Anti-histone h3 antibody or antigen-binding fragment thereof, and use thereof

Abstract

The present invention relates to the field of biomedicine, especially to an anti-histone H3 antibody or an antigen-binding fragment thereof, and the use thereof. The provided anti-histone H3 antibody or the antigen-binding fragment thereof contains a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and the light chain variable region comprises LCDR1, LCDR2 and LCDR3. A high-purity and high-affinity anti-histone H3 antibody is obtained by means of the screening of a humanized recombinant antibody library, and preparation, identification and modification, which antibody can significantly ameliorate related diseases mediated by histone H3.

Description

An anti-histone H3 antibody or its antigen-binding fragment and its application Technical Field

[0001] This invention relates to the field of biomedicine, specifically to an anti-histone H3 antibody or its antigen-binding fragment and its applications. Background Technology

[0002] Histones are strong cationic proteins found in the nucleus of eukaryotic cells, including H2A, H2B, H3, and H4, and are highly conserved across species. Within the nucleus, histones are major protein components of chromatin, acting as the threads that wind up DNA and playing a crucial role in gene regulation. Histones can be released into the extracellular space in three forms: free release, as nucleosomes wrapped around DNA, or as part of neutrophil extracellular traps (NETs). All three forms can be detected in serum following severe cell death (such as sepsis, trauma, ischemia / reperfusion injury, and autoimmune diseases). Once in the extracellular space, histones act as damage-associated molecular pattern (DAMP) proteins, activating the immune system and causing further cytotoxicity. Blocking histone release, neutralizing circulating histones, or blocking histone signal transduction may effectively reduce mortality and disability rates in animal models of acute organ injury or inflammatory diseases.

[0003] Studies have found that extracellular histones are potential mediators of death in diseases such as sepsis, especially circulating histone H3 levels, which are closely associated with multi-organ failure and death in sepsis patients. Besides sepsis, the accumulation of extracellular histones (NETs) due to excessive production or reduced clearance can also cause harm to the human body. For example, NETs can promote vasculitis and thrombosis, are associated with aseptic inflammatory diseases such as rheumatoid arthritis and systemic lupus erythematosus, and participate in the development of hematologic malignancies and non-malignant hematologic diseases. Therefore, detecting, identifying, and blocking the biological functions of extracellular histone H3 is particularly important for medical testing. Summary of the Invention

[0004] In view of the shortcomings of the prior art described above, the purpose of this invention is to provide an anti-histone H3 antibody or its antigen-binding fragment and its application, in order to solve the problems in the prior art.

[0005] To achieve the above and other related objectives, the first aspect of the present invention provides an antibody against histone H3 or an antigen-binding fragment thereof, the antibody or antigen-binding fragment thereof comprising a heavy chain variable region and a light chain variable region, the heavy chain variable region comprising HCDR1, HCDR2 and HCDR3, and the light chain variable region comprising LCDR1, LCDR2 and LCDR3.

[0006] The sequence of HCDR1 is shown in SEQ ID NO:1, SEQ ID NO:17, SEQ ID NO:32, SEQ ID NO:48, SEQ ID NO:71 or SEQ ID NO:83;

[0007] The sequences of HCDR2 are shown in SEQ ID NO:2, SEQ ID NO:18, SEQ ID NO:33, SEQ ID NO:49 or SEQ ID NO:72;

[0008] The sequences of HCDR3 are shown in SEQ ID NO:3, SEQ ID NO:19, SEQ ID NO:34, SEQ ID NO:50, SEQ ID NO:77, SEQ ID NO:85 or SEQ ID NO:89;

[0009] The sequence of LCDR1 is shown in SEQ ID NO:4, SEQ ID NO:20, SEQ ID NO:35 or SEQ ID NO:51;

[0010] The sequence of LCDR2 is shown in SEQ ID NO:5, SEQ ID NO:21, SEQ ID NO:36, SEQ ID NO:52, SEQ ID NO:73 or SEQ ID NO:79;

[0011] The sequences of LCDR3 are shown in SEQ ID NO:6, SEQ ID NO:22, SEQ ID NO:37, SEQ ID NO:53, SEQ ID NO:86, SEQ ID NO:91, SEQ ID NO:93, SEQ ID NO:95 or SEQ ID NO:97.

[0012] A second aspect of the present invention provides an isolated polynucleotide encoding the aforementioned anti-histone H3 antibody or a binding fragment thereof.

[0013] A third aspect of the present invention provides a nucleic acid construct containing the isolated polynucleotide.

[0014] A fourth aspect of the present invention provides an isolated engineered cell containing the aforementioned nucleic acid construct or the aforementioned polynucleotide.

[0015] The fifth aspect of the present invention provides the use of the aforementioned anti-histone H3 antibody or its antigen-binding fragment, the aforementioned polynucleotide or the aforementioned nucleic acid construct in the preparation of disease diagnostic, compound or therapeutic products.

[0016] As described above, the anti-histone H3 antibody or its antigen-binding fragment of the present invention and its application have the following beneficial effects:

[0017] This invention obtains high-purity and high-affinity anti-histone H3 antibodies through screening, preparation, and identification of humanized recombinant antibody libraries, which can significantly improve histone H3-mediated diseases. Attached Figure Description

[0018] Figure 1 shows the survival results analysis in the LPS-induced sepsis model;

[0019] Figure 2 shows the results of routine blood tests in mice in the LPS-induced sepsis model;

[0020] Figure 3 shows the results of liver and kidney function, myocardial and blood biochemical indicators in mice in the LPS-induced sepsis model, compared with the control (negative). *** P < 0.001, ** P < 0.01, * P < 0.05, ns: P < 0.001;

[0021] Figure 4 shows the blood biochemical results of mice in a LPS-induced sepsis model using mutant antibodies. *** P < 0.001, ** P < 0.01, * P < 0.05, ns: P < 0.001;

[0022] Figure 5 shows the blood biochemical results of the dose-dependent effect of the mutant antibody on the LPS-induced sepsis model in mice. *** P < 0.001, ** P < 0.01, * P < 0.05, ns: P < 0.001;

[0023] Figure 6 shows the efficacy results of the LPS-induced sepsis model of the antibody B029-139-B029-046 of this application;

[0024] Figure 7 shows the effect of antibody B029-122-B029-046 of this application on a rat model of acute gouty arthritis induced by sodium urate. In the figure, A represents the effect of a single tail vein injection of the test substance on the plantar pain threshold of rats with acute gouty arthritis induced by sodium urate. B represents the effect of a single tail vein injection of the test substance on paw volume in rats with sodium urate-induced acute gouty arthritis. C represents the effect of a single tail vein injection of the test substance on the rate of paw edema in rats with sodium urate-induced acute gouty arthritis. D represents the effect of a single tail vein injection of the test substance on the inhibition rate of paw swelling in rats with sodium urate-induced acute gouty arthritis.

[0025] Figure 8 shows the results of the cerebral ischemia-reperfusion experiment in mice with antibody B029-122-B029-046. A shows the effect of a single tail vein injection on the balance beam test in stroke mice (n=8, Mean±SEM); B shows the effect of a single tail vein injection on the cerebral blood flow difference in stroke mice (n=8, Mean±SEM); C shows the effect of a single tail vein injection on the infarct area ratio in stroke mice (n=8, Mean±SEM).

[0026] Figure 9 shows a TTC staining image of a brain slice in a mouse brain ischemia-reperfusion experiment using the B029-122-B029-046 antibody.

[0027] Figure 10 shows the effects of antibody B029-122-B029-046 on ulcerative colitis in mice. A is a graph showing the changes in body weight in the model group and the antibody B029-122-B029-046 group; B is a graph showing the DAI score of feces; C is a graph showing colon length and colon weight; and D is a graph showing the measurement of colon length.

[0028] Figure 11 shows the experimental results of the effect of antibody B029-139-B029-046 on acute pancreatitis in mice. A is the body weight change of the model group and the antibody group in the L-arginine-induced model; B is the serum amylase and lipase detection results in the L-arginine-induced model; C is the body weight change of the model group and the antibody group in the sodium taurocholate-induced model; D is the serum amylase and lipase detection results in the sodium taurocholate-induced model; E is the 24-hour animal mortality rate statistics; F is the 24-hour pathological score statistics.

[0029] Figure 12 shows the experimental results of the effect of antibody B029-139-B029-046 on rheumatoid arthritis in rats. A is a statistical result of rat paw volume, relative paw volume, and clinical arthritis score; B is a result of joint pathology score.

[0030] Figure 13 shows the experimental results of the effects of antibody B029-139-B029-046 on inflammatory cells and lung injury in mice with COPD.

[0031] Figure 14 shows the experimental results of the effect of antibody B029-139-B029-046 on acute kidney injury in mice.

[0032] Figure 15 shows the experimental results of the effect of antibody B029-139-B029-046 on systemic lupus erythematosus in mice; A is the statistical results of mouse body weight, spleen index, and kidney index; B is the detection results of UTP and mALB / Crea; C is the detection results of serum dsDNA.

[0033] Figure 16 shows the experimental results of the effect of antibody B029-139-B029-046 on pulmonary fibrosis in mice; A is the statistical results of mouse body weight and survival rate; B is the results of the detection of white blood cells, lymphocytes, neutrophils and eosinophils in BALF.

[0034] Figure 17 shows the experimental results of the effect of antibody B029-139-B029-046 on atopic dermatitis in mice.

[0035] Figure 18 shows the experimental results of the effect of antibody B029-139-B029-046 on asthma in mice; A is the result of inflammatory cell detection in BALF solution; B is the result of pathological analysis of lung tissue.

[0036] Figure 19 shows the experimental results of the effect of antibody B029-139-B029-046 on psoriasis in mice.

[0037] Figure 20 shows the experimental results of the effect of antibody B029-122-B029-046 on CLP-induced sepsis in mice.

[0038] Figure 21 shows the experimental results of the effect of antibody B029-139-B029-046 on acute respiratory distress syndrome in mice; A is the statistical results of mouse body weight change; B is the results of inflammatory cell detection in BALF solution; C is the results of endpoint pulmonary function test; D is the results of comprehensive pathological score of mouse lung tissue. Detailed Implementation

[0039] In this article, the term "antibody" is used in its broadest sense, specifically covering intact monoclonal antibodies, polyclonal antibodies, multispecific antibodies (such as bispecific antibodies) formed from at least two intact antibodies, and antibody fragments, as long as they exhibit the desired biological activity.

[0040] An antibody that "binds" to a target antigen, such as histone H3, is an antibody that can bind to the antigen with sufficient affinity so that it can be used as a therapeutic agent targeting cells expressing that antigen. If the antibody is one that binds to histone H3, it will typically bind to histone H3 preferentially over other antibodies.

[0041] The term "monoclonal antibody," as used herein, refers to an antibody obtained from a substantially homogeneous group of antibodies, meaning that the individual antibodies constituting the group are identical and, apart from possible naturally occurring mutations, are typically present in very small amounts. Monoclonal antibodies are highly specific, targeting a single antigenic site. Furthermore, unlike polyclonal antibody products which contain different antibodies targeting different determinants (epitopes), each monoclonal antibody targets a single determinant on the antigen. In addition to their specificity, the advantage of monoclonal antibodies lies in their ability to be synthesized without contamination from other antibodies. The modifier "monoclonal" indicates that the antibody is obtained from a substantially homogeneous group of antibodies and is not interpreted as requiring any particular method for antibody production.

[0042] Monoclonal antibodies in this article explicitly include “chimeric” antibodies, wherein a portion of the heavy and / or light chain is identical or homologous to the corresponding sequence in an antibody derived from a particular species or belonging to a particular antibody class or subclass, while the remaining portion of the chain is identical or homologous to the corresponding sequence in an antibody derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, provided they exhibit the desired biological activity.

[0043] A "complete" antibody is an antibody that contains an antigen-binding variable region as well as a light chain constant region (C) and heavy chain constant regions CH1, CH2, and CH3. The constant regions can be natural sequence constant regions (e.g., human natural sequence constant regions) or amino acid sequence variants thereof. Preferably, a complete antibody has one or more effector functions.

[0044] An "antibody fragment" comprises a portion of a complete antibody, preferably including its antigen-binding or variable region. Examples of antibody fragments include Fab, Fab′, F(ab′)2, Fv fragments, linear antibodies, and single-chain antibodies (singlechain Fv, scFv).

[0045] The “Fv” fragment is an antibody fragment containing a complete antigen recognition and binding site. This region consists of a dimer composed of a heavy chain and a light chain variable region tightly linked together, and this linkage (as in scFv) can be covalent. In this configuration, the three CDRs of each variable region interact to define the antigen binding site on the surface of the VH-VL dimer.

[0046] The “Fab” fragment comprises the variable and constant regions of the light chain and the variable and first constant region (CH1) of the heavy chain. The F(ab')2 antibody fragment comprises a pair of Fab fragments, which are typically covalently linked near their carboxyl ends via a hinge cysteine ​​residue between them. Other chemical conjugation methods for antibody fragments are also known in the art.

[0047] A single-chain antibody, or "scFv," is an antibody fragment containing the antibody's VH and VL domains. Typically, single-chain antibodies also include a peptide linker between the VH and VL domains, which allows the scFv to form an ideal structure for binding antigens.

[0048] The term "linear antibody" comprises paired tandem Fd segments (VH-CH1-VH-CH1) that, together with a complementary light chain polypeptide, form paired antigen-binding regions. Linear antibodies can be bispecific or monospecific.

