Radical cascade protocol for the synthesis of (5′S)- and (5′R)-5′,8-cyclo-2′-deoxyguanosine derivatives

Article abstract of DOI:10.1071/CH12494

The reaction of the appropriately substituted 8-bromo-2′- deoxyguanosine with Bu3SnH/2,2′-azobisisobutyronitrile (AIBN) can be favourably tuned to give the analogous 5′,8-cyclo-2′-deoxyguanosine derivatives in good yields, thus providing easy access to modified nucleosides that constitute an important DNA lesion. A large excess of AIBN is necessary. The creation of the new C5′-C8 bond is a non-chain radical cascade protocol.

Full text of DOI:10.1071/CH12494

RESEARCH FRONT
CSIRO PUBLISHING
Aust. J. Chem. 2013, 66, 330–335
Full Paper
http://dx.doi.org/10.1071/CH12494
Radical Cascade Protocol for the Synthesis of (59S)-
and (59R)-59,8-Cyclo-29-deoxyguanosine Derivatives*
A
,
A B
Michael A. Terzidis and Chryssostomos Chatgilialoglu
A
Istituto per la Sintesi Organica e la Fotoreattivit a` , Consiglio Nazionale delle Ricerche,
Via P. Gobetti 101, 40129 Bologna, Italy.
B
Corresponding author. Email: chrys@isof.cnr.it
0
0
The reaction of the appropriately substituted 8-bromo-2 -deoxyguanosine with Bu SnH/2,2 -azobisisobutyronitrile
3
0 0
(AIBN) can be favourably tuned to give the analogous 5 ,8-cyclo-2 -deoxyguanosine derivatives in good yields, thus
providing easy access to modified nucleosides that constitute an important DNA lesion. A large excess of AIBN is
0
necessary. The creation of the new C5 –C8 bond is a non-chain radical cascade protocol.
Manuscript received: 2 November 2012.
Manuscript accepted: 6 December 2012.
Published online: 4 February 2013.
Introduction
Chemical studies play a central role in the identification and
and the C8 purine carbon atoms is present. Recently, the rate
constant for the cyclization step1 - 2 at the nucleoside level was
[1,2]
5
ꢂ1 [6]
measurement of the main classes of DNA oxidative damage.
0
measured to be 6.1 ꢁ 10 s
. Lesion 3 exists in two diaste-
0
0
0
0
Purine 5 ,8-cyclonucleosides are an important class of ionizing
[
reomeric forms, i.e. (5 R)- and (5 S)-cdGuo (5 ,8-cyclo-2 -
deoxyguanosine).
3]
radiation-induced tandem DNA lesions. Their potential to
become free-radical stress markers is a matter of fundamental
0
0
Synthetic access to both (5 R)-cdGuo and (5 S)-cdGuo
lesions and their phosphoramidites for incorporation in oligo-
nucleotides models is still a challenge and limits the full
development of biochemical, biological, and biophysical
[
research. Scheme 1 shows the case of the guanine moiety. The
4]
0
ꢀ
attack at H5 of DNA by HO radicals is estimated to be 55 % of
0
[
5]
the H-atom abstraction occurs at the sugar moieties. After C5
[
3]
0
0
radical (1) formation, intramolecular addition to the C8–N7
double bond of the purine base and subsequent oxidation of the
intermediate create the lesion. Therefore, as well as the usual
studies. Unprotected (5 R)-cdGuo and (5 S)-cdGuo have been
synthesized by photolysis of 8-bromo-2 -deoxyguanosine in
0
0
0
aqueous solutions in 26 % yield and an 5 S : 5 R ratio of 1 : 8
0
[7]
(Scheme 2). Apart from the problem of low yield, several
glycosidic bond, another covalent bond between the C5 sugar
attempts to selectively protect the two secondary OH groups in
both diastereomeric forms also failed. Here, we report an
ex novo synthetic strategy based on a radical cascade reaction
that provides easy access to these building blocks for DNA
modification.
O
N
O
N
N
N
N
N
NH
NH
O
O
NH2
NH2
O
HO
O
Results and Discussion
O
O
Our approach was based on previous experience in our labora-
[7,8]
tory with the reactivity of 8-bromopurine derivatives.
An
initial choice was the cyclization reaction in organic medium of
1
0
compound 6, obtained from 8-bromo-2 -deoxyguanosine and
t
[8]
BuMe SiCl following a standard procedure in 98 % yield.
2
O
N
O
N
N
N
N
N
O
N
O
N
NH
NH
N
N
N
N
O
O
NH
NH
NH2
NH2
Br
O
oxidation
O
HO
HO
NH2
NH2
O
hν (UV)
O
O
O
H O
2
OH
OH
5 26 % (5ꢀS/5ꢀR ꢁ 1:8)
3
2
NaI, 2 h
4
0
0
Scheme 1. Mode of formation of 5 ,8-cyclo-2 -deoxyguanosine (cdGuo)
Scheme 2.
in DNA.
*
This work is dedicated to the memory of Athel Beckwith, an inspiring teacher and a friend.
Journal compilation Ó CSIRO 2013
www.publish.csiro.au/journals/ajc

0
0
Synthesis of 5 ,8-Cyclo-2 -deoxyguanosine Derivatives
331
O
O
N
O
N
N
N
N
N
N
NH
NH
NH
Br
RO
Bu SnH (2.4 equiv.)
3
RO
RO
N
N
NH2
NH2
NH2
O
AIBN (2.4 equiv.)
