Synthesis of novel 6′-spirocyclopropyl-5′-norcarbocyclic adenosine phosphonic acid analogues as potent anti-HIV agents

Article abstract of DOI:10.1080/15257770.2011.602656

Novel 5′-norcarbocyclic adenosine phosphonic acid analogues with 6′-electropositive moiety such as spirocyclopropane were designed and synthesized from the commercially available diethylmalonate 5. Regioselective Mitsunobu reaction proceeded in the presence of an allylic functional group at a low reaction temperature in polar cosolvent [dimethylformamide (DMF)/1,4-dioxane] to give purine analogue 15. To improve cellular permeability and enhance the anti-human immunodeficiency virus (HIV) activity of this phosphonic acid, a SATE phosphonodiester nucleoside prodrug 23 was prepared. The synthesized adenosine phosphonic acids analogues 18-21 and 23 were subjected to antiviral screening against HIV-1. Copyright Taylor and Francis Group, LLC.

Full text of DOI:10.1080/15257770.2011.602656

Nucleosides, Nucleotides and Nucleic Acids, 30:784–797, 2011
Copyright ꢀC Taylor and Francis Group, LLC
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770.2011.602656
SYNTHESIS OF NOVEL 6ꢀ-SPIROCYCLOPROPYL-5ꢀ-NORCARBOCYCLIC
ADENOSINE PHOSPHONIC ACID ANALOGUES AS POTENT
ANTI-HIV AGENTS
Lian Jin Liu, Eunae Kim, and Joon Hee Hong
BK-21 Project Team, College of Pharmacy, Chosun University, Kwangju, Republic of Korea
Novel 5ꢁ-norcarbocyclic adenosine phosphonic acid analogues with 6ꢁ-electropositive moiety such
2
as spirocyclopropane were designed and synthesized from the commercially available diethylmalonate
5. Regioselective Mitsunobu reaction proceeded in the presence of an allylic functional group at a
low reaction temperature in polar cosolvent [dimethylformamide (DMF)/1,4-dioxane] to give purine
analogue 15. To improve cellular permeability and enhance the anti-human immunodeficiency
virus (HIV) activity of this phosphonic acid, a SATE phosphonodiester nucleoside prodrug 23 was
prepared. The synthesized adenosine phosphonic acids analogues 18–21 and 23 were subjected to
antiviral screening against HIV-1.
Keywords Anti-HIV agents; 6ꢁ-spirocyclopropyl nucleosides; bis-SATE prodrug; mit-
sunobu reaction
INTRODUCTION
Much attention has been paid to unusual nucleosides since 6ꢁ-modified
nucleosides were reported to be promising anti-human immunodeficiency
virus (anti-HIV) and anti-hepatitis B virus (anti-HBV) agents. Among these
compounds, entecavir[1] (1) is being clinically used in anti-HBV drugs
(Figure 1). Carbovir analogue[2] 2 also exhibited potent anti-HIV activity,
but its cytotoxicity hindered it from being further developed as an antivi-
ral agent. Recently, an isonucleoside with an exomethylene[3] 3 or with a
spirocyclopane[4] 4 moiety in place of a carbon atom of a furanose ring
was reported to show antiviral activity, especially anti-HIV activity. Molecular
modeling studies demonstrated the presence of an electropositive moiety at
the 6ꢁ-position that could accommodate these substitutions and contribute
to the observed enhancement in potency in anti-HIV activity.[4] The spatial
location of the oxygen atom, namely the β-position from the phosphorus
atom in the nucleoside analogue, plays a critical role in antiviral activity.
Received 9 May 2011; accepted 30 June 2011.
Address correspondence to Dr. Joon Hee Hong, College of Pharmacy, Chosun University, Kwangju
501-759, Republic of Korea. E-mail: hongjh@chosun.ac.kr
784

6ꢁ-Spirocyclopropyl-5ꢁ-Norcarbocyclic Adenosine Analogues
785
O
O
N
N
N
N
N
NH
NH2
NH
NH2
HO
N
HO
HO
2
1
HO
NH2
N
NH2
N
N
N
N
N
N
OH
N
HO
4
O
O
3
FIGURE 1 Structures of nucleoside analogues as potent antiviral agents.
This increased antiviral activity due to this oxygen atom may be attributed
to the increased binding capacity of the phosphonate analogues to target
enzymes.[5] Moreover, a phosphonate nucleoside analogue can skip the req-
uisite first phosphorylation, which is a crucial step for the activation of nu-
cleosides. This is frequently a limiting step in the phosphorylation sequence,
which ultimately leads to triphosphates.[6] Though triphosphates of several
of the nucleoside analogues exhibit excellent RT inhibitory potency, only a
few nucleoside derivatives have exhibited biological activity in cell culture as-
says. This might be due to poor cellular penetration coupled with insufficient
metabolism of these nucleoside derivatives to 5ꢁ-triphosphates.
Stimulated by these findings that 6ꢁ-electropositive nucleoside analogues
and 5ꢁ-norcarbocyclic nucleoside phosphonate have excellent biological ac-
tivities, we sought to synthesize a novel class of nucleosides comprising 6ꢁ-
spirocyclopropyl-5ꢁ-norcarbocyclic phosphonic acid analogues in order to
search for more effective therapeutics against HIV and to provide analogues
for probing the conformational preferences of enzymes associated with the
nucleoside kinases of nucleosides and nucleotides.
As depicted in Scheme 1, the target compounds were prepared from
diethylmalonate 5.[7] Monosilylation of diol 6 and subsequent oxidation of
corresponding alcohol 7 gave aldehyde 8.
The aldehyde functional group of 8 was subjected to a carbonyl addi-
tion reaction by vinylmagnesium bromide to furnish the secondary alcohol 9,
which was successfully protected using p-methoxybenzyl chloride (PMBCl)[8]
to provide compound 10. Removal of the silyl protecting group of 10 using
t-butylammonium fluoride (TBAF) gave the primary alcohol 11, which was
oxidized to the aldehyde 12 using Swern oxidation conditions[9] [dimethyl