[0049] As used herein, the term "antibody variable region" refers to portions of the light and heavy chains of an antibody molecule, including the amino acid sequences of complementarity-determining regions (CDRs: namely CDR1, CDR2, and CDR3) and framework regions (FRs). VH refers to the variable region of the heavy chain. VL refers to the variable region of the light chain. According to the method used in this invention, the amino acid sites specified by CDRs and FRs can be defined by the numbering system described in Abm et al. (Sequences of Proteins of Immunological Interest, 5th edition, Public Health Service, National Institutes of Health, Bethesda, MD (1991)).

[0050] As used herein, the term "complementarity-determining regions" (CDRs: i.e., CDR1, CDR2, and CDR3) refers to the amino acid residues in the antibody variable region that are essential for antigen binding. Each variable region typically has three CDR regions identified as CDR1, CDR2, and CDR3. Each complementarity-determining region may contain amino acid residues from the Kabat-defined "complementarity-determining region" (i.e., approximately residues 24-34 (L1), 50-56 (L2), and 89-97 (L3) in the light chain variable region and residues 31-35 (H1), 50-65 (H2), and 95-102 (H3) in the heavy chain variable region).

[0051] The term "antigen-binding fragment" refers to a portion or segment of a complete antibody with fewer amino acid residues than the complete antibody, capable of binding an antigen or competing with the complete antibody (i.e., the complete antibody from which the antigen-binding fragment originates) for antigen binding. Antigen-binding fragments can be prepared using recombinant DNA technology or by enzymatic or chemical cleavage of complete antibodies. Antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab')2, Fv, single-chain Fv (scFv), single-chain Fab, diabody antibodies, single-domain antibodies (sdAb, nanobodies), camel Ig, IgNAR, F(ab)'3 fragments, bis-scFv, (scFv)2, microantibodies, bifunctional antibodies, trifunctional antibodies, tetrafunctional antibodies, and disulfide-stabilized Fv proteins ("dsFv"). The term also includes genetically engineered forms, such as chimeric antibodies (e.g., humanized mouse antibodies), hybrid binding antibodies (e.g., bispecific antibodies), and their antigen-binding fragments.

[0052] An "antigen binding site" refers to a site on an antibody that provides an interaction with an antigen, typically one or more amino acid residues. For example, the antigen binding site of an antibody may contain amino acid residues from the "complementarity-determining region" (CDR). A natural immunoglobulin molecule generally has two antigen binding sites, while a Fab molecule typically has a single antigen binding site.

[0053] The terms “whole antibody,” “full-length antibody,” “complete antibody,” and “intact antibody” are used interchangeably herein to refer to glycoproteins comprising at least two heavy chains (HC) and two light chains (LC) linked together by disulfide bonds. Each heavy chain consists of a heavy chain variable region (abbreviated as VH) and a heavy chain constant region. The heavy chain constant region consists of three domains: CH1, CH2, and CH3. Each light chain consists of a light chain variable region (abbreviated as VL) and a light chain constant region. The light chain constant region consists of one domain: CL. Mammalian heavy chains are classified as α, δ, ε, γ, and μ. Mammalian light chains are classified as λ or κ. Immunoglobulins containing α, δ, ε, γ, and μ heavy chains are classified as immunoglobulin (Ig) A, IgD, IgE, IgG, and IgM. Complete antibodies form a “Y” shape. The stem of Y consists of the second and third constant regions (and a fourth constant region for IgE and IgM) of two heavy chains linked together, with disulfide bonds (interchain) forming hinges. Heavy chains γ, α, and δ have constant regions consisting of three tandem (in a row) Ig domains, and hinge regions for increased flexibility; heavy chains μ and ε have constant regions consisting of four immunoglobulin domains. The second and third constant regions are referred to as the "CH2 domain" and the "CH3 domain," respectively. Each arm of Y includes a variable region of a single heavy chain and a first constant region that binds to a variable and constant region of a single light chain. The variable regions of the light and heavy chains are responsible for antigen binding.

[0054] The light chain and heavy chain variable regions each contain a "framework" region interspersed with three hypervariable regions (also known as "complementarity-determining regions" or "CDRs"). A "complementarity-determining region" or "CDR" or "hypervariable region" (which can be used interchangeably with "HVR" in this text) is a region within the antibody variable domain that is sequence-hypervariant and forms a structurally defined loop ("hypervariant loop") and / or contains antigen contact residues ("antigen contact sites"). CDRs are primarily responsible for binding to antigen epitopes. The heavy and light chain CDRs are typically referred to as CDR1, CDR2, and CDR3, numbered sequentially starting from the N-terminus. CDRs located within the antibody heavy chain variable domain are referred to as HCDR1, HCDR2, and HCDR3, while those located within the antibody light chain variable domain are referred to as LCDR1, LCDR2, and LCDR3. In a given light chain variable region or heavy chain variable region amino acid sequence, the precise amino acid sequence boundaries of each CDR can be determined using any of a number of known antibody CDR assignment systems or combinations thereof, including, for example, Chothia based on the three-dimensional structure of the antibody and the topology of the CDR ring.

[0055] The “humanized” form of non-human (e.g., rodent) antibodies refers to chimeric antibodies that contain at least a sequence derived from a non-human immunoglobulin. To a large extent, humanized antibodies refer to immunoglobulins in which hypervariable residues of a human immunoglobulin (receptor antibody) are replaced with hypervariable residues of a non-human species (donor antibody) such as mouse, rat, rabbit, or non-human primate, possessing the desired specificity, affinity, and capability. In some cases, framework region (FR) residues of the human immunoglobulin are replaced with corresponding non-human residues. Furthermore, humanized antibodies may contain residues not found in the receptor or donor antibody. These modifications are made to further improve antibody performance. Typically, humanized antibodies will contain substantially no less than one, usually two, variable regions, wherein all or substantially all of the hypervariable loops correspond to the hypervariable loops of the non-human immunoglobulin, and all or substantially all of the FRs are FRs of the human immunoglobulin sequence. Optionally, humanized antibodies will also contain at least a portion of the immunoglobulin constant region (Fc), typically the constant region of a human immunoglobulin.

[0056] The present invention provides an antibody against histone H3 or an antigen-binding fragment thereof, the antibody or antigen-binding fragment thereof comprising a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and the light chain variable region comprises LCDR1, LCDR2 and LCDR3.

[0057] The sequence of HCDR1 is shown in SEQ ID NO:1, SEQ ID NO:17, SEQ ID NO:32, SEQ ID NO:48, SEQ ID NO:71 or SEQ ID NO:83;

[0058] The sequences of HCDR2 are shown in SEQ ID NO:2, SEQ ID NO:18, SEQ ID NO:33, SEQ ID NO:49 or SEQ ID NO:72;

[0059] The sequences of HCDR3 are shown in SEQ ID NO:3, SEQ ID NO:19, SEQ ID NO:34, SEQ ID NO:50, SEQ ID NO:77, SEQ ID NO:85 or SEQ ID NO:89;

[0060] The sequence of LCDR1 is shown in SEQ ID NO:4, SEQ ID NO:20, SEQ ID NO:35 or SEQ ID NO:51;

[0061] The sequence of LCDR2 is shown in SEQ ID NO:5, SEQ ID NO:21, SEQ ID NO:36, SEQ ID NO:52, SEQ ID NO:73 or SEQ ID NO:79;

[0062] The sequences of LCDR3 are shown in SEQ ID NO:6, SEQ ID NO:22, SEQ ID NO:37, SEQ ID NO:53, SEQ ID NO:86, SEQ ID NO:91, SEQ ID NO:93, SEQ ID NO:95 or SEQ ID NO:97.

[0063] In the antibody against histone H3 provided in this application, or its antigen-binding fragment, the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region are selected from any one of the following:

[0064] (1) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, and the sequence of HCDR3 is shown in SEQ ID NO:3;

[0065] (2) The sequence of HCDR1 is shown in SEQ ID NO:17, the sequence of HCDR2 is shown in SEQ ID NO:18, and the sequence of HCDR3 is shown in SEQ ID NO:19.

[0066] (3) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, and the sequence of HCDR3 is shown in SEQ ID NO:34.

[0067] (4) The sequence of HCDR1 is shown in SEQ ID NO:48, the sequence of HCDR2 is shown in SEQ ID NO:49, and the sequence of HCDR3 is shown in SEQ ID NO:50.

[0068] (5) The sequence of HCDR1 is shown in SEQ ID NO:71, the sequence of HCDR2 is shown in SEQ ID NO:72, and the sequence of HCDR3 is shown in SEQ ID NO:3;

[0069] (6) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, and the sequence of HCDR3 is shown in SEQ ID NO:77.

[0070] (7) The sequence of HCDR1 is shown in SEQ ID NO:83, the sequence of HCDR2 is shown in SEQ ID NO:2, and the sequence of HCDR3 is shown in SEQ ID NO:3.

[0071] (8) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, and the sequence of HCDR3 is shown in SEQ ID NO:85.

[0072] (9) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, and the sequence of HCDR3 is shown in SEQ ID NO:89.

[0073] In the antibody against histone H3 or its antigen-binding fragment provided in this application, the LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from any one of the following:

[0074] (1) The sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:5, and the sequence of LCDR3 is shown in SEQ ID NO:6;

[0075] (2) The sequence of LCDR1 is shown in SEQ ID NO:20, the sequence of LCDR2 is shown in SEQ ID NO:21, and the sequence of LCDR3 is shown in SEQ ID NO:22;

[0076] (3) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:37.

[0077] (4) The sequence of LCDR1 is shown in SEQ ID NO:51, the sequence of LCDR2 is shown in SEQ ID NO:52, and the sequence of LCDR3 is shown in SEQ ID NO:53.

[0078] (5) The sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:6;

[0079] (6) The sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:79, and the sequence of LCDR3 is shown in SEQ ID NO:6;

[0080] (7) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:86.

[0081] (8) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:91;

[0082] (9) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:93;

[0083] (10) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:95;

[0084] (11) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:97.

[0085] In the antibody against histone H3 or its antigen-binding fragment provided in this application, the HCDR1, HCDR2, and HCDR3 of the heavy chain variable region and the LCDR1, LCDR2, and LCDR3 of the light chain variable region are selected from any one of the following:

[0086] (1) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:5, and the sequence of LCDR3 is shown in SEQ ID NO:6.

[0087] (2) The sequence of HCDR1 is shown in SEQ ID NO:17, the sequence of HCDR2 is shown in SEQ ID NO:18, the sequence of HCDR3 is shown in SEQ ID NO:19, the sequence of LCDR1 is shown in SEQ ID NO:20, the sequence of LCDR2 is shown in SEQ ID NO:21, and the sequence of LCDR3 is shown in SEQ ID NO:22.

[0088] (3) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:34, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:37.

[0089] (4) The sequence of HCDR1 is shown in SEQ ID NO:48, the sequence of HCDR2 is shown in SEQ ID NO:49, the sequence of HCDR3 is shown in SEQ ID NO:50, the sequence of LCDR1 is shown in SEQ ID NO:51, the sequence of LCDR2 is shown in SEQ ID NO:52, and the sequence of LCDR3 is shown in SEQ ID NO:53.

[0090] (5) The sequence of HCDR1 is shown in SEQ ID NO:71, the sequence of HCDR2 is shown in SEQ ID NO:72, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:6.

[0091] (6) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:77, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:6.

[0092] (7) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:79, and the sequence of LCDR3 is shown in SEQ ID NO:6.

[0093] (8) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:77, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:79, and the sequence of LCDR3 is shown in SEQ ID NO:6.

[0094] (9) The sequence of HCDR1 is shown in SEQ ID NO:83, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:6.

[0095] (10) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:85, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:86.

[0096] (11) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:86.

[0097] (12) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:91.

[0098] (13) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:93.

[0099] (14) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:85, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:95.

[0100] (15) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:95.

[0101] (16) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:97.

[0102] The antibody against histone H3 or its antigen-binding fragment provided in this application further includes a heavy chain variable region comprising the framework regions HFR1-HFR4; the amino acid sequence of HFR1 is as shown in SEQ ID NO:7, SEQ ID NO:23, SEQ ID NO:38, SEQ ID NO:54 or SEQ ID NO:80; the amino acid sequence of HFR2 is as shown in SEQ ID NO:8 or SEQ ID NO:39; the amino acid sequence of HFR3 is as shown in SEQ ID NO:9, SEQ ID NO:24, SEQ ID NO:40 or SEQ ID NO:56; and the amino acid sequence of HFR4 is as shown in SEQ ID NO:10, SEQ ID NO:26, SEQ ID NO:41 or SEQ ID NO:56.

[0103] In the antibody against histone H3 or its antigen-binding fragment provided in this application, the framework region of the heavy chain variable region is selected from any of the following:

[0104] (1) The amino acid sequence of HFR1 is shown in SEQ ID NO:7, the amino acid sequence of HFR2 is shown in SEQ ID NO:8, the amino acid sequence of HFR3 is shown in SEQ ID NO:9, and the amino acid sequence of HFR4 is shown in SEQ ID NO:10.

[0105] (2) The amino acid sequence of HFR1 is shown in SEQ ID NO:23, the amino acid sequence of HFR2 is shown in SEQ ID NO:8, the amino acid sequence of HFR3 is shown in SEQ ID NO:24, and the amino acid sequence of HFR4 is shown in SEQ ID NO:25.

[0106] (3) The amino acid sequence of HFR1 is shown in SEQ ID NO:38, the amino acid sequence of HFR2 is shown in SEQ ID NO:39, the amino acid sequence of HFR3 is shown in SEQ ID NO:40, and the amino acid sequence of HFR4 is shown in SEQ ID NO:41.

