FcRn Binding Molecule and Method of Use

Abstract

Binding molecules are provided herein that include a human neonatal Fc receptor (FcRn) binding molecule and a portion that does not specifically bind to a human antigen. Polynucleotides, vectors, host cells, and production methods are also provided herein. Further provided are methods of treating antibody-mediated disorders with the FcRn binding molecule or the FcRn / antigen binding molecule.

Description

Technical Field

[0001] The present disclosure relates to human neonatal Fc receptor (FcRn) binding molecules and methods of using them.

Background Art

[0002] Immunoglobulin gamma (IgG) antibodies play important roles in the pathology of many diseases, such as autoimmune diseases, inflammatory diseases, and disorders where the pathology is characterized by overexpression of IgG antibodies.

[0003] The half-life of IgG in serum is extended relative to the serum half-lives of other plasma proteins, due in part to the binding of the Fc region of IgG to the Fc receptor, FcRn. FcRn binds IgG and protects IgG from transport to degradative lysosomes by recycling IgG to the extracellular compartment. This recycling is facilitated by the pH-dependent binding of IgG to FcRn, and the IgG / FcRn interaction is stronger at acidic endosomal pH than at extracellular physiological pH.

[0004] When the serum concentration of IgG reaches levels that exceed the available FcRn molecules, unbound IgG is not protected from lysosomal degradation and thus has a reduced serum half-life. Thus, inhibition of IgG binding to FcRn reduces the serum half-life of IgG by preventing endosomal recycling of IgG. Agents that antagonize the binding of IgG to FcRn, such as FcRn binding molecules, are useful for modulating, treating, or preventing antibody-mediated disorders, such as autoimmune or inflammatory diseases.

[0005] Efgartigimod is an engineered Fc from the za allotype of human immunoglobulin (Ig) gamma (IgG)1 that binds to human FcRn with nanomolar affinity. Efgartigimod encompasses the IgG1 Fc region and has been engineered using ABDEG technology to increase its affinity for FcRn at both physiological and acidic pH. The increased affinity of efgartigimod for FcRn at both acidic and physiological pH results in blockade of FcRn-mediated recycling of IgG. Efgartigimod is approved in the United States and Japan as a weekly intravenous injection for use in the treatment of generalized myasthenia gravis and is in development for the treatment of several other antibody-mediated disorders.

[0006] FcRn also binds to and recycles serum albumin, a regulator of serum cholesterol levels. Efgartigimod advantageously has no effect on serum albumin levels in human subjects. However, recently, it has been shown that anti-FcRn antibodies can cause a reduction in serum albumin levels and an accompanying increase in serum cholesterol levels in human subjects, both of which are undesirable.

[0007] Accordingly, there is a need in the art for improved agents for use in the treatment of antibody-mediated disorders that antagonize FcRn binding to IgG and have a longer half-life, lower dose, less frequent dosing, better maintenance of albumin levels, and / or reduced or eliminated FcRn catabolism. SUMMARY OF THE INVENTION

[0008] Broadly, the present disclosure is directed to neonatal Fc receptor (FcRn) binding molecules linked to moieties that do not specifically bind to human antigens and methods of using them. Unexpectedly, it is shown for the first time in the present application that including a single non-human antigen-binding moiety can increase the stability and FcRn occupancy of the FcRn binding molecule.

[0009] In one aspect, provided herein is an FcRn-binding molecule linked to a moiety that does not specifically bind to a human antigen, wherein the molecular weight of the FcRn-binding molecule linked to the moiety is from about 50 to about 140 kilodaltons (kDa). In some embodiments, the FcRn-binding molecule is an FcRn antagonist. In some embodiments, the FcRn-binding molecule is a variant Fc region that binds to FcRn with increased affinity and reduced pH-dependence relative to the wild-type Fc region. In some embodiments, the moiety is not polyethylene glycol. In some embodiments, the moiety is a polypeptide and optionally an antigen-binding domain that does not specifically bind to a human antigen. In some embodiments, the molecule that does not specifically bind to a human antigen is an antigen-binding domain that specifically binds to a non-human antigen.

[0010] In some embodiments, provided is an FcRn / antigen-binding molecule comprising an FcRn-binding molecule and only one antigen-binding domain linked to the FcRn-binding molecule. In some embodiments, the antigen-binding domain specifically binds to a non-human antigen.

[0011] In some embodiments, the pharmacokinetics and / or pharmacodynamics of the FcRn-binding molecule are improved compared to efgartigimod.

[0012] In some embodiments, the FcRn / antigen-binding molecule has a molecular weight of 50 to 140 kilodaltons (kDa). In some embodiments, the FcRn / antigen-binding molecule or the FcRn-binding molecule has a molecular weight of about 60 kDa to about 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or the FcRn-binding molecule has a molecular weight of 60 kDa to 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or the FcRn-binding molecule has a molecular weight of about 60 kDa. In some embodiments, the FcRn / antigen-binding molecule or the FcRn-binding molecule has a molecular weight of about 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or the FcRn-binding molecule has a molecular weight of 60 kDa. In some embodiments, the FcRn / antigen-binding molecule or the FcRn-binding molecule has a molecular weight of 104 kDa.

[0013] In some embodiments, the FcRn-binding molecule is a variant Fc region, and the variant Fc region includes a first Fc domain and a second Fc domain that form a dimer, and at least one of the Fc domains includes amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively.

[0014] In some embodiments, the variant Fc region consists of a first Fc domain and a second Fc domain that form a dimer.

[0015] In some embodiments, administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or the FcRn-binding molecule to a subject results in a reduction of serum IgG in the subject. In some embodiments, administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or the FcRn-binding molecule to a subject results in a reduction of serum IgG in the subject of at least about 35%, about 40%, about 50%, about 55%, or about 60% compared to the baseline level of serum IgG in the subject.

[0016] In some embodiments, administration of a therapeutically effective amount of an FcRn / antigen binding molecule or an FcRn binding molecule to a subject does not reduce the FcRn level and / or albumin level in the subject.

[0017] In some embodiments, the clearance of an FcRn / antigen binding molecule or an FcRn binding molecule in a subject is less than about 2, about 1.9, about 1.8, about 1.7, about 1.6, about 1.5, about 1.4, about 1.3, about 1.2, about 1.1, about 1, about 0.9, about 0.8, about 0.7, about 0.6, or about 0.5 mL / hour / kg after a single administration of a therapeutically effective amount of the FcRn / antigen binding molecule or the FcRn binding molecule to the subject.

[0018] In some embodiments, the half-life (t 1 / 2 ) of an FcRn / antigen binding molecule or an FcRn binding molecule in a subject is greater than about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, or about 100 hours after a single administration of a therapeutically effective amount of the FcRn / antigen binding molecule or the FcRn binding molecule to the subject.

[0019] In some embodiments, an FcRn / antigen binding molecule or an FcRn binding molecule binds to FcRn at pH 5.5 and / or pH 6.0 and, optionally, binds with a higher affinity to FcRn at pH 5.5 and / or pH 6.0 than the affinity of efgartigimod for FcRn at pH 5.5 and / or pH 6.0 as measured by surface plasmon resonance.

[0020] In some embodiments, an FcRn / antigen binding molecule or an FcRn binding molecule binds to FcRn at pH 5.5 and / or pH 6.0 and, optionally, binds with a lower affinity to FcRn at pH 5.5 and / or pH 6.0 than the affinity of efgartigimod for FcRn at pH 5.5 and / or pH 6.0 as measured by surface plasmon resonance.

[0021] In some embodiments, the antigen-binding domain is selected from Fab domains, sdAbs, scFvs, and antibody mimetics. In some embodiments, the antibody mimetic is an anticalin or a DARPin. In some embodiments, the sdAb is a VHH fragment.

[0022] In some embodiments, the non-human antigen is a non-human antigen that can be found in humans. In some embodiments, the non-human antigen that can be found in humans is a viral antigen.

[0023] In some embodiments, the non-human antigen is a non-human antigen that is not found in humans. In some embodiments, the non-human antigen that is not found in humans is selected from hen egg white lysozyme (HEL) or ovalbumin. In some embodiments, the non-human antigen that is not found in humans is HEL, the antigen-binding domain is a Fab fragment or an scFv, and the Fab fragment or scFv comprises the HCDR1 amino acid sequence, the HCDR2 amino acid sequence, and the HCDR3 amino acid sequence of VH comprising the amino acid sequence of SEQ ID NO: 25, and the LCDR1 amino acid sequence, the LCDR2 amino acid sequence, and the LCDR3 amino acid sequence of VL comprising the amino acid sequence of SEQ ID NO: 24.

[0024] In some embodiments, the non-human antigen that is not found in humans is HEL, the antigen-binding domain is a Fab or scFv domain, and the Fab or scFv domain comprises VH and VL, and the VH domain and the VL domain comprise CDR sequences selected from the group consisting of HCDR1 (SEQ ID NO: 18), HCDR2 (SEQ ID NO: 19), HCDR3 (SEQ ID NO: 20), LCDR1 (SEQ ID NO: 21), LCDR2 (SEQ ID NO: 22), and LCDR3 (SEQ ID NO: 23).

[0025] In some embodiments, VH comprises, or consists of, an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 25, and VL comprises, or consists of, an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 24.

[0026] In some embodiments, VH comprises, or consists of, the amino acid sequence of SEQ ID NO: 25, and VL comprises, or consists of, the amino acid sequence of SEQ ID NO: 24.

[0027] In some embodiments, the antigen-binding domain is fused to the C-terminus of the first Fc domain or the second Fc domain. In some embodiments, the N-terminus of the antigen-binding domain is fused to the C-terminus of the first Fc domain. In some embodiments, the N-terminus of the antigen-binding domain is fused to the C-terminus of the second Fc domain.

[0028] In some embodiments, the antigen-binding domain is fused to the N-terminus of the first Fc domain or the second Fc domain. In some embodiments, the N-terminus of the first Fc domain is fused to the C-terminus of the antigen-binding domain. In some embodiments, the N-terminus of the second Fc domain is fused to the C-terminus of the antigen-binding domain.

[0029] In some embodiments, the first Fc domain and / or the second Fc domain is an IgG Fc domain, such as an IgG1 Fc domain. In some embodiments, the first Fc domain and / or the second Fc domain is a human IgG Fc domain, such as a human IgG1 Fc domain.

[0030] In some embodiments, the variant Fc region comprises or consists of a first Fc domain and a second Fc domain that form a dimer, and the first Fc domain and / or the second Fc domain comprise amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively. In some embodiments, the first Fc domain and / or the second Fc domain comprise amino acids Y, T, E, K, F, and Y at EU positions 252, 254, 256, 433, 434, and 436, respectively.

[0031] In some embodiments, both the first Fc domain and the second Fc domain comprise amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively. In some embodiments, both the first Fc domain and the second Fc domain comprise amino acids Y, T, E, K, F, and Y at EU positions 252, 254, 256, 433, 434, and 436, respectively.

[0032] In some embodiments, both the first Fc domain and the second Fc domain are IgG Fc domains, such as human IgG Fc domains. In some embodiments, both the first Fc domain and the second Fc domain are IgG1 Fc domains, such as human IgG1 Fc domains.

[0033] In some embodiments, the antigen-binding domain is fused to the first Fc domain or the second Fc domain via a linker. In some embodiments, the linker is a non-cleavable linker. In some embodiments, the linker is a peptide linker. In some embodiments, the peptide linker is a GS linker and is optionally 8 to 40 amino acids in length and optionally 20 or 30 amino acids in length.

[0034] In some embodiments, the antigen-binding domain is fused to the first Fc domain or the second Fc domain via an IgG hinge region or a portion thereof.

[0035] In some embodiments, the first Fc domain and / or the second Fc domain comprises an amino acid sequence independently selected from the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3. In some embodiments, the first Fc domain and / or the second Fc domain comprises the amino acid sequence of SEQ ID NO: 2.

[0036] In some embodiments, both the first Fc domain and the second Fc domain comprise an amino acid sequence independently selected from the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3. In some embodiments, both the first Fc domain and the second Fc domain comprise the amino acid sequence of SEQ ID NO: 2.

[0037] In some embodiments, the amino acid sequence of each of the first Fc domain and the second Fc domain consists of an amino acid sequence independently selected from the amino acid sequences set forth in SEQ ID NOs: 1, 2, and 3. In some embodiments, the amino acid sequence of the first Fc domain or the amino acid sequence of the second Fc domain consists of SEQ ID NO: 2.

[0038] In some embodiments, the amino acid sequences of both the first Fc domain and the second Fc domain consist of SEQ ID NO: 2.

[0039] In some embodiments, the variant Fc region comprises one or more mutations of amino acid residues that form the interface of the CH3 domain of the Fc domain. In some embodiments, the variant Fc region comprises a knob-into-hole mutation.

[0040] In some embodiments, the amino acid sequence of the first Fc domain further comprises amino acid W at EU position 366.

[0041] In some embodiments, the amino acid sequence of the first Fc domain comprises an amino acid sequence selected from the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6. In some embodiments, the amino acid sequence of the first Fc domain comprises the amino acid sequence of SEQ ID NO: 5.

[0042] In some embodiments, the amino acid sequence of the first Fc domain consists of an amino acid sequence selected from the amino acid sequences set forth in SEQ ID NOs: 4, 5, and 6. In some embodiments, the amino acid sequence of the first Fc domain consists of the amino acid sequence of SEQ ID NO: 5.

[0043] In some embodiments, the amino acid sequence of the second Fc domain further comprises amino acids S, A, and V at EU positions 366, 368, and 407, respectively.

[0044] In some embodiments, the amino acid sequence of the second Fc domain comprises an amino acid sequence selected from the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9. In some embodiments, the amino acid sequence of the second Fc domain comprises the amino acid sequence of SEQ ID NO: 8.

[0045] In some embodiments, the amino acid sequence of the second Fc domain consists of an amino acid sequence selected from the amino acid sequences set forth in SEQ ID NOs: 7, 8, and 9. In some embodiments, the amino acid sequence of the second Fc domain consists of the amino acid sequence of SEQ ID NO: 8.

[0046] In one aspect, provided herein is an FcRn / antigen-binding molecule comprising a first heavy chain, wherein the first heavy chain comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 34. In some embodiments, the first heavy chain comprises or consists of the amino acid sequence of SEQ ID NO: 34. In some embodiments, the FcRn / antigen-binding molecule further comprises a second heavy chain, and the second heavy chain consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 8. In some embodiments, the second heavy chain consists of the amino acid sequence of SEQ ID NO: 8.

[0047] In some embodiments, the FcRn / antigen-binding molecule further comprises a light chain, and the light chain comprises, or consists of, an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 24. In some embodiments, the light chain comprises, or consists of, the amino acid sequence of SEQ ID NO: 24.

[0048] Also provided are one or more isolated polynucleotides encoding any FcRn / antigen-binding molecule described herein, or any antigen-binding domain described herein, or any FcRn-binding molecule described herein.

[0049] Also provided is an expression vector comprising any one or more isolated polynucleotides described herein.

[0050] Also provided is a host cell comprising any one or more isolated polynucleotides or any expression vector described herein.

[0051] Also provided is a method for producing an FcRn / antigen-binding molecule, or an antigen-binding domain, or an FcRn-binding molecule, the method comprising culturing a host cell described herein under conditions that allow expression of the FcRn / antigen-binding molecule, or the antigen-binding domain, or the FcRn-binding molecule.

[0052] Also provided is a pharmaceutical composition comprising an FcRn / antigen-binding molecule described herein, or an antigen-binding domain described herein, or an FcRn-binding molecule described herein, and at least one pharmaceutically acceptable carrier.

[0053] Also provided is an FcRn / antigen-binding molecule described herein, or an antigen-binding domain described herein, or an FcRn-binding molecule described herein, or a pharmaceutical composition thereof, for use as a medicament.

[0054] Also provided is a method for reducing serum IgG in a subject in need thereof, the method comprising administering to the subject in need thereof a therapeutically effective amount of an FcRn / antigen-binding molecule described herein, or an antigen-binding domain described herein, or an FcRn-binding molecule described herein, or a pharmaceutical composition thereof.

[0055] Also provided is a method for treating an antibody-mediated disorder in a subject in need thereof, the method comprising administering to the subject in need thereof a therapeutically effective amount of an FcRn / antigen-binding molecule described herein, or an antigen-binding domain described herein, or an FcRn-binding molecule described herein, or a pharmaceutical composition thereof.

[0056] In some embodiments, the antibody-mediated disorder is an IgG-mediated disorder. In some embodiments, the antibody-mediated disorder is an autoimmune disease. In some embodiments, the autoimmune disease is allograft rejection, alopecia areata, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Addison's disease, Alzheimer's disease, antineutrophil cytoplasmic autoantibody (ANCA), autoimmune diseases of the adrenal gland, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune myocarditis, autoimmune neutropenia, autoimmune oophoritis and orchitis, immune thrombocytopenia (ITP or idiopathic thrombocytopenic purpura), idiopathic thrombocytopeniapurpura), immune-mediated thrombocytopenia, or idiopathic immune thrombocytopenia), autoimmune urticaria, Behçet's disease, bullous pemphigoid (BP), myocarditis, Castleman's disease, celiac sprue dermatitis, chronic fatigue immune dysfunction syndrome, chronic inflammatory demyelinating polyneuropathy (CIDP), Churg-Strauss syndrome, cicatricial pemphigoid, CREST syndrome, cold agglutinin disease, Crohn's disease, dilated cardiomyopathy, discoid lupus erythematosus, acquired epidermolysis bullosa, essential mixed cryoglobulinemia, factor VIII deficiency, fibromyalgia-fibromyositis, glomerulonephritis, Graves' disease, Guillain-Barré syndrome, Goodpasture's syndrome, graft-versus-host disease (GVHD), Hashimoto's thyroiditis, hemophilia A, idiopathic inflammatory myopathy (IIM), idiopathic membranous neuropathy, idiopathic pulmonary fibrosis, IgA neuropathy, IgM polyneuropathy, immune-mediated necrotizing myopathy (IMNM), juvenile arthritis, Kawasaki disease, lichen planus, lichen sclerosus, lupus erythematosus, lupus nephritis, Ménière's disease, mixed connective tissue disease, mucous membrane pemphigoid, multiple sclerosis, type 1 diabetes, multifocal motor neuropathy (MMN), myasthenia gravis (MG), generalized myasthenia gravis (gMG), myositis, paraneoplastic bullous pemphigoid, pemphigoid gestationis, pemphigus vulgaris (PV), pemphigus foliaceus (PF), pernicious anemia, polyarteritis nodosa, polychondritis, polyglandular syndrome, polymyalgia rheumatica, polymyositis, dermatomyositis (DM), necrotizing autoimmune myopathy (NAM), antisynthetase syndrome (ASyS), primary agammaglobulinemia, primary biliary cirrhosis, psoriasis, psoriatic arthritis, relapsing polychondritis, Raynaud's phenomenon, Reiter's syndrome, rheumatoid arthritis, sarcoidosis, scleroderma, Sjögren's syndrome, solid organ transplant rejection, stiff-man syndrome, systemic lupus erythematosus, Takayasu arteritis, toxic epidermal necrolysis (TEN), Stevens-Johnson syndrome (SJS), temporal arteritis / giant cell arteritis, thrombotic thrombocytopenic purpura, ulcerative colitis, uveitis, dermatitis herpetiformis vasculitis, antineutrophil cytoplasmic antibody-associated vasculitis, vitiligo, and Wegener's granulomatosis.

[0057] In some embodiments, the FcRn / antigen-binding molecule, or antigen-binding domain, or FcRn-binding molecule is administered to a subject either concomitantly with or sequentially to an additional therapeutic agent.

[0058] Also provided is an FcRn / antigen-binding molecule described herein, or an antigen-binding domain described herein, or an FcRn-binding molecule described herein for use in the treatment of antibody-mediated disorders.

[0059] Also provided is the use of an FcRn / antigen-binding molecule described herein, or an antigen-binding domain described herein, or an FcRn-binding molecule described herein in the manufacture of a medicament for treating an antibody-mediated disorder.