786
L. J. Liu et al.
ref.7
EtO
OEt
i
OH
HO
OP
HO
80%
O
O
6
5
7
P = TBDMS
Diethyl malonate
81%
ii
OP
iii
OP
HO
PMBO
iv
72%
OP
O
73%
8
10
9
v
91%
O
iii
OH
OH
PMBO
13
PMBO
PMBO
ii
90%
74%
12
11
Reagents: i) TBDMSCl, imidazole, DMF; ii) (COCl)2, DMSO, TEA; iii)
vinylMgBr, THF; iv) PMBCl, NaH, DMF; v) TBAF, THF.
SCHEME 1 Synthesis of cyclopropyl divinyl intermediate 13.
sulfoxide (DMSO), oxalyl chloride, TEA]. The aldehyde 12 was subjected
to nucleophilic Grignard conditions[10] with vinylmagnesium bromide to
give divinyl 13, which was subjected to ring-closing metathesis (RCM) condi-
tions using a 2nd generation Grubbs catalyst (C46H65Cl2N2PRu)[11] to pro-
vide spirocyclopropyl cyclopentenol 14a (40%) and 14b (39%), which were
readily separated by silica gel column chromatography. The nuclear Over-
hauser enhancement (NOE) experiments with cyclopentenols 14a and 14b
confirmed these assignments. As expected, NOEs were found between the
cis-oriented hydrogens. Upon irradiation of C4-H, weak NOE patterns were
observed at the proximal hydrogens of compound 14b [C7-CH- (1.5%)]
versus those of compound 14a [C7-CH- (2.1%)] (Figure 2).
To synthesize the desired 5ꢁ-norcarbocyclic adenosine nucleoside ana-
logues, the protected cyclopentenol 14b was treated with 6-chloropurine
H
OH
PMBO
PMBO
H
1.5%
4
H
7
OH
H
2.1%
a
b
FIGURE 2 NOE differences between the proximal hydrogens.

6ꢁ-Spirocyclopropyl-5ꢁ-Norcarbocyclic Adenosine Analogues
787
under Mitsunobu conditions[12] [diethyl azodicarboxylate (DEAD) and
PPh3]. An appropriate choice of the solvent system, temperature, and pro-
cedure is essential for the regioselectivity as well as for the yield. In purine
synthesis, a mixture of dioxane and DMF was used as the solvent for the cou-
pling of the cyclopentenol 14b with 6-chloropurine, instead of tetrahydrofu-
ran (THF). The heterocyclic bases had better solubility in the dioxane-DMF
mixture, resulting in better yields. The slow addition of diethyl azodicar-
boxylate (DEAD) to a mixture of cyclopentenol 14b, triphenylphosphine,
and the 6-chloropurine in anhydrous cosolvent (dioxane-DMF) gave a yel-
low solution, which was stirred for 1.5 hours at −40◦C and further stirred
overnight at room temperature (rt) to give the protected 6-chloropurine
analogue 15 as N 9-regioisomer [UV (MeOH) λmax 265.0 nm].[13] The
PMB protection group was removed, along with 2,3-dichloro-5,6-dicyano-
p-benzoquinone (DDQ),[14] to produce the 5ꢁ-nornucleoside analogue 16,
which was treated with diethylphosphonomethyl triflate[15] using lithium
t-butoxide to yield the nucleoside phosphonate analogue 17 (Scheme 2).
The chlorine group of 17 was then converted to amine with methanolic
ammonia at 70◦C to give the corresponding adenine phosphonate deriva-
tive 18. Hydrolysis of 18 by treatment with bromotrimethylsilane in CH3CN
in the presence of 2,6-lutidine gave an adenine phosphonic acid derivative
19 (Scheme 2).[16]
In order to synthesize the 2ꢁ,3ꢁ-dihydroxy nucleoside analogues, the pro-
tected nucleoside 18 was subjected to vicinal hydroxylation conditions[17]
using a catalytic amount of OsO4 and NMO to give 20 (33%) and 20ꢁ (31%),
respectively.[18] As shown in Figure 3, the stereochemistry was readily de-
termined by the NOE experiment. Upon irradiation of C4-H, a relatively
strong NOE was observed at C5-H and C6-H of 20ꢁ, which showed 1ꢁ,2ꢁ,3ꢁ-cis
relationships. But a relatively weak NOE was observed at C5-H and C6-H of
20, which indicates 1ꢁ,2ꢁ- and 1ꢁ,3ꢁ-trans relationships. The adenosine phos-
phonic acid 21 was synthesized from 20 by similar procedures described for
19 (Scheme 3).
To synthesize the thioester prodrug analogue, compound 19 was reacted
with thioester 22[19] in the presence of 1-(2-mesitylenesulfonyl)-3-nitro-1H-
1,2,4-triazole (MSNT)[20] to provide the bis(SATE) derivative as a target
compound 23 (Scheme 4).
The synthesized nucleoside phosphonate and phosphonic acid ana-
logues 18–21 and 23 were then evaluated for antiviral activity against HIV.
The procedures for measuring the antiviral activity toward wild-type HIV and
cytotoxicity have been reported previously.[21] As shown in Table 1, nucleo-
side phosphonic acid 23 exhibited increased anti-HIV activity compared with
its parent nucleoside phosphonic acid 19. However, nucleoside analogues
18, 20, and 21 did not show anti-HIV activity or cytotoxicity at concentrations
up to 100 µM.

