An antibody and uses thereof

Abstract

This invention relates to an antibody that primarily binds to plasma kallikrein, and the use of such antibodies in the preparation of medicaments for the prevention or treatment of diseases related to plasma kallikrein or plasma prekallikrein in subjects.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and more specifically to antibodies that primarily bind to plasma kallikrein and their uses. Background Technology

[0002] Plasma kallikrein (PK) belongs to the serine protease family and was first discovered in mammalian plasma. It is encoded by a single gene (KLKB1) located on chromosome 4q35 and is primarily synthesized in the liver. PK is a key enzyme in the kallikrein-kinin system (KKS), acting on high-molecular-weight kininogen (KH) to activate and release small-molecule bradykinin (BK). BK then participates in biological processes such as coagulation, fibrinolysis, complement activation, and inflammation by acting on bradykinin receptors. In recent years, with more in-depth research into the genetics, molecular biology, and pharmacology of plasma kallikrein, our understanding of its physiological and pathological roles has deepened. Studies have shown that plasma kallikrein-kininase is closely related to a variety of diseases, including inflammatory diseases, tumors, cardiovascular diseases, kidney diseases, central nervous system diseases, retinopathy, and diabetic retinopathy (Costa-Neto, CME et al. Participation of kallikrein-kinin system in different pathologies. Int. Immunopharmacol. 2008, 8, 135-142). For example, hereditary angioedema (HAE) is an autosomal dominant inherited condition mainly caused by a deficiency of C1-INH in the body, which weakens its inhibitory effect on plasma kallikrein-kininase, leading to uncontrolled activation of the KKS system, release of vasoactive substances, and increased vascular permeability, resulting in typical swelling (Farkas, H. Orphan drugs for the treatment of hereditary angioedema. Expert Opinion on Orphan Drugs, 2015, 1, 141-156). For example, in the vitreous humor of patients with diabetic macular edema, overactivation of the KKS system has been found, leading to increased retinal vascular permeability and retinal thickening. In recent years, numerous studies have been published indicating that plasma kallikrein inhibitors can reduce retinal vascular permeability and are used to treat diabetic retinopathy and diabetic macular edema (Feener, E. Plasma kallikrein and diabetic macular edema. Curr. Diab. Rep. 2010, 10, 270-275; Liu J. et al. Plasmakallikrein-kinin system and diabetic retinopathy. Biol. Chem. 2013, 394, 319-328).

[0003] Currently, some PK (kinin-releasing enzyme) protein or peptide drugs have entered clinical trials or been successfully marketed. For example, large-molecule plasma kallikrein inhibitors such as icaratide and lanadelumab are already on the market for the treatment of hereditary angioedema, with significant efficacy. Bicycle's THR-149, which uses a bicyclic peptide ligand that specifically binds to plasma kallikrein for the treatment of diabetic macular edema, is currently in Phase II clinical trials. Summary of the Invention

[0004] This invention provides an antibody or antigen-binding fragment that binds to plasma kallikrein. The antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region. The heavy chain variable region includes three complementarity-determining regions (CDRs): CDR1 contains the sequence X1X2X3X4X5, where X1 is D or N, X2 is T, I, or Y, X3 is Y or W, X4 is I or M, and X5 is F or H, or CDR1 is TSMMGVS; CDR2 contains the sequence Y1IY2PY3Y4Y5Y6Y7Y8Y9Y. 10 Y 11 Y 12 Y 13 Y 14 Y 15 Where Y1 is R or A, Y2 is D or Y, Y3 is E or G, Y4 is N or D, Y5 is D, G or S, Y6 is N or D, Y7 is T or I, Y8 is K, V or S, Y9 is Y or F, Y 10 For D or N, Y 11 For P or Q, Y 12 For K or R, Y 13 For I or F, Y 14 For Q or K, Y 15 For example, CDR2 is G or D, or CDR3 is HIYWDDDKRYNPSLKS; CDR3 contains the sequence Z1GZ2Z3Z4Z5Y, where Z1 is G or A, Z2 is G or S, Z3 is L or I, Z4 is F or P, and Z5 is A or S, or CDR3 is PHYYAFDGFGY or ERAYYRYDEDFDY;

[0005] The light chain variable region includes the following three complementarity-determining regions (CDRs): CDR1 contains the sequence L1ASL2DINL3YL4L5, where L1 is E or K, L2 is H or Q, L3 is N, K or S, L4 is I or L, and L5 is A or S, or CDR1 is SATSIINSNYFH; CDR2 contains the sequence M1M2M3M4LM5M6, where M1 is Y, F or R, M2 is T or A, M3 is S or N, M4 is T, S, R or N, M5 is Q, H, V or A, and M6 is S, Q, D or P; CDR3 contains the sequence LQYDN1LN2T, where N1 is N or D, N2 is F or W, or CDR3 is VQYDEFPLT or QQGSSLPRT.

[0006] Furthermore, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises the following three complementarity-determining regions (CDRs): CDR1 contains any one of the sequences SEQ ID NO: 1-5, CDR2 contains any one of the sequences SEQ ID NO: 6-11, and CDR3 contains any one of the sequences SEQ ID NO: 12-15; the light chain variable region comprises the following three complementarity-determining regions (CDRs): CDR1 contains any one of the sequences SEQ ID NO: 16-21, CDR2 contains any one of the sequences SEQ ID NO: 22-27, and CDR3 contains any one of the sequences SEQ ID NO: 28-31.

[0007] Furthermore, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region includes the following three complementarity-determining regions (CDRs): CDR1 contains the sequence SEQ ID NO:1; CDR2 contains the sequence SEQ ID NO:6; CDR3 contains the sequence SEQ ID NO:12; the light chain variable region includes the following three complementarity-determining regions (CDRs): CDR1 contains the sequence SEQ ID NO:16; CDR2 contains the sequence SEQ ID NO:22; CDR3 contains the sequence SEQ ID NO:28;

[0008] The variable region of the heavy chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:1; CDR2 contains the sequence SEQ ID NO:7; CDR3 contains the sequence SEQ ID NO:12; the variable region of the light chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:17; CDR2 contains the sequence SEQ ID NO:23; CDR3 contains the sequence SEQ ID NO:28;

[0009] The variable region of the heavy chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:2; CDR2 contains the sequence SEQ ID NO:8; CDR3 contains the sequence SEQ ID NO:13; the variable region of the light chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:18; CDR2 contains the sequence SEQ ID NO:24; CDR3 contains the sequence SEQ ID NO:29;

[0010] The variable region of the heavy chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:3; CDR2 contains the sequence SEQ ID NO:9; CDR3 contains the sequence SEQ ID NO:14; the variable region of the light chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:19; CDR2 contains the sequence SEQ ID NO:25; CDR3 contains the sequence SEQ ID NO:30;

[0011] The variable region of the heavy chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:4; CDR2 contains the sequence SEQ ID NO:10; CDR3 contains the sequence SEQ ID NO:12; the variable region of the light chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:20; CDR2 contains the sequence SEQ ID NO:26; CDR3 contains the sequence SEQ ID NO:29;

[0012] The variable region of the heavy chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:5; CDR2 contains the sequence SEQ ID NO:11; and CDR3 contains the sequence SEQ ID NO:15. The variable region of the light chain includes the following three complementary determinant regions (CDRs): CDR1 contains the sequence SEQ ID NO:21; CDR2 contains the sequence SEQ ID NO:27; and CDR3 contains the sequence SEQ ID NO:31.