Brief Description of the Drawings

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Figure 1A

Figure 1B

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Figure 2

DETAILED DESCRIPTION OF THE INVENTION

[0062] The present disclosure provides engineered FcRn-binding molecules linked to a portion that does not specifically bind to a human antigen. In one aspect, the FcRn-binding molecule is linked to one antigen-binding domain (FcRn / antigen-binding molecule). In some embodiments, provided are FcRn / antigen-binding molecules comprising an FcRn-binding molecule linked to the antigen-binding domain at the C-terminus, or the N-terminus, or at a position other than the C- or N-terminus. In some embodiments, the antigen-binding domain specifically binds to a non-human antigen. Also provided herein are nucleic acids, vectors, host cells, methods of manufacture encoding such FcRn-binding molecules or FcRn / antigen-binding molecules, and methods for their use in the treatment of antibody-mediated disorders.

[0063] Definitions As used herein, the term "FcRn" refers to the neonatal Fc receptor. Exemplary FcRn molecules include human FcRn encoded by the FCGRT gene as shown in RefSeq NM 004107. The amino acid sequence of the corresponding protein is shown in RefSeq NP_004098.

[0064] As used herein, the term "FcRn binding molecule" refers to any agent that specifically binds to FcRn. As used herein, the term "FcRn antagonist" refers to any agent that specifically binds to FcRn and inhibits the binding of an immunoglobulin to FcRn (e.g., human FcRn). In one embodiment, the FcRn antagonist comprises an Fc region (e.g., a variant Fc region disclosed herein), specifically binds to FcRn via the Fc region, and inhibits the binding of an immunoglobulin to FcRn. In one embodiment, the FcRn antagonist is not a full-length IgG antibody. In one embodiment, the FcRn antagonist comprises an antigen-binding domain that binds to a target antigen and a variant Fc region. In one embodiment, the term "FcRn antagonist" refers to an antibody or an antigen-binding fragment thereof that specifically binds to FcRn via its antigen-binding domain or its Fc region and inhibits the binding of the Fc region of an immunoglobulin (e.g., an IgG autoantibody) to FcRn. As used herein, the term "FcRn / antigen binding molecule" refers to any agent that specifically binds to FcRn and specifically binds to another human antigen that is not a human antigen. In some embodiments, the antigen is HEL. As used herein, the term "affinity" or "binding affinity" refers to the strength of the binding interaction between two molecules. As used herein, the term "equilibrium dissociation constant" or "K D " refers to the tendency of the binding complex of two molecules to dissociate into two free molecules. Thus, as the binding affinity increases, K D decreases.

[0065] As used herein, the term "specifically binds" refers to the ability of any molecule to preferentially bind to a given target. For example, a molecule that specifically binds to a given target can bind to other molecules and generally can bind with a lower affinity, as determined by, for example, an immunoassay, BIAcore™, KinExA 3000 instrument (Sapidyne Instruments, Boise, ID), or other assays known in the art. In a specific embodiment, a molecule that specifically binds to a given target has a K D or K D less than when binding to an antigen by at least 2 logs, 2.5 logs, 3 logs, 4 logs.

[0066] As used herein, the term "operably linked" refers to the linkage of polynucleotide sequence elements that are in a functional relationship. For example, a polynucleotide sequence is operably linked when placed in a functional relationship with another polynucleotide sequence. In some embodiments, a transcriptional regulatory polynucleotide sequence, such as a promoter, enhancer, or other expression control element, is operably linked to a polynucleotide sequence encoding a protein if it affects the transcription of the polynucleotide sequence encoding the protein. The operably linked elements can be continuous or discontinuous.

[0067] As used herein, the term "linked" refers to a physical linkage (e.g., directly or indirectly linked) between amino acid sequences (e.g., different segments, regions, fragments, or domains). The linked regions, fragments, domains, and segments of the FcRn / antigen-binding molecules of the present disclosure can be continuous or discontinuous (e.g., linked to each other via a linker). In some embodiments, the linkage is a covalent bond. In some embodiments, the linkage is a non-covalent bond.

[0068] As used herein, the term "covalently linked" refers to the linking of two molecules or chemical moieties by a covalent bond. In some embodiments, the covalent bond is a peptide bond or a disulfide bond. As used herein, the term "fused" refers to the linking of two peptides by a peptide bond or a peptide linker. In some embodiments, two proteins are fused together directly and continuously by a peptide bond. In some embodiments, two proteins are fused indirectly and discontinuously via a peptide linker. In some embodiments, one protein is fused to the peptide linker at a first position by a peptide bond, and a second protein is fused to the peptide linker at a second position by a peptide bond. As used herein, the term "non-covalently linked" refers to the linking of two molecules or chemical moieties by a non-covalent interaction or a non-covalent bond. In some embodiments, the non-covalent interaction or non-covalent bond includes hydrogen bonding, electrostatic bonding or interaction, halogen bonding, pi-stacking, and van der Waals interactions.

[0069] The determination of the "percent identity" between two sequences (e.g., amino acid or nucleic acid sequences) can be accomplished using a mathematical algorithm. Specific, non-limiting examples of mathematical algorithms used for comparing two sequences are the algorithms of Karlin S & Altschul SF, (1990) PNAS 87:2264-2268, as modified in Karlin S & Altschul SF, (1993) PNAS 90:5873-5877, each of which is incorporated herein by reference in its entirety. Such algorithms are incorporated into the NBLAST and XBLAST programs of Altschul SF et al., (1990) J Mol Biol 215:403, which is incorporated herein by reference in its entirety. BLAST nucleotide searches can be performed with the NBLAST nucleotide program parameters set, e.g., score = 100, wordlength = 12, to obtain nucleotide sequences homologous to the nucleic acid molecules described herein. BLAST protein searches can be performed with the XBLAST program parameters set, e.g., score = 50, wordlength = 3, to obtain amino acid sequences homologous to the protein molecules described herein. To obtain gapped alignments for comparison purposes, gapped BLAST can be utilized as described in Altschul SF et al., (1997) Nuc Acids Res 25:3389-3402. Alternatively, PSI BLAST can be used to perform iterative searches for detecting distant relationships between molecules. Id. When utilizing the BLAST, gapped BLAST, and PSI BLAST programs, the default parameters of each program (e.g., of XBLAST and NBLAST) can be used (see, e.g., National Center for Biotechnology Information (NCBI) on the worldwide web, ncbi.nlm.nih.gov).Another specific non-limiting example of a mathematical algorithm used for array comparison is the algorithm of Myers and Miller, (1988) CABIOS 4:11-17, which is hereby incorporated by reference in its entirety. Such an algorithm is incorporated into the ALIGN program (version 2.0), which is part of the GCG sequence alignment software package. When utilizing the ALIGN program to compare amino acid sequences, a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4 may be used.

[0070] Percent identity between two arrays can be determined using techniques similar to those above, with or without allowing gaps. In calculating percent identity, typically only exact matches are counted.

[0071] As used herein, the terms “antibody” and “antibodies” include full-length antibodies, antigen-binding fragments of full-length antibodies, and molecules comprising antibody CDRs, VH domains (VH), or VL domains (VL). Examples of antibodies include monoclonal antibodies, recombinantly produced antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), human antibodies, humanized antibodies, chimeric antibodies, immunoglobulins, synthetic antibodies, tetrameric antibodies comprising two heavy chains and two light chain molecules, antibody light chain monomers, antibody heavy chain monomers, antibody light chain dimers, antibody heavy chain dimers, antibody light chain-heavy chain pairs, intrabodies, heteroconjugate antibodies, antibody-drug conjugates, single domain antibodies (sdAb), monovalent antibodies, single-chain antibodies or single-chain Fv (scFv), camelid antibodies, humanized antibodies, affibody molecules, VHH fragments, Fab fragments, F(ab’)2 fragments, disulfide-linked Fv (sdFv), anti-idiotype (anti-Id) antibodies (including, e.g., anti-anti-Id antibodies), and antigen-binding fragments of any of the foregoing. Antibodies can be of any isotype (e.g., IgG, IgE, IgM, IgD, IgA, or IgY), any subclass (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, or IgA2), or species (e.g., mouse IgG2a or IgG 2b can be of an immunoglobulin molecule of ().

[0072] As used herein, the term "antigen-binding domain" (or "antigen binding domain") refers to any polypeptide that specifically binds to an antigen. Examples of antigen-binding domains include antibodies, such as Fab fragments, F(ab’)2 fragments, disulfide-linked Fv (sdFv), single-chain Fv (scFv), CDRs, VH domains (VH), VL domains (VL), single-domain antibodies (sdAb), VHH fragments, camelid antibodies, and polypeptides derived from any of the above antigen-binding fragments. The term also encompasses synthetic antigen-binding proteins or antibody mimetic proteins, such as anticalins and DARPins.

[0073] In some embodiments, the antigen-binding domain is a VHH fragment. In some embodiments, the VHH fragment has one or more additional amino acids at its C-terminus. In some embodiments, the one or more additional amino acids are selected from the group consisting of A, AG, GG, and PP.

[0074] As used herein, the term "Fc region" refers to the portion of an immunoglobulin formed by the Fc domains of its two heavy chains. The Fc region can be a wild-type Fc region (native Fc region) or a variant Fc region. The native Fc region is a homodimer. The Fc region can be derived from any native immunoglobulin. In some embodiments, the Fc region is formed from an IgA, IgD, IgE, or IgG heavy chain constant region. In some embodiments, the Fc region is formed from an IgG heavy chain constant region. In some embodiments, the IgG heavy chain is an IgG1, IgG2, IgG3, or IgG4 heavy chain constant region. In some embodiments, the Fc region is formed from an IgG1 heavy chain constant region. In some embodiments, the IgG1 heavy chain constant region includes the G1m1(a), G1m2(x), G1m3(f), or G1m17(z) allotype. See, e.g., Jefferis and Lefranc (2009) mAbs 1(4):332-338, and de Taeye et al., (2020) Front Immunol. 11:740, which are incorporated herein by reference in their entirety.

[0075] As used herein, the term "variant Fc region" refers to a variant of the Fc region that has one or more changes relative to the native Fc region. The modifications can include amino acid substitutions, additions and / or deletions, ligation of additional moieties, and / or modification of native glycans. This term encompasses heterodimeric Fc regions in which each of the constituent Fc domains is different. This term also encompasses single-chain Fc regions in which the constituent Fc domains are linked together by a linker moiety.

[0076] As used herein, the term "Fc domain" refers to the portion of a single immunoglobulin heavy chain that includes both the CH2 and CH3 domains of the antibody. In some embodiments, the Fc domain includes at least a portion of the hinge (e.g., upper, middle, and / or lower hinge region) region, the CH2 domain, and the CH3 domain. In some embodiments, the Fc domain does not include the hinge region.

[0077] As used herein, the term "hinge region" refers to the portion of the heavy chain molecule that links the CH1 domain to the CH2 domain. In some embodiments, the hinge region is up to 70 amino acid residues in length. In some embodiments, this hinge region contains approximately 11-17 amino acid residues, is flexible, and thus allows the two N-terminal antigen-binding regions to move independently. In some embodiments, the hinge region is 12 amino acid residues in length. In some embodiments, the hinge region is 15 amino acid residues in length. In some embodiments, the hinge region is 62 amino acid residues in length. The hinge region can be subdivided into three different domains: an upper, middle, and lower hinge domain. The FcRn / antigen-binding molecules of the present disclosure can include all or any portion of the hinge region. In some embodiments, the hinge region is derived from an IgG1 antibody. In some embodiments, the hinge region comprises the amino acid sequence EPKSCDKTHTCPPCP (SEQ ID NO: 33).

[0078] As used herein, the term "FcRn-binding fragment" refers to a portion of an FcRn-binding molecule, e.g., a portion of the Fc region, that is sufficient to confer FcRn binding.

[0079] As used herein, the terms "one-armed", "one armed", "one-arm", "one arm", or "OA" refer to an FcRn / antigen-binding molecule comprising an FcRn-binding molecule linked to only one antigen-binding domain. In some embodiments, "one-armed", "one armed", "one-arm", "one arm", or "OA" refers to an FcRn / antigen-binding molecule comprising an Fc region comprising the Fc domains of two heavy chains, wherein one of the Fc domains of the two heavy chains is linked to the antigen-binding domain and the other Fc domain of the two heavy chains is not linked to the antigen-binding domain. In some embodiments, the antigen-binding domain is linked to the C-terminus of one of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is linked to the N-terminus of one of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is linked to a position other than the N-terminus or C-terminus of one of the Fc domains of the two heavy chains. The linkage can be a covalent or non-covalent bond. In some embodiments, the antigen-binding domain is fused to the C-terminus of one of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is fused to the N-terminus of one of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is fused to a position other than the N-terminus or C-terminus of one of the Fc domains of the two heavy chains.

[0080] As used herein, the terms "two-armed", "two armed", "two-arm", "two arm", or "TA" refer to an FcRn / antigen-binding molecule that includes an FcRn-binding molecule linked to two antigen-binding domains. In some embodiments, "two-armed", "two armed", "two-arm", "two arm", or "TA" refers to an FcRn / antigen-binding molecule that includes an Fc region that includes the Fc domains of two heavy chains, wherein each of the Fc domains of the two heavy chains is linked to an antigen-binding domain. In some embodiments, the antigen-binding domain is linked to the C-terminus of each of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is linked to the N-terminus of each of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is linked to a position other than the N-terminus or C-terminus of each of the Fc domains of the two heavy chains. In some embodiments, one of the antigen-binding domains is linked to the N-terminus of one of the Fc domains of the two heavy chains, and the other antigen-binding domain is linked to the C-terminus of the other Fc domain of the two heavy chains. In some embodiments, one of the antigen-binding domains is linked to a position other than the N-terminus or C-terminus of one of the Fc domains of the two heavy chains, and the other antigen-binding domain is linked to the N-terminus of the other Fc domain of the two heavy chains. In some embodiments, one of the antigen-binding domains is linked to a position other than the N-terminus or C-terminus of one of the Fc domains of the two heavy chains, and the other antigen-binding domain is linked to the C-terminus of the other Fc domain of the two heavy chains. The linkage can be a covalent or non-covalent bond. In some embodiments, the antigen-binding domain is fused to the C-terminus of each of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is fused to the N-terminus of each of the Fc domains of the two heavy chains. In some embodiments, the antigen-binding domain is fused to a position other than the N-terminus or C-terminus of each of the Fc domains of the two heavy chains.In some embodiments, one of the antigen-binding domains is fused to the N-terminus of one of the Fc domains of the two heavy chains, and the other antigen-binding domain is fused to the C-terminus of the other Fc domain of the two heavy chains. In some embodiments, one of the antigen-binding domains is fused at a position other than the N-terminus or C-terminus of one of the Fc domains of the two heavy chains, and the other antigen-binding domain is fused to the N-terminus of the other Fc domain of the two heavy chains. In some embodiments, one of the antigen-binding domains is fused at a position other than the N-terminus or C-terminus of one of the Fc domains of the two heavy chains, and the other antigen-binding domain is fused to the C-terminus of the other Fc domain of the two heavy chains.

[0081] As used herein, the term "EU position" refers to the amino acid positions in the EU numbering notation for the Fc region as described in Edelman, G. M. et al. Proc. Natl. Acad. USA, 63, 78 - 85 (1969), and Rabat et al., "Sequences of Proteins of Immunological Interest," U.S. Dept. Health and Human Services, 5 th edition, 1991.

[0082] As used herein, the term "antibody-mediated disorder" refers to any disorder in which the symptoms of the disorder are caused by one or more (auto)antibodies at abnormal levels in a subject. As used herein, the term "autoantibody-mediated disorder" refers to any disease or disorder in which the underlying pathology is at least partially caused by pathogenic IgG autoantibodies.

[0083] As used herein, the terms "treating," "treatment," and "treat" refer to the therapeutic or prophylactic measures described herein. A "treatment" method uses the administration of a polypeptide to a subject having a disease or disorder, or susceptible to such a disease or disorder, to prevent, cure, delay, reduce the severity of, or improve one or more symptoms of the disease or disorder, or recurrent disease or disorder, or to extend the lifespan of the subject beyond that expected in the absence of such treatment. In some embodiments, a "treatment" method uses the administration of a polypeptide to a subject having a disease or disorder, or predisposed to having such a disease or disorder, to prevent, cure, delay, reduce the severity of, or remit the disease or disorder or recurrent disease or disorder.

[0084] As used herein, in the context of performing a therapy on a subject, the term "effective amount" refers to the amount of the therapy that achieves the desired prophylactic or therapeutic effect.

[0085] As used herein, the term "dosage" or "dose" refers to the amount of a drug administered to a subject in a single administration.

[0086] As used herein, both the terms "fixed dose" and "uniform dose" refer to a dose that does not vary based on characteristics of the subject (e.g., weight, e.g., within a set range; gender; age, e.g., within a set range, etc.).

[0087] As used herein, the term "equivalent dose" refers to the doses of the first and second therapeutic agents where the number of molecules of the first and second therapeutic agents is substantially the same. In some embodiments, the equivalent dose is an equimolar dose. As used herein, the term "equimolar dose" refers to the doses of the first and second therapeutic agents where the number of moles of the first and second agents is the same. In some embodiments, the first agent is an FcRn / antigen-binding molecule and the second agent is efgartigimod. In some embodiments, the equivalent dose is calculated using the observed molecular weights of the first and second agents. In some embodiments, the equivalent dose is calculated using the predicted molecular weights of the first and second agents. In some embodiments, the equivalent dose is calculated using the observed molecular weight of the first agent and the predicted molecular weight of the second agent. In some embodiments, the equivalent dose is calculated using the predicted molecular weight of the first agent and the observed molecular weight of the second agent.

[0088] As used herein, the terms "pharmacodynamics" and "PD" refer to the biological effects of a therapeutic agent on an organism. In some embodiments, the biological effect is the modulation of the amount of circulating IgG in an organism to which the therapeutic agent has been administered. In some embodiments, the biological effect is the modulation of the amount of circulating albumin in an organism to which the therapeutic agent has been administered. As used herein, the term "improved pharmacodynamics" or "improved PD" refers to an improvement in the desired biological effect in an organism to which the therapeutic agent has been administered. In some embodiments, improved pharmacodynamics includes a reduction in the amount of circulating IgG in a subject. In some embodiments, improved pharmacodynamics includes the maintenance of the amount of circulating albumin in a subject. In some embodiments, improved pharmacodynamics includes a reduction in the amount of circulating IgG in a subject and the maintenance of the amount of circulating albumin in a subject. In some embodiments, the therapeutic agent is an FcRn / antigen-binding molecule.

[0089] As used herein, the terms "pharmacokinetics" and "PK" refer to the effect of an organism on a therapeutic agent administered to the organism. In some embodiments, the effect is the metabolism and / or clearance of the therapeutic agent. In some embodiments, PK refers to the rate of metabolism and / or clearance of the therapeutic agent. As used herein, the term "improved pharmacokinetics" or "improved PK" refers to an improvement in the desired effect of an organism on a therapeutic agent administered to the organism. In some embodiments, improved pharmacokinetics includes an increase in the half-life (T 1 / 2 1 / 2 ) of the therapeutic agent, clearance, or area under the curve (AUC) in a subject. In some embodiments, the therapeutic agent is an FcRn / antigen-binding molecule.

[0090] As used herein, the term "subject", or "patient", or "participant" includes any human or non-human animal. In one embodiment, the subject, or patient, or participant is a human or non-human mammal. In one embodiment, the subject, or patient, or participant is a human.

[0091] As used herein, the term "about" or "approximately" when referring to a measurable value such as a dosage amount, encompasses a variation of ±20%, ±15%, ±10%, ±5%, ±1%, or ±0.1% of the given value or range as appropriate to carry out the methods disclosed herein.

[0092] As used herein, the term "molecular weight" can refer to "predicted molecular weight" or "observed molecular weight". The "predicted molecular weight" of a protein is the sum of the molecular weights of all amino acids in the protein. In certain situations, the "predicted molecular weight" can differ from the "observed molecular weight" of the molecule. In some embodiments, these differences can occur in a protein due to changes in glycosylation, glycanation, ubiquitination, phosphorylation, or protein cleavage of the protein or an additional protein complex with a given protein.

[0093] FcRn-binding molecule The FcRn-binding molecules disclosed herein include any molecule that binds to FcRn, and the molecule includes, but is not limited to, any anti-FcRn antibody, any anti-FcRn binding region, or any Fc domain or Fc region.

