Binding domain-immunoglobulin fusion proteins
View Patent ↗The invention relates to novel binding domain-immunoglobulin fusion proteins that feature a binding domain for a cognate structure such as an antigen, a counterreceptor or the like, a wild-type IgG1, IGA or IgE hinge region polypeptide or a mutant IgG1 hinge region polypeptide having either zero, one or two cysteine residues, and immunoglobulin CH2 and CH3 domains, and that are capable of ADCC and/or CDC while occurring predominantly as polypeptides that are compromised in their ability to form disulfide-linked multimers. The fusion proteins can be recombinantly produced at high expression levels. Also provided are related compositions and methods, including cell surface forms of the fusion proteins and immunotherapeutic applications of the fusion proteins and of polynucleotides encoding such fusion proteins.
1. A fusion protein, comprising:
(a) a binding domain polypeptide that binds CD20, wherein the binding domain comprises amino acids 23-265 of SEQ ID NO:397 except that leucine at position 155 of SEQ ID NO:397 is substituted with a serine,
(b) a human IgG1 hinge peptide comprising a mutated core sequence as set forth in amino acids 279-282 of SEQ ID NO:397, 398 or 582,
(c) an immunoglobulin heavy chain CH2 constant region polypeptide as set forth in amino acids 284-393 of SEQ ID NO:397 fused to the hinge peptide,
(d) an immunoglobulin heavy chain CH3 constant region polypeptide as set forth in amino acids 39-500 of SEQ ID NO:397 fused to the CH2 polypeptide, and
wherein the fusion protein promotes antibody dependent cell-mediated cytotoxicity, complement fixation, or both.
2. A composition comprising a fusion protein of claim 1 and a pharmaceutically acceptable excipient, diluent, or carrier.
3. A fusion protein, comprising:
(a) a binding domain polypeptide that binds CD20, wherein the binding domain comprises complementary determining regions of amino acids 46-55, 71-76, and 110-118 of an immunoglobulin light chain variable region as set forth in amino acids 23-128 of SEQ ID NO:397 fused via a linker polypeptide to a sequence comprising complementary determining regions of amino acids 175-179, 194-210, and 243-255 of a full-length immunoglobulin heavy chain variable region as set forth in amino acids 145-265 of SEQ ID NO:397, wherein the linker comprises amino acids 129-144 of SEQ ID NO:397,
(b) a human IgG1 hinge peptide comprising a mutated core sequence as set forth in amino acids 279-282 of SEQ ID NO:397,
(c) an immunoglobulin heavy chain CH2 constant region polypeptide as set forth in amino acids 284-393 of SEQ ID NO:397 fused to the hinge peptide,
(d) an immunoglobulin heavy chain CH3 constant region polypeptide as set forth in amino acids 394-500 of SEQ ID NO:397 fused to the CH2 polypeptide, and
wherein the fusion protein promotes antibody dependent cell-mediated cytotoxicity, complement fixation, or both.
4. The fusion protein of claim 1 wherein the hinge peptide comprises amino acids 279-282 as set forth in SEQ ID NO:397.
5. The fusion protein of claim 1 wherein the hinge peptide comprises amino acids 279-282 as set forth in SEQ ID NO:398.
6. The fusion protein of claim 1 wherein the hinge peptide comprises amino acids 279-282 as set forth in SEQ ID NO:582.
7. The fusion protein of claim 3 wherein the binding domain polypeptide has a serine at position 11 in the immunoglobulin heavy chain variable region polypeptide.
8. A composition comprising a fusion protein of claim 3 and a pharmaceutically acceptable excipient, diluent, or carrier.