An efficient access to protected disialylated glycohexaosyl threonine present on the leukosialin of activated T-lymphocytes

Article abstract of DOI:10.1016/S0008-6215(99)00313-4

The total synthesis of the threonine-linked core 2 class disialylated hexasaccharide in a completely protected form was accomplished for the first time. The L-threonine conjugate, N-(9-fluorenylmethoxycarbonyl)-O-{(5-acetamido-4,7,8,9-tetra-O-benzyl-3,5-d

Full text of DOI:10.1016/S0008-6215(99)00313-4

www.elsevier.nl/locate/carres
Carbohydrate Research 325 (2000) 132–142
An efficient access to protected disialylated
glycohexaosyl threonine present on the leukosialin of
activated T-lymphocytes
Latika Singh a, Yuko Nakahara a, Yukishige Ito a, Yoshiaki Nakahara a,b,
*
a The Institute of Physical and Chemical Research (RIKEN), Hirosawa 2-1, Wako-shi, Saitama 351-0198, Japan
b Department of Industrial Chemistry, CREST, Japan Science and Technology Corporation (JST), Tokai Uni6ersity,
Kitakaname 1117, Hiratsuka-shi, Kanagawa 259-1292, Japan
Received 9 September 1999; accepted 18 November 1999
Abstract
The total synthesis of the threonine-linked core 2 class disialylated hexasaccharide in a completely protected form
was accomplished for the first time. The
4,7,8,9-tetra-O-benzyl-3,5-dideoxy- -glycero-a-
galactopyranosyl)-(1“4)-2-acetamido-2-deoxy-3,6-di-O-benzyl-b-
tetra-O-benzyl-3,5-dideoxy- -glycero-a- -galacto-2-nonulopyranosylonic acid)-(2“3)-2,6-di-O-benzyl-b-
pyranosyl-(1“3)]-2-acetamido-2-deoxy-a- -galactopyranosyl-(1d“4c:1f“4e)-dilactone}- -threonine allyl ester was
synthesized via stereocontrolled glycosylations employing readily accessible monosaccharidic blocks; t-butyl-
diphenylsilyl-2-azido-2-deoxy-3,6-di-O-benzyl-b- -glucopyranose, N-(9-fluorenylmethoxycarbonyl)-O-(2-azido-6-O-
t-butyldimethylsilyl-2-deoxy-a- -galactopyranosyl)- -threonine allyl ester, 8, and N-(9-fluorenylmethoxy-
carbonyl)-O-(2-azido-4,6-O-benzylidene-3-O-chloroacetyl-2-deoxy-a- -galactopyranosyl)- -threonine allyl ester. For
L
-threonine conjugate, N-(9-fluorenylmethoxycarbonyl)-O-{(5-acetamido-
-galacto-2-nonulopyranosylonic acid)-(2“3)-(2,6-di-O-benzyl-b-
-glucopyranosyl-(1“6)-[(5-acetamido-4,7,8,9-
-galacto-
D
D
D-
D
D
D
D
D
L
D
D
L
9
D
L
the introduction of the amino acid, the azide group was used to temporarily mask the amino group of GalNAc so
as to obtain an a-glycosidic linkage without participation from the C-2 substituent. The threonine was attached to
the sugar unit at the monosaccharide stage to avoid loss of oligosaccharide at a later stage. The Fmoc and allyl ester
protected amino acid at the reducing end facilitates efficient glycopeptide synthesis on solid-phase support. © 2000
Elsevier Science Ltd. All rights reserved.
Keywords: Core class 2; Sialic acid; Leukosialin; O-Linked glycoprotein
1. Introduction
cell-type and lineage specific [1]. The aberrant
expression of carbohydrate structures is often
associated with various pathological condi-
tions and immunodeficiency [1,2].
Carbohydrate ligands on the surface of
blood cells have been the subject of extensive
studies. Leukosialin is a major sialoglyco-
protein found on the surface of T-lympho-
cytes, granulocytes and monocytes [3]. Nor-
mal resting T-lymphocytes express on
Cell-surface glycoproteins and glycolipids
play an essential role in maintaining the func-
tion as well as the structure of cells. The
carbohydrate structures on glycoproteins and
glycolipids undergo marked alterations during
development and differentiation, and are also
* Corresponding author. Tel./fax: +81-463-502075.
E-mail address: yonak@keyaki.cc.u-tokai.ac.jp (Y. Naka-
hara)
leukosialin the disialotetrasaccharides a-
D
-
NeuAc-(2“3)-b- -Gal-(1“3)-[a- -NeuAc-
D
D
0008-6215/00/$ - see front matter © 2000 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(99)00313-4

L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
133
(2“6)]-a-
D
-GalNAc-
L
-Ser/Thr. Activation of
and 4 to afford the hexasaccharide backbone
[11]. As the a- -NeuAc-(2“3)- -Gal linkage
human T-lymphocytes by anti-CD3 antibodies
and interleukin-2 results in an almost exclu-
sive expression of the title core 2 disialylated
D
D
appears in both the key intermediates 3 and 4,
we decided to construct this sequence first via
the building blocks 8, 9 and 10. The disaccha-
ride thus built was converted to its tri-
chloroacetimidate 5, which served as a com-
mon donor for the construction of 3 as well as
4 via coupling to the acceptors 6 and 7, re-
spectively. The azide groups, which are com-
patible with the sialic acid functionality, were
used to mask the amino groups of GlcNAc 6
and GalNAc 7 (Scheme 1).
hexasaccharide a-
(1“4)-b- -GlcNAc-(1“6) [a-
3)-b- -Gal-(1“3)]-a- -GalNAc-
D
-NeuAc-(2“3)-b-
-NeuAc-(2“
-Ser/Thr (1)
D-Gal-
D
D
D
D
L
[4]. Overexpression of core 2 O-glycans on
T-lymphocyte cell-surface glycoproteins has
been associated with certain pathological phe-
nomena including immunodeficient syndromes
[5] such as the Wiskott–Aldrich syndrome [6],
leukemia [7], AIDS [8], and malignant trans-
formation [9]. In these diseases the core 2
O-glycans are highly expressed on resting T
cells, whereas they are not expressed in nor-
mal, healthy individuals.
The synthesis of the hexasaccharide 2 com-
menced with the sialylation of the two galac-
topyranosyl acceptors 9 and 10 with the donor
8. The neighboring group assisted sialylation
of 9 and 10 with the donor 8 proceeded
stereoselectively under well-established condi-
tions [12] to afford the desired a-(2“3) sialyl-
ated disaccharides 12 and 13 in high yields.
