Fusion protein constructs for complement-related disorders

Abstract

The present invention provides a fusion protein construct that can bind to complement-related antigens, comprises a target-directing moiety, and a complement regulator protein or a fragment thereof or a variant thereof. [Solution] A monomeric fusion protein construct that binds to a complement-related antigen is provided, comprising a first polypeptide oriented AB from the N-terminus to the C-terminus, and a second polypeptide arranged EF from the N-terminus to the C-terminus, wherein at least one of A, B, E, or F may be conjugated to R, A may contain VH or an antigen-binding fragment thereof, B may contain a CH1 amino acid sequence, R may contain a complement regulator polypeptide, E may contain VL or an antigen-binding fragment thereof, and F may contain CL1, wherein B of the first polypeptide and F of the second polypeptide are linked by one or more disulfide bonds.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the benefits of U.S. Provisional Application No. 62 / 778,014, filed on 11 December 2018, which is incorporated herein by reference in its entirety.

[0002] Sequence List This application includes an electronically submitted sequence listing in ASCII format, which is incorporated herein by reference in its entirety. The above ASCII copy was created on December 11, 2019, named 53542_707_601_SL.txt, and has a size of 950,242 bytes. [Overview of the project]

[0003] In one embodiment, this specification provides a monomeric fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide containing domains A and B aligned in an AB orientation from the N-terminus to the C-terminus, and a second polypeptide containing domains E and F aligned in an EF orientation from the N-terminus to the C-terminus, wherein at least one of domains A, B, E, or F may be conjugated to domain R, and domain A may contain a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B may contain a heavy chain CH1 constant region amino acid sequence, domain R may contain a complement regulator polypeptide, domain E may contain a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F may contain a light chain constant region amino acid sequence (CL1), wherein domain B of the first polypeptide and domain F of the second polypeptide are linked by one or more disulfide bonds.

[0004] In some embodiments, the first polypeptide comprises domains A, B, and R, the domains of the first polypeptide may be aligned in an RAB orientation from the N-terminus to the C-terminus, and domain R and domain A may be conjugated. In some embodiments, the first polypeptide comprises domains A, B, and R, the domains of the first polypeptide may be aligned in an ABR orientation from the N-terminus to the C-terminus, and domain B and domain R may be conjugated. In some embodiments, the second polypeptide comprises domains E, F, and R, the domains of the second polypeptide may be aligned in an REF orientation from the N-terminus to the C-terminus, and domain E and domain R may be conjugated. In some embodiments, the second polypeptide comprises domains E, F, and R, the domains of the second polypeptide may be aligned in an EFR orientation from the N-terminus to the C-terminus, and domain F and domain R may be conjugated. In some embodiments, the monomeric fusion protein construct further comprises a second complement regulatory polypeptide, the second complement regulatory polypeptide and domain R may be the same or different.

[0005] One embodiment is a fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, domain R, hinge region, domain C, and domain D, and the domains A, B, hinge region, C, and D of the first polypeptide are aligned from the N-terminus to the C-terminus in an AB-hinge-region-CD orientation, and domain A may contain a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B may contain a heavy chain CH1 constant region amino acid sequence, and domain C may contain a heavy chain CH2 constant region amino acid sequence The present invention provides a fusion protein construct wherein domain D may contain a heavy chain CH3 constant region amino acid sequence, domain R may contain a complement regulator polypeptide, (1) domain A and domain R may be conjugated, or (2) domain D and domain R may be conjugated, the second polypeptide may contain domains E and F, domains E and F of the second polypeptide are aligned EF-oriented from the N-terminus to the C-terminus, (i) domain E may contain a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) domain F may contain a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide are linked by one or more disulfide bonds.

[0006] In some embodiments, the fusion protein construct may be a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain D and domain R may be conjugated in at least one of the primary polypeptides, and the two primary polypeptides are linked by one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct may further comprise domain R1, which may comprise a secondary complement regulatory polypeptide, and domain R1 and domain R may be the same or different.

[0007] In some embodiments, domain A and domain R may be conjugated, and domain D and domain R1 may be conjugated. In some embodiments, domain D and domain R may be conjugated, and domain A and domain R1 may be conjugated. In some embodiments, the fusion protein construct may be a tetravalent heterodimer fusion protein construct comprising c)(i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide are aligned AB-hinge region-CD from the N-terminus to the C-terminus, and d) a fourth polypeptide comprising domains E and F, wherein domains E and F of the fourth polypeptide are aligned EF from the N-terminus to the C-terminus, and domain B of the third polypeptide and domain F of the fourth polypeptide are linked by one or more disulfide bonds, and the first and third polypeptides are linked by one or more disulfide bonds in the hinge region. In some embodiments, domain D and domain R may be conjugated in the first polypeptide. In some embodiments, domain A and domain R may be conjugated in the first polypeptide.

[0008] In some embodiments, the fusion protein construct may further include domain R1, which includes a second complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain R1 may be conjugated to a first polypeptide. In some embodiments, domain R1 may be conjugated to a second polypeptide. In some embodiments, domain R1 is conjugated to a third polypeptide. In some embodiments, domain R1 is conjugated to a fourth polypeptide.

[0009] Another embodiment is a fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, domain R, a hinge region and domain C, and domain A of the first polypeptide The present invention provides a fusion protein construct wherein B, the hinge region and C may be aligned in an AB-hinge region-C orientation from the N-terminus to the C-terminus, domain A contains a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B contains a heavy chain CH1 constant region amino acid sequence, domain C contains a heavy chain CH2 constant region amino acid sequence, domain R contains a complement regulator polypeptide, (1) domain A and domain R may be conjugated or (2) domain C and domain R may be conjugated, the second polypeptide contains domains E and F, domains E and F may be aligned in an EF orientation from the N-terminus to the C-terminus, (i) domain E contains a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) domain F contains a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0010] In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain C and domain R may be conjugated in at least one of the primary polypeptides, and the two primary polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain A and domain R may be conjugated in at least one of the primary polypeptides, and the two primary polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct further comprises domain R1, which may comprise a secondary complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain A and domain R may be conjugated, and domain C and domain R1 may be conjugated. In some embodiments, domain C and domain R may be conjugated, and domain A and domain R1 may be conjugated.

[0011] In some embodiments, the fusion protein construct is a tetravalent heterodimer fusion protein construct comprising c)(i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide may be aligned AB-hinge region-CD from the N-terminus to the C-terminus, and d) a fourth polypeptide comprising domains E and F, wherein domains E and F of the fourth polypeptide may be aligned EF from the N-terminus to the C-terminus, and domain B of the third polypeptide and domain F of the fourth polypeptide may be linked by one or more disulfide bonds, and the first and third polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0012] In some embodiments, domain C and domain R may be conjugated in the first polypeptide. In some embodiments, domain A and domain R may be conjugated in the first polypeptide. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a second complement regulatory polypeptide, and domains R1 and R may be the same or different. In some embodiments, domain R1 is conjugated to the first polypeptide. In some embodiments, domain R1 is conjugated to the second polypeptide. In some embodiments, domain R1 is conjugated to the third polypeptide. In some embodiments, domain R1 is conjugated to the fourth polypeptide.

[0013] Another embodiment is a fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, domain R and a hinge region, and the domains A, B and the hinge region of the first polypeptide may be aligned from the N-terminus to the C-terminus in an AB-hinged orientation, domain A may comprise a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B may comprise a heavy chain CH1 constant region amino acid sequence, and domain R may comprise a complement regulator polypeptide, (1) domain A The invention provides a fusion protein in which (1) the domain R and domain B may be conjoined, or (2) the hinge region and domain R may be conjoined, the second polypeptide comprises domain E and domain F, the domains E and F may be aligned from the N-terminus to the C-terminus in the orientation of EF, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, domain F comprises a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0014] In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two first polypeptides and two second polypeptides, wherein domain A and domain R may be conjugated in at least one of the first polypeptides, and the two first polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two first polypeptides and two second polypeptides, wherein the hinge region and domain R may be conjugated in at least one of the first polypeptides, and the two first polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a second complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain A and domain R may be conjugated, and the hinge domain and domain R1 may be conjugated. In some embodiments, the hinge domain and domain R may be conjugated, and domain A and domain R1 may be conjugated. In some embodiments, the fusion protein construct is a tetravalent heterodimer fusion protein construct comprising c) (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide are aligned AB-hinge region-CD from the N-terminus to the C-terminus, and d) a fourth polypeptide comprising domains E and F, wherein domains E and F of the fourth polypeptide are aligned EF from the N-terminus to the C-terminus, domain B of the third polypeptide and domain F of the fourth polypeptide may be linked by one or more disulfide bonds, and the first and third polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0015] In some embodiments, the hinge domain and domain R may be conjugated in the first polypeptide. In some embodiments, domain A and domain R may be conjugated in the first polypeptide. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a second complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain R1 is conjugated to the first polypeptide. In some embodiments, domain R1 is conjugated to the second polypeptide. In some embodiments, domain R1 is conjugated to the third polypeptide. In some embodiments, domain R1 is conjugated to the fourth polypeptide.

[0016] Another embodiment is a trivalent heterodimeric fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, domain C, domain D, and domain R, and the domains A, B, the hinge region, C, and D of the first polypeptide may be aligned in an AB-hinge-CD orientation from the N-terminus to the C-terminus, and (1) domain A and domain R may be conjugated or (2) domain D and Domain R may be conjugated, the second polypeptide may contain domains E and F, and domains E and F of the second polypeptide may be aligned EF-oriented from the N-terminus to the C-terminus, the third polypeptide may contain (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, and domains A, B, hinge region, C and D of the third polypeptide may be aligned EF-oriented from the N-terminus to the C-terminus The AB-hinge region-CD orientation may be arranged, the 4th polypeptide may contain domains E and F, the 4th polypeptide may be arranged EF orientation from N-terminus to C-terminus, the domain B of the 1st polypeptide and the domain F of the 2nd polypeptide may be linked by one or more disulfide bonds, the domain B of the 3rd polypeptide and the domain F of the 4th polypeptide may be linked by one or more disulfide bonds, the 1st and 3rd polypeptides may be linked in the hinge region by one or more disulfide bonds, domain A contains a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B contains a heavy chain CH1 constant region amino acid sequence, domain R contains a complement regulator polypeptide, domain C contains a heavy chain CH2 constant region amino acid sequence, domain D contains a heavy chain CH3 constant region amino acid sequence, domain E contains a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F isThe present invention provides a trivalent heterodimer fusion protein construct containing the light chain constant region amino acid sequence (CL1).

[0017] In some embodiments, domain A and domain R of the first polypeptide may be conjugated. In some embodiments, domain D and domain R of the first polypeptide may be conjugated.

[0018] Another embodiment is a trivalent heterodimeric fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, domain C, and domain R, and the domains A, B, the hinge region, and C of the first polypeptide may be aligned AB-hinge-region-C from the N-terminus to the C-terminus, (1) domain A and domain R may be conjugated, or (2) domain C and domain R may be conjugated, the second polypeptide comprises domain E and domain F, and the domains E and F of the second polypeptide may be aligned EF from the N-terminus to the C-terminus, and the third polypeptide comprises (i) domain A, domain B, hinge region (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide may be aligned AB-hinge region-CD from the N-terminus to the C-terminus, and the fourth polypeptide may contain domains E and F, wherein domains E and F of the fourth polypeptide may be aligned EF from the N-terminus to the C-terminus, and domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds, and domain B of the third polypeptide and domain F of the fourth polypeptide may be linked by one or more disulfide bonds, and the first and third polypeptides have a hinge region The present invention provides a trivalent heterodimer fusion protein construct in which the domains are linked by one or more disulfide bonds, and in which domain A contains a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B contains a heavy chain CH1 constant region amino acid sequence, domain R contains a complement regulator polypeptide, domain C contains a heavy chain CH2 constant region amino acid sequence, domain E contains a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F contains a light chain constant region amino acid sequence (CL1).

[0019] In some embodiments, domain A and domain R may be conjugated in the first polypeptide. In some embodiments, domain C and domain R may be conjugated in the first polypeptide.

[0020] Another embodiment is a trivalent heterodimeric fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, and domain R, and the domains A, B, and hinge region of the first polypeptide may be aligned from the N-terminus to the C-terminus in an AB-hinged orientation, and (1) domain A and domain R may be conjugated or (2) the hinge region and domain R may be conjugated The second polypeptide may be modified, and the second polypeptide may contain domains E and F, and the domains E and F of the second polypeptide may be aligned EF-oriented from the N-terminus to the C-terminus, and the third polypeptide may contain (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, and the domains A, B, hinge region, C and D of the third polypeptide may be aligned A from the N-terminus to the C-terminus The present invention provides a trivalent heterodimer fusion protein construct wherein the B-hinge region-CD orientation may be arranged, the fourth polypeptide comprises domains E and F, the domains E and F of the fourth polypeptide are arranged EF orientation from the N-terminus to the C-terminus, the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds, the domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, the first and third polypeptides are linked by one or more disulfide bonds in the hinge region, domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprises a heavy chain CH1 constant region amino acid sequence, domain R comprises a complement regulator polypeptide, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F comprises a light chain constant region amino acid sequence (CL1).

[0021] In some embodiments, domain A and domain R may be conjugated in the first polypeptide. In some embodiments, the hinge region and domain R may be conjugated in the first polypeptide.

[0022] Another embodiment is a complement-associated antigen-binding fusion protein construct comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, hinge region, domain C and domain D, and domains A, B, hinge region, C and D may be aligned from the N-terminus to the C-terminus in an AB-hinge-region-CD orientation, domain A comprising a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprising a heavy chain CH1 constant region amino acid sequence, domain C comprising a heavy chain CH2 constant region amino acid sequence, domain D comprising a heavy chain CH3 constant region amino acid sequence, and the second polypeptide comprising domains E, F and domain R, wherein domains E and F are aligned from the N-terminus to the C-terminus The present invention provides a fusion protein construct in which the EFs may be aligned in an oriented manner, (1) domain E and domain R may be conjugated, or (2) domain F and domain R may be conjugated, domain E contains a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, domain F contains a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0023] In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain E and domain R may be conjugated in at least one of the secondary polypeptides, and the two primary polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain F and domain R may be conjugated in at least one of the secondary polypeptides, and the two primary polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a secondary complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain E and domain R may be conjugated, and domain F and domain R1 may be conjugated. In some embodiments, domain F and domain R may be conjugated, and domain E and domain R1 may be conjugated. In some embodiments, the fusion protein construct is a tetravalent heterodimer fusion protein construct comprising c) (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide are aligned AB-hinge region-CD from the N-terminus to the C-terminus, and d) a fourth polypeptide comprising domains E and domain F, wherein domains E and F of the fourth polypeptide are aligned EF from the N-terminus to the C-terminus, domain B of the third polypeptide and domain F of the fourth polypeptide may be linked by one or more disulfide bonds, and the first and third polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0024] In some embodiments, domain E and domain R may be conjugated in the first polypeptide. In some embodiments, domain F and domain R may be conjugated in the first polypeptide. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a second complement regulatory polypeptide, and domains R1 and R may be the same or different. In some embodiments, domain R1 is conjugated to the first polypeptide. In some embodiments, domain R1 is conjugated to the second polypeptide. In some embodiments, domain R1 is conjugated to the third polypeptide. In some embodiments, domain R1 is conjugated to the fourth polypeptide.

[0025] Another embodiment is a fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region and domain C, and domains A, B, the hinge region and C may be aligned in an AB-hinge-region-C orientation from the N-terminus to the C-terminus, domain A comprising a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprising a heavy chain CH1 constant region amino acid sequence, domain C comprising a heavy chain CH2 constant region amino acid sequence, and the second polypeptide comprising domain E, domain The present invention provides a fusion protein construct comprising domains F and R, wherein domains E and F may be aligned from the N-terminus to the C-terminus in an EF orientation, (1) domain E and domain R may be conjugated, or (2) domain F and domain R may be conjugated, wherein domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F comprises a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0026] In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain E and domain R may be conjugated in at least one of the secondary polypeptides, and the two primary polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain F and domain R may be conjugated in at least one of the secondary polypeptides, and the two primary polypeptides may be linked together via one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a secondary complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain E and domain R may be conjugated, and domain F and domain R1 may be conjugated. In some embodiments, domain F and domain R of the fusion protein construct may be conjugated, and domain E and domain R1 may be conjugated. In some embodiments, the fusion protein construct is a tetravalent heterodimer fusion protein construct comprising c)(i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide may be aligned AB-hinge region-CD from the N-terminus to the C-terminus, and d) a fourth polypeptide comprising domains E and domain F, wherein domains E and F of the fourth polypeptide may be aligned EF from the N-terminus to the C-terminus, domain B of the third polypeptide and domain F of the fourth polypeptide may be linked by one or more disulfide bonds, and the first and third polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0027] In some embodiments, domain E and domain R may be conjugated in the first polypeptide. In some embodiments, domain F and domain R may be conjugated in the first polypeptide. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a second complement regulatory polypeptide, and domains R1 and R may be the same or different. In some embodiments, domain R1 is conjugated in the first polypeptide. In some embodiments, domain R1 is conjugated in the second polypeptide. In some embodiments, domain R1 is conjugated in the third polypeptide. In some embodiments, domain R1 is conjugated in the fourth polypeptide.

[0028] Another embodiment is a fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B and a hinge region, and domains A, B and the hinge region may be aligned from the N-terminus to the C-terminus in an AB-hinged orientation, domain A comprising a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprising a heavy chain CH1 constant region amino acid sequence, and domain C comprising a heavy chain CH2 constant region amino acid sequence. The present invention provides a fusion protein construct comprising a no-acid sequence, wherein the second polypeptide comprises domains E, F, and R, and domains E and F may be aligned from the N-terminus to the C-terminus in an E-F orientation, (1) domain E and domain R may be conjugated, or (2) domain F and domain R may be conjugated, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, domain F comprises a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0029] In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain E and domain R may be conjugated in at least one of the secondary polypeptides, and the two primary polypeptides may be linked by one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct is a tetravalent homodimer fusion protein construct comprising two primary polypeptides and two secondary polypeptides, wherein domain F and domain R may be conjugated in at least one of the secondary polypeptides, and the two primary polypeptides may be linked by one or more disulfide bonds in a hinge region. In some embodiments, the fusion protein construct further comprises domain R1, which comprises a secondary complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain E and domain R may be conjugated, and domain F and domain R1 may be conjugated. In some embodiments, domain F and domain R may be conjugated, and domain E and domain R1 may be conjugated.

[0030] In some embodiments, the fusion protein construct is a tetravalent heterodimer fusion protein construct comprising c) (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region, wherein domains A, B, hinge region, C and D of the third polypeptide are aligned AB-hinge region-CD from the N-terminus to the C-terminus, and d) a fourth polypeptide comprising domains E and F, wherein domains E and F of the fourth polypeptide are aligned EF from the N-terminus to the C-terminus, domain B of the third polypeptide and domain F of the fourth polypeptide are linked by one or more disulfide bonds, and the first and third polypeptides are linked by one or more disulfide bonds in the hinge region.

[0031] In some embodiments, domain E and domain R may be conjugated in the first polypeptide. In some embodiments, domain F and domain R may be conjugated in the first polypeptide.

[0032] In some embodiments, the fusion protein construct further comprises domain R1, which comprises a second complement regulatory polypeptide, and domain R1 and domain R may be the same or different. In some embodiments, domain R1 is conjugated to a first polypeptide. In some embodiments, domain R1 is conjugated to a second polypeptide. In some embodiments, domain R1 is conjugated to a third polypeptide. In some embodiments, domain R1 is conjugated to a fourth polypeptide.

[0033] Another embodiment is a trivalent heterodimeric fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide. The first polypeptide comprises domain A, domain B, hinge region, domain C, and domain D, and the domains A, B, hinge region, C, and D of the first polypeptide may be aligned from the N-terminus to the C-terminus in an AB-hinge region-CD orientation, and the second polypeptide comprises domain E, domain F, and domain R, and the domains E and F are aligned from the N-terminus to the C-terminus in an EF orientation, and (1) domain E and domain R may be conjugated or (2) domain F and domain R The third polypeptide may be conjugated, and the third polypeptide may consist of (i) domain A, domain B, domain C, domain D and a hinge region, (ii) domain A, domain B, domain C and a hinge region, or (iii) domain A, domain B and a hinge region, wherein the domains A, B, hinge region, C and D of the third polypeptide may be aligned from the N-terminus to the C-terminus in the orientation AB-hinge region-CD, and the fourth polypeptide may consist of domain E and domain F, and the domain E and The present invention provides a trivalent heterodimer fusion protein construct wherein domains F and B may be aligned from the N-terminus to the C-terminus in an E-F orientation, domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds, domain B of the third polypeptide and domain F of the fourth polypeptide may be linked by one or more disulfide bonds, and the first and third polypeptides may be linked together in a hinge region by one or more disulfide bonds, and domain A contains a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B contains a heavy chain CH1 constant region amino acid sequence, domain C contains a heavy chain CH2 constant region amino acid sequence, domain D contains a heavy chain CH3 constant region amino acid sequence, domain R contains a complement regulator polypeptide, domain E contains a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F contains a light chain constant region amino acid sequence (CL1).

[0034] In some embodiments, domains E and R of the second polypeptide may be conjugated. In some embodiments, domains F and R of the second polypeptide may be conjugated.

[0035] Another embodiment is a trivalent heterodimeric fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, and domain C, and the domains A, B, the hinge region, and C of the first polypeptide may be aligned AB-hinge-region-C from the N-terminus to the C-terminus; the second polypeptide comprises domain E, domain F, and domain R, and the domains E and F may be aligned EF from the N-terminus to the C-terminus, and (1) domain E and domain R may be conjugated, or (2) domain F and domain R may be conjugated; and the third polypeptide comprises (i) domain A, domain B, domain C, domain D, and a hinge region, (ii) domain A, domain B, domain C, and a hinge region, or (iii) domain A, domain B, and a hinge region The third polypeptide may include a region, and the domains A, B, hinge region, C, and D of the third polypeptide may be aligned in an AB-hinge region-CD orientation from the N-terminus to the C-terminus; the fourth polypeptide may include domains E and F, and the domains E and F of the fourth polypeptide may be aligned in an EF orientation from the N-terminus to the C-terminus; the domain B of the first polypeptide and the domain F of the second polypeptide may be linked by one or more disulfide bonds; the domain B of the third polypeptide and the domain F of the fourth polypeptide may be linked by one or more disulfide bonds; the first and third polypeptides may be linked by one or more disulfide bonds in the hinge region; domain A includes a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof; domain B includes a heavy chain CH1 constant region amino acid sequence; and domain C includes a heavy chain CH2 constant region amino acid sequence The present invention provides a trivalent heterodimer fusion protein construct comprising, wherein domain R comprises a complement regulator polypeptide, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F comprises a light chain constant region amino acid sequence (CL1).

[0036] In some embodiments, domains E and R of the second polypeptide may be conjugated. In some embodiments, domains F and R of the second polypeptide may be conjugated.

[0037] Another embodiment is a trivalent heterodimeric fusion protein construct that binds to a complement-associated antigen, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, and a hinge region, and the domains A, B, and the hinge region of the first polypeptide may be aligned in an AB-hinge region orientation from the N-terminus to the C-terminus; the second polypeptide comprises domain E, domain F, and domain R, and the domains E and F may be aligned in an EF orientation from the N-terminus to the C-terminus, and (1) domain E and domain R may be conjugated, or (2) domain F and domain R may be conjugated; the third polypeptide comprises (i) domain A, domain B, domain C, domain D, and a hinge region, (ii) domain A, domain B, domain C, and a hinge region, or (iii) domain A, domain B, and a hinge region, and the domains A, B, the hinge region, C, and D of the third polypeptide may be aligned in an AB-hinge region orientation from the N-terminus to the C-terminus The present invention provides a trivalent heterodimer fusion protein construct wherein the domains may be aligned in a domain-CD orientation, the fourth polypeptide comprises domains E and F, the domains E and F of the fourth polypeptide may be aligned in an EF orientation from the N-terminus to the C-terminus, the domain B of the first polypeptide and the domain F of the second polypeptide may be linked by one or more disulfide bonds, the domain B of the third polypeptide and the domain F of the fourth polypeptide may be linked by one or more disulfide bonds, the first and third polypeptides may be linked together in a hinge region by one or more disulfide bonds, domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprises a heavy chain CH1 constant region amino acid sequence, domain R comprises a complement regulator polypeptide, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F comprises a light chain constant region amino acid sequence (CL1).

[0038] In some embodiments, domain E and domain R may be conjugated in the second polypeptide. In some embodiments, domain F and domain R may be conjugated in the second polypeptide.

[0039] Another embodiment is a fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide, a second polypeptide, and a third polypeptide, wherein the first polypeptide comprises domain R, a hinge region, domain C, and domain D, and domains R, the hinge region, C, and D may be aligned in an R-hinge-CD orientation from the N-terminus to the C-terminus, domain R comprises a complement regulator polypeptide, domain C comprises a heavy chain CH2 constant region amino acid sequence, domain D comprises a heavy chain CH3 constant region amino acid sequence, and domain R and the hinge domain may be conjugated, the second polypeptide comprises domain A, domain B, a hinge region, domain C, and domain D, and domains A, B, the hinge region, C, and D may be aligned in an AB-hinge-CD orientation from the N-terminus to the C-terminus, domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprises a heavy chain CH1 constant region amino acid sequence, and the third polypeptide comprises domains E and F The present invention provides a fusion protein construct comprising the following: the domains may be aligned in an E-F orientation from the N-terminus to the C-terminus; domain E may contain a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof; domain F may contain a light chain constant region amino acid sequence (CL1); domain B of the second polypeptide and domain F of the third polypeptide may be linked by one or more disulfide bonds; and the first and second polypeptides may be linked together in a hinge region by one or more disulfide bonds.

[0040] Another embodiment provides a monomeric fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide comprising domains A, B and a hinge region aligned in an AB-hinge orientation from the N-terminus to the C-terminus, and a second polypeptide comprising domains E and F aligned in an EF orientation from the N-terminus to the C-terminus, wherein at least one of domains A, the hinge domain, domain E or domain F is conjugated to domain R, and domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprises a heavy chain CH1 constant region amino acid sequence, domain R comprises a complement regulator polypeptide, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F comprises a light chain constant region amino acid sequence (CL1), wherein domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0041] In some embodiments, the first polypeptide comprises domain A, domain B, a hinge domain, and domain R, and the domains of the first polypeptide may be aligned in an RAB-hinge orientation from the N-terminus to the C-terminus, and domain R and domain A may be conjugated. In some embodiments, the first polypeptide comprises domain A, domain B, a hinge domain, and domain R, and the domains of the first polypeptide may be aligned in an AB-hinge-R orientation from the N-terminus to the C-terminus, and the hinge domain and domain R may be conjugated. In some embodiments, the second polypeptide comprises domain E, domain F, and domain R, and the domains of the second polypeptide may be aligned in an REF orientation from the N-terminus to the C-terminus, and domain E and domain R may be conjugated. In some embodiments, the second polypeptide comprises domain E, domain F, and domain R, and the domains of the second polypeptide may be aligned in an EFR orientation from the N-terminus to the C-terminus, and domain F and domain R may be conjugated. In some embodiments, the fusion protein construct further comprises a complement-2 regulatory polypeptide, the complement-2 regulatory polypeptide and domain R may be the same or different.

[0042] Another embodiment provides a monomeric fusion protein construct that binds to a complement-related antigen, comprising a first polypeptide comprising domain A, domain B, hinge region and domain C aligned in an AB-hinge-C region orientation from the N-terminus to the C-terminus, and a second polypeptide comprising domains E and F aligned in an EF orientation from the N-terminus to the C-terminus, wherein at least one of domains A, C, E or F is conjugated to domain R, and domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprises a heavy chain CH1 constant region amino acid sequence, domain C comprises a heavy chain CH2 constant region amino acid sequence, domain R comprises a complement regulator polypeptide, domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F comprises a light chain constant region amino acid sequence (CL1), wherein domain B of the first polypeptide and domain F of the second polypeptide may be linked by one or more disulfide bonds.

[0043] In some embodiments, the first polypeptide comprises domain A, domain B, hinge domain, domain C, and domain R, and the domains of the first polypeptide may be aligned in an RAB-hinge-C orientation from the N-terminus to the C-terminus, and domain R and domain A may be conjugated. In some embodiments, the first polypeptide comprises domain A, domain B, hinge domain, domain C, and domain R, and the domains of the first polypeptide may be aligned in an AB-hinge-CR orientation from the N-terminus to the C-terminus, and domain C and domain R may be conjugated. In some embodiments, the second polypeptide comprises domain E, domain F, and domain R, and the domains of the second polypeptide may be aligned in an REF orientation from the N-terminus to the C-terminus, and domain E and domain R may be conjugated. In some embodiments, the second polypeptide comprises domain E, domain F, and domain R, and the domains of the second polypeptide may be aligned in an EFR orientation from the N-terminus to the C-terminus, and domain F and domain R may be conjugated. In some embodiments, the fusion protein construct further comprises a complement-2 regulatory polypeptide, the complement-2 regulatory polypeptide and domain R may be the same or different.

[0044] Another embodiment is a monomeric fusion protein construct that binds to a complement-associated antigen, comprising: a) a first polypeptide comprising domain A, domain B, hinge region, domain C, and domain D aligned in an AB-hinge-CD region orientation from the N-terminus to the C-terminus; and b) a second polypeptide comprising domain E and domain F aligned in an EF orientation from the N-terminus to the C-terminus, wherein at least one of domains A, D, E, or F is conjugated to domain R, and (i) domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, and (ii) domain B comprises a heavy chain CH1 constant region The present invention provides a monomeric fusion protein construct comprising a regional amino acid sequence, wherein (iii) domain C comprises a heavy chain CH2 constant region amino acid sequence, (iv) domain D comprises a heavy chain CH3 constant region amino acid sequence, (v) domain R comprises a complement regulator polypeptide, (vi) domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and (vii) domain F comprises a light chain constant region amino acid sequence (CL1), wherein domain B of the first polypeptide and domain F of the second polypeptide may be linked via one or more disulfide bonds.

[0045] In some embodiments, the first polypeptide comprises domain A, domain B, hinge domain, domain C, domain D, and domain R, and the domains of the first polypeptide may be aligned in a RAB-hinge-CD orientation from the N-terminus to the C-terminus, and domain R and domain A may be conjugated. In some embodiments, the first polypeptide comprises domain A, domain B, hinge domain, domain C, domain D, and domain R, and the domains of the first polypeptide may be aligned in an AB-hinge-CDR orientation from the N-terminus to the C-terminus, and domain C and domain R may be conjugated. In some embodiments, the second polypeptide comprises domain E, domain F, and domain R, and the domains of the second polypeptide may be aligned in a REF orientation from the N-terminus to the C-terminus, and domain E and domain R may be conjugated. In some embodiments, the second polypeptide comprises domain E, domain F, and domain R, and the domains of the second polypeptide may be aligned in an EFR orientation from the N-terminus to the C-terminus, and domain F and domain R may be conjugated. In some embodiments, the fusion protein construct further comprises a complement-2 regulatory polypeptide, the complement-2 regulatory polypeptide and domain R may be the same or different.

[0046] In some embodiments, conjugation involves linking two domains with a peptide linker. This includes linking without a linker, enzymatic conjugation, chemical conjugation, or a combination thereof. In some embodiments, the complement-related antigen is C3d, iC3b, C3dg, or a fragment thereof, or a variant thereof. In some embodiments, the fusion protein construct binds to C3 and C3b with lower affinity than to C3d. In some embodiments, the fusion protein construct binds to C3 and C3b with approximately 10 -3 It binds with a KD affinity of M or higher. In some embodiments, the fusion protein construct binds to iC3b, C3dg, or both. -8It binds with a KD affinity of M or less. In some embodiments, the fusion protein construct modulates surrogate complement activity in a subject when administered to the subject or a pharmaceutical composition containing the fusion protein construct. In some embodiments, the fusion protein construct modulates classical complement activity in a subject when administered to the subject or a pharmaceutical composition containing the fusion protein construct. In some embodiments, the fusion protein construct modulates lectin complement activity in a subject when administered to the subject or a pharmaceutical composition containing the fusion protein construct. In some embodiments, the fusion protein construct binds to the domain of a mammalian annexin protein.

[0047] In some embodiments, the fusion protein construct has 10 units in the domain of mammalian annexin protein. -8 It binds with a KD affinity of M or less. In some embodiments, the domain is an annexin core domain. In some embodiments, the annexin core domain includes an alpha-helical domain. In some embodiments, the annexin core domain includes a calcium-binding site and a membrane-binding site. In some embodiments, the annexin core domain includes at least one annexin repeat. In some embodiments, the fusion protein construct binds to the annexin repeat sequence in the domain. In some embodiments, the fusion protein construct binds to a phospholipid. In some embodiments, the fusion protein construct binds to a phospholipid. -8Binding occurs with a KD affinity of M or less. In some embodiments, the phospholipid is selected from the group consisting of phosphatidylethanolamine (PE), cardiolipin (CL), phosphatidylcholine (PC), phosphatidylinositol, phosphatidylglycerol, phosphatidylserine, phosphatidic acid, and malondialdehyde (MDA). In some embodiments, the complement regulator peptide comprises domain A of the complement receptor 1 (CR1) protein, or a fragment thereof holding at least three short consensus repeats (SCRs) of domain A. In some embodiments, the fusion protein construct further comprises domain B of the CR1 protein, or a fragment thereof holding at least three SCRs of domain B.

[0048] In some embodiments, the fusion protein construct further comprises domain C of the CR1 protein, or a fragment thereof holding at least three SCRs of domain C. In some embodiments, the fusion protein construct further comprises domain D of CR1, or a fragment thereof holding at least three SCRs of domain D. In some embodiments, the complement regulator polypeptide comprises the first three SCRs of domain A, the first three SCRs of domain B, and the first three SCRs of domain C of the CR1 protein. In some embodiments, the complement regulator polypeptide is CR1(1-10). In some embodiments, the complement regulator polypeptide is CR1(1-17). In some embodiments, the complement regulator peptide comprises the amino acid sequence of SEQ ID NO: 41 or SEQ ID NO: 91, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator polypeptide comprises the amino acid sequence of SEQ ID NO: 42 or SEQ ID NO: 92, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is a complement enhancer (DAF) or a bioactive fragment thereof. In some embodiments, the DAF is a human DAF. In some embodiments, the bioactive fragment of human DAF is at least one of the short consensus repeat (SCR) domain and the O-glycosylated serine / threonine-rich domain of the full-length human DAF. Includes one.

[0049] In some embodiments, the bioactive fragment of DAF comprises SCR1-4 or SCR2-4 of full-length human DAF. In some embodiments, the bioactive fragment of DAF comprises the amino acid sequence of SEQ ID NO: 184, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is factor H or a bioactive fragment thereof. In some embodiments, factor H is human factor H. In some embodiments, the bioactive fragment of human factor H comprises one or more groups of short consensus repeats (SCRs) including SCR1-20, SCR1-2, SCR2-3, SCR3-4, SCR4-5, SCR5-6, SCR6-7, SCR7-8, SCR8-9, SCR9-10, SCR10-11, SCR11-12, SCR12-13, SCR13-14, SCR14-15, SCR15-16, SCR16-17, SCR17-18, SCR19-20, or any combination of SCR1-20 of full-length human factor H. In some embodiments, the bioactive fragment of human factor H comprises SCR1-4 or SCR1-5 of full-length human factor H. In some embodiments, the bioactive fragment of human factor H includes a sequence of amino acids selected from the group consisting of amino acids 21-266, 21-320, 21-509, or 19-1106 of SEQ ID NO: 9, or a variant thereof having an amino acid sequence that is at least 85% identical to said sequence of amino acids.

[0050] In some embodiments, factor H or its bioactive fragment comprises the amino acid sequence of SEQ ID NO: 72 or SEQ ID NO: 108, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is MCP or its bioactive fragment. In some embodiments, MCP is human MCP. In some embodiments, the bioactive fragment of human MCP comprises at least one short consensus repeat (SCR) domain of full-length human MCP. In some embodiments, the bioactive fragment of human MCP comprises SCR3-4 of full-length human MCP. In some embodiments, MCP comprises the amino acid sequence of SEQ ID NO: 187, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is MAP44 or its bioactive fragment. In some embodiments, MAP44 is human MAP44. In some embodiments, MAP44 comprises the amino acid sequence of SEQ ID NO: 186, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is CD59 or its bioactive fragment.

[0051] In some embodiments, CD59 is human CD59.

[0052] In some embodiments, CD59 includes the amino acid sequence of SEQ ID NO: 185, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the fusion protein construct includes a human antibody or an antigen-binding fragment thereof. In some embodiments, the fusion protein construct includes a humanized antibody or an antigen-binding fragment thereof. In some embodiments, the first and third polypeptides each include at least one orthogonal modification that is favorable to heterodimer formation compared to homodimer formation. In some embodiments, the first polypeptide includes a knob modification and the third polypeptide includes a hole modification, or the third polypeptide includes a knob modification and the first polypeptide includes a hole modification. In some embodiments, the first and third polypeptides include modifications that result in charge or surface complementarity.

[0053] In some embodiments, a) the first polypeptide is (i) SEQ ID NOs: 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 259 and 31; or 29, 260 and three heavy chain complementarity-determining regions (CDRs) having amino acid sequences 31, or (ii) SEQ ID NOs: 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 2 b) The second polypeptide comprises three heavy chain CDRs having amino acid sequences that differ in that one of 20, 221 and 222; 228, 229 and 230; 29, 259 and 31; or 29, 260 and 31 has a single conserved amino acid substitution, and b) the second polypeptide comprises (i) SEQ ID NOs: 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 32, 33 and 34; 38, 39 and (ii) Three light chain complementarity-determining regions (CDRs) having the amino acid sequences of (i) 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233, or (ii) SEQ ID NOs: 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; Includes three light chain CDRs having amino acid sequences that differ in that one of them contains a single conserved amino acid substitution: 32, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233.

