TL1a antibody and use thereof

Abstract

A TL1A antibody and use thereof. Provided are an antibody or antigen-binding fragment that specifically binds to human TL1A, a multi-characteristic antigen-binding molecule, a chimeric antigen receptor, an immune effector cell, a nucleic acid molecule, a vector, a cell, a preparation method, a pharmaceutical composition, a pharmaceutical use, and a disease treatment method. The present invention is of great significance for developing a drug for treating TL1A immune-mediated inflammatory diseases (IMIDs) (for example, autoimmune diseases and inflammatory diseases).

Description

TL1A antibodies and uses thereof

[0001] Cross-reference to Related Applications

[0002] This disclosure claims priority to and the benefit of Chinese Patent Application No. 202411264964.7, filed September 10, 2024, the patent application file of which is incorporated herein by reference in its entirety. TECHNICAL FIELD

[0003] The present disclosure relates to the field of medicine, in particular, to antibodies that specifically bind to human TNF-like ligand 1A (TL1A) and block its activity, and their use in treating autoimmune or inflammatory diseases. BACKGROUND

[0004] Inflammatory bowel disease (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), is an autoimmune disease. Common symptoms are diarrhea, bloody stool, and long-term inflammation can affect the whole body organs. IBD, as a chronic non-specific intestinal inflammatory disease, is recurrent, and there is no complete cure method at present, and the global disease burden is heavy, and the incidence and prevalence continue to increase. Traditional treatment drugs for IBD include glucocorticoids, antibiotics, immunosuppressants, aminosalicylic acid, etc. After infliximab and adalimumab targeting TNF-a were approved for UC / CD indications, IBD has entered the “biological agent” era. However, there is still a considerable proportion of target patient groups who do not respond or will lose response over time to existing biological agents, and there is still a great unmet medical need in the clinic.

[0005] Tumor necrosis factor-like cytokine 1A (TL1A) is a member of the tumor necrosis factor family. It is expressed in different immune cells, such as monocytes, macrophages, dendritic cells, T cells, and non-immune cells, such as synovial fibroblasts, endothelial cells, etc. The basal level of TL1A is low, but it will rapidly increase after immune activation. Studies have shown that the expression level of TL1A in the colon tissue of IBD patients is related to the severity of inflammation.

[0006] Given that TL1A plays an important role in biological processes related to several important human diseases, such as IBD, and the clinical medical needs of IBD are still strong, antibodies that more strongly inhibit the activity of TL1A have high potential clinical value. SUMMARY

[0007] The present disclosure provides antibodies or antigen-binding fragments that specifically bind to TL1A, corresponding multispecific antigen-binding molecules, chimeric antigen receptors, immune effector cells, nucleic acid fragments, vectors, host cells, pharmaceutical compositions, methods of making the same, pharmaceutical uses and applications in the treatment of immune-mediated inflammatory diseases (IMIDs), e.g., autoimmune and inflammatory diseases.

[0008] In a first aspect, the present disclosure provides an antibody or antigen-binding fragment that specifically binds to TL1A, comprising:

[0009] (a) a heavy chain variable region VH comprising a HCDR1, a HCDR2 and a HCDR3; and / or,

[0010] (b) a light chain variable region VL comprising a LCDR1, a LCDR2 and a LCDR3.

[0011] In some embodiments, the HCDR1, HCDR2 and HCDR3 have the amino acid sequence of the HCDR1, HCDR2 and HCDR3 of a VH as set forth in any one of SEQ ID NOs: 12, 14, 16, 18, 20, 22, 120, 121, 122, 123, 133, 134, 135, 136, 144, 145, 146, 147, 148, 149, 158, 159, 160, 161 and 162.

[0012] In some embodiments, the LCDR1, LCDR2 and LCDR3 have the amino acid sequence of the LCDR1, LCDR2 and LCDR3 of a VL as set forth in any one of SEQ ID NOs: 13, 15, 17, 19, 21, 23, 117, 118, 119, 129, 130, 131, 132, 141, 142, 143, 155, 156 and 157.

[0013] Preferably, the HCDR1-HCDR3 and / or the LCDR1-LCDR3 are selected from Table 5, Table 6, Table 7, Table 11, Table 13, Table 15 or Table 17.

[0014] In some embodiments, the antibody or antigen-binding fragment comprises the HCDR1-HCDR3 of a VH as set forth in SEQ ID NO: 12, 120, 121, 122 or 123, and the LCDR1-LCDR3 of a VL as set forth in SEQ ID NO: 13, 117, 118 or 119.

[0015] In some embodiments, the antibody or antigen-binding fragment comprises the amino acid sequences of HCDR1-HCDR3 of a VH set forth in SEQ ID NO: 14, and LCDR1-LCDR3 of a VL set forth in SEQ ID NO: 15.

[0016] In some embodiments, the antibody or antigen-binding fragment comprises the amino acid sequences of HCDR1-HCDR3 of a VH set forth in SEQ ID NO: 16, and LCDR1-LCDR3 of a VL set forth in SEQ ID NO: 17.

[0017] In some embodiments, the antibody or antigen-binding fragment comprises the amino acid sequences of HCDR1-HCDR3 of a VH set forth in SEQ ID NO: 18, 133, 134, 135, or 136, and LCDR1-LCDR3 of a VL set forth in SEQ ID NO: 19, 129, 130, 131, or 132.

[0018] In some embodiments, the antibody or antigen-binding fragment comprises the amino acid sequences of HCDR1-HCDR3 of a VH set forth in SEQ ID NO: 20, 144, 145, 146, 147, 148, or 149, and LCDR1-LCDR3 of a VL set forth in SEQ ID NO: 21, 141, 142, or 143.

[0019] In some embodiments, the antibody or antigen-binding fragment comprises the amino acid sequences of HCDR1-HCDR3 of a VH set forth in SEQ ID NO: 22, 158, 159, 160, 161, or 162, and LCDR1-LCDR3 of a VL set forth in SEQ ID NO: 23, 155, 156, or 157.

[0020] Preferably, the HCDR1-HCDR3 and / or the LCDR1-LCDR3 are determined according to the Kabat, IMGT, or Chothia system.

[0021] In some specific embodiments, the HCDR1-HCDR3 have the amino acid sequences selected from the group consisting of SEQ ID NOs: 39-41; SEQ ID NOs: 75-77; or SEQ ID NOs: 105-106 and 41, respectively.

[0022] In some specific embodiments, the HCDR1-HCDR3 have the amino acid sequences selected from the group consisting of SEQ ID NOs: 45-47; SEQ ID NOs: 80-82; or SEQ ID NOs: 107-108 and 47, respectively.

[0023] In some particular embodiments, the HCDR1-HCDR3 have amino acid sequences selected from the group consisting of: SEQ ID NOs: 51-53; SEQ ID NOs: 85-87; or SEQ ID NOs: 109-110 and 53, respectively.

[0024] In some particular embodiments, the HCDR1-HCDR3 have amino acid sequences selected from the group consisting of: SEQ ID NOs: 57-59; SEQ ID NOs: 90-92; or SEQ ID NOs: 111-112 and 59, respectively.

[0025] In some particular embodiments, the HCDR1-HCDR3 have amino acid sequences selected from the group consisting of: SEQ ID NOs: 63-65; SEQ ID NOs: 95-97; or SEQ ID NOs: 113-114 and 65, respectively.

[0026] In some particular embodiments, the HCDR1-HCDR3 have amino acid sequences selected from the group consisting of: SEQ ID NOs: 69-71; SEQ ID NOs: 100-102; SEQ ID NOs: 115-116 and 71; or SEQ ID NOs: 69, 154 and 71, respectively.

[0027] In some particular embodiments, the LCDR1-LCDR3 have sequences selected from the group consisting of: SEQ ID NOs: 42-44; or SEQ ID NOs: 78-79 and 44.

[0028] In some particular embodiments, the LCDR1-LCDR3 have sequences selected from the group consisting of: SEQ ID NOs: 48-50; or SEQ ID NOs: 83-84 and 50.

[0029] In some particular embodiments, the LCDR1-LCDR3 have sequences selected from the group consisting of: SEQ ID NOs: 54-56; or SEQ ID NOs: 88-89 and 56.

[0030] In some particular embodiments, the LCDR1-LCDR3 have sequences selected from the group consisting of: SEQ ID NOs: 60-62; or SEQ ID NOs: 93-94 and 62.

[0031] In some specific embodiments, the LCDR1-LCDR3 have a sequence selected from the group consisting of: SEQ ID NO: 66-68; SEQ ID NO: 98-99 and 68; or SEQ ID NO: 140 and 67-68; or

[0032] In some specific embodiments, the LCDR1-LCDR3 have a sequence selected from the group consisting of: SEQ ID NO: 72-74; or SEQ ID NO: 103-104 and 74.

[0033] In preferred embodiments, the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NO: 39-41, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NO: 42-44, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NO: 75-77, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NO: 78-79 and 44, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NO: 105-106 and 41, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NO: 42-44, respectively;

[0034] In preferred embodiments, the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NO: 45-47, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NO: 48-50, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NO: 80-82, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NO: 83-84 and 50, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NO: 107-108 and 47, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NO: 48-50, respectively;

[0035] In preferred embodiments, the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 51-53, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 54-56, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 85-87, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 88-89 and 56, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 109-110 and 53, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 54-56, respectively;

[0036] In preferred embodiments, the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 57-59, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 60-62, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 90-92, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 93-94 and 62, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 111-112 and 59, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 60-62, respectively;

[0037] In preferred embodiments, the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 63-65, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 66-68, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 95-97, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 98-99 and 68, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 113-114 and 65, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 66-68, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 63-65, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 140 and 67-68, respectively; or

[0038] In preferred embodiments, the HCDR1-HCDR3 have the amino acid sequences set forth in SEQ ID NOs: 69-71, respectively, and the LCDR1-LCDR3 have the amino acid sequences set forth in SEQ ID NOs: 72-74, respectively; the HCDR1-HCDR3 have the amino acid sequences set forth in SEQ ID NOs: 100-102, respectively, and the LCDR1-LCDR3 have the amino acid sequences set forth in SEQ ID NOs: 103-104 and 74, respectively; the HCDR1-HCDR3 have the amino acid sequences set forth in SEQ ID NOs: 115-116 and 71, respectively, and the LCDR1-LCDR3 have the amino acid sequences set forth in SEQ ID NOs: 72-74, respectively; or the HCDR1-HCDR3 have the amino acid sequences set forth in SEQ ID NOs: 69, 154 and 71, respectively, and the LCDR1-LCDR3 have the amino acid sequences set forth in SEQ ID NOs: 72-74, respectively.

[0039] Preferably, the HCDR1-3 and / or the LCDR1-3 are determined according to the Kabat, Chothia or IMGT approach.

[0040] In some specific embodiments, the heavy chain variable region VHcomprises an amino acid sequence as set forth in any one of SEQ ID NOs: 12, 14, 16, 18, 20, 22, 120, 121, 122, 123, 133, 134, 135, 136, 144, 145, 146, 147, 148, 149, 158, 159, 160, 161 and 162, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations thereto; and / or,

[0041] the light chain variable region VLcomprises an amino acid sequence as set forth in any one of SEQ ID NOs: 13, 15, 17, 19, 21, 23, 117, 118, 119, 129, 130, 131, 132, 141, 142, 143, 155, 156 and 157, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations thereto.

[0042] In preferred embodiments, the heavy chain variable region VHcomprises an amino acid sequence as set forth in SEQ ID NO: 12, and the light chain variable region VLcomprises an amino acid sequence as set forth in SEQ ID NO: 13.

[0043] In preferred embodiments, the heavy chain variable region VHcomprises an amino acid sequence as set forth in SEQ ID NO: 14, and the light chain variable region VLcomprises an amino acid sequence as set forth in SEQ ID NO: 15.

[0044] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in SEQ ID NO: 16, and the light chain variable region VLcomprises the amino acid sequence set forth in SEQ ID NO: 17.

[0045] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in SEQ ID NO: 18, and the light chain variable region VLcomprises the amino acid sequence set forth in SEQ ID NO: 19.

[0046] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in SEQ ID NO: 20, and the light chain variable region VLcomprises the amino acid sequence set forth in SEQ ID NO: 21.

[0047] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in SEQ ID NO: 22, and the light chain variable region VLcomprises the amino acid sequence set forth in SEQ ID NO: 23.

[0048] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 120-123, and the light chain variable region VLcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 117-119.

[0049] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 133-136, and the light chain variable region VLcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 129-132.

[0050] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 144-149, and the light chain variable region VLcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 141-143.

[0051] In preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 158-162, and the light chain variable region VLcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 155-157.

[0052] In more preferred embodiments, the heavy chain variable region VHcomprises the amino acid sequence set forth in any one of SEQ ID NOs: 120-123, and the light chain variable region VLcomprises the amino acid sequence set forth in SEQ ID NO: 119.

[0053] In more preferred embodiments, the heavy chain variable region VHcomprises an amino acid sequence as set forth in any one of SEQ ID NOs: 133-136, and the light chain variable region VLcomprises an amino acid sequence as set forth in SEQ ID NO: 129.

[0054] In more preferred embodiments, the heavy chain variable region VHcomprises an amino acid sequence as set forth in any one of SEQ ID NOs: 144-146, and the light chain variable region VLcomprises an amino acid sequence as set forth in SEQ ID NO: 142.

[0055] In more preferred embodiments, the heavy chain variable region VHcomprises an amino acid sequence as set forth in SEQ ID NO: 160, and the light chain variable region VLcomprises an amino acid sequence as set forth in any one of SEQ ID NOs: 155-157.

[0056] In some specific embodiments, the antibody or antigen binding fragment further comprises a heavy chain constant region and / or a light chain constant region; optionally, the heavy chain constant region and / or the light chain constant region is selected from a complete constant region sequence or a fragment thereof, the constant region fragment including CH1, hinge region, CH2, CH3, CL and / or Fc region; optionally, the heavy chain constant region is selected from a human or murine IgG1, IgG2, IgG3 or IgG4 constant region, the light chain constant region is selected from a human or murine kappa constant region or lamda constant region.

[0057] In some specific embodiments, the CH1has an amino acid sequence as set forth in SEQ ID NO: 24, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations compared thereto.

[0058] In some specific embodiments, the CLhas an amino acid sequence as set forth in SEQ ID NO: 25, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations compared thereto.

[0059] In some specific embodiments, the Fc region has an amino acid sequence as set forth in SEQ ID NO: 26, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations compared thereto.

[0060] In some specific embodiments, the antibody or antigen binding fragment comprises a heavy chain HC and a light chain LC, wherein,

[0061] the heavy chain HC has an amino acid sequence as set forth in SEQ ID NO: 27, 29, 31, 33, 35 or 37, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations compared thereto; and / or

[0062] The light chain LC has an amino acid sequence as shown in SEQ ID NO: 28, 30, 32, 34, 36, or 38, or an amino acid sequence having at least 70% identity or at most 15 amino acid mutations thereto.

