CA2510893A1 - Binding agents which inhibit myostatin - Google Patents

Binding agents which inhibit myostatin Download PDF

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CA2510893A1
CA2510893A1 CA002510893A CA2510893A CA2510893A1 CA 2510893 A1 CA2510893 A1 CA 2510893A1 CA 002510893 A CA002510893 A CA 002510893A CA 2510893 A CA2510893 A CA 2510893A CA 2510893 A1 CA2510893 A1 CA 2510893A1
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amino acid
peptide
neutral
absent
myostatin
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CA002510893A
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CA2510893C (en
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Hq Han
Hosung Min
Thomas Charles Boone
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Amgen Inc
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Amgen, Inc.
Hq Han
Hosung Min
Thomas Charles Boone
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Priority to CA2774928A priority Critical patent/CA2774928A1/en
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    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/566—Immunoassay; Biospecific binding assay; Materials therefor using specific carrier or receptor proteins as ligand binding reagents where possible specific carrier or receptor proteins are classified with their target compounds
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Abstract

The present invention provides binding agents comprising peptides capable of binding myostatin and inhibiting its activity. In one embodiment the binding agent comprises at least one myostatin-binding peptide attached directly or indirectly to at least one vehicle such as a polymer or an Fc domain. The binding agents of the present invention produced increased lean muscle mass when administered to animals and decreased fat to muscle ratios. Therepeutic compositions containing the binding agents of the present invention are useful for treating muscle-wasting disorders and other metabolic disorders including diabetes and obesity.

Claims (36)

1. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the amino acid sequence WMCPP (SEQ ID NO: 633), and physiologically acceptable salts thereof.
2. The binding agent of claim 1, wherein the peptide is between 5 and 50 amino acids in length.
3. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the amino acid sequence Ca1a2Wa3WMCPP (SEQ ID NO: 352), wherein a1, a2 and a3 are selected from a neutral hydrophobic, neutral polar, or basic amino acid, and wherein the peptide is between 10 and 50 amino acids in length, and physiologically acceptable salts thereof.
4. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence Cb1b2Wb3WMCPP (SEQ ID NO: 353), wherein b1 is selected from any one of the amino acids T, I, or R;
b2 is selected from any one of R, S, Q;
b3 is selected from any one of P, R and Q, and wherein the peptide is beween 10 and 50 amino acids in length, and physiologically acceptable salts thereof.
5. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence c1c2c3c4c5c6Cc7c8Wc9WMCPPc10c11c12c13 (SEQ
ID NO: 354), wherein:
c1 is absent or any amino acid;
c2 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
c3 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
c4 is absent or any amino acid;
c5 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
c6 is absent or a neutral hydrophobic, neutral polar, or basic amino acid;
c7 is a neutral hydrophobic, neutral polar, or basic amino acid;
c8 is a neutral hydrophobic, neutral polar, or basic amino acid;
c9 is a neutral hydrophobic, neutral polar or basic amino acid; and c10 to c13 is any amino acid; and wherein the peptide is between 20 and 50 amino acids in length, and physiologically acceptable salts thereof.
6. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence d1d2d3d4d5d6Cd7d8Wd9WMCPP d10d11d12d13 (SEQ ID NO: 355), wherein d1 is absent or any amino acid;
d2 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
d3 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
d4 is absent or any amino acid;
d5 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
d6 is absent or a neutral hydrophobic, neutral polar, or basic amino acid;
d7 is selected from any one of the amino acids T, I, or R;
d8 is selected from any one of R, S, Q;
d9 is selected from any one of P, R and Q, and d10 to d13 is selected from any amino acid, and wherein the peptide is between 20 and 50 amino acids in length, and physiologically acceptable salts thereof.
7. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence WYe1e2Ye3G, (SEQ ID NO: 356) wherein e1 is P, S or Y, e2 is C or Q, and e3 is G or H, and wherein the peptide is between 7 and 50 amino acids in length, and physiologically acceptable salts thereof.
8. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence f1EMLf2SLf3f4LL, (SEQ ID NO: 455), wherein f1 is M or I, f2 is any amino acid, f3 is L or F, f4 is E, Q or D;
and wherein the peptide is between 7 and 50 amino acids in length, and physiologically acceptable salts thereof.
9. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence Lg1g2LLg3g4L, (SEQ ID NO: 456), wherein g1 is Q, D or E, g2 is S, Q, D or E, g3 is any amino acid, g4 is L, W, F, or Y, and wherein the peptide is between 8 and 50 amino acids in length, and physiologically acceptable salts thereof.
10. A binding agent comprising at least one peptide capable of binding myostatin, wherein the peptide comprises the sequence h1h2h3h4h5h6h7h8h9 (SEQ ID NO:
457), wherein h1 is R or D, h2 is any amino acid, h3 is A, T S or Q, h4 is L or M, h5 is L or S, h6 is any amino acid, h7 is F or E, h8 is W, F or C, h9 is L, F, M or K, and wherein the peptide is between 9 and 50 amino acids in length, and physiologically acceptable salts thereof.
11. A binding agent wherein said agent has the structure:
(X1)a F1-(X2)b, or multimers thereof;
wherein F1 is a vehicle; and X1 and X2 are each independently selected from -(L1)c- P1;
-(L1)c-P1-(L2)d -P2;
-(LI)c-P1-(L2)d-P2-(L3)e-P3;
and -(L1)c-P1-(L2)d-P2-(L3)e -P3-(L4)f-P4;
wherein P1, P2, P3, and P4 are peptides capable of binding myostatin, and each independently comprise the amino acid sequence WMCPP (SEQ ID NO: 633);
wherein L1, L2, L3, and L4 are each linkers;
and a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a and b is 1, and physiologically acceptable salts thereof.
12. The binding agent of claim 11 wherein one or more of the myostatin binding peptides comprise the amino acid sequence Ca1a2Wa3WMCPP (SEQ ID NO: 352), wherein a1, a2 and a3 are selected from a neutral hydrophobic, neutral polar, or basic amino acid, and wherein the peptide is between 10 and 50 amino acids in length, and physiologically acceptable salts thereof.
13. The binding agent of claim 11, wherein one or more of the myostatin binding peptides each independently comprise the amino acid sequence Cb1b2Wb3WMCPP
(SEQ ID NO:
353), wherein b1 is selected from any one of the amino acids T, I, or R;
b2 is selected from any one of R, S, Q;
b3 is selected from any one of P, R and Q, and wherein the peptide is beween 10 and 50 amino acids in length, and physiologically acceptable salts thereof.
14. The binding agent of claim 11, wherein one or more of the myostatin binding peptides each independently comprise the sequence c1c2c3c4c5c6c7c8Wc9WMCPPc10c11c12c13 (SEQ ID
NO: 354), wherein:
c1 is absent or any amino acid;
c2 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
c3 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
c4 is absent or any amino acid;
c5 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
c6 is absent or a neutral hydrophobic, neutral polar, or basic amino acid;
c7 is a neutral hydrophobic, neutral polar, or basic amino acid;
c8 is a neutral hydrophobic, neutral polar, or basic amino acid;
c9 is a neutral hydrophobic, neutral polar or basic amino acid; and c10 to c13 is any amino acid; and wherein the peptide is between 20 and 50 amino acids in length, and physiologically acceptable salts thereof..
15. The binding agent of claim 11, wherein one or more of the myostatin binding peptides each independently comprise the sequence d1d2d3d4d5d6Cd7d8Wd9WMCPP
d10d11d12d13 (SEQ ID NO: 355), wherein d1 is absent or any amino acid;
d2 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
d3 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
d4 is absent or any amino acid;
d5 is absent or a neutral hydrophobic, neutral polar, or acidic amino acid;
d6 is absent or a neutral hydrophobic, neutral polar, or basic amino acid;
d7 is selected from any one of the amino acids T, I, or R;
d8 is selected from any one of R, S, Q;
d9 is selected from any one of P, R and Q, and d10 to d13 is selected from any amino acid, and wherein the peptide is between 20 and 50 amino acids in length, and physiologically acceptable salts thereof.
16. A binding agent wherein said agent has the structure:
(X1)a-F1-(X2)b, or multimers thereof;
wherein F1 is a vehicle; and X1 and X2 are each independently selected from -(L1)c- P1;
-(L1)c-P1-(L2)d -P2;
-(L1)c-P1-(L2)d-P2-(L3)e-P3;