[0107] (4) The amino acid sequence of HFR1 is shown in SEQ ID NO:54, the amino acid sequence of HFR2 is shown in SEQ ID NO:8, the amino acid sequence of HFR3 is shown in SEQ ID NO:55, and the amino acid sequence of HFR4 is shown in SEQ ID NO:56.

[0108] (5) The amino acid sequence of HFR1 is shown in SEQ ID NO:80, the amino acid sequence of HFR2 is shown in SEQ ID NO:8, the amino acid sequence of HFR3 is shown in SEQ ID NO:9, and the amino acid sequence of HFR4 is shown in SEQ ID NO:10.

[0109] The antibody against histone H3 or its antigen-binding fragment provided in this application further includes a light chain variable region comprising framework regions LFR1-LFR4; the amino acid sequence of LFR1 is shown in SEQ ID NO:11, SEQ ID NO:26, SEQ ID NO:42, or SEQ ID NO:57; the amino acid sequence of LFR2 is shown in SEQ ID NO:12, SEQ ID NO:27, SEQ ID NO:43, SEQ ID NO:58, or SEQ ID NO:74; the amino acid sequence of LFR3 is shown in SEQ ID NO:13, SEQ ID NO:28, SEQ ID NO:44, or SEQ ID NO:59; and the amino acid sequence of LFR4 is shown in SEQ ID NO:14, SEQ ID NO:29, SEQ ID NO:45, or SEQ ID NO:60.

[0110] In the anti-histone H3 antibody or its antigen-binding fragment provided in this application, the framework region of the light chain variable region is selected from any of the following:

[0111] (1) The amino acid sequence of LFR1 is shown in SEQ ID NO:11, the amino acid sequence of LFR2 is shown in SEQ ID NO:12, the amino acid sequence of LFR3 is shown in SEQ ID NO:13, and the amino acid sequence of LFR4 is shown in SEQ ID NO:14.

[0112] (2) The amino acid sequence of LFR1 is shown in SEQ ID NO:26, the amino acid sequence of LFR2 is shown in SEQ ID NO:27, the amino acid sequence of LFR3 is shown in SEQ ID NO:28, and the amino acid sequence of LFR4 is shown in SEQ ID NO:29.

[0113] (3) The amino acid sequence of LFR1 is shown in SEQ ID NO:42, the amino acid sequence of LFR2 is shown in SEQ ID NO:43, the amino acid sequence of LFR3 is shown in SEQ ID NO:44, and the amino acid sequence of LFR4 is shown in SEQ ID NO:45.

[0114] (4) The amino acid sequence of LFR1 is shown in SEQ ID NO:57, the amino acid sequence of LFR2 is shown in SEQ ID NO:58, the amino acid sequence of LFR3 is shown in SEQ ID NO:59, and the amino acid sequence of LFR4 is shown in SEQ ID NO:60.

[0115] (5) The amino acid sequence of LFR1 is shown in SEQ ID NO:11, the amino acid sequence of LFR2 is shown in SEQ ID NO:74, the amino acid sequence of LFR3 is shown in SEQ ID NO:13, and the amino acid sequence of LFR4 is shown in SEQ ID NO:14.

[0116] The amino acid sequence of the heavy chain variable region in the anti-histone H3 antibody or its antigen-binding fragment provided in this application is shown in SEQ ID NO:15, SEQ ID NO:30, SEQ ID NO:46, SEQ ID NO:61, SEQ ID NO:75, SEQ ID NO:78, SEQ ID NO:81, SEQ ID NO:84, SEQ ID NO:87 or SEQ ID NO:90.

[0117] The amino acid sequence of the light chain variable region in the anti-histone H3 antibody or its antigen-binding fragment provided in this application is shown in SEQ ID NO:16, SEQ ID NO:31, SEQ ID NO:47, SEQ ID NO:62, SEQ ID NO:76, SEQ ID NO:82, SEQ ID NO:88, SEQ ID NO:92, SEQ ID NO:94, SEQ ID NO:96 or SEQ ID NO:98.

[0118] In the antibody against histone H3 or its antigen-binding fragment provided in this application, the amino acid sequences of the heavy chain variable region and the light chain variable region are selected from any of the following:

[0119] (1) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:15; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:16;

[0120] (2) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:30; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:31;

[0121] (3) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:46; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:47;

[0122] (4) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:61; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:62;

[0123] (5) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:75; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:76;

[0124] (6) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:78; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:76;

[0125] (7) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:81; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:82;

[0126] (8) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:78; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:82;

[0127] (9) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:84; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:76;

[0128] (10) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:87; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:88;

[0129] (11) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:88;

[0130] (12) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:92;

[0131] (13) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:94;

[0132] (14) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:87; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:96;

[0133] (15) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:96;

[0134] (16) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:98.

[0135] The amino acid sequences of the heavy chains in the anti-histone H3 antibodies or antigen-binding fragments provided in this application are as shown in SEQ ID NO:63, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, SEQ ID NO:99, SEQ ID NO:101, SEQ ID NO:102, SEQ ID NO:104, SEQ ID NO:105 or SEQ ID NO:107.

[0136] The amino acid sequences of the light chains in the anti-histone H3 antibodies or antigen-binding fragments provided in this application are as shown in SEQ ID NO:64, SEQ ID NO:66, SEQ ID NO:68, SEQ ID NO:70, SEQ ID NO:100, SEQ ID NO:103, SEQ ID NO:106, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110, or SEQ ID NO:111.

[0137] In the anti-histone H3 antibody or its antigen-binding fragment provided in this application, the amino acid sequences of the heavy and light chains of the antibody are selected from any of the following:

[0138] (1) The amino acid sequence of the heavy chain is shown in SEQ ID NO:63, and the amino acid sequence of the light chain is shown in SEQ ID NO:64, namely H_PM_B029 of this application.

[0139] (2) The amino acid sequence of the heavy chain is shown in SEQ ID NO:65, and the amino acid sequence of the light chain is shown in SEQ ID NO:66, namely H_PM_C011 of this application;

[0140] (3) The amino acid sequence of the heavy chain is shown in SEQ ID NO:67, and the amino acid sequence of the light chain is shown in SEQ ID NO:68, namely H_PM_C053 of this application.

[0141] (4) The amino acid sequence of the heavy chain is shown in SEQ ID NO:69, and the amino acid sequence of the light chain is shown in SEQ ID NO:70, namely H_PM_C019 of this application;

[0142] (5) The amino acid sequence of the heavy chain is shown in SEQ ID NO:99, and the amino acid sequence of the light chain is shown in SEQ ID NO:100, namely B029-073-B029-046 of this application.

[0143] (6) The amino acid sequence of the heavy chain is shown in SEQ ID NO:101, and the amino acid sequence of the light chain is shown in SEQ ID NO:100, namely B029-122-B029-046 of this application.

[0144] (7) The amino acid sequence of the heavy chain is shown in SEQ ID NO:102, and the amino acid sequence of the light chain is shown in SEQ ID NO:103, namely B029-075-B029-122 of this application.

[0145] (8) The amino acid sequence of the heavy chain is shown in SEQ ID NO:101, and the amino acid sequence of the light chain is shown in SEQ ID NO:103, namely B029-122-B029-122 of this application;

[0146] (9) The amino acid sequence of the heavy chain is shown in SEQ ID NO:104, and the amino acid sequence of the light chain is shown in SEQ ID NO:100, namely B029-139-B029-046 of this application.

[0147] (10) The amino acid sequence of the heavy chain is shown in SEQ ID NO:105, and the amino acid sequence of the light chain is shown in SEQ ID NO:106, namely C019-084-C019-001 of this application;

[0148] (11) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:106, namely C019-108-C019-001 of this application;

[0149] (12) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:108, namely C019-108-C019-003 of this application.

[0150] (13) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:109, namely C019-108-C019-045 of this application;

[0151] (14) The amino acid sequence of the heavy chain is shown in SEQ ID NO:105, and the amino acid sequence of the light chain is shown in SEQ ID NO:110, namely C019-084-C019-123 of this application.

[0152] (15) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:110, namely C019-108-C019-123 of this application.

[0153] (16) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:111, namely C019-108-C019-130 of this application.

[0154] In a preferred embodiment, the aforementioned B029-073-B029-046, B029-122-B029-046, B029-075-B029-122, B029-122-B029-122, and B029-138-B029-046 are mutants of H_PM_B029.

[0155] The aforementioned C019-084-C019-001, C019-108-C019-001, C019-108-C019-003, C019-0108-C019-045, C019-084-C019-123, C019-108-C019-123, and C019-108-C019-130 are mutants of H_PM_C019.

[0156] In this invention, the amino acid sequences of HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3 are defined using AbM.

[0157] In this application, the aforementioned antibody may be a full-length antibody, a Fab fragment, a Fab' fragment, an F(ab')2 fragment, an Fv fragment, or an scFv fragment or a single-domain antibody.

[0158] In some specific embodiments, the antibody is a full-length antibody, wherein the heavy chain constant region of the aforementioned full-length antibody is derived from the heavy chain of a human antibody or a variant thereof, and the light chain constant region of the aforementioned full-length antibody is derived from the κ chain of a human antibody or a variant thereof.

[0159] In this application, the antibody is a monoclonal antibody, a bispecific antibody, a multispecific antibody, a recombinant antibody, a chimeric antibody, a labeled antibody, an anti-unique antibody, or a fusion protein. In some specific embodiments, the antibody is a monoclonal antibody suitable for specifically binding to histone H3 antigen.

[0160] In this application, the antibody against histone H3 is an IgG1, IgG2, IgG3, or IgG4 antibody; preferably, it is an IgG2 or IgG4 antibody; more preferably, it is an IgG4 antibody, for example, a human IgG4 antibody.

[0161] This application also provides an isolated polynucleotide encoding the aforementioned antihistone H3 antibody or its antigen-binding fragment.

[0162] The polynucleotide is DNA or RNA.

[0163] The present invention also provides a nucleic acid construct containing the isolated polynucleotide.

[0164] In some specific embodiments, the nucleic acid construct is constructed by inserting the aforementioned isolated polynucleotides into the multiple cloning site of the expression vector. The expression vector can be transformed, transduced, or transfected into host cells, allowing its carried genetic material elements to be expressed within the host cells. The construct can be a viral vector or a non-viral vector. For example, non-viral vectors include: plasmids, bacteriophages, Cos plasmids, artificial chromosomes such as yeast artificial chromosomes (YAC), bacterial artificial chromosomes (BAC), or P1-derived artificial chromosomes (PAC), bacteriophages such as λ phage or M13 phage, and animal viruses, etc. Viral vectors include: retroviruses (including lentiviruses), adenoviruses, adeno-associated viruses, herpesviruses (such as herpes simplex virus), poxviruses, baculoviruses, papillomaviruses, and papillomaviruses (such as SV40). The vector may contain various elements controlling expression, including promoter sequences, transcription initiation sequences, enhancer sequences, selection elements, and reporter genes. Additionally, the vector may contain a replication initiation site. The vector may also include components that facilitate its entry into the cell, including but not limited to viral particles, liposomes, or protein coats.

[0165] This application also provides an isolated engineered cell containing the aforementioned nucleic acid construct or the aforementioned polynucleotide.

[0166] In this invention, the engineered cell can be obtained by introducing the aforementioned nucleic acid construct into a host cell, or the engineered cell can be obtained by integrating the aforementioned exogenous polynucleotides into its genome.

[0167] Any cell suitable for expression via the expression vector can serve as a host cell. For example, the host cell can be a prokaryotic cell, such as a bacterial cell; a lower eukaryotic cell, such as a yeast cell; or a higher eukaryotic cell, such as a mammalian cell. The host cell includes many cell types, such as prokaryotic cells like *Escherichia coli* or *Bacillus subtilis*, fungal cells like yeast or *Aspergillus*, insect cells like S2 ​​*Drosophila* or Sf9, or animal cells like fibroblasts, HO cells, COS cells, NSO cells, HeLa cells, BHK cells, HEK293 cells, or human cells.

[0168] The present invention also provides the use of the aforementioned anti-histone H3 antibody or its antigen-binding fragment, the aforementioned polynucleotide or the aforementioned nucleic acid construct in the preparation of disease diagnostic, compound or therapeutic products.

[0169] The therapeutic or diagnostic drug may be a drug that targets histone H3 antigen, binds to or acts on the aforementioned histone H3 antigen, thereby treating and / or preventing the indication.

[0170] In some embodiments of the present invention, the disease is selected from one or more of the following: sepsis, gouty arthritis, acute pancreatitis, rheumatoid arthritis, chronic obstructive pulmonary disease, ischemic cerebrovascular disease, inflammatory bowel disease, acute kidney injury, systemic lupus erythematosus, pulmonary fibrosis, atopic dermatitis, asthma, myocardial infarction, acute respiratory distress syndrome, or psoriasis.

[0171] This application also provides a detection kit containing the aforementioned anti-histone H3 antibody.

[0172] The detection kit typically targets histone H3 antigen, using it as a biomarker for detection. The diagnostic kit may also include a label for anti-histone H3 antibodies. This label can typically be used to label anti-histone H3 antibodies, and the types of labels available include, but are not limited to, combinations of one or more of fluorescent labels, radioactive labels, enzyme-linked immunosorbent assay (ELISA) labels, and chemiluminescent labels. Depending on the detection principle of the kit, the kit may also contain one or more reagents required for the detection. Furthermore, the kit may, as needed, include: containers, controls (negative or positive controls), buffers, and auxiliary agents, which can be selected by those skilled in the art according to specific circumstances.

