Synthesis of 5′-O-(2-azido-2-deoxy-α-D-glycosyl)nucleosides and their antitumor activities

Article abstract of DOI:10.1002/hlca.200390163

The 1,3,4,6-tetra-O-acetyl-2-azido-2-deoxy-β-D-mannopyranose (4) or the mixture of 1,3,6-tri-O-acetyl-2-azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-β-D- galactopyranosyl)-β-D-mannopyranose (10) and the corresponding a-D-glucopyranose-type glycosyl donor 9/1

Full text of DOI:10.1002/hlca.200390163

Helvetica Chimica Acta Vol. 86 (2003)
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Synthesis of 5'-O-(2-Azido-2-deoxy-a-d-glycosyl)nucleosides and Their
Antitumor Activities
by Gui-Sheng Zhang, Jie Chen, Ji-Mei Min, and Li-He Zhang*
National Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University,
Beijing 100083, P.R. China
The 1,3,4,6-tetra-O-acetyl-2-azido-2-deoxy-b-d-mannopyranose (4) or the mixture of 1,3,6-tri-O-acetyl-2-
azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl)-b-d-mannopyranose (10) and the correspond-
ing a-d-glucopyranose-type glycosyl donor 9/10 reacted at room temperature with protected nucleosides 12 15
in CH2Cl2 solution in the presence of BF3 ¥ OEt2 as promoter to give 5'-O-(2-azido-2-deoxy-a-d-glycosyl)nu-
cleosides in reasonable yields (Schemes 2 and 3). Only the 5'-O-(a-d-mannopyranosyl)nucleosides were
obtained. Compounds 21, 28, 30, and 31 showed growth inhibition of HeLa cells and hepatoma Bel-7402 cells at
a concentration of 10 mm in vitro.
1. Introduction. Nucleoside analogues play an important role in antiviral and
anticancer chemotherapy. Among numerous nucleoside analogs, the nucleosides
connected to oligosaccharides have received much attention owing to their profound
biological activities. There are many carbohydrate receptors on the cell surface; e.g.,
the asialoglycoprotein receptor exists on the surface of hepatocytes and can bind to
terminal galactose residues [1]. Some activities of glycoproteins related to the
attachment, migration, proliferation of cancer cells, and the repeating units of
polysaccharides have been investigated for the prevention of tumor metastasis [2]. It
has been found that many natural antibiotics possessing significant antitumor and
antiviral activities contain O-glycosylated nucleoside substructures [3 5]. It would be
interesting to study whether the glycosyl residue could be used as a targeting moiety
towards glycosyl-binding proteins. In this way, some glycosylated antitumor or antiviral
nucleoside analogues might be expected to have a higher therapeutic index together
with lesser side effects and toxicity. We have reported the syntheses of galactosyl
phosphate diester derivatives of nucleoside [6], lactosyl phosphate diester derivatives
of nucleoside [7], and 5'-O-glycosylnucleosides [8]. It was found that the galactosyl
phosphate diesters of arabinosylcytosine were active in the human cytomegalovirus
(HCMV) assay. The IC50 of some compounds for anti-HCMVactivity were 0.1 0.2 mm
[6]. Amino and azido groups are potential active groups in many nucleoside analogues
possessing bioactivities, such as AZT, ezomycin A1, and liposidomycin C, etc. [9]. We
report here the investigation of the synthesis and antitumor activity of nucleoside
analogues containing azido-oligosaccharides.
2. Results and Discussion. Glycal is a very versatile synthetic intermediate and has
proven to be useful in the synthesis of glycoconjugates. The common method for the
synthesis of 2-azido-2-deoxy-sugars consists of the introduction of an azido group at

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Helvetica Chimica Acta Vol. 86 (2003)
C(2) of a per-O-acetylglycal [10 14] by addition of sodium azide in the presence of
cerium(IV) ammonium nitrate (CAN); subsequently, the C(1)-nitro group can be
substituted by an acetoxy group via the reaction with AcOH and NaOAc at 1008. This is
illustrated by the conversion of 1 to 2 4 (see Scheme 1 and below). However, the
azido-nitration usually yields a product mixture that is difficult to separate. Thus, 3,4-di-
O-acetyl-2,6-anhydro-5-deoxy-d-lyxo-hexo-5-enonate gave an inseparable mixture of
azido nitrates on treatment with CAN and sodium azide in MeCN at À158 under N2
[14]; after treatment with NaOAc in glacial AcOH, the NMR spectrum of the
corresponding azido acetate mixture indicated the presence of a-d-glucose, b-d-
glucose, and a-d-mannose-type anomers in the ratio 5 :2 :1. In the case of hexa-O-
acetyl-d-lactal (5), the three possible products 6 8 were obtained, namely of the d-
glucose and d-mannose type, respectively, depending on the different orientation of the
azido group at C(2) (Scheme 1). Arnap and Lˆnngren [12] reported that compounds 6
and 7 were obtained in the ratio 8:1. The d-mannose-type product 7 was the minor
component in the mixture of these isomers.
Scheme 1
We performed the azido-nitration/acetolysis sequence with tri-O-acetyl-d-glucal 1
and obtained the azido acetates 2/3 and 4 (Scheme 1). However, when the glycosyl
donors 2/3 (d-glucose type) or 4 (d-mannose type) were treated with glycosyl receptor

Helvetica Chimica Acta Vol. 86 (2003)
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12 (see below), only the reaction of the d-mannose-type glycosyl donor 4 proceeded in
good yield. Thus, the azido-nitration conditions were optimized to increase the
formation of d-mannose-type isomer 4 from 1. The ratio of the isomeric azido nitrate
precursors of 2 4 varied with temperature, and the d-mannose-type precursor of 4
was the major product at room temperature. Therefore, 1 was treated with CAN and
sodium azide at room temperature under Ar, and the crude azido nitrates were then
treated with anhydrous NaOAc in AcOH at 1008 to give a mixture 2/3/4 in a ratio of ca.
