Antibody and use thereof

Abstract

The present application provides an isolated antigen-binding protein, comprising: a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain specifically binds to TNF-like protein A (TL1A), and the first antigen binding domain and the second antigen binding domain are different.

Description

[0001] Attorney Docket No.: 769273: SA9-943PC

[0002] ANTIBODY AND USE THEREOF

[0003] RELATED APPLICATIONS

[0004] This application claims priority to Chinese Patent Application Nos. 202510717116.5, filed May 30, 2025, 202510130736.9, filed February 5, 2025, 202510127573.9, filed January 27, 2025, 202411388891.2, filed September 30, 2024, and 202411195305.2, filed August 28, 2024, the entire disclosures of which are hereby incorporated herein by reference.

[0005] SEQUENCE LISTING

[0006] The instant application contains a Sequence Listing which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML file, created on August 27, 2025, is named 769273_SA9-943PC_ST26.xml and is 572,968 bytes in size.

[0007] FIELD OF THE INVENTION

[0008] The present application relates to the field of biomedicine, and specifically to a TL1 A antigen-binding protein.

[0009] BACKGROUND OF THE INVENTION

[0010] TL1 A is encoded by the TNFSF15 gene and is also known as vascular growth inhibitory factor (VEGI), a member of the tumor necrosis factor (TNF) ligand superfamily.TLl A is a type II transmembrane protein.TLl Ais initially expressed on the surface of the cell as a membranebound protein, and then is sheared by extracellular matrix metalloproteinases to form a soluble form of TLlA.lt has been shown that TL1 Ais mainly expressed in antigen-presenting cells and lymphocytes, as well as endothelial cells and fibroblasts stimulated by inflammatory factors. TL1A is mainly expressed in antigen-presenting cells and lymphocytes, as well as Attorney Docket No.: 769273: SA9-943PC endothelial cells and fibroblasts stimulated by inflammatory factors. TL1A induces the activation of downstream NF-KB and MAPK cascade (e.g., ERK, p38, JNK) signaling pathways by binding to and activating its receptor DR3, which regulates the cell proliferation, survival, or apoptosis of the cells. TL1A-DR3 signaling plays an important role in the regulation of the immune system. The TL1A-DR3 signaling pathway plays an important role in the regulation of the immune system, which is mainly reflected in the following aspects: 1) synergistically promoting Thl activation and cytokine secretion. Under the condition of TCR activation, TL1 A can induce the production and secretion of proinflammatory factors, mainly including: IFN-y, GM-CSF, TNF-cr, etc. It can also synergize with IL12 / IL18 factors to promote IFN-y production by T cells, and the effect is not dependent on the presence of antigen.2) Synergistically promoting Th2 activation, sustained overexpression of TLlAin T or DC cells in mice produces an intestinal inflammation that is dependent on the Th2 cytokine IL13.In addition, TL1A also promotes Th2 immune responses and allergen-specific antibody production in a mouse model of asthma.3) Synergistically promotes the proliferation and activation of Thl7-type cells, and inhibition of TLlA helps to alleviate the symptoms of DSS- induced intestinal inflammation in mice.4) TL1 A also promotes the activation and cell-killing capacity of NK and NKT cells.5) TL1A also promotes the activation and cell-killing capacity of NK and NKT cells.6) TL1 A also promotes the activation of NK and NKT cells.

[0011] Antibodies targeting inhibition of TLlA have entered clinical trials and achieved positive therapeutic effects in patients with inflammatory bowel disease, and still have some efficacy in patients with resistance to or relapse of frontline biologies, but as of now, all kinds of drugs targeting inhibition of TLI A are still in the clinical stage, and there are no drugs on the market. For example, the results of phase II clinical trial of PRA023 / MK7240 for Crohn's disease showed that 49% of patients achieved clinical remission and 26% of patients achieved endoscopic remission; while the results of phase II clinical trial of PRA023 / MK7240 for ulcerative colitis showed that 26.5% of patients achieved clinical remission and 36.8% of patients achieved endoscopic remission. Similar therapeutic effects were also seen in the Phase II clinical trial of RVT-3 I0I / RG663 I. Overall, monoclonal antibodies targeting TLI A Attorney Docket No.: 769273: SA9-943PC inhibition have shown positive therapeutic effects in re-inflammatory bowel disease, but only a fraction of patients have achieved clinical benefit, and the overall efficacy needs to be further improved.

[0012] Integrins are transmembrane glycoproteins expressed on the cell surface that have important functions in immune cell proliferation, signaling, and cell migration by mediating cell-cell interactions. Integrins are heterodimers consisting of an a-subunit and a P- subunit .Integrin a4p7, a heterodimer of a4 and P7 subunits, is mainly expressed in memory T cells and memory B cells, and is important for the localization and migration of immune cells to intestinal mucosal tissues. a4p7 mediates the migration of immune cells by interacting with its ligand, MAdCAM-1, which is mainly expressed in the vascular endothelial cells of the small intestine. Currently, vedolizumab, a monoclonal antibody targeting inhibition of a4p7, is approved and marketed for the treatment of moderate-to-severe ulcerative colitis and Crohn's disease. In ulcerative colitis, vedolizumab resulted in better clinical response and disease remission in 21% of patients after 6 weeks of treatment and in 42% of patients after extending treatment to 52 weeks. In contrast, in Crohn's disease, Vedolizumab treatment for 6 weeks resulted in a better response and disease remission in 15% of patients, and after extending treatment to 52 weeks, a better clinical response and disease remission was achieved in 39% of patients.

[0013] Over the past two decades, biomolecules represented by vedolizumab, a monoclonal antibody targeting integrin a4p7, have achieved remarkable efficacy in the treatment of ulcerative colitis, driving new breakthroughs in clinical remission rates in ulcerative colitis. However, most current treatments have 1-year clinical remission rates between 30% and 50%, and more than 50% of patients still have inadequate clinical response, suggesting that the therapeutic ceiling may have been reached through the use of a single agent. In addition, there is still a huge unmet clinical need for effective yet safe therapeutic agents for patients with moderate-to-severe disease, and newer drug development is urgently needed to improve treatment outcomes. Attorney Docket No.: 769273: SA9-943PC

[0014] Ulcerative colitis is the result of multiple pathways driving immune-mediated inflammatory processes, and drugs that are effective in intestinal disease may be ineffective in extraintestinal manifestations or concomitant immune-mediated inflammatory disease. With this in mind, the concept of combination therapy based on the use of two biologies may be a promising strategy for patients with refractory ulcerative colitis, patients at high risk of developing complications, or patients with concomitant uncontrolled immune-mediated inflammatory disease. This strategy is currently being applied in real-world treatment with good results.2022 A systematic review and meta-analysis of combination therapy published in 2022 (Ahmed, W., et al. Clin Gastroenterol Hepatol. 2022) showed that in 279 patients who received combination therapy in a total of 30 included studies. Clinical remission and endoscopic remission rates reached 59% and 34%, respectively. This included 81% of patients with refractory ulcerative colitis and 12% of patients with ulcerative colitis complicated by extraintestinal manifestations or rheumatologic disease. The most common combinations in the treatment regimen included anti-integrin monoclonal antibody + anti-TNF-a monoclonal antibody (48%) and anti-integrin monoclonal antibody + Ustekinumab (19%).

[0015] SUMMARY OF THE INVENTION

[0016] An isolated antigen-binding protein comprising: a first antigen-binding domain and a second antigen -binding domain, the first antigen -binding domain specifically binds to TNF- like protein A (TL1A), and the first antigen-binding domain is different from the second antigen-binding domain.

[0017] In some embodiments, wherein,

[0018] 1) in the presence of monomeric TL1 A and / or trimeric TL1 A, an effect of down-regulating the expression of cell membrane a4p7 is enhanced, preferably, said antigen-binding protein or protein comprising the first antigen-binding domain forms a crosslinked structure with the monomeric TL1 A and / or trimeric TL1 A; or,

[0019] 2) in the presence of a4p7, an effect of down-regulating the expression of monomeric

[0020] TL1A and / or trimeric TL1A is enhanced, preferably, said antigen-binding protein or protein Attorney Docket No.: 769273: SA9-943PC comprising the second antigen-binding domain forms a crosslinked structure with the a4p7 protein.

[0021] In some embodiments, the isolated antigen-binding protein mediates internalization of the cell membrane a4p7 or TL1 A.

[0022] In some embodiments, wherein,

[0023] 1) in the presence of monomeric TL1 A and / or trimeric TL1 A, an effect of mediating the occurrence of the internalization in the cell membrane a4p7 of the subject is enhanced, or;

[0024] 2) in the presence of a4p7, an effect of mediating the occurrence of the internalization in the cell membrane a4p7 of the subject is enhanced.

[0025] In some embodiments, the isolated antigen-binding protein specifically binds to the a4p7 protein with a KD value of approximately 5E-08M or less.

[0026] In some embodiments, the isolated antigen-binding protein specifically binds to the monomeric TLlA / or trimeric TL1A with a KD value of approximately 5E-08M or less.

[0027] In some embodiments, a binding site of the first antigen-binding domain of the antigenbinding protein to the TLlAis selected from one or more of the following: R32, Q33, P35, T36, Q37, P44, E49, E51, L54, A56, S89, K102, P103, Y117, M146, S148, D161, 1162, S163, L164 and Y167; and the binding sites are numbered according to an amino acid sequence of TL1A as set forth in SEQ ID NO: 327.

[0028] In some embodiments, a binding site of the first antigen-binding domain to the TL1 A is selected from one or more of the following: R32, Q33, P35, T36, Q37, P44, S89, K102, P103, D161, 1162, SI 63 and LI 64, including any one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve or thirteen of the above 13 sites; and the binding sites are numbered according to the amino acid sequence of TL1A as set forth in SEQ ID NO: 327.

[0029] In some embodiments, a binding site of the antigen-binding protein to the TLlAis selected from one or more of the following: R32, E49, E51, L54, A56, Y117, M146, S148, and Y167, including any one, two, three, four, five, six, seven, eight, and nine of the above 9 sites; and the binding sites are numbered according to the amino acid sequence of TL1 A as set forth in SEQ ID NO: 327. Attorney Docket No.: 769273: SA9-943PC

[0030] In some embodiments, the isolated antigen-binding protein downregulates the expression of the a4p7 or the monomeric TL1 A and / or the trimer TL1 A.

[0031] In some embodiments, the isolated antigen-binding protein downregulates the expression of the membrane a4p7 or the monomeric TL1 A and / or the trimer TL1 A.

[0032] In some embodiments, the isolated antigen-binding protein comprises one or more polypeptides, wherein one of the polypeptides comprises at least one first antigen-binding domain and at least one second antigen-binding domain.

[0033] In some embodiments, the isolated antigen-binding protein comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises the first antigen-binding domain and the second antigen-binding domain, and the second polypeptide comprises a first antigen-binding domain or a second antigen-binding domain, and one or more of the following conditions are selected:

[0034] (1) the first antigen-binding domain of the first polypeptide is located at the C-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the C-terminus of the first polypeptide;

[0035] (2) the first antigen-binding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the N-terminus of the first polypeptide;

[0036] (3) the second antigen-binding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the second antigen-binding domain of the second polypeptide is located at the N-terminus of the second polypeptide;

[0037] (4) the second antigen-binding domain of the first polypeptide is located at the C-terminus of the second polypeptide, and the second antigen-binding domain of the second polypeptide is located at the C-terminus of the second polypeptide.

[0038] In some embodiments, the isolated antigen-binding protein comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises the first antigen-binding domain and the second antigen-binding domain, and the second polypeptide comprises the first Attorney Docket No.: 769273: SA9-943PC antigen-binding domain and the second antigen-binding domain, and one or more of the following conditions are selected:

[0039] (1) the first antigen-binding domain of the first polypeptide is located at the C-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the C-terminus of the first polypeptide;

[0040] (2) the first antigen-binding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the N-terminus of the first polypeptide;

[0041] (3) the second antigen-binding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the second antigen-binding domain of the second polypeptide is located at the N-terminus of the second polypeptide;

[0042] (4) the second antigen-binding domain of the first polypeptide is located at the C-terminus of the second polypeptide, and the second antigen-binding domain of the second polypeptide is located at the C-terminus of the second polypeptide.

[0043] In some embodiments, the isolated antigen-binding protein comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises the first antigen-binding domain and does not include the second antigen-binding domain, and the second polypeptide comprises the second antigen-binding domain and does not include the first antigen-binding domain.

[0044] In some embodiments, the first antigen-binding domain or the second antigen-binding domain comprises one or any combination of the following:

[0045] Fab, Fab', F(ab')2, Fd and Fv fragments, disulfide bond stabilized Fv fragments (dsFv)., (dsFv)2, bispecific dsFv (dsFv-dsFv1), disulfide bond stabilized double-stranded antibody (diabody) (ds double-stranded antibody), single-chain antibody molecule (scFv), Single domain antibody (SDAB), scFv dimer (divalent double-stranded antibody), single domain antibody or nanobody, domain antibody ( domain antibody (dAb), shark variable IgNAR domain, camelized VH domain, chicken heavy chain antibody domain, VHH domain, minimum identification unit composed of CDR of mimetic antibody ( minimal recognition Attorney Docket No.: 769273: SA9-943PC unit), alternative scaffolds for binding antigens, bivalent domain antibodies, pre-designed ankyrin repeat proteins (DARPins), multispecific proteins (including antigen-binding fragments or any other antibody fragments bound to antigens, but not intact antibody structures).

[0046] In some embodiments, the first antigen-binding domain comprises a heavy chain variable region (VH) containing an HCDR3 comprising an amino acid sequence of SEQ ID NO: 14.

[0047] In some embodiments, the HCDR3 comprises an amino acid sequence of any one of SEQ ID NO: 15-18.

[0048] In some embodiments, the VH comprises HCDR2 comprising an amino acid sequence of SEQ ID NO:5.

[0049] In some embodiments, the HCDR2 comprises an amino acid sequence of any one of SEQ ID NO:6-13.

[0050] In some embodiments, the VH comprises HCDR1 comprising an amino acid sequence of SEQ ID NO: 1.

[0051] In some embodiments, the HCDR1 comprises an amino acid sequence of any one of the SEQ ID NO:2-4.

[0052] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 15.

[0053] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 7, and the HCDR3 as shown in SEQ ID NO: 15.

[0054] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 8, and the HCDR3 as shown in SEQ ID NO: 15.

[0055] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 3, the HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 15.

[0056] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 16.

[0057] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the

[0058] HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 17. Attorney Docket No.: 769273: SA9-943PC

[0059] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the

[0060] HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 18.

[0061] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 4, the HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 15.

[0062] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 9, and the HCDR3 as shown in SEQ ID NO: 15.

[0063] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 10, and the HCDR3 as shown in SEQ ID NO: 15.

[0064] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 11, and the HCDR3 as shown in SEQ ID NO: 15.

[0065] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 12, and the HCDR3 as shown in SEQ ID NO: 15.

[0066] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 13, and the HCDR3 as shown in SEQ ID NO: 15.

[0067] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO:3, the HCDR2 as shown in SEQ ID NO:6, and the HCDR3 as shown in SEQ ID NO: 16.

[0068] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 3, the HCDR2 as shown in SEQ ID NO: 6, and the HCDR3 as shown in SEQ ID NO: 17.

[0069] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 3, the HCDR2 as shown in SEQ ID NO: 9, and the HCDR3 as shown in SEQ ID NO: 15.

[0070] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO:3, the HCDR2 as shown in SEQ ID NO: 10 and the HCDR3 as shown in SEQ ID NO: 15.

[0071] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 3, the HCDR2 as shown in SEQ ID NO: 11, and the HCDR3 as shown in SEQ ID NO: 15.

[0072] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 9, and the HCDR3 as shown in SEQ ID NO: 16.

[0073] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 10, and the HCDR3 as shown in SEQ ID NO: 16. Attorney Docket No.: 769273: SA9-943PC

[0074] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO:2, the

[0075] HCDR2 as shown in SEQ ID NO: 11, and the HCDR3 as shown in SEQ ID NO: 16.

[0076] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 2, the HCDR2 as shown in SEQ ID NO: 12, and the HCDR3 as shown in SEQ ID NO: 16.

[0077] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO:2, the HCDR2 as shown in SEQ ID NO: 10, and the HCDR3 as shown in SEQ ID NO: 17.

[0078] In some embodiments, the VH comprises an HFR1 containing an amino acid sequence of SEQ ID NO: 19.

[0079] In some embodiments, the HFR1 is selected from an amino acid sequence shown in any one of SEQ ID NO:20-28.

[0080] In some embodiments, the VH comprises an HFR2, and the HFR2 comprises an amino acid sequence of SEQ ID NO:29.

[0081] In some embodiments, the HFR2 is selected from an amino acid sequence shown in any one of SEQ ID NO:30-32.

[0082] In some embodiments, the VH comprises an HFR3 comprising an amino acid sequence of SEQ ID NO:33.

[0083] In some embodiments, the HFR3 is selected from an amino acid sequence shown in any one of SEQ ID NO:34-40.

[0084] In some embodiments, the VH comprises an HFR4 comprising an amino acid sequence shown in SEQ ID NO:41.

[0085] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 20, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 34, and the HFR4 as shown in SEQ ID NO: 41.

[0086] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 21, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41. Attorney Docket No.: 769273: SA9-943PC

[0087] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 22, the HFR2 as shown in SEQ ID NO: 32, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41.

[0088] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO:23, the HFR2 as shown in SEQ ID NO:31, the HFR3 as shown in SEQ ID NO:36, and the HFR4 as shown in SEQ ID NO:41.

[0089] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 23, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 37, and the HFR4 as shown in SEQ ID NO: 41.

[0090] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 23, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41.

[0091] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 24, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 38, and the HFR4 as shown in SEQ ID NO: 41.

[0092] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 25, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 37, and the HFR4 as shown in SEQ ID NO: 41.

[0093] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 25, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 39, and the HFR4 as shown in SEQ ID NO: 41.

[0094] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 25, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 40, and the HFR4 as shown in SEQ ID NO: 41.

[0095] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 26, the

[0096] HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41. Attorney Docket No.: 769273: SA9-943PC

[0097] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 25, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41.

[0098] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 27, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41.

[0099] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 28, the HFR2 as shown in SEQ ID NO: 31, the HFR3 as shown in SEQ ID NO: 35, and the HFR4 as shown in SEQ ID NO: 41.

[0100] In some embodiments, the first antigen-binding domain of the isolated antigen-binding fragment further comprises a light chain variable region (VL) containing at least one light chain complementarity-determining region (LCDR) containing amino acid sequences selected from an amino acid sequence shown in SEQ ID NO:42, SEQ ID NO:43, and SEQ ID NO:44.

[0101] In some embodiments, the VL comprises an LCDR3 comprising an amino acid sequence of SEQ ID NO:44.

[0102] In some embodiments, the VL comprises an LCDR2 comprising an amino acid sequence of SEQ ID NO:43.

[0103] In some embodiments, the VL comprises an LCDR1 comprising an amino acid sequence of SEQ ID NO:42.

[0104] In some embodiments, the VL comprises the LFR1 comprising an amino acid sequence of SEQ ID NO:45.

[0105] In some embodiments, the LFR1 comprises an amino acid sequence shown in either of the SEQ ID NO:46-47.

[0106] In some embodiments, the VL comprises the LFR2 comprising an amino acid sequence of SEQ ID NO:48.

[0107] In some embodiments, the LFR2 comprises an amino acid sequence shown in either SEQ

[0108] ID NO: 49-52. Attorney Docket No.: 769273: SA9-943PC

[0109] In some embodiments, the VL comprises the LFR3 comprising an amino acid sequence of SEQ ID NO:53.

[0110] In some embodiments, the LFR3 comprises an amino acid sequence shown in either of the SEQ ID NO: 54-57.

[0111] In some embodiments, the VL comprises the LFR4 comprising an amino acid sequence of SEQ ID NO:58.

[0112] In some embodiments, the LFR1 comprise an amino acid as set forth in any one of SEQ ID NO:59-61.

[0113] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 46, the LFR2 as shown in SEQ ID NO: 49, the LFR3 as shown in SEQ ID NO: 54, and the LFR4 as shown in SEQ ID NO: 59.

[0114] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 47, the LFR2 as shown in SEQ ID NO: 50, the LFR3 as shown in SEQ ID NO: 55, and the LFR4 as shown in SEQ ID NO: 60.

[0115] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 47, the LFR2 as shown in SEQ ID NO: 51, the LFR3 as shown in SEQ ID NO: 56, and the LFR4 as shown in SEQ ID NO: 61.

[0116] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 47, the LFR2 as shown in SEQ ID NO: 52, the LFR3 as shown in SEQ ID NO: 57, and the LFR4 as shown in SEQ ID NO: 61.

[0117] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 47, the LFR2 as shown in SEQ ID NO: 52, the LFR3 as shown in SEQ ID NO: 56, and the LFR4 as shown in SEQ ID NO: 61.

[0118] In some embodiments, the VH comprises a sequence as described in any one of SEQ ID NO:62-99.

[0119] In some embodiments, the VL comprises a sequence as described in any one of SEQ ID

[0120] NO: 102-106. Attorney Docket No.: 769273: SA9-943PC

[0121] An isolated antigen-binding protein comprising: a first antigen-binding domain and a second antigen -binding domain, the first antigen -binding domain specifically binds to TNF- like protein A (TL1A), and the first antigen-binding domain is different from the second antigen-binding domain.

[0122] In some embodiments, the first antigen-binding domain comprises a heavy chain variable region (VH) containing an amino acid sequence of an HCDR3 which comprises SEQ ID NO: 120.

[0123] In some embodiments, the VH comprises an HCDR2 comprising an amino acid sequence of SEQ ID NO: 115.

[0124] In some embodiments, the HCDR2 comprises an amino acid sequence of any one of SEQ ID NO: 116-119.

[0125] In some embodiments, the VH comprises an HCDR1 comprising an amino acid sequence of SEQ ID NO: 107.

[0126] In some embodiments, the HCDR1 comprises an amino acid sequence of any one of SEQ ID NO: 108-114.

[0127] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 108, the HCDR2 as shown in SEQ ID NO: 116, and the HCDR3 as shown in SEQ ID NO: 120.

[0128] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 109, the HCDR2 as shown in SEQ ID NO: 118, and the HCDR3 as shown in SEQ ID NO: 120.

[0129] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 108, the HCDR2 as shown in SEQ ID NO: 119, and the HCDR3 as shown in SEQ ID NO: 120.

[0130] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 110, the HCDR2 as shown in SEQ ID NO: 118, and the HCDR3 as shown in SEQ ID NO: 120.

[0131] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 111, the HCDR2 as shown in SEQ ID NO: 117, and the HCDR3 as shown in SEQ ID NO: 120.

[0132] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 112, the HCDR2 as shown in SEQ ID NO: 117, and the HCDR3 as shown in SEQ ID NO: 120. Attorney Docket No.: 769273: SA9-943PC

[0133] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 113, the

[0134] HCDR2 as shown in SEQ ID NO: 117, and the HCDR3 as shown in SEQ ID NO: 120.

[0135] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 108, the HCDR2 as shown in SEQ ID NO: 117, and the HCDR3 as shown in SEQ ID NO: 120.

[0136] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 114, the HCDR2 as shown in SEQ ID NO: 117 and the HCDR3 as shown in SEQ ID NO: 120.

[0137] In some embodiments, the VH comprises an HFR1 which comprises an amino acid sequence of SEQ ID NO : 121.

[0138] In some embodiments, the HFR1 is selected from an amino acid sequence shown in any one of SEQ ID NO: 122-132.

[0139] In some embodiments, the VH comprises an HFR2, and the HFR2 comprises an amino acid sequence of SEQ ID NO: 133.

[0140] In some embodiments, the HFR2 is selected from an amino acid sequence shown in any one of SEQ ID NO: 134-135, 30.

[0141] In some embodiments, the VH comprises an HFR3 comprising an amino acid sequence of SEQ ID NO: 136.

[0142] In some embodiments, the HFR3 is selected from an amino acid sequence shown in any one of SEQ ID NO: 137-148.

[0143] In some embodiments, the VH comprises HFR4 comprising an amino acid sequence shown in SEQ ID NO: 149.

[0144] In some embodiments, the HFR4 is selected from an amino acid sequence shown in ANY ONE of SEQ ID NO: 150-153, 41.

[0145] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 122, the HFR2 as shown in SEQ ID NO: 134, the HFR3 as shown in SEQ ID NO: 137, and the HFR4 as shown in SEQ ID NO: 150.

[0146] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 123, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 138, and the HFR4 as shown in SEQ ID NO: 41. Attorney Docket No.: 769273: SA9-943PC

[0147] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 124, the

[0148] HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 139, and the HFR4 as shown in SEQ ID NO: 151.

[0149] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 125, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 142, and the HFR4 as shown in SEQ ID NO: 41.

[0150] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 123, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 140, and the HFR4 as shown in SEQ ID NO: 41.

[0151] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 123, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 141, and the HFR4 as shown in SEQ ID NO: 41.

[0152] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 126, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 142, and the HFR4 as shown in SEQ ID NO: 152.

[0153] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 127, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 143, and the HFR4 as shown in SEQ ID NO: 41.

[0154] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 128, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 144, and the HFR4 as shown in SEQ ID NO: 41.

[0155] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 129, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 145, and the HFR4 as shown in SEQ ID NO: 41.

[0156] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 127, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 146, and the HFR4 as shown in SEQ ID NO: 41. Attorney Docket No.: 769273: SA9-943PC

[0157] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 125, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 142, and the HFR4 as shown in SEQ ID NO: 152.

[0158] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 130, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 147, and the HFR4 as shown in SEQ ID NO: 41.

[0159] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 131, the HFR2 as shown in SEQ ID NO: 135, the HFR3 as shown in SEQ ID NO: 148, and the HFR4 as shown in SEQ ID NO: 153.

[0160] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 132, the HFR2 as shown in SEQ ID NO: 30, the HFR3 as shown in SEQ ID NO: 145, and the HFR4 as shown in SEQ ID NO: 41.

[0161] In some embodiments, the first antigen-binding domain of the isolated antigen-binding fragment further comprises a light chain variable region (VL) containing at least one light chain complementarity-determining region (LCDR) containing amino acid sequences selected from SEQ ID NO: 154, SEQ ID NO: 159, and SEQ ID NO: 163.

[0162] In some embodiments, the VL comprises an LCDR3 comprising an amino acid sequence of SEQ ID NO: 163.

[0163] In some embodiments, the LCDR3 comprises an amino acid sequence shown in any one of SEQ ID NO: 164-168.

[0164] 1 In some embodiments, the VL comprises an LCDR2 comprising an amino acid sequence of SEQ ID NO: 159.

[0165] In some embodiments, the LCDR2 comprises an amino acid sequence shown in any one of SEQ ID NO: 160-162.

[0166] In some embodiments, the VL comprises an LCDR1 comprising an amino acid sequence of SEQ ID NO: 154.

[0167] In some embodiments, the LCDR1 comprises an amino acid sequence shown in any one of SEQ ID NO: 155-158. Attorney Docket No.: 769273: SA9-943PC

[0168] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 157, the LCDR2 as shown in SEQ ID NO: 161 and the LCDR3 as shown in SEQ ID NO: 164.

[0169] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 157, the LCDR2 as shown in SEQ ID NO: 162 and the LCDR3 as shown in SEQ ID NO: 164.

[0170] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 156, the LCDR2 as shown in SEQ ID NO: 161 and the LCDR3 as shown in SEQ ID NO: 164.

[0171] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 158, the LCDR2 as shown in SEQ ID NO: 160 and the LCDR3 as shown in SEQ ID NO: 164.

[0172] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 155, the LCDR2 as shown in SEQ ID NO: 160 and the LCDR3 as shown in SEQ ID NO: 164.

[0173] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 156, the LCDR2 as shown in SEQ ID NO: 162 and the LCDR3 as shown in SEQ ID NO: 164.

[0174] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 155, the LCDR2 as shown in SEQ ID NO: 160 and the LCDR3 as shown in SEQ ID NO: 165.

[0175] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 155, the LCDR2 as shown in SEQ ID NO: 160 and the LCDR3 as shown in SEQ ID NO: 166.

[0176] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 155, the LCDR2 as shown in SEQ ID NO: 160 and the LCDR3 as shown in SEQ ID NO: 167.

[0177] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 155, the LCDR2 as shown in SEQ ID NO: 160 and the LCDR3 as shown in SEQ ID NO: 168.

[0178] In some embodiments, the VL comprises the LFR1 comprising an amino acid sequence of SEQ ID NO: 169.

[0179] In some embodiments, the LFR1 comprises an amino acid sequence shown in any one of SEQ ID NO: 170-179.

[0180] In some embodiments, the VL comprises the LFR2 comprising an amino acid sequence of SEQ ID NO: 180.

[0181] In some embodiments, the LFR2 comprises an amino acid sequence shown in either of

[0182] SEQ ID NO: 181-188. Attorney Docket No.: 769273: SA9-943PC

[0183] In some embodiments, the VL comprises the LFR3 comprising an amino acid sequence of SEQ ID NO: 189.

[0184] In some embodiments, the LFR3 comprises an amino acid sequence shown in any one of SEQ ID NO: 190-197, 57.

[0185] In some embodiments, the VL comprises the LFR4 comprising an amino acid sequence of SEQ ID NO: 198.

[0186] In some embodiments, the LFR4 comprises an amino acid sequence shown in any one of the SEQ IDS NO: 199-201, 61.

[0187] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 170, the LFR2 as shown in SEQ ID NO: 181, the LFR3 as shown in SEQ ID NO: 190, and the LFR4 as shown in SEQ ID NO: 199.

[0188] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 171, the LFR2 as shown in SEQ ID NO: 182, the LFR3 as shown in SEQ ID NO: 191, and the LFR4 as shown in SEQ ID NO: 200.

[0189] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 172, the LFR2 as shown in SEQ ID NO: 183, the LFR3 as shown in SEQ ID NO: 192, and the LFR4 as shown in SEQ ID NO: 61.

[0190] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 174, the LFR2 as shown in SEQ ID NO: 184, the LFR3 as shown in SEQ ID NO: 57, and the LFR4 as shown in SEQ ID NO: 199.

[0191] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 175, the LFR2 as shown in SEQ ID NO: 185, the LFR3 as shown in SEQ ID NO: 193, and the LFR4 as shown in SEQ ID NO: 199.

[0192] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 176, the LFR2 as shown in SEQ ID NO: 186, the LFR3 as shown in SEQ ID NO: 194, and the LFR4 as shown in SEQ ID NO: 199. Attorney Docket No.: 769273: SA9-943PC

[0193] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 177, the LFR2 as shown in SEQ ID NO: 187, the LFR3 as shown in SEQ ID NO: 195, and the LFR4 as shown in SEQ ID NO: 199.

[0194] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 178, the LFR2 as shown in SEQ ID NO: 188, the LFR3 as shown in SEQ ID NO: 196, and the LFR4 as shown in SEQ ID NO: 199.

[0195] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 179, LFR2 as shown in SEQ ID NO: 187, LFR3 as shown in SEQ ID NO: 195, and LFR4 as shown in SEQ ID NO: 199.

[0196] In some embodiments, the VL comprises LFR1 as shown in SEQ ID NO: 173, LFR2 as shown in SEQ ID NO: 183, LFR3 as shown in SEQ ID NO: 192, and LFR4 as shown in SEQ ID NO: 201.

[0197] In some embodiments, the VL comprises LFR1 as shown in SEQ ID NO: 173, LFR2 as shown in SEQ ID NO: 183, LFR3 as shown in SEQ ID NO: 192, and LFR4 as shown in SEQ ID NO: 199.

[0198] In some embodiments, the VH comprises a sequence as described in any one of SEQ ID N0:202-220.

[0199] In some embodiments, the VL comprises a sequence as described in any one of SEQ ID NO: 221-234.

[0200] An isolated antigen-binding protein comprising: a first antigen-binding domain and a second antigen -binding domain, the first antigen -binding domain specifically binds to TNF- like protein A (TL1A), and the first antigen-binding domain is different from the second antigen-binding domain.

[0201] In some embodiments, the first antigen binding domain comprises a heavy chain variable region (VH) containing an HCDR3 which comprises an amino acid sequence of SEQ ID NO:242.

[0202] In some embodiments, the VH comprises an HCDR2 comprising an amino acid sequence of SEQ ID NO:238. Attorney Docket No.: 769273: SA9-943PC

[0203] In some embodiments, the HCDR2 comprises an amino acid sequence of any one of SEQ

[0204] ID NO: 239-241.

[0205] In some embodiments, the VH comprises an HCDR1 comprising an amino acid sequence of SEQ ID NO:235.

[0206] In some embodiments, the HCDR1 comprises an amino acid sequence of any one of SEQ ID NO: 236-237.

[0207] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 108, the HCDR2 as shown in SEQ ID NO: 116, and the HCDR3 as shown in SEQ ID NO: 120.

[0208] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 236, the HCDR2 as shown in SEQ ID NO: 239, and the HCDR3 as shown in SEQ ID NO: 242.

[0209] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 236, the HCDR2 as shown in SEQ ID NO: 240, and the HCDR3 as shown in SEQ ID NO: 242.

[0210] In some embodiments, the VH comprises the HCDR1 as shown in SEQ ID NO: 237, the HCDR2 as shown in SEQ ID NO: 241, and the HCDR3 as shown in SEQ ID NO: 242.

[0211] In some embodiments, the VH comprises an HFR1 comprising an amino acid sequence of SEQ ID NO:243.

[0212] In some embodiments, the HFR1 is selected from an amino acid sequence shown in any one of SEQ ID NO:244-250.

[0213] In some embodiments, the VH comprises an HFR2, and the HFR2 comprises an amino acid sequence of SEQ ID NO:251.

[0214] In some embodiments, the HFR2 is selected from an amino acid sequence shown in any one of SEQ ID NO: 252-255.

[0215] In some embodiments, the VH comprises an HFR3 comprising an amino acid sequence of SEQ ID NO:256.

[0216] In some embodiments, the HFR3 is selected from an amino acid sequence shown in any one of SEQ ID NO: 257-262.

[0217] In some embodiments, the VH comprises an HFR4 comprising an amino acid sequence shown in SEQ ID NO:263. Attorney Docket No.: 769273: SA9-943PC

[0218] In some embodiments, the HFR4 is selected from an amino acid sequence shown in any one of SEQ ID NO: 264-268.

[0219] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 244, the HFR2 as shown in SEQ ID NO: 252, the HFR3 as shown in SEQ ID NO: 257, and the HFR4 as shown in SEQ ID NO: 264.

[0220] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 245, the HFR2 as shown in SEQ ID NO: 252, the HFR3 as shown in SEQ ID NO: 258, and the HFR4 as shown in SEQ ID NO: 265.

[0221] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 246, the HFR2 as shown in SEQ ID NO: 253, the HFR3 as shown in SEQ ID NO: 259, and the HFR4 as shown in SEQ ID NO: 268.