[0173] This application also provides a pharmaceutical composition comprising the above-described antihistone H3 antibody or its antigen-binding fragment and a pharmaceutically acceptable carrier.

[0174] In some embodiments, the pharmaceutical composition of this application may be administered as a monotherapy (e.g., without the concomitant administration of any other therapeutic agent, or without the concomitant administration of any other therapeutic agent against the same disease to be treated or prevented by the pharmaceutical composition of this application). In some embodiments, the pharmaceutical composition of this application may also be administered in combination with or simultaneously with one or more other therapeutic agents.

[0175] Pharmaceutically acceptable carriers can be commonly used drug carriers, such as polyethylene glycol, including polyethylene glycols with molecular weights in the range of about 200 to about 5000 Da (e.g., PEG200, PEG300, PEG400, or PEG600), ethylene glycol, propylene glycol, glycerin, nonionic surfactants, tylosap, polysorbate 80, polyethylene glycol-15-hydroxy, stearate, phospholipids, lecithin, dimyristoyl phosphatidylcholine, dipalmitoyl phosphatidylcholine, distearate phosphatidylcholine, cyclodextrin, α-cyclodextrin, β-cyclodextrin, γ-cyclodextrin, hydroxyethyl-β-cyclodextrin, hydroxypropyl-β-cyclodextrin, hydroxyethyl-γ-cyclodextrin, etc. Dextrin, hydroxypropyl-γ-cyclodextrin, dihydroxypropyl-β-cyclodextrin, sulfobutyl ether-β-cyclodextrin, sulfobutyl ether-γ-cyclodextrin, glucosyl-α-cyclodextrin, glucosyl-β-cyclodextrin, disaccharosyl-β-cyclodextrin, maltosyl-α-cyclodextrin, maltosyl-β-cyclodextrin, maltosyl-γ-cyclodextrin, maltotriose-β-cyclodextrin, maltotriose-γ-cyclodextrin, disaccharosyl-β-cyclodextrin, methyl-β-cyclodextrin, carboxyalkyl sulfides, hydroxypropyl methylcellulose, hydroxypropyl cellulose, polyvinylpyrrolidone, vinyl acetate copolymer, vinylpyrrolidone, sodium lauryl sulfate, sodium dioctyl sulfosuccinate, or any combination thereof. Those skilled in the art can typically determine the formulation of pharmaceutical compositions based on dosage form requirements.

[0176] This application also provides a treatment method for a disease, the treatment method comprising administering to a subject a therapeutically effective amount of an antihistone H3 antibody or an antigen-binding fragment thereof.

[0177] The disease is selected from one or more of the following: sepsis, gouty arthritis, acute pancreatitis, rheumatoid arthritis, chronic obstructive pulmonary disease, ischemic cerebrovascular disease, inflammatory bowel disease, acute kidney injury, non-alcoholic fatty liver disease, systemic lupus erythematosus, pulmonary fibrosis, atopic dermatitis, asthma, myocardial infarction, acute respiratory distress syndrome, or psoriasis.

[0178] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0179] Before further describing specific embodiments of the present invention, it should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terminology used in the embodiments of the present invention is for describing specific embodiments and not for limiting the scope of protection of the present invention; in the specification and claims of the present invention, unless otherwise expressly stated in the text, the singular forms "a", "an" and "this" include the plural forms.

[0180] When numerical ranges are given in the embodiments, it should be understood that, unless otherwise stated in the present invention, both endpoints of each numerical range and any value between the two endpoints may be selected. Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art. In addition to the specific methods, apparatus, and materials used in the embodiments, based on the knowledge of the prior art possessed by one of ordinary skill in the art and the description of this invention, any prior art methods, apparatus, and materials similar to or equivalent to those described, apparatus, and materials in the embodiments of this invention may be used to implement the present invention.

[0181] In the following examples, H_PR_B029, H_PM_B029, or B029 refer to the same antibody; H_PR_C011, H_PM_C011, or C011 refer to the same antibody; H_PR_C019, H_PM_C019, or C019 refer to the same antibody; and H_PR_C053, H_PM_C053, or C053 refer to the same antibody.

[0182] Example 1 - Construction and Screening of Humanized Recombinant Antibody Library

[0183] In this embodiment, a phage-displaying recombinant antibody gene library was constructed, and Histone(H3&H4)-Avi / histag, Histone-H3-Avi / histag, and Histone-H4-Avi / histag were used as screening antigens to screen the library, obtaining multiple antibody molecules that specifically bind to histones H3&H4, H3, or H4.

[0184] 1) Screening of antibody gene phage display libraries using magnetic bead method

[0185] After biotin labeling the antigen protein, it is then combined with magnetic beads conjugated with streptavidin. The magnetic beads bound to the antigen and the phage display library containing the antibody gene are incubated, washed, and eluted. After three rounds of screening, the specific monoclonal antibodies against the antigen are enriched.

[0186] The specific implementation method for antibody screening is as follows:

[0187] First, biotin-labeled Histone (H3&H4)-Avi / histag, Histone-H3-Avi / histag, and Histone-H4-Avi / histag antigens (human H3 and H4 DNA sequences were obtained through NCBI search, expression vectors were constructed, H3 and H4 were expressed using a prokaryotic expression system, and then purified) were incubated with streptavidin-conjugated magnetic beads to allow the biotin-labeled Histone to bind to the magnetic beads. The histone-binding magnetic beads and the constructed phage library were incubated at room temperature for 2 hours. After washing 6-8 times with PBST to remove non-specifically adsorbed phages, Trypsin was added, the mixture was gently mixed, and the reaction was allowed to proceed for 20 minutes to elute specifically bound antibodies and display the phages. Subsequently, the eluted phages were used to infect SS320 cells in the logarithmic growth phase and allowed to stand for 30 min. Then, the cells were cultured at 220 rpm for 1 h. VSCM13 helper phages were added and allowed to stand for 30 min. The cells were then cultured at 220 rpm for another 1 h. After centrifugation, the medium was replaced with C+ / K+2-YT medium. The resulting phages were then used for the next round of panning.

[0188] 2) Screening of antibody gene phage display libraries using the immunotube method

[0189] Both the immunotube method and the magnetic bead method aim to enrich specific antibodies against antigens, serving as two complementary and validation experimental methods. The principle of the immunotube screening method involves coating Histone (H3&H4)-Avi / histag, Histone-H3-Avi / histag, and Histone-H4-Avi / histag antigens onto the surface of highly absorbent immunotubes. Then, the prepared phages are added to the immunotubes for incubation, washing, and elution. After three rounds of panning, specific monoclonal antibodies against the antigens are enriched.

[0190] The specific implementation method is as follows:

[0191] In the first round of screening, 1 mL of 100 μg / mL Histone (H3&H4)-Avi / histag, Histone-H3-Avi / histag, or Histone-H4-Avi / histag was added to the immunotherapy tubes and incubated overnight at 4°C. The next day, the coating solution was discarded, and the tubes were blocked with 5% milk PBS for 2 hours. After washing twice with PBS, the constructed phage library containing the fully human antibody gene was added and incubated for 2 hours. The tubes were washed 8 times with PBS to remove non-specifically bound phages. Then, 0.8 mL of trypsin digestion solution containing 0.05% EDTA was added to the immunotherapy tubes to elute phages specifically bound to the target antigen. The tubes were then inoculated with logarithmic-phase SS320 cells and incubated at 37°C for 30 minutes. The cells were then cultured at 220 rpm for 1 hour, centrifuged, and transferred to C+ / K+2-YT medium, and cultured overnight at 30°C and 220 rpm. The phages were precipitated the next day for the subsequent two rounds of screening. The antigen coating concentrations used for the second and third rounds of phage screening typically decrease sequentially, to 30 μg / mL and 10 μg / mL, respectively. In addition, the PBS rinsing intensity gradually increases, with PBS elution times of 12 and 16 times, respectively.

[0192] The enrichment effect was assessed using ELISA on the selected dataset. The process included plate coating, blocking, incubation, addition of secondary antibody, color development, termination, and OD value detection. A standard curve was generated using serially diluted known standards to quantify the Fab expression supernatant. The supernatant concentration was plotted on the x-axis, and the OD value on the y-axis, with negative values ​​used as a reference to evaluate the enrichment level of the dataset.

[0193] 3) Initial screening results of monoclonal antibodies

[0194] For datasets with enrichment, single clones were selected for screening using Histone(H3&H4)-Avi / histag, Histone-H3Avi / histag, and Histone-H4-Avi / histag antigen coating methods.

[0195] In the first initial screening, 6 plates were selected, yielding a total of 524 clones. 17 positive clones were found to bind to the Histone (H3&H4) antigen, 16 to the Histone-H3 antigen, and 10 to the Histone-H4 antigen. Eighteen positive clones were selected (10 cross-binding to Histone (H3&H4), Histone-H3, and Histone-H4; 5 cross-binding to Histone (H3&H4) and Histone-H3; 2 binding only to Histone (H3&H4); and 1 binding only to Histone-H3) for sequencing analysis. Seven molecules with unique sequences were obtained. Sequence diversity analysis revealed that two contained cysteine ​​residues and were not recommended for construction, while the remaining five were suitable for full-length construction.

[0196] In the second initial screening, 6 plates were selected, yielding a total of 552 clones. 39 positive clones were found to bind to the Histone (H3&H4) antigen, 44 to the Histone-H3 antigen, and 28 to the Histone-H4 antigen. Fifty positive clones were selected (23 cross-binding to Histone (H3&H4), Histone-H3, and Histone-H4; 13 cross-binding to Histone (H3&H4) and Histone-H3; 1 cross-binding to Histone-H3 and Histone-H4; 4 single-binding to Histone (H3&H4); 5 single-binding to Histone-H3; and 4 single-binding to Histone-H4) for sequencing analysis. Seventeen molecules with unique sequences were obtained. Sequence diversity analysis revealed one glycosylation site, one cysteine ​​residue, one identical CDR region, and one CDR region with ≤3 amino acid differences (construction was not recommended). Thirteen clones were suitable for full-length construction.

[0197] In the third initial screening, 6 plates were selected, yielding a total of 538 clones. 51 positive clones were found to bind to Histone (H3 & H4) antigen, 56 to Histone-H3 antigen, and 37 to Histone-H4 antigen. 59 positive clones were selected (36 cross-binding to Histone (H3 & H4), Histone-H3, and Histone-H4; 16 cross-binding to Histone (H3 & H4) and Histone-H3; 1 cross-binding to Histone-H3 & Histone-H4; 4 single-binding to Histone-H3; and 2 single-binding to Histone-H4) for sequencing. Thirteen molecules with unique sequences were obtained. Sequence diversity analysis revealed 3 molecules containing glycosylation sites, which were not recommended for construction; 10 molecules were suitable for full-length construction.

[0198] A total of 1614 single clones were initially screened by ELISA. 127 positive clones were selected for sequencing analysis, yielding 37 molecules with unique sequences. Sequence diversity analysis further selected 30 molecules for full-length construction. Molecules not selected contained post-translational modifications or had poor diversity and were not included in full-length construction. The amino acid sequences of H_PR_B029, H_PR_C011, H_PM_C019, and H_PM_C053 are shown in Tables 1 and 2.

[0199] Table 1. Amino acid sequences of the light chain and heavy chain variable regions of anti-histone H3 antibodies.

[0200] Table 2. Amino acid sequences of the heavy and light chains of anti-histone H3 antibodies (underlined regions are CDR regions, bolded regions are constant regions)

[0201] Example 2 - Construction, expression, and purification of full-length antibodies

[0202] In this embodiment, 30 molecules specifically targeting Histone (H3&H4), Histone-H3 and Histone-H4 antigens obtained in Example 1 were selected and their full lengths were constructed to prepare target antibodies that meet the requirements.

[0203] 1) Plasmid construction

[0204] From the LSXY-Ab antibody-containing strains obtained through screening, the variable regions of the antibody light and heavy chains were amplified by PCR. These regions were then constructed into the modified eukaryotic expression vector pcDNA3.4 containing the constant regions of the light and heavy chains, respectively, using homologous recombination to obtain protein expression plasmids.

[0205] In addition to PCR amplification, those skilled in the art know that recombinant vectors can also be constructed by synthesizing gene fragments of variable regions.

[0206] 2) Plasmid preparation

[0207] The constructed vector containing the antibody light and heavy chain genes was transformed into E. coli SS320 and cultured overnight at 37°C. The plasmids were extracted using an endotoxin-free plasmid extraction kit to obtain endotoxin-free antibody light and heavy chain plasmids for eukaryotic expression.

[0208] 3) Antibody expression and purification

[0209] The constructed candidate antibody plasmid was transfected into ExpiCHO cells. On the day of transfection, the cell density was confirmed to be 7 × 10⁶. 6 Up to 1×10 7 With approximately 100 viable cells / mL and a cell viability >98%, the cells were then adjusted to a final concentration of 6 × 10⁶ cells / mL using fresh ExpiCHO expression medium pre-warmed at 37°C. 6 Cells / mL. OptiPRO pre-cooled to 4°C TM SFM dilutes the target plasmid (add 1 μg / mL plasmid to 1 mL of culture medium). Simultaneously, OptiPRO is used. TM SFM dilution of ExpiFectamin TM CHO, then mix the two in equal volumes and gently blow to mix well to prepare ExpiFectamin. TM Incubate the CHO / plasmid DNA mixture at room temperature for 1-5 minutes, then slowly add it to the prepared cell suspension while gently shaking. Finally, place the mixture in a cell culture shaker and incubate at 37°C and 8% CO2.