The
D-galactopyranosyl derivative 9 having a
4-methoxyphenyl group at the anomeric posi-
tion was prepared as earlier reported [13].
Glycosylation of 9 with the donor 8 to the
disaccharide 12 and its subsequent conversion
to the trichloroacetimidate 5 was accom-
plished according to the reported protocol
[14]. Loss of material during oxidative cleav-
age of the anomeric 4-methoxyphenyl group
(77% yield) from the disaccharide 12 led us to
choose the galactopyranose acceptor 10 hav-
ing the tert-butyldiphenylsilyl ether at the
anomeric position, which can be efficiently
Heterogeneity and low abundance of gly-
copeptides preclude the isolation and charac-
terization of homogeneous glycans; therefore,
synthesis of well-defined glycopeptide frag-
ments remains the method of choice for struc-
ture–activity relationship studies. In the
framework of our project designed to eluci-
date the nature of the functional importance
of oligosaccharide structures on cell-surface
removed by the fluoride anion. The b- -galac-
D
topyranosyl derivative 10 was prepared from 9
by a series of standard functional group ma-
nipulations (1, Ac2O–pyridine, 0 °C–room
temperature, 10 h, 97%; 2, ceric ammonium
nitrate, CH3CN–H2O, 0 °C, 10 min, 77%; 3,
TBDPSCl, imidazole, 45 °C, 12 h, 94%; 4,
NaOMe, MeOH–toluene, 99%). The NIS–
TfOH-promoted sialylation of the acceptor 10
with the sialyl donor 8 afforded the disaccha-
ride 13 in 77% yield. Triphenyltin hydride (5
equivalents)-promoted desulfurization of 13
led exclusively to the lactone 15 in 80% yield,
thereby excluding the need for an additional
step of conversion of the ester 14 to lactone
15. Use of larger amounts of the promoter led
to a mixture of the desulfurized ester 14 and
glycoproteins, the protected -threonine con-
L
jugate 2 of the title disialylated core 2 hexa-
saccharide was constructed as a building block
for glycopeptide synthesis. There have been
earlier reports on the syntheses of the asialo
core 2 structures [10]; however, the sialylated
analog has not been reported so far. We
present here for the first time the total synthe-
sis of the completely sialylated core 2
hexasaccharide.
2. Results and discussion
The synthesis of the hexasaccharide 2 was
designed to combine the two trisaccharides 3

134
L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
lactone 15, which could be separated by chro-
matography but required an additional lac-
tonization reaction with DBU for conversion
of the benzyl ester 14 to the corresponding
lactone 15. Near-quantitative removal of the
anomeric silyl ether was achieved with tetra-
butylammonium fluoride in the presence of
acetic acid in THF to afford the hemiacetal
16. Subsequent conversion to the trichloroace-
timidate 5 was accomplished as before
(Scheme 2).
Glycosylation of the
D
-glucosamine accep-
tor 6 with the imidate 5 was then followed up.
The acceptor 6 was prepared in a manner
Scheme 1. TBDPS, t-butyldiphenylsilyl; Fmoc, 9-fluorenylmethoxycarbonyl; All, allyl; R, MP(4-methoxyphenyl) or TBDPS
(t-butyldiphenylsilyl); TBDMS, t-butyldimethylsilyl; CA, chloroacetyl.

L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
135
Scheme 2.
pared in three steps (1, hydrolysis of chloro-
acetate; 2, debenzylidenation; 3, regioselective
silylation) from 11. Glycosylation of the a
imidate 5 with 2 equivalents of 7 promoted by
BF3·OEt2 (0.8 equivalents) in 2:1 toluene–
CHCl3 at −15 °C gave the b-glycoside 21b
(54%) and the a-glycoside 21a (11%). The use
of Me3SiOTf as an alternative promoter for
the above reaction led to an in situ cleavage of
the tert-butyldimethylsilyl ether at the 6 posi-
tion of the GalNAc acceptor 7, followed by an
additional glycosylation at this position to
yield a pentasaccharide. The formation of the
pentasaccharide was confirmed by FABMS.
Compound 21b was desilylated (80% aqueous
CF3CO2H, 0 °C, 90%) to afford 4, which was
used as such as acceptor for the final glycosyl-
ation reaction (Scheme 4).
The hexasaccharidic backbone was con-
structed in a final glycosylation reaction of the
two key trisaccharidic intermediates 3 and 4.
The Me3SiOTf (0.5 equivalents)-promoted re-
action in CH2Cl2 at −40 °C furnished a mix-
ture of 22 and 23 (3:1) in 80% yield. To
facilitate separation, the mixture of the two
stereoisomers was treated with freshly distilled
thioacetic acid in pyridine to reduce the azide
into N-acetyl groups. Purification by column
chromatography gave the target hexasaccha-
ride 2 (68%) and 24 (22%) (Scheme 5). Use of
thioacetic acid contaminated with traces of
dithioacetic acid led to monothioacetylated
byproduct [18] along with the expected prod-
ucts. The structural assignments were made
similar to the one used to prepare the
analogous tert-butyldimethylsilyl ether [15]
with a slight modification. In this case sodium
cyanoborohydride-mediated reductive ring
opening of the 4,6-O-benzylidene precursor
afforded 6 regioselectively in 97% yield. The
Me3SiOTf-catalyzed reaction of the a-
trichloroacetimidate 5 with 6 afforded the
trisaccharides 17 (b:a=2:1) in 75% yield as an
inseparable mixture. The anomeric ratio of 17
was ascertained on the basis of the H-4b
1
(b-Gal) signals in the H NMR spectrum. The
borontrifluoride diethyletherate-mediated re-
action of 5 with 6 furnished 17 as a mixture
(b:a=6:1) in 29% yield. A major part of the
trichloroacetimidate was converted to its cor-
responding a and b fluorides. Glycosylation of
6 with this fluoride donor resulted in low
yields of the trisaccharide 17. Separation of
the trisaccharides 17 could be achieved after
cleavage of the anomeric silyl ether to afford
18b. This hemiacetal was then converted to its
trichloroacetimidate 3 as a 1:1 anomeric mix-
ture, which was used as such for the next
glycosylation (Scheme 3).
