(Z)- and (E)-2-(1,2-dihydroxyethyl)methylenecyclopropane analogues of 2′-deoxyadenosine and 2′-deoxyguanosine. Synthesis of all stereoisomers, absolute configuration, and antiviral activity

Article abstract of DOI:10.1021/jm900126v

Chiral Z- and E-stereoisomers of (1,2-dihydroxyethyl)methylenecyclopropane analogues of 2′-deoxyadenosine and 2′-deoxyguanosine were synthesized, and their antiviral activity was investigated. (S)- Methylenecyclopropylcarbinol (16) was converted in seven

Full text of DOI:10.1021/jm900126v

J. Med. Chem. 2009, 52, 3397–3407
3397
(Z)- and (E)-2-(1,2-Dihydroxyethyl)methylenecyclopropane Analogues of 2′-Deoxyadenosine and
2′-Deoxyguanosine. Synthesis of All Stereoisomers, Absolute Configuration, and Antiviral
Activity
Shaoman Zhou,† John C. Drach,‡ Mark N. Prichard,§ and Jiri Zemlicka*,†
DeVelopmental Therapeutics Program, Barbara Ann Karmanos Cancer Institute, Wayne State UniVersity School of Medicine, Detroit,
Michigan 48201-1379, Department of Biologic and Materials Science, School of Dentistry, UniVersity of Michigan, Ann Arbor,
Michigan 48018-3218, and Department of Pediatrics, The UniVersity of Alabama School of Medicine, Birmingham, Alabama 35233
ReceiVed February 2, 2009
Chiral Z- and E-stereoisomers of (1,2-dihydroxyethyl)methylenecyclopropane analogues of 2′-deoxyadenosine
and 2′-deoxyguanosine were synthesized, and their antiviral activity was investigated. (S)-Methylenecyclo-
propylcarbinol (16) was converted in seven steps to reagents 26 and 27, which were used for
alkylation-elimination of adenine and 2-amino-6-chloropurine to get ultimately analogues 12a, 12b, 13a,
13b, 14a, 14b, 15a, and 15b. The enantiomeric series ent-12a, ent-12b, ent-13a, ent-13b, ent-14a, ent-14b,
ent-15a, and ent-15b was obtained by similar procedures starting from (R)-methylenecyclopropylcarbinol
(ent-16). The Z-isomer ent-12b was an inhibitor of two strains of human cytomegalovirus (HCMV) with
EC50 of 6.8 and 7.5 µM and of murine cytomegalovirus (MCMV) with EC50 of 11.3 µM. It was less active
against HCMV with mutated gene UL97. It inhibited Epstein-Barr virus (EBV) with EC50 of 8 µM. The
E-isomers ent-15a, ent-13a, and 15b were less effective. All adenine analogues with the exception of the
Z-isomers ent-12a and ent-14a were moderate substrates for adenosine deaminase.
Introduction
The analogy between C-O-C grouping of acyclovir (9) or
ganciclovir (10) and methylenecyclopropane moiety was crucial
in designing analogues 1-4. It was therefore of interest whether
this type of relationship can be extended to other antiviral
nucleoside analogues. We determined that 9-[(2,3-dihydroxy-
1-propoxy)methyl]guanine (11) would be a good starting point
for such a design. Structurally, it is a positional isomer of
ganciclovir (10) with a strong potency against herpes simplex
viruses 1 and 2 (HSV-1, HSV-2) and HCMV.9,10 The S-
enantiomer of 11 is more potent than the R-enantiomer. A design
of methylenecyclopropane analogues based on compound 11
is stereochemically more complex. Thus, from a single hetero-
cyclic base, eight stereoisomers can be derived given the two
chiral centers present and taking Z and E isomerism into account
(Chart 2, 12-15 and ent-12 to ent-15). All 16 analogues
comprising two bases, adenine and guanine, were synthesized
and tested for antiviral activity. It is important to note that
vertical relationships in Chart 2 (columns) are enantiomeric,
whereas horizontal (rows) correspond to diastereoisomers or
Z,E-isomers.
Methylenecyclopropane analogues of nucleosides are estab-
lished antiviral agents1 effective against ꢀ-herpesviruses (cy-
tomegalovirus, CMV;a human herpes virus 6, HHV-6) and
γ-herpesviruses (Epstein-Barr virus, EBV; human herpes virus
8, HHV-8). In the first generation analogues, which have only
a single hydroxymethyl group, the Z-isomers 1 (Chart 1) are
most effective whereas the E-isomers 2 are active only
exceptionally. In the second generation group, the antiviral
activity is more narrow but the trend was similar. The Z-isomers
3 were effective against CMV and EBV, whereas the anti-EBV
effect was also found in some of the E-isomers 4. In this group
of methylenecyclopropanes containing two geminal hydroxym-
ethyl functions, the most potent analogue, cyclopropavir (3b),
is now in preclinical development as a potential drug against
human cytomegalovirus (HCMV) infections.2-5 Vicinal cis-bis-
hydroxymethyl analogues 5a and 5b were ineffective as antiviral
agents.6 Adenine trans analogue 6 was also synthesized, but
antiviral activity has not been reported.7 Addition of a third
hydroxymethyl group (Z- and E-isomers 7 and 8) led to a loss
of antiviral activity.8 Analogues 1 and 2 can be regarded as
analogues of antiherpetic drug acyclovir (9), whereas bis-
hydroxymethyl methylenecyclopropanes 3 and 4, and particu-
larly cyclopropavir (3b), are related to ganciclovir (10), a drug
used against HCMV.
Synthesis
Enantiomeric (methylenecyclopropyl)methanols11-13 16 and
ent-16 served as convenient starting materials for synthesis of
all stereoisomeric analogues reported herein. Importantly, the
originally assigned11,12 absolute configurations of 16 and ent-
16 were later reversed.13 In the 4′S series of analogues 12-15,
(S)-(+)-(methylenecyclopropyl)methanol (16) was first oxidized
to the respective aldehyde 17 using oxalyl chloride and DMSO
reagent12 (Scheme 1). The crude aldehyde 17 was transformed
to the diastereoisomeric cyanohydrin 18 using NaCN under
phase transfer conditions14 in 54% yield. Hydrolysis gave the
corresponding acid 19, which in turn were converted to benzyl
esters 20 and 21 by the action of benzyl bromide, NBu4I, and
K2CO3 in DMF. Both diastereoisomers were readily separated
by column chromatography on silica gel to give the 1S,2S-
* To whom correspondence should be addressed. Telephone: (313) 578-
4288. Fax: (313) 832-7294. E-mail: zemlicka@karmanos.org.
†
Wayne State University School of Medicine.
University of Michigan.
The University of Alabama School of Medicine.
‡
§
a Abbreviations: CMV, cytomegalovirus; HCMV, human cytomegalovi-
rus; MCMV, murine cytomegalovirus; UL97, gene for human cytomega-
lovirus phosphotransferase, pUL97; phosphotransferase enzyme; EBV,
Epstein-Barr virus, EBV; HSV-1, herpes simplex virus 1; HSV 2, herpes
simplex virus 2; HHV 6, human herpes virus 6; HHV-8, human herpes
virus 8; VZV, varicella zoster virus; HFF, human foreskin fibroblasts; MEF,
mouse embryonic fibroblasts; CPE, cytopathic effect; ADA, adenosine
deaminase; NOE, nuclear Overhauser effect.
10.1021/jm900126v CCC: $40.75
2009 American Chemical Society
Published on Web 04/27/2009

3398 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10
Zhou et al.
Chart 1a
a Series a: B ) Ade. Series b: B ) Gua. Series c: B ) 2-amino-6-chloropurine.
Table 1. Chiral HPLC of Adenine Stereoisomersa 12a-15a and ent-12a
to ent-15a
Chart 2a
stereoisomer
tR (min)b
ee (%)c
12a
13a
14a
15a
ent-12a
ent-13a
ent-14a
ent-15a
10.62
11.94
11.69
7.80
8.10
8.0
95.3
96.5
99.0
96.4
100.0
100.0
100.0
100.0
7.21
6.37
a Chiralpak AD column, 25 cm × 0.46 cm, methanol, 1.0 mL/min,
detection at 277 nm. b tR, retention time. c ee, enantiomeric enhancement.
a Seriesa:B)Ade.Seriesb:B)Gua.Seriesc:B)2-amino-6-chloropurine.
tribromide in CH2Cl2 furnished a mixture of the Z/E-isomers
26 (86%), whereas 25 gave dibromides 27 in 76% yield.
The 4′S,5′S series of analogues 12 and 13 were obtained using
dibromide 26. Alkylation-elimination protocol with adenine
and 26 (K2CO3 in DMF at 100-105 °C) gave an inseparable
mixture of the Z- and E-isomeric acetates 28a in 56% yield. In
a similar fashion, 2-amino-6-chloropurine and 26 afforded
intermediates 28b (57% yield). Deacetylation of 28a with K2CO3
in methanol furnished, after chromatographic separation, the
Z-isomer 12a (36%) and E-isomer 13a (49%). Deprotection of
28b was performed using NH3 in methanol to give the Z- and
E-isomers 12c and 13c in 41% and 48% yields, respectively.
Hydrolysis of 12c and 13c in 80% formic acid at 80 °C followed
by treatment with NH3 in methanol gave guanine Z- and
E-isomers 12b and 13b (83% and 84%).
Scheme 1
In the 4′S,5′R series 14 and 15, reaction of adenine with
dibromide 27 was performed under the conditions similar to
those used for dibromide 26 to give the Z/E isomers 29a (58%).
2-Amino-6-chloropurine and 27 gave the isomeric mixture 29b
(55% yield). Deacetylation of 29a gave the Z- and E-isomers
14a and 15a in 35% and 48% yields, respectively, whereas 29b
afforded the Z- and E-isomers 14c and 15c (34% and 45%).
Hydrolytic dechlorination furnished guanine analogues 14b and
15b (86% and 83%).
Synthesis of the enantiomeric series ent-12 through ent-15
started from (R)-(-)-(methylenecyclopropyl)methanol (ent-16)
and followed the procedures outlined in Scheme 1 (ent-16
through ent-29). The enantiomeric enhancement (ee) of adenine
stereoisomers 12a-15a and ent-12a through ent-15a determined
by chiral HPLC was >95% (Table 1).
stereoisomer 20 (35%) and 1S,2R-stereoisomer 21 (38%).
