Structure-activity relationships of 2′-modified-4′- selenoarabinofuranosyl-pyrimidines as anticancer agents

Article abstract of DOI:10.1016/j.ejmech.2014.06.031

Based on the potent anticancer activity of the D-arabino-configured cytosine nucleoside ara-C, novel 2′-substituted-4′- selenoarabinofuranosyl pyrimidines 3a-3u, comprising azido, fluoro, and hydroxyl substituents at C-2′ were designed, synthesized, and evaluated for anticancer activity. The 2′-azido group was stereoselectively introduced by the Mitsunobu reaction using diphenylphosphoryl azide (DPPA), and the 2′-fluoro group was stereoselectively introduced through the double inversions of stereochemistry via the episelenium intermediate, which was formed by the participation of the selenium atom. Among the compounds tested, the 2′-fluoro derivative 3t (X = NH₂, Y = H, R = F) was found to be the most potent anticancer agent and showed more potent anticancer activity than the control, ara-C in all tested human cancer cell lines (HCT116, A549, SNU638, T47D, and PC-3) except the leukemia cell lines (K562). The anticancer activity of the 2′-substituted-4′-selenonucleosides is in the following order: 2′-F > 2′-OH > 2′-N₃.

Full text of DOI:10.1016/j.ejmech.2014.06.031

Accepted Manuscript
Structure-Activity Relationships of 2'-Modified-4'-selenoarabinofuranosyl-pyrimidines
as Anticancer Agents
Jin-Hee Kim, Jinha Yu, Varughese Alexander, Jung Hee Choi, Jayoung Song, Hyuk
Woo Lee, Hea Ok Kim, Jungwon Choi, Sang Kook Lee, Lak Shin Jeong, Ph.D
PII:
S0223-5234(14)00550-9
10.1016/j.ejmech.2014.06.031
EJMECH 7074
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 30 April 2014
Revised Date: 11 June 2014
Accepted Date: 13 June 2014
Please cite this article as: J.-H. Kim, J. Yu, V. Alexander, J.H. Choi, J. Song, H.W. Lee, H.O. Kim, J.
Choi, S.K. Lee, L.S. Jeong, Structure-Activity Relationships of 2'-Modified-4'-selenoarabinofuranosyl-
pyrimidines as Anticancer Agents, European Journal of Medicinal Chemistry (2014), doi: 10.1016/
j.ejmech.2014.06.031.
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ACCEPTED MANUSCRIPT
Graphical Abstract
Structure-Activity Relationships of 2'-Modified-4'-selenoarabinofuranosyl-pyrimidines
as Anticancer Agents
Jin-Hee Kim, Jinha Yu, Varughese Alexander, Jung Hee Choi, Jayoung Song, Hyuk
*
Woo Lee, Hea Ok Kim, Jungwon Choi, Sang Kook Lee, and Lak Shin Jeong

ACCEPTED MANUSCRIPT
Eur. J. Med. Chem.-revised
Structure-Activity Relationships of 2'-Modified-4'-selenoarabinofuranosyl-
ǂ
pyrimidines as Anticancer Agents
1
1
1
1
2
Jin-Hee Kim, Jinha Yu, Varughese Alexander, Jung Hee Choi, Jayoung Song, Hyuk
1
1
3
2
1,2,*
Woo Lee, Hea Ok Kim, Jungwon Choi, Sang Kook Lee, and Lak Shin Jeong
1
2
College of Pharmacy, Ewha Womans University, Seoul 120-750, Korea, College of
3
Pharmacy, Seoul National University, Seoul 151-742, Korea, and Department of
Chemistry, The University of Suwon, Kyunggi-do 445-743, Korea
*
To whom correspondence should be addressed:
Lak Shin Jeong, Ph.D.
College of Pharmacy
Seoul National University
Seoul 151-742, Korea
Tel) 82-2-880-7850
Fax) 82-2-884-8334
E-mail) lakjeong@snu.ac.kr
Key Words: 4'-selenonucleosides; antimetabolite; Mitsunobu reaction; structure-
activity relationship; stereoselective fluorination
ǂ
J.-H. Kim and J. Yu equally contributed to this work

ACCEPTED MANUSCRIPT
Abstract
Based on the potent anticancer activity of the
D
-arabino-configured cytosine nucleoside
ara-C, novel 2'-substituted-4'-selenoarabinofuranosyl pyrimidines 3a-3u, comprising
azido, fluoro, and hydroxyl substituents at C-2' were designed, synthesized, and
evaluated for anticancer activity. The 2'-azido group was stereoselectively introduced by
the Mitsunobu reaction using diphenylphosphoryl azide (DPPA), and the 2'-fluoro
group was stereoselectively introduced through the double inversions of stereochemistry
via the episelenium intermediate, which was formed by the participation of the selenium
atom. Among the compounds tested, the 2'-fluoro derivative 3t (X = NH , Y = H, R =
2
F) was found to be the most potent anticancer agent and showed more potent anticancer
activity than the control, ara-C in all tested human cancer cell lines (HCT116, A549,
SNU638, T47D, and PC-3) except the leukemia cell lines (K562). The anticancer
activity of the 2'-substituted-4'-selenonucleosides is in the following order: 2'-F > 2'-OH
>
2'-N3.

ACCEPTED MANUSCRIPT
Introduction
DNA and RNA building blocks have long been regarded as valuable resources
towards the development of therapeutically useful modified nucleosides. The major
mechanism of action of these modified nucleosides for a variety of biological activities
is to act as antimetabolites interfering with cellular or viral metabolism. Based on this
mechanism, many anticancer or antiviral nucleosides have clinically been developed as
1
antimetabolites.
Uridine (1a, X = OH) and cytidine (1b, X = NH ) are essential RNA
2
pyrimidine building blocks and have also been served as important templates for the
development of new antiviral and anticancer agents. On the basis of the structure of
these templates, modifications have largely been done on the 2' or 3' position and 2'-
2
modified nucleosides generally showed better biological activities. For example, 1-ꢀ-
D
-arabinofuranosyl cytosine (ara-C, 2, X = NH , Y = O, R = OH) with an arabino
2
configuration is one of the representative nucleosides and is being used clinically as an
3
4
anticancer agent. Its bioisosteric 2'-fluoro (Y = O, R = F) and 2'-azido (Y = O, R =
5
N ) analogues 2 also showed good antiviral or anticancer activities (Figure 1).
3

ACCEPTED MANUSCRIPT
Figure 1. The rationale for the design of the target nucleoside 3
On the basis of bioisosteric rationale, the corresponding 4'-methylene
6
7
(
carbocyclic) and 4'-thionucleosides 2 were also synthesized and reported to show
various biological activities. Although they possessed better enzymatic and chemical
stability than the corresponding 4'-oxonucleosides, only a few nucleosides showed
attractive biological activity. Thus, the discovery of a new template to replace known
modified nucleosides has highly been desirable.

