Anti-human MUC1 antibody Fab fragment

• US 10,517,966 B2
• Filed: 11/17/2017
• Issued: 12/31/2019
• Est. Priority Date: 11/18/2016
• Status: Active Grant

• Current Assignee: Astellas Pharma Incorporated
• Original Assignee: Astellas Pharma Incorporated
• Inventors: Morinaka, Akifumi, Shirai, Hiroki, Hirayama, Kazunori, Hosogai, Naomi, Doihara, Hitoshi
• Primary Examiner(s): Puttlitz, Karl J
• Application Number:
  US16/462,143
• Publication Number:
  US 20190307906A1
• Time in Patent Office: 774 Days
• Field of Search: None
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 39/395 Antibodies agglutinins A61K38/36 ; as drug carriers A61K47/50; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
• CPC Class Codes: A61K 47/54 the modifying agent being an organic compound
• CPC Class Codes: A61K 47/6851 the antibody targeting a determinant of a tumour cell
• CPC Class Codes: A61K 47/6855 the tumour determinant being from breast cancer cell
• CPC Class Codes: A61K 49/0021 the fluorescent group being a small organic molecule
• CPC Class Codes: A61K 49/0032 Methine dyes, e.g. cyanine dyes
• CPC Class Codes: A61K 49/0036 Porphyrins used in photodynamic therapy A61K41/0071 or A61K41/0076; used as targeting group or modifying agent for targeting a therapeutic compound A61K47/546
• CPC Class Codes: A61K 49/0058 Antibodies
• CPC Class Codes: A61K 49/16 Antibodies; Immunoglobulins; Fragments thereof
• CPC Class Codes: A61K 51/1051 the tumor cell being from breast, e.g. the antibody being herceptin
• CPC Class Codes: A61K 51/106 the tumor cell being from kidney or bladder
• CPC Class Codes: A61K 51/1093 conjugates with carriers being antibodies
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/18 against material from animals or humans
• CPC Class Codes: C07K 16/28 against receptors, cell surface antigens or cell surface determinants
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 16/3092 against tumour-associated mucins
• CPC Class Codes: C07K 17/00 Carrier-bound or immobilised peptides carrier-bound or immobilised enzymes C12N11/00; Preparation thereof
• CPC Class Codes: C07K 2317/21 from primates, e.g. man
• CPC Class Codes: C07K 2317/24 : containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/40 : characterized by post-translational modification
• CPC Class Codes: C07K 2317/55 : Fab or Fab'
• CPC Class Codes: C07K 2317/76 : Antagonist effect on antigen, e.g. neutralization or inhibition of binding
• CPC Class Codes: C07K 2317/92 : Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• CPC Class Codes: C12N 15/09 : Recombinant DNA-technology
• CPC Class Codes: C12N 15/85 : for animal cells
• CPC Class Codes: C12N 5/10 : Cells modified by introduction of foreign genetic material
• CPC Class Codes: C12P 21/02 : having a known sequence of two or more amino acids, e.g. glutathione
• CPC Class Codes: G01N 2333/4725 : Mucins, e.g. human intestinal mucin
• CPC Class Codes: G01N 33/57407 : Specifically defined cancers
• CPC Class Codes: G01N 33/57415 : of breast

Abstract

The problem to be solved is to provide an anti-human MUC1 antibody Fab fragment that is expected to be useful in the diagnosis and/or treatment of a cancer, particularly, the diagnosis and/or treatment of breast cancer or bladder cancer, and a diagnosis approach and/or a treatment approach using a conjugate comprising the Fab fragment. The solution is an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment comprising a heavy chain variable region consisting of the amino acid sequence represented by SEQ ID NO: 8 or 10, and a light chain comprising a light chain variable region consisting of the amino acid sequence represented by SEQ ID NO: 12, and a conjugate comprising the Fab fragment.

Claims

1. An anti-human MUC1 antibody Fab fragment selected from the group consisting of the following (a) and (b):

(a) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment comprising a heavy chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

8 or SEQ ID NO;

10 and a light chain comprising a light chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

12; and

(b) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment comprising a heavy chain variable region derived from a heavy chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

8 or SEQ ID NO;

10 by the modification of glutamine at amino acid position 1 of SEQ ID NO;

8 or SEQ ID NO;

10 into pyroglutamic acid, and a light chain comprising a light chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

12.

2. The anti-human MUC1 antibody Fab fragment according to claim 1 which is selected from the group consisting of the following (a) and (b):

(a) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment consisting of the amino acid sequence represented by SEQ ID NO;

2 or SEQ ID NO;

4 and a light chain consisting of the amino acid sequence represented by SEQ ID NO;

6; and

(b) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment derived from a heavy chain fragment consisting of the amino acid sequence represented by SEQ ID NO;

2 or SEQ ID NO;

4 by the modification of glutamine at amino acid position 1 of SEQ ID NO;

2 or SEQ ID NO;

4 into pyroglutamic acid, and a light chain consisting of the amino acid sequence represented by SEQ ID NO;

6.

3. The anti-human MUC1 antibody Fab fragment according to claim 1 which is selected from the group consisting of the following (a) and (b):

(a) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment comprising a heavy chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

10 and a light chain comprising a light chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

12; and

(b) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment comprising a heavy chain variable region derived from a heavy chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

10 by the modification of glutamine at amino acid position 1 of SEQ ID NO;

10 into pyroglutamic acid, and a light chain comprising a light chain variable region consisting of the amino acid sequence represented by SEQ ID NO;

12.