[0094] In some embodiments, the FcRn-binding molecules disclosed herein are linked to a moiety that does not specifically bind to a human antigen. In some embodiments, the moiety that does not specifically bind to a human antigen increases the half-life of the FcRn-binding molecule without inducing any biological effect of its own. In some embodiments, the moiety does not specifically bind to an antigen. In some embodiments, the moiety is a polypeptide. In some embodiments, the polypeptide has the structure of an antigen-binding domain but does not specifically bind to an antigen. In some embodiments, the polypeptide is an antigen-binding domain that specifically binds to a non-human antigen. Non-limiting examples of antigen-binding domains that specifically bind to non-human antigens are provided herein.

[0095] In some embodiments, the FcRn-binding molecule is an FcRn antagonist that includes any molecule that binds to and inhibits FcRn, and the molecule includes, but is not limited to, any anti-FcRn antibody, any anti-FcRn binding region, or any Fc domain or Fc region.

[0096] In some embodiments, the FcRn-binding molecules disclosed herein include 2, 3, or 4 FcRn-binding regions, e.g., Fc regions.

[0097] In some embodiments, the FcRn-binding molecules disclosed herein include one or more Fc regions or FcRn-binding fragments thereof in combination with one or more antigen-binding domains (e.g., sdAb, Fab fragment, scFv, or antibody mimetic).

[0098] Any Fc region can be mutated to produce a variant Fc region disclosed herein. Generally, the Fc region or its FcRn-binding fragment is from a human immunoglobulin. However, it is understood that the Fc region can be derived from the immunoglobulin of any other mammalian species, including, for example, camelid species, rodents (e.g., mice, rats, rabbits, guinea pigs) or non-human primate (e.g., chimpanzee, macaque) species. Also, the Fc region or its FcRn-binding portion can be derived from any immunoglobulin class, including IgM, IgG, IgD, IgA, and IgE, and any immunoglobulin isotype, including IgG1, IgG2, IgG3, and IgG4. In one embodiment, the Fc region is an IgG Fc region (e.g., a human IgG region). In one embodiment, the Fc region is an IgG1 Fc region (e.g., a human IgG1 region). In one embodiment, the Fc region is a chimeric Fc region that includes portions of several different Fc regions. Suitable examples of chimeric Fc regions are described in US2011 / 0243966A1, which is incorporated herein by reference in its entirety. Various Fc region gene sequences (e.g., human constant region gene sequences) are available in the form of publicly available deposits.

[0099] The Fc region may be further cleaved to produce its minimal FcRn-binding fragment or may be internally deleted. The ability of an Fc region fragment to bind to FcRn can be determined using any binding assay recognized in the art, such as ELISA.

[0100] To enhance the manufacturability of the FcRn-binding molecules and FcRn / antigen-binding molecules containing them disclosed herein, the Fc region as a composition preferably does not contain any non-disulfide-bonded cysteine residues. Thus, in one embodiment, the Fc region does not contain free cysteine residues.

[0101] In some embodiments, any Fc variant or its FcRn-binding fragment that specifically binds to FcRn with increased affinity and reduced pH-dependence relative to the native Fc region can be used herein. In one embodiment, the variant Fc region comprises amino acid changes, substitutions, insertions, and / or deletions that confer the desired characteristics. In some embodiments, the FcRn-binding molecule comprises a variant Fc region or its FcRn-binding fragment, and the variant Fc region or its FcRn-binding fragment binds to FcRn with higher affinity at pH 5.5 compared to the corresponding wild-type Fc region. In some embodiments, the FcRn-binding molecule comprises a variant Fc region or its FcRn-binding fragment, and the variant Fc region or its FcRn-binding fragment binds to FcRn with higher affinity at pH 6.0 and / or pH 7.4 compared to the corresponding wild-type Fc region. In some embodiments, the FcRn-binding molecule comprises a variant Fc region or its FcRn-binding fragment, and the variant Fc region or its FcRn-binding fragment binds to FcRn with higher affinity at both acidic and neutral pHs compared to the corresponding wild-type Fc region.

[0102] In some embodiments, the variant Fc region is derived from the Fc region of any native immunoglobulin. In some embodiments, the native immunoglobulin is a human immunoglobulin. In some embodiments, the immunoglobulin is IgA, IgD, IgE, or IgG. In some embodiments, the immunoglobulin is IgG. In some embodiments, the immunoglobulin is human IgA, human IgD, human IgE, or human IgG. In some embodiments, the immunoglobulin is human IgG. In some embodiments, IgG is IgG1, IgG2, IgG3, or IgG4. In some embodiments, human IgG is human IgG1, human IgG2, human IgG3, or human IgG4. In some embodiments, the variant Fc region varies from the human IgG1 Fc region. In some embodiments, the human IgG1 Fc region comprises the G1m1(a), G1m2(x), G1m3(f), or G1m17(z) allotype.

[0103] In some embodiments, the FcRn binding molecule is an FcRn antagonist.

[0104] Other known Fc domain variants that can be used in the compositions disclosed herein are described in Ghetie et al, 1997, Nat. Biotech. 15:637-40, Duncan et al, 1988, Nature 332:563-564, Lund et al, 1991, J. Immunol, 147:2657-2662, Lund et al, 1992, Mol. Immunol, 29:53-59, Alegre et al, 1994, Transplantation 57:1537-1543, Hutchins et al, 1995, Proc Natl. Acad Sci USA, 92:11980-11984, Jefferis et al, 1995, Immunol Lett., 44:111-117, Lund et al, 1995, Faseb J., 9:115-119, Jefferis et al, 1996, Immunol Lett., 54:101-104, Lund et al, 1996, J. Immunol, 157:4963-4969, Armour et al, 1999, Eur J Immunol 29:2613-2624, Idusogie et al, 2000, J. Immunol, 164:4178-4184, Reddy et al, 2000, J. Immunol, 164:1925-1933, Xu et al, 2000, Cell Immunol, 200:16-26, Idusogie et al, 2001, J. Immunol, 166:2571-2575, Shields et al, 2001, J Biol. Chem., 276:6591-6604, Jefferis et al, 2002, Immunol Lett., 82:57-65, Presta et al, 2002, Biochem Soc Trans., 30:487-490), U.S. Patent No. 5,624,821, No. 5,885,573, No. 5,677,425, No. 6,165,745, No. 6,277,375, No. 5,869,046, No. 6,121,022, No. 5,624,821, No. 5,648,260, No. 6,528,624, No. 6,194,551, No. 6,737,056, No. 6,821,505, No. 6,277,375, U.S. Patent Publication No. 2004 / 0002587, and PCT Publication Nos. 94 / 29351, 99 / 58572, 00 / 42072, 02 / 060919, 04 / 029207, 04 / 099249, 04 / 063351, including but not limited to those disclosed therein, the contents of these documents are hereby incorporated by reference in their entirety.

[0105] In one embodiment, the variant Fc region or its FcRn-binding fragment comprises or consists of two Fc domains. In one embodiment, the variant Fc region or its FcRn-binding fragment comprises at least one Fc domain, and at least one Fc domain comprises the amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively. In one embodiment, the variant Fc region or its FcRn-binding fragment comprises at least one Fc domain, and at least one Fc domain comprises the amino acids Y, T, E, K, F, and Y at EU positions 252, 254, 256, 433, 434, and 436, respectively. In one embodiment, the variant Fc region or its FcRn-binding fragment consists of two Fc domains, both of which comprise the amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively. In one embodiment, the variant Fc region or its FcRn-binding fragment consists of two Fc domains, both of which comprise the amino acids Y, T, E, K, F, and Y at EU positions 252, 254, 256, 433, 434, and 436, respectively.

[0106]

[0107] In one embodiment, the amino acid sequence of the Fc domain of the variant Fc region comprises the amino acid sequence of SEQ ID NO: 1. In one embodiment, the amino acid sequence of the Fc domain of the variant Fc region consists of the amino acid sequence of SEQ ID NO: 1. In one embodiment, the amino acid sequence of the Fc domain of the variant Fc region comprises the amino acid sequence of SEQ ID NO: 2. In one embodiment, the amino acid sequence of the Fc domain of the variant Fc region consists of the amino acid sequence of SEQ ID NO: 2. In one embodiment, the amino acid sequence of the Fc domain of the variant Fc region comprises the amino acid sequence of SEQ ID NO: 3. In one embodiment, the amino acid sequence of the Fc domain of the variant Fc region consists of the amino acid sequence of SEQ ID NO: 3.

[0108] In one embodiment, the FcRn binding molecule comprises a variant Fc region, the variant Fc region comprises two Fc domains, and the amino acid sequence of each of the Fc domains is independently selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3.

[0109] In certain embodiments, the variant Fc region is a heterodimer and the constituent Fc domains are different from each other. Methods for producing Fc heterodimers are known in the art (see, e.g., US8,216,805, which is hereby incorporated by reference in its entirety). In one embodiment, the FcRn binding molecule consists of a variant Fc region, the variant Fc region consists of two Fc domains that form a heterodimer, and the amino acid sequence of each of the Fc domains is independently selected from SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3. In one embodiment, the FcRn binding molecule consists of or comprises a variant Fc region, the variant Fc region consists of or comprises two Fc domains that form a heterodimer, the amino acid sequence of the first Fc domain consists of or comprises the amino acid sequence of SEQ ID NO: 1, and the amino acid sequence of the second Fc domain consists of or comprises the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 3. In one embodiment, the FcRn binding molecule consists of or comprises a variant Fc region, the variant Fc region consists of or comprises two Fc domains that form a heterodimer, the amino acid sequence of the first Fc domain consists of or comprises the amino acid sequence of SEQ ID NO: 2, and the amino acid sequence of the second Fc domain consists of or comprises the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 3. In one embodiment, the FcRn binding molecule consists of or comprises a variant Fc region, the variant Fc region consists of or comprises two Fc domains that form a heterodimer, the amino acid sequence of the first Fc domain consists of or comprises the amino acid sequence of SEQ ID NO: 3, and the amino acid sequence of the second Fc domain consists of or comprises the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2.

[0110] In one embodiment, the FcRn binding molecule comprises a variant Fc region, the variant Fc region consists of or comprises two Fc domains that form a homodimer, and the amino acid sequence of each of the Fc domains consists of or comprises the amino acid sequence of SEQ ID NO: 1.

[0111] In one embodiment, the FcRn binding molecule comprises a variant Fc region, the variant Fc region consisting of or comprising two Fc domains that form a homodimer, and the amino acid sequence of each of the Fc domains consisting of or comprising the amino acid sequence of SEQ ID NO: 2.

[0112] In one embodiment, the FcRn binding molecule comprises a variant Fc region, the variant Fc region consisting of or comprising two Fc domains that form a homodimer, and the amino acid sequence of each of the Fc domains consisting of or comprising the amino acid sequence of SEQ ID NO: 3.

[0113] In one embodiment, the FcRn binding molecule comprises a variant Fc region, the variant Fc region comprising or consisting of efgartigimod (CAS Registry No. 1821402-21-4). As used herein, the term "efgartigimod" is interchangeable with "efgartigimod alpha". In some embodiments, efgartigimod is efgartigimod alpha-fcab.

[0114] In one embodiment, the variant Fc region is modified to promote heterodimerization. Such modifications are known in the art and any suitable means for promoting heterodimerization can be used to generate the FcRn / antigen-binding molecules described herein. In some embodiments, the variant Fc region comprises one or more mutations of amino acid residues that form the interface of the CH3 domain of the Fc domain. In some embodiments, the variant Fc region comprises a knob-into-hole mutation (see, e.g., WO 2006 / 028936, which is incorporated herein by reference in its entirety). In this technique, mispairing of Ig heavy chains is reduced by mutating selected amino acids that form the interface of the CH3 domain in IgG. At positions within the CH3 domain where two heavy chains interact directly, one or more amino acids with small side chains (holes) are introduced into the sequence of one heavy chain, and one or more amino acids with large side chains (knobs) are introduced within the corresponding interacting residue position(s) on the other heavy chain. The Fc domain of the Fc region can be composed of immunoglobulin chains of the same subclass (e.g., IgG1 or IgG3) or different subclasses (e.g., IgG1 and IgG3, or IgG3 and IgG4).

[0115] In some embodiments, the variant Fc region comprises or consists of two Fc domains, and one of the Fc domains comprises the amino acid W at EU position 366. In some embodiments, the variant Fc region comprises or consists of two Fc domains, and one of the Fc domains comprises the amino acids S, A, and V at EU positions 366, 368, and 407, respectively. In some embodiments, the variant Fc region comprises or consists of two Fc domains, one Fc domain comprises the amino acid W at EU position 366, and the other Fc domain comprises the amino acids S, A, and V at EU positions 366, 368, and 407, respectively.

[0116] In some embodiments, the variant Fc region comprises or consists of two Fc domains, wherein one Fc domain comprises amino acids E and D at EU positions 370 and 409, respectively, and the other Fc domain comprises amino acid K at EU positions 357 and 399. In some embodiments, the variant Fc region comprises or consists of two Fc domains, wherein one Fc domain comprises H and A at EU positions 364 and 405, respectively, and the other Fc domain comprises amino acids T and F at EU positions 349 and 394, respectively. In some embodiments, the variant Fc region comprises or consists of two Fc domains, wherein one Fc domain comprises amino acids V, Y, A, and V at EU positions 350, 351, 405, and 407, respectively, and the other Fc domain comprises amino acids V, L, L, and W at EU positions 350, 366, 392, and 394, respectively. In some embodiments, the variant Fc region comprises or consists of two Fc domains, wherein one Fc domain comprises amino acids D, M, and A at EU positions 360, 399, and 407, respectively, and the other Fc domain comprises amino acids R, R, V, and V at EU positions 345, 347, 366, and 409, respectively. In some embodiments, the variant Fc region comprises or consists of two Fc domains, wherein one Fc domain comprises amino acid D at EU positions 409 and 392, and the other Fc domain comprises amino acid K at EU positions 399 and 356. In some embodiments, the variant Fc region comprises or consists of two Fc domains, wherein one Fc domain comprises amino acids E, W, and C at EU positions 360, 409, and 349, respectively, and the other Fc domain comprises amino acids R, V, T, and C at EU positions 347, 399, 405, and 354, respectively.In some embodiments, the variant Fc region comprises or consists of two Fc domains, one Fc domain containing amino acids E and W at EU positions 370 and 409, respectively, and the other Fc domain containing amino acids N, V, and T at EU positions 357, 399, and 405, respectively.

[0117] In one embodiment, the FcRn-binding molecule consists of a variant Fc region, the variant Fc region comprising or consisting of two Fc domains that form a heterodimer, wherein the amino acid sequence of the first Fc domain is selected from the amino acid sequences comprising or consisting of SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, and / or the amino acid sequence of the second Fc domain is selected from the amino acid sequences comprising or consisting of SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9. In one embodiment, the FcRn-binding molecule consists of a variant Fc region, the variant Fc region comprising or consisting of two Fc domains that form a heterodimer, wherein the amino acid sequence of the first Fc domain is selected from the amino acid sequences comprising or consisting of SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9, and / or the amino acid sequence of the second Fc domain is selected from the amino acid sequences comprising or consisting of SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6. In one embodiment, the FcRn-binding molecule consists of a variant Fc region, the variant Fc region comprising or consisting of two Fc domains that form a heterodimer, wherein the amino acid sequence of the first Fc domain comprises or consists of SEQ ID NO: 4, and the amino acid sequence of the second Fc domain comprises or consists of SEQ ID NO: 7. In one embodiment, the FcRn-binding molecule consists of a variant Fc region, the variant Fc region comprising or consisting of two Fc domains that form a heterodimer, wherein the amino acid sequence of the first Fc domain comprises or consists of SEQ ID NO: 5, and the amino acid sequence of the second Fc domain comprises or consists of SEQ ID NO: 8. In one embodiment, the FcRn-binding molecule consists of a variant Fc region, the variant Fc region comprising or consisting of two Fc domains that form a heterodimer, wherein the amino acid sequence of the first Fc domain comprises or consists of SEQ ID NO: 6, and the amino acid sequence of the second Fc domain comprises or consists of SEQ ID NO: 9. In some embodiments, the FcRn-binding molecule is an FcRn antagonist. Table 1. Amino acid sequences of variant Fc regions [Table 1]

[0118] In some embodiments, the variant Fc region comprises a first Fc domain comprising the amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively, and a second Fc domain comprising the amino acids K and F at EU positions 433 and 434, respectively. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 1. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 2. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 3.

[0119] In some embodiments, the variant Fc region comprises a first Fc domain comprising the amino acids Y, T, E, W, K, and F at EU positions 252, 254, 256, 366, 433, and 434, respectively, and a second Fc domain comprising the amino acids S, A, V, K, and F at EU positions 366, 368, 407, 433, and 434, respectively. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 4. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 5. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 6.

[0120] In some embodiments, the variant Fc region comprises a first Fc domain comprising the amino acids Y, T, E, S, A, V, K, and F at EU positions 252, 254, 256, 366, 368, 407, 433, and 434, respectively, and a second Fc domain comprising the amino acids W, K, and F at EU positions 366, 433, and 434, respectively. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 7. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 8. In some embodiments, the first Fc domain comprises the amino acid sequence of SEQ ID NO: 9.

[0121] In one embodiment, the anti-FcRn antibody is rozanolixizumab (UCB7665), nipocalimab (M281), orilanolimab (ALXN1830 / SYNT001), or batoclimab (IMVT-1401 / RVT1401 / HBM9161).

[0122] In one embodiment, the antibody that specifically binds to FcRn and inhibits the binding of the Fc region of immunoglobulin to FcRn is nipocalimab, also known as M281. Nipocalimab is a full-length "Fc dead" IgG1 monoclonal antibody. Nipocalimab is being administered as an intravenous infusion in Phase 2 / 3 clinical trials for the treatment of myasthenia gravis (MG) and warm antibody autoimmune hemolytic anemia (WAIHA), and in Phase 2 clinical trials for the treatment of fetal and neonatal hemolytic disease (HDFN), systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and Sjögren's syndrome (SS). Nipocalimab contains the light chain (SEQ ID NO: 10) and heavy chain (SEQ ID NO: 11) sequences shown in Table 2 below (VL of SEQ ID NO: 10 and VH of SEQ ID NO: 11 are underlined). Table 2. Heavy and light chain sequences of nipocalimab

Table 2

[0123] In one embodiment, the antibody that specifically binds to FcRn and inhibits the binding of the Fc region of immunoglobulin to FcRn is rozanolixizumab, also known as UCB 7665. Rozanolixizumab is a full-length humanized IgG4 monoclonal antibody. Rozanolixizumab is being administered as a subcutaneous injection in clinical trials for MG, immune thrombocytopenia (ITP), chronic inflammatory demyelinating polyneuropathy (CIDP), autoimmune encephalitis (AIE), and myelin oligodendrocyte glycoprotein-related disease (MOG-AD). Rozanolixizumab contains the light chain (SEQ ID NO: 12) and heavy chain (SEQ ID NO: 13) sequences shown in Table 3 below (VL of SEQ ID NO: 12 and VH of SEQ ID NO: 13 are underlined). Table 3. Heavy and light chain sequences of rozanolixizumab

Table 3

[0124] In one embodiment, the antibody that specifically binds to FcRn and inhibits the binding of the Fc region of immunoglobulin to FcRn is orilanolimab, also known as SYNT001 / ALXN1830. Orilanolimab is another full-length humanized IgG4 monoclonal antibody. Orilanolimab is being administered as an intravenous infusion in Phase 2 clinical trials for the treatment of WAIHA and pemphigus. Orilanolimab contains the light chain (SEQ ID NO: 14) and heavy chain (SEQ ID NO: 15) sequences shown in Table 4 below (the VL of SEQ ID NO: 14 and the VH of SEQ ID NO: 15 are underlined). Table 4. Heavy and Light Chain Sequences of Orilanolimab

Table 4

[0125] In one embodiment, the antibody that specifically binds to FcRn and inhibits the binding of the Fc region of immunoglobulin to FcRn is batoclimab, also known as IMVT1401 / RVT1401 / HBM9161. Batoclimab is another full-length "Fc dead" IgG1 monoclonal antibody. Batoclimab is being administered as a subcutaneous injection in Phase 2 clinical trials for the treatment of MG, ITP, Graves' ophthalmopathy, thyroid ophthalmopathy, and neuromyelitis optica spectrum disorder (NMOSD). Batoclimab contains the light chain (SEQ ID NO: 16) and heavy chain (SEQ ID NO: 17) sequences shown in Table 5 below (the VL of SEQ ID NO: 16 and the VH of SEQ ID NO: 17 are underlined). Table 5. Heavy and Light Chain Sequences of Batoclimab

Table 5

[0126] FcRn / antigen binding molecule In one aspect, the present disclosure provides an FcRn / antigen-binding molecule or a fragment thereof. In some embodiments, the FcRn / antigen-binding molecules disclosed herein comprise an FcRn-binding molecule and only one antigen-binding domain. The FcRn-binding molecule can be any of the FcRn-binding molecules described herein. Similarly, the antigen-binding domain can be any of the antigen-binding domains described herein.

