Synthesis and antiviral evaluation of novel conformationally locked nucleosides and masked 5′-phosphate derivatives thereof

Article abstract of DOI:10.1039/b203484k

As part of a programme towards evaluating the potential of conformationally locked 3′-deoxy- and 3′-azido-3′-deoxy-nucleoside derivatives as prodrugs of potential 5′-O-triphosphorylated anti-HIV drugs, novel nucleoside derivatives with locked N-type (nort

Full text of DOI:10.1039/b203484k

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Synthesis and antiviral evaluation of novel conformationally locked
nucleosides and masked 5Ј-phosphate derivatives thereof
Torsten Bryld,a Marianne H. Sørensen,a Poul Nielsen,a Troels Koch,b Claus Nielsenc and
Jesper Wengel*a
1
a Nucleic Acid Center†, Department of Chemistry, University of Southern Denmark,
DK-5230 Odense M, Denmark
b Cureon A/S, Fruebjergvej 3, DK-2100 Copenhagen, Denmark
c Retrovirus Laboratory, Department of Virology, Statens Serum Institut,
DK-2300 Copenhagen, Denmark
Received (in Cambridge, UK) 9th April 2002, Accepted 31st May 2002
First published as an Advance Article on the web 19th June 2002
As part of a programme towards evaluating the potential of conformationally locked 3Ј-deoxy- and 3Ј-azido-3Ј-
deoxy-nucleoside derivatives as prodrugs of potential 5Ј-O-triphosphorylated anti-HIV drugs, novel nucleoside
derivatives with locked N-type (north-type, C3Ј-endo) furanose conformation were prepared using convergent
synthetic strategies. In addition, masked 5Ј-monophosphate derivatives of these, and of a conformationally restricted
3Ј-azido-3Ј-deoxynucleoside with E-type (eastern-type, O4Ј-endo) furanose conformation, were prepared in order to
potentially circumvent the first phosphorylation step. However, neither the free 5Ј-hydroxy derivatives nor the masked
5Ј-monophosphates showed anti-HIV activity in MT-4 cells.
Molecular modelling studies based on the crystal structure of
Introduction
HIV-1 RT complexed with AZT2 and studies of conformation-
In the treatment of HIV infection, 2Ј,3Ј-dideoxynucleosides
such as 2Ј,3Ј-dideoxycytidine (ddC), 2Ј,3Ј-dideoxyinosine (ddI),
2Ј,3Ј-didehydro-3Ј-deoxythymidine (d4T) and 3Ј-azido-3Ј-
ally restricted NTP analogues (Fig. 1; derivatives 1 and 2) have
shown that the polymerase domain of HIV-1 RT preferentially
binds NTPs adopting an N-type (north-type; C3Ј-endo) furan-
deoxythymidine (AZT) (Fig. 1) are important drugs used in
ose conformation (Fig. 2).3,4 Analogues of AZT and other
Fig. 2 Conformational equilibrium of a 2Ј,3Ј-dideoxynucleoside, the
locked N-type furanose conformation of nucleoside analogues 3–6, and
the preferred E-type furanose conformation of AZT analogue 7. For a
detailed description of furanose conformations and the pseudo-
rotational cycle the reader is referred to, e.g., refs 3, 4, 7 and 8.
Fig. 1 Structures of AZT and conformationally locked bicyclic
nucleoside analogues 1–8. T = thymin-1-yl. A = adenin-9-yl.
either monotherapy or in combination therapy.1 These nucleo-
sides are prodrugs of the active nucleoside triphosphates
(NTPs) and therefore need to be triphosphorylated in vivo
before interacting with the HIV-encoded enzyme HIV-1 reverse
transciptase (HIV-1 RT). Unfortunately, their use is correlated
with toxic side effects and the development of drug-resistant
viruses. Therefore, there is a need for improved new anti-HIV
drugs and a deeper understanding of the pharmacological
effects of the different nucleoside analogues.
anti-HIV-active nucleosides with a locked N-type furanose
conformation are therefore promising drug candidates to be
studied. Marquez et al. showed that the bicyclic nucleoside 1
with a fixed N-type furanose conformation was devoid of activ-
ity whereas the corresponding 5Ј-triphosphorylated derivative
showed activity similar to that of AZT (as an inhibitor of
HIV-1 RT).3 In contrast, the corresponding AZT analogue
2
with a locked S-type (south-type; C3Ј-exo) furanose
conformation showed no activity either as the free nucleoside
2 or as the corresponding 5Ј-O-triphosphorylated derivative.3
It was therefore suggested that the lack of activity of analogue
1 could be caused by the inability of kinases to convert
† A research center for studies of nucleic acid chemical biology funded
by The Danish National Research Foundation
DOI: 10.1039/b203484k
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This journal is © The Royal Society of Chemistry 2002
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Scheme 1 Reagents and conditions (and yields): i) 80% AcOH; ii) a) NaIO4, H2O, THF, b) CH2O, NaOH, 1,4-dioxane; iii) MsCl, pyridine;
iv) a) 80% TFA, b) Ac2O, pyridine; v) a) thymine, N,O-bis(trimethylsilyl)acetamide, TMS triflate, acetonitrile (23% from 9) or b) adenine, SnCl4,
acetonitrile; vi) saturated methanolic ammonia; vii) NaH, DMF [B = thymin-1-yl (64%, 2 steps); B = adenin-9-yl]; viii) KOAc, 18-crown-6,
1,4-dioxane [B = thymin-1-yl (66%), B = adenin-9-yl]; ix) saturated methanolic ammonia [B = thymin-1-yl (65%), B = adenin-9-yl (3% from 9)].
nucleosides with locked N-type conformation into the corre-
sponding 5Ј-O-triphosphorylated derivatives3 which requires
three sequential enzymatic steps involving different cellular
kinases.5 Accordingly, further examples of non-phosphorylated
free nucleoside analogues of AZT with a locked or restricted
furanose conformation (Fig. 1; derivatives 3 and 7) have been
shown to be devoid of anti-HIV activity.6,7
cleavage of the generated diol, aldol reaction and Cannizzaro
reaction,18,19 and finally mesylation of the two primary hydroxy
groups. The key intermediate 13 was obtained by cleavage of
the isopropylidene group using 80% trifluoroacetic acid (TFA)
followed by acetylation with acetic anhydride in pyridine.
Coupling between furanose 13 and thymine by standard
Vorbrüggen-type reaction20 using trimethylsilyl triflate as Lewis
acid and N,O-bis(trimethylsilyl)acetamide as silylating agent
provided stereoselectively the thymine nucleoside 14 in 23%
yield (from 9). The acetyl group of 14 was removed by treat-
ment with a mixture of saturated methanolic ammonia and
methanol (1 : 2, v/v) to give nucleoside 15, which was converted
into the bicyclic derivative 16 using NaH (64% yield from 14).