1
1.75 :1.00 :2.12 (by H-NMR). After chromatography (silica gel), 4 and 2/3 were
obtained in 26.6 and 27% yields, respectively. Similarly, 5 prepared from lactose [15]
was submitted to the same azido-nitration/acetolysis procedure; the mixture 6/7 could
be separated from 8 by chromatography after the reaction. Then, the mixture 6/7 was
1
treated with NaOAc in AcOH at 1008 to give 9/10 (ca. 1:1.1 by H-NMR) in 18.4%
yield from lactose (Scheme 1). The mixture 9/10 could not be separated by
chromatography.
The protected nucleosides 12 15 [16][17] were glycosylated with donor 4 or 9/10 in
CH2Cl2 solution at room temperature in the presence of BF3 ¥ Et2O (Schemes 2 and 3).
No reaction occurred when the mixture 2/3 was treated with 12 15 by this same
procedure. With donor 4, glycosylnucleosides 16 19 were obtained from 12 15 in 86.5,
65.0, 51.0, and 79.7% yield, respectively (Scheme 2). Of the mixture 9/10, only the d-
mannose-type glycosyl donor 10 reacted with 12 15 to give the corresponding glycosyl
nucleosides 24 27 in 46.7, 42.6, 83.0, and 65.2% yield, respectively (Scheme 3); the
unreacted 9 could be recovered from the reaction mixture. No reaction occurred when
the recovered 9 was resubmitted to the same procedure. The NMR data of compounds
16 19 and 24 27 confirmed the a-d-configuration (see Table 1) [13][18] of the
glycosyl bonds formed in above reaction. It is suggested that the formation of 16 19
and 24 27 occurs via an SN1 mechanism through an oxacarbenium cation as the
reaction intermediate, and the a-d-selectivity of the mannosylation is the result of a
steric effect of the axial substituent at C(2), and the steric hindrance leads to the failure
of the reactions of glycosyl donors 2/3 or 9 with the nucleosides.
Table 1. 1H-NMR Coupling Constants J(1'',2'') and 13C-NMR Chemical Shifts of Glycosylnucleosides 16 19 and
24 27 in CDCl3
16
1.5
98.21
17
18
19
1.5
98.16
24
2.0
98.19
25
1.5
98.18
26
1.75
97.97
27
2.0
97.96
J(1'',2'') [Hz]
d(C(1'')) [ppm]
s
s
98.14
98.12
The protected glycosylation products 16 19 and 24 27 were treated with 25 30%
MeNH2 in abs. EtOH at room temperature to give the target compounds 20 23
(Scheme 2) and 28 31 (Scheme 3), respectively.
The results indicate that the reactivity of the d-mannose-type glycosyl donors 4 and
10 lead to a convenient and efficient approach to the synthesis of azido-oligoglycosyl-
nucleosides with the advantages of mild conditions, easy workup, high selectivity, and
reasonable yields.
Compounds 20 23 and 28 31 were evaluated for their antitumor activities by cell-
culture bioassay. Compounds 21, 28, and 30 showed inhibition of the growth of Hela

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Helvetica Chimica Acta Vol. 86 (2003)
Scheme 2
cells by 31.92, 42.41, and 37.86%, respectively, at a concentration of 10 mm, and
compound 31 indicated a 46.05% growth inhibition of hepatoma Bel-7402 cells at a
concentration of 10 mm (Table 2).
Table 2. Percentage of Growth Inhibition of HeLa Cells and Hepatoma Bel-7402 Cells by Glycosylnucleosides
20 23 and 28 31 at a Concentration of 10 mma)
20
21
22
23
28
29
30
31
HeLa
4.03
31.92
6.86
0.10
42.41
16.26
37.86
16.28
Bel-7402
7.484.96
6.19
6.71
13.76
4.33
20.07
46.05
a) HeLa and Bel-7402 cells were seeded in 96-well culture plates at a concentration of 5000/well. After overnight
culture, the compounds were dissolved in sterilized phosphate-buffered saline and added to the well. After 5 h
incubation at 378, the inhibition of the cell growth was measured by the MTT method [19].
Experimental Part
1. General. CC ˆ Column chromotography. TLC: silica gel GF-254 (Qing Dao Chemical Company, China)
plates with detection by UV, or charting with 5% phosphomolybdic acid hydrate in EtOH. Optical rotations:
Perkin-Elmer 243B polarimeter. IR Spectra: DE-983G spectrophotometer; KBr pellets; in cmÀ1. NMR Spectra:
Varian VXR-300 or Varian INOVA-500; d in ppm rel. to SiMe4 as an internal standard J in Hz. MS: PE SCLEX-
QSTAR and Autospec UltimaETOF spectrometers; in m/z.

Helvetica Chimica Acta Vol. 86 (2003)
2077
Scheme 3
2. 1,3,4,6-Tetra-O-acetyl-2-azido-2-deoxy-b-d-mannopyranose (4). A soln. of 1 (2.11 g, 7.7 mmol) in dry
MeCN (100 ml) was added to a mixture of CAN (12.65 g, 28.1 mmol) and NaN3 (0.75 g, 11.5 mol) in MeCN
(30 ml), which was cooled in an ice-salt bath, with vigorous stirring under Ar. Then, the mixture was kept for
24 h at r.t. Et2O (100 ml) and ice-cold H2O (50 ml) were added, and the org. phase was washed with H2O (3 Â
50 ml) and evaporated to give a syrup (2.2 g).
