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WO2024146458A1 - Anti-il-11 cytokine humanized antibody - Google Patents

Anti-il-11 cytokine humanized antibody Download PDF

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Publication number
WO2024146458A1
WO2024146458A1 PCT/CN2023/142999 CN2023142999W WO2024146458A1 WO 2024146458 A1 WO2024146458 A1 WO 2024146458A1 CN 2023142999 W CN2023142999 W CN 2023142999W WO 2024146458 A1 WO2024146458 A1 WO 2024146458A1
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seq
antibody
amino acid
variable region
chain variable
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PCT/CN2023/142999
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French (fr)
Chinese (zh)
Inventor
许玉丽
徐愉林
李凯峰
杨子兰
柯志
宁荣良
张晓焰
陈斯琪
汪淑慧
卢伟琦
李文佳
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Sunshine Lake Pharma Co Ltd
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Sunshine Lake Pharma Co Ltd
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/24—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against cytokines, lymphokines or interferons
    • C07K16/244—Interleukins [IL]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/04—Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
    • C07K16/24—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against cytokines, lymphokines or interferons
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6863—Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors
    • G01N33/6869—Interleukin
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/52—Constant or Fc region; Isotype
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
    • C07K2317/565—Complementarity determining region [CDR]
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
    • C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
    • C07K2317/567—Framework region [FR]
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/08—Hepato-biliairy disorders other than hepatitis
    • G01N2800/085—Liver diseases, e.g. portal hypertension, fibrosis, cirrhosis, bilirubin
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/12—Pulmonary diseases
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/32—Cardiovascular disorders
    • G01N2800/325—Heart failure or cardiac arrest, e.g. cardiomyopathy, congestive heart failure

Definitions

  • the present invention relates to the field of biotechnology, specifically, the present invention relates to antibodies against IL-11 and applications thereof, more specifically, the present invention relates to antibodies or antigen-binding fragments thereof, nucleic acid molecules, expression vectors, recombinant cells, pharmaceutical compositions, pharmaceutical uses and kits for detecting IL-11 that can specifically recognize IL-11.
  • interleukin 11 The true physiological role of interleukin 11 (IL-11) is unclear. IL-11 is most closely associated with hematopoietic cell activation and platelet production, but has also been found to have pro- and anti-inflammatory, pro-angiogenic effects, and is important for neoplasia. It is known that TGF ⁇ 1 or tissue injury can induce the expression of IL-11. From the published literature, the role of IL-11 in fibrosis is unclear. IL-11 is considered to be important for lung fibrosis and inflammation, and its expression level is correlated with collagen levels in the skin and respiratory system. However, most studies have shown that IL-11 is anti-fibrotic in the heart and kidney and anti-inflammatory in several tissues and chronic inflammatory diseases. In general, the molecular mode of action of IL-11 is considered to be the mRNA level of RNA expression through STAT3-mediated transcriptional regulation.
  • the present invention develops a humanized Anti-IL-11 antibody with high neutralizing activity, in order to be used for the treatment or prevention of fibrosis-related diseases.
  • the present invention provides an antibody or an antigen-binding fragment thereof that can specifically recognize IL-11.
  • the antibody contains a CDR sequence selected from at least one of the following or an amino acid sequence having at least 95% identity thereto, as well as a heavy chain constant region and a light chain constant region, the light chain constant region of the antibody is derived from a human Kappa light chain constant region, the heavy chain constant region is derived from a human IgG4 heavy chain constant region, the antibody or its antigen-binding fragment The synthesized fragment has humanized modifications.
  • the antibody comprises:
  • the heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 7, 8 and 9, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 7, 8 and 9; or
  • the heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NOs: 13, 14 and 15, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 13, 14 and 15; or
  • the heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 16, 17 and 18, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 16, 17 and 18; or
  • the heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 19, 20 and 21, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 19, 20 and 21; or
  • the heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 28, 29 and 30, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 28, 29 and 30; or
  • the light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 52, 53 and 54, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 52, 53 and 54; or
  • the immunogenicity of the humanized antibody according to the embodiment of the present invention is reduced, and the antibody can specifically target and bind to IL-11, blocking the binding between IL-11 and IL-11 receptor.
  • the use of humanized IgG4 Anti-IL-11 antibody can reduce the risk of thrombocytopenia.
  • GYTFTTNG (SEQ ID NO:1).
  • ARVRSGNDALDF SEQ ID NO:6
  • GYTFTNYG (SEQ ID NO:7).
  • GFSLSSSGMS (SEQ ID NO: 22).
  • the above-mentioned antibody or antigen-binding fragment may further include at least one of the following additional technical features:
  • the antibody contains at least one of a heavy chain framework region sequence and a light chain framework region sequence, and at least a portion of at least one of the heavy chain framework region sequence and the light chain framework region sequence is derived from at least one of a murine antibody, a human antibody, a primate antibody or a mutant thereof.
  • the heavy chain framework region includes FL1, FL2, FL3 and FL4, and the heavy chain framework region FL1, FL2, FL3 and FL4 have the amino acid sequences shown in SEQ ID NOs: 67, 68, 69 and 70, respectively,
  • the antibody has a heavy chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 75 to 78.
  • the full-length sequence of the antibody constant region is shown in SEQ ID NO:83 or 84.
  • the above nucleic acid molecule may further include at least one of the following additional technical features:
  • the nucleic acid molecule is DNA.
  • nucleic acid molecule having the nucleotide sequence shown in SEQ ID NO:93 encodes the heavy chain of the antibody
  • nucleic acid molecule having the nucleotide sequence shown in SEQ ID NO:94 encodes the light chain of the antibody.
  • the above expression vector may further include at least one of the following additional technical features:
  • the expression vector is a eukaryotic expression vector, thereby achieving the expression of the above-mentioned antibody or antigen-binding fragment thereof that specifically recognizes IL-11 in eukaryotic cells, such as CHO cells.
  • the present invention provides a recombinant cell.
  • the recombinant cell carries the aforementioned nucleic acid molecule, or expresses the aforementioned antibody or antigen-binding fragment thereof.
  • the recombinant cell according to an embodiment of the present invention can be used for in vitro expression and mass production of the aforementioned antibody or antigen-binding fragment that specifically recognizes IL-11. get.
  • the above-mentioned recombinant cell may further include at least one of the following additional technical features:
  • the recombinant cell is obtained by introducing the aforementioned expression vector into a host cell.
  • the expression vector is introduced into the host cell by an electrotransduction method.
  • the recombinant cell is a eukaryotic cell.
  • the recombinant cell is a mammalian cell.
  • the present invention provides a pharmaceutical composition.
  • the pharmaceutical composition contains the aforementioned antibody or antigen-binding fragment thereof, the aforementioned nucleic acid molecule, the aforementioned expression vector or the aforementioned recombinant cell.
  • the antibody or expressed antibody contained in the pharmaceutical composition according to an embodiment of the present invention can specifically target and bind to IL-11, blocking the binding of IL-11 to the IL-11 receptor.
  • the present invention proposes a kit for detecting IL-11.
  • the kit includes any of the antibodies described above.
  • the IL-11 antibody described above can specifically target and bind to IL-11.
  • the kit according to an embodiment of the present invention can achieve specific detection of IL-11.
  • a fluorescent detection device can be used to achieve localization or real-time detection of IL-11.
  • the present invention proposes the use of the aforementioned antibody, the aforementioned nucleic acid molecule, the aforementioned expression vector or the aforementioned recombinant cell in the preparation of a kit for detecting IL-11 or diagnosing IL-11-related diseases.
  • FIG1 is a diagram showing the competitive blocking activity of a humanized antibody according to Example 3 of the present invention.
  • FIG2 is a lung inflammation injury score of SD rats after treatment with the humanized antibody according to Example 4 of the present invention.
  • FIG. 3 is a graph showing lung fibrosis scores in SD rats after treatment with the humanized antibody according to Example 4 of the present invention.
  • first and second are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Therefore, the features defined as “first” and “second” may explicitly or implicitly include at least one of the features. In the description of the present invention, the meaning of “plurality” is at least two, such as two, three, etc., unless otherwise clearly and specifically defined.
  • the amino acid composition and arrangement order of some areas have a higher degree of variation, which is called the hypervariable region (HVR).
  • the hypervariable region is the location where the antigen and antibody bind, so it is also called the complementarity-determining region (CDR).
  • CDR complementarity-determining region
  • the amino acid composition and arrangement order outside the hypervariable region in the antibody variable region vary relatively little, which is called the framework region (FR).
  • FR framework region
  • the present invention humanizes the Anti-IL-11 antibody IgG4 with high neutralizing activity, so that the obtained humanized antibody has lower species dependence and reduces the risk of thrombocytopenia.
  • immunogenicity refers to the ability to cause an immune response, that is, the antigen can stimulate specific immune cells, activate, proliferate, and differentiate the immune cells, and ultimately produce immune effector antibodies and sensitized lymphocytes.
  • the inventors of the present application further humanized the screened new mouse-derived neutralizing antibody sequence (humanized modification) to obtain a humanized antibody that retains the affinity and specificity of the parent mouse monoclonal antibody, and greatly reduces its immunogenicity and improves its safety.
  • the present invention provides a humanized antibody or antigen-binding fragment thereof that can specifically recognize IL-11, wherein the antibody contains a CDR sequence selected from at least one of the following or an amino acid sequence that has at least 95% identity thereto: heavy chain variable region CDR sequence: SEQ ID NO: 1-33, light chain variable region CDR sequence: SEQ ID NO: 34-66, and the antibody or antigen-binding fragment thereof has humanized modifications.
  • the antibody or antigen-binding fragment provided by the present invention has conservative amino acid substitutions compared to the above-mentioned heavy chain and light chain.
  • Antigen-binding fragment refers to a humanized antibody or antigen-binding fragment that retains specific Antibody fragments with heterotropic binding antigen ability.
  • Constant amino acid substitution refers to the replacement of an amino acid by another amino acid residue that is biologically, chemically or structurally similar.
  • Biological similarity means that the substitution does not destroy the biological activity of the IL-11 antibody or the binding to the IL-11 antigen.
  • Structurally similar means that the amino acids have side chains of similar length, such as alanine, glycine or serine, or have side chains of similar size.
  • Chemical similarity means that the amino acids have the same charge or are hydrophilic or hydrophobic.
  • the hydrophobic residues isoleucine, valine, leucine or methionine replace each other.
  • polar amino acids replace each other, such as arginine replacing lysine, glutamic acid replacing aspartic acid, glutamine replacing asparagine, serine replacing threonine, etc.
  • the present invention provides an antibody or antigen-binding fragment, which has a heavy chain variable region with an amino acid sequence as shown in any one of SEQ ID NOs: 75 to 78 and a light chain variable region with an amino acid sequence as shown in any one of SEQ ID NOs: 79 to 82.
  • the inventors can obtain the CDR region of the above-mentioned heavy chain variable region sequence (as shown in SEQ ID NOs: 4 to 6) and the CDR region of the light chain variable region sequence (as shown in SEQ ID NOs: 37 to 39) through the antibody sequence comparison database (NCBI, IMGT).
  • the heavy chain variable region sequence of the antibody or antigen-binding fragment has conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NOs: 75 to 78.
  • the light chain variable region sequence of the antibody or antigen-binding fragment has conservative amino acid substitutions compared with the amino acid sequence shown in any one of SEQ ID NOs: 79 to 82.
  • these conservative amino acid substitutions will not change the biological function of the antibody or antigen-binding fragment.
  • these conservative amino acid substitutions may occur in amino acids outside the CDR regions in the heavy chain variable region and the light chain variable region.
  • the present invention provides an anti-IL-11 antibody having a heavy chain having an amino acid sequence shown in any one of SEQ ID NOs: 85 to 88 and a light chain having an amino acid sequence shown in any one of SEQ ID NOs: 89 to 92.
  • nucleic acid molecules expressing these antibodies can be connected to different vectors and then expressed in different cells to obtain the corresponding antibodies.
  • the present invention also provides an isolated nucleic acid molecule, which encodes the antibody or antigen-binding fragment described above.
  • the isolated nucleic acid molecule has at least 90% homology, preferably 95% homology, more preferably 98% or 99% homology with the nucleotide sequence shown in SEQ ID NO: 93.
  • the isolated polynucleotide has at least 90% homology, preferably 95% homology, more preferably 98% or 99% homology with the nucleotide sequence shown in SEQ ID NO: 94.
  • sequences that are homologous to the nucleotide sequence shown in SEQ ID NO: 93 or SEQ ID NO: 94 can express amino acids as shown in the present invention or similar to the present invention, thereby being able to specifically bind to the IL-11 antigen and achieve the targeting function of the antibody.
  • the antibodies of the invention can be incorporated into a pharmaceutical composition suitable for parenteral administration (e.g., intravenous, subcutaneous, intraperitoneal, intramuscular)
  • a pharmaceutical composition suitable for parenteral administration e.g., intravenous, subcutaneous, intraperitoneal, intramuscular
  • the pharmaceutical compositions can be prepared in various forms.
  • liquid, semi-solid and solid dosage forms including but not limited to liquid solutions (e.g., injection solutions and infusion solutions), dispersants or suspensions, tablets, pills, powders, liposomes and suppositories.
  • Typical pharmaceutical compositions are in the form of injection solutions or infusion solutions.
  • the antibodies can be administered by intravenous infusion or injection or intramuscular or subcutaneous injection.
  • the assay method includes the following steps:
  • Example 4 Preferred antibodies inhibit pulmonary fibrosis in SD rats
  • HE staining mainly indicates the level of inflammatory infiltration in the lung tissue.
  • Masson's trichrome staining is mainly used to evaluate the level of pulmonary tissue fibrosis.
  • IgG1 at a dose of 100 mg/kg caused a decrease in PLT (platelets) in rats, which dropped to a minimum of 0.8% after 24 hours, and a recovery trend was seen after 48 hours.

