Patent ID: 11965030
Assignee: SANOFI
Field: Biotechnology (Chemistry)
Classification: CPC A  C | IPC C

Claim 4:
5. A multispecific antigen-binding protein comprising
a) a first light chain (LC1)/heavy chain (HC1) pair comprising:
(1) a first VL region (VL1) paired with first VH region (VH1) to form a first antigen-binding site;
(2) a first constant heavy chain region 1 (CH1-1) operatively linked to VH1 and a first constant light chain region (CL1) operatively linked to VL1, and

b) a second light chain (LC2)/heavy chain (HC2) pair comprising:
(3) a second VL region (VL2) paired with a second VH region (VH2) to form a second antigen-binding site; and
(4) a second constant heavy chain region 1 (CH1-2) operatively linked to VH2 and a second constant light chain region (CL2) operatively linked to VL2,

wherein the C terminus of CH1-1 is operatively linked to the N terminus of VH2,
wherein one or more cysteine residues are engineered into the one or both of VH1/VL1 and VH2/VL2 pairs to form one or more disulfide bonds,
wherein one or both of the VL1 and VH1 pair and the VL2 and VH2 pair comprise opposite charged mutations that facilitate pairing, said opposite charged mutations comprising (1) a mutated residue in the VH region at Kabat position 39 selected from E, D, K, R, or H, and (2) a mutated residue in the VL region at Kabat position 38 selected from E, D, K, R, or H, and wherein the mutated residue in the VH region has an opposite charge from the mutated residue in the VL region,
wherein one or both of the CL1 and CH1-1 pair and the CL2 and CH1-2 pair comprise mutations that facilitate pairing, and
wherein when both of CL1 and CH1-1 pair and of CL2 and CH1-2 pair comprise mutations to facilitate pairing, the mutations in CH1-1 and CL1 are different than the mutations in CH1-2 and CL2, wherein:
i) CH-1 comprises a T192E mutation and CL1 comprises N137K and S114A mutations and/or CH1-2 comprises a T192E mutation and CL2 comprises N137K and S114A mutations;
ii) CH-1 comprises L143Q and S188V mutations and CL1 comprises V133T and S176V mutations and/or CH1-2 comprises L143Q and S188V mutations and CL2 comprises V133T and S176V mutations;
iii) CH1-1 comprises T192E, L143Q and S188V mutations and CL1 comprises N137K, S114A, V133T and S176V mutations and/or CH1-2 comprises T192E, L143Q and S188V mutations and CL2 comprises N137K, S114A, V133T and S176V mutations;
iv) CH1-1 comprises a K221E mutation and CL1 comprises a E123K mutation and/or CH1-2 comprises a K221E mutation and CL2 comprises a E123K mutation;
v) CH1-1 comprises T192E and K221E mutations and CL1 comprises N137K, S114A and E123K mutations and/or CH1-2 comprises T192E and K221E mutations and CL2 comprises N137K, S114A and E123K mutations;
vi) CH1-1 comprises a K228D mutation and CL1 comprises a D122K mutation and/or CH1-2 comprises a K228D mutation and CL2 comprises a D122K mutation;
vii) CH1-1 comprises K221E and K228D mutations and CL1 comprises D122K and E123K mutations and/or CH1-2 comprises K221E and K228D mutations and CL2 comprises D122K and E123K mutations;
viii) CH1-1 comprises a L143E, a L143D, a L143K, a L143R, or a L143H mutation, and CL1 comprises a S176E, a S176D, a S176K, a S176R, or a S176H mutation and/or CH1-2 comprises a L143E, a L143D, a L143K, a L143R, or a L143H mutation, and CL2 comprises a S176E, a S176D, a S176K, a S176R, or a S176H mutation; and/or
ix) CH1-1 comprises a L124E, a L124D, a L124K, a L124R, or a L124H mutation, and CL1 comprises a V133E, a V133D, a V133K, a V133R, or a V133H mutation and/or CH1-2 comprises a L124E, a L124D, a L124K, a L124R, or a L124H mutation, and CL2 comprises a V133E, a V133D, a V133K, a V133R, or a V133H mutation,
wherein the mutations are according to Kabat numbering.