ANTIBODY AND PHARMACEUTICAL COMPOSITION CONTAINING IT FOR THE TREATMENT OF IMMUNE-MEDIATED DISEASES

Abstract

The present invention provides an antibody capable of effectively preventing or treating immune diseases associated with decreased autoantigen functions by competitively inhibiting autoantibodies without side effects, and a pharmaceutical composition comprising same for treating immune diseases.

Description

Antibodies and pharmaceutical compositions containing the same for treating immune diseases

[0001] The present invention relates to an antibody and a pharmaceutical composition containing the same for treating an immune disease.

[0002] The formation of autoantibodies against body substances plays a crucial role in the pathogenesis of various autoimmune diseases. Specifically, when the antigen proteins of autoantibodies present on the cell surface form multimeric complexes, the antigen and antibody bind, resulting in cross-linking on the cell surface. This cross-linking can trigger complement-dependent cytotoxicity (CDC), antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), and antigen internalization through complement activation, leading to immune responses against the cells containing the antigen, or the loss of antigen function, which can cause disease.

[0003] To treat autoantibody-induced diseases, IVIg, which rapidly removes antibodies from the body, FcRN inhibitors, complement inhibitors, and antibodies that eliminate antibody-producing B cells are being used. However, there is a continuing need for therapeutics that can specifically suppress the pathological effects of autoantibodies. In this regard, Korean Patent Publication No. 2020-0098604 discloses an FcRn antibody and a method of use thereof.

[0004] However, in the case of the above prior art, there is a problem in that it causes many side effects because it has a mechanism that suppresses the immune function in the body rather than directly removing the cause of the disease.

[0005] The present invention is intended to solve various problems including the above-mentioned problems, and was made possible with the support of the National New Drug Development Project of the National Drug Development Fund of the Ministry of Science and ICT, the Ministry of Trade, Industry and Energy, and the Ministry of Health and Welfare (Project Identification Number: RS-2023-00283006). The purpose of the present invention is to provide an antibody that can competitively inhibit autoantibodies without side effects and effectively prevent or treat autoimmune diseases related to a decrease in the function of autoantigens, and a pharmaceutical composition for treating autoimmune diseases containing the same. However, these tasks are exemplary and the scope of the present invention is not limited thereby.

[0006] According to one aspect of the present invention, a recombinant homodimeric antibody having specificity for an autoantigen, which is derived from an autoantibody having specificity for an autoantigen and comprises a light chain and a heavy chain,

[0007] The above heavy chain is the light chain variable region (V) of the above autoantibody L ) comprising at least a portion of the heavy chain variable region (V) of the autoantibody, wherein the light chain comprises a heavy chain variable region (V) of the autoantibody. H ) is provided, wherein the homodimeric antibody comprises at least a portion of the region of the antibody.

[0008] According to another aspect of the present invention, a pharmaceutical composition for preventing or treating an autoimmune disease is provided, comprising the homodimeric antibody and a pharmaceutically acceptable carrier.

[0009] According to another aspect of the present invention, there is provided a recombinant dimeric antibody comprising a light chain variable region and a heavy chain variable region and having specificity for an autoantigen,

[0010] At least one monomer of the recombinant dimeric antibody has specificity for the autoantigen and comprises a light chain variable region (V) of an autoantibody having pathogenicity for an autoimmune disease. L ) and at least a portion of the heavy chain variable region (V) of the autoantibody H) are provided, wherein at least some regions of the antibodies are exchanged with each other.

[0011] The antibody of the present invention, as described above, and the pharmaceutical composition containing the antibody for treating immune diseases can suppress or eliminate the pathological effects of autoantibodies without side effects, and thus can be effectively used to prevent or treat immune diseases associated with decreased autoantigen function. Of course, the scope of the present invention is not limited by these effects.

[0012] Figure 1 is a schematic diagram illustrating an antibody structure according to one embodiment of the present invention. Figure 1A illustrates an autoantibody against an autoantigen, and Figures 1B to 1D illustrate antibodies according to one embodiment.

[0013] Figure 2a is a schematic diagram illustrating the binding orientation of an autoantibody (A) or therapeutic antibody to an autoantigen according to one embodiment of the present invention.

[0014] Figure 2b is a schematic diagram showing the difference in binding to an autoantigen that occurs depending on the difference in the binding orientation of the autoantibody of the present invention to an autoantigen (A) and the binding orientation of the therapeutic antibody to an autoantigen (B).

[0015] Figure 2c is a schematic diagram (A) showing an antigen-antibody complex formed by cross-linking an autoantigen of the autoantibody of the present invention, and a schematic diagram (B) showing a mechanism by which a therapeutic antibody inhibits cross-linking with an autoantigen.

[0016] Figure 2d is a schematic diagram showing antigen internalization by the autoantibody of the present invention (A) and a schematic diagram showing the mechanism by which the therapeutic antibody inhibits antigen internalization (B).

[0017] Figure 2e is a schematic diagram showing complement activation by the autoantibody of the present invention (A) and a schematic diagram showing the mechanism for inhibiting complement-dependent cytotoxicity (B).

[0018] Figure 2f is a schematic diagram showing antibody-dependent cellular cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP) by an autoantibody of the present invention (A) and a schematic diagram showing a mechanism for inhibiting dependent cellular cytotoxicity or antibody-dependent cellular phagocytosis by a therapeutic antibody (B).

[0019] Figure 3 is a graph showing the results of analyzing the competitive binding of autoantibody mAb637 and recombinant antibodies mAb637 B and mAb637 C to the autoantigen AChR. The vertical axis represents the normalized geometric mean Alexa647 signal change according to the concentration of the candidate antibody by diluting the candidate antibody at various concentrations, and the horizontal axis represents the value expressed as the dilution factor in log scale (X-fold molar excess).

[0020] Figure 4a is a graph analyzing the ratio of C5b-9 positive cells induced after contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes (CD46, CD55, CD59) were knocked out with autoantibody mAb35, or autoantibody mAb35 and the recombinant antibody of the present invention.

[0021] Figure 4b is a graph showing the results of analyzing the rate of cell death induced by CDC after contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes (CD46, CD55, CD59) were knocked out with autoantibody mAb35, or autoantibody mAb35 and the recombinant antibody of the present invention.

[0022] Figure 4c is a graph showing the results of analyzing the level of inhibition of C5b-9 positive cells induced by autoantibodies by the competitive recombinant antibodies by contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes are knocked out with the autoantibodies mAb35 humanized antibody and the competitive recombinant antibody mAb192 C IgG2 / 4 and mutants.

[0023] Figure 4d is a graph showing the results of analyzing the level of inhibition of CDC-induced cell death by contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes are knocked out with the autoantibodies mAb35 humanized antibody and the competitive recombinant antibody mAb192 C IgG2 / 4 and mutants.

[0024] Figure 5 is a graph showing the results of analyzing cell viability by contacting U87MG cells expressing aquaporin-4 with the NMO pathogenic autoantibody rAb58 or the recombinant antibody rAb53 C that competitively binds to the aquaporin-4 antigen.

[0025] Figure 6a is a graph showing the results of analyzing the internalization of AChR into cells by contacting autoantibodies mAb192 or mAb192 variants with 293T cells expressing the AChR gene and having three complement inhibitor genes knocked out. The graph shows, from top to bottom, the degree of internalization over time of mAb192, mAb192 C IgG1, mAb192 IgG2 / 4, mAb192 C IgG4, and mAb192 C IgG2 / 4.

[0026] Figure 6b is a graph showing the results of analyzing the aspect in which the internalization of AChR induced into cells by the autoantibody mAb35 is inhibited by the mAb192 variant, mAb192-IgG2 / 4, by contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes are knocked out with the autoantibody mAb35 alone or with both mAb35 and mAb192 variants.

[0027] Figure 6c is a graph showing the results of analyzing the aspect in which the internalization of AChR induced into cells by the autoantibody mAb35 is inhibited by the mAb192 C-IgG1, a mAb192 variant, by contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes are knocked out with the autoantibody mAb35 alone or with both mAb35 and mAb192 variants.

[0028] Figure 6d is a graph showing the results of analyzing the aspect in which the internalization of AChR induced into cells by the autoantibody mAb35 was inhibited by the mAb192 variant, mAb192 C, by contacting 293T cells expressing the AChR gene and in which three complement inhibitor genes were knocked out with the autoantibody mAb35 alone or with both mAb35 and mAb192 variants.

[0029] Figure 7a is a graph showing the results of analyzing the change in body weight over time after administering therapeutic antibody mAb35 C (5 mg / kg, Ab) to Lewis rats four times at 3.5-day intervals, followed by administration of myasthenia gravis pathogenic autoantibody mAb35 (1.5 mg / kg, MG).

[0030] Figure 7b is a graph showing the results of analyzing the change in clinical scores over time after administering therapeutic antibody mAb35 C (5 mg / kg, Ab) to Lewis rats four times at 3.5-day intervals, followed by administration of myasthenia gravis pathogenic autoantibody mAb35 (1.5 mg / kg, MG).

[0031] Figure 7c is a graph showing the results of analyzing the change in body weight (A) and clinical score (B) over time after administering therapeutic antibody mAb35 C (0.125 to 2 mg.kg) to Lewis rats four times at 3.5-day intervals, followed by administration of myasthenia gravis pathogenic autoantibody mAb35 (1.5 mg / kg).

[0032] Definition of terms

[0033] Unless otherwise defined herein, technical and scientific terms used herein have the same meaning as commonly used in the art to which this disclosure belongs. For the purpose of understanding this disclosure, the following definitions will apply, and terms used in the singular include the plural unless the context clearly indicates otherwise, and vice versa. Terms such as first, second, etc. may be used to describe various components, but the components should not be limited by these terms. These terms are used only for the purpose of distinguishing one component from another. For example, a first component could be referred to as a second component, and similarly, a second component could also be referred to as a first component, without departing from the scope of the present invention.

[0034] The term "and / or" as used in this document includes all combinations that the associated constructs may define.

[0035] As used herein, the terms 'polypeptide' and 'protein' may be used interchangeably to refer to a long chain peptide having the amino acid sequence of a native protein or an amino acid sequence having one or more mutations, such as deletions, substitutions, additions and / or insertions of one or more amino acid residues.

[0036] The term 'antibody' as used in this document includes a whole antibody, any antigen-binding fragment or a single chain thereof. The 'antibody' may comprise a glycoprotein comprising at least two heavy (H) chains and two light (L) chains or antigen-binding portions thereof, which are interconnected by disulfide bonds. Each heavy chain comprises a heavy chain variable region (V H or HV) and a heavy chain constant region. The heavy chain constant region may include four regions, namely CH1, hinge, CH2 and CH3. Each light chain may include a light chain variable region (V Lor LV) and a light chain constant region. The light chain constant region may consist of one region, namely CL. V H and V L Regions may be interspersed with more conserved regions called framework regions (FR), and further subdivided into hypervariable regions (HVRs) called complementarity determining regions (CDRs). Each V H and V L An immunoglobulin can comprise three CDRs and four FRs arranged from amino terminus to carboxy terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable regions of the heavy and light chains contain binding domains that interact with antigens. The constant region can mediate binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system.

[0037] The term "half-antibody fragment" as used in this document refers to an antigen-binding fragment that contains only one of the two antigen-binding sites of a whole antibody. The half-antibody fragment comprises the light chain variable region (V L ) and heavy chain variable region (V H ) and may have binding specificity for an autoantigen. The dimeric antibody of the present invention may comprise two half antibody fragments. The half antibody fragments may further comprise all or part of a light chain constant region and a heavy chain constant region. The half antibody fragments may further comprise a light chain constant region (CL), and at least one of heavy chain constant regions CH1, CH2, and CH3. The half antibody fragments may be V L -CL and V H -CH1, or V L -CL and V H -CH1-CH2-CH3 may be included. The above V LBetween CL and V H The boundary region between CH1 and CH1, or between one or more of the domains of CH1-CH2-CH3, may comprise a linker. The linker may be, for example, an SS sequence.

[0038] The term "monoclonal antibody," as used herein, refers to an antibody obtained from a substantially homogeneous population of antibodies, wherein the individual antibodies within the population are identical and / or bind to the same epitope, except for a small number of variant antibodies. For example, monoclonal antibodies can be produced by a variety of techniques, including hybridoma methods, recombinant DNA methods, phage-display methods, and methods utilizing transgenic animals containing all or part of the human immunoglobulin locus.

[0039] The term "binding affinity" as used herein refers to the intrinsic binding affinity that reflects the 1:1 interaction between members of a binding pair (e.g., an antibody and an antigen). The affinity of a molecule X for its partner Y can generally be expressed as a dissociation constant (Kd). Affinity can be measured by conventional methods known in the art, including those described herein.

[0040] The terms 'Fc', 'Fc region', and 'Fc domain' used in this document refer to the portion of an antibody molecule consisting of the hinge region or a portion thereof, CH2, and CH3 regions. The Fc region of the IgG class refers to, but is not limited to, the region from the 226th cysteine ​​to the C-terminus, or from the 230th proline to the C-terminus, using EU numbering (also called EU INDEX). The Fc region can be suitably obtained by partially digesting IgG1, IgG2, IgG3, and IgG4 monoclonal antibodies with a proteolytic enzyme such as pepsin or papain, and then re-eluting the fraction adsorbed to a protein A column or a protein G column.