Compounds 20 and 21 were reduced with diisobutylaluminum
hydride (DIBALH) in hexanes to give diols 22 and 23 in 74%
and 79% yields, respectively. Acetylation afforded diacetates
24 and 25 (84% and 86%). Bromination of 24 using pyridinium
Assignment of Absolute Configuration and Z/E
Isomerism
The absolute configuration of diastereoisomers 12-15 and
ent-12 to ent-15 were assigned as follows. The configurations

Methylenecyclopropane Analogues
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10 3399
Scheme 2
Table 2. Selected Chemical Shifts (δ, ppm) of Adenine and Guanine
Analogues 12a-15a and 12b-15b
at the cyclopropane carbon atom C-4′ of analogues were
guaranteed by enantiomeric starting materials 16 and ent-16 (see
carbon C-1 in structures 20 and 21). To establish configuration
at the neighboring exocyclic carbon C-5′ (C-2 of of 20 and 21),
both esters were transformed to conformationally locked ox-
abicyclohexanes 30 and 31 (Scheme 2). Stereoisomer 20 was
first converted to cis,trans-dibromide 32, which was in turn
cyclized to oxabicyclohexane 30 (35%). Similarly, compound
21 afforded isomeric derivative 31 via dibromide 33 in 32%
yield. In both cases, only the cis isomers of 32 and 33 can
undergo cyclization. A similar reaction of unprotected racemic
methylenecyclopropyl alcohols with iodine followed by cy-
clization was described.15 Structure and configuration of both
stereoisomers 30 and 31 followed from NMR spectroscopy
including nuclear Overhauser effect (NOE) experiments. A
strong NOE enhancement (2.79% and 3.07%) was found in
compound 31 between the cis-configured H-1 and H-2, whereas
none was present in isomer 30 where these hydrogens are trans.
This established that absolute configuration is 1S,2R in 30 and
1R,2R in 31, respectively. Consequently, the configurations of
esters 20 and 21 are 1S,2S and 1S,2R. The enantiomers ent-20
and ent-21 have then opposite configurations. Thus, the absolute
configurations of all four key intermediates 20, 21, ent-20, and
ent-21 were secured.
compd (isomer)
H1′
H8
5′-OH
C3′
C4′
12a(Z)
13a(E)
14a(Z)
15a(E)
12b(Z)
13b(E)
14b(Z)
15b(E)
7.38
7.49
7.36
7.42
7.10
7.24
7.09
7.17
8.96
8.46
8.46
8.47
8.56
8.02
8.01
8.04
5.23
4.83
4.91
4.90
5.10
4.78
4.85
4.85
6.7
8.7
5.8
9.6
6.6
8.5
5.7
9.5
20.8
19.0
21.2
19.8
20.7
18.8
20.9
19.8
14b by the 5′-OH. At any rate, these differences may help in
distinguishing both stereoisomeric series of analogues. No
significant change in the 6′-OH chemical shifts was observed
in any of these analogues. The C4′ chemical shifts C4′(Z) > C4′(E)
then followed the trend seen in other methylenecyclopropane
analogues.17 Needless to say, all these patterns were also
reflected in the corresponding enantiomeric series 4′R,5′R and
4′R,5′S.
The assignment of the Z- and E-isomers was unequivocally
confirmed by the NOE results with guanine analogues 12b,
13b, 14b, and 15b (Tables 3 and 4). As expected,16 the
interactions of the cis-related protons with the H8 of purine
in an anti-like conformation were crucial for assignment. In
the Z-isomers 12b and 14b, strong effects were seen between
the H8 and 5′-OH, H6′(H5′), and H4′ complemented by NOE
enhancements between the H1′ and H3′. By contrast, interac-
tions between the H8 and H3′ as well as between H1′ and H4′
were typical for the E-isomers 12b and 15b. In the case of
15b, the long-range effects between the H1′ and H5′(H6′) and
even 6′-OH assisted possibly by the rigidity of methylenecy-
clopropane scaffold were also observed.
Dibromides 26, 27, ent-26, and ent-27 obtained from these
intermediates (Scheme 1) were used for synthesis of all
stereoisomeric analogues. Consequently, two pairs of adenine
Z- and E-enantiomers 12a, ent-12a and 13a, ent-13a were
readily identified and their relationship with 14a, 15a and ent-
14a, ent-15a must be diastereoisomeric. Similar conclusions
apply for the respective guanine analogues.
As observed wih other methylenecyclopropane analogues,16
the cis-(Z)-isomers were less polar, moving more quickly when
chromatographed on silica gel than trans-(E)-isomers. Final
isomeric assignment was confirmed by NMR spectroscopy
(Table 2). The chemical shifts of H1′ and C3′ followed the trend
observed earlier:2,16 H1′(E) > H1′(Z) and C3′(E) > C3′(Z).
Nevertheless, the “usual” pattern of chemical shifts of H8 and
5′-OH /H8(Z) > H8(E) and 5′-OH(Z) > 5′-OH(E) held only for
the 4′S,5′S series 12a, 13a, 12b, and 13b. In the 4′S,5′R series
14a, 15a, 14b, and 15b, no significant differences in these
chemical shifts were found for the Z- or E-isomers. This
indicates a lack of deshielding of H8 of the Z-isomers 14a and
The final confirmation of absolute configuration and Z,E-
isomeric structure came from X-ray diffraction of a single
crystal of diacetate 34 derived from adenine analogue ent-
12a (Figure 1). The X-ray confirmed the Z-configuration of
34 as well as the anti-like conformation of the purine base.
Given the fact that diacetate 34 does not contain any heavy
atom and molybdenum tube was used in the experiment,
determination of only relative configuration at the C-4′ and
C-5′ (C-9 and C-10 in Figure 1) was possible. Nevertheless,
as indicated above, the configuration at the C-4′ is R by virtue

3400 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10
Zhou et al.
Table 3. NOE Data of the Z- and E-Isomers 12b and 13b (DMSO-d6,
300 MHz)
compd
Hirr
δ, ppm
8.56
8.56
8.56
8.56
5.10
3.39-3.46
3.10
1.90
1.42
8.02
8.02
1.79-1.84
1.36-1.42
1.61
Hobs
δ, ppm
5.10
3.39-3.46
3.10
1.90
8.56
8.56
8.56
8.56
7.10
1.36-1.40
1.61
7.24
8.02
8.02
1.79-1.84
NOE, %
12b
H8
H8
H8
H8
5′-OH
H6′ (H5′)
H6′ (H5′)
H4′
H3′
H8
H8
H4′
5′-OH
H6′ (H5′)
H6′ (H5′)
H4′
H8
H8
H8′
H8
H1′
H3′
2.07
4.18
1.33
3.61
3.29
0.72
1.38
3.50
1.59
0.56
1.01
0.88
1.47
2.77
1.67
Figure 1. ORTEP-3 view of crystal structure of (4′R,5′R)-diacetate
34 molecule showing 50% probability displacement ellipsoids.
Table 5. Inhibition of HCMV and EBV Replication by Stereoisomeric
1,2-(Dihydroxyethyl)methylenecyclopropane Analogues of Nucleosides
EC50/CC50 (µM/µM)
13b
HCMV/HFF
H3′
H1′
H8
H8
compd configuration
Townea,b
AD169c,d
EBV/Akatae
H3′
H3′
H1′
12a
13a
14a
15a
12b
13b
14b
15b
ent-12a
ent-13a
ent-14a
ent-15a
ent-12b
ent-13b
ent-14b
ent-15b
3b
Z,4′S,5′S
E,4′S,5′S
Z,4′S,5′R
E,4′S,5′R
Z,4′S,5′S
E,4′S,5′S
Z,4′S,5′R
E,4′S,5′R
Z,4′R,5′R
E,4′R,5′R
Z,4′R,5′S
E,4′R,5′S
Z,4′R,5′R
Z,4′R,5′R
Z,4′R,5′S
E,4′R,5′S
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
>100/>100
6.8/>100f
300/>300
>20/50.5
>20/57
43/55
50.4/50.6
43/54
>20/67
>20/56
18/61
52.6/61.5
17/73
>60/>300
>60/>300
>300/>300
>300/>300
>60/>300
>300/>300
>300/>300
>60/194
7.24
H4′
Table 4. NOE Data of the Z- and E-Isomers 14b and 15b (DMSO-d6,
300 MHz)
>60/264
>60/206
60/93.5
9.8/72.7
8/78
>60/194
7.5/299a,g
>60/298
>100/>100
>100/>300
>100/>300
0.46/>100h
40/75
>300/>300
>300/>300
0.49/>100h
>20/62.6
58.7/86.6
0.22//46i
3.3/>100k
control
1.2-2.4/>100j 0.04-1.5/>100j
compd
Hirr
δ, ppm
8.01
8.01
8.01
8.01
4.84
4.64
3.51
3.25-3.31
2.05-2.09
1.30-1.38
7.09
8.04
8.04
Hobs
δ, ppm
4.84
4.64
3.51
2.05-2.09
8.01
8.01
8.01
8.01
8.01
7.09
1.30-1.38
1.64
NOE, %
a Plaque reduction assay in HFF cells. b Visual cytotoxicity in uninfected
HFFs used in plaque reduction assays. c Cytopathic effect (CPE) inhibition
assay. d Cytotoxicity by neutral red uptake. e DNA hybridization assay.
f Average of three experiments. g EC50 of 11.3 µM by plaque reduction assay
in MCMV/ MEF. EC50 of ganciclovir was 5.5 µM. h Reference 2. i Reference
28. j Ganciclovir. k Acyclovir.
14b
H8
H8
H8
H8
5′-OH
6′-OH
H5′ (H6′)
H5′ (H6′)
H4′
H3′
H1′
H8
H8
6′-OH
H5′ (H6′)
H5′ (H6′)
H4′
H3′
H3′
5′-OH
6′-OH
H6′ (H5′)
H4′
H8
H8
H8
H8
H8
H1′
H3′
H3′
H3′
H1′
H1′
H1′
H1′
H8
0.17
0.18
2.33
1.62
0.71
4.33
5.05
5.84
3.45
1.41
1.65
1.07
0.82
2.05
1.27
2.62
0.74
1.48
2.30
mers 14a, 15a and ent-14a, ent-15a must then have the
4′S,5′R and 4′R,5′S configurations, respectively.
