Anti-DOCK8 monoclonal antibodies and uses thereof

Abstract

The present invention provides a novel anti-DOCK8 monoclonal antibody and a preventive and / or therapeutic agent for autoimmune diseases which contains the anti-DOCK8 monoclonal antibody as an active ingredient. [Solution] An anti-DOCK8 monoclonal antibody having heavy chain CDR1 to 3 and light chain CDR1 to 3 represented by specific amino acid sequences is used.

Description

[Technical field]

[0001] The present invention relates to an anti-DOCK8 monoclonal antibody and uses thereof. [Background technology]

[0002] The immune system exists as a defense mechanism for the body against the invasion of harmful foreign substances from the outside world. However, sometimes the function of this immune system can end up being harmful to the body. It is known that the body can mount an immune response not only to foreign substances from the outside world, but also to its own components; this is called an autoimmune phenomenon. Diseases in which certain pathological conditions are caused by autoimmunity are called autoimmune diseases.

[0003] Autoimmune diseases are systemic diseases, but are broadly divided into two categories: organ-specific autoimmune diseases and non-organ-specific autoimmune diseases. Many of these autoimmune diseases cause lesions in tissues and are accompanied by tissue damage.

[0004] Systemic lupus erythematosus (SLE), a non-organ-specific autoimmune disease, is known to increase the production of autoantibodies such as anti-dsDNA antibodies. There is hope for molecular biological research into the process that causes SLE, elucidation of its relationship with various pathological conditions, and even the development of treatments.

[0005] When laboratory animals are repeatedly immunized with the same antigen, the immune response reaches a peak and eventually becomes exhausted, which is known to result in various autoimmune diseases (pathologies) accompanied by tissue damage (see, for example, Patent Document 1 and Non-Patent Document 1).

[0006] Research into tissue damage caused by autoimmune diseases is currently progressing, and it has been revealed that cross-presentation of antigens by antigen-presenting cells is involved in tissue damage. Specifically, (1) when patients suffer from autoimmune diseases, cross-presentation of antigens in dendritic cells is enhanced, and (2) enhanced cross-presentation in dendritic cells leads to CD8 + T cells are activated and the CD8 + It has been shown that cytotoxicity is induced by T cells (see, for example, Patent Document 2 and Non-Patent Document 1).

[0007] The present inventors have previously reported that repeated immunization of mice with antigens generates a new type of T cell, autoreactive / autoantibody-inducing CD4 T cells (aiCD4 T cells), in the periphery, and that the aiCD4 T cells induce a variety of autoantibodies and mature cytotoxic T cells (CTLs), causing various organ disorders and leading to the onset of SLE (Non-Patent Document 1). Furthermore, the inventors have reported that the aiCD4 T cells inhibit PD-1 + CD45RB low 122 low (PD-1 + CD45RB lo 122 lo They have found that autoantibodies are present in a CD4 T cell subpopulation of the IL-16-positive mice, and that when this cell subpopulation was adoptively transferred to naive mice, autoantibodies were observed in the serum of the recipient mice two weeks after transfer (see Non-Patent Document 2).

[0008] Furthermore, the inventors have discovered a marker to more easily identify the cells that cause the onset of autoimmune diseases, and have succeeded in developing a technology based on this that enables objective and accurate diagnosis of autoimmune diseases (see Patent Document 3).

[0009] Furthermore, the present inventors have demonstrated that anti-DOCK8 polyclonal antibodies can be used as prophylactic and / or therapeutic agents for autoimmune diseases (see Patent Document 4).

Prior technical literature

Chartered Documents

[0010] [Patent Document 1] Japanese Patent Publication No. 2006-288382 (published on October 26, 2006) [Patent Document 2] Japanese Patent Publication No. 2010-004750 (published on January 14, 2010) [Patent Document 3] Japanese Patent Publication No. 2017-003329 (published on January 5, 2017) [Patent Document 4] WO2018 / 159549A1 (published on September 7, 2018)

Non-licensed documents

[0011]

Non-patent document 1

Non-patent document 2

Non-patent document 3

Non-patent document 4

Non-patent document 5

Non-patent document 6

[0012] However, the drugs used for the prevention and / or treatment of autoimmune diseases are still insufficient, and further development of new drugs has been desired.

[0013] An object of the present invention is to provide a novel anti-DOCK8 monoclonal antibody and a preventive and / or therapeutic agent for autoimmune diseases which contains the anti-DOCK8 monoclonal antibody as an active ingredient. [Means for solving the problem]

[0014] As a result of intensive research, the present inventors have succeeded in obtaining a novel anti-DOCK8 monoclonal antibody, and have found that the use of said anti-DOCK8 monoclonal antibody can improve symptoms of autoimmune diseases, thereby completing the present invention. One embodiment of the present invention includes the following configuration.

[0015] [1] An anti-DOCK8 monoclonal antibody having heavy chain CDR1 to 3 and light chain CDR1 to 3 as shown in (1) or (2) below: (1) heavy chain CDR1 to 3 having amino acid sequences consisting of SEQ ID NOs: 1 to 3, and light chain CDR1 to 3 having amino acid sequences consisting of SEQ ID NOs: 4 to 6; (2) Heavy chain CDRs 1 to 3 having the amino acid sequences of SEQ ID NOs: 7 to 9, and light chain CDRs 1 to 3 having the amino acid sequences of SEQ ID NOs: 10 to 12.

[0016] [2] The anti-DOCK8 monoclonal antibody according to [1], having a heavy chain variable region and a light chain variable region shown in the following (1') or (2'): (1') a heavy chain variable region having the amino acid sequence of SEQ ID NO: 13, and a light chain variable region having the amino acid sequence of SEQ ID NO: 14; (2') a heavy chain variable region having an amino acid sequence consisting of SEQ ID NO: 15, and a light chain variable region having an amino acid sequence consisting of SEQ ID NO: 16.

[0017] [3] The anti-DOCK8 monoclonal antibody according to [1] or [2], having a heavy chain constant region having an amino acid sequence consisting of SEQ ID NO: 17.