[0222] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 246, the HFR2 as shown in SEQ ID NO: 254, the HFR3 as shown in SEQ ID NO: 260, and the HFR4 as shown in SEQ ID NO: 266.

[0223] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 247, the HFR2 as shown in SEQ ID NO: 254, the HFR3 as shown in SEQ ID NO: 261, and the HFR4 as shown in SEQ ID NO: 268.

[0224] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 248, the HFR2 as shown in SEQ ID NO: 252, the HFR3 as shown in SEQ ID NO: 258, and the HFR4 as shown in SEQ ID NO: 265.

[0225] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 246, the HFR2 as shown in SEQ ID NO: 255, the HFR3 as shown in SEQ ID NO: 260, and the HFR4 as shown in SEQ ID NO: 268.

[0226] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 245, the HFR2 as shown in SEQ ID NO: 252, the HFR3 as shown in SEQ ID NO: 258, and the HFR4 as shown in SEQ ID NO: 267. Attorney Docket No.: 769273: SA9-943PC

[0227] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 249, the

[0228] HFR2 as shown in SEQ ID NO: 254, the HFR3 as shown in SEQ ID NO: 260, and the HFR4 as shown in SEQ ID NO: 268.

[0229] In some embodiments, the VH comprises the HFR1 as shown in SEQ ID NO: 250, the HFR2 as shown in SEQ ID NO: 253, the HFR3 as shown in SEQ ID NO: 262, and the HFR4 as shown in SEQ ID NO: 266.

[0230] In some embodiments, the isolated antigen-binding fragments further include a light chain variable region (VL) containing at least one light chain complementarity-determining region (LCDR) containing amino acid sequences selected from an amino acid sequences shown in SEQ ID NO: 269, SEQ ID NO: 274, and SEQ ID NO: 277.

[0231] In some embodiments, the VL comprises an LCDR3 comprising an amino acid sequence of SEQ ID NO:277.

[0232] In some embodiments, the VL comprises an LCDR2 comprising an amino acid sequence of SEQ ID NO:274.

[0233] In some embodiments, the LCDR2 comprises an amino acid sequence shown in any one of SEQ ID NO:275-276.

[0234] In some embodiments, the VL comprises an LCDR1 comprising an amino acid sequence of SEQ ID NO:269.

[0235] In some embodiments, the LCDR1 comprises an amino acid sequence shown in any one of SEQ ID NO:270-273.

[0236] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 270, the LCDR2 as shown in SEQ ID NO: 275 and the LCDR3 as shown in SEQ ID NO: 277.

[0237] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 271, the LCDR2 as shown in SEQ ID NO: 275 and the LCDR3 as shown in SEQ ID NO: 277.

[0238] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 272, the LCDR2 as shown in SEQ ID NO: 275 and the LCDR3 as shown in SEQ ID NO: 277.

[0239] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 273, the LCDR2 as shown in SEQ ID NO: 275 and the LCDR3 as shown in SEQ ID NO: 277. Attorney Docket No.: 769273: SA9-943PC

[0240] In some embodiments, the VL comprises the LCDR1 as shown in SEQ ID NO: 271, the

[0241] LCDR2 as shown in SEQ ID NO: 276 and the LCDR3 as shown in SEQ ID NO: 277.

[0242] In some embodiments, the VL comprises the LFR1 comprising an amino acid sequence of SEQ ID NO:278.

[0243] In some embodiments, the LFR1 comprises an amino acid sequence as shown in any of SEQ ID NO: 47, 173, 279-283.

[0244] In some embodiments, the VL comprises the LFR2 comprising an amino acid sequence of SEQ ID NO:284.

[0245] In some embodiments, the LFR2 comprises amino acid sequences as shown in any of SEQ ID NO: 52, 285-287.

[0246] In some embodiments, the VL comprises the LFR3 comprising an amino acid sequence of SEQ ID NO:288.

[0247] In some embodiments, the LFR3 comprises an amino acid sequence shown in any one of SEQ ID NO: 289-293, 57.

[0248] In some embodiments, the VL comprises the LFR4 comprising an amino acid sequence of SEQ ID NO:294.

[0249] In some embodiments, the LFR4 comprises an amino acid sequence shown in any one of SEQ ID NO: 201, 295-296.

[0250] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 170, the LFR2 as shown in SEQ ID NO: 181, the LFR3 as shown in SEQ ID NO: 190, and the LFR4 as shown in SEQ ID NO: 199.

[0251] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 171, the LFR2 as shown in SEQ ID NO: 182, the LFR3 as shown in SEQ ID NO: 191, and the LFR4 as shown in SEQ ID NO: 200.

[0252] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 172, the LFR2 as shown in SEQ ID NO: 183, the LFR3 as shown in SEQ ID NO: 192, and the LFR4 as shown in SEQ ID NO: 61. Attorney Docket No.: 769273: SA9-943PC

[0253] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 174, the LFR2 as shown in SEQ ID NO: 184, the LFR3 as shown in SEQ ID NO: 57, and the LFR4 as shown in SEQ ID NO: 199.

[0254] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 175, LFR2 as shown in SEQ ID NO: 185, LFR3 as shown in SEQ ID NO: 193, and LFR4 as shown in SEQ ID NO: 199.

[0255] In some embodiments, the VL comprises LFR1 as shown in SEQ ID NO: 176, LFR2 as shown in SEQ ID NO: 186, LFR3 as shown in SEQ ID NO: 194, and LFR4 as shown in SEQ ID NO: 199.

[0256] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 177, the LFR2 as shown in SEQ ID NO: 187, the LFR3 as shown in SEQ ID NO: 195, and the LFR4 as shown in SEQ ID NO: 199.

[0257] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 178, the LFR2 as shown in SEQ ID NO: 188, the LFR3 as shown in SEQ ID NO: 196, and the LFR4 as shown in SEQ ID NO: 199.

[0258] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 179, the LFR2 as shown in SEQ ID NO: 187, the LFR3 as shown in SEQ ID NO: 195, and the LFR4 as shown in SEQ ID NO: 199.

[0259] In some embodiments, the VL comprises the LFR1 as shown in SEQ ID NO: 173, the LFR2 as shown in SEQ ID NO: 183, the LFR3 as shown in SEQ ID NO: 192, and the LFR4 as shown in SEQ ID NO: 201.

[0260] In some embodiments, the VH comprises a sequence as described in any one of SEQ ID N0:202-220.

[0261] In some embodiments, the VL comprises a sequence as described in any one of SEQ ID NO: 221-234.

[0262] In some embodiments, it also comprises a heavy chain constant region for IgG and / or a light chain constant region for human antibodies. Attorney Docket No.: 769273: SA9-943PC

[0263] In some embodiments, the isolated antigen-binding fragment comprises a heavy chain constant region of IgGl, IgG2, IgG3, or IgG4.

[0264] In some embodiments, the isolated antigen-binding fragment comprises a Kappa or Lambda light chain constant region.

[0265] In some embodiments, the isolated antigen-binding fragment is IgG, IgM, IgA, IgD, or IgE.

[0266] In some embodiments, the antigen-binding protein comprises one or more first heavy chain and one or more first light chain, and the first heavy chain comprises VH of the first antigen-binding domain and / or VH of the second antigen binding domain and / or scFv of the first antigen-binding domain and / or scFv of the second antigen binding domain; the first light chain comprises a VL of the first antigen-binding domain or a VL of the second antigen binding domain.

[0267] In some embodiments, the first heavy chain comprises VH of the first antigen-binding domain and scFv of the second antigen binding domain, or the first heavy chain comprises VH of the second antigen binding domain and scFv of the first antigen-binding domain.

[0268] In some embodiments, the N-terminus of the scFv of the first antigen-binding domain or the second antigen binding domain is either directly or by linker to the C-terminus of the first heavy chain, or the C-terminus of the scFv of the first antigen-binding domain or the second antigen binding domain is connected directly or by a linker to the N-terminus of the first heavy chain.

[0269] In some embodiments,

[0270] ( 1 ) the C-terminus of the first antigen-binding domain VH is connected directly or by linkers to the N-terminus of the first heavy chain, and the N-terminus of the second antigen binding domain of scFv is connected directly or by linkers to the C-terminus of the first heavy chain; The C-terminus of the VL of the first antigen-binding domain is attached to the N- terminus of the constant region of the light chain;

[0271] ( 2 ) the C-terminus of the second antigen binding domain VH is connected directly or by linker to the N-terminus of the first heavy chain, and the N-terminus of scFv of the first Attorney Docket No.: 769273: SA9-943PC antigen-binding domain is connected directly or by linker to the C-terminus of the first heavy chain; The C-terminus of the VL of the second antigen binding domain is connected to the N- terminus of the constant region of the light chain;

[0272] (3 ) the C-terminus of scFv of the first antigen-binding domain is connected directly or by a linker to the N-terminus of the second antigen binding domain VH, and the C-terminus of the VL of the second antigen binding domain is connected to the N-terminus of the constant region of the light chain;

[0273] (4) the C-terminus of the scFv of the second antigen binding domain is connected directly or by a linker to the N-terminus of the first antigen-binding domain VH, and the C- terminus of the VL of the first antigen-binding domain is connected to the N-terminus of the constant region of the light chain.

[0274] In some embodiments, the first heavy chain is selected from a sequence described in any one of SEQ ID NO: 350-366 and 378.

[0275] In some embodiments, the antigen-binding fragment is selected from one of the following:

[0276] 1) a light chain as shown in SEQ ID NO: 348 and heavy chain as shown in SEQ ID NO: 350;

[0277] 2) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 351;

[0278] 3) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 352;

[0279] 4) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 353;

[0280] 5) a light chain as shown in SEQ ID NO: 349 and a heavy chain as shown in SEQ ID NO: 354;

[0281] 6) a light chain as shown in SEQ ID NO: 349 and a heavy chain as shown in SEQ ID NO: 355;

[0282] 7) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 356; Attorney Docket No.: 769273: SA9-943PC

[0283] 8) a light chain as shown in SEQ ID NO: 372 and a heavy chain as shown in SEQ ID

[0284] NO: 357;

[0285] 9) a light chain as shown in SEQ ID NO: 349 and a heavy chain as shown in SEQ ID NO: 358;

[0286] 10) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 359;

[0287] 11) a light chain as shown in SEQ ID NO: 372 and a heavy chain as shown in SEQ ID NO: 360;

[0288] 12) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 361;

[0289] 13) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 362;

[0290] 14) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 363;

[0291] 15) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 364;

[0292] 16) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 365;

[0293] 17) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 366; 18) a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 378.

[0294] In some embodiments, the isolated antigen-binding protein includes a light chain as shown in SEQ ID NO: 348 and heavy chain as shown in SEQ ID NO: 363.

[0295] In some embodiments, the antigen-binding protein comprises one or more first light chains, one or more third heavy chains, and one or more fourth heavy chains, wherein:

[0296] 1) the third heavy chain comprises VH for the first antigen binding domain or the second antigen binding domain, and the fourth heavy chain comprises scFv for the first or second antigen binding domain, or; Attorney Docket No.: 769273: SA9-943PC

[0297] 2) the third heavy chain comprises scFv for the first antigen binding domain or the second antigen binding domain, and the fourth heavy chain comprises VH for the first or second antigen binding domain.

[0298] In some embodiments, the N-terminus of the VH of the first antigen-binding domain or the second antigen binding domain is connected directly or through a linker to the C-terminus of the third heavy chain, or the N-terminus of scFv of the first or second antigen binding domain is connected directly or through a linker to the C-terminus of the fourth heavy chain.

[0299] In some embodiments, wherein

[0300] 1) the third heavy chain comprises VH for the first antigen-binding domain, and the fourth heavy comprises scFv for the second antigen binding domain, or;

[0301] 2) the third heavy chain comprises scFv for the first antigen-binding domain, and the fourth heavy chain comprises VH for the second antigen binding domain, or;

[0302] 3) the third heavy chain comprises VH for the second antigen binding domain, and the fourth heavy comprises scFv for the first antigen-binding domain, or

[0303] 4) the third heavy chain comprises scFv for the second antigen binding domain, and the fourth heavy comprises VH for the first antigen-binding domain.

[0304] In some embodiments, the antigen-binding fragment is selected from one of the following:

[0305] 1) a light chain as shown in SEQ ID NO: 349, a third heavy chain as shown in SEQ ID NO: 367 and a fourth heavy chain as shown in SEQ ID NO: 368;

[0306] 2) a light chain as shown in SEQ ID NO: 348, a third heavy chain as shown in SEQ ID NO: 369 and a fourth heavy chain as shown in SEQ ID NO: 370.

[0307] In some embodiments, the first antigen-binding domain comprises HCDR1 as set forth in SEQ ID NO: 2, HCDR2 as set forth in SEQ ID NO: 6, HCDR3 as set forth in SEQ ID NO: 15, LCDR1 as set forth in SEQ ID NO: 42, LCDR2 as set forth in SEQ ID NO: 43, and LCDR3 as set forth in SEQ ID NO: 44; the second antigen-binding domain comprises HCDR1 as set forth in SEQ ID NO: 332, HCDR2 as set forth in SEQ ID NO: 333, HCDR3 as set forth in SEQ ID NO: 334, LCDR1 as set forth in SEQ ID NO: 336, LCDR2 as set forth in SEQ ID NO: 337, and LCDR3 as set forth in SEQ ID NO: 338. Attorney Docket No.: 769273: SA9-943PC

[0308] In some embodiments, the first antigen-binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106, and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339.

[0309] In some embodiments, (1) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 340 and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or, (2) the first antigenbinding domain comprises an scFv as set forth in SEQ ID NO: 341 and the second antigenbinding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or, (3) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 342; and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or, (4) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 343 and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or, (5) the first antigen binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106 and the second antigen binding domain comprises an scFv as set forth in 346; or, (6) the first antigen binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106 and the second antigen binding domain comprises an scFv as set forth in SEQ ID NO:347.

[0310] In some embodiments, the isolated antigen-binding protein includes a light chain as shown in SEQ ID NO: 348 and heavy chain as shown in SEQ ID NO: 363.

[0311] In another aspect, the present application provides an isolated nucleic acid molecule encoding the isolated antigen-binding fragment.

[0312] In another aspect, the present application provides an expression vector which comprises the aforementioned nucleic acids.

[0313] In another aspect, the present application provides a host cell containing the aforementioned isolated nucleic acids and / or the expression vector. Attorney Docket No.: 769273: SA9-943PC

[0314] In another aspect, the present application provides a composition containing the aforementioned isolated antigen-binding fragment and optionally a pharmaceutically acceptable carrier.

[0315] In another aspect, the application provides a method for preparing antigen-binding fragments isolated from TL1A, comprising: culturing the host cells claimed above, and recovering the isolated antigen-binding fragments.

[0316] In another aspect, the present application provides a method for the treatment of TL1A- mediated disease or symptoms, which comprises: administrating of the aforementioned isolated antigen-binding fragments, aforementioned isolated nucleic acids, and / or aforementioned expression vectors, aforementioned host cells, or aforementioned combinations.

[0317] Antigen-binding fragments isolated as described above, nucleic acids isolated as described above, and / or expression vectors described above, host cells as described above, or combinations described above, which are used for the treatment of TL1 A-mediated diseases or symptoms.

[0318] The use of the aforementioned isolated antigen-binding fragments, aforementioned isolated nucleic acids, and / or aforementioned expression vectors, aforementioned host cells, or aforementioned compositions in the treatment of TL1 A-mediated diseases or symptoms.

[0319] The use of aforementioned isolated antigen-binding fragments, the aforementioned isolated nucleic acids, and / or2 the aforementioned expression vectors, aforementioned host cells, or the aforementioned combinations in the preparation of drugs for the treatment of TL1 A-mediated diseases or conditions.

[0320] In some embodiments, the disease or symptom mediated by TLlA is inflammatory.

[0321] In some embodiments, TLlA-mediated inflammatory disease is selected from one of the following: Non-limiting examples of inflammatory disease include, allergy, ankylosing spondylitis, asthma, atopic dermatitis, autoimmune diseases or disorders, cancer, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, diabetes Attorney Docket No.: 769273: SA9-943PC

[0322] (e.g., type 1 diabetes and type 2 diabetes), glomerulonephritis, gout, hepatitis (e.g., active hepatitis), an immune-mediated disease or disorder, inflammatory bowel disease (IBD) such as Crohn's disease and ulcerative colitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), multiple sclerosis, neurodegenerative diseases of aging, periodontal disease (e.g., periodontitis), preperfusion injury transplant rejection, psoriasis, pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis), rheumatic disease, scleroderma, sinusitis, tuberculosis.

[0323] In some embodiments, inflammatory bowel disease (IBD) comprises ulcerative colitis (UC) and Crohn's disease (CD).

[0324] In some embodiments, the disease or symptom mediated by TL1A is an autoimmune disease.

[0325] In some embodiments, TLlA-mediated autoimmune disease is selected from one of the following: achalasia, Addison's disease, Adult Stil's disease, agammaglobulinemia, alopecia areata, amyloidosis, ankylosing spondylitis, anti-GBM / anti-TBM nephritis, antiphospholipid syndrome, autoimmune angioedema, autoimmune familial autonomic dysfunction, autoimmune encephalomyelitis, autoimmune hepatitis, autoimmune inner ear disease (AIED), autoimmune myocarditis, autoimmune oophoritis, autoimmune orchitis, autoimmune pancreatitis, autoimmune retinopathy, autoimmune urticaria, axonal and neuronal neuropathy (AMAN), Barlow disease (Bal 6 disease), Behcet's disease, benign mucous membrane pemphigoid, bullous pemphigoid, Castleman's disease ( Castleman disease (CD), celiac disease, Chagas disease, chronic inflammatory demyelinating polyneuropathy (CIDP), chronic recurrent multifocal osteomyelitis (CRMO), Churg-Strauss syndrome (CSS) or eosinophilic granulomatous disease (EGPA), cicatricial pemphigoid, and Cogan's syndrome), cold agglutinin disease, congenital heart block, coxsackienew myocarditis, CREST syndrome, Crohn's disease, dermatitis herpetiformis, dermatomyositis, Devic's disease (neuromyelitis optica), discoid lupus, Dressier's syndrome, endometriosis, eosinophilic esophagitis (EoE), eosinophilic fasciitis, erythema nodosum, primary mixed cryoglobulinemia, Evans syndrome, fibromyalgia, fibrotic alveolitis, giant cell arteritis (temporal arteritis), giant cell myocarditis, glomerulonephritis, Goodpasture's syndrome, granulomatosis with polyangiitis, Graves' Attorney Docket No.: 769273: SA9-943PC disease, Guillain-Barre syndrome syndrome), Hashimoto's thyroiditis, hemolytic anemia, Henoch- Schonl ein purpura (HSP), herpes gravilens or pemphigoid gestationum (PG), hidradenitis suppurativa (HS) (paradoxical acne), hypogammaglobulinemia, IgA nephropathy, IgG4-associated sclerotic disease, immune thrombocytopenic purpura (ITP), Inclusion body myositis (IBM), interstitial cystitis (IC), juvenile arthritis, juvenile diabetes mellitus (type 1 diabetes), juvenile myositis (JM), Kawasaki disease, Lambert-Eaton syndrome, leukocytoclastic vasculitis, lichen planus, lichen sclerosus, lignoconjunctivitis, linear IgA disease (LAD), lupus, chronic Lyme disease ( Lymedisease chronic), Meniere's disease, microscopic polyangiitis (MPA), mixed connective tissue disease (MCTD), Mooren's ulcer, Mucha-Habermann disease, multifocal motor neuropathy (MMN) or MMNCB, multiple sclerosis, myasthenia gravis, myositis, narcolepsy, neonatal lupus, neuromyelitis optica, neutropenia, ocular cicatricial pemphigoid, optic neuritis, recurrent rheumatism (PR), PandAS, paraneoplastic cerebellar degeneration (PCD), paroxysmal nocturnal hemoglobinuria (PNH), Parry Romberg syndrome, ciliary planitis (peripheral uveitis), Parsonage-Turner syndrome), pemphigus, peripheral neuropathy, perivenous encephalomyelitis, pernicious anemia (PA), POEMS syndrome, polyarteritis nodosa, polyglandular syndrome type I, II, type III, polymyalgia rheumatica, polymyositis, post-myocardial infarction syndrome, postpericardiotomy syndrome, primary biliary cirrhosis, primary sclerosing cholangitis, progesterone dermatitis, psoriasis, psoriatic arthritis, pure red blood cell aplasia (PRCA), pyoderma gangrenosum, Raynaud's phenomenon, reactive arthritis, reflex sympathetic dystrophy, relapsing polychondritis, restless legs syndrome (RLS), retroperitoneal fibrosis, rheumatic fever, rheumatoid arthritis, sarcoidosis, Schmidt syndrome, scleritis, scleroderma, Sjogren's syndrome, sperm and testicular autoimmunity, stiff-person syndrome (SPS), subacute bacterial endocarditis (SBE), Susac's syndrome syndrome), sympathetic ophthalmia (SO), Takayasu's arteritis, temporal arteritis / giant cell arteritis, thrombocytopenic purpura (TTP), Tolosa-Hunt syndrome (THS), transverse myelitis, type 1 diabetes mellitus, ulcerative colitis (UC), undifferentiated connective tissue disease (UCTD), uveitis, vasculitis, Vitiligo and Vogt- Koyanagi-Harada disease. Attorney Docket No.: 769273: SA9-943PC

[0326] In some embodiments, the disease or symptom mediated by TLlA is cancer.

[0327] In some embodiments, TLlA-mediated cancers are selected from one or more of the following: non-restrictive examples of cancer include adenoid cystic carcinoma, adrenal carcinoma, amyloidosis, anal cancer, ataxia-telangiectasia, atypical nevus syndrome, basal cell carcinoma, cholangiocarcinoma, Birt Hogg Dube syndrome, bladder cancer, bone cancer, brain tumor, breast cancer, male breast cancer, carcinoid tumor, cervical cancer, colorectal cancer, ductal cancer, Endometrial Cancer, Esophageal Cancer, Gastric Cancer, Gastrointestinal Stromal Tumor (GIST), HER2 -Positive Breast Cancer, Pancreatic Islet Cell Tumor, Juvenile Polyposis Syndrome, Kidney Cancer, Laryngeal Cancer, Leukemia-Acute Lymphoblastic Leukemia, Acute Lymphoblastic Leukemia (ALL), Acute Myeloid Leukemia AML, Adult Leukemia, Childhood Leukemia, 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's lymphoma (Lymphoma-Hodgkin's), non-Hodgkin's lymphoma (Lymphoma-Non-Hodgkin ' s), 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 tumors, polycythemia vera, prostate cancer, renal cell carcinoma, retinoblastoma, salivary gland carcinoma, sarcoma, Sarcoma-Kaposi, skin cancer, small bowel cancer, stomach cancer, testicular cancer, thymoma, thyroid cancer, uterine (endometrial) cancer, vaginal cancer, and Wilms’ Tumor.

[0328] On the other hand, the present application provides a method for downregulating the expression of a4p7 and / or TL1 A, comprising providing a TL1 Ainhibitor and an a4p7 inhibitor to a subject.

[0329] On the other hand, the present application provides a method for inducing the endocytosis of cell surface a4p7 and / or TL1 A, comprising providing a TL1 Ainhibitor and an a4p7 inhibitor to a subject. Attorney Docket No.: 769273: SA9-943PC

[0330] In some embodiments, the method further comprises: providing an antigen binding protein that can specifically bind to a4p7 or providing an antigen binding protein that can specifically bind to TL1A.

[0331] In some embodiments, the method further comprises: providing a TL1A protein or an a4p7 protein.

[0332] In some embodiments, the isolated antigen binding protein that can specifically bind to TL1 A and the antigen binding protein that can specifically bind to a4p7 are the same protein.

[0333] In some embodiments, the isolated antigen binding protein comprises one or more polypeptides, wherein one polypeptide comprises at least a first antigen binding domain and at least one second antigen binding domain.

[0334] In some embodiments, the isolated antigen-binding protein comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a first antigen-binding domain and a second antigen-binding domain, and the second polypeptide comprises a first antigen-binding domain or a second antigen-binding domain, and one or more of the following situations are selected: (1) the first antigen-binding domain of the first polypeptide is located at the C-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the C-terminus of the first polypeptide; (2) the first antigen-binding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the N-terminus of the first polypeptide; (3) the second antigen-binding domain of the first polypeptide is located at the N- terminus of the first polypeptide, and the second antigen-binding domain of the second polypeptide is located at the N-terminus of the second polypeptide; (4) the second antigenbinding domain of the first polypeptide is located at the C-terminus of the second polypeptide, and the second antigen-binding domain of the second polypeptide is located at the C-terminus of the second polypeptide.

[0335] In some embodiments, the isolated antigen-binding protein comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises a first antigen-binding domain and a second antigen-binding domain, and the second polypeptide comprises a first Attorney Docket No.: 769273: SA9-943PC antigen-binding domain and a second antigen-binding domain, and one or more of the following situations are selected: (1) the first antigen-binding domain of the first polypeptide is located at the C-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the C-terminus of the first polypeptide; (2) the first antigenbinding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the first antigen-binding domain of the second polypeptide is located at the N-terminus of the first polypeptide; (3) the second antigen-binding domain of the first polypeptide is located at the N-terminus of the first polypeptide, and the second antigen-binding domain of the second polypeptide is located at the N-terminus of the second polypeptide; (4) the second antigenbinding domain of the first polypeptide is located at the C-terminus of the second polypeptide, and the second antigen-binding domain of the second polypeptide is located at the C-terminus of the second polypeptide.

[0336] In some embodiments, the isolated antigen-binding protein comprises a first polypeptide and a second polypeptide, wherein the first polypeptide comprises the first antigen-binding domain and does not include the second antigen-binding domain, and the second polypeptide comprises the second antigen-binding domain and does not include the first antigen-binding domain.

[0337] In some embodiments, the isolated antigen binding protein specifically binds to the a4p7 protein with a KD value of about 5E-08M or less.

[0338] In some embodiments, the isolated antigen binding protein specifically binds to monomeric TL1A and / or trimeric TL1A with a KD value of about 5E-08M or less.

[0339] On the other hand, the present application provides a use of the antigen binding protein to induce the endocytosis of cell surface a4p7 and / or TL1A.

[0340] On the other hand, the present application provides a use of the antigen binding protein to downregulate the expression of a4p7 and / or TL1A. Attorney Docket No.: 769273: SA9-943PC

[0341] A bispecific antibody, comprising: a first antigen-binding domain and a second antigenbinding domain, wherein the first antigen-binding domain specifically binds to TNF-like protein A (TL1 A), and wherein the second antigen binding domain specifically binds to a4p7.

[0342] In some embodiments, the bispecific antibody comprises: a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain specifically binds to TNF-like protein A (TL1 A), and the second antigen binding domain specifically binds to a4p7, wherein the first antigen-binding domain comprises a heavy chain variable region (VH), and the VH comprises an HCDR3 SEQ ID NO: 14.

[0343] In some embodiments, the HCDR3 of the first antigen-binding domain of the bispecific antibody comprises an amino acid sequence of any one of SEQ ID NOs: 15-18.

[0344] In some embodiments, the VH of the first antigen-binding domain of the bispecific antibody comprises an HCDR2 comprising an amino acid sequence of SEQ ID NO:5.

[0345] In some embodiments, the HCDR2 of the first antigen-binding domain of the bispecific antibody comprises an amino acid sequence of any one of SEQ ID NOs: 6-13.

[0346] In some embodiments, the VH of the first antigen-binding domain of the bispecific antibody comprises an HCDR1 comprising an amino acid sequence of SEQ ID NO: 1.

[0347] In some embodiments, the HCDR1 of the first antigen-binding domain of the bispecific antibody comprises an amino acid sequence of any one of SEQ ID NOs:2-4.

[0348] In some embodiments;

[0349] (i) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 15;

[0350] (ii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 7, and the HCDR3 as set forth in SEQ ID NO: 15; Attorney Docket No.: 769273: SA9-943PC

[0351] (iii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 8, and the HCDR3 as set forth in SEQ ID NO: 15;

[0352] (iv) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 6 and the HCDR3 as set forth in SEQ ID NO: 15;

[0353] (v) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 16;

[0354] (vi) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 17;

[0355] (vii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 18;

[0356] (viii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 4, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 15;

[0357] (ix) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 9, and the HCDR3 as set forth in SEQ ID NO: 15;

[0358] (x) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 15;

[0359] (xi) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 11, and the HCDR3 as set forth in SEQ ID NO: 15;

[0360] (xii) the VH of the first antigen-binding domain of the bispecific antibody comprises the

[0361] HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 12, and the

[0362] HCDR3 as set forth in SEQ ID NO: 15; Attorney Docket No.: 769273: SA9-943PC

[0363] (xiii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 13, and the HCDR3 as set forth in SEQ ID NO: 15;

[0364] (xiv) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 16;

[0365] (xv) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 17;

[0366] (xvi) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 9, and the HCDR3 as set forth in SEQ ID NO: 15;

[0367] (xvii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 15;

[0368] (xviii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 11, and the HCDR3 as set forth in SEQ ID NO: 15;

[0369] (xix) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 9, and the HCDR3 as set forth in SEQ ID NO: 16;

[0370] (xx) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 16;

[0371] (xxi) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 11, and the HCDR3 as set forth in SEQ ID NO: 16;

[0372] (xxii) the VH of the first antigen-binding domain of the bispecific antibody comprises the

[0373] HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 12, and the

[0374] HCDR3 as set forth in SEQ ID NO: 16 or Attorney Docket No.: 769273: SA9-943PC

[0375] (xxiii) the VH of the first antigen-binding domain of the bispecific antibody comprises the HCDR1 as set forth in SEQ ID NO:2, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 17.

[0376] In some embodiments, the first antigen-binding domain of the bispecific antibody further comprises a light chain variable region (VL), wherein the VL of the first antigen-binding domain comprises at least one light chain complementary determining region (LCDR) comprising an amino acid sequence selected from SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44.

[0377] In some embodiments, the VH of the first antigen-binding domain of the bispecific antibody comprises a sequence as set forth in any one of SEQ ID NOs: 62-99 or 379.

[0378] In some embodiments, the VL of the first antigen-binding domain of the bispecific antibody comprises a sequence as set forth in in any one of SEQ ID NOs: 102-106 or 380.

[0379] In some embodiments, the second antigen binding domain of the bispecific antibody comprises a heavy chain variable region (VH), and the VH of the second antigen binding domain comprises an HCDR1 as set forth in SEQ ID NO: 332, an HCDR2 as set forth in SEQ ID NO: 333, and an HCDR3 as set forth in SEQ ID NO: 334, optionally wherein the second antigen binding domain comprises a VH as set forth in SEQ ID NO: 335.

[0380] In some embodiments, wherein the second antigen binding domain of the bispecific antibody comprises a light chain variable region (VL), and the VL of the second antigen binding domain comprises an LCDR1 as set forth in SEQ ID NO: 336, an LCDR2 as set forth in SEQ ID NO: 337, and an LCDR3 as set forth in SEQ ID NO: 338, optionally wherein the second antigen binding domain comprises a VL as set forth in SEQ ID NO: 339.

[0381] In some embodiments, the bispecific antibody comprises a Fab, Fab’, Fv fragment,

[0382] F(ab)’2, scFv, di-scFv and / or dAb. Attorney Docket No.: 769273: SA9-943PC

[0383] In some embodiments, the bispecific antibody comprises the scFv of the first antigen binding domain or the scFv of the second antigen binding domain.

[0384] In some embodiments, the scFv of the first antigen-binding domain of the bispecific antibody comprises a sequence as set forth in any one of SEQ ID NOs: 340-345.

[0385] In some embodiments, the scFv of the first antigen-binding domain of the bispecific antibody comprises a sequence as set forth in SEQ ID NO: 342.

[0386] In some embodiments, the first antigen-binding domain of the bispecific antibody is selected from the combination of light chain and heavy chain of one of the following: a) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:350; b) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:351; c) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:352; d) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:353; e) a light chain of SEQ ID NO:349 and a heavy chain of SEQ ID NO:354; f) a light chain of SEQ ID NO:349 and a heavy chain of SEQ ID NO:355; g) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:356; h) a light chain of SEQ ID NO:372 and a heavy chain of SEQ ID NO:357; i) a light chain of SEQ ID NO:349 and a heavy chain of SEQ ID NO:358; j) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:359; k) a light chain of SEQ ID NO: 372 and a heavy chain of SEQ ID NO: 360; l) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:361; m) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:362; n) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:363; o) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:364; p) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:365; q) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:366; r) a light chain of SEQ ID NO:348 and a heavy chain as set forth in SEQ ID NO:378. Attorney Docket No.: 769273: SA9-943PC

[0387] In some embodiments, the first antigen-binding domain of the bispecific antibody comprises HCDR1 as set forth in SEQ ID NO: 2, HCDR2 as set forth in SEQ ID NO: 6, HCDR3 as set forth in SEQ ID NO: 15, LCDR1 as set forth in SEQ ID NO: 42, LCDR2 as set forth in SEQ ID NO: 43, and LCDR3 as set forth in SEQ ID NO: 44; and, the second antigen-binding domain of the bispecific antibody comprises HCDR1 as set forth in SEQ ID NO: 332, HCDR2 as set forth in SEQ ID NO: 333, HCDR3 as set forth in SEQ ID NO: 334, LCDR1 as set forth in SEQ ID NO: 336, LCDR2 as set forth in SEQ ID NO: 337, and LCDR3 as set forth in SEQ ID NO: 338.

[0388] In some embodiments, the first antigen-binding domain of the bispecific antibody comprises a VH as set forth in SEQ ID NO: 379 and a VL as set forth in SEQ ID NO: 380; and the second antigen-binding domain of the bispecific antibody comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339.

[0389] In some embodiments;

[0390] (1) the first antigen-binding domain of the bispecific antibody comprises an scFv as set forth in SEQ ID NO: 340; and the second antigen-binding domain of the bispecific antibody comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,

[0391] (2) the first antigen-binding domain of the bispecific antibody comprises an scFv as set forth in SEQ ID NO: 341; and the second antigen-binding domain of the bispecific antibody comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,

[0392] (3) the first antigen-binding domain of the bispecific antibody comprises an scFv as set forth in SEQ ID NO: 342; and the second antigen-binding domain of the bispecific antibody comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,

[0393] (4) the first antigen-binding domain of the bispecific antibody comprises an scFv as set forth in SEQ ID NO: 343; and the second antigen-binding domain of the bispecific antibody comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or, Attorney Docket No.: 769273: SA9-943PC

[0394] (5) the first antigen binding domain of the bispecific antibody comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106; and the second antigen binding domain of the bispecific antibody comprises an scFv as set forth in 346; or,

[0395] (6) the first antigen binding domain of the bispecific antibody comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106; and the second antigen binding domain of the bispecific antibody comprises an scFv as set forth in SEQ ID NO:347.

[0396] In some embodiments, the bispecific antibody includes a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 363.

[0397] In another aspect, the application provides an isolated nucleic acid molecule encoding a bispecific antibody as described herein.

[0398] In another aspect, the application provides an expression vector comprising an isolated nucleic acid molecule as described herein.

