Synthesis and antiviral evaluation of cis-substituted cyclohexenyl and cyclohexanyl nucleosides

Article abstract of DOI:10.1021/jm0493966

Starting from commercially available (rac)-3-cyclohexene-1-carboxylic acid, a series of purine and pyrimidine cis-substituted cyclohexenyl and cyclohexanyl nucleosides were synthesized through a key Mitsunobu reaction. Antiviral evaluations were performed

Full text of DOI:10.1021/jm0493966

4
50
J. Med. Chem. 2005, 48, 450-456
Synthesis and Antiviral Evaluation of Cis-Substituted Cyclohexenyl and
Cyclohexanyl Nucleosides
†
‡
‡
§
§
§
Karine Barral, J e´ r oˆ me Courcambeck, G e´ rard P e` pe, Jan Balzarini, Johan Neyts, Erik De Clercq, and
,
†
Michel Camplo*
Laboratoire des Mat e´ riaux Mol e´ culaires et des Biomat e´ riaux, GCOM2, UMR CNRS 6114, Universit e´ de la M e´ diterran e´ e, case
9
01, 163 av. de Luminy, 13288 Marseille Cedex 9, France, Rega Institute for Medical Research, Katholieke Universiteit Leuven,
B-3000 Leuven, Belgium, and Genoscience, Marseille, France
Received July 28, 2004
Starting from commercially available (rac)-3-cyclohexene-1-carboxylic acid, a series of purine
and pyrimidine cis-substituted cyclohexenyl and cyclohexanyl nucleosides were synthesized
through a key Mitsunobu reaction. Antiviral evaluations were performed on HIV, coxsackie
B3, and herpes viruses (HSV-1, HSV-2, VZV, HCMV). Three compounds showed moderate
activity against HSV-1 and coxsackie viruses. Specific computer modeling studies were
performed on HSV-1 thymidine kinase in order to understand the enzyme activation of an
analogue showing moderate antiviral activity.
Introduction
Nucleoside analogues play a major role in antiviral
chemotherapy. Although interest in the design of these
analogues have relatively decreased during the past few
years, the emergence of resistance to currently FDA-
1
approved drugs has generated new interest for the
Figure 1. D- and L-Cyclohexenylguanine nucleosides.
search of new active nucleoside analogues. Despite the
fact that the carbocyclic analogues has led to the
2
discovery of carbovir as anti-HIV agent, less efforts
have been directed toward the synthesis of six-mem-
3
4
bered carbocyclic analogues. However, Herdewijn et al.
have described D- and L-5-hydroxy-4-hydroxymethyl-2-
cyclohexenylguanine (Figure 1) which selectively showed
potent and selective anti-herpes virus activity (HSV-1,
HSV-2, VZV, CMV).
Figure 2. General structure of cyclohexenyl and cyclo-
hexanyl nucleosides.
The rationale of these findings stems mostly from the
using 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), afforded
9
fact that the cyclohexene ring can be considered as a
the unsaturated lactone 3 in quantitative yield. Reduc-
5
(
bio)isostere of a saturated furanose ring. Cyclohexene
tion of 3 with lithium aluminum hydride, followed by
the protection of the primary alcohol function of 4,
provided 5 in 58% yield from compound 3. The prepara-
tion of the trans derivative 6 was accomplished using a
Mitsunobu-type reaction on the allylic alcohol 5. Thus,
the introduction of a benzoyl protective group allowed
an inversion of configuration.10 The allylic alcohol 5 was
reacted with benzoic acid in the presence of diethyl
azodicarboxylate (DEAD) and triphenylphosphine (PPh3)
in dry THF to give 6. Alkaline hydrolysis of 6 afforded
in 93% yield the trans allylic alcohol 7. It should be
noted that partial hydrolysis was achieved with am-
monia solution. The use of sodium hydroxide was
necessary to complete hydrolysis.
nucleosides analogues can also confer protection from
resistance to hydrolysis, since glycosidic bond cleavage
is a frequently encountered degradative pathway of
nucleoside antivirals, particularly for the 2′,3′-dideoxy-
6
nucleosides. Moreover, the conformational flexibility of
these cyclohexene derivatives proved to be important
7
for antiviral activity.
The aim of this article is to describe the synthesis and
antiviral evaluation of six-membered nucleoside con-
taining cyclohexenyl and cyclohexanyl sugars (Figure
2
).