[0041] The term "linker," as used herein, refers to a nucleic acid, amino acid, or non-peptide residue that can be inserted between one or more molecules, e.g., one or more component regions. For example, a linker can be used to provide a desired site of interest between the components to facilitate manipulation. A linker can also serve to enhance expression of the fusion protein from a host cell and / or to reduce steric hindrance so that the components can assume their optimal tertiary structure and / or interact appropriately with a target molecule. A linker sequence may comprise one or more amino acids naturally linked to the receptor component, or may be an added sequence used to enhance expression of the fusion protein, provide a desired site of interest, enable the component regions to form an optimal tertiary structure, and / or enhance interaction of the component with its target molecule. Preferably, the linker increases the flexibility of the fusion protein components without interfering with the structure of each functional component within the fusion protein.

[0042] An “antibody that binds to the same epitope” as a reference antibody can refer to an antibody that blocks binding of the reference antibody to its antigen by at least 10%, at least 20%, at least 30%, at least 40%, or at least 50% in a competition assay, and the reference antibody can refer to an antibody that blocks binding of the antibody to its antigen by at least 10%, at least 20%, at least 30%, at least 40%, or at least 50%. For example, an “antibody that binds to the same epitope” as a reference antibody can refer to an antibody that blocks binding of the reference antibody to its antigen by at least 50% in a competition assay, and the reference antibody can refer to an antibody that blocks binding of the antibody to its antigen by at least 50%.

[0043] The term 'human antibody' as used in this document is an antibody having an amino acid sequence corresponding to the amino acid sequence of an antibody produced by a human or human cell or derived from a non-human source utilizing the human antibody repertoire or other human antibody-coding sequences.

[0044] The term 'humanized antibody' as used herein refers to a molecule in which the antigen-binding portion of the molecule is substantially derived from an immunoglobulin from a non-human species, while the remainder of the immunoglobulin structure of the molecule is based on the structure and / or sequence of a human immunoglobulin.

[0045] The term 'chimeric antibody' as used herein refers to an antibody in which a portion of the heavy and / or light chain is derived from a particular source or species, while the remainder of the heavy and / or light chain is derived from another source or species.

[0046] The term "host cell" as used herein refers to a cell into which an exogenous nucleic acid has been introduced. Host cells include transformants and transformed cells, including the primary transformed cell and its progeny, regardless of the number of passages.

[0047] The term "pharmaceutical composition" as used herein refers to a preparation containing an active ingredient. Furthermore, the term "preparation" means that at least one additional ingredient is present in the pharmaceutical composition in addition to the active ingredient. A pharmaceutical composition is a preparation suitable for administration to a subject, such as a human patient. The pharmaceutical composition may be in a lyophilized form, for example, a solution formed after reconstitution of a lyophilized pharmaceutical composition using saline or water, or a solution that does not require reconstitution. The pharmaceutical composition may be liquid or solid.

[0048] The terms "administration" or "administering" as used herein refer to the step of providing a pharmaceutical composition or active ingredient to a subject. The pharmaceutical composition may be administered via various suitable routes.

[0049] The term 'therapeutically effective amount' as used herein means the level, amount or concentration of a pharmaceutical composition containing an agent required to treat a disease, disorder or condition without causing significant negative or adverse side effects.

[0050] As used herein, the terms 'treat', 'treating' or 'treatment' mean the alleviation or reduction (including partial reduction, substantial reduction, almost complete reduction and complete reduction), resolution or prevention (whether temporary or permanent) of a disease, disorder or condition so as to achieve a desired therapeutic result, for example, by healing injured or damaged tissue, or by altering, changing, strengthening, improving, ameliorating and / or beautifying a pre-existing or recognized disease, disorder or condition. 'Prevention' means the delay of the onset of a disease, disorder or condition. Prevention may be considered complete if the onset of the disease, disorder or condition is delayed for a predetermined period of time.

[0051] The term "combination" as used herein refers to any form of two or more different therapeutic agents that allows the second agent to be administered while the previously administered agent remains effective in the body. For example, two treatments are simultaneously effective in the subject, which may include a synergistic effect of the two agents. The different treatments may be administered simultaneously or sequentially as a single agent or as separate agents.

[0052] According to one aspect of the present invention, a recombinant homodimeric antibody having specificity for an autoantigen, which is derived from an autoantibody having specificity for an autoantigen and comprises a light chain and a heavy chain,

[0053] The above heavy chain is the light chain variable region (V) of the above autoantibody L ) comprising at least a portion of the heavy chain variable region (V) of the autoantibody, wherein the light chain comprises a heavy chain variable region (V) of the autoantibody. H ) is provided. For example, some regions of the variable regions of the heavy and light chains of the autoantibody may be exchanged with each other.

[0054] The above autoantibodies may have pathogenic properties for autoimmune diseases.

[0055] In one embodiment, the antibody of the present invention may have a variable region (e.g., a complementary determining region) derived from the autoantibody. That is, the antibody of one embodiment may include an amino acid sequence corresponding to a variable region of the autoantibody. For example, the heavy chain variable region of the antibody of one embodiment may be derived from a light chain variable region of the autoantibody, and the light chain variable region may be derived from a heavy chain variable region of the autoantibody. When the antibody of one embodiment includes a constant region, the constant region may be derived from the autoantibody, and in this case, the constant region may have a sequence that is preserved as is of the autoantibody, or may further include amino acid mutations (e.g., amino acid substitutions, deletions, and / or insertions). It should be understood that in the antibody of one embodiment, the constant region may not be derived from the autoantibody.

[0056] Among the individual functional domains of the antibody, the domains derived from the autoantibody may have 99% or more, 98% or more, 95% or more, or 90% or more homology with the corresponding amino acid sequence of the autoantibody. In this case, the activity of the mutated antibody may be 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, or 95% or more identical to the activity (e.g., binding affinity to an autoantigen) of the unmutated homodimeric antibody.

[0057] In the above homodimeric antibody, the autoantigen may be a membrane protein. A plurality of the autoantigens may be cross-linked by the autoantibody, forming an antigen-antibody complex structure through the cross-linking, and the antigen-antibody complex may be internalized through the cross-linking.

[0058] In the above homodimeric antibody, complement binding can be induced by the cross-linking, and the homodimeric antibody can bind to an epitope identical to the epitope of the autoantigen to which the autoantibody binds or a site adjacent to the identical epitope, thereby inhibiting the autoantibody from binding to the autoantigen.

[0059] That is, the homodimeric antibody may reduce or prevent the formation of a cross-linking structure by binding of multiple autoantigens to the autoantibody. In addition, the homodimeric antibody may reduce or prevent the internalization of the antigen-antibody complex by the cross-linking.

[0060] In the above homodimeric antibody, the binding orientation of at least one monomer of the homodimeric antibody to the autoantigen may be different from the binding orientation of the autoantibody to the autoantigen, and the binding of one monomer of the homodimeric antibody to the first autoantigen may have a binding orientation that prevents the remaining monomer of the homodimeric antibody from binding to the second autoantigen present on the cell surface.

[0061] In the above homodimeric antibody, it may have a conformation that does not bind to multiple complex autoantigens among the complex autoantigens present on the cell surface, may not cross-link two or more complex autoantigens present on the cell surface, and the Fc region of the heavy chain may be IgG1 Fc, IgG4 Fc, or IgG2 / 4 hybrid Fc.

[0062] In the above homodimeric antibody, the autoantibody comprises a first light chain variable region (V L ) and the first heavy chain variable region (V H ) and has specificity for an autoantigen; and a second half-antibody fragment comprising a second light chain variable region (VL) and a second heavy chain variable region (VH) and has specificity for the autoantigen,

[0063] The above homodimeric antibody may be characterized by comprising one or more of the following half-antibody fragments:

[0064] i) The first light chain variable region (V) L ) comprising a heavy chain, and the first heavy chain variable region (V H ) containing light chains;

[0065] ii) The second light chain variable region (V L ) comprising a heavy chain, and the second heavy chain variable region (V H ) containing light chains;

[0066] iii) The first light chain variable region (V L ) comprising at least one of CDRL1, CDRL2 and CDRL3, and the first heavy chain variable region (V L ) and a light chain comprising at least one of CDRH1, CDRH2 and CDRH3 and the remaining CDRL portion not included in the heavy chain; and

[0067] iv) The second light chain variable region (V)L ) comprising at least one of CDRL1, CDRL2 and CDRL3, and the second heavy chain variable region (V L ) and a light chain comprising at least one of CDRH1, CDRH2 and CDRH3 and the remaining CDRL portion not included in the heavy chain.

[0068] In the above homodimeric antibody, the autoantibody may include a first half-antibody fragment comprising a first light chain variable region and a first heavy chain variable region and having specificity for the autoantigen; and a second half-antibody fragment comprising a second light chain variable region and a second heavy chain variable region and having specificity for the autoantigen.

[0069] The above homodimeric antibody is a V of at least one half-antibody fragment among the half-antibody fragments of the above autoantibody. L -CL and V H -CH1 may have been exchanged with each other.

[0070] According to another aspect of the present invention, a pharmaceutical composition for preventing or treating an autoimmune disease is provided, comprising the homodimeric antibody and a pharmaceutically acceptable carrier.

[0071] According to another aspect of the present invention, there is provided a recombinant dimeric antibody comprising a light chain variable region and a heavy chain variable region and having specificity for an autoantigen,

[0072] At least one monomer of the recombinant dimeric antibody has specificity for the autoantigen and comprises a light chain variable region (V) of an autoantibody having pathogenicity for an autoimmune disease. L ) and at least a portion of the heavy chain variable region (V) of the autoantibody H ) are provided, wherein at least some regions of the antibodies are exchanged with each other.

[0073] According to another aspect of the present invention, a method for treating an autoimmune disease is provided, comprising administering the pharmaceutical composition to a subject suffering from an autoimmune disease.

[0074] According to another aspect of the present invention, a dimeric antibody comprising a light chain variable region and a heavy chain variable region and having specificity for an autoantigen is provided. The dimeric antibody may be a recombinant dimeric antibody. The dimeric antibody may be a heterodimeric antibody or a homodimeric antibody. For example, the dimeric antibody may be a homodimeric antibody. The dimeric antibody may be one in which at least a portion of the light chain variable region of an autoantibody having specificity for the autoantigen and at least a portion of the heavy chain variable region of the autoantibody are exchanged with each other. The autoantibody may have specificity for the autoantigen and pathogenicity for an autoimmune disease. When the dimeric antibody is a heterodimeric antibody, one monomer may be one in which at least a portion of the light chain variable region of the autoantibody and at least a portion of the heavy chain variable region of the autoantibody are exchanged with each other, and the other monomer may be one in which the amino acid sequence of the autoantibody is preserved. Alternatively, the heterodimeric antibody may be one in which at least a portion of the light chain variable region of the autoantibody and at least a portion of the heavy chain variable region of the autoantibody are exchanged between the two monomers, but the exchanged regions may be different.

[0075] Unless otherwise stated herein, the description of a heterodimeric antibody may be equally applicable to a homodimeric antibody, and the description of a homodimeric antibody may be equally applicable to a heterodimeric antibody. In the present specification, the homodimeric antibody may be a two-monomer antibody in which at least a portion of the light chain variable region of the autoantibody and at least a portion of the heavy chain variable region of the autoantibody are exchanged, and the exchanged regions may be identical.

[0076] The dimeric antibody may comprise two monomers. For example, the dimeric antibody may comprise a first half antibody fragment comprising a first light chain variable region and a first heavy chain variable region, and a second half antibody fragment comprising a second light chain variable region and a second heavy chain variable region. In one embodiment, at least one monomer of the dimeric antibody (e.g., the first half antibody fragment and / or the second half antibody fragment) may be a monomer in which at least a portion of a light chain variable region of the autoantibody that binds to the autoantibody and at least a portion of a heavy chain variable region of the autoantibody are exchanged with each other. That is, the first light chain variable region and / or the second light chain variable region of the dimeric antibody may be a sequence corresponding to at least a portion of a heavy chain variable region of the autoantibody, and the first heavy chain variable region and / or the second heavy chain variable region of the dimeric antibody may be a sequence corresponding to at least a portion of a light chain variable region of the autoantibody. The above autoantibody may have specificity for the above autoantigen and may have pathogenicity for an autoimmune disease.

[0077] In one embodiment of the present invention, when the dimeric antibody is a heterodimeric antibody, one of the monomers of the heterodimeric antibody (e.g., the first half antibody fragment or the second half antibody fragment) may be a monomer in which at least a portion of the light chain variable region of an autoantibody having specificity for the autoantigen and at least a portion of the heavy chain variable region of the autoantibody are exchanged with each other.

[0078] The 'dimeric antibody' of the above embodiment binds to an autoantigen and competitively inhibits the autoantibody as described below, and may be referred to herein as a 'competitive autoantibody inhibitor', 'candidate antibody', or 'therapeutic antibody'.

[0079] The autoantibody having specificity for the above autoantigen may be naturally found in an individual. The individual may be a mammal, including a human. The autoantibody may be a pathogenic autoantibodies for an autoimmune disease. For example, the autoantibody may be a pathogenic autoantibody for an autoimmune disease associated with a decrease in the function of the autoantigen for the autoimmune disease. For example, the autoantibody may be a pathogenic autoantibody that binds to the autoantigen, aquaporin-4, and causes antigen internalization, induction of ADCC, induction of CDC, and / or reduction of ADCP, thereby causing the autoimmune disease. For example, the autoantibody may be a pathogenic autoantibody for myasthenia gravis and / or neuromyelitis optica. The autoantibody may be a whole antibody. The autoantibody may comprise a light chain variable region - a light chain constant region (V). L -CL), and heavy chain variable region-heavy chain constant region 1 - heavy chain constant region 2 - heavy chain constant region 3 (V H -CH1-CH2-CH3). The autoantibody may have two antigen binding sites. Each antigen binding site of the autoantibody may bind to the same epitope.

