Synthesis of novel (2R,4R)- and (2S,4S)-iso dideoxynucleosides with exocyclic methylene as potential antiviral agents

Article abstract of DOI:10.1016/S0968-0896(01)00266-8

Novel (2R,4R)- and (2S,4S)-iso dideoxynucleosides with exocyclic methylene have been designed and synthesized, based on the lead BMS-200475 (3) which exhibited potent anti-HBV activity. For the synthesis of D types of (2R,4R)-nucleosides, L-xylose was converted to the key intermediate 14. The intermediate 14 was converted to the uracil derivative 4a and the cytosine derivative 4b. Compound 14 was also converted to the purine derivatives such as adenine derivative 4c, hypoxanthine derivative 4d, and guanine derivative 4e. The corresponding L types of (2S,4S)-enantiomers were more efficiently synthesized from the commercially available 1,2-isopropylidene-D-xylose (20) than the synthetic method used in the synthesis of (2R,4R)-nucleosides. The key intermediate 25 was converted to the pyrimidine analogues 5a and 5b and the purine derivatives 5c, 5d, and 5e using the similar method used in the preparation of 4c, 4d, and 4e. The synthesized final (2R,4R)- and (2S,4S)-nucleosides were tested against several viruses such as HIV-1, HSV-1, HSV-2, HCMV and HBV, (2R,4R)-Adenine analogue 4c exhibited potent anti-HBV activity (EC₅₀ = 1.5 μM in 2.2.15 cells) among compounds tested, while (2R,4R)-uracil derivative 4a was the most active against HCMV among compounds tested and (2R,4R)-adenine derivative 4c was found to be moderately active against the same virus. However, the corresponding (2S,4S)-isomers were found to be totally inactive against all tested viruses. Both (2R,4R)-adenine derivative 4c and (2S,4S)-adenine analogue 5c were totally resistant to the adenosine deaminase like iso-ddA (1). From the molecular modeling study the hydroxymethyl side chains of BMS-200475 (3) and 4c were almost overlapped, indicating that 4c may be suitable for phosphorylation by cellular kinases like the lead 3, but some discrepancy between two bases was observed, indicating why 4c is less potent against HBV than 3. It is concluded that discovery of (2R,4R)-adenine analogue 4c as potent anti-HBV agent suggested that the sugar moiety of this series can be regarded as a novel template for the development of new anti-HBV agent and oxygen atom can be acted as a bioisoster of C-OH. Copyright

Full text of DOI:10.1016/S0968-0896(01)00266-8

Bioorganic & Medicinal Chemistry10 (2002) 215–226
Synthesis of Novel (2R,4R)- and (2S,4S)-iso Dideoxynucleosides
with Exocyclic Methylene as Potential Antiviral Agents1
Su Jeong Yoo,a Hea Ok Kim,b Yoongho Lim,c Jeongmin Kimd and Lak Shin Jeonga,*
aLaboratory of Medicinal Chemistry, College of Pharmacy, Ewha Womans University, Seoul 120-750, Republic of Korea
bDepartment of Chemistry and Molecular Engineering, Seoul National University, Seoul 151-742, Republic of Korea
cDepartment of Applied Biology & Chemistry, Konkuk University, Seoul 143-701, Republic of Korea
dLG Chem/Biotech Research Institute, Taejon, 305-380, Republic of Korea
Received 1 June 2001; accepted 21 July2001
Abstract—Novel (2R,4R)- and (2S,4S)-iso dideoxynucleosides with exocyclic methylene have been designed and synthesized, based
on the lead BMS-200475 (3) which exhibited potent anti-HBV activity. For the synthesis of d types of (2R,4R)-nucleosides, l-xylose
was converted to the keyintermediate 14. The intermediate 14 was converted to the uracil derivative 4a and the cytosine derivative
4b. Compound 14 was also converted to the purine derivatives such as adenine derivative 4c, hypoxanthine derivative 4d, and
guanine derivative 4e. The corresponding l types of (2S,4S)-enantiomers were more efficientlysynthesized from the commercially
available 1,2-isopropylidene-d-xylose (20) than the synthetic method used in the synthesis of (2R,4R)-nucleosides. The keyinter-
mediate 25 was converted to the pyrimidine analogues 5a and 5b and the purine derivatives 5c, 5d, and 5e using the similar method
used in the preparation of 4c, 4d, and 4e. The synthesized final (2R,4R)- and (2S,4S)-nucleosides were tested against several viruses
such as HIV-1, HSV-1, HSV-2, HCMV and HBV. (2R,4R)-Adenine analogue 4c exhibited potent anti-HBV activity(EC 50=1.5 mM
in 2.2.15 cells) among compounds tested, while (2R,4R)-uracil derivative 4a was the most active against HCMV among compounds
tested and (2R,4R)-adenine derivative 4c was found to be moderatelyactive against the same virus. However, the corresponding
(2S,4S)-isomers were found to be totallyinactive against all tested viruses. Both (2 R,4R)-adenine derivative 4c and (2S,4S)-adenine
analogue 5c were totallyresistant to the adenosine deaminase like iso-ddA (1). From the molecular modeling studythe hydro-
xymethyl side chains of BMS-200475 (3) and 4c were almost overlapped, indicating that 4c maybe suitable for phosphorylation by
cellular kinases like the lead 3, but some discrepancybetween two bases was observed, indicating why 4c is less potent against HBV
than 3. It is concluded that discoveryof (2 R,4R)-adenine analogue 4c as potent anti-HBV agent suggested that the sugar moietyof
this series can be regarded as a novel template for the development of new anti-HBV agent and oxygen atom can be acted as a
bioisostere of C–OH. # 2001 Elsevier Science Ltd. All rights reserved.
Introduction
Recently, d- and l-carbocyclic nucleosides with exo-
cyclic methylene in place of furanose oxygen were syn-
thesized and evaluated for antiviral activities, among
which d-guanine derivative (BMS-200475, 3) was very
active against hepatitis B virus (HBV) and was 100 times
Iso dideoxynucleosides2ꢀ6 belong to unique nucleosides
in that furanose oxygen was moved to the C3 position,
among which adenine analogue (iso-ddA, 1)2,4 and
guanine analogue (iso-ddG, 2)4 exhibited potent anti-
HIV activitycomparable to 2 0,30-dideoxyadenosine
(ddA) and 20,30-dideoxyguanosine (ddG), respectively.
Besides potent anti-HIV activity, this class of nucleo-
sides also possess chemical and metabolic advantages
such as glycosyl bond stabilization and resistance to the
adenosine deaminase, which 20,30-dideoxynucleosides do
not have (Fig. 1).2ꢀ4
7
more potent than clinicallyavailable drug, lamivudine.
Based on these findings and bioisosteric concept, it was
of great interest to design and synthesize d types of
(2R,4R)-iso dideoxynucleosides 4 substituted with exo-
cyclic methylene from the lead 3 because C–OH was
known to serve as a bioisostere of oxygen as in the case
of 1,3-dioxolanyl nucleosides.8 It was also interesting to
synthesize their l types of the corresponding (2S,4S)-
enantiomers 5 for comparison of their antiviral activity
since many l-nucleosides9 were found to be more potent
than the corresponding d-nucleosides. Here, we report
*Corresponding author. Tel.: +82-2-3277-3466; fax: +82-2-3277-
2851; e-mail: lakjeong@mm.ewha.ac.kr
0968-0896/02/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0968-0896(01)00266-8

216
S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
the full accounts of synthesis and antiviral activity of
novel (2R,4R)- and (2S,4S)-iso dideoxynucleosides with
exocyclic methylene.
12 with boron trichloride in methylene chloride at
ꢀ78 ꢁC gave the diol 13 which was selectivelysilylated
with TBDPSCl to yield the key intermediate 14.
Synthesis of the desired d-nucleosides from d-allylic
alcohol 14 was achieved bythe Mitsunobu reaction
and is well shown in Scheme 2.
14
Results and Discussion
Chemistry
Condensation of 14 with N3-benzoyluracil under the
standard Mitsunobu conditions afforded the protected
uracil derivative 15 (63%) with concomminant forma-
tion of O-glycosylated product (10%). The regioisomers
was easilyconfirmed bythe comparison of the UV lit-
erature data.14 Debenzoylation of 15 with sodium
methoxide in methanol followed bydesillyation with
tetra-n-butylammonium fluoride produced the final d-
Our synthetic plan to the desired (2R,4R)- and (2S,4S)-
iso dideoxynucleosides is to synthesize the glycosyl
donor from a chiral template, carbohydrate precursor
and then to condense with nucleosidic bases. Synthesis
of the key d-glycosyl donor 14 from l-xylose is shown
in Scheme 1.
