A solid supported reagent for internucleoside H-phosphonate linkage formation

Article abstract of DOI:10.1081/NCN-200059257

A fast and convenient procedure for synthesis of dinucleoside H-phosphonates is obtained through use of the novel polystyrene supported 5-carboxy-5-methyl-2-oxo-2-chloro-1,3,2-dioxaphosphorinane reagent. Virtually quantitative H-phosphonate condensations

Full text of DOI:10.1081/NCN-200059257

This article was downloaded by: [University of Colorado at Boulder Libraries]
On: 29 December 2014, At: 05:06
Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,
37-41 Mortimer Street, London W1T 3JH, UK
Nucleosides, Nucleotides and Nucleic Acids
Publication details, including instructions for authors and subscription information:
http://www.tandfonline.com/loi/lncn20
A SOLID SUPPORTED REAGENT FOR INTERNUCLEOSIDE
H-PHOSPHONATE LINKAGE FORMATION
Nikhil Mohe a , Petri Heinonen a , Yogesh S. Sanghvi b & Roger Strömberg a
a Division of Organic and Bioorganic Chemistry, MBB, Scheele Laboratory , Karolinska
Institutet , Stockholm, Sweden
b Isis Pharmaceuticals, Inc. , Carlsbad, California, USA
Published online: 15 Nov 2011.
To cite this article: Nikhil Mohe , Petri Heinonen , Yogesh S. Sanghvi & Roger Strömberg (2005) A SOLID SUPPORTED REAGENT
FOR INTERNUCLEOSIDE H-PHOSPHONATE LINKAGE FORMATION, Nucleosides, Nucleotides and Nucleic Acids, 24:5-7, 897-899,
DOI: 10.1081/NCN-200059257
To link to this article: http://dx.doi.org/10.1081/NCN-200059257
PLEASE SCROLL DOWN FOR ARTICLE
Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) contained
in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no
representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the
Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and
are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and
should be independently verified with primary sources of information. Taylor and Francis shall not be liable for
any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever
or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of
the Content.
This article may be used for research, teaching, and private study purposes. Any substantial or systematic
reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any
form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://
www.tandfonline.com/page/terms-and-conditions

Nucleosides, Nucleotides, and Nucleic Acids, 24 (5–7):897–899, (2005)
Copyright D Taylor & Francis, Inc.
ISSN: 1525-7770 print/ 1532-2335 online
DOI: 10.1081/NCN-200059257
A SOLID SUPPORTED REAGENT FOR INTERNUCLEOSIDE
H-PHOSPHONATE LINKAGE FORMATION
5
Nikhil Mohe and Petri Heinonen
Division of Organic and Bioorganic Chemistry,
MBB, Scheele Laboratory, Karolinska Institutet, Stockholm, Sweden
5
Yogesh S. Sanghvi
Isis Pharmaceuticals, Inc., Carlsbad, California, USA
Division of Organic and Bioorganic Chemistry, MBB, Scheele
5
Roger Str¨omberg
Laboratory, Karolinska Institutet, Stockholm, Sweden
5
A fast and convenient procedure for synthesis of dinucleoside H-phosphonates is obtained through use
of the novel polystyrene supported 5-carboxy-5-methyl-2-oxo-2-chloro-1,3,2-dioxaphosphorinane reagent.
Virtually quantitative H-phosphonate condensations are obtained leading to excellent isolated yields and
with only a simple filtration as the purification procedure. This provides for a convenient and high-
yielding procedure that should be suited for solution-phase synthesis of oligonucleotides.
RESULTS AND DISCUSSION
We describe the use of the novel polymer bound 5-carboxy-5-methyl-2-oxo-2-
chloro-1,3,2-dioxaphosphorinane reagent[1] in synthesis of dinucleoside H-phos-
phonates.[2,3] This is to our knowledge the first reported solid supported
chlorophosphate. This reagent is synthesized from isopropylidene-2,2-bis(me-
thoxy)propionic acid, which is first coupled to aminomethylpolystyrene with
HBTU/DIPEA in DMF. The isopropylidene group is removed with p-toluene-
sulfonic acid in CH2Cl2/CH3OH and the support is then treated with POCl3 in
CH3CN/collidine. This chlorophosphate reagent was utilised in preparation of
dinucleoside H-phosphonate diesters (Scheme 1).
Reactions were carried out in acetonitrile-pyridine (1:1 or 3:1 v/v, depending on
solubility) using 1.78 mg of reagent/mmol of H-phosphonate. Virtually quantitative
coupling was obtained in 1 h as judged by 31P NMR analysis of the reaction
mixtures. In the preparations a reaction time of 2 h was used. Filtering off of the
polymer and washing with phosphate buffer (pH = 7.0) followed by drying and
Address correspondence to Roger Stro¨mberg, Division of Organic and Bioorganic Chemistry, MBB, Scheele
Laboratory, Karolinska Institutet, S-17177, Stockholm, Sweden; Fax: +46-8-6089127; E-mail: Roger.Stromberg@
bioci.ki.se
897
Order reprints of this article at www.copyright.rightslink.com

