Fluoroanalogues of anti-cytomegalovirus agent cyclopropavir: Synthesis and antiviral activity of (E)- and (Z)-9-{[2,2-bis(hydroxymethyl)-3- fluorocyclopropylidene]methyl}-adenines and guanines

Article abstract of DOI:10.1080/15257770701257210

Synthesis of fluorinated cyclopropavir analogues 13a, 13b, 14a, and 14b is described starting from alkene 15. Addition of carbene derived from dibromofluoromethane gave bromofluoro cyclopropane 16. Reduction (compound 17) followed by desilylation gave int

Full text of DOI:10.1080/15257770701257210

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Fluoroanalogues of Anti-Cytomegalovirus
Agent Cyclopropavir: Synthesis
and Antiviral Activity of (E)- and
(Z)-9-{[2,2-Bis(Hydroxymethyl)-3-
Fluorocyclopropylidene]Methyl}-
Adenines and Guanines
Shaoman Zhou a , Jiri Zemlicka a , Earl R. Kern b & John C. Drach c
a Developmental Therapeutics Program, Barbara Ann Karmanos
Cancer Institute, Wayne State University School of Medicine,
Detroit, Michigan, USA
b Department of Pediatrics, The University of Alabama School of
Medicine, Birmingham, Alabama, USA
c Department of Biologic and Materials Sciences, School of Dentistry,
University of Michigan, Ann Arbor, Michigan, USA
Published online: 04 Apr 2007.
To cite this article: Shaoman Zhou , Jiri Zemlicka , Earl R. Kern & John C. Drach (2007)
Fluoroanalogues of Anti-Cytomegalovirus Agent Cyclopropavir: Synthesis and Antiviral Activity of
(E)- and (Z)-9-{[2,2-Bis(Hydroxymethyl)-3-Fluorocyclopropylidene]Methyl}-Adenines and Guanines,
Nucleosides, Nucleotides and Nucleic Acids, 26:3, 231-243, DOI: 10.1080/15257770701257210
To link to this article: http://dx.doi.org/10.1080/15257770701257210
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Nucleosides, Nucleotides, and Nucleic Acids, 26:231–243, 2007
Copyright ꢀC Taylor & Francis Group, LLC
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770701257210
FLUOROANALOGUES OF ANTI-CYTOMEGALOVIRUS AGENT
CYCLOPROPAVIR: SYNTHESIS AND ANTIVIRAL ACTIVITY OF (E)-
AND (Z)-9-{[2,2-BIS(HYDROXYMETHYL)-3-
FLUOROCYCLOPROPYLIDENE]METHYL}-ADENINES
AND GUANINES
Shaoman Zhou and Jiri Zemlicka
Developmental Therapeutics Program,
2
Barbara Ann Karmanos Cancer Institute, Wayne State University School of Medicine,
Detroit, Michigan, USA
Earl R. Kern
Department of Pediatrics, The University of Alabama School
2
of Medicine, Birmingham, Alabama, USA
John C. Drach Department of Biologic and Materials Sciences, School of Dentistry,
2
University of Michigan, Ann Arbor, Michigan, USA
Synthesis of fluorinated cyclopropavir analogues 13a, 13b, 14a, and 14b is described
2
starting from alkene 15. Addition of carbene derived from dibromofluoromethane gave bromofluoro
cyclopropane 16. Reduction (compound 17) followed by desilylation gave intermediate 18, which
was transformed to 2-nitrophenylselenenyl derivative 19. Oxidation to selenoxide 20 was followed by
β-elimination to afford methylenecyclopropane 21. Addition of bromine provided compound 22 for
alkylation-elimination of adenine and 2-amino-6-chloropurine. The resultant E,Z isomeric mixtures
of methylenecyclopropanes 23a + 24a and 23c + 24c were resolved and the individual isomers
were deprotected to give adenine analogues 13a and 14a as well as compounds 13c and 14c.
Hydrolytic dechlorination of 13c and 14c furnished guanine analogues 13b and 14b. The only
significant antiviral effects were observed with analogue 13a against HCMV and 14a against VZV
in cytopathic inhibition assays.
Keywords 3-Fluoromethylenecyclopropanes; cyclopropavir; nucleoside analogues;
antivirals
Received 14 July 2006; accepted 3 November 2006.
We thank L. M. Hrihorczuk from the Central Instrumentation Facility, Department of Chemistry,
Wayne State University (D.M. Coleman, Director) for mass spectra. We also thank research assistants
and associates in Drs. Drach and Kern laboratories for expert performance of antiviral and cytotoxicity
assays. The work described herein was supported by U.S. Public Health Service grants RO1-CA32779
(J.Z.) from the National Cancer Institute, contracts NO1-AI85347, NO1-AI30049 (E.R.K.) and program
project PO1-AI46390 (J.C.D.) from the National Institute of Allergy and Infectious Diseases, National
Institutes of Health, Bethesda, Maryland 20892.
Address correspondence to Jiri Zemlicka, Barbara Ann Karmanos Cancer Institute, 110 E. Warren
Ave., Detroit, MI 48201-1379. E-mail: zemlicka@karmanos.org
231

