Anti-human IL-15 antibodies and their use

Abstract

The present invention relates to an antibody that binds to human IL-15, a nucleic acid molecule encoding the antibody, a vector containing the nucleic acid molecule, a host cell containing the nucleic acid molecule or vector, a method for producing and purifying the antibody, and the use of the antibody.

Description

[Technical Field]

[0001] (Cross-reference of related applications) This application claims priority to Chinese Patent Application No. 202310498953.4, filed on 5 May 2023, the entire contents of which are incorporated herein by reference.

[0002] This invention generally relates to the fields of genetic engineering and antibody drugs, and more specifically to antibodies that bind to human IL-15 and the use thereof. [Background technology]

[0003] The IL-15 gene is located on human chromosome 4q31 and consists of nine exons (1-8 and 4A) and eight introns, four of which exons (5-8) encode the mature protein. IL-15 protein expression is limited, tightly regulated during transcription and translation processes, and is primarily expressed in monocytes / macrophages and dendritic cells. [1] IL-15 is a four-alpha-helix bundle cytokine. The IL-15 receptor consists of IL-15Rα, IL-2Rβ, and γc. IL-15Rα is the specific receptor for IL-15, IL-2Rβ is the co-receptor for IL-2 and IL-15, and γc (CD132) is the common receptor for many cytokines, including IL-2, IL-4, IL-7, IL-9, and IL-21. IL-15Rα binds to IL-15 to form a stable, high-affinity complex, which is then presented to IL-2Rβ and γc on activated T cells or NK cells to form high-affinity immune synapses. [2] By sharing receptor subunits (IL-2Rβ and γc), IL-2 and IL-15 trigger several similar downstream signaling pathways, including Janus kinase (JAK) signaling proteins and activating transcription proteins (STAT), exerting physiological functions such as promoting proliferation and suppressing apoptosis.

[0004] Vitiligo is a chronic autoimmune disease characterized by loss of skin pigmentation, resulting in white patches of various shapes and sizes appearing on the skin and mucous membranes. This is caused by the loss and destruction of pigment-producing cells—melanocytes—and affects cosmetic appearance. Vitiligo affects approximately 0.5% to 2.0% of the world's population, with no significant difference between sexes, and its incidence varies by geographical region. Vitiligo is classified into segmental vitiligo, unsegmental vitiligo, and mixed vitiligo. Unsegmental vitiligo accounts for approximately 87% of patients under the age of 30, and is bilaterally symmetrical and distributed throughout the body. [3] .

[0005] JM Richmond's research group found that CD8 in skin tissue. + We discovered the existence of tissue-resident memory T cells (TRm), which can remain in the tissue for at least six months after formation and increase in inflammatory environments. Biopsies of lesional skin from vitiligo patients confirmed the presence of large amounts of active TRm in the lesional tissue. [4] Vitiligo often recurs in the same area, suggesting that TRm plays a crucial role in disease progression. IL-15 is a cytokine necessary for the long-term survival of TRm; IL-15 binds to receptors on the surface of TRm cells, maintaining TRm survival and stimulating TRm to secrete IFN-γ / granzyme / perforin, damaging melanocytes. Simultaneously, IFN-γ can also activate the downstream JAK-STAT pathway, causing skin keratinocytes to secrete CXCL9 and CXCL10, leading to more melanocyte-specific CD8 + It mobilizes T cells and increases inflammation through a positive feedback loop. [5] IL-15 is a potential new target for vitiligo treatment.

[0006] Therefore, the development of functional antibodies against IL-15 is important for the treatment of IL-15-mediated diseases. [Overview of the project]

[0007] According to a first aspect, the present invention is an antibody that binds to human IL-15, comprising a heavy chain variable region containing the amino acid sequences of HCDR1, HCDR2, and HCDR3, and a light chain variable region containing the amino acid sequences of LCDR1, LCDR2, and LCDR3, The amino acid sequence of HCDR1 is shown in SEQ ID NO:26, the amino acid sequence of HCDR2 is shown in SEQ ID NO:27, the amino acid sequence of HCDR3 is shown in SEQ ID NO:28, the amino acid sequence of LCDR1 is shown in SEQ ID NO:34, the amino acid sequence of LCDR2 is shown in SEQ ID NO:35, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:36. The amino acid sequence of HCDR1 is shown in SEQ ID NO:26, the amino acid sequence of HCDR2 is shown in SEQ ID NO:27, the amino acid sequence of HCDR3 is shown in SEQ ID NO:29, the amino acid sequence of LCDR1 is shown in SEQ ID NO:37, the amino acid sequence of LCDR2 is shown in SEQ ID NO:38, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:39.

[0008] The amino acid sequence of HCDR1 is shown in SEQ ID NO:26, the amino acid sequence of HCDR2 is shown in SEQ ID NO:27, the amino acid sequence of HCDR3 is shown in SEQ ID NO:30, the amino acid sequence of LCDR1 is shown in SEQ ID NO:37, the amino acid sequence of LCDR2 is shown in SEQ ID NO:38, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:39. The amino acid sequence of HCDR1 is shown in SEQ ID NO:31, the amino acid sequence of HCDR2 is shown in SEQ ID NO:32, the amino acid sequence of HCDR3 is shown in SEQ ID NO:33, the amino acid sequence of LCDR1 is shown in SEQ ID NO:40, the amino acid sequence of LCDR2 is shown in SEQ ID NO:41, the amino acid sequence of LCDR3 is shown in SEQ ID NO:42, or, The amino acid sequence of HCDR1 is shown in SEQ ID NO:31, the amino acid sequence of HCDR2 is shown in SEQ ID NO:45, the amino acid sequence of HCDR3 is shown in SEQ ID NO:46, the amino acid sequence of LCDR1 is shown in SEQ ID NO:47, the amino acid sequence of LCDR2 is shown in SEQ ID NO:48, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:49. Here, the amino acid sequences of HCDR and LCDR are provided, as defined by Kabat, to provide the antibodies.

[0009] In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is indicated by SEQ ID NO: 13, 19, 21, 22, or 43. In some embodiments of the first aspect, the amino acid sequence of the light chain variable region of the antibody is indicated by SEQ ID NO: 14, 20, 23, or 44.

[0010] In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:13, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:14. The amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:19, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:20. The amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:21, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:20. The amino acid sequence of the heavy chain variable region of the antibody is shown in SEQ ID NO:22, and the amino acid sequence of the light chain variable region of the antibody is shown in SEQ ID NO:23, or, The amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO:43, and the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:44.

[0011] According to a second aspect, the present invention provides an antibody that binds to human IL-15, wherein the amino acid sequence of the heavy chain variable region of the antibody has at least 90% identity with any one of SEQ ID NO: 13, 19, 21, 22, and 43, and the amino acid sequence of the light chain variable region of the antibody has at least 90% identity with any one of SEQ ID NO: 14, 20, 23, and 44.

[0012] In some embodiments of the first and second aspects, the antibody is a full antibody, Fab fragment, F(ab’)2 fragment, or single-chain Fv fragment (scFv), and / or the antibody is a monoclonal antibody, and / or the antibody further comprises a heavy chain constant region selected from IgG1 subtype, IgG2 subtype, or IgG4 subtype, and / or the 234th, 235th, and 331st amino acids of the Fc fragment are F, E, and S, respectively, and / or the 252nd, 254th, and 256th amino acids of the heavy chain constant region are Y, T, and E, respectively, and / or the antibody further comprises a light chain constant region selected from κ subtype or λ subtype, where the amino acid positions of the constant region of the antibody are determined according to EU numbering.

[0013] In some embodiments of the first and second aspects, the antibody binds to human IL-15 and / or simian IL-15, and / or the antibody can inhibit the activity of IL-15. According to a third aspect, the present invention provides a nucleic acid molecule encoding the antibody described in the first or second aspect. According to a fourth aspect, the present invention provides a pharmaceutical composition comprising the antibody described in the first or second aspect and a pharmaceutically acceptable excipient, diluent, or carrier.

