Systematic syntheses of influenza neuraminidase inhibitors: A series of carbosilane dendrimers uniformly functionalized with thioglycoside-type sialic acid moieties

Article abstract of DOI:10.1016/j.bmc.2009.06.036

In order to develop novel influenza sialidase inhibitors, we constructed a library of glycoclusters composed of twelve types of sialylated dendrimers with thioglycosidic linkage that are resistant to hydrolysis by the sialidases. These sialodendrimers were synthesized by condensation reaction between a thiosialoside modified on the aglycon terminal end by a thioacetyl group and twelve types of carbosilane dendrimers having brominated terminal ends under deacetylation conditions, and temporal re-protection was performed for purification. Removal of all protection of the glycodendrimers was accomplished by transesterification and subsequent saponification to provide corresponding water-soluble glycodendrimers in good yields. For investigation of the structure-activity relationship, dendrimer scaffolds having differences in number of the sugar moieties, such as 3-, 4-, 6- and 12-functionalized dendrimers, and in linkage patterns, such as normal aliphatic linkage, ether- and amide-linkages. Biological evaluations of these glycodendrimers showed that all of the ether- and amide-elongated compounds had inhibitory potencies for the influenza sialidases in the mM range, while compounds having normal aliphatic linkage did not have any activities except for a 12-functionalized compound.

Full text of DOI:10.1016/j.bmc.2009.06.036

Bioorganic & Medicinal Chemistry 17 (2009) 5451–5464
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Systematic syntheses of influenza neuraminidase inhibitors: A series of
carbosilane dendrimers uniformly functionalized with thioglycoside-type
sialic acid moieties q
Jun-Ichi Sakamoto a, Tetsuo Koyama a, Daisei Miyamoto b,c, Sangchai Yingsakmongkon d,
Kazuya I. P. J. Hidari b,c, Wipawee Jampangern e, Takashi Suzuki b,c, Yasuo Suzuki c,f, Yasuaki Esumi g,
Takemichi Nakamura g, Ken Hatano a, Daiyo Terunuma a, Koji Matsuoka a,
*
a Division of Material Science, Graduate School of Science and Engineering, Saitama University, Sakura, Saitama 338-8570, Japan
b Department of Biochemistry, University of Shizuoka, School of Pharmaceutical Sciences, and Global COE Program for Innovation in Human Health Sciences, Suruga,
Shizuoka 422-8526, Japan
c Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012, Japan
d Department of Microbiology and Immunology, Faculty of Veterinary Medicine, Kasetsart University, Bangkok 10900, Thailand
e Department of Medical Sciences, National Institute of Health, Ministry of Public Health, Nonthaburi 11000, Thailand
f Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, Kasugai, Aichi 487-8501, Japan
g The Institute of Physical and Chemical Research (RIKEN), Wako, Saitama 351-0198, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
In order to develop novel influenza sialidase inhibitors, we constructed a library of glycoclusters com-
posed of twelve types of sialylated dendrimers with thioglycosidic linkage that are resistant to hydrolysis
by the sialidases. These sialodendrimers were synthesized by condensation reaction between a thiosialo-
side modified on the aglycon terminal end by a thioacetyl group and twelve types of carbosilane dendri-
mers having brominated terminal ends under deacetylation conditions, and temporal re-protection was
performed for purification. Removal of all protection of the glycodendrimers was accomplished by
transesterification and subsequent saponification to provide corresponding water-soluble glycodendri-
mers in good yields. For investigation of the structure–activity relationship, dendrimer scaffolds having
differences in number of the sugar moieties, such as 3-, 4-, 6- and 12-functionalized dendrimers, and
in linkage patterns, such as normal aliphatic linkage, ether- and amide-linkages. Biological evaluations
of these glycodendrimers showed that all of the ether- and amide-elongated compounds had inhibitory
potencies for the influenza sialidases in the mM range, while compounds having normal aliphatic linkage
did not have any activities except for a 12-functionalized compound.
Received 5 June 2009
Revised 17 June 2009
Accepted 18 June 2009
Available online 21 June 2009
Keywords:
Thioglycosides
Sialic acid
Glycodendrimers
Sialidase inhibitors
Influenza virus
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
receptors on host cells with viral infection beginning.6 Then in
order to spread infection, replicated viruses are released from the
There has been much concern recently about the possibility of a
worldwide influenza outbreak leading to pandemic flu. The most
catastrophic pandemic was Spanish flu in 1918–1919, which
claimed the lives of at least 20 million people.1 If a new type of
pandemic flu occurs, it is expected that the flu will kill about 62
million people worldwide.2 Medicines that might be useful for a
forthcoming pandemic are TamifluÒ3 and RelenzaÒ,4 which are
used as inhibitors against influenza viral glycosidases. Influenza
A and B viruses have two types of sialic acid-recognizing glycopro-
teins on their surfaces.5 One of them is hemagglutinin (HA), which
is a kind of lectin and binds to sialyloligosaccharides as specific
infected cells by enzymatic cleavage of the sialic acid residues of
the sialyloligosaccharides by the other surface glycoprotein,
referred to as either sialidase or neuraminidase (NA).7 TamifluÒ
and RelenzaÒ have potent inhibitory activities against infection
spread because they plug the active sites of neuraminidases by
interaction with the amino acids forming the active sites and
thereby inhibit the viral particle release process and cut off the
virus multiplication cycle. However, Tamiflu-resistant viruses,
which have a hidden potential to cause an outbreak of infection,
have already been isolated.8 Therefore, treatment of influenza
infection requires other novel anti-viral agents to prepare for an
influenza outbreak.
Although molecular designs of TamifluÒ and RelenzaÒ are based
on the transition state analogue of sialic acid in enzymatic hydro-
lysis of sialoside by neuraminidases, substrates for neuraminidases
q
Glyco-silicon functional materials, Part 11. For Part 10, see Ref. 16b.
* Corresponding author. Tel./fax: +81 48 858 3099.
E-mail address: koji@fms.saitama-u.ac.jp (K. Matsuoka).
0968-0896/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.06.036

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J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
are absolutely natural sialic acids. Consequently, we focused on
thioglycoside of sialic acid as a wild-type artificial substrate for
neuraminidases because thioglycosidic linkage is generally resis-
tant to hydrolysis by glycosidases, such as neuramindases.9 The
substrates plug active sites of neuraminidases and prevent infec-
tion spread. In addition, we planned to use the glyco-clustering
effect10 since monomeric sialoside does not have inhibitory po-
tency for neuraminidases. Glycoclusters generally express greater
affinity between glycosides and sugar-recognition proteins than
those of corresponding monomeric glycosides. Influenza HAs and
NAs also have trimeric11 and tetrameric12 structures, respectively,
in order to enhance sialic acid-recognition activity. Therefore, the
thioglycosidic sialocluster is expected to be a promising multiva-
lent-type therapeutic agent for influenza disease. In fact, we have
designed and synthesized a linear polymer expressing pendant-
type thiosialoside moieties that showed inhibitory activity against
influenza virus NAs.13
Recently, we have synthesized various carbosilane dendrimers
having bioactive carbohydrate moieties on the terminal ends14
and investigated their biological activities.15 For example, it was
found that sialyllactose-modified dendrimers16a have potent bind-
ing activities for influenza HAs.16b As a part of our studies on
glycodendrimers, we previously reported six types of carbosilane
dendrimers having thioglycoside-type sialic acid moieties and
their inhibitory activities against human influenza NAs.17 In this
paper, we describe a systematic synthesis of a series of sialodendri-
mers based on carbosilane scaffolds and the preliminary results of
their inhibition assays for human influenza NAs.
the ability to support carbohydrate moieties in our ongoing
synthetic studies on glycoclusters. For investigation of the struc-
ture–activity relationship (SAR) for human influenza NAs, we
selected
a series of carbosilane dendrimers that consist of
Fan(0)3, Ball(0)4, Dumbbell(1)6 and Ball(1)12, named by their
shapes, generation number and number of terminal ends (Fig. 1).
In addition, on each framework, novel ether-linkage dendrimers17
as well as known normal aliphatic dendrimers14 and amide-link-
age dendrimers18 were used for clustering of thiosialosides. There-
fore, we thought that a library of thiosialoside clusters could be
constructed and that the most appropriate structure for a neur-
aminidase inhibitor would be determined.
2.1. Synthesis of sialoside having thioglycosidic linkage
Synthesis of thioglycosidic sialoside for clustering on carbosi-
lane dendrimers is shown in Scheme 1. Known b-sialyl-chloride
1719 was converted into
a-anomeric thioacetate 18 according to
Hasegawa’s method,20 which was SN2-type reaction in the pres-
ence of potassium thioacetate in CH2Cl2. In this method, produced
thioacetate 18 was contaminated with glycal product 19 (5ꢀ10%)
formed by elimination reaction of chloride 17.21 Chromatographic
purification or re-crystallization for separation of thioacetate 18
from glycal 19 was unsuccessful at this stage. Therefore, a mixture
of 18 and 19 was treated with potassium carbonate in methanol to
generate potassium thiolate, which reacted with 5-bromo-1-pen-
tene. After re-acetylation as temporal protection for purification,
pure pentenyl thiosialoside 20 was obtained in 84.2% yield. The
sialoside 20 has slightly better mobility on TLC than that of glycal
19 and could be isolated from the afforded mixture by flash col-
umn chromatography on silica gel. The 1H NMR spectrum of 20
indicated that the newly produced thioglycosidic linkage of sialic
2. Results and discussion
We designed a series of oligomeric thiosialosides using carbos-
ilane dendrimers as core scaffolds, since the carbosilanes showed
acid is
a
-configuration based on the empirical rule22: J7,8 = 8.2 Hz,
R
Si
R
R
R
OH
COOH
S
OH
O
R
R
HO
Si
R
R
R
AcHN
HO
Si
1
6: R=R1
7: R=R2
8: R=R3
3: R=R1
4: R=R2
5: R=R3
R
R
R
R
R
Fan(0)3
Ball(0)4
R
R
Si
Si
Si
Si
OH
OH
COOH
S
HO
AcHN
O
R
R
R
R
HO
S
S
Me
COOH
S
OH
R
Si
Si
Si
OH
12: R=R1
13: R=R2
14: R=R3
9: R=R1
10: R=R2
11: R=R3
O
R
R
Me
HO
AcHN
R
HO
R
2
Dumbbell(1)6
Ball(1)12
OH
OH
COOH
COOH
S
OH
OH
O
R1 =
R2 =
O
HO
HO
S
S
O
S
AcHN
AcHN
HO
HO
ether-elongated
normal
OH
COOH
S
H
N
OH
O
HO
R3 =
S
AcHN
HO
O
amide-elongated
Figure 1. Synthetic substrate of thioglycoside-type sialic acid residue on terminal ends.

J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
5453
OH
OAc
AcO
OAc
AcO
OH
Cl
COOMe
OH
OH
OH
O
OAc
OAc
OH
O
ii)
i)
iii)
HO
AcO
AcO
HO
O
O
19
SAc +
COOMe
COOMe
COOH
AcHN
AcHN
AcHN
AcHN
HO
HO
15 (Neu5Ac)
17
18
16
OAc
OAc
AcO
OAc
AcO
COOMe
S
COOMe
OAc
OAc
OAc
O
iv)
v)
COOMe
AcO
AcO
AcO
O
O
S
SAc
AcHN
AcHN
AcHN
AcO
vi)
19
20
21
1
Scheme 1. Reagents and conditions: (i) MeOH, Dowex-50 (H+), rt, overnight; (ii) AcCl, rt, 24 h; (iii) AcSK, CH2Cl2, rt, 24 h ; (iv) 5-bromo-1-pentene, K2CO3, MeOH, 0 °C?rt, 3 h,
then Ac2O, Pyr., rt, overnight; (v) AcSH, AIBN, 1,4-dioxane, 50 °C?80 °C, 3 h; (vi) NaOMe, MeOH, rt, 0.5 h, then 0.5 M aq NaOH, rt, overnight.
D
d |H-9a–H9b| = 0.20 ppm. Finally, radical addition of thioacetic
2.3. Elongation of spacer-arm of carbosilane dendrimer by
amide-linkage
acid16 into terminal olefin of pentenyl sialoside 20, using AIBN as
a radical initiator, gave corresponding primary thioacetate 21 in
quantitative yield. Further transformation of 20 was performed
by means of transesterification and subsequent saponification to
afford water-soluble thiosialoside 1 in quantitative yield, which
was used as a monomeric substrate for the biological assay.
Amide-elongation was carried out by the method reported pre-
viously18 (Scheme 3). Mesylate of Dumbbell(1)6 converted from
known hexaol 26 was replaced by azide anion in DMF to give hexa-
azide 28 in good yield. Reduction of azide groups of 28 in Stauding-
er’s condition followed by treatment with aqueous HCl solution
quantitatively yielded a hexaamine hydrochloride salt, which
was condensed with 6-bromohexanoyl chloride to afford the
desired amide-elongated dendrimer 37. Syntheses of twelve types
of carbosilane dendrimers having differences in the number of bro-
mo atoms on each terminal end, spacer lengths and linkage pat-
terns, including normal aliphatic (29,14b 32,14c 35,14c 3814c),
2.2. Elongation of spacer-arm of carbosilane dendrimers by
ether-linkage
A synthetic scheme of ether-linked carbosilane dendrimers is
shown in Scheme 2. Known triol 2214b was reacted with allyl bro-
mide in THF under sodium hydride-mediated Williamson’s ether
synthesis condition to afford allyl ether 23. Sequential functional-
ity conversion for preparation of carbosilane dendrimers having
bromo atoms on the terminal ends was carried out. Thus, the ter-
minal olefins were converted into a hydroxyl group by hydrobora-
tion using sterically-hindered dicyclohexylborane, which
improved the terminal selectivity to afford ether-elongated triol
24 after oxidation under an alkali condition. Triol 24 was treated
with methanesulfonyl chloride in pyridine to give mesylate, which
underwent replacement reaction by sodium bromide in DMF to
give corresponding tribrominated dendrimer 30. The other three
types of hydroxylated dendrimers 2518a, 2614c and 2723 also under-
went similar chemical conversion to give ether-elongated dendri-
mers 33, 36, and 39, respectively.
ether- (30, 33, 36, 39) and amide-linked (31,18a 34,18a 37, 4018b
dendrimers, were efficiently accomplished and the structures of
the carbosilanes are shown in Table 1.
)
2.4. Construction of carbosilane dendrimers having
thiosialosides
Having prepared the thiosialoside and a series of carbosilane
dendrimers, we carried out a reaction in which the sialoside was
introduced into the dendrimers. The thioacetate 21 was condensed
with known tribromodendrimer 2914b under a deacetylation con-
dition in the presence of sodium methoxide in DMF–methanol
(Scheme 4). In this reaction, generated thiolates of the sialoside
i)
Si
O
Si
OH
3
3
22
23
iii), iv)
ii)
Si
O
Br
Si
O
OH
3
3
30
24
v)
Si
O
Br
Si
OH
4
2
4
33
25
Me
Me
Me
v)
v)
Si
Si
O
Br
Si
Si
OH
3
3
2
Me
26
36
Si
Si
Si
O
Br
Si
OH
3
3
4
4
27
39
Scheme 2. Reagents and conditions: (i) NaH, AllBr, THF, reflux temperature, 3 h; (ii) dicyclohexylborane, THF, rt, 3 h, then 3 M aq NaOH, 30% aq H2O2, 60 °C, 2 h; (iii) MsCl,
Pyr., 0 °C, 3 h; (iv) NaBr, DMF, 60 °C, 3 h; (v) (i)ꢀ(iv).