O
O
ꢂ
CH CN,
3
2
.5 h, reflux
OR
OR
OR
7
3 % (7/8 ꢁ 5:1)
6
R ꢁ SiMe tBu
7 R ꢁ SiMe2tBu
8 R ꢁ SiMe2tBu
2
TBAF, THF
TBAF, THF
4
h, rt, 91 %
4 h, rt, 91 %
O
O
N
N
N
NH
NH
HO
HO
N
NH2
N
N
NH2
O
O
ꢂ
OH
OH
(5ꢀS)-5
(5ꢀR)-5
Scheme 3.
The cyclization reaction was initially tested using typical radical
chain conditions, i.e. a deoxygenated solution of 6 with Bu SnH
with respect to hydride concentration, in order to quantitatively
convert the starting material into the products. Based on previ-
3
0
[9,11]
(2.4 equiv.) and 2,2 -azobisisobutyronitrile (AIBN, 0.2 equiv.)
in refluxing CH CN. After 2 h, low conversion of the starting
bromide was observed. By increasing the amount of AIBN, the
ous findings,
drawn in Scheme 5 for the cyclization step. The Bu Sn radical,
these results suggest the reaction mechanism
ꢀ
3
3
ꢀ
initially generated from the reaction of the Me C( )CN radical
2
[
9]
reaction outcome improved. When the bis-protected deriva-
with tin hydride, abstracts a bromine atom from 6 or 9 to give an
aryl-type radical 16. Radical 16 abstracts a hydrogen atom either
tive 6 was treated with stoichiometric amounts of Bu SnH
3
0
0
(2.4 equiv.) and AIBN (2.4 equiv.) in CH CN under reflux for
3
intramolecularly from the 5 position to generate the C5 radical
17 or from Bu SnH to give the reduction product. Cyclization of
2
.5 h, the protected cyclopurine diastereomers 7 and 8 were
3
0
0
0
0
obtained in 73 % yield and in a ratio 5 S/5 R of 5 : 1 (Scheme 3),
in addition to 25 % yield of debrominated product. When
replacing Bu SnH with (TMS) SiH, the cyclonucleosides were
obtained in much lower yields (7 þ 8, 15 %). Compounds 7
and 8 were separated by reverse-phase chromatography and
17 is expected to produce radical 18 (a mixture of 5 S and 5 R
isomers). In the end, the oxidized products 7 þ 8 or 10 þ 11
ꢀ
derive from the reaction of radical 18 with Me C( )CN in
2
3
3
[
10]
accordance with the previous findings on Bu SnH-mediated
3
[
9,11]
intramolecular-homolytic substitution. In other words, the
reaction mechanism in Scheme 5 is an unusual non-chain
radical cascade protocol involving 1,6-H shift and 6-exo-trig
fully characterized. Bis-deprotection of the two secondary
0
0
silyloxy groups afforded the compounds (5 S)-5 and (5 R)-5
[
7]
[12]
cyclization. The reason for the low yield with (TMS) SiH as
respectively in high yields (Scheme 3). In order to use the
cyclopurines as building blocks for the synthesis of oligo-
nucleotides, thus obtaining the insertion of the lesion in a DNA
sequence, selective mono-deprotection of the 5 secondary
sililoxy group in 7 and 8 is necessary. Several attempts at this
reaction failed.
3
mediator is unknown; the interference of (TMS) SiBr as
3
by-product with the exocyclic amino group of the guanine
moiety could be play a role.
0
The radical cyclization 17 - 18 reported in Scheme 5 is
worthy of an additional comment. The rate constant for the
5
ꢂ1
Therefore, we changed strategy, introducing two different
protections for the OH groups before the cyclization step. For
example, compound 9 was prepared in two steps from 6 by
cyclization has been measured to be 6.1 ꢁ 10 s at 208C by
[
time-resolved spectroscopy for the unprotected radical and
6]
6
ꢂ1
t
,1 ꢁ 10 s at 308C by competitive kinetics for the BuMe Si-
2
0
0
[13]
primary 5 OH deprotection followed by a new 5 -protection, in
2 and 91 % yield respectively. Derivative 9 under the cycliza-
tion conditions described above afforded the protected cdGuo
diprotected radical.
Scheme 6 shows the chair transition
0
0
9
states in the pro-(5 S) and pro-(5 R) conformers of the cycliza-
0
tion. The cyclization of the pro-(5 S) conformer of unprotected
0
0
0
0
diastereomers 10 and 11 in 48 % yield and in a 5 S/5 R ratio of
6.5 : 1, together with a 50 % yield of debrominated product.
Column chromatography using reverse phase silica gel afforded
0 and 11 as pure compounds. The selective deprotection of the
2 -deoxyguanosin-5 -yl radicals was also studied theoretically
using the DFT B3LYP formalism with the 6-311þþG(d,p) basis
set and including Conductor Polarizable Continuum Model bulk
solvation effects. It was found that the solvation effects have
a little influence on reaction and activation energies. We found
that the solvent and nature of the substituent have a profound
,
[
14]
1
Et SiO moiety was achieved for each diastereomer after
3
coupling the amino group with DMF (Scheme 4).
It is worth recalling that in the presence of small amounts of
AIBN (0.1–0.2 equiv. with respect to Bu SnH), in a manner
3
0
effect on the stereoselectivity of the C5 radical cyclization.