0
l-Xylose (6) was converted to compound 7 in three
steps.10 Methylation of 6 at the anomeric position with
0.5% methanolic HCl, treatment of the triol with
CuSO4 and p-TsOH in acetone, and benzylation at the
C2 position with benzyl bromide afforded 7 (85% from
6). Hydrolysis of the 3,5-acetonide in 7 with 70% acetic
acid gave the diol 8 in quantitative yield. Selective ben-
zylation of 8 was achieved using well-known organotin
chemistry11 to give 9 in 99% yield. To remove the
methoxygroup at the anomeric position, compound 9
was first refluxed with hexamethyldisilazane (HMDS) to
protect the C3-position in situ to the trimethylsilyl
(TMS) ether and then without purification, treated with
triethylsilane in the presence of TMSOTf at room tem-
perature for 2 h to give 10 in 80% yield after the
uracil derivative 4a. The stereochemistryof the C2
position in compound 4a was confirmed byNOESY
experiment, proving that Mitsunobu condensation of
the allylic alcohol was proceeded in pure SN2 reaction,
not SN1 or SN20 reaction.15 For the preparation of the
cytosine derivative 4b, compound 4a was treated with
acetic anhydride in pyridine to protect the hydroxyl
group followed bytreatment of the acetate with 1,2,4-
triazole and phosphorous oxytrichloride in triethyl
amine to give the triazole derivative. Without purifica-
tion, triazole derivative was treated with ammonium
hydroxide in dioxane followed by deacetylation with
methanolic ammonia to give the final d-cytosine deri-
vative 4b. For the synthesis of the d-purine derivatives,
d-glycosyl donor 14 was condensed with 6-chloropurine
and 2-amino-6-chloropurine under the same Mitsunobu
12
purification bysilica gel column chromatograph.y
This procedure was proved to be more efficient than the
following method: benzoylation of 9 followed bytreat-
ment of the resulting benzoate with triethylsilane and
TMSOTf and then debenzoylation to give the same
product 10 (61% from 9). Oxidation of 10 with DMSO
and acetic anhydride produced ketone 11 (80%). Wittig
reaction to convert the ketone 11 to the olefin 12 was
found to be greatlyaffected bythe base used in the
reaction. When 11 was treated with methyl triphenyl-
phosphonium bromide in the presence of NaH or n-BuLi,
extensive decomposition on TLC was observed with low
yield (30–40%) of the desired product. However, addition
of t-amyl alcohol in the presence of NaH and methyl tri-
phenylphosphonium bromide to the reaction mixture
resulted in high yield (92%) of 12.13 Debenzylation of
Scheme 1. Reagents: a) (i) HCl, MeOH, rt. (ii) CH3COCH3, p-TsOH,
CuSO4. (iii) BnBr, NaH, n-Bu4NI. b) 70% AcOH, 60 ꢁC. c) (i) n-
Bu2SnO, toluene, reflux. (ii) n-Bu4NBr, BnBr, 100 ꢁC. d) (i) HMDS,
reflux. (ii) Et3SiH, TMSOTf, rt. e) DMSO, Ac2O, rt. f) Ph3PCH3Br,
NaH, t-amyl alcohol, 0 ꢁC. g) BCl3, CH2Cl2, ꢀ78 ꢁC. h) TBDPSCl,
imidazole, DMF, 0 ꢁC.
Figure 1. Rationale to the design of the target nucleosides.

S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
217
Scheme 2. Reagents: a) N3-benzoyluracil or 2-amino-6-chloropurine, PPh3, DEAD, THF, ꢀ10 ꢁC. b,c) NaOMe, MeOH, 0 ꢁC. d) (i) Ac2O, pyridine,
rt. (ii) POCl3, 1,2,4-triazole, Et3N, rt. (iii) NH4OH, dioxane, rt. (iv) NaOMe, MeOH, 0 ꢁC. c) n-Bu4NF, THF, 0 ꢁC. e) NH3/MeOH, 100 ꢁC. f) 1 N
NaOH, reflux.
conditions14 to give d-6-chloropurine analogue 16
(75%) and d-2-amino-6-chloropurine analogue 17
(50%), respectively. In case of 6-chloropurine, N7-iso-
mer was not formed during the Mitsunobu reaction,
while 2-amino-6-chloropurine condensation produced
the N7-substituted product (5%), in which UV of N9-
isomer was appeared at 308 nm, while that of N7-isomer
was shown at 323 nm.16 Desilylation of 16 with tetra-n-
butylammonium fluoride gave d-6-chloropurine deriva-
tive 18. Compound 18 was converted to the d-adenine
analogue 4c and the d-hypoxanthine analogue 4d by
treating 18 with methanolic ammonia at 100 ꢁC and
refluxing with 1 N NaOH, respectively. The d-guanine
derivative 4e was obtained byrefluxing 19 obtained
from the desilylation of 17, with 1 N NaOH.
Condensation of 25 with N3-benzoyluracil under the
Mitsunobu conditions gave the protected nucleoside 26
which was debenzoylated with sodium methoxide to
afford the final l-uracil derivative 5a. The final l-cyto-
sine derivative 5b was obtained from 5a using the same
method used in Scheme 2. Mitsunobu condensation of
25 with 6-chloropurine and 2-acetamido-6-chloropurine
yielded the protected nucleosides 27 and 28, respec-
tively. After deblocking of 27 and 28 with sodium
methoxide, l-6-chloropurine derivative 29 and l-2-
amino-6-chloropurine derivative 30 were obtained,
respectively. Treatment of compound 29 with metha-
nolic ammonia at 100 ꢁC afforded the adenine derivative
The corresponding (2S,4S)-enantiomers of the synthe-
sized (2R,4R)-nucleosides could be more efficientlysyn-
thesized starting from commerciallyavailable 1,2-
isopropylidene-d-xylose (20) than the method used in
the preparation of (2R,4R)-nucleosides.17
As shown in Scheme 3, 1,2-isopropylidene-d-xylose (20)
was converted to the ketone 21 according to the known
procedure.18 Wittig reaction of 21 with methyl triphe-
nylphosphonium bromide, t-amyl alcohol and NaH was
smoothlyproceeded to yield the desired olefin 23 with
the debenzoylated olefin 22 which could be rebenzoyl-
ated to 23 in quantitative yield. Acid-catalyzed hydro-
lysis (95%) of 1,2-acetonide of 23 followed byanomeric
demethoxylation12 (80%) of the resulting methoxide 24
gave the keyintermediate 25 in verygood yield. This
method was proved much better than that used in
Scheme 1 in terms of total steps and overall yield.
Synthesis of the (2S,4S)-nucleosides from the inter-
mediate 25 was achieved according to the similar pro-
cedure used in Scheme 2 and is depicted in Scheme 4.
Scheme 3. Reagents: a) Ph3PCH3Br, NaH, t-amyl alcohol, 0 ꢁC. b)
BzCl, pyridine, rt. c) 1% HCl, MeOH, rt. d) (i) HMDS, (NH4)2SO4.
(ii) Et3SiH,TMSOTf, rt.

218
S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
Scheme 4. Reagents: a) N3-benzoyluracil or 2-acetamido-6-chloropurine, PPh3, DEAD, THF, ꢀ10 ꢁC. b) NaOMe, MeOH, 0 ꢁC. c) (i) Ac2O, pyri-
dine, rt. (ii) POCl3, 1,2,4-triazole, Et3N, rt. (iii) NH4OH, dioxane, rt. (iv) NaOMe, MeOH, 0 ꢁC. d) NH3/MeOH, 100 ꢁC. e) 1 N NaOH, reflux.
5c. l-Hypoxanthine analogue 5d was directlyobtained
from the protected nucleoside 27 byrefluxing it with 1 N
NaOH. Similarly, l-2-amino-6-chloropurine 30 was
transformed to the l-guanine derivative 5e.
stand why 4c is active against HBV and less active than
BMS-200475.
As seen in Figure 2, we superimposed the lead 3 with
(2R,4R)-adenine analogue 4c to find out whether
hydroxymethyl side chains of both compounds overlap
well. As expected, two compounds were well overlapped
and their hydroxymethyl side chains were almost coin-
cident, which maybe suitable for phosphorylation by
cellular kinases, but their bases did not match perfectly
and the value of the root mean square deviation
between two structures is 0.11 A, which mayexplain
lower anti-HBV activityof compound 4c than that of
the lead 3. The anti-HBV activityof compound 4c may
Antiviral activity
All synthesized (2R,4R)- and (2S,4S)-nucleosides were
tested against several viruses such as HIV-1 (MT-4
cells), HSV-1 (CCL81 cells), HSV-2 (CCL81 cells),
HCMV (AD-169) and HBV (2.2.15 cells). None of the
final nucleosides was found to be active against HIV-1,
HSV-1 and HSV-2 up to 100 mg/mL, but manynucleo-
sides exhibited weak to potent antiviral activities against
HCMV and HBV as shown in Table 1.