The remaining methylsulfonyloxy group was replaced by an
acetate group by treatment of nucleoside 16 with a mixture of
potassium acetate and 18-crown-6 in 1,4-dioxane under reflux,
affording nucleoside 17 in 66% yield. Deacetylation by treat-
ment with a mixture of saturated methanolic ammonia and
methanol (2 : 1, v/v) yielded the target bicyclic nucleoside 5 in
65% yield. The applicability of di-O-(methylsulfonyl)furanose
intermediates for the synthesis of locked nucleic acid (LNA)-
type nucleosides has been demonstrated earlier.21,22
Coupling of furanose 13 with adenine using SnCl4 in
acetonitrile afforded nucleoside 18. As expected from the
presence of the 2-acetoxy group in 13, the Vorbrüggen-type
coupling reaction also, in this case, proceeded stereoselectively
with no indication of the formation of the anomeric nucleo-
side. It was possible to assign the obtained product as the
N 9-regioisomer from the 13C NMR data [δC 155.55 (C-6),
152.95 (C-2), 148.86 (C-4), 139.7 (C-8) and 120.04 (C-5)]
and comparison of these with literature data.23 The target
adenine 3Ј-deoxynucleoside 6 was subsequently obtained in
3% yield (9 steps from 9) via derivatives 19–21 using similar
procedures as described above for the corresponding thymine
3Ј-deoxynucleoside 5.
To further evaluate the relation between furanose conform-
ation and anti-HIV activity, we describe herein synthesis of
analogues of known anti-HIV active nucleosides with the
furanose moieties locked in the N-type (Fig. 1; derivatives 3,6,9
4, 5, 6 and 8) or in the E-type (east-type; O4Ј-endo) (Fig. 1;
derivative 77) furanose conformation. The 3Ј-deoxy--nucleo-
side analogue 8 (Fig. 1), also with locked N-type furanose con-
formation, was included in this study as a 3Ј-hydroxy derivative
of 8 has been shown to structurally mimic the corresponding
-ribo-configured nucleoside.10 In addition, we have synthesized
masked 5Ј-monophosphate derivatives of nucleosides 3, 4, 5
and 7 in order to study if the first phosphorylation step is the
major obstacle for in vivo activity of these derivatives. The
masked monophosphates prepared are of the cyclosal-type
introduced Meier et al.11,12 and of the aryloxy phosphor-
amidate-type introduced by McGuigan et al.,13–16 both of which
have been successfully applied to other nucleosides being
inactive unless masked.11–16 We decided to prepare these novel
compounds despite the results of an interesting molecular
dynamics study indicating that a substituent in the 2Ј-α-
position of a nucleoside NTP (e.g., a hydroxy or a fluoro sub-
stituent) is likely to collide with a tyrosine moiety of the active
site in HIV-1 RT, thereby preventing efficient binding and
thus anti-HIV activity.4 However, the 2Ј-deoxy-2Ј-fluororibo-
nucleotide used in this molecular dynamics study4 preferen-
tially adopts a twisted C2Ј-exo-C3Ј-endo conformation which,
though of the N-type, differs from the locked envelope-type
C3Ј-endo furanose conformation of nucleosides 3–6.
Synthesis of 1-(3-azido-3-deoxy-2-O,4-C-methylene-β--
ribofuranosyl)thymine 36,9 was accomplished using a published
procedure.6 However, for the synthesis of 9-(3-azido-3-deoxy-2-
O,4-C-methylene-β--ribofuranosyl)adenine (4; Scheme 2) we
found it more convenient to employ the furanoside 22 (with
methylsulfonyloxy groups instead of benzoyl groups6) for the
coupling reaction with adenine, and we used the known method
for synthesis of the di-O-methylsulfonyl derivative 22.24 Treat-
ment of 22 with 80% TFA to remove the isopropylidene group,
Results and discussion
In order to prepare the conformationally locked 3Ј-deoxy-
nucleosides 5 and 6 (Scheme 1) we chose as the first step to
remove the 3-hydroxy group of 1,2;5,6-di-O-isopropylidene-α-
-glucofuranose to give the 3-deoxyfuranose 9 as described
earlier.17 Preparation of derivative 12 via 10 and 11 was accom-
plished by hydrolysis of the primary acetonide, oxidative
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methanolic ammonia furnished the target adenine 3Ј-azido-3Ј-
deoxynucleoside 4 in 79% yield.
We also prepared the 3Ј-deoxynucleoside derivative 8, a
stereoisomer of derivative 5 (Scheme 3). Thus, the tri-O-(methyl-
Scheme 3 Reagents and conditions (and yields): i) MsCl, pyridine;
ii) 1 M NaOH, 1,4-dioxane (15% from 14 via 15).
sulfonyl)nucleoside 27 was prepared by methylsulfonylation of
derivative 15 using 2.6 equivalents of methylsulfonyl chloride in
pyridine. Compound 27 was subsequently dissolved in a mix-
ture of 1,4-dioxane and 1 M NaOH and heated at 90 ЊC for 18 h
whereupon the thymine 3Ј-deoxyderivative 8 was isolated in
15% yield (from 14 via 15), implying C2Ј-inversion, hydrolysis
of the putative 2Ј-C,2-O-anhydro intermediate, cyclization, and
conversion of the remaining methylsulfonyloxy group into a
hydroxy group in a one pot reaction. We have previously
reported similar cascade-type reactions during synthesis of
stereoisomeric LNA nucleosides.21
As none of the 5Ј-hydroxynucleosides 3–7 showed activity
against HIV-1 (vide infra), we decided to synthesize masked
5Ј-monophosphate derivatives (Scheme 4) in order to investi-
gate if the first phosphorylation step towards the corresponding
5Ј-triphosphates is the major obstacle for anti-HIV activity of
these locked nucleosides. We chose two different strategies,
namely the ‘cyclosal approach’ developed by Meier et al.11,12
and the ‘aryloxy phosphoramidate approach’ developed
McGuigan et al.,13–15 both of which have previously been used
successfully. The cyclosal approach allows release of the
nucleoside 5Ј-monophosphate inside the cell by chemical cleav-
age of the cyclosal group functioning as a phosphate-protecting
Scheme 2 Reagents and conditions (and yields): i) a) 80% TFA,
b) Ac2O, pyridine (90%, 2 steps); ii) adenine, SnCl4, acetonitrile,
60 ЊC (35%); iii) sat. methanolic ammonia; iv) 18-crown-6, KOAc,
1,4-dioxane (91%, 2 steps); v) saturated methanolic ammonia (79%).
followed by acetylation using acetic anhydride in pyridine,
afforded furanose 23 in 90% yield (2 steps). Coupling with
adenine was performed under the same conditions as described
above for nucleoside 18, furnishing nucleoside 24 in limited
(35%) yield. The product was subsequently dissolved in satur-
ated methanolic ammonia. After 3 h, two reaction products
were detected by analytical TLC, which were tentatively
assigned as the desired ring-closed nucleoside 25 and the 2-O-
deacetylated derivative of 24. When the reaction mixture was
instead stirred for three days only one product was observed
according to analytical TLC. This product was not purified, but
on the basis of NMR of the crude product it was identified as
nucleoside 25, and a subsequent substitution reaction using
potassium acetate as described above (synthesis of nucleo-
sides 17 and 21) afforded nucleoside 26 in 91% yield (from 24).
