Synthesis of 6′-branched locked nucleic acid by a radical TEMPO-scavanged stereoselective mercury cyclization

Article abstract of DOI:10.1021/jo801081t

(Chemical Equation Presented) A 6′(R)-hydroxymethyl derivative of the locked nucleic acid (LNA)-thymidine monomer has been synthesized by a stereoselective mercury cyclization and subsequent use of TEMPO as a radical scavenger. This compound was converted to an azide derivative, which in a Huisgen-type [3 + 2] cycloaddition afforded a double-headed nucleoside with a triazole linking an additional thymine to the opposition of the LNA-nucleoside monomer.

Full text of DOI:10.1021/jo801081t

Synthesis of 6′-Branched Locked Nucleic Acid by
a Radical TEMPO-Scavanged Stereoselective
Mercury Cyclization
FIGURE 1. An LNA monomer, an amino-LNA monomer, and a 6′-
branched LNA monomer.
Gerald Enderlin and Poul Nielsen*
Nucleic Acid Center, Department of Physics and Chemistry,
UniVersity of Southern Denmark, 5230 Odense M, Denmark
Figure 1) constitutes a hitherto unexplored center for function-
alization. This carbon complements the 2′-amino atom as well
as the 3′-position, giving two different positions (pro-S and pro-
R) for displaying nucleic acid decorations in the minor groove.
The preparation of a universal building block for branching LNA
in the 6′-position is however a challenging task. Herein we
demonstrate the first synthesis of a 6′-hydroxymethyl LNA
monomer8 and its conversion into a 6′-azidomethyl analogue,
whose potential for Huisgen [3 + 2] cycloaddition is demon-
strated in the preparation of a double-headed nucleoside.9-11
In the design of a general tool for branching LNA on the
6′-position, we envisioned the synthesis of a 6′-hydroxymethyl
derivate as the key goal. The hydroxymethyl group can easily
be further derivatized in numerous ways (oxidation, alkylation,
activation/substitution). We based the preparation on the known
4′-C-vinylribofuranose derivative 112 (Scheme 1), taking ad-
vantage of two benzyl protecting groups to be removed
simultaneously at the end of the synthesis. Compound 1 was
efficiently synthesized in seven steps from 1,2;5,6-di-O-isopro-
pylidene-R-D-allofuranose using the well-known aldol conden-
sation with formaldehyde13 as applied in the preparation of
LNA,5 a selective benzylation,5,14 and a high-yielding oxidation/
Wittig olefination sequence.12 Earlier studies on the preparation
of LNA and derivatives thereof have shown that the introduction
of the nucleobase cannot be performed after the ring-closing
step forming the 2′,4′-bridge.15 Therefore, compound 1 was
converted to the diacetate 2, which was coupled to thymine using
pon@ifk.sdu.dk
ReceiVed May 19, 2008
A 6′(R)-hydroxymethyl derivative of the locked nucleic acid
(LNA)-thymidine monomer has been synthesized by a
stereoselective mercury cyclization and subsequent use of
TEMPO as a radical scavenger. This compound was con-
verted to an azide derivative, which in a Huisgen-type [3 +
2] cycloaddition afforded a double-headed nucleoside with
a triazole linking an additional thymine to the 6′-position of
the LNA-nucleoside monomer.
Locked nucleic acid (LNA) has received significant attention
as a nucleic acid analogue displaying unprecedented recognition
of complementary nucleic acids.1 Therapeutic antisense proper-
ties2 of LNA are very promising,3 but also the potential in
nanoscale engineering and microarray construction has been
recognized.4 LNA is defined as oligonucleotides containing at
least one LNA monomer (Figure 1,) which is a bicyclic
nucleoside analogue conformationally locked in the N-type
conformation and therefore preorganizing the LNA for A-type
duplex formation.5 Hence, the 2′-amino group of amino-LNA
(see monomer in Figure 1) has been used as the branching point
for a diverse series of functionalized oligonucleotides.6 The 3′-
position of LNA has also been used for functionalization.7 The
carbon atom in the 2′,4′-bridge (herein defined as the 6′-carbon,
(6) (a) Sørensen, M. D.; Petersen, M.; Wengel, J. Chem. Commun. 2003,
2130–2131. (b) Hrdlicka, P. J.; Babu, B. R.; Sørensen, M. D.; Harrit, N.; Wengel,