[0018] [4] The anti-DOCK8 monoclonal antibody according to any one of [1] to [3], wherein the light chain constant region is a λ chain having an amino acid sequence consisting of SEQ ID NO: 18 or a κ chain having an amino acid sequence consisting of SEQ ID NO: 19.

[0019] [5] The anti-DOCK8 monoclonal antibody according to any one of [1] to [4], having a heavy chain and a light chain represented by the following (1'') or (2''): (1'') a heavy chain having an amino acid sequence consisting of SEQ ID NO:20, and a light chain having an amino acid sequence consisting of SEQ ID NO:21; (2'') a heavy chain having an amino acid sequence consisting of SEQ ID NO:22, and a light chain having an amino acid sequence consisting of SEQ ID NO:23.

[0020] [6] A preventive and / or therapeutic agent for an autoimmune disease, comprising the anti-DOCK8 monoclonal antibody according to any one of [1] to [5] as an active ingredient.

[0021] [7] The preventive and / or therapeutic agent for an autoimmune disease according to [6], wherein the autoimmune disease is systemic lupus erythematosus (SLE). Effect of the Invention

[0022] According to one embodiment of the present invention, it is possible to provide a novel anti-DOCK8 monoclonal antibody and a preventive and / or therapeutic agent for autoimmune diseases comprising the anti-DOCK8 monoclonal antibody as an active ingredient. [Brief description of the drawings]

[0023] [Figure 1] 1 is a graph showing the antigen recognition ability of monoclonal antibodies obtained in an example of the present invention (ELISA). [Diagram 2] 1 is a graph showing the antigen recognition ability of monoclonal antibodies obtained in an example of the present invention (FCM). [Diagram 3] 3 shows a tissue staining image (301) and a graph (302) showing the percentage of DOCK8-positive CD4 T cells in a patient with an autoimmune disease (active disease stage) in an example of the present invention. [Figure 4] 1 is a graph showing the percentage of DOCK8-positive CD4 T cells in patients with autoimmune disease (active disease stage) in an Example of the present invention. [Diagram 5] 1 is a graph showing the pharmacological effects of monoclonal antibodies in an example of the present invention. [Figure 6] 1 is a graph showing the pharmacological effects of monoclonal antibodies in an example of the present invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0024] An embodiment of the present invention will be described below, but the present invention is not limited thereto. The present invention is not limited to each configuration described below, and various modifications are possible within the scope of the claims, and embodiments and examples obtained by appropriately combining the technical means disclosed in different embodiments and examples are also included in the technical scope of the present invention. In addition, all academic literature and patent documents described in this specification are incorporated herein by reference. In addition, unless otherwise specified in this specification, "A to B" representing a numerical range means "A or more, B or less."

[0025] [1. Anti-DOCK8 monoclonal antibody] DOCK8 protein is a member of the DOCK family of proteins and is a 238 kDa protein that functions as a guanine nucleotide exchange factor (GEF). DOCK8 protein is known to control actin cytoskeleton polymerization through activation of Rho family small G proteins. Reorganization of the actin cytoskeleton is involved in various cellular functions such as cell differentiation, proliferation, migration, and signal transduction.

[0026] In recent years, it has been discovered that deficiency in the human DOCK8 gene causes severe combined immunodeficiency disease, and since then, several immunological research results on DOCK8 have been reported (see Non-Patent Documents 3 to 8).

[0027] In this specification, unless otherwise specified, "DOCK8" refers to both the DOCK8 protein, and the DNA and RNA (gene) that encodes the DOCK8 protein.

[0028] Compared with an anti-DOCK8 polyclonal antibody, an anti-DOCK8 monoclonal antibody according to one embodiment of the present invention can reduce nonspecific binding of the antibody to targets other than DOCK8 protein and can homogenize the physical properties of drugs containing the antibody, thereby improving the pharmacological effect of preventive and / or therapeutic agents for autoimmune diseases and suppressing the occurrence of side effects of preventive and / or therapeutic agents for autoimmune diseases.

[0029] More specifically, the anti-DOCK8 monoclonal antibody of one embodiment of the present invention is a fully human autoantibody and is therefore not foreign to humans. Compared to an anti-DOCK8 polyclonal antibody, it can reduce nonspecific binding of the antibody to targets other than DOCK8 protein, and it can homogenize the physical properties of drugs containing the antibody, thereby improving the pharmacological effect of preventive and / or therapeutic agents for autoimmune diseases and suppressing the occurrence of side effects of preventive and / or therapeutic agents for autoimmune diseases.

[0030] Furthermore, since the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention can be produced easily and in large quantities compared to an anti-DOCK8 polyclonal antibody, a preventive and / or therapeutic agent for an autoimmune disease can be produced easily and in large quantities.

[0031] An anti-DOCK8 monoclonal antibody according to one embodiment of the present invention is an anti-DOCK8 monoclonal antibody having heavy chain CDR1 to 3 and light chain CDR1 to 3 as shown in the following (1) or (2): (1) heavy chain CDR1 to 3 having amino acid sequences consisting of SEQ ID NOs: 1 to 3, and light chain CDR1 to 3 having amino acid sequences consisting of SEQ ID NOs: 4 to 6; (2) Heavy chain CDRs 1 to 3 having the amino acid sequences of SEQ ID NOs: 7 to 9, and light chain CDRs 1 to 3 having the amino acid sequences of SEQ ID NOs: 10 to 12.

[0032] Regarding the anti-DOC8 monoclonal antibody described in (1) above, the amino acid sequences shown in SEQ ID NOs: 1 to 6 are specifically as follows: SEQ ID NO: 1: GFTFSSYA, SEQ ID NO:2: ISGSGGST, SEQ ID NO:3: AKNLRSLIYMDD, SEQ ID NO:4: SLRSYY, SEQ ID NO:5: GKN, Sequence number 6: QSSSSSSHFDV.

[0033] Regarding the anti-DOC8 monoclonal antibody described in (2) above, the amino acid sequences shown in SEQ ID NOs: 7 to 12 are specifically as follows: SEQ ID NO: 7: GFTFSSYA, SEQ ID NO: 8: ISGSGGST, SEQ ID NO: 9: AKAGQEAYMDD, SEQ ID NO:10: QGISSW, SEQ ID NO:11: AAS, Sequence number 12: QQYSSSPIT.

