Cells of which genome is modified

• US 20040110704A1
• Filed: 04/09/2003
• Published: 06/10/2004
• Est. Priority Date: 04/09/2002
• Status: Abandoned Application

• Current Assignee: Kyowa Hakko Kogyo Company Limited (Kirin Holdings Company Limited)
• Original Assignee: Kyowa Hakko Kogyo Company Limited (Kirin Holdings Company Limited)
• Inventors: Yamane, Naoko, Yamano, Kazuya, Satoh, Mitsuo, Mori, Katsuhiro
• Application Number:
  US10/409,609
• Publication Number:
  US 20040110704A1
• Time in Patent Office: Days
• US Class Current: 514/44
• CPC Class Codes: A01K 2217/075 inducing loss of function, i.e. knock out
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 37/08 Antiallergic agents antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 16/2866 against receptors for cytokines, lymphokines, interferons
• CPC Class Codes: C07K 16/2896 against molecules with a "CD"-designation, not provided for elsewhere
• CPC Class Codes: C07K 16/3084 against tumour-associated gangliosides
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/41 Glycosylation, sialylation, or fucosylation
• CPC Class Codes: C07K 2317/732 Antibody-dependent cellular cytotoxicity [ADCC]
• CPC Class Codes: C12N 2510/00 Genetically modified cells
• CPC Class Codes: C12N 9/1051 Hexosyltransferases (2.4.1)
• CPC Class Codes: C12Y 204/01068 Glycoprotein 6-alpha-L-fucosyltransferase (2.4.1.68), i.e. FUT8

Abstract

A cell in which genome is modified so as to have a more decreased or deleted activity of an enzyme relating to modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α-bond in a complex N-glycoside-linked sugar chain than its parent cell, and a process for producing an antibody composition using the cell.

Claims

1. A cell in which genome is modified so as to have a more decreased or deleted activity of an enzyme relating to modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain than its parent cell.

2. The cell according to claim 1, wherein a genomic gene encoding an enzyme relating to modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain is knocked out.

3. The cell according to claim 1 or 2, wherein all of alleles on a genome encoding an enzyme relating to modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain are knocked out.

4. The cell according to any one of claims 1 to 3, wherein the enzyme relating to modification of a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain is α

1,6-fucosyltransferase.

5. The cell according to claim 4, wherein the α

1,6-fucosyltransferase is a protein encoded by a DNA selected from the group consisting of the following (a) to (d);

(a) a DNA comprising the nucleotide sequence represented by SEQ ID NO;

1;

(b) a DNA comprising the nucleotide sequence represented by SEQ ID NO;

2;

(c) a DNA which hybridizes with a DNA comprising the nucleotide sequence represented by SEQ ID NO;

1 under stringent conditions and having an α

1,6-fucosyltransferase activity;

(d) a DNA which hybridizes with a DNA comprising the nucleotide sequence represented by SEQ ID NO;

2 under stringent conditions and having an α

1,6-fucosyltransferase activity.

6. The cell according to claim 4, wherein the α

1,6-fucosyltransferase is a protein selected from the group consisting of the following (a), (b), (c), (d), (e) and (f);

(a) a protein comprising the amino acid sequence represented by SEQ ID NO;

4;

(b) a protein comprising the amino acid sequence represented by SEQ ID NO;

5;

(c) a protein which comprises an amino acid sequence in which at least one amino acid is deleted, substituted, inserted and/or added in the amino acid sequence represented by SEQ ID NO;

4 and has an α

1,6-fucosyltransferase activity;

(d) a protein which comprises an amino acid sequence in which at least one amino acid is deleted, substituted, inserted and/or added in the amino acid sequence represented by SEQ ID NO;

5 and has an α

1,6-fucosyltransferase activity;

(e) a protein which comprises an amino acid sequence having a homology of 80% or more with the amino acid sequence represented by SEQ ID NO;

4 and has an α

1,6-fucosyltransferase activity;

(f) a protein which comprises an amino acid sequence having a homology of 80% or more with the amino acid sequence represented by SEQ ID NO;

5 and has an α

1,6-fucosyltransferase activity.

7. The cell according to any one of claims 1 to 6, which is resistant to a lectin which recognizes a sugar chain in which 1-position of fucose is bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain.

8. The cell according to claim 7, which is resistant to at least one lectin selected from the group consisting of the following (a) to (d):

(a) a Lens culinaris lectin;

(b) a Pisum sativum lectin;

(c) a Vicia faba lectin,

(d) an Aleuria aurantia lectin.

9. The cell according to any one of claims 1 to 8, which is selected from the group consisting of the following (a) to (j):

(a) a CHO cell derived from a Chinese hamster ovary tissue;

(b) a rat myeloma cell line YB2/3HL.P2.G11.16Ag.20 cell;

(c) a mouse myeloma cell line NS0 cell;

(d) a mouse myeloma cell line SP2/0-Ag14 cell;

(e) a BHK cell derived from a Syrian hamster kidney tissue;

(f) a hybridoma cell which produces an antibody,

(g) a human leukemic cell line Namalwa cell;

(h) an embryonic stem cell;

(i) a fertilized egg cell;

(j) a plant cell.

10. The cell according to any one of claims 1 to 9, which contains a gene encoding an antibody molecule.

11. The cell according to claim 10, wherein the antibody molecule is selected from the group consisting of the following (a) to (d):

(a) a human antibody;

(b) a humanized antibody;

(c) an antibody fragment comprising the Fc region of (a) or (b), (d) a fusion protein comprising the Fc region of (a) or (b).

