Synthesis and antiviral activity of purine 2',3'-dideoxy-2',3'-difluoro-D- arabinofuranosyl nucleosides

Article abstract of DOI:10.1080/15257770903053979

9-(2',3'-Dideoxy-2',3'-difluoro-D-arabinofuranosyl)adenine (20), 2-chloro-9-(2',3'-dideoxy-2,3-difluoro-D-arabinofuranosyl)adenine (22), as well as their respective -anomers 21 and 23, were synthesized by the nucleobase anion glycosylation of intermediate

Full text of DOI:10.1080/15257770903053979

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Synthesis and Antiviral Activity of
Purine 2′,3′-Dideoxy-2′,3′-Difluoro-D-
Arabinofuranosyl Nucleosides
Grigorii G. Sivets a , Elena N. Kalinichenko a , Igor A. Mikhailopulo a
,
Mervi A. Detorio b , Tami R. McBrayer b , Tony Whitaker b & Raymond
F. Schinazi b
a Institute of Bioorganic Chemistry , National Academy of Sciences ,
Minsk, Belarus
b Center for AIDS Research , Emory University School of Medicine/
Veterans Affairs Medical Center , Decatur, Georgia, USA
Published online: 29 Jul 2009.
To cite this article: Grigorii G. Sivets , Elena N. Kalinichenko , Igor A. Mikhailopulo , Mervi A.
Detorio , Tami R. McBrayer , Tony Whitaker & Raymond F. Schinazi (2009) Synthesis and Antiviral
Activity of Purine 2′,3′-Dideoxy-2′,3′-Difluoro-D-Arabinofuranosyl Nucleosides, Nucleosides,
Nucleotides and Nucleic Acids, 28:5-7, 519-536, DOI: 10.1080/15257770903053979
To link to this article: http://dx.doi.org/10.1080/15257770903053979
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Nucleosides, Nucleotides and Nucleic Acids, 28:519–536, 2009
Copyright ꢀC Taylor & Francis Group, LLC
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770903053979
SYNTHESIS AND ANTIVIRAL ACTIVITY OF PURINE
2ꢀ,3ꢀ-DIDEOXY-2ꢀ,3ꢀ-DIFLUORO-D-ARABINOFURANOSYL
NUCLEOSIDES
Grigorii G. Sivets,1 Elena N. Kalinichenko,1 Igor A. Mikhailopulo,1
Mervi A. Detorio,2 Tami R. McBrayer,2 Tony Whitaker,2 and Raymond
F. Schinazi2
1Institute of Bioorganic Chemistry, National Academy of Sciences, Minsk, Belarus
2Center for AIDS Research, Emory University School of Medicine/Veterans Affairs Medical
Center, Decatur, Georgia, USA
9-(2 ꢁ,3 ꢁ-Dideoxy-2 ꢁ,3 ꢁ-difluoro-β-D-arabinofuranosyl)adenine (20), 2-chloro-9-(2 ꢁ,3 ꢁ-dideoxy-
2
2,3-difluoro-β-D-arabinofuranosyl)adenine (22), as well as their respective α-anomers 21 and 23,
were synthesized by the nucleobase anion glycosylation of intermediate 5-O-benzoyl-2,3-dideoxy-
2,3-difluoro-α-D-arabinofuranosyl bromide (13) starting from methyl 5-O-benzyl-3-deoxy-3-fluoro-
α-D-ribofuranoside (3) and methyl 5-O-benzoyl-α-D-xylofuranoside (10). These compounds were
evaluated as potential inhibitors of HIV-1 and hepatitis C virus in human PBM and Huh-7
Replicon cells, respectively. The adenosine analog 20 demonstrated potent activity against HIV-1 in
primary human lymphocytes with no apparent cytotoxicity. Conformation of pentofuranose ring of
nucleoside 20 in solution was studied by PSEUROT calculations.
Keywords Fluoro-nucleosides; purines; antiviral activity; conformational analysis
INTRODUCTION
Fluorine substitution of a hydrogen atom and/or a hydroxyl group in
a nucleoside analog can impart a significant influence on its biological
Received 6 April 2009; accepted 13 May 2009.
This paper is dedicated to Dr. Morris Robbins, our colleague and friend, and to honor all his
contributions to the field of nucleoside chemistry and biology.
This work was supported in part by NIH grant 2P30-AI-50409 (RFS), 5R37-AI-041980 (RFS)
and Department of Veterans Affairs (RFS) and Belarus State Program of FOI “Physiological Active
Compounds” (Grant 2.04). R.F.S. is the Founder and Director of RFS Pharma, LLC, and received no
compensation for performing this work.
Address correspondence to Raymond F. Schinazi, Center for AIDS Research, VA Medical
Center, Medical Research 151H, 1670 Clairmont Road, Decatur, Georgia, 30033, USA. E-mail:
rschina@emory.edu
519

520
G. G. Sivets et al.
NH2
N
NH2
N
N
N
N
N
N
HO
N
Cl
HO
O
O
F
F
HO
2
1
FIGURE 1 Biologically active purine 2ꢁ-β-fluoro nucleosides.
properties. Addition of a fluorine atom to the sugar moiety of nucleosides
can improve their antiviral properties by enhancing potency and selectivity
of modified nucleosides against HIV or other viruses.[1–4] The structure-
activity relationships for pyrimidine and purine monofluoro 2ꢁ,3ꢁ-dideoxy
nucleosides as potential anti-HIV agents has been previously reported.[1,5,6]
Of the different fluoro-substituted nucleosides, purine 2ꢁ-β-fluoro nucle-
osides attract particular attention in view of their interesting biological
activity, primarily due to the stabilization of the glycosidic bond. For
example, lodenosine 1 is anti-HIV agent which possesses increased chemical
and metabolic stability (Figure 1).[7]
Among dideoxydifluoro nucleosides with β-d-arabino-configuration, a
set of pyrimidine nucleosides have been synthesized and evaluated against
HIV-1.[5] However, synthesis and study of the antiviral properties of purine
nucleosides is of interest for a full biological evaluation of the antiviral
potential for this family of nucleosides. Thus, we have synthesized 9-(2ꢁ,3ꢁ-
dideoxy-2ꢁ,3ꢁ-difluoro-β-d-arabinofuranosyl)adenine (20) and its 2-chloro
derivative 22 which can also be considered as new, closely related nucleoside
analogues of purine 2ꢁ-β-fluoro nucleosides, lodenosine 1 and clofarabine
2,[8] respectively, the latter being used as anticancer agent for treatment of
pediatric acute leukemia (Figure 1).
We describe herein the synthesis of purine 2ꢁ,3ꢁ-dideoxy-2ꢁ,3ꢁ-difluoro
nucleosides with β-d-arabino-configuration 20 and 22, their α-anomers
21 and 23, along with the in vitro anti-HIV and anti-HCV evaluation.
RESULTS AND DISCUSSION
The synthetic route to adenine nucleoside 20 and its α-anomer 21
was briefly reported earlier from methyl 5-O-benzyl-2-deoxy-2-fluoro-α-
d-arabinofuranoside.[9] Herein, we studied alternative pathways for the
preparation of difluoride 9 from the benzyl derivative of methyl 3-fluoro-
3-deoxy-d-ribofuranoside (3) and methyl 5-O-benzoyl-α-d-xylofuranoside
(10) (Scheme 1). Riboside 3, prepared from methyl 5-O-benzyl-3-deoxy-3-
fluoro-α-d-arabinofuranoside,[10] gave the 2-O-Ts derivative 4 under stan-
dard tosylation conditions. The 2-O-SO2Imd derivative 5 was synthesized