[0127] In some embodiments, the antigen-binding domain is linked to the C-terminus of the FcRn-binding molecule. In some embodiments, the antigen-binding domain is linked to the N-terminus of the FcRn-binding molecule. In some embodiments, the antigen-binding domain is linked to the FcRn-binding molecule at a position other than the C-terminus or N-terminus. The antigen-binding domain may be covalently or non-covalently linked to the FcRn-binding molecule.

[0128] In some embodiments, the antigen-binding domain is fused to the C-terminus of the FcRn-binding molecule. In some embodiments, the antigen-binding domain is fused to the N-terminus of the FcRn-binding molecule. In some embodiments, the antigen-binding domain is fused to the FcRn-binding molecule at a position other than the C-terminus or N-terminus.

[0129] In some embodiments, the FcRn-binding molecule is an Fc region, such as a variant Fc region. In some embodiments, the antigen-binding domain is linked or fused to the C-terminus of one of the Fc domains of the variant Fc region. In some embodiments, the antigen-binding domain is linked or fused to the N-terminus of one of the Fc domains of the variant Fc region. In some embodiments, the antigen-binding domain is linked or fused to the FcRn-binding molecule at a position other than the C-terminus or N-terminus.

[0130] In some embodiments, the antigen-binding domain may be directly linked to or directly fused with the N-terminus or C-terminus of the FcRn-binding molecule. In some embodiments, the antigen-binding domain is linked to the N-terminus or C-terminus of the FcRn-binding molecule via a linker. In some embodiments, the linker is a non-cleavable linker.

[0131] In some embodiments, the antigen-binding domain may be directly linked (e.g., fused) to the N-terminus or C-terminus of the Fc domain. In some embodiments, the antigen-binding domain is linked to the N-terminus or C-terminus of the Fc domain via a linker. The linker can be any suitable linker including those described herein.

[0132] Antigen-binding domain In one aspect, an antigen-binding domain is provided by the present disclosure. In some embodiments, the FcRn / antigen-binding molecules disclosed herein comprise one or more FcRn-binding molecules in combination with one or more antigen-binding domains. In some embodiments, the FcRn / antigen-binding molecules disclosed herein comprise one or more Fc regions or their FcRn-binding fragments in combination with one or more antigen-binding domains. In some embodiments, the antigen-binding domain is a polypeptide derived from an antibody, and the antibody includes, but is not limited to, sdAb (e.g., VHH fragment), Fab fragment, scFv, VH, or VL. In some embodiments, the antigen-binding domain is a synthetic antigen-binding protein or an antibody mimetic protein, and the synthetic antigen-binding protein or antibody mimetic protein includes, but is not limited to, anticalin or DARPin.

[0133] In some embodiments, the antigen-binding domain further comprises one or more amino acids added at its C-terminus. In some embodiments, the antigen-binding domain further comprises one or more amino acids added at the C-terminus, and the one or more amino acids are selected from A, AG, GG, and PP.

[0134] The antigen-binding domain can bind to any antigen. In some embodiments, the antigen is a non-human antigen, for example, a protein or a fragment thereof that is not normally expressed by humans and is not normally found in humans. In some embodiments, the non-human antigen is a protein or a fragment thereof that is not normally expressed by humans but can be found in humans. Examples of non-human antigens that can be found in humans include proteins or fragments thereof expressed by pathogens, such as bacterial or viral proteins or fragments thereof. These pathogenic proteins or fragments thereof can be found in humans due to infection and / or immunization. Thus, in some embodiments, the non-human antigen that can be found in humans is a viral antigen. In some embodiments, the virus is the rabies virus. In some embodiments, the antigen-binding domain that binds to the rabies virus is a VHH fragment. In some embodiments, the VHH fragment that binds to the rabies virus contains the amino acid sequence of SEQ ID NO: 29 shown in Table 6 below (the CDRs are underlined). In some embodiments, the VHH fragment that binds to the rabies virus contains a combination of CDR1, CDR2, and CDR3, where CDR1 is SSDMS (SEQ ID NO: 26), CDR2 is GINSGGGRTLYADSVKG (SEQ ID NO: 27), and CDR3 is SWYTDY (SEQ ID NO: 28). Table 6.3 Rab VHH

Table 6

[0135] In some embodiments, the non-human antigen is a non-human antigen that is not found in humans. Examples of non-human antigens that are not found in humans include non-pathogenic proteins or fragments thereof that do not have a human counterpart, such as hen egg white lysozyme (HEL) or ovalbumin.

[0136] In some embodiments, the antigen-binding domain specifically binds to HEL. In some embodiments, the antigen-binding domain specifically binds to HEL and is selected from a Fab fragment, a scFv, and an sdAb. In some embodiments, the antigen-binding domain specifically binds to HEL and is a Fab fragment or a scFv.

[0137] In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv comprises the HCDR1 amino acid sequence, the HCDR2 amino acid sequence, and the HCDR3 amino acid sequence of VH comprising the amino acid sequence of SEQ ID NO: 25, and comprises the LCDR1 amino acid sequence, the LCDR2 amino acid sequence, and the LCDR3 amino acid sequence of VL comprising the amino acid sequence of SEQ ID NO: 24.

[0138] In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv comprises a combination of VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, and VLCDR3, and 1, 2, 3, 4, or 5 amino acids differ in at least one of the amino acid sequences selected from the amino acid sequences of SEQ ID NOs: 18, 19, 20, 21, 22, and 23. In some embodiments, in some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv comprises a combination of VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, and VLCDR3 selected from SEQ ID NOs: 18, 19, 20, 21, 22, and 23.

[0139] In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv consists of a combination of VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, and VLCDR3, and 1, 2, 3, 4, or 5 amino acids differ in at least one of the amino acid sequences selected from SEQ ID NOs: 18, 19, 20, 21, 22, and 23. In some embodiments, in some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv consists of a combination of VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, and VLCDR3 selected from SEQ ID NOs: 18, 19, 20, 21, 22, and 23.

[0140] In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv comprises VH and VL, and VH comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 25, and VL comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 24. In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, and the Fab fragment or scFv comprises VH and VL, and VH comprises the amino acid sequence of SEQ ID NO: 25, and VL comprises the amino acid sequence of SEQ ID NO: 24.

[0141] In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, the Fab fragment or scFv comprises VH and VL, VH consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence selected from SEQ ID NO: 25, and VL consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence selected from SEQ ID NO: 24. In some embodiments, the antigen-binding domain is a Fab fragment or a scFv, the Fab fragment or scFv comprises VH and VL, VH consists of the amino acid sequence of SEQ ID NO: 25, and VL consists of the amino acid sequence of SEQ ID NO: 24.

[0142] In some embodiments, the antigen-binding domain specifically binds to a rabies virus protein, such as the envelope protein of the rabies virus. In some embodiments, the rabies virus protein is the rabies G protein. In some embodiments, the antigen-binding domain specifically binds to a rabies virus protein and is selected from a Fab fragment, a scFv, and an sdAb. In some embodiments, the antigen-binding domain specifically binds to a rabies virus protein and is an sdAb, such as a VHH fragment.

[0143] In some embodiments, the antigen-binding domain is a VHH fragment, and the VHH fragment comprises the CDR1 amino acid sequence, the CDR2 amino acid sequence, and the CDR3 amino acid sequence of the VHH fragment comprising the amino acid sequence of SEQ ID NO: 29.

[0144] In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment comprises a combination of CDR1, CDR2, and CDR3, and 1, 2, 3, 4, or 5 amino acids are different in at least one of the amino acid sequences selected from SEQ ID NOs: 26, 27, and 28. In some embodiments, the antigen-binding domain is a VHH fragment, and the VHH fragment comprises a combination of CDR1, CDR2, and CDR3 selected from SEQ ID NOs: 26, 27, and 28.

[0145] In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment consists of a combination of CDR1, CDR2, and CDR3, and 1, 2, 3, 4, or 5 amino acids differ in at least one of the amino acid sequences selected from SEQ ID NOs: 26, 27, and 28. In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment consists of a combination of CDR1, CDR2, and CDR3 selected from SEQ ID NOs: 26, 27, and 28.

[0146] In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 29. In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment comprises the amino acid sequence of SEQ ID NO: 29.

[0147] In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 29. In some embodiments, the antigen-binding domain is a VHH fragment, the VHH fragment consists of the amino acid sequence of SEQ ID NO: 29.

[0148] Linker The antigen-binding domain can be linked to the N-terminus or C-terminus of an FcRn-binding molecule (e.g., an Fc domain). Alternatively, the antigen-binding domain can be linked at a position other than the N-terminus or C-terminus of the FcRn-binding molecule (e.g., an Fc domain).

[0149] In some embodiments, the antigen-binding domain can be non-covalently linked to the FcRn-binding molecule. In some embodiments, the antigen-binding domain can be covalently linked to the FcRn-binding molecule.

[0150] In some embodiments, the antigen-binding domain can be directly linked (e.g., fused) to the N-terminus or C-terminus of the FcRn-binding molecule. In some embodiments, the antigen-binding domain is linked to the N-terminus or C-terminus of the FcRn-binding molecule via a linker. In some embodiments, the linker is a non-cleavable linker.

[0151] In some embodiments, the antigen-binding domain can be directly linked (e.g., fused) to the N-terminus or C-terminus of the Fc domain. In some embodiments, the antigen-binding domain is linked to the N-terminus or C-terminus of the Fc domain via a linker. In some embodiments, the linker is a non-cleavable linker. As used herein, the term "non-cleavable linker" refers to a linker that is not readily cleaved by one or more of a given enzyme, chemical agent, or light irradiation. In some embodiments, the enzyme is a protease.

[0152] In some embodiments, the linker is a synthetic compound linker, such as a chemical crosslinking agent. Non-limiting examples of suitable commercially available crosslinking agents include N-hydroxysuccinimide (NHS), disuccinimidyl suberate (DSS), bis(sulfosuccinimidyl) suberate (BS3), dithiobis(succinimidyl propionate) (DSP), dithiobis(sulfosuccinimidyl propionate) (DTSSP), ethylene glycol bis(succinimidyl succinate) (EGS), ethylene glycol bis(sulfosuccinimidyl succinate) (sulfo-EGS), disuccinimidyl tartrate (DST), disulfosuccinimidyl tartrate (sulfo-DST), bis[2-(succinimidooxycarbonyloxy)ethyl] sulfone (BSOCOES), and bis[2-(sulfosuccinimidooxycarbonyloxy)ethyl] sulfone (sulfo-BSOCOES).

[0153] As described above, the Fc domain disclosed herein may include a portion of the hinge region. Thus, the antigen-binding domain may be linked to the N-terminus of the Fc domain via this hinge region. In some embodiments, one or more amino acids are included between the C-terminus of the antigen-binding domain and the N-terminus of the Fc domain. In some embodiments, the one or more amino acids included between the C-terminus of the antigen-binding domain and the N-terminus of the Fc domain are amino acids of the native hinge region. In some embodiments, the C-terminus of the antigen-binding domain is fused to the N-terminus of the Fc domain via a hinge region or a portion thereof. In some embodiments, the hinge region is the IgG hinge region, such as the human IgG hinge region.

[0154] In some embodiments, the linker is a peptide linker. Examples of peptide linkers are well known, and one of ordinary skill in the art can select a suitable peptide linker for use in linking an antigen-binding domain to an FcRn-binding molecule, such as an Fc domain.

[0155] The peptide linker can be of any length. In some embodiments, the length and amino acid composition of the linker peptide sequence can be optimized to vary the orientation and / or proximity of the polypeptide domains to each other to achieve the desired activity of the FcRn / antigen-binding molecule. In some embodiments, the peptide linker is about 1 to about 100 amino acids in length, about 8 to about 40 amino acids in length, or about 15 amino acids to about 25 amino acids in length. In some embodiments, the peptide linker is 1 to 100 amino acids in length, 8 to 40 amino acids in length, or 15 to 25 amino acids in length. In some embodiments, the peptide linker is about 8 amino acids in length, about 9 amino acids in length, about 10 amino acids in length, about 11 amino acids in length, about 12 amino acids in length, about 13 amino acids in length, about 14 amino acids in length, about 15 amino acids in length, about 16 amino acids in length, about 17 amino acids in length, about 18 amino acids in length, about 19 amino acids in length, about 20 amino acids in length, about 21 amino acids in length, about 22 amino acids in length, about 23 amino acids in length, about 24 amino acids in length, about 25 amino acids in length, about 26 amino acids in length, about 27 amino acids in length, about 28 amino acids in length, about 29 amino acids in length, about 30 amino acids in length, about 31 amino acids in length, about 32 amino acids in length, about 33 amino acids in length, about 34 amino acids in length, about 35 amino acids in length, about 36 amino acids in length, about 37 amino acids in length, about 38 amino acids in length, about 39 amino acids in length, or about 40 amino acids in length. In some embodiments, the peptide linker is 8 amino acids in length, 9 amino acids in length, 10 amino acids in length, 11 amino acids in length, 12 amino acids in length, 13 amino acids in length, 14 amino acids in length, 15 amino acids in length, 16 amino acids in length, 17 amino acids in length, 18 amino acids in length, 19 amino acids in length, 20 amino acids in length, 21 amino acids in length, 22 amino acids in length, 23 amino acids in length, 24 amino acids in length, 25 amino acids in length, 26 amino acids in length, 27 amino acids in length, 28 amino acids in length, 29 amino acids in length, 30 amino acids in length, 31 amino acids in length, 32 amino acids in length, 33 amino acids in length, 34 amino acids in length, 35 amino acids in length, 36 amino acids in length, 37 amino acids in length, 38 amino acids in length, 39 amino acids in length, or 40 amino acids in length.

[0156] In some embodiments, the peptide linker contains only glycine residues and / or serine residues (e.g., glycine-serine linker or GS linker). Examples of such peptide linkers include Gly(x)Ser (where x is from 0 to 6), or Ser Gly(x) (where x is from 0 to 6), (Gly Gly Gly Gly Ser)n (where n is an integer of 1 or more), and (Ser Gly Gly Gly Gly)n (where n is an integer of 1 or more). In some embodiments, the peptide linker comprises an amino acid sequence selected from the group consisting of (GGGGS)n and (SGGGG)n, where n is from 1 to 8. In some embodiments, the linker peptide is modified such that the amino acid sequence GSG (which occurs at the junction of conventional Gly / Ser linker peptide repeats) is absent. For example, in some embodiments, the peptide linker comprises an amino acid sequence selected from the group consisting of (GGGXX)nGGGGS and GGGGS(XGGGS)n (where X is any amino acid that can be inserted into the sequence and does not result in a polypeptide containing the sequence GSG, and n is from 0 to 4). In some embodiments, the sequence of the linker peptide is (GGGX1X2)nGGGGS, where X1 is P, X2 is S, and n is from 0 to 4. In some other embodiments, the sequence of the linker peptide is (GGGX1X2)nGGGGS, where X1 is G, X2 is Q, and n is from 0 to 4. In some other embodiments, the sequence of the linker peptide is (GGGX1X2)nGGGGS, where X1 is G, X2 is A, and n is from 0 to 4. In yet other embodiments, the sequence of the linker peptide is GGGGS(XGGGS)n, where X is P, and n is from 0 to 4. In some embodiments, the linker peptide of the present disclosure comprises or consists of the amino acid sequence (GGGG A)2GGGGS. In some embodiments, the linker peptide comprises or consists of the amino acid sequence (GGGGQ)2GGGGS. In another embodiment, the linker peptide comprises or consists of the amino acid sequence (GGG P S)2GGGGS. In another embodiment, the linker peptide comprises or consists of the amino acid sequence GGGGS(P G GGS)2.In yet another embodiment, the linker peptide comprises or consists of the amino acid sequence GSGGS or SGGSGS. In some embodiments, the linker peptide comprises or consists of the amino acid sequence GGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 30), GGGGSGGGGS (SEQ ID NO: 31), or GGGGSGGGGSGGGGSGGGGSGGGGSGGGGS (SEQ ID NO: 32).

[0157] In some embodiments, the peptide linker is a GS linker that is about 20 or about 30 amino acids in length. In some embodiments, the peptide linker is a GS linker that is 20 or 30 amino acids in length.

[0158] Heavy chain molecule In some embodiments, the FcRn / antigen-binding molecule can comprise the first heavy chain described herein. In some embodiments, the first heavy chain comprises an Fc domain and an antigen-binding domain linked by a linker. In some embodiments, the FcRn / antigen-binding molecule can further comprise the second heavy chain described herein. In some embodiments, the second heavy chain comprises an Fc domain and an antigen-binding domain linked by a linker. In some embodiments, the second heavy chain comprises an Fc domain. In some embodiments, the first and second heavy chains are the same. In some embodiments, the first and second heavy chains are different.

[0159] In some embodiments, the first and second heavy chains have the same Fc domain. In some embodiments, the first and second heavy chains have different Fc domains. In some embodiments, the first heavy chain comprises an Fc domain and an antigen-binding domain, and the second heavy chain comprises an Fc but does not comprise an antigen-binding domain. In some embodiments, the first heavy chain comprises an Fc domain and an antigen-binding domain, and the second heavy chain comprises an Fc domain but does not comprise an antigen-binding domain or a linker. In some embodiments, the first heavy chain comprises an Fc domain, an antigen-binding domain, and a linker, and the second heavy chain comprises an Fc domain but does not comprise an antigen-binding domain or a linker.

[0160] In some embodiments, the antigen-binding domain is linked to the N-terminus of the Fc domain. In some embodiments, the antigen-binding domain is linked to the C-terminus of the Fc domain. In some embodiments, the antigen-binding domain is linked to a position other than the N-terminus or C-terminus of the Fc domain.

[0161] In some embodiments, the antigen-binding domain is fused to the N-terminus of the Fc domain. In some embodiments, the antigen-binding domain is fused to the C-terminus of the Fc domain. In some embodiments, the antigen-binding domain is fused to a position other than the N-terminus or C-terminus of the Fc domain.

[0162] In some embodiments, the antigen-binding domain is linked to the N-terminus of the Fc domain by a linker. In some embodiments, the antigen-binding domain is linked to the C-terminus of the Fc domain by a linker. In some embodiments, the antigen-binding domain is linked to a position other than the N-terminus or C-terminus of the Fc domain by a linker.

[0163] In some embodiments, the antigen-binding domain is fused to the N-terminus of the Fc domain by a peptide linker. In some embodiments, the antigen-binding domain is fused to the C-terminus of the Fc domain by a peptide linker. In some embodiments, the antigen-binding domain is fused to a position other than the N-terminus or C-terminus of the Fc domain by a peptide linker.

[0164] In some embodiments, the Fc domain comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 1-9. In some embodiments, the Fc domain consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 1-9.

[0165] In some embodiments, the Fc domain comprises an amino acid sequence of any one of SEQ ID NOs: 1-9. In some embodiments, the Fc domain consists of an amino acid sequence of any one of SEQ ID NOs: 1-9.