[0054] In some embodiments, the second polypeptide (light chain containing domains E and F) comprises at least three CDRs, where the light chain CDRs are defined as CDR-L1, CDR-L2, and CDR-L3, respectively, and in SEQ ID NOs. 279, 68, 287, and 59, the CDRs comprise residues 27-37 (CDR-L1), 55-57 (CDR-L2), and 94-102 (CDR-L3), and in SEQ ID NO. 289, the CDRs comprise residues 27-38 (CDR-L1), 56-58 (CDR-L2), and 95-102 (CDR-L3). In some embodiments, the first polypeptide (a heavy chain including at least domains A and B) comprises at least three CDRs, where the heavy chain CDRs are defined as CDR-H1, CDR-H2, and CDR-H3, respectively, and in SEQ ID NOs. 280, 281, 282, 284, 285, 286, 73, and 288, the CDR comprises residues 26-33 (CDR-H1), 51-58 (CDR-H2), and 97-100 (CDR-H3); in SEQ ID NOs. 244, the CDR comprises residues 26-33 (CDR-H1), 51-58 (CDR-H2), and 97-102 (CDR-H3); and in SEQ ID NOs. 290, the CDR comprises residues 26-33 (CDR-H1), 51-58 (CDR-H2), and 97-110 (CDR-H3). In some embodiments, the first polypeptide (heavy chain including at least domains A and B) includes at least three CDRs, where the heavy chain CDRs are defined as CDR-H1, CDR-H2, and CDR-H3, respectively, and in SEQ ID NO: 342, it includes SEQ ID NO: 23, SEQ ID NO: 24, and SEQ ID NO: 25. In some embodiments, the second polypeptide (light chain including at least domains E and F) includes at least three CDRs, where the light chain CDRs are defined as CDR-L1, CDR-L2, and CDR-L3, respectively, and it includes SEQ ID NO: 26, SEQ ID NO: 27, and SEQ ID NO: 28.

[0055] In some embodiments, the fusion protein construct further comprises at least one amino acid linker, the at least one linker comprising any of the amino acid sequences of SEQ ID NO: 138, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169, SEQ ID NO: 170, SEQ ID NO: 171, SEQ ID NO: 172, SEQ ID NO: 173, SEQ ID NO: 174, SEQ ID NO: 175, SEQ ID NO: 176, SEQ ID NO: 177, SEQ ID NO: 178, SEQ ID NO: 179, SEQ ID NO: 180, SEQ ID NO: 181, SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 241, or SEQ ID NO: 242.

[0056] In the Grove embodiment, the fusion protein constructs are SEQ ID NOs: 43, 44, 46, 47, 48, 49, 50, and 52. , SEQ ID NO: 53, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 63, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 90, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: Sequence IDs 107, 109, 110, 111, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 140, 141, 142, 143, and 144 may contain at least one amino acid sequence. In some embodiments, the first polypeptide comprises the sequence of SEQ ID NO: 282 (domains A, B, C, and D) conjugated to a complement regulator polypeptide containing a sequence (domain R) selected from the group consisting of SEQ ID NOs: 41, 42, and 72, and the second polypeptide comprises the sequence of SEQ ID NO: 279 (domains E and F). In some embodiments, the first polypeptide comprises the sequence of SEQ ID NO: 282 (domains A, B, C, and D), and the second polypeptide comprises the sequence of SEQ ID NO: 279 (domains E and F) conjugated to a complement regulator polypeptide containing a sequence (domain R) selected from the group consisting of SEQ ID NOs: 72.

[0057] Another embodiment provides a pharmaceutical composition comprising a fusion protein construct according to any one of the above embodiments. Yet another embodiment provides a polynucleotide encoding a fusion protein. Another embodiment provides a therapeutic method comprising providing a therapeutically effective amount of the pharmaceutical composition to a target.

[0058] In some embodiments, the subjects suffer from complement-mediated disease or complement-mediated inflammation. In some embodiments, the fusion protein construct is specifically 10 C3d. -8 It binds with a KD affinity of M or less, and the target suffers from complement-mediated disease, which is characterized by increased deposition of C3d. In some embodiments, the fusion protein construct specifically binds to C2 antibody-reactive phospholipids. -8 The subjects bind with a KD affinity of M or less, and suffer from complement-mediated diseases, which are characterized by increased deposition of C2 antibody-reactive phospholipids. In some embodiments, subjects suffer from complement-mediated inflammation, which includes inflammatory fibrous diseases, including focal segmental glomerulosclerosis, primary sclerosing cholangitis, or membranoproliferative glomerulonephritis. In some embodiments, subjects suffer from complement-mediated autoimmune diseases, including rheumatoid arthritis, systemic lupus erythematosus, lupus nephritis, or pemphigus vulgaris. In some embodiments, subjects suffer from complement-mediated renal diseases, including membranoproliferative glomerulonephritis or complement-3 glomerulopathy. In some embodiments, subjects suffer from complement-mediated cardiovascular diseases, which include atherosclerosis or thrombosis. In some embodiments, subjects suffer from complement-mediated skin diseases.

[0059] In some embodiments, skin diseases include psoriasis, acne anomaly, lupus erythematous, cutaneous microvasculitis, urticaria, urticarial vasculitis, or bullous pemphigoid. In some embodiments, the subject suffers from complement-mediated inflammation, which includes ischemia / reperfusion injury, burns, endotoxemia and septic shock, adult respiratory distress syndrome, cardiopulmonary bypass, hemodialysis, anaphylactic shock, asthma, angioedema, Crohn's disease, sickle cell anemia, glomerulonephritis, and membranous This refers to symptoms or diseases selected from the group consisting of nephritis, pancreatitis, graft rejection, hyperacute xenograft rejection, recurrent miscarriage, pre-eclampsia, drug allergy, IL-2-induced vasoleap syndrome, X-ray contrast agent allergy, myasthenia gravis, Alzheimer's disease, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, insulin-dependent diabetes mellitus, acute disseminated encephalomyelitis, Addison's disease, antiphospholipid syndrome, autoimmune hepatitis, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, Hashimoto's disease, idiopathic thrombocytopenic purpura, pemphigus, Sjögren's syndrome, Takayasu's arteritis, myocardial infarction, stroke, acute respiratory distress syndrome, sepsis, plasmapheresis, platelet ferresis, leukocytapheresis, extracorporeal membrane oxygenation, heparinized extracorporeal LDL precipitation, enteritis, urticarial disorders, vasculitis, and lupus nephritis.

[0060] In some embodiments, the subjects include ischemia-reperfusion injury, rheumatoid arthritis (RA), lupus nephritis, atypical hemolytic uremic syndrome (aHUS), typical or infectious hemolytic uremic syndrome (tHUS), dense deposit disease (DDD), paroxysmal nocturnal hemoglobinuria (PNH), multiple sclerosis (MS), macular degeneration, and hemolysis, elevated liver enzymes, and low platelet count (HELLP) syndrome. Group, sepsis, dermatomyositis, diabetic retinopathy, thrombotic thrombocytopenic purpura (TTP), spontaneous miscarriage, microvasculitis, epidermolysis bullosa, recurrent miscarriage, multiple sclerosis (MS), traumatic brain injury, cardiovascular disease, myocarditis, cerebrovascular disease, peripheral vascular disease, renal vascular disease, mesenteric / intestinal vascular disease, vascular regeneration for grafts and / or replants, vasculitis, Henoch-Schönlein purpura nephritis, systemic The individual suffers from one of the following conditions or diseases selected from the group consisting of lupus erythematosus-associated vasculitis, vasculitis associated with rheumatoid arthritis, immune complex vasculitis, Takayasu's arthritis, capillary leak syndrome, dilated cardiomyopathy, diabetic vascular disease, thoracoabdominal aortic aneurysm, Kawasaki disease (arthritis), venous gas embolism (VGE), and restenosis after stent placement, rotational atherosclerosis resection, percutaneous transluminal coronary angioplasty (PTCA), myasthenia gravis, cold agglutinin disease (CAD), paroxysmal cold hemoglobinuria (PCH), dermatomyositis, scleroderma, warm autoimmune hemolytic anemia, Graves' disease, Hashimoto's thyroiditis, type 1 diabetes mellitus, psoriasis, pemphigus, autoimmune hemolytic anemia (AIHA), idiopathic thrombocytopenic purpura (ITP), Goodpasture syndrome, antiphospholipid syndrome (APS), Degos disease, and fulminant APS (CAPS).

[0061] In some embodiments, subjects suffer from a condition or disease selected from the group consisting of age-related macular degeneration (AMD), membranoproliferative glomerulonephritis type II (MPGN II), hemolytic uremic syndrome (HUS), asthma, amyloidosis, and thrombotic thrombocytopenic purpura. In some embodiments, hemolytic uremic syndrome (HUS) is atypical hemolytic uremic syndrome (aHUS). In some embodiments, subjects suffer from drusen-related diseases, or diseases associated with drusen. In some embodiments, drusen-related diseases include amyloidosis, fibrous elastica, dense deposit disease, glomerulonephritis, atherosclerosis, or drusen-related eye diseases.

[0062] Another embodiment provides a pharmaceutical composition comprising a tetravalent fusion protein construct for regulating complement activity, wherein the tetravalent fusion protein construct comprises (i) an antibody or antigen-binding fragment thereof that binds to a complement-related antigen, and (ii) a first complement regulator peptide and a second complement regulator peptide, wherein the first and second complement regulator polypeptides are either the same or different.

[0063] In some embodiments, at least one of the first and second complement-modulating peptides is conjugated to an antibody or antigen-binding fragment by a linker.

[0064] Another embodiment provides a pharmaceutical composition comprising a trivalent fusion protein construct for regulating complement activity, wherein the tetravalent fusion protein construct comprises (i) an antibody or antigen-binding fragment thereof that binds to a complement-related antigen, and (ii) a complement-modulating peptide.

[0065] In some embodiments, an antibody or its antigen-binding fragment may be conjugated with a complement-modulating peptide by a linker.

[0066] Another embodiment provides a pharmaceutical composition comprising a trivalent fusion protein construct for regulating complement activity, wherein the trivalent fusion protein construct comprises (i) a Fab that binds to a complement-related antigen, (ii) an antibody Fc domain, and (iii) a complement-modulating peptide conjugated to the Fab or Fc domain.

[0067] In some embodiments, the Fab or Fc domain and the complement-modulating peptide may be conjugated by a linker.

[0068] Another embodiment provides a pharmaceutical composition comprising a trivalent fusion protein construct, wherein a) a first polypeptide monomer comprising a CH2 or CH3 domain of an antibody, the domain having at least one orthogonal modification that favors heterodimer formation compared to homodimer formation; and b) a second polypeptide monomer comprising a CH2 or CH3 domain of an antibody, the domain having at least one orthogonal modification that favors heterodimer formation with the first polypeptide monomer compared to homodimer formation, wherein one of the first and second polypeptides further comprises a complement regulator peptide, and the trivalent fusion protein construct binds to a complement-related antigen.

[0069] In some embodiments, the first polypeptide includes a knob modification, and the second polypeptide includes a hole modification. In some embodiments, the first and second polypeptides include modifications that result in charge or surface complementarity. In some embodiments, a complement modifier peptide is linked to one of the first and second polypeptides by an amino acid linker.

[0070] Another embodiment includes (i) residues 26-33 (CDR-H1) of SEQ ID NOs: 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 259 and 31; 29, 260 and 31; residues 26-33 (CDR-H1) of SEQ ID NOs: 280, 281, 282, 284, 285, 286, 73 or 288, Three heavy chain complementarity determining regions (CDRs) having amino acid sequences of residues 26-33 (CDR-H1), 51-58 (CDR-H2), and 97-102 (CDR-H3) of SEQ ID NO: 244; or residues 26-33 (CDR-H1), 51-58 (CDR-H2), and 97-110 (CDR-H3) of SEQ ID NO: 290, or (ii) SEQ ID NOs: 11, 12, and 13; 17, 18, and 19; 23, 24, and 25; 29, 30, and 31; 35, 36, and 37; 147, 148, and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 259 and 31; or 29, 260 and 31; residues 26-33 (CDR-H1), 51-58 (CDR-H2) and 97-100 (CDR-H3) of SEQ ID NOs. 280, 281, 282, 284, 285, 286, 73 or 288; residues 26-33 (CDR-H1), 51-58 (CDR-H2) and 97- 102(CDR-H3); or three heavy chain CDRs having amino acid sequences that differ in that there is a single conserved amino acid substitution in one of the residues 26-33(CDR-H1), 51-58(CDR-H2), and 97-110(CDR-H3) of SEQ ID NO: 290; and (ii) SEQ ID NOs: 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 32, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233; residues 27-37 (CDR-L1), 55-57 (CDR-L2) and 94-102 (CDR-L3) of SEQ ID NOs. 279, 68, 287 or 59; or three light chain complementarity determining regions (CDRs) having amino acid sequences of residues 27-38 (CDR-L1), 56-58 (CDR-L2), and 95-102 (CDR-L3) of SEQ ID NOs. 289, or (ii) SEQ ID NOs. 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 32, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201 The present invention provides a pharmaceutical composition comprising an antibody or its antigen-binding fragment containing three light chain CDRs having amino acid sequences that differ in that one of the residues 27-37 (CDR-L1), 55-57 (CDR-L2), and 94-102 (CDR-L3) of SEQ ID NOs. 207, 208, and 209; 215, 216, and 217; 223, 224, and 225; or 231, 232, and 233; residues 27-37 (CDR-L1), 55-57 (CDR-L2), and 94-102 (CDR-L3) of SEQ ID NOs. 279, 68, 287, or 59 contains a single conserved amino acid substitution, or one of the residues 27-38 (CDR-L1), 56-58 (CDR-L2), and 95-102 (CDR-L3) of SEQ ID NO. 289, and a fusion protein construct containing a complement-modulating peptide.

[0071] Another embodiment is (a) Sequence IDs 54, 58, 67, 73, 74, 81, 88, 89, 98, 99, 112, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290 and (b) A heavy chain variable region comprising at least one amino acid sequence from among 342, and a light chain variable region comprising at least one amino acid sequence from among SEQ ID NOs: 45, 59, 68, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277 and 278, 279, 287 and 289, or (c) SEQ ID NOs: 54, 58, 67, 73, 74, 81, 88, 89, 98, 99, 112, 145 The amino acid sequences differ in that at least one of the following has one or more conserved amino acid substitutions: 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290, and 342. The present invention provides a pharmaceutical composition comprising a fusion protein construct comprising an antibody or an antigen-binding fragment thereof, comprising a light chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs. 45, 59, 68, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277 and 278, 279, 287 and 289 contains one or more conserved amino acid substitutions.

[0072] Another embodiment is sequence numbers 45, 51, 54, 58, 59, 62, 64, 67, 68, 73, 74, 79, 75, 81, 88, 89, 98, 99, 112, 121, 194, 195, 202, 203, 210, 211, 218, 219, 237, 226, 226, 227, 234, 235, 238, 239, 240, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255 The present invention provides a pharmaceutical composition comprising a fusion protein construct containing a sequence selected from the group consisting of 256, 257, 258, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277 and 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290 and 342, as well as a complement-modulating peptide.

[0073] In some embodiments, the complement modulator peptide is linked to an antibody or its antigen-binding fragment by an amino acid linker. In some embodiments, the amino acid linker includes one of the amino acid sequences of SEQ ID NO: 138, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169, SEQ ID NO: 170, SEQ ID NO: 171, SEQ ID NO: 172, SEQ ID NO: 173, SEQ ID NO: 174, SEQ ID NO: 175, SEQ ID NO: 176, SEQ ID NO: 177, SEQ ID NO: 178, SEQ ID NO: 179, SEQ ID NO: 180, SEQ ID NO: 181, SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 241, or SEQ ID NO: 242. In some embodiments, the pharmaceutical composition includes an antigen-binding fragment, which includes an Fv fragment, Fab, Fab', F(ab')2, or scFv. In some embodiments, when the pharmaceutical composition is administered to a subject, the fusion protein construct modulates surrogate complement activity in the subject. In some embodiments, the antibody or its antigen-binding fragment, or Fab, binds to the domain of the mammalian annexin protein. -8It binds with a KD affinity of less than M. In some embodiments, the domain is an annexin core domain. In some embodiments, the annexin core domain includes an alpha helical domain. In some embodiments, the annexin core domain includes a calcium binding site and a membrane binding site. In some embodiments, the annexin core domain includes at least one annexin repeat sequence. In some embodiments, an antibody or an antigen-binding fragment thereof, or a Fab binds to at least one annexin repeat sequence.

[0074] In some embodiments, the complement-related antigen includes a phospholipid, and an antibody or an antigen-binding fragment thereof, or a Fab binds to the phospholipid. In some embodiments, the antibody or an antigen-binding fragment thereof, or a Fab binds to the phospholipid with a KD affinity of less than 10 -8 It binds with a KD affinity of less than M. In some embodiments, the phospholipid is selected from the group consisting of phosphatidylethanolamine (PE), cardiolipin (CL), phosphatidylcholine (PC), and malondialdehyde (MDA). In some embodiments, the complement-related antigen includes a C3 complement protein or a fragment thereof, and an antibody or an antigen-binding fragment thereof, or a Fab binds to the C3 complement protein or a fragment thereof. In some embodiments, the antibody or an antigen-binding fragment thereof, or a Fab binds to the C3 complement protein or a fragment thereof with a KD affinity of less than 10 -8 It binds with a KD affinity of less than M. In some embodiments, the C3 complement protein fragment is C3d. In some embodiments, the complement regulator peptide includes a complement receptor 1 (CR1) protein. In some embodiments, the complement regulator peptide includes domain A of the CR1 protein, or a fragment thereof that retains at least three short consensus repeats (SCRs) of domain A. In some embodiments, the complement regulator peptide includes domain B of the CR1 protein, or a fragment thereof that retains at least three SCRs of domain B.

[0075] In some embodiments, the complement regulator peptide comprises domain C of the CR1 protein, or a fragment thereof holding at least three SCRs of domain C. In some embodiments, the pharmaceutical composition further comprises domain D of the CR1 protein, or a fragment thereof holding at least three SCRs of domain D. In some embodiments, the complement regulator peptide comprises the first three SCRs of domain A, the first three SCRs of domain B, and the first three SCRs of domain C of the CR1 protein. In some embodiments, the CR1 protein is human CR1 protein. In some embodiments, the complement regulator polypeptide is CR1(1-10). In some embodiments, the complement regulator polypeptide is CR1(1-17). In some embodiments, CR1(1-10) comprises the amino acid sequence of SEQ ID NO: 41 or SEQ ID NO: 91, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, CR1(1-17) comprises the amino acid sequence of SEQ ID NO: 42 or SEQ ID NO: 92, or at least 85% identical. This includes variants having the amino acid sequence. In some embodiments, the complement regulator peptide is a complement breakdown promoter (DAF) or a bioactive fragment thereof. In some embodiments, the DAF is human DAF.

[0076] In some embodiments, the bioactive fragment of human DAF comprises at least one of the short consensus repeat (SCR) domain and the O-glycosylated serine / threonine-rich domain of full-length human DAF. In some embodiments, the bioactive fragment of human DAF comprises SCR1-4 or SCR2-4 of full-length human DAF. In some embodiments, the bioactive fragment of human DAF comprises the amino acid sequence of SEQ ID NO: 184, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is factor H or a bioactive fragment thereof. In some embodiments, factor H is human factor H. In some embodiments, the bioactive fragment of human factor H comprises a sequence of amino acids selected from the group consisting of amino acids 21-266, 21-320, 21-509, or 19-1106 of SEQ ID NO: 9, or a variant thereof having an amino acid sequence that is at least 85% identical to said sequence of amino acids. In some embodiments, the bioactive fragment of human factor H comprises one or more groups of short consensus repeats (SCRs) including SCR1-20, SCR1-2, SCR2-3, SCR3-4, SCR4-5, SCR5-6, SCR6-7, SCR7-8, SCR8-9, SCR9-10, SCR10-11, SCR11-12, SCR12-13, SCR13-14, SCR14-15, SCR15-16, SCR16-17, SCR17-18, SCR19-20, or any combination of SCR1-20 of full-length human factor H.

[0077] In some embodiments, the bioactive fragment of human factor H includes SCR1-4 of full-length human factor H. In some embodiments, the bioactive fragment of human factor H includes SCR1-5 of full-length human factor H. In some embodiments, the bioactive fragment of human factor H includes the amino acid sequence of SEQ ID NO: 72 or SEQ ID NO: 108, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is MCP or a bioactive fragment thereof. In some embodiments, MCP is human MCP. In some embodiments, the bioactive fragment of human MCP includes at least one short consensus repeat (SCR) domain of full-length human MCP. In some embodiments, the bioactive fragment of human MCP includes SCR3-4 of full-length human MCP. In some embodiments, the bioactive fragment of human MCP includes the amino acid sequence of SEQ ID NO: 187, or a variant thereof having an amino acid sequence that is at least 85% identical.

[0078] In some embodiments, the complement regulator peptide is Map44 or a bioactive fragment thereof. In some embodiments, Map44 is human Map44. In some embodiments, Map44 includes the amino acid sequence of SEQ ID NO: 186, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the complement regulator peptide is CD59 or a bioactive fragment thereof. In some embodiments, CD59 is human CD59. In some embodiments, CD59 includes the amino acid sequence of SEQ ID NO: 185, or a variant thereof having an amino acid sequence that is at least 85% identical. In some embodiments, the antibody or its antigen-binding fragment is a human antibody or its antigen-binding fragment. In some embodiments, the antibody or its antigen-binding fragment is a humanized antibody or its antigen-binding fragment.

[0079] Another embodiment provides a polynucleotide encoding a fusion protein. Another embodiment provides a therapeutic method comprising providing a therapeutically effective amount of a pharmaceutical composition to a subject. In some embodiments, the subject suffers from complement-mediated inflammation. In some embodiments, complement-mediated inflammation includes inflammatory fibrous diseases, and inflammatory fibrous diseases include focal segmental glomerulosclerosis, primary sclerosing cholangitis, or membranoproliferative glomerulonephritis. In some embodiments In some embodiments, subjects suffer from complement-mediated autoimmune diseases, including rheumatoid arthritis, systemic lupus erythematosus, lupus nephritis, or pemphigus vulgaris. In some embodiments, subjects suffer from complement-mediated renal diseases, including membranoproliferative glomerulonephritis or complement triglomerulopathy. In some embodiments, subjects suffer from complement-mediated cardiovascular diseases, including atherosclerosis or thrombosis. In some embodiments, subjects suffer from complement-mediated skin diseases, including psoriasis, acne-like erythematosus, lupus erythematosus, cutaneous microvasculitis, urticaria, urticarial vasculitis, and bullous pemphigoid.

[0080] In some embodiments, complement-mediated inflammation is associated with ischemia / reperfusion injury, burns, endotoxemia and septic shock, adult respiratory distress syndrome, cardiopulmonary bypass, hemodialysis, anaphylactic shock, asthma, angioedema, Crohn's disease, sickle cell anemia, glomerulonephritis, membranous nephritis, pancreatitis, graft rejection, hyperacute xenograft rejection, recurrent miscarriage, preeclampsia, drug allergies, IL-2-induced vasoleap syndrome, X-ray contrast agent allergies, myasthenia gravis, Alzheimer's disease, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, insulin This refers to symptoms or diseases selected from the group consisting of diabetes mellitus vera, acute disseminated encephalomyelitis, Addison's disease, antiphospholipid syndrome, autoimmune hepatitis, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, Hashimoto's disease, idiopathic thrombocytopenic purpura, pemphigus, Sjögren's syndrome, Takayasu's arteritis, myocardial infarction, stroke, acute respiratory distress syndrome, sepsis, plasmapheresis, platelet fertilization, leukocytapheresis, extracorporeal membrane oxygenation, heparinized extracorporeal LDL precipitation, enteritis, urticarial and vasculitis, and lupus nephritis.

[0081] In some embodiments, the subjects include ischemia-reperfusion injury, rheumatoid arthritis (RA), lupus nephritis, atypical hemolytic uremic syndrome (aHUS), typical or infectious hemolytic uremic syndrome (tHUS), dense deposit disease (DDD), paroxysmal nocturnal hemoglobinuria (PNH), multiple sclerosis (MS), macular degeneration, and hemolysis, elevated liver enzymes, and low platelet count (HELLP) syndrome. Group, sepsis, dermatomyositis, diabetic retinopathy, thrombotic thrombocytopenic purpura (TTP), spontaneous miscarriage, microvasculitis, epidermolysis bullosa, recurrent miscarriage, multiple sclerosis (MS), traumatic brain injury, cardiovascular disease, myocarditis, cerebrovascular disease, peripheral vascular disease, renal vascular disease, mesenteric / intestinal vascular disease, vascular regeneration for grafts and / or replants, vasculitis, Henoch-Schönlein purpura nephritis, systemic The individual suffers from one of the following conditions or diseases selected from the group consisting of lupus erythematosus-associated vasculitis, vasculitis associated with rheumatoid arthritis, immune complex vasculitis, Takayasu's arthritis, capillary leak syndrome, dilated cardiomyopathy, diabetic vascular disease, thoracoabdominal aortic aneurysm, Kawasaki disease (arthritis), venous gas embolism (VGE), and restenosis after stent placement, rotational atherosclerosis resection, percutaneous transluminal coronary angioplasty (PTCA), myasthenia gravis, cold agglutinin disease (CAD), paroxysmal cold hemoglobinuria (PCH), dermatomyositis, scleroderma, warm autoimmune hemolytic anemia, Graves' disease, Hashimoto's thyroiditis, type 1 diabetes mellitus, psoriasis, pemphigus, autoimmune hemolytic anemia (AIHA), idiopathic thrombocytopenic purpura (ITP), Goodpasture syndrome, antiphospholipid syndrome (APS), Degos disease, and fulminant APS (CAPS).

[0082] In some embodiments, subjects may have a condition or disease selected from the group consisting of age-related macular degeneration (AMD), membranoproliferative glomerulonephritis type II (MPGN II), hemolytic uremic syndrome (HUS), asthma, amyloidosis, and thrombotic thrombocytopenic purpura. In some embodiments, hemolytic uremic syndrome (HUS) is atypical hemolytic uremic syndrome (aHUS). In some embodiments, subjects may have a drusen-related disease or a disease associated with drusen. In some embodiments, a drusen-related disease is amyloidosis, fibrosis elastica, dense deposit disease, glomerulonephritis, atherosclerosis, or a drusen-related eye disease.

[0083] Further embodiments include a fusion protein construct that binds to a complement-related antigen, comprising a first polypter comprising domains A and B aligned in an AB orientation from the N-terminus to the C-terminus. The present invention provides a monomeric fusion protein construct comprising a first polypeptide, a second polypeptide comprising domains E and F aligned in an E-F orientation from the N-terminus to the C-terminus, wherein at least one of domains A and E may be conjugated to domain R, domain A may contain a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B may contain a heavy chain CH1 constant region amino acid sequence, domain R may contain a complement regulator polypeptide, domain E may contain a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and domain F may contain a light chain constant region amino acid sequence (CL1). In some embodiments, the first polypeptide comprises domains A, B and R, the domains of the first polypeptide aligned in an RAB orientation from the N-terminus to the C-terminus, and domain R and domain A are conjugated. In some embodiments, the second polypeptide comprises domains E, F, and R, the domains of the second polypeptide are aligned in a REF orientation from the N-terminus to the C-terminus, and domains E and R are conjugated.

[0084] Another embodiment provides a fusion protein construct comprising an antibody or its antigen-binding fragment that specifically binds to complement protein 3d (c3d), and two molecules of a complement regulator polypeptide, each molecule of which contains a physiologically active fragment of a complement protein selected from the group consisting of CR1, DAF, MCP, Crry, MAp44, MAp19, CD59, and factor H, wherein the median inhibitory concentration in a complement assay is lower than that of a control protein construct that is identical except for the absence of an antibody.

[0085] In some embodiments, each molecule of the complement-modulating polypeptide may be conjugated to the heavy chain of an antibody or its antigen-binding fragment. In some embodiments, each molecule of the complement-modulating polypeptide may be complexed to the C-terminus of the heavy chain.

[0086] In some embodiments, the antibody or its antigen-binding fragment may comprise a first polypeptide and a second polypeptide, the first polypeptide may comprise a heavy chain sequence comprising at least one amino acid sequence from among SEQ ID NOs. 54, SEQ ID NOs. 58, SEQ ID NOs. 60, SEQ ID NOs. 61, SEQ ID NOs. 64, SEQ ID NOs. 65, SEQ ID NOs. 67, SEQ ID NOs. 69, SEQ ID NOs. 70, SEQ ID NOs. 71, SEQ ID NOs. 73, SEQ ID NOs. 74, SEQ ID NOs. 75, SEQ ID NOs. 76, SEQ ID NOs. 78, SEQ ID NOs. 80, SEQ ID NOs. 81, SEQ ID NOs. 82, SEQ ID NOs. 85, SEQ ID NOs. 87, SEQ ID NOs. 88, SEQ ID NOs. 132, SEQ ID NOs. 133, SEQ ID NOs. 134, SEQ ID NOs. 243, SEQ ID NOs. 282, SEQ ID NOs. 284, SEQ ID NOs. 285, and SEQ ID NOs. 286, the second polypeptide may comprise a light chain sequence comprising at least one amino acid sequence from among SEQ ID NOs. 59, SEQ ID NOs. 68, SEQ ID NOs. 77, SEQ ID NOs. 79, SEQ ID NOs. 84, SEQ ID NOs. 86, SEQ ID NOs. 135, and SEQ ID NOs. 27.

[0087] In some embodiments, the first polypeptide may contain at least two amino acid sequences selected from the group consisting of SEQ ID NOs: 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 74, 75, 76, 78, 80, 81, 82, 85, 87, 88, 132, 133, 134, 243, 282, 284, 285, and 286, and the second polypeptide may contain at least two amino acid sequences selected from the group consisting of SEQ ID NOs: 59, 68, 77, 79, 84, 86, 135, and 279.

[0088] In some embodiments, at least two amino acid sequences of the first polypeptide may be the same amino acid sequence, and at least two amino acid sequences of the second polypeptide may be the same amino acid sequence. The sequences may be such. In some embodiments, at least two amino acid sequences of the first polypeptide may be SEQ ID NO: 282 or SEQ ID NO: 285, and at least two amino acid sequences of the second polypeptide may be SEQ ID NO: 279.

[0089] In some embodiments, the fusion protein construct further includes a linker. In some embodiments, the linker may include any of the amino acid sequences of SEQ ID NO: 138, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169, SEQ ID NO: 170, SEQ ID NO: 171, SEQ ID NO: 172, SEQ ID NO: 173, SEQ ID NO: 174, SEQ ID NO: 175, SEQ ID NO: 176, SEQ ID NO: 177, SEQ ID NO: 178, SEQ ID NO: 179, SEQ ID NO: 180, SEQ ID NO: 181, SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 241, or SEQ ID NO: 242.

[0090] In some embodiments, the complement protein is factor H or a bioactive fragment thereof. In some embodiments, factor H or a bioactive fragment thereof may include the amino acid sequence of SEQ ID NO: 72 or SEQ ID NO: 108, or a variant thereof having an amino acid sequence that is at least 85% identical.

[0091] In some embodiments, the complement protein is CR1 or a fragment thereof. In some embodiments, the complement protein may include the amino acid sequence of SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 91, or SEQ ID NO: 92, or a variant thereof having an amino acid sequence that is at least 85% identical.

[0092] In the step embodiment, the fusion protein construct is SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 63, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 90, SEQ ID NO: 93, SEQ ID NO: 94, SEQ ID NO: 95, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 10 2. Sequence IDs 103, 104, 105, 106, 107, 109, 110, 111, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 140, 141, 142, 143, 144, 279, 282, and 285 may contain at least one amino acid sequence.

[0093] Another embodiment is a fusion protein construct comprising an antibody or antigen-binding fragment that specifically binds to a complement-related antigen, wherein the antibody comprises a first polypeptide and a second polypeptide, each comprising a heavy chain and a light chain; further comprising a first molecule and a second molecule of a complement modulator polypeptide, each comprising a physiologically active fragment of a complement protein selected from the group consisting of CR1, DAF, MCP, Crry, MAp44, MAp19, CD59 and factor H, wherein when the fusion protein construct is administered to a subject with a disease, the albumin-to-creatinine ratio of a urine sample from the subject with the disease is equivalent to that of a subject administered with an equivalent fusion protein construct. The present invention provides a fusion protein construct that has a lower albumin-to-creatine ratio than that of a urine sample from a subject, and is identical to an equivalent fusion protein construct except that it does not contain an antibody or its antigen-binding fragment.

[0094] In some embodiments, the disease is a complement-mediated renal disease. In some embodiments, the complement-mediated renal disease is membranoproliferative glomerulonephritis or complement triglomerulopathy.

[0095] In some embodiments, the albumin-to-creatinine ratio of urine samples from subjects with the disease may be at least about 1%, 2%, 3%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, and 85% lower (compared to subjects administered with an equivalent fusion protein construct).

[0096] In the first embodiment, sequence numbers 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 74, 75, 76, 78, 80, 81, 82, 85, 87, 88, 89, 98, 99, 112, 132, 133, 134, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, A heavy chain containing at least one amino acid sequence from among 282, 283, 284, 285, 286, 288, 290 and 342, and a light chain containing at least one amino acid sequence from among SEQ ID NOs: 45, 59, 68, 77, 79, 84, 86, 135, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, 278, 279, 287 and 289, or SEQ ID NOs: 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 7 At least one of 4, 75, 76, 78, 80, 81, 82, 85, 87, 88, 89, 98, 99, 112, 132, 133, 134, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290, and 342 The present invention provides a fusion protein construct comprising an antibody or an antigen-binding fragment thereof, comprising a heavy chain containing an amino acid sequence that differs in having a conserved amino acid substitution, and a light chain containing an amino acid sequence that differs in having one or more conserved amino acid substitutions in at least one of SEQ ID NOs: 45, 59, 68, 77, 79, 84, 86, 135, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, 278, 279, 287, and 289.

[0097] In some embodiments, the fusion protein construct may further comprise a complement regulator polypeptide, which comprises at least one of the complement receptor 1 (CR1) protein, DAF, MCP, Crry, MAp44, MAp19, CD59, factor H, and its physiologically active fragments.

[0098] Reference All publications, patents, patent applications, and NCBI accession numbers referenced herein are incorporated herein by reference to the same extent as they are incorporated specifically and individually with respect to each publication, patent, or patent application, and to the same extent as they are incorporated as a whole. In the event of any conflict between terms used herein and terms defined in the references to which they are incorporated, the definitions in this disclosure shall prevail.