[0014] According to a fifth aspect, the present invention provides the use of the antibody according to the first or second aspect, or the pharmaceutical composition according to the fourth aspect, in the manufacture of a medicament for preventing or treating an IL-15-mediated disease. According to a sixth aspect, the present invention provides a method for preventing or treating an IL-15-mediated disease, comprising administering to an individual in need thereof the antibody according to the first or second aspect, or the pharmaceutical composition according to the fourth aspect. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] [Figure 1] It is a diagram showing the binding ability of an anti-human IL-15 monoclonal antibody to IL-15 of different species. [Figure 2] It is a diagram showing the results of epitope analysis of an anti-human IL-15 monoclonal antibody. Here, FIG. 2A shows the results of blocking the binding of an anti-human IL-15 monoclonal antibody to AMG714 purified phage and human IL-15, and FIG. 2B shows the results of blocking the binding of an anti-human IL-15 monoclonal antibody to CALY-002 purified phage and human IL-15. [Figure 3] It is a diagram showing the activity results of an anti-human IL-15 monoclonal antibody against HEK-blue IL-2 cells. [Figure 4] It is a diagram showing the activity results of an anti-human IL-15 monoclonal antibody against NK-92 cells. MODE FOR CARRYING OUT THE INVENTION

[0016] 〔Description of Sequences〕 SEQ ID NO:1 shows the amino acid sequence of the extracellular region of human (homo sapiens) IL-15 (hIL-15). SEQ ID NO:2 shows the amino acid sequence of the extracellular region of human (homo sapiens) IL-15Ra (hIL-15Ra). SEQ ID NO:3 shows the amino acid sequence of the extracellular domain of IL-15 (mfIL-15) in cynomolgus monkeys (Macaca fascicularis). SEQ ID NO:4 shows the amino acid sequence of the extracellular domain of IL-15Ra (mfIL-15Ra) in cynomolgus monkeys (Macaca fascicularis). SEQ ID NO:5 shows the amino acid sequence of the extracellular domain (mIL-15) of mouse (Mus musculus).

[0017] SEQ ID NO:6 shows the amino acid sequence of the extracellular domain (mIL-15Ra) of mouse (Mus musculus). SEQ ID NO:7 indicates the amino acid sequence of the His tag (His). SEQ ID NO:8 shows the amino acid sequence of the Fc segment (mFc1) of the mouse antibody IgG1. SEQ ID NO:9 shows the amino acid sequence of the constant region of the heavy chain of the human (homo sapiens) IgG1 subtype. SEQ ID NO:10 shows the amino acid sequence of the constant region of the heavy chain of the human IgG1m3 subtype.

[0018] SEQ ID NO:11 shows the amino acid sequence of the constant region of the human (homo sapiens) κ subtype light chain. SEQ ID NO:12 shows the amino acid sequence of the constant region of the light chain of the human (homo sapiens) λ subtype. SEQ ID NO:13 shows the amino acid sequence of the heavy chain variable region R1A3VH of the anti-human IL-15 monoclonal antibody R1A3. SEQ ID NO:14 shows the amino acid sequence of the light chain variable region R1A3VK of the anti-human IL-15 monoclonal antibody R1A3. SEQ ID NO:15 shows the amino acid sequence of the heavy chain variable region of the anti-human IL-15 monoclonal antibody AMG714.

[0019] SEQ ID NO:16 shows the amino acid sequence of the light chain variable region of the anti-human IL-15 monoclonal antibody AMG714. SEQ ID NO:17 shows the amino acid sequence of the heavy chain variable region of the anti-human IL-15 monoclonal antibody CALY-002. SEQ ID NO:18 shows the amino acid sequence of the light chain variable region of the anti-human IL-15 monoclonal antibody CALY-002. SEQ ID NO:19 shows the amino acid sequence of the heavy chain variable region R4G4VH of the anti-human IL-15 monoclonal antibody R4G4VH+R22F11VK. SEQ ID NO:20 shows the amino acid sequence of the light chain variable region R22F11VK of the anti-human IL-15 monoclonal antibodies R4G4VH+R22F11VK and R2H2VH+R22F11VK.

[0020] SEQ ID NO:21 shows the amino acid sequence of the heavy chain variable region R2H2VH of the anti-human IL-15 monoclonal antibody R2H2VH+R22F11VK. SEQ ID NO:22 shows the amino acid sequence of the heavy chain variable region R26H10VH of the anti-human IL-15 monoclonal antibody R26H10. SEQ ID NO:23 shows the amino acid sequence of the light chain variable region R26H10VK of the anti-human IL-15 monoclonal antibody R26H10. SEQ ID NO:24 shows the nucleotide sequence of primer PmCGR. SEQ ID NO:25 shows the nucleotide sequence of primer PmCKR.

[0021] SEQ ID NO:26 shows the amino acid sequences of the heavy chain variable regions R1A3VH, R4G4VH, and R2H2VH of HCDR1. SEQ ID NO:27 shows the amino acid sequences of the heavy chain variable regions R1A3VH, R4G4VH, and R2H2VH of HCDR2. SEQ ID NO:28 shows the amino acid sequence of HCDR3 in the heavy chain variable region R1A3VH. SEQ ID NO:29 shows the amino acid sequence of HCDR3 in the heavy chain variable region R4G4VH. SEQ ID NO:30 shows the amino acid sequence of HCDR3 in the heavy chain variable region R2H2VH.

[0022] SEQ ID NO:31 shows the amino acid sequence of HCDR1 in the heavy chain variable regions R26H10VH and R6E11VH. SEQ ID NO:32-33 each represent the amino acid sequences of HCDR2 and HCDR3 in the heavy chain variable region R26H10VH. SEQ ID NO:34-36 each represent the amino acid sequences of LCDR1, LCDR2, and LCDR3 in the light chain variable region R1A3VK. SEQ ID NO:37-39 each represent the amino acid sequences of LCDR1, LCDR2, and LCDR3 in the light chain variable region R22F11VK. SEQ ID NO:40~42 each represents the amino acid sequences of LCDR1, LCDR2, and LCDR3 in the light chain variable region R26H10VK. SEQ ID NO:43 shows the amino acid sequence of the heavy chain variable region R6E11VH of the anti-human IL-15 monoclonal antibody R6E11.

[0023] SEQ ID NO:44 shows the amino acid sequence of the light chain variable region R6E11VK of the anti-human IL-15 monoclonal antibody R6E11. SEQ ID NO:45-46 each represent the amino acid sequences of HCDR2 and HCDR3 in the heavy chain variable region R6E11VH. SEQ ID NO:47-49 each represent the amino acid sequences of LCDR1, LCDR2, and LCDR3 in the light chain variable region R6E11VK. SEQ ID NO:50 indicates the amino acid sequence of the heavy chain variable region of the DP47 antibody. SEQ ID NO:51 shows the amino acid sequence of the light chain variable region of the DP47 antibody.

[0024] The inventors of the present invention have obtained a novel anti-human IL-15 antibody by antibody engineering technology. Various aspects of the present invention provide a novel anti-human IL-15 antibody, a nucleic acid molecule encoding the antibody, a vector containing the nucleic acid molecule, a host cell containing the nucleic acid molecule or vector, a method for producing and purifying the antibody, and the medical and biological use of the antibody. According to the amino acid sequence of the variable region of the antibody according to the present invention, a full-length antibody molecule can be constructed as a drug for preventing or treating human IL-15-mediated diseases.

[0025] Unless otherwise specified, the implementation of this invention utilizes techniques of molecular biology, microbiology, cell biology, biochemistry, and immunology that are conventional in the art. Unless otherwise specified, terms used in this application have the meanings generally understood by those skilled in the art.

[0026] [Definition] As used herein, the term “antibody” refers to an immunoglobulin molecule that can specifically bind to a target via at least one antigen-recognition site located in the variable region of the immunoglobulin molecule. Targets include, but are not limited to, carbohydrates, polynucleotides, lipids, polypeptides, etc. As used herein, “antibody” includes not only intact antibodies (i.e., full-length antibodies) but also their antigen-binding fragments (e.g., Fab, Fab', F(ab')2, Fv), their variants, fusion proteins containing the antibody moiety, humanized antibodies, chimeric antibodies, bivalent antibodies, linear antibodies, single-chain antibodies, multispecific antibodies (e.g., bispecific antibodies), and any other modified configurations of immunoglobulin molecules containing an antigen-recognition site with the desired specificity, such as glycosylated variants of antibodies, amino acid sequence variants of antibodies, and covalently modified antibodies.

[0027] Typically, intact antibodies or full-length antibodies contain two heavy chains and two light chains. Each heavy chain contains a heavy chain variable region (VH) and first, second, and third constant regions (CH1, CH2, and CH3). Each light chain contains a light chain variable region (VL) and a constant region (CL). Full-length antibodies may be antibodies of any class, such as IgD, IgE, IgG, IgA, or IgM (or the subclasses listed above), but antibodies do not need to belong to any particular class. Depending on the antibody amino acid sequence of the heavy chain constant domain, immunoglobulins can be assigned to various classes. Typically, there are five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM. Some of these classes can be further classified into subclasses (isotypes), such as IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant domains corresponding to different immunoglobulin classes are called α, δ, ε, γ, and μ, respectively. The subunit structures and three-dimensional structures of different classes of immunoglobulins are publicly known.