[0399] In another aspect, the application provides a host cell including an isolated nucleic acid molecule and / or an expression vector as described herein.

[0400] In another aspect, the application provides a composition comprising a bispecific antibody as described herein and optionally pharmaceutical acceptable carrier.

[0401] In another aspect, the application provides a preparation method for bispecific antibodies as described herein, comprising culturing a host cell as described herein, and recovering the bispecific antibodies.

[0402] In another aspect, the application provides a method of treating a disease or condition comprising administering a bispecific antibody, an isolated nucleic acid molecule, an expression vector, a host cell or a composition as described herein to a subject.

[0403] In another aspect, the application provides a bispecific antibody as described herein for use in the treatment of a disease or condition.

[0404] In another aspect, the application provides an isolated nucleic acid molecule, an expression Attorney Docket No.: 769273: SA9-943PC vector, a host cell or a composition as described herein for use in the treatment of a disease or condition.

[0405] In another aspect, the application provides a bispecific antibody, an isolated nucleic acid molecule, an expression vector, a host cell or a composition as described herein for the treatment of disease or condition or in the preparation of a drug for the treatment of disease or condition.

[0406] In some embodiments, the disease or the condition is a TL1 A-mediated disease or condition.

[0407] In some embodiments, the TL1 A-mediated disease or condition is inflammatory disease, optionally wherein the TL1 A-mediated inflammatory disease is selected from one of the following: Non-limiting examples of inflammatory disease include, allergy, ankylosing spondylitis, asthma, atopic dermatitis, autoimmune diseases or disorders, cancer, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, diabetes (e.g., type 1 diabetes and type 2 diabetes), glomerulonephritis, gout, hepatitis (e.g., active hepatitis), an immune-mediated disease or disorder, inflammatory bowel disease (IBD) such as Crohn's disease and ulcerative colitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), multiple sclerosis, neurodegenerative diseases of aging, periodontal disease (e.g., periodontitis), preperfusion injury transplant rejection, psoriasis, pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis), rheumatic disease, scleroderma, sinusitis, tuberculosis.

[0408] In some embodiments, the TL1 A-mediated disease or condition is autoimmune disease, optionally wherein the TLlA-mediated autoimmune diseases are selected from one of the following: achalasia, Addison's disease, Adult Stil's disease, agammaglobulinemia, alopecia areata, amyloidosis, ankylosing spondylitis, anti-GBM / anti-TBM nephritis, antiphospholipid syndrome, autoimmune angioedema, autoimmune familial autonomic dysfunction, autoimmune encephalomyelitis, autoimmune hepatitis, autoimmune inner ear disease (AIED), autoimmune myocarditis, autoimmune oophoritis, autoimmune orchitis, autoimmune pancreatitis, autoimmune retinopathy, autoimmune urticaria, axonal and neuronal neuropathy (AMAN), Barlow disease (Balddisease), Behcet's disease, benign mucous membrane pemphigoid, bullous pemphigoid, Castleman's disease (Castleman disease (CD), celiac disease, Chagas disease, chronic inflammatory demyelinating polyneuropathy (CIDP), Attorney Docket No.: 769273: SA9-943PC chronic recurrent multifocal osteomyelitis (CRMO), Churg-Strauss syndrome (CSS) or eosinophilic granulomatous disease (EGPA), cicatricial pemphigoid, and Cogan's syndrome), cold agglutinin disease, congenital heart block, coxsackienew myocarditis, CREST syndrome, Crohn's disease, dermatitis herpetiformis, dermatomyositis, Devic's disease (neuromyelitis optica), discoid lupus, Dressier's syndrome, endometriosis, eosinophilic esophagitis (EoE), eosinophilic fasciitis, erythema nodosum, primary mixed cryoglobulinemia, Evans syndrome, fibromyalgia, fibrotic alveolitis, giant cell arteritis (temporal arteritis), giant cell myocarditis, glomerulonephritis, Goodpasture's syndrome, granulomatosis with polyangiitis, Graves' disease, Guillain-Barre syndrome syndrome), Hashimoto's thyroiditis, hemolytic anemia, Henoch-Schonlein purpura (HSP), herpes gravilens or pemphigoid gestationum (PG), hidradenitis suppurativa (HS) (paradoxical acne), hypogammaglobulinemia, IgA nephropathy, IgG4-associated sclerotic disease, immune thrombocytopenic purpura (ITP), Inclusion body myositis (IBM), interstitial cystitis (IC), juvenile arthritis, juvenile diabetes mellitus (type 1 diabetes), juvenile myositis (JM), Kawasaki disease, Lambert-Eaton syndrome, leukocytoclastic vasculitis, lichen planus, lichen sclerosus, lignoconjunctivitis, linear IgA disease (LAD), lupus, chronic Lyme disease ( Lyme disease chronic), Meniere's disease, microscopic polyangiitis (MPA), mixed connective tissue disease (MCTD), Mooren's ulcer, Mucha-Habermann disease, multifocal motor neuropathy (MMN) or MMNCB, multiple sclerosis, myasthenia gravis, myositis, narcolepsy, neonatal lupus, neuromyelitis optica, neutropenia, ocular cicatricial pemphigoid, optic neuritis, recurrent rheumatism (PR), PandAS, paraneoplastic cerebellar degeneration (PCD), paroxysmal nocturnal hemoglobinuria (PNH), Parry Romberg syndrome, ciliary planitis (peripheral uveitis), Parsonage-Turner syndrome), pemphigus, peripheral neuropathy, perivenous encephalomyelitis, pernicious anemia (PA), POEMS syndrome, polyarteritis nodosa, polyglandular syndrome type I, II, type III, polymyalgia rheumatica, polymyositis, post-myocardial infarction syndrome, postpericardiotomy syndrome, primary biliary cirrhosis, primary sclerosing cholangitis, progesterone dermatitis, psoriasis, psoriatic arthritis, pure red blood cell aplasia (PRCA), pyoderma gangrenosum, Raynaud's phenomenon, reactive arthritis, reflex sympathetic dystrophy, relapsing polychondritis, restless legs syndrome (RLS), retroperitoneal fibrosis, rheumatic fever, rheumatoid arthritis, sarcoidosis, Attorney Docket No.: 769273: SA9-943PC

[0409] Schmidt syndrome, scleritis, scleroderma, Sjogren's syndrome, sperm and testicular autoimmunity, stiff-person syndrome (SPS), subacute bacterial endocarditis (SBE), Susac's syndrome syndrome), sympathetic ophthalmia (SO), Takayasu's arteritis, temporal arteritis / giant cell arteritis, thrombocytopenic purpura (TTP), Tolosa-Hunt syndrome (THS), transverse myelitis, type 1 diabetes mellitus, ulcerative colitis (UC), undifferentiated connective tissue disease (UCTD), uveitis, vasculitis, Vitiligo and Vogt-Koyanagi-Harada disease.

[0410] In some embodiments, the TL1 A-mediated disease or condition is cancer, optionally wherein

[0411] TL1 A-mediated cancers are selected from one or more of the following: non-restrictive examples of cancer include adenoid cystic carcinoma, adrenal carcinoma, amyloidosis, anal cancer, ataxia telangiectasia, atypical nevus syndrome, basal cell carcinoma, cholangiocarcinoma, Birt Hogg Dube syndrome, bladder cancer, bone cancer, brain tumor, breast cancer, male breast cancer, carcinoid tumor, cervical cancer, colorectal cancer, ductal cancer, Endometrial Cancer, Esophageal Cancer, Gastric Cancer, Gastrointestinal Stromal Tumor (GIST), HER2 -Positive Breast Cancer, Pancreatic Islet Cell Tumor, Juvenile Polyposis Syndrome, Kidney Cancer, Laryngeal Cancer, Leukemia-Acute Lymphoblastic Leukemia, Acute

[0412] Lymphoblastic Leukemia (ALL), Acute Myeloid Leukemia AML,

[0413] Adult Leukemia, Childhood Leukemia, 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's lymphoma (Lymphoma- Hodgkin's), non Hodgkin's lymphoma (Lymphoma-Non-Hodgkin’s), 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 tumors, polycythemia vera, prostate cancer, renal cell carcinoma, retinoblastoma, salivary gland carcinoma, sarcoma, Sarcoma- Attorney Docket No.: 769273: SA9-943PC

[0414] Kaposi, skin cancer, small bowel cancer, stomach cancer, testicular cancer, thymoma, thyroid cancer, uterine (endometrial) cancer, vaginal cancer, and Wilms’ Tumor.

[0415] Other aspects and advantages of the present application will become readily apparent to those skilled in the art from the following detailed description. Only exemplary embodiments of the present application are shown and described in the following detailed description. As will be recognized by those skilled in the art, the contents of the present application enable those skilled in the art to make modifications to the specific embodiments disclosed without departing from the spirit and scope of the present application to which the present application pertains. Accordingly, the drawings and description herein are to be regarded as illustrative in nature, and not as restrictive.

[0416] BRIEF DESCRIPTION OF THE DRAWINGS

[0417] Specific features of the present application to which the present application relates are as set forth in the appended claims. The features and advantages of the present application to which the present application relates will be better understood by reference to the exemplary embodiments and drawings described in detail below. A brief description of the drawings is as follows:

[0418] Figure 1 shows a schematic diagram of the bi specific antibody configuration described in this application;

[0419] Figure 2 shows the predicted structure of the TLA0015 monoclonal antibody and TL1A antigen complex in this application;

[0420] Figure 3 shows the ELISA test results of the binding of different bispecific antibodies described in this application to human TL1 A-his protein;

[0421] Figures 4A-4C show the binding and dissociation curves of different anti-TLl A antibodies described in this application to human TL1 A-his protein;

[0422] Figures 5A-5C show the binding and dissociation curves of different anti-TLl A antibodies described in this application to monkey TL1 A-his protein; Attorney Docket No.: 769273: SA9-943PC

[0423] Figures 6A-6B show the binding and dissociation curves of the antibodies described in this application to human a4p7 protein, isolation curve;

[0424] Figure 7A-7B shows that the bispecific antibody described in this application blocks the binding of TLlA-his protein to HEK-293-DR3 cells;

[0425] Figure 8A-8D shows the effect of a4p7 protein on the blocking activity of the bispecific antibody described in the application;

[0426] Figure 9 shows that the bispecific antibody described in this application inhibits the activity of Caspase3 / 7 activated by TL1A;

[0427] Figure 10 shows that the bispecific antibody described in this application inhibits the NF- KB signaling pathway activated by TLlA-his protein;

[0428] Figure 11 shows the ELISA test results of the binding of different bispecific antibodies described in this application to a4p7 protein;

[0429] Figure 12 shows the binding of the bispecific antibody described in this application to Hut 78 cell binding curve;

[0430] Figure 13 shows the bispecific antibody described in the present application blocking the binding of a4p7 and ligand MAdCAM-1;

[0431] Figure 14 shows the bispecific antibody described in the present application blocking MAdCAM-1 -mediated cell adhesion;

[0432] Figure 15 shows the effect of TL1A protein on the bispecific antibody described in the present application blocking cell adhesion function;

[0433] Figures 16A-16B show the endocytic activity of different bispecific antibodies described in the present application on HuT 78 cells;

[0434] Figures 17A-17D show the effect of adding TL1 A protein on the endocytic activity of the bispecific antibody described in the present application on HuT 78 cells and primary T cells;

[0435] Figures 18 and 19 show the endocytic activity of different bispecific antibodies described in the present application on HuT 78 cells;

[0436] Figure 20 shows the bispecific antibody described in the present application downregulating HuT 78 cell membrane, and after adding TL1A protein, the endocytosis Attorney Docket No.: 769273: SA9-943PC activity increased, which can more obviously down-regulate the expression of a4p7 on the membrane;

[0437] Figure 21A-Figure 21B show that the bispecific antibody described in this application down-regulates the total expression of a4p7 in HuT 78 cells, and after adding TL1 A protein, the endocytosis activity increased, which can more obviously down-regulate the total expression of a4p7;

[0438] Figure 22A shows the recovery of a4p7 on the HuT 78 cell membrane at different times after endocytosis (antibody concentration 5nM);

[0439] Figure 22B shows the proportion of positive cells that recover the expression of a4p7 on the HuT 78 cell membrane at different times after endocytosis (antibody concentration 5nM);

[0440] Figure 23 A shows the recovery of a4p7 on the HuT 78 cell membrane at different times after endocytosis (antibody concentration 50nM);

[0441] Figure 23B shows the recovery of a4p7 on the HuT 78 cell membrane at different times after endocytosis (antibody concentration 50nM); 78 The proportion of positive cells that restored expression of a4p7 on the cell membrane at different times (antibody concentration 50nM);

[0442] Figure 24A shows the restoration of expression of a4p7 on the PanT cell membrane at different times after endocytosis (antibody concentration 5nM);

[0443] Figure 24B shows the proportion of positive cells that restored expression of a4p7 on the PanT cell membrane at different times after endocytosis (antibody concentration 5nM);

[0444] Figure 25 shows the inhibitory activity of the antibody on PanT cell migration;

[0445] Figure 26 shows the endocytosis activity of the antibody in PanT cells (Donorl);

[0446] Figures 27A-27B show the endocytosis of the antibody, The recovery of a4p7 expression on the membrane of PanT cells at different times (Donorl);

[0447] Figures 28A-28B show the migration of PanT cells at different times (Donorl);

[0448] Figure 29 shows the endocytosis activity of antibodies in PanT cells (Donor2);

[0449] Figures 30A-30B show the endocytosis of antibodies and the recovery of a4p7 expression on the membrane of PanT cells at different times (Donor2); Attorney Docket No.: 769273: SA9-943PC

[0450] Figures 31A-31B show the migration of PanT cells at different times (Donor2);

[0451] Figure 32A shows the inhibitory activity of the antibodies disclosed in this application on CD4 T cell activation;

[0452] Figure 32B shows the inhibitory activity of the antibodies disclosed in this application on CD4 T cell proliferation;

[0453] Figure 32C shows the inhibitory activity of the antibodies on CD4 T cell activation and proliferation inhibitory activity;

[0454] Figure 33 shows the inhibitory activity of the bispecific antibody described in this application on IFN-y secretion;

[0455] Figures 34A-34C show the killing effect of the bispecific antibody described in this application on PBMC under different conditions;

[0456] Figures 35A-35F show the activation of different T cells after the bispecific antibody described in this application is co-incubated with CHO-K1 cells and PBMC antibodies;

[0457] Figures 36A-36F show the activation of different T cells after the bispecific antibody described in this application is co-incubated with CHO-K1-TL1 A cells and PBMC antibodies;

[0458] Figures 37A-37F show the activation of different T cells after the bispecific antibody described in this application is co-incubated with PBMC antibodies;

[0459] Figures 38A-38E show the activation of different T cells after the bispecific antibody described in this application is co-incubated with CHO-K1 cells

[0460] Figure 39A-39E shows the release of different cytokines after the bispecific antibody described in this application is co-incubated with CHO-K1-TL1 A cells and PBMC antibodies;

[0461] Figure 40 shows the PK detection of TLb0045 described in this application in rats;

[0462] Figure 41 shows the PK detection of TLb0065 described in this application in rats;

[0463] Figure 42 shows the PK detection of TLb0045 described in this application and TLb0065 described in this application in rats;

[0464] Figure 43 shows the PK detection of TLb0045 described in this application in hFcRn mice;

[0465] Figure 44 shows the PK detection of TLb0045-l described in this application in hFcRn mice. Attorney Docket No.: 769273: SA9-943PC

[0466] Figures 45A-45B show the weight changes of the antibody in the DSS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment;

[0467] Figure 46 shows the daily disease score of the antibody in the DSS-induced hTLl A / ha4p7 transgenic mouse enteritis efficacy experiment;

[0468] Figures 47A-47B show the colorectal length changes of the antibody in the DSS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment;

[0469] Figures 48A-48B show the colorectal weight changes of the antibody in the DSS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment;

[0470] Figures 49A-49B show the weight changes of the antibody in the TNBS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment;

[0471] Figure 50 shows the colorectal length changes of the antibody in the TNBS-induced hTLIA / Daily disease scores in the ha4p7 transgenic mouse enteritis efficacy experiment;

[0472] Figure 51 shows the mouse survival curve results in the TNBS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment;

[0473] Figures 52A-52B show the changes in colorectal length in the TNBS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment;

[0474] Figures 53A-53B show the changes in colorectal weight in the TNBS-induced hTLlA / ha4p7 transgenic mouse enteritis efficacy experiment.

[0475] DETAILED DESCRIPTION OF THE INVENTION

[0476] The embodiments of the invention of the present application are illustrated by particular examples. Other advantages and effects of the present application will become readily apparent to those skilled in the art from disclosure of the present specification.

[0477] Definition of Terms

[0478] In the present application, the term “isolated” generally refers to a product obtained from a natural state by artificial means. If an “isolated” substance or component occurs in nature, it may be altered from its natural environment, or the substance may be isolated from its natural environment, or both. For example, an unisolated polynucleotide or polypeptide naturally Attorney Docket No.: 769273: SA9-943PC occurs in a living animal and the same polynucleotide or polypeptide isolated from its natural state in high purity is the to be isolated. The term “isolated” does not exclude the admixture of artificial or synthetic substances, nor the presence of other impure substances which do not affect the activity of the substance.

[0479] In the present application, the term “antigen-binding protein” generally refers to a polypeptide molecule capable of specifically recognizing and / or neutralizing a particular antigen. In the present application, the term “antigen-binding protein” may include an “antibody” or an “antigen-binding fragment”. For example, the antibody may comprise an immunoglobulin composed of at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds, and may include any molecule comprising an antigenbinding portion thereof. The term “antibody” may include monoclonal antibodies, antibody fragments, or antibody derivatives, including, but not limited to, murine antibodies, human antibodies (fully human antibodies), humanized antibodies, chimeric antibodies, single chain antibodies (e.g., scFv), and antibody fragments that bind to an antigen (e.g., Fab, Fab ' , and (Fab)2 fragments). The term “antibody” may also include all recombinant forms of antibodies, such as antibodies expressed in prokaryotic cells, non-glycosylated antibodies, and any antigen-binding antibody fragments and derivatives thereof described herein. Each heavy chain may be composed of a heavy chain variable region and a heavy chain constant region. Each light chain may be composed of a light chain variable region and a light chain constant region. The VH and VL regions can be further distinguished as hypervariable regions called complementarity determining regions (CDRs), interspersed with more conserved regions called framework regions (FRs). Each VH and VL may be composed of three CDRs and four FRs, which may be arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The variable regions of the heavy and light chains contain binding domains that interact with an antigen. The constant region of an antibody can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component of the classical complement system (Clq). The exact boundaries of the CDRs have been defined differently Attorney Docket No.: 769273: SA9-943PC for different systems. The system described by Kabat (Kabat et al., Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987) and (1991)) provides not only an unambiguous residue numbering system applicable to any variable region of an antigen-binding fragment, but also provides precise residue boundaries defining CDRs. These CDRs may be referred to as Kabat CDRs. Chothia and co-workers (Chothia and Lesk, J. Mol. Biol. 196: 901-917(1987) and Chothia et al., Nature 342: 877-883(1989)) found that certain sub-portions within Kabat CDRs adopt nearly identical conformations in the backbone of a peptide despite the large diversity at the amino acid sequence level. These sub-portions are designated as LI, L2, and L3 or Hl, H2, and H3, where “L” and “H” refer to the light chain and heavy chain regions, respectively. These regions may be referred to as Chothia CDRs, which have boundaries that overlap with Kabat CDRs. Other boundaries that define CDRs and overlap with Kabat CDRs have been described by Padlan (FASEB J. 9: 133-139 (1995)) and MacCallum (J Mol Biol 262(5): 732-45(1996)). In addition, other CDR boundary definitions may not strictly follow one of the above systems, but will still overlap with Kabat CDRs, although they may be shortened or lengthened in light of prediction or experimental findings that a particular residue or group of residues or even the entire CDRs, do not significantly affect the antigen binding. In the present application, said CDR may be defined through the Kabat numbering system.

[0480] In the present application, the term “antigen-binding fragment” generally refers to one or more fragments of an antibody that function to specifically bind to an antigen. The antigenbinding function of an antibody can be achieved by a full-length fragment of the antibody. The antigen-binding function of an antibody may also be achieved by: a heavy chain comprising a fragment of Fv, scFv, dsFv, Fab, Fab ' or F (ab ' )2, or a light chain comprising a fragment of Fv, scFv, dsFv, Fab, Fab ' , or F(ab ' )2. (1) An Fab fragment, i.e., a monovalent fragment consisting of the VL, VH, CL, and CH domains; (2) an F(ab ' )2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bond at the hinge region; (3) an Fd fragment consisting of the VH and CH domains; (4) an Fv fragment consisting of the VL and VH domains of a single arm of an antibody; (5) a dAb fragment consisting of VH domains Attorney Docket No.: 769273: SA9-943PC

[0481] (Ward et al., (1989) Nature 341 : 544-546); (6) an isolated complementarity determining region (CDR), and (7) a combination of two or more isolated CDRs which may optionally be joined by a linker. For example, monovalent single chain molecules Fv (scFv) formed by the pairing of VL and VH may also be included (see, Bird et al., (1988) Science 242: 423-426; and Huston et al., (1988) Proc. Natl. Acad. Sci. 85: 5879-5883). For example, a class of antibody VHH lacking the antibody light chain but only the heavy chain variable region can also be included (see, e.g., Kang Xiaozhen et al., Chinese Journal of Biotechnology, 2018, 34 (12): 1974-1984). The “antigen-binding portion” may also include an immunoglobulin fusion protein comprising a binding domain selected from the group consisting of: (1) a binding domain polypeptide fused to an immunoglobulin hinge region polypeptide; (2) an immunoglobulin heavy chain CH2 constant region fused to the hinge region; and (3) an immunoglobulin heavy chain CH3 constant region fused to the CH2 constant region.

[0482] The basic 4-chain antibody unit is a heterotetrameric glycoprotein composed of two identical light (L) chains and two identical heavy (H) chains. IgM antibodies consist of 5 basic heterotetrameric units and another polypeptide called the J chain and contain 10 antigen binding sites, while IgA antibodies consist of 2-5 basic 4-chain units, which can polymerize to form a multivalent assembly bound to the J chain. In the case of IgG, the 4-chain unit is usually about 150,000 Daltons. Each L chain is linked to the H chain by one covalent disulfide bond, while the two H chains are linked to each other by one or more disulfide bonds, depending on the isotype of the H chain. Each H and L chain also has regularly spaced intrachain disulfide bridges. Each H chain has a variable domain (VH) at the N-terminus, followed by three constant domains (CH) in each a and y chain, and four CH domains in the p and a isotypes. Each L chain has a variable domain (VL) at the N-terminus and then a constant domain at the other end. VL is aligned with VH, and CL is aligned with the first constant region (CHI) of the heavy chain. Specific amino acid residues are believed to form an interface between the light chain variable domain and the heavy chain variable domain. The pairing of VH and VL together forms a single antigen binding site. For the structure and properties of different classes of antibodies, see, for example, Basic and Clinical Immunology, 8th edition, Daniel P. Sties, Abba Attorney Docket No.: 769273: SA9-943PC

[0483] I. Terr and Tristram G. Parsolw (eds.), Appleton & Lange, Norwalk, Conn., 1994, page 71 and Chapter 6. The L chains of any vertebrate species can be divided into two clearly distinct types, called kappa and lambda, based on the amino acid sequence of their constant regions. Immunoglobulins can be divided into different classes or isotypes based on the amino acid sequence of their heavy chain constant domain (CH). There are five classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, with heavy chains designated a, 5, a, y, and p, respectively. Based on relatively minor differences in the sequence and function of CH, the gamma and alpha classes are further divided into subclasses, for example, humans express the following subclasses: IgGl, IgG2A, IgG2B, IgG3, IgG4, IgAl, and IgKl.

[0484] In the present application, the term “monoclonal antibody” generally refers to a population of substantially homologous antibodies, i.e., the individual antibodies comprising the population are identical except for possible naturally occurring mutations that may be present in minor amounts. Monoclonal antibodies are highly specific, being directed against a single antigenic site. For example, the monoclonal antibodies can be prepared by hybridoma techniques or can be produced in bacteria, eukaryotic animal or plant cells using recombinant DNA methods, or can be derived from phage antibody libraries using the techniques, for example, described in Clackson et al., Nature, 352:624-628 (1991) and Marks et al., Mol. Biol., 222:581-597 (1991).

[0485] In the present application, the term “chimeric antibody” generally refers to an antibody in which a portion of the amino acid sequence of each heavy or light chain is homologous to the corresponding amino acid sequence in an antibody from a particular species, or belongs to a particular class, while the remaining segments of the chain are homologous to the corresponding sequence in another species. For example, the variable regions of both the light and heavy chains are derived from the variable region of an antibody of one animal species (e.g., mouse, rat, etc.), while the constant portions are homologous to antibody sequences from another species (e.g., human). For example, to obtain chimeric antibodies, non-human-derived B cells or hybridoma cells can be used to generate variable regions, while which the constant regions in combination therewith are of human origin. The variable region has the advantage Attorney Docket No.: 769273: SA9-943PC of being easy to prepare and its specificity is not influenced by the origin of the constant region with which it is combined. Also, since the constant region of the chimeric antibody can be derived from human, the chimeric antibodies are less likely to elicit an immune response upon injection than using antibodies whose constant regions are of non-human origin.

[0486] In the present application, the term “humanized antibody” generally refers to a chimeric antibody that contains fewer sequences from non-human immunoglobulins, thereby reducing the immunogenicity of a xenogenous antibody when introduced into humans, while maintaining the full antigen-binding affinity and specificity of the antibody. For example, non- human binding domains can be humanized using the technical means, such as CDR transplantion (Jones et al., Nature 321 :522(1986)) and variants thereof; including “reshaping”, (Verhoeyen, et al., 1988 Science 239: 1534-1536; Riechmann, et al., 1988 Nature 332:323-337; Tempest, et al., Bio / Technol 1991 9:266-271), “hyperchimerization” (Queen, et al., 1989 Proc Natl Acad Sci USA 86: 10029-10033; Co, et al., 1991 Proc Natl Acad Sci USA 88:2869-2873; Co, et al., 1992 J Immunol 148: 1149-1154), and “veneering” (Mark, et al., “Derivation of therapeutically active humanized and veneered anti-CD18 antibodies.” In: Metcalf B W, Dalton B J, eds. Cellular adhesion: molecular definition to therapeutic potential. New York: Plenum Press, 1994: 291-312), and resurfacing (U.S. Pat. No. 5,639,641). Other regions, such as hinge and constant region domains, may also be humanized if they are also derived from non-human sources.

[0487] In the present application, the term “murine antibody” generally refers to an antibody in which the variable region framework and CDR regions are derived from mouse germline immunoglobulin sequences. In addition, if the antibody comprises constant regions, which are also derived from mouse germline immunoglobulin sequences. The murine antibodies of the present application may comprise amino acid residues not encoded by the mouse germline immunoglobulin sequences, such as mutations introduced by random or point mutations in vitro or by somatic mutation in vivo.

[0488] In the present application, the term "directly linked" is used in contrast to the term indirect linked", which usually means a direct connection. For example, said direct connection Attorney Docket No.: 769273: SA9-943PC may be a direct connection between substances without spacers. Said spacer may be a linker. For example, said linker may be a peptide linker. The term "indirect linkage" generally refers to the case where there is no direct linkage between substances. For example, said indirect linkage may be a linkage through a spacer. For example, in the isolated antigen-binding proteins described in the present application, the C-terminal end of said L-FR1 and the N- terminal end of said LCDR1 may be directly or indirectly linked.

[0489] In this application, the term "TL1A" refers to TNF-like protein 1A. TL1A (TNFSF15) is a member of the TNF family that is expressed predominantly by endothelial cells, macrophages and dendritic cells (DCs). Its expression is induced by immune complexes (ICs) and cytokines. TL1 A receptor DR3 is expressed mainly on T cells and NKT cells. In vitro, TL1A has been shown to enhance T cell proliferation and cytokine production in both humans and mice. In vivo, TL1 A transgenic mice produce an IBD phenotype similar to human Crohn's disease. In addition, treatment with recombinant TL1 A protein exacerbated colitis in mdrl- / - mice. The TL1 A described in this application may be a monomer or trimer of TLlA.The TL1A described in this application can be a human, mouse or monkey TL1 A.

[0490] The cell surface molecule "a4p7 integrin" or "a4p7" is a heterodimer formed by the a4 chain (CD49D, ITGA4) and the P7 chain (ITGB7). a4 and P7 chains also form a heterodimer a4pi or aEp7 with the alternative integrin chains pi or aE, respectively. aE to form a heterodimer a4pi or aEp7. human a4 and P7 genes (GenBank (National Center for Biotechnology Information, Bethesda, MD) RefSeq Nos. NM_000885 and NM_000889, respectively) by B and T lymphocytes, especially memory CD4+ lymphocytes. As typical of many integrins, a4p7 can exist in either a quiescent or activated state, ligands for a4p7 include vascular cell adhesion molecule (VCAM), fibronectin, and mucosal addressin (MAdCAM (e.g., MAdCAM-1)). The a4p7 described in the present application may be human, murine or monkey a4p7.

[0491] In this application, the term "scFv" generally refers to a fusion protein comprising at least one antibody fragment comprising a light chain variable region and at least one antibody fragment comprising a heavy chain variable region, wherein said light chain and heavy chain Attorney Docket No.: 769273: SA9-943PC variable regions are adjacent (e.g., via synthetic junctions, such as short flexible peptide junctions) and are capable of being expressed as a single-chain polypeptide, and said scFv retains the specificity of the intact antibody from which it is derived. Unless specifically stated, as used in this application, an scFv may have said VL and VH variable regions in any order (e.g., relative to the N-terminus and the C-terminus of the polypeptide), and an scFv may comprise a VL-linker-VH or may comprise a VH-linker-VL.

[0492] In this application, the terms "specific binding" or "specific" generally refer to measurable and reproducible interactions, such as binding between a target and an antibody, that may determine the presence of a target in the presence of a heterogeneous population of molecules, including biomolecules. For example, an antibody that specifically binds a target (which may be an epitope) may be an antibody that binds that target with greater affinity, affinity, more readily, and / or for a greater duration than it binds other targets. In some embodiments, the antibody specifically binds an epitope on a protein, said epitope being conserved across different species of proteins. In certain embodiments, specific binding may include, but is not required to exclusively bind.

[0493] In addition to the specific proteins and nucleotides mentioned herein, the present application may also include functional variants, derivatives, analogs, homologs, and fragments thereof.

[0494] The term “functional variant” refers to an amino acid sequence that is substantially identical to a naturally occurring sequence or a polypeptide encoded by a substantially identical nucleotide sequence and capable of having one or more activities of the naturally occurring sequence. In the context of the present application, a variant of any given sequence refers to a sequence in which the particular sequence of residues, whether amino acid or nucleotide residues, has been modified such that the polypeptide or polynucleotide substantially retains at least one endogenous function. Variant sequences may be obtained by the addition, deletion, substitution, modification, substitution and / or variation of at least one amino acid residue and / or nucleotide residue present in a naturally occurring protein and / or polynucleotide, so long as the original functional activity is retained. Attorney Docket No.: 769273: SA9-943PC

[0495] In the present application, the term “derivative” generally refers to a polypeptide or polynucleotide of the present application comprising any substitution, variation, modification, replacement, deletion and / or addition of one (or more) amino acid residues from / on the sequence, so long as the resulting polypeptide or polynucleotide substantially retains at least one of its endogenous functions.

[0496] In the present application, the term “analog” generally refers to a polypeptide or polynucleotide including any mimetic of a polypeptide or polynucleotide, i.e., a chemical compound that possesses at least one endogenous function of the polypeptide or polynucleotide that the mimetic mimics.

[0497] Generally, amino acid substitutions, e.g., at least one (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 20 or more) amino acid substitution can be made, so long as the modified sequence substantially retains the desired activity or ability. Amino acid substitutions can include the use of non-naturally occurring analogs.

[0498] In the present application, the term “homologue” generally refers to an amino acid sequence or nucleotide sequence having certain homology to a naturally occurring sequence. The term “homology” can be equivalent to sequence “identity”. Homologous sequences may include amino acid sequences that may be at least 80%, 85%, 90%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9% identical to the subject sequence. Typically, the homologue will comprise the same active site or the like as the subject amino acid sequence. Homology may be considered in terms of similarity (i.e., amino acid residues having similar chemical properties / functions) or may be expressed in terms of sequence identity. In the present application, a sequence having percent identity in any one of the SEQ ID NO of a mentioned amino acid sequence or nucleotide sequence refers to a sequence having the percent identity over the entire length of the mentioned SEQ ID NO. To determine sequence identity, sequence alignments may be performed by a variety of ways known to those skilled in the art, e.g., using BLAST, BLAST-2, ALIGN, NEEDLE, or Megalign (DNASTAR) software, etc. Those skilled in the art can determine the appropriate parameters for the alignment, including any algorithm required to achieve the optimal alignment over the full length of the sequence being compared. Attorney Docket No.: 769273: SA9-943PC

[0499] The proteins or polypeptides used in the present application may also have deletions, insertions, or substitutions of amino acid residues which produce silent changes and result in functionally equivalent proteins. Intentional amino acid substitutions may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, and / or amphipathic nature of the residues, so long as the endogenous function is retained. For example, negatively charged amino acids include aspartic acid and glutamic acid; positively charged amino acids include lysine and arginine; and amino acids containing uncharged polar head groups with similar hydrophilicity values include asparagine, glutamine, serine, threonine, and tyrosine.

[0500] In the present application, the term “immunoconjugate” generally refers to a conjugate formed by the conjugation of other agents (e.g., a chemotherapeutic agent, a radioactive element, a cytostatic agent, and a cytotoxic agent) to The isolated antigen-binding protein (e.g., via covalent attachment of a linking molecule), wherein the conjugate can deliver the other agents to a target cell (e.g., a tumor cell) via specific binding of The isolated antigen-binding protein to an antigen on the target cell. The immunoconjugate then undergoes such internalization and eventually enters the interior of the target cell (e.g., into vesicles such as a lysosome), at which point the linker molecule in the immunoconjugate can be cleaved to release the other agent, thereby exerting its cytotoxic effect. In addition, the antigen may also be secreted by the target cell and located in the space outside the target cell.

[0501] In the present application, the term “subject” generally refers to human or non-human animals, including, but not limited to, cats, dogs, horses, pigs, cows, caprid, rabbits, mice, rats, or monkeys.

[0502] In the present application, the term “nucleic acid molecule” generally refers to an isolated form of nucleotides, deoxyribonucleotides, or ribonucleotides or analogs thereof, of any length, isolated from their natural environment or artificially synthesized.