Eventually, 5Ј-O-deacetylation by treatment with saturated
Scheme 4 Reagents and conditions (and yields): i) 2-chloro-4H-1,3,2-benzodioxaphosphorine 28, DIPEA, acetonitrile; 0 ЊC; ii) tert-butyl hydroper-
oxide (yields 3a: 34%, 5a: 30%, 7a: 58%, 2 steps); iii) phenyl methoxyalaninylphosphorochloridate 29, N-methylimidazole, THF (3b: 38%, 4b: 16%,
5b: 22%).
J. Chem. Soc., Perkin Trans. 1, 2002, 1655–1662
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group during penetration into the cell. The procedure used for
synthesis of derivatives 3a, 5a and 7a (Scheme 4) was adopted
from the literature.11 Thus, the relevant nucleoside was dis-
solved in acetonitrile containing N,N-diisopropylethylamine
(DIPEA), and the phosphorylating reagent 2-chloro-4H-1,3,2-
benzodioxaphosphorine (28)11 was added followed after 1 h by
addition of tert-butyl hydroperoxide to give, after troublesome
column chromatographic purification, the masked 5Ј-mono-
phosphate derivatives 3a (34% yield), 5a (30% yield) and 7a
(58% yield) as diastereoisomeric mixtures.
Many different derivatives of the aryloxy phosphoramidate-
type of masked 5Ј-monophosphates have been prepared, e.g.,
by introducing substituents into the phenyl moiety or by chan-
ging the amino acid involved.14,15 They have been reported
to be enzymatically cleaved inside the cell, liberating the free
5Ј-monophosphates.13 We chose to synthesize the masked
monophosphate derivatives 3b, 4b and 5b (Scheme 4) con-
taining the parent phosphoramidate group because of their
relativelystraightforwardsynthesis.Thereagentphenylmethoxy-
alaninylphosphorochloridate (29)13,25 was treated with nucleo-
sides 3, 4 and 5 in THF to give the target molecules 3b (38%), 4b
(16%) and 5b (22%) as diastereoisomeric mixtures.
The bicyclic nucleosides 3a, 3b, 4, 4b, 5, 5a, 5b, 6, 7a and 8
were evaluated for antiviral activity against HIV-1 in MT-4
cells as described earlier.26 All compounds were inactive against
HIV-1 at 300 µM. Earlier, nucleosides 3 and 7 likewise tested
inactive against HIV-1.6,7 These results suggest that the parent
nucleosides and/or the nucleoside 5Ј-monophosphates and/or
the nucleoside diphosphates are not substrates for nucleoside
kinases, and/or that the nucleoside triphosphates are not recog-
nized by HIV-1 RT. However, as the 5Ј-triphosphate of the
N-type mimicking AZT analogue 1 is active against HIV-1,3
a possible explanation for the lack of activity for the ribo-
configured derivatives 3, 3a, 3b, 4, 4b, 5, 5a, 5b and 6 is
unfavourable steric interactions at the acitve site in HIV-1 RT
preventing efficient binding and thus anti-HIV activity, as
shown before for other ribo-configured nucleosides.4 Likewise,
the inactivity of the E-type nucleoside 7 and the derivatives 7a
and 8 suggests structural incompatibility between the core
bicyclic structures of these nucleosides and the active site of
one or more of the relevant enzymes.
purification. Petroleum ether of the distillation range 60–80 ЊC
was used. The silica gel (0.040–0.063 mm) used for column
chromatography was purchased from Merck. After drying of
organic phases using Na2SO4, filtration was performed. After
column chromatography, fractions containing product were
pooled, evaporated under reduced pressure, and dried over-
night under high vacuum (oil pump) to give the product unless
otherwise specified. 1H NMR spectra were recorded at 300
MHz, 13C NMR spectra at 75.5 MHz, and 31P NMR spectra at
121.5 MHz. Chemical shifts are reported in ppm relative to
either tetramethylsilane or the deuterated solvent as internal
standard for 1H NMR and 13C NMR, and relative to 85%
H3PO4 as external standard for 31P NMR. J-values are given
in Hz. Assignments of NMR spectra, when given, are based
on 2D spectra and follow the standard carbohydrate/
nucleoside nomenclature. Fast-atom bombardment mass
spectra (FAB-MS) were recorded in positive-ion mode.
3-Deoxy-5-O-methylsulfonyl-4-[(methylsulfonyloxy)methyl]-1,2-
O-isopropylidene-ꢀ-D-glycero-pentofuranose (12)
3-Deoxy-1,2;5,6-di-O-isopropylidene-α--ribo-hexofuranose17
(9) (11.0 g, 45.0 mmol) was dissolved in 80% acetic acid
(250 cm3) and stirring was continued for 20 h. The mixture was
evaporated to dryness under reduced pressure and the residue
was coevaporated with EtOH (3 × 100 cm3). The residue was
dissolved in a mixture of H2O (100 cm3) and THF (100 cm3),
the resulting mixture was cooled to 0 ЊC, and sodium periodate
(9.10 g, 42.5 mmol) was added. The mixture was stirred for 1 h
at 0 ЊC and then for 30 min at rt whereupon ethylene glycol
(6 cm3) was added. The mixture was filtered and the aqueous
phase was extracted with EtOAc (3 × 250 cm3). The combined
organic phase was evaporated to dryness under reduced pres-
sure. The residue was suspended in 1,4-dioxane (100 cm3), and
formaldehyde (37%; 9.00 cm3, 111 mmol) was added together
with 1 M NaOH (80 cm3). The mixture was stirred for 20 h
and was then extracted with EtOAc (6 × 150 cm3). The com-
bined organic phase was dried (Na2SO4), and evaporated to
dryness under reduced pressure. The residue was dissolved in
pyridine (150 cm3) and MsCl (10.5 cm3, 135 mmol) was added.
The mixture was stirred for 2 h at rt whereupon H2O (10 cm3)
was added. The mixture was evaporated to dryness under
reduced pressure, the residue was dissolved in DCM (300 cm3),
and washing was performed using saturated aq. NaHCO3
(3 × 150 cm3). The organic phase was dried (Na2SO4) and evap-
orated to dryness under reduced pressure to give a crude prod-
uct, which was purified by column chromatography and elution
Conclusions
The synthesis of a series of novel conformationally locked
3Ј-deoxy- and 3Ј-azido-3Ј-deoxy-nucleoside analogues has been
accomplished. The analogues were all devoid of any activity
against HIV-1 in MT-4 cells. Although nucleosides are known
preferentially to adopt N-type furanose conformation when
bound to the active site of HIV-1 RT, even masked 5Ј-mono-
phosphate derivatives of the locked N-type nucleosides contain-
ing a 2Ј-oxygen atom are inactive. The results obtained herein
indicate that the subsequent phosphorylation steps and/or the
interaction with HIV-1 RT are incompatible with the ribo-
configured locked N-type nucleoside structure. Unfortunately,
also the masked 5Ј-monophosphate derivative 7a of the con-
formationally restricted but somewhat flexible 3Ј-azido-3Ј-
deoxy E-type nucleoside 7 is devoid of anti-HIV activity.
with 0
2% MeOH in DCM (v/v) to give pentofuranose 12 as
a white solid material, which was used without further purifi-
cation in the next step; δC(CDCl3) 112.98, 107.06, 84.22, 80.61,
70.24, 69.72, 37.68, 35.48, 26.60, 25.59; δH(CDCl3) 5.90 (1H, d,
J 4.0), 4.84 (1H, d, J 2.1), 4.59–4.16 (4H, m), 3.13 (3H, s), 3.07
(3H, s), 1.61 (3H, s), 1.32 (3H, s); FAB-MS m/z 361 [M ϩ H]ϩ.
