Methods to increase or decrease bone density

• US 20030229041A1
• Filed: 02/28/2003
• Published: 12/11/2003
• Est. Priority Date: 03/01/2002
• Status: Active Grant

• Current Assignee: Celltech Group Limited (UCB SA)
• Original Assignee: Celltech Group Limited (UCB SA)
• Inventors: Geoghegan, James C., Latham, John, Yu, Changpu, Sutherland, May S. Kung
• Granted Patent: US 7,332,276 B2
• Time in Patent Office: Days
• US Class Current: 514/44
• CPC Class Codes: A61K 31/557 Eicosanoids, e.g. leukotrienes or prostaglandins
• CPC Class Codes: A61K 31/573 substituted in position 21, e.g. cortisone, dexamethasone, prednisone or aldosterone
• CPC Class Codes: A61K 38/40 Transferrins, e.g. lactoferrins, ovotransferrins
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 19/08 for bone diseases, e.g. rachitism, Paget's disease
• CPC Class Codes: A61P 19/10 for osteoporosis
• 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 2310/14 interfering N.A.
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 1/6897 involving reporter genes operably linked to promoters
• CPC Class Codes: C12Q 2600/136 Screening for pharmacological compounds
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: G01N 2333/51 Bone morphogenetic factor; Osteogenins; Osteogenic factor; Bone-inducing factor
• CPC Class Codes: G01N 33/6893 related to diseases not provided for elsewhere

Abstract

The SOST gene gives rise to sclerostin, a protein that leads to apoptosis of bone progenitor cells. The invention provides antagonists to the sclerostin protein, and methods for identifying new sclerostin antagonists. The invention also provides molecules that can depress expression of the SOST gene, as well as methods for identifying such molecules. Such molecules and antagonists are useful for increasing bone mineralization in mammals, for example, in the treatment of osteoporosis.

Claims

1. A pharmaceutical composition comprising a carrier and a compound of the formula I:

2. The pharmaceutical composition of claim 1, wherein R₁, R₂, R₃, R₄, R₅, R₆, R₇, and R₈ groups are each separately carbonyl, hydroxy, hydrogen, or lower hydroxy alkyl.

3. The pharmaceutical composition of claim 1, wherein R₁ is carbonyl or hydroxy.

4. The pharmaceutical composition of claim 1, wherein R₈ is fluorine, hydrogen or halo.

5. A pharmaceutical composition for decreasing SOST expression in a mammal comprising a therapeutically effective amount of a glucocorticoid.

6. The pharmaceutical composition of claim 5, wherein the glucocorticoid can bind to the glucocorticoid receptor.

7. The pharmaceutical composition of claim 5, wherein the glucocorticoid is fluocinolone acetonide, triamcinolone or dexamethasone.

8. A pharmaceutical composition comprising a carrier and a prostaglandin comprising formula II:

9. The pharmaceutical composition of claim 8, wherein R₁ and R₄ are each separately carbonyl, hydroxy, hydrogen, or lower hydroxy alkyl.

10. The pharmaceutical composition of claim 8, wherein the prostaglandin is prostaglandin E2.

11. A pharmaceutical composition comprising a carrier and a bile salt of formula III:

12. The pharmaceutical composition of claim 11, wherein R₁, R₂, R₃, R₄, R₅, R₆, R₇ and R₉ are each separately carbonyl, hydroxy, hydrogen, or lower hydroxy alkyl.

13. The pharmaceutical composition of claim 11, wherein the bile salt is ursodeoxycholic acid.

14. A pharmaceutical composition comprising a carrier and a siRNA comprising SEQ ID NO:

11, SEQ ID NO;

14, SEQ ID NO;

17, SEQ ID NO;

20, SEQ ID NO;

23, SEQ ID NO;

26, SEQ ID NO;

29, SEQ ID NO;

32, SEQ ID NO;

35, SEQ ID NO;

38, SEQ ID NO;

41 or SEQ ID NO;

44, wherein the siRNA can modulate SOST expression.