Chemistry
The corresponding cis-cyclohexenyl nucleosides were
obtained by use of a second Mitsunobu-type reaction
between the allylic alcohol 7 and pyrimidine and purine
bases.11 The synthesis of the cytosin-1-yl (10), thymin-
The known allylic alcohol derivative 7 (Scheme 1) is
a requisite for the Mitsunobu coupling reaction of
8
various nucleoside bases. Iodolactonization of the com-
mercially available precursor 1 (rac)-3-cyclohexene-1-
carboxylic acid, followed by elimination of the iodide 2
1
-yl (13), adenin-9-yl (17), and guanin-9-yl (19) deriva-
tives are illustrated in Schemes 2 and 3.
*
To whom correspondence should be addressed. Tel: 04 91 82 95
Condensation of the common intermediate, allylic
8
6, Fax: 04 91 82 95 80. E-mail: camplo@luminy.univ-mrs.fr.
4
3
alcohol 7, respectively with N -benzoylcytosine and N -
benzoylthymine12 in the presence of DEAD and tri-
phenylphosphine in THF gave the cis racemic cytosine
†
Laboratoire des Mat e´ riaux Mol e´ culaires et des Biomat e´ riaux
Rega Institute.
Genoscience.
‡
§
1
0.1021/jm0493966 CCC: $30.25 © 2005 American Chemical Society
Published on Web 12/23/2004

Antiviral Cyclohexenyl and Cyclohexanyl Nucleosides
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 2 451
Scheme 1a
Scheme 3a
a
Reagents and conditions: (a) aq NaHCO3. then aq KI/I2; (b)
DBU, toluene reflux 10 h; (c) AlLiH4, THF, rt, 2 h ; (d) TBDMSCl,
imidazole, DMF, rt, 5 h; (e) DEAD, PPh3, BzOH, THF rt, 5 h ; (f)
NH3/MeOH, NaOH 2 N, rt, 3 h.
Scheme 2a
a
Reagents and conditions: (a) 6-chloropurine, DEAD, PPh3,
THF, rt, 18 h; (b) 2-amino-6-chloropurine, DEAD, PPh3, dioxane,
rt, 48 h; (c) NH3/MeOH, 80 °C, 24 h; (d) TFA/H2O (3/1), rt, 72 h;
(
e) TBAF, THF, 4 h; (f) 10% Pd/C, H2, EtOH/EtOAc, rt, 24 h.
1
2 in EtOAc over 10% palladium on carbon gave the
saturated cytosine derivative 10 and the saturated
thymine derivative 13 in 56% and 77% yield, respec-
tively. It should be noted that hydrogenation in meth-
anol or ethanol rapidly led to a rapid C1′-N cleavage.
Using the same conditions, the Mitsunobu-type reac-
tion on the allylic alcohol 7 with 6-chloropurine and
2
-amino-6-chloropurine gave the protected 6-chloro-
purine derivatives 14 and 18 in 39% and 40% yield,
respectively (Scheme 3).14
Construction of the adenine ring from 14 was ac-
complished by treatment with methanolic ammonia in
a sealed reaction vessel for 1 day to give 15 in 52% yield.
The material was converted to pure adenine-cyclohexene
nucleoside 16 by treatment with TBAF in THF. The
a
4
Reagents and conditions: (a) N -benzoylcytosine, DEAD, PPh3,
3
THF, rt, 18 h; (b) N -benzoylthymine, DEAD, PPh3, THF, rt, 18
h; (c) TBAF, THF, 3 h then sat. NH3/MeOH, 20 h; (d) 10% Pd/C,
H2, EtOAc, 24 h.
2
-amino-6-chloropurine derivative 18 was converted to
the guanine cyclohexene nucleoside 19 by treatment
with TFA-H2O (3:1). Under these conditions, the TB-
DMS protecting group was simultaneously removed
with an overall yield of 36%. Hydrogenation of com-
pound 16 over palladium on activated carbon gave the
saturated adenine derivative 17 in 67% yield. However,
hydrogenation of compound 19 either in methanol or
and thymine derivatives 8 and 11 in 55% and 45% yield
(
Scheme 2). Compounds 8 and 11 were converted to
compounds 9 and 12 by treatment with TBAF in THF
followed by a saturated ammonia solution in MeOH. The
overall yields, starting from 8 and 11, were 34% and
1
3
3
6% respectively. Hydrogenation of compounds 9 and

4
52 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 2
Barral et al.