[0063] In some specific embodiments, the at least 70% identity is at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity.

[0064] In some specific embodiments, the at most 3 mutations are preferably at most 3, 2, 1, or 0 mutations.

[0065] In some specific embodiments, the at most 15 mutations are at most 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, or 0 mutations.

[0066] Preferably, the mutations are selected from substitution, deletion, or insertion mutations; preferably, the mutations are back mutations or hot spot mutations; preferably, the substitutions are conservative amino acid substitutions.

[0067] In some specific embodiments, the antibody or antigen binding fragment of the present disclosure comprises:

[0068] (1) a chimeric antibody or fragment thereof; and / or

[0069] (2) a humanized antibody or fragment thereof; and / or,

[0070] (3) a fully human antibody or fragment thereof.

[0071] In some specific embodiments, the antibody or antigen binding fragment is selected from a monoclonal antibody, a polyclonal antibody, a natural antibody, an engineered antibody, a monospecific antibody, a multispecific antibody (e.g., a bispecific antibody), a monovalent antibody, a multivalent antibody, a full-length antibody, an antibody fragment, a naked antibody, a conjugated antibody, a humanized antibody, a fully human antibody, a Fab, a Fab’, a F(ab’)2, a Fd, a Fv, a scFv, a diabody, or a single domain antibody.

[0072] In some specific embodiments, the antibody or antigen binding fragment specifically binds to TL1A; preferably, the TL1A is human TL1A, murine TL1A, and / or monkey TL1A.

[0073] In some specific embodiments, the antibody or antigen-binding fragment exhibits one or more effector functions selected from the group consisting of antibody-dependent cell-mediated cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), and antibody-dependent cellular phagocytosis (ADCP).

[0074] In some specific embodiments, the antibody or antigen-binding fragment is further conjugated to a therapeutic agent or a tracer; preferably, the therapeutic agent is selected from the group consisting of a radioisotope, a chemotherapeutic drug, or an immunomodulatory agent, and preferably, the tracer is selected from the group consisting of a radiological contrast agent, a paramagnetic ion, a metal, a fluorescent label, a chemiluminescent label, an ultrasound contrast agent, and a photosensitizer.

[0075] In some specific embodiments, the antibody or antigen-binding fragment specifically binds to human TL1A, monkey TL1A, and / or murine TL1A, preferably, the antibody or antigen-binding fragment has a KD of less than 1E-6 M, 1E-7 M, 1E-8 M, 1E-9 M, 2E-9 M, 3E-9 M, 4E-9 M, 5E-9 M, 6E-9 M, 7E-9 M, 8E-9 M, 9E-9 M, 1E-10 M, 2E-10 M, 3E-10 M, 4E-10 M, 5E-10 M, 6E-10 M, 8E-10 M, 9E-10 M, 1E-11 M, or 1E-12 M to human TL1A, monkey TL1A, and / or murine TL1A.

[0076] In a second aspect, the present disclosure provides a multispecific antigen-binding molecule comprising the aforementioned antibody or antigen-binding fragment; and another antigen-binding molecule that binds to another antigen other than TL1A, or another antigen-binding molecule that binds to a different epitope of TL1A than the aforementioned antibody or antigen-binding fragment; preferably, the other antigen can be selected from the group consisting of a tumor-specific antigen (TSA), a tumor-associated antigen (TAA), an immune checkpoint, and / or a target that recruits and / or activates an immune cell.

[0077] Preferably, the other antigen-binding molecule is an antibody or an antigen-binding fragment.

[0078] Preferably, the multispecific antigen-binding molecule can be bispecific, trispecific, or tetraspecific.

[0079] Preferably, the multispecific antigen-binding molecule can be bivalent, tetravalent, or hexavalent.

[0080] In a third aspect, the present disclosure provides a chimeric antigen receptor (CAR) comprising an extracellular antigen-binding domain, a transmembrane domain, and an intracellular signaling domain, wherein the extracellular antigen-binding domain comprises the aforementioned antibody or antigen-binding fragment.

[0081] In a fourth aspect, the present disclosure provides an immune effector cell expressing the aforementioned CAR or comprising a nucleic acid fragment encoding the aforementioned CAR; preferably, the immune effector cell is selected from a T cell, a NK cell, a natural killer cell, a NKT cell, a natural killer T cell, a monocyte, a macrophage, a dendritic cell, or a mast cell;

[0082] Preferably, the immune effector cell is an autologous immune effector cell or an allogeneic immune effector cell.

[0083] Preferably, the T cell is selected from a cytotoxic T cell, a regulatory T cell, or a helper T cell.

[0084] In a fifth aspect, the present disclosure provides an isolated nucleic acid fragment encoding the aforementioned antibody or antigen binding fragment, multi-specific antigen binding molecule, or chimeric antigen receptor.

[0085] In a sixth aspect, the present disclosure provides a vector comprising the aforementioned nucleic acid fragment.

[0086] In a seventh aspect, the present disclosure provides a host cell comprising the aforementioned nucleic acid fragment or the aforementioned vector; preferably, the cell is a prokaryotic cell or a eukaryotic cell, such as a bacterium (E. coli), a fungus (yeast), an insect cell, or a mammalian cell (CHO cell line or 293T cell line); preferably, the cell lacks a fucosyltransferase, such as FUT8.

[0087] In an eighth aspect, the present disclosure provides a method of producing the aforementioned antibody or antigen binding fragment, multi-specific antigen binding molecule, the method comprising culturing the aforementioned cell, and isolating the antibody or antigen binding fragment expressed by the cell, or isolating the multi-specific antigen binding molecule expressed by the cell.

[0088] In a ninth aspect, the present disclosure provides a method of producing the aforementioned immune effector cell, the method comprising introducing into the immune effector cell a nucleic acid fragment encoding the aforementioned CAR, and optionally, the method further comprises initiating the immune effector cell to express the aforementioned CAR.

[0089] In a tenth aspect, the present disclosure provides a pharmaceutical composition comprising the aforementioned antibody or antigen binding fragment, multi-specific antigen binding molecule, chimeric antigen receptor, immune effector cell, nucleic acid fragment, vector, or cell; preferably, the composition further comprises a pharmaceutically acceptable carrier, diluent, or adjuvant; and optionally, the pharmaceutical composition further comprises an additional therapeutic agent.

[0090] In an eleventh aspect, the present disclosure provides use of the aforementioned antibody or antigen binding fragment, multispecific antigen binding molecule, chimeric antibody receptor, immune effector cell, nucleic acid fragment, vector, or cell in the manufacture of a medicament for treating an immune-mediated inflammatory disease (IMID); preferably, the immune-mediated inflammatory disease (IMID) can be selected from one or more of inflammatory bowel disease (such as Crohn’s disease and ulcerative colitis), multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes (e.g., type 1 diabetes and type 2 diabetes), vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis (e.g., active hepatitis), myositis, osteoarthritis, pelvic inflammatory disease (PID), an age-related neurodegenerative disease, periodontal disease (e.g., periodontitis), pre-perfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatism, scleroderma, sinusitis, tuberculosis, arteriosclerosis, and uveitis.

[0091] In a twelfth aspect, the present disclosure provides a method of treating an immune-mediated inflammatory disease (IMID), the method comprising administering to a subject an effective amount of the aforementioned antibody or antigen binding fragment, multispecific antigen binding molecule, chimeric antibody receptor, immune effector cell, nucleic acid fragment, vector, or cell; preferably, the immune-mediated inflammatory disease (IMID) can be selected from one or more of inflammatory bowel disease (such as Crohn’s disease and ulcerative colitis), multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes (e.g., type 1 diabetes and type 2 diabetes), vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis (e.g., active hepatitis), myositis, osteoarthritis, pelvic inflammatory disease (PID), an age-related neurodegenerative disease, periodontal disease (e.g., periodontitis), pre-perfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatism, scleroderma, sinusitis, tuberculosis, arteriosclerosis, and uveitis.

[0092] In a thirteenth aspect, the present disclosure provides the aforementioned antibody or antigen binding fragment, multispecific antigen binding molecule, chimeric antigen receptor, immune effector cell, nucleic acid fragment, nucleic acid vector, or host cell for use in the treatment of an immune-mediated inflammatory disease (IMID); preferably, the immune-mediated inflammatory disease (IMID) can be selected from one or more of inflammatory bowel disease (such as Crohn’s disease and ulcerative colitis), multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes (e.g., type 1 diabetes and type 2 diabetes), vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis (e.g., active hepatitis), myositis, osteoarthritis, pelvic inflammatory disease (PID), neurodegenerative disease of aging, periodontal disease (e.g., periodontitis), pre-perfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatism, scleroderma, sinusitis, tuberculosis, arteriosclerosis, and uveitis.

[0093] In a fourteenth aspect, the present disclosure provides a kit comprising the aforementioned antibody or antigen binding fragment, multispecific antigen binding molecule, immune effector cell, nucleic acid fragment, vector, product of manufacture or pharmaceutical composition prepared according to the aforementioned methods.

[0094] Definitions and explanations of terms

[0095] Unless otherwise defined herein, scientific and technical terms used in connection with the present disclosure shall have the meanings that are commonly understood by those of ordinary skill in the art.

[0096] Further, unless otherwise noted, singular forms of terms herein are intended to include the plural forms as well, and vice versa. More specifically, as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless explicitly stated otherwise.

[0097] The terms "comprise", "contain", and "have" are used interchangeably in this document and are intended to mean the inclusion of the elements specified in the scheme, meaning that the scheme can have other elements in addition to the listed elements. It should also be understood that the use of "comprise", "contain" and "have" is described herein also provides a "consisting of" scheme. Exemplarily, "a composition comprising A and B" should be understood as a technical scheme consisting of a composition of A and B, and a composition containing other components in addition to A and B, both of which fall within the scope of the aforementioned "a composition".

[0098] The term "and / or", when used herein, includes the meaning of "and", "or", and "all or any other combination of the elements linked to the term".

[0099] The term "TLIA", also known as "TNF ligand-related molecule 1 and vascular endothelial growth inhibitor (VEGI)" or "tumor necrosis factor superfamily member 15 (TNFSF15)", belongs to the tumor necrosis factor family, and is a ligand of DR3 and decoy receptor TR6 / DcR3. "TL1A" herein includes TL1A full-length protein or its mutants (e.g. point mutation, insertion mutation or deletion mutation), splice variants, Orthologs and fragments of TL1A. "TL1A" herein can be derived from human, primates (e.g. cynomolgus monkey, rhesus monkey) and rodents (e.g. mouse, rat). Exemplarily, the human TL1A amino acid sequence can be found at Uniprot No. O95159, the cynomolgus monkey TL1A amino acid sequence can be found at Uniprot No. G7PRK8, and the mouse TL1A amino acid sequence can be found at Uniprot No. Q5UBV8.

[0100] The terms "specific binding", "immunobinding" and "immunobinding property" herein refer to the type of non-covalent interaction that occurs between immunoglobulin molecules and antigens for which the immunoglobulin has specificity, and are antigen-binding molecules (e.g. bodies) that specifically bind to an antigen and substantially the same antigen generally with high affinity, but do not bind to unrelated antigens with high affinity. The strength or affinity of an immunobinding interaction is usually reflected in the equilibrium dissociation constant (KD), where a lower KD indicates a higher affinity, and the immunobinding property of a selected polypeptide can be quantitatively determined using methods well known in the art. Taking antibodies as an example, high affinity usually refers to a KD of about 10 -8 M or less, about 1 x 10 -9 M or less, about 1 x 10 -10 M or less, 1 x 10 -11 M or less, or 1 x 10 -12 M or less. The KD is calculated as follows: KD = Kd / Ka, where Kd represents the dissociation rate and Ka represents the association rate. The equilibrium dissociation constant KD can be measured by methods well known in the art, such as surface plasmon resonance (e.g. Biacore) or equilibrium dialysis, and exemplary methods for obtaining KD values are shown in Example 8 herein.

[0101] The term "antigen binding molecule" is used herein in the broadest sense, and refers to a molecule that specifically binds to an antigen. Exemplary, antigen binding molecules include, but are not limited to, antibodies or antibody mimetics. An "antibody mimetic" refers to an organic compound or binding domain that is capable of specifically binding to an antigen, but is not related to the structure of an antibody. Exemplary, antibody mimetics include, but are not limited to, affibodies, affitins, affilins, designed ankyrin repeat proteins (DARPins), aptamers, or Kunitz domain peptides.

[0102] The term "antibody" is used herein in the broadest sense, and refers to a polypeptide or combination of polypeptides that contains a sufficient sequence from a heavy chain variable region of an immunoglobulin and / or a sufficient sequence from a light chain variable region of an immunoglobulin to be capable of specifically binding to an antigen. "Antibody" herein encompasses various forms and various structures, as long as they exhibit the desired antigen binding activity. The terms "full-length antibody," "intact antibody," and "whole antibody" are used herein interchangeably to refer to an antibody having a structure substantially similar to a native antibody structure. "Antibody" herein includes, but is not limited to, a monoclonal antibody, a polyclonal antibody, a monospecific antibody, a multispecific antibody (e.g., a bispecific antibody), a monovalent antibody, a multivalent antibody, a whole antibody, a fragment of a whole antibody, a naked antibody, a conjugated antibody, a chimeric antibody, a humanized antibody, or a fully human antibody.

[0103] The terms "antibody fragment" or "antigen binding fragment" are used herein interchangeably, and refer to a portion of an antibody that does not possess the full structure of an intact antibody, but only contains a portion or a local variant of an intact antibody that possesses the ability to bind to the same antigen recognized by the intact antibody. "Antibody fragment" or "antigen binding fragment" also includes any synthetic or genetically engineered protein that functions like an antibody by binding to a particular antigen to form a complex. For example, antibody fragments include isolated fragments consisting of the light chain variable region, "Fv" fragments consisting of the heavy and light chain variable regions, recombinant single chain polypeptide molecules (scFv) in which the light and heavy chain variable regions are connected by a peptide linker, and minimal recognition units consisting of the amino acid residues that mimic the hypervariable region. "Antigen binding fragment" or "antibody fragment" herein includes, but is not limited to, Fab, F(ab')2, Fab', Fab'-SH, Fd, Fv, scFv, diabodies, and single domain antibodies.

[0104] An "antibody" herein also includes surrogate protein scaffolds or artificial scaffolds with grafted complementarity determining regions (CDRs) or CDR derivatives. Such scaffolds include antibody-derived scaffolds (which comprise mutations introduced to, for example, stabilize the three-dimensional structure of an antibody) as well as wholly synthetic scaffolds comprising, for example, biocompatible polymers. See, for example, Korndorfer et al., 2003, Proteins: Structure, Function, and Bioinformatics, 53(1): 121-129 (2003); Roque et al., Biotechnol. Prog. 20:639-654 (2004). Such scaffolds can also include non-antibody-derived scaffolds, such as scaffold proteins known in the art to be useful for grafting CDRs, including but not limited to, tenascin, fibronectin, peptide aptamers, and the like.

