Process for oligonucleo tide synthesis using phosphormidite intermediates

• US 4,973,679 A
• Filed: 09/18/1986
• Issued: 11/27/1990
• Est. Priority Date: 03/27/1981
• Status: Expired due to Term

• Current Assignee: University Patents, Inc.
• Original Assignee: University Patents, Inc.
• Inventors: Caruthers, Marvin H., Beaucage, Serge L.
• Primary Examiner(s): Rollins, John W.
• Assistant Examiner(s): Crane, L. Eric
• Application Number:
  US06/909,070
• Time in Patent Office: 1,531 Days
• Field of Search: 536/27, 536/28, 536/29
• US Class Current: 536/26.71
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids

Abstract

A new class of nucleoside phosphoramidites which are relatively stable to permit isolation thereof and storage at room temperature. The phosphoramidites are derivatives of saturated secondary amines.

Claims

1. In a process for producing oligonucleotides which comprises the step of condensing the 3'"'"'-OH or 5'"'"'-OH group of a nucleoside or oligonucleotide with a nucleoside phosphite, the improvement being that the nucleoside phosphite is a compound of one of the formulae:

##STR24## wherein B is a nucleoside or deoxynucleoside base;

A is H OH or OR₄ in which R₄ is a blocking group;

R is a blocking group;

R₁ '"'"' is a hydrocarbyl radical containing up to about carbon atoms; and

X is NR₂ '"'"'R₃ '"'"', wherein R₂ '"'"' and R₃ '"'"' taken separately each represent alkyl, aryl, aralkyl, cycloalkyl and cycloalkylalkyl containing up to 10 carbon atoms;

R₂ '"'"' and R₃ '"'"' when taken together form an alkylene chain containing up to 5 carbon atoms in the principal chain and a total of up to 10 carbon atoms with both terminal valence bonds of said chain being attached to the nitrogen atom to which R₂ '"'"' and R₃ '"'"' are attached; and

R₂ '"'"' and R₃ '"'"' when taken together with the nitrogen atom to which they are attached form a saturated nitrogen heterocycle including at least one additional heteroatom from the group consisting of nitrogen, oxygen and sulfur.

2. The process according to claim 1 wherein R is a trityl group.

3. The process according to claim 1 wherein R is a di-p-anisylphenylmethyl.

4. The process according to claim 1 wherein R is p-anisyldiphenylmethyl.

5. The process according to claim 1 wherein R₁ '"'"' is lower alkyl.

6. The process according to claim 1 wherein X is di-lower alkylamino.

7. The process according to claim 1 wherein X is a saturated nitrogen heterocyclic.

8. The process according to claim 1 wherein B is adenine, guanine, cytosine, uracil or thymine.

9. The process according to claim 1 wherein R is di-p-anisylphenylmethyl, B is 9-(N-6-benzoyladeninyl), R₁ '"'"' is methyl, A is H and X is morpholino.

10. The process according to claim 1 wherein R is di-p-anisylphenylmethyl, B is thyminyl, R₁ is methyl, A is H and X is dimethylamino.

11. The process according to claim 1 wherein R is di-p-anisylphenylmethyl, B is 1-(N-4-benzoylcytosinyl), R₁ '"'"' is methyl, A is H and X is dimethylamino.

12. The process according to claim 1 wherein R is di-p-anisylphenylmethyl, B is 9-(N-6-benzoyladeninyl), R₁ '"'"' is methyl, A is H and X is dimethylamino.

13. The process according to claim 1 wherein R is di-p-anisylphenylmethyl, B is 9-(N-2-isobutyrylguaninyl), R₁ '"'"' is methyl, A is H and X is dimethylamino.

14. The process according to claim 1 wherein said nucleoside or oligonucleotide is covalently bonded to an inorganic polymer.

15. The process according to claim 14 wherein said nucleoside or oligonucleotide is linked to the inorganic polymer through a base hydrolyzable covalent bond.

16. The process according to claim 15 wherein the base hydrolyzable covalent bond is an ester linkage.