ACCEPTED MANUSCRIPT
Recently, the 4'-selenonucleoside has been reported as the next generation
nucleoside for the development of new therapeutic agents as well as new biolo
8
gical tools. Among these, the 4'-seleno analogue 3 (X = NH , Y = H, R = F)
2
of 2'-fluoro-ara C showed potent anticancer activity in a panel of human tumor
8
c
cell lines. Thus, based on these findings, it was of great interest to study the s
tructure-activity relationship of the 4'-selenoarabinofuranosyl-pyrimidines 3 substitu
ted with bioisosteric azido, fluoro, or hydroxyl group at the 2' position as antica
ncer agents. The 2'-azido group was stereoselectively introduced by the Mitsunob
u reaction using diphenylphosphoryl azide (DPPA) with inversion of configuration
and the 2'-fluoro group was stereoselectively introduced by N,N-diethylaminosufu
r trifluoride (DAST) reaction via an episelenium ion intermediate through double
inversions of configuration. In this article, we report the full accounts of the str
ucture-activity relationships of 2'-modified-4'-selenoarabibofuranosyl pyrimidines 3
as anticancer agents.
Results and discussion
Chemistry

ACCEPTED MANUSCRIPT
For the synthesis of the target nucleosides, the key intermediate 4-selenosugar 9 was
first synthesized from D-ribose according to our previously reported procedure (Scheme
8
e
1
). Commercially available 2,3-O-isopropylidene-
D
-ribonolactone (4) was converted
9
to known 2,3-O-isopropylidene-
L
-lyxonolactone (5) in two steps. Protection of the
hydroxyl group of 5 with TBDPS group gave 6 which was treated with NaBH to give
4
diol 7. Treatment of 7 with MsCl followed by the treatment with the resulting
dimesylate 8 with Na Se (prepared in situ with Se and NaBH ), afforded the 4-
2
4
8
e
selenosugar 9 in excellent yield.

ACCEPTED MANUSCRIPT
The key intermediate 9 was first converted to the key precursors, 4'-
selenoribofuranosyl pyrimidines 12a-12f for the modification of the 2'-position, using a
8
Pummerer type condensation reaction as a key step, as shown in Scheme 2.
Scheme 2
O
O
X
NH
X
O
NH
O
RO
RO
N
Se
Se
RO
O
a
b
c
N
Se
HO
85%
4'
1'
Se
O
O
O
O
O
O
OH OH
9
10
R = TBDPS
11a (X = H) (65%)
12a (X = H) (81%)
12b (X = CH ) (94%)
12c (X = F) (98%)
12d (X = Cl) (92%)
12e (X = Br) (93%)
12f (X = I) (82%)
1
1
1
1
1
1b (X = CH ) (66%)
3
3
1c (X = F) (59%)
1d (X = Cl) (44%)
1e (X = Br) (45%)
1f (X = I) (42%)
o
Reagent and conditions: a) mCPBA, CH Cl , -78 C, 45 min; b) Base, Et N, TMSOTf, CH Cl , toluene,
2
2
3
2
2
o
0
C to rt; c) 50% TFA, THF, rt.
8
a,8e
Oxidation of 9 with m-CPBA gave the glycosyl donor, 4-selenoxide 10
.
The Pummerer-type condensation of 10 with various pyrimidine bases such as uracil,
thymine, and 5-halouracils in the presence of trimethylsilyl trifluoromethanesulfonate
(
TMSOTf) and Et N yielded the β-anomers 11a-11f, exclusively. The anomeric
3
configurations of 11a-11f were easily confirmed by the NOE experiment between 1'-H

ACCEPTED MANUSCRIPT
and 4'-H. Removal of the protecting groups of 11a-11f with 50% TFA yielded the 4'-
selenoribofuranosyl pyrimidines 12a-12f.
For the synthesis of the 2'-azido-4'-selenoarabinofuranosyl pyrimidines 3a-3f,
the ribo analogues 12a-12f were treated with TIPDSCl to give the 3',5'-O-TIPDS
2
protected nucleosides 13a-13f, respectively (Scheme 3).
Scheme 3
O
O
N
O
N
X
X
X
NBz
O
O
N
NH
NBz
O
X
a
b
c
NH
O
O
N
O
O
O
Se
Se
Se
Si
Si
Si
N3
HO
Se
OH OH
2a (X = H)
O
O
O
O
O
OH
O
OH
Si
Si
Si
1
13a (X = H, 86%)
13b (X = CH , 69%)
13c (X = F, 60%)
13d (X = Cl, 78%)
13e (X = Br, 76%)
13f (X = I, 87%)
14a (X = H, 82%)
14b (X = CH , 79%)
14c (X = F, 42%)
14d (X = Cl, 45%)
14e (X = Br, 43%)
14f (X = I, 41%)
15a (X = H, 51%)
15b (X = CH , 52%)
15c (X = F, 46%)
15d (X = Cl, 37%)
15e
(X = Br, 35%)
15f (X = I, 32%)
1
2b (X = CH
2c (X = F)
2d (X = Cl)
2e (X = Br)
2f (X = I)
3
)
3
3
3
1
1
1
1
d
O
X
O
X
NH
NBz
O
N
O
N
HO
HO
Se
Se
e
N3
N3
OH
a (X = H, 77%)
HO
3
3
3
3
3
3
16a (X = H, 80%)
16b (X = CH , 82%)
6c (X = F, 84%)
16d (X = Cl, 78%)
16e (X = Br, 80%)
16f (X = I, 73%)
b (X = CH , 76%)
3
3
1
c (X = F, 74%)
d (X = Cl, 80%)
e (X = Br, 82%)
f (X = I, 78%)
+
-
Reagents and conditions: a)TIPDSCl , pyridine, rt; b) BzCl, Et N Br , in aqueous Na CO /CH Cl , rt for 13a, 13c-13f and
2
4
2
3
2
2
o
BzCl, Et N, CH Cl , rt for 13b; c) DEAD, DPPA, PPh , THF, rt; d) 3HF.Et N, Et N, THF, 0 C; e) NH /MeOH, rt.
3
2
2
3
3
3
3
To prevent the formation of O2,2'-anhydro nucleosides during the Mitsunobu
reaction with diphenylphosphoryl azide (DPPA), the N-3 position of 13a-13f was
selectively protected with electron-withdrawing benzoyl group in the presence of the 2'-
hydroxyl group to give 14a-14f. Treatment of 14a-14f with DPPA under the Mitsunobu

ACCEPTED MANUSCRIPT
conditions afforded the desired 2'-azidoarabinofuranosyl derivatives 15a-15f with
inversion of stereochemistry, respectively. The removal of TIPDS groups of 15a-15f
with 3HF·Et N yielded 16a-16f, which were treated with methanolic ammonia to afford
3
the final 2'-azido-4'-selenoarabinofuranosyl pyrimidines 3a-3f, respectively. The
configuration of the 2'-azido group of 3a-3f was unambiguously confirmed by the
1
comparison of H NOE experiments of 3b. Irradiation on 2'-H in 3b gave the peak
enhancement by 6.0% on 4'-H, while no NOE between 2'-H and H-6 was observed,
indicating that the sugar moiety possesses an arabino configuration.
For the synthesis of 2'-fluoro-4'-selenoarabinofuranosyl pyrimidines 3g-3l, the
same intermediates 13a-13f were converted to 21g-21l, respectively which serve as the
substrates for DAST fluorination, as shown in Scheme 4. Treatment of 13a-13f with
MsCl/DBU gave the O2,2'-anhydro nucleosides 17g-17l. After the removal of the
TIPDS groups of 17g-17l with 3HF·Et N, the resulting diols 18g-18l were protected
3
with trityl (Tr) and THP groups to give 20g-20l, respectively. Opening of the O2,2'-
anhydro ring in 20g-20l with 1 M NaOH afforded 21g-21l with an arabino
configuration at the 2'-position, respectively, which serve as the substrates for DAST
fluorination.