[0166] In some embodiments, the first and second heavy chains comprise the same Fc domain. In some embodiments, both the first and second heavy chains comprise an Fc domain that comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence of any one of SEQ ID NOs: 1-3. In some embodiments, both the first and second heavy chains comprise an Fc domain that consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to an amino acid sequence of any one of SEQ ID NOs: 1-3. In some embodiments, both the first and second heavy chains comprise an Fc domain that comprises an amino acid sequence of any one of SEQ ID NOs: 1-3. In some embodiments, both the first and second heavy chains comprise an Fc domain that consists of an amino acid sequence of any one of SEQ ID NOs: 1-3.

[0167] In some embodiments, the first and second heavy chains comprise different Fc domains. In some embodiments, the first heavy chain comprises an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 4-6, and the second heavy chain comprises an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 7-9. In some embodiments, the first heavy chain comprises an Fc domain consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 4-6, and the second heavy chain comprises an Fc domain consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 7-9. In some embodiments, the first heavy chain comprises an Fc domain comprising an amino acid sequence of any one of SEQ ID NOs: 4-6, and the second heavy chain comprises an Fc domain comprising an amino acid sequence of any one of SEQ ID NOs: 7-9. In some embodiments, when the first heavy chain comprises an Fc domain comprising the amino acid sequence of SEQ ID NO: 4 or a variant thereof, the second heavy chain comprises an Fc domain comprising SEQ ID NO: 7 or a variant thereof. In some embodiments, when the first heavy chain comprises an Fc domain comprising the amino acid sequence of SEQ ID NO: 5 or a variant thereof, the second heavy chain comprises an Fc domain comprising SEQ ID NO: 8 or a variant thereof. In some embodiments, when the first heavy chain comprises an Fc domain comprising the amino acid sequence of SEQ ID NO: 6 or a variant thereof, the second heavy chain comprises an Fc domain comprising SEQ ID NO: 9 or a variant thereof.

[0168] In some embodiments, the first and second heavy chains further comprise a peptide linker. In some embodiments, the first and second heavy chains further comprise the same peptide linker. In some embodiments, the first and second heavy chains further comprise different peptide linkers. In some embodiments, the first heavy chain comprises an Fc domain, a peptide linker, and an antigen-binding domain, and the second heavy chain comprises an Fc domain but does not comprise a peptide linker or an antigen-binding domain. The peptide linker encoded by the first and heavy chains can be any of those described herein. In some embodiments, the linker comprises the amino acid sequence of SEQ ID NO: 30, 31, or 32.

[0169] In some embodiments, the FcRn / antigen-binding molecule comprises, or consists of, an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 34. In some embodiments, the FcRn / antigen-binding molecule comprises, or consists of, the amino acid sequence of SEQ ID NO: 34.

[0170] In some embodiments, the FcRn / antigen-binding molecule comprises the amino acid sequence of SEQ ID NO: 34 or a variant thereof and one or more amino acids added at the C-terminus. In some embodiments, the FcRn / antigen-binding molecule comprises the amino acid sequence of SEQ ID NO: 34 or a variant thereof and one or more amino acids added at the C-terminus, and the one or more amino acids are selected from A, AG, GG, and PP.

[0171] In some embodiments, the first heavy chain of the FcRn / antigen-binding molecule comprises or consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 34, and the second heavy chain of the FcRn / antigen-binding molecule does not contain an antigen-binding domain. In some embodiments, the second heavy chain of the FcRn / antigen-binding molecule contains an Fc domain but does not contain an antigen-binding domain. Optionally, the second heavy chain of the FcRn / antigen-binding molecule comprises or consists of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 8.

[0172] In some embodiments, the first heavy chain of the FcRn / antigen-binding molecule comprises or consists of the amino acid sequence of SEQ ID NO: 34, and the second heavy chain of the FcRn / antigen-binding molecule does not contain an antigen-binding domain. In some embodiments, the second heavy chain of the FcRn / antigen-binding molecule contains an Fc domain but does not contain an antigen-binding domain. Optionally, the second heavy chain of the FcRn / antigen-binding molecule comprises or consists of the amino acid sequence of SEQ ID NO: 8.

[0173] Polynucleotide, vector, and production method The present disclosure also provides polynucleotides encoding the FcRn / antigen-binding molecules or fragments thereof disclosed herein. In some embodiments, the polynucleotide encodes an antigen-binding domain of the present disclosure. In some embodiments, the polynucleotide encodes an FcRn-binding molecule of the present disclosure. In some embodiments, the polynucleotide encodes an Fc region of the present disclosure. In some embodiments, the polynucleotide encodes an Fc domain of the present disclosure. In some embodiments, the polynucleotide encodes one or more of an antigen-binding domain, an FcRn-binding molecule, and a linker. In some embodiments, the polynucleotide encodes an antigen-binding domain and an FcRn-binding molecule, and optionally encodes a linker. In some embodiments, the polynucleotide encodes one or more of an antigen-binding domain, an Fc region, and a linker. In some embodiments, the polynucleotide encodes an antigen-binding domain and an Fc region, and optionally encodes a linker. In some embodiments, the polynucleotide encodes an FcRn / antigen-binding molecule comprising one antigen-binding domain and an Fc region. In some embodiments, the polynucleotide encodes one or more of an antigen-binding domain, an Fc domain, and a linker. In some embodiments, the polynucleotide encodes an antigen-binding domain and an Fc domain, and optionally encodes a linker. In some embodiments, the polynucleotide encodes an FcRn / antigen-binding molecule comprising one antigen-binding domain and one or more Fc domains. In some embodiments, the polynucleotide encodes one or more heavy chains of the present disclosure.

[0174] As used herein, an "isolated" polynucleotide or nucleic acid molecule is separated from other nucleic acid molecules that are present in a natural source of the nucleic acid molecule (e.g., in a mouse or human). Also, an "isolated" nucleic acid molecule, e.g., a cDNA molecule, may be substantially free of other cellular material or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemicals when chemically synthesized. For example, the phrase "substantially free of" means that a preparation of a polynucleotide or nucleic acid molecule has less than about 15%, 10%, 5%, 2%, 1%, 0.5%, or 0.1% (particularly less than about 10%) of other materials, such as cellular material, culture medium, other nucleic acid molecules, chemical precursors, and / or other chemicals. In one embodiment, a nucleic acid molecule(s) encoding a polypeptide described herein is isolated or purified.

[0175] In one aspect, a polynucleotide is provided herein that comprises a nucleotide sequence encoding an FcRn binding molecule or an FcRn / antigen binding molecule described herein. In another aspect, a polynucleotide is provided herein that comprises a nucleotide sequence encoding an antigen binding domain described herein. In another aspect, a polynucleotide is provided herein that comprises a nucleotide sequence encoding an FcRn / antigen binding molecule described herein. In another aspect, a polynucleotide is provided herein that comprises a nucleotide sequence encoding an FcRn binding molecule or an FcRn / antigen binding molecule described herein.

[0176] The polynucleotide can comprise a nucleotide sequence encoding an sdAb (e.g., a VHH fragment), a Fab fragment, a scFv, a VH, or a VL that includes the FRs and CDRs of the antigen-binding domain described herein. The polynucleotide can also comprise a nucleotide sequence encoding an antibody mimetic described herein. In some embodiments, the polynucleotide can comprise a nucleotide sequence encoding a VHH fragment that includes the FRs and CDRs of the antigen-binding domain described herein. In some embodiments, the polynucleotide can comprise a nucleotide sequence encoding a light chain that includes the VL FRs and CDRs of the antigen-binding domain described herein, or a nucleotide sequence encoding a heavy chain that includes the VH FRs and CDRs of the antigen-binding domain and / or the Fc domain described herein. In one embodiment, the polynucleotide encodes a VH, a VL, a heavy chain, and / or a light chain of the antigen-binding domain described herein. In one embodiment, the polynucleotide encodes a first VH and a first VL of the antigen-binding domain described herein. In one embodiment, the polynucleotide encodes a second VH and a second VL of the antigen-binding domain described herein. In one embodiment, the polynucleotide encodes a first heavy chain and a first light chain of the antigen-binding domain described herein. In one embodiment, the polynucleotide encodes a second heavy chain and a second light chain of the antigen-binding domain described herein. In one embodiment, the polynucleotide encodes a VH and / or a VL, or a heavy chain and / or a light chain of the antigen-binding domain described herein.

[0177] In some embodiments, the polynucleotide can comprise a nucleotide sequence encoding the first heavy chain described herein. In some embodiments, the first heavy chain comprises an Fc domain and an antigen-binding domain linked by a linker. In some embodiments, the polynucleotide can comprise a nucleotide sequence encoding the second heavy chain described herein. In some embodiments, the second heavy chain comprises an Fc domain and an antigen-binding domain linked by a linker. In some embodiments, the first and second heavy chains are the same. In some embodiments, the first and second heavy chains are different.

[0178] In some embodiments, the first and second heavy chains have the same Fc domain. In some embodiments, the first and second heavy chains have different Fc domains. In some embodiments, the second heavy chain comprises an Fc domain but does not comprise an antigen-binding domain, and the first heavy chain comprises an Fc domain and an antigen-binding domain. In some embodiments, the second heavy chain comprises an Fc domain but does not comprise an antigen-binding domain or a linker, and the first heavy chain comprises an Fc domain and an antigen-binding domain. In some embodiments, the second heavy chain comprises an Fc domain but does not comprise an antigen-binding domain or a linker, and the first heavy chain comprises an Fc domain, an antigen-binding domain, and a linker.

[0179] In some embodiments, the polynucleotide comprises a nucleotide sequence encoding an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 1-9. In some embodiments, the polynucleotide consists of a nucleotide sequence encoding an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 1-9.

[0180] In some embodiments, the polynucleotide comprises a nucleotide sequence encoding an Fc domain comprising any one of the amino acid sequences of SEQ ID NOs: 1-9. In some embodiments, the polynucleotide comprises a nucleotide sequence encoding an Fc domain consisting of any one of the amino acid sequences of SEQ ID NOs: 1-9.

[0181] In some embodiments, the polynucleotide comprises a nucleotide sequence encoding two or more Fc domains. In some embodiments, the polynucleotide comprises a nucleotide sequence encoding two Fc domains. In some embodiments, the polynucleotide comprises a first nucleotide sequence encoding a first Fc domain and a second nucleotide sequence encoding a second Fc domain. In some embodiments, the first nucleotide sequence and the second nucleotide sequence are contained within different nucleic acid molecules. In some embodiments, the first nucleotide sequence and the second nucleotide sequence are contained within the same nucleic acid molecule.

[0182] In some embodiments, the first and second nucleotide sequences encode the same Fc domain. In some embodiments, both the first and second nucleotide sequences encode an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 1-3. In some embodiments, both the first and second nucleotide sequences encode an Fc domain consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 1-3. In some embodiments, both the first and second nucleotide sequences encode an Fc domain comprising any one of the amino acid sequences of SEQ ID NOs: 1-3. In some embodiments, both the first and second nucleotide sequences encode an Fc domain consisting of any one of the amino acid sequences of SEQ ID NOs: 1-3.

[0183] In some embodiments, the first and second nucleotide sequences encode different Fc domains. In some embodiments, the first nucleotide sequence encodes an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 4-6, and the second nucleotide sequence encodes an Fc domain comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 7-9. In some embodiments, the first nucleotide sequence encodes an Fc domain consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 4-6, and the second nucleotide sequence encodes an Fc domain consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of the amino acid sequences of SEQ ID NOs: 7-9. In some embodiments, the first nucleotide sequence encodes an Fc domain comprising any one of the amino acid sequences of SEQ ID NOs: 4-6, and the second nucleotide sequence encodes an Fc domain comprising any one of the amino acid sequences of SEQ ID NOs: 7-9. In some embodiments, the first nucleotide sequence encodes an Fc domain consisting of any one of the amino acid sequences of SEQ ID NOs: 4-6, and the second nucleotide sequence encodes an Fc domain consisting of any one of the amino acid sequences of SEQ ID NOs: 7-9. In some embodiments, when the first nucleotide sequence encodes the amino acid sequence of SEQ ID NO: 4 or a variant thereof, the second nucleotide sequence encodes SEQ ID NO: 7 or a variant thereof. In some embodiments, when the first nucleotide sequence encodes the amino acid sequence of SEQ ID NO: 5 or a variant thereof, the second nucleotide sequence encodes SEQ ID NO: 8 or a variant thereof.In some embodiments, when the first nucleotide sequence encodes the amino acid sequence of SEQ ID NO: 6 or a variant thereof, the second nucleotide sequence encodes SEQ ID NO: 9 or a variant thereof.

[0184] In some embodiments, the first nucleotide sequence encodes an Fc domain and an antigen-binding domain, and the second nucleotide sequence encodes an Fc domain but does not encode an antigen-binding domain. The antigen-binding domain encoded by the first and / or second nucleotide sequence can be any of those described herein.

[0185] In some embodiments, the first and second nucleotide sequences also encode a peptide linker. In some embodiments, the first and second nucleotide sequences encode the same peptide linker. In some embodiments, the first and second nucleotide sequences encode different peptide linkers. In some embodiments, the first nucleotide sequence encodes an Fc domain, a peptide linker, and an antigen-binding domain, and the second nucleotide sequence encodes an Fc domain but does not encode a peptide linker or an antigen-binding domain. In some embodiments, the first nucleotide sequence encodes an antigen-binding domain, a peptide linker, and an Fc domain, and the second nucleotide sequence encodes an Fc domain but does not encode a peptide linker or an antigen-binding domain. The peptide linker encoded by the first and / or second nucleotide sequence can be any of those described herein. In some embodiments, the linker comprises the amino acid sequence of SEQ ID NO: 30, 31, or 32.

[0186] In some embodiments, the polynucleotide comprises, or consists of, a nucleotide sequence encoding a protein that comprises an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 34.

[0187] In some embodiments, the polynucleotide comprises a nucleotide sequence encoding a protein comprising or consisting of the amino acid sequence of SEQ ID NO: 34.

[0188] In some embodiments, the first nucleotide sequence encodes a protein comprising or consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 34, and the second nucleotide sequence encodes an Fc domain but does not encode an antigen-binding domain. Optionally, the second nucleotide sequence encodes a protein comprising an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 8.

[0189] In some embodiments, the first nucleotide sequence encodes a protein comprising the amino acid sequence of SEQ ID NO: 34, and the second nucleotide sequence encodes an Fc domain but does not encode an antigen-binding domain. Optionally, the second nucleotide sequence encodes a protein comprising the amino acid sequence of SEQ ID NO: 8. In some embodiments, the first nucleotide sequence comprises a nucleotide sequence encoding a protein consisting of the amino acid sequence of SEQ ID NO: 34, and the second nucleotide sequence encodes an Fc domain but does not encode an antigen-binding domain. Optionally, the second nucleotide sequence encodes a protein consisting of the amino acid sequence of SEQ ID NO: 8.

[0190] In some embodiments, a third nucleotide sequence encoding a light chain is provided. In some embodiments, the third nucleotide sequence encodes a protein comprising or consisting of an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to the amino acid sequence of SEQ ID NO: 24. In some embodiments, the third nucleotide sequence encodes a protein comprising or consisting of the amino acid sequence of SEQ ID NO: 24.

[0191] Also provided herein are polynucleotides encoding the polypeptides provided above, which are optimized, for example, by codon / RNA optimization, replacement with heterologous signal sequences, and elimination of mRNA instability elements. Thus, methods for generating optimized nucleic acids for recombinant expression by introduction of codon changes and / or elimination of inhibitory regions in mRNA can be carried out, for example, by adapting the optimization methods described in U.S. Patent Nos. 5,965,726, 6,174,666, 6,291,664, 6,414,132, and 6,794,498, all of which are hereby incorporated by reference in their entirety. For example, potential splice sites and instability elements (e.g., A / T or A / U rich elements) within the RNA can be mutated without changing the amino acids encoded by the nucleic acid sequence to increase the stability of the RNA for recombinant expression. The changes use the degeneracy of the genetic code and, for example, alternative codons for the same amino acid are used. In one embodiment, it may be desirable to change one or more codons to encode conservative mutations, such as similar amino acids having a similar chemical structure and properties and / or function as the original amino acid.

[0192] A polynucleotide can be obtained by any method known in the art, and the nucleotide sequence of the polynucleotide can be determined by any method known in the art. The nucleotide sequences encoding the proteins described herein, and modified versions of these antibodies, can be determined using methods well known in the art, i.e., nucleotide codons known to encode specific amino acids are assembled to generate a nucleic acid encoding the protein. Such polynucleotides encoding a protein can be assembled from chemically synthesized oligonucleotides (e.g., as described in Kutmeier G et al., (1994) BioTechniques 17:242-6, which is incorporated herein by reference in its entirety), briefly, this involves the synthesis of overlapping oligonucleotides containing portions of the sequence encoding the antibody, the annealing and ligation of these oligonucleotides, and then the amplification of the ligated oligonucleotides by PCR.

[0193] Alternatively, the polynucleotides encoding the proteins described herein can be generated from nucleic acids from a suitable source (e.g., hybridoma) using methods well known in the art (e.g., PCR and other molecular cloning methods). For example, PCR amplification using synthetic primers that hybridize to the 3' and 5' ends of a known sequence can be performed using genomic DNA obtained from hybridoma cells that produce the polypeptide of interest. Such PCR amplification methods can be used to obtain a nucleic acid containing the sequence encoding the polypeptide. The amplified nucleic acid can be cloned into a vector for expression in a host cell and further cloning.

[0194] Although clones containing nucleic acids encoding a specific polypeptide are not available, if the sequence of the polypeptide is known, the nucleic acid encoding the polypeptide can be obtained from a suitable source (e.g., a cDNA library generated from any tissue or cell expressing the polypeptide described herein, or a nucleic acid isolated therefrom, preferably polyA+ RNA), by PCR amplification using synthetic primers capable of hybridizing to the 3' and 5' ends of the sequence, or by using an oligonucleotide probe specific for a particular gene sequence, for example, by cloning to identify a cDNA clone from a cDNA library encoding the polypeptide. The amplified nucleic acid generated by PCR can then be cloned into a replicable cloning vector using any method well known in the art.

[0195] The DNA encoding the proteins described herein can be readily isolated and sequenced using conventional procedures. Hybridoma cells can function as a source of such DNA. When isolated, the DNA can be placed into an expression vector, which can then be transfected into a host cell, such as E. coli cells, monkey COS cells, Chinese hamster ovary (CHO) cells (e.g., CHO cells from the CHO GS System™ (Lonza)), or otherwise myeloma cells that do not produce the proteins described herein.

[0196] Also provided are polynucleotides that hybridize to a polynucleotide encoding a protein described herein under high stringency, intermediate, or low stringency hybridization conditions.

[0197] Hybridization conditions are described in the art and are known to those skilled in the art. For example, hybridization under stringent conditions can include hybridization to filter-bound DNA in 6x sodium chloride / sodium citrate (SSC) at about 45°C, followed by one or more washes in 0.2x SSC / 0.1% SDS at about 50 - 65°C, and hybridization under highly stringent conditions can include hybridization to filter-bound nucleic acid in 6x SSC at about 45°C, followed by one or more washes in 0.1x SSC / 0.2% SDS at about 68°C. Hybridization under other stringent hybridization conditions is known to and described by those skilled in the art. See, for example, Ausubel FM et al., eds., (1989) Current Protocols in Molecular Biology, Vol. I, Green Publishing Associates, Inc. and John Wiley & Sons, Inc., New York, pages 6.3.1 - 6.3.6 and 2.10.3, which is hereby incorporated by reference in its entirety.

[0198] In one aspect, provided herein are cells (e.g., host cells) that express (e.g., recombinantly express) the proteins described herein, as well as related polynucleotides and expression vectors. Also provided herein are vectors (e.g., expression vectors) for recombinant expression in a host cell, preferably a mammalian cell (e.g., a CHO cell), that contain a polynucleotide that includes a nucleotide sequence encoding a protein described herein. Also provided herein are host cells for recombinant expression of a protein described herein that contain such a vector. In one aspect, provided herein is a method for producing a protein described herein, the method including expressing a polypeptide from a host cell.