0
0
Indeed, the 5 S /5 R isomer ratio varies from 1 : 8 for the
unprotected radical in water at room temperature (Scheme 2)
typical of radical chain reactions, the reaction does not proceed.
Stoichiometric amounts of AIBN and Bu SnH are necessary,
0
to 5 : 1 or 6.5 : 1 for the bulky 5 -OSiR substituent in CH CN at
3
3
3
ꢀ
which means nearly twice the amount of Me C( )CN radicals
2
808C (see Schemes 3 and 4). We suggest that a bulky silyloxy

3
32
M. A. Terzidis and C. Chatgilialoglu
O
N
O
N
O
N
N
N
N
N
NH
NH
NH
Br
1
1
1
R O
Bu SnH (2.4 equiv.)
R O
R O
3
N
N
NH2
NH2
NH2
O
AIBN (2.4 equiv.)
O
O
ꢂ
CH CN,
3
2
.5 h, reflux
OR
R ꢁ SiMe tBu, R1 ꢁ SiEt3
OR
10 R ꢁ SiMe2tBu, R1 ꢁ SiEt3
OR
4
8 % (10/11 ꢁ 6.5:1)
9
11 R ꢁ SiMe2tBu, R1 ꢁ SiEt3
2
(
CH ) NCH(OCH ) , THF
(CH ) NCH(OCH ) , THF
3
2
3
2
3
2
3 2
2.5 h, rt, 88 %
2.5 h, rt, 93 %
O
N
O
N
N
N
NH
NH
1
1
R O
R O
N
N
N
N
N
N
O
O
OR
OR
1
2 R ꢁ SiMe2tBu, R1 ꢁ SiEt3
13 R ꢁ SiMe2tBu, R1 ꢁ SiEt3
TBAF, THF
0 min, ꢃ18ꢄC
TBAF, THF
30 min, ꢃ18ꢄC
89 %
3
88 %
O
N
O
N
N
NH
NH
HO
N
HO
N
N
N
N
N
N
O
O
OR
OR
1
4 R ꢁ SiMe2tBu
15 R ꢁ SiMe2tBu
Scheme 4.
O
N
H N
H N
2
2
ꢂ
H
δ
N
N
NH
NH
7
NH
N
N
H
O
1
ꢃ
R O
O
δ
H
N
O
NH2
O
O
O
N
5ꢀ
O
6
or 9
N
N
7
R SiO
8
H
HO
5ꢀ
8
H
OR
3
Bu Sn
Bu SnBr
R SiO
3
3
3
1
6
1
9
20
O
N
O
N
0
0
Scheme 6. Chair transition states in the pro-(5 S) and pro-(5 R) confor-
N
N
N
N
mers of the cyclization.
H
NH
NH
1
1
R O
R O
NH2
NH2
substituent in the equatorial position favours the conformer 19 in
the transition state, thus affording a diastereomeric ratio favour-
O
O
0
ing the 5 S isomer, whereas in aqueous solution, only the
0
OR
OR
pro-(5 R) conformer can be stabilized by hydrogen bonding,
involving either the N3 of the base (such as in 20) or the oxygen
of the sugar ring.
1
8
17
Me CCN
2
Conclusions
Me C(H)CN
7 ꢂ 8 or 10 ꢂ 11
2
We have described a short and efficient synthetic sequence for
building blocks 14 and 15. The key step is a non-chain radical
Scheme 5. Reaction mechanism for the radical cascade protocol.

0
0
Synthesis of 5 ,8-Cyclo-2 -deoxyguanosine Derivatives
333
cascade involving bromine atom abstraction and formation of a
0
quantity of warm acetonitrile and subjected to column chro-
matography on silica C18 (CH CN/H O 7 : 3) to afford the two
pure diastereomers. Flash chromatography using the automated
system CombiFlash also works well (see Supplementary
Material).
C8 radical, radical translocation to the C5 position, and radical
cyclization to the aromatic ring, with a final oxidation step.
3
2
Ò
Incorporation of building blocks 14 and 15 into oligonucleotides
[
can be obtained following previously published procedures.
15]
0
0
0
0
Our efficient route to (5 R)- and (5 S)-5 ,8-cyclo-2 -
deoxyguanosine lesions represents an important facilitation for
further biochemical, biological, and biophysical studies. Indeed,
it was recently found that these tandem lesions accumulate
with aging in a tissue-specific manner (liver . kidney . brain),
providing evidence that DNA repair mechanisms are inadequate
7: d (400 MHz, CDCl ) 11.46 (bs, 1H), 6.17 (d, J 4.7, 1H),
H
3
6.04 (bs, 2H), 5.12 (d, J 6.1, 1H), 4.84 (dd, J 7.1, 4.2, 1H), 4.51
(d, J 6.1, 1H), 2.50 (dd, J 13.1, 7.3, 1H), 2.15 (dt, J 13.0, 4.6, 1H),
1.00 (s, 9H), 0.88 (s, 9H), 0.30 (s, 3H), 0.26 (s, 3H), 0.07 (s, 3H),
0.04 (s, 3H). d (101 MHz, CDCl ) 158.4, 153.5, 149.4, 144.7,
C
3
116.8, 87.1, 84.8, 69.6, 66.2, 46.5, 26.0, 25.7, 18.6, 17.8, ꢂ4.4,
[
16,17]
þ
to preserve the genetic material from these lesions. Results
from competitive transcription and adduct bypass assays
revealed that (5 S)-cdGuo strongly inhibits transcription in vitro
and in mammalian cells and induces transcriptional mutagenesis
ꢂ4.4, ꢂ4.6, ꢂ4.8. m/z (ESIþ) 494 [M þ H] . m/z (MS/MS) 362,
294, 264, 230, 187. Anal. Calc. for C H N O Si : C 53.52,
2
2
39
5
4
2
0
H 7.96, N 14.18. Found: C 53.50, H 7.95, N 14.22 %.