232
S. Zhou et al.
INTRODUCTION
Among methylenecyclopropane analogues of nucleosides, the purine
Z-isomers 1 and 3 were identified as effective antivirals whereas the
E-isomers 2 and 4 were active in only a few cases.[1,2] The most po-
tent analogue,[3,4] cyclopropavir (3b), is presently a subject of mecha-
nism of action studies[5–8] and preclinical development as a therapeutic
agent against human cytomegalovirus (HCMV). It also is effective against
Epstein-Barr virus (EBV),[3] and human herpes virus 6 and 8 (HHV-6
and HHV-8).[3,6] A considerable attention also has been paid to fluori-
nated methylenecyclopropanes. The geminal difluoro analogues 5a, 5b,
6a, and 6b are with the exception of 5a devoid of antiviral activity.[9]
By contrast, monofluoro derivatives 7a through 12a and 7b through
12b exhibit varying degree of antiviral[10–12] effects particularly against
HCMV (7a), Epstein-Barr virus (EBV, 7a), varicella zoster virus (VZV, 9a)
and HIV-1 (7a and 9a). Some antiviral activity has been noted with the
trans-isomers 8b and 12a. Consequently, the synthesis of fluorinated 2,2-
bis(hydroxymethyl)methylenecyclopropanes 13a, 13b, 14a, and 14b related
to cyclopropavir (3b) was of interest.
RESULTS AND DISCUSSION
Synthesis
The synthesis of these analogues commenced with addition of carbene
derived from CHBr2F to alkene 15 in CH2Cl2 under phase-transfer condi-
tions (Bu4NI and 50% NaOH) to give bromofluorocyclopropane 16 (72%,
Scheme 1). Reduction with Bu3SnH using AIBN as a catalyst gave fluoro
derivative 17 in 83% yield. The TBDMS group was removed using Bu4NF in
THF to give hydroxymethylcyclopropane 18 in 94% yield. These procedures
followed a similar sequence that started from chlorofluorocyclopropane 16
(Br Cl).[13] Reaction with 2-nitrophenyl selenocyanate and Bu3P in THF[9]
afforded intermediate 19 (91%). Oxidation with H2O2 in THF furnished
crude selenoxide 20 which was converted to methylenecyclopropane 21
(57%) by a thermolysis at 80◦C in toluene. Addition of bromine via
pyridinium perbromide in CH2Cl2 gave dibromo derivative 22 in 91%
yield.
Alkylation-elimination of adenine (K2CO3, DMF, 100◦C) with 22 pro-
vided, after chromatographic separation, the E- and Z-isomers 23a and 24a
in 21 and 35% yield, respectively. The separated isomers 23a and 24a were
debenzylated using BCl3·SMe2 complex in CH2Cl2 to give compounds 13a
and 14a (83 and 84% yield). In a similar fashion, alkylation-elimination of 2-
amino-6-chloropurine with dibromo derivative 22 gave the E- and Z-isomers
23c and 24c in 17.5 and 37% yield, respectively. Somewhat surprisingly, the

Fluoroanalogues of Cyclopropavir
233
Chart 1
SCHEME 1

234
S. Zhou et al.
TABLE 1 Chemical shifts (δ) of the relevant 1H NMR signals of
fluorinated methylenecyclopropanes 13a, 14a, 13b, 14b, 13c, and 14c
Compounda
H8
OH
ꢁ
H1
13a
14a
13b
14b
13c
14c
7.89
7.53
7.55
7.26
7.69
7.37
8.82
8.34
8.42
7.90
8.79
8.30
5.18, 5.22
4.89
5.23
4.87
5.20
4.90
aCD3SOCD3 as solvent. For numbering of signals see Table 2.
yields of the E(cis) isomers 23a, 23c are lower than those of the Z(trans)
isomers 24a, 24c. Debenzylation of 23c and 24c furnished compounds 13c
(86%) and 14c (84%). Hydrolytic dechlorination using 80% formic acid at
80◦C gave guanine analogues 13b and 14b in 90.5 and 85% yield.
E- and Z-Isomeric Assignment
Methylenecyclopropane nucleoside analogues with a cis-configuration
of the nucleobase and hydroxymethyl groups 1 and 3 are always less polar
moving faster on silica gel than the trans-configured isomers[1,2] 2 and 4.
1
In the H NMR spectra, the H8 of purine cis-isomers 1 and 3 are more
deshielded than the corresponding signals of the trans-isomers 2 and 4. Both
trends (polarity and H8 chemical shifts) are preserved in the fluorinated
analogues[9,10,12] 5 through 12. It is then not surprising that similar patterns
of the H8 chemical shifts were also found in analogues 13 and 14 (Table 1).
Interestingly, the OH groups of 13a are not equivalent. In addition, the
OH signals of the E(cis) isomers 13 are more deshielded than those of
the Z(trans) isomers 14 as also found in fluorinated and nonfluorinated
methylenecyclopropanes alike.[3,9,10,12] The H1 chemical shifts of 13 and 14
ꢁ
then follow the trend observed in 3ꢁ-fluorinated analogues[9,12] 5, 6, and
ꢁ
9 through 12. The H1 configured cis relative to F in the E-isomer 13 is
more deshielded than in Z-isomer 14 where the orientation of both atoms
is trans. An unambiguous confirmation of the isomeric structure came from
the nuclear Overhauser effect (NOE) experiments with adenine analogues
13a and 14a (Table 2). As expected, the NOE enhancements were observed
ꢁ
between the H8 in anti-like conformation and cis-orientated OH and H5
ꢁ
ꢁ
protons of the E-isomer 13a. By contrast, the interactions H8 − H3 and H5
ꢁ
− H1 were strongest in the Z-isomer 14a.
Antiviral Activity
To our surprise, introduction of a fluorine atom in the 3ꢁ position
(compound 13b) abolished the antiviral activity of cyclopropavir (3b). Some

