Humanized anti-LT-β-R antibodies

• US 7,429,644 B2
• Filed: 04/10/2003
• Issued: 09/30/2008
• Est. Priority Date: 10/13/2000
• Status: Expired due to Fees

• Current Assignee: Biogen Idec MA Inc. (Biogen, Inc.)
• Original Assignee: Biogen Idec MA Inc. (Biogen, Inc.)
• Inventors: Garber, Ellen, Saldanha, Jose William, Lyne, Paul
• Primary Examiner(s): Blanchard; David J.
• Application Number:
  US10/412,406
• Publication Number:
  US 20040058394A1
• Time in Patent Office: 2,000 Days
• Field of Search: 530/387.3, 530/388.1, 530/388.22, 530/388.8, 424/133.1, 424/141.1, 424/143.1, 424/155.1, 435/69.1, 435/69.6, 435/70.21
• US Class Current: 530/387.3
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/2866 against receptors for cytokines, lymphokines, interferons
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered

Abstract

Humanized antibodies to LT-β-R and methods of use thereof are provided.

Claims

1. A humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen-binding fragment thereof, comprising

a light chain variable domain sequence as set forth in SEQ ID NO;

8, and

a heavy chain comprising complementary determining regions as set forth in SEQ ID NO;

2,

wherein the antibody comprises at least one of the following residues in its heavy chain;

F37, T40, A49, M89 and V93 (Kabat numbering convention), or a conservative amino acid substitution thereof.

3. The humanized antibody of claim 1, wherein the antibody further comprises a heavy chain variable domain sequence as set forth in SEQ ID NO:

16.

7. The humanized antibody according to any one of claims 1-4 wherein the antibody substantially retains the binding properties of the parent antibody.

8. An immunotoxin comprising the humanized antibody according to any one of claims 1-4, 5 or 6.

9. The humanized antibody according to any one of claims 1-4, 5 or 6, wherein the antibody is linked to a chemotherapeutic drug.

10. A pharmaceutical composition comprising the humanized antibody of any one of claims 1-4, 5 or 6, and a pharmaceutically acceptable carrier.

13. The humanized antibody of claim 9, wherein the chemotherapeutic drug is selected from the group consisting of adriamycin, 5FU, vinbiastine, actinomycin D, etoposide, cisplatin, methotrexate, and doxorubicin.

14. The humanized antibody of any one of claims 1-4, wherein the antibody is an IgG1.

15. The humanized antibody, or antigen binding fragment thereof, of any one of claims 1-4, wherein the antigen binding fragment is selected from the group consisting of a Fab fragment, an F(ab'"'"')_2, and an Fv fragment.

2. A humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen-binding fragment thereof, comprising

a heavy chain variable domain sequence as set forth in SEQ ID NO;

16, and

a light chain comprising complementary determining regions as set forth in SEQ ID NO;

1, and residue Y71 (Kabat numbering convention), or a conservative amino acid substitution thereof.

30. The humanized antibody, or antigen-binding fragment of claim 2, further comprising at least one of the following residues in the light chain:

K3, W41, I46, and Q69 (Kabat numbering convention), or a conservative amino acid substitution thereof.

31. The humanized antibody, or antigen-binding fragment of claim 2, wherein the antibody further comprises a light chain variable domain sequence as set forth in SEQ ID NO:

8.

4. An antibody, or an antigen-binding fragment thereof, comprising the same heavy and light variable region sequences as an antibody produced by cell line E46.4 (ATCC patent deposit designation PTA-3357) or cell line E77.4 (ATCC patent deposit designation PTA-3765).

20. The humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen binding fragment thereof, of any one of claims 4, 5, or 6, wherein the humanized antibody, or antigen-binding fragment thereof, significantly increases survival of an animal in an in vivo tumor growth assay.

5. A humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody comprising

a) the light chain amino acid sequence set forth in SEQ ID NO;

32 and

b) the heavy chain amino acid sequence set forth in SEQ ID NO;

33.

28. The humanized antibody of claim 5 or 6, wherein the antibody is an IgG1.

29. The humanized antibody, or antigen binding fragment thereof, of claim 5 or 6, wherein the antigen binding fragment is selected from the group consisting of a Fab fragment, an F(ab'"'"')₂, and an Fv fragment.