Table 1. Anti-HCMV and anti-HBV activities of the synthesized
d- and l-nucleosides
d-Adenine analogue 4c exhibited the most potent anti-
HBV activityamong compounds tested although it was
less potent than the control, lamivudine. To the best of
our knowledge, since none of the iso dideoxynucleosides
was reported to show anti-HBV activityso far, com-
pound 4c is the first example to show anti-HBV activity
in this type of nucleosides. No anti-HIV activity of 4c
unlike iso-ddA might be explained bythe fact that HIV-1
prefers flexible conformation of iso-ddA like 20,30-
dideoxynucleosides to the rigid conformation of 4c. d-
Uracil derivative 4a was the most potent against HCMV
among tested and d-adenine derivative 4c was found to
be moderatelyactive against the same virus. It is inter-
esting to note that (2R,4R)-isomers exhibited potent
antiviral activities, while the corresponding (2S,4S)-iso-
mers were found to be totallyinactive against all tested
viruses, indicating that only(2 R,4R)-derivatives might
be phosphorylated by cellular kinases. We also exam-
ined the affinityto the adenosine deaminase for the
adenine derivatives 4c and 5c. Theywere found to be
resistant to adenosine deaminase like iso-ddA (1).2ꢀ4 We
have done the molecular modeling study19 to under-
HCMVa
HBVb
Cytotoxiciy
EC50 (mg/mL) EC50 (mM) CC50 (mg/mL)
d-Uracil (4a)
10.6
>100
33.3
> 100
>100
>100
>100
>100
>100
>100
>10
>10
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
d-Cytosine (4b)
d-Adenine (4c)
d-Hypoxanthine (4d)
d-Guanine (4e)
l-Uracil (5a)
1.5
>10
>10
>10
>10
>10
>10
>10
l-Cytosine (5b)
l-Adenine (5c)
l-Hypoxanthine (5d)
l-Guanine (5e)
Ganciclovir
0.74
ND
ND
0.05
Lamivudine
aAD-169 infected cells.
b2.2.15 cells.

S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
219
2-O-Benzyl-1-O-methyl-3,5-O-isopropylidene-ꢀ,ꢁ-L-xy-
lofuranoside (7). To a stirred solution of l-xylose (10 g,
66.61 mmol) in anhydrous methanol (46 mL) was added
acetyl chloride (0.31 mL) dropwise at ambient tempera-
ture under nitrogen and the reaction mixture was stirred
overnight at ambient temperature under nitrogen. The
reaction mixture was neutralized with silver nitrate and
filtrated through a Celite pad. The filtrate was evapo-
rated to dryness to give a crude mixture of 1-methyl-
a,b-l-xylose as a sticky oil, which, without purification,
.
was treated with p-TsOH H2O (508 mg, 2.67 mmol) and
anhydrous copper(II) sulfate (205 g, 1.286 mol) in anhy-
drous acetone (60 mL) at ambient temperature under
nitrogen. The reaction mixture was stirred overnight at
ambient temperature under nitrogen. The solid was
removed byfiltration and the filtrate was neutralized
with ammonia water and evaporated. The residue was
extracted with methylene chloride, washed with water,
dried with anhydrous MgSO4 and evaporated. The
residue was purified byflash silica gel column chroma-
tography(hexanes/EtOAc=4:1) to give methyl-3,5- O-
isopropylidene-a,b-l-xylofuranoside (13.6 g, 100%) as a
yellow oil. To a stirred solution of methyl-3,5-O-iso-
propylidene-a,b-l-xylofuranoside (6 g, 29.38 mmol) in
dryTHF (50 mL) were added NaH (1.76 g, 60% in oil,
44.07 mmol) and BnBr (4.19 mL, 35.26 mmol), followed
by n-Bu4NI (1.1 g, 2.94 mmol) at ice bath under nitrogen
and the reaction mixture was stirred for 2 h at ambient
temperature under nitrogen. The reaction mixture was
neutralized with acetic acid, extracted with EtOAc,
washed with water, dried with anhydrous MgSO4 and
evaporated. The residue was purified byflash silica gel
column chromatography(hexanes/EtOAc=7:1 to 4:1 to
Figure 2. Superimposition of two structures obtained through com-
puter aided molecular modeling calculations (black; BMS-200475, red;
compound 4c).
be also attributed to the rigid sugar conformation
endowed byexoccylic double bond as in the case of
BMS-200475. To our best knowledge, since none of the
iso dideoxynucleosides was reported to show anti-HBV
activityso far, compound 4c is the first example to show
anti-HBV activityin this type of nucleosides. No anti-
HIV activityof 4c unlike iso-ddA might be explained by
the fact that HIV-1 prefers flexible conformation of iso-
ddA like 20,30-dideoxynucleosides to the rigid con-
formation of 4c.
Conclusion
1
2:1) to give 7 (7.35 g, 85.0%) as a yellow oil: H NMR
In summary, we have completed the synthesis and
structure–activityrelationship studyof (2 R,4R)- and
(2S,4S)-iso dideoxynucleosides with exocyclic methylene
as antiviral agents. Although we could not find excellent
antiviral agents, when compared to the control com-
pounds, several compounds including (2R,4R)-adenine
analogue 4c exhibited significant antiviral activities,
indicating that the sugar moietyof this series can be
regarded as a novel template for the development of
new antiviral agents and oxygen atom can be acted as a
bioisostere of C–OH.
(CDCl3) d 1.35 (s, 3H, a isomer CH3), 1.36 (s, 3H, a
isomer CH3), 1.38 (s, 3H, b isomer CH3), 1.39 (s, 3H, b
isomer CH3), 3.42 (s, 3H, b isomer OCH3), 3.46 (s, 3H,
a isomer OCH3), 3.76–4.00 (m, 3H, a and b isomer 2-H
and 5-H), 4.15–4.19 (m, 1H, a and b isomer 4-H), 4.24
(d, 1H, J=4.0 Hz, b isomer 3-H), 4.29 (dd, 1H,
J=2.4 Hz, J=4.4 Hz, a isomer 3-H), 4.61 (d, 2H,
J=1.2 Hz, b isomer benzylic H), 4.66 (d, 2H, J=6.0 Hz,
a isomer benzylic H), 4.98 (s, 1H, b isomer 1-H), 5.01
(d, 1H, J=4.4 Hz, a isomer 1-H), 7.29–7.39 (m, 5H, a
and b isomer Ph). Anal. calcd for C16H22O5: C, 65.29;
H, 7.53. Found: C, 65.29; H, 7.77.
2-O-Benzyl-1-O-methyl-ꢀ,ꢁ-L-xylofuranoside (8).
A
Experimental
solution of 7 (5.9 g, 16.99 mmol) in 70% acetic acid
(25 mL) was stirred for 1 h at 60 ꢁC. The reaction mix-
ture was evaporated and the residue was further dried
bycoevaporating with toluene and ethanol. The residue
was purified byflash silica gel column chromatography
(chloroform/methanol=100:1 to 50:1) to give 8 (5.10 g,
100%) as a yellow oil, which was subjected to the next
reaction.
Ultra violet (UV) spectra were recorded on a Beckman
DU-68 spectrophotometer and 1H and 13C spectra were
recorded on Varian-400 (250 and 100 MHz) spectro-
meter, respectively, using CDCl3 or DMSO-d6 and che-
mical shifts are reported in parts per million (d)
downfield from tetramethylsilane as internal standard.
FAB mass spectra were recorded on Jeol HX 110 spec-
trometer. Elemental analyses were performed at the
general instrument laboratoryof Ewha Womans Uni-
versity, Korea. TLC was performed on Merck pre-
coated 60F254 plates. Column chromatographywas
performed using silica gel 60 (230–400 mesh, Merck).
All the anhydrous solvents were distilled over CaH2 or
P2O5 or Na/benzophenone prior to the reaction.
2,5-Di-O-benzyl-1-O-methyl-ꢀ,ꢁ-L-xylofuranoside (9).
To a stirred solution of 8 (3 g, 11.80 mmol) in toluene
(50 mL) was added n-Bu2SnO (4.41 g, 17.70 mmol) and
the reaction mixture was heated for 5 h at 140–150 ꢁC
under nitrogen. After the oil bath was cooled to 90–
100 ꢁC, n-Bu4NBr (1.90 g, 5.90 mmol) and BnBr

220
S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
(2.22 mL, 17.70 mmol) were added to the reaction mix-
ture. The whole mixture was heated overnight at 90–
100 ꢁC under nitrogen. The solvent was evaporated and
the residue was purified byflash silica gel column chro-
matography(hexanes/EtOAc=3:1) to give 9 (4.01 g,
99.0%) as a yellow oil.