[0013] The antibody or antigen-binding fragments provided by this invention include murine antibodies, chimeric antibodies, humanized antibodies, or fully humanized antibodies or their antigen-binding fragments.

[0014] Furthermore, the antibody or antigen-binding fragment provided by the present invention has a heavy chain variable region sequence selected from any one of SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 35, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 46, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 54 or SEQ ID NO: 56, and a light chain variable region sequence selected from any one of SEQ ID NO: 34, SEQ ID NO: 36-42, SEQ ID NO: 45, SEQ ID NO: 47-49, SEQ ID NO: 53, SEQ ID NO: 55 or SEQ ID NO: 57.

[0015] Furthermore, the antibody or antigen-binding fragment provided by the present invention has a heavy chain variable region sequence selected from SEQ ID NO: 35 and a light chain variable region sequence selected from any one of SEQ ID NO: 36-42; or a heavy chain variable region sequence selected from SEQ ID NO: 44 or 46 and a light chain variable region sequence of SEQ ID NO: 45; or a heavy chain variable region sequence selected from SEQ ID NO: 50 or SEQ ID NO: 32 and a light chain variable region sequence selected from any one of SEQ ID NO: 47-49; or a heavy chain variable region sequence of SEQ ID NO: 33 and a light chain variable region sequence of SEQ ID NO: 34; or a heavy chain variable region sequence of SEQ ID NO: 43 and a light chain variable region sequence of SEQ ID NO: 42; or a heavy chain variable region sequence of SEQ ID NO: 51 and a light chain variable region sequence of SEQ ID NO: 48; or a heavy chain variable region sequence of SEQ ID NO: 52 and a light chain variable region sequence of SEQ ID NO: 53; or a heavy chain variable region sequence of SEQ ID NO: 54 and a light chain variable region sequence of SEQ ID NO: 48. NO: 55; or the heavy chain variable region sequence is SEQ ID NO: 56, and the light chain variable region sequence is SEQ ID NO: 57.

[0016] The antigen-binding fragments described in this invention are preferably derived from Fab, Fab', Fv, scFv, or (Fab')2 fragments.

[0017] The present invention also provides nucleic acid molecules encoding the aforementioned antibody or antigen-binding fragments; further provides expression vectors for expressing the nucleic acids; and further provides host cells containing the vectors, the host cells comprising prokaryotic or eukaryotic cells, more preferably selected from yeast cells, mammalian cells (e.g., 293 cells or CHO cells).

[0018] The invention further provides light chain constant regions such as SEQ ID NO: 61 and its mutants; heavy chain constant regions such as Human IgG1, Human IgG2, Human IgG4, etc., wherein Human IgG1 is preferably SEQ ID NO: 58 and its mutants; Human IgG2 is preferably SEQ ID NO: 59 and its mutants; and Human IgG4 is preferably SEQ ID NO: 60 and its mutants.

[0019] The sequence correspondence between the antibody and antigen-binding fragments given in the examples of this invention is shown in Table 1:

[0020] Table 1. Sequences of some antibodies and antigen-binding fragments

[0021] KH01 SEQ ID NO:33 SEQ ID NO:34 KH12 SEQ ID NO:32 SEQ ID NO:47 KH02 SEQ ID NO:35 SEQ ID NO:36 KH13 SEQ ID NO:32 SEQ ID NO:48 KH03 SEQ ID NO:35 SEQ ID NO:37 KH14 SEQ ID NO:32 SEQ ID NO:49 KH04 SEQ ID NO:35 SEQ ID NO:38 KH15 SEQ ID NO:50 SEQ ID NO: ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​ ​

[0022] The light chain constant region of antibody KH01-18 involved in the examples provided in this invention is SEQ ID NO: 61; the heavy chain constant region of KH10 and KH12-14 is Human IgG1 (SEQ ID NO: 58); the heavy chain constant region of KH01-KH09, KH11, and KH15-18 is Human IgG4 (SEQ ID NO: 60); and KH19-21 is a murine antibody.

[0023] The present invention also provides a pharmaceutical composition comprising the above-described antibody or antigen-binding fragment and a pharmaceutically acceptable excipient; the pharmaceutical composition is preferably an intravitreal injection formulation, a subretinal injection formulation, a choroidal injection formulation, an intravenous injection formulation, an intratumoral injection formulation, or an intramuscular injection formulation.

[0024] The present invention also provides the use of the aforementioned somatic or antigen-binding fragment or pharmaceutical composition in the preparation of a medicament for the prevention or treatment of diseases associated with plasma kallikrein or plasma prokallikrein in subjects; wherein the diseases associated with plasma kallikrein or plasma prokallikrein are preferably edema, rheumatoid arthritis, gout, intestinal diseases, stomatitis, neuropathic pain, inflammatory pain, spinal stenosis-degenerative spinal disease, diabetes, arterial or venous thrombosis, aortic aneurysm, osteoarthritis, vasculitis, pulmonary embolism, stroke, sepsis, systemic lupus erythematosus nephritis and burns, and retinal diseases; wherein the edema is preferably hereditary angioedema, cerebral edema, or head trauma; wherein the retinal diseases are preferably diabetic macular edema, retinal vein occlusion, age-related macular degeneration, macular edema secondary to retinal vein occlusion, uveitis, endophthalmitis, or polypoid choroidal vascular disease.

[0025] The MHL (Lanadelumab) mentioned in this invention can be found in existing technologies (such as patents WO2011085103A, WO2014113701A, WO2017100679A, etc.) and can be constructed and prepared according to conventional technical means in the field.

[0026] definition

[0027] The term "binding" or "specific binding" as used in this invention refers to the binding of an antibody to an epitope of an antigen (such as human PPK) in an in vitro assay. The affinity of the binding is characterized by KD (binding rate) or Kd (dissociation constant). The antibody binding or specific binding of human plasma kallikrein (PK) described in this invention does not preclude its possible binding to other antigens or epitopes, such as plasma prokallikrein (PPK).

[0028] In this invention, "epitope" refers to the portion of an antigen that specifically binds to an antibody. An epitope typically consists of chemically active (e.g., polar, nonpolar, or hydrophobic) surface groups of a moiety (such as an amino acid or polysaccharide side chain) and may possess specific three-dimensional structural features and specific charge characteristics. An epitope can be composed of continuous and / or discontinuous amino acids forming conformational spatial units. For discontinuous epitopes, amino acids from different portions of the linear sequence of the antigen are close together in three-dimensional space due to the folding of the protein molecule.

[0029] The term "antibody" as used in this invention refers to a binding protein containing an antigen-binding site. The term "binding site" or "antigen-binding site" refers to the region of the antibody molecule to which the ligand actually binds. The term "antigen-binding site" comprises an antibody heavy chain variable domain (VH) and / or an antibody light chain variable domain (VL) or a VH / VL pair, and may be derived from an intact antibody or antibody fragment such as a single-chain Fv, VH domain and / or VL domain, Fab, or (Fab)2. In one embodiment of the invention, each antigen-binding site comprises an antibody heavy chain variable domain (VH) and / or an antibody light chain variable domain (VL), and preferably consists of a pair of antibody light chain variable domains (VL) and antibody heavy chain variable domains (VH).