J. J. Am. Chem. Soc. 2005, 127, 13293–13299. (c) Lindegaard, D.; Madsen,
A. S.; Astakhova, I. V.; Malakhov, A. D.; Babu, B. R.; Korshun, V. A.; Wengel,
J. Bioorg. Med. Chem. 2008, 16, 94–99.
(7) (a) Meldgaard, M.; Hansen, F. G.; Wengel, J. J. Org. Chem. 2004, 69,
6310–6322. (b) Sharma, P. K.; Kumar, S.; Nielsen, P. Nucleosides, Nucleotides
Nucleic Acids 2007, 26, 1505–1508.
(8) Recently, a PCT Int. Appl. demonstrating an alternative synthetic method
towards 6′-branching LNA has been published. A non-stereoselective multistep
synthesis has been applied based on dihydroxylation of a differently protected
analogue of 1 and multiple protecting group manipulations affording 6′-epimeric
mixtures: Swayze, E. E.; Seth, P. P. WO2007090071, 2007.
(9) We define nucleosides with two nucleobases as double-headed nucleo-
sides. These have recently been studied as building blocks in oligonucleotides
by the group of Herdewijn10 and us.11
(10) (a) Tongfei, W.; Froeyen, M.; Schepers, G.; Mullens, K.; Rozenski, J.;
Busson, R.; Van Aerschot, A.; Herdewijn, P. Org. Lett. 2004, 6, 51–54. (b)
Tongfei, W.; Nauwelaerts, K.; Van Aerschot, A.; Froeyen, M.; Lescrinier, E.;
Herdewijn, P. J. Org. Chem. 2006, 71, 5423–5431.
(11) (a) Pedersen, S. L.; Nielsen, P. Org. Biomol. Chem. 2005, 3, 3570–
3575. (b) Christensen, M. S.; Madsen, C. M.; Nielsen, P. Org. Biomol. Chem.
2007, 5, 1586–1594. (c) Christensen, M. S.; Bond, A.; Nielsen, P. Org. Biomol.
Chem. 2008, 6, 81–91.
(1) (a) Petersen, M.; Wengel, J. Trends Biotechnol. 2003, 21, 74–81. (b)
Kaur, H.; Babu, B. R.; Maiti, S. Chem. ReV. 2007, 107, 4672–4697.
(2) For a general review on antisense, see: Kurreck, J. Eur. J. Biochem. 2003,
270, 1628–1644.
(3) (a) Jepsen, J. S.; Wengel, J. Curr. Opin. Drug DiscoVery DeV. 2004, 7,
188–194. (b) Vester, B.; Wengel, J. Biochemistry 2004, 43, 13233–13241.
(4) (a) Wengel, J. Org. Biomol. Chem. 2004, 2, 277–280. (b) Seferos, D. S.;
Giljohann, D. A.; Rosi, N. L.; Mirkin, C. A. ChemBioChem 2007, 8, 1230–
1232. (c) Ahlborn, C.; Siegmund, K.; Richert, C. J. Am. Chem. Soc. 2007, 129,
15218–15232.
(12) Morita, K.; Takagi, M.; Hasegawa, C.; Kaneko, M.; Tsutsumi, S.; Sone,
J.; Ishikawa, T.; Imanishi, T.; Koizumi, M. Bioorg. Med. Chem. 2003, 11, 2211–
2226.
(5) (a) Singh, S. K.; Nielsen, P.; Koshkin, A. A.; Wengel, J. Chem. Commun.
1998, 455–456. (b) Koshkin, A. A.; Singh, S. K.; Nielsen, P.; Rajwanshi, V. K.;
Kumar, R.; Meldgaard, M.; Olsen, C. E.; Wengel, J. Tetrahedron 1998, 54, 3607–
3630. (c) Obika, S.; Nanbu, D.; Hari, Y.; Andoh, J.; Morio, K.; Doi, T.; Imanishi,
T. Tetrahedron Lett. 1998, 39, 5401–5404.
(13) Youssefyeh, R. D.; Verheyden, J. P. H.; Moffatt, J. G. J. Org. Chem.