A soln. of the above syrup (2.1 g) in AcOH (15 ml) containing anh. NaOAc (1.0 g) was heated at 1008 for
2 h and then cooled. The mixture was extracted with CH2Cl2 (100 ml), the extract washed with H2O, aq.
NaHCO3 soln., and H2O, dried (MgSO4), and evaporated: syrup (1.87 g) containing 2, 3, and 4 (ca.
1.75 :1.00 :2.12 by 1H-NMR (CDCl3): 6.21 (d, J(1,2) ˆ 3.54, HÀC(1) of 2); 5.80 (d, J(1,2) ˆ 8.55, HÀC(1) of 3);
5.96 (d, J(1,2) ˆ 1.43, HÀC(4) of 4). The syrup was purified by CC (silica gel, petroleum ether (60 908)/AcOEt
4 :1): 2/3 (753 mg, 27% from 1) and 4 (743 mg, 26.6% from 1). 2/3: 1H-NMR (CDCl3): 6.30 (d, J(1,2) ˆ 3.55,
1
HÀC(1) of 2); 5.56 (d, J(1,2) ˆ 8.55, HÀC(1) of 3). 4: H-NMR (CDCl3): 6.12 (d, J(1,2) ˆ 1.3, HÀC(1)).
3. 1,3,6-Tri-O-acetyl-2-azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl)-a-d-glucopyranose
(9) and 1,3,6-Tri-O-acetyl-2-azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl)-b-d-mannopyra-
nose (10). Lactose (5.86 g, 32.52 mmol) was treated according to a published method [15] to give a syrup 5
(13.3 g). The crude 5 in MeCN (50 ml) was added to a mixture of CAN (31.85 g, 70.7 mmol) and NaN3 (2.25 g,
34.5 mmol) in MeCN (30 ml), which was cooled in an ice-salt bath, with vigorously stirring under Ar. Then, the
mixture was kept for 24 h at r.t. Et2O (250 ml) and ice-cold H2O (50 ml) were added, and the org. phase was

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Helvetica Chimica Acta Vol. 86 (2003)
washed with H2O (3 Â 50 ml) and evaporated. The residue was purified by CC (silica gel, petroleum ether (60
908)/AcOEt 3 :2): 8 and 6/7.
The mixture 6/7 obtained above in AcOH (60 ml) containing anh. NaOAc (4.0 g) was heated at 1008 for 2 h
and then cooled. The mixture was extracted with CH2Cl2 (300 ml), the extract washed with H2O, aq. NaHCO3
soln., and H2O, dried (MgSO4), and evaporated, and the syrup obtained was purified by CC (silica gel,
petroleum ether (60 908)/AcOEt 4 :1): 9/10 (3.7 g, 18.4% from lactose) (ca. 1.0 :1.1 by 1H-NMR (CDCl3): 6.14
(d, J(1,2) ˆ 3.9, HÀC(1) of 9); 5.86 (br., HÀC(1) of 10).
4. Glycosylations of Nucleosides 12 15: General Procedure. A soln. of one of the glycosyl receptors 12 15
(50 mg) and 2 equiv. of glycosyl donor 4 or 3 equiv. of 9/10 in a soln. of dry CH2Cl2 (5 ml) and BF3 ¥ OEt2
(0.2 ml) was stirred at r.t. until the disappearance of the glycosyl receptor (TLC monitoring). After cooling in an
ice bath and neutralization with sat. NaHCO3 soln., the mixture was extracted with CH2Cl2, the org. layer was
dried (Na2SO4) and evaporated, and the residue purified by CC (silica gel): 16 19 or 24 27, resp.
5'-O-(3,4,6-Tri-O-acetyl-2-azido-2-deoxy-a-d-mannopyranosyl)uridine 2',3'-Diacetate (16). Yield 86.5%.
White solid. [a]D18 ˆ À42 (c ˆ 0.024, H2O). 1H-NMR (CDCl3): 8.67 (s, NH); 7.42 (d, J(5,6) ˆ 8.0, HÀC(6)); 6.06
(d, J(1',2') ˆ 5.5, HÀC(1')); 5.97 (d, HÀC(5)); 4.92 (d, J(1'',2'') ˆ 1.5, HÀC(1'')); 5.45 5.27 (m, HÀC(3''),
HÀC(3'), HÀC(2'), HÀC(4'')); 4.26 (m, HaÀC(6''), HÀC(4')); 4.16 (m, HÀC(2'')); 4.11 (m, HbÀC(6'')); 4.01
3.96 (m, HÀC(5''), HaÀC(5')); 3.81 (m, HbÀC(5')); 2.12 2.08(5 s, 5 AcO). 13C-NMR (CDCl3): 170.64 162.50
(CˆO); 150.08(C(6)); 139.71 (C(4)); 103.92 (C(5)); 98.21 (C(1 '')); 87.59 (C(1')); 80.69 (C(4')); 72.67 (C(3'));
71.22 (C(3'')); 69.95 (C(2')); 69.40 (C(5'')); 66.69 (C(5'')); 65.09 (C(4'')); 62.01 (C(6'')); 60.92 (C(2'')); 20.73
‡
(MeCO). ESI-TOF-MS: 664.0792 ([M ‡ Na] ).