Table 1. Antiviral Activity of Cyclohexenyl and Cyclohexanyl
Nucleosides in Comparison with Standard and Approved
Antiviral Drugs: Effective Concentration or Concentration
Required To Inhibit 50% of Virus-Induced Cytopathicity (EC50,
a
µM)
compd
Coxsackie B3 (Vero cells) HSV-1 Kos (Hel cells)
(
(
(
(
()-cis 9
>200
>200
>200
168
>200
>200
>200
>200
62
()-cis 10
()-cis 12
()-cis 13
63
>200
>200
>200
>200
Figure 3. NOE effects on cyclohexenyl and cyclohexa-
nyl derivatives.
(()-cis 16
(()-cis 17
(
()-cis 19
DPN
5.84
n.d.
n.d.
n.d.
0.32
0.06
ethanol led to decomposition whereas the use of neat
EtOAc was unsuitable since 19 was insoluble in this
solvent. Attempts using a mixture of EtOH/EtOAc were
ineffective too.
acyclovir
gancyclovir
a
n.d. ) Not determined. DPN: 2-(3,4-dichlorophenoxy)-5-ni-
trobenzonitrile.
Conformational Study
with the bases and the hydroxymethyl group orientated
in equatorial position (cis configuration). Similar results
were obtained for compounds 9, 10, 13, 16, and 19.
The configuration of the synthesized cis-substituted
cyclohexenyl and cyclohexanyl nucleosides 9, 10, 12, 13,
1
1
6, 17, and 19 were confirmed by 1D and 2D H NMR
spectroscopy. Data are given for each compound in the
Results and Discussion
1
Experimental Section. A detailed H NMR conforma-
Antiviral Activity. Evaluation of antiviral activity
was done as previously described.15 The target com-
pounds were evaluated against HIV-1 and HIV-2, her-
pes simplex virus type 1 (HSV-1) and type 2 (HSV-2),
vaccinia virus, coxsackie B3 virus in Vero cells, and
human cytomegalovirus in Hel cells. No significant
cytotoxicities were reported for any of the compounds.
None of these compounds were active against any of the
tested viruses except for coxsackie and herpes simplex
virus type 1. Compound 13 displayed a moderate
activity against coxsackie whereas 10 and 12 displayed
moderate activity against wild-type HSV-1. The weak
but consistent activity of these compounds against
HSV-1 is encouraging and may suggest that these new
nucleosides are metabolized to their 5′-triphosphates or
as such recognized by the viral DNA polymerases. As
intracellular phosphorylation in the virus-infected cells
is postulated to be an important step in the enzymatic
activation of these nucleosides, we carried out molecular
modeling studies using compound 12 and HSV-1 thy-
midine kinase.
Molecular Modeling. Molecular modeling was per-
formed on thymidine cyclohexenyl derivative 12 in the
HSV-1 thymidine kinase (TK) active site, using “Gen-
Mol” (www.3dgenoscience.com) software with its all
atoms force field, from the X-ray crystal structure of
wild-type binary complex TKHSV-1/5-iodouridine anhy-
drohexitol (AHIU) derivative (PDB ID: 1KI6).16 HSV-1
thymidine kinase (TKHSV-1) activates antiviral nucleo-
sides such as acyclovir to the 5′-monophosphate. Further
phosphorylation by cellular kinases yields the 5′-tri-
phosphate that is a selective inhibitor of the viral
polymerase. Phosphorylation of compound 12 by TKHSV-1
may be a prerequisite for antiviral activity. We therefore
carried out a molecular modeling study.
tional study was carried out on compounds 12 and 17
in DMSO-d6 at room temperature.
For compound 12 the H-2′ resonance at δ ) 5.49 ppm
appears as a doublet with one large coupling constant
J2′-3′ ) 9.94 Hz.
The absence of coupling between H-1′ and H-2′
indicates, according to the Karplus equation, that the
dihedral angle between C-1′-H-1′ and C-2′-H-2′ is close
to 90° (involving a quasi-null constant of coupling). That
is possible only if proton H-1′ assumes a pseudoaxial
position with the thymine base in a pseudoequatorial
orientation. Moreover, the proton H-6′ax at δ ) 1.27 ppm
represents a ddd which appears as a broad quadruplet
because the coupling constant J6′ax-6′eq is equivalent to
the constant J6′ax-1′ = J6′ax-5′ ) 11.6 Hz. These data
suggest a pseudoaxial position of H-1′ and H-5′. There-
fore, it can be concluded that compound 12 exists
predominantly in a pseudo-chair conformation with the
thymine base and the hydroxymethyl group in pseudo-
equatorial positions. These conclusions are further
1
1
supported by examination of 2D H- H NOESY spectra.