[0503] In the present application, the term “vector” generally refers to a nucleic acid molecule capable of self-replication in a suitable host. The vector can transfer the inserted nucleic acid molecule into and / or between cells. The vector may include a vector for primarily inserting DNA or RNA into a cell, a vector for primarily replicating DNA or RNA, and a vector for Attorney Docket No.: 769273: SA9-943PC primarily expressing transcription and / or translation of DNA or RNA. The vector can be a polynucleotide capable of being transcribed and translated into a polypeptide when introduced into an appropriate cell. In general, the vector may produce the desired expression product by culturing an appropriate cell containing the vector. In the present application, the vector may include a lentiviral vector.

[0504] In the present application, the term “cell” generally refers to an individual cell, cell line or cell culture that may or may already contain a plasmid or vector comprising a nucleic acid molecule as described herein, or that is capable of expressing a polypeptide as described herein or an antigen-binding protein as described herein. The cell may include the progeny of a single cell. Due to natural, accidental, or deliberate mutations, the progeny cells may not necessarily be identical in morphology or in genome to the original parent cell, but are capable of expressing the polypeptide or antigen-binding protein as described herein. The cells can be obtained by transfecting cells in vitro with the vectors as described herein. The cells may be prokaryotic cells (e.g., E. coli) or eukaryotic cells (e.g., yeast cell, COS cells, Chinese Hamster Ovary (CHO) cells, HeLa cells, HEK293 cells, COS-1 cells, NSO cells, or myeloma cells). In some embodiments, the cells may be immune cells. For example, the immune cell may be selected from the group consisting of T cells, B cells, natural killer cells (NK cells), macrophages, NKT cells, monocytes, dendritic cells, granulocytes, lymphocytes, leukocytes and / or peripheral blood mononuclear cells.

[0505] In the present application, the term “treatment” generally refers to: (i) the prevention of the development of a disease, disorder, and / or condition in a patient who may be susceptible to, but has not yet been diagnosed with, that disease, disorder, or condition; (ii) the suppression of the disease, disorder, or condition, i.e., the curb of the development; and (iii) remission of the disease, disorder, or condition, i.e., causing regression of the disease, disorder, and / or condition and / or symptoms associated with the disease, disorder, and / or condition.

[0506] In the present application, the terms “polypeptide”, “peptide”, and “protein” are used interchangeably and generally refer to a polymer of amino acids of any length. The polymer may be linear or branched, and may comprise modified amino acids and it may be interrupted Attorney Docket No.: 769273: SA9-943PC by non-amino acids. These terms also encompass amino acid polymers that have been modified. These modifications may comprise: disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation (e.g., binding to a labeling component). The term “amino acid” includes natural and / or non-natural or synthetic amino acids, including glycine as well as the D and L optical isomers, as well as amino acid analogs and peptidomimetics.

[0507] In the present application, the terms “polynucleotide”, “nucleotide”, “nucleotide sequence”, “nucleic acid”, and “oligonucleotide” are used interchangeably and refer generally to polymeric forms of nucleotides of any length, such as deoxyribonucleotides or ribonucleotides, or analogs thereof. A polynucleotide may have any three-dimensional structure and may perform any function that is known or unknown. Non-limiting examples of polynucleotides are as follows: a coding or noncoding region of a gene or gene fragment, a plurality of loci (one locus) as defined by ligation analysis, exons, introns, messenger RNA (mRNA), transport RNA, ribosomal RNA, short interfering RNA (siRNA), short hairpin RNA (shRNA), micro-RNA (miRNA), ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes, and primers. A polynucleotide may comprise one or more modified nucleotides, such as methylated nucleotides and nucleotide analogs. If present, modification of the nucleotide structure may be performed before or after assembly of the polymer. The sequence of nucleotides may be interrupted by non-nucleotide components. The polynucleotides may be further modified after polymerization, e.g., by conjugation to labeled components.

[0508] In the present application, the term “KD” (likewise, “KD” or “KD”) generally refers to an “affinity constant” or an “equilibrium dissociation constant” and refers to a value obtained at equilibrium in a titration measurement, or by dividing the dissociation rate constant (kd) by the binding rate constant (ka). The binding affinity of a binding protein (e.g., an isolated antigenbinding protein as described herein) for an antigen is expressed using an association rate constant (ka), a dissociation rate constant (kd), and an equilibrium dissociation constant (KD). Attorney Docket No.: 769273: SA9-943PC

[0509] Methods for determining association and dissociation rate constants are well known in the art. The use of fluorescence-based techniques provides high sensitivity and the ability to examine samples at equilibrium in physiological buffers. For example, the KD value can be determined by Octet assay, and other experimental approaches and instruments such as BIAcore (Biomolecular Interaction Analysis) can be used (e.g., instruments available from BIAcorelnternationalAB, aGEHealthcarecompany, Uppsala, Sweden). Alternatively, the KD value can be determined using KinExA (dynamic exclusion assay (KineticExclusi on Assay)) available from Sapidynelnstruments (Boise, Idaho) or using a surface plasmon resonance (SPR) instrument.

[0510] In the present application, the term "IC50 value" or the term "half-maximal inhibitory concentration" (IC50) indicates the concentration of a particular compound that is required to achieve 50% inhibition of a biological process in vitro. The IC50 value can be logarithmically converted to a pIC50 value (-log IC50), where higher values indicate exponentially greater potency. The IC50 value is not an absolute value, but rather depends on experimental conditions, such as the concentration used. The IC50 value can be converted to an absolute inhibitory constant (Ki) using the Cheng-Prusoff equation (Biochem. Pharmacol. (1973) 22:3099).

[0511] The term "EC50", in the context of in vitro or in vivo analyses using isolated antigenbinding fragments, refers to the concentration of the antibody or antigen-binding portion thereof when the induced response is 50% of the maximal response, i.e., halfway between the maximal response and the baseline.

[0512] The terms "a4p7 inhibitor" and "a4p7 antagonist" are used interchangeably. Each is a molecule that detectably inhibits at least one function of a4p7. Conversely, an "a4p7 agonist" is a molecule that detectably enhances at least one function of a4p7.The inhibition caused by the a4p7 inhibitor need not be complete, as long as it is detectable by, for example, employing an assay. Any assay of a4p7 function may be employed, examples of which are provided herein. Examples of a4p7 functions that can be inhibited by a4p7 inhibitors (or enhanced by a4p7 agonists) include ligand binding (i.e., binding to MAdCAM-1), adhesion to cells expressing the ligand, transport to specific compartments (e.g., intestines), release of cytokines, Attorney Docket No.: 769273: SA9-943PC chemokines, and other mediators, exacerbation or worsening of inflammatory responses and tissue damage, and the like. a4p7 inhibitors and a4p7 agonists of the Examples of types include, but are not limited to, a4p7-binding polypeptides, such as antigen-binding proteins (e.g., a4p7 antigen-binding proteins), antibodies, antibody fragments, and antibody derivatives.

[0513] The terms "TL1A inhibitor" and "TL1A antagonist" are used interchangeably. Each is a molecule that detectably inhibits at least one function of TL1 A. Conversely, a "TL1A agonist" is a molecule that detectably enhances at least one function of TLlA.The inhibition caused by the TL1 A inhibitor need not be complete, as long as it is detectable by, for example, using an assay. Any assay for a4p7 function may be employed, examples of which are provided herein. Examples of functions of TL1 Athat can be inhibited by TL1 A inhibitors (or enhanced by TL1 A agonists) include ligand binding (i.e., binding to MAdCAM-1), adhesion to cells expressing the ligand, transport to specific compartments (e.g., intestines), release of cytokines, chemokines, and other mediators, exacerbation or worsening of inflammatory responses and tissue damage, and the like. The TL1 A inhibitors and the TL1 A agonists of the Examples of types include, but are not limited to, TLlA-binding polypeptides, such as antigen-binding proteins (e.g., TL1 A antigen-binding proteins), antibodies, antibody fragments, and antibody derivatives.

[0514] The term "endocytosis" or “internalization” usually refers to the process by which the cell membrane transfers substances from outside the cell into the cell through deformation movements. Said endocytosis may involve both phagocytosis and cytotoxicity. For example, the cell may be depressed to form "vesicles" that encapsulates said macromolecules (e.g., proteins), but then detach from the cell membrane and enter the cell interior.

[0515] The degree of degradation by endocytosis can be evaluated by means of an "endocytosis degradation rate". For a cell-derived macromolecule (e.g., a soluble protein) interacting with a cell, the "endocytosis degradation rate" of said macromolecule may refer to the amount of said macromolecule that enters the cell via said endocytosis as a percentage of the total amount of said macromolecule interacting with said cell (which may generally be the sum of the amount endocytosed and the amount bound to the surface of said cell and the amount of said Attorney Docket No.: 769273: SA9-943PC macromolecule bound to the surface of said cell). (generally the total amount can be the sum of the amount endocytosed and the amount bound to the surface of said cell) as a proportion (usually expressed as a percentage) of the total amount of said macromolecule interacting with said cell. For a macromolecule on the surface of a cell membrane (e.g. a membrane protein on said cell), said "endocytosis degradation rate" of a macromolecule may refer to the ratio of the amount of said macromolecule which enters the cell via said endocytosis and is degraded to the total amount of said macromolecule on said cell (which generally may be the sum of the amount endocytosed and the amount of the macromolecule which is endocytosed and degraded). (generally the total amount can be the sum of the amount degraded by endocytosis and the amount remaining on the surface of the cell membrane after endocytosis; or it can also be expressed as the total amount on the surface of the cell membrane in the absence of endocytotic degradation), which is usually expressed as a percentage.)

[0516] As used herein, the term "epitope" or "antigenic determinant cluster" generally refers to a site on an antigen to which an antibody binds. Epitopes may be formed from contiguous amino acids (linear epitopes) or discontinuous amino acids (conformational epitopes) adjacent to each other by tertiary folding of the protein. Epitopes formed from contiguous amino acids are typically retained upon exposure to denaturing solvents, whereas epitopes formed by tertiary folding are typically lost upon treatment with denaturing solvents. Epitopes typically include at least 3, and more typically at least 5 or 8-10 amino acids in a unique spatial conformation. Methods for determining the spatial conformation of epitopes include, for example, x-ray crystallography and two-dimensional nuclear magnetic resonance. See, e.g., Epitope Mapping Protocols in Methods in Molecular Biology, vol. 66, edited by Glenn E. Morris (1996).

[0517] As used herein, the term "conformational epitope" generally refers to non-contiguous amino acid residues of an antigen (such as the PD-L1 antigen) that are adjacent to the protein by tertiary folding. These non-contiguous amino acid residues may converge on the surface when the polypeptide chain folds to form a natural protein. Conformational epitopes include, but are not limited to, functional epitopes. Attorney Docket No.: 769273: SA9-943PC

[0518] As used herein, the term "cross-linking" refers to the process by which an antigen-binding protein binds to more than one antigen at the same time, or an antigen binds to more than one antigen-binding protein at the same time. For example, RVT-3101 (Analogue) monoclonal antibody binds to huTLIA-his antigen, which can be seen as three monoclonal antibodies bound to a single TL1 A trimer, i.e., a 1 Ag (Ag represents the antigen) + 3 Ab (Ab represents the antibody) complex. tla0015-ll monoclonal antibody binds to the huTLIA-his antigen, which results in the formation of a multiple-antigen binding protein that binds to a number of antigen-binding proteins cross-linked complexes.

[0519] As used herein, the term "functional epitope" generally refers to the amino acid residues of an antigen that actively contributes to the binding of an antibody, i.e., form an "active epitope." Mutation of any active promoting residue of the antigen to alanine will destroy the binding of the antibody, so that the relative KD ratio of the antibody (KD mutant / KD wild type) can be, for example, greater than 2 times, such as greater than 3 times, greater than 4 times, greater than 6 times , greater than 10 times, greater than 20 times, greater than 30 times, greater than 40 times, greater than 50 times, greater than 60 times, greater than 70 times, greater than 80 times, greater than 90 times, greater than 100 times, greater than 150 times, greater than 200 times or more.

[0520] In the present application, the term “and / or” should be understood as meaning any one of the alternatives or both of the alternatives.

[0521] In the present application, the term “comprising” or “containing” generally refers to the inclusion of explicitly specified features, but not excluding other elements. In certain instances, “comprising” or “contain” also encompasses the inclusion of only the specified components.

[0522] In the present application, the term “about” generally refers to a range from 0.5% to 10% above or below the specified value, for example, a range of 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, or 10% above orbelow the specified value. Attorney Docket No.: 769273: SA9-943PC

[0523] In the present application, the term “including” generally means comprising, summing up, encompassing or covering. In certain instances, the meaning of “being”, “consisting of . . . ” is also indicated.

[0524] DETAILED DESCRIPTION

[0525] First antigen binding domain

[0526] In one aspect, the present application provides an isolated antigen-binding protein comprising a first antigen-binding domain which specifically binds to TL1 A. The first binding domain includes a heavy chain variable region (VH). The VH includes at least one heavy chain complementarity determining regions (HCDR). The VH may include at least one, two or three of HCDR1, HCDR2 or HCDR3. In some embodiments, HCDR 3, HCDR2 and HCDR1 are sequences as set forth in SEQ ID NO: 14, 5 and 1 respectively.

[0527] An HCDR3 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 14. For example, the sequence of HCDR3 of the antigen-binding protein may be defined by Kabat.

[0528] In some embodiments, the HCDR3 may be GX1X2DAMDY(SEQ ID NO: 14). XI may be D or L or N, X2 may be F or Y.

[0529] In some embodiments, the HCDR3 may include an amino acid sequence as set forth in any one of SEQ ID NOs: 15-18.

[0530] An HCDR2 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 5. For example, the sequence of the HCDR2 of the antigen-binding protein may be defined by Kabat.

[0531] In some embodiments, the HCDR2 may be X3IWGFGX4TX5YX6X7ALKS(SEQ ID NO:5). X3 may be R or V, and X4 may be G or K, and X5 may be D or H or N, and X6 may be N or Q, and X7 may be A or P or S.

[0532] In some embodiments, the HCDR2 may include an amino acid sequence as set forth in any one of SEQ ID NOs: 6-13. Attorney Docket No.: 769273: SA9-943PC

[0533] An HCDR1 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 1. For example, the sequence of the HCDR1 of the antigen-binding protein may be defined by Kabat.

[0534] In some embodiments, the HCDR1 may be X8YGVD (SEQ ID NO: 1). X8 may be S or T or Y

[0535] In some embodiments, the HCDR1 may include an amino acid as set forth in any one of SEQ ID NO:2-4.

[0536] In some embodiments, the VH comprises the HFR1, and C-terminus of the HFR1 is directly or indirectly linked to the N-terminus of the HCDR1. For example, the sequence of HFR1 of the antigen-binding protein may be defined by Kabat. HFR1 comprises an amino acid as set forth in SEQ ID NO: 19.

[0537] In some embodiments, the HFR1 is EX9QLX10ESGGGLXl lQPGGSLRX12SCAVSSGFXxlLXx2 (SEQ ID NO: 19). X9 may be I or V, X10 may be L or V, XI 1 may be I or V, X12 may be I or L. Xxl is D or S, Xx2 is I or K or S.

[0538] In some embodiments, the HFR1 is selected from an amino acid sequence as set forth in any one of SEQ ID NO:20-28.

[0539] In some embodiments, the VH comprises the HFR2, and the HFR2 is located between the HCDR1 and the HCDR2. For example, the sequence of the HFR2 of the antigen-binding protein may be defined by Kabat. the HFR2 comprises an amino acid sequence as set forth in SEQ ID NO:29.

[0540] In some embodiments, the HFR2 is WVRQX13PGKGLEWX14G(SEQ ID NO:29).X13 may be A or S, and X14 may be L or V.

[0541] In some embodiments, the HFR2is selected from an amino acid sequence as set forth in any one of SEQ ID NO:30-32.

[0542] In some embodiments, the VH comprises the HFR3, and the HFR3 is located between the HCDR2 and the HCDR3. For example, the sequence of the HFR3 of the antigen-binding Attorney Docket No.: 769273: SA9-943PC protein may be defined by Kabat. the HFR3 comprises an amino acid as set forth in SEQ ID NO:33.

[0543] In some embodiments, the HFR3 is RX15TISX16DNSKNTX17YLQMNSLRAEDTAVYYCAX18(SEQ ID NO:33. X15 may be F or L,X16 may be A or K or R or V, X17 may be L or V, X18 may be R or S.

[0544] In some embodiments, the HFR3is selected from an amino acid sequence as set forth in any one of SEQ ID NO:34-40.

[0545] In some embodiments, VH comprises the HFR4, and N-terminus of the HFR4 is directly or indirectly linked to C-terminus of the HCDR3. For example, the sequence of the HFR4 of the antigen-binding protein may be defined by Kabat. the HFR4 comprises an amino acid sequence as set forth in SEQ ID NO:41.

[0546] In some embodiments, the isolated antigen-binding protein further comprises a light chain variable region(VL). The VL comprises at least one light chain complementarity determining regions (LCDR). VL comprises an LCDR1, an LCDR2 and an LCDR3, which is selected from an amino acid sequence as set forth in any one of SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44 accordingly.

[0547] In some embodiments, the VL comprises an LCDR3, including an amino acid sequence as set forth in SEQ ID NO: 44. For example, the sequence of the LCDR3 of the antigenbinding protein may be defined by Kabat.

[0548] In some embodiments, the VL comprises an LCDR2, including an amino acid sequence as set forth in SEQ ID NO:43. For example, the sequence of the LCDR2 of the antigenbinding protein may be defined by Kabat.

[0549] In some embodiments, the VL comprises an LCDR1, including an amino acid sequence as set forth in SEQ ID NO:42. For example, the sequence of the LCDR1 of the antigenbinding protein may be defined by Kabat.

[0550] In some embodiments, VL comprises the LFR1, and C-terminus of the LFR1 is directly or indirectly linked to N-terminus of the LCDR1. For example, the sequence of the LFR1 of Attorney Docket No.: 769273: SA9-943PC the antigen-binding protein may be defined by Kabat. The LFR1 comprises an amino acid sequence as set forth in SEQ ID NO:45.

[0551] In some embodiments, the LFR1 is DIQMTQX19X20SSLSASVGDRVTIX21C(SEQ ID NO:45).X19 may be S or T, X20 may be P or T, X21 may be T or S.

[0552] In some embodiments, the LFR1 comprises an amino acid sequence as set forth in any one of SEQ ID NO:46-47.

[0553] In some embodiments, the VL comprises the LFR2, and the LFR2 is located between the LCDR1 and the LCDR2. For example, the sequence of the LFR2 of the antigen-binding protein may be defined by Kabat, the LFR2 comprises an amino acid sequence as set forth in SEQ ID NO:48.

[0554] In some embodiments, the LFR2 is WYQQKPGKX22X23KLLIY(SEQ ID NO:48). X22 may be A or V, and X23 may be P or V.

[0555] In some embodiments, the LFR2 comprises an amino acid sequence as set forth in any one of SEQ ID NO:49-52.

[0556] In some embodiments, the VL comprises the LFR3,the LFR3 is located between the LCDR2 and the LCDR3. For example, the sequence of the LFR3 of the antigen-binding protein may be defined by Kabat. The LFR3 comprises an amino acid sequence as set forth in SEQ ID NO 53.

[0557] In some embodiments, the LFR3 is GVPSRFSGSGSGTDX24TLTISSLQPEDX25ATYX26C(SEQ ID NO:53).X24 may be F or Y,X25 may be F or V,X26 may be F or Y.

[0558] In some embodiments, the LFR3 comprises an amino acid sequence as set forth in any one of SEQ ID NO:54-57.

[0559] In some embodiments, the isolated antigen-binding fragment comprises the LFR4, and N- terminus of the LFR4 is directly or indirectly linked to C terminus of the LCDR3. For example, the sequence of the LFR4 of the antigen-binding protein may be defined by Kabat. The LFR4 comprises an amino acid sequence as set forth in SEQ ID NO: 58. Attorney Docket No.: 769273: SA9-943PC

[0560] In some embodiments, the LFR4 is FGX27GTKLEIK(SEQ ID NO:58). X27 may be P or Q

[0561] In some embodiments, the LFR4 comprises an amino acid sequence as set forth in any one of SEQ ID NO:59-61. In some embodiments, the VH comprises sequences as set forth in SEQ ID NO:62-99.

[0562] In some embodiments, the VL comprises sequences as set forth in SEQ ID NO: 102-106.

[0563] In some embodiments, the isolated antigen-binding protein comprises a VH as shown in SEQ ID NO: 70 and a VL as shown in SEQ ID NO: 106.

[0564] In some embodiments, the VH of the isolated antigen-binding protein comprises a HCDR1 as shown in SEQ ID NO: 2, a HCDR2 as shown in SEQ ID NO: 6, and a HCDR3 as shown in SEQ ID NO: 15, and further comprises a light chain variable region (VL), wherein the VL comprises at least one light chain complementary determining region (LCDR), which comprises a LCDR1 as shown in SEQ ID NO: 42, a LCDR2 as shown in SEQ ID NO: 43, and a LCDR3 as shown in SEQ ID NO: 44. In some embodiments, the antigen-binding proteins described above are described in table A and table B .

[0565] In some embodiments, the distribution of antibodies for the TLA-0015 line is shown in the following table, with Table A showing the distribution of heavy chain components and Table B showing the distribution of light chain components. Table A Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0566] Table B Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0567] In one aspect, the present application provides an isolated antigen-binding protein comprising a first antigen-binding domain which specifically binds to TL1 A. The first binding domain includes a heavy chain variable region (VH). The VH comprises at least one heavy chain complementarity determining regions (HCDR). The VH may include at least one, two or three of HCDR1, HCDR2 or HCDR3. In some embodiments, the HCDR 1, the

[0568] HCDR3 and the HCDR3 are sequences as set forth in SEQ ID NO: 107, 115 and 120 respectively.

[0569] The HCDR3 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 120. For example, the sequence of the HCDR3 of the antigen-binding protein may be defined by Kabat. Attorney Docket No.: 769273: SA9-943PC

[0570] The HCDR2 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 115. For example, the sequence of the HCDR2 of the antigen-binding protein may be defined by Kabat.

[0571] In some embodiments, the HCDR2 may be QIRLKSDNYATHYAX28X29VKG(SEQ ID NO: 115). X28 may be A or D or E. X29 may be P or S.

[0572] In some embodiments, the HCDR2 may include an amino acid sequence as set forth in any one of SEQ ID NOs: 116-119. the HCDR1 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 107. For example, the sequence of the HCDR1 of the antigen-binding protein may be defined by Kabat.

[0573] In some embodiments, the HCDR1 may be NYX30MX31 (SEQ ID NO: 107). X30 may be I or L or M or Y or W. X31 may be N or H or S.

[0574] In some embodiments, the HCDR1 may include an amino acid as set forth in any one of SEQ ID NO: 108-114.

[0575] In some embodiments, the VH comprises the HFR1, and C-terminus of the HFR1 is directly or indirectly linked to the N-terminus of the HCDR1. For example, the sequence of the HFR1 of the antigen-binding protein may be defined by Kabat. the HFR1 comprises an amino acid as set forth in SEQ ID NO: 121.

[0576] In some embodiments, the HFR1 is X32VX33X34X35ESGGGX36VX37PGX38X39LX40LSCX41ASGFTFS (SEQ ID NO: 121). X32 may be E or Q. X33 may be K or Q.X34 may be L or V.X35 may be E or V.X36 may be L or V.X37 may be K or Q. X38 may be G or R.X39 may be A or S.X40 may be K or R.X41 may be A or I.

[0577] In some embodiments, the HFR1 is selected from an amino acid sequence as set forth in any one of SEQ ID NO: 122-132.

[0578] In some embodiments, VH comprises the HFR2, and the HFR2 is located between the HCDR1 and the HCDR2. For example, the sequence of the HFR2 of the antigen-binding Attorney Docket No.: 769273: SA9-943PC protein may be defined by Kabat. the HFR2 comprises an amino acid sequence as set forth in SEQ ID NO: 133.

[0579] In some embodiments, the HFR2 is WVRQX42PX43KGLEWVX44(SEQ ID NO: 133). X42 may be A or S. X43 may be E or G.X44 may be A or G.

[0580] In some embodiments, the HFR2 is selected from an amino acid sequence as set forth in any one of SEQ ID NO: 134-135 and 30.

[0581] In some embodiments, VH comprises the HFR3, and the HFR3 is located between the HCDR2 and the HCDR3. For example, the sequence of the HFR3 of the antigen-binding protein may be defined by Kabat. the HFR3 comprises an amino acid as set forth in SEQ ID NO: 136.

[0582] In some embodiments, the HFR3 is X45FX46ISRDX47X48KX49X50X51 YLQMNSLX52X53EDX54AVYYCTP(SEQ ID NO: 136). X45 may be N or R.X46 may be A or T.X47 may be D or N.X48 may be A or S.X49 may be N or S.X50 may be S or T.X51 may be A or L or V.X52 may be K or R.X53 may be A or D or T.X54 may be M or T.

[0583] In some embodiments, the HFR3 is selected from an amino acid sequence as set forth in any one of SEQ ID NO: 137-148.

[0584] In some embodiments, VH comprises the HFR4, and N-terminus of the HFR4 is directly or indirectly linked to C-terminus of the HCDR3. For example, the sequence of the HFR4 of the antigen-binding protein may be defined by Kabat. The HFR4 comprises an amino acid sequence as set forth in SEQ ID NO: 149.

[0585] In some embodiments, the HFR4 is WGQGT X55X56TVSS (SEQ ID NO: 149). X55 may be L or M OR H or T.X56 may be L or V.

[0586] In some embodiments, the isolated antigen-binding fragment further comprises a light chain variable region(VL), and the VL comprises at least one light chain complementarity determining regions (LCDR). VL comprises an LCDR1, an LCDR2 and an LCDR3, which is selected from an amino acid sequence as set forth in any one of SEQ ID NO: 154, SEQ ID NO: 159 and SEQ ID NO: 163 accordingly. Attorney Docket No.: 769273: SA9-943PC

[0587] In some embodiments, the VL comprises the LCDR3, including an amino acid sequence as set forth in SEQ ID NO: 163. For example, the sequence of the LCDR3 of the antigenbinding protein may be defined by Kabat.

[0588] In some embodiments, the LCDR3 is FQEX57X58HPFT(SEQ ID NO: 163). X57 may be N or S or E.X58 may be G or E or A.

[0589] In some embodiments, the LCDR3 is selected from an amino acid sequence as set forth in any one of SEQ ID NO : 164- 168.

[0590] In some embodiments, the VL comprises the LCDR2, including an amino acid sequence as set forth in SEQ ID NO: 159. For example, the sequence of the LCDR2 of the antigenbinding protein may be defined by Kabat.

[0591] In some embodiments, the LCDR2 is DTSNX59AX60 (SEQ ID NO: 159). X59 may be L or R. X60 may be S or T.

[0592] In some embodiments, the LCDR2 is selected from an amino acid sequence as set forth in any one of SEQ ID NO: 160-162.

[0593] In some embodiments, the VL comprises the LCDR1, including an amino acid sequence as set forth in SEQ ID NO:42. For example, the sequence of the LCDR1 of the antigenbinding protein may be defined by Kabat.

[0594] In some embodiments, the LCDR1 is SASSSVSYX61X62 (SEQ ID NO: 154). X61 may be L or M. X62 may be A or H or T.

[0595] In some embodiments, the LCDR1 is selected from an amino acid sequence as set forth in any one of SEQ ID NO : 155 - 158.

[0596] In some embodiments, the VL comprises the LFR1, and C-terminus of the LFR1 is directly or indirectly linked to N-terminus of the LCDR1. For example, the sequence of the LFR1 of the antigen-binding protein may be defined by Kabat. The LFR1 comprises an amino acid sequence as set forth in SEQ ID NO: 169.

[0597] In some embodiments, the LFR1 is X63X64X65X66TQSPX67X68X69SX70X71X72GX73X74X75TX76X77C(SEQ ID NO: 169). X63 may be E or D.X64 may be I or N or T or V.X65 may be Q or V.X66 may be L Attorney Docket No.: 769273: SA9-943PC or M.X67 may be A or S. X68 may be F or I or S or T.X69 may be L or M or V. X70 may be A or L or V. X71 may be S or T.X72 may be P or V.X73 may be D or E. X74 may be K or R.X75 may be A or V.X76 may be I or M.X77 may be S or T.

[0598] In some embodiments, the LFR1 comprises an amino acid sequence as set forth in any one of SEQ ID NO : 170- 179.

[0599] In some embodiments, the VL comprises the LFR2, and the LFR2 is located between the LCDR1 and the LCDR2. For example, the sequence of the LFR2 of the antigen-binding protein may be defined by Kabat, the LFR2 comprises an amino acid sequence as set forth in SEQ ID NO: 180.

[0600] In some embodiments, the LFR2 is WYQQKP X78 X79 X80P X81LWIY(SEQ ID NO: 180).X78 may be D or G.X79 may be K or Q.X80 may be A or S.X81 may be K or Q or R.

[0601] In some embodiments, the LFR2 comprises an amino acid sequence as set forth in any one of SEQ ID NO: 181-188.

[0602] In some embodiments, the VL comprises the LFR3, and the LFR3 is located between the LCDR2 and the LCDR3. For example, the sequence of the LFR3 of the antigen-binding protein may be defined by Kabat. The LFR3 comprises an amino acid sequence as set forth in SEQ ID NO: 189.

[0603] In some embodiments, the LFR3 is GVP X82RFSGSGSGX83X84X85TLTISSX86X87X88EDX89AX90YYC(SEQ ID NO: 189). X82 may be A or D or G or S. X83 may be N or T.X84 may be D or S. X85 may be F or Y. X86 may be L or M.X87 may be E or Q. X88 may be A or P.X89 may be A or F or V. X90 may be T or V.

[0604] In some embodiments, the LFR3 comprises an amino acid sequence as set forth in any one of SEQ ID NO: 190-197 and 57.

[0605] In some embodiments, the isolated antigen-binding fragment comprises the LFR4, and N-terminus of the LFR4 is directly or indirectly linked to C terminus of the LCDR3. For Attorney Docket No.: 769273: SA9-943PC example, the sequence of the LFR4 of the antigen-binding protein may be defined by Kabat. The LFR4 comprises an amino acid sequence as set forth in SEQ ID NO: 198.

[0606] In some embodiments, the LFR4 is FG X91GTK X92EX93K(SEQ ID NO: 198).X91 may be G or Q. X92 may be L or V. X93 may be I or M. In some embodiments, the LFR4 comprises an amino acid sequence as set forth in any one of SEQ ID NO : 199-201 and 61.

[0607] In some embodiments, VH comprises sequences as set forth in SEQ ID N0:202-220.

[0608] In some embodiments, VL comprises sequences as set forth in SEQ ID NO :221-234.

[0609] In some embodiments, the antigen-binding proteins described above are as described in Tables C and D.

[0610] In some embodiments, the distribution of antibodies for the TLA-0238 line is shown in the following table, with Table C showing the distribution of heavy chain components and Table D showing the distribution of light chain components.

[0611] Table C Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0612] Table D Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0613] In one aspect, the present application provides an isolated antigen-binding protein comprising a first antigen-binding domain which specifically binds to TL1 A. The first binding domain includes a heavy chain variable region (VH). The VH comprises at least one heavy chain complementarity determining regions (HCDR). The VH may include at least one, two or three of the HCDR1, the HCDR2 or the HCDR3. In some embodiments, the HCDR 1, the HCDR3 and the HCDR3 are sequences as set forth in SEQ ID NO:235, 238 and 242 respectively.

[0614] The HCDR3 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO: 242. For example, the sequence of the HCDR3 of the antigen-binding protein may be defined by Kabat. Attorney Docket No.: 769273: SA9-943PC

[0615] The HCDR2 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO:238. For example, the sequence of the HCDR2 of the antigen-binding protein may be defined by Kabat.

[0616] In some embodiments, the HCDR2 may be DINPNNGRTTYX94X95X96X97X98G(SEQ ID NO:238). X94 may be A or N. X95 may be D or Q. X96 may be K or S. X97 may be F or V. X98 may be K or Q or T.

[0617] In some embodiments, the HCDR2 may include an amino acid sequence as set forth in any one of SEQ ID NOs:239-241. the HCDR1 of the antigen-binding protein may include amino acid sequence as set forth in SEQ ID NO:235. For example, the sequence of the HCDR1 of the antigen-binding protein may be defined by Kabat.

[0618] In some embodiments, the HCDR1 may be DYY X99 X100(SEQ ID NO:235). X99 may be L or M. X100 may be N or H.

[0619] In some embodiments, the HCDR1 may include an amino acid as set forth in any one of SEQ ID NO:236-237.

[0620] In some embodiments, the VH comprises HFR1, and C-terminus of HFR1 is directly or indirectly linked to the N-terminus of the HCDR1. For example, the sequence of HFR1 of the antigen-binding protein may be defined by Kabat. HFR1 comprises an amino acid as set forth in SEQ ID NO:243.

[0621] In some embodiments, HFR1 is X101IX102 X103TQSPX104 X105 X106S X107S X108G X109 X110VTITC(SEQ ID NO: 243). X101 may be A or D or E. X102 may be V or Q. X103 may be L or M. X104 may be A or G or P or S or T. X105 may be S or T. X106 may be L or V. X107 may be A or L. X108 may be P or V. X109 may be D or E. XI 10 may be K or R.

[0622] In some embodiments, HFR1 is selected from an amino acid sequence as set forth in any one of SEQ ID NO:244-250.

[0623] In some embodiments, VH comprises the HFR2, and the HFR2 is located between the HCDR1 and the HCDR2. For example, the sequence of the HFR2 of the antigen-binding Attorney Docket No.: 769273: SA9-943PC protein may be defined by Kabat. HFR2 comprises an amino acid sequence as set forth in SEQ ID NO:251.

[0624] In some embodiments, the HFR2 is WYQQKP XI 11 XI 12 X113P X114LLIY(SEQ ID NO:251). XI 11 may be D or G. XI 12 may be K or Q. XI 13 may be A or S. XI 14 may be K or Q.

[0625] In some embodiments, the HFR2 is selected from an amino acid sequence as set forth in any one of SEQ ID NO: 252-255.

[0626] In some embodiments, VH comprises the HFR3, and the HFR3 is located between the HCDR2 and the HCDR3. For example, the sequence of the HFR3 of the antigen-binding protein may be defined by Kabat. the HFR3 comprises an amino acid as set forth in SEQ ID NO:256.

[0627] In some embodiments, the HFR3 is

[0628] GVPX1 15RFSGSGSGTX116FTLTIX117SLX118X119ED X120ATYYC(SEQ ID NO:256). X115 may be A or D or S. X116 may be D or E or S. X117 may be S or N. X118 may be E or Q. XI 19 may be A or P. X120 may be A or F.

[0629] In some embodiments, the HFR3 is selected from an amino acid sequence as set forth in any one of SEQ ID NO:257-262.

[0630] In some embodiments, VH comprises the HFR4, and N-terminus of the HFR4 is directly or indirectly linked to C-terminus of the HCDR3. For example, the sequence of the HFR4 of the antigen-binding protein may be defined by Kabat. the HFR4 comprises an amino acid sequence as set forth in SEQ ID NO:263.