Biological Activity
15b
1.35-1.40
7.17
7.17
7.17
7.17
A. Antiviral Effects. Analogues 12a-15a, 12b-15b, and
enantiomers ent-12a to ent-15a, ent-12b to ent-15b were tested
for antiviral activity against the following viruses in vitro: herpes
simplex viruses 1 and 2 (HSV-1, HSV-2), varicella zoster virus
(VZV), human cytomegalovirus (HCMV), Epstein-Barr virus
(EBV), hepatitis B and C virus (HBV, HCV). The results for
HCMV and EBV are summarized in Table 5. Activity against
HCMV was found only for the 4′R,5′R guanine Z-isomer ent-
12b. It was effective in plaque reduction assay against Towne
and AD169 strain of HCMV with EC50 of 6.8 and 7.5 µM,
respectively. It was also active in murine cytomegalovirus
(MCMV) assay (EC50 of 11.5 µM). It is not cytotoxic, and its
efficacy is somewhat lower than that of ganciclovir. The fact
that the new active anti-HCMV agent ent-12b is the Z-isomer
is not surprising because the Z-guanine analogues 1b and 3b
were highly effective.1 None of the E-isomers of the present
4.63
3.37-3.46
2.97-3.03
1.72-1.77
1.64
8.04
8.04
1.30-1.38
H8
of the starting (R)-(-)-(methylenecyclopropyl)methanol (ent-
16) used for the synthesis of analogue ent-12a and diacetate
34. The absolute configuration at carbon C-5′ is then R, thus
1
confirming the H NMR spectroscopic assignment based on
intermediates 20, 21, and respective enantiomers ent-20, ent-
21 (see above). This established not only the 4′R,5′R
configuration and structure of the E-isomer ent-13a but also
of the opposite 4′S,5′S enantiomers 12a and 13a. Stereoiso-

Methylenecyclopropane Analogues
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10 3401
Table 6. Activity of ent-12b against Drug Resistant HCMV
EC50 (µM)a in virus strain
analogues were effective against HCMV, which is in line with
the previous results with other methylenecyclopropane ana-
logues1 such as 2 and 4. More intriguing is a lack of HCMV
potency of adenine analogues 12a-15a and ent-12a to ent-15a
because analogues 1a and 3a were effective against HCMV.1
The very narrow range of potent anti-HCMV activity limited
to a single compound ent-12b with unique 4′R,5′R stereochem-
istry from a total of eight Z-isomeric analogues is a hallmark
of this series. The rigidity of methylenecyclopropane system
coupled with two centers of chirality is probably responsible
for this marked stereoselectivity. Interestingly, an opposite trend,
decreased enantioselectivity, was observed in less rigid dihy-
droxyanalogue 11 with only a single center of asymmetry where
both enantiomers were active against HCMV.9
compd
Towneb
2696rc
E8d
ent-12b
cyclopropavir (3b)
ganciclovir (10)
8
0.6
2
>100
28
42
72
8
22
a Data from a plaque reduction assay with four drug concentrations in
duplicate. b Wild-type virus from which isolates 2696r and E8 were obtained.
c Virus isolated for resistance to cyclopropavir (3b) that has a truncated
UL97 gene. d Virus with two point mutations introduced into gene UL97.
Table 7. Deamination of Adenine 1,2-(Dihydroxyethyl)methylene-
cyclopropane Analogues of Nucleosides by Adenosine Deaminasea
deamination, %
compd
isomer
configuration
24 h
48 h
Analogue ent-12b was also the most effective against EBV
in Akata cell culture with EC50 of 8 µM (Table 5). It is
somewhat less potent than acyclovir (9). The other less potent
analogues included the E-isomers ent-15a, ent-13a, and 15b with
EC50 of 9.8-18 µM indicating that the 4′R,5′R stereochemistry
and Z/E isomerism are less important for anti-EBV than an anti-
HCMV effect. It is noted that effectivity of the E-isomers of
methylenecyclopropane analogues against EBV was observed
before in several instances.1,18 Interestingly, the patterns of anti-
HCMV and anti-EBV effects of analogue ent-12b parallel those
of cyclopropavir 3b, although the former is a weaker antiviral.
Compound ent-12b also exhibited a moderate effect against
HSV-1 (EC50/CC50 of 50/>100 µM) in BSC-1 cells (ELISA),
but it was inactive against HSV-1 and HSV-2 in human foreskin
fibroblast (HFF) culture. It was also somewhat effective in VZV/
HFF assay (EC50/CC50 of 47/284 µM). None of the analogues
was active against hepatitis B or C virus. It is noted that
somewhat similar cyclopropane analogue 35 (n ) 0, B ) Gua)
described in the patent literature19 as a mixture of diastereoi-
somers was without significant activity against HSV-1. The
1′S,2′R,4′S stereoisomers 35 (n ) 0 or 1, B ) Thy, Ura, Ade,
or Gua) were also devoid of antiviral activity.20,21 Nevertheless,
it is emphasized that HCMV and EBV assays with 35 have not
been reported.
12a
13a
14a
15a
ent-12a
ent-13a
ent-14a
ent-15a
Z
E
Z
E
Z
E
Z
E
4′S,5′S
4′S,5′S
4′S,5′R
4′S,5′R
4′R,4′R
4′R,4′R
4′R,5,S
4′R,5′S
45
85
65
100
0
100
0
65
80
100
80
100
0
100
0
85
a For details, see Experimental Section.
much on the stereochemistry of the side chain. Thus, all
E-isomers were deaminated from 80% to 100% within 24-48
h, whereas the Z-analogues ent-12a and ent-14a were resistant
to deamination.
Experimental Section
General Methods. The UV spectra were measured in ethanol,
and NMR spectra were determined in CD3SOCD3 at 400 MHz (1H)
and 100 MHz (13C) unless stated otherwise. Mass spectra were
determined in electron-impact (EI-MS) or electrospray ionization
(ESI-MS, methanol-AcONa) mode. Optical rotations were mea-
sured on JASCO digital polarimeter DIP-370. The (S)-(+)- and
(R)-(-)-(methylenecyclopropyl)methanols (16 and ent-16) were
prepared as described11-13 from (S)-(+)- and (R)-(-)-epichloro-
hydrin. Enantiomeric epichlorohydrins were obtained as reported.27
They were converted to benzyl (S)- and (R)-glycidyl ethers27
using a procedure for racemic compounds,28 and enantiomeric
enhancements (ee) were determined by chiral HPLC (Chiralcel AD
column was used instead of Chjralcel OD27). The ee values of
benzyl (S)- and (R)-glycidyl ether were 96% and 99%, respectively.
A characterization of enantiomeric compounds by UV (where
Regarding the mechanism of antiviral action of active 1,2-
dihydroxyethyl analogues, we hypothesize that the phosphory-
lation cascade generally applicable for activation of nucleosides
(including methylenecyclopropane analogues1), phosphate f
diphosphate f triphosphate, is also operative here. Because none
of the compounds is active against HSV-1 or HSV-2, viral
thymidine kinase likely seems unable to effect the first phos-
phorylation in contrast to ganciclovir analogue 11 where the
hydroxymethyl group is phosphorylated by this enzyme.10 There
is evidence that methylenecyclopropanes 1b, 2b, and 3b are
phosphorylated by the action3,22-25 of HCMV-encoded phos-
photransferase pUL97, and this appears to be the case with ent-
12b as well. Comparison of the activity of ent-12b to that of
cyclopropavir (3b) and ganciclovir (10) in two HCMV strains
with mutated UL97 genes showed that all three compounds were
significantly less active against HCMV with UL97 mutations23,25
than to wild-type virus (Table 6). Together with the known
substrate specificity26 of pUL97 for ganciclovir (10), we take
this as a strong evidence that this enzyme phosphorylates ent-
12b.
applicable), H, 13C NMR, and MS is reported for only one set of
1
enantiomers. The data of the opposite series were essentially
identical. The prefixes “ent-” before compound numbers indicate
opposite enantiomers. Optical rotation was determined for all
enantiomeric compounds. All biologically tested analogues were
g95% pure as indicated by C, H, N analyses. The optical purity of
all adenine analogues was at least 95% as shown by chiral HPLC
(Table 1).
(-)-(R,S)-2-Hydroxy-2-[(S)-2-methylenecyclopropyl]acetoni-
trile (18). To a solution of oxalyl chloride (7.5 mL, 85.0 mmol) in
CH2Cl2 (100 mL) at -60 °C was added DMSO (12.5 mL, 175.0
mmol) in CH2Cl2 (25 mL) with stirring at -60 °C followed, after
45 min, by a dropwise addition of (S)-(+)-(methylenecyclopro-
pyl)methanol (16, 4.2 g, 50.0 mmol) in CH2Cl2 (25 mL). The stirring
was continued for 45 min, whereupon triethylamine (35 mL, 0.25
mol) was added and the reaction mixture containing crude aldehyde
17 was allowed to warm to room temperature. Then 1 M HCl
saturated with NaCl (275 mL) was added and the aqueous layer
was extracted with CH2Cl2 (4 × 50 mL). The organic layers were
combined, and a saturated solution of NH4Cl (30 mL) was added
with stirring and external ice-cooling followed by NaCN (3.67 g,
75 mmol). The stirring was continued for 30 min, the layers were
separated, and the aqueous layer was extracted with CH2Cl2 (6 ×
B. Adenosine Deaminase (ADA). Adenine analogues
12a-15a and ent-12a to ent-15a were tested for substrate
activity toward ADA (Table 7). All active analogues are only
moderate substrates for the enzyme. As observed in the previous
cases of methylenecyclopropane analogues, the E-isomers are
more reactive than the Z-isomers. This effect does not depend

3402 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10
Zhou et al.
25 mL). The combined organic phase was dried over MgSO4. The
solvent was evaporated and the crude product was chromatographed
on a silica gel column using hexanes-diethyl ether (10:1 to 1:1)
to give nitrile 18 as a colorless oil (2.96 g, 54.3%). [R]2D6 -11.8° (c
(c 1.03, CHCl3). 1H NMR (CDCl3) δ 0.97-1.01 (m, 1H), 1.27 (tt,
1H, J ) 8.8, 1.6 Hz), 1.57-1.62 (m, 1H, cyclopropane), 3.10 (bs,
2H, OH), 3.32 (dt, 1H, J ) 7.6, 3.2 Hz), 3.56 (dd, 1H, J ) 11.4,
8.2 Hz), 3.73 (dd, 1H, J ) 11.2, 3.2 Hz, CHO, CH2O), 5.44, 5.53
(2m, 2H, dCH2). 13C NMR 7.3, 17.9 (cyclopropane), 66.4 (CH2O),
74.9 (CHO), 104.7 (dCH), 132.6 (dCH2). EI-HRMS calcd for
C6H8O (M - H2O) 96.0575, found 96.0572. Anal. (C6H10O2 ·
0.2H2O) C, H.
(+)-(R)-1-[(R)-2-Methylenecyclopropyl]ethane-1,2-diol (ent-
22). The procedure described above was performed with (R,R)-
ester ent-20 (1.65 g, 7.6 mmol) to furnish diol ent-22 (630 mg,
73%). [R]2D5 2.7° (c 1.05, CHCl3).
1
1.0, CHCl3). H NMR (300 MHz, CDCl3) δ 1.17-1.27 (m, 1H),
1.43-1.52 (m, 1H), 1.93-2.01 (m, 1H, cyclopropane), 3.90 (bs,
1H, OH), 4.10, 4.29 (2d, J ) 8.4, 7.2 Hz, 1H, CHO), 5.53-5.55
(m, 1H), 5.61-5.65 (m, 1H, dCH2). 13C NMR (75 MHz) 7.5, 9.2,
18.9, 19.2 (cyclopropane), 63.4, 63.7 (CHO), 107.00, 107.04
(dCH2), 118.7, 119.1 (dC), 128.6, 129.5 (CΝ). EI-HRMS calcd
for C6H7NO (M) 109.0528, found 109.0529.
(+)-(R,S)-2-Hydroxy-2-[(R)-2-methylenecyclopropyl]acetoni-
trile (ent-18). The procedure described above was performed with
(R)-(-)-(methylenecyclopropyl)methanol (ent-16, 4.8 g, 57 mmol)
to give nitrile ent-18 via aldehyde ent-17 (3.55 g, 57%), [R]2D3 11.4°
(c 1.58, CHCl3).