788
L. J. Liu et al.
Cl
N
i
ii
X
Y
PMBO
N
13
N
PMBO
36%
N
14a: X = OH, Y = H (40%)
14b: X = H, Y = OH (39%)
15
iii
70%
Cl
Cl
N
N
O
iv
N
N
N
N
HO
EtO P
EtO
O
49%
N
N
16
17
v
55%
NH2
NH2
N
N
N
O
O
N
vi
N
N
EtO P
EtO
O
HO P
HO
O
79%
N
N
19
18
Reagents: i) Grubbs (II), CH2Cl2; ii) 6-chloropurine, DEAD, PPh3,
1,4-dioxane/DMF; iii) DDQ, CH2Cl2/H2O (10:1); iv) (EtO)2POCH2OTf,
LiO-t-Bu, THF; v) NH3/MeOH, 70 oC; vi) TMSBr, 2,6-lutidine, CH3CN;
SCHEME 2 Synthesis of 6ꢁ-spirocyclopropyl cyclopentenyl adenine phosphonic acid.
In summary, on the basis of the potent anti-HIV activity of 6ꢁ-
electropositive nucleosides and 5ꢁ-norcarbocyclic nucleoside analogues, we
have designed and successfully synthesized novel 6ꢁ-spirocyclopropyl-5ꢁ-
norcarbocyclic nucleoside analogues starting from the commercially avail-
able diethylmalonate 5.
O
O
EtO P
EtO P
A
EtO O
A
EtO O
4
4
7
7
H
H
H
H
5
6
5
6
H
H
OH
HO
HO OH
H
H
20'
20
FIGURE 3 NOE relationships between the proximal hydrogens.

6ꢁ-Spirocyclopropyl-5ꢁ-Norcarbocyclic Adenosine Analogues
NH2
789
N
O
EtO P
EtO
N
i
N
O
18
N
HO
OH
ii
20 (33%)
64%
and
NH2
NH2
N
N
N
O
HO P
HO
O
EtO P
EtO
N
N
O
N
O
N
N
HO
OH
HO
OH
20' (31%)
21
Reagents: i) OsO4, NMO, acetone/t-BuOH/H2O (6:1:1);
ii) TMSBr, 2,6-lutidine, CH3CN;
SCHEME 3 Synthesis of 6ꢁ-cyclopropyl-2ꢁ,3ꢁ-dihydroxy-cyclopentanyl adenine phosphonic acid.
The synthesized nucleoside prodrug 23 exhibited slight improvement in
cell-based activity compared with phosphonic acid 19. Although the SATE
protecting group as a prodrug scaffold was introduced, the antiviral activity
was slightly increased.
EXPERIMENTAL
Melting points were determined on a Mel-temp II laboratory device
and are uncorrected. NMR spectra were recorded on a JEOL 300 Fourier
NH2
NH2
N
O
O
HO P
N
N
i
O
N
N
N
S
36%
O P
HO O
N
O
N
O O
O
S
OH
S
22
19
23
Reagents: i) thioester, 22, 1-(2-mesitylenesulfonyl)-3-nitro-1H-1,2,4-triazole,
pyridine.
SCHEME 4 Synthesis of the target bis(SATE) prodrug of adenine analogue 23.

790
L. J. Liu et al.
TABLE 1 Anti-HIV activity of synthesized compounds
Compound no.
Anti-HIV EC50 (µM)c
Cytotoxicity CC50 (µM)d
18
19
20
21
80.0
19.1
88.9
90
12.3
0.01
0.51
98
80
100
100
50
23
AZTa
PMEAb
100
10
aAZT: azidothymidine.
bPMEA: 9-[2-(phosphonomethoxy)ethyl]adenine.
cEC50(µM): Concentration (µM) required to inhibit the replication of HIV-1 by 50%.
dCC50(µM): Concentration (µM) required to reduce the viability of unaffected cells by 50%.
transform spectrometer (JEOL, Tokyo, Japan); chemical shifts are reported
in parts per million (δ) and signals are reported as s (singlet), d (doublet), t
(triplet), q (quartet), m (multiplet), and dd (doublet of doublets). UV spec-
tra were obtained on a Beckman DU-7 spectrophotometer (Beckman, South
Pasadena, CA, USA). MS spectra were collected in electrospray ionization
(ESI) mode. Elemental analyses were performed using a Perkin–Elmer 2400
analyzer (Perkin–Elmer, Norwalk, CT, USA). TLC was performed on Uni-
plates (silica gel), purchased from Analtech Co. (7558, Newark, DE, USA).
All reactions were carried out in an atmosphere of nitrogen unless otherwise
specified. Dry dichloromethane, benzene, and pyridine were obtained by
distillation from CaH2. Dry THF was obtained by distillation from Na and
benzophenone immediately prior to use.
[1-(t-Butyldimethylsilyloxymethyl) cyclopropyl]methanol (7). t-Butyldi-
methylsilyl chloride (743 mg, 4.93 mmol) was added to a stirred solution
of compound 6 (480 mg, 4.7 mmol) and imidazole (477 mg, 7.02 mmol)
in CH2Cl2 (40 mL) at 0◦C. The mixture was stirred at rt for 3 hours, and
quenched by adding a NaHCO3 solution (5 mL). The mixture was extracted
using an EtOAc/water system, dried over MgSO4, filtered, and then con-
centrated. The residue was purified by silica gel column chromatography
(EtOAc/hexane, 1:5) to give compound 7 (813 mg, 80%) as a colorless
syrup: 1H NMR (CDCl3, 300 MHz) δ 3.54 (s, 2H), 3.49 (s, 2H), 0.84 (s, 9H),
0.44 (d, J = 3.9 Hz, 2H), 0.39 (d, J = 3.9 Hz, 2H), 0.02 (s, 6H); 13C NMR
(CDCl3) δ 76.6, 66.1, 25.8, 24.7, 18.2, 13.7, −5.5; MS m/z 217 (M+H)+.
˚
1-(t-Butyldimethylsilanyloxymethyl) cyclopropanecarbaldehyde (8). 4A
molecular sieves (1.9 g) and PCC (1.95 g, 9.06 mmol) were added slowly
to a solution of compound 7 (779 mg, 3.6 mmol) in CH2Cl2 (40 mL) at 0◦C,
and the mixture was stirred at rt for 10 hours. An excess of diethyl ether
(120 mL) was then added to the mixture. The mixture was stirred vigorously
at rt for 2 hours, and the resulting solid was filtered through a short silica
gel column. The filtrate was concentrated under vacuum and purified by