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Abstract

Provided is an antibody capable of specifically recognizing IL-11, or an antigen-binding fragment thereof. The antibody contains a CDR sequence selected from at least one of the following or an amino acid sequence having at least 95% identity thereto, and a heavy chain constant region and a light chain constant region; the light chain constant region of the antibody is derived from a human Kappa light chain constant region, and the heavy chain constant region of the antibody is derived from a human IgG4 heavy chain constant region. The CDR sequence of a heavy chain variable region is as shown in SEQ ID NOs: 1 to 33, the CDR sequence of a light chain variable region is as shown in SEQ IN NOs: 34 to 66, and the antibody or the antigen-binding fragment thereof has a humanization modification. The antibody has low immunogenicity and can specifically target and bind to IL-11 and block the binding of IL-11 to IL-11 receptors.

Description

抗IL-11细胞因子人源化抗体Anti-IL-11 cytokine humanized antibody 技术领域Technical Field

本发明涉及生物技术领域，具体地，本发明涉及IL-11的抗体及其应用，更具体地，本发明涉及能够特异性识别IL-11的抗体或其抗原结合片段、核酸分子、表达载体、重组细胞、药物组合物、制药用途以及检测IL-11的试剂盒。The present invention relates to the field of biotechnology, specifically, the present invention relates to antibodies against IL-11 and applications thereof, more specifically, the present invention relates to antibodies or antigen-binding fragments thereof, nucleic acid molecules, expression vectors, recombinant cells, pharmaceutical compositions, pharmaceutical uses and kits for detecting IL-11 that can specifically recognize IL-11.

背景技术Background technique

纤维化是一个重要的过程，是伤口愈合的关键部分。过度纤维化在许多罕见和常见疾病中很常见，并且在疾病发病机制中很重要。以过度纤维化为特征的疾病包括但不限于：系统性硬化症、硬皮病、肥厚性心肌病、扩张型心肌病(DCM)、心房纤维性颤动、心室纤维性颤动、心肌炎、肝硬化、肾脏疾病、眼部疾病、哮喘、囊性纤维化、关节炎和特发性肺纤维化。尽管对人类健康影响很大，但对纤维化的治疗和诊断方法的医疗需求仍然未得到满足。Fibrosis is an important process and a critical part of wound healing. Excessive fibrosis is common in many rare and common diseases and is important in disease pathogenesis. Diseases characterized by excessive fibrosis include, but are not limited to: systemic sclerosis, scleroderma, hypertrophic cardiomyopathy, dilated cardiomyopathy (DCM), atrial fibrillation, ventricular fibrillation, myocarditis, cirrhosis of the liver, kidney disease, eye disease, asthma, cystic fibrosis, arthritis, and idiopathic pulmonary fibrosis. Despite the significant impact on human health, there remains an unmet medical need for treatments and diagnostics for fibrosis.

白细胞介素11(IL-11)的真正生理作用尚不清楚。IL-11与造血细胞活化和血小板生成的关系最为密切，但也被发现具有促炎和抗炎、促血管生成的作用，并对瘤形成很重要。已知TGFβ1或组织损伤可诱导IL-11的表达。从已发表的文献来看，IL-11在纤维化中的作用尚不清楚。IL-11被认为对肺纤维化和炎症很重要，其表达水平与皮肤和呼吸系统中的胶原水平相关。然而，大多数研究表明IL-11在心脏和肾脏中是抗纤维化的，在几种组织和慢性炎症疾病中的是抗炎的。一般来说，IL-11的分子作用模式被认为是通过STAT3介导的转录调节RNA表达的mRNA水平。The true physiological role of interleukin 11 (IL-11) is unclear. IL-11 is most closely associated with hematopoietic cell activation and platelet production, but has also been found to have pro- and anti-inflammatory, pro-angiogenic effects, and is important for neoplasia. It is known that TGFβ1 or tissue injury can induce the expression of IL-11. From the published literature, the role of IL-11 in fibrosis is unclear. IL-11 is considered to be important for lung fibrosis and inflammation, and its expression level is correlated with collagen levels in the skin and respiratory system. However, most studies have shown that IL-11 is anti-fibrotic in the heart and kidney and anti-inflammatory in several tissues and chronic inflammatory diseases. In general, the molecular mode of action of IL-11 is considered to be the mRNA level of RNA expression through STAT3-mediated transcriptional regulation.

已有研究表明，施用能够抑制白细胞介素11(IL-11)作用的试剂来治疗、预防或缓解需要治疗的受试者中的纤维化。能够抑制IL-11作用的试剂可以阻止或减少IL-11与IL-11受体的结合。Studies have shown that administering agents that inhibit the action of interleukin 11 (IL-11) can treat, prevent or alleviate fibrosis in a subject in need of treatment. Agents that inhibit the action of IL-11 can prevent or reduce the binding of IL-11 to the IL-11 receptor.

发明内容Summary of the invention

本发明开发一款具有高中和活性人源化的Anti-IL-11的抗体，以期用于纤维化相关疾病的治疗或预防。The present invention develops a humanized Anti-IL-11 antibody with high neutralizing activity, in order to be used for the treatment or prevention of fibrosis-related diseases.

在本发明的的第一方面，本发明提出了一种能够特异性识别IL-11的抗体或其抗原结合片段。根据本发明的实施例，所述抗体含有选自下列至少之一的CDR序列或与其具有至少95％同一性的氨基酸序列，以及重链恒定区和轻链恒定区，所述抗体的轻链恒定区来自于人源的Kappa轻链恒定区，重链恒定区来自于人源IgG4的重链恒定区，所述抗体或其抗原结 合片段具有人源化修饰。In the first aspect of the present invention, the present invention provides an antibody or an antigen-binding fragment thereof that can specifically recognize IL-11. According to an embodiment of the present invention, the antibody contains a CDR sequence selected from at least one of the following or an amino acid sequence having at least 95% identity thereto, as well as a heavy chain constant region and a light chain constant region, the light chain constant region of the antibody is derived from a human Kappa light chain constant region, the heavy chain constant region is derived from a human IgG4 heavy chain constant region, the antibody or its antigen-binding fragment The synthesized fragment has humanized modifications.

根据本发明的实施例，所述抗体包括：According to an embodiment of the present invention, the antibody comprises:

分别如SEQ ID NO:1、2和3或者与SEQ ID NO:1、2和3具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 1, 2 and 3, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 1, 2 and 3; or

分别如SEQ ID NO:4、5和6或者与SEQ ID NO:4、5和6具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 4, 5 and 6, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 4, 5 and 6; or

分别如SEQ ID NO:7、8和9或者与SEQ ID NO:7、8和9具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 7, 8 and 9, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 7, 8 and 9; or

分别如SEQ ID NO:10、11和12或者与SEQ ID NO:10、11和12具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NOs: 10, 11 and 12, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 10, 11 and 12; or

分别如SEQ ID NO:13、14和15或者与SEQ ID NO:13、14和15具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NOs: 13, 14 and 15, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 13, 14 and 15; or

分别如SEQ ID NO:16、17和18或者与SEQ ID NO:16、17和18具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 16, 17 and 18, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 16, 17 and 18; or

分别如SEQ ID NO19、20和21或者与SEQ ID NO:19、20和21具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 19, 20 and 21, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 19, 20 and 21; or

分别如SEQ ID NO:22、23和24或者与SEQ ID NO:22、23和24具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 22, 23 and 24, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 22, 23 and 24; or

分别如SEQ ID NO:25、26和27或者与SEQ ID NO:25、26和27具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 25, 26 and 27, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 25, 26 and 27; or

分别如SEQ ID NO:28、29和30或者与SEQ ID NO:28、29和30具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 28, 29 and 30, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 28, 29 and 30; or

分别如SEQ ID NO:31、32和33或者与SEQ ID NO:31、32和33具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列。The heavy chain variable region CDR1, CDR2, and CDR3 sequences are respectively shown in SEQ ID NO: 31, 32, and 33, or amino acid sequences that are at least 95% identical to SEQ ID NO: 31, 32, and 33.

根据本发明的实施例，所述抗体包括：According to an embodiment of the present invention, the antibody comprises:

分别如SEQ ID NO:34、35和36或者与SEQ ID NO:34、35和36具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 34, 35 and 36, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 34, 35 and 36; or

分别如SEQ ID NO:37、38和39或者与SEQ ID NO:37、38和39具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 37, 38 and 39, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 37, 38 and 39; or

分别如SEQ ID NO:40、41和42或者与SEQ ID NO:40、41和42具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者 The light chain variable region CDR1, CDR2, CDR3 sequences shown in SEQ ID NOs: 40, 41 and 42, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 40, 41 and 42; or

分别如SEQ ID NO:43、44和45或者与SEQ ID NO:43、44和45具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 43, 44 and 45, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 43, 44 and 45; or

分别如SEQ ID NO:46、47和48或者与SEQ ID NO:46、47和48具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NOs: 46, 47 and 48, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 46, 47 and 48; or

分别如SEQ ID NO:49、50和51或者与SEQ ID NO:49、50和51具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 49, 50 and 51, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 49, 50 and 51; or

分别如SEQ ID NO:52、53和54或者与SEQ ID NO:52、53和54具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 52, 53 and 54, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 52, 53 and 54; or

分别如SEQ ID NO:55、56和57或者与SEQ ID NO:55、56和57具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 55, 56 and 57, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 55, 56 and 57; or

分别如SEQ ID NO:58、59和60或者与SEQ ID NO:58、59和60具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 58, 59 and 60, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 58, 59 and 60; or

分别如SEQ ID NO:61、62和63或者与SEQ ID NO:61、62和63具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 61, 62 and 63, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 61, 62 and 63; or

分别如SEQ ID NO:64、65和66或者与SEQ ID NO:64、65和66具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列。The light chain variable region CDR1, CDR2, and CDR3 sequences are respectively shown in SEQ ID NO: 64, 65, and 66, or amino acid sequences that are at least 95% identical to SEQ ID NO: 64, 65, and 66.