3-Deoxy-1,2-di-O-acetyl-5-O-methylsulfonyl-4-[(methyl-
sulfonyloxy)methyl]-ꢀ,ꢁ-D-glycero-pentofuranose (13)
Pentofuranose 12 (10.0 g) was dissolved in 80% TFA (100 cm3)
and stirring at rt was continued for 2 h at which point analytical
TLC showed complete conversion of the starting material. The
mixture was evaporated to dryness under reduced pressure and
the residue was coevaporated first with EtOH (2 × 100 cm3) and
then with pyridine (200 cm3). The residue was dissolved in
anhydrous pyridine (250 cm3) at rt and Ac2O (20.0 cm3,
211 mmol) was added. The reaction was quenched after 22 h at
rt by addition of H2O (20 cm3). The mixture was evaporated to
dryness under reduced pressure and the residue was dissolved
in EtOAc (500 cm3) and washed with saturated aq. NaHCO3
(3 × 150 cm3). The organic phase was dried (Na2SO4), and
evaporated to dryness under reduced pressure. The product was
obtained as a white solid material after dry column vacuum
Experimental‡
General
Reactions were conducted under an atmosphere of nitrogen
when anhydrous solvents were used. All reagents were obtained
from commercial suppliers and were used without further
‡ Copies of the 13C NMR spectra of compounds 4, 5, 6, 8, 18, 24, 25, 26
and 31P NMR spectra of compounds 3a, 3b, 4b, 5a, 5b and 7a
were enclosed with this manuscript on submission in order to verify
satisfactory purities.
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chromatography,27 and elution with 0
100% EtOAc in
1-(5-O-Acetyl-3-deoxy-2-O,4-C-methylene-ꢁ-D-erythro-pento-
petroleum ether (v/v) to give pentofuranose 13 (7.70 g) as a
white solid material. The not completely clean mixture was used
without any further purification in the next step; δC(CDCl3)
170.29, 169.95, 169.69, 169.20, 99.59, 94.14, 84.52, 82.57,
79.55, 76.95, 70.76, 70.52, 68.97, 68.37, 64.15, 63.52, 37.87,
37.60, 37.52, 20.95, 20.88, 20.74, 20.54; FAB-MS m/z 421
[M ϩ H]ϩ.
furanosyl)thymine (17)
Nucleoside 16 (370 mg, 1.1 mmol) was dissolved in anhydrous
1,4-dioxane (20 cm3). KOAc (870 mg, 8.90 mmol) and 18-
crown-6 (450 mg, 1.70 mmol) were added to the solution and
the mixture was heated under reflux for 6 h. The mixture was
then cooled to rt and the solvent was removed under reduced
pressure. The residue was dissolved in DCM (200 cm3) and
washed with saturated aq. NaHCO3 (3 × 100 cm3). The organic
phase was dried (Na2SO4), and evaporated to dryness under
reduced pressure to give a crude product, which was purified by
dry column vacuum chromatography and elution with 50
1-{2-O-Acetyl-3-deoxy-5-O-methylsulfonyl-4-
[(methylsulfonyloxy)methyl]-ꢁ-D-glycero-pentofuranoyl}thymine
(14)
100% EtOAc in petroleum ether (v/v) followed by 0
2.0%
Pentofuranose 13 (1.9 g) and thymine (1.2 g, 9.5 mmol) were
suspended in anhydrous acetonitrile (40 cm3) and N,O-bis-
(trimethylsilyl)acetamide (9.0 cm3, 36.4 mmol) was added. The
mixture was heated under reflux for 1 h and was then cooled
to 0 ЊC. Trimethylsilyl trifluoromethanesulfonate (2.70 cm3,
17.5 mmol) was added dropwise to the stirred mixture at 0 ЊC
whereupon the temperature was raised to 60 ЊC (for 24 h) and
then to reflux (for 48 h). After cooling of the mixture to rt,
additional trimethylsilyl trifluoromethanesulfonate (2.00 cm3,
11.1 mmol) was added and the mixture was heated under reflux
for an additional 24 h. After cooling to rt, the mixture was
evaporated to dryness under reduced pressure, and the residue
was dissolved in EtOAc (200 cm3) and washed with saturated
aq. NaHCO3 (3 × 75 cm3). The organic phase was dried
(Na2SO4), and evaporated to dryness under reduced pressure to
give a crude product, which was purified by column chrom-
MeOH in EtOAc (v/v) to give nucleoside 17 (220 mg, 66%) as a
white solid material, δC(CDCl3) 170.25, 163.97, 149.94, 134.40,
110.19, 88.40, 87.32, 78.37, 73.68, 61.04, 33.78, 20.77, 12.88;
δH(CDCl3) 8.52 (1H, s), 7.58 (1H, d, J 1.1), 5.67 (1H, s), 4.70
(1H, d, J 1.4), 3.95 (1H, d, J 10.9), 3.83–3.61 (3H, m), 1.93 (3H,
s), 1.89–1.76 (1H, m), 1.32–1.24 (4H, m); FAB-MS m/z 297
[M ϩ H]ϩ [Found (%): C, 52.52; H, 5.44; N, 9.16. C13H16N2O6
requires C, 52.70; H, 5.44; N, 9.16].
1-(3-Deoxy-2-O,4-C-methylene-ꢁ-D-erythro-pentofuranosyl)-
thymine (5)
Nucleoside 17 (50 mg, 0.17 mmol) was dissolved in saturated
methanolic ammonia (5 cm3). The mixture was stirred for 4 h at
rt and was then evaporated to dryness under reduced pressure.