[0210] Add ExpiCHO to the culture medium 18-22 hours after transfection. TM Enhancer and ExpiCHO TM Feed, and place the shake flasks in a shaker at 32°C and 5% CO2 for further incubation. On day 5 post-transfection, add the same volume of ExpiCHO. TM Feed was added slowly while gently mixing the cell suspension. 7-15 days after transfection, the cell culture supernatant expressing the target protein was centrifuged at 15000g for 10 min. The supernatant was then purified using MabSelect SuReLX affinity chromatography, followed by elution with 100mM sodium acetate (pH 3.0), neutralization with 1M Tris-HCl, and finally, the protein was transferred to PBS buffer via ultrafiltration (Millipore, UFC901096). Results showed that the purified protein yields ranged from 0.04 mg to 2.18 mg for 30 target proteins.

[0211] 4) Antibody purity detection

[0212] SDS-PAGE purity identification

[0213] The purified antibody protein concentration was determined according to A280. 1 μg of the non-reduced sample was added to 4×LDS loading buffer and iodoacetamide (final concentration 40 mM), and heated in a dry bath at 75℃ for 10 min. 2 μg of the reduced sample was added to 4×LDS loading buffer and DTT (final concentration 5 mM), and heated in a dry bath at 100℃ for 10 min. Electrophoresis was performed at 140V for 75 min. The gel was stained with Coomassie Brilliant Blue, destained, and then scanned using an EPSON V550 color scanner. The purity of the reduced band, or the purity of the reduced heavy chain plus the light chain, was calculated using ImageJ according to the peak area normalization method. The reference IPI non-reduced band had a molecular weight of approximately 150 kDa and a purity greater than 90%; the reduced heavy chain had a molecular weight of approximately 55 kDa, and the light chain had a molecular weight of approximately 25 kDa, with a heavy chain plus light chain purity greater than 90%. The results are shown in Table 3; the purity of all 28 submitted antibodies was greater than 95.0%.

[0214] Table 3 Results of purity identification of the antibodies to be tested

[0215] SEC purity identification

[0216] Mobile phase preparation: Prepare 0.15 MPa + NaCl, adjust pH to 6.0. Sample preparation: Dilute the sample concentration to 0.5 mg / mL. Column conditions: Use XBridgeBEH. The SEC column was 3.5 μm, 7.8 × 300 mm, and the column temperature was set to 20℃, resulting in a stable detection baseline. Parameter settings included: flow rate of 0.8 mL / min; sample injection volume of 20 μL; detection wavelength of 280 nm with a bandwidth of 4 nm; reference wavelength of 360 nm with a bandwidth of 100 nm; peak width (response time) > 0.1 min (2 s response time); slit width of 4 nm; and a negative absorbance baseline of 100 mAU. If the purity of the reference Herceptin monomer was greater than 95%, the resolution between the BSA monomer and dimer was greater than 1.5, and the baseline was stable, the system was considered to have passed adaptability. The results are shown in Table 3; the purity of all 22 submitted antibodies was greater than 90.0%.

[0217] 5) ELISA-based detection of the affinity activity of candidate antibodies

[0218] Plate preparation: Dilute the antigen sample to a concentration of 2 μg / mL with 1×PBS, add 30 μL / well to each well of a 96-well ELISA plate, and incubate overnight (or more than 12 h) at 4°C. Blocking: Wash the plate 3 times with PBST, add blocking buffer (5% PBSM), and block at room temperature for 2 h. Incubation: Wash the plate 3 times with PBST, add 30 μL / well of candidate antibody diluted with 1% PBSM, and incubate at room temperature for 60 min. Secondary antibody: Wash the plate 3 times with PBST, add diluted Goat-human-IgG-Fc-HRP secondary antibody (purchased from Abcam), and incubate at room temperature for 50 min. Colorimetric development: Wash the plate 3 times with PBST, add 30 μL TMB (purchased from SurModics) to each well. Stop reading: Add 2M stop solution to stop the reaction and measure OD450. ELISA activity assays revealed that among the 30 purified antibodies, 25 antibodies exhibited strong binding activity to the antigen Histone(H3&H4)-Avi / histag; 27 antibodies showed strong binding activity to the antigen Histone-H3-Avi / histag; and 15 antibodies showed strong binding activity to the antigen Histone-H4Avi / histag. The results are shown in Table 4. H_PM_B029 showed strong binding activity to antigens Histone(H3&H4) and Histone-H3, with higher activity than the positive control antibody, but no binding activity to Histone-H4, indicating good specificity for Histone-H3. Similarly, H_PM_C011 showed strong binding activity to antigens Histone(H3&H4) and Histone-H3, with higher activity than the positive control antibody, but weaker binding activity to Histone-H4, indicating poor specificity for Histone-H3. Histone-H3 exhibits good specificity; H_PM_C019 binds to antigens Histone(H3&H4), Histone-H3, and Histone-H3, but not Histone-H4, indicating that H_PM_C019 has good specificity against Histone-H3; H_PM_C053 binds to antigens Histone(H3&H4), Histone-H3, and Histone-H4, and the binding activity is higher than that of the positive control monoclonal antibody, indicating good specificity against Histone-H3.

[0219] Table 4 Summary of ELISA affinity test results for four candidate antibodies - EC 50

[0220] Notes: 1) N represents no binding; 2) Unit: μg / mL;

[0221] Example 3 - Survival analysis of candidate antibodies in a mouse LPS-induced sepsis model

[0222] Ten candidate antibodies with high affinity were selected for survival analysis. Sixty-six wild-type male C57BL / 6 mice (20g ± 1g) were divided into a control LPS group and ten candidate antibody groups. Mice in the LPS group received a direct intravenous injection of 200 μg LPS (10 mg / kg) into the right orbital vein. Mice in the monoclonal antibody group received a direct intravenous injection of 400 μg monoclonal antibody (diluted to 100 μl PBS) into the left orbital vein, followed by a direct intravenous injection of 200 μg LPS into the right orbital vein. The 66 mice were then observed in 66 separate cages, with survival status monitored every 12 hours for three days. The survival rate of each group of mice was recorded: As shown in Figure 1, the survival rate of the LPS model group was 20%, and the monoclonal antibody groups H_PM_B029, H_PM_C011, H_PM_C019 and H_PM_C053 showed significant protective effects with a survival rate of more than 50%.

[0223] Example 4 - In vivo protective effects of candidate antibodies H_PM_B029, H_PM_C011, H_PM_C019 and H_PM_C053 on an LPS-induced sepsis model

[0224] Survival analysis showed that the monoclonal antibody groups H_PM_B029, H_PM_C011, H_PM_C019, and H_PM_C053 provided significant protection. The above four candidate antibodies were used in a sepsis assay. Thirty C57BL / 6 mice weighing 20g ± 1g were divided into a control LPS group and four candidate antibody groups (H_PM_B029, H_PM_C011, H_PM_C019, and H_PM_C053). In the LPS group, mice were directly injected with 200 μg of LPS (10 mg / kg) into the right orbital vein. In the monoclonal antibody group, mice were first injected with 400 μg of monoclonal antibody (diluted to 100 μl PBS) into the left orbital vein, followed by an injection of 200 μg of LPS into the right orbital vein. The mice were then observed in 15 cages. After 24 hours, 1 ml of whole blood was drawn from the inferior vena cava to measure blood routine (WBC / Lym / Neu / Mon / Plt), blood biochemistry (ALT / AST / LDH / BUN / CREA / CK / CRP), and IL-6 levels. After cardiac perfusion, the liver, lungs, kidneys, and other organs of the mice were harvested.

[0225] The results, shown in Figures 2 and 3, indicate that compared to the LPS model group, the liver and kidney functions (ALT, AST, LDH, BUN, CREA) of mice in the monoclonal antibody groups H_PM_B029, H_PM_C011, H_PM_C019, and H_PM_C053 were significantly reduced, while the inflammatory response (CRP) was significantly restored, and blood routine levels were significantly restored compared to the LPS model group. This demonstrates that anti-histone H3 antibodies can effectively alleviate LPS-induced liver and kidney dysfunction and inflammatory damage.

[0226] Example 5 - Affinity Modification of Candidate Antibodies H_PM_B029 and H_PM_C019

[0227] Library Construction

[0228] Four phage display mutant antibody libraries were designed based on the library construction strategy. The H_PM_B029 molecular library was numbered AM2095, AM2096, AM2097, and AM2098, and the H_PM_C019 molecular library was numbered AM2099, AM2100, AM2101, and AM2102, respectively. After electroporation, the library volume was determined by dilution plate method, and the correct insertion rate of antibody genes was verified by single-clone sequencing analysis.

[0229] Document filtering

[0230] Using the immunotube and magnetic bead screening instrument described in the examples, phage mutant antibody libraries were screened using three methods: solid-phase, liquid-phase, and solid-phase-liquid cross-concentration. Antibody molecules that specifically bind to SIGMA-Histone and have a higher affinity than the parent antibody were screened at the ELISA level. Sequence alignment and sequence diversity analysis yielded 39 unique candidate molecules for H_PM_B029 (6 of which were obtained through optimized combinations of light and heavy chains), and 49 unique candidate molecules for H_PM_C019 (16 of which were obtained through optimized combinations of light and heavy chains). Full-length construction was then performed on the candidate molecules from H_PM_B029 and H_PM_C019. Among them, five mutants of H_PM_B029 (B029-073-B029-046, B029-122-B029-046, B029-075-B029-122, B029-122-B029-122, B029-139-B029-046) and seven mutants of H_PM_C019 (C) are mutants of H_PM_C019. The amino acid sequences of (019-084-C019-001, C019-108-C019-001, C019-108-C019-003, C019-108-C019-045, C019-084-C019-123, C019-108-C019-123, C019-108-C019-130) are shown in Tables 5 and 6.

[0231] Table 5. Amino acid sequences of the light chain variable region and heavy chain variable region of the H_PM_B029 and H_PM_C019 mutants.

[0232] Table 6. Amino acid sequences of the heavy and light chains of H_PM_B029 and H_PM_C019 mutants (underlined regions are CDR regions, bolded regions are constant regions)

[0233] Candidate molecule validation

[0234] After eukaryotic expression and purification, candidate molecules were identified by SDS-PAGE, SEC and DSF, detected by ELISA and affinity kinetics.

[0235] The results are shown in Tables 7 and 8. In the H_PM_B029 molecular affinity maturation experiment, a total of 39 candidate molecules were constructed in this antibody modification. The SDS-PAGE purity of all submitted antibodies was greater than 95%, and the purity of all antibodies except 3 antibodies in SEC was greater than 90.0%. ELISA results showed that, except for 9 molecules, the affinity of the remaining molecules was superior to that of the control antibody and the maternal antibody. Affinity kinetics analysis showed that 7 antibodies (B029-073-B029-P, B029-075-B029-046, B029-122-B029-122, B029-073-B029-046, B029-122-B029-046, B029-075-B029-122, B029-122-B029-122, B029-139-B029-046) had an affinity that was more than 5 times higher than that of the maternal antibody. In the H_PM_C019 molecular affinity maturation assay, 49 candidate molecules were constructed through antibody modification. The SDS-PAGE purity of all submitted antibodies was greater than 95%, and the purity of all antibodies except 6 in the SEC assay was greater than 90.0%. The Tm1 values ​​identified by DSF ranged from 62.47℃ to 70.05℃. ELISA results showed that, except for 8 molecules, the affinity of the remaining molecules was superior to the control antibody and the parent antibody. Affinity kinetics... The test showed that seven antibodies (C019-084-C019-001, C019-108-C019-001, C019-108-C019-003, C019-108-C019-045, C019-084-C019-123, C019-108-C019-123, C019-108-C019-130) had an affinity that was more than 5 times higher than that of the maternal antibody.

[0236] Table 7 Summary of ELISA affinity assay results for H_PM_B029 modified and optimized antibody Notes: 1) "N" means not bound; 2) Unit: μg / mL.

[0237] Table 8 Summary of ELISA affinity assay results for H_PM_C019 modified and optimized antibodies

[0238] Example 6 - In vivo protective effect of antibodies H_PM_B029 and H_PM_C019 mutants against an LPS-induced sepsis model

[0239] Affinity experiments showed that the H_PM_B029 mutant (B029-073-B029-046, B029-122-B029-046, B029-075-B029-122, B029-122-B029-122, B029-139-B029-046) and the H_PM_C019 mutant (C019-084-C019-0) were compatible with each other. 01, C019-108-C019-001, C019-108-C019-003, C019-108-C019-045, C019-084-C019-123, C019-108-C019-123, C019-108-C019-130) showed significantly improved affinity compared to the parental lines H_PM_B029 and H_PM_C019. Antibodies C019-108-C019-001, C019-108-C019-045, C019-108-C019-123, C019-108-C019-130, B029-073-B029-046, B029-122-B029-046, B029-075-B029-122, and B029-122-B029-122 were used for acute inflammation experiments. Thirty C57BL / 6 mice weighing 20g±1g were used and divided into negative control, positive control LPS group, and 8 mutant antibody groups. In the negative control group, mice were injected with 100 μl PBS into the left and right orbital veins. In the LPS group, mice were first injected with 250 μg LPS (10 mg / kg) into the right orbital vein, followed by 100 μl PBS into the left orbital vein. In the monoclonal antibody group, mice were first injected with 250 μg LPS (10 mg / kg) into the right orbital vein, followed by 200 μg monoclonal antibody (diluted to 100 μl PBS) into the left orbital vein. 24 h later, 0.5 ml of whole blood was drawn from the inferior vena cava, centrifuged at 12000 rpm for 10 min, and the supernatant plasma was used to determine blood biochemistry (ALT / AST / LDH / BUN / CREA).