We next focused our attention on the con-
struction of the second trisaccharide 4. The
acceptor 7 was synthesized in a manner simi-
lar to that used for the analogous serine
derivative [14]. The known glycosyl fluoride 19
[12b] and Fmoc threonine allyl ester 20 [16]
were coupled in the presence of Cp2ZrCl2–
AgClO4 [17] to give a-glycoside 11 (76%). The
D-galactosamine-derived acceptor 7 was pre-

136
L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
Scheme 3.
Scheme 4.

L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
137
Scheme 5.
13
from C NMR measurements, and their com-
ppm downfield. FAB mass spectra were mea-
sured on a Jeol JMS-HX-110 mass spectrome-
ter. 3-Nitrobenzyl alcohol was used as the
matrix. Molecular sieves were purchased from
Nacarai Chemical Co. and activated at 180 °C
under vacuum, immediately prior to use. Gly-
cosylation reactions were performed under an
atmosphere of Ar in anhydr solvents.**
parison with those of the disaccharide, N-(9-
fluorenylmethoxycarbonyl)-O-[2-acetamido-4-
O-acetyl-3,6-di-O-benzyl-2-deoxy-b- -gluco-
D
pyranosyl-(1“6)-2-acetamido-3-O-acetyl-2-
deoxy-a-
D
-galactopyranosyl]- -threonine allyl
L
ester.
In conclusion, an efficient route to the syn-
thesis of core class 2 hexasaccharide 2 in a
fully protected form has been achieved for the
first time.
tert-Butyldiphenylsilyl-2,6-di-O-benzyl-i-
galactopyranoside (10).—To a soln of 3,4-di-
O-acetyl-2,6-di-O-benzyl- -galactopyranose
D
-
D
(4.67 g, 10.51 mmol) and imidazole (1.57 g,
23.12 mmol) in dry DMF (40 mL) was added
tert-butyldiphenylsilyl chloride (4.1 mL, 15.76
mmol), and the reaction mixture was allowed
to stir for 12 h. The reaction was quenched
with water, and the DMF was evaporated
under high vacuum. The dark residual oil was
extracted with EtOAc and washed successively
with water and brine, dried (Na2SO4), and
concentrated in vacuo. The crude product was
chromatographed on silica gel with 5:1 hex-
ane–EtOAc to 2:1 hexane–EtOAc to yield
6.74 g (94%) of the silyl ether. To a soln of the
silyl ether in MeOH–toluene (66–14 mL) was
added 0.04 equiv of 0.2 N NaOMe in MeOH
3. Experimental
General.—Optical rotations were deter-
mined with a Jasco DIP-370 polarimeter for
solutions in CHCl3. Column chromatography
was performed on Silica Gel 60 (E. Merck
70–230 mesh). Thin-layer chromatography
(TLC) was performed on Silica Gel 60 F254 (E.
Merck). 1H and 13C NMR spectra were
recorded on a Jeol a 400, a 600 MHz or
EX270 MHz spectrometer. Spectra were
recorded in CDCl3 using Me4Si as an internal
standard. Chemical shifts are expressed in

138
L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
1 H, J 9.4, J 4.1 Hz, H-3a), 3.30–3.38 (m, 1
H, H-2a), 3.30–3.38 (m, 1 H, H-6a), 3.14 (dd,
1 H, J 9.8, J 6.2 Hz, H-6a%), 2.80 (m, 1 H,
H-5a), 2.20 (dd, 1 H, J 4.8, J 13.3 Hz, H-3b
eq), 1.65 (m, 1 H, H-3b ax), 1.64 (s, 3 H,
NHCOCH3), 1.02 (s, 9 H, t-Bu). 13C NMR
(67.5 MHz): l 170.0, 165.5 (OCꢀO, NHCꢀO),
97.1 (C-1a), 95.3 (C-2b). Anal. Calcd for
C75H81NO13Si·0.5 H2O: C, 72.56; H, 6.66; N,
1.13. Found: C, 72.51; H, 6.60; N, 1.29.
and the reaction was allowed to stir for 12 h.
The reaction was neutralized with Amberlyst
15 (H+) resin, filtered, concentrated and chro-
matographed on silica gel with 1:1 hexane–
EtOAc to obtain 10 (5.86 g, 93%). Rf 0.51 (1:1
hexane–EtOAc); [h]D +21.7° (c 0.6); 1H
NMR (270 MHz): l 7.09–7.23 (m, 20 H, Ar),
4.88 (d, 1 H, J 11.3 Hz, OCH2Ph), 4.63 (d, 1
H, J 11.3 Hz, OCH2Ph), 4.46 (d, 1 H, J 7.4
Hz, H-1), 4.25 (s, 1 H, OCH2Ph), 4.24 (s, 1 H,
OCH2Ph), 3.70 (d, 1 H, J 2.9 Hz, H-4), 3.50
(m, 1 H, H-2), 3.44 (m, 1 H, H-6), 3.31 (m, 2
H, H-3, H-6%), 3.07 (m, 1 H, H-5), 1.02 (s, 9 H,
t-Bu). 13C NMR (67.5 MHz): l 97.9 (C-1),
81.0 (C-2), 74.7 (OCH2Ph), 73.4 (OCH2Ph),
73.1 (C-3), 72.9 (C-5), 69.1 (C-4), 69.0 (C-6).
Anal. Calcd for C36H42O6Si·0.5 H2O: C, 71.14;
H, 7.13. Found: C, 71.51; H, 7.11.
5 - Acetamido - 4,7,8,9 - tetra - O - benzyl - 3,5-
dideoxy-
D
-glycero-h- -galacto-2-nonulopyr-
D
anosylonic acid-(2“3)-2,6-di-O-benzyl-h and
i- -galactopyranosyl trichloroacetimidate-
D
(1b“4a)-lactone (5).—A soln of 15 (846 mg,
694 mmol), AcOH (600 mL, 10.5 mmol) and
TBAF (3.8 mL, 3.8 mmol, as its mol soln in
THF) in dry THF (15 mL) was stirred for 12
h at room temperature (rt). The reaction mix-
ture was then concentrated in vacuo, co-evap-
orated with toluene and chromatographed on
silica gel with 4:1 toluene–EtOAc to give the
hemiacetals 16 (685 mg, 99%). To a stirred
mixture of 16 (117 mg, 117 mmol) and DBU
(1.9 mL, 13.3 mmol) in dry CH2Cl2 (3 mL), was
added CCl3CN (118 mL, 1.17 mmol) at
−10 °C. After stirring for 2 h, the mixture
was chromatographed on silica gel 4:1
toluene–EtOAc to obtain 5 as two fractions
of pure a-imidate and a:b (1:9) mixture (94
mg, 30 mg, respectively, 93%). The physical
data of the product were identical to those
previously reported [14].