[0030] The antibody of the present invention may comprise six complementarity-determining regions (CDRs), including three heavy chain variable domain CDRs (CDRH1, CDRH2, and CDRH3) and three light chain variable domain CDRs (CDRL1, CDRL2, and CDRL3). CDRs and framework regions (FRs) together constitute the heavy or light chain variable domains. Amino acid residues in the “complementarity-determining regions” or “CDRs” are responsible for antigen binding. The “framework” or “FR” regions are the variable domain regions outside the complementarity-determining regions. FRs are less variable than CDRs; there are four FR molecules: FR1, FR2, FR3, and FR4. The light and heavy chains of the antibody, from the N-terminus to the C-terminus, include the domains FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The CDRs on each chain are separated by the framework amino acids. During antibody recognition, the four FR molecules coil to bring the CDR molecules closer together. In particular, CDR3 of the heavy chain is the region most conducive to antigen binding.

[0031] In this invention, the term "Fab" refers to a polypeptide comprising an antibody heavy chain variable domain (VH), an antibody constant domain 1 (CH1), an antibody light chain variable domain (VL), and an antibody light chain constant domain (CL), wherein the heavy and light chain domains are stabilized by disulfide bonds. One Fab can form one antigen-binding site, and two "Fab"s can form "(Fab')2" through disulfide bonds, thus having two antigen-binding sites. The "F(ab')2" fragment can be further reduced to form two Fab' fragments.

[0032] In this invention, the term "scFv" refers to a polypeptide containing an antibody heavy chain variable domain (VH) and a light chain variable domain (VL), which are stabilized by a short peptide. The scFv is a key region for antigen binding. "Fv" refers to a polypeptide containing an antibody heavy chain variable domain (VH) and a light chain variable domain (VL), which are linked together by non-covalent bonds.

[0033] The binding molecules or antibodies of the present invention further comprise immunoglobulin constant regions of one or more immunoglobulin species. Immunoglobulin species include IgG, IgM, IgA, IgD, and IgE isotypes, and in the case of IgG and IgA, include their subtypes. In a preferred embodiment, the antibody of the present invention has a constant domain structure of an IgG-type antibody.

[0034] The term "constant region" in this invention refers to the sum of antibody domains excluding variable regions. Constant regions are not directly involved in antigen binding but exhibit different effector functions. Antibodies are classified into the following categories based on the amino acid sequence of the constant region of the heavy chain: IgA, IgD, IgE, IgG, and IgM, with IgG and IgA further subdivided into the following subtypes: IgG1, IgG2, IgG3, and IgG4, IgA1, and IgA2. The heavy chain constant regions corresponding to different antibody classes are referred to as α, δ, ε, γ, and μ, respectively. The light chain constant regions of all five antibody classes are referred to as κ (kappa) and λ (lambda). In this invention, constant regions derived from humans refer to the heavy chain constant regions and / or κ or λ light chain constant regions of human antibodies subclasses IgG1, IgG2, IgG3, or IgG4.

[0035] The antibody or antigen-binding fragments according to the present invention also include binding molecules or antibodies ("variants") having "conserved sequence modifications." This means nucleotide and amino acid sequence modifications that do not affect or alter the aforementioned characteristics. Nucleotides or amino acids can be modified using techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Non-essential amino acid residues in the antibody or antigen-binding fragment can preferably be substituted with another amino acid residue from the same side chain family. Therefore, a "variant" antibody or antigen-binding fragment refers to an amino acid sequence having one or more amino acid additions, deletions, and / or substitutions compared to the "parent" amino acid sequence. Therefore, the nucleotide or amino acid sequences of the antibody or antigen-binding fragments of the present invention all include sequences having "conserved sequence modifications" with at least 80%, 85%, 90%, 95%, 98%, 99%, or higher homology to the parent nucleotide or amino acid.

[0036] The term "chimeric antibody" in this invention refers to an antibody comprising an antibody variable domain from one source or species and at least a portion of a constant region derived from another source or species, typically prepared using recombinant DNA technology. For example, in one embodiment, a chimeric antibody comprising a mouse variable region and a human constant region.

[0037] In this invention, the term "humanized antibody" refers to an antibody whose constant region (i.e., the CH and CL regions) or which is entirely encoded by a human antibody gene. Humanized antibodies can significantly reduce the immune response induced by heterologous antibodies in the human body.

[0038] The term "host cell" in this invention refers to any cell line that can be modified to produce the antibodies described in this invention. In one embodiment, HEK293 cells and CHO cells are used as host cells. In this application, the terms "cell," "cell line," and "cell culture" are used interchangeably and include their progeny.

[0039] The term "pharmaceutically acceptable excipient" in this invention refers to any formulation or carrier medium capable of delivering an effective amount of the active molecule of this invention without interfering with the biological activity of the active molecule and without toxic side effects on the host or patient. This includes any solvents, dispersion media, surfactants, antioxidants, preservatives (e.g., antibacterial agents, antifungal agents), isotonics, absorption delay agents, salts, preservatives, pharmaceuticals, pharmaceutical stabilizers, binders, excipients, disintegrants, lubricants, sweeteners, flavoring agents, dyes, and combinations thereof known to those skilled in the art. Preferably, the excipient is suitable for administration via intraocular, intravenous, intramuscular, subcutaneous, parenteral, or intra-articular routes.

[0040] Instruction manual illustrations

[0041] ​ Images of fundus fluorescein angiography in rhesus monkeys in Example 6.

[0042] ​ Results of retinal microvascular leakage in Example 6

[0043] ​ Scoring criteria for fundus fluorescein angiography in rhesus monkeys (scores range from 0 to 10 based on the severity of retinal microvascular leakage; 0 for the saline group, 8 for the group given only CA-I, 9 for incomplete light transmission and blurred fundus, and 10 for complete lack of light transmission (animal has no light perception)).

[0044] ​ This is a fundus fluorescein angiography image of a rhesus monkey in Example 7. Detailed Implementation

[0045] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0046] The antibody heavy and light chain CDR transplantation, PCR introduction of site mutations, and screening of mutant libraries described in this invention are accomplished through conventional gene recombination technology and immunological techniques based on antigen-antibody interactions. Specific experimental methods and steps are as described in <<Molecular Cloning>> 3rd Edition (Joseph Sambrook, Science Press, August 1, 2002) and similar experimental manuals.

[0047] Example 1: Mouse Immunization and Hybridoma Screening: BALB / C female mice (6-8 weeks old, purchased from Dashuo Laboratory Animal Co., Ltd.) were immunized using either standard intraperitoneal or plantar immunization methods. Each mouse received 10 μg of Human kallikrein antigen (Enzyme Research, catalog number: 3792A) every 2-3 weeks. For the initial immunization, all mice were immunized with an equal volume of CFA (SIGMA, catalog number: SLBW7430) and Human kallikrein antigen (Enzyme Research, catalog number: 3792A), emulsified together. Booster immunizations before the sprint immunization (approximately the second to fourth immunizations) were performed using IFA (SIGMA, catalog number: F5506). Three days before fusion, mice underwent a sprint immunization. The Human kallikrein antigen used for the pre-fusion sprint immunization was prepared with PBS (20 μg / mouse). Mice were euthanized by twisting their necks and disinfected by immersion in 70% ethanol for 5 minutes. Spleen and lymph nodes were harvested, and the spleen and lymph nodes were appropriately ground. The obtained B lymphocytes or spleen B cells were then mixed with P3X63Ag8.653 myeloma cells (product number: CRL-1580 (purchased from ATCC) or FO myeloma cells (catalog number: TCM31, purchased from the Chinese Academy of Sciences Cell Bank) were mixed at a certain ratio and fused using an electrofusion instrument. When the hybridoma cells recovered to their optimal state, ELISA was performed. Based on the ELISA results, positive hybridoma cells were selected for expanded culture. A second test (combining ELISA and functional activity) was performed 2-3 days later, and functionally active hybridoma cell lines were expanded. Total RNA was extracted from hybridoma cells using the Trizol method, and 5 μg of total RNA was reverse transcribed to obtain cDNA. Then, using cDNA as a template, PCR amplification was performed using 5' RACE technology. The PCR product was ligated into the restriction enzyme vector pcDNA3.4, and the ligation product was transformed into competent Stellar Competent Cells by heat shock. Colony PCR identification was performed, and a small amount of plasmid was extracted from positive clones for sequencing to obtain the light and heavy chain variable region sequences of the hybridoma antibody gene.

