High affinity nucleic acid ligands to lectins

• US 6,001,988 A
• Filed: 06/07/1995
• Issued: 12/14/1999
• Est. Priority Date: 06/11/1990
• Status: Expired due to Fees

• Current Assignee: Gilead Sciences Inc.
• Original Assignee: NeXstar Pharmaceuticals, Inc. (Gilead Sciences Inc.)
• Inventors: Parma, David H., Hicke, Brian J., Gold, Larry, Bridonneau, Philippe
• Primary Examiner(s): Zitomer, Stephanie W.
• Application Number:
  US08/472,256
• Time in Patent Office: 1,651 Days
• Field of Search: 536/22.1, 536/25.4, 536/24.3, 435/6, 435/91.2, 935/77, 935/78
• US Class Current: 536/24.3
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 47/547 Chelates, e.g. Gd-DOTA or Zinc-amino acid chelates; Chelate-forming compounds, e.g. DOTA or ethylenediamine being covalently linked or complexed to the pharmacologically- or therapeutically-active agent
• CPC Class Codes: A61K 47/549 Sugars, nucleosides, nucleotides or nucleic acids
• CPC Class Codes: B82Y 5/00 Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
• CPC Class Codes: C07H 19/06 Pyrimidine radicals
• CPC Class Codes: C07H 19/10 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: C07K 14/001 by chemical synthesis
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C07K 14/72 for hormones for neuromediators C07K14/70571
• CPC Class Codes: C07K 7/22 Tachykinins, e.g. Eledoisins , Substance P; Related peptides
• CPC Class Codes: C12N 15/1048 SELEX
• CPC Class Codes: C12N 15/115 Aptamers, i.e. nucleic acids binding a target molecule specifically and with high affinity without hybridising therewith ; Nucleic acids binding to non-nucleic acids, e.g. aptamers
• CPC Class Codes: C12N 2310/13 Decoys
• CPC Class Codes: C12N 2310/322 2'-R Modification
• CPC Class Codes: C12N 2310/53 partially self-complementary or closed
• CPC Class Codes: C12N 2740/16222 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 9/1276 RNA-directed DNA polymerase (2.7.7.49), i.e. reverse transcriptase or telomerase
• CPC Class Codes: C12Q 1/6811 Selection methods for production or design of target specific oligonucleotides or binding molecules
• CPC Class Codes: C40B 40/00 Libraries per se, e.g. arrays, mixtures
• CPC Class Codes: F02B 2075/027 : four
• CPC Class Codes: G01N 2333/918 : Carboxylic ester hydrolases (3.1.1)

Abstract

This invention discloses high-affinity oligonucleotide ligands to lectins, specifically nucleic acid ligands having the ability to bind to the lectins, wheat germ agglutinin, L-selectin, E-selectin and P-selectin. Also disclosed are the methods for obtaining such ligands.

Claims

1. A purified and isolated non-naturally occurring nucleic acid ligand to a lectin selected from the group consisting of wheat germ agglutinin, serum mannose binding protein, and a selectin.

2. The nucleic acid ligand of claim 1 which is a non-naturally occurring nucleic acid ligand having a specific binding affinity for said lectin, such lectin being a three dimensional chemical structure other than a polynucleotide that binds to said nucleic acid ligand through a mechanism which predominantly depends on Watson/Crick base pairing or triple helix binding, wherein said nucleic acid ligand is not a nucleic acid having the known physiological function of being bound by said lectin.

3. The nucleic acid ligand of claim 1 wherein said lectin is wheat germ agglutinin.

4. The nucleic acid ligand to wheat germ agglutinin of claim 3 wherein said nucleic acid ligand is a ribonucleic acid (RNA).

5. The nucleic acid ligand to wheat germ agglutinin of claim 4 wherein said ligand is an RNA ligand selected from the group consisting of the nucleotide sequences set forth in Table II.

6. The nucleic acid ligand of claim 5 wherein said ligand is selected from the group consisting of SEQ ID NOS:

4-55.

