function that is isostructural and isoelectrostatic to a natural
backbone phosphodiester function. This modification is
expected to block the â-elimination reaction at abasic sites,
thus inhibiting strand scission.
3′-Deoxy-3′-hydroxymethylene nucleosides and corre-
sponding phosphonates were synthesized before. Their
syntheses typically started from the natural nucleosides using
different C-1 synthons.2 DNA oligonucleotides containing
phosphonate backbone linkages were also prepared before
in the context of antisense research, and it was shown that
they slightly enhance duplex stability,3 indicating that the
mutation of a 3′-oxygen into a methylene group does not
interfere with double helix formation.
during deprotection and detachment from the solid support
of the oligonucleotide. The synthesis of building block 7 is
outlined in Scheme 1.
While the synthesis up to iodide 6 proceeded smoothly
and in good yields, the following Arbusov reaction proved
to be difficult. Reaction of 6 with in situ prepared bis-
trimethylsilylphosphonite (BTSP) led to only ca. 20% of the
desired building block 7 after aqueous workup, even after
considerable efforts of optimization. A major side reaction
observed was the (known) reduction by BTSP to the
corresponding 3′-methyl derivative.5 Changing the leaving
group to a tosylate inhibited the reduction pathway but
surprisingly did not improve the yield. Attempts to use the
more active triflate leaving group failed due to the instability
of the corresponding compound. To test whether a TMS-
based Lewis acid originating from the TMS phosphinate
intermediate of 7 was responsible for the low yield, we also
subjected lactone 5 to BTSP treatment. But again the
corresponding Arbusov product could only be isolated in ca.
25% yield, indicating that Lewis acid-catalyzed elimination
of the anomeric acetate group is not a significant side reaction
responsible for the low yields of 7. Despite the difficulties
in the last synthetic step, 7 could be produced in sufficient
quantities for the following synthetic and biophysical studies.
Here, we report on the successful synthesis of oligode-
oxynucleotides containing the 3′-methylene abasic site
analogue X (Figure 1) and the characterization of its
functional properties and its chemical stability.
Scheme 1. Synthesis of Building Block 7
In a series of model experiments using 3′-O-TBDMS-
protected thymidine and 7 we first tested a variety of
carbodiimide-, uronium-, and phosphonium-based coupling
reagents and were surprised to find that, contrary to the
reported high coupling efficiencies in the ribo series, building
block 7 reacted not at all under standard coupling conditions
(30 min, rt, 20-fold molar excess). It thus seems that a
missing electronegative substituent at the remote 2′-position
decelerates the condensation reaction for a yet unknown
reason. A way out was finally found by adding a nucleophilic
catalyst to the coupling mixture as was recently reported for
the synthesis of boranophosphates.6 We found that the
combination of BOP-Cl and 3-nitro-1,2,4-triazole was per-
fectly suited for coupling the H-phosphinate intermediate
(Figure 2), leading to complete conversion within 30 min.
The related NEP-Cl was under these conditions less active
and needed 24 h for complete conversion.
We planned to use the H-phosphinate 7 (Scheme 1) as a
building block for X, reasoning that it could be directly used
in the automated synthesis of oligodeoxynucleotides and that
it would be compatible with standard phosphoramidite
chemistry. For the elaboration of the C-C bond between
the methylene unit and the deoxyribose ring we intended to
utilize the photoinduced radical conjugate addition of
methanol, previously developed by Mann.4 As a protecting
group for the anomeric center we chose an acetyl group
which would liberate the hemiacetal function concomitantly
Having optimized the coupling conditions in solution, we
next approached the solid-phase oligonucleotide synthesis.
Within the amidite protocol, a separate coupling and oxida-
tion step for 7 had to be accommodated. Coupling was
effected with BOP-Cl and 3-nitro-1,2,4-triazole as activator
while oxidation was performed using iodine (200 mM) under
basic (1 M NEt3) conditions for 5 min.7 The detailed protocol
is described in the Supporting Information. With this
protocol, we prepared the two oligonucleotides 8 and 9
(Table 1). Coupling efficiencies for the incorporation of 7
as determined from the trityl assay were typically >95%.
Oligonucleotides were detached from solid support and
deprotected under standard conditions (NH3 concd, 55 °C,
(2) (a) Sanghvi, Y. S.; Bharadwaj, R.; Debart, F.; De Mesmaeker, A.
Synthesis 1994, 1163. (b) Collingwood, S. P.; Douglas, M. E.; Natt, F.;
Pieles, U. Phosphorus, Sulfur Silicon Relat. Elem. 1999, 144-146, 645.
(c) An, H.; Wang, T.; Dan Cook, P. Tetrahedron Lett. 2000, 41, 7813. (d)
Winquist, A.; Stro¨mberg, R. Eur. J. Org. Chem. 2001, 4305.
(3) An, H.; Wang, T.; Maier, M. A.; Manoharan, M.; Ross, B. S.; Dan
Cook, P. J. Org. Chem. 2001, 66, 2789.
(5) Winqvist, A.; Stro¨mberg, R. Eur. J. Org. Chem. 2002, 1509.
(6) Shimizu, M.; Wada, T.; Oka, N.; Saigo, K. J. Org. Chem. 2004, 69,
5261.
(4) (a) Mann, J.; Weymouth-Wilson, A. Synlett 1992, 67. (b) Gould, J.
H. M.; Mann, J. Nucleosides, Nucleotides 1997, 16, 193.
(7) Winqvist, A.; Stro¨mberg, R. Eur. J. Org. Chem. 2002, 3140.
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Org. Lett., Vol. 7, No. 23, 2005