N4-Acetyl-5'-O-(3,4,6-tri-O-acetyl-2-azido-2-deoxy-a-d-mannopyranosyl)cytidine 2',3'-Diacetate (17):
Yield 65%. Colorless. [a]1D8 ˆ ‡4.4 (c ˆ 0.031, CHCl3). IR: 2111.15 (N3). 1H-NMR (CDCl3): 10.01 (br., AcNH);
7.90 (d, J(5,6) ˆ 6.5, HÀC(6)); 7.56 (d, HÀC(5)); 6.12 (d, J(1',2') ˆ 1.5, HÀC(1')); 5.38( m, HÀC(2'),
HÀC(3')); 5.35 (dd, J(3'',4'') ˆ 10.0, J(4'',5'') ˆ 12.0, HÀC(4'')); 5.27 (dd, J(2'',3'') ˆ 8.5, HÀC(3'')); 4.91
(s, HÀC(1'')); 4.31 (br., HÀC(4')); 4.23 (m, 1 H, HaÀC(6'')); 4.15 (m, HÀC(2'')); 4.05 (m, HbÀC(6')); 3.96
(m, HaÀC(5'), HÀC(5'')); 3.84 (m, HbÀC(5')); 2.27 2.04 (6s, 18H, 5 AcO, 1 AcNH). 13C-NMR (CDCl3):
170.64 163.16 (6 CˆO); 144.26 (C(6)); 98.14 (C(1'')); 97.74 (C(5)); 89.03 (C(1')); 80.53 (C(4')); 73.62 (C(3'));
71.00 (C(3'')); 69.64 (C(2')); 69.18(C(5 '')); 66.52 (C(5')); 65.23 (C(4'')); 61.96 (C(6'')); 60.91 (C(2'')); 24.87
‡
(MeCONH); 20.66 20.39 (5 MeCO). ESI-TOF-MS: 681.3033 ([M À 1] ).
5'-O-(3,4,6-Tri-O-acetyl-2-azido-2-deoxy-a-d-mannopyranosyl)thymidine 3'-Acetate (18): Yield 51.0%.
White solid. [a]1D8 ˆ À52.9 (c ˆ 0.021, H2O). 1H-NMR (CDCl3): 9.04 (s, NH); 7.23 (s, HÀC(6)); 6.31
(m, HÀC(1')); 5.59 5.25 (m, HÀC(4'')); HÀC(3'), HÀC(3'')); 4.94 (s, HÀC(1'')); 4.27 4.22 (m, HaÀC(6''));
4.17 4.15 (m, HbÀC(6''), HaÀC(5')); 4.07 4.04 (m, HÀC(2''), HÀC(4')); 3.92 (m, HÀC(5'')); 3.74
(m, HbÀC(5')); 2.25 (m, 2 HÀC(2')); 2.12 1.93 (5s, 4 AcO, Me). 13C-NMR (CDCl3): 170.73 163.43 (4
CˆO), 150.32 (C(4)); 134.65 (C(6)); 111.61 (C(5)); 98.12 (C(1'')); 86.68 (C(1')); 84.7 (C(4')); 82.89 (C(3'));
74.27 (C(3'')); 69.44 (C(5'')); 67.78(C(5 ')); 65.20 (C(4'')); 62.08(C(6 '')); 61.17 (C(2'')); 37.08(C(2 ')); 20.89
‡
‡
(4 MeCO); 12.56 (Me). ESI-TOF-MS: 598.2132 ([M ‡ 1] ), 620.1924 ([M ‡ Na] ).
N4-Benzoyl-1-[2,3-di-O-benzoyl-5-O-(3,4,6-tri-O-acetyl-2-azido-2-deoxy-a-D-mannopyranosyl)-b-d-arabi-
nofuranosyl]cytosine (19): Yield 79.7%. White solid. [a]1D8 ˆ ‡18.5 (c ˆ 0.024, CHCl3). 1H-NMR (CDCl3): 8.75
(br., PhNH); 8.18 7.38 (m, 15 arom. H); 8.08 (d, J(5,6) ˆ 8.0, HÀC(6)); 7.84 (dd, HÀC(5)); 6.61 (d, J(1',2') ˆ
4.5, HÀC(1')); 6.05 (dd, J(2',3') ˆ 2.0, HÀC(2')); 5.59 (br., HÀC(3')); 5.35 (m, HÀC(4'), HÀC(3'')); 4.99
(d, J(1'',2'') ˆ 1.5, HÀC(1'')); 4.38( dd, J(3'',4'') ˆ 8.5, J(4'',5'') ˆ 5.0, HÀC(4'')); 4.28( m, HaÀC(6'')); 4.18
(m, HaÀC(5')); 4.12 (m, HbÀC(6''), HÀC(5''), HÀC(2'')); 4.03 (m, HbÀC(5')); 2.09, 2.07, 2.00 (3s, 3 AcO).
13C-NMR (CDCl3): 170.68 162.51 (6 CO); 145.18 (C(6)); 133.95 127.53 (12 C, Ph); 98.16 (C(1 '')); 97.27
(C(5)); 85.50 (C(1')); 81.18 (C(4')); 75.21 (C(3')); 70.88 (C(3'')); 68.94 (C(2'), C(5'')); 66.96 (C(5')); 65.64
‡
(C(4'')); 62.01 (C(6'')); 61.13 (C(2'')); 20.67 20.48(3 MeCO). ESI-TOF-MS: 869.0676 ([M ‡ 1] ), 891.0727
‡
([M ‡ Na] ).