[0034] In an anti-DOCK8 monoclonal antibody according to one embodiment of the present invention, the amino acid sequences of the heavy chain variable region and the light chain variable region are not limited. The anti-DOCK8 monoclonal antibody according to one embodiment of the present invention is preferably an anti-DOCK8 monoclonal antibody having the heavy chain variable region and the light chain variable region shown in the following (1') or (2'): (1') a heavy chain variable region having the amino acid sequence of SEQ ID NO: 13, and a light chain variable region having the amino acid sequence of SEQ ID NO: 14; (2') a heavy chain variable region having an amino acid sequence consisting of SEQ ID NO: 15, and a light chain variable region having an amino acid sequence consisting of SEQ ID NO: 16.

[0035] Regarding the anti-DOC8 monoclonal antibody shown in (1') above, the amino acid sequences shown in SEQ ID NOs: 13 to 14 are specifically as follows: SEQ ID NO: 13: EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKNLRSLIYMDDWGQGTLVTVSS, SEQ ID NO:14: SSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCQSSSSSSHFDVFGGGTKLTVL.

[0036] Regarding the anti-DOC8 monoclonal antibody described in (2') above, the amino acid sequences shown in SEQ ID NOs: 15 to 16 are specifically as follows: SEQ ID NO: 15: EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKAGQEAYMDDWGQGTLVTVSS, SEQ ID NO:16:DIQMTQSPSSLSASVGDRVTITCRASQGISSWLAWYQQKPEKAPKSLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQYSSSPITFGQGTKLEIK.

[0037] In the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention, the amino acid sequence of the heavy chain constant region is not limited. The anti-DOCK8 monoclonal antibody according to one embodiment of the present invention preferably has a heavy chain constant region having the amino acid sequence of SEQ ID NO: 17; SEQ ID NO:17: TKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAPPAAAPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG.

[0038] In the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention, the amino acid sequence of the light chain constant region is not limited. In the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention, the light chain constant region is preferably a λ chain having the amino acid sequence of SEQ ID NO: 18 or a κ chain having the amino acid sequence of SEQ ID NO: 19; SEQ ID NO: 18: APSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS, SEQ ID NO:19: VAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC.

[0039] In the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention, the amino acid sequences of the heavy and light chains are not limited. The anti-DOCK8 monoclonal antibody according to one embodiment of the present invention preferably has the heavy and light chains shown in (1'') or (2'') below: (1'') a heavy chain having an amino acid sequence consisting of SEQ ID NO:20, and a light chain having an amino acid sequence consisting of SEQ ID NO:21; (2'') a heavy chain having an amino acid sequence consisting of SEQ ID NO:22, and a light chain having an amino acid sequence consisting of SEQ ID NO:23.

[0040] Regarding the anti-DOC8 monoclonal antibody shown in (1'') above, the amino acid sequences shown in SEQ ID NOs: 20 to 21 are specifically as follows: SEQ ID NO: 20: EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKNLRSLIYMDDWGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVER KCCVECPPCPAPPAAAPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEK TISKTKGQPREPQVYTLPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG, SEQ ID NO:21: SSELTQDPAVSVALGQTVRITCQGDSLRSYYASWYQQKPGQAPVLVIYGKNNRPSGIPDRFSGSSSGNTASLTITGAQAEDEADYYCQSSSSSSHFDVFGGGTKLTVLGQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS.

[0041] Regarding the anti-DOC8 monoclonal antibody described in (2'') above, the amino acid sequences shown in SEQ ID NOs: 22 to 23 are specifically as follows: SEQ ID NO: 22: EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKAGQEAYMDDWGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVER KCCVECPPCPAPPAAAPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEK TISKTKGQPREPQVYTLPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG, SEQ ID NO:23:DIQMTQSPSSLSASVGDRVTITCRASQGISSWLAWYQQKPEKAPKSLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQYSSSPITFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC.

[0042] An anti-DOCK8 monoclonal antibody according to one embodiment of the present invention may be an anti-DOCK8 monoclonal antibody that binds (e.g., specifically binds) to an antigen consisting of the amino acid sequence shown in SEQ ID NO: 24. The amino acid sequence shown in SEQ ID NO: 24 is specifically as follows: SEQ ID NO:24: RNLLYVYPQRLNFASKLASARNITIKIQFMCGEDPSNAMPVIFGKSSGPEFLQEVYTAITYHNKSPDFYEEVKIKLPAKLTVNHHLLFTFYHISCQQKQGASGESLLGYSWLPILLNERLQTGSYCLPVALEKLPPNYSIHSAEKVPLQNPPIKWAEGHKGVFNIEVQAV.

[0043] As used herein, the term "specifically binds" refers to an antibody that binds to an antigen with higher affinity than to other polypeptides. Here, "high affinity" refers to, for example, a binding constant (Ka) of 10 7 M -1 More than 10, preferably 8 M -1 More than 10, more preferably 9 M -1 More than 10, more preferably 10 M -1 More than 10, more preferably 11 M -1 More than 10, more preferably 12 M -1 More than 10, most preferably 13 M -1 The affinity is intended to be equal to or greater than this.

[0044] The anti-DOCK8 monoclonal antibody according to one embodiment of the present invention may comprise the whole anti-DOCK8 monoclonal antibody, consist of the whole anti-DOCK8 monoclonal antibody, comprise a fragment of the anti-DOCK8 monoclonal antibody, or consist of a fragment of the anti-DOCK8 monoclonal antibody. For example, the anti-DOCK8 monoclonal antibody may be a chimeric antibody, a humanized antibody, or a fragment thereof (e.g., F(ab')2, Fab', Fab, or Fv). The anti-DOCK8 monoclonal antibody may be IgA, IgD, IgE, IgG, IgM, or a fragment thereof.

[0045] Anti-DOCK8 monoclonal antibodies can be prepared according to well-known methods (e.g., (1) HarLow et al., "Antibodies: A laboratory manual, Cold Spring Harbor Laboratory, New York (1988)" and (2) Iwasaki et al., "Monoclonal Antibodies: Hybridomas and ELISA, Kodansha (1991)"). Of course, commercially available antibodies can also be used as anti-DOCK8 antibodies.

