TREATMENT OF LIPID TRANSPORT AND METABOLISM GENE RELATED DISEASES BY INHIBITION OF NATURAL ANTISENSE TRANSCRIPT TO A LIPID TRANSPORT AND METABOLISM GENE

• US 20120046344A1
• Filed: 05/06/2010
• Published: 02/23/2012
• Est. Priority Date: 05/06/2009
• Status: Active Grant

• Current Assignee: CURNA, Inc. (Opko Health, Inc.)
• Original Assignee: CURNA, Inc. (Opko Health, Inc.)
• Inventors: Collard, Joseph, Khorkova Sherman, Olga
• Granted Patent: US 9,155,754 B2
• Time in Patent Office: Days
• US Class Current: 514/44.A
• CPC Class Codes: A61K 31/713 Double-stranded nucleic acids or oligonucleotides
• CPC Class Codes: A61K 48/00 Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
• CPC Class Codes: A61P 1/04 for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 11/06 Antiasthmatics
• CPC Class Codes: A61P 13/10 of the bladder
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/06 Antipsoriatics
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• 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 25/30 for treating abuse or dependence
• CPC Class Codes: A61P 27/06 Antiglaucoma agents or miotics
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 3/04 Anorexiants; Antiobesity agents
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 3/10 : for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/00 : Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/04 : Antibacterial agents
• CPC Class Codes: A61P 33/00 : Antiparasitic agents
• CPC Class Codes: A61P 35/00 : Antineoplastic agents
• CPC Class Codes: A61P 37/00 : Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 37/02 : Immunomodulators
• CPC Class Codes: A61P 37/06 : Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 37/08 : Antiallergic agents antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14
• CPC Class Codes: A61P 43/00 : Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/14 : of the thyroid hormones, e.g. T3, T4
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• 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 15/113 : 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 15/1137 : against enzymes viral enzymes C12N15/1131; receptors C12N15/1138
• CPC Class Codes: C12N 15/1138 : against receptors or cell surface proteins
• CPC Class Codes: C12N 2310/11 : Antisense
• CPC Class Codes: C12N 2310/14 : interfering N.A.
• CPC Class Codes: C12N 2310/313 : Phosphorodithioates
• CPC Class Codes: C12N 2310/314 : Phosphoramidates
• CPC Class Codes: C12N 2310/315 : Phosphorothioates
• CPC Class Codes: C12N 2310/321 : 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 : 2'-R Modification
• CPC Class Codes: C12Y 203/01043 : Phosphatidylcholine-sterol O-acyltransferase (2.3.1.43), i.e. lecithin-cholesterol acyltransferase or LCAT

Abstract

The present invention relates to antisense oligonucleotides that modulate the expression of and/or function of a Lipid transport and metabolism gene, in particular, by targeting natural antisense polynucleotides of a Lipid transport and metabolism gene. The invention also relates to the identification of these antisense oligonucleotides and their use in treating diseases and disorders associated with the expression of a Lipid transport and metabolism genes.

Claims

1. A method of modulating a function of and/or the expression of a Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro comprising:

contacting said cells or tissues with at least one antisense oligonucleotide 5 to 30 nucleotides in length wherein said at least one oligonucleotide has at least 50% sequence identity to a reverse complement of a polynucleotide comprising 5 to 30 consecutive nucleotides within nucleotides 1 to 1299 of SEQ ID NO;

8, 1 to 918 of SEQ ID NO;

9, 1 to 1550 of SEQ ID NO;

10, 1 to 329 of SEQ ID NO;

11, 1 to 1826 of SEQ ID NO;

12, 1 to 536 of SEQ ID NO;

13, 1 to 551 of SEQ ID NO;

14, 1 to 672 of SEQ ID NO;

15, 1 to 616 of SEQ ID NO;

16, 1 to 471 of SEQ ID NO;

17, 1 to 707 of SEQ ID NO;

18, 1 to 741 of SEQ ID NO;

19, 1 to 346 of SEQ ID NO;

20, 1 to 867 of SEQ ID NO;

21, 1 to 563 of SEQ ID NO;

22 (FIG. 3);

thereby modulating a function of and/or the expression of the Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro.

13. The method of claim 1, wherein the at least one oligonucleotide comprises at least one oligonucleotide sequences set forth as SEQ ID NOS:

23 to 263.