6ꢁ-Spirocyclopropyl-5ꢁ-Norcarbocyclic Adenosine Analogues
791
silica gel column chromatography (EtOAc/hexane, 1:35) to give compound
1
8 (617 mg, 81%) as a colorless oil: H NMR (CDCl3, 300 MHz) δ 9.58 (s,
1H), 4.06 (s, 2H), 0.82 (s, 9H), 0.75 (d, J = 3.8 Hz, 2H), 0.58 (d, J = 3.8 Hz,
2H), 0.01 (s, 6H); 13C NMR (CDCl3, 75 MHz) δ 204.2, 71.5, 40.6, 25.3, 18.5,
13.5, −5.3; MS m/z 215 (M+H)+.
( )-1-[1-(t-Butyldimethylsilanyloxymethyl)cyclopropyl]prop-2-en-1-ol
(9). To a solution of 8 (1.64 g, 7.66 mmol) in dry THF (40 mL), vinyl-
magnesium bromide (9.2 mL, 1.0 M solution in THF) was slowly added
at −20◦C and stirred at 0◦C for 4 hours. A saturated NH4Cl solution (10
mL) was added to the mixture, which was slowly warmed to rt. The mixture
was diluted with water (80 mL) and extracted with EtOAc (2 × 80 mL).
The combined organic layer was washed with brine, dried over anhydrous
MgSO4, filtered, and evaporated in vacuo. The residue was purified by silica
gel column chromatography (EtOAc/hexane, 1:12) to give 9 (1.35 g, 73%)
as a colorless oil: 1H NMR (CDCl3, 300 MHz) δ 5.90 (m, 1H), 5.27–5.18 (m,
2H), 3.89 (d, J = 4.2 Hz, 1H), 3.73 (m, 2H), 0.82 (s, 9H), 0.72–0.67 (m,
2H), 0.44–0.36 (m, 2H), 0.02 (s, 6H); 13C NMR (CDCl3, 75 MHz) δ 141.8,
114.5, 82.7, 73.4, 30.3, 25.4, 9.2, –5.5; MS m/z 243 (M+H)+.
( )-t-Butyl-{1-[1-(4-methoxybenzyloxy)allyl]cyclopropylmethoxy}
dimethylsilane (10). NaH (60% in mineral oil, 264 mg, 6.64 mmol)
was added portion-wise to a cooled (0◦C) solution of secondary alcohol 9
(1.34 g, 5.54 mmol) and p-methoxybenzyl chloride (0.82 mL, 6.09 mmol)
in anhydrous DMF (20 mL). The reaction mixture was stirred overnight
at rt. The solvent was removed in vacuo and the residue was diluted with
H2O (100 mL), followed by extraction with diethyl ether (2 × 80 mL).
The combined organic layer was washed with brine, dried over anhydrous
MgSO4, and concentrated under vacuum. The residue was purified by silica
gel column chromatography (EtOAc/hexane, 1:15) to give 10 (1.47 g,
1
72%) as a colorless oil. H NMR (CDCl3, 300 MHz) δ 7.29–7.23 (m, 2H),
6.92–6.86 (m, 2H), 5.88 (m, 1H), 5.29–5.23 (m, 2H), 4.50 (s, 2H), 3.79 (s,
3H), 3.64–3.52 (m, 3H), 0.83 (s, 9H), 0.74-0.69 (m, 2H), 0.48–0.40 (m, 2H),
0.01 (s, 6H); 13C NMR (CDCl3, 75 MHz) δ 159.6, 143.4, 131.7, 128.7, 117.5,
114.5, 86.7, 77.4, 75.2, 55.9, 25.6, 24.2, 18.5, 10.5, 9.9, –5.3; MS m/z 363
(M+H)+.
( )-–1-[1-(4-Methoxybenzyloxy)allyl]cyclopropyl˝methanol (11). To a
solution of 10 (1.2 g, 3.31 mmol) in THF (15 mL), TBAF (4.96 mL, 1.0 M
solution in THF) was added at 0◦C. The mixture was stirred overnight at rt
and concentrated in vacuo. The residue was purified by silica gel column
chromatography (Hexane/EtOAc, 7:1) to give 11 (747.9 mg, 91%): 1H NMR
(CDCl3, 300 MHz) δ 7.33–7.28 (m, 2H), 6.96–6.87 (m, 2H), 5.91 (m, 1H),
5.28–5.21 (m, 2H), 4.61 (s, 2H), 3.76 (s, 3H), 3.55–3.47 (m, 3H), 0.70-0.64
(m, 2H), 0.49–0.41 (m, 2H); 13C NMR (CDCl3, 75 MHz) δ 159.5, 143.0,
131.5, 129.4, 118.6, 114.3, 87.1, 74.8, 70.8, 57.1, 25.5, 10.0, 9.4; MS m/z 249
(M+H)+.