[0046] Anti-DOCK8 monoclonal antibodies can be produced according to methods well known in the art (e.g., (1) the hybridoma method (Kohler, G. and Milstein, C., Nature 256, 495-497 (1975)), (2) the trioma method, (3) the human B cell hybridoma method (Kozbor, Immunology Today 4, 72 (1983)), and (4) the EBV-hybridoma method (Monoclonal Antibodies and Cancer Therapy, Alan R Liss, Inc., 77-96 (1985))).

[0047] Anti-DOCK8 monoclonal antibodies can also be produced according to the phage display method. A known method may be used to produce monoclonal antibodies using the phage display method. An example of a method for producing monoclonal antibodies using the phage display method is described below. First, mRNA is prepared from an animal immunized with DOCK8 protein, and cDNA is prepared using the mRNA as a template, and a single-chain antibody (scFV) gene encoding only the variable region of the antibody is produced from the cDNA. Next, the single-chain antibody gene is inserted into a phagemid vector, and the phagemid vector is introduced into E. coli, and the E. coli is infected with a phage. This allows the scFV antibody to be expressed on the phage coat. Anti-DOCK8 monoclonal antibodies can be produced by screening the scFV antibodies expressed on the phage coat for those that bind to the DOCK8 protein. Preparation of mRNA, preparation of cDNA, insertion of the single-chain antibody gene into a phagemid vector, introduction of the phagemid vector into E. coli, infection with phage, and screening of the antibody can be carried out according to well-known methods.

[0048] By chimeric antibodies is intended antibodies in which the constant regions have been replaced with the constant regions of a human antibody. Chimeric antibodies can be produced according to well known methods (see, for example, European Patent Publication EP 0125023).

[0049] A humanized antibody is intended to mean an antibody in which the structures other than the complementarity determining regions (CDRs) of the H and L chains have been replaced with those of a human antibody.

[0050] By human antibody is intended an antibody made using transgenic animals into which genes responsible for human antibody production have been introduced (see, for example, European Patent Publication No. EP0546073).

[0051] F(ab')2, Fab', Fab, and Fv can be obtained by degrading the whole antibody with a protease (e.g., papain or pepsin) and, if necessary, further reducing the degraded product. Alternatively, F(ab')2, Fab', Fab, and Fv can be obtained by isolating cDNA from an antibody-producing hybridoma, inserting the cDNA into an expression vector, and introducing the expression vector into a host. In this case, the antibody fragment can also be obtained as a fusion protein of the antibody fragment and another protein.

[0052] 2. Preventive and / or therapeutic agents for autoimmune diseases The prophylactic and / or therapeutic agent for autoimmune diseases according to one embodiment of the present invention contains the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention as an active ingredient. Note that the anti-DOCK8 monoclonal antibody according to one embodiment of the present invention has been explained in the above section [1. Anti-DOCK8 monoclonal antibody], and therefore will not be explained further below.

[0053] As used herein, the term "therapeutic agent" refers to a drug that provides a therapeutic effect. The above-mentioned therapeutic effect refers to, but is not limited to, the following examples: (1) Reducing the severity of one or more symptoms associated with a disease, compared to when the therapeutic agent is not administered; (2) Preventing an increase in the severity of one or more symptoms associated with a disease, compared to when the therapeutic agent is not administered; (3) Reducing the rate of increase in the severity of one or more symptoms associated with a disease, compared to when the therapeutic agent is not administered.

[0054] As used herein, the term "prophylactic agent" refers to a drug that provides a prophylactic effect. The above-mentioned prophylactic effect refers to, but is not limited to, the following examples: (1) Preventing the onset of one or more symptoms associated with a disease, or reducing the risk of onset, compared to when the prophylactic agent is not administered; (2) Preventing the recurrence of one or more symptoms associated with a disease, or reducing the risk of recurrence, compared to when the prophylactic agent is not administered; (3) Preventing the onset of symptoms of one or more symptoms associated with a disease, or reducing the risk of onset, compared to when the prophylactic agent is not administered.

[0055] As used herein, the term "autoimmune disease" refers to a disease in which autoantibodies (e.g., anti-Sm antibodies, anti-dsDNA antibodies, rheumatoid factors (RF)) against self-components (e.g., immunoglobulins, etc.) are detected and autoimmunity is involved in the pathology. Autoimmune diseases include organ-specific autoimmune diseases (e.g., chronic thyroiditis, primary mucosal edema, thyrotoxicosis, pernicious anemia, Goodpasture's syndrome, acute progressive glomerulonephritis, myasthenia gravis, pemphigus vulgaris, bullous pemphigoid, insulin-resistant diabetes mellitus, juvenile diabetes mellitus, Addison's disease, atrophic gastritis, male infertility, premature menopause, lensogenic uveitis, sympathetic phlebitis, multiple sclerosis, ulcerative colitis, primary biliary cirrhosis, chronic active hepatitis, autoimmune hemolytic anemia, paroxysmal hemoglobinuria, idiopathic thrombocytopenic purpura, and Sjogren's syndrome), and organ-nonspecific autoimmune diseases (e.g., rheumatoid arthritis, systemic lupus erythematosus (SLE), discoid lupus erythematosus, polymyositis, scleroderma, and mixed connective tissue disease). Rheumatoid arthritis, systemic lupus erythematosus, discoid lupus erythematosus, polymyositis, scleroderma, and mixed connective tissue disease are known as collagen diseases. In other words, collagen diseases are included in systemic autoimmune diseases. In one embodiment of the present invention, the autoimmune disease is preferably a collagen disease, and more preferably systemic lupus erythematosus.

[0056] The amount of anti-DOCK8 monoclonal antibody contained in the preventive agent and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention is not particularly limited, and may be, for example, 0.001% by weight to 100% by weight, 0.01% by weight to 100% by weight, 0.1% by weight to 100% by weight, 0.1% by weight to 95% by weight, 0.1% by weight to 90% by weight, 0.1% by weight to 80% by weight, 0.1% by weight to 70% by weight, 0.1% by weight to 60% by weight, 0.1% by weight to 50% by weight, 0.1% by weight to 40% by weight, 0.1% by weight to 30% by weight, 0.1% by weight to 20% by weight, or 0.1% by weight to 10% by weight, assuming the preventive agent and / or therapeutic agent to be 100% by weight.