8: d (400 MHz, CDCl ) 11.72 (bs, 1H), 6.27 (d, J 4.5, 1H),
6.03 (bs, 2H), 4.68 (d, J 0.8, 1H), 4.51 (s, 1H), 4.25 – 4.13 (m,
H
3
[
18]
both in vitro and in vivo.
1
0
H), 2.38 (dd, J 13.1, 7.1, 1H), 2.21–2.09 (m, 1H), 0.93 (s, 9H),
.89 (s, 9H), 0.19 (s, 3H), 0.12 (s, 3H), 0.07 (s, 3H), 0.05 (s, 3H).
Experimental
d (101 MHz, CDCl ) 159.0, 153.6, 149.2, 142.9, 116.8, 71.4,
C
3
0
0
0
8
-Bromo-3 ,5 -O-bis(tert-butyldimethylsilyl)-2 -
67.0, 45.5, 25.9, 25.8, 18.4, 18.1, 1.1, ꢂ4.2, ꢂ4.7, ꢂ4.7, ꢂ4.9.
þ
deoxyguanosine 6
m/z (ESIþ) 494 [M þ H] . m/z (MS/MS) 362, 294, 264, 230,
0
187. Anal. Calc. for C H N O Si : C 53.52, H 7.96, N 14.18.
22 39 5 4 2
8
-Bromo-2 -deoxyguanosine (3.115 g, 9 mmol), imidazole
1.84 g, 27 mmol), and 4-dimethylaminopyridine (269 mg,
.2 mmol) were mixed in dry DMF (40 mL) and stirred until
homogenization. Next, tert-butyldimethylsilyl chloride (4.07 g,
7 mmol) was added and the reaction mixture was stirred at
Found: C 53.46, H 7.95, N 14.20 %.
(
2
0
0
0
0
(
5 S)- or (5 R)-5 ,8-Cyclo-2 -deoxyguanosine 5
Compound 7 or 8 (49.4 mg, 0.1 mmol) was dissolved in THF
5 mL). A solution of TBAF in THF (1 M, 15 mL, 0.3 mmol)
2
room temperature under an Ar atmosphere. After 18 h, the
reaction mixture was poured into ice water (100 mL) and the
precipitate was filtered and dried under vacuum. After chro-
matography on silica gel (AcOEt/CH Cl 6 : 4 v/v), 5.06 g
(
was then added, and the mixture was stirred at room temper-
ature. After 2 h, the starting material was consumed and the
solvent was removed. The resulting residue was chromato-
graphed on C18 reverse-phase silica (CH CN/H O 2 : 8) to
give 91 % yield of (5 S)-5 or (5 R)-5. Spectroscopic and
analytical data are identical with the previously described
2
2
(98 %) of a light yellow foam was obtained. dH (400 MHz,
CDCl ) 11.87 (bs, 1H), 6.23 (t, J 6.9, 1H), 6.12 (bs, 2H),
3
2
3
0
0
4
1
(
.78–4.69 (m, 1H), 3.92 (dd, J 9.2, 5.5, 1H), 3.85 (dd, J 10.7, 6.6,
H), 3.71 (dd, J 10.9, 4.8, 1H), 3.45 (dt, J 13.0, 6.5, 1H), 2.16
ddd, J 12.8, 6.8, 3.8, 1H), 0.93 (s, 9H), 0.87 (s, 9H), 0.14 (s, 6H),
[7]
reference compounds.
0
1
1
.03 (s, 3H), 0.01 (s, 3H). dC (101 MHz, CDCl ) 157.8,
3
53.2, 152.5, 122.2, 118.1, 87.8, 86.0, 72.8, 62.6, 36.6, 25.8,
þ
0
0
8
-Bromo-3 -O-tert-butyldimethylsilyl-2 -deoxyguanosine
8.1, 6.7, 4.4, ꢂ4.8. m/z (ESIþ) 576, 574 [M þ H] . m/z
0
0
0
8
(
-Bromo-3 ,5 -O-bis(tert-butyldimethylsilyl)-2 -deoxyguanosine
5.96 g, 10 mmol) was taken up in THF (125 mL). The mixture
was stirred until all material had dissolved and then water
62.5 mL) and acetic acid (62.5 mL) were added sequentially
final ratio of THF/H O/AcOH 2 : 1 : 1). The reaction mixture
was stirred at room temperature and was monitored with
HPLC-UV, reaching after 20 h a bis-protected/mono-protected/
unprotected ratio of 29 : 62 : 7. Dichloromethane (150 mL) was
added to the reaction mixture and subsequently the mixture was
(MS/MS) 230, 232.
0 0 0 0
5 S)-3 ,5 -O-Bis(tert-butyldimethylsilyl)-5 ,8-
(
(
(
0
0
0
0
cyclo-2 -deoxyguanosine 7 and (5 R)-3 ,5 -O-Bis(tert-
butyldimethylsilyl)-5 ,8-cyclo-2 -deoxyguanosine 8
2
0
0
0
0
0
8
(
-Bromo-3 ,5 -O-bis(tert-butyldimethylsilyl)-2 -deoxyguanosine
2.87 g, 5 mmol) was added to a round-bottom flask (1 L) con-
taining acetonitrile (500 mL) (previously flushed with Ar for
0 min) to give a 10 mM concentration. The mixture was heated
3
quenched carefully with saturated NaHCO solution to pH 7.5.
