Soluble interferon receptor and its use

Abstract

The present disclosure provides soluble interferon receptors. The methods of the present disclosure can be used to treat or prevent disorders associated with abnormal immune responses.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of U.S. Provisional Application Serial No. 62 / 548,737, filed on August 22, 2017. The entire contents of the above-mentioned provisional patent application are incorporated herein by reference. Background Art

[0003] Systemic lupus erythematosus (SLE) is a chronic, multi-system, autoimmune disease characterized by diverse disease manifestations affecting multiple organs, including skin, CNS, joints, vasculature and kidneys. There is evidence that interferon (IFN) is overproduced in subjects with SLE and causes systemic inflammation characteristic of SLE. In particular, type I interferon is an important cytokine in SLE and is closely related to disease activity and nephritis. Type I interferon is a subset of interferon protein, including IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν; they are all combined with IFN-α receptors (IFNARs) composed of two different polypeptide chains IFNAR1 and IFNAR2. Therefore, there is a need for removing the interferon and / or reducing inflammation in subjects in need. Summary of the invention

[0004] The present disclosure relates in part to soluble interferon receptors, which can bind interferon (e.g., IFN-α, IFN-β). Such soluble interferon receptors can be used to inhibit interferon activity. In some aspects, the soluble interferon receptor of the present disclosure is a heterodimerization construct. In some aspects, the present disclosure relates to soluble interferon receptors useful in the treatment of diseases (e.g., SLE, Sjögren's syndrome) characterized by interferon.

[0005] The present disclosure provides heterodimers that bind to type I interferons, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an interferon receptor 1 (IFNAR1) domain operably coupled to a mutant Fc domain, with or without a linker domain, and wherein the second polypeptide comprises an interferon receptor 2 (IFNAR2) domain operably coupled to a mutant Fc domain, with or without a linker domain.

[0006] In some respects, the disclosure provides a heterodimer in conjunction with type I interferon, it comprises a first polypeptide and a second polypeptide, wherein the first polypeptide is included in an IFNAR1 domain operably coupled to a mutant Fc domain with or without a joint domain, and wherein the second polypeptide is included in an IFNAR2 domain operably coupled to a mutant Fc domain with or without a joint domain. In some respects, the heterodimer of the disclosure (wherein the first and second polypeptides are each directly (without a joint) connected to a mutant Fc domain) has an increased combination with type I interferon relative to a heterodimer in which the first and second polypeptides each comprise a polypeptide joint domain.

[0007] In some aspects, the heterodimers of the present disclosure comprise a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant Fc domain with a linker domain (e.g., a polypeptide linker), and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant Fc domain with a linker domain (e.g., a polypeptide linker). In some aspects, the heterodimers of the present disclosure comprise a first polypeptide, wherein the first polypeptide comprises a polypeptide linker (e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5). In some aspects, the heterodimer of the present disclosure comprises a second polypeptide, wherein the second polypeptide comprises a polypeptide linker (e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5). In some aspects, the polypeptide linker is about 1-50, about 5-40, about 10-30 or about 15-20 amino acids in length. In some aspects, the polypeptide linker is about 20 or less amino acids, about 15 or less amino acids, about 10 or less amino acids or about 5 or less amino acids in length. In some aspects, the polypeptide linker is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 amino acids in length.

[0008] In some aspects, the heterodimers of the present disclosure bind to type I interferons selected from interferon-α (INFα), interferon-β (INFβ), or both INFα and INFβ. In some aspects, type I interferon is INFα. In some aspects, type I interferon is INFβ. In some aspects, type I interferon is both INFα and INFβ. In other aspects, the heterodimers of the present disclosure inhibit the activity of INFα. In some aspects, the heterodimers of the present disclosure inhibit the activity of INFβ. In some aspects, the heterodimers of the present disclosure inhibit the activity of both INFα and INFβ. In some aspects, the heterodimers of the present disclosure inhibit the induction of type I interferon (IFN) gene expression.

[0009] In some aspects, the heterodimer of the present disclosure includes the first polypeptide and the second polypeptide, wherein the first polypeptide is included in the IFNAR1 domain that is operably coupled to the mutant Fc domain with or without a joint domain (e.g., a polypeptide joint), and wherein the second polypeptide is included in the IFNAR2 domain that is operably coupled to the mutant Fc domain with or without a joint domain (e.g., a polypeptide joint). In some aspects, the mutant Fc domain of the first and second polypeptides respectively includes the mutant human immunoglobulin Fc domain. In some aspects, the mutant Fc domain of the first and second polypeptides respectively includes one or more mutations in the CH2 domain. In some aspects, the mutant Fc domain of the first and second polypeptides respectively includes one or more mutations in the CH3 domain. In some aspects, the mutant Fc domain of the first and second polypeptides respectively includes one or more mutations in the CH2 domain and one or more mutations in the CH3 domain. In some aspects, the mutant Fc domain of the first and second polypeptides respectively includes the mutant human IgG1 Fc domain. In some aspects, the mutant Fc domain of the first and second polypeptides respectively includes the mutant human IgG4 domain.

[0010] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n, wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising the mutation Y407T. In some aspects, the first and second polypeptides each comprise a mutant human IgG1 Fc domain. In some aspects, the first and second polypeptides each comprise a mutant human IgG4 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0011] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising the mutation T366Y. In some aspects, the first and second polypeptides each comprise a mutant human IgG1 Fc domain. In some aspects, the first and second polypeptides each comprise a mutant human IgG4 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0012] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n, wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutations T366S, L368A and Y407V. In some aspects, the first and second polypeptides each comprise a mutant human IgG1 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0013] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain comprising a mutant Fc domain comprising mutations T366S, L368A and Y407 with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutation T366W. In some aspects, the first and second polypeptides each comprise a mutant human IgG1 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0014] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n, wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first and second polypeptides each comprise a mutant human IgG1 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0015] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first and second polypeptides each comprise a mutant human IgG1 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0016] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n, wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutations T366S, L368A and Y407V. In some aspects, the first and second polypeptides each comprise a mutant human IgG4 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0017] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain comprising a mutant Fc domain comprising mutations T366S, L368A and Y407V, with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising the mutation T366Y. In some aspects, the first and second polypeptides each comprise a mutant human IgG4 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0018] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n, wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first and second polypeptides each comprise a mutant human IgG4 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0019] In some aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide is comprised in a polypeptide having or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) according to EU numbering) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), and wherein the second polypeptide is comprised in an IFNAR1 domain with or without a linker domain (e.g., a polypeptide linker, e.g., a Gly / Ser linker, e.g., (G4S) n , wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5), operably coupled to an IFNAR2 domain comprising a mutant Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first and second polypeptides each comprise a mutant human IgG4 Fc domain. In some aspects, the first and second polypeptides each comprise a polypeptide linker, wherein the polypeptide linker is a Gly / Ser linker, e.g., (G4S) n , where n is 1-10, 2-5, 1, 2, 3, 4 or 5.

[0020] In any of the aforementioned or related aspects, the heterodimer of the present disclosure comprises a first polypeptide containing an IFNAR1 domain and a second polypeptide containing an IFNAF2 domain, wherein the first polypeptide and the second polypeptide are each operably coupled to a mutant Fc domain comprising one or more mutations (e.g., one or more CH2 mutations, one or more CH3 mutations, or one or more CH2 and CH3 mutations) with or without a joint, wherein relative to a heterodimer comprising a first polypeptide containing an IFNAR1 domain and a second polypeptide containing an IFNAF2 domain (the first polypeptide and the second polypeptide each comprising a wild-type Fc domain (e.g., an Fc domain of the same isotype (e.g., human IgG1 or human IgG4) without the one or more Fc mutations)), the one or more Fc mutations promote, increase or enhance the formation of heterodimers. In some aspects, the mutant Fc domain is a mutant human IgG1 Fc domain. In some aspects, the mutant Fc domain is a mutant human IgG4 Fc domain.

[0021] In any of the aforementioned or related aspects, the heterodimer of the present disclosure comprises a first polypeptide containing an IFNAR1 domain and a second polypeptide containing an IFNAF2 domain, wherein the first polypeptide and the second polypeptide are each operably coupled to a mutant Fc domain comprising one or more mutations (e.g., one or more CH2 mutations, one or more CH3 mutations, or one or more CH2 and CH3 mutations) with or without a joint, wherein the mutant Fc domain of the first polypeptide and / or the mutant Fc domain of the second polypeptide also comprises one or more mutations that promote, increase or enhance the stability of the Fc domain and / or reduce the binding to an Fc receptor. In some aspects, according to EU numbering, mutations are selected from: C220S, C226S, C229S, P238S and P331S, and combinations thereof. In some aspects, mutations include C220S, P238S and P331S. In some aspects, mutations include C220S, C226S, C229S, P238S and P331S. In some aspects, the first polypeptide and the second polypeptide each comprise a mutant Fc domain comprising mutations C220S, P238S and P331S. In some aspects, the first polypeptide and the second polypeptide each comprise a mutant Fc domain comprising mutations C220S, C226S, C229S, P238S and P331S.

[0022] In any of the foregoing or related aspects, the heterodimer of the present disclosure comprises a first polypeptide comprising an IFNAR1 domain and a second polypeptide comprising an IFNAF2 domain, wherein each of the first and second polypeptides is operably coupled to a mutant Fc domain with or without a linker, wherein the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12.

[0023] In some aspects, the present disclosure provides a heterodimer binding to type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation T366Y, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation Y407T. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 107.

[0024] In some aspects, the disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation Y407T, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation T336Y. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 107.

[0025] In some aspects, the present disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation T366W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T366S, L368A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 109.

[0026] In some aspects, the present disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutation T336W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 109.

[0027] In some aspects, the present disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, T366L, K392L and T394W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10.

[0028] In some aspects, the present disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, L351Y, F405A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10.

[0029] In some aspects, the disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising a mutation T366Y, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising a mutation Y407T. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 123.

[0030] In some aspects, the disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising a mutation Y407T, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising a mutation T336Y. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 123.

[0031] In some aspects, the disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T366W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T366S, L368A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain containing the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain containing the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 125.

[0032] In some aspects, the disclosure provides a heterodimer in conjunction with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T366S, L368A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T336W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 125.

[0033] In some aspects, the disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, T366L, K392L and T394W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121.

[0034] In some aspects, the present disclosure provides a heterodimer in combination with type I interferon, comprising a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, L351Y, F405A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121.

[0035] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 107.

[0036] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 107.

[0037] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 109.

[0038] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 109.

[0039] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10.

[0040] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10.

[0041] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 123.

[0042] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 123.

[0043] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 125.

[0044] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 125.

[0045] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121.

[0046] In some aspects, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) according to EU numbering). nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121.

[0047] In other aspects, the present disclosure provides a heterodimer that binds type I interferon, comprising a first polypeptide and a second polypeptide selected from the group consisting of:

[0048] (i) a first polypeptide comprising the amino acid sequence of SEQ ID NO:40 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:43;

[0049] (ii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:41 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:42;

[0050] (iii) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 53 and a second polypeptide comprising the amino acid sequence of SEQ ID NO: 55; and

[0051] (iv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:57 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:59;

[0052] (v) a first polypeptide comprising the amino acid sequence of SEQ ID NO:40 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:55 and SEQ ID NO:59;

[0053] (vi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:43 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:53 and SEQ ID NO:57;

[0054] (vii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:42 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:45 and SEQ ID NO:47;

[0055] (viii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:41 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:49 and SEQ ID NO:51;

[0056] (ix) a first polypeptide comprising the amino acid sequence of SEQ ID NO:53 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:55 and SEQ ID NO:59;

[0057] (x) a first polypeptide comprising the amino acid sequence of SEQ ID NO:55 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:53 and SEQ ID NO:57;

[0058] (xi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:57 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:55 and SEQ ID NO:59;

[0059] (xii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:59 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:53 and SEQ ID NO:57;

[0060] (xiii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:45 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:49 and SEQ ID NO:51;

[0061] (xiv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:47 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:49 and SEQ ID NO:51;

[0062] (xv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:49 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:45 and SEQ ID NO:47;

[0063] (xvi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:51 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:45 and SEQ ID NO:47;

[0064] (xvii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:61 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:75 and SEQ ID NO:82;

[0065] (xviii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:63 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:65, SEQ ID NO:73 and SEQ ID NO:81;

[0066] (xix) a first polypeptide comprising the amino acid sequence of SEQ ID NO:65 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:63, SEQ ID NO:71 and SEQ ID NO:79;

[0067] (xx) a first polypeptide comprising the amino acid sequence of SEQ ID NO:67 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:61, SEQ ID NO:69 and SEQ ID NO:77;

[0068] (xxi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:69 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:75 and SEQ ID NO:82;

[0069] (xxii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:71 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:65, SEQ ID NO:73 and SEQ ID NO:81;

[0070] (xxiii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:73 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:63, SEQ ID NO:71 and SEQ ID NO:79;

[0071] (xxiv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:75 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:61, SEQ ID NO:69 and SEQ ID NO:77;

[0072] (xxv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:77 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:75 and SEQ ID NO:82;

[0073] (xxvi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:79 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:65, SEQ ID NO:73 and SEQ ID NO:81;

[0074] (xxvii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:81 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:63, SEQ ID NO:71 and SEQ ID NO:79;

[0075] (xxviii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:82 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:61, SEQ ID NO:69 and SEQ ID NO:77;

[0076] (xxix) a first polypeptide comprising the amino acid sequence of SEQ ID NO:84 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:97 and SEQ ID NO:105;

[0077] (xxx) a first polypeptide comprising the amino acid sequence of SEQ ID NO:85 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:87, SEQ ID NO:95 and SEQ ID NO:103;

[0078] (xxxi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:87 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:85, SEQ ID NO:93 and SEQ ID NO:101;

[0079] (xxxii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:89 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:84, SEQ ID NO:91 and SEQ ID NO:99;

[0080] (xxxiii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:91 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:97 and SEQ ID NO:105;

[0081] (xxxiv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:93 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:87, SEQ ID NO:95 and SEQ ID NO:103;

[0082] (xxxv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:95 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:85, SEQ ID NO:93 and SEQ ID NO:101;

[0083] (xxxvi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:97 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:84, SEQ ID NO:91 and SEQ ID NO:99;

[0084] (xxxvii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:99 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:97 and SEQ ID NO:105;

[0085] (xxxviii) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 101 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 87, SEQ ID NO: 95 and SEQ ID NO: 103;

[0086] (xxxix) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 103 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 85, SEQ ID NO: 93 and SEQ ID NO: 101; and

[0087] (xl) a first polypeptide comprising the amino acid sequence of SEQ ID NO:105 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:84, SEQ ID NO:91 and SEQ ID NO:99.

[0088] In some aspects, the disclosure provides a type I interferon-binding heterodimer comprising a first polypeptide comprising the amino acid sequence of SEQ ID NO:40 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:43.

[0089] In some aspects, the disclosure provides a type I interferon-binding heterodimer comprising a first polypeptide comprising the amino acid sequence of SEQ ID NO:41 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:42.

[0090] In some aspects, the present disclosure provides a type I interferon-binding heterodimer comprising a first polypeptide comprising the amino acid sequence of SEQ ID NO:53 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:55.

[0091] In some aspects, the disclosure provides a type I interferon-binding heterodimer comprising a first polypeptide comprising the amino acid sequence of SEQ ID NO:57 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:59.

[0092] In any of the aforementioned or related aspects, the present disclosure provides a heterodimer in combination with type 1 interferon, wherein type I interferon is selected from interferon-α (INFα), interferon-β (INFβ) or both INFα and INFβ. In some aspects, type I interferon is INFα. In some aspects, type I interferon is INFβ. In some aspects, type I interferon is both INFα and INFβ. In some aspects, heterodimers inhibit the activity of INFα. In some aspects, heterodimers inhibit the activity of INFβ. In some aspects, heterodimers inhibit the activity of both INFα and INFβ. In some aspects, heterodimers inhibit the induction of type I interferon (IFN) gene expression.

[0093] In any of the foregoing or related aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first and a second polypeptide, wherein the first and second polypeptides are each directly (without a linker) linked to a mutant Fc domain, and wherein the heterodimer has increased binding to type I interferon relative to a heterodimer in which the first and second polypeptides each comprise a polypeptide linker domain.

[0094] In any of the foregoing or related aspects, the present disclosure provides a heterodimer that binds to type 1 interferon, comprising a first polypeptide comprising an IFNAR1 domain and a second polypeptide comprising an IFNAF2 domain, wherein the first and second polypeptides are each operably coupled to a mutant Fc domain comprising one or more mutations (e.g., one or more CH2 mutations, one or more CH3 mutations, or one or more CH2 and CH3 mutations), and wherein the one or more Fc mutations promote, increase or enhance the formation of the heterodimer relative to a heterodimer comprising a first polypeptide comprising an IFNAR1 domain and a second polypeptide comprising an IFNAF2 domain, each of which comprises a wild-type Fc domain.

[0095] In any of the aforementioned or related aspects, the heterodimer of the present disclosure comprises a first polypeptide containing an IFNAR1 domain and a second polypeptide containing an IFNAF2 domain, wherein the first polypeptide and the second polypeptide are each operably coupled to a mutant Fc domain comprising one or more mutations (e.g., one or more CH2 mutations, one or more CH3 mutations, or one or more CH2 and CH3 mutations) with or without a joint, wherein the mutant Fc domain of the first polypeptide and / or the mutant Fc domain of the second polypeptide also comprises one or more mutations that promote, increase or enhance the stability of the Fc domain and / or reduce the binding to an Fc receptor. In some aspects, according to EU numbering, mutations are selected from: C220S, C226S, C229S, P238S and P331S, and combinations thereof. In some aspects, mutations include C220S, P238S and P331S. In some aspects, mutations include C220S, C226S, C229S, P238S and P331S. In some aspects, the first polypeptide and the second polypeptide each comprise a mutant Fc domain comprising mutations C220S, P238S and P331S. In some aspects, the first polypeptide and the second polypeptide each comprise a mutant Fc domain comprising mutations C220S, C226S, C229S, P238S and P331S.

[0096] In other aspects, the present disclosure provides compositions comprising a heterodimer of the present disclosure and a pharmaceutically acceptable carrier.

[0097] In other aspects, the present disclosure provides a nucleic acid comprising a nucleotide sequence encoding a first polypeptide of a heterodimer, wherein the first polypeptide is comprised in a polypeptide having or without a linker (e.g., a Gly / Ser linker (e.g., (G4S) n A linker, wherein n is 1-5, 1, 2, 3, 4 or 5) is operably coupled to an IFNAR1 domain of a mutant Fc domain (e.g., a mutant human IgG1 Fc domain or a mutant human IgG4 Fc domain). In some aspects, the first polypeptide comprising an IFNAR1 domain comprises the amino acid sequence in SEQ ID NO: 11.

[0098] In other aspects, the present disclosure provides a nucleic acid encoding a nucleotide sequence of a second polypeptide of the heterodimer, wherein the second polypeptide is comprised in a polypeptide having or without a linker (e.g., a Gly / Ser linker (e.g., (G4S) n A linker, wherein n is 1-5, 1, 2, 3, 4 or 5) is operably coupled to an IFNAR2 domain of a mutant Fc domain (e.g., a mutant human IgG1 Fc domain or a mutant human IgG4 Fc domain). In some aspects, the second polypeptide comprising the IFNAR2 domain comprises the amino acid sequence in SEQ ID NO: 12.

[0099] Other aspects of the present disclosure provide recombinant expression vectors and host cells comprising nucleic acids of the present disclosure. In some aspects, the recombinant expression vectors and host cells comprise a nucleic acid comprising a nucleotide sequence encoding a first polypeptide of a heterodimer, wherein the first polypeptide is comprised in a polypeptide having or without a linker (e.g., a Gly / Ser linker (e.g., (G4S) n linker, wherein n is 1-5, 1, 2, 3, 4 or 5) operably coupled to an IFNAR1 domain of a mutant Fc domain (e.g., a mutant human IgG1 Fc domain or a mutant human IgG4 Fc domain). In some aspects, the recombinant expression vector and host cell comprise a nucleic acid comprising a nucleotide sequence encoding a second polypeptide of the heterodimer, wherein the second polypeptide is comprised in a polypeptide having or without a linker (e.g., a Gly / Ser linker (e.g., (G4S) n In some aspects, the recombinant expression vector and host cell comprise a nucleic acid containing a nucleotide sequence of a first polypeptide encoding a heterodimer and a nucleic acid containing a nucleotide sequence of a second polypeptide encoding a heterodimer. In some aspects, the first polypeptide comprising the IFNAR1 domain comprises the amino acid sequence in SEQ ID NO:11. In some aspects, the second polypeptide comprising the IFNAR2 domain comprises the amino acid sequence in SEQ ID NO:12.

[0100] Other aspects of the present disclosure provide methods of reducing, decreasing or inhibiting interferon (IFN) gene expression in a subject in need thereof, comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier.

[0101] Other aspects of the present disclosure provide methods for reducing, lowering or inhibiting type I interferon in a subject in need, including administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier. In some aspects, type I interferon is INFα. In some aspects, type I interferon is INFβ. In some aspects, type I interferon is both INFα and INFβ.

