An efficient synthesis of 3′-amino-3′-deoxythymidine derivatives

Article abstract of DOI:10.1007/s11171-005-0018-4

An efficient method of reduction of 3′-azido-3′-deoxythymidine and its 5′-protected derivatives to 3′-aminothymidine derivatives on a palladium catalyst using ammonium formate as a source of hydrogen was suggested.

Full text of DOI:10.1007/s11171-005-0018-4

Russian Journal of Bioorganic Chemistry, Vol. 31, No. 2, 2005, pp. 135–138. Translated from Bioorganicheskaya Khimiya, Vol. 31, No. 2, 2005, pp. 147–150.
Original Russian Text Copyright © 2005 by Seregin, Chudinov, Yurkevich, Shvets.
An Efficient Synthesis of 3'-Amino-3'-Deoxythymidine
Derivatives
1
K. V. Seregin, M. V. Chudinov, A. M. Yurkevich, and V. I. Shvets
Lomonosov State Academy of Fine Chemical Technology, pr. Vernadskogo 86, Moscow, 117571 Russia
Received April 20, 2004; in final form, June 10, 2004
Abstract—An efficient method of reduction of 3'-azido-3'-deoxythymidine and its 5'-protected derivatives to
3'-aminothymidine derivatives on a palladium catalyst using ammonium formate as a source of hydrogen was
suggested.
Key words: 3'-azido-3'-deoxythymidine, modified nucleosides
1
INTRODUCTION
The reduction of azide group is more effective and is
used much more often.
3'-Amino-3'-deoxythymidine stands out from other
synthetic analogues of natural nucleosides owing to its
wide spectrum of biological activities. Its antineoplas-
tic properties are well-known; they have been demon-
strated on, e.g., L1210 murine leukemia [1] and CCRF-
HSB-2 and KB lymphoblastoid leukemia [2] cell lines.
3'-Amino-3'-deoxythymidine has been found to be one
of the main metabolites of 3'-azido-3'-deoxythymidine,
which is probably generated in liver cells upon the
reduction with cytochrome P-450 [3–5]. 3'-Amino-3'-
deoxythymidine possesses high cytotoxic effect [3–5]
and inhibits thymidine transport into erythrocytes [6].
This compound itself is not used in medical practice
because of its high toxicity; however, the search for
potential pharmacophores among its derivatives has
extensively been performed [7–9].
Catalytic hydrogenation over platinum or palladium
catalysts is the most abundant method for the azide con-
version into amine. The reaction was carried out in var-
ious solvents and at various values of hydrogen pres-
sure in 51–90% yield [16–20]. Among other proce-
dures, the reduction with SnCl2 in methanol resulting in
85% yield of the product, the reduction using SnCl2–
PhSH–Et3N system in acetonitrile (85%) [21], the radi-
cal reduction with Bun3 SnH [22], and the reductions
with NaBH4 (93%) [7] and thiols, β-mercaptoethanol
(46%) [8] or dithiothreitol (96%) [9] should be men-
tioned. The reduction with triphenylphosphine attached
to a polymer matrix (70%) has also been described
[23]. A more detailed information on the methods of
reduction of azidonucleosides to aminonucleosides can
be found in review [24].
3'-Amino-3'-deoxythymidine can be used as a build-
ing block in the synthesis of oligonucleotides with
nitrogen-containing internucleotide linkers [10–12],
for the introduction of label upon DNA sequencing
[13], and for the attachment of oligonucleotides with
the 3'-terminal amino group to the carrier in the solid-
phase synthesis [14, 15].
The method of 3'-amino-3'-deoxythymidine synthe-
sis by the catalytic reduction of 3'-azido-3'-deoxythy-
midine over the palladium catalyst using ammonium
formate as a source of hydrogen, we herein suggest,
results in almost quantitative (97–98%) yield, and the
reaction time is significantly less than in the case of
reduction with dithiothreitol, which is the most effec-
tive method among those aforementioned. Note that the
isolation procedure of the reaction product is also sim-
plified.