15. A pharmaceutical composition comprising a carrier and a siRNA that is selectively hybridizable under stringent conditions to an RNA derived from a DNA comprising SEQ ID NO:

9, SEQ ID NO;

12, SEQ ID NO;

15, SEQ ID NO;

18, SEQ ID NO;

21, SEQ ID NO;

24, SEQ ID NO;

27, SEQ ID NO;

30, SEQ ID NO;

33, SEQ ID NO;

36, SEQ ID NO;

39, or SEQ ID NO;

42, wherein the siRNA can modulate SOST expression.

16. A method for modulating SOST expression in a mammal that comprises administering to the mammal ursodeoxycholic acid, fluocinolone acetonide, triamcinolone, prostaglandin E2 or dexamethasone.

17. A method for modulating SOST expression in a mammal that comprises administering to the mammal a compound of formula I:

18. A method for modulating SOST expression in a mammal that comprises administering to the mammal a compound of formula II:

19. A method for modulating SOST expression in a mammal that comprises administering to the mammal a compound of formula III:

20. A method for modulating SOST expression in a mammal that comprises administering to the mammal a glucocorticoid.

21. The method of claim 20, wherein the glucocorticoid is cortisol or dexamethasone.

22. A method for modulating SOST expression in a mammal that comprises administering to the mammal a siRNA comprising SEQ ID NO:

11, SEQ ID NO;

14, SEQ ID NO;

17, SEQ ID NO;

20, SEQ ID NO;

23, SEQ ID NO;

26, SEQ ID NO;

29, SEQ ID NO;

32, SEQ ID NO;

35, SEQ ID NO;

38, SEQ ID NO;

41 or SEQ ID NO;

44, wherein the siRNA can modulate SOST expression.

23. A method for modulating SOST expression in a mammal that comprises administering to the mammal a siRNA that is selectively hybridizable under stringent conditions to an RNA derived from a DNA comprising SEQ ID NO:

9, SEQ ID NO;

12, SEQ ID NO;

15, SEQ ID NO;

18, SEQ ID NO;

21, SEQ ID NO;

24, SEQ ID NO;

27, SEQ ID NO;

30, SEQ ID NO;

33, SEQ ID NO;

36, SEQ ID NO;

39, or SEQ ID NO;

42, wherein the siRNA can modulate SOST expression.

24. A method for increasing bone density in a mammal that comprises administering ursodeoxycholic acid, fluocinolone acetonide, triamcinolone, prostaglandin E2 or dexamethasone to the mammal.

25. A method for increasing bone density in a mammal that comprises administering to the mammal a compound of formula I:

26. A method for increasing bone density in a mammal that comprises administering to the mammal a compound of formula II:

27. A method for increasing bone density in a mammal that comprises administering to the mammal a compound of formula III:

28. A method for increasing bone density in a mammal that comprises administering to the mammal a glucocorticoid, wherein the glucocorticoid modulates SOST expression.

29. The method of claim 28, wherein the glucocorticoid is cortisol or dexamethasone.

30. A method for increasing bone density in a mammal that comprises administering to the mammal a siRNA comprising SEQ ID NO:

11, SEQ ID NO;

14, SEQ ID NO;

17, SEQ ID NO;

20, SEQ ID NO;

23, SEQ ID NO;

26, SEQ ID NO;

29, SEQ ID NO;

32, SEQ ID NO;

35, SEQ ID NO;

38, SEQ ID NO;

41 or SEQ ID NO;

44, wherein the siRNA can modulate SOST expression.

31. A method for increasing bone density in a mammal that comprises administering to the mammal a siRNA that is selectively hybridizable under stringent conditions to an RNA derived from a DNA comprising SEQ ID NO:

9, SEQ ID NO;

12, SEQ ID NO;

15, SEQ ID NO;

18, SEQ ID NO;

21, SEQ ID NO;

24, SEQ ID NO;

27, SEQ ID NO;

30, SEQ ID NO;

33, SEQ ID NO;

36, SEQ ID NO;

39, or SEQ ID NO;

42, wherein the siRNA can modulate SOST expression.

32. A method for decreasing apoptosis of bone cells in a mammal that comprises administering ursodeoxycholic acid, fluocinolone acetonide, triamcinolone, prostaglandin E2 or dexamethasone to the mammal.