12. The cell according to claim 10 or 11, wherein the antibody molecule belongs to an IgG class.

13. The cell according to any one of claims 1 to 12, which produces an antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity than an antibody composition produced by its parent cell.

14. The cell according to claim 13, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity has a higher ratio of a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chain among total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition than an antibody composition produced by its parent cell.

15. The cell according to claim 14, wherein the sugar chain in which fucose is not bound is a sugar chain in which 1-position of the fucose is not bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain.

16. The cell according to any one of claims 13 to 15, wherein the ratio of a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end through α

-bond is 20% or more of total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition.

17. The cell according to any one of claims 13 to 16, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition which has no sugar chain in which fucose is bound to N-acetylglucosamine in the reducing end in the sugar chain.

18. A process for producing an antibody composition, which comprises using the cell according to any one of claims 10 to 17.

19. A process for producing an antibody composition, which comprises culturing the cell according to any one of claims 10 to 18 in a medium to form and accumulate an antibody composition in the culture, and recovering the antibody composition from the culture.

20. The process according to claim 18 or 19, wherein the antibody composition is an antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity than an antibody composition produced by its parent cell.

21. The process according to claim 20, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity has a higher ratio of a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chain among total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition than an antibody composition produced by its parent cell.

22. The process according to claim 21, wherein the sugar chain in which fucose is not bound is a sugar chain in which 1-position of the fucose is not bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain.

23. The cell according to any one of claims 20 to 22, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity is a an antibody composition having a ratio of a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end through α

-bond of 20% or more of total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition.

24. The cell according to any one of claims 20 to 23, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition which has no sugar chain in which fucose is bound to N-acetylglucosamine in the reducing end in the sugar chain.

25. A transgenic non-human animal or plant or the progenies thereof, which is produced by using the cell according to any one of claims 1 to 9.

26. The transgenic non-human animal or plant or the progenies thereof according to claim 24, wherein the transgenic non-human animal is an animal selected from the group consisting of cattle, sheep, goat, pig, horse, mouse, rat, fowl, monkey and rabbit.

27. The transgenic non-human animal or plant or the progenies thereof according to claim 25 or 26, which is introduced with a gene encoding an antibody molecule.

28. The transgenic non-human animal or plant or the progenies thereof according to claim 27, wherein the antibody molecule is selected from the group consisting of the following (a) to (d):

(a) a human antibody;

(b) a humanized antibody;

(c) an antibody fragment comprising the Fc region of (a) or (b);

(d) a fusion protein comprising the Fc region of (a) or (b).

29. The transgenic non-human animal or plant or the progenies thereof according to claim 27 or 28, wherein the antibody molecule belongs to an IgG class.

30. A process for producing an antibody composition, which comprises rearing the transgenic non-human animal or plant according to any one of claims 27 to 29, isolating a tissue or body fluid comprising an antibody molecule introduced from the reared animal or plant- and recovering an antibody composition comprising a desired antibody molecule from the isolated tissue or body fluid.

31. A process for producing an antibody composition, which comprises isolating an antibody-producing cell from the transgenic non-human animal or plant or the progenies thereof according to any one of claims 26 to 29;

culturing the isolated antibody-producing cell in a medium to form and accumulate an antibody composition in the culture, and recovering the antibody composition from the culture.

32. The process according to claim 30 or 31, wherein the antibody composition is an antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity than an antibody composition produced by a transgenic non-human animal or plant or the progenies thereof in which genome is not modified.

33. The process according to claim 32, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity has a higher ratio of a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chain among total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition than an antibody composition produced by a transgenic non-human animal or plant or the progenies thereof in which genome is not modified.

34. The process according to claim 33, wherein the sugar chain in which fucose is not bound is a sugar chain in which 1-position of the fucose is not bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain.

35. The process according to any one of claims 32 to 34, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition in which a ratio of a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chain is 20% or more of total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition.

36. The cell according to any one of claims 32 to 35, wherein the antibody composition having a higher antibody-dependent cell-mediated cytotoxic activity is an antibody composition which has no sugar chain in which fucose is bound to N-acetylglucosamine in the reducing end in the sugar chain.

39. An antibody composition produced by the process of any one of claims 18 to 24.

40. An antibody composition produced by the process of any one of claims 18 to 24.

37. An antibody composition comprising an antibody molecule having an glycoside-linked sugar chain in the Fc region, which has a sugar chain in which fucose is not bound to N-acetylglucosamine in the reducing end in the sugar chain among total complex N-glycoside-linked sugar chains bound to the Fc region in the antibody composition.

38. The antibody composition according to claim 37, wherein the sugar chain in which fucose is not bound is a sugar chain in which 1-position of the fucose is not bound to 6-position of N-acetylglucosamine in the reducing end through α

-bond in a complex N-glycoside-linked sugar chain.

41. A medicament comprising as an active ingredient the antibody composition according to any one of claims 37 to 40.

42. The medicament according to claim 41, which is a diagnostic agent, an preventing agent or a treating agent for tumor-accompanied diseases, allergy-accompanied diseases, inflammatory-accompanied diseases, autoimmune diseases, cardiovascular diseases, viral infection-accompanied diseases or bacterial infection-accompanied diseases.

43. Use of the antibody composition according to any one of claims 37 to 40 in the manufacture of the medicament according to claim 41 or 42.