根据本发明实施例的上述人源化抗体的免疫源性降低，抗体能够特异性的靶向结合IL-11，阻断IL-11和IL-11受体的结合。The immunogenicity of the humanized antibody according to the embodiment of the present invention is reduced, and the antibody can specifically target and bind to IL-11, blocking the binding between IL-11 and IL-11 receptor.

需要说明的是，本申请所述的“人源化修饰”是指能够使得所述抗体或其抗原结合片段的免疫原性降低的氨基酸的改变，包括氨基酸的突变、插入、缺失、化学基团的赘合等等。It should be noted that the “humanization modification” described in the present application refers to changes in amino acids that can reduce the immunogenicity of the antibody or its antigen-binding fragment, including amino acid mutations, insertions, deletions, chemical group superpositions, etc.

根据本发明的实施例，使用人源化IgG4的Anti-IL-11的抗体可降低血小板减少的风险。According to an embodiment of the present invention, the use of humanized IgG4 Anti-IL-11 antibody can reduce the risk of thrombocytopenia.

GYTFTTNG(SEQ ID NO:1)。GYTFTTNG (SEQ ID NO:1).

INTNTGEP(SEQ ID NO:2)。INTNTGEP(SEQ ID NO:2).

AREGAFGLDY(SEQ ID NO:3)。AREGAFGLDY(SEQ ID NO:3).

DYTFTNYW(SEQ ID NO:4)。DYTFTNYW (SEQ ID NO:4).

IYPGGGYT(SEQ ID NO:5)。IYPGGGYT (SEQ ID NO:5).

ARVRSGNDALDF(SEQ ID NO:6)。ARVRSGNDALDF (SEQ ID NO:6).

GYTFTNYG(SEQ ID NO:7)。GYTFTNYG (SEQ ID NO:7).

INTNTGEP(SEQ ID NO:8)。INTNTGEP (SEQ ID NO:8).

SREGIYGMDS(SEQ ID NO:9)。SREGIYGMDS (SEQ ID NO:9).

GFSLTSYG(SEQ ID NO:10)。 GFSLTSYG (SEQ ID NO: 10).

IWAGGRT(SEQ ID NO:11)。IWAGGRT (SEQ ID NO: 11).

AREGGYDYDGDLWT(SEQ ID NO:12)。AREGGYDYDGDLWT (SEQ ID NO:12).

GFSLTSYG(SEQ ID NO:13)。GFSLTSYG (SEQ ID NO: 13).

IWSGGIT(SEQ ID NO:14)。IWSGGIT (SEQ ID NO: 14).

ARNFDGYYYSIDY(SEQ ID NO:15)。ARNFDGYYYSIDY(SEQ ID NO:15).

GFSLTNYG(SEQ ID NO:16)。GFSLTNYG (SEQ ID NO: 16).

IWRGGST(SEQ ID NO:17)。IWRGGST (SEQ ID NO: 17).

AKNLYGPAAMDY(SEQ ID NO:18)。AKNLYGPAAMDY (SEQ ID NO:18).

GDTFTNYA(SEQ ID NO:19)。GDTFTNYA (SEQ ID NO: 19).

IDTYTGDP(SEQ ID NO:20)。IDTYTGDP(SEQ ID NO:20).

AGDTWFAY(SEQ ID NO:21)。AGDTWFAY (SEQ ID NO: 21).

GFSLSSSGMS(SEQ ID NO:22)。GFSLSSSGMS (SEQ ID NO: 22).

IWWSDDK(SEQ ID NO:23)。IWWSDDK (SEQ ID NO: 23).

AREGSLGYGLDY(SEQ ID NO:24)。AREGSLGYGLDY (SEQ ID NO:24).

GFDFRRYW(SEQ ID NO:25)。GFDFRRYW (SEQ ID NO: 25).

INPDSSMI(SEQ ID NO:26)。INPDSSMI (SEQ ID NO: 26).

ATYGHHATDS(SEQ ID NO:27)。ATYGHHATDS (SEQ ID NO:27).

GYTFTSYW(SEQ ID NO:28)。GYTFTSYW (SEQ ID NO: 28).

IYPGSDST(SEQ ID NO:29)。IYPGSDST (SEQ ID NO: 29).

ALDSSGYGFAY(SEQ ID NO:30)。ALDSSGYGFAY (SEQ ID NO:30).

DYEFPSHD(SEQ ID NO:31)。DYEFPSHD (SEQ ID NO:31).

INSDGGNT(SEQ ID NO:32)。INSDGGNT (SEQ ID NO:32).

ARPRPTIGTTATGSSM(SEQ ID NO:33)。ARPRPTIGTTATGSSM (SEQ ID NO:33).

QSVSND(SEQ ID NO:34)。QSVSND (SEQ ID NO:34).

YGS(SEQ ID NO:35)。YGS (SEQ ID NO:35).

QQDFFSPWT(SEQ ID NO:36)。QQDFFSPWT (SEQ ID NO:36).

ESVDEYGISF(SEQ ID NO:37)。ESVDEYGISF (SEQ ID NO:37).

SAS(SEQ ID NO:38)。SAS (SEQ ID NO:38).

QQSKEVPWT(SEQ ID NO:39)。QQSKEVPWT (SEQ ID NO:39).

QSVSND(SEQ ID NO:40)。QSVSND (SEQ ID NO:40).

YAS(SEQ ID NO:41)。 YAS (SEQ ID NO:41).

QQDYYSPWT(SEQ ID NO:42)。QQDYYSPWT (SEQ ID NO:42).

QSLVHSNGNTY(SEQ ID NO:43)。QSLVHSNGNTY (SEQ ID NO:43).

KVS(SEQ ID NO:44)。KVS (SEQ ID NO:44).

SQSTHVPPD(SEQ ID NO:45)。SQSTHVPPD (SEQ ID NO:45).

QDISNS(SEQ ID NO:46)。QDISNS (SEQ ID NO:46).

YTS(SEQ ID NO:47)。YTS (SEQ ID NO:47).

QQGNTLPLT(SEQ ID NO:48)。QQGNTLPLT(SEQ ID NO:48).

QNIYVW(SEQ ID NO:49)。QNIYVW (SEQ ID NO:49).

EAS(SEQ ID NO:50)。EAS (SEQ ID NO:50).

QQGQSYPYT(SEQ ID NO:51)。QQGQSYPYT (SEQ ID NO:51).

QSLLYSRNQKNR(SEQ ID NO:52)。QSLLYSRNQKNR (SEQ ID NO:52).

WAS(SEQ ID NO:53)。WAS (SEQ ID NO:53).

QQYYSYPRT(SEQ ID NO:54)。QQYYSYPRT (SEQ ID NO:54).

QSIVHTDGNTY(SEQ ID NO:55)。QSIVHTDGNTY (SEQ ID NO:55).

KVS(SEQ ID NO:56)。KVS (SEQ ID NO:56).

FQGSHVPWT(SEQ ID NO:57)。FQGSHVPWT (SEQ ID NO:57).

ESVDSYGNSF(SEQ ID NO:58)。ESVDSYGNSF (SEQ ID NO:58).

RAS(SEQ ID NO:59)。RAS (SEQ ID NO:59).

QQSNEDPFT(SEQ ID NO:60)。QQSNEDPFT (SEQ ID NO:60).

QSVSND(SEQ ID NO:61)。QSVSND (SEQ ID NO:61).

YAS(SEQ ID NO:62)。YAS (SEQ ID NO:62).

QQDYSSPFT(SEQ ID NO:63)。QQDYSSPFT (SEQ ID NO:63).

QSVSTSTYNY(SEQ ID NO:64)。QSVSTSTYNY (SEQ ID NO:64).

YAS(SEQ ID NO:65)。YAS (SEQ ID NO:65).

QHSWEIPFT(SEQ ID NO:66)。QHSWEIPFT (SEQ ID NO:66).

根据本发明的实施例，上述抗体或抗原结合片段还可以进一步包括如下附加技术特征至少之一：According to an embodiment of the present invention, the above-mentioned antibody or antigen-binding fragment may further include at least one of the following additional technical features:

根据本发明的实施例，所述抗体含有重链框架区序列和轻链框架区序列的至少之一，所述重链框架区序列和轻链框架区序列的至少之一的至少一部分来自于鼠源抗体、人源抗体、灵长目源抗体或其突变体的至少之一。According to an embodiment of the present invention, the antibody contains at least one of a heavy chain framework region sequence and a light chain framework region sequence, and at least a portion of at least one of the heavy chain framework region sequence and the light chain framework region sequence is derived from at least one of a murine antibody, a human antibody, a primate antibody or a mutant thereof.

根据本发明的实施例，所述重链框架区包括FL1、FL2、FL3和FL4，所述重链框架区 FL1、FL2、FL3和FL4分别具有SEQ ID NO:67、68、69和70所示的氨基酸序列，According to an embodiment of the present invention, the heavy chain framework region includes FL1, FL2, FL3 and FL4, and the heavy chain framework region FL1, FL2, FL3 and FL4 have the amino acid sequences shown in SEQ ID NOs: 67, 68, 69 and 70, respectively,

QVQLVQSGAEVKKPGASVKVSCKAS(SEQ ID NO:67)，QVQLVQSGAEVKKPGASVKVSCKAS (SEQ ID NO: 67),

LGWVRQAPGX1GLEWX2GH(SEQ ID NO:68)，LGWVRQAPGX1GLEWX2GH (SEQ ID NO: 68),

NYNEKFKGRX3TX4TX5DTSTSTVYMELSSLRSEDTAVYX6C(SEQ ID NO:69)，NYNEKFKGRX3TX4TX5DTSTSTVYMELSSLRSEDTAVYX6C (SEQ ID NO: 69),

WGQGTLVTVSS(SEQ ID NO:70)，WGQGTLVTVSS (SEQ ID NO: 70),

其中，X1为Q或H，X2为M或I，X3为V或A，X4为M或L，X5为R或A，X6为Y或F。Among them, X1 is Q or H, X2 is M or I, X3 is V or A, X4 is M or L, X5 is R or A, and X6 is Y or F.

根据本发明的实施例，所述轻链框架区包括FL1、FL2、FL3和FL4，所述轻链框架区FL1、FL2、FL3和FL4分别具有SEQ ID NO:71、72、73和74所示的氨基酸序列，According to an embodiment of the present invention, the light chain framework region includes FL1, FL2, FL3 and FL4, and the light chain framework regions FL1, FL2, FL3 and FL4 have the amino acid sequences shown in SEQ ID NO: 71, 72, 73 and 74, respectively.

EIVLTQSPATLSLSPGERATLSCRAS(SEQ ID NO:71)，EIVLTQSPATLSLSPGERATLSCRAS (SEQ ID NO:71),

MNWX7QQKPGQAPRLLIY(SEQ ID NO:72)，MNWX7QQKPGQAPRLLIY (SEQ ID NO: 72),

NQGSGIPARFSGSGSGTDFTLTISSLEPEDFAX8YX9C(SEQ ID NO:73)，NQGSGIPARFSGSGSGTDFTLTISSLEPEDFAX8YX9C (SEQ ID NO:73),

FGGGTKLEIK(SEQ ID NO:74)，FGGGTKLEIK (SEQ ID NO: 74),

其中，X7为Y或F，X8为V或M，X9为Y或F。Among them, X7 is Y or F, X8 is V or M, and X9 is Y or F.

根据本发明的实施例，所述抗体具有如SEQ ID NO:75～78任一项所示氨基酸序列的重链可变区。
According to an embodiment of the present invention, the antibody has a heavy chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 75 to 78.