[0057] The amount of anti-DOCK8 monoclonal antibody contained in the preventive agent and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention is not particularly limited, and may be, for example, 0.01 mg to 1000 mg, 0.1 mg to 1000 mg, 1 mg to 1000 mg, 1 mg to 900 mg, 1 mg to 800 mg, 1 mg to 700 mg, 1 mg to 600 mg, 1 mg to 500 mg, 1 mg to 400 mg, 1 mg to 300 mg, 1 mg to 200 mg, or 1 mg to 100 mg.

[0058] The preventive and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention may contain components other than the anti-DOCK8 monoclonal antibody (e.g., a pharma- ceutically acceptable carrier). When the preventive and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention is a solid preparation, the components other than the anti-DOCK8 monoclonal antibody may include excipients, lubricants, binders, and disintegrants, and when the preventive and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention is a liquid preparation, the components may include solvents, solubilizers, suspending agents, isotonicity agents, buffers, and soothing agents. Other components other than the anti-DOCK8 monoclonal antibody may include preservatives, antioxidants, and stabilizers.

[0059] Examples of the above-mentioned "excipient" include, but are not limited to, lactose, sucrose, D-mannitol, xylitol, sorbitol, erythritol, starch and crystalline cellulose.

[0060] Examples of the "lubricant" include, but are not limited to, magnesium stearate, calcium stearate, wax, talc and colloidal silica.

[0061] Examples of the "binder" include, but are not limited to, pregelatinized starch, methylcellulose, crystalline cellulose, sucrose, D-mannitol, trehalose, dextrin, hydroxypropyl cellulose, hydroxypropylmethyl cellulose, and polyvinylpyrrolidone.

[0062] Examples of the "disintegrant" include, but are not limited to, starch, carboxymethylcellulose, low-substituted hydroxypropylcellulose, calcium carboxymethylcellulose, croscarmellose sodium, and sodium carboxymethylstarch.

[0063] Examples of the "solvent" include, but are not limited to, water for injection, alcohol, propylene glycol, macrogol, sesame oil, corn oil and tricaprylin.

[0064] Examples of the "solubilizing agent" include, but are not limited to, polyethylene glycol, propylene glycol, D-mannitol, trehalose, benzyl benzoate, ethanol, trisaminomethane, cholesterol, triethanolamine, sodium carbonate, and sodium citrate.

[0065] Examples of the "suspending agent" include, but are not limited to, surfactants (e.g., stearyl triethanolamine, sodium lauryl sulfate, lauryl aminopropionic acid, lecithin, benzalkonium chloride, benzethonium chloride, glycerin monostearate) and hydrophilic polymers (e.g., polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethylcellulose, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose).

[0066] Examples of the "isotonicity agent" include, but are not limited to, sodium chloride, glycerin, and D-mannitol.

[0067] The above-mentioned "buffers" include, but are not limited to, for example, phosphates, acetates, carbonates and citrates.

[0068] The above-mentioned "soothing agent" includes, but is not limited to, benzyl alcohol.

[0069] Examples of the "preservative" include, but are not limited to, paraoxybenzoic acid esters, chlorobutanol, benzyl alcohol, phenethyl alcohol, dehydroacetic acid, and sorbic acid.

[0070] The above-mentioned "antioxidants" include, but are not limited to, for example, sulfites and ascorbic acid.

[0071] The above-mentioned "stabilizer" is not particularly limited as long as it is one that is commonly used in the pharmaceutical field.

[0072] The amount of components other than the anti-DOCK8 monoclonal antibody contained in the preventive agent and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention is not particularly limited, and may be, for example, 0% by weight to 99.999% by weight, 0% by weight to 99.99% by weight, 0% by weight to 99.99% by weight, 0% by weight to 99.9% by weight, 5% by weight to 99.9% by weight, 10% by weight to 99.9% by weight, 20% by weight to 99.9% by weight, 30% by weight to 99.9% by weight, 40% by weight to 99.9% by weight, 50% by weight to 99.9% by weight, 60% by weight to 99.9% by weight, 70% by weight to 99.9% by weight, 80% by weight to 99.9% by weight, or 90% by weight to 99.9% by weight.

[0073] The amount of components other than the anti-DOCK8 monoclonal antibody contained in the preventive agent and / or therapeutic agent for autoimmune disease according to one embodiment of the present invention is not particularly limited, and may be, for example, 0.01 mg to 1000 mg, 0.1 mg to 1000 mg, 1 mg to 1000 mg, 1 mg to 900 mg, 1 mg to 800 mg, 1 mg to 700 mg, 1 mg to 600 mg, 1 mg to 500 mg, 1 mg to 400 mg, 1 mg to 300 mg, 1 mg to 200 mg, or 1 mg to 100 mg.

[0074] The method of administration of the prophylactic and / or therapeutic agent for autoimmune diseases according to one embodiment of the present invention is not particularly limited, and the agent may be administered orally or parenterally, intravenously, intrarectally, intraperitoneally, intramuscularly, intranasally, or subcutaneously.

[0075] The dosage form of the prophylactic and / or therapeutic agent for autoimmune diseases according to one embodiment of the present invention is not particularly limited, and may be, for example, an injection (e.g., an intraperitoneal injection, an intravenous injection, a subcutaneous injection, an intradermal injection, an intramuscular injection, or a drip injection), a nasal drop, or a suppository.

[0076] The administration interval of the prophylactic and / or therapeutic agent for autoimmune diseases according to one embodiment of the present invention is not limited, and may be, for example, once every hour, once every 2 hours, once every 3 hours, once every 6 hours, once every 12 hours, once every day, once every 2 days, once every 3 days, once every 4 days, once every 5 days, once every 6 days, once every week, once every 2 weeks, once every 3 weeks, once every month, once every 2 months, once every 3 months, once every 4 months, once every 5 months, or once every 6 months.

