Nucleic acid ligand complexes

• US 6,465,188 B1
• Filed: 05/10/2000
• Issued: 10/15/2002
• Est. Priority Date: 06/11/1990
• Status: Expired due to Fees

• Current Assignee: Gilead Sciences Inc.
• Original Assignee: Gilead Sciences Inc.
• Inventors: Schmidt, Paul G, Janjic, Nebojsa, Gold, Larry
• Primary Examiner(s): ZITOMER, STEPHANIE W
• Application Number:
  US09/569,572
• Time in Patent Office: 888 Days
• Field of Search: 435/6, 435/91.2, 536/23.1, 536/25.4, 514/44
• US Class Current: 435/6.11
• 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: A61K 9/1271 Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers liposome as conjugate A61K47/6911
• CPC Class Codes: A61K 9/1272 with substantial amounts of non-phosphatidyl, i.e. non-acylglycerophosphate, surfactants as bilayer-forming substances, e.g. cationic lipids with cholesterol as the only non-phosphatidyl surfactant A61K9/127; lipids as modifying agent A61K47/543
• 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: 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 9/1276 RNA-directed DNA polymerase (2.7.7.49), i.e. reverse transcriptase or telomerase
• CPC Class Codes: C12Q 1/37 involving peptidase or proteinase
• CPC Class Codes: C12Q 1/6811 Selection methods for production or design of target specific oligonucleotides or binding molecules
• CPC Class Codes: C12Q 2541/101 Selex
• CPC Class Codes: C12Q 2563/161 Vesicles, e.g. liposome
• CPC Class Codes: C40B 40/00 Libraries per se, e.g. arrays, mixtures
• CPC Class Codes: F02B 2075/027 four
• CPC Class Codes: G01N 2333/163 : Regulatory proteins, e.g. tat, nef, rev, vif, vpu, vpr, vpt, vpx
• CPC Class Codes: G01N 2333/575 : Hormones derived from pro-opiomelanocortin, pro-enkephalin or pro-dynorphin G01N2333/665, corticotropin G01N2333/695
• CPC Class Codes: G01N 2333/62 : Insulins
• CPC Class Codes: G01N 2333/8125 : Alpha-1-antitrypsin
• CPC Class Codes: G01N 2333/96433 : Serine endopeptidases (3.4.21)
• CPC Class Codes: G01N 2333/96455 : Kallikrein (3.4.21.34; 3.4.21.35)
• CPC Class Codes: G01N 2333/966 : Elastase
• CPC Class Codes: G01N 2333/9726 : Tissue plasminogen activator
• CPC Class Codes: G01N 2333/974 : Thrombin
• CPC Class Codes: G01N 2333/976 : Trypsin; Chymotrypsin
• CPC Class Codes: G01N 33/531 : Production of immunochemical test materials
• CPC Class Codes: G01N 33/532 : Production of labelled immunochemicals
• CPC Class Codes: G01N 33/535 : with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
• CPC Class Codes: G01N 33/56988 : HIV or HTLV
• CPC Class Codes: G01N 33/68 : involving proteins, peptides or amino acids involving lipoproteins G01N33/92
• CPC Class Codes: G01N 33/76 : Human chorionic gonadotropin including luteinising hormone, follicle stimulating hormone, thyroid stimulating hormone or their receptors

Abstract

This invention discloses a method for preparing a therapeutic or diagnostic complex comprised of a nucleic acid ligand and a lipophilic compound or non-immunogenic, high molecular weight compound by identifying a nucleic acid ligand by SELEX methodology and associating the nucleic acid ligand with a lipophilic compound or a non-immunogenic, high molecular weight compound. The invention further discloses complexes comprising one or more nucleic acid ligands in association with a lipophilic compound or non-immunogenic, high molecular weight compound.

Claims

1. A method for the preparation of a therapeutic or diagnostic Complex comprised of a Nucleic Acid Ligand and a Lipophilic Compound or a Non-Immunogenic, High Molecular Weight Compound, said method comprising:

identifying a Nucleic Acid Ligand from a Candidate Mixture of Nucleic Acids, said Nucleic Acid Ligand being a ligand of a given Target by the method comprising;

a) contacting the Candidate Mixture with the Target, wherein Nucleic Acids having an increased affinity to the Target 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 ligand-enriched mixture of Nucleic Acids; and

d) covalently linking said identified Nucleic Acid Ligand with a Lipophilic Compound or a Non-Immunogenic, High Molecular Weight Compound.

2. The method of claim 1 wherein said Complex is comprised of a Nucleic Acid Ligand and Non-Immnunogenic, High Molecular Weight Compound.

3. The method of claim 2 wherein said Non-Immunogenic, High Molecular Weight Compound is selected from the group consisting of polyethylene glycol, dextran, albumin, and magnetite.

4. The method of claim 3 wherein said Non-Immnunogenic, High Molecular Weight Compound is polyethylene glycol.

5. The method of claim 4, which further comprises attaching an additional therapeutic or diagnostic agent with said Nucleic Acid Ligand.

6. The method of claim 5 wherein said therapeutic or diagnostic agent is covalently linked to said polyethylene glycol.

7. The method of claim 1 wherein said Complex is comprised of a Nucleic Acid Ligand and Lipophilic Compound.

8. The method of claim 7 wherein said Lipophilic Compound is selected from the group consisting of cholesterol, dialkyl glycerol and diacyl glycerol.

9. The method of claim 8 wherein said Lipophilic Compound is cholesterol.

10. The method of claim 8 wherein said Lipophilic Compound is dialkyl glycerol.

11. The method of claim 1 wherein said Lipophilic Compound is a Lipid Construct.

12. The method of claim 11 wherein the Lipid Construct is a Lipid Bilayer Vesicle.

13. The method of claim 12 wherein said Lipid Bilayer Vesicle is a Liposome.

14. The method of claim 13 wherein said Nucleic Acid Ligand is projecting from the exterior surface of said Liposome.

15. The method of claim 13, which further comprises attaching an additional therapeutic or diagnostic agent with said nucleic acid ligand.

16. The method of claim 1 wherein said Nucleic Acid Ligand targets the Complex to a preselected location.

17. A method for improving the Pharmacokinetic Properties of a Nucleic Acid Ligand said method comprising:

identifying a Nucleic Acid Ligand from a Candidate Mixture of Nucleic Acids, said Nucleic Acid Ligand being a ligand of a given Target by the method comprising;

a) contacting the Candidate Mixture with the Target, wherein Nucleic Acids having an increased affinity to the Target 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 ligand-enriched mixture of Nucleic Acids; and

d) covalently linking said identified Nucleic Acid Ligand with a Lipophilic Compound or a Non-Immunogenic, High Molecular Weight Compound wherein the Pharmacokinetic Properties of said Complex are improved relative to the Nucleic Acid Ligand alone.