根据本发明的实施例，所述抗体具有如SEQ ID NO:79～82任一项所示氨基酸序列的轻链可变区。

According to an embodiment of the present invention, the antibody has a light chain variable region having an amino acid sequence as shown in any one of SEQ ID NOs: 79 to 82.

根据本发明的实施例，所述抗体恒定区的全长序列如SEQ ID NO:83或84所示。
According to an embodiment of the present invention, the full-length sequence of the antibody constant region is shown in SEQ ID NO:83 or 84.

其中，上述SEQ ID NO:83所示氨基酸序列为抗体重链恒定区序列；上述SEQ ID NO:84所示的氨基酸序列为抗体轻链恒定区序列。Among them, the amino acid sequence shown in the above SEQ ID NO:83 is the antibody heavy chain constant region sequence; the amino acid sequence shown in the above SEQ ID NO:84 is the antibody light chain constant region sequence.

根据本发明的实施例，所述抗体具有SEQ ID NO:85～88任一项所示氨基酸序列的重链和具有SEQ ID NO:89～92任一项所示氨基酸序列的轻链。


According to an embodiment of the present invention, the antibody has a heavy chain having an amino acid sequence shown in any one of SEQ ID NOs: 85 to 88 and a light chain having an amino acid sequence shown in any one of SEQ ID NOs: 89 to 92.


根据本发明的实施例，所述抗体为单链抗体、多聚体抗体、CDR移植抗体或小分子抗体。According to an embodiment of the present invention, the antibody is a single-chain antibody, a multimeric antibody, a CDR-grafted antibody or a small molecule antibody.

根据本发明的实施例，所述小分子抗体包括Fab抗体、Fv抗体、单链抗体以及最小识别单位的至少之一。According to an embodiment of the present invention, the small molecule antibody includes at least one of a Fab antibody, a Fv antibody, a single-chain antibody and a minimum recognition unit.

在本发明的第二方面，本发明提出了一种核酸分子。根据本发明的实施例，所述核酸分子编码前面所述的抗体或其抗原结合片段。根据本发明实施例的核酸分子所编码的抗体或抗原结合片段免疫源性低，可特异性靶向结合IL-11，阻断IL-11和IL-11受体的结合。In a second aspect of the present invention, the present invention provides a nucleic acid molecule. According to an embodiment of the present invention, the nucleic acid molecule encodes the above-mentioned antibody or antigen-binding fragment thereof. The antibody or antigen-binding fragment encoded by the nucleic acid molecule according to an embodiment of the present invention has low immunogenicity, can specifically target and bind to IL-11, and block the binding of IL-11 and IL-11 receptor.

根据本发明的实施例，上述核酸分子还可以进一步包括如下附加技术特征至少之一：According to an embodiment of the present invention, the above nucleic acid molecule may further include at least one of the following additional technical features:

根据本发明的实施例，所述核酸分子为DNA。According to an embodiment of the present invention, the nucleic acid molecule is DNA.

根据本发明的实施例，所述核酸分子具有如SEQ ID NO:93或94所示核苷酸序列。

According to an embodiment of the present invention, the nucleic acid molecule has a nucleotide sequence as shown in SEQ ID NO:93 or 94.

其中，具有SEQ ID NO:93所示的核苷酸序列的核酸分子编码抗体的重链；具有SEQ ID NO:94所示的核苷酸序列的核酸分子编码抗体的轻链。Among them, the nucleic acid molecule having the nucleotide sequence shown in SEQ ID NO:93 encodes the heavy chain of the antibody; the nucleic acid molecule having the nucleotide sequence shown in SEQ ID NO:94 encodes the light chain of the antibody.

在本发明的第三方面，本发明提出了一种表达载体。根据本发明的实施例，所述表达载体携带前面所述的核酸分子。根据本发明实施例的表达载体导入合适的受体细胞后，可在调控系统的介导下，有效实现前面所述的特异性识别IL-11的抗体或其抗原结合片段表达，进而实现所述抗体或抗原结合片段的体外大量获得。In the third aspect of the present invention, the present invention proposes an expression vector. According to an embodiment of the present invention, the expression vector carries the aforementioned nucleic acid molecule. After the expression vector according to the embodiment of the present invention is introduced into a suitable recipient cell, the aforementioned antibody or antigen-binding fragment thereof that specifically recognizes IL-11 can be effectively expressed under the mediation of a regulatory system, thereby achieving a large amount of in vitro acquisition of the antibody or antigen-binding fragment.

根据本发明的实施例，上述表达载体还可以进一步包括如下附加技术特征至少之一：According to an embodiment of the present invention, the above expression vector may further include at least one of the following additional technical features:

根据本发明的实施例，所述表达载体为真核表达载体。进而实现前面所述的特异性识别IL-11的抗体或其抗原结合片段在真核细胞中的表达，如CHO细胞。According to an embodiment of the present invention, the expression vector is a eukaryotic expression vector, thereby achieving the expression of the above-mentioned antibody or antigen-binding fragment thereof that specifically recognizes IL-11 in eukaryotic cells, such as CHO cells.

在本发明的第四方面，本发明提出了一种重组细胞。根据本发明的实施例，所述重组细胞携带前面所述的核酸分子，或者表达前面所述的抗体或其抗原结合片段。根据本发明实施例的重组细胞可用于前面所述的特异性识别IL-11的抗体或其抗原结合片段体外表达和大量 获得。In a fourth aspect of the present invention, the present invention provides a recombinant cell. According to an embodiment of the present invention, the recombinant cell carries the aforementioned nucleic acid molecule, or expresses the aforementioned antibody or antigen-binding fragment thereof. The recombinant cell according to an embodiment of the present invention can be used for in vitro expression and mass production of the aforementioned antibody or antigen-binding fragment that specifically recognizes IL-11. get.

根据本发明的实施例，上述重组细胞还可以进一步包括如下附加技术特征至少之一：According to an embodiment of the present invention, the above-mentioned recombinant cell may further include at least one of the following additional technical features:

根据本发明的实施例，所述重组细胞是通过将前面所述的表达载体引入至宿主细胞中而获得的。According to an embodiment of the present invention, the recombinant cell is obtained by introducing the aforementioned expression vector into a host cell.

根据本发明的实施例，通过电转导的方法将所述表达载体引入所述宿主细胞中。According to an embodiment of the present invention, the expression vector is introduced into the host cell by an electrotransduction method.

根据本发明的实施例，所述重组细胞为真核细胞。According to an embodiment of the present invention, the recombinant cell is a eukaryotic cell.

根据本发明的实施例，所述重组细胞为哺乳动物细胞。According to an embodiment of the present invention, the recombinant cell is a mammalian cell.

在本发明的第五方面，本发明提出了一种药物组合物。根据本发明的实施例，所述药物组合物含有前面所述的抗体或其抗原结合片段，前面所述的核酸分子，前面所述的表达载体或前面所述的重组细胞。根据本发明实施例的药物组合物中所包含的抗体或表达的抗体能够特异性的靶向结合IL-11，阻断IL-11和IL-11受体的结合。In a fifth aspect of the present invention, the present invention provides a pharmaceutical composition. According to an embodiment of the present invention, the pharmaceutical composition contains the aforementioned antibody or antigen-binding fragment thereof, the aforementioned nucleic acid molecule, the aforementioned expression vector or the aforementioned recombinant cell. The antibody or expressed antibody contained in the pharmaceutical composition according to an embodiment of the present invention can specifically target and bind to IL-11, blocking the binding of IL-11 to the IL-11 receptor.

在本发明的第六方面，本发明提出了前面所述的抗体或其抗原结合片段、前面所述的核酸分子、前面所述的表达载体或前面所述的重组细胞、前面所述的药物组合物在制备药物中的用途，所述药物用于治疗或者肺纤维化(Pulmonary fibrosis)，非酒精性脂肪肝炎(non-alcoholic steatohepatitis,NASH)，慢性心力衰竭(Chronic heart failure,CHF)或由成纤维细胞向纤维化转化所导致的疾病。In the sixth aspect of the present invention, the present invention proposes the use of the aforementioned antibody or antigen-binding fragment thereof, the aforementioned nucleic acid molecule, the aforementioned expression vector or the aforementioned recombinant cell, and the aforementioned pharmaceutical composition in the preparation of a drug, wherein the drug is used to treat pulmonary fibrosis (Pulmonary fibrosis), non-alcoholic steatohepatitis (NASH), chronic heart failure (Chronic heart failure, CHF) or diseases caused by the transformation of fibroblasts to fibrosis.

在本发明的第七方面，本发明提出了一种检测IL-11的试剂盒。根据本发明的实施例，所述试剂盒包括前面任一所述的抗体。前面所述的IL-11抗体能够特异性靶向结合IL-11，根据本发明实施例的试剂盒可以实现IL-11的特异性检测，如当抗体结合有荧光基团时，可以采用荧光检测装置实现对IL-11的定位或实时检测。In the seventh aspect of the present invention, the present invention proposes a kit for detecting IL-11. According to an embodiment of the present invention, the kit includes any of the antibodies described above. The IL-11 antibody described above can specifically target and bind to IL-11. The kit according to an embodiment of the present invention can achieve specific detection of IL-11. For example, when the antibody is bound to a fluorescent group, a fluorescent detection device can be used to achieve localization or real-time detection of IL-11.

在本发明的第八方面，本发明提出了前面所述的抗体、前面所述的核酸分子、前面所述的表达载体或前面所述的重组细胞在制备试剂盒中的用途，所述试剂盒用于检测IL-11或者诊断IL-11相关的疾病。In the eighth aspect of the present invention, the present invention proposes the use of the aforementioned antibody, the aforementioned nucleic acid molecule, the aforementioned expression vector or the aforementioned recombinant cell in the preparation of a kit for detecting IL-11 or diagnosing IL-11-related diseases.

本发明的附加方面和优点将在下面的描述中部分给出，部分将从下面的描述中变得明显，或通过本发明的实践了解到。Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention.

附图说明BRIEF DESCRIPTION OF THE DRAWINGS

本发明的上述和/或附加的方面和优点从结合下面附图对实施例的描述中将变得明显和容易理解，其中：The above and/or additional aspects and advantages of the present invention will become apparent and easily understood from the description of the embodiments in conjunction with the following drawings, in which:

图1是根据本发明实施例3的人源化抗体竞争性阻断活性；FIG1 is a diagram showing the competitive blocking activity of a humanized antibody according to Example 3 of the present invention;

图2是根据本发明实施例4的人源化抗体治疗后SD大鼠肺部炎症损伤评分； FIG2 is a lung inflammation injury score of SD rats after treatment with the humanized antibody according to Example 4 of the present invention;

图3是根据本发明实施例4的人源化抗体治疗后SD大鼠肺部纤维化评分。FIG. 3 is a graph showing lung fibrosis scores in SD rats after treatment with the humanized antibody according to Example 4 of the present invention.

具体实施方式Detailed ways

下面详细描述本发明的实施例，所述实施例的示例在附图中示出，其中自始至终相同或类似的标号表示相同或类似的元件或具有相同或类似功能的元件。Embodiments of the present invention are described in detail below, examples of which are shown in the accompanying drawings, wherein the same or similar reference numerals throughout represent the same or similar elements or elements having the same or similar functions.

此外，术语“第一”、“第二”仅用于描述目的，而不能理解为指示或暗示相对重要性或者隐含指明所指示的技术特征的数量。由此，限定有“第一”、“第二”的特征可以明示或者隐含地包括至少一个该特征。在本发明的描述中，“多个”的含义是至少两个，例如两个，三个等，除非另有明确具体的限定。In addition, the terms "first" and "second" are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Therefore, the features defined as "first" and "second" may explicitly or implicitly include at least one of the features. In the description of the present invention, the meaning of "plurality" is at least two, such as two, three, etc., unless otherwise clearly and specifically defined.