[0199] The recombinant expression of the proteins described herein generally involves the construction of an expression vector containing a polynucleotide encoding a polypeptide. When a polynucleotide encoding a polypeptide described herein is obtained, a vector for the production of the polypeptide can be produced by recombinant DNA techniques using techniques well known in the art. Accordingly, methods for preparing a protein by expressing a polynucleotide containing a polypeptide encoding a nucleotide sequence are described herein. Methods well known to those skilled in the art can be used to construct an expression vector containing a polypeptide coding sequence and appropriate transcriptional and translational control signals. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo gene recombination. Also provided is a replicable vector comprising a nucleotide sequence encoding a polypeptide described herein, wherein the nucleotide sequence is operably linked to a promoter. Such a vector can include, for example, a nucleotide sequence encoding the first heavy chain of the present disclosure (see, for example, International Publication Nos. 86 / 05807 and 89 / 01036, and U.S. Patent No. 5,122,464, which are hereby incorporated by reference in their entirety), and the second heavy chain of the present disclosure can be cloned into such a vector for the expression of the first heavy chain, the second heavy chain, or both the first and second heavy chains.

[0200] In one embodiment, the vector comprises a polynucleotide encoding an sdAb, Fab fragment, scFv, VHH fragment, VH, VL, heavy chain, and / or light chain of a polypeptide described herein. In another embodiment, the vector comprises a polynucleotide encoding VH and VL of a polypeptide described herein. In another embodiment, the vector comprises a polynucleotide encoding the heavy chain and light chain of a polypeptide described herein.

[0201] An expression vector can be transferred into a cell (e.g., a host cell) by conventional techniques, and then the obtained cell can be cultured by conventional techniques to produce the polypeptide or a fragment thereof described herein. Accordingly, provided herein are host cells containing a polynucleotide encoding the polypeptide or a fragment thereof, or a heavy or light chain thereof, or a fragment thereof, or a single-chain antibody described herein, wherein the polynucleotide is operably linked to a promoter for expression of such a sequence in the host cell.

[0202] In one embodiment, the host cell comprises a polynucleotide comprising one of the above first nucleotide sequences and one of the above second nucleotide sequences. In another embodiment, the host cell comprises a first polynucleotide comprising one of the above first nucleotide sequences and a second polynucleotide comprising one of the above first nucleotide sequences. In another embodiment, the host cell comprises a first vector comprising one of the above first nucleotide sequences and one of the above second nucleotide sequences. In another embodiment, the host cell comprises a first vector comprising one of the above first nucleotide sequences and one of the above second nucleotide sequences, and a second vector comprising a second polynucleotide comprising one of the above first nucleotide sequences.

[0203] In some embodiments, the FcRn / antigen-binding molecule expressed by the first host cell associates with the FcRn / antigen-binding molecule expressed by the second host cell to form a two-armed FcRn / antigen-binding molecule. In some embodiments, the FcRn / antigen-binding molecule expressed by the first host cell associates with the FcRn-binding molecule expressed by the second host cell to form a one-armed FcRn / antigen-binding molecule. In some embodiments, provided herein is a population of host cells comprising such a first host cell and such a second host cell.

[0204] In some embodiments, a population of vectors is provided herein, comprising a first vector comprising a polynucleotide encoding an FcRn / antigen-binding molecule and a second vector comprising a polynucleotide encoding an FcRn / antigen-binding molecule. In some embodiments, a population of vectors is provided herein, comprising a first vector comprising a polynucleotide encoding an FcRn / antigen-binding molecule and a second vector comprising a polynucleotide encoding an FcRn-binding molecule. In some embodiments, a population of vectors is provided herein, comprising a first vector comprising a polynucleotide encoding an FcRn / antigen-binding molecule and a polynucleotide encoding an FcRn / antigen-binding molecule. In some embodiments, a population of vectors is provided herein, comprising a first vector comprising a polynucleotide encoding two FcRn / antigen-binding molecules.

[0205] A variety of host expression vector systems can be used to express the polypeptides described herein (see, e.g., U.S. Patent No. 5,807,715, which is incorporated herein by reference in its entirety). Such host expression systems represent a medium in which the desired coding sequence can be produced and then purified, and also represent cells that can express the polypeptides described herein in situ when transformed or transfected with the appropriate nucleotide coding sequence. These include microorganisms such as bacteria (e.g., E. coli and B. subtilis), bacteria transformed with recombinant bacteriophage DNA, plasmid DNA, or cosmid DNA expression vectors containing, for example, an FcRn / antigen-binding molecule coding sequence; yeasts (e.g., Saccharomyces and Pichia), yeasts transformed with recombinant yeast expression vectors containing, for example, an FcRn / antigen-binding molecule coding sequence; insect cell lines, such as insect cell lines infected with recombinant virus expression vectors (e.g., baculovirus) containing an FcRn / antigen-binding molecule coding sequence; plant cell lines (e.g., green algae, e.g., Chlamydomonas reinhardtii), plant cell lines infected with recombinant virus expression vectors (e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus, TMV) containing, for example, an FcRn / antigen-binding molecule coding sequence, or plant cell lines transformed with recombinant plasmid expression vectors (e.g., Ti plasmid) containing, for example, an FcRn / antigen-binding molecule coding sequence; or mammalian cell lines (e.g., COS (e.g., COS1 or COS), CHO, BHK, MDCK, HEK 293, NS0, PER.C6, VERO, CRL7O3O, HsS78Bst, HeLa, NIH 3T3, HEK-293T, HepG2, SP210, R1.1, B-W, L-M, BSC1, BSC40, YB / 20, and BMT10 cells), including, but not limited to, mammalian cell lines carrying recombinant expression constructs containing a promoter derived from the genome of a mammalian cell (e.g., the metallothionein promoter) or a promoter derived from a mammalian virus (e.g., the adenovirus late promoter; the vaccinia virus 7.5K promoter).In one embodiment, the cells for expressing the FcRn / antigen-binding molecule described herein are Chinese hamster ovary (CHO) cells, e.g., CHO cells from the CHO GS System™ (Lonza). In one embodiment, the heavy and / or light chains produced by CHO cells may have N-terminal glutamine or glutamate residues replaced by pyroglutamic acid. In one embodiment, the cells for expressing the polypeptides described herein are human cells, e.g., human cell lines. In one embodiment, the mammalian expression vector is pOptiVEC™ or pcDNA3.3. In one embodiment, bacterial cells, e.g., Escherichia coli, or eukaryotic cells (e.g., mammalian cells) are used for the expression of recombinant polypeptides. For example, mammalian cells, e.g., CHO cells, are an effective expression system for antibodies together with a vector, e.g., the major immediate early gene promoter element from human cytomegalovirus (Foecking MK & Hofstetter H (1986) Gene 45:101-5, and Cockett MI et al., (1990) Biotechnology 8(7):662-7, each of these references is incorporated herein by reference in its entirety). In one embodiment, the polypeptides described herein are produced by CHO cells or NS0 cells. In one embodiment, the expression of the nucleotide sequence encoding the polypeptides described herein that contain two, three, or four binding sites to human FcRn is regulated by a constitutive promoter, an inducible promoter, or a tissue-specific promoter.

[0206] In the bacterial system, several expression vectors can be advantageously selected depending on their intended use for the expression of molecules. For example, when large amounts of such polypeptides are produced for the generation of pharmaceutical compositions of antibody molecules, a vector that directs the expression of a high level of fusion protein product that is easily purified may be desirable. Such vectors include, but are not limited to, the E. coli expression vector pUR278 (Ruether U & Mueller-Hill B (1983) EMBO J 2:1791-1794), in which the coding sequence can be individually ligated into the vector in-frame with the lac Z coding region, whereby a fusion protein is produced, pUR278; and pIN vectors (Inouye S & Inouye M (1985) Nuc Acids Res 13:3101-3109, Van Heeke G & Schuster SM (1989) J Biol Chem 24:5503-5509), all of these references are incorporated herein by reference in their entirety. Also, for example, the pGEX vector can be used to express a foreign polypeptide as a fusion protein with glutathione S-transferase (GST). Generally, such fusion proteins are soluble and can be easily purified from lysed cells by adsorption and binding to matrix glutathione agarose beads, followed by elution in the presence of free glutathione. The pGEX vector is designed to contain a thrombin or factor Xa protease cleavage site, whereby the cloned target gene product can be released from the GST moiety.

[0207] In the insect system, for example, the Autographa californica nuclear polyhedrosis virus (AcNPV) can be used as a vector to express foreign genes. The virus replicates in Spodoptera frugiperda cells. The coding sequences can be individually cloned into non-essential regions of the virus (e.g., the polyhedrin gene) and placed under the control of an AcNPV promoter (e.g., the polyhedrin promoter).

[0208] In mammalian host cells, several virus-based expression systems can be used. When adenovirus is used as an expression vector, the coding sequence of interest can be ligated to an adenovirus transcription / translation control complex, such as a late promoter and a tripartite leader sequence. This chimeric gene can then be inserted into the adenovirus genome by in vitro or in vivo recombination. Insertion into a non-essential region of the viral genome (e.g., region E1 or E3) results in a recombinant virus that is viable and capable of expressing the molecule in the infected host (see, for example, Logan J & Shenk T (1984) PNAS 81(12):3655-9, which is incorporated herein by reference in its entirety). Also, specific initiation signals may be required for efficient translation of the inserted coding sequence. These signals include the ATG initiation codon and adjacent sequences. Furthermore, the initiation codon must be in frame with the reading frame of the desired coding sequence to ensure translation of the entire insert. These exogenous translation control signals and initiation codons can be of various origins, both natural and synthetic. The efficiency of expression can be enhanced by including appropriate transcriptional enhancer elements, transcriptional terminators, etc. (see, for example, Bitter G et al., (1987) Methods Enzymol. 153:516-544, which is incorporated herein by reference in its entirety).

[0209] In addition, a host cell line can be selected that modulates the expression of the inserted array or modifies and processes the gene product in a desired specific manner. Such modifications (e.g., glycosylation) and processing (e.g., cleavage) of the protein product can be important for the function of the protein. Different host cells have characteristic and specific mechanisms for the post-translational processing and modification of proteins and gene products. An appropriate cell line or host system can be selected to ensure the correct modification and processing of the foreign protein being expressed. For this purpose, eukaryotic host cells having cell machinery for the appropriate processing of primary transcripts, glycosylation, and phosphorylation of gene products can be used. Such mammalian host cells include, but are not limited to, CHO, VERO, BHK, HeLa, MDCK, HEK 293, NIH 3T3, W138, BT483, Hs578T, HTB2, BT2O, and T47D, NS0 (a mouse myeloma cell line that does not endogenously produce any immunoglobulin chains), CRL7O3O, COS (e.g., COS1 or COS), PER.C6, VERO, HsS78Bst, HEK-293T, HepG2, SP210, R1.1, B-W, L-M, BSC1, BSC40, YB / 20, BMT10, and HsS78Bst cells. In one embodiment, the proteins described herein are produced in mammalian cells, e.g., CHO cells.

[0210] In one embodiment, the polypeptides described herein comprise a portion of an antibody having a reduced fucose content or no fucose content. Such proteins can be produced using techniques known to those of skill in the art. For example, the protein can be expressed in cells that are deficient or lacking in the ability to fucosylate. In one example, a cell line having knockout of both alleles of α1,6-fucosyltransferase can be used to produce an antibody having a reduced fucose content. The Potelligent® system (Lonza) is an example of such a system that can be used to produce an antibody having a reduced fucose content.

[0211] Stable expressing cells can be generated for the long-term high-yield production of recombinant proteins. For example, cell lines that stably express the proteins described herein can be manipulated. In one embodiment, the cells provided herein stably express an antigen-binding domain, an FcRn / antigen-binding molecule, or an FcRn-binding molecule, and the antigen-binding domain, FcRn / antigen-binding molecule, or FcRn-binding molecule associate to form the one-arm polypeptides described herein.

[0212] In certain embodiments, rather than using an expression vector containing a viral origin of replication, the host cells can be transformed with DNA controlled by appropriate expression control elements (e.g., promoters, enhancers, sequences, transcription terminators, polyadenylation sites, etc.) and selectable markers. After introduction of the foreign DNA / polynucleotide, the engineered cells can be allowed to grow in enriched medium for 1-2 days and then switched to selective medium. The selectable marker in the recombinant plasmid confers resistance to the selection, allowing the cells to stably integrate the plasmid into their chromosomes, grow, and form foci, which can then be cloned and grown into cell lines. This method can advantageously be used to engineer cell lines that express polypeptides containing 2, 3, or 4 binding sites to the human FcRn or fragments thereof described herein. Such engineered cell lines can be particularly useful in screening and evaluating compositions that interact directly or indirectly with the polypeptide.

[0213] Several selection systems can be used, and the selection systems each include, but are not limited to, the herpes simplex virus thymidine kinase (Wigler M et al., (1977) Cell 11(1):223-32), hypoxanthine-guanine phosphoribosyltransferase (Szybalska EH & Szybalski W (1962) PNAS 48(12):2026-2034), and adenine phosphoribosyltransferase (Lowy I et al., (1980) Cell 22(3):817-23) genes in tk, hgprt, or aprt cells, and all of these references are incorporated herein by reference in their entirety. Also, metabolic antagonist resistance can be used as the basis for selection for the following genes, dhfr that confers resistance to methotrexate (Wigler M et al., (1980) PNAS 77(6):3567-70, O’Hare K et al., (1981) PNAS 78:1527-31); gpt that confers resistance to mycophenolic acid (Mulligan RC & Berg P (1981) PNAS 78(4):2072-6); neo that confers resistance to aminoglycoside G-418 (Wu GY & Wu CH (1991) Biotherapy 3:87-95, Tolstoshev P (1993) Ann Rev Pharmacol Toxicol 32:573-596, Mulligan RC (1993) Science 260:926-932, and Morgan RA & Anderson WF (1993) Ann Rev Biochem 62:191-217, Nabel GJ & Felgner PL (1993) Trends Biotechnol 11(5):211-5); and hygro that confers resistance to hygromycin (Santerre RF et al., (1984) Gene 30(1-3):147-56), and all of these references are incorporated herein by reference in their entirety.Methods generally known in the art of recombinant DNA technology are commonly applied to select the desired recombinant clone, such methods are described, for example, in Ausubel FM et al., (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993), Kriegler M, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990), and Chapters 12 and 13, Dracopoli NC et al., (eds.), Current Protocols in Human Genetics, John Wiley & Sons, NY (1994), Colbere-Garapin F et al., (1981) J Mol Biol 150:1-14, and all of these references are hereby incorporated by reference in their entirety.

[0214] The expression level of the polypeptide can be increased by vector amplification (for a review, see Bebbington CR & Hentschel CCG, The use of vectors based on gene amplification for the expression of cloned genes in mammalian cells in DNA cloning, p.163-188. DNA Cloning, Vol III, A Practical Approach. D.M. Glover (ed.) (Academic Press, New York, 1987), which reference is hereby incorporated by reference in its entirety). When the marker in the vector system is amplifiable, an increase in the level of inhibitor present in the culture of host cells increases the number of copies of the marker gene. Also, since the amplified region is related to the gene of interest, the production of the polypeptide increases (Crouse GF et al., (1983) Mol Cell Biol 3:257-66, which reference is hereby incorporated by reference in its entirety).

[0215] The host cell can be co-transfected with two or more expression vectors described herein. The two vectors can contain the same selectable marker, and the same selectable marker enables equal expression of polypeptides, such as the first heavy chain and the second heavy chain polypeptides. The host cell can be co-transfected with different amounts of two or more expression vectors. For example, the host cell can be transfected with any one of the following ratios of the first expression vector and the second expression vector: about 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:12, 1:15, 1:20, 1:25, 1:30, 1:35, 1:40, 1:45, or 1:50.

[0216] Alternatively, a single vector that can encode and express both polypeptides can be used. The coding sequence can include cDNA or genomic DNA. The expression vector can be monocistronic or polycistronic. The polycistronic nucleic acid construct can encode in the range of 2, 3, 4, 5, 6, 7, 8, 9, 10, or more genes / nucleotide sequences, or in the range of 2 - 5, 5 - 10, or 10 - 20 genes / nucleotide sequences. For example, a bicistronic nucleic acid construct can include a promoter, a first gene, and a second gene in the following order. In such an expression vector, the transcription of both genes can be driven by the promoter, the translation of mRNA from the first gene can be by a cap-dependent scanning mechanism, and the translation of mRNA from the second gene can be by a cap-independent mechanism, such as an IRES.

[0217] The polypeptides described herein, when produced by recombinant expression, can be purified by any method known in the art for protein purification, such as chromatography (e.g., ion exchange, affinity, particularly affinity for a specific antigen after Protein A, and sizing column chromatography), centrifugation, differential solubility, or any other standard technique for protein purification. Further, the polypeptides described herein can be fused to the heterologous polypeptide sequences described herein, or otherwise heterologous polypeptide sequences known in the art, to facilitate purification.

[0218] In one embodiment, the polypeptides described herein are isolated or purified. In one embodiment, an isolated polypeptide is substantially free of other polypeptides having antigenic specificities different from that of the isolated polypeptide. For example, in certain embodiments, preparations of the proteins described herein are substantially free of cellular material and / or chemical precursors. The expression "substantially free of cellular material" includes preparations of a polypeptide, wherein the polypeptide is isolated from the cellular components of the cells from which it was isolated or recombinantly produced. Thus, a polypeptide substantially free of cellular material has less than about 30%, 20%, 10%, 5%, 2%, 1%, 0.5%, or 0.1% (by dry weight) of heterologous proteins (also referred to herein as "contaminating proteins") and / or variants of the polypeptide, e.g., polypeptides in different post-translational modified forms or other different versions of the polypeptide (e.g., polypeptide fragments). Also, when recombinantly produced, the polypeptide is generally substantially free of the culture medium, i.e., the culture medium represents less than about 20%, 10%, 2%, 1%, 0.5%, or 0.1% of the volume of the protein preparation. When produced by chemical synthesis, the polypeptide is generally substantially free of chemical precursors or other chemicals, i.e., is separated from the chemical precursors or other chemicals involved in the synthesis of the protein. Thus, such preparations of the protein have less than about 30%, 20%, 10%, or 5% (by dry weight) of chemical precursors or compounds other than the molecule of interest. In one embodiment, the polypeptides described herein are isolated or purified.

[0219] The polypeptides described herein can be produced by any method known in the art for the synthesis of antibodies, e.g., by chemical synthesis or recombinant expression techniques. The methods described herein use conventional techniques in molecular biology, microbiology, genetic analysis, recombinant DNA, organic chemistry, biochemistry, PCR, oligonucleotide synthesis and modification, nucleic acid hybridization, and related fields within the art, unless otherwise indicated. These techniques are described, for example, in the references cited herein and are fully explained in the literature.For example, reference is made to Maniatis T et al., (1982) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Sambrook J et al., (1989), Molecular Cloning: A Laboratory Manual, Second Edition, Cold Spring Harbor Laboratory Press, Sambrook J et al., (2001) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, Ausubel FM et al., Current Protocols in Molecular Biology, John Wiley & Sons (1987 and annual updates), Current Protocols in Immunology, John Wiley & Sons (1987 and annual updates), Gait (ed.) (1984) Oligonucleotide Synthesis: A Practical Approach, IRL Press, Eckstein (ed.) (1991) Oligonucleotides and Analogues: A Practical Approach, IRL Press, Birren B et al., (ed.) (1999) Genome Analysis: A Laboratory Manual, Cold Spring Harbor Laboratory Press, all of these references are hereby incorporated by reference in their entirety.

[0220] In one embodiment, the polypeptides described herein are prepared, expressed, produced, or isolated by any means including making, for example, by synthesis of a DNA sequence, through genetic manipulation. In one embodiment, such polypeptides include sequences (e.g., DNA sequences or amino acid sequences) that do not naturally occur within the antibody germline repertoire of an animal or mammal (e.g., human) in vivo.

[0221] Pharmaceutical composition In one aspect, the present disclosure provides a pharmaceutical composition comprising an FcRn / antigen-binding molecule or an FcRn-binding molecule disclosed herein for use in a method of treating an antibody-mediated disorder (e.g., an autoantibody-mediated disorder). In certain embodiments, these compositions comprise an FcRn-binding molecule and an FcRn / antigen-binding molecule comprising an antigen-binding domain. In some embodiments, these compositions comprise an FcRn-binding molecule conjugated to a molecule that does not specifically bind to a human protein. In some embodiments, the FcRn-binding molecule is an FcRn antagonist. In some embodiments, the FcRn antagonist comprises, or consists of, a variant Fc region or an FcRn-binding fragment thereof that inhibits the binding of the Fc region of an immunoglobulin to FcRn. Generally, these FcRn antagonists inhibit the binding of Fc-containing agents (e.g., antibodies and immunoadhesins) to FcRn in vivo, which results in an increased rate of degradation of the Fc-containing agent and, concomitantly, a reduced serum level of these agents.