[0240] As shown in Figure 4, compared with the negative control group, the liver and kidney functions (ALT, AST, LDH, BUN, CREA) of mice in the positive control group were significantly increased, and the inflammatory response (CRP) was obvious. The blood biochemical levels of liver and kidney function in the mutant antibody group were not significantly different from those in the negative control group, indicating that the inflammatory response in the mutant antibody group was significantly restored.

[0241] Two mutant antibodies (B029-122-B029-122 and B029-122-B029-046) that significantly restored inflammatory damage were used in a dose-dependent experiment. Fifty-six C57BL / 6 mice weighing 20g±1g were randomly divided into a negative control group, a positive control group (LPS group), a wild-type group, and a mutant antibody group. Three doses (50μg, 100μg, and 200μg) were administered to both the wild-type and mutant antibody groups. Mice in the LPS positive control group were first injected intravenously into the right orbital vein. Mice in the wild-type and mutant antibody groups were injected with LPS at a dose of 250 μg (i.e., 10 mg / kg) via the left orbital vein, followed by an injection of 100 μL of PBS. Mice in the wild-type and mutant antibody groups were injected with LPS at a dose of 250 μg (i.e., 10 mg / kg) via the right orbital vein, followed by an injection of 50 / 100 / 200 μg of monoclonal antibody via the left orbital vein. 24 hours later, 0.5 ml of whole blood was drawn from the inferior vena cava and centrifuged at 12000 rpm for 10 min. The supernatant plasma was used to determine blood biochemistry (ALT / AST / LDH / BUN / CREA).

[0242] As shown in Figure 5, compared with the negative control group, the liver and kidney functions (ALT, AST, LDH, BUN, CREA) of mice in the positive control group were significantly increased, and the inflammatory response (CRP) was obvious. The blood biochemical levels of liver and kidney function in the mutant antibody group at each dose showed no significant change compared with the negative control group, indicating that the inflammatory response was significantly restored in the mutant antibody group. As mentioned above, the mutants (B029-122-B029-122 and B029-122-B029-046) have significant anti-inflammatory effects compared with wild-type antibodies.

[0243] Example 7 - In vivo protective effect of antibody H_PM_B029 mutant against LPS-induced sepsis model

[0244] A dose-dependent experiment was conducted using the mutant antibody B029-139-B029-046. Fifteen C57BL / 6 mice weighing 20g±1g were divided into a positive control LPS group and a mutant antibody group. The mutant antibody group was set with three doses (50μg, 100μg, and 200μg). Mice in the LPS positive control group were first injected with 250μg of LPS (i.e., 10mg / kg) into the right orbital vein, followed by 100μl of PBS into the left orbital vein. Mice in the mutant antibody group were first injected with 250μg of LPS (i.e., 10mg / kg) into the right orbital vein, followed by 50 / 100 / 200μg of monoclonal antibody into the left orbital vein. 24h later, 0.5ml of whole blood was drawn from the inferior vena cava and centrifuged at 12000rpm for 10min. The supernatant plasma was used to determine blood biochemistry (ALT / AST / LDH / BUN / CREA).

[0245] As shown in Figure 6, compared with the positive control group, the inflammatory response of liver and kidney function (ALT, AST, LDH, BUN, CREA) in mice in the mutant antibody group was significantly restored. The blood biochemical levels of liver and kidney function in the mutant antibody group at each dose were significantly restored compared with the positive group, and the mutant B029-139-B029-046 exhibited significant anti-inflammatory effects.

[0246] Example 8 - In vivo protective effect of antibody H_PM_B029 mutant against sodium urate-induced acute gouty arthritis model in rats.

[0247] Antibody B029-122-B029-046 was used for an acute gouty arthritis test.

[0248] After acclimatization, the baseline pain threshold of the right hind paw was tested using the Von Frey method. The test was performed three times (with intervals of at least 10 minutes), and the average value was taken as the baseline pain threshold. Paw volume was also measured. Twelve animals were randomly divided into two groups based on body weight, baseline pain threshold, and paw volume: a model group and an antibody 10 mg / kg group, with six animals in each group. After grouping, the rats were anesthetized with isoflurane, and 50 μl of sodium urate solution was injected into the joint cavity at the posterior aspect of the right ankle joint. The test substance was then administered via a single tail vein injection. The paw volume was measured at 2 h, 5 h, 8 h, 24 h, and 48 h post-inflammatory hyperplasia using a paw edema meter, and the swelling inhibition rate was calculated. Simultaneously, the plantar pain threshold was tested using the Von Frey method at the same time points to observe the anti-inflammatory and analgesic effects of the test substance and their time-dependent relationship.

[0249] As shown in Figure 7, the tail vein injection of antibody 10 mg / kg significantly inhibited the reduction of plantar pain threshold and foot volume in acute gouty arthritis induced by sodium urate injection into the ankle joint. Specifically, the plantar pain threshold, foot volume (8-24 hours post-modeling), and swelling rate (5-48 hours post-modeling) in the antibody 10 mg / kg group showed statistically significant differences compared to the model group.

[0250] Example 9 - In vivo protective effect of antibody H_PM_B029 mutant against mouse cerebral ischemia-reperfusion model

[0251] Antibody B029-122-B029-046 was used in a mouse cerebral ischemia-reperfusion experiment.

[0252] After acclimatization, experimental mice were grouped according to their body weight. Mice were fasted overnight before surgery but had free access to water. Preoperatively, mice were anesthetized with isoflurane inhalation, and the MCAO model was established using the Longa method. Thirty minutes after infarction, each group received either saline control or antibody B029-122-B029-046 (10 mg / kg) via tail vein injection. One hour after infarction, the suture embolus was removed, thus completing the cerebral ischemia-reperfusion injury model.

[0253] At the end of the experiment (24 hours post-surgery), changes in cerebral blood flow were measured in all animals using Doppler flowmeter to assess the ameliorative effect of the test substance on the cerebral ischemia-reperfusion model. Behavioral functional tests were conducted using a balance beam. After the balance beam test, whole brain tissue was harvested for brain volume measurement. Subsequently, the brain tissue was prepared into serial coronal sections, stained with TTC, photographed, and the infarct area was measured using Image-J software. The total infarct area was calculated, and the percentage of the infarct area to the total brain area was calculated.

[0254] The results are shown in Figures 8 and 9. In a mouse model of cerebral ischemia-reperfusion injury, a single tail vein injection of antibody B029-122-B029-046 at a dose of 10 mg / kg showed the following: ① The antibody at a dose of 10 mg / kg shortened the average time to cross the beam in the model mice, improving behavioral function (Figure 8A); ② The antibody at a dose of 10 mg / kg significantly reduced postoperative cerebral blood flow difference in the model mice (Figure 8B); ③ As shown in Figures 8C and 9, the antibody at a dose of 10 mg / kg effectively reduced the postoperative infarct area in the model mice.

[0255] Example 10 - Pharmacological study on the effect of antibody H_PM_B029 mutant on DSS-induced ulcerative colitis (Crohn's disease) in mice.

[0256] Antibody B029-122-B029-046 was used in a mouse ulcerative colitis experiment.

[0257] After acclimatization, 12 male C57BL / 6N mice of suitable weight were randomly divided into two groups of 6 mice each. After grouping, the animals were given DSS solution for free drinking to establish the model (day 1 of modeling). The modeling agent was changed every 2 days for 7 consecutive days. During the experiment, the mice's weight was monitored daily, and fecal shape and presence of bleeding were observed to determine the DAI score. On day 1 (D1), mice in the model control group and the test substance B029-122-B029-046 (10 mg / kg) group were treated with the corresponding drug via tail vein injection 30 minutes before drinking the DSS solution (single dose). Mice in the model control group received an equal volume of physiological saline. At the end of the experiment (D8), all mice were anesthetized with deep CO2 inhalation. The abdominal cavity was opened, and the entire colon was removed from the appendix and photographed. The colon was dissected along the mesentery side, and the feces were washed with NS, dried, and the entire colon was weighed and its length measured.

[0258] DAI scoring criteria: (1) Stool condition: formed, pellet (0 points), soft stool (1 point), loose and unformed (2 points), loose stool (3 points); (2) Bleeding condition: no bleeding (0 points), bloody stool or blood around the anus (1 point), severe bleeding (2 points); (3) Weight loss (%): weight loss <1% is 0 points, weight loss 1% to 5% is 1 point, weight loss 5% to 10% is 2 points, weight loss 10% to 15% is 3 points, weight loss >15% is 4 points.

[0259] The experimental results are shown in Figure 10. B029-122-B029-046 did not significantly improve the weight loss induced by DSS in the model mice. However, the test substance group showed improvement in fecal shape and bleeding on the sixth day after drug treatment, and reduced their DAI scores, with the effect lasting until the end of the experiment. The colon length and weight of the mice in the test substance group were significantly greater than those in the model control group, effectively improving colonic atrophy in mice with ulcerative colitis and demonstrating a significant improvement effect on ulcerative colitis in mice.

[0260] Example 11 - Pharmacological study on the effect of antibody H_PM_B029 mutant on acute pancreatitis in mice

[0261] Antibody B029-139-B029-046 was used in an acute pancreatitis test in mice.

[0262] 1. Pharmacological study of a mouse model of acute pancreatitis constructed using L-arginine.

[0263] After acclimatization, Balb / c mice of suitable weight were randomly divided into a model control group and a treatment group, with 8 mice in each group. Both groups were fasted but allowed free access to water for 20 hours. An acute pancreatitis model was established by intraperitoneal injection of 5 g / kg of 20% L-arginine solution, followed by a second injection of 5 g / kg of 20% L-arginine solution 1 hour later. The model control group received intravenous PBS immediately after model establishment, while the treatment group received intravenous injection of 10 mg / kg of antibody drug B029-139-B029-046 immediately after model establishment. Serum amylase and lipase levels were observed in both groups at 16 and 24 hours post-treatment.

[0264] The results, shown in Figures 11A-11B, indicate that administration of the B029-139-B029-046 antibody significantly alleviated the weight loss symptoms in the model mice and significantly reduced the elevation of serum amylase and lipase levels in arginine-induced acute pancreatitis model mice, demonstrating a protective effect. Furthermore, visual observation revealed that the animals in the treatment group exhibited significantly better condition and activity levels than the model group.

[0265] 2. Pharmacological study of an acute pancreatitis model in mice constructed using sodium taurine.

[0266] After acclimatization, male C57BL / 6 mice of suitable weight were randomly divided into a model control group and a drug treatment group, with 8 mice in each group. Acute pancreatitis models were established in both groups using the standard retrograde bile duct injection method with sodium taurocholate. The model control group received intravenous PBS immediately after model establishment, while the drug treatment group received intravenous injection of 10 mg / kg of antibody drug B029-139-B029-046 immediately after model establishment. The mortality rate, serum amylase, and lipase levels were observed 24 hours after drug administration.

[0267] The results, shown in Figures 11C-11F, indicate that administration of the B029-139-B029-046 antibody significantly alleviated weight loss symptoms in the model mice, reduced the mortality rate, and significantly reduced the elevation of serum amylase and lipase levels in arginine-induced acute pancreatitis model mice (excluding the influence of dying animals), demonstrating a protective effect. 24-hour pathological scores also showed that the treated group exhibited mild or slight pancreatic inflammation with lower scores, significantly alleviating the inflammatory response compared to the model group.

[0268] Example 12 - Pharmacological study of antibody H_PM_B029 mutant in a rat model of rheumatoid arthritis

[0269] Antibody B029-139-B029-046 was used in a rat rheumatoid arthritis experiment.

[0270] Female Lewis rats were immunized on days 0 and 7 after acclimatization with 250 μL of an emulsion consisting of 8 mg / ml bovine type II collagen (CII) and an equal volume of Freund's incomplete adjuvant. When the average clinical score (based on the degree of redness and swelling at the lesion site and joint deformity) reached 0.5 or higher, they were randomly assigned to two groups based on clinical score and body weight: a model control group (Vehicle) and a B029-139-B029-046 antibody administration group (n=6 per group). On day 14 after modeling, mice in each group were intravenously administered B029-139-B029-046 at 10 mg / kg (single dose). Mice in the normal control group and model control group received an equal volume of PBS. Animal health was monitored from day 14 to day 28, with clinical scores and paw volume recorded every two days. At the end of the experiment, the hind paws of each rat were collected, weighed, and subjected to pathological analysis.

[0271] The results are shown in Figure 12. Administration of B029-139-B029-046 significantly reduced foot volume, improved toe swelling symptoms, reduced clinical arthritis scores, and relieved joint swelling (Figure 12A). Pathological analysis of arthritis in the hind claw showed a decrease in arthritis scores in all treatment groups (Figure 12B).

[0272] Example 13 - Pharmacodynamic study of antibody H_PM_B029 mutant against a mouse model of chronic obstructive pulmonary disease (COPD)

[0273] Antibody B029-139-B029-046 was used in a mouse COPD experiment.