[0102] Other aspects of the present disclosure provide a method for treating a disease characterized by type I interferon or inhibiting its progress and a method for treating an autoimmune disease of a subject in need, including administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier. In some aspects, type I interferon is INFα. In some aspects, type I interferon is INFβ. In some aspects, type I interferon is both INFα and INFβ. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0103] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation T366Y, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising a mutation Y407T. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 107. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0104] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutation Y407T, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutation T336Y. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 107. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0105] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutation T366W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T366S, L368A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 109. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0106] In some aspects, the disclosure provides a method of reducing, reducing or inhibiting interferon (IFN) gene expression, a method of reducing, reducing or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutation T336W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 109. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0107] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, T366L, K392L and T394W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0108] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, L351Y, F405A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0109] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising a mutation T366Y, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising a mutation Y407T. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 123. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0110] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutation Y407T, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T336Y. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 123. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0111] In some aspects, the disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T366W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T366S, L368A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 125. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0112] In some aspects, the disclosure provides a method for reducing, reducing or inhibiting interferon (IFN) gene expression, a method for reducing, reducing or inhibiting type I interferon (e.g., INFα, INFβ or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) of a subject in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T366S, L368A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutation T336W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 125. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome. In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, T366L, K392L and T394W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, L351Y, F405A and Y407V, according to EU numbering. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence set forth in SEQ ID NO: 121.In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0113] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises an IFNAR1 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, L351Y, F405A and Y407V, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0114] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO:106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO:107. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0115] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO:106. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO:107. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0116] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG1 Fc domain comprising a mutation T366W, and the second polypeptide comprises an IFNAR2 domain comprising a mutant IgG1 Fc domain comprising a mutation T366S, L368A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 109. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0117] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and the second polypeptide comprises an IFNAR2 domain comprising a mutant IgG1 Fc domain comprising mutation T336W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 108. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 109. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0118] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG1 Fc domain comprising mutations T350V, T366L, K392L and T394W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG1 Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 10. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0119] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO:9. In some aspects, the mutant IgG1 Fc domain comprises the amino acid sequence shown in SEQ ID NO:10. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0120] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG4 Fc domain comprising a mutation T366Y, and the second polypeptide comprises an IFNAR2 domain comprising a mutant IgG4 Fc domain comprising a mutation Y407T. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:123. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0121] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG4 Fc domain comprising a mutation Y407T, and the second polypeptide comprises an IFNAR2 domain comprising a mutant IgG4 Fc domain comprising a mutation T336Y. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:122. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:123. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0122] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG4 Fc domain comprising a mutation T366W, and the second polypeptide comprises an IFNAR2 domain comprising a mutant IgG4 Fc domain comprising a mutation T366S, L368A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:125. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0123] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) nIn some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG4 Fc domain comprising mutations T366S, L368A and Y407V, and the second polypeptide comprises an IFNAR2 domain comprising a mutant IgG4 Fc domain comprising mutation T336W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO:11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO:12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:124. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO:125. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0124] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG4 Fc domain comprising mutations T350V, T366L, K392L and T394W, and wherein the second polypeptide comprises an IFNAR2 domain operably coupled to a mutant IgG4 Fc domain comprising mutations T350V, L351Y, F405A and Y407V. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0125] In some aspects, the present disclosure provides a method for reducing, lowering or inhibiting interferon (IFN) gene expression, a method for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method for treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein according to EU numbering, the first polypeptide comprises a polypeptide having a Gly / Ser linker (e.g., (G4S) n In some aspects, the first polypeptide comprises an IFNAR1 domain comprising a mutant IgG4 Fc domain comprising mutations T350V, T366L, K392L and T394W. In some aspects, the first polypeptide comprises an IFNAR1 domain comprising the amino acid sequence in SEQ ID NO: 11 and the second polypeptide comprises an IFNAR2 domain comprising the amino acid sequence in SEQ ID NO: 12. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 120. In some aspects, the mutant IgG4 Fc domain comprises the amino acid sequence shown in SEQ ID NO: 121. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0126] In some aspects, the present disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need thereof, the method comprising administering to the subject a heterodimer of the present disclosure or a composition of the present disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide and a second polypeptide selected from the following:

[0127] (i) a first polypeptide comprising the amino acid sequence of SEQ ID NO:40 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:43;

[0128] (ii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:41 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:42;

[0129] (iii) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 53 and a second polypeptide comprising the amino acid sequence of SEQ ID NO: 55; and

[0130] (iv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:57 and a second polypeptide comprising the amino acid sequence of SEQ ID NO:59;

[0131] (v) a first polypeptide comprising the amino acid sequence of SEQ ID NO:40 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:55 and SEQ ID NO:59;

[0132] (vi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:43 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:53 and SEQ ID NO:57;

[0133] (vii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:42 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:45 and SEQ ID NO:47;

[0134] (viii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:41 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:49 and SEQ ID NO:51;

[0135] (ix) a first polypeptide comprising the amino acid sequence of SEQ ID NO:53 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:55 and SEQ ID NO:59;

[0136] (x) a first polypeptide comprising the amino acid sequence of SEQ ID NO:55 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:53 and SEQ ID NO:57;

[0137] (xi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:57 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:55 and SEQ ID NO:59;

[0138] (xii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:59 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:40, SEQ ID NO:53 and SEQ ID NO:57;

[0139] (xiii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:45 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:49 and SEQ ID NO:51;

[0140] (xiv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:47 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:42, SEQ ID NO:49 and SEQ ID NO:51;

[0141] (xv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:49 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:45 and SEQ ID NO:47;

[0142] (xvi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:51 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:41, SEQ ID NO:45 and SEQ ID NO:47;

[0143] (xvii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:61 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:75 and SEQ ID NO:82;

[0144] (xviii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:63 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:65, SEQ ID NO:73 and SEQ ID NO:81;

[0145] (xix) a first polypeptide comprising the amino acid sequence of SEQ ID NO:65 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:63, SEQ ID NO:71 and SEQ ID NO:79;

[0146] (xx) a first polypeptide comprising the amino acid sequence of SEQ ID NO:67 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:61, SEQ ID NO:69 and SEQ ID NO:77;

[0147] (xxi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:69 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:75 and SEQ ID NO:82;

[0148] (xxii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:71 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:65, SEQ ID NO:73 and SEQ ID NO:81;

[0149] (xxiii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:73 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:63, SEQ ID NO:71 and SEQ ID NO:79;

[0150] (xxiv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:75 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:61, SEQ ID NO:69 and SEQ ID NO:77;

[0151] (xxv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:77 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:67, SEQ ID NO:75 and SEQ ID NO:82;

[0152] (xxvi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:79 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:65, SEQ ID NO:73 and SEQ ID NO:81;

[0153] (xxvii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:81 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:63, SEQ ID NO:71 and SEQ ID NO:79;

[0154] (xxviii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:82 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:61, SEQ ID NO:69 and SEQ ID NO:77;

[0155] (xxix) a first polypeptide comprising the amino acid sequence of SEQ ID NO:84 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:97 and SEQ ID NO:105;

[0156] (xxx) a first polypeptide comprising the amino acid sequence of SEQ ID NO:85 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:87, SEQ ID NO:95 and SEQ ID NO:103;

[0157] (xxxi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:87 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:85, SEQ ID NO:93 and SEQ ID NO:101;

[0158] (xxxii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:89 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:84, SEQ ID NO:91 and SEQ ID NO:99;

[0159] (xxxiii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:91 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:97 and SEQ ID NO:105;

[0160] (xxxiv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:93 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:87, SEQ ID NO:95 and SEQ ID NO:103;

[0161] (xxxv) a first polypeptide comprising the amino acid sequence of SEQ ID NO:95 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:85, SEQ ID NO:93 and SEQ ID NO:101;

[0162] (xxxvi) a first polypeptide comprising the amino acid sequence of SEQ ID NO:97 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:84, SEQ ID NO:91 and SEQ ID NO:99;

[0163] (xxxvii) a first polypeptide comprising the amino acid sequence of SEQ ID NO:99 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO:89, SEQ ID NO:97 and SEQ ID NO:105;

[0164] (xxxviii) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 101 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 87, SEQ ID NO: 95 and SEQ ID NO: 103;

[0165] (xxxix) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 103 and a second polypeptide comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 85, SEQ ID NO: 93 and SEQ ID NO: 101; and

[0166] (xl) a first polypeptide comprising the amino acid sequence of SEQ ID NO: 105 and a second polypeptide comprising an amino acid sequence selected from SEQ ID NO: 84, SEQ ID NO: 91 and SEQ ID NO: 99. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0167] In some aspects, the disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide comprising an amino acid sequence of SEQ ID NO: 40 and a second polypeptide comprising an amino acid sequence of SEQ ID NO: 43. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0168] In some aspects, the disclosure provides a method of reducing, lowering or inhibiting interferon (IFN) gene expression, a method of reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and a method of treating an autoimmune disease (e.g., SLE, Sjögren's syndrome) in a subject in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide comprising an amino acid sequence of SEQ ID NO: 41 and a second polypeptide comprising an amino acid sequence of SEQ ID NO: 42. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0169] In some aspects, the disclosure provides methods for reducing, lowering or inhibiting interferon (IFN) gene expression, methods for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and methods for treating autoimmune diseases (e.g., SLE, Sjögren's syndrome) in subjects in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide comprising an amino acid sequence of SEQ ID NO: 53 and a second polypeptide comprising an amino acid sequence of SEQ ID NO: 55. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0170] In some aspects, the disclosure provides methods for reducing, lowering or inhibiting interferon (IFN) gene expression, methods for reducing, lowering or inhibiting type I interferon (e.g., INFα, INFβ, or both INFα and INFβ), and methods for treating autoimmune diseases (e.g., SLE, Sjögren's syndrome) in subjects in need, the method comprising administering to the subject a heterodimer of the disclosure or a composition of the disclosure comprising a heterodimer and a pharmaceutically acceptable carrier, wherein the heterodimer comprises a first polypeptide comprising an amino acid sequence of SEQ ID NO: 57 and a second polypeptide comprising an amino acid sequence of SEQ ID NO: 59. In some aspects, the autoimmune disease is SLE. In some aspects, the autoimmune disease is Sjögren's syndrome.

[0171] Other aspects of the present disclosure provide use of the heterodimer of the present disclosure and an optional pharmaceutically acceptable carrier in the preparation of a medicament for treating an autoimmune disease in a subject in need thereof or delaying its progression, wherein the treatment comprises administering the medicament to a subject in need thereof.

[0172] Other aspects of the disclosure provide kits including a medicament comprising a heterodimer of the disclosure and optionally a pharmaceutically acceptable carrier, and a package insert comprising instructions for administering the medicament to treat or delay progression of an autoimmune disease in a subject in need thereof.

[0173] In some aspects, the disclosure provides kits comprising a container comprising a heterodimer of the disclosure and optionally a pharmaceutically acceptable carrier, and a package insert comprising instructions for administering the heterodimer to treat or delay progression of an autoimmune disease in a subject in need thereof.

[0174] BRIEF DESCRIPTION OF THE DRAWINGS

[0175] These and other features, aspects and advantages of the present invention will be better understood with reference to the following description and accompanying drawings, in which:

[0176] Figure 1 Depicted are exemplary soluble interferon receptors RSLV 601-604, RSLV 602-603, RSLV 606-611, and RSLV 608-613.

[0177] Figure 2 Graph depicts inhibition of IFNα-induced SEAP production in HEK-Blue α / β cells by inhibitors RSLV 601-604, RSLV 602-603 and anti-human IFNα positive control. Human IgG was used as negative control.

[0178] Figure 3 Graph depicts inhibition of IFNβ-induced SEAP production in HEK-Blue α / β cells by inhibitors RSLV 601-604, RSLV 602-603 and anti-human IFNβ positive control. Human IgG was used as a negative control.

[0179] Figure 4 Graph depicts the effect of linker length on inhibition of IFNα-induced SEAP production in HEK-Blue α / β cells.

[0180] Figure 5 Graph depicts the effect of linker length on inhibition of IFNβ-induced SEAP production in HEK-Blueα / β cells.

[0181] Figure 6 Inhibition of SLE serum-induced interferon gene expression in PBMCs by the soluble interferon receptor RSLV 601-604 is depicted.

[0182] Figure 7 Depicted is the inhibition of SLE serum-induced interferon gene expression in PBMCs by the soluble interferon receptor RSLV 608-613. DETAILED DESCRIPTION

[0183] The present disclosure provides soluble heterodimers that bind to type I interferons and methods for reducing, lowering or inhibiting interferon (IFN) gene expression, methods for reducing, lowering or inhibiting type I interferons and / or type I interferon activity (e.g., INFα, INFβ, or both INFα and INFβ), and methods for treating a disease characterized by type I interferon production in a subject in need thereof.

[0184] The present disclosure is based at least in part on the following findings: heterodimers comprising an IFNAR1 domain and an IFNAR2 domain, each operably coupled to a mutant Fc domain, effectively inhibit IFNα and IFNβ activity and inhibit interferon gene expression induced by SLE serum. These results indicate that the heterodimers of the present disclosure are capable of binding to type I interferons in subjects with autoimmune diseases such as SLE and inhibiting their activity.

[0185] It is also found that, relative to the heterodimers in which IFNAR1 domain and IFNAR2 domain are each operably coupled to the mutant Fc domain in the case of having a polypeptide linker, the heterodimers comprising the IFNAR1 domain and IFNAR2 domains each operably directly coupled (i.e., without a polypeptide linker) to the mutant Fc domain have increased binding affinity and effectiveness to type I interferon such as INFα. Not bound by theory, it is believed that IFNAR1 and IRNAR2 require a certain degree of flexibility to form a pocket for interacting with type I interferon. The extracellular domains of known IFNAR1 and IFNAR2 bind type I interferon and form a ternary complex, which causes the intracellular domain to be close to the receptor and regulate the signal transduction by the receptor (Li et al., J.Mol.Biol. (2017) 429, 2571-2589). It has also been shown that ligand-induced conformational changes in receptor components propagate to the membrane-proximal Ig domain of IFNAR1, although this domain does not interact with the ligand (ibid., p. 2572). Notably, the crystal structure of the heterotrimeric complex of IFNAR1, IFNAR2, and IFNα2 depicts a linker-like domain between the extracellular and transmembrane domains of the two IFNARs (see Li et al., 2572). Figure 1 ; See also Piehler et al., Immunological Reviews (2012) 250, 317-334, Figure 5 ). Not bound by theory, it is assumed that the joint-like domain can provide a certain degree of flexibility to facilitate the formation of a ternary complex with an appropriate conformation for ligand binding and signal transduction. Therefore, a study was designed to investigate the impact of various polypeptide linker lengths on IFNAR1 and IFNAR2 ligand binding. Unexpectedly, it was found that the heterodimer of the IFNAR1 domain and the IFNAR2 domain comprising each directly coupled to the mutant Fc domain without a joint, relative to the heterodimer of the IFNAR1 domain and the IFNAR2 domain each being operably coupled to the mutant Fc domain with a polypeptide linker (such as a length of 10 or 20 amino acids), demonstrated the increased binding affinity and effectiveness of type I interferon. In particular, when comparing constructs with different polypeptide linker lengths, it was observed that IFN-α binding affinity and effectiveness increased as the linker length decreased.

[0186] Therefore, the present disclosure provides a heterodimer that binds to type I interferon, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an IFNAR1 domain that is operably coupled to a mutant Fc domain with or without a joint domain, and wherein the second polypeptide comprises an IFNAR2 domain that is operably coupled to a mutant Fc domain with or without a joint domain. As described herein, the heterodimer of the present disclosure can be used in methods for inhibiting type I interferon gene expression, methods for inhibiting type I interferon activity, and methods for treating autoimmune diseases.

[0187] definition

[0188] Unless otherwise stated, the terms used in the claims and specification are defined as follows.

[0189] "Amino acid" refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code, as well as those amino acids that are later modified, such as hydroxyproline, γ-carboxyglutamate, and O-phosphoserine. Amino acid analogs refer to compounds that have the same basic chemical structure as naturally occurring amino acids, i.e., an alpha carbon bound to a hydrogen, a carboxyl group, an amino group, and an R group, such as homoserine, norleucine, methionine sulfoxide, methionine methylsulfonium. Such analogs have modified R groups (e.g., norleucine) or modified peptide backbones, but retain the same basic chemical structure as naturally occurring amino acids. Amino acid mimetics refer to chemical compounds that have a structure that is different from the general chemical structure of an amino acid but function in a manner similar to naturally occurring amino acids.

[0190] Amino acids may be referred to herein by either their commonly known three letter symbols or the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Likewise, nucleotides may be referred to by their commonly recognized single-letter codes.

[0191] "Amino acid substitution" refers to the replacement of at least one existing amino acid residue in a predetermined amino acid sequence (the amino acid sequence of the starting polypeptide) by a second, different "replacement" amino acid residue. "Amino acid insertion" refers to the integration of at least one additional amino acid into a predetermined amino acid sequence. Although insertions typically consist of the insertion of one or two amino acid residues, larger "peptide insertions" may be performed, such as insertions of about three to about five or even up to about ten, fifteen or twenty amino acid residues. The inserted residues may be naturally occurring or non-naturally occurring, as disclosed above. "Amino acid deletion" refers to the removal of at least one amino acid residue from a predetermined amino acid sequence.

[0192] "Polypeptide", "peptide" and "protein" are used interchangeably herein to refer to a polymer of amino acid residues. The term applies to amino acid polymers in which one or more amino acid residues is an artificial chemical mimetic of the corresponding naturally occurring amino acid, as well as naturally occurring amino acid polymers and non-naturally occurring amino acid polymers.

[0193] The term "nucleic acid" refers to deoxyribonucleotides or ribonucleotides and polymers thereof in single-stranded or double-stranded form, unless otherwise specifically limited, and includes nucleic acids containing known analogs of natural nucleotides, the analogs having binding properties similar to reference nucleic acids and metabolized in a manner similar to naturally occurring nucleotides. Unless otherwise indicated, a specific nucleic acid sequence also implicitly includes conservatively modified variants thereof (e.g., degenerate codon substitutions) and complementary sequences as well as sequences explicitly indicated. Specifically, degenerate codon substitutions can be achieved by generating sequences in which the third position of one or more selected (or all) codons is replaced by mixed bases and / or deoxyinosine residues (Batzer et al., Nucleic Acid Res 1991; 19: 5081; Ohtsuka et al., JBC 1985; 260: 2605-8; Rossolini et al., Mol Cell Probes 1994; 8: 91-8). For arginine and leucine, the modification at the second base can also be conservative. The term nucleic acid is used interchangeably with gene, cDNA, and mRNA encoded by a gene.

[0194] The polynucleotide of the present invention can be composed of any polyribonucleotide or polydeoxyribonucleotide, which can be unmodified RNA or DNA or modified RNA or DNA. For example, the polynucleotide can be composed of single-stranded and double-stranded DNA, DNA as a mixture of single-stranded and double-stranded regions, single-stranded and double-stranded RNA, and RNA as a mixture of single-stranded and double-stranded regions, or comprise a hybrid molecule of DNA and RNA that can be single-stranded or more generally double-stranded or a mixture of single-stranded and double-stranded regions. In addition, the polynucleotide can be composed of a triple-stranded region comprising RNA or DNA or both RNA and DNA. The polynucleotide can also include one or more modified bases or DNA or RNA backbones modified for stability or for other reasons."Modified" bases include, for example, tritylated bases and uncommon bases such as inosine. DNA and RNA can be subjected to a variety of modifications; therefore, "polynucleotides" include chemical, enzymatic or metabolically modified forms.

[0195] As used herein, the term "operably linked" or "operably coupled" refers to a juxtaposition wherein the components described are in a relationship permitting them to function in their intended manner.

[0196] As used herein, the term "glycosylation" or "glycosylated" refers to the process or result of adding a sugar moiety to a molecule.

[0197] As used herein, the term "altered glycosylation" refers to aglycosylated, deglycosylated, or hypoglycosylated molecules.

[0198] As used herein, "glycosylation site" refers to sites that can potentially accept carbohydrate moieties as well as sites within a protein to which carbohydrate moieties have actually been attached, and includes any amino acid sequence that can act as an acceptor for oligosaccharides and / or carbohydrates.

[0199] As used herein, the term "aglycosylation" or "aglycosylated" refers to the production of a molecule in an unglycosylated form (e.g., by engineering a protein or polypeptide to lack amino acid residues that act as glycosylation acceptors). Alternatively, a protein or polypeptide can be expressed in, for example, E. coli to produce an aglycosylated protein or polypeptide.

[0200] As used herein, the term "deglycosylation" or "deglycosylated" refers to the process or result of enzymatic removal of sugar moieties on a molecule.

[0201] As used herein, the term "hypoglycosylation" or "hypoglycosylated" refers to a molecule in which one or more carbohydrate structures normally present when produced in a mammalian cell have been omitted, removed, modified or masked.

[0202] As used herein, the terms "Fc region" and "Fc domain" are portions of natural immunoglobulins formed by the corresponding Fc domains (or Fc portions) of the two heavy chains of an immunoglobulin without the variable regions that bind to an antigen. In some embodiments, the Fc domain begins at the hinge region just upstream of the papain cleavage site and ends at the C-terminus of the antibody. Thus, a complete Fc domain comprises at least a hinge domain, a CH2 domain, and a CH3 domain. In certain embodiments, the Fc domain comprises at least one of a hinge (e.g., an upper, middle, and / or lower hinge region) domain, a CH2 domain, a CH3 domain, a CH4 domain, or a variant, portion, or fragment thereof. In other embodiments, the Fc domain comprises a complete Fc domain (i.e., a hinge domain, a CH2 domain, and a CH3 domain). In one embodiment, the Fc domain comprises a hinge domain (or a portion thereof) fused to a CH3 domain (or a portion thereof). In another embodiment, the Fc domain comprises a CH2 domain (or a portion thereof) fused to a CH3 domain (or a portion thereof). In another embodiment, the Fc domain consists of a CH3 domain or a portion thereof. In another embodiment, the Fc domain consists of a hinge domain (or a portion thereof) and a CH3 domain (or a portion thereof). In another embodiment, the Fc domain consists of a CH2 domain (or a portion thereof) and a CH3 domain. In another embodiment, the Fc domain consists of a hinge domain (or a portion thereof) and a CH2 domain (or a portion thereof). In one embodiment, the Fc domain lacks at least a portion of the CH2 domain (e.g., all or part of the CH2 domain). In one embodiment, the Fc domain of the present invention comprises at least a portion of the Fc molecule required for FcRn binding known in the art. In one embodiment, the Fc domain of the present invention comprises at least a portion of the Fc molecule required for protein A binding known in the art. In one embodiment, the Fc domain of the present invention comprises at least a portion of the Fc molecule required for protein G binding known in the art. The Fc domain herein generally refers to a polypeptide comprising all or part of the Fc domain of an immunoglobulin heavy chain. This includes, but is not limited to, polypeptides comprising the entire CH1, hinge, CH2 and / or CH3 domains, and fragments of such peptides comprising only, for example, hinge, CH2 and CH3 domains. The Fc domain may be derived from an immunoglobulin of any species and / or any subtype, including but not limited to human IgG1, IgG2, IgG3, IgG4, IgD, IgA, IgE or IgM antibodies. The Fc domain encompasses natural Fc and Fc variant molecules. Like Fc variants and natural Fc, the term Fc domain includes molecules in monomeric or multimeric form, whether digested from a whole antibody or produced by other means.

[0203] As described herein, one of ordinary skill in the art will appreciate that any Fc domain can be modified such that its amino acid sequence changes from the amino acid sequence of a native Fc domain of a naturally occurring immunoglobulin molecule.