Fluoroanalogues of Cyclopropavir
235
TABLE 2 The NOE enhancements of relevant 1H NMR signals of fluorinated
adenine methylenecyclopropanes 13a and 14a
Compounda
Hiir
δ
Hobs
OH
δ
NOE (%)
13a
H8
H8
OH
8.82
8.82
5.18–5.22
3.53, 3.74
8.82
2.37
ꢁ
H5
H8
H8
H3
H8
H1
1.16, 2.37
2.79, 2.89
2.13
3, 3.2
4.10, 2.48
1.32, 1.60
5.18, 5.22
3.74
ꢁ
H5
H8
H3
H5
8.82
ꢁ
ꢁ
14a
8.34
5.34, 5.51a
8.34
5.34, 5.51b
3.53, 3.64
ꢁ
ꢁ
7.53
aBoth halves of the doublet were observed separately.
bBoth halves of the doublet were irradiated separately.
antiviral effects were seen only in adenine analogues 13a and 14a. Thus, the
Z-isomer 13a had a moderate anti-HCMV effect in Towne strain of the virus
(Table 3). The Z-isomer 13a and E-isomer 14a were effective against AD169
strain of HCMV (EC50 1.8 µM) and VZV (EC50 2.7 µM), respectively, but
TABLE 3 Anti-herpesvirus activity of 2,2-bis(hydroxymethyl)-3-fluoromethylenecyclopropane
analogues of nucleosides
EC50/CC50 (µM)
HCMV/HFF
Townec,d
EBV/Akataa
VZV/HFFb
Compound
AD169e,f
13a
14a
13b
14b
51/>100
>100/>100
>100/>100
>100/>100
2.5/>100i
1.8/170g
>60/>300
>300/260
>300/197
0.05/>100i
84.5/>100
97.3/>100
>100/>100
>100/>100
14/j
>60
2.7h
>300
238.5
0.25j
Control
aDNA hybridization assay.
bFor cytotoxicity in HFF cells see HCMV/HFF (AD169).
cPlaque reduction assay.
dVisual cytotoxicity.
eCytopathic effect (CPE) inhibition assay in stationary HFF cells.
fCytotoxicity by neutral red uptake.
gThe EC50 in plaque reduction assay was >20 µM.
hThe EC50 in plaque reduction assay was >100 µM.
iGanciclovir.
jAcyclovir.

236
S. Zhou et al.
only in cytopathic effect inhibition assays. In both cases, no activity in plaque
reduction assays was seen. Little effect was also observed against EBV and all
analogues were inactive against HSV-1, HSV-2, HBV, or HIV-1. It is possible
that compounds 13a, 13b, 14a and 14b are not effective substrates for
activation enzymes responsible for phosphorylation or their triphosphates
are not capable of inhibiting the viral DNA polymerase. Isomers 12a and
13a were not substrates for adenosine deaminase.
CONCLUSION
3-Fluoromethylenecyclopropane analogues 13a, 13b, 14a, and 14b were
prepared and their antiviral activity was investigated. In contrast to nonflu-
orinated analogues 3a and 3b, they were devoid of significant anti-herpetic
activity.
EXPERIMENTAL SECTION
General Methods
The UV spectra were measured in ethanol and NMR spectra were
determined at 300 or 400 MHz (1H), 75 or 100 MHz (13C), and 376 MHz
(19F) in CD3SOCD3 unless stated otherwise. For 19F NMR, CFCl3 was used
as a reference. Mass spectra were determined in electron-impact (EI-MS) or
electrospray ionization (ESI-MS, methanol-NaCl) mode.
Reagents
For abbreviations of common reagents and protecting groups see
the Journal of. Organic Chemistry, volume 70, pages 26A–27A (2005).
Dibromofluoromethane,[14] diphenyl diselenide,[15] and 2-nitrophenyl
selenocyanate[16] were prepared as described.
(cis,trans)-2,2-Bis(benzyloxymethyl)-3-bromo-1-tert-butyldimethylsilylox-
ymethyl-3-fluorocyclopropane (16).[17] The protocol for the corresponding
chloro derivative[13] (16, Br = Cl) was modified as follows. A mixture
of CHBr2F (8.64 g, 45 mmol), alkene 15 (12.4 g, 30 mmol) and NBu4I
(1.11 g, 3.0 mmol) in CH2Cl2 (22 mL) and 50% aqueous NaOH (22 mL)
was stirred at room temperature for 12 hours. Water (100 mL) was then
added at 0◦C, the organic layer was separated and the aqueous portion
was extracted with CH2Cl2 (5 × 40 mL). The combined organic phase
was washed with saturated NaCl and it was dried (MgSO4). The solvent
was evaporated and the crude product was chromatographed on a silica
gel column in hexanes-ether (60 : 1 to 10 : 1) to give compound 16 as a
1
colorless oil (11.30 g, 72%). H NMR (CDCl3) δ 0.05–0.07 (m, 6H, SiMe2,
0.88 and 0.89 (2s, 9 H, Me of t-Bu), 1.60, 1.64–1.76 (2m, 1H, H1), 3.49–3.91

