Vascular endothelial growth factor (VEGF) nucleic acid ligand complexes

• US 5,859,228 A
• Filed: 10/25/1996
• Issued: 01/12/1999
• Est. Priority Date: 05/04/1995
• Status: Expired due to Term

• Current Assignee: Gilead Sciences Inc.
• Original Assignee: NeXstar Pharmaceuticals, Inc. (Gilead Sciences Inc.)
• Inventors: Janjic, Nebojsa, Gold, Larry, Schmidt, Paul
• Primary Examiner(s): ZITOMER, STEPHANIE W
• Application Number:
  US08/739,109
• Time in Patent Office: 809 Days
• Field of Search: 435/6, 435/91.2, 536/25.4, 536/24.3, 935/77, 935/78, 514/44
• 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 47/544 Phospholipids
• 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 14/52 Cytokines; Lymphokines; Interferons
• 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 15/88 using microencapsulation, e.g. using amphiphile liposome vesicle
• CPC Class Codes: C12N 2310/3515 Lipophilic moiety, e.g. cholesterol
• CPC Class Codes: C12Q 1/6811 Selection methods for production or design of target specific oligonucleotides or binding molecules

Abstract

This invention discloses a method for preparing a complex comprised of a VEGF Nucleic Acid Ligand and a Lipophilic Compound by identifying a VEGF Nucleic Acid Ligand by SELEX methodology and associating the VEGF Nucleic Acid Ligand with a Lipophilic Compound. The invention further discloses Complexes comprising one or more VEGF Nucleic Acid Ligands in association with a Lipophilic Compound. The invention further includes a Lipid construct comprising a VEGF Nucleic Acid Ligand or Complex and methods for making the same.

Claims

1. A method for the preparation of a Complex comprised of a vascular endothelial growth factor (VEGF) Nucleic Acid Ligand and a Lipophilic Compound, said method comprising:

a) contacting the Candidate Mixture with VEGF, wherein Nucleic Acids having an increased affinity to VEGF relative to the Candidate Mixture may be partitioned from the remainder of the Candidate Mixture;

b) partitioning the increased affinity VEGF Nucleic Acids from the remainder of the Candidate Mixture;

c) amplifying the increased affinity VEGF Nuclcic Acids to yield a ligand-enriched mixture of Nucleic Acids; and

d) covalently linking said identified VEGF Nucleic Acid Ligand with a Lipophilic Compound.

2. The method of claim 1 wherein said Lipophilic Compound is a lipid.

3. The method of claim 2 wherein said lipid is selected from the group consisting of dialkylglycerol and diacyiglycerol.

4. The method of claim 1 wherein said Complex is further associated with a Lipid Construct.

5. The method of claim 4 wherein said Lipid Construct is a Lipid Bilayer Vesicle.

6. The method of claim 5 wherein said Lipid Bilayer Vesicle is a Liposome.

7. A method for preparing a Lipid Construct comprising a Complex comprised of a VEGF Nucleic Acid Ligand and a Lipophilic Compound, said method comprising:

a) contacting the Candidate Mixture with VEGF, wherein Nucleic Acids having an increased affinity to VEGF relative to the Candidate Mixture may be partitioned from the rermainider of the Candidate Mixture;

b) partitioning the increased affinity VEGF Nucleic Acids from the remnainder of the Candidate Mixture;

c) amplifying the increased affinity VEGF Nucleic Acids to yield a ligand-enriched mixture of Nucleic Acids;

d) covalently linking said identified VEGF Nucleic Acid Ligand with a Lipophilic Compound to form a Complex; and

e) forming a Lipid Construct comprising said Complex.