Compositions and methods for recognition of RNA using triple helical peptide nucleic acids

• US 10,260,089 B2
• Filed: 10/29/2013
• Issued: 04/16/2019
• Est. Priority Date: 10/29/2012
• Status: Active Grant

• Current Assignee: The Research Foundation for The State University of New York (State University of New York)
• Original Assignee: The Research Foundation for The State University of New York (State University of New York)
• Inventors: Rozners, Eriks, Zengeya, Thomas
• Primary Examiner(s): Switzer, Juliet C
• Application Number:
  US14/066,006
• Publication Number:
  US 20140206838A1
• Time in Patent Office: 1,995 Days
• Field of Search: None
• CPC Class Codes: C07K 14/003 Peptide-nucleic acids (PNAs)
• CPC Class Codes: C07K 7/08 having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68
• CPC Class Codes: C07K 9/00 Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof
• CPC Class Codes: C12Q 1/6839 Triple helix formation or other higher order conformations in hybridisation assays
• CPC Class Codes: C12Q 2525/107 incorporating a peptide nucleic acid

Abstract

Peptide nucleic acids containing thymidine and 2-aminopyridine (M) nucleobases formed stable and sequence selective triple helices with double stranded RNA at physiologically relevant conditions. The M-modified PNA displayed unique RNA selectivity by having two orders of magnitude higher affinity for the double stranded RNAs than for the same DNA sequences. Preliminary results suggested that nucleobase-modified PNA could bind and recognize double helical precursors of microRNAs.

Claims

1. A method of forming a pH 7.4-stable peptide nucleic acid-double stranded ribonucleic acid triple helix, comprising:

providing a peptide nucleic acid sequence having at least one basic amino acid covalently linked to a nucleic acid sequence portion configured to selectively bind to a complementary nucleic acid sequence portion, wherein the nucleic acid sequence portion comprises at least one 2-aminopyridine nucleobase; and

contacting a peptide nucleic acid having the peptide nucleic acid sequence with double stranded ribonucleic acid having the complementary nucleic acid sequence portion, to form a peptide nucleic acid double-stranded ribonucleic acid triple helix, in a nucleic acid sequence portion selective manner.

2. The method according to claim 1, wherein the peptide nucleic acid sequence portion has a higher affinity for the double stranded ribonucleic acid having the complementary nucleic acid sequence portion than for a double stranded deoxyribonucleic acid having a corresponding complementary nucleic acid sequence portion.

3. The method according to claim 1, wherein the peptide nucleic acid comprises at least six nucleobases including at least three 2-aminopyridine nucleobases.

4. The method according to claim 1, wherein the at least one amino acid comprises lysine.

5. A method of forming a peptide nucleic acid-double stranded nucleic acid triple helix, comprising:

providing a peptide nucleic acid, comprising a nucleic acid sequence-selective binding portion, the nucleic acid sequence-selective binding portion comprising a plurality of nucleobases and at least one 2-aminopyridine nucleobase, conjugated to at least one basic amino acid; and

interacting the peptide nucleic acid with a double stranded nucleic acid having a nucleic acid sequence corresponding to the nucleic acid sequence-selective binding portion, to form the peptide nucleic acid-double stranded nucleic acid triple helix selectively in dependence on the nucleic acid sequence-selective binding portion.

6. The method according to claim 5, wherein the nucleic acid sequence-selective binding portion comprises at least six nucleobases.

7. The method according to claim 5, wherein the nucleic acid sequence-selective binding portion comprises at least three 2-aminopyridine nucleobases.

8. The method according to claim 5, wherein the nucleic acid sequence-selective binding portion comprises at least six nucleobases including at least three 2-aminopyridine nucleobases.

9. The method according to claim 5, wherein the nucleic acid sequence-selective binding portion comprises a copolymer comprising 2-aminopyridine and thymidine.

10. The method according to claim 5, wherein the peptide nucleic acid-double stranded nucleic acid triple helix is stable at pH 7.4 at temperatures less than about 35°

C. in a phosphate buffer solution containing 2 mM magnesium chloride, 90 mM potassium chloride, 10 mM sodium chloride, and 50 mM potassium phosphate.

11. The method according to claim 5, wherein the peptide nucleic acid has a higher affinity for a double stranded ribonucleic acid containing the nucleic acid sequence corresponding to the nucleic acid sequence-selective binding portion than for a double stranded deoxyribonucleic acid containing a deoxyribonucleic acid sequence corresponding to the nucleic acid sequence-selective binding portion.

12. The method according to claim 5, wherein the at least one 2-aminopyridine selectively binds to guanosine nucleobases of the nucleic acid sequence.

13. The method according to claim 5, wherein the at least one basic amino acid comprises lysine.