tert-Butyldiphenylsilyl-5-acetamido-4,7,8,9-
tetra-O-benzyl-3,5-dideoxy-
D
-glycero-h- -
D
galacto-2-nonulopyranosylonic acid-(2“3)-2,-
6-di-O-benzyl-i- -galactopyranoside-(1b“4a)-
D
lactone (15).—A mixture of 8 (1 g, 1.1 mmol),
,
10 (1 g, 1.66 mmol) and 3 A molecular sieves
(1 g) and NIS (375 mg 1.67 mmol) in dry
CH3CN (12 mL) was allowed to stir for 1 h at
−40 °C. To the cooled reaction mixture was
added TfOH (19.6 mL, 222 mmol). After 1 h
the reaction was quenched with aq NaHCO3,
filtered through Celite and concentrated in
vacuo. The residue was extracted with EtOAc
and washed successively with aq Na2S2O3, 1
M Na2CO3 and brine, dried (Na2SO4), concen-
trated and purified by silica gel column chro-
matography with 4:1 toluene–EtOAc to
obtain 1.23 g (77%) of 13. A mixture of 13
(115 mg, 80 mmol), Ph3SnH (140 mg, 400
mmol, as its 1 M soln in benzene) and AIBN
(3.6 mg, 22 mmol) in dry benzene (3 mL) were
heated at 90 °C for 1 h. An additional 1.5 mg
of AIBN was added after every 30 min for 3
h, and the reaction was allowed to stir at
90 °C for a total of 7 h. The reaction mixture
was then cooled to ambient temperature,
filtered through Celite, concentrated in vacuo,
and chromatographed on silica gel with 9:1
toluene–EtOAc to 4:1 toluene–EtOAc to ob-
tain 78 mg (80%) of 15. Rf 0.53 (7:3 toluene–
tert-Butyldiphenylsilyl-2-azido-2-deoxy-3,6-
di-O-benzyl-i- -glucopyranose (6).—To an
D
ice-cooled mixture of tert-butyldiphenylsilyl 2-
azido-2-deoxy-3-O-benzyl-4,6-O-benzylidene-
b- -glucopyranoside (3.3 g, 5.3 mmol),
D
,
NaBH3CN (3.33 g, 53 mmol) and 4 A molecu-
lar sieves in dry THF (100 mL) was slowly
added a mol soln of HCl in dioxane until
effervescence ceased. The reaction mixture
was then filtered through Celite and concen-
trated in vacuo. The residual syrup was ex-
tracted with EtOAc and washed successively
with aq NaHCO3, water and brine, dried
(Na2SO4), concentrated and purified by silica
gel column chromatography with 4:1 hexane–
EtOAc to 2:1 hexane–EtOAc to obtain 6 (3.2
g, 97%). Rf 0.35 (4:1 hexane–EtOAc); [h]D
1
EtOAc); [h]D +27.3° (c 0.1); H NMR (270
MHz): l 7.61–7.11 (m, 40 H, Ar), 5.01 (d, 1
H, J 3.4 Hz, H-4a), 4.25 (1 H, H-1a), 3.76 (dd,
1
−12.8° (c 1.1); H NMR (270 MHz): l 7.72–

L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
139
with EtOAc, concentrated in vacuo, extracted
with EtOAc and washed successively with wa-
ter and brine, dried (Na2SO4), concentrated
and purified by silica gel column chromatog-
raphy with 10:1 CHCl3–tert-butylmethyl
ether to obtain 18 as an anomeric mixture.
This mixture was purified by preparative TLC
using CHCl3–tert-butylmethyl ether to afford
the hemiacetals as two fractions consisting of
pure 18b and 18a (20 mg and 16 mg, respec-
tively, 94%). To a soln of 18b (14 mg, 10.3
mmol), CCl3CN (10.3 mL, 103 mmol) in
CH2Cl2 (1 mL) at −10 °C was added DBU
(0.3 mL, 2.05 mmol as a soln in CH2Cl2). The
reaction mixture was chromatographed on sil-
ica gel with 4:1 toluene–EtOAc to abtain 3 as
an a:b 1:1 mixture (14.8 mg, 96%). Rf 0.54 (7:3
7.23 (m, 40 H, Ar), 4.87 (d, 1 H, J 11.2 Hz,
OCH2Ph), 4.74 (d, 1 H, J 11.2 Hz, OCH2Ph),
4.43 (d, 1 H, J 11.8 Hz, OCH2Ph), 4.37 (d, 1
H, J 7.6 Hz, H-1), 4.35 (d, 1 H, J 11.8 Hz,
OCH2Ph), 3.64 (ddd, 1 H, J 2.3, J 9.5, J 8.9
Hz, H-4), 3.52–3.40 (m, 3 H, H-2, H-6, H-6%),
3.14 (dd, 1 H, J 8.9, J 9.9 Hz, H-3), 2.99 (m,
1 H, H-5), 2.55 (d, 1 H, J 2.3 Hz, OH), 1.11
(s, 9 H, t-Bu). Anal. Calcd for C36H41N3O5Si:
C, 69.31; H, 6.62; N, 6.74. Found: C, 69.29;
H, 6.65; N, 6.76.
tert-Butyldiphenylsilyl 5-acetamido-4,7,8,9-
tetra-O-benzyl-3,5-dideoxy-
D
-glycero-h-
acid-(2“3)-
-galactopyranosyl-
(1“4)-2-azido-2-deoxy-3,6-di-O-benzyl-i-
-glucopyranoside-(1b“4a)-lactone (17).—To
D
-
galacto-2-nonulopyranosylonic
2,6-di-O-benzyl-i- and -h-
D
D
1
,
a mixture of 6 (46 mg, 73.7 mmol) and 4 A
molecular sieves (50 mg) in 1:1 CH2Cl2–hex-
ane (1.2 mL) at −40 °C was added Me3SiOTf
(4.9 mL, 27.6 mmol). The mixture was allowed
to stir for 15 min, and to it was added 5 (63
mg, 55.3 mmol). The reaction was quenched
with NaHCO3, diluted with EtOAc and
filtered through Celite. The organic phase was
washed successively with aq NaHCO3 water
and brine, dried (Na2SO4), concentrated and
purified by silica gel column chromatography
with 4:1 toluene–EtOAc to afford 17 (66 mg,
75%, b:a=2:1). Rf 0.38 (4:1 toluene–EtOAc);
toluene–EtOAc); 3 H NMR (400 MHz): l
8.63 (s, 2 H, 2 CꢀNH), 6.27 (d, 1 H, J 3.4 Hz,
H-1a a), 5.50 (d, 1 H, J 8.2 Hz, H-1a b), 5.08
(d, 2 H, J 3.9 Hz, H-4b a,b), 2.32–2.21 (m, 2
H, H-3c eq a,b), 1.69 (s, 3 H, NHCOCH3),
1.68 (s, 3 H, NHCOCH3), 1.69 (m, H-3c ax
a,b).