[0028] As used herein, the term “antigen-binding fragment or antigen-binding moiety” refers to a portion or region of an intact antibody molecule involved in binding to an antigen. The antigen-binding domain may include a heavy chain variable region (VH), a light chain variable region (VL), or both. Each of the VH and VL typically contains three complementarity-determining regions: CDR1, CDR2, and CDR3.

[0029] Those skilled in the art know that the complementarity-determining regions (CDRs, typically CDR1, CDR2, and CDR3) are the regions within the variable region that have the greatest influence on antibody affinity and specificity. There are two common methods for defining the CDR amino acid sequences of VH or VL: the Chothia definition and the Kabat definition. For example, Kabat's definition is found in "Sequences of Proteins of Immunological Interest," National Institutes of Health, Bethesda, Md. (1991). 7", "A1-Lazikani et al, J. Mol. Biol. 273:927-948 (1997) 8 ", and "Martin et al, Proc. Natl. Acad. Sci.USA86:9268-9272 (1989) 9 See ". For the variable region amino acid sequence of a given antibody, the CDR amino acid sequences in the VH and VL amino acid sequences can be determined according to the Chothia definition or the Kabat definition. In an embodiment of the present invention, Kabat is used to define the CDR amino acid sequence.

[0030] For the variable region amino acid sequence of a given antibody, the CDR amino acid sequences of the variable region amino acid sequence can be analyzed in various ways. For example, the online software Abysis (http: / / www.abysis.org / ) can be used.

[0031] Actual examples of antigen-binding fragments include, but are not limited to, the following five examples: (1) a Fab fragment, which may be a monovalent fragment having a VL-CL chain and a VH-CH1 chain; (2) an F(ab’)2 fragment, which may be a bivalent fragment having two Fab’ fragments linked by a disulfide bridge in the hinge region (i.e., a dimer of Fab’); (3) an Fv fragment having the VL and VH domains of one arm of an antibody; (4) a single-chain Fv (scFv), which may be a single polypeptide chain composed of a VH domain and a VL domain via a peptide linker; (5) (scFv)2, which may include two VH domains linked by a peptide linker and two VL domains bound to the two VH domains via disulfide bridges.

[0032] As used herein, the term "specific binding" refers to a non-random binding reaction between two molecules, such as the binding of an antibody to an antigen epitope. As used herein, the term “monoclonal antibody” refers to an antibody obtained from a substantially homogeneous antibody population; that is, the individual antibodies constituting the population are identical except for naturally occurring mutations that may be present in a small number of individuals.

[0033] As used herein, the term "humanized antibody" refers to an antibody in which the constant region (i.e., the CH region and CL region) or the entire antibody is encoded by a human antibody gene. Humanized antibodies can significantly reduce the immunological side effects of heterologous antibodies on the human body. Humanized antibodies include chimeric antibodies, modified antibodies, and fully humanized antibodies.

[0034] According to a first aspect, the present invention provides an antibody that binds to human IL-15, comprising a heavy chain variable region containing the amino acid sequences of HCDR1, HCDR2, and HCDR3, and a light chain variable region containing the amino acid sequences of LCDR1, LCDR2, and LCDR3. The amino acid sequence of HCDR1 is shown in SEQ ID NO:26, the amino acid sequence of HCDR2 is shown in SEQ ID NO:27, the amino acid sequence of HCDR3 is shown in SEQ ID NO:28, the amino acid sequence of LCDR1 is shown in SEQ ID NO:34, the amino acid sequence of LCDR2 is shown in SEQ ID NO:35, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:36. The amino acid sequence of HCDR1 is shown in SEQ ID NO:26, the amino acid sequence of HCDR2 is shown in SEQ ID NO:27, the amino acid sequence of HCDR3 is shown in SEQ ID NO:29, the amino acid sequence of LCDR1 is shown in SEQ ID NO:37, the amino acid sequence of LCDR2 is shown in SEQ ID NO:38, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:39.

[0035] The amino acid sequence of HCDR1 is shown in SEQ ID NO:26, the amino acid sequence of HCDR2 is shown in SEQ ID NO:27, the amino acid sequence of HCDR3 is shown in SEQ ID NO:30, the amino acid sequence of LCDR1 is shown in SEQ ID NO:37, the amino acid sequence of LCDR2 is shown in SEQ ID NO:38, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:39. The amino acid sequence of HCDR1 is shown in SEQ ID NO:31, the amino acid sequence of HCDR2 is shown in SEQ ID NO:32, the amino acid sequence of HCDR3 is shown in SEQ ID NO:33, the amino acid sequence of LCDR1 is shown in SEQ ID NO:40, the amino acid sequence of LCDR2 is shown in SEQ ID NO:41, the amino acid sequence of LCDR3 is shown in SEQ ID NO:42, or, The amino acid sequence of HCDR1 is shown in SEQ ID NO:31, the amino acid sequence of HCDR2 is shown in SEQ ID NO:45, the amino acid sequence of HCDR3 is shown in SEQ ID NO:46, the amino acid sequence of LCDR1 is shown in SEQ ID NO:47, the amino acid sequence of LCDR2 is shown in SEQ ID NO:48, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:49. Here, the amino acid sequences of HCDR and LCDR are defined by Kabat.

[0036] In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO:13. In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is indicated by SEQ ID NO:19. In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is indicated by SEQ ID NO:21.

[0037] In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO:22. In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO:43. In some embodiments of the first aspect, the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:14.

[0038] In some embodiments of the first aspect, the amino acid sequence of the light chain variable region of the antibody is indicated by SEQ ID NO:20. In some embodiments of the first aspect, the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:23. In some embodiments of the first aspect, the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:44.

[0039] In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:13, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:14. In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:19, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:20. In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:21, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:20.

[0040] In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:22, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:23. In some embodiments of the first aspect, the amino acid sequence of the heavy chain variable region of the antibody is shown by SEQ ID NO:43, and the amino acid sequence of the light chain variable region of the antibody is shown by SEQ ID NO:44. According to a second aspect, the present invention provides an antibody that binds to human IL-15, wherein the amino acid sequence of the heavy chain variable region of the antibody has at least 90% identity with any one of SEQ ID NO: 13, 19, 21, 22, and 43, and the amino acid sequence of the light chain variable region of the antibody has at least 90% identity with any one of SEQ ID NO: 14, 20, 23, and 44.

[0041] In some embodiments of the second aspect, the amino acid sequence of the heavy chain variable region of the antibody has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more identity with any one of SEQ ID NO: 13, 19, 21, 22, and 43. In some embodiments of the second aspect, the amino acid sequence of the light chain variable region of the antibody has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more identity with any one of SEQ ID NO: 14, 20, 23, and 44.

[0042] In some embodiments of the second aspect, the amino acid sequence of the heavy chain variable region of the antibody differs from the amino acid sequence shown in any one of SEQ ID NO: 13, 19, 21, 22, and 43 by about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, and / or additions. In some embodiments of the second aspect, the amino acid sequence of the light chain variable region of the antibody differs from the amino acid sequence shown in any one of SEQ ID NO: 14, 20, 23, and 44 by about 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions, deletions, and / or additions.

[0043] In some embodiments of the second aspect, the C-terminal or N-terminal region of the amino acid sequence shown in any one of SEQ ID NO: 13, 19, 21, 22, and 43 may be further truncated by about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25 or more amino acids, and a function similar to that of the heavy chain variable region of the antibody is still maintained. In some embodiments of the second aspect, one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, four, fifteen, twenty, twenty, fifteen or more amino acids may be further added to the C-terminal or N-terminal region of the amino acid sequence shown in any one of SEQ ID NO: 13, 19, 21, 22, and 43, and the resulting amino acid sequence still maintains a function similar to that of the heavy chain variable region of the antibody. In some embodiments of the second aspect, the modified amino acid sequence may have 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25 or more amino acids added or deleted in a region other than the C-terminus or N-terminus of the amino acid sequence shown in any one of SEQ ID NO: 13, 19, 21, 22, and 43, as long as the modified amino acid sequence substantially maintains a function similar to that of the heavy chain variable region of the antibody. In some embodiments of the second aspect, the C-terminal or N-terminal region of the amino acid sequence represented by any one of SEQ ID NO: 14, 20, 23, and 44 may be further cleaved by about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25 or more amino acids, and a function similar to that of the light chain variable region of the antibody is still maintained. In some embodiments of the second aspect, one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, four, fifteen, twenty, twenty, fifteen, or more amino acids may be further added to the C-terminal or N-terminal region of the amino acid sequence represented by any one of SEQ ID NO: 14, 20, 23, and 44, and the resulting amino acid sequence still maintains a function similar to that of the light chain variable region of the antibody.