3
to boiling point and stirred until the solid was fully dissolved.
Next, AIBN (1.97 g, 12 mmol) and tributyltin hydride (3228 mL,
Next, the mixture was extracted with dichloromethane
(2 ꢁ 200 mL), the organic layer was dried over Na SO and
2
4
1
2 mmol) were added and the reaction mixture was stirred at
the solvent was removed by rotary evaporation. The crude
reaction product was purified by column chromatography on
silica gel (AcOEt/CH Cl 6 : 4, changed to AcOEt/CH Cl /
reflux temperature under an Ar atmosphere. The reaction
was monitored with HPLC-UV and after consumption of the
starting material (2.5 h), the reaction mixture was cooled, the
solvent was removed by rotary evaporation and the crude
residue was chromatographed on silica gel (AcOEt/hexane 5 : 1)
2
2
2
2
MeOH 2 : 1 : 1) and 2.76 g (60 %, or 92 % based on starting
0
material recovered) of 8-bromo-3 -O-tert-butyldimethylsilyl-
0
2 -deoxyguanosine was obtained as a light-brown foam. dH
(400 MHz, CDCl ) 12.00 (bs, 1H), 6.66 (bs, 2H), 6.28 (dd, J 9.5,
0
0
0
0
0
to give 1.799 g (73 %) of the (5 S)- and (5 R)-3 ,5 -O-bis(tert-
0
3
butyldimethylsilyl)-5 ,8-cyclo-2 -deoxyguanosine in a 5 : 1
mixture (based on HPLC-UV analysis) as a white foam.
5.6, 1H), 4.64 (d, J 5.1, 1H), 4.11 (s, 1H), 3.97 (d, J 12.0, 1H),
3.71 (d, J 12.3, 1H), 3.00–2.81 (m, 1H), 2.14 (dd, J 12.8, 5.3,
1H), 1.88 (s, 1H), 0.93 (s, 9H), 0.12 (s, 3H), 0.12 (s, 3H). dC
0
0
0
A second fraction of 3 ,5 -O-bis(tert-butyldimethylsilyl)-2 -
deoxyguanosine (620 mg, 25 %) was isolated as a white solid.
The (5 S)- and (5 R)-3 ,5 -O-bis(tert-butyldimethylsilyl)-5 ,8-
cyclo-2 -deoxyguanosine mixture was dissolved in a small
(101 MHz, CDCl ) 157.7, 153.5, 151.1, 121.2, 118.3, 90.0, 88.2,
3
0
0
0
0
0
þ
63.2, 25.8, 18.0, ꢂ4.7, ꢂ4.7. m/z (ESIþ) 482, 484 [M þ Na] .
0
m/z (MS/MS) 230, 232.

3
34
M. A. Terzidis and C. Chatgilialoglu
0
0
8
-Bromo-3 -O-tert-butyldimethylsilyl-5 -O-
dropwise and the reaction mixture was stirred under an Ar
atmosphere. The reaction was monitored by TLC (CH Cl /
0
triethylsilyl-2 -deoxyguanosine 9
2
2
0
0
MeOH 97 : 3). After consumption of the starting material (2.5 h),
the solvent was removed by rotary evaporation and the mixture
was subjected to silica gel chromatography (CH Cl /MeOH
8
5
-Bromo-3 -O-tert-butyldimethylsilyl-2 -deoxyguanosine (2.3 g,
mmol) and imidazole (1.02 g, 15 mmol) were taken up in dry
dichloromethane (30 mL) and stirred until homogenization.
Next, triethylsilyl chloride (2.517 mL, 15 mmol) was added
dropwise and the reaction mixture was stirred at room temper-
ature under an Ar atmosphere. After 45 min, TLC showed the
consumption of the starting material, so the mixture was
quenched with saturated NaHCO3 solution, extracted with
dichloromethane and washed with 5 % NaHCO3 solution
2
2
0
6
from 99 : 1 to 97 : 3) to give 966 mg (88 %) of (5 S)-N -
0
0
0
dimethylformyl-3 -O-tert-butyldimethylsilyl-5 -O-triethylsilyl-2 -
deoxyguanosine as a white foam. d (400 MHz, CDCl ) 9.55 (s,
H
3
1
H), 8.55 (s, 1H), 6.21 (d, J 4.6, 1H), 5.12 (d, J 6.0, 1H), 4.87 (dd,
J 7.0, 4.2, 1H), 4.50 (d, J 6.0, 1H), 3.18 (s, 3H), 3.09 (s, 3H), 2.49
dd, J 13.0, 7.3, 1H), 2.14 (dt, J 12.7, 4.4, 1H), 1.03 (t, J 7.8, 9H),
.87 (s, 9H), 0.80 (td, J 15.1, 7.3, 6H), 0.07 (s, 3H), 0.04 (s, 3H).
d (101 MHz, CDCl ) 157.9, 157.9, 156.6, 147.9, 144.8, 119.9,
(
0
(
2 ꢁ 50 mL). The organic layer was dried over Na SO and the
2
4
solvent was removed by rotary evaporation. The crude reaction
mixture was chromatographed on silica gel (AcOEt/CH Cl /
C
3
8
7.1, 84.6, 69.6, 65.8, 46.4, 41.3, 35.1, 25.7, 17.9, 6.9, 4.8, ꢂ4.7,
2
2
þ
0
ꢂ4.9. m/z (ESIþ) 418 [M þ H] . m/z (MS/MS) 349, 319, 285.