7. The nucleic acid ligand of claim 4 wherein said ligand comprises sequences selected from the group consisting of SEQ ID NOS:

56-63.

8. The nucleic acid ligand to wheat germ agglutinin of claim 3 wherein said ligand is substantially homologous to and has substantially the same ability to bind said wheat germ agglutinin as a ligand selected from the group consisting of the sequences set forth in Table II.

9. The nucleic acid ligand to wheat germ agglutinin of claim 3 wherein said ligand has substantially the same structure and has substantially the same ability to bind said wheat germ agglutinin as a ligand selected from the group consisting of the sequences set forth in Table II.

10. The nucleic acid ligand of claim 1 wherein said lectin is a selectin.

11. The nucleic acid ligand of claim 10 wherein said selectin is selected from the group consisting of L-selectin, E-selectin and P-selectin.

12. The nucleic acid ligand of claim 11 wherein said selectin is L-selectin.

13. The nucleic acid ligand to L-selectin of claim 12 wherein said nucleic acid ligand is a ribonucleic acid (RNA).

14. The nucleic acid ligand to L-selectin of claim 12 wherein said nucleic acid ligand is a deoxyribonucleic acid (DNA).

15. The nucleic acid ligand to L-selectin of claim 13 wherein said ligand is an RNA ligand selected from the group consisting of the nucleotide sequences set forth in Table VIII.

16. The nucleic acid ligand of claim 15 wherein said ligand is selected from the group consisting of SEQ ID NOS:

67-117.

17. The nucleic acid ligand of claim 13 wherein said ligand comprises sequences selected from the group consisting of SEQ ID NOS:

118-125.

18. The nucleic acid ligand to L-selectin of claim 14 wherein said ligand is a DNA ligand selected from the group consisting of the nucleotide sequences set forth in Table XII.

19. The nucleic acid ligand of claim 18 wherein said ligand is selected from the group consisting of SEQ ID NOS:

129-167.

20. The nucleic acid ligand of claim 14 wherein said ligand comprises sequences selected from the group consisting of SEQ ID NOS:

168-171.

21. The nucleic acid ligand of claim 1 wherein said ligand is a ribonucleic acid ligand.

22. The nucleic acid ligand of claim 1 wherein said ligand is a deoxyribonucleic acid.

23. The nucleic acid ligand of claim 1 wherein said ligand comprises 2'"'"'-amino (2'"'"'-NH₂) modified ligands, 2'"'"'-fluoro (2'"'"'-F) modified ligands and 2'"'"'-O-methyl (2'"'"'-OMe) modified ligands.

25. The nucleic acid ligand to L-selectin of claim 12 wherein said ligand is substantially homologous to and has substantially the same ability to bind said L-selectin as a ligand selected from the group consisting of the sequences set forth in Table VIII.

26. The nucleic acid ligand to L-selectin of claim 12 wherein said ligand has substantially the same structure and substantially the same ability to bind said L-selectin as a ligand selected from the group consisting of the sequences set forth in Table VIII.

27. The nucleic acid ligand to L-selectin of claim 12 wherein said ligand is substantially homologous to and has substantially the same ability to bind said L-selectin as a ligand selected from the group consisting of the sequences set forth in Table XII.

28. The nucleic acid ligand to L-selectin of claim 12 wherein said ligand has substantially the same structure and substantially the same ability to bind said L-selectin as a ligand selected from the group consisting of the sequences set forth in Table XII.

24. A nucleic acid ligand to a lectin selected from the group consisting of wheat germ agglutinin, serum mannose binding protein, L-selectin, E-selectin and P-selectin, identified according to the method comprising:

a) contacting a candidate mixture of nucleic acids with said lectin, wherein nucleic acids having an increased affinity to said lectin relative to the candidate mixture may be partitioned from the remainder of the candidate mixture;

b) partitioning the increased affinity nucleic acids from the remainder of the candidate mixture; and

c) amplifying the increased affinity nucleic acids to yield a mixture of nucleic acids enriched for nucleic acid sequences with increased affinity and specificity for binding to said lectin, whereby nucleic acid ligands of said lectin may be identified.