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
521
BzO
O
BnO
a or b
c;d
BnO
O
O
OMe
OR
6, R=Ts
OMe
OR
4, R=Ts
OMe
OH
F
F
F
3
7, R=SO2Imd
5, R=SO2Imd
e;f
h
BzO
BzO
O
BzO
O
O
F
+
+
OMe
OMe
OMe
F
OH
11
F
F
8
9
g
BzO
O
OH
OMe
OH
10
SCHEME 1 Reagents and conditions: a) TsCl/Py, room temperature 18 hours, (4, 84%); b)
SO2Cl2/CH2Cl2, −40◦C to room temperature, Imd, 0◦C to room temperature (5, 64%); c)
20% Pd(OH)2/C/cycloxehene, EtOH, reflux, 2–3 hours; d) BzCl/Py, room temperature (c + d,
6 ꢀ70%; 7 ꢀ64%); e) KHF2/45%HF/2,3-butanediol, 160◦C, 45 minutes (9 from 7, 49%); f)
CsF/anh.DMSO/anh.HMPA (11:1,v/v), 190◦C, 1 hour (9, 10%; 8, 26%); g) DAST, CH2Cl2, room
temperature and 27–29◦C, 9, 34%; 11, 19%; h) SO2Cl2/CH2Cl2/DMF, −40◦C to room temperature,
Imd, 0◦C to room temperature (7, 92%).
from 3 using the method proposed by Hanessian for the preparation
of imidazolylsulfonate derivatives of sugars.[11] Debenzylation of 4 and 5
by 20% Pd(OH)2/C in ethanol in the presence of cyclohexene followed
by benzoylation resulted in benzoates 6 and 7. Further, two approaches
involving nucleophilic displacement of the 2-OTs group of the riboside
6 by CsF in DMSO/HMPA[12,13] or 2-O-SO2Imd group of the riboside
7 under treatment with KHF2/HF/2,3-butanediol[14] were tested for the
introduction of a fluorine atom at C-2 position. Difluoride 9 was synthesized
under these conditions from the tosylate 6 and the imidazolylsulfonate
7 in 10 and 49% yields (Scheme 1), respectively, after chromatographic
isolation.
Concomitant with the nucleophilic displacement of the 2-O-tosyl group
in 6 by the fluoride anion, elimination of the tosyl group generated the
the 3-fluorovinyl compound 8 as a by-product isolated in 26% yield after
chromatography. Thus, the investigated four-step route to 9 from riboside
3 via imidazolylsulfonate 7 resulted in the target difluoride in a moderate
20% combined yield.[9]
The most efficient synthetic route to difluoride 9 was from derivative of
methyl α-d-xylofuranoside 10, which is readily available from D-xylose.[15]
The fluorination of 10 was accomplished with excess of DAST in methylene
chloride under 27–29◦C for 20 hours resulted in the target sugar 9 and

522
G. G. Sivets et al.
methyl 5-O-benzoyl-3-deoxy-3-fluoro-α-d-ribofuranoside (11) with 34% and
19% yields, respectively. The synthesis of difluoride 9 from 10 involves
fluorination of 10 at C-3 resulting in the formation of intermediate 3-
fluorodeoxy sugar 11 followed by introduction of a fluorine atom with DAST
at C-2 of 11 under mild heating. It should be noted that the preparation of 9
under these conditions is the first example of relatively effective synthesis of
vicinal substituted 2,3-dideoxy-2,3-difluoro d-pentofuranose via a double flu-
orination of a 5-O-acyl derivative of d-pentofuranose with double inversion
at the C-2 and C-3 atoms resulting from of a single treatment of the starting
sugar by a fluorinating agent. The riboside 11 prepared from xyloside 10
can also be converted into difluoride 9 via imidazolylsulfonate 7. Synthesis
of sulfonate 7 was performed by treatment of 11 with SO2Cl2 and imidazole
in CH2Cl2-DMF in high yield (92%) after column chromatography on silica
gel.
The condensation of difluoride 9 with persilylated N 6-benzoyladenine
in the presence of SnCl4 in acetonitrile afforded a mixture of β- and
α-nucleosides 14 and 15 which was separated into individual anomers
by column chromatography on silica gel.[9] The formation of desired β-
nucleoside 14 was observed in low yield as a result of the glycosylation of
N 6-benzoyladenine by α-methyl glycoside 9. Therefore, another and more
effective approach to β-nucleoside 14 was studied via bromide 13 and exper-
imental details of it are reported in this communication (Scheme 2). Con-
ventional acetolysis of 9 gave a mixture of acetates 12 after chromatography
R1
N
N
N
BzO
g
N
h
O
BzO
R2
f
BzO
O
O
F
OAc
F
9
F
Br
F
F
F
12
13
14 β, R1= NHBz; R2 = H
15 α, R1= NHBz; R2 = H
16 β, R1=R2=Cl
j
NH2
N
N
N
17 α, R1=R2=Cl
18 β, R1= NH2; R2=Cl
19 α, R1= NH2; R2=Cl
i
j
HO
O
+
F
N
R
HO
O
F
N
F
N
N
F
R
N
NH2
20, R= H
22, R= Cl
21, R= H
23, R= Cl
SCHEME 2 Reagents and conditions: f) AcOH/Ac2O/concn. H2SO4 (77%); g) TMS-Br/CDCl3, 2
weeks, room temperature; (13, ≈50%); h) 13/Na-salt of N 6BzAde/THF, reflux, 5 hours (14, 23%; 15,
9%); h) 11/Na-salt of 2,6-di-Cl purine/CH3CN, room temperature (1:1.04), 18 hours (16+17, 48%);
i) saturated NH3/1,2-DME, room temperature (18, 67%; 19, 25%); j) saturated NH3/MeOH, room
temperature (20, 88%, 21, 81%, 22, 75%, 23, 80%).

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
523
on silica gel, bromination (TMSiBr/CDCl3)[16] of the latter generated
intermediate glycosyl bromide 13 that was reacted, without isolation, with
the sodium salt of N 6-benzoyladenine under reflux in tetrahydrofuran to
give a mixture of β- and α-nucleosides 14 and 15. The target β-nucleoside
14 and its α-anomer 15 were isolated by column chromatography on silica
gel in 23% and 9% yield, respectively. An analogous route via coupling
of 1-α-bromo sugar with the nucleobase was used for the preparation of
2-chloro derivative of 18.
The condensation of bromide 13 with the sodium salt of 2,6-
dichloropurine in acetonitrile[17] when compared to the one of N 6-
benzoyladenine gave more complex mixture of products from which a
mixture of protected N9-β/α-nucleosides 16 and 17 (β/α-ratio ≈ 2.9:1
1
according to H NMR data) was isolated in 48% overall yield after column
chromatography on silica gel (Scheme 2). Treatment of the 16/17 mixture
with a saturated solution of ammonia in 1,2-dimethoxyethane[18] at room
temperature for 24 hours afforded protected β- and α-nucleosides of 2-
chloroadenine 18 and 19 which were successively isolated by chromatog-
raphy on silica gel in 67 and 25% yield, respectively.
Standard deprotection of individual blocked nucleosides 14, 15, 18, and
19 with methanolic ammonia and subsequent chromatographic purifica-
tion gave pure 9-(2ꢁ,3ꢁ-dideoxy-2ꢁ,3ꢁ-difluoro-β-d-arabinofuranosyl)adenine
(20) and its α-anomer 21, 2-chloro-9-(2ꢁ,3ꢁ-dideoxy-2ꢁ,3ꢁ-difluoro-β-d-
arabinofuranosyl)adenine (22) and its α-anomer 23, respectively. The struc-
1
ture of nucleosides 20 and 21, 22, and 23 was verified by H, 19F, 13C NMR
and by mass spectroscopy, UV, CD.
The assignments of the configurations of nucleosides 20–23 at the
anomeric centers were based upon NMR data. The diagnostic for the β-
5
anomeric configurations of 20 and 22 is the J H,F long-range coupling of
H-C(8) to a 2ꢁ-β-fluorine atom of 2.3 and 2.27 Hz, respectively, exhibited in
their 1H NMR spectra (Tables 1 and 2). Similarly, the presence of the similar
five-bond coupling of 2.9 and 3.0 Hz (see Experimental) for intermediate
protected nucleosides 14 and 18 supported their β-structural assignment.
This coupling is generally indicative of a spatial proximity of the nuclei
involved and is not observed in the α-anomers 15 and 19, 21, and 23. The
most informative feature of the 1H NMR spectra for the ones in comparison
with the corresponding β-nucleosides 14 and 19, 20, and 22 is the shift of
H-2ꢁ and H-4ꢁ resonances in a lower field for the α-anomers.[19,20]
Of interest are some other observations from the NMR data (Table 2).
1
Long-range coupling of H-1ꢁ to 3ꢁ-fluorine atom of 1.86 and 0.9 Hz in H
NMR spectra of nucleosides 20 and 22 argues for W-shape configuration
of H-1ꢁ and F-3ꢁ, and the β-anomeric configuration of adenine and 2-
chloroadenine nucleosides. A four-bond coupling (0.91 Hz) between H-1
1
and H-3 atoms of difluoride 9 is exhibited in its H NMR spectrum due to
W-arrangement[21] between these protons in the case of α-methyl glycoside