MeOH/TEA 5 :3 :1 : 0.05) to give 2.248 g (91%) of 8-bromo-3 -
0
0
O-tert-butyldimethylsilyl-5 -O-triethylsilyl-2 -deoxyguanosine
as a light-brown foam. d (400 MHz, CDCl ) 11.85 (bs, 1H),
0
0
(5 R)-N-Dimethylformamidine-3 -O-tert-
H
3
0
0
0
butyldimethylsilyl-5 -O-triethylsilyl-5 ,8-cyclo-2 -
deoxyguanosine 13
6.48 (bs, 2H), 6.21 (t, J 7.0, 1H), 4.77–4.65 (m, 1H), 3.93 (ddd,
J 6.8, 5.3, 3.2, 1H), 3.83 (dd, J 10.7, 6.9, 1H), 3.69 (dd, J 10.7,
.2, 1H), 3.48 (dt, J 13.1, 6.5, 1H), 2.16 (ddd, J 13.0, 6.7, 3.5,
H), 1.00–0.83 (m, 18H), 0.56 (q, J 7.9, 6H), 0.13 (s, 6H). dC
101 MHz, CDCl ) 157.8, 153.2, 152.5, 122.2, 118.1, 87.8, 86,
0
0
The procedure described for (5 S)-3 -O-tert-butyldimethylsilyl-
0
5
1
(
7
5
0
0
5
starting from 140 mg (0.28 mmol) of the 5 R diastereomer. After
-O-triethylsilyl-5 ,8-cyclo-2 -deoxyguanosine was followed
0
3
0
6
purification, 142 mg (91 %) of (5 R)-N -dimethylformyl-3 -O-
0
2.8, 62.6, 36.6, 25.8, 18.1, 6.7, 4.3, ꢂ4.6, ꢂ4.7. m/z (ESIþ)
0
0
þ
tert-butyldimethylsilyl-5 -O-triethylsilyl-2 -deoxyguanosine was
obtained as a white foam. d (400 MHz, CDCl ) 9.63 (bs, 1H),
76, 574 [M þ H] . m/z (MS/MS) 230, 232.
H
3
0 0 0 0
5 S)-3 -O-tert-Butyldimethylsilyl-5 -O-triethylsilyl-5 ,8-
8.54 (s, 1H), 6.31 (d, J 4.5, 1H), 4.68 (d, J 1.3, 1H), 4.51 (s, 1H),
4.17 (dd, J 7.2, 4.4, 1H), 3.17 (s, 3H), 3.08 (d, J 0.4, 3H), 2.31
(dd, J 13.0, 7.3, 1H), 2.20–2.06 (m, 1H), 0.96 (t, J 7.9, 9H), 0.86
(
0
0
0
cyclo-2 -deoxyguanosine 10 and (5 R)-3 -O-tert-
Butyldimethylsilyl-5 -O-triethylsilyl-5 ,8-cyclo-2 -
deoxyguanosine 11
0
0
0
(s, 9H), 0.75–0.65 (m, 6H), 0.04 (s, 3H), 0.02 (s, 3H). dC
(101 MHz, CDCl ) 158.1, 158, 156.9, 147.8, 142.4, 119.9, 90.3,
3
Starting from 9, 2.87 g (5 mmol) and applying the same condi-
0
8
4.8, 71.3, 66.5, 45.7, 41.4, 35.2, 25.8, 25.7, 18.0, 6.8, 4.8, ꢂ4.8,
0
0
0
tions described for the synthesis of (5 R)- and (5 S)-3 ,5 -O-
0
þ
ꢂ5. m/z (ESIþ) 418 [M þ H] . m/z (MS/MS) 349, 319, 285.
0
bis(tert-butyldimethylsilyl)5 ,8-cyclo-2 -deoxyguanosine, the
0
0
0
0
0
0
synthesis of (5 S)- and (5 R)-3 -O-tert-butyldimethylsilyl-5 -O-
0
(5 S)-N-Dimethylformamidine-3 -O-tert-butyldimethylsilyl-
0 0
5 ,8-cyclo-2 -deoxyguanosine 14
0
triethylsilyl-5 ,8-cyclo-2 -deoxyguanosine was achieved. After
purification by silica gel chromatography (AcOEt/hexanes
0
6
5 S)-N -Dimethylformyl-3 -O-tert-butyldimethylsilyl-5 -O-
0
0
(
0
5
: 1), 1.11 g (45 %) of a diastereomeric mixture of (5 S) and
0
0
triethylsilyl-2 -deoxyguanosine (549 mg, 1 mmol) was taken up
in THF (50 mL). The solution was cooled down to ꢂ188C, 1 M
solution of TBAF in THF (0.52 mL, 0.52 mmol) was added, and
the mixture was stirred at this temperature. The reaction was
monitored by TLC (CH Cl /CH OH 93 : 7), and after 30 min,
(
5 R) in a 6.5 : 1 ratio (based on HPLC-UV analysis) was
obtained as a white foam along with 1.24 g (50 %) reduced
product. The diastereomeric mixture was separated by C18
reverse-phase silica gel chromatography (CH CN/H O 7 : 3).
