderson, Int. J.
Cancer 1994, 58, 108–115.
[18] J. G. Krupnick, I. Damjanov, A. Damjanov, Z. M. Zhu,
B. A. Fenderson, Int. J. Cancer 1994, 59, 692–698.
[19] R. Kannagi, S. B. Levery, F. Ishigami, S. Hakomori,
L. H. Shevinsky, B. B. Knowles, D. Solter, J. Biol.
Chem. 1983, 258, 8934–8942.
[20] S. Saito, H. Aoki, A. Ito, S. Ueno, T. Wada, K. Mitsuzu-
ka, M. Satoh, Y. Arai, T. Miyagi, J. Biol. Chem. 2003,
278, 26474–26479.
[21] W. F. Steelant, Y. Kawakami, A. Ito, K. Handa, E. A.
Bruyneel, M. Mareel, S. Hakomori, FEBS Lett. 2002,
531, 93–98.
[22] S. Saito, S. Orikasa, M. Satoh, C. Ohyama, A. Ito, T.
Takahashi, Jpn. J. Cancer Res. 1997, 88, 652–659.
[23] S. Saito, S. Orikasa, C. Ohyama, M. Satoh, Y. Fukushi,
Int. J. Cancer 1991, 49, 329–334.
[24] M. Kiso, H. Ishida, A. Hiromne, in: Carbohydrate-
Based Drug Discovery, Vol. 1, (Ed.: C.-H. Wong),
Wiley-VCH, Weinheim, 2003, pp 37–54.
[25] J. R. Allen, J. G. Allen, X.-F. Zhang, L. J. Williams, A.
Zatorski, G. Ragupathi, P. O. Livingston, S. J. Danishef-
sky, Chem. Eur. J. 2000, 6, 1366–1375.
[43] D. R. Mootoo, P. Konradsson, U. Udodong, B. Fraser-
Reid, J. Am. Chem. Soc. 1988, 110, 5583–5584.
[44] X. Huang, L. Huang, H. Wang, X.-S. Ye, Angew. Chem.
2004, 116, 5333–5336; Angew. Chem. Int. Ed. 2004, 43,
5221–5224.
[45] J. D. C. Codꢅe, L. J. van den Bos, R. E. J. N. Litjens,
H. S. Overkleeft, C. A. A. van Boeckel, J. H. van Boom,
G. A. van der Marel, Tetrahedron 2004, 60, 1057–1064.
[46] S. Yamago, T. Yamada, T. Maruyama, J.-I. Yoshida,
Angew. Chem. 2004, 116, 2197–2200; Angew. Chem.
Int. Ed. 2004, 43, 2145–2148.
[47] H. M. Nguyen, J. L. Poole, D. Y. Gin, Angew. Chem.
2001, 113, 428–431; Angew. Chem. Int. Ed. 2001, 40,
414–417.
[48] D. Crich, S. Sun, Tetrahedron 1998, 54, 8321–8348.
[49] L. Huang, X. Huang, Chem. Eur. J. 2007, 13, 529–540.
[50] A. Miermont, Y. Zeng, Y. Jing, X.-S. Ye, X. Huang, J.
Org. Chem. 2007, 72, 8958–8961.
[51] N. Teumelsan, X. Huang, J. Org. Chem. 2007, 72, 8976–
8979.
[52] L. Huang, Z. Wang, X. Li, X.-S. Ye, X. Huang, Carbo-
hydr. Res. 2006, 341, 1669–1679.
[53] L. Huang, Z. Wang, X. Huang, Chem. Commun. 2004,
17, 1960–1961.
[54] V. Martichonok, G. M. Whitesides, J. Org. Chem. 1996,
61, 1702–1706.
[55] D. Crich, M. Smith, J. Am. Chem. Soc. 2001, 123, 9015–
9020.
[56] D. Crich, M. Smith, Org. Lett. 2000, 2, 4067–4069.
[57] C. Wang, H. Wang, X. Huang, L.-H. Zhang, X.-S. Ye,
Synlett 2006, 2846–2850.
[58] J. D. C. Codꢅe, R. E. J. N. Litjens, R. den Heeten, H. S.
Overkleeft, J. H. van Boom, G. A. van der Marel, Org.
Lett. 2003, 5, 1519–1522.
[59] J. D. C. Codꢅe, L. J. van den Bos, R. E. J. N. Litjens,
H. S. Overkleeft, J. H. van Boom, G. A. van der Marel,
Org. Lett. 2003, 5, 1947–1950.
[60] P. Pornsuriyasak, A. V. Demchenko, Carbohydr. Res.
2006, 341, 1458–1466, and references cited therein.
[61] D. Crich, M. Smith, Q. Yao, J. Picione, Synthesis 2001,
323–326.
[62] Y. Gen, L.-H. Zhang, X.-S. Ye, Chem. Commun. 2008,
597–599.
[63] J. Li, J. Wang, P. G. Czyryca, H. Chang, T. W. Orsak, R.
Evanson, C.-W. T. Chang, Org. Lett. 2004, 6, 1381–
1384.