524
G. G. Sivets et al.
TABLE 1 1H NMR chemical shifts of 2,3-dideoxy-2,3-difluoro sugars 9, 12 and nucleosides 20–23 with
d-arabino-configurations. δ in ppm; J in Hz
H-1 or
H-1ꢁ
H-2 or
H-2ꢁ
H-3 or
H-3ꢁ
H-4 or
H-4ꢁ
H-5a or
H-5ꢁa
H-5b or
H-5ꢁb
Cmpd.
Others
9
5.13
br.d
5.08
ddd
5.10
dddt
4.49–4.61
3.46 (s, OMe), 8.08 (m,
1H), 7.60 (m, 2H), 7.48
(m, 2H, Bz)
m
12
6.42
br.d
5.21
dd
5.20
ddd
4.70
ddt
4.57
dd
4.55
dd
2.12 (s, 3H, CH3CO),
8.04 (d, 1H), 7.57 (t,
2H), 7.44 (t, 2H, Bz)
8.30 (d, 1H, J = 2.3,
H-8), 8.21 (s, 1H, H-2)
8.27 (d, 1H, J = 2.27,
H-8), 7.94 (br.s, 2H,
20
22
6.53
ddd
6.42
ddd
5.50
dddd
5.74
5.46
ddd
5.57
dddd
4.30
dm
4.20
dm
3.89
ddd
3.72
br.m
3.85
dd
3.69
br.m
dddd
NH2),5.27 (br.s, 1H,
gem
5ꢁ-OH),
J
∼ 13
ꢁ
ꢁ
5 a, 5 b
21
23
6.53
dd
6.35
dd
6.04
ddt
6.06
ddt
5.47
dddd
5.45
ddt
4.81
ddt
4.63
dm
3.83
dd
3.61
dd
3.79
ddd
3.58
dd
8.40 (s, 1H, H-8) 8.39 (s,
4
ꢁ
ꢁ
1H, H-2) J 5 b, F3 = 1.15
8.31 (s, 1H, H-8), 5.26
(t, 1H, J = 5.66, 5ꢁ-OH),
7.90 (br.s, 2H, NH2)
Spectra were obtained in CDCl3 for sugars 9 and 12, in CD3OD and DMSO-d6 for nucleosides 20, 21
and 22, 23, respectively. Spectral data for pure α-anomer of 12 are presented.
TABLE 2 Coupling constants (in Hz) for 1H NMR data of 2,3-dideoxy-2,3-difluoro sugars 9, 12 and
nucleosides 20–23 with d-arabino-configurations
3J (H,H)
3J (H,F)
4,5a/4,5b H1,F2 H3,F2 H2,F3 H4,F3
1,2 or 2,3 or 3,4 or or 4ꢁ,5ꢁa/
or
or
or
or
1ꢁ,2ꢁ
2ꢁ,3ꢁ
3ꢁ,4ꢁ
4ꢁ,5ꢁb
H1ꢁ,F2 H3ꢁ,F2 H2ꢁ,F3 H4ꢁ,F3
Others
9
12
<0.8 0.91
<1.0 <1.0
4.11
3.24
n.d.
4.5/4.69
10.03
10.57
19.76
17.49
13.44
12.21
22.60 4J 3,1 = 0.91
gem
gem
gem
5
24.14
J
J
J
= 48.17
= 51.0
2,F2
3,F3
= 12.71
H5a,H5b
ꢁ
20
22
3.97
4.76
2.17
3.35
3.76
4.73
4.27/4.85 17.67
15.29
16.58
12.82
14.77
24.80
J F2 ,H8 = 2.3
4
ꢁ
ꢁ
J 1 ,F3 = 1.86
gem
gem
ꢁ
ꢁ
ꢁ
J
J
ꢁ
= 50.52
= 51.24
2 ,F2
ꢁ
3 ,F3
ꢁ
J H5 a,H5 b = 12.8
5
ꢁ
4.78
14.54
22.31
J F2 ,H8 = 2.27
4
ꢁ
ꢁ
J 1 ,F3 = 0.9
gem
gem
gem
gem
gem
gem
gem
gem
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
J
J
J
J
J
J
J
J
= 50.86
= 52.16
= 49.70
= 51.70
ꢁ
2 ,F2
ꢁ
3 ,F3
ꢁ
21
23
2.42
3.24
2.74
3.37
3.90
4.32
5.04/4.98 15.04
6.30/5.26 15.11
15.90
15.39
14.0
21.16
20.44
2 ,F2
ꢁ
3 ,F3
ꢁ
= 12.8
= 2.29
H5 a,H5 b
ꢁ
ꢁ
17.06
H5 a,H5 b
ꢁ
ꢁ
ꢁ
= 52.35
3 ,F3
ꢁ
= 50.28
2 ,F2

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
525
TABLE 3 The 13C NMR data (chemical shifts in ppm)
Chemical shifts, δTMS, ppm [J (C,F)in Hz]
C-1 or
C-1ꢁ
C-2 or
C-2ꢁ
C-3 or
C-3ꢁ
C-4 or
C-4ꢁ
C-5 or
C-5ꢁ
Compd.
Others
9
106.03
dd
96.81 dd
95.04 dd
80.19 d
62.95 d
166.14 (s, Ph-C O, arom),
133.30, 129.78, 129.55,
128.47 (Ph-C O), 54.98 (s,
OCH3)
12
20
22
21
23
98.84 dd
83.07 dd
81.75 dd
87.95 dd
86.58 dd
96.86 dd
94.38 dd
82.76 d
62.81 d
169.34 (s, CH3-C O),
166.21 (s, Ph-C O),
133.54, 129.91, 129.8,
128.59 (Ph-C O), 21.05 (s,
CH3-C O)
93.57 dd
93.93 dd
96.90 dd
97.03 dd
92.69 dd
92.99 dd
93.93 dd
94.46 dd
82.15 dd
81.11 dd
85.13 dd
84.04 dd
60.26 d
60.40 d
60.30 d
60.66 d
156.09 (C-6)
152.83 (C-2)
149.13 (C-4)
5
ꢁ
140.26 (d, J C8,F2 = 4.6,
C-8) 118.49 (C-5)
157.38(C-6)
153.95(C-2)
150.70(C-4)
5
ꢁ
140.69 (d, J C8,F2 = 3.75,
C-8) 117.89 (C-5)
152.93(C-6)
148.68(C-2)
147.94(C-4)
5
ꢁ
141.02 (s, J C8,F2 <1.0,
C-8) 119.37 (C-5)
157.42(C-6) 153.87(C-2)
150.56(C-4) 140.06 (s, C-8)
118.69 (d, J = 5.51, C-5)
9, but it is not found for α-1-O-acetate 12. Interesting stereochemical
1
peculiarities of compounds 20, 22, and 9 is well represented by H NMR
data described above.
13C NMR data presented in Tables 3 and 4 provide further support for
the assignments of the configurations of nucleosides 20–23 at the anomeric
1
centers which were made in terms of H NMR data, long-range coupling
constants between C-8 of heterocyclic base and 2ꢁ-β-fluorine atom (4.6 and
3.75 Hz) that were observed only for β-nucleosides 20 and 22, respectively.
The carbon resonances of the bases in 13C NMR spectra of nucleosides
20–23 are in good accord with 13C NMR spectral data obtained earlier for
closely related purine 2ꢁ-β-fluoro nucleosides.[20] 19F NMR data (see Experi-
mental) of 2,3-dideoxy-2,3-difluoro sugars 9, 12 and nucleosides 20–23 with
D-arabino-configurations are evidence in favour of the assigned structures
of synthesized sugars and nucleosides. F-2ꢁ and F-3ꢁ resonance signals of
sugars and nucleosides with two fluorine atoms in trans-arrangement are