[0048] Example 2: Construction of Chimeric Antibody: Primers were designed based on the variable region gene sequence of mouse hybridoma antibody and the constant region of human light and heavy chains. The variable region gene sequence of mouse hybridoma antibody was amplified by PCR to obtain the chimeric antibody Q fragment. The constant regions of human light and heavy chains (hIgG4 or hIgG1) were amplified by PCR to obtain the chimeric antibody H fragment. The Q and H fragments were simultaneously ligated into the restriction enzyme vector pcDNA3.4 using homologous recombination. The correctly sequenced plasmid was transfected into expiCHO or HEK293 cells for protein expression. Cells were cultured for approximately 7-10 days, and the cell supernatant was collected by centrifugation. The cell supernatant was purified using an AKTA protein purifier to obtain the chimeric antibody protein.

[0049] Example 3: Antibody Humanization: Since murine antibodies easily induce HAMA reactions in humans, humanization is necessary to reduce their immunogenicity in humans. This example employs traditional methods such as CDR transplantation, framework reorganization, and key amino acid reverse mutation design to humanize multiple candidate molecules. Specifically, the method involves first comparing the CDR regions of the murine antibody light and heavy chains with the FR region of a highly similar human germline gene, obtaining the humanized antibody variable regions. Then, homologous recombination is used to splice the constant regions of the light and heavy chains with the corresponding antibody variable regions and ligate them into expression vectors. Colony PCR screening and sequencing are then used to identify the correct antibody light and heavy chain expression plasmids. The correctly sequenced light and heavy chain plasmids are then transfected into expiCHO or HEK293 cells for expression. After 7-10 days, the cell supernatant is purified using an AKTA protein purifier to obtain the humanized antibody protein. The humanized antibody activity is then measured. The selected antibody molecules were compared with the parent antibody molecules in sequence. Based on factors such as amino acid position, polarity, molecular weight, and spatial conformation, the amino acid molecules that have a greater impact on activity were identified. These amino acids were then reverse-mutated to further increase the activity of the antibody molecules. Humanized antibody molecules were finally obtained through combination screening.

[0050] Example 4: Comparison with Humans and Affinity Test

[0051] The affinity of the candidate antibody for the antigen Human kallikrein (Enzymeresearch, catalog number: HPK1302) was detected using the Octet QKe refurbisher. The candidate antibody was loaded onto the sensor and subjected to binding and dissociation reactions with different concentrations of antigen under the following conditions: baseline time: 120 s, loading time: 300 s, association time: 600 s, and dissociation time: 2000 s. After the reaction was completed, the results were analyzed using Octet analysis software, and the results are shown in Table 2.

[0052] Table 2

[0053]

[0054] MHL (Lanadelumab) can be found in existing technologies (such as patents WO2011085103A, WO2014113701A, WO2017100679A, etc.) and can be constructed and prepared according to conventional technical means in this field.

[0055] Example 5: PK Activity Detection Method

[0056] PK specifically cleaves the H-Pro-Phe-Arg-AMC substrate from the C-terminus of arginine, releasing AMC (7-amino-4-methylcoumarin). AMC emits fluorescence (excitation at 360 nm, emission at 480 nm). Detecting changes in AMC fluorescence reflects the activity level of PK, thus achieving the purpose of detecting the biological activity of PK inhibitors. First, antibody samples are prepared. The antibody protein is diluted to 200 μg / ml as the starting concentration using detection buffer (20 mM Tris-HCl, pH 7.50, 50 mM NaCl, 1 mM EDTA, 0.1% PEG-8000, and 0.1% Triton X-100). A four-fold serial dilution is then performed, resulting in 11 concentration gradients. The diluted solutions are then set aside. Next, PK is diluted to a solution of 250 ng / ml using detection buffer, and the H-Pro-Phe-Arg-AMC substrate is diluted to 1000 μM using detection buffer. Pipette 50 μl of detection buffer (negative control) and each diluted protein solution of various concentration gradients into a 96-well opaque ELISA plate; add 40 μl of 250 ng / ml PK dilution buffer (final concentration 100 ng / ml) to the same 96-well plate; finally, add 10 μl of 1000 μM H-Pro-Phe-Arg-AMC substrate (final concentration 100 μM) to the same 96-well plate. Place the prepared 96-well plate into a multi-functional microplate reader (Spectra Max i3X) for detection. Detection parameters: excitation light 360 nm, emission light 480 nm, read every 60 s for a total of 10 min. After normalizing the enzyme reaction rate (slope), perform a four-parameter curve fitting with the concentration (nM) to calculate the IC50 (nM). The detection results are shown in Table 2.

[0057] Table 2 shows the activity results of some antibody pKa.

[0058] ​ 0.297 0.235 0.227 0.188 0.174 0.221 0.398 0.224 0.230 0.870 1.1 0.82 0.990

[0059] Example 6: Human Plasma Activity Detection Method

[0060] Human plasma contains proteins such as PPK and PK, but detection has shown that PK does not exhibit activity in either undiluted or diluted human plasma, possibly indicating an equilibrium state of enzyme inhibition. Adding Factor XIIa disrupts this equilibrium, producing PK protein with enzymatic activity. This principle is used to detect the activity of PK inhibitors. The first step is antibody sample preparation. The antibody protein is diluted to 200 μg / ml using detection buffer (20 mM Tris-HCl, pH 7.50, 150 mM NaCl, 1 mM EDTA, 0.1% PEG-8000, and 0.1% Triton X-100) as the starting concentration. A four-fold serial dilution is then performed, resulting in 11 concentration gradients. The diluted human plasma is then diluted 40-fold using detection buffer, Factor XIIa is diluted to 100 ng / ml, and the substrate peptide (H-Pro-Phe-Arg-AMC) is diluted to 1000 μM for later use. Add 50 μl of detection buffer and diluted antibody protein solutions of various concentrations to each well of a 96-well opaque ELISA plate; add 20 μl of 40-fold diluted human plasma to each well of the same 96-well plate and incubate at 37°C for 30 min; add 20 μl of 100 ng / ml Factor XIIa and incubate at 37°C for 45 min; finally, add 10 μl of 1000 μM substrate peptide (H-Pro-Phe-Arg-AMC) to each well of the 96-well opaque ELISA plate. Place the prepared 96-well plate into a multi-functional microplate reader (Spectra Max i3X) for detection. Detection parameters: excitation light 360 nm, emission light 480 nm, read every 60 s for a total of 10 min. Normalize the enzyme reaction rate (slope) and perform a four-parameter curve fitting with the concentration (nM) to calculate the IC50 (nM). The results are shown in Table 3.