(+)-(R)-1-[(S)-2-Methylenecyclopropyl]ethane-1,2-diol (23).
The reduction of (R,S)-ester 21 (1.6 g, 7.34 mmol) followed the
above procedure to give diol 23 (660 mg, 79%) as a white solid:
mp 69-71 °C, [R]2D7 25.4° (c 1.0, CHCl3). H NMR (CDCl3) δ
1
1.06-1.10 (m, 1H), 1.34 (tt, 1H, J ) 8.8, 1.6 Hz), 1.54-1.60 (m,
1H, cyclopropane), 3.01, 3.10 (2bs, 2H, OH), 3.21 (dt, 1H, 8.0, J
) 2.7 Hz), 3.59 (dd, 1H, J ) 10.8, 7.2 Hz), 3.69 (poorly resolved
dd, 1H, J ) 10.4 Hz, CHO, CH2O), 5.40 (d, 1H, J ) 2.4 Hz), 5.42
(d, 1H, J ) 1.6 Hz, dCH2). 13C NMR 8.3, 18.5 (cyclopropane),
66.7 (CH2O), 75.2 (CHO), 104.8 (dCH), 132.0 (dCH2). EI-HRMS
calcd for C6H8O (M - H2O) 96.0575, found 96.0580. Anal.
(C6H10O2 ·0.45H2O) C, H.
(-)-(S)-1-[(R)-2-Methylenecyclopropyl]ethane-1,2-diol (ent-
23). The reduction of (S,R)-ester ent-21 (2.0 g, 9.17 mmol) followed
the above procedure to give diol ent-23 (805 mg, 77%): mp 73-74
°C, [R]2D6 -25.0° (c 1.05, CHCl3).
(-)-Benzyl (S)-2-Hydroxy-2-[(S)-2-methylenecyclopropyl)]ac-
etate (20) and (-)-Benzyl (R)-2-Hydroxy-2-[(S)-2-methylenecy-
clopropyl)]acetate (21). A solution of nitrile 18 (2.76 g, 25.32
mmol) in CH2Cl2 (100 mL) was stirred with 37% HCl (25 mL) at
room temperature for 2 h. The volatiles were evaporated at 18 torr
and room temperature, and a solution of the residue in ether was
passed through a 5 cm silica gel pad. The pad was washed with
hexanes-Et2O (1:1, 180 mL), and the filtrate was concentrated to
give acid 19 (2.91 g, 90%), which was directly used in the next
step.
Benzyl bromide (8.4 mL, 70.3 mmol) was added dropwise with
stirring to acid 19 (2.8 g, 21.9 mmol) in DMF (33 mL), anhydrous
K2CO3 (5.07 g, 36.7 mmol), and NBu4I (2.60 g, 7.03 mmol) at
room temperature. The stirring was continued for 3.5 h, the solvent
was evaporated, and the residue was partitioned between water (60
mL) and Et2O (4 × 60 mL). The organic phase was washed with
10% HCl (25 mL), 5% NaHCO3 (25 mL), and brine (20 mL). After
the mixture was dried (MgSO)4, the solvent was evaporated and
the crude product was chromatographed on a silica gel column in
hexanes-Et2O (10:1 to 5:1) to give the (S,S)-isomer 20 (1.67 g,
35%) followed by (S,R)-isomer 21 (1.81 g, 38%).
(+)-(S)-1-[(S)-2-Methylenecyclopropyl]ethane-1,2-diyl Diac-
etate (24). Acetic anhydride (3 mL) was added dropwise to a stirred
solution of diol 22 (520 mg, 4.56 mmol) in pyridine (1.5 mL) at
room temperature. The stirring was continued for 10 h, the reaction
was quenched with water, and product was extracted with ice cold
Et2O (100 mL). The combined organic phase was washed succes-
sively with saturated aqueous CuSO4, 5% HCl (4 × 25 mL),
aqueous NaHCO3 (3 × 20 mL), and brine (3 × 20 mL). It was
dried (MgSO4), the solvent was evaporated, and the residue was
chromatographed on a silica gel column (hexanes-Et2O, 50:1 to
10:1) to give diacetate 24 (760 mg, 84%) as a colorless oil: [R]D24
1
(S,S)-Isomer 20: [R]D26 -7.3° (c 1.0, CHCl3). H NMR (CDCl3)
δ 1.28-1.34 (m, 2H), 1.82-1.85 (m, 1H, cyclopropane), 2.73 (s,
1H, OH), 4.04 (d, 1H, J ) 5.6 Hz, CHO), 5.23 (AB, 2H, JAB ) 12
Hz, CH2 of Bn), 5.46 (d, 2H, J ) 2.4 Hz, dCH2), 7.37 (m, 5H,
Ph). 13C NMR 6.8, 18.8 (cyclopropane), 67.5 (CHO), 71.0 (CH2
of Bn), 105.7 (dCH), 128.5, 128.7, 128.9, 130.9, 135.5 (dC, Ph),
174.3 (CdO). ESI-MS 241.3 (M + Na, 100.0). Anal. Calcd for
C13H14O3 ·0.2H2O.
1
80.9° (c 0.97, CHCl3). H NMR (CDCl3) δ 1.06-1.09 (m, 1H),
1.37 (tt, 1H, J ) 9.2, 1.6 Hz), 1.71 (m, 1H, cyclopropane), 2.05,
2.07 (2s, 6H, CH3), 4.09 (dd, 1H, J ) 12.0, 7.2 Hz), 4.34 (dd, 1H,
J ) 12.4, 2.8 Hz,), 4.71 (dt, 1H, J ) 8.8, 3.2 Hz, CHO, CH2O),
5.42, 5.44 (2s, 2H, dCH2). 13C NMR 7.9, 15.8 (cyclopropane),
21.0, 21.3 (CH3), 65.0 (CH2O), 73.8 (CHO), 105.2 (dCH), 131.2
(dC), 170.5, 171.0 (CdO). ESI-MS 221 (M + Na, 100.0). EI-
HRMS calcd for C10H15O4 199.0970, found 199.0966. Anal.
(C10H14O4) C, H.
(S,R)-Isomer 21: [R]2D5 -45.3° (c, 1.02, CHCl3). 1H NMR (CDCl3)
δ 1.19 (m, 1H), 1.37 (tt, 1H, J ) 8.8, 2.4 Hz), 1.72-1.79 (m, 1H,
cyclopropane), 2.93 (s, 1H, OH), 3.80 (d, 1H, J ) 7.2 Hz, CHO),
5.20 (AB, 2H, JAB ) 12 Hz, CH2 of benzyl), 5.40, (s, 1H), 5.43 (d,
1H, J ) 1.6 Hz, dCH2), 7.37 (m, 5H, Ph). 13C NMR 8.0, 19.4
(cyclopropane), 67.8 (CHO), 72.6 (CH2 of Bn), 106.0 (dCH2),
128.6, 128.8, 128.9, 130.9, 135.4 (dC, Ph), 174.2 (CdO). ESI-
MS 241.3 (M + Na, 100.0). Anal. (C13H14O3 × 0.3 H2O) C, H.
(+)-Benzyl (R)-2-Hydroxy-2-[(R)-2-methylenecyclopropyl)]-
acetate (ent-20) and (+)-Benzyl (S)-2-Hydroxy-2-[(R)-2-meth-
ylenecyclopropyl)]acetate (ent-21). The protocol described above
was repeated with nitrile ent-18 (3.10 g, 28 mmol) to give esters
ent-20 (1.71 g, 28%) and ent-21 (2.52 g, 41%) via acid ent-19 as
an intermediate. ent-20: [R]2D6 6.3° (c 1.04, CHCl3). ent-21: [R]2D6
44.9° (c 1.02, CHCl3).
(-)-(R)-1-[(R)-2-Methylenecyclopropyl]ethane-1,2-diyl Diac-
etate (ent-24). The protocol described above was followed with
diol ent-22 (600 mg, 5.26 mmol) to give diacetate ent-24 (840 mg,
81%), [R]2D7 -78.1° (c 1.0, CHCl3).
(+)-(R)-1-[(S)-2-Methylenecyclopropyl]ethane-1,2-diyl Diac-
etate (25). The procedure described above was performed with diol
23 (610 mg, 5.35 mmol) to give diacetate 25 (910 mg, 86%) as a
colorless oil: [R]D26 31.6° (c 1.0, CHCl3). 1H NMR (CDCl3) δ
1.10-1.14 (m, 1H), 1.28 (tt, 1H, J ) 8.8, 2.4 Hz), 1.58-1.63 (m,
1H, cyclopropane), 1.99, 2.00 (2s, 6H, CH3), 4.08-4.19 (m, 2H),
4.53-4.68 (m, 1H, CH2O, CHO), 5.40 (m, 2H, dCH2). 13C NMR
8.8, 16.5 (cyclopropane), 20.9, 21.2 (CH3), 65.4 (CH2O), 73.6
(CHO), 105.6 (dCH), 131.0 (dC), 170.6, 170.7 (CdO). ESI-MS
221 (M + Na, 100.0). Anal. (C10H14O4) C, H.
(-)-(S)-1-[(R)-2-Methylenecyclopropyl]ethane-1,2-diyl Diac-
etate (ent-25). The protocol described above was followed with
diol ent-23 (720 mg, 6.32 mmol) to give diacetate ent-25 (1.04 g,
81%), [R]2D7 -33.9° (c 0.98, CHCl3).
(S)-1-[(1R,2S)-2-Bromo-2-(bromomethyl)cyclopropyl]ethane-
1,2-diyl Diacetate + (S)-1-[(1R,2R)-2-Bromo-2-(bromomethyl)-
cyclopropyl]ethane-1,2-diyl Diacetate (26). Pyridinium tribromide
(1.64 g, 5.15 mmol) was added with stirring to a solution of
diacetate 24 (680 mg, 3.43 mmol) in CH2Cl2 (18 mL) at -20 °C.
(-)-(S)-1-[(S)-2-Methylenecyclopropyl]ethane-1,2-diol (22).
DIBALH in hexanes (1M, 24 mL, 24 mmol) was added to a solution
of the (S,S)-ester 20 (1.50 g, 6.9 mmol) in THF (40 mL) with stirring
at 0 °C during 10 min under N2. The stirring was continued for
1 h. The reaction was quenched by a dropwise addition of HCl
(5%, 35 mL). After stirring for 2 h, it was extracted with Et2O (15
× 30 mL). The organic phase was washed successively with
saturated NaHCO3 (2 × 30 mL) and brine (2 × 30 mL). After
drying (MgSO4), the solvents were evaporated and the crude product
was chromatographed on a silica gel column in hexanes-Et2O (2:1
to 1:3) to give diol 22 (580 mg, 74%) as a colorless oil, [R]2D4 -3.5°

Methylenecyclopropane Analogues
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10 3403
The reaction mixture was allowed to warm to room temperature.