[0204] The Fc domain of the interferon receptor Fc construct of the present disclosure can be derived from different immunoglobulin molecules. For example, the Fc domain of the interferon receptor Fc construct can include a CH2 and / or CH3 domain derived from an IgG1 molecule and a hinge region derived from an IgG3 molecule. In another example, the Fc domain can include a chimeric hinge region partially derived from an IgG1 molecule and partially derived from an IgG3 molecule. In another example, the Fc domain can include a chimeric hinge partially derived from an IgG1 molecule and partially derived from an IgG4 molecule. The wild-type human IgG1 Fc domain has the amino acid sequence shown in SEQ ID NO: 26.

[0205] As used herein, the term "serum half-life" refers to the time required for the serum soluble interferon receptor concentration in vivo to decrease by 50%. The shorter the serum half-life of the soluble interferon receptor, the shorter the time it exerts its therapeutic effect.

[0206] As used herein, the term "soluble interferon receptor" or "heterodimer" refers to a molecule comprising a first IFNAR domain or a variant or fragment thereof, and a second IFNAR domain or a variant or fragment thereof. In some embodiments, the IFNAR domain is the extracellular domain of IFNAR. In some embodiments, the soluble interferon receptor comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an extracellular domain of IFNAR or a variant or fragment thereof, and the second polypeptide comprises an extracellular domain of IFNAR or a variant or fragment thereof. In some embodiments, the first and second polypeptides interact to form a dimer (e.g., a heterodimer). In some embodiments, the soluble interferon receptor comprises a first fragment, which is included in an IFNAR domain or a variant or fragment thereof operably connected to an immunoglobulin Fc domain or a variant or fragment thereof with or without a joint domain, and wherein the second polypeptide is included in an IFNAR domain or a variant or fragment thereof operably connected to an immunoglobulin Fc domain or a variant or fragment thereof with or without a joint domain; and nucleic acids encoding such polypeptides. In some embodiments, the first polypeptide is included in INFAR1 or its variant or fragment operably connected to the Fc domain or its variant or fragment with or without a joint domain. In some embodiments, the second polypeptide is included in INFAR2 or its variant or fragment operably connected to the Fc domain or its variant or fragment with or without a joint domain. In some embodiments, the first and second polypeptides dimerize to form a heterodimer construct. In some embodiments, the first and / or second polypeptides may or may not include a leader sequence. In some embodiments, a soluble interferon receptor may or may not include a leader sequence. In some embodiments, a soluble interferon receptor comprises two or more interferon receptor Fc constructs. In some embodiments, a soluble interferon receptor binds type I interferons, for example, IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν.

[0207] As used herein, the term "interferon receptor Fc construct" refers to a polypeptide comprising an IFNAR domain (e.g., IFNAR1 or IFNAR2) or a variant or fragment thereof operably connected to an Fc domain or its variant or fragment with or without a joint domain, and a nucleic acid encoding such a polypeptide. In some embodiments, the IFNAR domain is the extracellular domain of IFNAR. The interferon receptor Fc construct may or may not include a leader sequence. The interferon receptor Fc construct may also be referred to as an "interferon receptor Fc protein".

[0208] As used herein, the term "type I interferon" or "type I IFN" refers to a proinflammatory cytokine that binds to a heterodimeric cell surface receptor IFN-α receptor (IFNAR) comprising IFNAR1 and IFNAR2. There are 16 types of type I interferons in the human body, including IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν. Type I IFN is rapidly produced in a variety of different cell types, and is known to have a wide variety of effects. Although all type I IFNs bind to the same receptor and form a very similar ternary complex in structure, the IFN activity of the difference is due to the different life spans and ligand affinity on each receptor chain, which determines the assembly and dynamics of the signaling complex (Piehler, J. et al. (2012) Immunological Reviews 250: 317-334; Li, H. et al. (2017) J Mol Biol 429: 2571-2589).

[0209] As used herein, the term "type I IFN activity" refers to the biological activity produced when type I IFN interacts with a receptor (IFNAR) (including but not limited to those described herein). The typical consequences of type I IFN production in vivo are the activation of antimicrobial cell programs and the occurrence of innate and adaptive immune responses. Type I interferons regulate innate immune cell activation (e.g., maturation of dendritic cells) to promote antigen presentation and natural killer cell function. Type I IFN also promotes the development of high-affinity antigen-specific T and B cell responses and immune memory (Ivashkiv and Donlin (2014) Nat Rev Immunol 14 (1): 36-49).

[0210] It has been shown that type I IFN activates dendritic cells (DC) and promotes their T cell stimulating ability through autocrine signaling (Montoya et al., (2002) Blood 99: 3263-3271). Type I IFN exposure promotes the maturation of DC by increasing the expression of chemokine receptors and adhesion molecules (e.g., promoting DC migration to draining lymph nodes), co-stimulatory molecules, and MHC class I and class II antigen presentation. DCs that mature after type I IFN exposure can effectively induce protective T cell responses (Wijesundara et al., (2014) Front Immunol 29 (412) and references therein). Therefore, in some embodiments, type I IFN activity is an increase or induction of DC activation and / or maturation. In some embodiments, type I IFN activity is an increase or induction of DC migration to draining lymph nodes.

[0211] In addition, type I IFN can promote T cell activation, proliferation, differentiation and survival (Crouse et al., (2015) Nat Rev Immunol 15: 231-242). Early studies revealed that the expression of MHC-1 is upregulated in response to type I IFN in a variety of cell types (Lindahl et al., (1976), J Infect Dis 133 (Suppl): A66-A68; Lindahl et al., (1976) Proc Natl Acad Sci USA 17: 1284-1287), which is required for optimal T cell stimulation, differentiation, amplification and cytolytic activity. In addition, type I IFN can play an effective costimulatory role on CD8 T cells, thereby enhancing CD8 T cell proliferation and differentiation (Curtsinger et al., (2005) J Immunol 174: 4465-4469; Kolumam et al., (2005) J Exp Med 202: 637-650). Therefore, in some embodiments, type I IFN activity is an increase or induction of T cell proliferation. In some embodiments, type I IFN activity is an increase or induction of CD8+T cell proliferation. In some embodiments, type I IFN activity is an increase or induction of T cell differentiation. In some embodiments, type I IFN activity is an increase or induction of CD8+T cell differentiation.

[0212] For B cells, type I IFN exposure has been shown to promote B cell activation, antibody production, and isotype switching following viral infection or following experimental immunization (Le Bon et al., (2006) J Immunol 176:4:2074-2078; Swanson et al., (2010) J Exp Med 207:1485-1500). Thus, in some embodiments, type I IFN activity is an increase or induction of antibody production.

[0213] In some embodiments, type I IFN activity is produced when IFN α is combined with IFNAR. In some embodiments, type I IFN activity is produced when IFN β is combined with IFNAR. In some embodiments, type I IFN activity is selected from: (i) increase or induction of T cell proliferation (e.g., CD8+T cell proliferation); (ii) increase or induction of DC maturation; (iii) increase or induction of DC migration to draining lymph nodes; (iv) increase or induction of T cell differentiation (e.g., CD8+T cell differentiation); (v) increase or induction of antibody production; and (vi) (i)-(v) any combination. In some embodiments, type I IFN activity is measured in vitro. In some embodiments, type I IFN activity is measured in vivo.

[0214] In some embodiments, the heterodimers described herein inhibit or reduce type I IFN activity. For example, in some embodiments, the heterodimers described herein: (i) inhibit or reduce T cell proliferation (e.g., CD8+T cell proliferation); (ii) inhibit or reduce DC maturation; (iii) inhibit or reduce DC migration to draining lymph nodes; (iv) inhibit or reduce T cell differentiation (e.g., CD8+T cell differentiation); (v) inhibit or reduce IFN-mediated antibody production; or (vi) (i)-(v) any combination. In some embodiments, the inhibition or reduction of type I IFN activity by the heterodimers described herein is relative to the activity in the absence of heterodimers. Methods for determining the inhibition or reduction of T cell proliferation, DC maturation, DC migration, T cell differentiation, and IFN-mediated antibody production are known to those skilled in the art.

[0215] As used herein, the term "dimer" refers to a macromolecular complex formed by two macromolecules (e.g., polypeptides). A "homodimer" refers to a dimer formed by two identical macromolecules (e.g., polypeptides). A "heterodimer" refers to a dimer formed by two different macromolecules (e.g., polypeptides).

[0216] As used herein, the term "variant" refers to a polypeptide derived from a wild-type interferon receptor or Fc domain, and differs from the wild type in one or more changes at one or more positions, i.e., substitution, insertion and / or deletion. Substitution refers to the replacement of an amino acid occupying a position with a different amino acid. Deletion refers to the removal of an amino acid occupying a position. Insertion refers to the addition of 1 or more, such as 1-3 amino acids, to an amino acid adjacent to an amino acid occupying a position. Variant polypeptides must have less than 100% sequence identity or similarity with wild-type polypeptides. In some embodiments, variant polypeptides have an amino acid sequence identity or similarity of about 75% to less than 100% with the amino acid sequence of a wild-type polypeptide, or about 80% to less than 100%, or about 85% to less than 100%, or about 90% to less than 100% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%), or about 95% to less than 100% of an amino acid sequence, for example, over the entire length of a variant polypeptide.

[0217] In some aspects, interferon receptor Fc constructs use one or more "joint domains", such as polypeptide linkers. As used herein, the term "joint domain" refers to one or more amino acids connecting two or more peptide domains in a linear polypeptide sequence. As used herein, the term "polypeptide linker" refers to a peptide or polypeptide sequence (e.g., a synthetic peptide or polypeptide sequence) connecting two or more polypeptide domains in a linear amino acid sequence of a protein. For example, a polypeptide linker can be used to operably connect interferon receptor to an Fc domain. In some embodiments, such a polypeptide linker provides flexibility for a polypeptide molecule. In some embodiments, a polypeptide linker is used to connect (e.g., genetic fusion) such as IFNAR1 domain and Fc domain and / or IFNAR2 domain and Fc domain. Interferon receptor Fc constructs may include more than one joint domain or peptide linker. Various peptide linkers are known in the art.

[0218] As used herein, the term "gly-ser polypeptide linker" refers to a peptide consisting of glycine and serine residues. An exemplary gly / ser polypeptide linker comprises the amino acid sequence (Gly4Ser) n In some embodiments, n is 1 or greater, such as 2 or greater, 3 or greater, 4 or greater, 5 or greater, 6 or greater, 7 or greater, 8 or greater, 9 or greater or 10 or greater (e.g., (Gly4Ser) 10 ). Another exemplary gly / ser polypeptide linker comprises the amino acid sequence Ser (Gly4Ser) n In some embodiments, n is 1 or greater, such as 2 or greater, 3 or greater, 4 or greater, 5 or greater, 6 or greater, 7 or greater, 8 or greater, 9 or greater, or 10 or greater (e.g., Ser(Gly4Ser) 10 ).

[0219] As used herein, the terms "coupled," "conjugated," "linked," "fused," or "fused" are used interchangeably. These terms refer to joining two or more elements or components or domains together by any means including chemical conjugation or recombinant means. Methods of chemical conjugation (e.g., using heterobifunctional cross-linking agents) are known in the art.

[0220] A polypeptide or amino acid sequence "derived from" a specified polypeptide or protein refers to the source of the polypeptide. Preferably, a polypeptide or amino acid sequence derived from a specific sequence has an amino acid sequence substantially identical to that sequence or a portion thereof, wherein the portion consists of at least 10-20 amino acids, preferably at least 20-30 amino acids, more preferably at least 30-50 amino acids, or can be otherwise determined by one of ordinary skill in the art to have the origin of its sequence. A polypeptide derived from another polypeptide may have one or more mutations relative to the starting polypeptide, e.g., an amino acid residue substituted by another amino acid residue or having one or more amino acid residues inserted or deleted.

[0221] In one embodiment, there is one amino acid difference between the starting polypeptide sequence and the sequence derived therefrom. Identity or similarity relative to the sequence is defined herein as the percentage of amino acid residues (i.e., identical residues) in the candidate sequence that are identical to the starting amino acid residue after aligning the sequences and introducing spaces where necessary to obtain maximum percent sequence identity.

[0222] In one embodiment, the polypeptide of the present disclosure is composed of, consists essentially of, or comprises an amino acid sequence and a functionally active variant thereof listed in a sequence listing or a sequence table disclosed herein. In one embodiment, the polypeptide comprises an amino acid sequence that is at least 80%, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identical to an amino acid sequence listed in a sequence listing or a sequence table disclosed herein. In some embodiments, the polypeptide comprises a contiguous amino acid sequence that is at least 80%, e.g., at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identical to a contiguous amino acid sequence listed in a sequence listing or sequence listing disclosed herein. In some embodiments, the polypeptide includes an amino acid sequence having at least 10, e.g., at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 55, at least 60, at least 65, at least 70, at least 75, at least 80, at least 85, at least 90, at least 95, at least 100, at least 200, at least 300, at least 400, or at least 500 (or any integer within these numbers) consecutive amino acids of an amino acid sequence listed in a sequence listing or sequence listing disclosed herein.

[0223] In some embodiments, the interferon receptor Fc construct of the present disclosure is encoded by a nucleotide sequence. The nucleotide sequence of the present disclosure can be used for many applications, including: cloning, gene therapy, protein expression and purification, mutation introduction, DNA vaccination of the required host, antibody production for passive immunity, PCR, primer and probe production, siRNA design and production (e.g., see Dharmacon siDesign website), etc. In some embodiments, the nucleotide sequence of the present disclosure comprises a nucleotide sequence encoding an amino acid sequence of an interferon receptor Fc construct selected from a sequence table or a sequence table, is composed of, or is essentially composed of. In some embodiments, the nucleotide sequence comprises a nucleotide sequence that is at least 80%, e.g., at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identical to a nucleotide sequence encoding an amino acid sequence of a sequence listing disclosed herein or a sequence listing. In some embodiments, the nucleotide sequence includes a contiguous nucleotide sequence that is at least 80%, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identical to a contiguous nucleotide sequence encoding an amino acid sequence listed in a sequence listing or sequence listing disclosed herein. In some embodiments, the nucleotide sequence includes a nucleotide sequence having at least 10, such as at least 15, such as at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 55, at least 60, at least 65, at least 70, at least 75, at least 80, at least 85, at least 90, at least 95, at least 100, at least 200, at least 300, at least 400 or at least 500 (or any integer within these numbers) consecutive nucleotides of a nucleotide sequence encoding an amino acid sequence listed in a sequence listing or sequence listing disclosed herein.

[0224] Those of ordinary skill in the art will also understand that the interferon receptor Fc constructs can be changed so that they are different from the naturally occurring or original sequences from which their components (e.g., interferon receptor domains, joint domains, and Fc domains) are derived in sequence, while retaining the desired activity of the original sequence. For example, conservative substitutions or altered nucleotides or amino acid substitutions that result in "non-essential" amino acid residues can be performed. The isolated nucleic acid molecules encoding non-natural variants can be produced by introducing one or more nucleotide substitutions, additions, or deletions into the nucleotide sequence of the interferon receptor Fc construct so that one or more amino acid substitutions, additions, or deletions are introduced into the encoded protein. Mutations can be introduced by standard techniques, such as site-directed mutagenesis and PCR-mediated mutagenesis.

[0225] Interferon receptor Fc constructs may include conservative amino acid replacements at one or more amino acid residues (e.g., essential or non-essential amino acid residues). "Conservative amino acid replacement" is an amino acid replacement in which an amino acid residue is replaced by an amino acid residue with a similar side chain. Families of amino acid residues with similar side chains have been defined in the art, including basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), non-polar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), β-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Therefore, the non-essential amino acid residue in the interferon receptor Fc construct is preferably replaced by another amino acid residue from the same side chain family. In another embodiment, the string of amino acids can be replaced with a structurally similar string that differs in the order and / or composition of the side chain family members. Alternatively, in another embodiment, mutations can be introduced randomly along all or part of the coding sequence, for example by saturation mutagenesis, and the resulting mutants can be incorporated into an interferon receptor Fc construct and screened for their ability to bind to the desired target.

[0226] The term "amelioration" refers to any therapeutically beneficial outcome in the treatment of a disease state, such as an autoimmune disease state (eg, SLE, Sjögren's syndrome), including prevention, lessening of its severity or progression, regression, or cure.

[0227] The term "in situ" refers to processes that occur in living cells grown separately from a living organism, such as grown in tissue culture.

[0228] The term "in vivo" refers to processes that occur in a living organism.

[0229] As used herein, the term "mammal" or "subject" or "patient" includes humans and non-humans, and includes, but is not limited to, humans, non-human primates, canines, felines, murines, bovines, equines, and porcines.

[0230] In the case of two or more nucleic acids or polypeptide sequences, the term percent "identity" refers to two or more sequences or subsequences having a specified percentage of identical nucleotides or amino acid residues when compared and aligned for maximum correspondence, as measured using one of the sequence comparison algorithms described below (e.g., BLASTP and BLASTN or other algorithms available to those skilled in the art) or by visual inspection. Depending on the application, percent "identity" may exist over regions of the sequences being compared, such as functional domains, or over the full length of the two sequences to be compared.

[0231] To perform sequence comparison, typically one sequence is used as a reference sequence to which a test sequence is compared. When a sequence comparison algorithm is used, the test sequence and the reference sequence are input into a computer, subsequence coordinates are specified if necessary, and sequence algorithm program parameters are specified. The sequence comparison algorithm then calculates the percent sequence identity of the test sequence relative to the reference sequence based on the specified program parameters.

[0232] For example, optimal alignment of sequences for comparison can be performed by the local homology algorithm of Smith & Waterman, Adv Appl Math 1981; 2: 482, by the homology alignment algorithm of Needleman & Wunsch, J Mol Biol 1970; 48: 443, by the similarity search method of Pearson & Lipman, PNAS 1988; 85: 2444, by computer implementations of these algorithms (GAP, BESTFIT, FASTA and TFASTA, Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.), or by visual inspection (see generally Ausubel et al., below).

[0233] An example of an algorithm suitable for determining percent sequence identity and sequence similarity is the BLAST algorithm, which is described in Altschul et al., J Mol Biol 1990; 215: 403-10. Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information website.

[0234] The term "sufficient amount" refers to an amount sufficient to produce the desired effect.

[0235] The term "therapeutically effective amount" is an amount effective to ameliorate symptoms of a disease. A therapeutically effective amount can be a "prophylactically effective amount" because prevention can be considered treatment.

[0236] The term "about" is understood by the skilled artisan and will vary to some extent depending on the context in which it is used. If there is a use of the term that is unclear to the skilled artisan given the context in which it is used, "about" means ±10% of the particular value.

[0237] It must be noted that, as used in the specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0238] Soluble interferon receptor

[0239] In some embodiments, the soluble interferon receptor of the present disclosure includes a first polypeptide and a second polypeptide, wherein each polypeptide comprises an interferon receptor Fc construct with or without a leader sequence. Interferon receptor Fc constructs include interferon receptor domains (e.g., IFNAR1 or IFNAR2) (with or without a leader sequence) or variants or fragments thereof, which are operably coupled with Fc domains or variants or fragments thereof with or without a joint domain. In some embodiments, the first and second polypeptides dimerize to form a dimer; e.g., a heterodimer.

[0240] In some embodiments, the compositions of the present disclosure include a soluble interferon receptor.

[0241] In some embodiments, the interferon receptor domain is operably coupled to the Fc domain, or variant or fragment thereof, in the absence of a linker domain.

[0242] In some embodiments, the interferon receptor domain is operably coupled to the Fc domain or its variant or fragment via a linker domain. In some embodiments, the linker domain is a linker peptide. In some embodiments, the linker domain is a linker nucleotide.

[0243] In some embodiments, the joint domain includes (gly4ser) 2, 3, 4 or 5 variants that change the length of the joint according to 5 amino acid increments. In another embodiment, the joint domain is about 18 amino acids in length and includes a glycosylation site that is connected by N-, which can be sensitive to protease cleavage in vivo. In some embodiments, the glycosylation site that N- is connected can protect soluble interferon receptors from cutting in the joint domain. In some embodiments, the glycosylation site that N- is connected can help separate the folding of independent functional domains separated by the joint domain.

[0244] In some embodiments, the linker domain is an NLG linker (VDGASSPVNVSSPSVQDI) (SEQ ID NO: 18).

[0245] In some embodiments, the interferon receptor Fc construct includes a leader molecule, for example, a leader peptide. In some embodiments, the leader molecule is a leader peptide located at the N-terminal of the interferon receptor domain. In some embodiments, the interferon receptor Fc construct of the present invention is included in the leader peptide at the N-terminal of the molecule, wherein the leader peptide is subsequently cut from the interferon receptor Fc construct. The method for generating the nucleic acid sequence encoding the leader peptide fused to the recombinant protein is well known in the art. In some embodiments, any interferon receptor Fc construct of the present invention can be expressed or synthesized with or without the leader sequence fused to its N-terminal. The protein sequence of the interferon receptor Fc construct of the present disclosure after the fusion leader peptide cutting can be predicted and / or deduced by those skilled in the art.

[0246] In some embodiments, the leader peptide comprises the amino acid sequence: MDWTWRILFLVAAATGTHA (SEQ ID NO: 13). In some embodiments, the leader sequence is a VK3 leader peptide (VK3LP), which comprises the amino acid sequence: METPAQLLFLLLLWLPDTTG (SEQ ID NO: 14). In some embodiments, the leader peptide is fused to the N-terminus of the interferon receptor Fc construct. Such a leader sequence can increase the synthesis and secretion levels of the interferon receptor Fc construct in mammalian cells. In some embodiments, the leader sequence is cleaved to produce an interferon receptor Fc construct. In some embodiments, the interferon receptor Fc construct of the present invention is expressed without a leader peptide fused to its N-terminus, and the resulting interferon receptor Fc construct has an N-terminal methionine.

[0247] In some embodiments, the soluble interferon receptor of the present disclosure includes the first and second polypeptides, wherein each polypeptide comprises an interferon receptor domain (e.g., IFNAR1, IFNAR2) (with or without a leader sequence) or a variant or fragment thereof, which is operably coupled to the N- or C-terminus of an immunoglobulin Fc domain or its variant or fragment with or without a joint domain. In some embodiments, the interferon receptor domains of the two polypeptides are different, e.g., IFNAR1 and IFNAR2. In some embodiments, the soluble interferon receptor is a dimeric molecule. In some embodiments, the first and second polypeptides dimerize to form a heterodimer.