ACCEPTED MANUSCRIPT
Scheme 4
O
N
O
O
N
X
X
X
NH
N
N
O
N
O
Se
O
Se
O
O
HO
Se
a, b
c
Si
Si
O
O
O
OH
O
HO
Si
Si
1
8g (X = H, 69%)
1
1
1
1
1
1
3a (X = H)
3b (X = CH3)
3c (X = F)
3d (X = Cl)
3e (X = Br)
3f (X = I)
17g (X = H, 77%)
1
1
1
1
1
8h (X = CH , 76%)
17h (X = CH , 85%)
3
3
8i (X = F, 79%)
8j (X = Cl, 85%)
8k (X = Br, 86%)
8l (X = I, 89%)
17i (X = F, 65%)
17j (X = Cl, 64%)
17k (X = Br, 72%)
17l (X = I, 80%)
d
O
X
O
N
O
N
X
X
NH
N
O
f
O
Se
N
e
O
Se
N
TrO
TrO
TrO
Se
OH
THPO
THPO
HO
2
2
2
2
2
2
0g (X = H, 88%)
2
2
2
2
2
2
1g (X = H, 94%)
1
9g (X = H, 69%)
0h (X = CH , 96%)
3
1h (X = CH , 96%)
3
19h (X = CH , 77%)
0i (X = F, 82%)
0j (X = Cl, 90%)
0k (X = Br, 75%)
0l (X = I, 74%)
3
1i (X = F, 95%)
1j (X = Cl, 84%)
1k (X = Br, 72%)
1l (X = I, 88%)
19i (X = F, 81%)
19j (X = Cl, 74%)
19k (X = Br, 83%)
19l (X = I, 69%)
o
.
Reagents and conditions: a) MsCl, DMAP, Et N, MC, 0 C; b) DBU, acetone, rt; c) 3HF Et N, Et N,
3
3
3
o
o
THF, 0 C; d) TrCl, DMAP, pyridine, 60 C; e) DHP, MC, rt; f) 1N NaOH, CH CN, rt
3
Treatment of 21g-21l with DAST gave the desired 2'-fluoro derivatives 22g-
2
2l with arabino configurations, which were formed by double S 2 reactions via
N
8
c
episelenium ion intermediates resulting from the participation of the selenium atom, as
shown in Scheme 5. The stereochemistry of the fluoro group of 22g-22l was confirmed
by the comparison of the X-ray crystal structure of 3g formed after the removal of all
the protecting groups in 22g (Figure 2). Treatment of 22g-22l with 80% acetic acid
afforded the final 2'-fluoro-4'-selenoarabinofuranosyl pyrimidines 3g-3l, respectively.

ACCEPTED MANUSCRIPT
For the synthesis of the 4'-selenarabinofuranosyl derivatives, 18g-18k were
treated with 1 M NaOH to yield the final 4'-selenarabinofuranosyl pyrimidines 3m-3q,
respectively (Scheme 6). Surprisingly, the reaction of 5-iodo derivative 18l under these
set of conditions resulted in the formation of the dehalogenated uracil compound (3m,
5
4%) instead of giving 3r. Thus, the synthesis of the 5-iodo derivative 3r was

ACCEPTED MANUSCRIPT
completed by the Mitsunobu reaction on 14f followed by the removals of all protecting
groups, as shown in Scheme 7.
Scheme 6
O
O
N
X
N
X
NH
O
Se
N
1 N NaOH
HO
O
HO
CH CN
Se
OH
3
OH
OH
1
1
1
1
1
1
8g (X = H)
8h (X = CH3)
8i (X = F)
3m (X = H, 94%)
3n (X = CH , 99%)
3
3
o (X = F, 95%)
8j (X = Cl)
(X = Br)
3p (X = Cl, 61%)
3q (X = Br, 57%)
8k
8l (X = I)
3
m (X = H, 54%)

ACCEPTED MANUSCRIPT
Scheme 8
O
N
O
NH
NH
O
N
O
a
AcO
HO
Se
Se
OH
R
R
OAc
2
2
2
3s (R = N3)
3t (R = F)
3u (R = OH)
3
3
3
a (R = N3)
g (R = F)
m (R = OH)
b
N
NH2
N
N
N
N
c, d
N
O
HO
N
O
Se
OH
AcO
R
Se
R
OAc
24s (R = N3)
4t (R = F)
4u (R = OH)
3
3
3
s (R = N , 46% from 3a)
t (R = F, 59% from 3g)
u (R = OH, 54% from 3m)
3
2
2
Reagents and conditions: a) Ac O, rt; b) 1,2,4-triazole, Et N,
2
3
POCl , CH CN, rt; c) NH OH, 1,4-dioxane, rt; d) NH /MeOH, rt.
3
3
4
3
The uracil derivatives 3a, 3g, and 3m were converted to the corresponding
8
c
cytosine derivatives 3s, 3t, and 3u, respectively, as illustrated in Scheme 8. The uracil
derivatives 3a, 3g, and 3m were peracetylated to give 23s, 23t, and 23u, respectively.
Treatment of 23s, 23t, and 23u with POCl , 1,2,4-triazole, and Et N gave the
3
3
corresponding triazole derivatives 24s, 24t, and 24u, which were immediately used in
the next step just after work-up because they are generally unstable during the

ACCEPTED MANUSCRIPT
purification by silica gel column chromatography. Compounds 24s, 24t, and 24u were
successively treated with NH OH in 1,4-dioxane and methanolic ammonia to afford the
4
8
c
cytosine derivatives 3s, 3t, and 3u, respectively.
Antitumor activity evaluation
All the final 2'-modified-4'-selenoarabinofuranosyl pyrimidines 3a-3u were
assayed for cytotoxic effects in several human cancer cell lines such as colon cancer
(HCT116), lung cancer (A549), stomach cancer (SNU638), breast cancer (T47D),
prostate cancer (PC-3), and leukemia (K562) cells, using sulforhodamine B (SRB)
1
0
protein staining method (Table 1).
Table 1. Anticancer activity of all the final 2'-modified-4'-selenonucleosides, 3a-3u in
several human cancer cell lines.
a
IC (ꢀM)
5
0
Compound No.
b
c
d
e
f
g
HCT116
> 100
> 100
> 100
> 100
> 100
> 100
> 100
85.2
A549
SNU638
> 100
> 100
> 100
> 100
> 100
> 100
> 100
77.2
T47D
PC-3
K562
3
3
3
3
3
3
3
3
a (X = OH, Y = H, R = N3)
b (X = OH, Y = Me, R = N3)
c (X = OH, Y = F, R = N3)
d (X = OH, Y = Cl, R = N3)
e (X = OH, Y = Br, R = N3)
f (X = OH, Y = I, R = N3)
> 100
> 100
> 100
> 100
> 100
> 100
> 100
76.4
> 100
> 100
> 100
> 100
> 100
> 100
> 100
65.3
> 100
> 100
> 100
> 100
> 100
> 100
> 100
88.0
> 100
> 100
> 100
> 100
> 100
> 100
> 100
76.3
h
g (X = OH, Y = H, R = F)
h (X = OH, Y = Me, R = F)