5454
J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
O
Me
Si
Me
Me
Me
i), ii)
iii), iv), v)
Br
26
Si
N
H
Si
Si
N
3
3
3
2
2
28
37
Scheme 3. Reagents and conditions: (i) MsCl, Pyr., 0 °C, 3 h; (ii) NaN3, DMF, 60 °C, 3 h; (iii) PPh3, H2O, THF, rt, overnight, then 1 N HCl aq; (v) Et3N, 6-bromohexanoyl chloride,
MeOH, ꢀ20 °C, 1.5 h.
substituted bromine atoms on terminal ends of dendrimers to give
fully protected three-functional sialodendrimer 41 in 80.2% yield
after re-protection. At the same time, fully protected dimeric
byproduct 53 having disulfide linkage was also obtained. In the
1H NMR spectrum of 41, the integral ratio due to corresponding
protons elucidated that one dendrimer core scaffold had three
sialoside moieties; SiCH2/CH2SCH2/Ph/H-4(Neu) = 6:12:5:3. In
addition to the NMR spectrum, a high-resolution ESI-MS spectrum
of 41 proved construction of this compound; calcd for [M+H]+:
2056.71535; found m/z: 2056.72008. In the final step, removal of
all protections of sialodendrimer 41 was carried out by a combina-
tion of transesterification and subsequent saponification to give
water-soluble sialodendrimer 3 in 87.2% yield after size exclusion
chromatography using a Sephadex G-25 column followed by
lyophilization. Production of 3 was supported by the results of
the high-resolution ESI-MS spectroscopy; calcd for [M+H]+:
1510.54162; found m/z: 1510.53494. Each of the other eleven
types of sialodendrimers 4ꢀ14 was efficiently synthesized by sim-
ilar condensation reaction and subsequent de-protection, and
details are shown in Section 4. A library of thiosialodedrimers
having twelve different types of core scaffolds was successfully
constructed. Furthermore, removal of the protective groups in 53
was carried out by a method similar to that described for the sialo-
dendrimer to afford water-soluble dimeric sialoside 2 in high yield
(Scheme 4), which was also used as a comparative substrate for
biological assays.
2.5. Inhibitory sialidase activity assay
Since a series of thiosialoside clusters 3ꢀ14 as well as thiogly-
cosidic monomer 1 and dimer 2 had been prepared, biological
evaluation of the thiosialosides for inhibitory activity against
Table 1
Brominated carbosilane dendrimers 29ꢀ40 and corresponding sialodendrimers (protected 41ꢀ52 and free 3ꢀ14)
OAc
AcO
OH
COOMe
S
COOH
S
OAc
O
OH
O
AcO
HO
A =
B =
S
S
AcHN
AcHN
HO
Normal
Ether-elongated
Amide-elongated
O
Si
R
Si
O
R
3
R
3
Si
N
H
3
Fan(0)3
29: R = Br
41: R = A
3: R = B
30: R = Br
42: R = A
4: R = B
31: R = Br
43: R = A
5: R = B
O
Si
R
Si
O
R
4
R
4
Si
N
H
4
Ball(0)4
32: R = Br
44: R = A
6: R = B
33: R = Br
45: R = A
7: R = B
34: R = Br
46: R = A
8: R = B
Me
Si
Me
Me
Si
Me
O
Me
Si
Me
Si
Si
O
R
R
R
3
2
3
Si
N
H
2
3
2
Dumbbell(1)6
35: R = Br
47: R = A
9: R = B
36: R = Br
48: R = A
10: R = B
37: R = Br
49: R = A
11: R = B
O
Si
Si
R
Si
Si
O
R
R
3
3
4
Si
Si
N
4
3
H
4
Ball(1)12
38: R = Br,
50: R = A,
12: R = B
39: R = Br
51: R = A
13: R = B
40: R = Br
52: R = A
14: R = B

J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
5455
1
1
OR
1
2
OR
1
1
2
OR COOR
OR
O
COOR
S
1
i)
1
R O
R O
O
S
Si
Br
S
Si
3
S
AcHN
+
AcHN
3
1
R O
R O
2
53: R1 = Ac, R2 = Me
2: R1 = R2 = H
29
41: R1 = Ac, R2 = Me
3: R1 = R2 = H
ii)
ii)
Scheme 4. Reagents and conditions: (i) 20, MeONa, MeOH, DMF, rt, overnight, then Ac2O, Pyr., rt, overnight; then CH2N2 in Et2O, MeOH, 0 °C; (ii) MeONa, MeOH, rt, 3 h, then
0.5 M aq NaOH, rt, overnight.
Table 2
neuraminidases of two subtypes of human influenza virus (H1N1
Preliminary results of inhibition assays of a series of glycodendrimers against human
influenza virus sialidases
and H3N2) was performed The IC50 values measured by a previ-
ously reported method24 are summarized in Table 2. Interestingly,
Compound
Inhibitory potency (mM)a
all of the ether- and amide-elongated sialodendrimers and
Ball(1)12-S-Neu5Ac12 (12) showed inhibitory potencies not only
for H3N2-type sialidase but also for H1N1-type sialidase in the
mM range, although monomeric sialoside 1 and dimeric sialoside
2 as well as normal aliphatic-type dendrimers 3, 6, 9 did not show
any inhibitory activities. The most potent inhibitory activities were
observed when Dumbbell(1)6-ether-S-Neu5Ac6 (10) for H3N2-
type sialidase and Fan(0)3-amide-S-Neu3 (5) as well as Dumb-
bell(1)6-amide-S-Neu6 (11) for H1N1-type sialidase were used as
substrates. Among the normal aliphatic-type dendrimers (3, 6, 9,
12), only Ball(1)12-type dendrimer 12 showed inhibitory activity,
indicating the glycocluster effect was effectively functioned with
enhancing the binding affinity. However, among ether- and
amide-elongated dendrimers, Ball(1)12-ether-Neu5Ac12 (13) and
Ball(1)12-amide-Neu5Ac12 (14) interestingly showed the weakest
inhibition potencies, respectively, indicating that the cluster effect
functioned toward adverse direction. According to these results,
elongation of the dendrimer may have a more favorable influence
than increase in the number of sialic acid moieties for inhibitory
activities against sialidases. Therefore, it is thought that the dis-
tance between the sialic acid moieties as well as the degree of free-
dom of the sialic acid moieties are important for effective binding
to active sites on the tetrameric sialidases on virus surfaces.
A/Memphis/1/71
(H3N2)
A/PR/8/34
(H1N1)
S-Neu5Ac (1)
S-Neu5Ac2 (2)
N.D.b
N.D.b
N.D.b
5.00
7.1
N.D.b
N.D.b
N.D.b
5.00
2.7
Fan(0)3-S-Neu5Ac3 (3)
Fan(0)3-ether-S-Neu5Ac3 (4)
Fan(0)3-amide-S-Neu5Ac3 (5)
Ball(0)4-S-Neu5Ac4 (6)
Ball(0)4-ether-S-Neu5Ac4 (7)
Ball(0)4-amide-S-Neu5Ac4 (8)
N.D.b
5.00
8.8
N.D.b
5.00
4.8
Dumbbell(1)6-S-Neu5Ac6 (9)
Dumbbell(1)6-ether-S-Neu5Ac6 (10)
Dumbbell(1)6-amide-S-Neu5Ac6 (11)
N.D.
1.25
8.1
N.D.
5.00
2.8
Ball(1)12-S-Neu5Ac12 (12)
Ball(1)12-ether-S-Neu5Ac12 (13)
Ball(1)12-amide-S-Neu5Ac12 (14)
7.0
6.5
10.0<
5.0
6.4
7.2
a
IC50 value are indicated in millimolar concentration (mM) based on a mono-
meric sugar unit concentration.
b
N.D. means not determined due to weak inhibition.
reagents were used without further purification. Pyridine (Pyr.)
and dimethylformamide (DMF) were stored over molecular sieves
(4 Å MS), and methanol (MeOH) was stored over 3 Å MS prior to
use. Tetrahydrofuran (THF) was dried over sodium benzophenone
ketyl under argon atmosphere and distilled prior to use. 20-(4-
3. Conclusion
Systematic construction of a library of a series of carbosilane
dendrimers carrying thioglycosidic sialic acid moieties has been
efficiently accomplished. These glycodendrimers were synthesized
on the basis of introduction of thiosialoside functionalized on the
aglycon terminal end with a thioacetyl group into twelve types
of brominated carbosilane dendrimers having difference in the
dendrimer frame and linkage pattern. Biological evaluation of
these sialodendrimers against human influenza virus sialidases
was carried out, and the results showed that all of the ether- and
amide-elongated dendrimers have inhibitory activity against the
sialidase. Dumbbell(1)6-ether-S-Neu6 (10) for H3N2-type sialidase
and Fan(0)3-amide-S-Neu3 (5) as well as Dumbbell(1)6-amide-S-
Neu6 (11) for H1N1-type sialidase showed the most potent
inhibition.
Methylumbelliferyl)-a-D-N-acetylneuraminic acid (4-MU-Neu5Ac)
was purchased from Sigma. Reactions were monitored by TLC
using Merck silica gel plates 60 F254 (0.25 mm in layer thickness).
Detection was carried out by UV light when applicable and treat-
ment with a solution of 85:10:5 (v/v/v) MeOH/p-anisaldehyde/
concd sulfuric acid followed by charring or exposure to an iodide
atmosphere for visualization. Purification by column chromatog-
raphy was performed on silica gel (Silica Gel 60; 63ꢀ200
Merck). Flash column chromatography was performed on silica
gel (Silica Gel 60, spherical neutral; 40ꢀ100 m, E. Merck). The
lm, E.
l
ratio between silica gel and crude product is 20ꢀ50:1 (w/w). IR
spectra were recorded on a Shimadzu IR-Prestige 21 using KBr
pellet (for solid) or as neat sample (for liquid). 1H and 13C NMR
spectra were recorded using a Bruker AM 400 and a Buruker
DRX 400 (at 400 MHz for 1H and 100 MHz for 13C) or a Varian
Gemini 2000 (at 200 MHz for 1H and 50 MHz for 13C) in CDCl3
or D2O. Chemical shifts are expressed as parts per million (ppm,
d) and are relative to an internal tetrametylsilane (TMS) in CDCl3
(d 0.0) or HDO in D2O (d 4.78) for 1H, and CDCl3 (d 77.0), MeOD (d
49.0) or acetone (d 215.0) for 13C. FAB, ESI and MALDI-TOF mass
spectra were obtained with a JEOL JMS-700AM spectrometer, a
JEOL JMS-HX110A and a Buruker Autoflex spectrometer, respec-
Further investigation of different types of thioglycosidic sialo-
sides and the clustering of the sialosides using dendric core scaf-
folds is now in progress. Details of the syntheses and their
biological evaluation will be reported elsewhere.
4. Experimental
4.1. General methods
tively. Elemental analyses were performed with
a Fisons
EA1108 on samples extensively dried in vacuo at 40ꢀ50 °C over
All reactions were carried out in dried solvents under an argon
atmosphere. Unless otherwise stated, all commercial solvents and
phosphorus pentoxide for 4ꢀ5 h.

5456
J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
4.1.1. Methyl (n-pentenyl 5-acetamido-4,7,8,9-tetra-O-acetyl-
21.04, 20.73, 20.65.; MS (FAB): calcd for [M+H]+: 651.2, found:
m/z 652.0.
Anal. Calcd for C27H41O13NS2: C, 49.76; H, 6.34; N, 2.15. Found:
C, 49.83; H, 6.33; N, 2.17.
3,5-dideoxy-2-thio-
onate (20)
a-D-glycero-D-galacto-2-nonulopyranosid)-
To a solution of anomeric thioacetate 18 (2.26 g, 4.11 mmol)
and 5-bromo-1-pentene (974 L, 8.22 mmol) in MeOH (22 mL)
l
was added potassium carbonate (568 mg, 4.11 mmol) at 0 °C, and
the mixture was stirred for 3 h at room temperature. After quench-
ing of the reaction with acetic acid (50 lL), the reaction mixture
4.1.3. n-Pentenyl 5-acetamido-3,5-dideoxy-2-thio-
-galacto-2-nonulopyranoside sodium salt (1)
Fully protected n-pentenyl S-sialoside 20 (300 mg, 0.521 mmol)
a-D-glycero-
D
was concentrated. The residue was treated with acetic anhydride
(20 mL) in pyridine (20 mL) at room temperature overnight. After
concentration (co-evaporation with toluene), the residual mixture
was extracted with CHCl3. The extraction solution was washed
with 1 M aq H2SO4, satd aq NaHCO3 and brine, dried over anhyd
MgSO4, filtered, and concentrated. The residue was purified by
silica gel column chromatography [2:3 (v/v) toluene/EtOAc] to give
n-pentenyl thiosialoside 20 (1.99 g, 84.2%) as a white foam: Rf 0.52
was dissolved in 1 M NaOMe in MeOH (1 mL), and the solution was
stirred for 0.5 h at room temperature. Then 0.5 M aq solution of
NaOH was added and the reaction mixture was stirred overnight
at room temperature. The solution was treated with IR-120B (H+)
resin and adjusted to pH ꢃ4, and then the suspension was filtered.
The resin was washed with water, and the filtrate and washings
were combined and concentrated. The resulting mixture was puri-
fied by size exclusion column chromatography on Sephadex G-15
with 5% aq AcOH as an eluent to give de-protected n-pentenyl
S-sialoside 1 (199 mg, quantitative) as a white powder after lyoph-
[5:4:1 (v/v/v) CHCl3ꢀEtOAcꢀMeOH]; ½a D27
ꢁ
+29.2 (c 1.13, CHCl3); IR
C@O, amide I), 1549 (dNꢀH
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz,
(KBr): 2940 (
amide II), 1227 (
m
C–H), 1742 (
m
C@O), 1663 (
m
,
mCꢀO), 1038 (
m
;
ilization: Rf 0.47 [8:6:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D25
ꢁ
+21.3
C@O),
1638 ( CꢀO), 1034
;
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz, D2O): d 5.87 (m, 1H, CH@CH2),
CDCl3): d 5.78 (m, 1H, CH@CH2), 5.37 (ddd, 1H, J7,8 = 8.2 Hz,
J8,9a = 2.4 Hz, J8,9b = 4.9 Hz, H-8), 5.33 (dd, 1H, J6,7 = 2.0 Hz, H-7),
5.24 (d, 1H, J5,NH = 10.4 Hz, NH), 5.03, 4.97 (2 ꢂ dd, 2H,
Jtrans = 17.1 Hz, Jcis = 10.2 Hz, Jgem = 1.7 Hz, CH@CH2), 4.86 (ddd,
1H, J4,5 = 10.4 Hz, J3ax,4 = 12.3 Hz, J3eq,4 = 4.6 Hz, H-4), 4.32 (dd, 1H,
J9a,9b = 12.4 Hz, H-9a), 4.12 (dd, 1H, H-9b), 4.05 (q, 1H,
J5,6 = 10.4 Hz, H-5), 3.84 (dd, 1H, H-6), 3.80 (s, 3H, OMe),
2.79ꢀ2.72 (m, 1H, SCHa), 2.72 (dd, 1H, J3ax,3eq = 12.3 Hz, H-3eq),
2.60ꢀ2.53 (m, 1H, SCHb), 2.21ꢀ2.08 (m, 2H, SCH2CH2CH2), 2.16,
2.14, 2.04, 2.03 (4 ꢂ s, 12H, OAc), 1.99 (t, 1H, H-3ax), 1.88 (s, 3H,
NAc), 1.71ꢀ1.53 (m, 2H, SCH2CH2); 13C NMR (100 MHz, CDCl3): d
170.86, 170.54, 170.11, 170.08, 169.98, 168.59, 137.59, 111.14,
83.23, 74.20, 69.69, 68.90, 67.48, 62.19, 52.83, 49.50, 38.16,
32.65, 28.56, 28.34, 23.14, 21.08, 20.78, 20.69; MS (FAB): calcd
for [M+H]+: 576.2, found: m/z 576.0; calcd for [M+Na]+: 598.2,
found: m/z 597.9.
(c 1.07, H2O); IR (KBr): 3399 (mOꢀH), 2930 (mCꢀH), 1707 (m
mC@O, amide I), 1560 (dNꢀH, amide II), 1279 (m
(m
5.09, 5.03 (dd and d, 1H, Jtrans = 17.3 Hz, Jcis = 10.2 Hz, Jgem = 1.6 Hz,
CH@CH2), 3.87ꢀ3.58 (m, 7H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4),
2.81 (dd, 1H, J3eq,4 = 4.8 Hz, J3ax,3eq = 12.8 Hz, H-3eq), 2.83ꢀ2.73 (m,
1H, SCHa), 2.68ꢀ2.61 (m, 1H, SCHb), 2.21ꢀ2.09 (m, 2H,
CH2CH@CH2), 2.04 (s, 3H, NAc), 1.78 (t, J3ax,4 = 11.7 Hz, H-3ax),
1.74ꢀ1.63 (m, 2H, SCH2CH2CH2); 13C NMR (100 MHz, D2O): d
174.68, 173.18, 138.08, 114.71, 84.95, 74.47, 71.30, 67.86, 67.72,
62.19, 51.33, 40.38, 31.80, 28.48, 27.94, 21.64; MS (MALDI-TOF):
calcd for [M+Na]+: 438.117, found: m/z 437.996; Calcd for
[M+K]+: 454.091, found: m/z 453.980.
Anal. Calcd for C16H26NO8SNaꢄ1.0H2O: C, 44.34; H, 6.51; N, 3.23.
Found: C, 44.53; H, 6.62; N, 3.19.
Anal. Calcd for C25H37NO12S: C, 52.16; H, 6.48; N, 2.43. Found: C,
52.02; H, 6.48; N, 2.38.
4.1.4. Tris(4-oxa-6-heptenyl)phenylsilane (23, Fan(0)3-ether-
All)
Fifty five percent of NaH dispersion in mineral oil (1.07 g,
24.52 mol) was washed with n-hexane and suspended in THF
(10 mL). A solution of triol 22 (1.15 g, 4.07 mmol) in THF (15 mL)
was dropwise added to the suspension at 0 °C, followed by addition
of 3-bromopropene (2.11 mL, 24.38 mmol), and the mixture was
stirred for 3 h at reflux temperature (ca. 70 °C). The reaction was
quenched by dropwise addition of MeOH (ꢃ5 mL) at 0 °C. The con-
centrated mixture was dissolved in water and twice extracted with
Et2O. The combined extract was washed with brine, dried over an-
hyd MgSO4, filtered, and concentrated. The syrupy residue was
chromatographed on silica gel [15:1 (v/v) n-hexaneꢀEtOAc] to
afford allyl ether 23 (1.44 g, 87.9%) as a colorless liquid: Rf 0.49
4.1.2. Methyl (5-acetylthiopentyl 5-acetamido-4,7,8,9-tetra-O-
acetyl-3,5-dideoxy-2-thio-a-D-glycero-D-galacto-2-
nonulopyranosid)onate (21)
AIBN (996 mg, 6.08 mmol) was added to a stirred solution of n-
pentenyl sialoside 20 (1.75 g, 3.04 mmol) and thioacetic acid
(7.4 mL, 103.53 mmol) in 1,4-dioxane (7.4 mL) at 50 °C. The mix-
ture was stirred for 2 h at 80 °C. After addition of cyclohexene
(20.9 mL, 206.32 mmol) at 0 °C, the mixture was stirred at room
temperature for a few minutes and concentrated. The resulting
mixture was purified by silica gel column chromatography
[1:0 to 0:1 (v/v) toluene/EtOAc] to afford thioacetate 21 (1.98 g,
quantitative) as a white foam: Rf 0.57 [5:4:1 (v/v/v) CHCl3ꢀ
[5:1 (v/v) n-hexaneꢀEtOAc]; IR (neat): 2930 (
mCꢀH), 2853 (mCꢀH),
1107 (
m
CꢀOꢀC) cmꢀ1
;
1H NMR (200 MHz, CDCl3): d 7.52ꢀ7.29 (m,
EtOAcꢀMeOH]; ½a D28
ꢁ
+25.4 (c 1.11, CHCl3); IR (KBr): 2938 (
C@O, amide I), 1541 (dN–H, amide II), 1227 (mCꢀO,
mCꢀH),
5H, SiPh), 5.99ꢀ5.80 (m, 3H, CH@CH2), 5.30ꢀ5.11 (m, 6H,
CH@CH2), 3.95ꢀ3.90 (m, 6H, CH2CH@CH2), 3.37 (t, 6H, J = 6.9 Hz,
SiCH2CH2CH2O), 1.69ꢀ1.54 (m, 6H, SiCH2CH2CH2O), 0.87ꢀ0.79
(m, 6H, SiCH2); 13C NMR (50 MHz, CDCl3): d 136.58, 134.94,
133.97, 128.87, 127.69, 116.56, 73.06, 71.63, 23.90, 8.21; MS
(FAB): calcd for [M+Na]+: 425.2, found: m/z 425.1.
1744 (
m
C@O), 1663 (
m
ester), 1036 (m
CꢀOꢀC) cmꢀ1; 1H NMR (400 MHz, CDCl3): d 5.36 (ddd,
1H, J7,8 = 8.6 Hz, J8,9a = 2.1 Hz, J8,9b = 4.3 Hz, H-8), 5.33 (dd, 1H,
J6,7 = 1.6 Hz, H-7), 5.26 (d, 1H, J5,NH = 10.7 Hz, NH), 4.86 (ddd, 1H,
J4,5 = 10.7 Hz, J3ax,4 = 12.3 Hz, J3eq,4 = 4.6 Hz, H-4), 4.31 (dd, 1H,
J9a,9b = 12.9 Hz, H-9a), 4.11 (dd, 1H, H-9b), 4.05 (q, 1H,
J5,6 = 10.7 Hz, H-5), 3.83 (dd, 1H, H-6), 3.80 (s, 3H, OMe), 2.86 (t,
2H, J = 7.2 Hz, CH2SAc), 2.77ꢀ2.69 (m, 1H, SCHa), 2.71 (dd, 1H,
J3ax,3eq = 12.6 Hz, H-3eq), 2.57ꢀ2.50 (m, 1H, SCHb), 2.32 (s, 3H,
SAc), 2.16, 2.14, 2.04, 2.03 (4 ꢂ s, 12H, OAc), 1.98 (t, 1H, H-3ax),
1.88 (s, 3H, NAc), 1.61ꢀ1.40 (m, 6H, SCH2CH2CH2CH2CH2SAc); 13C
NMR (100 MHz, CDCl3): d 195.66, 170.80, 170.48, 170.07, 170.04,
169.92, 168.54, 83.17, 74.16, 69.69, 68.82, 67.45, 62.21, 52.79,
49.44, 38.11, 30.50, 28.97, 28.85, 28.75, 28.63, 27.83, 23.08,
Anal. Calcd for C25H37NO12S: C, 71.59; H, 9.51. Found: C, 71.64;
H, 9.70.
4.1.5. Tris(7-hydroxy-4-oxaheptyl)phenylsilane (24, Fan(0)3-
ether-OH)
To a 1 M solution of BH3–THF complex in THF (2.24 mL) was
added cyclohexene (453 lL, 4.47 mmol) at 0 °C, and the mixture
was stirred for 2.5 h at 0 °C. A solution of allyl ether 23 (150 mg,
0.373 mmol) in THF (5 mL) was dropwise added at 0 °C, and the