1979, 44, 1301–1309.
(14) Waga, T.; Nishizaki, T.; Miyakawa, I.; Ohrui, H.; Meguro, H. Biosci.
Biotechnol. Biochem. 1993, 57, 1433–1438.
(15) Nielsen, P.; Wengel, J. Chem. Commun. 1998, 2645–2646.
10.1021/jo801081t CCC: $40.75
Published on Web 08/02/2008
2008 American Chemical Society
J. Org. Chem. 2008, 73, 6891–6894 6891

SCHEME 1. Synthesis of the 6′(R)-Hydroxymethyl
constitute a hydroxylated analogue of the ring-enlarged analogue
of LNA called ENA.12 Finally, the hydroxy group appeared as
a triplet proving a primary alcohol and the LNA-constitution
of 6. The 6′-configuration was not verified at this stage, as NOE-
difference spectroscopy was hampered by the overlap of crucial
1H NMR signals. Later derivatization (see below) proved a
6′(R)-configuration.
LNA-Thymidine Monomera
Even though the target ring closure was obtained by the
epoxidation method, the yield was too low for any practical
use. A protection of the nucleobase as its 4-O-(2,6-dimeth-
yl)phenyl derivative did not improve the epoxidation (see
Supporting Information). This protecting group has been used
by Sekine and co-workers, allowing an efficient dihydroxylation
of a 2′-O-allyl group of a uridine derivative,21 but in our case
it let to even more pronounced nucleobase oxidation. Dihy-
droxylation on 4 using OsO4 afforded also pronounced nucleo-
base oxidation. Iodocyclization based on the addition of iodine
and following ring closure failed to give any product. Electro-
philic activation of the double bond based on NBS or NIS also
failed. The solution was found in activation by mercury(II)
trifluoroacetate.
a Reagents and conditions: (a) Ac2O, H2SO4, AcOH (90%); (b) thymine,
N,O-bis(trimethylsilyl)acetamide, TMS-triflate, 1,2-DCE (85%); (c)
NaOCH3, CH3OH (95%); (d) mCPBA, DCM; (e) CSA, DCM (21% from
4); (f) Hg(OCOCF3)2, THF (aq), then 10% NaBr (aq), DCM (98%); (g)
NaBH4, O2, DMF (16% 6 and 55% 4); (h) NaBH4, TEMPO, DMF (77%);
(i) Zn, AcOH, THF (aq) (83%); (j) H2, Pd(OH)2/C, EtOAc (65%). T )
thymin-1-yl.
Hanessian and co-workers have earlier reported the prepara-
tion of a six-membered ring forming a bicyclic nucleoside based
on a mercury cyclization with mercury(II) acetate.22 We decided
to apply the more reactive trifluoroacetate in a similar protocol
and observed a quantitative reaction. The protocol had to be
changed, however, in order to extract the mercury bromide
derivative 7 from a sodium bromide solution, giving almost
quantitative yield. The subsequent reduction of the mercuric
derivative by NaBH4 in a solution of DMF saturated by
continuous bubbling with oxygen23 afforded, however, the
desired product 6 in only 16% yield as well as the starting
compound 4 in 55% yield. Thus, the primary radical formed in
the reduction has been reorganized into a more stable form by
reopening the constrained ring previously formed. With a large
excess of TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxyl) in the
reduction reaction,23 however, the primary radical was very
efficiently scavenged as the TEMPO-derivative 8 in 77% yield.
Cleavage of the oxygen-nitrogen bond occurred thereafter in
a good yield by the application of Zn dust in neat acetic acid to
give the 6′-hydroxymethyl LNA derivative 6. The overall yield
of 6 was 63% from 4 as compared to the 21% yield obtained
by the epoxidation method. The method was completely
stereoselective affording the same 6′-epimer as obtained by the
epoxidation method. Global debenzylation by hydrogenation
afforded the branched LNA-thymidine monomer 9.