Strong NOE interactions were observed between H-1′,
H-5′, and H-6′eq suggesting the cis configuration of the
nucleosides 12 (Figure 3).
For compound 17, the H-1′ resonance at δ ) 3.69 ppm
appears as a triplet of triplets with coupling constants:
J1′-2′ax = J1′-6′ax ) 12.15 Hz and J1′-2′eq = J1′-6′eq ) 3.6
Hz. Altogether, these coupling constants confirm that
H-1′ occupies an axial position in a chair conformation.
As previously mentioned, H-2′ax at δ ) 0.81 represents
a ddd which appears as broad quadruplet resulting from
three approximately equal coupling constants J2′ax-2′eq
=
J2′ax-1′ = J2′ax-3′ ) 12.1 Hz. These data suggest a chair
conformation with the heterocyclic moiety and the
hydroxymethyl group in equatorial orientations. Strong
NOE interactions were observed between H-1′, H-2′eq.
and H-3′, and no correlation appears between H-1′ and
H-2′ax (Figure 3). These findings definitively exclude any
boatlike conformation.
The crystal structure of HSV-1 thymidine kinase
complexed with a iodouridine anhydrohexitol derivative
and dT was used as a model for this molecular modeling
study (Figure 4). Indeed, since compound 12 is a six
membered ring, it seemed reasonable to use a TKHSV-1
crystal structure with a six membered ring. The TKHSV-1
active site was then adapted to this type of ribose
In summary, the values of vicinal H,H coupling
constants and NOE measurements lead to the conclu-
sion that both 12 and 17 are in a chair conformation

Antiviral Cyclohexenyl and Cyclohexanyl Nucleosides
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 2 453
Figure 4. Structures of AHIU and compound 12 used
for the molecular modeling study.
Figure 6. Molecular modeling of compound 12 and dT in the
TKHSV-1 active site.
lacks the hydrogen bond network with the 3′OH TKHSV-1
binding pocket.
The molecular model of compound 12 is superimposed
with the minimized 2′-deoxythymidine-TKHSV-1 crystal
structure (PDB ID: 1KIM).16 Natural nucleoside sub-
strate dT of TKHSV-1 is displayed in brown and the
model of 12 in the TKHSV-1 active site in atom type.
Water molecules are shown with its hydrogen (oxygen
in red and hydrogen in cyan). For clarity, only the
hydrogen bonds between 12 and TKHSV-1 active site are
shown (green dotted line).
Figure 5. Molecular modeling of compound 12 and AHIU in
the TKHSV-1 active site.
This picture were generated with Swiss-PdbViewer
and rendered with PovRay 3.5.
mimicking ring. Compound 12 binds in the deep pocket
of the TK active site and is stabilized by hydrophobic
van der Waals interactions and by direct or indirect
hydrogen bonds through structural water molecules.
The active conformation of compound 12 in the
TKHSV-1 shows a thymidine base with a pseudoequato-
rial orientation which is consistent with the iodouridine
conformation of AHIU in TKHSV-1 structure (Figure 5).
Moreover, the hydroxymethyl moiety of compound 12
is in an axial conformation allowing close interactions
in the 5′OH binding site of TKHSV-1: Glu-83, Arg-222,
Arg-163 (Figure 5).
In the case of AHIU (Figure 5), its 3′OH makes
hydrogen bonds with Tyr-101 and Glu-225. These
specific interactions increase the binding affinity as well
as the stability of the complex with TKHSV-1. However,
the cyclohexenyl of TKHSV-1 ring has fine geometrical
complementarities with the ribose ring binding pocket.
The cyclohexenyl ring provides favorable hydrophobic
van der Waals interactions with the hydrophobic pocket
of TKHSV-1 shaped by the side chain of hydrophobic
residue Trp-88, Met-85, Ile-97 and alkyl side chain of
17
Arg-222, as previously described (Figures 5 and 6). The
The molecular model of compound 12 is superimposed
with the minimized AHIU-TKHSV-1 crystal structure
thymine moiety of compound 12 undergoes π-stacking
interactions with Tyr-172 and van der Waals contacts
with Met-128 as well as Ile-100. Structurally, thymine,
which is sandwiched between Tyr-172 and Met-128
(Figure 5) allows a fine orientation of the nucleobase
for the Watson-Crick-like hydrogen bonds to Gln-125.
Thymine is strongly hydrogen bonded to N3 and O4
with Ãꢀ1 and Nꢀ2 of Gln-125 respectively (Figure 4).