2. A method of modulating a function of and/or the expression of a Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro comprising:

contacting said cells or tissues with at least one antisense oligonucleotide 5 to 30 nucleotides in length wherein said at least one oligonucleotide has at least 50% sequence identity to a reverse complement of a natural antisense of a Lipid transport and metabolism gene polynucleotide;

thereby modulating a function of and/or the expression of the Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro.

3. A method of modulating a function of and/or the expression of a Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro comprising:

contacting said cells or tissues with at least one antisense oligonucleotide 5 to 30 nucleotides in length wherein said oligonucleotide has at least 50% sequence identity to an antisense oligonucleotide to the Lipid transport and metabolism gene polynucleotide;

thereby modulating a function of and/or the expression of the Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro.

4. A method of modulating a function of and/or the expression of a Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro comprising:

contacting said cells or tissues with at least one antisense oligonucleotide that targets a region of a natural antisense oligonucleotide of the Lipid transport and metabolism gene polynucleotide;

thereby modulating a function of and/or the expression of the Lipid transport and metabolism gene polynucleotide in patient cells or tissues in vivo or in vitro.

5. The method of claim 4, wherein a function of and/or the expression of the Lipid transport and metabolism gene is increased in vivo or in vitro with respect to a control.

6. The method of claim 4, wherein the at least one antisense oligonucleotide targets a natural antisense sequence of a Lipid transport and metabolism gene polynucleotide.

7. The method of claim 4, wherein the at least one antisense oligonucleotide targets a nucleic acid sequence comprising coding and/or non-coding nucleic acid sequences of a Lipid transport and metabolism gene polynucleotide.

8. The method of claim 4, wherein the at least one antisense oligonucleotide targets overlapping and/or non-overlapping sequences of a Lipid transport and metabolism gene polynucleotide.

9. The method of claim 4, wherein the at least one antisense oligonucleotide comprises one or more modifications selected from:

at least one modified sugar moiety, at least one modified internucleoside linkage, at least one modified nucleotide, and combinations thereof.

10. The method of claim 9, wherein the one or more modifications comprise at least one modified sugar moiety selected from:

a 2′

-O-methoxyethyl modified sugar moiety, a 2′

-methoxy modified sugar moiety, a 2′

-O-alkyl modified sugar moiety, a bicyclic sugar moiety, and combinations thereof.

11. The method of claim 9, wherein the one or more modifications comprise at least one modified internucleoside linkage selected from:

a phosphorothioate, 2′

-Omethoxyethyl (MOE), 2′

-fluoro, alkylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, carboxymethyl ester, and combinations thereof.

12. The method of claim 9, wherein the one or more modifications comprise at least one modified nucleotide selected from:

a peptide nucleic acid (PNA), a locked nucleic acid (LNA), an arabino-nucleic acid (FANA), an analogue, a derivative, and combinations thereof.

14. A method of modulating a function of and/or the expression of a Lipid transport and metabolism gene in mammalian cells or tissues in vivo or in vitro comprising:

contacting said cells or tissues with at least one short interfering RNA (siRNA) oligonucleotide 5 to 30 nucleotides in length, said at least one siRNA oligonucleotide being specific for an antisense polynucleotide of a Lipid transport and metabolism gene polynucleotide, wherein said at least one siRNA oligonucleotide has at least 50% sequence identity to a complementary sequence of at least about five consecutive nucleic acids of the antisense and/or sense nucleic acid molecule of the Lipid transport and metabolism gene polynucleotide; and

, modulating a function of and/or the expression of a Lipid transport and metabolism gene in mammalian cells or tissues in vivo or in vitro.

15. The method of claim 14, wherein said oligonucleotide has at least 80% sequence identity to a sequence of at least about five consecutive nucleic acids that is complementary to the antisense and/or sense nucleic acid molecule of the Lipid transport and metabolism gene polynucleotide.

16. A method of modulating a function of and/or the expression of a Lipid transport and metabolism gene in mammalian cells or tissues in vivo or in vitro comprising:

contacting said cells or tissues with at least one antisense oligonucleotide of about 5 to 30 nucleotides in length specific for noncoding and/or coding sequences of a sense and/or natural antisense strand of a Lipid transport and metabolism gene polynucleotide wherein said at least one antisense oligonucleotide has at least 50% sequence identity to at least one nucleic acid sequence set forth as SEQ ID NOS;

1 to 7 and 8 to 22; and

, modulating the function and/or expression of the Lipid transport and metabolism gene in mammalian cells or tissues in vivo or in vitro.

