IgG1 FC monomer and application thereof
An IgG1 Fc monomer, a preparation method therefor and an application thereof. Causing an Fc dimer of a novel IgG1 Fc monomer sequence for which an antibody IgG1 constant region is modified to become an Fc monomer by means of the modification on a human antibody IgG1 constant region Fc that uses antibody engineering technology, and maintaining an FcRn binding function; the present application has the feature of very low non-specific binding with unrelated proteins, and the main features of the Fc monomer comprise the T366, L368, P395, and K409 positions in a CH3 region of the constant region of the antibody having mutations, and the monomer being highly efficiently expressed in prokaryotic cells; the monomer may bind to FcRn by using a pH-dependent specific binding mode, and has the feature of very low non-specific binding. By using said Fc monomer, the same may be fused or coupled with various proteins, polypeptides, small molecules, nucleic acids and the like for different targets such that the fused or coupled molecules have the feature of being capable of binding to FcRn by using pH dependence.
1. An IgG1 Fc monomer polypeptide comprising CH2 and CH3 domains, wherein the polypeptide comprises arginine (R), histidine (H), lysine (K) and threonine (T) at positions 366, 368, 395 and 409 of IgG1, respectively, wherein the numbering is according to EU numbering and the Fc monomer polypeptide is capable of binding to a neonatal Fc receptor (FcRn), wherein the binding affinity of the IgG1 Fc monomer polypeptide to at least one antigen selected from the group consisting of 5T4, PD-L1 and OX40 is lower than that of a wild-type IgG1 Fc, as measured by an enzyme-linked immunosorbent assay (ELISA) with 100 ng of the antigen and up to 223 nM of the IgG1 Fc monomer polypeptide.
2. The IgG1 Fc monomer polypeptide according to claim 1 , further comprising an amino acid substitution at position 428 of IgG1.
3. A fusion protein, comprising the IgG1 Fc monomer polypeptide according to claim 1 and a heterologous protein.
4. The fusion protein of claim 3 , wherein the IgG1 Fc monomer polypeptide further comprises methionine (M) at position 428 of IgG1.
5. The fusion protein of claim 3 , wherein the heterologous protein comprises a heavy chain variable region and a light chain variable region and the heterologous protein is capable of specifically binding to an antigen of interest; the heterologous protein is an antigen, a cytokine, a soluble receptor, a growth factor, a label, a human interferon, an erythropoietin, a soluble tumor necrosis factor receptor, CTLA-4, a soluble IL-4 receptor or a factor I; or the fusion protein comprises a toxin.
6. The fusion protein of claim 3 , wherein the heterologous protein is a cytokine.
7. The fusion protein of claim 4 , wherein the heterologous protein is a cytokine.
8. An IgG1 Fc monomer polypeptide comprising CH2 and CH3 domains, wherein the CH3 domain comprises the amino acid sequence having residues 111 to 217 of SEQ ID NO: 2, which correspond to positions 341 to 447 of IgG1 according to EU numbering, wherein the binding affinity of the IgG1 Fc monomer polypeptide to at least one antigen selected from the group consisting of 5T4, PD-L1 and OX40 is lower than that of a wild-type IgG1 Fc, as measured by an enzyme-linked immunosorbent assay (ELISA) with 100 ng of the antigen and up to 223 nM of the IgG1 Fc monomer polypeptide.
9. A fusion protein, comprising the IgG1 Fc monomer polypeptide according to claim 8 and a heterologous protein.
10. The fusion protein of claim 9 , wherein the heterologous protein comprises a heavy chain variable region and a light chain variable region and the heterologous protein is capable of specifically binding to an antigen of interest; the heterologous protein is an antigen, a cytokine, a soluble receptor, a growth factor, a label, a human interferon, an erythropoietin, a soluble tumor necrosis factor receptor, CTLA-4, a soluble IL-4 receptor or a factor I; or the fusion protein comprises a toxin.
11. An IgG1 Fc monomer polypeptide comprising the amino acid sequence of SEQ ID NO: 2.
12. A fusion protein, comprising the IgG1 Fc monomer polypeptide according to claim 11 and a heterologous protein.
13. The fusion protein of claim 12 , wherein the heterologous protein comprises a heavy chain variable region and a light chain variable region and the heterologous protein is capable of specifically binding to an antigen of interest; the heterologous protein is an antigen, a cytokine, a soluble receptor, a growth factor, a label, a human interferon, an erythropoietin, a soluble tumor necrosis factor receptor, CTLA-4, a soluble IL-4 receptor or a factor I; or the fusion protein comprises a toxin.