5'-O-[3,6-Di-O-acetyl-2-azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl)-a-d-mannopyra-
nosyl]uridine 2',3'-Diacetate (24): Yield 46.7%. Yellow solid. [a]1D8 ˆ ‡27.2 (c ˆ 0.035, CHCl3). 1H-NMR
(CDCl3): 8.87 (s, NH); 7.44 (d, J(5,6) ˆ 6.0, HÀC(6)); 6.09 (d, J(1',2') ˆ 6.5, HÀC(1')); 5.97 (dd, HÀC(5)); 5.40
(dd, HÀC(3')); 5.38( dd, HÀC(4''')); 5.30 5.28( m, HÀC(3''), HÀC(2')); 5.17 (dd, HÀC(2''')); 4.99
(dd, HÀC(3''')); 4.85 (d, J(1'',2'') ˆ 2.0, HÀC(1'')); 4.59 (d, J(1''',2''') ˆ 8.50, HÀC(1''')); 4.46 (m, HaÀC(6'''));
4.25 4.16 (m, HÀC(4'), HbÀC(6'''), HaÀC(6'')); 4.11 4.07 (m, HÀC(2''), HbÀC(6'')); 4.02 4.01
(m, HÀC(4''), HÀC(5''')); 3.98 3.94 ( m, HÀC(5''), HaÀC(5')); 3.81 (m, HbÀC(5')); 2.26 1.97 (8s, 8AcO);
13C-NMR (CDCl3): 170.54 162.60 (8C ˆO); 150.41 (C(4)); 139.67 (C(6)); 103.98(C(5)); 101.15 (C(1 ''')); 98.19
(C(1'')); 87.10 (C(1')); 80.76 (C(4')); 73.75 (C(4')); 72.60 (C(2')); 71.03 (C(3''), C(3''')); 70.59 (C(5'')); 70.52

Helvetica Chimica Acta Vol. 86 (2003)
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(C(3')); 69.97 (C(5''')); 68.89 (C(2''')); 67.19 (C(5')); 66.82 (C(4''')); 62.03 (C(6''')); 61.12 (C(2'')); 60.92 (C(6''));
‡
20.81 20.25 (MeCO). ESI-TOF-MS: 930.0867 ([M ‡ 1] ).
N4-Acetyl-5'-O-[3,6-di-O-acetyl-2-azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl)-a-d-
mannopyranosyl]cytidine 2',3'-Diacetate (25): Yield 42.6%. Yellow solid. [a]1D8 ˆ ‡36.33 (c ˆ 0.033, CHCl3).
1H-NMR (CDCl3): 9.01 (br., AcNH); 7.87 (d, J(5,6) ˆ 7.5, HÀC(6)); 7.55 (dd, HÀC(5)); 6.16 (d, J(1',2') ˆ 4.0,
HÀC(1')); 5.40 5.30 (m, HÀC(3''), HÀC(4'''), HÀC(3'), HÀC(2')); 5.15 (dd, HÀC(2''')); 4.99
(dd, HÀC(3''')); 4.85 (d, J(1'',2'') ˆ 1.5, HÀC(1'')); 4.58( d, J(1''',2''') ˆ 8.0, HÀC(1''')); 4.42 (m, HaÀC(6'''));
4.31 (m, HÀC(4')); 4.19 4.14 (m, HaÀC(6''), HbÀC(6''')); 4.11 4.07 (m, HÀC(2''), HbÀC(6'')); 4.02 3.94
(m, HÀC(5''), HÀC(4''), HÀC(5'''), HaÀC(5')); 3.82 (m, HbÀC(5')); 2.15 1.97 (9s, 8AcO, AcNH). 13C-NMR
(CDCl3): 170.52 162.60 (9 CˆO); 144.42 (C(6)); 101.141 (C(1''')); 98.18 (C(1'')); 97.552 (C(1')); 88.94 (C(5));
80.71 (C(4')); 73.94 (C(4'')); 73.67 (C(2')); 71.04 (C(3'')); 70.89 (C(3''')); 70.59 (C(5'')); 70.26 (C(3')); 69.79
(C(5''')); 68.93 (C(2''')); 66.95 (C(5')); 66.81 (C(4''')); 62.06 (C(6''')); 61.09 (C(2'')); 60.96 (C(6'')); 24.97
‡
‡
(MeCONH); 20.82 20.34 (MeCO). ESI-TOF-MS: 971.0698([ M ‡ 1] ), 993.0771 ([M ‡ Na] ).
5'-O-[3,6-Di-O-acetyl-2-azido-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl)-a-d-mannopy-
ranosyl]thymidine 3'-Acetate (26): Yield 83%. White solid. [a]1D8 ˆ ‡26.4 (c ˆ 0.024, CHCl3). 1H-NMR (CDCl3):
8.59 (s, NH); 7.22 (s, HÀC(6)); 6.33 (m, HÀC(1')); 5.37 (m, HÀC(4''')); 5.31 (m, HÀC(3'')); 5.22
(m, HÀC(3')); 5.15 (dd, HÀC(2''')); 4.98( dd, HÀC(3''')); 4.88 (d, J(1''2'') ˆ 1.75, HÀC(1'')); 4.57
(d, J(1''',2''') ˆ 7.50, H ÀC(1''')); 4.45 (m, HaÀC(6''')); 4.21 3.88 (m, HÀC(4'), HbÀC(6'''), HaÀC(6''),
HÀC(2''), HbÀC(6''), HÀC(4''), HÀC(5'''), HÀC(5''), HaÀC(5')); 3.68( m, HbÀC(5')); 2.40 2.10
(m, HÀC(2')); 2.15 1.93 (8s, 7 AcO, Me). 13C-NMR (CDCl3): 170.49 163.22 (7 CˆO); 150.17 (C(4));
134.69 (C(6)); 111.62 (C(5)); 101.58(C(1 ''')); 97.97 (C(1'')); 84.57 (C(1')); 83.01 (C(4')); 74.34 (C(3')); 74.11
(C(4'')); 71.26 (C(3'')); 70.94 (C(3''')); 70.72 (C(5''')); 70.01 (C(5'')); 69.04 (C(2''')); 67.66 (C(5')); 66.73 (C(4'''));
61.93 (C(6''')); 61.17 (C(6''')); 61.02 (C(2'')); 36.96 (C(2')); 20.86 20.49 (7 MeCO); 12.49 (Me). ESI-TOF-MS:
‡
‡
886.1050 ([M ‡ 1] ), 908.0852 ([M ‡ Na] ).