[0248] In some embodiments, the interferon receptor Fc construct includes substantially all or at least active fragments of the interferon receptor. In some embodiments, the interferon receptor is IFNAR1, such as the extracellular domain of human IFNAR1 (SEQ ID NO: 11), or a variant or fragment thereof. In some embodiments, the interferon receptor is IFNAR2, such as the extracellular domain of human IFNAR2 (SEQID NO: 12), or a variant or fragment thereof. In some embodiments, IFNAR-joint-Fc containing 20 or 25aa joint domains is prepared. In some embodiments, IFNAR-joint-Fc containing 10aa joint domains is prepared. In some embodiments, IFNAR-joint-Fc containing 5aa joint domains is prepared. In some embodiments, IFNAR-Fc without joint domains is prepared. In some embodiments, the interferon receptor may include or not include a leader sequence.

[0249] In some embodiments, the interferon receptor Fc construct includes IFNAR-linker-Fc, wherein the IFNAR domain is located on the COOH side of Fc. In other embodiments, the interferon receptor Fc construct includes IFNAR-linker-Fc, wherein the IFNAR domain is located on the NH2 side of Fc. In some embodiments, soluble interferon receptors include: IFNAR1-Fc and IFNAR2-Fc; IFNAR1-linker-Fc and IFNAR2-linker-Fc; Fc-IFNAR1 and Fc-IFNAR2; Fc-linker-IFNAR1 and Fc-linker-IFNAR2. The interferon receptor Fc construct may or may not include a leader sequence.

[0250] In some embodiments, the fusion junction between the interferon receptor domain and other domains of the interferon receptor Fc construct is optimized.

[0251] Figure 1 Exemplary configurations of soluble interferon receptors are shown, and the sequence listing provides the sequences of exemplary interferon receptor Fc constructs of various configurations.

[0252] In one embodiment, the interferon receptor domain is operably coupled (e.g., chemically conjugated or genetically fused (e.g., directly or via a linker, such as a polypeptide linker)) to the N-terminus of the Fc domain, or a variant or fragment thereof. In another embodiment, the interferon receptor domain is operably coupled (e.g., chemically conjugated or genetically fused (e.g., directly or via a linker, such as a polypeptide linker)) to the C-terminus of the Fc domain, or a variant or fragment thereof. In other embodiments, the interferon receptor domain is operably coupled (e.g., chemically conjugated or genetically fused (e.g., directly or via a linker, such as a polypeptide linker)) through the amino acid side chains of the Fc domain, or a variant or fragment thereof.

[0253] In some embodiments, the soluble interferon receptor comprises at least two identical or different interferon receptor domains (e.g., IFNAR1 and IFNAR2) or variants or fragments thereof, and at least two identical or different Fc domains or variants or fragments thereof, with an optional linker domain between the interferon receptor domains and the Fc domains.

[0254] In some embodiments, the interferon receptor Fc construct forms heterodimers.

[0255] In some embodiments, as described herein, the soluble interferon receptor of the present disclosure comprises an Fc domain or a variant or fragment thereof, thereby increasing the serum half-life and bioavailability of the soluble interferon receptor.

[0256] Those skilled in the art will appreciate that other configurations of the interferon receptor domain and the Fc domain are possible, including an optional linker between the interferon receptor domain and the Fc domain. It should also be understood that the domain orientation can be changed, as long as the interferon receptor domain is active in the specific configuration tested.

[0257] In certain embodiments, the soluble interferon receptor of the present disclosure has at least one interferon receptor domain specific to a target molecule mediating a biological effect. In another embodiment, the combination of the soluble interferon receptor of the present disclosure and a target molecule such as type I interferon (e.g., IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν) causes the target molecule to be reduced or eliminated, for example, from a cell, tissue, or circulation.

[0258] In other embodiments, the interferon receptor Fc constructs of the present disclosure can be assembled together or assembled with other interferon receptor Fc constructs or polypeptides to form a binding protein ("multimer") having two or more polypeptides, wherein at least one polypeptide of the multimer is an interferon receptor Fc construct of the present disclosure. Exemplary multimeric forms include altered binding proteins of dimeric, trimeric, tetrameric and hexameric, etc. In one embodiment, the polypeptides of the multimer are the same (i.e., homomeric binding proteins, e.g., homodimers, homotetramers). In another embodiment, the polypeptides of the multimer are different (e.g., heteromeric binding proteins, e.g., heterodimers).

[0259] In some embodiments, the serum half-life of the soluble interferon receptor is increased by at least about 1.5 times, e.g., at least 3 times, at least 5 times, at least 10 times, at least about 20 times, at least about 50 times, at least about 100 times, at least about 200 times, at least about 300 times, at least about 400 times, at least about 500 times, at least about 600 times, at least about 700 times, at least about 800 times, at least about 900 times, at least about 1000 times, or 1000 times or more relative to a corresponding interferon receptor molecule not fused to an Fc domain, or a variant or fragment thereof. In other embodiments, the serum half-life of the soluble interferon receptor is reduced by at least about 1.5 times, such as at least 3 times, at least 5 times, at least 10 times, at least about 20 times, at least about 50 times, at least about 100 times, at least about 200 times, at least about 300 times, at least about 400 times, at least about 500 times, or 500 times or less relative to the corresponding interferon receptor molecule not fused to the Fc domain or its variant or fragment. The serum half-life of the soluble interferon receptor of the present disclosure can be determined using conventional methods recognized in the art.

[0260] In some embodiments, the activity of the interferon receptor (e.g., IFNAR1 or IFNAR2) in the soluble interferon receptor is not less than about 10 times lower than the activity of the naturally occurring wild-type interferon receptor molecule, for example, 9 times lower, 8 times lower, 7 times lower, 6 times lower, 5 times lower, 4 times lower, 3 times lower or 2 times lower. In some embodiments, the activity of the interferon receptor in the soluble interferon receptor is approximately equal to the activity of the naturally occurring wild-type interferon receptor molecule.

[0261] In some embodiments, soluble interferon receptors can bind interferon and reduce the effects of interferon.

[0262] In some embodiments, the activity of the soluble interferon receptor is detectable in vitro and / or in vivo. In some embodiments, the soluble interferon receptor construct binds to a cell, a diseased cell, a malignant cell, or a cancer cell and interferes with its biological activity.

[0263] In another aspect, a multifunctional soluble interferon receptor is provided, which is linked to an enzyme or antibody with binding specificity, such as a scFv targeting type I interferon. For example, type I interferons include IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν.

[0264] In some embodiments, the target of the IFNAR domain of a soluble interferon receptor is mainly extracellular type I interferon. For example, IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν. In some embodiments, the soluble interferon receptor is active in the acidic environment of endocytic vesicles. In some embodiments, the soluble interferon receptor including the Fc domain or its variant or fragment is adapted to be active in the extracellular and endocytic environment. In some embodiments, the IFNAR domain of a soluble interferon receptor is not active in the cytoplasm of a cell.

[0265] In some embodiments, soluble interferon receptors include both IFNAR1 and IFNAR2. In some embodiments, these soluble interferon receptors improve the treatment of SLE because they bind interferons (IFNs), e.g., IFNα or IFNβ, and eliminate or reduce the inflammatory effects of cytokines.

[0266] Interferon receptor domain

[0267] In some embodiments, the interferon receptor Fc construct of the present disclosure is included in one or more interferon receptor domains (IFNAR) or its variant or fragment operably coupled with immunoglobulin Fc domain or its variant or fragment with or without a joint domain. In some embodiments, the interferon receptor domain is a variant or fragment of the interferon receptor domain. In some embodiments, the interferon receptor domain includes the extracellular domain of interferon receptor. For example, the extracellular domain of IFNAR1 or the extracellular domain of IFNAR2. In some embodiments, the interferon receptor domain includes a leader sequence. In some embodiments, the interferon receptor domain does not include a leader sequence.

[0268] Suitable IFNAR domains are well known in the art and include, but are not limited to, IFNAR1 and IFNAR2. For example, IFNAR domains are discussed in deWeerd et al., Type I Interferon Receptors: Biochemistry and Biological Functions, J. of Biol. Chem., Vol. 282, No. 28, 20053-20057 (2007), the entire contents of which are hereby incorporated by reference.

[0269] Type I interferon-α / β receptor (IFNAR) is a heteromeric cell surface receptor composed of multiple components (called IFNAR1 and IFNAR2). The INFAR complex is unique in cytokine receptors because it binds to and / or mediates signal transduction through more than 15 different but related type I interferon (IFN) ligands, including, for example, IFN-α and IFN-β, several IFNα subtypes, IFN-ω, IFN-ε, IFN-κ and others. After type I IFN binds, IFNAR activates the JAK-STAT signal transduction pathway.

[0270] In some embodiments, the interferon receptor Fc construct of the present disclosure binds to interferon (e.g., IFN-α, INF-β, IFN-ε, -κ, -τ, -ζ, IFN-ω, IFN-ν) when complexed with another interferon receptor Fc construct in heterodimeric form. For example, a heterodimer of the present disclosure capable of binding to interferon comprises a first interferon receptor Fc construct (wherein IFNAR is IFNAR1) and a second interferon receptor Fc construct (wherein IFNAR is IFNAR2).

[0271] In some embodiments, the IFNAR domain of the interferon receptor Fc construct is a naturally occurring wild-type IFNAR1 protein, whose amino acid sequence is shown in SEQ ID NO: 5. In some embodiments, the IFNAR domain is a variant or fragment of the IFNAR1 protein. In some embodiments, the IFNAR domain comprises 50-100, 100-150, 150-200, 200-250, 250-300, 300-350, 350-400, 400-450, 450-500 or 450-557 amino acids of the amino acid sequence shown in SEQ ID NO: 5. In some embodiments, the IFNAR domain comprises amino acids 350-360, 360-370, 370-380, 380-390, 390-400, 400-410, 410-420, 420-430, 430-440, 440-450, 450-470, 460-470, 470-480, 480-490, or 490-500 of the amino acid sequence of SEQ ID NO: 5. In some embodiments, the IFNAR1 domain comprises amino acids 28-436 of the amino acid sequence of SEQ ID NO: 5.

[0272] In some embodiments, the IFNAR domain comprises an amino acid sequence that is at least 70% identical to the amino acid sequence set forth in SEQ ID NO:5, e.g., 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or at least 99.5% identical.

[0273] In some embodiments, the IFNAR domain of the interferon receptor Fc construct comprises the amino acid sequence set forth in SEQ ID NO: 7. In some embodiments, the IFNAR domain comprises the amino acid sequence set forth in SEQ ID NO: 11. In some embodiments, the IFNAR domain comprises an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO: 7 or SEQ ID NO: 11, such as 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence set forth in SEQ ID NO: 7 or SEQ ID NO: 11.

[0274] In some embodiments, the IFNAR domain is a naturally occurring human wild-type IFNAR2 protein, whose amino acid sequence is shown in SEQ ID NO: 6. In some embodiments, the IFNAR domain is a variant or fragment of the IFNAR2 protein. In some embodiments, the IFNAR domain comprises 50-100, 100-150, 150-200, 200-250, 250-300, 300-350, 350-400, 400-450, 450-500 or 450-515 amino acids of the amino acid sequence shown in SEQ ID NO: 6. In some embodiments, the IFNAR domain comprises 150-160, 160-170, 170-180, 180-190, 190-200, 200-210, 210-220, 220-230, 230-240, 240-250, 250-260, 260-270, 270-280, 280-290, 290-300 amino acids of the amino acid sequence shown in SEQ ID NO: 6. In some embodiments, the IFNAR2 domain comprises amino acids 27-243 of the amino acid sequence shown in SEQ ID NO: 6.

[0275] In some embodiments, the IFNAR domain comprises an amino acid sequence that is at least 70% identical to the amino acid sequence set forth in SEQ ID NO:6, e.g., 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or at least 99.5% identical.

[0276] In some embodiments, the IFNAR domain of the interferon receptor Fc construct comprises the amino acid sequence set forth in SEQ ID NO: 8. In some embodiments, the IFNAR domain comprises the amino acid sequence set forth in SEQ ID NO: 12. In some embodiments, the IFNAR domain comprises an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO: 8 or SEQ ID NO: 12, such as 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence set forth in SEQ ID NO: 8 or SEQ ID NO: 12.

[0277] In some embodiments, the interferon receptor domain includes the extracellular domain of interferon receptor. In some embodiments, the interferon receptor domain includes the extracellular domain of IFNAR1. In some embodiments, the interferon receptor domain includes the extracellular domain of IFNAR2. In some embodiments, the extracellular domain of interferon receptor (e.g., IFNAR1 or IFNAR2) is substantially free of transmembrane domain and cytoplasmic domain. In some embodiments, the extracellular domain of interferon receptor (e.g., IFNAR1 or IFNAR2) is by no means transmembrane domain and cytoplasmic domain.

[0278] In some embodiments, the extracellular domain of IFNAR1 comprises the amino acid sequence set forth in SEQ ID NO: 7 or SEQ ID NO: 11. In some embodiments, the extracellular domain of IFNAR1 comprises amino acids 28-436 of the amino acid sequence set forth in SEQ ID NO: 5. In some embodiments, the extracellular domain of IFNAR1 comprises amino acids 1-436 of the amino acid sequence set forth in SEQ ID NO: 5.

[0279] In some embodiments, the extracellular domain of IFNAR2 comprises the amino acid sequence set forth in SEQ ID NO: 8 or SEQ ID NO: 12. In some embodiments, the extracellular domain of IFNAR2 comprises amino acids 27-243 of the amino acid sequence set forth in SEQ ID NO: 6. In some embodiments, the extracellular domain of IFNAR2 comprises amino acids 1-243 of the amino acid sequence set forth in SEQ ID NO: 6.

[0280] In some embodiments, the IFNAR domain is changed or modified, for example, by causing a mutation that amino acids are added, deleted or replaced. As used herein, the term "IFNAR domain variant" refers to an IFNAR domain having at least one amino acid modification (such as amino acid replacement) compared to the wild-type IFNAR or its fragment derived from the IFNAR domain. For example, in the case where the IFNAR domain is derived from human wild-type IFNAR, compared to the wild-type amino acid at the corresponding position of human wild-type IFNAR, the variant includes at least one amino acid mutation (for example, replacement). For example, in the case where the IFNAR domain is derived from the extracellular domain of human IFNAR, compared to the amino acid at the corresponding position of extracellular IFNAR, the variant includes at least one amino acid mutation (for example, replacement).

[0281] In some embodiments, the IFNAR variant comprises one or more amino acid substitutions at an amino acid position located in the extracellular domain of IFNAR.

[0282] In some embodiments of the present disclosure, the soluble interferon receptor includes one or more IFNAR domains. In some embodiments, the soluble interferon receptor includes two IFNAR domains. In some embodiments, the IFNAR domains of the soluble interferon receptor are different (e.g., IFNAR1 and IFNAR2).

[0283] In some embodiments, the soluble interferon receptor includes a first polypeptide and a second polypeptide. In some embodiments, both the first polypeptide and the second polypeptide include an interferon receptor Fc construct. In some embodiments, the IFNAR domain of the interferon receptor Fc construct is IFNAR1 or a variant or fragment thereof. In some embodiments, the IFNAR domain of the interferon receptor Fc construct is IFNAR2 or a variant or fragment thereof. In some embodiments, the IFNAR domain of the first polypeptide of the soluble interferon receptor includes IFNAR1 or a variant or fragment thereof. In some embodiments, the IFNAR domain of the second polypeptide of the soluble interferon receptor includes IFNAR2 or a variant or fragment thereof. In some embodiments, the IFNAR domain of the interferon receptor Fc construct is human IFNAR1 or human IFNAR2. In some embodiments, the IFNAR domain of the interferon receptor Fc construct is a fragment or variant of IFNAR1 or a fragment or variant of IFNAR2. In some embodiments, the IFNAR domain of the interferon receptor Fc construct is the extracellular domain of human IFNAR1 or the extracellular domain of human IFNAR2.

[0284] Linker domain

[0285] In some embodiments, the interferon receptor Fc construct includes a joint domain. In some embodiments, the interferon receptor Fc construct includes multiple joint domains. In some embodiments, the joint domain is a polypeptide joint. In some aspects, it is desirable to use a polypeptide joint to fuse Fc or its variants or fragments with one or more interferon receptor domains to form an interferon receptor Fc construct. In some embodiments, the interferon receptor Fc construct does not include a joint domain.

[0286] In one embodiment, the polypeptide linker is synthetic. As used herein, the term "synthetic" with respect to a polypeptide linker includes a peptide (or polypeptide) comprising an amino acid sequence (which may or may not be naturally occurring) connected to a sequence (which may or may not be naturally occurring) (e.g., an Fc sequence) that is not naturally connected to it in nature with a linear amino acid sequence. For example, a polypeptide linker may include a non-naturally occurring polypeptide, which is a modified form of a naturally occurring polypeptide (e.g., including mutations such as additions, substitutions, or deletions), or it includes a first amino acid sequence (which may or may not be naturally occurring). The polypeptide linker of the present invention can be used, for example, to ensure that Fc or its variants or fragments are juxtaposed to ensure the correct folding and formation of functional Fc or its variants or fragments. Preferably, a polypeptide linker compatible with the present invention is relatively non-immunogenic and does not inhibit any non-covalent binding between the monomer subunits of the binding protein.

[0287] In certain embodiments, the interferon receptor Fc construct employs an NLG linker as shown in SEQ ID NO:18.

[0288] In certain embodiments, the interferon receptor Fc construct of the present disclosure adopts a polypeptide linker to join any two or more domains in a single polypeptide chain frame. In one embodiment, two or more domains can be independently selected from any Fc domain or its variant or fragment or interferon receptor domain discussed herein. For example, in certain embodiments, a polypeptide linker can be used to fuse the Fc domain or its variant or fragment with IFNAR. In some embodiments, the polypeptide linker of the present invention can be used to genetically fuse the C-terminus of IFNAR with the N-terminus of the Fc domain or its variant or fragment to form an interferon receptor Fc construct. In other embodiments, the polypeptide linker of the present invention can be used to genetically fuse the C-terminus of the Fc domain or its variant or fragment with the N-terminus of IFNAR to form an interferon receptor Fc construct. In other embodiments, the polypeptide linker of the present invention can be used to genetically fuse the C-terminus of IFNAR with the C-terminus of the Fc domain or its variant or fragment to form an interferon receptor Fc construct. In other embodiments, the polypeptide linker of the present invention can be used to genetically fuse the N-terminus of IFNAR with the N-terminus of the Fc domain or its variant or fragment to form an interferon receptor Fc construct.

[0289] In one embodiment, the polypeptide linker comprises a portion of an Fc domain or a variant or fragment thereof. For example, in one embodiment, the polypeptide linker may comprise an Fc fragment (e.g., a C or N domain), or different portions of an Fc domain or a variant thereof.

[0290] In another embodiment, the polypeptide linker comprises or consists of a gly-ser linker. As used herein, the term "gly-ser linker" refers to a peptide consisting of glycine and serine residues. An exemplary gly / ser linker comprises an amino acid sequence of the formula (Gly4Ser)n, wherein n is a positive integer (e.g., 1-10, 1, 2, 3, 4, or 5). A preferred gly / ser linker is (Gly4Ser)1. Another preferred gly / ser linker is (Gly4Ser)2. Another preferred gly / ser linker is (Gly4Ser)3. Another preferred gly / ser linker is (Gly4Ser)4. Another preferred gly / ser linker is (Gly4Ser)5. In certain embodiments, a gly-ser linker can be inserted between two other polypeptide linker sequences (e.g., any polypeptide linker sequences described herein). In other embodiments, a gly-ser linker is connected to one or both ends of another sequence of a polypeptide linker (e.g., any polypeptide linker sequences described herein). In still other embodiments, two or more gly-ser linkers are incorporated in tandem into the polypeptide linker.

[0291] In other embodiments, the polypeptide linker of the present invention comprises a biologically relevant peptide sequence or a sequence portion thereof. For example, a biologically relevant peptide sequence may include but is not limited to a sequence derived from an anti-rejection or anti-inflammatory peptide. The anti-rejection or anti-inflammatory peptide may be selected from a cytokine inhibitory peptide, a cell adhesion inhibitory peptide, a thrombin inhibitory peptide, and a platelet inhibitory peptide. In a preferred embodiment, the polypeptide linker comprises a peptide sequence selected from IL-1 inhibition or antagonist peptide sequence, erythropoietin (EPO) mimetic peptide sequence, thrombopoietin (TPO) mimetic peptide sequence, G-CSF mimetic peptide sequence, TNF antagonist peptide sequence, integrin binding peptide sequence, selectin antagonist peptide sequence, antipathogenic peptide sequence, vasoactive intestinal peptide (VIP) mimetic peptide sequence, calmodulin antagonist peptide sequence, mast cell antagonist, SH3 antagonist peptide sequence, urokinase receptor (UKR) antagonist peptide sequence, somatostatin or cortistatin mimetic peptide sequence, and macrophage and / or T cell inhibitory peptide sequence. Exemplary peptide sequences, any of which can be used as a polypeptide linker, are disclosed in US Pat. No. 6,660,843, which is incorporated herein by reference.

[0292] Other linkers suitable for use in interferon receptor Fc constructs are known in the art, for example, the serine-rich linkers disclosed in US 5,525,491, the helix-forming peptide linkers disclosed in Arai et al., Protein Eng 2001; 14: 529-32 (e.g., A(EAAAK)nA(n=2-5)), and the stabilizing linkers disclosed in Chen et al., Mol Pharm 2011; 8: 457-65, namely the dipeptide linker LE, the thrombin-sensitive disulfide cyclic peptide linker, and the α-helix-forming linker LEA(EAAAK)4ALEA(EAAAK)4ALE (SEQ ID NO: 19).

[0293] Other exemplary linkers include GS linkers (ie, (GS)n), GGSG (SEQ ID NO: 27) linkers (ie, (GGSG)n), GSAT linkers (SEQ ID NO: 28), SEG linkers, and GGS linkers (ie, (GGSGGS)n), where n is a positive integer (e.g., 1, 2, 3, 4, or 5). Other suitable linkers for use in interferon receptor Fc constructs can be found using public databases such as the Linker Database (ibi.vu.nl / programs / linkerdbwww). The Linker Database is a database of domain-interconnected linkers in multifunctional enzymes that serve as potential linkers in novel fusion proteins (see, e.g., George et al., Protein Engineering 2002; 15: 871-9).

[0294] It should be understood that the variant forms of these exemplary polypeptide linkers can be introduced into the nucleotide sequence of the encoding polypeptide linker by one or more nucleotide substitutions, additions or deletions so that one or more amino acid replacements, additions or deletions are introduced into the polypeptide linker to produce. Sudden changes can be introduced by standard techniques such as site-directed mutagenesis and PCR-mediated mutagenesis.

