METHODS AND COMPOSITIONS CONCERNING siRNA'S AS MEDIATORS OF RNA INTERFERENCE

• US 20100159591A1
• Filed: 06/15/2009
• Published: 06/24/2010
• Est. Priority Date: 12/23/2004
• Status: Active Grant

• Current Assignee: Applied Biosystems LLC (Thermo Fisher Scientific Incorporated)
• Original Assignee: Life Technologies Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: Ford, Lance P., Krebs, Joseph
• Granted Patent: US 8,058,255 B2
• Time in Patent Office: Days
• US Class Current: 435/375
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/1137 against enzymes viral enzymes C12N15/1131; receptors C12N15/1138
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2330/30 chemically synthesised
• CPC Class Codes: C12Y 102/01012 Glyceraldehyde-3-phosphate dehydrogenase (phosphorylating) (1.2.1.12)
• CPC Class Codes: C12Y 502/01008 Peptidylprolyl isomerase (5.2.1.8), i.e. cyclophilin

Abstract

The present invention concerns an isolated siRNA of from about 5 to about 20 nucleotides that mediates RNA interference. Also disclosed are methods of reducing expression of a target gene in a cell comprising obtaining at least one siRNA of 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 basepairs in length; and delivering the siRNA into the cell. The siRNAs can be chemically synthesized RNA or an analog of a naturally occurring RNA.

Claims

1. An isolated RNA of from about 5 to about 20 nucleotides that mediates RNA interference of a target mRNA.

2. The isolated RNA of claim 1 further comprising a terminal 3′

hydroxyl group.

3. The isolated RNA of claim 1 which is chemically synthesized RNA or an analog of a naturally occurring RNA.

4. The isolated RNA of claim 1, wherein the RNA is from about 12 to about 18 nucleotides in length.

5. The isolated RNA of claim 1, wherein the RNA is from about 14 to about 16 nucleotides in length.

6. The isolated RNA of claim 1, wherein the RNA is 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, or 19 nucleotides in length.

7. The isolated RNA of claim 1, wherein the RNA is an siRNA.

8. The isolated RNA of claim 1, wherein the RNA is a single-stranded.

9. The isolated RNA of claim 1, wherein the RNA is a double-stranded.

10. The isolated RNA of claim 9, wherein the 3′

ends of the double stranded RNA comprises a 2, 3, 4, or 5 nucleotide overhang.

11. The isolated RNA of claim 10, wherein the nucleotide overhang is a 2 nucleotide overhang.

12. The isolated RNA of claim 11, wherein the 2 nucleotides are thymine.

13. The isolated RNA of claim 9, wherein the 5′

ends of the double stranded RNA comprises a 2, 3, 4, or 5 nucleotide overhang.

14. The isolated RNA of claim 13, wherein the nucleotide overhang is a 2 nucleotide overhang.

15. The isolated RNA of claim 14, wherein the 2 nucleotides are thymine.

16. The isolated RNA of claim 1, wherein the RNA is formulated into a pharmaceutically acceptable composition.

17. The isolated RNA of claim 1, wherein the RNA associates with a protein complex.

18. The isolated RNA of claim 17, wherein the protein complex is RNA-induced silencing complex (RISC).

19. The isolated RNA of claim 1, wherein the isolated RNA comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:

1, SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

6, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, SEQ ID NO;

12, SEQ ID NO;

13;

SEQ ID NO;

14, SEQ ID NO;

15, SEQ ID NO;

16, SEQ ID NO;

17;

SEQ ID NO;

18, SEQ ID NO;

19, SEQ ID NO;

20, SEQ ID NO;

21, SEQ ID NO;

22, SEQ ID NO;

23, SEQ ID NO;

24, SEQ ID NO;

25, SEQ ID NO;

26, SEQ ID NO;

27, SEQ ID NO;

28, SEQ ID NO;

29, SEQ ID NO;

30;

SEQ ID NO;

31, SEQ ID NO;

32, SEQ ID NO;

33, SEQ ID NO;

34, SEQ ID NO 35, SEQ ID NO;

36;

SEQ ID NO;

37, SEQ ID NO;

38, SEQ ID NO;

39, SEQ ID NO;

40, SEQ ID NO;

41, SEQ ID NO;

42, SEQ ID NO;

43, SEQ ID NO;

44, SEQ ID NO;

45, SEQ ID NO;

46, SEQ ID NO;

47, SEQ ID NO;

48, SEQ ID NO;

49, SEQ ID NO;

50, SEQ ID NO;

51, SEQ ID NO;

52, SEQ ID NO;

53, and SEQ ID NO;

54.

20. A method of reducing expression of a target gene in a cell comprising:

a) obtaining at least one siRNA of 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides in length; and

b) delivering the siRNA into the cell.