[0631] In some embodiments, the HFR4 is FGX121GTX122X123EX124K(SEQ ID NO:263). X121 may be A or Q. X122 may be K or R. X123 may be L or V. X124 may be I or L.

[0632] In some embodiments, the HFR4 is selected from an amino acid sequence as set forth in any one of SEQ ID NO:264-268.

[0633] In some embodiments, the isolated antigen-binding fragment further comprises a light chain variable region(VL), an VL comprises at least one light chain complementarity determining regions (LCDR). VL comprises an LCDR1, an LCDR2 and an LCDR3, which is Attorney Docket No.: 769273: SA9-943PC selected from an amino acid sequence as set forth in any one of SEQ ID NO: 269, SEQ ID NO:274 and SEQ ID NO:277 accordingly.

[0634] In some embodiments, VL comprises the LCDR3, including an amino acid sequence as set forth in SEQ ID NO:277. For example, the sequence of the LCDR3 of the antigen-binding protein may be defined by Kabat.

[0635] In some embodiments, VL comprises the LCDR2, including an amino acid sequence as set forth in SEQ ID NO:274. For example, the sequence of the LCDR2 of the antigen-binding protein may be defined by Kabat.

[0636] In some embodiments, the LCDR2 is KTSN X125PS(SEQ ID NO:274). X125 may be L or R.

[0637] In some embodiments, the LCDR2 is selected from an amino acid sequence as set forth in any one of SEQ ID NO:275-276.

[0638] In some embodiments, VL comprises the LCDR1, including an amino acid sequence as set forth in SEQ ID NO:269. For example, the sequence of the LCDR1 of the antigen-binding protein may be defined by Kabat.

[0639] In some embodiments, the LCDR1 is SX126SSSIISNYX127X128(SEQ ID NO:269). X126 may be A or T. X127 may be L or S. X128 may be A or H.

[0640] In some embodiments, the LCDR1 is selected from an amino acid sequence as set forth in any one of SEQ ID NO:270-273.

[0641] In some embodiments, VL comprises the LFR1, and C-terminus of the LFR1 is directly or indirectly linked to N-terminus of the LCDR1. For example, the sequence of the LFR1 of the antigen-binding protein may be defined by Kabat. the LFR1 comprises an amino acid sequence as set forth in SEQ ID NO:278.

[0642] In some embodiments, the LFR1 is X129IX130X131TQSPX132X133X134SX135SX136GX137X138VTITC(SEQ ID NO:278).

[0643] X129 may be A or E or D. X130 may be Q or V. X131 may be L or M. X132 may be A or G or P or S or T. X133 may be S or T. X134 may be L or V. X135 may be L or A. X136 may be P or V. X137 may be D or E. X138 may be K or R. Attorney Docket No.: 769273: SA9-943PC

[0644] In some embodiments, the LFR1 comprises an amino acid sequence as set forth in any one of SEQ ID NO: 47, 173, 279-284.

[0645] In some embodiments, VL comprises the LFR2, and the LFR2 is located between the LCDR1 and the LCDR2. For example, the sequence of the LFR2 of the antigen-binding protein may be defined by Kabat, the LFR2 comprises an amino acid sequence as set forth in SEQ ID NO:284.

[0646] In some embodiments, the LFR2 is WYQQKP X139 X140 X141P X142LLIY(SEQ ID NO:284). X139 may be D or G. X140 may be K or Q. X141 may be A or S. X142 may be K or Q.

[0647] In some embodiments, the LFR2 comprises an amino acid sequence as set forth in any one of SEQ ID NO:52, 285-287.

[0648] In some embodiments, VL comprises the LFR3,the LFR3 is located between the LCDR2 and the LCDR3. For example, the sequence of the LFR3 of the antigen-binding protein may be defined by Kabat. The LFR3 comprises an amino acid sequence as set forth in SEQ ID NO: 288.

[0649] In some embodiments, the LFR3 is GVPX143RFSGSGSGTX144FTLTIX145SLX146X147ED X148ATYYC(SEQ ID NO:288). X143 may be A or D or S. X144 may be D or E or S. X145 may be N or S. X146 may be E or Q. X147 may be A or P. X148 may be A or F.

[0650] In some embodiments, the LFR3 comprises an amino acid sequence as set forth in any one of SEQ ID NO:289-293 and 57.

[0651] In some embodiments, the isolated antigen-binding fragment comprises the LFR4, and N-terminus of the LFR4 is directly or indirectly linked to C terminus of the LCDR3. For example, the sequence of the LFR4 of the antigen-binding protein may be defined by Kabat. The LFR4 comprises an amino acid sequence as set forth in SEQ ID NO:294.

[0652] In some embodiments, the LFR4 is FG FGX148GTX149X150EX151K(SEQ ID NO:294). X148 may be A or Q. X149 may be K or R. XI 50 may be L or V. XI 51 may be I or L. Attorney Docket No.: 769273: SA9-943PC

[0653] In some embodiments, the LFR4 comprises an amino acid sequence as set forth in any one of SEQ ID NO:295-296 and 201.

[0654] In some embodiments, VH comprises sequences as set forth in SEQ ID NO:297-230.

[0655] In some embodiments, VL comprises sequences as set forth in SEQ ID NO:308-317. In some embodiments, the antigen-binding proteins described above are as described in Tables E and F.

[0656] In some embodiments, the distribution of antibodies for the TLA-0238 series is shown in the following table, with Table E showing the distribution of heavy chain components and Table F showing the distribution of light chain components. Table E Attorney Docket No.: 769273: SA9-943PC

[0657] Table F Attorney Docket No.: 769273: SA9-943PC

[0658] In some embodiments, said first antigen-binding domain may be a binding domain of an RVT3101 antibody, e.g., comprising VH as shown in SEQ ID NO:322 and VL as shown in SEQ ID NO:323. Attorney Docket No.: 769273: SA9-943PC

[0659] Second antigen binding domain

[0660] In the present application, said isolated antigen-binding protein may comprise a second antigen-binding domain.

[0661] The second antigen-binding domain does not specifically bind to TL1 A.

[0662] The second antigen-binding domain specifically binds to a4p7 .

[0663] In the present application, the second antigen-binding domain of said isolated antigenbinding protein may comprise scFv.

[0664] In the present application, in said isolated antigen-binding protein wherein said second antigen binding domain VH comprises an HCDR1 as shown in SEQ ID NO: 332, an HCDR2 as shown in SEQ ID NO: 333, and an HCDR3 as shown in SEQ ID NO: 334.

[0665] In the present application, in said isolated antigen-binding protein, said second antigenbinding domain may comprise a heavy chain variable region VH, and said VH may comprise the amino acid sequence shown in SEQ ID NO: 335.

[0666] In the present application, in said isolated antigen-binding protein, said second antigenbinding domain may comprise the LCDR3, and said the LCDR3 may comprise the amino acid sequence shown in SEQ ID NO: 34.

[0667] The second antigen binding domain VL comprises the LCDR1 as shown in SEQ ID NO: 336, the LCDR2 as shown in SEQ ID NO: 337, and the LCDR3 as shown in SEQ ID NO: 338.

[0668] In some embodiments, the second antigen binding domain VH comprises the HFR1 as shown in SEQ ID NO: 382, the HFR2 as shown in SEQ ID NO: 383, the HFR3 as shown in SEQ ID NO: 384, the HFR4 as shown in SEQ ID NO: 385 and / or the second antigen binding domain VL comprises the LFR1 as shown in SEQ ID NO: 386, the LFR2 as shown in SEQ ID NO: 387, the LFR3 as shown in SEQ ID NO: 388, the LFR4 as shown in SEQ ID NO: 389.

[0669] In the present application, said second antigen binding domain comprises VL as shown in SEQ ID NO: 339.

[0670] In some embodiments, the first antigen-binding domain comprises HCDR1 as set forth in SEQ ID NO: 2, HCDR2 as set forth in SEQ ID NO: 6, HCDR3 as set forth in SEQ ID NO: 15, LCDR1 as set forth in SEQ ID NO: 42, LCDR2 as set forth in SEQ ID NO: 43, and Attorney Docket No.: 769273: SA9-943PC

[0671] LCDR3 as set forth in SEQ ID NO: 44; and, the second antigen-binding domain comprises HCDR1 as set forth in SEQ ID NO: 332, HCDR2 as set forth in SEQ ID NO: 333, HCDR3 as set forth in SEQ ID NO: 334, LCDR1 as set forth in SEQ ID NO: 336, LCDR2 as set forth in SEQ ID NO: 337, and LCDR3 as set forth in SEQ ID NO: 338.

[0672] In some embodiments, the first antigen-binding domain comprises a VH as set forth in SEQ ID NO: 70 and a VL as set forth in SEQ ID NO: 106; and the second antigen-binding domain comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339.

[0673] In some embodiments, (1) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 340; and the second antigen-binding domain comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339; or, (2) the first antigenbinding domain comprises an scFv as set forth in SEQ ID NO: 341; and the second antigenbinding domain comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339; or, (3) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 342; and the second antigen-binding domain comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339; or, (4) the first antigen -binding domain comprises an scFv as set forth in SEQ ID NO: 343; and the second antigen-binding domain comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339; or, (5) the first antigen binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106; and the second antigen binding domain comprises an scFv as set forth in SEQ ID NO:346; or, (6) the first antigen binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106; and the second antigen binding domain comprises an scFv as set forth in SEQ ID NO:347.

[0674] Isolated Antigen-Binding Protein

[0675] In one aspect, the present application provides an isolated antigen-binding protein that can bind to a monomer TL1A and / or trimer TL1A with a KD value of about 1.0E-8M or less (For example, said KD is no greater than about 5E-08M, no greater than about 4.5E-08M, no greater than about 4E-08M, no greater than about 3.5E-08M, no greater than about 3E-08M, no greater than about 2E-08M, no greater than about 2E-08M, no greater than about 1.5E-08M, no greater than about 1E-08M, no greater than about 5E-09M, no greater than about4.5E-09M, no greater Attorney Docket No.: 769273: SA9-943PC than about 4E-09M, no greater than about 3.5E-09M, no greater than about 3E-09M, no greater than about 2.5E-09M, no greater than about 2E-09M, no greater than about 1.5E-09M, no greater than about 1E-09M, no greater than about 9E-10M, no greater than about 5E-10M, no greater than about 1E-10M, no greater than about5E-HM, no greater than about 1E-11M, or no greater than 5E-12M or less.)

[0676] In the present application, the isolated antigen-binding protein of the present disclosure is an antagonist of the TL1A receptor, including, but not limited to, DR3 and TR6 / DcR3. In the present application, the isolated antigen-binding protein of the present disclosure blocks the interaction of TL1A with death receptor 3 ("DR3"). In the present application, said isolated antigen-binding protein blocks monomeric TLlA-protein and / or trimeric TL1A in a flow- through assay at an IC50 value of about 50 nM or less (e.g., said IC50 value is no higher than about 50 nM, no higher than about 45 nM, no higher than about 40 nM, no higher than about 35 nM, no higher than about 30 nM, no higher than about 25 nM or less) binding to DR3 (DR3 : UniprotKB NO. Q93038).

[0677] In one aspect, the present application provides an isolated antigen-binding protein that can bind to a a4p7 protein with a KD value of about 1.0E-8M or less (For example, said KD is no greater than about 5E-08M, no greater than about 4.5E-08M, no greater than about 4E-08M, no greater than about 3.5E-08M, no greater than about 3E-08M, no greater than about 2E-08M, no greater than about 2E-08M, no greater than about 1.5E-08M, no greater than about 1E-08M, no greater than about 5E-09M, no greater than about4.5E-09M, no greater than about 4E-09M, no greater than about 3.5E-09M, no greater than about 3E-09M, no greater than about 2.5E- 09M, no greater than about 2E-09M, no greater than about 1.5E-09M, no greater than about 1E-09M, no greater than about 9E-10M, no greater than about 5E-10M, no greater than about 1E-10M, no greater than about5E-HM, no greater than about 1E-11M, or no greater than 5E- 12M or less.).

[0678] In the present application, the isolated antigen-binding protein is an antagonist of a4p7 protein receptors, including, but not limited to, MAdCAM-1. In the present application, said isolated antigen-binding protein blocks the interaction of TL1A with mucosal addressin cell Attorney Docket No.: 769273: SA9-943PC adhesion molecule-1 ("MAdCAM-1"). In the present application, the isolated antigen-binding protein blocks binding of a4p7 protein and MAdCAM-1 in a flow-through assay at an IC50 value of about 50 nM or less (e.g., said IC50 value is no higher than about 50 nM, no higher than about 45 nM, no higher than about 40 nM, no higher than about 35 nM, no higher than about 30 nM, no higher than about 25 nM or less).

[0679] In the present application, the a4p7 protein does not affect the blocking activity of isolated antigen-binding proteins against TL1 A proteins.

[0680] In the present application, the isolated antigen-binding proteins can inhibit the activity of the soluble TLlA-activated NF-KB signaling pathway. For example, the soluble TL1A- activated NF-KB signaling pathway can be tested for inhibition by a luciferase reporter gene system.

[0681] In the present application, said isolated antigen-binding protein can inhibit the activity of the membrane-bound TLlA-activated NF-KB signaling pathway. For example, whether the membrane-bound TLlA-activated NF-KB signaling pathway is inhibited can be detected by a luciferase reporter gene system.

[0682] In the present application, said isolated antigen-binding protein may function to inhibit Caspase3 / 7 activity. For example, the function of the Caspase3 / 7 activity can be tested for inhibition by a luciferase reporter gene system.

[0683] In the present application, said isolated antigen-binding protein blocks MAdCAM-1 mediated cell adhesion. The blocking ability can be determined, for example, by comparing curves of different antibody concentrations blocking MAdCAM-1 mediated cell adhesion to Hut 78.

[0684] In the present application, said isolated antigen-binding protein down-regulates the expression of a4p7. Said expression of a4p7 can be total expression or membrane expression.

[0685] In the present application, said isolated antigen-binding protein can inhibit the activity of CD4T cells.

[0686] In the present application, said isolated antigen-binding protein inhibits the activity of

[0687] IFN-y. Attorney Docket No.: 769273: SA9-943PC

[0688] In another aspect, the present application provides an antigen-binding protein which competitively binds a4p7 protein with the antigen-binding protein described in the present application.

[0689] In another aspect, the present application provides an antigen-binding protein which competitively binds TL1 A with the antigen -binding protein described in the present application.

[0690] A binding site of the first antigen-binding domain binding to the TL1 A is selected from one or more of the following: R32, Q33, P35, T36, Q37, P44, S89, K102, P103, D161, 1162, SI 63 and LI 64, including any one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve or thirteen of the above mentioned sites; and the binding sites are numbered according to the amino acid sequence of TL1A as set forth in SEQ ID NO: 327.

[0691] A binding site of the antigen-binding protein binding to the TLlA is selected from one or more of the following:R32, E49, E51, L54, A56, Y117, M146, S148, and Y167, including any one, two, three, four, five, six, seven, eight, and nine of the above mentioned site; and the binding sites are numbered according to the amino acid sequence of TL1 A as set forth in SEQ ID NO: 327.

[0692] In the present application, the first antigen-binding domain or the second antigen-binding domain comprises one or any combination of the following: Fab, Fab', F(ab')2, Fd and Fv fragments, disulfide bond stabilized Fv fragments (dsFv)., (dsFv)2, bispecific dsFv (dsFv- dsFv'), disulfide bond stabilized double-stranded antibody (diabody) (ds double-stranded antibody), single-chain antibody molecule (scFv), Single domain antibody (SDAB), scFv dimer (divalent double-stranded antibody), single domain antibody or nanobody, domain antibody ( domain antibody (dAb), shark variable IgNAR domain, camelized VH domain, chicken heavy chain antibody domain, VHH domain, minimum identification unit composed of CDR of mimetic antibody ( minimal recognition unit), alternative scaffolds for binding antigens, bivalent domain antibodies, pre-designed ankyrin repeat proteins (DARPins), multispecific proteins (including antigen-binding fragments or any other antibody fragments bound to antigens, but not intact antibody structures). Attorney Docket No.: 769273: SA9-943PC

[0693] In the present application, the antigen-binding protein comprises the scFv of the first antigen binding domain or the scFv of the second antigen binding domain.

[0694] In the present application, the scFv of the first antigen-binding domain comprises VL and VH of the first antigen-binding domain.

[0695] In the present application, in the first antigen-binding domain, the C-terminus of the VL of the first antigen-binding domain is linked directly or via a linker to the N-terminus of the VH of the first antigen-binding domain, or the N-terminus of the VL of the first antigen-binding domain is linked directly or via a linker to the C-terminus of the VH of the first antigen-binding domain.

[0696] In the present application, scFv of the first antigen-binding domains comprises sequences as set forth in any one of SEQ ID NO: 340-345.

[0697] In the present application, the scFv of the second antigen-binding domain comprises VL and VH of the second antigen-binding domain.

[0698] In the present application, in the second antigen binding domain, the C-terminus of the VL of the second antigen binding domain is linked directly or via a linker to the N-terminus of the VH of the second antigen binding domain, or the N-terminus of the VL of the second antigen binding domain is linked directly or via a linker to the C-terminus of the VH of the second antigen binding domain.

[0699] In the present application, scFv of the second antigen-binding domains comprises sequences as set forth in any one of SEQ ID NO: 346-347.

[0700] In the present application, the isolated antigen-binding protein comprises a heavy chain constant region of IgG and / or the light chain constant region of a human antibody

[0701] In the present application, the isolated antigen-binding protein comprises heavy chain constant region of IgGl, IgG2, IgG3 or IgG4

[0702] In the present application, the isolated antigen-binding protein comprises a Kappa or Lambda light chain constant region.

[0703] In the present application, the isolated antigen-binding protein is derived from IgG, IgM,

[0704] IgA, IgD or IgE. Attorney Docket No.: 769273: SA9-943PC

[0705] In the present application, the isolated antigen-binding protein comprises one or more first heavy chains and one or more first light chains.

[0706] The first heavy chain comprises VH of the first antigen-binding domain and / or VH of the second antigen binding domain and / or scFv of the first antigen-binding domain and / or scFv of the second antigen binding domain; the first light chain comprises a VL of the first antigen-binding domain or a VL of the second antigen binding domain.

[0707] In the present application, the first heavy chain comprises VH of the first antigen-binding domain and scFv of the second antigen binding domain, or the first heavy chain comprises VH of the second antigen binding domain and scFv of the first antigen-binding domain.

[0708] In the present application, the N-terminus of the scFv of the first antigen-binding domain or the second antigen binding domain is linked directly or via a linker to the C-terminus of the first heavy chain, or the C-terminus of the scFv of the first antigen-binding domain or the second antigen binding domain is linked directly or via a linker to the N-terminus of the first heavy chain.

[0709] In the present application, the isolated antigen-binding protein is selected from one of the following: the C-terminus of the first antigen-binding domain VH is linked to the N-terminus of said first heavy chain directly or via a linker, and the N-terminus of the scFv of the second antigen binding domain is linked to the C-terminus of said first heavy chain directly or via a linker; the C-terminus of the VL of the first antigen-binding domain is located at the N-terminus of the constant region of the light chain;

[0710] (2) the C-terminus of the second antigen binding domain VH is linked to the N-terminus of said first heavy chain directly or via a linker, and the N-terminus of the scFv of the first antigen-binding domain is linked to the C-terminus of said first heavy chain directly or via a linker; the C-terminus of the VL of the second antigen binding domain is located at the N- terminus of the light-chain constant region; Attorney Docket No.: 769273: SA9-943PC

[0711] (3) the C-terminus of the scFv of the first antigen-binding domain is linked, directly or via a linker, to the N-terminus of said second antigen binding domain VH, and the C-terminus of the VL of the second antigen binding domain is linked to the N-terminus of the light chain constant region;

[0712] (4) the C-terminus of the scFv of the second antigen binding domain is connected, directly or via a linker, to the N-terminus of said first antigen-binding domain VH, and the C-terminus of the VL of the first antigen-binding domain is linked to the N-terminus of the light chain constant region.

[0713] The first heavy chain is selected from the sequence as set forth in any one of the SEQ ID NO: 350-366 and 378.

[0714] The antigen-binding fragment is selected from one of the following:

[0715] 1) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO: 350;

[0716] 2) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:351;

[0717] 3) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:352;

[0718] 4) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:353;

[0719] 5) light chain of SEQ ID NO:349 and heavy chain of SEQ ID NO:354;

[0720] 6) light chain of SEQ ID NO:349 and heavy chain of SEQ ID NO:355;

[0721] 7) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:356;

[0722] 8) light chain of SEQ ID NO:372 and heavy chain of SEQ ID NO:357;

[0723] 9) light chain of SEQ ID NO:349 and heavy chain of SEQ ID NO:358;

[0724] 10) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:359;

[0725] 11) light chain of SEQ ID NO: 372 and heavy chain of SEQ ID NO: 360;

[0726] 12) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:361;

[0727] 13) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:362;

[0728] 14) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:363;

[0729] 15) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:364;

[0730] 16) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:365;

[0731] 17) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:366; Attorney Docket No.: 769273: SA9-943PC

[0732] 18) light chain of SEQ ID NO:348 and heavy chain of SEQ ID NO:378.

[0733] In some embodiments, the isolated antigen-binding protein is selected from one of the following: a light chain as shown in SEQ ID NO: 348 and heavy chain as shown in SEQ ID NO: 363.

[0734] In some embodiments, the antigen-binding proteins described in the present application have one, two, three, or four combinations of the light and heavy chains described above.

[0735] In the present application, antigen-binding proteins comprises one or more first light chains, one or more third heavy chains, and one or more fourth heavy chains, wherein:

[0736] 1) the third heavy chain comprises VH for the first antigen binding domain or the second antigen binding domain, and the fourth heavy chain comprises the ScFv of the first or second antigen binding domain;

[0737] 2) the third heavy chain comprises ScFv of the first or second antigen binding domain, the fourth heavy chain comprises the VH of the first or second antigen binding domain.

[0738] In the present application, the N-terminus of the first antigen-binding domain or the VH of the second antigen binding domain is linked directly or via a linker to the C-terminus of the third heavy chain, or the N-terminus of the scFv of the first or second antigen binding domains is linked directly or via a linker to the C-terminus of the fourth heavy chain.

[0739] In the present application,

[0740] 1) the third heavy chain comprises the VH of the first antigen-binding domain, and the fourth heavy chain comprises the scFv of the second antigen binding domain; or,

[0741] 2) the third heavy chain comprises the scFv of the first antigen binding domain, and the fourth chain comprises the VH of the second antigen binding domain, or;

[0742] 3) The third heavy chain comprises VH of the second antigen binding domain, and the fourth heavy chain comprises the scFv of the first antigen binding domain; or Attorney Docket No.: 769273: SA9-943PC

[0743] 4) The third chain comprises the scFv of the second antigen binding domain, and the fourth heavy chain comprises VH of the first antigen-binding domain.

[0744] In the present application, antigen-binding protein is selected from one of the following:

[0745] 1) light chain of SEQ ID NO:349, third heavy chain of SEQ ID NO:367 and fourth heavy chain of SEQ ID NO:368;

[0746] 2) light chain of SEQ ID NO:348, third heavy chain of SEQ ID NO:369 and fourth heavy chain of SEQ ID NO:370.

[0747] As shown in FIG. 1, the antigen-binding proteins of the present application may have the following structures (not including linkers):

[0748] (1) Heavy chain from N-terminal to C-terminal: VH of the second antigen binding domain, CHI, CH2, CH3,VH of the first antigen binding domain scFv, VL of the first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of the second antigen binding domain, light chain constant region;

[0749] (2) Heavy chain from N-terminal to C-terminal: VH of the second antigen binding domain, CHI, CH2, CH3,VL of the first antigen binding domain scFv, VH of the first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of the second antigen binding domain, light chain constant region;

[0750] (3) Heavy chain from N-terminal to C-terminal: VL of the first antigen binding domain scFv, VH of the first antigen binding domain scFv, VH of the second antigen binding domain, CHI, CH2, CH3; light chain from N-terminal to C-terminal: VL of the second antigen binding domain, light chain constant region.

[0751] (4) Heavy chain from N-terminal to C-terminal: VH of the first antigen binding domain, CHI, CH2, CH3,VH of the second antigen binding domain scFv, VL of the second antigen binding domain scFv; light chain from N-terminal to C-terminal: VL of the first antigen binding domain, light chain constant region;

[0752] (5) Heavy chain from N-terminal to C-terminal: VH of the first antigen binding domain, CHI, CH2, CH3, VL of the second antigen binding domain scFv, VH of the second antigen binding domain scFv; light chain from N-terminal to C-terminal: VL of the first antigen binding domain, light chain constant region; Attorney Docket No.: 769273: SA9-943PC

[0753] (6) Heavy chain from N-terminal to C-terminal: VL of the second antigen binding domain scFv, VH of the second antigen binding domain scFv, VH of the first antigen binding domain, CHI, CH2, CH3; light chain from N-terminal to C-terminal: VL of the first antigen binding domain, light chain constant region.

[0754] Specifically, the antigen-binding protein of the present application as shown in FIG. 1 may have the following structure (not counting linkers). Wherein specific first antigen binding domains and or second antigen binding domains of the species are shown in the Figure.

[0755] TLb0045, heavy chain from N-terminal to C-terminal: VH of the second antigen binding domain, CHI, CH2, CH3,VH of the first antigen binding domain scFv, VL of the first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of the second antigen binding domain, light chain constant region.

[0756] TLb0046, heavy chain from N-terminal to C-terminal: VL of first antigen binding domain scFv, VH of first antigen binding domain scFv, VH of second antigen binding domain, CHI, CH2, CH3; light chain from N-terminal to C-terminal: VL of second antigen binding domain, light chain constant region.

[0757] TLb0065, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of second antigen binding domain, light chain constant region.

[0758] TLb0066, heavy chain from N-terminal to C-terminal: VL of first antigen binding domain scFv, VH of first antigen binding domain scFv, VH of second antigen binding domain, CHI, CH2, CH3; light chain from N-terminal to C-terminal: VL of second antigen binding domain, light chain constant region.

[0759] TLb0067, Heavy chain from N-terminal to C-terminal: VH of the first antigen binding domain, CHI, CH2, CH3, VL of the second antigen binding domain scFv, VH of the second antigen binding domain scFv; Light chain from N-terminal to C-terminal: VL of the first antigen binding domain, light chain constant region. Attorney Docket No.: 769273: SA9-943PC

[0760] TLb0068 Heavy chain from N-terminal to C-terminal: VL of the second antigen binding domain scFv, VH of the second antigen binding domain scFv, VH of the first antigen binding domain, CHI, CH2, CH3; Light chain from N-terminal to C-terminal: VL of the first antigen binding domain, light chain constant region.

[0761] TLb0081, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of second antigen binding domain, light chain constant region.

[0762] TLb0085, heavy chain from N-terminal to C-terminal: VH of first antigen binding domain, CHI, CH2, CH3, VL of second antigen binding domain scFv, VH of second antigen binding domain scFv; light chain from N-terminal to C-terminal: VL of first antigen binding domain, light chain constant region.

[0763] TLb0086, heavy chain from N-terminal to C-terminal: VH of first antigen binding domain, CHI, CH2, CH3, VL of second antigen binding domain scFv, VH of second antigen binding domain scFv, ; light chain from N-terminal to C-terminal: VL of first antigen binding domain, light chain constant region.

[0764] TLb0087, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of second antigen binding domain, light chain constant region.

[0765] TLb0088, heavy chain from N-terminal to C-terminal: VH of first antigen binding domain, CHI, CH2, CH3, VL of second antigen binding domain scFv, VH of second antigen binding domain scFv; light chain from N-terminal to C-terminal: VL of first antigen binding domain, light chain constant region.

[0766] TLb0093, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VL of first antigen binding domain scFv, VH of first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of second antigen binding domain, light chain constant region. Attorney Docket No.: 769273: SA9-943PC

[0767] TLb0094, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VL of first antigen binding domain scFv, VH of first antigen binding domain scFv; light chain from N-terminal to VL of second antigen binding domain at C- terminal, light chain constant region.

[0768] TLbOOlll, VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of second antigen binding domain, light chain constant region.

[0769] TLb0045-l, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal VL of second antigen binding domain, light chain constant region.

[0770] TLb0045-2, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal second antigen binding domain, VL of light chain constant region.

[0771] TLb0045-3, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal second antigen binding domain, light chain constant region.

[0772] TLb0061-l, heavy chain from N-terminal to C-terminal: VH of second antigen binding domain, CHI, CH2, CH3,VH of first antigen binding domain scFv, VL of first antigen binding domain scFv; light chain from N-terminal to C-terminal second antigen binding domain, light chain constant region.

[0773] Recombinant Antibodies

[0774] In the present application, said antigen binding protein may include a heavy chain constant region, said heavy chain constant region may include a constant region derived from IgG or a constant region derived from IgY. Attorney Docket No.: 769273: SA9-943PC

[0775] For example, said antigen-binding protein may comprise a constant region derived from IgG. For example, said antigen-binding protein may comprise a heavy chain. The heavy chain may comprise a heavy chain variable region and a heavy chain constant region. The heavy chain variable region and said heavy chain constant region may be directly or indirectly linked. For example, said heavy chain variable region may be directly linked to said heavy chain constant region. For example, said heavy chain constant region may comprise a constant region derived from a protein selected from the following group: IgGl, IgG2, IgG3, and IgG4. In some embodiments, said heavy chain constant region may comprise a constant region derived from a protein selected from the following group: IgGl. Fc constant region of an antigenbinding protein may comprise an amino acid sequence as shown in SEQ ID NO:320 or SEQ ID NO:321 or SEQ ID NO:371, 373, 374, and CHI may be an amino acid sequence as shown in SEQ ID NO:319.

[0776] In the present application, said antigen binding protein may include a light chain constant region, said light chain constant region may include a constant region derived from IgK or a constant region derived from IgA. For example, said light chain constant region may include a Lambda light chain constant region, such as SEQ ID NO:318.

[0777] For example, the light chain constant region of said antigen-binding protein comprises the amino acid sequence shown in SEQ ID NO:318.

[0778] Peptides and immunoconjugates

[0779] In another aspect, the present application provides one or more polypeptides, which may comprise an isolated antigen-binding protein of the present application.

[0780] In another aspect, the present application provides one or more immunoconjugates, which may comprise an isolated antigen-binding protein of the present application. In some embodiments, the immunoconjugate also comprises a pharmaceutically acceptable therapeutic agent.

[0781] Nucleic Acid, Vector, and Cell

[0782] In another aspect, the present application also provides one or more isolated nucleic acid molecules that may encode the isolated antigen-binding proteins described herein. For example, Attorney Docket No.: 769273: SA9-943PC each of the one or more nucleic acid molecules may encode the entire antigen-binding protein or a portion thereof (e.g., one or more of the HCDR1-3 and the heavy chain variable regions).

[0783] The nucleic acid molecules described herein may be isolated. For example, it may be produced or synthesized by the following methods: (i) in vitro amplification, e.g., by polymerase chain reaction (PCR) amplification, (ii) clonal recombination, (iii) purification, e.g., by digestion and gel electrophoresis fractionation, or (iv) synthesis, e.g., by chemical synthesis. For example, the isolated nucleic acid may be a nucleic acid molecule prepared by recombinant DNA technology.

[0784] In the present application, the nucleic acids encoding the isolated antigen-binding proteins may be prepared by a variety of methods known in the art including, but not limited to, using reverse transcription PCR and PCR to obtain nucleic acid molecules of the isolated antigenbinding proteins described herein.

[0785] In another aspect, the present application provides one or more vectors comprising one or more nucleic acid molecules described herein. Each vector may comprise one or more of the nucleic acid molecules. In addition, the vector may also comprise other genes, such as marker genes that allow for selection of the vector in an appropriate host cell and under appropriate conditions. In addition, the vector may also comprise expression control elements that allow for the proper expression of the coding region in an appropriate host. Such control elements are well known to those skilled in the art and may include, for example, promoters, ribosome binding sites, enhancers, and other control elements that regulate gene transcription or mRNA translation, etc. In some embodiments, the expression control sequence is a regulatable element. The specific structure of the expression control sequences may vary depending on the function of the species or cell type, but typically comprise 5' non-transcribed sequences and 5' and 3' non-translated sequences involved in transcription and translation initiation, respectively, such as TATA cassettes, capping sequences, CAAT sequences, etc. For example, the 5' nontranscribed expression control sequence may comprise a promoter region, which may comprise a promoter sequence for transcription control of a functionally linked nucleic acid. The expression control sequences may also include enhancer sequences or upstream activator Attorney Docket No.: 769273: SA9-943PC sequences. In the present application, suitable promoters may include, for example, promoters for SP6, T3, and T7 polymerases, human U6RNA promoters, CMV promoters, and artificial hybrid promoters thereof (e.g., CMV), wherein some portion of the promoter may be fused to some portion of the promoter of a gene for another cellular proteins (e.g., human GAPDH, glyceraldehyde-3 -phosphate dehydrogenase), which may or may not comprise an additional intron. One or more nucleic acid molecules described herein can be operably linked to the expression control elements.

[0786] Such vectors may include, for example, plasmids, cosmids, viruses, phages, or other vectors commonly used in, for example, genetic engineering. For example, the vector may be an expression vector. For example, the vector may be a viral vector. The patient may be administered directly (in vivo) with the viral vector or may be administered indirectly, e.g., the patient may be administered with the cell treated with the virus in vitro (ex vivo). Viral vector technology is well known in the art and is described, for example, in Sambrook et al., (2001, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, New York) and other virology and molecular biology manuals. Conventional virus-based systems may include retroviral vectors, lentiviral vectors, adenoviral vectors, adeno-associated viral vectors, and herpes simplex viral vectors for gene transfer. In some cases, retroviral, lentiviral, and adeno- associated viral methods can be used to transfer and integrate gene into the host genome for long term expression of the inserted gene. Lentiviral vectors are retroviral vectors capable of transducing or infecting non-dividing cells and typically producing higher viral titers. Lentiviral vectors may comprise a long terminal repeat 5' LTR and a truncated 3' LTR, a RRE, a rev response element (cPPT), a central termination sequence (CTS), and / or a post- translational regulatory element (WPRE). The vectors described herein can be introduced into cells.

[0787] According to another aspect, the present application provides a cell. The cell may comprise an isolated antigen-binding protein as described herein, a polypeptide as described herein, an immunoconjugate as described herein, one or more nucleic acid molecules and / or one or more vectors as described herein. For example, each or every cell may comprise a Attorney Docket No.: 769273: SA9-943PC nucleic acid molecule or vector described herein. For example, each or every cell may comprise many (e.g., 2 or more) or multiple (e.g., 2 or more) kinds of nucleic acid molecules or vectors described herein. For example, the vectors described herein can be introduced into said host cells, such as prokaryotic cells (e.g., bacterial cells), CHO cells, NS / 0 cells, HEK293T cells, 293F cells, or HEK293A cells, or other eukaryotic cells, such as cells from plants, fungal or yeast cells, etc. The vectors described herein can be introduced into the host cells by methods known in the art, such as electroporation, lipofectine transfection, lipofectamin transfection, etc. For example, the cells may include yeast cells. For example, the cells may include d. coli cells. For example, the cells may include mammalian cells. For example, the cells may include immune cells.