[0295] The length of polypeptide linker of the present disclosure is at least one amino acid, and can have different lengths. In one embodiment, polypeptide linker of the present disclosure is about 1 to about 50 amino acid length. As used in this case, the term "about" represents + / - two amino acid residues. Since the linker length must be a positive integer, the length of about 1 to 50 amino acid means that the length is 1 to 48-52 amino acid. In another embodiment, polypeptide linker of the present disclosure is about 5-10 amino acid length. In another embodiment, polypeptide linker of the present disclosure is about 10-20 amino acid length. In another embodiment, polypeptide linker of the present disclosure is about 15-30 amino acid length. In another embodiment, polypeptide linker of the present disclosure is about 15 to about 50 amino acid length.

[0296] In another embodiment, the polypeptide linker of the present disclosure is about 20 to about 45 amino acids in length. In another embodiment, the polypeptide linker of the present disclosure is about 15 to about 25 amino acids in length. In another embodiment, the polypeptide linker of the present disclosure is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60 or 61 or more amino acids in length.

[0297] Polypeptide linkers can be introduced into the polypeptide sequence using techniques known in the art. Modifications can be confirmed by DNA sequence analysis. Plasmid DNA can be used to transform host cells for stable production of the produced polypeptide.

[0298] Fc domain

[0299] In some embodiments, a polypeptide comprising one or more interferon receptor domains or variants or fragments thereof is operably coupled to an Fc domain with or without a linker domain, which serves as a scaffold and a means of increasing the serum half-life of the polypeptide. In some embodiments, one or more interferon receptor domains and / or Fc domains are non-glycosylated, deglycosylated or hypoglycosylated. In some embodiments, the Fc domain is a mutant or variant Fc domain or a fragment of an Fc domain.

[0300] Suitable Fc domains are well known in the art, and include, but are not limited to, Fc and Fc variants, such as those disclosed in WO2011 / 053982, WO 02 / 060955, WO 02 / 096948, WO05 / 047327, WO05 / 018572, and US 2007 / 0111281 (the foregoing are incorporated herein by reference). It is within the capabilities of the skilled person to introduce (e.g., clone, conjugate) an Fc domain into an interferon receptor Fc construct disclosed herein (with or without altered glycosylation) using conventional methods.

[0301] In some embodiments, the Fc domain is a wild-type human IgG1 Fc, as set forth in SEQ ID NO: 26. In some embodiments, the Fc domain is a wild-type human IgG4 Fc, as set forth in SEQ ID NO:112.

[0302] In some embodiments, the Fc domain is changed or modified by, for example, a mutation that causes the addition, deletion or replacement of an amino acid. As used herein, the term "Fc domain variant" refers to an Fc domain having at least one amino acid modification (e.g., amino acid replacement) compared to the wild-type Fc from which the Fc domain is derived. For example, in the case where the Fc domain is derived from a human IgG1 antibody, the variant comprises at least one amino acid mutation (e.g., replacement) compared to the wild-type amino acid at the corresponding position in the human IgG1 Fc region. The amino acid replacement of the Fc variant can be located within the Fc domain at a position referred to as the position number (according to the EU index number) given in the Fc region of the antibody corresponding to the residue.

[0303] In one embodiment, the Fc variant comprises one or more amino acid replacements at an amino acid position located in the hinge region or a portion thereof. In another embodiment, the Fc variant comprises one or more amino acid replacements at an amino acid position located in the CH2 domain or a portion thereof. In another embodiment, the Fc variant comprises one or more amino acid replacements at an amino acid position located in the CH3 domain or a portion thereof. In another embodiment, the Fc variant comprises one or more amino acid replacements at an amino acid position located in the CH4 domain or a portion thereof.

[0304] In some embodiments, the Fc variant comprises one or more of the following amino acid substitutions: T350V, L351Y, F405A, and Y407V. In some embodiments, the Fc variant comprises one or more of the following amino acid substitutions: T350V, T366L, K392L, and T394W. In some embodiments, the Fc region has a mutation at N83 (i.e., N297 by Kabat numbering) to produce an aglycosylated Fc region (e.g., Fc N83S).

[0305] In some embodiments, the Fc domain includes a mutation in one or more of the three hinge region cysteines (residues 220, 226, and 229, numbered according to the EU index). In some embodiments, one or more of the three hinge cysteines of the Fc domain may be mutated to SCC (SEQ ID NO: 20) or SSS (SEQ ID NO: 21), wherein "S" represents an amino acid substitution of cysteine ​​to serine (where CCC refers to the three cysteines present in the wild-type hinge domain). Thus, "SCC" indicates an amino acid substitution of only the first cysteine ​​of the three hinge region cysteines (residues 220, 226, and 229, numbered according to the EU index) to serine, while "SSS" indicates that all three cysteines in the hinge region are replaced by serine (residues 220, 226, and 229, numbered according to the EU index).

[0306] In some aspects, the Fc domain is a mutant human IgG1 Fc domain.

[0307] In some aspects, the mutant Fc domain comprises one or more mutations in the hinge, CH2, and / or CH3 domains.

[0308] In some aspects, the Fc domain is a mutant human IgG4 Fc domain. In some embodiments, the mutation in the IgG4 Fc domain includes one or more mutations selected from the following mutation groups: F296Y, E356K, R409K and H345R. In some embodiments, the mutation in the IgG4 Fc domain includes one or more mutations selected from the following mutation groups: F296Y, R409K and K439E. In some embodiments, the soluble interferon receptor disclosed herein includes a first polypeptide comprising a mutant IgG4 Fc domain, wherein the Fc domain includes mutations F296Y, E356K, R409K and H345R, and a second polypeptide includes a mutant IgG4 Fc domain, wherein the CH3 domain includes mutations F296Y, R409K and K439E. In some embodiments, the mutant IgG4 Fc domain includes one or more mutations in the hinge, CH2 and and / or CH3 domains.

[0309] A. CH2 substitution

[0310] In some aspects, the mutant Fc domain includes a P238S mutation. In some aspects, the mutant Fc domain includes a P331S mutation. In some aspects, the mutant Fc domain includes a P238S mutation and a P331S mutation. In some aspects, the mutant Fc domain includes P238S and / or P331S, and may include one or more mutations in three hinge cysteines (residues 220, 226 and 229), according to the EU index numbering. In some aspects, the mutant Fc domain includes P238S and / or P331S and / or one or more mutations in three hinge cysteines (residues 220, 226 and 229), according to the EU index numbering. In some aspects, the mutant Fc domain includes a mutation in P238S and / or P331S and / or hinge cysteines to SCC or three hinge cysteines to SSS. In some aspects, the mutant Fc domain comprises P238S and P331S and a mutation in at least one of the three hinge cysteines. In some aspects, the mutant Fc domain comprises P238S and P331S and SCC. In some aspects, the mutant Fc domain comprises P238S and P331S and SSS. In some aspects, the mutant Fc domain comprises P238S and SCC or SSS. In some aspects, the mutant Fc domain comprises P331S and SCC or SSS (all numberings are according to EU index).

[0311] In some aspects, the mutant Fc domain includes a mutation at an N-linked glycosylation site such as N297, for example, asparagine is substituted with another amino acid such as serine, for example, N297S. In some aspects, the mutant Fc domain includes a mutation at an N-linked glycosylation site such as N297, for example, asparagine is substituted with another amino acid such as serine, for example, N297S, and a mutation in one or more of the three hinge cysteines. In some aspects, the mutant Fc domain includes a mutation at an N-linked glycosylation site such as N297, for example, asparagine is substituted with another amino acid such as serine, for example, N297S, and a mutation of one of the three hinge cysteines to SCC or all three cysteines to SSS. In some aspects, the mutant Fc domain includes a mutation at an N-linked glycosylation site such as N297, for example, asparagine is substituted with another amino acid such as serine, for example, N297, and one or more mutations in the CH2 domain that reduce FcγR binding and / or complement activation, such as a mutation of P238 or P331 or both, for example, P238S or P331S or both P238S and P331S. In some aspects, such mutant Fc domains may further include a mutation in the hinge region, for example, SCC or SSS (all numberings are according to the EU index). In some aspects, the mutant Fc domain is as shown in the sequence table or sequence table herein.

[0312] B. CH3 substitution

[0313] In some embodiments, heterodimers are formed by mutations in the CH3 domain of the Fc domain on the interferon receptor Fc construct disclosed herein. The heavy chain is first engineered for heterodimerization using a "knob-and-hole" strategy (Rigway B et al., Protein Eng., 9 (1996) pp. 617-621, incorporated herein by reference). The term "knob-and-hole" refers to a technique for pairing two polypeptides together in vitro or in vivo by introducing a protrusion (knob) into one polypeptide and a cavity (hole) into another polypeptide at the interface where the two polypeptides interact. See, for example, WO 96 / 027011, WO 98 / 050431, US 5,731,168, US2007 / 0178552, WO2009089004, US20090182127. In particular, the combination of mutations in the CH3 domain can be used to form heterodimers, for example, S354C, T366W in the "knob" heavy chain and Y349C, T366S, L368A, Y407V in the "hole" heavy chain. In another example, T366Y in the "knob" heavy chain and Y407T in the "hole" heavy chain. In some embodiments, the soluble interferon receptor disclosed herein includes a first CH3 domain with a knob mutation T366W and a second CH3 domain with a hole mutation T366S, L368A and Y407V (numbered according to the EU index). In some embodiments, the soluble interferon receptor disclosed herein includes a first CH3 domain with a knob mutation T366Y and a second CH3 domain with a hole mutation Y407T.

[0314] In some embodiments, the CH3 mutations are those described in US 2012 / 0149876A1, US 2017 / 0158779, US9574010, and US 9562109 (each of which is incorporated herein by reference in its entirety); and Von Kreudenstein, TS et al. mABs, 5 (2013), pp. 646-654 (incorporated herein by reference), and include the following mutations: T350V, L351Y, F405A, and Y407V (first CH3 domain); and T350V, T366L, K392L, T394W (second CH3 domain). In some embodiments, a soluble interferon receptor disclosed herein comprises a first CH3 domain having T350V, L351Y, F405A, and Y407V mutations and a second CH3 domain having T350V, T366L, K392L, T394W mutations (numbered according to the EU index).

[0315] In some embodiments, heterodimers are formed by mutations in the CH3 domain of the Fc domain on the interferon receptor Fc construct disclosed herein. In particular, the combination of mutations in the CH3 domain can be used to form heterodimers with high heterodimer stability and purity; for example, see, for example, Von Kreudenstein et al., mAbs 5:5,646-654, September-October 2013, and US 2012 / 0149876A1, US 2017 / 0158779, US 9574010 and US 9562109, each of which is incorporated herein by reference in its entirety. In some embodiments, the mutation in the Fc domain includes one or more mutations selected from the following mutation groups: T350V, L351Y, F405A and Y407V. In some embodiments, the mutation in the Fc domain includes one or more mutations selected from the following mutation groups: T350V, T366L, K392L and T394W. In some embodiments, the interferon receptor Fc construct disclosed herein includes a CH3 domain with mutations T350V, L351Y, F405A and Y407V. In some embodiments, the interferon receptor Fc construct disclosed herein includes a CH3 domain with mutations T350V, T366L, K392L and T394W. In some embodiments, the soluble interferon receptor disclosed herein includes a first polypeptide comprising a mutant Fc domain (wherein the CH3 domain comprises mutations T350V, L351Y, F405A and Y407V); and a second polypeptide comprising a mutant Fc domain (wherein the CH3 domain comprises mutations T350V, T366L, K392L and T394W).

[0316] Other mutations in the CH3 domain of the Fc domain are expected to preferentially form heterodimers. For example, see, for example, Von Kreudenstein et al., mAbs 5:5, 646-654; September-October 2013, incorporated herein by reference. In some embodiments, the mutations in the Fc domain of the first polypeptide include one or more mutations selected from the following mutation groups: T350V, L351Y, F405A, and Y407V, and the mutations in the Fc domain of the second polypeptide include one or more mutations selected from the following mutation groups: T350V, T366L, K392M, and T394W. In some embodiments, the mutations in the Fc domain of the first polypeptide include one or more mutations selected from the following mutation groups: L351Y, F405A, and Y407V, and the mutations in the Fc domain of the second polypeptide include one or more mutations selected from the following mutation groups: T366L, K392M, and T394W.

[0317] In some embodiments, CH3 mutations are those described in Moore, GL et al. (mABs, 3 (2011), pp. 546-557), and include the following mutations: S364H and F405A (first CH3 domain); and Y349T and T394F (second CH3 domain). In some embodiments, the interferon receptor Fc construct disclosed herein includes a first CH3 domain with S364H and F405A mutations and a second CH3 domain with Y349T and T394F mutations (numbered according to the EU index).

[0318] In some embodiments, the CH3 mutations are those described by Gunasekaran, K. et al. (J. Biol. Chem., 285 (2010), pp. 19637-19646), and include the following mutations: K409D and K392D (first CH3 domain); and D399K and E365K (second CH3 domain). In some embodiments, the interferon receptor Fc construct disclosed herein includes a first CH3 domain having K409D and K392D mutations and a second CH3 domain having D399K and E365K mutations (numbered according to the EU index).

[0319] Interferon receptor Fc constructs disclosed herein can use art-recognized Fc variants that are known to confer effector function and / or alterations in FcR binding. For example, International PCT Publications WO88 / 07089A1, WO96 / 14339A1, WO98 / 05787A1, WO98 / 23289A1, WO99 / 51642A1, WO99 / 58572A1, WO00 / 09560A2, WO00 / 32767A1, WO00 / 42072A2, WO02 / 44215A2, WO02 / 060919A2, WO03 / 074569A2, WO04 / 016750A2 , WO04 / 029207A2, WO04 / 035752A2, WO04 / 063351A2, WO04 / 074455A2, WO04 / 099249A2, WO05 / 040217A2, WO04 / 044859, WO05 / 070963A1, WO05 / 077981A2, WO05 / 092925A2, WO05 / 123780A2, WO06 / 019447A1, WO06 / 047350A2 and WO06 / 085967A2; US Patent Publication Nos. US2007 / 0231329, US2007 / 0231329, US2007 / 0237765, US2007 / 0237766, US2007 / 0237767, US2007 / 0243188, US20070248603, US20070286859, US20080057056; or US Patent Nos. 5,648,260; 5,739,27 7; 5,834,250; 5,869,046; 6,096,871; 6,121,022; 6,194,551; 6,242,195; 6,277,375; 6,528,624; 6,538,124; 6,737,056; 6,821,505; 6,998,253; 7,083,784; and 7,317,091 (each of which is incorporated herein by reference). In one embodiment, a specific change can be made at one or more of the disclosed amino acid positions (e.g., a specific substitution of one or more amino acid positions disclosed in the art). In another embodiment, a different change can be made at one or more of the disclosed amino acid positions (e.g., a different substitution of one or more amino acid positions disclosed in the art).

[0320] Other amino acid mutations in the Fc domain are expected to reduce binding to Fcγ receptors and Fcγ receptor subtypes. The assignment of amino acid residue numbers to the Fc domain is according to the definition of Kabat. See, for example, Sequences of Proteins of Immunological Interest (Table of Contents, Introduction and Constant Region Sequences sections), 5th edition, Bethesda, MD: NIH vol. 1: 647-723 (1991); Kabat et al., "Introduction" Sequences of Proteins of Immunological Interest, US Dept of Health and Human Services, NIH, 5th edition, Bethesda, MD vol. 1: xiii-xcvi (1991); Chothia & Lesk, J. Mol. Biol. 196: 901-917 (1987); Chothia et al., Nature 342: 878-883 (1989), each of which is incorporated herein by reference for all purposes.

[0321] For example, positions 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 279, 279, 280, 283, 285, 298, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 312, 315, 322, 324, 327, 329 Mutations at , 330, 331, 333, 334, 335, 337, 338, 340, 356, 360, 373, 376, 378, 379, 382, ​​388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438 or 439 can alter binding as described in U.S. Pat. No. 6,737,056, issued May 18, 2004, the entire contents of which are incorporated herein by reference. The patent reports that changing Pro331 in IgG3 to Ser resulted in a six-fold lower affinity compared to unmutated IgG3, indicating that Pro331 is involved in FcγRI binding. Additionally, U.S. Patent No. 5,624,821, issued April 29, 1997 and incorporated herein by reference in its entirety, discloses that amino acid modifications at positions 234, 235, 236 and 237, 297, 318, 320 and 322 potentially alter receptor binding affinity (numbering according to the EU index).

[0322] Other mutations contemplated for use include, for example, those described in U.S. Patent Application Publication No. 2006 / 0235208, published on October 19, 2006, and incorporated herein by reference in its entirety. This publication describes Fc variants that exhibit reduced binding to Fcγ receptors, reduced antibody-dependent cell-mediated cytotoxicity, or reduced complement-dependent cytotoxicity, comprising at least one amino acid modification in the Fc region, including 232G, 234G, 234H, 235D, 235G, 235H, 236I, 236N, 236P, 236R, 237K, 237L, 237N, 237P, 238K, 239R, 265G, 267R, 269R, 270H, 297S, 299A, 299I, 299V, 325A, 325L, 327 R, 328R, 329K, 330I, 330L, 330N, 330P, 330R and 331L (numbering is according to the EU index), and the double mutants 236R / 237K, 236R / 325L, 236R / 328R, 237K / 325L, 237K / 328R, 325L / 328R, 235G / 236R, 267R / 269R, 234G / 235G, 236R / 237K / 325L, 236R / 325L / 328R, 235G / 236R / 237K and 237K / 325L / 328R. Other mutations contemplated for use as described in the disclosure include 227G, 234D, 234E, 234G, 234I, 234Y, 235D, 235I, 235S, 236S, 239D, 246H, 255Y, 258H, 260H, 264I, 267D, 267E, 268D, 268E, 272H, 272I, 272R, 281D, 282G, 283H, 284E, 293R, 295E, 304T, 324G, 324I, 327D, 327A, 328A, 328D, 32 8E, 328F, 328I, 328M, 328N, 328Q, 328T, 328V, 328Y, 330I, 330L, 330Y, 332D, 332E,335D, insert G between positions 235 and 236, insert A between positions 235 and 236, insert S between positions 235 and 236, insert T between positions 235 and 236, insert N between positions 235 and 236, insert D between positions 235 and 236, insert V between positions 235 and 236, insert L between positions 235 and 236, insert G between positions 235 and 236, insert A between positions 235 and 236, insert S between positions 235 and 236, insert T between positions 235 and 236, insert and E between positions 326 and 327 (numbering is according to the EU index). In addition, mutations described in U.S. Patent Application Publication No. 2006 / 0235208 include 227G / 332E, 234D / 332E, 234E / 332E, 234Y / 332E, 234I / 332E, 234G / 332E, 235I / 332E, 235S / 332E, 235D / 332E, 235E / 332E, 236S / 332E, 236Y / 332E, 236I / 332E, 236G ...G / 332E, 236 6A / 332E, 236S / 332D, 236A / 332D, 239D / 268E, 246H / 332E, 255Y / 332E, 258H / 332E, 260H / 332E, 264I / 332E, 267E / 332E, 267D / 332E, 268D / 332D, 268E / 332D, 268E / 332E, 268D / 332 E, 268E / 330Y, 268D / 330Y, 272R / 332E, 272H / 332E, 283H / 332E, 284E / 332E, 293R / 332E, 2 95E / 332E, 304T / 332E, 324I / 332E, 324G / 332E, 324I / 332D, 324G / 332D, 327D / 332E, 328A / 332E, 328T / 332E, 328V / 332E, 328I / 332E, 328F / 332E, 328Y / 332E, 328M / 332E, 328D / 33 2E, 328E / 332E, 328N / 332E, 328Q / 332E, 328A / 332D, 328T / 332D, 328V / 332D, 328I / 332D,328F / 332D, 328Y / 332D, 328M / 332D, 328D / 332D, 328E / 332D, 328N / 332D, 328Q / 332D, 330L / 332E, 330Y / 332E, 330I / 332E, 3 32D / 330Y, 335D / 332E, 239D / 332E, 239D / 332E / 330Y, 239D / 332E / 330L, 239D / 332E / 330I, 239D / 332E / 268E, 239D / 332E / 26 8D, 239D / 332E / 327D, 239D / 332E / 284E, 239D / 268E / 330Y, 239D / 332E / 268E / 330Y, 239D / 332E / 327A, 239D / 332E / 268E / 327A, 239D / 332E / 330Y / 327A, 332E / 330Y / 268E / 327A, 239D / 332E / 268E / 330Y / 327A, insert G>297-298 / 332E, insert A>297-298 / 332E, insert Insert S>297-298 / 332E, insert D>297-298 / 332E, insert G>326-327 / 332E, insert A>326-327 / 332E, insert T>326-327 / 332E, insert D>326-327 / 332E, insert E>326-327 / 332E, insert G>235-236 / 332E, insert A>235-236 / 332E, insert S>235-236 / 332E, insert T>235-236 / 332E, insert N>235-236 / 332E, Insertion D>235-236 / 332E, insertion V>235-236 / 332E, insertion L>235-236 / 332E, insertion G>235-236 / 332D, insertion A>235-236 / 332D, insertion S>235-236 / 332D, insertion T>235-236 / 332D, insertion N>235-236 / 332D, insertion D>235-236 / 332D, insertion V>235-236 / 332D and insertion L>235-236 / 332D (numbered according to the EU index) are contemplated for use. Mutant L234A / L235A is described, for example, in U.S. Patent Application Publication No. 2003 / 0108548, published on June 12, 2003, the entire contents of which are incorporated herein by reference. In an embodiment, the modifications described include, individually or in combination (numbered according to the EU index).

[0323] C. Alternative stents

[0324] The modular structure of immunoglobulins can be used to generate alternative scaffolds. For example, alternative scaffolds can be alternative Fc formats. In some embodiments, alternative scaffolds can be used to generate heterodimer constructs.

[0325] In some embodiments of the present disclosure, the soluble interferon receptor includes an alternative Fc domain. In some embodiments, the alternative Fc domain is any suitable alternative Fc form known in the art. For example, the Fc domain may be included in any alternative Fc form discussed in Spiess et al. Molecular Immunology, 2015, Oct; 67 (2Pt A) 95-106, the entire contents of which are incorporated herein by reference. For example, the alternative Fc form can be selected from any one of the following forms: (i) bispecific IgG (BsIgG), (ii) attached IgG, (iii) bispecific fragments, (iv) bispecific fusion proteins, (v) bispecific conjugates, (vi) and IgG4 Fab arm exchange and (vii) alternative scaffolds.