[0099] Novel features of this disclosure are described in detail in the claims set forth separately. Features of this disclosure and A better understanding of the advantages will be obtained by referring to the following detailed description illustrating exemplary embodiments in which the principles of this disclosure are utilized, and to the accompanying drawings. [Brief explanation of the drawing]

[0100] [Figure 1] An exemplary monomeric fusion protein construct of this disclosure is shown, comprising a linkage between the target-directed light chain and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the N-terminus of the light chain via the linker. [Figure 2] An exemplary monomeric fusion protein construct of this disclosure is shown, comprising a linkage between the heavy chain of the target-directed moiety and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the N-terminus of the heavy chain via the linker. [Figure 3] An exemplary monomeric fusion protein construct of this disclosure is shown, comprising a linkage between the heavy chain of the target-directed moiety and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the C-terminus of the heavy chain via the linker. [Figure 4]An exemplary monomeric fusion protein construct of this disclosure is shown, comprising a linkage between the target-directed light chain and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the C-terminus of the light chain via the linker. [Figure 5] An exemplary tetravalent homodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the heavy chain of the target-directed moiety and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the C-terminus of the heavy chain via the linker. [Figure 6] An exemplary tetravalent homodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the heavy chain of the target-directed moiety and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the N-terminus of the heavy chain via the linker. [Figure 7] An exemplary trivalent heterodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the heavy chain of the target-directed moiety and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the C-terminus of the heavy chain via the linker. [Figure 8] An exemplary trivalent heterodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the heavy chain of the target-directed moiety and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the N-terminus of the heavy chain via the linker. [Figure 9] An exemplary tetravalent homodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the target-directed light chain and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the C-terminus of the light chain via the linker. [Figure 10] An exemplary tetravalent homodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the target-directed light chain and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the N-terminus of the light chain via the linker. [Figure 11] An exemplary trivalent heterodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the target-directed light chain and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the N-terminus of the light chain via the linker. [Figure 12]An exemplary trivalent heterodimeric fusion protein construct of this disclosure is shown, comprising a linkage between the target-directed light chain and the complement regulator polypeptide. The complement regulator polypeptide is ligated to the C-terminus of the light chain via the linker. [Figure 13] An exemplary fusion protein construct of this disclosure is shown, which includes a linkage between the N-terminus of the Fc region of the target-directing moiety and a complement regulator polypeptide. [Figure 14] This disclosure shows an exemplary design of a C2-complement regulator fusion protein construct. [Figure 15] This disclosure shows an exemplary design of an anti-C3d-complement regulator fusion protein construct. [Figure 16] This disclosure shows an exemplary design of an anti-C3d complement regulator fusion protein construct. [Figure 17] This disclosure shows an exemplary design of an anti-C3d complement regulator fusion protein construct. [Figure 18] This disclosure shows an exemplary design of an anti-C3d complement regulator fusion protein construct. [Figure 19] An exemplary fusion protein construct of this disclosure is shown, which is a tetravalent heterodimer containing two different complement regulator polypeptides linked to the C-terminus of a heavy chain via a polypeptide linker. [Figure 20] This report presents results from PEGperMAP® linear epitope mapping and five mouse IgG1 antibodies against the C3dg extension. Linear C3d epitopes were determined using PEPperPRINT technology (PEPperPRINT GmbH, Heidelberg, Germany). A 15-amino acid peptide derived from the C3dg primary sequence, offset by one amino acid, was synthesized in two replicates on a PEPperPRINT microarray. The array was then incubated with antibodies (e.g., 3d8b, 3d9a, 3d29) and stained with secondary DyLight680-labeled goat anti-mouse IgG(H+L) antibody. The microarrays were read using the LI-COR Odyssey imaging system and analyzed by PEPperPRINT. [Figure 21]This graph illustrates the recognition of the C-terminal epitope of C3dg by the exemplary anti-C3d antibody 3d29. Sequence numbers 301-341 are disclosed on the x-axis of the graph in order of appearance, with sequence number 299 being the sequence "NLDVSLQLPS". [Figure 22] This graph illustrates the recognition of the C-terminal epitope of C3dg by the exemplary anti-C3d antibody 3d8b. Sequence numbers 301-341 are disclosed on the x-axis of the graph in order of appearance, with sequence number 299 being the sequence "NLDVSLQLPS". [Figure 23] This graph illustrates the recognition of the C-terminal epitope of C3dg by the exemplary anti-C3d antibody 3d9a. Sequence numbers 301-341 are disclosed on the x-axis of the graph in order of appearance, with sequence number 299 being the sequence "NLDVSLQLPS". [Figure 24] This graph shows epitope mapping using negative control anti-C3d antibodies that did not bind to linear epitopes, tested at various concentrations. Sequence IDs 301-341 are displayed on the x-axis of the graph in order of appearance. [Figure 25] This graph shows epitope mapping using negative control anti-C4d antibodies tested at various concentrations. Sequence IDs 301-341 are displayed on the x-axis of the graph in order of appearance. [Figure 26]A-L show exemplary designs of the C2-complement regulator (CR1) fusion protein constructs of the present disclosure. A shows a fusion protein construct comprising CR1(1-10) polypeptide and exemplary C2-scFv, where the CR1(1-10) polypeptide is ligated to the C-terminus of the light chain variable domain of exemplary C2-scFv. B shows a fusion protein construct comprising Crry polypeptide and exemplary C2-scFv, where the Crry polypeptide is ligated to the C-terminus of the light chain variable domain of exemplary C2-scFv. C shows a fusion protein construct comprising Crry polypeptide and exemplary C2-Fab, where the Crry polypeptide is ligated to the C-terminus of the heavy chain of exemplary C2-Fab. D shows a fusion protein construct comprising CR1(1-10) polypeptide and exemplary C2-Fab, where the CR1(1-10) polypeptide is ligated to the N-terminus of the heavy chain of exemplary C2-Fab. E shows a fusion protein construct containing CR1(1-10) polypeptide and exemplary C2-Fab, where CR1(1-10) is ligated to the C-terminus of the heavy chain of exemplary C2-Fab. F shows a fusion protein construct containing CR1(1-17) polypeptide and exemplary C2-Fab, where CR1(1-17) polypeptide is ligated to the N-terminus of the heavy chain of exemplary C2-Fab. G shows a fusion protein construct containing CR1(1-17) polypeptide and exemplary C2-Fab, where CR1(1-17) polypeptide is ligated to the C-terminus of the heavy chain of exemplary C2-Fab. H shows a fusion protein construct containing a single CR1(1-10) polypeptide and exemplary full-length C2 antibody, where CR1(1-10) is ligated to the C-terminus of the heavy chain of exemplary full-length C2 antibody, and the fusion protein construct includes a knob-into-hole heterodimer antibody construct. Figure I shows a fusion protein construct comprising two CR1(1-10) polypeptides and an exemplary full-length C2 antibody, where each C-terminus of each heavy chain of the exemplary full-length C2 antibody is ligated to a single CR1(10) polypeptide, and the fusion protein construct contains a knob-into-hole heterodimer antibody construct.J shows a fusion protein construct comprising a single CR1(1-10) polypeptide and an exemplary C2 antibody fragment containing a variable heavy chain region, a variable light chain region, and a constant region, with the single CR1(10) polypeptide ligated to the N-terminus of the CH2-CH3 constant region of the exemplary C2 antibody fragment. K shows a fusion protein construct comprising a single CR1(1-10) polypeptide and an exemplary full-length C2 antibody, with the single CR1(10) polypeptide ligated to one N-terminus of the light chain of the exemplary full-length C2 antibody. L shows a fusion protein construct comprising a single CR1(1-10) polypeptide and an exemplary full-length C2 antibody, with the single CR1(10) polypeptide ligated to one N-terminus of the heavy chain of the exemplary full-length C2 antibody. [Figure 27] A–E illustrate exemplary designs of the anti-C3d(3d29)-complement regulator (CR1 or factor H) fusion protein constructs of the present disclosure. A shows a fusion protein construct comprising a Crry polypeptide and exemplary anti-C3d-Fab, where the Crry polypeptide is ligated to the C-terminus of the heavy chain of exemplary anti-C3d-Fab. B shows a fusion protein construct comprising a CR1(1-10) polypeptide and exemplary anti-C3d Fab, where the CR1(1-10) polypeptide is ligated to the C-terminus of the heavy chain of exemplary anti-C3d Fab. C shows a fusion protein construct comprising factor H and exemplary anti-C3d Fab, where factor H is ligated to the C-terminus of the heavy chain of exemplary anti-C3d Fab. D shows a fusion protein construct comprising a single factor H polypeptide and exemplary full-length anti-C3d antibody, where the single factor H polypeptide is ligated to the C-terminus of the heavy chain of exemplary full-length anti-C3d antibody. Figure E shows a fusion protein construct containing two single H factor polypeptides and an exemplary full-length anti-C3d antibody, where the C-terminus of each heavy chain of the exemplary full-length anti-C3d antibody is ligated to a single H factor polypeptide. [Figure 28]A–H show exemplary designs of the anti-C3d(3d8b)-complement regulator (CR1 or factor H) fusion protein constructs of the present disclosure. A shows a fusion protein construct comprising the Crry polypeptide and exemplary anti-C3d Fab, where Crry is ligated to the C-terminus of the heavy chain of exemplary anti-C3d Fab. B shows a fusion protein construct comprising the CR1(1-10) polypeptide and exemplary anti-C3d Fab, where the CR1(1-10) polypeptide is ligated to the C-terminus of the heavy chain of exemplary anti-C3d Fab. C shows a fusion protein construct comprising the factor H polypeptide and exemplary anti-C3d Fab, where the factor H polypeptide is ligated to the C-terminus of the heavy chain of exemplary anti-C3d Fab. D shows a fusion protein construct comprising a single CR1(1-10) polypeptide and exemplary full-length anti-C3d antibody, where the single CR1(1-10) polypeptide is ligated to one C-terminus of the light chain of exemplary full-length anti-C3d antibody. E shows a fusion protein construct containing a single CR1(1-10) polypeptide and an exemplary full-length anti-C3d antibody, where the single CR1(1-10) polypeptide is ligated to one C-terminus of the heavy chain of the exemplary full-length antibody. F shows a fusion protein construct containing two CR1(1-10) polypeptides and an exemplary full-length anti-C3d antibody, where the C-terminus of each heavy chain of the exemplary full-length anti-C3d antibody is ligated to a single CR1(1-10) polypeptide. G shows a fusion protein construct containing a single H factor polypeptide and an exemplary full-length anti-C3d antibody, where the single H factor polypeptide is ligated to the C-terminus of the heavy chain of the exemplary full-length anti-C3d antibody. H shows a fusion protein construct containing two H factor polypeptides and an exemplary full-length anti-C3d antibody, where the C-terminus of each heavy chain of the exemplary full-length anti-C3d antibody is ligated to a single H factor polypeptide. [Figure 29]Figures A-C illustrate exemplary designs of the anti-C3d(3d8b)-complement regulator (factor H) fusion protein constructs of the present disclosure. A shows the fH1-5-3d8b heavy chain mouse IgG1 (the complement regulator is linked to the N-terminus of the heavy chain via a linker). B shows the 3d8b kappa light chain-fH1-5 (the complement regulator is linked to the C-terminus of the light chain via a linker). C shows the fH1-5-3d8b kappa light chain (the complement regulator is linked to the N-terminus of the light chain via a linker). [Figure 30] The present disclosure shows an exemplary design of the IgG1-complement regulator (Factor H) fusion protein construct, in which the complement regulator (Factor H) is linked to the hinge region of IgG1 via a linker. [Figure 31]Figures A-F illustrate exemplary designs of the anti-C3d(3d8b)-complement modifier (CR1 1-17) fusion protein constructs of the present disclosure. Figure A shows a fusion protein construct comprising one CR1(1-17) polypeptide and an exemplary Fab fragment of the anti-C3d antibody 3d8b, with the C-terminus of the heavy chain of the exemplary Fab fragment ligated to a single CR1(1-17) polypeptide. Figure B shows a fusion protein construct comprising one CR1(1-17) polypeptide and an exemplary full-length anti-C3d 3d8b antibody, with one C-terminus of the heavy chain of the exemplary full-length anti-C3d 3d8b antibody ligated to a single CR1(1-17) polypeptide. Figure C shows a fusion protein construct comprising two CR1(1-17) polypeptides and exemplary full-length anti-C3d 3d8b antibodies, with the C-terminus of each heavy chain of the exemplary full-length anti-C3d 3d8b antibodies ligated to a single CR1(1-17) polypeptide. D shows a fusion protein construct containing two CR1(1-17) polypeptides and an exemplary full-length anti-C3d 3d8b antibody, where the C-terminus of each light chain of the exemplary full-length anti-C3d 3d8b antibody is ligated to a single CR1(1-17) polypeptide. E shows a fusion protein construct containing two CR1(1-17) polypeptides and an exemplary full-length anti-C3d 3d8b antibody, where the N-terminus of each heavy chain of the exemplary full-length anti-C3d 3d8b antibody is ligated to a single CR1(1-17) polypeptide. F shows a fusion protein construct containing two CR1(1-17) polypeptides and an exemplary full-length anti-C3d 3d8b antibody, where the N-terminus of each light chain of the exemplary full-length anti-C3d 3d8b antibody is ligated to a single CR1(1-17) polypeptide. [Figure 32] Figures A and B show that the quantification of C3-positive staining is expressed as a percentage of total glomerular area (ImageJ software analysis) or as a semi-quantitative score. Figure A shows the staining of glomerular C3 deposits in a collected kidney, measured by immunofluorescence (IF). Figure B shows C3 deposits in a collected liver using IF. [Figure 33]Figures A and B show that the quantification of C3-positive staining is expressed as a percentage of total glomerular area (ImageJ software analysis) or as a semi-quantitative score. Figure A shows the staining of glomerular C3 deposits in a collected kidney as measured by IF. Figure B shows C3 deposits in a collected liver using IF. [Figure 34] This shows the percentage of infarct volume in Balb / c mice. [Figure 35] This shows the difference in urinary albumin:creatinine ratio (uACR) in Adriamycin-damaged mice between day 8 and day 22 of the study. Day 8 represents the pre-treatment state, and day 22 represents 12 days after administration of the complement inhibitor (treatment). [Modes for carrying out the invention]

[0101] This disclosure provides a fusion protein construct comprising an antibody or its antigen-binding fragment and two molecules of a complement-modulating polypeptide. The antibody or its antigen-binding fragment can specifically bind to a complement protein. The complement protein may be, for example, complement protein 3d (c3d). In some cases, the molecules of the complement-modulating polypeptide may include at least a physiologically active fragment of the complement protein. The complement protein may be, for example, CR1, DAF, MCP, Crry, MAp44, MAp19, CD59, or factor H. In some cases, the median inhibitory concentration (e.g., based on a complement assay) of the fusion protein construct is lower than that of a control protein construct that is identical except for the absence of an antibody. In some cases, the molecules of the complement-modulating polypeptide may be conjugated to the heavy chain of the antibody or its antigen-binding fragment. In some cases, the molecules of the complement-modulating polypeptide may be conjugated to the light chain of the antibody or its antigen-binding fragment. In some cases, the molecules of the complement-modulating polypeptide may be conjugated to the C-terminus of the heavy chain and / or light chain. In some cases, the complement-modulating polypeptide molecule is conjugated to the N-terminus of the heavy and / or light chain.

[0102] This disclosure provides a fusion protein construct comprising an antibody or its antigen-binding fragment that specifically binds to a complement-related antigen, and a first and second molecule of a complement-modulating polypeptide. In some cases, the antibody may comprise a first and second polypeptide. In some cases, the first and second polypeptides may each comprise a heavy chain and a light chain. In some cases, the first and second molecules may each comprise a physiologically active fragment of a complement protein. The complement protein may be CR1, DAF, MCP, Crry, MAp44, MAp19, CD59, or factor H. In some cases, the albumin-to-creatine ratio may be measured in a biological sample from a subject. In some cases, the albumin-to-creatine ratio may be determined at some point in time after administration of the fusion protein construct of this disclosure or after administration of an equivalent fusion protein construct. In some cases, the albumin-to-creatine ratio may be compared between two or more biological samples from the same subject or from different subjects. In some cases, the biological sample may be a urine sample. In some cases, the subject may have one of the diseases disclosed in this disclosure. In some cases, the subject may not have one of the diseases. In some cases, the albumin-to-creatinine ratio of urine samples from subjects with a disease (e.g., renal disease) after administration of the fusion protein construct of this disclosure is lower than the albumin-to-creatine ratio of urine samples from subjects administered an equivalent fusion protein construct. In some cases, an equivalent fusion protein construct is identical to the fusion protein construct of this disclosure except that it does not contain an antibody or its antigen-binding fragment.

[0103] This disclosure provides a fusion protein construct comprising at least two components fused together: (i) a target-directing moiety that binds to a complement-related antigen, and (ii) a complement modifier. The components of the construct may be fused together by covalent or non-covalent interactions. The fusion construct protein may be engineered to optimize and adapt its target-binding valency depending on the target, conditions, and desired amount of complement modifier. For example, a fusion protein construct may be designed as divalent, trivalent, or tetravalent, each valency relating to the target-binding component or the complement modifier. When a fusion protein construct is expressed using valency, it is the sum of the valencies of each component. In some cases, the target-directing moiety may be divalent while the complement modifier is monovalent, resulting in a trivalent fusion protein construct. In other specific examples, the target-directing moiety may be monovalent or divalent while the complement modifier is divalent, resulting in a trivalent or tetravalent fusion construct, respectively.

[0104] In addition, the target-directing moiety in the fusion protein construct can specifically bind to one or more antigens, thereby allowing the fusion protein construct to be a bispecific, tripspecific, or multispecific molecule. The fusion protein constructs of this disclosure can bind to multiple targets, equipped with their improved specificity and increased valency. In certain examples, the fusion protein constructs may have additional beneficial properties, such as suitability for production and pharmaceutical composition, through their improved stability, low aggregation, pharmacokinetic and biological properties, or any combination thereof.

[0105] The target-directed moiety may be an antibody or its antigen-binding fragment that can specifically bind to one or more complement-related antigens, such as antigens locally presented at or near sites associated with complement activation. An example of a target-directed moiety may include an antibody or its antigen-binding fragment that is specific to a domain of a mammalian annexin protein. Alternatively, the target-directed moiety may include an antibody or its antigen-binding fragment that is specific to a phospholipid. The target-directed moiety may include an antibody or its antigen-binding fragment that is specific to a complement protein, such as a C3 fragment associated with complement activation, such as C3b, iC3b, C3d, and C3dg. The target-directed moiety may also be a bispecific or triplicate antibody or its antigen-binding fragment that is specific to any combination of the following targets: a domain of a mammalian annexin protein, a phospholipid, and a complement protein, such as a C3 fragment, such as C3d, iC3b, C3dg, C3a, C3b, C3c, or C3f. In addition... Furthermore, it is also intended that the target-directing moiety, which is polyvalent, for example trivalent or tetravalent, may be part of the fusion protein construct described herein.

[0106] Some examples of fusion protein constructs may be trivalent and bispecific, comprising (i) a bivalent antibody or its antigen-binding fragment (e.g., antibody Fab region), (ii) an antibody Fc domain, and (iii) a complement regulator fused to the antibody or its antigen-binding fragment or the Fc domain. The bivalent antibody or its antigen-binding fragment may be the target-directed portion of the trivalent bispecific fusion protein construct and may bind to two target molecules, such as the domains of mammalian annexin proteins (e.g., annexin IV, annexin-2), phospholipids, complement proteins, or fragments of complement proteins (e.g., iC3b, C3d, C3dg, C3a, C3b, C3c, or C3f). The complement regulator of the trivalent bispecific fusion protein construct may be an inhibitor of complement activation. Further variants of the trivalent bispecific fusion protein construct may include a) a first polypeptide monomer comprising a CH2 or CH3 domain of an antibody, wherein the domain has at least one amino acid modification resulting in a hole; b) a second polypeptide monomer comprising a CH2 or CH3 domain of an antibody, wherein the domain has at least one amino acid modification resulting in a knob; thereby the knob interacts with the hole during heterodimerization of the first and second polypeptide monomers; and a bispecific trivalent heterodimer polypeptide construct in which one of the first and second polypeptides is further fused to a complement regulator.

[0107] In another example, the fusion protein construct is a tetravalent construct for regulating complement activity, comprising (i) a bivalent antibody or its antigen-binding fragment and (ii) two complement modifiers. The bivalent antibody in the tetravalent fusion construct may be a full-length bivalent antibody or its antigen-binding fragment, such as a variable region fragment or a bivalent single-strand variable fragment. The two complement modifiers may be the same protein or different, and may form a bispecific tetravalent or tripspecific tetravalent fusion protein construct. In yet another example, the fusion protein construct is a monomer and includes one or more amino acid modifications that eliminate the disulfide bond that occurs when forming a homodimeric construct.

[0108] An antibody consists of four polypeptides: two heavy chains and two light chains. The antigen-binding portion of the antibody is formed by the light chain variable domain (VL) and the heavy chain variable domain (VH). At one end of these domains, six loops form the antigen-binding site, also known as the complementarity-determining region (CDR). Three CDRs are located on the VH domains (H1, H2, and H3), and the other three are located on the VL domains (L1, L2, and L3). During B cell development, somatic recombination known as V(D)J gene rearrangement forms a unique immunoglobulin region. The variable regions of the immunoglobulin heavy or light chain are encoded by different gene segments. The heavy chain is encoded by three segments called the variable (V), diverse (D), and binding (J) segments, while the light chain variable region is formed by only two segments, V and J. Recombination between one of multiple copies of the V, D, and J segments present in the genome can generate numerous antibody paratopes (also referred to herein as antigen-binding sites). The V segment encodes CDR1 and CDR2, but CDR3 is generated by a recombination event. During the immune response, further variability is introduced to the antigen-binding site through a process called somatic hypermutation (SHM). During this process, point mutations are introduced into the variable genes of the heavy and light chains, specifically the region encoding CDR. This additional variability allows for the selection and proliferation of B cells that express antibody variants with improved affinity for congener antigens. The vast majority of immunoglobulins are bivalent monospecific molecules that possess the same specificity in both arms, as they consist of two identical heavy chain polypeptides and two identical light chain polypeptides. However, very early in the development of hybridoma technology, hybrid hybridomas were created by fusion events between two hybridomas. It was recognized that such a combination could be produced (see, for example, MRSuresh et al., Methods Enzymol 1986;121:210-228). These "quadromas" express two different heavy chains and two different light chains, and therefore produce a wide variety of antibody species due to the random pairing of heavy and light chains. Among these different species, bispecific antibodies (bsAb) are produced, each possessing different specificity on each arm. Another naturally occurring exception is the IgG4 isotype immunoglobulin, which can undergo heavy chain exchange due to less stable dimerization mediated by the hinge region of its isotype (see, for example, van der Neut Kolfschoten M (See et al., Science. 2007 317(5844):1554-7). This exchange appears to occur in vivo, but its biological significance remains unclear. Monoclonal antibodies have emerged as a successful and attractive class of molecules for therapeutic intervention in several areas of human disease. However, targeting or neutralizing a single protein is not always sufficient to achieve efficacy in specific diseases, which limits the therapeutic use of monoclonal antibodies. It is becoming increasingly clear that neutralizing one component of a biological system is not sufficient to achieve efficacy in multiple indications. One solution to this problem is the co-administration of several monoclonal antibodies. However, this approach is complicated by regulatory issues if the antibodies used in combination have not been approved individually beforehand. Moreover, the co-administration method is also costly from a manufacturing standpoint. Therefore, there is a need for antibodies and therapeutics that enable targeting of multiple antigens with a single molecule.

[0109] This disclosure provides molecules comprising at least divalent, trivalent, and tetravalent fusion protein constructs that have enhanced therapeutic capabilities to address complement system-related diseases, disorders, or symptoms.

[0110] Complement regulators The complement system is a major effector of humoral and innate immunity. The complement system has three independent pathways for complement activation: the classical pathway, the alternative pathway, and the lectin pathway. While each of these three pathways differs in its initiation event, all three converge on the cleavage of complement component C3. Key to complement system activity is the covalent binding of complement C3 and / or C4, processed protein fragments derived from serum proteins, to the tissue site of complement activation. This unique property stems from the presence of a thioester bond in C3, which, when cleaved during C3 activation, converts C3 into a form called C3b, which can subsequently bind to cell and tissue-binding molecules via ester or amide bonds. Once covalently bound, C3b is rapidly processed into iC3b, C3dg, and C3d forms, each remaining covalently bound to the target tissue site. This process "marks" the tissue as being affected by inflammatory damage or other complement-related processes.

[0111] Complement can be activated by one of three pathways: classical, lectin, and alternative pathways. The classical pathway is activated when the complement protein C1q binds to antigen-antibody complexes, pentraxin, or apoptotic cells. Examples of pentraxin include C-reactive protein and serum amyloid-P component. The lectin pathway is initiated when carbohydrates bind to mannose-binding lectins, or when ficolin or collectin bind to carbohydrates or acetylated molecules.

[0112] The alternative pathway is activated on the surface of pathogens that neither express nor contain complement inhibitors. This results from a process called "tickover" of C3, which spontaneously occurs with the interaction of conformationally altered C3 and factor B, leading to the fixation of active C3b on or on the pathogen. The alternative pathway involves specific antibodies activating the endogenous regulatory mechanism of IgA It can also be initiated when it is blocked by a containing immune complex or when the expression of complement regulatory proteins is reduced. In addition, the alternative pathway is activated by a mechanism called the “amplification loop” when C3b deposited on the target via the classical or lectin pathway or, of course, the tick-over process itself, binds to factor B. See Muller-Eberhard (1988) Ann. Rev. Biochem. 57:321. For example, Holers and colleagues showed that the alternative pathway is amplified at the site of local injury when inflammatory cells are recruited following initial complement activation. See Girardi et al., J. Clin. Invest. 2003, 112:1644. Then, dramatic complement amplification by the alternative pathway occurs, either by a mechanism involving further development of complement-fixing injured cells or local synthesis of alternative pathway components, or more likely by pre-formed C3 and properdin-bearing infiltrating inflammatory cells specifically and significantly increasing activation at that site.

[0113] Alternate pathway amplification is initiated when circulating factor B binds to activated C3b. This complex is then cleaved by circulating factor D to produce the enzymatically active C3-converting enzyme complex, C3bBb. C3bBb further cleaves C3 to produce C3b, which promotes inflammation and further amplifies the activation process, creating a positive feedback loop. Factor H is a key regulator (inhibitor) of the alternative complement pathway activation and initiation mechanism, competing with factor B for binding to conformally altered C3 in the tick-over mechanism and to C3b in the amplification loop. Binding of C3b to factor H also leads to degradation of C3b by factor I into the inactive form iC3b (also called C3bi), thus further inhibiting complement activation. Factor H regulates complement in the fluid phase while circulating in plasma at concentrations of approximately 400–600 μg / ml. However, its binding to cells is a regulated phenomenon enhanced by the presence of negatively charged surfaces and fixed C3b, iC3b, C3dg, or C3d. See Jozsi et al., Histopathol. (2004) 19:251-258.

[0114] Complement activation, C3 fragment fixation, and complement-mediated inflammation are involved in the pathogenesis and progression of many diseases. Downregulation of complement activation is associated with, for example, systemic lupus erythematosus and glomerulonephritis (Y. Wang et al., Proc. Nat'l Acad. Sci. USA (1996) 93:8563-8568), rheumatoid arthritis (Y. Wang et al., Proc. Nat'l Acad. Sci. USA (1995) 92:8955-8959), cardiopulmonary bypass and hemodialysis (CSRinder, J. Clin. Invest. (1995) 96:1564-1572), hyperacute rejection in organ transplantation (T. J. Kroshus et al., Transplantation (1995) 60:1194-1202), and myocardial infarction (J. W. Homeister et al., J. Immunol. (1993) 150:1055-1064, HF Weisman et al. It has been shown to be effective in treating several diseases, including ischemia / reperfusion injury (EAAmsterdam et al., Am.J.Physiol.(1995)268:H448-H457), antibody-mediated allograft rejection, such as in the kidney (JBColvin, J.Am.Soc.Nephrol.(2007)18(4):1046-56), and adult respiratory distress syndrome (R.Rabinovici et al., J.Immunol.(1992)149:1744-1750), in animal models and in vitro studies. Furthermore, other inflammatory conditions and autoimmune / immune complex diseases, including but not limited to burns, severe asthma, anaphylactic shock, enteritis, urticaria, angioedema, vasculitis, multiple sclerosis, myasthenia gravis, myocarditis, membranoproliferative glomerulonephritis, atypical hemolytic uremic syndrome, Sjögren's syndrome, renal and pulmonary ischemia / reperfusion, and other organ-specific inflammatory disorders, are also closely associated with complement activation (BPMorgan.Eur.J.Clin.Invest.(1994)24:219-228). Currently, local tissue C3 activation and inflammation It is unclear whether complement activation is essential for the onset and injury of all diseases involving damage, but nevertheless, C3 fragment fixation is frequently found as an associated event.

[0115] Endogenous membrane-bound proteins that regulate alternative pathway activation include catalytic degeneration (DAF / CD55), membrane cofactor protein (MCP / CD46), and complement receptor 1 (CR1). Other endogenous proteins that regulate alternative pathway activation include factor H, a 155 kDa circulating glycoprotein that regulates alternative pathway activation in the fluid phase and on tissue surfaces. See J. Alexander et al., Mol. Immunol. (2006) 44:123-132. Unregulated alternative pathway activation is thought to be associated with the development of a diverse range of diseases, including age-related macular degeneration (AMD), atypical hemolytic uremic syndrome (aHUS), membranoproliferative glomerulonephritis type II (MPGN II), asthma, and renal ischemia / reperfusion (I / R) injury. See J. M. Hurman et al., J. Immunol. (2006) 176:1305-1310. Damage to host tissue via alternative pathways suggests inadequate local control of these alternative pathways by the target tissue. In fact, mutations in CRP are a strong risk factor for aHUS (MCPickering et al., J.Exp.Med.(2007)204:1249-1256) and MPGN II (RJ.Smith et al., J.Am.Soc.Nephrol.(2007)18:2447-2456), and functional polymorphism in factor H, a regulator of circulating alternative pathways, is associated with the progression of AMD (RJKlein et al., Science(2005)308:385-389, AOEdwards et al., Science(2005)308:421-424, JLHaines et al., Science(2005)308:419-421, GSHageman et al., Proc.Nat'l Acad.Sci This has been demonstrated in recent studies (USA (2005) 102:7227-7232)).

[0116] Ischemic acute kidney injury (AKI) is associated with the activation of alternative pathways on the basolateral surface of damaged tubular cells in rodents (JMThurman et al., J.Immunol. (2003) 170:1517-1523, JMThurman et al., Am.Soc.Nephrol. (2006) 17:707-715) and humans (JMThurman et al., Kidney Int. (2005) 67:524-530). In mice, the gene / protein y (Crry, a rodent analog of human MCP and CR1) related to complement receptor 1 has been found to be the only C-reactive protein (CRP) expressed by proximal tubular epithelial cells, and ischemia / reperfusion has been shown to cause a decrease in the expression of this protein on the surface. See JMThurman et al., J.Clin.Invest. (2006) 116:357-368. Mice with a congenital deficiency of Crry (Crry+ / -) are more susceptible to ischemic acute renal failure (Id.) than wild-type controls, highlighting the importance of basal Crry in regulating this alternative pathway on the surface. It remains unclear whether polymorphism or mutation in CRP may lead to an increased risk of AKI progression in humans. Nevertheless, unregulated activation of alternative pathways in the context of reduced surface Crry suggests that the protective capacity of circulating factor H against the surface of hypoxic tubular epithelial cells is limited.

[0117] Factor H circulates at high concentrations (over 400-600 μg / ml) and is a potent inhibitor of the alternative complement pathway. See J.A. Alexander et al., Mol.Immunol.(2006)44:123-132. However, inhibitory effect of factor H on the alternative pathway at the cell surface requires that it binds properly to that surface. Several regions in the factor H protein bind to anionic surfaces, such as heparin sulfate or sialic acid-rich surfaces, and to C3b on the surface. See S.Meri et al., Proc.Nat'l Acad.Sci USA(1990)87:3982-3986, MK See Pangburn et al., Immunol. (2000) 164:4742-4751. Activation of alternative pathways on specific surfaces is strongly influenced by the affinity of factor H to that surface. The polymorphisms and mutations associated with AMD and aHUS, respectively, most often involve the region of factor H required for anionic surface binding, rather than the complement regulatory region. See MCPickering et al., J.Exp.Med. (2007) 204:1249-1256 and APSjoberg et al., J.Biol.Chem. (2007) 282:10894-10900. Thus, certain tissues or cell types require factor H to regulate the activation of alternative pathways on their surfaces. Different binding regions of the factor H protein may be required for complement regulation in those tissues or cell types. In some cases, the binding of factor H to the surface of a particular tissue may be influenced by other proteins. Identifying putative tissue-specific binding partners for factor H may provide potential mechanisms for modulating, i.e., stimulating or inhibiting the activity of alternative complement pathways in different tissues.

[0118] Thus, the complement modulators of this disclosure may include complement modulator proteins or complement modulator polypeptides (e.g., complement inhibitor polypeptides), such as membrane cofactor proteins (MCPs) (SEQ ID NO: 1) (UniProtKB / Swiss-Prot accession number P15529), degrading factors (DAFs), CD59, Crry, CR1, CR2, factor H, or their variants or fragments. Further complement modulators may include anti-C5 antibodies, eculizumab, pexerizumab, anti-C3b antibodies, anti-C6 antibodies, anti-C7 antibodies, anti-C8 antibodies, anti-C9 antibodies, anti-factor B antibodies, anti-MASP antibodies, anti-factor D antibodies, and anti-properdin antibodies, anti-MBL antibodies, factor I, linear peptides, cyclic peptides, compstatin or its analogues, N-acetylaspartylglutamic acid (NAAGA), and any of the above-mentioned physiologically active fragments. In some cases, the complement inhibitor may be a human complement inhibitor (e.g., human MCP, human DAF, human CD59, human CR1, human H factor, human Map44, human Map19, or another complement inhibitor derived from humans). In further examples, the complement inhibitor may be a mammalian complement inhibitor (e.g., mouse DAF, mouse CD59 (also known as isoform A), mouse CD59 isoform B, mouse Crry, mouse H factor, mouse Map44, mouse Map19, or another complement inhibitor derived from mice). Thus, the polypeptide may be a human polypeptide or a polypeptide of a non-human species. For example, a complement regulator polypeptide may be from a non-human primate (e.g., orangutan, chimpanzee, rhesus monkey, gorilla, lemur or gibbon), horse, cattle, pig, sheep, goat, dog, cattle, or rodent (e.g., mouse, rabbit, hamster, gerbil, guinea pig or rat).

[0119] The complement regulator of the fusion protein construct may be a mutant complement regulator. A mutant complement regulator polypeptide may have one or more amino acid substitutions compared to the corresponding wild-type sequence (e.g., 59 or fewer, 58, 57, 56, 55, 54, 53, 52, 51, 50, 49, 48, 47, 46, 45, 44, 43, 42, 41, 40, 39, 38, 37, The mutant polypeptide may contain 36, 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 amino acid substitution. The amino acid substitution may be conserved, non-conserved, or a mixture of both. In some embodiments, the mutant polypeptide may contain one or more deletions or additions, or a combination of one or more deletions, additions, and substitutions. In some examples, the mutant complement regulator polypeptide may contain one or more amino acid deletions, additions, or substitutions per 100 amino acids of the polypeptide. In some cases where complement-modulating polypeptides contain short consensus repeats (SCRs), the mutant polypeptide is, The complement-modulating polypeptide SCR may contain no substitutions, deletions, or additions whatsoever.

[0120] A mutant complement regulator polypeptide may contain an amino acid sequence that is at least 70% (e.g., at least 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%) identical to the corresponding wild-type amino acid sequence. The functional fragments of complement regulator polypeptides or mutant polypeptides described herein are shorter than full-length polypeptides. A mutant complement regulator polypeptide may contain an amino acid sequence that is at least 90%, 95%, or 98% identical to one or more functional (bioactive) fragments or domains of complement regulator polypeptides identified herein. Mutant polypeptides and functional fragments of wild-type proteins or mutants retain at least 50% (e.g., at least 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99)% of the complement regulatory activity of the corresponding wild-type polypeptide.

[0121] Mutant complement regulator polypeptides, or functional fragments of complement regulator polypeptides, may exhibit a complement activity-modulating ability exceeding 100% compared to the corresponding wild-type protein. Methods for detecting and / or quantifying complement activity are known in the art and are described herein.

[0122] Other examples of sequences useful as complement moduloproliferators in this disclosure include one or more short consensus repeat (SCR) domains derived from one or more of the following complement-related proteins: factor H, complement receptor 1, complement receptor 2, factor B, DAF, etc. In some cases, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17 SCR regions are incorporated into the complement moduloproliferators described herein. In some examples where the complement moduloproliferator polypeptide includes “membrane cofactor protein,” “MCP,” or “CD46,” this means a broadly distributed C3b / C4b-binding cell surface glycoprotein, including its homologs, that can inhibit complement activation in host cells and act as a cofactor for factor I-mediated cleavage of C3b and C4b. MCP may belong to a family known as complement activation moduloproliferators (“RCAs”). Family members may share specific structural features, including varying numbers of short consensus repeat (SCR) domains typically 60–70 amino acids in length. MCPs may comprise four SCRs starting from their amino terminus, a serine / threonine / proline-rich region, a region of unknown function, a transmembrane hydrophobic domain, a cytoplasmic anchor, and a cytoplasmic tail. It is understood that the peptides, polypeptides, and proteins of this disclosure exhibit species and lineage diversity, and that human MCPs or their bioactive fragments may encompass all species and lineage diversity.

[0123] Sequence ID 1 represents an exemplary sequence of a full-length human MCP (see, for example, UniProtKB / Swiss-Prot accession number P15529). Amino acids 1-34 may correspond to the signal peptide, amino acids 35-343 may correspond to the extracellular domain, amino acids 344-366 may correspond to the transmembrane domain, and amino acids 367-392 may correspond to the cytoplasmic domain. In the extracellular domain, amino acids 35-96 may correspond to SCR1, amino acids 97-159 may correspond to SCR2, amino acids 160-225 may correspond to SCR3, amino acids 226-285 may correspond to SCR4, and amino acids 302-326 may correspond to the serine / threonine-rich domain. The peptides, polypeptides and proteins disclosed herein exhibit species and lineage diversity, as well as MCP or its physiological activity. It is understood that the fragments may encompass diversity across all species and lineages. As used herein, the term “bioactive” fragment of MCP may refer to any soluble fragment lacking the cytoplasmic and transmembrane domains, including fragments that are essentially derived from or consist of fragments that have some or all of the complement inhibitory activity of the full-length human MCP protein, with or without the serine / threonine-rich domain, and contain one, two, three, or four SCR domains. In some embodiments, the complement inhibitory portion includes the full-length human MCP (amino acids 35-392 of SEQ ID NO: 1), the extracellular domain of human MCP (amino acids 35-343 of SEQ ID NO: 1), or SCR1-4 of human MCP (amino acids 35-285 of SEQ ID NO: 1).

[0124] In some cases, complement regulatory polypeptides include complement-aggravating factors (DAF / CD55), also known as CD55 (SEQ ID NO: 2 and SEQ ID NO: 3), which are membrane-bound glycoproteins with a molecular weight of approximately 70 kilodaltons (kDa) that inhibit complement activation in host cells. Like several other complement regulatory proteins, DAFs contain several repeating motifs of approximately 60 amino acids called short consensus repeats (SCRs).

[0125] As used herein, the terms “Disintegration Promoter,” “DAF,” or “CD55” refer to a 70 kilodalton (”kDa”) membrane glycoprotein comprising four short consensus repeat (SCR) domains followed by a highly O-glycosylated serine / threonine-rich C-terminal domain that lifts molecules from the membrane surface, followed by a glycosylphosphatidylinositol (”GPI”) anchor. DAF protects the cell surface from complement activation by dissociating membrane-bound C3-converting enzymes required for complement protein C3 cleavage and complement cascade amplification. DAF prevents or promotes the aggregation of both C3 and C5-converting enzymes of the alternative and classical complement pathways.

[0126] Sequence ID 2 represents an exemplary sequence of a full-length human DAF (see, for example, UniProtKB / Swiss-Prot accession number P08173), and Sequence ID 3 represents an exemplary sequence of a full-length mouse DAF (see, for example, UniProtKB / Swiss-Prot accession number Q61475). In the human DAF sequence, amino acids 1-34 may correspond to a signal peptide, amino acids 35-353 may appear in the mature protein, and amino acids 354-381 may be removed from the polypeptide post-translation. In the mature protein, amino acids 35-96 may correspond to SCR1, amino acids 96-160 may correspond to SCR2, amino acids 161-222 may correspond to SCR3, amino acids 223-285 may correspond to SCR4, and amino acids 287-353 may correspond to an O-glycosylated serine / threonine-rich domain. The GPI anchor can bind to human DAF at serine position 353. In the mouse DAF sequence, amino acids 1-34 may correspond to a signal peptide, amino acids 35-362 may appear in the mature protein, and amino acids 363-390 may be removed from the polypeptide post-translation. In the mature protein, amino acids 35-96 may correspond to SCR1, amino acids 97-160 may correspond to SCR2, amino acids 161-222 may correspond to SCR3, amino acids 223-286 may correspond to SCR4, and amino acids 288-362 may correspond to an O-glycosylated serine / threonine-rich domain. The GPI anchor can bind to mouse DAF at serine position 362. It is understood that there is species and lineage diversity in the peptides, polypeptides and proteins of this disclosure, and that DAFs or their bioactive fragments may encompass all species and lineage diversity. As used herein, the term “bioactive” fragment of DAF may refer to any fragment of DAF lacking the GPI anchor, the amino acid to which it is bound (e.g., Ser-353), or both, including, but not limited to, a full-length DAF protein containing one, two, three, or four SCR domains, which have some or all of the complement inhibitory activity of the full-length DAF protein, with or without the O-glycosylated serine / threonine-rich domain, and which is essentially derived from or consists of a full-length DAF protein. This also includes any fragment of the DAF protein.