[0788] The cells may include immune cells. In some cases, the cells may include immune cells. For example, the cells may include T cells, B cells, natural killer cells (NK cells), macrophages, NKT cells, monocytes, dendritic cells, granulocytes, lymphocytes, leukocytes and / or peripheral blood mononuclear cells.

[0789] Pharmaceutical Composition and Pharmaceutical Combination

[0790] In another aspect, the present application provides a pharmaceutical composition. The pharmaceutical composition may comprise the isolated antigen-binding protein, the polypeptide, the immunoconjugate, the isolated nucleic acid molecule, the vector, the cell, and / or the pharmaceutically acceptable adjuvant and / or excipient as described herein. In the present application, the pharmaceutically acceptable adjuvants may include buffers, antioxidants, preservatives, low molecular weight polypeptides, proteins, hydrophilic polymers, amino acids, sugars, chelating agents, counter ions, metal complexes and / or nonionic surfactants. Any conventional media or agent is contemplated for the pharmaceutical compositions of the present application, unless they are incompatible with the cells described herein. In the present application, the pharmaceutically acceptable excipients may include an additive other than the main drug in the pharmaceutical preparation, which may also be referred to as pharmaceutical necessities. For example, the excipients may include binders, fillers, disintegrants, and lubricants in tablets. For example, the excipients may include alcohol, Attorney Docket No.: 769273: SA9-943PC vinegar, medicinal juice, etc. in traditional Chinese medicine pills. For example, the excipients may include the base portion of semi-solid formulation ointments and creams. For example, the excipients may include preservatives, antioxidants, flavoring agents, perfuming agents, solubilizing assistant, emulsifiers, solubilizers, osmotic pressure regulators, and colorants in liquid formulations.

[0791] Kit, use and method

[0792] In one aspect, the present application provides a method for detecting or assaying TL1A, said method may comprise using said isolated antigen-binding protein or said peptide.

[0793] In the present application, said method may comprise an in vitro method, an ex vivo method, a method for non-diagnostic or non-therapeutic purposes.

[0794] For example, said methods may comprise methods for detecting the presence and / or amount of TLlA for non-diagnostic purposes, which may comprise the following steps:

[0795] 1) contacting a sample with an antigen-binding protein of the present application; and

[0796] 2) detecting the presence and / or amount of said antigen-binding protein bound by the sample to determine the presence and / or expression level of TLlA in a sample obtained from a subject.

[0797] In another aspect, the present application provides a kit for TL1A, which may comprise the use of said isolated antigen-binding protein or said peptide.

[0798] In the present application, said kit may also include instructions for use, said instructions for use documenting a method for detecting the presence and / or level of TL1A. For example, said methods may include in vitro methods, ex vivo methods, methods for non-diagnostic or non-therapeutic purposes.

[0799] In another aspect, the present application provides a use of said isolated antigen-binding protein or said polypeptide in the preparation of a kit, said kit being usable for methods for detecting the presence and / or the amount of TL1A. For example, said methods may include in vitro methods, ex vivo methods, methods for non-diagnostic or non-therapeutic purposes.

[0800] In another aspect, the present application provides a method of inhibiting the binding of

[0801] TL1A to DR3 / TR6 / DcR3 comprising administering to a subject in need thereof an effective Attorney Docket No.: 769273: SA9-943PC amount of said isolated antigen-binding protein, said peptide, said immuno-fix, said isolated nucleic acid molecule, said vector, and / or said cell. Said methods may be in vitro or ex vivo methods.

[0802] In another aspect, the present application provides a method for detecting or assaying a4p7, said method may comprise using said isolated antigen-binding protein or said peptide.

[0803] In the present application, said method may comprise an in vitro method, an ex vivo method, a method for non-diagnostic or non-therapeutic purposes.

[0804] For example, said methods may comprise methods for detecting the presence and / or amount of a4p7 for non-diagnostic purposes, which may comprise the following steps:

[0805] 1) contacting a sample with an antigen-binding protein of the present application; and

[0806] 2) detecting the presence and / or amount of said antigen-binding protein bound by the sample to determine the presence and / or expression level of a4p7 in a sample obtained from a subject.

[0807] In another aspect, the present application provides a kit for a4p7, which may comprise the use of said isolated antigen-binding protein or said peptide.

[0808] In the present application, said kit may also include instructions for use, said instructions for use documenting a method for detecting the presence and / or level of a4p7. For example, said methods may include in vitro methods, ex vivo methods, methods for non-diagnostic or non-therapeutic purposes.

[0809] In another aspect, the present application provides a use of said isolated antigen-binding protein or said polypeptide in the preparation of a kit, said kit being usable for methods for detecting the presence and / or the amount of a4p7. For example, said methods may include in vitro methods, ex vivo methods, methods for non-diagnostic or non-therapeutic purposes.

[0810] In another aspect, the present application provides a method of inhibiting the binding of a4p7 to MAdCAM-1 comprising administering to a subject in need thereof an effective amount of said isolated antigen-binding protein, said peptide, said immuno-fix, said isolated nucleic acid molecule, said vector, and / or said cell. Said methods may be in vitro or ex vivo methods. Attorney Docket No.: 769273: SA9-943PC

[0811] In another aspect, in the present application, said kits and / or said pharmaceutical compositions are used for the prevention, mitigation and / or treatment of a disease or condition.

[0812] In another aspect, the present application provides the use of a pharmaceutical composition in the preparation of a drug, said drug being used for the prevention, alleviation and / or treatment of a disease or condition.

[0813] In another aspect, the present application provides a method of preventing and / or treating a disease or condition comprising administering to a subject in need thereof said isolated antigen-binding protein, said isolated nucleic acid molecule, said vector, said cell, said pharmaceutical composition.

[0814] The disease or condition disclosed herein may be an inflammatory disease, a fibrostenotic disease, or a fibrotic disease. In some instances, the disease or the condition is a TL1A- mediated disease or condition. The term, “TLlA-mediated disease or condition” refers to a disease or a condition pathology or pathogenesis that is driven, at least in part, by TL1A signaling. In some instances, the disease or the condition is immune-mediated disease or condition, such as those mediated by TL1 A.

[0815] In some embodiments, the disease or the condition is an inflammatory disease or disorder that is mediated, at least in part, by TL1 A signaling. Non-limiting examples of inflammatory disease include, allergy, ankylosing spondylitis, asthma, atopic dermatitis, autoimmune diseases or disorders, cancer, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, diabetes (e.g., type 1 diabetes and type 2 diabetes), glomerulonephritis, gout, hepatitis (e.g., active hepatitis), an immune-mediated disease or disorder, inflammatory bowel disease (IBD) such as Crohn's disease and ulcerative colitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), multiple sclerosis, neurodegenerative diseases of aging, periodontal disease (e.g., periodontitis), preperfusion injury transplant rejection, psoriasis, pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis), rheumatic disease, scleroderma, sinusitis, tuberculosis.

[0816] In some embodiments, the disease or the condition is an autoimmune disease that is mediated, at least in part, by TL1A signaling. Non-limiting examples of autoimmune disease Attorney Docket No.: 769273: SA9-943PC or disorder include Achalasia, Addison's disease, Adult Still's disease, Agammaglobulinemia, Alopecia areata, Amyloidosis, Ankylosing spondylitis, Anti-GBM / Anti-TBM nephritis, Antiphospholipid syndrome, Autoimmune angioedema, Autoimmune dysautonomia, Autoimmune encephalomyelitis, Autoimmune hepatitis, Autoimmune inner ear disease (AIED), Autoimmune myocarditis, Autoimmune oophoritis, Autoimmune orchitis, Autoimmune pancreatitis, Autoimmune retinopathy, Autoimmune urticaria, Axonal & neuronal neuropathy (AMAN), Balo disease, Behcet's disease, Benign mucosal pemphigoid, Bullous pemphigoid, Castleman disease (CD), Celiac disease, Chagas disease, Chronic inflammatory demyelinating polyneuropathy (CIDP), Chronic recurrent multifocal osteomyelitis (CRMO), Churg-Strauss Syndrome (CSS) or Eosinophilic Granulomatosis (EGPA), Cicatricial pemphigoid, Cogan's syndrome, Cold agglutinin disease, Congenital heart block, Coxsackie myocarditis, CREST syndrome, Crohn's disease, Dermatitis herpetiformis, Dermatomyositis, Devic's disease (neuromyelitis optica), Discoid lupus, Dressier's syndrome, Endometriosis, Eosinophilic esophagitis (EoE), Eosinophilic fasciitis, Erythema nodosum, Essential mixed cryoglobulinemia, Evans syndrome, Fibromyalgia, Fibrosing alveolitis, Giant cell arteritis (temporal arteritis), Giant cell myocarditis, Glomerulonephritis, Goodpasture's syndrome, Granulomatosis with Polyangiitis, Graves' disease, Guillain-Barre syndrome, Hashimoto's thyroiditis, Hemolytic anemia, Henoch-Schonlein purpura (HSP), Herpes gestationis or pemphigoid gestationis (PG), Hidradenitis Suppurativa (HS) (Acne Inversa), Hypogammalglobulinemia, IgA Nephropathy, IgG4-related sclerosing disease, Immune thrombocytopenic purpura (ITP), Inclusion body myositis (IBM), Interstitial cystitis (IC), Juvenile arthritis, Juvenile diabetes (Type 1 diabetes), Juvenile myositis (JM), Kawasaki disease, Lambert-Eaton syndrome, Leukocytoclastic vasculitis, Lichen planus, Lichen sclerosus, Ligneous conjunctivitis, Linear IgA disease (LAD), Lupus, Lyme disease chronic, Meniere's disease, Microscopic polyangiitis (MPA), Mixed connective tissue disease (MCTD), Mooren's ulcer, Mucha-Habermann disease, Multifocal MotorNeuropathy (MMN) or MMNCB, Multiple sclerosis, Myasthenia gravis, Myositis, Narcolepsy, Neonatal Lupus, Neuromyelitis optica, Neutropenia, Ocular cicatricial pemphigoid, Optic neuritis, Palindromic Attorney Docket No.: 769273: SA9-943PC rheumatism (PR), PANDAS, Paraneoplastic cerebellar degeneration (PCD), Paroxysmal nocturnal hemoglobinuria (PNH), Parry Romberg syndrome, Pars planitis (peripheral uveitis), Parsonage-Turner syndrome, Pemphigus, Peripheral neuropathy, Perivenous encephalomyelitis, Pernicious anemia (PA), POEMS syndrome, Polyarteritis nodosa, Polyglandular syndromes type I, II, III, Polymyalgia rheumatica, Polymyositis, Postmyocardial infarction syndrome, Postpericardiotomy syndrome, Primary biliary cirrhosis, Primary sclerosing cholangitis, Progesterone dermatitis, Psoriasis, Psoriatic arthritis, Pure red cell aplasia (PRCA), Pyoderma gangrenosum, Raynaud's phenomenon, Reactive Arthritis, Reflex sympathetic dystrophy, Relapsing polychondritis, Restless legs syndrome (RLS), Retroperitoneal fibrosis, Rheumatic fever, Rheumatoid arthritis, Sarcoidosis, Schmidt syndrome, Scleritis, Scleroderma, Sjogren's syndrome, Sperm & testicular autoimmunity, Stiff person syndrome (SPS), Subacute bacterial endocarditis (SBE), Susac's syndrome, Sympathetic ophthalmia (SO), Takayasu's arteritis, Temporal arteritis / Giant cell arteritis, Thrombocytopenic purpura (TTP), Tolosa-Hunt syndrome (THS), Transverse myelitis, Type 1 diabetes, Ulcerative colitis (UC), Undifferentiated connective tissue disease (UCTD), Uveitis, Vasculitis, Vitiligo, and Vogt-Koyanagi-Harada Disease.

[0817] In some embodiments, the disease or the condition is a cancer that is mediated, at least in part, by TL1 A signaling. Non-limiting examples of cancers include Adenoid Cystic Carcinoma, Adrenal Gland Cancer, Amyloidosis, Anal Cancer, Ataxia-Telangiectasia, Atypical Mole Syndrome, Basal Cell Carcinoma, Bile Duct Cancer, Birt Hogg Dube Syndrome, Bladder Cancer, Bone Cancer, Brain Tumor, Breast Cancer, Breast Cancer in Men, 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, Leukemia — Acute Lymphoblastic Leukemia, Leukemia — Acute Lymphocytic (ALL), Leukemia — Acute Myeloid AML, Leukemia — Adult, Leukemia — Childhood, Leukemia — Chronic Lymphocytic (CLL), Leukemia — Chronic Myeloid (CIVIL), Liver Cancer, Lobular Carcinoma, Lung Cancer, Lung Cancer — Small Cell (SCLC), Lung Cancer — Non-small Cell Attorney Docket No.: 769273: SA9-943PC

[0818] (NSCLC), Lymphoma — Hodgkin's, Lymphoma — Non-Hodgkin's, Malignant Glioma, Melanoma, Meningioma, Multiple Myeloma, Myelodysplastic Syndrome (MDS), Nasopharyngeal Cancer, Neuroendocrine Tumor, Oral Cancer, Osteosarcoma, Ovarian Cancer, Pancreatic Cancer, Pancreatic Neuroendocrine Tumors, Parathyroid Cancer, Penile Cancer, Peritoneal Cancer, Peutz-Jeghers Syndrome, Pituitary Gland Tumor, Polycythemia Vera, Prostate Cancer, Renal Cell Carcinoma, Retinoblastoma, Salivary Gland Cancer, Sarcoma, Sarcoma — Kaposi, Skin Cancer, Small Intestine Cancer, Stomach Cancer, Testicular Cancer, Thymoma, Thyroid Cancer, Uterine (Endometrial) Cancer, Vaginal Cancer, and Wilms' Tumor.

[0819] In some embodiments, the disease or the condition is an inflammatory bowel disease, such as Crohn's disease (CD) or ulcerative colitis (UC). A subject may suffer from fibrosis, fibrostenosis, or a fibrotic disease, either isolated or in combination with an inflammatory disease. In some cases, the CD is severe CD. The severe CD may result from inflammation that has led to the formation of scar tissue in the intestinal wall (fibrostenosis) and / or swelling. In some cases, the severe CD is characterized by the presence of fibrotic and / or inflammatory strictures. The strictures may be determined by computed tomography enterography (CTE), and magnetic resonance imaging enterography (MRE). The disease or condition may be characterized as refractory, which in some cases, means the disease is resistant to a standard treatment (e.g., anti-TNFa therapy). Non-limiting examples of standard treatment include glucocorticosteriods, anti-TNF therapy, anti-a4-b7 therapy (vedolizumab), anti-IL12p40 therapy (ustekinumab), Thalidomide, and Cytoxin.

[0820] In the present application, the subject may include a human or non-human animal. For example, the non-human animal may be selected from the group consisting of: a monkey, a chicken, a goose, a cat, a dog, a mouse, and a rat. In addition, a non-human animal may also include any animal species other than human, such as livestock animals, or rodents, or primates, or domestic animals, or poultry animals. The human can be Caucasian, African, Asian, Sumerian, or other ethnicity, or a hybrid of various ethnicities. As another example, the human may be the elderly, adults, adolescents, children or infants. Attorney Docket No.: 769273: SA9-943PC

[0821] Effective amounts in human can be extrapolated from the effective amounts in experimental animals. For example, Freireich et al. described the interrelationship of dosages for animals and humans (based on milligrams per meter squared of body surface) (Freireich et al., Cancer Chemother. Rep. 50, 219 (1966)). Body surface area can be approximately determined from the height and weight of the patient. See, e.g., Scientific Tables, Geigy Pharmaceuticals, Ardsley, N.Y, 537 (1970).

[0822] Without intending to be bound by any theory, the following examples are intended merely to illustrate the fusion proteins, methods of preparation, uses, etc., of the present application and are not intended to limit the scope of the present application.

[0823] On the other hand, the present application provides a method for downregulating the expression of a4p7 / TLl A, comprising: providing an isolated antigen-binding protein that can specifically bind to TL1 A. In some embodiments, the method further comprises: providing an antigen-binding protein that can specifically bind to a4p7. In some embodiments, the method further comprises: providing a TL1A protein. In some embodiments, the isolated antigenbinding protein that can specifically bind to TL1A and the antigen-binding protein that can specifically bind to a4p7 are the same protein. In some embodiments, the same protein is the aforementioned antigen-binding protein.

[0824] On the other hand, the present application provides a method for inducing endocytosis of cell surface a4p7 / TLlA, comprising: providing an isolated antigen-binding protein that can specifically bind to TL1 A. In some embodiments, the method further comprises: providing an antigen-binding protein that can specifically bind to a4p7. In some embodiments, the method further comprises: providing a TL1A protein. In some embodiments, the isolated antigenbinding protein that can specifically bind to TL1A and the antigen-binding protein that can specifically bind to a4p7 are the same protein. In some embodiments, the same protein is the aforementioned antigen-binding protein.

[0825] In the present application, the isolated antigen binding protein can induce the endocytosis of cell surface a4p7 / TLlA. In some embodiments, the endocytosis of the protein can be determined by a fluorescent labeling method. For example, the efficiency of the antigen binding Attorney Docket No.: 769273: SA9-943PC protein in inducing the endocytosis of cell surface a4p7 / TLl A can be obtained by detecting the fluorescence intensity of the cell using flow cytometry. In some cases, the isolated antigen binding protein can induce the endocytosis of a4p7 / TLlA with an efficiency of about 50% or more.

[0826] On the other hand, the present application provides the use of the antigen binding protein to induce the endocytosis of cell surface a4p7 / TLl A.

[0827] On the other hand, the present application provides the use of the antigen binding protein to downregulate the expression of a4p7 / TLl A.

[0828] Without being limited by any theory, the following examples are only for illustrating the fusion protein, preparation method and use of the present application, and are not intended to limit the scope of the invention of the present application.

[0829] Example

[0830] Example 1 anti-TLIA and antibody «4p7 monoclonal antibody sequence

[0831] Anti-TLIA monoclonal antibodies: Anti-TLIA monoclonal antibodies were screened using TLA0015-11 and TLA0238-5 monoclonal antibodies, and their VH and VL amino acid sequences, scFv configuration sequences, and scFv sequences after the addition of disulfide bonds are shown in Table 1. Anti-a4p7 monoclonal antibody: Anti- Anti-a4p7 monoclonal antibody uses Vedolizumab, whose VH and VL amino acid sequences, scFv configuration sequences, and scFv sequences after the addition of disulfide bonds are shown in Table 1. In addition, the amino acid sequences of human IgGl antibody CK, CHI and Fc, and the sequences of Fc mutants are shown in Table 2.

[0832] Table 1 Amino acid sequence of TLA0015-ll,TLA0238-5 and Vedolizumab Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0833] Table2. Amino acid sequence of the constant region of Human IgGl Attorney Docket No.: 769273: SA9-943PC

[0834] Example 2 design of anti-TLlAx«4p7 bispecific antibody

[0835] Anti-TLIA antibodies TLA0015-11 and TLA0238-5 were assembled with vedolizumab monoclonal antibodies into bispecific antibody molecules of different configurations, the amino acid sequences of each bispecific antibody are shown in Table 3A and 3B, and the schematic configuration of each bispecific antibody is shown in Figure 1.

[0836] Table 3 A. Bispecific antibody amino acid sequence with format 2+2 Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0837] Table 3B. Bispecific antibody amino acid sequence with format 1+1 Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0838] Example 3 Expression of anti-TLlAx«4p7 bispecific antibody

[0839] The designed bispecific antibody was expressed, and the purity of the bispecific antibody with a purity of less than 95% after one-step purification was purified in two steps, and the expression yield and purity of the bispecific antibody were counted in Table 4. Most of the bispecific antibodies express a volume of 100 ml, and about 20 mg of purity can be obtained to meet the requirements of the bispecific antibody.

[0840] Table 4. Bispecific antibody expression details Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

[0841] Example 4 Prediction of sites where TLA0015 binds to TL1A antigen

[0842] The Al-based antigen-antibody complex structure prediction model predicts the complex structure of the TL1 A monomer crystal structure and the monoclonal antibody TLA0015, and obtains a high-quality complex structure. Its binding schematic diagram in different directions is shown in Figure 2. TLA0015 binds to the TL1A monomer, but it will shift toward the gap between the two monomers in the TLlAtrimer, so it has good activity in blocking the binding of TLlA to the receptor DR3. In addition, based on the results of the complex structure, the key sites for TLA0015 monoclonal antibody to bind to the TL1 A antigen were obtained. These key sites on the TL1 A antigen include: R32, Q33, P35, T36, Q37, P44, S89, K102, P103, D161, 1162, S163, and L164.

[0843] Example 5 ELISA detects the binding ability of bispecific antibody to TL1A protein

[0844] The binding ability of a plurality of bispecific antibodies prepared by example 1-3 to human TL1 A (purchased from Acrobiosy stems, Cat. No. TLA-H5243) was detected based on an ELISA assay. The binding capacity of different concentrations of bispecific antibodies is determined by comparing the binding curves of different concentrations to TL1 A-his protein.

[0845] Human TL1 A-his protein was diluted to 2 pg / ml in PBS at 100 pl / well and added to a 96- well plate and coated overnight at 4 °C. The next day, remove the coating solution, add 5% BS A blocking solution at 200 pl / well, and incubate for 2 hours at room temperature. Remove the blocking solution, wash three times at 250 pl / well with 0.5%o PBST, and dilute the bispecific antibody to 100 nM with the blocking solution, 10-fold dilution to form 8 concentration gradients (maximum concentration 100 nM) at lOOpl / The wells were added to the closed plate and incubated for 60 minutes at room temperature. After washing the plate 3 times with PBST, pat it dry and incubate at 100 pl / well with 10,000-fold dilution of HRP- Attorney Docket No.: 769273: SA9-943PC labeled goat anti-human IgG secondary antibody for 60 min at room temperature. Wash the plate 5 times with 0.5%o PBST at 100 pl / well, add TMB solution, incubate at room temperature in the dark for 10 min, at 100 pl / well, add stop solution, stop the substrate chromogenic reaction, read the OD value at 450 nm with a microplate reader, analyze and plot the data with a GraphPad and calculate EC50. As shown in Figure 3, the bispecific antibody has good binding activity to the TL1 A protein.

[0846] Example 6 Biacore detects the affinity of the bispecific antibody with the TL1A and a 4p7 protein

[0847] The affinity of the bispecific antibodies prepared by the present invention through example 1-3 with human TLlA-his protein, monkey TLlA-his and human a4p7 protein is detected based on the Biacore biomolecular interaction instrument.

[0848] A certain amount of antibody to be tested was captured affinity using the biosensing chip Protein A (Cytiva, 29127556), and then a series of different concentrations of antigen (50 nM, 25 nM, 12.5 nM, 6.25 nM, 3.125 nM, 1.5625 nM) were passed on the surface of the chip to obtain binding and dissociation curves by detecting the reaction signal in real time with Biacore (Cytiva, IK). After the completion of each cycle of dissociation, the biochips are washed and regenerated with 10 mM glycine-hydrochloric acid solution pH 1.5 (Cytiva, BR-1003-54). The experimental data were fitted to a 1 : 1 model using Biacore Insight Evaluation Software 5.0 to calculate the value of the affinity. The results are shown in Figures 4 and 5 and Table 5. TLb0045 and TLb0065 bispecific antibodies have high affinity for both human TL1A and monkey TL1 A proteins. The affinity results of Vedolizumab (analog) and bispecific antibody TLb0045-l to human a4p7 are shown in Figures 6A-6B and Table 6. The affinity of TLb0045- 1 to human a4p7 is comparable to that of Vedolizumab (analog). Attorney Docket No.: 769273: SA9-943PC

[0849] Table 5 Affinity of antibodies to TL1 A proteins of different species

[0850] Table 6 Affinity of antibodies to human a4p protein

[0851] Example 7 FACS detects the ability of bispecific antibody to block the binding of TLlA-his to 293-DR3 cells

[0852] The ability of multiple bispecific antibodies prepared by example 1-3 to block the binding of hu TLlA-his protein to human DR3 (DR3: UniprotKB NO. Q93038) expressed on the surface of Expi293 cells was detected based on flow cytometry assay. The blocking ability of Attorney Docket No.: 769273: SA9-943PC huTLIA-his protein is determined by comparing the curves of different antibody concentrations blocking huTLIA-his protein to 293-huDR3 cells.

[0853] The DNA sequence of the receptor DR3 extracellular domain protein encoding TL1 A was cloned into the mDeZ-TM (with transmembrane sequence and Zeocin resistance gene) vector, transiently transfected into Expi293 cells, and 100 pg / mL Zeocin (final concentration) was added 24 h later for 7 days to obtain the Expi293 cell line (293-huDR3) expressing human DR3 on the cell surface. The huTLIA-his-Biotin protein was obtained by Biotin conjugation using a biotin labeling kit (ThermoFisher, A35389). FACS Binding was used to determine the EC80 concentration of protein binding between huTLIA-his-Biotin protein and 293-huDR3 cells. Wash 293-huDR3 cells three times with staining buffer (PBS + 2% FBS (Gibco, Cat #10091148) + 5 mM EDTA (Gibco, Cat #15575020)) at 300 g each for 5 min and discard the supernatant. Resuspend cells in PBS, adjust the cell density to 5^ 105 cells / mL, 100 pl / well, add to a 96- well plate, centrifuge to remove the supernatant, and add 2*EC80 concentration of huTLlA- his-Biotin protein, 50 pl / well. Dilute the bispecific antibody and the positive control antibody to 400 nM in 2-fold increments over 12 gradients (up to a final concentration of 200 nM), 50pl / well, add to a 96-well plate and mix well with huTLIA-his-Biotin protein as well as 293- huDR3 cells. Incubate at 4 °C for 30 min. Cells are washed twice in PBS to remove unbound antibodies to be tested. Add another 100 pl / well of Streptavidin-PE secondary antibody and incubate at 4 °C for 30 min. Centrifuge at 300 g for 5 min and wash the cells twice in PBS to remove unbound secondary antibody. Finally, the cells are resuspended in 200 pl PBS, and the ability of multiple antibodies to block the binding of TL1 A protein to 293-huDR3 cells is determined by a Beckman Coulter CytoFLEX flow cytometer. The resulting data were fitted and analyzed by GraphPad Prism software. As shown in Figure 7A, the antibodies all have the ability to block the binding of TLlA-his protein to 293-DR3 cells. In addition, there was no difference in the blocking activity between TLb0045 and TLb0045-l ( Figure 7B ).

[0854] In addition, 10 nM of a4p7 protein (purchased from Acrobiosy stems, item number MAM- H52H4) was added at the same time as incubation of the bispecific antibody with huTLl A- his-biotin protein and 293-DR3 cells, and was used to verify that binding of a4p7 protein to Attorney Docket No.: 769273: SA9-943PC the bispecific antibody interfered with the binding of the bispecific antibody to huTLlA-his- biotin. protein, the results are shown in Figure 8, a4p7 protein binding to the bispecific antibodies does not interfere with the blocking activity of the bi specific antibodies on the TL1 A protein.

[0855] Example 8 detects the ability of bispecific antibody to inhibit TLlA-DR3-activated caspase3 / 7

[0856] A method based on the Caspase-Gio® 3 / 7 Assay System Activity Assay Kit (Promega, Cat#G8092) was used to assay the ability of the plurality of bispecific antibodies of the present invention, prepared by means of Examples 1-3, to inhibit the TLlA-induced apoptosis signaling pathway in TF-1 (endogenously expressing DR3) cells. The inhibitory capacity was determined by comparing the curves of inhibition of caspase3 / 7 activity in TF-1 cells with different antibody concentrations.

[0857] The huTLIA-his protein was configured to a 4X working concentration of 16nM; the Cycloheximide small molecule drug was configured to a 4X working concentration of lOpg / ml. 25 pl of the mixture was added to each well in a white-bottomed 96-well plate. The starting concentration of the antibody was configured as 200nM (final concentration was 50nM), 4X dilution of four concentration points, then 2X dilution for a total of 10 concentrations, and 25 pl of each well was added to the white bottom 96-well plate. Pre-incubate the above mixture in a 37°C incubator for Ih. At the same time, centrifuge the TF-1 cells (300g for 5min) and resuspend them with buffer (1640 medium+10% FBS) and count them, and adjust the density of cells to 1E6 cells / ml. When the incubation time of the antigen-antibody is over, take out the white 96-well flat-bottomed plate, and inoculate TF-1 cells according to the adjusted density of 50pl / well, and then add 25 pl of each well into the white bottom 96-well plate. After incubation, take out the white 96-well flat-bottom plate, inoculate with the adjusted density of TF-1 cells at 50pl / well, and continue to incubate for 6h in a carbon dioxide incubator. After incubation, take out the plate from the incubator and equilibrate the plate at room temperature for about 30min, and take out the Caspase-Gio® 3 / 7 Assay System reagent beforehand and Attorney Docket No.: 769273: SA9-943PC equilibrate the plate at room temperature. Add Caspase-Gio® 3 / 7 Assay System reagent, 50pl per well, shake the bed at 300rpm for 30s, and then leave it for 15-30min to make the cells fully lysed, and then carry out the reading of the RLU value on a multifunctional enzyme labeling instrument, and then carry out a four-parameter fit plot of the logarithm of the sample concentration to the measured RLU value with Graphpad. The results are shown in Figure 9 and Table 7, the bispecific antibodies such as TLb0045, TLb0065 and TLb0081 all functioned well to inhibit Caspase3 / 7 activity.

[0858] Table 7. IC50 of the inhibitory effect on TL1 A-activated Caspase3 / 7 activity

[0859] Example 9 Fluorescent reporter method to detect the ability of bispecific antibodies to inhibit the NF-KB signaling pathway activated by TL1A

[0860] A luciferase reporter gene-based method was used to test the ability of multiple bispecific antibodies obtained by the present invention prepared by Examples 1-3 to inhibit the NF-KB signaling pathway in TF-1 (endogenously expressing DR3) cells activated by the TLlA-his protein. The TF-1-NFKB-LUC cell line was purchased from Yoshimitsu Biotechnology Ltd (GM-C30289, H_TNFSF15(TL1A) Reporter Cell Line) and screened using NF -KB-Luciferase lentiviral infection of TF-1 cells. In this study, TF 1-NFKB-1UC cells were used to detect the biological activity of the DR3-NF KB signaling pathway activated by TLlA-his protein by bispecific antibody. In this experiment, 3 x of the analytical concentration of huTLIA-his antigen solution was prepared with analytical buffer (RPMI 1640 medium + 1% FBS) at 0.9 Attorney Docket No.: 769273: SA9-943PC pg / ml (final concentration of 0.3 pg / ml) and added to a white 96-well flat-bottom plate at 40 pl / well. Prepare bispecific antibodies at 3 - analytical concentrations with a maximum concentration of 300 nM (final concentration of 100 nM) and a 5-fold serial dilution for a total of 8 concentration points. After the antibody is diluted, add 40 pl / well to the white 96-well flatbottom plate described above, then place the white 96-well flat-bottom plate in a carbon dioxide incubator, and pre-incubate the bispecific antibody and antigen for Ih. During bispecific antibody and antigen pre-incubation, TF 1-NFKB-1UC cells were centrifuged (300 g, 5 min) and then resuspended with analysis buffer and counted, adjusting the cell density to 5E5 cells / ml. After the antigen and antibody incubation time was over, the white 96-well flat-bottom plate was removed, andTFl-NFKB-luc cells with adjusted density were inoculated at 40 pl / well, and placed in a carbon dioxide incubator for 6 h. At the end of the incubation, remove the white 96- well flat-bottom plate from the incubator, add 60 pl of Bio-Lite Luciferase Assay reagent (Novozan, DD 1201-02) to each well, shake slightly for 5 minutes, and read the RLU values on a multimode microplate reader. The four-parameter fitting plot of the logarithm of the sample concentration with Graphpad was performed with the measured signal value, and the results are shown in Figure 10: TLb0045, TLb0065, and TLb0081 bispecific antibodies all have good activity in inhibiting the NF-KB signaling pathway activated by TL1 A-his protein.

[0861] Example 10 ELISA detects the binding ability of bispecific antibody to «4p7 protein

[0862] Based on the ELISA assay method to detect the binding ability of a plurality of bispecific antibodies prepared by example 1-3 of the present invention and human a4p7 protein (purchased from Acrobiosystems, Cat. No. IT7-H52W4). The binding capacity of different bispecific antibodies is determined by comparing the binding curves of different bispecific antibodies to a4p7 protein.

[0863] Dilute a4p7 protein to 2 pg / ml with coating solution (PBS + 1 mM CaC12 + 1 mM MgC12) at 100 pl / well, add to the microplate and incubate overnight at 4 °C. Remove the coating solution and incubate at 200pl / well with 5% B SA blocking solution (containing 1 mM MnC12) for 2 hours. Remove the blocking solution, wash three times at 250 pl / well with 0.5%o PBST Attorney Docket No.: 769273: SA9-943PC

[0864] (containing 1 mM MnC12), and dilute the bispecific antibody to 100 nM with the blocking solution, 10-fold dilution to form 8 concentration gradients (maximum concentration 100 nM) at lOOpl / The wells were added to the closed plate and incubated for 60 minutes at room temperature. Wash the plate 3 times with 0.5%o PBST (containing 1 mM MnC12) and pat dry, add HRP-labeled goat anti -human IgG antibody at 100 pl / well, and incubate for 60 min at room temperature. Wash the plate 5 times with 0.5%o PBST (containing 1 mM MnC12) at 100 pl / well, add TMB solution, incubate for 10 min at room temperature in the dark, at 100 pl / well, add stop solution, stop the substrate chromogenic reaction, read the OD value at 450 nm with a microplate reader, analyze and plot the data with a GraphPad and calculate EC50. As shown in Figure 11, the bispecific antibodies have good binding activity to the a4p7 protein, as with the vedolizumab monoclonal antibody.

[0865] Example 11 FACS was used to detect the binding ability of bispecific antibody to cells endogenously expressing «4p7

[0866] The binding ability of a plurality of bi specific antibodies prepared by the present invention through example 1-3 to cells endogenously expressing a4p7 is detected based on flow cytometry assay. Hut 78 cells (purchased from the Cell Bank of the Chinese Academy of Sciences, Cat# TCHu206) are a T lymphocyte line with a4p7 molecules endogenously expressed on the cell membrane surface. The binding capacity of different concentrations of antibodies is determined by comparing their binding curves with those of the Hut78 cell line.