N-(9-Fluorenylmethoxycarbonyl)-O-(2-azido-
4,6-O-benzylidene-3-O-chloroacetyl-2-deoxy-h-
D
-galactopyranosyl)- -threonine allyl ester
L
(11).—A mixture of Cp2ZrCl2 (2.1 g, 7.2
mmol), AgClO4 (1.48 g, 7.2 mmol), and dried,
,
powdered 4 A molecular sieves (5.5 g) in dry
1
17b: H NMR (400 MHz): l 5.15 (d, 1 H, J
CH2Cl2 (53 mL) was stirred at rt for 30 min
under Ar, then cooled in an ice–MeOH bath
(−15 °C). A soln of 19 (1.33 g, 3.6 mmol)
and 20 (1.60 g, 4.3 mmol) in dry CH2Cl2 (70
mL) was added, and the mixture was stirred
between −15 °C and rt overnight. After the
reaction was quenched with aq NaHCO3, the
mixture was diluted with EtOAc, and filtered
through Celite. The filtrate was washed with
water and brine, dried (Na2SO4), and concen-
trated in vacuo. The crude product was chro-
matographed on silica gel with 93:7
toluene–EtOAc to afford crystalline 11 (1.70
g, 76%). Compound 11: Rf 0.30 (9:1 toluene–
EtOAc); mp 78–80 °C; [h]D +161.2° (c 1.0);
1H NMR (400 MHz): l 7.8–7.3 (m, 13 H,
Ar), 5.94 (m, 1 H, ꢁCHꢀCH2), 5.69 (d, 1 H, J
9.6 Hz, NH), 5.56 (s, 1 H, PhCH(O)2), 5.36
(brd, 1 H, J 17.2 Hz, CHꢀCH2), 5.28 (brd, 1
H, J 10.2 Hz, CHꢀCH2), 5.12 (d, 1 H, J 3.6
Hz, H-1), 4.68 (brd, 2 H, J 5.6 Hz,
4.4 Hz, H-4b), 4.25–4.40 (m, H-1a, H-1b),
2.28 (dd, 1 H, J 5.1, J 13.4 Hz, H-3c eq), 1.75
(s, 3 H, NHCOCH3), 1.1 (s, 9 H, t-Bu); 13C
NMR (100 MHz): 101.7 (C-1b), 96.7 (C-1a),
95.3 (C-2c), 23.7 (CH3CꢀO), 37.7 (C-3c); 17a
1H NMR (400 MHz): l 5.52 (d, 1 H, J 3.9 Hz,
H-1b), 5.26 (d, 1 H, J 3.4 Hz, H-4b), 4.25–
4.40 (d, H-1a), 13C NMR (100 MHz): 96.8
(C-1a), 96.9 (C-1b), 95.3 (C-2c); FABMS
[M+Na]+ 1622.3.
5 - Acetamido - 4,7,8,9 - tetra - O - benzyl - 3,5-
dideoxy-
anosylonic acid-(2“3)-2,6-di-O-benzyl-i-
galactopyranosyl-(1“4)-2-azido-2-deoxy-3,6-
di-O-benzyl-i- -glucopyranosyl trichloroace-
D
-glycero-h-
D
-galacto-2-nonulopyr-
D-
D
timidate-(1c“4b)-lactone (3).—A mixture of
17 (45 mg, 28 mmol), AcOH, (24 mL, 422
mmol) and 1 M TBAF soln in THF (155 mL,
155 mmol) in dry THF (5 mL) was stirred at rt
for 12 h. The reaction mixture was diluted

140
L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
ꢁCH2CHꢀCH2), 4.16 (s, 2 H, ꢁCH2Cl), 4.02
(dd, 1 H, J 3.6, J 11.2 Hz, H-2), 3.82 (brs, 1
H, H-4), 1.33 (d. 3 H, J 6.3 Hz, ThrꢁCH3);
Anal. Calcd for C37H37ClN4O10: C, 60.61; H,
5.09; N, 7.64; Cl, 4.84. Found: C, 61.18; H,
5.17; N, 7.26; Cl, 4.91.
N - (9 - Fluorenylmethoxycarbonyl) - O - [(5-
acetamido-4,7,8,9-tetra-O-benzyl-3,5-dideoxy-
D
-glycero-h-
acid)-(2“3)-(2,6-di-O-benzyl-i- and h-
galactopyranosyl) - (1“3) - 2 - azido - 6 - O - tert-
butyldimethylsilyl-2-deoxy-h- -galactopyran-
osyl-(1c“4b)-lactone]- -threonine allyl ester
D
-galacto-2-nonulopyranosylonic
D
-
D
N-(9-Fluorenylmethoxycarbonyl)-O-(2-azido-
L
6-O-tert-butyldimethylsilyl-2-deoxy-h- -galac-
D
(21).—A mixture of 5 (50 mg, 44 mmol), 7 (36
mg, 53 mmol), and powdered AW 300 molecu-
lar sieves (550 mg) in 2:1 toluene–CH2Cl2 (2
mL) was stirred at rt for 1 h and then cooled
to −15 °C. To the mixture was added 0.2 M
BF3·OEt2 (176 mL, 35.2 mmol). The reaction
mixture was diluted with ether and filtered
through Celite. The filtrate was washed suc-
cessively with aq NaHCO3, water and brine,
dried (Na2SO4) and concentrated in vacuo.
The residue was chromatographed on silica
gel with 4:1 toluene–EtOAc to afford 21a (7.6
mg, 11%) and 21b (40 mg, 54%) and 7 (16 mg,
43% recovery). 21b: Rf 0.38 (4:1 toluene–
topyranosyl)- -threonine allyl ester (7).—A
L
stirred mixture of 11 (860 mg, 1.17 mmol) and
thiourea (500 mg, 6.45 mmol) in dry DMF (30
mL) was heated at 70 °C under Ar for 1 h.