[0127] Sequence ID 4 represents an exemplary sequence of full-length human CD59 (see, for example, UniProtKB / Swiss-Prot accession number P13987), Sequence ID 5 represents an exemplary sequence of full-length mouse CD59 isoform A (see, for example, UniProtKB / Swiss-Prot accession number O55186), and Sequence ID 6 represents an exemplary sequence of full-length mouse CD59 isoform B (see, for example, UniProtKB / Swiss-Prot accession number P58019). In the human CD59 sequence, amino acids 1-25 of Sequence ID 4 may correspond to the leader peptide, amino acids 26-102 of Sequence ID 4 may correspond to the mature protein, and amino acids 103-128 of Sequence ID 4 may be removed post-translation. The GPI anchor may bind to CD59 at asparagine at position 102 of Sequence ID 4. In mouse CD59 isoform A, amino acids 1-23 of SEQ ID NO: 5 may correspond to the leader peptide, amino acids 24-96 of SEQ ID NO: 5 may correspond to the mature protein, and amino acids 97-123 of SEQ ID NO: 5 may be removed post-translation. The GPI anchor can bind to CD59 at serine at position 96 of SEQ ID NO: 5. In mouse CD59 sequence isoform B, amino acids 1-23 of SEQ ID NO: 6 may correspond to the leader peptide, amino acids 24-104 of SEQ ID NO: 6 may correspond to the mature protein, and amino acids 105-129 of SEQ ID NO: 6 may be removed post-translation. The GPI anchor can bind to CD59 at asparagine at position 104 of SEQ ID NO: 6. It is understood that the peptides, polypeptides, and proteins of this disclosure exhibit species and lineage diversity, and that CD59 or its bioactive fragments may encompass all species and lineage diversity.As used herein, the term “bioactive” fragment of human CD59 may refer to any fragment of human CD59 lacking the GPI anchor, the amino acid to which it is bound (e.g., Asn-102), or both, including any fragment of the full-length human CD59 protein having some or all of the complement inhibitory activity of the full-length CD59 protein; the term “bioactive” fragment of mouse CD59 may refer to any fragment of mouse CD59 isoform A or isoform B lacking the GPI anchor and / or the amino acid to which it is bound (e.g., Ser-96 in isoform A, or Asp-104 in isoform B), including any fragment of either full-length mouse CD59 protein isoform having some or all of the complement inhibitory activity of the full-length CD59 protein.

[0128] Sequence ID 7 represents an exemplary sequence of the full-length mouse Crry protein. Amino acids 1-40 may correspond to the leader peptide, and amino acids 41-483 of Sequence ID 7 may correspond to the mature protein, which includes amino acids 41-405 of Sequence ID 7 that may correspond to the extracellular domain, amino acids 406-426 that may correspond to the transmembrane domain, and amino acids 427-483 that may correspond to the cytoplasmic domain. In the extracellular domain, amino acids 83-143 of Sequence ID 7 may correspond to SCR1, amino acids 144-205 of Sequence ID 7 may correspond to SCR2, amino acids 206-276 of Sequence ID 7 may correspond to SCR3, amino acids 277-338 of Sequence ID 7 may correspond to SCR4, and amino acids 339-400 of Sequence ID 7 may correspond to SCR5. It is understood that there is species and strain diversity in the peptides, polypeptides and proteins of this disclosure, and that the mouse Crry protein or its bioactive fragments may encompass all species and strain diversity. As used herein, the term “bioactive” fragment of the mouse Crry protein may refer to any soluble fragment of mouse Crry lacking the transmembrane domain and cytoplasmic domain, including any fragment of the full-length mouse Crry protein having some or all of the complement inhibitory activity of the full-length Crry protein, as well as fragments containing, essentially derived from, or consisting of 1, 2, 3, 4, or 5 SCR domains.

[0129] As used herein, the terms "complement receptor 1," "CR1," or "CD35" are used in this specification. The term can refer to a human gene encoding a 2039-amino acid protein with a predicted molecular weight of 220 kilodaltons ("kDa"), including its homologs. The gene can, in principle, be expressed on erythrocytes, monocytes, neutrophils, and B cells, but may also be present on some T lymphocytes, mast cells, and glomerular podocytes. Typically, 100 to 1000 copies of the CR1 protein may be expressed per cell. CR1 may be the primary system for the processing and clearance of complement-opsonized immune complexes. CR1 can negatively regulate the complement cascade, mediate immunoadhesion and phagocytosis, and inhibit all complement pathways. The full-length CR1 protein may consist of a 42-amino acid signal peptide, a 1930-amino acid extracellular domain, a 25-amino acid transmembrane domain, and a 43-amino acid C-terminal cytoplasmic domain. The extracellular domain of CR1 may contain 25 potential N-glycosylation signal sequences and 30 short consensus repeat ("SCR") domains, each 60–70 amino acids long, also known as complement regulatory protein (CCP) repeats or success domains. Sequence homology between SCRs can range from 60–99 percent. The 30 SCR domains may be further classified into four longer regions called long homologous repeats ("LHR"), each encoding a segment of the CR1 protein of approximately 45 kDa, and are designated LHR-A, -B, -C, and -D (see, e.g., Krych-Goldberg et al., 274(44):31160–31168, 1999). The first three LHRs may each contain seven SCR domains, while LHR-D may contain nine SCR domains. The active sites on the extracellular domain of the CR1 protein may include C4b binding sites in SCR1-3 containing amino acids 42-234 with lower affinity for C3b, C3b binding sites in SCR8-11 containing amino acids 490-745 with lower affinity for C4b, C3b binding sites in SCR15-18 containing amino acids 940-1196 with lower affinity for C4b, and C1q binding sites in SCR22-28 containing amino acids 1394-1842.

[0130] Sequence ID 8 represents an exemplary sequence of full-length human CR1 (see, for example, UniProtKB / Swiss-Prot accession number P17927). Amino acids 1-41 may correspond to a signal peptide, and amino acids 42-2039 may correspond to a mature protein containing amino acids 42-1971 which may correspond to an extracellular domain, amino acids 1972-1996 which may correspond to a transmembrane domain, and amino acids 1997-2039 which may correspond to a cytoplasmic domain. In the extracellular domain, amino acids 42-101 may correspond to SCR1, 102-163 to SCR2, 164-234 to SCR3, 236-295 to SCR4, 295-355 to SCR5, 356-418 to SCR6, 419-489 to SCR7, and 491-551 to SCR8. Amino acids 552-613 may correspond to SCR9, amino acids 614-684 may correspond to SCR10, amino acids 686-745 may correspond to SCR11, amino acids 745-805 may correspond to SCR12, amino acids 806-868 may correspond to SCR13, amino acids 869-939 may correspond to SCR14, amino acids 941-1001 may correspond to SCR15, and amino acids 1002-1063 are SC Amino acids 1064-1134 can correspond to SCR17, amino acids 1136-1195 can correspond to SCR18, amino acids 1195-1255 can correspond to SCR19, amino acids 1256-1318 can correspond to SCR20, amino acids 1319-1389 can correspond to SCR21, amino acids 1394-1454 can correspond to SCR22, and amino acids 1455-1516 can correspond to SCR23. The peptides, polypeptides, and proteins of this disclosure exhibit species and lineage diversity. It is understood that such fragments exist, and that the CR1 protein or its bioactive fragments may encompass diversity across all species and lineages. As used herein, the term “bioactive” fragment of the CR1 protein may refer to any soluble fragment of CR1 lacking the transmembrane domain and cytoplasmic domain, including any fragment of the full-length CR1 protein having some or all of the complement inhibitory activity of the full-length CR1 protein, as well as fragments containing, essentially derived from, or consisting of 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 SCR domains. Functional fragments may include SCR1 and 2; SCR1, 2, 3 and 4; SCR1, 2, 3, 4, 5, 6, and 7; SCR1, 2, 3, 4, 5, 6, 7, 8, 9 and 10 ("CR1 1-10"); SCR6, 7, 8, 9, 10, 11 and 12; SCR8 and 9; SCR8, 9, 10 and 11; SCR8, 9, 10, 11, 12, 13 and 14; SCR15 and 16; SCR12, 13, 14, 15, 16 and 17; SCR15, 16, 17, 18 and 19; SCR1-17 ("CR1 1-17"); SCR1-23, SCR1-28. An exemplary mutant polypeptide comprises at least three SCRs in each of domains A and B; at least three SCRs in each of domains A, B, and C; at least the first three SCRs in domains A, B, and C; or an amino acid sequence that is at least 90% identical to any of the above.

[0131] As used herein, the terms “complement H factor,” “H factor,” or “FH” may refer to complement H factor, a single-chain polypeptide plasma glycoprotein, including its homologs. The protein may consist of 20 conserved short consensus repeat (SCR) domains of approximately 60 amino acids, arranged in a continuous manner resembling beads on a thread, separated by short linker sequences of 2–6 amino acids each. Factor H can bind to C3b and promote the breakdown of alternative pathway C3-converting enzymes (C3bBb) and alternative pathway C5-converting enzymes (C3bBb3b), and may act as a cofactor for the proteolytic inactivation of C3b. In the presence of factor H, proteolysis by factor I may result in the cleavage and inactivation of C3b. Factor H may have at least three distinct binding domains to C3b, which may be located within any one of SCR1–20, SCR1–4, SCR5–8, and SCR19–20. Each domain can bind to a distinct region within the C3b protein: the N-terminal region can bind to native C3b, the second region located in the intermediate region of factor H can bind to the C3c fragment, and the regions located within SCR19 and 20 can bind to the C3d region. In addition, factor H may also contain a heparin binding site, which can be located within SCR7, SCR5-12, and SCR20 of factor H, and may overlap with those of the C3b binding site. Structural and functional analyses have shown that the domain for complement inhibitory activity of factor H can be located within the first four N-terminal SCR domains.

[0132] Sequence ID 9 represents an exemplary amino acid sequence of the full-length human factor H protein (see, for example, UniProtKB / Swiss-Prot accession number P08603), and Sequence ID 10 represents an exemplary amino acid sequence of the full-length mouse factor H protein (see, for example, UniProtKB / Swiss-Prot accession number P06909). In the human factor H sequence, amino acids 1-18 of Sequence ID 9 may correspond to a signal peptide, and amino acids 19-1231 of Sequence ID 9 may correspond to a mature protein. Within that protein, amino acids 21-80 of Sequence ID 9 may correspond to SCR1, amino acids 85-141 may correspond to SCR2, amino acids 146-205 may correspond to SCR3, amino acids 210-262 may correspond to SCR4, and amino acids 267-320 may correspond to SCR5. In the mouse H factor sequence, amino acids 1-18 of SEQ ID NO: 10 may correspond to a signal peptide, and amino acids 19-1234 of SEQ ID NO: 10 may correspond to a mature protein. Within that protein, amino acids 19-82 of SEQ ID NO: 10 may correspond to SCR1, and amino acids 83-143 of SEQ ID NO: 10 may correspond to SCR2. For example, amino acids 144-207 of SEQ ID NO: 10 may correspond to SCR3, amino acids 208-264 of SEQ ID NO: 10 may correspond to SCR4, and amino acids 265-322 of SEQ ID NO: 10 may correspond to SCR5. It is understood that the peptides, polypeptides, and proteins of this disclosure exhibit species and lineage diversity, and that H factor or its bioactive fragments may encompass diversity across all species and lineages. As used herein, the term “bioactive” fragment of H factor may refer to any portion of the H factor protein having some or all of the complement inhibitory activity of the full-length H factor protein, and may include, but is not limited to, H factor fragments including SCR1-4, SCR1-5, SCR1-8, SCR1-18, SCR19-20, or any naturally occurring homologs or fragments thereof of H factor as described below. In some examples of fusion protein constructs, the bioactive fragment of factor H may have one or more of the following properties: (1) binding to C-reactive protein (CRP), (2) binding to C3b and / or its fragments, (3) binding to heparin, (4) binding to sialic acid, (5) binding to the surface of endothelial cells, (6) binding to integrin receptors on cells, (7) binding to pathogens, (8) C3b cofactor activity, (9) C3 and C5 alternative pathway-converting enzyme disruption-promoting activity, and (10) inhibition of alternative complement pathways.

[0133] In some examples of fusion protein constructs, the complement regulator portion of the construct may include a complement inhibitor or a bioactive fragment thereof. In some examples of fusion protein constructs, the complement inhibitor may be selected from human MCP, human DAF, mouse DAF, human CD59, mouse CD59 isoform A, mouse CD59 isoform B, mouse Crry protein, human CR1, human factor H, mouse factor H, its bioactive fragment, and its variants.

[0134] In some cases, the complement inhibitory portion of the fusion protein construct may contain full-length human MCP (SEQ ID NO: 1). In other cases, the complement inhibitory portion of the fusion protein construct may contain a bioactive fragment of human MCP (SEQ ID NO: 1). In other cases, the bioactive fragment of human MCP may be selected from SCR1-4 (amino acids 35-285 of SEQ ID NO: 1), SCR1-4 and serine / threonine-rich domains (amino acids 35-326 of SEQ ID NO: 1), and the extracellular domain of MCP (amino acids 35-343 of SEQ ID NO: 1), as well as any combination thereof.

[0135] In some cases, the complement inhibitory portion of the fusion protein construct may contain a full-length human DAF. In some cases, the complement inhibitory portion of the fusion protein construct may contain a bioactive fragment of a human DAF (SEQ ID NO: 2). In some cases, the bioactive fragment of the human DAF may be selected from SCR1-4 (amino acids 25-285 of SEQ ID NO: 2), SCR1-4 and the O-glycosylated serine / threonine-rich domain (amino acids 25-353 of SEQ ID NO: 2), and any combination thereof. In some cases, the complement inhibitory portion of the construct may contain a full-length mouse DAF (SEQ ID NO: 3). In some cases, the complement inhibitory portion of the construct may contain a bioactive fragment of a mouse DAF. In some cases, the bioactive fragment of the mouse DAF may be selected from SCR1-4 (amino acids 35-286 of SEQ ID NO: 3), SCR1-4 and the O-glycosylated serine / threonine-rich domain (amino acids 35-362 of SEQ ID NO: 3), and any combination thereof.

[0136] In some cases, the complement inhibitory portion of the fusion protein construct may contain full-length human CR1 (SEQ ID NO: 8). In some cases, the complement inhibitory portion of the fusion protein construct may contain a bioactive fragment of human CR1 (SEQ ID NO: 8). In some cases, the bioactive fragment of human CR1 may be SCR1 (amino acids 42-101 of SEQ ID NO: 8), SCR2 (amino acids 102-163 of SEQ ID NO: 8), SCR3 (amino acids 164-234 of SEQ ID NO: 8), SCR4 (amino acids 236-295 of SEQ ID NO: 8), SCR5 (amino acids 295-355 of SEQ ID NO: 8), SCR6 (amino acids 356-418 of SEQ ID NO: 8), SCR7 (amino acids 419-489 of SEQ ID NO: 8), SCR8 (amino acids 491-551 of SEQ ID NO: 8), SCR9 (amino acids 552-613 of SEQ ID NO: 8), SCR10 (amino acids 614-684 of SEQ ID NO: 8), SCR11 (amino acids 686-745 of SEQ ID NO: 8), SCR12 (amino acids 745-805 of SEQ ID NO: 8), SCR13 (amino acids 806-868 of SEQ ID NO: 8), SCR14 (amino acids 869-939 of SEQ ID NO: 8), SCR15 (amino acids 941-1001 of SEQ ID NO: 8), SCR16 (amino acids 1002-1063 of SEQ ID NO: 8), SCR17 (amino acids 1064-1134 of SEQ ID NO: 8), SCR18 (amino acids 1136-1195 of SEQ ID NO: 8), SCR19 (amino acids 1195-1255 of SEQ ID NO: 8), SCR20 (amino acids 1256-1318 of SEQ ID NO: 8), SCR21 (amino acids 686-745 of SEQ ID NO: 8) These may be amino acids 1319-1389, SCR22 (amino acids 1394-1454 of SEQ ID NO: 8), SCR23 (amino acids 1455-1516 of SEQ ID NO: 8), SCR24 (amino acids 1517-1587 of SEQ ID NO: 8), SCR25 (amino acids 1589-1648 of SEQ ID NO: 8), SCR26 (amino acids 1648-1708 of SEQ ID NO: 8), SCR27 (amino acids 1709-1771 of SEQ ID NO: 8), SCR28 (amino acids 1772-1842 of SEQ ID NO: 8), SCR29 (amino acids 1846-1906 of SEQ ID NO: 8), SCR30 (amino acids 1907-1967 of SEQ ID NO: 8), or any combination thereof.

[0137] In some cases, the complement inhibitory portion of the fusion protein construct may contain full-length human factor H (SEQ ID NO: 9). In some cases, the complement inhibitory portion of the fusion protein construct may contain a bioactive fragment of human factor H (SEQ ID NO: 9). In some cases, the complement inhibitory portion of the fusion protein construct may contain full-length mouse factor H (SEQ ID NO: 10). In some cases, the complement inhibitory portion of the fusion protein construct may contain a bioactive fragment of mouse factor H (SEQ ID NO: 10). The bioactive fragment of factor H may include SCR1-4, SCR1-5, SCR1-8, SCR1-18, SCR19-20 of factor H, or any homolog or fragment thereof of naturally occurring factor H, or any combination thereof.

[0138] Targeting part The target-directed portion of a multivalent construct can be responsible for the targeted delivery of a complement system modifier to the site of action, such as the site of complement activation. The complement modifier may have therapeutic activity, such as specifically inhibiting complement activation. Therefore, the multivalent construct described herein as a whole has a dual function: binding to an epitope recognized by the antibody described herein and exerting therapeutic activity by inhibiting complement activation. The target-directed portion may be a human, mouse, humanized or camelized antibody, or an antigen-binding fragment thereof.

[0139] The epitopes recognized by the antibody or its antigen-binding fragments may be domains of mammalian annexin proteins, phospholipids, such as one or more of the C2 antibody-reactive phospholipids described below, or complement proteins, such as C3d, C3 fragments (e.g., deposited C3 fragments - C3b, iC3b, C3d, C3dg; free or undeposited C3 fragments - C3a, C3b, C3c, or C3f).

[0140] An antibody or its antigen-binding fragment may specifically bind to the domain of the mammalian annexin protein. Annexin is Ca 2+ The binding site is the same as most other Ca 2+ Calcium (Ca) is different from binding proteins. 2+) and a family of phospholipid-binding proteins. The Ca family of annexins 2+ The binding site has a unique structural configuration that allows annexin family members to be reversibly tethered to the outer surface of the membrane of cells and / or organelles. Annexin family members have a conserved Ca characteristic. 2+ The binding site is located in the annexin core domain and contains four annexin repeats, each 70 amino acids long. The annexin core domain is α-helical, forming a small, rounded disc with a convex surface that is Ca 2+ It also includes a membrane-binding site, with the concave side facing outward towards the membrane, making it available for other types of interactions. Annexin family members also typically have an amino-terminal domain of indeterminate length preceding the annexin core domain, which varies in sequence and structure. In vertebrates, this includes annexin IV and annexin 2. The properties of the 12 annexin subfamilies have been elucidated, and each has a different amino-terminal domain and Ca at different positions. 2+ It has different splice variants in which the binding site is located. An antibody or its antigen-binding fragment that specifically binds to a domain in annexin IV protein or recognizes an epitope within it may be a B4 mAb or an antigen-binding fragment derived from a B4 mAb. An antibody or its antigen-binding fragment that specifically binds to a domain in annexin IV protein (e.g., human annexin IV protein) or recognizes an epitope within it may be a B4 mAb or an antigen-binding fragment derived from a B4 mAb, as described in Kulik et al., J Immunol. 182(9):5363 (2009). Exemplary CDRs of B4 mAbs are shown in SEQ ID NOs: 11-16. The target-directing portion may further be an antibody or its antigen-binding fragment that specifically binds to or recognizes an epitope in annexin 2 protein (e.g., human annexin 2 protein).

[0141] Antibodies or their antigen-binding fragments may also specifically bind to phospholipids (e.g., phosphatidylethanolamine (PE), cardiolipin (CL), phosphatidylcholine (PC), phosphatidylinositol, phosphatidylglycerol, phosphatidylserine, or phosphatidic acid) or malondialdehyde (MDA). Antibodies or their antigen-binding fragments that specifically bind to phospholipids may be C2 mAbs or derivatives thereof. Phospholipids may be present in cells in or adjacent to tissues that are (or at risk of) tissue damage (e.g., non-ischemic injury), oxidative damage, or any combination thereof, on the surface of basement membranes (e.g., Bruch's membranes), or in pathological structures (e.g., in drusen). Phospholipids may be neutral, negatively or positively charged, or oxidized. Antibodies or antigen-binding fragments that specifically bind to phospholipids are C2 mAbs, or C2 mAbs, as described in Elvington et al., J Immunol., 188(3):1460-1468 (2012). These may be antigen-binding fragments derived from mAbs. Exemplary CDRs of C2 mAbs are shown in SEQ ID NOs: 17-22. C2 mAbs recognize a subset of phospholipids exposed after complement activation or ischemia, which is referred to herein as "C2 antibody-reactive phospholipids." C2 mAbs have been shown to recognize a subset of phospholipids that include phosphatidylcholine, phosphatidylethanolamine, and cardiolipin, but do not include phosphatidylglycerol or phosphatidylserine.

[0142] In some cases, the target-directed portion may be an antibody or its antigen-binding fragment that specifically binds to deposited or opsonized C3 fragments, e.g., C3b, iC3b, C3d, or C3dg, but may or may not bind to free, circulating, or undeposited C3 fragments, e.g., C3a, C3b, C3c, or C3f. In some examples, a fusion protein construct containing an anti-C3d or anti-C3dg antibody or its antigen-binding fragment can bind to deposited C3 fragments with relatively high affinity compared to free C3 or C3 fragments. For example, an antibody or its antigen-binding fragment may bind to C3 and C3b with binding affinity about 1 / 5, 1 / 6, 1 / 7, 1 / 8, 1 / 9, or 1 / 10 compared to C3d. In some embodiments, the antibody or antigen-binding fragment binds to C3 and / or C3b with 10 -4 M or above, 10 -3 M or higher or 10 -2 Joined by M or greater KD, with 10 in iC3b, C3dg, or both. -8 M or less, 10 -9 M or less or 10 -10 It binds with a binding affinity (KD) of M or less.

[0143] In some cases, fusion protein constructs containing anti-C3d or anti-C3dg antibodies or their antigen-binding fragments can bind to both deposited C3 fragments and free C3 or C3(H2O) fragments. Furthermore, the anti-C3d or anti-C3dg antibodies or their antigen-binding fragments in exemplary fusion protein constructs can bind to complement fragment C3d, distinguishing between tissue-bound C3 fragments and circulating C3 (e.g., C3, C3b, or C3(H2O)). It may also be possible that the antibody has the ability to do so. Examples of anti-C3d or anti-C3dg antibodies or their antigen-binding fragments include, but are not limited to, mAb 3d9a, 3d29, and 3d8b (see, for example, US9815890). In some cases, the anti-C3d or anti-C3dg antibodies of this disclosure can bind to C3d with higher specificity than commercially available anti-C3d antibodies, such as the anti-C3d antibodies represented by Quidel catalog numbers A207 and A250, commercially available from Quidel Corporation (Quidel Corp., San Diego and Santa Clara, Calif.). In some cases, the target-directing portion may include an antibody that is specific to C3d and / or other C3 fragments (such as C3b, iC3b, C3c, C3dg), for example, antibody C8D3 described in U.S. Patent Application Publication No. 2016 / 0333082. The antibody C8D3 can bind to the epitope on C3d with high affinity (see, for example, Figures 3 and 8 in US2016 / 0333082), which overlaps with the CR2-binding epitope. The heavy chain sequence of C8D3 may be SEQ ID NO: 154, the light chain sequence SEQ ID NO: 155, CDRH1, CDRH2, and CDRH3 of C8D3 are SEQ ID NOs: 156, 157, and 158, respectively, and CDRL1, CDRL2, and CDRL3 are SEQ ID NOs: 159, 160, and 161, respectively. This publication also describes four hybridoma clones (i.e., clones B7, C2, C6, and C8) that produce antibodies against C3d. The heavy chain sequence of C6 is sequence number 162 and the light chain sequence is sequence number 163. CDRH1, CDRH2, and CDRH3 of C6 are sequence numbers 164, 165, and 166, respectively, and CDRL1, CDRL2, and CDRL3 are sequence numbers 167, 168, and 169, respectively.

[0144] In some cases, the C3d antibody may be a neoanti-C3d antibody ("neoanti-C3d"), as described in U.S. Patent Application Publication 2015 / 0139899, ​​which binds to C3d but not to C3c. The binding affinity of the C3d antibody to its epitope is approximately 100 pM to 500 pM, e.g., 447 pM. The C3d antibody may be a neoanti-iC3b antibody ("neoanti-iC3b"), which in some cases binds to that epitope with an affinity of approximately 100 pM to 500 pM, e.g., 262 pM, but does not bind to C3c or C3d.

[0145] In some cases, C3d antibodies can be monoclonal antibodies M130 that are specific to the antigenic determinants represented by C3bi, C3dg, and C3d, which are almost undetectable by C3 and C3b. M130 has been shown to have a higher affinity for C3d and iC3b than for C3(H2O) (JDTamerius et al., J.Immunol.135(3):2015-19(1985)), and can bind to residues 1209-1236 and 1217-1232 of C3d (Lambris et al., PNAS, 82(12):4235-9(1985).

[0146] In some cases, the C3d antibody may be a monoclonal antibody C3-12.2 (Hidalgo et al., Eur.J.Immunol.47(3):504-15(2017)) that binds to human, rat, and mouse C3dg fragments. It was prepared using C3-deficient mice immunized with a mixture of human C3b, iC3b, and C3dg proteins and measured by BiaCore with a K2 of approximately 95 nM. D The antibodies C3-12.2, as well as antibodies 3d29, 3d8b, and 3d9a, can recognize one or more overlapping C3 fragments or variants, and Fab appears to bind to the same or adjacent epitopes as CR2.

[0147] In some cases, the C3d antibody may be a monoclonal antibody 15-39-06, assembled in wild-type rats using a synthetic peptide derived from human C3dg and conjugated to diphtheria toxin (KJRassmussen et al., J.Im). munol.Methods,444:51-5(2017). It may have specificity for C3dg complement division products.

[0148] In some cases, C3d antibodies may be commercially available antibodies that are specific to C3d and / or other C3 fragments (i.e., C3b, iC3b, C3c, C3dg, etc.). Examples include antibodies available from Quidel, where monoclonal antibodies A250 and A209 have been reported to be specific to the neoepitope C3d and iC3b, respectively. Antibody A250 has been shown to agglutinate EC3bi, EC3b, and EC3d cells in indirect hemagglutination assays, and has also been shown to bind to radiolabeled purified iC3b, C3b, and C3d but not to similarly labeled C3 or C3c. Antibody A209 has been shown to agglutinate EC3bi but not EC3b or EC3d cells in indirect hemagglutination assays, and has also been shown to bind to radiolabeled purified iC3b but not to similarly labeled C3, C3b, C3d, or C3c. Further examples include the anti-C3d antibody 7C10, which has an unknown epitope, and the anti-C3d antibody [E28-P](ab136916), which has been reported to bind to the N-terminal epitope of C3d, both of which can be obtained from Abcam. In some cases, commercially available antibodies may be the anti-C3d antibody 053A-514.3.1.4 and the iC3b antibody 013III-1.16 (formerly MCA2607), both of which are available from BioRad and have been reported to be specific to the neoantigens C3d and iC3b, respectively. Antibody 3E7 is available from Sigma and has been reported to recognize both C3b and iC3b. Origene offers the monoclonal antibody AM26358PU-N, which reacts with neoantigens on iC3(C3(H2O)), iC3b, C3dg, and / or C3g, recognizing iC3b, C3dg, and C3g in plasma, but not C3 or C3b. (USBiological Life) The C0010-19 rat anti-C3g antibody (which recognizes iC3, iC3b, and C3dg) is available from Sciences. The inactive C7850-13V-ML550 mouse anti-complement C3b antibody is also available from USBiological Life Sciences. This antibody has been reported to recognize the human inactivated complement C3b (iC3b) neoantigen in serum. Hycult offers several antibodies; for example, antibody HM2199 reacts with neoantigens on iC3, iC3b, C3dg, and C3g, recognizing iC3b, C3dg, and C3g in serum, but not C3 or C3b, making it an anti-human C3g mAb 9(YB2 / 90-5-20); monoclonal antibody HM2198 has been reported to react with linear determinants in C3d found on C3, C3b, iC3b, C3dg, and C3d, recognizing C3, C3b, iC3b, C3dg, and C3d, but not C3c, making it an anti-human C3d,mAb. 3(YB2 / 39-11-1-7); antibody HM2168 is an active human C3, clone bH6 specific to C3 neoepitopes expressed on C3b, iC3b, and C3c cleavage fragments, rather than C3dg and C3f (P. Garred et al., Scand J Immunol 1988, 27:319); antibody HM2257 is an active human C3, mAb I3 / I5 that recognizes active complement protein C3, or more specifically, neoepitopes on C3b, iC3b, and C3dg that are not present in native C3. From Meridian, antibody H54189M is available, which has been reported to react with the alpha chain of C3b but not with C3a or C3d, making it possible to demonstrate C3 deposition in tissues, cells, microorganisms, and immune complexes.

[0149] Each of the 3d9a, 3d29, and 3d8b antibodies may be able to bind to inflamed renal tissue when injected intravenously into mice, and may be able to bind to C3 opsonized zymosan, which is known to express iC3b but not C3b. Therefore, these antibodies may be able to distinguish between tissue-bound fragment C3d, circulating naive C3, and fragment C3b. C3-binding antibodies or their antigen-binding properties The fragments can bind to C3d or C3dg from multiple species (species cross-reactivity). Anti-C3d or anti-C3dg antibodies or their antigen-binding fragments can bind to C3d or C3dg from at least one species selected from humans, non-human mammals (e.g., cynomolgus monkey or cynomolgus macaque, rhesus monkey, ape, baboon, chimpanzee, orangutan, or gorilla), rodents (e.g., mouse, rat, hamster, guinea pig, gerbil, or rabbit), cattle, sheep, goats, donkeys, pigs, dogs, cattle, horses, and camels. Anti-C3d or anti-C3dg antibodies or their antigen-binding fragments can bind to cynomolgus monkey C3d or C3dg. The anti-C3d or anti-C3dg antibodies or their antigen-binding fragments described herein bind to C3d or C3dg from at least two species selected from those listed above. The anti-C3d or anti-C3dg antibodies or their antigen-binding fragments bind to both human and cynomolgus monkey C3d or C3dg. The anti-C3d or anti-C3dg antibodies or their antigen-binding fragments may be antibodies selected from 3d8b, 3d9a, 3d29, 3d11, 3d31, 3d3, 3d15, 3d10, and 3d16 (see, for example, US9815890). The anti-C3d or anti-C3dg antibodies or their antigen-binding fragments may be antibodies selected from 3d9a, 3d29, and 3d8b. The anti-C3d or anti-C3dg antibody or its antigen-binding fragment may be 3d29.

[0150] In some examples, an anti-C3d or anti-C3dg antibody or its antigen-binding fragment of a fusion protein construct may compete with CR2 for binding to C3d or C3dg. Such an antibody or its antigen-binding fragment may reduce the binding ability of the CR2 protein to human complement components C3d or C3dg by more than 50% (e.g., more than 55, 60, 65, 70, 75, 80, 85, 90, or 95%). For example, CR2-C3d binding may be reduced to at least 60%, at least 40%, or a percentage in between. An anti-C3d or anti-C3dg antibody or its antigen-binding fragment may significantly inhibit or block the binding of CR2 to C3d. In some embodiments, such antibodies are 3d9a, 3d29, or 3d8b. In some cases, exemplary fusion protein constructs comprising an anti-C3d or anti-C3dg target-directed domain and a complement regulator may compete more effectively with CR2 binding to C3d or C3dg compared to the anti-C3d or anti-C3dg target-directed domain alone.

[0151] Fusion protein construct design A fusion protein construct comprising a target-directing moiety and a complement modifier may include an antibody or its antigen-binding fragment as the target-directing moiety. Examples of target-directing moieties include, but are not limited to, monoclonal antibodies or antibody fragments, diabodies, chimeric or chimeric antibodies or antibody fragments, humanized antibodies or antibody fragments, immunized human antibodies or antibody fragments, fully human antibodies or antibody fragments, bispecific antibodies or antibody fragments, monovalent antibodies or antibody fragments, single-chain antibodies, immunoglobulin G1 (IgG1) heavy chains, single-chain variable fragments (i.e., scFv), sc(fv)2, tandem scFv, diabodies, VHH domains, VH domains, Fv, Fd, Fab heavy chains, Fab light chains, Fab, Fab', and Fab' light chains, Fab' heavy chains, and F(ab')2. The antibody or antigen-binding fragment forming the target-directing moiety may be a human antibody, a humanized antibody, or a mouse antibody. A fusion protein construct may contain one or more target-directed moieties, each containing an antibody or antigen-binding fragment specific to the target. In addition, a fusion protein construct may contain more than one complement modifier. If a fusion protein construct contains more than one complement modifier, it may contain multiple molecules of the same complement modifier, or it may contain different types of complement modifiers. Similarly, a fusion protein construct having multiple target-directed moieties may contain several molecules of the same type of target-directed moiety, thereby being polyvalent with respect to the target-directed moieties, or it may contain several molecules of different types of target-directed moieties, thereby being polyvalent with respect to the target-directed moieties. This is also acceptable. The targets of the fusion protein construct may be, for example, complement proteins or fragments thereof, domains of mammalian annexin proteins, or phospholipids. Since the fusion protein construct may have the ability to specifically bind to more than one target, it is conceivable that in some examples at least two of the above targets may be specifically bound by the fusion protein construct described herein. Therefore, the fusion protein construct may be bivalent, triplicate, quadruple-specific, etc. Furthermore, the fusion protein construct can be polyvalent, for example, bivalent, trivalent, tetravalent, etc. For example, the target-directing portion of a bivalent fusion protein construct may be specific to a target, for example, a domain of mammalian annexin, a phospholipid, or a C3 complement protein fragment (e.g., C3d), ​​and the complement regulator of the fusion protein construct may be specific to another protein in the complement pathway. A tetravalent fusion protein construct may have a bivalent target-directing portion, for example, a bivalent antibody or its antigen-binding fragment having two binding domains to a target protein, and two molecules of a complement regulator. Furthermore, the fusion protein construct can be a bispecific trivalent protein in which the target-directing portion contains a bivalent antibody or its antigen-binding fragment and the constant domain (Fc) of the antibody, and the complement regulator is fused to the bivalent antibody or the Fc domain.

[0152] Other examples of fusion protein constructs include heterodimerized Fc regions containing various modifications that promote the formation of heterodimerized Fc regions rather than homodimerized Fc regions. See, for example, JH Ha et al. Immunoglobulin Fc Heterodimer Platform Technology: From Design to Applications in Therapeutic Antibodies and Proteins. Front.Immuno.7:394 (2016). Conventional IgG antibodies are polyvalent and monospecific, and their assembly relies on the homodimerization of two identical heavy chains (HCs) in vivo, which is mediated by homodimerized association between CH3 domains in B cells and subsequent disulfide linkage between each HC and each light chain (LC). Therefore, the development of bsAbs using unmodified IgG forms with wild-type HCs and LCs involves HC-HC and HC VH-CH1-This may involve problems with incorrect LC pairing. In addition, the development of trivalent fusion constructs having complement regulators ligated to only one of the two HCs or only one of the two LCs may involve problems with incorrect pairing. Therefore, the heterodimeric Fc regions of the target-directed moieties disclosed herein may be beneficial in terms of avoiding problems with incorrect HC pairing. While the wild-type Fc region is a polypeptide homodimer, the Fc domains disclosed herein include amino acid substitutions such that, in some examples, they do not form homodimers. The monomeric Fc domains Fc1 and Fc2 are IgG Fc in some embodiments. In some embodiments, the monomeric Fc domains Fc1 and Fc2 are from other immunoglobulin subclasses, including IgA, IgE, IgD, and IgM. The heterodimeric Fc domain of the target-directed moiety described herein includes a mutant CH3 constant domain and a CH2 constant domain, both containing amino acid mutations that promote the formation of the heterodimer with stability comparable to that of the naturally occurring homodimeric Fc. Wild-type Fc is inherently homodimeric, a characteristic promoted by both hydrophobic interactions at the center of the CH3 contact surface and symmetric electrostatic interactions around the hydrophobic core. The Fc domains described herein may include amino acid substitutions such that they do not form homodimers. The Fc domains described herein may include amino acid substitutions that are more favorable to heterodimer formation than homodimer formation. In certain examples, heterodimeric Fc domains may (i) be stereocomplementary designs that make symmetry asymmetric (e.g., KiH, HA-TF, and ZW1) (e.g., Klein et al. Progress in overcoming the chain association issue in bispecific heterodimeric IgG antibodies. mAbs 4(6):653-66 (2012), GLMoore et al. A novel bispec See (iv) (iv) ific antibody format enables simultaneous bivalent and monovalent co-engagement of distinct target antigens. mAbs 3(6)546-557 (2011), TS von Kreudenstein et al. Improving biophysical properties of a bispecific antibody scaffold to aid developability: Quality by molecular design. mAbs 5(5):646-654 (2013), HJ Choi et al. A heterodimeric Fc-based bispecific antibody simultaneously targeting VEGFR-2 and Met exhibits potent antitumor activity. Mol Cancer Ther 12(12):2748-59 (2013)., (ii) is generated using charge-to-charge exchange (e.g., DD-KK), (iii) switching from charge to stereocomplementarity and additional broad electrostatic interactions (e.g., EW-RVT), and (iv) isotype chain exchange [e.g., chain exchange operation domain (SEED)]. Examples of strand exchange mutations include 45 residues derived from IgA on IgG1 CH3-Fc1 and 57 residues derived from IgG1 on IgA CH3-Fc2, or vice versa. Examples of stereocomplementary mutations that create asymmetry include HA-TF (S364H / F405A in Fc1-CH3 or CH3A and Y349T / T394F in Fc2-CH3 or CH3B) and ZW1 (T350V / L351Y / F405A / Y407V in Fc1-CH3 or CH3A and T350V / T366L / K392L / T394W in Fc2-CH3 or CH3B).In other examples, Fc variants can be created using the "knob-into-hole (KiH)" technique, in which case Fc1 contains a "knob" mutation of T366W in Fc1-CH3 or CH3A, and Fc2 contains a "hole" mutation of T366S / L368A / Y407V in the Fc2-CH3 or CH3B domain. In further examples, Fc variants can be created using the "knob-into-hole (KiH)" and disulfide bond techniques, KiH. s-s These can be produced using , in which case Fc1 contains a "knob" mutation of T366W / S354C in Fc1-CH3 or CH3A, and Fc2 contains a "hole" mutation of T366S / L368A / Y407V / Y349C in the Fc2-CH3 or CH3B domain. In such examples, heterodimerization proceeds favorably due to hydrophobic interactions in the core at the contact surface between Fc1-CH3 or CH3A and Fc2-CH3 or CH3B. An example of a charge-to-charge exchange mutation when the Fc heterodimer that favors the interaction is based on electrostatic complementarity is DD-KK (K409D / K392D in Fc1-CH3 or CH3A and D399K / E356K in Fc2-CH3 or CH3B, or vice versa). Examples of mutations resulting from the switch from charge to stereocomplementarity and additional broad electrostatic interactions include EW-RVT (K360E / K409W in Fc1-CH3 or CH3A and Q347R / D399V / F405T in Fc2-CH3 or CH3B, or vice versa); EW-RVT containing CH3 inter-SS bonds. s-sExamples include K360E / K409W / Y349C in Fc1-CH3 or CH3A and Q347R / D399V / F405T / S354C in Fc2-CH3 or CH3B, or vice versa. In yet another example, Fc mutants can be created using hydrophobic or steric complementarity and electrostatic complementarity, e.g., 7.8.60 (K360D / D399M / Y407A in Fc1-CH3 or CH3A and E345R / Q347R / T366V / K409V in Fc2-CH3 or CH3B, or vice versa). Heterodimers forming the Fc mutants described herein can also be created by directed evolution combined with yeast surface presentation and high-speed, high-volume screening. For example, a combinatorial heterodimer type Fc library presentation system can be developed by pairing two haploid yeast cell lines, where one haploid cell line presents an Fc chain library (CH3-Fc1 or CH3A) with a mutation in one CH3 domain on the surface of the yeast cell. One cell line is presented, and the other secretes an Fc chain library (CH3-Fc2 or CH3B) with a mutation in the CH3 domain of the other cell line. In paired cells, the secreted CH3-Fc2 or CH3B can be presented on the cell surface by heterodimerization with the presented CH3-Fc1 or CH3A. Detection of this interaction based on fluorescence allows for screening of the library for heterodimeric Fc variants by flow cytometry. Antibodies or antigen-binding fragments containing the wild-type Fc domain have the ability to interact with the embryonic Fc receptor (FcRn) in a pH-dependent manner, and this interaction can result in an extended serum half-life. Residues crucial for the high-affinity interaction between the Fc domain and FcγR are located in the CH2 domain. Therefore, in some cases, the target-directed portion of the Fc heterodimer contains a CH2 domain with a wild-type IgG sequence.