Fluoroanalogues of Cyclopropavir
237
(cluster of m, 6 H, CH2O), 4.43–4.57 (m, 4 H, CH2Ph), 7.26–7.35 (m, 10H,
Ph). 13C NMR −5.2, −5.1, −5.0 (3s, SiMe2, 18.4 (quaternary C of t-Bu),
26.1 (Me of t-Bu), 34.4, 34.9 (2d, J = 8.2, 8.6 Hz), 35.7, 37.4 (2d, J = 9.7 Hz,
C1, C2), 57.2 (d, J = 7.5 Hz), 60.9, 64.0 (d, J = 9.0 Hz), 68.2, 68.27, 68.34
(CH2OSi, CH2OBn), 72.5, 72.8, 73.1, 73.3, 73.5 (CH2Ph), 91.3 (d, J = 304.4
Hz, C3), 127.78, 127.83, 127.9, 128.0, 128.5, 128.6, 138.3, 138.5 (Ph). 19F
3
NMR −132.68 (d, JF,H-cis = 20.0 Hz, cis-isomer), −149.57 (s, trans-isomer).
ESI-MS 523, 525 (M + H, 5.1, 5.3), 545, 547 (M + Na, 95.0, 100.0).
(cis, trans)-2,2-Bis(benzyloxymethyl)-3-fluoro-1-(2-nitrophenylselenen-
ylmethyl) cyclopropane (17). The procedure used for reduction of the
corresponding chloro derivative[13] (16, Br = Cl) was modified as follows.
Compound 16 (11.0 g, 21.0 mmol) was heated with a catalytic amount
of AIBN (110 mg, 0.67 mmol) under N2 at 90◦C. Bu3SnH (6.13 mL,
23.1 mmol) was then added and the mixture was stirred for 4 hours. After
cooling, silica gel (4 g) was added and the mixture was put on the top of a
silica gel column. Chromatography was performed in hexanes-ether (100: 1
to 30 : 1) to give compound 17 as a colorless oil (7.74 g, 83%). The 1H, 13
C
and 19F NMR (CDCl3) corresponded to the mixture of E and Z-isomers 7
described[13] as individual components. ESI-MS 445 (M + H, 5.6), 467 (M +
Na, 100.0).
(cis, trans)-2,2-Bis(benzyloxymethyl)-3-fluoro-1-(hydroxymethyl) cyclo-
propane (18). The described procedure[13] was modified as follows. A
solution of compound 17 (7.6 g, 17.11 mmol) and Bu4NF (1.0 M in THF,
20 mL, 20 mmol) in THF (40 mL) was stirred at room temperature for
8 hours. The solvent was evaporated and the residue was chromatographed
on a silica gel column using hexanes-ether (10 : 1 to 1 : 1) to give compound
1
18 as a colorless oil (5.31 g, 94%). The H, 13C and 19F NMR spectra
corresponded to those described.[13] ESI-MS 331 (M + H, 6.3), 353 (M +
Na, 100.0).
(cis, trans)-2,2-Bis(benzyloxymethy)l-3-fluoro-1-(2-nitrophenylselenenyl-
methyl)-cyclopropane (19). A mixture of compound 18 (5.0 g, 15.2 mmol),
2-nitrophenyl selenocyanate (4.14 g, 18.24 mmol) and Bu3P (4.5 mL, 18.24
mmol) in THF (30 mL) was stirred for 4 hours at room temperature. The
solvents were removed and the residue was chromatographed on a silica
gel column using hexanes-EtOAc (10 : 1 to 8 : 1) to give the selenide 19
1
(7.1g, 91%) as a pale yellow oil. H NMR (CDCl3) δ 1.38, 1.55 (m, 1H,
H3), 2.85–3.21 (m, 2H, CH2Se), 3.33–3.56, 3.77–3.89 (2m, 4H, CH2OBn,
4.45–4.73 (m, 5H of Bn, H1), 7.27–7.36 (m, 11H, C6H5), 7.43–7.54m, 2H),
8.29 (d, 1H, J = 8.0 Hz, NO2C6H4, aromatic H’s). 19F NMR −215.33 (dd, J
= 64.0, 21.5 Hz), −232.11 (dd, J = 64.0 Hz, 6.0 Hz). ESI-MS 538 (M + Na,
100.0).
2,2-Bis(benzyloxymethyl)-3-fluoro-1-methylenecyclopropane (21).
A
mixture of selenide 19 (7.0 g, 14.0 mmol) and 30% H2O2 (9.3 mL, 79.1
mmol) in THF (42 mL) was stirred for 14 hours at room temperature. Ethyl