After cooling, the mixture was concentrated in
vacuo. The residue was extracted with 1:1
ether–EtOAc, washed with water and brine,
dried (Na2SO4), concentrated, and purified by
silica gel column chromatography with 4:1
toluene–EtOAc to give the de-chloroacetyl-
ated product as crystals (630 mg, 82%); Rf
0.46 (7:3 toluene–EtOAc); mp 73–74 °C; [h]D
+125.0° (c 1.0).
A mixture of the crystals (130 mg, 198
mmol), 80% aq CF3CO2H (3 mL) and CH2Cl2
(1 mL) was stirred at 0 °C for 3 h, diluted with
water and toluene, and concentrated in vacuo.
The residue was extracted in EtOAc and
washed successively with aq NaHCO3, water
and brine, dried (Na2SO4), concentrated and
purified by silica gel column chromatography
with 10:1 CHCl3–MeOH to afford the triol
(109 mg), which was dissolved in DMF (1.75
mL) and stirred with tert-BuMe2SiCl (65 mL,
249 mmol) and imidazole (29 mg, 422 mmol)
for 4 h at 40 °C. The reaction was quenched
with water and concentrated in vacuo. The
residue was extracted in EtOAc and washed
successively with water and brine, dried
(Na2SO4), concentrated and purified by silica
gel column chromatography with 2:1 hexane–
EtOAc to afford 7 (139 mg, 87%). Rf 0.48
1
EtOAc); [h]D +18.7° (c 2.1); H NMR (400
MHz): l 7.75 (d, 2 H, J 7.3 Hz, Ar), 7.61
(d, 2 H, J 7.3 Hz, Ar) 5.92 (m, 1 H,
CH2CHꢀCH2), 5.68 (d, 1 H, J 9.8 Hz, NH),
5.35 (d, 1 H, J 16.8 Hz, CꢀCH2), 5.25 (d, 1 H,
J 10.6 Hz, ꢁCꢀCH2), 5.20 (d, 1 H, J 3.6 Hz,
H-4b), 5.03 (d, 1 H, J 3.6 Hz, H-1a), 4.42 (m,
H-1b), 2.19 (dd, 1 H, J 4.6, J 14.1 Hz, H-3c
eq), 1.69 (s, 3 H, NHCOCH3), 1.35 (d, 3 H, J
6.5 Hz, ThrꢁCH3), 0.86 (s, 9 H, t-Bu).13C
NMR (100 MHz): l 102.9 (C-1b, J 158.5 Hz,),
100.0 (C-1a, J 171.4 Hz), 95.4 (C-2c), 37.5
(C-3c), 23.6 (NHCOCH3), 19.0 (CH3ꢁThr).
FABMS [M+Na]+ 1681.3; 21a: Rf 0.39 (4:1
1
toluene–EtOAc); H NMR (400 MHz): l 7.72
(d, 2 H, J 7.6 Hz, Ar), 7.58 (d, 2 H, J 7.5 Hz,
Ar), 5.93 (m, 1 H, ꢁCHꢀCH2), 5.63 (d, 1 H, J
9.8 Hz, NHꢁThr), 5.34 (m, 2 H, H-4b,
ꢁCHꢀCHH%), 5.22 (d, J 10.3 Hz, ꢁCHꢀCHH%),
4.96 (d, 1 H, J 3.8 Hz, C-1a), 4.91 (d, 1 H, J
3.7 Hz, H-1b), 4.51 (m, H-3b), 4.02 (m, H-3a),
3.59 (dd, 1 H, J 3.8, J 9.6 Hz, H-2b), 3.42 (m,
H-2a), 2.40 (dd, 1 H, J 5.1, J 13.2 Hz, H-3c
eq), 1.87 (dd, 1 H, J 10.7, J 13.2 Hz, H-3c ax),
1.77 (s, 3 H, NHCOCH3), 0.90 (s, 9 H, t-Bu),
1
(10:1 CHCl3 –MeOH); [h]D +56.1° (c 3.3); H
NMR (270 MHz): l 7.76 (m, 2 H, Ar), 7.62
(m, 2 H, Ar), 7.40 (m, 2 H, Ar), 7.31 (m, 2 H,
Ar), 5.92 (m, 1 H, ꢁCHꢀCH2), 5.70 (d, J 9.5
Hz, NH), 5.36 (brd, 1 H, J 17.1 Hz, ꢁCꢀCH2),
5.27 (brd, 1 H, J 10.2 Hz, ꢁCꢀCH2), 4.97. (d,
1 H, J 3.6 Hz, H-1), 4.70 (d, 2 H, J 5.9 Hz,
CO2CH2), 3.52 (dd, J 3.6, J 10.2 Hz, H-2),
1.30 (d, 3 H, J 6.3 Hz, ThrꢁCH3), 0.90 (s, 9 H,
t-Bu), 0.10 (s, 6 H, 2 Me). Anal. Calcd for
C34H46N4O9Si·0.5 H2O: C, 59.03; H, 6.85; N,
8.10. Found: C, 58.89; H, 6.77; N, 7.99.
13
0.07 (s, 6 H, 2 CH3). C NMR (100 MHz): l
169.7, 169.6, 165.1 (OꢁCꢀO, NHCꢀO, CO2 all
not assigned), 99.8 (C-1a), 95.2 (C-1b), 95.1
(C-2c), 38.1 (C-2c), 25.9 (t-Bu), 23.8
(NHCOCH3), 18.9 (CH3ꢁThr).