[0061] Table 3 shows the results of antibody activity in human plasma.

[0062] ​ 0.753 0.52 0.949 0.987 0.921 0.918 0.863 0.578 ​ ​ ​ ​ ​ ​ ​ ​ ​ 0.764 0.1032 0.419 0.93 1.09 1.29 1.17

[0063] Example 7: Animal Efficacy Study

[0064] 1. Construction of primate disease models

[0065] In the vitreous body of patients with diabetic retinopathy, there is significant overexpression of carbonic anhydrase I (CA-I). Increased CA-I expression alters the vitreous microenvironment, thereby activating the KKS signaling pathway, leading to increased retinal vascular permeability, resulting in retinal vascular leakage and macular edema (DME). In rhesus monkeys (4 years old, 3.2-4.9 kg), intravitreal injection of carbonic anhydrase at doses of 0.4 mg–0.55 mg / eye was performed, and fluorescein fundus angiography (FFA) 30 minutes later revealed more severe microvascular leakage.

[0066] 2. The excipient formulations containing the test drug are shown in Table 4:

[0067] Table 4. Excipient formulations of the test drugs

[0068] ​ ​ ​ 8.8％ ​ 0.01％

[0069] 3. Administration method

[0070] The model was established by intravitreal injection of carbonic anhydrase (CA-I) 30 min after administration of the test drug (KH02) (500 μg / eye). Fundus fluorescein angiography (FFA) was performed 30 min after modeling to observe the inhibitory effect of the drug molecule on CA-I-induced retinal vascular leakage. Three rhesus monkeys were divided into a drug group, an MHL group, and a negative group (the negative group received a drug-free buffer solution), with one monkey (two eyes) in each group.

[0071] 4. Experimental Results

[0072] See experimental fundus fluorescein angiography images. ​ See fundus leakage condition ​ The drug of this invention can effectively inhibit leakage.

[0073] Example 8: Animal Efficacy Study

[0074] See Example 7 for details on animal model construction, etc.

[0075] Fundus fluorescein angiography scoring criteria: Scoring is based on the severity of retinal microvascular leakage, ranging from 0 to 10. The saline group receives 0 points for FFA images, the CA-I-only group receives 8 points, incomplete light transmission and blurred fundus image receive 9 points, and complete lack of light transmission (animal has no light perception) receives 10 points. See the detailed scoring criteria below. ​ .