[0105] An "antibody" herein includes a typical "four-chain antibody" which belongs to an immunoglobulin composed of two heavy chains (HC) and two light chains (LC); a heavy chain refers to a polypeptide chain which consists of, in the direction from N-terminus to C-terminus, a heavy chain variable region (VH), a heavy chain constant region CH1 domain, a hinge region (HR), a heavy chain constant region CH2 domain, a heavy chain constant region CH3 domain; and, when the full-length antibody is of an IgE isotype, a heavy chain constant region CH4 domain is optionally included; a light chain is a polypeptide chain which consists of, in the direction from N-terminus to C-terminus, a light chain variable region (VL) and a light chain constant region (CL); the heavy chains and the heavy chains, the heavy chains and the light chains are connected by disulfide bonds, forming a "Y" shape. Due to the difference in the amino acid composition and the order of arrangement of the immunoglobulin heavy chain constant region, the antigenicity is also different. Accordingly, the "immunoglobulin" herein can be divided into five categories, or called isotypes of immunoglobulin, namely IgM, IgD, IgG, IgA and IgE, and the corresponding heavy chains are μ chain, δ chain, γ chain, α chain and ε chain, respectively. The same category of Ig can be divided into different subcategories according to the difference in the amino acid composition of the hinge region and the number and position of the heavy chain disulfide bond, such as IgG can be divided into IgG1, IgG2, IgG3, IgG4, IgA can be divided into IgA1 and IgA2. The light chain is divided into κ chain or λ chain through the constant region. Each of the five categories of Ig can have κ chain or λ chain.

[0106] The "antibodies" herein also include antibodies that do not comprise light chains, such as heavy-chain antibodies (HCAbs) produced by Camelus dromedarius, Camelus bactrianus, Lama glama, Lama guanicoe, and Vicugna pacos, and the like, and immunoglobulin new antigen receptor (IgNAR) found in cartilaginous fishes such as sharks.

[0107] The "antibodies" herein can be derived from any animal, including but not limited to humans and non-human animals, which can be selected from primates, mammals, rodents, and vertebrates, such as a camelid, a llama, a guanaco, an alpaca, a sheep, a rabbit, a mouse, a rat, or a cartilaginous fish (e.g., a shark).

[0108] The term "monoclonal antibody" as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and / or bind the same epitope, except for possible variants that can arise during production of the antibody, such variants are typically present in minor amounts. In contrast to polyclonal antibody preparations, which typically include different antibodies directed against different determinants (epitopes), each monoclonal antibody of a monoclonal antibody preparation is directed against a single determinant on an antigen. The modifier "monoclonal" is not to be construed as requiring production of the antibody by any particular method. For example, the monoclonal antibodies to be used in accordance with the present application can be made by a variety of techniques, including but not limited to the hybridoma method, recombinant DNA methods, phage-display methods, and transgenic animals containing all or part of the human immunoglobulin loci, and other methods known in the art.

[0109] The term "natural antibody" as used herein refers to an antibody produced and paired by the immune system of a multicellular organism. The term "engineered antibody" as used herein refers to a non-natural antibody obtained by genetic engineering, antibody engineering, and the like. Exemplarily, the "engineered antibody" includes a chimeric antibody, a humanized antibody, an antibody fragment (e.g., scFv, sdAb, and the like), a bispecific antibody, and the like.

[0110] The term "monospecific" refers to having one or more binding sites, wherein each binding site binds the same epitope of the same antigen.

[0111] The term "multispecific antibody" refers to an antibody having at least two antigen binding sites, each of which binds to a different epitope of the same antigen or to different epitopes of different antigens. Thus, terms such as "bispecific", "trispecific", "quadrispecific", and the like refer to the number of different epitopes to which the antibody / antigen binding molecule can bind.

[0112] The term "valency" denotes the presence of a specified number of binding sites for an antibody / antigen binding molecule. Thus, the terms "monovalent", "bivalent", "tetravalent", and "hexavalent" refer to the presence of one, two, four, and six binding sites, respectively, in an antibody / antigen binding molecule.

[0113] Papain digestion of intact antibodies produces two identical antigen binding fragments, called "Fab" fragments, each with a heavy and a light chain variable domain, and half a constant domain. Thus, the term "Fab fragment" herein refers to a light chain fragment which comprises the VL and CL domains of the light chain and a heavy chain fragment which comprises the VH and CH1 domains of the heavy chain. Fab' fragments differ from Fab fragments by the addition of a few residues at the carboxy terminus of the CH1 domain of the heavy chain, including one or more cysteines from the antibody hinge region. Fab'-SH is the designation herein for Fab' in which the cysteine residue(s) of the constant domains carry(s) a free thiol group. Pepsin treatment yields an F(ab')2 fragment that has two antigen binding sites (two Fab fragments) and a part of the Fc region.

[0114] The term "Fd" herein refers to an antibody consisting of the VH and CH1 domains. The term "Fv" herein refers to an antibody fragment which consists of the VL and VH domains of a single arm of an antibody. The Fv fragment typically is considered the smallest antibody fragment that is capable of forming a complete antigen binding site, the Fv fragment has the same binding characteristics as a Fab and a similar three-dimensional binding characteristic, the VH and VL chains of the Fv fragment are held together by noncovalent interactions. It is generally considered that the six CDRs confer the antigen binding specificity to an antibody. However, even a single variable domain (e.g., an Fd fragment which contains only three CDRs specific for an antigen) is capable of recognizing and binding antigen, although at a lower affinity than the entire binding site.

[0115] The term "scFv" (single-chain variable fragment) herein refers to a single polypeptide chain comprising a VL and a VH domain, wherein the VL and VH are connected by a linker. Such scFv molecules can have the general structure: NH2-VL-linker-VH-COOH or NH2-VH-linker-VL-COOH. Suitable prior art linkers consist of repeating GGGGS amino acid sequences or variants thereof. For example, a linker having the amino acid sequence (GGGGS)4 can be used, but variants thereof can also be used. Other linkers useful in the present disclosure are described by Alfthan et al. (1995), Protein Eng. 8:725-731, Choi et al. (2001), Eur. J. Immunol. 31 :94-106, Hu et al. (1996), Cancer Res. 56:3055-3061, Kipriyanov et al. (1999), J. Mol. Biol. 293:41-56, and Roovers et al. (2001), Cancer Immunol. In some cases, a disulfide bond between the VH and VL of the scFv can also be present, forming a disulfide-bonded Fv (dsFv).

[0116] The term "diabody" herein, the VH and VL domains of which are expressed in a single polypeptide chain, but using a too short linker such that it does not allow pairing between the two domains of the same chain, thereby forcing the domains to pair with the complement domains of another chain and creating two antigen binding sites.

[0117] The terms "single domain antibody", "VHH" and "nanobody" have the same meaning and are used interchangeably herein and refer to the variable region of the heavy chain of a cloned antibody, which is constructed to consist of only one heavy chain variable region, which is the smallest antigen binding fragment with full functionality. Usually, the variable region of the heavy chain of an antibody is cloned after obtaining an antibody naturally missing the light chain and the constant region 1 (CH1) of the heavy chain, which is constructed to consist of only one heavy chain variable region. Single domain antibodies can be derived from camelid heavy chain antibodies or cartilaginous fish IgNAR.

[0118] The term "chimeric antibody" herein refers to an antibody in which a portion of the light or / and heavy chain is derived from one antibody (which can be derived from a particular species or belong to a particular antibody class or subclass), and the other portion of the light or / and heavy chain is derived from another antibody (which can be derived from the same or a different species or belong to the same or a different antibody class or subclass), but retains the binding activity to the target antigen. For example, the term "chimeric antibody" can include an antibody (e.g., human murine chimeric antibody) in which the heavy and light chain variable regions of the antibody are from a first antibody (e.g., a murine antibody), while the heavy and light chain constant regions of the antibody are from a second antibody (e.g., a human antibody).

[0119] The term "humanized antibody" herein refers to a non-human-derived antibody that has been genetically engineered to have an amino acid sequence that is modified to increase homology to the sequence of a human-derived antibody. Typically, a humanized antibody has all or a portion of the CDR regions from a non-human-derived antibody (donor antibody) and all or a portion of the non-CDR regions (e.g., FRs in the variable region and / or constant region) from a human-derived immunoglobulin (acceptor antibody). A humanized antibody typically retains or partially retains the desired properties of the donor antibody, including but not limited to, antigen specificity, affinity, reactivity, ability to enhance immune cell activity, ability to enhance immune response, etc.

[0120] The term "fully human antibody" herein refers to an antibody having a variable region in which both the FR and CDR are derived from human germline immunoglobulin sequences. In addition, if the antibody comprises a constant region, the constant region is also derived from human germline immunoglobulin sequences. A "fully human antibody" herein can include amino acid residues that are not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by in vitro random or site-specific mutagenesis or by in vivo somatic mutation). However, a "fully human antibody" herein does not include antibodies in which CDR sequences derived from the germline of another mammalian species (e.g., mouse) have been grafted onto human framework sequences.

[0121] The term "naked antibody" herein refers to an antibody that is not linked, fused, or conjugated to another agent or molecule (e.g., a label or a drug), a peptide, or a polypeptide. In particular embodiments, a naked antibody expressed by a mammalian host cell can be glycosylated by the glycosylation machinery (e.g., glycosylation enzymes) of the host cell. In certain embodiments, a naked antibody is not glycosylated when expressed by a host cell that does not have its own glycosylation machinery (e.g., glycosylation enzymes). In certain embodiments, a naked antibody is a whole antibody, while in other embodiments, a naked antibody is an antigen-binding fragment of a whole antibody.

[0122] The term "variable region" herein refers to the region of an antibody heavy or light chain that is involved in binding the antibody to an antigen, "variable region of the heavy chain" and "VH" or "HCVR" are used interchangeably, and "variable region of the light chain" and "VL" or "LCVR" are used interchangeably. The variable domains of the heavy chain and light chain (VH and VL, respectively) in a native antibody generally have similar structures and each domain comprises four conserved framework regions (FRs) and three hypervariable regions (HVRs). A single VH or VL domain can be sufficient to confer antigen-binding specificity. The term "complementarity determining region" and "CDR" are used interchangeably herein to refer to the hypervariable regions of a heavy chain variable region (VH) or light chain variable region (VL), which are also referred to as hypervariable loops (HVRs), because of their high degree of sequence variability in antibodies that specifically bind to antigens, and their ability to form structurally constrained, antigen binding sites on antibody-antigen interaction. The heavy chain variable region CDRs can be abbreviated as HCDR and the light chain variable region CDRs can be abbreviated as LCDR. The term "framework region" or "FR region" is interchangeable and refers to those amino acid residues in a variable region of a heavy chain or light chain of an antibody that are outside the CDRs. Typically, a canonical antibody variable region is composed of 4 FR regions and 3 CDR regions in the following order: FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4.

[0123] The CDRs herein can be annotated and defined in ways known in the art, including but not limited to the Kabat numbering system, the Chothia numbering system, or the IMGT numbering system, using tools websites including but not limited to the AbRSA website (http: / / cao.labshare.cn / AbRSA / cdrs.php), the abYsis website (www.abysis.org / abysis / sequence_input / key_annotation / key_annotation.cgi), and the IMGT website (http: / / www.imgt.org / 3Dstructure-DB / cgi / DomainGapAlign.cgi#results). The CDRs herein include overlaps and subsets of amino acid residues defined in different ways.

[0124] The term "Kabat numbering system" herein generally refers to the immunoglobulin alignment and numbering system proposed by Elvin A. Kabat (see, e.g., Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991).

[0125] The term "Chothia numbering system" herein generally refers to an immunoglobulin numbering system proposed by Chothia et al. that is based on the location of structural loops to identify CDR region boundaries (see, e.g., Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883).

[0126] The term "IMGT numbering system" herein generally refers to a numbering system based on The international ImMunoGeneTics information system (IMGT) initiated by Lefranc et al., see Lefranc et al., Dev. Comparat. Immunol. 27:55-77, 2003.

[0127] The term "heavy chain constant region" herein refers to the carboxy-terminal portion of an antibody heavy chain that is not directly involved in binding of an antibody to an antigen, but exhibits effector functions such as interaction with Fc receptors, which has a more conserved amino acid sequence relative to the variable domain of the antibody. The "heavy chain constant region" comprises at least: a CH1 domain, a hinge region, a CH2 domain, a CH3 domain, or a variant or fragment thereof. The "heavy chain constant region" includes both "full length heavy chain constant region" and "heavy chain constant region fragment", the former has substantially similar structure as the native antibody constant region, while the latter only includes "a portion of the full length heavy chain constant region". Exemplarily, a typical "full length antibody heavy chain constant region" consists of CH1 domain-hinge region-CH2 domain-CH3 domain; when the antibody is IgE, it further includes CH4 domain; when the antibody is heavy chain antibody, it does not include CH1 domain. Exemplarily, a typical "heavy chain constant region fragment" can be selected from CH1, Fc or CH3 domain.

[0128] The term "light chain constant region" herein refers to the carboxy-terminal portion of an antibody light chain that is not directly involved in binding of an antibody to an antigen, which can be selected from constant kappa domain or constant lambda domain.

[0129] The term "Fc" herein refers to the carboxy-terminal portion of an intact antibody that results from papain digestion, typically comprising the CH3 and CH2 domains of an antibody. The Fc region includes, e.g., native-sequence Fc regions, recombinant Fc regions, and variant Fc regions. Although the boundaries of the Fc region of an immunoglobulin heavy chain might vary slightly, the Fc region of a human IgG heavy chain typically extends from the amino acid residue at position Cys226, or from Pro230 to its carboxy-terminus. The C-terminal lysine (residue 447 according to the Kabat numbering system) of the Fc region can be removed, e.g., during production or purification of the antibody, or by recombinant engineering of nucleic acid encoding the antibody heavy chain, and therefore, the Fc region can include or not include the C-terminal lysine (Lys447), or the C-terminal glycine (Gly446) and lysine (Lys447).

[0130] The term "epitope" herein includes any protein determinant capable of specific binding to an immunoglobulin, scFv or T-cell receptor. The term "epitope" includes any protein determinant capable of specific binding to an immunoglobulin or T-cell receptor. Epitopic determinants usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three-dimensional structural characteristics, as well as specific charge characteristics. For example, an antibody can be raised against an N-terminal or C-terminal peptide of a polypeptide.

[0131] The term "mutation" herein includes genetic mutation and amino acid mutation, wherein genetic mutation refers to deletion, insertion of heterologous nucleic acid, inversion or substitution, which can lead to changes in amino acid sequence in the corresponding protein product; amino acid mutation, also known as non-synonymous single nucleotide mutation, is due to the change of some single bases, resulting in changes in amino acid sequence in the protein product. Changes in amino acids affect protein stability, interactions and enzyme activity, leading to the occurrence of diseases.