O2 thymine nucleobase as well provides three additional
water-mediated hydrogen bonds with Arg-176, Oú Tyr-
101, and Oꢀ1 of Gln-125. This strong hydrogen bond
network on thymine affords an excellent anchorage
point for compound 12 as well as a fine orientation in
regard with Gln-125 for the Watson-Crick-like interac-
tions. The existence of these specific interactions allows
compound 12 to have favorable interaction with the
nucleoside binding site of TKHSV-1 and a correct orien-
tation of its 5′OH for the phosphorylation with ATP
cofactor. The relatively weak activity of compound 12
could be explained by the loss of 3′OH interactions with
residue Glu-225 and Tyr-101 of TKHSV-1 3′OH binding
pocket (Figures 5 and 6). Moreover, the 1,3 substitution
(
PDB ID: 1KI6).16 Nucleoside analogue AHIU is dis-
played in brown and the model of 12 in the TKHSV-1
active site in atom type. Water molecules are shown
with its hydrogen (oxygen in red and hydrogen in cyan).
For clarity, only the hydrogen bonds between 12 and
the TKHSV-1 active site are shown (green dotted line).
This picture was generated with Swiss-PdbViewer and
rendered with PovRay 3.5.
Thus, compound 12 adopts a half chair conformation
and allows this compound to mimic the active South
3
′exo conformation for the natural nucleoside substrates
1
6
of TKHSV-1 such as dT (PDB ID: 1KIM, Figure 6). The
′OH hydroxymethyl of compound 12 is hydrogen
5
bonded to Glu-83 and via a water molecule to Lys 62,
Arg 163, Arg 222, and the sulfate anion that is located
in a phosphate binding loop, residues 59 g 63 (GMGKT)
corresponding to the phosphate binding site of the ATP
cofactor. A second water molecule allows to extend the
hydrogen bond network to Asp-162 (Figure 5). Com-
pound 12 is a dideoxynucleoside analogue (ddN) and

4
54 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 2
Barral et al.
of cyclohexenyl ring requires a half chair conformation
to mimic the south 3′OH exo active conformation of
natural nucleoside substrates for TKHSV-1. Therefore,
this energy consuming half chair conformation de-
creases the stability of the active complex 12/TKHSV-1
in regard with the phosphorylation.
In summary, the syntheses of cyclohexenyl and cyclo-
hexyl nucleoside derivatives were successfully achieved.
Conformational studies, as well as molecular modeling
performed on thymidine cyclohexenyl derivative 12 in
the HSV-1 thymidine kinase (TK) active site, allowed
us to understand the observed moderate antiviral activ-
ity. These results will be taken into account in order to
optimize the encouraging activity profile of compound
solvent in vacuo yielded 2 (9.60 g, 97%) as a yellow solid: mp
+
+
+
1
33 °C; MS (GT, FAB ): 125 (M - I) , 253 (M + 1H) .
(
()-6-Oxabicyclo[3.2.1]oct-3-en-7-one (3). Iodolactone 2
13.6 g, 54 mmol) was dissolved in dry toluene (120 mL)
containing 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) (12.48 g,
1 mmol), the mixture was heated at reflux for 6 h, cooled,
(
8
and filtered, and the filtrate was concentrated under reduced
pressure to give an oil. This crude oil was directly used in the
next step.
(
()-cis-5-(Hydroxymethyl)-2-cyclohexen-1-ol (4). The
crude lactone 3 (6.7 g, 54 mmol) was dissolved in dry THF
200 mL) and added dropwise to LiAlH (3.07 g, 81 mmol)
(
4
dissolved in dry THF (200 mL) at 0 °C. A white precipitate
formed. After 1 h, water (6.7 mL), 15% aqueous NaOH (6.7
mL), and water (20 mL) were added sequentially, and the
mixture was allowed to warm to room temperature with
stirring over several hours. The salts were removed by
12.
4
filtration, and the solution was dried over MgSO and concen-
trated at room temperature. Recrystallization from cyclohex-
Experimental Section
ane/acetone gave 4 (5.4 g, quantitative) as white solid: mp 82
°
C.