(10 mL) was added NaOMe (1 M solution in MeOH,
0.08 mL, 0.08 mmol) at 0 ꢁC under nitrogen and the
reaction mixture was stirred for 2 h at 0 ꢁC under nitro-
gen. The reaction mixture was neutralized with acetic
acid and evaporated. The residue was dissolved in
methylene chloride and the organic layer was washed
with water, dried with anhydrous MgSO4 and evapo-
rated. The residue was purified byflash silica gel column
a-Isomer. 1H NMR (CDCl3) d 3.04 (d, 1H, J=9.8 Hz,
OH), 3.33 (s, 3H, OCH3), 3.68 (dd, 1H, J=6.2, 10.4 Hz,
5-Ha), 3.82 (dd, 1H, J=4.9, 10.4 Hz, 5-Hb), 3.92 (s, 1H,
2-H), 4.20 (m, 1H, 4-H), 4.60 (dd, 1H, J=4.8, 10.7 Hz,
3-H), 4.63–4.57 (m, 4H, benzylic H), 4.93 (s, 1H, 1-H),
7.34–7.24 (m, 10H, 2ꢂPh). Anal. calcd for C20H24O5: C,
69.75; H, 7.02. Found: C, 69.78; H, 7.18.
chromatography(hexanes/EtOAc=2:1) to give
10
1
(110 mg, 85%) as a white oil: H NMR (CDCl3) d 3.48
(d, 1H, J=3.9 Hz, OH), 3.83 (dd, 1H, J=2.2, 9.8 Hz, 1-
Ha), 3.89 (d, 2H, J=4.9 Hz, 5-H), 4.03 (m, 1H, 2-H),
4.12 (q, 1H, J=3.9 Hz, 3-H), 4.22 (dd, 1H, J=4.9,
9.8 Hz, 1-Hb), 4.35 (m, 1H, 4-H), 4.53–4.67 (m, 4H,
benzylic H), 7.27–7.39 (m, 10H, 2ꢂPh). Anal. calcd for
C19H22O4: C, 72.59; H, 7.05. Found: C, 72.86; H, 6.97.
b-Isomer. 3.08 (d, 1H, J=7.6 Hz, OH), 3.38 (s, 3H,
OCH3), 3.72 (d, 2H, J=3.8 Hz, 5-H), 3.87 (dd, 1H,
J=4.3, 6.3 Hz, 2-H), 4.11 (m, 1H, 4-H), 4.28 (dd, 1H,
J=3.7, 7.6 Hz, 3-H), 4.60–4.49 (m, 4H, benzylic H),
4.79 (d, 1H, J=4.3, 1-H), 7.35–7.25 (m, 10H, 2ꢂPh).
Anal. calcd for C20H24O5: C, 69.75; H, 7.02. Found: C,
69.45; H, 7.42.
Method B. A mixture of 9 (1 g, 2.90 mmol) and
ammonium sulfate (catalytic amount) in HMDS (2 mL)
was refluxed for 1 h under nitrogen. The reaction mix-
ture was cooled to ambient temperature and the solvent
was evaporated under anhydrous conditions. The resi-
due was dissolved in anhydrous methylene chloride
(15 mL) and triethylsilane (2.32 mL, 14.50 mmol) was
added to this solution followed byadding slowly
1,4-Anhydro-2,5-di-O-benzyl-L-xylitol (10). Method A.
To a stirred solution of 9 (3.7 g, 10.74 mmol) in pyri-
dine (40 mL) was added benzoyl chloride (1.87 mL,
16.12 mmol) dropwise at ambient temperature under
nitrogen and the reaction mixture was stirred for 2 h at
ambient temperature under nitrogen. The reaction mix-
ture was quenched with methanol and evaporated. The
residue was dissolved in methylene chloride and the
organic layer was washed with water, dried with anhy-
drous MgSO4 and evaporated. The residue was purified
byflash silica gel column chromatography(hexanes/
EtOAc=7:1 to 5:1) to give methyl-3-O-benzoyl-2,5-di-
O-benzyl-a,b-l-xylofuranoside (3.60 g, 75.0%) as a white
oil. To a stirred solution of methyl-3-O-benzoyl-2,5-di-
O-benzyl-a,b- l-xylofuranoside (200 mg, 0.44 mmol) in
anhydrous methylene chloride (10 mL) was added tri-
ethylsilane (0.36 mL, 2.22 mmol) followed by slowly
adding trimethylsilyl trifluoromethansulfonate (0.44 mL,
2.22 mmol) at 0 ꢁC under nitrogen and the reaction mix-
ture was stirred for 3 h at ambient temperature under
nitrogen. The reaction mixture was quenched with satu-
rated NaHCO3 solution and extracted with methylene
chloride. The organic layer was washed with water,
dried with anhydrous MgSO4 and evaporated. The
residue was purified byflash silica gel column chroma-
tography(hexanes/EtOAc=5:1) to give 1,4-anhydro-3-
O-benzoyl-2,5-di-O-benzyl-l-xylitol (180 mg, 96.0%) as
trimethylsilyl
trifluoromethansulfonate
(2.62 mL,
14.50 mmol) at ambient temperature under nitrogen.
After the mixture was stirred for 2 h at ambient tem-
perature under nitrogen, it was poured into saturated
NaHCO3 solution and stirred for 30 min. The mixture
was extracted with methylene chloride and the organic
layer was washed with brine and water, dried with
anhydrous MgSO4 and evaporated. The residue was
purified byflash silica gel column chromatography
(hexanes/EtOAc=2:1) to give 10 (730 mg, 80.0%) as a
white oil.
1,4-Anhydro-2,5-di-O-benzyl-L-xylofurano-3-ulose (11).
To a stirred solution of 10 (250 mg, 0.8 mmol) in
DMSO (2 mL) was added Ac2O (2 mL, 22.3 mmol)
dropwise at ambient temperature under nitrogen and
the reaction mixture was stirred for 9 h at ambient tem-
perature under nitrogen. The reaction mixture was
quenched with saturated NaHCO3 solution and extrac-
ted with ether, dried with anhydrous MgSO4 and eva-
porated. The residue was purified byflash silica gel
column chromatography(hexanes/EtOAc=5:1) to give
11 (200 mg, 80.0%) as a white oil, which was subjected
to the next reaction.
1,4-Anhydro-2,5-di-O-benzyl-3-deoxy-3-C-methylene-L-
xylitol (12). Method A. To a stirred suspension of
1
a white oil: H NMR (CDCl3) d 3.76 (d, 2H, J=5.9 Hz,
5-H), 3.87 (dd, 1H, J=2.4, 10.0 Hz, 1-Ha), 4.16 (dd, 1H,
J=1.7, 5.3 Hz, 2-H), 4.26 (dd, 1H, J=5.4, 10.0 Hz, 1-
Hb), 4.46 (d, 1H, J=11.9 Hz, benzylic Hc), 4.43 (m, 1H,
4-H), 4.59 (d, 1H, J=11.9 Hz, benzylic Hd), 4.66 (d, 1H,
J=11.9 Hz, benzylic Hb), 4.84 (d, 1H, J=11.9 Hz,
benzylic Ha), 5.58 (d, 1H, J=3.7 Hz, 3-H), 8.00–7.18
(m, 15H, 3ꢂPh).
methyl
triphenylphosphonium
bromide
(1.27 g,
3.56 mmol) in dryTHF (10 mL) was added n-BuLi
(1.6 M solution in hexane, 2.62 mL, 4.19 mmol) at 0 ꢁC
under nitrogen and followed bydropwise addition of a
solution of 11 (654 mg, 2.09 mmol) in dryTHF (3 mL)
through cannula for 30 min at 0 ꢁC under nitrogen. The
mixture was stirred for 30 min at ambient temperature
under nitrogen. Ether was added to the mixture to pre-
cipitate out triphenylphosphine oxide and the mixture
was filtered off. The filtrate was evaporated and the
To a stirred solution of 1,4-anhydro-3-O-benzoyl-2,5-di-
O-benzyl-l-xylitol (170 mg, 0.41 mmol) in methanol

S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
221
residue was purified byflash silica gel column chroma-
tography(hexanes/EtOAc=5:1) to give 12 (200 mg,
32.0%) as a white oil: 1H NMR (CDCl3) d 3.53 (dd, 1H,
J=5.9, 10.4 Hz, 5-Ha), 3.59 (dd, 1H, J=3.8, 10.4 Hz, 5-
Hb), 3.82 (dd, 1H, J=4.3, 9.5 Hz, 1-Ha), 4.04 (dd, 1H,
J=5.2 Hz, J=9.5 Hz, 1-Hb), 4.37 (m, 1H, 2-H), 4.50 (d,
1H, J=11.9 Hz, benzylic Ha), 4.58 (s, 2H, benzylic Hc
and Hd), 4.63 (d, 1H, J=11.9 Hz, benzylic Hb), 4.68 (m,
1H, 4-H), 5.17 (m, 1H, vinylic Ha), 5.34 (m, 1H, vinylic
Hb), 7.22–7.34 (m, 10H, 2ꢂPh). Anal. calcd for
C20H22O3: C, 77.39; H, 7.14. Found: C, 77.79; H, 6.75.
Ha), 5.36 (m, 1H, vinylic Hb), 7.34–7.43 (m, 6H, Ph),
7.65–7.70 (m, 4H, Ph). Anal. calcd for C22H2O3: C,
71.70; H, 7.66. Found: C, 71.98; H, 7.35.