[0132] The term "amino acid" as used herein refers to the basic unit that makes up a protein, conferring a specific molecular structure to the protein, which imparts biochemical activity to the molecule. In chemistry, amino acids refer to organic compounds that contain an amino group (-NH2) and a carboxyl group (-COOH) in their structure. Depending on the position of the amino group attached to the carbon atom in the carboxylic acid, amino acids can be classified as alpha, beta, gamma, delta, etc. In alpha amino acids, the amino and carboxyl groups are attached to the same carbon atom, in beta amino acids, the amino and carboxyl groups are attached to adjacent carbon atoms, and so on. In biology, amino acids usually refer specifically to alpha amino acids, i.e., amino acids in which the amino and carboxyl groups are attached directly to the same -CH- structure, and have the general formula H2NCHRCOOH (R represents some organic substituent). Exemplary, the common 20 amino acids include glycine, alanine, valine, leucine, isoleucine, phenylalanine, tryptophan, tyrosine, aspartate, histidine, asparagine, glutamate, lysine, glutamine, methionine, arginine, serine, threonine, cysteine, and proline.

[0133] The term "amino acid substitution" as used herein refers to those in which at least one amino acid residue in the native or starting sequence is removed and a different amino acid is inserted in its place. Substitutions can be single, in which only one amino acid in the molecule has been substituted, or they can be multiple, in which two or more amino acids in the same molecule have been substituted.

[0134] The term "conservative amino acid substitution" herein refers to the replacement of an amino acid normally present in a sequence with a different amino acid having similar size, charge, or polarity. Examples of conservative substitutions include the substitution of non-polar (hydrophobic) residues such as isoleucine, valine, and leucine; the substitution of one polar (hydrophilic) residue for another such as between arginine and lysine, between glutamine and asparagine, and between glycine and serine. Additionally, the substitution of a basic residue such as lysine, arginine, or histidine for another, or the substitution of one acidic residue such as aspartic acid or glutamic acid for another, are additional examples of conservative substitutions. Examples of non-conservative substitutions include the substitution of a non-polar (hydrophobic) amino acid residue such as isoleucine, valine, leucine, alanine, methionine for a polar (hydrophilic) residue such as cysteine, glutamine, glutamic acid, or lysine, and / or the substitution of a polar residue for a non-polar residue.

[0135] The term "mutant" herein refers to a "variant" of the protein or peptide can have at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99% amino acid identity to the amino acid sequence of the protein or peptide.

[0136] The term "nucleic acid" herein includes any compound and / or substance comprising a polymer of nucleotides. Each nucleotide is composed of a base, in particular a purine or pyrimidine base (i.e. cytosine (C), guanine (G), adenine (A), thymine (T) or uracil (U)), a sugar (i.e. deoxyribose or ribose) and a phosphate group. Typically, a nucleic acid molecule is described by the sequence of bases, whereby the bases represent the primary structure (linear structure) of the nucleic acid molecule. The sequence of bases is usually denoted 5' to 3'. In the present context, the term nucleic acid molecule encompasses deoxyribonucleic acids (DNA), including, for example, complementary DNA (cDNA) and genomic DNA, ribonucleic acids (RNA), in particular messenger RNA (mRNA), synthetic forms of DNA or RNA, and polymers comprising a mixture of two or more of these molecules. The nucleic acid molecule can be linear or circular. Furthermore, the term nucleic acid molecule includes both the sense and anti-sense strands, as well as single- and double-stranded forms. Also, the nucleic acid molecules described herein can contain naturally-occurring or non-naturally-occurring nucleotides. Examples of non-naturally-occurring nucleotides include modified nucleotide bases with derivatized sugar or phosphate backbone linkages or chemically modified residues. The nucleic acid molecule also encompasses DNA and RNA molecules which are suitable as vectors for the direct expression of an antibody of the present disclosure in vitro and / or in vivo, e.g. in a host or patient. Such DNA (e.g. cDNA) or RNA (e.g. mRNA) vectors can be unmodified or modified. For example, the mRNA can be chemically modified to enhance the stability of the RNA vector and / or the expression of the encoded molecule, so that the mRNA can be injected into a subject to produce the antibody in vivo.

[0137] The term "identity" herein can be calculated by determining the percent "identity" of two amino acid sequences or two nucleic acid sequences, by aligning the sequences for optimal comparison purposes (e.g., gaps can be introduced in the sequence of one or both of the first and second amino acid sequences or nucleic acid sequences for optimal alignment and non-homologous sequences can be disregarded for comparison purposes). The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position.

[0138] The percent identity between two sequences is varied as a function of the number of identical positions shared by the sequences, taking into account the number of gaps required to achieve an optimal alignment of the two sequences and the length of each gap.

[0139] Sequence comparisons and percent identity calculations between two sequences can be accomplished using a mathematical algorithm. For instance, the percent identity between two amino acid sequences can be determined using the Needleman and Wunsch ((1970) J. Mol. Biol. 48: 444-453) algorithm which has been incorporated into the GAP program in the GCG software package (available at www.gcg.com), using either a Blossum 62 matrix or a PAM250 matrix, and a gap penalty of 16, 14, 12, 10, 8, 6, or 4, and a gap length penalty of 1, 2, 3, 4, 5, or 6. By way of further example, the percent identity between two nucleotide sequences can be determined using the GAP program (available at www.gcg.com), using a NWSgapdna.CMP matrix and gap weight of 40, 50, 60, 70, or 80, and a gap length penalty of 1, 2, 3, 4, 5, or 6. An especially preferred parameter set (and the one that should be used unless otherwise indicated) is a Blossum 62 scoring matrix with a gap penalty of 12, a gap extend penalty of 4, and a frameshift gap penalty of 5.

[0140] The percent identity between two amino acid or nucleotide sequences can also be determined using the algorithm of E. Meyers and W. Miller ((1989) CABIOS, 4: 11-17) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4.

[0141] Additionally or alternatively, the nucleic acid sequences and protein sequences described in the present disclosure can be further used as a "query sequence" to perform a search against public databases to, for example, identify other family members or related sequences. Such searches can be performed using the NBLAST and XBLAST programs (version 2.0) of Altschul et al. ((1990) J. Mol. Biol. 215:403-10). BLAST nucleotide searches can be performed with the NBLAST program. BLAST protein searches can be performed with the XBLAST program to achieve amino acid sequences homologous to the protein molecules of the present disclosure. When using BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used. See www.ncbi.nlm.nih.gov.

[0142] The term "antigen chimeric receptor (CAR)" herein refers to an artificial cell surface receptor engineered to be expressed on an immune effector cell and specifically binds an antigen, which comprises at least (1) an extracellular antigen binding domain, such as a variable heavy chain or light chain of an antibody, (2) a transmembrane domain anchoring the CAR into the immune effector cell, and (3) an intracellular signaling domain. CARs are capable of redirecting T cells and other immune effector cells to a selected target, such as a cancer cell, in a non-MHC restricted manner using the extracellular antigen binding domain.

[0143] As used herein, the term "vector" includes nucleic acid vectors, such as DNA vectors (e.g., plasmids), RNA vectors, viruses, or other suitable replicons (e.g., viral vectors). A variety of vectors have been developed for the delivery of polynucleotides encoding exogenous proteins into prokaryotic or eukaryotic cells. The expression vectors of the present disclosure contain polynucleotide sequences as well as additional sequence elements, such as for example, for the expression of proteins and / or for the integration of these polynucleotide sequences into the genome of a mammalian cell. Certain vectors that can be used for the expression of the antibodies and antibody fragments of the present disclosure include plasmids containing regulatory sequences that direct the transcription of genes, such as promoter and enhancer regions. Other useful vectors for the expression of antibodies and antibody fragments contain polynucleotide sequences that enhance the rate of translation of these genes or improve the stability or nuclear export of mRNA produced from the transcription of genes. These sequence elements include, for example, 5' and 3' untranslated regions, internal ribosome entry sites (IRES), and polyadenylation signal sites in order to direct efficient transcription of genes carried on the expression vectors. The expression vectors of the present disclosure can also contain a polynucleotide that encodes a marker for the selection of cells containing such vectors. Examples of suitable markers include genes that encode antibiotic (e.g., ampicillin, chloramphenicol, kanamycin, or neomycin) resistance.

[0144] The term "host cell" herein refers to a cell into which a foreign nucleic acid has been introduced, including the progeny of such a cell. Host cells include "transformants" and "transformed cells," which include both the primary transformed cell and progeny of the primary transformed cell that are not identical to the parent cell in that they do not contain the original nucleic acid content of the parent cell. The progeny can not be completely identical to the parent cell in nucleic acid content, but can contain mutations. Mutant progeny that have the same function or biological activity as screened or selected for in the original transformed cell are included herein.

[0145] The term "pharmaceutical composition" herein refers to a preparation which is in a form suitable for its administration to a subject in need of said pharmaceutical composition and which allows the biological activity of the active ingredients contained therein to be effective, and which does not contain additional ingredients which are unacceptable with respect to toxicity to the subject to which the pharmaceutical composition is administered.

[0146] The term "pharmaceutically acceptable excipient," "pharmaceutically acceptable carrier," or "pharmaceutically acceptable carrier, diluent or adjuvant" as used herein refers to an excipient that does not stimulate an undesirable response in an organism, and does not substantially interfere with the biological activity and / or property of the active compound. Suitable excipients are well known to those skilled in the art, and include, for example, carbohydrates, waxes, water soluble and / or swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, and the like.

[0147] The term "treatment" as used herein refers to a surgical or therapeutic treatment whose purpose is to prevent, slow down (reduce), or halt the progression of an undesirable physiological change or condition, such as cancer, immune-mediated inflammatory diseases (IMIDs) (e.g., autoimmune and inflammatory diseases), and viral infections, in a subject. Beneficial or desired results include, but are not limited to, alleviation of symptoms, diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, and remission (whether partial or total), whether detectable or undetectable. A subject in need of treatment includes a subject who has a condition or disease, as well as a subject who is predisposed to having a condition or disease or who is intended to prevent a condition or disease. When referring to the terms slow down, reduce, diminish, palliate, alleviate, and the like, the meaning also includes elimination, disappearance, nonoccurrence, and the like.

[0148] The term "subject," "object," "patient" as used herein refers to an organism that receives treatment for a particular disease or condition as described in the present disclosure. Examples of subjects and patients include mammals, such as humans, primates (e.g., monkeys), or non-primate mammals, that receive treatment for a disease or condition. "Patient" also refers to an organism that receives treatment for a particular disease or condition as described herein, such as an immune-mediated inflammatory disease (IMID) or cancer.

[0149] The term "effective amount" as used herein refers to that amount of a therapeutic agent, alone or in combination with another therapeutic agent, which is effective to prevent or alleviate a disease condition or the progression of the disease in a cell, tissue, or subject. An "effective amount" also refers to the amount of a compound which is sufficient to reduce symptoms, e.g., to treat, cure, prevent, or alleviate the relevant medical condition, or to increase the rate of treatment, cure, prevention, or alleviation of these conditions. When the active ingredient is administered individually to an individual, the therapeutically effective dose refers to the amount of the ingredient alone. When a combination is used, the therapeutically effective dose refers to the combined amount of the active ingredients that produces the therapeutic effect, whether administered in combination, sequentially, or simultaneously.

[0150] The term "immune-mediated inflammatory disease" or "IMID" herein refers to any group of conditions or diseases that lack a clear etiology but are characterized by common inflammatory pathways that lead to inflammation, and which can result from or be triggered by a dysregulation of the normal immune response. "Immune-mediated inflammatory disease" herein encompasses autoimmune diseases and inflammatory diseases.

[0151] "Autoimmune disease" herein refers to a condition in which a subject mounts an immune response against its own cells, tissues and / or organs, resulting in damage to the cells, tissues and / or organs. "Inflammatory disease" herein refers to a condition in a subject characterized by inflammation, preferably chronic inflammation. An autoimmune condition can or can not be accompanied by inflammation. Further, inflammation can or can not be caused by an autoimmune condition. The terms "autoimmune disease", "autoimmune disorder" and "autoimmune-related disease" can be used interchangeably herein.

[0152] In some embodiments, exemplary immune-mediated inflammatory diseases include inflammatory bowel disease (such as Crohn's disease and ulcerative colitis), multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes (e.g., type 1 diabetes and type 2 diabetes), vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis (e.g., active hepatitis), myositis, osteoarthritis, pelvic inflammatory disease (PID), neurodegenerative diseases of aging, periodontal disease (e.g., periodontitis), preperfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatic disease, scleroderma, sinusitis, tuberculosis, arteriosclerosis, uveitis, and the like.

[0153] The term "cancer" in this document refers to or describes a physiological condition in mammals characterized by unregulated cell growth. This definition includes both benign and malignant cancers. The term "tumor" or "neoplastic tumor" in this document refers to all neoplasmic cell growth and proliferation, whether malignant or benign, and all pre-cancerous and cancerous cells and tissues. The terms "cancer" and "tumor" are not mutually exclusive when used herein. In some embodiments, exemplary cancers include: adenoid cystic carcinoma, adrenal carcinoma, amyloidosis, anal cancer, ataxia-telangiectasia, atypical nevus syndrome, basal cell carcinoma, cholangiocarcinoma, and Birt Hogg's disease. Dube syndrome, bladder cancer, bone cancer, brain tumor, breast cancer, male breast cancer, carcinoid tumor, cervical cancer, colorectal cancer, ductal carcinoma, endometrial cancer, esophageal cancer, gastric cancer, gastrointestinal stromal tumor (GIST), HER2-positive breast cancer, islet cell tumor, juvenile polyposis syndrome, kidney cancer, laryngeal cancer, acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myeloid leukemia (CML), liver cancer, lobular carcinoma, lung cancer, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), Hodgkin lymphoma, non-Hodgkin's lymphoma. Golden lymphoma, malignant glioma, melanoma, meningioma, multiple myeloma, myelodysplastic syndrome (MDS), nasopharyngeal carcinoma, neuroendocrine tumor, oral cancer, osteosarcoma, ovarian cancer, pancreatic cancer, pancreatic neuroendocrine tumor, parathyroid carcinoma, penile cancer, peritoneal cancer, Peutz-Jeghers syndrome, pituitary adenoma, polycythemia vera, prostate cancer, renal cell carcinoma, retinoblastoma, salivary gland cancer, sarcoma, Kaposi's sarcoma, skin cancer, small intestine cancer, gastric cancer, testicular cancer, thymoma, thyroid cancer, uterine (endometrial) cancer, vaginal cancer, and nephroblastoma, etc. Attached Figure Description

[0154] Figure 1 shows the binding activity of the anti-TL1A chimeric antibody to the human TL1A fusion protein.

[0155] Figure 2 shows the binding activity of the anti-TL1A chimeric antibody to the cynomolgus monkey TL1A fusion protein.

[0156] Figure 3 shows the binding activity of the anti-TL1A chimeric antibody to the mouse TL1A fusion protein.