[0326] In some embodiments, heterodimers of the present disclosure can be formed by utilizing alternative Fc forms. In some embodiments, the Fc domain can be engineered to promote heterodimerization. In some embodiments, heterodimers are formed by mutations in the CH3 domain of the Fc domain of a soluble interferon receptor disclosed herein.

[0327] (i) Bispecific IgG (BsIgG)

[0328] Bispecific IgG is an alternative Fc format that can be used to promote heterodimerization of two Fc domains and overcome homodimerization. In some embodiments, the CH3 domain of the Fc can be mutated to promote heterodimerization.

[0329] In some embodiments, "knob-hole" can be used to promote heterodimerization of Fc domains. The combination of mutations in the CH3 domain can be used to form heterodimers. In some embodiments, the "knob" is produced by replacing the small amino acid side chains at the interface between the CH3 domains with larger amino acid side chains, and the "hole" is produced by replacing the larger amino acid side chains at the interface between the CH3 domains with smaller amino acid side chains. In some embodiments, the soluble interferon receptor of the present disclosure includes a first interferon receptor Fc construct comprising a first CH3 domain with a knob mutation T366W and a second interferon receptor Fc construct comprising a second CH3 domain with a hole mutation T366S, L368A and Y407V (Ridgeway et al., (1996) Prot. Eng. 9, 617-621 and Atwell et al., (1997) J. Mol. Biol. 270, 26-35, both of which are incorporated herein by reference). In some embodiments, the soluble interferon receptor of the present disclosure includes a first interferon receptor Fc construct comprising a first CH3 domain having a knob mutation T366Y and a second interferon receptor Fc construct comprising a second CH3 domain having a hole mutation Y407V (Ridgeway et al., (1996) Prot. Eng. 9, 617-621). The "hole" mutation provides efficient pairing with the "knob" mutation to promote heterodimerization of the Fc domain.

[0330] In certain embodiments, "bispecific antibodies (duobody)" can also be used to promote heterodimerization of Fc domains (Labrijn et al., (2013) Proc. Natl. Acad. Sci. USA 110, 5145-5150, the entire contents of which are incorporated herein by reference). For example, the Fc domain of the first interferon receptor Fc construct can include a F405L mutation, while the Fc domain of the second interferon receptor Fc construct can include a K409R mutation.

[0331] In other embodiments, "azymetric mutations" can be introduced into the interferon receptor Fc construct to promote the formation of heterodimers (Von Kreudenstein et al., (2013) mAbs 5, 646-654, the entire contents of which are incorporated herein by reference; and US 2012 / 0149876 A1, US 2017 / 0158779, US 9574010 and US 9562109, each of which is incorporated herein by reference in its entirety). In some embodiments, the azymetric mutations include T350V, L351Y, F405A and Y407V in the Fc domain of the first interferon receptor Fc construct, and T350V, T366L, K392L and T394W in the Fc domain of the second interferon receptor Fc construct.

[0332] In some embodiments, "charge pair" mutations determined by rational design of electrostatic steering mutations can also be introduced into interferon receptor Fc constructs to promote heterodimerization (Gunasekaran et al., (2010) J. Biol. Chem. 285, 19637-19646 and Strop et al., (2012) J. Mol. Biol. 420, 204-219, both of which are incorporated herein by reference). "Charge pair" mutations produce altered charge polarity on the Fc dimer interface, so that co-expression of electrostatically matched Fc chains supports favorable attractive interactions, thereby promoting the formation of Fc heterodimers. Unfavorable repulsive charge interactions inhibit the formation of homodimers (Gunasekaran et al., (2010)). For example, in some embodiments, the Fc domain of the first interferon receptor Fc construct may include K409D and K392D, and the Fc domain of the second interferon receptor Fc construct may include D399K and E356K. In other embodiments, the Fc domain of the first interferon receptor Fc construct may include D221E, P228E, L368E, and the Fc domain of the second interferon receptor Fc construct may include D221R, P228R and K409R.

[0333] Heterodimerization of the Fc domains of at least two interferon receptor Fc constructs can also be promoted by introducing "HA-TF" mutations (Moore et al., (2011) mAbs 3, 546-557, the entire contents of which are incorporated herein by reference). For example, in some embodiments, the Fc domain of the first interferon receptor Fc construct includes S364H and F405A, while the Fc domain of the second interferon receptor Fc construct includes Y349T and T394F.

[0334] The formation of heterodimeric soluble interferon receptors can also be promoted by utilizing the structural similarity and sequence divergence between different classes of immunoglobulins. For example, the SEED platform utilizes the sequence divergence but structural similarity of the CH3 domains of IgG and IgA (Davis et al., (2010) Protein Eng. Des. Sel. 23, 195-202, the entire contents of which are incorporated herein by reference). In some embodiments, the Fc domain of the first interferon receptor Fc construct includes an IgG / A chimera, and the Fc domain of the second interferon receptor Fc construct includes an IgA / G chimera.

[0335] In other embodiments, the inability of IgG3 to bind to protein A can be used for differential labeling of the Fc domain to enable efficient purification of heterodimers (Davis et al., 2013, U.S. Patent No. 8,586,713, the entire contents of which are incorporated herein by reference). For example, in some embodiments, the Fc domain of the interferon receptor Fc construct includes the H354R mutation.

[0336] In some embodiments, CH3 domain mutations can be introduced into IgG1, IgG2 and IgG4 constant regions to promote heterodimerization. In some embodiments, the Fc domain of the first interferon receptor Fc construct may include a 407A replacement, and the Fc domain of the second interferon receptor Fc construct may include a 366V or 366M replacement and a 409V replacement. In some embodiments, the Fc domain of the first interferon receptor Fc construct may include a 407A replacement and one or more replacements selected from the following replacement groups: 356G, 357D, 360D, 364Q, 364R and 399M. In some embodiments, the Fc domain of the second interferon receptor Fc construct may include a 366V or 366M replacement and a 409V replacement and one or more replacements selected from the following replacement groups: 345R, 347R, 349S, 366V, 370Y and 399M. In some embodiments, the Fc domain of the second interferon receptor Fc construct may include 366V and 409V substitutions and one or more substitutions selected from the following substitution groups: 345R, 347R, 349S, 366V, 370Y and 399M. In some embodiments, the Fc domain of the second interferon receptor Fc construct may include 366M and 409V substitutions and one or more substitutions selected from the following substitution groups: 345R, 347R, 349S, 366V, 370Y and 399M (see US2018 / 0009908 and Lewis et al., nature Biotechnology (2014) 32 (2), 191-198, both of which are incorporated herein by reference in their entirety).

[0337] (ii) Attachment of IgG

[0338] In some embodiments, the Fc domain can be engineered to include two different binding domains by attaching one or more binding domains to the amino and / or carboxyl termini of the Fc domain. In some embodiments, the one or more binding domains can be attached to the Fc domain via a peptide linker.

[0339] In some embodiments, the binding domain is an IFNAR (eg, IFNAR1 or IFNAR2) or a variant or fragment thereof. In some embodiments, the binding domain is the extracellular domain of IFNAR1 or IFNAR2.

[0340] (iii) Bispecific fragments

[0341] In some embodiments, the soluble interferon receptor may lack some or all of the Fc domain. In some embodiments, the binding domain of the soluble interferon receptor lacking some or all of the Fc domain and the partial Fc domain can be connected by a peptide linker. In some embodiments, the soluble interferon receptor can be produced using a polypeptide linker to connect each domain (e.g., binding domain, Fc domain) of the soluble interferon receptor and thereby generate a single polypeptide chain.

[0342] In some embodiments, the binding domain is an IFNAR (eg, IFNAR1 or IFNAR2) or a variant or fragment thereof. In some embodiments, the binding domain is the extracellular domain of IFNAR1 or IFNAR2.

[0343] (iv) Bispecific fusion proteins

[0344] In some embodiments, the binding domain of soluble interferon receptor is connected with other proteins to increase additional functionality or specificity. In some embodiments, the binding domain is IFNAR (e.g., IFNAR1 or IFNAR2) or its variant or fragment. In some embodiments, the binding domain is the extracellular domain of IFNAR1 or IFNAR2.

[0345] (v) Bispecific conjugates

[0346] In some embodiments, the binding domains of soluble interferon receptors are chemically conjugated to each other and / or to the Fc domain. In some embodiments, the binding domain is IFNAR (e.g., IFNAR1 or IFNAR2) or a variant or fragment thereof. In some embodiments, the binding domain is the extracellular domain of IFNAR1 or IFNAR2.

[0347] (vi) IgG4 Fab arm exchange

[0348] In some embodiments, Fab arm exchange can be used to promote the heterodimerization of Fc domain. Fab arm exchange is a post-translational modification of IgG4 antibody, which includes the third constant domain of IgG4 in addition to the hinge region of IgG4 and requires a reducing environment to be activated (van der Neut Kolfschoten et al. (2007) Science 317, 1554-1557, the entire contents of which are incorporated herein by reference). IgG4 antibody exchanges Fab arms by exchanging the heavy chain and the light chain connected with the heavy chain of another molecule, which produces bispecific antibodies (van der Neut Kolfschoten et al. (2007)). In some embodiments, the CH3 domain in IgG1 Fc can be replaced with the CH3 domain in IgG4 Fc and the IgG1 core hinge sequence can be replaced with the IgG4 sequence (i.e., by replacing Pro228 (P228S) with Ser) to promote the heterodimerization of soluble interferon receptor of the present disclosure.

[0349] (vii) Other alternative stents

[0350] In some embodiments, engineered protein scaffolds (eg, Affibodies, DARPins, Adnectins) can be used to generate molecules having at least two IFNAR binding domains. In some embodiments, the binding domain is the extracellular domain of IFNAR1 or IFNAR2.

[0351] PK part

[0352] In some embodiments, the soluble interferon receptor is operably coupled to a PK moiety that serves as a scaffold as well as a means to increase the serum half-life of the soluble interferon receptor.

[0353] Suitable PK moieties are well known in the art and include, but are not limited to, albumin, transferrin, Fc and variants thereof, and polyethylene glycol (PEG) and its derivatives. Suitable PK moieties include, but are not limited to, HSA or variants or fragments thereof, such as those disclosed in US 5,876,969, WO 2011 / 124718, and WO 2011 / 0514789; Fc and Fc variants, such as those disclosed in WO2011 / 053982, WO 02 / 060955, WO 02 / 096948, WO05 / 047327, WO05 / 018572, and US 2007 / 0111281; transferrin or variants or fragments thereof, such as those disclosed in US 7,176,278 and US 8,158,579; and PEG or derivatives, such as those disclosed in Zalipsky et al. ("Use of Functionalized Poly(Ethylene Glycols) for Modification of Polypeptides" in Polyethylene Glycol Chemistry: Biotechnical and Biomedical Applications, JM Harris, Plenus Press, New York (1992)), Zalipsky et al. (Advanced Drug Reviews 1995: 16: 157-182), and those disclosed in U.S. Pat. Nos. 4,640,835, 4,496,689, 4,301,144, 4,670,417, 4,791,192, 4,179,337 and 5,932,462 (the contents of the above are incorporated herein by reference). It is within the capabilities of a skilled artisan to introduce (e.g., clone, conjugate) a PK moiety into a soluble interferon receptor of the present invention using conventional methods.

[0354] In some embodiments, the PK moiety is HSA, which is naturally non-glycosylated.

[0355] In some embodiments, the PK moiety is wild-type Fc (SEQ ID NO: 26).

[0356] In certain embodiments, the Fc domain is altered or modified by, for example, amino acid mutations (e.g., additions, deletions, or substitutions). As used herein, the term "Fc domain variant" refers to an Fc domain having at least one amino acid modification, such as an amino acid substitution, compared to the wild-type Fc from which the Fc domain is derived. For example, in the case where the Fc domain is derived from a human IgG1 antibody, the variant comprises at least one amino acid mutation (e.g., substitution) compared to the wild-type amino acid at the corresponding position of the human IgG1 Fc region.

[0357] In some embodiments, the PK moiety is any of the Fc variants described herein.

[0358] In some embodiments, the PK moiety is wild-type HST. In other embodiments, the PK moiety is a HST with mutations of N413 and / or N611 and / or S12 (S12 is a potential O-linked glycosylation site), thereby generating HST with altered glycosylation (i.e., HST N413S, HST N611S, HST N413S / N611S, and HST S12A / N413S / N611S).

[0359] Exemplary Soluble Interferon Receptors

[0360] The soluble interferon receptors of the present disclosure can be configured to be incorporated into various interferon receptor Fc constructs. Likewise, the interferon receptor Fc constructs of the present invention can be configured to be incorporated into various domains.

[0361] For example, in one embodiment, the interferon receptor Fc construct may include the IFNAR1 domain shown in (SEQ ID NO: 11). In another embodiment, the interferon receptor Fc construct may include the IFNAR2 domain shown in (SEQ ID NO: 12). In another embodiment, the interferon receptor Fc construct may include a linker domain. In another embodiment, the IFNAR1 domain is operably coupled to the mutant Fc domain with a linker domain (e.g., a polypeptide linker). In some embodiments, the IFNAR2 domain is operably coupled to the mutant Fc domain with a linker domain (e.g., a polypeptide linker). In some embodiments, the linker domain is a polypeptide linker (e.g., a Gly / ser linker, e.g., (G4S) n, wherein n is 1-10, 2-5, 1, 2, 3, 4 or 5). In some embodiments, the length of the polypeptide linker is about 1-50, about 5-40, about 10-30 or about 15-20 amino acids. In some aspects, the length of the polypeptide linker is about 20 or less amino acids, about 15 or less amino acids, about 10 or less amino acids or about 5 or less amino acids. In some aspects, the polypeptide linker is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 amino acids in length. In another embodiment, the interferon receptor Fc construct may include the (Gly4Ser)4 linker domain shown in SEQ ID NO: 15. In another embodiment, the interferon receptor Fc construct may include a (Gly4Ser)2 linker domain as shown in SEQ ID NO: 38. In another embodiment, the interferon receptor Fc construct does not include a linker domain.

[0362] In another embodiment, the interferon receptor Fc construct may include the leader sequence shown in SEQ ID NO:13.

[0363] In some embodiments, the interferon receptor Fc construct may include an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W as shown in (SEQ ID NO: 10). In some embodiments, the interferon receptor Fc construct may include an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V as shown in (SEQ ID NO: 9). In some embodiments, the interferon Fc receptor construct may include an IgG1 Fc domain comprising mutation T366Y as shown in SEQ ID NO: 106, and optionally one or more mutations selected from C220S, P238S and P331S. In some embodiments, the interferon Fc receptor construct may include an IgG1 Fc domain comprising a mutation Y407T as shown in SEQ ID NO: 107, and optionally one or more mutations selected from C220S, P238S and P331S. In some embodiments, the interferon Fc receptor construct may include an IgG1 Fc domain comprising a mutation T366W as shown in SEQ ID NO: 108, and optionally one or more mutations selected from C220S, P238S and P331S. In some embodiments, the interferon Fc construct may include an IgG1 Fc domain comprising mutations T366S, L368A and Y407V as shown in SEQ ID NO: 109, and optionally one or more mutations selected from C220S, P238S and P331S.

[0364] In some embodiments, the interferon Fc receptor construct comprises a mutant IgG4 Fc domain comprising mutations S228P, F296Y, E356K, R409K, H435R and L445P according to EU numbering. In some embodiments, the heterodimer comprises a mutant IgG4 Fc domain comprising mutations T366S, L368A and Y407V according to EU numbering. In some embodiments, the interferon Fc construct comprises an IgG4 Fc domain comprising mutations S228P, F234A, L235A, L445P (EU) and K478del (Kabat) as shown in SEQ ID NO: 113. In some embodiments, the interferon Fc construct comprises an IgG4 Fc domain having mutations F296Y, E356K, R409K and H435R (EU) as shown in SEQ ID NO: 116. In some embodiments, the interferon Fc construct comprises an IgG4 Fc domain having mutations F296Y, R409K and K439E (EU) as shown in SEQ ID NO: 117. In some embodiments, the interferon Fc construct comprises an IgG4 Fc domain having mutations S228P, F296Y, E356K, R409K, H435R, L445P, G446del (EU) and K478del (Kabat) as shown in SEQ ID NO: 118. In some embodiments, the interferon Fc construct comprises an IgG4 Fc domain having mutations S228P, F296Y, R409K, K439E, L445P, G446del (EU) and K478del (Kabat) as shown in SEQ ID NO: 119.

[0365] Those skilled in the art will appreciate that these single domains can be operably coupled to each other in any order to form an interferon receptor Fc construct with enzymatic activity. For example, as described in detail in the specific examples below, the IFNAR1 extracellular domain can be operably coupled to the Fc domain via (Gly4Ser)4 joint domains. In another example, the IFNAR2 extracellular domain can be operably coupled to the Fc domain by (Gly4Ser)4 joint domains. In another example, the IFNAR1 extracellular domain can be operably coupled to the Fc domain by (Gly4Ser)2 joint domains. In another example, the IFNAR2 extracellular domain can be operably coupled to the Fc domain by (Gly4Ser)2 joint domains. In another example, the IFNAR1 extracellular domain can be operably coupled to the Fc domain. In another example, the IFNAR2 extracellular domain can be operably coupled to the Fc domain by (Gly4Ser)2 joint domains. In another example, the IFNAR1 extracellular domain can be operably coupled to the Fc domain. In another example, the IFNAR2 extracellular domain can be operably coupled to the Fc domain. Various other configurations are possible, with Figure 1and non-limiting exemplary configurations disclosed in the Sequence Listing.

[0366] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0367] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0368] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W, with or without a leader sequence, in the absence of a linker domain.

[0369] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0370] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0371] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W, with or without a leader sequence, in the absence of a linker domain.

[0372] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0373] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0374] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V, without a linker domain, with or without a leader sequence.

[0375] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0376] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0377] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V, with or without a leader sequence, in the absence of a linker domain.

[0378] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence via a linker domain, such as a (Gly4Ser)4 linker domain.

[0379] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence via a linker domain, such as a (Gly4Ser)2 linker domain.

[0380] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising the mutation T366Y, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence.

[0381] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence via a linker domain, such as a (Gly4Ser)4 linker domain.

[0382] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence via a linker domain, such as a (Gly4Ser)2 linker domain.

[0383] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising the mutation T366Y, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence.

[0384] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation Y407T, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0385] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation Y407T, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0386] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising the mutation Y407T, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence.

[0387] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation Y407T, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0388] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation Y407T, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0389] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising the mutation Y407T, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence.

[0390] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0391] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0392] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising the mutation T366W, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence.

[0393] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0394] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0395] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising the mutation T366W and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence.

[0396] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0397] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0398] In some embodiments, the interferon receptor Fc construct comprises an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain without a linker domain, the IgG1 Fc domain comprising mutations T366S, L368A, and Y407V, and optionally one or more mutations selected from C220S, P238S, and P331S, with or without a leader sequence.

[0399] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence via a linker domain, such as a (Gly4Ser)4 linker domain.

[0400] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A and Y407V, and optionally one or more mutations selected from C220S, P238S and P331S, with or without a leader sequence via a linker domain, such as a (Gly4Ser)2 linker domain.

[0401] In some embodiments, the interferon receptor Fc construct comprises an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A, and Y407V and optionally one or more mutations selected from C220S, P238S, and P331S, with or without a leader sequence, in the absence of a linker domain.

[0402] In some embodiments, the interferon receptor Fc construct comprises IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0403] In some embodiments, the interferon receptor Fc construct comprises IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0404] In some embodiments, the interferon receptor Fc construct comprises IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R, with or without a leader sequence, without a linker domain.

[0405] In some embodiments, the interferon receptor Fc construct comprises IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0406] In some embodiments, the interferon receptor Fc construct comprises IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0407] In some embodiments, the interferon receptor Fc construct comprises IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R, with or without a leader sequence, without a linker domain.

[0408] In some embodiments, the interferon receptor Fc construct comprises IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0409] In some embodiments, the interferon receptor Fc construct comprises IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0410] In some embodiments, the interferon receptor Fc construct comprises IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E, with or without a leader sequence, without a linker domain.

[0411] In some embodiments, the interferon receptor Fc construct comprises IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)4 linker domain.

[0412] In some embodiments, the interferon receptor Fc construct comprises IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E, with or without a leader sequence, via a linker domain, such as a (Gly4Ser)2 linker domain.

[0413] In some embodiments, the interferon receptor Fc construct comprises IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E, with or without a leader sequence, without a linker domain.

[0414] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 1 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 1.

[0415] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 2 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 2.

[0416] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 3 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 3.

[0417] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 4 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 4.

[0418] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 52 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 52.

[0419] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 54 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 54.

[0420] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 56 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 56.

[0421] In some embodiments, the interferon receptor Fc construct comprises the amino acid sequence shown in SEQ ID NO: 58 with or without a leader sequence, and a nucleic acid encoding the amino acid sequence shown in SEQ ID NO: 58.

[0422] In some embodiments, the interferon receptor Fc construct dimerizes to form a soluble interferon receptor, eg, a heterodimeric soluble interferon receptor. For example, in some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V via a linker domain, such as a (Gly4Ser)4 linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V via a linker domain, such as a (Gly4Ser)4 linker domain.

[0423] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V via a linker domain, such as a (Gly4Ser)2 linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V via a linker domain, such as a (Gly4Ser)2 linker domain.

[0424] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V in the absence of a linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, T366L, K392L and T394W in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations C220S, P238S, P331S and T350V, L351Y, F405A and Y407V in the absence of a linker domain.

[0425] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain coupled to an IgG1 Fc domain comprising the mutation Y407T via a linker domain, such as a (Gly4Ser)4 linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation Y407T via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y via a linker domain, such as a (Gly4Ser)4 linker domain.

[0426] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation Y407T via a linker domain, such as a (Gly4Ser)2 linker domain. For example, in some embodiments, the soluble interferon receptor Fc construct comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation Y407T via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y via a linker domain, such as a (Gly4Ser)2 linker domain.

[0427] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation Y407T in the absence of a linker domain. For example, in some embodiments, the soluble interferon receptor Fc construct comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an Fc domain comprising the mutation Y407T in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366Y in the absence of a linker domain.

[0428] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutations T366S, L368A, and Y407V via a linker domain, such as a (Gly4Ser)4 linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A, and Y407V via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation T366W via a linker domain, such as a (Gly4Ser)4 linker domain.

[0429] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutations T366S, L368A, and Y407V via a linker domain, such as a (Gly4Ser)2 linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A, and Y407V via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation T366W via a linker domain, such as a (Gly4Ser)2 linker domain.