[0153] In some examples, the "CH3 domain" in the Fc region, comprising a sequence of residues on the C-terminal side of the CH2 domain (i.e., from amino acid residue approximately 341 to amino acid approximately 447 in IgG), may be a CH3 domain of the native sequence or a mutant CH3 domain (e.g., a CH3 domain having a "projection" introduced on one strand and a corresponding "hole" introduced on the other strand). Such a mutant CH3 domain may, in some examples, be part of the heterodimeric Fc domain described herein. Thus, in some examples, the fusion protein construct includes an antibody having a projection and a hole to accept it, or an antigen-binding fragment thereof.

[0154] In some cases, the Fc region may contain a human or mouse IgG1 or human IgG4 sequence. In some cases, the Fc region may contain a human IgG1 or IgG4 or mouse IgG1 sequence that contains amino acid substitutions compared to the wild-type sequence. Examples of such Fc regions are shown in SEQ ID NOs: 247 and 248.

[0155] Domains and structure of fusion protein constructs A fusion protein construct comprises one or more polypeptides, each containing a domain and linked, for example, by one or more disulfide bonds. For example, in some cases, the fusion protein construct may comprise a first polypeptide chain comprising domains A, B, C, D, and R, where domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, domain B comprises a heavy chain CH1 constant region amino acid sequence, domain C comprises a heavy chain CH2 constant region amino acid sequence, domain D comprises a heavy chain CH3 constant region amino acid sequence, and domain R comprises a complement regulator polypeptide. In some embodiments, one or more of domains B, C, and D are optional. The first polypeptide may further comprise a hinge region, and the first polypeptide may further be linked to a second polypeptide comprising further domains, such as domain E containing a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, and an optional domain F containing a light chain constant region amino acid sequence (CL1). Linking between the first and second polypeptides may be achieved by one or more disulfide bonds between domain B and domain F. In some cases, the first and second polypeptides described above are combined using various orientations to create fusion protein constructs such as monomers, trivalent homodimers, trivalent homodimers, tetravalent homodimers, and tetravalent heterodimers.

[0156] In various examples of multivalent fusion protein constructs (e.g., bivalent, trivalent, tetravalent), the multivalent nature of the fusion protein construct can improve its avidity to a specific target. As used herein, "avidity" may refer to the overall strength of the interaction between two or more molecules, for example, a fusion protein construct containing a bivalent target-directing moiety, and avidity may be the cumulative strength of the interaction given by the affinity of the two antigen-binding sites. Avidity can be used, for example, to determine affinity. It can be measured in the same way as used. In certain cases, avidity may be the cumulative strength of the interaction, expressed as the affinity of multiple antigen-binding sites to distinct antigens on a common specific target or complex, such as distinct antigens found on individual cells. In certain cases, avidity may be the cumulative strength of the interaction, expressed as the affinity of multiple antigen-binding sites to distinct epitopes on individual antigens on a common individual antigen.

[0157] Domain A (VH) Domain A of the fusion protein constructs described herein may include a VH amino acid sequence, for example, an antibody heavy chain variable domain sequence. In typical antibody compositions, both in natural and in the fusion protein constructs described herein, a specific VH amino acid sequence may associate with a specific VL amino acid sequence to form an antigen-binding site. In various examples, the VH amino acid sequence may be a mammalian sequence, including a human sequence, or a synthetic sequence, or a combination of non-human mammalian, mammalian, and synthetic sequences. In various examples, the VH amino acid sequence may be a mutant sequence of a naturally occurring sequence that retains a CDR sequence sufficient to maintain the desired antigen-binding affinity. In some examples, the VH sequence may be from an IgA1, IgA2, IgD, IgE, IgG1, IgG2, IgG3, IgG4, or IgM isotype. In some examples, the VH sequence may be from an IgG1 isotype.

[0158] Domain E(VL) Domain E of the fusion protein constructs described herein may include a VL amino acid sequence, such as an antibody light chain variable domain sequence. In a typical configuration, in both natural antibodies and the fusion protein constructs described herein, a specific VL amino acid sequence may associate with a specific VH amino acid sequence to form an antigen-binding site. In various examples, the VL amino acid sequence may be a mammalian sequence including a human sequence, a synthetic sequence, or a combination of human, non-human mammalian, mammalian, and synthetic sequences. In various examples, the VL amino acid sequence may be a mutant sequence of a naturally occurring sequence that retains at least 70%, 75%, 80%, 85%, 90%, 95%, or more amino acid identity. In certain examples, the VL amino acid sequence may be a lambda (λ) light chain variable domain sequence. In certain examples, the VL amino acid sequence may be a kappa (κ) light chain variable domain sequence. In some examples, the VL sequence may be from IgA1, IgA2, IgD, IgE, IgG1, IgG2, IgG3, IgG4, or IgM isotypes. In some cases, the VL sequence may originate from the IgG1 isotype.

[0159] Domain B (CH1) The CH1 amino acid sequence of the fusion protein constructs described herein may be the sequence of the second domain of the antibody heavy chain when viewed from the N-terminus to the C-terminus. In certain examples, the CH1 sequence may be an endogenous sequence or a mutant sequence thereof that retains at least 70%, 75%, 80%, 85%, 90%, 95% or more of amino acid identity. In some examples, the CH1 sequence may be a mammalian sequence, including, but not limited to, sequences from mouse, rat, hamster, rabbit, camel, donkey, goat, and human. In some examples, the CH1 sequence may be a human sequence. In some examples, the CH1 sequence may be from an IgA1, IgA2, IgD, IgE, IgG1, IgG2, IgG3, IgG4, or IgM isotype. In some examples, the CH1 sequence may be from an IgG1 isotype. In some examples, the CH1 sequence may be from an IgG4 isotype.

[0160] Domain F (CL1) The CL amino acid sequence of the fusion protein construct described herein may be the antibody light chain constant domain sequence. In certain examples, the CL sequence may be an endogenous sequence, or at least 70%, 7%. The mutant sequence may retain 5%, 80%, 85%, 90%, 95% or more of amino acid identity. In some examples, the CL sequence may be a mammalian sequence, including, but not limited to, sequences from mice, rats, hamsters, rabbits, camels, donkeys, goats, and humans. In some examples, the CL sequence may be a human sequence. In some examples, the CL amino acid sequence may be a lambda (λ) light chain constant domain sequence. In some examples, the CL amino acid sequence may be a human lambda light chain constant domain sequence. In certain embodiments, the CL amino acid sequence is a kappa (κ) light chain constant domain sequence. In preferred embodiments, the CL amino acid sequence is a human kappa (κ) light chain constant domain sequence.

[0161] Domain C (CH2) In the fusion protein constructs described herein, domain C may have a CH2 amino acid sequence. The CH2 sequence may be an endogenous sequence or a mutant sequence thereof that retains at least 70%, 75%, 80%, 85%, 90%, 95% or more of amino acid identity. In some examples, the CH2 sequence may be a mammalian sequence, including, but not limited to, a human sequence. In some examples, the CH2 amino acid sequence may have an N-terminal hinge region that ligates domain C to domain B, for example, by ligating the C-terminus of domain B (CL1 sequence) to the N-terminus of domain C (CH2 sequence). In some examples, the CH2 sequence may be from an IgA1, IgA2, IgD, IgE, IgG1, IgG2, IgG3, IgG4, or IgM isotype. In some examples, the CH2 sequence may be from an IgG1 isotype. In some examples, the CH2 sequence may be from an IgG4 isotype.

[0162] Domain D (CH3) In the fusion protein constructs described herein, domain D may comprise a constant region amino acid sequence, such as a CH3 amino acid sequence. The CH3 sequence may be an endogenous sequence or a mutant sequence thereof that retains at least 70%, 75%, 80%, 85%, 90%, 95% or more of amino acid identity. In some examples, the CH3 sequence may be a mammalian sequence, including, but not limited to, a human sequence. In some examples, domain D may comprise a constant region sequence that is a CH3 sequence comprising a knob-hole orthogonal mutation; an isoallotyped mutation; and a mutation of either S354C or Y349C, or any combination thereof, that forms an operational disulfide bridge with the CH3 domain containing the orthogonal mutation. In some examples, the knob-hole orthogonal mutation in combination with an isoallotyped mutation may include mutations such as: D356E, L358M, T366S, L368A, and Y407V. In some cases, the CH3 sequence may be from the IgA1, IgA2, IgD, IgE, IgG1, IgG2, IgG3, IgG4, or IgM isotype. In some cases, the CH3 sequence may be from the IgG1 isotype. In some cases, the CH3 sequence may be from the IgG4 isotype. In any of these examples, the CH1, CH2, and CH3 sequences may be from the same isotype, for example, IgG1 or IgG3.

[0163] Complementarity determination regions within Domains A and F The VH (domain A) and VL (domain F) amino acid sequences of the various fusion protein constructs described herein may include highly variable sequences referred to as "complementarity-determining regions" (CDRs), typically three CDRs (CDR1, CD2, and CDR3). In various examples, the CDRs may be mammalian sequences, including, but not limited to, sequences from mouse, rat, hamster, rabbit, camel, donkey, goat, and human. In some examples, the CDRs may be human sequences. In some examples, the CDRs may be naturally occurring sequences. In some examples, the CDRs may be naturally occurring sequences that have mutated, altering the binding affinity of the antigen-binding site to a particular antigen or epitope. In certain examples, naturally occurring CDRs may have mutated in vivo in the host through affinity maturation and somatic hypermutation. In certain cases, CDRs may be mutated in vitro by methods including, but not limited to, PCR mutagenesis and chemical mutagenesis. In various cases, CDRs may include, but not limited to, synthetic sequences obtained from random sequence CDR libraries and reasonably designed CDR libraries.

[0164] Framework domain and CDR grafting VH and VL amino acid sequences may further contain “framework region” (FR) sequences. FRs are often conserved sequence regions that can serve as scaffolding for scattered CDRs and are typically arranged in the order FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4 (from N-terminus to C-terminus). In various examples, FRs can be mammalian sequences, including, but not limited to, sequences from mouse, rat, hamster, rabbit, camel, donkey, goat, and human. In some examples, FRs can be human sequences. In various examples, FRs can be naturally occurring sequences. In various examples, FRs can be synthetic sequences, including, but not limited to, reasonably designed sequences.

[0165] In various examples, both FR and CDR may be the same naturally occurring variable domain sequence. In various embodiments, FR and CDR may be from different variable domain sequences, with CDR being grafted onto the FR scaffold to provide specificity for a particular antigen. In certain examples, any grafted CDR may originate from the same naturally occurring variable domain sequence. In certain examples, grafted CDRs may originate from different variable domain sequences. In certain examples, grafted CDRs may be synthetic sequences, including, but not limited to, random sequence CDR libraries and CDRs obtained from reasonably designed CDR libraries. In certain examples, grafted CDRs and FRs may be from the same species. In certain examples, grafted CDRs and FRs may be from different species. In certain examples, various domains of the fusion protein construct may be "humanized," with grafted CDRs being non-human mammalian sequences, including, but not limited to, mouse, rat, hamster, rabbit, camel, donkey, and goat sequences, and FR being a human sequence. In various cases, a portion or specific sequence of FR from one species may be used to replace a portion or specific sequence of FR from another species.

[0166] Homodimerization or heterodimerization of the two domains D (CH3 domain) In the fusion protein constructs described herein, two domains D may associate to form a dimeric construct. In various examples, all domains D have identical amino acid sequences. In certain examples, domain D contains an endogenous CH3 sequence. In some examples, if the fusion protein construct is a heterodimer or is heterodimeric, two domains D with different amino acid sequences can associate, each containing orthogonal modifications within its endogenous CH3 sequence, one domain D interacts with the other domain with a different sequence, and neither domain interacts much with the other domain D (CH3 domain) which does not have orthogonal modifications.

[0167] As used herein, “orthogonal modification” or synonymously “orthogonal mutation” may be one or more manipulated mutations in the amino acid sequence of an antibody domain that can increase the affinity of binding between a first domain having an orthogonal modification and a second domain having a complementary orthogonal modification. In certain cases, an orthogonal modification may decrease the affinity between a domain having an orthogonal modification and a domain without a complementary orthogonal modification. In certain cases, an orthogonal modification may be a mutation in the endogenous antibody domain sequence. In various examples, an orthogonal modification may be a modification of the N-terminus or C-terminus of the endogenous antibody domain sequence, including, but not limited to, amino acid additions or deletions. In some examples, an orthogonal modification may include, but not limited to, manipulated disulfide crosslinks, knob-in-hole mutations, and charge-pair mutations. In some cases, orthogonal Modifications may include, but are not limited to, combinations of orthogonal modifications selected from manipulative disulfide crosslinks, knob-in-hole mutations, and charge-pair mutations. In some examples, orthogonal modifications may be combined with amino acid substitutions that reduce immunogenicity, such as isoallotype mutations.

[0168] In certain cases, when the fusion protein construct is a heterodimer, orthogonal modifications may include mutations that produce an operational disulfide bridge between the first and second domains. As used herein, “operational disulfide bridge” may include mutations in two or more domains that give a non-endogenous cysteine ​​amino acid such that a non-natural disulfide bond is formed when the two or more domains associate. In certain cases, operational disulfide bridges can improve orthogonal association between specific domains. In some cases, mutations that can produce an operational disulfide bridge may include a K392C mutation in one of the first or second CH3 domains and a D399C mutation in the other CH3 domain. In one example, mutations that can produce an operational disulfide bridge may include an S354C mutation in one of the first or second CH3 domains and a Y349C mutation in the other CH3 domain. In another example, mutations that can result in manipulative disulfide crosslinking may include the 447C mutation, which results from an extension of the C-terminus of the CH3 domain incorporating a KSC tripeptide sequence within both the first and second CH3 domains.

[0169] In some examples, orthogonal modifications may include knob-hole (synonymously, knob-in-hole) mutations. As described herein, knob-hole mutations may include mutations that alter the steric features of the surface of a first domain such that the first domain associates with a second domain having a complementary steric mutation preferentially over association with a domain that does not have a complementary steric mutation. In various examples, knob-hole mutations may be combined with manipulative disulfide crosslinks. In various embodiments, knob-hole mutations, isoallotype mutations, and manipulative disulfide mutations may be combined to produce heterodimeric fusion protein constructs. In certain examples, knob-in-hole mutations may include a T366Y mutation in one CH3 domain and a Y407T mutation in another CH3 domain. In certain examples, knob-in-hole mutations may include F405A in one CH3 domain and T394W in another CH3 domain. In certain embodiments, the knob-in-hole mutations may include the T366Y and F405A mutations in one CH3 domain, and the T394W and Y407T mutations in another CH3 domain. In certain examples, the knob-in-hole mutations may include the T366W mutation in one CH3 domain and the Y407A mutation in another CH3 domain. In certain embodiments, the combined knob-in-hole mutations and manipulated disulfide mutations may include the S354C and T366W mutations in one CH3 domain, and the Y349C, T366S, L368A, and Y407V mutations in another CH3 domain. In some examples, combined knob-in-hole mutations, isoallotype mutations, and manipulative disulfide mutations may include S354C and T366W mutations in one CH3 domain, as well as Y349C, D356E, L358M, T366S, L368A, and Y407V mutations in the other CH3 domain. In various embodiments, orthogonal modifications may be charge-pair mutations.As used herein, a charge-pair mutation can be a mutation that affects the charge of an amino acid on the surface of a domain such that the domain preferentially associates with a second domain having a complementary charge-pair mutation over association with a domain that does not have a complementary charge-pair mutation. In certain embodiments, a charge-pair mutation can improve orthogonal association between certain domains. In certain embodiments, a charge-pair mutation can improve stability between certain domains. In some examples, the charge-pair mutation is a T366K mutation in one CH3 domain and another. This is an L351D mutation in the CH3 domain.

[0170] Monomeric fusion protein constructs Exemplary monomeric fusion protein constructs are shown in Figures 2-3 and comprise a first and second polypeptide, the first polypeptide comprising the following domains: domain A, domain B, and domain R, and the second polypeptide comprising the following domains: domain E and domain F. In the fusion protein construct shown in Figure 2, domain A comprises the heavy chain variable region amino acid sequence (VH) or its antigen-binding fragment, domain B comprises the heavy chain CH1 constant region amino acid sequence, domain R comprises the complement regulator polypeptide, domain E comprises the light chain variable region amino acid sequence (VL) or its antigen-binding fragment, and domain F comprises the light chain constant region amino acid sequence (CL1). Furthermore, domains B and F of the first and second polypeptides may each be linked by one or more disulfide bonds. The domains of the first polypeptide are aligned from the N-terminus to the C-terminus in an ABR or RAB orientation, and the domains of the second polypeptide are aligned from the N-terminus to the C-terminus in an EF orientation. In the example shown in Figure 2, the monomeric fusion protein construct includes a linkage between domain A and domain R, such as a chemical linkage or a peptide linker. In a further example, as shown in Figure 3, a monomeric fusion protein construct is provided that includes a linkage between domain R and domain B.

[0171] Further exemplary monomeric fusion protein constructs are shown in Figures 1 and 4, comprising a first and second polypeptide, the first polypeptide comprising the following domains: domain A and domain B, and the second polypeptide comprising the following domains: domain E, domain F, and domain R. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The domains of the first polypeptide are aligned in an AB orientation from the N-terminus to the C-terminus, and the domains of the second polypeptide are aligned in an EF orientation from the N-terminus to the C-terminus. In the example shown in Figure 4, the monomeric fusion protein construct includes a linkage between domain F and domain R, such as a chemical linkage or a peptide linker. Further examples provide a monomeric fusion protein construct including a linkage between domain E and domain R, as shown in Figure 1.

[0172] Further examples provide fusion protein constructs containing more complement regulator polypeptides, i.e., one more than one domain R. In such cases, the fusion protein construct comprises a first polypeptide having domains A, B and a first complement regulator polypeptide (domain R1); a second polypeptide having domains E, F and a second complement regulator polypeptide (domain R2); and the fusion protein construct comprises linkages between domain R1 and domain A, or between domain R1 and domain B; and between domain R2 and domain F, or between domain R2 and domain E.

[0173] In further examples, the fusion protein construct comprises a first polypeptide containing domain A, domain B, a first complement regulator polypeptide (domain R1), and a second complement regulator polypeptide (domain R2); and a second polypeptide containing domain E, domain F, a third complement regulator polypeptide (domain R3), and a fourth complement regulator polypeptide (domain R4). The fusion protein construct may include linkages between domain R1 and domain B and between domain R2 and domain A, or between domain R2 and domain B and between domain R1 and domain A; as well as between domain R3 and domain E and between domain R4 and domain F, or between domain R4 and domain E and between domain R3 and domain E.

[0174] Homodimeric fusion protein constructs containing complement regulators bound to heavy chain polypeptides Exemplary tetravalent homodimer fusion protein constructs are shown in Figures 5-6, and two po The peptide comprises a first polypeptide containing domain A, domain B, hinge region, domain C, domain D, and domain R; and a second polypeptide containing domains E and F, wherein the first polypeptide is aligned AB-hinge domain-CDR orientation from N-terminus to C-terminus and includes a linkage between domain D and domain R; or aligned RAB-hinge region-CD orientation and includes a linkage between domain R and domain A. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The second polypeptide may be aligned EF orientation from N-terminus to C-terminus. The two first polypeptides may be linked by one or more disulfide bonds in the hinge region. Further exemplary tetravalent homodimer fusion protein constructs comprise two polypeptides, each comprising a first polypeptide containing domain A, domain B, a hinge region, domain C, and domain R; and a second polypeptide containing domains E and F, wherein the first polypeptide is aligned AB-hinge-domain-CR from N-terminus to C-terminus and includes a linkage between domain D and domain R; or aligned RAB-hinge-domain-C and includes a linkage between domain R and domain A. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The second polypeptide may be aligned EF from N-terminus to C-terminus. The two first polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0175] Further exemplary tetravalent homodimer fusion protein constructs comprise two polypeptides, each polypeptide chain comprising: a first polypeptide containing domain A, domain B, a hinge region, and domain R; and a second polypeptide containing domains E and F, wherein the first polypeptide is aligned AB-hinge-domain-R from N-terminus to C-terminus and includes a linkage between the hinge region and domain R; or aligned RAB-hinge-domain and includes a linkage between domain R and domain A. The second polypeptide may be aligned EF-F from N-terminus to C-terminus. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The two first polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0176] Heterodimeric fusion protein constructs containing complement regulators bound to heavy chain polypeptides Exemplary trivalent heterodimer fusion protein constructs are shown in Figures 7-8 and comprise a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, domain C, domain D, and domain R, which are aligned from the N-terminus to the C-terminus in a RAB-hinge region-CD orientation (as in Figure 8) and contain a linkage between domain A and domain R, or aligned from the N-terminus to the C-terminus in an AB-hinge region-CDR orientation and contain a linkage between domain D and domain R; the second polypeptide comprises domains E and F aligned from the N-terminus to the C-terminus in an EF orientation; the third polypeptide comprises domains A, domain B, a hinge region, domain C, and domain D aligned from the N-terminus to the C-terminus in an AB-hinge region-CD orientation; and the fourth polypeptide comprises domains E and F aligned from the N-terminus to the C-terminus in an EF orientation. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds, and domains B and F of the third and fourth polypeptides may be linked by one or more disulfide bonds. The first and third polypeptides may be linked together by one or more disulfide bonds in the hinge region.

[0177] Further exemplary trivalent heterodimer fusion protein constructs include the first polypeptide, and the second polypeptide. The polypeptide comprises two polypeptides, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, domain C, and domain R, which are aligned in a RAB-hinge region-C orientation from the N-terminus to the C-terminus and contain a linkage between domain A and domain R, or aligned in an AB-hinge region-C orientation and contain a linkage between domain C and domain R; the second polypeptide comprises domain E and domain F aligned in an EF orientation from the N-terminus to the C-terminus; the third polypeptide comprises domain A, domain B, a hinge region, and domain C aligned in an AB-hinge region-C orientation from the N-terminus to the C-terminus; and the fourth polypeptide comprises domain E and domain F aligned in an EF orientation from the N-terminus to the C-terminus. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds, and domains B and F of the third and fourth polypeptides may be linked by one or more disulfide bonds. The first and third polypeptides may be linked together via one or more disulfide bonds in the hinge region.

[0178] Further exemplary trivalent heterodimer fusion protein constructs include a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, and domain R, which are aligned in an RAB-hinge region orientation from the N-terminus to the C-terminus and contain a linkage between domain A and domain R, or aligned in an AB-hinge region-R orientation and contain a linkage between the hinge region and domain R; the second polypeptide comprises domains E and F aligned in an EF orientation from the N-terminus to the C-terminus; the third polypeptide comprises domains A, B, and a hinge region aligned in an AB-hinge region orientation from the N-terminus to the C-terminus; and the fourth polypeptide comprises domains E and F aligned in an EF orientation from the N-terminus to the C-terminus. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds, and domains B and F of the third and fourth polypeptides may be linked by one or more disulfide bonds. The third polypeptide may be linked together via one or more disulfide bonds in the hinge region.

[0179] Fusion protein constructs containing complement regulators bound to light chain polypeptides Further exemplary tetravalent homodimer fusion protein constructs shown in Figure 9 comprise two polypeptides, each comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, domain C, and domain D, and the second polypeptide comprises domain E, domain F, and domain R, wherein the first polypeptide is aligned in an AB-hinge-domain-CD orientation from the N-terminus to the C-terminus, and the second polypeptide may be aligned in an EFR orientation from the N-terminus to the C-terminus (as shown in Figure 9) and contain a linkage between domain F and domain R, or may be aligned in an REF orientation (as shown in Figure 10) and contain a linkage between domain E and domain R. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The two first polypeptides may be linked together in the hinge region by one or more disulfide bonds.

[0180] Further exemplary tetravalent homodimer fusion protein constructs comprise two polypeptides, each comprising a first polypeptide and a second polypeptide, wherein the first polypeptide comprises domain A, domain B, hinge region and domain C, and the second polypeptide comprises domain E, domain F and domain R, wherein the first polypeptide is aligned AB-hinged-domain-C from the N-terminus to the C-terminus, and the second polypeptide is aligned EFR from the N-terminus to the C-terminus between domain F and domain R. Linkages may be included, or linkages may be included between domain E and domain R aligned in the REF orientation. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The two first polypeptides may be linked together by one or more disulfide bonds in the hinge region.

[0181] Further exemplary tetravalent homodimer fusion protein constructs comprise two polypeptides, each comprising a first polypeptide and a second polypeptide, the first polypeptide comprising domain A, domain B and a hinge region, and the second polypeptide comprising domain E, domain F and domain R, wherein the first polypeptide is aligned in an AB-hinge region orientation from the N-terminus to the C-terminus, and the second polypeptide may be aligned in an EFR orientation from the N-terminus to the C-terminus and contain a linkage between domain F and domain R, or aligned in a REF orientation and contain a linkage between domain E and domain R. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds. The two first polypeptides may be linked by one or more disulfide bonds in the hinge region.

[0182] An exemplary heterodimer construct is provided, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, domain C, and domain D; the second polypeptide comprises domain E, domain F, and domain R; the third polypeptide comprises domain A, domain B, a hinge region, domain C, and domain D; and the fourth polypeptide comprises domain E and domain F, wherein the first and third polypeptides are aligned AB-hinge region-CD from the N-terminus to the C-terminus; the second polypeptide is aligned EFR from the N-terminus to the C-terminus (as shown in Figure 12) and includes a linkage between domain F and domain R, or is aligned REF from the N-terminus to the C-terminus (as shown in Figure 11) and includes a linkage between domain E and domain R; and the fourth polypeptide is aligned EF from the N-terminus to the C-terminus. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds, and domains B and F of the third and fourth polypeptides may be linked by one or more disulfide bonds. The first polypeptide and the third polypeptide may be linked together in the hinge region by one or more disulfide bonds.

[0183] The present invention provides an exemplary heterodimer construct comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, a hinge region, and domain C; the second polypeptide comprises domain E, domain F, and domain R; the third polypeptide comprises domain A, domain B, a hinge region, and domain C; the fourth polypeptide comprises domain E and domain F; the first and third polypeptides are aligned in an AB-hinge-region-C orientation from the N-terminus to the C-terminus; the second polypeptide is aligned in an EFR orientation from the N-terminus to the C-terminus and includes a linkage between domain F and domain R, or is aligned in an REF orientation and includes a linkage between domain E and domain R; and the fourth polypeptide is aligned in an EF orientation from the N-terminus to the C-terminus. Domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds, and domains B and F of the third and fourth polypeptides may be linked by one or more disulfide bonds. The first polypeptide and the third polypeptide may be linked together in the hinge region by one or more disulfide bonds.

[0184] The present invention provides an exemplary heterodimer construct comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, wherein the first polypeptide comprises domain A, domain B, and a hinge region; the second polypeptide comprises domain E, domain F, and domain R; the third polypeptide comprises domain A, domain B, and a hinge region; and the fourth polypeptide comprises domain E and domain F, wherein the first and third polypeptides are aligned in an AB-hinge region orientation from the N-terminus to the C-terminus; the second polypeptide is aligned in an EFR orientation from the N-terminus to the C-terminus and contains a linkage between domain F and domain R, or is aligned in an REF orientation and contains a linkage between domain E and domain R; and the fourth polypeptide is aligned in an EF orientation from the N-terminus to the C-terminus. The domains B and F of the first and second polypeptides may be linked by one or more disulfide bonds, and the domains B and F of the third and fourth polypeptides may be linked by one or more disulfide bonds. The first polypeptide and the third polypeptide may be linked together in the hinge region via one or more disulfide bonds.

[0185] Fusion protein construct containing a complement regulator conjugated to the constant region of an antibody This specification provides an exemplary fusion protein, shown in Figure 13, comprising a first polypeptide, a second polypeptide, and a third polypeptide, wherein the first polypeptide comprises domain R, a hinge region, domain C, and domain D; the second polypeptide comprises domain A, domain B, a hinge region, domain C, and domain D; and the third polypeptide comprises domain E and domain F, wherein the first polypeptide may be aligned R-hinge-region-CD from the N-terminus to the C-terminus and include a linkage between domain R and the hinge region; the second polypeptide may be aligned AB-hinge-region-CD from the N-terminus to the C-terminus; and the third polypeptide may be aligned EF from the N-terminus to the C-terminus. Domains B and F of the second and third polypeptides may each be linked by one or more disulfide bonds. The first and second polypeptides may be linked by one or more disulfide bonds in the hinge region. In some examples, the fusion protein contains a linkage between the complement regulator and the constant region, and the complement regulator may be CR1(1-10), CR1(1-17), factor H, MCP, or DAF linked to the constant region of an immunoglobulin molecule containing, for example, two CH3 domains, two CH2 domains, and a hinge region.

[0186] Exemplary structures Specific examples of fusion protein constructs include fusions with complement regulators CR1(1-10), CR1(1-17), and C2 antibodies or their antigen-binding fragments, e.g., C2scFv-CR1 1-10, C2scFv-CR1 1-17, C2scFv-Crry, C2 IgG1 heavy chain-CR1 1-10, C2 IgG1 heavy chain-CR1 1-17, C2 IgG1 heavy chain (whole)-CR1 1-10 [paired with Fc domain containing a knob], C2 IgG1 heavy chain (whole)-CR1 1-17 [paired with Fc domain containing a knob], C2 IgG1 heavy chain (nob) [paired with Fc domain containing a hole], C2 IgG1 Fab heavy chain-Crry, C2 IgG1 Fab heavy chain-CR1 1-10, C2 IgG1 Fab heavy chain-CR1 1-17, CR1 1-10-IgG1 Fc (hole) [paired with Fc domain containing a knob], CR1 1-17-IgG1 Fc (hole) [paired with Fc domain containing a knob], CR1 1-10-C2 IgG1 heavy chain (hole) [paired with construct containing Fc domain with a knob], CR1 1-17-C2 IgG1 heavy chain (hole) [paired with construct containing Fc domain with a knob], CR1 1-10-C2 IgG1 Fab heavy chain, CR1 1-17-C2 IgG1 Fab heavy chain, CR1 The 1-10-C2 kappa light chain and the CR1 1-17-C2 kappa light chain are shown in Figure 30. Constructs that combine complement regulator factor H and anti-C3d antibody (3d29) or its antigen-binding fragment, e.g., 3d29 kappa light chain, 3d29 Fab heavy chain mouse IgG1, 3d29 Fab heavy chain mouse IgG1-Crry, 3d29 Fab heavy chain mouse IgG1-CR1 1-10, 3d29 Fab heavy chain mouse IgG1-CR1 1-17, 3d29 The construct shown in Figure 31 contains Fab heavy chain mouse IgG1-fH1-5, 3d29 heavy chain mouse IgG1 (knob), 3d29 heavy chain mouse IgG1 (whole)-fH1-5, and 3d29 heavy chain mouse IgG1-fH1-5;Fusions of anti-C3d antibody (3d8b) or its antigen-binding fragment, 3d8b Fab heavy chain mouse IgG1-Crry, 3d8b Fab heavy chain mouse IgG1-CR1 1-10, 3d8b Fab heavy chain mouse IgG1-fH1-5, 3d8b heavy chain mouse IgG1, 3d8b kappa light chain-CR1 1-10, 3d8b kappa light chain-CR1 1-17, 3d8b heavy chain mouse IgG1 (nob) paired with 3d8b heavy chain mouse IgG1 (hole), 3d8b heavy chain mouse IgG1-CR1 1-10, 3d8b heavy chain mouse IgG1-CR1 1-17, 3d8b heavy chain mouse IgG1 (hole)-fH1-5 paired with 3d8b heavy chain mouse IgG1 (nob), 3d8b heavy chain mouse IgG1-fH1-5, CR1 1-10-3d8b heavy chain mouse IgG1, CR1; 1-10-3d8b kappa light chain, CR1; 1-17-3d8b kappa light chain, 3d29 heavy chain mouse IgG1-CR1; 1-10; 3d29 heavy chain mouse IgG1-CR1; 1-10 + His tag; 3d29 heavy chain mouse IgG1 (knob)-CR1; 1-10 + His tag; 3d29 heavy chain mouse IgG1 (whole); 3d29 heavy chain mouse IgG1-CR1; 1-17; 3d29 heavy chain mouse IgG1-CR1; 1-17 + His tag; 3d29 heavy chain mouse IgG1 (knob)-CR1 1-17+His tag, 3d29 heavy chain mouse IgG1 (whole), fH1-5-3d8b heavy chain mouse IgG1, 3d8b kappa light chain-fH1-5, and fH1-5-3d8b kappa light chain, CR1 1-10-mouse IgG1 Fc (nob), 3d8b heavy chain mouse IgG1 (whole), 3d8b kappa light chain (mouse), fH1-5-mouse IgG1 Fc (nob), 3d8b Fab heavy chain mouse IgG1-CR1 1-17, 3d8b heavy chain mouse IgG1 (whole)-CR1 1-17, 3d8b heavy chain mouse IgG1 (nob), 3d8b heavy chain mouse IgG1-CR1 1-17, 3d8b heavy chain mouse IgG1, 3d8b kappa light chain-CR1 1-17, CR1 1-17-3d8b heavy chain mouse IgG1, CR1 The constructs shown in Figure 32, including the 1-17-3d8b kappa light chain, may include, but are not limited to, these. The amino acid sequences of exemplary constructs are shown in SEQ ID NOs. 103–149, and Figures 30–35 show illustrative diagrams.

[0187] Table 1 lists exemplary constructs containing an anti-C3d antibody or its antigen-binding fragment; and fusion protein constructs containing an anti-C3d antibody or its antigen-binding fragment and a complement regulator or its bioactive fragment. The sequences of the antibody heavy and light chains are shown, including the sequences of the complement regulator polypeptide or its bioactive fragment conjugated to the anti-C3d antibody (e.g., one or more complement regulators conjugated to the C-terminus or N-terminus of the heavy chain of the exemplary anti-C3d antibody; one or more complement regulators conjugated to the C-terminus or N-terminus of the light chain of the exemplary anti-C3d antibody; one or more complement regulators conjugated to the hinge region of the anti-C3d antibody). Fusion protein constructs containing an anti-C3d antibody may contain an anti-C3d antibody containing two heavy chains and two light chains, may contain an anti-C3d Fab fragment, or may contain anti-C3d Contains scFv. In some cases, the construct contains two molecules of a primary polypeptide (two heavy chains, identified in the table as HC1 or HC1+ complement regulator and HC2 or HC2+ complement regulator) and two molecules of a secondary polypeptide (two light chains, identified in the table as LC1 or LC1+ complement regulator and LC2 or LC2+ complement regulator).

[0188] [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6]

[0189] Variable domains and CDR sequences in fusion protein constructs In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 11, 12, and 13, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 11, 12, or 13 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 14, 15, and 16, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 14, 15, or 16 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0190] In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 11, 12, and 13, each having a single conserved amino acid substitution in heavy chain CDR1, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 14, 15, and 16. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 11, 12, and 13, each having a single conserved amino acid substitution in heavy chain CDR2. It comprises a first polypeptide containing three heavy chain CDRs, SEQ ID NOs. 11, 12, and 13, each having a single conservative amino acid substitution in 3, and a second polypeptide containing three light chain CDRs, SEQ ID NOs. 14, 15, and 16.

[0191] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 17, 18, and 19, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 17, 18, or 19 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 20, 21, and 22, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 20, 21, or 22 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0192] In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 17, 18, and 19, each having a single conserved amino acid substitution in heavy chain CDR1, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 20, 21, and 22. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 17, 18, and 19, each having a single conserved amino acid substitution in heavy chain CDR2, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 20, 21, and 22. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 17, 18, and 19, each having a single conserved amino acid substitution in heavy chain CDR3, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 20, 21, and 22.

[0193] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 23, 24, and 25, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 23, 24, and 25 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 26, 27, and 28, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 26, 27, and 28 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0194] In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs, SEQ ID NOs. 23, 24, and 25, with a single conserved amino acid substitution in heavy chain CDR1, and a second polypeptide comprising three light chain CDRs, SEQ ID NOs. 26, 27, and 28. In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs, SEQ ID NOs. 23, 24, and 25, with a single conserved amino acid substitution in heavy chain CDR2, and a second polypeptide comprising three light chain CDRs, SEQ ID NOs. 26, 27, and 28. In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs, SEQ ID NOs. 23, 24, and 25, with a single conserved amino acid substitution in heavy chain CDR3, and a second polypeptide comprising three light chain CDRs, SEQ ID NOs. 26, 27, and 28.