[0080] The autoantibody may bind to the autoantigen. The autoantibody may be a dimeric antibody, with each fragment of the autoantibody binding to the same epitope of the autoantigen. Accordingly, the autoantibody may be a bivalent antibody. For example, the autoantibody may be a monospecific bivalent antibody.

[0081] The dimeric antibody of the present invention may bind to the autoantigen. The first half-antibody fragment or the second half-antibody fragment may each bind to the same epitope of the autoantigen. The competitive autoantibody inhibitor may be a bivalent antibody. For example, the competitive autoantibody inhibitor may be a monospecific bivalent antibody.

[0082] The autoantibody may bind to a complex autoantigen. The complex autoantigen may be an autoantigen comprising multiple subunit proteins. The complex autoantigen may comprise multiple subunit proteins, and the subunit proteins may comprise two or more identical subunits. For example, the complex autoantigen may be an acetylcholine receptor (AChR) comprising five subunits, including two alpha subunits. The complex autoantigen may be an integral transmembrane protein. The complex autoantigen may have an epitope on the cell surface. The epitope may be, for example, the main immunogenic region (MIR) of the alpha subunit of the AChR. The autoantibody may be, for example, a monoclonal antibody IgG1-637 that binds to the main immunogenic region (MIR) of the alpha subunit of human AChR. The antibody-binding site in the MIR may be residues 1-32 or 60-81. An autoantibody that binds to the alpha subunit of the AChR may cause myasthenia gravis.

[0083] Additionally, the complex autoantigen may be aquaporin-4 (AQP4). The production of autoantibodies against aquaporin-4, a water channel in nerve cells, can lead to the development of neuromyelitis optica (NMO). Aquaporin-4 exists in isoforms called M1 and M23. Among them, M23 forms cluster and have increased binding affinity to autoantibodies. Therefore, the M23 form can be considered pathogenic.

[0084] In the above neuromyelitis optica patients, loss of aquaporin-4 in the central nervous system is observed. Aquaporin-4 antibodies are found in more than 95% of neuromyelitis optica patients and are autoantibodies with CDC or NK-based cytotoxicity. Removal of aquaporin-4 antibodies from plasma improves symptoms of neuromyelitis optica, and transfer of aquaporin-4 antibodies to healthy animals results in symptoms similar to those of neuromyelitis optica. Loss of aquaporin-4 in the central nervous system is observed in neuromyelitis optica patients. The aquaporin-4 may have a tetrameric structure. The complex autoantigen may have an epitope on the cell surface. Residues 148-149 and 151 of extracellular loop C of aquaporin-4, residues 226 and 228 of loop E, residues 63-65 and 69 of extracellular loop A, residues 141, 151 and 154 of loop C, and / or residues 230-231 of loop E may be important residues for binding to pathogenic autoantibodies that bind to aquaporin-4. For example, an epitope of aquaporin-4 that binds to a pathogenic autoantibody may include residues 148-149, 151 of extracellular loop C of aquaporin-4, residues 226, 228 of loop E, residues 63-65, 69 of extracellular loop A, residues 141, 151, 154 of loop C, and / or residues 230-231 of loop E. A non-limiting example of such autoantibodies is rAb53, a rat-derived sequence that binds to aquaporin-4. The two major isoforms of aquaporin-4 mentioned above are expressed in astrocytes. The long isoform (M1) initiates translation at Met-1, and the short isoform (M23) initiates translation at Met-23. When expressed, M23 aquaporin-4 particles assemble in the membrane as regular square arrays called orthogonal arrays of particles (OAP).OAP formation by M23 results from a tetramer-tetramer interaction involving residues just downstream of Met-23 in the cytoplasmic N-terminus, whereas residues just upstream of Met-23 in M1 disrupt this interaction. M1 does not form OAP on its own, but co-assembles with M23 into a heterotetramer that limits OAP size. A hallmark of neuromyelitis optica (NMO) is the presence of autoantibodies to aquaporin-4 (NMO-IgG) in the serum. The presence of NMO-IgG is specific for NMO, and some studies have shown that serum NMO-IgG titers correlate with NMO disease activity. NMO-IgG is pathogenic for NMO. NMO-IgG can bind to the extracellular domain of M1 or M23, and is known to have particularly high affinity for M23.

[0085] Hereinafter, various structures of dimeric antibodies included in the antibody platform of the present invention and their use in preventing or treating various autoimmune diseases will be exemplified with reference to the drawings.

[0086] Figure 1 is a schematic diagram illustrating an antibody structure according to one embodiment. Figure 1A illustrates an autoantibody against an autoantigen, and Figures 1B to 1D illustrate antibodies according to one embodiment. While Figure 1 illustrates a homodimeric antibody structure as an example, it should be understood that the embodiment is not limited thereto.

[0087] As described above, in one embodiment, at least one monomer or both monomers of the antibody may comprise at least a portion of the light chain variable region of the autoantigen that binds to the autoantibody and at least a portion of the heavy chain variable region of the autoantibody. The antibody may be one in which portions of the variable regions of the heavy and light chains of the autoantibody are exchanged with each other.

[0088] In one embodiment, the exchange may be an exchange of one or more, two or more, or three or more consecutive amino acid residues, or functional domains. The exchange may be such that the complex formed by binding of the autoantigen and the autoantibody has a different conformation from the conformation of the complex while maintaining the binding affinity to the antigen. The different conformation may be such that when one antigen-binding site binds to a complex autoantigen present on the cell surface, the other antigen-binding site has an orientation that prevents binding to another complex autoantigen.

[0089] In one embodiment, the exchanged one or more, two or more, or three or more consecutive amino acid residues, or functional domains, of the antibody may comprise additional mutations. For example, the sequence of the exchanged one or more, two or more, or three or more consecutive amino acid residues, or functional domain may have at least 99% homology, 98% homology, 97% homology, 96% homology, 95% homology, or 90% homology with the sequence of the consecutive amino acid residues, or functional domain of the autoantibody, while maintaining at least 50%, 60%, 70%, 80%, 90%, or 95% binding affinity to the antigen.

[0090] Referring to Figure 1, an antibody of one embodiment comprises a light chain variable region (V) of the autoantigen. L) and the light chain constant region (CL) and heavy chain variable region (V H ) and heavy chain constant region 1 (CH1) are exchanged with each other (B in Fig. 1), and the light chain variable region (V) of the autoantigen L ) and heavy chain variable region (V H ) are interchanged antibodies (C in Fig. 1), or light chain variable regions (V L ) and one or more of CDRL1, CDRL2 and CDRL3 and the heavy chain variable region (V H ) may include an antibody (D in Figure 1) in which one or more of CDRH1, CDRH2 and CDRH3 are exchanged with each other. In the case of the antibody shown in D in Figure 1, the light chain variable region (V L ) and the heavy chain variable region (V) of CDRL1, CDRL2 and CDRL3 H ) may have exchanged CDRH1, CDRH2, and CDRH3.

[0091] In addition, although not shown, each monomer of the dimeric antibody may have different regions exchanged. For example, one monomer of the dimeric antibody may have the light chain variable region (V) of the autoantigen as shown in B of FIG. 1 L ) and the light chain constant region (CL) and heavy chain variable region (V H ) and heavy chain constant region 1 (CH1) are exchanged with each other, and the remaining one monomer is the light chain variable region (V) of the autoantigen as shown in C of Figure 1. L ) and heavy chain variable region (V H ) may be interchangeable antibodies.

[0092] Additionally, although FIG. 1 illustrates that the aforementioned exchange occurs in both monomers of the antibody of one embodiment, such exchange may occur in only one monomer of the dimer.

[0093] In one embodiment of the dimeric antibody, the two monomers may be referred to as a first half antibody fragment and a second half antibody fragment, respectively. The first half antibody fragment may further comprise a first light chain constant region (CL) and a first heavy chain constant region (CH), and the second half antibody fragment may further comprise a second light chain constant region (CL) and a second heavy chain constant region (CH). The first heavy chain constant region (CH) and the second heavy chain constant region (CH) may each comprise heavy chain constant region 1 (CH1), heavy chain constant region 2 (CH2), and heavy chain constant region 3 (CH3). In one embodiment, the antibody comprises V in the first half antibody fragment. L -CL and V H -CH1 is an antibody with an exchanged second half antibody fragment, V L -CL and V H -CH1 is an antibody with an exchanged CH1, or V in the first half antibody fragment L -CL and V H -CH1 is exchanged with each other and V in the second half antibody fragment L -CL and V H - May include antibodies in which CH1 is exchanged with each other.

[0094] The autoantigen may be a membrane protein. The autoantigen may be an integral membrane protein or an extrinsic membrane protein, and specifically, may be a transmembrane protein or an integral monomeric protein (e.g., an integral monomeric protein attached to the outside of a cell membrane). The monotopic protein may interact with one layer of a phospholipid bilayer. For example, the epitope for the autoantibody of the autoantigen may be distributed on the cell surface. The autoantigen may be a complex autoantigen comprising multiple subunit proteins. The complex autoantigen comprises multiple subunit proteins, and the subunit proteins may comprise two or more identical subunits.

[0095] The autoantibody may bind to the same subunit of the first complex antigen and to the same subunit of the second complex antigen, thereby cross-linking the first complex antigen and the second complex antigen. By the cross-linking, the complex of the autoantibody and the complex antigen may be internalized. The same subunit may be, for example, the alpha subunit of the AChR, and the autoantibody may bind to a MIR epitope of the alpha subunit.

[0096] Complement binding may be induced by the above cross-linking. The complement may bind to the Fc region of the antibody bound to the first complex autoantigen and the second complex autoantigen. Antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) may be induced by the above binding.

[0097] In one embodiment, the antibody may avoid this problem by not binding to multiple autoantigens. This is exemplified below with reference to FIGS. 2A through 2F.

[0098] Figures 2a to 2f are schematic diagrams explaining the treatment mechanism of an immune disease caused by an autoantibody, and are schematic diagrams of a change in binding orientation between an antibody and an autoantigen and the resulting treatment mechanism according to one example.

[0099] Figure 2a is a schematic diagram illustrating the binding orientation of an antibody (B of Figure 2a) or an autoantibody (A of Figure 2a) to an autoantigen according to one embodiment.

[0100] FIG. 2B is a schematic diagram illustrating differences in binding patterns between antibodies and autoantigens that occur depending on differences in binding orientations between antibodies and autoantigens (B of FIG. 2B) and binding orientations between autoantibodies and autoantigens (A of FIG. 2B) according to one embodiment.

[0101] Figure 2c is a schematic diagram illustrating a mechanism by which an antibody according to one embodiment inhibits the binding of an autoantigen and an autoantibody to form a cross-linking structure.

[0102] Figure 2d is a schematic diagram illustrating a mechanism by which an antibody inhibits internalization of an antigen according to one embodiment.

[0103] Figure 2e is a schematic diagram illustrating a mechanism by which an antibody according to one embodiment inhibits complement-dependent cytotoxicity by an autoantibody.

[0104] FIG. 2f is a schematic diagram illustrating a mechanism by which an antibody inhibits antibody-dependent cytotoxicity or antibody-dependent cellular phagocytosis according to one embodiment.

[0105] Referring to FIGS. 2A and 2B , in the antibody of one embodiment, the binding orientation of at least one monomer of the dimer of the dimeric antibody of one embodiment to the autoantigen may be different from the binding orientation of the autoantibody to the autoantigen. Accordingly, the binding of one monomer of the dimer to the first autoantigen may have an orientation that prevents the remaining monomer of the dimer from binding to a second autoantigen present on the cell surface (an autoantigen identical to the first autoantigen). For example, the binding orientation of the first half-antibody fragment or the second half-antibody fragment to the autoantigen may be different from the binding orientation of the autoantibody to the autoantigen.

[0106] Accordingly, the first half antibody fragment and / or the second half antibody fragment may bind to the same epitope as the epitope to which the autoantibody fragment binds or to an epitope located in close proximity to the same epitope, thereby inhibiting the autoantibody from binding to the autoantigen. For example, the first half antibody fragment and the second half antibody fragment may bind to the same epitope as the epitope to which the autoantibody fragment binds. Since the binding orientation of the first half antibody fragment (or the second half antibody fragment) to the first autoantigen is different from the binding orientation of the autoantibody to the autoantigen, the second half antibody fragment (or the first half antibody fragment) that does not bind to the autoantigen may not bind to the second autoantigen present on the cell surface.

[0107] That is, the antibody of one embodiment may have a conformation that makes it difficult or impossible to bind to multiple (e.g., two) complex autoantigens present on the cell surface.

[0108] Referring to FIG. 2C, an antibody of one embodiment can inhibit the cross-linking of multiple autoantigens by binding an autoantibody to an autoantigen. For example, an antibody of one embodiment may not cross-link multiple (e.g., two, three, four, or five or more) complex autoantigens present on a cell surface. Since an antibody of one embodiment can inhibit the binding of an autoantibody to an autoantigen to form a cross-linking structure even with a small dose, a preventive or therapeutic effect can be achieved with a low dose. That is, an antibody or competitive autoantibody inhibitor of one embodiment can have a low effective dose.