[64] T. Y. Park, I. J. Kim, S. Hu, M. T. Bilodeau, J. T. Ran-
dolph, O. Kwon, S. J. Danishefsky, J. Am. Chem. Soc.
1996, 118, 11488–11500.
[26] J. M. Lassaletta, R. R. Schmidt, Liebigs Ann. Chem.
1996, 1417–1423.
[27] T. Zhu, G. J. Boons, Angew. Chem. 1999, 111, 3704–
3707; Angew. Chem. Int. Ed. 1999, 38, 3495–3497.
[28] D. D. Werz, B. Castagner, P. H. Seeberger, J. Am.
Chem. Soc. 2007, 129, 2770–2771.
[29] C.-Y. Huang, D. A. Thayer, A. Y. Chang, M. Best, L.
Hoffmann, S. Head, C.-H. Wong, Proc. Natl. Acad. Sci.
USA 2006, 103, 15–20.
[30] Z. Wang, L. Zhou, K. El-boubbou, X.-S. Ye, X. Huang,
J. Org. Chem. 2007, 72, 6409–6420.
[31] D. Crich, B. Wu, Tetrahedron 2008, 64, 2042–2047 and
references cited therein.
[32] G. J. Boons, A. Demchenko, Chem. Rev. 2000, 100,
4539–4565.
[33] J. Seifert, M. Lergenmꢄller, Y. Ito, Angew. Chem. 2000,
112, 541–544; Angew. Chem. Int. Ed. 2000, 39, 531–
534.
[34] H. Ishida, R. Miyawaki, M. Kiso, A. Hasegawa, J. Car-
bohydr. Chem. 1996, 15, 163–182.
[35] J. M. Lassaletta, K. Carlsson, P. J. Garegg, R. R.
Schmidt, J. Org. Chem. 1996, 61, 6873–6880.
[36] K. M. Koeller, C.-H. Wong, Chem. Rev. 2000, 100,
4465–4493, and references cited therein.
[37] N. Wymer, E. J. Toone, Curr. Opin. Chem. Biol. 2000,
4, 110–119.
[65] B. Garcia, D. Y. Gin, J. Am. Chem. Soc. 2000, 122,
4269–4279.
[66] A. A.-H. Abdel-Rahman, S. J. Jonke, E. S. H. El Ashry,
R. R. Schmidt, Angew. Chem. 2004, 116, 4489; Angew.
Chem. Int. Ed. 2004, 43, 4389.
[67] D. Crich, J. Picione, Org. Lett. 2003, 5, 781–784.
[68] Z. Zhang, I. R. Ollman, X.-S. Ye, R. Wischnat, T.
Baasov, C.-H. Wong, J. Am. Chem. Soc. 1999, 121, 734–
753.
[38] R. I. Duclos, Carbohydr. Res. 2000, 328, 489–507.
[39] O. Blixt, D. Vasiliu, K. Allin, N. Jacobsen, D. Warnock,
N. Razi, J. C. Paulson, S. Bernatchez, M. Gilbert, W.
Wakarchuk, Carbohydr. Res. 2005, 340, 1963–1972.
[40] V. P. Kamath, R. E. Yeske, J. M. Gregson, R. M. Rat-
cliffe, Y. R. Fang, M. M. Palcic, Carbohydr. Res. 2004,
339, 1141–1146, and references cited therein.
[41] M. Randriantsoa, S. Drouillard, C. Breton, E. Samain,
FEBS Lett. 2007, 581, 2652–2656.
Adv. Synth. Catal. 2008, 350, 1717 – 1728
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
1727

DEDICATED CLUSTER
FULL PAPERS
Zhen Wang et al.
[69] D. Crich, V. Dudkin, Tetrahedron Lett. 2000, 41, 5643–
5646.
[70] D. Crich, W. Li, H. Li, J. Am. Chem. Soc. 2004, 126,
15081–15086.
[71] W. Yi, J. Shao, L. Zhu, M. Li, M. Singh, Y. Lu, S. Lin,
H. Li, K. Ryu, J. Shen, H. Guo, Q. Yao, C. A. Bush,
P. G. Wang, J. Am. Chem. Soc. 2005, 127, 2040–2041.
[72] H. Yu, H. Chokhawala, H. Karpel, H. Yu, B. Wu, J.
Zhang, Y. Zhang, Q. Jia, X. Chen, J. Am. Chem. Soc.
2005, 127, 17618–17619.
[73] H. Yu, S. Huang, H. Chokhawala, M. Sun, H. Zheng,
X. Chen, Angew. Chem. Int. Ed. 2005, 44, 3938–3944.
[74] D. M. Su, H. Eguchi, W. Yi, L. Li, P. G. Wang, C. Xia,
Org. Lett. 2008, 10, in press.
[75] K. Bock, C. Pedersen, J. Chem. Soc. Perkin Trans. 2
1974, 293–297.
1728
asc.wiley-vch.de
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Adv. Synth. Catal. 2008, 350, 1717 – 1728