After 12 h, it was diluted with AcOEt (100 mL), and the resultant
solution was washed sequentially with saturated Na2S2O3 (2 × 25
mL), NaHCO3 (2 × 20 mL), and water (20 mL). The organic phase
was dried over MgSO4, and the solvents were evaporated. The crude
9-{(Z)-(S)-2-[(Z)-(R)-1,2-Diacetoxyethyl)cyclopropylidene]methyl}-
adenine + 9-{(E)-(S)-2-[(R)-1,2-Diacetoxyethyl)cyclopropyli-
dene]methyl}adenine (29a). The reaction of adenine with dibro-
mide 27 was performed on the same scale as described for
compound 28a (reaction time was 12 h at room temperature and
10 h at 100-105 °C) to give the Z,E-isomers 29a (273 mg, 58%):
mp 135-139 °C, [R]D22 27.8° (c, 1.0, MeOH). UV λmax 226 nm (ε
26 500), 263 (ε 12 700), 278 (ε 9400). 1H NMR (CDCl3) δ
1.51-1.55 (m), 1.60 (dt, J ) 9.2, 2.0 Hz), 1.66-1.70 (m), 1.79
(dt, J ) 8.8, 2.4 Hz, H3′), 2.00, 2.01, 2.10, 2.11 (4s, 6H, CH3),
2.07-2.10 (m, partly overlapped with 2.10), 2.18-2.23 (m, 1H,
H4′), 4.24, 4.01 and 4.23, 4.03 (2AB, JAB ) 12.2 Hz, 2H, H6′),
4.72, 5.21 (2s, 1H, H5′), 6.28, 6.34 (2bs, 2H, NH2), 7.49, 7.61 (2d,
J ) 1.6 Hz. 1H, H1′), 8.23, 8.24, 8.37 (3s, 2H, H2, H8). 13C NMR
10.62, 10.5, 16.6, 18.3, 20.9, 20.96, 21.02, 64.6, 65.1, 70.1, 73.0,
112.51, 112.54, 113.1, 114.3, 119.3, 119.4, 137.6, 138.8, 148.9,
151.8., 152.2, 154.9, 155.02, 170.1, 170.6, 170.7, 170.8. ESI-MS
332 (M + H, 38.9), 354 (M + Na, 100.0), 685 (2 M + Na, 56.5).
9-{(Z)-(R)-2-[(S)-1,2-Diacetoxyethyl)cyclopropylidene]methyl}-
adenine + 9-{(E)-(R)-2-[(S)-1,2-Diacetoxyethyl)cyclopropyli-
dene]methyl}adenine (ent-29a). A mixture of adenine (220 mg,
1.63 mmol), dibromide ent-27 (550 mg, 1.54 mmol), and K2CO3
(1.28 g, 9.3 mmol) in DMF (8 mL) was stirred under N2 for 24 h
at room temperature and 8 h at 100-105 °C. The workup followed
the above procedure to furnish the Z,E-isomers ent-29a (280 mg,
55%): mp 152-156 °C, [R]2D2 -36.6° (c, 1.0, MeOH).
2-Amino-6-chloro-9-{(Z)-(S)-2-[(S)-1,2-diacetoxyethyl)cyclo-
propylidene]methyl}purine + 2-Amino-6-chloro-9-{(E)-(S)-2-
[(S)-1,2-diacetoxyethyl)cyclopropylidene]methyl}purine (28b).
The reaction was performed as described for adenine Z,E-isomers
28a with 2-amino-6-choropurine (270 mg, 1.6 mmol), dibromide
26 (550 mg, 1.54 mmol), and K2CO3 (1.27 g, 9.2 mmol) in DMF
(8 mL). Reaction time was 24 h at room temperature and 6 h at
100-105 °C. The crude product was chromatographed in hexanes-
AcOEt (1:1 to 1:2) to give the E,Z-isomers 28b (320 mg, 57%):
mp 142-147 °C, [R]D26 80.0° (c 1.18, CHCl3). UV λmax 311 nm (ε
7800), 228 (ε 29 000). 1H NMR (CDCl3) δ 1.43-1.47, 1.51-1.55
(2m), 1.69, 1.81 (2dt, 1H, J ) 8.8, 1.6 Hz, H3′), 2.25-2.31 (m,
1H, H4′), 1.96, 2.06, 2.09, 2.12 (4s, 6H, CH3), 4.14-4.20, 4.34-4.44
(2m, 2H, H6′), 4.71-4.75, 4.78-4.82 (2m, 1H, H5′), 5.36 (bs, 2H,
NH2), 7.31, 7.38 (2 poorly resolved d, 1H, H1′), 8.17, 8.29 (2s, 1H,
H8). 13C NMR 7.5, 9.2, 16.0, 17.9, 20.9, 21.0, 21.1, 21.2, 64.5,
64.8, 72.9, 74.3, 111.7, 111.8, 113.1, 113.6, 125.1, 125.2, 138.9,
139.5, 151.6, 151.7, 152.5, 159.50, 159.51, 170.3, 170.4, 170.85,
170.89. ESI-MS 366, 368 (M + H, 22.2, 7.4), 388, 390 (M + Na,
100.0, 30.0).
2-Amino-6-chloro-9-{(Z)-(R)-2-[(R)-1,2-diacetoxyethyl)cyclo-
propylidene]methyl}purine + 2-Amino-6-chloro-9-{(E)-(R)-2-
[(R)-1,2-diacetoxyethyl)cyclopropylidene]methyl}purine (ent-
28b). A reaction of 2-amino-6-chloropurine (350 mg, 2.07 mmol)
with dibromide ent-26 (720 mg, 2.0 mmol) and K2CO3 was
performed as indicated above to give the E,Z-isomers ent-28b (360
mg, 46%): mp 136-140 °C, [R]2D6 -87.5° (c 1.01, CHCl3).
2-Amino-6-chloro-9-{(Z)-(S)-2-[(R)-1,2-diacetoxyethyl)cyclo-
propylidene]methyl}purine + 2-Amino-6-chloro-9-{(E)-(S)-2-
[(R)-1,2-diacetoxyethyl)cyclopropylidene]methyl}purine (29b).
The protocol described above was followed with 2-amino-6-
chloropurine (310 mg, 1.83 mmol), dibromide 27 (620 mg, 1.73
mmol), and K2CO3 (1.43 g, 10.4 mmol) at room temperature for
4 h and at 100-105 °C for 7 h. After chromatography in
hexanes-AcOEt (1:1 to 1:3), the E,Z-isomers 29b were obtained
(348 mg, 55%): mp 153-156 °C, [R]2D7 44.7 ° (c 1.1, CHCl3). UV
λmax 311 nm (ε 8000), 228 (ε 35 400). 1H NMR (CDCl3) δ
1.50-1.60 (m), 1.67-1.68 (m), 1.78 (t, J ) 8.9 Hz, 3H, cyclo-
propane), 2.06-2.11 (m, partially overlapped with 2.04), 2.35-2.41
(2m, 3H, cyclopropane), 2.00, 2.03, 2.10, 2.12 (4s, 6H, CH3),
3.96-4.01 (dd, J ) 12.0, 6.2 Hz), 4.24-4.30 (m), 4.73-4.76 (m,
3H, H6′,H5′), 5.27, 5.26 (2bs, 2H, NH2), 7.44, 7.32 (2bs, 1H, J )
1.6 Hz, H1′), 8.18, 8.19 (2s, 1H, H8). 13C NMR 6.2, 10.4, 16.5,
18.4, 20.9.1, 21.0, 21.2, 64.3, 65.2, 70.3, 72.9, 112.3, 112.6, 114.0,
125.25, 125.27, 138.8, 140.0, 151.8, 152.6, 152.7, 159.57, 159.61,
product was chromatographed on
a silica gel column in
hexanes-Et2O (20:1 to 5:1) to afford cis,trans-dibromide 26 (1.06
1
g, 86%) as a colorless oil: [R]2D5 46.5° (c 1.1, CHCl3). H NMR
(CDCl3) δ 1.00 (t, J ) 6.4 Hz), 1.22-1.31 (m), 1.34-1.43 (m),
1.60-1.64 (m, 3H, cyclopropane), 2.06, 2.07, 2.09, 2.10 (4s, 6H,
CH3), 3.64-3.80 (m, 2H, CH2Br), 4.03-4.14 (m), 4.35-4.46 (m,
2H), 4.68-4.73 (m), 4.81-4.85 m, 1H, CH2O, CHO). 13C NMR
20.9, 21.0, 21.2, 21.3, 22.3, 22.6, 27.1, 32.0, 35.5, 38.0, 40.2, 43.4,
64.6, 64.7, 70.8, 74.7, 170.1, 170.4, 170.8, 170.9. ESI-MS (MeOH
+ AcOK) 395, 397, 399 (M + K, 48.8, 100.0, 55.6).
(R)-1-[(1S,2R)-2-Bromo-2-(bromomethyl)cyclopropyl]ethane-
1,2-diyl Diacetate + (R)-1-[(1S,2S)-2-Bromo-2-(bromomethyl)-
cyclopropyl]ethane-1,2-diyl Diacetate (ent-26). The reaction was
performed as described above with diacetate ent-24 (740 mg, 3.74
mmol) to give cis,trans-dibromide ent-26 (1.14 g, 85%): [R]D25
-52.8° (c 1.05, CHCl3).
(R)-1-[(1R,2S)-2-Bromo-2-(bromomethyl)cyclopropyl]ethane-
1,2-diyl Diacetate + (R)-1-[(1R,2R)-2-Bromo-2-(bromomethyl)-
cyclopropyl]ethane-1,2-diyl Diacetate (27). The protocol described
above was performed with diacetate 25 (810 mg, 4.09 mmol) to
give the cis,trans-dibromide 27 (1.11 g, 76%) as a colorless oil,\:
[R]2D7 11.6° (c 1.0, CHCl3). H NMR (CDCl3) δ 1.12 (t, J ) 7.5
1
Hz), 1.15-1.22 (m), 1.31-1.36 (m), 1.41-4.53 (m), 1.83-1.90
(m, 3H, cyclopropane), 2.005, 2.009, 2.02 (3s, 6, CH3), 3.46, 3.74
(AB, JAB ) 11.2 Hz), 3.70, 3.85 (AB, 2H, JAB ) 12.4 Hz, CH2Br),
4.17-4.25 (m), 4.36-4.40 (2d, J ) 3.2, 4.4 Hz, 2H), 4.77-4.82,
4.84-4.89 (2m, 1H, CH2O, CHO). 13C NMR 20.95, 21.00, 21.17,
21.23, 23.4, 23.7, 26.2, 31.8, 35.8, 37.4, 40.9, 44.0, 64.5, 65.6, 69.8,
74.6, 170.3, 170.6, 170.7. ESI-MS (MeOH + AcOK) 395, 397,
399 (M + K, 50.0, 100.0, 56.9).
(R)-1-[(1S,2R)-2-Bromo-2-(bromomethyl)cyclopropyl]ethane-
1,2-diyl Diacetate + (S)-1-[(1S,2S)-2-Bromo-2-(bromomethyl)-
cyclopropyl]ethane-1,2-diyl Diacetate (ent-27). The reaction was
performed as described above with diacetate ent-25 (740 mg, 3.74
mmol) to give cis,trans-dibromide ent-27 (1.14 g, 85%): [R]2D5 -8.9°
(c 1.0, CHCl3).