6. A humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen binding fragment thereof, comprising

a) a light chain variable region comprising complementary determining regions (CDRs) as set forth in SEQ ID NO;

1, and a framework region from a human acceptor light chain, wherein the human acceptor light chain is antibody TNF-A1'"'"'CL (Kabat ID

     004770), and

b) a heavy chain variable region comprising CDRs as set forth in SEQ ID NO;

2, and a framework region from a human acceptor heavy chain, wherein the human acceptor heavy chain is antibody FLA-IgG'"'"'CL (Kabat ID

     040003),

wherein the framework region of a) or b) comprises at least one framework residue from the mouse CBE11 antibody.

16. The humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen binding fragment thereof, of claim 6, wherein the at least one framework residue from the mouse CBE11 antibody is selected from the group consisting of

i) an unusual murine residue at the interface between the variable light and heavy chains;

ii) an unusual murine residue close to the antigen binding site; and

iii) both an unusual murine residue at the interface between the variable light and heavy chains and an unusual murine residue close to the antigen binding site.

17. The humanized antibody, or an antigen binding fragment thereof, of claim 16, wherein the unusual murine residue between the variable light and heavy chains is selected from the group consisting of S34, I46, L89, and H91 in the light chain or Y35, F37, V93, and E95 in the heavy chain.

18. The humanized antibody, or an antigen binding fragment thereof, of claim 16, wherein the unusual murine residue close to the antigen binding site is selected from the group consisting of K3, M11, Y12, W41, Q69, S72, D81, and T83 in the light chain, or F37, T40, E42, A49, and N77 in the heavy chain.

19. The humanized antibody, or an antigen binding fragment thereof, of claim 16, wherein the unusual murine residue at the interface between the variable light and heavy chains and the unusual residue close to the antigen binding site is F37 in the heavy chain.

11. The cell line E46.4 (ATCC patent deposit designation PTA-3357) or cell line E77.4 (ATCC patent deposit designation PTA-3765).

12. A humanized antibody, or antigen binding fragment thereof, comprising

a) a light chain comprising complementary determining regions (CDRs) as set forth in SEQ ID NO:

1, and residues K3, W41, I46, O69, and Y71 (Kabat numbering convention), or a conservative amino acid substitution thereof, wherein the remainder of the light chain is from a human acceptor framework, and

b) a heavy chain comprising CDRs as set forth in SEQ ID NO;

2, residues F37, T40, A49, M89, and residue V93 (Kabat numbering convention), or a conservative amino acid substitution thereof, wherein the remainder of the heavy chain is from a human acceptor framework.

21. A humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen binding fragment thereof, comprising a light chain comprising the light chain variable region as set forth in SEQ ID NO;

8, and a heavy chain comprising CDRs as set forth in SEQ ID NO;

2.

22. The humanized antibody of claim 21, wherein the heavy chain further comprises residues F37 and A49 (Kabat numbering convention), or a conservative amino acid substitution thereof, within the framework region.

23. The humanized antibody of claim 21, wherein the heavy chain comprises the heavy chain variable region as set forth in SEQ ID NO:

16.

25. A pharmaceutical composition comprising the humanized antibody, or antigen binding fragment thereof, of claim 21 or 24, and a pharmaceutically acceptable carrier.

26. The humanized antibody of claim 21 or 24, wherein the antibody is an IgG1.

27. The humanized antibody, or antigen binding fragment thereof, of claim 21 or 24, wherein the antigen binding fragment is selected from the group consisting of a Fab fragment, an F(ab'"'"')₂, and an Fv fragment.

24. A humanized anti-lymphotoxin-beta receptor (LT-β

-R) antibody, or an antigen binding fragment thereof, comprising a light chain comprising CDRs as set forth in SEQ ID NO;

1, and a heavy chain comprising a heavy chain variable region as set forth in SEQ ID NO;

16.

32. The humanized antibody of claim 24, wherein the light chain further comprises residue Y71 (Kabat numbering convention), or a conservative amino acid substitution thereof, within the framework region.

33. The humanized antibody of claim 24, wherein the light chain comprises the light chain variable region as set forth in SEQ ID NO:

8.