N4-Benzoyl-1-[O-2,3,4,6-tetra-O-acetyl-b-d-galactopyranosyl-(1 ! 4)-O-3,6-di-O-acetyl-2-azido-2-deoxy-
a-d-mannopyranosyl-(1 ! 5)-2,3-di-O-benzoyl-b-d-arabinofuranosyl]cytosine (27). Yield 65.3%. Yellow solid.
[a]1D8 ˆ ‡8.35 (c ˆ 0.092, CHCl3). 1H-NMR (CDCl3): 8.84 (br. AcNH); 8.08 (d, J(5,6) ˆ 7.5, HÀC(6)); 7.85
(dd, HÀC(5)); 8.16 7.41 (15 arom. H); 6.60 (d, J(1',2') ˆ 3.50, HÀC(1')); 6.03 (dd, HÀC(2')); 5.60 (br.,
HÀC(3')); 5.42 5.38( m, HÀC(3''), HÀC(4''')); 5.14 (dd, J(2''',3''') ˆ 10.5, HÀC(2''')); 5.00 (dd, HÀC(3'''));
4.93 (d, J(1'',2'') ˆ 2.0, HÀC(1'')); 4.57 (d, J(1''',2''') ˆ 8.0, HÀC(1''')); 4.42 4.39 (m, HaÀC(6'''), HÀC(4'));
4.19 4.12 (m, HaÀC(6'')), HbÀC(6'''), HaÀC(5'), HbÀC(6'')); 4.02 3.94 (m, HÀC(5''), HÀC(4''), HÀC(5'''),
HÀC(2''), HbÀC(5')); 2.15 1.97 (6s, 6 AcO, AcN). 13C-NMR (CDCl3): 170.52 162.00 (9 CˆO); 133.85
127.55 (9 C, Ph); 129.93 (C(6)); 129.63 (C(5)); 101.46 (C(1''')); 97.96 (C(1'')); 85.55 (C(1')); 81.16 (C(4')); 76.44
(C(3')); 75.09 (C(2')); 74.47 (C(4'')); 70.90 (C(3''), C(3''')); 70.59 (C(5''')); 69.52 (C(5'')); 68.94 (C(2''')); 66.84
(C(5')); 66.74 (C(4''')); 62.06 (C(6''')); 60.97 (C(6'')); 60.88 (C(2'')); 20.82 20.34 (MeCO). ESI-TOF-MS:
‡
‡
1157.0925 ([M ‡ 1] ), 1179.0927 ([M ‡ Na] ).
5. General Procedure for Deprotection. At r.t., one of the compounds 16 19 or 24 27 (50 mg) was added to
10 ml of 25 30% MeNH2 in abs. EtOH. After stirring for 8h, the mixture was evaporated and the residue was
submitted to CC (silica gel, CH2Cl2/MeOH 10 :1 (removal of all impurities), then CH2Cl2/MeOH 1:1). The
eluted product was purified by reversed-phase CC (short C-18 column, H2O): 20 23 and 28 31, resp.
5'-O-(2-Azido-2-deoxy-a-d-mannopyranosyl)uridine (20): Yield 83.3%. White solid. [a]1D8 ˆ ‡66.0 (c ˆ
0.015, H2O). 1H-NMR (D2O): 7.76 (d, J(5,6) ˆ 8.13, HÀC(6)); 5.80 (d, HÀC(5)); 5.77 (d, J(1',2') ˆ 3.8,
HÀC(1')); 4.96 (d, J(1'',2'') ˆ 1.41, HÀC(1'')); 4.22 4.19 (m, HÀC(2'), HÀC(3'), HÀC(4')); 3.96
(dd, HÀC(2'')); 3.90 3.86 (m, HaÀC(5'), HÀC(3'')); 3.79 3.74 (m, HaÀC(6''), HbÀC(5')); 3.65
(m, HbÀC(6'')); 3.54 (t, HÀC(4'')); 3.42 (m, HÀC(5'')). 13C-NMR (D2O): 166.62, 151.84 (C(4), C(2)); 141.86
(C(6)); 102.22 (C(5)); 98.17 (C(1'')); 90.36 (C(1')); 82.50 (C(3')); 74.29 (C(2')); 73.44 (C(5'')); 70.86 (C(3''));
69.46 (C(4')); 66.95 (C(4'')); 66.13 (C(5')); 64.03 (C(2'')); 60.98(C(6 '')). HR-FAB-MS (pos. mode): 432.1306
‡
([C15H22O10N5 ‡ H] ; calc. 432.1366).
5'-O-(2-Azido-2-deoxy-a-d-mannopyranosyl)cytidine (21): Yield 62.1%. White solid. [a]1D8 ˆ ‡26.8( c ˆ
0.020, H2O). 1H-NMR (D2O): 7.63 (d, J(5,6) ˆ 7.50, HÀC(6)); 5.81 (d, HÀC(5)); 5.63 (s, HÀC(1')); 4.83
(s, HÀC(1'')); 4.05 (br., HÀC(2'), HÀC(3'), HÀC(4')); 3.82 3.63 (m, HÀC(2''), HaÀC(5'), HÀC(3'')); 3.69
3.63 (m, HaÀC(6''), HbÀC(5')); 3.55 3.49 (m, HbÀC(6'')); 3.47 3.40 (m, HÀC(4')); 3.35 (m, HÀC(5'')).
