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Synlett 2002(5): 0763-0766
DOI: 10.1055/s-2002-25332
LETTER
© Georg Thieme Verlag Stuttgart · New York

Nucleic Acid Containing 3′-C-P-N-5′ Ethyl Phosphonamidate Ester and 2′-Methoxy Modifications in Combination; Synthesis and Hybridisation
Properties

Robin A. Fairhurst*, Stephen P. Collingwood, David Lambert, Elke Wissler
Novartis Pharmaceuticals, Central Research Laboratories, Hulley Road, Macclesfield, Cheshire, SK10 2NX, UK
Fax: +44(1403)323307; e-Mail: robin.fairhurst@pharma.novartis.com;
Further Information

Publication History

Received 29 January 2002
Publication Date:
07 February 2007 (online)

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Abstract

The preparation of thymidine-thymidine and thymidine-5-methylcytidine dinucleosides containing a 3′-C-P-N-5′ ethyl phosphonamidate ester linkage, with defined phosphorus stereochemistry, in combination with a 2′-methoxy substituent in the lower sugar residue, is described. Incorporation of these dinucleosides into DNA oligonucleotides and the effect upon duplex stability with complimentary RNA is reported.

Key words

antisense - modified oligonucleotides - nucleosides - phosphorus - stereoselective

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1

New address: Novartis Horsham Research Centre, Wimblehurst Road, Horsham, West Sussex, RH12 5AB, UK.

11

Reaction conditions were as described in ref. [5] Flash column chromatography was performed using Merck Silica Gel 60 (0.040-0.063 mm). NMR spectra were recorded with a Brucker AC400 instrument. Key distinguishing 1H resonances for each diastereoisomer are assigned. 31P NMR shifts are given as ppm values relative to phosphoric acid. Mass spectroscopy was carried out using a Fisons Instruments VG Platform II spectrometer. A reaction carried out on a 3.11 mmol scale gave;
5: White amorphous foam; 1.07 g; 31P NMR (CDCl3, 162 MHz): δ = 32.52 ppm; 1H NMR (CDCl3, 400 MHz): δ = 9.67 (br s, 1 H), 9.52 (s, br, 1 H), 7.69-7.61 (m, 4 H), 7.45-7.34 (m, 7 H), 7.13 (s, 1 H), 6.10-6.02 (m, 1 H, H1′ upper sugar), 5.50 (d, J = 2 Hz, 1 H, H1′ lower sugar), 4.26-3.64 (m, 8 H), 3.50 (s, 3 H), 3.36-3.17 (m, 3 H), 2.74-2.61 (m, 1 H), 2.51-2.42 (m, 1 H), 2.32-2.20 (m, 1 H), 2.04-1.82 (m, 2 H), 1.84 (s, 3 H), 1.74-1.59 (m, 1 H), 1.58 (s, 3 H), 1.27 (t, J = 7 Hz, 3 H), 1.04 (s, 9 H). MS (ES+): m/z (%) = 840(27) [M + H], 862(100) [M + Na].
6: White amorphous foam; 0.94 g; 31P NMR (CDCl3, 162 MHz): δ = 32.81 ppm; 1H NMR (CDCl3, 400 MHz): δ = 9.38 (br s, 1 H), 9.17 (s, br, 1 H), 7.70-7.62 (m, 4 H), 7.44-7.34 (m, 7 H), 7.13 (s, 1 H), 6.14-6.06 (m, 1 H, H1′ upper sugar), 5.55 (d, J = 2 Hz, 1 H, H1′ lower sugar), 4.24-3.64 (m, 8 H), 3.51 (s, 3 H), 3.50-3.18 (m, 3 H), 2.80-2.68 (m, 1 H), 2.50-2.39 (m, 1 H), 2.34-2.23 (m, 1 H), 2.01-1.84 (m, 1 H), 1.84 (s, 3 H), 1.77-1.58 (m, 2 H), 1.61 (s, 3 H), 1.20-1.10 (m, 3 H), 1.04 (s, 9 H). MS (ES+): m/z (%) = 840(8) [M + H], 862(100) [M + Na].

13

Reaction conditions were as described in ref. [5] A reaction carried out on a 1.46 mmol scale gave;
13: White amorphous foam; 1.54 g; 31P NMR (CDCl3, 162 MHz): δ = 32.70 ppm; 1H NMR (CDCl3, 400 MHz): δ = 8.62 (s, 1 H), 8.15 (d, 2 H, J = 7 Hz), 7.58-7.44 (m, 8 H), 7.35-7.31 (m, 1 H), 7.30-7.17 (m, 16 H), 7.08 (s, 1 H), 5.96-5.90 (m, 1 H, H1′ upper sugar), 5.42 (d, 1 H, J = 2 Hz, H1′ lower sugar), 3.97-3.89 (m, 1 H), 3.86-3.77 (m, 2 H), 3.69-3.53 (m, 4 H), 3.10 (s, 3 H), 2.98-2.81 (m, 2 H), 2.76-2.63 (m, 1 H), 2.59-2.48 (m, 1 H), 2.29-2.20 (m, 1 H), 2.03-1.94 (m, 1 H), 1.91 (s, 3 H), 1.76-1.60 (m, 1 H), 1.44 (s, 3 H), 1.38-1.14 (m, 1 H), 1.07-0.99 (m, 3 H), 0.94 (s, 9 H), 0.91 (s, 9 H). Minor (S P)-diastereoisomer; 31P NMR (CDCl3, 162 MHz): δ = 31.71 ppm; 1H NMR (CDCl3, 400 MHz): key distinguishing resonances δ = 5.90-5.83 (m, 1 H, H1′ upper sugar), 5.29 (d, 1 H, J = 2 Hz, H1′ lower sugar).