17. A synthetic, modified oligonucleotide comprising at least one modification wherein the at least one modification is selected from:

at least one modified sugar moiety;

at least one modified internucleotide linkage;

at least one modified nucleotide, and combinations thereof;

wherein said oligonucleotide is an antisense compound which hybridizes to and modulates the function and/or expression of a Lipid transport and metabolism gene in vivo or in vitro as compared to a normal control.

18. The oligonucleotide of claim 17, wherein the at least one modification comprises an internucleotide linkage selected from the group consisting of phosphorothioate, alkylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, carboxymethyl ester, and combinations thereof.

19. The oligonucleotide of claim 17, wherein said oligonucleotide comprises at least one phosphorothioate internucleotide linkage.

20. The oligonucleotide of claim 17, wherein said oligonucleotide comprises a backbone of phosphorothioate internucleotide linkages.

21. The oligonucleotide of claim 17, wherein the oligonucleotide comprises at least one modified nucleotide, said modified nucleotide selected from:

a peptide nucleic acid, a locked nucleic acid (LNA), analogue, derivative, and a combination thereof.

22. The oligonucleotide of claim 17, wherein the oligonucleotide comprises a plurality of modifications, wherein said modifications comprise modified nucleotides selected from:

phosphorothioate, alkylphosphonate, phosphorodithioate, alkylphosphonothioate, phosphoramidate, carbamate, carbonate, phosphate triester, acetamidate, carboxymethyl ester, and a combination thereof.

23. The oligonucleotide of claim 17, wherein the oligonucleotide comprises a plurality of modifications, wherein said modifications comprise modified nucleotides selected from:

peptide nucleic acids, locked nucleic acids (LNA), analogues, derivatives, and a combination thereof.

24. The oligonucleotide of claim 17, wherein the oligonucleotide comprises at least one modified sugar moiety selected from:

a 2′

-O-methoxyethyl modified sugar moiety, a 2′

-methoxy modified sugar moiety, a 2′

-O-alkyl modified sugar moiety, a bicyclic sugar moiety, and a combination thereof.

25. The oligonucleotide of claim 17, wherein the oligonucleotide comprises a plurality of modifications, wherein said modifications comprise modified sugar moieties selected from:

a 2′

-O-methoxyethyl modified sugar moiety, a 2′

-methoxy modified sugar moiety, a 2′

-O-alkyl modified sugar moiety, a bicyclic sugar moiety, and a combination thereof.

26. The oligonucleotide of claim 17, wherein the oligonucleotide is of at least about 5 to 30 nucleotides in length and hybridizes to an antisense and/or sense strand of a Lipid transport and metabolism gene polynucleotide wherein said oligonucleotide has at least about 20% sequence identity to a complementary sequence of at least about five consecutive nucleic acids of the antisense and/or sense coding and/or noncoding nucleic acid sequences of the Lipid transport and metabolism gene polynucleotide.

27. The oligonucleotide of claim 17, wherein the oligonucleotide has at least about 80% sequence identity to a complementary sequence of at least about five consecutive nucleic acids of the antisense and/or sense coding and/or noncoding nucleic acid sequence of the Lipid transport and metabolism gene polynucleotide.

28. The oligonucleotide of claim 17, wherein said oligonucleotide hybridizes to and modulates expression and/or function of at least one Lipid transport and metabolism gene polynucleotide in vivo or in vitro, as compared to a normal control.

29. The oligonucleotide of claim 17, wherein the oligonucleotide comprises the sequences set forth as SEQ ID NOS:

23 to 263.

30. A composition comprising one or more oligonucleotides specific for one or more Lipid transport and metabolism gene polynucleotides, said polynucleotides comprising antisense sequences, complementary sequences, alleles, homologs, isoforms, variants, derivatives, mutants, fragments, or combinations thereof.

31. The composition of claim 30, wherein the oligonucleotides have at least about 40% sequence identity as compared to any one of the nucleotide sequences set forth as SEQ ID NOS:

23 to 263.

32. The composition of claim 30, wherein the oligonucleotides comprise nucleotide sequences set forth as SEQ ID NOS:

23 to 263.