[0157] Figure 4 shows the binding activity of the anti-TL1A chimeric antibody to the CHO-K1 human TL1A-overexpressing cell line.

[0158] Figure 5 shows the effect of the anti-TL1A chimeric antibody in blocking the interaction between human TL1A and CHO-K1-human DR3 cells.

[0159] Figure 6 shows the effect of anti-TLlA chimeric antibodies on blocking the interaction of human TLlA with DcR3 protein.

[0160] Figure 7 shows the effect of anti-TLlA chimeric antibodies on inhibiting NFκB activity in TF-1-NFκB-luciferase cells.

[0161] Figure 8 shows the effect of anti-TLlA chimeric antibodies on inhibiting apoptosis activity in TF-1 cells.

[0162] Figure 9 shows the effect of anti-TLlA chimeric antibodies on inhibiting IFN-γ secretion from PBMC stimulated by human TLlA and IL-12 and IL-18.

[0163] Figure 10 shows the binding activity of anti-TLlA humanized antibodies to human TLlA fusion protein.

[0164] Figure 11 shows the binding activity of anti-TLlA humanized antibodies to cynomolgus TLlA fusion protein.

[0165] Figure 12 shows the binding activity of anti-TLlA humanized antibodies to mouse TLlA fusion protein.

[0166] Figure 13 shows the binding activity of anti-TLlA humanized antibodies to CHO-K1 human TLlA high expression cell line.

[0167] Figure 14 shows the effect of anti-TLlA humanized antibodies on blocking the interaction of human TLlA with CHO-K1-human DR3 cells.

[0168] Figure 15 shows the effect of anti-TLlA humanized antibodies on blocking the interaction of human TLlA with DcR3 protein.

[0169] Figure 16 shows the effect of anti-TLlA humanized antibodies on inhibiting NFκB activity in TF-1-NFκB-luciferase cells.

[0170] Figure 17 shows the effect of anti-TLlA humanized antibodies on inhibiting apoptosis activity in TF-1 cells.

[0171] Figure 18 shows the effect of anti-TLlA humanized antibodies on inhibiting IFN-γ secretion from PBMC stimulated by human TLlA and IL-12 and IL-18. DETAILED DESCRIPTION

[0172] The advantages and features of the present disclosure will become apparent from the description which follows, in conjunction with the accompanying drawings. In the description, the specific conditions not mentioned in the examples are carried out according to the conventional conditions or the conditions recommended by the manufacturers. The reagents or instruments not mentioned by the manufacturers are all conventional products that can be purchased on the market.

[0173] The embodiments of the present disclosure are merely exemplary, and do not constitute any limitation on the scope of the present disclosure. Those skilled in the art should understand that the details and forms of the technical solutions of the present disclosure can be modified or replaced without departing from the spirit and scope of the present disclosure, and such modifications and replacements all fall within the protection scope of the present disclosure.

[0174] Preparation of TL1A antigen

[0175] The human TL1A protein extracellular region (UniProt sequence number: O95150, SEQ ID NO: 1), the cynomolgus monkey TL1A protein extracellular region (UniProt sequence number: G7PRK8, SEQ ID NO: 2), and the mouse TL1A protein extracellular region (UniProt sequence number: Q5UBV8, SEQ ID NO: 3) are used as templates for TL1A in the embodiments of the present disclosure (Table 1). The amino acid sequences of the antigens and detection proteins involved in the present disclosure are designed, and optionally, different tags (such as his tags) are fused to the TL1A protein. The sequences are cloned into the PTT5 vector (Invitrogen) for expression in 293 cells or stable expression and purification in CHO cells to obtain the antigens and detection proteins of the present disclosure.

[0176] Table 1 TL1A template sequences

[0177] Construction of CHO-K1 engineering cell strain

[0178] The nucleotide sequences corresponding to the human TL1A full-length amino acid sequence (UniProt sequence number: O95150, SEQ ID NO: 4) and the human DR3 full-length amino acid sequence (UniProt sequence number: Q93038, SEQ ID NO: 5) (Table 2) are cloned into the pcDNA3.1 vector (purchased from Clontech) to prepare plasmids. After transfecting the CHO-K1 cell line (purchased from the Chinese Academy of Sciences) with the plasmids (3000 Transfection Kit, purchased from Invitrogen, item number: L3000-015), the cells are cultured in a DMEM / F12 medium containing 10% (w / w) fetal bovine serum and 10 μg / ml puromycin for 2 weeks, and then the single clone cells are plated in a 96-well plate and cultured at 37°C and 5% (v / v) CO2. After about 2 weeks, some of the single clone wells are selected for amplification. The amplified clones are screened by flow cytometry. The cell lines with good growth, high fluorescence intensity, and single clones are selected for further culture and cryopreservation in liquid nitrogen. 3000 Transfection Kit, purchased from Invitrogen, item number: L3000-015) and cultured in a DMEM / F12 medium containing 10% (w / w) fetal bovine serum and 10 μg / ml puromycin for 2 weeks, and then the single clone cells are plated in a 96-well plate and cultured at 37°C and 5% (v / v) CO2. After about 2 weeks, some of the single clone wells are selected for amplification. The amplified clones are screened by flow cytometry. The cell lines with good growth, high fluorescence intensity, and single clones are selected for further culture and cryopreservation in liquid nitrogen.

[0179] Table 2 Human TL1A and human DR3 full-length amino acid sequences

[0180] Example 3 Construction of TF1-NFκB luciferase reporter cell line

[0181] The NF-kB re-nanoLuc plasmid (purchased from Promega) was transfected into the TF-1 cell line (Kocia, Catalog #CBP60808) using the 3000 Transfection Kit (purchased from Invitrogen, Catalog #L3000-015) and the transfected cells were selected in 1640 medium containing 10% (w / w) fetal bovine serum with 1 μg / ml puromycin for 2 weeks. The single cell clones were then plated into 96-well plates and cultured at 37°C, 5% (v / v) CO2. After about 2 weeks, some of the single clone wells were selected for expansion. The expanded clones were screened by flow cytometry. The cell lines with good growth, high reporter signal and single clone were selected for further expansion and cryopreservation in liquid nitrogen.

[0182] Example 4 Preparation of control antibodies and detection antibodies

[0183] The sequences of control antibodies RVT3101 (Publication No. CN113150144A), PRA023 (Publication No. CN114901311A / WO2022178159A1) and Teva-C320-168 (Publication No. US20140255302A1) were used as templates and constructed into the PPT5-hIgG1_LALAGA vector (provided by General Biotech (Anhui) Co., Ltd.) for expression in Expi293F cells or stable expression in CHO cells for purification to obtain positive control antibodies (Table 3).

[0184] Table 3 Control antibody sequences

[0185] Example 5 Generation of anti-human TL1A murine monoclonal antibodies

[0186] Anti-human TL1A antibodies were obtained by hybridoma technology. Mice were immunized with human TL1A-his fusion protein, and the spleen cells of the immunized mice were isolated. The spleen cells were fused with mouse myeloma cells by electrofusion, and cultured in HAT selection medium. The culture supernatant was taken for identification, and the clones secreting the target antibodies were subcloned. Finally, the murine monoclonal antibodies were obtained by production and purification.

[0187] The detailed description is as follows:

[0188] 5.1 Animal immunization

[0189] ​SJL mice, female, 6-8 weeks old (Shanghai Slac Animal Laboratory Co., Ltd., Animal Production License No: SCXK (Shanghai) 2017-0005) were used for experiments. The feeding environment: SPF level. After the mice were purchased, they were adaptively fed in the laboratory environment for 1 week in the experimental animal room (Shanghai Genechem Co., Ltd.), 12 / 12 hour light / dark cycle regulation, temperature 20-25°C; humidity 40-60%. The mice were immunized with TL1A antigen (prepared based on Example 1) in a conventional manner in the art.

[0190] 5.2 B cell fusion

[0191] The spleen and lymph node cells were fused with myeloma cells SP2 / 0 cells (ATCC® CRL-1581) to obtain hybridoma cells using an optimized electrofusion (BTX ECM2001+) procedure.

[0192] 5.3 Screening of hybridoma cells

[0193] After 7-10 days of fusion, the supernatant of the hybridoma cells was taken and the binding activity with human TL1A-his (prepared based on Example 1) was detected by ELISA method, and positive clones were selected; the next day, the binding activity of the supernatant of the positive clones with cynomolgus monkey TL1A protein, mouse TL1A protein (prepared based on Example 1), CHO-K1 human TL1A (prepared based on Example 2), the effect of blocking the binding of human TL1A protein and CHO-K1 human DR3 (prepared based on Example 2) were detected, and target clones were subcloned in semi-solid medium (Stemcell, item #03810). After 7-10 days of culture of the subcloned cells, the clones that grew were picked into 96-well culture plates one by one, and were cultured in DMEM medium containing 10% (w / w) fetal bovine serum, 1xHT (Sigma, item #H0137-10VL) to expand the culture, and target hybridoma monoclonal cells were selected by ELISA method, and were expanded to 24-well culture, 2-3 days later, the binding activity of the culture supernatant with human TL1A-his protein, mouse TL1A protein, cynomolgus monkey TL1A protein, the binding activity with CHO-K1 human TL1A (prepared based on Example 2), the effect of blocking the interaction of human TL1A protein and DcR3 protein, the effect of blocking the interaction of human TL1A protein and CHO-K1 human DR3 (prepared based on Example 2), and the inhibitory activity on the NFkB activity in TF-1-NFkB-luciferase cells (prepared based on Example 3) were detected. The target clones were selected for production and purification to obtain 230 monoclonal antibodies.

[0194] 5.4 Identification of murine monoclonal antibodies

[0195] ​The 230 monoclonal antibodies obtained above were further identified after purification, and 30 murine monoclonal candidate antibodies were obtained.

[0196] (a) Detection of the binding activity of anti-TL1A murine monoclonal antibodies to human TL1A-his protein, cynomolgus monkey TL1A-his protein and mouse TL1A-his protein.

[0197] The human TL1A-his protein / cynomolgus monkey TL1A-his protein or mouse TL1A-his protein prepared based on Example 1 was diluted with PBS to a final concentration of 2 μg / mL, and then added to a 96-well ELISA plate at 50 μL / well. Incubate overnight at 4°C with plastic film, and the next day wash the plate twice with PBST, add blocking solution [PBS + 5% (w / w) skim milk] and incubate at room temperature for 1 hour. Wash the plate 3 times with PBST, and add 50 μL / well of 100 nM starting 8-fold gradient diluted anti-TL1A murine monoclonal antibodies or control antibodies. After incubation at 37°C for 1 hour, wash the plate 3 times with PBST. Add HRP (horseradish peroxidase) labeled secondary antibody (Jackson Immuno, Cat. #115-035-003), and after incubation at 37°C for 1 hour, wash the plate 5 times with PBST. Add TMB substrate at 50 μL / well. After incubation at room temperature for 4 minutes, add stop solution (1.0 N HCl) at 50 μL / well. Read the OD450nm values with an ELISA plate reader (Multimode Plate Reader, EnSight, purchased from Perkin Elmer). Analyze the data by software (GraphPad Prism 9) for data fitting to calculate the EC50 value. The experimental results show that the anti-TL1A murine monoclonal antibodies can effectively bind to human TL1A-his protein and cynomolgus monkey TL1A-his protein, and some antibodies can bind to mouse TL1A-his protein.

[0198] (b) Detection of the binding activity of anti-TL1A murine monoclonal antibodies to CHO-K1 human TL1A high expression cell strain by FACS method

[0199] The CHO-K1-human TL1A cells (clone 1F2, human TL1A high expression cell strain) were expanded in a T-75 cell culture flask to 90% confluence, the culture medium was aspirated, and the cells were washed twice with PBS buffer, then the cells were trypsinized, and after stopping the digestion, the cells were washed twice with PBS buffer. After cell counting of the cells in the previous step, the cell pellet was resuspended to 2 x 10 6Cells / mL were injected at a rate of 50 μL / well into a 96-well plate. 50 μL / well of the initial 200 nM 4-fold serially diluted sample was added to each well, and the plate was incubated on ice for 1 hour. After washing three times with PBS buffer, 50 μL / well of Alexa Fluor 647-labeled secondary antibody (Jackson Immuno, catalog number #115-605-003) was added, and the plate was incubated on ice for 1 hour. After washing five times with PBS buffer, the results were detected and analyzed using flow cytometry (FACS Canto™, BD). Data analysis was performed using FlowJo software to obtain the mean fluorescence intensity (MFI) of the cells. Further analysis was performed using GraphPad Prism9 software to fit the data and calculate the EC50 value. The experimental results show that the purified murine anti-TL1A monoclonal antibody can effectively bind to the CHO-K1 human TL1A-overexpressing cell line.

[0200] (c) FACS method for detecting the interaction between human TL1A and CHO-K1 human DR3 cells blocked by anti-TL1A murine monoclonal antibody.

[0201] CHO-K1 human DR3 cells were cultured in T-75 cell culture flasks until 90% confluence. The culture medium was aspirated, and the cells were washed twice with PBS buffer. The cells were then digested with trypsin, and after digestion, washed twice with PBS buffer. Cell counting was performed, and the cells were diluted to 2 × 10⁻⁶ with blocking buffer. 6 Cells / mL, 50 μL / well added to a 96-well plate. The test sample, starting at 400 nM, was 3-fold diluted and mixed 1:1 with hTL1A-his (prepared based on Example 1) diluted to 0.4 μg / mL with PBS, and incubated at room temperature for 30 minutes. After incubation, 50 μL of the mixture was added to the plate and incubated on ice for 1 hour. The cells were washed three times with PBS buffer, and 50 μL / well of Alexa Fluor 647-labeled secondary antibody (GenScript, catalog number #A01802) was added, and incubated on ice for 1 hour. The cells were washed five times with PBS buffer, and the results were detected and analyzed using flow cytometry (FACS Canto™, purchased from BD). Data analysis was performed using software (FlowJo) to obtain the mean fluorescence intensity (MFI) of the cells. Further analysis was performed using software (GraphPad Prism9) to perform data fitting and calculate the IC50 value. Experimental results show that the purified murine monoclonal antibody can effectively block the binding of TL1A protein to CHO-K1-human DR3 cells.

[0202] (d) ELISA method to detect the inhibition of human TL1A-DCR3 interaction by anti-TL1A murine monoclonal antibody.