Molecular Modeling. The molecular modeling was per-
formed with GenMol (www.3dgenoscience.com) with its all
atoms force field, from the X-ray crystal structure of wild-type
binary complex TKHSV-1/AHIU derivative (PDB ID: 1KI6). The
first entity (A) of the TKHSV-1/AHIU crystal structure in the
asymmetric crystal unit was used for the molecular modeling
study. All crystallographic water molecules corresponding to
the first entity (A) were kept. All hydrogen atoms were added
to the enzyme and water from the Biopolymer module of
GenMol. Hydrogen positions were then optimized. Energy
refinements were done in a vacuum with a sigmoid distance-
(()-cis-5-[(tert-Butyldimethylsilyloxy)methyl]-2-cyclo-
hexen-1-ol (5). Diol 4 (2 g, 15.6 mmol) was dissolved in dry
DMF (30 mL). Imidazole (2.12 g, 31 mmol) was added and
dissolved. The solution was cooled to 0 °C, tert-butyldimethyl-
silyl chloride (2.35 g, 15.6 mmol, in 5 mL DMF) was added,
and the reaction mixture was allowed to warm to room
temperature and stirred for 10 h. Brine was added and the
solution extracted with ether and washed with water. MgSO
4
was added, and the solution was filtered and concentrated
under reduced pressure. The resulting oil was purified by flash
chromatography (dichloromethane) to give 5 (2.2 g, 58%).
(()-trans-1-Benzoyloxy-5-[(tert-butyldimethylsilyl-oxy)-
methyl]-2-cyclohexene (6). A solution of diethyl azodicar-
boxylate (DEAD) (2.86 mL, 18.14 mmol) in THF (10 mL) was
added dropwise to a stirred solution of the alcohol 5 (2.2 g,
9.07 mmol), triphenylphosphine (4.76 g, 18.14 mmol), and
benzoic acid (2.21 g, 18.14 mmol) in THF (40 mL) at room
temperature. After 14 h the mixture was partitioned between
chloroform (200 mL) and saturated aqueous sodium hydrogen
carbonate (200 mL). The aqueous layer was further extracted
with chloroform, and the combined organic extracts were dried,
filtered, and evaporated to give a white residue. This latter
was purified by flash chromatography (dichloromethane) to
1
8
dependent dielectric model. The construction of the nucleo-
side analogue 12 was based on X-ray conformation of AHIU
in the crystal structure (1KI6).16 All atom charges were
computed with GenMol for TK, nucleoside, water molecules,
and phosphate. Finally, compound 12 was docked into TKHSV-1
active site from the positioning of AHIU in the X-ray structure
binary complex 1KI6. The resulting binary complex was then
optimized using GenMol all atom force field with a gradient
of energy up to 0.01 kcal.mol-1
Å .
-1
Chemistry. General Methods. Melting points were de-
termined in capillary tubes with a 9100 Electrothermal (Fisher
1
Scientific) apparatus and are uncorrected. The H NMR and
13
C NMR spectra were determined with a BRUKER AMX 200
+
+
give 6 (2.6 g, 78%). MS (GT, FAB ): 225 (M - OBz) ; 347 (M
MHz and referenced to the solvent. Chemical shifts are
expressed in ppm and coupling constants (J) are in hertz (s )
singlet, d ) doublet, dd ) double doublet, ddd ) double double
doublet, t ) triplet, dt ) double triplet, tt ) triple triplet, m
+
+
1H) .
(()-trans-5-[(tert-Butyldimethylsilyl)oxy]methyl]-2-cy-
clohexen-1-ol (7). Compound 6 (2.2 g, 9.07 mmol) was
dissolved in THF (8 mL). A saturated ammonia solution in
MeOH (10 mL) and a solution of NaOH 2N (15 mL) were added
at room temperature. The mixture was stirred for 3 h until
no starting material appears on TLC. It was then evaporated,
and the residue was partitioned between chloroform and water.
The aqueous layer was further extracted with chloroform, and
)
multiplet, dm ) double multiplet, triple multiplet). FAB+
mass spectra (MS) were obtained on a JEOL DX-100 mass
spectrometer (Laboratoire de Mesures Physiques RMN, Dr.
Astier, USTL, Montpellier, France) using a cesium ion source
and a glycerol-thioglycerol (GT) matrix. Elemental microanal-
yses were determined by Service Central d’Analyse CNRS
Vernaison-Lyon France and gave combustion values for C, H,
N within 0.4% of the theoretical values. Preparative flash
column chromatographies were performed using silica gel
4
the combined organic extracts were dried over MgSO . The
residue was purified by flash chromatography (dichloromethane/
+
MeOH: 9.5/0.5) to afford 7 (1.35 g, 93%). MS (GT, FAB ): 109
+
+
+
(
Merck) G60 230-240 mesh. Analytical thin layer chromatog-
raphies were performed on silica gel 60F 254 aluminum plates
Merck) of 0.2 mm thickness. The spots were examined with
(M - TBDMSiO) ; 154 (M - tBu) ; 225 (M - OH) ; 243 (M +
+
1H) .