The residue was coevaporated with EtOH (3 × 5 cm3) and the
crude product was purified by column chromatography and
elution with 3% MeOH in DCM (v/v), affording nucleoside 5
(28 mg, 65%) as a white solid material, δC(CD3OD) 166.50,
151.87, 136.90, 110.49, 91.51, 89.53, 79.73, 74.69, 59.77, 33.78,
12.59: δH(CD3OD) 7.52 (1H, d, J 1.2), 5.43 (1H, s), 4.67 (1H, s),
3.94 (1H, d, J 12.8), 3.85 (1H, d, J 12.8), 3.61–3.54 (1H, m),
1.78–1.66 (5H, m); MALDI-HRMS m/z 277.0794 ([M ϩ Na]ϩ,
C11H14N2O5ؒNaϩ Calc. 277.0795).
atography and elution with 0
3% MeOH in DCM (v/v) to
give nucleoside 14 (1.12 g, 23% from 9) as a white solid material,
δC(CDCl3) 170.55, 163.81, 150.44, 138.08, 111.62, 95.23, 84.24,
69.82, 68.89, 37.74, 37.49, 35.46, 20.76, 12.22 76; δH(CDCl3)
9.64 (1H, s), 7.28 (1H, s), 7.09 (1H, s), 5.65 (1H, d, J 3.0),
5.51–5.31 (1H, m), 4.47–4.27 (3H, m), 3.11 (3H, s), 3.09
(3H, s), 2.88–2.80 (1H, m), 2.22–2.16 (1H, m), 2.13 (3H, s), 1.92
(3H, s); FAB-MS m/z 471 [M ϩ H]ϩ [Found (%): C, 38.41; H,
4.71; N, 6.14. C15H22N2O11S2 requires C, 38.29; H, 4.71; N,
5.95%].
1-[3-Deoxy-2-O,4-C-methylene-5-O-(2-oxo-4H-1,3,2-benzo-
dioxaphosphorin-2-yl)-ꢁ-D-erythro-pentofuranosyl]thymine (5a)
1-{3-Deoxy-5-O-methylsulfonyl-4-[(methylsulfonyloxy)methyl]-
ꢁ-D-glycero-pentofuranosyl}thymine (15)
Nucleoside 5 (65 mg, 0.25 mmol) was dissolved in anhydrous
acetonitrile (4 cm3) under stirring and DIPEA (0.1 cm3) was
added. After cooling of the mixture to 0 ЊC, 2-chloro-4H-1,3,2-
benzodioxaphosphorine11 (28, 107 mg, 0.56 mmol) was added.
After 45 min at 0 ЊC the mixture was treated with tert-butyl
hydroperoxide (0.1 cm3, 5.0–6.0 M solution in THF) added
slowly. The mixture was allowed to reach rt, was stirred for an
additional 1 h, and then was evaporated to dryness under
reduced pressure. The crude product was purified by column
Nucleoside 14 (1.00 g, 2.12 mmol) was dissolved in a mixture of
MeOH (70 cm3) and saturated methanolic ammonia (40 cm3).
The mixture was stirred for 5 h and then evaporated to dryness
under reduced pressure. The crude product tentatively assigned
as nucleoside 15 was coevaporated with EtOH (3 × 100 cm3)
and used without further purification in the next step;
δC(CDCl3) 164.39, 151.14, 136.23, 110.91, 93.02, 82.86, 79.45,
73.70, 69.76, 69.49, 48.24, 36.79, 11.49.
chromatography and elution with 0
1.75% MeOH in DCM
(v/v) to give nucleoside 5a (32 mg, 30%) as a clear oil, δP(CDCl3)
Ϫ7.65, Ϫ7.87; δC(CDCl3) 163.88, 149.83, 134.32, 130.17,
125.47, 124.72, 118.62, 118.50, 110.11, 88.27, 88.21, 87.69,
87.63, 87.60, 87.53, 78.35, 73.15, 73.10, 68.72, 68.63, 65.08,
65.01, 64.87, 64.80, 33.48, 33.36, 12.51, 12.37; MALDI-HRMS
m/z 445.0772 ([M ϩ Na]ϩ, C18H19N2O8PؒNaϩ Calc. 445.0771).
1-(3-Deoxy-5-O-methylsulfonyl-2-O,4-C-methylene-ꢁ-D-
erythro-pentofuranosyl)thymine (16)
Nucleoside 15 (200 mg) was dissolved in anhydrous DMF (8
cm3), and NaH (60% suspension in oil; 110 mg, 2.75 mmol) was
added. After stirring for 3 h at rt the mixture was diluted with
DCM (100 cm3) and the resulting organic phase was washed
with saturated aq. NaHCO3 (3 × 50 cm3). The organic phase
was dried (Na2SO4), and evaporated to dryness under reduced
pressure to give a crude product, which was purified by column
chromatography and elution with 3% MeOH in DCM (v/v) to
give nucleoside 16 (101 mg, 64% from 14) as a white solid
material, δC(CD3OD) 163.80, 149.91, 134.47, 108.58, 87.60,
86.44, 77.85, 72.81, 72.72, 67.07, 36.83, 33.33, 12.20;
δH(CD3OD) 11.37 (1H, s), 7.44 (1H, s), 5.53 (1H, s), 4.94 (1H,
d, J 12.0), 4.82 (1H, d, J 11.6), 4.60 (1H, s), 3.75 (2H, dd, J 7.6
and 19.6), 3.29 (3H, s), 1.91–1.75 (5H, m); FAB-MS m/z 333 [M
ϩ H]ϩ [Found (%): C, 43.30; H, 4.83; N, 8.47. C12H16N2O7S
requires C, 43.37; H, 4.85; N, 8.43].
1-(3-Deoxy-2-O,4-C-methylene-5-O-{phenoxy[1-(methoxy-
carbonyl)ethylamino]phophoryl}-ꢁ-D-erythro-pentofuranosyl)-
thymine (5b)
Nucleoside 5 (20.0 mg, 0.107 mmol) was dissolved in a solution
of phenyl methoxyalaninylphosphorochloridate13,25 (29, 0.184
M solution in anhydrous THF; 2 cm3, 0.736 mmol). N-Methyl-
imidazole (0.1 cm3) and anhydrous pyridine (1 cm3) were added
to the solution and stirring was continued for 20 h at rt. The
temperature was raised to 45 ЊC and the mixture was stirred for
an additional 3 h. The mixture was evaporated to dryness under
reduced pressure and the residue was re-dissolved in DCM
(10 cm3) and washed successively with 1 M aq. HCl (2 × 5 cm3)
J. Chem. Soc., Perkin Trans. 1, 2002, 1655–1662
1659

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and saturated aq. NaHCO3 (2 × 5 cm3). The organic phase was
dried (Na2SO4), and evaporated to dryness under reduced pres-
sure to give a crude product, which was purified by column
any further purification in the next step; δ (CDCl ) 169.92 (C᎐
᎐
C
3
O), 155.55 (C-6), 152.95 (C-2), 148.86 (C-4), 139.70 (C-8),
120.04 (C-5), 90.67 (C-1Ј), 84.69 (C-4Ј), 76.38 (C-2Ј), 69.03,
68.27 (C-5Ј,C-5Љ), 37.56, 37.17 (2 × Ms), 35.22 (C-3Ј), 20.57
(CH3); δH(CDCl3) 8.33 (1H, s, H-8), 7.90 (1H, s, H-2), 7.28 (2H,
s, NH2), 6.15 (1H, d J 2.1, H-1), 5.90 (1H, m, H-2Ј), 4.56–4.13
(4H, m, H2-5Ј, H2-5Љ), 3.13 (3H, s, Ms), 2.95 (3H, s, Ms), 3.26–
2.98 (2H, m, H2-3Ј), 2.15 (3H, s, CH3CO); FAB-MS m/z 480 [M
ϩ H]ϩ.
chromatography: 0
3% MeOH in DCM (v/v) to give com-
pound 5b as a white solid material (12 mg, 22%), δP(CDCl3)
3.70, 3.48; MALDI-HRMS m/z 518.1284 ([M ϩ Na]ϩ, C21H26-
N3O9PؒNaϩ Calc. 518.1299).