The versatility of the hydroxymethyl compound 6 as the
starting point for various modifications was demonstrated by
its conversion to the azide (Scheme 2). Hence, mesylation
afforded 10, on which a microwave-heated nucleophilic sub-
stitution with sodium azide gave the azide derivative 11. The
azide was reduced to the primary amine 12 by applying Zn dust.
The two derivatives 11 and 12 were used in the final solution
of the 6′-configuration of the present branched LNA derivatives.
Thus, NOE-difference spectroscopy revealed in both cases
strong mutual contacts between the H1′ and one of the H7′-
protons of 3-4%. No contacts were seen between H1′ and H6′.
a Vorbru¨ggen-type16 protocol, affording the protected 4′-
vinylribonucleoside 3. Deacetylation gave compound 4 consti-
tuting a key material for a ring closure giving the 2′,4′-ring and
a 6′-hydroxymethyl group.
Our first attempt was based on the epoxidation of the terminal
olefin followed by a ring-opening nucleophilic attack from the
2′-hydroxy group. However, the vinyl group demonstrated a
low reactivity toward all attempted reagents, yielding a maxi-
mum of 20% of the target epoxide 5 by the application of
mCPBA. Protocols based on DMDO,17 tBuOOH,18 or H2O2
(Payne oxidation)19 all failed to improve this situation, though
another 20% yield of 5 was obtained with DMDO. Some
oxidation of the nucleobase was also detected in all cases.
Similar difficulties have been observed with the epoxidation of
5′-olefinated thymidine derivatives.11b,20 Despite the low yield,
compound 5 was obtained as only one stereoisomer, revealing
a substrate for the following ring closure. This was performed
under acidic conditions using CSA, giving the targeted 6′-
hydroxymethyl-LNA monomer 6 in 50% yield. A one-pot
epoxidation using mCPBA and CSA was optimized to give 6
in 21% overall yield from 4. The constitution of compound 6
was verified by NMR spectroscopy. The singlets for H1′ and
H3′ are characteristics of the constrained N-type conformation
of an LNA derivative,5 and the chemical shift for the H1′ at
5.56 ppm is characteristic for LNA (5.51 ppm reported for the
thymidine monomer)5 and not for ENA, for which shifts around
6.0 ppm have been reported.12 Hence, an alternative ring closure
forming a six-membered ring and a secondary alcohol would
(16) Vorbru¨ggen, H.; Krolikiewicz, K.; Bennua, B. Chem. Ber. 1981, 114,
1234–1255.
(17) Adam, W.; Hadjiarapoglou, L.; Smerz, A. Chem. Ber. 1991, 124, 227–
232.
(18) Crimminns, M. T.; Caussanel, F. J. Am. Chem. Soc. 2006, 128, 3128–
3129.
(19) (a) Payne, G. B.; Williams, P. J. Org. Chem. 1961, 26, 651–658. (b)
(21) Okamoto, I.; Shohda, K.-i.; Seio, K.; Sekine, M. J. Org. Chem. 2003,
68, 9971–9982.
(22) Hanessian, S.; Kloss, J.; Sugawara, T. J. Am. Chem. Soc. 1986, 108,
2758–2759.
Ravn, J.; Thorup, N.; Nielsen, P. J. Chem. Soc., Perkin Trans. 1 2001, 1855–
1861.
(20) Trafelet, H.; Stulz, E.; Leumann, C. HelV. Chim. Acta 2001, 84, 87–
105.
(23) Hill, C. L.; Whitesides, G. M. J. Am. Chem. Soc. 1974, 96, 870–876.
6892 J. Org. Chem. Vol. 73, No. 17, 2008

SCHEME 2. Synthesis of the 6′(R)-Azidomethyl
LNA-Thymidine Monomer and a Double-Headed
Nucleosidea
from 6. The potential of the 6′(R)-position for the design of
functional LNA and subsequent nanostructures will be illustrated
in the near future. Nevertheless, simple modeling starting from
LNA-modified duplexes1,28 indicates the present pro-R position
of the 6′-carbon to be the most favorable position of the two
pointing into the minor groove. Hence, we consider the present
method for stereoselective preparation of 6′(R)-branched LNA
as extremely convenient, and the application of the key
compounds 6, 11 and 12 for the preparation of functionalized
LNA derivatives is in progress.
a Reagents and conditions: (a) MsCl, pyridine, DCM (90%); (b) NaN3,
DMF (57%); (c) Zn, AcOH (31%); (d) N1-propragylthymine, Na ascorbate,
CuSO4 ·5H2O, tBuOH, H2O (75%); (e) H2, Pd/C, EtOH (67%).