(
cis-(()-1-{5′-[(tert Butyldimethylsilyloxy)methyl]-2′-
4
UV light and Ceric Dip spray. Preparative TLC was done on
glass plates EM Science silica gel 60F 254 (1.0 mm or 2.0 mm
layer).
cyclohexenyl}N -benzoylcytosine (8). DEAD (2.16 mL,
13.77 mmol) dissolved in freshly distilled THF (15 mL) was
added dropwise over 5 min to a 0 °C stirred suspension of 7
4
(
()-4-Iodo-6-oxabicyclo[3.2.1]octan-7-one (2). 3-Cyclo-
hexene-1-carboxylic acid 1 (5 g, 39.7 mmol) was added to a
solution of NaHCO (9.95 g, 119 mmol) in water (96 mL), and
(1.67 g, 6.88 mmol), N -benzoylcytosine (2.96 g, 13.77 mmol),
and triphenylphosphine (3.61 g, 13.77 mmol) in anhydrous
THF (50 mL) under argon. The mixture was stirred for 3 h at
0 °C and then at room temperature for 20 h. Evaporation of
the volatiles and purification by flash chromatography (10-
15% EtOAc/cyclohexane) yielded 8 (1.89 g, 60%). MS (GT,
3
the resulting mixture was stirred until it became homoge-
neous. The flask was then protected from light, and the
mixture was treated in one portion with a solution of KI (39.50
g, 238 mmol) and iodine (10.57 g, 41.7 mmol) in water (96 mL).
The reaction mixture was stirred at room temperature for 16
+
+
+
+
FAB ): 216 (BBz + 1H) ; 382 (M - tBu) ; 440 (M + 1H) .
cis-(()-1-[5′-(Hydroxymethyl)-2′-cyclohexenyl]cy-
tosine (9). Compound 8 (595 mg, 1.05 mmol) dissolved in dry
THF (7 mL) was stirred with a 1 M solution of TBAF (1.58
mL, 1.58 mmoL) for 4 h at room temperature. The solution
h and then extracted with CHCl
combined, washed with 10% aqueous Na
NaHCO , and water, and then dried (MgSO
3
. The organic extracts were
, 10% aqueous
). Removal of the
2 2 3
S O
4
3

Antiviral Cyclohexenyl and Cyclohexanyl Nucleosides
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 2 455
trated to afford compound 17 (30 mg, 67%): mp 168 °C. MS
was then evaporated and the residue treated with a saturated
ammonia solution in MeOH (10 mL) for 24 h. Evaporation of
the volatiles and purification by flash chromatography (EtOAc)
+
+
+
(GT, FAB ): 136 (B + 1H) , 248 (M + 1H) . Anal. (C12
17 5
H N O)
C, H, N.
+
yielded 9 (80 mg, 34%): mp 167-168 °C. MS (GT, FAB ): 112
cis-(()-2-Amino-6-chloro-9-{5′-[(tert-butyldimethylsilyl-
oxy)methyl]-2′-cyclohexenyl}purine (18). DEAD (600 µL,
618 g, 3.55 mmol) dissolved in dry dioxane (10 mL) was added
dropwise over a period 2 h to a stirred suspension of 7 (430
mg, 1.775 mmol), 2-amino-6-chloropurine (602 mg, 355 mmol),
and triphenylphosphine (931 mg, 3.55 mmol) in anhydrous
dioxane (30 mL) under argon at room temperature. The
mixture was stirred for 48 h. Evaporation of the volatiles and
purification by flash chromatography (2% MeOH/dichloro-
methane) yielded 18 (280 mg, 40%) as a white foam: mp 135
+
+
(
15 3 2
B + 1H) ; 222 (M + 1H) . Anal. (C11H N O ) C, H, N.
cis-(()-1-[3′-(Hydroxymethyl)cyclohexanyl]cytosine
10). A suspension of compound 9 (18 mg, 0.08 mmol) and 10%
(
palladium on carbon (8 mg) in EtOAc (5 mL) was hydrogenated
under atmospheric pressure for 24 h. The reaction mixture
was filtered through Celite and concentrated to afford com-
pound 10 (10 mg, 56%): mp 166 °C. MS (GT, FAB+): 112 (B
+
+
+
1H) ; 224 (M + 1H) . Anal. (C11
17 3 2
H N O ) C, H, N.