[0109] Referring to Figure 2d, it can be confirmed that even if the antibody of one embodiment binds to a complex of autoantigens present on the cell surface, internalization of the antigen is not induced because multiple autoantigens are not cross-linked. Therefore, the antibody of one embodiment can induce an immune response against the cell in which the antigen is present as the antigen is internalized, or can prevent or treat an immune disease caused by the loss of antigen function.

[0110] The antibody of one embodiment can prevent autoantigens from being cross-linked and internalized, and thus can exhibit excellent preventive or therapeutic effects on diseases associated with autoantigens densely formed in cells.

[0111] Referring to FIGS. 2E and 2F, the antibody of one embodiment, unlike the autoantibody, does not bind to an autoantigen to form a cross-linking structure. Therefore, complement activation is inhibited by preventing complement such as C1q from binding to the antibody. In addition, the antibody of one embodiment does not bind to FcγR and inhibits antigen binding of the autoantibody, thereby inhibiting ADCC and / or CDC through FcγR. Therefore, even if the competitive autoantibody inhibitor binds to a complex autoantigen present on the cell surface, ADCC and / or CDC may not be induced. Therefore, it is effective in the prevention or treatment of autoimmune diseases (e.g., myasthenia gravis and / or neuromyelitis optica), in which CDC and / or ADCC are one of the important etiologies.

[0112] In one embodiment, the antibody may lack effector functions of an intact antibody. For example, mutations may be introduced into the Fc region that reduce binding of the antibody to activating Fcγ receptors (FcγRs) and reduce Fc effector functions, such as C1q binding, complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC), or antibody-dependent phagocytosis (ADCP). Fc positions that can be mutated to reduce binding of the antibody to activating FcγRs and subsequently reduce effector function include positions 214, 233, 234, 235, 236, 237, 238, 265, 267, 268, 270, 295, 297, 309, 327, 328, 329, 330, 331, and 365. Exemplary mutations that can be made singly or in combination are the mutations K214T, E233P, L234V, L234A, deletion of G236, V234A, F234A, L235A, G237A, P238A, P238S, D265A, S267E, H268A, H268Q, Q268A, N297A, A327Q, P329A, D270A, Q295A, V309L, A327S, L328F, A330S and P331S in IgG1, IgG2, IgG3 or IgG4.Exemplary combination mutations resulting in antibodies with reduced ADCC include mutations L234A / L235A on IgG1, V234A / G237A / P238S / H268A / V309L / A330S / P331S on IgG2, F234A / L235A on IgG4, S228P / F234A / L235A on IgG4, N297A on all Ig isotypes, V234A / G237A on IgG2, K214T / E233P / L234V / L235A / G236-deletion / A327G / P331A / D365E / L358M on IgG1, H268Q / V309L / A330S / P331S on IgG2, S267E / L328F on IgG1, L234F / L235E / D265A on IgG1, L234A / L235A / G237A / P238S / H268A / A330S / P331S on IgG1, S228P / F234A / L235A / G237A / P238S on IgG4, and S228P / F234A / L235A / G236-deletion / G237A / P238S on IgG4. A hybrid IgG2 / 4 Fc domain, such as an Fc having residues 117-260 from IgG2 and residues 261-447 from IgG4, may also be used. An exemplary mutation resulting in an antibody with reduced CDC is the K322A mutation. The well-known S228P mutation in an IgG4 antibody may be made to improve IgG4 stability.

[0113] The antibody Fc region may comprise a mutated Fc region lacking effector function, such as an IgG1 sequence comprising the L234A / L235A amino acid substitution, the L234A / L235A / G237A amino acid substitution, and / or the K322A amino acid substitution. The inhibitor may also comprise an IgG2 or IgG3 having a substitution in the Fc region that eliminates an effector function, such as complement activation. Alternatively, the inhibitor may be an IgG4 antibody. The inhibitor may be optimized for antigen binding, for example, AChR or aquaporin-4 binding, by mutating some residues in the antigen binding region. The antibody may be humanized.

[0114] The antibody may comprise mutations in the Fc region that modulate half-life. Fc positions that may be mutated to modulate antibody half-life (e.g., binding to FcRn) include positions 250, 252, 253, 254, 256, 257, 307, 376, 380, 428, 434, and 435. Exemplary mutations that may be made singly or in combination are mutations T250Q, M252Y, I253A, S254T, T256E, P257I, T307A, D376V, E380A, M428L, H433K, N434S, N434A, N434H, N434F, H435A, and H435R. Exemplary single or combination mutations that can be made to reduce the half-life of an antibody are the mutations H435A, P257I / N434H, D376V / N434H, M252Y / S254T / T256E / H433K / N434F, T308P / N434A and H435R.

[0115] Hereinafter, antibodies according to one embodiment will be described with specific examples. The antibodies described in one embodiment below may be animal-derived antibodies, but the present invention may include humanized antibodies thereof.

[0116] The above antibodies may be mAb637-IgG1, a myasthenia gravis autoantibody, and mAb637 C and mAb637 B that competitively bind to the AChR antigen. mAb637 C comprises a heavy chain of SEQ ID NO: 19 and a light chain of SEQ ID NO: 20, and mAb637 B comprises a heavy chain of SEQ ID NO: 17 and a light chain of SEQ ID NO: 18, respectively.

[0117] The antibodies may be mAb192 C IgG2 / 4 and mAb192 B that competitively bind to the AChR antigen with the myasthenia gravis autoantibody mAb637-IgG1. mAb192 C IgG2 / 4 comprises a heavy chain of SEQ ID NO: 3 and a light chain of SEQ ID NO: 4. mAb192 B comprises a heavy chain of SEQ ID NO: 5 and a light chain of SEQ ID NO: 6, respectively. In addition, the antibodies may be competitive inhibitory recombinant antibodies mAb35 B and mAb35 C of the myasthenia gravis pathogenic autoantibody mAb35 that competitively bind to the AChR antigen with the mAb35. The recombinant antibody mAb35 B comprises a heavy chain of SEQ ID NO: 13 and a light chain of SEQ ID NO: 14, and the recombinant antibody mAb35 C comprises a heavy chain of SEQ ID NO: 15 and a light chain of SEQ ID NO: 16. Additionally, the antibody may be rAb53 C, which competitively binds to the neuromyelitis optica pathogenic autoantibody rAb58 and the aquaporin-4 antigen. rAb53 C comprises a heavy chain of SEQ ID NO: 9 and a light chain of SEQ ID NO: 10. The rAb53 autoantibody may have a heavy chain of SEQ ID NO: 47 and a light chain of SEQ ID NO: 48.

[0118] The above antibodies may be mAb637-IgG1, a myasthenia gravis autoantibody, and mAb637 C and mAb637 B that competitively bind to the AChR antigen. mAb637 C and mAb637 B comprise complementarity determining regions of SEQ ID NOs: 21, 22, 23, 24, 25, and 26.

[0119] V of the above mAb637 B and mAb637 C LThe VH comprises a light chain CDR1 of SEQ ID NO: 21, a light chain CDR2 of SEQ ID NO: 22, and a light chain CDR3 of SEQ ID NO: 23, and the VH comprises a heavy chain CDR1 of SEQ ID NO: 24, a heavy chain CDR2 (EDN) of SEQ ID NO: 25, and a heavy chain CDR3 of SEQ ID NO: 26.

[0120] The above antibodies may be mAb192 C IgG2 / 4 and mAb192 B, which competitively bind to the AChR antigen with the myasthenia gravis autoantibody mAb637-IgG1. mAb192 C IgG2 / 4 and mAb192 B comprise complementarity determining regions of SEQ ID NOs: 27, 28, 29, 30, 31, and 32.

[0121] V of the above mAb192 B and mAb192 C IgG2 / 4 L It comprises a light chain CDR1 of SEQ ID NO: 27, a light chain CDR2 of SEQ ID NO: 28 and a light chain CDR3 of SEQ ID NO: 29, and V H The antibody may comprise a heavy chain CDR1 of SEQ ID NO: 30, a heavy chain CDR2 of SEQ ID NO: 31, and a heavy chain CDR3 of SEQ ID NO: 32. In addition, the antibody may be a competitive inhibitory recombinant antibody mAb35 B and mAb35 C of myasthenia gravis pathogenic autoantibody mAb35 and mAb35 that competitively bind to the AChR antigen. The recombinant antibodies mAb35 B and mAb35 C comprise complementarity determining regions of SEQ ID NOs: 33, 34, 35, 36, 37, and 38.

[0122] V of the above mAb35 B and mAb35 C L It comprises a light chain CDR1 of SEQ ID NO: 33, a light chain CDR2 of SEQ ID NO: 34, and a light chain CDR3 of SEQ ID NO: 35, and V H may include a heavy chain CDR1 of SEQ ID NO: 36, a heavy chain CDR2 (KTN) of SEQ ID NO: 37, and a heavy chain CDR3 of SEQ ID NO: 38.

[0123] Additionally, the antibody may be rAb53 C, which competitively binds to the neuromyelitis optica pathogenic autoantibody rAb58 and the aquaporin-4 antigen. rAb53 C comprises complementarity determining regions of SEQ ID NOs: 39, 40, 41, 42, 43, and 44. The rAb53 autoantibody may have a heavy chain of SEQ ID NO: 47 and a light chain of SEQ ID NO: 48.

[0124] V of the above rAb53 C L It comprises a light chain CDR1 of SEQ ID NO: 39, a light chain CDR2 of SEQ ID NO: 40, and a light chain CDR3 of SEQ ID NO: 41, and V H may include a heavy chain CDR1 of SEQ ID NO: 42, a heavy chain CDR2 (GAS) of SEQ ID NO: 43, and a heavy chain CDR3 of SEQ ID NO: 44.

[0125] According to another aspect of the present invention, there is provided a pharmaceutical composition for use in preventing or treating a disease associated with a decrease in the function of an autoantigen, comprising the competitive autoantibody inhibitor as an active ingredient and a pharmaceutically acceptable carrier.

[0126] Pharmaceutically acceptable carriers enhance or stabilize the composition, or facilitate its manufacture. Such carriers include physiologically compatible solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like.

[0127] The composition of the present invention can be administered by various methods known in the art. The route and / or method of administration will vary depending on the desired result. Administration may be intravenous, intramuscular, intraperitoneal, or subcutaneous, or administered adjacent to the target site. The carrier may be suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion). The composition may be sterile and fluid. The composition may be in lyophilized form. The composition may include an isotonic agent, such as a sugar, a polyalcohol such as mannitol or sorbitol, and sodium chloride. The composition of the present invention may be prepared according to methods well known and commonly practiced in the art.

[0128] The dosage level of the active ingredient in the composition may be varied to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and route of administration without being toxic to the patient. The selected dosage level will depend on various pharmacodynamic factors, such as the particular composition of the present disclosure employed, or the route of administration, time of administration, rate of excretion, duration of treatment, other drugs, compounds, and / or substances used in combination, the age, sex, weight, condition, general health, and prior medical history of the patient to be treated, and other factors. As a non-limiting example, dosages range from about 0.0001 to 100 mg / kg body weight, more typically from 0.1 to 20 mg / kg body weight. An exemplary treatment regimen involves administration once weekly, once every two weeks, once monthly, or once every three to six months.