3
2
2
2
3
1
0: d (400 MHz, CDCl ) 11.38 (bs, 1H), 6.17 (d, J 4.6, 3H),
H 3
when the starting material was consumed, was quenched with a
saturated solution of NaHCO and extracted with ethyl acetate
5
2
9
0
1
.09 (d, J 6.0, 1H), 4.83 (dd, J 7.0, 4.5, 1H), 4.48 (d, J 6.1, 1H),
.47 (dd, J 13.1, 7.3, 1H), 2.12 (dt, J 12.9, 4.3, 1H), 1.03 (t, J 7.9,
H), 0.87 (s, 9H), 0.78 (ddd, J 12.9, 7.9, 4.5, 6H), 0.06 (s, 3H),
.03 (s, 3H). d (101 MHz, CDCl ) 158.3, 153.7, 149.3, 144.4,
3
(
2 ꢁ 200 mL). The organic layer was dried over anhydrous
Na SO , filtered, and the solvent was subsequently removed
2
4
C
3
by rotary evaporation. The resulting residue was subjected
to column chromatography on silica gel (CH Cl /CH OH
16.7, 86.9, 84.7, 69.6, 65.8, 46.2, 25.7, 17.8, 6.8, 4.8, ꢂ4.7,
þ
2
2
3
ꢂ4.9. m/z (ESIþ) 494 [M þ H] . m/z (MS/MS) 362, 294,
0
6
from 100 : 0 to 92 : 8) to give 382 mg (88 %) of (5 S)-N -
2
64, 230, 230.
1: d (400 MHz, CDCl ) 11.65 (bs, 1H), 6.28 (d, J 4.5, 1H),
0
0
dimethylformyl-3 -O-tert-butyldimethylsilyl-2 -deoxyguanosine
as a white foam. d (400 MHz, CDCl ) 8.79 (bs, 1H), 8.56 (s,
1
H
3
H
3
6
4
.05 (bs, 2H), 4.68 (d, J 1.0, 1H), 4.52 (s, 1H), 4.20 (dd, J 6.9,
.6, 1H), 2.37 (dd, J 13.1, 7.2, 1H), 2.2 ꢂ2.09 (m, 1H), 0.97
1
1
2
0
1
2
2
H), 6.24 (d, J 4.8, 1H), 5.23 (d, J 6.2, 1H), 4.87 (dd, J 7.4, 4.3,
H), 4.68 (d, J 6.2, 1H), 4.36 (bs, 1H), 3.19 (s, 3H), 3.09 (s, 3H),
.51 (dd, J 13.3, 7.4, 1H), 2.20 (dt, J 13.1, 4.6, 1H), 0.88 (s, 9H),
.11 (s, 3H), 0.08 (s, 3H). d (101 MHz, CDCl ) 158, 157.4,
(t, J 7.9, 8H), 0.88 (s, 9H), 0.76–0.63 (m, 6H), 0.07 (s, 3H), 0.04
(
s, 3H). d (101 MHz, CDCl ) 158.9, 153.7, 149.2, 142.3, 116.8,
C
3
C
3
9
0.2, 84.9, 71.4, 66.6, 45.5, 25.7, 18.0, 6.8, 4.9, ꢂ4.7, ꢂ4.9. m/z
57.0, 147.7, 146.4, 119.5, 86.3, 69.4, 64.2, 46.5, 41.4, 35.2,
þ
þ
(ESIþ) 494 [M þ H] . m/z (MS/MS) 362, 294, 264, 230, 230.
5.8, 18.0, ꢂ4.7, ꢂ5. m/z (ESIþ) 435 [M þ H] . m/z (MS/MS)
85, 235. Anal. Calc. for C H N O Si: C 52.51, H 6.96,
1
9 30 6 4
0 0
5 S)-N-Dimethylformamidine-3 -O-tert-butyldimethylsilyl-
(
N 19.34. Found: C 52.55, H 6.96, N 19.30 %.
0
0
0
5
-O-triethylsilyl-5 ,8-cyclo-2 -deoxyguanosine 12
0 0
(5 R)-N-Dimethylformamidine-3 -O-tert-
0
0
0
0
(
5 S)-3 -O-tert-Butyldimethylsilyl-5 -O-triethylsilyl-5 ,8-cyclo-
0
0
0
butyldimethylsilyl-5 ,8-cyclo-2 -deoxyguanosine 15
2 -deoxyguanosine (988 mg, 2 mmol) was dissolved in dry
THF (15 mL) and stirred at room temperature. Dimethylfor-
0
6
0
The same procedure as for (5 S)-N -dimethylformyl-3 -O-
0 0
tert-butyldimethylsilyl-5 -O-triethylsilyl-2 -deoxyguanosine
mamide diethylacetal (1371 mL, 8 mmol) was then added

0
0
Synthesis of 5 ,8-Cyclo-2 -deoxyguanosine Derivatives
335
0
was followed for the 5 R diastereomer. Starting from
1
[4] C. Chatgilialoglu, C. Ferreri, M. A. Terzidis, Chem. Soc. Rev. 2011, 40,
1368. doi:10.1039/C0CS00061B
[5] B. Aydogan, D. T. Marshall, S. G. Swarts, J. E. Turner, A. J. Boone,
0
6
0
10 mg (0.2 mmol) of (5 R)-N -dimethylformyl-3 -O-tert-
0
0
butyldimethylsilyl-5 -O-triethylsilyl-2 -deoxyguanosine, 77 mg
89 %) of product was obtained as a white foam. d (400 MHz,
CDCl ) 9.70 (bs, 1H), 8.54 (s, 1H), 6.29 (d, J 4.7, 1H), 4.71
N. G. Richards, W. E. Bolch, Radiat. Res. 2002, 157, 38. doi:10.1667/
(
H
0
033-7587(2002)157[0038:SSOATT]2.0.CO;2
3
[
6] C. Chatgilialoglu, M. D’Angelantonio, G. Kciuk, K. Bobrowski,
Chem. Res. Toxicol. 2011, 24, 2200. doi:10.1021/TX2003245
7] C. Chatgilialoglu, R. Bazzanini, L. B. Jimenez, M. A. Miranda, Chem.