[0077] The subjects of administration of the prophylactic and / or therapeutic agent for autoimmune diseases according to one embodiment of the present invention are not limited, and examples thereof include humans and non-human animals (e.g., livestock, pets, and laboratory animals). Examples of non-human animals include monkeys, chimpanzees, cows, pigs, sheep, goats, horses, dogs, cats, rabbits, mice, and rats.

[0078] [3. Other] The present invention can also be configured as follows.

[0079] <1> A method for preventing and / or treating an autoimmune disease, comprising the step of administering to a subject (e.g., a human or a non-human animal) an agent for preventing and / or treating an autoimmune disease, the agent comprising an anti-DOCK8 monoclonal antibody according to one embodiment of the present invention as an active ingredient.

[0080] <2> The autoimmune disease is systemic lupus erythematosus (SLE). <1> A method for preventing and / or treating an autoimmune disease according to the above. EXAMPLES

[0081] 1. Preparation of antigen The antigens were prepared according to known methods, and the outline of the antigen preparation is described below.

[0082] A DNA fragment encoding the DHR1 domain of mouse DOCK8 protein (amino acid sequence: SEQ ID NO: 24) was inserted into a vector for expressing a fusion protein with a His tag and GST. The vector was used to transform E. coli, and the DHR1 domain-His tag-GST fusion protein was expressed in the E. coli.

[0083] The E. coli was disrupted in a lysis buffer, and the lysis buffer was then centrifuged to separate the precipitate from the supernatant, and the supernatant was obtained.

[0084] The supernatant was applied to a glutathione sepharose column to bind the DHR1 domain-His tag-GST fusion protein to the glutathione sepharose column, which was then washed with a washing buffer to remove unnecessary substances from the glutathione sepharose column.

[0085] A protease was added to the washed glutathione Sepharose column to cleave the bond between the His tag and GST in the DHR1 domain-His tag-GST fusion protein, and the DHR1 domain-His tag fusion protein was eluted from the glutathione Sepharose column.

[0086] The eluate containing the DHR1 domain-His tag fusion protein was applied to a gel filtration column to obtain a highly pure DHR1 domain-His tag fusion protein, which was used as an antigen in the following tests.

[0087] 2. Phage display Phage display was performed according to a known method. A human antibody fragment (scFV) that specifically binds to the DHR1 domain was obtained by screening using the DHR1 domain-His tag fusion protein and a completely human antibody synthetic phage library as follows.

[0088] Through the first to fourth pannings, phages expressing scFV antibodies that bind to the DHR1 domain (amino acid sequence: SEQ ID NO: 24) of mouse DOCK8 protein were enriched.

[0089] Specifically, in the first panning, phages that bound to tosyl beads (Dynabeads (registered trademark) M-280 Tosylactivated, manufactured by Thermo Fisher Scientific) linked with the DHR1 domain of mouse DOCK8 protein were collected. In the second panning, the phages collected in the first panning were amplified, and then the phages were applied to new tosyl beads linked with the DHR1 domain of mouse DOCK8 protein (amino acid sequence: SEQ ID NO: 24), and the phages that bound to the tosyl beads were collected. In the third panning, the phages collected in the second panning were amplified, and then the phages were applied to new tosyl beads linked with the DHR1 domain of mouse DOCK8 protein (amino acid sequence: SEQ ID NO: 24), and the phages that bound to the tosyl beads were collected.

[0090] In the fourth panning, the phages recovered in the third panning were amplified and then subjected to an immunotube (Nunc immunotube, Thermo Fisher Scientific) linked to the DHR1 domain of mouse DOCK8 protein (amino acid sequence: SEQ ID NO: 24), and the phages that bound to the immunotube were recovered.

[0091] After amplifying the phages recovered in the fourth panning, the phages were subjected to ELISA (enzyme-linked immunosorbent assay) using the DHR1 domain of mouse DOCK8 protein (amino acid sequence: SEQ ID NO: 24) to screen for phages that bind to the DHR1 domain of mouse DOCK8 protein.

[0092] By the fourth panning, we succeeded in obtaining two phage clones containing at least the heavy chain CDR3 and the light chain CDR3 that bind to the DHR1 domain (amino acid sequence: SEQ ID NO: 24) of mouse DOCK8 protein. The amino acid sequences of the heavy chain CDR3 and the light chain CDR3 of each clone were as follows.

[0093] [Table 1]

[0094] <3. Preparation of IgG type monoclonal antibodies and Fc type monoclonal antibodies> Monoclonal antibodies were produced using the obtained clones 3 and 5. Clones 3 and 5 were used as IgG type monoclonal antibodies.

[0095] To prepare an expression plasmid for an IgG monoclonal antibody, the DNA sequences encoding the heavy and light chains were cloned into the pcDNA3.4 vector (Thermo Fisher Scientific).

[0096] Each of the prepared expression plasmids for the IgG monoclonal antibodies was introduced into Expi293F cells (Thermo Fisher Scientific) to express the IgG monoclonal antibodies.

[0097] According to a known method, an IgG type monoclonal antibody was purified using the recovered supernatant of Expi293F cells and MabSelectSuRe (Cytiva).

[0098] The IgG type monoclonal antibodies derived from clones 3 and 5 were designated monoclonal antibody 03.6 and monoclonal antibody 05.6, respectively.

[0099] The amino acid sequences of the heavy chain CDR1 to 3, light chain CDR1 to 3, heavy chain variable region, light chain variable region, heavy chain constant region, light chain constant region, heavy chain, and light chain of each monoclonal antibody were as follows.

[0100] [Table 2]

[0101] <4. Confirmation of the antigen recognition ability of monoclonal antibodies (ELISA)> The ability of the two monoclonal antibodies to recognize DOCK8 protein was confirmed by a known enzyme-linked immunosorbent assay (ELISA).

[0102] (i) the DHR1 domain of mouse DOCK8 protein (amino acid sequence: SEQ ID NO: 24) and (ii) SEAP (Secreted Alkaline Phosphatase) protein were immobilized on the substrate as targets for recognition by the monoclonal antibodies.

[0103] Solutions containing various concentrations of monoclonal antibodies were added onto the substrate, and the substrate was then washed with a washing solution to remove monoclonal antibodies that did not bind to the target of recognition.

