ANTIBODY HAVING ENHANCED ACTIVITY, AND METHOD FOR MODIFYING SAME
As a result of producing ACE910 variants in which various sites of the constant regions were modified, the inventors discovered bispecific antibodies having FVIII mimetic activity higher than that of ACE910. The inventors also identified mutation positions that elevate the FVIII mimetic activity and discovered methods for elevating the FVIII mimetic activity by using the mutations.
1 . A modified bispecific antibody against blood coagulation factor IX and/or activated blood coagulation factor IX and blood coagulation factor X, wherein the antibody comprises a modification of at least one amino acid residue in the constant region of the antibody, and wherein the antibody has higher blood coagulation factor VIII mimetic activity than that of a bispecific antibody, which is an antibody before the modification, in which an H chain comprising the amino acid sequence of SEQ ID NO: 1 associates with an L chain comprising the amino acid sequence of SEQ ID NO: 3, and an H chain comprising the amino acid sequence of SEQ ID NO: 2 associates with an L chain comprising the amino acid sequence of SEQ ID NO: 3.
2 . The antibody of claim 1 , wherein the modification is a modification in a CH3 region and/or hinge region.
3 . The antibody of claim 2 , wherein the modification in the CH3 region is a modification at position 434 according to EU numbering.
4 . The antibody of claim 3 , wherein the modification at position 434 according to EU numbering is a modification to Tyr (Y).
5 . The antibody of claim 2 , wherein the modification in the hinge region is a modification of positions 216 to 230 according to EU numbering to AAAC.
6 . A pharmaceutical composition comprising the antibody of any one of claims 1 to 5 , and a pharmaceutically acceptable carrier.
7 . The composition of claim 6 , which is a pharmaceutical composition for use in prevention and/or treatment of bleeding, a disease accompanying bleeding, or a disease caused by bleeding, wherein the disease is a disease that develops and/or progresses by a decrease or deficiency in the activity of blood coagulation factor VIII and/or activated blood coagulation factor VIII.
8 . The composition of claim 7 , wherein the disease that develops and/or progresses by a decrease or deficiency in the activity of blood coagulation factor VIII and/or activated blood coagulation factor VIII is hemophilia A, acquired hemophilia, von Willebrand's disease, or a disease in which an inhibitor against blood coagulation factor VIII and/or activated blood coagulation factor VIII has been raised.
9 . A kit for use in a method for prevention and/or treatment of bleeding, a disease accompanying bleeding, or a disease caused by bleeding, wherein the kit comprises at least the bispecific antibody of any one of claims 1 to 5 .
10 . A method for enhancing the blood coagulation factor VIII mimetic activity of a bispecific antibody against blood coagulation factor IX and/or activated blood coagulation factor IX and blood coagulation factor X, wherein the method modifies at least one amino acid residue in the constant region of a bispecific antibody in which an H chain comprising the amino acid sequence of SEQ ID NO: 1 associates with an L chain comprising the amino acid sequence of SEQ ID NO: 3, and an H chain comprising the amino acid sequence of SEQ ID NO: 2 associates with an L chain comprising the amino acid sequence of SEQ ID NO: 3.
11 . The method of claim 10 , wherein the modification is a modification of at least one amino acid residue in a CH3 region and/or hinge region.
12 . The method of claim 11 , wherein the modification in the CH3 region is a modification at position 434 according to EU numbering.
13 . The method of claim 12 , wherein the modification at position 434 is a modification to Tyr (Y).
14 . The method of claim 11 , wherein the modification in the hinge region is a modification of positions 216 to 230 according to EU numbering to AAAC.
15 . A method for producing a bispecific antibody against blood coagulation factor IX and/or activated blood coagulation factor IX and blood coagulation factor X, wherein the method comprises: (a) modifying at least one amino acid residue in the constant region of a bispecific antibody in which an H chain comprising the amino acid sequence of SEQ ID NO: 1 associates with an L chain comprising the amino acid sequence of SEQ ID NO: 3, and an H chain comprising the amino acid sequence of SEQ ID NO: 2 associates with an L chain comprising the amino acid sequence of SEQ ID NO: 3, which is an antibody before the modification, so that the blood coagulation factor VIII mimetic activity is enhanced compared to the antibody before the modification, by modifying a nucleic acid encoding an antibody comprising said amino acid residue; (b) culturing a host cell to express the nucleic acid; and (c) collecting the antibody from a culture of the host cell.