792
L. J. Liu et al.
( )-1-[1-(4-Methoxybenzyloxy)allyl]cyclopropanecarbaldehyde (12). To
a stirred solution of oxalyl chloride (258 mg, 2.04 mmol) in CH2Cl2 (17 mL)
was added a solution of DMSO (318 mg, 4.08 mmol) in CH2Cl2 (6.0 mL)
dropwise at –78◦C. The resulting solution was stirred at –78◦C for 30 min-
utes, and a solution of alcohol 11 (253 mg, 1.02 mmol) in CH2Cl2 (12 mL)
was added dropwise. The mixture was stirred at –78◦C for 30 min and TEA
(1.14 mL, 8.16 mmol) was added. The resulting mixture was warmed to 0◦C
and stirred for 30 minutes. H2O (15 mL) was added, and the solution was
stirred for 30 min at rt. The mixture was diluted with water (150 mL) and
then extracted with EtOAc (2 × 150 mL). The combined organic layer was
washed with brine, dried over anhydrous MgSO4 and filtered. The filtrate
was concentrated in vacuo and the residue was purified by silica gel col-
umn chromatography (EtOAc/hexane, 1:20) to give aldehyde compound
1
12 (226 mg, 90%) as a colorless oil: H NMR (CDCl3, 300 MHz) δ 9.68 (s,
1H), 7.29–7.22 (m, 2H), 6.91–6.86 (m, 2H), 5.89 (m, 1H), 5.26-5.19 (m,
2H), 4.69 (s, 2H), 3.75 (s, 3H), 3.71 (d, J = 4.0 Hz, 1H), 0.73–0.67 (m,
2H), 0.49-0.41 (m, 2H); 13C NMR (CDCl3, 75 MHz) δ 203.1, 159.6, 143.5,
132.0, 128.2, 117.8, 114.5, 83.2, 74.7, 56.9, 39.3, 10.7, 10.2; MS m/z 247
(M+H)+.
(rel)-(3R and 3S,5S)- 5-(4-Methoxybenzyloxy)-4-cyclopropyl-hepta-1,6-
dien-3-ol (13). Divinyl analogue 13 was synthesized as a diastereomeric mix-
ture from aldehyde 12 by a procedure similar to that described for 8 as a
diastereomeric mixture: yield 74%; 1H NMR (CDCl3, 300MHz) δ 7.32–7.24
(m, 2H), 6.92–6.83 (m, 2H), 5.93–5.85 (m, 2H), 5.23–5.15 (m, 4H), 4.65
(s, 2H), 3.89 (d, J = 4.1 Hz, 1H), 3.76–3.68 (m, 4H), 0.75–0.65 (m, 2H),
0.48–0.37 (m, 2H).
(rel)-(4S,7S)-7-(4-Methoxybenzyloxy)-spiro[2.4]hept-5-en-4-ol (14a) and
(rel)-(4R,7S)-7-(4-methoxybenzyloxy)-spiro[2.4]hept-5-en-4-ol (14b). To a so-
lution of 13 (296 mg, 1.08 mmol) in dry methylene chloride (7 mL) was
added a 2nd generation Grubbs catalyst (40.0 mg, 0.0471 mmol). The re-
action mixture was refluxed overnight and cooled to rt. The mixture was
concentrated in vacuo, and the residue was purified by silica gel column
chromatography (EtOAc/hexane, 1:10) to give cyclopentenol 14a (106 mg,
40%) and 14b (103 mg, 39%). Data for 14a: 1H NMR (CDCl3, 300 MHz) δ
7.31–7.26 (m, 2H), 6.89–6.83 (m, 2H), 5.66 (dd, J = 2.8, 5.3 Hz, 1H), 5.38
(m, 1H), 4.65 (s, 2H), 4.02 (d, J = 2.9 Hz, 1H), 3.77 (s, 3H), 3.62 (d, J = 3.1
Hz, 1H), 2.20 (dd, J = 13.6. 8.2 Hz, 1H), 0.77–0.71 (m, 2H), 0.51–0.47 (m,
2H); 13C NMR (CDCl3, 75 MHz) δ 159.6, 138.1, 137.0, 132.2, 127.6, 118.4,
87.4, 82.6, 74.5, 57.7, 24.7, 11.5, 9.9; MS m/z 247 (M+H)+.
Data for 14b: 1H NMR (CDCl3, 300 MHz) δ 7.29–7.21 (m, 2H), 6.88–6.83
(m, 2H), 5.66 (dd, J = 2.8, 5.4 Hz, 1H), 5.34 (m, 1H), 4.68 (s, 2H), 4.00 (dd,
J = 4.2, 2.6 Hz, 1H), 3.73-3.65 (m, 4H), 0.78–0.73 (m, 2H), 0.49–0.44 (m,
2H); 13C NMR (CDCl3, 75 MHz) δ 159.8, 138.7, 136.1, 134.5, 129.8, 118.4,
88.2, 73.8, 27.5, 8.8, 7.7; MS m/z 247 (M+H)+.