1-[3-Azido-3-deoxy-2-O,4-C-methylene-5-O-(2-oxo-4H-1,3,2-
benzodioxaphosphorin-2-yl)-ꢁ-D-ribofuranosyl]thymine (3a)
9-(3-Deoxy-2-O,4-C-methylene-ꢁ-D-erythro-pentofuranosyl)-
adenine (6)
1-(3-Azido-2-O,4-C-methylene-β--ribofuranosyl)thymine6
3
(50 mg, 0.17 mmol) was dissolved in anhydrous acetonitrile
(4 cm3) and DIPEA (0.1 cm3) was added. The mixture was
cooled to 0 ЊC, and 2-chloro-4H-1,3,2-benzodioxaphosphor-
ine11 (28, 107 mg, 0.56 mmol) was added. Stirring was con-
tinued for 45 min at 0 ЊC, whereupon tert-butyl hydroperoxide
(0.15 cm3; 5.0–6.0 M solution in THF) was slowly added. The
mixture was allowed to reach rt and was then stirred for
an additional 1 h. After evaporation to dryness under
reduced pressure and coevaporation with anhydrous aceto-
nitrile (5 cm3), the crude product was purified by column
Nucleoside 18 (600 mg) was dissolved in saturated methanolic
ammonia (50 cm3) and the solution was stirred under N2 for
1 h. The mixture was then evaporated to dryness under
reduced pressure and the residue was coevaporated with EtOH
(2 × 50 cm3). This gave an intermediate white solid material
tentatively assigned as nucleoside 19, which was dissolved in
anhydrous DMF (20 cm3) followed by addition of NaH (60%
suspension in oil; 250 mg, 6.2 mmol). After stirring for 3 h at rt
the mixture was diluted with DCM (100 cm3) and the mixture
was washed with saturated aq. NaHCO3 (3 × 50 cm3). The
combined water phase was extracted with DCM (50 cm3), and
the combined organic phases were dried (Na2SO4), and evapor-
ated to dryness under reduced pressure. The residue was
purified by dry column vacuum chromatography and elution,
chromatography and elution with 1
1.5% MeOH in DCM
(v/v) to give nucleoside 3a (16 mg, 34%) as a white solid
material, δP(CDCl3) Ϫ8.01, Ϫ8.16; δC(CDCl3) 163.50, 149.61,
133.83, 130.14, 125.39, 124.72, 118.60, 118.46, 110.75, 87.30,
78,47, 70.95, 68.67, 62.42, 60.27, 12.26; MALDI-HRMS m/z
486.0765 ([M ϩ Na]ϩ, C18H18N5O8PؒNaϩ Calc. 486.0785).
first, with 80
100% EtOAc (v/v) in petroleum ether and then
0
1.0% MeOH in EtOAc (v/v) to give a slightly yellow oil
tentatively assigned as nucleoside 20. The product 20 was dis-
solved in anhydrous 1,4-dioxane (15 cm3), KOAc and 18-crown-
6 (450 mg, 1.7 mmol) were added under stirring, and the
mixture was heated with reflux for 3 h. The mixture was then
cooled to rt and evaporated to dryness under reduced pressure.
The residue was dissolved in EtOAc (60 cm3) and washed with
saturated aq. NaHCO3 (3 × 20 cm3). The organic phase was
dried (Na2SO4) and evaporated to dryness under reduced pres-
sure. The residue was purified by column chromatography and
1-(3-Azido-3-deoxy-2-O,4-C-methylene-5-O-{phenoxy-
[1-(methoxycarbonyl)ethylamino]phosphoryl}-ꢁ-D-ribo-
furanosyl)thymine (3b)
1-(3-Azido-2-O,4-C-methylene-β--ribofuranosyl)thymine6
3
(50 mg, 0.17 mmol) was dissolved in a solution of phenyl
methoxyalaninylphosphorochloridate13,25 (29, 0.184 M solution
in anhydrous THF; 4 cm3, 0.736 mmol). To the mixture were
added N-methylimidazole (0.1 cm3) and anhydrous pyridine
(1 cm3). Stirring was continued for 20 h at rt whereupon the
temperature was raised to 45 ЊC and stirring was continued for
an additional 3 h. The mixture was evaporated to dryness under
reduced pressure, the residue was dissolved in DCM (20 cm3),
and the solution was washed successively with 1 M aq. HCl
(2 × 7 cm3) and saturated aq. NaHCO3 (2 × 7 cm3). The organic
phase was dried (Na2SO4), and evaporated to dryness under
reduced pressure to give a crude product, which was purified by
column chromatography and elution with 3% MeOH in DCM
(v/v). The crude product obtained was again purified by column
elution with 2
5% MeOH in DCM (v/v) to give an inter-
mediate tentatively assigned as nucleoside 21 (110 mg) as a
white solid material. This material (100 mg) was dissolved in
saturated methanolic ammonia (10 cm3) and the mixture was
stirred at rt for 2 h and then evaporated to dryness under
reduced pressure. The residue was coevaporated with EtOH
(3 × 5 cm3) and the crude product was purified by column
chromatography and elution with 3% MeOH in DCM (v/v) to
give nucleoside 6 (42 mg, 3% from 9) as a white solid material,
δC((CD3)2SO) 155.86, 152.55, 148.30, 137.55, 89.30, 86.61,
78.07, 73.38, 58.29, 33.86; δH((CD3)2SO) 8.18 (1H, s), 8.14
(1H, s), 7.29 (2H, br s), 5.93 (1H, s), 4.76 (1H, s), 3.92 (2H, m),
3.76 (1H, d, J 7.8), 3.73 (1H, d, J 7.4), 3.31 (1H, s, overlapping
with H2O signal), 2.04 (1H, d, J 11.6), 1.94 (1H, d, J 11.3);
MALDI-HRMS m/z 286.0910 ([M ϩ Na]ϩ, C11H13N5O3ؒNaϩ
Calc. 286.0911).
chromatography; this time, elution with 0
3% MeOH in
DCM (v/v) afforded compound 3b (35mg, 38%), δP(CDCl3)
3.58, 3.50; MALDI-HRMS m/z 559.1291 ([M ϩ Na]ϩ, C21H25-
N6O9PؒNaϩ Calc. 559.1313).
