Bifunctional Molecules Containing a Variant of IL-7

Abstract

The present invention relates to IL-7 variants, bifunctional molecules containing it and their use.

Claims

1. A bifunctional molecule comprising a variant of interleukin 7 (IL-7) conjugated to a linking group, wherein: the binding group comprises an antagonist antibody that binds human PD-1 or an antagonist human PD-1 binding fragment thereof that contains an antigen-binding domain that binds PD-1, an IL-7 variant comprises at least 90% identity to a wild-type human IL-7 (wth-IL-7) comprising or consisting of the amino acid sequence shown in SEQ ID NO: 1, wherein the variant comprises at least one amino acid mutation selected from the group consisting of (i) W142H, W142F or W142Y, (ii) C2S-C141S and C47S-C92S, C2S-C141S and C34S-C129S, or C47S-C92S and C34S-C129S, (iii) D74E, D74Q or D74N, iv) Q11E, Y12F, M17L, Q22E and / or K81R; or any combination thereof, wherein the amino acid numbering corresponds to that in SEQ ID NO:

1.

2. The bifunctional molecule of claim 1, wherein the antagonist antibody fragment is selected from a Fab, Fab', single-chain variable fragment (scFV) or single-domain antibody (sdAb) domain.

3. The bifunctional molecule of claim 1 or 2, wherein said antagonist antibody binding human PD-1 or antagonist human PD-1 binding fragment thereof is selected from BI-754091, pembrolizumab, nivolumab, pidilizumab, cemiplimab, camrelizumab, AUNP12, AMP-224, AGEN-2034, BGB-A317, PDR001, MK-3477, SCH-900475, PF-06801591, JNJ-63723283, genolimzumab, LZM-009, BCD-100, SHR-1201, BAT-1306, AK-103, MEDI-0680, MEDI0608, JS001, CBT-501, INCSHR1210, TSR-042, GLS-010, AM- 0001, STI-1110, AGEN2034, MGA012 or IBI308, monoclonal antibodies 5C4, 17D8, 2D3, 4H1, 4A11, 7D3 and 5F4, RG7769, XmAb20717, MEDI5752, FS118, SL-279252 and XmAb23104 or an antagonist to human PD-1 binding fragment of any of the above.

4. A bifunctional molecule according to claim 1, 2 or 3, characterized in that the IL-7 variant contains an amino acid substitution selected from the group consisting of W142H, W142F and W142Y, wherein the amino acid numbering corresponds to that in SEQ ID NO:

1.

5. A bifunctional molecule according to any one of claims 1 to 4, characterized in that the IL-7 variant contains a group of amino acid substitutions selected from the group consisting of C2S-C141S and C47S-C92S, C2S-C141S and C34S-C129S, and C47S-C92S and C34S-C129S, wherein the amino acid numbering corresponds to that in SEQ ID NO:

1.

6. A bifunctional molecule according to any one of claims 1 to 5, characterized in that the IL-7 variant contains an amino acid substitution selected from the group consisting of D74E, D74Q and D74N, wherein the amino acid numbering corresponds to that in SEQ ID NO:

1.

7. A bifunctional molecule according to any one of claims 1-6, characterized in that the IL-7 variant comprises or consists of the amino acid sequence represented by SEQ ID NO: 2-15.

8. A bifunctional molecule according to any one of claims 1 to 7, characterized in that the antagonist antibody that binds human PD-1 or the antagonist binding fragment with respect to human PD-1 comprises a heavy chain constant domain.

9. A bifunctional molecule according to claim 8, which comprises the Fc domain of human IgG1.

10. The bifunctional molecule of claim 9, wherein the Fc domain of human IgG1 comprises a substitution or a combination of substitutions selected from the group consisting of T250Q / M428L; M252Y / S254T / T256E + H433K / N434F; E233P / L234V / L235A / G236A + A327G / A330S / P331S; E333A; S239D / A330L / I332E; P257I / Q311; K326W / E333S; S239D / I332E / G236A; N297A; L234A / L235A; N297A + M252Y / S254T / T256E; K322A and K444A, N297A and N297A in combination with M252Y / S254T / T256E and L234A / L235A.

11. A bifunctional molecule according to any one of claims 1 to 7, characterized in that the antagonist antibody that binds human PD-1 or the antagonist binding fragment with respect to human PD-1 comprises the Fc domain of human IgG4.

12. The bifunctional molecule of claim 11, wherein the Fc domain of human IgG4 comprises a substitution or a combination of substitutions selected from the group consisting of S228P; L234A / L235A, S228P + M252Y / S254T / T256E.17 and K444A.

13. A bifunctional molecule according to any one of claims 1 to 12, characterized in that the N-terminus of the IL-7 variant is directly covalently linked to the C-terminus of the constant domain of the heavy or light chain of an antagonist antibody that binds human PD-1, or an antagonist binding fragment with respect to human PD-1, or is linked to said antibody indirectly via a peptide linker.

14. A bifunctional molecule according to claim 13, characterized in that the N-terminus of the IL-7 variant is directly covalently linked to the C-terminus of the constant domain of the heavy chain of an antagonist antibody that binds human PD-1, or an antagonist binding fragment with respect to human PD-1, or is linked to said antibody indirectly via a peptide linker.