6ꢁ-Spirocyclopropyl-5ꢁ-Norcarbocyclic Adenosine Analogues
793
(rel)-(4ꢁR,7ꢁS)-9-[7-(4-Methoxybenzyloxy)-spiro[2.4]hept-5-en-4-yl]
6-
chloropurine (15). To a solution containing compound 14b (183 mg, 0.744
mmol), triphenylphosphine (528 mg, 2.016 mmol) and 6-chloropurine
(229 mg, 1.488 mmol) in anhydrous cosolvent (1,4-dioxane, 8.0 mL and
DMF, 6.0 mL), DEAD (0.271 mL, 1.488 mmol) was added dropwise at
–40◦C for 10 minutes under nitrogen. The reaction mixture was stirred
at the same temperature for 1.5 hours under nitrogen and further stirred
overnight at rt. The solvent was concentrated in vacuo and the residue was
purified by silica gel column chromatography (EtOAc/hexane, 3:1) to give
compound 15 (102 mg, 36%): mp 162–164◦C; UV (MeOH) λmax 265.0 nm:
1H NMR (CDCl3, 300 MHz) δ 8.73 (s, 1H), 8.36 (s, 1H), 7.20–7.24 (m, 2H),
6.93–6.86 (m, 2H), 5.67 (d, J = 5.4 Hz, 1H), 5.34 (dd, J = 3.2, 5.5 Hz, 1H),
4.66 (s, 2H), 4.45 (d, J = 3.3 Hz, 1H), 3.74 (s, 3H), 0.78–0.72 (m, 2H),
0.48–0.43 (m, 2H); 13C NMR (CDCl3, 75 MHz) δ 159.7, 151.4, 149.9, 143.8,
137.5, 135.7, 132.3, 131.5, 128.5, 118.4, 89.5, 75.1, 67.7, 56.3, 21.5, 8.6, 8.0;
MS m/z 383 (M+H)+.
(rel)-(4ꢁR,7ꢁS)-[9-(7-Hydroxy)-spiro[2.4]hept-5-en-4-yl] 6-chloropurine
(16). To a solution of compound 15 (324 mg, 0.846 mmol) in CH2Cl2/H2O
(10 mL, 10:1 v/v) was added DDQ (286 mg, 1.264 mmol), and the mixture
was stirred overnight at room temperature. Saturated NaHCO3 (1.7 mL)
was added to quench the reaction, which was then stirred at rt for 2 hours.
The mixture was diluted with water (150 mL) and extracted with CH2Cl2 (3
× 150 mL). The combined organic layer was dried over anhydrous MgSO4
and filtered. The filtrate was concentrated in vacuo and the residue was
purified by silica gel column chromatography (EtOAc/hexane/MeOH,
3:1:0.05) to give compound 16 (155 mg, 70%): mp 179–181◦C; UV (MeOH)
λmax 264.0 nm; 1H NMR (DMSO-d6, 300 MHz) δ 8.67 (s, 1H), 8.29 (s, 1H),
5.62 (d, J = 5.5 Hz, 1H), 5.34 (dd, J = 5.6, 3.9 Hz, 1H), 4.50 (d, J = 5.4 Hz,
1H), 3.98 (d, J = 5.2 Hz, 1H), 0.39–0.29 (m, 4H); 13C NMR (DMSO-d6,
75 MHz) δ 152.0, 151.5, 149.9, 145.4, 137.5, 135.2, 132.4, 83.8, 67.4, 21.9,
8.5, 7.8; MS m/z 263 (M+H)+.
(rel)-(4ꢁR,7ꢁS)-Diethyl [9-(7-hydroxymethyl)-spiro[2.4]hept-5-en-4-yl) 6-
chloropurine] phosphonate (17). Both LiOt-Bu (2.98 mL of 0.5 M solution
in THF, 1.488 mmol) and a solution of diethyl phosphonomethyltriflate
(417 mg, 1.392 mmol) in 11.0 mL of THF were slowly added to a solution
of 6-chloropurine analogue 16 (183 mg, 0.696 mmol) in 9.0 mL of THF at
–25◦C and stirred overnight at rt under nitrogen. The mixture was quenched
by adding a saturated NH4Cl solution (7 mL) and further diluted with ad-
ditional H2O (150 mL). The aqueous layer was extracted with EtOAc (3 ×
150 mL). The combined organic layer was dried over anhydrous MgSO4 and
concentrated in vacuo. The residue was purified by silica gel column chro-
matography (MeOH/Hexane/EtOAc, 0.04:3:1) to give 17 (140 mg, 49%) as
a foam: 1H NMR (DMSO-d6, 300 MHz) δ 8.78 (s, 1H), 8.49 (s, 1H), 5.63 (d,
J = 5.4 Hz, 1H), 5.38 (dd, J = 5.3, 4.2 Hz, 1H), 4.48 (d, J = 4.2 Hz, 1H),