33. The composition of claim 32, wherein the oligonucleotides set forth as SEQ ID NOS:

23 to 263 comprise one or more modifications or substitutions.

34. The composition of claim 33, wherein the one or more modifications are selected from:

phosphorothioate, methylphosphonate, peptide nucleic acid, locked nucleic acid (LNA) molecules, and combinations thereof.

35. A method of preventing or treating a disease associated with at least one Lipid transport and metabolism gene polynucleotide and/or at least one encoded product thereof, comprising:

administering to a patient a therapeutically effective dose of at least one antisense oligonucleotide that binds to a natural antisense sequence of said at least one Lipid transport and metabolism gene polynucleotide and modulates expression of said at least one Lipid transport and metabolism gene polynucleotide;

thereby preventing or treating the disease associated with the at least one Lipid transport and metabolism gene polynucleotide and/or at least one encoded product thereof.

36. The method of claim 35, wherein a disease associated with the at least one Lipid transport and metabolism gene polynucleotide is selected from:

a cardiovascular disease or disorder, a metabolic disease or disorder (e.g., diabetes, obesity, dyslipidemia, hyperglycemia, hyperinsulinemia, hypercholesterolemia etc.), a disease or disorder associated with impaired lipid metabolism, a coronary artery disease, atherosclerosis, an HDL metabolism disease or disorder (e.g., familial HDL deficiency (FHD), Sea-blue histiocytosis, Tangier'"'"'s Disease, Fish-eye disease, LCAT deficiency, low-HDL cholesterolemia etc.), a disease or disorder associated with cellular cholesterol and/or phospholipid homeostasis, Familial amyloid nephropathy, a disease or disorder associated with impaired cholesterol regulation, a disease or disorder associated with a deficiency of the Lipid transport and metabolism gene transporter, Apolipoprotein A-I deficiency, a disease or disorder associated with abnormally fast or abnormally slow rate of cholesterol efflux in a cell, a disease or disorder associated with pancreatic beta cell function, diabetes, a metabolic disease or disorder, arthritis, inflammation, an autoimmune disease or disorder, acquired immune deficiency syndrome (AIDS), inflammation, a neurological disease or disorder, a neurodegenerative disease or disorder, cancer, dyslipidiemia, metabolic syndrome, a senile plaque, cerebral amyloid angiopathy, Amyloidosis, glioblastoma, a disease or disorder associated with amyloid deposition, neurofibrillary tangles, choriocarcinoma, astrocytoma, amyloidosis, hyperlipidemia, neoplastic transformation, atherosclerotic plaque, obstruction, metastasis, pulmonary fibrosis, necrosis, shock, melanoma, genetic susceptibility, psoriasis, glioma, neuropathology, a vascular disease, cell damage, Nonsmall cell lung carcinomas (NSCLCs), liposarcoma, an immunodeficiency disease or disorder, an allergy, an organ transplant rejection, glomerulonephritis, venous thrombosis, pathological processes or leukemia, a skeletal disease or disorder, a muscular disease or disorder, a disease or disorder associated with infectious organisms, an immune related disease or disorder, nerve repair and paralysis, neuroendocrine differentiation, systemic non-neuropathic amyloidosis, an amyloid disease, tumor growth dependent on angiogenesis, non-cancerous diseases with symptoms include an increase in angiogenesis, e.g., psoriasis, retinopathy of prematurity, a Choroid disease, neovascular glaucoma, diabetic retinopathy, substance abuse, impaired cognitive function, and reduced neurite outgrowth, ApoE abnormal expression, function, activity as compared to a normal control, psoriasis, a disease or disorder caused by foreign organisms such as viral, bacterial, parasitic, fungal, and the like.

37. A method of identifying and selecting at least one oligonucleotide for in vivo administration comprising:

selecting a target polynucleotide associated with a disease state;

identifying at least one oligonucleotide comprising at least five consecutive nucleotides which are complementary to the selected target polynucleotide or to a polynucleotide that is antisense to the selected target polynucleotide;

measuring the thermal melting point of a hybrid of an antisense oligonucleotide and the target polynucleotide or the polynucleotide that is antisense to the selected target polynucleotide under stringent hybridization conditions; and

selecting at least one oligonucleotide for in vivo administration based on the information obtained.