Trispecific and/or trivalent binding proteins

A trispecific binding protein comprising four polypeptide chains that form three antigen binding sites that specifically bind one or more target proteins, wherein a first polypeptide chain of the binding protein comprises a structure represented by the formula:

• VL2-L1-VL1-L2-CL [I]
• and a second polypeptide chain of the binding protein comprises a structure represented by the formula:
• VH1-L3-VH2-L4-CH1-hinge-CH2-CH3 [II]
• and a third polypeptide chain of the binding protein comprises a structure represented by the formula:
• VH3-CH1-hinge-CH2-CH3 [III]
• and a fourth polypeptide chain of the binding protein comprises a structure represented by the formula:
• VL3-CL [IV]
• wherein:
• VL1 is a first immunoglobulin light chain variable domain;
• VL2 is a second immunoglobulin light chain variable domain;
• VL3 is a third immunoglobulin light chain variable domain;
• VH1 is a first immunoglobulin heavy chain variable domain;
• VH2 is a second immunoglobulin heavy chain variable domain;
• VH3 is a third immunoglobulin heavy chain variable domain;
• CL is an immunoglobulin light chain constant domain;
• CH1 is an immunoglobulin CH1 heavy chain constant domain;
• CH2 is an immunoglobulin CH2 heavy chain constant domain;
• CH3 is an immunoglobulin CH3 heavy chain constant domain;
• hinge is an immunoglobulin hinge region connecting the CH1 and CH2 domains;
• and
• L1, L2, L3 and L4 are each independently amino acid linkers or zero amino acids in length;
• wherein the polypeptide of formula I and the polypeptide of formula II form a cross-over light chain-heavy chain pair of the binding protein;
• wherein VH1 and VL1 form a binding pair and a first antigen binding site, and VH2 and VL2 form a binding pair and a second antigen binding site;
• and wherein VH3 and VL3 form a binding pair and a third antigen binding site.