526
G. G. Sivets et al.
TABLE 4 The 13C NMR data (coupling constants in Hz)
F-2
F-3
2J (C,F)
2J (C,F)
3J (C,F)
4J (C,F)
3J (C,F)
3J (C,F)
C1,F2 or C3,F2 or C4,F2 or C5,F2 or C1,F3 or
C2,F3 or C4,F3 or C5,F3 or
Compd.
C1ꢁ,F2
C3ꢁ,F2
C4ꢁ,F2
C5ꢁ,F2
C1ꢁ,F3
C2ꢁ,F3
C4ꢁ,F3
C5ꢁ,F3
9
12
20
22
21
23
35.30
36.87
17.35
17.42
36.40
35.50
30.45
30.85
28.9
27.30
28.02
27.36
<1.0
<1.0
2.84
3.35
2.08
3.40
<1.0
<1.0
<1.0
<1.0
<1.0
<1.0
4.20
1.52
3.55
5.89
4.98
6.42
28.10
29.43
28.70
27.63
28.71
27.11
27.20
28.23
25.51
24.36
24.55
24.30
5.40
6.49
6.39
4.63
5.66
4.68
revealed as complex multiplets in their 19F NMR spectra, but the magnitude
3
of J F-2,F-3 (8.08–8.48 Hz) did not profoundly differ for sugars 9, 12, and
target nucleosides.
Compounds 20–23 were evaluated as potential inhibitors of HIV-1 and
HCV in primary human peripheral blood mononuclear (PBM) and Huh-
7 Replicon cells, respectively. The adenine nucleoside 20 demonstrated
potent activity against HIV-1 in primary human lymphocytes with a median
effective concentration (EC50) of 0.72 µM with no apparent cytotoxicity in
three different cell systems up to 100 µM. The 2-chloro derivative 22 was
moderately active with an EC50 of 14.1 µM, but antiviral activity could be
secondary to its cytotoxicity in human PBM cells (Table 5).
TABLE 5 Anti-HIV and anti-HCV activities of nucleosides 20–23
Anti-HIV-1 activity in human PBM cells and
cytotoxicity in
Anti-HCV activity and toxicity
in replicon assay Huh7 cells
CC50(µM)
b
EC50
EC90
EC50
EC90
CC50
Compound
(µM)
(µM)
PBM
CEM
Vero
(µM)
(µM)
(µM)
20
21
22
23
0.72
5.6
14.1
60.1
4.4
10.3
>100
46.3
>100
—
>100
>100
13.6
9.4
>100
>100
—
>100
>100
70.2
>100
—
>100
>100
>100
>100
—
>10
>10
>10
>10
—
>10
>10
>10
>10
—
>10
>10
>10
>10
—
2ꢁ-β-FddAa
AZTc
0.0026
—
0.010
—
14.3
—
56.0
—
—
1.3
—
5.4
—
>10
2ꢁ-C-Me-Cc
[23]
aThe anti-HIV activity of lodenosine (2ꢁ-β-FddA) 1 in PBM cells was taken from literature
presented for comparison.
and
bToxicity in Huch-7 Replicon cells.
cAZT and 2ꢁ-C-Me C were used as positive controls for the HIV and HCV assays.

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
527
It should be stressed that nucleoside 20 as an analogue of lodenosine
1 displays higher antiviral activity than 2ꢁ-β-FddA[23] with similar cytotoxic
effect in the same cell system (Table 5). None of the compounds were
effective against HCV replication in a Huh-7 based replicon system when
tested at 10 µM. The methodologies for evaluating the antiviral activity
against HIV, HCV and cytotoxicity have been published elsewhere.[24, 25]
Considering that 9-(2ꢁ,3ꢁ-dideoxy-2ꢁ,3ꢁ-difluoro-β-d-arabinofuranosyl)
adenine (20) as new closely related nucleoside analogue of known anti-
HIV agent, lodenosine 1, both exhibit anti-HIV activity in vitro tests, it is
interesting to compare the conformational peculiarities of the both 2ꢁ-β-
fluoro nucleosides possessing antiviral activity, namely, N /S equilibrium of
their pentofuranose rings in solution.
Conformational analysis of the pentofuranose ring of the adenine β-
nucleoside 20 was performed employing the PSEUROT 6.3 program[26] and
the results have been compared with those for closely related deoxyfluoro
nucleoside 1[22] (Table 6). All calculations have been performed aiming
at achieving: (i) the minimal rms and |ꢁJ | values, and (ii) maximal
correspondence of the pseudorotational parameters obtained using only
four vicinal [H,F] couplings and simultaneous analysis of seven [H,H] and
[H,F] couplings.[27,28] Scale factors for the 3J H,H and 3J H,F were 1.0 and 0.2,
respectively.
According to the data of PSEUROT analysis for nucleosides 20 and
1
1 with application of H NMR data prepared in the same deuterated
solvent it can be concluded that pentofuranose rings of the both purine
nucleosides are predominantly in the S-type conformations, the degree of
populations being very similar (76% and 71%). It is noteworthy that the
dominating conformations for N /S equilibrium of the pentofuranose rings
20 and 1 occupy a narrow close segment of pseudorotational wheel for these
nucleosides (Table 6).
Further, comparison of N ↔S pseudorotational equilibrium of
2ꢁ-β-FddA 1 and difluoride 20 possesing anti-HIV activity with the
one reported earlier[13,27,29] for isomeric adenine nucleosides with
2ꢁ-α-fluoro atom and without antiviral activity, 9-(2ꢁ,3ꢁ-dideoxy-2ꢁ-fluoro-
β-d-erythro-pentofuranosyl)adenine (2ꢁ-α-FddA) and 2ꢁ,3ꢁ-dideoxy-2ꢁ,3ꢁ-
difluoro–adenosine, permits us to note that the conformation of
pentofuranose rings of 2ꢁ-β-fluoro anti-HIV nucleosides in solution
differ from the stereochemistry of inactive adenine nucleosides by
pseudorotational parameters.
In conclusion, the preferred conformation of adenine 2ꢁ-β-fluoro nu-
cleosides 20 and 1 appear to play an important role in their anti-HIV
activity. The synthesis of new purine dideoxydifluoro nucleosides with β-d-
arabino-configuration is of interest for the development of novel nucleoside
analogues with potential antiviral activity and for the continuing search

528

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
529
for a stereochemical rationale[30] for the activity of anti-HIV nucleosides
fluorinated on carbohydrate moiety.
EXPERIMENTAL
Column chromatography was performed on silica gel 60 H (70–230
mesh; Merck, Germany). TLC: aluminium-backed silica gel 60 F254 sheets
(Merck, Germany); eluents: hexane/AcOEt 3:1 (A), hexane/AcOEt 1:2
(B), CHCl3/MeOH 4:1 (C). All the anhydrous solvents were distilled over
CaH2, P2O5 or sodium prior to the reaction. The UV and CD spectra were
recorded on Specord M-400 (Carl Zeiss, Germany) and a J-20 (JASCO,
Japan) spectropolarimeter, respectively.1H, 13C, and 19F NMR Spectra were
recorded in CDCl3, CD3OD and (D6)DMSO with Bruker Avance-500-DRX
spectrometer at 500.13, 126.76, and 470.593 MHz, respectively. NMR (δ val-
ues) are in ppm downfield from internal SiMe4 (1H, 13C) or external CFCl3
(19F). J values are reported in Hz. All NMR assignments were confirmed
1
1
by two-dimensional (1H, H and H, 13C) correlation spectroscopy. Mass
spectra were recorded on a chromatomass spectrometer with HPLC-Accela
and LCQ Fleet mass detector (Thermo Electron Corporation, USA).
Methyl 5-O-benzoyl-3-deoxy-3-fluoro-2-O-
[(4-methylphenyl)sulfonyl]-α-D-ribofuranoside (6)
Standard tosylation of 3 (1.49 g, 5.83 mmol) with TsCl in pyridine
followed by column chromatography on silica gel (80 mL), using for elution
mixture EtOAc/hexane 1:3 and 1:1, afforded tosylate 4 (2.0 g, 84%) as a
syrup. Rf 0.31 (A). 1H NMR (CDCl3): 7.78 (d, 2H, CH3C6H4SO2-), 7.22–7.35
(m, 7H, C6H5CH2-, CH3C6H4SO2-), 4.89 (d, 1H, H-1, J 1,2 = 4.8, H-1), 4.80
(ddd, 1H, J 3,2 = 4.8, J 3,4 = 1.2, J 3,F = 60.0, H-3), 4.73 (dm, 1H, J 2,F
=
19.2, H-2), 4.55 (d,1H, PhCH2), 4.45 (d,1H, PhCH2), 4.38 (dm, 1H, H-4,
ꢁ
J 4,F = 25.8), 3.58 (dd, 1H, H-5, J 5,4 = 2.5, J 5,5 = 10.8), 3.53 (dd, 1H,
H-5ꢁ, J 5,4 = 2.9), 3.33 (s, 3H, OCH3), 2.43 (s, 3H, CH3C6H4SO2-). 13C
(CDCl3): 145.4, 137.5, 129.9, 129.3, 128.7, 128.3, 128.1 and 127.8 (s, Bzl
and CH3C6H4SO2-),101.3 (s, C-1), 89.1 (d, J C-3,F = 192.3, C-3), 81.8 (d,
J C-2, F = 23.6, C-2), 75.5 (d, J C-4, F = 14.7, C-4), 73.8 (s, -CH2C6H5), 69.1
(d, J C-5, F = 9.5, C-5), 55.9 (s, OCH3), 21.8 (s, CH3C6H4SO2-). Anal. calc.
for C20H23FO6S: C, 58.52; H, 5.65; Found : C, 58.63; H, 5.58. 19F (CDCl3):
-193.45 (dt, F-3).
To a solution of 4 (1.8 g, 4.38 mmol) in 85 mL anhydrous ethanol,
20% Pd(OH)2/C (3.26 g) and 85 mL cyclohexene was added. The reaction
mixture was refluxed for 150 minutes, catalyst was filtered off and washed
with EtOH (100 mL). The filtrate was evaporated and coevaporated with
toluene (2 × 30 mL). Standard benzoylation of oil product (1.5 g) followed

