The use of gaseous ammonia for the deprotection and cleavage steps during the solid-phase synthesis of oligonucleotides, and analogs

Bioorganic and Medicinal Chemistry Letters

Volumn 7, Issue 11, 1997, Pages 1443-1448

  Iyer, Radhakrishnan P ^a   Yu, Dong ^a   Xie, Jin ^a   Zhou, Wen ^a   Agrawal, Sudhir ^a  

^a HYBRIDON INC   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

AMMONIA; OLIGONUCLEOTIDE;

ARTICLE; DRUG SYNTHESIS; METHODOLOGY; REACTION ANALYSIS;

EID: 0030946199     PISSN: 0960894X     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0960-894X(97)00236-9     Document Type: Article

References (25)

1. (a) Beaucage, S. L.; Caruthers, M. H. Tetrahedron Lett. 1981, 22, 1859;
2. (b) Sinha, N. D.; Biernat, J. P.; McManus, J.; Koster, H. Nucl. Acids Res. 1984, 12, 4539;
3. (c) Caruthers, M. H. Science 1985, 250, 281.
4. (a) Pon, R. T.; Buck, G. A.; Niece, R. L.; Robertson, M.; Smith, A. J.; Spicer, E. Biotechniques, 1994, 17, 526;
5. (b) Agrawal, S.; Iyer, R. P. Curr. Op. Biotech. 1995, 6, 12;
6. (c) Antisense Therapeutics, Agrawal, S., Ed.; Humana: Totowa, NJ., 1996, pp 1-11;
7. (a) For a review, see: Beaucage, S. L.; Iyer, R. P. Tetrahedron 1992, 48, 2223;
8. (b) For recent reports on improved purification protocols for PS oligonucleotides, see: (i) Green, A. P. ; Burzynski, J. ; Helveston, N. M.; Prior, G. M.; Wunner, W. H.; Thompson, J. A. Biotechniques, 1995, 19, 836, and references therein; Gerstner, J. A.; Pedroso, P.; Morris, J.; Bergot, B. J. Nucl. Acids Res. 1995, 23, 2292, and references therein.
9. (b) For recent reports on improved purification protocols for PS oligonucleotides, see: (i) Green, A. P. ; Burzynski, J. ; Helveston, N. M.; Prior, G. M.; Wunner, W. H.; Thompson, J. A. Biotechniques, 1995, 19, 836, and references therein; Gerstner, J. A.; Pedroso, P.; Morris, J.; Bergot, B. J. Nucl. Acids Res. 1995, 23, 2292, and references therein.
10. (a) Devlin, T.; Iyer, R. P.; Johnson, S.; Agrawal, S. Bioorg. Med. Chem. Lett. 1996, 22, 2663;
11. (b) For other reports on these analogs, see: Peyrottes, S.; Vasseur, J.-J.; Imbach, J.-L.; Rayner, B. Nucl. Acids Res. 1996, 24, 1841, and references therein.
12. Boal, J. H.; Wilk, A.; Harindranath, N.; Max, E. E.; Kempe, T.; Beaucage, S. L. Nucl. Acids Res. 1996, 24, 3115.
13. (a) It is expected that by operating under pressures higher than that used in the present study, significant reduction in the time of deprotection, and cleavage can be achieved. Work is ongoing in this regard;
14. (b) Kodra, J. T.; Kehler, J.; Dahl, O. Nucl. Acids Res. 1995, 23, 3349;
15. (c) Following deprotection, the reaction vessel was depressurized in a well-ventilated hood.
16. The Parr apparatus was purchased from Ace Glass Inc., Vineland, New Jersey.
17. For conditions of analytical, and preparative reversed-phase HPLC analysis, see: (a) Iyer, R. P.; Yu, D.; Agrawal, S. Bioorg. Med. Chem. Lett. 1994, 4, 2471;
18. (b) Iyer, R. P.; Yu, D.; Agrawal, S. Bioorg. Chem. 1995, 23, 1.
19. (a) DNA synthesis was performed on an EXPEDITE synthesizer (PerSeptive Biosystems);
20. (b) For details see: Iyer, R. P.; Yu, D.; Habus, I.; Ho, N.-H.; Johnson, S.; Devlin, T.; Jiang, Z.; Zhou, W.; Xie, J.; Agrawal, S. Tetrahedron 1997, 55, 2731.
21. (a) It is reported that some detritylationof the DMT-on full-length oligonucleotide might occur during the processing step that follows aqueous ammonia deprotection; the resulting DMT-off material being bereft of the "hydrophobic handle" might co-elute with the failure sequences, and not recovered. Therefore, certain protocols recommend the addition of triethylamine to the solution of the digonucleotide in ammonium hydroxide, prior to concentration, to minimize this premature detritylation, see: (i) Warren, W. J.; Vella, G. In Protocols for Oligonucleotide Conjugates, Agrawal, S., Ed.; Humana: Totowa, NJ., 1994, pp 233-264;
22. Applied Biosystems User Bulletin, No. 50, August 1988;
23. (b) This ratio was ca. 3 to 5% higher when such a comparative study was done using PNT nucleoside monomers in each case.
24. Several factors may contribute to the reduced N - 1 content; For a discussion, see reference 9b.
25. CGE analysis was done on a Beckman P/ace 2200 instrument operating at 14.1 Kv; before CGE, the samples were desalted using a SEP-PAK cartridge.