[0195] In any of the fusion protein constructs described herein, the first polypeptide is (i) a three-heavy chain CDR having the amino acid sequences of SEQ ID NOs. 29, 30 and 31, or (i i) may comprise three heavy chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs: 29, 30, and 31 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light chain CDRs having the amino acid sequences of SEQ ID NOs: 32, 33, and 34, or (ii) three light chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs: 32, 33, and 34 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0196] In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 29, 30, and 31, each having a single conserved amino acid substitution in heavy chain CDR1, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 32, 33, and 34. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 29, 30, and 31, each having a single conserved amino acid substitution in heavy chain CDR2, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 32, 33, and 34. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 29, 30, and 31, each having a single conserved amino acid substitution in heavy chain CDR3, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 32, 33, and 34.

[0197] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 29, 259, and 31, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 29, 259, and 31 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 32, 33, and 34, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 32, 33, and 34 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0198] In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs, SEQ ID NOs. 29, 259, and 31, with a single conserved amino acid substitution in heavy chain CDR1, and a second polypeptide containing three light chain CDRs, SEQ ID NOs. 32, 33, and 34. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs, SEQ ID NOs. 29, 259, and 31, with a single conserved amino acid substitution in heavy chain CDR2, and a second polypeptide containing three light chain CDRs, SEQ ID NOs. 32, 33, and 34. In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs, SEQ ID NOs. 29, 259, and 31, with a single conserved amino acid substitution in heavy chain CDR3, and a second polypeptide containing three light chain CDRs, SEQ ID NOs. 32, 33, and 34.

[0199] In any of the fusion protein constructs described herein, the first polypeptide can comprise three heavy chain CDRs having the amino acid sequences of SEQ ID NOs: 29, 260, and 31, or three heavy chain CDRs having amino acid sequences that differ in that one, two, or three conservative amino acid substitutions are present in one or more of SEQ ID NOs: 29, 260, and 31; the second polypeptide can comprise three light chain CDRs having the amino acid sequences of SEQ ID NOs: 32, 33, and 34, or three light chain CDRs having amino acid sequences that differ in that one, two, or three conservative amino acid substitutions are present in one or more of SEQ ID NOs: 32, 33, and 34. In some embodiments, a single conservative amino acid substitution is present in one or more of the CDRs. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0200] In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs of SEQ ID NOs: 29, 260, and 31 with a single conservative amino acid substitution in heavy chain CDR1, and a second polypeptide comprising three light chain CDRs of SEQ ID NOs: 32, 33, and 34. In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs of SEQ ID NOs: 29, 260, and 31 with a single conservative amino acid substitution in heavy chain CDR2, and a second polypeptide comprising three light chain CDRs of SEQ ID NOs: 32, 33, and 34. In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs of SEQ ID NOs: 29, 260, and 31 with a single conservative amino acid substitution in heavy chain CDR3, and a second polypeptide comprising three light chain CDRs of SEQ ID NOs: 32, 33, and 34.

[0201] In any of the fusion protein constructs described herein, the first polypeptide can comprise three heavy chain CDRs having the amino acid sequences of SEQ ID NOs: 35, 36, and 37, or three heavy chain CDRs having amino acid sequences that differ in that one, two, or three conservative amino acid substitutions are present in one or more of SEQ ID NOs: 35, 36, and 37; the second polypeptide can comprise three light chain CDRs having the amino acid sequences of SEQ ID NOs: 38, 39, and 40, or three light chain CDRs having amino acid sequences that differ in that one, two, or three conservative amino acid substitutions are present in one or more of SEQ ID NOs: 38, 39, and 40. In some embodiments, a single conservative amino acid substitution is present in one or more of the CDRs. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0202] In some embodiments, the fusion protein construct has a first polypeptide that comprises three heavy chain CDRs of SEQ ID NOs: 35, 36, and 37 with a single conservative amino acid substitution in heavy chain CDR1, and a second polypeptide that comprises three light chain CDRs of SEQ ID NOs: 38, 39, and 40. In some embodiments, the fusion protein construct has a first polypeptide that comprises three heavy chain CDRs of SEQ ID NOs: 35, 36, and 37 with a single conservative amino acid substitution in heavy chain CDR2, and a second polypeptide that comprises three light chain CDRs of SEQ ID NOs: 38, 39, and 40. In some embodiments, the fusion protein construct has a first polypeptide that comprises three heavy chain CDRs of SEQ ID NOs: 35, 36, and 37 with a single conservative amino acid substitution in heavy chain CDR3, and a second polypeptide that comprises three light chain CDRs of SEQ ID NOs: 38, 39, and 40.

[0203] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 147, 148, and 149, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 150, 151, and 152 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 147, 148, and 149, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 150, 151, and 152 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0204] In some embodiments, the fusion protein construct comprises a first polypeptide containing three heavy chain CDRs of SEQ ID NOs. 147, 148, and 149, each having a single conserved amino acid substitution in the heavy chain CDR1, and a second polypeptide containing three light chain CDRs of SEQ ID NOs. 150, 151, and 152. The fusion protein construct has two polypeptides. In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs of SEQ ID NOs. 147, 148, and 149, having a single conserved amino acid substitution in the heavy chain CDR2, and a second polypeptide comprising three light chain CDRs of SEQ ID NOs. 150, 151, and 152. In some embodiments, the fusion protein construct has a first polypeptide comprising three heavy chain CDRs of SEQ ID NOs. 147, 148, and 149, having a single conserved amino acid substitution in the heavy chain CDR3, and a second polypeptide comprising three light chain CDRs of SEQ ID NOs. 150, 151, and 152.

[0205] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 188, 199, and 190, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 188, 199, and 190 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 191, 192, and 193, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 191, 192, and 193 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions (substitutions of original residues by histidine) are present in the CDRs.

[0206] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs: 196, 197, and 198, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs: 196, 197, and 198 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs: 199, 200, and 201, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs: 199, 200, and 201 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0207] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 204 or 343, 205, and 206, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 204 or 343, 205, and 206 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 207, 208, and 209, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 207, 208, and 209 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0208] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy chain CDRs having the amino acid sequences of SEQ ID NOs. 212, 213, and 214, or (ii) three heavy chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 212, 213, and 214 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light chain CDRs having the amino acid sequences of SEQ ID NOs. 215, 216, and 217, or (ii) one or more of SEQ ID NOs. 215, 216, and 217 contain one, two, or three conserved amino acid substitutions It may contain three light chain CDRs having amino acid sequences that differ in a point. In some embodiments, one or more CDRs contain a single conservative amino acid substitution. In other embodiments, the CDRs contain one, two, three, or four additional histidine substitutions.

[0209] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 220, 221, and 222, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 220, 221, and 222 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 223, 224, and 225, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 223, 224, and 225 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0210] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) three heavy-chain CDRs having the amino acid sequences of SEQ ID NOs. 228, 229, and 230, or (ii) three heavy-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 228, 229, and 230 contain one, two, or three conserved amino acid substitutions; the second polypeptide may comprise (i) three light-chain CDRs having the amino acid sequences of SEQ ID NOs. 231, 232, and 233, or (ii) three light-chain CDRs having amino acid sequences that differ in that one or more of SEQ ID NOs. 231, 232, and 233 contain one, two, or three conserved amino acid substitutions. In some embodiments, one or more CDRs contain a single conserved amino acid substitution. In other embodiments, one, two, three, or four additional histidine substitutions are present in the CDRs.

[0211] In any of the fusion protein constructs described herein, the first polypeptide may include (i) a heavy chain variable region comprising at least one amino acid sequence from among SEQ ID NOs: 194, 202, 210, 218, 226, 234, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, and 274, and (ii) a light chain variable region comprising at least one amino acid sequence from among SEQ ID NOs: 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, and 278, or (i) SEQ ID NO: 194 (ii) a heavy chain variable region comprising an amino acid sequence that differs in that at least one of 202, 210, 218, 226, 234, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273 and 274 contains one or more conserved amino acid substitutions, and (ii) a light chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277 and 278 contains one or more conserved amino acid substitutions.

[0212] In any of the fusion protein constructs described herein, the first polypeptide may include (i) a humanized heavy chain variable region comprising at least one amino acid sequence from among SEQ ID NOs: 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, and 274, and (ii) a humanized light chain variable region comprising at least one amino acid sequence from among SEQ ID NOs: 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, and 278, or (i) SEQ ID NOs: 246, 247, 248, 249, 250, 25 (ii) A humanized heavy chain variable region comprising an amino acid sequence that differs in that at least one of 1, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, and 274 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, and 278 contains one or more conserved amino acid substitutions.

[0213] In any of the fusion protein constructs described herein, the first polypeptide may include (i) a humanized heavy chain variable region comprising at least one amino acid sequence from SEQ ID NOs: 246, 247, 248, 249, 250, 251, 252, 253, 254, and 255, and (ii) a light chain variable region comprising at least one amino acid sequence from SEQ ID NOs: 256, 257, and 258; or (i) a humanized heavy chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 246, 247, 248, 249, 250, 251, 252, 253, 254, and 255 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 256, 257, and 258 contains one or more conserved amino acid substitutions.

[0214] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 254 and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 258, or may comprise a humanized heavy chain variable region comprising (i) an amino acid sequence that differs in that SEQ ID NO: 254 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that SEQ ID NO: 258 contains one or more conserved amino acid substitutions.

[0215] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 251 and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 258, or may comprise a humanized heavy chain variable region comprising (i) an amino acid sequence that differs in that SEQ ID NO: 251 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that SEQ ID NO: 258 contains one or more conserved amino acid substitutions.

[0216] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain variable region comprising at least one amino acid sequence from SEQ ID NOs: 261, 262, 263, 264, and 265, and (ii) a light chain variable region comprising at least one amino acid sequence from SEQ ID NOs: 266, 267, 268, and 269; or (i) a humanized heavy chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 261, 262, 263, 264, and 265 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 266, 267, 268, and 269 contains one or more conserved amino acid substitutions.

[0217] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 264 and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 269, or may comprise a humanized heavy chain variable region comprising (i) an amino acid sequence that differs in that SEQ ID NO: 264 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that SEQ ID NO: 269 contains one or more conserved amino acid substitutions.

[0218] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 264 and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 268, or may comprise a humanized heavy chain variable region comprising (i) an amino acid sequence that differs in that SEQ ID NO: 264 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that SEQ ID NO: 268 contains one or more conserved amino acid substitutions.

[0219] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain variable region comprising the amino acid sequence of SEQ ID NO: 263 and (ii) a light chain variable region comprising the amino acid sequence of SEQ ID NO: 269, or may comprise a humanized heavy chain variable region comprising (i) an amino acid sequence that differs in that SEQ ID NO: 263 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain variable region comprising an amino acid sequence that differs in that SEQ ID NO: 269 contains one or more conserved amino acid substitutions.

[0220] In any of the fusion protein constructs described herein, the first polypeptide may comprise (i) a humanized heavy chain sequence comprising at least one amino acid sequence from SEQ ID NOs: 280, 281, 282, 237, 283, 284, 285, and 286, and (ii) a light chain sequence comprising the amino acid sequence of SEQ ID NOs: 279, or (i) a humanized heavy chain sequence comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 280, 281, 282, 237, 283, 284, 285, and 286 contains one or more conserved amino acid substitutions, and (ii) a humanized light chain sequence comprising an amino acid sequence that differs in that one or more conserved amino acid substitutions are present in SEQ ID NOs: 279.

[0221] In some embodiments, the fusion protein constructs described herein include a humanized heavy chain sequence comprising at least one amino acid sequence of SEQ ID NO: 280, 281, 282, 283, 284, 285, or 286, or a humanized heavy chain sequence comprising an amino acid sequence that differs in that it has one or more conservative amino acid substitutions in at least one of SEQ ID NO: 280, 281, 282, 237, 283, 284, 285, and 286. In some embodiments, the fusion protein constructs described herein include a humanized light chain sequence comprising the amino acid sequence of SEQ ID NO: 279, or a humanized light chain sequence comprising an amino acid sequence that differs in that it has one or more conservative amino acid substitutions in SEQ ID NO: 279.

[0222] In any of the fusion protein constructs described herein, the first polypeptide can include (i) a mouse heavy chain sequence comprising at least one amino acid sequence of SEQ ID NO: 73, 288, 244, 290, or 342, and (ii) a mouse light chain sequence comprising at least one amino acid sequence of SEQ ID NO: 68, 287, 59, or 289, or (i) a mouse heavy chain sequence comprising an amino acid sequence that differs in that it has one or more conservative amino acid substitutions in at least one of SEQ ID NO: 73, 288, 244, 290, and 342, and (ii) a mouse light chain sequence comprising an amino acid sequence that differs in that it has one or more conservative amino acid substitutions in at least one of SEQ ID NO: 68, 287, 59, and 289.

[0223] The fusion protein constructs described herein include, in some embodiments, a mouse heavy chain sequence comprising at least one amino acid sequence from SEQ ID NOs: 73, 288, 244, 290, and 342, or a mouse heavy chain sequence comprising an amino acid sequence that differs in that at least one of SEQ ID NOs: 73, 288, 244, 290, and 342 contains one or more conserved amino acid substitutions. The fusion protein constructs described herein include, in some embodiments, a mouse light chain sequence comprising at least one amino acid sequence from SEQ ID NOs: 68, 287, 59, and 289, or an amino acid sequence that differs in that at least one of SEQ ID NOs: 68, 287, 59, and 289 contains one or more conserved amino acid substitutions. Includes mouse light chain sequences containing columns.

[0224] Conservative amino acid substitutions are substitutions that change a given amino acid to a different amino acid with similar biochemical properties (e.g., charge, hydrophobicity, size). For example, any one of the following categories of amino acids is considered a conservative amino acid substitution: polar (hydrophilic) neutral amino acids: serine (Ser), threonine (Thr), cysteine ​​(Cys), histidine (His), asparagine (Asn), glutamine (Gln), and tyrosine (Tyr); polar and negatively charged amino acids: aspartic acid (Asp), glutamic acid (Glu); polar and positively charged amino acids: histidine (His), lysine (Lys), arginine (Arg); hydrophobic amino acids: glycine (Gly), alanine (Ala), valine (Val). ), leucine (Leu), isoleucine (Ile), proline (Pro), phenylalanine (Phe), methionine (Met), and tryptophan (Trp); hydrophobic amino acids with aliphatic side chains: alanine (Ala), valine (Val), leucine (Leu), isoleucine (Ile); hydrophobic amino acids with aromatic side chains: phenylalanine (Phe), tryptophan (Trp); small amino acids: alanine (Ala), cysteine ​​(Cys), glycine (Gly), proline (Pro), serine (Ser), and threonine (Thr).

[0225] Conjugation of target-directing portion and complement regulator The linkage between the target-directed moiety and the complement modulator may include (1) conjugation by direct fusion of two protein sequences, or (2) fusion by an intervening linker / linker sequence / spacer / tether-forming sequence. As used herein, the terms “linker,” “linker sequence,” “spacer,” or “tether-forming sequence” may mean a molecule or group of molecules (e.g., monomer or polymer) that links two molecules and often serves to position them in a favorable configuration. Several methods may be used to covalently link molecules together. These include, but are not limited to, polypeptide linkage between the N-terminus and C-terminus of a protein or protein domain, linkage via disulfide bonds, and linkage via chemical crosslinking reagents. In one embodiment, the linker is a peptide bond produced by recombinant technology or peptide synthesis.

[0226] In some examples, the complement regulator and the target-directing moiety may be conjugated via a linker peptide, such as a linker peptide that can directly link the target-directing moiety and the complement regulator. The linker peptide may contain amino acid residues that provide flexibility. Therefore, the linker peptide may contain the following amino acid residues: glycine, serine, alanine, or threonine. The linker peptide must be long enough to link the two molecules in such a way that they adopt the correct conformation relative to each other so that they maintain the desired activity. A length suitable for this purpose may include at least 1 to about 100 amino acid residues or more. The linker may be 1 to 30 amino acids long. The linker may be about 1 to 20 amino acids long. The linker peptide may be included as a spacer between two protein moieties. The linker peptide may promote proper protein folding, stability, expression, and biological activity of the constituent protein moieties. Long, flexible linker peptides may consist of glycine, serine, or threonine, with multiple glycine residues providing extremely flexible conformations. Serine or threonine residues provide polar surface regions that limit hydrophobic interactions with the peptide or its constituent fusion protein moiety. The amino acid residues selected for inclusion in the linker peptide may exhibit properties that do not significantly interfere with polypeptide activity. Therefore, the linker peptide may not exhibit a charge that is inconsistent with polypeptide activity, interferes with internal folding, or forms binding or other interactions with amino acid residues in target-directed moieties or complement modifiers, thereby seriously interfering with the binding of the moiety to its target. The non-limiting range of sequences that can act as linkers Examples may include short peptides with a length of approximately 2 to 15 amino acids. Among the peptide sequences that can be used as linkers in this disclosure, n=0, 1, 2, 3, 4, 5, 6, 7, or 8 (Gly-Ser) are particularly used. n (Sequence code 292); n=0, 1, 2, 3, or 4 (GlyGlyGlySer) n (Sequence code 293); n=0, 1, 2, 3, or 4 (GlySerSerGly) n(SEQ ID NO: 294) is an example. In some fusion protein constructs, the linker sequence is (GlyGlyGlyGlySerGlyGlyGlyGlySer)(SEQ ID NO: 138). In some cases, tethered blockers for shaker potassium channels can be used, including glycine-alanine polymers, alanine-serine polymers, and other flexible linkers, such as panels of quaternary ammonium (QA) linked to maleimide by polyglycine tethers of varying lengths (see, e.g., TJMorin and WRKobertz Tethering Chemistry and K+ Channels J.Biol.Chem. 283(37):25105-25109 (2008)), and a variety of other flexible linkers. Glycine-serine polymers can be used because neither amino acid is relatively organized and therefore can potentially serve as a neutral tether between the components. Secondly, serine is hydrophilic and therefore can solubilize what may be spherical glycine chains. In some cases, the linker may contain the sequence shown in any one of sequence numbers 171-193, where in some cases n can be at least 4, and in other cases n can be 1-8 or 1-5. For example, in sequence number 161 n can be at least 4, in sequence number 164 n can be 1-8, in sequence number 165 n can be 1-5, and in sequence number 166 n can be 1-5. In sequence number 168 X can be A (alanine), K (lysine), or E (glutamic acid), where n can be 5-17. In some cases, in sequence numbers 161, 164, 165, 166, 168, 170, 171, 174, and 179 n can be in the range of 1-17, 1-8, 1-5, at least 4, or 5-17.

[0227] Suitable linkers can also be identified by screening a database of known three-dimensional structures to find naturally occurring motifs that can fill the gap between two polypeptide chains, such as linkers derived from naturally occurring multidomain proteins. In some cases, the linker is not immunogenic when administered to human patients. Thus, linkers that have low immunogenicity, or are thought to have low immunogenicity, can be selected. For example, linkers that are naturally present in humans can be selected. In some cases, the linker may have a sequence of the hinge region of the antibody, which is the sequence that connects the Fab and Fc regions of the antibody, or the linker may have a sequence that includes a part of the hinge region of the antibody, or a sequence that is substantially similar to the hinge region. Another way to obtain a suitable linker is to use a simple linker, e.g., (Gly4Ser) n This is achieved by optimizing (SEQ ID NO: 295) by random mutagenesis. Alternatively, a suitable polypeptide linker may be identified first, and then further linker polypeptides may be created to select amino acids that more optimally interact with the domain to be linked. Other types of linkers that may be used in the present invention include artificial polypeptide linkers and inteins. In some cases, the complement regulator and the target-directing moiety may be conjugated using enzymatic site-directed conjugation methods, including the use of mammalian or bacterial transglutaminase enzymes. Microbial transglutaminases (mTGs) are versatile tools in modern research and biotechnology. The availability of relatively pure enzymes in large quantities, ease of use, and lack of regulation by calcium and guanosine-5'-triphosphate (GTP) have made mTGs the primary crosslinking enzyme used in both the food industry and biotechnology. Currently, mTGs are used in many applications to conjugate proteins and peptides to small molecules, polymers, surfaces, DNA, and other proteins. For example, Pavel Strop, Veracity of microbial transglutaminase, Bioconjugate Chem. 25(5) See pages 855-862.

[0228] In some examples, fusion protein constructs are provided in which a constant region contains a target-directed moiety containing receptor glutamine, which can later be conjugated to a complement protein via a lysine linker (e.g., any primary amine chain that is a substrate of the TG-ase, e.g., including alkylamines and oxoamines), and conjugation occurs exclusively on one or more receptor glutamine residues present in the target-directed moiety outside the antigen-binding site (e.g., outside the variable region, within the constant region). Therefore, conjugation does not occur on glutamine within the variable region, e.g., glutamine with at least partially exposed surfaces. The conjugate can be formed by reacting the target-directed moiety with the lysine linker in the presence of a TG-ase. The lysine linker may contain, for example, a primary amine in addition to, e.g., alkylamines, oxoamines, peptides, polypeptides, any organic molecule, a drug or diagnostic moiety, or a reactive moiety that can later react with a complement modifier.

[0229] In another embodiment, a disulfide bond can be designed to connect two molecules. In some cases, the linker is a chemical crosslinking agent. For example, a variety of bifunctional protein coupling agents can be used, including but not limited to N-succinimidyl-3-(2-pyridyldithiol)propionate (SPDP), succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate, iminothiolane (IT), difunctional derivatives of imide esters (e.g., dimethyl HCl adipiimidoate), active esters (e.g., disuccinimidyl sberate), aldehydes (e.g., glutaraldehyde), bisazide compounds (e.g., bis(p-azidobenzoyl)hexanediamine), bisdiazonium derivatives (e.g., bis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (e.g., toluene 2,6-diisocyanate), and bis-active fluorine compounds (e.g., 1,5-difluoro-2,4-dinitrobenzene). For example, lysine immunotoxins can be prepared as described in Vitetta et al., 1971, Science 238:1098. Chemical linkers can enable isotopic chelation. For example, carbon-14 labeled 1-isothiocyanatobenzyl-3-methyldiethylenetriaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent for conjugating radioactive nucleotides to antibodies (see, e.g., WO94 / 11026). Linkers may be cleavable, thereby facilitating the release of cytotoxic agents in cells. For example, acid-unstable linkers, peptidase-sensitive linkers, dimethyl linkers, or disulfide-containing linkers (Chari et al., 1992, Cancer Research 52:127-131) may be used. Alternatively, various non-proteinaceous polymers, such as but not limited to polyethylene glycol (PEG), polypropylene glycol, polyoxyalkylene, or copolymers of polyethylene glycol and polypropylene glycol, may be used as linkers, that is, they may be used to link the target-directed moieties of the Disclosure to fusion or complex partners, such as complement modifiers, to create the fusion protein constructs of the Disclosure.

[0230] When the target-directing portion and the active portion are directly joined, a hybrid vector may be created in which the DNA encoding the target-directing portion and the complement regulator are directly ligated to each other. When a linker is used, a hybrid vector may be created in which the DNA encoding the target-directing portion is ligated to the DNA encoding one end of the linker portion, and the DNA encoding the complement regulator is ligated to the other end of the linker portion. Such ligation may be performed sequentially or as a three-component ligation.

[0231] Methods for manufacturing, refining, and characterization The fusion protein constructs described herein can be manufactured using a variety of techniques. For example, The nucleic acid encoding the fusion protein construct described herein may be inserted into an expression vector containing, for example, a transcription and translation regulatory sequence including a promoter sequence, a ribosome binding site, transcription start and stop sequences, translation start and stop sequences, a transcription termination signal, a polyadenylation signal, and an enhancer or activation sequence. The regulatory sequence includes the promoter, as well as the transcription start and stop sequences. In addition, the expression vector may include one or more replication systems that allow its control in two different organisms, for example, mammalian or insect cells for expression and a prokaryotic host for cloning and amplification.

[0232] Several suitable vector systems are available for the expression of fusion proteins from nucleic acids in mammalian cells. One type of vector relies on the integration of the desired gene sequence into the host cell genome. Cells with stably integrated DNA can be selected by simultaneously introducing drug resistance genes such as E. coli gpt (see Mulligan and Berg Proc. Natl. Acad. Sci. USA 78:2072 (1981)) or Tn5 neo (see Southern and Berg Mol. Appl. Genet. 1:327 (1982)). The selection marker gene can be either ligated to the DNA gene sequence to be expressed or introduced into the same cell by co-transfection (Wigler et al. Cell 16:77 (1979)). A second type of vector utilizes DNA elements that confer autonomous replication ability to extrachromosomal plasmids. These vectors may be derived from animal viruses, such as bovine papillomavirus (Sarver et al. Proc. Natl. Acad. Sci. USA, 79:7147 (1982)), polyomavirus (Deans et al. Proc. Natl. Acad. Sci. USA 81:1292 (1984)), or SV40 virus (Lusky and Botchan Nature 293:79 (1981)).

[0233] Expression vectors can be introduced into cells using a method suitable for subsequent nucleic acid expression. The introduction method is generally determined by the type of target cell, as described below. Examples of methods include calcium phosphate precipitation, liposome fusion, lipofection, electroporation, viral infection, dextran-mediated transfection, polyblen-mediated transfection, protoplast fusion, and direct microinjection.

[0234] Suitable host cells for the expression of fusion proteins include yeast, bacteria, insects, plants, and mammalian cells as described above. Of particular interest are bacteria, e.g., E. coli; fungi, e.g., Saccharomyces cerevisiae and Pichia pastoris; insect cells, e.g., SF9; mammalian cell lines (e.g., human cell lines); and primate cell lines (e.g., primate mammalian cells). In some embodiments, the fusion protein may be expressed in Chinese hamster ovary (CHO) cells or in a preferred myeloma cell line, e.g., (NS0). Preferred cell lines also include, for example, HEK cells (human embryonic kidney 293 cells), BHK-21 (baby hamster kidney) cells, 293 (human embryonic kidney) cells, HMEpC (human mammalian epithelial cells), and 3T3 (mouse embryonic fibroblast) cells.

[0235] As will be recognized by those skilled in the art, some complement modulojugates that can be used in this disclosure exist naturally as secreted proteins, as propeptides, or both, together with signal or leader peptides, and undergo further intracellular or extracellular processing. In such cases, the hybrid vectors of this disclosure may include one or more DNA sequences encoding such signal or leader peptides, one or more DNA sequences encoding such propeptide sequences, or both, depending on whether such secretion, processing, or a combination of secretion and processing is desired. Alternatively, the hybrid vectors of this disclosure may include different signal or leader peptides, propeptides, etc. It may include DNA sequences encoding either or both, or sequences selected to optimize the expression and localization of the fusion protein. In most cases, the signal peptide may be omitted, as the target-directing portion will provide sufficient information to direct the complement regulator to the desired tissues and cells in the target body.

[0236] Exemplary production methods may involve expressing the fusion protein constructs described herein in transgenic animals (e.g., transgenic mammals) and purifying them therefrom. For example, the fusion proteins described herein can be produced in transgenic non-human mammals (e.g., rodents, sheep, or goats) and isolated from their milk, as described in, for example, Houdebine Curr. Opin. Biotechnol. 13(6):625-629 (2002), van Kuik-Romeijn et al. Transgenic Res. 9(2):155-159 (2000), and Pollock et al. J Immunol. Methods 231(1-2):147-157 (1999).

[0237] The fusion protein constructs described herein can be produced from cells by culturing host cells transformed with an expression vector containing a nucleic acid encoding an antibody or its antigen-binding fragment, under conditions and for a duration sufficient to enable protein expression. For example, polypeptides expressed in E. coli can be refolded from inclusion bodies (see, e.g., Hou et al. Cytokine 10:319-30 (1998)). Bacterial expression systems and methods of using them are well known in the art (Current Protocols in Molecular Biology, Wiley & Sons, and Molecular Cloning-A). See Laboratory Manual - 3rd Ed., Cold Spring Harbor Laboratory Press, New York (2001). The selection of codons, suitable expression vectors, and suitable host cells will vary depending on the number of factors and can be easily optimized as needed. The fusion protein constructs described herein may be expressed in mammalian cells or in other expression systems, including but not limited to yeast and baculovirus, and in vitro expression systems (see, for example, Kaszubska et al. Protein Expression and Purification 18:213-220 (2000)).

[0238] After expression, the fusion protein construct can be isolated. The terms “purified” or “isolated” applied to any of the proteins described herein (e.g., the fusion protein constructs described herein) may refer to polypeptides isolated or purified from naturally associated components (e.g., proteins or other naturally occurring biomolecules or organic molecules), such as other proteins, lipids, and nucleic acids in the prokaryote expressing the protein. Typically, a polypeptide is purified if it constitutes at least 60% by weight (e.g., at least 65, 70, 75, 80, 85, 90, 92, 95, 97, or 99) of the total protein in the sample. The fusion protein constructs described herein can be isolated or purified in a variety of ways depending on what other components are present in the sample. Standard purification methods include electrophoresis, molecular techniques, immunological techniques, and chromatographic techniques, such as ion-exchange, hydrophobic, affinity, and reverse-phase HPLC chromatography. For example, fusion proteins can be purified using standard anti-fusion protein antibody affinity columns. Ultrafiltration and diafiltration techniques are also useful in conjunction with protein enrichment. For example, Scopes (1994) "Protein Purification, 3 rdSee edition, “Springer-Verlag, New York City, NY.” The required degree of purification will vary depending on the desired application. In some cases, purification of the expressed polypeptide may not be necessary.

[0239] Methods for determining the yield or purity of purified polypeptides may include, for example, the Bradford assay, UV spectroscopy, Biuret protein assay, Lowry protein assay, Amido Black protein assay, high-pressure liquid chromatography (HPLC), mass spectrometry (MS), and gel electrophoresis (e.g., using protein stains such as Coomassie blue or colloidal silver stain).

[0240] In other exemplary manufacturing methods, the entirety or a portion of the fusion protein constructs described herein can be synthesized from scratch using chemical methods. For example, the constituent amino acid sequences can be synthesized by solid-phase technology, cleaved from a resin, purified by preparative high-performance liquid chromatography, and then chemically linked to form the desired polypeptide. The composition of the synthesized peptide can be confirmed by amino acid analysis or sequencing.

[0241] The fusion protein constructs described herein may be evaluated for any one of several desired properties using in vitro or in vivo assays, such as those described herein, either at the time of expression and purification, or after purification following expression. For example, the fusion proteins described herein may be evaluated for their ability to inhibit C5 convertase, as described, for example, in Heinen et al. Factor H-related protein 1 (CFHR-1) inhibits complement C5 convertase activity and terminal complex formation. Blood 114(12):2439-47 (2009). Endotoxins can be removed from fusion protein construct preparations using a variety of commercially available reagents, including but not limited to ProteoSpin® endotoxin removal kits (Norgen Biotek Corporation), Detoxi-Gel endotoxin removal gels (Thermo Scientific, Pierce Protein Research Products), MiraCLEAN® endotoxin removal kits (Mirus), or Acrodisc®-Mustang® E membranes (Pall Corporation).

[0242] Methods for detecting and / or measuring the amount of endotoxins present in a sample (both before and after purification) may be based on commercially available kits. For example, the concentration of endotoxins in a protein sample can be determined using the QCL-1000 colorimetric kit (BioWhittaker), kits based on limulus amebocyte lysate (LAL), such as Pyrotell®, Pyrotell®-T, Pyrochrome®, Chromo-LAL, and CSE kits available from Associates of Cape Cod Incorporated.

[0243] Following expression and purification, the fusion protein constructs described herein may be modified. The modifications may be covalent or non-covalent. Such modifications may be introduced into the fusion protein, for example, by reacting target amino acid residues of the polypeptide with an organic derivatizing agent capable of reacting with selected side-chain or terminal residues. Suitable modification sites may be selected using any of a variety of criteria, including, for example, structural analysis or amino acid sequence analysis of the fusion protein described herein.

[0244] In some exemplary manufacturing methods, the fusion protein constructs described herein may be conjugated to a heterologous moiety. The heterologous moiety may be, for example, a heterologous polypeptide, a therapeutic agent (e.g., a toxin or drug), or a detectable label, such as, but not limited to, a radiolabel, an enzymatic label, a fluorescent label, or a luminescent label. Suitable heterologous polypeptides include, for example, antigenic tags for use in antibody purification (e.g., FLAG, polyhistidine, hemagglutinin (HA), glutathione-S-transferase (GST), or maltose-binding proteins). This may include polypeptides (MBP). Heterogeneous polypeptides may also include polypeptides useful as diagnostic or detectable markers, such as luciferase, green fluorescent protein (GFP), or chloramphenicol acetyltransferase (CAT). If the heterogeneous portion is a polypeptide, that portion may be incorporated into the fusion protein described herein to form the fusion protein.

[0245] Pharmaceutical preparations Pharmaceutical formulations comprising the fusion protein constructs described herein are also provided in this disclosure. Pharmaceutical formulations of the fusion protein constructs of this disclosure are typically prepared as unit injectable dosage forms for parenteral administration, i.e., bolus, intravenous, intratumoral, or subcutaneous injection, together with a pharmaceutically acceptable parenteral vehicle. The fusion protein constructs may optionally be mixed with pharmaceutically acceptable diluents, carriers, excipients, or stabilizers (Remington's Pharmaceutical Sciences (1980) 16th edition, Osol, A. Ed.) in the form of lyophilized formulations or aqueous solutions. Acceptable diluents, carriers, excipients and stabilizers are non-toxic to the recipient at the dosage and concentration used, and include buffers, e.g., phosphates, citrates and other organic acids; antioxidants, including ascorbic acid and methionine; preservatives (e.g., octadecyldimethylbenzylammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkylparabens, e.g., methyl or propylparaben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, e.g. This includes serum albumin, gelatin, or immunoglobulin; hydrophilic polymers, e.g., polyvinylpyrrolidone; amino acids, e.g., glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates, e.g., glucose, mannose, or dextrin; chelating agents, e.g., EDTA; sugars, e.g., sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions, e.g., sodium; metal complexes (e.g., Zn-protein complexes); nonionic surfactants, e.g., TWEEN®, PLURONICS®, or polyethylene glycol (PEG), or any combination thereof.

[0246] Furthermore, the fusion protein construct may be encapsulated in microcapsules prepared, for example, by coacervation technology or interfacial polymerization, such as hydroxymethylcellulose or gelatin microcapsules and poly(methyl methacrylate) microcapsules, in colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules), or in macroemulsions. Such techniques are disclosed in Remington's Pharmaceutical Sciences 16th edition, Osol, A. Ed. (1980).

[0247] Sustained-release formulations may be prepared. A preferred example of a sustained-release formulation is a semipermeable matrix of a solid hydrophobic polymer containing a fusion protein construct, the semipermeable matrix being in the form of a molded article, such as a thin film or microcapsule. Examples of sustained-release matrices include polyesters, hydrogels (e.g., poly(2-hydroxyethyl methacrylate) or poly(vinyl alcohol)), polylactide (U.S. Patent No. 3,773,919), copolymers of L-glutamic acid and gamma-ethyl-L-glutamate, non-degradable ethylene vinyl acetate, degradable lactic acid-glycolic acid copolymers, such as LUPRON DEPOT (trademark) (injectable microspheres consisting of lactic acid-glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3-hydroxybutyric acid.

[0248] Formulations used for in vivo administration are sterile, and this may involve filtration using a sterile filtration membrane. Formulations include those suitable for the above-mentioned routes of administration. Formulations may be conveniently provided in unit dosage forms and may be prepared by any of the methods commonly used in the preparation of pharmaceutical formulations as unit doses. The techniques and formulations can be broadly found in Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, Pa.). Such methods involve the step of combining the active ingredient with a carrier constituting one or more auxiliary components. Generally, formulations are prepared by homogeneously and densely combining a fusion protein construct with a liquid carrier or a micronized solid carrier or both, and, if necessary, subsequently shaking the product.

[0249] The aqueous suspensions of this disclosure contain a fusion protein construct as a miscion with an excipient suitable for the preparation of an aqueous suspension. Such excipients include suspending agents, such as sodium carboxymethylcellulose, croscarmellose, povidone, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, tragacanth gum, and acacia gum, as well as dispersants or wetting agents, such as naturally occurring phosphatides (e.g., lecithin), condensation products of alkylene oxides and fatty acids (e.g., polyoxyethylene stearate), condensation products of ethylene oxides and long-chain aliphatic alcohols (e.g., heptadecaethyleneoxycetanol), and condensation products of ethylene oxides and partial esters derived from fatty acids and anhydrous hexitol (e.g., polyoxyethylene sorbitan monooleate). The aqueous suspension may also include one or more preservatives, such as ethyl p-hydroxybenzoate or n-propyl, one or more colorants, one or more flavorings, and one or more sweeteners, such as sucrose or saccharin.

[0250] Pharmaceutical formulations containing fusion protein constructs may be in the form of sterile injectable formulations, such as sterile aqueous or oily injectable suspensions. These suspensions may be formulated according to the methods using the preferred dispersants or wetting agents and suspending agents described above. Sterile injectable formulations may also be sterile injectable solutions or suspensions prepared with non-toxic, parenterally acceptable diluents or solvents, such as a solution made with 1,3-butane-diol, or prepared as lyophilized powders. Among the acceptable vehicles and solvents that may be employed are, in particular, water, Ringer's solution, and isotonic sodium chloride solution. In addition, it may be advantageous to employ sterile fixative oils as solvents or suspension media. For this purpose, any sterile fixative oil, including synthetic mono- or diglycerides, may be employed. Furthermore, fatty acids, such as oleic acid, may also be used in the preparation of injectable formulations.

[0251] The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will vary depending on the host being treated and the specific mode of administration. For example, an aqueous solution intended for intravenous infusion may contain approximately 3 to 500 μg of active ingredient per milliliter of solution to produce a suitable infusion rate of approximately 30 mL / hour.

[0252] Formulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions that may contain antioxidants, buffers, bacteriostatic agents, and solutes that are isotonic with the blood of the recipient to whom the formulation is intended; as well as aqueous and non-aqueous sterile suspensions that may contain suspending agents and thickeners.

[0253] Oral administration of protein-based therapeutic agents is generally avoided due to hydrolysis or denaturation in the stomach. However, formulations of fusion protein constructs suitable for oral administration can be prepared as discontinuous units, such as capsules, cachets, or tablets, each containing a predetermined amount of the fusion protein construct.

[0254] The formulation may be packed in single-dose or multi-dose containers, such as sealed ampoules and vials, and may be stored under lyophilization conditions requiring only the addition of a sterile liquid carrier for injection, such as water, immediately before use. In some cases, the formulation may be delivered, for example, by controlled delivery. The product may be packaged in an infusion device such as an infusion pump. Instantaneous injection solutions and suspensions are prepared from sterile powders, granules, and tablets of the above class. Preferred unit dose formulations contain the daily dose or unit daily sub-dose or an appropriate portion thereof of the active ingredient as listed above in this specification.