Experimental Section
Synthesisof(1R,3R,4R,6S,7S)-7-benzyloxy-1-benzyloxymethyl-
6-bromomercurimethyl-3-(thymin-1-yl)-2,5-dioxabicyclo[2.2.1]-
heptane (7). To a solution of the olefin 4 (2.82 g, 6.1 mmol) in
THF (150 mL) was added mercury(II) trifluoroacetate (3.90 g, 9.11
mmol), and the mixture was stirred for 12 h. The mixture was
concentrated under reduced pressure, and the residue was dissolved
in DCM (150 mL). A 10% aqueous solution of NaBr (50 mL) was
added, and the mixture was stirred vigorously for 30 min. The two
phases were separated, and the organic phase was treated with
another 10% aqueous solution of NaBr (50 mL). After separation,
the organic phase was washed with water (3 × 50 mL), dried
(Na2SO4), and concentrated under reduced pressure to give the
product (4.45 g, 98%) as a white foam: Rf 0.42 (EtOAc in hexane,
FIGURE 2. N-Type conformation and mutual NOE contacts indicated
in the 6′-methylenebranched LNA derivatives (based on NMR experi-
ments on 11 and 12, R ) N3 or NH2).
Both observations proved the 6′(R)-configuration as demon-
strated in Figure 2.
Obviously, both the hydroxy group of 6 and the amine of 12
can be used in nucleophilic derivatizations. Nevertheless, we
decided to apply the azide functionality of 11 in combination
with terminal alkynes in Huisgen-type [3 + 2] cycloadditions,
which can be performed with complete regioselectivity, giving
only 1,4-disubstituted triazoles by a Cu(I) catalysis.24 Recently,
this reaction type has been intensively applied as a result of the
concept of click chemistry introduced by Sharpless and co-
workers.25 The reaction is very robust, it tolerates almost all
functional groups, it can be performed in aqueous solutions,
and it displays high atom economy giving no side products. In
line of our ongoing program on double-headed nucleosides,9,11
we decided to use the present azide for the decoration of LNA
with an additional nucleobase. N1-Propargylthymine26 and
compound 11 were reacted following a standard protocol with
microwave heating24,27 to give efficiently the double-headed
nucleoside 13. Global deprotection by hydrogenolysis afforded
compound 14. This compound is currently being studied as a
building block in nucleic acid secondary structures such as our
recently investigated three-way junctions or DNA-zipper motifs.11
In summary, we have proved the applicability of mercury(II)
trifluoroacetate for forming a 6′-hydroxymethyl modified LNA
monomer that is otherwise obtainable only in lower yields and
a higher number of steps.8 Hence, compound 6 was obtained
in approximately 22% overall yield over the 13 steps from
1,2;5,6-di-O-isopropylidene-R-D-allofuranose, whereas the dif-
ferently protected analogue by Swayze and Seth was obtained
in 19% yield as an epimeric mixture over 17 steps.8 Furthermore,
the utility of compound 6 for derivatization was illustrated by
the preparation of first the azide 11 and then of the double-
headed nucleoside 14 in 26% overall yield over the four steps
1
1:1 v/v); H NMR (300 MHz, CDCl3) δ 9.10 (m, 1H, NH), 7.49
(s, 1H, H-6), 7.31-7.18 (m, 10H, Ph), 5.57 (s, 1H, H-1′), 4.61-4.55
(m, 6H, H-2′, 2 × CH2Ph, H-6′), 3.99 (s, 1H, H-3′), 3.68 (s, 2H,
2 × H-5′), 2.25 (m, 1H, H-7′), 1.91 (m, 1H, H-7′), 1.55 (s, 3H,
CH3), 13C NMR (75 MHz, CDCl3) δ 163.8 (C-4), 149.9 (C-2),
137.3, 136.8 (Ph), 134.7 (C-6), 128.8, 128.8, 128.7, 128.7, 128.6,
128.3, 128.0, 127.9 (Ph), 110.2 (C-5), 87.3, 86.7 (C-1′, C-4′), 79.8,
78.3, 77.5 (C-2′, C-3′, C-6′), 73.95, 72.5 (2 × CH2Ph), 64.7 (C-
5′), 31.9 (C-7′), 12.4 (CH3); HRMALDI MS m/z calcd for
C26H27HgBrN2O6 [M + Na]+ 767.0651, found 767.0618.