[0274] After acclimatization, C57BL / 6J mice were used to induce a COPD model using smoke exposure combined with LPS. From D1-D14, D16-D28, and D30-D35, mice were exposed to smoke twice daily (with an interval of more than 3 hours between exposures), with each exposure lasting approximately 40 minutes. On D15 and D29, 50 μL of 0.45 mg / mL LPS was injected into the airway. Mice were divided into a model control group, a single-dose group, and a double-dose group, with 8 mice in each group and 2 mice in each group. The single-dose group received an intravenous injection of 10 mg / kg LPS 0.5 hours before the tracheal injection on D15. The double-dose group received 10 mg / kg LPS 0.5 hours before the tracheal injection on D15 and D29. The model control group received an equal volume of PBS. Lung function was assessed on D36. The main trachea was surgically bluntly dissected, and lung tissue was lavaged with PBS. Bronchoalveolar lavage fluid (BALF) was collected for inflammatory cell detection, and lung tissue was collected for pathological analysis.

[0275] As shown in Figure 13, after administration of B029-139-B029-046, the number of inflammatory cells (white blood cells (WBC), neutrophils, eosinophils, lymphocytes, and monocytes) in the bronchoalveolar lavage fluid significantly decreased, with the two-dose group showing a more significant effect. Pathological analysis showed that B029-139-B029-046 also showed a trend of improving lung injury, significantly improving the mean interalveolar septal wall distance, delaying the onset of emphysema, and with the two-dose group showing better efficacy.

[0276] Example 14 - Pharmacodynamic study of antibody H_PM_B029 mutant in a mouse model of acute kidney injury

[0277] Antibody B029-139-B029-046 was used in an acute kidney injury experiment in mice.

[0278] Acute kidney injury model in BABL / C mice induced by cisplatin

[0279] After acclimatization, BABL / C mice were used to induce an acute kidney injury model using cisplatin. The model was established by a single intraperitoneal injection of 15 mg / kg per mouse. Thirty minutes before cisplatin injection, mice were given 10 mg / kg of cisplatin (B029-139-B029-046). The model control group received an equal volume of physiological saline. Each group consisted of 15 mice. The protective effect of cisplatin on animal survival was observed 72 hours after administration (the experimental endpoint).

[0280] As shown in Figure 14, after administration of B029-139-B029-046, the survival rate of mice in the treatment group increased by 100%, demonstrating a significant protective effect.

[0281] Example 15 - Pharmacological study of antibody H_PM_B029 mutant against mouse model of systemic lupus erythematosus (SLE)

[0282] Antibody B029-139-B029-046 was used in a mouse systemic lupus erythematosus (SLE) experiment.

[0283] After acclimatization, male Balb / c mice were induced to develop SLE (strained leukemia) by collecting urine and serum samples and applying IMQ (imiquimod, Sichuan Mingxin) three times a week. Two weeks after induction, mice were randomly assigned to groups based on anti-dsDNA and mALB / Crea ratios. The treatment group (n=8) received a single intravenous injection of B029-139-B029-046 10 mg / kg once in week 3. The negative control group (n=3) received no induction or treatment. The model group (n=8) received the same volume of PBS concurrently with the treatment group. Throughout the experiment, from grouping and administration to the end of the experiment, the animals were continuously weighed, urine was collected, and serum was collected. At the experimental endpoint, urine biochemistry (mALB / Crea, UTP) and serum ELISA (anti-dsDNA) were measured.

[0284] As shown in Figure 15, after administration of B029-139-B029-046, there were no significant differences in body weight, spleen index, and kidney index between the administration group and the model group and the normal group (Figure 15A). After 5 to 7 weeks, UTP and mALB / CREA were significantly improved in the administration group (Figure 15B). The mean dsDNA content in the administration group was also significantly reduced compared with the model group (Figure 15C), reflecting the efficacy of the drug.

[0285] Example 16 - Pharmacodynamic study of antibody H_PM_B029 mutant against a mouse pulmonary fibrosis model

[0286] Antibody B029-139-B029-046 was used in a mouse pulmonary fibrosis experiment.

[0287] Male C57BL / 6 mice were acclimatized and then used to induce a mouse model of pulmonary fibrosis using BLM. On day 0, all animals were intravenously infused with BLM (1 mg / mL) into their trachea at a dose of 1 mg / kg. Mice were weighed daily after model establishment, and their weight, weight changes, and survival rate were recorded. Starting on day 7, 16 animals were selected based on their weight and randomly divided into two groups, with the average weight of each group being as consistent as possible. On day 7, mice were intravenously administered B029-139-B029-046 10 mg / kg once, while the model group received PBS. There were 8 mice in each group. Survival rates were recorded. On day 21, plasma and bronchoalveolar lavage fluid (BALF) were collected, and CBC analysis was performed to detect the number of white blood cells (WBA), lymphocytes, monocytes, and eosinophils in the BALF.

[0288] The results are shown in Figure 16. After administration of B029-139-B029-046, the survival rate of mice was significantly improved (Figure 16A). The number of inflammatory cells in BALF was detected. The number of white blood cells, lymphocytes, monocytes and eosinophils in BALF were significantly reduced compared with the model group (Figure 16B), indicating that the anti-inflammatory effect of the drug was significant.

[0289] Example 17 - Pharmacological study of antibody H_PM_B029 mutant against a mouse model of atopic dermatitis (AD).

[0290] The experimental results of the effect of antibody B029-139-B029-046 on atopic dermatitis in mice are shown in the figure.

[0291] Female Balb / c mice were acclimatized and then induced with atopic dermatitis by applying 1% OXA (Oxazolone, MCE) to their right ears three times a week on days 0, 2, 4, 7, 9, 11, 14, and 16. Day 1 of modeling was designated as D0. On the day of modeling (D0), mice were divided into groups for drug administration. Three mice in the normal control group were not induced. Eight mice in the model control group were intravenously injected with PBS. Eight mice in each of the single-dose and double-dose groups of B029-139-B029-046 were intravenously injected with 10 mg / kg of B029-139-B029-046. On day 7, eight mice in the double-dose group of B029-139-B029-046 were intravenously injected with 10 mg / kg of B029-139-B029-046. After induction and modeling, the patient was weighed once every day, and symptoms such as ear dryness, rash, ulceration and edema were scored three times a week.

[0292] As shown in Figure 17, after administration of B029-139-B029-046, there was no significant difference in body weight change between the treatment group and the model group and the normal group. The two injections showed significant effects in improving the EASI scores (scaling score change, erythemas score change, and edema score change), especially during the period from Day 11 to Day 16.

[0293] Example 18 - Pharmacodynamic study of antibody H_PM_B029 mutant against a mouse asthma model

[0294] Antibody B029-139-B029-046 was used in a mouse asthma experiment.

[0295] Female Balb / c mice were acclimatized and then used to induce an asthma model using OVA: Sensitization phase: On days 0, 7, and 14, mice were intraperitoneally injected with 200 μL OVA + Al(OH)3 adjuvant to establish the model. On day 20, 16 mice were randomly divided into two groups of 8 mice each, with the weight of each group being as uniform as possible. Animals with excessively large or small weight differences were excluded. Challenge phase: From days 21 to 25, mice were nebulized with 5% OVA for 30 minutes once daily. On day 21, 0.5 hours before nebulization, mice were intravenously administered B029-139-B029-046 10 mg / kg, while the model group received PBS. On day 26, plasma, serum, and bronchoalveolar lavage fluid (BALF) were collected. CBC counts of leukocytes, lymphocytes, neutrophils, and eosinophils in the BALF were performed. Lung tissue was collected for pathological analysis.

[0296] The results are shown in Figure 18. After administration of B029-139-B029-046, the number of inflammatory cells in the BALF fluid of the administration group was reduced compared with that of the model group (Figure 18A). Pathological analysis showed that the airway inflammation was slightly relieved after administration, and the distribution of mucus in the lung tissue of mice was significantly reduced (Figure 18B).

[0297] Example 19 - Pharmacodynamic study of antibody H_PM_B029 mutant in a rat model of myocardial infarction

[0298] Antibody B029-139-B029-046 was used in a rat myocardial infarction experiment.

[0299] Male SD rats were acclimatized and then routinely anesthetized and analgesic before undergoing routine myocardial infarction surgery. The treatment group received a 10 mg / kg drug via tail vein injection immediately post-surgery. The control group received an equal volume of placebo (PBS) immediately post-surgery. Each group consisted of 15 rats. After drug administration, both groups were placed on warming mats until recovery, and then observed for 48 hours for survival.

[0300] The mortality results showed that after administration of B029-139-B029-046, excluding deaths caused by anesthesia stress, the mortality rate in the model group at 48 hours was 22.2%, while no animal deaths occurred in the administration group. B029-139-B029-046 can significantly reduce the mortality rate in rats in the myocardial infarction model and has a significant protective effect.

[0301] Example 20 - Pharmacological study of antibody H_PM_B029 mutant against a mouse model of psoriasis.

[0302] Antibody B029-139-B029-046 was used in a mouse psoriasis experiment.

[0303] Female Balb / c mice were acclimatized and their backs were shaved (2cm x 3cm) one day before model induction. IMQ (Imiquimod, 3M) was applied to the backs once daily for 7 consecutive days to induce a psoriasis mouse model. Day 1 of model induction was designated as D0. Animals were grouped before initial model induction or drug administration, excluding those with significant damage to their backs due to hair growth or shaving. The remaining animals were grouped according to body weight. On the day of grouping, the drug administration group (8 mice) received a single intravenous injection of B029-139-B029-046 10 mg / kg once. The control group (3 mice) did not undergo induction, and the negative control group (8 mice) received PBS after induction. From the initial IMQ application until the end of the experiment, photographs were taken of the backs of the mice on D0, D3, G5, and D7, and the symptoms of erythema, thickening, and desquamation were scored.

[0304] As shown in Figure 19, after administration of B029-139-B029-046, the erythema score, scaling score, and PASI score of the treatment group were significantly lower than those of the model group on days 5 to 6, demonstrating the alleviating effect of the drug on the symptoms of the psoriasis model.

[0305] Example 21 - Pharmacodynamic study of antibody H_PM_B029 mutant against mouse CLP-induced sepsis model

[0306] Antibody B029-122-B029-046 was used in a mouse CLP-induced sepsis experiment.

[0307] After acclimatization, male C57BL / 6 mice were used to establish a cecal ligation-puncture (CLP) model: On day 1 (D1), all mice underwent cecal ligation-puncture surgery. The cecum was ligated with 4-0 silk suture at the midpoint between the cecal tip and the ileocecal valve. Two punctures were made along the midline of the cecum, 0.5 cm apart. Pressure was then applied to the cecum to expel fecal contents approximately 2 mm in size. Within half an hour after the surgery, 8 mice in the treatment group received a single intravenous injection of B029-122-B029-046 10 mg / kg, while 8 mice in the model control group received an equal volume of saline. Blood samples were collected on day 7 to measure AST, ALT, ALP, Tbil, CREA, and UA.

[0308] As shown in Figure 20, after administration of B029-122-B029-046, a significant increase in body weight was observed in the treated group mice at the end of the experiment compared to the model group. B029-122-B029-046 had a protective effect against the weight loss in CLP-induced sepsis model mice. Regarding blood biochemical indicators, B029-122-B029-046 showed a protective effect on liver function in ALP, ALT, and AST levels, while CREA levels also demonstrated a protective effect on kidney function, both delaying further increases in these blood biochemical indicators.

[0309] Example 22 - Pharmacodynamic study of antibody H_PM_B029 mutant against LPS-induced mouse acute respiratory distress syndrome (ARDS) model

[0310] Antibody B029-139-B029-046 was used in an experiment to treat acute respiratory distress syndrome in mice.

[0311] Male C57BL / 6 mice were acclimatized and divided into a control group and a model group. An LPS-induced mouse acute respiratory distress syndrome (ARDS) model was established. On day 0, LPS was instilled orally into the trachea of ​​the model group at a dose of 2.5 mg / kg. The model mice were randomly divided into two groups: a treatment group and a model group, with 8 mice in each group, and the average body weight of the animals in each group was kept as consistent as possible. LPS was administered within 0.5 hours after instillation; the treatment group received 10 mg / kg intravenously, while the model group received PBS. Lung function was assessed 48 hours after LPS instillation. Serum and bronchoalveolar lavage fluid (BALF) were collected. CBC counts of white blood cells, lymphocytes, neutrophils, and eosinophils in the BALF were performed. Lung tissue was collected for HE staining and scoring.

[0312] The results are shown in Figure 21. After administration of B029-139-B029-046, there was no significant difference in body weight change between the treatment group and the model group (G1, PBS) (Figure 21A). Compared with the normal group, the absolute counts of WBC, NEU, and LYM in the BALF of the model group were significantly increased (p<0.05), indicating that the acute respiratory distress syndrome (ARDS) model successfully induced a pulmonary inflammatory response. However, the number of inflammatory cells (WBC, NEU, LYM, MON, BAS) in the BALF of the B029-139-B029-046 group was reduced compared with that of the model group (Figure 21B), indicating that the drug has an inhibitory effect on the pulmonary inflammatory response. In pulmonary function tests, Compared with the model group, the drug-treated group showed improvement in elastic resistance (Ers) index. Simultaneously, the small airway-related index FEF0.1 was significantly increased in the drug-treated group compared with the model group (p<0.05), suggesting that the drug may have a regulatory effect on bronchiolar function (Figure 21C). Compared with the normal group, the model group (G1, PBS) exhibited significant ARDS-like pathological damage, with a significantly increased comprehensive pathological score (p<0.05). Compared with the model group (G1, PBS), after intervention with the test substance, the comprehensive pathological score of mouse lung tissue was significantly reduced (p<0.05) (Figure 21D), indicating that B029-139-B029-046 has a significant effect on improving lung function.