[0430] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutation T366W in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising the mutations T366S, L368A, and Y407V in the absence of a linker domain. For example, in some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR1 extracellular domain operably coupled to an IgG1 Fc domain comprising mutations T366S, L368A, and Y407V in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising an IFNAR2 extracellular domain operably coupled to an IgG1 Fc domain comprising mutation T366W in the absence of a linker domain.

[0431] In some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E via a linker domain, such as a (Gly4Ser)4 linker domain. For example, in some embodiments, a soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E via a linker domain, such as a (Gly4Ser)4 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R via a linker domain, such as a (Gly4Ser)4 linker domain.

[0432] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E via a linker domain, such as a (Gly4Ser)2 linker domain. For example, in some embodiments, the soluble interferon receptor Fc construct comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E via a linker domain, such as a (Gly4Ser)2 linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R via a linker domain, such as a (Gly4Ser)2 linker domain.

[0433] In some embodiments, the soluble interferon receptor comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E in the absence of a linker domain. For example, in some embodiments, the soluble interferon receptor Fc construct comprises a heterodimer comprising an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR1 operably coupled to an IgG4 Fc domain comprising mutations F296Y, R409K, and K439E in the absence of a linker domain, and an interferon receptor Fc construct (with or without a leader sequence) comprising IFNAR2 operably coupled to an IgG4 Fc domain comprising mutations F296Y, E356K, R409K, and H435R in the absence of a linker domain.

[0434] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:1 and a polypeptide comprising the amino acid sequence of SEQ ID NO:4.

[0435] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:2 and a polypeptide comprising the amino acid sequence of SEQ ID NO:3.

[0436] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:52 and a polypeptide comprising the amino acid sequence of SEQ ID NO:54.

[0437] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:56 and a polypeptide comprising the amino acid sequence of SEQ ID NO:58.

[0438] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:40 and a polypeptide comprising the amino acid sequence of SEQ ID NO:43.

[0439] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:41 and a polypeptide comprising the amino acid sequence of SEQ ID NO:42.

[0440] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:53 and a polypeptide comprising the amino acid sequence of SEQ ID NO:55.

[0441] In some embodiments, the soluble interferon receptor is a heterodimer comprising a polypeptide comprising the amino acid sequence of SEQ ID NO:57 and a polypeptide comprising the amino acid sequence of SEQ ID NO:59.

[0442] In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence at least 70% identical to the amino acid sequence of SEQ ID NO: 1, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence at least 70% identical to the amino acid sequence of SEQ ID NO: 2, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 3. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 4. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 52. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 54. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 56. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 58.In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 40. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 43. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence at least 70% identical to the amino acid sequence of SEQ ID NO: 41, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence at least 70% identical to the amino acid sequence of SEQ ID NO: 42, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 53. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, such as 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 55. In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical to the amino acid sequence of SEQ ID NO: 57, e.g., 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical.In some embodiments, the interferon receptor Fc construct comprises a polypeptide having an amino acid sequence that is at least 70% identical, e.g., 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or at least 99.5% identical to the amino acid sequence of SEQ ID NO: 59. In some embodiments, the polypeptide comprises an amino acid sequence selected from any one of SEQ ID NO: 1-4, 52, 54, 56, 58, 40, 43, 41, 42, 43, 55, 57 or 59.

[0443] In some embodiments, the interferon receptor Fc construct has a leader sequence. In some embodiments, the interferon receptor Fc construct does not have a leader sequence.

[0444] Those of ordinary skill in the art will appreciate that leader sequences and linker sequences are optional and are not limited to those described in the above embodiments. For example, IFNAR domains (e.g., IFNAR1, IFNAR2) can be directly fused to the N- and / or C-terminal of the Fc domain or its variant or fragment; the leader domain can be any of those leader domains known in the art that can be used for its intended purpose, for example, to increase protein expression and / or secretion (e.g., MDWTWRILFLVAAATGTHA; SEQ ID NO: 13); the linker can be any linker known in the art, for example, (Gly4Ser) n, NLG (VDGASSPVNVSSPSVQDI; SEQ ID NO: 18), LE, thrombin-sensitive disulfide cyclic peptide linker, LEA (EAAAK) 4ALEA (EAAAK) 4 (SEQ ID NO: 19) or a disulfide linker that can be cleaved in vivo as described herein. It should also be understood that the use of conventional cloning and recombination methods to make corresponding changes to the amino acid sequence of the interferon receptor Fc construct is within the capabilities of the technician.

[0445] Method for preparing soluble interferon receptor

[0446] Recombinant DNA technology can be used to prepare the interferon receptor Fc construct of the present disclosure in a large number of transformed or transfected host cells. For this reason, a recombinant DNA molecule encoding the peptide is prepared. The method for preparing such DNA molecules is well known in the art. For example, a suitable restriction enzyme can be used to cut the sequence encoding the interferon receptor Fc construct from DNA. Alternatively, chemical synthesis techniques, such as phosphoramidate method synthetic DNA molecules, can be used. Moreover, a combination of these technologies can be used.

[0447] The present invention also includes vectors capable of expressing interferon receptor Fc constructs in suitable hosts. The vector comprises a DNA molecule encoding interferon receptor Fc constructs operably coupled to a suitable expression control sequence. Before or after the DNA molecule is inserted into the vector, the method for achieving such effective connection is well known. The expression control sequence comprises a promoter, an activator, an enhancer, an operator, a ribosomal nuclease domain, a start signal, a stop signal, a cap signal, a polyadenylation signal and other signals related to the control of transcription or translation.

[0448] The resulting vector with the DNA molecule thereon is used to transform or transfect a suitable host. In some embodiments, the soluble interferon receptor of the present disclosure can be prepared by co-transfecting or co-transforming two or more expression vectors comprising the DNA encoding the interferon receptor Fc construct into a suitable host. Such transformation or transfection can be performed using methods well known in the art.

[0449] Any of a large number of available and well-known host cells can be used to implement the present invention. The selection of a specific host depends on many factors recognized in the art. These include, for example, compatibility with the selected expression vector, toxicity of the interferon receptor Fc construct encoded by the DNA molecule, the rate of conversion or transfection, the recovery convenience of the interferon receptor Fc construct, expression characteristics, biosafety and cost. Under the premise that it is understood that not all hosts may be equally effective for the expression of a specific DNA sequence, it is necessary to balance between these factors. Within these general criteria, useful microbial hosts include bacteria (such as Escherichia coli), yeast (such as saccharomyces cerevisiae) and other fungi, insects, plants, mammalian (including humans) cells in culture or other hosts known in the art. In some embodiments, the interferon receptor Fc construct is produced in CHO cells.

[0450] Next, the transformed or transfected host is cultured and purified. The host cell can be cultured under conventional fermentation or culture conditions to express the desired compound. Such fermentation and culture conditions are well known in the art. Finally, the interferon receptor Fc construct is purified from the culture by methods well known in the art.

[0451] Compounds can also be prepared by synthetic methods. For example, solid phase synthesis techniques can be used. Suitable techniques are well known in the art and include those described in Merrifield (1973), Chem. Polypeptides, pp. 335-61 (Katsoyannis and Panayotis eds.); Merrifield (1963), J. Am. Chem. Soc. 85: 2149; Davis et al., Biochem Intl 1985; 10: 394-414; Stewart and Young (1969), Solid Phase Peptide Synthesis; U.S. Patent No. 3,941,763; Finn et al. (1976), The Proteins (3rd ed.) 2: 105-253; and Erickson et al. (1976), The Proteins (3rd ed.) 2: 257-527. In some embodiments, derivatized peptide-containing or non-peptide-containing compounds can be synthesized by well-known organic chemistry techniques.

[0452] Other methods of molecule expression / synthesis are generally known to those of ordinary skill in the art.

[0453] Soluble interferon receptor with altered glycosylation

[0454] Glycosylation (e.g., O-linked or N-linked glycosylation) can, for example, affect the serum half-life of the soluble interferon receptor of the present disclosure by minimizing its removal from the circulation through mannose and asialoglycoprotein receptors and other lectin-like receptors. Therefore, in some embodiments, the soluble interferon receptor of the present disclosure is prepared in a non-glycosylated, deglycosylated or hypoglycosylated form. Preferably, the N-linked glycosylation is altered and the soluble interferon receptor is non-glycosylated.

[0455] In some embodiments, all asparagine residues in the soluble interferon receptor that conform to the consensus sequence of Asn-X-Ser / Thr (X can be any other naturally occurring amino acid other than Pro) are mutated to residues that do not serve as receptors for N-linked glycosylation (e.g., serine, glutamine), thereby eliminating glycosylation of the soluble interferon receptor when synthesized in a cell that glycosylates proteins.

[0456] In some embodiments, a soluble interferon receptor lacking an N-linked glycosylation site is produced in a mammalian cell. In one embodiment, the mammalian cell is a CHO cell. Thus, in a specific embodiment, a soluble interferon receptor without glycosylation is produced in a CHO cell.

[0457] In other embodiments, reduction or absence of N-glycosylation is achieved by producing a soluble interferon receptor, for example, in a host (e.g., a bacterium such as E. coli), a mammalian cell engineered to lack one or more enzymes important for glycosylation, or a mammalian cell treated with an agent that prevents glycosylation, such as tunicamycin (an inhibitor of Dol-PP-GlcNAc formation).

[0458] In some embodiments, soluble interferon receptors are produced in lower eukaryotic organisms engineered to produce glycoproteins with complex N-glycans rather than high mannose-type sugars (see, eg, US 2007 / 0105127).

[0459] In some embodiments, glycosylated soluble interferon receptors (e.g., those produced in mammalian cells such as CHO cells) are chemically or enzymatically treated to remove one or more sugar residues (e.g., one or more mannose, fucose and / or N-acetylglucosamine residues) or to modify or mask one or more sugar residues. Such modification or masking can reduce the binding of soluble interferon receptors to mannose receptors, and / or asialoglycoprotein receptors, and / or other lectin-like receptors. Chemical deglycosylation can be achieved by treating soluble interferon receptors with trifluoromethanesulfonic acid (TFMS) as disclosed in, for example, Sojar et al., JBC 1989; 264: 2552-9 and Sojar et al., Methods Enzymol 1987; 138: 341-50, or by treating with hydrogen fluoride as disclosed in Sojar et al. (1987, supra). As disclosed in Thotakura et al. (Methods Enzymol 1987; 138: 350-9), N-linked carbohydrates can be enzymatically removed from soluble interferon receptors by treating the soluble interferon receptors with protein N-glycosidase (PNGase) A or F. Other suitable commercially available deglycosylation enzymes recognized in the art include: endo-α-N-acetylgalactosaminidase, endoglycosidase F1, endoglycosidase F2, endoglycosidase F3, and endoglycosidase H. In some embodiments, one or more of these enzymes can be used to deglycosylate the soluble interferon receptors of the present disclosure. For example, alternative methods for deglycosylation are disclosed in US 8,198,063.

[0460] In some embodiments, the soluble interferon receptor is partially deglycosylated. Partial deglycosylation can be achieved by treating the soluble interferon receptor with an endoglycosidase (e.g., endoglycosidase H), which cuts the high mannose carbohydrates connected by N-, but does not cut the complex type of carbohydrates, thereby leaving a single GlcNAc residue connected to asparagine. The soluble interferon receptor treated with endoglycosidase H will lack high mannose carbohydrates, thereby resulting in a reduction in the interaction with the liver mannose receptor. Although the receptor recognizes terminal GlcNAc, the probability of productive interaction with a single GlcNAc on the protein surface is not as high as a complete high mannose structure.

[0461] In other embodiments, the glycosylation of the soluble interferon receptor is modified, for example, by oxidation, reduction, dehydration, substitution, esterification, alkylation, sialylation, carbon-carbon bond cleavage, etc. to reduce the clearance of soluble interferon from the blood. In some embodiments, the soluble interferon receptor is treated with periodate and sodium borohydride to modify the carbohydrate structure. Periodate treatment oxidizes vicinal diols, thereby cleaving carbon-carbon bonds and replacing hydroxyls with aldehyde groups; borohydrides reduce aldehydes to hydroxyls. Many sugar residues include vicinal diols and are therefore cleaved by this treatment. The use of periodate and sodium borohydride to extend serum half-life is exemplified by sequential treatment of the lysosomal enzyme β-glucuronidase with these agents (see, e.g., Houba et al. (1996) Bioconjug Chem 1996:7:606-11; Stahl et al. PNAS 1976;73:4045-9; Achord et al. Pediat. Res 1977;11:816-22; Achord et al. Cell 1978;15:269-78). Treatment with periodate and sodium borohydride is disclosed in Hickman et al., BBRC 1974;57:55-61. Treatment with periodate and cyanoborohydride, which increases the serum half-life and tissue distribution of ricin, is disclosed in Thorpe et al. Eur J Biochem 1985;147:197-206.

[0462] In one embodiment, the carbohydrate structure of a soluble interferon receptor can be masked by adding one or more additional moieties (e.g., carbohydrate groups, phosphate groups, alkyl groups, etc.) that interfere with recognition of the structure by mannose or asialoglycoprotein receptors or other lectin-like receptors.

[0463] In some embodiments, one or more potential glycosylation sites are removed by mutation of nucleic acids encoding soluble interferon receptors, thereby reducing the glycosylation (hypoglycosylation) of soluble interferon receptors when synthesized in cells that glycosylate proteins, for example, mammalian cells such as CHO cells. In some embodiments, if, for example, a soluble interferon receptor with low glycosylation exhibits increased activity or causes an increase in serum half-life, it may be necessary to selectively make the soluble interferon receptor low glycosylation by mutating potential N-linked glycosylation sites therein. In other embodiments, if, for example, low glycosylation improves the serum half-life of a soluble interferon receptor, it may be desirable to modify the portion of the soluble interferon receptor so that certain domains lack N-glycosylation. Alternatively, other amino acids near the glycosylated receptor may be modified to destroy the recognition motif of a glycosylated enzyme without having to change the amino acids that are usually glycosylated.

[0464] In some embodiments, the glycosylation of the soluble interferon receptor can be changed by introducing a glycosylation site. For example, the amino acid sequence of the soluble interferon receptor can be modified to introduce a consensus site for N-linked glycosylation of Asn-X-Ser / Thr (X is any amino acid other than proline). Additional N-linked glycosylation sites can be added at any position in the entire amino acid sequence of the soluble interferon receptor. Preferably, the glycosylation site is introduced at a position in the amino acid sequence that does not substantially reduce the activity of the soluble interferon receptor.

[0465] It has been reported that the addition of O-linked glycosylation sites changes the serum half-life of proteins, such as growth hormone, follicle stimulating hormone, IGFBP-6, factor IX, and many other proteins (e.g., as disclosed in Okada et al., Endocr Rev 2011; 32: 2-342; Weenen et al., J Clin Endocrinol Metab 2004; 89: 5204-12; Marinaro et al., European Journal of Endocrinology 2000; 142: 512-6; US 2011 / 0154516). Thus, in some embodiments, the O-linked glycosylation (on serine / threonine residues) of the soluble interferon receptor is altered. Methods for altering O-linked glycosylation are routine in the art and can be, for example, by β-elimination (see, e.g., Huang et al., Rapid Communications in Mass Spectrometry 2002; 16: 1199-204; Conrad, Curr Protoc Mol Biol 2001; Chapter 17: Unit 17.15A; Fukuda, Curr Protoc Mol Biol 2001; Chapter 17; Unit 17.15B; Zachara et al., Curr Protoc Mol Biol 2011; Unit 17.6); by using commercially available kits (e.g., GlycoProfile TMBeta-Elimination Kit, Sigma); or by treating the soluble interferon receptor with a series of glycosidic exonucleases, such as, but not limited to, β1-4 galactosidase and β-N-acetylglucosaminidase, until only Galβ1-3GalNAc and / or GlcNAcβ1-3GalNAc is left, followed by treatment with, for example, endo-α-N-acetylgalactosaminidase (i.e., O-glycosidase). Such enzymes are commercially available from, for example, New England Biolabs. In yet other embodiments, the soluble interferon receptor is altered to introduce O-linked glycosylation in the soluble interferon receptor, as disclosed in, for example, Okada et al. (supra), Weenen et al. (supra), US2008 / 0274958, and US2011 / 0171218. In some embodiments, one or more O-linked glycosylation consensus sites are introduced into the soluble interferon receptor, such as CXXGGT / SC (SEQ ID NO: 29) (van den Steen et al., Critical Reviews in Biochemistry and Molecular Biology, Michael Cox, ed., 1998; 33: 151-208), NST-E / DA (SEQ ID NO: 30), NITQS (SEQ ID NO: 31), QSTQS (SEQ ID NO: 32), D / E-FT-R / KV (SEQ ID NO: 33), CE / D-SN (SEQ ID NO: 34) and GGSC-K / R (SEQ ID NO: 35). Additional O-linked glycosylation sites can be added at any position in the entire amino acid sequence of the soluble interferon receptor. Preferably, the glycosylation site is introduced at a position in the amino acid sequence that does not substantially reduce the activity of the soluble interferon receptor. Alternatively, O-linked sugar moieties may be introduced by chemically modifying the amino acids in a soluble interferon receptor as described, for example, in WO 87 / 05330 and Aplin et al., CRC Crit Rev Biochem 1981; 259-306.

[0466] In some embodiments, both N-linked and O-linked glycosylation sites are introduced into the soluble interferon receptor, preferably at positions in the amino acid sequence that do not substantially reduce the activity of the soluble interferon receptor.

[0467] It is within the capabilities of the skilled artisan to introduce, reduce or eliminate glycosylation (e.g., N-linked or O-linked glycosylation) in a soluble interferon receptor and determine whether such a change in glycosylation state increases or decreases the activity or serum half-life of the soluble interferon receptor using routine methods in the art.

[0468] In some embodiments, a soluble interferon receptor can comprise altered glycoforms (eg, hypofucosylated or fucose-free glycans).

[0469] In some embodiments, the serum half-life of the soluble interferon receptor with altered glycosylation is increased by at least about 1.5 times relative to the corresponding glycosylated soluble interferon receptor (e.g., a soluble interferon receptor in which the potential N-linked glycosylation site is not mutated), for example, at least 3 times, at least 5 times, at least 10 times, at least about 20 times, at least about 50 times, at least about 100 times, at least about 200 times, at least about 300 times, at least about 400 times, at least about 500 times, at least about 600 times, at least about 700 times, at least about 800 times, at least about 900 times, at least about 1000 times or 1000 times or more. Conventional methods recognized in the art can be used to determine the serum half-life of a soluble interferon receptor with an altered glycosylation state.

[0470] In some embodiments, a soluble interferon receptor with altered glycosylation (e.g., a soluble interferon receptor that is aglycosylated, deglycosylated, or hypoglycosylated) retains at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or 100% of the activity of a corresponding glycosylated soluble interferon receptor (e.g., a soluble interferon receptor in which there is no mutation in a potential N-linked glycosylation site).

[0471] In some embodiments, changing the glycosylation state of the soluble interferon receptor can increase its activity by directly increasing activity or increasing bioavailability (e.g., serum half-life). Therefore, in some embodiments, the activity of the soluble interferon receptor of the glycosylation change is increased by at least 1.3 times relative to the corresponding glycosylated soluble interferon receptor (e.g., wherein the potential N-connected glycosylation site has no soluble interferon receptor mutation), for example, at least 1.5 times, at least 2 times, at least 2.5 times. At least 3 times, at least 3.5 times, at least 4 times, at least 4.5 times, at least 5 times, at least 5.5 times, at least 6 times, at least 6.5 times, at least 7 times, at least 7.5 times, at least 8 times, at least 8.5 times, at least 9 times, at least 9.5 times or 10 times or more.

[0472] Technicians can easily determine the glycosylation state of soluble interferon receptors using methods recognized in the art. In a preferred embodiment, mass spectrometry is used to determine the glycosylation state. In other embodiments, the interaction with concanavalin A (Con A) can be evaluated to determine whether the soluble interferon receptor is hypoglycosylated. Compared with the corresponding glycosylated soluble interferon receptor, the hypoglycosylated soluble interferon receptor is expected to show reduced binding to Con A-agarose. SDS-PAGE analysis can also be used to compare the mobility of hypoglycosylated proteins and corresponding glycosylated proteins. Compared with glycosylated proteins, hypoglycosylated proteins are expected to have greater mobility in SDS-PAGE. For example, Roth et al., International Journal of Carbohydrate Chemistry 2012; 1-10 discloses other suitable methods recognized in the art for analyzing the glycosylation state of proteins.

[0473] The pharmacokinetics, such as serum half-life, of soluble interferon receptors with different glycosylation states can be determined using conventional methods, for example, by introducing soluble interferon receptors into mice (e.g., intravenously), obtaining blood samples at predetermined time points, and determining and comparing the levels and / or activity of the soluble interferon receptors in the samples.

[0474] Pharmaceutical composition

[0475] In certain embodiments, soluble interferon receptor is administered alone. In certain embodiments, soluble interferon receptor is administered before administering at least one other therapeutic agent. In certain embodiments, soluble interferon receptor is administered simultaneously with the administration of at least one other therapeutic agent. In certain embodiments, soluble interferon receptor is administered after administering at least one other therapeutic agent. In other embodiments, soluble interferon receptor is administered before administering at least one other therapeutic agent. As will be appreciated by those skilled in the art, in some embodiments, soluble interferon receptor is combined with other agents / compounds. In some embodiments, soluble interferon receptor and other agents are administered simultaneously. In some embodiments, soluble interferon receptor and other agents are not administered simultaneously, but soluble interferon receptor is administered before or after the agent is administered. In some embodiments, the subject receives both soluble interferon receptor and other agents in the same period of prevention, disease occurrence and / or treatment period.

[0476] The pharmaceutical composition of the present disclosure can be used in a combination therapy, i.e., in combination with other agents. In certain embodiments, the combination therapy includes a soluble interferon receptor in combination with at least one other agent. Agents include, but are not limited to, chemical compositions, antibodies, antigen binding regions, combinations thereof, and conjugates prepared by in vitro synthesis. In certain embodiments, agents can act as agonists, antagonists, allosteric modulators, or toxins.

[0477] In certain embodiments, the present disclosure provides a pharmaceutical composition comprising a soluble interferon receptor and a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative and / or adjuvant.

[0478] In certain embodiments, the present invention provides a pharmaceutical composition comprising a soluble interferon receptor and a therapeutically effective amount of at least one other therapeutic agent, as well as a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative and / or adjuvant.