[0867] In this experiment, Hut 78 cells endogenously expressing a4p7 were used as target cells. Wash cells three times with staining buffer (PBS + 2% FBS (Gibco, Cat #10091148) + 5 mM EDTA (Gibco, Cat #15575020)) at 450 g each time for 5 min and discard the supernatant. Resuspend cells in staining buffer, adjust cell density to 1 ^ 106 cells / mL, 100 pl / well, and add to a 96-well plate. Bispecific antibody and positive control antibody were diluted to 200 nM, 10-fold stepwise dilution of 8 gradients (maximum concentration 200 nM), lOOpl / well, added to a 96-well plate, and mixed well with Hut 78 cells. After 30 min of incubation at 4 °C, the cells are washed twice in PBS to remove unbound antibodies to be tested. Add another 100 Attorney Docket No.: 769273: SA9-943PC pl / well of goat anti-human IgG-PE and incubate at 4 °C for 30 min. Centrifuge at 300 g for 5 min and wash the cells twice in PBS to remove unbound secondary antibody. Finally, the cells were resuspended in 200pl PBS, and the binding capacity of the bispecific antibody to Hut 78 cells was determined by Beckman Coulter CytoFLEX flow cytometry. The resulting data were fitted and analyzed by GraphPad Prism software. As shown in Figure 12, both TLb0045 and TLb0065 bispecific antibodies bind well to Hut 78 cells, as with the vedolizumab monoclonal antibody.

[0868] Example 12 ELISA detection of the binding of bispecific antibody blocking «4p7 to the ligand MAdCAM-1

[0869] The ability of multiple bispecific antibodies prepared by example 1-3 to block the binding of a4p7 protein and the ligand MAdCAM-1 protein (purchased from Acrobiosystems, Cat. No. MAM-H5253) is detected by ELISA assay by comparing different concentrations of bispecific antibodies. The curve of a4p7 protein binding to the ligand MAdCAM-1 was used to determine its blocking ability.

[0870] Biotin conjugation of MAdCAM-l-Fc protein was performed using a biotin labeling kit (ThermoFisher, A35389) to obtain MAdCAM-1 -Fc-Biotin protein backup. Dilute a4p7 protein to 2 pg / ml with PBS (containing 1 mM CaC12 and 1 mM MgC12) at 100 pl / well, add to the microplate and incubate overnight at 4 °C. Remove the coating solution and incubate at 200pl / well with 5% BSA blocking solution for 2 h at room temperature. Remove the blocking solution and wash three times at 250 pl / well by adding PBST wash (PBST containing 0.5%o Tween-20 and 1 mM MnC12) and pat dry for later use. Dilute the bispecific antibody to 20 nM with blocking solution, form 8 concentration gradients (maximum concentration of 20 nM) with 3-fold dilution, and add sequentially to the blocked plate at 50 pl / well, while dilute the MAdCAM-1 -Fc-Biotin protein to 1 pg / ml using blocking solution, add sequentially to the plate at 50 pl / well, and incubate for 1 hour at room temperature. Wash the plate 3 times with PBST wash containing MnC12 (remove residual droplets with absorbent paper) at lOOpl / wells, add SA-HRP secondary antibody containing 5,000-fold dilution, and incubate for 60 min at room Attorney Docket No.: 769273: SA9-943PC temperature. Wash the plate 3 times with PBST wash containing MnC12 at 100 pl / well, add TMB solution, incubate for 10 min at room temperature in the dark, at 100 pl / well, add stop solution, stop the substrate chromogenic reaction, read the OD value at 450 nm with a microplate reader, analyze the data with GraphPad and calculate EC50. As shown in Figure 13, both TLb0045 and TLb0065 bispecific antibodies have a good ability to block the binding of a4p7 to MAdCAM-1, as with vedolizumab monoclonal antibodies.

[0871] Example 13 Detection of bispecific antibody blockade of MAdCAM-l-mediated cell adhesion

[0872] The blocking ability of MAdCAM-1 -mediated cell adhesion function mediated by multiple bispecific antibodies prepared by example 1-3 was detected based on CTG bioluminescence detection technology, and its blocking ability was determined by comparing the curves of different antibody concentrations blocking MAdCAM-1 -mediated adhesion of Hut 78 cells.

[0873] In this experiment, Hut 78 cells endogenously expressing a4p7 were used as target cells. MAdCAM-1 -Fc protein was diluted to 2 pg / ml in PBS at 100 pl / well and added to a 96-well plate and coated overnight at 4 °C. Remove the coating solution and incubate at 200pl / well with 5% BSA blocking solution for 2 h at room temperature. At the end of the incubation, remove the blocking solution and wash three times with PBST wash at 250 pl / well and pat dry for later use. Dilute cultured Hut 78 cells to 3E5 / ml, 100 pl / ml with FACS buffer (DPBS+1% BSA+1 mM MnC12). The wells were plated into a 96-well cell plate, and the supernatant was discarded by centrifugation at 450 g for 5 min; At the same time, dilute the antibody to 100 nM with FACS buffer, dilute 4-fold to form 12 concentration gradients, add to a 96-well cell plate, 100 pl / well, resuspend the cells, and then transfer to a coated microplate plate, incubate in a cell culture incubator for 40 min, centrifuge at 450g for 5 min, discard the supernatant, wash the cells five times with FACS buffer, and then add CTG (Vazyme, Cat#DDl 101-02), 50pl / wells, microplate reader for reading. Analyze the data with GraphPad, graph it, and calculate the EC50. As shown in Figure 14, TLb0045, TLb0065, and TLb0081 bispecific antibodies all have a good blockade of MAdCAM-1 -mediated cell adhesion function, as shown in Figure 12. Attorney Docket No.: 769273: SA9-943PC

[0874] At the same time as the cell adhesion experiment, TL1 A protein was added and incubated with bispecific antibody and Hut 78 cells to verify the effect of TL1A protein on the cell adhesion function of bispecific antibody blockade. As shown in Figure 15, the addition of TL1A protein had little effect on the blockade of MAdCAM-1 -mediated cell adhesion by bispecific antibodies.

[0875] Example 14 Detection of the internalization activity of the bispecific antibody

[0876] The method based on pH Dye is used to detect the internalization activity of a plurality of bispecific antibodies prepared by example 1-3 of the present invention. The internalization capacity of bispecific antibodies is determined by comparing the internalization signal curves of different concentrations.

[0877] Antibody pH Dye labeling: antibodies were labeled using pHAb Amine Reactive Dye (Promega, Cat#G9845) reagent. Take 120 pg of the antibody to be labeled and add it to a 30 kDa protein concentrate tube, then add 400 pl of 10 mM sodium bicarbonate buffer (pH 8.5), centrifuge at 7500 rpm for 5 min, repeat the above steps twice (total 3 changes), centrifuge to a volume of about 100 pl; transfer the antibody to a 1.5 mL centrifuge tube. Add 1 pl of pHAb Amine Reactive Dye solution to each tube, mix thoroughly, and shake the reaction at low speed for 60 minutes at room temperature and away from light; transfer the antibody to a 30 kDa protein concentrate tube, add 400 pl of PBS, centrifuge at 7500 rpm for 5 minutes, and repeat the above steps for 4 times (totaling 5 changes of solution); centrifuge until the volume of the solution is about 100 pl, and then transfer the antibody to a 30 kDa protein concentrate tube. Transfer the antibody to a 1.5 mL centrifuge tube, keep it away from light, test the absorbance of A280 and A532 of the labeled antibody, calculate the concentration of the labeled antibody, and set aside.

[0878] The cultured Hut 78 cells were centrifuged at 300 g for 5 min, the supernatant was discarded, resuspended in freshly prepared RPMH640 medium, the cell density was adjusted to 2E6 / mL, and the cell density was adjusted to a 96-well low-adsorption U bottom plate, a volume of 50pl was added to each well, and the bispecific antibody and the positive control antibody were diluted to 66 nM with RPMH640 medium. Dilute 11 gradients (maximum final Attorney Docket No.: 769273: SA9-943PC concentration 33 nM) in 3-fold steps, 50pl / well, add to a 96-well plate, mix well with Hut 78 cells, and place in a 37°C incubator for incubation. The cell plates incubated for 6 h and 24 h were removed, centrifuged at 450 g for 5 min, the supernatant was discarded, 200 pl of FACS buffer was added to each well, 450 g was centrifuged for 5 min, the supernatant was discarded, and the step was repeated. Prepare Live / Dead dye (Invitrogen, CAT#L34976) at a ratio of 1 : 1,000, add 100 pl per well, and incubate at 4 °C for 30 min. After the end of the incubation, centrifuge at 450 g for 5 min, discard the supernatant, repeat this step twice, resuspend the cells with 100 pl FACS buffer, and detect the PE channel by flow cytometry. Analyze the data with GraphPad, graph it, and calculate the EC50. The 6-hour internalization is shown in Figure 16A, and the 24-hour internalization result is shown in Figure 16B, and the bispecific antibodies have good internalization activity.

[0879] In addition, during the incubation of the internalization experimental antibody with Hut 78 cells, 10 nM TL1A protein was added to verify the effect of TL1A protein on the internalization activity of the bispecific antibody, the internalization results of 6 hours are shown in Figure 17A, and the results of internalization in 24 hours are shown in Figure 17B, and the internalization activity of the bispecific antibody TLb0045 and TLb0065 can be significantly improved after the addition of TL1 A protein. The results of the endocytic activity of the bispecific antibody TLb0045-l on Hut 78 cells are shown in Figure 17C. The addition of TL1 A protein significantly enhanced the endocytic activity of TLb0045-l. Using healthy human peripheral blood mononuclear cells, Pan T cells were sorted and tested for the endocytic activity of the bispecific antibody on T cells. The results of the endocytic activity of the bispecific antibody TLb0045-l on primary T cells are shown in Figure 17D. Consistent with the endocytic results on Hut 78 cells, the addition of TL1 A protein significantly enhanced the endocytic activity of TLb0045-l. Attorney Docket No.: 769273: SA9-943PC

[0880] Example 15 Detection of bispecific antibody endocytosis downregulation of «4p7 expression

[0881] Based on flow cytometry assay, the effect of internalization of a plurality of bispecific antibodies prepared by example 1-3 of the present invention on the cell membrane and the total a4p7 expression of cells is detected.

[0882] Cultured Hut 78 cells were centrifuged at 450 g for 5 min, the supernatant was discarded, resuspended in freshly prepared RPMI1640 medium, the cell density was adjusted to 4E6 / mL, and added to a 96-well low-adsorption U bottom plate, with a volume of 25pl per well. Dilute pH Dye-labeled bispecific antibody and positive control antibody to 10 nM in RPMH640 medium, 50pl / well and add to a 96-well plate. TL1A protein was diluted to 40 nM with RPMH640 medium and 25pl / well was added to the 96-well plate described above; For wells that do not require TL1 A protein, add 25pl of RPMH640 medium, mix well with Hut 78 cells, and incubate in a 37°C incubator for 16 hours (three 96-well plates, one for internalization activity, another for a4p7 expression on the membrane, and the last for total a4p7 expression).

[0883] Internalization activity assay: After the end of incubation, the first cell plate was removed, 450g centrifuged for 5min, the supernatant was discarded, 200pl FACS Buffer was added to each well, 450g centrifuged for 5min, the supernatant was discarded, and the step was repeated. Configure the Live / Dead dye at a ratio of 1 : 1,000, add 100 pl per well, and incubate at 4 °C for 30 min. After the end of incubation, centrifuge at 450 g for 5 min, discard the supernatant, repeat this step twice, resuspend the cells using 100 pl FACS buffer, and FACS to detect antibody internalization activity. The data were analyzed and graphed with GraphPad, and the results are shown in Figure 18, where the vedolizumab monoclonal antibody has internalization activity and is not affected by the TL1A protein; TLA0015-11 monoclonal antibody has no internalization activity; Internalization in the Vedolizumab+TLA0015-ll combo group is mediated by Vedolizumab monoclonal antibody and is therefore also not affected by the TL1 A protein; TLb0045-l and TLb0065-l bispecific antibodies have good internalization activity in the absence of TL1A protein, and the increase of internalization activity is significantly enhanced after the addition of TL1A protein. The reason is that after the bispecific antibodies Attorney Docket No.: 769273: SA9-943PC

[0884] TLb0045-l or TLb0065-l bind to the TL1A antigen, they will promote endocytosis activity better through cross-linking. As shown in Figure 19, the endocytosis activity of the bispecific antibodies increased significantly after the addition of TL1A protein. The reason is that when the TL1 A monoclonal antibodies in these four bispecific antibodies bind to the TL1 A protein, they will form a complex antigen-antibody complex through cross-linking. Therefore, complex antigen-antibody complexes will also be formed in the bispecific antibodies through crosslinking, thereby promoting endocytosis.

[0885] Detection of a4p7 expression on the membrane: after the end of incubation, the second cell plate was removed, 450g centrifuged for 5min, the supernatant was discarded, 200 pl FACS Buffer was added to each well, 450g was centrifuged for 5min, the supernatant was discarded, and the step was repeated; Configure the Live / Dead dye at a ratio of 1 : 1,000 while adding 9.87 pl of Vedolizumab (at a concentration of 4.56 mg / mL) for a final concentration of 100 nM, adding 100 pl per well, and incubating at 4 °C for 60 min; Centrifuge at 450 g for 5 min, discard the supernatant, add 200 pl of FACS Buffer per well, and repeat this step three times; Dilute AF488 Goat anti-human Fc secondary antibody at a ratio of 1 :200, add 100 pl per well, and incubate at 4 °C for 30 min; Centrifuge at 450 g for 5 min, discard the supernatant, add 200 pl of FACS Buffer per well, repeat this step three times, resuspend the cells with 100 pl FACS Buffer, and detect the fluorescence signal by flow cytometry. The data were analyzed and graphed with GraphPad, and the results are shown in Figure 20, which shows that Vedolizumab monoclonal antibody has internalization activity and can down-regulate the expression of a4p7 on Hut 78 membrane, and is not affected by TL1A protein. Internalization in the Vedolizumab+TLA0015-ll combo group is mediated by Vedolizumab monoclonal antibody and is therefore also not affected by the TL1 A protein; TLb0045-l and TLb0065-l bispecific antibodies have good internalization activity, so in the absence of TL1 A protein, the expression of a4p7 on the membrane can be down-regulated, and after the addition of TL1 A protein, the internalization activity increases, and the expression of a4p7 on the membrane can be more significantly down-regulated. Attorney Docket No.: 769273: SA9-943PC

[0886] Detection of total a4p7: the second cell plate was removed at the end of incubation, centrifuged at 450g for 5min, discarded the supernatant, 200pl of FACS Buffer was added to each well, centrifuged at 450g for 5min, discarded the supernatant, and repeated the step once; Live / Dead dye was configured at a ratio of 1 : 1000, and at the same time, 9.87pl of Vedolizumab (concentration of 4.56 mg / mL), the final concentration of 100 nM, 100 pl of each well, incubation at 4 °C for 60 min; 450g centrifugation for 5 min, discard the supernatant, 200 pl of FACS Buffer per well, repeat the step three times; 100 pl of 2% PFA Fix Solution per well, incubation at room temperature Add lOOpl 2% PFA Fix Solution per well, incubate at room temperature for lOmin, centrifuge at 600g for 5min, discard the supernatant, add 200pl FACS Buffer per well, repeat the step twice; add lOOpl 0.1% Triton X-100 per well, incubate at ice for 7min, centrifuge at 600g for 5min, discard the supernatant, add 200pl FACS Buffer per well, repeat the step twice; add lOOpl 0.1% Triton X-100 per well, incubate at a final concentration of lOOnM Add Vedolizumab at a final concentration of 100 nM to each well and incubate at 4°C for 60 min; after incubation, centrifuge at 600g for 5 min, discard the supernatant, and add 200 pl of FACS Buffer to each well and repeat the procedure three times; Dilute the AF488 Goat anti -human Fc secondary antibody at a ratio of 1 :200 and add 100 pl to each well and incubate for 30 min at 4°C, and then add the AF488 Goat anti-human Fc secondary antibody to each well. After incubation for 30 min at °C, centrifuge at 600 g for 5 min, discard the supernatant, add 200 pl of FACS Buffer to each well, repeat the procedure three times, resuspend the cells with 100 pl of FACS Buffer, and detect the fluorescence signal by flow- through. The data were analyzed and plotted using GraphPad, and the results without TL1A protein are shown in Figure 21A. TLb0045-l and TLb0065-l bispecific antibodies downregulated the expression of total a4p7 as well as Vedolizumab monoclonal antibody. With the addition of TL1A protein, the results are shown in Figure 21B, the bispecific antibodies TLb0045-l and TLb0065-l could significantly down-regulate the expression of total a4p7 and were superior to Vedolizumab monoclonal antibody. Attorney Docket No.: 769273: SA9-943PC

[0887] Example 16 The reverted expression of «4p7 on the membrane after internalization of bispecific antibody was detected

[0888] Based on flow cytometry assay, the relationship between the amount of a4p7 reverting to expression and time on the cell membrane after internalization of a plurality of bispecific antibodies prepared by example 1-3 of the present invention is detected.

[0889] Cultured Hut 78 cells were centrifuged at 450 g for 5 min, the supernatant was discarded, resuspended in freshly prepared RPMI1640 medium, the cell density was adjusted to 4E6 / mL, and added to a 96-well low-adsorption U bottom plate, with a volume of 25 pl per well. pH Dye-labeled bispecific antibody and positive control antibody were diluted to 10 nM and 100 nM in RPMH640 medium, respectively, and 50pl / well were added to a 96-well plate. TL1A protein was diluted to 40 nM with RPMH640 medium and 25pl / well was added to the 96-well plate described above; For wells that do not require TL1A protein, add 25pl of RPMH640 medium, mix well with Hut 78 cells, and incubate in a 37°C incubator for 24 hours. After the end of incubation, 450 g was centrifuged for 5 min, the supernatant was discarded, and the cells were washed twice using freshly prepared RPMH640 medium. The first group is incubated with Vedolizumab-488 fluorescently labeled antibody at 4 °C for 60 min. After the end of incubation, centrifuge at 450g for 5min, discard the supernatant, wash the cells twice with flow cytometry, add an appropriate amount of flow cytometry buffer to resuspend the cells, and detect the fluorescence signal by flow cytometry. The second, third, and fourth groups were supplemented with 200 pl of RPMH640 medium per well, and on the second, third, and fourth days, the newly expressed a4p7 molecule on the membrane was detected using Vedolizumab- 488 fluorescently labeled antibody, respectively.

[0890] The results of antibody incubation concentration of 5 nM are shown in Figure 22A (fluorescent signal), after 3 days of washing off the incubated antibody, the a4p7 molecule on the cell membrane surface of the cell membrane basically returned to the original level, and the addition of TL1A protein had no effect on the expression of a4p7 molecule on the membrane surface; Vedolizumab + TLA0015-11 in combination with vedolizumab monoclonal antibody results; The expression of a4p7 molecule on the membrane of TLb0045-l bispecific antibody Attorney Docket No.: 769273: SA9-943PC did not return to the original level after 3 days of washing off the incubated antibody The results indicated that TLb0045-l bispecific antibody had a stronger inhibitory effect on a4p7 expression on the membrane surface than that of Vedolizumab monoclonal antibody group and Vedolizumab+TLA0015-ll group. TLb0045-l had a strong inhibitory effect on the expression of a4p7 molecules on the upper surface of the membrane when TL1 A protein was added, and the expression of a4p7 molecules on the membrane was still at a very low level after 3 days of washing off the incubated antibody and TL1 A protein. The results of the proportion of positive cells are shown in Figure 22B, and the trend of the results is consistent with the results of the fluorescence signal in Figure 22 A, TLb0045-l has a good inhibition effect on the expression of a4p7 molecule, and the inhibition effect will be more obvious when TL1 A protein is added.

[0891] The results of the antibody incubation at 50 nM are shown in Figure 23A (fluorescent signal) and Figure 23B (proportion of positive cells), and the trend is consistent with the results of the 5 nM incubation, but due to the increase in antibody concentration, the expression of a4p7 molecule on the membrane surface has not returned to the original level after 3 days of washing off the incubation antibody. Among them, TLb0045-l bispecific antibody had a stronger inhibitory effect on the expression of a4p7 molecule on the membrane than that of Vedolizumab monoclonal antibody group and Vedolizumab + TLA0015-11 group, and the inhibitory effect was more obvious after the addition of TL1 A protein.

[0892] At the same time, PanT cells isolated from PBMC cells were used to detect the recovery of a4p7 expression on the membrane. The results were consistent with the results on HuT 78. As shown in Figures 24A and 24B, after the addition of TL1 A protein, the inhibitory effect of TLb0045-l bispecific antibody was more obvious. On day 3, the expression of Free a4p7 was still not detected on the cell surface.

[0893] Example 17 Detection of the inhibitory activity of bispecific antibodies on PanT cell migration

[0894] Based on the Trans-well experimental method, the inhibitory activity of multiple bispecific antibodies prepared by Examples 1-3 of the present invention on T cell migration was detected. Attorney Docket No.: 769273: SA9-943PC

[0895] Cell adhesion and Trans-well combined experiment: MAdCAM-1 protein was diluted to 2pg / ml with PBS, lOOpl / well, added to the chamber, and coated overnight at 4°C. The next day, the antibody was diluted with assay buffer (DPBS (w / o Ca2+,Mg2+)+l% BSA+0.5mM MnC12) to prepare an antibody concentration of 20nM (final concentration of 5nM); TL1A protein was prepared at a concentration of 40nM (final concentration of lOnM). According to the experimental design, 30pl of the above-mentioned antibody and 30pl of TL1 A protein were added to each well of a 96-well round-bottom plate (30pl assay buffer was added to the group without TL1A), and the 96-well round-bottom plate was placed on ice. Then, Pan T was resuspended in assay buffer (DPBS (w / o Ca2+,Mg2+)+l% BSA+0.5mM MnC12) to a density of 3.33x 106 cells / mL, and 60pl / well was inoculated in the above-mentioned 96-well roundbottom plate. After mixing, incubation was carried out on ice for 30 minutes. During the incubation process, the unbound proteins were washed away from the chamber coated with MAdCAM-1 his with PBST (PBS+0.5%o Tween-20) for three times. After the incubation, lOOpl of the mixture in the 96-well round-bottom plate was transferred to the chamber coated with MAdCAM-1 and incubated on ice for 45 minutes. After the incubation time, the unbound Pan T-antibody mixture in the chamber was washed away with assay buffer and washed 4 times. Then add lOOpl of DPBS+2%FBS medium to the chamber, and add 600pl of DPBS+2%FBS+ 500ng / ml CCL25 medium to the 24-well plate below the chamber. Incubate the 24-well plate in a carbon dioxide incubator overnight. On the third day, carefully remove the chamber from the 24-well plate, add 300pl of Bio-Lite Luciferase Assay reagent (Novozyme, DD1201-02) to each well, shake gently for 5 minutes, transfer 200pl to each well in a new 96-well white plate, and then read the RLU value on a multifunctional microplate reader. Use Graphpad to make a bar graph of the sample and the measured signal value. The results of the combined cell adhesion and Trans-well experiment are shown in Figure 25. Vedolizumab (Analogue), TLb0045-l and TLA0015-ll+Vedolizumab can all be seen to inhibit T cell migration.

[0896] Combined experiment of a4p7 recovery expression on membrane surface and Trans-well: The a4p7 recovery experiment on membrane surface in Example 16 was combined with the

[0897] Trans-well experiment. On day 0 and day 3, the a4p7 recovery expression on membrane Attorney Docket No.: 769273: SA9-943PC surface was detected and the Trans-well experiment was performed. The results of the 16-hour endocytosis experiment are shown in Figure 26, which shows that TL1A protein can specifically enhance the endocytosis activity of TLb0045-l and TLb0045-2 bispecific antibodies; the expression levels of a4p7 on the membrane surface of PanT cells on Day 0 and Day 3 are shown in Figures 27A and 27B, which shows that TL1A protein can specifically enhance the inhibitory activity of TLb0045-l and TLb0045-2 bispecific antibodies on the restoration of a4p7 expression on the membrane surface; the results of the Trans-well experiment on Day 0 are shown in Figure 28A, which shows that on Day 0, Vedolizumab monoclonal antibody, TLA0015-11 + Vedolizumab combination, TLb0045-l and TLb0045-2 bispecific antibodies have basically the same inhibitory activity on T cell migration; The results of the Trans-well experiment on Day 4 are shown in Figure 28B. It can be seen that the inhibitory activity on T cell migration in the monoclonal antibody group and the monoclonal antibody combination group is very weak in the presence or absence of TL1 A because the expression of a4p7 on the membrane has been restored; TLb0045-l and TLbO45-2 bispecific antibodies are equivalent to monoclonal antibodies in the absence of TL1 A, but in the presence of TL1 A protein, it can be seen that the bispecific antibodies still have a strong inhibitory effect on T cell migration. The experiment was repeated on another PanT (Donor 2), and the results were completely consistent. The specific endocytosis results are shown in Figure 29, the expression level of a4p7 on the membrane surface is shown in Figures 30A and 30B, and the results of Trans-well day 0 and day 4 are shown in Figures 31 A and 3 IB, respectively.

[0898] Example 18 Detection of the inhibitory activity of bispecific antibody on CD4T cells

[0899] In this study, the inhibitory ability of bispecific antibodies on immune cell activation was evaluated by measuring the amount of cytokines in CD4+ T cells isolated from cryopreserved healthy human peripheral blood and their culture supernatants.

[0900] Dilute Anti-Human CD3 (Aero, CDE-M120a) with HBSS at a concentration of 5 ug / ml, add 96 cell culture flat-bottom plates, add 100 ul per well, and coat overnight at 4°C. Cryopreserved healthy human PBMCs (purchased from Saili Bio) were recovered into RPMI 1640 complete medium and incubated overnight at 37°C. The next day, after centrifugation of Attorney Docket No.: 769273: SA9-943PC

[0901] PBMCs (500 g for 5 min), discard the supernatant, resuspend the cells with EasySep™ Buffer (StemCell, Cat#20144), count them, and adjust the cell density to 5E7 cells / mL. CD4+ T cells were sorted according to the kit instructions using the CD4 negative selection kit (StemCell, Cat#17952). CD4+ T cells were washed twice with PBS, centrifuged at 500 g each for 5 min and discarded supernatant. Resuspend CD4+ T cells in RPMI 1640 complete medium, count, adjust cell density to 5E5 cells / mL for later use. Serial dilution of antibodies with 1640 complete medium, up to a maximum concentration of 400 nM, 10-fold dilution for a total of 5 points, set aside. Dilute huTLIA-his with 1640 complete medium at a concentration of 40 nM for later use. After the coated 96-well plate was washed once with 200 ul / well HBSS, the diluted MadCAM-l-his (Aero, MAM-H52H4-1 mg) diluted with HBSS was added at a concentration of 10 nM, 100 ul per well was added, and incubated at 37 degrees for 1 hour. After incubation, wash once with 200 ul / well HBSS, add CD4+ T cells lOOpl per well (final concentration is 5 w cells / well); Add huTLIA-his, 50 pl per well (final concentration is 10 nM); Add antibody 50 ul per well (final concentration up to 100 nM). Remove after 4 days of incubation at 37 °C. Centrifuge at 500 g for 5 min, aspirate the cell supernatant and place on ice for subsequent cytokine detection.

[0902] Cells were stained using PE anti-human CD25 (BioLegend, 356104) and Live / Dead dye and incubated at 4°C for 30min protected from light; FACS buffer was washed twice, and the membrane was fixed and ruptured using eBioscience™ Foxp3 / Transcri ption Factor Flow Fixation Rupture Kit (Invitrogen, 00-5523-00) according to the instructions. 00-5523-00) for membrane fixation and rupture, resuspend the cells in 200pl, incubate at 4°C for 60min under light protection; 1 x rupture solution working solution was washed once, 1 x rupture solution working solution diluted with flow-through antibody Alexa Fluor® 488 anti-human Ki-67 Antibody (BioLegend, 350508), incubate at room temperature under light protection for The cells were incubated for 30 min at room temperature and protected from light, washed twice with 1 x membrane-breaking solution working solution and the supernatant was discarded, and finally the cells were resuspended with 100 pl FACS buffer, and the fluorescence signals of CD25 and Ki-67 were detected by cell flow meter. The FACS results of the assay are shown in Attorney Docket No.: 769273: SA9-943PC

[0903] Figure 32A-C. TLb0045-l bispecific antibody had a very good inhibitory effect on both proliferation and activation of CD4T cells, and was superior to Vedolizumab monoclonal antibody and TLA0015-11 monoclonal antibody.

[0904] Cytokine IFN-y levels in the supernatant were also assayed using the BD™ Cytometric Bead Array (human IFN-y) kit. Configure the standards according to the instructions, dilute the samples, 50pl / well, and add to a 96-well plate. Configure Capture beads solution, 50pl / well per well, add to 96-well plate and co-incubate the standard / sample with Capture beads solution for 1 hour. Dilute the Detection Antibody solution according to the instructions, 50pl / well per well, add to the 96-well plate and co-incubate for 2 hours. Centrifuge at 300g for 5 minutes, discard supernatant and wash again with wash buffer. After centrifugation again at 300g for 5 minutes, the supernatant was discarded and resuspended in wash buffer, 70pl / well, and the amount of cytokine IFN-y was measured by Beckman CytoFLEX flow cytometer. The data obtained were fitted and analyzed by GraphPad Prism software, and the results of the IFN-y concentration assay are shown in Figure 33, which shows that the TLb0045-l bispecific antibody has a good inhibitory effect on the secretion of IFN-y.

[0905] Example 19 LDH method to detect the T cell-dependent cytotoxic killing effect mediated by bispecific antibodies

[0906] This experiment uses the LDH method to detect whether bispecific antibodies will produce T cell-dependent cytotoxic killing effects. One end of the TLb0045-l bispecific antibody targets TL1 A, which is mainly expressed by endothelial cells, such as umbilical vein endothelial cells (HUVECs), monocytes, macrophages, dendritic cells (DC), T cells, chondrocytes and synovial fibroblasts, among which HUVECs have the highest expression level; the other end targets integrin « 4 f> 7, which is mainly expressed on the surface of lymphocytes and is responsible for mediating lymphocytes from the blood circulation into the intestines and central nervous system. Its abnormal function is closely related to human autoimmune diseases. In this experiment, TL1A overexpressing cell lines were co-incubated with PBMC under the condition of adding bispecific antibodies, and then LDH detection was Attorney Docket No.: 769273: SA9-943PC used to evaluate whether bispecific antibodies would produce T cell -dependent cytotoxic killing effects.

[0907] The target cells used in the experiment were CHO-K1 (purchased from Nanjing Kebai Biotechnology) and CHO-K1-TL1A (overexpression of human full-length TL1A was constructed on CHO-K1 cells), and the effector cells were PBMCs from different donors. CHO-K1 and CHO-K1-TL1A cells were first digested and centrifuged (300g, 5min), resuspended in F-12K complete medium and counted, the cell density was adjusted to 1E5 cells / ml, and 100 1 / well was inoculated in a 96-well flat-bottom plate and cultured overnight. The next day, the antibody was diluted with 1640 complete medium to prepare an antibody of 2 X analytical concentration, with a top dose of 2 u M (final of 1 u M), 10-fold gradient dilution, and a total of 9 concentration points. After the antibody was diluted, the 96-well flatbottom plate inoculated with CHO-K1-TL1A cells was removed, the culture supernatant was aspirated, and 80 u 1 of the above-diluted antibody was added to each well. It is necessary to set up 2 target cell only wells (Tmax and Tmin wells) and 1 target cell + PBMC well (Buffer well). Continue to pre-incubate the 96-well flat-bottom plate in the CO2 incubator for 30 minutes. During the pre-incubation process, centrifuge the PBMCs that were revived one day in advance (300g, 5min), resuspend and count them with 1640 complete medium, and adjust the cell density to 2.5E6 cells / ml. After the pre-incubation time is over, take out the 96-well flat-bottom plate and inoculate the PBMCs with the adjusted density at 80 u 1 / well. Take out a new 96-well plate and add 80 u 1 of the above-diluted antibody and 80 u 1 of PBMCs with the adjusted density to each well. Place the three groups of 96-well plates in the CO2 incubator and continue to incubate for 48h. 30min~lh before the end of incubation, add 20 u 1 of 10 X Lysis buffer (Tmax) to one of the target cell only wells and continue to incubate in the CO2 incubator. After the incubation, the 96-well flat-bottom plate was removed from the incubator, centrifuged (400g, 5min), and 50 u 1 of the culture supernatant was aspirated from each well into a new 96-well flat-bottom plate. 50 u 1 of CytoTox 96® Non-Radioactive Cytotoxicity Assay (promega, G1780) substrate solution was added to each well of the 96-well flat-bottom plate. After incubation at room temperature and in the dark for 20min, 50 u 1 of Stop Solution Attorney Docket No.: 769273: SA9-943PC was added to each well after the color development time to terminate the reaction, and the absorbance value at 490nm was detected by an enzyme reader. The killing ratio was calculated as follows: Lysis %=(OD490Sample - OD490Buffer) / (OD490Tmax- OD490Tmin) X 100%. The logarithm of the antibody concentration was taken using Graphpad to perform a four- parameter fitting plot with the killing ratio. The killing results are shown in Figure 33, where the killing of CHO-K1 cells after co-incubation with PBMC antibodies is shown in Figure 34A. Except for the positive control, other antibodies have no killing effect; the killing of CHO-K1- TL1 A cells after co-incubation with PBMC antibodies is shown in Figure 34B. Except for the positive control, other antibodies have no killing effect; the results of co-incubation of simple antibodies with PBMC are shown in Figure 34C. Except for the positive control, other antibodies have no killing effect.

[0908] Bispecific antibody-mediated T cell activation detection: Prepare antibody mixture: Brilliant Violet 785™ anti-human CD3 Antibody (BioLegend, Cat#344842), Brilliant Violet 421™ anti-human CD8 Antibody (BioLegend, Cat#344748), BD Pharmingen™ APC Mouse Anti-Human CD69 (BD, Cat#555533), BD Pharmingen™ PE Mouse Anti-Human CD25 (BD, Cat#555432) at a ratio of 1 :200, and dilute them with FACS buffer (PBS+1%FBS) for later use. In the above T cell-dependent cytotoxic killing experiment, the cell pellet after centrifugation (400g, 5min) was resuspended with antibody mixture, incubated at 4 °C for 30min, washed with PBS to remove unbound antibodies 3 times, and finally resuspended with 80 u 1 PBS. The samples were collected on a flow cytometer (Agilent, NovoCyte Penteon) to analyze the proportion of CD25+CD69+ in the CD4+ cell population and the proportion of CD25+CD69+ in the CD8+ cell population. The four-parameter fitting plot was performed with Graphpad after taking the logarithm of the antibody concentration and the CD25+CD69+ ratio. The different T cell activation detection after CHO-K1 cells were co-incubated with PBMC antibodies is shown in Figure 35; the different T cell activation detection after CHO-K1-TL1 A cells were co-incubated with PBMC antibodies is shown in Figure 36; the results of the simple antibody co-incubated with PBMC are shown in Figure 37. The above experimental results are Attorney Docket No.: 769273: SA9-943PC consistent. Except for the positive control, other antibodies have no function of activating T cells.