Res. Toxicol. 2007, 20, 1820. doi:10.1021/TX700282X
(
(
s, 1H), 4.67 (s, 1H), 4.43 (bs, 1H), 4.26 (dd, J 7.1, 4.3, 1H), 3.19
s, 3H), 3.07 (s, 3H), 2.39 (dd, J 13.2, 7.2, 1H), 2.15 (dt, J 13.7,
4.5, 1H), 0.87 (s, 9H), 0.06 (s, 3H), 0.03 (s, 3H). d (101 MHz,
C
CDCl ) 158.2, 157.9, 157.2, 147.7, 143.0, 119.5, 89.1, 84.6,
[
3
[8] M. L. Navacchia, C. Chatgilialoglu, P. Montevecchi, J. Org. Chem.
7
4
1.6, 65.6, 45.7, 41.5, 35.2, 25.7, 18.0, ꢂ4.8, ꢂ4.9. m/z (ESIþ)
2006, 71, 4445. doi:10.1021/JO060197Z
þ
35 [M þ H] . m/z (MS/MS) 285, 235. Anal. Calc. for
[9] A. L. J. Beckwith, V. W. Bowry, W. R. Bowman, E. Mann, J. Parr,
J. M. D. Storey, Angew. Chem. Int. Ed. 2004, 43, 95. doi:10.1002/
ANIE.200352419
C H N O Si: C 52.51, H 6.96, N 19.34. Found: C 52.50,
1
H 6.95, N 19.35 %.
9 30 6 4
[
[
10] C. Chatgilialoglu, Chem.–Eur. J. 2008, 14, 2310. doi:10.1002/CHEM.
2
00701415
11] M. L. Navacchia, A. Manetto, P. Montevecchi, C. Chatgilialoglu, Eur.
Supplementary Material
Further details about experimental procedures and data analysis
are available on the Journal’s website.
J. Org. Chem. 2005, 4640. doi:10.1002/EJOC.200500398
[12] A. Baralle, A. Baroudi, M. Daniel, L. Fensterbank, J.-P. Goddard,
E. Lac oˆ te, M.-H. Larraufie, G. Maestri, M. Malacria, C. Ollivier,
in Encyclopedia of Radicals in Chemistry, Biology and Materials (Eds
C. Chatgilialoglu, A. Studer) 2012, pp. 729–765. (Wiley: Chichester).
Acknowledgements
Financial support from the Ministero dell’Istruzione, dell’Universit a` della
Ricerca (PRIN-2009K3RH7N_002) and Marie Curie Intra-European
Fellowship (CYCLOGUO–298555) is gratefully acknowledged.
[
13] A. Manetto, D. Georganakis, T. Gimisis, L. Leondiadis, P. Mayer,
T. Carell, C. Chatgilialoglu, J. Org. Chem. 2007, 72, 3659. doi:10.1021/
JO062518C
[
14] R. B. Zhang, L. A. Eriksson, Chem. Phys. Lett. 2006, 417, 303.
doi:10.1016/J.CPLETT.2005.10.020
[15] A. Romieu, D. Gasparutto, J. Cadet, Chem. Res. Toxicol. 1999, 12,
References
[
1] J. Cadet, T. Douki, J.-L. Ravanat, Acc. Chem. Res. 2008, 41, 1075.
doi:10.1021/AR700245E
412. doi:10.1021/TX9802668
[
2] J. Cadet, T. Douki, J.-L. Ravanat, R. Wagner, D. Gasparutto, in
[16] J. Wang, B. Yuan, C. Guerrero, R. Bahde, S. Gupta, Y. Wang, Anal.
Chem. 2011, 83, 2201. doi:10.1021/AC103099S
Encyclopedia of Radicals in Chemistry, Biology and Materials
(
Eds C. Chatgilialoglu, A. Studer) 2012, pp. 1319–1344. (Wiley:
[17] J. Wang, C. L. Clauson, P. D. Robbins, L. J. Niedernhofer, Y. Wang,
Aging Cell 2012, 11, 714. doi:10.1111/J.1474-9726.2012.00828.X
[18] C. You, X. Dai, B. Yuan, J. Wang, J. Wang, P. J. Brooks,
L. J. Niedernhofer, Y. Wang, Nat. Chem. Biol. 2012, 8, 817.
doi:10.1038/NCHEMBIO.1046
Chichester).
[
3] N. Belmadoui, F. Boussicault, M. Guerra, J.-L. Ravanat, C. Chatgi-
lialoglu, J. Cadet, Org. Biomol. Chem. 2010, 8, 3211. doi:10.1039/
C004531D