[0104] An anti-human IgG antibody labeled with HRP (Peroxidase AffiniPure Donkey Anti-Human IgG (H+L) manufactured by Jackson ImmunoResearch) was added as a secondary antibody onto the substrate. The substrate was then washed with a washing solution. This removed the secondary antibody that did not bind to the monoclonal antibody.

[0105] The amount of resist IgG antibody bound to the substrate, in other words, the amount of monoclonal antibody bound to the substrate, was determined by measuring the absorbance at 450 nm.

[0106] The test results are shown in Figure 1. In Figure 1, "a mDHR1_03.6" and "a mDHR1_05.6" correspond to monoclonal antibody 03.6 and monoclonal antibody 05.6, respectively.

[0107] As is clear from 101 to 102 in FIG. 1, the two monoclonal antibodies bound to the antigen protein (i) in a monoclonal antibody concentration-dependent manner. On the other hand, the two monoclonal antibodies did not bind to the non-antigen protein (ii) at all. This demonstrated that the two monoclonal antibodies specifically recognized the DOCK8 protein.

[0108] <5. Confirmation of the antigen recognition ability of monoclonal antibodies (FCM)> The ability of the five monoclonal antibodies to recognize DOCK8 protein present on the cell surface and intracellularly was confirmed by known flow cytometry (FCM).

[0109] <5-1. DOCK8 protein present on the cell surface> The spleens were removed from mice aged 17 to 18 weeks. The spleens were left to stand at room temperature for 5 minutes in an ammonium chloride hemolytic agent, and then washed twice with a washing solution (1 mM EDTA / PBS) to collect splenocytes.

[0110] The splenocytes were subjected to blocking treatment using a blocking solution (10 μg / mL Anti-Mouse CD16 / CD32 / 5% mouse serum / 5% rat serum / 1% Fetal Bovine Serum / 1 mM EDTA / PBS).

[0111] A monoclonal antibody was added as a primary antibody to the splenocytes, and the cells were allowed to stand on ice for 20 minutes.

[0112] The splenocytes were washed with a washing solution (1% Fetal Bovine Serum / 1 mM EDTA / PBS) three times.

[0113] A secondary antibody (R-Phycoerythrin AffiniPure F(ab')2 Fragment Donkey Anti-Human IgG (H+L) manufactured by Jackson ImmunoResearch) was added to the above splenocytes, and the cells were allowed to stand on ice for 20 minutes.

[0114] The splenocytes were washed with a washing solution (1% Fetal Bovine Serum / 1 mM EDTA / PBS) three times.

[0115] The splenocytes were subjected to blocking treatment using a blocking solution (100 μg / mL human Ig / 5% mouse serum / 5% rat serum / 1% Fetal Bovine Serum / 1 mM EDTA / PBS).

[0116] To detect the T cell surface markers CD4 and CD3 of the splenocytes, an anti-CD4 antibody (BioLegend, Brilliant Violet 421 (registered trademark) anti-mouse CD4 Antibody) and an anti-CD3 antibody (Thermo Fisher Scientific, CD3e Monoclonal Antibody (145-2C11), FITC) were added and the cells were allowed to stand on ice for 20 minutes.

[0117] The splenocytes were washed with a washing solution (1% Fetal Bovine Serum / 1 mM EDTA / PBS) three times.

[0118] In the above series of treatments, the cell membrane of the splenocytes is not destroyed but maintained, and therefore, the DOCK8 protein present on the surface of the splenocytes (in other words, the outside of the splenocytes) is stained.

[0119] The splenocytes that had been subjected to the above-mentioned series of treatments were subjected to FCM using an LSR Fortessa X-20, and the number of stained splenocytes was counted.

[0120] The test results are shown in 201 of Figure 2. In addition, in 201 of Figure 2, "03.6" and "05.6" correspond to monoclonal antibodies 03.6 and 05.6, respectively. As shown in 201 of Figure 2, the two monoclonal antibodies did not recognize the DOCK8 protein present on the cell surface.

[0121] <5-2. DOCK8 protein present in cells> The spleens were removed from mice aged 17 to 18 weeks. The spleens were left to stand at room temperature for 5 minutes in an ammonium chloride hemolytic agent, and then washed twice with a washing solution (1 mM EDTA / PBS) to collect splenocytes.

[0122] The splenocytes were subjected to blocking treatment using a blocking solution (10 μg / mL Anti-Mouse CD16 / CD32 / 5% mouse serum / 5% rat serum / 1% Fetal Bovine Serum / 1 mM EDTA / PBS).

[0123] To detect the T cell surface markers CD4 and CD3 of the splenocytes, an anti-CD4 antibody (BioLegend, Brilliant Violet 421 (registered trademark) anti-mouse CD4 Antibody) and an anti-CD3 antibody (Thermo Fisher Scientific, CD3e Monoclonal Antibody (145-2C11), FITC) were added and the cells were allowed to stand on ice for 20 minutes.

[0124] The splenocytes were washed with a washing solution (1% Fetal Bovine Serum / 1 mM EDTA / PBS) three times.

[0125] The splenocytes were suspended in a fixation / permeabilization solution (BD Biosciences, BD Cytofix / Cytoperm (registered trademark) Fixation and Permeabilization Solution) and left to stand on ice for 20 minutes to perform fixation and permeabilization.

[0126] The splenocytes were washed using a washing solution (BD Biosciences, BD Perm / Wash (registered trademark) Perm / Wash Buffer). This washing process was performed twice.

[0127] The splenocytes were subjected to a blocking treatment using a blocking solution (5% mouse serum / 5% rat serum / BD Perm / Wash (registered trademark) Perm / Wash Buffer).

[0128] A monoclonal antibody was added as a primary antibody to the splenocytes, and the cells were allowed to stand on ice for 20 minutes.

[0129] The splenocytes were washed using a washing solution (BD Biosciences, BD Perm / Wash (registered trademark) Perm / Wash Buffer). This washing process was repeated three times.

[0130] A secondary antibody (R-Phycoerythrin AffiniPure F(ab')2 Fragment Donkey Anti-Human IgG (H+L) manufactured by Jackson ImmunoResearch) was added to the above splenocytes, and the cells were allowed to stand on ice for 20 minutes.

