Compositions and methods for growth factor modulation

Patent metadata

Field	Value
Publication number	US-10981981-B2
Application number	US-201816209078-A
Country	US
Kind code	B2
Filing date	Dec 4, 2018
Priority date	May 6, 2013
Publication date	Apr 20, 2021
Grant date	Apr 20, 2021

Abstract

Provided herein are proteins, antibodies, assays and methods useful for modulating growth factor levels and/or activities. In some embodiments, such growth factors are members of the TGF-β superfamily of proteins.

First claim

The invention claimed is:

1. A method of making a pharmaceutical composition comprising an antibody, or an antigen-binding fragment thereof, that binds to pro/latent GDF8 and inhibits GDF8 signaling, the method comprising the steps of:
(i) providing a panel of antibodies, or antigen-binding fragments thereof;
(ii) screening for an antibody, or an antigen-binding fragment thereof, that binds to pro/latent GDF8 but does not bind to mature GDF8, wherein pro/latent GDF8 comprises a complex comprising a latency associated peptide (LAP) domain associated with mature GDF8;
(iii) screening the antibody, or the antigen-binding fragment thereof, that binds to pro/latent GDF8 but does not bind to mature GDF8 in step (ii) for inhibitors of GDF8 signaling; and
(iv) formulating the antibody, or the antigen-binding fragment thereof, that inhibits GDF8 signaling in step (iii) into a pharmaceutical composition that further comprises a pharmaceutically acceptable excipient.

2. The method of claim 1 , wherein the panel of antibodies is an antibody library.

3. The method of claim 2 , wherein the antibody library is a phage display library.

4. The method of claim 1 , further comprising a step of subjecting the antibody, or the antigen-binding fragment thereof, to humanization and/or affinity maturation.

5. The method of claim 1 , wherein step (iii) comprises screening for an antibody, or an antigen-binding fragment thereof, that inhibits release of mature GDF8 growth factor from the pro/latent GDF8.

6. The method of claim 1 , wherein step (ii) comprises conducting a negative selection to exclude an antibody, or an antigen-binding fragment thereof, that binds to an antigen selected from GDF8 prodomain, mature GDF8 growth factor, murine proGDF8, proGDF11, proTGFβ1, and a protein comprising an amino acid sequence selected from any one of SEQ ID NOs: 207-230.

7. The method of claim 1 , wherein step (ii) comprises a binding assay selected from an enzyme-linked immunosorbent assay, a surface plasmon resonance assay, and a flow cytometry assay.

8. The method of claim 1 , further comprising a step of introducing at least one amino acid substitution to the antibody, or the antigen-binding fragment thereof, wherein the antibody, or the antigen-binding fragment thereof, comprises at least one CDR amino acid sequence with at least one amino acid deletion, addition, or substitution relative to a corresponding CDR amino acid sequence in the antibody or the antigen-binding fragment resulting from step (ii) or step (iii).

9. The method of claim 1 , wherein the antibody, or the antigen-binding fragment thereof, is an isotype selected from IgG1, IgG2, IgG3, IgG4, IgA, IgA2, IgD, IgE, or IgM.

10. The method of claim 1 , further comprising a step of incorporating a second antibody or antigen-binding fragment thereof, wherein the resultant antibody, or the antigen-binding fragment thereof, is a bispecific antibody, or an antigen-binding fragment thereof.

11. The method of claim 1 , further comprising a step of immunizing a host with an antigen comprising the pro/latent GDF8.

12. The method of claim 1 , wherein the antibody, or the antigen-binding fragment thereof, is a fully human or humanized antibody, or an antigen-binding fragment thereof.

13. The method of claim 1 , wherein the antibody, or the antigen-binding fragment thereof, is a stabilizing antibody, or an antigen-binding fragment thereof.

14. The method of claim 1 , wherein the antibody, or the antigen-binding fragment thereof, inhibits GDF8 signaling in step (iii) by:
inhibiting cleavage of a GDF8 prodomain;
inhibiting release of mature GDF8 growth factor from pro/latent GDF8; and/or
inhibiting activation of pro/latent GDF8 to mature GDF8.

15. The method of claim 14 , wherein the cleavage, release, and/or activation comprises cleavage of a GDF8 prodomain by one or more enzymes.

16. The method of claim 15 , wherein the one or more enzymes comprise a BMP/tolloid proteinase.

17. The method of claim 15 , wherein the one or more enzymes are selected from the group consisting of: furin, bone morphogenetic protein-1 (BMP-1), mammalian tolloid protein (mTLD), mammalian tolloid-like 1 (mTLL1), and mammalian tolloid-like 2 (mTLL2).

18. The method of claim 15 , wherein the activation comprises sequential enzymatic cleavage with at least two enzymes.

19. The method of claim 15 , wherein the cleavage of the GDF8 prodomain comprises BMP/tolloid proteolytic cleavage between residues Arg 75 and Asp 76 of the GDF8 prodomain.

20. The method of claim 1 , wherein step (iii) comprises identifying an antibody, or an antigen-binding fragment thereof, that inhibits BMP/tolloid proteolytic cleavage between residues Arg 75 and Asp 76 of a GDF8 prodomain.

21. The method of claim 1 , further comprising the steps of contacting the antibody, or the antigen-binding fragment thereof, resulting from step (ii) or step (iii) with pro/latent GDF8 and a BMP/tolloid proteinase; and confirming that the antibody, or the antigen-binding fragment thereof, blocks cleavage of the pro/latent GDF8 by the BMP/tolloid proteinase.

22. The method of claim 21 , wherein the BMP/tolloid proteinase is selected from the group consisting of: furin, bone morphogenetic protein-1 (BMP-1), mammalian tolloid protein (mTLD), mammalian tolloid-like 1 (mTLL1), and mammalian tolloid-like 2 (mTLL2).

23. A method of screening for an antibody, or an antigen-binding fragment thereof, that binds to pro/latent GDF8 and inhibits GDF8 signaling, the method comprising the steps of:
(i) providing a panel of antibodies;
(ii) screening for an antibody, or an antigen-binding fragment thereof, that binds to pro/latent GDF8 but does not bind to mature GDF8, wherein pro/latent GDF8 comprises a complex comprising a latency associated peptide (LAP) domain associated with mature GDF8; and
(iii) screening the antibody, or the antigen-binding fragment thereof, that binds to pro/latent GDF8 but does not bind to mature GDF8 in step (ii) for inhibition of GDF8 signaling, wherein the antibody, or the antigen-binding fragment thereof, inhibits GDF8 signaling by:
inhibiting cleavage of a GDF8 prodomain;
inhibiting release of mature GDF8 growth factor from pro/latent GDF8; and/or
inhibiting activation of pro/latent GDF8 to mature GDF8.

24. The method of claim 23 , further comprising a step of formulating the antibody, or the antigen-binding fragment thereof, that inhibits GDF8 signaling in step (iii) into a pharmaceutical composition that further comprises a pharmaceutically acceptable excipient.

Assignees

• Scholar Rock Inc

Inventors

• Schurpf Thomas
• Mahanthappa Nagesh K
• Straub Michelle

Classifications

• C07K2317/76

  Antagonist effect on antigen, e.g. neutralization or inhibition of binding · CPC title

• C07K2317/33

  Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity · CPC title

• C07K2317/32

  specific for a neo-epitope on a complex, e.g. antibody-antigen or ligand-receptor · CPC title

• C07K16/2863

  against receptors for growth factors, growth regulators · CPC title

• C07K16/22 Primary

  against growth factors {; against growth regulators} · CPC title