[0129] In the pharmaceutical composition of the present invention, the disease may be an autoimmune disease. The autoimmune disease may be pemphigus (pemphigus vulgaris, pemphigus foliaceus, or paraneoplastic pemphigus), Crohn's disease, idiopathic thrombocytopenic purpura (ITP), heparin-induced thrombocytopenia (HIT), thrombotic thrombocytopenic purpura (TTP), myasthenia gravis (MG), and chronic inflammatory demyelinating polyneuropathy (CIDP). Additional non-limiting autoimmune diseases include autoimmune thrombocytopenia, immune neutropenia, antihemophilic FVIII inhibitors, antiphospholipid syndrome, Kawasaki syndrome, ANCA-associated disease, polymyositis, bullous pemphigoid, multiple sclerosis (MS), Guillain-Barré syndrome, chronic polyneuropathy, ulcerative colitis, diabetes mellitus, autoimmune thyroiditis, Graves' ophthalmopathy, rheumatoid arthritis, ulcerative colitis, primary sclerosing cholangitis, systemic lupus erythematosus (SLE), autoimmune encephalomyelitis, Hashimoto's thyroiditis, Goodpasture's syndrome, autoimmune hemolytic anemia, scleroderma with anticollagen antibodies, mixed connective tissue disease, pernicious anemia, idiopathic Addison's disease, autoimmune-associated infertility, glomerulonephritis (e.g., crescentic glomerulonephritis, proliferative glomerulonephritis), insulin resistance, and Autoimmune diabetes (type 1 diabetes, insulin-dependent diabetes). Autoimmune diseases have also been recognized to include atherosclerosis and Alzheimer's disease. In another embodiment, the autoimmune disease is hepatitis, autoimmune hemophilia, autoimmune lymphoproliferative syndrome (ALPS), autoimmune uveitis, glomerulonephritis, agammaglobulinemia, alopecia areata, amyloidosis, ankylosing spondylitis, autoimmune angioedema, autoimmune aplastic anemia, autoimmune dysautonomia, autoimmune hyperlipidemia, autoimmune immunodeficiency, autoimmune inner ear disease (AIED), autoimmune myocarditis, autoimmune pancreatitis, autoimmune retinopathy, autoimmune urticaria, autoimmune urticarial neuropathy, autoimmune axonal neuropathy, Ballot disease, Bennett's disease, Castleman's disease, celiac disease, Chagas' disease, chronic relapsing multiple myelitis (CRMO), Churg-Strauss syndrome,Cicatricial pemphigoid, benign mucosal pemphigoid, Cogan's syndrome, cold agglutinin disease, Coxsackie myocarditis, CREST disease, essential mixed cryoglobulinemia, dermatitis herpetiformis, dermatomyositis, Devic's disease (neuromyelitis optica), dilated cardiomyopathy, discoid lupus, Dressler's syndrome, endometriosis, eosinophilic angiocentric fibrosis, eosinophilic fasciitis, erythema nodosum, Evans syndrome, fibrosing alveolitis, giant cell arteritis (temporal arteritis), Hashimoto's encephalitis, Henoch-Schonlein purpura, herpes gestationis, idiopathic hypocomplementary ureteronephritis, multiple myeloma, multiple motor neuropathy, NMDA receptor antibody encephalitis, IgG4-related disease, IgG4-related sclerosing disease, inflammatory aortic aneurysm, inflammatory pseudotumor, inclusion body myositis, interstitial cystitis, juvenile arthritis, Kuttner's tumor, Lambert-Eaton syndrome, leukocytoclastic vasculitis, lichen planus, lichen sclerosus, conjunctivitis, linear IgA disease (LAD), Lyme disease, chronic, mediastinal fibrosis, Meniere's disease, microscopic polyangiitis, Mikulicz syndrome, Mooren's ulcer, Mucha-Habermann disease, multiple sclerosis, narcolepsy, optic neuritis, Ormond disease (retroperitoneal fibrosis), palindromic rheumatism, PANDAS (Pediatric autoimmune neuropsychiatric disorders associated with streptococci), paraneoplastic cerebellar degeneration, paraproteinemic polyneuropathy, paroxysmal nocturnal hemoglobinuria (PNH), Parry-Romberg syndrome, Parsonage-Turner syndrome, arthritis, periareolaritis, peripheral neuropathy, perivenous encephalomyelitis, POEMS syndrome, polyarteritis nodosa, types I, II, & III autoimmune polyglandular syndromes, Polymyalgia rheumatica, postpericardiotomy syndrome, progesterone dermatitis, primary biliary cirrhosis, psoriasis, psoriatic arthritis, idiopathic pulmonary fibrosis, pyoderma gangrenosum, pure red cell aplasia, Raynaud's phenomenon, reflex sympathetic dystrophy, Reiter's syndrome, relapsing polychondritis, restless legs syndrome, rheumatic fever, Riede's thyroiditis, sarcoidosis, Schmidt syndrome, scleritis, Sjögren's syndrome, sperm and testicular autoimmunity, stiff person syndrome, subacute bacterial endocarditis (SBE), Susac's syndrome, sympathetic ophthalmia,It may be selected from the group consisting of Takayasu arteritis, Tolosa-Hunt syndrome, transverse myelitis, undifferentiated connective tissue disease (UCTD), bullous dermatosis, vitiligo, Rasmussen's encephalitis, and Waldenström macroglobulinemia.

[0130] For example, the diseases include pemphigus vulgaris, pemphigus leafhopper, neoplastic pemphigus, rheumatoid arthritis, systemic lupus erythematosus, Crohn's disease, idiopathic thrombocytopenic purpura (ITP), heparin-induced thrombocytopenia (HIT), thrombotic thrombocytopenic purpura (TTP), autoimmune hemolytic anemia (AIHA), myasthenia gravis (MG), chronic inflammatory demyelinating polyneuropathy (CIDP), multiple motor neuropathy, neuromyelitis optica, autoimmune thrombocytopenia, immune neutropenia, antihemophilic FVIII inhibitors, antiphospholipid syndrome, Kawasaki syndrome, ANCA-related diseases, polymyositis, dermatomyositis, bullous pemphigoid, multiple sclerosis (MS), Guillain-Barré syndrome, chronic polyneuropathy, ulcerative colitis, and true It may be selected from the group consisting of diabetes mellitus, autoimmune thyroiditis, Graves' ophthalmopathy, autoimmune urticaria, vasculitis and Rasmussen's encephalitis.

[0131] According to another aspect of the present invention, a method for preventing or treating a disease associated with a decrease in the function of an autoantigen in a subject is provided, comprising administering to the subject an amount of the antibody effective to prevent or treat the disease associated with a decrease in the function of the autoantigen.

[0132] According to another aspect of the present invention, the present invention provides a use for preventing or treating a disease associated with a decrease in the function of an autoantigen of the antibody.

[0133] According to another aspect of the present invention, the use of the antibody is provided in the manufacture of a medicament for preventing or treating a disease associated with a decrease in the function of an autoantigen.

[0134] In this specification, the description of the antibody described above may be equally applied to the pharmaceutical composition, the method for prevention or treatment, the use for prevention or treatment, and the use in the manufacture of a medicament for prevention or treatment.

[0135] In the above method, the disease may be an autoimmune disease, and the disease may be pemphigus (pemphigus vulgaris, pemphigus foliaceus or paraneoplastic pemphigus), Crohn's disease, idiopathic thrombocytopenic purpura (ITP), heparin-induced thrombocytopenia (HIT), thrombotic thrombocytopenic purpura (TTP), myasthenia gravis (MG) and chronic inflammatory demyelinating polyneuropathy (CIDP). Additional non-limiting autoimmune diseases include autoimmune thrombocytopenia, immune neutropenia, antihemophilic FVIII inhibitors, antiphospholipid syndrome, Kawasaki syndrome, ANCA-associated disease, polymyositis, bullous pemphigoid, multiple sclerosis (MS), Guillain-Barré syndrome, chronic polyneuropathy, ulcerative colitis, diabetes mellitus, autoimmune thyroiditis, Graves' ophthalmopathy, rheumatoid arthritis, ulcerative colitis, primary sclerosing cholangitis, systemic lupus erythematosus (SLE), autoimmune encephalomyelitis, Hashimoto's thyroiditis, Goodpasture's syndrome, autoimmune hemolytic anemia, scleroderma with anticollagen antibodies, mixed connective tissue disease, pernicious anemia, idiopathic Addison's disease, autoimmune-associated infertility, glomerulonephritis (e.g., crescentic glomerulonephritis, proliferative glomerulonephritis), insulin resistance, and Autoimmune diabetes (type 1 diabetes, insulin-dependent diabetes). Autoimmune diseases have also been recognized to include atherosclerosis and Alzheimer's disease. In another embodiment, the autoimmune disease is hepatitis, autoimmune hemophilia, autoimmune lymphoproliferative syndrome (ALPS), autoimmune uveitis, glomerulonephritis, agammaglobulinemia, alopecia areata, amyloidosis, ankylosing spondylitis, autoimmune angioedema, autoimmune aplastic anemia, autoimmune dysautonomia, autoimmune hyperlipidemia, autoimmune immunodeficiency, autoimmune inner ear disease (AIED), autoimmune myocarditis, autoimmune pancreatitis, autoimmune retinopathy, autoimmune urticaria, autoimmune urticarial neuropathy, autoimmune axonal neuropathy, Ballot disease, Bennett's disease, Castleman's disease, celiac disease, Chagas' disease, chronic relapsing multiple myelitis (CRMO), Churg-Strauss syndrome, cicatricial pemphigoid,Benign mucosal pemphigoid, Cogan's syndrome, cold agglutinin disease, Coxsackie myocarditis, CREST disease, essential mixed cryoglobulinemia, dermatitis herpetiformis, dermatomyositis, Devic's disease (neuromyelitis optica), dilated cardiomyopathy, discoid lupus, Dressler's syndrome, endometriosis, eosinophilic angiocentric fibrosis, eosinophilic fasciitis, erythema nodosum, Evans syndrome, fibrosing alveolitis, giant cell arteritis (temporal arteritis), Hashimoto's encephalitis, Henoch-Schonlein purpura, herpes gestationis, idiopathic hypocomplementary ureteronephritis, multiple myeloma, multiple motor neuropathy, NMDA receptor antibody encephalitis, IgG4-related disease, IgG4-related sclerosing disease, inflammatory aortic aneurysm, inflammatory pseudotumor, inclusion body myositis, interstitial cystitis, juvenile arthritis, Kuttner's tumor, Lambert-Eaton syndrome, leukocytoclastic vasculitis, lichen planus, lichen sclerosus, conjunctivitis, linear IgA disease (LAD), Lyme disease, chronic, mediastinal fibrosis, Meniere's disease, microscopic polyangiitis, Mikulicz syndrome, Mooren's ulcer, Mucha-Habermann disease, multiple sclerosis, narcolepsy, optic neuritis, Ormond disease (retroperitoneal fibrosis), palindromic rheumatism, PANDAS (pediatric autoimmune neuropsychiatric disorders associated with streptococci), paraneoplastic cerebellar degeneration, paraproteinemic polyneuropathy, paroxysmal nocturnal hemoglobinuria (PNH), Parry-Romberg syndrome, Parsonage-Turner syndrome, arthritis, periareolitis, peripheral neuropathy, perivenous encephalomyelitis, POEMS syndrome, polyarteritis nodosa, types I, II, & III autoimmune polyglandular syndromes, rheumatoid Polymyalgia, postpericardiotomy syndrome, progesterone dermatitis, primary biliary cirrhosis, psoriasis, psoriatic arthritis, idiopathic pulmonary fibrosis, pyoderma gangrenosum, pure red cell aplasia, Raynaud's phenomenon, reflex sympathetic dystrophy, Reiter's syndrome, relapsing polychondritis, restless legs syndrome, rheumatic fever, Riede's thyroiditis, sarcoidosis, Schmidt syndrome, scleritis, Sjögren's syndrome, sperm and testicular autoimmunity, stiff person syndrome, subacute bacterial endocarditis (SBE), Susac's syndrome, sympathetic ophthalmia, Takayasu's arteritis,It may be selected from the group consisting of Tolosa-Hunt syndrome, transverse myelitis, undifferentiated connective tissue disease (UCTD), bullous dermatosis, vitiligo, Rasmussen's encephalitis, and Waldenström macroglobulinemia.

[0136] For example, the diseases include pemphigus vulgaris, pemphigus leafhopper, neoplastic pemphigus, rheumatoid arthritis, systemic lupus erythematosus, Crohn's disease, idiopathic thrombocytopenic purpura (ITP), heparin-induced thrombocytopenia (HIT), thrombotic thrombocytopenic purpura (TTP), autoimmune hemolytic anemia (AIHA), myasthenia gravis (MG), chronic inflammatory demyelinating polyneuropathy (CIDP), multiple motor neuropathy, neuromyelitis optica, autoimmune thrombocytopenia, immune neutropenia, antihemophilic FVIII inhibitors, antiphospholipid syndrome, Kawasaki syndrome, ANCA-related diseases, polymyositis, dermatomyositis, bullous pemphigoid, multiple sclerosis (MS), Guillain-Barré syndrome, chronic polyneuropathy, ulcerative colitis, and true It may be selected from the group consisting of diabetes mellitus, autoimmune thyroiditis, Graves' ophthalmopathy, autoimmune urticaria, vasculitis and Rasmussen's encephalitis.

[0137] The above object may be an animal, the animal may be a mammal, and the mammal may be a human, a dog, a cat, a cow, a pig, a monkey, a rat or a horse.

[0138] The above administration may be administered by various methods known in the relevant art. The route and / or method of administration will vary depending on the desired result. Administration may be intravenous, intramuscular, intraperitoneal, or subcutaneous, or administered adjacent to the target site. The administration may be by administering a composition comprising the antibody described above. A pharmaceutically acceptable carrier may be suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion). The composition may be sterile and fluid. The composition may be in lyophilized form. Isotonic agents, such as sugars, sugar alcohols such as mannitol or sorbitol, and sodium chloride may be included in the composition.

[0139] In the above method, the dosage level of the active ingredient can be varied to obtain an amount of the active ingredient that is effective to achieve the desired therapeutic response for a particular patient, composition, and route of administration without being toxic to the patient. The selected dosage level will depend on various pharmacodynamic factors, such as the particular active ingredient of the present disclosure employed or the composition comprising the same, or the route of administration, time of administration, rate of excretion, duration of treatment, other drugs, compounds, and / or substances used in combination, the age, sex, weight, condition, general health, and prior medical history of the patient to be treated, and other factors. As a non-limiting example, the dosage ranges from about 0.0001 to 100 mg / kg body weight, more typically from 0.1 to 20 mg / kg body weight. An exemplary treatment regimen may involve administration once weekly, once every two weeks, once monthly, or once every three to six months.

[0140] According to another aspect of the present invention, there is provided a use of the antibody in the manufacture of a pharmaceutical composition for preventing or treating a disease associated with a decrease in the function of an autoantigen.

[0141] The present invention will be described in more detail below through examples. However, these examples are intended to exemplify the present invention and the scope of the present invention is not limited to these examples.