L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
141
dried (Na2SO4), concentrated in vacuo, and
purified by column chromatography on silica
gel with 1:1 hexane–EtOAc to afford 22 and
23 as a mixture (20 mg, b:a=3:1, 80%). Com-
pounds 22 and 23 (9 mg, 3.1 mmol) were
stirred with freshly distilled AcSH (152 mL)
and pyridine (76 mL) for 12 h under a N2
atmosphere. The reaction mixture was diluted
and co-evaporated with toluene, and the resid-
ual yellow liquid was purified by column chro-
matography on silica gel over a gradient of
1:1, 1:2, 1:3 hexane–EtOAc to afford 2 (6.2
mg, 68%) and 24 (2.0 mg, 22%). 2 Rf 0.30 (1:3
hexane–EtOAc); [h]D +27.5° (c 0.4); 1H
NMR (600 MHz): l 7.74 (d, 2 H, J 7.3 Hz,
Ar), 7.61 (m, 2 H, Ar) 5.85 (m, 1 H,
CHꢀCH2), 5.32 (d, 2 H, J 17.1 Hz, ꢁCꢀCH2),
5.27 (d, 2 H, J 10.2 Hz, ꢀCH2), 5.14 (d, 1 H J
3.4 Hz, H-4, Gal), 5.12 (d, J 3.4 Hz, H-4,
Gal), 4.81 (m, 2 H, H-1, GlcNAc, H-1, Gal-
NAc), 3.88–4.02 (m, H-4, NeuNAc, H-4, Neu-
NAc% not assigned), 2.29 (m, 1 H, H-3eq,
NeuNAc), 2.07 (m, 1 H, H-3eq, NeuNAc%)
1.84 (s, 3 H, NHCOCH3), 1.82 (s, 3 H,
NHCOCH3), 1.75 (s, 3 H, NHCOCH3), 1.70
(s, 3 H, NHCOCH3), 1.77 (t, 1 H, J 10.7
H-3ax, NeuNAc), 1.62 (t, 1 H, J 10.7 H-3ax,
NeuNAc%), 1.28 (3 H, ThrꢁCH3). 13C NMR
(150 MHz): l 103.4, 102.0 (C-1 Gal, C-1, Gal%
not assigned), 100.6 (C-1, GalNAc), 99.8 (C-1,
GlcNAc), 95.3 (C-2, NeuAc, C-2, NeuAc% not
assigned), 23.4, 23.6, 23.7 (NHCOCH3×4),
18.3 (ThrꢁCH3). FABMS [M+1]+ 2920.5,
[M+Na]+ 2942.9; 24 Rf 0.40 (1:3 hexane–
N - (9 - Fluorenylmethoxycarbonyl) - O - [(5-
acetamido-4,7,8,9-tetra-O-benzyl-3,5-dideoxy-
D
-glycero-h-
acid)-(2“3)-(2,6-di-O-benzyl-i-
pyranosyl) - (1“3) - 2 - azido - 2 - deoxy - h -
galactopyranosyl-(1c“4b)-lactone]- -threonine
D
-galacto-2-nonulopyranosylonic
D
-galacto-
D-
L
allyl ester (4).—To a soln of 21b (37 mg, 21
mmol) in CH2Cl2 at 0 °C was added 80% aq
CF3CO2H (2 mL) and the soln was stirred for
3 h. It was then neutralized with solid
NaHCO3, diluted, and extracted with EtOAc.
The organic phase was washed successively
with water and brine, dried (Na2SO4), concen-
trated in vacuo and purified by column chro-
matography on silica gel with 3:2 toluene–
EtOAc to afford 4 (28.8 mg, 90%). Rf 0.35 (3:2
toluene–EtOAc); [h]D +65.1° (c 0.7); 1H
NMR (400 MHz): l 7.68 (d, 2 H, J 7.4 Hz,
Ar), 7.54 (d, 2 H, J 7.2 Hz, Ar), 5.75 (m, 1 H,
ꢁCHꢀCH2), 5.63 (d, 1 H, J 9.5 Hz, NH), 5.28
(d, 1 H, J 16.9 Hz, ꢁCꢀCH2), 5.18 (d, 1 H, J
10.1 Hz, ꢀCH2), 5.18 (d, 1 H, J 3.8 Hz, H-4b),
4.96 (d, 1 H, J 3.6 Hz, H-1a), 2.13 (dd, 1 H, J
4.8, J 13.5 Hz, H-3c eq.), 1.61 (m, H-3c ax),
1.60 (s, 3 H, NHCOCH3). 1.24 (d, 3 H, J 6.2
13
Hz, ThrꢁCH3); C NMR (100 MHz): l 170.5,
170.4, 165.4 (NHCO, OꢁCꢀO, CO2All), 103.1
(C-1b), 100.0 (C-1a), 95.9 (C-2c), 23.9
(NHCOCH3), 19.2 (CH3ꢁThr). Anal. Calcd
for C87H93N5O21: C, 67.64; H, 6.06; N, 4.53.
Found: C, 67.23; H, 6.19; N, 4.32.
N - (9 - Fluorenylmethoxycarbonyl) - O - {(5-
acetamido-4,7,8,9-tetra-O-benzyl-3,5-dideoxy-
D
-glycero-h-
acid)-(2“3)-(2,6-di-O-benzyl-i-
pyranosyl)-(1“4)-2-acetamido-2-deoxy-3,6-
di - O - benzyl - i - - glucopyranosyl - (1“6)-
[(5 - acetamido - 4,7,8,9 - tetra - O - benzyl - 3,5-
dideoxy- -glycero-h- -galacto-2-nonulopyr-
anosylonic acid)-(2“3)-2,6-di-O-benzyl-i-
galactopyranosyl - (1“3)] - 2 - acetamido - 2-
deoxy-h- -galactopyranosyl-(1d“4c:1f“4e)-
dilactone}- -threonine allyl ester (2).—To a
D
-galacto-2-nonulopyranosylonic
1
D
-galacto-
EtOAc); [h]D +33.9° (c 0.2); H NMR (600
MHz): l 7.77 (m, 2 H, Ar), 7.63 (m, 2 H, Ar)
5.72 (m, 1 H, CHꢀCH2), 5.22 (m, 2 H,
ꢁCHꢀCH2), 5.15 (d, J 3.4 Hz, H-4, Gal), 5.11
(d, J 2.9 Hz, H-4, Gal%), 4.75–4.81 (m, 2 H,
H-1, GlcNAc, H-1, GalNAc not assigned),
3.97–4.78 (m, H-4, NeuNAc, H-4, NeuNAc%
not assigned), 2.30 (m, 1 H, H-3eq, NeuNAc),
2.05 (m, 1 H, H-3eq, NeuNAc%), 1.83 (s, 3 H,
NHCOCH3), 1.81 (s, 3 H, NHCOCH3), 1.75
(s, 3 H, NHCOCH3), 1.75 (m, H-3ax, Neu-
NAc), 1.69 (s, 3 H, NHCOCH3), 1.58 (m, 1 H,
H-3ax, NeuNAc), 1.27 (3 H, ThrꢁCH3). 13C
NMR (150 MHz): l 103.3, 102.2 (C-1 Gal,
C-1 Gal% not assigned), 100.6 (C-1, GlcNAc),
97.2 (C-1, GalNAc), 95.3 (C-2, NeuAc, C-2,
NeuAc% not assigned), 23.7, 23.5, 23.4, 23.0
D
D
D
D-
D
L
mixture of 3 (13 mg, 8.6 mmol), 4 (15.3 mg, 9.9
mmol) and 4 A molecular sieves (60 mg) in
,
CH2Cl2 (1.6 mL) at −40 °C was added
Me3SiOTf (0.08 mL, 4.6 mmol, as its soln in
CH2Cl2). After 1 h the reaction was quenched
with NaHCO3, diluted with EtOAc and
filtered through Celite. The organic phase was
washed successively with water and brine,

142
L. Singh et al. / Carbohydrate Research 325 (2000) 132–142
[8] (a) J.-C. Lefebvre, V. Giordanengo, M. Limouse, A.