530
G. G. Sivets et al.
by silica gel (90 mL) column chromatography, using for elution mixture
EtOAc/hexane 1:6, 1:2, and 1:1, afforded benzoyl derivative 6 (1.3 g, 70%)
1
as syrup. Rf 0.32 (A). H NMR (CDCl3): 8.04 (d, 2H, Bz), 7.80 (d, 2H, J =
8.1, CH3C6H4SO2-), 7.63 (m, 1H, Bz), 7.48 (m, 2H, Bz), 7.30 (d, 2H, J = 8.1,
CH3C6H4SO2-),4.94 (d, 1H, H-1, J 1,2 = 4.6), 4.91 (ddd, 1H, J 3,2 = 5.4, J 3,4
= 1.2, J 3,F = 56.4, H-3), 4.76 (ddd, 1H, J 2,F = 22.8, H-2), 4.60 (dm, 1H, H-4,
J 4,F = 25.2), 4.51 (dd, 1H, H-5, J 5,4 = 3.5, J 5,5 = 12.0), 4.43 (dd, 1H, H-5ꢁ,
ꢁ
J 5,4 = 3.6), 3.39 (s, 3H, OCH3), 2.44 (s, 3H, CH3C6H4SO2-). 13C (CDCl3):
165.9 (s, C = O, Bz), 145.6, 133.7, 132.9, 130.0, 129.8, 129.3, 128.8 and 128.2
(s, Bz and CH3C6H4SO2-), 101.2 (s, C-1), 88.4 (d, J C-3, F = 194.6, C-3), 80.3
(d, J C-2, F = 25.0, C-2), 75.2 (d, J C-4, F = 15.0, C-4), 63.5 (d, J C-5, F = 9.1,
C-5), 56.0 (s, OCH3), 21.8 (s, CH3C6H4SO2-). Anal. calc. for C20H21FO7S :
C, 56.59; H, 4.99; Found : C, 56.63; H, 5.08. 19F (CDCl3): -192.97 (ddd, F-3).
Methyl 5-O-benzoyl-3-deoxy-3-fluoro-2-O-(imidazolylsulfonyl)-
α-D-ribofuranoside (7)
Method A. Sulfuryl chloride (0.8 mL, 9.87 mmol) was added dropwise to
a solution of fluoride 3 (1.24 g, 4.84 mmol) in anhydrous CH2Cl2 (15 mL)
at −40◦C. The reaction mixture was stirred at this temperature during 1
hour and then temperature was gradually raised to room temperature for
3 hours. After cooling to 0◦C, imidazole (3.35 g, 49.1 mmol) was added to
prepared solution and the reaction mixture was stirred at room temperature
for 18 hours. The solution was diluted CH2Cl2 (20 mL), washed cold water
(10 mL), the aqueous phase was extracted with CH2Cl2 (3 × 30 mL). The
combined organic extracts were dried over anh. Na2SO4 and evaporated to
dryness. The residue was chromatographed on a silica gel (70 mL), using a
linear gradient of EtOAc (0→50%, v/v; 500 mL) in hexane, to afford syrupy
1
5 (1.2 g, 64%). Rf 0.73 (B). H NMR (CDCl3): 7.16–7.90 (m, 8H, Ar-H),
4.89 (d, 1H, H-1, J 1,2 = 4.6), 4.82 (ddd, 1H, J 3,2 = 5.3, J 3,4 = 1.0, J 3,F
=
59.6, H-3), 4.81 (dt, 1H, J 2,F = 21.3, H-2), 4.57 (d,1H, PhCH2), 4.48 (d,1H,
ꢁ
PhCH2), 4.43 (dm, 1H, H-4, J 4,F = 25.4), 3.63 (dd, 1H, H-5, J 5,4 = 2.4, J 5,5
= 10.7), 3.59 (dd, 1H, H-5ꢁ, J 5,4 = 1.8), 3.37 (s, 3H, OCH3). 13C (CDCl3):
137.5, 137.1, 131.4, 128.8, 128.7, 128.3, 127.9 and 118.4 (s, C6H5CH2- and
C3H3N2SO2-), 100.4 (s, C-1), 88.5 (d, J C-3, F = 194.3, C-3), 81.8 (d, J C-2,F
=
23.5, C-2), 78.4 (d, J C-4,F = 14.5, C-4), 73.9 (s, -CH2C6H5), 69.0 (d, J C-5, F
=
9.5, C-5), 55.8 (s, OCH3). 19F (CDCl3): -193.83 (ddd, F-3). Anal. calc. for
C16H19FN2O6S: C, 49.73; H, 4.96; N, 7.25; Found: C, 49.70; H, 5.02; N, 7.35.
To a solution of imidazolylsulfonate 5 (1.2 g, 3.12 mmol) in 57 mL
anhydrous ethanol, 20% Pd(OH)2/C (2.4 g) and 57 mL cyclohexene was
added and the reaction mixture was refluxed for 150 minutes, catalyst was
filtered off and washed with EtOH (100 mL). The filtrate was evaporated
and coevaporated with toluene (2 × 30 mL). Standard benzoylation of oil
product (1.2 g) followed by silica gel (70 mL) column chromatography

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
531
1
afforded benzoyl derivative 7 (0.75 g, 64%) as syrup. Rf 0.79 (B). H NMR
(CDCl3): 7.10–7.98 (m, 8H, Ar-H), 4.97 (ddd, 1H, J 3,2 = 5.7, J 3,4 = 1.8, J 3,F
=
56.4, H-3), 4.91 (d, 1H, H-1, J 1,2 = 4.6), 4.78 (dt, 1H, J 2,F = 20.0, H-2), 4.62
ꢁ
(dm, 1H, H-4, J 4,F = 24.8), 4.57 (dd, 1H, H-5, J 5,4 = 3.4, J5,5 = 12.2), 4.43
(dd, 1H, H-5ꢁ, J 5,4 = 3.2), 3.38 (s, 3H, OCH3). 13C (CDCl3): 165.8 (s, C =
O, Bz), 137.1, 133.9, 131.5, 129.6, 129.2, 128.9 and 118.3 (s, C6H5CO- and
C3H3N2SO2-), 100.2 (s, C-1), 87.8 (d, J C-3,F = 196.7, C-3), 80.1 (d, J C-2,F
=
24.9, C-2), 78.0 (d, J C-4, F = 14.9, C-4), 63.1 (d, J C-5,F = 8.3, C-5), 56.0 (s,
OCH3). 19F (CDCl3): -193.55 (dt, F-3). Anal. calc. for C16H17FN2O7S : C,
48.00; H, 4.28; N, 6.99; Found : C, 48.09; H, 4.32; N, 7.15.
Method B. Sulfuryl chloride (0.15 ml, 1.85 mmol) was added dropwise
to a solution of fluorosugar 11 (0.17 g, 0.63 mmol) in anhydrous CH2Cl2
(2 mL) and DMF (0.53 ml) at −40◦C. The reaction mixture was stirred
at this temperature during 50 minutes and then for 90 minutes at room
temperature. After cooling to 0◦C, 0.55 g (8.08 mmol) imidazole was
added to prepared solution and the reaction mixture was stirred at room
temperature for 18 hours. After standard work-up and removal of solvent,
the residue was chromatographed on a silica gel (25 mL), using for elution
mixture EtOAc/hexane 1:3, 1:2, and 2:1, to afford syrupy 7 (0.232 g, 92%)
which was identical to imidazolylsulfonate 7 prepared from fluoride 3.
Methyl 5-O-benzoyl-2,3-dideoxy-2,3-difluoro-α-
D-arabinofuranoside (9)
Method A. To a stirred solution of 6 (1.2 g, 2.83 mmol) in anhydrous
DMSO (8 ml) and HMPA (0.7 mL) was added freshly dried cesium
fluoride (2.3 g, 15.1 mmol). The reaction mixture was stirred at 190◦C
for 70 minutes. After cooling to room temperature the reaction mixture
was poured into water (30 ml), and extracted with EtOAc (4 × 90 mL).
The combined organic extracts was dried over anhydrous Na2SO4 and
evaporated to dryness. The residue was chromatographed on a silica gel
(200 mL), using a linear gradient of EtOAc (0→33%, v/v; 600 mL) in
hexane, to afford syrupy difluoride 9 (0.08 g, 10%). Rf 0.74 (A). 19F (CDCl3):
−195.78 (F-2, m), −194.28 (F-3, m, J F-2,F-3 = 8.08). HPLC/APCI-MS, m/z
272 M+, 241 (M-OCH3)+. Anal. calc. for C13H14F2O4: C, 57.35; H, 5.18.
Found: C, 56.90; H, 5.02.
Vinyl fluoride 8 (0.183 g, 26%) as syrup. Rf 0.65 (A). 1H NMR (CDCl3):
8.04 (d, 2H, Bz), 7.59 (m, 1H, Bz), 7.48 (m, 2H, Bz), 5.79 (m, 1H, H-2,
J 2,1 = 1.3, J 2,4 = J 2,F = 4.2), 5.35 (t, 1H, H-1, J 1,F < 1.0, J 1,4 = 1.3), 5.09 (m,
H-4), 4.65 (dd, 1H, H-5, J 5,4 = 2.8, J5,5 = 12.1), 4.41 (dd, 1H, H-5ꢁ, J 5 ,4
=
=
ꢁ
ꢁ
4.0), 3.44 (s, 3H, OCH3). 13C(CDCl3): 166.2 (s, C O, Bz), 161.5 (d, J C-3,F
287.0, C-3), 133.4, 129.9, 129.8, 128.6 (s, C6H5CO-), 106.4 (d, J C-2,F = 15.1,
C-2), 101.8 (d, J C-1,F = 7.2, C-1), 76.6 (d, J C-4,F = 25.4, C-4), 63.4 (d, J C-5,F
=
1.9, C-5), 54.3 (s, OCH3). 19F (CDCl3): −131.6 (m, F-3). HPLC/APCI-MS,