[0222] The FcRn / antigen-binding molecule or FcRn-binding molecule of the present disclosure has a molecular weight in the range of about 50 kDa to about 140 kDa, which is about one-third of the molecular weight of full-length IgG (MW of about 150 kDa). In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of 50 kDa to 140 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of about 60 kDa to about 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of 60 kDa to 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of about 60 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of about 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of 60 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a molecular weight of 104 kDa.

[0223] In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule of the present disclosure has a predicted molecular weight in the range of about 50 kDa to about 140 kDa, which is about one-third of the molecular weight of full-length IgG (MW of about 150 kDa). In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of 50 kDa to 140 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of about 60 kDa to about 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of 60 kDa to 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of about 60 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of about 104 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of 60 kDa. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule has a predicted molecular weight of 104 kDa.

[0224] The formulations disclosed herein include bulk pharmaceutical compositions useful in the manufacture of pharmaceutical compositions (e.g., compositions suitable for administration to a subject or patient) that can be used in the preparation of unit dosage forms. In one embodiment, the compositions of the invention are pharmaceutical compositions. Such compositions include a prophylactically or therapeutically effective amount of one or more prophylactic or therapeutic agents of the invention (e.g., an FcRn / antigen-binding molecule) (or other prophylactic or therapeutic agent), and a pharmaceutically acceptable carrier.

[0225] In some embodiments, the pharmaceutical composition is formulated for administration via any suitable route of administration to a subject, and the routes of administration include, but are not limited to, intramuscular, intravenous, intradermal, intraperitoneal, subcutaneous, epidural, nasal, oral, rectal, topical, inhalation, buccal (e.g., sublingual), and transdermal administration. In one embodiment, the pharmaceutical composition is formulated to be suitable for intravenous administration to a subject. In one embodiment, the pharmaceutical composition is formulated to be suitable for subcutaneous administration to a subject.

[0226] Method of treatment The present disclosure also provides a method for treating an antibody-mediated disorder (e.g., an autoantibody-mediated disorder) in a subject, the method comprising administering to the subject a therapeutically effective amount of an FcRn / antigen-binding molecule or FcRn-binding molecule according to the present disclosure, or a pharmaceutical composition comprising the same.

[0227] In some embodiments, the antibody-mediated disorder is an autoimmune disease. In some embodiments, the autoimmune disease is allograft rejection, alopecia areata, ankylosing spondylitis, antiphospholipid syndrome, autoimmune Addison's disease, Alzheimer's disease, antineutrophil cytoplasmic autoantibody (ANCA), autoimmune diseases of the adrenal gland, autoimmune hemolytic anemia, autoimmune hepatitis, autoimmune myocarditis, autoimmune neutropenia, autoimmune oophoritis and orchitis, immune thrombocytopenia (ITP or idiopathic thrombocytopenic purpura), idiopathic thrombocytopeniaselected from the group consisting of purpura), immune-mediated thrombocytopenia, or idiopathic immune thrombocytopenia), autoimmune urticaria, Behçet's disease, bullous pemphigoid (BP), cardiomyopathy, Castleman disease, celiac sprue dermatitis, chronic fatigue immune dysfunction syndrome, chronic inflammatory demyelinating polyneuropathy (CIDP), Churg-Strauss syndrome, cicatricial pemphigoid, CREST syndrome, cold agglutinin disease, Crohn's disease, dilated cardiomyopathy, discoid lupus erythematosus, acquired epidermolysis bullosa, essential mixed cryoglobulinemia, factor VIII deficiency, fibromyalgia-fibromyositis, glomerulonephritis, Graves' disease, Guillain-Barré syndrome, Goodpasture syndrome, graft-versus-host disease (GVHD), Hashimoto's thyroiditis, hemophilia A, idiopathic inflammatory myopathy (IIM), idiopathic membranous neuropathy, idiopathic pulmonary fibrosis, IgA neuropathy, IgM polyneuropathy, immune-mediated necrotizing myopathy (IMNM), juvenile arthritis, Kawasaki disease, lichen planus, lichen sclerosus, lupus erythematosus, lupus nephritis, Ménière's disease, mixed connective tissue disease, mucous membrane pemphigoid, multiple sclerosis, type 1 diabetes, multifocal motor neuropathy (MMN), myasthenia gravis (MG), generalized myasthenia gravis (gMG), myositis, paraneoplastic bullous pemphigoid, pemphigoid gestationis, pemphigus vulgaris (PV), pemphigus foliaceus (PF), pernicious anemia, polyarteritis nodosa, polychondritis, polyglandular syndrome, polymyalgia rheumatica, polymyositis, dermatomyositis (DM), necrotizing autoimmune myopathy (NAM), anti-synthetase syndrome (ASyS), primary agammaglobulinemia, primary biliary cirrhosis, psoriasis, psoriatic arthritis, relapsing polychondritis, Raynaud phenomenon, Reiter syndrome, rheumatoid arthritis, sarcoidosis, scleroderma, Sjögren syndrome, solid organ transplant rejection, stiff-person syndrome, systemic lupus erythematosus, Takayasu arteritis, toxic epidermal necrolysis (TEN), Stevens-Johnson syndrome (SJS), temporal arteritis / giant cell arteritis, thrombotic thrombocytopenic purpura, ulcerative colitis, uveitis, dermatitis herpetiformis vasculitis, antineutrophil cytoplasmic antibody-related vasculitis, vitiligo, and Wegener granulomatosis.

[0228] In one embodiment, the FcRn / antigen-binding molecule antagonizes the binding of FcRn to the Fc region of the antibody. In one embodiment, the FcRn / antigen-binding molecule does not antagonize the binding of FcRn to albumin.

[0229] The present disclosure provides a method for reducing serum IgG in a subject, the method comprising administering to the subject a therapeutically effective amount of an FcRn / antigen-binding molecule or an FcRn-binding molecule according to the present disclosure, or a pharmaceutical composition comprising the same. In one embodiment, at least one of the IgG subtypes is reduced in the subject after administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In some embodiments, IgG1, IgG2, IgG3, IgG4, or any combination thereof is reduced. In some embodiments, the administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule is a single administration (e.g., a single therapeutic administration) of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In one embodiment, the level of serum IgG decreases in the subject after administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule as compared to the baseline level of serum IgG.

[0230] In one embodiment, at least about 35% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 45% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 40% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 45% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 50% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 55% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 60% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 65%, about 70%, about 75%, or about 80% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 65% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 70% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 75% total serum IgG reduction is obtained as compared to baseline serum IgG level. In one embodiment, at least about 80% total serum IgG reduction is obtained as compared to baseline serum IgG level.

[0231] In one embodiment, the level of serum IgG decreases in a subject, relative to the baseline level of serum IgG, following administration of the FcRn / antigen-binding molecule or FcRn-binding molecule. In one embodiment, a total serum IgG reduction of about 35% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 40% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 45% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 50% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 55% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 60% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 65%, about 70%, about 75%, or about 80% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 65% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 70% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 75% is obtained relative to the baseline serum IgG level. In one embodiment, a total serum IgG reduction of about 80% is obtained relative to the baseline serum IgG level.

[0232] In one embodiment, the level of FcRn does not decrease in a subject, relative to the baseline level of FcRn, following administration of the FcRn / antigen-binding molecule or FcRn-binding molecule. In one embodiment, an FcRn reduction of less than about 1%, 2%, 3%, 4%, or 5% is observed relative to the baseline FcRn level. In one embodiment, an FcRn reduction of less than about 10% is observed relative to the baseline FcRn level.

[0233] In one embodiment, the albumin level does not decrease in the subject after administration of the FcRn / antigen-binding molecule or FcRn-binding molecule as compared to the baseline level of albumin. In one embodiment, an albumin reduction of less than about 1%, 2%, 3%, 4%, or 5% is observed as compared to the baseline albumin level. In one embodiment, an albumin reduction of less than about 10% is observed as compared to the baseline albumin level.

[0234] In one embodiment, the total IgG, FcRn / antigen-binding molecule, FcRn-binding molecule, FcRn, or albumin in a patient's serum sample is analyzed using a bioanalytical method. In one embodiment, the total IgG, FcRn / antigen-binding molecule, FcRn-binding molecule, FcRn, or albumin in a patient's serum sample is analyzed using ELISA or an automated diagnostic analyzer (IVD). In one embodiment, the total IgG, FcRn / antigen-binding molecule, FcRn-binding molecule, FcRn, or albumin in a patient's serum sample is analyzed using ELISA. In one embodiment, the total IgG, FcRn / antigen-binding molecule, FcRn-binding molecule, FcRn, or albumin in a patient's serum sample is analyzed using an automated diagnostic analyzer (IVD). In one embodiment, the total FcRn in a patient's blood sample is analyzed using a bioanalytical method, preferably flow cytometry, microscopy, or immunoblotting.

[0235] In some embodiments, the reduction of total serum IgG is measured by the area under the reduction percentage curve (AUEC). In some embodiments, the reduction of total serum IgG is measured by the clearance (CL) of total serum IgG.

[0236] In some embodiments, the clearance of total serum IgG increases in a subject after administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule. In some embodiments, the clearance of total serum IgG in a subject after a single therapeutic administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule is equivalent to the clearance of total serum IgG in a subject after a single therapeutic administration of efgartigimod. In some embodiments, the clearance of total serum IgG in a subject after a single therapeutic administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule is similar or the same as the clearance of total serum IgG in a subject after a single therapeutic administration of efgartigimod. In some embodiments, the clearance of total serum IgG increases in a subject after a single therapeutic administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule as compared to the clearance of total serum IgG after a single therapeutic administration of efgartigimod. In some embodiments, the clearance of total serum IgG increases in a subject after a single therapeutic administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 125%, at least 150%, or at least 200% as compared to the clearance of total serum IgG after a single therapeutic administration of efgartigimod.

[0237] In some embodiments, the clearance of total serum IgG in a subject after a single administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule is equivalent to the clearance of total serum IgG in a subject after a single administration of an equivalent amount of efalizumab. In some embodiments, the clearance of total serum IgG in a subject after a single administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule is similar or the same as the clearance of total serum IgG in a subject after a single administration of an equivalent amount of efalizumab. In some embodiments, the clearance of total serum IgG increases in a subject after a single administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule as compared to the clearance of total serum IgG after a single administration of an equivalent amount of efalizumab. In some embodiments, the clearance of total serum IgG increases in a subject after a single administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 125%, at least 150%, or at least 200% as compared to the clearance of total serum IgG after a single administration of an equivalent amount of efalizumab.

[0238] In some embodiments, the clearance of an FcRn / antigen-binding molecule or an FcRn-binding molecule decreases in a subject after a single therapeutic administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule as compared to the clearance of efalizumab after a single therapeutic administration of efalizumab. In some embodiments, the clearance of an FcRn / antigen-binding molecule or an FcRn-binding molecule decreases in a subject after a single therapeutic administration of an FcRn / antigen-binding molecule or an FcRn-binding molecule by at least 1-fold, at least 1.5-fold, at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 12-fold, at least 15-fold, or at least 20-fold as compared to the clearance of efalizumab after a single therapeutic administration of efalizumab.

[0239] In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is decreased in a subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, as compared to the clearance of efgartigimod after a single administration of an equivalent amount of efgartigimod. In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is decreased in a subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, by at least 1-fold, at least 1.5-fold, at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 12-fold, at least 15-fold, or at least 20-fold as compared to the clearance of efgartigimod after a single administration of an equivalent amount of efgartigimod.

[0240] In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is less than about 0.2, about 0.19, about 0.18, about 0.17, about 0.16, about 0.15, about 0.14, about 0.13, about 0.12, about 0.11, about 0.1, about 0.09, about 0.08, about 0.07, about 0.06, or about 0.05 l / hour in a subject after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is less than 0.2, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11, 0.1, 0.09, 0.08, 0.07, 0.06, or 0.05 l / hour in a subject after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is in the range of about 0.05 to about 0.2 l / hour after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is about 0.2, about 0.19, about 0.18, about 0.17, about 0.16, about 0.15, about 0.14, about 0.13, about 0.12, about 0.11, about 0.1, about 0.09, about 0.08, about 0.07, about 0.06, or about 0.05 l / hour in a subject after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is in the range of 0.05 to 0.2 l / hour after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule. In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule is 0.2, 0.19, 0.18, 0.17, 0.16, 0.15, 0.14, 0.13, 0.12, 0.11, 0.1, 0.09, 0.08, 0.07, 0.06, or 0.05 l / hour in a subject after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule.

[0241] In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule in a subject is less than about 2.0, about 1.9, about 1.8, about 1.7, about 1.6, about 1.5, about 1.4, about 1.3, about 1.2, about 1.1, about 1.0, about 0.9, about 0.8, about 0.7, about 0.6, or about 0.5 mL / hour / kg after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or the FcRn-binding molecule to the subject.

[0242] In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule in a subject is less than 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, or 0.5 mL / hour / kg after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or the FcRn-binding molecule to the subject.

[0243] In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule in a subject is about 2.0, about 1.9, about 1.8, about 1.7, about 1.6, about 1.5, about 1.4, about 1.3, about 1.2, about 1.1, about 1.0, about 0.9, about 0.8, about 0.7, about 0.6, or about 0.5 mL / hour / kg after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or the FcRn-binding molecule to the subject.

[0244] In some embodiments, the clearance of the FcRn / antigen-binding molecule or the FcRn-binding molecule in a subject is 2.0, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, or 0.5 mL / hour / kg after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or the FcRn-binding molecule to the subject.

[0245] In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z in a subject, after a single therapeutic administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, is the t of efgartigimod after a single therapeutic administration of efgartigimod 1 / 2,zincreases as compared to. In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z in a subject, after a single therapeutic administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, is at least 0.5-fold, at least 1-fold, at least 1.5-fold, at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 12-fold, at least 15-fold, or at least 20-fold increased as compared to the t of efgartigimod after a single therapeutic administration of efgartigimod. 1 / 2,z

[0246] In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z in a subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, increases as compared to the t of efgartigimod after a single administration of an equivalent amount of efgartigimod. 1 / 2,z In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z in a subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, is at least 0.5-fold, at least 1-fold, at least 1.5-fold, at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, at least 6-fold, at least 7-fold, at least 8-fold, at least 9-fold, at least 10-fold, at least 12-fold, at least 15-fold, or at least 20-fold increased as compared to the t of efgartigimod after a single administration of an equivalent amount of efgartigimod. 1 / 2,z

[0247] In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z ​​In the subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, it is about 3 days, about 3.5 days, about 4 days, about 4.5 days, about 5 days, about 5.5 days, about 6 days, about 6.5 days, about 7 days, about 7.5 days, about 8 days, about 8.5 days, about 9 days, about 9.5 days, about 10 days, about 10.5 days, about 11 days, about 11.5 days, about 12 days, about 12.5 days, about 13 days, about 13.5 days, about 14 days, about 14.5 days, about 15 days, about 15.5 days, about 16 days, about 16.5 days, about 17 days, about 17.5 days, about 18 days, about 18.5 days, about 19 days, about 19.5 days, about 20 days, about 20.5 days, about 21 days, about 21.5 days, about 22 days, about 22.5 days, about 23 days, about 23.5 days, about 24 days, about 24.5 days, about 25 days, about 25.5 days, about 26 days, about 26.5 days, about 27 days, about 27.5 days, about 28 days, about 28.5 days, about 29 days, about 29.5 days, or more than about 30 days.

[0248] In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z In the subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, it is 3 days, 3.5 days, 4 days, 4.5 days, 5 days, 5.5 days, 6 days, 6.5 days, 7 days, 7.5 days, 8 days, 8.5 days, 9 days, 9.5 days, 10 days, 10.5 days, 11 days, 11.5 days, 12 days, 12.5 days, 13 days, 13.5 days, 14 days, 14.5 days, 15 days, 15.5 days, 16 days, 16.5 days, 17 days, 17.5 days, 18 days, 18.5 days, 19 days, 19.5 days, 20 days, 20.5 days, 21 days, 21.5 days, 22 days, 22.5 days, 23 days, 23.5 days, 24 days, 24.5 days, 25 days, 25.5 days, 26 days, 26.5 days, 27 days, 27.5 days, 28 days, 28.5 days, 29 days, 29.5 days, or more than 30 days.

[0249] In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z is in the range of about 3 days to about 30 days. In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,zIn the subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, it is about 3 days, about 3.5 days, about 4 days, about 4.5 days, about 5 days, about 5.5 days, about 6 days, about 6.5 days, about 7 days, about 7.5 days, about 8 days, about 8.5 days, about 9 days, about 9.5 days, about 10 days, about 10.5 days, about 11 days, about 11.5 days, about 12 days, about 12.5 days, about 13 days, about 13.5 days, about 14 days, about 14.5 days, about 15 days, about 15.5 days, about 16 days, about 16.5 days, about 17 days, about 17.5 days, about 18 days, about 18.5 days, about 19 days, about 19.5 days, about 20 days, about 20.5 days, about 21 days, about 21.5 days, about 22 days, about 22.5 days, about 23 days, about 23.5 days, about 24 days, about 24.5 days, about 25 days, about 25.5 days, about 26 days, about 26.5 days, about 27 days, about 27.5 days, about 28 days, about 28.5 days, about 29 days, about 29.5 days, or about 30 days.

[0250] In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z is in the range of 3 to 30 days. In some embodiments, the t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2,z In the subject, after a single administration of the FcRn / antigen-binding molecule or the FcRn-binding molecule, it is 3 days, 3.5 days, 4 days, 4.5 days, 5 days, 5.5 days, 6 days, 6.5 days, 7 days, 7.5 days, 8 days, 8.5 days, 9 days, 9.5 days, 10 days, 10.5 days, 11 days, 11.5 days, 12 days, 12.5 days, 13 days, 13.5 days, 14 days, 14.5 days, 15 days, 15.5 days, 16 days, 16.5 days, 17 days, 17.5 days, 18 days, 18.5 days, 19 days, 19.5 days, 20 days, 20.5 days, 21 days, 21.5 days, 22 days, 22.5 days, 23 days, 23.5 days, 24 days, 24.5 days, 25 days, 25.5 days, 26 days, 26.5 days, 27 days, 27.5 days, 28 days, 28.5 days, 29 days, 29.5 days, or 30 days.

[0251] In some embodiments, the half-life (t of the FcRn / antigen-binding molecule or the FcRn-binding molecule 1 / 2) is greater than about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, or about 100 hours after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or FcRn-binding molecule to a subject.

[0252] In some embodiments, the t 1 / 2 of the FcRn / antigen-binding molecule or FcRn-binding molecule is greater than 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 hours after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or FcRn-binding molecule to a subject.

[0253] In some embodiments, the half-life (t 1 / 2 ) of the FcRn / antigen-binding molecule or FcRn-binding molecule is about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, or about 100 hours after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or FcRn-binding molecule to a subject.

[0254] In some embodiments, the t 1 / 2 of the FcRn / antigen-binding molecule or FcRn-binding molecule is 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 hours after a single administration of a therapeutically effective amount of the FcRn / antigen-binding molecule or FcRn-binding molecule to a subject.

[0255] In some embodiments, the one-armed FcRn / antigen-binding molecule of the present disclosure sweeps the antigen more efficiently than a two-armed FcRn / antigen-binding molecule (e.g., a full-length antibody). In some embodiments, the one-armed FcRn / antigen-binding molecule sweeps the antigen more efficiently than the corresponding two-armed FcRn / antigen-binding molecule. In other words, in some embodiments, the removal of one arm of the two-armed FcRn / antigen-binding molecule results in a molecule that sweeps the antigen more efficiently than the two-armed FcRn / antigen-binding molecule.