and -(L1)c-P1-(L2)d-P2-(L3)e -P3-(L4)f-P4;
wherein P1, P2, P3, and P4 are peptides capable of binding myostatin, and wherein one or more of P1, P2, P3, and P4 each independently are selected from the group consisting of (a) a peptide comprising the amino acid sequence WYe1e2Ye3G, (SEQ ID NO: 356), wherein e1 is P, S or Y, e2 is C or Q, and e3 is G or H;
(b) a peptide comprising the amino acid sequence f1EMLf2SLf3f4LL, (SEQ ID NO:
455), wherein f1 is M or I, f2 is any amino acid, f3 is L or F, and f4 is E, Q or D;
(c) a peptide comprising the amino acid sequence Lg1g2LLg3g4L, (SEQ ID NO:
456), wherein g1 is Q, D or E, g2 is S, Q, D or E, g3 is any amino acid, and g4 is L, W, F, or Y;
and (d) a peptide comprising the amino acid sequence h1h2h3h4h5h6h7h8h9 (SEQ ID
NO: 457), wherein h1 is R or D, h2 is any amino acid, h3 is A, T S or Q, h4 is L or M, h5 is L or S, h6 is any amino acid, h7 is F or E, h8 is W, F or C, and h9 is L, F, M or K;
wherein the peptide is between 9 and 50 amino acids in length, wherein L1, L2, L3, and L4 are each linkers;
and a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a and b is 1, and physiologically acceptable salts thereof.
17. A binding agent wherein said agent has the structure:
(X1)n-F1-(X2)b, or multimers thereof;
wherein F1 is a vehicle; and X1 and X2 are each independently selected from -(L1)c- P1 -(L1)c-P1-(L2)d -P2;
-(L1)c-P1-(L2)d-P2-(L3)e-P3;
and -(L1)c-P1-(L2)d-P2-(L3)e -P3-(L4)t-P4;
wherein P1, P2, P3, and P4 are peptides capable of binding myostatin, and are independently selected from SEQ ID NO: 305 through 351 and SEQ ID NO: 357 through 454;
wherein L1, L2, L3, and L4 are each linkers;
and a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a and b is 1, and physiologically acceptable salts thereof.
18. The binding agent of any one of claims 11 through 17 , wherein the vehicle is an Fc domain.
19. A polynucleotide sequence encoding the binding agent of claim 18.
20. An expression vector comprising the polynucleotide of claim 19.
21. A host cell comprising the expression vector of claim 20.
22. The host cell of claim 21, wherein the cell is a procaryotic cell.
23. The host cell of claim 21, wherein the cell is a eucaryotic cell.
24. A pharmaceutical composition comprising an effective amount of the binding agent of any one of claims 1 or 11 in admixture with a pharmaceutically acceptable carrier thereof.
25. The pharmaceutical composition of claim 24, wherein the vehicle is an Fc domain.
26. A method of inhibiting myostatin activity in a subject comprising administering an effective amount of the binding agent of any one of claims 1, 3 or 11 to the subject.
27. A method of increasing lean muscle mass in a subject comprising administering the composition of claim 24 to the subject.
28. The method of claim 27, wherein the subject is a food animal.
29. A method of increasing the ratio of lean muscle mass to fat in a subject comprising administering a therapeutically effective amount of the composition of claim 24 to the subject.
30. A method of treating a muscle-wasting disease in a subject comprising administering a therapeutically effective amount of the composition of claim 24 to the subject.
31. The method of claim 30, wherein the disease is selected from muscular dystrophy amyotrophic lateral sclerosis, congestive obstructive pulmonary disease, chronic heart failure, cancer, AIDs, renal failure, uremia, rheumatoid arthritis, age-related sarcopenia, and muscle-wasting due to prolonged bedrest, spinal chord injury, stroke, bone fracture, and aging.
32. A method of treating a myostatin-related metabolic disorder in a subject comprising administering a therapeutically effective amount of the composition of claim 24 to the subject.
33. The method of claim 32 wherein the metabolic disorder is selected from diabetes, obesity, hyperglycemia, and bone loss.
34. A method of detecting myostatin in a sample comprising contacting the sample with a binding agent of claims 1 or 11, and detecting the bound complex.
35. A method of measuring myostatin in a sample comprising contacting the sample with a binding agent of claims 1 or 11, and measuring the bound complex.
36. A method of diagnosing a myostatin related disorder in a subject comprising contacting a sample taken from the subject with a binding agent of claims 1 or 11, and detecting the bound complex.
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