[0255] This disclosure further provides animal compositions comprising at least one active ingredient as defined above, together with an animal carrier therefor. The animal carrier is a material useful for the purpose of administering the composition and may be a solid, liquid, or gaseous material, or it may be inert or acceptable in the field of veterinary medicine and compatible with the active ingredient. These animal compositions may be administered parenterally, orally, or by any other desired route.

[0256] Methods for treating complement-related disorders, conditions, or symptoms The fusion protein constructs described herein may be used to treat a variety of complement-related disorders, including, but not limited to, ischemia-reperfusion injury, rheumatoid arthritis (RA); lupus nephritis; atypical hemolytic uremic syndrome (aHUS); typical or infectious hemolytic uremic syndrome (tHUS); dense deposit disease (DDD); paroxysmal nocturnal hemoglobinuria (PNH); multiple sclerosis (MS); macular degeneration (e.g., age-related macular degeneration (AMD), geographic atrophy (also known as atrophic age-related macular degeneration or progressive dry AMD); hemolytic, elevated liver enzymes, and hypothrombocytopenic (HELLP) syndrome; sepsis; dermatomyositis; diabetic retinopathy; thrombotic thrombocytopenic purpura (TTP); spontaneous miscarriage; microimmunovasculitis; epidermolysis bullosa; recurrent miscarriage; multiple sclerosis (MS); and traumatic brain injury.

[0257] Furthermore, complement-mediated vascular disorders, such as, but not limited to, cardiovascular disorders, myocarditis, cerebrovascular disorders, peripheral (e.g., musculoskeletal) vascular disorders, renal vascular disorders, mesenteric / intestinal vascular disorders, vascular regeneration for grafts and / or replants, vasculitis, Henoch-Schönlein purpura nephritis, lupus erythematosus-associated vasculitis, vasculitis associated with rheumatoid arthritis, immune complex vasculitis, Takayasu's arteriovenous arthritis, capillary leak syndrome, dilated cardiomyopathy, diabetic vascular disorders, thoracoabdominal aortic aneurysms, Kawasaki disease (arthritis), venous gas embolism (VGE), and restenosis after stent placement, rotational atherosclerosis, and percutaneous transluminal coronary angioplasty (PTCA), can also be treated using the fusion protein constructs of this disclosure.

[0258] Complement-related disorders can also be myasthenia gravis, cold agglutinin disease (CAD), paroxysmal cold hemoglobinuria (PCH), idiopathic inflammatory myopathy including dermatomyositis and polymyositis, scleroderma, warm autoimmune hemolytic anemia, Graves' disease, Hashimoto's thyroiditis, type 1 diabetes mellitus, psoriasis, pemphigus, autoimmune hemolytic anemia (AIHA), idiopathic thrombocytopenic purpura (ITP), Goodpasture syndrome, antiphospholipid syndrome (APS), Degos disease, and fulminant APS (CAPS).

[0259] Ischemia-reperfusion (IR) injury can refer to tissue damage that occurs when blood supply is restored to a tissue after a period of ischemia (restriction of blood supply). Deficiency of oxygen and nutrients from the blood creates conditions in which the restoration of circulation leads to inflammation and oxidative damage rather than a restoration of normal function. Ischemia-reperfusion injury is a significant medical problem and can be associated with traumatic injuries, including hemorrhagic shock, and m...

Claims

1. A monomeric fusion protein construct that binds to complement-related antigens, a) A first polypeptide comprising domains A and B aligned in an A-B orientation from the N-terminus to the C-terminus, b) Second polypeptide containing domains E and F aligned in an E-F orientation from the N-terminus to the C-terminus. The domains include, and at least one of the domains A, B, E, or F is conjugated to domain R. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (v) The domain F contains the light chain constant region amino acid sequence (CL1), The monomeric fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

2. The monomeric fusion protein construct according to claim 1, wherein the first polypeptide comprises domain A, domain B, and domain R, the domains of the first polypeptide are arranged in an R-A-B orientation from the N-terminus to the C-terminus, and domain R and domain A are conjugated.

3. The monomeric fusion protein construct according to claim 1, wherein the first polypeptide comprises domain A, domain B, and domain R, the domains of the first polypeptide are arranged in an A-B-R orientation from the N-terminus to the C-terminus, and domain B and domain R are conjugated.

4. The monomeric fusion protein construct according to claim 1, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an R-E-F orientation from the N-terminus to the C-terminus, and domain E and domain R are conjugated.

5. The monomeric fusion protein construct according to claim 1, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an E-F-R orientation from the N-terminus to the C-terminus, and domain F and domain R are conjugated.

6. A monomeric fusion protein construct according to claim 2, 3, 4, or 5, further comprising a second complement regulatory polypeptide, wherein the second complement regulatory polypeptide and the domain R are the same or different.

7. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide and a second polypeptide, a) The first polypeptide comprises domain A, domain B, domain R, hinge region, domain C and domain D, The domains A, B, hinge region, C, and D of the first polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. (i) Domain A is a heavy chain variable region amino acid sequence (VH) or its antigen-binding properties It contains fragments, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain D contains a heavy chain CH3 constant region amino acid sequence, (v) The domain R contains a complement regulator polypeptide, (1) Domain A and Domain R are combined, or (2) Domain D and Domain R are combined, b) The second polypeptide comprises domain E and domain F, The domains E and F of the second polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

8. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain D and domain R are conjugated in at least one of the first polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 7.

9. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain A and domain R are conjugated in at least one of the first polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 7.

10. A fusion protein construct according to claim 8 or 9, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

11. The fusion protein construct according to claim 10, wherein domain A and domain R are conjugated, and domain D and domain R1 are conjugated.

12. The fusion protein according to claim 10, wherein domain D and domain R are conjugated, and domain A and domain R1 are conjugated.

13. c) (i) Domain A, Domain B, Hinge region, Domain C and Domain D, (ii) Domain A, Domain B, Hinge region and Domain C, or (iii) Domain A, Domain B and Hinge region, a third polypeptide comprising these domains. The third polypeptide further comprises the domains A, B, hinge region, C, and D of the third polypeptide arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus, and d) A fourth polypeptide including domains E and F The fourth polypeptide further comprises the domains E and F of the fourth polypeptide being aligned in an E-F orientation from the N-terminus to the C-terminus. The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides each have one or more disulfide bonds in the hinge region. They are connected through each other. The fusion protein construct according to claim 7, which is a tetravalent heterodimer type fusion protein construct.

14. The fusion protein construct according to claim 13, wherein the domain D and the domain R are conjugated in the first polypeptide.

15. The fusion protein construct according to claim 13, wherein the domain A and the domain R are conjugated in the first polypeptide.

16. A fusion protein construct according to claim 14 or 15, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

17. The fusion protein construct according to claim 16, wherein the domain R1 is conjugated to the first polypeptide.

18. The fusion protein construct according to claim 16, wherein the domain R1 is conjugated to the second polypeptide.

19. The fusion protein construct according to claim 16, wherein the domain R1 is conjugated to the third polypeptide.

20. The fusion protein construct according to claim 16, wherein the domain R1 is conjugated to the fourth polypeptide.

21. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide and a second polypeptide, a) The first polypeptide comprises domain A, domain B, domain R, hinge region and domain C, The domains A, B, hinge region, and C of the first polypeptide are arranged in the orientation A-B-hinge region-C from the N-terminus to the C-terminus. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain R contains a complement regulator polypeptide, (1) Domain A and Domain R are combined, or (2) Domain C and Domain R are combined, b) The second polypeptide comprises domain E and domain F, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

22. A tetravalent homodimer fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein at least one of the first polypeptides contains the same The fusion protein construct according to claim 21, wherein in C and domain R are conjugated, and the two first polypeptides are linked together in the hinge region via one or more disulfide bonds.

23. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain A and domain R are conjugated in at least one of the first polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 21.

24. The fusion protein construct according to claim 21, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

25. The fusion protein construct according to claim 24, wherein domain A and domain R are conjugated, and domain C and domain R1 are conjugated.

26. The fusion protein according to claim 24, wherein the domain C and the domain R are conjugated, and the domain A and the domain R1 are conjugated.

27. c) (i) Domain A, Domain B, Hinge region, Domain C and Domain D, (ii) Domain A, Domain B, Hinge region and Domain C, or (iii) Domain A, Domain B and Hinge region, a third polypeptide comprising these domains. The third polypeptide further comprises the domains A, B, hinge region, C, and D of the third polypeptide arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus, and d) A fourth polypeptide including domains E and F The fourth polypeptide further comprises the domains E and F of the fourth polypeptide being aligned in an E-F orientation from the N-terminus to the C-terminus. The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds. The fusion protein construct according to claim 21, which is a tetravalent heterodimer type fusion protein construct.

28. The fusion protein construct according to claim 27, wherein the domain C and the domain R are conjugated in the first polypeptide.

29. The fusion protein construct according to claim 27, wherein the domain A and the domain R are conjugated in the first polypeptide.

30. A fusion protein construct according to claim 28 or 29, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

31. The fusion protein construct according to claim 30, wherein the domain R1 is conjugated to the first polypeptide.

32. The fusion protein construct according to claim 30, wherein the domain R1 is conjugated to the second polypeptide.

33. The fusion protein construct according to claim 30, wherein the domain R1 is conjugated to the third polypeptide.

34. The fusion protein construct according to claim 30, wherein the domain R1 is conjugated to the fourth polypeptide.

35. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide and a second polypeptide, a) The first polypeptide comprises domain A, domain B, domain R and a hinge region, The domains A, B and hinge region of the first polypeptide are arranged in an A-B-hinge region orientation from the N-terminus to the C-terminus. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (1) Domain A and Domain R are combined into one, or (2) The hinge region and Domain R are combined into one. b) The second polypeptide comprises domain E and domain F, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The fusion protein wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

36. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain A and domain R are conjugated in at least one of the first polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 35.

37. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein the hinge region and the domain R are conjugated in at least one of the first polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 35.

38. A fusion protein construct according to claim 36 or 37, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

39. The fusion protein construct according to claim 38, wherein the domain A and the domain R are conjugated, and the hinge domain and the domain R1 are conjugated.

40. The fusion protein according to claim 38, wherein the hinge domain and domain R are conjugated, and domain A and domain R1 are conjugated.

41. c) (i) Domain A, Domain B, Hinge region, Domain C and Domain D, (ii) Domain A, Domain B, Hinge region and Domain C, or (iii) Domain A, Domain B and Hinge region, a third polypeptide comprising these domains. The third polypeptide further comprises the domains A, B, hinge region, C, and D of the third polypeptide arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus, and d) A fourth polypeptide including domains E and F The fourth polypeptide further comprises the domains E and F of the fourth polypeptide being aligned in an E-F orientation from the N-terminus to the C-terminus. The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds. The fusion protein construct according to claim 35, which is a tetravalent heterodimer type fusion protein construct.

42. The fusion protein construct according to claim 41, wherein the hinge domain and the domain R are conjugated in the first polypeptide.

43. The fusion protein construct according to claim 41, wherein the domain A and the domain R are conjugated in the first polypeptide.

44. A fusion protein construct according to claim 42 or 43, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

45. The fusion protein construct according to claim 44, wherein the domain R1 is conjugated to the first polypeptide.

46. The fusion protein construct according to claim 44, wherein the domain R1 is conjugated to the second polypeptide.

47. The fusion protein construct according to claim 44, wherein the domain R1 is conjugated to the third polypeptide.

48. The fusion protein construct according to claim 44, wherein the domain R1 is conjugated to the fourth polypeptide.

49. A trivalent heterodimeric fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region, domain C, domain D and domain R, The domains A, B, hinge region, C, and D of the first polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. (1) Domain A and Domain R are combined, or (2) Domain D and Domain R are combined, b) The second polypeptide comprises domain E and domain F, The domains E and F of the second polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. c) The third polypeptide comprises (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, and (iii) includes domain A, domain B and hinge region, The domains A, B, hinge region, C, and D of the third polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. d) The fourth polypeptide comprises domain E and domain F, The domains E and F of the fourth polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. The domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds, The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain C contains the heavy chain CH2 constant region amino acid sequence, (v) The domain D contains a heavy chain CH3 constant region amino acid sequence, (vi) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (vii) The domain F contains the light chain constant region amino acid sequence (CL1), The aforementioned trivalent heterodimer fusion protein construct.

50. The trivalent heterodimer fusion protein construct according to claim 49, wherein the domain A and the domain R of the first polypeptide are conjugated.

51. The trivalent heterodimer fusion protein construct according to claim 49, wherein the domain D and the domain R of the first polypeptide are conjugated.

52. A trivalent heterodimeric fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region, domain C and domain R, The domains A, B, hinge region, and C of the first polypeptide are arranged in the orientation A-B-hinge region-C from the N-terminus to the C-terminus. (1) Domain A and Domain R are combined, or (2) Domain C and Domain R are combined, b) The second polypeptide comprises domain E and domain F, The domains E and F of the second polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. c) The third polypeptide comprises (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region. The domains A, B, hinge region, C, and D of the third polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. d) The fourth polypeptide comprises domain E and domain F, The domains E and F of the fourth polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. The domain B of the first polypeptide and the domain F of the second polypeptide are 1 They are connected by more than one disulfide bond, The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain C contains the heavy chain CH2 constant region amino acid sequence, (v) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (vi) The trivalent heterodimer fusion protein construct wherein domain F contains the light chain constant region amino acid sequence (CL1).

53. The trivalent heterodimer fusion protein construct according to claim 52, wherein the domain A and the domain R are conjugated in the first polypeptide.

54. The trivalent heterodimer fusion protein construct according to claim 52, wherein the domain C and the domain R are conjugated in the first polypeptide.

55. A trivalent heterodimeric fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region and domain R, The domains A, B and hinge region of the first polypeptide are arranged in an A-B-hinge region orientation from the N-terminus to the C-terminus. (1) Domain A and Domain R are combined into one, or (2) The hinge region and Domain R are combined into one. b) The second polypeptide comprises domain E and domain F, The domains E and F of the second polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. c) The third polypeptide comprises (i) domain A, domain B, hinge region, domain C and domain D, (ii) domain A, domain B, hinge region and domain C, or (iii) domain A, domain B and hinge region. The domains A, B, hinge region, C, and D of the third polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. d) The fourth polypeptide comprises domain E and domain F, The domains E and F of the fourth polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. The domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds, The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (v) The domain F contains the light chain constant region amino acid sequence (CL1), The aforementioned trivalent heterodimer fusion protein construct.

56. The trivalent heterodimer fusion protein construct according to claim 55, wherein the domain A and the domain R are conjugated in the first polypeptide.

57. The trivalent heterodimer fusion protein construct according to claim 55, wherein the hinge region and the domain R are conjugated in the first polypeptide.

58. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide and a second polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region, domain C and domain D, The domains A, B, hinge region, C, and D are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain D contains a heavy chain CH3 constant region amino acid sequence, b) The second polypeptide comprises domain E, domain F and domain R, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (1) Domain E and Domain R are combined, or (2) Domain F and Domain R are combined, (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

59. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein the domain E and the domain R are conjugated in at least one of the second polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 58.

60. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain F and domain R are conjugated in at least one of the second polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 58.

61. A fusion protein construct according to claim 58 or 59, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

62. The fusion protein construct according to claim 61, wherein the domain E and the domain R are conjugated, and the domain F and the domain R1 are conjugated.

63. The fusion protein according to claim 61, wherein the domain F and the domain R are conjugated, and the domain E and the domain R1 are conjugated.

64. c) (i) Domain A, Domain B, Hinge region, Domain C and Domain D, (ii) Domain A, Domain B, Hinge region and Domain C, or (iii) Domain A, Domain B and Hinge region, a third polypeptide comprising these domains. The third polypeptide further comprises the domains A, B, hinge region, C, and D of the third polypeptide arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus, and d) A fourth polypeptide including domains E and F The fourth polypeptide further comprises the domains E and F of the fourth polypeptide being aligned in an E-F orientation from the N-terminus to the C-terminus. The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds. The fusion protein construct according to claim 58, which is a tetravalent heterodimer type fusion protein construct.

65. The fusion protein construct according to claim 64, wherein the domain E and the domain R are conjugated in the first polypeptide.

66. The fusion protein construct according to claim 64, wherein the domain F and the domain R are conjugated in the first polypeptide.

67. A fusion protein construct according to claim 65 or claim 66, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

68. The fusion protein construct according to claim 67, wherein the domain R1 is conjugated to the first polypeptide.

69. The fusion protein construct according to claim 67, wherein the domain R1 is conjugated to the second polypeptide.

70. The fusion protein construct according to claim 67, wherein the domain R1 is conjugated to the third polypeptide.

71. The fusion protein construct according to claim 67, wherein the domain R1 is conjugated to the fourth polypeptide.

72. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide and a second polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region and domain C, The domains A, B, hinge region, and C are arranged in the orientation A-B-hinge region-C from the N-terminus to the C-terminus. (i) Domain A is a heavy chain variable region amino acid sequence (VH) or its antigen-binding properties It contains fragments, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, b) The second polypeptide comprises domain E, domain F and domain R, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (1) Domain E and Domain R are combined, or (2) Domain F and Domain R are combined, (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

73. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein the domain E and the domain R are conjugated in at least one of the second polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 72.

74. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain F and domain R are conjugated in at least one of the second polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 72.

75. A fusion protein construct according to claim 73 or 74, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

76. The fusion protein construct according to claim 75, wherein the domain E and the domain R are conjugated, and the domain F and the domain R1 are conjugated.

77. The fusion protein according to claim 75, wherein the domain F and the domain R are conjugated, and the domain E and the domain R1 are conjugated.

78. c) (i) Domain A, Domain B, Hinge region, Domain C and Domain D, (ii) Domain A, Domain B, Hinge region and Domain C, or (iii) Domain A, Domain B and Hinge region, a third polypeptide comprising these domains. The third polypeptide further comprises the domains A, B, hinge region, C, and D of the third polypeptide arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus, and d) A fourth polypeptide including domains E and F The fourth polypeptide further comprises the domains E and F of the fourth polypeptide being aligned in an E-F orientation from the N-terminus to the C-terminus. The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds. The fusion protein construct according to claim 72 is a tetravalent heterodimer type fusion protein construct. construction.

79. The fusion protein construct according to claim 78, wherein the domain E and the domain R are conjugated in the first polypeptide.

80. The fusion protein construct according to claim 78, wherein the domain F and the domain R are conjugated in the first polypeptide.

81. A fusion protein construct according to claim 79 or 80, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

82. The fusion protein construct according to claim 81, wherein the domain R1 is conjugated to the first polypeptide.

83. The fusion protein construct according to claim 81, wherein the domain R1 is conjugated to the second polypeptide.

84. The fusion protein construct according to claim 81, wherein the domain R1 is conjugated to the third polypeptide.

85. The fusion protein construct according to claim 81, wherein the domain R1 is conjugated to the fourth polypeptide.

86. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide and a second polypeptide, a) The first polypeptide comprises domain A, domain B and a hinge region, The domains A, B and the hinge region are arranged in an A-B-hinge region orientation from the N-terminus to the C-terminus. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, b) The second polypeptide comprises domain E, domain F and domain R, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (1) Domain E and Domain R are combined, or (2) Domain F and Domain R are combined, (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

87. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain E and domain R are conjugated in at least one of the second polypeptides, and the two first polypeptides are linked together in the hinge region via one or more disulfide bonds, as described in claim 86. The fusion protein construct shown.

88. A tetravalent homodimeric fusion protein construct comprising two of the first polypeptides and two of the second polypeptides, wherein domain F and domain R are conjugated in at least one of the second polypeptides, and the two first polypeptides are linked together via one or more disulfide bonds in the hinge region, according to claim 86.

89. A fusion protein construct according to claim 87 or claim 88, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

90. The fusion protein construct according to claim 89, wherein the domain E and the domain R are conjugated, and the domain F and the domain R1 are conjugated.

91. The fusion protein according to claim 89, wherein the domain F and the domain R are conjugated, and the domain E and the domain R1 are conjugated.

92. c) (i) Domain A, Domain B, Hinge region, Domain C and Domain D, (ii) Domain A, Domain B, Hinge region and Domain C, or (iii) Domain A, Domain B and Hinge region, a third polypeptide comprising these domains. The third polypeptide further comprises the domains A, B, hinge region, C, and D of the third polypeptide arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus, and d) A fourth polypeptide including domains E and F The fourth polypeptide further comprises the domains E and F of the fourth polypeptide being aligned in an E-F orientation from the N-terminus to the C-terminus. The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds. The fusion protein construct according to claim 86, which is a tetravalent heterodimer type fusion protein construct.

93. The fusion protein construct according to claim 92, wherein the domain E and the domain R are conjugated in the first polypeptide.

94. The fusion protein construct according to claim 92, wherein the domain F and the domain R are conjugated in the first polypeptide.

95. A fusion protein construct according to claim 93 or claim 94, further comprising domain R1, wherein domain R1 comprises a second complement regulatory polypeptide, and domain R1 and domain R are the same or different.

96. The fusion protein construct according to claim 95, wherein the domain R1 is conjugated to the first polypeptide.

97. The fusion protein construct according to claim 95, wherein the domain R1 is conjugated to the second polypeptide.

98. The domain R1 is conjugated to the third polypeptide, as described in claim 95. Fusion protein construct.

99. The fusion protein construct according to claim 95, wherein the domain R1 is conjugated to the fourth polypeptide.

100. A trivalent heterodimeric fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region, domain C and domain D, The domains A, B, hinge region, C, and D of the first polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. b) The second polypeptide comprises domain E, domain F and domain R, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (1) Domain E and Domain R are combined, or (2) Domain F and Domain R are combined, c) The third polypeptide comprises (i) domain A, domain B, domain C, domain D and hinge region, (ii) domain A, domain B, domain C and hinge region, or (iii) domain A, domain B and hinge region. The domains A, B, hinge region, C, and D of the third polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. d) The fourth polypeptide comprises domain E and domain F, The domains E and F of the fourth polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. The domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds, The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain D contains a heavy chain CH3 constant region amino acid sequence, (v) The domain R contains a complement regulator polypeptide, (vi) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (vii) The domain F contains the light chain constant region amino acid sequence (CL1), The aforementioned trivalent heterodimer fusion protein construct.

101. The trivalent heterodimer fusion protein construct according to claim 100, wherein the domain E and the domain R of the second polypeptide are conjugated.

102. The trivalent heterodimer fusion protein construct according to claim 100, wherein the domain F and the domain R of the second polypeptide are conjugated.

103. A trivalent heterodimeric fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, a) The first polypeptide comprises domain A, domain B, hinge region and domain C, The domains A, B, hinge region, and C of the first polypeptide are arranged in the orientation A-B-hinge region-C from the N-terminus to the C-terminus. b) The second polypeptide comprises domain E, domain F and domain R, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (1) Domain E and Domain R are combined, or (2) Domain F and Domain R are combined, c) The third polypeptide comprises (i) domain A, domain B, domain C, domain D and hinge region, (ii) domain A, domain B, domain C and hinge region, or (iii) domain A, domain B and hinge region. The domains A, B, hinge region, C, and D of the third polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. d) The fourth polypeptide comprises domain E and domain F, The domains E and F of the fourth polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. The domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds, The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain R contains a complement regulator polypeptide, (v) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (vi) The trivalent heterodimer fusion protein construct wherein domain F contains the light chain constant region amino acid sequence (CL1).

104. The trivalent heterodimer fusion protein construct according to claim 103, wherein the domain E and the domain R of the second polypeptide are conjugated.

105. The trivalent heterodimer fusion protein construct according to claim 103, wherein the domain F and the domain R of the second polypeptide are conjugated.

106. A trivalent heterodimeric fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, a third polypeptide, and a fourth polypeptide, a) The first polypeptide comprises domain A, domain B and a hinge region, The domains A, B and hinge region of the first polypeptide are arranged in an A-B-hinge region orientation from the N-terminus to the C-terminus. b) The second polypeptide comprises domain E, domain F and domain R, The domains E and F are arranged in an E-F orientation from the N-terminus toward the C-terminus. (1) Domain E and Domain R are combined, or (2) Domain F and Domain R are combined, c) The third polypeptide comprises (i) domain A, domain B, domain C, domain D and hinge region, (ii) domain A, domain B, domain C and hinge region, or (iii) domain A, domain B and hinge region. The domains A, B, hinge region, C, and D of the third polypeptide are arranged in the orientation A-B-hinge region-C-D from the N-terminus to the C-terminus. d) The fourth polypeptide comprises domain E and domain F, The domains E and F of the fourth polypeptide are arranged in an E-F orientation from the N-terminus to the C-terminus. The domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds, The domain B of the third polypeptide and the domain F of the fourth polypeptide are linked by one or more disulfide bonds, The first and third polypeptides are linked together in the hinge region via one or more disulfide bonds, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (v) The domain F contains the light chain constant region amino acid sequence (CL1), The aforementioned trivalent heterodimer fusion protein construct.

107. The trivalent heterodimer fusion protein construct according to claim 106, wherein the domain E and the domain R are conjugated in the second polypeptide.

108. The trivalent heterodimer fusion protein construct according to claim 106, wherein the domain F and the domain R are conjugated in the second polypeptide.

109. A fusion protein construct that binds to complement-related antigens, comprising a first polypeptide, a second polypeptide, and a third polypeptide, a) The first polypeptide comprises domain R, hinge region, domain C and domain D, The domains R, hinge region, C, and D are arranged in the R-hinge-C-D orientation from the N-terminus to the C-terminus. (i) The domain R contains a complement modulator polypeptide, (ii) The domain C contains a heavy chain CH2 constant region amino acid sequence, (iii) The domain D contains a heavy chain CH3 constant region amino acid sequence, and the domain R and the hinge region are conjugated, b) The second polypeptide comprises domain A, domain B, hinge region, domain C and domain D, The domains A, B, hinge region, C, and D are arranged in the orientation A-B-hinge-C-D from the N-terminus to the C-terminus. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, c) The third polypeptide comprises domain E and domain F, The aforementioned domains are arranged in an E-F orientation from the N-terminus to the C-terminus. (i) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (ii) The domain F contains the light chain constant region amino acid sequence (CL1), The domain B of the second polypeptide and the domain F of the third polypeptide are linked by one or more disulfide bonds, The fusion protein construct wherein the first and second polypeptides are linked together in the hinge region via one or more disulfide bonds.

110. A monomeric fusion protein construct that binds to complement-related antigens, a) A first polypeptide comprising domain A, domain B, and a hinge region arranged in an A-B-hinge orientation from the N-terminus to the C-terminus, b) A second polypeptide comprising domains E and F aligned in an E-F orientation from the N-terminus to the C-terminus. The domain includes, wherein at least one of the domains A, the hinge domain, the domain E, or the domain F is combined with the domain R. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (v) The domain F contains the light chain constant region amino acid sequence (CL1), The monomeric fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

111. The monomeric fusion protein construct according to claim 110, wherein the first polypeptide comprises domain A, domain B, hinge domain and domain R, the domains of the first polypeptide are arranged in an R-A-B-hinge orientation from the N-terminus to the C-terminus, and domain R and domain A are conjugated.

112. The monomeric fusion protein construct according to claim 110, wherein the first polypeptide comprises domain A, domain B, hinge domain and domain R, the domains of the first polypeptide are arranged in an A-B-hinge-R orientation from the N-terminus to the C-terminus, and the hinge domain and domain R are conjugated.

113. The monomeric fusion protein construct according to claim 110, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an R-E-F orientation from the N-terminus to the C-terminus, and domain E and domain R are conjugated.

114. The monomeric fusion protein construct according to claim 110, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an E-F-R orientation from the N-terminus to the C-terminus, and domain F and domain R are conjugated.

115. A monomeric fusion protein construct according to claim 111, 112, 113, or 114, further comprising a second complement regulatory polypeptide, wherein the second complement regulatory polypeptide and the domain R are the same or different.

116. A monomeric fusion protein construct that binds to complement-related antigens, a) Domains A arranged in an A-B-hinge-C region orientation from the N-terminus to the C-terminus, A first polypeptide comprising domain B, hinge region and domain C, b) A second polypeptide comprising domains E and F aligned in an E-F orientation from the N-terminus to the C-terminus. The domains include, and at least one of the domains A, C, E, or F is conjugated to domain R. (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain R contains a complement regulator polypeptide, (v) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (vi) The monomeric fusion protein construct wherein domain F contains a light chain constant region amino acid sequence (CL1), and domain B of the first polypeptide and domain F of the second polypeptide are linked by one or more disulfide bonds.

117. The monomeric fusion protein construct according to claim 116, wherein the first polypeptide comprises domain A, domain B, hinge domain, domain C, and domain R, the domains of the first polypeptide are arranged in an R-A-B-hinge-C orientation from the N-terminus to the C-terminus, and domain R and domain A are conjugated.

118. The monomeric fusion protein construct according to claim 116, wherein the first polypeptide comprises domain A, domain B, hinge domain, domain C, and domain R, the domains of the first polypeptide are arranged in an A-B-hinge-C-R orientation from the N-terminus to the C-terminus, and domain C and domain R are conjugated.

119. The monomeric fusion protein construct according to claim 116, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an R-E-F orientation from the N-terminus to the C-terminus, and domain E and domain R are conjugated.

120. The monomeric fusion protein construct according to claim 116, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an E-F-R orientation from the N-terminus to the C-terminus, and domain F and domain R are conjugated.

121. A monomeric fusion protein construct according to claim 117, 118, 119, or 120, further comprising a second complement regulatory polypeptide, wherein the second complement regulatory polypeptide and the domain R are the same or different.

122. A monomeric fusion protein construct that binds to complement-related antigens, a) A first polypeptide comprising domains A, B, a hinge region, domain C, and D arranged in an A-B-hinge-C-D region orientation from the N-terminus to the C-terminus, b) A second polypeptide comprising domains E and F aligned in an E-F orientation from the N-terminus to the C-terminus. The domains include, and at least one of the domains A, D, E, or F is combined with the domain R. (i) Domain A is a heavy chain variable region amino acid sequence (VH) or an antigen-binding cleavage thereof It contains pieces, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain C contains the heavy chain CH2 constant region amino acid sequence, (iv) The domain D contains a heavy chain CH3 constant region amino acid sequence, (v) The domain R contains a complement regulator polypeptide, (vi) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (vii) The domain F contains the light chain constant region amino acid sequence (CL1), The monomeric fusion protein construct wherein the domain B of the first polypeptide and the domain F of the second polypeptide are linked by one or more disulfide bonds.

123. The monomeric fusion protein construct according to claim 116, wherein the first polypeptide comprises domain A, domain B, hinge domain, domain C, domain D, and domain R, the domains of the first polypeptide are arranged in the orientation R-A-B-hinge-C-D from the N-terminus to the C-terminus, and domain R and domain A are conjugated.

124. The monomeric fusion protein construct according to claim 116, wherein the first polypeptide comprises domain A, domain B, hinge domain, domain C, domain D, and domain R, the domains of the first polypeptide are arranged in an A-B-hinge-C-D-R orientation from the N-terminus to the C-terminus, and domain C and domain R are conjugated.

125. The monomeric fusion protein construct according to claim 116, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an R-E-F orientation from the N-terminus to the C-terminus, and domain E and domain R are conjugated.

126. The monomeric fusion protein construct according to claim 116, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an E-F-R orientation from the N-terminus to the C-terminus, and domain F and domain R are conjugated.

127. A monomeric fusion protein construct according to claim 117, 118, 119, or 120, further comprising a second complement regulatory polypeptide, wherein the second complement regulatory polypeptide and the domain R are the same or different.

128. The aforementioned combination is Linking two domains with a peptide linker, linking them without a linker, enzymatic conjugation, chemical conjugation, or a combination thereof. A fusion protein construct according to any one of claims 1 to 120, comprising:

129. The fusion protein construct according to any one of claims 1 to 128, wherein the complement-related antigen is C3d, iC3b, C3dg, a fragment thereof, or a variant thereof.

130. The fusion protein construct according to claim 129, wherein it binds to C3 and C3b with lower affinity than to C3d.

131. Approximately 10 －３ K above M Ｄ The fusion according to claim 130, which is bonded by affinity. Synthetic protein construct.

132. iC3b, C3dg, or both: 10 －８ K below M Ｄ A fusion protein construct according to claim 130, which binds by affinity.

133. A fusion protein construct according to any one of claims 1 to 132, which modulates surrogate complement activity in a subject when administered to the subject, the fusion protein construct or a pharmaceutical composition containing the fusion protein construct.

134. A fusion protein construct according to any one of claims 1 to 132, which modulates classical complement activity in a subject when administered to the subject, the fusion protein construct or a pharmaceutical composition containing the fusion protein construct.

135. A fusion protein construct according to any one of claims 1 to 132, which modulates lectin complement activity in a subject when administered to the subject, the fusion protein construct or a pharmaceutical composition containing the fusion protein construct.

136. The fusion protein construct according to any one of claims 1 to 135, wherein the fusion protein construct binds to the domain of a mammalian annexin protein.

137. The aforementioned fusion protein construct has 10 in the domain of mammalian annexin protein. －８ K below M Ｄ A fusion protein construct according to claim 136, which binds by affinity.

138. The fusion protein according to claim 136, wherein the domain is an annexin core domain.

139. The fusion protein construct according to claim 138, wherein the annexin core domain comprises an alpha-helical domain.

140. The fusion protein construct according to claim 139, wherein the annexin core domain includes a calcium-binding site and a membrane-binding site.

141. The fusion protein construct according to claim 139 or claim 140, wherein the annexin core domain comprises at least one annexin repeat.

142. The fusion protein construct according to claim 141, wherein the fusion protein construct binds to the annexin repeat sequence in the domain.

143. The fusion protein construct according to any one of claims 1 to 135, wherein the fusion protein construct binds to a phospholipid.

144. The aforementioned fusion protein construct is 10 phospholipids －８ K below M Ｄ A fusion protein construct according to claim 143, which binds by affinity.

145. The fusion protein construct according to claim 143, wherein the phospholipid is selected from the group consisting of phosphatidylethanolamine (PE), cardiolipin (CL), phosphatidylcholine (PC), phosphatidylinositol, phosphatidylglycerol, phosphatidylserine, phosphatidic acid, and malondialdehyde (MDA).

146. The complement regulatory peptide is the domain A of the complement receptor 1 (CR1) protein, or A fusion protein construct according to any one of claims 1 to 145, comprising a fragment thereof that retains at least three short consensus repeats (SCRs) of domain A.

147. The fusion protein construct according to claim 146, further comprising domain B of the CR1 protein, or a fragment thereof holding at least three SCRs of domain B.

148. The fusion protein construct according to claim 146 or claim 147, further comprising domain C of the CR1 protein, or a fragment thereof holding at least three SCRs of domain C.

149. The fusion protein construct according to claim 146, 147, or 148, further comprising domain D of CR1, or a fragment thereof holding at least three SCRs of domain D.

150. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulator polypeptide comprises the first three SCRs of domain A, the first three SCRs of domain B, and the first three SCRs of domain C of the CR1 protein.

151. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulator polypeptide is CR1-10.

152. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulator polypeptide is CR1-17.

153. The complement regulatory peptide is The amino acid sequence of SEQ ID NO: 41 or SEQ ID NO: 91, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 151, comprising:

154. The complement-modulating polypeptide is The amino acid sequence of SEQ ID NO: 42 or SEQ ID NO: 92, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 152, comprising:

155. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulatory peptide is a dysintegration factor (DAF) or a bioactive fragment thereof.

156. The fusion protein construct according to claim 153, wherein the DAF is human DAF.

157. The fusion protein construct according to claim 155 or claim 156, wherein the bioactive fragment of human DAF comprises at least one of a short consensus repeat (SCR) domain and an O-glycosylated serine / threonine-rich domain of full-length human DAF.

158. The fusion protein construct according to claim 155 or claim 156, wherein the physiologically active fragment of DAF comprises SCR1-4 or SCR2-4 of full-length human DAF.

159. The physiologically active fragment of DAF is The amino acid sequence of sequence number 184, or a variant thereof having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 155, comprising:

160. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulatory peptide is factor H or a bioactive fragment thereof.

161. The fusion protein construct according to claim 160, wherein the H factor is human H factor.

162. The physiologically active fragment of the human factor H is Full-length human H factor SCR1-20, SCR1-2, SCR2-3, SCR3-4, SCR4-5, SCR5-6, SCR6-7, SCR7-8, SCR8-9, SCR9-10, SCR10-11, SCR11-12, SCR12-13, SCR13-14, SCR14-15, SCR15-16, SCR16-17, SCR17-18, SCR19-20, or any combination of SCR1-20. A fusion protein construct according to claim 161, comprising one or more groups of short consensus repeats (SCRs) including the following.

163. The fusion protein construct according to claim 161, wherein the physiologically active fragment of the human H factor comprises SCR1-4 or SCR1-5 of the full-length human H factor.

164. The physiologically active fragment of the human factor H is A sequence of amino acids selected from the group consisting of amino acids 21-266, 21-320, 21-509, or 19-1106 of SEQ ID NO: 9, or The fusion protein construct according to claim 161, comprising a variant having an amino acid sequence that is at least 85% identical to the aforementioned sequence of amino acids.

165. The aforementioned H factor or its physiologically active fragment The amino acid sequence of SEQ ID NO: 72 or SEQ ID NO: 108, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 160, comprising:

166. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulatory peptide is MCP or a physiologically active fragment thereof.

167. The fusion protein construct according to claim 166, wherein the MCP is human MCP.

168. The fusion protein construct according to claim 167, wherein the bioactive fragment of the human MCP comprises at least one short consensus repeat (SCR) domain of a full-length human MCP.

169. The fusion protein construct according to claim 168, wherein the physiologically active fragment of the human MCP comprises SCR3-4 of the full-length human MCP.

170. The aforementioned MCP, The amino acid sequence of SEQ ID NO: 187, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 166, comprising:

171. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulatory peptide is Map44 or a physiologically active fragment thereof.

172. The fusion protein construct according to claim 171, wherein the Map44 is human Map44. 。

173. The aforementioned Map 44 is, The amino acid sequence of SEQ ID NO: 186, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 171, comprising:

174. The fusion protein construct according to any one of claims 1 to 145, wherein the complement regulatory peptide is CD59 or a bioactive fragment thereof.

