Synthesis and incorporation of a simple acyclic furan containing phosphoramidite

Article abstract of DOI:10.1080/15257770701533917

A novel furan containing phosphoramidite was synthesized and incorporated into model oligonucleotides. This glycol nucleic acid based building block contains a furan unit substituting the natural base, and can be used for post synthetic oligonucleotide mo

Full text of DOI:10.1080/15257770701533917

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Synthesis and Incorporation of a
Simple Acyclic Furan Containing
Phosphoramidite
K. Stevens a & A. Madder a
a Laboratory for Organic and Biomimetic Synthesis, Department of
Organic Chemistry , Ghent University , Ghent, Belgium
Published online: 05 Dec 2007.
To cite this article: K. Stevens & A. Madder (2007) Synthesis and Incorporation of a Simple Acyclic
Furan Containing Phosphoramidite, Nucleosides, Nucleotides and Nucleic Acids, 26:10-12, 1359-1362,
DOI: 10.1080/15257770701533917
To link to this article: http://dx.doi.org/10.1080/15257770701533917
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Nucleosides, Nucleotides, and Nucleic Acids, 26:1359–1362, 2007
Copyright ꢀ Taylor & Francis Group, LLC
C
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770701533917
SYNTHESIS AND INCORPORATION OF A SIMPLE ACYCLIC FURAN
CONTAINING PHOSPHORAMIDITE
K. Stevens and A. Madder
Laboratory for Organic and Biomimetic Synthesis,
2
Department of Organic Chemistry, Ghent University, Ghent, Belgium
A novel furan containing phosphoramidite was synthesized and incorporated into model
2
oligonucleotides. This glycol nucleic acid based building block contains a furan unit substitut-
ing the natural base, and can be used for post synthetic oligonucleotide modifications by orthogonal
chemistries such as Schiff base formation after in situ oxidation or Diels-Alder cycloadditions.
Keywords Phosphoramidite; furan; cross-linking; modified oligonucleotide
INTRODUCTION
Interstrand cross-links have been extensively studied in the past since
they are among the most toxic DNA damage.[1] They are lesions that can be
formed by various anti tumor drugs, such as cis-diaminedichloroplatinum
(II) and mitomycin C.[2] For the study of DNA-repair, sufficient amounts of
uniformly cross-linked DNA are needed. Since isolation of cross-linked DNA
from natural sources is difficult, various interstrand cross-linked duplexes
were chemically synthesized.[3–6]
Recently our group developed a new method for DNA interstrand
cross-linking based on the use of a furan moiety as masked reactive
functionality.[7] Through selective oxidation the furan can be transformed
into a 4-oxo-enal. This unsaturated aldehyde can react with various nucle-
ophiles, resulting in interstrand cross-links when positioned inside a DNA-
duplex (Figure 1). Furan containing oligonucleotides further find applica-
tions for the conjugation of fluorescent labels[8,9] and surface enhanced
resonance Raman scattering-probes[10] via Diels-Alder strategies and in
the development of new orthogonal base pairs.[11]
K. Stevens thanks the University of Ghent for a position as research fellow (BOF 01115203). Support
from the FWO-Vlaanderen (KAN 1.5.146.06 and KAN 1.5.186.03) is gratefully acknowledged.
Address correspondence to A. Madder, Laboratory for Organic and Biomimetic Synthesis, Depart-
ment of Organic Chemistry, Ghent University, Krijgslaan 281 (S4); B 9000 Ghent, Belgium. E-mail:
annemieke.madder@ugent.be
1359

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K. Stevens and A. Madder
FIGURE 1 Furan based cross-link strategy.
Previously, we concentrated on a nucleoside building block containing a
furan moiety linked via an amide bond to the 2ꢁ position of the ribose. New
furan containing building blocks should be easily synthetically accessible
and display a high atom economy. Inspired by the glycol nucleic acid[12]
(GNA) building blocks, developed by the Meggers group, we here present a
simplified furan phosphoramidite accessible via a very short and convenient
synthesis route (Figure 2). In order to ensure accessibility of the oxidized
furan moiety by the complementary bases a two carbon linker between the
formal C3ꢁ and the furan was introduced.
RESULTS AND DISCUSSION
The synthesis is outlined in Scheme 1. The first step involves tosylation of
the commercially available S-(+)-3,4-O-isopropylidenedioxybutan-1-ol. Di-
rect substitution of the tosylate with furyllithium failed to give conversion
to the desired product. Therefore, upon conversion to a bromide smooth
substitution with furyllithium yields compound 3 in satisfactory yields. De-
protection of the acetonide was performed at 0◦C since cleavage of the fu-
ran ring can occur at higher temperatures under these acidic conditions.
In order to avoid formation of the double dimethoxytritylated compound,
also this protection reaction was performed at low temperature. In this way
dimethoxytritylation of the secondary alcohol can be minimised. Although
FIGURE 2 Structure of glycol nucleic acid building block S-GNA and the proposed furan derivative.