[0076] The model was established by intravitreal injection of 100 μg / eye of the test drug (KH02) followed by intravitreal injection of carbonic anhydrase (CA-I) 30 min after drug administration. Fundus fluorescein angiography (FFA) was performed 30 min after modeling to observe the inhibitory effect of the drug molecule on CA-I-induced retinal vascular leakage. The model was re-established with CA-I 14 days later, with a second FFA analysis performed 30 min after modeling. Participants were divided into a drug group (2 rhesus monkeys, 4 eyes), a control group (saline group), and a CA-I group. Fundus fluorescein angiography images are shown below. ​ The drug of this invention can significantly inhibit leakage. SEQUENCE LISTING <110> Chengdu Kanghong Biotechnology Co., Ltd. <120> An antibody and its uses <130> KH20220106 <160> 61 <170> PatentIn version 3.3 <210> 1 <211> 5 <212> PRT <213> Artificial sequence <400> 1 Asp Thr Tyr Ile Phe 1 5 <210> 2 <211> 5 <212> PRT <213> Artificial sequence <400> 2 Asp Tyr Tyr Met Phe 1 5 <210> 3 <211> 7 <212> PRT <213> Artificial sequence <400> 3 Thr Ser Gly Met Gly Val Ser 1 5 <210> 4 <211> 5 <212> PRT <213> Artificial sequence <400> 4 Asp Ile Tyr Met Phe 1 5 <210> 5 <211> 5 <212> PRT <213> Artificial sequence <400> 5 Asn Tyr Trp Met His 1 5 <210> 6 <211> 17 <212> PRT <213> Artificial sequence <400> 6 Arg Ile Asp Pro Glu Asn Asp Asn Thr Lys Tyr Asp Pro Lys Ile Gln 1 5 10 15 Gly <210> 7 <211> 17 <212> PRT <213> Artificial sequence <400> 7 Arg Ile Asp Pro Glu Asn Asp Asn Thr Lys Phe Asp Pro Lys Ile Gln 1 5 10 15 Gly <210> 8 <211> 17 <212> PRT <213> Artificial sequence <400> 8 Arg Ile Asp Pro Glu Asn Gly Asn Thr Val Tyr Asp Pro Arg Phe Gln 1 5 10 15 Asp <210> 9 <211> 16 <212> PRT <213> Artificial sequence <400> 9 His Ile Tyr Trp Asp Asp Asp Lys Arg Tyr Asn Pro Ser Leu Lys Ser 1 5 10 15 <210> 10 <211> 17 <212> PRT <213> Artificial sequence <400> 10 Arg Ile Asp Pro Glu Asn Gly Asn Ile Lys Tyr Asp Pro Lys Phe Gln 1 5 10 15 Gly <210> 11 <211> 17 <212> PRT <213> Artificial sequence <400> 11 Ala Ile Tyr Pro Gly Asp Ser Asp Thr Ser Tyr Asn Gln Lys Phe Lys 1 5 10 15 Gly <210> 12 <211> 7 <212> PRT <213> Artificial sequence <400> 12 Gly Gly Gly Leu Phe Ala Tyr 1 5 <210> 13 <211> 7 <212> PRT <213> Artificial sequence <400> 13 Ala Gly Ser Ile Pro Ser Tyr 1 5 <210> 14 <211> 11 <212> PRT <213> Artificial sequence <400> 14 Pro His Tyr Tyr Ala Phe Asp Gly Phe Gly Tyr 1 5 10 <210> 15 <211> 13 <212> PRT <213> Artificial sequence <400> 15 Glu Arg Ala Tyr Tyr Arg Tyr Asp Glu Asp Phe Asp Tyr 1 5 10 <210> 16 <211> 11 <212> PRT <213> Artificial sequence <400> 16 Glu Ala Ser His Asp Ile Asn Asn Tyr Ile Ala 1 5 10 <210> 17 <211> 11 <212> PRT <213> Artificial sequence <400> 17 Lys Ala Ser His Asp Ile Asn Lys Tyr Ile Ala 1 5 10 <210> 18 <211> 11 <212> PRT <213> Artificial sequence <400> 18 Lys Ala Ser Gln Asp Ile Asn Lys Tyr Ile Ala 1 5 10 <210> 19 <211> 11 <212> PRT <213> Artificial sequence <400> 19 Lys Ala Ser Gln Asp Ile Asn Ser Tyr Leu Ser 1 5 10 <210> 20 <211> 11 <212> PRT <213> Artificial sequence <400> 20 Lys Ala Ser Gln Asp Ile Asn Lys Tyr Ile Ala 1 5 10 <210> twenty one <211> 12 <212> PRT <213> Artificial sequence <400> twenty one Ser Ala Thr Ser Ile Ile Asn Ser Asn Tyr Phe His 1 5 10 <210> twenty two <211> 7 <212> PRT <213> Artificial sequence <400> twenty two Tyr Thr Ser Thr Leu Gln Ser 1 5 <210> twenty three <211> 7 <212> PRT <213> Artificial sequence <400> twenty three Phe Thr Ser Thr Leu Gln Ser 1 5 <210> twenty four <211> 7 <212> PRT <213> Artificial sequence <400> twenty four Tyr Thr Ser Ser Leu His Gln 1 5 <210> 25 <211> 7 <212> PRT <213> Artificial sequence <400> 25 Arg Ala Asn Arg Leu Val Asp 1 5 <210> 26 <211> 7 <212> PRT <213> Artificial sequence <400> 26 Tyr Thr Ser Thr Leu Gln Pro 1 5 <210> 27 <211> 7 <212> PRT <213> Artificial sequence <400> 27 Arg Thr Ser Asn Leu Ala Ser 1 5 <210> 28 <211> 8 <212> PRT <213> Artificial sequence <400> 28 Leu Gln Tyr Asp Asn Leu Phe Thr 1 5 <210> 29 <211> 8 <212> PRT <213> Artificial sequence <400> 29 Leu Gln Tyr Asp Asp Leu Trp Thr 1 5 <210> 30 <211> 9 <212> PRT <213> Artificial sequence <400> 30 Val Gln Tyr Asp Glu Phe Pro Leu Thr 1 5 <210> 31 <211> 9 <212> PRT <213> Artificial sequence <400> 31 Gln Gln Gly Ser Ser Leu Pro Arg Thr 1 5 <210> 32 <211> 122 <212> PRT <213> Artificial sequence <400> 32 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asn Tyr 20 25 30 Trp Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Ala Ile Tyr Pro Gly Asp Ser Asp Thr Ser Tyr Asn Gln Lys Phe 50 55 60 Lys Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Val Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Glu Arg Ala Tyr Tyr Arg Tyr Asp Glu Asp Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 33 <211> 116 <212> PRT <213> Artificial Sequence <400> 33 Glu Val Gln Leu His Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala 1 5 10 15 Ser Val Arg Leu Ser Cys Thr Ala Ser Gly Phe Asn Ile Lys Asp Thr 20 25 30 Tyr Ile Phe Trp Val Lys Gln Arg Pro Glu Gln Gly Leu Glu Trp Ile 35 40 45 Gly Arg Ile Asp Pro Glu Asn Asp Asn Thr Lys Phe Asp Pro Lys Ile 50 55 60 Gln Gly Lys Ala Thr Leu Thr Ala Asp Thr Ser Ser Asn Thr Ala Tyr 65 70 75 80 Leu Gln Leu Gly Ser Leu Thr Ser Glu Asp Ser Ala Ile Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Gly Leu Phe Ala Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Ser Val Ser Ala<00​​​​​​​​​​​​​​​​​​​​​​​​​​​​​ 65 70 75 80 Glu Asp Ile Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 35 <211> 116 <212> PRT <213> Artificial Sequence <400> 35 Glu Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala 1 5 10 15 Ser Val Arg Leu Ser Cys Thr Ala Ser Gly Phe Asn Ile Lys Asp Thr 20 25 30 Tyr Ile Phe Trp Val Lys Gln Arg Pro Lys Gln Gly Leu Glu Trp Ile 35 40 45 Gly Arg Ile Asp Pro Glu Asn Asp Asn Thr Lys Tyr Asp Pro Lys Ile 50 55 60 Gln Gly Lys Ala Thr Leu Thr Ala Asp Thr Ser Ser Asn Thr Ala Tyr 65 70 75 80 Leu Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Ile Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Gly Leu Phe Ala Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Ser Val Ser Ala 115 <210> 36 <211> 106 <212> PRT <213> Synthetic sequence <400> 36 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Thr Ser Leu Gly 1 5 10 15 Gly Lys Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln His Lys Pro Gly Lys Gly Pro Arg Leu Leu Ile 35 40 45 His Tyr Thr Ser Thr Leu Gln Ser Gly Ile Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Lys Asp Tyr Ser Phe Ser Ile Ser Asn Leu Glu Pro 65 70 75 80 Glu Asp Ile Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 37 <211> 106 <212> PRT <213> Synthetic sequence <400> 37 Asp Ile Gln Met Thr Gln Ser Pro Ser Ala Met Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Asn Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 38 <211> 106 <212> PRT <213> Artificial Sequence <400> 38 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Asn Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 39 <211> 106 <212> PRT <213> Artificial Sequence <400> 39 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln Gln Lys Pro Gly Gln Pro Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Glu Phe Thr Leu Thr Ile Ser Asn Leu Gln Pro 65 70 75 80 Glu Asp Phe Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 40 <211> 106 <212> PRT <213> Artificial sequence <400> 40 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45<00�0688> Tyr Tyr Thr Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro<00%06%1>65 70 75 80 Glu Asp Ile Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 41 <211> 106 <212> PRT It should be noted that there are some incorrect tags in the original text, such as <00�0688> and <00%06%1>, which may cause problems in the translation and understanding. It is recommended to correct these tags before further processing.<213> Artificial Sequence <400> 41 Asp Ile Gln Met Thr Gln Ser Pro Ser Ala Met Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln Gln Lys Pro Gly Gln Pro Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Ile Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 42 <211> 106 <212> PRT <213> Artificial Sequence <400> 42 Asp Ile Gln Met Thr Gln Ser Pro Ser Ala Met Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Glu Ala Ser His Asp Ile Asn Asn Tyr 20 25 30 Ile Ala Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile 35 40 45 Tyr Tyr Thr Ser Thr Leu Gln Ser Gly Val Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln Pro 65 70 75 80 Glu Asp Ile Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asn Leu Phe Thr 85 90 95 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 43 <211> 116 <212> PRT <213> Artificial sequence <400> 43 Glu Val Gln Leu Val Glu Ser Gly Gly Ala Leu Val Lys Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Asn Ile Lys Asp Thr 20 25 30 Tyr Ile Phe Trp Val Arg Gln Met Pro Arg Lys Gly Leu Glu Trp Ile 35 40 45 Gly Arg Ile Asp Pro Glu Asn Asp Asn Thr Lys Tyr Asp Pro Lys Ile 50 55 60 Gln Gly Lys Ala Thr Ile Thr Ala Asp Thr Ser Thr Asp Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Gly Gly Gly Leu Phe Ala Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ser 115 <2 10> 44 <2 11> 122 <2 12> PRT <2 13> Artificial Sequence <4 00> 44 Glu Val Gln Leu Gln Gln Ser Gly Thr Val Leu Ala Arg Pro Gly Ala 1 5 10 15 Ser Val Lys Met Ser Cys Lys Ala Ser Gly Tyr Ser Phe Thr Asn Tyr 20 25 30 Trp Met His Trp Val Asn Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Ala Ile Tyr Pro Gly Asp Ser Asp Thr Ser Tyr Asn Gln Lys Phe 50 55 60 [[ID=​​​Met Glu Leu Ser Ser Leu Thr Ile Glu Asp Ser Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Arg Ala Tyr Tyr Arg Tyr Asp Glu Asp Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Thr Leu Thr Val Ser Ser 115 120 <210> 45 <211> 108 <212> PRT <213> Artificial Sequence <400> 45 Glu Ile Val Leu Thr Gln Ser Pro Thr Thr Met Ala Ala Ser Pro Gly 1 5 10 15 Glu Lys Ile Thr Ile Thr Cys Ser Ala Thr Ser Ile Ile Asn Ser Asn 20 25 30 Tyr Phe His Trp Tyr Gln Gln Lys Pro Gly Phe Ser Pro Lys Leu Leu 35 40 45 Ile Tyr Arg Thr Ser Asn Leu Ala Ser Gly Val Pro Ala Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Ser Tyr Ser Leu Thr Ile Gly Thr Met Glu 65 70 75 80 Ala Glu Asp Val Ala Thr Tyr Tyr Cys Gln Gln Gly Ser Ser Leu Pro 85 90 95 Arg Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 46 <211> 122 <212> PRT <213> Artificial Sequence <400> 46 Glu Val Gln Leu Gln Gln Ser Gly Thr Val Leu Ala Arg Pro Gly Ala 1 5 10 15 Ser Val Lys Met Ser Cys Lys Ala Ser Gly Tyr Ser Phe Thr Asn Tyr 20 25 30 Trp Met His Trp Val Asn Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Ala Ile Tyr Pro Gly Asp Ser Asp Thr Ser Tyr Asn Gln Lys Phe 50 55 60 Lys Gly Lys Ala Lys Leu Thr Ala Val Thr Ser Ala Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Thr Ile Glu Asp Ser Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Arg Ala Tyr Tyr Arg Tyr Asp Glu Asp Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Thr Leu Thr Val Ser Ser 