[0142] Example 1: Competitive autoantibodies and their efficacy

[0143] 1-1: Autoantibody mAb637 antibody and its competitive autoantibody inhibitor antibody

[0144] The antibody used was a monoclonal mAb637, which induces myasthenia gravis (MG) in humans. mAb637 is an AChR-specific monoclonal autoantibody isolated from a human MG patient. The mAb637 antibody, also known as IgG1-637, is a G1 class antibody with effector function. It binds to the MIR of the AChR alpha subunit. In this experiment, the mAb637 antibody was produced in-house by cloning DNA based on the published sequence, transiently expressing it in ExpiCHO cells, and purifying it using Mabselect Sure, a column used to purify or separate biopharmaceuticals. The mAb637 antibody induces MG when passively administered to a subject. mAb637 comprises a heavy chain having the amino acid sequence of SEQ ID NO: 1 and a light chain having the amino acid sequence of SEQ ID NO: 2. V of mAb637 H and V L Each comprises an amino acid sequence of positions 1 to 126 of SEQ ID NO: 1 and an amino acid sequence of positions 1 to 112 of SEQ ID NO: 2.

[0145] 1-2: Recombinant antibody according to one embodiment for competitively inhibiting myasthenia gravis autoantibodies for use in in vitro CDC assay and competition assay

[0146] The above mAb637 C: VL -SS-CH1-hinge-CH2-CH3(IgG2 / 4) + V H -kappa structure. That is, the structure of mAb637 C is V of mAb637, a human myasthenia gravis-inducing antibody. L -CL structure light chain (SEQ ID NO: 2) and V H -V in the heavy chain (SEQ ID NO: 1) of the CH1-hinge-CH2-CH3 structure L and V H exchange with each other, and as a result V H Go V L V in the heavy chain substituted with L The CH1-hinge-CH2-CH3 connection site is connected with a linker -SS-, and the CH1, CH2, and CH3 regions are changed from IgG1 to IgG2 / 4. Here, CL is a kappa chain. The heavy chain and light chain of the mAb637 C have the amino acid sequence of SEQ ID NO: 19 and the amino acid sequence of SEQ ID NO: 20, respectively.

[0147] mAb637 B: V L -kappa-hinge-CH2+CH3(IgG2 / 4) + V H -It has a CH1(IgG2 / 4) structure.

[0148] That is, the structure of mAb637 B is V of mAb637, a human myasthenia gravis-inducing antibody. L -CL structure light chain (SEQ ID NO: 2) and V H In the heavy chain of the -CH1-hinge-CH2-CH3 structure (SEQ ID NO: 1), V L -CL and V H -CH1 is exchanged, and the resulting V is obtained L In the heavy chain of -CL-hinge-CH2-CH3 and the light chain of VH-CH1, the CH1 and CH2-CH3 regions were each modified to the IgG2 / 4 type. The heavy and light chains of mAb637 B have the amino acid sequences of SEQ ID NO. 17 and SEQ ID NO. 18, respectively.

[0149] mAb192 C IgG2 / 4: VL -SS-IgG2 / 4 + V H -kappa structure. That is, the structure of mAb192 C is V of mAb192, a rat-derived myasthenia gravis-inducing antibody. L -CL structure light chain (SEQ ID NO: 46) and V H In the heavy chain of the -CH1-hinge-CH2-CH3 structure (SEQ ID NO: 45), VL and V H exchange, and as a result V H Go V L In the heavy chain substituted with , the VL and CH1-hinge-CH2-CH3 junction regions are connected with a linker -SS-, and the CH1, CH2, and CH3 regions are changed from G1 to IgG2 / 4. Here, CL is a kappa chain. The heavy and light chains of mAb192 C IgG2 / 4 have the amino acid sequences of SEQ ID NO: 3 and SEQ ID NO: 4, respectively.

[0150] mAb192 B: V L -kappa-hinge-CH2+CH3(IgG2 / 4) + V H -It has a CH1(IgG2 / 4) structure.

[0151] That is, the structure of mAb192 B is V of mAb192, a rat-derived myasthenia gravis-inducing antibody. L -CL structure light chain (SEQ ID NO: 46) and V H In the heavy chain (SEQ ID NO: 45) of the -CH1-hinge-CH2-CH3 structure, VL-CL and V H -CH1 is exchanged, and the resulting V is obtained L -CL-hinge-CH2-CH3 heavy chain and V H In the light chain of -CH1, the CH1 and CH2-CH3 regions were each modified to the IgG2 / 4 type. The heavy and light chains of mAb192 B have the amino acid sequences of SEQ ID NO: 5 and SEQ ID NO: 6, respectively.

[0152] mAb192 IgG2 / 4: It has a human IgG2 / 4 structure. That is, the CH1, CH2, and CH3 regions of mAb192, a rat-derived myasthenia gravis-inducing antibody, are changed from IgG1 to IgG2 / 4. The heavy chain of mAb192 IgG2 / 4 has the amino acid sequence of SEQ ID NO: 65, and the light chain has the amino acid sequence of SEQ ID NO: 66.

[0153] mAb192 C IgG1: V L -SS-IgG1 + V H -kappa structure, i.e., V of mAb192, a rat-derived myasthenia gravis-inducing antibody L -CL structure light chain (SEQ ID NO: 46) and V H In the heavy chain of the -CH1-hinge-CH2-CH3 structure (SEQ ID NO: 45), V L and V H , and as a result VH becomes V L V in the heavy chain substituted with L The connecting portions of CH1-hinge-CH2-CH3 are connected with a linker -SS-. Here, the CL is a kappa chain. The heavy chain of the mAb192 C IgG1 has an amino acid sequence of SEQ ID NO: 69, and the light chain has an amino acid sequence of SEQ ID NO: 70.

[0154] mAb192 C IgG4: V L -SS-IgG4 + V H -kappa structure. That is, the structure of mAb192 C is V of mAb192, a rat-derived myasthenia gravis-inducing antibody. L -CL structure light chain (SEQ ID NO: 46) and V H In the heavy chain of the -CH1-hinge-CH2-CH3 structure (SEQ ID NO: 45), V L and V H exchange, and as a result V H Go V L V in the heavy chain substituted with LThe CH1-hinge-CH2-CH3 junction region is connected by a linker -SS-, and the CH1, CH2, and CH3 regions are changed to IgG4 by introducing mutations that inhibit Fab arm exchange in IgG1. Here, CL is a kappa chain. The heavy chain of mAb192 C IgG4 has the amino acid sequence of SEQ ID NO: 71, and the light chain has the amino acid sequence of SEQ ID NO: 72.

[0155] Humanized antibody of mAb35: Human IgG1 The mAb35 antibody is an autoantibody that binds to the alpha subunit of rat AChR and comprises a heavy chain having an amino acid sequence of SEQ ID NO: 11 and a light chain having an amino acid sequence of SEQ ID NO: 12. mAb35 is an antibody that also binds to human AChR, and to create a humanized variant, the CH1, CH2, and CH3 regions were changed from rat IgG to human IgG1. The heavy chain of the mAb35 humanized antibody has an amino acid sequence of SEQ ID NO: 73, and the light chain has an amino acid sequence of SEQ ID NO: 74.

[0156] In vitro neuromyelitis optica (NMO) pathogenic autoantibodies

[0157] rAb58: V H -IgG1 + V L -kappa structure. The above rAb53 and rAb58 antibodies are autoantibodies against aquaporin-4 obtained from plasma blast clones isolated from the cerebrospinal fluid of aquaporin-4 antibody-positive patients. They bind to the aquaporin-4 M23 isoform and cause antibody-dependent cytotoxicity or complement-dependent cytotoxicity. rAb58 is an antibody against patient-derived aquaporin-4. The rAb58 antibody comprises a heavy chain having an amino acid sequence of SEQ ID NO: 7 and a light chain having an amino acid sequence of SEQ ID NO: 8. The VH and V of rAb58 L Each has an amino acid sequence of positions 1 to 115 of SEQ ID NO: 7 and an amino acid sequence of positions 1 to 110 of SEQ ID NO: 8.

[0158] 1-3: One embodiment of a recombinant antibody competitively inhibiting neuromyelitis optica (NMO) autoantibody rAb58 for use in in vitro ADCC assays.

[0159] The rAb53 antibody has a heavy chain of SEQ ID NO: 47 and a light chain of SEQ ID NO: 48. rAb53 C: V L -SS-IgG2 / 4 + V H -kappa structure. rAb53 C antibody is V of rAb53 antibody. H and VL are exchanged with each other, and V is obtained from the obtained heavy chain L The CH1-hinge-CH2-CH3 connection region is connected with a linker -SS-, and the CH1, CH2, and CH3 regions are modified from IgG1 to IgG2 / G4. The rAb53 C is a recombinant antibody that competitively binds to aquaporin-4 with the autoantibody rAb58 or rAb53. The rAb53 C antibody comprises a heavy chain having an amino acid sequence of SEQ ID NO: 9 and a light chain having an amino acid sequence of SEQ ID NO: 10. The V of rAb53 C H and V L Each has an amino acid sequence of positions 1 to 124 of SEQ ID NO: 9 and an amino acid sequence of positions 1 to 108 of SEQ ID NO: 10.

[0160] 1-4: In vivo myasthenia gravis pathogenic autoantibodies

[0161] mAb35 antibody: rat IgG

[0162] The mAb35 antibody is an autoantibody that binds to the alpha subunit of the rat AChR. The mAb35 antibody comprises a heavy chain having the amino acid sequence of SEQ ID NO: 11 and a light chain having the amino acid sequence of SEQ ID NO: 12. The V of the mAb35 antibody H and V L Each has an amino acid sequence of positions 1 to 121 of SEQ ID NO: 11 and an amino acid sequence of positions 1 to 106 of SEQ ID NO: 12.

[0163] 1-5: Recombinant antibodies that competitively bind to in vivo mAb35 and autoantigens

[0164] Competitive recombinant antibody mAb35 B: V L -kappa-CH2-CH3 + V H -CH1

[0165] mAb35 B is the heavy chain V of the mAb35 antibody. H -CH1 and light chain V L -CL is an IgG1 antibody that is exchanged with each other. The mAb35 B has heavy and light chains having amino acid sequences of SEQ ID NOs: 13 and 14, respectively. The V of the mAb35 B antibody H and V L Each has an amino acid sequence of positions 1 to 122 of SEQ ID NO: 13 and an amino acid sequence of positions 1 to 105 of SEQ ID NO: 14.

[0166] Competitive recombinant antibody mAb35 C: V of mAb35 L -SS-CH1-CH2-CH3(IgG2 / 4) + VH-kappa

[0167] mAb35 C is the heavy chain V of the mAb35 antibody. H -CH1 and light chain V L -CL is exchanged with each other, and V is obtained from the obtained heavy chain L The connecting portion of -CH1-hinge-CH2-CH3 is connected with a linker -SS-, and the CH1, CH2, and CH3 regions are modified from IgG1 to IgG2 / G4. The mAb35 C has heavy and light chains having amino acid sequences of SEQ ID NOs: 15 and 16, respectively. The V of the mAb35 C antibody H and V L Each has an amino acid sequence of positions 1 to 122 of SEQ ID NO: 15 and an amino acid sequence of positions 1 to 105 of SEQ ID NO: 16.

[0168] Example 2: Competition assay

[0169] For competitive analysis, TE671 cells, which are human medulloblastoma cells expressing fetal acetylcholine receptors (AChRs) on their cell membranes and are used to verify antibodies that induce myasthenia gravis, were prepared by detaching them from culture dishes using 10 mM EDTA. Afterwards, DMEM containing FBS was used as the culture medium, and the detached cells were prepared in PBS. Then, the prepared cells were seeded in a 96-well plate at a density of 1 x 10 5 Cells were seeded so that they were 10 cells / well. The therapeutic antibody, which is a competitive autoantibody inhibitor, was serially diluted to prepare various concentrations of the therapeutic antibody, which was added to the wells and incubated at 4°C for 20 minutes to allow binding to the antigen. Next, biotinylated-mAb637 (Biolegend) was added to the wells without a washing step to 8 μg / ml and incubated at 4°C for 20 minutes to allow competition between the therapeutic antibody and the biotinylated-mAb637 antibody for the autoantigen.

[0170] Streptavidin-Allophycocyanin (SA-APC) was diluted 1:1000 in FACS buffer, added to the wells, and incubated at 4°C for 20 minutes to allow reaction with the biotinylated-mAb637 antibody. After washing the wells, fluorescence-activated cell sorting (FACS) analysis was performed on the cells to analyze the degree to which the therapeutic antibody was competed with mAb637.

[0171] Example 3: Complement-dependent cytotoxicity (CDC) assay

[0172] The cells used for the complement-dependent cytotoxicity assay were a recombinant 293T cell line. The recombinant cell line was created by introducing a gene into the genome of the 293T cell line using a lentiviral system, thereby allowing the expression of the extracellular domain of the AChR. Specifically, the 293T cell line was recombined to co-express four types of AChR (A, B, D, and E) by introducing four types of DNA to express AChR.

[0173] Additionally, the CD45, CD55, and CD59 genes were knocked out of the obtained 293T cells using the CRISPR / Cas9 system. The CRISPR / Cas9 system uses gRNA to specifically target genomic DNA, creating nicks that induce double-strand breaks, which are then repaired by the cell's DNA repair system with some nucleotides missing. This process causes frame shifts and other mutations, knocking out normal gene expression.