[0313] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any form or substance. It should be noted that those skilled in the art can make various improvements and additions without departing from the method of the present invention, and these improvements and additions should also be considered within the scope of protection of the present invention. Any modifications, alterations, and equivalent changes made by those skilled in the art based on the above-disclosed technical content without departing from the spirit and scope of the present invention are equivalent embodiments of the present invention. Furthermore, any modifications, alterations, and evolutions made to the above embodiments based on the essential technology of the present invention still fall within the scope of the technical solution of the present invention.

Claims

1. An antibody against histone H3 or an antigen-binding fragment thereof, said antibody or antigen-binding fragment comprising a heavy chain variable region and a light chain variable region, said heavy chain variable region comprising HCDR1, HCDR2 and HCDR3, and said light chain variable region comprising LCDR1, LCDR2 and LCDR3; in, The sequences of HCDR1 are shown in SEQ ID NO:1, SEQ ID NO:17, SEQ ID NO:32, SEQ ID NO:48, SEQ ID NO:71 or SEQ ID NO:83; The sequences of HCDR2 are shown in SEQ ID NO:2, SEQ ID NO:18, SEQ ID NO:33, SEQ ID NO:49 or SEQ ID NO:72; The sequences of HCDR3 are shown in SEQ ID NO:3, SEQ ID NO:19, SEQ ID NO:34, SEQ ID NO:50, SEQ ID NO:77, SEQ ID NO:85 or SEQ ID NO:89; The sequence of LCDR1 is shown in SEQ ID NO:4, SEQ ID NO:20, SEQ ID NO:35 or SEQ ID NO:51; The sequence of LCDR2 is shown in SEQ ID NO:5, SEQ ID NO:21, SEQ ID NO:36, SEQ ID NO:52, SEQ ID NO:73 or SEQ ID NO:79; The sequences of LCDR3 are shown in SEQ ID NO:6, SEQ ID NO:22, SEQ ID NO:37, SEQ ID NO:53, SEQ ID NO:86, SEQ ID NO:91, SEQ ID NO:93, SEQ ID NO:95 or SEQ ID NO:

97.

2. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The HCDR1, HCDR2, and HCDR3 of the heavy chain variable region are selected from any one of the following: (1) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, and the sequence of HCDR3 is shown in SEQ ID NO:3; (2) The sequence of HCDR1 is shown in SEQ ID NO:17, the sequence of HCDR2 is shown in SEQ ID NO:18, and the sequence of HCDR3 is shown in SEQ ID NO:

19. (3) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, and the sequence of HCDR3 is shown in SEQ ID NO:

34. (4) The sequence of HCDR1 is shown in SEQ ID NO:48, the sequence of HCDR2 is shown in SEQ ID NO:49, and the sequence of HCDR3 is shown in SEQ ID NO:

50. (5) The sequence of HCDR1 is shown in SEQ ID NO:71, the sequence of HCDR2 is shown in SEQ ID NO:72, and the sequence of HCDR3 is shown in SEQ ID NO:3; (6) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, and the sequence of HCDR3 is shown in SEQ ID NO:

77. (7) The sequence of HCDR1 is shown in SEQ ID NO:83, the sequence of HCDR2 is shown in SEQ ID NO:2, and the sequence of HCDR3 is shown in SEQ ID NO:

3. (8) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, and the sequence of HCDR3 is shown in SEQ ID NO:

85. (9) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, and the sequence of HCDR3 is shown in SEQ ID NO:

89. And / or, the LCDR1, LCDR2, and LCDR3 of the light chain variable regions are selected from any one of the following: (1) The sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:5, and the sequence of LCDR3 is shown in SEQ ID NO:6; (2) The sequence of LCDR1 is shown in SEQ ID NO:20, the sequence of LCDR2 is shown in SEQ ID NO:21, and the sequence of LCDR3 is shown in SEQ ID NO:22; (3) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

37. (4) The sequence of LCDR1 is shown in SEQ ID NO:51, the sequence of LCDR2 is shown in SEQ ID NO:52, and the sequence of LCDR3 is shown in SEQ ID NO:

53. (5) The sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:6; (6) The sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:79, and the sequence of LCDR3 is shown in SEQ ID NO:6; (7) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

86. (8) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:91; (9) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:93; (10) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:95; (11) The sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

97.

3. The antibody or its antigen-binding fragment according to claim 2, characterized in that, The heavy chain variable regions HCDR1, HCDR2, and HCDR3 and the light chain variable regions LCDR1, LCDR2, and LCDR3 are selected from any one of the following: (1) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:5, and the sequence of LCDR3 is shown in SEQ ID NO:

6. (2) The sequence of HCDR1 is shown in SEQ ID NO:17, the sequence of HCDR2 is shown in SEQ ID NO:18, the sequence of HCDR3 is shown in SEQ ID NO:19, the sequence of LCDR1 is shown in SEQ ID NO:20, the sequence of LCDR2 is shown in SEQ ID NO:21, and the sequence of LCDR3 is shown in SEQ ID NO:

22. (3) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:34, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

37. (4) The sequence of HCDR1 is shown in SEQ ID NO:48, the sequence of HCDR2 is shown in SEQ ID NO:49, the sequence of HCDR3 is shown in SEQ ID NO:50, the sequence of LCDR1 is shown in SEQ ID NO:51, the sequence of LCDR2 is shown in SEQ ID NO:52, and the sequence of LCDR3 is shown in SEQ ID NO:

53. (5) The sequence of HCDR1 is shown in SEQ ID NO:71, the sequence of HCDR2 is shown in SEQ ID NO:72, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:

6. (6) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:77, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:

6. (7) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:79, and the sequence of LCDR3 is shown in SEQ ID NO:

6. (8) The sequence of HCDR1 is shown in SEQ ID NO:1, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:77, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:79, and the sequence of LCDR3 is shown in SEQ ID NO:

6. (9) The sequence of HCDR1 is shown in SEQ ID NO:83, the sequence of HCDR2 is shown in SEQ ID NO:2, the sequence of HCDR3 is shown in SEQ ID NO:3, the sequence of LCDR1 is shown in SEQ ID NO:4, the sequence of LCDR2 is shown in SEQ ID NO:73, and the sequence of LCDR3 is shown in SEQ ID NO:

6. (10) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:85, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

86. (11) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

86. (12) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

91. (13) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

93. (14) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:85, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

95. (15) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

95. (16) The sequence of HCDR1 is shown in SEQ ID NO:32, the sequence of HCDR2 is shown in SEQ ID NO:33, the sequence of HCDR3 is shown in SEQ ID NO:89, the sequence of LCDR1 is shown in SEQ ID NO:35, the sequence of LCDR2 is shown in SEQ ID NO:36, and the sequence of LCDR3 is shown in SEQ ID NO:

97.

4. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:15, SEQ ID NO:30, SEQ ID NO:46, SEQ ID NO:61, SEQ ID NO:75, SEQ ID NO:78, SEQ ID NO:81, SEQ ID NO:84, SEQ ID NO:87 or SEQ ID NO:90; and / or, the amino acid sequence of the light chain variable region is shown in SEQ ID NO:16, SEQ ID NO:31, SEQ ID NO:47, SEQ ID NO:62, SEQ ID NO:76, SEQ ID NO:82, SEQ ID NO:88, SEQ ID NO:92, SEQ ID NO:94, SEQ ID NO:96 or SEQ ID NO:

98.

5. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The amino acid sequences of the heavy chain variable region and the light chain variable region are selected from any of the following: (1) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:15; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:16; (2) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:30; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:31; (3) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:46; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:47; (4) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:61; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:62; (5) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:75; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:76; (6) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:78; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:76; (7) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:81; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:82; (8) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:78; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:82; (9) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:84; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:76; (10) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:87; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:88; (11) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:88; (12) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:92; (13) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:94; (14) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:87; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:96; (15) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:96; (16) The amino acid sequence of the heavy chain variable region is shown in SEQ ID NO:90; the amino acid sequence of the light chain variable region is shown in SEQ ID NO:

98.

6. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The amino acid sequence of the heavy chain of the antibody or antigen-binding fragment is shown in SEQ ID NO:63, SEQ ID NO:65, SEQ ID NO:67, SEQ ID NO:69, SEQ ID NO:99, SEQ ID NO:101, SEQ ID NO:102, SEQ ID NO:104, SEQ ID NO:105 or SEQ ID NO:107; and / or, the amino acid sequence of the light chain of the antibody or antigen-binding fragment is shown in SEQ ID NO:64, SEQ ID NO:66, SEQ ID NO:68, SEQ ID NO:70, SEQ ID NO:100, SEQ ID NO:103, SEQ ID NO:106, SEQ ID NO:108, SEQ ID NO:109, SEQ ID NO:110 or SEQ ID NO:

111.

7. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The amino acid sequences of the heavy and light chains of the antibody or its antigen-binding fragment are selected from any of the following: (1) The amino acid sequence of the heavy chain is shown in SEQ ID NO:63, and the amino acid sequence of the light chain is shown in SEQ ID NO:64; (2) The amino acid sequence of the heavy chain is shown in SEQ ID NO:65, and the amino acid sequence of the light chain is shown in SEQ ID NO:

66. (3) The amino acid sequence of the heavy chain is shown in SEQ ID NO:67, and the amino acid sequence of the light chain is shown in SEQ ID NO:

68. (4) The amino acid sequence of the heavy chain is shown in SEQ ID NO:69, and the amino acid sequence of the light chain is shown in SEQ ID NO:70; (5) The amino acid sequence of the heavy chain is shown in SEQ ID NO:99, and the amino acid sequence of the light chain is shown in SEQ ID NO:

100. (6) The amino acid sequence of the heavy chain is shown in SEQ ID NO:101, and the amino acid sequence of the light chain is shown in SEQ ID NO:100; (7) The amino acid sequence of the heavy chain is shown in SEQ ID NO:102, and the amino acid sequence of the light chain is shown in SEQ ID NO:103; (8) The amino acid sequence of the heavy chain is shown in SEQ ID NO:101, and the amino acid sequence of the light chain is shown in SEQ ID NO:103; (9) The amino acid sequence of the heavy chain is shown in SEQ ID NO:104, and the amino acid sequence of the light chain is shown in SEQ ID NO:

100. (10) The amino acid sequence of the heavy chain is shown in SEQ ID NO:105, and the amino acid sequence of the light chain is shown in SEQ ID NO:

106. (11) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:106; (12) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:108; (13) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:109; (14) The amino acid sequence of the heavy chain is shown in SEQ ID NO:105, and the amino acid sequence of the light chain is shown in SEQ ID NO:

110. (15) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:

110. (16) The amino acid sequence of the heavy chain is shown in SEQ ID NO:107, and the amino acid sequence of the light chain is shown in SEQ ID NO:

111.

8. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The antibody or its antigen-binding fragment may be a full-length antibody, a Fab fragment, an F(ab)2 fragment, an Fv fragment, a scFv fragment, or a single-domain antibody.

9. The antibody or its antigen-binding fragment according to claim 1, characterized in that, The antibody or its antigen-binding fragment is a monoclonal antibody, bispecific antibody, multispecific antibody, recombinant antibody, chimeric antibody, labeled antibody, bivalent antibody, anti-idiotype antibody, or fusion protein.

10. An isolated polynucleotide, characterized in that, The polynucleotide encodes an antibody against histone H3 as described in any one of claims 1-9, or an antigen-binding fragment thereof.

11. A nucleic acid construct, characterized in that, The nucleic acid construct contains the polynucleotide as described in claim 10.

12. An isolated engineered cell, characterized in that, The engineered cells contain the nucleic acid construct as described in claim 11 or the polynucleotide as described in claim 10.

13. Use of the antibody against histone H3 as described in any one of claims 1-9 or its antigen-binding fragment, the polynucleotide as described in claim 10, or the nucleic acid construct as described in claim 11 in the preparation of disease diagnostic, preventive, or therapeutic products.

14. The use according to claim 13, characterized in that, The disease is a histone H3-mediated disease; and / or the disease is selected from one or more of the following: sepsis, gouty arthritis, acute pancreatitis, rheumatoid arthritis, chronic obstructive pulmonary disease, ischemic cerebrovascular disease, inflammatory bowel disease, acute kidney injury, systemic lupus erythematosus, pulmonary fibrosis, atopic dermatitis, asthma, myocardial infarction, acute respiratory distress syndrome, or psoriasis.

15. A test kit, characterized in that, The test kit contains an antibody against histone H3 as described in any one of claims 1-9, or an antigen-binding fragment thereof.

16. A pharmaceutical composition comprising an antibody against histone H3 as described in claims 1-9 or an antigen-binding fragment thereof and a pharmaceutically acceptable carrier.

17. A treatment method for a disease, characterized in that, The treatment method comprises administering to a subject a therapeutically effective amount of the antibody against histone H3 as described in any one of claims 1 to 9 or an antigen-binding fragment thereof.

18. The treatment method according to claim 17, characterized in that, The disease is a histone H3-mediated disease; and / or the disease is selected from one or more of the following: sepsis, gouty arthritis, acute pancreatitis, rheumatoid arthritis, chronic obstructive pulmonary disease, ischemic cerebrovascular disease, inflammatory bowel disease, acute kidney injury, systemic lupus erythematosus, pulmonary fibrosis, atopic dermatitis, asthma, myocardial infarction, acute respiratory distress syndrome, or psoriasis.

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