115 120 <210> 47 <211> 108 <212> PRT <213> Artificial sequence <400> 47 Glu Ile Val Met Thr Gln Ser Pro Pro Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Val Thr Leu Ser Cys Ser Ala Thr Ser Ile Ile Asn Ser Asn 20 25 30 Tyr Phe His Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35 40 45 Ile Tyr Arg Thr Ser Asn Leu Ala Ser Gly Ile Pro Ala Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gln 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Gly Ser Ser Leu Pro 85 90 95 Arg Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 48 <211> 108 <212> PRT <213> Artificial sequence <400> 48 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15 Asp Arg Val Thr Ile Thr Cys Ser Ala Thr Ser Ile Ile Asn Ser Asn 20 25 30 Tyr Phe His Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro Lys Leu Leu 35 40 45 Ile Tyr Arg Thr Ser Asn Leu Ala Ser Gly Val Pro Ser Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Phe Thr Ile Ser Ser Leu Gln 65 70 75 80 Pro Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Gly Ser Ser Leu Pro 85 90 95 Arg Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 49 <211> 108 <212> PRT <213> Artificial Sequence <400> 49 Glu Ile Val Leu Thr Gln Ser Pro Ala Ile Met Ser Ala Ser Pro Gly 1 5 10 15 Glu Arg Val Thr Met Thr Cys Ser Ala Thr Ser Ile Ile Asn Ser Asn 20 25 30 Tyr Phe His Trp Tyr Gln Gln Lys Pro Gly Thr Ser Pro Arg Arg Trp 35 40 45 Ile Tyr Arg Thr Ser Asn Leu Ala Ser Gly Val Pro Ala Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Tyr Ser Leu Thr Ile Ser Ser Met Glu 65 70 75 80 Pro Glu Asp Ala Ala Thr Tyr Tyr Cys Gln Gln Gly Ser Ser Leu Pro 85 90 95 Arg Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 <210> 50 <211> 122 <212> PRT <213> Synthetic sequence <400> 50 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Thr Val Lys Ile Ser Cys Lys Val Ser Gly Tyr Thr Phe Thr Asn Tyr 20 25 30 Trp Met His Trp Val Gln Gln Ala Pro Gly Lys Gly Leu Glu Trp Met 35 40 45 Gly Ala Ile Tyr Pro Gly Asp Ser Asp Thr Ser Tyr Asn Gln Lys Phe 50 55 60 Lys Gly Arg Val Thr Ile Thr Ala Asp Thr Ser Thr Asp Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Thr Arg Glu Arg Ala Tyr Tyr Arg Tyr Asp Glu Asp Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 51 <211> 122 <212> PRT <213> artificial sequence <400> 51 Gln Val Gln Leu Leu Glu Ser Gly Ala Glu Leu Val Arg Pro Gly Ser 1 5 10 15 Ser Val Lys Ile Ser Cys Lys Ala Ser Gly Tyr Ala Phe Thr Asn Tyr 20 25 30 Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Ala Ile Tyr Pro Gly Asp Ser Asp Thr Ser Tyr Asn Gln Lys Phe 50 55 60 Lys Gly Lys Ala Thr Leu Thr Ala Asp Glu Ser Ser Ser Thr Ala Tyr 65 70 75 80 Met Gln Leu Ser Ser Leu Arg Ser Glu Asp Ser Ala Val Tyr Ser Cys 85 90 95 Ala Arg Glu Arg Ala Tyr Tyr Arg Tyr Asp Glu Asp Phe Asp Tyr Trp 100 105 110 Gly Gln Gly Thr Thr Val Thr Val Ser Ser 115 120 <210> 52 <211> 116 <212> PRT <213> artificial sequence <400> 52 Glu Val Gln Leu Gln Gln Ser Gly Ala Glu Val Val Arg Pro Gly Ala 1 5 10 15 Ser Val Lys Leu Ser Cys Lys Pro Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met Phe Trp Leu Arg Gln Arg Pro Asp Gln Gly Leu Glu Trp Ile 35 40 45 Gly Arg Ile Asp Pro Glu Asn Gly Asn Thr Val Tyr Asp Pro Arg Phe 50 55 60 Gln Asp Arg Ala Ser Ile Thr Ala Val Thr Ser Ser Asn Thr Ala Phe 65 70 75 80 Leu Gln Val Asn Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Ala Gly Ser Ile Pro Ser Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ala 115 <210> 53 <211> 106 <212> PRT <213> artificial sequence <400> 53 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Leu Gly 1 5 10 15 Asp Lys Ile Thr Ile Thr Cys Lys Ala Ser Gln Asp Ile Asn Lys Tyr 20 25 30 Ile Ala Trp Tyr Gln His Lys Pro Gly Lys Gly Pro Arg Leu Leu Ile 35 40 45 His Tyr Thr Ser Ser Leu His Gln Asp Ile Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Arg Asp Tyr Ser Phe Ser Ile Ser Asn Leu Glu Pro 65 70 75 80 Glu Asp Phe Ala Ile Tyr Tyr Cys Leu Gln Tyr Asp Asp Leu Trp Thr 85 90 95 Phe Gly Gly Gly Thr Lys Val Glu Ile Lys 100 105 <210> 54 <211> 121 <212> PRT <213> Artificial Sequence <400> 54 Gln Ile Thr Leu Lys Glu Ser Gly Pro Gly Ile Leu Gln Pro Ser Gln 1 5 10 15 Thr Leu Ser Leu Thr Cys Ser Phe Ser Gly Phe Ser Leu Ser Thr Ser 20 25 30 Gly Met Gly Val Ser Trp Ile Arg Gln Ser Ser Gly Lys Gly Leu Glu 35 40 45 Trp Leu Ala His Ile Tyr Trp Asp Asp Asp Lys Arg Tyr Asn Pro Ser 50 55 60 Leu Lys Ser Arg Leu Thr Ile Ser Lys Asp Thr Ser Arg Asp Gln Val 65 70 75 80 Phe Leu Lys Ile Thr Ser Val Asp Thr Ala Asp Thr Ala Thr Tyr Tyr 85 90 95 Cys Val Arg Pro His Tyr Tyr Ala Phe Asp Gly Phe Gly Tyr Trp Gly 100 105 110 Gln Gly Thr Leu Val Thr Val Ser Ala 115 120 <210> 55 <211> 107 <212> PRT <213> Artificial Sequence <400> 55 Asp Ile Lys Met Thr Gln Ser Pro Ser Ser Met Ser Ala Ser Pro Gly 1 5 10 15 Glu Arg Val Thr Ile Thr Cys Lys Ala Ser Gln Asp Ile Asn Ser Tyr 20 25 30 Leu Ser Trp Phe Gln Gln Lys Ser Gly Lys Phe Pro Lys Thr Leu Ile ​​Tyr Arg Ala Asn Arg Leu Val Asp Gly Val Ala Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Gln Asp Tyr Ser Leu Thr Ile Ser Ser Leu Asp Tyr 65 70 75 80 Glu Asp Met Gly Ile Tyr Tyr Cys Val Gln Tyr Asp Glu Phe Pro Leu 85 90 95 Thr Phe Gly Ala Gly Thr Lys Leu Glu Leu Lys 100 105 <210> 56 <211> 116 <212> PRT <213> Artificial Sequence <400> 56 Glu Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Leu Ser Cys Thr Gly Ser Gly Phe Asn Ile Lys Asp Ile [[ID=X]]20 25 30 Tyr Met Phe Trp Val Lys Gln Arg Pro Glu Gln Gly Leu Glu Trp Ile 35 40 45 Gly Arg Ile Asp Pro Glu Asn Gly Asn Ile Lys Tyr Asp Pro Lys Phe 50 55 60 Gln Gly Lys Ala Thr Ile Thr Ala Glu Thr Ser Ser Asn Thr Val Tyr 65 70 75 80 It seems there is a small error in your original text where the "X" in line 32 should probably be "32" for proper numbering consistency. I've translated it as is but noted this for your reference.Leu Gln Leu Asn Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ser Arg Gly Gly Gly Leu Phe Ala Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Ser Ala<用 115 <210> 57 <211> 106 <212> PRT <213> Artificial Sequence <400> 57 Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Leu Gly 1 5 10 15 Gly Lys Val Thr Ile Thr Cys Lys Ala Ser Gln Asp Ile Asn Lys Tyr 20 25 30 Ile Ala Trp Tyr Gln His Lys Pro Gly Lys Gly Pro Arg Leu Leu Ile 35 40 45 His Tyr Thr Ser Thr Leu Gln Pro Gly Ile Pro Ser Arg Phe Ser Gly 50 55 60 Ser Gly Ser Gly Ser Asp Tyr Ser Phe Ser Ile Ser Thr Leu Glu Pro 65 70 75 80 Glu Asp Ile Ala Thr Tyr Tyr Cys Leu Gln Tyr Asp Asp Leu Trp Thr 85 90 95 Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 用05 It should be noted that there seems to be an error in the original text where "<用 " and "<用000103", and the translated text has been adjusted as accurately as possible based on the available content. <210> 58 <211> 329 <212> PRT <213> IG1 <400> 58 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 1 5 10 15 Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr 65 70 75 80 Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110 Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125 Lys Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys 130 135 140 Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160 Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175 Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190 His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205 Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly 210 215 220 Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu 225 230 235 240 Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255 Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn 260 265 270 Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285 Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn 290 295 300 Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320 Gln Lys Ser Leu Ser Leu Ser Pro Gly 325 <210> 59 <211> 326 <212> PRT <213> IG2 <400> 59 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser Arg 1 5 10 15 Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Asn Phe Gly Thr Gln Thr 65 70 75 80 Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Thr Val Glu Arg Lys Cys Cys Val Glu Cys Pro Pro Cys Pro Ala Pro 100 105 110 Pro Val Ala Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp 115 120 125 Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp 130 135 140 Val Ser His Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp Gly 145 150 155 160 Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe Asn 165 170 175 Ser Thr Phe Arg Val Val Ser Val Leu Thr Val Val His Gln Asp Trp 180 185 190 Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu Pro 195 200 205 Ala Pro Ile Glu Lys Thr Ile Ser Lys Thr Lys Gly Gln Pro Arg Glu 210 215 220 Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn 225 230 235 240 Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile 245 250 255 Ser Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr 260 265 270 Thr Pro Pro Met Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys 275 280 285 Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys 290 295 300 Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu 305 310 315 320 Ser Leu Ser Pro Gly Lys 325 <210> 60 <211> 327 <212> PRT <213> IG4 <400> 60 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Cys Ser Arg 1 5 10 15 Ser Thr Ser Glu Ser Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Lys Thr 65 70 75 80 Tyr Thr Cys Asn Val Asp His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Arg Val Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala Pro 100 105 110 Glu Phe Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys 115 120 125 Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val 130 135 140 Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp 145 150 155 160 Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe 165 170 175 Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp 180 185 190 Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu 195 200 205 Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg 210 215 220 Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr Lys 225 230 235 240 Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 245 250 255 Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys 260 265 270 Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser[[ID=W]] 275 280 285 Arg Leu Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe Ser 290 295 300 Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser 305 310 315 320 Leu Ser Leu Ser Leu Gly Lys 325 <210> 61 <211> 107 <212> PRT <213> Synthetic Sequence <400> 61 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 1 5 10 15 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe [[ID=W]] Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 35 40 45 ​Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 50 55 60 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 65 70 75 80 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 85 90 95 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 100 105