13C-NMR (D2O): 168.77, 160.04 (C(4), C(2)); 143.76 (C(6)); 100.43 (C(5)); 98.30 (C(1'')); 93.31 (C(1')); 84.35
(C(3')); 77.02 (C(2')); 75.71 (C(5'')); 73.16 (C(3')); 71.52 (C(4')); 69.25 (C(4'')); 68.23 (C(5')); 66.34 (C(2''));
‡
63.24 (C(6'')). HR-FAB-MS (pos. mode): 431.1523 ([C15H23O9N6 ‡ H] ; calc. 431.1526).

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Helvetica Chimica Acta Vol. 86 (2003)
5'-O-(2-Azido-2-deoxy-a-d-mannopyranosyl)thymidine (22): Yield 72.7%. White solid. [a]1D8 ˆ ‡61.4 (c ˆ
0.016, H2O). 1H-NMR (D2O): 7.46 (s, HÀC(6)); 6.15 (t, J(1',2') ˆ 6.54, HÀC(1')); 4.90 (s, HÀC(1'')); 4.43
(dd, HÀC(3')); 4.07 (dd, HÀC(4')); 3.88 3.84 (m, HÀC(2''), HaÀC(5'), HÀC(3'')); 3.76 3.67 (m, HaÀC(6''),
HbÀC(5')); 3.64 (m, HbÀC(6'')); 3.55 (dd, HÀC(4'')); 3.42 (m, HÀC(5'')); 2.33 2.23 (m, 2 HÀC(2')); 1.82
(s, Me). 13C-NMR (D2O): 166.76, 151.92 (C(4), C(2)); 137.49 (C(6)); 111.54 (C(5)); 98.33 (C(1'')); 85.91 (C(1'));
85.27 (C(4')); 73.41 (C(5'')); 71.11 (C(3')); 70.82 (C(3'')); 67.97 (C(5'')); 67.17 (C(5')); 66.99 (C(4'')); 64.14
‡
(C(2'')); 60.96 (C(6'')); 39.28(C(2 ')); 12.11 (Me). HR-FAB-MS (pos. mode): 430.1597 ([C16H24O9N5 ‡ H] ;
calc. 430.1574).
1-[5-O-(2-Azido-2-deoxy-a-d-Mannopyranosyl)-b-d-arabinofuranosyl]cytosine (23): Yield 87.0%. White
solid. [a]1D8 ˆ ‡16.61 (c ˆ 0.028, MeOH). 1H-NMR (D2O): 7.59 (d, J(5,6) ˆ 7.50, HÀC(6)); 6.04 (d, J(1',2') ˆ 5.4,
HÀC(1')); 5.85 (d, HÀC(5)); 4.87 (s, HÀC(1'')); 4.26 (t, HÀC(3')); 3.99 3.79 (m, HÀC(2'), HÀC(4'),
HÀC(2''), HaÀC(5'), HÀC(3'')); 3.71 3.47 (m, HaÀC(6''), HbÀC(5'), HÀC(5''), HbÀC(6''), HÀC(4'')).
13C-NMR (D2O): 166.80, 158.13 (C(4), C(2)); 142.99 (C(6)); 98.64 (C(1'')); 96.06 (C(1')); 86.14 (C(5)); 81.25
(C(3')); 76.26 (C(2')); 75.46 (C(5'')); 73.77 (C(3'')); 71.18(C(4 ')); 67.35 (C(4'')); 66.74 (C(5')); 64.41 (C(2''));
‡
61.32 (C(6''); HR-FAB-MS (pos. mode): 431.1505 ([C15H23O9N6 ‡ H] ; calc. 431.1526).
5'-O-[2-Azido-2-deoxy-4-O-(b-d-galactopyranosyl)-a-d-mannopyranosyl]uridine (28): Yield 67.2%. White
solid. [a]1D8 ˆ ‡66.0 (c ˆ 0.015, MeOH). 1H-NMR (D2O): 7.73 (d, J(5,6) ˆ 8.0, HÀC(6)); 5.77 (d, HÀC(5)); 5.73
(d, J(1',2') ˆ 3.0, HÀC(1')); 4.93 (s, HÀC(1'')); 4.31 (d, J(1''',2''') ˆ 7.50, HÀC(1''')); 4.20 4.14 (m, HÀC(3''),
HÀC(4'''), HÀC(3')); 3.99 3.96 (m, HÀC(2'), HÀC(2''')); 3.84 (dd, HÀC(3''')); 3.79 3.52 (m, HaÀC(6'''),
HÀC(4'), HbÀC(6'''), HaÀC(6''), HÀC(2''), HbÀC(6''), HÀC(4''), HÀC(5'''), HÀC(5''), HaÀC(5')); 3.38
(m, HbÀC(5')). 13C-NMR (D2O): 169.08, 154.25 (C(2), C(4)); 144.26 (C(6)); 105.82 (C(5)); 104.53 (C(1'''));
100.31 (C(1'')); 92.79 (C(1')); 84.78 (C(3')); 78.94 (C(5''')); 78.11 (C(2')); 76.63 (C(5'')); 75.21 (C(3'')); 74.41
(C(3''')); 73.68(C(4 ')); 72.03 (C(2''')); 71.74 (C(4'')); 71.28(C(5 ')); 68.48 (C(4''')); 65.78(C(2 '')); 63.82 (C(6''));
‡
62.68(C(6 ''')). HR-FAB-MS (pos. mode): 594.1852 ([C21H32O15N5 ‡ H] ; calc. 594.1895).
5'-O-[2-Azido-2-deoxy-4-O-(b-d-galactopyranosyl)-a-d-mannopyranosyl]cytidine (29): Yield 62.1%.
