Antibody and antibody composition production method
[Problem] To provide: an antibody comprising at least two kinds of Fab, and in particular having restricted light chain-heavy chain combinations; a corresponding antibody composition; and production methods for same. [Solution] The present invention provides production methods for (1) an antibody or (2) an antibody composition, the methods using non-natural disulfide bonds.
1. A method for making an antibody comprising a first Fab region which comprises a first light chain and heavy chain, and a second Fab region which comprises a second light chain and heavy chain each being different from said first light chain and heavy chain; the method comprising:
a) substituting at least one amino acid residue other than cysteine in a CL region and a CH1 region in the first Fab region of a parent antibody of said antibody with a cysteine residue which forms a disulfide bond, and
b) forming a non-natural disulfide bond in the first Fab region by said cysteine residue which forms a disulfide bond,
wherein due to the presence of said non-natural disulfide bond, the first Fab region forms a disulfide bond at a position different from the second Fab region,
wherein the non-natural disulfide bond is formed by cysteine residues introduced in at least one set of light chain-heavy chain positions selected from light chain position 124-heavy chain position 126, and light chain position 162-heavy chain position 170, and
wherein the light chain-heavy chain positions are numbered based on Kabat EU numbering system.
2. The method according to claim 1 , wherein the antibody is a bispecific antibody.
3. The method according to claim 1 , wherein the antibody is one wherein at least two antibody fragments are connected through a linker or directly.
4. The method according to claim 1 , wherein the antibody is an antibody fragment.
5. The method according to claim 1 , wherein a Fc region of the antibody is substituted with another molecule.
6. The method according to claim 1 , wherein the antibody is a chimeric antibody, a humanized antibody or a human antibody.
7. The method according to claim 1 , wherein a natural disulfide bond is not formed between a CL region and a CH1 region of at least one Fab region.
8. The method according to claim 1 , wherein the position of a disulfide bond between a CL region and a CH1 region in one Fab region is entirely different from the position of a disulfide bond between a CL region and a CH1 region in at least one other Fab region.
9. An antibody comprising at least two different Fab regions,
wherein at least one Fab region comprises a cysteine residue which forms a non-natural disulfide bond between a CL region and a CH1 region, thereby forming a non-natural disulfide bond,
wherein due to the presence of said non-natural disulfide bond, the position of a disulfide bond between a CL region and a CH1 region in a Fab region is different from the position of a disulfide bond between a CL region and a CH1 region in at least one other Fab region,
wherein the non-natural disulfide bond is formed by cysteine residues introduced in at least one set of light chain-heavy chain positions selected from light chain position 124-heavy chain position 126, and light chain position 162-heavy chain position 170, and
wherein the light chain-heavy chain positions are numbered based on Kabat EU numbering system.
10. The antibody according to claim 9 , which comprises two different Fab regions.
11. The antibody according to claim 9 , wherein the antibody is a bispecific antibody.
12. The antibody according to claim 9 , wherein the antibody is one wherein at least two antibody fragments are connected through a linker or directly.
13. The antibody according to claim 9 , wherein the antibody is an antibody fragment.
14. The antibody according to claim 9 , wherein an Fe region of the antibody is substituted with another molecule.
15. The antibody according to claim 9 , wherein the antibody is a chimeric antibody, a humanized antibody or a human antibody.
16. The antibody according to claim 9 , wherein a natural disulfide bond is not formed between a CL region and a CH1 region of at least one Fab region.
17. The antibody according to claim 9 , wherein the position of a disulfide bond between a CL region and a CH1 region in one Fab region is entirely different from the position of a disulfide bond between a CL region and a CH1 region in at least one other Fab region.
18. The antibody according to claim 9 , which specifically binds to at least CD20.
19. A composition comprising a mixture of at least two antibodies comprising Fab regions, the Fab regions being different between the at least two antibodies,
wherein at least one Fab region comprises a cysteine residue which forms a non-natural disulfide bond between a light chain and a heavy chain, thereby forming a non-natural disulfide bond,
wherein due to the presence of said non-natural disulfide bond, the position of a disulfide bond between a light chain and a heavy chain of a Fab region is different from the position of a disulfide bond between a light chain and a heavy chain of at least one other Fab region,
wherein the non-natural disulfide bond is formed by cysteine residues introduced in at least one set of light chain-heavy chain positions selected from light chain position 124-heavy chain position 126, and light chain position 162-heavy chain position 170, and
wherein the light chain-heavy chain positions are numbered based on Kabat EU numbering system.