Claims

1. An antibody or antigen-binding fragment that binds human plasma kallikrein, characterized in that, The antibody or antigen-binding fragment comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region includes the following three complementarity-determining regions (CDRs): CDR1 sequence is SEQ ID NO:1, CDR2 sequence is SEQ ID NO:6, and CDR3 sequence is SEQ ID NO:12; the light chain variable region includes the following three complementarity-determining regions (CDRs): CDR1 sequence is SEQ ID NO:16, CDR2 sequence is SEQ ID NO:22, and CDR3 sequence is SEQ ID NO:

28.

2. The antibody or antigen-binding fragment of claim 1, wherein, The antibody or antigen-binding fragment is a murine antibody, a chimeric antibody, or a humanized antibody or its antigen-binding fragment.

3. The antibody or antigen-binding fragment of claim 1, wherein The heavy chain variable region sequence is SEQ ID NO:35, and the light chain variable region sequence is selected from any one of SEQ ID NO:36-42; or the heavy chain variable region sequence is SEQ ID NO:43, and the light chain variable region sequence is SEQ ID NO:

42.

4. The antibody or antigen binding fragment of any one of claims 1-3, wherein, The antigen-binding fragment is selected from Fab, Fab', Fv, scFv, or (Fab')2 fragments.

5. An isolated nucleic acid that encodes an antibody or antigen-binding fragment as described in any one of claims 1-4.

6. A vector comprising the nucleic acid of claim 5, wherein the vector is an expression vector.

7. A host cell comprising the vector of claim 6.

8. The host cell of claim 7, wherein, The host cell is a prokaryotic cell or a eukaryotic cell.

9. The host cell of claim 7, wherein, The host cell is selected from yeast cells or mammalian cells.

10. A pharmaceutical composition, characterized by It comprises an antibody or antigen-binding fragment as described in any one of claims 1-4, and a pharmaceutically acceptable excipient.

11. The pharmaceutical composition of claim 10, wherein, The pharmaceutical composition is an intravitreal injection preparation, a subretinal injection preparation, a choroidal injection preparation, an intravenous injection preparation, an intratumoral injection preparation, or an intramuscular injection preparation.

12. Use of the antibody or antigen-binding fragment of any one of claims 1-4 in the preparation of a medicament for treating macular edema.

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