33. A method for decreasing apoptosis of bone cells in a mammal that comprises administering to the mammal a compound of formula I:

34. A method for decreasing apoptosis of bone cells in a mammal that comprises administering to the mammal a compound of formula II:

35. A method for decreasing apoptosis of bone cells in a mammal that comprises administering to the mammal a compound of formula III:

36. A method for decreasing apoptosis of bone cells in a mammal that comprises administering to the mammal a glucocorticoid, wherein the glucocorticoid modulates SOST expression.

37. The method of claim 36, wherein the glucocorticoid is cortisol or dexamethasone.

38. A method for decreasing apoptosis of bone cells in a mammal that comprises administering to the mammal a siRNA comprising SEQ ID NO:

11, SEQ ID NO;

14, SEQ ID NO;

17, SEQ ID NO;

20, SEQ ID NO;

23, SEQ ID NO;

26, SEQ ID NO;

29, SEQ ID NO;

32, SEQ ID NO;

35, SEQ ID NO;

38, SEQ ID NO;

41 or SEQ ID NO;

44, wherein the siRNA can modulate SOST expression.

39. A method for decreasing apoptosis of bone cells in a mammal that comprises administering to the mammal a siRNA that is selectively hybridizable under stringent conditions to an RNA derived from a DNA comprising SEQ ID NO:

9, SEQ ID NO;

12, SEQ ID NO;

15, SEQ ID NO;

18, SEQ ID NO;

21, SEQ ID NO;

24, SEQ ID NO;

27, SEQ ID NO;

30, SEQ ID NO;

33, SEQ ID NO;

36, SEQ ID NO;

39, or SEQ ID NO;

42, wherein the siRNA can modulate SOST expression.

40. A method for identifying a factor that decreases SOST expression comprising:

(1) providing a cell that comprises a nucleic acid construct comprising a SOST promoter comprising SEQ ID NO;

4 that is operably linked to a nucleic acid encoding a detectable marker;

(2) contacting the cell with a test agent; and

(3) detecting whether the test agent decreases expression of the detectable marker relative to a control that comprises a cell that has not been contacted with a test agent.

41. The method of claim 40, wherein the cell is responsive to a bone morphogenetic protein.

42. The method of claim 40, wherein the cell is a C2C12 cell from a cell line having ATCC Deposit No. CRL-1772 or a C3H10T1/2 cell from a cell line having ATCC Deposit No. CCL-226.

43. The method of claim 40, wherein the cell is a human mesenchymal (hMSC) cell.

44. The method of claim 40, wherein the nucleic acid construct further comprises a nucleic acid segment encoding an osteoblastic-specific steroid response element.

45. The method of claim 40, wherein the nucleic acid construct further comprises a nucleic acid segment encoding glucocorticoid response element.

46. A method for identifying a factor that decreases SOST expression comprising:

(1) contacting a cell with a test agent; and

(2) detecting whether endogenous SOST expression decreases, wherein the cell comprises a nucleic acid encoding a protein comprising SEQ ID NO;

3 or SEQ ID NO;

6 or SEQ ID NO;

8.

47. The method of claim 46, wherein the nucleic acid comprises SEQ ID NO:

1 or SEQ ID NO;

2 or SEQ ID NO;

5 or SEQ ID NO;

7.

48. A method for identifying an antagonist for sclerostin comprising:

(1) contacting a cell with sclerostin and a test agent; and

(2) detecting whether the test agent prevents apoptosis of the cell.

49. A method for identifying a molecule that binds to sclerostin comprising:

(1) contacting a test agent with a sclerostin polypeptide comprising SEQ ID NO;

3 or SEQ ID NO;

6 or SEQ ID NO;

8; and

(2) determining whether the test agent binds to the polypeptide.

50. A marker for osteoblast differentiation comprising a sclerostin polypeptide comprising SEQ ID NO:

3 or SEQ ID NO;

6 or SEQ ID NO;

8.

51. A marker for osteoblast differentiation comprising a nucleic acid comprising SEQ ID NO:

1 or SEQ ID NO;

2 or SEQ ID NO;

5 or SEQ ID NO;

7.