Single variable domain immunoglobulin polypeptides targeting IL-13 and TSLP

Claims

1. A polypeptide comprising or consisting of at least one immunoglobulin single variable domain (ISVD) that specifically binds to IL13 or TSLP, wherein said ISVD comprises three complementarity determining regions (CDR1 to CDR3, respectively); and wherein at least one ISVD comprises: a) CDR1, which is the amino acid sequence of SEQ ID NO: 7 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 7; CDR2, which is the amino acid sequence of SEQ ID NO: 12 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 12; and CDR3, which is the amino acid sequence of SEQ ID NO: 17 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 17, b) CDR1, which is the amino acid sequence of SEQ ID NO: 8 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 8; CDR2, which is the amino acid sequence of SEQ ID NO: 13 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 13; and CDR3, which is the amino acid sequence of SEQ ID NO: 18 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 18, c) CDR1, which is the amino acid sequence of SEQ ID NO: 9 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 9; CDR2, which is the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 14; and CDR3, which is the amino acid sequence of SEQ ID NO: 19 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 19, or d) CDR1, which is the amino acid sequence of SEQ ID NO: 11 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 11; CDR2, which is the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 16; and CDR3, which is the amino acid sequence of SEQ ID NO: 21 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO:

21.

2. The polypeptide of claim 1, wherein at least one ISVD that specifically binds to IL-13 or TSLP comprises: a) CDR1, which is the amino acid sequence of SEQ ID NO: 7, CDR2, which is the amino acid sequence of SEQ ID NO: 12, and CDR3, which is the amino acid sequence of SEQ ID NO: 17, b) CDR1, which is the amino acid sequence of SEQ ID NO: 8, CDR2, which is the amino acid sequence of SEQ ID NO: 13, and CDR3, which is the amino acid sequence of SEQ ID NO: 18, c) CDR1, which is the amino acid sequence of SEQ ID NO: 9, CDR2, which is the amino acid sequence of SEQ ID NO: 14, and CDR3, which is the amino acid sequence of SEQ ID NO: 19, or d) CDR1, which is the amino acid sequence of SEQ ID NO: 11, CDR2, which is the amino acid sequence of SEQ ID NO: 16, and CDR3, which is the amino acid sequence of SEQ ID NO:

21.

3. The polypeptide of claim 1 or 2, wherein the amino acid sequence of at least one ISVD that specifically binds to IL-13 or TSLP is characterized by: a) more than 90% sequence identity with SEQ ID NO: 2, b) more than 90% sequence identity with SEQ ID NO: 3, c) more than 90% sequence identity with SEQ ID NO: 4, or d) more than 90% sequence identity with SEQ ID NO:

6.

4. The polypeptide of any one of claims 1-3, wherein said at least one ISVD that specifically binds to IL-13 or TSLP comprises: a) the amino acid sequence of SEQ ID NO: 2, b) the amino acid sequence of SEQ ID NO: 3, c) the amino acid sequence of SEQ ID NO: 4, or d) the amino acid sequence of SEQ ID NO:

6.

5. The polypeptide of claim 1, wherein the polypeptide comprises or consists of at least two ISVDs, wherein each of said ISVDs comprises three complementarity determining regions (CDR1 to CDR3, respectively), wherein the at least two ISVDs are optionally linked via one or more peptide linkers, and wherein: a) The first and second ISVDs specifically bind to IL-13 and contain: i. CDR1, which is the amino acid sequence of SEQ ID NO: 7 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 7; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 12 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 12; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 17 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 17, b) the first and second ISVDs specifically bind to IL-13 and contain: i. CDR1, which is the amino acid sequence of SEQ ID NO: 8 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 8; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 13 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 13; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 18 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 18, c) the first ISVD specifically binds to IL-13 and contains: i. CDR1, which is the amino acid sequence of SEQ ID NO: 7 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 7; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 12 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 12; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 17 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 17, and The second ISVD binds specifically to IL-13 and contains: iv. CDR1, which is the amino acid sequence of SEQ ID NO: 8 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 8; v. CDR2, which is the amino acid sequence of SEQ ID NO: 13 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 13; and vi. CDR3, which is the amino acid sequence of SEQ ID NO: 18 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 18, d) the first ISVD specifically binds to IL-13 and contains: i. CDR1, which is the amino acid sequence of SEQ ID NO: 8 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 8; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 13 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 13; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 18 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 18, and The second ISVD binds specifically to IL-13 and contains: iv. CDR1, which is the amino acid sequence of SEQ ID NO: 7 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 7; v. CDR2, which is the amino acid sequence of SEQ ID NO: 12 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 12; and vi. CDR3, which is the amino acid sequence of SEQ ID NO: 17 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 17, e) the first ISVD specifically binds to TSLP and contains: i. CDR1, which is the amino acid sequence of SEQ ID NO: 11 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 11; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 16; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 21 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 21, and The second ISVD specifically binds to TSLP and contains: iv. CDR1, which is the amino acid sequence of SEQ ID NO: 9 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 9; v. CDR2, which is the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 14; and vi. CDR3, which is the amino acid sequence of SEQ ID NO: 19 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 19, or f) the first ISVD specifically binds to TSLP and contains: i. CDR1, which is the amino acid sequence of SEQ ID NO: 9 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 9; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 14; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 19 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 19, and The second ISVD specifically binds to TSLP and contains: iv. CDR1, which is the amino acid sequence of SEQ ID NO: 11 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 11; v. CDR2, which is the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 16; and vi. CDR3, which is the amino acid sequence of SEQ ID NO: 21 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 21, where the order of the ISVDs indicates their relative positions with respect to each other, considered from the N-terminus to the C-terminus of the said polypeptide.