抗体Antibody

本文中，术语“抗体”是能够与特异性抗原结合的免疫球蛋白分子。包括两条分子量较轻的轻链和两条分子量较重的重链，重链(H链)和轻链(L链)由二硫键连接形成一个四肽链分子。其中，肽链的氨基端(N端)氨基酸序列变化很大，称为可变区(V区)，羧基端(C端)相对稳定，变化很小，称为恒定区(C区)。L链和H链的V区分别称为VL和VH。As used herein, the term "antibody" refers to an immunoglobulin molecule that can bind to a specific antigen. It includes two light chains with a relatively low molecular weight and two heavy chains with a relatively high molecular weight. The heavy chain (H chain) and the light chain (L chain) are connected by disulfide bonds to form a tetrapeptide chain molecule. Among them, the amino acid sequence at the amino end (N end) of the peptide chain varies greatly, which is called the variable region (V region), while the carboxyl end (C end) is relatively stable and varies little, which is called the constant region (C region). The V regions of the L chain and H chain are called VL and VH, respectively.

在可变区中某些区域氨基酸组成和排列顺序具有更高的变化程度，称为高变区(Hypervariable region，HVR)，高变区为抗原和抗体结合的位置，因此也称为决定簇互补区(complementarity-determining region，CDR)。重链可变区和轻链可变区上均有三个CDR区。抗体可变区内高变区之外的氨基酸组成和排列顺序变化相对较小，称为框架区(FR)。VH和VL内各有4个框架区，分别以FR1、FR2、FR3和FR4表示。In the variable region, the amino acid composition and arrangement order of some areas have a higher degree of variation, which is called the hypervariable region (HVR). The hypervariable region is the location where the antigen and antibody bind, so it is also called the complementarity-determining region (CDR). There are three CDR regions in the heavy chain variable region and the light chain variable region. The amino acid composition and arrangement order outside the hypervariable region in the antibody variable region vary relatively little, which is called the framework region (FR). There are 4 framework regions in VH and VL, represented by FR1, FR2, FR3 and FR4 respectively.

本发明通过对具有高中和活性的Anti-IL-11抗体IgG4人源化改造，所获得的人源化抗体具有较低的种属依赖性，降低了血小板减少的风险。The present invention humanizes the Anti-IL-11 antibody IgG4 with high neutralizing activity, so that the obtained humanized antibody has lower species dependence and reduces the risk of thrombocytopenia.

需要说明的是，“免疫原性”是指能引起免疫应答的性能，即抗原能刺激特定的免疫细胞，使免疫细胞活化、增殖、分化，最终产生免疫效应物质抗体和致敏淋巴细胞的特性。本申请的发明人进一步对筛选得到的鼠源的全新中和抗体序列进行人源化改造(人源化修饰)，得到人源化抗体，该人源化抗体既能保留亲本鼠单克隆抗体的亲和力和特异性，又大幅降低了其免疫原性，提高了其安全性。It should be noted that "immunogenicity" refers to the ability to cause an immune response, that is, the antigen can stimulate specific immune cells, activate, proliferate, and differentiate the immune cells, and ultimately produce immune effector antibodies and sensitized lymphocytes. The inventors of the present application further humanized the screened new mouse-derived neutralizing antibody sequence (humanized modification) to obtain a humanized antibody that retains the affinity and specificity of the parent mouse monoclonal antibody, and greatly reduces its immunogenicity and improves its safety.

在一些实施方案中，本发明提供了一种能够特异性识别IL-11的人源化抗体或其抗原结合片段，所述抗体含有选自下列至少之一的CDR序列或与其具有至少95％同一性的氨基酸序列：重链可变区CDR序列：SEQ ID NO:1～33，轻链可变区CDR序列：SEQ IN NO:34～66，所述抗体或其抗原结合片段具有人源化修饰。在另一些实施方案中，本发明所提供的抗体或者抗原结合片段与上述重链和轻链相比，具有保守氨基酸取代。“抗原结合片段”是指保持特 异性结合抗原能力的抗体片段。“保守氨基酸取代”指的是氨基酸被另一氨基酸发生生物学上、化学上或者结构上相似的残基所取代。生物学上相似的指的是该取代不破坏IL-11抗体或者与IL-11抗原结合的生物学活性。结构上相似指的是氨基酸具有相似长度的侧链，如丙氨酸、甘氨酸或丝氨酸，或具有相似大小的侧链。化学相似性指的是氨基酸具有相同的电荷或者都是亲水或者疏水的。例如疏水残基异亮氨酸、缬氨酸、亮氨酸或者甲硫氨酸相互取代。或者极性氨基酸相互取代，例如用精氨酸取代赖氨酸、谷氨酸取代天冬氨酸、谷氨酰胺取代天冬酰胺，丝氨酸取代苏氨酸等等。In some embodiments, the present invention provides a humanized antibody or antigen-binding fragment thereof that can specifically recognize IL-11, wherein the antibody contains a CDR sequence selected from at least one of the following or an amino acid sequence that has at least 95% identity thereto: heavy chain variable region CDR sequence: SEQ ID NO: 1-33, light chain variable region CDR sequence: SEQ ID NO: 34-66, and the antibody or antigen-binding fragment thereof has humanized modifications. In other embodiments, the antibody or antigen-binding fragment provided by the present invention has conservative amino acid substitutions compared to the above-mentioned heavy chain and light chain. "Antigen-binding fragment" refers to a humanized antibody or antigen-binding fragment that retains specific Antibody fragments with heterotropic binding antigen ability. "Conservative amino acid substitution" refers to the replacement of an amino acid by another amino acid residue that is biologically, chemically or structurally similar. Biological similarity means that the substitution does not destroy the biological activity of the IL-11 antibody or the binding to the IL-11 antigen. Structurally similar means that the amino acids have side chains of similar length, such as alanine, glycine or serine, or have side chains of similar size. Chemical similarity means that the amino acids have the same charge or are hydrophilic or hydrophobic. For example, the hydrophobic residues isoleucine, valine, leucine or methionine replace each other. Or polar amino acids replace each other, such as arginine replacing lysine, glutamic acid replacing aspartic acid, glutamine replacing asparagine, serine replacing threonine, etc.

在一些实施方案中，本发明提供了一种抗体或抗原结合片段，所述抗体或抗原结合片段具有SEQ ID NO:75～78任一项所示氨基酸序列的重链可变区和具有如SEQ ID NO:79～82任一项所示氨基酸序列的轻链可变区。发明人通过抗体序列比对数据库(NCBI、IMGT)可得到上述抗重链可变区序列的CDR区(如SEQ ID NO:4～6所示)和轻链可变区序列的CDR区(如SEQ ID NO:37～39所示)。在另一些实施方案中，所述抗体或抗原结合片段的重链可变区序列与SEQ ID NO:75～78所示氨基酸序列相比，具有保守氨基酸取代。在一些实施方案中，所述抗体或抗原结合片段的轻链可变区序列与SEQ ID NO:79～82任一项所示氨基酸序列相比，具有保守氨基酸取代。当然，这些保守氨基酸取代不会对抗体或者抗原结合片段的生物学功能带来改变。在一些具体方式中，这些保守氨基酸取代可以发生在重链可变区和轻链可变区中除了CDR区之外的氨基酸上。In some embodiments, the present invention provides an antibody or antigen-binding fragment, which has a heavy chain variable region with an amino acid sequence as shown in any one of SEQ ID NOs: 75 to 78 and a light chain variable region with an amino acid sequence as shown in any one of SEQ ID NOs: 79 to 82. The inventors can obtain the CDR region of the above-mentioned heavy chain variable region sequence (as shown in SEQ ID NOs: 4 to 6) and the CDR region of the light chain variable region sequence (as shown in SEQ ID NOs: 37 to 39) through the antibody sequence comparison database (NCBI, IMGT). In other embodiments, the heavy chain variable region sequence of the antibody or antigen-binding fragment has conservative amino acid substitutions compared with the amino acid sequence shown in SEQ ID NOs: 75 to 78. In some embodiments, the light chain variable region sequence of the antibody or antigen-binding fragment has conservative amino acid substitutions compared with the amino acid sequence shown in any one of SEQ ID NOs: 79 to 82. Of course, these conservative amino acid substitutions will not change the biological function of the antibody or antigen-binding fragment. In some specific embodiments, these conservative amino acid substitutions may occur in amino acids outside the CDR regions in the heavy chain variable region and the light chain variable region.

在一些优选方案中，本发明提供了一种抗IL-11抗体，该抗体具有SEQ ID NO:85～88任一项所示氨基酸序列的重链和具有SEQ ID NO:89～92任一项所示氨基酸序列的轻链。In some preferred embodiments, the present invention provides an anti-IL-11 antibody having a heavy chain having an amino acid sequence shown in any one of SEQ ID NOs: 85 to 88 and a light chain having an amino acid sequence shown in any one of SEQ ID NOs: 89 to 92.

在一些优选方案中，本发明提供了一种抗IL-11单链抗体。In some preferred embodiments, the present invention provides an anti-IL-11 single-chain antibody.

核酸分子、表达载体、重组细胞Nucleic acid molecules, expression vectors, recombinant cells

在制备或者获取这些抗体的过程中，可以利用表达这些抗体的核酸分子，与不同的载体连接，然后在不同细胞中表达，来获得相应抗体。In the process of preparing or obtaining these antibodies, nucleic acid molecules expressing these antibodies can be connected to different vectors and then expressed in different cells to obtain the corresponding antibodies.

为此，本发明还提供了一种分离的核酸分子，所述核酸分子编码上述所述的抗体或抗原结合片段。To this end, the present invention also provides an isolated nucleic acid molecule, which encodes the antibody or antigen-binding fragment described above.

在一些实施方案中，所述分离核酸分子具有如SEQ ID NO:93所示核苷酸序列或具有SEQ ID NO:94所示核苷酸序列。In some embodiments, the isolated nucleic acid molecule has a nucleotide sequence shown in SEQ ID NO:93 or has a nucleotide sequence shown in SEQ ID NO:94.

在一些实施方案中，所述分离的核酸分子与上述SEQ ID NO:93所示的核苷酸序列至少具有90％以上的同源性，优选具有95％以上的同源性，更优选具有98％、99％以上的同源性。在至少一些实施方案中，所述分离的多核苷酸与所述SEQ ID NO:94所示的核苷酸序列至少具有90％以上的同源性，优选具有95％以上的同源性，更优选具有98％、99％以上的同源性。 这些与SEQ ID NO:93或者SEQ ID NO:94所示核苷酸序列具有同源性的序列，能够表达如本发明所示或与本发明相似的氨基酸，从而能够与IL-11抗原特异性结合，实现抗体的靶向性功能。In some embodiments, the isolated nucleic acid molecule has at least 90% homology, preferably 95% homology, more preferably 98% or 99% homology with the nucleotide sequence shown in SEQ ID NO: 93. In at least some embodiments, the isolated polynucleotide has at least 90% homology, preferably 95% homology, more preferably 98% or 99% homology with the nucleotide sequence shown in SEQ ID NO: 94. These sequences that are homologous to the nucleotide sequence shown in SEQ ID NO: 93 or SEQ ID NO: 94 can express amino acids as shown in the present invention or similar to the present invention, thereby being able to specifically bind to the IL-11 antigen and achieve the targeting function of the antibody.

在一些优选实施方式中，所述分离的核酸分子具有SEQ ID NO:93所示的重链核苷酸序列和SEQ ID NO:94所示的轻链核苷酸序列。这些核苷酸序列经过种属优化，更易在哺乳动物细胞中表达。In some preferred embodiments, the isolated nucleic acid molecule has a heavy chain nucleotide sequence shown in SEQ ID NO: 93 and a light chain nucleotide sequence shown in SEQ ID NO: 94. These nucleotide sequences are species optimized and are more easily expressed in mammalian cells.