RESULTS AND DISCUSSION
Currently, two types of approaches to the obtaining
of 3'-amino-3'-deoxythymidine have been reported.
These are the syntheses with the reduction of 3'-azido-
3'-deoxythymidine [7–9, 16–24] and syntheses through
the alkylation of various nitrogen-containing com-
pounds by 3'-sulfonate derivatives of thymidine [25].
We reduced the 5'-protected azidothymidine deriva-
tives containing acyl protective groups (Ib)–(Id) in
almost quantitative yields. In the case of the derivatives
with trityl protective groups (Ie)–(If), the yields were
somewhat lower (70–80%), probably because of a par-
tial cleavage of these protective groups under the reac-
tion conditions.
1
Corresponding author; phone: +7 (095) 434-7146; e-mail:
kirill_7743@comail.ru
1068-1620/05/3102-0135 © 2005 Pleiades Publishing, Inc.

136
SEREGIN et al.
EXPERIMENTAL
room temperature. The reaction was monitored by TLC
(system A). Two hours after the gas liberation ceased,
the catalyst was removed by filtration through the
Celite layer, the residue was repeatedly washed with
methanol, and the solvent was removed in a vacuum.
The residue was dried in a vacuum (0.1 mmHg, 2 h);
3'-Azido-3'-deoxythymidine (Ia) was from Assotsi-
atsiya AZT (Russia); ammonium formate, from Riedel
de Haen (Germany); 10% palladium on carbon, from
Lancaster (UK); pivaloyl chloride, from Merck (Ger-
many); and trityl chloride and 4,4'-dimethoxytrityl
chloride, fromAcros (Belgium).Other reagents and sol-
vents of reagent or analytical grade were of domestic
1
yield 4.42 g (98%); H NMR (4 : 1 CDCl3–CD3OD):
7.58 (1 H, d, J 1.2, H6), 5.98 (1 H, dd, J1 5.2, J2 6.8,
H1'), 3.57–3.73 (3 H, m, H4' and H5'), 3.54 (1 H, m,
H3'), 1.96–2.20 (2 H, m, H2'), and 1.79 (3 H, d, J 1.2,
5-ëç3). Found, %: C 49.64, N 17.11, H 6.53.
C10H15N3O4. Calc., %: C 49.79, N 17.42, H 6.27.
manufacture.
5'-Acetyl-3'-azido-3'-deoxythymidine
(Ia) was synthesized by the procedure [26]; 3'-azido-5'-
benzoyl-3'-deoxythymidine (Id) was obtained as
described in [27]. TLC was carried out on Kieselgel 60
F254 (Merck, Germany) precoated plates in (A) 9 : 1
chloroform–methanol, (B) 19 : 1 chloroform–metha-
nol, (C) 3 : 1 chloroform–ethyl acetate, and (D) chloro-
form. Substance spots were visualized by spraying with
ninhydrin and UV irradiation (254 nm). Column chro-
matography was carried out on Kieselgel 60 (Merck,
Germany).
3'-Azido-3'-deoxy-5'-O-pivaloylthymidine (Ib).
Pivaloyl chloride (1.56 ml, 12.59 mmol) was added to
a solution of 3'-azido-3'-deoxythymidine (Ia) (2 g,
7.41 mmol) in anhydrous pyridine (10 ml) cooled in a
water bath; the solution was stirred for 12 h at room
temperature until the reaction completed (TLC moni-
toring, system B), diluted with water (50 ml), and
extracted with ethyl acetate (40 ml). The organic layer
was washed with 4% hydrochloric acid, dried with
Na2SO4, and evaporated in a vacuum. The solid white
residue of (Ib) was dried in a vacuum (1 mmHg, 1.5 h);
yield 1.99 g (76.51%); 1H NMR (CDCl3): 8.49 (1 H, br.
s, Nç), 7.21 (1 H, br. s, H6), 6.12 (1 H, t, J 6.5, H1'),
4.52 (2 H, m, H5'), 4.31 (1 H, m H4'), 4.19 (1 H, m H3'),
2.44 (2 H, m, H2'), 1.69 (3 H, s, 5-ëç3), and 1.21 (9 H,
s, pivaloyl). Found, %: C 50.95, N 19.82, H 6.05.
C15H21N5O5. Calc., %: C 51.28, N 19.93, H 6.02.
1H NMR spectra were registered on a Bruker MSL-
200 spectrometer at
a working frequency of
200.13 MHz using tetramethylsilane as an internal
standard. Chemical shifts are given in ppm (δ scale) and
spin coupling constants, J, in Hz.
3'-Amino-3'-deoxythymidine (IIa). 3'-Azido-3'-
deoxythymidine (Ia) (5 g, 19 mmol) was dissolved in
absolute methanol (120 ml), 10% palladium on carbon
(0.5 g) and ammonium formate (7.18 g, 114 mmol)
were added, and the reaction mixture was stirred at
3'-Azido-3'-deoxy-5'-O-tritylthymidine (Ie). Tri-
tyl chloride (0.18 g, 0.63 mmol) was added to a solution
of 3'-azido-3'-deoxythymidine (Ia) (0.1 g, 0.37 mmol)
in anhydrous pyridine (10 ml). The solution was stirred
overnight at 45°C until the reaction completed (TLC
monitoring, system C) and then poured into ice-cold
water (100 ml). The precipitated solid was filtered,
dried in a vacuum (10 mmHg, 12 h), and crystallized
from toluene. The light yellow fine crystalline precipi-
tate of (Ie) was dried in a vacuum (10 mmHg, 12 h);
yield 0.18 g (95%); mp 103–104°C; 1H NMR (CDCl3):
8.62 (1 H, br. s, Nç), 7.41–7.18 (15 H, m, trityl), 7.17
(1 H, br. s, H6), 6.16 (1 H, t, J 6.5, H1'), 4.60 (2 H, m,
H5'), 4.33 (1 H, m, H4'), 4.19 (1 H, m, H3'), 2.44 (2 H,
m, H2'), and 1.67 (3 H, s, 5-ëç3). Found, %: C 68.01,
N 13.50, H 5.31. C29H27N5O4. Calc., %: C 68.36, N
13.74, H 5.34.
O
N
O
N
HN
HN
RO
RO
O
O
O
O
Pd/C, HCOONH4
MeOH
N3
(Ia–If)
NH2
(IIa–IIf)
R
‡
b
c
d
e
f
H
3'-Azido-3'-deoxy-5'-O-(4,4'-dimethoxytrityl)thy-
midine (If). 4,4'-Dimethoxytrityl chloride (4.16 g,
12.27 mmol) was added to a solution of 3'-azido-3'-
deoxythymidine (I‡) (2.76 g, 10.22 mmol) in anhy-
drous pyridine (70 ml). The reaction mixture was
stirred for 18 h at 45°C until the reaction completed
(TLC monitoring, system A), methanol (50 ml) was
added, and the solvents were evaporated in a vacuum.
The residue was distributed between ëç2Cl2 (100 ml)
and water (100 ml). The organic layer was washed with
Piv
Ac
Bz
Tr
(MeO)2Tr
Scheme.
