Chiral control

• US 10,590,413 B2
• Filed: 03/17/2017
• Issued: 03/17/2020
• Est. Priority Date: 07/13/2012
• Status: Active Grant

• Current Assignee: WAVE Life Sciences Ltd.
• Original Assignee: WAVE Life Sciences Ltd.
• Inventors: Butler, David Charles Donnell, Iwamoto, Naoki, Meena, Svrzikapa, Nenad, Verdine, Gregory L., Zlatev, Ivan
• Primary Examiner(s): Bakshi, Pancham
• Application Number:
  US15/462,787
• Publication Number:
  US 20170275621A1
• Time in Patent Office: 1,096 Days
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07C 317/28 with sulfone or sulfoxide groups bound to acyclic carbon atoms of the carbon skeleton
• CPC Class Codes: C07D 295/088 to an acyclic saturated chain
• CPC Class Codes: C07H 1/02 Phosphorylation
• 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: C07H 21/02 with ribosyl as saccharide radical
• CPC Class Codes: C12N 15/11 DNA or RNA fragments; Modified forms thereof DNA or RNA not used in recombinant technology, C07H21/00; Non-coding nucleic acids having a biological activity
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding 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 2310/11 Antisense
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/31 of the backbone
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2330/30 chemically synthesised
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes

Abstract

The present invention relates to chirally controlled oligonucleotides, chirally controlled oligonucleotide compositions, and the method of making and using the same. The invention specifically encompasses the identification of the source of certain problems with prior methodologies for preparing chiral oligonucleotides, including problems that prohibit preparation of fully chirally controlled compositions, particularly compositions comprising a plurality of oligonucleotide types. In some embodiments, the present invention provides chirally controlled oligonucleotide compositions. In some embodiments, the present invention provides methods of making chirally controlled oligonucleotides and chirally controlled oligonucleotide compositions.

Claims

1. A method for making a chirally controlled oligonucleotide comprising steps of:

(1) coupling;

(2) capping;

(3) modifying;

(4) deblocking; and

(5) repeating steps (1)-(4) until a desired length is achieved;

wherein at least one cycle of (1) to (4) forms an internucleotidic linkage having the structure of formula I-c;

2. The method of claim 1, wherein the step of coupling comprises the use of N-cyanomethylpyrrolidinium triflate and

3. The method of claim 1, wherein the step of capping comprises capping of the amino group in the chiral auxiliary and capping of unreacted 5′

-OH.

4. The method of claim 3, wherein the step of capping comprises capping of the amino group in the chiral auxiliary using phenoxyacetic anhydride.

5. The method of claim 1, wherein the step of deblocking comprises use of acid.

6. The method of claim 1, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 10% of the crude product.

7. The method of claim 6, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 5% of the crude product.

8. The method of claim 6, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 4% of the crude product.

9. The method of claim 6, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 3% of the crude product.

10. The method of claim 6, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 2% of the crude product.

11. The method of claim 6, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 1% of the crude product.

12. The method of claim 6, wherein all oligonucleotides that are one nucleotide shorter than the desired length, when combined, are less than 0.5% of the crude product.

13. The method of claim 1, wherein the chirally controlled oligonucleotide is provided in a pre-determined amount.

14. The method of claim 1, wherein a phosphorothioate diester precursor is used for each phosphorothioate diester linkage in the chirally controlled oligonucleotide.

15. The method of claim 14, wherein the phosphorothioate diester precursor is

16. The method of claim 15, wherein each phosphorothioate diester precursor is converted to a phosphorothioate diester linkage after the desired oligonucleotide length is achieved.

17. The method of claim 1, wherein at least one modifying step is an oxidation step to install a phosphate diester linkage.

18. The method of claim 1, wherein the chirally controlled oligonucleotide is at least 10 nucleotide units in length.

19. The method of claim 1, wherein the step of capping comprises steps of capping of an amino group in a chiral auxiliary and capping of unreacted 5′

-OH, wherein the chiral auxiliary is of the structure;

20. The method of claim 19, wherein one of G¹ and G² is optionally substituted alkyl and the other is optionally substituted phenyl.