9-{(Z)-(S)-2-[(S)-1,2-Diacetoxyethyl)cyclopropylidene]methyl}-
adenine + 9-{(E)-(S)-2-[(S)-1,2-Diacetoxyethyl)cyclopropyli-
dene]methyl}adenine (28a). A mixture of adenine (200 mg, 1.48
mmol), dibromide 26 (490 mg, 1.37 mmol), and K2CO3 (1.13 g,
8.2 mmol) in DMF (7 mL) was stirred under N2 at 100-105 °C
for 7 h. After the mixture was cooled, the solids were filtered off
using a silica gel pad that was washed with DMF (60 mL). The
solvent was evaporated in vacuo and the residue was chromato-
graphed on a silica gel column in AcOEt-MeOH (50:1 to 20:1) to
give the Z,E-isomers 28a (254 mg, 56%): mp 141-145 °C, [R]D22
65.8° (c 1.06, MeOH). UV λmax 224 nm (ε 25 700), 262 (ε 12 800),
279 (ε 9100). 1H NMR (CDCl3) δ 1.43-1.47 (m), 1.52-1.56 (m),
1.70, 1.81 (dt, 1H, J ) 8.8, 2.4 Hz), 2.00-2.03 (m, partially
overlapped with CH3 at 2.06), 2.25-2.32 (m, 3H, cyclopropane),
1.91, 2.06, 2.09, and 2.11 (4s, 6H, CH3), 4.15-4.19 (m), 4.41, 4.44
(dt, 2H, J ) 12.0, 1.6 Hz, H6′), 4.64-4.69 (m), 4.77-4.81 (m, 1H,
H5′), 6.32, 6.40 (2bs, 2H, NH2), 7.49, 7.56 (m, 1H, H1′), 8.25, 8.36,
8.37, 8.40 (4s, 2H, H2, H8). 13C NMR 7.6, 9.3, 15.9, 17.6, 20.99,
21.03, 21.3, 64.6, 64.8, 73.0, 74.4, 112.0, 112.1, 112.9, 113.6, 119.3,
119.4, 137.5, 137.9, 148.9, 152.7, 152.9, 155.4, 170.0, 170.4,
170.87, 170.92. ESI-MS 332 (M + H, 44.5), 354 (M + Na, 100.0),
685 (2M + Na, 85.8).
9-{(Z)-(R)-2-[(R)-1,2-Diacetoxyethyl)cyclopropylidene]methyl}-
adenine + 9-{(E)-(R)-2-[(R)-1,2-Diacetoxyethyl)cyclopropyli-
dene]methyl}adenine (ent-28a). A mixture of adenine (230 mg,
1.7 mmol), dibromide ent-26 (580 mg, 1.62 mmol), and K2CO3
(1.34 g, 9.7 mmol) in DMF (8 mL) was stirred under N2 for 48 h
at room temperature and then for 8 h at 100-105 °C. The workup
followed the protocol described above to give the Z,E-isomers ent-
28a (327 mg, 61%): mp 163-165 °C, [R]2D2 -75.5° (c 1.0, MeOH).

3404 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10
Zhou et al.
170.2, 170.6, 170.7, 170.9. ESI-MS 366, 368 (M + H, 26.0, 8.3),
388, 390 (M + Na, 100.0, 32.3).
119.1, 137.9, 148.9, 153.6, 156.7 (adenine, C2′, C1′). ESI-MS 248
(M + H, 100.0), 270 (M + Na, 11.9). Anal. (C11H13N5O2) C,
H, N.
2-Amino-6-chloro-9-{(Z)-(R)-2-[(S)-1,2-diacetoxyethyl)cyclo-
propylidene]methyl}-purine + 2-Amino-6-chloro-9-{(E)-(R)-2-
[(S)-1,2-diacetoxyethyl)cyclopropylidene]methyl}purine (ent-
29b). The reaction was performed as described above with
dibromide ent-27. Reaction time was 24 h at room temperature and
7 h at 100-105 °C: yield 340 mg (53%) of the E,Z-isomers ent-
29b, mp 166-169 °C, [R]2D7 -47.0° (c 1.01, CHCl3).
(+)-9-{(Z)-(S)-2-[(S)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}adenine (12a) and (-)-9-{(E)-(S)-2-[(S)-1,2-Dihydro-
xyethyl)cyclopropylidene]methyl}adenine (13a). A mixture of the
Z,E-isomers 28a (240 mg, 0.72 mmol) and K2CO3 (200 mg, 1.45
mmol) in methanol (15 mL) was stirred for 45 min at room
temperature. The solvent was evaporated and the residue was chro-
matographed on a silica gel column using AcOEt-CH2Cl2-MeOH
(15:15:1 to 5:5:1) to give the Z-isomer 12a (64 mg, 36%) followed
by E-isomer 13a (88 mg, 49%).
(-)-9-{(Z)-(S)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}adenine (ent-14a) and (+)-9-{(E)-(S)-2-[(R)-1,2-Dihydro-
xyethyl)cyclopropylidene]methyl}adenine (ent-15a). The experi-
ment using the Z,E-isomers ent-29a (255 mg, 0.77 mmol) was
performed as described above to give the Z-isomer ent-14a (97
mg, 51%) and E-isomer ent-15a (65 mg, 35%).
Z-Isomer ent-14a: mp 221-223 °C, [R]2D6 -97.2° (c 0.48,
DMSO). Anal. (C11H13N5O2) C, H, N.
E-Isomer ent-15a: mp 230-231 °C, [R]2D8 7.2° (c 0.53, DMSO).
Anal. (C11H13N5O2) C, H, N.
(+)-2-Amino-6-chloro-9-{(Z)-(S)-2-[(S)-1,2-dihydroxyethyl)cy-
clopropylidene]methyl}purine (12c) and (+)-2-Amino-6-chloro-
9-{(E)-(S)-2-[(S)-1,2-dihydroxyethyl)cyclopropylidene]methyl}-
purine (13c). The Z,E-isomers 28b (280 mg, 0.77 mmol) were
dissolved in methanol (50 mL), NH3 in MeOH (20%, 35 mL) was
added, and the mixture was stirred for 13 h at room temperature.
The volatiles were evaporated and the residue was chromatographed
on a silica gel column in AcOEt-MeOH (60:1 to 20:1) to give the
Z-isomer 12c (90 mg, 41%) followed by E-isomer 13c (105 mg,
48%).
Z-Isomer 12a: mp 271-272 °C, [R]2D7 54.6° (c 1.09, DMSO).
UV λmax 278 nm (ε 8400), 258 (ε 11 900), 229 (ε 25 500). 1H NMR
δ 1.27 (poorly resolved t), 1.47 (t, 2H, J ) 8.0 Hz, H3′), 2.00 (poorly
resolved dd, 1H, H4′), 3.09-3.15 (m, 1H), 3.40-3.51 (m, 2H, H5′,
H6′), 4.67 (t, 1H, J ) 5.0 Hz, 6′-OH), 5.23 (d, J ) 4.8 Hz, 5′-OH),
7.31 (bs, 2H, NH2), 7.38 (s, 1H, H1′), 8.15 (s, 1H, H2), 8.96 (s, 1H,
H8). 13C NMR 6.7 (C3′), 20.8 (C4′), 66.5 (C6′), 75.1 (C5′), 110.8,
115.5, 119.0, 137.8, 148.7, 153.6, 156.7 (adenine, C1′, C2′). ESI-
MS 248 (M + H, 95.9), 270 (M + Na, 100.0). Anal. (C11H13N5O2)
C, H, N.
Z-Isomer 12c: mp 215-217 °C, [R]2D6 80.9° (c 0.75, DMF). UV
1
λmax 311 nm (ε 8100), 235 (ε 32 800). H NMR δ 1.27 (poorly
resolved m), 1.46 (t, 2H, J ) 8.4 Hz, H3′), 2.00 (poorly resolved
m, 1H, H4′), 3.11 (poorly resolved m, 1H), 3.80-3.47 (m, 2H, H5′,
H6′), 4.70 (t, 1H, J ) 4.8 Hz, 6′-OH), 5.14 (d, 1H, J ) 4.8, Hz,
5′-OH), 7.01 (s, 2H, NH2), 7.19 (s, 1H, H1′), 8.91 (s, 1H, H8). 13
C
E-isomer 13a: mp 245-247 °C, [R]2D7 -3.6° (c 0.67, DMSO).
UV λmax 278 nm (ε 8100), 262 (ε 11 100), 228 (ε 24 700). 1H NMR
δ 1.43-1.46 (m), 1.66 (t, 2H, J ) 8.8 Hz, H3′), 1.83-1.88 (m, 1H,
H4′), 3.17-3.22 (m, 1H), 3.42-3.46 (m, 2H, H5′, H6′), 4.66 (t, 1H,
J ) 4.8 Hz, 6′-OH), 4.83 (d, J ) 4.0 Hz, 5′-OH), 7.33 (bs, 2H,
NH2), 7.49 (s, 1H, H1′), 8.16 (s, 1H, H2), 8.46 (s, 1H, H8). 13C
NMR 8.7 (C3′), 19.0 (C4′), 66.3 C6′), 73.5 (C5′), 110.9, 116.0, 119.1,
137.8, 148.9, 153.7, 156.7 (adenine, C2′, C1′). ESI- MS 248 (M +
H, 100.0), 270 (M + Na, 85.1). Anal. (C11H13N5O2) C, H, N.
NMR 6.8 (C3′), 20.7 (C4′), 66.4 (C6′), 74.9 (C5′), 110.3, 116.4, 123.7,
140.9, 150.2, 153.0, 160.7 (purine, C2′, C1′). ESI-MS 282, 284 (M
+ H, 100.0, 31.8), 304, 306 (M + Na, 82.7, 23.8).
E-Isomer 13c: mp 258-260 °C, [R]2D5 23.5° (c 0.81, DMF). UV
λ
max 311 nm (ε 8100), 235 (ε 32 800). 1H NMR δ 1.42-1.45 (m),
1.67 (t, 2H, J ) 8.8 Hz, H3′), 1.83-1.88 (m, 1H, H4′), 3.19 (poorly
resolved m, 1H), 3.43 (poorly resolved m, 2H, H5′, H6′), 4.64 (t,
1H, J ) 6.0 Hz, 6′-OH), 4.81 (d, J ) 4.0 Hz, 5′-OH), 7.02 (s, 2H,
NH2), 7.33 (s, 1H, H1′), 8.43 (s, 1H, H8). 13C NMR 8.8 (C3′), 19.0
(C4′), 66.3 (C6′), 73.3 (C5′), 110.5, 117.0, 123.7, 140.2, 150.3, 153.1,
160.7 (purine, C2′, C1′). ESI-MS 282, 284 (M + H, 86.5, 25.2),
304, 306 (M + Na, 100.0, 31.4).