15. The bifunctional molecule of claim 13 or 14, wherein the IL-7 variant is fused to an antagonist antibody that binds human PD-1 or an antagonist binding fragment to human PD-1 via a peptide linker selected from the group consisting of GGGGS (SEQ ID NO: 68), GGGGSGGGS (SEQ ID NO: 67), GGGGSGGGGS (SEQ ID NO: 69) and GGGGSGGGGSGGGGS (SEQ ID NO: 70), preferably being (GGGGS)3.

16. A bifunctional molecule according to any one of claims 1 to 15, comprising a variant of interleukin 7 (IL-7) conjugated to an antagonist antibody that binds human PD-1 or an antagonist human PD-1 binding fragment of an antibody that contains an antigen-binding domain of an antibody that binds PD-1, wherein the molecule consists of a first monomer and a second monomer, wherein (i) the first monomer comprises an antagonist to PD-1 antigen-binding domain of an antibody that is directly covalently linked via its C-terminus to the N-terminus of a first heterodimeric Fc chain or indirectly covalently linked via a peptide linker to said, and wherein said first heterodimeric Fc chain is covalently linked via its C-terminus to the N-terminus of an IL-7 variant or indirectly covalently linked via a peptide linker to said, and (ii) the second monomer contains a complementary second heterodimeric Fc chain and does not contain an antigen-binding domain.

17. The bifunctional molecule of claim 16, wherein in the second monomer the complementary second heterodimeric Fc chain is covalently linked directly to the IL-7 variant or is covalently linked indirectly via a peptide linker to the IL-7 variant.

18. The bifunctional molecule of claim 17, wherein in the second monomer the complementary second heteromeric Fc chain is covalently directly linked through its C-terminus to the IL-7 variant or is indirectly covalently linked to said IL-7 variant via a peptide linker.

19. A bifunctional molecule according to any one of claims 1 to 15, comprising a variant of interleukin 7 (IL-7) conjugated to an antagonist antibody that binds PD-1 or an antagonist human PD-1 binding fragment of an antibody that comprises an antigen-binding domain of an antibody that binds PD-1, wherein the molecule consists of a first monomer and a second monomer, wherein (i) the first monomer comprises a PD-1 antigen-binding antagonist domain that is covalently linked via its C-terminus to the N-terminus of a first heterodimeric Fc chain, optionally via a peptide linker, wherein said first heterodimeric Fc chain does not comprise an IL-7 variant, and (ii) the second monomer comprises a complementary second heterodimeric Fc chain and does not comprise an antigen-binding domain, wherein said second heterodimeric Fc chain is directly covalently linked to the IL-7 variant or indirectly via a peptide linker is covalently linked to said.

20. The bifunctional molecule of claim 19, wherein in the second monomer the second heteromeric Fc chain is directly covalently linked through its C-terminus to the N-terminus of the IL-7 variant or is indirectly linked to said via a peptide linker.

21. A bifunctional molecule according to any one of claims 1 to 15, comprising a variant of interleukin 7 (IL-7) conjugated to an antagonist antibody that binds human PD-1, or an antagonist human PD-1 binding fragment of an antibody that contains an antigen-binding domain of an antibody that binds PD-1, wherein the molecule consists of a first monomer and a second monomer, wherein (i) the first monomer comprises a PD-1 antigen binding domain directly covalently linked via its C-terminus to the N-terminus of the first heterodimeric Fc chain or indirectly linked via a peptide linker to said, and (ii) the second monomer comprises a PD-1 antigen binding domain covalently linked via its C-terminus to the N-terminus of a complementary second heterodimeric Fc chain or indirectly linked via a peptide linker to said Only one of the heterodimeric Fc chains is covalently linked via the C-terminus to the N-terminus of the IL-7 variant.

22. The bifunctional molecule of claim 21, wherein only the first heterodimeric Fc chain is covalently linked through its C-terminus to the N-terminus of the IL-7 variant.

23. A bifunctional molecule according to any one of claims 16 to 22, characterized in that the antigen-binding domain to PD-1 is a Fab, Fab', single-chain variable fragment (scFV) or single-domain antibody (sdAb) domain.

24. An isolated nucleic acid that contains nucleic acids that encode for expression a bifunctional molecule according to any one of claims 1 to 23.

25. A host cell containing the isolated nucleic acid of claim 24.

26. A pharmaceutical composition comprising a bifunctional molecule according to any one of claims 1 to 23, a nucleic acid encoding said bifunctional molecule or a host cell containing said nucleic acid, and a pharmaceutically acceptable carrier.

27. The use of a molecule according to any one of claims 1 to 23, a nucleic acid encoding said molecule, or a pharmaceutical composition containing this molecule or this encoding nucleic acid, as a medicament for the treatment of cancer.

28. The use of a molecule according to any one of claims 1 to 23, a nucleic acid encoding said molecule, or a pharmaceutical composition containing this molecule or this encoding nucleic acid, as a medicinal product for the treatment of an infectious disease.

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