ACCEPTED MANUSCRIPT
3
3
3
3
3
3
3
3
3
3
3
3
3
i (X = OH, Y = F, R = F)
j (X = OH, Y = Cl, R = F)
k (X = OH, Y = Br, R = F)
l (X = OH, Y = I, R = F)
m (X = OH, Y = H, R = OH)
40.1
80.2
54.2
82.1
50.1
76.3
51.1
77.1
43.1
> 100
> 100
> 100
> 100
> 100
66.3
> 100
> 100
> 100
75.1
78.5
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
> 100
62.3
> 100
> 100
> 100
> 100
56.8
> 100
> 100
> 100
78.3
> 100
> 100
> 100
> 100
55.2
> 100
> 100
> 100
80.1
> 100
> 100
> 100
> 100
60.1
> 100
> 100
> 100
85.2
> 100
> 100
> 100
> 100
67.4
> 100
> 100
> 100
78.4
h
n (X = OH, Y = Me, R = OH)
o (X = OH, Y = F, R = OH)
p (X = OH, Y = Cl, R = OH)
q (X = OH, Y = Br, R = OH)
r (X = OH, Y = I, R = OH)
s (X = NH , Y = H, R = N )
2
3
h
t (X = NH , Y = H, R = F)
1.1
7.13
5.3
0.47
8.83
1.90
0.14
4.72
0.15
0.79
8.91
2.72
d
0.58
4.54
55.9
0.63
86.6
0.05
2
h
u (X = NH , Y = H, R = OH)
2
h
Ara-C (2)
a
b
c
measured using SRB method; human colon cancer cell lines; human lung cancer cell lines; human stomach cancer cell
e
f
g
h
lines; human breast cancer cell lines; human prostate cancer cell lines; human leukemia cell lines; ref. 8c; ND.
As shown in Table 1, all the cytosine derivatives 3s-3u showed significant
anticancer activity in human tumor cell lines tested, among which the 2'-fluoro
derivative 3t was the most potent and was more potent than the control, Ara-C (2)
8
c
except human leukemia cell lines (K562). The anticancer activity of the 2'-modified-
'-selenoarabinofuranosyl cytosines decreased in the following order: 2'-F derivative 3t
R = F) > 2'-OH derivative 3u (R = OH) > 2'-N derivative 3s (R = N ). In case of all
4
(
3
3
the uracil, thymine, and 5-halouracil derivatives, the same trend was also observed as
the cytosine derivatives: 2'-F derivative 3i (R = F) > 2'-OH derivative 3o (R = OH),
although they showed very weak activity. Interestingly, the 5-fluorouracil derivatives 3i
and 3o with 2'-F and 2'-OH substitution exhibited more potent anticancer activity than
other uracil, thymine, and 5-halouracil derivatives. All the 2'-azido derivatives did not

ACCEPTED MANUSCRIPT
show significant anticancer activity up to 100 ꢀM, except the cytosine derivative 3s
showing weak anticancer activity.
In order to determine if the anticancer activity of 3t depends on the conversion
to the corresponding triphosphate by cellular kinases, which are essential for the
inhibition of DNA and/or RNA cellular polymerase, 3t was incubated in the cell and the
metabolites were analyzed by LC-MS. Surprisingly, none of any mono-, di-, or
triphosphate was detected as the major metabolites unlike ara-C (2), which was
1
1
converted to the corresponding triphosphate, indicating that the anticancer activity of
3
t might come from different mechanism of action from that of ara-C (2) 16, lines used
in the treatment of leukemia. This cellular metabolism study might explain why 3t is
less potent against human leukemia cell lines (K562) and more potent against solid
tumor cell lines (HCT116, A549, SNU638, T47D, and PC-3) than ara-C (2). This result
also demonstrates that all synthesized compounds might not act as cellular DNA and/or
RNA polymerase inhibitors because of no phosphorylation by cellular kinases. No
phosphorylation of 3t by cellular kinases might be attributed to the different sugar
1
2
8
13
puckering of 4-selenosugar . As shown by the X-ray crystal structure of 3g (Figure
), bulky selenium atom pushes 3g to adopt the 2'-endo/3'-exo (South) conformation
with pseudorotation parameters of P = 0.033ꢀ and τ =46.2ꢀ. When compared with the
2
m

ACCEPTED MANUSCRIPT
corresponding 4'-oxo derivative (2’-deoxy-2’-fluorouridine), which takes the unusual 4'-
exo/1'-endo sugar puckering with P = 70.84ꢀ and τ =38.2ꢀ, the conformation of 3g
m
appears to be affected by selenium atom. Most of the 2’-fluoro derivatives including 2’-
deoxy-2’-fluorocytidine and 2’-deoxy-2’-fluoro-4’-thiocytidine shows typical 2’-exo/3’-
endo (North) conformation due to the gauche effect and this conformational difference
will affect the orientation of the 5'-OH which is essential for the phosphorylation.
Figure 2. The X-ray crystal structure of 2’-deoxy-2’-fluoro-4’-selenouridine (3g)
Conclusions
On the basis of potent anticancer activity of ara-C (2) with the 2'-arabino
configuration, the 2'-deoxy-4'-selenoarabinofuranosyl pyrimidines 3a-3u with the 2'-
azido, 2'-fluoro, or 2'-hydroxyl substitution were synthesized from
D-ribose and
evaluated for cytotoxic effects in a panel of human tumor cell lines. The highlight of our

ACCEPTED MANUSCRIPT
synthetic endeavor is the Pummerer-type base condensation, Mitsunobu reaction with
DPPA for the introduction of the 2'-azido group, and stereoselective fluorination via the
episelenium intermediate. Among compounds tested, the 2'-fluoro derivative 3t was
found to exhibit the most potent anticancer activity and showed better anticancer
activity than the reference, ara-C. Synthesis, conformational study, and cellular study
executed in this study will provide the medicinal chemist with great insight into the
design of novel modified nucleosides.
Experimental Section
General methods.
1
H-NMR Spectra (CDCl , CD OD or DMSO-d ) were recorded on Varian Unity Invoa
3
3
6
1
4
00 MHz. The H-NMR data are reported as peak multiplicities: s for singlet, d for doublet, dd
for doublet of doublets, t for triplet, q for quartet, br s for broad singlet and m for multiplet.
1
3
Coupling constants are reported in Hertz. C-NMR spectra (CDCl , CD OD or DMSO-d ) were
3
3
6
1
9
recorded on Varian Unity Inova 100 MHz. F-NMR spectra (CDCl , CD OD) were recorded on
3
3
Varian Unity Inova 376 MHz. The chemical shifts were reported as parts per million (δ) relative
to the solvent peak. Optical rotations were determined on Jasco III in appropriate solvent. UV
spectra were recorded on U-3000 made by Hitachi in methanol or water. Infrared spectra were

ACCEPTED MANUSCRIPT
recorded on FT-IR (FTS-135) made by Bio-Rad. Melting points were measured on B-540 made
by Buchi. Elemental analyses (C, H, and N) were used to determine purity of all synthesized
compounds, and the results were within ± 0.4% of the calculated values, confirming > 95%
purity. The progress of reactions was monitored via thin layer chromatography (TLC, Merck
precoated 60 F254 plates) and ethyl acetate:hexane or dichloromethane/methanol. The TLC
plates were visualized using UV lamp (254 nM) and/or staining in anisaldehyde solution with
acetic acid, sulfuric acid and methanol. Flash column chromatography was performed using
silica gel 60 (230-400 mesh, Merck) and ethyl acetate:hexane or dichloromethane/methanol
mixtures as the mobile phase. Reagents were purchased from Aldrich Chemical Company.
Solvents were obtained from local suppliers. All the anhydrous solvents were distilled over
CaH , P O or sodium/benzophenone prior to the reaction. All reactions sensitive to air or
2
2
5
moisture were conducted under nitrogen atmosphere, unless otherwise stated. The yields given
refer to purified products after column chromatography or recrystallization with appropriate
solvents. 2.3-O-isopropyledene-L-lyxono-1,4-lactone (5) was synthesized from D-ribose using a
9
known procedure. Compounds 6-9 were synthesized according to our previously reported
8
e
procedure.
General procedure for the synthesis of β-anomers 11a-11f.