本发明还提供了一种表达载体，所述表达载体包含上述分离的核酸分子。在将上述分离的多核苷酸连接到载体上时，可以将多核苷酸与载体上的控制元件直接或者间接相连，只要这些控制元件能够控制多核苷酸的翻译和表达等即可。当然这些控制元件可以直接来自于载体本身，也可以是外源性的，即并非来自于载体本身。当然，多核苷酸与控制元件进行可操作地连接即可。本文中“可操作地连接”是指将外源基因连接到载体上，使得载体内的控制元件，例如转录控制序列和翻译控制序列等等，能够发挥其预期的调节外源基因的转录和翻译的功能。当然用来编码抗体重链和轻链的多核苷酸，可以分别独立的插入到不同的载体上，常见的是插入到同一载体上。常用的载体例如可以为质粒、噬菌体等等。例如Plasmid-X质粒。The present invention also provides an expression vector, which comprises the above-mentioned isolated nucleic acid molecule. When the above-mentioned isolated polynucleotide is connected to the vector, the polynucleotide can be directly or indirectly connected to the control elements on the vector, as long as these control elements can control the translation and expression of the polynucleotide. Of course, these control elements can come directly from the vector itself, or they can be exogenous, that is, they are not from the vector itself. Of course, the polynucleotide can be operably connected to the control element. Herein, "operably connected" means connecting the exogenous gene to the vector so that the control elements in the vector, such as transcription control sequences and translation control sequences, etc., can play their expected functions of regulating the transcription and translation of the exogenous gene. Of course, the polynucleotides used to encode the heavy chain and light chain of the antibody can be inserted into different vectors separately and independently, and it is common to insert them into the same vector. Commonly used vectors can be, for example, plasmids, bacteriophages, etc. For example, Plasmid-X plasmid.

本发明还提供了一种重组细胞，该重组细胞中包含有该表达载体。可以将表达载体导入到哺乳动物细胞中，构建获得重组细胞，然后利用这些重组细胞表达本发明提供的抗体或者抗原结合片段。通过该重组细胞进行培养，即可以获得相应抗体。这些可用的哺乳动物细胞例如可以为CHO细胞等。The present invention also provides a recombinant cell, which contains the expression vector. The expression vector can be introduced into mammalian cells to construct recombinant cells, and then these recombinant cells are used to express the antibody or antigen-binding fragment provided by the present invention. The corresponding antibody can be obtained by culturing the recombinant cells. These available mammalian cells can be, for example, CHO cells.

药物组合物、试剂盒及制药用途和在制备试剂盒中的用途。Pharmaceutical composition, kit and pharmaceutical use and use in preparing a kit.

本发明还提供了一种药物组合物，所述药物组合物包括上述所述的抗体或者抗原结合片段和药学可接受的载体。The present invention also provides a pharmaceutical composition, which comprises the above-mentioned antibody or antigen-binding fragment and a pharmaceutically acceptable carrier.

本文提供的抗IL-11抗体可以掺入适合受试者施用的药物组合物中。通常，这些药物组合物包括本文提供的抗IL-11抗体以及药学上可接受的载体。The anti-IL-11 antibodies provided herein can be incorporated into pharmaceutical compositions suitable for administration to a subject. Generally, these pharmaceutical compositions include the anti-IL-11 antibodies provided herein and a pharmaceutically acceptable carrier.

“药学上可接受的载体”可以包括生理学上相容的任何和所有溶剂、分散介质、包衣、抗细菌剂和抗真菌剂、等渗剂和延迟吸收剂等等。具体实例可以是水、盐水、磷酸盐缓冲盐水、葡萄糖、甘油、乙醇等以及它们的组合物中的一种或多种。有许多情况下，药物组合物中包括等渗剂，例如糖类、多元醇(如甘露醇、山梨醇)或氯化钠等。当然药学上可接受的载体还可包括微量的辅助物质，例如润湿剂或乳化剂、防腐剂或缓冲剂，用来延长抗体的保存限期或效力。"Pharmaceutically acceptable carriers" may include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents and agents for delayed absorption, etc. that are physiologically compatible. Specific examples may be one or more of water, saline, phosphate buffered saline, glucose, glycerol, ethanol, etc., and combinations thereof. In many cases, isotonic agents such as sugars, polyols (such as mannitol, sorbitol) or sodium chloride are included in the pharmaceutical composition. Of course, pharmaceutically acceptable carriers may also include trace amounts of auxiliary substances, such as wetting agents or emulsifiers, preservatives or buffers, to extend the shelf life or efficacy of the antibody.

例如，本发明的抗体可掺入适用于胃肠外施用(例如，静脉内、皮下、腹膜内、肌肉内) 的药物组合物中。这些药物组合物可以被制备成各种形式。例如液体、半固体和固体剂型等，包括但不限于液体溶液(例如，注射溶液和输注溶液)、分散剂或悬浮剂、片剂、丸剂、粉末、脂质体和栓剂。典型的药物组合物为注射溶液或输注溶液形式。所述抗体可通过静脉输注或注射或肌肉内或皮下注射来施用。For example, the antibodies of the invention can be incorporated into a pharmaceutical composition suitable for parenteral administration (e.g., intravenous, subcutaneous, intraperitoneal, intramuscular) The pharmaceutical compositions can be prepared in various forms. For example, liquid, semi-solid and solid dosage forms, including but not limited to liquid solutions (e.g., injection solutions and infusion solutions), dispersants or suspensions, tablets, pills, powders, liposomes and suppositories. Typical pharmaceutical compositions are in the form of injection solutions or infusion solutions. The antibodies can be administered by intravenous infusion or injection or intramuscular or subcutaneous injection.

当然，本文中的抗IL-11抗体还可以根据需要被制成试剂盒或者其他诊断性试剂的一部分。根据本发明的实施例，本发明还提供了一种试剂盒，所述试剂盒包括上述IL-11抗体。应用本发明提供的试剂盒，例如可以用于免疫印迹、免疫沉淀等涉及到利用IL-11抗原和抗体特异性结合性能，来检测的试剂盒等。这些试剂盒可包含下列中的任意一种或多种：拮抗剂、抗IL-11抗体或者药物参照材料；蛋白纯化柱；免疫球蛋白亲和纯化缓冲剂；细胞的测定稀释剂；说明书或者文献等。抗IL-11抗体可被用于不同类型的诊断测试，例如可以在体外或者体内检测各种各样的疾病或者药物、毒素或者其他蛋白等的存在。例如可以通过对受试者的血清或者血液进行检测，用来测试相关疾病。这种相关疾病可包括IL-11相关疾病，例如肺纤维化等等。当然本文提供的抗体也可以用于上述疾病的放射免疫检测和放射免疫治疗等等。Of course, the anti-IL-11 antibodies herein can also be made into a kit or part of other diagnostic reagents as needed. According to an embodiment of the present invention, the present invention also provides a kit, which includes the above-mentioned IL-11 antibody. The kit provided by the present invention can be used, for example, for immunoblotting, immunoprecipitation, etc., which involve the use of IL-11 antigen and antibody specific binding properties to detect kits, etc. These kits may contain any one or more of the following: antagonists, anti-IL-11 antibodies or drug reference materials; protein purification columns; immunoglobulin affinity purification buffers; cell assay diluents; instructions or literature, etc. Anti-IL-11 antibodies can be used for different types of diagnostic tests, for example, various diseases or the presence of drugs, toxins or other proteins can be detected in vitro or in vivo. For example, the serum or blood of the subject can be tested to test related diseases. Such related diseases may include IL-11 related diseases, such as pulmonary fibrosis, etc. Of course, the antibodies provided herein can also be used for radioimmunoassay and radioimmunotherapy of the above-mentioned diseases, etc.

在利用本发明所提供的抗IL-11抗体治疗上述疾病时，可以将本发明提供的抗IL-11抗体提供给受试者即可。为此，本发明提供了一种用于治疗上述疾病的方法，包括向有需要的受试者施用本发明所提供的抗体或其抗原结合片段。When the anti-IL-11 antibody provided by the present invention is used to treat the above diseases, the anti-IL-11 antibody provided by the present invention can be provided to the subject. To this end, the present invention provides a method for treating the above diseases, comprising administering the antibody or antigen-binding fragment thereof provided by the present invention to a subject in need.

下面将结合实施例对本发明的方案进行解释。本领域技术人员将会理解，下面的实施例仅用于说明本发明，而不应视为限定本发明的范围。实施例中未注明具体技术或条件的，按照本领域内的文献所描述的技术或条件或者按照产品说明书进行。所用试剂或仪器未注明生产厂商者，均为可以通过市购获得的常规产品。The scheme of the present invention will be explained below in conjunction with the embodiments. It will be appreciated by those skilled in the art that the following embodiments are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. Where specific techniques or conditions are not indicated in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. The reagents or instruments used are not indicated by the manufacturer and are all conventional products that can be obtained commercially.

实施例1：IL-11抗体的改造Example 1: Modification of IL-11 Antibody

载体的构建：人源IL-11氨基酸序列通过全球公共数据库UniProt(P20809：124294)获得。以人的IgG4形式，使用基因工程方法，构建产生了克隆IgG4(SEQ ID NO:85；SEQ ID NO:91)全长人源化抗体。采用电转染的方法，将编码人源IL-11的载体转染至哺乳动物细胞CHO进行表达14天。表达完成后采用Protein A层析柱(NMab Pro，纳微)对收集的细胞培养液进行纯化，平衡液为20mM PBS，0.15M NaCl，pH 7.3，洗脱液为0.05M乙酸，pH 3.0±0.1，收集目标吸收峰下蛋白洗脱液，调整pH和浓度后，取部分样品进行亲和力检测。Vector construction: The amino acid sequence of human IL-11 was obtained through the global public database UniProt (P20809: 124294). In the form of human IgG4, a full-length humanized antibody cloned IgG4 (SEQ ID NO: 85; SEQ ID NO: 91) was constructed using genetic engineering methods. The vector encoding human IL-11 was transfected into mammalian CHO cells by electrofection for expression for 14 days. After the expression was completed, the collected cell culture fluid was purified using a Protein A chromatography column (NMab Pro, Nanomicro). The equilibrium solution was 20mM PBS, 0.15M NaCl, pH 7.3, and the eluent was 0.05M acetic acid, pH 3.0±0.1. The protein eluate under the target absorption peak was collected, and after adjusting the pH and concentration, some samples were taken for affinity detection.

表1：IgG4人源化抗体序列编号

Table 1: IgG4 humanized antibody sequence numbering

实施例2：人源化抗体亲和力检测Example 2: Humanized Antibody Affinity Testing

用生物膜干涉技术测定IgG4人源化抗体与IL-11的结合，得到亲和力常数KD(M)。测定方法包括如下步骤：The binding of IgG4 humanized antibody to IL-11 was measured by biomembrane interference technology to obtain the affinity constant KD (M). The determination method includes the following steps:

将人源化抗体用SD buffer(含有0.05％吐温-20的0.02M PBS缓冲液，pH＝7.4)稀释至25ug/mL，用SD buffer稀释IL11蛋白至5个浓度梯度溶液(3ug/mL～0.1875ug/mL，2倍梯度)，将人源化抗体溶液和IL11蛋白梯度稀释液分别加入至96孔黑色样品板中。通过分子相互作用仪，AHC抗人免疫球蛋白Fc段传感器捕获待测样品中人源化抗体后，自动检测其与IL11蛋白的结合情况，最后用10mM甘氨酸缓冲液洗脱传感器。The humanized antibody was diluted to 25ug/mL with SD buffer (0.02M PBS buffer containing 0.05% Tween-20, pH=7.4), and the IL11 protein was diluted to 5 concentration gradient solutions (3ug/mL～0.1875ug/mL, 2-fold gradient) with SD buffer. The humanized antibody solution and IL11 protein gradient dilution were added to the 96-well black sample plate. Through the molecular interaction instrument, the AHC anti-human immunoglobulin Fc segment sensor captured the humanized antibody in the sample to be tested, and automatically detected its binding with the IL11 protein, and finally eluted the sensor with 10mM glycine buffer.