(2R,4R)-3-Benzoyl-1-(2-tert-butyldiphenylsilyloxymethyl
-3-methylene-tetrahydrofuran-4-yl)-1H-pyrimidine-2,4-
dione (15). To a stirred solution of 14 (250 mg,
0.70 mmol), N3-benzoyluracil (421 mg, 1.61 mmol), and
triphenyl phosphine (552 mg, 2.10 mmol) in dry THF
(11 mL) was added diethyl azodicarboxylate (0.33 mL,
2.10 mmol) dropwise at ꢀ10 ꢁC under nitrogen and the
reaction mixture was stirred for 1 h at ꢀ10 ꢁC under
nitrogen. The solvent was evaporated and the residue
was purified byflash silica gel column chromatography
(hexanes/EtOAc=7:2) to give 15 (270 mg, 63.0%) as a
white oil and O-glycosylated product (40 mg, 10.0%) as
Method B. To a stirred suspension of methyl triphenyl-
phosphonium bromide (2.19 g, 6.13 mmol) and t-amyl
alcohol (0.73 mL, 6.68 mmol) in dryTHF (18 mL) was
added NaH (160 mg, 60% in oil, 6.68 mmol) at 0 ꢁC
under nitrogen and the reaction mixture was stirred for
2 h at ambient temperature under nitrogen. To this yel-
low phosphrous ylide was added a solution of 11
(580 mg, 1.86 mmol) in dryTHF (3 mL) dropwise
through cannula at 0 ꢁC under nitrogen. After the mix-
ture was stirred for 30 min at ambient temperature
under nitrogen, the reaction mixture was quenched with
saturated NaHCO3 solution and extracted with EtOAC.
The organic layer was washed with water, dried with
anhydrous MgSO4 and evaporated. The residue was
purified byflash silica gel column chromatography
(hexanes/EtOAc=7:1) to give 12 (510 mg, 92.0%) as a
white oil.
1
a white oil: UV (MeOH) lmax 271 nm (pH 7); H NMR
(CDCl3) d 1.14 (s, 9 H, t-butyl), 4.21–3.94 (m, 4H, 50-H
and 10-H), 4.46 (m, 1H, 40-H), 5.39 (m, 1H, vinylic Ha),
5.47 (m, 1H, vinylic Hb), 5.49 (d, 1H, J=7.4 Hz, H-5),
5.67 (m, 1H, 20-H), 7.53 (d, 1H, J=7.4 Hz, H-6), 7.98–
7.45 (m, 15H, 3ꢂPh). Anal. calcd for C33H34N2O5Si: C,
69.94; H, 6.05; N, 4.94. Found: C, 70.34; H, 6.42; N,
4.89.
(2R,4R)-6-Chloro-9-(2-tert-butyldiphenylsilyloxymethyl-
3-methylene-tetrahydrofuran-4-yl)-9H-purine (16). The
intermediate 14 (40 mg, 0.11 mmol) was converted to
compound 16 (hexanes/EtOAc=3:1, 40 mg, 75.0%) as a
white oil under the same Mitsunobu conditions used in
the synthesis of compound 15: UV (MeOH) lmax
1,4-Anhydro-3-deoxy-3-C-methylene-L-xylitol (13). To a
stirred solution of 12 (200 mg, 0.67 mmol) in anhydrous
methylene chloride (10 mL) was added boron trichloride
(2.68 mL, 1 M solution in methylene chloride,
2.68 mmol) at ꢀ78 ꢁC under nitrogen and the reaction
mixture was stirred for 30 min at ꢀ78 ꢁC under nitro-
gen. The reaction mixture was quenched with methanol,
neutralized with pyridine, and evaporated to dryness.
The residue was purified byflash silica gel column
chromatography(chloroform/methanol=10:1) to give
1
264 nm (pH 7); H NMR (CDCl3) d 1.08 (s, 9H, tert-
butyl), 3.94 (d, 2H, J=4.7 Hz, 50-H), 4.23 (d, 2H,
J=5.4 Hz, 10-H), 4.59 (m, 1 H, 40-H), 5.33 (m, 2H,
vinylic H), 5.72 (m, 1H, 20-H), 7.35–7.47 (m, 6H, Ph),
7.60–7.71 (m, 4H, Ph), 8.25 (s, 1H, H-2), 8.73 (s, 1H, H-
8). Anal. calcd for C27H29ClN4O2Si: C, 64.21; H, 5.79;
N, 11.09. Found: C, 64.05; H, 5.99; N, 11.49.
(2R,4R)-2-Amino-6-chloro-9-(2-tert-butyldiphenylsilylox-
ymethyl-3-methylene-tetrahydrofuran-4-yl)-9H-purine
(17). The intermediate 14 (30 mg, 0.08 mmol) was con-
verted to compound 17 (hexanes/EtOAc=2:1, 20 mg,
50.0%) as a white oil under the same Mitsunobu con-
ditions used in the synthesis of compound 15: UV
1
13 (60 mg, 75.0%) as a white oil: H NMR (CDCl3) d
2.31 (br s, 2H, 2ꢂOH), 3.59–3.77 (m, 3H, 1-Ha and 5-
H), 4.14 (dd, 1H, J=5.4, 9.3 Hz, 1-Hb), 4.57–4.67 (m,
2H, 2-H and 4-H), 5.17 (m, 1H, vinylic Ha), 5.43 (m,
1H, vinylic Hb). Anal. calcd for C6H10O3: C, 55.37; H,
7.74; N. Found: C, 55.16; H, 7.86.
1
(MeOH) lmax 308 nm (pH 7); H NMR (CDCl3) d 1.12
(s, 9H, tert-butyl), 3.94 (m, 2H, 50-H), 4.16 (dd, 1H,
J=4.6, 9.9 Hz, 10-Ha), 4.23 (dd, 1H, J=6.2, 9.9 Hz, 10-
Hb), 4.62 (m, 1H, 40-H), 5.17 (br s, 2H, NH2), 5.27 (m,
1H, vinylic Ha), 5.32 (m, 1H, vinylic Hb), 5.52 (m, 1H,
20-H), 7.39–7.50 (m, 6H, Ph), 7.71–7.75 (m, 4H, Ph),
7.88 (s, 1H, H-8). Anal. calcd for C27H30ClN5O2Si: C,
62.35; H, 5.81; N, 13.47. Found: C, 62.76; H, 5.45; N,
13.21.
1,4-Anhydro-5-O-(tert-butyldiphenylsilyl)-3-deoxy-3-C-
methylene-L-xylitol (14). To a stirred solution of 13
(60 mg, 0.51 mmol) and imidazole (104 mg, 1.53 mmol)
in DMF (5 mL) was added t-butyldiphenylsilyl chloride
(0.13 mL, 0.61 mmol) dropwise at 0 ꢁC under nitrogen
and the reaction mixture was stirred for 1 h at 0 ꢁC
under nitrogen. The reaction mixture was quenched
with water at 0 ꢁC and extracted with EtOAC. The
organic layer was washed with water, dried with anhy-
drous MgSO4 and evaporated. The residue was purified
byflash silica gel column chromatography(hexanes/
EtOAc=2:1) to give 14 (120 mg, 66.0%) as a white oil:
1H NMR (CDCl3) d 1.04 (s, 9H, tert-butyl), 1.76 (d, 1H,
J=5.9 Hz, OH, D2O exchangeable), 3.64–3.72 (m, 3H,
1-Ha and 5-H), 4.12 (dd, 1H, J=5.4, 9.3 Hz, 1-Hb),
4.61–4.63 (m, 2H, 2-H and 4-H), 5.11 (m, 1H, vinylic
(2R,4R)-6-Chloro-9-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-9H-purine (18). To a stirred solution of
16 (100 mg, 0.20 mmol) in THF (5 mL) was added tetra
n-butylammonium fluoride (1 M solution in THF,
0.24 mL, 0.24 mmol) at 0 ꢁC under nitrogen and the
reaction mixture was stirred for 30 min at 0 ꢁC under
nitrogen. The solvent was evaporated and the residue
was purified byflash silica gel column chromatography

222
S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
(chloroform/methanol=10:1) to give 18 (50 mg, 96.0%)
as a white stickyoil: UV (MeOH) lmax 265 nm (pH 7); 1H
NMR (CDCl3) d 2.29 (d, 1H, J=4.3Hz, OH), 4.00 (dd,
1H, J=3.8, 12.2Hz, 50-Ha), 4.12 (br d, 1H, J=12.2 Hz, 50-
Hb), 4.34 (pseudo t, 2H, J=4.3, 10.3 Hz, 10-H), 4.68 (m,
1H, 40-H), 5.31 (m, 1H, vinylic Ha), 5.40 (m, 1H, vinylic
Hb), 5.68 (m, 1H, 20-H), 8.20 (s, 1H, H-2), 8.56 (s, 1H, H-
8). Anal. calcd for C11H12ClN4O2: C, 49.54; H, 4.16; N,
21.01. Found: C, 49.15; H, 4.48; N, 21.32.
product. To a stirred mixture of 1,2,4-triazole (108mg,
1.56mmol), phosphorus oxytrichloride (0.12 mL,
1.30mmol) in dryacetonitrile (2 mL) was added triethyl-
amine (0.18 mL, 1.30 mmol) followed bydropwise addi-
tion of a solution of crude acetylated product in
acetonitrile at ambient temperature under nitrogen and
the whole mixture was stirred overnight at ambient
temperature under nitrogen. The reaction mixture was
quenched with water and triethylamine and evaporated.