[0203] Human DcR3 / TNFRSF6B, Fc Tag protein (Acro, Cat# TNB-H5255) was diluted with PBS to a final concentration of 2 pg / mL, then added to a 96-well ELISA plate at 50 pL / well. Incubate overnight at 4°C with plastic film, the next day wash the plate 2 times with PBST, add blocking solution [PBS + 5% (w / w) skim milk] to block for 1 hour at room temperature, wash the plate 3 times with PBST. Mix 400 nM starting 4-fold dilution of the antibody to be tested with 1 pg / mL biotinylated labeled hTL1A protein (prepared based on Example 1) at 1:1, incubate for 30 min. After incubation, add 50 pL of the mixture to the ELISA plate, incubate at 37°C for 1 hour, then wash the plate 3 times with PBST. Add HRP (horseradish peroxidase) labeled secondary antibody (Sigma, Cat# S2438), incubate at 37°C for 1 hour, then wash the plate 5 times with PBST. Add TMB substrate at 50 pL / well. Incubate at room temperature for 4 minutes, then add stop solution (1.0 N HC1) at 50 pL / well. Read the OD450nm value with an ELISA plate reader (Multimode Plate Reader, EnSight, purchased from Perkin Elmer). Then analyze and fit the data by software (GraphPad Prism 9) to calculate the IC50value. The experimental results show that most of the purified murine monoclonal antibodies have strong inhibitory effect on the binding of TL1A and DcR3.

[0204] (e) Detecting the inhibition of NFkB activity in TF-1-NFkB-luciferase cells by anti-TL1A murine monoclonal antibodies.

[0205] The TF-1-NFkB-luciferase cells (prepared based on Example 3) were expanded in T-75 cell culture flasks to the logarithmic growth phase, and the culture medium supernatant was directly centrifuged and discarded, and the cell pellet was washed 2 times with PBS. The cells were adjusted to a density of 8 x 105cells / mL with [1640 + 2% (w / w) FBS] medium, and 100 pL of the cell suspension was added to each well of a 96-well plate. Incubate at 37°C, 5% CO2for 24 hours, then discard the culture medium supernatant and wash the cells twice with PBS. The cells were adjusted to a density of 8 x 105cells / mL with [1640 + 2% (w / w) FBS] medium, and 100 pL of the cell suspension was added to each well of a 96-well plate. Incubate at 37°C, 5% CO2for 24 hours, then discard the culture medium supernatant and wash the cells twice with PBS. 5The total volume of the test system used in this example was 100 μL. Human TL1A protein (prepared based on Example 1) was prepared in a detection medium (RPMI1640 + 10% FBS + 1x Penicillin / Streptomycin) to a final concentration of 100 ng / mL, and 20 μL / well was added to a 96-well flat-bottom white plate (Corning, item #3917). The TL1A antibody to be tested was diluted with the detection medium to a maximum final concentration of 32 nM, 2-fold dilution, 8 concentration gradients, and 20 μL / well was added to the 96-well flat-bottom white plate to mix with the TL1A protein, and incubated at room temperature for 30 minutes. Logarithmic growth phase TF-1 cells were collected and resuspended in the detection medium, and CHX (MCE, item #HY-12320) was added to a final concentration of 10 μg / mL. 20,000 cells / well of TF-1 cells were seeded into the above 96-well flat-bottom white plate, and mixed evenly. The cell plate was placed in a 37°C, 5% CO2 incubator for 24 hours. After 24 hours, the cell plate was removed from the incubator and equilibrated to room temperature, and 100 μL / well of equilibrated cell viability detection reagent Cell Counting-Lite 2.0 (Nanjing Nvigan Biotech Co., Ltd., item #DD1101-03) was added, mixed on a shaker for 5 minutes, and incubated at room temperature for 10 minutes. The fluorescence signal was detected using an enzyme-labeled instrument (PerkinElmer, Envision). The results showed that the purified mouse monoclonal antibodies could inhibit the apoptosis activity of TL1A-induced TF-1 cells.

[0206] (f) Detection of the apoptosis activity of anti-TL1A mouse monoclonal antibodies on TF-1 cells.

[0207] The total volume of the test system used in this example was 100 μL. Human TL1A protein (prepared based on Example 1) was prepared in a detection medium (RPMI1640 + 10% FBS + 1x Penicillin / Streptomycin) to a final concentration of 100 ng / mL, and 20 μL / well was added to a 96-well flat-bottom white plate (Corning, item #3917). The TL1A antibody to be tested was diluted with the detection medium to a maximum final concentration of 32 nM, 2-fold dilution, 8 concentration gradients, and 20 μL / well was added to the 96-well flat-bottom white plate to mix with the TL1A protein, and incubated at room temperature for 30 minutes. Logarithmic growth phase TF-1 cells were collected and resuspended in the detection medium, and CHX (MCE, item #HY-12320) was added to a final concentration of 10 μg / mL. 20,000 cells / well of TF-1 cells were seeded into the above 96-well flat-bottom white plate, and mixed evenly. The cell plate was placed in a 37°C, 5% CO2 incubator for 24 hours. After 24 hours, the cell plate was removed from the incubator and equilibrated to room temperature, and 100 μL / well of equilibrated cell viability detection reagent Cell Counting-Lite 2.0 (Nanjing Nvigan Biotech Co., Ltd., item #DD1101-03) was added, mixed on a shaker for 5 minutes, and incubated at room temperature for 10 minutes. The fluorescence signal was detected using an enzyme-labeled instrument (PerkinElmer, Envision). The results showed that the purified mouse monoclonal antibodies could inhibit the apoptosis activity of TL1A-induced TF-1 cells.

[0208] The above detection results showed that the 30 TL1A mouse monoclonal antibodies exhibited good human TL1A binding ability and blocking activity.

[0209] Example 6 Identification of Anti-Human TLlA Chimeric Antibodies

[0210] The murine antibodies were used to identify 6 chimeric antibodies: Mab 001, Mab 002, Mab 003, Mab 004, Mab 005 and Mab 006. Table 4 shows the VH / VL sequences of the chimeric antibodies, and Tables 5 to 7 show the results of KABAT, IMGT and Chothia analysis of the CDRs of the chimeric antibodies, respectively.

[0211] Table 4 Chimeric Antibody Sequences

[0212] Table 5 KABAT Analysis of CDRs of Chimeric Antibodies

[0213] Table 6 IMGT Analysis of CDRs of Chimeric Antibodies

[0214] Table 7 Chothia Analysis of CDRs of Chimeric Antibodies

[0215] 6.1 Detection of binding activity of anti-TLlA chimeric antibodies to human TLlA-His protein

[0216] The specific method is described in Example 5.4(a). The chimeric antibodies were added to the 96-well plates coated with the protein at a gradient dilution, and after incubation, HRP enzyme-labeled Goat anti-human antibody (Jackson, Cat. #109-035-098) was added, followed by color development with TMB and OD450nm reading on a microplate reader. As shown in Table 8 and Figure 1, the results showed that the anti-TLlA chimeric antibodies could effectively bind to human TLlA-His protein, as compared with the negative control Isotype (i.e., an anti-FITC antibody used as a homologous control antibody).

[0217] 6.2 Detection of binding activity of anti-TLlA chimeric antibodies to cynomolgus TLlA-His protein

[0218] The specific method is described in Example 5.4(a). The chimeric antibodies were added to the 96-well plates coated with the protein at a gradient dilution, and after incubation, HRP enzyme-labeled Goat anti-human antibody (Jackson, Cat. #109-035-098) was added, followed by color development with TMB and OD450nm reading on a microplate reader. As shown in Table 8 and Figure 1, the results showed that the anti-TLlA chimeric antibodies could effectively bind to human TLlA-His protein, as compared with the negative control Isotype (i.e., an anti-FITC antibody used as a homologous control antibody).

[0219] 6.3 Detection of binding activity of anti-TLl A chimeric antibodies to mouse TLl A-His protein

[0220] The specific method is referred to Example 5.4(a). Gradient dilution of chimeric antibodies was added to the 96-well plate coated with protein, and after incubation, HRP enzyme labeled Goat anti human antibody (Jackson, item #109-035-098) was added, and TMB was used for color development, and OD450nm was read by the enzyme label instrument. As shown in Table 8 and Figure 3, the results showed that anti-TLl A chimeric antibodies Mab001, Mab004, Mab005 and Mab006 could effectively bind to mouse TLl A-His protein, while chimeric antibodies Mab002 and Mab003 could not bind to mouse TLl A-his protein.

[0221] 6.4 Detection of binding activity of anti-TLl A chimeric antibodies to CHO-K1-human TLl A cell strain by FACS method

[0222] The specific method is referred to Example 5.4(b). The collected cells were mixed with chimeric antibodies in the same volume, and after incubation, Alexa 647fluorescein-labeled Goat anti human secondary antibody (1:800) (Jackson, item #109-605-088) was added, and the cell sample was analyzed on a flow cytometer (BD, Canto II). As shown in Table 8 and Figure 4, the results showed that all chimeric antibodies could effectively bind to CHO-K1-human TLl A cell strain compared with the negative control Isotype.

[0223] 6.5 Detection of anti-TLl A chimeric antibodies inhibiting the interaction of human TLl A with CHO-K1 human DR3 by FACS method

[0224] The specific method is referred to Example 5.4(c). As shown in Table 8 and Figure 5, the results showed that all chimeric antibodies could effectively block the binding of TLl A protein to CHO-K1-human DR3 cells compared with the negative control Isotype.

[0225] 6.6 Detection of anti-TLl A chimeric antibodies inhibiting the interaction of human TLl A with DcR3 by ELISA method

[0226] The specific method is referred to Example 5.4(d). As shown in Table 8 and Figure 6, the results showed that all chimeric antibodies could block the interaction of human TLl A with DcR3.

[0227] 6.7 Detection of anti-TLl A chimeric antibodies inhibiting the activity of NFκB in TF-1-NFκB-luciferase cells

[0228] The procedure is described in Example 5.4(e). As shown in Table 8 and Figure 7, the results of the experiment indicated that all the chimeric antibodies effectively inhibited the NFKB activity in TF-1-NFKB-luciferase cells.

[0229] 6.8 Detection of the anti-TLl A chimeric antibodies' inhibition of the apoptotic activity of TLl A on TF-1 cells

[0230] The procedure is described in Example 5.4(f). As shown in Table 8 and Figure 8, the results of the experiment indicated that all the chimeric antibodies effectively inhibited the TLl A-induced apoptotic activity of TF-1 cells.

[0231] 6.9 Detection of the anti-TLl A chimeric antibodies' inhibition of the secretion of IFN-γ from human PBMC stimulated by TLl A, IL-12 and IL-18

[0232] Isolation of CD4+T cells from peripheral blood mononuclear cells (PBMC) of healthy human donors + T cells (Stemcell, Cat# 17952) were adjusted to a cell density of 1 million / mL, and the CD4+T cells were transferred to 96-well U-bottom cell culture plates, corresponding to 100 μL per well. Appropriate amounts of recombinant human interleukin 18 (rhIL-18, R&D, Cat# 9124-IL-050, working concentration 50 ng / mL) and recombinant human interleukin 12 (rhIL-4, Peprotech, Cat# 200-12, working concentration 2 ng / mL) and recombinant human TLl A (rhTLl A, prepared based on Example 1, working concentration 50 ng / mL) were prepared, and 50 μL of the rhIL-18 / rhIL-4 / rhTLl A cytokine mixture was added to each well to induce the activation of CD4+T cells. The test TLl A antibodies or isotype control antibodies were prepared at working concentrations of 10, 5, 2.5, 1.25, 0.625, 0.3125, 0.15625, 0.078125 nM, and 50 μL of the corresponding TLl A antibody was added to each well. After mixing, the cells were incubated in a 37°C, 5% CO2incubator for 2 days, after which the cell culture supernatant was collected, and the content of IFN-γ in the supernatant was detected using a Human IFN-γ Precoated ELISA Kit (Duo, Cat# 1110003) according to the instructions of the reagent. Figure 9 shows the results of the experiment, which indicated that the chimeric antibodies effectively inhibited the secretion of IFN-γ.

[0233] Table 8 Identification of TLl A chimeric antibodies

[0234] 6.10 BIAcore detection of the affinity of anti-TLl A chimeric antibodies for human, cynomolgus monkey and mouse TLl A proteins

[0235] Specific binding between the tested anti-TL1A chimeric antibodies and human, cynomolgus and mouse TL1A proteins was detected by BIAcore 8K. The experiment used a Protein A chip (purchased from cytiva, item # 29-1275-56) to capture antibodies, and gradient dilutions of human TL1A-His, cynomolgus TL1A-His or mouse TL1A-His proteins prepared in Example 1 were used to measure the affinity of the antibodies to the antigens using multi-cycle kinetics. In each cycle, after capturing the antibodies, gradient concentrations of TL1A proteins were injected to record the binding and dissociation process of the antigens and antibodies. After each cycle, the Protein A chip was regenerated (to remove the proteins on the chip) with Glycine pH 1.5 (purchased from cytiva, item # BR-1003-54). Finally, using a 1:1 binding model, the data was analyzed to fit the antibody-antigen binding kinetic parameters, including the association rate constant ka, the dissociation rate constant kd, the equilibrium dissociation constant KD, and the maximum binding signal Rmax. The results in Table 9 show that all the tested anti-TL1A chimeric antibodies have specific binding to human and cynomolgus TL1A proteins and have high affinity levels, and Mab001, Mab004, Mab005 and Mab006 also have strong affinity to mouse TL1A protein.

[0236] Table 9 Affinity of anti-TL1A chimeric antibodies to human, cynomolgus and mouse TL1A proteins

[0237] Example 7 Humanization of anti-human TL1A monoclonal antibodies

[0238] By comparing the IMGT (http: / / imgt.cines.fr) human antibody heavy and light chain variable region gene database, the heavy and light chain variable region genes with high homology to the murine antibodies were selected as templates, and the CDRs of the murine antibodies were transplanted into the corresponding human templates to form variable region sequences in the order of FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. According to the needs, the key amino acids in the framework sequence were back-mutated to the corresponding amino acids of the murine antibodies to ensure the original affinity, i.e. to obtain humanized anti-TL1A monoclonal antibodies, wherein the CDR amino acid residues of the antibodies are determined and annotated by the Kabat numbering system.

[0239] 7.1 Humanization of Mab001

[0240] The humanized light chain template of the mouse antibody Mab001 is IGKV4-1*01 / IGKV2D-29*01 and IGKJ2*01, and the humanized heavy chain template is IGHV1-3*01 and IGHJ6*01. The CDRs of the mouse antibody Mab001 are transplanted into the humanized templates, respectively, to obtain the corresponding humanized version. According to the needs, the key amino acids in the FR region sequence of the humanized antibody of Mab001 are back-mutated to the corresponding amino acids of the mouse antibody to ensure the original affinity. The specific back-mutation design is shown in Table 10.

[0241] Table 10 Back-mutation design of humanized antibody of Mab001 Note: Graft represents the transplantation of CDRs of the mouse antibody into the FR region sequence of the human template; I2T represents the mutation of I at the 2nd position of Graft to T, and the others are the same. The numbering of the back-mutation amino acids is the natural order.