[0256] As used herein, "sweep" refers to the ability of a molecule to remove an antigen from serum. "Sweeping" can be performed by a molecule (e.g., an antibody) having both pH-sensitive antigen binding and binding to FcRn at neutral or physiological pH at least at a threshold level. For example, a sweep molecule can bind to an antigen via an antigen-binding domain, bind to FcRn via an Fc region, and result in intracellular trafficking of the antigen / sweep antibody complex. The antigen can then be released from the complex and degraded in acidic endosomes. In some embodiments, the sweep molecule that is no longer bound to the antigen can then be released by the cell (e.g., by exocytosis) and returned to the serum.

[0257] In one embodiment, the FcRn / antigen-binding molecule or FcRn-binding molecule is administered to a subject simultaneously or sequentially with an additional therapeutic agent. In one embodiment, the additional therapeutic agent is an anti-inflammatory agent. In one embodiment, the additional therapeutic agent is a corticosteroid. In one embodiment, the additional therapeutic agent is rituximab, daclizumab, basiliximab, muromonab-CD3, infliximab, adalimumab, omalizumab, efalizumab, natalizumab, tocilizumab, eculizumab, golimumab, canakinumab, ustekinumab, or belimumab. In one embodiment, the additional therapeutic agent is a leukocyte-depleting agent.

[0258] In one embodiment, the additional therapeutic agent is a B cell-depleting agent. In one embodiment, the B cell-depleting agent is an antibody. In one embodiment, the B cell-depleting antibody is an antibody that specifically binds to CD10, CD19, CD20, CD21, CD22, CD23, CD24, CD37, CD53, CD70, CD72, CD74, CD75, CD77, CD79a, CD79b, CD80, CD81, CD82, CD83, CD84, CD85, or CD86.

[0259] In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule is administered intravenously. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule is administered intravenously once a week, once every two weeks, once every three weeks, once every four weeks, once a month, or once every six weeks.

[0260] In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule is administered subcutaneously. In some embodiments, the FcRn / antigen-binding molecule or FcRn-binding molecule is administered subcutaneously once a week, once every two weeks, once every three weeks, once every four weeks, once a month, or once every six weeks.

Example

[0261] The following examples are provided by way of illustration and not limitation.

[0262] Example 1 In Vitro Characterization of the OA-Fc-ABDEG Molecule Initial studies showed that OA-HEL-ABDEG, which contains one Fab fragment attached at the N-terminus to Fc-ABDEG via a native IgG1 hinge and specifically binds to hen egg white lysozyme (HEL), showed a two-fold improvement in FcRn occupancy and reached a level similar to that of efalizumab's FcRn occupancy (IC50 of TA-HEL-ABDEG = 14 nM, OA-HEL-ABDEG = 7 nM, efalizumab = 6 nM). HEL-ABDEG specifically binds to an antigen (hen egg white lysozyme) not found in humans, and thus it is not expected that the antigen-binding arm(s) of this molecule will bind to any protein found in humans. The improvement in FcRn occupancy upon removal of one arm of HEL-ABDEG is thought to be due at least in part to a reduction in steric hindrance.

[0263] The amino acid sequences of the CDRs of the HEL Fab fragment are as follows: VHCDR1 - GYGVN (SEQ ID NO: 18), VHCDR2 - MIWGDGNTDYNSALKS (SEQ ID NO: 19), VHCDR3 - ERDYRLDY (SEQ ID NO: 20), VLCDR1 - RASGNIHNYLA (SEQ ID NO: 21), VLCDR2 - YTTTLAD (SEQ ID NO: 22), and VLCDR3 - QHFWSTPRT (SEQ ID NO: 23).

[0264] The amino acid sequence of the bivalent (TA) HEL - ABDEG is as follows. HEL - Vk: DIQMTQSPSSLSASVGDRVTITCRASGNIHNYLAWYQQKPGKAPKLLIYYTTTLADGVPSRFSGSGSGTDYTFTISSLQPEDIATYYCQHFWSTPRTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 24) HEL - VH: QVQLQESGPGLVRPSQTLSLTCTVSGFSLTGYGVNWVRQPPGRGLEWIGMIWGDGNTDYNSALKSRVTMLKDTSKNQFSLRLSSVTAADTAVYYCARERDYRLDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC (SEQ ID NO: 25) HEL - VH - ABDEG: QVQLQESGPGLVRPSQTLSLTCTVSGFSLTGYGVNWVRQPPGRGLEWIGMIWGDGNTDYNSALKSRVTMLKDTSKNQFSLRLSSVTAADTAVYYCARERDYRLDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALKFHYTQKSLSLSPGK(SEQ ID NO: 35)

[0265] The amino acid sequence for the 1-arm type (OA) HEL-ABDEG is as follows. HEL-Vk: DIQMTQSPSSLSASVGDRVTITCRASGNIHNYLAWYQQKPGKAPKLLIYYTTTLADGVPSRFSGSGSGTDYTFTISSLQPEDIATYYCQHFWSTPRTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ ID NO: 24) HEL-VH: QVQLQESGPGLVRPSQTLSLTCTVSGFSLTGYGVNWVRQPPGRGLEWIGMIWGDGNTDYNSALKSRVTMLKDTSKNQFSLRLSSVTAADTAVYYCARERDYRLDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSC (SEQ ID NO: 25) HEL-VH-ABDEG-KNOB: QVQLQESGPGLVRPSQTLSLTCTVSGFSLTGYGVNWVRQPPGRGLEWIGMIWGDGNTDYNSALKSRVTMLKDTSKNQFSLRLSSVTAADTAVYYCARERDYRLDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLWCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALKFHYTQKSLSLSPG (SEQ ID NO: 34) Fc-Hole-ABDEG: DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLSCAVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLVSKLTVDKSRWQQGNVFSCSVMHEALKFHYTQKSLSLSPG (SEQ ID NO: 8)

[0266] Furthermore, previous studies have shown that the anti-IgE-ABDEG full-length antibody (mAb37) has a low FcRn occupancy (IC50 > 100 nM). mAb37 was previously described in WO / 2020 / 208177, the content of which is hereby incorporated by reference in its entirety. Three mAb37 variant constructs were developed: (1) a 1-armed construct (mAb37-OA-ABDEG) in which one mAb37 Fab fragment was removed, resulting in one Fab fragment remaining at the N-terminus of one of the Fc domains; (2) a 2-armed construct (Fc-ABDEG-mAb37) having two mAb37 scFvs fused via a 20GS linker to the C-terminus of each Fc domain; and (3) a 1-armed construct (OA-Fc-ABDEG-mAb37) having one mAb37 scFv fused via a 20GS linker to the C-terminus of one Fc domain. FcRn occupancy and degradation were measured using in vitro assays as described elsewhere herein. The FcRn occupancy results are shown in Table S1 and FIG. 1A below. The FcRn degradation results are shown in FIG. 1B. Table S1: FcRn occupancy of Fc-ABDEG-mAb37 variants

Table 7

[0267] The amino acid sequences for the CDRs of mAb37 are as follows: VHCDR1 - WHYLE (SEQ ID NO: 36), VHCDR2 - EIDPGTFTTNYNEKFKA (SEQ ID NO: 37), VHCDR3 - FSHFSGSNHDYFDY (SEQ ID NO: 38), VLCDR1 - RASQSIGTNIH (SEQ ID NO: 39), VLCDR2 - YASESIS (SEQ ID NO: 40), and VLCDR3 - QQSWSWPTT (SEQ ID NO: 41).

[0268] The amino acid sequence for mAb37-OA-ABDEG is as follows. mAb37-VL:

Chemistry

Chemistry

Chemistry

[0269] The amino acid sequence of OA-Fc-ABDEG-mAb37 is as follows. Knob-ABDEG-mAb37 scFv:

Chemistry

[0270] Example 2 Pharmacokinetics / Pharmacodynamics of OA-HEL-ADBEG in Cynomolgus Monkeys The objective of this study was to evaluate the pharmacokinetics (PK) and pharmacodynamics (PD) (total IgG) after single intravenous (IV) doses of OA-HEL-ABDEG at 10, 20, and 85 mg / kg in cynomolgus monkeys. After single-dose administration, an 8-week follow-up period ensued.

[0271] The study was conducted in an AAALAC-accredited facility under EU and French animal welfare regulations regarding the use of animals in experiments (European Directive 2010 / 63 / EU, and French Decree and Order 2013-118 of February 1, 2013, and French Decree and Order 2020-274 of March 17, 2020). Briefly, a total of 9 naive adult male cynomolgus monkeys (2.85 - 4.25 kg) were randomly assigned to one of three groups (n = 3 per group). The monkeys were dosed once by IV administration of 10, 20, or 85 mg / kg of OA-HEL-ABDEG. Blood samples were collected before treatment (pre-dose) and after treatment according to the schedule described in Table S2 for PK, PD, and immunogenicity (ADA) evaluations. Table S2: Summary of Sample Collection

Table 8

[0272] Pharmacodynamics were evaluated by measuring total cynomolgus monkey IgG serum levels using a validated sandwich ELISA. Briefly, plates were coated with mouse anti-monkey IgG antibody. The capture antibody binds to cynomolgus monkey IgG in the standards and samples. After washing any unbound material from the plates, an HRP-labeled anti-monkey IgG mouse monoclonal antibody was added to the wells. The plates were then washed and a TMB substrate solution was added to the wells. Color developed in direct proportion to the amount of bound cynomolgus monkey IgG in the standard or sample wells. A stop solution was then added over the wells and the plates were read on an ELISA plate reader at 450 nm (reference 620 nm).

[0273] The concentration of OA-HEL-ABDEG in monkey serum was determined using a validated PK assay. Briefly, anti-HN hFab was coated overnight on 96-well Maxisorp plates. The next day, the assay plates were washed and then blocked with 1% casein blocking buffer for 1 hour. Study samples, QC samples (two sets of LQC, MQC, and HQC), and calibration substances were diluted to 10 MRD in assay buffer (PBS + 0.1% casein). Samples were further diluted in sample diluent (PBS + 0.1% casein + 10% pooled cynomolgus monkey serum) if required. 50 μL of study samples, calibration substances, and QC samples were added in duplicate to the assay plates and incubated for 1 hour. The assay plates were washed and a secondary antibody solution (500 ng / mL in assay buffer) directed against HN was added for 1 hour. After washing the assay plates, 50 μL of detection solution (SA-HRP) was added to the plates for 30 minutes. After washing, TMB substrate was added. The color reaction was stopped after 15 minutes with sulfuric acid and the OD at 450 nm (reference 620) was measured on a suitable plate reader. The concentration of OA-HEL-ABDEG in study samples and QC samples was back-calculated from the calibration curve using a 5-parameter logistic fit with weighting of 1 / Y 2 using Graphpad Prism 8 software.

[0274] The presence of an immune response against OA-HEL-ABDEG was measured using a sandwich ELISA. Briefly, OA-HEL-ABDEG was coated overnight on ELISA plates and non-specific binding sites were blocked. Pre-dose and post-dose 1:100 dilutions of cynomolgus monkey serum in assay buffer were applied and incubated for 1 hour. The plates were washed, mouse anti-monkey IgG-HRP was added, and incubated for 1 hour. The plates were developed by adding TMB substrate. The enzyme reaction was stopped and the optical density values at 450 nm (reference 620 nm) were recorded.

[0275] To evaluate the pharmacodynamic effects of OA-HEL-ABDEG after single IV administration at doses of 10, 20, and 85 mg / kg, the levels of total circulating IgG in serum samples were determined at baseline (before dosing) and after dosing according to the blood sampling scheme in Table S2. The baseline IgG concentrations measured on Day -6 and the dosing day ranged from 12.2 to 27.4 mg / mL. On average, in Group 1 (10 mg / kg), the IgG concentration decreased to 64% of the pre-dose concentration within the median of 3 days, in Group 2 (20 mg / kg), it decreased to 50% of the pre-dose concentration within the median of 5 days, and in Group 3 (85 mg / kg), it decreased to 39% of the pre-dose concentration within the median of 7 days, resulting in mean IgG reductions of 36%, 50%, and 61% for Groups 1, 2, and 3, respectively. Subsequently, the IgG concentration increased to 100 - 218% of baseline in all groups until the end of the study. Values after Day 8 may have been affected by ADA and are therefore considered to be less certain. A summary of the PD data is provided in Table S3. Table S3: Summary of pharmacodynamic parameters of OA-HEL-ABDEG in cynomolgus monkeys

Table 9

[0276] To evaluate the PK profile of OA-HEL-ABDEG after single IV administration at doses of 10, 20, and 85 mg / kg, OA-HEL-ABDEG levels were determined in serum samples after dosing according to the blood sampling scheme in Table S2. Quantifiable concentrations were detected in most animals up to Day 15 of the study, after which they were below the lower limit of quantification (LLOQ). Exceptions were one animal in Group 1 where the last quantifiable concentration was measured on Day 29 of the study, one animal from Group 2 where quantification was possible up to Day 22 of the study, and one animal from Group 3 where the last quantifiable concentration was measured on Day 8 of the study. The obtained OA-HEL-ABDEG serum concentrations were plotted over time during the course of the study (Figure 2). Upon visual inspection of the pharmacokinetic profile, the mean serum OA-HEL-ABDEG increased as the dose increased. For each dose level, the individual profiles appeared to be homogeneous with respect to serum exposure, i.e., low inter-individual variability was observed. After the end of the infusion, in most animals, the concentration initially decreased rapidly and then more slowly in the second phase, followed by a steeper decline after Day 7 in the third phase. Since all animals in all groups were tested positive for ADA on and after Day 14, the steeper decline in the third phase could be explained by the presence of anti-drug antibodies (ADA). C max , AUC 0~tlast , t 1 / 2 , and values for CL are summarized in Table S4. Table S4: Summary of Pharmacokinetic Parameters of OA-HEL-ABDEG in Cynomolgus Monkeys

Table 10

[0277] A second NCA was performed using serum concentrations obtained after Day 7, i.e., without serum concentrations obtained prior to the detection of ADA. The adjusted calculations showed an increase of more than 20% compared to the calculations performed including values after the detection of ADA (Table S4). C 1 / 2 was more than 20% higher compared to the calculations performed including values after the detection of ADA (Table S4). C max , AUC 0~tlast , t1 / 2 The adjusted values for [substance name] and CL are summarized in Table S5. Table S5: Summary of Adjusted Pharmacokinetic Parameters of OA-HEL-ABDEG in Cynomolgus Monkeys

Table 11

[0278] As a conclusion, C max increased proportionally to the dose as the dose increased from 10 to 85 mg / kg. The AUC increased proportionally to the dose of 10 - 20 mg / kg but was less than the dose proportion of 20 - 85 mg / kg. IgG was quantifiable in all animals from 1 week before dosing to day 56. In Group 1 (10 mg / kg), the IgG concentration decreased to 64% of the pre-dose concentration within the median of 3 days, in Group 2 (20 mg / kg), it decreased to 50% of the pre-dose concentration within the median of 5 days, and in Group 3 (85 mg / kg), it decreased to 39% of the pre-dose concentration within the median of 7 days. Thereafter, the IgG concentration increased to 100 - 218% of the baseline until the end of the study in all groups. The values after day 8 of the study may have been affected by anti-OA-HEL-ABDEG antibodies (ADA, detected in all groups from day 15), and thus are considered to be less certain. ■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■■ ***

[0279] The present invention is, in scope, not limited by the specific embodiments described herein. Indeed, various modifications of the invention will become apparent to those skilled in the art from the foregoing description and the accompanying drawings. Such modifications are intended to fall within the scope of the appended claims.

Claims

1. An FcRn / antigen-binding molecule comprising an FcRn-binding molecule and a single antigen-binding domain linked to the FcRn-binding molecule, wherein the FcRn-binding molecule includes a variant Fc region that binds to FcRn with higher affinity at pH 6.0 and pH 7.4 compared to the corresponding wild-type Fc region, and the antigen-binding domain specifically binds to a non-human antigen.

2. The FcRn / antigen-binding molecule according to claim 1, wherein the variant Fc region comprises or consists of a first Fc domain and a second Fc domain that form a dimer, and the first Fc domain and / or the second Fc domain contain amino acids Y, T, E, K, and F at EU positions 252, 254, 256, 433, and 434, respectively.

3. The FcRn / antigen-binding molecule according to claim 1, wherein the variant Fc region comprises a first Fc domain and a second Fc domain that form a dimer.

4. The FcRn / antigen-binding molecule according to claim 1, wherein the non-human antigen is a non-human antigen that can be found in humans.

5. The FcRn / antigen-binding molecule according to claim 4, wherein the non-human antigen that can be found in the human is a viral antigen.

6. The FcRn / antigen-binding molecule according to claim 1, wherein the non-human antigen is a non-human antigen not found in humans.

7. The FcRn / antigen-binding molecule according to claim 6, wherein the non-human antigen not found in humans is selected from chicken egg white lysozyme (HEL) or ovalbumin.

8. The FcRn / antigen-binding molecule according to claim 7, wherein the non-human antigen not found in humans is HEL, the antigen-binding domain is a Fab fragment or scFv containing VH and VL, and the VH domain and VL domain contain a CDR sequence selected from the group consisting of HCDR1 (SEQ ID NO: 18), HCDR2 (SEQ ID NO: 19), HCDR3 (SEQ ID NO: 20), LCDR1 (SEQ ID NO: 21), LCDR2 (SEQ ID NO: 22), and LCDR3 (SEQ ID NO: 23).

9. The FcRn / antigen-binding molecule according to claim 3, wherein the antigen-binding domain is fused to the C-terminus of the first Fc domain or the second Fc domain.

10. The FcRn / antigen-binding molecule according to claim 3, wherein the antigen-binding domain is fused to the N-terminus of the first Fc domain or the second Fc domain.

11. The FcRn / antigen-binding molecule according to claim 3, wherein the first Fc domain and / or the second Fc domain is an IgG1 Fc domain.

12. The FcRn / antigen-binding molecule according to claim 3, wherein the antigen-binding domain is fused to the first Fc domain or the second Fc domain via a linker.

13. The FcRn / antigen-binding molecule according to claim 3, wherein the antigen-binding domain is fused to the first Fc domain or the second Fc domain via an IgG hinge region or a portion thereof.

14. The FcRn / antigen-binding molecule according to claim 3, wherein the first Fc domain and / or the second Fc domain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, 2, and 3.

15. The FcRn / antigen-binding molecule according to claim 2, wherein the amino acid sequence of the first Fc domain further comprises amino acid W at EU position 366.

16. The FcRn / antigen-binding molecule according to claim 15, wherein the amino acid sequence of the first Fc domain includes an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 5, and 6.

17. The FcRn / antigen-binding molecule according to claim 2, wherein the amino acid sequence of the second Fc domain further comprises amino acids S, A, and V at EU positions 366, 368, and 407, respectively.

18. The FcRn / antigen-binding molecule according to claim 17, wherein the amino acid sequence of the second Fc domain includes an amino acid sequence selected from the group consisting of SEQ ID NOs: 7, 8, and 9.

19. An FcRn / antigen-binding molecule according to claim 1, comprising a first heavy chain, a second heavy chain, and a light chain, (i) The first heavy chain consists of the amino acid sequence of SEQ ID NO: 34, (ii) The second heavy chain consists of the amino acid sequence of SEQ ID NO: 8, and (iii) The FcRn / antigen-binding molecule wherein the light chain consists of the amino acid sequence of SEQ ID NO:

24.

20. One or more isolated polynucleotides encoding an FcRn / antigen-binding molecule according to any one of claims 1 to 19.

21. An expression vector comprising one or more isolated polynucleotides as described in claim 20.

22. A host cell comprising one or more isolated polynucleotides as described in claim 20.

23. A method for producing an FcRn / antigen-binding molecule, comprising culturing the host cells described in claim 22 under conditions that enable the expression of the FcRn / antigen-binding molecule.

24. A pharmaceutical composition comprising an FcRn / antigen-binding molecule according to any one of claims 1 to 19 and at least one pharmaceutically acceptable carrier.

25. A pharmaceutical composition for use in the treatment of antibody-mediated disorders, comprising an FcRn / antigen-binding molecule according to any one of claims 1 to 19.

26. The pharmaceutical composition according to claim 25, wherein the antibody-mediated disorder is an IgG-mediated disorder.

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