Process for preparing oligoribonucleoside and oligodeoxyribonucleoside boranophosphates

• US 5,177,198 A
• Filed: 11/30/1989
• Issued: 01/05/1993
• Est. Priority Date: 11/30/1989
• Status: Expired due to Fees

• Current Assignee: Boron Biologicals Incorporated, Duke University, University of North Carolina At Chapel Hill (University of North Carolina System)
• Original Assignee: Boron Biologicals Incorporated, Duke University, University of North Carolina At Chapel Hill (University of North Carolina System)
• Inventors: Sood, Anup, Spielvogel, Bernard F., Hall, Iris H., Ramsay Shaw, Barbara
• Primary Examiner(s): Brown, Johnnie R.
• Assistant Examiner(s): Wilson, James O.
• Application Number:
  US07/443,781
• Time in Patent Office: 1,132 Days
• Field of Search: 435/6, 514/44, 514/64, 536/18.1, 536/27, 536/28, 536/29, 536/51, 536/122, 536/64, 604/20
• US Class Current: 536/25.33
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• 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: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 2310/31 of the backbone
• CPC Class Codes: C12N 2310/312 Phosphonates
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/353 linked to the nucleic acid via an atom other than carbon
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: Y02P 20/55 Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups

Abstract

A process of making oligoribonucleoside and deoxyribonucleoside boranophosphates and salts thereof is disclosed. The process comprises the steps of condensing a ribonucleotide or deoxyribonucleotide with a nucleoside phosphitylating agent, wherein the phosphitylating agent includes a 5'"'"' -OH group protected by an acid-labile protecting group and a moiety bonded to the phosphorus atom therein that is a stronger proton acceptor than the 5'"'"'-OH group of the ribonucleotide or deoxyribonucleotide to form a reaction intermediate, then oxidizing the reaction intermediate so formed with a compound comprising a Lewis base and a boron-containing substituent, wherein the Lewis base is a weaker electron donor than the phosphite phosphorus of the reaction intermediate, to form a ribonucleotide or deoxyribonucleotide boranophosphate. These steps are repeated on the nucleotide boranophosphate to form an oligonucleotide boranophosphate.

Claims

1. A process of making an oligonucleoside boranophosphate, comprising (a) condensing a compound of formula (i) ##STR8## wherein:

Ac is an acyl group or a solid support;

n is from 0 to 50;

N is a ribonucleoside or deoxyribonucleoside;

W is selected from the group consisting of ═

O, ═

S, --OPr, and --SPr where Pr is a base-labile protecting group;

X is selected from the group consisting of --OH, --BH₃, --BH₂ R₁, --BHR₁ R₂ and --BR₁ R₂ R₃ ;

R₁ is selected from the group consisting of --R₄, --COOOH, --COOR₄, --CONHR₄, --CON(R₄)₂, --CN⁺ R₄, --CN, carboxycholesteryl and carboxybenzyl, wherein R₄ is C1 to C18 alkyl;

R₂ is selected from the group consisting of --R₅, --COOH, --COOR₅, --CONHR₅, --CON(R₅)₂, --CN⁺ R₅, --CN, carboxychoesteryl and carboxybenzyl, wherein R₅ is C1 to C18 alkyl; and

R₃ is selected from the group consisting of C1 to C3 alkyl,

with a compound of formula (ii) ##STR9## wherein;

Pr'"'"' is a Lewis acid-labile protecting group;

N is a ribonucleoside or deoxyribonucleoside;

Y is selected from the group consisting of --OPr, and --SPr where Pr is a base-labile protecting group; and

Q is a proton acceptor which is a stronger proton acceptor than the 5'"'"' oxygen atom of compound (i);

to produce a compound of formula (iii), ##STR10## and then;

(b) oxidizing the compound of formula (iii) with a compound of the formula LX, wherein

L is a Lewis base which is a weaker electron donor than the phosphite phosphorus of compound (iii); and

X is selected from the group consisting of --BH₃, --BH₂ R₁, --BHR₁ R₂ and --BR₁ R₂ R₃ ;

wherein;

R₁ is selected from the group consisting of --R₄, --COOH, --COOR₄, --CONHR₄, --CON(R₄)₂, --CN⁺ R₄, --CN, carboxycholesteryl and carboxybenzyl, wherein R₄ is C1 to C18 alkyl;

R₂ is selected from the group consisting of --R₅, --COOH, --COOR₅, --CONHR₅, --CON(R₅)₂, --CN⁺ R₅, --CN, carboxycholesteryl and carboxybenzyl, wherein R₅ is C1 to C18 alkyl; and

R₃ is selected from the group consisting of C1 to C3 alkyl, to give a compound of formula (iv) ##STR11## and then (c) repeating at least step (a) by substituting the compound of formula (iv) for the compound of formula (i) in step (a).

2. A process according to claim 1, wherein n is from 0 to 30.

3. A process according to claim 1, wherein n is from 0 to 18.

4. A process according to claim 1, wherein W is selected from the group consisting of ═

O, ═

S, --OR'"'"', --SR'"'"' and --)CH₂ "CH₂ CN, wherein R'"'"' is C1 to C3 alkyl.

5. A process according to claim 1, wherein Ac is an acyl group.

6. A process according to claim 1, wherein Ac is acetyl.

7. A process according to claim 1, wherein Ac is a solid support.

8. A process according to claim 1, wherein Ac is controlled pore glass.

9. A process according to claim 1, wherein Pr is selected from the group consisting of C1 to C3 alkyl, 2,2,2-tribomoethyl-, benzyl-, 2-cyanoethyl-, 2-methylsulfonylethyl, 4-nitrophenylethyl, 2-chlorophenyl, p-chlorophenyl, pehnylethyl, and p-nitrophenylethyl.

10. A process according to claim 1, wherein Q is selected from the group consisting of chlorine, 1-substituted 1,2,4-trizole, and 1-substituted 1,2,3,4-tetrazole.

11. A process according to claim 1, wherein L is selected from the group consisting of --S(CH₃)₂ and --N(CH₃)₃.

12. A process according to claim 2, wherein N is a ribonucleoside or deoxyribonucleoside having a base selected from the group consisting adenine, thymine, cytosine, guanine, uracil, xanthine, hypoxanthine, 5-methylcytosine, 5-bromouracil, 8-azidoadene, and 2,6-diaminopurine.

13. A process according to claim 1, wherein:

n is from 0 to 18;

W is selected from the group consisting of ═

O, ═

S, --OR'"'"', --SR'"'"' and --)CH₂ CH₂ CN, wherein R'"'"' is C1 to C3 alkyl;

Ac is an acyl group;

Pr is selected from eh group consisting of C1 to C3 alkyl, 2,2,2-tribromoethyl-, benzyl-, 2-cyanoethyl-, 2-methylsulfonylethyl, 4-nitroophenylethyl, 2-chlorophenyl, p-chlorophenyl, phenylethyl, and p-nitrophenylethyl;

Q is selected from the group consisting of chlorine, 1-substituted 1,2,4-triazole, and 1-substituted 1,2,3,4-tetrazole;

L is selected from the group consisting of --S(CH₃)₂ and --N(CH₃)₃.

14. A process according to claim 13, wherein n is from 0 to 3.

15. A process according to claim 14, wherein Ac is acetyl.

16. A process according to claim 15, wherein N is a ribonucleoside or deoxyribonucleoside having a base selected from the group consisting adenine, thymine, cytosine, guanine, uracil, xanthine, hypoxanthine, 5-methylcytosine, 5-bromouracil, 8-azidoadenine, and 2,6-diaminopurine.