J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
5457
reaction mixture was stirred for 3 h at room temperature. To the
[5:1 (v/v) n-hexaneꢀEtOAc]; IR (neat): 2928 (
m
CꢀH), 2851 (
m
CꢀH),
mixture were dropwise added MeOH (0.2 mL), 3 M aq NaOH solu-
tion (2 mL) and 30% aq H2O2 (1 mL) at 0 °C, successively, and then
the heterogeneous mixture was stirred at 60 °C for 2 h. The organic
layer was separated from the reaction mixture, and the aq layer
was twice extracted with EtOAc. The organic layer and extract
were combined, washed with brine, dried over anhyd MgSO4, fil-
tered, and concentrated. The residual syrup was purified by flash
column chromatography on silica gel [50:1 (v/v) CHCl3ꢀMeOH]
to afford triol 24 (143 mg, 84.1%) as a light yellow syrup: Rf 0.33
1103 (
m
CꢀOꢀC) cmꢀ1
;
1H NMR (200 MHz, CDCl3): d 6.01ꢀ5.82 (m,
4H, CH@CH2), 5.32ꢀ5.13 (m, 8H, CH@CH2), 3.97ꢀ3.93 (m, 8H,
CH2CH@CH2), 3.36 (t, 8H, J = 7.0 Hz, SiCH2CH2CH2O), 1.66ꢀ1.50
(m, 8H, SiCH2CH2CH2O), 0.58ꢀ0.50 (m, 8H, SiCH2); 13C NMR
(50 MHz, CDCl3): d 134.94, 116.67, 73.29, 71.71, 23.99, 8.07; MS
(FAB): calcd for C24H45O4Si [M+H]+: 425.3, found: m/z 425.1.
Anal. Calcd for C24H44O4Si: C, 67.88; H, 10.44. Found: C, 68.02;
H, 10.73.
[10:1 (v/v) CHCl3ꢀMeOH]; IR (neat): 3395 (
m
OꢀH), 2930 (
mCꢀH),
4.1.9. Tetrakis(7-hydroxy-4-oxaheptenyl)silane (Ball(0)4-ether-
OH)
2866 ( CꢀH), 1111 (
m
m
CꢀOꢀC) cmꢀ1 1H NMR (200 MHz, CDCl3): d
;
7.49ꢀ7.29 (m, 5H, SiPh), 3.74 (t, 6H, J = 5.6 Hz, CH2OH), 3.56 (t,
6H, J = 5.8 Hz, OCH2CH2CH2OH), 3.38 (t, 6H, J = 6.7 Hz, SiCH2CH2-
CH2O), 3.00 (br s, 3H, OH), 1.80 (quint, 6H, J = 5.7 Hz, OCH2CH2-
CH2OH), 1.66ꢀ1.51 (m, 6H, SiCH2CH2CH2O), 0.86ꢀ0.77 (m, 6H,
SiCH2); 13C NMR (50 MHz, CDCl3): d 136.32, 133.86, 128.97,
127.76, 73.76, 69.63, 61.54, 31.90, 23.77, 8.06; MS (FAB): calcd
for [M+H]+: 457.3, found: m/z 457.1; Calcd for [M+Na]+: 479.3,
found: m/z 479.0.
Ball(0)4-ether-All (250 mg, 0.756 mmol) was subjected to
hydroxylation in the same manner as that described for 24 to give
Ball(0)4-ether-OH (256 mg, 87.7%) as a white solid: Rf 0.71 [5:1 (v/
v) CHCl3ꢀMeOH]; IR (KBr): 3339 (
mOꢀH), 2926 (mCꢀH), 2872 (mCꢀH),
1111 (
m
CꢀOꢀC) cmꢀ1 1H NMR (200 MHz, CDCl3): d 3.75 (t, 8H,
;
J = 5.6 Hz, CH2OH), 3.60 (t, 8H, J = 5.8 Hz, OCH2CH2CH2OH), 3.38
(t, 8H, J = 6.7 Hz, SiCH2CH2CH2O), 3.01 ((br s, 4H, OH), 1.82 (quint,
8H, J = 5.7 Hz, OCH2CH2CH2OH), 1.63ꢀ1.48 (m, 8H, SiCH2CH2-
CH2O), 0.58ꢀ0.49 (m, 8H, SiCH2); 13C NMR (50 MHz, CDCl3): d
73.93, 69.64, 61.55, 31.96, 23.88, 8.02; MS (FAB): calcd for
[M+H]+: 497.4, found: m/z 497.1; Calcd for [M+Na]+: 519.3, found:
m/z 519.1.
Anal. Calcd for C24H44O6Siꢄ0.5H2O: C, 61.90; H, 9.74. Found: C,
61.85; H, 10.03.
4.1.6. Tris(7-methylsulfonyloxy-4-oxaheptyl)phenylsilane
(Fan(0)3-ether-OMs)
Anal. Calcd for C24H52O8Si: C, 58.03; H, 10.55. Found: C, 58.16;
H, 10.78.
To a solution of triol 24 (67 mg, 0.147 mmol) in pyridine
(0.7 mL) was added methanesulfonyl chloride (68 lL, 0.879 mmol)
at 0 °C, and the reaction mixture was stirred for 3 h at 0 °C. The
resulting mixture was diluted in CHCl3, successively washed twice
with 5% aq H2SO4, satd aq NaHCO3 and brine, dried over anhyd
MgSO4, filtered, and concentrated to afford mesylate (107 mg,
quantitative) as a yellow syrup. This compound was used for the
next step without further purification.: Rf 0.66 [10:1 (v/v)
4.1.10. Tetrakis(7-methylsulfonyloxy-4-oxaheptyl)silane
(Ball(0)4-ether-OMs)
Ball(0)4-ether-OH (74 mg, 0.149 mmol) was subjected to mesy-
lation in the same manner as that described for mesylate of 24 to
give Ball(0)4-ether-OMs (121 mg, quantitative) as a yellow syrup:
Rf 0.63 [10:1 (v/v) CHCl3ꢀMeOH]; IR (neat): 2932 (
m
CꢀH), 2866
CHCl3ꢀMeOH]; IR (neat): 2934 (
m
CꢀH), 2866 (
;
m
CꢀH), 1354 (
m
O@S@O),
(m
CꢀH), 1350 ( O@S@O), 1175 ( O@S@O), 1115 ( ;
m
m
m
CꢀOꢀC) cmꢀ1
1H
1175 ( O@S@O), 1113 (
m
m
CꢀOꢀC) cmꢀ1
1H NMR (200 MHz, CDCl3): d
NMR (200 MHz, CDCl3): d 4.34 (t, 8H, J = 6.3 Hz, CH2OMs), 3.52 (t,
8H, J = 6.0 Hz, OCH2CH2CH2OMs), 3.36 (t, 8H, J = 6.8 Hz, SiCH2CH2-
CH2O), 3.02 (s, 12H, OMs), 2.00 (quint, 8H, J = 6.1 Hz, OCH2CH2-
CH2OMs), 1.62ꢀ1.47 (m, 8H, SiCH2CH2CH2O), 0.57ꢀ0.49 (m, 8H,
SiCH2); 13C NMR (50 MHz, CDCl3): d 73.90, 67.39, 65.86, 37.09,
29.42, 23.88, 8.02.
7.50ꢀ7.28 (m, 5H, SiPh), 4.32 (t, 6H, J = 6.3 Hz, CH2OMs), 3.49 (t,
6H, J = 6.0 Hz, OCH2CH2CH2OMs), 3.37 (t, 6H, J = 6.8 Hz, SiCH2CH2-
CH2O), 2.99 (s, 9H, OMs), 1.98 (quint, 6H, J = 6.1 Hz, OCH2CH2-
CH2OMs), 1.65ꢀ1.50 (m, 6H, SiCH2CH2CH2O), 0.85ꢀ0.77 (m, 6H,
SiCH2); 13C NMR (50 MHz, CDCl3): d 136.41, 133.90, 129.00,
127.75, 73.70, 67.39, 65.83, 37.06, 29.41, 23.83, 8.14.
4.1.11. Tetrakis(7-bromo-4-oxaheptyl)silane (33, Ball(0)4-
ether-Br)
Ball(0)4-ether-OMs (121 mg, 0.149 mmol) was subjected to
bromination in the same manner as that described for 30 to give
Ball(0)4-ether-Br 33 (84 mg, 75.7%) as a colorless syrup: Rf 0.39
4.1.7. Tris(7-bromo-4-oxaheptyl)phenylsilane (30, Fan(0)3-
ether-Br)
NaBr (227 mg, 2.205 mmol) was added to a solution of mesylate
(107 mg, 0.147 mmol) in DMF (1 mL), and the mixture was stirred
for 3 h at 60 °C. The resulting mixture was diluted in CHCl3, succes-
sively washed with water and brine, dried over anhyd MgSO4, fil-
tered, and concentrated. The syrupy residue was purified by flash
column chromatography on silica gel [20:1 (v/v) n-hexaneꢀEtOAc]
to give bromide 30 (74 mg, 77.9%) as a colorless syrup: Rf 0.39 [5:1
[4:1 (v/v) n-hexaneꢀEtOAc]; IR (neat): 2930 (
mCꢀH), 2860 (mCꢀH),
1113 (
m
CꢀOꢀC) cmꢀ1 1H NMR (200 MHz, CDCl3): d 3.53 (t, 8H,
;
J = 5.9 Hz, OCH2CH2CH2Br), 3.51 (t, 8H, J = 6.5 Hz, CH2Br), 3.37 (t,
8H, J = 6.9 Hz, SiCH2CH2CH2O), 2.09 (quint, 8H, J = 6.2 Hz,
OCH2CH2CH2Br), 1.64ꢀ1.48 (m, 8H, SiCH2CH2CH2O), 0.58ꢀ0.50
(m, 8H, SiCH2); 13C NMR (50 MHz, CDCl3): d 73.94, 67.95, 32.83,
30.74, 23.91, 8.04; MS (FAB): calcd for [M+H]+ 749, found: m/z
748.5.
(v/v) n-hexaneꢀEtOAc]; IR (neat): 2930 (
mCꢀH), 2862 (mCꢀH), 1110
(m
CꢀOꢀC) cmꢀ1
;
1H NMR (200 MHz, CDCl3): d 7.51ꢀ7.33 (m, 5H,
SiPh), 3.50 (t, 12H, J = 6.4 Hz, OCH2CH2CH2Br), 3.37 (t, J = 6.8 Hz,
6H, SiCH2CH2CH2O), 2.07 (quint, 6H, J = 6.1 Hz, OCH2CH2CH2Br),
1.66ꢀ1.51 (m, 6H, SiCH2CH2), 0.86ꢀ0.77 (m, 6H, SiCH2); 13C NMR
(50 MHz, CDCl3): d 136.54, 133.97, 128.97, 127.77, 73.75, 67.93,
32.84, 30.74, 23.85, 8.16; MS (FAB): calcd for [M+H]+: 643.1, found:
m/z 642.7.
Anal. Calcd for C24H48O4SiBr4: C, 38.52; H, 6.47. Found: C, 38.90;
H, 6.50.
4.1.12. Bis[tris(4-oxa-6-heptenyl)silylpropyl]dimethylsilane
(Dumbbell(1)6-ether-All)
Anal. Calcd for C24H41O3SiBr3: C, 44.67; H, 6.40. Found: C, 44.90;
H, 6.45.
Dumbbell(1)6-OH 26 (1.46 g, 2.60 mmol) was subjected to ally-
lation in the same manner as that described for 23 to afford Dumb-
bell(1)6-ether-All (1.87 g, 89.3%) as a colorless liquid: Rf 0.41 [5:1
4.1.8. Tetrakis(4-oxa-6-heptenyl)silane (Ball(0)4-ether-All)
Ball(0)4-OH 25 (200 mg, 0.756 mmol) was subjected to allyla-
tion in the same manner as that described for 23 to give
Ball(0)4-ether-All (263 mg, 81.9%) as a colorless liquid: Rf 0.46
(v/v) n-hexaneꢀEtOAc]; IR (neat): 2913 (
mCꢀH), 2866 (mCꢀH), 1103
(m
CꢀOꢀC) cmꢀ1
;
1H NMR (200 MHz, CDCl3): d 6.02ꢀ5.82 (m, 6H,
CH@CH2), 5.32ꢀ5.13 (m, 12H, CH@CH2), 3.98ꢀ3.94 (m, 12H,
CH2CH@CH2), 3.37 (t, 12H, J = 7.1 Hz, SiCH2CH2CH2O), 1.65ꢀ1.50