238
S. Zhou et al.
ether (200 mL) was added, the organic phase was washed with water (5 ×
40 mL) and it was dried (MgSO4). The solvent was evaporated to give the
crude selenoxide 20 as a yellow syrup. This product was heated in toluene
(30 mL) at 80◦C for 4 hours. The solvent was evaporated and the residue
was chromatographed on a silica gel column in hexanes-Et2 O (50 : 1 to
30 : 1) to give methylenecyclopropane 21 (2.93 g, 67%) as a colorless oil.
1H-NMR (CDCl3) δ 3.52, 3.84 (split AB, 2H, J = 10.0 Hz), 3.73 (poorly
resolved dd, 2H, J = 8.0), CH2OBn), 4.56 (s, 2H), 4.59 (AB, 2H, J = 11.6
Hz, CH2 of Bn), 4.95 (d, 1H, J = 68 Hz, H1), 5.77, 5.95 (2s, 2H, CH2),
7.31–7.40 (m, 10H, Ph). 13C NMR 32.6 (d, J = 14.2 Hz, C2), 66.9, 69.6 (d,
J = 4.5 Hz, CH2OBn), 72.5 (d, C3, J = 236.5 Hz), 73.0, 73.2 (CH2 of Bn),
111.4 (d, J = 2.2 Hz, CH2 ), 134.1 (C1), 127.8, 127.9, 128.6, 128.7, 138.5,
138.7 (Ph). 19F NMR −214.02 (d, J = 67.4 Hz). ESI-MS 313 (M + H, 12.0),
335 (M + Na, 100.0). Anal. Calcd for C20H21FO2: C, 76.90; H, 6.78. Found:
C, 76.85; H, 6.98.
(cis, trans)-2,2-Bis(benzyloxymethyl)-1-bromo-1-bromomethyl-3-fluorocy-
clopropane (22). Pyridinium tribromide (4.3 g, 13.5 mmol) was added to a
solution of compound 21 (2.8 g, 8.97 mmol) in CH2Cl2 (40 mL) with stirring
at −20◦C. The reaction mixture was allowed to warm to room temperature
and the stirring was continued for 10 hours. The solvent was evaporated and
the crude product was chromatographed in hexane-Et2 O (50 : 1 to 10 : 1) to
afford compound 22 as a colorless oil (3.92 g, 90.7%, isomeric ratio 1.6/1).
1H NMR (CDCl3) δ 3.75–4.07 (m, 6H, CH2OBn, CH2 Br), 4.53–4.60 (m, 4
H, CH2 of Bn), 4.58, 4.83 (2d, 1H, J = 64.0, 63.2 Hz, H3), 7.35–7.43 (m, 10
H, Ph). 13C NMR 35.7 (d, J = 6.7 Hz), 36.5 (d, J = 8.6 Hz), 37.6 (d, J =
10.4 Hz), 39.1 (C2, C1), 42.6 (d, J = 9.0 Hz), 43.7 (d, J = 9.8 Hz, CH2 Br),
63.6 (d, J = 8.3 Hz), 66.6, 69.8 (d, J = 5.9 Hz), 71.7 (CH2OBn), 73.2, 73.3,
73.6, 73.7 (CH2Ph), 76.7 (d, J = 241.1 Hz), 81.6 (d, J = 245.5 Hz, C3), 128.0,
128.1, 128.2, 128.3, 128.7, 128.8, 128.9, 137.6, 138.0, 138.2, 138.3 (Ph). 19F
NMR −211.55 (d, J = 64.0 Hz), −217.20 (d, J = 64.4 Hz). ESI-MS (MeOH
+ KOAc) 509, 511, 513 (M + K, 28.6, 100.0, 33.3).
(E)- and (Z)-9-{[2,2-Bis(benzyloxymethyl)-3-fluorocyclopropylidene]
methyl}adenine (23a) and (24a). A mixture of adenine (594 mg, 4.4 mmol),
compound 22 (1.92 g, 4.0 mmol) and K2CO3 (3.31 g, 24 mmol) in DMF
(20 mL) was stirred for 6 hours at room temperature under N2 and then
at 100–105◦C for 4 hours. The insoluble solid was filtered off using a silica
gel pad (5 g) which was washed with DMF (70 mL). The solvent from the
filtrate was evaporated in vacuo and the residue was chromatographed in
EtOAc-hexanes (1 : 3 to 2 : 1) to give the faster moving E-isomer 23a (380
mg, 21.3%) followed by Z-isomer 24a (630 mg, 35.4%).
E-Isomer 23a: m.p. 152–153◦C. UV λmax 242 nm (ε 24,400), 281 (ε
1
8,200). H NMR (CDCl3) δ 3.53, 3.93 (AB, 2H, JAB = 10.2Hz), 3.79, 3.84
ꢁ
(AB, 2H, J = 9.8 Hz, 10.0 Hz, H5 ), 4.53, 4.54, 4.55 (3s, 4H, CH2 of Bn), 5.05
ꢁ
(d, 1H, J = 69.6 Hz, H3 ), 6.54 (s, 2H, NH2), 7.25–7.34 (m, 10H, Ph), 8.00