[0909] Cytokine release detection mediated by bispecific antibodies: For the above T celldependent cytotoxic killing experiment, after centrifugation (400g, 5min), 50 u 1 of culture supernatant was aspirated from each well into a new 96-well V-bottom plate, and then processed according to the instructions of BD™ Cytometric Bead Array (CBA) Human Thl / Th2 Cytokine Cytometric Bead Array (CBA) Kit II (BD, Cat# 551809). Samples were collected on a flow cytometer (Agilent, NovoCyte Penteon), analyzed using FCAP Array v3 software, and finally Graphpad was used to perform a four-parameter fitting plot of the antibody concentration and the concentration of each cytokine. The specific results are shown in Figures 38 and 39. After CHO-K1 cells were co-incubated with PBMC antibodies, and after CHO-K1-TL1A cells were co-incubated with PBMC antibodies, the release of IL-2, IL-6, IL- 10, TNF- a , and IFN- y cytokines was not induced.

[0910] Example 20 Detection of bispecific antibody Tm value detection

[0911] The experiment is based on a Panta instrument to detect the Tm values of a plurality of bispecific antibodies prepared by example 1-3 of the present invention. The Prometheus Panta instrument (NanoTemper Technologies GmbH) includes micro-differential scanning fluorescence (nanoDSF) technology, which tracks the folding state of a protein by detecting its endogenous fluorescence. The ratio of the fluorescence signal changes with increasing temperature or chemical denaturant concentration, thus determining the protein stability parameter Tm value. The specific Tm values of each antibody are shown in Table 8.

[0912] Table 8. The body Tm value of the anti-TLl A antibody Attorney Docket No.: 769273: SA9-943PC

[0913] Example 21 Detection of bispecific antibody pharmacokinetics in rats

[0914] In this experiment, SD rats were used to study the pharmacokinetics of bispecific antibodies in rats. At the same time, the stability of bispecific antibody molecules in rats was detected. Female rats with SPF grades were purchased from Weitong Lihua Laboratory Animal Technology Co., Ltd., and after the animals were quarantined, 5 mice were randomly assigned for each antibody. Each rat was injected with Img / kg of bispecific antibodies TLb0045 and TLb0065 through the tail vein, and blood was subsequently collected at the time points in Table 9. Serum was collected by lOOpl / time point / only by jugular vein, placed at room temperature for 1 h, and centrifuged (8000 rpm, centrifugation for 5 min) at room temperature.

[0915] The first detection method: recombinant human TL1 A-his protein was diluted to 1 pg / ml with PBS, added to the microplate at 100 pl / well, and incubated overnight at 4 °C. Remove the coating solution, block at 200 pl / well, add 5% skim milk, and incubate for 2 h at room temperature. Remove the blocking solution and wash three times at 250 pl / well with 0.5%o PBST and pat dry for later use. Dilute the serum sample to the appropriate ratio using the blocking solution and add it to the blocked plate. The standard curves were added sequentially to the blocked plate at 100 pl / well using TLb0045 and TLb0065 starting at 100 nM for a total of 20 concentration gradients (up to 100 nM), respectively, and incubated for 1 h at room temperature. Wash the plate 3 times with 0.5%oPBST, add 10,000-fold dilution of HRP-labeled goat anti -human IgG antibody, 100 pl / well, and incubate for 60 min at room temperature. Wash the plate 5 times with 0.5%oPBST, add TMB at 100 pl / well, incubate at room temperature in the dark for 5 min, at 100 pl / well, add stop solution, stop the substrate chromogenic reaction, read the OD value at 450 nm with a microplate reader, analyze the data with a GraphPad, plot and calculate the half-life (T 1 / 2).

[0916] At the same time, the second method was used to quantify the antibody concentration in rat serum: first, Biotin conjugation of a4p7 protein was performed using a biotin labeling kit to obtain a4p7-Biotin protein for backup. Recombinant human TL1 A-his protein is diluted to 1 pg / ml in PBS at 100 pl / well, added to the plate and incubated overnight at 4 °C. Remove the Attorney Docket No.: 769273: SA9-943PC coating solution, block at 200 pl / well, add 5% skim milk, and incubate at room temperature for 2 h. Remove the blocking solution and wash three times at 250 pl / well with 0.5%o PBST and pat dry for later use. The diluted serum sample is added to the closed microplate label. The standard curves were added sequentially to the blocked plate at 100 pl / well using TLb0045 and TLb0065 starting at 100 nM for a total of 20 concentration gradients (up to 100 nM), respectively, and incubated for 1 h at room temperature. Wash the plate 3 times with 0.5%oPBST, dilute a4p7-Biotin protein to 2 pg / ml and incubate at 100 pl / well for 60 min at room temperature; Add 5,000-fold diluted SA-HRP solution, incubate at room temperature for 60 min, wash the plate 5 times with 0.5%oPBST at 100 pl / well, add TMB, and incubate at room temperature at 100 pl / well in the dark. The stop solution was added, the substrate chromogenic reaction was stopped, the OD value at 450 nm was read with a microplate reader, the data was analyzed with a GraphPad, plotted and the half-life (Tl / 2) was calculated.

[0917] As shown in Fig. 40 and Fig. 41, the concentrations of bispecific antibody TLb0045 and TLb0065 in serum were basically the same as those detected by the two detection methods, indicating that the bispecific antibody molecule was stable in rats and was not easy to break. When TLb0045 and TLb0065 bispecific antibodies were compared together, there was little difference in serum concentrations, as shown in Figure 42. The individual parameters of the PK calculation are shown in Table 10, and the half-life of the bispecific antibody is about 6-8 days.

[0918] Table 9. PK blood collection time point Attorney Docket No.: 769273: SA9-943PC

[0919] Table 10. PK test of TLb0045 and TLb0065 in rats

[0920] Example 22 Detection of pharmacokinetics of bispecific antibody in hFcRn mice

[0921] In this experiment, B-hFcRn transgenic mice were used to study the pharmacokinetics of bispecific antibodies. FcRn is the receptor for antibody IgG, and its binding is pH-dependent. In a weakly acidic environment, the IgG-Fc segment exhibits a high affinity for FcRn, while in a neutral or alkaline environment, the IgG-Fc segment exhibits a low affinity for FcRn. When IgG is non-specific pinocytosis by cells, it enters the acidic environment of the endosome, and shows high affinity binding to FcRn on the endosomal membrane, and then is transported to the cell membrane by the circulating endosome, at this time, under alkaline conditions, the IgG- Fc segment has a very low affinity with FcRn, and IgG is released from the FcRn into the blood circulation, thereby increasing the half-life of IgG. When M428L and N434S mutations are introduced into IgG-Fc, the binding ability of Fc to FcRn receptor can be enhanced, and the half-life of IgG can be better extended. B-hFcRn transgenic mice (Catalog: 110001) were purchased from Biocytogen, and the mouse FcRn gene was knocked out on the C57BL / 6 strain mice, and the human FcRn gene was knocked in, and the transgenic mouse only expressed the human FcRn receptor protein, but not the mouse FcRn receptor, so it can be used for metabolic kinetic studies of antibody drugs. Attorney Docket No.: 769273: SA9-943PC

[0922] Female B-hFcRn transgenic mice with SPF grade were purchased from Biocytogen (Beijing) Pharmaceutical Technology Co., Ltd., and after the animals were quarantined, 10 mice were randomly assigned to each antibody, divided into 2 groups, 5 mice in each group, and blood was collected alternately. Each mouse is injected with 1 mg / kg of bispecific antibody via the tail vein and blood is then collected alternately at the time points in Table 11. Blood was collected by mandibular vein by collecting 20 pl / time point / piece, leaving it at room temperature for 1 h, and centrifugation (8000 rpm, centrifugation for 5 min) at room temperature.

[0923] The first detection method: recombinant human TL1 A-his protein was diluted to 1 pg / ml with PBS, added to the microplate at 100 pl / well, and incubated overnight at 4 °C. Remove the coating solution, block at 200 pl / well, add 5% skim milk, and incubate for 2 h at room temperature. Remove the blocking solution and wash three times at 250 pl / well with 0.5%o PBST and pat dry for later use. Dilute the serum sample to the appropriate ratio using the blocking solution and add it to the blocked plate. The standard curves were added sequentially at 100 pl / well to the closed plate using TLb0045 and TLb0045-l, respectively, with 2-fold dilutions for a total of 20 concentration gradients (maximum concentration of 100 nM), at 100 pl / well, and incubated for 1 hour at room temperature. Wash the plate 3 times with 0.5%oPBST, add 10,000-fold dilution of HRP -labeled goat anti -human IgG antibody, 100 pl / well, and incubate for 60 min at room temperature. Wash the plate 5 times with 0.5%oPBST, add TMB at 100 pl / well, incubate at room temperature in the dark for 5 min, at 100 pl / well, add stop solution, stop the substrate chromogenic reaction, read the OD value at 450 nm with a microplate reader, analyze the data with a GraphPad, plot and calculate the half-life (T 1 / 2).

[0924] At the same time, the second method was used to increase antibody concentrations in mouse serum: recombinant human TLlA-his protein was diluted to 1 pg / ml in PBS at 100 pl / well, added to a microplate plate, and incubated overnight at 4 °C. Remove the coating solution, block at 200 pl / well, add 5% skim milk, and incubate at room temperature for 2 h. Remove the blocking solution and wash three times at 250 pl / well with 0.5%o PBST and pat dry for later use. The diluted serum sample is added to the closed microplate label. The standard Attorney Docket No.: 769273: SA9-943PC curves were added sequentially at 100 pl / well to the closed plate using TLb0045 and TLb0045- 1, respectively, with 2-fold dilutions for a total of 20 concentration gradients (maximum concentration of 100 nM), at 100 pl / well, and incubated for 1 hour at room temperature. Wash the plate 3 times with 0.5%oPBST, dilute a4p7-Biotin protein to 2 pg / ml and incubate at 100 pl / well for 60 min at room temperature; Add 5,000-fold diluted SA-HRP solution, incubate at room temperature for 60 min, wash the plate 5 times with 0.5%oPBST at 100 pl / well, add TMB, and incubate at room temperature at 100 pl / well in the dark. The stop solution was added, the substrate chromogenic reaction was stopped, the OD value at 450 nm was read with a microplate reader, the data was analyzed with a GraphPad, plotted and the half-life (Tl / 2) was calculated.

[0925] As shown in Figures 43 and 44, the concentrations of the bispecific antibodies TLb0045 and TLb0045-l in serum were not significantly different. The half-life calculated by fitting the two test results of TLb0045 is 5-6 days, and the half-life calculated by fitting the two test results of TLb0045-l is 9.7 days. The individual parameters of PK calculation are shown in Table 11.

[0926] Table 11. PK blood collection time point Attorney Docket No.: 769273: SA9-943PC

[0927] Table 12. PK test of TLb0045 and TLb0045-l in hFcRn mice

[0928] Example 23 Detection of the pharmacodynamic activity of the bispecific antibody in the DSS-induced hTLlA / h«4p7 mouse enteritis model This experiment used hTLlA / ha4p7 transgenic mice to study the in vivo pharmacodynamic activity of the bispecific antibody in the DSS-induced enteritis model. hTLlA / ha4p7 transgenic mice were purchased from Biocytogen (Beijing) Pharmaceutical Technology Co., Ltd. The humanized transgenic mice are C57BL / 6 strain mice, which are hybridized from B-hTLl A transgenic mice and B-ha4p7 transgenic mice. The transgenic mice can express human TL1A and human a4p7 proteins at the same time. The mouse enteritis model induced by dextran sulfate sodium salt (DSS) is the most widely used chemically induced mouse IBD model. Acute ulcerative enteritis or chronic colitis is induced by dissolving DSS in drinking water, destroying the intestinal epithelial cells of mice, and nonspecific immune cells releasing cytokines, which ultimately leads to the destruction of the integrity of the mucosal barrier. The animals show obvious weight loss, loose stools, blood in stools, and granulocyte infiltration, which are extremely similar to human ulcerative colitis in clinical symptoms and pathological characteristics. Therefore, the DSS-induced enteritis model can be used to evaluate the in vivo pharmacodynamic activity of the bispecific antibody. Attorney Docket No.: 769273: SA9-943PC

[0929] 32 female hTLlA / ha4p7 transgenic mice were randomly divided into six groups according to their body weight, including two mice in the G1 negative control group, G2 modeling group, G3 TLA0015-11 drug group, G4 Vedolizumab (analog) drug group, G5 TLA0015-11 + Vedolizumab drug combination group, and G6 TLb0045-l drug group, each with six mice. The detailed grouping of mice is shown in Table 13. The drug was injected intraperitoneally on days 0, 3, and 6, with a dose of 10 mg / kg. From day 0 to day 7, mice were given drinking water containing 3% DSS, which was replaced with normal drinking water from day 8 to day 9, and the experiment ended on day 9. During the experiment, the weight of the experimental animals was measured and recorded every two days, and the daily activities of the animals were observed; the fecal characteristics of the experimental animals were scored every day (0 = normal, 1 = moist / sticky, 2 = soft, 3 = liquid). After the experiment ended on day 9, all experimental animals were euthanized by asphyxiation with excessive carbon dioxide, and then the abdominal cavity was cut open, the colon was removed, and the colon length was recorded.

[0930] After the experiment, the results were statistically analyzed, and the results of the weight changes of mice are shown in Figures 45A and 45B. Compared with the G2 modeling group, the weight loss of mice in the four drug-administered groups was alleviated, and TLA0015-11 had the best efficacy. From the daily activity score results (Figure 46), the TLb0045-l drug- administered group had the lowest score and the best efficacy. At the end of the 9th day of the experiment, all mice were alive. The length and weight of the colorectum were measured after dissection. The results of the colorectal length of mice are shown in Figures 47A and 47B, and the results of the colorectal weight are shown in Figures 48A and 48B. The G2 modeling group can see that the colorectal length is significantly shortened and the weight is significantly increased. The TLA0015-11 and TLb0045-l administration groups can see the efficacy of the drug in the colorectal length and weight indicators of mice. Attorney Docket No.: 769273: SA9-943PC

[0931] Table 13 Mouse grouping and drug administration details (DSS-induced enteritis model)

[0932] Example 24 Detection of the pharmacodynamic activity of the bispecific antibody in the TNBS-induced hTLlA / h«4p7 mouse enteritis model

[0933] This experiment used hTLlA / ha4p7 transgenic mice to study the in vivo pharmacodynamic activity of the bispecific antibody in the TNBS-induced enteritis model. TNBS-treated mice can establish a preclinical model that simulates clinical Crohn's disease (CD). The immune response produced is Thl-mediated, characterized by the infiltration of CD4+T cells, neutrophils, and macrophages, forming a horizontally progressive inflammation, leading to colitis. Therefore, the TNBS-induced enteritis model can be used to evaluate the in vivo pharmacodynamic activity of the bispecific antibody.

[0934] Twenty-seven female hTLlA / ha4p7 transgenic mice were randomly divided into six groups according to their body weight, including two mice in the G1 negative control group, five mice in the G2 modeling group, five mice in the G3 TLA0015-11 drug group, five mice in the G4 Vedolizumab (analog) drug group, five mice in the G5 TLA0015-ll+Vedolizumab drug combination group, and five mice in the G6 TLb0045-l drug administration group. The detailed grouping of mice is shown in Table 14. Intraperitoneal administration began on Day 1, and the drug was administered once every three days for a total of three times. All animals were fasted on Day 0; on Day 1, 100 pl 60% ethanol was given rectal administration (i.r.) to the G1 negative control group, and 100 pl 2% TNBS was given rectal administration (i.r.) to the G2 to Attorney Docket No.: 769273: SA9-943PC

[0935] G6 groups, and all animals were given a feed to induce acute enteritis. During the experiment, the weight of the experimental animals was measured and recorded every day, and the daily activities of the animals were observed; the fecal characteristics of the experimental animals were scored every day (0 = normal, 1 = moist / sticky, 2 = soft, 3 = liquid). After the experiment ended on the 9th day, all experimental animals were euthanized by asphyxiation with excessive carbon dioxide, and then the abdominal cavity was cut open, the colon was removed, and the colon length was recorded.

[0936] After the experiment, the results were statistically analyzed, and the results of the weight change of mice are shown in Figures 49A and 49B. Compared with the G2 modeling group, the weight of mice in the TLA0015-11 drug group, the Vedolizumab (analog) drug group and the TLb0045-l drug group can be seen to have recovered. From the daily activity score results (Figure 50), the scores of the TLA0015-11 drug group, the Vedolizumab (analog) drug group and the TLb0045-l drug group began to decrease after the 4th day. The animal survival curve results are shown in Figure 51, where all mice in the modeling group and the combination group died on the 5th day. At the end of the 9th day, some other mice also died. After dissection, the length and weight of the colorectum were measured. The results of the colorectal length of mice are shown in Figures 52A and 52B, and the results of the colorectal weight are shown in Figures 53 A and 53B. The G2 modeling group can see that the colorectal length is significantly shortened and the weight is significantly increased. TLA0015-11, Vedolizumab (analog) and bispecific antibody TLb0045-l can all see the efficacy.

[0937] Table 14. Mouse grouping and drug administration details (TNBS-induced enteritis model) Attorney Docket No.: 769273: SA9-943PC

[0938] In summary, the sequences used in the present application are shown in the table below, wherein [] represents that the position can be arbitrarily replaced by the amino acids shown in []•

[0939] SEQUENCE LISTING Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC Attorney Docket No.: 769273: SA9-943PC

Claims

1. Attorney Docket No.: 769273: SA9-943PCClaims1. An isolated antigen-binding protein, comprising: a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain specifically binds to TNF-like protein A (TL1A), and wherein the second antigen binding domain specifically binds to a4p7.

2. The isolated antigen-binding protein of claim 1, comprising: a first antigen-binding domain and a second antigen-binding domain, wherein the first antigen-binding domain specifically binds to TNF-like protein A (TL1A), and the second antigen binding domain specifically binds to a4p7, wherein the first antigen-binding domain comprises a heavy chain variable region (VH), and the VH comprises an HCDR3 SEQ ID NO: 14.

3. The isolated antigen-binding protein according to claim 1 or claim 2, wherein the HCDR3 comprises an amino acid sequence of any one of SEQ ID NOs: 15-18.

4. The isolated antigen-binding protein according to any one of claims 1-3, wherein the VH of the first antigen-binding domain comprises an HCDR2 comprising an amino acid sequence of SEQ ID NO: 5.

5. The isolated antigen-binding protein of claim 4, wherein the HCDR2 comprises an amino acid sequence of any one of SEQ ID NOs: 6-13.

6. The isolated antigen-binding protein according to any one of claims 2-5, wherein the VH of the first antigen-binding domain comprises an HCDR1 comprising an amino acid sequence of SEQ ID NO: 1.

7. The isolated antigen-binding protein of claim 6, wherein the HCDR1 comprises an amino acid sequence of any one of SEQ ID NOs:2-4.

8. The isolated antigen-binding protein according to any one of claims 2-7, wherein(i) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 15;(ii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQAttorney Docket No.: 769273: SA9-943PCID NO: 2, the HCDR2 as set forth in SEQ ID NO: 7, and the HCDR3 as set forth in SEQ ID NO: 15;(iii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 8, and the HCDR3 as set forth in SEQ ID NO: 15;(iv) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 6 and the HCDR3 as set forth in SEQ ID NO: 15;(v) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 16;(vi) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 17;(vii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 18;(viii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 4, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 15;(ix) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 9, and the HCDR3 as set forth in SEQ ID NO: 15;(x) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 15;(xi) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 11, and the HCDR3 as set forth in SEQ ID NO: 15;(xii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ IDAttorney Docket No.: 769273: SA9-943PCNO: 2, the HCDR2 as set forth in SEQ ID NO: 12, and the HCDR3 as set forth in SEQ ID NO: 15;(xiii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 13, and the HCDR3 as set forth in SEQ ID NO: 15;(xiv) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 16;(xv) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 6, and the HCDR3 as set forth in SEQ ID NO: 17;(xvi) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 9, and the HCDR3 as set forth in SEQ ID NO: 15;(xvii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 15;(xviii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 3, the HCDR2 as set forth in SEQ ID NO: 11, and the HCDR3 as set forth in SEQ ID NO: 15;(xix) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 9, and the HCDR3 as set forth in SEQ ID NO: 16;(xx) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 16;(xxi) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO: 2, the HCDR2 as set forth in SEQ ID NO: 11, and the HCDR3 as set forth in SEQ ID NO: 16;(xxii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQAttorney Docket No.: 769273: SA9-943PCID NO: 2, the HCDR2 as set forth in SEQ ID NO: 12, and the HCDR3 as set forth in SEQ IDNO: 16 or(xxiii) the VH of the first antigen-binding domain comprises the HCDR1 as set forth in SEQ ID NO:2, the HCDR2 as set forth in SEQ ID NO: 10, and the HCDR3 as set forth in SEQ ID NO: 17.

9. The isolated antigen-binding protein according to any one of claims 2-8 wherein the first antigen-binding domain further comprises a light chain variable region (VL), wherein the VL of the first antigen-binding domain comprises at least one light chain complementary determining region (LCDR) comprising an amino acid sequence selected from SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44.

10. The isolated antigen-binding protein according to any one of claims 2-9, wherein the VH of the first antigen -binding domain comprises a sequence as set forth in any one of SEQ ID NOs: 62-99 or 379.

11. The isolated antigen-binding protein according to any one of claims 2-10, wherein the VL of the first antigen-binding domain comprises a sequence as set forth in in any one of SEQ ID NOs: 102-106 or 380.

12. The isolated antigen-binding protein according to any one of claims 1-11, wherein the second antigen binding domain comprises a heavy chain variable region (VH), and the VH of the second antigen binding domain comprises an HCDR1 as set forth in SEQ ID NO: 332, an HCDR2 as set forth in SEQ ID NO: 333, and an HCDR3 as set forth in SEQ ID NO: 334, optionally wherein the second antigen binding domain comprises a VH as set forth in SEQ ID NO: 335.

13. The isolated antigen-binding protein according to any one of claims 1-12, wherein the second antigen binding domain comprises a light chain variable region (VL), and the VL of the second antigen binding domain comprises an LCDR1 as set forth in SEQ ID NO: 336, an LCDR2 as set forth in SEQ ID NO: 337, and an LCDR3 as set forth in SEQ ID NO: 338, optionally wherein the second antigen binding domain comprises a VL as set forth in SEQ ID NO: 339.

14. The isolated antigen-binding protein of claim 1-13, wherein the antigen-binding protein comprises a Fab, Fab’, Fv fragment, F(ab)’2, scFv, di-scFv and / or dAb.Attorney Docket No.: 769273: SA9-943PC15. The isolated antigen-binding protein according to claims 1-14, wherein the antigenbinding protein comprises the scFv of the first antigen binding domain or the scFv of the second antigen binding domain.

16. The isolated antigen-binding protein according to claim 15, wherein the scFv of the first antigen-binding domain comprises a sequence as set forth in any one of SEQ ID NOs: 340- 345.

17. The isolated antigen-binding protein according to claim 16, wherein the scFv of the first antigen-binding domain comprises a sequence as set forth in SEQ ID NO: 342.

18. The isolated antigen-binding protein according to any one of claims 1-17, wherein the first antigen-binding domain is selected from the combination of light chain and heavy chain of one of the following: s) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:350; t) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:351; u) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:352; v) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:353; w) a light chain of SEQ ID NO:349 and a heavy chain of SEQ ID NO:354; x) a light chain of SEQ ID NO:349 and a heavy chain of SEQ ID NO:355; y) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:356; z) a light chain of SEQ ID NO:372 and a heavy chain of SEQ ID NO:357; aa) a light chain of SEQ ID NO:349 and a heavy chain of SEQ ID NO:358; bb) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:359; cc) a light chain of SEQ ID NO: 372 and a heavy chain of SEQ ID NO: 360; dd) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:361; ee) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:362; ff) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:363; gg) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:364; hh) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:365; ii) a light chain of SEQ ID NO:348 and a heavy chain of SEQ ID NO:366; jj) a light chain of SEQ ID NO:348 and a heavy chain as set forth in SEQ ID NO:378.

19. The isolated antigen-binding protein according to claim 1, whereinAttorney Docket No.: 769273: SA9-943PC the first antigen-binding domain comprises HCDR1 as set forth in SEQ ID NO: 2, HCDR2 as set forth in SEQ ID NO: 6, HCDR3 as set forth in SEQ ID NO: 15, LCDR1 as set forth in SEQ ID NO: 42, LCDR2 as set forth in SEQ ID NO: 43, and LCDR3 as set forth inSEQ ID NO: 44; and, the second antigen-binding domain comprises HCDR1 as set forth in SEQ ID NO: 332, HCDR2 as set forth in SEQ ID NO: 333, HCDR3 as set forth in SEQ ID NO: 334, LCDR1 as set forth in SEQ ID NO: 336, LCDR2 as set forth in SEQ ID NO: 337, and LCDR3 as set forth in SEQ ID NO: 338.

20. The isolated antigen-binding protein according to claim 19, wherein the first antigenbinding domain comprises a VH as set forth in SEQ ID NO: 379 and a VL as set forth in SEQ ID NO: 380; and the second antigen-binding domain comprises a VH as set forth in SEQ ID NO: 335 and a VL as set forth in SEQ ID NO: 339.

21. The isolated antigen-binding protein according to any one of claims 19-20, wherein(1) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 340; and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,(2) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 341; and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,(3) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 342; and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,(4) the first antigen-binding domain comprises an scFv as set forth in SEQ ID NO: 343; and the second antigen-binding domain comprises the VH as set forth in SEQ ID NO: 335 and the VL as set forth in SEQ ID NO: 339; or,(5) the first antigen binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106; and the second antigen binding domain comprises an scFv as set forth in 346; or,Attorney Docket No.: 769273: SA9-943PC(6) the first antigen binding domain comprises the VH as set forth in SEQ ID NO: 70 and the VL as set forth in SEQ ID NO: 106; and the second antigen binding domain comprises an scFv as set forth in SEQ ID NO:347.

22. The isolated antigen-binding protein according to any one of claims 19-20, wherein the isolated antigen-binding protein includes a light chain as shown in SEQ ID NO: 348 and a heavy chain as shown in SEQ ID NO: 363.

23. An isolated nucleic acid molecule encoding an isolated antigen-binding protein as described in any one of claims 1-22.

24. An expression vector comprising the isolated nucleic acid molecule according to claim 23.

25. A host cell including the isolated nucleic acid molecule according to claim 23 and / or the expression vector according to claim 24.

26. A composition comprising the isolated antigen-binding protein according to claims 1-22 and an optionally pharmaceutical acceptable carrier.

27. A preparation method for antigen-binding proteins, comprising: culturing the host cell of claim 25, and recovering the isolated antigen-binding proteins.

28. A method of treating a disease or conditions comprising: administering the isolated antigen-binding protein according to any one of claims 1-22, the isolated nucleic acid molecule according to claim 23, and / or the expression vector according to claim 24, the host cell according to claim 25, or the composition according to claim 26.

29. The isolated antigen-binding protein according to any one of claims 1-22 for use in the treatment of a disease or condition.

30. The isolated nucleic acid molecule according to claim 23, and / or the expression vector according to claim 24, the host cell according to claim 25, or the composition according to claim 26, for use in the treatment of a disease or condition.

31. Use of the isolated antigen-binding protein according to any one of claims 1-22, the isolated nucleic acid molecule according to claim 23, and / or the expression vector according to claim 24, the host cell according to claim 25, or the composition according to claim 26 for the treatment of disease or condition or in the preparation of a drug for the treatment of disease or condition.Attorney Docket No.: 769273: SA9-943PC32. The method according to claim 28, the isolated antigen-binding protein for the use of claim 29, the isolated nucleic acid molecule, expression vector, host cell or composition of claim 30 or the use according to claim 31, wherein the disease or the condition is TL1 A- mediated disease or condition.

33. The method, use or the isolated antigen-binding protein, isolated nucleic acid molecule, expression vector, host cell or composition for the use according to claim 32, wherein the TLlA-mediated disease or condition is inflammatory disease, optionally wherein the TL1 A- mediated inflammatory disease is selected from one of the following: Non-limiting examples of inflammatory disease include, allergy, ankylosing spondylitis, asthma, atopic dermatitis, autoimmune diseases or disorders, cancer, celiac disease, chronic obstructive pulmonary disease (COPD), chronic peptic ulcer, cystic fibrosis, diabetes (e.g., type 1 diabetes and type 2 diabetes), glomerulonephritis, gout, hepatitis (e.g., active hepatitis), an immune-mediated disease or disorder, inflammatory bowel disease (IBD) such as Crohn's disease and ulcerative colitis, myositis, osteoarthritis, pelvic inflammatory disease (PID), multiple sclerosis, neurodegenerative diseases of aging, periodontal disease (e.g., periodontitis), preperfusion injury transplant rejection, psoriasis, pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis), rheumatic disease, scleroderma, sinusitis, tuberculosis.

34. The method, use or the isolated nucleic acid molecule, expression vector, host cell or composition for the use according to claim 32, wherein the TL1 A-mediated disease or condition is autoimmune disease, optionally wherein the TL1 A-mediated autoimmune diseases are selected from one of the following: achalasia, Addison's disease, Adult Stil's disease, agammaglobulinemia, alopecia areata, amyloidosis, ankylosing spondylitis, anti- GBM / anti-TBM nephritis, antiphospholipid syndrome, autoimmune angioedema, autoimmune familial autonomic dysfunction, autoimmune encephalomyelitis, autoimmune hepatitis, autoimmune inner ear disease (AIED), autoimmune myocarditis, autoimmune oophoritis, autoimmune orchitis, autoimmune pancreatitis, autoimmune retinopathy, autoimmune urticaria, axonal and neuronal neuropathy (AMAN), Barlow disease (Balo disease), Behcet's disease, benign mucous membrane pemphigoid, bullous pemphigoid, Castleman's disease (Castleman disease (CD), celiac disease, Chagas disease, chronic inflammatory demyelinating polyneuropathy (CIDP), chronic recurrent multifocalAttorney Docket No.: 769273: SA9-943PC osteomyelitis (CRMO), Churg-Strauss syndrome (CSS) or eosinophilic granulomatous disease (EGPA), cicatricial pemphigoid, and Cogan's syndrome), cold agglutinin disease, congenital heart block, coxsackienew myocarditis, CREST syndrome, Crohn's disease, dermatitis herpetiformis, dermatomyositis, Devic's disease (neuromyelitis optica), discoid lupus, Dressier's syndrome, endometriosis, eosinophilic esophagitis (EoE), eosinophilic fasciitis, erythema nodosum, primary mixed cryoglobulinemia, Evans syndrome, fibromyalgia, fibrotic alveolitis, giant cell arteritis (temporal arteritis), giant cell myocarditis, glomerulonephritis, Goodpasture's syndrome, granulomatosis with polyangiitis, Graves' disease, Guillain-Barre syndrome syndrome), Hashimoto's thyroiditis, hemolytic anemia, Henoch- Schonl ein purpura (HSP), herpes gravilens or pemphigoid gestationum (PG), hidradenitis suppurativa (HS) (paradoxical acne), hypogammaglobulinemia, IgA nephropathy, IgG4-associated sclerotic disease, immune thrombocytopenic purpura (ITP), Inclusion body myositis (IBM), interstitial cystitis (IC), juvenile arthritis, juvenile diabetes mellitus (type 1 diabetes), juvenile myositis (JM), Kawasaki disease, Lambert-Eaton syndrome, leukocytoclastic vasculitis, lichen planus, lichen sclerosus, lignoconjunctivitis, linear IgA disease (LAD), lupus, chronic Lyme disease ( Lyme disease chronic), Meniere's disease, microscopic polyangiitis (MPA), mixed connective tissue disease (MCTD), Mooren's ulcer, Mucha-Habermann disease, multifocal motor neuropathy (MMN) or MMNCB, multiple sclerosis, myasthenia gravis, myositis, narcolepsy, neonatal lupus, neuromyelitis optica, neutropenia, ocular cicatricial pemphigoid, optic neuritis, recurrent rheumatism (PR), PandAS, paraneoplastic cerebellar degeneration (PCD), paroxysmal nocturnal hemoglobinuria (PNH), Parry Romberg syndrome, ciliary planitis (peripheral uveitis), Parsonage-Turner syndrome), pemphigus, peripheral neuropathy, perivenous encephalomyelitis, pernicious anemia (PA), POEMS syndrome, polyarteritis nodosa, polyglandular syndrome type I, II, type III, polymyalgia rheumatica, polymyositis, post- myocardial infarction syndrome, postpericardiotomy syndrome, primary biliary cirrhosis, primary sclerosing cholangitis, progesterone dermatitis, psoriasis, psoriatic arthritis, pure red blood cell aplasia (PRCA), pyoderma gangrenosum, Raynaud's phenomenon, reactive arthritis, reflex sympathetic dystrophy, relapsing polychondritis, restless legs syndrome (RES), retroperitoneal fibrosis, rheumatic fever, rheumatoid arthritis, sarcoidosis, SchmidtAttorney Docket No.: 769273: SA9-943PC syndrome, scleritis, scleroderma, Sjogren's syndrome, sperm and testicular autoimmunity, stiff-person syndrome (SPS), subacute bacterial endocarditis (SBE), Susac's syndrome syndrome), sympathetic ophthalmia (SO), Takayasu's arteritis, temporal arteritis / giant cell arteritis, thrombocytopenic purpura (TTP), Tolosa-Hunt syndrome (THS), transverse myelitis, type 1 diabetes mellitus, ulcerative colitis (UC), undifferentiated connective tissue disease (UCTD), uveitis, vasculitis, Vitiligo and Vogt-Koyanagi-Harada disease.

35. The method, use or the isolated nucleic acid molecule, expression vector, host cell or composition for the use according to claim 32, wherein the TL1 A-mediated disease or condition is cancer, optionally wherein TL1 A-mediated cancers are selected from one or more of the following: non-restrictive examples of cancer include adenoid cystic carcinoma, adrenal carcinoma, amyloidosis, anal cancer, ataxia-telangiectasia, atypical nevus syndrome, basal cell carcinoma, cholangiocarcinoma, Birt Hogg Dube syndrome, bladder cancer, bone cancer, brain tumor, breast cancer, male breast cancer, carcinoid tumor, cervical cancer, colorectal cancer, ductal cancer, Endometrial Cancer, Esophageal Cancer, Gastric Cancer, Gastrointestinal Stromal Tumor (GIST), HER2 -Positive Breast Cancer, Pancreatic Islet Cell Tumor, Juvenile Polyposis Syndrome, Kidney Cancer, Laryngeal Cancer, Leukemia-Acute Lymphoblastic Leukemia, Acute Lymphoblastic Leukemia (ALL), Acute Myeloid Leukemia AML, Adult Leukemia, Childhood Leukemia, 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's lymphoma (Lymphoma-Hodgkin's), non-Hodgkin's lymphoma (Lymphoma-Non-Hodgkin’s), 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 tumors, polycythemia vera, prostate cancer, renal cell carcinoma, retinoblastoma, salivary gland carcinoma, sarcoma, Sarcoma- Kaposi, skin cancer, small bowel cancer, stomach cancer, testicular cancer, thymoma, thyroid cancer, uterine (endometrial) cancer, vaginal cancer, and Wilms’ Tumor.