898
N. Mohe et al.
SCHEME 1 Use of solid supported 5-carboxy-5-methyl-2-oxo-2-chloro-1,3,2-dioxaphosphorinane for synthesis of
di (deoxynucleoside) H-phosphonates.
concentration led to isolation of the products. Only traces of remaining H-
phosphonate monoesters were detected (primarily because of its use in slight
excess) in the crude material. The material was sufficiently pure for direct use in
further transformations, but the remaining monoesters were also readily removed
by filtration through silica gel with 1,4 dioxane as the solvent. This procedure gave
excellent yields (76–81%) of the isolated H-phosphonate diesters 1–4. The prod-
1
ucts were all characterized by 31P NMR, H NMR, and ESI-MS analysis. In prep-
aration of compound 4 no side product from potential reaction on the lactam
function of the guanine base was detected even after keeping the reaction mixture
in contact with the solid supported chlorophosphate for 8 h.
The above procedure provides for a convenient and high-yielding method for
synthesis of dinucleoside H-phosphonates. Distinct advantages are that the reagent
can be made in a few steps with relatively inexpensive materials and that the H-
phosphonate diesters generated can be conveniently isolated in highly pure form by
a simple filtration step. The compounds can then typically be used directly for
further transformations to dinucleotides or different modified analogues. In
addition, the procedure provides an excellent alternative for the coupling step in
solution-phase synthesis of oligonucleotides. The use of the H-phosphonate
method[2,4–8] especially in a modified form[9,10] has been actualised as an
economic alternative for the large scale synthesis of therapeutic oligonucleotides.[11]
REFERENCES
1. Mohe, N.; Heinonen, P.; Sanghvi, Y.; Stro¨mberg, R. A universal solid supported condensing agent. Angew.
Chem., to be submitted.

N. Mohe et al.
899
2. Garegg, P.J.; Regberg, T.; Stawinski, J.; Stro¨mberg, R. Formation of internucleotidic bonds via phosphonate
intermediates. Chem. Scr. 1985, 25, 280.
3. Stawinski, J.; Stro¨mberg, R.; Thelin, M.; Westman, E. Studies on the t-butyl-dimethylsilyl group as 2’-O-
protection in oligoribonucleotide synthesis via the H-phosphonate approach. Nucleic Acids Res. 1988, 16,
9285.
4. Garegg, P.J.; Regberg, T.; Stawinski, J.; Stro¨mberg, R. Nucleoside hydrogenphosphonates in oligonucleotide
synthesis. Chem. Scr. 1986, 26, 59.
5. Garegg, P.J.; Lindh, I.; Regberg, T.; Stawinski, J.; Stro¨mberg, R.; Henrichson, C. Nucleoside H-phosphonates.
III. Chemical synthesis of oligodeoxyribonucleotides by the hydrogenphosphonate approach. Tetrahedron
Lett. 1986, 27, 4051–4054.
6. Garegg, P.J.; Lindh, I.; Regberg, T.; Stawinski, J.; Stro¨mberg, R.; Henrichson, C. Nucleoside H-phosphonates.
IV. Automated solid phase synthesis of oligo-ribonucleotides by the hydrogenphosphonate approach.
Tetrahedron Lett. 1986, 27, 4055–4058.
7. Froehler, B.C.; Matteucci, M.D. Nucleoside H-phosphonates: valuable intermediates in the synthesis of
deoxyoligonucleotides. Tetrahedron Lett. 1986, 27, 469–472.
8. Froehler, B.C.; Ng, P.G.; Matteucci, M.D. Synthesis of DNA via deoxynucleoside H-phosphonate
intermediates. Nucleic Acids Res. 1986, 14, 5399–5407.
9. Reese, C.B.; Song, Q. A new approach to the synthesis of oligonucleotides and their phosphorothioate
analogs in solution. Bioorg. Med. Chem. Lett. 1997, 7, 2787.
10. Reese, C.B.; Song, Q. The H-phosphonate approach to the synthesis of oligonucleotides and their
phosphorothioate analogs in solution. J. Chem. Soc., Perkin Trans. 1 1999, 1477.
11. Reese, C.B.; Yan, H. Solution phase synthesis of ISIS 2922 (Vitravene) by the modified H-phosphonate
approach. J. Chem. Soc., Perkin Trans. 1 2002, 2619.