结果如表2所示，人源化抗体IgG4及阳性对照3C6与IL11相互作用的KD(M)值均处于低摩尔浓度(9.39E-10～5.35E-09)，表明抗体与IL11蛋白均有很高的亲和力，亲和力相当。阳性对照3C6来源于CN113056481A中的3C6抗体序列。The results are shown in Table 2. The KD (M) values of the interaction between humanized antibody IgG4 and positive control 3C6 and IL11 are both at low molar concentrations (9.39E-10 to 5.35E-09), indicating that the antibody and IL11 protein have high affinity and comparable affinity. The positive control 3C6 is derived from the 3C6 antibody sequence in CN113056481A.

表2：人源化抗体与IL-11抗原的亲和力
Table 2: Affinity of humanized antibodies to IL-11 antigen

其中，3C6重链氨基酸序列如SEQ ID NO:95所示；3C6轻链氨基酸序列如SEQ ID NO:96所示。

Among them, the 3C6 heavy chain amino acid sequence is shown in SEQ ID NO:95; the 3C6 light chain amino acid sequence is shown in SEQ ID NO:96.

IgG1的全长重链源于CN2021109237370的SEQ ID NO:151；IgG1的全长轻链源于专利CN2021109237370中的SEQ ID NO:157。The full-length heavy chain of IgG1 is derived from SEQ ID NO: 151 in CN2021109237370; the full-length light chain of IgG1 is derived from SEQ ID NO: 157 in patent CN2021109237370.

实施例3：人源化抗体竞争性阻断IL-11与IL-11R结合Example 3: Humanized antibody competitively blocks the binding of IL-11 to IL-11R

为了评估优选人源化抗体能否阻断IL-11与IL-11R的结合。本实施例构建了稳定表达IL-11R的细胞株membrane-IL-11R CHO。测定方法包括如下步骤：In order to evaluate whether the preferred humanized antibody can block the binding of IL-11 to IL-11R. In this example, a cell line membrane-IL-11R CHO stably expressing IL-11R was constructed. The assay method includes the following steps:

取浓度为20μg/mL的IL-11蛋白(His标签)10μL/孔加入到96孔U型板，与90μL梯度稀释的优选人源化抗体(2400μg/mL、800μg/mL、266.67μg/mL、88.89μg/mL、29.63μg/mL、9.88μg/mL、3.29μg/mL、1.10μg/mL、0.37μg/mL、0.12μg/mL)室温孵育1h。然后与1.5*10^5个membrane-IL-11R CHO细胞(100μL/孔)室温孵育1h后，1500rpm离心5min，用PBST清洗2次，加入1:100稀释的流式二抗PE anti-6x His tag antibody(Abcam，ab72467)，200μL/孔，孵育1h，1500rpm离心5min，用PBST清洗3次，重悬细胞于180μL分析缓冲液中(含有1％FBS的PBS缓冲液)，流式上机检测。Take 10 μL/well of IL-11 protein (His tag) with a concentration of 20 μg/mL and add it to a 96-well U-shaped plate, and incubate it with 90 μL of gradient diluted preferred humanized antibody (2400 μg/mL, 800 μg/mL, 266.67 μg/mL, 88.89 μg/mL, 29.63 μg/mL, 9.88 μg/mL, 3.29 μg/mL, 1.10 μg/mL, 0.37 μg/mL, 0.12 μg/mL) at room temperature for 1 hour. Then incubate it with 1.5*10^5 membrane-IL-11R CHO cells (100 μL/well) at room temperature for 1 hour, centrifuge at 1500 rpm for 5 minutes, wash twice with PBST, add 1:100 diluted flow secondary antibody PE anti-6x His tag antibody (Abcam, ab72467), 200 μL/well, The cells were incubated for 1 hour, centrifuged at 1500 rpm for 5 minutes, washed three times with PBST, and resuspended in 180 μL of analysis buffer (PBS buffer containing 1% FBS) for detection on a flow cytometer.

结果绘制S曲线见图1，IC50值见表3，结果表明人源化抗体IgG4及阳性对照3C6均可有效阻断IL-11与IL-11R的结合，且IgG4阻断效果更强。The results of the S-curve plot are shown in Figure 1, and the IC50 values are shown in Table 3. The results show that both the humanized antibody IgG4 and the positive control 3C6 can effectively block the binding of IL-11 to IL-11R, and the blocking effect of IgG4 is stronger.

表3：人源化抗体竞争性阻断IL-11与IL-11R结合的IC50
Table 3: IC50 of humanized antibodies competitively blocking the binding of IL-11 to IL-11R

实施例4：优选抗体在SD大鼠中抑制肺纤维化Example 4: Preferred antibodies inhibit pulmonary fibrosis in SD rats

在博来霉素(Bleomycin)诱导SD大鼠单侧肺纤维化的动物模型中，当给予人源化抗体治疗后，可以明显观察到人源化抗体减缓大鼠肺的炎症反应及纤维化进程。在实施例中，采用2.5％异氟烷将大鼠进行麻醉，然后手术打开主气管后，注射适量的溶媒或博来霉素进行造模。手术完成后观察大鼠恢复情况。手术后的第13天进行灌胃或腹腔给药进行治疗25天，第26天解剖。具体动物分组及给药剂量汇总如下表4。In the animal model of unilateral pulmonary fibrosis induced by Bleomycin in SD rats, after treatment with humanized antibodies, it can be clearly observed that humanized antibodies slow down the inflammatory response and fibrosis process of the rat lungs. In the embodiment, the rats were anesthetized with 2.5% isoflurane, and then the main trachea was opened surgically, and an appropriate amount of solvent or bleomycin was injected for modeling. After the operation, the recovery of the rats was observed. On the 13th day after the operation, oral gavage or intraperitoneal administration was performed for 25 days of treatment, and dissection was performed on the 26th day. The specific animal grouping and dosage are summarized in Table 4 below.

治疗结束后，取大鼠的肺进行石蜡包埋后切片，并进行HE染色及Masson三色染色。其中，HE染色主要指示肺组织中炎症的浸润水平。Masson三染色主要用于评价肺组织纤维化的水平。After the treatment, the lungs of the rats were taken and embedded in paraffin, sliced, and stained with HE and Masson's trichrome. HE staining mainly indicates the level of inflammatory infiltration in the lung tissue. Masson's trichrome staining is mainly used to evaluate the level of pulmonary tissue fibrosis.

结果如图2与图3所示，表明优选人源化抗体能很好的降低博来霉素所引起的大鼠肺炎 症损伤及纤维化。The results are shown in Figures 2 and 3, indicating that the preferred humanized antibody can effectively reduce bleomycin-induced pneumonia in rats Damage and fibrosis.

表4：人源化抗体(IgG4)大鼠药效实验方案

注：Yes代表进行了造模处理；NA代表未进行给药；IP代表腹膜内给药；QOD代表隔日一
次。Table 4: Humanized antibody (IgG4) rat efficacy test plan

Note: Yes means modeling was performed; NA means no medication was given; IP means intraperitoneal administration; QOD means once every other day.

实施例5：人源化Anti-IL-11抗体对SD大鼠血小板功能影响检测Example 5: Detection of the effect of humanized Anti-IL-11 antibody on platelet function in SD rats

本实施例使用雄性SD大鼠12只，随机分为3组，4只/组。单次静脉滴注溶媒对照、人源化抗IL11抗体IgG1(简称IgG1)和人源化抗IL11抗体IgG4(简称IgG4)，药物组剂量为100mg/kg，给药容量为10mL/kg。每天观察死亡和濒死情况1次，临床症状2次，体重给药前称重一次，在给药后5min、30min、5h、24h、48h和120h进行血液学指标检测。In this example, 12 male SD rats were used and randomly divided into 3 groups, 4 rats/group. A single intravenous drip of vehicle control, humanized anti-IL11 antibody IgG1 (abbreviated as IgG1) and humanized anti-IL11 antibody IgG4 (abbreviated as IgG4) was administered, the drug group dose was 100 mg/kg, and the administration volume was 10 mL/kg. Death and dying were observed once a day, clinical symptoms were observed twice, body weight was weighed once before administration, and hematological indicators were detected at 5 min, 30 min, 5 h, 24 h, 48 h and 120 h after administration.

实验结果：Experimental results:

死亡或濒死情况：试验期间，无动物死亡；Death or dying conditions: No animals died during the experiment;

临床症状：试验期间，动物临床症状未见异常；Clinical symptoms: During the test period, no abnormal clinical symptoms were observed in the animals;

血液学：IgG1在100mg/kg剂量下会引起大鼠PLT(血小板)下降，24h降至最低点0.8％，48h可见恢复趋势，IgG4不会引起PLT明显变化。Hematology: IgG1 at a dose of 100 mg/kg caused a decrease in PLT (platelets) in rats, which dropped to a minimum of 0.8% after 24 hours, and a recovery trend was seen after 48 hours. IgG4 did not cause significant changes in PLT.

在本说明书的描述中，参考术语“一个实施例”、“一些实施例”、“示例”、“具体示例”、或“一些示例”等的描述意指结合该实施例或示例描述的具体特征、结构、材料或者特点包含于本发明的至少一个实施例或示例中。在本说明书中，对上述术语的示意性表述不必须针对的是相同的实施例或示例。而且，描述的具体特征、结构、材料或者特点可以在任一个或多个实施例或示例中以合适的方式结合。此外，在不相互矛盾的情况下，本领域的技术人员可以将本说明书中描述的不同实施例或示例以及不同实施例或示例的特征进行结合和组合。In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine the different embodiments or examples described in this specification and the features of the different embodiments or examples, without contradiction.

尽管上面已经示出和描述了本发明的实施例，可以理解的是，上述实施例是示例性的，不能理解为对本发明的限制，本领域的普通技术人员在本发明的范围内可以对上述实施例进行变化、修改、替换和变型。 Although the embodiments of the present invention have been shown and described above, it is to be understood that the above embodiments are exemplary and are not to be construed as limitations of the present invention. A person skilled in the art may change, modify, replace and vary the above embodiments within the scope of the present invention.