5458
J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
(m, 12H, SiCH2CH2CH2O), 1.38ꢀ1.22 (m, 4H, SiCH2CH2CH2Si),
0.63ꢀ0.48 (m, 20H, SiCH2), ꢀ0.07 (s, 6H, SiCH3); 13C NMR
(50 MHz, CDCl3): d 134.35, 116.61, 73.34, 71.66, 24.02, 20.29,
18.24, 16.99, 8.14, ꢀ3.30; MS (FAB): calcd for [M+H]+: 793.1,
found: m/z 793.1.
(
m
CꢀOꢀC) cmꢀ1
;
1H NMR (400 MHz, CDCl3): d 5.96ꢀ5.86 (m, 12H,
CH@CH2), 5.29ꢀ5.15 (m, 24H, CH@CH2), 3.96 (d, 24H, J = 5.6 Hz,
CH2CH@CH2), 3.37 (t, 24H, J = 6.9 Hz, SiCH2CH2CH2O), 1.61ꢀ1.50
(m, 8H, SiCH2CH2CH2O), 1.30ꢀ1.21 (m, 8H, SiCH2CH2CH2Si),
0.66ꢀ0.50 (m, 40H, SiCH2); 13C NMR (100 MHz, CDCl3):
d
Anal. Calcd for C44H84O6Si3: C, 66.61; H, 10.67. Found: C, 66.44;
H, 10.95.
135.903, 116.61, 73.40, 71.72, 24.10, 18.43, 17.64, 17.47, 8.22
(SiC4); HRMS (ESI): calcd for [M+2Na]2+/2: 771.51138, found: m/z
771.5130.
4.1.13. Bis[tris(7-hydroxy-4-oxaheptyl)silylpropyl]dimethylsi-
lane (Dumbbell(1)6-ether-OH)
Anal. Calcd for C84H156O12Si5: C, 67.33; H, 10.49. Found: C,
67.25; H, 10.57.
Dumbbell(1)6-ether-All (200 mg, 0.271 mmol) was subjected to
hydroxylation in the same manner as that described for 24 to give
Dumbbell(1)6-ether-OH (185 mg, 80.8%) as a yellow syrup: Rf 0.58
4.1.17. Tetrakis[tris(7-hydroxy-4-oxaheptyl)silylpropyl]silane
(Ball(1)12-ether-OH)
[5:1 (v/v) CHCl3ꢀMeOH]; IR (neat): 3381 (
m
OꢀH), 2926 (
m
CꢀH), 2868
Ball(1)12-ether-All (280 mg, 0.187 mmol) was subjected to
hydroxylation in the same manner as that described for 24 to give
Ball(1)12-ether-OH (210 mg, 65.6%) as a yellow solid: Rf 0.34 [5:1
(m
CꢀH), 1115 (
m
CꢀOꢀC) cmꢀ1 1H NMR (200 MHz, CDCl3): d 3.74 (t,
;
J = 5.8 Hz, 12H, CH2OH), 3.59 (t, 12H, J = 5.9 Hz, OCH2CH2CH2OH),
3.38 (t, 12H, J = 6.8 Hz, SiCH2CH2CH2O), 3.07 ((br s, 6H, OH), 1.82
(quint, 12H, J = 5.8 Hz, OCH2CH2CH2OH), 1.63ꢀ1.47 (m, 12H,
SiCH2CH2CH2O), 1.38ꢀ1.22 (m, 4H, SiCH2CH2CH2Si), 0.63ꢀ0.47
(m, 20H, SiCH2), ꢀ0.06 (s, 6H, SiCH3); 13C NMR (50 MHz, CDCl3):
d 74.00, 69.43, 61.30, 31.94, 23.87, 20.16, 18.20, 16.85, 8.02,
ꢀ3.32; MS (FAB): calcd for [M+H]+: 901.6, found: 901.3; calcd for
[M+Na]+: 923.6, found: m/z 923.3.
(v/v) CHCl3ꢀMeOH]; IR (KBr): 3389 (
mOꢀH), 2926 (mCꢀH), 2868
(m
CꢀH), 1115 (
m
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz, CDCl3): d 3.74 (t,
;
24H, J = 5.7 Hz, CH2OH), 3.59 (t, 24H, J = 5.9 Hz, OCH2CH2CH2OH),
3.39 (t, 24H, J = 6.8 Hz, SiCH2CH2CH2O), 3.17 ((br s, 12H, OH),
1.82 (quint, 24H, J = 5.8 Hz, OCH2CH2CH2OH), 1.59ꢀ1.52 (m, 24H,
SiCH2CH2CH2O), 1.32ꢀ1.23 (m, 8H, SiCH2CH2CH2Si), 0.62ꢀ0.50
(m, 40H, SiCH2); 13C NMR (100 MHz, CDCl3): d 74.07, 69.49,
61.31, 32.09, 24.03, 18.47, 17.60, 17.34, 8.19; HRMS (ESI): calcd
for [M+2Na]2+/2: 879.57477, found: m/z 879.5757.
Anal. Calcd for C44H96O12Si3ꢄ1.5H2O: C, 56.92; H, 10.75. Found:
C, 57.02; H, 10.95.
Anal. Calcd for C84H180O24Si5ꢄ2H2O: C, 57.63; H, 10.59. Found: C,
4.1.14. Bis[tris(7-methylsulfonyloxy-4-oxaheptyl)silylpropyl]-
dimethylsilane (Dumbbell(1)6-ether-OMs)
57.48; H, 10.45.
Dumbbell(1)6-ether-OH (224 mg, 0.265 mmol) was subjected
to mesylation in the same manner as that described for mesylate
of 24 to give Dumbbell(1)6-ether-OMs (363 mg, quantitative) as
a yellow syrup: Rf 0.63 [10:1 (v/v) CHCl3ꢀMeOH]; IR (neat):
4.1.18. Tetrakis[tris(7-methylsulfonyloxy-4-
oxaheptyl)silylpropyl]silane (Ball(1)12-ether-OMs)
Ball(1)12-ether-OH (150 mg, 0.087 mmol) was subjected to
mesylation in the same manner as that described for mesylate of
24 to give Ball(1)12-ether-OMs (225 mg, 97.0%) as a yellow syrup:
2928 (
m
CꢀH), 2868 (
m
CꢀH), 1354 (
m
O@S@O), 1175 (mO@S@O), 1116
(mCꢀOꢀC
)
cmꢀ1 1H NMR (200 MHz, CDCl3):
;
d
4.33 (t, 12H,
Rf 0.48 [10:1 (v/v) CHCl3ꢀMeOH]; IR (neat): 2928 (
m
CꢀH), 2868
J = 6.2 Hz, CH2OMs), 3.52 (t, 12H, J = 6.0 Hz, OCH2CH2CH2OMs),
3.37 (t, 12H, J = 6.8 Hz, SiCH2CH2CH2O), 3.01 (s, 18H, OMs), 2.00
(quint, 12H, J = 6.1 Hz, OCH2CH2CH2OMs), 1.62ꢀ1.46 (m, 6H,
SiCH2CH2CH2O), 1.38ꢀ1.26 (m, 4H, SiCH2CH2CH2Si). 0.64ꢀ0.47
(m, 20H, SiCH2), ꢀ0.05 (s, 6H, SiCH3); 13C NMR (50 MHz, CDCl3):
d 73.83, 67.35, 65.72, 36.91, 29.28, 23.81, 20.13, 18.09, 16.80,
7.97, ꢀ3.44.
(m
CꢀH), 1356 ( O@S@O), 1177 ( O@S@O), 1117 ( ;
m
m
m
CꢀOꢀC) cmꢀ1
1H
NMR (400 MHz, CDCl3): d 4.33 (t, 24H, J = 6.3 Hz, CH2OMs), 3.52
(t, 24H, J = 6.0 Hz, OCH2CH2CH2Ms), 3.37 (t, 24H, J = 6.9 Hz,
SiCH2CH2CH2O), 3.02 (s, 36H, OMs), 2.00 (quint, 24H, J = 6.1 Hz,
OCH2CH2CH2OMs),
1.58ꢀ1.50
(m,
24H,
SiCH2CH2CH2O),
1.29ꢀ1.21 (m, 8H, SiCH2CH2CH2Si), 0.61ꢀ0.49 (m, 40H, SiCH2);
13C NMR (100 MHz, CDCl3): d 74.07, 67.46, 66.02, 37.17, 29.53,
24.05, 18.43, 17.66, 17.51, 8.15; HRMS (ESI): calcd for [M+2Na]2+
/
4.1.15. Bis[tris(7-bromo-4-oxaheptyl)silylpropyl]dimethyl-
silane (36, Dumbbell(1)6-ether-Br)
2: 1347.44007, found: m/z 1347.4434.
Dumbbell(1)6-ether-OMs (363 mg, 0.265 mmol) was subjected
to bromination in the same manner as that described for 30 to af-
ford Dumbbell(1)6-ether-Br (197 mg, 58.1%) as a colorless syrup:
4.1.19. Tetrakis[tris(7-bromo-4-oxaheptyl)silylpropyl]silane
(39, Ball(1)12-ether-Br)
Ball(1)12-ether-OMs (225 mg, 0.085 mmol) was subjected to
bromination in the same manner as that described for 30 to give
Ball(1)12-ether-Br (129 mg, 61.4%) as a colorless syrup: Rf 0.63
Rf 0.43 [4:1 (v/v) n-hexaneꢀEtOAc]; IR (neat): 2910 (
mCꢀH), 2862
(m
CꢀH), 1113 (
m
CꢀOꢀC) cmꢀ1 1H NMR (200 MHz, CDCl3): d 3.54 (t,
;
12H, J = 5.9 Hz, OCH2CH2CH2Br), 3.51 (t, 12H, J = 6.6 Hz, CH2Br),
3.38 (t, 12H, J = 6.9 Hz, SiCH2CH2CH2O), 2.10 (quint, 12H,
J = 6.2 Hz, OCH2CH2CH2Br), 1.63ꢀ1.48 (m, 12H, SiCH2CH2CH2O),
1.39ꢀ1.22 (m, 4H, SiCH2CH2CH2Si), 0.64ꢀ0.47 (m, 20H, SiCH2),
ꢀ0.06 (s, 6H, SiCH3); 13C NMR (50 MHz, CDCl3): d 74.07, 67.97,
32.89, 30.72, 24.00, 20.34, 18.30, 17.03, 8.19 , ꢀ3.25; MS (MALDI-
TOF) Calcd for [M+Na]+: 1301.099, found: m/z 1301.119; Calcd
for [M+K]+: 1315.074, found: m/z 1315.092.
[2:1 (v/v) n-hexaneꢀEtOAc]; IR (neat): 2916 (
mCꢀH), 2862 (mCꢀH),
1111 (
m
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz, CDCl3): d 3.54 (t, 24H,
;
J = 5.9 Hz, OCH2CH2CH2Br), 3.51 (t, 24H, J = 6.5 Hz, CH2Br), 3.38 (t,
24H, J = 6.9 Hz, SiCH2CH2CH2O), 2.10 (quint, 24H, J = 6.2 Hz,
OCH2CH2CH2Br), 1.60ꢀ1.52 (m, 24H, SiCH2CH2CH2O), 1.31ꢀ1.23
(m, 24H, SiCH2CH2CH2Si), 0.62ꢀ0.50 (m, 40H, SiCH2); 13C NMR
(100 MHz, CDCl3): d 74.08, 68.03, 32.92, 30.75, 24.08, 18.47,
17.66, 17.53, 8.22; HRMS (ESI): calcd for [M+2Na]2+/2:
1258.0630, found: m/z 1258.0664.
Anal. Calcd for C44H90O6Si3Br6: C, 41.32; H, 7.09. Found: C,
41.69; H, 7.16.
Anal. Calcd for C84H168Br12O12Si5: C, 40.85; H, 6.86. Found: C,
41.16; H, 6.83.
4.1.16. Tetrakis[tris(4-oxa-6-heptenyl)silylpropyl]silane
(Ball(1)12-ether-All)
Ball(1)12-OH 27 (500 mg, 0.492 mmol) was subjected to allyla-
tion in the same manner as that described for 23 to give Ball(1)12-
ether-All (429 mg, 58.3%) as a colorless liquid: Rf 0.58 [3:1 (v/v)
4.1.20. Bis[tris(3-azidepropyl)silylpropyl]dimethylsilane (28,
Dumbbell(1)6-N3)
NaN3 (706 mg, 10.862 mmol) was added to a solution of Dumb-
bell(1)6-ether-OMs (311 mg, 0.361 mmol) in DMF (5 mL), and the
mixture was stirred for 1 h at 80 °C. The resulting mixture was
n-hexaneꢀEtOAc]. IR (neat): 2916
(mCꢀH), 2855 (mCꢀH), 1103

J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
5459
poured into ice water and extracted with CHCl3. The extract was
successively washed with water and brine, dried over anhyd
MgSO4, filtered, and concentrated. The syrupy residue was purified
by flash column chromatography on silica gel [15:1 (v/v) n-hex-
aneꢀEtOAc] to give hexaazide 28 (211 mg, 83.1%) as a colorless
(dd, 3H, H-6), 3.80 (s, 9H, OMe), 2.77ꢀ2.71 (m, 3H, SCHa), 2.72
(dd, 3H, J3ax,3eq = 12.7 Hz, H-3eq), 2.57ꢀ2.50 (m, 3H, SCHb), 2.50
(t, 6H, J = 7.2 Hz, CH2SCH2), 2.44 (t, 6H, J = 7.2 Hz, CH2SCH2), 2.16,
2.14, 2.04, 2.03 (4 ꢂ s, 36H, OAc), 1.98 (t, 3H, H-3ax), 1.87 (s, 9H,
NAc), 1.61ꢀ1.41 (2 ꢂ m, 24H, SCH2CH2CH2CH2CH2SCH2CH2),
0.93ꢀ0.88 (m, 6H, SiCH2); 13C NMR (100 MHz, CDCl3): d 170.89,
170.57, 170.13, 170.07, 169.97, 168.44, 136.28, 133.98, 129.10,
127.86, 83.12, 74.08, 69.66, 68.62, 67.28, 62.12, 52.89, 49.35,
38.05, 35.86, 31.89, 29.24, 28.82, 28.69, 28.03, 24.03, 23.15,
21.15, 20.82, 20.75, 11.89; HRMS (ESI): calcd for [M+H]+:
2056.71535, found: m/z 2056.72008.
syrup: Rf 0.39 [10:1 (v/v) n-hexaneꢀEtOAc]; IR (neat): 2922 (
mCꢀH),
2874 (m (m
CꢀH), 2097 cmꢀ1 N@N@N); 1H NMR (400 MHz, CDCl3): d
3.26 (t, 12H, J = 6.8 Hz, CH2N3), 1.62ꢀ1.54 (m, 12H, SiCH2CH2-
CH2N3), 1.36ꢀ1.28 (m, 4H, SiCH2CH2CH2Si), 0.67ꢀ0.55 (m, 20H,
SiCH2), ꢀ0.03 (s, 6H, SiCH3); 13C NMR (50 MHz, CDCl3): d 54.44,
23.57, 20.25, 18.28, 16.74, 9.38, ꢀ3.30.
Anal. Calcd for C26H54N18Si3: C, 44.42; H, 7.74; N, 35.86. Found:
C, 44.70; H, 7.88; N, 35.96.
Anal. Calcd for C90H137N3O36S6Si: C, 52.54; H, 6.71; N, 2.04.
Found: C, 52.33; H, 6.75; N, 2.00.
A disulfide dimer 53 (35 mg) as a white foam was obtained as a
4.1.21. Bis{tris[3-(6-bromohexanoylamino)propyl]silylpropyl}-
dimethylsilane (37, Dumbbell(1)6-amide-Br)
byproduct; Rf 0.39 [5:4:1 (v/v/v) CHCl3ꢀEtOAcꢀMeOH]; ½a D28
ꢁ
+28.1
(c 1.13, CHCl3); IR (KBr): 2936 (
m
CꢀH), 1740 (
m
C@O), 1665 (mC@O
CꢀOꢀC) cmꢀ1
5.37 (ddd, 2H, J7,8 = 8.6 Hz,
,
To a solution of Dumbbell(1)6-N3 28 (276 mg, 0.393 mmol) in
6:1(v/v) THF–water (7 mL) was added triphenylphosphine
(680 mg, 2.593 mmol) at 0 °C. The reaction mixture was stirred
overnight at room temperature and was concentrated in vacuo.
The residue was suspended in CHCl3 and extracted with 1 M aq
HCl. Concentration of the aqueous extract gave white foamy amine
hydrochloride (300 mg).
amide I), 1541 (dNꢀH, amide II), 1227 (
m
CꢀO), 1038 (
m
;
1H NMR (400 MHz, CDCl3):
d
J8,9a = 2.1 Hz, J8,9b = 4.6 Hz, H-8), 5.33 (dd, 2H, J6,7 = 2.1 Hz, H-7),
5.18 (d, 2H, J5,NH = 10.4 Hz, NH), 4.86 (ddd, 2H, J4,5 = 10.4 Hz,
J3ax,4 = 12.3 Hz, J3eq,4 = 4.6 Hz, H-4), 4.31 (dd, 2H, J9a,9b = 12.6 Hz,
H-9a), 4.11 (dd, 2H, H-9b), 4.05 (q, 2H, J5,6 = 10.4 Hz, H-5), 3.83
(dd, 2H, H-6), 3.81 (s, 6H, OMe), 2.78ꢀ2.71 (m, 2H, SCHa), 2.72
(dd, 2H, J3ax,3eq = 12.6 Hz, H-3eq), 2.67 (t, 4H, J = 7.2 Hz, CH2S2CH2),
2.58ꢀ2.51 (m, 2H, SCHb), 2.17, 2.14, 2.04, 2.03 (4 ꢂ s, 24H, OAc),
1.98 (t, 2H, H-3ax), 1.88 (s, 6H, NAc), 1.69ꢀ1.59 (m, 8H,
SCH2CH2CH2CH2CH2S), 1.51ꢀ1.43 (m, 4H, SCH2CH2CH2); 13C NMR
The amine hydrochloride (300 mg) was dissolved in MeOH
(6 mL) with Et3N (723 lL), and 6-bromohexanoyl choloride was
added dropwise to the mixture at ꢀ20 °C. The reaction mixture
was stirred at ꢀ20 °C for 1 h and then diluted with CHCl3 and
poured into ice water. The organic layer was separated and washed
with satd aq NaHCO3 and brine, dried over anhyd MgSO4, filtered,
and concentrated. The residual syrup was purified by flash column
chromatography on silica gel [15:4:1 (v/v/v) CHCl3ꢀEtOAcꢀMeOH]
to give bromide 37 (627 mg, 99.2%) as a colorless syrup: Rf 0.53
[5:4:1 (v/v) CHCl3ꢀEtOAcꢀMeOH]; 1H NMR (400 MHz, CDCl3): d
6.45 (t, 12H, J = 5.6 Hz, NH), 3.42 (t, 24H, CH2Br), 3.19 (br q, 12H,
J = 6.6 Hz, CH2N), 2.22 (t, 12H, J = 7.5 Hz, CH2CO), 1.88 (quint,
12H, J = 7.1 Hz, CH2CH2Br), 1.67 (quint, 12H, J = 7.6 Hz, CH2CH2CO),
1.48 (m, 24H, CH2CH2CH2Br, CH2CH2N), 1.32ꢀ1.22 (m, 4H,
SiCH2CH2), 0.59ꢀ0.48 (m, 20H, SiCH2), ꢀ0.06 (s, 6H, SiCH3); 13C
NMR (100 MHz, CDCl3): d 173.05, 42.64, 36.34, 33.70, 32.40,
27.76, 24.86, 24.05, 20.17, 18.43, 16.96, 9.39, ꢀ3.00; MS (MALDI-
TOF): calcd for [M+Na]+: 1633.351, found: m/z 1633.398; Calcd
for [M+K]+: 1649.325, found: m/z 1649.370.
(100 MHz, CDCl3):
d 170.88, 170.57, 170.13, 170.07, 169.99,
168.45, 83.11, 74.08, 69.66, 68.63, 67.29, 62.14, 52.89, 49.33,
38.66, 38.04, 28.82, 28.67, 27.58, 23.14, 21.14, 20.81, 20.74; MS
(FAB): calcd for [M+H]+ 1217.4, found: m/z 1218.0.
Anal. Calcd for C50H76N2O24S4: C, 49.33; H, 6.29; N, 2.30. Found:
C, 49.01; H, 6.27; N, 2.24.
4.1.23. Fan(0)3-s-neu5ac3 (3)
Solution of NaOMe (0.5 M) in MeOH was added to a solution of
fully protected Fan(0)3-S-Neu5Ac3(OAc, OMe) 41 (36.6 mg,
17.79 lmol) in MeOH (1 mL) at 0 °C. The reaction mixture was stir-
red for 3 h at room temperature, and then 0.5 M aq solution of
NaOH was added and reaction mixture was stirred overnight at
room temperature. The solution was treated with IR-120B (H+)
and adjusted to pH ꢃ4, and then the suspension was filtered. The
resin was washed with water, and the filtrate and washings were
combined and concentrated. The resulting mixture was purified
by size exclusion column chromatography on Sephadex G-25 with
5% aq AcOH as an eluent to give de-protected Fan(0)3-Neu5Ac 3
(23.5 mg, 87.4%) as a white powder after lyophilization: Rf 0.63
Anal. Calcd for C62H120N6O6Br6Si3: C, 46.27; H, 7.52; N, 5.22.
Found: C, 45.89; H, 7.47; N, 4.89.
4.1.22. Fan(0)3-S-Neu5Ac3(OAc, OMe) (41)
NaOMe (17.5 mg, 0.325 mmol) was added to a solution of thios-
ialoside 21 (192 mg, 0.295 mmol) and Fan(0)3-Br dendrimer 29
(23 mg, 0.049 mmol) in 1:1 (v/v) DMFꢀMeOH (0.8 mL) at 0 °C.
The reaction mixture was stirred overnight at room temperature,
and then the reaction was quenched by addition of acetic acid
[3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D29
ꢁ
+27.7 (c 1.03, H2O); IR
C@O), 1636 ( C@O, amide
CꢀO), 1032 (
CꢀOꢀC) cmꢀ1 1H
(KBr): 3420 (
m
OꢀH), 2924 (
m
CꢀH), 1697 (
m
m
I), 1557 (dNꢀH, amide II), 1271 (
m
m
;
NMR (400 MHz, D2O): d 7.38, 7.17 (2 ꢂ br s, 5H, SiPh), 3.90ꢀ3.62
(m, 21H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4), 2.82 (br s, 6H, SCHa
and H-3eq), 2.66 (br s, 3H, SCHb), 2.41 (br s, 12H, CH2SCH2), 2.04 (s,
9H, NAc), 1.88 (br s, 3H, H-3ax), 1.51, 1.41 (2 ꢂ br s, 24H,
SCH2CH2CH2CH2CH2SCH2CH2), 0.84 (br s, 6H, SiCH2); 13C NMR
(100 MHz, D2O): d 174.66, 172.03, 133.70, 127.61, 83.44, 74.68,
71.00, 67.76, 62.25, 51.45, 40.42, 35.29, 31.35, 29.06, 28.87,
28.45, 28.14, 27.81, 23.67, 21.82, 11.33; HRMS (ESI): calcd for
[M+H]+: 1510.54162, found: m/z 1510.53494.
(50 lL). After removal of MeOH by evaporation, the residual mix-
ture was treated with acetic anhydride (1.5 mL) in pyridine
(2 mL) at room temperature overnight. After co-evaporation with
toluene in vacuo to dryness, the residue was treated with ethereal
CH2N2. Purification of the concentrated crude product by column
chromatography on silica gel [40:1 (v/v) CHCl3ꢀMeOH] afforded
41 (81 mg, 80.2%) as
a
ꢁ
white foam: Rf 0.28 [5:4:1 (v/v/v)
+26.2 (c 1.10, CHCl3); IR (KBr): 2932
CHCl3ꢀEtOAcꢀMeOH]; ½a D33
(m
CꢀH), 1740 (mC@O), 1663 (
m
C@O, amide I), 1545 (dNꢀH, amide II),
1229 (
m
CꢀO), 1038 (
m
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz, CDCl3): d
;
4.1.24. Bis[n-pentenyl 5-acetamido-3,5-dideoxy-2-thio-
glycero- -galacto-2-nonulopyranosidyl]disulfide (2)
The de-protection of disulfide dimer (OAc, OMe) 53 (21.1 mg,
4.90 mol) was carried out by a similar method similar to that
a-D-
7.47ꢀ7.33 (2 ꢂ m, 5H, SiPh), 5.36 (ddd, 3H, J7,8 = 8.4 Hz,
J8,9a = 2.3 Hz, J8,9b = 4.6 Hz, H-8), 5.33 (dd, 3H, J6,7 = 1.8 Hz, H-7),
5.25 (d, 3H, J5,NH = 10.4 Hz, NH), 4.86 (ddd, 3H, J4,5 = 10.4 Hz,
J3ax,4 = 12.2 Hz, J3eq,4 = 4.8 Hz, H-4), 4.31 (dd, 3H, J9a,9b = 12.5 Hz,
H-9a), 4.11 (dd, 3H, H-9b), 4.05 (m, 3H, J5,6 = 10.4 Hz, H-5), 3.83
D
l
described for the preparation of 1 to give de-protected disulfide
dimer 2 (16.2 mg, quantitative) as a white powder after lyophiliza-