Synthesis of (1S,3R,4R,6R,7S)-7-Benzyloxy-1-benzyloxymethyl-
6-(2,2,6,6-tetramethylpiperidin-1-yl)oxymethyl-3-(thymin-1-yl)-2,5-
dioxabicyclo[2.2.1]heptane (8). To a stirred solution of the mercuric
derivative 7 (2.0 g, 2.68 mmol) in anhydrous DMF (13 mL) was
added TEMPO (3.15 g, 20.2 mmol). The reaction mixture was
stirred at 0 °C, and a solution of sodium boron hydride (477 mg,
12.6 mmol) and TEMPO (3.15 g, 20.2 mmol) in anhydrous DMF
(13 mL) was added dropwise. The mixture was stirred at 0 °C for
1 h and filtered through celite with precaution to remove the
mercury particles. The sinter was rinsed with dichloromethane (150
mL). The combined filtrates were concentrated under reduced
pressure and coevaporated with xylene. The residue was purified
by column chromatography (20-50% EtOAc in hexane) to afford
the product (2.50 g, 77%) as a white foam: Rf 0.46 (EtOAc in
hexane, 1:1 v/v); IR (KBr,νmax cm-1) 3433.9, 3187.5, 3065.9,
2929.4, 1693.9, 1455.14, 1361.2, 1269.7, 1103.4, 1055.9; 1H NMR
(300 MHz, CDCl3) δ 7.59 (s, 1H, H-6), 7.36-7.25 (m, 10H, Ph),
5.54 (s, 1H, H-1′), 4.69-4.55 (m, 4H, CH2Ph), 4.36 (s, 1H, H-2′),
4.31 (m, 1H, H-6′), 4.13-4.08 (m, 2H, H-3′, H-5′), 3.99 (m, 2H,
H-7′), 3.79 (d, 1H, J ) 11.4 Hz, H-5′), 1.60 (s, 3H, CH3), 1.43 (br
s, 6H, 3 × CH2), 1.13 (br s, 6H, 2 × CH3), 1.00 (br s, 6H, 2 ×
CH3); 13C NMR (75 MHz, CDCl3) δ 163.7 (C-4), 149.7 (C-2),
137.8, 137.0 (Ph), 135.1 (C-6), 128.7, 128.6, 128.3, 128.1, 128.0,
127.7 (Ph), 110.2 (C-5), 87.9 (C-4′), 86.6 (C-1′), 77.6 (C-3′), 76.7
(C-6′), 76.5 (C-2′), 74.0 (CH2Ph), 73.7 (C-7′), 72.6 (CH2Ph), 64.8
(C-5′), 60.1, 60.0 (C(CH3)2), 39.8, 39.7 (CH2C(CH3)2), 33.4, 32.8
(C(CH3)2), 20.5, 20.1 (C(CH3)2), 17.2 (CH2CH2CH2), 12.4 (CH3);
HRMALDI MS m/z calcd for C35H45N3O7 [M + Na]+ 642.3150,
(24) (a) Tornøe, C. W.; Christensen, C.; Meldal, M. J. Org. Chem. 2002,
67, 3057–3064. (b) Rostovtsev, V. V.; Green, L. G.; Fokin, V. V.; Sharpless,
K. B. Angew. Chem., Int. Ed. 2002, 41, 2596–2599.
(25) Kolb, H. C.; Finn, M. G.; Sharpless, K. B. Angew. Chem., Int. Ed. 2001,
40, 2004–2021.