532
G. G. Sivets et al.
m/z 252 M+. Anal. calc. for C13H13FO4: C, 61.90; H, 5.19. Found: C, 62.10;
H, 5.27.
Method B. To a solution of 7 (0.75 g, 1.87 mmol) in freshly distilled 2,3-
butanediol (7.2 mL) was added KHF2 (0.61 g, 7.8 mmol) and suspension
was stirred under argon at 160◦C for several minutes, and then HF (0.36 mL,
46% in H2O) was added to this mixture. The reaction mixture was stirred
at 160◦C during 50 minutes. After the standard work up and removal of
solvent, the residue was chromatographed on a silica gel (100 mL), using for
elution mixture EtOAc/hexane 1:8, 1:6 to afford syrup difluoride 9 (0.25 g,
49%) which was identical to the one described above.
Method C. To a solution of xyloside 10 (0.3 g, 1.12 mmol) in anhydrous
CH2Cl2 (6.5 mL) at room temperature was added under argon (0.82 ml,
6.19 mmol) DAST and the reaction mixture was stirred at this temperature
during 150 minutes and then for 18 hours at 27–29◦C under argon. After
cooling, the reaction mixture was poured into saturated cooled aqueous
NaHCO3 (75 mL). When evolution of gas ceased, it extracted with CH2Cl2
(3 × 30 mL). The combined organic extracts were dried over anh. Na2SO4
and evaporated to dryness. The residue was chromatographed on silica gel
(90 mL), using for a linear gradient of hexane/EtOAc/2:1 (v/v; 500 mL) in
hexane/EtOAc/8:1, to afford difluoride 9 (0.103 g, 34%) and fluoride 11
(0.058 g, 19%) which was identical to the one prepared earlier.[15] 19F for 11
(CDCl3): -195.58 (dt, F-3).
1-O-Acetyl-5-O-benzoyl-2,3-dideoxy-2,3-difluoro-α/β-
D-arabinofuranoside (12)
Concentrated H2SO4 (0.1 mL) was added to a solution of fluoride
9 (0.337 g, 1.24 mmol) in acetic acid (2.33 mL) and acetic anhydride
(0.59 mL) at 0◦C. The reaction mixture was stirred at this temperature
during 30 minutes and left at +4◦C for 18 hours, then it was poured into
mixture ice/water. After ice melted, the aqueous phase was extracted with
CH2Cl2 (3 × 40 mL). The combined organic extracts was washed aqueous
NaHCO3, dried over anh. Na2SO4 and evaporated to dryness. The residue
was chromatographed on a silica gel (55 mL), using for elution mixture
EtOAc/hexane 1:3 and 1:1 to afford 12 (0.286 g, 77%) as syrup. Rf 0.48 (A).
1H NMR (CDCl3): (α, β ratio ca. 3:1), 7.43–8.08 (m, ArH), 6.42 (d, H-1 α,
J 1,F-2 = 10.57), 6.39 (d, H-1 β, J 1,2 = 4.7), 5.29–5.49 (m, H-2 β and H-3 β),
5.21 (dd, H-2 α), 5.20 (ddd, H-3 α), 4.70 (ddt, H-4 α), 4.40–4.60 (m, H-5 α,
β and H-5ꢁ α, β, H-4 β), 2.12 (s, OAc α), 2.00 (s, OAc β). HPLC/APCI-MS,
m/z 241 (M-CH3COO)+. 19F (CDCl3, for α-anomer): −195.3 (F-2, m),
−192.9 (F-3, m, J F-2,F-3 = 8.28). Anal. calc. for C14H14F2O5: C, 56.00; H, 4.70;
Found : C, 56.11; H, 4.65.

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
533
9-(2,3-dideoxy-2,3-difluoro-β-D-arabinofuranosyl)adenine (20)
and its α-anomer (21)
TMSiBr (0.2 ml, 1.5 mmol) was added to a solution of 12 (0.19 g,
0.64 mmol) in CDCl3 (1.4 mL) at 0◦C and the mixture was stirred at this
temperature for 30 minutes. After standing at room temperature for 14 days,
the reaction mixture was evaporated, and coevaporated with anhydrous
toluene (3 × 4 mL) and 13 (content ≈50% according to TLC data and
1H NMR data, 6.55 ppm, d, J 1,F = 12.6, H-1 and 4.85 ppm, ddt, H-4 for
1-α-bromo sugar 13) was used in the next step without purification.
The solution of 13 in anhydrous THF (10 mL) was added to sodium
salt of N 6-benzoyladenine, prepared from 0.098 g (0.40 mmol) N 6-
benzoyladenine and NaH in oil (14 mg of 80% in oil, 0.46 mmol). The
reaction mixture was refluxed for 5 hours, filtered off, and washed with
CH2Cl2. After evaporation of combined filtrates, the residue was chro-
matographed on silica gel (120 mL), using a linear gradient of EtOAc
(0→66%, v/v; 500 mL) in hexane, to afford syrupy nucleoside 15 (14 mg,
1
9%). Rf 0.44 (B). H NMR (CDCl3): 9.09(s, 1H, NH), 8.84 (s, 1H, H-2),
ꢁ
ꢁ
8.15 (s, 1H, H-8), 7.48–8.08 (5m, 10H, 2Bz), 6.55 (dd, 1H, J 1 ,2 = 1.1,
J 1 ,F-2 = 15.6, H-1ꢁ), 6.03 (ddm, 1H, H-2ꢁ), 5.46 (ddm, 1H, H-3ꢁ), 5.11 (dm,
1H, H-4ꢁ), 4.64 (dd, 1H, H-5ꢁ), 4.60 (dd, 1H, H-5ꢁꢁ). HPLC/APCI-MS, m/z
479 M+.
ꢁ
ꢁ
1
14 (36 mg, 23%) as syrup. Rf 0.33 (B) H NMR (CDCl3): 9.16 (s, 1H,
ꢁ
NH), 8.14 (s, 1H, H-2), 8.25 (d, 1H, J F2 ,H8 = 2.9, H-8), 7.47–8.08 (5m, 10H,
2Bz), 6.68 (dt, 1H, J 1 ,F-2 = 22.3, J 1 ,2 = J 1 ,F-3 = 2.67, H-1ꢁ), 5.47 (dm, 1H, H-
2ꢁ), 5.38 (ddd, 1H, H-3ꢁ), 4.62–4.75 (m, 3H, H-4ꢁ, 2H-5ꢁ). HPLC/APCI-MS,
m/z 479 M+.
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
Deprotection of 14 and 15 with methanol saturated at 0◦C by am-
monia and subsequent chromatographic purification on silica gel using
for elution mixture CHCl3:MeOH-20:1 and 6:1 afforded adenine nucleo-
sides 20 (18 mg, 88%) and 21 (6.4 mg, 81%), respectively. Compound
20. Rf 0.64 (C). m.p. 166–169◦C (from EtOH/ether); UV (EtOH) λmax
,
nm (ε): 259 (14000), λmin, nm (ε): 227 (2000). CD (EtOH), λ, nm
([θ]·10−3): 205 (+6.4), 214(0), 218 (-2.2), 226 (0), 248 (+1.3), 252 (0),
263 (−1.7), 288 (0). HPLC/APCI-MS, m/z 271 M+. 19F (CD3OD):−204.6
(m, F-C2ꢁ or F-C3ꢁ, J F-2 ,F-3 is not determined), −195.8 (m, F-C2ꢁ or F-
C3ꢁ). Anal. calc. for C10H11F2N5O2: C, 44.28; H, 4.09; Found: C, 44.19; H,
4.00.
ꢁ
ꢁ
Compound 21. Rf 0.64 (C). m.p. 96–100◦C (from EtOH/ether);
UV (EtOH) λmax, nm (ε): 259 (14300), λmin, nm (ε): 227 (2100). CD
(EtOH)), λ, nm ([θ]·10−3): 205 (+6.2), 214(0), 217 (-4.7), 220 (0), 230
(+1.9), 255 (+0.8), 260 and 288 (0). HPLC/APCI-MS, m/z 271 M+. 19F
(CD3OD):−187.29 (m, F-C2ꢁ or F-C3ꢁ), −183.08 (m, F-C2ꢁ or F-C3ꢁ, J F-2 ,F-3
ꢁ
ꢁ
is not determined). Anal. calc. for C10H11F2N5O2 : C, 44.28; H, 4.09; Found:
C, 44.35; H, 4.17.