[0242] Table 11 CDR sequences of humanized antibody of Mab001 (Kabat numbering)

[0243] The specific sequences of the variable regions of the Hu001 humanized antibody are as follows:

[0244] The Hu001.VL1 amino acid sequence is shown as SEQ ID NO: 117:

[0245] The Hu001.VL2 amino acid sequence is shown as SEQ ID NO: 118:

[0246] The Hu001.VL3 amino acid sequence is shown as SEQ ID NO: 119:

[0247] The Hu001.VH1 amino acid sequence is shown as SEQ ID NO: 120:

[0248] The Hu001.VH2 amino acid sequence is shown as SEQ ID NO: 121:

[0249] The Hu001.VH3 amino acid sequence is shown as SEQ ID NO: 122:

[0250] The Hu001.VH4 amino acid sequence is shown as SEQ ID NO: 123:

[0251] The amino acid sequence of the humanized light chain template IGKV4-1*01 is shown as SEQ ID NO: 124:

[0252] The amino acid sequence of the humanized light chain template IGKV2D-29*01 is shown as SEQ ID NO: 125:

[0253] The amino acid sequence of the humanized light chain template IGKJ2*01 is shown as SEQ ID NO: 126:

[0254] The amino acid sequence of the humanized heavy chain template IGHV1-3*01 is shown as SEQ ID NO: 127:

[0255] The amino acid sequence of the humanized heavy chain template IGHJ6*01 is shown as SEQ ID NO: 128:

[0256] 7.2 Humanization of Mab004

[0257] The humanized light chain templates of murine antibody Mab004 are IGKV1-33*01 / IGKV3-15*01 and IGKJ2*01, and the humanized heavy chain template is IGHV1-69*02 and IGHJ6*01. The CDRs of murine antibody Mab004 are grafted into the humanized templates, respectively, to obtain the corresponding humanized version. According to the need, the key amino acids in the FR region sequence of the humanized antibody of Mab004 are back-mutated to the corresponding amino acids of the murine antibody to ensure the original affinity. The specific back-mutation design is shown in Table 12.

[0258] Table 12 Back-mutation design of humanized antibody of Mab004 Note: Graft represents the grafting of CDRs of murine antibody into the FR region sequence of human germline template; I2V means mutating I at position 2 of Graft to V, and the others are in the same manner. The numbering of back-mutation amino acids is the natural order.

[0259] Table 13 CDR sequences of humanized antibody of Mab004 (Kabat numbering)

[0260] The specific sequences of the variable regions of the Hu004 humanized antibody are as follows:

[0261] The amino acid sequence of Hu004.VL1 is shown as SEQ ID NO: 129:

[0262] The amino acid sequence of Hu004.VL2 is shown in SEQ ID NO:130:

[0263] The amino acid sequence of Hu004.VL3 is shown in SEQ ID NO:131:

[0264] The amino acid sequence of Hu004.VL4 is shown in SEQ ID NO:132:

[0265] The amino acid sequence of Hu004.VH1 is shown in SEQ ID NO:133:

[0266] The amino acid sequence of Hu004.VH2 is shown in SEQ ID NO:134:

[0267] The amino acid sequence of Hu004.VH3 is shown in SEQ ID NO:135:

[0268] The amino acid sequence of Hu004.VH4 is shown in SEQ ID NO:136:

[0269] The amino acid sequence of the humanized light chain template IGKV1-33*01 is shown in SEQ ID NO:137:

[0270] The amino acid sequence of the humanized light chain template IGKV3-15*01 is shown in SEQ ID NO:138:

[0271] The amino acid sequence of the humanized light chain template IGKJ2*01 is shown in SEQ ID NO:126:

[0272] The amino acid sequence of the humanized heavy chain template IGHV1-69*02 is shown in SEQ ID NO:139:

[0273] The amino acid sequence of the humanized heavy chain template IGHJ6*01 is shown in SEQ ID NO:128:

[0274] 7.3 Humanization of Mab005

[0275] The humanized light chain template of the mouse antibody Mab005 is IGKV2-29*02 / IGKV4-1*01 and IGKJ4*01, and the humanized heavy chain template is IGHV2-26*01 and IGHJ1*01. The CDRs of the mouse antibody Mab005 are transplanted into the humanized templates, respectively, to obtain the corresponding humanized version. According to the needs, the key amino acids in the FR region sequence of the humanized antibody of Mab005 are back-mutated to the corresponding amino acids of the mouse antibody to ensure the original affinity, and the sites prone to chemical modification are point-mutated to eliminate the risk of modification. The specific back-mutation design is shown in Table 14.

[0276] Table 14 Back-mutation design of humanized antibody of Mab005 Note: Graft represents the transplantation of CDRs of the mouse antibody into the FR region sequence of the human template; I2V represents the mutation of I at position 2 of Graft to V, and the others are similar. The numbering of the back-mutation amino acids is the natural order.

[0277] Table 15 CDR sequences of humanized antibody of Mab005 (Kabat numbering)

[0278] The specific sequences of the variable regions of the humanized antibody Hu005 are as follows:

[0279] The amino acid sequence of Hu005.VL1 is shown as SEQ ID NO: 141:

[0280] The amino acid sequence of Hu005.VL1a is shown as SEQ ID NO: 142:

[0281] The amino acid sequence of Hu005.VL2 is shown as SEQ ID NO: 143:

[0282] The amino acid sequence of Hu005.VH1 is shown as SEQ ID NO: 144:

[0283] The amino acid sequence of Hu005.VH2 is shown as SEQ ID NO: 145:

[0284] The amino acid sequence of Hu005.VH3 is shown as SEQ ID NO: 146:

[0285] The amino acid sequence of Hu005.VH4 is shown as SEQ ID NO: 147:

[0286] The VH5 amino acid sequence of Hu005 is shown as SEQ ID NO: 148:

[0287] The VH6 amino acid sequence of Hu005 is shown as SEQ ID NO: 149:

[0288] The humanized light chain template IGKV2-29*02 amino acid sequence is shown as SEQ ID NO: 150:

[0289] The humanized light chain template IGKV4-1*01 amino acid sequence is shown as SEQ ID NO: 124:

[0290] The humanized light chain template IGKJ4*01 amino acid sequence is shown as SEQ ID NO: 151:

[0291] The humanized heavy chain template IGHV2-26*01 amino acid sequence is shown as SEQ ID NO: 152:

[0292] The humanized heavy chain template IGHJ1*01 amino acid sequence is shown as SEQ ID NO: 153:

[0293] 7.4 Humanization of Mab006

[0294] The humanized light chain templates of murine antibody Mab006 are IGKV3-11*01 / IGKV6-21*01 and IGKJ2*01, and the humanized heavy chain templates are IGKV3-11*01 / IGKV6-21*01 and IGKJ2*01. The CDRs of the murine antibody Mab006 are grafted into the humanized templates, respectively, to obtain the corresponding humanized versions. According to the need, the key amino acids in the FR region sequence of the humanized antibody of Mab006 are back-mutated to the corresponding amino acids of the murine antibody to ensure the original affinity, and the sites prone to chemical modification are point-mutated to eliminate the risk of modification. The specific back-mutation design is shown in Table 16.

[0295] Table 16 Back-mutation design of humanized antibody of Mab006 Note: Graft represents the grafting of CDRs of the murine antibody into the FR region sequence of the human germline template; K50Y means mutating K at position 50 of Graft to Y, and the others are similar. The numbering of the back-mutation amino acids is the natural order.

[0296] Table 17 Mab006 humanized antibody CDR sequences (Kabat numbering)

[0297] The specific sequences of the variable regions of the Hu006 humanized antibodies are as follows:

[0298] The Hu006.VL1 amino acid sequence is set forth in SEQ ID NO: 155:

[0299] The Hu006.VL2 amino acid sequence is set forth in SEQ ID NO: 156:

[0300] The Hu006.VL3 amino acid sequence is set forth in SEQ ID NO: 157:

[0301] The Hu006.VH1 amino acid sequence is set forth in SEQ ID NO: 158:

[0302] The Hu006.VH2 amino acid sequence is set forth in SEQ ID NO: 159:

[0303] The Hu006.VH2a amino acid sequence is set forth in SEQ ID NO: 160:

[0304] The Hu006.VH3 amino acid sequence is set forth in SEQ ID NO: 161:

[0305] The Hu006.VH4 amino acid sequence is set forth in SEQ ID NO: 162:

[0306] The humanized light chain template IGKV3-11*01 amino acid sequence is set forth in SEQ ID NO: 163:

[0307] The humanized light chain template IGKV6-21*01 amino acid sequence is set forth in SEQ ID NO: 164:

[0308] The humanized light chain template IGKJ2*01 amino acid sequence is set forth in SEQ ID NO: 126:

[0309] The humanized heavy chain template IGHV1-3*01 amino acid sequence is set forth in SEQ ID NO: 127:

[0310] The humanized heavy chain template IGHJ6*01 amino acid sequence is shown in SEQ ID NO: 128:

[0311] Example 8 Identification of anti-TL1A humanized antibodies

[0312] After expression, protein binding and blocking identification of all humanized molecules of the antibodies, part of the humanized molecules of each molecule were selected for further identification.

[0313] 8.1 Detection of binding activity of anti-TL1A humanized antibodies to human TL1A-His protein

[0314] The specific method is referred to Example 5.4(a). Gradient diluted humanized antibodies were added to the 96-well plate coated with the protein, and after the incubation was completed, HRP enzyme-labeled Goat anti-human antibody (Jackson, item #109-035-098) was added, and TMB was used for color development, and OD450nm was read by the enzyme-labeled instrument. The results are shown in Table 18 and Figure 10, and all anti-TL1A humanized antibodies can effectively bind to human TL1A-His protein.

[0315] 8.2 Detection of binding activity of anti-TL1A humanized antibodies to cynomolgus TL1A-His protein

[0316] The specific method is referred to Example 5.4(a). Gradient diluted humanized antibodies were added to the 96-well plate coated with the protein, and after the incubation was completed, HRP enzyme-labeled Goat anti-human antibody (Jackson, item #109-035-098) was added, and TMB was used for color development, and OD450nm was read by the enzyme-labeled instrument. The results are shown in Table 18 and Figure 11, and all anti-TL1A humanized antibodies can effectively bind to cynomolgus TL1A-His protein.

[0317] 8.3 Detection of binding activity of anti-TL1A humanized antibodies to mouse TL1A-His protein

[0318] The specific method is referred to Example 5.4(a). Gradient diluted humanized antibodies were added to the 96-well plate coated with the protein, and after the incubation was completed, HRP enzyme-labeled Goat anti-human antibody (Jackson, item #109-035-098) was added, and TMB was used for color development, and OD450nm was read by the enzyme-labeled instrument. The results are shown in Table 18 and Figure 12, and the binding activity of anti-TL1A antibody Hu005 humanized molecules to mouse TL1A-his protein decreased slightly, and other anti-TL1A humanized antibodies can effectively bind to mouse TL1A-His protein.

[0319] 8.4 FACS method to detect the binding activity of anti-TLl A humanized antibodies to CHO-K1 -human TLl A high expression cell line. Refer to Example 5.4(b) for the detailed method. The collected cells were mixed with the humanized antibodies in equal volume, after incubation, Alexa 647fluorescein-labeled Goat anti-human secondary antibody (1 :800) (Jackson, Cat# 109-605-088) was added, and the cell samples were analyzed on a flow cytometer (BD, Canto II). As shown in Table 18 and Figure 13, the results indicated that all the humanized antibodies could effectively bind to CHO-K1 -human TLl A high expression cell line.

[0320] 8.5 FACS method to detect the inhibition of the interaction of human TLl A with human TLl A and CHO-K1 human DR3 by anti-TLl A humanized antibodies

[0321] Refer to Example 5.4(c) for the detailed method. As shown in Table 18 and Figure 14, the experimental results indicated that the humanized antibodies effectively blocked the binding of TLl A protein to CHO-K1 -human DR3 cells.

[0322] 8.6 ELISA method to detect the inhibition of the interaction of human TLl A with DcR3 by anti-TLl A humanized antibodies

[0323] Refer to Example 5.4(d) for the detailed method. As shown in Table 18 and Figure 15, the experimental results indicated that the humanized antibodies could all block the binding of TLl A protein to DcR3 protein.

[0324] 8.7 Detection of the inhibition of NFκB activity in TF-1-NFκB-luciferase cells by anti-TLl A humanized antibodies.

[0325] Refer to Example 5.4(e) for the detailed method. As shown in Table 18 and Figure 16, the experimental results indicated that the humanized antibodies could all effectively inhibit the NFκB activity in TF-1-NFκB-luciferase cells.

[0326] 8.8 Detection of the inhibition of apoptosis activity of TF-1 cells by anti-TLl A humanized antibodies.

[0327] Refer to Example 5.4(f) for the detailed method. As shown in Table 18 and Figure 17, except that the Hu005-L1aH1 and Hu005-L1aH2 humanized antibodies were slightly weaker than the chimeric antibody in inhibiting the apoptosis activity of TLl A-induced TF-1 cells, the other humanized antibodies could all effectively inhibit the apoptosis activity of TLl A-induced TF-1 cells.

[0328] Table 18 Identification of anti-TLl A humanized antibodies

[0329] 8.9 Detection of inhibition of IFN-g secretion by anti-TLl A humanized antibodies on human TLl A and IL-12 and IL-18 stimulated PBMC.

[0330] The specific method is referred to Example 6.9. As shown in Figure 18, all humanized antibodies can effectively inhibit IFN-g secretion by human TLl A and IL-12 and IL-18 stimulated PBMC.

[0331] 8.10 BIAcore detection of affinity of anti-TLl A humanized antibodies to human, cynomolgus monkey and mouse TLl A proteins

[0332] The specific method is referred to Example 6.10. The results in Table 19 show that specific binding and high affinity levels exist between all tested anti-TLl A humanized antibodies and human, cynomolgus monkey or mouse TLl A proteins.

[0333] Table 19 Affinity of anti-TLl A humanized antibodies to human, cynomolgus monkey and mouse TLl A proteins

[0334] The teachings of all patents, published applications and references cited herein are incorporated by reference in their entirety.

[0335] While example embodiments have been particularly shown and described, a person of ordinary skill in the art will understand that various changes in form and detail can be made thereto without departing from the scope of the embodiments encompassed by the appended claims.

Claims

1. An antibody or antigen binding fragment that specifically binds to TL1A, characterized in that, The antibody or antigen-binding fragment comprises a heavy chain variable region VH comprising HCDR1-HCDR3, and / or a light chain variable region VL comprising LCDR1-LCDR3, The HCDR1-HCDR3 has the amino acid sequence of HCDR1, HCDR2 and HCDR3 of VH as set forth in any one of SEQ ID NOs: 12, 14, 16, 18, 20, 22, 120, 121, 122, 123, 133, 134, 135, 136, 144, 145, 146, 147, 148, 149, 158, 159, 160, 161 and 162; and / or; The LCDR1-LCDR3 has the amino acid sequence of LCDR1, LCDR2 and LCDR3 of VL as set forth in any one of SEQ ID NOs: 13, 15, 17, 19, 21, 23, 117, 118, 119, 129, 130, 131, 132, 141, 142, 143, 155, 156 and 157.