794
L. J. Liu et al.
4.32 (m, 4H), 3.94 (d, J = 8.1 Hz, 2H), 3.67 (d, J = 5.1 Hz, 1H), 1.38 (m
6H), 0.41–0.32 (m, 4H); 13C NMR (DMSO-d6, 75 MHz) δ 151.7, 151.0, 150.2,
146.5, 137.8, 134.2, 132.2, 90.4, 69.1, 65.4, 64.6, 59.2, 22.1, 14.7, 9.1, 8.3; MS
m/z 413 (M+H)+.
(rel)-(4ꢁR,7ꢁS)-Diethyl
[9-(7-hydroxymethyl)spiro[2.4]hept-5-en-4-yl)-
adenine]-phosphonate (18). A solution of 17 (180 mg, 0.436 mmol) in
saturated methanolic ammonia (10 mL) was stirred overnight at 65◦C in a
steel bomb, and the volatiles were evaporated. The residue was purified by
silica gel column chromatography (MeOH/CH2Cl2, 1:7) to give 18 (94 mg,
55%) as a white solid: mp 144–146◦C; UV (MeOH) λmax 261.0 nm; 1H NMR
(DMSO-d6, 300 MHz) δ 8.28 (s, 1H), 8.13 (s, 1H), 5.66 (d, J = 5.4 Hz,
1H), 5.37 (m, 1H), 4.43 (d, J = 5.2 Hz, 1H), 4.35–4.30 (m, 4H), 4.00 (d,
J = 8.2 Hz, 2H), 3.59 (d, J = 5.0 Hz, 1H), 1.41–1.37 (m 6H), 0.41–0.36
(m, 4H); 13C NMR (DMSO-d6, 75 MHz) δ 154.6, 152.7, 150.1, 142.2, 137.3,
132.9, 119.4, 90.5, 67.7, 64.6, 63.7, 57.7, 21.5, 15.2, 8.0, 7.2; Anal. Calc. for
C17H24N5O4P (+0.5 MeOH): C, 51.39; H, 6.40; N, 17.11; Found: C, 51.44;
H, 6.42; N, 17.09; MS m/z 394 (M+H)+.
(rel)-(4ꢁR,7ꢁS)-[9-(7-Hydroxymethyl)-spiro[2.4]hept-5-en-4-yl)-adenine]-
7-phosphonic acid (19). To a solution of the phosphonate 18 (82.5 mg, 0.21
mmol) in anhydrous CH3CN (7 mL) and 2,6-lutidine (0.489 mL, 4.2 mmol)
was added trimethylsilyl bromide (0.321 mg, 2.1 mmol). The mixture was
heated overnight at 60◦C under nitrogen gas and then concentrated in
vacuo. The residue was partitioned between CH2Cl2 (50 mL) and distilled
purified water (50 mL). The aqueous layer was washed with CH2Cl2 (2 × 30
mL) and then freeze-dried to give phosphonic acid 19 (56 mg, 79%) as a
1
yellowish foam: UV (H2O) λmax 260.5 nm; H NMR (DMSO-d6, 300 MHz)
δ 8.29 (s, 1H), 8.10 (s, 1H), 5.67 (d, J = 5.3 Hz, 1H), 5.35 (dd, J = 5.4,
4.3 Hz, 1H), 4.49 (d, J = 4.5 Hz, 1H), 4.15 (d, J = 8.2 Hz, 2H), 0.39–0.32
(m, 4H); 13C NMR (DMSO-d6, 75 MHz) δ 154.7, 152.1, 150.2, 141.7, 136.9,
133.6, 120.0, 90.2, 67.3, 57.8, 21.4, 8.7, 7.6; Anal. Calc. for C13H16N5O4P
(+3.0 H2O): C, 39.92; H, 5.67; N, 18.76; Found: C, 39.89; H, 5.66; N, 18.78;
MS m/z 338 (M+H)+.
(rel)-(4ꢁR,5ꢁR,6ꢁS,7ꢁS)-Diethyl [9-(7-hydroxymethyl)-5,6-dihydroxy-spiro
[2.4]hept-5-en-4-yl)-adenine] phosphonate (20) and (rel)-(4ꢁR,5ꢁS,6ꢁR,7ꢁS)-
diethyl
[9-(7-hydroxymethyl)-5,6-dihydroxy-spiro[2.4]hept-5-en-4-yl)-
adenine] phosphonate (20ꢁ). Compound 18 (205 mg, 0.522 mmol)
was dissolved in a cosolvent system (10 mL) (acetone: t-BuOH:H2O =
6:1:1) along with 4-methylmorpholine N -oxide (121 mg, 1.044 mmol).
Subsequently, OsO4 (0.261 mL, 4% wt. % in H2O) was added. The mixture
was stirred overnight at rt and quenched with a saturated Na2SO3 solution
(5 mL). The resulting solid was removed by filtration through a pad of
Celite, and the filtrate was concentrated in vacuo. The residue was purified
by silica gel column chromatography (MeOH/CH2Cl2, 1:4) to give 20 (73
mg, 33%) and 20’ (66 mg, 31%) as a solid: compound 20: mp 147–149◦C;