5460
J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
tion: Rf 0.34 [8:6:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D30
ꢁ
+25.2 (c 1.03,
C@O), 1636 (mC@O
CꢀO), 1034 (
CꢀOꢀC) cmꢀ1
carried out by a method similar to that described for the prepara-
tion of 41 to give Fan(0)3-amide-S-Neu5Ac3(OAc, OMe) 43 (95 mg,
80.5%) as a white foam: Rf 0.33 [5:4:1 (v/v/v) CHCl3ꢀEtOAcꢀMe-
H2O); IR (KBr): 3420 (
m
OꢀH), 2930 (
m
CꢀH), 1705 (
m
,
;
amide I), 1558 (dNꢀH, amide II), 1273 (
m
m
1H NMR (400 MHz, D2O): d 3.90ꢀ3.59 (m, 14H, H-9a, H-9b, H-8,
H-7, H-6, H-5, H-4), 2.81 (dd, 2H, J3eq,4 = 4.8 Hz, J3ax,3eq = 12.9 Hz,
H-3eq), 2.83ꢀ2.78 (m, 2H, SCHa), 2.73ꢀ2.66 (m, 2H, SCHb), 2.02
(s, 6H, NAc), 1.86 (t, 2H, J3ax,4 = 11.8 Hz, H-3ax), 1.69ꢀ1.59 (m,
8H, SCH2CH2CH2CH2CH2S), 1.51–1.43 (m, 4H, SCH2CH2CH2); 13C
NMR (100 MHz, D2O): d 174.74, 171.98, 83.53, 74.75, 71.10,
67.97, 67.65, 62.43, 51.53, 40.27, 38.23, 28.85, 28.37, 28.09,
26.99, 21.82; MS (FAB): calcd for [M+H]+: 853.3, found: 853.0;
Calcd for [M+Na]+: 875.2, found: m/z 874.9.
OH]; ½a 3D0
ꢁ
+21.5 (c 1.11, CHCl3); IR (KBr): 3292 (
m
NꢀH), 2934 (
C@O, amide I), 1549 (dNꢀH, amide
CꢀO), 1036 (mCꢀOꢀC
cmꢀ1 1H NMR
mCꢀH),
2859 (
m
CꢀH), 1744 (mC@O), 1651 (m
II), 1261 (
m
CꢀN), 1227 (
m
)
;
(400 MHz, CDCl3): d 7.44ꢀ7.32 (m, 5H, SiPh), 5.91(t, 3H, J =
5.7 Hz, NHCH2), 5.37ꢀ5.32 (m, 9H, J8,9a = 2.1 Hz, J8,9b = 4.4 Hz,
J6,7 = 1.8 Hz, J5,NH = 10.4 Hz, H-8, H-7 and NH), 4.86 (ddd,
3H, J4,5 = 10.4 Hz, J3ax,4 = 12.3 Hz, J3eq,4 = 4.7 Hz, H-4), 4.31 (dd, 3H,
J9a,9b = 12.5 Hz, H-9a), 4.11 (dd, 3H, H-9b), 4.05 (q, 3H, J5,6
=
10.4 Hz, H-5), 3.83 (dd, 3H, H-6), 3.80 (s, 9H, OMe), 3.20
(q, 6H, J = 6.6 Hz, CH2N)), 2.77ꢀ2.70 (m, 3H, SCHa), 2.72 (dd, 3H,
J3ax,3eq = 12.3 Hz, H-3eq), 2.57ꢀ2.50 (m, 3H, SCHb), 2.484 (t, 6H,
J = 7.3 Hz, CH2SCH2), 2.478 (t, 6H, J = 7.3 Hz, CH2SCH2), 2.18 (t,
6H, J = 7.5 Hz, CH2CO), 2.16, 2.14, 2.04, 2.03 (4 ꢂ s, 36H, OAc),
1.98 (t, 3H, H-3ax), 1.88 (s, 9H, NAc), 1.67ꢀ1.36 (m, 42H,
SCH2CH2CH2CH2CH2SCH2CH2CH2CH2CH2CONHCH2CH2), 0.82ꢀ0.78
(m, 6H, CH2Si); 13C NMR (100 MHz, CDCl3): d 173.05, 170.87,
170.64, 170.19, 170.07, 170.01, 168.44, 136.04, 133.85, 129.27,
127.98, 83.10, 74.04, 69.66, 68.59, 67.27, 62.10, 52.88, 49.29,
42.36, 38.04, 36.54, 31.87, 29.29, 29.13, 28.80, 28.66, 28.49,
27.98, 25.32, 23.81, 23.14, 21.15, 20.82, 20.76, 9.13; HRMS (ESI):
calcd for [M+3Na]3+/3: 821.30913, found: m/z 821.3100.
Anal. Calcd for C32H56O16N2S4ꢄ1.4H2O: C, 43.76; H, 6.75; N, 3.19.
Found: C, 43.65; H, 6.64; N, 3.10.
4.1.25. Fan(0)3-ether-S-Neu5Ac3(OAc, OMe) (42)
The condensation reaction between thiosialoside 21 (327 mg,
0.502 mmol) and Fan(0)3-ether-Br 30 (44 mg, 0.068 mmol) was
carried out by
a method similar to that described for the
preparation of 41 to give Fan(0)3-ether-S-Neu5Ac3(OAc, OMe) 42
(85 mg, 55.9%) as
a white foam: Rf 0.38 [5:4:1 (v/v/v)
CHCl3ꢀEtOAcꢀMeOH]; ½a D27
ꢁ
+23.1 (c 1.11, CHCl3); IR (KBr): 2934
(
m
CꢀH), 2859 (
m
CꢀH), 1742 (
m
C@O), 1663 (
CꢀOꢀC), 1036 (
(400 MHz, CDCl3): d 7.48ꢀ7.30 (m, 5H, SiPh), 5.36 (ddd, 3H, J7,8
m
C@O, amide I), 1545 (dNꢀH
,
amide II), 1227 (
m
CꢀO), 1109 (
m
m
CꢀOꢀC) cmꢀ1; 1H NMR
=
8.4 Hz, J8,9a = 2.1 Hz, J8,9b = 4.6 Hz, H-8), 5.33 (dd, 3H, J6,7 = 1.9 Hz,
4.1.28. Fan(0)3-amide-S-Neu5Ac3 (5)
The de-protection of Fan(0)3-amide-S-Neu5Ac3(OAc, OMe) 43
(32.5 mg, 13.56 lmol) was carried out by a method similar to that
H-7), 5.29 (d, 3H, J5,NH = 10.4 Hz, NH), 4.86 (ddd, 3H, J4,5
10.4 Hz, J3ax,4 = 12.4 Hz, J3eq,4 = 4.4 Hz, H-4), 4.31 (dd, 3H, J9a,9b
=
=
12.3 Hz, H-9a), 4.11 (dd, 3H, H-9b), 4.05 (q, 3H, J5,6 = 10.4 Hz, H-
5), 3.83 (dd, 3H, H-6), 3.80 (s, 9H, OMe), 3.45 (t, 6H, J = 6.3 Hz,
OCH2CH2CH2S), 3.35 (t, 6H, J = 6.9 Hz, SiCH2CH2CH2O), 2.77ꢀ2.71
(m, 3H, SCHa), 2.71 (dd, 3H, J3ax,3eq = 12.7 Hz, H-3eq), 2.57ꢀ2.50
(m, 3H, SCHb), 2.56 (t, 6H, J = 7.3 Hz, CH2SCH2), 2.49 (t, 6H,
J = 7.3 Hz, CH2SCH2), 2.16, 2.14, 2.04, 2.03 (4 ꢂ s, 36H, OAc), 1.98
(t, 3H, H-3ax), 1.88 (s, 9H, NAc), 1.83 (quint, 6H, OCH2CH2CH2S),
1.61ꢀ1.42 (m, 24H, SCH2CH2CH2CH2CH2SCH2CH2), 0.82ꢀ0.77 (m,
6H, Si CH2); 13C NMR (100 MHz, CDCl3): d 170.87, 170.55, 170.14,
170.07, 169.97, 168.46, 136.65, 134.01, 128.94, 127.75, 83.13,
74.10, 73.75, 69.66, 69.25, 68.70, 67.34, 62.14, 52.86, 49.37,
38.06, 35.86, 31.88, 29.79, 29.13, 28.82, 28.69, 28.01, 23.92,
23.13, 21.12, 20.79, 20.72, 8.24; HRMS (ESI): calcd for [M+Na]+
2252.82289, found: m/z 2252.82345.
described for the preparation of 3 to give Fan(0)3-amide-S-
Neu5Ac3 5 (25.1 mg, quantitative) as a white powder after lyophi-
lization: Rf 0.61 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D25
ꢁ
+23.0 (c
1.09, H2O); IR (KBr): 3420 (mOꢀH), 2924 (mCꢀH), 1697 (mC@O), 1636
(mC@O, amide I), 1557 (dNꢀH, amide II), 1271 (mCꢀO), 1032 (mCꢀOꢀC)
cmꢀ1
;
1H NMR (400 MHz, D2O): d 7.40, 7.30 (2 ꢂ br s, 5H, SiPh),
3.84ꢀ3.61 (m, 21H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4), 3.12 (br
s, 6H, CH2N), 2.78 (br s, 6H, SCHa and H-3eq), 2.63 (br s, 3H, SCHb),
2.41 (br s, 12H, CH2SCH2), 2.13 (br s, 6H, CH2CO), 2.03 (s, 9H, NAc),
1.84 (br s, 3H, H-3ax), 1.49, 1.39 (2 ꢂ br s, 42H, SCH2CH2CH2CH2
CH2SCH2CH2CH2CH2CH2CONHCH2CH2), 0.83 (br s, 6H, SiCH2); 13C
NMR (100 MHz, CDCl3): d 175.11 (2 ꢂ C,), 172.36, 133.93, 128.18,
83.68, 75.17, 71.56, 68.16, 52.84, 52.07, 42.12, 40.80, 36.11,
31.79, 29.29, 28.87, 28.14, 25.81, 23.50, 22.32, 9.68; HRMS (ESI):
calcd for [M+2Na]2+/2: 947.38222, found: m/z 947.3817.
4.1.26. Fan(0)3-ether-S-Neu5Ac3 (4)
The de-protection of Fan(0)3-ether-S-Neu5Ac3(OAc, OMe) 42
4.1.29. Ball(0)4-S-Neu5Ac4(OAc, OMe) (44)
(32.6 mg, 14.61 lmol) was carried out by a method similar to that
The condensation reaction between thiosialoside 21 (273 mg,
0.419 mmol) and Ball(0)4-Br 32 (27 mg, 0.052 mmol) was carried
out by a method similar to that described for the preparation of
41 to give Ball(0)4-S-Neu5Ac4(OAc, OMe) 44 (103 mg, 75.2%) as a
described for the preparation of 3 to give Fan(0)3-ether-S-Neu5Ac3
4 (24.6 mg, quantitative) as a white powder after lyophilization: Rf
0.52 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D24
ꢁ
+25.4 (c 1.07, H2O);
CꢀH), 1700 ( C@O), 1634
(mC@O, amide I), 1558 (dNꢀH, amide II), 1275 (mCꢀO), 1110 (mCꢀOꢀC),
IR (KBr): 3414 (
m
O-H), 2930 (
m
CꢀH), 2857 (
m
m
white foam: Rf 0.13 [5:4:1 (v/v/v) CHCl3ꢀEtOAcꢀMeOH]; ½a D24
ꢁ
1032 (m ;
CꢀOꢀC) cmꢀ1
1H NMR (400 MHz, D2O): d 7.37, 7.18 (2 ꢂ br
+26.5 (c 1.19, CHCl3); IR (KBr): 2932 (mCꢀH), 1742 (mC@O), 1667
(m
C@O, amide I), 1549 (dNꢀH, amide II), 1229 (
mCꢀO), 1038 (mCꢀOꢀC)
s, 5H, SiPh), 3.82ꢀ3.58 (m, 21H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-
4), 3.35 (t, 6H, OCH2CH2CH2S), 3.25 (t, 6H, SiCH2CH2CH2O), 2.77 (br
s, 6H, SCHa and H-3eq), 2.62 (br s, 3H, SCHb), 2.45, 2.43 (2 ꢂ br s,
12H, CH2SCH2), 2.00 (s, 9H, NAc), 1.83 (br s, 3H, H-3ax), 1.71 (br s,
6H, OCH2CH2CH2S), 1.50, 1.39 (2 ꢂ br s, 24H, SCH2CH2CH2CH2CH2
SCH2CH2), 0.73 (br s, 6H, SiCH2); 13C NMR (100 MHz, D2O): d
174.78, 172.09, 136.48, 133.64, 127.58, 83.66, 74.78, 72.98, 71.12,
68.71, 67.93, 67.77, 62.38, 51.61, 40.49, 31.42, 29.39, 28.84, 28.48,
28.27, 27.69, 23.54, 21.88, 8.11; HRMS (ESI): calcd for [M+Na]+:
1706.64916, found: m/z 1706.65283.
cmꢀ1 1H NMR (400 MHz, CDCl3): d 5.36 (ddd, 3H, J7,8 = 8.4 Hz,
;
J8,9a = 2.1 Hz, J8,9b = 4.6 Hz, H-8), 5.33 (dd, 3H, J6,7 = 1.6 Hz, H-7),
5.25 (d, 4H, J5,NH = 10.3 Hz, NH), 4.86 (ddd, 4H, J4,5 = 10.3 Hz,
J3ax,4 = 12.1 Hz, J3eq,4 4.5 = Hz, H-4), 4.31 (dd, 4H, J9a,9b = 12.5 Hz,
H-9a), 4.11 (dd, 4H, H-9b), 4.05 (q, 4H, J5,6 = 10.3 Hz, H-5), 3.84
(dd, 4H, H-6), 3.81 (s, 12H, OMe), 2.78ꢀ2.71 (m, 4H, SCHa), 2.72
(dd, 4H, J3ax,3eq = 12.7 Hz, H-3eq), 2.58ꢀ2.53 (m, 4H, SCHb), 2.495
(t, 8H, J = 7.1 Hz, CH2SCH2), 2.487 (t, 8H, J = 7.2 Hz, CH2SCH2),
2.17, 2.14, 2.04, 2.03 (4 ꢂ s, 48H, OAc), 1.98 (t, 4H, H-3ax), 1.87
(s, 12H, NAc), 1.61ꢀ1.42 (m, 32H, SCH2CH2CH2CH2CH2SCH2CH2),
0.62 (m, 8H, SiCH2); 13C NMR (100 MHz, CDCl3):
d 170.89,
4.1.27. Fan(0)3-amide-S-Neu5Ac3(OAc, OMe) (43)
170.60, 170.15, 170.08, 169.98, 168.46, 83.14, 74.09, 69.67, 68.62,
67.28, 62.12, 52.91, 49.37, 38.06, 36.03, 32.03, 29.28, 28.86,
The condensation reaction between thiosialoside 20 (145 mg,
0.222 mmol) and Fan(0)3-amide-Br 31 (40 mg, 0.049 mmol) was