Simple Acyclic Furan
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SCHEME 1 Synthesis a) pTsCl (1.15 eq.), pyridine, 0◦C, 2.25 hours, 91%; b) LiBr (5 eq.), DMF, 60◦C,
1 hour; c) Furan, BuLi, THF, −78–0◦C, 16 hours, 62% over two steps; d) 4M HCl, THF, 3 hours, 0◦C,
89%; e) DMTCl (1 eq.), pyridine, 0◦C, 1.25 hours, 66%; f) (iPr2N)(NCCH2CH2O)PCl (2.5 eq.), DIPEA,
CH2Cl2, 1.5 hours, rt, 79%.
no completely selective reaction takes place, separation of mono- and
ditritylated compounds was achieved by flash chromatography yielding the
desired monoprotected 5 in good yield. The correct positioning of the DMT-
group was confirmed by NMR. The phosphoramidite could be easily gener-
ated under standard conditions which yielded 6 as a colorless oil.
The building block was then incorporated into oligonucleotide strands
on an Applied Biosystems 394 DNA synthesizer. For a 1 µmol synthesis,
0.4 ml of a 50 mM solution of 6 in acetonitrile was dried over molecular
˚
sieves (3 A) for 1 hour. A 1 M 4,5-dicyanoimidazole solution in acetonitrile
was used as activator.[13] The coupling of the modification was performed
manually with a prolonged coupling time of 10 minutes. The coupling yields
under these conditions were estimated to be more than 99%. The modi-
fied phosphoramidite was incorporated into four different sequences (see
Table 1) varying each time the nearest neighboring bases. It was thus con-
firmed that coupling to different bases did not result in observed differ-
ences in coupling yield. These oligonucleotide sequences will be further
used to explore the specificity and sequence context dependence of cross-
linking. All masses of the synthesized oligonucleotides were confirmed by
MALDI-TOF.
In conclusion we have synthesized a novel furan containing phospho-
ramidite which is easily accessible via a short and convenient route and
TABLE 1 Synthesized oligonucleotide sequences where X represents the modified
residue. Comparison of the calculated masses with the masses obtained with MALDI-TOF
Sequence
Calculated mass (Da)
Measured [M+1] (m/z)
5ꢁ-CTGACGGXGTGC-3ꢁ
5ꢁ-CTGACGTXTTGC-3ꢁ
5ꢁ-CTGACGCXCTGC-3ꢁ
5ꢁ-CTGACGAXATGC-3ꢁ
3589.63
3539.62
3509.54
3557.56
3589.44
3540.51
3509.86
3557.53

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K. Stevens and A. Madder
which can be efficiently incorporated into oligonucleotide strands. The cur-
rent building block allows for internal introduction of a furan moiety into an
oligonucleotide. Selective reactions with furan, such as Diels-Alder cycload-
dition or oxidative transformation into a 4-oxo-enal, allow for versatile and
orthogonal postsynthetic chemistries. Studies on the influence of the newly
incorporated building block on the duplex stability and cross-link potential
are currently in progress.
REFERENCES
1. Dronkert, M.L.G.; Kanaar, R. Mutation Research 2001, 486, 217–247.
2. Rajski, S.R.; Williams, R.M. Chem. Rev. 1998, 98, 2723–2795.
3. Lee, B.L.; Murakami, A.; Blake, K.R.; Lin, S.B.; Miller, P.S. Biochemistry 1988, 27(9), 3197–3203.
4. Kozekov, I.D.; Nechev, L.V.; Moseley, M.S.; Harris, C.M.; Rizzo, C.J.; Stone, M.P.; Harris, T.M. J. Am.
Chem. Soc. 2003, 125, 50–61.
5. Noronha, A.M.; Noll, D.M.; Miller, P.S. Nucleosides, Nucleotides Nucleic Acids 2001, 20(4–7), 1303–1307.
6. Kirchner, J.J.; Sigurdsson, S.T.; Hopkins, P.B. J. Am. Chem. Soc. 1992, 114, 4021–4027.
7. Halila, S.; Velasco, T.; De Clercq, P.; Madder, A. Chem. Comm. 2005, 7, 336–338.
8. Graham, D.; Grondin, A.; McHugh, C.; Fruk, L.; Smith, W.E. Tetrahedron Lett. 2002, 43, 4785–4788.
9. Anderson, E.; Picken, D. Nucleosides Nucleotides Nucleic Acids 2005, 24, 761–765.
10. Graham, D.; Fruk, L.; Smith, W.E. Analyst 2003, 128, 692–699.
11. Berger, M.; Luzzi, S.D.; Henry, A.A.; Romesberg, F.E. J. Am. Chem. Soc. 2002, 124, 1222–1226.
12. Zhang, L.; Peritz, A.; Meggers, E. J. Am. Chem. Soc. 2005, 127, 4174–4175.
13. Vargeese, C.; Carter, J.; Yegge, J.; Krivjansky, S.; Settle, A.; Kropp, E.; Peterson, K.; Pieken, W. Nucl.
Acids Res. 1998, 26(4), 1046–1050.