White solid. [a]1D8 ˆ ‡76.1 (c ˆ 0.011, MeOH). 1H-NMR (D2O): 7.73 (d, J(5,6) ˆ 7.5, H ÀC(6)); 5.91
(d, HÀC(5)); 5.72 (d, J(1',2') ˆ 2.0, HÀC(1')); 4.93 (s, HÀC(1'')); 4.31 (d, J(1''',2''') ˆ 8.00, HÀC(1''')); 4.17
4.11 (m, HÀC(3''), HÀC(4'''), HÀC(3')); 3.99 3.95 (m, HÀC(2'), HÀC(2''')); 3.86 3.51 (m, HÀC(3'''),
HaÀC(6'''), HÀC(4'), HbÀC(6'''), HaÀC(6''), HÀC(2''), HbÀC(6''), HÀC(4''), HÀC(5'''), HÀC(5''),
HaÀC(5')); 3.38( m, HbÀC(5)). 13C-NMR (D2O): 166.94, 158.19 (C(2), C(4)); 141.91 (C(6)); 103.80 (C(5));
98.30 (C(1''')); 96.42 (C(1'')); 91.42 (C(1')); 82.39 (C(3')); 76.95 (C(5''')); 76.10 (C(2')); 75.02 (C(5'')); 73.21
(C(3'')); 72.38(C(3 ''')); 71.67 (C(4')); 70.05 (C(2''')); 69.54 (C(4')); 69.27 (C(5')); 66.36 (C(4''')); 63.79 (C(2''));
‡
61.82 (C(6'')); 60.67 (C(6''')). HR-FAB-MS (pos. mode): 593.2054 ([C21H33O14N6 ‡ H] ; calc. 593.2055).
5'-O-[2-Azido-2-deoxy-4-O-(b-d-galactopyranosyl)-a-d-mannopyranosyl]thymidine (30): Yield 88.0%.
White solid. [a]D18 ˆ ‡14.04 (c ˆ 0.027, MeOH). 1H-NMR (D2O): 7.40 (s, HÀC(6)); 6.11 (t, J(1',2') ˆ 6.6,
HÀC(1')); 4.87 (d, J(1'',2'') ˆ 1.5, HÀC(1'')); 4.63 (dd, HÀC(4''')); 4.28( d, J(1''',2''') ˆ 7.50, HÀC(1''')); 4.01
(dd, HÀC(3')); 3.94 (dd, HÀC(3'')); 3.86 (dd, HÀC(2''')); 3.81 3.73 (m, HÀC(3'''), HaÀC(6''')); 3.70 3.48
(m, HÀC(4'), HbÀC(6'''), HaÀC(6''), HÀC(2''), HbÀC(6''), HÀC(4''), HÀC(5'''), HÀC(5''), HaÀC(5')); 3.36
(m, HbÀC(5')); 2.25 (m, 2 HÀC(2')); 1.77 (s, Me). 13C-NMR (D2O): 169.20, 154.36 (C(4), C(2)); 139.94 (C(6));
114.01 (C(5)); 105.83 (C(1''')); 100.57 (C(1'')); 88.24 (C(1')); 87.59 (C(3')); 79.01 (C(5''')); 78.12 (C(5'')); 75.29
(C(3'')); 74.42 (C(3''')); 73.73 (C(4')); 73.36 (C(4'')); 72.05 (C(2''')); 71.33 (C(5')); 69.61 (C(4''')); 65.91 (C(2''));
63.84 (C(6'')); 62.73 (C(6''')); 41.58(C(2 ')); 14.51 (Me). HR-FAB-MS (pos. mode): 592.2108([C 22H34O14N5 ‡
‡
H] ; calc. 592.2102).
1-[O-b-d-Galactopyranosyl-(1 ! 4)-O-2-azido-2-deoxy-a-d-mannopyranosyl-(1 ! 5)-b-d-arabinofurano-
syl]cytosine (31): Yield 91.0%. White solid. [a]1D8 ˆ ‡37.48( c ˆ 0.021, MeOH). 1H-NMR (D2O): 7.63
(d, J(5,6) ˆ 8.0, HÀC(6)); 6.09 (d, J(1',2') ˆ 5.50, HÀC(1')); 5.90 (dd, HÀC(5)); 4.93 (d, J(1'',2'') ˆ 1.0,
HÀC(1'')); 4.32 4.29 (m, HÀC(1'''), HÀC(2')); 4.05 3.96 (m, HÀC(3'), HÀC(3''), HÀC(4'''), HÀC(2'''));
3.86 (dd, HÀC(3''')); 3.81 3.59 (m, HaÀC(6'''), HÀC(4'), HaÀC(6''), HbÀC(6'''), HÀC(2''), HbÀC(6''),
HÀC(5''), HÀC(4''), HÀC(5''')); 3.53 (HaÀC(5')); 3.39 (HbÀC(5')). 13C-NMR (D2O): 168.79, 160.14 (C(2),
C(4)); 145.04 (C(6)); 105.81 (C(5)); 100.52 (C(1''')); 98.07 (C(1'')); 88.10 (C(1')); 83.22 (C(3')); 78.96 (C(5'''));
78.29 (C(2')); 78.11 (C(5'')); 77.41 (C(3'')); 75.24 (C(3''')); 74.41 (C(4')); 73.69 (C(2''')); 72.02 (C(4'')); 71.31
(C(5')); 68.86 (C(4''')); 65.82 (C(2'')); 63.83 (C(6'')); 62.70 (C(6''')). HR-FAB-MS (pos. mode): 593.2053
‡
([C21H33O14N6 ‡ H] ; calc. 593.2054).
5. Antitumor Activity. Inhibition of tumor-cell growth by the designed compounds was carried out according
to published methods [20][21].

Helvetica Chimica Acta Vol. 86 (2003)
2081
This work was supported by the National Natural Science Foundation of China.
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Received October 20, 2002