cis-(()-1-{5′-[(tert-Butyldimethylsilyloxy)methyl]-2′-cy-
+
+
+
3
°C. MS (GT, FAB ): 170 (B + 1H) ; 394 (M + 1H) ; 782 (2M
clohexenyl}-N -benzoylthymine (11). DEAD (571 µL, 3.63
mmol) dissolved in anhydrous dioxane (5 mL) was added
dropwise over 15 min to a 0 °C stirred suspension of 7 (440
+
+
1H) .
cis-(()-9-[5′-(Hydroxymethyl)-2′-cyclohexenyl]gua-
3
nine (19). Compound 18 (250 mg, 0.634 mmol) was treated
mg, 1.81 mmol), N -benzoylthymine (836 mg, 3.63 mmol), and
2
with TFA/H O (3:1, 8 mL) for 72 h at room temperature. The
triphenylphosphine (952 mg, 3.63 mmol) in anhydrous dioxane
reaction mixture was then concentrated and coevaporated
twice with toluene. The residue was cooled and treated with
(
20 mL) under argon. The mixture was stirred for 3 h at 0 °C
and then at room temperature for 30 h. Evaporation of the
volatiles and purification by flash chromatography (10-20%
EtOAc/cyclohexane) yielded 11 (355 mg, 45%).
4
MeOH/NH OH (10:1, 15 mL). Volatiles were removed and the
residue was purified by flash chromatography (MeOH/dichloro-
methane 1:4) yielded 19 (60 mg, 36%): mp 174 °C. MS (GT,
cis-(()-1-[5′-(Hydroxymethyl)-2′-cyclohexenyl]thym-
ine (12). Compound 11 (350 mg, 0.77 mmol) dissolved in dry
THF (10 mL) was stirred with a 1 M solution of TBAF (1.15
mL, 1.15 mmoL) for 4 h at room temperature. The solution
was then evaporated and the residue treated with a saturated
ammonia solution in MeOH (10 mL) for 48 h. Evaporation of
the volatiles and purification by flash chromatography (dichloro-
methane/methanol: 98/2) yielded 12 (80 mg, 44%) as a white
+
+
+
FAB ): 262 (M + 1H) ; 562 (2M + 1H) . Anal. (C12
15 5 2
H N O )
C, H, N.
Acknowledgment. We wish to thank Dr. Robert
Faure (Universit e´ Aix-Marseille III, St. J e´ r oˆ me,
Marseille, France) for his expertise in conducting NMR
experiments and Mrs. Miette Stuyck, Mrs. Ann Absillis,
Mrs. Anita Vanlierde, and Mrs. Frieda De Meyer for
excellent technical assistance.
+
+
solid: mp 171-172 °C. MS (GT, FAB ): 127 (B + 1H) , 237
+
+
(
16 2 3
M + 1H) , 473 (2M) . Anal. (C12H N O ) C, N, H.
cis-(()-1-[3′-(Hydroxymethyl)cyclohexanyl]thymine
(
1
13). A suspension of compound 12 (30 mg, 0.13 mmol) and
Supporting Information Available: Elemental analysis
and spectroscopic data. This material is available free of charge
via the Internet at http://pubs.acs.org.
0% palladium on carbon (20 mg) in EtOAc (15 mL) was
hydrogenated at atmospheric pressure for 24 h. The reaction
mixture was filtered through Celite and concentrated to afford
+
References
compound 13 (24 mg, 77%): mp 171 °C. MS (GT, FAB ): 127
+
+
+
(
B + 1H) , 239 (M + 1H) , 477 (2M) . Anal. (C12
H
18
N
2
O
3
) C,
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(
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hexane) yielded 14 (420 mg, 39%): mp 99 °C. MS (GT, FAB ):
+
+
1
55 (B + 1H) , 379 (M + 1H) .
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19 (2M) .
cis-(()-9-[5′-(Hydroxymethyl)-2′-cyclohexenyl]ade-
nine (16). Compound 15 (200 mg, 0.55 mmol) dissolved in dry
THF (5 mL) was stirred with a 1 M solution of TBAF (834 µL,
0
.834 mmol) for 4 h at room temperature. The solution was
then evaporated, and the residue was dissolved in EtOAc,
washed with 10% aqueous citric acid, and water, and then
4
dried (MgSO ). Evaporation of the volatiles and purification
by flash chromatography (10% MeOH/dichloromethane) yielded
1
1
(
+
6 (40 mg, 30%): mp 173-174 °C. MS (GT, FAB ): 136 (B +
+
+
+
H) , 246 (M + 1H) , 491 (2M) . Anal. (C12
15 5
H N O) C, H, N.
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(
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