Doglio, M. Cucchiarini, F. Monpoux, R. Mariani, J.-F.
Peyron, J. Exp. Med., 180 (1994) 1609–1617. (b) B.
Ardman, M.A. Sikorski, M. Settles, D.E. Staunton, J.
Exp. Med., 172 (1990) 1151–1158.
(NHCOCH3×4), 18.5 (ThrꢁCH3). FABMS
[M+Na]+ 2942.9.
[9] S. Yousefi, E. Higgins, Z. Daoling, A. Pollex-Kruger, O.
Hindsgaul, J.W. Dennis, J. Biol. Chem., 266 (1991)
1772–1782.
Acknowledgements
[10] (a) Z.-G. Wang, X.-F. Zhang, Y. Ito, Y. Nakahara, T.
Ogawa, Carbohydr. Res., 295 (1996) 25–39. (b) E. Mein-
johanns, M. Meldal, A. Schleyer, H. Paulsen, K. Bock, J.
Chem. Soc., Perkin Trans. 1, (1996) 985–983. (c) D. Qiu,
R.R. Koganty, Tetrahedron Lett., 38 (1997) 45–48. (d)
N. Mathieux, H. Paulsen, M. Meldal, K. Bock, J. Chem.
Soc., Perkin Trans. 1, (1997) 2359–2368. (e) W. Kinzy,
R.R. Schimdt, Carbohydr. Res., 166 (1987) 265–276. (f)
H. Paulsen, W. Rauwald, U. Weichert, Justus Liebigs
Ann. Chem., (1988) 75–86.
We are grateful to Dr J. Uzawa, Dr H.
Koshino, and T. Chijimatsu for the NMR and
Dr Y. Ohnishi for the FABMS measurements.
We thank A. Takahashi for technical
assistance.
References
[11] L. Singh, Y. Nakahara, Y. Ito, Y. Nakahara, Tetrahe-
dron Lett., 40 (1999) 3769–3772.
[1] (a) M. Fukuda, Cancer Res., 56 (1996) 2237–2244. (b) T.
Feizi, Nature, 314 (1985) 53–57. (c) S. Hakomori, Ann.
Re6. Immunol., 2 (1984) 103–126.
[2] T.W. Rademacher, R.B. Parekh, R.A. Dwek, Ann. Re6.
Biochem., 57 (1988) 785–838.
[3] (a) S.R. Carlsson, M. Fukuda, J. Biol. Chem., 261 (1986)
12,779–12,786. (b) A. Bettaieb, F. Farace, M.T. Mit-
javila, Z. Mishal, M.C. Dokhelar, T. Tursz, J. Breton-
Gorius, W. Vainchenkerk, N. Kieffer, Blood, 71 (1988)
1226–1233. (c) S.R. Carlsson, H. Sasaki, M. Fukuda, J.
Biol. Chem., 261 (1986) 12,787–12,795.
[12] (a) Y. Ito, T. Ogawa, Tetrahedron, 46 (1990) 89–102. (b)
Z.-G. Wang, X.-F. Zhang, Y. Ito, Y. Nakahara, T.
Ogawa, Bioorg. Med. Chem., 4 (1996) 1901–1908.
[13] (a) C. Murakata, T. Ogawa, Carbohydr. Res., 235 (1992)
95–114. (b) Y. Nakahara, H. Iijima, T. Ogawa, Tetrahe-
dron Lett., 35 (1994) 3321–3324.
[14] Y. Nakahara, Y. Nakahara, Y. Ito, T. Ogawa, Carbo-
hydr. Res., 309 (1998) 287–296.
[15] (a) W. Kinzy, R.R. Schmidt, Liebigs Ann. Chem., (1985)
1537–1545. (b) C. Murakata, T. Ogawa, Carbohydr.
Res., 234 (1992) 75–91.
[4] F. Piller, V. Piller, R.I. Fox, M. Fukuda, J. Biol. Chem.,
263 (1988) 15,146–15,150.
[5] S. Tsuboi, M. Fukuda, J. Biol. Chem., 273 (1998)
30,680–30,687.
[16] (a) B.G. de la Torre, J.L. Torres, E. Bardaji, P. Clape´s,
N. Xaus, X. Jorba, S. Calvet, F. Albericio, G. Valentia,
J. Chem. Soc., Chem. Commun., (1990) 965–967. (b) M.
Ciommer, H. Kunz, Synlett, (1991) 593–595.
[6] E.A. Higgins, K.A. Siminovitch, D. Zhuang, I. Brock-
hausen, J.W. Dennis, J. Biol. Chem., 266 (1991) 6280–
6290.
[7] (a) O. Saitoh, F. Piller, R.I. Fox, M. Fukuda, Blood, 77
(1991) 1491–1499. (b) I. Brockhausen, W. Kuhns, H.
Schachter, K.L. Matta, D.R. Sutherland, M.A. Baker,
Cancer Res., 51 (1991) 1257–1263.
[17] (a) K. Suzuki, H. Maeta, T. Matsumoto, G. Tsuchihashi,
Tetrahedron Lett., 29 (1988) 3567–3570. (b) K. Suzuki,
H. Maeta, T. Matsumoto, G. Tsuchihashi, Tetrahedron
Lett., 29 (1988) 3571–3574. (c) K. Suzuki, H. Maeta, T.
Matsumoto, Tetrahedron Lett., 30 (1989) 4853–4856.
[18] H. Paulsen, T. Biedfeldt, S. Peters, M. Meldal, K. Bock,
Justus Liebigs Ann. Chem., (1994) 369–379.
.