6. Polypeptide according to claim 5, where: a) the first and second ISVDs that specifically bind to IL-13 comprise CDR1, which is the amino acid sequence of SEQ ID NO: 7, CDR2, which is the amino acid sequence of SEQ ID NO: 12, and CDR3, which is the amino acid sequence of SEQ ID NO: 17, b) the first and second ISVDs that specifically bind to IL-13 comprise CDR1, which is the amino acid sequence of SEQ ID NO: 8, CDR2, which is the amino acid sequence of SEQ ID NO: 13, and CDR3, which is the amino acid sequence of SEQ ID NO: 18, c) the first ISVD that specifically binds to IL-13 comprises CDR1, which is the amino acid sequence of SEQ ID NO: 7, CDR2, which is the amino acid sequence of SEQ ID NO: 12, and CDR3, which is the amino acid sequence of SEQ ID NO: 17, and the second ISVD that specifically binds to IL-13 comprises CDR1, which is the amino acid sequence of SEQ ID NO: 8, CDR2, which is the amino acid sequence of SEQ ID NO: 13, and CDR3, which is the amino acid sequence of SEQ ID NO: 18, d) the first ISVD that specifically binds to IL-13 comprises CDR1, which is the amino acid sequence of SEQ ID NO: 8, CDR2, which is the amino acid sequence of SEQ ID NO: 13, and CDR3, which is the amino acid sequence of SEQ ID NO: 18, and the second ISVD that specifically binds to IL-13 comprises CDR1, which is the amino acid sequence of SEQ ID NO: 7, CDR2, which is the amino acid sequence of SEQ ID NO: 12, and CDR3, which is the amino acid sequence of SEQ ID NO: 17, e) a first ISVD that specifically binds to TSLP comprises a CDR1 that is the amino acid sequence of SEQ ID NO: 11, a CDR2 that is the amino acid sequence of SEQ ID NO: 16, and a CDR3 that is the amino acid sequence of SEQ ID NO: 21, and a second ISVD that specifically binds to TSLP comprises a CDR1 that is the amino acid sequence of SEQ ID NO: 9, a CDR2 that is the amino acid sequence of SEQ ID NO: 14, and a CDR3 that is the amino acid sequence of SEQ ID NO: 19, or f) the first ISVD that specifically binds to TSLP comprises CDR1, which is the amino acid sequence under SEQ ID NO: 9, CDR2, which is the amino acid sequence under SEQ ID NO: 14, and CDR3, which is the amino acid sequence under SEQ ID NO: 19, and the second ISVD that specifically binds to TSLP comprises CDR1, which is the amino acid sequence under SEQ ID NO: 11, CDR2, which is the amino acid sequence under SEQ ID NO: 16, and CDR3, which is the amino acid sequence under SEQ ID NO:

21.