2. The antibody or antigen-binding fragment of claim 1, wherein, The antibody or antigen-binding fragment comprises: (1) the amino acid sequence of HCDR1-HCDR3 of VH as set forth in SEQ ID NO: 12, 120, 121, 122 or 123, and the amino acid sequence of LCDR1-LCDR3 of VL as set forth in SEQ ID NO: 13, 117, 118 or 119; or, (2) the amino acid sequence of HCDR1-HCDR3 of VH as set forth in SEQ ID NO: 14, and the amino acid sequence of LCDR1-LCDR3 of VL as set forth in SEQ ID NO: 15; or, (3) the amino acid sequence of HCDR1-HCDR3 of VH as set forth in SEQ ID NO: 16, and the amino acid sequence of LCDR1-LCDR3 of VL as set forth in SEQ ID NO: 17; or, (4) the amino acid sequence of HCDR1-HCDR3 of VH as set forth in SEQ ID NO: 18, 133, 134, 135 or 136, and the amino acid sequence of LCDR1-LCDR3 of VL as set forth in SEQ ID NO: 19, 129, 130, 131 or 132; or, (5) the amino acid sequence of HCDR1-HCDR3 of VH as set forth in SEQ ID NO: 20, 144, 145, 146, 147, 148 or 149, and the amino acid sequence of LCDR1-LCDR3 of VL as set forth in SEQ ID NO: 21, 141, 142 or 143; or, (6) the amino acid sequence of HCDR1-HCDR3 of VH as set forth in SEQ ID NO: 22, 158, 159, 160, 161 or 162, and the amino acid sequence of LCDR1-LCDR3 of VL as set forth in SEQ ID NO: 23, 155, 156 or 157; Preferably, the HCDR1-HCDR3 and / or the LCDR1-LCDR3 are determined according to the Kabat, IMGT or Chothia system.

3. The antibody or antigen binding fragment of claim 1 or 2, characterized in that, The antibody or antigen binding fragment comprises the heavy chain variable region VH and the light chain variable region VL, wherein, the HCDR1-HCDR3 have amino acid sequences selected from the group consisting of: (1) SEQ ID NOs: 39-41, 75-77, or 105-106 and 41; or, (2) SEQ ID NOs: 45-47, 80-82, or 107-108 and 47; or, (3) SEQ ID NOs: 51-53, 85-87, or 109-110 and 53; or, (4) SEQ ID NOs: 57-59, 90-92, or 111-112 and 59; or, (5) SEQ ID NOs: 63-65, 95-97, or 113-114 and 65; or, (6) SEQ ID NOs: 69-71, 100-102, 115-116 and 71, or SEQ ID NOs: 69, 154 and 71; and / or the LCDR1-LCDR3 have sequences selected from the group consisting of: (1) SEQ ID NOs: 42-44 or 78-79 and 44; or, (2) SEQ ID NOs: 48-50 or 83-84 and 50; or, (3) SEQ ID NOs: 54-56 or 88-89 and 56; or, (4) SEQ ID NOs: 60-62; or 93-94 and 62; or, (5) SEQ ID NOs: 66-68, 98-99 and 68, or SEQ ID NOs: 140 and 67-68; or, (6) SEQ ID NOs: 72-74 or 103-104 and 74; Preferably, (1) the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 39-41, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 42-44, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 75-77, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 78-79 and 44, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 105-106 and 41, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 42-44, respectively; (2) the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 45-47, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 48-50, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 80-82, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 83-84 and 50, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 107-108 and 47, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 48-50, respectively; (3) the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 51-53, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 54-56, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 85-87, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 88-89 and 56, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 109-110 and 53, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 54-56, respectively; (4) the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 57-59, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 60-62, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 90-92, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 93-94 and 62, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 111-112 and 59, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 60-62, respectively; (5) the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 63-65, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 66-68, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 95-97, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 98-99 and 68, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 113-114 and 65, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 66-68, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 63-65, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 140 and 67-68, respectively; or (6) the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 69-71, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 72-74, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 100-102, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 103-104 and 74, respectively; the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 115-116 and 71, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 72-74, respectively; or the HCDR1-HCDR3 have the amino acid sequences of SEQ ID NOs: 69, 154 and 71, respectively, and the LCDR1-LCDR3 have the amino acid sequences of SEQ ID NOs: 72-74, respectively.

4. The antibody or antigen binding fragment of any one of claims 1-3, wherein, the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 12, 14, 16, 18, 20, 22, 120, 121, 122, 123, 133, 134, 135, 136, 144, 145, 146, 147, 148, 149, 158, 159, 160, 161, and 162, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations, and / or, the light chain variable region VL comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 13, 15, 17, 19, 21, 23, 117, 118, 119, 129, 130, 131, 132, 141, 142, 143, 155, 156, and 157, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations, preferably, (1) the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 12, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 13; (2) the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 14, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 15; (3) the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 16, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 17; (4) the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 18, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 19; (5) the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 20, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 21; (6) the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 22, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 23; (7) the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 120-123, and the light chain variable region VL comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 117-119; (8) the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 133-136, and the light chain variable region VL comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 129-132; (9) the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 144-149, and the light chain variable region VL comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 141-143; (10) or the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 158-162, and the light chain variable region VL comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 155-157; Preferably, the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 120-123, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 119; or the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 133-136, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 129; or the heavy chain variable region VH comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 144-146, and the light chain variable region VL comprises an amino acid sequence as set forth in SEQ ID NO: 142; or the heavy chain variable region VH comprises an amino acid sequence as set forth in SEQ ID NO: 160, and the light chain variable region VL comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 155-157.

5. The antibody or antigen binding fragment of any one of claims 1-4, wherein, the antibody or antigen binding fragment further comprises a heavy chain constant region and / or a light chain constant region; optionally, the heavy chain constant region and / or the light chain constant region is selected from a complete constant region sequence or a fragment thereof, the constant region fragment including CH1, hinge region, CH2, CH3, CL and / or Fc region; Preferably, the heavy chain constant region is selected from a human or murine IgG1, IgG2, IgG3 or IgG4 constant region; and / or the light chain constant region is selected from a human or murine kappa constant region or lamda constant region; Preferably, the CH1 has an amino acid sequence as set forth in SEQ ID NO: 24, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations; Preferably, the CL has an amino acid sequence as set forth in SEQ ID NO: 25, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations; Preferably, the Fc region has an amino acid sequence as set forth in SEQ ID NO: 26, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations.

6. The antibody or antigen binding fragment of any one of claims 1-5, wherein, the antibody or antigen binding fragment comprises a heavy chain HC and a light chain LC, wherein, the heavy chain HC has an amino acid sequence as set forth in SEQ ID NO: 27, 29, 31, 33, 35 or 37, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations; and / or the light chain LC has an amino acid sequence as set forth in SEQ ID NO: 28, 30, 32, 34, 36 or 38, or an amino acid sequence having at least 70% identity thereto or at most 15 amino acid mutations.

7. The antibody or antigen binding fragment of any one of claims 1-6, wherein, The at least 70% identity is at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity; the at most 3 mutations are preferably at most 3, 2, 1, or 0 mutations; the at most 15 mutations are preferably at most 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, or 0 mutations; preferably, the mutations are selected from substitution, deletion, or insertion mutations; preferably, the mutations are back mutations or hot spot mutations; preferably, the substitutions are conservative amino acid substitutions.

8. The antibody or antigen binding fragment of any one of claims 1-7, wherein, The antibody or antigen binding fragment is selected from a monoclonal antibody, a polyclonal antibody, a natural antibody, an engineered antibody, a monospecific antibody, a multispecific antibody (e.g., a bispecific antibody), a monovalent antibody, a multivalent antibody, an intact antibody, a fragment of an intact antibody, a naked antibody, a conjugated antibody, a chimeric antibody, a humanized antibody, a fully human antibody, a Fab, a Fab', a Fab'-SH, a F(ab')2, a Fd, a Fv, a scFv, a diabody, or a single domain antibody.

9. The antibody or antigen binding fragment of any one of claims 1-8, wherein, The antibody or antigen binding fragment is capable of binding to human TL1A, monkey TL1A, and / or murine TL1A.

10. The antibody or antigen binding fragment of any one of claims 1-9, wherein, The antibody or antigen binding fragment further comprises a conjugate; preferably, the conjugate is selected from a therapeutic agent or a tracer; preferably, the therapeutic agent is selected from a radioisotope, a chemotherapeutic agent, or an immunomodulatory agent; preferably, the tracer is selected from a radiological contrast agent, a paramagnetic ion, a metal, a fluorescent label, a chemiluminescent label, an ultrasound contrast agent, and a photosensitizer.

11. The antibody or antigen binding fragment of any one of claims 1-10, wherein, The antibody or antigen binding fragment binds to human TL1A, monkey TL1A, and / or murine TL1A with a KD value of less than 1E-08 M, 1E-09 M, 1E-10 M, 1E-11 M, or 1E-12 M.

12. A multispecific antigen-binding molecule, characterized in that, The multispecific antigen binding molecule comprises the antibody or antigen binding fragment of any one of claims 1-11; and a further antigen binding molecule that binds to an antigen other than TL1A, or a further antigen binding molecule that binds to a different epitope of TL1A than the preceding antibody or antigen binding fragment; preferably, the further antigen binding molecule is an antibody or antigen binding fragment; Preferably, the further antigen is selected from the group consisting of: (1) a tumor specific antigen (TSA) or a tumor associated antigen (TAA); (2) an immune checkpoint; and (3) a target that recruits and / or activates an immune cell; Preferably, the multispecific antigen binding molecule is bispecific, trispecific, or tetraspecific; Preferably, the multispecific antigen binding molecule is bivalent, tetravalent, or hexavalent.

13. A chimeric antigen receptor (CAR), characterized in that, The chimeric antigen receptor comprises at least an extracellular antigen binding domain, a transmembrane domain, and an intracellular signaling domain, the extracellular antigen binding domain comprising the antibody or antigen binding fragment of any one of claims 1-11 or the multispecific antigen binding molecule of claim 11.

14. An immune effector cell, wherein, The immune effector cell expresses and / or comprises a nucleic acid molecule encoding the chimeric antigen receptor of claim 13. Preferably, the immune effector cell is an autologous immune effector cell or an allogeneic immune effector cell. Preferably, the immune effector cell is an autologous immune effector cell or an allogeneic immune effector cell.

15. An isolated nucleic acid molecule, comprising, The nucleic acid molecule encodes the antibody or antigen binding fragment of claims 1-11, the multispecific antigen binding molecule of claim 12, or the chimeric antigen receptor of claim 13.

16. A vector, characterized in that, The vector comprises the nucleic acid molecule of claim 15.

17. A cell, wherein, The cell comprises the nucleic acid molecule of claim 15 or the vector of claim 16.

18. A method of making the antibody or antigen binding fragment of claims 1-11, or the multispecific antigen binding molecule of claim 12, comprising, The method comprises: (1) culturing the cell of claim 17 and / or (2) isolating the antibody or antigen binding fragment, or the multispecific antigen binding molecule, expressed by the cell.

19. A method of making the immune effector cell of claim 14, comprising, The method comprises introducing into the immune effector cell a nucleic acid molecule encoding the chimeric antigen receptor of claim 13, and / or initiating expression of the chimeric antigen receptor by the immune effector cell.

20. A pharmaceutical composition comprising, The pharmaceutical composition comprises the antibody or antigen binding molecule of any one of claims 1-11, or the multispecific antigen binding molecule of claim 12, or the immune effector cell of claim 14, or the nucleic acid molecule of claim 15, or the vector of claim 16, or the cell of claim 17, or a product produced by the method of any one of claims 18-19; preferably, the composition further comprises a pharmaceutically acceptable carrier, diluent, or adjuvant.

21. Use of the antibody or antigen binding fragment of any one of claims 1-11, the multispecific antigen binding molecule of claim 12, the immune effector cell of claim 14, the nucleic acid molecule of claim 15, the vector of claim 16, the cell of claim 17, the pharmaceutical composition of claim 20, or a product produced by the method of any one of claims 18-19, in the manufacture of a medicament for treating an immune-mediated inflammatory disease (IMID); preferably, the immune-mediated inflammatory disease (IMID) is an autoimmune disease and / or an inflammatory disease; more preferably, the immune-mediated inflammatory disease (IMID) is selected from one or more of inflammatory bowel disease, multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes, vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), a neurodegenerative disease of aging, periodontal disease, preperfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatism, scleroderma, sinusitis, tuberculosis, arteriosclerosis, and uveitis.

22. A method of treating an immune-mediated inflammatory disease (IMID), comprising administering to a subject in need thereof a therapeutically effective amount of a compound of any one of claims 1-21, or a pharmaceutically acceptable salt thereof. The method comprises administering to the subject an effective amount of the antibody or antigen binding fragment of any one of claims 1-11, the multispecific antigen binding molecule of claim 12, the immune effector cell of claim 14, the nucleic acid molecule of claim 15, the vector of claim 16, the cell of claim 17, the pharmaceutical composition of claim 20, or a product produced by a process according to any one of claims 18-19, preferably, the immune-mediated inflammatory disease (IMID) is an autoimmune disease and / or an inflammatory disease; more preferably, the immune-mediated inflammatory disease (IMID) is selected from one or more of inflammatory bowel disease, multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes, vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), a neurodegenerative disease of aging, periodontal disease, pre-perfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatic disease, scleroderma, sinusitis, tuberculosis, arteriosclerosis, and uveitis.

23. The antibody or antigen binding fragment of any one of claims 1-11, the multispecific antigen binding molecule of claim 12, the immune effector cell of claim 14, the nucleic acid molecule of claim 15, the vector of claim 16, the cell of claim 17, the pharmaceutical composition of claim 20, or the product produced by a process according to any one of claims 18-19, characterized in that, The method comprises administering to the subject an effective amount of the antibody or antigen binding fragment of any one of claims 1-11, the multispecific antigen binding molecule of claim 12, the immune effector cell of claim 14, the nucleic acid molecule of claim 15, the vector of claim 16, the cell of claim 17, the pharmaceutical composition of claim 20, or a product produced by a process according to any one of claims 18-19, preferably, the immune-mediated inflammatory disease (IMID) is an autoimmune disease and / or an inflammatory disease; more preferably, the immune-mediated inflammatory disease (IMID) is selected from one or more of inflammatory bowel disease, multiple sclerosis, psoriasis, rheumatoid arthritis, psoriatic arthritis, systemic lupus erythematosus, diabetes, vasculitis, asthma, eczema, atopic dermatitis, fibrosis, transplant rejection, graft-versus-host disease, allergy, ankylosing spondylitis, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, glomerulonephritis, gout, hepatitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), a neurodegenerative disease of aging, periodontal disease, pre-perfusion injury transplant rejection, psoriasis, pulmonary fibrosis, rheumatic disease, scleroderma, sinusitis, tuberculosis, arteriosclerosis, and uveitis.

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