9-{2-O-Acetyl-3-deoxy-5-O-methylsulfonyl-4-[(methylsulfonyl-
oxy)methyl]-ꢁ-D-glycero-pentofuranosyl}adenine (18)
3-Azido-3-deoxy-1,2-di-O-acetyl-6-O-methylsulfonyl-4-
[(methylsulfonyloxy)methyl]-ꢀ,ꢁ-D-erythro-pentofuranose (23)
Pentofuranose 13 (1.0 g) was coevaporated with anhydrous
acetonitrile (15 cm3) and dissolved in anhydrous acetonitrile
(15 cm3). Adenine (675 mg, 5.0 mmol) and then SnCl4 (0.75
cm3, 6.4 mmol) were added under stirring at rt. After 6 h, addi-
tional SnCl4 (0.75 cm3, 6.4 mmol) was added and stirring was
continued for an additional 20 h. Saturated aq. NaHCO3
(approx. 100 cm3) was slowly added until evolution of gas
ceased. The mixture was filtered through compressed Celite 545
which, after filtration, was flushed with DCM (800 cm3). The
combined organic phase was separated, dried (Na2SO4), and
evaporated to dryness under reduced pressure. The crude prod-
uct was purified by dry column vacuum chromatography and
Ribofuranose 2224 (700 mg, 1.7 mmol) was dissolved in
TFA (50 cm3; 80%) and after stirring for 1 h the mixture was
evaporated to dryness under reduced pressure. The residue was
coevaporated successively with CH3CN (2 × 50 cm3) and pyrid-
ine (50 cm3). The residue was dissolved in anhydrous pyridine
(50 cm3), and Ac2O (5 cm3, 52.8 mmol) was added. The mixture
was stirred at rt for 42 h whereupon H2O (15 cm3) was added.
The mixture was evaporated to dryness under reduced pressure
and the residue was dissolved EtOAc (50 cm3) and washing was
performed using saturated aq. NaHCO3 (3 × 20 cm3). The
organic phase was dried (Na2SO4), and evaporated to dryness
under reduced pressure. The crude product was purified by dry
elution with, first, 20
and then 0 5.0% MeOH in EtOAc (v/v) to give nucleoside 18
(604 mg) as a white solid material, and this was used without
100% EtOAc in petroleum ether (v/v)
column vacuum chromatography and elution with 0
70%
1660
J. Chem. Soc., Perkin Trans. 1, 2002, 1655–1662

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EtOAc in petroleumether (v/v) to afford ribofuranose 23 (687
mg, 90%) as a white solid material, which was not completely
clean but was used without any further purification in the next
step; δC(CDCl3) (data for major anomer) 169.26, 168.74, 97.54,
83.45, 75.17, 68.20, 67.84, 63.43, 37.95, 37.85, 20.99, 20.61;
MALDI-MS m/z 486 [M ϩ Na]ϩ.
(1H, s), 5.29 (1H, br s), 4.80 (1H, s), 4.58 (1H, s), 3.90–3.81 (4H,
m); MALDI-MS m/z 305.1 [M ϩ H]ϩ, 327.1 [M ϩ Na]ϩ.
3Ј-Azido-3Ј-deoxy-2Ј-O,4Ј-C-methylene-5Ј-O-{phenoxy-
[1-(methoxycarbonyl)ethylamino]phosphoryl}adenosine (4b)
Nucleoside 4 (20.0 mg, 0.065 mmol) was dissolved in a solu-
tion of phenyl methoxyalaninylphosphorochloridate13,25 29 in
anhydrous THF (0.184 M solution; 1.1 cm3, 0.20 mmol). To the
solution was added N-methylimidazole (0.05 cm3) and the mix-
ture was stirred at rt for 16 h before being evaporated to dryness
under reduced pressure. The residue was re-dissolved in DCM
(15 cm3) and washing was performed using saturated aq.
NaHCO3 (3 × 8 cm3). The organic phase was dried (Na2SO4),
and evaporated to dryness under reduced pressure. The crude
product was purified by column chromatography and elution
with 3% MeOH in DCM (v/v) to give compound 4b (6 mg,
16%) as a white solid material and starting nucleoside 4
(recovery 10 mg), δP((CD3)2SO) 4.84, 4.63; MALDI-HRMS
m/z 568.1432 ([M ϩ Na]ϩ, C21H24N9O7PؒNaϩ Calc. 568.1429).
9-{(2-O-Acetyl-3-azido-3-deoxy-5-O-methylsulfonyl-4-[(methyl-
sulfonyloxy)methyl]-ꢁ-D-erythro-pentofuranosyl}adenine (24)
Ribofuranose 23 (600mg, 1.34 mmol) was dissolved in
anhydrous CH3CN (15 cm3) and adenine (435 mg, 3.2 mmol)
was added. To the mixture was added SnCl4 (0.75 cm3,
6.4 mmol) and the temperature was raised to 60 ЊC. After con-
tinued stirring for 16 h at 60 ЊC, the temperature was changed to
rt and saturated aq. NaHCO3 (approx. 80 cm3) was added until
evolution of gas ceased. The mixture was filtered through com-
pressed Celite 545 which, after filtration, was flushed with
DCM (500 cm3). The combined organic phase was separated,
dried (Na2SO4), and evaporated to dryness under reduced
pressure. The product was purified by dry column vacuum
1-{3-Deoxy-2,5-di-O-methylsulfonyl-4-[(methylsulfonyloxy)-
methyl]-ꢁ-D-glycero-pentofuranosyl}thymine (27)
chromatography and elution, first, with 20
100% EtOAc
in petroleum ether (v/v) and then 0 5.0% MeOH in EtOAc
(v/v) to give nucleoside 24 (247 mg, 35%) as a white solid
material, δC((CD3)2SO) 169.411, 156.19, 152.93, 149.05, 139.67,
118.98, 85.28, 73.60, 69.06, 67.76, 62.08, 36.84, 36.72, 20.18;
δH((CD3)2SO) 8.31 (1H, s), 8.13 (1H, s), 7.36 (2H, s), 6.27 (1H,
d, J 5.3), 6.14 (1H, d, J 5.7), 5.21 (1H, d, J 6.5), 4.48 (2H, s),
4.38 (2H, s), 3.27 (3H, s), 3.23 (3H, s), 2.06 (3H, s); MALDI-
MS m/z 543 [M ϩ Na]ϩ.
Nucleoside 15 (700 mg) was dissolved in anhydrous pyridine
(20 cm3) and MsCl (0.2 cm3, 2.6 mmol) was added. The mixture
was stirred at rt for 48 h and was then evaporated to dryness.
The residue was dissolved in EtOAc (150 cm3) and washing was
performed using saturated aq. NaHCO3 (3 × 50 cm3). The
organic phase was dried (Na2SO4), and evaporated to dryness
under reduced pressure. The crude product was purified by dry
column vacuum chromatography and elution with, first, 50
9-(5-O-Acetyl-3-azido-3-deoxy-2-O,4-C-methylene-ꢁ-D-ribo-
furanosyl)adenine (26)
100% EtOAc in petroleum ether (v/v) and then 0
1.5%
MeOH in EtOAc (v/v) to give nucleoside 27 (320 mg) as a white
solid material. This was used without any further purification in
the next step; δC(CD3OD) 164.14, 151.32, 136.56, 110.62, 90.07,
81.25, 72.16, 71.09, 71.01, 37.23, 36.46, 32,85, 12.51; δH(CD3-
OD) 11.38 (1H, s), 7.49 (1H, d, J 0.71), 5.88 (1H, d, J 5.9), 5.73
(1H, d, J 4.8), 4.46–4.27 (4H, m), 3.59 (3H, s), 3.27 (3H, s),
3.25 (3H, s), 2.56–2.32 (2H, m), 1.79 (3H, s); FAB-MS m/z 507
[M ϩ H]ϩ.