4. The anti-human MUC1 antibody Fab fragment according to claim 3 which is selected from the group consisting of the following (a) and (b):

(a) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment consisting of the amino acid sequence represented by SEQ ID NO;

4 and a light chain consisting of the amino acid sequence represented by SEQ ID NO;

6; and

(b) an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment derived from a heavy chain fragment consisting of the amino acid sequence represented by SEQ ID NO;

4 by the modification of glutamine at amino acid position 1 of SEQ ID NO;

4 into pyroglutamic acid, and a light chain consisting of the amino acid sequence represented by SEQ ID NO;

6.

5. The anti-human MUC1 antibody Fab fragment according to claim 4 which is an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment consisting of the amino acid sequence represented by SEQ ID NO:

4 and a light chain consisting of the amino acid sequence represented by SEQ ID NO;

6.

6. The anti-human MUC1 antibody Fab fragment according to claim 4 which is an anti-human MUC1 antibody Fab fragment comprising a heavy chain fragment derived from a heavy chain fragment consisting of the amino acid sequence represented by SEQ ID NO:

4 by the modification of glutamine at amino acid position I of SEQ ID NO;

4 into pyroglutamic acid, and a light chain consisting of the amino acid sequence represented by SEQ ID NO;

6.

7. A conjugate comprising one or more labeling moiety and the anti-human MUC1 antibody Fab fragment according to any one of claims 1 to 6.

8. The conjugate according to claim 7, wherein the labeling moiety is (i) a ligand and a linker or (ii) a ligand.

9. The conjugate according to claim 8, wherein the ligand is a ligand represented by the following formula (A):

10. The conjugate according to claim 9, wherein the labeling moiety is a ligand and a linker represented by the following formula (A′

);

11. The conjugate according to claim 10, wherein the anti-human MUC1 antibody Fab fragment is bound via an amino group thereof to the carbon atom of a labeling moiety terminal C(═

S) group.

12. A conjugate selected from the group consisting of the following (a) to (c):

(a) the conjugate according to claim 11 wherein the anti-human MUC1 antibody Fab fragment is the anti-human MUC1 antibody Fab fragment according to claim 5;

(b) the conjugate according to claim 11 wherein the anti-human MUC1 antibody Fab fragment is the anti-human MUC1 antibody Fab fragment according to claim 6; and

(c) a conjugate which is a mixture of (a) and (b).

13. The conjugate according to any one of claims 10 to 12, further comprising a metal.

14. The conjugate according to claim 13, wherein the metal is a metal radioisotope.

15. The conjugate according to claim 14, wherein the metal is ⁸⁹Zr.

16. The conjugate according to claim 12, further comprising ⁸⁹Zr.

17. A polynucleotide selected from the group consisting of the following (a) and (b):

(a) a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1; and

(b) a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1.

18. A polynucleotide selected from the group consisting of the following (a) and (b):

(a) a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5; and

(b) a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5.

19. An expression vector comprising the following (a) and/or (b):

(a) a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1; and

(b) a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1.

20. An expression vector comprising the following (a) and/or (b):

(a) a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5; and

(b) a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5.

21. A host cell selected from the group consisting of the following (a) to (d):

(a) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1;

(b) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1;

(c) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1 and a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1; and

(d) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1 and an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1.

22. A host cell selected from the group consisting of the following (a) to (d):

(a) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5;

(b) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5;

(c) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5 and a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5; and

(d) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5 and an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5.

23. A method for producing an anti-human MUC1 antibody Fab fragment comprising the step of culturing a host cell selected from the group consisting of the following (a) to (c) to express the anti-human MUC1 antibody Fab fragment:

(a) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1 and a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1;

(b) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1 and an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1; and

(c) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 1, and a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 1.

24. A method for producing an anti-human MUC1 antibody Fab fragment comprising the step of culturing a host cell selected from the group consisting of the following (a) to (c) to express the anti-human MUC1 antibody Fab fragment:

(a) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5 and a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5;

(b) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5 and an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5; and

(c) a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the heavy chain fragment of the anti-human MUC1 antibody Fab fragment according to claim 5, and a host cell transformed with an expression vector comprising a polynucleotide comprising a nucleotide sequence encoding the light chain of the anti-human MUC1 antibody Fab fragment according to claim 5.

25. A method for producing a conjugate comprising a labeling moiety and an anti-human MUC1 antibody Fab fragment, comprising the steps of:

producing the anti-human MUC1 antibody Fab fragment by the method according to claim 23 or 24; and

covalently binding the Fab fragment to the labeling moiety.

26. The method for producing a conjugate according to claim 25, wherein the step of covalently binding the Fab fragment to the labeling moiety is the step of i) binding the Fab fragment via a linker to a ligand or ii) covalently binding the Fab fragment directly to a ligand.

27. The method for producing a conjugate according to claim 26, further comprising the step of labeling the ligand of the conjugate with a metal radioisotope.

28. The method for producing a conjugate according to claim 25, wherein the step of covalently binding the Fab fragment to the labeling moiety is the step of i) binding the Fab fragment via a linker to a fluorescent dye or ii) covalently binding the Fab fragment directly to a fluorescent dye.

29. A pharmaceutical composition comprising one or more conjugate according claim 7, and a pharmaceutically acceptable carrier.

30. A pharmaceutical composition comprising one or more conjugate according to claim 16, and a pharmaceutically acceptable carrier.

31. A method for diagnosing breast cancer or bladder cancer, comprising preoperatively or intraoperatively administering the conjugate according to claim 7 to a subject.

32. A method for treating breast cancer or bladder cancer, comprising the step of administering a therapeutically effective amount of the conjugate according to claim 7.