7. The polypeptide according to any one of paragraphs 5 or 6, wherein: a) the amino acid sequences of the first and second ISVDs that specifically bind to IL-13 are characterized by more than 90% sequence identity with SEQ ID NO: 2, b) the amino acid sequences of the first and second ISVDs that specifically bind to IL-13 are characterized by more than 90% sequence identity with SEQ ID NO: 3, c) the amino acid sequence of the first ISVD that specifically binds to IL-13 is characterized by more than 90% sequence identity with SEQ ID NO: 2, and the second ISVD that specifically binds to IL-13 is characterized by more than 90% sequence identity with SEQ ID NO: 3, d) the amino acid sequence of the first ISVD that specifically binds to IL-13 is characterized by more than 90% sequence identity with SEQ ID NO: 3, and the second ISVD that specifically binds to IL-13 is characterized by more than 90% sequence identity with SEQ ID NO: 2, e) the amino acid sequence of the first ISVD that specifically binds to TSLP is characterized by more than 90% sequence identity with SEQ ID NO: 6, and the second ISVD that specifically binds to TSLP is characterized by more than 90% sequence identity with SEQ ID NO: 4, f) the amino acid sequence of the first ISVD that specifically binds to TSLP is characterized by more than 90% sequence identity with SEQ ID NO: 4, and the second ISVD that specifically binds to TSLP is characterized by more than 90% sequence identity with SEQ ID NO:

6.

8. The polypeptide according to any one of paragraphs 5-7, wherein: a) the first and second ISVDs that specifically bind to IL-13 comprise the amino acid sequence of SEQ ID NO: 2, b) the first and second ISVDs that specifically bind to IL-13 comprise the amino acid sequence of SEQ ID NO: 3, c) a first ISVD that specifically binds to IL-13 comprises the amino acid sequence of SEQ ID NO: 2, and a second ISVD that specifically binds to IL-13 comprises the amino acid sequence of SEQ ID NO: 3, d) a first ISVD that specifically binds to IL-13 comprises the amino acid sequence of SEQ ID NO: 3, and a second ISVD that specifically binds to IL-13 comprises the amino acid sequence of SEQ ID NO: 2, e) a first ISVD that specifically binds to TSLP comprises the amino acid sequence of SEQ ID NO: 6, and a second ISVD that specifically binds to TSLP comprises the amino acid sequence of SEQ ID NO: 4, or f) the first ISVD, which specifically binds to TSLP, comprises the amino acid sequence of SEQ ID NO: 4, and the second ISVD, which specifically binds to TSLP, comprises the amino acid sequence of SEQ ID NO:

6.

9. The polypeptide of any one of claims 5-8, wherein the polypeptide comprises: a) the amino acid sequence of SEQ ID NO: 148, b) the amino acid sequence of SEQ ID NO: 149, c) the amino acid sequence of SEQ ID NO: 150, d) the amino acid sequence of SEQ ID NO: 151, e) the amino acid sequence of SEQ ID NO: 152, f) the amino acid sequence of SEQ ID NO: 153, g) the amino acid sequence of SEQ ID NO: 154, h) the amino acid sequence of SEQ ID NO: 155, i) the amino acid sequence of SEQ ID NO: 156, j) the amino acid sequence of SEQ ID NO: 157, k) the amino acid sequence of SEQ ID NO: 158 or l) the amino acid sequence of SEQ ID NO: 159 or consists of them.

10. The polypeptide of any one of claims 1 or 5, wherein the polypeptide comprises or consists of at least four ISVDs, wherein each of said ISVDs comprises three complementarity determining regions (CDR1 to CDR3, respectively), wherein the at least four ISVDs are optionally linked via one or more peptide linkers, and wherein: a) The first ISVD specifically binds to IL-13 and contains: i. CDR1, which is the amino acid sequence of SEQ ID NO: 7 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 7; ii. CDR2, which is the amino acid sequence of SEQ ID NO: 12 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 12; and iii. CDR3, which is the amino acid sequence of SEQ ID NO: 17 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 17; b) the second ISVD specifically binds to IL-13 and contains: iv. CDR1, which is the amino acid sequence of SEQ ID NO: 8 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 8; v. CDR2, which is the amino acid sequence of SEQ ID NO: 13 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 13; and vi. CDR3, which is the amino acid sequence of SEQ ID NO: 18 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 18; c) the third ISVD specifically binds to TSLP and contains: vii. CDR1, which is the amino acid sequence of SEQ ID NO: 9 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 9; viii. CDR2, which is the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 14; and ix. CDR3, which is the amino acid sequence of SEQ ID NO: 19 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 19; and d) the fourth ISVD specifically binds to TSLP and contains: x. CDR1, which is the amino acid sequence of SEQ ID NO: 11 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 11; xi. CDR2, which is the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 16; and xii. CDR3, which is the amino acid sequence of SEQ ID NO: 21 or an amino acid sequence having 2 or 1 amino acid difference from SEQ ID NO: 21, where the order of the ISVDs indicates their relative positions with respect to each other, considered from the N-terminus to the C-terminus of the said polypeptide.