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 31 No. 2 2005

AN EFFICIENT SYNTHESIS OF 3'-AMINO-3'-DEOXYTHYMIDINE DERIVATIVES
137
5% Na2CO3 (70 ml), dried with Na2SO4, and evapo- 8.38, H 6.23. C29H29N3O4. Calc., %: C 72.03, N 8.69, H
rated in a vacuum. The residue was crystallized from 6.04.
toluene, and the white-pink amorphous precipitate of
3'-Amino-3'-deoxy-5'-O-(4,4'-dimethoxytrityl)thy-
(If) was dried in a vacuum (10 mmHg, 12 h); yield
midine (IIf) was obtained as described for (IIb) from
4.95 g (84.97%); 1H NMR (CDCl3): 11.1 (1 H, s, NH),
3'-azido-3'-deoxy-5'-O-(4,4'-dimethoxytrityl)thymidine
6.95–7.70 (14 H, m, trityl and H6), 6.12 (1 H, t, J 6.2,
H1'), 4.56 (2 H, m, H5'), 4.28 (1 H, m, H4'), 4.15 (1 H,
m, H3'), 3.60 (6 H, s, ëç3é), 2.22 (2 H, m, H2'), and
1.72 (3 H, s, 5-ëç3). Found, %: C 65.01, N 12.27, H
5.51. C31H31N5O6. Calc., %: C 65.37, N 12.30, H 5.49.
(If) (0.28 g). The reaction product was additionally
purified by column chromatography on silica gel using
ethyl acetate as an eluent; yield 0.19 g (71%); 1H NMR
(CDCl3): 11.1 (1 H, s, NH), 6.95–7.70 (14 H, m, trityl
and H6), 6.11 (1 H, t, J 6.2, H1'), 4.56 (2 H, m, H5'),
4.08 (1 H, m, H4'), 3.59 (6 H, s, NH3I), 3.47 (1 H, m,
H3'), 2.24 (2 H, m, H2'), 1.67 (3 H, d, J 1.2, 5-NH3).
3'-Amino-3'-deoxy-5'-O-pivaloylthymidine (IIb).
3'-Azido-3'-deoxy-5'-O-pivaloylthymidine (Ib) (1 g,
2.84 mmol) was dissolved in absolute methanol Found, %: C 67.75, N 6.89, H 6.44. C31H33N3O6. Cal-
(12 ml), and 10% palladium on carbon (100 mg) was
added. Ammonium formate (1.07 g, 17.04 mmol) was
added, and the reaction mixture was refluxed (TLC
monitoring, system D). After 2.5 h, the catalyst was
removed by filtration through a silica gel layer (0.5 cm),
the residue was repeatedly washed with methanol, and
the solvent was evaporated in a vacuum; yield of (IIb)
0.9 g (97%); 1H NMR (CDCl3): 11.3 (1 H, br. s, NH),
7.43 (1 H, br. s, H6), 6.11 (1 H, t, J 6.2, H1'), 4.36–4.65
(2 H, m, H5'), 4.14 (1 H, m, H4'), 3.75 (1 H, m, H3'),
2.12–2.38 (2 H, m, H2'), 1.70 (3 H, d, J 1.2, 5-ëç3),
and 1.18 (9 H, s, pivaloyl). Found, %: C 54.69, N 16.11,
H 7.29. C15H23N3O5. Calc., %: C 55.37, N 12.91, H
7.13.
5'-O-Acetyl-3'-amino-3'-deoxythymidine (IIc)
was obtained as described for (IIb) from 5'-O-acetyl-3'-
azido-3'-deoxythymidine (Ic) (0.3 g); yield 0.26 g
(95%); 1H NMR (4 : 1 CDCl3–CD3OD): 7.56 (1 H, br.
s, H6), 6.18 (1 H, t, J 6.2, H1'), 4.17–4.29 (2 H, m, H5'),
3.91 (1 H, m, H4'), 3.78 (1 H, m, H3'), 2.65 (3 H, s, Ac),
2.11–2.28 (2 H, m, H2'), and 1.76 (3 H, d, J 1.2, 5-CI3).
Found, %: C 49.95, N 14.50, H 6.51. C12H17N3O5. Cal-
culated, %: C 50.88, N 14.83, H 6.05.
culated, %: C 68.49, N 7.73, H 6.12.
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1
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