534
G. G. Sivets et al.
2-Chloro-9-(2,3-dideoxy-2,3-difluoro-β-D-arabinofuranosyl)
adenine (22) and its α-anomer (23)
A suspension of 2,6-dichloropurine (0.064 g, 0.332 mmol) in anhydrous
acetonitrile (3.0 mL) at room temperature was treated NaH (11.0 mg of
80% in oil, 0.034 mmol), and the mixture was stirred for 30 minutes under
argon. To this stirred suspension, a solution bromide 13 prepared from 12
(0.19 g, 0.64 mmol) as described above was added in anhydrous acetonitrile
(4 mL) and the reaction mixture was stirred at room temperature overnight.
Insoluble material was removed by filtration and washed with acetonitrile (5
mL). The combined filtrate and washings were evaporated and the residue
was chromatographed on silica gel (130 mL) using for elution mixture
EtOAc/hexane 1:3 and 2:3 to afford mixture nucleosides 16 and 17 (0.066
g, 48%) as syrup. Rf 0.48 (A). 1H NMR (CDCl3): 8.32 (d, H-8β, J F2 ,H8 = 2.9,
ꢁ
ꢁ
ꢁ
2.9H), 8.26 (s, H-8α, 1H), 7.44–8.06 (m, 4Bz), 6.59 (dt, 1H, J 1 ,F-2 = 21.97,
J 1 ,2 = J 1 ,F-3 = 2.6, H-1ꢁ β), 6.56 (br. d, 1H’α, J 1 ,2 < 1.0, J 1 ,F-2 = 19.1,
H-1ꢁ), 5.75–5.93 (m, H-2ꢁα and H-3ꢁα), 5.46 (dd, H-2ꢁβ), 5.36(ddd, H-3ꢁβ),
5.09 (dm, H-4ꢁα), 4.52–4.75 (m, 2H-5ꢁα, 2H-5ꢁβ and H-4ꢁβ). (UV (EtOH)
λmax, nm (ε): 274 (5660), 231 (7300).
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
ꢁ
A solution of mixture nucleosides 16 and 17 (0.066 g, 0.154 mmol)
in anhydrous 1,2-dimethoxyethane (10 mL) was saturated by dry ammonia
for 4 hours and then left for 18 hours at room temperature. The reaction
mixture was filtered off and washed with CH2Cl2. The combined filtrate and
washings were evaporated and the residue was chromatographed on silica
gel (130 mL), using a linear gradient of EtOAc (0→66%, v/v; 500 mL) in
hexane, to afford nucleoside 18 as foam (42 mg, 67%). Rf 0.60 (B). 1H NMR
ꢁ
(CDCl3): 7.45–8.06 (3m, 5H, Bz), 7.99 (d, 1H, J
= 3.0, H-8), 6.52 (dt,
F2 ,H8
1H, J 1 ,F2 = 12.5, H-1ꢁ), 6.32 (br. s, 2H, NH2), 5.41 (dd, H-2ꢁ), 5.31 (ddd,
H-3ꢁ), 4.54–4.70 (m, 2H-5ꢁand H-4ꢁ). HPLC/APCI-MS, m/z, 410 and 412,
Cl35/Cl37 ratio ≈ 3:1, M+. Nucleoside 19 (16 mg, 25%) as foam. Rf 0.46 (B).
1H NMR (CDCl3): 7.47–8.10 (3m, 5H, Bz), 7.97 (s, 1H, H-8), 6.48 (d, 1H,
ꢁ
ꢁ
J 1 ,F2 = 14.9, H-1ꢁ), 6.18 (br. s, 2H, NH2), 5.88 (dm, 1H, H-2ꢁ), 5.44 (dm,
1H, H-3ꢁ), 5.07 (dm, 1H, H-4ꢁ), 4.63 (dd, 1H, H-5ꢁ), 4.58 (dd, 1H, H-5ꢁꢁ).
HPLC/APCI-MS, m/z 410 and 412, Cl35/Cl37 ratio ≈ 3:1, M+.
ꢁ
ꢁ
Solution of nucleoside 18 (36 mg, 0.088 mmol) in 10 mL methanol satu-
rated at 0◦C by ammonia was kept during 195 minutes at room temperature
and then was evaporated. The residue was chromatographed on silica gel
(40 mL) using for elution mixture CHCl3:MeOH-20:1 and 15:1 to afford
nucleoside 22 (20 mg, 75%). Rf 0.75 (c). m.p. 171–173◦C (CHCl3/EtOH).
UV λmax, nm (ε): 263 (14900) at pH 7, 263 (15000) at pH 13, 263 (13900) at
pH 1. CD (EtOH), λ, nm ([θ]·10−3): 207 (+12.9), 212(0), 217 (−4.0), 231
(0), 260 (−1.7), 288 (0). HPLC/APCI-MS, m/z 307 and 309, Cl35/Cl37 ratio
≈ 3:1, (M+H)+. 19F (DMSO-d6): −202.47 (m, F-C2ꢁ), −196.40 (m, F-C3ꢁ,
ꢁ
ꢁ
J F-2 ,F-3 = 8.48). Anal. calc. for C10H10F2ClN5O2: C, 39.29; H, 3.30; Cl, 11.60;
Found : C, 39.40; H, 3.47; Cl, 11.72.

