Structure based and combinatorially selected oligonucleoside phosphorothioate and phosphorodithioate aptamer targeting AP-1 transcription factors

• US 9,567,579 B2
• Filed: 01/26/2011
• Issued: 02/14/2017
• Est. Priority Date: 05/23/2003
• Status: Active Grant

• Current Assignee: Board of Regents of the University of Texas System (University of Texas System)
• Original Assignee: Board of Regents of the University of Texas System (University of Texas System)
• Inventors: Gorenstein, David G., Luxon, Bruce A., Leary, James, Yang, Xianbin
• Primary Examiner(s): Gross, Christopher M
• Application Number:
  US13/014,597
• Publication Number:
  US 20110212843A1
• Time in Patent Office: 2,211 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: C12N 15/1034 Isolating an individual clone by screening libraries
• CPC Class Codes: C12N 15/1048 SELEX
• CPC Class Codes: G01N 33/6818 Sequencing of polypeptides
• CPC Class Codes: G01N 33/6848 Methods of protein analysis involving mass spectrometry
• CPC Class Codes: G01N 33/6851 Methods of protein analysis involving laser desorption ionisation mass spectrometry

Abstract

The present invention includes composition and methods for making and using a combinatorial library to identify modified thioaptamers that bind to, and affect the immune response of a host animal, transcription factors such as IL-6, NF-κB, AP-1 and the like. Composition and methods are also provided for the treatment of viral infections, as well as, vaccines and vaccine adjuvants are provided that modify host immune responses.

Claims

1. A system for identifying one or more proteins that interact with a specific oligonucleotide on a bead comprising:

an oligonucleotide-bead library for binding one or more proteins, wherein each bead of the oligonucleotide-bead library comprises more than one copy of a unique oligonucleotide having a unique sequence, wherein the unique oligonucleotides are attached to the beads by a hexaethyleneglycol linker, and wherein the oligonucleotides attached to the bead are not released during phosphoroamidite deprotection; and

a bead separator wherein the one or more beads of the bead library to which protein has bound are separated by the bead separator.

2. The system of claim 1, further comprising a nucleic acid sequencer to determine the sequence of the oligonucleotide.

3. The system of claim 1, wherein the sequence of the oligonucleotides on the one or more beads that bind to known proteins is determined.

4. The system of claim 1, wherein the one or more proteins bound to the oligonucleotide are extracted, separated and identified by liquid chromatography, mass spectrometry, or liquid chromatography and time of flight (TOF) mass spectrometry (MS) MALDI-TOF.

5. The system of claim 1, further comprising a mass spectrometer to identify the proteins that have bound to the one or more oligonucleotide-beads.

6. The system of claim 1, wherein the oligonucleotide is defined further as comprising an at least partially thio-modified oligonucleotide.

7. The system of claim 1, wherein the one or more proteins is further defined as being a protein extracted from a cell.

8. The system of claim 1, wherein the one or more proteins is further defined as being a protein extracted from a virally infected cell.

9. The system of claim 1, wherein the one or more proteins is further defined as being a protein extracted from a diseased cell.

10. The system of claim 1, wherein the bead library is substantially protein-free.

11. The system of claim 1, wherein the bead library comprises an [S]-oligonucleotide or an [S₂]-oligonucleotide combinatorial bead library.

12. The system of claim 1, wherein the oligonucleotides of the bead library are single stranded.

13. The system of claim 1, wherein the oligonucleotides of the bead library comprises RNA.

14. The system of claim 1, wherein the bead library comprises sequence motifs for high affinity with cellular proteins selected from the group consisting essentially of proteins that are members of the AP-1, RBP-Jκ

, NF-κ

B, NF IL-6 , CREB and GRE protein families, and combinations thereof.