[0174] The above three proteins are representative complement inhibitors. Accordingly, recombinant cells were obtained in which the AChR-ECM gene was expressed on the cell surface and the three complement inhibitory protein genes were knocked out. The AChR is a nicotinic acetylcholine receptor type AChR, which recognizes acetylcholine, a neurotransmitter secreted from nerve cells at the neuromuscular junction, and functions to open calcium channels to receive signals for muscle contraction. Next, the recombinant cells were detached from the culture dish using Trypsin-EDTA in DMEM medium (Welgene, cat. no. LB001-02), and then 1 x 10 per well were seeded. 5The cells were added to the wells of a 48-well plate at a concentration of 100 μg / ml. Cells were cultured in DMEM medium containing AOPI, glutamine, and FBS. AOPI was used to measure cell viability during cell culture. Normal human serum (10%) 10% (v / v) (Innovative research, CISER10ML-31004) and antibodies were added at the indicated concentrations to the wells to which the recombinant cells were added. The plates were incubated at 37°C for 6 hours to induce CDC. The serum is for providing complement. The antibodies are autoantibodies that cause autoimmune diseases (e.g., mAb35) and / or their competitively binding recombinant antibodies.

[0175] Next, anti-C5b-9 antibody (Abcam, ab66768) was diluted 1:200 in FACs buffer (2% FBS in PBS), added to each well, and incubated at 4°C for 15 minutes. After washing each well with mouse anti-C5b-9 antibody, secondary antibody anti-mouse IgG2A Alexa647 (R&D SYSTEMS, IC003R) and eBioscience™ Fixable Viability Dye eFluor™ 450 (FVD450) (Invitrogen, 65-0863-14) for apoptotic cell staining were diluted 1:100 in PBS buffer, added to each well, and incubated at 4°C for 15 minutes. After incubation, the cells were analyzed by flow cytometry to measure the percentage of C5b-9-positive cells and the percentage of apoptotic cells. Flow cytometry was performed by reading the fluorescence signal from cells diluted in FACs buffer using FACs analysis.

[0176] Example 4: Antibody internalization assay

[0177] The cells for antibody internalization analysis were recombinant 293T cell lines. The recombinant cell line was introduced into the genome of the 293T cell line using a lentiviral system, and the extracellular domain of AChR was expressed. Specifically, the 293T cell line was recombined to co-express four types of AChR (A, B, D, and E) by introducing four types of DNA to express AChR. The recombinant cells were detached from a culture dish using Trypsin-EDTA in DMEM medium (Welgene, cat. no. LB001-02), and then seeded at 1x10 per well. 4 The cells were added to the wells of a 96-well plate at a concentration of 100 μg / ml. Cell culture was performed in DMEM medium containing AOPI, glutamine, and FBS. AOPI was used to measure cell viability during cell culture.

[0178] After that, in order to confirm the internalization of the antibody by fluorescence, Incucyte Human Fabfluor-pH Red (STORIUS, Cat. no. 4722), which emits red fluorescence when the Fc portion of the antibody is internalized into cells and the pH is lowered, was mixed in a molar ratio of 1:2 and labeled at 37°C for 15 minutes. Then, 4 μg / ml of Fabfluor-labeled competitive recombinant antibody mAb192 C IgG2 / 4 and its variants or 4 μg / ml of Fabfluor-labeled humanized mAb35 antibody and serial dilutions of various concentrations of mAb192 C IgG2 / 4 and its variants were added to the wells to which recombinant cells were added, and incubated at 37°C for 24 hours, and the internalization of the antibody was measured by fluorescence at 30-minute intervals using an Incucyte device.

[0179] Example 5: In vivo efficacy test

[0180] Myasthenia gravis was induced in 4-week-old Lewis rats, which are used as an autoimmune disease model, by intraperitoneal administration of 1.5 mg / kg mAb35 antibody mixed in PBS. Lewis rats are a species derived from the Wistar rat, have white fur, and are susceptible to obesity induced by high-fat diets and autoimmune myocarditis. Lewis rats are widely used as models for multiple sclerosis and autoimmune diseases, and are the most commonly used species for inducing myasthenia gravis. The degree of myasthenia gravis induction was measured by adding body weight and clinical score. mAb35 antibody is an autoantibody against rat AChR of rat origin and is a pathogenic antibody that induces myasthenia gravis when passively administered to rats. Individuals administered the mAb35 antibody were observed for symptoms, and the test was conducted for up to 48 hours to obtain experimental rats induced with myasthenia gravis.

[0181] The clinical scores were awarded as follows: After performing the exercise of grasping the wire mesh of the cage several times, the condition after the exercise was checked and a total of 5-level scores were assigned. Grade 0: Normal muscle activity after the exercise. Grade 1: After the exercise, the subject touches the floor with the chin, cannot lift the head, bends the waist, and has reduced mobility. Grade 2: Rests without moving after the exercise. Grade 3: Signs of severe muscle weakness (paralysis, loss of grasping ability) and dyspnea during the exercise. Grade 4: Death.

[0182] To confirm the therapeutic and preventive effects of the antibodies of the present invention on autoimmune diseases, each therapeutic antibody was administered intravenously before administration of the mAb35 antibody, i.e., before induction of myasthenia gravis. Typically, clinical symptoms of myasthenia gravis appear 24 hours after administration of an autoantibody such as the mAb35 antibody. To confirm its preventive as well as therapeutic use, the antibody was administered before mAb35. The therapeutic antibody was administered once or four times repeatedly, depending on the experimental configuration. Blood samples were collected by jugular vein sampling before administration of the therapeutic antibody or the disease-causing antibody mAb35 and at regular intervals after antibody administration.

[0183] The collected blood was coagulated, and the serum was separated. The concentration of therapeutic or pathogenic antibodies in the serum was measured using ELISA. Furthermore, the rat tibialis anterior (TA) muscle was isolated and imaged using ELISA and immunostaining. This was done to determine whether the antibody binds to rat muscle AChR and to determine the antibody concentration in the muscle.

[0184] Example 6: ELISA for AChR

[0185] The AChR used in the present invention was derived from the Pacific electric ray (Torpedo californica). Specifically, an AChR-enriched membrane fraction obtained from the electric organ of Torpedo californica was added to each well of an ELISA plate at 5 to 10 μg / ml PBS and incubated overnight at 4°C to coat the wells with AChR. After washing the wells three times with washing buffer, a buffer containing 2% BSA was added and incubated at room temperature for 1 hour to block the surface. Thereafter, the sample was diluted to an appropriate concentration, added to the well, and incubated at 37°C for 2 hours. Human serum or rat tibialis anterior muscle lysate (TA lysate) was used as the sample. After washing the wells five times with washing buffer, anti-rat HRP or anti-human HRP antibodies were added at a dilution of 1:1000 to 1:5000 and incubated at 37°C for 1 hour. After washing five times with washing buffer, TMB substrate was added and incubated for 15 minutes. The reaction was stopped by adding 2 N H2SO4 and the absorbance was measured at 450 nm. HRP stands for horse radish peroxidase and TMB stands for 3,3',5,5'-tetramethylbenzidine.

[0186] Example 7: In vitro efficacy against myasthenia gravis and neuromyelitis optica

[0187] 7-1: Competitive Analysis

[0188] The present inventors analyzed the competitive binding of autoantibodies and recombinant antibodies to autoantigens. Specifically, the competitive binding of mAb637, an autoantibody to autoantigen AChR, and its competitive recombinant antibodies mAb637 B and mAb637 C was analyzed. The autoantibody is mAb637, which induces myasthenia gravis in humans, and the recombinant antibodies are mAb637 B, mAb637 C, and mAb192 C IgG2 / 4, and the autoantigen is AChR. As shown in Figure 3, mAb637 B binds to the V heavy chain of mAb637. H -CH1 and V of light chain L -kappa is a structure in which the two are exchanged (i.e., V L -kappa-CH2-CH3 heavy chain structure and V H -CH1 light chain structure), and mAb637 C is V of the heavy chain in mAb637. H and V of light chain L These interchanged structures (i.e., V L -CH1-CH2-CH3 heavy chain structure and V H-kappa light chain structure). Specifically, biotinylated pathogenic antibodies and native recombinant antibodies were used, and pathogenic antibodies were detected with Alexa647 conjugated-streptavidin. After analyzing the signal intensity through fluorescence-activated cell sorters (FACs), the signal intensity when the pathogenic antibody was 100% bound was set to 100, and the signal intensity when the candidate antibody was 100% bound was replaced with 0, and the degree of signal decrease when the binding ability of the pathogenic antibody was inhibited by the candidate antibody was observed. Specifically, the candidate antibodies were diluted by concentration and the change in signal according to the candidate antibody concentration was observed. Figure 3 shows the results of confirming the competitive binding ability for the antigen between the biotinylated pathogenic antibodies and the native candidate antibodies. The pathogenic antibody was detected with Alexa647 conjugated streptavidin, and the signal intensity was analyzed through FACs. The signal intensity when the pathogenic antibody was 100% bound was set to 100, and the signal intensity when the candidate antibody was 100% bound was replaced with 0, and the degree of signal decrease when the binding ability of the pathogenic antibody was inhibited by the candidate antibody was observed.

[0189] As a result, as shown in Figure 3, both mAb637 B and mAb637 C antibodies competitively bound to autoantigens with mAb637, thereby inhibiting the binding of mAb637 to autoantigens. The above results suggest that the antibodies of the present invention can prevent or treat autoimmune diseases by competitively inhibiting antibodies that bind to autoantigens and cause a decrease in the function of the autoantigens.

[0190] 7-2: Complement-dependent cytotoxicity assay (CDC assay)

[0191] For CDC analysis, the present inventors contacted 293T cell lines expressing the AChR gene and having three complement inhibitor genes knocked out with autoantibodies, mAb35 humanized antibody and mAb35 humanized antibody and competitive recombinant antibody mAb192 C IgG2 / 4, respectively, and confirmed the degree of CDC induction in vitro.

[0192] Specifically, the cell line was contacted with the pathogenic antibody mAb35 humanized antibody and the humanized mAb35 antibody and the competitive recombinant antibody mAb192 C IgG2 / 4, respectively, and human normal serum and complement were added and incubated at 37°C for 6 hours to induce CDC. After washing the cells, anti-C5b-9 antibody was added and incubated, and the proportion of C5b-9 positive cells and the proportion of apoptotic cells by adding FVD450 were measured using a fluorescence-activated cell sorter (FACS). Thereafter, when 293T cells expressing the AChR gene and knocking out three complement inhibitor genes were contacted with the pathogenic antibody mAb35 humanized antibody, or the humanized mAb35 antibody and the competitive recombinant antibody mAb192 C IgG2 / 4, the proportion of induced C5b-9 positive cells and the proportion of apoptotic cells were measured.

[0193] The concentrations of the above-mentioned pathogenic antibodies are 0 μg / ml on the left and 10 μg / ml thereafter, and the concentrations of mAb192 C IgG2 / 4 are 0.00 μg / ml, 0.00 μg / ml, 0.04 μg / ml, 0.15 μg / ml, 0.62 μg / ml, 2.5 μg / ml, and 10 μg / ml from left to right, respectively, and complement was added to human serum at a concentration of 10%. The pathogenic antibodies refer to serum containing the humanized antibody mAb35. mAb192 C IgG2 / 4 refers to a recombinant antibody containing a light chain of SEQ ID NO: 3 and a heavy chain of SEQ ID NO: 4.

[0194] As a result, it was confirmed that treatment with the humanized mAb35 antibody increased the proportion of C5b-9-positive cells (Fig. 4a) and the proportion of cells killed by CDC (Fig. 4b) depending on the concentration, whereas co-treatment with the competitive recombinant antibody mAb192 C IgG2 / 4 suppressed the increase in C5b-9-positive cells and the apoptosis rate. In addition, when the humanized mAb35 autoantibody and the competitive recombinant antibody mAb192 C IgG2 / 4 and its variants (mAb192 C IgG1 and mAb192 C IgG4) were each contacted with 293T cells expressing the AChR gene and knocked out of three complement inhibitor genes, the proportion of induced C5b-9-positive cells suppressed was analyzed. The CDC inhibitory activity was calculated as 100% when only the pathogenic antibody and complement were added without the competitive recombinant antibody mAb192 C IgG2 / 4 and the above mutants, and 0% when only the complement was added. Then, the relative signal by mAb192 C IgG2 / 4 and the above mutants was calculated to indicate the degree of CDC inhibitory activity.

[0195] As a result, as shown in Figures 4c and 4d, the proportion of C5b-9 positive cells and apoptosis increased by treating with mAb35 humanized antibody V L and V H When 0.15 μg / ml of mAb192 C IgG2 / 4 or the above variants were treated together, which exchanged CH1, CH2, and CH3 regions from IgG1 to IgG2 / 4, C5b-9 positive cells were reduced by about 60% or more and apoptosis was reduced by about 37% or more. The results are shown in V L and V H It shows the excellent CDC inhibitory ability of the candidate antibody that was exchanged.

[0196] 7-3: Analysis of ADCC inhibition ability of recombinant antibodies using a neuromyelitis optica cell model

[0197] Neuromyelitis optica (NMO) spectrum disorder is a chronic disease of the brain and spinal cord caused by inflammation of the optic nerve (optic neuritis) and the spinal cord (myelitis), resulting in damage or destruction of the myelin sheath and underlying nerve fibers. It is also known as Devic's disease. It is known that NMO is induced by autoantibodies to aquaporin-4 (ADCC). To confirm the inhibitory effect of therapeutic antibodies against NMO, we established an in vitro ADCC model using an aquaporin-4 CHOK1 (AQP4-CHOK1) cell line expressing pathogenic aquaporin-4. First, model cells were cultured in the presence of NMO pathogenic autoantibodies, recombinant antibodies that competitively bind to the autoantibodies against aquaporin-4, or recombinant antibodies that competitively bind to the autoantibodies against NMO pathogenic autoantibodies against aquaporin-4, and the degree of ADCC occurrence was measured. Specifically, AQP4-CHOK1, pathogenesis-inducing antibodies (together with or alone the present antibody), and NK92 cells were co-cultured for 1 hour, and the ratio of AQP4-CHOK1 cells killed by NK92 cells was measured (viability assay). rAb58, an antibody against rat-derived aquaporin-4, was used as the NMO pathogenic autoantibody. The recombinant antibody that competitively binds to the autoantibodies against aquaporin-4 is rAb53 C antibody. The rAb53 C antibody has the V of the heavy chain of the rAb53 antibody. H V of domain and light chain L It has a structure in which domains are exchanged, and the heavy chain constant region is replaced from IgG1 to IgG2 / 4.