The residue was dissolved in methylene chloride and the
organic layer was washed with water, dried with anhy-
drous MgSO4 and evaporated to give the triazole inter-
mediate. This triazole intermediate was treated with
dioxane (1.16 mL) and ammonium hydroxide (0.39 mL)
and stirred overnight at ambient temperature. The
reaction mixture was evaporated and the residue was
dissolved in methanol (3 mL) followed bytreatment
with NaOMe (1.0 M solution in MeOH, 0.16 mL,
0.16 mmol). The mixture was stirred for 4 h at ambient
temperature under nitrogen. The reaction mixture was
neutralized with acetic acid and evaporated. The residue
was purified byflash silica gel column chromatography
(chloroform/methanol=8:1) to give 4b (35 mg, 100%)
as white solid which was crystallized from ether: MS
(FAB) m/z 224 (MH+); UV (H2O) lmax 273 nm (e 7170)
(pH 7); 282 nm (e 9800) (pH 2); 273 nm (e 6940) (pH
(2R,4R)-2-Amino-6-chloro-9-(2-hydroxymethyl-3-methyl-
ene-tetrahydrofuran-4-yl)-9H-purine (19). Compound 17
(80 mg, 0.15 mmol) was converted to compound 19
(chloroform/methanol=10:1, 40 mg, 93.0%) as a white
1
stickyoil: UV (MeOH) lmax 310 nm (pH 7); H NMR
(CD3OD) d 3.85 (dd, 1H, J=7.9, 12.3 Hz, 50-Ha), 3.91
(dd, 1H, J=9.1, 12.3 Hz, 50-Hb), 4.15 (dd, 1H, J=5.9,
10.1 Hz, 10-Ha), 4.25 (dd, 1H, J=3.2, 10.1 Hz, 10-Hb), 4.51
(m, 1H, 40-H), 5.37 (m, 2H, vinylic H), 5.57 (m, 1H, 20-H),
8.26 (s, 1H, H-8). Anal. calcd for C11H12ClN5O2: C, 46.90;
H, 4.29; N, 24.86. Found: C, 46.91; H, 4.48; N, 24.99.
(2R,4R)-1-(2-Hydroxymethyl-3-methylene-tetrahydro-
furan-4-yl)-1H-pyrimidine-2,4-dione (4a). To a stirred
solution of 15 (250 mg, 0.41 mmol) in methanol (10 mL)
was added NaOMe (1.0 M solution in MeOH, 0.49 mL,
0.49 mmol) dropwise at 0 ꢁC under nitrogen and the
reaction mixture was stirred for 2 h at 0 ꢁC under nitro-
gen. The reaction mixture was neutralized with acetic
acid and evaporated. The residue was dissolved in
methylene chloride, and the organic layer was washed
with water and brine, dried with anhydrous MgSO4 and
evaporated. To a stirred solution of this crude deben-
zoylated product in THF (6 mL) was added tetra n-
butylammonium fluoride (1.0 M solution in THF,
0.49 mL, 0.49 mmol) at 0 ꢁC under nitrogen and the
reaction mixture was stirred for 4 h at 0 ꢁC under nitro-
gen. The mixture was evaporated and the residue was
purified byflash silica gel column chromatography
(chloroform/methanol=10:1) to give 4a (80 mg, 75.0%)
as a white solid which was crystallꢁized from ether: MS
(FAB) m/z 225 (MH+); mp 176.5 C (methanol/ether);
UV (H2O) lmax 266 nm (e 12,490) (pH 7); 265 nm (e
12,090) (pH 2); 264 nm (e 9240) (pH 11); [a]2D5 ꢀ18.5ꢁ (c
0.13); 1H NMR (DMSO-d6) d 3.67 (dd, 1H, J=4.0,
12.0 Hz, 50-Ha), 3.73 (dd, 1H, J=2.4, 12.0 Hz, 50-Hb),
3.88 (dd, 1H, J=4.0, 9.6 Hz, 10-Ha), 3.98 (dd, 1H,
J=6.8, 9.6 Hz, 10-Hb), 4.32 (m, 1H, 40-H), 5.03 (br s,
1H, OH), 5.23 (m, 1H, vinylic Ha), 5.32 (m, 1H, vinylic
Hb), 5.45 (m, 1H, 20-H), 5.57 (d, 1H, J=7.6 Hz, H-5),
7.74 (d, 1H, J=7.6 Hz, H-6), 11.32 (br s, 1 H, NH); 13C
NMR (DMSO-d6) d 57.65, 63.28, 71.09, 81.88, 102.35,
111.64, 143.22, 148.07, 151.97, 163.87. Anal. calcd for
C10H12N2O4: C, 53.57; H, 5.39; N, 12.49. Found: C,
53.32; H, 5.68; N, 12.10.
11); [a]2D5 +9.2ꢁ (c 0.13); H NMR (DMSO-d6) d 3.66
1
(dd, 1H, J=4.4, 11.6 Hz, 50-Ha), 3.71 (dd, 1H, J=3.6,
11.6 Hz, 50-Hb), 3.78 (dd, 1H, J=4.4, 9.6 Hz, 10-Ha),
3.96 (dd, 1H, J=7.2, 9.6 Hz, 10-Hb), 4.32 (m, 1H, 40-H),
5.01 (br s, 1H, OH), 5.11 (m, 1H, vinylic Ha), 5.28 (m,
1H, vinylic Hb), 5.51 (m, 1H, 20-H), 5.68 (d, 1H,
J=7.6 Hz, H-5), 7.08 (br d, 2H, J=40.8 Hz, NH2), 7.58
(d, 1 H, J=7.6 Hz, H-6); 13C NMR (DMSO-d6) d 58.43,
63.43, 71.68, 82.04, 94.63, 110.89, 143.65, 149.04,
156.59, 166.05. Anal. calcd for C10H13N3O3: C, 53.80;
H, 5.87; N, 18.82. Found: C, 53.81; H, 5.93; N, 18.77.
(2R,4R)-6-Amino-9-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-9H-purine (4c).
A solution of 18
(50 mg, 0.20 mmol in methanolic ammonia (6 mL) was
stirred at 100 ꢁC for 2 days. The solvent was evaporated
and the residue was purified byflash silica gel column
chromatography(chloroform/methanol=10:1) to give
4c (33 mg, 70%) as a white solid which was crystallized
from ether: MS (FAB) m/z 248 (MH+); mp 251.5 ꢁC
(methanol/ether); UV (H2O) lmax 260 nm (e 17,430) (pH
7); 259 nm (ꢁe 17,050) (pH 2); 260 nm (e 17,120) (pH 11);
1
[a]2D5 +16.7 (c 0.12); H NMR (DMSO-d6) d 3.73 (m,
2H, 50-H), 4.11 (dd, 1H, J=6.0, 9.6 Hz, 10-Ha), 4.15 (dd,
1H, J=4.4, 9.6 Hz, 10-Hb), 4.44 (m, 1H, 40-H), 5.07 (t,
1H, J=5.6 Hz, OH), 5.15 (m, 1H, vinylic Ha), 5.30 (m,
1H, vinylic Hb), 5.56 (t, 1H, J=4.8 Hz, 20-H), 7.26 (br s,
2H, NH2), 8.15 (s, 1H, H-2), 8.16 (s, 1H, H-8); 13C NMR
(DMSO-d6) d 57.09, 63.56, 71.41, 82.08, 111.41, 119.22,
139.86, 148.43, 150.00, 153.10, 156.64. Anal. calcd for
C11H13N5O2: C, 53.43; H, 5.30; N, 28.32. Found: C,
53.83; H, 5.38; N, 28.11.
(2R,4R)-4-Amino-1-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-1H-pyrimidine-2-one (4b). To a stirred
solution of 4a (35 mg, 0.13 mmol) in pyridine (1 mL)
was added acetic anhydride (0.05 mL) at ambient tem-
perature under nitrogen and the reaction mixture was
stirred for 3 h at ambient temperature under nitrogen.
The solvent was evaporated to give the crude acetylated
(2R,4R)-9-(2-Hydroxymethyl-3-methylene-tetrahydro-
furan-4-yl)-1,9-dihydro-purine-6-one (4d). A solution of
18 (20 mg, 0.08 mmol) in 1 N sodium hydroxide solution
(2 mL) was refluxed for 1 h. The reaction mixture was

S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
223
neutralized with 1 N hydrochloride and evaporated. The
residue was dissolved in methanol and NaCl salt was
filtrated off. The filtrate was evaporated and the residue
was purified byflash silica gel column chromatography
(chloroform/methanol=8:1) to give 4d (17 mg, 90%) as
a white solid which was crystallized from ether: MS
(FAB) m/z 237 (MH+); mp 232.3 ꢁC (methanol/ether);
UV (H2O) lmax 248 nm (e 13,770) (pH 7); 247 nm (e
13,280) (pH 2); 253 nm (e 13,820) (pH 11); [a]2D5 ꢀ35.4ꢁ
(c 0.13); 1H NMR (DMSO-d6) d), 3.72 (d, 2H,
J=4.0 Hz, 50-H), 4.12 (pseudo t, 2H, J=4.0, 6.0 Hz, 10-
H), 4.42 (m, 1H, 40-H), 4.77 (t, 1H, J=3.6 Hz, OH), 5.18
(m, 1H, vinylic Hb), 5.32 (m, 1H, vinylic Ha), 5.54 (t,
1H, J=4.8 Hz, 20-H), 8.06 (s, 1H, H-2), 8.07 (s, 1H, H-
8), 11.52 (br s, 1H, NH); 13C NMR (DMSO-d6) d 57.41,
71.58, 82.10, 89.45, 111.77, 126.01, 129.75, 139.22,
146.36, 148.25, 157.37. Anal. calcd for C11H12N4O3: C,
53.22; H, 4.87; N, 22.57. Found: C, 52.98; H, 5.17; N,
22.18.
