Methods and compositions involving miRNA and miRNA inhibitor molecules

• US 8,765,709 B2
• Filed: 08/28/2013
• Issued: 07/01/2014
• Est. Priority Date: 11/12/2004
• Status: Expired due to Fees

• Current Assignee: Asuragen, Inc.
• Original Assignee: Asuragen, Inc.
• Inventors: Brown, David, Ford, Lance, Cheng, Angie, Jarvis, Rich, Byrom, Mike, Ovcharenko, Dmitriy, Devroe, Eric, Kelnar, Kevin
• Primary Examiner(s): WHITEMAN, BRIAN A
• Application Number:
  US14/012,302
• Publication Number:
  US 20140031415A1
• Time in Patent Office: 307 Days
• Field of Search: 514/44.A
• US Class Current: 514/44.A
• CPC Class Codes: A61K 31/7088 Compounds having three or more nucleosides or nucleotides
• CPC Class Codes: A61K 31/7105 Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
• CPC Class Codes: A61K 31/713 Double-stranded nucleic acids or oligonucleotides
• CPC Class Codes: A61K 45/06 Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• CPC Class Codes: A61K 9/127 Liposomes
• CPC Class Codes: A61N 5/10 X-ray therapy; Gamma-ray therapy; Particle-irradiation therapy A61N5/01 takes precedence
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 15/08 for gonadal disorders or for enhancing fertility, e.g. inducers of ovulation or of spermatogenesis
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/06 for tuberculosis
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/113 : Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/1136 : against growth factors, growth regulators, cytokines, lymphokines or hormones
• CPC Class Codes: C12N 2310/14 : interfering N.A.
• CPC Class Codes: C12N 2310/141 : MicroRNAs, miRNAs
• CPC Class Codes: C12N 2310/312 : Phosphonates
• CPC Class Codes: C12N 2310/321 : 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 : 2'-R Modification
• CPC Class Codes: C12N 2310/33 : of the base
• CPC Class Codes: C12N 2310/344 : Position-specific modifications, e.g. on every purine, at the 3'-end
• CPC Class Codes: C12N 2310/35 : Nature of the modification
• CPC Class Codes: C12N 2310/351 : Conjugate
• CPC Class Codes: C12N 2310/3521 : Methyl
• CPC Class Codes: C12N 2310/3527 : Other alkyl chain
• CPC Class Codes: C12N 2310/3535 : Nitrogen
• CPC Class Codes: C12N 2310/533 : having a mismatch or nick in at least one of the strands
• CPC Class Codes: C12N 2320/12 : in functional genomics, i.e. for the determination of gene function
• CPC Class Codes: C12N 2320/30 : Special therapeutic applications
• CPC Class Codes: C12N 2320/50 : Methods for regulating/modulating their activity
• CPC Class Codes: C12N 2330/10 : naturally occurring
• CPC Class Codes: C12Q 1/68 : involving nucleic acids
• CPC Class Codes: C12Q 1/6876 : Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 2600/158 : Expression markers
• CPC Class Codes: C12Q 2600/178 : miRNA, siRNA or ncRNA

Abstract

The present invention concerns methods and compositions for introducing miRNA activity or function into cells using synthetic nucleic acid molecules. Moreover, the present invention concerns methods and compositions for identifying miRNAs with specific cellular functions that are relevant to therapeutic, diagnostic, and prognostic applications wherein synthetic miRNAs and/or miRNA inhibitors are used in library screening assays.

Claims

1. A method of decreasing lung cancer or thyroid cancer cell proliferation, comprising providing to a lung cancer or thyroid cancer cell an effective amount of an miRNA inhibitor molecule between 15 and 130 nucleotide residues in length, having a sequence that is at least 90% complementary to the sequence of nucleotides 65-87 of SEQ ID NO:

170, thereby decreasing lung cancer or thyroid cancer cell proliferation.

2. The method of claim 1 wherein the miRNA inhibitor molecule consists of a modified oligonucleotide.

3. The method of claim 1 wherein the miRNA inhibitor molecule is 17 to 25 nucleotides in length.

4. The method of claim 1 wherein the miRNA inhibitor molecule is 100% complementary to the sequence of nucleotides 65-87 of SEQ ID NO:

170.

5. The method of claim 1 wherein at least one internucleoside linkage is a modified internucleoside linkage.

6. The method of claim 5 wherein the modified internucleoside linkage is a phosphorothioate internucleoside linkage.

7. The method of claim 4 wherein at least one internucleoside linkage is a modified internucleoside linkage.

8. The method of claim 7 wherein the modified internucleoside linkage is a phosphorothioate internucleoside linkage.

9. The method of claim 1 wherein at least one nucleoside comprises a modified sugar.

10. The method of claim 9 wherein each modified sugar is independently selected from a 2′

-fluoro or a 2′

-O-methyl sugar moiety.

11. The method of claim 1 wherein each of a plurality of nucleosides comprises a modified sugar.

12. The method of claim 11 wherein each modified sugar is independently selected from a 2′

-fluoro or a 2′

O-methyl sugar moiety.

13. The method of claim 1 wherein each nucleoside comprises a modified sugar.

14. The method of claim 13 wherein each modified sugar is independently selected from a 2′

-fluoro or a 2′

-O-methyl sugar moiety.

15. The method of claim 1 wherein at least one nucleoside comprises a modified nucleobase.

16. The method of claim 15 wherein the modified nucleobase is a 5-methylcytosine.

17. The method of claim 1 wherein the cell is present in a subject and the providing compromises intravenous or subcutaneous administration.

18. The method of claim 1 wherein the subject has lung cancer.

20. The method of claim 1 wherein the subject has thyroid cancer.

19. A method of decreasing lung cancer or thyroid cancer cell proliferation, comprising providing to a lures cancer or thyroid cancer cell an effective amount of miRNA inhibitor molecule between 17 and 25 nucleotide residues in length, having a sequence that is at least 90% complementary to the sequence of nucleotides 65-87 of SEQ ID NO:

170, thereby decreasing lung cancer or thyroid cancer cell proliferation, wherein;

a) the lung cancer or thyroid cancer cell is present in a subject;

b) the providing comprises intravenous or subcutaneous administration and c) the subject has lung cancer or thyroid cancer.

21. The method of claim 19 wherein the subject has lung cancer.

22. The method of claim 19 wherein the subject has thyroid cancer.