[0044] In some embodiments of the second aspect, the modified amino acid sequence may have 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25 or more amino acids further added or deleted from a region other than the C-terminus or N-terminus of the amino acid sequence shown in any one of SEQ ID NO: 14, 20, 23, and 44, insofar as the modified amino acid sequence substantially maintains a function similar to that of the light chain variable region of the antibody. In some embodiments of the first and second embodiments, the antibody is a complete antibody, a Fab fragment, an F(ab')2 fragment, or a single-stranded Fv fragment (scFv). In some embodiments of the first and second embodiments, the antibody is a fully human antibody or a humanized antibody. In some embodiments of the first and second embodiments, the antibody is a monoclonal antibody. In some embodiments of the first and second embodiments, the antibody further comprises a heavy chain constant region selected from the IgG1 subtype, the IgG2 subtype, or the IgG4 subtype.

[0045] In some specific embodiments of the first and second embodiments, the heavy chain constant region is an IgG1 subtype, for example, an IgG1m3 subtype. In some embodiments of the first and second embodiments, the amino acids at positions 234, 235, and 331 of the Fc fragment are F, E, and S, respectively, where the amino acid positions of the antibody constant region are determined according to EU numbering. In some embodiments of the first and second embodiments, the amino acids at positions 252, 254, and 256 of the heavy chain constant region are Y, T, and E, respectively, where the amino acid positions of the antibody constant region are determined according to EU numbering. In some embodiments of the first and second embodiments, the antibody further comprises a light chain constant region selected from a κ subtype or a λ subtype. In some specific embodiments of the first and second embodiments, the antibody further comprises a light chain constant region selected from the κ subtype.

[0046] In some embodiments of the first and second embodiments, the antibody binds to human IL-15. In some specific embodiments of the first and second embodiments, the antibody binds to recombinant human IL-15 (SEQ ID NO:1). In some embodiments of the first and second embodiments, the antibody binds to monkey IL-15. In some specific embodiments of the first and second embodiments, the antibody binds to recombinant monkey IL-15 (SEQ ID NO:3). In some embodiments of the first and second embodiments, the antibody binds to an IL-15 and IL-15α complex, such as a complex of human IL-15 and human IL-15α, or a complex of monkey IL-15 and monkey IL-15α.

[0047] In some embodiments of the first and second embodiments, the antibody can inhibit the activity of IL-15. In some specific embodiments of the first and second embodiments, the antibody inhibits IL-15's ability to induce the secretion of secretory embryonic alkaline phosphatase (SEAP). In some specific embodiments of the first and second embodiments, the antibody inhibits IL-15's ability to induce proliferation of NK92 cells. In some embodiments of the first and second embodiments, the antibody does not bind to mouse IL-15, such as recombinant mouse IL-15 (SEQ ID NO: 5). According to a third aspect, the present invention provides a nucleic acid molecule encoding an antibody according to the first or second aspect.

[0048] In some embodiments of the third aspect, the nucleic acid molecule may include DNA molecules and RNA molecules. The nucleic acid molecule may be single-stranded or double-stranded, or it may be cDNA. In some embodiments of the third aspect, the nucleic acid molecule is operably linked to a regulatory amino acid sequence, which can be identified by a host cell transformed with the vector. According to a fourth aspect, the present invention provides a pharmaceutical composition comprising an antibody according to the first or second aspect and a pharmaceutically acceptable excipient, diluent, or carrier. In some embodiments of the fourth aspect, the pharmaceutical composition is used to prevent or treat IL-15-mediated diseases. In some embodiments of the fourth aspect, the IL-15-mediated disease is selected from vitiligo, celiac disease, eosinophilic esophagitis, and macrogranular lymphocytic leukemia.

[0049] In some embodiments of the fourth aspect, the pharmaceutical composition may further contain one or more of the following: lubricants (e.g., talc, magnesium stearate, and mineral oil), wetting agents, emulsifiers, suspending agents, preservatives (e.g., benzoic acid, sorbic acid, calcium propionate), sweeteners and / or flavoring agents. In some embodiments of the fourth aspect, the pharmaceutical composition according to the present invention may be prepared in the form of tablets, pills, powders, lozenges, elixirs, suspensions, emulsions, solutions, syrups, suppositories, or capsules. In some embodiments of the fourth aspect, the pharmaceutical composition according to the present invention can be delivered by any physiologically acceptable method of administration (including, but not limited to, oral administration, parenteral administration, nasal administration, rectal administration, intraperitoneal administration, intravascular injection, subcutaneous administration, transdermal administration, inhalation administration, etc.).

[0050] In some embodiments of the fourth aspect, the therapeutic pharmaceutical composition can be formulated for storage in the form of a lyophilized preparation or an aqueous solution by mixing a reagent of the required purity with any pharmaceutically acceptable carrier, excipient, etc. According to the fifth aspect, the present invention provides the use of an antibody according to the first or second aspect, or a pharmaceutical composition according to the fourth aspect, in the manufacture of a pharmaceutical for the prevention or treatment of IL-15-mediated diseases.

[0051] In some embodiments of the fifth aspect, the IL-15-mediated disease is selected from vitiligo, celiac disease, eosinophilic esophagitis, and macrogranular lymphocytic leukemia. According to the sixth aspect, the present invention provides a method for preventing or treating an IL-15-mediated disease, comprising administering an antibody described in the first or second aspect, or a pharmaceutical composition described in the fourth aspect, to an individual in need thereof. In some embodiments of the sixth aspect, the IL-15-mediated disease is selected from vitiligo, celiac disease, eosinophilic esophagitis, and macrogranular lymphocytic leukemia.

[0052] In other embodiments, the present invention further provides a vector comprising the nucleic acid molecule, a host cell comprising the nucleic acid molecule or the vector, and a method for producing the antibody. In some embodiments, the method for producing the antibody comprises culturing the host cell to facilitate nucleic acid expression. In some embodiments, the method for producing the antibody further comprises recovering the antibody from the host cell culture medium.

[0053] The above detailed description is intended only to make the present invention clearly understandable to those skilled in the art, and should be understood not to limit any aspect. Those skilled in the art can make various modifications and changes to the above embodiments. [Examples]

[0054] The following examples illustrate, but do not limit, the scope of the present invention.

[0055] Example 1: Preparation of recombinant protein The process of preparing and identifying IL-15-specific antibodies requires the use of various different recombinant proteins, including superagonists composed of co-expression of human IL-15 (hIL15, SEQ ID NO:1) and human IL-15Rα (hIL-15Rα, SEQ ID NO:2), superagonists composed of co-expression of monkey IL-15 (mfIL-15, SEQ ID NO:3) and monkey IL-15Rα (mfIL-15Rα, SEQ ID NO:4), and superagonists composed of co-expression of mouse IL-15 (mIL-15, SEQ ID NO:5) and mouse IL-15Rα (mIL-15Rα, SEQ ID NO:6). Adding a His tag (His, SEQ ID NO:7) or the Fc segment of mouse antibody IgG1 (mFc1, SEQ ID NO:8) to the C-terminus of IL-15Rα is advantageous for the purification and functional identification of recombinant proteins. When preparing recombinant antibodies, the antibody heavy chain constant region may be a human IgG1 subtype (SEQ ID NO:9) or various variants of a selected human IgG1 subtype (e.g., IgG1m3 (SEQ ID NO:10)), and the light chain constant region may be a human κ subtype (SEQ ID NO:11) or a human λ subtype (SEQ ID NO:12).

[0056] Based on the amino acid sequences of recombinant proteins in the Uniprot database, genes for various recombinant proteins (including His or mFc1 tags) were designed and synthesized. Using standard molecular biology techniques, the synthesized recombinant protein genes were cloned into appropriate eukaryotic expression vectors (e.g., Invitrogen's pcDNA3.1). Subsequently, the prepared recombinant protein expression plasmids were transfected into HEK293 cells (e.g., Invitrogen's HEK293F) using liposomes (e.g., Invitrogen's 293fectin) or other cationic transfection reagents (e.g., PEI), and cultured under serum-free suspension conditions for 3-4 days. The culture supernatant was then obtained by centrifugation or other methods.