J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
5461
28.71, 28.07, 24.23, 23.16, 21.17, 20.83, 20.77, 12.00; HRMS (ESI):
4.1.33. Ball(0)4-amide-S-Neu5Ac4(OAc, OMe) (46)
calcd for [M+Na]+: 2651.88866, found: m/z 2651.87993.
The condensation reaction between thiosialoside 21 (203 mg,
0.311 mmol) and Ball(0)4-amide-Br 34 (42 mg, 0.043 mmol) was
carried out by a method similar to that described for the prepara-
tion of 41 to give Ball(0)4-amide-S-Neu5Ac4(OAc, OMe) 46
4.1.30. Ball(0)4-s-neu5ac4 (6)
The de-protection of Ball(0)4-S-Neu5Ac4(OAc, OMe) 44
(30.0 mg, 11.04
l
mol) was carried out by a method similar to that
(106 mg, 80.9%) as a white foam: Rf: 0.37 [7:2:1 (v/v/v)
described for the preparation of 3 to give Ball(0)4-S-Neu5Ac4
6
CHCl3ꢀEtOAcꢀMeOH]; ½a D27
ꢁ
+24.0 (c 1.15, CHCl3); IR (KBr): 3289
CꢀH), 1740 ( C@O), 1651 ( C@O, amide
CꢀN), 1227 ( CꢀO), 1036 (mCꢀOꢀC
;
1H NMR (400 MHz, CDCl3): d 6.12 (t, 4H, J = 5.7 Hz, NHCH2),
(21.6 mg, 99.5%) as a white powder after lyophilization: Rf 0.52
(
m
NꢀH), 2932 (
m
CꢀH), 2859 (
m
m
m
[3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D31
ꢁ
+23.2 (c 1.06, H2O); IR
C@O), 1636 ( C@O, amide
CꢀO), 1034 (
CꢀOꢀC) cmꢀ1 1H
I), 1549 (dNꢀH, amide II), 1261 (
m
m
)
(KBr): 3424 (mOꢀH), 2927 (
m
CꢀH), 1701 (
m
m
cmꢀ1
I), 1560 (dNꢀH, amide II), 1279 (
m
m
;
5.43 (d, 4H, J5,NH = 10.5 Hz, NH), 5.38ꢀ5.32 (m, 8H, J8,9a = 2.1 Hz,
J8,9b = 4.4 Hz, J6,7 = 1.7 Hz, H-8 and H-7), 4.86 (ddd, 4H,
J4,5 = 10.5 Hz, J3ax,4 = 12.2 Hz, J3eq,4 4.6 = Hz, H-4), 4.31 (dd, 4H,
J9a,9b = 12.4 Hz, H-9a), 4.11 (dd, 4H, H-9b), 4.05 (q, 4H,
J5,6 = 10.5 Hz, H-5), 3.83 (dd, 4H, H-6), 3.80 (s, 12H, OMe), 3.18
(q, 8H, J = 6.6 Hz, CH2N)), 2.78–2.70 (m, 4H, SCHa), 2.72 (dd, 4H,
J3ax,3eq = 12.2 Hz, H-3eq), 2.57–2.50 (m, 4H, SCHb), 2.50 (t, 8H,
J = 7.3 Hz, CH2SCH2), 2.49 (t, 8H, J = 7.2 Hz, CH2SCH2), 2.18 (t, 8H,
J = 7.5 Hz, CH2CO), 2.17, 2.14, 2.05, 2.03 (4 ꢂ s, 48H, OAc), 1.98 (t,
4H, H-3ax), 1.88 (s, 12H, NAc), 1.68ꢀ1.41 (m, 56H,
SCH2CH2CH2CH2CH2SCH2CH2CH2CH2CH2CONHCH2CH2), 0.53–0.49
(m, 8H, Si CH2); 13C NMR (100 MHz, CDCl3): d 173.12, 170.87,
170.65, 170.18, 170.07, 169.99, 168.44, 83.10, 74.05, 69.67, 68.56,
67.27, 62.11, 52.90, 49.30, 42.46, 40.71, 38.05, 36.55, 31.90,
29.33, 29.15, 28.83, 28.67, 28.53, 28.00, 25.36, 23.89, 23.16,
21.17, 20.84, 20.78, 9.15; HRMS (ESI): calcd for [M+3Na]3+/3:
1050.06760, found: m/z 1050.0695.
NMR (400 MHz, D2O): d 3.86ꢀ3.62 (m, 28H, H-9a, H-9b, H-8, H-
7, H-6, H-5, H-4), 2.82 (br s, 8H, SCHa and H-3eq), 2.71 (br s, 4H,
SCHb), 2.54 (br s, 16H, CH2SCH2), 2.03 (s, 12H, NAc), 1.87 (br s,
4H, H-3ax), 1.60, 1.49 (2 ꢂ br s, 32H, SCH2CH2CH2CH2CH2SCH2CH2),
0.69 (br s, 6H, SiCH2); 13C NMR (100 MHz, D2O): d 174.86, 172.13,
83.59, 74.70, 71.04, 67.87, 67.75, 62.34, 51.50, 40.41, 35.50, 31.45,
29.11, 28.86, 28.51, 27.80, 23.97, 21.83, 11.62; HRMS (ESI): calcd
for [M+H]+: 1901.67508, found: m/z 1901.66668.
4.1.31. Ball(0)4-ether-S-Neu5Ac4(OAc, OMe) (45)
The condensation reaction between thiosialoside 21 (321 mg,
0.493 mmol) and Ball(0)4-ether-Br 33 (64 mg, 0.069 mmol) was
carried out by
a method similar to that described for the
preparation of 41 to give Ball(0)4-ether-S-Neu5Ac4(OAc, OMe) 45
(113 mg, 59.2%) as a white foam: Rf 0.23 [5:4:1 (v/v/v) CHCl3ꢀ
EtOAcꢀMeOH]; ½a D25
ꢁ
+23.3 (c 1.06, CHCl3); IR (KBr): 2932 (mCꢀH),
2857 (mCꢀH), 1742 (
m
C@O), 1667 (
m
C@O, amide I), 1549 (dNꢀH, amide
II), 1227 (
m
CꢀO), 1109 (
m
CꢀOꢀC), 1036 (
m
CꢀOꢀC) cmꢀ1 1H NMR
;
4.1.34. Ball(0)4-amide-S-Neu5Ac4 (8)
(400 MHz, CDCl3): d 5.39 (d, 4H, J5,NH = 10.4 Hz, NH), 5.36ꢀ5.31
(m, 8H, J8,9a = 2.1 Hz, J8,9b = 4.6 Hz, J6,7 = 1.9 Hz, H-8 and H-7),
4.86 (ddd, 4H, J4,5 = 10.4 Hz, J3ax,4 = 12.3 Hz, J3eq,4 4.6 = Hz, H-4),
4.32 (dd, 4H, J9a,9b = 12.3 Hz, H-9a), 4.11 (dd, 4H, H-9b), 4.04 (q,
4H, J5,6 = 10.4 Hz, H-5), 3.84 (dd, 4H, H-6), 3.80 (s, 12H, OMe),
3.47 (t, 8H, J = 6.3 Hz, OCH2CH2CH2S), 3.34 (t, 8H, J = 7.0 Hz,
The de-protection of Ball(0)4-amide-S-Neu5Ac4(OAc, OMe) 46
(30.7 mg, 9.96 lmol) was carried out by a method similar to that
described for the preparation of 3 to give Ball(0)4-amide-S-
Neu5Ac4 8 (23.4 mg, quantitative) as a white powder after lyophi-
lization: Rf 0.54 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D26
ꢁ
+22.4 (c
CꢀH), 1701
mC@O, amide I), 1555 (dNꢀH, amide II), 1273 (mCꢀO),
1.41, H2O); IR (KBr): 3399 (mOꢀH), 2930 (mCꢀH), 2857 (m
SiCH2CH2CH2O), 2.78ꢀ2.70 (m, 4H, SCHa), 2.72 (dd, 4H, J3ax,3eq
=
(m
C@O), 1638 (
12.7 Hz, H-3eq), 2.59ꢀ2.48 (m, 4H, SCHb), 2.57 (t, 8H, J = 7.3 Hz,
CH2SCH2), 2.50 (t, 8H, J = 7.3 Hz, CH2SCH2), 2.16, 2.14, 2.04, 2.03
(4 ꢂ s, 48H, OAc), 1.98 (m, 4H, H-3ax), 1.87 (s, 12H, NAc), 1.83
(quint, 8H, OCH2CH2CH2S), 1.62ꢀ1.41 (m, 32H, SCH2CH2CH2CH2-
CH2SCH2CH2), 0.53ꢀ0.49 (m, 8H, SiCH2); 13C NMR (100 MHz,
CDCl3): d 170.80, 170.49, 170.10, 170.03, 169.95, 168.49, 83.13,
74.14, 73.94, 69.67, 69.29, 68.82, 67.41, 62.16, 52.80, 49.37,
38.07, 31.88, 29.81, 29.12, 28.85, 28.82, 28.96, 27.99, 23.98,
1036 (
m
CꢀOꢀC) cmꢀ1
;
1H NMR (400 MHz, D2O): d 3.84ꢀ3.61 (m,
28H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4), 3.15 (br s, 8H, CH2N),
2.80 (br s, 8H, SCHa and H-3eq), 2.68 (br s, 4H, SCHb), 2.52 (br s,
16H, CH2SCH2), 2.24 (br s, 8H, CH2CO), 2.04 (s, 12H, NAc), 1.84
(br s, 4H, H-3ax), 1.58, 1.45 (2 ꢂ br s, 56H, SCH2CH2CH2CH2-
CH2SCH2CH2CH2CH2CH2CONHCH2CH2), 0.53 (br s, 8H, SiCH2); 13C
NMR (100 MHz, CDCl3): d 175.14 (2 ꢂ C), 172.52, 83.89, 75.21,
71.54, 68.13, 62.81, 52.03, 42.78, 40.83, 36.19, 31.85, 29.29,
28.91, 28.40, 28.13, 25.69, 23.74, 22.29, 9.23; HRMS (ESI): calcd
for [M+2Na]2+/2: 1199.49098, found: m/z 1199.4899.
23.07, 21.06, 20.74, 20.66, 8.15; HRMS (ESI): calcd for [M+2Na]2+
/
2: 1453.52295, found: m/z 1453.51809.
4.1.32. Ball(0)4-ether-S-Neu5Ac4 (7)
4.1.35. Dumbbell(1)6-S-Neu5Ac6(OAc, OMe) (47)
The de-protection of Ball(0)4-ether-S-Neu5Ac4(OAc, OMe) 45
(33.2 mg, 11.59 mol) was carried out by a method similar to that
described for the preparation of 3 to give Ball(0)4-ether-S-Neu5Ac4
The condensation reaction between thiosialoside 21 (457 mg,
0.701 mmol) and Dumbbell(1)6-Br 35 (64 mg, 0.069 mmol) was
carried out by a method similar to that described for the prepara-
tion of 41 to give Dumbbell(1)6-S-Neu5Ac6(OAc, OMe) 47 (151 mg,
l
7 (24.8 mg, quantitative) as a white powder after lyophilization: Rf
0.59 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D25
ꢁ
+28.6 (c 1.12, H2O);
CꢀH), 1699 ( C@O),
CꢀO), 1113
1H NMR (400 MHz, D2O):
53.5%) as
a
white foam: Rf 0.27 [7:2:1 (v/v/v) CHCl3ꢀ
IR (KBr): 3420 (
m
OꢀH), 2928 (
m
CꢀH), 2857 (
m
m
EtOAcꢀMeOH]; ½a D24
ꢁ
+23.6 (c 0.66, CHCl3); IR (KBr): 2931 (
C@O, amide I), 1545 (dNꢀH, amide II), 1221
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz, CDCl3): d 5.37
mCꢀH),
1636 (
m
C@O, amide I), 1558 (dNꢀH, amide II), 1273 (
m
1736 (mC@O), 1665 (m
(m
CꢀOꢀC), 1032
(mCꢀOꢀC
)
cmꢀ1
;
d
(m
CꢀO), 1036 (
m
;
3.84ꢀ3.63 (m, 28H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4), 3.51 (t,
8H, OCH2CH2CH2S), 3.40 (t, 8H, SiCH2CH2CH2O), 2.82 (br s, 8H,
SCHa and H-3eq), 2.70 (br s, 4H, SCHb), 2.59, 2.54 (2 ꢂ br s, 16H,
CH2SCH2), 2.04 (s, 12H, NAc), 1.84 (br s, 12H, H-3ax, OCH2CH2
CH2S), 1.60, 1.50 (2 ꢂ br s, 32H, SCH2CH2CH2CH2CH2SCH2CH2),
0.58 (br s, 6H, SiCH2); 13C NMR (100 MHz, D2O): d 174.68,
171.82, 83.19, 74.72, 73.25, 70.99, 68.71, 67.69, 62.30, 61.48,
40.24, 31.36, 29.35, 28.89, 28.76, 28.44, 28.19, 27.76, 23.61,
21.80, 20.05, 7.98; HRMS (ESI): calcd for [M+Na]+: 2155.82448,
found: m/z 2155.81847.
(ddd, 6H, J7,8 = 8.5 Hz, J8,9a = 2.2 Hz, J8,9b = 4.6 Hz, H-8), 5.33 (dd,
6H, J6,7 = 1.7 Hz, H-7), 5.28 (d, 6H, J5,NH = 10.4 Hz, NH), 4.86 (ddd,
6H, J4,5 = 10.4 Hz, J3ax,4 = 12.1 Hz, J3eq,4 = 4.5 Hz, H-4), 4.31 (dd, 6H,
J9a,9b = 12.3 Hz, H-9a), 4.11 (dd, 6H, H-9b), 4.05 (q, 6H,
J5,6 = 10.4 Hz, H-5), 3.83 (dd, 6H, H-6), 3.81 (s, 18H, OMe), 2.78–
2.71 (m, 6H, SCHa), 2.72 (dd, 6H, J3ax,3eq = 12.7 Hz, H-3eq),
2.58ꢀ2.47 (m, 8H, SCHb), 2.50 (t, 12H, J = 7.1 Hz, CH2SCH2), 2.49
(t, 12H, J = 7.3 Hz, CH2SCH2), 2.17, 2.14, 2.05, 2.03 (4 ꢂ s, 72H,
OAc), 1.98 (t, 6H, H-3ax), 1.87 (s, 18H, NAc), 1.62ꢀ1.43 (m, 48H,
SCH2CH2CH2CH2CH2SCH2CH2), 1.32ꢀ1.23 (m, 4H, SiCH2CH2CH2Si),

