JPWO2020082057A5 - - Google Patents

Claims (22)

An IL-10 polypeptide homodimer of two IL-10 polypeptides each comprising a non-naturally encoded amino acid, wherein said IL-10 polypeptides are homodimers of each IL-10 polypeptide linked via a linker covalently attached to said non-naturally encoded amino acid;
said non-naturally encoded amino acid of each IL-10 polypeptide is incorporated at position 59, 63, 66, 70, or 74;
An IL-10 polypeptide homodimer wherein all references to amino acid positions are based on SEQ ID NO:1 . 2. The IL-10 polypeptide homodimer of claim 1, wherein the amino acid sequence of said IL-10 polypeptide is at least 90% identical to the amino acid sequence of SEQ ID NO:1, SEQ ID NO:2 or SEQ ID NO:5. 3. The IL-10 polypeptide homodimer of claim 2, wherein the amino acid sequence of said IL-10 polypeptide is at least 95%, at least 98%, or at least 99% identical to the amino acid sequence of SEQ ID NO:2. . 4. The IL-10 polypeptide homozygote of any one of claims 1-3 , wherein said linker covalently attached to each said non-naturally encoded amino acid of said IL-10 polypeptide comprises a water -soluble polymer. Dimer . 5. The IL-10 polypeptide homodimer of claim 4 , wherein said water-soluble polymer is PEG. 6. The IL-10 polypeptide homodimer of claim 5 , wherein said PEG has a molecular weight of 0.1 kDa to 100 kDa. 6. The IL-10 polypeptide homodimer of claim 5 , wherein said PEG has a molecular weight of 0.1 kDa to 50 kDa. The IL-10 polypeptide homodimer of any one of claims 5-7, wherein said PEG is linear or branched. IL-10 according to any one of claims 1 to 8 , wherein said non-naturally encoded amino acid comprises a carbonyl group, an aminooxy group, a hydrazine group, a hydrazide group, a semicarbazide group, an azide group or an alkyne group. Polypeptide homodimer . said non-naturally encoded amino acid is para-acetylphenylalanine, p-nitrophenylalanine, p-sulfotyrosine, p-carboxyphenylalanine, o-nitrophenylalanine, m-nitrophenylalanine, p-boronylphenylalanine, o-boronylphenylalanine , m-boronylphenylalanine, p-aminophenylalanine, o-aminophenylalanine, m-aminophenylalanine, p-acylphenylalanine, o-acylphenylalanine, m-acylphenylalanine, p-OMe phenylalanine, o-OMe phenylalanine, m-OMe Phenylalanine, p-sulfophenylalanine, o-sulfophenylalanine, m-sulfophenylalanine, 5-nitroHis, 3-nitroTyr, 2-nitroTyr, nitro-substituted Leu, nitro-substituted His, nitro-substituted De, nitro-substituted Trp, 2 -NitroTrp, 4-NitroTrp, 5-NitroTrp, 6-NitroTrp, 7-NitroTrp, 3-Aminotyrosine, 2-Aminotyrosine, O-Sulfotyrosine, 2-Sulfoxyphenylalanine, 3-Sulfoxyphenylalanine , o-carboxyphenylalanine, m-carboxyphenylalanine, p-acetyl-L-phenylalanine, p-propargyl-phenylalanine, O-methyl-L-tyrosine, L-3-(2-naphthyl)alanine, 3-methyl-phenylalanine, O-4-aryl-L-tyrosine, 4-propyl-L-tyrosine, tri-O-acetyl-GlcNAcβ-serine, L-dopa, fluorinated phenylalanine, isopropyl-L-phenylalanine, p-azido-L- Phenylalanine, p-acyl-L-phenylalanine, p-benzoyl-L-phenylalanine, L-phosphoserine, phosphonoserine, phosphonotyrosine, p-iodo-phenylalanine, p-bromophenylalanine, p-amino-L-phenylalanine, isopropyl-L -phenylalanine , and p- propargyloxy - L-phenylalanine. 11. The IL-10 polypeptide homodimer of claim 10 , wherein said non-naturally encoded amino acid is para-acetyl-phenylalanine or para-azidomethyl-phenylalanine. said non-naturally encoded amino acid is reactive with a linker, polymer or biologically active molecule that is not reactive with any of the 20 common amino acids in polypeptides The IL-10 polypeptide homodimer of any one of claims 1-11 . 13. The IL-10 polypeptide homodimer of any one of claims 4-12, wherein said water-soluble polymer is linked to said IL-10 polypeptide by a linker. 14. The IL-10 polypeptide homodimer of claim 13 , wherein said linker is a cleavable or non-cleavable linker. 15. The IL-10 polypeptide homodimer of claim 13 or 14 , wherein said linker is a bifunctional octyne linker. The IL-10 polypeptide dimer of any one of claims 1 and 4-15, wherein the amino acid sequence of said IL-10 polypeptide is any one of the amino acid sequences of SEQ ID NOS: 10-44. . 2. A method of making the IL-10 polypeptide homodimer of claim 1, comprising: a first isolated IL-10 polypeptide comprising a non-naturally encoded amino acid; and contacting with a second isolated IL -10 polypeptide comprising a non-naturally encoded amino acid of each IL-10 polypeptide via a linker covalently attached to said non-naturally encoded amino acid of each IL-10 polypeptide; A method comprising linking one and a second IL-10 polypeptide. PEGylated IL-10 polypeptide homodimer of the IL-10 polypeptide homodimer of claim 1. The IL-10 polypeptide homodimer according to any one of claims 1 to 16 or the PEGylated IL-10 polypeptide homodimer according to claim 18, and a pharmaceutically acceptable carrier and a composition comprising: 20. The composition of Claim 19, further comprising a chemotherapeutic or immunotherapeutic agent. The IL-10 polypeptide homodimer of any one of claims 1-16, the PEGylated IL-10 polypeptide homodimer of claim 18 or the composition of claim 19 or 20. A pharmaceutical composition for treating cancer, comprising: An IL-10 polypeptide homodimer according to any one of claims 1 to 16, a PEGylated IL-10 polypeptide homodimer according to claim 18 or a composition according to claim 19 or 20. A pharmaceutical composition for treating an immune or inflammatory disease, disorder or condition, comprising: