Methods and compositions for targeting polyubiquitin

Patent metadata

Field	Value
Publication number	US-10035849-B2
Application number	US-201615153644-A
Country	US
Kind code	B2
Filing date	May 12, 2016
Priority date	Jan 18, 2008
Publication date	Jul 31, 2018
Grant date	Jul 31, 2018

Abstract

Anti-K63-linked polyubiquitin monoclonal antibodies, and methods for using the antibodies, are provided.

First claim

We claim:

1. An isolated antibody or antigen-binding fragment thereof that specifically binds to K63-linked polyubiquitin, comprising:
the HVR-H1 sequence of SEQ ID NO: 5;
the HVR-H2 sequence of SEQ ID NO: 61, optionally having one substitution selected from the following:
(i) a D at position 1,
(ii) an A at position 3,
(iii) a T at position 3, or
(iv) an L at position 6;

the HVR-H3 sequence of SEQ ID NO: 6;
the HVR-L1 sequence of SEQ ID NO: 3;
the HVR-L2 sequence of SEQ ID NO: 9, wherein:
(a) if HVR-H2 is SEQ ID NO: 61, then HVR-L2 optionally has a substitution of an R at position 4,
(b) if HVR-H2 is SEQ ID NO: 61 substituted with a D at position 1, then HVR-L2 optionally has a substitution of a A, S, or T at position 4 of SEQ ID NO: 9;
(c) if HVR-H2 is SEQ ID NO: 61 substituted with an A at position 3, then HVR-L2 optionally has a substitution of an A, L, or N at position 4 of SEQ ID NO: 9; and
(d) if HVR-H2 is SEQ ID NO: 61 substituted with a T at position 3, then HVR-L2 optionally has a substitution of a T at position 4 of SEQ ID NO: 9;

and
the HVR-L3 sequence of SEQ ID NO: 4.

2. An isolated antibody or antigen-binding fragment that binds to the same antigenic determinant on polyubiquitin as the antibody or antigen-binding fragment of claim 1 , wherein the antibody or antigen-binding fragment does not specifically bind to K48-linked polyubiquitin or monoubiquitin.

3. An isolated antibody or antigen-binding fragment that competes with the antibody of claim 1 for binding to polyubiquitin, wherein the antibody or antigen-binding fragment does not specifically bind to monoubiquitin.

4. The antibody or antigen-binding fragment of claim 1 , wherein the antibody or antigen-binding fragment specifically binds to a K63-linked polyubiquitinated protein.

5. The antibody or antigen-binding fragment of claim 4 , wherein the antibody or antigen-binding fragment has at least one property selected from inhibiting degradation of the K63-linked polyubiquitinated protein, modulating at least one polyubiquitin-mediated signaling pathway, inhibiting at least one polyubiquitin-mediated signaling pathway, and stimulating at least one polyubiquitin-mediated signaling pathway.

6. A method of producing the antibody or antigen-binding fragment of claim 1 , comprising culturing a host cell comprising a nucleic acid molecule encoding the antibody or antigen-binding fragment under conditions wherein the antibody or antigen-binding fragment is produced.

7. A composition comprising an effective amount of the antibody or antigen-binding fragment of claim 1 and a pharmaceutically acceptable carrier.

8. A method of identifying the presence of K63-linked polyubiquitin or a K63-linked polyubiquitinated protein in a sample, comprising contacting the sample with at least one antibody or antigen-binding fragment of claim 1 .

9. A method of determining the presence of a K63-linked polyubiquitin or a K63-linked polyubiquitinated protein in a sample suspected of containing a K63-linked polyubiquitin or K63-linked polyubiquitinated protein, comprising exposing the sample to at least one antibody or antigen-binding fragment of claim 1 and determining the binding of the at least one antibody or antigen-binding fragment to a K63-linked polyubiquitin or K63-linked polyubiquitinated protein in the sample.

10. A method of separating K63-linked polyubiquitinated protein from non-K63-linked polyubiquitinated protein in a sample, comprising contacting the sample with at least one antibody or antigen-binding fragment of claim 1 .

11. A method of determining the function and/or activity of K63-linked polyubiquitin in a cell or sample comprising contacting the cell or sample with at least one antibody or antigen-binding fragment of claim 1 and assessing the effect of said contacting step on the cell or sample.

12. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 and the HVR-L2 sequence is unsubstituted SEQ ID NO: 9.

13. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an R at position 4.

14. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with a D at position 1 and the HVR-L2 sequence is SEQ ID NO: 9.

15. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with a D at position 1 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an A at position 4.

16. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with a D at position 1 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an S at position 4.

17. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with a D at position 1 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an T at position 4.

18. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with an A at position 3 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an A at position 4.

19. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with an A at position 3 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an L at position 4.

20. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with an A at position 3 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an N at position 4.

21. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with a T at position 3 and the HVR-L2 sequence is SEQ ID NO: 9 substituted with an T at position 4.

22. The antibody of claim 1 , wherein the HVR-H2 sequence is SEQ ID NO: 61 substituted with an L at position 6 and the HVR-L2 sequence is SEQ ID NO: 9.

23. The antibody of claim 1 , which has a Kd for K63-linked polyubiquitin of about 10 nM to about 90 nM.

Assignees

• Genentech Inc

Inventors

• Kelley Robert F
• Matsumoto Marissa

Classifications

• A61P43/00

  Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00 · CPC title

• A61P37/00

  Drugs for immunological or allergic disorders · CPC title

• A61P35/02

  specific for leukemia · CPC title

• A61P35/00

  Antineoplastic agents · CPC title

• A61P29/00

  Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID] · CPC title