[0198] FIG. 5 of the present invention shows the results of analyzing cell viability when aquaporin-4 expressing U87MG cells are contacted with NMO pathogenic autoantibody rAb58 or recombinant antibody rAb53 C that competitively binds to the aquaporin-4 antigen. When cells are contacted with rAb58, when they are contacted with rAb53 C, and when they are contacted with rAb53 C and then with rAb58, the relative cell viability is calculated as 100% when only NK92 cells and AQP4-CHOK1 cells are co-incubated without antibodies, and the viability when all cells are killed is calculated as 0%. Then, the ratio of cell viability when each antibody is added is relatively calculated and displayed on the graph.

[0199] As a result, as shown in Fig. 5, when cells were brought into contact with rAb58, the relative viability of cells decreased depending on the concentration, but when brought into contact with rAb53 C, the relative viability tended to be maintained. In addition, when cells were brought into contact with rAb53 C and then brought into contact with rAb58, the decrease in relative cell viability tended to be suppressed. The above results appear to be because rAb53 C binds to autoantigens, thereby preventing autoantibodies from binding to autoantigens, thereby reducing cell internalization, and because ADCC of cells is not observed by rAb53 C with the effector function removed.

[0200] 7-4: Antibody internalization assay

[0201] For antibody internalization analysis, the inventors contacted 293T cell lines expressing the AChR gene with Fabfluor-labeled pathogenic substances or candidate antibody variants (mAb192 C IgG1, mAb192 C IgG4, and mAb192 C IgG2 / G4) and confirmed the degree of internalization induction in vitro. Specifically, the cell line was contacted with 4 ㎍ / ml of Fabfluor-labeled pathogenesis-inducing antibody mAb192 IgG1 and competitive recombinant antibody mAb192 C IgG2 / 4 and its variants (mAb192 C IgG1 and mAb192 C IgG4), respectively, and incubated at 37°C for 24 hours, and the degree of antibody internalization was measured by fluorescence at 30-minute intervals. In addition, Fabfluor-labeled mAb35 humanized antibody and serially diluted concentrations of mAb192 C IgG2 / 4 and its variants were contacted, and incubated at 37°C for 24 hours, and the degree of antibody internalization was measured by fluorescence at 30-minute intervals. Figure 6a shows the pathogenesis-inducing substance mAb192, competitive recombinant antibody mAb192 C IgG2 / 4 and its variant mAb192 C IgG1, mAb192 C IgG4 represent the levels of internalization by binding to AChR, respectively.

[0202] As a result, as shown in Fig. 6a, the greatest internalization was observed when mAb192 contacted the cells, and V L and V H or exchange (mAb192 C IgG1 and mAb192 C Ig4), V L and V H The substance (mAb192 C IgG2 / 4) with the CH1, CH2, and CH3 domains exchanged and IgG2 / 4 showed reduced internalization compared to the pathogenic substance. In particular, V L and V HAmong the mutants that were exchanged, the level of internalization was the lowest for mAb192 C IgG2 / 4, a competitive recombinant antibody in which the CH1, CH2, and CH3 regions were exchanged with IgG2 / 4, which suggests that the possibility of internalization of AChR by antibody binding is also very low.

[0203] Figures 6b to 6d show the level at which internalization of the humanized antibody mAb35, a pathogenic agent, is inhibited by the competitive recombinant antibody mAb192 C IgG2 / 4 and its variants mAb192 C IgG1 and mAb192 C IgG4.

[0204] Figure 6b shows that the inhibition of internalization of mAb35 humanized antibody increases with increasing concentration of mAb192 C IgG2 / 4 or its variants, and the internalization of most pathogenic substances was inhibited by 40 μg / ml of mAb192 C IgG2 / 4 or its variants. In particular, V L and V H The recombinants that were exchanged had an excellent effect on suppressing the internalization of pathogenic substances, which was V L and V H The recombinants that were exchanged are shown to be effective in suppressing the internalization of pathogenic substances. When looking at Figures 6a to 6d in a comprehensive manner, V L and V H mAb192 C IgG2 / 4, which exchanged the CH1, CH2, and CH3 regions with IgG2 / 4, shows that the substance itself is not well internalized, and effectively inhibits the internalization of the pathogenesis-causing substance, thereby showing that it is the form that can best inhibit the pathogenesis-causing substance from binding to AChR and being internalized together, causing AChR loss.

[0205] Example 8: In vivo efficacy trial in myasthenia gravis

[0206] 8-1: In vivo efficacy of competitive recombinant antibodies

[0207] Lewis rats were administered 5 mg / kg of therapeutic recombinant antibody four times at 3.5-day intervals, followed by 1.5 mg / kg of myasthenia gravis pathogenic autoantibody, and body weight changes and clinical scores of myasthenia gravis were observed over time. The pathogenic autoantibody was mAb35, and the therapeutic recombinant antibody was mAb35, which contained the V heavy chain. H V of domain and light chain L This is an antibody in which domains are exchanged and the heavy chain constant region CH2-CH3 domain is replaced with IgG2 / 4.

[0208] Figure 7a shows the change in body weight over time in Lewis rats administered 5 mg / kg of therapeutic antibody four times at 3.5-day intervals, followed by 1.5 mg / kg of myasthenia gravis pathogenic autoantibody mAb35. Figure 7b shows the change in clinical scores over time in Lewis rats administered 5 mg / kg of therapeutic antibody four times at 3.5-day intervals, followed by 1.5 mg / kg of myasthenia gravis pathogenic autoantibody mAb35. Figure 7c shows the change in body weight and clinical scores according to the administered dose of therapeutic antibody. In Figures 7a and 7b, G1 was administered only PBS as a carrier, G2 was administered PBS followed by mAb35, G3 was administered the therapeutic antibody, and G4 was administered mAb35 and the therapeutic antibody. As shown in Figures 7a and 7b, when only PBS was administered (G1), weight loss due to administration stress was observed, but there was an overall tendency for weight to increase during the administration period. When mAb35 antibody was administered after PBS administration (G2), weight loss and induction of myasthenia gravis symptoms were confirmed. When the therapeutic antibody was repeatedly administered four times (G3), weight gain almost similar to that of the PBS administration group was observed, and induction of myasthenia gravis symptoms was not confirmed. When the therapeutic antibody was repeatedly administered four times and mAb35 antibody was administered (G4), weight loss and myasthenia gravis symptoms due to mAb35 antibody were not observed at all. This indicates that the therapeutic antibody not only does not induce symptoms but also has the effect of preventing the induction of symptoms due to autoantibodies.

[0209] When mAb35 antibody was administered after repeated administration of therapeutic antibodies, body weight decreased rapidly only in the group that did not receive therapeutic antibodies, while there was no significant change in the group that did. Furthermore, clinical scores also appeared only in the group that did not receive therapeutic antibodies, and some individuals ultimately died (clinical score = 4). Clinical scores were not measured in any group except the group administered pathogenic antibody mAb35, and thus are not shown in Figure 7b.

[0210] In addition, as shown in Fig. 7c, in order to confirm the dose-dependent effect of the therapeutic antibody, the therapeutic antibody was administered four times over two weeks at doses of 0 mg / kg, 0.125 mg / kg, 0.25 mg / kg, 0.5 mg / kg, 1 mg / kg, and 2 mg / kg, and the suppression of symptoms by the mAb35 antibody was confirmed. As a result, clinical symptoms and weight loss due to mAb35 administration were suppressed at a dose of 0.25 mg / kg, and no clinical symptoms were observed at concentrations higher than 0.5 mg / kg.

[0211] According to the above results, it is confirmed from the above-described examples that the antibody of the present invention is effective in treating autoimmune diseases associated with decreased function of autoantigens. In particular, the antibody platform of the present invention shows excellent effects in preventing or treating autoimmune diseases caused by autoantibodies such as autoantibodies mAb35, mAb637, mAb192, or rAb53, and thus it can be confirmed that autoimmune diseases caused by various autoantibodies can be prevented or treated. In addition, the antibody platform of the present invention has been confirmed to have preventive or therapeutic effects on various diseases such as myasthenia gravis or neuromyelitis optica, and thus can be utilized in the prevention or treatment of various types of autoimmune diseases.

[0212] While the present invention has been described with reference to the above-described embodiments, these are merely exemplary, and those skilled in the art will understand that various modifications and equivalent alternative embodiments are possible. Therefore, the true scope of technical protection of the present invention should be determined by the technical spirit of the appended claims.

Claims

1. A recombinant homodimeric antibody derived from an autoantibody having specificity for an autoantigen, which comprises a light chain and a heavy chain, wherein said recombinant homodimeric antibody has specificity for said autoantigen, wherein the heavy chain comprises at least a portion of the variable region of the light chain (V L ) autoantibodies, and the light chain contains at least part of the variable region of the heavy chain (V H ) autoantibodies.

2. A homodimeric antibody according to claim 1, wherein the autoantigen is a membrane protein.

3. The homodimeric antibody of claim 1, which reduces or prevents the formation of a cross-linking structure by binding of the autoantibody to multiple autoantigens.

4. A homodimeric antibody according to claim 3, which reduces or prevents internalization of an antigen-antibody complex caused by cross-linking.

5. The homodimeric antibody of claim 3, wherein the autoantibody induces complement fixation by cross-linking.

6. A homodimeric antibody according to claim 1, which inhibits the binding of an autoantibody to an autoantigen by binding to the same site as the epitope of the autoantigen to which the autoantibody binds, or to a site located near said epitope.

7. The homodimeric antibody of claim 1, wherein the binding orientation of at least one monomer of the homodimeric antibody relative to the autoantigen differs from the binding orientation of the autoantibody relative to the autoantigen.

8. The homodimeric antibody of claim 7, wherein the binding of one monomer of the homodimeric antibody to a first autoantigen results in a binding orientation that prevents the binding of the remaining monomer of the homodimeric antibody to a second autoantigen located on the cell surface.

9. A homodimeric antibody according to claim 7, having a conformation that does not bind to a plurality of complex-forming autoantigens among the complex-forming autoantigens located on the cell surface.

10. A homodimeric antibody according to claim 9, which does not cross-link two or more autoantigens that form a complex located on the cell surface.

11. The homodimeric antibody of claim 1, wherein the heavy chain Fc region is an IgG1 Fc, an IgG4 Fc, or an IgG2 / 4 hybrid Fc.

12. The homodimeric antibody of claim 1, wherein the autoantibody comprises a first half-antibody fragment comprising a first light chain variable region (V L ) and the first variable region of the heavy chain (V H ) and has specificity for the autoantigen, and a second fragment of the half-antibody containing the second variable region of the light chain (V L ) and the second variable region of the heavy chain (V H) and has specificity for the autoantigen; wherein said homodimeric antibody comprises at least one of the following half-antibody fragments: i) a heavy chain comprising a first light chain variable region (V L ), and a light chain containing the first variable region of the heavy chain (V H ); ii) a heavy chain containing a second variable region of the light chain (V L ), and a light chain containing the second variable region of the heavy chain (V H ); iii) a heavy chain comprising at least one of CDRL1, CDRL2 and CDRL3 of the first variable region of the light chain (V L ), and a light chain comprising at least one of CDRH1, CDRH2 and CDRH3 of the first variable region of the heavy chain (V H ) and the remaining CDRL fragments not included in the heavy chain; and iv) a heavy chain comprising at least one of CDRL1, CDRL2 and CDRL3 of a second variable region of a light chain (VL ), and a light chain comprising at least one of CDRH1, CDRH2 and CDRH3 of the second variable region of the heavy chain (V H ) and the remaining parts of CDRL not included in the heavy chain.

13. The homodimeric antibody of claim 1, wherein portions of the variable regions of the heavy chain and light chain of the autoantibody are substituted for each other.

14. The homodimeric antibody of claim 1, wherein the autoantibody comprises a first half-antibody fragment comprising a first variable region of a light chain and a first variable region of a heavy chain and having specificity for an autoantigen; and a second half-antibody fragment comprising a second variable region of a light chain and a second variable region of a heavy chain and having specificity for an autoantigen, wherein in the homodimeric antibody V L -CL and V H -CH1 of at least one half-antibody fragment among the half-antibody fragments of the autoantibody are replaced by each other.

15. A pharmaceutical composition for the prevention or treatment of an autoimmune disease, comprising a homodimeric antibody according to any one of claims 1-13 and a pharmaceutically acceptable carrier.

16. A recombinant dimeric antibody comprising a variable region of a light chain and a variable region of a heavy chain and having specificity for an autoantigen, wherein in at least one monomer of the recombinant dimeric antibody at least part of the variable region of the light chain (V L ) an autoantibody that has specificity for an autoantigen and pathogenicity for an autoimmune disease, and at least a portion of the variable region of the heavy chain (V H ) the specified autoantibodies are replaced by each other.