[0479] In certain embodiments, acceptable formulation materials are preferably non-toxic to the recipient at the dosage and concentration employed. In some embodiments, formulation materials are used for sc and / or IV administration. In certain embodiments, pharmaceutical compositions may include formulation materials for changing, maintaining or preserving, for example, the pH, osmotic concentration, viscosity, clarity, color, isotonicity, odor, sterility, stability, dissolution rate or release rate, adsorption or penetration of the composition. In certain embodiments, suitable formulation materials include, but are not limited to, amino acids (e.g., glycine, glutamine, asparagine, arginine, or lysine); antimicrobial agents; antioxidants (e.g., ascorbic acid, sodium sulfite, or sodium bisulfite); buffers (e.g., borates, bicarbonates, Tris-HCl, citrates, phosphates, or other organic acids); bulking agents (e.g., mannitol or glycine); chelating agents (e.g., ethylenediaminetetraacetic acid (EDTA)); complexing agents (e.g., caffeine, polyvinylpyrrolidone, β-cyclodextrin, or hydroxypropyl-β-cyclodextrin); fillers; monosaccharides; disaccharides; and other carbohydrates (e.g., glucose, mannose, or dextrin); proteins (e.g., gelatin); coloring, flavoring, and diluents; emulsifiers; hydrophilic polymers (e.g., polyvinylpyrrolidone); low molecular weight the polypeptide; a salt-forming counterion (e.g., sodium); a preservative (e.g., benzalkonium chloride, benzoic acid, salicylic acid, thimerosal, phenylethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid, or hydrogen peroxide); a solvent (e.g., glycerol, propylene glycol, or polyethylene glycol); a sugar alcohol (e.g., mannitol or sorbitol); a suspending agent; a surfactant or wetting agent (e.g., pluronic, PEG, sorbitan esters, polysorbates such as polysorbate 20, polysorbate 80, triton, tromethamine, lecithin, cholesterol, tiloxapor); a stability enhancer (e.g., sucrose or sorbitol); a tonicity enhancer (e.g., an alkali metal halide, preferably sodium chloride or potassium chloride, mannitol, sorbitol); a delivery vehicle; a diluent; an excipient and / or a pharmaceutical adjuvant (Remington's Pharmaceutical Sciences, 18th Edition, AR Gennaro, ed., Mack Publishing Company (1995). In some embodiments, the formulation comprises PBS; 20 mM NaOAC, pH 5.2, 50 mM NaCl; and / or 10 mM NAOAC, pH 5.2, 9% sucrose.

[0480] In certain embodiments, the soluble interferon receptor and / or therapeutic molecule are connected to a half-life extension medium known in the art. Such vehicles include, but are not limited to, polyethylene glycol, glycogen (e.g., glycosylation of the soluble interferon receptor) and dextran. For example, such vehicles are described in U.S. Application Serial No. 09 / 428,082, current U.S. Patent No. 6,660,843 and disclosed application WO 99 / 25044.

[0481] In certain embodiments, the optimal pharmaceutical composition is determined by one skilled in the art based on, for example, the intended route of administration, delivery form, and desired dosage. See, for example, Remington's Pharmaceutical Sciences, supra. In certain embodiments, such compositions may affect the physical state, stability, in vivo release rate, and in vivo clearance of the antibodies of the invention.

[0482] In certain embodiments, the main vehicle or carrier in the pharmaceutical composition can be aqueous or non-aqueous in nature. For example, in certain embodiments, a suitable vehicle or carrier can be water for injection, saline solution or artificial cerebrospinal fluid, which may supplement other substances common in the composition for parenteral administration. In some embodiments, saline includes isotonic phosphate buffered saline. In certain embodiments, the pharmaceutical composition comprises a Tris buffer of about pH 7.0-8.5, or an acetate buffer of about pH 4.0-5.5, which may further include sorbitol or its suitable substitute. In certain embodiments, the composition comprising a soluble interferon receptor (with or without at least one other therapeutic agent) can be prepared for storage in the form of a lyophilized cake or aqueous solution by mixing the selected composition with the desired purity level with an optional formulation reagent (Remington's Pharmaceutical Sciences, the same). In addition, in certain embodiments, the composition comprising a soluble interferon receptor (with or without at least one other therapeutic agent) can be formulated as a lyophilized product using an appropriate excipient such as sucrose.

[0483] In certain embodiments, pharmaceutical compositions can be selected for parenteral delivery. In certain embodiments, compositions can be selected for suction or for delivery by the digestive tract, such as oral. The preparation of such pharmaceutically acceptable compositions is within the capabilities of those skilled in the art.

[0484] In certain embodiments, the formulation components are present in concentrations acceptable to the site of administration. In certain embodiments, a buffer is used to maintain the composition at physiological pH or slightly lower pH, typically in the pH range of about 5 to about 8.

[0485] In certain embodiments, when considering parenteral administration, the therapeutic composition can be in the form of a pyrogen-free, parenterally acceptable aqueous solution in a pharmaceutically acceptable carrier, which comprises the required soluble interferon receptor in a pharmaceutically acceptable medium, with or without another therapeutic agent. In certain embodiments, the medium for parenteral injection is sterile distilled water, wherein the soluble interferon receptor (with or without and at least one other therapeutic agent) is formulated into a sterile isotonic solution of suitable preservation. In certain embodiments, preparation can include preparing the required molecule together with reagents such as injectable microspheres, bioerodible particles, polymeric compounds (such as polylactic acid or polyglycolic acid), beads or liposomes, which can be used for the controlled or sustained release of the product that can then be delivered by long-acting injection. In certain embodiments, hyaluronic acid can also be used, and the effect of the duration in the circulation can be promoted. In certain embodiments, an implantable drug delivery device can be used for introducing the required molecule.

[0486] In certain embodiments, the pharmaceutical composition can be formulated for inhalation. In certain embodiments, the soluble interferon receptor can be formulated as a dry powder for inhalation with or without at least one additional therapeutic agent. In certain embodiments, an inhalation solution comprising a soluble interferon receptor with or without at least one additional therapeutic agent can be formulated for aerosol delivery with a propellant. In certain embodiments, the solution can be atomized. Pulmonary administration is further described in PCT Application No. PCT / US94 / 001875, which describes pulmonary delivery of chemically modified proteins.

[0487] In certain embodiments, it is expected that the preparation can be orally administered. In certain embodiments, the soluble interferon receptor with or without at least one other therapeutic agent used in this way can be prepared with or without those carriers that are generally used for solid dosage forms such as tablets and capsules. In certain embodiments, the capsule can be designed to release the active part of the preparation at a point when bioavailability is maximized in the gastrointestinal tract and degradation is minimized before the system. In certain embodiments, at least one other reagent may be included to promote the absorption of soluble interferon receptor and / or any other therapeutic agent. In certain embodiments, diluents, flavoring agents, low melting point waxes, vegetable oils, lubricants, suspending agents, tablet disintegrating agents and adhesives may also be used.

[0488] In certain embodiments, the pharmaceutical composition may involve an effective amount of a soluble interferon receptor in a mixture with a non-toxic excipient suitable for making tablets with or without at least one additional therapeutic agent. In certain embodiments, the solution may be prepared in unit dosage form by dissolving the tablet in sterile water or another suitable medium. In certain embodiments, suitable excipients include, but are not limited to, inert diluents such as calcium carbonate, sodium carbonate or sodium bicarbonate, lactose or calcium phosphate; or adhesives such as starch, gelatin or gum arabic; or lubricants such as magnesium stearate, stearic acid or talc.

[0489] For those skilled in the art, other pharmaceutical compositions will be apparent, including preparations including soluble interferon receptors in a sustained delivery or controlled delivery preparation with or without at least one other therapeutic agent. In certain embodiments, the technology for preparing a variety of other sustained or controlled delivery modes, such as liposome carriers, microparticles or porous beads and reservoir injections, is also known to those skilled in the art. See, for example, PCT application number PCT / US93 / 00829, which describes the controlled release of porous polymer microparticles for delivery of pharmaceutical compositions. In certain embodiments, sustained release formulations may include a semipermeable polymer matrix in the form of a molded article (e.g., film or microcapsule). Sustained-release matrices may include polyesters, hydrogels, polylactides (U.S. Pat. No. 3,773,919 and EP 058,481), copolymers of L-glutamic acid and γ-ethyl-L-glutamic acid (Sidman et al., Biopolymers, 22: 547-556 (1983)), poly (2-hydroxyethyl-methacrylate) (Langer et al., J Biomed Mater Res, 15: 167-277 (1981) and Langer, Chem Tech, 12: 98-105 (1982)), ethylene vinyl acetate (Langer et al., supra) or poly-D (-) -3-hydroxybutyric acid (EP 133,988). In certain embodiments, sustained-release compositions may also include liposomes, which may be prepared by any of several methods known in the art. See, e.g., Eppstein et al., PNAS, 82:3688-3692 (1985); EP 036,676; EP 088,046 and EP 143,949.

[0490] The pharmaceutical composition for in vivo administration is usually sterile. In certain embodiments, this can be accomplished by filtration with a sterile filtration membrane. In certain embodiments, when the composition is freeze-dried, the method can be used for sterilization before or after freeze-drying and reconstruction. In certain embodiments, the composition for parenteral administration can be stored in a lyophilized form or in a solution form. In certain embodiments, parenteral compositions are usually placed in a container with a sterile access port, for example, an intravenous solution bag or a vial with a stopper that a hypodermic needle can pierce.

[0491] In certain embodiments, once the pharmaceutical composition is formulated, it can be stored in a sterile vial as a solution, suspension, gel, emulsion, solid, or dehydrated or lyophilized powder. In certain embodiments, such preparations can be stored in a ready-to-use form or in a form (e.g., lyophilized form) reconstituted before administration.

[0492] In certain embodiments, a kit for producing a single-dose administration unit is provided. In certain embodiments, the kit can include a first container with a dried protein and a second container with an aqueous formulation. In certain embodiments, a kit containing a single-chamber and multi-chamber prefilled syringe (e.g., a liquid syringe and a lyosyringe syringe) is included.

[0493] In certain embodiments, the effective amount of the pharmaceutical composition containing a soluble interferon receptor with or without at least one additional therapeutic agent for therapeutic use depends on, for example, the treatment background and the target. It will be appreciated by those skilled in the art that, according to certain embodiments, the appropriate dosage level for treatment is therefore partially based on the delivered molecule, the indication used for the soluble interferon receptor with or without at least one additional therapeutic agent, the route of administration, and the patient's stature (weight, body surface area or organ size) and / or condition (age and overall health) and changes. In certain embodiments, clinicians can titrate the dose and change the route of administration to obtain the best therapeutic effect. In certain embodiments, depending on the above factors, the range of a typical dose can be from about 0.1 μg / kg to up to about 100 mg / kg or higher. In certain embodiments, the dosage can be from 0.1 μg / kg to a maximum of about 100 mg / kg; or 1 μg / kg to a maximum of about 100 mg / kg; or 5 μg / kg to a maximum of about 100 mg / kg.

[0494] In certain embodiments, the frequency of administration will take into account the pharmacokinetic parameters of the soluble interferon receptor in the preparation used and / or any other therapeutic agent. In certain embodiments, the clinician applies the composition until the dosage for achieving the desired effect is reached. Therefore, in certain embodiments, the composition can be used in a single dose, or over time with two or more dosages (which may or may not include the desired molecule of the same amount) or by the continuous infusion of an implant or cannula. The further optimization of suitable doses is routinely performed by those of ordinary skill in the art, and within the scope of the tasks they routinely perform. In certain embodiments, suitable dosage can be determined by using appropriate dose-response data.

[0495] In certain embodiments, the route of administration of the pharmaceutical composition is according to known methods, such as oral administration; injection by intravenous, intraperitoneal, intracerebral (intraparenchymal), intraventricular, intramuscular, subcutaneous, intraocular, intraarterial, intraportal or intralesional routes; by sustained release system; or by implant device. In certain embodiments, the composition can be administered by bolus injection or continuously by infusion or by implant device.

[0496] In certain embodiments, the composition can be applied topically by implanting a membrane, sponge, or another suitable material into which the desired molecule has been absorbed or encapsulated. In certain embodiments, where an implant device is used, the device can be implanted in any suitable tissue or organ, and the delivery of the desired molecule can be carried out by diffusion, timed release bolus, or continuous administration.

[0497] In certain embodiments, it may be desirable to use a pharmaceutical composition comprising a soluble interferon receptor (with or without at least one additional therapeutic agent) in an ex vivo manner. In these cases, cells, tissues and / or organs removed from a patient are exposed to a pharmaceutical composition containing a soluble interferon receptor with or without at least one additional therapeutic agent, after which the cells, tissues and / or organs are implanted back into the patient.

[0498] In certain embodiments, soluble interferon receptor and / or any other therapeutic agent can be delivered by implanting certain cells that have been genetically engineered to express and secrete polypeptides using those methods as described herein. In certain embodiments, such cells can be animal or human cells, and can be autologous, heterologous or xenogeneic. In certain embodiments, cells can be immortalized. In certain embodiments, in order to reduce the possibility of immune response, cells can be encapsulated to avoid the infiltration of surrounding tissues. In certain embodiments, encapsulating material is typically biocompatible, semipermeable polymer shell or membrane, which allows the release of protein products, but prevents cells from being destroyed by the patient's immune system or other harmful factors from surrounding tissues.

[0499] In vitro analysis

[0500] Various in vitro assays known in the art can be used to assess the efficacy of the soluble interferon receptors of the invention.

[0501] For example, the ability of soluble interferon receptors to inhibit IFN-α and / or IFN-β-induced secreted alkaline phosphatase (SEAP) production in HEK-Blue α / β cells can be assessed. HEK-Blue IFN-α / β cells (Invivogen, catalog number hkb-ifnab) produce and secrete SEAP in response to stimulation with IFN-α or IFN-β.

[0502] To evaluate the ability of soluble interferon receptors to inhibit IFN-α and / or IFN-β activity, IFN-α and / or IFN-β can be added to a tissue culture plate along with inhibitors (e.g., soluble interferon receptors (e.g., RSLV-601-604, RSLV-602-603) and control molecules (e.g., anti-IFN-α, anti-IFN-β, and human IgG). HEK-Blue IFN-α / β cells can then be added to the plate and incubated at 37° C. for a predetermined time. To evaluate SEAP activity, cell supernatants can then be added to QUANTI-Blue reagent and incubated at 37° C. for a predetermined time. SEAP activity can be detected by measuring absorbance at 620 nm.

[0503] The effectiveness of soluble interferon receptors is demonstrated by comparing the assay results of cells treated with soluble interferon receptors disclosed herein with the assay results of cells treated with controls. After treatment with effective soluble interferon receptors, the levels of IFN-α and / or IFN-β are generally comparable to the levels measured after anti-IFN-α or anti-IFN-β control treatment. After treatment with effective soluble interferon receptors, the levels of IFN-α and / or IFN-β are generally reduced relative to the levels measured after treatment with negative controls (e.g., human IgG).

[0504] Treatment

[0505] The soluble interferon receptors disclosed herein are particularly effective for the treatment of autoimmune disorders or abnormal immune responses. In this regard, it should be understood that the soluble interferon receptors disclosed herein can be used to control, inhibit, regulate, treat or eliminate immune response disorders caused by excessive production of interferon.

[0506] On the other hand, soluble interferon receptors are adapted to prevent (preventative) or treat (therapeutic) diseases or disorders, such as autoimmune diseases, in mammals by administering soluble interferon receptors to mammals in need in a therapeutically effective amount or sufficient amount, wherein the disease is prevented or treated. The present invention contemplates any route of administration (e.g., intravenous, intramuscular, subcutaneous) suitable for achieving the desired effect. Treatment of a disease condition may result in alleviation of symptoms associated with the condition, which may be long-term or short-term, or even transient beneficial effects.

[0507] Many disease conditions are amenable to treatment using the soluble interferon receptors of the present disclosure.For example, in some aspects, the disease or disorder is an autoimmune disease or cancer. In some such aspects, the autoimmune disease is insulin-dependent diabetes mellitus, multiple sclerosis, experimental autoimmune encephalomyelitis, rheumatoid arthritis, experimental autoimmune arthritis, myasthenia gravis, thyroiditis, experimental uveoretinitis, Hashimoto's thyroiditis, primary mucosal edema, thyrotoxicosis, pernicious anemia, autoimmune atrophic gastritis, Addison's disease, premature menopause, male infertility, juvenile diabetes, Goodpasture's syndrome, pemphigus vulgaris, pemphigus, sympathetic ophthalmia, lenticular uveitis, autoimmune hemolytic anemia, idiopathic leukopenia, primary biliary cirrhosis, chronic active hepatitis HBs-ve, cryptogenic cirrhosis, ulcerative colitis, Sjögren's syndrome, scleroderma, Wegener's granulomatosis, polymyositis, dermatomyositis, discoid LE, SLE, or connective tissue disease.

[0508] In a specific embodiment, soluble interferon receptors are used to prevent or treat SLE or Sjogren's syndrome. The effectiveness of soluble interferon receptors is demonstrated by comparing the expression levels of certain known IFN-regulated genes in mammals treated with soluble interferon receptors disclosed herein and in mammals treated with control preparations. In some embodiments, the expression levels of one, two, three, four, five or more IFN-regulated genes are measured. For example, in some embodiments, the expression levels of three or more IFN-regulated genes (e.g., HERC5, EPSTI, CMPK2) are measured. In some embodiments, IFN-regulated genes include those described in Lupus Science and Medicine, 2015, such as Bennett et al., J. Exp. Med., Vol. 197, No. 6, 711-723, March 2003 and Kennedy et al., 2015; 2: e00080. Doi: 10.1136 / lupus-2014-000080, both of which are incorporated herein by reference.

[0509] For exa...

Claims

1. A heterodimer that binds type I interferons, interferon-α (INFα) and interferon-β (INFβ), wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an interferon receptor 1 (IFNAR1) domain that is operably coupled to a mutant Fc domain with a joint domain, and wherein the second polypeptide comprises an interferon receptor 2 (IFNAR2) domain that is operably coupled to a mutant Fc domain with a joint domain; and wherein: (i) the first polypeptide consists of the amino acid sequence of SEQ ID NO: 40 and the second polypeptide consists of the amino acid sequence of SEQ ID NO: 43; (ii) the first polypeptide consists of the amino acid sequence of SEQ ID NO: 41 and the second polypeptide consists of the amino acid sequence of SEQ ID NO: 42; or (iii) the first polypeptide consists of the amino acid sequence of SEQ ID NO: 53 and the second polypeptide consists of the amino acid sequence of SEQ ID NO:

55.

2. A heterodimer that binds type I interferon interferon-α (INFα) and interferon-β (INFβ), wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an interferon receptor 1 (IFNAR1) domain that is operably coupled to a mutant Fc domain with a joint domain, and wherein the second polypeptide comprises an interferon receptor 2 (IFNAR2) domain that is operably coupled to a mutant Fc domain with a joint domain, wherein the first polypeptide consists of the amino acid sequence of SEQ ID NO: 40 and the second polypeptide consists of the amino acid sequence of SEQ ID NO:

43.

3. A heterodimer that combines type I interferon interferon-α (INFα) and interferon-β (INFβ), wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein the first polypeptide is included in an interferon receptor 1 (IFNAR1) domain that is operably coupled to a mutant Fc domain with a joint domain, and wherein the second polypeptide is included in an interferon receptor 2 (IFNAR2) domain that is operably coupled to a mutant Fc domain with a joint domain, wherein the first polypeptide consists of the amino acid sequence of SEQ ID NO: 41 and the second polypeptide consists of the amino acid sequence of SEQ ID NO:

42.

4. A heterodimer that combines type I interferon interferon-α (INFα) and interferon-β (INFβ), wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein the first polypeptide is included in an interferon receptor 1 (IFNAR1) domain that is operably coupled to a mutant Fc domain with a joint domain, and wherein the second polypeptide is included in an interferon receptor 2 (IFNAR2) domain that is operably coupled to a mutant Fc domain with a joint domain, wherein the first polypeptide consists of the amino acid sequence of SEQ ID NO: 53 and the second polypeptide consists of the amino acid sequence of SEQ ID NO:

55.

5. A heterodimer that binds type I interferon interferon-α (INFα) and interferon-β (INFβ), wherein the heterodimer comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises an interferon receptor 1 (IFNAR1) domain that is operably coupled to a mutant Fc domain without a joint domain, and wherein the second polypeptide comprises an interferon receptor 2 (IFNAR2) domain that is operably coupled to a mutant Fc domain without a joint domain, wherein the first polypeptide consists of the amino acid sequence of SEQ ID NO: 57 and the second polypeptide consists of the amino acid sequence of SEQ ID NO:

59. The heterodimer according to any one of claims 1 to 5 , wherein the heterodimer inhibits the activity of INFα.

7. The heterodimer of any one of claims 1-5, wherein the heterodimer inhibits the activity of INFβ.

8. The heterodimer of any one of claims 1-5, wherein the heterodimer inhibits the activities of both INFα and INFβ.

9. A heterodimer as described in any one of claims 1-5, wherein the heterodimer inhibits the induction of type I interferon (IFN) gene expression.

10. A composition comprising the heterodimer according to any one of claims 1 to 9 and a pharmaceutically acceptable carrier.

11. A nucleic acid molecule comprising a nucleotide sequence encoding the heterodimer according to any one of claims 1 to 9. A recombinant expression vector comprising the nucleic acid molecule according to claim 11 .

13. A host cell transformed with the recombinant expression vector according to claim 12.

14. Use of the heterodimer of any one of claims 1 to 9 or the composition of claim 10 in the preparation of a medicament for treating SLE in a subject.

15. Use of the heterodimer of any one of claims 1-9 and an optional pharmaceutically acceptable carrier in the preparation of a medicament for treating SLE in a subject in need thereof or delaying its progression, wherein the treatment comprises administering the medicament to a subject in need thereof.

16. A kit comprising a medicament comprising the heterodimer of any one of claims 1-9 and optionally a pharmaceutically acceptable carrier, and a package insert comprising instructions for administering the medicament to treat SLE or delay the progression thereof in a subject in need thereof.

17. A kit comprising a container comprising the heterodimer of any one of claims 1-9 and optionally a pharmaceutically acceptable carrier, and a package insert comprising instructions for administering the heterodimer to treat SLE or delay the progression thereof in a subject in need thereof.

18. A method for producing the heterodimer of any one of claims 1-9, comprising: Providing a host cell comprising a nucleic acid sequence encoding the heterodimer; and maintaining the host cell under conditions whereby the heterodimer is expressed.

19. The method of claim 18, further comprising obtaining the heterodimer.

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