[0057] Recombinant proteins expressed by His tag fusion in the culture supernatant were purified in a single step using a metal chelate affinity chromatography column (e.g., GE's HisTrap FF). Recombinant proteins expressed by antibody-Fc fusion were purified in a single step using a Protein A / G affinity chromatography column (e.g., GE's Mabselect SURE). Subsequently, the recombinant protein storage buffer was changed to PBS (pH 7.0) or another suitable buffer using a desalting column (e.g., GE's Hitrap desaulting). After filter sterilization, the samples were packaged and stored at -20°C for later use.

[0058] Example 2: Screening of anti-hIL-15 monoclonal antibodies from a fully human Fab phage library 2.1 Screening of a fully human Fab library The recombinant human IL-15+IL-15Rα superagonist prepared in Example 1 was used as the antigen, and a solid-phase screening strategy was employed (for the experimental scheme, see "Phage Display: A Practical Approach" edited by Clackson, T. and Lowman, H.B., translated by Ma Lan et al., Chemical Industry Press, May 2008). [6] Using this method, a prepared complete human Fab phage library was screened (see Chinese Patent Application No. 202210871809.6). [7] A total of three rounds of screening were performed, involving binding, elution, neutralization, infection, and amplification, ultimately yielding one strain of monoclonal antibody R1A3 (the amino acid sequence of the heavy chain variable region R1A3VH is shown in SEQ ID NO:13, and the amino acid sequence of the light chain variable region R1A3VK is shown in SEQ ID NO:14) that specifically binds to human IL-15.

[0059] Using conventional molecular biological techniques, R1A3 was prepared as a complete antibody of the IgG1m3 subtype, and patent patent No. 7153507B2 was issued. [8] Referring to [reference], the heavy chain variable region (SEQ ID NO: 15) and light chain variable region (SEQ ID NO: 16) of AMG714 were synthesized to prepare a complete antibody AMG714 of the IgG1m3 subtype for control studies. Similarly, U.S. Patent No. US10301384B2 [9] Referring to the above, the heavy chain variable region (SEQ ID NO: 17) and light chain variable region (SEQ ID NO: 18) of CALY-002 were synthesized to prepare CALY-002, a complete antibody for the IgG1m3 subtype, for control studies.

[0060] 2.2 Affinity analysis of the anti-human IL-15 monoclonal antibody R1A3 The affinity of anti-IL-15 monoclonal antibodies was measured using surface plasmon resonance technology with a Biacore T200. All related reagents and consumables, including the amino coupling kit (BR-1000-50), human antibody capture kit (BR-1008-39), S-series CM5 tip (14100530), and pH 7.4 10×HBS-EP (BR100669), were purchased from GE Healthcare. Following the kit's instructions, the surface of the carboxylated CM5 chip was activated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). Anti-human IgG (Fc) antibody (capture antibody) was diluted to 25 μg / mL with 10 mM sodium acetate (pH 5.0) and then injected at a flow rate of 10 μL / min to achieve a binding amount of approximately 10,000 response units (RUs). After antibody injection and capture, unreacted groups were blocked by injecting 1 M ethanolamine. In kinetic measurements, anti-IL-15 monoclonal antibody was diluted to 1 μg / mL and injected at 10 μL / min, ensuring that approximately 200 RUs of antibody were captured by the anti-human Fc antibody. Subsequently, the IL-15+IL-15Rα complex was injected at a flow rate of 30 μL / min from low to high concentrations, with a series of concentration gradients (e.g., 1.23 nM, 3.7 nM, 11.1 nM, 33.3 nM, and 100 nM). The chip surface was regenerated by injecting 3 M MgCl2 at a flow rate of 10 μL / min for 30 seconds, with a binding time of 90 seconds and a dissociation time of 1200 seconds. Using Biacore T200 evaluation software version 3.2.1, the binding and dissociation sensorgrams were fitted using a 1:1 binding model to determine the binding rate (K a ) and dissociation rate (K d The ratio K was calculated. d / K a The dissociation equilibrium constant (K D The value was calculated. The fitting results are shown in Table 1.

[0061] [Table 1]

[0062] Example 3: Preparation of R1A3 mutant 3.1 Construction of an R1A3 recombinant mutant library Using standard molecular biological techniques, we constructed a CDR3 mutation library based on R1A3VH by introducing mutations into the CDR3 region of the heavy chain variable region R1A3VH. The designed mutation scheme is shown in Table 2. The volume of the constructed library was 2E+7, and the accuracy was 82%.

[0063] [Table 2]

[0064] 3.2 Screening of R1A3 Recombinant Mutation Libraries A recombinant Fab mutant library was constructed by recombining a prepared R1A3 heavy chain mutant library with a fully human light chain antibody library based on a recombinant library phage presentation system (see Example 1 of Chinese Patent Application No. 202210871809.6). Using the recombinant human IL-15+IL-15Rα superagonist prepared in Example 1 as the antigen, the prepared recombinant Fab mutation library was screened using a solid-phase screening strategy (for the experimental scheme, see "Phage Display: A Practical Approach" edited by Clackson, T. and Lowman, H.B., translated by Ma Lan et al., Chemical Industry Press, 2008.5). A total of three rounds of screening were performed by binding, elution, neutralization, infection, and amplification. Anti-human IL15 monoclonal antibodies R4G4VH+R22F11VK (the amino acid sequence of the heavy chain variable region R4G4VH is shown in SEQ ID NO:19, and the amino acid sequence of the light chain variable region R22F11VK is shown in SEQ ID NO:20) and R2H2VH+R22F11VK (the amino acid sequence of the heavy chain variable region R2H2VH is shown in SEQ ID NO:20) were identified. The amino acid sequence of the light chain variable region R22F11VK (shown as NO:21 and SEQ ID NO:20) was ultimately obtained.

[0065] 3.3 Affinity analysis of R1A3 mutants Referring to Example 2.2, affinity analysis was performed using Biacore T200 for the human IL-15 monoclonal antibody R1A3 and its variants, and the results are shown in Table 3.

[0066] [Table 3]

[0067] Example 4: Screening of mouse anti-human IL-15 monoclonal antibodies from a mouse immunology library 4.1 Preparation of hIL-15 mouse immunolibrary Recombinant proteins hIL-15+hIL-15Rα and mfIL-15+mfIL-15Rα superagonists prepared in Example 1 were used as antigens. 6-8 week old BALB / c mice were immunized with an immunodose of 50 μg / mouse, with booster immunizations every 14 days. Eight weeks after the initial immunization, the mice were sacrificed and spleen cells were collected. Mouse spleen lymphocytes were isolated using mouse lymphocyte isolate (Dakewei Biotechnology Co., Ltd., CAT#DKW33-R0100), and total RNA extraction was performed on the isolated lymphocytes using a total RNA extraction kit (Tiangen Biochemistry Technology (Beijing) Co., Ltd., CAT#DP430). Using the extracted total RNA as a template, the antibody heavy chain variable region and light chain variable region were synthesized using a first-strand cDNA synthesis kit (Thermo Scientific, CAT#K1621). Gene-specific reverse transcription primers were employed, and the primer pairing regions were located in the constant region of the antibody heavy chain and the constant region of the antibody light chain, respectively. The specific sequences were PmCGR:TGCATTTGAACTCCTTGCC (SEQ ID NO:24) and PmCKR:CCATCAATCTTCCACTTGAC (SEQ ID No:25). The synthesized cDNA was immediately stored at -70°C for subsequent use. Subsequently, the cDNA obtained by reverse transcription was used as a template, as referenced in (Krebber A, Bornhauser S, Burmester J, et al., Reliable cloning of functional antibody variable domains from hybridomas and spleen cell repertoires employing a reengineered phage display system. J Immunol Methods. 1997;201(1):35-55).

[10] Primers were synthesized with reference to the (all contents of this document are incorporated herein by reference), the nucleotide sequences encoding mouse antibodies VH and VK were amplified by PCR, and then cloned into recombinant plasmids pHGDisn-attP-new and pHKb-attB-new, respectively (see Example 1 of Chinese Patent Application No. 202210871809.6 for a method of preparing recombinant plasmids).

[0068] A recombinant mouse phage library was prepared by recombining the prepared PHGDisn-attP-new-mVHs heavy chain library (library volume: 6.1E+7, accuracy: 63%) and the PHKb-attB-new-mVKs light chain library (library volume: 7.4E+7, accuracy: 78%) (see Example 1 of Chinese Patent Application No. 202210871809.6). The constructed library volume reached 2.7E+11, and the sequencing accuracy was 60%.