(26) Lindsell, W. E.; Murray, C.; Preston, P. N.; Woodman, T. A. J.
Tetrahedron 2000, 56, 1233–1245.
(27) Kocˇalka, P.; Andersen, N. K.; Jensen, F.; Nielsen, P. ChemBioChem
2007, 8, 2106–2116.
(28) Nielsen, K. E.; Singh, S. K.; Wengel, J.; Jacobsen, J. P. Bioconjugate
Chem. 2001, 11, 228–238.
J. Org. Chem. Vol. 73, No. 17, 2008 6893

found 642.3120. Anal. for C35H45N3O7 ·0.25 H2O: found C, 67.35;
H, 7.33; N, 6.80; requires C, 67.34; H, 7.35; N, 6.73.
137.4, 136.9 (Ph), 134.8 (C-6), 128.8, 128.7, 128.7, 128.6, 128.4,
128.2, 127.9, 127.8 (Ph), 110.45 (C-5), 87.4 (C-4′), 86.8 (C-1′),
81.0 (C-6′), 77.8 (C-3′), 76.6 (C-2′), 73.9, 72.4 (2 × CH2Ph), 65.0
(C-5′), 60.5 (C-7′), 12.4 (CH3); HRMALDI MS m/z calcd for
C26H28N2O7 [M + Na]+ 503.1789, found 503.1782.
Synthesis of (1S,3R,4R,6R,7S)-7-Benzyloxy-1-benzyloxymethyl-
3-(thymin-1-yl)-6-hydroxymethyl-2,5-dioxabicyclo[2.2.1]heptane (6).
To a stirred solution of the TEMPO derivative 8 (1.8 g, 2.97 mmol)
in a mixture of AcOH, H2O, and THF (95 mL, 3:1:1 v/v/v) was
added (10 µm) zinc dust (1.67 g, 26.4 mmol) in small portions.
The mixture was stirred at 65 °C for 1.5 h and then filtered through
a filter (0.45 µm) on a syringe adapter. The filter was washed with
THF, and the combined filtrates were concentrated under reduced
pressure. The residue was dissolved in EtOAc (250 mL), and the
solution was washed with water (50 mL), a saturated aqueous
solution of NaHCO3 (50 mL), and brine (50 mL), dried (Na2SO4)
and concentrated under reduced pressure. The residue was purified
by column chromatography (20-60% EtOAc in hexane) to afford
the product (1.18 g, 83%) as a white foam; Rf 0.20 (EtOAc in
hexane, 7:3 v/v); IR (KBr, νmax cm-1) 3437.9, 3183.4, 3065.1,
2926.6, 1690.1, 1455.26, 1365.9, 1271.1, 1105.4, 1057.6; 1H NMR
(300 MHz, CDCl3) δ 8.7 (br s, 1H, NH), 7.45 (s, 1H, H-6),
7.36-7.25 (m, 10H, Ph), 5.56 (s, 1H, H-1′), 4.68-4.52 (m, 4H,
CH2Ph), 4.45 (s, 1H, H-2′), 4.32 (m, 1H, H-6′), 4.04 (s, 1H, H-3′),
3.98-3.82 (m, 4H, H-5′, H-7′), 2.25 (m, 1H, OH), 1.62 (s, 3H,
CH3); 13C NMR (75 MHz, CDCl3) δ 164.1 (C-4), 150.0 (C-2),
Acknowledgment. The project was supported by The Danish
Research Agency′s Programme for Young Researchers, Nucleic
Acid Center and The Danish National Research Foundation,
NAC-DRUG (Nucleic Acid Based Drug Design Training
Center) in the Sixth Framework Programme Marie Curie Host
Fellowships for Early Stage Research Training under contract
no. MEST-CT-2004-504018, and Møllerens Fond. The Nucleic
Acid Center is funded by the Danish National Research
Foundation for studies on nucleic acid chemical biology.
Supporting Information Available: General experimental
section including additional experimental procedures and copies
of NMR spectra. This material is available free of charge via
the Internet at http://pubs.acs.org.
JO801081T
6894 J. Org. Chem. Vol. 73, No. 17, 2008