[0131] The splenocytes were washed using a washing solution (BD Biosciences, BD Perm / Wash (registered trademark) Perm / Wash Buffer). This washing process was repeated three times.

[0132] In the above series of treatments, the cell membrane of the splenocytes is destroyed, and therefore, the DOCK8 protein present in the splenocytes is stained.

[0133] The test results are shown in 202 of Figure 2. In addition, in 202 of Figure 2, "03.6" and "05.6" correspond to monoclonal antibodies 03.6 and 05.6, respectively. As shown in 202 of Figure 2, the two monoclonal antibodies recognized the DOCK8 protein present in cells.

[0134] <6. DOCK8-positive CD4 T cells in patients with autoimmune diseases (active disease)> We investigated whether DOCK8-positive CD4 T cells were present in spleens collected from patients with an autoimmune disease (systemic lupus erythematosus: SLE) in the active stage before starting treatment for the autoimmune disease.

[0135] The collected spleen was immersed in a 10% aqueous formalin solution for fixation, and then deparaffinized using xylene and alcohol.

[0136] The spleen was then treated with a biotinylated anti-DOCK8 monoclonal antibody (specifically, <3> Tissue staining was performed using anti-DOCK8 monoclonal antibody 03.6 (for DOCK8 staining) prepared by NIH, anti-IFNγ antibody (for T cell staining), alkaline phosphatase avidin D, and anti-Novolink antibody.

[0137] The results of the tissue staining are shown in 301 of Figure 3. As indicated by the arrow in 301 of Figure 3, there were many DOCK8-positive CD4 T cells in the spleen of patients with autoimmune diseases in the active stage of the disease. This suggests that DOCK8 is involved in autoimmune diseases.

[0138] The relationship between the ratio of the number of DOCK8-positive CD4 T cells to the number of CD4 T cells in the peripheral blood of patients with an autoimmune disease (systemic lupus erythematosus: SLE) and the SLE disease activity index was examined according to a known method.

[0139] The test results are shown in 302 of Figure 3 (test results for anti-DOCK8 monoclonal antibody 03.6) and in Figure 4 (test results for anti-DOCK8 monoclonal antibody 05.6). In 302 of Figure 3, "black circles" indicate test results for SLE patients. In 302 of Figure 3, "black squares" indicate test results for rheumatoid arthritis patients, "black triangles" indicate test results for connective tissue disorder patients, "white triangles" indicate test results for polymyositis patients, "black diamonds" indicate test results for systemic sclerosis patients, and "white squares" indicate test results for microscopic polyangiitis patients.

[0140] As shown in Figure 3, 302 and Figure 4, it was revealed that the ratio of the number of DOCK8-positive CD4 T cells to the number of CD4 T cells in the peripheral blood of patients with autoimmune diseases increased as the SLE activity index value increased, suggesting that DOCK8 is involved in autoimmune diseases.

[0141] 7. Pharmacological effects of monoclonal antibodies We repeatedly intraperitoneally administered ovalbumin (OVA: grade V; Sigma) to BALB / c mice (Charles River Japan Inc.) to generate mice exhibiting symptoms of autoimmune disease.

[0142] During the 15th to 24th week after the start of OVA administration, 100 μg of anti-DOCK8 monoclonal antibody (specifically, <3> Anti-DOCK8 monoclonal antibody 03.6 or 05.6 prepared in the above manner, or 100 μg of control rabbit IgG was administered intraperitoneally to BALB / c mice.

[0143] Between the 15th and 26th weeks after the start of OVA administration, we detected anti-Sm antibodies, anti-dsDNA antibodies, rheumatoid factor (RF), and proteinuria (albuminuria), which are indicators of autoimmune disease.

[0144] The test results are shown in Figure 5 (test results for anti-DOCK8 monoclonal antibody 03.6) and Figure 6 (anti-DOCK8 monoclonal antibody 05.6). As is clear from Figures 5 and 6, when the control rabbit IgG was administered, various indicators of autoimmune disease increased. On the other hand, when the anti-DOCK8 monoclonal antibody was administered, the increase in various indicators of autoimmune disease was significantly suppressed. This indicates that the anti-DOCK8 monoclonal antibody can be used as a preventive and / or therapeutic agent for autoimmune diseases. [Industrial Applicability]

[0145] One embodiment of the present invention may be used in the prevention and / or treatment of autoimmune diseases.

Claims

1. A preventive and / or therapeutic agent for systemic lupus erythematosus (SLE), comprising as an active ingredient an anti-DOCK8 monoclonal antibody having heavy chain CDR1-3 and light chain CDR1-3 as shown in (2) below: (2) Heavy chain CDR1 to 3 having the amino acid sequences of SEQ ID NOs: 7 to 9, and light chain CDR1 to 3 having the amino acid sequences of SEQ ID NOs: 10 to 12.

2. The preventive and / or therapeutic agent for systemic lupus erythematosus (SLE) according to claim 1, wherein the anti-DOCK8 monoclonal antibody has a heavy chain variable region and a light chain variable region shown in the following (2'): (2') a heavy chain variable region having an amino acid sequence consisting of SEQ ID NO:15, and a light chain variable region having an amino acid sequence consisting of SEQ ID NO:

16.

3. The anti-DOCK8 monoclonal antibody is a preventive and / or therapeutic agent for systemic lupus erythematosus (SLE) described in claim 1, having a heavy chain constant region having an amino acid sequence consisting of sequence number 17.

4. The anti-DOCK8 monoclonal antibody is a preventive and / or therapeutic agent for systemic lupus erythematosus (SLE) as described in claim 1, wherein the light chain constant region is a κ chain having an amino acid sequence consisting of SEQ ID NO:

19.

5. The preventive and / or therapeutic agent for systemic lupus erythematosus (SLE) according to claim 1, wherein the anti-DOCK8 monoclonal antibody has a heavy chain and a light chain shown in the following (2''): (2'') A heavy chain having an amino acid sequence consisting of SEQ ID NO:22, and a light chain having an amino acid sequence consisting of SEQ ID NO:23.

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