ACCEPTED MANUSCRIPT
To a stirred solution of 4-selenosugar 9 in dichloromethane was added a solution of m-CPBA
°
(
1.1 equiv.) in dichloromethane at -78 C and the mixture was stirred at the same temperature for
4
5 min. The reaction mixture was quenched with saturated NaHCO solution and then extracted
3
with dichloromethane. The organic layer was washed with saturated NaHCO , brine, dried over
3
MgSO , filtered and evaporated under reduced pressure. The residue was purified on flash silica
4
gel column chromatography (dichloromethane:methanol = 30:1) to give selenoxide 10 (85%) as
a colorless syrup. Due to the unstable nature of selenoxide 10, it was immediately used for the
next step.
To a suspension of appropriate pyrimidine base (2 equiv.) in toluene were added
triethylamine (4.1 equiv.) and trimethylsilyl trifluoromethanesulfonate (6.1 equiv.) and the
mixture were stirred at room temperature for 1 h. The silylated base solution was diluted with
additional dichloromethane and this solution was then added to a solution of 10 in
°
dichloromethane dropwise over a period of 20 min at 0 C. An additional amount of
triethylamine (2.1 equiv.) was added dropwise to the reaction mixture to initiate the Pummerer
°
reaction at 0 C. After the reaction mixture was stirred at room temperature for 15 h, the reaction
mixture was quenched with saturated NaHCO and extracted with dichloromethane. The organic
3
layers were washed with saturated NaHCO solution, water and brine, dried over MgSO ,
3
4
filtered and evaporated under reduced pressure. The residue was purified on flash silica gel

ACCEPTED MANUSCRIPT
column chromatography (hexane:ethyl acetate = 2:1) to give the β-anomers 11a-11f.
(
-)-1-[5-O-(tert-Butyldiphenylsilyl)-2,3-O-isopropylidene-4-seleno-
D
-ribofuranosyl]uracil (11a).
2
0
1
Light yellow foam; Yield: 65%; [α]D -47.74 (c 0.22, CH OH); UV (CH OH) λ 265 nm; H
3
3
max
NMR (400 MHz, CDCl ) δ 8.45 (brs, 1H), 7.69-7.63 (m, 4H), 7.52 (d, J = 8.4 Hz, 1H), 7.45-
3
7
1
.37 (m, 6H), 6.35 (d, J = 4.0 Hz, 1H), 5.56 (dd, J = 2.0, 8.0 Hz, 1H), 4.73 (dd, J = 4.2, 5.2 Hz,
H), 4.67 (dd, J = 4.2, 5.2 Hz, 1H), 4.02-3.95 (m, 3H), 1.56 (s, 3H), 1.28 (s, 3H), 1.08 (s, 9H);
1
3
C NMR (100 MHz, CDCl ) δ 150.0, 142.0, 135.8, 135.7, 133.2, 132.8, 130.3, 128.2, 128.1,
3
1
5
4
12.4, 103.5, 90.1, 85.3, 76.6, 66.1, 59.7, 50.3, 28.0, 27.1, 27.0, 26.8, 25.5 19.5; MS (FAB) m/z
+
87 [M+H] ; Found: C, 57.04; H, 5.86; N, 4.66. Calc. for C H N O SeSi: C, 57.43; H, 5.85; N,
2
8
34
2
5
.78%.
(
(
-)-1-[5-O-(tert-Butyldiphenylsilyl)-2,3-O-isopropylidene-4-seleno-
D
-ribofuranosyl]thymine
11b).
2
0
1
Light yellow foam; Yield: 66%; [α]D -29.79 (c 5.83, CH Cl ); UV (CH OH) λ 278 nm; H
2
2
3
max
NMR (400 MHz, CDCl ) δ 8.13 (s, 1H), 7.69-7.67 (m, 4H), 7.43-7.37 (m, 6H), 6.33 (s, 1H),
3
1
3
4
.71 (m, 2H), 4.04 (m, 2H), 3.92 (m, 1H), 1.83 (s, 3H), 1.56 (s, 3H), 1.08 (s, 9 H); C NMR
(
100 MHz, CD OD) δ 140.1, 136.9, 136.8, 134.6, 134.3, 131.3, 131.2, 129.1, 129.0, 112.9,
3
1
12.6, 90.9, 87.0, 79.7, 67.7, 61.2, 52.4, 28.2, 27.4, 25.6, 20.2, 12.5; MS (FAB) m/z 601
+
[
M+H] ; Found: C, 58.30; H, 6.48; N, 4.37. Calc. for C H N O SeSi: C, 58.09; H, 6.05; N,
2
9
36
2
5
4
.67%.
(
-)-5-Fluoro-1-[5-O-(tert-butyldiphenylsilyl)-2,3-O-isopropylidene-4-seleno-
ribofuranosyl]uracil (11c).
Light yellow foam; Yield: 59%; [α]D -21.52 (c 7.15, CH Cl ); UV (CH Cl ) λ 284 nm; H
D-
2
0
1
2
2
2
2
max
NMR (400 MHz, CDCl ) δ 7.69-7.66 (m, 4H), 7.61 (d, J = 2.0 Hz, 1H), 7.48-7.38 (m, 6H), 6.34
3