Nucleoside 24 (200 mg, 0.38 mmol) was dissolved in saturated
methanolic ammonia (20 cm3). The mixture was stirred for 72 h
at rt whereupon the mixture was evaporated to dryness under
reduced pressure. The product was dried in vacuum (oil pump)
to give a white solid material tentatively assigned as nucleoside
25, which was used in the next step without further purification.
Data for 25: δC((CD3)2SO) 156.08, 152.84, 148.12, 138.17,
118.84, 85.62, 85.20, 78.97, 71.20, 65.58, 62.03, 36.91; δH((CD3)2-
SO) 8.30 (1H, s), 8.15 (1H, s), 7.34 (2H, s), 6.04 (1H, s), 4.92
(1H, s), 4.77 (1H, d, J 11.6), 4.76 (1H, s), 4.63 (1H, d, J 12.1),
3.94 (2H, dd, J 8.5 and 3.2), 3.24 (3H, s).
Compound 25 was dissolved in anhydrous 1,4-dioxane
(10 cm3) together with 18-crown-6 (200 mg, 0.75 mmol) and
KOAc (300 mg, 3.00 mmol). The mixture was heated under
reflux for 3 h and was then cooled to rt. The mixture was evap-
orated to dryness under reduced pressure and the residue was
dissolved in EtOAc (20 cm3). Washing was performed using
saturated aq. NaHCO3 (3 × 8 cm3), and the organic phase was
dried (Na2SO4), and evaporated to dryness under reduced pres-
sure to afford nucleoside 26 (122 mg, 91% from 24) as a white
solid material, δC(CDOD3) 170.94, 153.36, 138.20, 87.04, 86.98,
79.22, 72.27, 62.36, 59.62, 20.70; δH(CDOD3) 8.28 (1H, s), 8.06
(1H, s), 6.08 (1H, s), 5.00 (1H, s), 4.58 (1H, d, J 13.0), 4.50 (1H,
d, J 12.7), 4.39 (1H, s), 4.28 (1H, s), 4.09 (1H, d, J 8.5), 4.04
(1H, d, J 8.5), 2.18 (3H, s); MALDI-MS m/z 369.4 [M ϩ Na]ϩ.
1-(3-Deoxy-2-O,4-C-methylene-ꢀ-L-erythro-pentofuranosyl)-
thymine (8)
Nucleoside 27 (300 mg, 0.59 mmol) was dissolved in 1,4-
dioxane (10 cm3) and to the solution was added 1 M aq. NaOH
(6 cm3). The mixture was stirred for 4 h at rt and then for 18 h at
90 ЊC. After neutralization using 10% aq. HCl, the mixture was
evaporated to dryness and the crude product was purified by
column chromatography and elution with 3
4% MeOH in
DCM (v/v) to give nucleoside 8 (61 mg, 15% from 14) as a white
solid material, δC(CD3OD) 166.54, 153.07, 138.52, 110.37,
93.55, 92.40, 79.39, 76.67, 61.83, 61.72, 14.91; δH(CD3OD)
7.60 (1H, m), 5.82 (1H, d, J 4.8), 4.55 (1H, m), 4.04–3.76
(4H, m), 2.20–1.73 (5H, m); MALDI-HRMS m/z 277.09797
([M ϩ Na]ϩ, C11H14N2O5ؒNaϩ Calc. 277.0795).
1-[3-Azido-3-deoxy-2-O,3-C-methylene-5-O-(2-oxo-4H-1,3,2-
benzodioxaphosphorin-2-yl)-ꢁ-D-arabinofuranosyl]thymine (7a)
Nucleoside 77 (50 mg, 0.19 mmol) was dissolved in anhydrous
acetonitrile (4 cm3) and DIPEA (0.1 cm3) was added. The
stirred mixture was cooled to 0 ЊC and 2-chloro-4H-1,3,2-
benzodioxaphosphorine11 28 (90 mg, 0.47 mmol) was added.
The mixture was stirred for 1.5 h at 0 ЊC whereupon a solution
of tert-butyl hydroperoxide THF (5.0–6.0 M in anhydrous
THF; 0.1 cm3, 0.5–0.6 mmol) was added. The resulting mixture
was stirred for an additional 2 h during which time the mixture
was allowed to reach rt. The mixture was evaporated to dryness
under reduced pressure and the crude product was purified by
9-(3-Azido-3-deoxy-2-O,4-C-methylene-ꢁ-D-ribofuranosyl)-
adenine (4)
Nucleoside 26 (122 mg, 0.35 mmol) was dissolved in saturated
methanolic ammonia (5 cm3). The mixture was stirred for 24 h
at rt and then evaporated to dryness under reduced pressure.
The residue was crystallized from 40% MeOH in DCM to give
nucleoside 4 (85 mg, 79%) as a white solid material. Selected IR
signal: νmax 2118 cmϪ1 (azido group); δC((CD3)2SO) 156.03,
152.72, 148.71, 138.06, 118.95, 88.96, 85.09, 78.74, 71.46, 61.39,
56.57; δH((CD3)2SO) 8.30 (1H, s), 8.16 (1H, s), 7.35 (2H, s), 5.99
column chromatography and elution with 0
1% MeOH in
J. Chem. Soc., Perkin Trans. 1, 2002, 1655–1662
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12 C. Meier, E. De Clerq and J. Balzarini, Eur. J. Org. Chem., 1998, 837.
13 C. McGuigan, R. N. Pathinara, N. Mahmood, K. G. Devine and
A. J. Hay, Antiviral Res., 1992, 17, 107.
14 C. McGuigan, R. N. Pathirana, J. Balzarini and E. De Clerq, J. Med.
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15 C. McGuigan, H. W. Tsang, D. Cahard, K. Turner, S. Velazquez,
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DCM (v/v) to give compound 7a (46 mg, 58%), δP(CDCl3)
Ϫ8.43; δC(CDCl3) 163.74, 150.31, 137.04, 136.98, 130.15,
130.13, 125.49, 124.77, 118.84, 118.79, 118.72, 118.66, 110.49,
110.45, 87.15, 83.57, 83.52, 78.29, 78.19, 72.77, 72.70,
69.13, 69.04, 68.95, 66.80, 66.76, 64.81, 64.73, 64.66, 12.67;
MALDI-HRMS m/z 486.0787 ([M ϩ Na]ϩ, C18H18N5O8PؒNaϩ
Calc. 486.0785).
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