Fluoroanalogues of Cyclopropavir
239
(s, 1H, H1 ), 8.37 (s, 1H, H2), 8.91 (s, 1H, H8). 13C NMR 34.1 (d, J = 12.7
ꢁ
ꢁ
ꢁ
Hz, C4 ), 66.6, 69.4 (d, J = 3.7 Hz, C5 ), 71.29, 71.34 (2d, J = 237.3, 238.0
ꢁ
ꢁ
ꢁ
Hz, C3 ), 73.5, 73.6 (CH2 of Bn), 112.2 (d, J = 3.0 Hz, C2 ), 117.4 (C1 ),
119.5 (C5), 127.95, 127.97, 128.09, 128.18, 128.70, 128.75, 137.74, 137.96
(Ph), 138.6 (C8), 149.2 (C4), 153.7 (C2), 156.1 (C6). 19F NMR −210.95 (d,
J = 70.0 Hz). ESI-MS (MeOH + KOAc) 446 (M + H, 100.0), 484 (M + K,
100.0).
Z-isomer 24a: m.p. 160–162◦C. UV λmax 244 nm (ε 24,900), 281 (ε
1
ꢁ
8,300). H NMR (CDCl3) δ 3.62–3.81 (m, 4H, H5 ), 4.52, 4.55, 4.56 (3s, 4H,
ꢁ
CH2Ph), 5.26 (d, 1 H, J = 68.0 Hz, H3 ), 6.77 (s, 2H, NH2), 7.26–7.32 (m,
10 H, Ph), 7.63 (s, 1H, H1 ), 8.26 (s, 1H, H2), 8.39 (s, 1H, H8). 13C NMR
ꢁ
ꢁ
ꢁ
33.4 (d, J = 13.7 Hz, C4 ), 66.7, 69.2 (d, J = 3.7 Hz, C5 ), 72.1 (d, J = 238.9
ꢁ
ꢁ
ꢁ
Hz, C3 ), 73.3, 73.5 (CH2 of Bn), 113.9 (C2 ), 116.1 (C1 ), 119.6 (C5), 127.89,
127.93, 128.03, 128.6, 128.7, 138.0, 138.1, 138.2 (C8), 149.1 (C4), 153.7 (C2),
155.6 (C6). 19F NMR −211.88 (d, J = 68.5 Hz). ESI-MS 446 (M + H, 44.3),
468 (M+ Na, 100.0).
(E)-9-{[2,2-Bis(hydroxymethyl)-3-fluorocyclopropylidene]methyl}aden-
ine (13a). A solution of the BCl3·SMe2 complex (2.0 M in CH2Cl2, 3.35
mL, 6.73 mmol) was added dropwise to the E-isomer (23a, 300 mg, 0.673
mmol) in CH2Cl2 (30 mL) at room temperature under N2 over 10 minutes
with stirring which was then continued for 10 hours. NaHCO3 (2.8 g, 33.3
mmol) and methanol (30 mL) were added to quench the reaction and the
reaction mixture was stirred for 4 hours. The insoluble solid was filtered
off using a silica gel pad (3.5 g) which was washed with CH2Cl2-MeOH (2 :
1, 60 mL). The solvents from the filtrate were evaporated and the residue
was chromatographed using EtOAc-MeOH (10 : 1) to give product 13a
(147 mg, 82.7%), m.p. 243–245◦C. UV λmax 237 nm (ε 23,100), 280 (ε
1
ꢁ
8,100). H NMR δ 3.51–3.55, 3.69–3.76 (2m, 4H, H5 ), 5.18, 5.22 (2t, 2H, J
ꢁ
= 5.4, 6.6 Hz, OH), 5.21 (d, 1H, J = 70.4 Hz, H3 ), 7.41 (s, 2H, NH2), 7.89
(s, 1H, H1 ), 8.19 (s, 1H, H2), 8.82 (s, 1H, H8). 13C NMR 37.9 (d, J = 12.0
ꢁ
ꢁ
ꢁ
ꢁ
Hz, C4 ), 58.2, 60.8 (d, J = 4.5 Hz, C5 ), 72.3 (d, J = 232.1 Hz, C3 ), 114.2
ꢁ
ꢁ
(d, J = 3.0 Hz, C2 ), 116.8 (d, J = 3.0 Hz, C1 ), 119.2 (C5), 138.5 (C8), 148.9
(C4), 153.9 (C2), 156.8 (C6). 19F NMR −212.07 (d, J = 71.0 Hz). EI-MS 266
(M + H, 100.0), 288 (M + Na, 73.4), 553 (2M + Na, 20.3). Anal. Calcd for
C11H12FN5O2×0.1 H2O: C, 50.67; H, 4.34; N, 29.55. Found: C, 50.54; H,
4.30; N, 29.31.
(Z)-9-{[2,2-Bis(hydroxymethyl)-3-fluorocyclopropylidene]methyl}aden-
ine (14a). The procedure described above for the E-isomer 13a was
repeated with the Z-isomer 24a to give analogue 14a (150 mg, 84.4%), m.p.
1
230–232◦C. UV λmax 237 nm (ε 23,400), 280 (ε 8,200). H NMR δ 3.53 (d,
ꢁ
2H, J = 5.6 Hz), 3.64 (m, 2H, H5 ), 4.89 (t, 2 H, J = 5.6 Hz, OH), 5.42 (d,
ꢁ
ꢁ
1 H, J = 68.8 Hz, H3 ), 7.44 (s, 2 H, NH2), 7.53 (s, 1 H, H1 ), 8.20 (s, 1 H,
13
ꢁ
H2), 8.34 (s, 1 H, H8). C NMR 36.8 (d, J = 11.9 Hz, C4 ), 58.2, 60.9 (d,
ꢁ
ꢁ
J = 5.2 Hz), 73.2 (d, J = 233.6 Hz, C3 ), 115.0 (d, J = 3.7 Hz, C2 ), 119.1