ACCEPTED MANUSCRIPT
(
dd, J = 1.2, 4.4 Hz, 1H), 4.68 (dd, J = 3.2, 5.6 Hz, 1H), 4.62 (t, J = 4.4 Hz, 1H), 4.05-4.00 (m,
1
3
2
H), 3.92 (dd, J = 9.2, 12.4 Hz, 1H), 1.57 (s, 3H), 1.28 (s, 3H), 1.09 (s, 9H); C NMR (100
MHz, CDCl ) δ 156.67 (d, J = 27.3 Hz), 148.9, 140.6 (d, J = 239.3 Hz), 135.8 (d, J = 8.8 Hz),
3
1
6
33.0 (d, J = 11.0 Hz), 130.2, 128.1 (d, J = 1.5 Hz), 126.0 (d, J = 33.8 Hz), 112.7, 89.7, 84.7,
1
9
6.0, 59.8, 50.1, 28.0, 27.1, 25.5, 19.5; F NMR (376 MHz, CDCl ) δ -161.88 (d, J = 5.2 Hz);
3
+
MS (ESI) m/z 643.0957 [M+K] ; Found: C, 55.78; H, 5.14; N, 4.62. Calc. for
C H FN O SeSi: C, 55.71; H, 5.51; N, 4.64%.
28
33
2
5
(
-)-5-Chloro-1-[5-O-(tert-butyldiphenylsilyl)-2,3-O-isopropylidene-4-seleno-
ribofuranosyl]uracil (11d).
Light yellow foam; Yield: 44%; [α]D -29.31 (c 1.01, CH Cl ); UV (CH Cl ) λ 283 nm; H
D-
2
0
1
2
2
2
2
max
NMR (400 MHz, CDCl ) δ 8.64 (brs, 1H), 7.81 (s, 1H), 7.67 (m, 4H), 7.42 (m, 6H), 6.30 (d, J =
3
4
.8 Hz, 1H), 4.69 (dd, J = 3.2, 5.6 Hz, 1H), 4.63 (dd, J = 4.4, 6.0 Hz, 1H), 4.03 (m, 2H), 3.91
1
3
(
dd, J = 9.2, 12.4 Hz, 1H), 1.56 (s, 3H), 1.27 (s, 3H), 1.08 (s, 9H); C NMR (100 MHz, CDCl )
3
δ 158.7, 149.5, 138.8, 135.9, 133.1, 130.2, 128.1, 112.7, 110.1, 89.7, 84.6, 65.9, 59.7, 50.4, 27.9,
2
5
7.1, 25.5, 19.5; Found: C, 54.12; H, 5.67; N, 4.23. Calcd for C H ClN O SeSi: C, 54.24; H,
28 33 2 5
.36; N, 4.52%.
(
-)-5-Bromo-1-[5-O-(tert-butyldiphenylsilyl)-2,3-O-isopropylidene-4-seleno-
ribofuranosyl]uracil (11e).
Light yellow foam; Yield: 45%; [α]D -25.41 (c 6.49, CH Cl ); UV (CH Cl ) λ 285 nm; H
D-
2
0
1
2
2
2
2
max
NMR (400 MHz, CDCl ) δ 9.41 (s, 1H), 7.82 (s, 1H), 7.69-7.66 (m, 4H), 7.47-7.38 (m, 6H),
3
6
.29 (d, J = 4.4 Hz, 1H), 4.72-4.65 (m, 2H), 4.07-3.90 (m, 3H), 1.56 (s, 3H), 1.28 (s, 3H), 1.09
13
(
s, 9H); C NMR (100 MHz, CDCl ) δ 158.9, 149.7, 141.4, 135.8, 133.0, 130.2, 128.1, 112.7,
3
9
7.9. 89.7, 84.6, 66.0, 59.7, 50.5, 28.0, 27.1, 25.5, 19.5; Found: C, 50.66; H, 5.00; N, 4.21. Calc.
for C H BrN O SeSi: C, 50.61; H, 5.01; N, 4.22%.
28
33
2
5

ACCEPTED MANUSCRIPT
(
-)-5-Iodo-1-[5-O-(tert-butyldiphenylsilyl)-2,3-O-isopropylidene-4-seleno-D-
ribofuranosyl]uracil (11f).
Light yellow foam; Yield: 42%; [α]D -52.4 (c 0.11, CH OH); UV λ (CH OH) 284 nm; H
2
0
1
3
max
3
NMR (400 MHz, CDCl ) δ 8.71 (s, 1H), 7.89 (s, 1H), 7.69-7.66 (m, 4H), 7.48-7.38 (m, 6H),
3
6
1
1
2
3
.24 (d, J = 4.4 Hz, 1H), 4.71-4.63 (m, 2H), 4.06-4.01 (m, 2H), 3.95-3.89 (m, 1H), 1.55 (s, 3H),
1
3
.27 (s, 3H), 1.09 (s, 9H); C NMR (100 MHz, CDCl ) δ 159.6, 149.8, 146.5, 135.9, 135.8,
3
33.1, 133.0, 130.24, 130.21, 128.1, 128.0, 112.8, 89.7, 84.6, 69.4, 66.0, 59.6, 50.6, 28.0, 27.1,
5.5, 19.5; Found C, 46.99; H, 4.97; N, 3.99. Calc. for C H IN O SeSi: C, 47.26; H, 4.67; N,
2
8
33
2
5
.94%.
General procedure for the synthesis of ribofuranosyl analogues 12a-12f.
To a solution of compound 11a-11f in tetrahydrofuran was added 50% aqueous trifluoroacetic
o
acid at 0 C and the mixture was allowed to stir at room temperature for 15 h. The solvent was
evaporated under reduced pressure and then co-evaporated under reduced pressure three times
with dry toluene to give 12a-12f.
(
-)-1-(4-Seleno-
D
-ribofuranosyl)uracil (12a).
o
20
White solid; Yield: 81%; mp 198-200 C; [α] -113.93 (c 0.33, CH OH); UV (CH OH) λ
D
3
3
max
1
2
67 nm; H NMR (400 MHz, CD OD) δ 8.00 (d, J = 8.0 Hz, 1H), 6.11 (d, J = 8.8 Hz, 1H), 5.77
3
(
d, J = 8.0 Hz, 1H), 4.26 (dd, J = 3.6, 8.4 Hz, 1H), 4.19 (t, J = 2.5 Hz, 1H), 3.75 (dd, J = 7.8,
1
3
1
1.4 Hz, 1H), 3.63 (dd, J = 7.8, 11.4 Hz, 1H), 3.44-3.40 (m, 1H); C NMR (100 MHz, CD OD)
3
+
δ 163.0, 150.9, 142.1, 102.1, 77.0, 73.5, 63.6, 55.3, 48.8; MS (FAB) m/z 307 (M ); Found: C,
3
4.93; H, 3.82; N, 9.07. Calc. for C H N O Se: C, 35.19; H, 3.94; N, 9.12%.
9 12 2 5
(
-)-1-(4-Seleno-D-ribofuranosyl)thymine (12b).

ACCEPTED MANUSCRIPT
o
20
White solid, Yield: 94%; mp 128-130 C; [α] -111.62; (c 0.86, CH OH); UV (CH OH) λ
max
D
3
3
1
2
8
1
7
8
75 nm; H NMR (400 MHz, CD OD) δ 7.91 (s, 1H), 6.29 (d, J = 8.4 Hz, 1H), 4.38 (dd, J = 3.6,
3
.0 Hz, 1H), 4.26 (t, J = 2.8 Hz, 1H), 3.88 (dd, J = 6.4, 11.6 Hz, 1H), 3.82 (dd, J = 5.6, 11.6 Hz,
1
3
H), 3.56-3.53 (m, 1H), 1.91 (s, 3H); C NMR (CD OD) δ 166.3, 153.1, 139.6, 112.2, 80.1,
3
+
6.4, 65.2, 57.6, 30.8, 12.6; MS (ESI) m/z 344.9961 [M+Na] ; Found: C, 37.53; H, 4.02; N,
.47. Calc. for C H N O Se: C, 37.39; H, 4.39; N, 8.72%.
1
0
14
2
5
(
-)-1-(4-Seleno-
D
-ribofuranosyl)-5-fluorouracil (12c).
o
20
White solid; Yield: 98%; mp 199-200 C; [α] -26.84 (c 0.79, CH OH); UV (CH OH) λ 275
D
3
3
max
1
nm; H NMR (400 MHz, CD OD) δ 8.31 (t, J = 3.4 Hz, 1H), 6.27 (dd, J = 1.6, 7.6 Hz, 1H),
3
4
.37 (dd, J = 3.4, 7.8 Hz, 1H), 4.26 (t, J = 3.0 Hz, 1H), 3.90-3.81 (m, 2H), 3.58-3.54 (m, 1H);
1
3
C NMR (100 MHz, CD OD) δ 159.2 (d, J = 101.6 Hz), 151.3, 142.5, 140.2, 127.7 (d, J =
3
1
9
1
3
3
39.6 Hz), 80.2, 76.3, 64.6, 58.3; F NMR (CD OD) δ -166.65 (t, J = 5.65 Hz); MS (ESI) m/z
3
+
64.9437 [M+K] ; Found: C, 33.64; H, 3.79; N, 8.43. Calc. for C H FN O Se: C, 33.24; H,
9
11
2
5
.41; N, 8.62%.
(
-)-1-(4-Seleno-D-ribofuranosyl)-5-chlorouracil (12d).
°
25
White solid; Yield: 92%; mp 220-223 C; [α]D -27.87 (c 0.24, CH OH); UV (CH OH) λ 280
3
3
max
1
nm; H NMR (400 MHz, CD OD) δ 8.42 (s, 1H), 6.25 (d, J = 8.0 Hz, 1H), 4.39 (dd, J = 3.2, 7.6
3
1
3
Hz, 1H), 4.24 (t, J = 3.2 Hz, 1H), 3.90-3.81 (m, 2H), 3.59-3.55 (m, 1H); C NMR (100 MHz,
CD OD) δ 161.4, 152.0, 141.1, 109.7, 80.4, 76.5, 64.8, 58.4, 50.1; MS (ESI) m/z 364.9613
3
+
[
M+Na] ; Found: C, 31.89; H, 3.33; N, 8.23. Calc. for C H ClN O Se: C, 31.64; H, 3.25; N,
9
11
2
5
8
.20%.
(
-)-1-(4-Seleno-D-ribofuranosyl)-5-chlorouracil (12e).
°
20
White solid; Yield: 93%; mp 169-173 C; [α] -61.81 (c 1.11, CH OH+H O); UV (CH OH)
D
3
2
3
1
λmax 282 nm; H NMR (400 MHz, CD OD) δ 8.51 (s, 1H), 6.25 (d, J = 7.6 Hz, 1H), 4.39 (dd, J
3