[0069] 4.2 Screening of hIL-15 mouse immunolibraries Using recombinant human IL-15+IL-15Rα and monkey IL-15+IL-15Rα superagonists prepared in Example 1 as antigens, a solid-phase screening strategy (for the experimental scheme, see "Phage Display: A Practical Approach" edited by Clackson, T. and Lowman, H.B., translated by Ma Lan et al., Chemical Industry Press, 2008.5) was used to screen the prepared mouse recombinant phage library. Three rounds of screening were performed by binding, elution, neutralization, infection, and amplification, ultimately yielding one strain of mouse monoclonal antibody R6E11 (the amino acid sequence of the heavy chain variable region R6E11VH is shown in SEQ ID NO:43, and the amino acid sequence of the light chain variable region R6E11VK is shown in SEQ ID NO:44) that specifically binds to human IL-15.

[0070] 4.3 Affinity Analysis of Mouse Anti-Human IL-15 Monoclonal Antibody R6E11 Referring to Example 2.2, affinity analysis was performed for the anti-IL-15 monoclonal antibody R6E11 using Biacore T200, and the results are shown in Table 4.

[0071] [Table 4]

[0072] Example 5: Humanization of mouse monoclonal antibody R6E11 5.1 Humanization of R6E11 The mouse monoclonal antibody R6E11 was humanized to reduce its immunogenicity. A classical framework transplantation strategy was employed in the humanization scheme (J Immunol, 2002 Jul 15;169(2):1119-25).

[11] The heavy chain variable region and light chain variable region of R6E11 were compared with human antibody germ cell gene sequences in the IMGT database, respectively. Appropriate germ cell gene sequences were selected to provide the antibody framework regions 1-3 (FR1+FR2+FR3), and an appropriate J region gene sequence was selected to provide framework region 4 (FR4). This template can be selected based on various factors, such as the relative length of the antibody, the size of the CDR, the amino acid residues located at the junction between the antibody framework region (FR) and hypervariable region (CDR), and the homology of the entire sequence. The selected template may be a mixture of multiple sequences or a common template in order to maintain the appropriate conformation of the complementarity-determining region (CDR) as much as possible. Finally, one humanized variant heavy chain variable region variant R6E11VH-h3 and one humanized light chain variable region variant R6E11VK-h3 were obtained.

[0073] 5.2 Screening of humanized R6E11 mutants Considering that the heavy chain sequences of the humanized mutants R6E11VH-h3+R6E11VK-h3 contain deamination sites NG and NA, mutations were designed to improve the physicochemical properties of R6E11VH-h3. The mutation scheme is shown in Table 5. The constructed library volume was 2E+7, and the accuracy was 47%.

[0074] [Table 5]

[0075] A recombinant Fab mutant library was constructed by recombining a prepared R6E11VH-h3 mutant library with a full human light chain antibody library based on a recombinant library phage presentation system (see Example 1 of Chinese Patent Application No. 202210871809.6). The constructed recombinant Fab mutant library underwent a total of three rounds of screening and enrichment using a solid-phase screening method with a human IL-15 + IL-15Rα superagonist antigen. Ultimately, a monoclonal antibody R26H10 containing a full human light chain with improved physicochemical properties was obtained (the amino acid sequence of the heavy chain variable region R26H10VH is shown in SEQ ID NO:22, and the amino acid sequence of the light chain variable region R26H10VK is shown in SEQ ID NO:23).

[0076] 5.3 Affinity analysis of the R6E11 humanized mutant Referring to Example 2.2, affinity analysis was performed on the anti-human IL-15 mouse monoclonal antibody R6E11 and its humanized variant using Biacore T200, and the results are shown in Table 6.

[0077] [Table 6]

[0078] Example 6: Binding and affinity analysis of anti-human IL-15 monoclonal antibodies against different species of IL-15 Prepared human IL-15 (hIL-15+hIL-15Rα-His complex), cynomolgus monkey IL-15 (mfIL-15+mfIL-15Rα-His complex), and mouse IL-15 (mIL-15+mIL-15Rα-mFc1 complex) were coated onto 96-well ELISA plates at 1 μg / mL and 100 μL / well, respectively, and coated overnight at 4°C. After blocking at 37°C for 1 hour using blocking solution (3% skim milk-PBST), each anti-IL-15 monoclonal antibody was added and conjugated at 37°C for 1 hour. The ELISA plates were washed with PBST, and HRP mouse anti-human IgG (Beijing Boaosen Biotechnology Co., Ltd., bsm-0297M-HRP) was added and conjugated at 37°C for 1 hour. ELISA plates were washed with PBST, OPD substrate chromogenic solution was added, and after 5-10 minutes, color development was terminated with 1M H2SO4. Optical density values ​​were measured at dual wavelengths of 492nm / 630nm using a microplate reader. The results of the ELISA analysis (Figure 1) showed that the anti-human IL-15 antibodies R1A3, R2H2VH+R22F11VK, R4G4VH+R22F11VK, and R26H10 cross-recognized human IL-15 and cynomolgus monkey IL-15, while none of the molecules recognized mouse IL-15.

[0079] Referring to Example 2.2, the affinity of anti-human IL-15 monoclonal antibodies that bind to human IL-15 and cynomolgus monkey IL-15 was analyzed using Biacore T200, and the results are shown in Tables 7 and 8.

[0080] [Table 7]

[0081] [Table 8]

[0082] Example 7: Epitope analysis of anti-human IL-15 monoclonal antibody Human IL-15 (hIL-15 + hIL-15Rα-his complex) was coated onto 96-well ELISA plates at 1 μg / mL, 100 μL / well, and coated overnight at 4°C. Blocking was performed using a blocking solution (3% skim milk-PBST) at 37°C for 1 hour. Fixed-concentration purified anti-human IL-15 phage (AMG714 was 4 × 10⁶) was then used. 11 cfu / mL, CALY-002 is 1 × 10 10 Anti-human IL-15 monoclonal antibodies (R1A3, R2H2VH+R22F11VK, R4G4VH+R22F11VK, R26H10, AMG714, and CALY-002) were serially diluted using cfu / mL, starting at a concentration of 30 μg / mL, followed by 3-fold serial dilutions, totaling 11 concentration gradients. These were added to well-blocked 96-well ELISA plates at 100 μL / well and incubated at 37°C for 1 hour. The ELISA plates were washed with PBST, and then HRP-labeled anti-M13 secondary antibody (Beijing Yiqiao Shenzhou Technology Co., Ltd., 11973-MM05T-H) was added and incubated at 37°C for 1 hour. The ELISA plate was washed with PBST, OPD substrate chromogenic solution was added, and after 5-10 minutes, color development was terminated with 1M H2SO4. The optical density was measured at a dual wavelength of 492nm / 630nm using a microplate reader. The results of the ELISA analysis are shown in Figure 2. The epitope of R26H10 that binds to human IL-15 is different from that of AMG714 (Figure 2A) and similar to or cross-relating with CALY-002 (Figure 2B). The epitopes of R1A3, R2H2VH+R22F11VK, and R4G4VH+R22F11VK that bind to human IL-15 are similar to or cross-relating with AMG714 (Figure 2A) and different from CALY-002 (Figure 2B). The epitope of AMG714 that binds to human IL-15 is different from that of CALY-002 (Figure 2B).

[0083] Example 8: Activity evaluation of anti-human IL-15 monoclonal antibody against HEK-Blue IL-2 cells HEK-Blue IL-2 cells are reliably transfected with human CD25 (IL-2Rα), CD122 (IL-2Rβ), and CD132 (IL-2Rγ) genes into HEK293-derived cells, enabling IL-2 signaling via the JAK-STAT5 pathway. Simultaneously, HEK-Blue cells are also introduced with a STAT5-inducible secreted embryonic alkaline phosphatase (SEAP) reporter gene. Under stimulation with IL-2 or IL-15, HEK-Blue IL-2 cells can activate JAK / STAT5 and secrete SEAP. SEAP detection kit QUANTI-Blue TM The solution can be used to monitor the amount of SEAP, enabling activity monitoring of IL-2 or IL-15.