240
S. Zhou et al.
(C5), 119.2 (C1 ), 137.9 (C8), 148.9 (C4), 154.2 (C2), 156.8 (C6). 19F NMR
−212.87 (d, J = 68.5 Hz). ESI-MS (MeOH + KOAc) 266 (M + H, 14.3), 304
(M + K, 100.0). Anal. Calcd for C11H12FN5O2 × 0.1 H2O: C, 50.67; H, 4.34;
N, 29.55. Found: C, 50.89; H, 4.43; N, 29.33.
ꢁ
(E)- and (Z)-2-Amino-6-chloro-9-{[2,2-(bis(benzyloxymethyl)-3-fluorocy-
clopropylidene-methyl}purine (23c) and (24c). A mixture of 2-amino-6-
chloropurine (430 mg, 2.55 mmol), K2CO3 (1.05 g, 7.62 mmol) and
compound 22 (1.2 g, 2.54 mmol) in DMF (7.5 mL) was stirred under N2
for 7 hours at room temperature, then at 60◦C for 1 hour and, finally, at
100–105◦C for 2.5 hours. After cooling, the insoluble portion was filtered
off using a silica gel pad (5 g) which was washed with DMF (70 mL).
DMF was evaporated in vacuo and the residue was chromatographed in
hexanes-EtOAc (10 : 1 to 3 : 1) to give the faster moving E-isomer 23c
(210 mg, 17.5%) and slower moving Z-isomer 24c (440 mg, 36.7%) as white
solids.
E-Isomer 23c: m.p. 135–137◦C. UV λmax 242 nm (ε 32,800), 310 (ε
7,800). 1H NMR (CDCl3) δ 3.47, 3.85 (split AB, 2H, J = 12.0, 2.0 Hz), 3.72,
ꢁ
3.79 (AB, 2 H, J = 10.2 Hz, H5 ), 4.51, 4.53 (2s, 4H, CH2 of Bn), 5.01 (d, 1
ꢁ
H, J = 68.8 Hz, H3 ), 5.22 (bs, 2H, NH2), 7.22–7.33 (m, 10H, Ph), 7.78 (s,
13
ꢁ
ꢁ
1H, H1 ), 8.89 (s, 1 H, H8). C NMR δ 34.2 (d, J = 12.7 Hz, C4 ), 66.4, 69.2
ꢁ
ꢁ
(J = 5.2 Hz, C5 ), 71.2 (d, J = 238.9 Hz, C3 ), 73.5, 73.6 (CH2 of Bn), 112.8
ꢁ
ꢁ
(d, J = 3.7 Hz, C2 ), 116.9 (d, J = 3.0 Hz, C1 ), 125.4 (C5), 127.97, 128.05,
128.2, 128.67, 128.72, 137.6, 137.8 (Ph), 140.5 (C8), 151.8, 152.8 (C4, C2),
159.6 (C6). 19F NMR −211.22 (d, J = 70.4 Hz). ESI-MS (MeOH) 480, 482
(M + H, 100.0, 41.1). Anal. Calcd for C25H23ClFN5O2: C, 62.56; H, 4.83; N,
14.59. Found: C, 62.38; H, 4.77; N, 14.56.
Z-Isomer 24c: m.p. 145–146◦C. UV λmax 242 nm (ε 34,100), 310
1
ꢁ
(ε 7,900). H NMR (CDCl3) δ 3.59–3.81, m, 4 H, H5 ), 4.52 (s), 4.53, 4.58
ꢁ
(AB, 4 H, J = 12.0 Hz, CH2 of Bn), 5.21 (d, 1H, J = 68.8 Hz, H3 ), 5.31
ꢁ
(s, 2H, NH2), 7.26–7.34 (m, 10H, Ph), 7.46 (s, 1 H, H1 ), 8.11 (s, 1 H, H8).
13
ꢁ
ꢁ
C NMR 33.4 (d, J = 12.7 Hz, C4 ), 66.6, 69.2 (J = 4.5 Hz, C5 ), 72.1 (d,
ꢁ
ꢁ
J = 237.3 Hz, C3 ), 73.3, 73.5 (CH2 of Bn), 114.0, 115.6 (d, J = 3.0 Hz, C1 ,
ꢁ
C2 ), 125.4 (C5), 127.9, 127.97, 128.06, 128.6, 128.7, 138.0, 138.2 (Ph), 139.5
(C8), 152.0, 152.5 (C4, C2), 159.8 (C6). 19F NMR −211.29 (d, J = 67.4 Hz).
ESI-MS 480, 482 (M, 100.0, 38.1), 502, 504 (M + Na, 97.6, 36.9). Anal. Calcd
for C25H23ClFN5O2: C, 62.56; H, 4.83; N, 14.59. Found: C, 62.51; H, 4.83; N,
14.58.
(E)-2-Amino-6-chloro-9-{[2,2-bis(hydroxymethyl)-3-fluorocyclopropylid-
ene]methyl}purine (13c). A solution of BCl3·SMe2 complex (2.0 M
in CH2Cl2, 2.1 mL, 4.2 mmol) was added dropwise to a solution of the
E-isomer 23c (200 mg, 0.418 mmol) in CH2Cl2 (8 mL) at room temperature
under N2 over 10 minutes with stirring. The stirring was continued for
6 hours. The reaction was quenched with methanol (20 mL) and NaHCO3
(4.0 g, 47.6 mmol). After 10 hours of stirring, the insoluble portion was