(-)-9-{(Z)-(R)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}adenine (ent-12a) and (+)-9-{(E)-(R)-2-[(R)-1,2-Dihydro-
xyethyl)cyclopropylidene]methyl}adenine (ent-13a). The reaction
was performed as described above using the Z,E-isomers ent-28a
(300 mg, 0.91 mmol) and K2CO3 (250 mg, 1.81 mmol) in methanol
(18 mL) to give the Z-isomer ent-12a (94 mg, 42%) and E-isomer
ent-13a (80 mg, 36%).
(-)-2-Amino-6-chloro-9-{(Z)-(R)-2-[(R)-1,2-dihydroxyethyl)-
cyclopropylidene]methyl}purine (ent-12c) and (-)-2-Amino-6-
chloro-9-{(E)-(R)-2-[(R)-1,2-dihydroxyethyl)cyclopropylidene]-
methyl}purine (ent-13c). The above procedure starting from the
Z,E-isomers ent-28b (340 mg, 0.93 mmol) afforded after chroma-
tography in AcOEt-MeOH (30:1 to 20:1) the Z-isomer ent-12c
(95 mg, 36%) followed by E-isomer ent-13c (130 mg, 50%).
Z-Isomer ent-12c: mp 210-212 °C, [R]2D8 -87.4° (c 0.85, DMF).
E-Isomer ent-13c: mp 250-251 °C, [R]2D8 -25.3° (c 0.67, DMF).
(+)-2-Amino-6-chloro-9-{(Z)-(S)-2-[(R)-1,2-dihydroxyethyl)cy-
clopropylidene]methyl}purine (14c) and (+)-2-Amino-6-chloro-
9-{(E)-(S)-2-[(R)-1,2-dihydroxyethyl)cyclopropylidene]methyl}-
purine (15c). The procedure described above was performed with
the Z,E-isomers 29b (320 mg, 0.87 mmol) to give the faster moving
Z-isomer 14c (85 mg, 34%) and slower moving E-isomer 15c (110
mg, 45%).
Z-Isomer ent-12a: mp 273-274 °C, [R]2D5 -56.6° (c 1.13,
DMSO). Anal. (C11H13N5O2) C, H, N.
E-Isomer ent-13a: mp 251-252 °C, [R]2D6 5.0° (c 0.71, DMSO).
Anal. (C11H13N5O2) C, H, N.
(+)-9-{(Z)-(S)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}adenine (14a) and (-)-9-{(E)-(S)-2-[(R)-1,2-Dihydroxy-
ethyl)cyclopropylidene]methyl}adenine (15a). The reaction was
performed as in the previous experiments using the Z,E-isomers
29a (260 mg, 0.7 mmol) to give the Z-isomer 14a (68 mg, 35%)
and E-isomer 15a (93 mg, 48%).
Z-Isomer 14a: mp 217-219 °C, [R]2D7 94.7° (c 0.46, DMSO). UV
1
λmax 278 nm (ε 8700), 261 (ε 12 800), 224 (ε 26 900). H NMR δ
Z-Isomer 14c: mp 205-207 °C, [R]2D4 41.3° (c 0.8, DMF). UV
1.35-1.43 (m, 2H, H3′), 2.10-2.15 (m, 1H, H4′), 3.22-3.32 (m, 2H,
H6′), 3.53-3.58 (m, 1H, H5′), 4.66 (t, 1H, J ) 5.6 Hz, 6′-OH), 4.91
(d, 1H, J ) 4.8 Hz, 5′-OH), 7.32 (bs, 2H, NH2), 7.36 (d, 1H, J ) 1.6
Hz, H1′), 8.16 (s, 1H, H2), 8.46 (s, 1H, H8). 13C NMR 5.8 (C3′), 21.2
(C4′), 66.0 (C6′), 71.3 (C5′), 110.3, 116.5, 119.0, 138.6, 148.9, 153.6,
156.7 (adenine, C2′, C1′). ESI-MS 248 (M + H, 100.0), 270 (M + Na,
11.2). Anal. (C11H13N5O2) C, H, N.
1
λmax 311 nm (ε 7900), 234 (ε 31 700). H NMR δ 1.34-1.43 (m,
2H, H3′), 2.10-2.15 (m, 1H, H4′), 3.20-3.31 (m, 2H), 3.47-3.50
(m, overlapped with H2O, H5′, H6′), 4.63 (t, J ) 5.2 Hz, 6′-OH),
4.86 (d, 1H, J ) 4.0 Hz, 5′-OH), 7.00 (s, 2H, NH2), 7.19 (d, 1H,
J ) 1.6 Hz, H1′), 8.41 (s, 1H, H8). 13C NMR 6.0 (C3′), 21.3 (C4′),
66.0 (H6′), 71.4 (H5′), 110.9, 117.5, 123.7, 140.9, 150.2, 153.3, 160.7
(purine, C2′, C1′). ESI-MS (MeOH + AcOK) 282, 284 (M + H,
16.6, 5.0), 320, 322 (M + K, 100.0, 41.4).
E-isomer 15a: mp 233-235 °C, [R]2D7 -5.8° (c 0.43, DMSO).
UV λmax 278 nm (ε 8600), 262 (ε 11 700), 227 (ε 26 400). 1H NMR
δ 1.43-1.47 (m), 1.68 (dt, 2H, J ) 8.8, 2.4 Hz, H3′), 1.78-1.84
(m, 1H, H4′), 3.04-3.09 (m, 1H), 3.42-3.45 (m, 2H, H5′, H6′), 4.69
(t, 1H, J ) 4.8 Hz, 6′-OH), 4.90 (d, 1H, J ) 4.8 Hz, 5′-OH), 7.33
(s, 2H, NH2), 7.42 (s, 1H, H1′), 8.15 (s, 1H, H2), 8.47 (s, 1H, H8).
13C NMR 9.6 (C3′), 19.8 (C4′), 66.4 (C6′), 74.2 (C5′), 111.3, 116.0,
E-Isomer 15c: mp 243-245 °C, [R]2D5 ) 51.8 ° (c 1.1, DMF).
1
UV λmax 311 nm (ε 8000), 234 (ε 32 200). H NMR δ 1.42-1.45
(m), 1.69 (t, J ) 8.8 Hz, 2H, H3′), 1.77-1.82 (m, 1H, H4′), 3.03
(m, 1H), 3.50 (m, partly overlapped with H2O, 2H, H5′, H6′), 4.67
(poorly resolved t, 1H, 6′-OH), 4.89 (d, J ) 4.8 Hz, 5′-OH), 7.02

Methylenecyclopropane Analogues
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10 3405
(s, 2H, NH2), 7.25 (s, 1H, H1′), 8.43 (s, 1H, H8). 13C NMR 9.7
(C3′), 20.0 (C4′), 66.4 (C6′), 74.2 (C5′), 111.0, 116.9, 123.7, 140.2,
150.3, 153.1, 160.7 (purine, C2′, C1′). ESI-MS 304, 306 (M + Na+,
100.0, 32.5), 585, 587 (2 M + Na, 57.4, 39.9).
(+)-9-{(E)-(S)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (15b). The procedure described above was
performed with the E-isomer 15c (100 mg, 0.35 mmol) to give the
title compound 15b (78 mg, 83%): mp >300 °C, [R]2D7 17.5° (c
0.31, DMSO). UV λmax 270 nm (ε 9400), 231 (ε 24 300). 1H NMR
δ 1.35-1.40 (m, 1H), 1.64 (poorly resolved dt, 1H, H3′), 1.72-1.77
(m, 1H, H4′), 2.97-3.03 (m, 1H), 3.37-3.46 (m, partly overlapped
with H2O, 2H, H5′, H6′), 4.63 (t, 1H, J ) 5.2 Hz, 6′-OH), 4.85 (d,
1H, J ) 4.0 Hz, 5′-OH), 6.54 (s, 2H, NH2), 7.17 (s, 1H, H1′), 8.04
(s, 1H, H8), 10.64 (s, 1H, NH). 13C NMR 9.5 (C3′), 19.8 (C4′), 66.4
(C6′), 74.3 (C5′), 111.3, 115.5, 116.9, 134.4, 150.5, 154.6, 157.4
(guanine, C2′, C1′). ESI-MS 264 (M + H, 47.0), 286 (M + Na,
100.0). Anal. (C11H13N5O3 ·0.5H2O) C, H, N.
(-)-2-Amino-6-chloro-9-{(Z)-(R)-2-[(S)-1,2-dihydroxyethyl)-
cyclopropylidene]methyl}purine (ent-14c) and (-)-2-Amino-6-
chloro-9-{(E)-(R)-2-[(S)-1,2-dihydroxyethyl)cyclopropylidene]me-
thyl}purine (ent-15c). The experiment described above was
performed with the Z,E-enantiomers ent-29b (300 mg, 0.82 mmol)
to furnish the Z-isomer ent-14c (94 mg, 41%) and E-isomer ent-
15c (106 mg, 46%).
Z-Isomer ent-14c: mp 199-200 °C, [R]D27 -39.3° (c 0.75,
DMF).
E-Isomer ent-15c: mp 248-250 °C, [R]2D7-49.7° (c 1.0, DMF).
(-)-9-{(E)-(R)-2-[(S)-1,2-dihydroxyethyl)cyclopropylidene]-
methyl}guanine (ent-15b). The procedure described above was
performed with the E-isomer ent-15c (95 mg, 0.34 mol) to give
the title compound ent-15b (75 mg, 84%): mp >300 °C, [R]D27
-19.4° (c 0.41, DMSO). Anal. (C11H13N5O3 ·0.4H2O) C, H, N.
Benzyl (1S,2R,5S)-5-Bromo-3-oxabicyclo[3.1.0]hexane-2-car-
boxylate (30). Pyridinium tribromide (85 mg, 0.27 mmol) was
added with stirring to a solution of (1S,2S)-ester 20 (50 mg, 0.23
mmol) in CH2Cl2 (1 mL) at 0 °C. The stirring was continued for
1 h at room temperature. The mixture was then filtered through a
silica gel pad (1 cm), which was washed with CH2Cl2 (10 mL).
The solvent was evaporated, and the crude dibromide 32 and K2CO3
(50 mg, 0.36 mmol) were refluxed in THF (1.5 mL) with stirring
for 5 min. After cooling, the mixture was chromatographed on a
silica gel column in hexanes-Et2O (10:1) to furnish compound 30
(22 mg, 32%) as a colorless oil. [R]2D5 -17.1° (c 0.70, CHCl3). 1H
NMR (CDCl3 δ 1.14 (t, J ) 5.6 Hz), 1.38 (dd, 2H, J ) 8.0 Hz,
(+)-9-{(Z)-{(S)-2-[(S)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (12b). A solution of the Z-isomer 12c (85 mg,
0.3 mmol) in 80% formic acid (80%, 8 mL) was heated at 80 °C
for 4 h. The volatiles were evaporated in vacuo, the crude product
was dissolved in NH3 in methanol (5%, 40 mL), and the mixture
was stirred for 30 min at 0 °C. After removal of solvents, the solid
was recrystallized from NH4OH (28%) to give the guanine Z-isomer
12b (66 mg, 83%): mp >300 °C, [R]2D7 98.6° (c 0.56, DMSO). UV
λ
max 271 nm (ε 10 000), 231 (ε 28 100). 1H NMR δ 1.22 (m), 1.42
(t, 2H, J ) 8.0 Hz, H3′), 1.94 (m, 1H, H4′) 3.10 (m, 1H), 3.39-3.46
(m, 2H, partly overlapped with H2O, H5′, H6′), 4.68 (bs, 1H, 6′-
OH), 5.10 (bs, 1H, 5′-OH), 6.51 (s, 2H, NH2), 7.10 (s, 1H, H1′),
8.56 (s, 1H, H8), 10.64 (bs, 1H, NH). 13C NMR 6.6 (C3′), 20.7
(C4′), 66.5 (C6′), 75.0 (C5′), 110.8, 115.1, 116.8, 135.4, 150.3, 154.5,
157.4 (guanine, C2′, C1′). ESI-MS 264 (M + H, 100.0), 286 (M +
Na, 65.7). Anal. (C11H13N5O3 ·0.5H2O) C, H, N.