6ꢁ-Spirocyclopropyl-5ꢁ-Norcarbocyclic Adenosine Analogues
795
1
UV (H2O) λmax 260.0 nm; H NMR (DMSO-d6, 300 MHz) δ 8.30 (s, 1H),
8.19 (s, 1H), 4.24–4.17 (m, 4H), 4.09 (d, J = 8.1 Hz, 2H), 3.79 (d, J = 5.0
Hz, 1H), 3.67 (m, 1H), 3.30 (dd, J = 6.8, 4.6 Hz, 1H), 2.96 (d, J = 6.1 Hz,
1H), 1.36–1.30 (m 6H), 0.38-0.31 (m, 4H); 13C NMR (DMSO-d6, 75 MHz) δ
154.8, 152.2, 149.5, 142.7, 119.8, 84.6, 67.6, 66.2, 64.2, 63.7, 59.3, 20.4, 9.2,
8.5; Anal. Calc. for C17H26N5O6P (+0.5 MeOH): C, 47.42; H, 6.37; N, 15.80;
Found: C, 47.39; H, 6.39; N, 15.78; MS m/z 428 (M+H)+.
1
Compound 20ꢁ: mp 153–154◦C; UV (H2O) λmax 260.5 nm; H NMR
(DMSO-d6, 300 MHz) δ 8.29 (s, 1H), 8.15 (s, 1H), 4.35-4.28 (m, 4H), 4.15
(d, J = 8.0 Hz, 2H), 3.76 (br s, 1H), 3.59–3.46 (m, 2H), 1.38–1.34 (m 6H),
0.41-0.37 (m, 4H); 13C NMR (DMSO-d6, 75 MHz) δ 154.6, 152.5, 150.7, 143.2,
120.2, 85.2, 68.1, 65.3, 64.7, 60.8, 19.9, 8.0, 7.3; Anal. Calc. for C17H26N5O6P
(+1.0 MeOH): C, 47.08; H, 6.58; N, 15.25; Found: C, 47.11; H, 6.60; N, 15.27;
MS m/z 428 (M+H)+.
(rel)-(4ꢁR,5ꢁR,6ꢁS,7ꢁS)-[9-(7-Hydroxymethyl)-5,6-dihydroxy-spiro[2.4]
hept-5-en-4-yl)-adenine] phosphonic acid (21). The final adenosine phos-
phonic acid 21 was synthesized from 20 using a similar procedure described
for 19 as a light yellow foamy solid: yield 64%; UV (H2O) λmax 261.5 nm; 1H
NMR (DMSO-d6, 300 MHz) δ 8.29 (s, 1H), 8.17 (s, 1H), 4.14 (d, J = 8.2 Hz,
2H), 3.80 (d, J = 5.5 Hz, 1H), 3.66 (dd, J = 5.8, 4.8 Hz, 1H), 3.39 (dd, J =
5.4, 4.7 Hz, 1H), 0.41–0.34 (m, 4H); 13C NMR (DMSO-d6, 75 MHz) δ 154.5,
152.6, 151.7, 142.5, 120.0, 84.2, 67.4, 64.4, 63.7, 59.8, 19.6, 8.4, 7.3; Anal.
Calc. for C13H18N5O6P (+2.0 H2O): C, 38.35; H, 5.44; N, 17.20; Found: C,
38.32; H, 5.46; N, 17.17; MS m/z 372 (M+H)+.
(rel)-(4ꢁR,7ꢁS)-Bis(SATE) phosphoester of [9-(7-methyloxyphosphonate-
spiro[2.4]hept-5-en-4-yl)]-adenine (23). A solution of adenine phosphonic
acid derivative 19 (57 mg, 0.169 mmol) and tri-n-butylamine (94 mg, 0.510
mmol) in methanol (3.9 mL) was mixed for 30 min and concentrated un-
der reduced pressure. The residue was thoroughly dried with anhydrous
ethanol and toluene. The resulting foamy solid was dissolved in anhy-
drous pyridine (10 mL) to which thioester 22 (519 mg, 3.2 mmol) and 1-
(2-mesitylenesulfonyl)-3-nitro-1H-1,2,4-triazole (201 mg, 0.678 mmol) were
added. The mixture was stirred overnight at rt and quenched with tetra-
butylammonium bicarbonate buffer (10.0 mL, 1 M solution, pH 8.0). The
mixture was concentrated under reduced pressure and the residue was di-
luted with water (60 mL) and extracted with CHCl3 (70 mL) two times.
The combined organic layer was washed with brine, dried over Na2SO4,
filtered, and evaporated. The residue was purified by silica gel column chro-
matography (MeOH/Hexane/EtOAc, 0.05:4:1) to give 23 (38 mg, 36%) as
a white solid: mp 130–133◦C; UV (MeOH) λmax 260.5 nm; 1H NMR (CDCl3,
300 MHz) δ 8.30 (s, 1H), 8.12 (s, 1H), 5.54 (d, J = 5.7 Hz, 1H), 5.31 (dd,
J = 5.6, 4.4 Hz, 1H), 4.49 (m, 1H), 4.22 (d, J = 8.0 Hz, 2H), 3.93-3.90 (m,
4H), 3.17–3.14 (m, 4H), 1.22–1.18 (m, 18), 0.88 (m, 1H), 0.39-0.34 (m, 4H);
13C NMR (CDCl3, 75 MHz) δ 203.8, 154.7, 152.3, 148.5, 143.8, 127.6, 123.7,

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L. J. Liu et al.
119.8, 84.7, 66.8, 64.5, 62.8, 58.5, 48.3, 35.8, 26.2, 19.7, 8.8, 7.5; Anal. Calc.
for C27H40N5O6PS2 (+0.5 MeOH): C, 51.46; H, 6.59; N, 10.91; Found: C,
51.49; H, 6.61; N, 10.89; MS m/z 626 (M+H)+.
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