Fluoroanalogues of Cyclopropavir
241
filtered off through a silica gel pad (3.5 g) which was washed with CH2Cl2 –
MeOH (2 : 1, 60 mL). After removal of solvents from the filtrate, the
residue was chromatographed using hexane-EtOAc (1 : 1 to 100% EtOAc)
to afford compound 13c (107 mg, 86%), m.p. 238–239◦C. UV λmax 241
1
nm (ε 25,700), 309 (ε 7,400). H NMR δ 3.49–3.53, 3.70–3.77 (2m, 4H,
ꢁ
H5 ), 5.11 (t, 1H, J = 5.6 Hz), 5.16 (t, 1H, J = 4.8 Hz, OH), 5.20 (d partly
ꢁ
overlapped with δ 5.11, J = 70.4 Hz, H3 ), 7.08 (s, 2H, NH2), 7.69 (d, 1H, J
13
ꢁ
ꢁ
= 1.6 Hz, H1 ), 8.79 (s, 1H, H8). C NMR 37.9 (d, J = 12.0 Hz, C4 ), 58.2,
ꢁ
ꢁ
60.8 (d, J = 3.7 Hz, C5 ), 72.2 (d, J = 232.8 Hz, C3 ), 114.8 (d, J = 3.0 Hz),
ꢁ
ꢁ
116.2 (d, J = 2.9 Hz, C1 , C2 ), 123.8 (C5), 140.3 (C8), 150.4 (C4), 153.3 (C2),
160.9 (C6). 19F NMR −212.79 (d, J = 70.0 Hz). ESI-MS 300, 302 (M + H,
90.1, 37.0), 322, 324 (M + Na, 100.0, 33.1).
(Z)-2-Amino-6-chloro-9-{[3-fluoro-2,2-bis(hydroxymethyl)cyclopropylid-
ene]methyl}purine (14c). The procedure described above for the E-isomer
13c was followed using the Z-isomer 24c (400 mg, 0.83 mmol) to give
compound 14c (210 mg, 84%), m.p. 247–249◦C. UV λmax 242 nm
1
ꢁ
(ε 27,400), 310 (ε 7,200). H NMR δ 3.49–3.52, 3.58–3.69 (2 m, 4 H, H5 ),
ꢁ
4.90 (t, 2 H, J = 5.6 Hz, OH), 5.43 (d, J = 68.0 Hz, H3 ), 7.09 (s, 2 H, NH2),
13
ꢁ
ꢁ
7.37 (s, 1 H, H1 ), 8.30 (s, 1 H, H8). C NMR 37.0 (d, J = 12.7 Hz, C4 ),
ꢁ
ꢁ
58.1, 60.9 (d, J = 5.2 Hz, C5 ), 73.3 (d, J = 233.5 Hz, C3 ), 114.6 (d, J = 3.0
ꢁ
ꢁ
Hz), 115.8 (C1 , C2 ), 123.8 (C5), 140.0 (C8), 150.6 (C4), 153.1 (C2), 161.0
(C6). 19F NMR −212.37 (d, J = 68.5 Hz). ESI-MS 300, 302 (M + H, 100.0,
29.8), 322, 324 (M + Na, 88.4, 23.5).
(E)-9-{[2,2-Bis(hydroxymethyl)-3-fluorocyclopropylidene]methyl}guan-
ine (13b). A solution of the E-isomer 13e (100 mg, 0.33 mmol) in 80%
formic acid (5 mL) was heated at 80◦C with stirring for 4 hours. After
cooling, formic acid and water were evaporated in vacuo and the crude
product was dissolved in methanol (10 mL). Ammonia in methanol (20%,
10 mL) was added at 0◦C and the reaction mixture was stirred for 4 hours.
The volatile components were evaporated, the crude product was dried at
0.01–0.03 torr at room temperature whereupon it was recrystallized from
methanol to give compound 13b (85 mg, 90.5%), m.p. >300◦C. UV 243 (ε
22,300), 274 nm (ε 7,700). H NMR δ 3.44–3.49, 3.64–3.74 (2 m, 4 H, H5 ),
1
ꢁ
5.16 (d partly overlapped with δ 5.23, 1 H, J = 70.8 Hz, H3), 5.23 (t, 2 H,
ꢁ
OH), 6.89 (s, 2 H, NH2), 7.55 (s, 1 H, H1 ), 8.42 (s, 1 H, H8), 11.01 (bs, 1
H, NH). 13 C NMR 37.7 (d, J = 11.3 Hz, C4 ), 58.0, 60.7 (d, J = 4.5 Hz, C5 ),
ꢁ
ꢁ
ꢁ
ꢁ
72.3 (d, J = 232.0 Hz, C3 ), 113.8 (d, J = 3.0 Hz), 116.4 (d, J = 2.2 Hz, C1 ,
C2 ), 117.0 (C5), 134.7 (C8), 150.7 (C4), 155.1 (C2), 157.2 (C6). 19F NMR
ꢁ
−212.63 (d, J = 71.9 Hz). ESI-MS 282 (M + H, 17.0), 304 (M + Na, 100.0).
Anal. Calcd for C11H12N5FO3: C, 46.98; H, 4.30; N, 24.90. Found: C, 47.07;
H, 4.50; N, 25.03.
(Z)-9-{[2,2-Bis(hydroxymethyl)-3-fluorocyclopropylidene]methyl}guan-
ine (14b). The procedure described above for the E-isomer 13b was
followed using the Z-isomer 14e (150 mg, 0.5 mmol) to give compound 14b

242
S. Zhou et al.
1
(120 mg, 85%), m.p. >300◦C. UV 243 (ε 22,800), 274 nm (ε 7,900).
H
ꢁ
NMR δ 3.50 (poorly resolved d, 2 H), 3.56–3.67 (m, 2 H, H5 ), 4.87 (t, 2 H,
ꢁ
J = 5.6 Hz, OH), 5.38 (d, 1 H, J = 69.2 Hz, H3 ), 6.56 (s, 2 H, NH2), 7.26 (s,
1 H, H1 ), 7.90 (s, 1 H, H8), 10.74 (bs, 1 H, NH). 13C NMR 36.7 (d, J = 12.0
ꢁ
ꢁ
ꢁ
ꢁ
Hz, C4 ), 58.1, 60.9 (d, J = 5.2 Hz, C5 ), 73.1 (d, J = 232.8 Hz, C3 ), 114.7
ꢁ
ꢁ
(d, J = 1.4 Hz), 114.9 d, J = 2.9 Hz, C1 , C2 ), 117.1 (C5), 134.3 (C8), 150.6
(C4), 154.9 (C2), 157.3 (C6). 19F NMR −212.86 (d, J = 68.9 Hz). ESI-MS
282 (M + H, 32.1), 304 (M + Na, 100.0), 585 (2M + Na, 32.0). Anal. Calcd
for C11H12N5FO3: C, 46.98; H, 4.30; N, 24.90. Found: C, 46.76; H, 4.50; N,
24.90.
Antiviral Assays
The antiviral assays were performed as described previously.[10] The
HCMV assays were run in HFF culture with two strains of virus, Towne and
AD169, in a plaque reduction or cytopathic effect (CPE) inhibition assay.
The EBV assays were performed in Akata instead of Daudi cells using DNA
hybridization. The VZV was assayed in HFF cells by CPE inhibition or plaque
reduction. For cytotoxicity assays, HFF and Akata cells were employed.
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