ACCEPTED MANUSCRIPT
1
3
=
3.6, 7.6 Hz, 1H), 4.24 (t, J = 3.0 Hz, 1H), 3.90-3.81 (m, 2H), 3.59-3.55 (m, 1H); C NMR
(
100 MHz, CD OD) δ 161.5, 152.2, 143.7, 97.4, 80.5, 76.5, 64.7, 58.4, 50.1; MS (ESI) m/z
3
+
4
2
08.8908 [M+Na] ; Found: C, 27.89; H, 3.03; N, 7.44. Calc. for C H BrN O Se: C, 28.00; H,
9
11
2
5
.87; N, 7.26%.
(
-)-1-(4-Seleno-D-ribofuranosyl)-5-iodouracil (12f).
°
20
White solid; Yield: 82%; mp 242-244 C; [α]
-83.50 (c 0.61, CH OH+H O); UV λ
3 2 max
D
1
(
(
CH OH) 292 nm; H NMR (400 MHz, CD OD) δ 8.55 (s, 1H), 6.22 (d, J = 7.6 Hz, 1H), 4.38
3
3
1
3
dd, J = 3.6, 7.6 Hz, 1H), 4.23 (t, J = 3.2 Hz, 1H), 3.90-3.81 (m, 2H), 3.59-3.56 (m, 1H); C
NMR (100 MHz, CD OD) δ 160.4, 150.8, 146.4, 77.5, 73.8, 69.9, 63.4, 55.9, 49.3; MS (ESI)
3
+
m/z 456.8775 [M+Na] ; Found: C, 25.04; H, 2.54; N, 6.07. Calc. for C H IN O Se: C, 24.96; H,
9
11
2
5
2
.56; N, 6.47%.
General procedure for the synthesis of 13a-13f.
To a solution of compound 12a-12f in pyridine was added 1,3-dichloro-1,1,3,3-tetraisopropyl
°
disiloxane (1.5 equiv.) at 0 C and the mixture was allowed to stir at room temperature for 3 h.
The reaction mixture was concentrated, and the residue co-evaporated under reduced pressure
three times with toluene. The residue was purified by silica gel column chromatography
(
hexane:ethyl acetate = 2:1) to give 13a-13f.
(
-)-1-[3,5-O-(1,1,3,3-Tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
D
-ribofuranosyl]uracil (13a).
2
0
1
White foam; Yield: 86%; [α]D -1.31; (c 0.84, CH Cl ); UV (CH Cl ) λ 265 nm; H NMR
2
2
2
2
max
(
400 MHz, CDCl ) δ 9.29 (brs, 1H), 8.20 (d, J = 8.0 Hz, 1H), 6.06 (s, 1H), 5.72 (d, J = 8.0 Hz,
3
1
H), 4.34 (dd, J = 3.2, 9.2 Hz, 1H), 4.21 (t, J = 1.2 Hz, 1H), 4.14 (dd, J = 3.2, 13.2 Hz, 1H),

ACCEPTED MANUSCRIPT
4
.02 (d, J = 1.6, 12.8 Hz, 1H), 3.88 (dt, J = 2.4, 9.6 Hz, 1H), 3.19 (d, J = 1.6 Hz, 1H), 1.14-0.97
1
3
(
m, 28H); C NMR (100 MHz, CDCl ) δ 163.3, 150.7, 143.0, 102.4, 80.3, 74.3, 59.0, 57.7, 46.4,
3
1
5
6
7.8, 17.7, 17.55, 17.53, 17.35, 17.27, 17.22, 17.1, 13.6, 13.5, 13.3, 12.7; MS (ESI) m/z
+
73.1326 [M+Na] ; Found: C, 45.89; H, 6.99; N, 5.45. Calc. for C H N O SeSi : C, 45.89; H,
2
1
38
2
6
2
.97; N, 5.10%.
(
(
+)-1-[3,5-O-(1,1,3,3-Tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
D
-ribofuranosyl]thymine
13b).
2
0
1
White foam; Yield: 69%; [α]D +12.70 (c 7.78, CH Cl ); UV (CH Cl ) λ 271 nm; H NMR
2
2
2
2
max
(
400 MHz, CDCl ) δ 8.63 (brs, 1H), 7.71 (d, J = 1.2 Hz, 1H), 6.11 (d, J = 1.6 Hz, 1H), 4.28 (dd,
3
J = 3.6, 9.2 Hz, 1H), 4.22-4.11 (m, 2H), 4.04-4.00 (m, 1H), 3.92-3.89 (m, 1H), 2.91 (brs, 1H),
1
3
1
1
1
.90 (d, J =1.2 Hz, 3H), 1.15-0.99 (m, 28H); C NMR (100 MHz, CDCl ) δ 163.7, 150.6, 138.1,
3
11.4, 80.2, 74.8, 59.1, 56.9, 46.9, 17.7, 17.69, 17.6, 17.5, 17.4, 17.3, 17.2, 17.1, 13.6, 13.5,
+
3.1, 12.7, 12.6; MS (ESI) m/z 587.1504 [M+Na] ; Found: C, 46.76; H, 7.32; N, 5.01. Calc. for
C H N O SeSi : C, 46.88; H, 7.15; N, 4.97%.
22
40
2
6
2
(
+)-5-Fluoro-1-[3,5-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
ribofuranosyl]uracil (13c).
White foam; Yield: 60%; [α]D +21.48 (c 2.91, CH Cl ); UV (CH Cl ) λ 272 nm; H NMR
D-
2
0
1
2
2
2
2
max
(
400 MHz, CDCl ) δ 8.31 (d, J = 6.4 Hz, 1H), 6.04 (s, 1H), 4.32 (dd, J = 3.2, 9.2 Hz, 1H), 4.22
3
(
d, J = 3.2 Hz, 1H), 4.14 (dd, J = 3.2, 12.8 Hz, 1H), 4.02 (dd, J = 1.6, 12.8 Hz, 1H), 3.89 (m,
13
1
H), 1.14-0.98 (m, 28H); C NMR (100 MHz, CDCl ) δ 156.9 (d, J = 27.3 Hz), 149.2, 140.0 (d,
3
J = 237.9 Hz), 127.2 (d, J = 35.3 Hz), 80.3, 74.3, 58.8, 58.1, 46.8, 17.7, 17.6, 17.5, 17.4, 17.3,
1
9
1
7.2, 17.1, 13.6, 13.5, 13.3, 12.7; F NMR (376 MHz, CDCl ) δ -163.0 (d, J = 6.02 Hz); MS
3
+
(
ESI) m/z 569.1416 [M+H] ; Found: C, 44.13; H, 6.34; N, 4.76. Calc. for C H FN O SeSi : C,
2
1
37
2
6
2
4
4.43; H, 6.57; N, 4.93%.