Claims (16)

一种能够特异性识别IL-11的抗体或其抗原结合片段，其特征在于，所述抗体含有选自下列至少之一的CDR序列或与其具有至少95％同一性的氨基酸序列；以及An antibody or an antigen-binding fragment thereof capable of specifically recognizing IL-11, characterized in that the antibody comprises a CDR sequence selected from at least one of the following or an amino acid sequence having at least 95% identity therewith; and 重链恒定区和轻链恒定区，所述抗体的轻链恒定区来自于人源的Kappa轻链恒定区，重链恒定区来自于人源IgG4的重链恒定区；A heavy chain constant region and a light chain constant region, wherein the light chain constant region of the antibody is derived from a human Kappa light chain constant region, and the heavy chain constant region is derived from a human IgG4 heavy chain constant region; 所述抗体或其抗原结合片段具有人源化修饰；The antibody or antigen-binding fragment thereof has humanized modification; 所述抗体包括：The antibodies include: 分别如SEQ ID NO:1、2和3或者与SEQ ID NO:1、2和3具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 1, 2 and 3, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 1, 2 and 3; or 分别如SEQ ID NO:4、5和6或者与SEQ ID NO:4、5和6具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 4, 5 and 6, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 4, 5 and 6; or 分别如SEQ ID NO:7、8和9或者与SEQ ID NO:7、8和9具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 7, 8 and 9, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 7, 8 and 9; or 分别如SEQ ID NO:10、11和12或者与SEQ ID NO:10、11和12具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NOs: 10, 11 and 12, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 10, 11 and 12; or 分别如SEQ ID NO:13、14和15或者与SEQ ID NO:13、14和15具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 13, 14 and 15, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 13, 14 and 15; or 分别如SEQ ID NO:16、17和18或者与SEQ ID NO:16、17和18具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 16, 17 and 18, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 16, 17 and 18; or 分别如SEQ ID NO19、20和21或者与SEQ ID NO:19、20和21具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 19, 20 and 21, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 19, 20 and 21; or 分别如SEQ ID NO:22、23和24或者与SEQ ID NO:22、23和24具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 22, 23 and 24, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 22, 23 and 24; or 分别如SEQ ID NO:25、26和27或者与SEQ ID NO:25、26和27具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 25, 26 and 27, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 25, 26 and 27; or 分别如SEQ ID NO:28、29和30或者与SEQ ID NO:28、29和30具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；或者The heavy chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 28, 29 and 30, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 28, 29 and 30; or 分别如SEQ ID NO:31、32和33或者与SEQ ID NO:31、32和33具有至少95％同一性的氨基酸序列所示的重链可变区CDR1、CDR2、CDR3序列；The heavy chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO:31, 32 and 33, respectively, or amino acid sequences having at least 95% identity with SEQ ID NO:31, 32 and 33; 和and 分别如SEQ ID NO:34、35和36或者与SEQ ID NO:34、35和36具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者 The light chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NOs: 34, 35 and 36, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NOs: 34, 35 and 36; or 分别如SEQ ID NO:37、38和39或者与SEQ ID NO:37、38和39具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 37, 38 and 39, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 37, 38 and 39; or 分别如SEQ ID NO:40、41和42或者与SEQ ID NO:40、41和42具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 40, 41 and 42, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 40, 41 and 42; or 分别如SEQ ID NO:43、44和45或者与SEQ ID NO:43、44和45具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 43, 44 and 45, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 43, 44 and 45; or 分别如SEQ ID NO:46、47和48或者与SEQ ID NO:46、47和48具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 46, 47 and 48, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 46, 47 and 48; or 分别如SEQ ID NO:49、50和51或者与SEQ ID NO:49、50和51具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences as shown in SEQ ID NO: 49, 50 and 51, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 49, 50 and 51; or 分别如SEQ ID NO:52、53和54或者与SEQ ID NO:52、53和54具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 52, 53 and 54, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 52, 53 and 54; or 分别如SEQ ID NO:55、56和57或者与SEQ ID NO:55、56和57具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 55, 56 and 57, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 55, 56 and 57; or 分别如SEQ ID NO:58、59和60或者与SEQ ID NO:58、59和60具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are as shown in SEQ ID NO: 58, 59 and 60, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 58, 59 and 60; or 分别如SEQ ID NO:61、62和63或者与SEQ ID NO:61、62和63具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列；或者The light chain variable region CDR1, CDR2, CDR3 sequences are shown in SEQ ID NO: 61, 62 and 63, respectively, or amino acid sequences that are at least 95% identical to SEQ ID NO: 61, 62 and 63; or 分别如SEQ ID NO:64、65和66或者与SEQ ID NO:64、65和66具有至少95％同一性的氨基酸序列所示的轻链可变区CDR1、CDR2、CDR3序列。The light chain variable region CDR1, CDR2, and CDR3 sequences are respectively shown in SEQ ID NO: 64, 65, and 66, or amino acid sequences that are at least 95% identical to SEQ ID NO: 64, 65, and 66. 根据权利要求1所述的抗体或其抗原结合片段，其特征在于，所述抗体含有重链框架区序列和轻链框架区序列的至少之一，所述重链框架区序列和轻链框架区序列的至少之一的至少一部分来自于鼠源抗体、人源抗体、灵长目源抗体或其突变体的至少之一。The antibody or antigen-binding fragment thereof according to claim 1, characterized in that the antibody contains at least one of a heavy chain framework region sequence and a light chain framework region sequence, and at least a portion of at least one of the heavy chain framework region sequence and the light chain framework region sequence is derived from at least one of a murine antibody, a human antibody, a primate antibody or a mutant thereof. 根据权利要求2所述的抗体或其抗原结合片段，其特征在于，所述重链框架区包括FL1、FL2、FL3和FL4，所述重链框架区FL1、FL2、FL3和FL4分别具有SEQ ID NO:67、68、69和70所示的氨基酸序列，
QVQLVQSGAEVKKPGASVKVSCKAS(SEQ ID NO:67)，
LGWVRQAPGX1GLEWX2GH(SEQ ID NO:68)，
NYNEKFKGRX3TX4TX5DTSTSTVYMELSSLRSEDTAVYX6C(SEQ ID NO:69)，
WGQGTLVTVSS(SEQ ID NO:70)，The antibody or antigen-binding fragment thereof according to claim 2, characterized in that the heavy chain framework region comprises FL1, FL2, FL3 and FL4, and the heavy chain framework regions FL1, FL2, FL3 and FL4 have the amino acid sequences shown in SEQ ID NOs: 67, 68, 69 and 70, respectively,
QVQLVQSGAEVKKPGASVKVSCKAS (SEQ ID NO: 67),
LGWVRQAPGX1GLEWX2GH (SEQ ID NO: 68),
NYNEKFKGRX3TX4TX5DTSTSTVYMELSSLRSEDTAVYX6C (SEQ ID NO: 69),
WGQGTLVTVSS (SEQ ID NO: 70), 其中，X1为Q或H，X2为M或I，X3为V或A，X4为M或L，X5为R或A，X6为 Y或F。Among them, X1 is Q or H, X2 is M or I, X3 is V or A, X4 is M or L, X5 is R or A, X6 is Y or F. 根据权利要求2所述的抗体或其抗原结合片段，其特征在于，所述轻链框架区包括FL1、FL2、FL3和FL4，所述轻链框架区FL1、FL2、FL3和FL4分别具有SEQ ID NO:71、72、73和74所示的氨基酸序列，
EIVLTQSPATLSLSPGERATLSCRAS(SEQ ID NO:71)，
MNWX7QQKPGQAPRLLIY(SEQ ID NO:72)，
NQGSGIPARFSGSGSGTDFTLTISSLEPEDFAX8YX9C(SEQ ID NO:73)，
FGGGTKLEIK(SEQ ID NO:74)，The antibody or antigen-binding fragment thereof according to claim 2, characterized in that the light chain framework region comprises FL1, FL2, FL3 and FL4, and the light chain framework regions FL1, FL2, FL3 and FL4 have the amino acid sequences shown in SEQ ID NOs: 71, 72, 73 and 74, respectively,
EIVLTQSPATLSLSPGERATLSCRAS (SEQ ID NO: 71),
MNWX7QQKPGQAPRLLIY (SEQ ID NO: 72),
NQGSGIPARFSGSGSGTDFTLTISSLEPEDFAX8YX9C (SEQ ID NO: 73),
FGGGTKLEIK (SEQ ID NO: 74), 其中，X7为Y或F，X8为V或M，X9为Y或F。Among them, X7 is Y or F, X8 is V or M, and X9 is Y or F. 根据权利要求1所述的抗体或其抗原结合片段，其特征在于，所述抗体具有如SEQ ID NO:75～78任一项所示氨基酸序列的重链可变区。The antibody or antigen-binding fragment thereof according to claim 1, characterized in that the antibody has a heavy chain variable region with an amino acid sequence as shown in any one of SEQ ID NO: 75 to 78. 根据权利要求1所述的抗体或其抗原结合片段，其特征在于，所述抗体具有如SEQ ID NO:79～82任一项所示氨基酸序列的轻链可变区。The antibody or antigen-binding fragment thereof according to claim 1, characterized in that the antibody has a light chain variable region with an amino acid sequence as shown in any one of SEQ ID NO: 79 to 82. 根据权利要求1所述的抗体或其抗原结合片段，其特征在于，所述抗体恒定区的全长序列如SEQ ID NO:83或84所示。The antibody or antigen-binding fragment thereof according to claim 1, characterized in that the full-length sequence of the antibody constant region is shown in SEQ ID NO: 83 or 84. 根据权利要求1所述的抗体或其抗原结合片段，其特征在于，所述抗体具有SEQ ID NO:85～88任一项所示氨基酸序列的重链和具有SEQ ID NO:89～92任一项所示氨基酸序列的轻链。The antibody or antigen-binding fragment thereof according to claim 1, characterized in that the antibody has a heavy chain having an amino acid sequence shown in any one of SEQ ID NOs: 85 to 88 and a light chain having an amino acid sequence shown in any one of SEQ ID NOs: 89 to 92. 一种核酸分子，其特征在于，所述核酸分子编码权利要求1～8任一项所述的抗体或其抗原结合片段。A nucleic acid molecule, characterized in that the nucleic acid molecule encodes the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8. 根据权利要求9所述的核酸分子，其特征在于，所述核酸分子为DNA；所述核酸分子具有如SEQ ID NO:93或94所示核苷酸序列。The nucleic acid molecule according to claim 9 is characterized in that the nucleic acid molecule is DNA; the nucleic acid molecule has a nucleotide sequence shown in SEQ ID NO:93 or 94. 一种表达载体，其特征在于，携带权利要求9～10任一项所述的核酸分子。An expression vector, characterized in that it carries the nucleic acid molecule according to any one of claims 9 to 10. 一种重组细胞，其特征在于，所述重组细胞表达权利要求1～8任一项所述的抗体或其抗原结合片段，或者携带权利要求9～10任一项所述的核酸分子；A recombinant cell, characterized in that the recombinant cell expresses the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, or carries the nucleic acid molecule according to any one of claims 9 to 10; 优选地，所述重组细胞是通过将权利要求11所述的表达载体引入至宿主细胞中而获得的。Preferably, the recombinant cell is obtained by introducing the expression vector according to claim 11 into a host cell. 一种药物组合物，其特征在于，含有权利要求1～8任一项所述的抗体或其抗原结合片段，权利要求9～10任一项所述的核酸分子，权利要求11所述的表达载体或权利要求12所述的重组细胞。A pharmaceutical composition, characterized in that it contains the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, the nucleic acid molecule according to any one of claims 9 to 10, the expression vector according to claim 11 or the recombinant cell according to claim 12. 权利要求1～8任一项所述的抗体或其抗原结合片段、权利要求9～10任一项所述的 核酸分子、权利要求11所述的表达载体或权利要求12所述的重组细胞、权利要求13所述的药物组合物在制备药物中的用途，所述药物用于治疗或者预防肺纤维化，非酒精性脂肪肝炎，慢性心力衰竭或由成纤维细胞向纤维化转化所导致的疾病。The antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, or the antibody or antigen-binding fragment thereof according to any one of claims 9 to 10 Use of the nucleic acid molecule, the expression vector according to claim 11 or the recombinant cell according to claim 12, and the pharmaceutical composition according to claim 13 in the preparation of a drug for treating or preventing pulmonary fibrosis, non-alcoholic fatty hepatitis, chronic heart failure, or a disease caused by the transformation of fibroblasts to fibrosis. 一种检测IL-11的试剂盒，其特征在于，包括权利要求1～8任一项所述的抗体。A kit for detecting IL-11, characterized in that it comprises the antibody according to any one of claims 1 to 8. 权利要求1～8任一项所述的抗体、权利要求9～10任一项所述的核酸分子、权利要求11所述的表达载体或权利要求12所述的重组细胞在制备试剂盒中的用途，所述试剂盒用于检测IL-11或者诊断IL-11相关的疾病。 Use of the antibody according to any one of claims 1 to 8, the nucleic acid molecule according to any one of claims 9 to 10, the expression vector according to claim 11 or the recombinant cell according to claim 12 in the preparation of a kit for detecting IL-11 or diagnosing IL-11-related diseases.
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YU JIFENG, SONG YONGPING, TIAN WENZHI: "How to select IgG subclasses in developing anti-tumor therapeutic antibodies", JOURNAL OF HEMATOLOGY & ONCOLOGY, vol. 13, no. 1, 1 December 2020 (2020-12-01), XP055884120, DOI: 10.1186/s13045-020-00876-4 *

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