5462
J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
0.63ꢀ0.51 (m, 20H, SiCH2), ꢀ0.06 (s, 6H, SiCH3); 13C NMR
(2 ꢂ br s, 52H, SCH2CH2CH2CH2CH2SCH2CH2 and SiCH2CH2CH2Si),
0.58 (br s, 20H, SiCH2), ꢀ0.02 (s, 6H, SiCH3); 13C NMR (100 MHz,
D2O): d 174.72, 171.79, 83.32, 74.81, 73.23, 71.06, 68.75, 67.89,
67.63, 66.45, 62.90, 62.41, 61.56, 40.34, 31.44, 29.47, 28.90,
28.76, 28.47, 28.30, 27.72, 23.69, 21.86, 2.78, 8.17, ꢀ3.01; HRMS
(ESI): calcd for [Mꢀ2H]2ꢀ/2: 1676.67135, found: m/z 1676.67301.
(100 MHz, CDCl3):
d 170.86, 170.58, 170.17, 170.05, 170.00,
168.44, 83.11, 74.07, 69.66, 68.67, 67.26, 62.10, 52.88, 49.27,
38.02, 36.03, 31.97, 29.25, 28.82, 28.67, 28.04, 24.27, 23.11,
21.14, 20.80, 20.74, 20.37, 18.29, 17.06, 12.08, ꢀ3.27; HRMS
(ESI): calcd for [M+2H]2+/2: 2072.71705, found: m/z 2072.71684.
4.1.36. Dumbbell(1)6-s-neu5ac6 (9)
4.1.39. Dumbbell(1)6-amide-S-Neu5Ac6(OAc, OMe) (49)
The de-protection of Dumbbell(1)6-S-Neu5Ac6(OAc, OMe) 47
(21.1 mg, 4.90 lmol) was carried out by a method similar to that
described for the preparation of 3 to give Dumbbell(1)6-S-Neu5Ac6
The condensation reaction between thiosialoside 21 (219 mg,
0.336 mmol) and Dumbbell(1)6-amide-Br 37 (50 mg, 0.031 mmol)
was carried out by a method similar to that described for the prep-
aration of 41 to give Dumbbell(1)6-amide-S-Neu5Ac6(OAc, OMe)
49 (104 mg, 70.0%) as a white foam: Rf 0.30 [7:2:1 (v/v/v)
9 (16.2 mg, quantitative) as a white powder after lyophilization: Rf
0.42 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D27
ꢁ
+23.2 (c 0.99, H2O);
C@O), 1636 ( C@O, amide
CꢀOꢀC) cmꢀ1; 1H NMR
IR (KBr): 3418 (
m
OꢀH), 2924 (
m
CꢀH), 1701 (
m
m
CHCl3ꢀEtOAcꢀMeOH]; ½a D27
ꢁ
+23.3 (c 1.04, CHCl3); IR (KBr): 3291
CꢀH), 1744 ( C@O), 1651 ( C@O, amide
CꢀN), 1227 ( CꢀO), 1036 (mCꢀOꢀC
1H NMR (400 MHz, CDCl3): d 6.22 (t, 6H, J = 5.3 Hz, NHCH2),
I), 1560 (dNꢀH, amide II), 1261 (
m
CꢀO), 1032 (
m
(
m
NꢀH), 2930 (
m
CꢀH), 2859 (
m
m
m
(400 MHz, D2O): d 3.88ꢀ3.63 (m, 42H, H-9a, H-9b, H-8, H-7, H-6,
H-5, H-4), 2.83 (br s, 12H, SCHa and H-3eq), 2.72 (br s, 6H, SCHb),
2.54 (br s, 24H, CH2SCH2), 2.04 (s, 18H, NAc), 1.88 (br s, 6H, H-
3ax), 1.61, 1.50 (2 ꢂ br s, 48H, SCH2CH2CH2CH2CH2SCH2CH2), 1.34
(m, 4H, SiCH2CH2CH2Si), 0.68 (br s, 20H, SiCH2), ꢀ0.07 (s, 6H,
SiCH3); 13C NMR (100 MHz, D2O): d 174.73, 172.07, 83.50, 74.72,
71.03, 67.83, 62.32, 51.52, 40.40, 35.59, 31.55, 29.19, 28.91,
28.59, 27.94, 24.04, 21.86, 20.01, 18.35, 17.19, 11.69, ꢀ2.68; HRMS
(ESI): calcd for [M+2H]2+/2: 1504.5614, found: m/z 1504.5558.
I), 1549 (dNꢀH, amide II), 1261 (
m
m
)
cmꢀ1
;
5.42 (d, 6H, J5,NH = 10.3 Hz, NH), 5.38–5.32 (m, 12H, J8,9a = 2.0 Hz,
J8,9b = 4.4 Hz, J6,7 = 1.6 Hz, H-8 and H-7), 4.86 (ddd, 6H,
J4,5 = 10.3 Hz, J3ax,4 = 12.3 Hz, J3eq,4 = 4.6 Hz, H-4), 4.32 (dd, 6H,
J9a,9b = 12.4 Hz, H-9a), 4.11 (dd, 6H, H-9b), 4.05 (q, 6H,
J5,6 = 10.3 Hz, H-5), 3.84 (dd, 6H, H-6), 3.80 (s, 18H, OMe), 3.18
(q, 12H, J = 6.4 Hz, CH2N), 2.78–2.70 (m, 6H, SCHa), 2.72 (dd, 6H,
J3ax,3eq = 12.3 Hz, H-3eq), 2.57ꢀ2.50 (m, 6H, SCHb), 2.50 (t, 12H,
J = 7.2 Hz, CH2SCH2), 2.49 (t, 12H, J = 7.3 Hz, CH2SCH2), 2.18 (t,
12H, J = 7.6 Hz, CH2CO), 2.16, 2.14, 2.04, 2.03 (4 ꢂ s, 72H, OAc),
1.97 (t, 6H, H-3ax), 1.88 (s, 18H, NAc), 1.68ꢀ1.41 (m, 84H,
SCH2CH2CH2CH2CH2SCH2CH2CH2CH2CH2CONHCH2CH2), 1.28ꢀ1.26
(m, 4H, SiCH2CH2CH2Si), 0.59ꢀ0.48 (m, 20H, SiCH2), ꢀ0.07 (s, 6H,
SiCH3); 13C NMR (100 MHz, CDCl3): d 173.17, 170.87, 170.66,
170.24, 170.09, 170.03, 168.48, 83.13, 74.04, 69.69, 68.64, 67.31,
62.13, 52.90, 49.31, 43.83, 42.61, 38.07, 36.53, 31.93, 29.36,
29.17, 28.84, 28.69, 28.57, 28.01, 25.40, 23.98, 23.15, 21.17,
20.83, 20.78, 20.16, 18.43, 16.99, 9.29, ꢀ3.02; HRMS (ESI): calcd
for [M+4Na]4+/4: 1217.47896, found: m/z 1217.4819.
4.1.37. Dumbbell(1)6-ether-S-Neu5Ac6(OAc, OMe) (48)
The condensation reaction between thiosialoside 21 (306 mg,
0.470 mmol) and Dumbbell(1)6-ether-Br 36 (50 mg, 0.039 mmol)
was carried out by a method similar to that described for the prep-
aration of 41 to give Dumbbell(1)6-ether-S-Neu5Ac6(OAc, OMe) 48
(55 mg, 31.6%) as
a white foam: Rf 0.33 [7:2:1 (v/v/v)
CHCl3ꢀEtOAcꢀMeOH]; ½a D21
ꢁ
+24.7 (c 1.29, CHCl3); IR (KBr): 2930
C@O), 1663 ( C@O, amide I), 1541 (dNꢀH
CꢀOꢀC), 1036 (
CꢀOꢀC) cmꢀ1; 1H NMR
(m
CꢀH), 2857 (
m
CꢀH), 1744 (
m
m
,
amide II), 1227 (
mCꢀO), 1111 (
m
m
(400 MHz, CDCl3): d 5.36 (ddd, 6H, J7,8 = 8.4 Hz, J8,9a = 2.2 Hz,
J8,9b = 4.6 Hz, H-8), 5.33 (dd, 6H, J6,7 = 2.0 Hz, H-7), 5.27 (d, 6H,
J5,NH = 10.4 Hz, NH), 4.86 (ddd, 6H, J4,5 = 10.4 Hz, J3ax,4 = 12.3 Hz,
J3eq,4 = 4.6 Hz, H-4), 4.31 (dd, 6H, J9a,9b = 12.4 Hz, H-9a), 4.11 (dd,
6H, H-9b), 4.05 (q, 6H, J5,6 = 10.4 Hz, H-5), 3.83 (dd, 6H, H-6),
3.80 (s, 18H, OMe), 3.48 (t, 12H, J = 6.3 Hz, OCH2CH2CH2S), 3.34
(t, 12H, J = 7.0 Hz, SiCH2CH2CH2O), 2.78ꢀ2.70 (m, 6H, SCHa), 2.72
(dd, 6H, J3ax,3eq = 12.7 Hz, H-3eq), 2.59–2.48 (m, 6H, SCHb), 2.57 (t,
12H, J = 7.2 Hz, CH2SCH2), 2.50 (t, 12H, J = 7.3 Hz, CH2SCH2), 2.17,
2.14, 2.04, 2.03 (4 ꢂ s, 72H, OAc), 1.98 (m, 6H, H-3ax), 1.87 (s,
18H, NAc), 1.83 (quint, 12H, OCH2CH2CH2S), 1.62ꢀ1.41 (m, 48H,
SCH2CH2CH2CH2CH2SCH2CH2), 1.32ꢀ1.24 (m, 4H, SiCH2CH2CH2Si),
0.60–0.47 (m, 20H, SiCH2), ꢀ0.07 (s, 6H, SiCH3); 13C NMR
4.1.40. Dumbbell(1)6-amide-S-Neu5Ac6 (11)
The de-protection of Dumbbell(1)6-amide-S-Neu5Ac6(OAc,
OMe) 49 (32.7 mg, 6.84 lmol) was carried out by a method similar
to that described for the preparation of 3 to give Dumbbell(1)6-
amide-S-Neu5Ac6 11 (24.9 mg, 98.8%) as a white powder after
lyophilization: Rf 0.53 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D26
ꢁ
+18.4 (c 1.01, H2O); IR (KBr): 3383 (mOꢀH), 2930 (mCꢀH), 2859 (mCꢀH),
1701 (
m
C@O), 1639 (
m
C@O, amide I), 1557 (dNꢀH, amide II), 1271
(mCꢀO), 1034
(mCꢀOꢀC
)
cmꢀ1 1H NMR (400 MHz, D2O):
;
d
3.85ꢀ3.62 (m, 42H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4), 2.82 (br
s, 12H, SCHa and H-3eq), 2.68 (br s, 6H, SCHb), 2.53 (br s, 24H,
CH2SCH2), 2.04 (s, 18H, NAc), 1.85 (br s, 6H, H-3ax), 1.59, 1.47
(2 ꢂ br s, 88H, SCH2CH2CH2CH2CH2SCH2CH2CH2CH2CH2CON-
HCH2CH2 and SiCH2CH2CH2Si), 0.58 (br s, 20H, SiCH2), ꢀ0.02 (s,
6H, SiMe); 13C NMR (100 MHz, D2O): d 175.16 (2 ꢂ C), 172.36,
83.67, 75.25, 71.60 68.16, 62.82, 52.23, 41.05, 40.70, 31.90, 29.37,
28.91, 28.19, 25.89, 22.30, 9.23, ꢀ2.65; HRMS (ESI): calcd for
[M+3Na]3+/3: 1251.52639, found: m/z 1251.5248.
(100 MHz, CDCl3):
d 170.88, 170.58, 170.14, 170.07, 169.96,
168.44, 83.11, 74.07, 69.66, 69.31, 68.59, 67.26, 62.11, 52.89,
49.34, 38.04, 31.86, 29.78, 29.13, 28.83, 28.69, 28.04, 24.04,
23.16, 21.16, 20.83, 20.76, 20.41, 18.28, 17.06, 8.15, ꢀ3.27; HRMS
(ESI): calcd for [M+3Na]3+/3: 1505.55837, found: m/z 1505.55195.
4.1.38. Dumbbell(1)6-ether-S-Neu5Ac6 (10)
The de-protection of Dumbbell(1)6-ether-S-Neu5Ac6(OAc,
OMe) 48 (26.2 mg, 5.89
lmol) was carried out by a method similar
4.1.41. Ball(1)12-S-Neu5Ac12(OAc, OMe) (50)
to that described for the preparation of 3 to give Dumbbell(1)6-
ether-S-Neu5Ac6 10 (20.0 mg, quantitative) as a white powder
after lyophilization: Rf 0.58 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O];
The condensation reaction between thiosialoside 21 (250 mg,
0.384 mmol) and Ball(1)12-Br 38 (40 mg, 0.023 mmol) was carried
out by a method similar to that described for the preparation of 41
to give Ball(1)12-S-Neu5Ac12(OAc, OMe) 50 (108 mg, 59.0%) as a
½
a 2D5
+20.0 (c 1.05, H2O); IR (KBr): 3408 (mOꢀH), 2928 (mCꢀH), 2855
ꢁ
(
m
CꢀH), 1699 (
m
C@O), 1636 (
m
C@O, amide I), 1558 (dNꢀH, amide II),
white foam: Rf 0.45 [7:1 (v/v) CHCl3ꢀMeOH]; ½a D26
ꢁ
+27.4 (c 1.05,
1273 CꢀO), 1113
(m
(mCꢀOꢀC), 1032
(mCꢀOꢀC
)
cmꢀ1 1H NMR
;
CHCl3); IR (KBr): 2924 (
1549 (dNꢀH, amide II), 1229 (
m
CꢀH), 1740 (
m
C@O), 1663 (
m
C@O, amide I),
(400 MHz, D2O): d 3.84ꢀ3.40 (m, 66H, H-9a, H-9b, H-8, H-7, H-6,
H-5, H-4, OCH2CH2CH2S, SiCH2CH2CH2O), 2.82 (br s, 12H, SCHa
and H-3eq), 2.70 (br s, 6H, SCHb), 2.54 (br s, 24H, CH2SCH2), 2.04
(s, 18H, NAc), 1.84 (br s, 18H, H-3ax, OCH2CH2CH2S), 1.61, 1.51
mCꢀO), 1038 (
m
CꢀOꢀC) cmꢀ1
;
1H NMR
(400 MHz, CDCl3): d 5.48 (d, 12H, J5,NH = 10.3 Hz, NH), 5.38ꢀ5.32
(m, 12H, J8,9b = 4.5 Hz, H-8 and H-7), 4.86 (ddd, 12H,
J4,5 = 10.3 Hz, J3ax,4 = 12.4 Hz, J3eq,4 = 4.6 Hz, H-4), 4.31 (d, 12H,