H3), 2.10 (dd, 1H, J ) 9.0, 5.0 Hz, H2), 4.23, 4.10 (AB, 2H, JAB
)
(-)-9-{(Z)-(R)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (ent-12b). The procedure described above was
performed with the Z-isomer ent-12c (90 mg, 0.32 mol) to give
the title compound ent-12b (73 mg, 87%): mp >300 °C, [R]D27
-101.7° (c 0.52, DMSO). Anal. (C11H13N5O3 ·0.45H2O) C, H, N.
7.8 Hz, H5), 4.31 (s, 1H, H1), 5.26, 5.20 (AB, 2H, JAB ) 12.0 Hz,
CH2Ph), 7.37 (m, 5H, Ph). 13C NMR 18.4 (C6), 29.1 (C1), 30.1
(C5), 67.2 (C4), 75.0 (CH2Ph), 78.1 (C2), 128.4, 128.7, 128.9, 135.5
(Ph), 171.5 (CdO). ESI-MS 319, 321 (M + Na, 100.0, 100.0),
615, 617, 619 (2M + Na, 38.0, 76.7, 40.0). Anal. (C13H13BrO3) C,
H.
(+)-9-{(E)-(S)-2-[(S)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (13b). The procedure described above was
performed with the E-isomer 13c (90 mg, 0.32 mmol) to give the
title compound 13b (71 mg, 84%): mp >300 °C, [R]2D7 8.8° (c 0.8,
Benzyl (1R,2R,5S)-5-Bromo-3-oxabicyclo[3.1.0]hexane-2-car-
boxylate (31). The above-described protocol was followed with
(1S,2R)-ester 21 (85 mg, 0.39 mmol) via dibromide 33 to give
compound 31 (40 mg, 35%) as a colorless oil: [R]2D6 48.5° (c 0.86,
CHCl3). 1H NMR (CDCl3) δ 1.23 (t, 1H, J ) 7.6 Hz), 1.34 (t, 1H,
J ) 5.8 Hz, H6, H6′), 2.19 (ddd, 1H, J ) 8.0, 4.4, 1.6 Hz, H1),
4.24, 3.98 (AB, 2H, JAB ) 8.6 Hz, H4), 4.67 (d, 1H, J ) 3.2 Hz,
H2), 5.20 (d, 2H, J ) 1.6 Hz, CH2Ph), 7.35 (m, 5H, Ph). 13C NMR
15.5 (C6), 28.6 (C1), 30.7 (C5), 67.2 (C4), 75.6 (CH2Ph), 76.9 (C2),
128.5, 128.7, 128.9, 135.5 (Ph), 169.4 (CdO). ESI-MS 319, 321
(M + Na, 100.0, 97.4), 615, 617, 619 (2M + Na, 23.7, 48.7, 25.7).
Anal. (C13H13BrO3) C, H.
1
DMSO). UV λmax 271 nm (ε 10 900), 231 (ε 28 200). H NMR δ
1.36-1.40 (m), 1.61 (dt, 2H, J ) 8.8, 1.6 Hz, H3′), 1.79-1.84 (m,
1H, H4′) 3.17 (m, 1H), 3.40-3.45 (m, partly overlapped with H2O,
2H, H5′, H6′), 4.62 (t, 1H, J ) 4.8 Hz, 6′-OH), 4.78 (d, 1H, J ) 4.8
Hz, 5′-OH), 6.53 (s, 2H, NH2), 7.24 (s, 1H, H1′), 8.02 (s, 1H, H8),
10.67 (bs, 1H, NH). 13C NMR 8.5 (C3′), 18.8 (C4′), 66.3 (C6′), 73.4
(C5′), 110.8, 115.5, 116.9, 134.4, 150.5, 154.6, 157.4 (guanine, C2′,
C1′). ESI-MS 264 (M + H, 81.4), 286 (M + Na, 100.0). Anal.
(C11H13N5O3 ·0.5H2O).
(-)-9-{(E)-(R)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (ent-13b). The procedure described above was
performed with the E-isomer ent-13c (110 mg, 0.39 mol) to give
the title compound ent-13b (86 mg, 83%): mp >300 °C, [R]D27
-10.8° (c 0.88, DMSO). Anal. (C11H13N5O3 ·0.45H2O) C, H, N.
(Z)-9-{(R)-2-[(R)-1,2-Diacetoxyethyl)cyclopropylidene]meth-
yl}adenine (34). A mixture of the Z-isomer ent-12a (16 mg, 0.065
mmol) and Ac2O (0.3 mL) in pyridine (1 mL) was stirred for 5 h
at room temperature. Solvent was removed at room temperature
and 0.04 torr. The crude product was chromatographed on a silica
gel column in AcOEt-methanol (50:1 to 20:1) to give compound
34 (16 mg, 75%). Single crystals for X-ray diffraction were obtained
by slow crystallization of a solution in AcOEt: mp 185-186 °C,
[R]2D5 -152.6° (c 1.03, MeOH). UV λmax 278 nm (ε 8600), 261 (ε
12 500), 227 (ε 28 200). 1H NMR (CDCl3) δ 1.43-1.46 (m), 1.70
(dt, 2H, J ) 8.8, 1.6 Hz, H3′), 1.92, 2.06 (2s, 6H, CH3), 2.25-2.31
(m, 1H, H4′), 4.43, 4.17 and 4.42, 4.18 (2AB, 2H, JAB ) 11.6 Hz,
H6′), 4.67 (m, 1H, H5′), 5.97 (bs, 2H, NH2), 7.49 (d, 1H, J ) 1.6
Hz, H1′), 8.38 (s, 2H, H2, H8). 13C NMR 7.6 (C3′), 17.6 (C4′), 21.0
(CH3), 64.6 (C6′), 74.5 (C5′), 112.1, 113.1, 119.5, 137.7, 149.1,
153.8, 155.8 (adenine, C1′, C2′), 170.0, 170.8 (CdO). ESI-MS: 332
(M + H, 68.9), 354 (M + Na, 100.0), 53.0, 685 (2M + Na, 55).
Anal. (C15H17N5O4) C, H, N.
(+)-9-{(Z)-(S)-2-[(R)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (14b). The above protocol was performed with
the Z-isomer 14c (80 mg, 0.28 mmol) to give the title compound
14b (64 mg, 86%), mp >300 °C, [R]2D7 62.4° (c 0.53, DMSO). UV
1
λmax 271 nm (ε 9700), 230 (ε 22 800). H NMR δ 1.30-1.38 (m,
2H, H3′), 2.05-2.09 (m, 1H, H4′), 3.25-3.31 (m, 2H, partially
overlapped with H2O), 3.51 (m, 1H, H5′, H6′), 4.64 (s, 1H, 6′-OH),
4.85 (s, 1H, 5′-OH), 6.53 (s, 2H, NH2), 7.09 (d, 1H, J ) 1.6 Hz,
H1′), 8.01 (s, 1H, H8), 10.71 (bs, 1H, NH). 13C NMR 5.7 (C3′),
20.9 (C4′), 66.0 (C6′), 71.4 (C5′), 111.2, 116.0, 116.9, 135.1, 150.6,
154.6, 157.5 (guanine, C2′, C1′). ESI-MS 264 (M + H, 100.0), 286
(M + Na, 39.6). Anal. (C11H13N5O3 ·0.5H2O) C, H, N.
(-)-9-{(Z)-(R)-2-[(S)-1,2-Dihydroxyethyl)cyclopropylidene]-
methyl}guanine (ent-14b). The procedure described above was
performed with the E-isomer ent-13c (85 mg, 0.3 mol) to give the
title compound ent-14b (68 mg, 86%): mp >300 °C, [R]2D7 -62.3°
(c 0.55, DMSO). Anal. (C11H13N5O3 ·0.15H2O) C, H, N.
Antiviral Assays. Antiviral assays were performed as described
previously.29,30 HCMV assays were run in human foreskin
fibroblast (HFF) cell culture with two strains of virus, Towne
and AD169, in a plaque reduction or cytopathic effect (CPE)

3406 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 10
Zhou et al.
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inhibition assay. Virus strains resistant to ganciclovir (10) and
cyclopropavir (3b) were isolated from Towne strain by growth
in synadenol (1a) (strain E8)23 or in 3b (isolate 2696r).24 The
former strain has two point mutations introduced into gene UL97;
the latter has a truncated UL97 gene. The MCMV was assayed
in mouse embryonic fibroblast (MEF) cells by plaque reduction.
The EBV DNA hybridization assay was run in Akata cells.30,31
Adenosine Deaminase (ADA) Assay.29 Adenine analogues
12a-15a and ent-12a to ent-15a (2 µmol) and ADA from calf
intestine (1.7 units) were magnetically stirred in 0.05 M Na2HPO4
(pH 7.5) in a total volume of 0.6 mL. Aliquots were removed
and diluted with buffer. UV spectra were recorded, and a
decrease of absorbance at 260 nm was followed. The results
are summarized in Table 7.
Acknowledgment. We thank M. J. Heeg from the Depart-
ment of Chemistry, Wayne State University, for X-ray
crystallographic studies and L. M. Hryhorczuk from the
Central Instrumentation Facility, Department of Chemistry,
Wayne State University, for mass spectra. We also thank
Kathy Borysko and Julie Breitenbach of the University of
Michigan for isolation and engineering of drug-resistant
HCMV and for the antiviral assays. The work described
herein was supported by U.S. Public Health Service Grant
RO1-CA32779 and Contract NO1-AI-30049 from the Na-
tional Institutes of Health, Bethesda, Maryland.
Supporting Information Available: Crystallographic data of
diacetate 34 (Tables 1-6) and elemental analysis results of all
biologically tested compounds and some key intermediates
(Table 7). This material is available free of charge via the Internet
at http://pubs.acs.org.
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Z-Isomers of 2-hydroxymethylcyclopropylidenemethyl adenine (syn-
adenol) and guanine (synguanol) are active against ganciclovir- and
foscarnet-resistant human cytomegalovirus UL97 mutants. AntiViral
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