[0084] The HEK-Blue IL-2 cell line was purchased from Invivogen. DMEM + 5% inactivated FBS was prepared as the test medium, and HEK-Blue IL-2 cells were incubated in the test medium at a rate of 5 × 10⁶. 5 The antibody was resuspended at 1 / mL. Using 5 ng / mL human IL-15 (hIL-15+hIL-15Rα complex) as a diluent, an antibody concentration gradient was created starting at 400 nM, with 10 concentration points produced by 4-fold dilutions. 100 μL of antibody and 100 μL of cells were mixed and cultured in a carbon dioxide incubator (37°C, 5% CO2) for 20 hours. Centrifugation was performed, and 20 μL of the supernatant was taken and mixed with 180 μL of Quanti-blue (Invivogen, product number: rep-qbs). The mixture was incubated at 37°C for 1-2 hours, and absorbance was detected at 630 nm using a microplate reader (Biotek, model: 800TS). Raw data were statistically processed using Graphpad Prism 7.0 software. DP47 antibody (negative control, US Patent Application US20160200833A1)

[12] Prepared with reference to the above, where the amino acid sequences of the heavy chain variable region and light chain variable region are shown in SEQ ID NO: 50 and 51, respectively). The results are shown in Figure 3, and the activity of the four molecules R1A3, R4G4VH+R22F11VK, R2H2VH+R22F11VK, and R26H10 was higher than that of CALY-002 and AMG-714, and specific IC 50 The data is shown in Table 9.

[0085] [Table 9]

[0086] Example 9: Activity evaluation of anti-human IL-15 monoclonal antibody against NK92 cells The NK92 cell line was purchased from Nanjing Kebai Biotechnology Co., Ltd. It is a cell line that depends on IL-2 proliferation, and IL-15 can promote cell proliferation when IL-2 is deprived. One day prior to the start of the experiment, NK92 cells were starved (by performing a blank culture without IL-2), and on the second day, the cells were resuspended at 5 × 10⁵ cells / mL in IL-2-free medium. The antibody was prepared using 12 ng / mL human IL-15 (hIL-15 + hIL-15Rα-his complex) as the diluent, starting the antibody concentration gradient at 400 nM, and 10 concentration points were created by 4-fold dilution. 100 μL of antibody and 100 μL of cells were mixed and cultured in a carbon dioxide incubator (37°C, 5% CO₂) for 2 days. Cell proliferation was detected using the Cell Titer-Glo® Luminescent Cell Viability Assay (Promega, product number G7571), and full-wavelength fluorescence detection was performed using a microplate reader (Molecular Devices, model SpectraMax I3X). Raw data were statistically processed using Graphpad Prism 7.0 software. DP47 antibody served as a negative control.

[0087] See Figure 4 for the results. The activity of the four anti-human IL-15 monoclonal antibodies, R1A3, R26H10, R4G4VH+R22F11VK, and R2H2VH+R22F11VK, was higher than that of AMG-714 and similar to that of CALY-002. Of these, CALY-002 was unable to completely inhibit the IL-15 activity of NK92 cells. Specific IC 50 The data is shown in Table 10.

[0088] [Table 10]

[0089] [Sequence Information] SEQ ID NO:1 NWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISLESGDASIHDTVENLIILANNSLSSNGNVTESGCKECEELEEKNIKEFLQSFVHIVQMFINTS SEQ ID NO:2 ITCPPPMSVEHADIWVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNVAHWTTPSLKCIRDPALVHQRPAPPSTVTTAGVTPQPESLSPSGKE SEQ ID NO:3 NWVNVISDLKKIEDLIQSMHIDATLYTESDVHPSCKVTAMKCFLLELQVISHESGDTDIHDTVENLIILANNILSSNGNITESGCKECEELEEKNIKEFLQSFVHIVQMFINTS SEQ ID NO:4 ITCPPPVSVEHADIRVKSYSLYSRERYICNSGFKRKAGTSSLTECVLNKATNIAHWTTPSLKCIRDPLLARQRPAPPFTVTTAGVTPQPESLSPSGKE SEQ ID NO:5 NWIDVRYDLEKIESLIQSIHIDTTLYTDSDFHPSCKVTAMNCFLLELQVILHEYSNMTLNETVRNVLYLANSTLSSNKNVAESGCKECEELEEKTFTEFLQSFIRIVQMFINTS SEQ ID NO:6 GTTCPPPVSIEHADIRVKNYSVNSRERYVCNSGFKRKAGTSTLIECVINKNTNVAHWTTPSLKCIRDPSLAHYSPVPTVVTPKVTSQPESPSPSAKE SEQ ID NO:7 HHHHHH SEQ ID NO:8 ASVPRDSGCKPCICTVPEVSSVFIFPPKPKDVLTITLTPKVTCVVVDISKDDPEVQFSWFVDDVEVHTAQTQPREEQFNSTFRSVSELPIMHQDWLNGKEFKCRVNSAAFPAPIEKTISKTKGRPKAPQVYTIPPPKEQMAKDKVSLTCMITDFFPEDITVEWQWNGQPAENYKNTQPIMDTDGSYFVYSKLNVQKSNWEAGNTFTCSVLHEGLHNHHTEKSLSHSPGK SEQ ID NO:9 ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO:10 ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEFEGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPASIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID NO:11 RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC SEQ ID NO:12 GQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS SEQ ID NO:13 EVQLVQSGAEVKKPGESLKISCKVSGYFFTTYWIGWVRQMPGKGLEYMGIIYPGDSDTRYSPSFQGQVTISADKSISTAYLQWSSLKASDTAMYYCARGGHWNAFDFWGQGTLVTVSS SEQ ID NO:14 EIVLTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASRRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQFDASQTFGQGTKLEIS SEQ ID NO:15 EVQLVQSGAEVKKPGESLKISCKVSGYFFTTYWIGWVRQMPGKGLEYMGIIYPGDSDTRYSPSFQGQVTISADKSISTAYLQWSSLKASDTAMYYCARGGNWNCFDYWGQGTLVTVSS SEQ ID NO:16 EIVLTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASRRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQRYGSSHTFGQGTKLEIS SEQ ID NO:17 EVQLVESGGGLVQPGGSLRLSCAASEFTFSNYAMSWVRQAPGKGLEWVATISRGGDYTYYPDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARRVSMITGGWAMDYWGQGTLVTVSS SEQ ID NO:18 DVVMTQSPLSLPVTLGQPASISCRSSQSIVDITGNTYLEWYQQRPGQSPRLLIYKVFNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQDSFVPYTFGQGTKLEIK SEQ ID NO:19 EVQLVQSGAEVKKPGESLKISCKVSGYFFTTYWIGWVRQMPGKGLEYMGIIYPGDSDTRYSPSFQGQVTISADKSISTAYLQWSSLKASDTAMYYCARGGHWNAFDIWGQGTLVTVSS SEQ ID NO:20 EIVLTQSPGTLSLSPGERATLSCRASQSVIGSYLAWYQQKPGQAPRLLIYSASKLASGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQHYSTPGTFGQGTKVEIK SEQ ID NO:21 EVQLVQSGAEVKKPGESLKIISCKVSGYFFTTYWIGWVRQMPGKGLEYMGIIYPGDSDTRYSPSFQGQVTISADKSISTAYLQWSSLKASDTAMYYCARGGHWNSFDTWGQGTLVTVSS SEQ ID NO:22 QVTLKESGPVLVKPTETLTLTTCTVSGFSLTTYAVHWIRQPPGKALEWLGVIWGAGSTDYNPSFMSRLTISKDTSKSQVVLTMTNMDPVDTATYYCAKNAAYYVMDYWGQGTTVTVSS SEQ ID NO:23 EIVLTQSPGTLSLSPGERATLLSCRASQSVIYSYLAWYQQKPGQAPRLLIYAASKLASGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQRASYPLTFGQGTKVEIK SEQ ID NO:24 TGCATTTGAACTCCTTGCC SEQ ID NO:25 CCATCAATCTTCCACTTGAC SEQ ID NO:26 TYWIG SEQ ID NO:27 IIYPGDSDTRYSPSFQG SEQ ID NO:28 GGHWNAFDF SEQ ID NO:29 GGHWNAFDI SEQ ID NO:30 GGHWNSFDT SEQ ID NO:31 TYAVH SEQ ID NO:32 VIWGAGSTDYNPSFMS SEQ ID NO:33 NAAYYVMDY SEQ ID NO:34 RASQSVSSSYLA SEQ ID NO:35 GASRRAT SEQ ID NO:36 QQFDASQT SEQ ID NO:37 RASQSVIGSYLA SEQ ID NO:38 SASKLAS SEQ ID NO:39 QQHYSTPGT SEQ ID NO:40 RASQSVIYSYLA SEQ ID NO:41 AASKLAS SEQ ID NO:42 QQRASYPLT SEQ ID NO:43 EVKLKESGPSLVQPSQSLSITCTVSGFSLTTYAVHWVRQSPGKGLEWLGVIWGAGSTDYNAAFMSRLSITKDNSKSQVFFKMNSLQADDTAIYFCAKNGAYYVMDYWGQGTSVTVSS SEQ ID NO:44 EIVLTQSPAIMSASPGEKVTITCSASSSVSYMHWFQQRPGTSPKLWIYSTSNLASGVPARFSGSGSGTSYSLTISRMEAEDAATYYCQQRSSYPLTFGAGTKLEIK SEQ ID NO:45 VIWGAGSTDYNAAFMS SEQ ID NO:46 NGAYYVMDY SEQ ID NO:47 SASSSVSYMH SEQ ID NO:48 STSNLAS SEQ ID NO:49 QQRSSYPLT SEQ ID NO:50 EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSAISGSGGSTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKGSGFDYWGQGTLVTVSS SEQ ID NO:51 EIVLTQSPGTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLEPEDFAVYYCQQYGSSPLTFGQGTKVEIK