175. The fusion protein construct according to claim 174, wherein the CD59 is human CD59.

176. The aforementioned CD59 is The amino acid sequence of SEQ ID NO: 185, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 174, comprising:

177. The fusion protein construct according to any one of claims 1 to 176, wherein the fusion protein construct comprises a human antibody or an antigen-binding fragment thereof.

178. The fusion protein construct according to any one of claims 1 to 177, wherein the fusion protein construct comprises a humanized antibody or an antigen-binding fragment thereof.

179. A fusion protein construct according to any one of claims 49 to 57 or 100 to 108, wherein each of the first and third polypeptides comprises at least one orthogonal modification that is advantageous for heterodimer formation compared to homodimer formation.

180. A trivalent fusion protein construct according to claim 179, wherein the first polypeptide comprises a knob modification and the third polypeptide comprises a hole modification, or the third polypeptide comprises a knob modification and the first polypeptide comprises a hole modification.

181. The trivalent fusion protein construct according to claim 180, wherein the first and third polypeptides include modifications that result in charge or surface complementarity.

182. a) The first polypeptide is (i) Three heavy chain complementarity determining regions (CDRs) having the amino acid sequences of SEQ ID NOs: 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 259 and 31; or 29, 260 and 31, or (ii) Three heavy chain CDRs having amino acid sequences that differ in that one of the following sequences contains a single conserved amino acid substitution: (ii) Sequence IDs 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 259 and 31; or 29, 260 and 31. Includes, b) The second polypeptide is (i) Sequence IDs 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 3 Three light chain complementarity-determining regions (CDRs) having amino acid sequences 2, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233, or (ii) Three light chain CDRs having amino acid sequences that differ in that one of the following sequences contains a single conserved amino acid substitution: (ii) Sequence IDs 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 32, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233. A fusion protein construct according to any one of claims 179 to 181, comprising:

183. A fusion protein construct according to any one of claims 1 to 182, further comprising at least one amino acid linker, wherein the at least one linker comprises any of the amino acid sequences of SEQ ID NO: 138, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169, SEQ ID NO: 170, SEQ ID NO: 171, SEQ ID NO: 172, SEQ ID NO: 173, SEQ ID NO: 174, SEQ ID NO: 175, SEQ ID NO: 176, SEQ ID NO: 177, SEQ ID NO: 178, SEQ ID NO: 179, SEQ ID NO: 180, SEQ ID NO: 181, SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 241, or SEQ ID NO:

242.

184. The aforementioned fusion protein construct is Sequence ID 43, Sequence ID 44, Sequence ID 46, Sequence ID 47, Sequence ID 48, Sequence ID 49, Sequence ID 50, Sequence ID 52, Sequence ID 53, Sequence ID 55, Sequence ID 56, Sequence ID 57, Sequence ID 60, Sequence ID 61, Sequence ID 63, Sequence ID 65, Sequence ID 66, Sequence ID 69, Sequence ID 70, Sequence ID 71, Sequence ID 75, Sequence ID 76, Sequence ID 77, Sequence ID 78, Sequence ID 79, Sequence ID 80, Sequence ID 82, Sequence ID 83, Sequence ID 84, Sequence ID 85, Sequence ID 86, Sequence ID 87, Sequence ID 88, Sequence ID 90, Sequence ID 93, Sequence ID 94, Sequence ID 95, Sequence ID 96, Sequence ID 97, Sequence ID 98, Sequence ID 99, Sequence ID 100, Sequence ID 101, Sequence ID 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143 and SEQ ID NO: 144 A fusion protein construct according to any one of claims 1 to 183, comprising at least one amino acid sequence from among the following.

185. A fusion protein construct according to any one of claims 1 to 181, wherein the first polypeptide comprises the sequence of SEQ ID NO: 282 (domains A, B, C, and D) conjugated to a complement regulator polypeptide containing a sequence (domain R) selected from the group consisting of SEQ ID NOs: 41, 42, and 72, and the second polypeptide comprises the sequence of SEQ ID NO: 279 (domains E and F).

186. The first polypeptide comprises the sequence of SEQ ID NO: 282 (domains A, B, C, and D), and the second polypeptide is conjugated to a complement regulator polypeptide comprising a sequence selected from the group consisting of SEQ ID NO: 72 (domain R), resulting in SEQ ID NO:

279. A fusion protein construct according to any one of claims 1 to 181, comprising the sequences (domain E and domain F).

187. A pharmaceutical composition comprising a fusion protein construct according to any one of claims 1 to 186.

188. A polynucleotide encoding a fusion protein according to any one of claims 1 to 186.

189. A therapeutic method comprising providing a therapeutically effective amount of the pharmaceutical composition described in claim 186 to a target.

190. The method according to claim 189, wherein the subject is suffering from a complement-mediated disease or complement-mediated inflammation.

191. The fusion protein construct binds specifically to C3d with a K affinity of 10 M or less, the subject has a complement-mediated disease, and the complement-mediated disease is characterized by increased deposition of C3d. The method according to claim 189 or claim 190. －８ M or less Ｄ 親和性で結合し、前記対象が補体媒介性疾患に罹患しており、前記補体媒介性疾患がC3dの増加した沈着を特徴とする、請求項189または請求項190に記載の方法。

192. The aforementioned fusion protein construct specifically reacts to C2 antibody-reactive phospholipids. －８ K below M Ｄ The method according to claim 189 or claim 190, wherein the substance binds by affinity, the target suffers from a complement-mediated disease, and the complement-mediated disease is characterized by increased deposition of C2 antibody-reactive phospholipids.

193. The method according to claim 190, wherein the subject is suffering from complement-mediated inflammation, the complement-mediated inflammation includes inflammatory fibrous disease, and the inflammatory fibrous disease includes focal segmental glomerulosclerosis, primary sclerosing cholangitis, or membranoproliferative glomerulonephritis.

194. The method according to claim 190, wherein the subject suffers from a complement-mediated autoimmune disease including rheumatoid arthritis, systemic lupus erythematosus, lupus nephritis, or pemphigus vulgaris.

195. The method according to claim 190, wherein the subject is suffering from a complement-mediated renal disease, including membranoproliferative glomerulonephritis or complement triglomerulopathy.

196. The method according to claim 190, wherein the subject is suffering from complement-mediated cardiovascular disease.

197. The method according to claim 196, wherein the cardiovascular disease includes atherosclerosis or thrombosis.

198. The method according to claim 190, wherein the subject is suffering from a complement-mediated skin disease.

199. The method according to claim 198, wherein the skin disease includes psoriasis, acne atypical, lupus erythematous, cutaneous microvasculitis, urticaria, urticarial vasculitis, and bullous pemphigoid.

200. The subject suffers from complement-mediated inflammation, and the complement-mediated inflammation is ischemia / reperfusion injury, burns, endotoxemia and septic shock, adult respiratory distress syndrome, cardiopulmonary bypass, hemodialysis, anaphylactic shock, asthma, angioedema, Crohn's disease, sickle cell anemia, glomerulonephritis, membranous nephritis, pancreatitis, graft rejection, hyperacute xenograft rejection, recurrent miscarriage, preeclampsia, drug allergy, IL-2-induced vasoleap syndrome, X-ray contrast agent allergy, myasthenia gravis, Alzheimer's disease, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, insulin-dependent diabetes mellitus, acute disseminated encephalomyelitis, Addison's disease, antiphospholipid antibody syndrome, etc. The method according to claim 190, relating to a symptom or disease selected from the group consisting of autoimmune hepatitis, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, Hashimoto's disease, idiopathic thrombocytopenic purpura, pemphigus, Sjögren's syndrome, Takayasu's arteritis, myocardial infarction, stroke, acute respiratory distress syndrome, sepsis, plasmapheresis, platelet ferresis, leukocytapheresis, extracorporeal membrane oxygenation, heparinized extracorporeal LDL precipitation, enteritis, urticarial and vasculitis, and lupus nephritis.

201. The aforementioned conditions include ischemia-reperfusion injury, rheumatoid arthritis (RA), lupus nephritis, atypical hemolytic uremic syndrome (aHUS), typical or infectious hemolytic uremic syndrome (tHUS), dense deposit disease (DDD), paroxysmal nocturnal hemoglobinuria (PNH), multiple sclerosis (MS), macular degeneration, hemolysis, elevated liver enzymes, and hypothrombocytopenia (HELLP) syndrome, sepsis, dermatomyositis, Diabetic retinopathy, thrombotic thrombocytopenic purpura (TTP), spontaneous miscarriage, microvascular vasculitis, epidermolysis bullosa, recurrent miscarriage, multiple sclerosis (MS), traumatic brain injury, cardiovascular disorders, myocarditis, cerebrovascular disorders, peripheral vascular disorders, renal vascular disorders, mesenteric / intestinal vascular disorders, vascular regeneration for grafts and / or replants, vasculitis, Henoch-Schönlein purpura nephritis, systemic lupus erythematosus-associated vasculitis, The method according to claim 189, wherein the patient is suffering from a condition or disease selected from the group consisting of vasculitis associated with rheumatoid arthritis, immune complex vasculitis, Takayasu's arthritis, capillary leak syndrome, dilated cardiomyopathy, diabetic vascular disease, thoracoabdominal aortic aneurysm, Kawasaki disease (arthritis), venous gas embolism (VGE), and restenosis after stent placement, rotational atherosclerosis resection, percutaneous transluminal coronary angioplasty (PTCA), myasthenia gravis, cold agglutinin disease (CAD), paroxysmal cold hemoglobinuria (PCH), dermatomyositis, scleroderma, warm autoimmune hemolytic anemia, Graves' disease, Hashimoto's thyroiditis, type 1 diabetes mellitus, psoriasis, pemphigus, autoimmune hemolytic anemia (AIHA), idiopathic thrombocytopenic purpura (ITP), Goodpasture syndrome, antiphospholipid syndrome (APS), Degos disease, and fulminant APS (CAPS).

202. The method according to claim 189, wherein the subject suffers from a symptom or disease selected from the group consisting of age-related macular degeneration (AMD), membranoproliferative glomerulonephritis type II (MPGN II), hemolytic uremic syndrome (HUS), asthma, amyloidosis, and thrombotic thrombocytopenic purpura.

203. The method according to claim 202, wherein the hemolytic uremic syndrome (HUS) is atypical hemolytic uremic syndrome (aHUS).

204. The method according to claim 189, wherein the subject is suffering from a drusen-related disease or a disease associated with drusen.

205. The method according to claim 204, wherein the disease related to the drusen is amyloidosis, elastic fibrosis, dense deposit disease, glomerulonephritis, atherosclerosis, or an eye disease related to the drusen.

206. A pharmaceutical composition comprising a tetravalent fusion protein construct for regulating complement activity, wherein the tetravalent fusion protein construct is a) an antibody or its antigen-binding fragment that binds to a complement-related antigen, and b) Complement regulator peptide 1 and complement regulator peptide 2 The pharmaceutical composition comprising the first and second complement modulating polypeptides, wherein the first and second complement modulating polypeptides are the same or different.

207. The pharmaceutical composition according to claim 206, wherein at least one of the first and second complement-modulating peptides is conjugated to the antibody or antigen-binding fragment by a linker.

208. A pharmaceutical composition comprising a trivalent fusion protein construct for regulating complement activity, wherein the tetravalent fusion protein construct is a) an antibody or its antigen-binding fragment that binds to a complement-related antigen, and b) A single complement-modulating peptide The pharmaceutical composition comprising the above.

209. The pharmaceutical composition according to claim 208, wherein the antibody or its antigen-binding fragment and the complement-modulating peptide are conjugated by a linker.

210. A pharmaceutical composition comprising a trivalent fusion protein construct for regulating complement activity, wherein the trivalent fusion protein construct is a) Fab that binds to complement-related antigens, b) Antibody Fc domain, and c) Complement-modulating peptides conjugated to the Fab or Fc domain. The pharmaceutical composition comprising the above.

211. The pharmaceutical composition according to claim 210, wherein the Fab or Fc domain and the complement regulatory peptide are conjugated by a linker.

212. a) A first polypeptide monomer containing the CH2 or CH3 domain of the antibody. The domain includes, wherein the domain has at least one orthogonal modification that is more favorable to the formation of a heterodimer compared to a homodimer; b) Second polypeptide monomer containing the CH2 or CH3 domain of the antibody The domain comprises, wherein the domain has at least one orthogonal modification that is advantageous for the formation of a heterodimer with the first polypeptide monomer compared to a homodimer. Trivalent fusion protein construct A pharmaceutical composition comprising, wherein one of the first and second polypeptides further comprises a complement regulator peptide, and the trivalent fusion protein construct binds to a complement-related antigen.

213. The pharmaceutical composition according to claim 212, wherein the first polypeptide comprises a knob modification and the second polypeptide comprises a hole modification.

214. The pharmaceutical composition according to claim 212, wherein the first and second polypeptides include modifications that provide charge or surface complementarity.

215. The pharmaceutical composition according to any one of claims 212 to 214, wherein the complement-modulating peptide is linked to one of the first and second polypeptides by an amino acid linker.

216. a) Three heavy chain complementarity-determining regions (CDRs) having the amino acid sequences of SEQ ID NOs: 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 259 and 31; or 29, 260 and 31, or b) Sequence IDs 11, 12 and 13; 17, 18 and 19; 23, 24 and 25; 29, 30 and 31; 35, 36 and 37; 147, 148 and 149; 188, 189 and 190; 196, 197 and 198; 204 or 343, 205 and 206; 212, 213 and 214; 220, 221 and 222; 228, 229 and 230; 29, 25 Three heavy chain CDRs having amino acid sequences that differ in that one of 9 and 31 has a single conserved amino acid substitution in one of 29, 260, and 31; and c) Three light chain complementarity determining regions (CDRs) having the amino acid sequences of SEQ ID NOs: 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 32, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233, or d) Three light chain CDRs having amino acid sequences that differ in that one of the following sequences contains a single conserved amino acid substitution: SEQ ID NOs: 14, 15 and 16; 20, 21 and 22; 26, 27 and 28; 32, 33 and 34; 38, 39 and 40; 150, 151 and 152; 191, 192 and 193; 199, 200 and 201; 207, 208 and 209; 215, 216 and 217; 223, 224 and 225; or 231, 232 and 233. Antibodies containing or antigen-binding fragments thereof; and Complement-modulating peptides A pharmaceutical composition comprising a fusion protein construct containing [specific component].

217. a) Heavy chains comprising at least one amino acid sequence from sequence numbers 54, 58, 67, 73, 74, 81, 88, 89, 98, 99, 112, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290 and 342, and b) Light chain containing at least one amino acid sequence from sequence numbers 45, 59, 68, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, 278, 279, 287, and 289 including, or c) Heavy chains containing amino acid sequences that differ in that at least one of sequence numbers 54, 58, 67, 73, 74, 81, 88, 89, 98, 99, 112, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290 and 342 has one or more conserved amino acid substitutions, and d) Light chains containing amino acid sequences that differ in that at least one of sequence numbers 45, 59, 68, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, 278, 279, 287, and 289 has one or more conserved amino acid substitutions. A pharmaceutical composition comprising a fusion protein construct containing an antibody or an antigen-binding fragment thereof.

218. Sequence numbers 45, 51, 54, 58, 59, 62, 64, 67, 68, 73, 74, 79, 81, 88, 89, 98, 99, 112, 194, 195, 202, 203, 210, 211, 218, 219, 226, 227, 234, 235, 238, 239, 240, 243, 244, 246, 247, 248, 249, 250, 251, 252, 253, 25 Sequences selected from the group consisting of 4, 255, 256, 257, 258, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290 and 342, and Complement-modulating peptides A pharmaceutical composition comprising a fusion protein construct containing [specific component].

219. The pharmaceutical composition according to claim 216 or claim 217, wherein the complement-modulating peptide is linked to the antibody or its antigen-binding fragment by an amino acid linker.

220. The pharmaceutical composition according to claim 207, 209, 211, 215, or 218, wherein the amino acid linker comprises any of the amino acid sequences of SEQ ID NO: 138, SEQ ID NO: 161, SEQ ID NO: 162, SEQ ID NO: 163, SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169, SEQ ID NO: 170, SEQ ID NO: 171, SEQ ID NO: 172, SEQ ID NO: 173, SEQ ID NO: 174, SEQ ID NO: 175, SEQ ID NO: 176, SEQ ID NO: 177, SEQ ID NO: 178, SEQ ID NO: 179, SEQ ID NO: 180, SEQ ID NO: 181, SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 241, or SEQ ID NO:

242.

221. The antigen-binding fragment comprises the antigen-binding fragment Fv fragment, Fab, Fab', F(ab') ２ A pharmaceutical composition according to any one of claims 206 to 212 or 214 to 219, comprising scFv.

222. The pharmaceutical composition according to any one of claims 206 to 220, wherein when the pharmaceutical composition is administered to a subject, the fusion protein construct modulates alternative complement activity in the subject.

223. The pharmaceutical composition according to any one of claims 206 to 222, wherein the antibody or its antigen-binding fragment, or the Fab, is bound to the domain of a mammalian annexin protein.

224. The antibody or its antigen-binding fragment, or the Fab, is attached to the domain of the mammalian annexin protein. －８ K below M Ｄ A pharmaceutical composition according to any one of claims 206 to 223, which binds by affinity.

225. The pharmaceutical composition according to claim 223 or claim 224, wherein the domain is an annexin core domain.

226. The pharmaceutical composition according to claim 225, wherein the annexin core domain comprises an alpha-helical domain.

227. The pharmaceutical composition according to claim 224 or claim 225, wherein the annexin core domain comprises a calcium-binding site and a membrane-binding site.

228. The pharmaceutical composition according to any one of claims 223 to 227, wherein the annexin core domain comprises at least one annexin repeat sequence.

229. The pharmaceutical composition according to claim 228, wherein the antibody or its antigen-binding fragment, or the Fab, is bound to the at least one annexin repeat sequence.

230. The pharmaceutical composition according to any one of claims 206 to 221, wherein the complement-related antigen comprises a phospholipid, and the antibody or an antigen-binding fragment thereof, or the Fab, is bound to the phospholipid.

231. The antibody or its antigen-binding fragment, or the Fab, is 10 times the amount of phospholipid. －８ K below M Ｄ A pharmaceutical composition according to any one of claims 206 to 222 or 230, which binds by affinity.

232. The pharmaceutical composition according to claim 231, wherein the phospholipid is selected from the group consisting of phosphatidylethanolamine (PE), cardiolipin (CL), phosphatidylcholine (PC), and malondialdehyde (MDA).

233. The pharmaceutical composition according to any one of claims 206 to 222, wherein the complement-related antigen comprises a C3 complement protein or a fragment thereof, and the antibody or an antigen-binding fragment thereof, or the Fab, is bound to the C3 complement protein or a fragment thereof.

234. The antibody or its antigen-binding fragment, or the Fab, is attached to the C3 complement protein or its fragment. －８ K below M Ｄ A pharmaceutical composition according to any one of claims 206 to 222 or 233, which binds by affinity.

235. The pharmaceutical composition according to claim 234, wherein the C3 complement protein fragment is C3d.

236. The pharmaceutical composition according to any one of claims 206 to 235, wherein the complement regulatory peptide comprises complement receptor 1 (CR1) protein.

237. The pharmaceutical composition according to claim 236, wherein the complement-modulating peptide comprises domain A of the CR1 protein, or a fragment thereof that holds at least three short consensus repeats (SCRs) of domain A.

238. The pharmaceutical composition according to claim 236 or claim 237, wherein the complement regulator peptide comprises domain B of the CR1 protein, or a fragment thereof that holds at least three SCRs of domain B.

239. The pharmaceutical composition according to any one of claims 236 to 238, wherein the complement regulator peptide comprises domain C of the CR1 protein, or a fragment thereof that holds at least three SCRs of domain C.

240. The pharmaceutical composition according to any one of claims 236 to 239, further comprising domain D of the CR1 protein, or a fragment thereof holding at least three SCRs of domain D.

241. The pharmaceutical composition according to claim 240, wherein the complement regulator peptide comprises the first three SCRs of domain A, the first three SCRs of domain B, and the first three SCRs of domain C of the CR1 protein.

242. The pharmaceutical composition according to any one of claims 236 to 241, wherein the CR1 protein is human CR1 protein.

243. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement-modulating polypeptide is CR1(1-10).

244. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement-modulating polypeptide is CR1(1-17).

245. The aforementioned CR1 (1-10) The amino acid sequence of SEQ ID NO: 41 or SEQ ID NO: 91, or Its variant having an amino acid sequence that is at least 85% identical. A pharmaceutical composition according to claim 244, comprising:

246. The aforementioned CR1 (1-17) The amino acid sequence of SEQ ID NO: 42 or SEQ ID NO: 92, or Its variant having an amino acid sequence that is at least 85% identical. A pharmaceutical composition according to claim 244, comprising:

247. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement regulatory peptide is a complement-associated complement factor (DAF) or a physiologically active fragment thereof.

248. The pharmaceutical composition according to claim 247, wherein the DAF is human DAF.

249. The pharmaceutical composition according to claim 247 or claim 248, wherein the physiologically active fragment of human DAF comprises at least one of a short consensus repeat (SCR) domain and an O-glycosylated serine / threonine-rich domain of full-length human DAF.

250. The pharmaceutical composition according to claim 248 or claim 249, wherein the physiologically active fragment of human DAF comprises SCR1-4 or SCR2-4 of full-length human DAF.

251. The aforementioned physiologically active fragment of human DAF is The amino acid sequence of SEQ ID NO: 184, or Its variant having an amino acid sequence that is at least 85% identical. A pharmaceutical composition according to any one of claims 247 to 250, comprising:

252. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement regulatory peptide is factor H or a physiologically active fragment thereof.

253. The pharmaceutical composition according to claim 252, wherein the H factor is human H factor.

254. The physiologically active fragment of the human factor H is A sequence of amino acids selected from the group consisting of amino acids 21-266, 21-320, 21-509, or 19-1106 of SEQ ID NO: 9, or The pharmaceutical composition according to claim 253, comprising a variant thereof having an amino acid sequence that is at least 85% identical to the aforementioned sequence of amino acids.

255. The physiologically active fragment of the human factor H is Full-length human H factor SCR1-20, SCR1-2, SCR2-3, SCR3-4, SCR4-5, SCR5-6, SCR6-7, SCR7-8, SCR8-9, SCR9-10, SCR10-11, SCR11-12, SCR12-13, SCR13-14, SCR14-15, SCR15-16, SCR16-17, SCR17-18, SCR19-20, or any combination of SCR1-20. The pharmaceutical composition according to claim 253, comprising one or more groups of short consensus repeats (SCRs) including the above.

256. The pharmaceutical composition according to claim 253, wherein the physiologically active fragment of the human H factor comprises SCR1 to SCR4 of the full-length human H factor.

257. The pharmaceutical composition according to claim 253, wherein the physiologically active fragment of the human H factor comprises SCR1 to SCR5 of the full-length human H factor.

258. The physiologically active fragment of the human factor H is The amino acid sequence of SEQ ID NO: 72 or SEQ ID NO: 108, or Its variant having an amino acid sequence that is at least 85% identical. A pharmaceutical composition according to any one of claims 252 to 257, comprising:

259. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement regulatory peptide is MCP or a physiologically active fragment thereof.

260. The pharmaceutical composition according to claim 259, wherein the MCP is human MCP.

261. The pharmaceutical composition according to claim 260, wherein the physiologically active fragment of the human MCP comprises at least one short consensus repeat (SCR) domain of a full-length human MCP.

262. The pharmaceutical composition according to claim 261, wherein the physiologically active fragment of the human MCP comprises SCR3 to SCR4 of the full-length human MCP.

263. The physiologically active fragment of the human MCP is The amino acid sequence of SEQ ID NO: 187, or Its variant having an amino acid sequence that is at least 85% identical. A pharmaceutical composition according to claim 261 or claim 262, comprising:

264. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement regulatory peptide is Map44 or a physiologically active fragment thereof.

265. The pharmaceutical composition according to claim 264, wherein the Map 44 is human Map 44.

266. The aforementioned Map 44 is, The amino acid sequence of SEQ ID NO: 186, or Its variant having an amino acid sequence that is at least 85% identical. The pharmaceutical composition according to claim 265, comprising:

267. The pharmaceutical composition according to any one of claims 206 to 242, wherein the complement-modulating peptide is CD59 or a physiologically active fragment thereof.

268. The pharmaceutical composition according to claim 267, wherein the CD59 is human CD59.

269. The aforementioned CD59 is The amino acid sequence of SEQ ID NO: 185, or Its variant having an amino acid sequence that is at least 85% identical. A pharmaceutical composition according to claim 267 or claim 268, comprising:

270. The pharmaceutical composition according to any one of claims 206 to 269, wherein the antibody or its antigen-binding fragment is a human antibody or its antigen-binding fragment.

271. The pharmaceutical composition according to any one of claims 206 to 269, wherein the antibody or its antigen-binding fragment is a humanized antibody or its antigen-binding fragment.

272. A polynucleotide encoding a fusion protein as described in any one of claims 206 to 271.

273. A therapeutic method comprising providing a therapeutically effective amount of the pharmaceutical composition described in any one of claims 206 to 271.

274. The method according to claim 269, wherein the subject is suffering from complement-mediated inflammation.

275. The method according to claim 274, wherein the complement-mediated inflammation includes an inflammatory fibrous disease, and the inflammatory fibrous disease includes focal segmental glomerulosclerosis, primary sclerosing cholangitis, or membranoproliferative glomerulonephritis.

276. The method according to claim 275, wherein the subject suffers from a complement-mediated autoimmune disease including rheumatoid arthritis, systemic lupus erythematosus, lupus nephritis, or pemphigus vulgaris.

277. The method according to claim 275, wherein the subject is suffering from a complement-mediated renal disease, including membranoproliferative glomerulonephritis or complement triglomerulopathy.

278. The method according to claim 275, wherein the subject is suffering from complement-mediated cardiovascular disease.

279. The method according to claim 278, wherein the cardiovascular disease includes atherosclerosis or thrombosis.

280. The method according to claim 275, wherein the subject is suffering from a complement-mediated skin disease.

281. The method according to claim 280, wherein the skin disease includes psoriasis, acne atypical, lupus erythematous, cutaneous microvasculitis, urticaria, urticarial vasculitis, and bullous pemphigoid.

282. The aforementioned complement-mediated inflammation can lead to ischemia / reperfusion injury, burns, endotoxemia and septic shock, adult respiratory distress syndrome, cardiopulmonary bypass, hemodialysis, anaphylactic shock, asthma, angioedema, Crohn's disease, sickle cell anemia, glomerulonephritis, membranous nephritis, pancreatitis, graft rejection, hyperacute xenograft rejection, recurrent miscarriage, pre-eclampsia, drug allergies, IL-2-induced vasoleap syndrome, X-ray contrast agent allergies, myasthenia gravis, Alzheimer's disease, multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, insulin-dependent diabetes mellitus, acute The method according to claim 274, relating to a symptom or disease selected from the group consisting of disseminated encephalomyelitis, Addison's disease, antiphospholipid antibody syndrome, autoimmune hepatitis, Goodpasture syndrome, Graves' disease, Guillain-Barré syndrome, Hashimoto's disease, idiopathic thrombocytopenic purpura, pemphigus, Sjögren's syndrome, Takayasu's arteritis, myocardial infarction, stroke, acute respiratory distress syndrome, sepsis, plasmapheresis, platelet ferresis, leukocytapheresis, extracorporeal membrane oxygenation, heparinized extracorporeal LDL precipitation, enteritis, urticarial and vasculitis, and lupus nephritis.

283. The aforementioned conditions include ischemia-reperfusion injury, rheumatoid arthritis (RA), lupus nephritis, atypical hemolytic uremic syndrome (aHUS), typical or infectious hemolytic uremic syndrome (tHUS), dense deposit disease (DDD), paroxysmal nocturnal hemoglobinuria (PNH), multiple sclerosis (MS), macular degeneration, hemolysis, elevated liver enzymes, and hypothrombocytopenia (HELLP) syndrome, sepsis, dermatomyositis, diabetic retinopathy, thrombotic thrombocytopenic purpura (TTP), spontaneous miscarriage, microvasculitis, epidermolysis bullosa, recurrent miscarriage, traumatic brain injury, and cardiovascular disorders. Damage, myocarditis, cerebrovascular disease, peripheral vascular disease, renal vascular disease, mesenteric / intestinal vascular disease, vascular regeneration for grafts and / or replants, vasculitis, Henoch-Schönlein purpura nephritis, lupus erythematosus-associated vasculitis, vasculitis associated with rheumatoid arthritis, immune complex vasculitis, Takayasu's arteritis, capillary leak syndrome, dilated cardiomyopathy, diabetic vascular disease, thoracoabdominal aortic aneurysm, Kawasaki disease (arthritis), venous gas embolism (VGE), and restenosis after stent placement, rotational atherosclerosis resection, percutaneous transluminal coronary angioplasty (PTCA), myasthenia gravis, cold agglutinin disease (CAD), paroxysmal cold The method according to claim 273, wherein the patient suffers from a condition or disease selected from the group consisting of hemoglobinuria (PCH), dermatomyositis, scleroderma, warm autoimmune hemolytic anemia, Graves' disease, Hashimoto's thyroiditis, type 1 diabetes mellitus, psoriasis, pemphigus, autoimmune hemolytic anemia (AIHA), idiopathic thrombocytopenic purpura (ITP), Goodpasture syndrome, antiphospholipid syndrome (APS), Degos disease, and fulminant APS (CAPS).

284. The method according to claim 273, wherein the subject suffers from a symptom or disease selected from the group consisting of age-related macular degeneration (AMD), membranoproliferative glomerulonephritis type II (MPGN II), hemolytic uremic syndrome (HUS), asthma, amyloidosis, and thrombotic thrombocytopenic purpura.

285. The method according to claim 284, wherein the hemolytic uremic syndrome (HUS) is atypical hemolytic uremic syndrome (aHUS).

286. The method according to claim 283, wherein the subject is suffering from a drusen-related disease or a disease associated with drusen.

287. The method according to claim 286, wherein the disease related to the drusen is amyloidosis, elastic fibrosis, dense deposit disease, glomerulonephritis, atherosclerosis, or an eye disease related to the drusen.

288. A monomeric fusion protein construct that binds to complement-related antigens, a) A first polypeptide comprising domains A and B aligned in an A-B orientation from the N-terminus to the C-terminus, b) Second polypeptide containing domains E and F aligned in an E-F orientation from the N-terminus to the C-terminus. It includes, and at least one of domains A and E is conjugated to domain R, (i) Domain A comprises a heavy chain variable region amino acid sequence (VH) or an antigen-binding fragment thereof, (ii) The domain B contains the heavy chain CH1 constant region amino acid sequence, (iii) The domain R contains a complement regulator polypeptide, (iv) The domain E comprises a light chain variable region amino acid sequence (VL) or an antigen-binding fragment thereof, (v) The domain F contains the light chain constant region amino acid sequence (CL1), The monomeric fusion protein construct.

289. The monomeric fusion protein construct according to claim 288, wherein the first polypeptide comprises domain A, domain B, and domain R, the domains of the first polypeptide are arranged in an R-A-B orientation from the N-terminus to the C-terminus, and domain R and domain A are conjugated.

290. The monomeric fusion protein construct according to claim 288, wherein the second polypeptide comprises domain E, domain F, and domain R, the domains of the second polypeptide are arranged in an R-E-F orientation from the N-terminus to the C-terminus, and domain E and domain R are conjugated.

291. A fusion protein construct, a) An antibody or its antigen-binding fragment that specifically binds to complement protein 3d (c3d), and b) Two complement-modulating polypeptide molecules The complement regulator polypeptide comprises a physiologically active fragment of a complement protein selected from the group consisting of CR1, DAF, MCP, Crry, Map44, Map19, CD59, and factor H. The median inhibitory concentration in the complement assay is lower than that of the control protein construct, which is identical except for the absence of the aforementioned antibody. The aforementioned fusion protein construct.

292. The fusion protein construct according to claim 291, wherein each molecule of the complement regulator polypeptide is conjugated to the heavy chain of the antibody or its antigen-binding fragment.

293. The fusion protein construct according to claim 292, wherein each molecule of the complement regulator polypeptide is complexed with the C-terminus of the heavy chain.

294. The antibody or its antigen-binding fragment comprises a first polypeptide and a second polypeptide, a) The first polypeptide comprises a heavy chain sequence containing at least one amino acid sequence from among SEQ ID NOs: 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 74, 75, 76, 78, 80, 81, 82, 85, 87, 88, 132, 133, 134, 243, 282, 284, 285, and 286, b) The second polypeptide comprises a light chain sequence containing at least one amino acid sequence from among SEQ ID NOs: 59, SEQ ID NOs: 68, SEQ ID NOs: 77, SEQ ID NOs: 79, SEQ ID NOs: 84, SEQ ID NOs: 86, SEQ ID NOs: 135 and SEQ ID NOs: 279, A fusion protein construct according to claim 291.

295. The fusion protein construct according to claim 294, wherein the first polypeptide comprises at least two amino acid sequences selected from the group consisting of SEQ ID NOs: 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 74, 75, 76, 78, 80, 81, 82, 85, 87, 88, 132, 133, 134, 243, 282, 284, 285, and 286, and the second polypeptide comprises at least two amino acid sequences selected from the group consisting of SEQ ID NOs: 59, 68, 77, 79, 84, 86, 135, and 279.

296. The fusion protein construct according to claim 295, wherein at least two amino acid sequences of the first polypeptide are the same amino acid sequence, and at least two amino acid sequences of the second polypeptide are the same amino acid sequence.

297. The fusion protein construct according to claim 296, wherein the at least two amino acid sequences of the first polypeptide are sequence number 282 or sequence number 285, and the at least two amino acid sequences of the second polypeptide are sequence number 279.

298. The fusion protein construct according to claim 291, further comprising a linker.

299. The linker is sequence number 138, sequence number 161, sequence number 162, sequence number 163 A fusion protein construct according to claim 298, comprising any of the amino acid sequences of SEQ ID NO: 164, SEQ ID NO: 165, SEQ ID NO: 166, SEQ ID NO: 167, SEQ ID NO: 168, SEQ ID NO: 169, SEQ ID NO: 170, SEQ ID NO: 171, SEQ ID NO: 172, SEQ ID NO: 173, SEQ ID NO: 174, SEQ ID NO: 175, SEQ ID NO: 176, SEQ ID NO: 177, SEQ ID NO: 178, SEQ ID NO: 179, SEQ ID NO: 180, SEQ ID NO: 181, SEQ ID NO: 182, SEQ ID NO: 183, SEQ ID NO: 241, or SEQ ID NO:

242.

300. The fusion protein construct according to claim 291, wherein the complement protein is factor H or a physiologically active fragment thereof.

301. The aforementioned H factor or its physiologically active fragment The amino acid sequence of SEQ ID NO: 72 or SEQ ID NO: 108, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 300, comprising:

302. The fusion protein construct according to claim 291, wherein the complement protein is CR1 or a fragment thereof.

303. The aforementioned complement protein, The amino acid sequences of SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 91, or SEQ ID NO: 92, or Its variant having an amino acid sequence that is at least 85% identical. A fusion protein construct according to claim 302, comprising:

304. The aforementioned fusion protein construct is sequence number 41, 42, 43, 44, 46, 47, 48, 49, 50, 52, 53, 54, 55, 56, 57, 60, 61, 63, 65, 66, 69, 70, 71, 72, 75, 76, 77, 78, 79, 80, 82, 83, 84, 85, 86, 87, 88, 90, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 1 A fusion protein construct according to claim 291, comprising at least one amino acid sequence from among 04, SEQ ID NO: 105, SEQ ID NO: 106, SEQ ID NO: 107, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO: 122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 138, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 279, SEQ ID NO: 282, and SEQ ID NO:

285.

305. A fusion protein construct, a) Antibodies or antigen-binding fragments that specifically bind to complement-related antigens. The antibody comprises a first polypeptide and a second polypeptide, and each of the first polypeptide and the second polypeptide comprises a heavy chain and a light chain; further, b) The first and second molecules of the complement-modulating polypeptide The first molecule and the second molecule each contain a physiologically active fragment of a complement protein selected from the group consisting of CR1, DAF, MCP, Crry, Map44, Map19, CD59, and factor H. The fusion protein construct, wherein when administered to a subject with a disease, the albumin-to-creatinine ratio of a urine sample from the subject with the disease is lower than the albumin-to-creatinine ratio of a urine sample from a subject administered an equivalent fusion protein construct, and the equivalent fusion protein construct is identical to the equivalent fusion protein construct except that it does not contain an antibody or its antigen-binding fragment.

306. The fusion protein construct according to claim 305, wherein the disease is complement-mediated renal disease.

307. The fusion protein construct according to claim 306, wherein the complement-mediated renal disease is membranoproliferative glomerulonephritis or complement triglomerulopathy.

308. The fusion protein construct according to claim 305, wherein the albumin-to-creatinine ratio of the urine sample from the subject having the disease is at least about 1%, 2%, 3%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85% lower.

309. a) Sequence numbers 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 74, 75, 76, 78, 80, 81, 82, 85, 87, 88, 89, 98, 99, 112, 132, 133, 134, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 2 A heavy chain comprising at least one amino acid sequence from among 47, 248, 249, 250, 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290, and 342, and b) Light chain containing at least one amino acid sequence from sequence numbers 45, 59, 68, 77, 79, 84, 86, 135, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, 278, 279, 287, and 289 including, or c) Sequence numbers 54, 58, 60, 61, 64, 65, 67, 69, 70, 71, 73, 74, 75, 76, 78, 80, 81, 82, 85, 87, 88, 89, 98, 99, 112, 132, 133, 134, 145, 153, 194, 202, 210, 218, 226, 234, 238, 243, 244, 246, 247, 248, 249, 250, Heavy chains containing amino acid sequences that differ in that at least one of 251, 252, 253, 254, 255, 261, 262, 263, 264, 265, 270, 271, 272, 273, 274, 280, 281, 282, 283, 284, 285, 286, 288, 290, and 342 has one or more conserved amino acid substitutions, and d) Light chains containing amino acid sequences that differ in that at least one of sequence numbers 45, 59, 68, 77, 79, 84, 86, 135, 195, 203, 211, 219, 227, 235, 256, 257, 258, 266, 267, 268, 269, 275, 276, 277, 278, 279, 287, and 289 has one or more conserved amino acid substitutions. A fusion protein construct comprising an antibody or its antigen-binding fragment.

310. The fusion protein construct further comprises a complement regulator polypeptide, wherein the complement regulator polypeptide comprises at least one of the following: complement receptor 1 (CR1) protein, DAF, MCP, Crry, Map44, Map19, CD59, factor H, and its physiologically active fragment. The fusion protein construct according to claim 309.

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