Purine 2ꢁ,3ꢁ-Dideoxy-2ꢁ,3ꢁ-Difluoro-D-Arabinofuranosyl Nucleosides
535
In a similar way, nucleoside 23 (6 mg, 80%) was prepared starting
from nucleoside 19 (10 mg, 0.024 mmol). Rf 0.75 (C). m.p. 176–178◦C
(CHCl3/EtOH). UV λmax, nm (ε): 263 (14500) at pH 7, 263 (14700) at
pH 13, 263 (13900) at pH 1. CD (EtOH), λ, nm ([θ]·10−3): 205 (+11.6),
212(0), 217 (−8.5), 228 (0), 255 (+1.1), 269 (0). HPLC/APCI-MS, m/z
307 and 309, Cl35/Cl37 ratio ≈ 3:1, (M+H)+. 19F (DMSO-d6): −195.21 (m,
F-C2ꢁ, J F-2 ,F-3 = 8.48), −195.76 (m, F-C3ꢁ). Anal. calc. for C10H10F2ClN5O2:
ꢁ
ꢁ
C, 39.29; H, 3.30; Cl, 11.60; Found : C, 39.38; H, 3.20; Cl, 11.69.
REFERENCES
1. Ray, A.S.; Schinazi, R.F.; Murakami, E.; Basavapathruni, A.; Shi, J.; et al. Probing the mechanistic
consequences of 5-fluorine substitution on cytidine nucleotide analogue incorporation by HIV-1
reverse transcriptase. Antiv. Chem. Chemother. 2003, 14, 115–125.
2. Lee, K.; Choi, Y.; Gumina, G.; Zhou, W.; Schinazi, R.F.; Chu, C.K. Structure-Activity Relationships of
2ꢁ-fluoro-2ꢁ,3ꢁ-unsaturated d-nucleosides as anti-HIV-1 agents. J. Med. Chem. 2002, 45, 1313–1320.
3. Clarck, J.L.; Hollecker, L.; Mason, J.C.; Stuyver, L.J.; Tharnish, P.M.; et al. Design, synthesis, and
antiviral activity of 2ꢁ-deoxy-2ꢁ-fluoro-2ꢁ-C-methylcytidine, a potent inhibitor of hepatitis C virus
replication. J. Med. Chem. 2005, 48, 5504–5508.
4. Shi, J.; Du, J.; Ma, T.; Pankiewicz, K.W.; Patterson, S.E.; et al. Synthesis and antiviral activity of a series
of D- and L-2ꢁ-deoxy-2ꢁ-fluororibonucleosides in the subgenomic HCV replicon system. Bioorg. Med.
Chem. 2005, 13, 1641–1652.
5. Martin, J.A.; Busnell, D.J.; Duncan, I.B.; Dunsdon, S.J.; Hall, M.J.; et al. Synthesis and antiviral
activity of monofluoro and difluoro analogues of pyrimidine deoxyribonucleosides against human
immunodeficiency virus (HIV-1). J. Med. Chem. 1990, 33, 2137–2145.
6. Herdewijn, P.; Pauwels, R.; Baba, M.; Balzarini, J.; De Clercq, E. Synthesis and anti-HIV activity of
various 2ꢁ- and 3ꢁ-substituted 2ꢁ,3ꢁ-dideoxyadenosines: A structure-activity analysis. J. Med. Chem. 1987,
30, 2131–2137.
7. Marquez, V.E.; Tseng, C.K.-H.; Kelly, J.A.; Mitsuya, H.; Broder, S.; et al. 2ꢁ,3ꢁ-Dideoxy-2ꢁ-fluoro-ara-
A. An acid-stable purine nucleoside active against human immunodeficiency virus (HIV). Biochem.
Pharmacol. 1987, 36, 2719–2722.
8. Bonate, P.L.; Arthaud, L.; Cantrell, W.R.; Stephenson, K.; Secrist J.A. III; Weitman, S. Discovery
and development of clofarabine: a nucleoside analogue for treating cancer. Nature Reviews 2006,
5, 855–863.
9. Sivets, G.G.; Kalinichenko, E.N.; Mikhailopulo, I.A. Synthesis of 9-(2,3-dideoxy-2,3-difluoro-β-d-
arabinofuranosyl)adenine. Nucleosides Nucleotides Nucleic Acids. 2007, 26, 1387–1389.
10. Mikhailopulo, I.A.; Sivets, G.G.; Poopeiko, N.E.; Khripach, N.B. Oxidation-reduction sequence for
the synthesis of peracylated fluorodeoxy pentofuranosides. Carbohyd. Res. 1995, 278, 71–89.
11. Hanessian, S.; Vatele, J.-M. Design and reactivity of organic functional groups: imidazolylsulfonate
(imidazylate)—an efficient and versatile leaving group. Tetrahedron Lett. 1981, 22, 3579–3582.
12. Su, T.-L.; Klein, R.S.; Fox, J.J. Improved synthesis of α-d-ribofuranosides via stereoselective alkylation
of a dibutylstannylene derivative for ready access to the 2-substituted 2-deoxyarabinofuranosides. J.
Org. Chem. 1981, 47, 1506–1509.
13. Sivets, G.G.; Kalinichenko, E.N.; Mikhailopulo, I.A. Synthesis and conformational analysis of 1ꢁ-
and 3ꢁ-substituted 2-deoxy-2-fluoro-d-ribofuranosyl nucleosides. Helv. Chim. Acta. 2007, 90, 1818–
1836.
14. Tann, C.H.; Brodfuehrer, P.R.; Brundidge, S.P.; Sapino, C. Jr.; Howell, H.G. An efficient synthe-
sis of 1-(2-deoxy-2-fluoro-β-d-arabinofuranosyl)-5-iodouracil (β-FIAU) and 1-(2-deoxy-2-fluoro-β-d-
arabinofuranosyl) thymine (β-FMAU). J. Org. Chem. 1985, 50, 3644–3647.
15. Mikahilopulo, I.A.; Sivets, G.G. A Novel route for the synthesis of deoxy fluoro sugars and
nucleosides. Helv. Chim. Acta. 1999, 82, 2052–2065.
16. Gillard, J.W.; Israel, M. Trimethylsilyl bromide as a mild, stereoselective anomeric brominating agent.
Tetrahedron Lett. 1981, 22, 513–516.

536
G. G. Sivets et al.
17. Kazimierczuk, Z.; Cottam, H.B.; Revankar, G.R.; Robins, R.K. Synthesis of 2ꢁ-Deoxytubercidin,
2ꢁ-deoxyadenosine, and related 2ꢁ-deoxynucleosides via a novel direct stereospecific sodium salt
glycosylation procedure. J. Am. Chem. Soc. 1984, 106, 6379–6382.
18. Zaitseva, G.V.; Sivets, G.G.; Kazimierczuk, Z.; Vilpo, J.A.; Mikahilopulo, I.A. Convergent syntheses
and cytostatic properties of 2-chloro-2ꢁ-deoxy-2ꢁ-fluoroadenosine and its N7-isomer. Bioorg. Med.
Chem. Lett. 1995, 5, 2999–3002.
19. Mikhailopulo, I.A.; Poopeiko, N.E.; Pricota, T.I.; Sivets, G.G.; Kvasyuk, E.I.; Balzarini, J.; De Clercq,
E. Synthesis and antiviral and cytostatic properties of 3ꢁ-deoxy-3ꢁ-fluoro- and 2ꢁ-azido-3ꢁ-fluoro-2ꢁ,3ꢁ-
dideoxy-d-ribofuranisides of natural heterocyclic bases. J. Med. Chem. 1991, 34, 2195–2202.
20. Tennila¨, T.; Azhayeva, E.; Vepsa¨la¨inen, J.; Laatikainen, R.; Azhayev, A.; Mikhailopulo, I.A. Oligonu-
cleotides containing 9-(2-deoxy-2-fluoro-β-d-arabinofuranosyl)-adenine and -guanine: synthesis, hy-
bridization and antisense properties. Nucleosides Nucleotides Nucleic Acids. 2000, 19, 1861–1884.
21. Michalik, M.; Hein, M.; Frank, M. NMR spectra of fluorinated carbohydrates. Carbohyd. Res. 2000,
327, 185–218.
22. Sivets, G.G.; Kalinichenko, E.N.; Mikahilopulo, I.A. Synthesis of C2ꢁ-β-fluoro-substituted adenine
nucleosides via pivaloyl derivatives of adenosine and 3ꢁ-deoxyadenosine. Lett. Org. Chem. 2006, 3,
402–408.
23. De Clercq, E. HIV inhibitors targeted at the reverse transcriptases. AIDS Res. Human Retroviruses
1992, 8, 119–134.
24. Schinazi, R.F.; Sommadossi, J.P.; Saalmann, V.; Cannon, D.L.; Xie, M.-W.; et al. Activities of 3ꢁ-azido-3ꢁ-
deoxythymidine nucleotide dimers in primary lymphocytes infected with human immunodeficiency
virus type 1. Antimicrob. Agents Chemother. 1990, 34, 1061–1067.
25. Stuyver, L.J.; Lostia, S.; Adams, M.; Mathew, J.; Pai, B.S.; et al. Antiviral activities and cellular toxicities
of modified 2ꢁ,3ꢁ-dideoxy-2ꢁ,3ꢁ-didehydrocytidine analogues. Antimicrob. Agents Chemother. 2002, 46,
3854–3860.
26. Van Wijk, J.; Haasnot, C.A.G.; de Leeuw, F.A.A.M.; Huckriede, B.D.; Westra Hoekzema, A.J.A.;
Altona, C. PSEUROT 6.3. A program for the conformational analysis of five-membered rings, Leiden
Institute of Chemistry: Leiden, the Netherlands, 1999.
27. Mikahilopulo, I.A.; Pricota, T.I.; Sivets, G.G.; Altona, C. 2ꢁ-Chloro-2ꢁ,3ꢁ-dideoxy-3ꢁ-fluoro-d-
ribofuranosides: synthesis, stereospecificity, some chemical transformations, and conformational
analysis. J. Org. Chem. 2003, 68, 5897–5908.
28. Thibaudeau, C.; Plavec, J.; Chattopadhyaya, J. A new generalized Karplus-type equation relating
vicinal proton-fluorine coupling constants to H-C-C-F torsion angles. J. Org. Chem. 1998, 63,
4967–4984.
29. Ford, H.; Dai, F.; Mu, L.; Siddiqui, M.A.; Nicklaus, M.C.; et al. Adenosine deaminase prefers a distinct
sugar ring conformation for binding and catalysis: kinetic and structural studies. Biochemistry 2000,
39, 2581–2592.
30. Barchi, J.J.; Karki, R.G.; Nicklaus, M.C.; Siddiqui, M.A.; Clifford, G.; et al. Comprehensive structural
studies of 2ꢁ,3ꢁ-difluorinated nucleosides: comparison of theory, solution and solid state. J. Am. Chem.
Soc. 2008, 130, 8048–8057.