Oligonucleotide synthesis with substituted aryl carboxylic acids as activators

• US 5,574,146 A
• Filed: 08/30/1994
• Issued: 11/12/1996
• Est. Priority Date: 08/30/1994
• Status: Expired due to Term

• Current Assignee: Beckman Instruments, Inc. (Danaher Corporation)
• Original Assignee: Beckman Instruments, Inc. (Danaher Corporation)
• Inventors: Farooqui, Firdous, Reddy, M. Parameswara
• Primary Examiner(s): Kunz, Gary L.
• Application Number:
  US08/298,545
• Time in Patent Office: 805 Days
• Field of Search: 536/26.1, 536/25.34, 536/55.3
• US Class Current: 536/25.34
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 19/20 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• 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
• CPC Class Codes: Y02P 20/55 Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Abstract

A process for synthesizing oligonucleotides by phosphoramidite chemistry wherein the improvement is the use of substituted aryl carboxylic acids as the activators. These activators produce in situ nucleotide intermediates in which the substituted arylcarbonyl group has displaced the amidite moiety.

Claims

1. A composition comprising a secondary amine and an in situ generated compound of formula Ia or Ib:

##STR7## wherein;

B is a purine or pyrimidine base;

R is a hydroxyl-protecting group;

R'"'"' is a substituted aryl;

R" is selected from the group consisting of lower alkyl, lower alkoxy, NCCH₂ CH₂ O--, NCCH₂ CH₂ MeO--, NCCH₂ CMe₂ O--, Cl₃ CCH₂ O--, Cl₃ CCMe₂ O--, C₆ H₅ SO₂ CH₂ CH₂ O--, MeSO₂ CH₂ CH₂ O--, and NO₂ C₆ H₄ CH₂ CH₂ O--.

2. The composition according to claim 1 wherein R'"'"' is an aryl group bearing at least one electron-withdrawing substituent which does not interfere with oligonucleotide synthesis.

3. The composition of claim 2, wherein aryl is selected from the group consisting of phenyl, naphthyl, and anthracyl.

4. The composition of claim 2, wherein the electron-withdrawing substitutent is selected from the group consisting of halogen or nitro.

5. The composition of claim 1, wherein R'"'"' is selected from the group consisting of 2-nitrophenyl;

3,5-dinitrophenyl;

2,4,5-trifluorophenyl;

2,3,6-trifluorophenyl;

2,3,5,6,-tetrafluorophenyl;

pentafluorophenyl;

3-nitrophenyl; and

2,4-dinitrophenyl.

6. The composition of claim 5, wherein R'"'"' is 2,4-dinitrophenyl.

7. The composition of claim 1, wherein R" is selected from the group consisting of lower alkoxy, NCCH₂ CH₂ O--, NCH₂ CH₂ MeO--, NCCH₂ CMe₂ O--, Cl₃ CCH₂ O--, Cl₃ CCMe₂ O--, C₆ H₅ SO₂ CH₂ CH₂ O--, MeSO₂ CH₂ CH₂ O--, and NO₂ C₆ H₄ CH₂ CH₂ O--.

8. The composition of claim 1, wherein R" is lower alkyl.

9. The composition of claim 1, wherein R is selected from the group consisting of 4,4'"'"'-dimethoxytrityl;

4,4'"'"',4"-tris(benzyloxy)trityl;

4,4'"'"',4"-tris-(4,5-dichlorophthalimido)trityl;

4,4'"'"',4"-tris-(levulinyloxy)trityl;

3-(imidazolylmethyl)-4,4'"'"',-dimethoxytrityl;

pixyl(9-phenylxanthen-9-yl);

9-(p-methoxyphenyl)xanthen-9-yl);

4-decyloxytrityl;

4-hexadecyloxytrityl;

9-(4-octadecyloxyphenyl)xanthene-9-yl;

1,1-bis-(4-methoxyhenyl)-1'"'"'-pyrenyl methyl;

p-phenylazophenyloxycarbonyl;

9-fluorenylmethoxycarbonyl;

2,4-dinitrophenylethoxylcarbonyl;

4-(methylthiomethoxy)butyryl;

2-(methylthiomethoxymethyl)-benzoyl;

2-(isopropylthiomethoxymethyl)benzoyl;

2-(2,4-dinitrobenzenesulphenyloxymethyl)benzoyl; and

levulinyl.

10. The composition of claim 9, wherein R is dimethoxytrityl.

11. The composition of claim 1, wherein B is selected from the group consisting of adenine, guanine, cytosine, and thymine.

12. A process for the preparation of the composition according to claim 1 containing the in situ genereated compound of formulae Ia or Ib comprising:

reacting the compound of either formula IIa or IIb ##STR8## wherein R and R" are defined as above and each R'"'"'" is lower alkyl or when taken together form the group --(CH₂)₂ --O--(CH₂)₂ --

with an acid of general formula R'"'"'--C--OH in a suitable solvent.

13. In a process of oligonucleotide synthesis in which nucleoside phosphoramidite monomer precursors are activated by reaction with an activating agent to form activated intermediates and the activated intermediates are sequentially added to form an oligonucleotide product, wherein the improvement comprises using an substituted aryl carboxylic acid as the acid catalyst and thus generating in situ the compound of formula Ia or Ib according to claim 1.