(10 mL) at ambient temperature under nitrogen: 1H
NMR (CDCl3) d 1.40 (s, 3H, CH3), 1.54 (s, 3H, CH3),
4.40 (dd, 1H, J=5.5, 11.9 Hz, 5-Ha), 4.55 (dd, 1H,
J=3.4, 11.9 Hz, 5-Hb), 4.96 (dd, 1H, J=0.9, 4.0 Hz, 2-
H), 5.08 (m, 1H, 4-H), 5.30 (m, 1H, vinylic Ha), 5.52 (m,
1H, vinylic Hb), 5.93 (d, 1H, J=4.0 Hz, 1-H), 7.41–8.06
(m, 5H, Ph). Anal. calcd for C16H18O5: C, 66.19; H,
6.25. Found: C, 66.38; H, 6.47.
5-O-Benzoyl-1-O-methyl-3-deoxy-3-C-methylene-D-xylo-
furanoside (24). To a stirred solution of 23 (2.9 g,
10.35 mmol) in anhydrous methanol (15 mL) was added
acetyl chloride (0.2 mL) equivalent to 1% HCl at ambi-
ent temprerature under nitrogen and the reaction mix-
ture was stirred for 6 h at ambient temperature under
nitrogen. The reaction mixture was neutralized with
pyridine and evaporated. The residue was purified by
flash silica gel column chromatography(hexanes/
EtOAc=2:1) to give 24 (2.5 g, 95%) as an oil: 1H NMR
(CDCl3) d 2.38 (br s, 1H, a and b isomer OH), 3.43 (s,
3H, b isomer OCH3), 3.49 (s, 3H, a isomer OCH3),
4.34–4.60 (m, 3H, b isomer 2-H, and a and b isomer 5-
H), 4.62 (m, a isomer 2-H), 4.86 (m, 1H, a isomer 4-H),
4.94 (s, 1H, b isomer 1-H), 5.01 (d, 1H, J=4.6 Hz, a
isomer 1-H), 5.06 (m, 1H, b isomer 4-H), 5.28 (m, 1H, a
isomer vinylic Ha), 5.36 (m, 1H, b isomer vinylic Ha),
5.46 (m, 1H, a isomer vinylic Hb), 5.56 (m, 1H, b isomer
vinylic Hb), 7.44–8.13 (m, 5H, a and b isomer Ph). Anal.
calcd for C14H16O5: C, 63.63; H, 6.10. Found: C, 63.33;
H, 6.50.
(2R,4R)-2-Amino-9-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-1,9-dihydro-purine-6-one (4e). A solu-
tion of 19 (40 mg, 0.14 mmol) in 1 N sodium hydroxide
solution (2 mL) was refluxed for 1 h. The reaction mix-
ture was neutralized with 1 N hydrochloride and eva-
porated. The residue was dissolved in methanol and
NaCl salt was filtrated off. The filtrate was evaporated
and the residue was purified byflash silica gel column
chromatography(chloroform/methanol=5:1) to give 4e
(25 mg, 70%) as a white solid which was crystallized
from ether: MS (FAB) m/z 264 (MH+); mp 269.4 ꢁC
(methanol/ether); UV (H2O) lmax 252 nm (e 10,770) (pH
7); 252 nm (e 10,460) (pH 2); 265 nm (e 8770) (pH 11);
[a]D25 ꢀ43.6ꢁ (c 0.11); 1H NMR (DMSO-d6) d 3.70
(pseudo t, 2H, J=5.6, 10.0 Hz, 50-H), 4.04 (pseudo t,
2H, J=2.8, 6.8 Hz, 10-H ), 4.39 (m, 1H, 40-H), 5.03 (t,
1H, J=5.6 Hz, OH), 5.15 (m, 1H, vinylic Hb), 5.29 (m,
2H, 20-H and vinylic Ha), 6.59 (br s, 2H, NH2), 7.69 (s,
1H, H-8), 10.67 (br s, 1H, NH); 13C NMR (DMSO-d6) d
56.64, 63.52, 71.58, 82.03, 111.38, 116.89, 136.08,
148.49, 151.77, 154.30, 157.39. Anal. calcd for
C11H13N5O3: C, 50.19; H, 4.98; N, 26.60. Found: C,
50.45; H, 4.76; N, 26.24.
1,4-Anhydro-5-O-benzoyl-3-deoxy-3-C-methylene-D-ery-
thro-pentitol (25). A mixture of 24 (3.6 g, 14.20 mmol)
and (NH4)2SO4 (0.2 g, 1.42 mmol) in HMDS (5 mL) was
refluxed for 2 h under nitrogen. The reaction mixture
was cooled to ambient temperature and HMDS was
evaporated with the exclusion of moisture. To a stirred
solution of the silylated residue in anhydrous methylene
chloride (15 mL) were added Et3SiH (11.3 mL,
70.96 mmol) and TMSOTf (12.8 mL, 70.96 mmol) at
ambient temperature under nitrogen and the reaction
mixture was stirred for 2 h at ambient temperature
under nitrogen and poured into saturated NaHCO3
solution with vigorouslystirring. The whole mixture
was extracted with methylene chloride and the organic
layer was washed with water, dried with MgSO4, and
evaporated. The residue was purified byflash silica gel
column chromatography(hexanes/EtOAc=1:1) to give
5-O-Benzoyl-3-deoxy-1,2-O-isopropylidene-3-C-methyl-
ene-ꢀ-D-xylofuranose (23). To a stirred solution of
methyl
triphenylphosphonium
bromide
(26.8 g,
80.16 mmol) and t-amyl alcohol (9.58 mL, 87.45 mmol)
in dryTHF (100 mL) was added NaH (60% in oil,
3.50 g, 87.45 mmol) at 0 ꢁC under nitrogen and the
reaction mixture was stirred for 2 h at ambient tem-
perature under nitrogen. To the yellow ylide was added
a solution of 21 (7.1 g, 24.29 mmol) in dryTHF (5 mL)
at 0 ꢁC under nitrogen and the mixture was stirred for
1 h at 0 ꢁC under nitrogen. The reaction mixture was
quenched with saturated NH4Cl solution and extracted
with EtOAc. The organic layer was washed with water,
dried with MgSO4, and evaporated. The residue was
purified byflash silica gel column chromatography
(hexanes/EtOAc=5:1) to give 22 (3.4 g, 79.4%) as an oil
and 23 (1.4 g, 20.7%) as an oil. Compound 22 was con-
verted to compound 23 in quantitative yield by treating
with benzoyl chloride (2.79 mL, 20.05 mmol) in pyridine
1
25 (2.5 g, 80%) as a white solid: H NMR (CDCl3) d
1.95 (d, J=5.0 Hz, OH), 3.75 (dd, 1H, J=5.2, 9.5 Hz, 1-
Ha), 4.22 (dd, 1H, J=5.7, 9.5 Hz, 1-Hb), 4.41 (dd, 1H,
J=6.0, 11.9 Hz, 5-Ha), 4.48 (dd, 1H, J=3.6, 9.5 Hz, 1-
Hb), 4.73 (m, 1H, 2-H), 4.72 (m, 1H, 4-H), 5.30 (m, 1H,
vinylic Ha), 5.49 (m, 1H, vinylic Hb), 7.45–8.10 (m, 5H,
Ph). Anal. calcd for C13H14O4: C, 66.66; H, 6.02.
Found: C, 66.34; H, 6.35.