ACCEPTED MANUSCRIPT
(
-)-5-Chloro-1-[3,5-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl)-4-seleno-D-
ribofuranosyl]uracil (13d).
White foam; Yield: 78%; [α]D -4.61 (c 6.77, CH Cl ); UV (CH Cl ) λ 281 nm; H NMR
2
0
1
2
2
2
2
max
(
400 MHz, CDCl ) δ 9.62 (brs, 1H), 8.34 (s, 1H), 6.05 (d, J = 1.6 Hz, 1H), 4.33 (dd, J = 3.6, 9.6
3
Hz, 1H), 4.23 (d, J = 2.4 Hz, 1H), 4.14 (dd, J = 3.2, 13.2 Hz, 1H), 4.02 (dd, J = 2.0, 13.2 Hz,
1
3
1
H), 3.93 (dt, J = 3.2, 9.6 Hz, 1H), 3.35 (brs, 1H), 1.14-0.98 (m, 28H); C NMR (100 MHz,
CDCl ) δ 159.0, 149.9, 139.5, 109.5, 80.1, 74.4, 58.7, 57.9, 47.1, 17.8, 17.7, 17.6, 17.5, 17.4,
3
1
7.3, 17.2, 17.1, 13.6, 13.5, 13.2, 12.7; Found: C, 43.58; H, 6.08; N, 4.43. Calc. for
C H ClN O SeSi : C, 43.18; H, 6.38; N, 4.80%.
21
37
2
6
2
(
-)-5-Bromo-1-[3,5-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
ribofuranosyl]uracil (13e).
White foam; Yield: 76%; [α]D -20.36 (c 4.70, CH Cl ); UV (CH Cl ) λ 283 nm; H NMR
D-
2
0
1
2
2
2
2
max
(
400 MHz, CDCl ) δ 9.7 (brs, 1H), 8.41 (s, 1H), 6.04 (s, 1H), 4.32 (dd, J = 3.4, 9.4 Hz, 1H),
3
4
.23 (d, J = 2.4 Hz, 1H), 4.14 (dd, J = 3.0, 13.0 Hz, 1H), 4.02 (dd, J = 2.0, 13.2 Hz, 1H), 3.96-
1
3
3
.92 (m, 1H), 3.42 (brs, 1H), 1.15-0.99 (m, 28H) ; C NMR (100 MHz, CDCl ) δ 159.1, 150.2,
3
1
42.1, 97.3, 80.0, 74.5, 58.7, 57.9, 47.1, 17.9, 17.8, 17.7, 17.5, 17.4, 17.3, 17.2, 17.1, 13.6, 13.5,
3.3, 12.7; Found: C, 40.01; H, 5.65; N, 4.77. Calc. for C H BrN O SeSi : C, 40.13; H, 5.93;
1
2
1
37
2
6
2
N, 4.46%.
(
(
-)-5-Iodo-1-[3,5-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
D
-ribofuranosyl]uracil
13f).
2
0
1
White foam; Yield: 87%; [α]D -46.9, (c 0.60, CH Cl ); UV λ (CH OH) 292 nm; H NMR
2
2
max
3
(
400 MHz, CDCl ) δ 8.92 (s, 1H), 8.37 (s, 1H), 6.06 (d, J = 1.6 Hz, 1H), 4.39 (dd, J = 3.6, 9.2
3
Hz, 1H), 4.21 (t, J = 1.6 Hz, 1H), 4.15-4.00 (m, 2H), 3.94-3.91 (m, 1H), 2.94 (d, J = 2.0 Hz,
1
3
1
H), 1.17-1.01 (m, 28H); C NMR (100 MHz, CDCl ) δ 159.9, 150.2, 146.9, 80.2, 74.9, 68.8,
3

ACCEPTED MANUSCRIPT
5
9.0, 57.1, 47.4, 18.03, 18.00, 17.7, 17.5, 17.4, 17.3, 17.2, 17.1, 13.6, 13.5, 13.3, 12.7; Found: C,
7.73; H, 5.12; N, 3.87. Calc. for C H IN O SeSi : C, 37.34; H, 5.52; N, 4.15%.
3
2
1
37
2
6
2
General procedure for the synthesis of 14a and 14c-14f.
To a solution of compound 13a, 13c-13f in dichloromethane were added benzoyl chloride (1.05
equiv.) and tetrabutylammonium bromide (0.3 equiv.) at room temperature and the mixture was
stirred at the same temperature for 15 h. The mixture was diluted with dichloromethane and the
organic layer was washed with water, brine, dried over MgSO , filtered and evaporated under
4
reduced pressure. The residue was purified by silica gel column chromatography (hexane:ethyl
acetate = 3:1) to give 14a and 14c-14f.
3
(
+)-N -Benzoyl-1-[3,5-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
D
-
ribofuranosyl]uracil (14a).
White foam; Yield: 82%; [α]D +22.64 (c 2.08, CH Cl ); UV (CH Cl ) λ 255 nm; H NMR
2
0
1
2
2
2
2
max
(
400 MHz, CDCl ) δ 8.26 (d, J = 8.0 Hz, 1H), 7.90 (d, J = 1.6 Hz, 2H), 7.66 (t, J = 1.2 Hz, 1H),
3
7
.50 (t, J = 1.6 Hz, 2H), 6.06 (s, 1H), 5.83 (d, J = 8.4 Hz, 1H), 4.41 (d, J = 3.6 Hz, 1H), 4.24 (t,
J = 2.0 Hz, 1H), 4.16-4.02 (m, 2H), 3.87 (d, J = 2.4 Hz, 1H), 2.68 (s, 1H), 0.96-1.15 (m, 28H);
1
3
C NMR (100 MHz, CDCl ) δ 168.5, 162.1, 149.5, 142.5, 135.4, 131.5, 130.7, 129.4, 102.3,
3
8
0.4, 74.4, 59.1, 57.6, 46.6, 17.6, 17.5, 17.4, 17.3, 17.2, 17.1, 13.6, 13.5, 13.3, 12.7; MS (ESI)
+
m/z 759.2037 [M+H] Found: C, 51.23; H, 6.56; N, 4.33. Calc. for C H N O SeSi : C, 51.44;
2
8
42
2
7
2
H, 6.48; N, 4.28%.
3
(
+)-N -Benzoyl-5-fluoro-1-[3,5-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl)-4-seleno-
D
-
ribofuranosyl]uracil (14c).
2
0
1
White foam; Yield: 42%; [α]D +47.17 (c 1.13, CH Cl ); UV (CH Cl ) λ 272 nm; H NMR
2
2
2
2
max