[0090] References 1.Steel JC, Waldmann TA, Morris JC. Interleukin-15 biology and its therapeutic implications in cancer. Trends Pharmacol Sci. 2012 Jan;33(1):35-41. 2.Fehniger TA, Caligiuri MA. Interleukin 15: biology and relevance to human disease. Blood. 2001 Jan 1;97(1):14-32. 3.Bergqvist C, Ezzedine K. Vitiligo: A focus on pathogenesis and its therapeutic implications. J Dermatol. 2021 Mar;48(3):252-270. 4.Richmond JM, Strassner JP, Rashighi M, et al. Resident Memory and Recirculating Memory T Cells Cooperate to Maintain Disease in a Mouse Model of Vitiligo. J Invest Dermatol. 2019 Apr;139(4):769-778. 5.Richmond JM, Strassner JP, Zapata L Jr, et al.. Antibody blockade of IL-15 signaling has the potential to durably reverse vitiligo. Sci Transl Med. 2018 Jul 18;10(450). 6. Phage Display: A General Experimental Guide / Edited by Clackson, T. and Lowman, HB. (USA), translated by Ma Lan et al. Chemical Industry Press, May 2008. 7.CN202210871809.6. 8.US7153507B2. 9.US10301384B2. 10.Krebber A, Bornhauser S, Burmester J, et al. Reliable cloning of functional antibody variable domains from hybridomas and spleen cell repertoires employing a reengineered phage display system. J Immunol Methods. 1997;201(1):35-55. 11.J Immunol,2002 Jul 15;169(2):1119-25.

Claims

1. An antibody that binds to human IL-15, It comprises a heavy chain variable region containing the amino acid sequences of HCDR1, HCDR2, and HCDR3, and a light chain variable region containing the amino acid sequences of LCDR1, LCDR2, and LCDR3, The amino acid sequence of HCDR1 is shown in SEQ ID NO: 26, the amino acid sequence of HCDR2 is shown in SEQ ID NO: 27, the amino acid sequence of HCDR3 is shown in SEQ ID NO: 29, the amino acid sequence of LCDR1 is shown in SEQ ID NO: 37, the amino acid sequence of LCDR2 is shown in SEQ ID NO: 38, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:

39. The amino acid sequence of HCDR1 is shown in SEQ ID NO: 26, the amino acid sequence of HCDR2 is shown in SEQ ID NO: 27, the amino acid sequence of HCDR3 is shown in SEQ ID NO: 28, the amino acid sequence of LCDR1 is shown in SEQ ID NO: 34, the amino acid sequence of LCDR2 is shown in SEQ ID NO: 35, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:

36. The amino acid sequence of HCDR1 is shown in SEQ ID NO: 26, the amino acid sequence of HCDR2 is shown in SEQ ID NO: 27, the amino acid sequence of HCDR3 is shown in SEQ ID NO: 30, the amino acid sequence of LCDR1 is shown in SEQ ID NO: 37, the amino acid sequence of LCDR2 is shown in SEQ ID NO: 38, and the amino acid sequence of LCDR3 is shown in SEQ ID NO:

39. The amino acid sequence of HCDR1 is shown in SEQ ID NO: 31, the amino acid sequence of HCDR2 is shown in SEQ ID NO: 32, the amino acid sequence of HCDR3 is shown in SEQ ID NO: 33, the amino acid sequence of LCDR1 is shown in SEQ ID NO: 40, the amino acid sequence of LCDR2 is shown in SEQ ID NO: 41, the amino acid sequence of LCDR3 is shown in SEQ ID NO: 42, or, The amino acid sequence of HCDR1 is shown as SEQ ID NO: 31, the amino acid sequence of HCDR2 is shown as SEQ ID NO: 45, the amino acid sequence of HCDR3 is shown as SEQ ID NO: 46, the amino acid sequence of LCDR1 is shown as SEQ ID NO: 47, the amino acid sequence of LCDR2 is shown as SEQ ID NO: 48, and the amino acid sequence of LCDR3 is shown as SEQ ID NO:

49. Here, the amino acid sequences of HCDR and LCDR are defined by Kabat. antibody.

2. The antibody according to claim 1, wherein the amino acid sequence of the heavy chain variable region of the antibody is shown in SEQ ID NO: 13, 19, 21, 22, or 43.

3. The antibody according to claim 1, wherein the amino acid sequence of the light chain variable region of the antibody is shown in SEQ ID NO: 14, 20, 23, or 44.

4. The amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO: 13, and the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:

14. The amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO: 19, and the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:

20. The amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO: 21, and the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:

20. The amino acid sequence of the heavy chain variable region of the antibody is shown in SEQ ID NO: 22, and the amino acid sequence of the light chain variable region of the antibody is shown in SEQ ID NO: 23, or, The amino acid sequence of the heavy chain variable region of the antibody is shown as SEQ ID NO: 43, and the amino acid sequence of the light chain variable region of the antibody is shown as SEQ ID NO:

44. The antibody according to any one of claims 1 to 3.

5. An antibody that binds to human IL-15, wherein the amino acid sequence of the heavy chain variable region of the antibody has at least 90% identity with one of SEQ ID NO: 13, 19, 21, 22, and 43, and the amino acid sequence of the light chain variable region of the antibody has at least 90% identity with one of SEQ ID NO: 14, 20, 23, and 44.

6. The aforementioned antibody is a complete antibody, Fab fragment, F(ab') ２ A fragment, or a single-stranded Fv fragment (scFv), preferably the antibody is a fully human antibody or a humanized antibody, and / or The antibody is a monoclonal antibody, and / or, The antibody further comprises a heavy chain constant region selected from IgG1 subtype, IgG2 subtype or IgG4 subtype, preferably the heavy chain constant region being IgG1 subtype, more preferably the heavy chain constant region being IgG1m3 subtype, and / or The 234th, 235th, and 331st amino acids of the Fc fragment are F, E, and S, respectively, and / or The 252nd, 254th, and 256th amino acids in the heavy chain constant region are Y, T, and E, respectively, and / or The antibody further comprises a light chain constant region selected from a κ subtype or a λ subtype, preferably the light chain constant region being a κ subtype. Here, the amino acid positions in the constant region of the antibody are determined according to EU numbering. The antibody according to any one of claims 1 to 5.

7. The antibody binds to human IL-15 and / or monkey IL-15, and / or The antibody can inhibit the activity of IL-15, for example, by inhibiting IL-15's ability to induce the secretion of secretory embryonic alkaline phosphatase (SEAP), and / or by inhibiting IL-15's ability to induce the proliferation of NK92 cells. The antibody according to any one of claims 1 to 6.

8. A nucleic acid molecule encoding an antibody according to any one of claims 1 to 7.

9. A pharmaceutical composition comprising an antibody according to any one of claims 1 to 7 and a pharmaceutically acceptable excipient, diluent, or carrier.

10. The pharmaceutical composition according to claim 9, which is used for the prevention or treatment of an IL-15-mediated disease, preferably the IL-15-mediated disease being selected from vitiligo, celiac disease, eosinophilic esophagitis, and macrogranular lymphocytic leukemia.

11. Use of the antibody according to any one of claims 1 to 7, or the pharmaceutical composition according to claim 9 or 10, in the manufacture of a pharmaceutical for the prevention or treatment of an IL-15-mediated disease. Preferably, the IL-15-mediated disease is selected from vitiligo, celiac disease, eosinophilic esophagitis, and large granular lymphocytic leukemia.

12. A method for preventing or treating IL-15-mediated diseases, A method comprising administering an antibody according to any one of claims 1 to 7, or a pharmaceutical composition according to claim 9 or 10, to an individual in need thereof. Preferably, the IL-15-mediated disease is selected from vitiligo, celiac disease, eosinophilic esophagitis, and macrogranular lymphocytic leukemia.

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