(2S,4S)-3-Benzoyl-1-(2-benzoylmethyl-3-methylene-tetra-
hydrofuran-4-yl)-1H-pyrimidine-2,4-dione (26). To
a
stirred solution of 25 (300 mg, 1.35 mmol), N3-benzoyl-
uracil (705 mg, 2.70 mmol), and triphenyl phosphine
(1.06 g, 4.05 mmol) in dryTHF (12 mL) was added die-
thylazodicarboxylate (0.64 mL, 4.05 mmol) slowly at

224
S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
ꢀ10 ꢁC under nitrogen and the reaction mixture was
stirred for 1 h at ꢀ10 to 5 ꢁC under nitrogen. The sol-
vent was evaporated and the residue was purified by
flash silica gel column chromatography(hexanes/
EtOAc=3:2) to give 26 (505 mg, 80.5%) as a colorless
oil contaminated with impurities, which was subjected
to the next reaction.
chromatography(CHCl 3: MeOH=20:1) to give 29
(148 mg, 96%) as a stickyoil: UV (MeOH) lmax 266 nm;
1H NMR (CDCl3) d 2.29 (d, 1H, J=4.3 Hz, OH), 4.00
(dd, 1H, J=3.8, 12.2 Hz, 50-Ha), 4.12 (br d, 1H,
J=12.2Hz, 50-Hb), 4.34 (pseudo t, 2H, J=4.3, 10.3Hz, 10-
H), 4.68 (m, 1H, 40-H), 5.31 (m, 1H, vinylic Ha), 5.40 (m,
1H, vinylic Hb), 5.68 (m, 1H, 20-H), 8.20 (s, 1H, H-2), 8.56
(s, 1H, H-8). Anal. calcd for C11H12ClN4O2: C, 49.54; H,
4.16; N, 21.01. Found: C, 49.34; H, 4.56; N, 21.11.
(2S,4S)-1-(2-Hydroxymethyl-3-methylene-tetrahydrofuran-
4-yl)-1H-pyrimidine-2,4-dione (5a). To a stirred solution
of impure 26 (505 mg, 1.09 mmol) in MeOH (8 mL) was
added NaOMe (1 M solution in MeOH, 2.60 mL,
2.60 mmol) at 0 ꢁC under nitrogen and the reaction
mixture was stirred for 1 h at 0 ꢁC under nitrogen. The
reaction mixture was neutralized with acetic acid and
evaporated. The residue was purified byflash silica gel
(2S,4S)-2-Amino-6-chloro-9-(2-benzoylmethyl-3-methyl-
ene-tetrahydrofuran-4-yl)-9H-purine (30). The impure
28 (300 mg) was converted to compound 30 (chloro-
form/methanol=10:1, 187 mg, 51% from 25) using the
same conditions used in the synthesis of compound 29
1
as white stickyoil: UV (MeOH) lmax 309 nm; H NMR
column chromatography(CHCl /MeOH=10:1) to give
3
(CD3OD) d 3.85 (dd, 1H, J=7.9, 12.3 Hz, 50-Ha), 3.91
(dd, 1H, J=9.1, 12.3 Hz, 50-Hb), 4.15 (dd, 1H,
J=5.9 Hz, J=10.1 Hz, 10-Ha), 4.25 (dd, 1H, J=3.2,
10.1 Hz, 10-Hb), 4.51 (m, 1H, 40-H), 5.37 (m, 2H, vinylic
H), 5.57 (m, 1H, 20-H), 8.26 (s, 1H, H-8). Anal. calcd for
C11H12ClN5O2: C, 46.90; H, 4.29; N, 24.86. Found: C,
46.76; H, 4.31; N, 24.75.
5a (199 mg, 61.5%) as a white solid which was crystal-
lized from ether, whose spectral data was identical to
those of 4a except optical rotation: [a]2D5 +18.1ꢁ (c 0.11).
Anal. calcd for C10H12N2O4: C, 53.57; H, 5.39; N,
12.49. Found: C, 53.43; H, 5.79; N, 12.26.
(2S,4S)-4-Amino-1-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-1H-pyrimidine-2-one (5b). Compound
5a (55 mg, 0.20 mmol) was converted to compound 5b
(CHCl3/MeOH=8:1, 35 mg, 65%) as a white solid
which was crystallized from ether according to the same
procedure used in the synthesis of 4b, whose spectral
data was identical to those of 4b except optical rotation:
[a]2D5 ꢀ9.6ꢁ (c 0.12). Anal. calcd for C10H13N3O3: C,
53.80; H, 5.87; N, 18.82. Found: C, 54.12; H, 5.86; N,
17.98.
(2S,4S)-6-Amino-9-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-9H-purine (5c). Compound 29 (30 mg,
0.12 mmol) was converted to compound 5c (CHCl3/
MeOH=10:1, 20 mg, 70%) as a white solid which was
crystallized from ether according to the same procedure
used in the synthesis of 4c, whose spectral data was
identical to those of 4c except optical rotation: [a]D25
ꢀ15.9ꢁ (c 0.15). Anal. calcd for C11H13N5O2: C, 53.43;
H, 5.30; N, 28.32. Found: C, 53.22; H, 5.16; N, 28.76.
(2S,4S)-6-Chloro-9-(2-benzoylmethyl-3-methylene-tetra-
hydrofuran-4-yl)-9H-purine (27). The intermediate 25
(300 mg, 1.35 mmol) was converted to compound 27
(hexanes/EtOAc=1:1, 408 mg, 84.3%) as a white solid
under the same Mitsunobu conditions used in the
synthesis of compound 26: UV (MeOH) lmax 265 nm;
1H NMR (CDCl3) d 4.34 (dd, 1H, J=5.8, 10.4 Hz, 10-
Ha), 4.43 (dd, 1H, J=3.4, 10.4 Hz, 10-Hb), 4.71 (pseudo
t, 2H, J=1.8, 3.8 Hz, 50-H), 4.91 (m, 1H, 40-H), 5.50 (m,
2H, vinylic H), 5.82 (m, 1H, 20-H), 7.46–8.04 (m, 5H,
Ph), 8.33 (s, 1H, H-2), 8.79 (s, 1H, H-8). Anal. calcd for
C18H15ClN4O3: C, 58.31; H, 4.08; N, 15.11. Found: C,
58.32; H, 4.37; N, 15.10.
(2S,4S)-9-(2-hydroxymethyl-3-methylene-tetrahydrofuran-
4-yl)-1,9-dihydropurine-6-one (5d). Compound 27
(100 mg, 0.28 mmol) was refluxed with 1 N NaOH to give
5d (CHCl3: MeOH=8:1, 40 mg, 61%) as a white solid,
which was crystallized from ether, whose spectral data
was identical to those of 4d except optical rotation:
[a]2D5 +35.6ꢁ (c 0.14). Anal. calcd for C11H12N4O3: C,
53.22; H, 4.87; N, 22.57. Found: C, 53.62; H, 4.85; N,
22.44.
(2S,4S)-2-Amino-9-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-1,9-dihydropurine-6-one (5e). Com-
pound 30 (178 mg, 0.66 mmol) was converted to
compound 5e (CHCl3/MeOH=8:1, 139 mg, 79%) as a
white solid which was crystallized from ether according
to the same procedure used in the synthesis of 4e, whose
spectral data was identical to those of 4e except optical
rotation: [a]2D5 +44.8ꢁ (c 0.15). Anal. calcd for
C11H13N5O3: C, 50.19; H, 4.98; N, 26.60. Found: C,
50.59; H, 5.12; N, 26.85.
(2S,4S)-2-Acetamido-6-chloro-9-(2-benzoylmethyl-3-
methylene-tetrahydrofuran-4-yl)-9H-purine (28). The
intermediate 25 (300 mg, 1.35 mmol) was converted to
the impure 28 (chloroform/methanol=20:1, 300 mg)
under the same Mitsunobu conditions used in the
synthesis of compound 26.
(2S,4S)-6-Chloro-9-(2-hydroxymethyl-3-methylene-tetra-
hydrofuran-4-yl)-9H-purine (29). To a stirred solution
of 27 (220 mg, 0.61 mmol) in MeOH (8 mL) was added
NaOMe (1 M solution in MeOH, 0.74 mL, 0.74 mmol)
at 0 ꢁC under nitrogen and the reaction mixture was
stirred for 1 h at 0 ꢁC under nitrogen. The reaction mix-
ture was neutralized with acetic acid and evaporated.
The residue was purified byflash silica gel column
Antiviral assays
Anti-HCMV assay. Anti-HCMV activitywas measured
as published previously.20
Anti-HBV assay. Cell culture and chemical treatment.
The HepG2 2.2.15 cell line was cultured according to
Korba’s protocol21 with the following procedural

S. J. Yoo et al. / Bioorg. Med. Chem. 10 (2002) 215–226
225
modifications. The cell line was maintained in DMEM
Acknowledgements
medium (Gibco BRL #430-2200) containing 10% fetal
bovine serum (FBS, Gibco BRL #16000-028), 1%
ABAM and 200 ug/mL G418 (Sigma, #G-9516). Cells
This research was supported bythe grant from the
Korea Science and Engineering Foundation (KOSEF
981-0716-126-2).
22
were routinelychecked for resistance to G418.
For the anti-HBV assay, cells were seeded into 96-well
tissue culture plates at approximately1 ꢂ10(exp4)/well
and grown to confluence. When cells become confluent,
medium was changed with fresh one containing 2%
FBS. During the 10 daytreatment period, the culture
medium was replaced with fresh one containing test
chemicals at a 2 days interval. Immediately prior to the
first dose of test chemical (day0), and after 2, 4, 6, 8,
and 10 days of treatment, culture media were collected
and stored at ꢀ70 ꢁC for HBV DNA analysis.
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