J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
5463
J9a,9b = 11.6 Hz, H-9a), 4.12 (dd, 12H, H-9b), 4.05 (q, 12H,
J5,6 = 10.3 Hz, H-5), 3.84 (d, 12H, H-6), 3.81 (s, 36H, OMe),
2.76ꢀ2.70 (m, 12H, SCHa), 2.72 (dd, 12H, J3ax,3eq = 12.4 Hz, H-3eq),
2.57ꢀ2.50 (m, 12H, SCHb), 2.50 (t, 24H, J = 7.2 Hz, CH2SCH2), 2.49
(t, 24H, J = 7.3 Hz, CH2SCH2), 2.17, 2.14, 2.05, 2.03 (4 ꢂ s, 144H,
OAc), 1.99 (t, 12H, H-3ax), 1.87 (s, 36H, NAc), 1.61ꢀ1.41 (m, 96H,
SCH2CH2CH2CH2CH2SCH2CH2CH2Si), 1.28ꢀ1.26 (m, 8H, SiCH2CH2
CH2Si), 0.64ꢀ0.51 (m, 40H, SiCH2); 13C NMR (100 MHz, CDCl3): d
170.86, 170.65, 170.24, 170.06, 169.99, 168.48, 83.14, 74.07,
69.69, 68.61, 67.23, 62.09, 52.93, 49.28, 38.03, 36.13, 32.08,
29.64, 29.33, 28.87, 28.70, 28.13, 24.31, 23.12, 21.19, 20.84,
20.79, 18.41, 17.59, 12.09; HRMS (ESI): calcd for [M+5Na]5+/5:
1646.3587, found: m/z 1646.3777.
1701 (
m
C@O), 1638 (
m
C@O, amide I), 1560 (dNꢀH, amide II), 1275
(mCꢀO), 1112 (mCꢀOꢀC), 1032 (m ;
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz,
D2O): d 3.86ꢀ3.38 (m, 132H, H-9a, H-9b, H-8, H-7, H-6, H-5, H-4,
CH2OCH2), 2.83 (br s, 24H, SCHa and H-3eq), 2.70 (br s, 12H, SCHb),
2.58 (br s, 48H, CH2SCH2), 2.04 (s, 36H, NAc), 1.85 (br s, 36H, H-3ax
and
OCH2CH2CH2S),
1.61,
1.50
(2 ꢂ br
s,
96H,
SCH2CH2CH2CH2CH2SCH2CH2), 1.39 (br s, 8H, SiCH2CH2CH2Si),
0.58 (br s, 40H, SiCH2); 13C NMR (100 MHz, D2O): d 175.14,
172.28, 83.66, 75.22, 74.67, 73.77, 71.47, 69.26, 68.19, 62.79,
51.94, 49.00 (MeOD), 40.80, 31.83, 30.28, 29.84, 29.38, 28.94,
28.70, 28.25, 24.25, 22.29, 24.25, 22.29, 18.00, 8.54; HRMS (ESI):
calcd for [Mꢀ6H]6ꢀ/6: 1103.7681, found: m/z 1103.7648.
4.1.45. Ball(1)12-amide-S-Neu5Ac12(OAc, OMe) (52)
4.1.42. Ball(1)12-s-neu5ac12 (12)
The de-protection of Ball(1)12-S-Neu5Ac12(OAc, OMe) 50
(39.7 mg, 4.89 lmol) was carried out by a method similar to that
The condensation reaction between thiosialoside 21 (175 mg,
0.269 mmol) and Ball(1)12-amide-Br 40 (46 mg, 0.015 mmol)
was carried out by a method similar to that described for the prep-
aration of 41 to give Ball(1)12-amide-S-Neu5Ac12(OAc, OMe) 52
described for the preparation of 3 to give Ball(1)12-S-Neu5Ac12
12 (27.2 mg, 93.8%) as a white powder after lyophilization: Rf
(88 mg, 63.3%) as a white foam: ½a D26
ꢁ
+20.5 (c 0.83, CHCl3); Rf
NꢀH), 2932 ( CꢀH),
C@O, amide I), 1545 (dNꢀH, amide
CꢀO), 1036 (mCꢀOꢀC
cmꢀ1 1H NMR
(400 MHz, CDCl3): d 6.75 (br s, 12H, NHCH2), 5.54 (d, 12H, J5,NH
10.3 Hz, NH), 5.38ꢀ5.32 (m, 24H, J8,9a = 1.9 Hz, J8,9b = 4.1 Hz, H-8
and H-7), 4.86 (ddd, 12H, J4,5 = 10.3 Hz, J3ax,4 = 12.3 Hz, J3eq,4
0.17 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D24
ꢁ
+30.7 (c 1.13, H2O);
C@O), 1636 ( C@O, amide
CꢀOꢀC) cmꢀ1; 1H NMR
0.47 [7:1 (v/v) CHCl3ꢀMeOH]; IR (KBr): 3275 (
m
m
IR (KBr): 3379 (
m
OꢀH), 2920 (
m
CꢀH), 1717 (
m
m
2860 (
m
CꢀH), 1742 (
mC@O), 1647 (m
I), 1558 (dNꢀH, amide II), 1279 (
m
CꢀO), 1034 (
m
II), 1261 (
m
CꢀN), 1227 (
m
)
;
(400 MHz, D2O): d 3.92ꢀ3.60 (m, 84H, H-9a, H-9b, H-8, H-7, H-6,
H-5 and H-4), 2.81 (br s, 24H, SCHa and H-3eq), 2.69 (br s, 12H,
SCHb), 2.56 (br s, 48H, CH2SCH2), 2.03 (s, 36H, NAc), 1.80 (br s,
12H, H-3ax), 1.62, 1.50 (2 ꢂ br s, 96H, SCH2CH2CH2CH2CH2SCH2CH2
CH2Si), 1.48 (br s, 8H, SiCH2CH2CH2Si), 0.70 (br s, 40H, SiCH2); 13C
NMR (100 MHz, CDCl3): d 175.15, 172.28, 83.67, 75.16, 71.43,
68.18, 62.78, 51.90, 49.00, 40.70, 36.08, 32.01, 29.63, 29.29,
28.97, 28.34, 24.47, 22.27, 20.52, 18.97, 12.18; HRMS(MALDI-
TOF): calcd for [M+Na]+: 5954.138, found: m/z 5954.212.
=
=
4.4 Hz, H-4), 4.31 (dd, 12H, J9a,9b = 12.4 Hz, H-9a), 4.11 (dd, 12H,
H-9b), 4.05 (q, 12H, J5,6 = 10.3 Hz, H-5), 3.84 (d, 12H, H-6), 3.81
(s, 36H, OMe), 3.16 (br d, 24H, J = 5.1 Hz, CH2N), 2.77ꢀ2.70 (m,
12H, SCHa), 2.72 (dd, 12H, J3ax,3eq = 12.3 Hz, H-3eq), 2.58ꢀ2.50 (m,
12H, SCHb), 2.50 (t, 24H, J = 7.2 Hz, CH2SCH2), 2.49 (t, 24H,
J = 7.2 Hz, CH2SCH2), 2.19 (t, 24H, J = 7.5 Hz, CH2CO), 2.17, 2.14,
2.05, 2.03 (4 ꢂ s, 144H, OAc), 1.97 (t, 12H, H-3ax), 1.88 (s, 18H,
NAc), 1.67ꢀ1.37 (m, 168H, SCH2CH2CH2CH2CH2SCH2CH2CH2CH2
CH2CONHCH2CH2), 1.29ꢀ1.24 (m, 8H, SiCH2CH2CH2Si), 0.50 (br d,
4.1.43. Ball(1)12-ether-S-Neu5Ac12(OAc, OMe) (51)
The condensation reaction between thiosialoside 21 (180 mg,
0.276 mmol) and Ball(1)12-ether-Br 39 (40 mg, 0.016 mmol) was
carried out by a method similar to that described for the prepara-
tion of 41 to give Ball(1)12-ether-S-Neu5Ac12(OAc, OMe) 51
(93 mg, 65.0%) as a white foam: Rf 0.43 [8:1 (v/v) CHCl3ꢀMeOH];
40H, SiCH2); 13C NMR (100 MHz, CDCl3):
d 173.37, 170.84,
170.66, 170.24, 170.07, 170.02, 168.46, 83.10, 74.04, 69.69, 68.59,
67.27, 62.11, 52.89, 49.24, 42.68, 38.03, 36.42, 31.91, 29.38,
29.15, 28.82, 28.65, 28.62, 28.01, 25.45, 23.92, 23.12, 21.17,
20.82, 20.78, 18.54, 17.49, 17.13, 9.38; HRMS (ESI): calcd for
[M+5Na]5+/5: 1918.0024, found: m/z 1918.1611.
½
a 2D7
ꢁ
+22.7 (c 1.09, CHCl3); IR (KBr): 2932 (
1742 ( C@O), 1665 (
CꢀO), 1111 ( CꢀOꢀC), 1036 (
m
CꢀH), 2859 (
C@O, amide I), 1545 (dNꢀH, amide II), 1227
CꢀOꢀC) cmꢀ1 1H NMR (400 MHz,
mCꢀH),
m
m
(
m
m
m
;
CDCl3): d 5.43 (d, 12H, J5,NH = 10.3 Hz, NH), 5.38ꢀ5.32 (m, 24H,
4.1.46. Ball(1)12-amide-S-Neu5Ac12 (14)
The de-protection of Ball(1)12-amide-S-Neu5Ac12(OAc, OMe)
52 (34.0 mg, 3.59 lmol) was carried out by a method similar to
J8,9a = 1.7 Hz, J8,9b = 4.4 Hz, H-8 and H-7), 4.86 (ddd, 12H, J4,5
10.3 Hz, J3ax,4 = 12.3 Hz, J3eq,4 = 4.7 Hz, H-4), 4.31 (dd, 12H, J9a,9b
=
=
12.4 Hz, H-9a), 4.12 (dd, 12H, H-9b), 4.05 (q, 12H, J5,6 = 10.4 Hz,
H-5), 3.83 (d, 12H, H-6), 3.81 (s, 36H, OMe), 3.47 (t, 24H,
J = 6.3 Hz, OCH2CH2CH2S), 3.34 (t, 24H, J = 7.0 Hz, SiCH2CH2CH2O),
2.78–2.70 (m, 12H, SCHa), 2.72 (dd, 12H, J3ax,3eq = 12.3 Hz, H-3eq),
2.59ꢀ2.48 (m, 12H, SCHb), 2.57 (t, 24H, J = 7.2 Hz, CH2SCH2), 2.50
(t, 24H, J = 7.3 Hz, CH2SCH2), 2.17, 2.14, 2.05, 2.03 (4 ꢂ s, 144H,
OAc), 1.98 (t, 12H, H-3ax), 1.87 (s, 36H, NAc), 1.83 (quint, 24H,
OCH2CH2CH2S), 1.62ꢀ1.43 (m, 96H, SCH2CH2CH2CH2CH2SCH2CH2),
that described for the preparation of 3 to give Ball(1)12-amide-S-
Neu5Ac12 14 (26.0 mg, 99.2%) as a white powder after lyophiliza-
tion: Rf 0.14 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D25
ꢁ
+12.6 (c
CꢀH), 1699
1.06, H2O); IR (KBr): 3318 (
m
OꢀH), 2928 (
m
CꢀH), 2859 (
m
(
m
C@O), 1636 (
m
C@O, amide I), 1558 (dNꢀH, amide II), 1273 (
mCꢀO),
1032 (
m
CꢀOꢀC) cmꢀ1
;
1H NMR (400 MHz, D2O): d 4.78 (s, HDO,
internal standard), 3.83ꢀ3.57 (m, 84H, H-9a, H-9b, H-8, H-7, H-6,
H-5 and H-4), 3.17 (br s, 24H, CH2N), 2.79 (br s, 24H, SCHa and
H-3eq), 2.66 (br s, 12H, SCHb), 2.53 (br s, 48H, CH2SCH2), 2.25 (br
s, 24H, CH2CO), 2.03 (s, 36H, NAc), 1.79 (br s, 12s H, H-3ax), 1.59,
1.45 (2 ꢂ br s, 168H, SCH2CH2CH2CH2CH2SCH2CH2CH2CH2CH2CON-
HCH2CH2), 1.25 (br s, 8H, SiCH2CH2CH2Si), 0.55 (br s, 40H, SiCH2);
13C NMR (100 MHz, CDCl3): d 175.14 (2 ꢂ C), 172.23, 84.81,
75.10, 71.73, 68.35, 68.27, 62.69, 51.98, 42.75, 41.09, 36.17,
31.77, 29.37, 29.21, 29.06, 28.30, 28.07, 25.71, 23.80, 22.24,
18.80, 17.44, 9.49; HRMS (ESI): calcd for [Mꢀ7H]7ꢀ/7: 1040.3013,
found: m/z 1040.2980.
1.28ꢀ1.21 (m, 8H, SiCH2CH2CH2Si), 0.58–0.46 (m, 40H, SiCH2); 13
C
NMR (100 MHz, CDCl3): d 170.87, 170.62, 170.19, 170.07, 169.98,
168.47, 83.11, 74.11, 74.08, 69.67, 69.39, 68.60, 67.26, 62.11,
52.91, 49.31, 38.04, 31.85, 29.78, 29.15, 28.84, 28.69, 28.07,
24.07, 23.14, 21.17, 20.83, 20.78, 18.39, 17.66, 17.53, 8.12; HRMS
(ESI): calcd for [M+5Na]5+/5: 1785.6593, found: m/z 1785.6934.
4.1.44. Ball(1)12-ether-S-Neu5Ac12 (13)
The de-protection of Ball(1)12-ether-S-Neu5Ac12(OAc, OMe) 51
(32.3 mg, 3.66 lmol) was carried out by a method similar to that
described for the preparation of 3 to give Ball(1)12-ether-S-
4.2. Viruses and sialidases
Neu5Ac12 13 (24.3 mg, quantitative) as a white powder after
lyophilization: Rf 0.17 [3:3:1 (v/v/v) CHCl3ꢀMeOHꢀH2O]; ½a D21
ꢁ
Influenza virus A/PR/8/34 (H1N1) and A/Memphis/1/71 (H3N2)
were cultured in 11-day-old embryonic cheken eggs, purified, and
+30.7 (c 1.12, H2O); IR (KBr): 3401 (mOꢀH), 2928 (mCꢀH), 2857 (mCꢀH),

5464
J.-I. Sakamoto et al. / Bioorg. Med. Chem. 17 (2009) 5451–5464
concentrated as described previously.24 The virus suspensions in
saline were used as the source of sialidase.
8. (a) Le, Q. M.; Kiso, M.; Someya, K.; Sakai, Y. T.; Nguyen, T. H.; Nguyen, K. H. L.;
Pham, N. D.; Ngyen, H. H.; Yamada, S.; Muramoto, Y.; Horimoto, T.; Takada, A.;
Goto, H.; Suzuki, T.; Suzuki, Y.; Kawaoka, Y. Nature (London) 2005, 437, 1108;
(b) de Jong, M. D.; Thanh, T. T.; Khanh, T. H.; Hien, V. M.; Smith, G. J. D.; Chau, N.
V.; Cam, B. V.; Qui, P. T.; Ha, D. Q.; Guan, Y.; Peiris, J. S. M.; Hien, T. T.; Farrar, J.
N. Engl. J. Med. 2005, 353, 2667.
4.3. Sialidase inhibition assay
9. Wilson, J. C.; Kiefel, M. J.; Angus, D. I.; von Itzstein, M. Org. Lett. 1999, 1, 443.
10. (a) Lee, Y. C. Carbohydr. Res. 1978, 67, 509; (b) Lee, Y. C.; Townsend, R. R.; Hardy,
M. R.; Lönngren, J.; Arnarp, J.; Haraldsson, M.; Lönn, H. J. Biol. Chem. 1983, 258,
199.
11. (a) Varghese, J. N.; Laver, W. G.; Colman, P. M. Nature (London) 1983, 303, 35;
(b) Colman, P. M.; Varghese, J. N.; Laver, W. G. Nature (London) 1983, 303, 41.
12. (a) Wilson, I. A.; Skehel, J. J.; Wiley, D. C. Nature (London) 1981, 289, 366; (b)
Wiley, D. C.; Wilson, I. A.; Skehel, J. J. Nature (London) 1981, 289, 373.
13. Matsuoka, K.; Takita, C.; Koyama, T.; Miyamoto, D.; Yingsakmongkon, S.;
Hidari, K. I. P. J.; Jampangern, W.; Suzuki, T.; Suzuki, Y.; Hatano, K.; Terunuma,
D. Bioorg. Med. Chem. Lett. 2007, 17, 3826.
14. (a) Matsuoka, K.; Terabatake, M.; Esumi, Y.; Terunuma, D.; Kuzuhara, H.
Tetrahedron Lett. 1999, 40, 7839; (b) Matsuoka, K.; Terabatake, M.; Umino, A.;
Esumi, Y.; Hatano, K.; Terunuma, D.; Kuzuhara, H. Biomacromolecules 2006, 7,
2274; (c) Matsuoka, K.; Terabatake, M.; Esumi, Y.; Hatano, K.; Terunuma, D.;
Kuzuhara, H. Biomacromolecules 2006, 7, 2284.
15. (a) Nishikawa, K.; Matsuoka, K.; Kita, E.; Okabe, N.; Mizuguchi, M.; Hino, K.;
Miyazawa, S.; Yamasaki, C.; Aoki, J.; Takashima, S.; Yamakawa, Y.; Nishijima,
M.; Terunuma, D.; Kuzuhara, H.; Natori, Y. Proc. Natl. Acad. Sci. U.S.A. 2002, 99,
7669; (b) Nishikawa, K.; Matsuoka, K.; Watanabe, M.; Igai, K.; Hino, K.; Hatano,
K.; Yamada, A.; Abe, N.; Terunuma, D.; Kuzuhara, H.; Natori, Y. J. Infect. Dis.
2005, 191, 2097.
Influenza A virus suspension (HAU = 26), suitable amounts of
glycodendrimers and 4-MU-Neu5Ac were mixed in 20 mM acetate
buffer (pH 5.2), and the mixture was adjusted to 0.4 mM of final
concentration of 4-MU-Neu5Ac and total volume 20 lL. The reac-
tion mixture was incubated at 37 °C for 30 min. The reaction was
stopped by the addition of 1 mL of 100 mM carbonate buffer (pH
10.7). The fuorescence of released 4-methylumbelliferone (4-MU)
was measured using a fuorescence spectrophotometer (Hitachi F-
4010) with excitation at 355 nm and emission at 460 nm. The con-
centration causing 50% inhibition (IC50) of sialidase activity using
4-MU-Neu5Ac was calculated graphically by plotting percent inhi-
bition versus inhibitor concentration.
Acknowledgment
We are grateful to Snow Brand Milk Products Co., Ltd for provid-
ing the sialic acid used for in this study.
16. (a) Matsuoka, K.; Oka, H.; Koyama, T.; Esumi, Y.; Terunuma, D. Tetrahedron Lett.
2001, 42, 3327; (b) Oka, H.; Onaga, T.; Koyama, T.; Guo, C.-T.; Suzuki, Y.; Esumi,
Y.; Hatano, K.; Terunuma, D.; Matsuoka, K. Bioorg. Med. Chem. Lett. 2008, 18,
4405.
References and notes
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Theriault, S.; Suzuki, H.; Nishimura, H.; Mitamura, K.; Sugaya, N.; Usui, T.;
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Feldmann, H.; Kawaoka, Y. Nature (London) 2004, 431, 703. and references cited
therein.
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C. J. Am. Chem. Soc. 1997, 119, 681.
4. von Itzstein, M.; Wu, W.-Y.; Kok, G. B.; Pegg, M. S.; Dyason, J. C.; Jin, B.; Phan, T.
V.; Smythe, M. L.; White, H. F.; Oliver, S. W.; Colman, P. M.; Varghese, J. N.;
Ryan, D. M.; Woods, J. M.; Bethell, R. C.; Hotham, V. J.; Cameron, J. M.; Penn, C.
R. Nature (London) 1993, 363, 418.
5. For example see: Webster, R. G.; Bean, W. J.; Gorman, O. T.; Chambers, T. M.;
Kawaoka, Y. Microbiol. Rev. 1992, 56, 152. and references cited therein.
6. For example see: Wiely, D. C.; Skehel, J. Annu. Rev. Biochem. 1987, 56, 365–394.
and references cited therein.
7. For example, see: ayden, F. G.; Calfee, D. P. Drugs 1998, 56, 537. and references
cited therein.